Orphan Biopharmaceuticals

& the CDMO

(Contract Development and Manufacturing Organization)

Abhinav A. Shukla, Ph.D.

Vice President, Process Development & Manufacturing

KBI Biopharma, Durham NC

Presented at: World Orphan Drug Congress, Washington DC, April 9-11, 2013

Why are orphan biopharmaceuticals unique? • Smaller material demand

• Fewer clinical batches reduced large scale manufacturing experience prior to BLA/MAA filing Flexible manufacturing at a smaller scale (< 2000L cell

culture volumes) needed (Single-Use Manufacturing Technologies) Increased focus on process knowledge from scale-down

experimentation (QbD)

• Limited ability to do clinical bridging studies • Process changes during clinical development are less

desirable since their clinical impact can often not be studied readily Getting the process right the first time (Building robustness

and scalability into the process right from the start)

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Biologics Commercialization

Pre-Clinical Phase I Phase II Phase III

Process Development Process

Characterization Process

Validation Process Monitoring

& Improvement

FIH Process • Deliver clinical process

quickly • Platform process • Clinical Supply

Submission & Approval

Lifecycle management

BLA Prep & PAI

Commercial Process • Deliver manufacturing process for

registrational trials and market • Design keeping large-scale manufacturing in

mind • Improve productivity, efficiency, robustness,

manufacturability, COGs • Analytical characterization and method

development

Process Characterization and Validation • Develop IPC strategy through understanding of process inputs and

outputs (design space) • Scale-down characterization and validation studies • Large-scale process validation to demonstrate process consistency • BLA preparation • Supporting documents for licensure inspections • Post-commercial process improvements (CI) • Post-commercial process monitoring

FIH process Commercial process

Gottschalk U., Brorson K., Shukla A. Nature Biotechnology, 30(6), 489-491, 2012

-Confidential-

Biologics Commercialization

Pre-Clinical Phase I Phase II Phase III

Process Development Process

Characterization Process

Validation Process Monitoring

& Improvement

FIH Process • Deliver clinical process

quickly • Platform process • Clinical Supply

Submission & Approval

Lifecycle management

BLA Prep & PAI

Commercial Process • Deliver manufacturing process for

registrational trials and market • Design keeping large-scale manufacturing in

mind • Improve productivity, efficiency, robustness,

manufacturability, COGs • Analytical characterization and method

development

Process Characterization and Validation • Develop IPC strategy through understanding of process inputs and

outputs (design space) • Scale-down characterization and validation studies • Large-scale process validation to demonstrate process consistency • BLA preparation • Supporting documents for licensure inspections • Post-commercial process improvements (CI) • Post-commercial process monitoring

FIH process Commercial process

A single development cycle Robust and complete process characterization package

Commercial manufacturing at smaller scales

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SINGLE-USE MANUFACTURING TECHNOLOGIES

Why are single-use manufacturing systems growing? • Lower capital and utility costs (up to 40% reduction*) • Increasing titers driving bioreactor scales smaller

• Single-use bioreactors now up to 2000L volume

• Increased universalization of biomanufacturing • Co-location of manufacturing with markets • Biosimilars (estimated $ 17 billion market by 2020) • Smaller market sizes for novel drugs in niche/personalized

applications • Market fragmentation making large single-product

manufacturing facilities redundant

• Single-use systems finding application in stainless steel facilities for enhanced operational flexibility

Laukel et al, BioProcess International, May 2011 Supplement, pp. 14-21.

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Media and Feed preparation utilizing disposable mixing, filtration and storage systems

Disposable shake flasks or disposable spinner flasks

MCB or WCB vial

Disposable expansion reactor

Disposable seed bioreactor

Disposable production bioreactor

Disposable fluid path centrifuge

Disposable depth filtration system

0,2 µm filter

Hold vessels (Bags)

Hold vessel (bag)

Disposable fluid path purification system

Disposable mixing tank

0,2 µm filter

Retentate

Permeate

PD

Disposable fluid path purification system

Disposable mixing tank

0,2 µm filter

BPC

Virus filter

BPC

0,2 µm filter

BPCBPC

Sterile bulk fill and sampling bags

Buffer preparation utilizing disposable mixing, filtration and storage systems

0,2 µm filter

Disposable fluid path UF/DF system

Aseptic connection

Hold vessel (bag)

Hold vessel (bag)

Hold vessel (bag)

Hold vessel (bag)

Hold vessel (bag)

Process Reproducibility

4 manufacturing runs in Single Use Bioreactors

Highly consistent process

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Scalability

•4 different scales •3L and 15L scales in non-disposable bioreactors

•Process performance with different working volumes is also reproducible

Single-use technologies in downstream processing • Centrifugation (kSep® Systems)

• Closed, continuous centrifuge with class VI product contact surfaces

• Counteraction of Centrifugal force and fluid flow force • Very low shear • Continuous operation • Reversal of flow direction empties the chamber • Up to 7.2 L/min

Single-use technologies in downstream processing • Depth filtration:

• Harvest depth filters have traditionally been single-use except for their holders

• Based on particle entrapment in a fibrous bed • Can be used as the primary cell separation step for smaller cell

culture harvest volumes • Millipore – POD® system • Pall - Stax® system • Sartorius – Sartoclear P ® • Cuno – Zeta Plus ®

Pall – Stax System

Millipore - POD

Single-use technologies in downstream processing • Chromatography

• Membrane adsorbers • Mustang® (Pall), Sartobind® (Sartorius), Chromasorb® (Millipore),

Adsept® (Natrix), • Q, S, HIC and salt-tolerant ion-exchange functionalities • Most widely used for trace impurity removal in a flow-through mode

(DNA, endotoxin, viral clearance) • Pre-packed chromatography columns

• ReadyToProcess (GE Healthcare), Opus (Repligen), GoPure (Life Technologies)

• Monoliths • CIM monoliths (BIA Separations), Uno monoliths (Biorad)

Up to 20 cm D available

Clinical and commercial manufacturing using single-use technologies • Smaller material demand drives reduced scale for

commercial manufacturing • Fidelity between clinical and commercial product

needed (ideally single facility that fits both needs) • Single-use manufacturing technologies reduce costs

and reduce risk of cross-contamination

-Confidential-

Shukla, A., Mostafa, S., Wilson, M., Lange, D. Vertical Integration of Disposables in Biopharmaceutical Drug Substance Manufacturing, Bioprocess International, 10(6), 34-47, 2012. Gottschalk, U., Shukla, A. Single-use disposable technologies for biopharmaceutical manufacturing, Trends in Biotechnology, 31(3), 147-154, 2013.

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QUALITY BY DESIGN (QBD)

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Quality by Design (QbD) • “Quality by design means designing and developing

manufacturing processes during the product development stage to consistently ensure a predefined quality at the end of the manufacturing process.” ICH Q10, FDA 2006

Process Design (Process Development)

Process Control Strategy Definition

Process Validation

Continued Process Verification

QbD

Critical Quality Attributes (CQAs)

Process Design Space

Linking CQAs to Clinical outcome

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Process design space Characterization Space

Control Space

Operating Range

Acceptable Range

Design Space

Process Parameters

Key Parameters

CPPs

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Integrative Approach Each step is influenced by the preceding step

Shake flask and seed bioreactor parameters may affect growth rate in the seed bioreactor.

Seed bioreactor and production bioreactor parameters may affect productivity and critical quality attributes.

Production bioreactor parameters may affect downstream steps.

Characterization studies are linked.

Vial Thaw

Shake Flasks Seed Bioreactor

Production Bioreactor

Downstream Steps

Biotechnology and Bioengineering, 106(6), 894-905, 2010.

Production Bioreactor

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Establishing A Process Control Strategy

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Application of Quality by Design (QbD): downstream resin lot variability

Biotechnology and Bioengineering, 107(6), 989-1001, 2010.

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Evolving expectations in Process Validation

• Q7A definition: “Process validation is the documented evidence that the process, operated within established parameters can perform effectively and reproducibly to produce an intermediate or API meeting its predetermined specifications and quality attributes” • FDA guidance, Jan 2011: “The collection and evaluation of data, from the process design stage through commercial production, which establishes scientific evidence that a process is capable of consistently delivering quality product” • Process validation is now viewed as a process that occurs throughout the lifecycle of a product

Process Design (Process Development)

Process Control Strategy Definition

Process Qualification

Continued Process Verification

Scale-Down Process Validation Studies

• Scale-down validation studies in addition to large-scale process validation (conformance lots)

• Probe extremes in the process and demonstrate them to be acceptable

• Examples • Reprocessing validation – combine hold times with process

conditions that create the greatest stress on the protein • Intermediate hold times – combine hold times and

demonstrate releasable drug substance • Viral clearance studies • Impurity clearance studies

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Validation of Host Cell Protein Clearance

Harvest

Column 1

Column 2

Column 3

Worst-case C1 eluate

Worst-case C2 eluate

Harvest

Column 1

Column 2

Column 3

Harvest

Column 1

Column 2

Column 3

Spiking Strategy • Some CHOP species in harvest

may not be encountered by C2 and C3 in Mfg

• LVR could be overstated for C2 and C3

Worst-case Strategy • CHOP species in eluate is relevant

to the next step • More accurate evaluation of LRV • Need process characterization to

identify worst-case condition

By-pass Strategy • HCP species in load are relevant to

that process step in case the previous step is by-passed (e.g. “resin bed channeling”)

• Represents most “challenged” scenario

Biotechnol. Progr., 24(3), 615 – 622, 2008

Worst-case harvest

Development Phase • Utilizing the right set of analytical tools for in-process

testing and release • Characterization assays are equally important • Utilizing a broad set of tools up front gives the best

chance of determining CQAs & linking them to the process

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Analytical Methods Portfolio • Protein Primary Structure

Peptide Sequencing via LC/MS/MS Amino Acid Analysis Peptide Mapping

• Biophysical Characterization CD, FTIR, DSC, DLS, fluorescence

spectroscopy

• Capillary and Slab Gel Electrophoresis CZE SDS-CGE cIEF and icIEF SDS-PAGE and IEF Western blot Microchip electrophoresis 2D gels and blots

• Glycan Analysis Oligosaccharide mapping Monosaccharide composition Sialic Acid Quantitation

• Process Residuals • ELISA (HCP, protein A etc.) • HPLC (antibiotics, IPTG, detergents, etc) • qPCR (DNA)

• HPLC • Size Exclusion (with MALLS) • Ion Exchange • Reverse Phase • Hydrophobic Interaction • Affinity

• Potency Assays • Binding Assays via ELISA, Biacore and

ForteBio • Cell Based Assays (e.g., proliferation,

cytokine release, etc.)

• Mass Spectrometry • Intact mass • Peptide mapping with LC/MS or

LC/MS/MS • Disulfide Mapping • Post translational modifications (e.g.,

oxidation, deamidation) • PEGylation site identification • Glycan Identification & site identification

• Particle measurements

• Visible & sub-visible particles

Comprehensive Analytics

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THE RIGHT SCIENCE FROM THE START

Designing more efficient HCP clearance into the downstream process • Most current chromatographic steps are designed to

remove impurities based on differential binding to the stationary phase surface

• Conventional wisdom: wash conditions are between binding and elution conditions

• Orthogonal approach disrupt impurity-product interactions

Washes that disrupt protein-protein interactions

Conventional washes

30

Enhancing HCP clearance across Protein A • HCPs form a diverse set of impurities • HCP clearance is a key concern in biopharmaceutical

separation processes

-Confidential-

Washes can be developed to disengage HCPs from the product rather than disrupt product-Protein A ligand interactions

96

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05000

100001500020000250003000035000400004500050000

Null supernatant MAbSelecteluate (load =

nullsupernatant)

MAbSelecteluate (load =

null supernatant+ product)

Prosep A eluate(load = null

supernatant)

Prosep A eluate(load = null

supernatant +product)

Hos

t Cel

l Pro

tein

s (n

g/m

L)

Normalized Yield vs. normalized CHOP for a variety of washes on MAbSelect Protein A

0%

20%

40%

60%

80%

100%

120%

140%

0% 20% 40% 60% 80% 100% 120%

Yield normalized to control experiment

CH

OP

(ppm

) nor

mal

ized

to

cont

rol e

xper

imen

t

Direction ofdesired trend

Biotechnology Progress, 24, 1115-1121, 2008.

Do HCPs co-elute with the product or co-associate with the product?

Enhancing HCP clearance across Protein A

Enhancing HCP clearance across Protein A • Use washes at high pH (pH > 7) to preserve Protein A –

mAb interactions • Include selective modulators (moderate concentrations of

urea, ethylene glycol, salts, arginine) in washes to disrupt HCP-mAb interactions

Shukla, A., Hinckley, P. Host cell protein clearance during Protein A resin chromatography: development of an Improved wash step, Biotechnology Progress, 24, 1115-1121, 2008.

Evaluation of intermediate washes at pH > 7.0

0%

20%

40%

60%

80%

100%

120%

140%

0% 20% 40% 60% 80% 100% 120%

Normalized yield % of control

Norm

alize

d CHO

P (%

of co

ntro

l)

Mixed Mode Chromatography

• Takes advantage of more than one type of interaction • Can reduce process steps • Provides enhanced selectivity, “pseudo-affinity” • Several mixed mode resins have recently been developed with:

» Increased loading capacities » Higher ionic strength tolerance

+

+ +

+ + Mixe

d Mode

GE Healthcare, Capto MMC ligand

Ionic interactions

Hydrophobic interactions

Hydrophobic interactions

Ionic interactions

GE Healthcare, Capto Adhere ligand

Log k’ vs Log [NaCl]

0.00

0.20

0.40

0.60

0.80

1.00

1.20

1.40

2.60 2.80 3.00 3.20 3.40 3.60

Log

k'

Log [NaCl]

Lysozyme

pH 7.0

1M urea

5% ethylene glycol

50mM arginine

-0.40

-0.20

0.00

0.20

0.40

0.60

0.80

1.00

1.20

1.40

1.60

1.50 2.00 2.50

Log

k'

Log [NaCl]

RNase

pH 7.0

1M urea

5% ethylene glycol

50mM arginine

-0.20

0.00

0.20

0.40

0.60

0.80

1.00

1.20

1.40

2.10 2.30 2.50 2.70

Log

k'

Log [NaCl]

Monoclonal antibody

pH 7.0

1M urea

5% ethylene glycol

50mM arginine

Wash development on mixed mode

0

50

100

150

200

250

300

350

400

450

500

0.0% 20.0% 40.0% 60.0% 80.0% 100.0%

HCP

(ppm

)

Recovery

Capto MMC HCP Clearance 25mM Tris pH 7.0 (baseline)

25mM Tris pH 7.0, 5% ethylene glycol

25mM Tris pH 7.0, 50mM arginine

25mM Tris pH 7.0, 50mM NaSCN

25mM Tris pH 7.0, 1M urea

25mM Tris pH 7.0, 1M ammonium sulfate

25mM Tris pH 7.0, 0.1M NaCl

25mM Tris pH 7.0, 0.5M ammonium sulfate

25mM Tris pH 7.0, 0.1M NaCl, 1M urea

25mM Tris pH 7.0, 0.1M NaCl, 1M urea, 5% ethylene glycol

25mM Tris pH 7.0, 0.1M NaCl, 1M urea, 5% glycerol

• Selective wash strategies can eliminate one chromatographic step in non-mAb processes • Designing quality into the process

Designing processes with the end in mind • Having the right analytical methods and product quality

profile in mind from the start • Keeping issues that can be encountered in large-scale

manufacturing in mind from the beginning

Process yields & robustness

Titer & downstream yields Reproducibility

Column loading and buffer needs

Column loading drives costs!

Raw material selection Potential for variability

Supply assurance Compatible with cGMP

Process impact

Transfer ready processes

Processes that can be compatible with many scales and facilities

Conclusions • Orphan biopharmaceutical development needs

particular emphasis on • Developing a process with the end in mind (licensure filing)

to avoid multiple changes along the way • Manufacturing costs • Demonstrating process robustness without recourse to an

extensive manufacturing history

• A dedicated CDMO with the right knowledge and capabilities can help smooth the development pathway