A Case Study of Scale-up and Tech Transfer from Non-Disposable to Fully
Disposable Systems
Sigma S. Mostafa, Ph.D., Director, Upstream Development Abhinav Shukla, Ph.D., Vice President , Process Development
Disposable Seed, Production, & Harvest
Shake Flasks (20 mL – 2000 mL)
Wave Bags (2L – 25L)
Bioreactors (40L – 2000L)
Depth Filters (Sized based on need)
Strategy for Tech Transfer to Disposable Bioreactors
• Detailed characterization of disposable bioreactors
• Development of in-house scalability data
• Determination of a robust scaling factor (e.g. Power per volume)
• Delivery of a comparable process through choosing appropriate aeration and agitation strategy
4
Characterization of Bioreactors
• Response surface DOE for Mass Transfer Coefficient (kLa)
• Salt solution with ionic strength similar to cell culture
• Sparging Rate, Agitation Rate, and Working Volumes
are key factors
200L Disposable Bioreactor with 20um Sparger
Sparger Selection
6
• Vendors offer multiple options for spargers:
2 μm, 20 μm, 0.5 mm, 1.0 mm
• Multiple disks of same porosity is available
• A dual sparger system: 1) micro and/or macro
sparger disks for oxygen; 2) wand (drilled pipe)
for air is available
• Each of these options will lead to different
mixing characteristics
Impact of Probes
7
• Depending on agitation rate, impeller
orientation, and working volume, probe
location may impact probe reading
• Disposable probes preferable; however,
accuracy is still under question
Case Study - I
• Tech Transfer project (200L)
• At scale data available from other bioreactor systems
• Shear damage a concern; as a result relatively low agitation is used
• Sparger disk with 2 μm porosity used
8
Viable Cell Density
9
• 4 GMP runs at 200L scale show minimal variability
• Data from KBI and Client match well (not shown)
Viability
10
• 4 GMP runs at 200L scale show minimal variability
• Data from KBI and Client match well (not shown)
Titer
11
• 4 GMP runs at 200L scale show minimal variability
• Data from KBI and Client match well (not shown)
Titer
12
• Titer data at KBI was similar to client’s data
2
2.1
2.2
2.3
2.4
2.5
2.6
2.7
2.8
2.9
3
Fin
al
Tite
r
Client KBI
Source
Onew ay Analysis of Final Titer By Source
Dissolved CO2
13
• KBI dCO2 high due to use of a microsparger and low agitation rate
50
70
90
110
130
150
170
190
Max
dC
O2
Client KBI
Source
Onew ay Analysis of Max dCO2 By Source
Product Quality
14
• KBI’s product quality data within range provided by client
60
65
70
75
80
85
90
95
100
G0
F
Client KBI
Source
Onew ay Analysis of G0F By Source
Case Study - II
15
• Tech Transfer project (200L, 2000L)
• Intermediate scale data available
• Dissolved CO2 a concern
• Dual sparging system used – i) Wand with drilled holes for air supply (for CO2 removal)
ii) Four 20 μm sparger disks used for O2 supply
Titer
16
• VCC, viability within ranges of
client data (not shown)
• Titer within range of client
data
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
Client KBI
Source
Onew ay Analysis of Final Titer By Source
Case Study - III
17
• Process Development project
• Data generated in 3L and 10L glass bioreactors, and 200L and 2000L bioreactors
• Oxygen uptake rate (per cell) was high; cells were sensitive to high dissolved CO2
• Antifoam addition was not an option
3L Glass Bioreactor Characterization
20
kL
a kLa
• Characterization of small scale bioreactors is a necessity
• Per cell oxygen uptake rate was determined at this scale
Comparability Across Scale – 3L, 15L, 200L, and 2000L
Viable Cell Density
• VCD data matches across scale
Viability
22
• Viability data matches across scale
Comparability Across Scale – 3L, 15L, 200L, and 2000L
Product Quality
24
• KBI’s product quality data within range provided by client
25
30
35
40
45
50
55
60
G0
F
Client KBI
Source
Onew ay Analysis of G0F By Source
Harvest Clarification
• Current platform: depth filtration
• Alternate platforms being evaluated: Disposable Centrifuges
25
Resistance vs. Throughput Chart
26
0.00
0.05
0.10
0.15
0.20
0.25
0 50 100 150 200 250 300
Throughput (L/m2)
Resis
tan
ce (
psi/L
MH
)
Depth Filtration Evaluation
• Resistance across different depth filters studied
Depth Filter Evaluation
27
Minimum Area Comparison
0.0
0.5
1.0
1.5
2.0
2.5
3.0
A1->B1->C A1->B2->C A2->C
Min
imu
m A
rea
(m
2)
Sterile Filter
Secondary Filter
Primary Filter
• Comparison was made of the total minimum area used for the depth and
sterile filters
Conclusions
• End-to-end disposable facility shortens change over duration and eliminates cross contamination risk
• A rigorous scientific approach in tech transfer from non-disposable to disposable leads to consistent success in the disposable platform.
28
Questions ?
Contact Information:
Sigma S. Mostafa, Ph.D.
Director, Process Development
KBI Biopharma Inc.
1101 Hamlin Road
Durham, NC 27704
29
Depth Filtration Evaluation
30
Turbidity vs. Throughput
0 50 100 150 200 250 300
Throughput (Volume/Surface Area)
Tu
rbid
ity
• Turbidity post depth filter was studied
0.22 μm Polishing Filter Study: Vmax Throughput vs. Time
31
0 5 10 15 20 25 30
Time (min)
Th
rou
gh
pu
t (l
/m 2
)
.
post filter 1
post filter 2
post filter 3
Polishing Filter Evaluation
• Throughput across polishing filters varied based on load material
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