Accelerating drug

development to FTIH: Potential

of new expression technologies

Lekan Daramola

Associate Director

Biopharmaceutical Development, Cell

Culture & Fermentation Sciences

CMC Strategy Forum Europe 2014

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Outline

Introduction

Potential strategies to accelerate FTIH

Can we use Transient expression platform?

– Pros and Cons

– MedImmune’s proprietary CHO transient system

Conclusion

Introduction

Conventional method for producing biologics for IND is based on

stable cell line expression platforms

Cell line development is typically time consuming (6 - 12 months) and

resource intensive

– Impact on accelerating drug candidates to IND

Clonal stable cell lines used to produce biologic for IND for ~20 years

despite significant improvements in other protein expression

technologies

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Stable Cell Line Based Drug Development –

Typical Critical Path

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Extensive Screening, Bulk up

Parental Cell lines

Gene optimisation, Vector construction

Transfection

Extensive Screening, Bulk up

Clonal Cell lines Cell Banking

Cell bank Characterisation

Phenotypic Characterisation

Genetic Characterisation

Product Quality

CLD timeline range from 6 – 12 months

Choose Manufacturing cell line

0

2

4

6

8

Clone 1

Clone 2

Clone 3

Clone 4

Bioreactor process

development, Scale up GMP Production

Cell Bank Safety

testing

Potential strategies to shorten drug development

timelines

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Non-Clonal transfected Cell

lines (Parental)

Stable pools (transfected cells) Plant protein

expression

Synthon

Microbial expression

Cell-free protein expression Transient expression

Baculovirus expression

Protein Sciences - http://www.genengnews.com/gen-articles/betting-on-baculovirus-expression/4776/

Transient vs. Stable Expression Systems

Transgene DNA integrated into host genome

Expression stably maintained over many months

Transgene DNA passed to progeny

Gram-amounts but time-consuming

Recombinant cell lines can be stored frozen

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Nucleus

Genomic

Integrated

Plasmid

DNA plasmid

Transfection STABLE

Nucleus

Genomic

Plasmids

Transfect in many copies of plasmid

Expression over a few days/weeks

Transcription without integration into host genome

Repeat transfection for repeat protein batches

Rapid for µg to gram amounts

TRANSIENT

Why consider Transient system now?

Low expression level no longer an issue

– Higher titres – up to 2g/L reported

Successful use of robust, industry standard cell line for transient expression

– CHO cell line

Same cell host as stable cell line can now be used for Transient expression

– Production process transferable from stable to transient

Scalable process

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Expression Strategy

CHO Stable cell line

multi gram amounts

CHO pool > gram

Transient gene expression –small amounts

Lead

Profiling Pre-clinical

Development

Lead

Isolation Lead

Optimisation Target

Evaluation

Pre-

project

Target

validation Clinical

Development

Transient gene expression – potential >gram amounts

Can Transient expression be used for GMP production?

Transient Expression Production Process:

What it could look like?

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Fully characterised cells and transfected with high quality DNA

Scaled up according to drug requirements

Transfection

(Chemical or

Electroporation)

Grams – Kg

of protein

DNA – GMP/non-

GMP grade

Cells – qualified host

cell bank

Transfection Production Harvest

Cell Bulk up

Day1 Up to Day20

Benefits: Transient Material and FTIH

Speed

– Potentially the fastest route to FTIH

• No cell line development required

• DNA purification and production phase

Increase in throughput

– Potential increase in capacity as CLD is no longer on critical

path?

• Organisational

Reduced Cost

– Projects with increase rate of attrition – speed

– Postpone resource intensive CLD to later clinical stages

– Potentially more cost effective to FTIH

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Other Benefits of Using Transient Material

Only one cell bank is required – transient host cell line

– Same host cell line transfected with different DNA

Only the host cell will require safety and genetic

characterisation

Wealth of information and experience from cell, gene

therapy and vaccine production e.g. GMP DNA

production, DNA stability, transfections

Ideal for personalised medicine, low dose indications,

orphan drugs

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www.microscopyu.com/galleries/

http://www.nature.com/news/2007/071127/full/news.2007.291.html

Personalise

d Medicine http://bizj.us/cnoqv/i/1 http://scienceprogress.org/2009/09/personalized-medicine/

Concerns

Low expression in transient systems

Process requirements: different to stable lines?

– Process consistency, robustness,

– Comparability risks

• Comparability with stable cell lines - later clinical stages

– Process development – upstream and downstream

Scalability

Process contaminants

– Transfection reagent

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Precedence

Vaccine Production using transient transfection of HEK293 cells for

clinical studies – NIH

– FDA approved VLP production (influenza) for Phase 1 studies

– Working on scale up for commercial production

Flu vaccine commercial production using baculovirus vector and

insect cells - transient expression approach

– FluBlok® the first FDA approved recombinant trivalent hemagglutinin

vaccine.

Cell/Gene therapy

– viral transduction, electroporation

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MedImmune Proprietary CHO Transient System

MedImmune has developed an industry leading proprietary CHO

transient expression system

– Highest titres reported/published in the industry (2g/L IgG)

– Scalable, easy to use transfection process

Successfully used to express several IgGs and non-IgGs for preclinical

studies over many years

Improved productivity over the years as a result of continuous process

development effort

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Transfection Process

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Simple, scalable transfection protocol

No media exchange required pre or post transfection

High yielding process, highest to-date 2g/L of IgG i.e. potentially up to 40g

crude from a 20L transfection culture

Examples of >1g/L IgG titre from CHO transient platform

Successfully used to fast-track research grade material production for

numerous projects over the past 7 years

– Scaled up to 250L

Product Quality

Robust process will produce consistent product quality

Literature and in-house data suggest IgG product quality is

comparable to material from stable cell lines

– Glycan data suggest transient material is comparable with stable cell

line material.

– Within the micro-heterogeneity expected in stables e.g clone to clone

micro-heterogeneity.

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Conclusion

Conventional stable cell generation can be time consuming

Alternative expression strategies exist to accelerate timelines to

FTIH

– Pools

– Cell-free expression system

– Different expression platforms

Propose that an high yielding scalable CHO transient system is a

potential viable option to accelerate timelines to FTIH

– Already in use for viral vaccine production e.g. viral recombinant

protein, Virus-Like Particles

Pros and Cons

Use of Transient expression for manufacturing is now technically

feasible

– Increase awareness

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Acknowledgements

Early Expression and Supply

Cell Culture and Fermentation Sciences (CCFS) Group,

MedImmune

Ray Field

Paul Varley

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