KBI Biopharma has implemented single-use fermentation technology across various scales to improve efficiency and flexibility in bioprocess development. By coupling process design to the operational boundaries of their 300L single-use fermentors, KBI has been able to develop fermentation processes capable of high cell densities and product titers. Their approach of platforming single-use fermentation processes has allowed for rapid scaling between systems and accelerated timelines to bring multiple client programs into GMP production.

1. Implementation of Single-Use
Fermentation Technology
Andrew Lowell
KBI Biopharma, Inc.
Boulder, Colorado

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• KBI Biopharma, Inc. Overview
• Drivers and Strategy for Single-Use Fermentation Implementation
• Coupling Process Design to Boundaries of at-scale Single-Use Fermentors
• Challenges and Success of Single-Use Fermentation (SUF) Systems at KBI
Implementation of Single-Use Fermentation Technology: Agenda
Coupled Process Design to the SUF equipment is enabling 6-7 g/L and 200 OD600nm in soluble processes

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Durham, NC (2004)
Mammalian
• Clinical & Commercial (pre-PAI)
cGMP Manufacturing
• Analytical, Formulation, Stability & QC
• Mass Spec Core Facility
• Cell Based Assays
Boulder, CO (2014)
Microbial
• Strain Development
• Process & Analytical Development
• Clinical & Commercial (Approved)
cGMP Manufacturing
• QC, Analytical, Formulation, Stability
• Particle Characterization Core
RTP, NC (2013)
Venture Center, NC (2018)
Mammalian
• Cell Line Development
• Process & Analytical Development
• Process Characterization
• Small scale Process Validation
The Woodlands, TX (2017)
Cell Therapy
• Process and Analytical Development
• cGMP Manufacturing & Testing
• Cell Based Assays
San Diego, CA (2017)
Alliance Protein Labs
• Analytical Technologies
• Leading AUC expertise
KBI Sites
KBI’s North America Locations
March 2018
Louisville, CO (2018)
Elion Labs
• Analytical Technologies
• Leading Biophysical Chara

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SelexisKBI
Selexis, Geneva, Switzerland
(2017- an affiliate of KBI)
Best in class cell line development &
candidate selection technologies
KBI Leuven, BE
(2018)
Analytical, Formulation and QC services for
GMP & non-GMP product testing
European Offerings

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• Process & Analytical Development for First in Human
and Clinical Programs
• Late stage commercialization: Process Characterization,
Process Validation
• Technology Transfer, NPI, Materials Management
• Two Manufacturing Streams: 1500L and 100L SS, 300L
Single-Use production fermentors
• QA/QC (EM, RM, IPC, Release, Stability)
• Validation (AM, IOQ, PV)
• Operations (MFG, Document Management, Training)
• MS&T (Process Engineering, Tech Ops, Modeling)
• Engineering, Calibration, Maintenance, Facilities
Microbial Development & Manufacturing Capabilities – Colorado, USA

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Microbial Process Development: Built for Speed with Proven Scalability
250mL Scale
• Ambr250®
• Screening
• Optimization
• DOE/PC/SSMQ
10L Scale
• Sartorius/Biolaffite
• Harvest development
• Scale-up confirmation
30L Scale – Single Use
• Thermo Fisher SUF
• Harvest development
• Scale-up confirmation
100L SS GMP (Custom)
1500L SS GMP (Custom)
300L GMP Scale – Single Use
(Thermo Fisher SUF)

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• Early-phase programs with relatively small pre-clinical and clinical material needs
• Quicker process design to GMP production – platformed and equipment-based
• Higher throughput through GMP and pilot suite resulting in more flexibility in schedule
• Validation requirements much less with single-use technology at GMP scale
• Decreased cross-contamination risk between products
• Ease of operational requirements equate to time and cost savings
• Greater flexibility for KBI as a CMO organization for rapid execution of projects
Drivers for Implementing Single-Use Fermentation Systems at KBI

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• 30-liter pilot system
located in
fermentation
development
laboratory (non-GMP)
• 300-liter SUF is a GMP
system in KBI’s small-
scale manufacturing
suite
Thermo Fisher Single-Use Fermentation Systems at KBI Boulder

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Development of fermentation process into our single-use fermentation platform begins as
early as cell line development
• Understanding boundaries of at-scale 300L SUF performance
• Cell line screening and initial evaluation in 250mL SUFs. Platforming and optimization of
fermentation with equipment-based process design follows.
• 30L SUF process demonstration and confirmation of design
Single-Use Fermentation Implementation Strategy

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• High microbial growth demands exist in aggressive microbial
fermentations
• High oxygen mass transfer rates are necessary
• Lack of pressurized system limits OUR potential
• Process Development approaches generally aligned
to scalability in stainless systems
Single-Use Fermentation Limitations Dictate Design Approach
Oxygen Demand
Biomass
OUR
At-scale SUF defines
boundaries of
platform PD approach
Parallelized development
designed within
at-scale constraints

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• Batch phase is
platformed to
support optimal
growth matching
culture demand
• Late growth phase
limits OUR capacity
through tightly
controlled feed
rates and
decreased
temperature
Platform for Single-Use Fermentations

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• Feed rate in growth phase
is optimized to maximize
culture growth while
limiting OUR
• Fermentation process
optimization is focused on
production phase
Platform for Single-Use Fermentations: Full Fermentation

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• Aeration and cooling performance of 300L
single-use system typically superior to 30L pilot
system
• Comparable biomass and product titers at both
scales (comparable to S-S outputs)
Scaling Single-Use Fermentation 30L (PD) 300L (GMP)
Parameter 30L 300L
Aeration (% max O2 pull) 85% < 60%
Cooldown time (h) 1-2 < 0.5-2
Titers (g/L) 3-6 ≈ 3-6
Cell density (OD600nm) 130 - 200 ≈ 130 – 200
Cell density (WCW) 22-29% ≈ 22-29%

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Performance across Single-Use Fermentation Systems at KBI
0
10
20
30
40
50
60
70
80
90
00:00 04:48 09:36 14:24 19:12 24:00 28:48
OpticalDensity(AU/cm)
EFT (hh:mm)
250mL SUF 30L SUF 300L SUF
Run ID
Final
WCW
(%)
Final
OD600
(AU/cm)
Final Titer
SDS-
PAGE
(g/L)
250mL SUF 14.4 80 3.1
30L SUF 15.6 85 3.1
300L SUF 12.3 75 2.7
Present SUF
Max Output*
29 200 6
*30L Thermo Fisher SUF
Comparable performance across single-use
systems in same fermentation process

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• Five Single-Use Fermentation Programs through USPD lab to date
400+ x 250mL SUF Fermentations
21 x 30L SUF Fermentations
6 x 300L GMP Fermentations
• Three new programs planned in next 2 months
• High throughput and success resulting from platforming our single-use fermentation
process design
Single-Use Fermentation Programs at KBI Boulder

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• Single-Use fermentation systems have allowed greater throughput and efficiency in
laboratory leading to accelerated project timelines to GMP production
• Coupling process design to the SUF equipment capabilities has enabled processes
capable of high cell densities and high titers. Platforming single-use fermentation
allows rapid scaling among single-use systems
• Improved flexibility of KBI as a CMO organization to address multiple and diverse
programs through our single-use systems
Success of Single-Use Fermentation Implementation at KBI Boulder

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Robert Todd – VP, Process Development Aaron Pilling - Director, USPD
Erik Nordwald - Scientist, USPD Emil Raubach – Scientist, USPD
Vanna Yu – PD Assoc, USPD Molly Kruse – PD Assoc, USPD
Thatsinee Johnson – PD Assoc, USPD Aliyah Dennert – Sr. PD Assoc, USPD
Michelle Yang – Sr. PD Assoc, USPD Cassie Riehl – PD Assoc, USPD
Michelle Giuntini – PD Assoc, USPD Aaron Kuhl – Engineer, MS&T
Craig Mays – Engineer, MS&T
Acknowledgements