EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

Epidemiudvalget 2021-22
EPI Alm.del Bilag 322
Offentligt

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

EUROPEAN MEDICINES AGENCY

SCJENCE

bilag 9

MEDICINES

HEAETH

Amsterdam, 30 November 2020

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

Committee for Medicinal Products for Human Use (CHMP)

Quality rolling review CHMP overview and list of questions

COVID-19 mRNA Vaccine BioNTech

BNT162b2, S’capped mRNA encoding full length SARS-C0V-2 Spike protein

Procedure No. EMEA/H/C/005735/RR/02

Applicant: BioNTech Manufacturing GmbH

Official address Domenico Scarlattilaen 6

•

1083 HS Amsterdam

•

The Netherlands

Address for

visits

and deliveries Refer to www.ema.europâEi.’ho,-to-hnd-us

Send usa question Go to ww; ema.europa.eu/contact Telephone +31 (0)88 781 6000

An agency of

Lhe

European Unjon

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

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Table ofContents

Administrative information

1. Executive summary

1.1. Scope of the rolling review submission

New active substance status

1.2. The development programme/compliance with CHMP guidance/scientific advice

1.3. General comments on compliance with GMP, GLP, GCP

1.4. Type of application and other comments on the submitted dossier

2. Scientific overview and discussion on new data

2.1. Quality aspects

2.1.1. Introduction

2.1.2. Active Substance

General Information

Manufacture, process controls and characterisation

Specification, analytical procedures, reference standards, batch analysis, and

container ciosure

Stability

2.1.3. Finished Medicinal Product

Description of the product and Pharmaceutical Development

Manufacture of the product and process controls

Product specification, analytical procedures, batch analysis

Stability of the product

Adventitious agents

GMO

N/A

Novel excipients

2.1.4. Discussion and conclusions on chemical, pharmaceutical and biological aspects

2.2. Non-cllnical aspects

2.2.1. Pharmacology

2.2.2. Pharmacokinetics

2.2.3. Toxicology

2.2.4. Ecotoxicity/environmental risk assessment

2.2.5. Discussion on non-clinical aspects

2.2.6. Conclusion on non-clinical aspects

2.3. Clinical aspects

2.4. Risk management plan

2.5. Pharmacovigilance system

3. SCIENTIFIC OVERVIEW

AND DISCUSSION on responses to questions

raised in previous cycle(s)

Benefit risk assessment

CHMP list of questions

5.1. Quality aspects

COVID-19 mRNA vaccine fnucleoside modified)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

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Major objections

Other concerns

5.2. Non-clinical aspects

Major objections

Other concerns

5.3. Cilnical aspects

5.4. Risk management plan

5.5. Pharmacovigilance system

5.6. New active substance status

.54

6. Recommended conditions for future marketing authorisation and product

information in case of a positive benefit risk assessment

7. Appendices (as appropriate)

COVID-19 mRNA vaccine (nucleoside modified)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

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Administrative informatïon

Invented name of the medicinal product:

INN

for common name) of the active

substance(s):

Applicant:

Applied Indication(s):

Pharmaco-therapeutic group

(ATC Code):

Pharmaceuticai form(s) and strength(s):

Rapporteur contact person:

COVID-19 mRNA Vaccine BioNTech

BNT162b2, 5’capped mRNA encoding full Iength

SRAS-CoV-2 Spike protein

BioNTech Manufacturing GmbH

TBD

]078X

Concentrate for suspension for injection

0,225 mg

Elin Blom

+46(0) 18 17 15 60

elin. bIomlakemedelsverket.se

Perrine Nuez

Perrine.NUEZ@’ansm.sante.fr

Vanessa Seguin

[email protected]

Co-Rapporteur contact person:

EMA Product Lead:

Names of the Rapporteur assessors

(internal and external):

Tel to ali: +46 18 17 46 00

Quality

Andreea Barbu

[email protected]

Anna Hiligren

anna. hilIgrenIakemedelsverket.se

Stefan Nilsson

stefan. [email protected]

Margareta Ramström Jonsson

[email protected]

Anna Karin Rehnström

anna-karin.rehnstromlakemedelsverket.se

Non-cl inical

Tom Meyerson Goldschmidt

tom. [email protected]

Dariush Mokhtari

dariush. mokhtarftlakemedelsverket.se

Birger Scholz

birqer.scholzclakemedelsverket.se

Clinical:

Efficacy & Safety

Helena Back

[email protected]

Charlotta Bergquist

charlotta [email protected]

Helena Faust

[email protected]

Jessica Mwinyi

[email protected]

Risk Management Plan

Helena Back

[email protected]

Jessica Mwinyi

COVID-19 mRNA vaccine (nucleoside modified)

Quahty rolling review CHMP overview and list of questions

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

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jessica. mwinvklakemedelsverket. se

Statistics

Linda Dalin

[email protected]

Maria Grünewald

[email protected]

Pharmacovigilance system

Elin Blom

elin. bIomIakemedelsverket.se

Product Information

Markus Forslund

markus.forslund©IakemedelsverkeLse

Kim Sherwood

[email protected]

CHMP members

Jean-Michel. RACE@ansm .sante.fr

alexandre.moreauansm.sante.fr

Coordinator

[email protected]

Assistant Director

Alban. DHANANIfansm.sante.fr

Head Vaccine division

[email protected]

Head European division

Vincent.gazinansm.sante.fr

Clinical efficacy/safety

[email protected]

Quality

[email protected]

Estelle. [email protected]

[email protected]

Francois.CANOansm.sante.fr

Christian. pitotansm.sante.fr

Dominipue.GARCIAtansm.sante.fr

Solene. MAITENAZansm .sante.fr

Viral safety

[email protected]

Frederik. [email protected]

[email protected]

Non-clinical

[email protected]

Matthew.BURBANKansm.sante.fr

Pirrvi,

cnresansmsantefr

Date 2020-11-19

Signature Filip Josephson

Names of the Co-Rapporteur assessors

(internal and external):

In accordance with Articie 6(3) of

Regulation (RC) No 726/2004, I the

Rapporteur hereby declare that I have

completed my assessment report in less

than 80 days.

COVID-19 mRNA

vaccine (nucleoside modified)

Quality rolling review CHMP overview end list of questions

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/64;856/2020

Page 5/81

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Decla ratio n s

This application includes an Active Substance Master File (ASMF):

Ves

EN0

The assessor confirms that proprietary information an, or reference to, third pafties (e.g. ASMF

holder) or products are not inciuded in this assessment, including the Product Information, unless

there are previous contracts and/ar agreements with the third party(ies).

The assessor conflrms that reference to ongoing assessments ar development plans for other

products is flot fncluded in this assessment report.

Whenever the above box is un-ticked please indicate section and page where confidential information

is located (including the Product Information document) here:

COV1D19 mRNA vaccine (nudeoside modified)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

Page 6/81

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List of abbreviations

5’ cap

AF4-MALS-QELS

5’ capping structure, (m27,3’-OGppp(m12’-O)ApG)

Asymmetric Flow Field-Flow Fractionation (AF4) Multi-Angle Static and Quasi

Elastic Light Scattering

PEG-lipid, 2-[(polyethylene glycol)-2000J-N,N-ditetradecylacetamide

Cationic lipid, ((4-hydroxybutyl)azanediyl)bis(hexane-6, 1-diyl)bis(2-

hexyldecanoate)

Acceptance Quality Limit

adenosine triphosphate

Area under the curve

Vaccine candidate encoding the SARS-CoV-2 full-length spike protein, modified

by 2 proline mutations (P2 5)

bovine spongiform encephalopathies

Cause and Effect Matrices

Cause and Effects

Charged Aerosol Detection

Container Closure Integrity

N,N-carbonyldiimidazole

Capillary Gel Electrophoresis

Conditional marketing authorisation

Certificate of Analysis

Coronavirus disease 2019

Critical process parameter

Critical Quality Attribute

CIlnical Reference Material

Clinical Trial Material

Droplet digital PCR

Detection Limit

Dynamic Light Scattering

Design of experiments

Drug Product

Drug Substance

Differential Scanning Calorimetry

ALC-0159

ALC-0315

AQL

ATP

AUC

BNT162b2

BSE

C&E

CAD

CCI

CDI

CGE

CMA

CoA

COVID-19

CPP

CQA

CRM

CTM

ddPCR

DLS

DOE

DSC

COVID-19 mRNA vaccine (riucleoside modified)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/ 2O2OEMA/CHMP/641856/2020

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DSPC

DVS

EDTA

ELSD

EVA

FID

Phospholipid, (1,2-distearoyl-sn-glycero-3-phosphocholine)

Dynamic Vapor sorption

Ethylenediaminetetraacetic acid

Evaporative Light Scattering Detection

Ethylene Vinyl Acetate

Ethylene vinyl acetate

Flame lonization Detector

Failure

Modes

and Effects Analysis

Fourier-Transform lnfrared

Gas Chromatography

Good Manufacturing Practice

Guanidine triphosphate

High Efficiency Particulate Arresting filter

N-(2-hydroxyethyl)-piperazine-N-(2-ethanesulfonic acid)

High Performance Liquid Chromatography

Heat Transfer Fluid

International Council for Harmonisation

Inductively Coupled Plasma

ion-paired reversed-phase high performance liquid chromatography

In-Process Tests for Control

In-Process Tests for Monitoring

Infrared spectroscopy

In-Vitro Expression

In vitro transcription

]apanese Pharmacopeia

Liquid Chromatography

Lipid nanoparticle

Marketing authorization application

Multi-factor-at-a-time

Master cell bank

Messenger RNA

Mass Spectroscopy

FMEA

FrIR

GMP

GTP

HEPA filter

HEPES

HPLC

HTF

ICH

ICP

IP-RP-HPLC

IPT-C

IPT-M

IVE

IVT

LNP

MAA

MFAT

MBC

mRNA

COVID-19 mRNA vaccine fnucleoside modified)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

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N/P

Molar ratio of the amme in the cationic lipid (N) to the phosphate in anionic

phosphodiester backbone of RNA (P)

Next Generation Sequencing

Nuclear Magnetic Resonance

Normal Operating Range

Nucleotide triphosphate

One-factor-at-a-time

Operational Qualification

Proven acceptable ranges

Phosphate-Buffered Salme

Polymerase Chain Reaction

Polyethylene glycol

Polyethersulfone filter

European Pharmacopeia

polyadenosmne

Process Performance Qualification

Performance Qualification

Primary Reference Materials

Polytetrafluoroethylene

Process Validation

Quality Attributes

Quality Control

Quantitation Limit

Quality Target Product Profile

Risk Priority Number

ion-pair reversed-phase high performance liquid chromatography- ultraviolet

light detection at 260 nm and online electrospray onization mass spectrometry

Rolling review

Residual Seal Force

Reverse Transcription Polymerase Chain Reaction

Overall ratio between root mean square radius (Rz) and hydrodynamic radius

NGS

NMR

NOR

NTP

OFAT

PAR

PBS

PCR

PEG

PES filter

Ph. Eur.

poly(A)

PPQ

PRM

PTFE

QTPP

RPN

RP-HPLC/UV-ESI

RSF

RT-PCR

Rz/Rh

COVID-19 mRNA vaccine fnucleoside modifled)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

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(

Rh)

SARS

SARS-CoV-2

TFF

TLC

TOC

TSE

UFDF

USP

UTP

UTR

WCB

WRM

XRD

Spike glycoprotein

Severe acute respiratory syndrome

SARS Coronavirus-2; virus causing the disease COVID-19

Tangential Flow Filtration

Thin Layer Chromatography

Total Organic Carbon

transmitting transmissible spongiform encephalopathies

ultrafiltration/diafiltration

United States Pharmacopeia

uridine triphosphate

Untranslated region

Ultraviolet

Working cell bank

Working Reference Materials

X-Ray Diffraction

COVID-

mRNA vaccine (nucleoside modified)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

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1. Executive summary

Rolling Review is an

ad hoc

procedure used in an emergency context. Rolling Review procedures allow

the CHMP to review quality, non-clinical and clinical data as they become available, before a formal

regulatory application is submitted to the Agency. The main objective of the rolling review is to

expedite the future assessment of the scientific data once submitted in the context of a formal

regulatory application. As such, the scientiHc standards and regulatory principles applied in rolling

reviews are the same as those applicable to formal regulatory procedures. Consequently, concepts

such as “Major Objections” or “Other Concerns” are used in this Rolling Review report to categorise, in

the same manner as in a formal regulatory application, the deficiencies identified by the CHMP on the

preliminary data submitted.

The assessment performed for Rolling Review procedures is without prejudice to additional

considerations that may be held during the subsequent assessment of the formal reg ulatory

application. Only the scientific opinion adopted on the formal regulatory application constitutes the final

view of the CHMP on whether the medicinal product satisfies the criteria for marketing authorisation.

1.1. Scope of the rolling review submission

RR1

—

conciuded on 06.11.2020

Non-clinical dossier has been submitted for the first rolling review cycle i (RR1).

RR2

—

subject of this assessment

For this second rolling review cycle (RR2) quality data has been submitted. This second rolling review

cycle is the first rolling review that contains quality documentation, RR2 (CMC1).

The applicant plans to update several sections in the Quality part of dossier as part of upcoming

submission for quality data package and states the following:

“Data for this section is pending and will be updated once the data has been generated, analyzed, and

verified”.

Only partial information on the quality and non-clinical data has been submitted. Other modules of the

dossier (e.g. clinical) have flot been submitted.

Only the opinion adopted by the CHMP in the context of the application for marketing authorisation

constitutes the final position of the committee on the quality, safety and efficacy of the medicinal

product.

New active substance status

Based on the review of the data the active substance BNT162b2, 5’capped mRNA encoding full length

SRAS-CoV-2 Spike protein contained in the medicinal product COVID-19 mRNA Vaccine BioNTech is

considered to be qualified as a new active substance in itself.

1.2. The development programme/compliance with CHMP

guidance/scientific advice

N/A

COVID-19 mRNA vaccine (nucleoside modified)

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1.3. General comments on compliance with GMP, GLP, GCP

Regarding the non-clinical dossier, the pivotal toxicological studies are stated to be GLP compliant.

There are some issues with repeat-dose toxicity study #38166 regarding the documentation which led

to a request for an GLP inspection of the laboratory site (as adopted by the CHMP).

The EMA Compliance and Inspection Service has reviewed the manufacturer information contained in

the application form and available certificates from the EEA National Competent Authorities. EMA

confirms that a GMP Distant Assessment (DA) of the US Andover and Chesterfield sites are on-going.

1.4. Type of application and other comments on the submitted dossier

Legal basis

The legal basis for this application will be provided as part of the marketing authorisation application

submission refers to:

Article 8.3 of Directive 2001/83/EC, as amended

New active substance status

The applicant requested the active substance BNT162b2, 5’capped mRNA encoding full length

SRAS

CoV-2 Spike protein contained in the above medicinal product to be considered as a new active

substance, as the applicant claims that it is flot a constituent of a medicinal product previously

authorised within the European Union.

complete and independent application.

1.5. Steps taken for the rolling review of the product

The Rapporteur and Co-Rapporteur appointed by the CHMP were:

Rapporteur: Filip ]osephson

CHMP Peer reviewer: Ingrid Wang

Submission of the first package (NC) via eCTD

Validation and start of ist RR round

Rapporteurs’ CHMP ARs and draft overviews to peer reviewer, ETF,

CHMP and EMA for 48 h consultation and comments

Deadline for comments

BWP extraordinary adobe: agreement on BWP report and draft L0Q &

proposals for SOBs/REC5

Updated joint draft overview and LoQ drafted by Rapporteurs and

circulated to CHMP and ETF

ETF discussions on the consolidated List of Questions

CHMP written procedure

Adoption of the 2nd interim opinion for this rolling review of COVID-19

mRNA Vaccine Bio NTech on

06 November 2020

07 November 2020

19 November 2020

Co-Rapporteur: Jean Michel RACE

23 November 2020

24 November 2020

25 November 2020

26 November 2020

27 November 2020

30 November 2020

COVID- 19 mRNA vaccine (nucleoside modified)

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2. Scientific overview and discussion on new data

2.1. Quality aspects

2.1.1. Introduction

The vaccine is based on the SARS CoV-2 spike glycoprotein (S) encoded in RNA and formulated in lipid

nanoparticies (LNPs), referred to as COVID-19 Vaccine (BNT162b2).

The finished product is presented as a preservative-free, multi-dose concentrate to be diluted for

intramuscular injection, intended for 5 doses. The finished product is a sterile dispersion of RNA

containing lipid nanoparticles (LNP5) in aqueous cryoprotectant buffer containing 30 lJg/dose of the

active substance BNT162b2, 5’capped mRNA encoding full length SARS-CoV-2 Spike protein as active

substance.

Other ingredients are: ALC-0315((4-hydroxybutyl)azanediyl)bis(hexane-6, 1-diyl)bis(2-

hexyldecanoate), ALC-0159 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide), DSPC (1,2-

distearoyl-sn-glycero-3-phosphocholine), cholesterol, sucrose, sodium chloride, potassium chioride,

disodium phosphate dihydrate, potassium dihydrogen phosphate and water.

The product is available in glass vial sealed with a bromobutyl rubber stopper and an aluminium seal

with flip-off plastic cap.

2.1.2. Active Substance

General Information

The active substance consists of a single-stranded, 5-capped mRNA that is translated nto a codon

optimized sequence encoding the spike antigen of SARS-CoV-2. Figure i illustrates the general

structure of the antigen-encoding RNA: In addition to the codon-optimized sequence encoding the

antigen, the RNA contains common structural elements optimized for mediating high RNA stability and

translational efficiency (5-cap, 5-UTR, 3’-UTR, poly(A)-tail; see below). Furthermore, an intrinsic

signal peptide (sec) is part of the open reading frame and is translated as an N-terminal peptide. The

RNA does not contain any uridines; instead of uridine the modified Ni-methylpseudouridine is used in

RNA synthesis.

Figure 1. General structure of the RNA

Ca p1

m273’°Gppp(m12°) ApG

RBPO2O.2

-KozakI sec

5152 protein

fF1 etfent

[ø:øi

Schematic illustration of the general structure of the BNT1G2b2 drug substance with 5-cap, 5’- and 3’-untranslated

regions (hAg-Kozak and FI element, respectively), coding sequence with mutations and intrinsic signal peptide (sec)

as well as poly(A)-tail (A30L70). Individual elements are not drawn to scale compared to their respective sequence

Iengths.

COVID-19 mRNA vaccine (nucleoside modified)

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Manufacture, process controls and characterisation

Manufacturers

The Drug Substance is manufactured and controlled by either Wyeth SioPharma Division, Andover,

United States ar by BioNTech Manufacturing GmbH, Mainz, Germany, (steps 1-3) and Rentschler

Biopharma SE, Laupheim, Germany (steps 4 and 5). Of note, the manufacturing process at the

European sites is flot yet included in the application.

Release and stability testing sites are listed. As Mutual Recognition Agreement is not in force for

human vaccines, the provided documentation for manufacturing and testing sites located in the USA is

not considered sufficient

(MO).

Descriptlon of manufacturing process and process controls

Information on the manufacturing process and process controls for the manufacturing site BNT Mainz &

Rentschler is not yet provided. Therefore, the comments below are related only to the Andover site. It

is expected that no significant differences between the two processes are envisaged. However, minor

process adaption could be accepted provided that they will be appropriately validated.

Overall description of the manufacturing process steps

The manufacturing process of BNT162b2 drug substance CDS) involves five major steps. The DS is

produced at a scale of 37.6 L. The RNA is first synthesized from linear DNA via an in vitro transcription

(IVT) step. It should be observed that the linear DNA template is defined as a starting material, and

therefore manufacturing of the template via plasmid DNA is flot included in the process. The

IVT

step

is followed by two enzymatic steps, i.e. the DNase I (this teaction is stopped with EDTA addition) and

proteinase K digestion steps, which aid in purification. The crude RNA is then purified through a two-

stage ultrafiltration/diafiltration (UFDF) step. Lastly, the RNA undergoes a final filtration before being

dispensed and stored trozen in EVA flexible containers.

A flow diagram is provided (Figure 3.2.5.2.2-1), presenting the process inputs and the process controls

for each step. The purpose of each step in the manufacturing process is sufficiently described. The hold

times, process parameters and corresponding acceptance criteria are listed for each step. It is noted

that flot alI process parameters are listed, but that the lists inciude alI critical and several non-critical

process parameters. In general, it is agreed that the key process parameters are described in section

3.2.S.2.2. However, for the

IVT

step, the added volumes of the enzymes T7 polymerase and

pyrophosphatase should be regarded as critical, unless justified. It should also be noted that future

changes to any at the process parameters listed in S.2.2, regardless of the ciassification as CPP or non

CPP, should be applied for as variation applications. A few concerns are raised regarding the incubation

time during GTP/N1-methylpseudo UTP bolus feeds, the transfers of the UFDF pool into a single PE

flexible container and DS filling volume range.

The Applicant explains that the UFDF membrane lifetime remain to be established and the concurrent

validation plan is found adequately described in the dossier. The strategy could be found acceptable,

provided that the Applicant will update the manufacturing process description with control of feed flow

rates, transmembrane pressure and membrane surface area.

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Figure 3.2.S.2.2-1. RNA Manufacturing Process

Step

ATP

LIlIQ

Ptot’ Inputs

heen CI?

Pioc Siep

Proce

(catiots

I mstb’.1psu&

UT?

cap

ho.-u

C,T1’

?roc ?arsimttrs

Ttmperslure I

Pgeczvrn

Àrtsusu Rste P T-rs,r.uie Ariatcr, Ratr

AT?

vciuuie

(I? a1uwe l,ar,I GI?

oleIia

vohsa

tsut,al

Nl-npci& UT?

;Iuticn rc1u,ne Z,r.tar

DNA

ccactatihc’ls,

Incebut,ou ross,

D’.guir GI? N!.mechyIpeudo

UT? Bota Trd ToiI

GI? 2.1. nttIyIpe& UT? Blu: Vehrm. Tir.rt TUT kcabusojs T,mt

—

bt,on, RNac wbt

IOX

aiptsoa buffer L,nesr UNA

Templ. Pvropbepbatse.

T’pc1tnersc

Watei

f’r

Iajecun

Cstcnnn

cbkudt

olutsca,

DNse I

EDTA

DNz:

I Dt:oi

Piocr

Ps1’n,,eH:

Tmpetanite I & 2, tsecn P,ste,

DNae

I Iant DNae I

Pttmat t

P,ctr.ei

Prot Psiosits

Tnsposrure 2 Aur.

,at.

Peu;e

h;tne.

tsas,K

Incutiz’a Tsae

IkI ixne

irsp

In-Prott

IPIM RNÄ

tr5tir,

hs:bacIa ea&-txn

Proc,- P,rfo,msnc,

AIt,,bui.

Asnsc,uss:sfsse d,tutti

buifri

fIac I tsffet fuiuIeca tnsff,i

—

[UFDF

Drug substance transpoftation

Reprocessing

Fuii

sad

flpt

]

Pro’ P.itm,te:

Osfilnation I

voTume

tats

H1d tuue

snap

FmiubnabuTezpH

In.l’,oc,

Ttst

IPIM

RNA

ctscert,ea

isuca

top ius

UFDF

atsOa) bsbtude

sxze

IPT-C ?.NA cancer,

i,c’nlI’TDf Pet

pie s

?roce’s P,its,tnsnce Air,ib,ue

Se1iYttId

Psoces Poramerer:

fl3I Ttsne Temp

Proce Le, tot cnanrt Air, ibure

Fiier

,oea,1r.’

set Sit’r

Y,e1

ino’.w

—-_______

Ding

Sebuance os

EVA

Ftexsbl. Ceatoosen l’FC

The drug substance is stored between -15 °C and -25

Transportation using an insulated shipper is

qualified for a shipping time up to 106 hours at -15 °C.

It is stated that if the post-use integrity test an the final 0.45/0.2 pm filter fails, refiltration is allowed.

It is clearly defined that reprocessing at the final filtration step is only aliowed once. This is found

acceptable.

Batch scale and definition

It is explained that commercial scale drug substance batches are executed at a scale of 37.6 L starting

volume for in vitro transcription (lyT). Ali material produced is purified by a single, two-stage

ultrafiltration/diafiltration (UFDF) to produce drug substance. The batch numbering system is

sufficiently described. Each batch is assigned one batch number for the entire process. This is found

acceptable. However, in addition, information an the final DS volume should be provided.

Control of materials

An adequate overview of the raw materials and solutions used in the Drug Substance manufacturing

process is provided. Limited acceptance criteria are iflcluded in a tabular format for ali raw materials

but represefitative CoAs should also be provided for the non-compendial materials. In general, the

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submitted information seem to support an appropriate quality of raw materials, however, several

concerns are raised at this point.

Startino materials:

The listed starting materials inciude ATP solution, CTP solution, GTP solution, N1-methyipseudo UTP

solution and 5’-cap solution and the linear DNA tempiate. The approach is acceptable. As the 5’-cap

structure is complex, additional information on its synthesis and discussion an its impurities are

requested. Clarifications are also requested on materials testing.

Linear DNA tern plate

BNT162b2 drug substance is rnanufactured by in vitro transcription using a linear DNA template,

produced via piasmid DNA (pST4-1525) from transformed DH1OB Escherichia coli cells.

The linear DNA template is flot part of the final product but defines the sequence of the mRNA product

and therefore it is fundamental to ensure its adequate control. Changes to the manufacturing process

of the linear DNA template (e.g. change to plasmid host cell) may result in a different impurity profile

in the active substance. Therefore, the level of details included in the dossier with respect to the

manufacturing process and the control strategy for this starting material, although shortly described, is

nat yet considered adequate to allow for a proper assessment.

The functionai elements of the pST4-1525 are sufficiently described in graphic and tabular forrnats and

the sequence is inciuded. However, details regarding the bacterial stram and the source and

generation of the pST4-1525 plasmid used rernain to be documented.

The cell banks involved in the plasmid manufacturing process are described. MCB and WCB

qualification tests are listed and include morphologic and genotypic identity, restriction map analysis

and DNA sequencinq, absence of contaminating bacteriophages, viability, plasmid retention and

plasmid copy number. Relevant specifications are set and data from the current MCB and WCE are

provided. The plasmid MCBs and WCBs are enrolled in a cell bank stability program consisting of

viability and plasmid retention assays conducted at ali stability time points. The strategy is, in general,

considered adequate, although some details are requested.

pST4-1525 is manufactured by a fed-batch fermentation process initiated from the bacterial working

celi bank

(WCB).

Foliowing fermentation, the cells are harvested and chemically lysed to recover the

piasmid DNA. After this lysis step, the circular plasmid DNA is purified by ultrafiltration/diafiltration

and anion exchange chrornatography. The circular plasmid DNA is filtered via 0.2 pm filtration and

stored frozen at -60 to -90 °C; the hold time for this intermediate is not deNned. The filtrate is

sampled for the circular plasmid DNA specifications. After thawing, the plasmid is linearized,

concentrated, filtered and stored frozen at -15 to -25 °C. No additional information nor data are

provided to support stabiiity. The filtrate is sampled for the linear DNA template specification. A list of

the raw materials as weli as the chromatography resins and filters used in the manufacture of the

linear DNA template is provided. Ali materials used are animal origin free and sourced from approved

su ppl iers.

Specifications for the circular plasmid DNA as well as for the DNA linear tempiate are provided.

Process- and product-related impurities including host cell genomic DNA, RNA, proteins, endotoxins,

bioburden and plasmid isoforms, for the piasmid DNA, are quantified routinely. The reference material

for plasmid identity testing is nat described. Resuits from three different batches are provided for the

circular and linearized piasmid and the proposed specification limits seem to be justified by the yet

limited available data. No descriptions of the analytical methods used for the control of the linear DNA

template nor evidence regarding their qualification/validation have been yet provided. This information

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is, however, considered criticai for quality of the final product. The Applicant is reminded that

impiementation of changes in the manufacture of the linear DNA tempiate should be appiled for in a

variation application.

Control of critical steps and intermediates

Process parameters and tests that are used to control the process and drug substance quality are

provided. The Appiicant claims that due to rapid development of additional process knowiedge, process

parameters and ranges are expected to be updated in a subsequent submission to the

MAA

prior to its

approval. This is found acceptable, but the Applicant is reminded that ali process parameters and

ranges should be sufticiently validated. Ali changes in future submissions prior to MAA or CMA approvai

should be clearly stated. Some ciarifications about the list of critical process parameters (CPPs), in

process tests for control (IPT-C), and hold times are already requested.

The in-process test methods are defined either as in-process testing for control (IPT-C) or in-process

testing for monitoring (IPT-M). The sole IPT-C is determination of RNA concentration in the

ultrafiltration/diafiltration (UFDF) pool (pre- or post-dilution) by UV spectroscopy. This method is

performed as described for the corresponding DS specification test. Three IPT-Ms are listed;

determination of RNA concentration in the proteinase K pooi by UV Spectroscopy (same as above),

bioburden and bacterial endotoxin testing. Ali three methods are applied to test the proteinase K pool

(post-hold), the UFDF pool (post-hold), and the UFDF end of diafiltration 2 retenate (pre-recovery)

samples. Bioburden and bacterial endotoxin testing are compendial methods.

Process validation

The process validation is ongoing at Wyeth BioPharma, Andover. For the process validation studies a

total of five vaiidation batches will be inciuded, ali these batches have been manufactured representing

the commercial batch size ot 37.6 L. Resuits are available for three out of the five consecutive batches.

The resuits from batches PPQ4 and PPQ5 are still pending.

No validation data are available to confirm consistent removai of impurities, which is not acceptable. Ifl

addition, residual DNA tempiate is present at higher level in PPQ3 batch (211 ng DNA

mg RNA) than

in PPQ1 and PPQ2 batches (10 and 23 ng/mg) which does flot confirm the robustness of DNase I

digestion.

The final filtration refiltration was validated at lab scale using a commercial scale filtration pooi and will

be confirmed at commerciai scale. This is acceptable.

Several validatfon studies are stili pending and will be updated once the data has been generated. ATP

and CTP volumes added at the beginning of

lyT

wete increased from the third PPQ batch and onwards.

The resuits for PPQ4 and PPQ5 batches are therefore necessary to confirm the consistency of the

process after this change. Therefore, a time-plan for the submission of these additional process

validation data should be provided before marketing authorisation approvai.

Hold times

It is stated that in-process pooi hold times are flot required for routine processing, but strategic hoids

in the process 24 hours to afd in manufacturing scheduling were validated. The small scale in-process

hold studies are intended to support biochemical stability at commercial scale. The hold times for the

Proteinase K pool, UFDF pool and DS before freezing as listed in S.2.2 are ali acceptably validated for

hold times 72 hours.

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Filter Qualification and Validation

The final filtration refiltration was validated at lab scale using a commercial scale filtration pool, and

will be confirmed at commercial scale, which is pending. This is acceptable.

Shitpin Performance Qualification

The shipping qualification strategy are described in detail and considered both thermal and mechanical

aspects of shipping. The shipping procedures and configuration for transport of frozen DS to the DP

manufacturing sites were validated to maintain product temperature in the acceptable tange for

durations up to 106 hours.

UFDF membrane lifetime

The strategy for UFDF membrane lifetime validation is to perform concurrent validation of the

membranes at commercial scale. Parameters related to performance and cleaning of membranes will

be evaluated as listed in Table 5.2.5-9. This strategy is found appropriate since control of process

parameters and IPC-tests are in place for every batch.

Manufacturing process development

Data for this section is pending.

Develorment history and Comparability

Process development changes were adequately summarised. Two DS processes have been used during

the development history; Process 1 and 2. Details about process differences, justifications for making

changes, and results from a comparability study is provided. The major changes between DS Ptocess i

and 2 are; increased process scale, DNA template changed from a PCR template to linearized plasmid

DNA, magnetic bead purification replaced with proteinase K digestion and UFDF steps.

No comparability study was provided for non-clinical versus clinical batches, but the batch analysis

results are provided.

The comparability study was performed between process 1 GMP batches and process 2 batches

manufactured at Andover and will be completed when ali PPQ data will be available.

In the comparability study a decrease in RNA integrit:y was observed for the Ptocess 2 batches

compared to Process i batches (78.1-82.8°h compared to 59.7%). After adjustment of process

parameters for

CTP

and ATP volumes batch 20Y513C501 (PPQ3) was manufactured with RNA integrity

level of 75%, more consistent with the Process 1 batches. No analysis of the capillary

electropherogram was provided. It is therefore not possible to conclude if the differences in RNA

integrity are quantitative or qualitative. Additional batch data are needed to confirm that the optimized

Process 2 allows to reach RNA integrity levels consistent with the Process 1 batches. (Part

MO).

Regarding the 5’ cap end of the DS, LC- UV/MS characterisation confirmed that the 5’-capped and

uncapped structures are the same in Process i and 2, but that there is a slight shift towards higher 5-

capping levels in Process 2. It is noted that the capped-intact RNA was flot measured, but only

deducted from the results of 5’-cap and RNA integrity. Therefore, this argument cannot be used to fully

confirm the comparability of Process 2 versus Process 1.

Furthermore, the poly(A)tail of the 3’ end was characterised by

LC-UV/MS. The

expected short (A30)

and long (L70) segments of the poly(A) tail were observed after RNase Ti cleavage. While the results

for the A30 segment were demonstrated to be comparable between Process i and Process 2 batches,

significant differences were identified for the L70 segment. As expected, poly(A) tail heterogeneity was

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observed both for Process i and Process 2 batches, due to transcriptional slippage. Longer poly(A) tails

were observed for the Process 2 batch, while the most abundant L70 segments of the Process i batch

were demonstrated to contain an additional cytidine residue. Differences in the poly(A)tail pattern were

observed when comparing the Process i and Process 2 DS batches. The differences in the extent of

cytidine monophosphate incorporation and transcriptional slippage needs to be further investigated and

the possible impact on efficacy and safety should be discussed. The only Process 2 DS inciuded in the

comparison was manufactured prior to the adjustment of CTP and ATP volumes. Resuits obtained an

the PPQ batches manufactured after adjustment (PPQ 3, 4 and 5) also needs to be presented.

The overall primary sequence of BNT162b2 drug substance was demonstrated to be comparable by LC/

MS/MS -oligonucleotide mapping. Circular dichroism (CD) spectroscopy confirmed that the higher

ordet structute of Process i and Process 2 DS batches were comparable.

To demonstrate functionality, the protein size after in-vitro expression of BNT162b2 drug substance

was determined using Western blot. The expressed protein sizes were demonstrated to be comparable

between Process i and Ptocess 2 batches. However, the method is only briefly described, and the

relevance of the resuits is therefore difficult to assess.

Critical Quality Attributes (CQAs)

A summary of the quality attributes with the rationale for the criticality assignment is provided. The

rationale for classification into CQA or QA is presented for each attribute and appears reasonable. The

identified CQAs are; RNA integrity, 5’-cap, PolyfA) tail, residual DNA template and double stranded

RNA (dsRNA). To be noted, for polyfA) tails, both percentage of Poly(A) positive mRNA molecules as

well as the length of the Poly(A) tails are considered CQAs. A related concern is raised in 5.4.

Process Development and Characterization

Data for this section is pending.

Process characterisation studies based an Cause and Effect Matrices (C&E) assessment, Failure Modes

and Effects Analysis (FMEA), design of experiments (DOE), using scale-down models of individual unit

operations, were/will be performed. To be noted, the overall control strategy inciuding the approaches

taken to identify critical process parameters (CPPs) are presented but some parameter and ranges

may be updated after PPQ and additional characterization studies are completed. As for assessment of

overall control strategy, a complete set of data and information is needed and therefore the final

evaluation of the control strategy cannot be made at this point.

It should also be noted that future changes to any of the process parameters listed in 5.2.2, regardless

of the classification of CPP or non-CPP, should be applied for as variation applications.

Initially, addition volumes for ATP and CTP were identified as non-CPPs as both were supplied in

theoretical excess. Following the Pfizer GMP campaigns and additional smalis scale studies it was

shown that these volumes could be limiting, and the ranges were widened at the higher end. The

approach to only change the higher end of the ranges need to be further justified and clarified. It is

noted that after the adjustment of these volumes the percentage of RNA integrity was increased to

levels more consistent with the Process i batches.

In the In vitro transcription (lyT) step T7 RNA polymerase and pyrophosphatase are added to start the

reaction. The ribonucleotide building blocks are assembled by the T7 polymerase. T7 polymerase is

magnesium dependent, but the magnesium can be chelated by pyrophosphate released by the addition

of each ribonucleotide to the growing chain. Pyrophosphatase is used to maintain sufficient levels of

free magnesium by breaking down the pyrophosphate. It is claimed that the added volumes of these

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two enzymes have been identified as non-CPPs as they are most likely to impact yield only. This

conciusion is flot agreed upon, the added volumes of the enzymes should be ciassifled as CPPs.

Risk Assessment of Process Related Imnurities

Data for this section is pending.

A safety risk assessment for potential process-related impurities included in the drug substance

process relative to patient safety is provided

ii,

this section. The potential impurities include small

molecules, enzymes and the NTP/Capping Structure. The sources of the impurities are sufficiently

addressed.

The safety risk assessment strategy involves comparison of the theoretical worst-case concentration at

impurities, assuming no removal, to calculated safety concern thresholds. If the worst-case level of an

impurity exceeds the pre-determined safety limits, any available commercial scale data for the specific

impurity will be provided in the relevant section and at a minimum will be monitored as part of process

validation to demonstrate consistent removal to acceptable levels.

The worst-case levels of NTPs, 5’ cap, small molecule process related impurities, RNase inhibitor,

DNase I and pyrophosphatase from the BNT162b2 drug substance manufacturing process were

calculated to be significantly below the pre-determined safety limits. This is found acceptable. The T7

RNA polymerase and proteinase K levels were further investigated and it was demonstrated that the

detected concentrations in the clinical, initial emergency supply and PPQ BNT162b2 DS batches were

well below the safety concern threshold. The Applicant states that data will be provided for additional

batches once testing is complete. This is found acceptable. However, the Applicant should provide data

on the T7 RNA polymerase and proteinase K levels in additional commercial scale DS batches, once

testing is complete. In addition, the Applicant should briefly describe the methods applied to determine

the concentrations of these two enzymes in the BNT162b2 DS samples.

Characterisation

Elucidation of structure and other characteristics

Analytical characterisation was performed an BNT162b2 drug substance batch 20Y513C101, which was

manufactured by DS Process 2 at commercial scale. This is found acceptable.

The physico-chemical characterisation involved primary structure, 5’ cap structure, poly(A)tail and

higher order structure evaluation. Primary structure was confirmed by oligonucleotide mapping and the

ofthogonal method, RNA sequencing using the Illumina MiSeq Next Generation Sequencing (NGS)

technology. The results confirm the RNA sequence. The 5’-cap and 3’ poly A tail were confirmed by two

separate LC-UV/MS-methods. It was demonstrated that the predominant form of the 5’ terminus is the

full-length nucleotide sequence with the 5’-Cap, but that there are also other minor species including

phosphorylated, truncated and extended species. Analysis of the 3’ poly A-tail demonstrated that

BNT162b2 DS contains the expected tail, but that there is some heterogeneity due to transcriptional

slippage. Un-capped RNA and/ar truncated/extended forms are possible at minor to trace levels but a

precise quantification of each uncapped ar incompletely capped specie was flot provided. It is also not

specified if and how these species contribute to the potency of the BNT162b2 DS. The higher order

structure of BNT162b2 mRNA DS was characterized in solution using circular dichroism (CD)

spectroscopy. Overall, state-of-the-art methods were applied for physico-chemical characterisation and

the results confirmed the expected sequence and quality attributes.

A severe deficiency of the characterisation section is that no biological characterisation is presented

and that the mode of action is not described. This is flot found acceptable and the dossier should be

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updated with relevant information. Even though full biological characterfsation is flot possibie to

perform on DS, the strategy to determine potency and relevant functional assay(s) should be

described in section 3.2.S.3. Results obtained on DP could be inciuded, to demonstrated functionality.

Furthermore, it is observed that in the Deveiopment History and Comparability section (3.2.S.2.6), the

expressed protein size is evaluated by in vitro expression followed by Western blot. Resuits obtained

by this method could be cegarded as biological charactecisation and should be included in section

3.2.S.3. The method needs further description and the results should be sufficiently characterized.

Impurities

Process-related and product-retated impurities as well as potential contaminants are described in this

section. Five batches were evaluated for impurities, Le. clinical, initial emergency supply and PPQ

batches. It is noted that this section is incomplete and will be updated after PPQ completion.

The sole product-related impurity addressed is double-stranded RNA, derived from the in-vitro

transcription reaction. Resuits from the five DS batches demonstrate that the level of double sttanded

RNA is low, acceptable and consistent.

In addition to double stranded RNA, there are more product-related impurities, i.e. truncated RNA, also

referred to as fragmented species. Truncated RNA is reflected in the DS specification in terms of RNA

integrity. However, the characterisation of BNT162b2 DS is currently not found acceptable in relation

to the CQA RNA integrity. Significant differences between batches manufactured by Process i and 2

are observed for this specific attribute. Even though two methods, namely agarose gel electrophoresis

and capillary gel electrophoresis, have been appiled to determine RNA integrity of BNT162b2 DS, no

characterisation data on RNA integrity and truncated forms is presented and the potential safety risks

associated with truncated RNA isoforms are not addressed. This is especially important considering

that the current DS and DP acceptance criteria attows for up to

fragmented species. Therefore,

the dossier should be updated with additional characterisation data and discussion ci, mRNA integrity,

this is considered a major objection.

Residual DNA template is

process-related impurity derived from the linearised DNA template added

to the in-vitro transcription reaction. Residual DNA template is controlled by qPCR as defined in the DS

specification, and the levels for ali five batches are demonstrated to be well below the acceptance

criteria. However, a drift towards higher level was observed for the third PPQ batch and therefore

additional batch data are needed to conciude on the consistent removal of this impurity. Additional

process-related impurities, including nucleoside triphosphates (NTP5) and capping structure, small

molecules, and enzymes, are evaluated and assessed in Section 3.2.S.2.6 Risk Assessment of Potential

Process Related Impurities. Taking section 3.2.S.2.6 into account, the process-related impurities are

sufficiently described. Some uncertainty remains regarding the approach to determine the levels of T7

RNA polymerase and proteinase K.

The potential contaminants described in this section are endotoxin and bioburden. Acceptable results

are shown for the Proteinase K pool, UF retentate pre recovery, UF-product pooi and the drug

substance.

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Specification, analytical procedures, reference standards, batch a nalysis,

and container ciosure

Specifications

Table S. 4-i. Specifications

Quality Attribute

Composition and Strength

Ciarity

Coloration

Appearance (Ciarity)

Appearance fColoration)a

6 NTU

Not more intensely coloured than evel 7

of the brown (B) colour standard

Content (RNA Concentration)

Potentiometry’

UV Spectroscopy

7.0

0.5

2.25

rir

Analytical Procedure

Acceptance Criteria

0.25 mg/mL

Identity

Identity of Encoded RNA

Seguence

RTPCRb

Identity conflrmed

Purity

RNA Integrity

5’- Cap

PoIy(A) Tail

Capillary Gel Electrophoresis

RP-HPLC

ddPCR

50%

intact RNA

50%

70%

Process Related Impurities

Residual DNA Template

qPCRb

330 ng DNA/mg RNA

Product Related Impurities

dsRNA

Safety

Bacterial Endotoxin

Bioburden

a. Compendial

b. Assay not performed on stability.

Abbreviations:

reaction; ddPCR

NTU

Immunoblot’

1000 pg dsRNA/]g RNA

Endotoxin (LAL)0

Bioburden

12.5 EU/mL

i CFU/ 10 mL

Nephelometric Turbidity Units; B

brown; RT-PCR

teverse transcription polymerase chain

droplet digital PCR; qPCR

quantitative PCR; dsRNA

double stranded RNA;

LAL

Limulus amebocyte lysate; EU

endotoxin unit; CFU

colony forming unit

The proposed specification

for

drug substance is at large found acceptable with respect to the analyses

chosen for routine release testing. The CQAs RNA integrity, 5’-cap, PoIy(A) tail, residual DNA template

and double stranded

RNA

(dsRNA) are ali inciuded in

the release specwication. However, the length of

the poly(A) tails in BNT162b2 DS is important for RNA stability and translational efficiency and

therefore should be included in DS release testing. It is also noted that no method references are

included, this needs to be updated.

Potency testing is flot inciuded in the control of DS but instead is performed at the level of DP release.

Considering the nature of this produci, the approach is endorsed.

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Analyucal procedures and reference standards

Analytical procedures

Ali analytical methods used for testing of the drug substance are described in the dossier.

The following tests are performed in accordance with Ph Eur; ciarity (Ph Eur 2.2.1), colour (Ph Eur

2.2.2), pH (Ph Eur 2.2.3), bacterial endotoxins (Ph Eur 2.6.14) and bioburden (Ph Eur 2.6.12).

A general comment which applies to ali non-compendial analytical methods is that rather brief details

are given. Some of the analytical methods are not presented in sufficient detail and often method

descriptions are based on “examples” of procedures, controls and standards as well as on “typical”

system operating parameters. This hampers a full understanding the operation or, sometimes, the

scientific basis of the assay. Furthermore, since several of these assays are none standard and

complex, this interferes with assessment of suitability. The lack of sufficient information on critical

reagents, standards or equipment hinders regulatory control of criticai aspects of the assays. Several

concerns are raised for specific assays requesting additional information an critical procedures,

reagents, standards and equipment.

It is claimed that the analytical methods were validated against the parameters presented in ICH

Q2(R1). However, the validation summaries presented are far too brief to be able to conclude on

suitability of the in-house analytical methods. The quality of BNT162b drug substance cannot be

properly assessed, if the reliability of the analytical methods cannot be guaranteed.

Capillary gel electrophoresis (CGE) is used to determine the percent integrity of RNA in both drug

substance (DS) and drug product (DP). The test sample is subjected to a denaturant containing

formamide that unfolds the RNA and dissociates non-covalent complexes. When subjected to an

electric field, the denatu red RNA species migrate through the gel matrix, as a function of Ienqth and

size, toward the anode. An intercalating dye binds to RNA and associated fragments during migration

allowing for fluorescence detection. The intact RNA is separated from any fragmented species allowing

for the quantitation of RNA integrity by determining the relative percent time corrected area for the

intact (main) peak.

Reversed Phase-High Performance Liquid Chromatography (RP-HPLC) is used to measure the relative

amount of 5’- capped RNA species. Test samples are digested using RNase H followed by affinity

purification and (RP-HPLC) with UV detection. After an annealing process to a biotinylated probe

complementary to the last 26 bases of the 5’ end of the RNA, samples are digested with RNase H,

followed by streptavidin-matrix based affinity purification of the resultant duplexes from the much

larger mRNA remnants. The short oligonucleotide capped, and uncapped species are eluted from the

streptavidin-matrix, and relative quantification of the 5’-cap is accomplished by RP-HPLC analysis of

the ensemble of RNA capped and un-capped molecules. The relative amount of capped species is

determined by dividing the capped species signal by the total species signal.

The in-house analytical methods for CGE and RP-HPLC are at large well described and includes details

on typical test conditions, operating parameters, representative electropherograms and

chromatograms as well as information on system suitability testing.

An RT-PCR method is used to determine the identity of the encoded RNA sequence, a quantitative

polymerase chain reaction (qPCR) analytical procedure is used to quantify the residual DNA template

and an immunoblot analytical procedure is used to detect double stranded RNA (d5RNA) in BNT162b2

drug substance. Ali these assays are deemed suitable for their intended purpose and, in general,

although brief, the descriptions provided are considered relevant. Several concerns regarding

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bilag 9

additional details on method description, controls and in some cases further clarifÏcations of criteria

established to support method suitability are raised.

The UUPCR technology is ptoposed for the quantification of the poly(A) tail in the messenger ribonucleic

acid (mRNA). The technical procedure is considered, in general, sufficiently described but the suitability

of this method for the intended purpose needs additional clarifications. The rationale by which the

method determines the percent poly(A) relative to the theoretical input (which is flot clearly described)

should be further addressed.

Release and stability testing can be performed at several testing sites. However, the method transfer

plan or activities was not submitted in the RR. It should be noted that, if method transfer was

will be

performed, the following information are requested. For the non-compendial tests, it should be

confirmed that the validation acceptance criteria for the receiving sites will be the same as for the

transferring site (which will be assessed during the RR). For the analytical methods where comparative

analysis will be proposed, it should be confirmed that the acceptance criteria will be the same as for

the intermediate precision validated at the transferring site (and assessed during RR).

Reference standard

The current reference standard is referred to as the Clinical Reference Material (CRM). It is stated that

the CRM will be used for clinical suppifes, process validation and initial commercial suppiles. The CRM is

prepared from the GMP BNT162b2 DS batch 20Y513C201. Release data is presented in the dossier.

The intended storage condition is -20 ±5 °C, but an alternative storage condition of -60 to -90 °C is

also evaluated. A stability protocol is provided. There are several concerns regarding the reference

standard, including the suitability of the batch chosen as CRM, If additional standards have been used

during early development and issues related to the formal stability protocol. It should also be clarified

for what release and stability testing methods the reference standard is used and will be used in

future.

In future, a two-tiered system for future commercial reference material will be implemented. A PRM

and an initial WRM will be established in 2021 for the drug substance reference material. The PRM will

be the standard against which WRMs are qualified and the PRM will be intended to last the lifetime of

the commercial product. The Applicant claims that further information on the selection, preparation,

qualification and stability of the PRM and WRM will be provided in the future.

The use of a two-tiered system is encouraged. Tt is understood that the PRM and WRM is not yet

established. The Applicant is reminded that the implementation of the two-tiered system should be

applied for in a Type II variation application. Alternatively, information on the preparation, qualification

and stability evaluation of the PRM and WRMs should be inciuded in a PACMP.

Batch analysis

Batch resuits are presented for DS batches used for nonclinical toxicology, clinical trials, process

performance qualification (PPQ), emergency supply, and stability.

In general, the results obtained using the commercial process CDS Process 2) demonstrate batch to

batch consistency with a few exceptions. The resuits for RNA integrity are higher for batch PPQ3

(20Y513C501) as a volume adjustments was made for ATP and

CTP

volumes before manufacturing of

this batch. Batch results should be presented for the two newly manufactured batches PPQ4 and PPQS

verify that the commercial manufacturing process consistently results in RNA integrity levels similar to

levels achieved in process i batches.

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Justification of specification

The rationale used to establish the acceptance criteria is described in detail and based a limited data

set representative of BNT162b2 DS manufactured at the intended commercial scale and process. It is

endorsed that the specification for BNT162b2 DS will be reassessed when more batches have been

manufactured. However, from the available data, it appears that RNA integrity, dsRNA, Poly(A) tail and

5’-cap acceptance criteria are too wide and need to be tightened yet to better reflect data obtained

from available lots used in cllnical studies (and considered cllnically qualified) and data from lots used

for PPQ.

Container ciosure

The drug substance is stored in 12 L or 16.6 L single-use, flexible, disposable bags of ethylene vinyl

acetate (EVA). Compliance with Ph. Eur. 3.1.7 Ethylene-VinylAcetate Copolymer for Containers and

Tubing for Parenteral Nutrition Preparations is claimed. Schematic drawings of the bags are provided in

the dossier but no specification or certificate of analysis for the container or the EVAM contact layer are

included.

The information regarding container closure system is in general acceptable. However, the Applicant

should verify compliance with Ph. Eur. 3.1.7 with a certificate of analysis for one representative batch

of the EVAM contact layer.

A controlled extraction study has been performed on the EVA container film; ali the compounds were

observed below the Safety Concern Threshold of 1.5 pg/day TDI. Considering that the intended

storage of the DS is -20 °C, a temperature which has a lower risk of leachables, it is reasonable that

no specific leachable compounds have been selected for further studies. Nevertheless, a leachable

study will be initiated to detect semi quantitate unexpected leachable compounds equal to or greater

than 1.5 pg/day TDI. This approach can be accepted.

Stability

The initial proposed commercial shelf life at the dtug substance is 6 months when stored at the

intended storage condition of -20 ± 5°C in EVA bags. The initial shelt life is based on the currently

available data from stability studies utilizing material from three clinical DS batches manufactured

using Process i and two cllnical DS batches (up to 3 months data presented) and three process

validation batches manufactured by Process 2 Cup to i month data presented).

It is claimed that the results of the comparability studies support that stability data generated using

drug substance manufactured using Process 1 can be considered predictive of the drug substance

manufactured by Process 2. This conclusion is not fully agreed with as detailed above in section S.2.6.

Based on the currently very limited stability data presented for process 2 batches (only 1-month data

available for one batch) no conclusion can be drawn in relation to the proposed shelf life for the DS.

Therefore, in ordet to support shelf life setting for drug substance updated reports from the ongoing

stability studies on the primary batches (including data from the ongoing process validation batches)

should be provided.

is stated that sponsor will extend the assigned shelt life without notification providing the real time

stability data at the intended storage condition is acceptable and within commercial specifications. This

kind of extensions can be accepted for clinical trials but flot atter marketing authorisation approval.

This statement should be removed from the dossier.

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2.L3. Finished Medicinal Product

Description of the product and Pharmaceutical Development

The BNT162b2 drug product is supplied as a preservative-free, multi-dose concentrate to be diluted for

intramuscular injection, intended for 5 doses. The drug product is a sterile dispersion of RNA

containing lipid nanoparticies (LNPs) in aqueous cryoprotectant buffer.

Each vial, containing 0.45 mL of the drug product at pH 7.4 is designed to deliver a total of 5 doses

after dilution by addition of 1.8 mL of sterile 0.9% sodium chloride solution tor a total votume of 2.25

mL, with each dose containing 30 pg of RNA in 0.3 mL. There is no manufacturing overage. The

justification for the overfill is discussed, but the final volume exceeding the nominal volume is

questioned.

The drug product

supplied in a 2 mL glass vial sealed with a bromobutyl rubber stopper and an

aluminum seal with flip-off plastic cap.

The composition of the drug product, including amounts per vial and function and quality standard

applicable to each component, are given in Table P.1-1.

AlI ingredients, including process aids used in the manufacture, should be specitied in the composition

together with a footnote that they are processing ald removed during manufacturing. Therefore,

ethanol and citrate buffer and the excipients present in the DS (HEPES and EDTA) should be added to

the composition.

Table P. 1-1. Composition of BNT1 62b2 drug product, multi-dose vial (225 ,ig/vial).

Name of

Ingi’edients

BNI162h2 dnie

stthstance

AL(-0315

AL(-0159

DSPC

Cbolesterol

Sucrose

Sodhun chioride

Potahuu ctdoride

Dibasic sodium

phosphate.

dilwdrateb

Refei’ence to

Standai’d

Iii-house

specification

Ifl-house

specificalion

Iii-house

specification

In-house

specification

Ph. Fur.

Functioii

Active hisredient

Fiuscijonal lipid

Funetional lipid

Stnicluial lipid

Stmctural lipid

C’ryop;otecta;it

Buffer component

Coucentratiou

(mg.mL)

0.5

.17

0.89

1,56

3. la

103

0.15

1.08

Amount

per

per vial

225

3.23 mg

0.4 nis

—

dose

30 ie

0.43 inc

0.05 mc

0.09 mc

0.2 me

6 mg

0.36

1112

0.01 mg

0.0 mg

1.4 mg

46 m

2.7 mg

0.0 mg

0.39 mg

Monohasic

Ph.

Fur.

Buffer component

0.15

0.0 mc

0.01 mg

potassium

pliosphate

Water for Injectioll

Ph. Fur.

Solvent. vehicle

g.s

q.s

a. Vahies are i’mmdecl to inaintain the same level of precision as the label dum, with

b’ailhlg zeros flot

s1ion,

where apphcable. for exaniple. 46 ing sucrose is rotmded from 46.35 mg (103 mg mi).

b. Dibasic sodium phosphate. clitydrate is uamed as disoclium phosphate diliydrate iii the Ph Fur.

c. Monobasic poiasimn phosphate is named as ijotassium dihydroceii phosphate in the Ph. Fur.

Ähhreviation:

ÄLC-03 15

(t3-hydroxvhulyl)azanedivhhisthexiine-6.1-divl)his(2-hexyldecanoate)

ALC-0159 2’[fpolvethvlene a1vco1l-2000]-N.N-ditetradec1acetamide

DSPC

i .2-ditearovl-su-clvceao-3-phosphocho1ine

q.s.

quannim satis cus much as mi suffice)

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Ali excipients except the functionai lipids ALC-0315 and ALC-0159 and the structurai lipid DSPC comply

Ph.

Eur. grade. The functional lipid excipients ALC-0315 and ALC-0159 are classified as novel

excipients. Both structural lipids DSPC and cholesteroi are used in several already approved drug

products. DSPC is used in several products approved in the

(Marqibo, Doxil, Ambisome, Onpattro),

though flot by the same route of administration. Furtherjustification that DSPC is flot a novei excipieflt

is requested.

The vial, stopper and seal components are compliant with the appropriate

Ph.

Eur. monographs for

primary containers and ciosures.

Pharmaceutical development

Formulation de velopment

The section on formulation deveiopment describes and justifies the chosen formulation and is

sufficiently comprehensive.

The formulation deveiopment studies of the RNA containing lipid nanoparticies have been thoroughiy

Uescribed. The development of a robust LNP formulation platform was performed at Acuitas

Therapeutics. Studies are comprehensiveiy described and were performed with availabie drug

substance, representative of the mRNA platform and inciuded in the drug product.

The LNPs consists of four lipids, each has a functional or structurai purpose. The ionizable cationic lipid

ALC-0315 interact electrostatically with negatively charged nucleic acids and encapsulate the mRNA.

The PEGylated lipid ALC-0 159 is preferably inserted at the LNP surface as a steric barnet to

interactions with surfaces or other LNPs to avoid aggregation during storage. The phospholipid DSPC

and cholesterol are structural lipids providing a stable bilayer and enabling mobility of the iipid

components in the LNP structure.

The formed RNA-containing LNPs are solid particies relatively homogeneous in size, largely sphericai in

shape and has a neanly neutral surface. Furthermore, the accumulated batch-data to date show a

consistent manufacturing of lipid nanoparticies both with respect to size and polydispersity.

Upid nanoparticie ILNP)

C5nk ii5i

i-e

per pd

60-120cm

Cnitical quality attnibutes related to LNP formation and payload deiivery ane primarily LNP size,

encapsulation efficiency, and in viva potency (RNA integrity). Additionaily, surface area is considered

critical to avoid aggregation both during storage and with serum components in vivo. The ratio cationic

lipid to RNA (N/P) is also critical for formation of LNP. An access of cationic lipid is required and a ratio

of about

is found reasonable.

The DP is stored frozen at the recommended storage temperature of

-90

-60°C.

Stability studies ate

ongoing for the determination of DP shelf-life.

The same DP formulation composition has been used throughout the nonclinical and clinical studies

and will also be used for the manufacturing of the pending full scale commercial PPQ-batches.

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There are no formula overages in the drug product, only an overfill which has been acceptably justified

ensurfng that five doses can be removed from the multi-dose vial and delivered.

Screening studies were performed to confirm that the ALC-0315/ALC-0159/DSPC/CHOL at molar ratio

47.5/10/40.7/1.8 with a ratio of cationic lipid to RNA (N/P ratio) of 6.3 provide LNP with acceptable

quality and stability. Physicochemical and biological properties were studied (density, viscosity, DSC

characteristics). Moreover, size distribution and particle shape were studied showing a narrow

distribution with a hydrodynamic radius and an almost spherical shape in the entire size distribution.

The zeta potential was narrow and monomodal. The pegylated surface of the LNP5 was studied

showing consistence with the proposed LNP architecture: presence at the surface of ÆG and

hydrophilic head of ALC-0315. While the

effort

made by the applicant to provide sufficient development

data in a very brief time is acknowledged, and taking into account that some additional heightened

characterization information will be added, the formulation development lacks some characterisation

studies showing the homogeneity of the suspension during storage at long-term or accelerated

conditions, after freeze/thaw, or after dilution with 0.9% NaCI.

Manufacturing process development

The development history of the drug product is sufficiently described.

The initial LNP and drug product formulation processes were developed at Acuitas Therapeutics,

followed by scale-up and manufacture at Polymun Scientific for clinical trial material and emergency

supply. The process has been transferred to Pfizer commercial facilities in Kalamazoo, MI, USA, and

Puurs, Belgium, for rnanufacture of later clinical materials (Puurs), emergency supply and commercial

supply.

The DP analytical comparability evaluation employed release testing and extended characterization

methods. It is agreed that comparability has been reasonable demonstrated between the cllnical

supply lots manufactured with the “classical” LNP process and the representative emergency supply bt

manufactured with the “upscale” LNP process with only small differences noted.

It is stated in the dossier that the applicant has a plan for a comprehensive demonstration of

comparability among cilnical supplies and the commercial product including an assessment of the

starting drug substance batches, raw materials (e.g. ALC-0315, DSPC and cholesterol) from different

vendors, process designs and comprehensive characterization of the resulting product quality. Data for

this section is pending and will be updated once the data has been generated, analyzed, and verified.

Four commercial PPQ-batches will be manufactured in November and December 2020. The results for

the comparability of the commercial PPQ-batches versus the clinical supply batches of DP is pending

and will be provided for assessment during the procedure.

In summary, no final conciusion on comparability can be drawn until ali comparability data

among clinicai suppiies and the commercial product (PPQ-batches) will be provided for

assessment.

Critical Quality Attributes inciude appearance, visible particulates, subvisible particles, pH, osmolality,

extractable volume, lipid identities and contents, RNA identity and content, LPN size and polydispersity,

RNA encapsulation, RNA integrity, 5’-cap, poly(A) tail, in vitro expression, endotoxins, sterility,

container closure integrity. Even though the risk assessment was not explained in detail, no issue is

raised on that point since the DP specification contains the expected parameters.

The development of the manufacturing process is extensively described, and critical process

parameters are defined. Process characterisation studies based on Cause and Effect Matrices (C&E)

assessment, Failure Modes and Effects Analysis (FMEA), design of experiments (DOE), using scale

covIo-19

mRNA vaccine fnucleoside modified)

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down models of individual unit operations, were

will be performed. It is noted that some resuits of

process characterisation studies are pending. The overall documentation related to criticality

assignment and NOR/PAR establishment will be assessed when completed. In addition, it is highlighted

that for the process characterisation studies already presented, the level of information was not

sufficient to allow assessment. Therefore, the PAR5 are flot considered acceptable at this stage.

The lipid nanopafticle (LNP) formation is one critical manufacturing step. The process development is

described and physicochemical properties (e.g. LNP size, polydispersity, RNA encapsulation, lipid to

RNA ratio (N/P) as well as LNP topology by X-ray scattering) has been evaluated during upscale. The

provided results are comparable. The tested parameters are considered relevant, covering the critical

attributes size, shape, encapsulation and lipid to RNA molar ration.

The in-process hold times, dilution and mixing of DS parameters, and lipid weight and organic phase

mixing parameters will be studied during PPQ. For buffer exchange and concentration step, residual

ethanol and citrate should be studied during PPQ and process validation. Process characterisation

studies were satisfactorily provided for DS thawing, sterile filtration, aseptic filling, stoppering, sealing

and capping, and freezing steps. However, PPQ data will be needed to verify the filling weight of

BNT162b2 filled at the commercial filling lines. Moreover, no development data showing homogeneity

of LNP or RNA concentration in the vials during filling process was provided. Drug product robustness

to freezing and warming during storage was studied and confirmed that BNT162b1 quality was flot

impacted by different thawing processes, but this will have to be confirmed for BNT162b2 DP.

Overall control strategy was presented but some pararneter and ranges may be updated after PPQ and

additional characterization studies completed. As for assessment of overall control strategy, a complete

set of data and information is needed, this document will be assessed when finalised.

The analytical testing strategy of drug product has changed throughout the development and these

changes have been described. Bridging studies have been performed for analytical tests that have

been changed or replaced (subvisible pafticles, identity of encoded RNA sequence and RNA integrity).

This is found acceptable.

Container closure system

The development of the container closure system is sufficiently presented. The primary packaging is

composed of glass vial and rubber stopper and are compliant with the compendial requirements of Ph.

Eu r.

Controlled extraction studies have been performed on the bromobutyl rubber stopper. Leachables

studies are planned to be set up the applicant should commit to provide the updated results from the

leachables study for assessment.

Microbioloqical attributes:

Sterility and endotoxin testing is performed at Drug Product (DP) release. A rapid sterility test may be

utilized. CCI will be verified by dye ingress testing or head-space analysis. These tests were

demonstrated to be able to detect CCI failure.

Compatibility

The drug product is frozen, and after thawing, the solution/suspension must be diluted with sterile

0.9%

sodium chloride solution. The studies described have been performed to assess physicochemical

stability of the DP after dilution with 0.9% sodium chioride solution in the original glass vial as well

with commonly used commercially available administration components and using worst-case

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conditions for dosage and administration in the cilnical setting. The thawed hold time (in-use period) of

und iluted DP are ongoing as part of the stability program in section P.8.

Resuits presented support physicochemical stability of DP diluted in

0.9%

sodium chioride solution for

up to 24 hours at ambient or refrigerated temperatures and compatibility with dosing components

(syringes and needles) for up to 6 hours. Furthermore, a microbiological in-use hold time study was

performed by a challenge test inciuding five compendial micro-organisms. No significant growth

(>0.510gb from the start-point) was observed for any of the microorganisms within 12 hours of

inoculation with storage at 20-25°C of diluted DP in

O.9%

sodium chloride solution. However, while the

representativity of 005 mg/mL concentration against the 0.1 mg/mL concentration is accepted, there

was no confirmation that the analytical methods are valid at this dilution, and the in-use specifications

should be the same as the sheif-life specifications. It is noted, however, that this section may be

updated as additional studies are completed.

Compatibility of drug product is at large acceptably demonstrated by the dilution and administration

simulation studies performed.

Manufacture of the product and process controls

Table P.3-1 Iists the sites that have responsibilities in the production of BNT162b2 drug product and

their specified functions.

COVID-19

mRNA

vaccine (nudeoside moditied)

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Table P3-1. Sites and responsibilities for BNT162b2 drug product manufacture

Sut

Pfirer .1anufactunng Belgium NV

Rijksweg 12

Puurs. 2SC

Belrium

Responsibffit

DIF rczucn and hulk dnag produnt formulaton

FilI and finash

Pranan niachnng

Sec cndar: packaeans

P.e1eae and atabilit-: testrna :Ccmgc2iticn.

Å±ventncur Åeenii

Batch releare b Quahfied Perron ur EEA [European

Economic Area

Release and rtabilitv testing (Comporition and

Sucne±, idenntv Potencv, Punft Ådtenutous

Åsen&i

Wvetk BicPhanna DivisirTa ofWve±

Pharmaceutic als LLC

1 Burt Road

Åndoven

:.IÅOIS!D

Unred Stater

Pfirer Inc

S_5 Cher.erfietd Parlo.vav West

Ches:erfield.MO

6301

United Stater

P&er Ireland Phamsareutcals

Granse Castle Buamera Paal

Ciondaikan. Dublin 22

Ireland

Hospua Zagreb Ltd.5

Pnidnidka

rene

10291

&iaone Brdr

sdr

Croatsa

SOS Lab Simon SÅ

Vieat Chenmn du Poste 10

l\asTe

1301

Belgaum

BiolClech ylanufatarmns GmbH

Kupferberetertasre 1-19

15116 Mamo

Oernaanv

Fele&e and stabikt-- teatina Comporition and

Snenat Idemitv. Poteucv. Punw Advenutous

»erna’

Releare and stabihtv testmg Jdentrt- Compositiont

Releare testing (Stenhtv)

Release teatma Stenbt

Batch releare by Qalified Person mEfl [European

Eronomir Årea1

a. The legal er.ut’ time change from Wvetb BioPhaama Dm non of Wve:h Pbamurccuticala was chanred a:

-Jac acqmaition by Pfizerm 2009, sance Uret the Wyech Pbarmaceuticala manufactunng site ur »doven

Masrachusetta belongs to Pfizefa produntion attea and is embedded ur Pficefs Ola? avatem POrer trall be

trd red ±rcush:ut the CTfl

b Ho;pL-ais a whollv onned subsidsan of POrer Inc

The DP is manufactured tested and batch released by Pfizer Manufacturing Belgium NV, Puurs,

Belgium. Batch release can also be done at BioNTech Manufacturing GmbH, Mainz, Germany. Several

testing sites are listed, in addition to Pflzer, Puurs, Belgium. Some clarifications are requested for GMP

activities of sites located in Europe (MO). Moreover, as Mutual Recognition Agreement is not in force

for human vaccines, the provided documentation for sites located in the USA is not considered

sufficient (MO).

The manufacturing process inciudes lipid nanoparticie (LNP) fabrication and bulk drug product

formulation followed by fill and finish. The target drug product batch size is 139 L (approximately

309,000 vials). The batch formula is provided but iacks process aids.

LNP fabricaUon and bulk drug product formulation

The frozen drug substance (mRNA) is thawed and diluted in water for injection to a target

concentration of 2.0 mg/mL. The lipids are diluted in ethanol. To form the LNPs the aqueous phase

with mRNA and the organic phase with the lipids are fed into one or more parallel T-mixers with pre

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set flow rates to get 3:1 volume ratio. The LNP bulk is then first buffer exchanged with citrate buffer to

remove ethanol from the suspension then with phosphate-buffered salme (PBS) at pH 7.4, suitable for

intramuscular administration. Sucrose is added as cryoprotectant, the concentration is adjusted, and

the solution mixed until homogenous. Hold times during the bulk drug product formulation process are

established.

Sterile filtration and aseptic filling

The bulk drug product is sterile filtered into a holding vessel using two sequential redundant sterilizing

grade filters. Integrity of these filters are controlled by pre- and post-use integrity testing. A sample is

taken for bioburden prior to filtration. The holding vessel is aseptically connected to the filling line and

then sterile filtered bulk drug product is aseptically filled into sterile vials and capped. Vials are l00%

inspected for defects either through automated visual inspection or manual visual inspection. Inspected

vials are mndividually labelled and packed. AlI hold times following sterile filtration will be within the

validated media fill times, ensuring acceptable microbial control during the drug product manufacturing

process.

Controls of critical steps and intermediates

Critical manufacturing steps are discussed, and relevant in-process controls are applied.

Residual ethanol is flot controlled in-process or in the final drug product specification. Data provided

demonstrates that ethanol is sufficiently removed in the final drug product. Absence oftest is therefore

considered acceptable.

The lipid nanoparticle (LNP) formation is one critical manufacturing step and some additional

information is requested regarding this step such as that a drawing of the T-mixer should be provided

as well as the number T-mixers defined.

Process validation and/or e valuation

No full commercial scale batches are included in section 3.2.P.3.5 and the applicant states that “Data

for this section is pending and will be updated once the data has been generated, analysed, and

verified.”

However, it is stated in the dossier that four commercial PPQ-batches will be manufactured in

November and December 2020. These batches will be executed according to defined protocols and

evaluated with predetermined acceptance criteria. Furthermore, these batches will be used both to

demonstrate the comparability of the commercial PPQ-batches vetsus the dinical supply batches as

well as for process validation of the manufacturing process of the drug product. In addition, validation

data on process hold-times, shipping validation and verification of mn-process test methods are

mncomplete. Since ali these vaiidation data are pending, no final conciusion on process

validation in section 3.2.P.3.5 can be drawn until these data are provided for assessment.

Media flIls have been performed to validate the aseptic filling process and were run

iri

accordance to

guidelines. Results have been provided from three consecutive simulation studies and gave satisfactory

results without any contaminated units. Results for the media fill cover the maximum process time for

the manufacturing of drug product (maximum filling time is 112 hours) and simulate worst-case

manufacturing conditions. The media fill validation demonstrated that aseptic conditions are

maintained during the filling process.

Acceptable information has been provided for filter validation on the 0.2 pm-filters used for sterile

filtration, describing the material, pote size and surface area. AlI study resuits met the predetetmined

acceptance ctitetia and the studies for microbial tetention, membtane compatibility, extractable

COVJD-19 mRNA vaccine fnucleoside moditied)

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substances and integrity test determination have shown that the 0.2 ijm-filters are appropriate for

sterile filtration of the drug product. However, the applicant should clarify if the 0.2 pm-filter used for

bioburden reduction is identical with the 0.2 pm-filters used for sterile filtration.

Control of exciients

ALC-0315 and ALC-0 159 are novel excipients, not previously used in an approved drug product within

LU. Additional information is provided separately in Section A.3.

DSPC is a non-compendial excipient sufficiently controlled by an in-house specification.

Cholesterol is sufficiently controlled according to the Ph. Eur. monograph with additional tests for

residual solvents and microbial contamination.

The other excipients (sucrose, sodium chioride, potassium chloride, disodium phosphate dihydrate,

potassium dihydrogen phosphate and water for injection) are controlled according to respective Ph.

Eur. monograph. However, appropriate documentation for processing aids (ethanol and citrate buffer)

and for drug substance buffer (HEPES and EDTA) should be provided.

Product specification, analytical procedures, batch analysis

The release and stability testing specifications for BNT162b2 drug product are provided in Table P.5-1.

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Table P.5-1. BNT1 62b2 drug product specifications.

Quality

Atitibute

Ànalvtical Procedure

Composition and Strength

Appearance

Appearance (Visual)

Appearance

Particulates)

Subvisible

Acceptance

Ciiteria

White

to off-white suspension

Essentialiv free from visible particulates

Particies ?t0 tm:

tVtstble

Appearance

(Particles)1’

Particies

Subvis;ble Particulate Mafter

6000 per

containez

Osmolality

LNP

Size

LNP ?olvdispersity

RNA

?otentiometrv

Osmometryb

Damic Liaht

Scattering

(DIS)

flynamic Light

Scatteiing

(DLS)

Encapsulation

content

Fluorescence

f1uoresccce

assay

assav

ALC-0315 content

ÀLC-O 159 conteut

DS?C

ILC-CAD

IWLC-CAD

COntent

Cholesterol

content

Container content for

injections

1LC-CAD

HPLC-CAD

Volume

of injections iii contamerst

Particies

25 tm: 00

per

contain&

6.9

7.9

425

625 mOsmol/kg

to ISO

0,3

80%

0 50 z 0.13 mg!mL

4.50

to 9.25

mg’mL

055 to 1.20

mgmL

090 to

2.05

mg!mL

—

180 to 3.90

mamL

Not lets than the sum of the nommal

vohimes of

HPLC-CAD

doses

consistent

with references

Identitv

Lipid identities

tRetention times

(ALC-03

DS?C)

Identity

15. ALC-O1

C hotesterol.

encoded

RNA

RI-PC

Identity

confirmed

sequence

Potency

lii Vitro Expression

Puijty

RNA Integrity

Adveutitious Ageuts

Bacierial Endotoxin

Sierilitv

Container Ciosure

hiteity

Cell-based flow

cvtontry

Capillaty Gel Elecwophoresis

Endotoxm (LAI)

Stenhty5

Dye mcursiont

‘‘

30% Cds Positive

50% intaet RNA

12.5 EUmL

Growth

Detected

Pass

Ali assays perfosmed en siability unless othenvise noted

b. Compendial

c. USP 7$] (obscuration method). and ahgned with

tipcoming (Jan

2021)

revision

Eur. 2.9.19

d US? 785

also iii accor&mce with

Eur. 2.2.35. with

minor

chiference sti

instnimetit

cahbrauon

e. Assay not perfomied on

stabihiv

f Procedure is aligned with Test

for Extractable

Volume ofPareiueral

Preparanous,

Tested ar release and on

stability

for

stabihry batches

only

Abbreviatmns:

LNP

Lipid nanoparticles. CAD

charged

aerosol

drrector: RT-PCR= reverse

tianscnption

polvmeiase cham reacnon. FÀC

fluorescence activated ccl sorter.

ddPCR

droplet digital PCR. qPC R

quantltative

PCR dsRi’A

double

stranded

RNA L41.

Limulus amebocvte lysaxe; EU

endotoxin unit

Specification and justification of specifications

The specifications document for drug product in section 3.2.P.5.1 inciudes a comprehensive panel of

relevant tests along with corresponding acceptance criteria.

With the exception of osmometry, volume of injections in containers, HPLC-CAD (lipid identities) and

RT-PCR (identity of encoded RNA sequence), which are performed only at DP release, ali other

analytical procedures are conducted at release and stability studies for drug product. It is stated by the

applicant that the acceptance criteria used for stability during sheif-life will be the same as the

acceptance criteria used for bt release, but this remains to be conflrmed.

Test method numbers are missing and should be given to ali analytical procedures used in the

specifications for release and end-of-sheif-life and should consequently be inserted in the drug product

specifications document and to the descriptions and validations of analyticab procedures.

LNP size for drug product is measured by dynamic light scattering fDLS) and the efficacy of the drug

product depends on the size of the LNP. The proposed acceptance criteria of 40 to 180 nm seem wide

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compared to clinical batch data that is found in the range of 59-74 nm for the small scale clinical

batches (“classical LNP process) and 68-71 nm for the emergency supply (“upscale” LNP process). The

acceptance criteria should therefore be tightened to be in line with what has been qualified in the

clinical studies or clinically qualified by other means and set such that a clinically qualified level is

assured throughout the shelf-life of the drug product.

Potency: In-vitro expression is a cell-based flow cytometry assay. The assay was implemented recently

and the proposed acceptance criteria of 30% cells positive seem wide compared to the limited batch

release data available to date, i.e. emergency supply lots that is in the range of 63-65%. In addition,

some data are presented for the small-scale clinical batches used in comparability testing, where data

are found in the range of 50-71°7o (Table 3.2.P.2.3-5 in the dossier). The proposed acceptance criteria

need to be thoroughly justified and tightened in line with the levels qualified in clinical studies or

clinically qualified by other means. This justification should include the applicant’s total current

knowledge of the drug product.

RNA encapsulation of drug product is measured by a fluorescence assay where free and total RNA are

determined and the difference between the total and free RNA corresponds to RNA encapsulation.

Encapsulation is used to ensure delivery of the RNA and improve the chances of transfection. The

proposed acceptance criteria of 80% seem wide compared to clinical batch data that is found in the

range of

Y2-94%.

The proposed acceptance criteria for RNA encapsulation should therefore be

tightened based on clinical qualification or clinically qualified by other means and set such that a

cllnically qualified level is assured throughout the sheif-life of the drug product.

The proposed acceptance criteria of 500/o intact RNA for RNA integrity as measured by capillary gel

electrophoresis seem wide compared to cllnical batch data that is found in the range of 69-81%. The

proposed acceptance criteria for RNA integrity should therefore be tightened based on clinical

qualification or cllnically qualified by other means and set such that a clinically qualified level is assured

throughout the shelf-life of the drug product. Additionally, it should also be clarified if the emergency

lots EE8492 and EE8493, both with results for RNA integrity of

55%,

have actually been used in the

clinical trials or not. In this context, it is also unclear whether there is a decrease in RNA integrity

during the manufacturing of DP or not and a consequential need for a more stringent DS specification.

The applicant should therefore discuss, and present comparative results for DS and DP, on RNA

integrity. Sections S.4.1 and P.5.1 in the dossier should be aligned and updated accordingly. (MO)

The proposed acceptance criteria for LNP polydispersity as measured by DLS are wide and should be

tightened in line with batch results for clinical batches, i.e. NMT 0.2 (0.22 observed on stability).

The proposed acceptance criteria for appearance, subvisible particles, pH, osmolality, volume of

injection in containers, identity of encoded RNA sequence, RNA content, bacterial endotoxin, sterility

and container closure integrity are ali found acceptable.

Upid content: Both safety and efficacy are dependent on the total amount of lipid relative to the RNA

DS. A consistent molar ratio of lipid/RNA is expected in the DP vial, driven by the encapsulation

process. Absolute lipid content may vary but composition (relative molar

D/)

of the four lipids remains

consistent. The acceptance criteria ranges have been calculated from worst-case low and high RNA

content. No batches manufactured to date have exhibited results at or below the low RNA content

estimate while the high RNA content level has been justified by development batches manufactured at

worst-case high RNA contents. Although the absolute range of each lipid appears somewhat broad, the

acceptance criteria are found acceptable. However, to further strengthen the control strategy given

that a fixed molar ratio of cationic lipid and RNA is critical for LNP formation, acceptance criteria for the

molar ratio NIP should be included in the specification unless further justified.

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A separate test for in vitro release is flot inciuded in the specification. This is considered acceptable

since test for potency is inciuded by a cell-based method.

Analytical procedures

Some of the analytical procedures are common to both DS and DP. Several analytical procedures are

specific to DP and are detalied and validation resuits are presented.

The compendial methods have been verified for use in accordance to the appropriate Ph. Eur. chapters.

It is claimed that ali non-compendial methods were validated against the parameters given in ICH Q2.

However, the validation summaries presented are far too brief to be able to conciude on the suitabiiity

of the analytical method. More comprehensive validation summaries of ali non-compendial methods,

for exampie in the form of short validation repofts should be provided. The validation summaries

should inciude ali relevant caiculations, acceptance criteria, description of and resuits obtained for

individual sampies. Chromatograms and dose response curves should be inciuded, where applicabie.

The dossier should be updated accordingly.

Furthermore, in ali of the in-house analytical methods used in the release of DP, method descriptions

are based on “examples” of procedures, controls and standards as weli as on “typical” system

operating parameters. These terms raise uncertainties regarding the deveiopmental stage, and the

control of critical steps of these assays. The analytical methods used in the control of DP are expected

to be finalized. The appiicant is requested to confirm this and to update the relevant parts of the

dossier with unequivocal method descriptions and additional details, if needed. The applicant should

also confirm that any significant changes in analytical procedures will be applied for in a variation

application.

In addition, it is stated in the dossier that a compiete description of the rapid sterility test is pending.

Therefore, method description and validation summary of the rapid sterility test should be provided

during the procedure.

Potency: Cell based flow cytometry is used to confirm the in vitro expression of SARS-CoV-2 spike

protein encoded by the RNA in BNT162b2 drug product (DP). Aithough the principie and method

procedure are, at iargely described, additional details are requested on criticai reagents (such as

antibodies), drug product control sampies, equipment, assay suitability, gating strategy as well as

further justification of the use of HEK293 ceils in the assay.

Batch analysis

Batch analysis data have been provided including DP batches used in toxicology studies, clinical trials,

emergency supply and stability. Ali these batches have been manufactured with the “classical” LNP

process (nonciinical, ciinicai suppiy iots) or the “upscaie” LNP process (emergency supply) and

comparability has been reasonable demonstrated between the ciinicai supply lots and the emergency

supply bt with only small differences noted. AlI DP batches manufactured and presented met the

acceptance criteria in the DP specification. However, no DP batches at the intended full commercial

scaie have been manufactured to date.

Characterisation of impurities

The impurity profile of the DP is based on the impurity profile of the materials that are used for the

manufacturing as webb as the bipid impurities.

There are four process-related impurities identified for the DP; ethanol, citrate, HEPES and EDTA.

Removab of ethanoi will be demonstrated during process validation against the ICH Q3C iimit (5000

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ppm, class 3 soivents). EDTA, citrate and HEPES have been shown through safety risk assessment and

theoretical worst-case caiculations to be significantly below established safety limits. This is found

acceptable.

The lipids are controlled via the acceptance criteria in their specifications. However, no information is

provided on the lipid-related impurities originating from the degradation of the lipid nanoparticies and

such data needs to be provided.

The applicant plans to update the dossier with further evaluations of lipid-related impurities and states

that for section 3.2.P.5.5 “Data for this section is pending and will be updated once the data has been

generated, analysed, and verified”. Until these data are available for assessment, no final conclusions

can be drawn on section 3.2.P.5.5.

A summary ot risk assessment on elemental impurities in line with the ICH Q3D is missing. A summary

of this risk assessment based on the general principles outlined in Section 5.1 of ICH Q3D should be

submitted.

In summary, no tinal conciusion on the section 3.2.P.55 can be drawn until ali data on the

characterization of impurities will be provided for assessment.

Reference standard

The current reference standard for the BNT162b2 drug product is the clinical batch EE8493, stability

data is being acquired. The applicant intends to establish a primary (PRS) and a working reference

standard (WRS). A question is raised on the preparation, qualification and stability ot PRS and WRS.

Stability of the product

The proposed initial sheif-life for drug product is 6 months when stored at the recommended storage

condition of -90 to -60°C.

The applicant has provided stability results up to 4 months at -80 to -60°C of one clinical batch and up

to 3 months of a non-clinical batch of drug product. Additionally, up to 3 months resuits at -80 to

60°C are also provided for suppoftive stability studies for two clinical lots of drug product.

The applicant has also initiated stability studies on two emergency supply lots (only release data exists

to date) and has plans to initiate stability studies on the future PPQ-batches.

In addition, stability data has also been provided at accelerated (-40°C to +5°C) and stressed (+25°C

to +30°C) storage conditions.

The stability studies are performed in accordance with ICH QSC (Quality of biotechnological products:

Stability testing of biotechnologicai/biological products) and the same or representative container

ciosure system are used in these stability studies as will be used for commercial batches.

Data is presented in P.2.5 for the container closure inciude extractables and leachabels, container

integrity, and for functional tests for the bromobutyl stopper (penetrability, fragmentation, and seif

sealing). A question is raised regarding the self-sealing test for the bromobutyl stopper atter freezing

and thawing.

Ali stability resuits for the cilnical and non-clinical batches as well as for the supportive stability studies

stored at -80 to -60°C complies with the clinical acceptance criteria in place at the time ot testing.

Overall, the presented stability data indicate no signs of degradation, significant trends or changes in

terms ot quality.

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At accelerated conditions of +5°C-storage and up to 4 months testing of a clinical batch of drug

product, LNP polydispersity and RNA integrity were out of specification at the 3 and 4 month-points.

As discussed, and conciuded in section 3.2.P.2.3, it is agreed that comparability has been reasonable

demonstrated between the clinical supply lots manufactured with the “ciassical” LNP process and the

representative emergency supply bt manufactured with the “upscale” LNP process. However, the

applicant has a plan for a comprehensive demonstration of comparability among clinical supplies and

the fuli commercial scale product but data for this section is pending. Four commercial PPQ-batches

will be manufactured in November and December 2020. In summary, no final conciusion on

comparability can be drawn until ali comparability data among clinical supplies and the commercial

product (PPQ-batches) of drug product will be provided for assessment. In addition, the claimed shelf

life is not yet acceptable since the batches are flot representative of commercial supply (manufacturer,

scale, drug substance process), the batches used represent less than

of the commercial scale, and

only very limited data is available.

Photostability testing as well as temperature cycling studies are planned, and results are pending to

date. While normally this data should be provided before the end of the RR procedure, it is

acknowledged that the outer container (carton box) will provide protection from light; this information

should be clearly stated in the SmPC/PIL.

Furthermore, it should be confirmed that future extensions of the assigned DP shelf life will be applied

for in formal variation applications. The following statement should be removed for Module 3.2.P.8.1 of

the dossier; “The sponsor will extend the assigned shelf life without notification providing the real time

stability data at the intended storage condition is acceptable and within commercial specifications.”

Post-appro vat stabil/ty protocol and stabil/ty commitment

A minimum of one batch of drug product will be added to the on-going post-approval stability program

annually. The annual post-approval stability protocol has been provided and found acceptable although

this protocol is part of GMP and therefore flot assessed in this report. However, the applicant should

confirm that they commit to continue alI the ongoing stability studies at long-term conditions until

completion.

Conciuding remarks on the proposed shelf-life and storage conditions

The proposed initial shelf-life for the drug product is 6 months at the recommended storage

temperature of -90 to -60°C. In order to support the suggested shelf-life for drug product, updated

reports from the ongoing stability studies should be provided.

Post approval change management protocol(s)

Not applicable.

Adventitious agents

Adventitious agents safety evaluation has been provided for the DS manufacturing site [Andover] and

for the DP manufacturing site [Puurs]. Information regarding the DP manufacturing site [ENT

&Rentschler] is pending.

Proteinase K used in DS manufacturing and LE broth used in the establishment of the pST4-1525 MCB

and WCB are the only materials of animal origin used in the manufacturing of ENT162b2. The applicant

has identified contamination of the product by Transmissible Spongiform Encephabopathy (TSE) agents

as the main theoretical risk associated with these ingredients, deemed minimal.

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No information is provided regarding viral safety of these materials. Considering the stringent

conditions routinely used in the heparin production, the risk for viral contamination is considered

negligible for this material. Additional ciarifications are requested for pyrophosphatase, T7 polymerase

and RNase inhibitor, spermidine, DNase I and excipients ALC-0315, ALC-O 159, DSPC and Cholesterol.

No information is inciuded in A.2 on the control of other non-viral adventitious agents and only sterility

testing performed at the level of DP is named. However, sufficient details on the aseptic validation

filling and media fills have been provided in P.3 Manufacture. Furthermore, adequate testing for

bioburden and endotoxin is performed at different stages of the manufacturing process, as described in

section S.2.4. Therefore, based on the information existing in other parts of the dossier and pending

new information regarding the BNT & Rentschler manufacturing site as well as new information

requested on the control of materials, the overall risk for contamination is considered minimal at this

point and no additional concerns are raised.

GMO

N/A

Novel excipients

Two novel excipients are inciuded in the drug product, the cationic lipid ALC-0315 the PEGylated lipid

ALC-0159. No f9nal conclusion can be drawn until alI data are provided. Some questions with regards to

batch size and validation of analytical methods are raised at this point. Additional information on

chemical synthesis, quality control of starting material, specification limits and retest period will be

provided for assessment during the procedure.

2.1.4. Discussion and conciusions on chemical, pharmaceutical and

biological aspects

Drug substance

Where data is submitted, the dossier is overall of acceptable quality. However, a substantial amount of

information is pending, due to the very short time frame of product development and will be submitted

in the subsequent submission(s). Information on the manufacturing process and process controls for

the manufacturing site Andover is provided, while the corresponding information for site BNT Mainz &

Rentschler is pending.

Based on the significant differences observed between batches manufactured by DS Process 1 and 2

for the CQA mRNA integrity, a MO is raised regarding comparability, characterisation and clinical

qualification of the proposed acceptance criteria of 50% intact RNA. Whilst some testing results of

biological activity/functionality has been submitted in support of comparability and potency testing is

part of the DP release specifications, biological characterisation of the active substance is limited, and

additional data and discussion is requested to address functionality.

The reference standard was poorly characterised, and the final two-tiered system is not yet in place.

The proposed initial shelf-life for the drug substance is 6 months at the recommended storage

temperature of -20°C. In order to support the proposed shelf-life for drug product, updated reports

from the ongoing stability studies should be provided.

CQVID-19 mRNA vaccine (nucleoside modified)

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Drug product

The drug product is a preservative-free, multi-dase concentrate to be diluted for intramuscular

injection, intended for 5 doses. The drug praduct is a sterile dispersion of RNA-containing lipid

nanoparticies (LNP5) in aqueous cryoprotectant buffer.

The formulation development studies of the RNA containing lipid nanoparticles have been thoroughly

described including studies that were performed with available drug substance, representative of the

mRNA platform and inciuded in the drug product.

The development of the manufacturing process is extensively described, and critical process

parameters are defined.

The manufacturing process includes lipid nanoparticle fabrication and bulk drug product formulation

followed by fill and finish, and the process has at large been acceptably described.

however, no drug product batches at the intended full commercial scale have been manufactured to

date. It is described in the dossier that four commercial PPQ-batches will be manufactured in

November and December 2020. These batches will be used both to demonstrate the comparability of

the commercial PPQ-batches versus the clinical supply batches as well as for process validation of the

manufacturing process of the drug product. Therefore, no final conciusion on drug product

comparability, process validation, and shelf life can be drawn until additional data will be

provided for assessment.

The specifications document for drug product includes a comprehensive panel of relevant tests along

with corresponding acceptance criteria. Several questions are raised concerning tightening of

acceptance criteria for LNP size, polydispersity, potency, RNA integrity and RNA encapsulation to be in

line with what has been qualified in the clinical studies or clinically qualified by other means.

The proposed initial shelt-life for the drug product is 6 months at the recommended storage

temperature of -90 to -60°C. In order to support the suggested sheif-life for drug product, updated

reports from the ongoing stability studies should be provided.

Conciusion

Three major objections are identified that would preclude a marketing authorisation: The first MO

relates to the GMP status of the DS and DP manufacturing sites. Comparability between clinical and

commercial material has nat yet been demonstrated, which is addressed in MO 2. In particular,

significant differences between batches manufactured by DS Process i and 2 are observed for the CQA

mRNA integrity. Characterisation of truncated forms, more comprehensive comparability data, resuits

an additional batches and impact an safety and efficacy is requested. The third MO concerns omission

of data an DP manufactured at the commercial site. Batch results at release, data an comparability af

commercial batches with clinical batches and additional stability data is required.

In addition, several deficiencies have been noted which should be appropriately addressed by the

applicant before a positive CHMP opinion can be granted.

2.2. Non-cilnical aspects

2.2.1. Pharmacology

The pharmacology dossier is based an initial studies of the functionality of the BNT162b2 (V9) RNA

based product and the encoded SARS-CoV-2 P2 5 protein as well as on supporting studies of SARS

CoV-2 P2 S protein structure. This is followed by characterization of the humoral and cellular immune

COVID-19 mRNA vaccine (nucleoside modified)

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response in mouse and nonhuman primate upon immunization with BNT162b2 (V9) and ends up with a

SARS-CoV-2 challenge study of BNT162b2 (V9) immunized nonhuman primates. No secondary

pharmacodynamic, safety pharmacology or pharmacodynamic drug interaction studies with BNT162b2

have been conducted due to the nature of the RNA-based vaccine product, which is accotding to

applicable guidelines.

Mechanism of action

The SARS-Cov-2 virus infect the body by the use of the Spike protein (S) to attach to specific ceil

surface receptors, especially, as recently suggested, the angiotensin converting enzyme 2 (ACE2). In

addition to the initial attachment to a host celi, the S protein is aiso responsible for vital envelope

fusion with the host celi membrane resulting in genome release. Due to its indispensabie role, the S

protein is a major target of virus neutralizing antibodies and has become a key antigen for vaccine

development. By immunization with BNT62b2, encoding for the S protein, the intention is to trigger a

strong and relatively long-lasting production of high affinity virus neutralizing antibodies, which can act

through blocking the S-protein and it’s receptor-binding domain (RBD) interaction with host cell

receptors but also by opsonization mediated virus clearance. In addition, the immunization with

BNT162b2 is also intended to elicit a concomitant T celi response of the Thi type, supporting the B

celis responsibie for the production of S-specific antibodies and cytotoxic T celis that kiil virus infected

celis.

The structurai eiements of the vector backbones of the BNT162b2 are optimized for prolonged and

strong translation of the antigen-encoding RNA. The potency of the RNA vaccine is further optimized by

encapsulation of the RNA into lipid nano particies (LNPs), which protects the RNA from degradation by

RNAses and enable transfection of host ceils after intramuscular (i.m.) delivery. BNT162b2 is

nucleoside-modified by a substitution of i-methyl-pseudouridine for uridine and thus its inherent

adjuvant activity mediated by binding to innate immune sensors such as toli-iike receptors (TLR5) 7

and 8, is dampened, but not abrogated.

The 5 protein is a trimeric class I fusion protein that exists in a metastable prefusion conformation

before engaging with a target ceil. BNT162b2 encodes a P2 mutant (P2 5) variant of 5 where two

consecutive proline mutations have been introduced in ordet to iock the RBD in the prefusion

conformation.

The RNA is formulated with functionai and structural iipids forming lipid nano particies (LNPs), which

protect the RNA from degradation and enabie transfection of the RNA into host celis after IM injection.

The composition of the LNP5 may aiso affect the distribution of injected ENTi62b2. In addition, it

cannot be exciuded the LNP composition contributes to the overall immunogenicity.

Primary oharmacodynamic studies in vitro

To confirm the functionality of the BNTi62b2 (VO) RNA-based product, protein expression, transfection

frequency from BNT162b2 and ceil surface expression of the SARS-CoV-2 P2 5 protein antigen was

assessed. Regarding the resuits obtained from the Western Blot, a semi quantitative anaiysis of the

results shouid be provided to improve the readabiiity of the protein expression and in the anaiysis of

the blot, some missing scientific information and expianations should be added by the applicant (OC).

BNTi62b2 (VO) transfection of HEK293T cells indicated SARS-C0V-2 P2 5 was correctiy expressed on

the ceil surface, as indicated by flow cytometry staining of non-permeabiiized ceils with an anti-Si

monocional antibody. In addition, the ceiluiar iocalization of expressed Sl protein was investigated.

The 5 protein co-localized with an ER marker, as detected by immunofluorescence experiments in

HEK293T celis expressing BNT162b2-RNA, suggesting the 5 protein is processed within the ER.

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In a set of supportive studies, it was investigated whether BNT162b2 RNA encodes for an amino acid

sequence that authentically express the ACE2 binding site. No study report for this data set could be

found and should be provided (OC). Recombinant P2 S was expressed from DNA encoding for the

same amino acid sequence as BNT162b2 RNA encodes for. Flow cytometry staining with spike protein

(S) binding agents, as human ACE2 and monocional antibodies known to bind to authentic S-protein ell

indicated an authentically presented P2 5 protein and ACE2 binding site. Low nano molar affinity of P2

5 binding to ACE2 PD and B38 mAb was demonstrated with the use of biolayer Interferometry.

To further structurally characterize the P2 spike protein, a cryo-electron microscopy (cryoEM)

investigation of purified P2 S, expressed from DNA, was conducted. The cryoEM revealed, according to

the Applicant, a particie population ciosely resembling the prefusion conformation of SARS-C0V-2 spike

protein. By fitting a previously published atomic model on to a processed and refined cryoEM dataset,

a rebuilt model was obtained showing good agreement with reported structures of prefusion fulI-Iength

wild type 5 and its ectodomain with P2 mutations. In the prefusion state the RED undergo hinge-like

conformational movements and can either be in an “up” position (open for receptor binding) or in a

“down” position (closed for receptor binding). Three-dimensional classification of the dataset showed a

class of particles that was in the conformation one RED ‘up’ and two RED ‘down”. This paftly open

conformation represented 20.4% of the trimeric molecules. The remainder were in the ali RED ‘down’

conformation. Although potent neutralizing epitopes have been described when the RED is in the

“heads down” ciosed conformation, the “heads up” receptor accessible conformation exposes a

potentially greater breadth of neutralizing antibody targets. It is conciuded that antibodies to both the

up and down conformations will potentially be formed upon immunization with the P2 5 encoding

ENT162b2. Regarding the Structural and Eiophysical Characterization, the applicant is asked to provide

a) A schematic description of the V8 and V9 variants,

as to identify the exact position of optimized

codons in the sequence, as well as the position of added cytosines nucleotides. The exact position of

these optimized codons inside the moURNA sequence should be provided. b) The exactly detailed

mRNA structure of ENT162b2, including coding and non-coding sequences. c) A comparison in the VS

and V9 codon sequences, highlighting their differences and mlJU residues. d) An estimation of mIPU

content in both VS and V9 sequences and discuss on the potential difference in immunogenicity

between these two sequences. E) A comparison on the protein expression obtain from both variants

(V8 and V9) to ensure that the expected protein is expressed in non-clinical models (OC).

Primary pharmacodynamic studies in vivo

The humoral and cellular immune response following IM administration of ENT162b2 (V9) was

investigated in mice and nonhuman primates.

Balb/c, females were immunized IM on day 0 with 0.2, 1 or 5 pg RNA/animal of BNT162b2 (V9), or

with buffer alone (n=8). Blood samples were collected on Days 7, 14, 21 and 28 efter immunization.

The IgG antibody response to SARS-CoV2- RED or Sl was analyzed by ELISA. Immunization with

BNT162b2 induced IgGs that bound to Sl and RED, as detected by ELISA, and on day 28 after

immunization showed a binding affinity of KD 12 nM or 0.99 nM (geometric mean) respectively, as

detected by surface plasmon resonance.

To further characterize the antibody response to ENT162b2 and its potential capacity to reduce SARS

Cov-2 infections, a pseudo virustype neutralization assay (pVNT) was used as a surrogate of virus

neutralization since studies with authentic SARS-CoV-2 requires a BSL3 containment. The pyNT was

based on a recombinant replication-deficient vesicular stomatitis virus (VSV) vector that had been

pseudotyped with SARS-CoV-2 5 protein according to published protocols. A dose-dependent increases

in SARS-CoV-2-S VSV pseudovirus neutralizing antibodies were observed in sera from BNT162b2-

immunized mice. On day 14, the difference of the group treated with 5 pg RNA compared to the buffer

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control was statistically signif9cant (p

0.0010). On days 21 and 28, the differences of the groups

treated with i ig and 5 pg BNT162b2 compared to the buffer control were statistically significant. The

relevance of the pseudovirus assay for authentic SARS-Cov-2 was not discussed. Concerning study R

20-0085 on the immunogenicity of the LNP formulated modRNA encoding the viral 5 protein (V9), the

applicant is asked to a) justify the absence of IgG2A and IgGi characterization for RBD; b) justify why

the resuits were flot expressed in titers that would also allowed comparisons across experiments.

Indeed, comparison with pyNT experiments expressing resuits in titers could help to determine the

levels of neutralizing and non-neutralizing antibodies present in the sera (OC). Immunization of mice

with BNT162b2 also induced IFN-y secreting celis of both the CD4+ and CD8+ T-celI subsets. This was

shown by ELISPOT atter ex vivo re-stimulation of spienocytes with an S-protein overlapping peptide

pool Day 28 after immunization. Cytokine profiling was also carried out by Multiplex analysis of

cytokine release from the Day 28 Spienocytes. High levels of the Thi cytokines IFNy and IL-2 but

minute amounts of the Th2 cytokines IL-4, IL-5 and IL-13 were detected after re-stimulation with S

but flot RBD overlapping peptide mix. In addition, an elevated secretion ofTNFa, GM-CSF, IL-113, IL

12p7O and IL-18 was recorded after re-stimulation. In order to characterize the immunophenotype of

B-and T-cells appearing in lymph nodes from mice immunized with BNT162b2 fV9), B- and T-cell

subsets in draining Iymph node ceils were quantified by flow cytometry 12 days after immunization.

Higher numbers of B cells were observed in the samples from mice that received BNT162b2 compared

to controls. That included plasma cells, class switched IgGi- and IgG2a-positive B ceils, and germinal

centre B celis. T-cell counts were elevated, particularly numbers ofT follicular helper (Tfh) cells,

inciuding subsets with ICOS upregulation, which play an essential role in the formation of germinal

centres (Hutloft 2015).

In the nonhuman primate (rhesus macaques) studies, 8NT162b2 (V9) was shown to be immunogenic

atter intramuscular administration. The serum concentrations of both Sl-binding and the SARS-CoV-2

neutralizing antibody titers were at least an order of magnitude higher atter BNT162b2 immunization

of rhesus macaques than for the panel of SARS-CoV-2 convalescent human sera. Regarding Study VR

VTR-10671 BNT162b2 (V9) Immunogenicity and Evaluation of Protection against SARS-CoV-2

Challenge in Rhesus Macaques, the applicant needs to a) precise for the Luminex data how the

reference curve for has been constructed, what does represent the arbitrary U/ml used and how it is

referring to the serum dilution factor; b) define the criteria for choosing a 10-30% infection rate of

Vero ceils; c) Methods to quantify antibody production in the different experiments dufter and

consequently cross-comparison between experiments is hardly impossible. Indeed, it is impoftant to

distinguish neutralizing antibodies from non-neutralizing antibodies. In this study, total antibody

response is measured using a luminex assay and resuits expressed on U/ml and for the neutralization

assay results are expressed in VNT 50. The applicant needs to provide an estimation of the non

neutralizing antibodies in the whole antibody response. d) Tt is important to notice that on figure 6 of

study report, neither panel A nor panel B highlight the consumption of IgG Sl binding antibodies after

challenge nor the increase due to B memories response following the challenge: this would need to be

further justified by the Applicant (OC).

Antigen specific S-reactive T-cell response after BNTi62b2 immunization of the macaques was

measured by ELISPOT and ICS. While S-specific T cells were low to undetectable in naïve animals,

strong IFNy but minimal IL-4 ELISpot responses were detected after the second 30 or 100 ig dose of

the 8NT162b2. Intra cellular staining (ICS) contirmed that BNT162b2 immunization elicited strong S

specific IFNy producing T cell responses, including a higher frequency ot CD4+ T cells that produced

IFNy, IL-2, or TNF-a but a lower frequency of CD4+ celis that produce IL-4. An S-specific IFNy

producing CD8+ T cell response was also recorded.

COvID-19 mRNA vaccine fnucleoside modified)

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A challenge study in rhesus macaques served as nonclinical proof of concept (PoC). Rhesus macaques

share a 100% homology with the human ACE2 sequence that interacts with the RBD of the S protein.

BNT162b2 (V9) immunized macaques were challenged with SARS-CoV-2 intra nasally and intra

tracheally

days after the second immunization with BNT62b2. Rhesus macaques were immunized

on days 0 and 21. Some other covid-19 vaccine candidates have different prime-boost intervals, such

as 4 weeks for both ChAdOxi (Graham et al., 2020) and mRNA-1273 (Corbett et al., 2020).

Considering that the time between the first and second vaccine dose may have a significant impact on

the immunological response, the applicant is asked to provide the rationale for the chosen prime-boost

interval (21 days) (OC). At the time of challenge, SARS-CoV-2 neutralizing titers ranged from 260 to

1,004 in the BNT162b2 (V9)-immunized animals. Neutralizing titers were undetectable in animals from

the control-immunized and sentinel groups. The presence of SARS-CoV-2 RNA was monitored by nasal

and oropharyngeal (OP) swabs and bronchoalveolar lavage (BAL). Vital RNA was detected in BAL fluid

from 2 of the 3 control-immunized macaques on Day 3 after challenge and from 1 of 3, on Day 6. At

no time point sampled was vital RNA detected in BAL fluid from the BNT162b2 (V9)-immunized and

SARS-CoV-2 challenged macaques. The difference in vital RNA detection in BAL fluid between

BNT162b2-immunized and conttol-immunized rhesus macaques aftet challenge is statistically

significant (p=O.0014). From control-immunized macaques, viral RNA was detected in nasal swabs

obtained on Days 1, 3, and 6 after SARS-CoV-2 challenge; from BNT162b2 (V9)-immunized macaques,

viral RNA was detected only in nasal swabs obtained on Day 1 after challenge and flot in swabs

obtained on Day 3 ot subsequently. The pattern of viral RNA detection from OP swabs was similar to

that for nasal swabs. No signs of viral RNA detected vaccine-elicited disease enhancement were

observed. The vital RNA levels between control-immunized and BNT162b2-immunized animals after

challenge were compared by a non-parametric aflalysis (Friedman’s test), and the p-values ate 0.0014

for BAL fluid, 0.2622 for nasal swabs, and 0.0007 for OP swabs. The data from the individual animals

should be provided for the RT-qPCR test for presence of SARS-C0V-2 RNA (OC).

Despite the presence of vital RNA in BAL fluid from challenged control animals, none of the challenged

animals, immunized or control, showed clinical signs of illness (weight change, body temperatute

change, blood oxygen saturation and heatt rate). The Applicant concluded, the absence of cllnical signs

in any of the challenged animals, immunized or control, despite the presence of vital RNA in BAL fluid

from challenged control animals, indicates that the 2-4 year old male rhesus monkey challenge model

appears to be an infection model, but flot a clinical disease model. However, a further investigation by

lufig radiograph and computerized tomography (CI) was conducted. Radiographic evidence of

pulmonary abnormality was observed in chailenged controls but flot in unchallenged sentinels flot in

challenged BNT162b2-immunized animals except for a CT-score signal in i of 6 pre infection and 2 out

of six at Day 10/EOP in BNT162b immunized animals. The CT score signal was at the same level as the

control at Day 10/EOP and is of unclear significance due to the presence in one animal before

chailenge. No radiographic evidence of vaccine-elicited enhanced disease was observed.

Histopathological examination of lung tissues is ongoing and will be submitted as an addendum (OC).

Overall, the challenge study is suboptimal as it comes with a number of uncertainties. The limitations

can be listed regarding the model: absence of sars-cov2-clinical signs in control and challenged NHP,

use ofjuveniles NHP, lack offemales NHP, one out of three age-matched salme control-immunized

(n=3) male rhesus macaques not responding to challenge (no viral RNA neither in the BAL and nasal

swab), low numbers of animals with a low statistical sigflificance, questionable selection of titer of the

viral challenge (1.05.

106

PFU). Moteover, some important data are missing to date like the absence of

cytokines measurement in the NHP BAL. The applicant is asked to discuss ali these limitations and

should provide further scientific information on the NHP model relevance. Although the model is

considered adequate to demonstrate immuflogenicity, and vital clearance, it is considered insufficient

to demonstrate efficacy against the disease (OC).

COVID-19 mRNA vaccine fnucleoside modified)

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Secondary yharmacodynamic studies

No secondary pharmacodynamics studies were conducted with BNT162b2. This is accepted.

Safety pharmacology programme

No safety pharmacology studies were conducted with BNT162b2. The Applicant refers to that they are

flot considered necessary according to the WHO guideline (WHO, 2005). Ifl addition, no findings on

vital organ functions have been recorded in the repeat dose toxicology studies. Thus, the absence at

safety pharmacology studies is accepted.

Pharmacodynamic drug interactions

No pharmacodynamics drug interaction studies were conducted with SNT162b2. This is accepted.

2.2.2. Pharmacokinetics

The applicant has determined the pharmacokinetics of the two novel LNP excipients ALC-0315

(aminolipid) and ALC-0159 (PEG-lipid) in plasma and iver as well as their elimination and metabolism

in rats. Furthermore, the applicant has studied the biodistribution at the two novel lipids

rats) and a

LNP-formulated surrogate luciferase RNA in mice. No traditional pharmacokinetic ar biodistribution

studies have been performed with the vaccine candidate BNT162b2.

No validated methods of analysis to suppoft the non-clinical PK/biodistribution studies have been

submitted. However, the applicant claims to have used a qualified LC-MS/MS method to support

quantitation of the two novel LNP excipients without providing such data

(OC).

PK studies with the two novel LNP-excip!ents ALC-0315 and ALC-0159: Wistar Han rats were IV bolus

injected with LNP formulated luciferase-encoding RNA at 1 mg/kg and ALC-0315 and ALC-0159

concentrations at 15,3 mg/kg and 1,96 mg/kg respectively. ALC-0315 and ALC-0 159 levels in plasma,

iver, urine and faeces were analysed by LC-MS/MS at different time-points up to 2-weeks. No other

organs besides the hver were investigated and therefore distribution to other organs cannot be

excluded. The clinical administration route is IM the PK study was performed with a different

administration route (IV)

(OC).

ALC-0315 and ALC-0159 were rapidly cleared from plasma during the first 24 hours with an initial

t1/2

of 1.62 and 1.72 h, respectively. 24 hours post-dosing, Iess than

of the maximum plasma

concentrations remained. A slower clearance rate was observed atter 24 hours with ALC-0315 and

ALC-0159 terminal elimination

t1/2

at 139 and 72.7 h, respectively.

Following plasma clearance, the hver appears to be to major organ to which ALC-0315 and ALC-0 159

distribute. The applicant has estimated the percent of dase distributed to the hver to be 6O% for ALC

0315 and

r..20°/o

for ALC-0159. The observed livet distribution is consistent with the observations from

the biodistribution study and the repeat-dose toxicology, both using IM administration.

For ALC-0315 (aminolipid), the maximum detected concentration in the livet (294 pg/g hver) was

teached 3 hours after IV injection. ALC-0315 was eliminated slowly from the livet and after 2-weeks

the concentration of ALC-0315 was still ‘-25% of the maximum concentration indicating that ALC-0315

would be eliminated from rat livet in approximately 6-weeks time. For ALC-0159 (PEG-hipid), the

maximum detected concentration in the hver (15.2 pg/g hver) was teached 30 minutes folhowing IV

injection. ALC-0159, was eliminated from the hver faster than ALC-0315 and atter 2-weeks the

concentration of ALC-0159 was only «0,04% at the maximum detected concentration. The applicant is

asked to comment an the differences in the kinetics at the two novel excipients as well as on the

relatively long livet chearance ofALC-0315

(OC).

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While there was no detectable excretion of either lipid in the utine, the percent of dose excreted

unchanged in faeces was

for ALC-0315 and

r5Q%

for ALC-0159.

Biodistribution ofa LNP-formulated Iuciferase surrogate reyorter: To determine the biodistribution of

the LNP-formulated

modRNA,

the applicant did flot study distribution of the modRNA used in the

vaccine candidate BNT162b2, but instead, in a non-GLP study, determined the biodistribution of a

surrogate luciferase modRNA formulated with a LNP with identical lipid composition used in BNT162b2.

Since several LNP formulations were tested in the study it is flot completely clear which of the LNP

formulation is used in the clinical version of BNT162b2 (OC).

The study used three female BALB-c mice per group and luciferase protein expression was determined

by in vivo biolumiflescence readouts using an In Vivo Imaging System (IVIS) tollowing ifljection ot the

luciferase substrate luciferine. The readouts were performed at 6h, 24h, 48h, 72h, 6d and 9d post IM

injection (intended clinical route) in the right and left hind leg with each i

lig

(total of 2pg) of LNP

formulated luciferase RNA. The biodistribution method has flot been validated or qualified and no

discussion on its sensitivity has been inciuded (OC).

In vivo luciferase expression was detected at different timepoints at the injection sites and in the hver

region indicating drainage to the livet. As expected with an mRNA product, the luciferase expression

was transient and decreased over time. Luciferase signals at the ifljection sites, most likely reflecting

distribution to the lymph nodes draining the injection sites, peaked 6h post injection with signals of

approximately 10 000 times of buffer control animals. The signal decreased slowly during the first 72

hours and atter 6 and 9 days the signals were further weakened to approximately levels of 18 and 7

times the signals obtained from animals injected with buffer control.

The signals from the hver region peaked 6h post injection and decreased to background levels 48h

after injection. The livet expression is also supportive of the data from the rat PK study and the

findings in the rat repeat-dose toxicological study showing reversible livet vacuolation and increased

gGGT levels.

Immunogenicity ofa LNP formulated luciferase modRNA: Activation of the innate immune system

following IM injection of a LNP-formuhated luciferase reporter RNA into mice was assessed in a

Luminex-based multiplex assay were serum samples (day -i (pte), 6 h and day 9) were tested for

levels of the following chemokines and cytokines: MCP-1, MIP-ip, TNF-a, IFN-a, IFN- y, IL-2, 11-6, IL

10, ILi-p, lP-lO. The apphicant tested 3 different LNP5, ali formulated together with luciferase RNA.

The results suggest that the LNP formulation used

BNT162b2 (LNP8) shightly increased levels of

MCP-1, IL-6, and lP-lO at 6h post immunisation. AlI chemokine/cytokine levels dropped to background

levels at day 9. The applicant is asked to charify issues regarding the data and discuss the possible

clinical relevance of the transiently increased IL-6 levels (OC). In addition to innate immune

activation, LNP formulated luciferase modRNA was able to induce IFN-y T-cell responses (when

chahlenged with MHC 1-specific luciferase peptide pools) measured in splenocytes isolated from the

mice at day 9.The LNP formulated luciferase modRNA did flot induce the formation of luciferase-specific

IgGs as measured by ELISA.

Metabolism of the two novel LNP-exciDients ALC-0315 and ALC-0159: Meta bolism studies were

conducted to evaluate the two novel lipids in the LNP, ALC-0315 (aminolipid) and ALC-0159 (PEG

lipid). No metabolic studies were performed with the moURNA or the other two lipids of the LNP.

Overall, it seems as both ALC-0159 and ALC-0315 are metabolised by hydrolytic metabolism of the

amide or ester functionalities, respectively, and this hydrolytic metabolism is observed across the

species evaluated.

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The metabolism of the novel excipients, ALC-0159 and ALC-0315, were examined in

vitro

using blood,

livet S9 fractions and hepatocytes, ali from mouse, rat, monkey and human. The in vivo metaboiism

was examined in rat plasma, urine, faeces, and livet from a rat pharmacokinetics study where a

luciferase-encoding modRNA formulated in an LNP was used.

Metabolism of ALC-0315 appears to occur via two sequential ester hydrolysis teactions, flrst yielding

the monoester metabolite foliowed by the doubly de-esterified metabolite. The monoester metaboiite

was observed in vitro in rat blood, monkey S9 fraction, and in vivo in rat plasma and rat livet. The

doubly de-estetified metabolite was observed in vitto in mouse and rat blood; monkey livet 59

fraction; and in vivo in rat plasma, urine, faeces and livet. Subsequent metabolism of the doubiy de

estetified metabolite resulted in a glucuronide metaboiite which was observed in urine only from the

rat pharmacokinetics study. Additionaily, 6-hexyidecanoic acid, the acid ptoduct of both hydrolysis

reactions of ALC-0315, was identified in vitro in mouse and rat blood; mouse, rat, monkey and human

hepatocytes; mouse, rat and human livet 59 fractions; and in vivo in rat plasma.

ALC-0315 was stable over 120 min (>93% remaining) in livet microsomes and S9 fractions and over

240 min (>93% remaining) in hepatocytes in ali species and test systems.

The primary route of metabolism for ALC-0159 appears to involve amide bond hydrolysis yielding

N,N

ditetradecylamine. This metabolite was identified in mouse and rat blood as weli as hepatocytes and

livet S9 from mouse, rat, monkey and human. Theoretical metabolites were arrived at via examination

of the excipient molecules and consideration of commonly observed biotransformations (hydroxylation,

N-dealkylation, hydtoiysis, glucuronidation, sulfation, oxidation and combinations thereoO. Given that

the acetamines have been repofted to be carcinogenic in animals, iflcluding livet tumors, potentially

based on genotoxic mechanism, the applicant is asked to provide a discussion on the distribution and

metabolism of the ALC-0159 focusing on the acetamide moiety (OC raised in toxicology section).

ALC-0159 was stable over 120 min (>82% remaining) in livet microsomes and 59 fractions and over

240 min (>87% remaining) in hepatocytes in ali species and test systems.

Excretion of the two novel LNP-excipients ALC-0315 and ALC-0159: Excretion ot the two novel lipids in

the LNP, ALC-0315 (aminolipid) and ALC-0159 (PEG-lipid) was studied in the rat PK study. No

excretion studies were performed with the modRNA ot the other two lipids of the LNP which is

considered acceptable.

While thete was no detectable exctetion of either lipid in the urine, the petcent of dose excteted

unchanged in faeces was «4% for ALC-0315 and ‘-50% for ALC-0159. Since almost no unchanged

ALC-3015 was detected in utine or faeces, metabolism may play a bigger role in the elimination of

ALC-0315 than ALC-0159.

2.2.3. Toxicology

The toxicological dossier for BNT162b2 is based on a total of three pivotal toxicological experimental

studies; two tepeat-dose toxicity rat studies (one full study submitted, one intermittent study

submitted) and one DART/EFD rat study (flot yet submitted beyond a study plan). The test substance

is BNT162b2 (variant 8 and the clinically relevant variant 9), a modified RNA in a lipid nanoparticle

(LNP) fotmulation. The differences between the variants are due to codon optimization. The LNP

contains four excipients wheteof two are novel (ALC-0315 and ALC-0159).

General toxicity: The two general/tepeat-dose toxicity studies involved i.m. exposure of Han Wistar

tats to BNT62b2 for a total of 17d (three exposures; lggr/w) followed by three weeks of recovery.

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One study used variant 8 of BNT162b2 (dose lOOug) and one study used variant 9 (3Oug). Overall, the

study designs only inciuded a single experimental group each with a variant of 3NT162b2, with no

dose-response assessment or specific experimental groups for the LNP alone or its novel excipients.

This somewhat limits the risk assessment but is acceptable. No test substance-linked mortality or

clinical signs were observed (except a slight increase

[<1C]

in body temperature). No ophthalmological

and auditory effects were found. The animal model of choice, the rat, has flot been assessed in the

pharmacological dossier but a limited absorption/distribution study has been conducted in

pharmacokinetics dossier. Immunogenicity was assessed in the toxicology studies.

Body weight and food intake: Exposure generated a slight reduction of absolute 8W within 24h after

ist exposure (-6.8% to -11.3%; BNT162b2 V8) alternatively a weak body weight increase reduction

[8NT162b2 v9]. No changes in food intake were observed.

Gross pathology and organ weights: At lOOug BNT162b2 V8 and 3Oug BNT162b2 V9, the tissue at the

injection site was thickened/enlarged with oedema and etythema at the end of exposure in a reversible

manner. The spleen was enlarged (reversible) with up to 60% for both vaccine variants and doses.

There was also an enlargement of the lymph nodes at lOOug. Overall, there were signs of a significant

immune response which is likely linked to and expected to a certain degree for the test substance.

There was a trend of slightly enlarged livet

ii,

females at lOOug (BNT162b2 V8)

—

Histopathology: At lOOug BNT162b2 V8, there were observations of various infiammatoty signs at the

injection site (e.g. fibrosis, myofiber degenetation, oedema, subcutis hypetplasia). Also, there was

infiammation of the perineural tissue of the sciatic nerve and surrounding bone in most rats at d17.

The bone marrow demonstrated incteased cellularity and the Iymph nodes showed plasmacytosis,

infiammation and incteased cellularity. The spleen demonstrated increased haematopolesis in half the

animals at d17. The livet showed hepatocellulat and periportal vacuolation at d17 (partially or fully

reversed ducing recovery) which may be related to hepatic cleatance of the PEGylated lipid in the LNP.

Only some organ and tissue samples from the main otgans wete used for BNT162b2 V8 histopathology

(adrenal gland, brain, epididymis, heart, kidney, livet, lungs, lymph nodes, ovary, pituitaty gland,

prostate, spleen, testicle, thymus, thyroid). Other tissues/structutes (nasal body cavity, ciltorial gland,

dorsal root ganglion, larynx, mandibular lymph node, tibial netve, preputial gland, ureter, Zymbal’s

gland) were preserved for additional histopathology if needed. The interim data for 3Oug ENT162b2 V9

did not inciude histopathology data and no specification of what tissues ate to be examined/stoted.

Immunogenicity: Treatment of rats with lOOug BNT162b2 V8 genetated SARS-C0V-2 S-binding IgG

antibodies against the Sl fragment and the RED (based on ELISA and pseudovirus neutralization test

on blood samples).

Haematology; At 3Oug ENT162b2 V9 and lOOug BNT162b2 V8, thete was a moderate to strong

teduction of reticulocytes (48-74°h, not specified for V9) coupled to lowered red cell mass parametets

(REC, HGB, and HCT). There was a very strong increase (>100%) in large unclassified cells [LUC;

295.5-319.5°h for V8, not specified for V9], neutrophils [606-680°h for V8, not specified for V9],

eosinophils [419-509% for V8, not specified for V9], basophils [105-147°/o for V8, not specified for V9]

and fibrinogen [160-205% for V8, flot specified for V9]. The changes were reversible (assessed for

V8). No effects on coagulation were observed for a V$ and a slight inctease in mainly males with V9.

CIinicaI pathology: A very strong but reversible increase (>100%) in pro-infiammatory acute phase

proteins in the blood (A1AGP, A2M) was seen with both 3Oug BNT162b2 V9 and lOOug ENT162b2 V8.

Also, indicative of pro-infiammation, a slight to moderate teduced albumin/globulin ratio was seen for

both variants. V8 (lODug) exposure generated increased levels of gGT (>200%) and increased gGT

enzyme activity and incteased AST levels (+

n49%).

V9 (3Oug) exposure led to slight to moderate

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increases in ALT and ALP levels, possible indicative of livet effects. There were no changes in cytokine

levels

(IFNg, TNFa, 11-ib, 116, 11-10)

after iOOug V8 exposure. For iOOug V8, there were no changes

measured in urine whereas there was a slight-moderate reduction in pH for 30ug V9.

Genotoxicity: No genotoxicity has been

and RNA that are flot expected to have

acetamide moiety which

classified as

genotoxic mechanism, which should be

provided. The components of the vaccine formulation are lipids

genotoxic potential. That being said, the novel lipids possess an

possible human carcinogen

(IARC

Group

2B)

with debated

discussed

further

(OC).

DART: No resuits have yet been submitted flor has a discussion been provided on choice of animal

model (rat) or experimental design

(OC).

Local tolerance: No dedicated local tolerance studies available but assessment inciuded in repeat-dose

toxicity studies. At iOOug BNT162b2 V8, there was mostly light to moderate oedemas but in some

cases severe oedema. The severity increased with the 2nd and 3rd injections. The interim data for

3Oug BNTi62b2 V9 exposure indicated similar effects.

2.2.4. Ecotoxicity/environmental risk assessment

As the active substance is a vaccine product (which additionally is based on naturally degradable

mRNA and lipids), no ERA is considered necessary.

2.2.5. Discussion on non-clinical aspects

Pharmacology: The proposed medical product is composed of a modRNA formulated with functional

and structural lipids forming lipid nano particles (LNP5), the latter having the purpose to protect the

modRNA from degradation and enable transfection of the modRNA into host cells after IM injection.

The composition of the LNPs is likely to affect the distribution of injected BNT162b2. In addition, it

cannot be excluded the LNP composition contributes to the overall immunogenicity (see also toxicology

below). Applicant should provide a more detailed clarification of the mode of action of 5NT162b2, e.g.

which cells types will take up the LNP, translate the moURNA and express the S-protein on the surface.

Moreover, which cell types/organs will be targeted by the immune defence system, when the vaccine is

in action. Further information on the potential activity/mode of action of the two novel excipients

should be provided

(OC).

Regarding the structural and biophysical characterization of the modRNA, some information is missing

(e.g. a schematic description and comparison of both vaccine variants including the position of added

cytosines nucleotides and optimized codons, coding and non-coding sequences, m4JU content, an

assessment of sequence structure relation to immunogenicity)(see

for details).

Regarding the

results obtained from the Western Blot in the in-vitro studies, a semi quantitative analysis of the

results should be provided and in the analysis of the blot, some missing scientific information and

explanations should be added by the applicant

(OC).

It can be noted that an overview of the structural

and biophysical characterization of P2 S as a vaccine antigen has been provided. While it is flot

considered to be of critical importance for the assessment in this procedure, it still provides a scientific

understanding supporting the nonclinical key studies of humoral and cellular immune response,

including SARS-CoV-2 neutralizing IgG, as well as SARS-CoV-2 challenge nonclinical P0C.

In-vivo pharmacodynamics: The humoral and cellular immune response following IM administration of

BNT162b2 (V9) was investigated in mice and nonhuman primates but a more in depth discussion on

the suitability of these pharmacological animal models has not been provided (e.g. susceptibility for

SARS-CoV-2 infection; potential bias for Thi- or Th2-skewed responses has been well characterized for

certain mice strains) and the relevance of the immunogenicity data for the clinic (e.g. only single

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immunisation in mice). Also, no or limited attention to the induction of long-term memory responses

nor immunogenicity and protection in aged animals has been paid

(OC).

That being said, the induction

of virus neutralizing antibodies in both mice (VSV-SARS-C0V-2 5) and primates (SARS-C0V-2)

indicated that BNT162b2 immunization has the potential to induce neutralizing antibodies also in

humans. Thus, vaccination with modRNA is expected to induce robust neutralising antibodies and a

concomitant T cell response to achieve protective immunity. Nevertheless, no fufther discussion was

provided regarding the possibility of autoimmune responses induced by the ModRNA. The Applicant is

invited to further discuss the risk that the mRNA vaccine can trigger potential autoimmune responses

and how they plan to possibly evaluate their occurrence (OC).

In mice, the immune response was assessed by single immunization only. Taking the phenotyping of B

and T cells in aggregate, the data indicates a concurrent induction of SARS-CoV-2 S-speciflc

neutralizing antibody titers and a Thl-driven T-cell response by immunization with BNT162b2 (this was

also seen in nonhuman primates).

There are some issues with study R-20-0085 regarding the immunogenicity of the LNP formulated

modRNA encoding the viral S protein (V9; e.g. regarding the absence of IgG2A and IgGi

characterization of RBD, experimental design that would allow an effective titer comparaison between

experiment) (see OC for details). There are also some issues with the study on multiplex analysis of

cytokine release from murine Splenocytes Day 28 after Immunization with BNT162b2 (the use of “5”

as compared to “i ug” BNT162b2 for the Luminex analysis in the Pharmacology written summary,

page 18 (last paragraph) as compared to in the repoft R-20-0085)

(OC).

Moreover, it is noted that

high levels of the Thi cytokines IFNy and IL-2 ifl multiplex immunoassays were detected after re

stimulation with the 5 but not RBD overlapping peptide mix, although RBD is part of the 5 protein. This

should also be clarified (OC).

Concerning the nonhuman primate (rhesus macaques) studies, the applicant considers the human

convalescent serum panel as an assessable benchmark to judge the quality of the immune response to

the vaccine. The reasoning behind this can be followed. The assumption that the immune response to

SARS-CoV-2 infection provides some measure of protection from disease upon subsequent exposure to

the virus, appears plausible. There were a number of specific questions regarding the NHP proof of

concept study (study VR-VTR-1067) which could be considered demonstrate immunogenicity and viral

clearance in NHP but insufficient to fully demonstrate efficacy against the disease (issues to consider

were e.g. on the report itseif, the animal model relevance, technical aspects, endpoints, immunological

aspects) (OC).

The applicant needs to a) precise for the Luminex data how the reference curve for has been

constructed, what does represent the arbitrary U/ml used and how it is referring to the serum dilution

factor; b) define the criteria for choosing a iO-30% infection rate of Vero cells; c) Methods to quantify

antibody production in the different experiments dufter and consequently cross-comparison between

experiments is hardly impossible. Indeed, it is important to distinguish neutralizing antibodies from

non-neutralizing antibodies. In this study, total antibody response is measured using a luminex assay

and results expressed on U/ml and for the neutralization assay resuits are expressed in VNT 50. The

applicant needs to provide an estimation of the non-neutralizing antibodies in the whole antibody

response (OC). d) Tt is important to notice that on figure 6 of study report, neither panel A nor panel B

highlight the consumption of IgG 51 binding antibodies after challenge nor the increase due to B

memory response following the challenge: this would need to be furtherjustified by the Applicant

(OC). The report VR-MQR-102i1, on Si-binding rhesus macaque serum IgG levels detected by a

direct binding Luminex immunoassay, was not provided. This should be submitted

(OC)

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Concerning the characterization of the T cell responses, the Applicant suggests the S-specific IFNy

producing T cell responses, including a high frequency of CD4+ T ceils that produced IFNy, IL-2, or

TNF-a but a low frequency of CD4+ cells that produce IL-4, indicates a Thl-biased response occurred

after the BNT162b2 (V9) immunization. This reasoning appears plausible, however, there was no

reference to what to expect from a typical Th2 biased response to enable a comparison of the current

data. Nevertheless, the role of such a Thi biased response was put in the context of antigen-specific T

cell responses playing an important role in generation of antigen-specific antibody response as well as

in elimination of infected cells to mediate protection against disease. However, the potential

importance of T-cell effector celis for a putative protection against SARS-Cov-2 infection after

BNT162b2 (V9) immunization was flot further investigated or discussed.

When immunized macaques were challenged with SARS-CoV-2, a clear and statistically significant

effect was observed on reduced presence of viral RNA in bronchoalveolar lavage (BAL), nasal and

oropharyngeal (OP) swabs. A clear effect was also recorded by blinded X ray scoring of the lungs. A

protective effect is also evident in the CT score Day 3 after challenge, however at Day 10/EOP, there

was a CT signal in 2 out of six BNTi62b immunized monkeys at the same level as observed in the

control group. That signal is of unclear significance since also in i out of 6 pre infection BNTi62b

immunized animals a similar CT-score signal was observed. The size of the study prevents any firm

conciusion on these observations. A histopathological examination of lung tissues is ongoing and a

submission ASAP as an addendum is awaited (OC). Furthermore, the data from the individual monkeys

should be provided for the RT-qPCR test for presence of SARS-CoV-2 RNA (OC).

In the NHP pharmacology study (Study VR-VTR-i0671), rhesus macaques were immunized on days 0

and 21. Some other covid-19 vaccine candidates have different prime-boost intervals, such as 4 weeks

for both ChAdOxi (Graham et al., 2020) and mRNA-i273 (Corbett et al., 2020). Considering that the

time between the first and second vaccine dose may have a significant impact on the immunological

response, the applicant is asked to provide the rationale for the chosen prime-boost interval (21 days).

(Graham et al., 2020: httls://www.ncbi.nlm.nfh.gov/tmc/articles/PMC7385486/ Corbett et al., 2020:

https://www.ncbi.nlm.nih.gov/nmc/articles/PMC7449230/

)

(OC). The Spike protein of SARS-CoV-2

undergo mutations, and it thus critically important to investigate the biological significance of these

variants in relation to the development of Spike-based covid-i9 vaccine candidates. For example,

Korber et al. present evidence that there are now more SARS-CoV-2 viruses circulating in the human

population globally that have the G6i4 form of the Spike protein versus the D614 form that was

originally identified from the first human cases in Wuhan, China. Further, Li et al., states that as of

May 6, 2020, 329 naturally occurring variants in Spike protein have been reported in the public

domain. The applicant is asked to discuss how the chosen Spike afltigen variant in BNT162b2 relates to

the Spike variants currently on the dominant SARS-CoV-2 viruses circulating in the human population.

(Korber et al., 2020: https://www.ncbi.nlm.nih.gov/mc/articles/PMC7332439/ Li et al., 2020: httns://

doi.org/iO.1016/i.cell.2020.07.012

)

(OC). The rhesus macaques were challenged with the SARS-CoV

2 USA-WA1/2020 isolate. To our knowledge, this stram does not contain the D614G mutation. This

mutation is reported to rapidly accumulate in the circulating SARS-CoV-2 strains and may increase the

infectivity several-fold compared to the original Wuhan-1 stram. The applicant is asked to discuss the

relevance of the NHP challenge study resuits in relation to the stram used for challenge and the strains

circulating in the human population (OC).

In conciusion of the preclinical pharmacology, the presented data, including immunogenicity, triggering

of neutralizing IgGs and Thi response and reduced presence of viral RNA in challenged animals as well

as radiological ung parameters (to be confirmed by histopathology), provide some support for the

vaccination approach. It can be noted that in the primary proof-of-concept study, the use ofjuvenile

rhesus monkeys with no or only mild cllnical symptoms for the preclinical efficacy testing has

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limitations in its value as a disease animal model for human Covid-19 (which is a clearly age stratified

disease, mostly affecting the elderly). In addition, the low number of animals of the male sex only that

were studied only for a short time period weakens the conclusiveness of the study. However, due to

species differences in the immune system between animal model species and humans, the final call on

whether this candidate vaccine will work sufficiently well in humans will entirely rely on the clinical

o utco m e.

Pharmacokinetic (regarding the two novel LNP excipients): The applicant is requested to provide

qualification data for LC-MS/MS method used to quantify the two novel LNP lipids in the non-clinical PK

study (DC). The Applicant is also asked to justify the choice of an IV study instead of an IM study,

which would have a more clinical relevance. The difference observed in terms of PK absorption should

be discussed (DC).

It is worth to notice that the lipid displaying a persistent kinetic over time in hver is ALC-0159, ie the

one that does flot contain any PEG, although PEG is known to be used to increase half-life of many

recombinants. The Applicant will have to justify this observation, as well as to discuss the difference of

kinetics profile between the two lipids. The Applicant is also requested to estimate the delay of the

clearance of the ALC-0315 from the hver, as this could have an impact on the safety profile (OC).

Biodistribution: As expected for an RNA, the expression of the surrogate luciferase reporter RNA was

transient and decreased over time. It is acknowledged that the biodistribution of the mRNA mostly will

be dependent on the composition of LNP and the apphicant has provided data that differences in LNP

formulation affects the biodistribution of the luciferase modRNA and luciferase protein expression. The

Applicant mentions that the LNP-formulated luciferase-encoding modRNA tested in this study have the

exact same lipid composition than BNT162b2. It is however not chear to understand which of the three

tested LNP formulation is present in the drug product, BNT162b2 variant VO. The Applicant should

comment (OC).

RNA stability and kinetics are not expected to be the same for alI RNAs and are influenced by the

nucleosides of the RNA and although expression of the full-length spike (S) protein is expected to

follow similar kinetics of that of the luciferase with a transient expression fading over time, it cannot be

excluded that differences in stability/persistence of the signal could differ between the luciferase

protein and the spike (S) protein. It can be noted that there is no information on the similarities of

the mRNA modifications of the non-coding regions between the luciferase modRNA used in the study

and the modRNA used in BNT162b2. The applicant is asked to provide more information on the

luciferase reporter RNA, in particular, whether the untranslated sequences are similar to that of the

BNT162b2 modRNA and therefore at least the stability of the mRNAs are somewhat comparable. (DC)

The biodistribution of the vaccine has been evaluated in mice, using 2 jig mRNA (encoding for

luciferase). In humans and in the repeat-dose study in rat using the V9 version, 30 ig (per

administration) was used. It is not clear if this difference in RNA concentration results differences in

the amount of LNP used. The applicant is therefore asked to clarify if there were differences in the

amount of LNP used in the biodistribution study and the repeat-dose study /chinical trials and if

so,

discuss how this could affect the distribution and safety evaluation observed in the clinic compared to

non-clinical data (DC).

The bioluminescence method used to determine the surrogate luciferase moURNA biodistribution has

flot been validated or quahified and no discussion on its sensitivity has been inchuded. Only three

females were investigated which is considered a low number. Moreover, only one dose level (given as

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single injection) was tested (compared to two injections given clinically). The sensitivity of the method

and dose propottionality effects have therefore flot been determined.

The applicant has only discussed distribution at the injection site and to the livet. To the unttained eye

it is not clear that the bioluminescence signal is solely livet specific. Although the signal appeats to be

in the livet region, from the data submitted it cannot be excluded that the bioluminescence signal

could inciude distribution to othet organs located in proximity of the livet.

Sevetal literature reports indicate that LNP-fotmulated RNAs can disttibute rathet non-specifically to

several organs such as spleen, heart, kidney, lung and btain. The observed extra-hepatic distribution

of modified RNAs have been detected at much lower levels compared to the livet when measuted with

techniques detecting nucleic acids (for example btanched DNA analysis). This raises the concetn If the

sensitivity of the bioluminescence method is sufficient to detect potentially weaket biodistribution to

sites besides the livet and injection sites?

The vaccine is intended to be given to patients twice with a boostet dose aftet 22 days. The booster

dose scheme could inctease the risk for infiammatoty teactions at sites of exptession and thetefore a

wide biodisttibution ptofile might flot be optimal ftom a safety perspective. The applicant is therefore

asked to provide mote information regarding the biodisttibution assay and should discuss the

sensitivity of the biodisttibution method. The choice of using a non-validated/non-qualified

bioluminescence method to determine the biodistribution of a suttogate lucifetase protein instead of

choosing to detect the actual modRNA used in the vaccine candidate BNT162b2 should be justified

(OC). Moteovet, the applicant is asked to consider the possibility of a wider biodistribution pattern

than observed and discuss the possible safety consequences of a wider biodistribution profile of

5NT162b2. (OC)

Results from the Luminex-based multiplex assay revealed that immunisation with LNP fotmulated

luciferase modRNA transiently increased levels of MCP-1, IL-6, lP-lO, at 6 houts post-immunization.

These tesults in mice suggest that the LNP can activate the nnate immune system of mice, by the

synthesis of pro-inflammatory cytokines. As the effect was ttansient, this effect could be considered as

an adjuvant-like effect. Overall, on the basis of above, the LNP fotmulation is expected to have nat

only a tole to protect modRNA from nucleases degradation, and facilitating cellulat ttansfection, but

also adjuvant like effects.

In view of potential acute immunotoxicity mediated by LNPs, does the Applicant possess data on other

timepoints (earliet than 6h ot beyond) regarding the cytokines measurements? (OC)

The Applicant is also asked to discuss the absence of an in vitto hPBMC stimulation assay in healthy

donots to assess reactogenicity (CC).

Extrapolating to clinics, the Applicant is requested to discuss the level of IL-6 cytokine induced by

LNPs, considering that asymptomatic but infected subjects candidate to vaccination, could display

higher IL-6 levels duting early phase infection (OC).

Toxicology: The fitst “V8”-repeat-dose toxicity rat study has some documentation issues that possible

would have to be followed up tegarding its GLP status (a GLP inspection has been initiated) and

thereby increasing the uncettainty of the interpretation of the tesutts (OC). That being said, as the

toxicological outcomes ftom the V8 and V9-studies ate ovetall similar (and the two studies wete

conducted at different sites), the V8-results ate consideted to be useful for tisk assessment. Thete is

also some uncertainty tegatding which production process (two possible) was used tot the test

substance in the V9 study (also dependent on the quality assessment which has yet to start).

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Only the whole formulation (modified RNA in LNPs) were used, so there is no toxicological data on the

LNP alone or its specific novel excipients. Overall, the V8 and V9 test substances invoked a strong but

mostly reversible immune-linked response in rats after 17d exposure. It is unclear If or how much of

that immune response is attributable to the LNP components (which are included in the formulation as

excipients). There is some pharmacokinetic data that indicates that the LNP has the potential to induce

a transient immune response, but it can also be noted that most exogenous biomolecules tend to

generate some degree of transient immune response, so such observations would not be unexpected.

While no extensive pharmacological assessment has been conducted in rat (only in mouse and non-

human primate, with no deeper discussion on the choice of animal models

[OCj),

immunogenicity tests

on blood samples in the V8 repeat-dose toxicity study indicate that rats generate SARS-CoV-2

antibodies, partly supporting the choice of animal model. Other SARS-C0V-2 immune responses in rat

remain unclear. The immune responses, especially at the injection sites (e.g. oedema, erythema),

seem to fncrease with each injection in the studies (n=3). There was a marked increase in acute phase

proteins, fibrinogen and reduced albumin-globulin ratio (but no increase in cytokines with V8, unclear

for V9). There was also a general increase in immune cells (LUC, neutrophils, eosinophils, basophils)

and a decrease in red blood cell parameters (reticulocytes, RGB, HGB, HCT). The spleen was enlarged

at both 3Oug V9 and lOOug V9 and the lymph nodes were enlarged mostly at lOOug (V8) but also in a

few animals at 30g (V9). While an immune response is expected from V8 and V9, the strong reaction

of the injection site and immune system in rat is difficult to interpret/risk assess as the vaccine

candidate(s) are derived from a novel vaccine platform. There is also the possibility, which is difficult to

assess non-clinically or effectively in-silico, that the generated antibodies may react with endogenous

proteins. An absence of dose-response designs in the studies increases the difficulty to interpret the

effects.

As the pharmacokinetic distribution study in rat was limited (mainly giving data on hver), the

distribution (and its effects) has to be inferred indirectly from the toxicological studies. There is some

uncertainty in this regard as not ali tissues have been investigated in the V8 study and

histopathological details are unknown for the V9 study and it is recommended to study as many tissues

as possible (the following tissues were flot studied: nasal body cavity, chitorial gland, dorsal root

ganglion, larynx, mandibular lymph node, tibial nerve, preputial gland, ureter, Zymbal’s gland). While

there was no severe pathogenesis in iver, there were some reversible functional hepatic and/or biliary

effects with V8 and V9 (enlarged hver, vacuohation, strongly increased gGT levels at >200Db and

activity, minor-moderate increase in levels of ALT and ALP) which may be hinked to the LNP. The gGT

changes were flot observed with 3Oug V9, which may be due to variant differences and/or a lower

dose. Considering that vaccines are expected to generate little or no toxicity (beyofld local tolerance

and immune response effects) and that BNT162b2 derives from a novel vaccine platform in the context

of a pandemic, further discussion mi these effects and a possible mention in the SmPC is required (see

OC for specific details). DART data remains to be submitted and the choice of animal model and

experimental design to bejustified

(OC).

With regard to the characterization of the novel LNP components, these are flot considered primarily as

adjuvant substances. While some degree of LNP-specific immune response cannot be ruled out (as

demonstrated in the

in vivo

biodistribution study

iii

Balb-C mice by pro-infiammatory cytokines

induction [MCP-1, MIP-1, TNF-a, IFN-y, IL-6, lP-bl at 6 h post immunization see study R-20-

0072), no further experimental toxicological studies are considered necessary as the use of the whoie

formulation (RNA

LNP) in the repeat-dose toxicity and DART studies is sufficient to quahify the novel

excipient hipids in combination that the overall effects are also being assessed in the ciinical trials. That

being said, as the hipids contain an acetamide moiety which has been hinked to carciflogenicity

ifl

—

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animals, inciuding livet tumors, potentially related to genotoxicity, and livet distribution and functional

effects have been observed in rat, an extended discussion ofthese lipids is tequested

(OC).

It is unclear at this stage of the rolling review how these effects are depicted in the SmPC.

2.2.6. Conclusïon on non-clïnical aspects

Based on the provided data so far there are no non-cilnical major objections. The applicant will need to

sufficiently address the other concerns raised to be granted MAA from a non-clinical perspective. Other

non-clinical elements in further rolling review cycles are expected to define the safety profile of the

vaccine.

2.3. Clinical aspects

N/A

2.4. Risk management plan

N/A

2.5. Pharmacovigilance system

N/A

SCIENTIFIC OVERVIEW AND DISCUSSION

an responses to

questions raised in previous cyde(s)

N/A

4. Benefit risk assessment

N/A

5. CHMP list of questions

5.1. Quality aspects

Major objections

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GMP

GMP status for DS and DP manufacturing sites is currently flot acceptably demonstrated:

A statement on GMP compliance issued by EU supervisory authority of the DS and DP

manufacturing and testing sites Wyeth BioPharma Division, Andover, United States and Pfizer

Inc, Chesterfield, United States should be available by adoption of the CHMP opinion.

b. The MIA for PHzer Puurs is limited to the formulation and filling only. Tt should be clarified if

authorisation will be extended to ali operations listed in 3.2.P.3.1, including LNP manufacturing.

Moreover, GMP certiticate or a statement of GMP compliance issued by the Supervisory

authority of BioNTech Manufacturing GmbH, Mainz, Germany should cover batch certification of

the DP.

Drug substance and Drug product

Comparability between clinical and commercial material has not yet been demonstrated, which

raises unceftainties about consistency of product quality and hence uncertainties as regards

product safety and efficacy of the commercial product. Significant differences between batches

manufactured by DS Process i and 2 are observed for the CQA mRNA integrity. In addition, the

characterisation of BNT162b2 DS is currently not found acceptable in relation to this quality

attribute. This is especially important considering that the current DS and DP acceptance criteria

allows for up to

5Q%

fragmented species. Therefore, the dossier should be updated with

additional characterisation data on mRNA integrity in sections 3.2.5.2.6 (comparability) and

3.2.S.3 of the dossier.

a. Truncated and modified RNA species should be regarded as product-related impurities. Even

though two methods, namely agarose gel electrophoresis and capillary gel electrophoresis

(CGE), have been applied to determine RNA integrity of BNT162b2 DS, no characterisation data

on truncated forms is presented. Results obtained on RNA integrity by CGE and agarose gels

should be included in the characterisation section (3.2.5.3). The truncated forms should be

sufficiently characterised, i.e. they should be described, and it should be discussed if the

fragmented species are expected to be similar between batches. In addition, the possibility of

translated proteins other than the intended spike protein (S1S2), resulting from truncated and!

ar modified mRNA species should be addressed and relevant protein characterization data for

predominant species should be provided, if available.

Upon changing to DS Process 2, a decrease in RNA integrity was observed (only numerical

values provided). Concerning this difference in RNA integrity between Process i and Process 2

DS batches the Applicant is requested to provide capillary electropherograms together with an

evaluation of any batch differences in peak patterns. The potential safety risks associated with

truncated RNA isoforms should be thoroughly discussed with reference to the batches used,

clinical experience and possibly literature data. The quantitative and qualitative differences

observed between Process i and 2 should be discussed with respect to their impact on safety

and efficacy.

For Process 2, the

CTP

and ATP volumes were adjusted before the manufacture of DS batch

PPQ3 to align better with RNA integrity results from Process 1. Additional batch data (from

PPQ4 and PPQ5) should be provided to confirm that the optimised Process 2 allows for reaching

RNA integrity ievels consistent with the Process i batches.

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vaccine (nucieoside modified)

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overvew and list ot questions

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After contact with the applicant it was confirmed that DP batches manufactured from early

Process 2 batches, with lower RNA integrity, have been recently introduced in clinical trials.

However, as the cut-off date for the clinical Interim Analysis (IA) was changed, the IA doesn’t

include data from subjects dosed with Process 2 material, and the Company does flot expect to

have Process 2 included in the Final Analysis dataset. Therefore, the proposed acceptance

criteria of

50%

intact RNA for RNA integrity is considered too wide compared to clinical batch

data, 69-81%. The proposed release and sheif-life acceptance criteria for the DP should

therefore be tightened based on the clinical data inciuded in the dossier ar cilnically qualified by

other means.

Release data provided for some of the DP batches indicates a possible decrease in mRNA

integrity during the manufacturing of DP. The applicant should therefore discuss possible root

causes, and present comparative resuits for DS and DP, an RNA integrity. A consequential need

for a more stringent DS specification should be considered. Sections 5.4.1 and P.5.1 in the

dossier should be aligned and updated accordingly.

Drug product batches manufactured at the commercial facility (whole manufacturing process at

the commercial site Pfizer, Puurs, at commercial scale, drug substance from process 2) were flot

presented. Process validation (PPQ) for commercial scale batches are already initiated and

validation data should be provided. Batch resuits for at least 2 commercial scale batches

representative of the commercial process should be presented. Comparability of commercial

batches with clinical batches should be demonstrated and the data should be provided. The

claimed sheif-life and storage condition are nat yet acceptable since no stability data is available

for batches from the commercial manufacturing site and scale and shelf-life is based an very

small scale (development) batches (less than

bio

of the commercial scale), nat representative of

the commercial batches (manufacturing site, scale, process for the drug substance). Additional

stability data (6 months at long-term storage condition) should be presented.

Other concerns

Drug substance

The applicant

plans to update a number of sections along

the dossier and states the following: “Data

for this section is pending and will be updated once the data has been generated, analysed, and

verified”. Until these data are available for assessment, no final conclusions can be drawn on the

concerned sections.

General information (Sl)

3. The proposed mechanism of action should be presented in 5.1 General Information.

Description of manufacturinq process and orocess controls (5.2.2)

Information an the final batch volume should be provided. The Applicant should state either the

total batch volume ar the approximate number of DS containers generated from one batch. Section

3.2.5.2.2 should be updated accordingty.

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It is noted that some parameters and ranges may be updated after PPQ and additional

characterization studies are completed. These updates could have an impact on overall assessment

of the manufacturing process description, leading to additional issues. However, the following

issues have already been identified and should be addressed:

It should be indicated that the incubation time during GTP/N1-methylpseudo UTP bolus feeds is

a global time for the 11 feeds

b. The strategy for UFDF membrane lifetime validation is to per-form concurrent validation of the

membranes at commercial scale. This is found acceptable, provided that the Applicant will

inciude control of the feed flow rates, transmembrane pressure and membrane surface area in

section 3.2.5.2.2. The dossier should be updated accordingly.

The transfers of the UFDF pool into a single PE flexibie container before and/or after 0.45/0.2

pm filtration should be ciarified and should appear in the DS process flow diagram.

d. The DS filling volume tange in the EVA fiexible containers should be defined in line with the

volumes validated for shipping.

Control of materials (5.2.3)

Representative CoAs or full specifications should be provided for starting and non-compendial raw

materials used in the manufacturing of BNT162b2 DS. It is expected that information regarding the

microbiological control is included. Additionally, ali raw materials should be demonstrated to be

free from contaminating RNases, unless otherwise justified.

It is noted that for starting and raw materials used at Andover, additional materiai testing will be

performed and provided when availabie. Where relevant, the applicant shouid consider in house

testing for the functional activity of starting and critical raw materials such as the enzymes used in

the manufacturing process. The information should also be completed with the analytical methods.

As the 5’-cap structure is complex, its synthesis should be descri bed. The impurities and by-

products generated during its synthesis should be discussed.

Linear DNA template

9. Additional details on relevant characteristics and origin of the E. coli stram DH1OE as weil as source

and an overall description of generation (flow chart of the successive steps) of the piasmid used as

template for the production of Drug Substance should be provided.

10. Release testing of plasmid MCB and WCB should be compieted with a percentage of the expected

sequence rather than “comparable to the refetence sequence”. Moreover 100 % homology is

requested for the coding sequence; for the other parts of the plasmid any mutation should be

assessed.

11. The specification for the future WCBs should be completed with the petcentage of via ble ceils with

an appropriate acceptance limit. Moreover, an acceptance limit for viable cell concentration should

be set, and a percentage of the expected sequence (% homology) for DNA sequencing as

requested for piasmid MCB and current WCB should be proposed. Finaily, the anaiytical methods

should be indicated.

12. The cell bank stability protocol (mncluding test parameters and corresponding acceptance criteria)

should be provided. Otherwise, the performance of the WCB should be checked during the

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manufacture of each batch of plasmid DNA, for example by following the trends in bacterial growth

and plasmid yield.

13. It is recommended that cell banks be stored in two ot more separate locations to minimize the

risks of their total loss as a result of a catastrophic event. It is indicated that Pfizer facility at 875

Chesterfield Parkway West, Chesterfield is the only proposed storage site for MCB and WCB. A

ciarification whether any risk amelioration strategies are in place to avoid the loss of cell banks

should be requested.

14. Information should be provided regarding the reference material used in the restriction map

analysis and DNA sequencing determination for MCE and WCB used for plasmid DNA template

production.

15. The manufacturing process to obtain the linear DNA template should be completed with the

following information:

The quantity of linear DNA template obtained in each batch should be stated

b. The chemical agent used for chemical lysis of the cells should be mentioned and its clearance

should be demonstrated to be sufficient.

The mention “or equivalent” for the restriction enzyme should be deleted.

d. The Applicant should confirm that implementation of changes in the manufacture of the linear

DNA template will be applied for in a variation application.

16. The specification for the linear DNA template should be revised with narrower limits for purity and

process-related impurities taking into account the batch analysis resuits. A high level of DNA

impurities could impact the activity of the T7 polymerase during the Transcription phase of the VS

production.

17. Appropriate descriptions of ali analytical methods used in the release control of the linear DNA

template as well as summaries of the results obtained in the method validation/qualification studies

should be provided.

18. The reference material for plasmid identity testing should be described. (Rapp Q10)

19. The stability of the linear DNA template and the stability of the filtered circular plasmid DNA

intermediate should be addressed. A sheif life for the linearized DNA template should be

established and a stability protocol covering the proposed storage period should be included.

Relevant available data should be provided to support this proposal.

Control of critical steps and intermediates (S.2.4

20. It is stated that 005 result for in-process controls would trigger an evaluation of the deviation to

determine if the batch could be further manufactured. It should be confirmed that 00S resuits will

lead to batch rejection.

Process validation and/or evaluation (S.2.5)

21. Several validation studies and full PPQ data are still pending for the manufacturing process at

Wyeth Biopharma, Andover. Therefore, additional information is needed:

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Results for PPQ4 and PPQ5 batches should be provided to confirm the consistency of the DS

manufacturing process after the change of ATP and CTP volumes in the IVT vessel at PPQ3 and

onwards. The description of deviations and investigation conciusions should be provided, as

well as the evaluation of removal of impurities for the five PPQ batches.

A time-plan for the submission additional process validation data should be provided before

marketing authorisation approval.

22. Residual DNA template is present at higher level in PPQ3 batch (211 ng DNA

mg RNA) than in

PPQ1 and PPQ2 batches (10 and 23 ng/mg); the robustness of DNase I digestion step should be

further investigated.

Manufacturing arocess develoment (S.2.6

23. It is noted that the ranges studied for addition volumes for CTP and ATP as stated in 3.2.S.2.6 are

81.0-143.8 and 90.0-135.1 mg/L respectively and that the acceptable ranges proposed are 85.4-

143.8 and 85.4-135.1 mg/L. It seems as if the lower acceptable tange of 85.4 mg/L proposed for

ATP volume have not been studied, this needs to be clarified. In addition, it needs to be justified

why the lower end of the ranges for both CTP and ATP volumes remained unchanged although the

target ranges were increased (from 90 to135.1 and 107.9 mg/L respectively), to avoid that these

nucleotides will be limiting in order to inctease the percentage of the RNA integrity.

These ranges need to be further justified and clarified and the dossier updated accordingly.

24. In the In vitro transcription (IVT) step, the magnesium dependent T7 RNA polymerase assembles

ribonucleotide building blocks. Since magnesium can be chelated by pyrophosphate released by the

addition of each ribonucleotide pyrophosphatase is important to maintain sufficient levels of free

magnesium. The Applicant states that added volumes of these two enzymes have been identified

as non-CPPs as they are most likely to impact yield only. This conclusion is flot entirely agreed

upon.

It needs to be further justified why these parameters are not classified as CPPs.

Regardless of the classification as non-CPPs or CPPs it is strongly recommended to include an

appropriate control of the added volumes of the enzymes T7 polymerase and pytophosphatase

in sections 3.2.S.2.2 and 3.2.5.2.4 of the dossier.

In addition,

needs to be ciarified if the actual volumes loaded are calculated based an enzyme

activity as stated in the certificates of the actual batch of the enzymes that are used. (See also

question in section 3.2.S.2.3 above).

25. The Applicant should provide data on the T7 RNA polymerase and proteinase K levels in additional

commercial scale DS batches, once testing is complete. In addition, the Applicant should briefly

describe that the methods applied to determine the concenttations of these two enzymes in the

BNT162b2 DS samples and confirm that these methods are fit for purpose.

26. Differences in the poly(A)tail pattern were observed when comparing the Process i and Process 2

DS batches. The differences in the extent of cytidine monophosphate incorporation and

transctiptional slippage should be further investigated and the possi ble impact an efficacy and

safety should be discussed. The only Process 2 DS included in the comparison was manufactured

prior to the adjustment of CTP and ATP volumes. Results obtained on the PPQ batches,

manufactured after adjustment (PPQ 3, 4 and 5) should also be presented and discussed.

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27. The level of information in the dossier presenting the available process characterisation studies is

flot sufficient to allow assessment: the results of the studies should be presented, preferably

summarised in figures ar tables.

28. An overall control strategy was presented but some parameter afld ranges may be updated after

PPQ and additional characterization studies completed. As for assessment of overall control

strategy, a complete set of data and information is needed, this document will be assessed when

finalised. A time plan for the submission of the final data set of the control strategy should be

provided.

Characterisation (5.3)

29. In the Development History and Comparability section (3.2.5.2.6), the expressed protein size is

evaluated by in vitro expression followed by Western blot. Results obtained by this method could

be regarded as biological characterisation and should be included in section 3.2.5.3. The method

needs further description and the results should be sufficiently characterized.

A brief method description including conditions for protein expression, gel separation, and

western blot assay should be provided.

b. The expected protein size should be stated and supported by theoretical caiculations.

The identities of the two distinct bands should be explained. If possible, the identities of the

bands should be confirmed and characterized by LC-MS/MS.

The Applicant should provide data on protein expression in terms of percentage of successfully

transduced HEK293 cells using the lipofectamine transfection system.

30. Even though biological characterisation might not be possible to perform on DS, the strategy to

determine potency and relevant functional assay(s) should be described in section 3.2.5.3. Resuits

obtained on DP could be included, to demonstrated functionality.

31. NGS technology has been used as an orthogonal method to confirm primary sequence but details

are missing about the resuits of this analysis in terms of coverage of the target genome. A brief

description of the NGS method, and the resuits obtained with it should be provided.

32. As regards 5’ end of the RNA, relative abundance of each species (capped, non-capped and/ar

incampletely capped) is given as major

(>50%)

for the expected 5’-cap structure, minor (5 to

SO%)

and trace (<5%) for other species. However, a more precise quantification of each uncapped

or incompletely capped species should be provided. Moreover, the potential contribution of

uncapped or incompletely capped structures to the potency of the 6NT162b2 DS should be

discussed.

33. The Applicant should discuss the relationship between 5’-cap heterogeneity and UsRNA production.

A risk assessment should be provided. This should be also taken into account in the justification of

DS specification.

34. It should be addressed whether, under expected storage conditions, individuat base modifications

occur (e.g. depurination, oxidation). Based on this discussion it may be necessary to review the

impurity methods and specifications for appropriateness to detect relevant degradation under long

term conditions.

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Control of drug substance, Specifications (5.4.1)

35. The proposed commercial drug substance specifications, the method descriptions and the method

validation summaries should be updated to inciude in-house method identification numbers for the

non-compendial methods. The information is required in order to provide a ciear link between the

specification and the descriptions and validations of analytical procedures used for routine testing.

Furthermore, for the compendial methods references to relevant parts of the Ph Eur should be

inciuded. Section 3.2.5.4.1, 3.2.5.4.2 and 3.2.5.4.3 of the dossier should be updated accordingly.

Control of drup substance, Analytical procedures (5.4.2)

36. In ali the in-house analytical methods used in the release of DS, method descriptions are based on

“examples” of procedures, controls and standards as well as on “typical” system operating

parameters. These terms raise uncertainties regarding the developmental stage, and the control of

critical steps of these assays. The analytical methods used in the control of DS are expected to be

finalized. The applicant is requested to confirm this and to update the relevant parts of the dossier

with unequivocal method descriptions, inciuding relevant lists of materials and additional details, if

needed. The applicant should also confirm that any significant changes in analytical procedures will

be applied for in a variation application.

37. Regarding the RT-PCR method for determination of DS and DP identity:

Information regarding the positive control used in the should be provided.

b. The proposed assay acceptance criteria for the quaiitative RT-PCR-based assay used for

determination of DS identity requires a Ct value for the positive PCR control of NMT than 32

simultaneous with a Ct value for the negative controls of NLT 32. These criteria are not

considered relevant to support method suitability. More stringent acceptance criteria should be

established and supported by relevant data.

The mRNA extraction step needed for determination of the identity of BNT162b2 DP should be

included in the description of the RT-PCR-based assay and this step should be appropriately

described and addressed in the method validation procedure. This question relates to the DP

part of the dossier.

38. Regarding the ddPCR-based method for determination of poly(A) tails in the mRNA DS:

Information regarding the internal control used in the should be provided.

From the limited description of the ddPCR-based assay for quantification of poly(A) tails it

seems that the cDNA generated using a poly(T) primer is used both as a template for further

amplification of the (poly(A) positive mRNA)-derived cDNA and also as the theoretical input

based on which the final caiculation of the Poly(A) tails is made. This strategy is not

understood. The suitability of this approach and the rationale by which the method is able to

determine the percent poly(A) tails in the mRNA DS relative to the input (which should be

clearly defined) needs to be better described.

With respect to the storage conditions of the cDNA prior ddPCR, storage at room temperature,

however with no hold time defined, is mentioned in the method description, but a storage time

of 3 days at —20°C is examined in the validation studies with respect to method robustness.

These discrepancies should be ciarified. Information on the qualified bt of linearized plasmid

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standard used in the qPCR-based method to quantify the residual DNA template in 5NT162 b2

DS should be provided.

39. Information on the qualifÏed bt of linearized plasmid standard used in the qPCR-based method to

quantify the residual DNA template in BNT162 b2 DS should be provided.

40. With respect to the immunobbot analytical method used for determination of dsRNA in BNT162b2

drug substance:

a. Additional information regarding the critical reagents (such as antibodies), standards and

equipment used as well as representative dot blots and standard curves should be highlighted

in the dossier. The robustness of the method should be appropriately demonstrated in the

validation exercise, if different reagents, e.g. different ciones or different vendors for the

antibodies, are envisaged.

An incubation time of >16h is defined for the primary antibody incubation step. An upper limit

should be defined as well. Unless otherwise justified, ali variable incubation times described in

the method should be considered in the validation exercise, in order to demonstrate the

robustness of the assay.

41. For the capillary gel electrophoresis method, it should be specified how the peaks are integrated to

allow quantitation of the RNA integrity. An integrated electropherogram should be provided as an

example.

Control of drug substance, Validation of analytical procedures (S.4.3)

42. The information in the dossier does flot support that any of the in-house analytical procedures

applied for drug substance has been properly validated in line with ICH Q2. The validation

summaries provided are far too brief and important details are missing. The Applicant should

submit more comprehensive validation summaries of ali non-compendial methods, for example in

the form of short validation reports. The validation summaries should inciude ali relevant

caiculations, acceptance criteria, description of and results obtained for individual samples.

Chromatograms and dose response curves should be inciuded, where applicable.

Module 3.2.S.4.3 of the dossier should be updated accordingly.

43. The method transfer plan or activities should be addressed. It should be noted that, If method

transfer was

will be performed, the folbowing information should be provided. For the non

compendial tests, it should be confirmed that the validation acceptance criteria for the receiving

sites will be the same as for the transferring site (which will be assessed during the RR). For the

anaiytical methods for which comparative analysis will be proposed, it should be confirmed that the

acceptance criteria will be the same as for the intermediate precision validated at the transferring

site (and assessed during RR).

Control of drug substance, Batch analyses (S.4.4)

44. Batch results should be presented for the two newly manufactured batches PPQ4 and PPQ5 to be

able to assess process consistency. This is considered specifically important to verify that the

volume adjustments made for ATP and CTP voiumes before manufacturing of PPQ3 (20Y513C501)

consistently provides reproducible resuits, in particular with RNA integrity levels simiiar to ievels

achieved in process i batches.

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Control of drug substance, Justification of specifications (S.4.5)

45. The length of the poly(A) tails in 8NT162b2 DS is important for RNA stability and translational

efficiency and this test should therefore be included in DS release specification.

46. The proposed acceptance criteria for the percentage afs’- Cap (50%), UsRNA (<1000 pg/pg

mRNA) and Poly(A) tail

(70%)

are flot considered justified and should be tightened to better

reflect the data presented for the DS material used in the manufacturing of the clinical and PPQ

batches. In addition, batch release results from two newly manufactured batches PPQ4 and PPQ5

should be inciuded in the reassessment of the acceptance criteria.

Reference standards (5.5)

47. It should be ciarified for what release and stability testing methods the reference standard is used

and will be used in future. The function of the reference standard should be briefly stated for each

assay, i.e. result evaluation/normalisation, sample compliance, assay control etc. The information

could be provided preferentially in a tabulated form.

48. It is noted that the CRM is derived from a Process 2 DS batch that was established in September

2020. Tt should be explained if another reference standard was used to perform release tests an

Process 1 DS batches. Ali initial reference materials should be listed.

49. The CRM is derived from an early Process 2 batch which has a slightly lower RNA integrity than the

clinical batches and possibly also to future batches, due to target value optimisation. The Applicant

should justify the suitability and address potential risks of using this material as a reference

standard.

50. Neither the storage condition, nor the shelf-life is established for the CRM. The Applicant should

explain if the reference standard is used in any of the methods inciuded in the formal stability

protocol. If this is the case, the Applicant should explain how compliance with the acceptance

criteria can be guaranteed.

51. Since the Applicant intends to establish primary and working reference materials, information on

the preparation, qualification and stability evaluation of the PRM and WRMs should be included in a

PACMP. Otherwise it should be confirmed that a variation application will be submitted in

connection with the introduction of these standards.

Container closure system (5.6)

52. The following additional information should be included in Module 3.2.5-6 of the dossier.

a. A certificate of analysis of one representative batch of the EVAM contact layer demonstrating

compliance with Ph. Eur. 3.1.7.

A specification for the container closure system inciuding dimensions (currently only schematic

drawings are included).

53. A commitment to submit for assessment any unexpected leachable compound from EVA container

closure system reproducibly observed above 1.5 pg/day TDI should be provided.

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Stability (S.7)

54. Process i batch is flot considered representative to process 2 batches. The only parameters studied

for process i batch are RNA integrity and RNA content and the cGE method for RNA integrity was

changed. Therefore, based on the currently very limited stability data presented for process 2

batches (only i-month data available for one batch) no conclusion can be drawn in relation to the

proposed sheif lite for the DS. Thus, in ordet to support shelf lite setting for dtug substance

updated repofts from the ongoing stability studies on the primary batches (including data from the

ongoing ptocess validation batches) should be ptovided.

55. It should be confirmed that future extensions of the assigned DS sheif life will be applied for in

formal variation applications. The following statement should be removed for Module 3.2.5.7.1 of

the dossier; “The sponsor will extend the assigned sheif life without notification pro ufding the real

time stabil/ty data at the intended storage condition is acceptable and within commercial

specifications.”

Drug product

The applicant plans to update a number of sections along the dossier and states the following: “Data

for this section is pending and will be updated once the data has been generated, analysed, and

verified”. Until these data are available for assessment, no final conclusions can be drawn on the

concerned sections.

P.1 Descriition and composition of the drug iroduct fP.1)

56. AlI ingredients, including ptocess aids used in the manufacture, should be specified in the

composition together with a footnote that they are removed during manufacturing. Therefore,

ethanol and components of cittate buffer should be added to the composition. Moreovet, HEPES

and EDTA (excipients used in the Urug substance buffer) should also be added to the composition

table. Section P.1 should be updated accordingly. Ali these ingredients should be mentioned in the

SmPC and PIL.

57. While the final volume of drug ptoduct after reconstitution (2.25 ml) exceeds the vial nominal

capacity (2 ml), it is expected that during ciinical trials it was demonstrated that the method of

preparation is feasible and is robust in ensuring efficient mixing and uniformity of the solution. This

issue should be addressed and if needed, appropriate insttuctions for use (IFU) should be given in

the SmPC and PIL.

Pharmaceutical development (P2)

58. Controlled extraction studies have been performed on the bromobutyl tubber stopper. Leachables

studies are planned to be set up to support the proposed DP shelf-life of 24 months, the TO will be

provided later on during the procedure. The applicant should commit to provide the updated resuits

from the leachables study for assessment.

59. It is noted that some additional heightened chatacterization information will be added in the

formulation development tue. However, the awaited data were flot detailed. Formulation

development should be compieted with characterisation studies showing the homogeneity of the

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suspension during storage at long term or accelerated conditions, after freeze/thaw, or after

dilution with

O.9%

NaCI should be studied.

60. Development data showing homogeneity of LNP or RNA concentration in the vials during filling

process should be provided.

61. Overall control strategy was presented but some parameter and ranges may be updated after PPQ

and additional characterization studies completed. As for assessment of overall control strategy, a

complete set of data and information is needed, this document will be assessed when finalised. A

time-plan for the submission of the final data set of the control strategy should be provided.

62. The compatibility studies of the diluted suspension in the vial and in syringes were performed with

DP diluted to 0.05 mg/mL while dilution for administration is intended to be 0.1 mg/mL: it should

be confirmed that the analytical methods are valid at this dilution. Moreover, the specifications

applied for RNA content and RNA integrity

(+1-

20% of TO) are flot acceptable; in use specifications

should be the same as the sheif-life specifications. It is noted, however, that this section may be

updated as additional studies are completed. The applicant still needs to define in P.8 and the

SmPC/PIL the in-use sheif-life and storage conditions after dilution and first use, in line with

available data.

Manufacture (P.3)

63. The batch formula should be completed with process aids.

64. The lipid nanoparticle (LNP) formation is one critical manufacturing step and some additional

information is requested regarding this step.

a. The tange number of DS bags and DS batches to be thawed should be stated.

According to pharmaceutical development (Section P.2.3.4) 2-8 parallel T-mixer may be used

depending on the batch size and manufacturers equipment. In the description of manufacturing

process (Section P.3.3) it is stated that “one or more” T-mixer(s) are used. The number of T

mixers should be defined in Section P.3.3 and the dossier should be updated accordingly.

A drawing of the T-mixer inciuding further details should be provided, e.g. geometry and

dimensions.

3. It is noted that some parameters and ranges may be updated after PPQ and additional

characterization studies completed. These updates could have an impact on overall assessment of

the manufacturing process description, leading to additional issues. From the first assessment, the

manufacturing process description should already be completed with the following information;

a. The environment grades should be indicated for each step;

holding times will be assessed when complementary data will be available.

4. The applicant should ciarify if the 0.2 pm-filter used for bioburden reduction is identical with the

0.2 pm-filters used for sterile filtration.

It is stated that OOS result for in-process controls would trigger an evaluation of the deviation to

determine if the batch could be further manufactured. It should be confirmed that OOS resuits for

acceptance criteria will lead to batch rejection.

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6. The validation protocol should be completed with the minimum number of consecutive batches at

commercial scale to be included in the PPQ validation process, which should flot be less than 3

batches. DS thaw parameters should be studied. Each thawing method (controlled room

temperature thaw or controlled thaw) should be validated on at least one batch. Moreover, the

mixing speed during dilution of DS shauld be added in the list of studied parameters.

For PPQ, to validate the TFF efflciency, residual ethanol and citrate should be measured with

appropriate limits. During aseptic filling, a homogeneity test of the filled vials should be added with

appropriate sampling and acceptance criteria. Finally, some acceptance criteria are “report results”

with limits to be developed after sufficient manufacturing experience. This is flot endorsed and

acceptance criteria should be fixed before PPQ validation.

8. Acceptance criteria for quality attributes that are requested to be narrowed in the DP specification

should be narrowed as well in the process validation protocol.

It should be confirmed that the Kleenpak Capsule with Supor EKV Membrane will be the one used

for routine DP manufacturing at Puurs. If other filters are used, the extractables

leachables

should be studied before use.

Control of exciients (P.4’l

10. It should be confirmed that cholesterol will be controlled in line with Ph. Eur. monograph

Cholesterol for parenteral use (2397) for future batches and not Ph. bur. monograph Cholesterol

(0993).

11. Additional test for microbial contamination should be included for alI compendial excipients, except

for water for injection. Further, where relevant, a test for bacterial endotoxins should be added

unless otherwise justified.

12. Appropriate documentation for the processing aid excipients ethanol and citrate buffer and the

excipients for drug substance buffer HEPES and EDTA is missing and should be provided.

13. DSPC is used in several medicinal products approved in EU and administered intravenously.

According to the guideline on excipients in the dossier (EMEA/CHMP/QWP/396951/2006), an

excipient used by a new route of administration may be considered as a novel excipient. Therefore,

further discussion should be provided to justify why DSPC administered intramuscularly is flot

considered as a novel excipient and how data from intravenous administration can support safety

of the excipient for this drug product.

14. Specifications for DSPC should include a test for purity of stearic acid, identity of phosphorus, and

the assay specification (9O.O-llO.O%) should be tightened in line with batch results from the

su pplier.

15. For cholesterol and DSPC, the analytical methods for residual solvents and microbial purity should

be described in detail (e.g. detailed chromatographic conditions for GC, sample and standatds

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preparation, detailed caiculation formulae for the SC method and respectively the actual method of

preparation and count for microbial purity).

16. Unless otherwise justified, controls for the absence of RNase should be inciuded in the specification

for excipients, especially Water for Injections.

Control of drun oroduct (P.5)

17. In ali of the in-house analytical methods used in the release of DP, method descriptions are based

on “examples” of procedures, controls and standards as weli as on “typical” system operating

parameters. These terms raise uncertainties regarding the developmental stage, and the control of

critical steps of these assays. The analytical methods used in the control of DP are expected to be

finalized. The applicant is requested to confirm this and to update the relevant parts of the dossier

with unequivocal method descriptions and additional details, If needed. The applicant should also

confirm that any significant changes in analytical procedures will be applied for in a variation

application.

18. The information in the dossier does not suppoft that any of the in-house analytical procedures

applied for DP has been properly validated in line with ICH Q2. The validation summaries provided

are far too brief and impoftant details are missing. The Applicant should submit more

comprehensive validation summaries of ali non-compendial methods, for example in the form of

short validation reports. The validation summaries should include ali relevant calculations,

acceptance criteria, description of and results obtained for individual samples. Chromatograms and

dose response curves should be included, where applicable. Module 3.2.P.5.3 of the dossier should

be updated accordingly.

19. With the exception of osmometry, volume of injections in containers, HPLC-CAD (lipid identities)

and RT-PCR (identity of encoded RNA sequence), which are performed only at DP release, ali other

analytical procedures are conducted at release and stability studies for drug product. It is stated by

the appiicant in section 3.2.P.5.6 that the acceptance criteria used for stability during sheif life will

be the same as the acceptance criteria used for bt release. This is found acceptable, however, the

applicant should confirm that the same acceptance criteria are valid both at release and end-of

sheif-life for the drug product. The specifications document in 3.2.P.5.1 could preferabiy be

updated to include a separate coiumn for the end-of-sheif-life specifications.

20. Test method numbers are missing and should be given to ali analytical procedures used in the

specifications for release and end-of-sheif-life and shouid consequentiy be inserted in the drug

product specifications document and to the descriptions and validations of analytical procedures.

Sections 3.2.P.5.1, 3.2.P.5.2 and 3.2.P.5.3 should be updated accordingly.

21. LNP size for drug product is measured by dynamic light scattering (DLS) and the efficacy of the

drug product depends on the size of the LNP. The proposed acceptance criteria of 40 to 180 nm

seem wide compared to clinical batch data that is found in the range of 59-74 nm for the small

scale clinical batches (“classical LNP process) and 68-71 nm for the emergency supply (“upscale”

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LNP process). The acceptance criteria should therefore be tightened to be in line with what has

been qualified in the clinical studies or clinically qualified by other means and set such that a

cllnically qualified level is assured throughout the shelf-life of the drug product.

22. The mRNA extraction step needed for determination of the identity of BNT62b2 DP should be

inciuded in the description of the RT-PCR-based assay and this step should be appropriately

addressed in the method validation procedure.

23. With respect to the cell-based flow cytometry method used to confirm the in vitro expression of

SARS-CoV-2 spike protein encoded by the RNA in BNT162b2 DP:

Information regarding critical reagents (such as antibodies), drug product control samples and

equipment used should be provided in the dossier. The robustness of the method should be

appropriately demonstrated in the validation exercise, if different reagents, e.g. different clones

or different vendors for the antibodies or different instruments, are envisaged.

It is stated that exact shapes and locations of gates are expected to be different between

instruments and that gates will be shaped and sized to select for the relevant cell populations.

The gating strategy should be established, clearly defined and a description of the rationale for

establishing the gating strategy should be provided. Possible changes observed between

different equipment should be appropriately cross-validated.

Complete examples of resuits (including the three population: P1, P2 and P3) should be

provided for NC, DPC and TS samples

In the table defining assay acceptance criteria, a limit of >30% is established for results

obtained using drug product control samples. In order to unequivocally demonstrate the

suitability of this method, the lower limit strategy should be replaced by a target/interval value.

A value of, or close to,

30%

is considered too low for the demonstration of method suitability

and should be updated based on relevant data.

e. The relevance of the results obtained in the in vitro expression test using a HEK293 cell line for

the in vivo intended targeted cell population should be further discussed and, ideally,

substantiated with characterization data, unless otherwise justified. Additionally, information on

characterisation of the HEK293 cell line used, including specifications should be provided.

The cell culture and transfection steps inciuded in the potency method should be appropriately

considered in the method validation strategy. For example, substantial variation in the culture

parameters (such as passage number and seeding densities) are allowed for HEK293 ceils used

in determining DP in vitro expression. Unless otherwise justified, these possible variations

should be addressed in the validation exercise when investigating assay robustness

High variability is claimed in the comparability exercise in P.2.2; in method validation

is noted

that variability (% RSD) decreases significantly with sample size (%RSD is 18% for iSOng

(sample size per method) and

7.1%

for 250ng). It should be discussed if the method is

optimized for the intended use and this should be confirmed with comparability results with

commercial scale batches.

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24. In-vitro expression is a cell-based flow cytometry assay. The assay was implemented recently and

the proposed acceptance criteria of

30%

cells positive seem wide compared to the limited batch

release data available to date, i.e. emergency supply lots that is in the tange of 63-65%. Ifl

addition, some data are presented for the small-scale clinical batches used in comparability testing,

where data are found in the tange of 50-71% (Table 3.2.P.2.3-5 in the dossiet). The proposed

acceptafice criteria need to be thoroughiy justified and tightened in line with the levels qualified in

clinical studies or clinically qualifled by other means. This justitication should include the applicant’s

total current knowledge of the drug product.

25. The proposed acceptance criteria of 80% for RNA encapsulation seem wide compared to clinicai

batch data that is found in the tange of 92-94%. The proposed acceptance criteria for RNA

encapsulation should therefore be tightened based on clinical qualification or clinically qualified by

other means and set such that a clinically qualified level is assured throughout the sheif-life of the

drug product.

26. The specification tange of each lipid appears somewhat broad, but the acceptance criteria are

found acceptable. However, to futther strengthen the control strategy given that a fixed molar ratio

of cationic lipid and RNA is critical for LNP formation, acceptance criteria for the molar ratio

NIP

should be included in the specification unless further justified.

27. The method description and validation summary of the rapid sterility test should be provided during

the ptocedure.

28. A specification should be included for free lipids or the applicant should justify that the control

sttategy is sufficient in this regard. In addition, no information and discussion are provided on the

lipid-related impurities originating from the degradation of the lipid nanoparticies and such data

needs to be provided.

29. A risk assessment with respect to the potential presence of elemental impurities in the drug

product based on the general prificiples outlined in Section 5.1 of ICH Q3D should be performed. A

summary of this risk assessment should be submitted. The risk assessment should cover ali

relevant elements and sources in accordance with the guideline. The summary must enable a

quantitative comparison of observed or predicted levels with the PDE:s given in the guideline. It

should contain what is necessary to evaluate the appropriateness and completeness of the risk

assessment, inciuding any assumptions, calculations etc, made. The control strategy for elemental

impurities should be justified based on the risk assessment.

30. The specification for LNP polydispersity index should be tightened in line with batch results for

clinical batches, i.e. NMT 0.2 (0.22 observed ori stability).

31. Detailed description of analytical methods should be provided in P.5.2; these details should be in

line with the validation data:

for ali methods, a list of materials needed for analysis

for the DLS method for particle size and polydispersity, futther details of the instrument and

the sample size

COVID-19 mRNA vaccine (nucleoside modified)

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for the fluorescence assay method: the surfactant and ts concentration, sample and standard

concentration and the tange of the calibration curve.

for the CAD method, the sample diluent.

for the potency ifl vitro by cell based flow cytometry: the Drug Product Control (DPC) (e.g.

qualification), for the flow cytometer acquisition: complete examples of results (including the

three population: P1, P2 and P3) should be provided for NC, DPC and TS samples, and Assay

and Sample acceptance criteria rationale should be explained and justified.

for the RT-PCR method: criteria for the selection of primers used for the test.

32. Validation data for the CGE (RNA integrity) is referred to the drug substance section S.4.3.

However, as the active substance is formulated (RNA is encapsulated in the LNP formula), the

appropriate validation parameters for the drug product (specificity, accuracy, sensitivity,

robustness) should be addressed.

33. Method transfer plan was flot submitted in the RR but is requested to be discussed in the next

submission. For the non-compendial tests, it should be confirmed that the validation acceptance

criteria for the receiving sites will be the same as for the transferring site fwhich will be assessed

during the RR). For the analytical methods where comparative analysis will be proposed, it should

be confirmed that the acceptance criteria will be the same as for the intermediate precision

validated at the transferring site (and assessed during RR).

Reference standards or materials (P.6)

34. It should be clarified for what release and stability testing methods the reference standard

(including the CRM) is used today and will be used in the future. The function of the reference

standard should be briefly stated for each assay, i.e. resuits of evaluation/normalisation, sample

compliance, assay control etc. This information could be provided preferentially in a tabulated

form.

35. Since the Applicant ntends to establish primary and working reference standards, information on

the preparation, qualification and stability of the PRS and WRSs should be provided.

Stability (P.8)

36.

The proposed initial shelf-life for the drug product is 6 months at the recommended storage

temperature of -90 to -60°C. In ordet to support the suggested shelf-life for drug product

updated reports from the ongoing stability studies should be provided.

37.

It should be confirmed that future extensions of the assigned DP shelt life will be appiled for in

formal variation applications. The following statement should be removed for Module 3.2.P.8.1 of

the dossier; “The sponsor will extend the assigned shelt life without notification providing the

COVID-19 mRNA vaccine (nucleoside modified)

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real time stability data at the intended storage condition is acceptable and within commercial

speciNcations.”

38.

Results on photostability testing as well as temperature cycling studies are pending to date and

needs to be provided for assessment.

The applicant should confirm that they commit to continue ali the ongoing stability studies at

long-term conditions until completion.

It should be confirmed that the specifications for the bromobutyl stopper include the tests in the

Ph Eur 3.2.9, including the seif-sealing test, and that the self-sealing test is still acceptable after

the stopper exposure to freezing (down to -90°C) and thawing, since the vial is a multi-dose

container intended for 5 doses.

41.

The applicant needs to clearly define in P.8 and in the future SmPC/PIL in line with available data

and practicai needs:

the sheif-life under recommended, refrigerated, and ambient conditions

the in-use shelf-life and storage conditions after dilution with salme and after first use

a storage condition to keep the vial in outer cafton and protect from light, before and after

dilution (smnce multi-dose container).

39.

40.

Appendices (3. 2.A)

Vital safety

42.

Regarding the Pyrophosphatase, T7 polymerase and RNase inhibitor, spermidine and DNase I

provide a cettificate stating that no product of biological origin has been used during the

manufacture (production and purification) or provide adequate virological documentation, with

regard to viruses and unconventional transmissibie agents (NCTA or prions, compliance with

EMEA/410/01 Rev.3 requirements) where applicable, for each of the components concerned.

43.

Regarding the four lipid excipients: ALC-0315, ALC-0159, DSPC and Cholesterol provide a

cettificate stating that no product of biological origin has been used during the manufacture

(production and purification) or provide adequate virological documentation, with regard to

viruses and unconventional transmissible agents (NCTA or prions, compliance with EMEA/410/01

Rev.3 requirements) where applicable, for each of the components concerned.

Novel excipient

—

ALC-0315

Based on the limited information no final conclusion can be drawn on chemical synthesis, quality

control of starting material, specification limits for impurities and retest period.

44.

45.

The commercial batch size should be provided.

The specification limit for assay (85-115%) is considered wide and should, if possible, be

tightened. The specification limit shouid be re-evaluated as more batch data are available and

COVID-19 mRNA vaccine tnudeoside modified)

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then specification limits for impurities are set, i.e. the mass balance should be taken into

account.

46.

47.

The method description should inciude the GC chromatography parameters.

A brief summary of validation of the GC method is provided. Extended information in form of a

short validation report including relevant data, chromatograms and calculations should be

su bm itted.

48.

It should be confirmed that the packaging materials are conform to Ph Eur or EU regulation

10/2011 amended.

Novel excipient

—

ALC-0159

Based on the limited information no final conclusion can be drawn on chemical synthesis, quality

control of starting material, specification limits for assay impurities and retest period.

49.

50.

51.

52.

The synthesis scheme is illegible, a readable scheme should be provided.

The commercial batch Size should be provided.

The method description should include the GC chromatography parameters.

A brief summary of validation of the GC method is provided. Extended information in form of a

short validation report including relevant data, chromatograms and calculations should be

su bmitted.

53.

54.

A test for molecular weight and polydispersity should be included unless otherwise justified.

It should be confirmed that the packaging materials are conform to Ph Eur or EU regulation

10/2011 amended.

5.2. Non-cilnical aspects

Major objections

None

Other concerns

Pharmacology

Applicant should provide a more detailed ciarification of the mode of action of BNT162b2, e.g.

which cells types will take up the LNP, translate the modRNA and express the S-protein on the

sur-face. Moreover, which cell types/organs will be targeted by the immune defence system, when

the vaccine is in action. Further information on the potential activity/mode of action of the two

novel excipients should be provided. ([confidential information deleted])

COVID- 19 mRNA vaccine (nucleoside modified)

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Ifl study 20-0211, regarding the results obtained from the Western Blot, a semi quantitative

analysis of the resuits to improve the readability of the protein expression should be provided and

in the analysis of the blot, some missing scientific information and explanations should be added

by the applicant ([confidential information deleted]):

a. The presence of the two bands for BNT62b2 ARN (at 100 KDa and 190 KDa respectively)

b. The 76.5 kDa bands are not observed in both BNT162b2 nor in 51 control anes, and the full

5 protein at 141.14 kDa is also flot observed in BNT162b2 lane.

c. The lack of an impoftant expression for the Sl protein ctrl at 76.5 Kda

Regarding the structural and biophysical characterization, the applicant is asked to provide

(fconfidenti& information deleted]):

a. A schematic description of both variants, (V8 and V9) so as to identify the exact position of

optimized codons in the sequence and inciuding coding and non-coding sequences...

b. A comparison between the VS and V9 codon sequeflces, highlighting their differences on m’1JU

and cytosine residues. The exact position of these optimized codons inside the modRNA

sequence should be provided

c. An estimation of mtlJU relative content in both V8 and V9 sequences and a discussion on the

potential difference in immunogenicity between these two variants. Changes in cytosine and

mL1JU content can significantly change the modRNAs immunogenicity.

d. A comparison on the protein expression obtain from both variants (V8 and V9) to ensure that

the expected protein is expressed in non-cilnical models.

4. The modRNA contains a substitution of 1-methyl-pseudouridine for uridine. This substitution

decreases recognitfon of the vaccine RNA by innate immune sensors, such as toll-like receptors

(TLRS) 7 and 8, resulting in decreased innate immune activation and increased protein translation.

Vaccination with modRNA is expected to induce robust neutralising antibodies and a concomitant I

cell response to achieve protective immunity. Nevertheless, no further discussion was provided

regarding the risk of autoimmune responses induced by the modRNA. The Applicant is invited to

fufther discuss the possibility that the mRNA vaccine can trigger potential autoimmune responses

and how do it plan to possibly evaluate their occurrence (Iconfidential information deleted]).

The applicant is requested to provide a more extended discussion on the choice and relevance of

the pharmacological animal models (also with regard to the choice of the rat as a toxicological

animal model) and chosen endpoint in the pharmacological-immunological assessment (eg. lack

of assessment of long-term memory responses, no assessment of old age-dependent effects)

([confidential information de/eted]).

6. Concerning study R-20-0085 on the immunogenicity in mice of the LNP formulated modRNA

encoding the viral S protein (V9):

a. The applicant is asked to justify the absence of IgG2A and IgGi characterization for RBD

(tconfidential information

deleted]);

b. The applicant is asked to justify why the resuits were flot expressed

iii

titers that would also

allowed comparisons across experiments. Indeed, comparison with pyNT experiments

expressing results in titers could help to determine the levels of neutralizing and non

neutralizing antibodies present in the sera ([confidential information deleted]).

c. Ifl the study report R-20-0085 section 4.5.3.1 a discrepancy was found between text (1, 5 or

10 ug/animal) and table of treatment schedule (0.2, i and 5 ug); the Applicant should ciarify

which is the correct piece of information ([confidential information deleted]).

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Concerning the Multiplex analysis of cytokine release from murine Spienocytes Day 28 after

Immunization with BNT62b2, it is referred to immunization with “5” as compared to “1 ug”

8NT162b2 for the Luminex analysis in the Pharmacology written summary, page 18 (last

paragraph) as compared to in the report R-20-0085, respectively, this discrepancy could be

clarified. Moreover, it is noted that high levels of the Thi cytokines IFNy and IL-2 in multiplex

immunoassays were detected after re-stimulation with the 5 but not RED overlapping peptide

mix, although RBD is part of the 5 protein. This could be further ciarified or commented

([confidential information deleted]).

7. Regarding Study VR-VTR-1067i: BNT162b2 (V9) Immunogenicity and Evaluation of Protection

against SARS-CoV-2 Challenge in Rhesus Macaques (tconfidential information deleted]):

a. The applicant needs to precise for the Luminex data how the reference curve has been

constructed, what does represent the arbitrary U/mI used and how it is referring to the serum

dilution factor;

b. The applicant is asked to define the criteria for choosing a 10-30% infection rate of Vero celis

c. Methods to quantify antibody production in the different experiments differ and consequently

cross-comparison between experiments is difficult. Indeed, it is important to distinguish

neutralizing antibodies from non-neutralizing antibodies. In this study, total antibody

response is measured using a luminex assay and results expressed on U/ml and for the

neutralization assay results are expressed in VNT 50. The applicant needs to provide an

estimation of the non-neutralizing antibodies in the whole antibody response.

d. It is important to notice that on figure 6 of study report, neither panel A flor panel B highlight

the consumption ot IgG Sl binding antibodies after challenge nor the increase due to B

memories response following the challenge: this would need to be further discussed by the

Applicant

8. The report VR-MQR-10211, on Sl-binding rhesus macaque serum IgG levels detected by a direct

binding Luminex immunoassay, was flot provided. This should be submitted (tconfidential

information deleted]).

9. The data from the individual animals should be provided for the RT-qPCR test for presence ofSARS

C0V-2 RNA after SARS-CoV-2 Challenge in BNT1G2b2 (V9) immunized nonhuman primates

([confidential information deleted]).

10. In the NHP pharmacology and in the toxicology studies the control group is immunized with PBS

and not with a mRNA in LNP expressing a non-correlated antigen. The Applicant is invited to

further discuss the potential effect of the formulated mRNA on the immune response and toxicity

([confidential information deleted]).

ii. The Spike protein of SARS-CoV-2 undergo mutations, and it thus critically important to investigate

the biological significance of these variants in relation to the development of Spike-based covid-19

vaccine candidates. For example, Korber et al. 2020 present evidence that there are flow more

SARS-CoV-2 viruses circulating in the human population globally that have the G614 form of the

Spike protein versus the D614 form that was originally identified from the first human cases in

Wuhan, China. Further, Li et al., states that as of May 6, 2020, 329 naturally occurring variants in

Spike protein have been reported in the public domain. The applicant is asked to discuss how the

chosen Spike antigen variant in BNT162b2 relates to the Spike variants currently on the dominant

SARS-CoV-2 viruses circulating in the human population ((confidential information deleted]).

mRNA vaccine (nucleoside modified)

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References: Korber et al., 2020: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7332439/ Li et

al., 2020: https://doi.or/10.1016/i.cell.2020.07.012

12. The rhesus macaques were challenged with the SARS-CoV-2 USA-WA1/2020 isolate. To our

knowiedge, this stram does flot contain the D614G mutation. This mutation is reported to rapidly

accumulate in the circulating SARS-CoV-2 strains and may increase the infectivity several-fold

compared to the original Wuhan-1 stram. The applicant is asked to discuss the relevance of the

NHP challenge study resuits in relation to the stram used for challenge and the strains circulating in

the human population ([confidential information deleted]).

13. Overall, the challenge study appears questionable in its design and hardly supports the robustness

of the immunological response. The above limitations can be listed regarding the model:

a) Absence of clinical signs in control and challenged NHP,

b) Use ofjuveniles NHP,

c) Lack of females NHP,

d) One out of three age-matched salme control-immunized (n=3) male rhesus macaques not

responding to challenge (no viral RNA neither in the BAL and nasal swab),

e) Low numbets of animals with a low statistical significance

f) Questionable selection of titer of the vital challenge (1.05.

106

PFU)

g) In the NHP phatmacology study (Study VR-VTR-10671), rhesus macaques were immunized

on days 0 and 21. Some other covid-19 vaccine candidates have diffetent prime-boost

intervals, such as 4 weeks for both ChAdOxl (Graham et al., 2020) and mRNA-1273

(Corbett et al., 2020). Considering that the time between the first and second vaccine dose

may have a significant impact on the immunological response, the applicant is asked to

provide the rationale for the chosen prime-boost interval (21 days)

([confidential

information deleted]). References: Graham et al., 2020:

https://www.ncbi.nlm.nih.gov/rmc/articles/PMC7385486/ Corbett et al., 2020:

htts

:/Iwww.

ncbi .nlm nih.gov/cmc/articles/PMC7449230/

Moreover, some important data are missing to date:

Lung histopathology and immunochemistry, mentioned by the Applicant as ongoing, should

be provided.

Absence of cytokines measurement in the NHP BAL

The applicant is asked to discuss ali these limitations and should provide further scientific information

on the NHP model relevance. Although the model is considered adequate to demonstrate

immunogenicity, and viral clearance, it is considered insufficient to demonstrate efficacy against the

disease ([confidential information deleted]).

Pharmacokinetics

Quantification ofALC-0315 and ALC-0159 in plasma, hver homogenates, urine, and faeces

homogenates was conducted by LC-MS/MS in an in vivo PK study (PF

07302048_063u120_072424). No validation data for the LC-MS/MS method in the non-GLP IV PK

study in rats (Study PF-07302048_06]u120_072424) had been presented. The Applicant is

requested to provide quahification data for this method

([confidential information deleted]).

COVID-19 mRNA vaccine fnucleoside modified)

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CHMP overview and list of questions

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The Applicant is asked to justify the choice of an IV study instead of an IM study in the non-GLP IV

PK study in rats (Study PF-07302048_063u120_072424), which would have a more clinical

relevance. The difference observed ifl terms Gf PK absorption should be discussed

((can

fidentla!

information deleted]).

It’s

worth to notice that the lipid displaying a persistent kinetic over time in iver is ALC-0159, ie

the one that does flot contain any PEG, although PEG is known to be used to increase half-life of

many recombinants. The Applicant will have to justify this observation, as well as to discuss the

difference of kinetics profile between the two lipids. The Applicant is also requested to estimate the

delay of the clearance of the ALC-0315 from the livet, as this could have an impact on the safety

proflie ([confidential information deleted])

The Applicant mentions that the LNP-formulated luciferase-encoding modRNA tested in this study

have the exact same lipid composition than BNT162b2. It is howevet flot cleat to understand which

of the three tested LNP formulation is present in the drug product, BNT162b2 variant V9. The

Applicant should comment. ([confidential information deleted])

There are uncertainties regarding the biodistribution study performed with the surrogate luciferase

reporter RNA. The applicant is therefore asked to provide more information regarding the

biodistribution assay:

a) The applicant is asked to justify and discuss the choice of using a non-validated/non

qualified bioluminescence method to determine the biodistribution of a reporter

luciferase protein instead of detecting the actual BNT162b2 modRNA. The justification

should inciude a discussion on the sensitivity of the method.

(Iconfidential information

dele ted])

b) It can be noted that there is no information on the similarities of the mRNA

modifications of the non-coding regions between the luciferase modRNA used in the

study and the modRNA used in BNT162b2. The applicant is asked to provide more

information on the luciferase reporter RNA, and in particular, whether the untranslated

sequences are similar to that of the BNT162b2 modRNA and therefore at least the

stability of the mRNA5 are somewhat comparable.

([confidential information deleted])

c) The biodistribution of the vaccine has been evaluated in mice, using 2 pg mRNA

(encoding for luciferase). Ifl humans and in the repeat-dose study in rat using the V9

version, 30 ]g (per administration) was used. It is not clear if this difference in RNA

concentration results differences in the amount of LNP used. The applicant is therefore

asked to ciarify if there were differences in the amount of LNP used in the

biodistribution study and the repeat-dose study /clinical trials and if so, discuss how

this could affect the distribution and safety evaluation observed in the clinic compared

to non-cl in ical data. (Iconfidential information deleted])

d) The applicant is asked to consider the possibility of a broader biodistribution pattern

than observed and discuss the possible safety consequences of a wider biodistribution

profile of BNT162b2.

(fconfidential information

de/eted])

Luminex-based multiplex assay:

COVID-19

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In view of potential acute immunotoxicity mediated by LNPs, does the Applicant

possess data on othet timepoints (eariier than 6h or beyond) regarding the cytokines

measurements? ([con fidential information deleted])

b) The Applicant is asked to discuss the absence of an in vitro hPBMC stimulation assay

on healthy donors to assess reactogenicity. ([confidential information deleted])

Extrapolating to clinics, the Applicant is requested to discuss the level of IL-6 cytokines

induced by LNPs considering that asymptomatic but infected subjects candidate to

vaccination, could display higher IL-6 levels during early phase intection. ([confidential

information deleted])

Toxicology

Further discussion is requested an how the immunological response to the vaccine observed in

rats, the species used in the toxicological studies, compares to that observed in Rhesus monkeys,

the species used in the virus chailenge study and, if possible, humans ([confidential information

deleted]).

The qualitative and quantitative compositiofi of the lipids constituting the LNP are flot specified in

the final study report #38166. It is thus flot possible to check the composition of LNP; this point

appears crucial as it is expected that the toxicity associated with modRNA formulated in LNP

formulations is expected to be driven primarily by the LNP composition; This will have to be

specitied ([confidential information deleted]).

Both the “V8” and “V9” repeat-dose toxicity rat studies indicate functional hepatic and/or biliary

effects (enlarged livet, vacuolation, increased gGT, ALT and ALP) which may or may not be linked

to the LNP and which requires further discussion ([confidential information deleted]).

The applicant is requested to provide a discussion an the ciinical relevance of these

findings, and the need for a notation in the SmPC ([confidential information deleted]).

The discussion should include the mechanism underlying the elevated plasma activity

at liver/biliary enzymes and ts potential relation to LNP lipids ([con fidential information

dele ted]).

The discussion should also include the findings of vacuolation at hepatocytes (minimal

to mild) that was present in the portal regions of livet for ali BNT162b2 (V8)-dosed

animals (19 of 20 animals) at the end of the dosing phase ([confidential information

deleted]):

Confirm that the same LNPs composition was used amongst ali treated groups (from a

qualitative and quantitative point of view);

Explain and discuss the difference in vacuolation occutrence between sexes for ali

treated groups as weil as the absence at vacuoles in the V9 study;

Justify the occurrence at vacuolation in hepatocytes, while this effect is usually seen

with phagocytes and nat hepatocytes;

Discuss the shaft delay at occurrence as well as the mechanism underlying these

vacuoies at the time at sacrifice (i.e. only 3 IM weekly injections) (i.e. Development of

anti-PEG antibodies? adaptive response with ar without functional change? imaging to

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determine If vacuoles contain PEG?). Of note, the accumulation in the livet was mainly

observed with ALS-0315 that does flot contain PEG, in conttast of ALC-0159 which

does contain PEG 2000. Discuss their potential toxicity from a non-clinical and clinical

point of view.

In light of the in vivo PK study, showing petsistence of ALC-315 inside livet (slow

elimination kinetics inside livet) and considering the presence of PEG in the formulation

of ALC-0159, the Applicant is requested to document more in depth the

role/implication of ALC-0159 and ALC-315 in the occutrence of vacuolation of pen

po tta I hepatocytes (tconfidential information deleted]).

10. The Applicant will also have to detail and furthet document the elevated serum levels of the

cytokines IFN-gamma, TNF-alpha, IL-ibeta, IL-6, and IL-lO that were noted in the control group of

study #38166 ([confidential information deleted]).

11. Complement (C) activation-related pseudoallergy (CARPA) can be a serious side effect of liposomal

drugs, biologicals, and many other modern therapeutic and diagnostic agents. The Applicant is

asked to discuss the absence of quantitative and targeted assays of C3c and C4 proteins

([confidential information deleted]).

12. The applicant is requested to provide an extended discussion on the distribution and metabolism of

the novel excipient lipids (ALC-0315 and ALC-0159), their potential genotoxicity of the acetamide

moiety in the lipids (which is classified as possible human carcinogen (IARC Group 25) with

debated genotoxic mechanism) in the context of the rat hver observations ([confidential

information deleted]).

13. Some toxicological studies remain to be submitted: the full report for the #20GR142 study and the

interim and full report for the DART study.

The Applicant is also asked to provide at the next NC roll of submission, a detailed

timeline for availability of preliminary data ([confidential information deleted]).

With regard to the DART study, a justification of the study design is requested to

determine the value of this study for evaluation of the developmental risk in humans.

Primarily the choice of the rat as relevant animal species (rodent placental antibody

ttansfer during the latter part of gestation is flot considered similar to human antibody

transfer during the third tnimester of gestation) and the design of the dose regimen

(whether this will lead to sufficient antibody transfer during lactation, which is

equivalent to the third trimester exposure in humans) will fleed attention ([confidential

information deleted]).

14. In terms of GLP compliance, concerns have been raised by the assessors during the review of the

non-clinical report amendment of the study #38166 ([confidential information deleted]).

About the test items and tormulations:

Page 26: The test item EE4 (G7) is desig ned as: LNP formulated modRNA encoding the RED

subunit of SARS-CoV-2 5 protein (BNT162b 2): what is the difference with bl designed by

the same terminology. b2 is flot supported to code the full-length spike S glycoprotein?

BNT162b 2 is also associated with another name in the pathology repont page 1563 (“LNP

modRNA Sp2”). Is it the same test item? The applicant is ask to ciarify the different

terminologies used for the test item EE4.

COVID-19

mRNA vaccine fnucleoside modified)

Quality rolling review CHMP

overview and list of questions

Quality rolling review CHMP overview and list of questons

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

Page 79/81

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bilag 9

Information about stabil ities of the test items during 6 hours at room temperature is flot

documented, whereas the test items were administrated 6 hours after thawing at room

temperature. Could you provide evidence that the test items ate stable for 6 hours at RT.

Calculation of dose concenttations to be administrated to animals are difficult to be understood

for Group7 (BNT162b2 as test item), if 200pI per animal is administrated, the total

concentratian seems to be llOjJg and flot lOQug as stated. The applicant should clarify.

About the management of the study:

A mistake in the conclusion page 61 concerning the sex of animal No.179 having eschar has

been observed. It seems to be a male, and flot a female as stated. And this presence of eschar

was not found in the table for individual clinical signs page 152 for this female No.179. Same

comment for male No.162 (reddened skin repofted page 62) flot found page 138. The

applicant should explain these discrepancies and correct these ssues.

The final report had been amended justified by changes qualified as minor ar/and corrections

following sponsor comments. Some corrections could be considered as nat minor but major,

because they put into relief real mistakes in the issuance of the final report: “eschar formation

was incorrectly described with accurrence an test days 14 and 15 instead of an test day 14

only” ; “an haematology and coagulation the finding of an increased number of eosinophils in

groups 4, 5 and 7 was missing” ; “an clinical chemistry the directions at changes for albumin

and globulin levels were incorrectly stated as an increase in albumin and a decrease in globulin

plasma levels instead af a decrease in albumin and an iflcrease in globulin plasma levels”; “the

incarrect test item ‘BNT162b1’ instead at BNT162a1’ was stated for Group 3”.

5.3. Cilnical aspects

N/A

5.4. Risk management plan

N/A

55.

Pharmacovigilance system

N/A

5.6. New active substance status

N/A

6. Recommended conditions for future marketing

authorisation and product ïnformation in case of a posïtive

benefit risk assessment

N/A

COVID- 19 mRNA vaccine fnucleoside modified)

Quality rolling review CHMP overview and list of questions

EMA/CHMP/641856/2O2OEMA/CHMP/641856/2020

Page 80/81

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bilag 9

7. Appendices (as appropriate)

N/A

COVID-19 mRNA vaccine (nudeoside modified)

Quality rolling review CKMP overview and list of questions

Quality rolling review CHMP overview and list of questions

EMA/CHMP1641856/2OZOEMA/CHMP/641856/ 2020

Page 81/81

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 12

loksiske Pfizer vaccinebatches

af Max Schmeling, Statistiker

Økonom, Cand.merc.(Iog)

Li5te over Toksiske Pfizer Covid-19 vaccinebatches (opdateret: 12-12-2021)

Batchkode

EK9788

ER1741

EM0477

EW4109

E]6797

ER1749

EP9605

ET8885

EW3143

EL14$4

FA1O27

Ff3319

EJ6134

EM4965

EJ6136

EP2163

EJ6788

EW4811

EW2239

EJ6790

EP9598

ER2659

EJ5789

ET3620

FD6840

EW6126

EJ6795

EP2166

EL0739

ET1$31

EJ6796

FE6208

Antal

bivirkninger

pr. batch

Batchkode

2410

EE8493

2088

EX8679

188$

EW8904

1864

1829

1719

1501

1481

1463

1419

1212

1162

1161

1157

1106

1099

1089

1087

1081

1064

1056

1024

996

992

990

989

988

985

968

963

EK4243

EK4244

e18723

EM6950

FC0681

EX3599

FE1573

FD5996

FC309$

FE151O

E18713

FC5089

FC1526

FA5833

E]1688

EL0141

EW2245

FF3318

EX3617

EX3510

FC5435

EY3014

EY4834

Fe8087

ET3045

EY5420

FA4597

FE8244

FC8889

Antal

bivirkninger

pt. batch

Batchkode

589

FE6975

582

FC8289

575

570

566

564

560

558

531

524

521

519

515

507

493

491

489

473

472

471

466

461

456

450

445

435

429

428

427

426

FE2090

FA2453

FF0900

FF2752

EX2405

FA7338

FE3065

FE$405

FE4728

FD9309

FG7387

FÉ7053

ER9449

FC3661

FC1131

FE7OS1

EK4176

EY5423

FA5843

FC5295

FC8736

FA5715

EW6326

FG4686

FC1440

Fh3220

Fi5782

FG4442

Fg3712

EY3860

Antal

bivirkninger

pr. batch

Batchkode

279

FK0596

278

Éi0724

278

EW0203

276

FF3620

273

249

247

244

243

240

237

236

230

229

228

227

221

21$

214

207

203

200

199

198

197

196

195

EN1194

EW6327

EK4237

EY7065

FD0350

FE8206

FE4721

FG4509

FD0927

FE8162

FF2382

EY0572

FG3716

EW0193

EW0221

EY0583

EW0199

FG0978

FHO161

FC9880

FHO151

FF9944

FC5436

FF9942

FE6029

FE3064

FG6270

F15790

Antal

bivirkninger

pr. batch

126

125

124

121

119

117

116

114

113

111

110

105

104

101

1 af 2

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

biag 12

Batchkode

EW2246

FE3380

ER9480

ET3674

FD0168

FD4555

EK1768

ER7449

EL0725

EW4815

ET6956

ER9470

EX0893

E17205

FD8813

FD1921

EX6537

EL1491

EY2173

ÉN1185

ER7934

EN3924

FA4598

FC9001

FA8016

EE8492

FA5831

EX7389

EX8680

ER7812

FF0680

FC2336

fd5613

Fc3143

EW9127

EX7823

Antal

bivirkninger

pt. batch

963

945

941

928

894

889

845

837

828

825

824

821

812

806

793

776

750

727

722

712

711

704

698

684

682

651

649

641

632

628

622

612

604

596

594

Batchkode

FF8222

FF8288

FE2625

FF0688

FD0785

E10553

FD9234

FE7O1O

FC3095

FD7958

FC2473

FA4632

EY2172

EY0779

FF4213

FD7959

FE2707

FA7082

FE2296

EJ3002

FC3558

FA7812

ET9096

EY7015

FE2083

EL7834

FE1248

EY5456

FE9174

FG4493

FA5829

FF2153

EW2243

FE8235

FG6273

FA5765

Antal

bivirkninger

pr. batch

424

422

419

410

404

403

400

387

386

379

378

376

373

372

371

358

357

354

350

342

340

330

327

322

312

309

306

302

290

288

Batchkode

EP6775

FG6431

FC1439

FF2834

FD8274

FC1433

FC5029

Fg3739

EY5422

FD0932

ER0641

EWO2O1

FE7O11

FF7481

FA8142

FC5947

EY0573

FC6984

FF3622

FC6997

FF2832

fh8469

EW0207

FC2229

FD0889

FA7083

FC1435

FF0843

ER1742

FF2782

EX6564

FHO114

FF4204

Fk0112

FC1436

EW5279

Antal

bivirkninger

pt. batch

186

184

183

179

178

177

172

169

167

166

162

161

156

151

147

145

142

141

140

139

135

133

131

130

127

Batchkode

Ek4241

fg9019

EK4245

FD0348

FD1945

EY0578

FF2018

FA7842

FJ7489

ET6924

EL1406

EN1198

FC2495

FA7478

FD7204

FA9091

FF5357

FH3023

FG7911

FC9909

Ew0206

EW0195

FF8871

Fd7206

EW3344

FA8721

EY0579

EP6017

Fg7898

FA9090

FKO1O8

EL0203

EX0438

Antal

bivirkninger

pt. batch

2 af2

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 13

Overdødelighed 2027

Samlet antal dødsfald i Danmark pr. måned

5800

-o

5600

5400

5200

5000

4800

4600

4400

4200

4000

3800

maj

jun

jul

aug

2021

2020

sep

okt

nov

dec

jan

feb

mat

apt

2007-19

Gennemsnit. 2007-21

Kilde: Danmarks Statistik DODC11.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 14

Guide til reproduktion af resultaterne.

Download datasættene: VAERS DATA (2O2VAERSData.csv)og VAERS Vaccine (2O21VAERSVAX.csv) fra

hjemmesiden: https://vaers.hhs.gov/data/datasets.html? Som er the Vaccine Adverse Event Reporting System

(VAERS).

•

Lav en Excel

og åbn denne. Det er nødvendigt, at anvende en 64 bits udgave af Excel grundet

datasættenes størrelse.

Anvend menuen Data->Get Data

From text/csv og vælg første fil 2D21VAERSData.csv. Tryk derefter

Load. Dataene bliver oprettet som en tabel, der har en forbindelse/”connection” til csv filen.

Gentag ovenstående punkt for filen 2O21VAERSVAX.csv

Begge datasæt en nu importeret i Excel.

Anvend igen menuen Data->Data tools->Relationships. Anvend dialogboksen til at lave en ny relation

mellem de to tabeller, der laver en relation på variablen VAERS ID fra begge tabeller.

Anvend menuen lnsert->PivotTable->From DataModel

I den oprettede pivot tabel indsættes VAX_TYPE og VAX_MANU under Filters, VAX LOT i Rows og

VAX LOT under Values.

Under filtre vælges for VAX_TYPE værdien COVID19 og for VAX_MANU vælges Moderne og/eller

P F IZE R/B IC NTECH

Ved at sortere på Values fås de to første grafer, der viser fordeling. Ved at sortere på Row Labels vises

systematikken.

Datafilerne kan også sammenkøres i de fleste statistikpakker, ved at matche på variablen VAERS_ID i

datafilerne.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 15

Åbent brev til:

Folketinget, Epidemiudvalget, Epidemikommissionen,

Sundhedsministeren, Statsministeren, Sundhedsstyrelsen og

Statens Serum Institut

Systematiske forskelle i antal bivirkninger mellem Covid-19

vaccinebatches på op til 5021 gange.

af Max Schmeling, Statistiker

Økonom, Cand.merc.(log)

Jeg henvender mig til dem angående et yderst foruroligende fund afforskelle i antallet af bivirkninger mellem

Covid-19 vaccinebatches. Fundene viser at det er op til en faktor 3648 i forskelle mellem antallet af bivirkninger

i de enkelte Covid-19 vaccinebatches fra Pfizer. For Modetna er der tale om op til en faktor 5021. Selv om at

dette fund i sin alvorlighed skulle synes slemt nok, er situationen langt værre, da det i denne forskellighed i

bivirkninger ses en systematik relateret til selve batchnumrene.

Indledning

Jeg arbejder som selvstændig uafhængig økonom og statistisk rådgiver, typisk for analysevirksomheder og

mediebureauer og større virksomheder. Disse resultater er kommet til mit kendskab gennem Craig Kooper

som er en engelsk, researcher ved Kingston University, London. Fordi implikationerne af resultaterne netop er

så alvorlige som er tilfældet, har jeg selv reproduceret og verificeret resultaterne og er kommet til samme

konklusion. Vigtigheden at disse resultater kan næppe underdrives, hvilket er grunden til min henvendelse.

Resultater i det følgende er udarbejdet af undertegnede.

Dataene, der analyseres, er fra den Amerikanske bivirkningsdatabase VAERS’. Dette fordi at den europæiske

bivirkningsdatabase EudraVigilance ikke umiddelbart rapporterer vaccinebatch numre. VAERS databasen er

ligesom det danske og europæiske bivirkningsrapporteringssystem begrænset på flere måder. De væsentligste

begrænsninger er at systemet er passivt, således at bivirkninger ikke automatisk indsamles, men kræver at en

sundhedsfaglig person foretager indberetningen, hvilket leder til andet problem, som er en stor grad at

underrapportering. Ingen af disse begrænsninger kan dog antages at påvirke denne analyses resultater

nævneværdigt da den bias, begrænsningerne giver må antages at være ens for alle vaccinebatches. Det største

problem ved analysen synes faktisk at være at datakvaliteten for vaccinebatchnummeret er påvirket af at

indrapporteringen af nummeret sker som fritekst og ikke prædefineret. Dette giver noget støj grundet

tastefejl. Alligevel er resultaterne tydelige nok til understøtte de viste resultater. Analysens metode er

beskrevet i bilag 1, således at denne let kan reproduceres.

‘

https://vaers.h hs.gov/data/datasets.html?

af 6

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 15

Bivirkninger pr. vaccinebatch

Nedenfor ses antallet af bivirkninger pr. vaccinebatch og sorteret efter antal

Moderna

6000

antal bivirkninger pr. batch

sooo.

4000

3000

1-4

—

(1-1-41-41-41-4

<ni

n —

1-4

I-

0 0 t 0

-4

—

1-4fl4

,-4

-..

-I

-4

—I 1-)

..J_J0

—

1-41-I

—

,-I

—

‘-‘

1-1

Pfizer

antal bivirkninger pt. batch

4000

35002

3000

2500

2000

1500

1000

500

ni ni

ni CO

ifl

0 CO ni NI

1-I

‘

,-4

0 0

ni_I

I.-

-1

0 ni 0

—

0 ni

—

CO NI 1-4 00 0

0 00

Lfl

ifl

Som det ses at graferne, er resultaterne ekstremt skævt fordelt. For langt de fleste batches, Pfizer: 95,9%

Moderna: 95,6% ses kun op til 10 bivirkninger, men for et mindretal at batchene Pfizer: 4,1% Moderna: 4,4%

ses op til Pfizer: 3648, Moderna: 5021 bivirkninger. Det kan altså konstateres, at et mindretal af

vaccinebatchene producerer op til 5021 gange flere bivirkninger end hvad man kan kalde baseline.

Konsekvenserne af dette resultat er helt åbenbar. Det et umuligt at give informeret samtykke ifm. vaccination

mod

COVID-19, da risikoen ganske enkelt ikke kan kendes. Dermed bliver vaccination mod COVID-19 jfr.

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 15

sundhedsloven kapitel 5, §15 formentlig ulovlig. Samtidig vil vaccination under disse forhold stride imod

Nürnberg kodekset, specielt punkt 1.

Systematik i forhold til Vaccinebatchnumre

Det antages i det følgende, at vaccinebatchnumre er genereret i en fortløbende nummereret sekvens,

indeholdende både bogstaver og tal. Den for Pfizer mest forekommende syntaks er formen AB1234, som

består af to bogstaver efterfulgt af 4 tal cifre. Dette er ikke fuldstændigt entydigt i data, bla. grundet

ovennævnte begrænsninger, men det er tydeligt at denne syntaks er den normale form for batchnummer for

Pfizer vaccinen.

I figuren nedenfor, er antallet af bivirkninger pr. batchnummer sorteret alphanumerisk ift., batchnummer

meget varierende. Dataene er de samme som ovenfor, med den forskel, at alle batches, som ikke overholder

syntaksen ovenfor er fjernet. Dette bevirker, at næsten alle fejl indtastninger fjernes, selv om noget reel data

også forsvinder. Billedet er helt identisk uanset om man renser data eller ej, men bliver lidt mere overskueligt

ift. farverne i nedenstående graf.

Da dataene er identiske ft. tidligere grafer er det alene er relevant at se på systematikken. Farvelægningen er

lavet for at adskille de største batchserier ift. bogstavskode. Datapunkterne for de enkelte batches er eks.

orange for alle EK#### batches, blå for alle EL#### batches, cyan for alle EN#### batches, osv. Som eksempel

kan EN serien (cyan) anvendes. Denne serie består af 320 batches startende med EN0101 og sluttende med

EN9899. I EN serien ses 12 batches, der hver har over 2000 bivirkninger pr. batch og for 11 af disse er

batchnumrene fortløbende nummereret fra EN6198 EN6208. Det er helt tydeligt, at vaccinebatches med

meget høje antal bivirkninger forekommer som isolerede klynger af batches, der er en del af en større

batchserie.

—

Pfizer

—

Antal bivirkninger

Antal bivirkninger pr. vaccine batch sorteret efter batchnummer

4000

3500

3000

2500

2000

1500

1000

500

— — r,,

r’

—

r-.

.—l

-J

r-.

tO tO

-I

—

‘

—

0 0

0000

—1

Dette skaber et tidsrum mellem hver af disse klynger hvor der kun ses vaccinebatches med meget få

bivirkninger. Samtidigt ses der, startende med batchserien EN fcyan), klynger af batches med mange

bivirkninger, men som klynge for klynge er faldende i niveau. Dette mønster er tydeligt relateret til

batchnumrene og dermed kan det konstateres at der er en systematik i fordelingen af batches med

EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 15

overnormalt antal bivirkninger set i forhold til batchnummeret. Der ses en lignende systematik for Moderna

vaccinerne, men denne er ikke bragt her, da Pfizer vaccinen er langt den dominerende vaccine i Danmark.

Alvorligheden af dette resultat kan ikke understreges nok, da det skaber et plausibelt argument for at mønstret

repræsenterer et forsøg. Den faldende tendens kan ikke forklares ved at man måske skulle forsøge at justere

på visse indholdsstoffer for at mindske bivirkninger, da baseline jo viser at det er det, der produceres i 96% af

tilfældene. Ej heller kan tendensen forklares som fejlagtige batches grundet mønstret. Tendensen kunne

indikere et dosseringsfotsøg, selv om at dette selvfølgelig ikke er muligt at bevise ud fra data. Skulle dette være

tilfældet foregår der altså en form for forsøg bag ved vaccineprogrammerne. Så utænkeligt som dette

forekommet ser jeg ikke umiddelbart andre forklaringer.

Hvorfor skulle bestemte tætte klynger af vaccinebatches producere op til 3648 gange flere bivirkninger end

hovedparten af batches i samme batchserie, samt være organiseret i et blokmønster med faldende antal

bivirkninger mellem klyngerne? Tilstedeværelsen af mønstret udelukker tilfældighed som forklaring og hvis

det ikke er tilfældigt, så er der en årsag.

Den umiddelbare indvendig mod resultatet vil utvivlsomt og naturligt være, at resultaterne ikke er

repræsentative for Danmark, da VAERS registrerer bivirkninger i USA, men denne kritik kan uden videre

tilsidesættes af flere årsager. For det første er Pfizer vaccinen af samme formulering i både USA og EU, så

resultaterne må alene ud fra denne observation være overførbare til Danmark.

Hertil skal tilføjes, at det fremgår aftabel DODC1 fra Danmarks Statistik, at der siden 1. maj 2021 har været en

uforklarlig, statistisk signifikant og i forhold ti! de sidste 15 år, rekordsættende overdødelighed på 2928

personer opgjort måned for måned. Sandsynligheden for denne overdødelighed skulle være et tilfældigt

udsving er 0,00000000001%. Der findes dermed et bestyrket mistankegrundlag for at resultaterne fra VAERS

også vil kunne findes i de danske bivirkningsdata og man bliver derfor, hvis ikke andet lægges til grund, nødt

til at antage at dette er rent faktisk er tilfældet.

Jeg skal derfor indtrængende anmode dem om straks af verificere disse resultater, samt undersøge om

tilsvarende resultater findes for det Danske CO VI D-19 vaccinationsprogram.

Jeg er naturligvis klar over hvilke implikationer dette vil kunne tænkes at få, samt hvor absurd det må

forekomme, at forestille sig at skulle stoppe COVID-19 vaccineprogrammet på nuværende tidspunkt. Men

forestil dem alternativet. Stil dem selv spørgsmålet om de ville føle dem tryg ved at lade deres eget barn

vaccinere? Hvad skal forældre hvis børn ender med svære skader tænke? Hvad ville de selv tænke?

Vi ser samtidig både i Danmark og på verdensplan et hastigt stigende pres på den gruppe af borgere, som ikke

har ladet sig vaccinere. I Danmark har statsministeren endda sat sig selv i spidsen for dette pres, mens den

danske stat har indgået kontrakter med Pfizer og Moderna der frataget disse for ethvert ansvar. Hvem vil tage

ansvaret? Vil de selv tage ansvaret? Hvad vil borgere, der er blevet presset til vaccination med skade til følge

gøre?

Personligt finder jeg resultaterne yderst forfærdende, idet at de kan underbygge en opfattelse om at

vaccinerne ikke bliver produceret eller givet med et beneficialt formål.

Jeg håber at have leveret tilstrækkelig information til at understrege alvorligheden af denne henvendelse og

jeg håber at de vil tage de rigtige valg og iværksætte de relevante initiativer i denne situation idet jeg henviser

til de konventioner, der i mange år har sikret vores rettigheder i forhold medicinske behandlinger.

Aarhus d.14.12 2021

Max Schmeling

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EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

bilag 15

Bilag i

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Guide til reproduktion af resultaterne.

Download datasættene: VAERS DATA (2O21VAERSData.csv)og VAERS Vaccine (2O21VAERSVAX.csv) fra

hjemmesiden: https://vaers.hhs.gov/data/datasets.html? Som er the Vaccine Adverse Event Reporting System

(VAERS).

•

Lav en Excel

og åbn denne. Det er nødvendigt, at anvende en 64 bits udgave af Excel grundet

datasættenes størrelse.

Anvend menuen Data->Get Data

From text/csv og vælg første fil 2O21VAERSData.csv. Tryk derefter

Load. Dataene bliver oprettet som en tabel, der har en forbindelse/”connection” til csv filen.

Gentag ovenstående punkt for filen 2O2VAERSVAX.csv

Begge datasæt en nu importeret i Excel.

Anvend igen menuen Data->Data tools->Relationships. Anvend dialogboksen til at lave en ny relation

mellem de to tabeller, der laver en relation på variablen VAERS_ID fra begge tabeller.

Anvend menuen lnsert->Pivotlable->From DataModel

I den oprettede pivot tabel indsættes VAX_TYPE og VAX_MANU under Filters, VAX LOT i Rows og

VAX LOT under Values.

Under filtre vælges for VAX_TYPE værdien COVID19 og for VAX_MANU vælges Moderne og/eller

PFIZER/BIONTECH

Ved at sortere på Values fås de to første grafer, der viser fordeling. Ved at sortere på Row Labels vises

systematikken.

Datafilerne kan også sammenkøres i de fleste statistikpakker, ved at matche på variablen VAERS_ID i

datafilerne.

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EPI, Alm.del - 2021-22 - Bilag 322: Henvendelse af 18/1-2022 fra Max Schmeling om sammenhæng mellem Covid-19 vaccineskader og bivirkninger for Covid-19 vaccinebatches.

Bilag 2

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Samlet antal dødsfald i Danmark pr. måned

Samlet antal dødsfald Danmark pr. måned

5800

Lå,

5400

5200

5000

4800

4600

4400

4200

4000

3800

maj

2020

—2021

jan

jul

feb

mat

apr

jun

aug

sep

okt

nov

dec

2007-19

Gennemsnit. 2007-21

Kilde: Danmarks Statistik DODC12.

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