Accelerated Path to Probe Biology through Deferred Cloning and Applying Platform Manufacturing Processes Rohini Deshpande, Ph.D.,

Executive Director Drug Substance Development, Amgen Thousand Oaks, CA

CMC Strategy Forum Europe May 7th, 2014

We need a product development strategy that meets the needs of a Tough Industry Environment

•Pressures on health care budgets

•Heightened payer focus on value

•Declining R&D productivity

•Global regulatory expectations

• Lengthy timelines for product development

•High cost to develop drugs due to low success rate

Objectives of Accelerated Process Development

• Reach ‘go/no go’ decisions early by probing biology in humans

• Enhance first-to-market probability

• Increase pipeline capacity

• Generate representative material efficiently

• Allow phase appropriate PD and Reg CMC investment

Can we shorten cycle time by moving to a dual cycle approach for cell line development?

Legacy Process: Opportunity for Phase Appropriate Approach

Molecule Assessment

FIH process development

Clinical Production

Phase I/II studies

Commercial Process

Development

Clinical Production

Pivotal studies

IND

Project Strategy Team

INDa

Cell Bank 1 and Process 1 Cell Bank 1 and Process 2

•Dual cycle investment for drug substance process development (P1 for FIH to P2 for commercial) •Single cycle approach for cell line development (commercial ready cell lines at FIH – opportunity?)

Generating phase appropriate clinical material earlier: Dual Cycle Cell Line Development

Transfections, selection and amplification

Cell Pools

Single cell cloning

Final clone selection

Toxicology and early phase

clinical studies

Pivotal studies and commercialization

Cell bank 1

Cell bank 2

Deferred Cloning

A

B

CA

B

C

A

B

CA

B

C

D

E

A

B

CA

B

CA

B

CA

B

C

A

B

CA

B

CA

B

CA

B

CA

B

CA

B

C

D

E

Compose and test

Cell line and

process selection

Purification

process definition

Database In silico designs

Process Dev. Platform and High Throughput Tools

Candidate selection to FIH: platform process for representative material

Platform verification/set point definition & supply

For Internal Use Only. Amgen Confidential. 7

Deferred Cloning and platform application reduces time to IND by ~8 months

Phase appropriate approach saves time for Tox and FIH material supply

Molecule, methods, process, and

formulation

Cell bank 1 and DS

manufacturing

Lead

Molecule selection

Cell bank1 and DS

manufacturing

Lead

Methods, process, and

formulation

Accelerated approach

Traditional approach

For Internal Use Only. Amgen Confidential. 8

Deferred Cloning & Comparability

•Is the product quality data for material generated from cell pools and clones comparable? •Does it pose a risk to comparability?

Phase I/II studies

CPD Clinical

Prod Pivotal studies

Molecule, process,

formulation, and methods

Tox and Clinical

Prod

IND PST

INDa

Accelerated approach

Process 1 and Cell Bank 1 • Material generated

from cell pools

Process 2 and Cell Bank 2 • Material generated

from clonal cell lines

For Internal Use Only. Amgen Confidential. 9

Cell bank 1 from cell pools: Measures for fit-for-use

•Plasmid quality is controlled and sequencing performed on single isolates •Multiple transfections are performed to ensure many parental cell pool options •Product quality attributes are tested using sensitive LC/MS/MS methods to detect microheterogeneity and confirm primary sequence •Product consistency is achieved by using defined population doublings for each run (one vial thaw/run) •ICH guidelines are followed for cell bank genetic characterization (DNA sequencing), release testing, and adventitious agent testing

Cell Bank 1 created from a cell pool will follow the same testing paradigm (no change is proposed)

Purpose Tests CB1

Identity Isoenzyme analysis X

Safety Sterility X

Mycoplasma X

In vitro viral assay X

In vivo viral assay X

Transmission electron microscopy of cells X

Mouse antibody production X

Hamster antibody production X

Cocultivation with reverse transcriptase and focus endpoints for retrovirus

detection

X

In vitro assay for bovine viruses* X

In vitro assay for porcine viruses* X

Genetic

stability

cDNA sequence confirmation X**

Expression system is designed to generate stable cell pools & clones

• Antibody and selection marker are produced as a single mRNA

• Only cells with functional selection marker survive • Provides a tight linkage between antibody light and heavy

chain genes and the survival gene i.e. they exist as part of the same mRNA

• Leads to the creation of highly productive and

homogeneous stable pools which can be used for large-scale production

Cell Pool Productivity & Homogeneity

*Flow cytometry Assay

GMD_30min.fcs

APC-A

Count

100

101

102

103

104

0

1

3

4

5

Non-expressing population

Expressing Cells

Freq

uen

cy

Antibody Secretion*

HC+LC

Amgen Expression System Alternate Expression System

LC

HC

APC-A

Count

100

101

102

103

104

0

16

32

47

63

Cell pool

Antibody Secretion*

Freq

uen

cy

Mock pool

Homogeneity of Cell Pools is an Enabler for the Accelerated Approach

Antibody Secretion

Similar FACS profile between cell pool and clones

Grey- Pool Red- Clone 1 Blue- Clone 2

Efficient generation of tox and GMP supply

0

4

8

12

16

GITR BAFF IL2-Mutein

(g

/L)

Titer

0

200

400

600

800

0 2 4 6 8 10 12 14 16 Culture Duration (Days)

VCD

High titers from cell pools allows use of a single lot for tox and FIH supply

Cell pools

•Charge variant

•Clipping

•Identity

•Glycans

•Chemical modification

•Disulfide integrity Total Ion Chromatogram

Extracted Ion Chromatograms

Modality independent platform analytical technology

Digest

Proteins

Analytical methodologies allow detection of sequence variants from cell pools

Multi-attribute method

• Change from profile to attributes • Utilize Orbitrap MS

• Detect low level protein micro-heterogeneity

• Quantify desired attributes • Detect Unknowns • Fully automated

• Suitable for product disposition

• Replaces conventional methods

• Fewer DS/DP release/stability testing

No sequence variants were detected in 3 pilot programs from material produced using cell pools

Cell pools & clones: similar PQA ranges %

a-f

uco

syla

ted

%

HM

W

% H

igh

Man

no

se

% G

alac

tosy

lati

on

(A

2G

1F)

Cell Pools Clones Cell Pools Clones

Cell Pools Clones Cell Pools Clones

Over 20 cell pools are generated for diversity and over hundred clones are screened to ensure comparability

Molecule 1: Comparable PQAs for material from cell pools & clones

Assay Attribute Cell pool Clone FIH Spec (DS/DP)

Purity

Size Exclusion

HMW 1.3 0.8 < 5%

Ion-exchange

Acidic 26.4 17.3

± 10% Main 60.4 76

Basic 13.1 6.6

rCE-SDS

LC+HC 97.3 97.3 ≥ 95%

LMW 0.1 0.1 N/A

NGHC 2.2 2.3 N/A

Glycan Afucosylation 2.5 1.6

N/A High Mannose 8.52 5.7

Impurity CHOP ng/mL 42 17 < 200 ppm

DNA pg/mg N/A < 0.2 10 ng/dose

Activity Bioassay Potency 75 88 60-140%

*Difference in CEX acid and main is from change in harvest operation

Molecule 1: Comparable Product Quality for material from cell pools & clones

AU

0.00

0.10

0.20

0.30

0.40

0.50

0.60

0.70

0.80

0.90

1.00

1.10

1.20

1.30

1.40

1.50

1.60

1.70

1.80

1.90

2.00

Minutes

10.00 15.00 20.00 25.00 30.00 35.00 40.00 45.00 50.00 55.00 60.00 65.00 70.00 75.00 80.00 85.00 90.00 95.00 100.00 105.00 110.00 115.00 120.00 125.00 130.00 135.00 140.00 145.00 150.00 155.00 160.00 165.00

Cell pool clone

• Peaks aligned for visualization • No significant new peaks or peak changes. • Slight differences in missed cleavage, minor PTMs, and glycosylation

Assay Attribute Amplified pool

(3-150) Clone*1 Clone*2

FIH Spec (DS/DP)

Purity

Size Exclusion HMW 1.2 2.1 2.7 < 5%

Ion-exchange

Acidic 13.2 15.2 15.7

± 10% Main 74.0 72.0 67.9

Basic 12.8 11.6 14.1

rCE-SDS

LC+HC 98.1 97.8 97.6 ≥ 95%

LMW 0.0 0.0 0.0 N/A

NGHC 0.8 0.8 1.0 N/A

nrCE-SDS

Pre-Peaks 5.5 5.5 4.8 < 5%

%HH 2.1 1.4 2.2 N/A

%HHL 1.6 1.9 1.7 N/A

Glycan

% Afucosylation 1.5 3.0 3.9

N/A % High Mannose 4.0 3.8 4.2

% Galactosylation 7.1 8.6 9.0

Impurity CHOP ng/mg 50 < 200 ppm

DNA pg/mg 0.7 < 10 ng/dose

Potency Binding % 107 60-140%

Molecule 2: Comparable PQAs for material from cell pools & clones

Assay Attribute Cell pool Clone 1 Clone 2 FIH Spec (DS/DP)

Purity

Size Exclusion

HMW 4.1 3.5 3.4 < 5%

Ion-exchange

Acidic 11.9 11.2 24.5

± 10% Main 65.8 65.3 54.6

Basic 22.3 23.4 20.8

rCE-SDS LC+HC 97.9 98.5 98.3 ≥ 95%

nrCE-SDS Pre-Peaks 1.7 3.2 3.1 < 5%

%Main 98.3 96.8 96.9 N/A

Glycan

% Afucosylation 1.6

1.3 1.74

N/A % High Mannose 6.4

7.4 10.7

% Galactosylation 13.1

11.9 20.8

Molecule 3: Comparable PQAs for material from cell pools & clones

0

1

2

3

4

5

6

PreMCB MCB Start of Production Bioreactor

End of Production Bioreactor

Titer

VCD

High Mannose

Galactosylation

Molecule 3: similar PQA profile from vial thaw to end of production bioreactors for cell pool and clones

A-Fucosylation

HMW Pool Clone

• Both cell sources performed similarly: • Cell Culture Process

• Purification Process

• Product Quality Attributes

• Purity /Impurity

• Stability

• Manufacturing and Quality controls are used to ensure no/low risk to patient safety and product consistency

• Next steps –

• Engagement and feedback from the regulatory agencies

• Application to molecules amenable to platform

• Molecules with less complex product quality attributes

• Risk based approached to comparability

In Summary, Product Quality from Cell Pools is comparable to clones

Biosimilars

Different:

Facility and equipment

Cell-line, vector and culture media

Fermentation/culture conditions

Downstream

processing/purification

Formulation and container/closure

Out of scope of comparability guidance (ICH Q5E)

Covered by EMA, FDA, and WHO Biosimilar Guidelines

[Adapted from presentation by G. Grampp (Amgen); Manufacturing Changes in the Era of Biosimilars,

BioProcess International Conference, November 2, 2011, Long Beach, CA]

Biosimilars

Change

filter

Raw

material

supplier

Replace

equipment

Site

Change

Change

cell

culture

media

Cell line

or

formulation

change

Nature of

Process

Change

Risk & Data

Requirements Low Risk

Commonly

implemented

- Analytical data

- Process studies

Moderate Risk

- Analytical data

- Process studies

- Stability data

High Risk

Less commonly implemented

- Analytical data

- Process studies

- Stability data

- Clinical data

Governed by comparability guidance (ICH Q5E), Para 2.2 of CHMP1

Innovator Process Change

Biosimilar

development requires

most if not all changes

“Abbreviated”

comparability

“Comprehensive” comparability

Biosimilar Development Transcends the Highest Risk Category of Process Change; experience can be applied to advancing NMEs

Acknowledgements

Discovery Research

Translational Sciences

Clinical Operations &

Operations Technology

Regulatory Affairs (CMC)

Product Quality

Questions