BPI West 2017 Presentation, Leslie S. Wolfe, Ph.D.
Principal Scientist, Downstream Process Development, KBI Biopharma & Abhinav A. Shukla, Ph.D. Senior Vice President, Process Development & Manufacturing, KBI Biopharma
9. • Parameters shaded in gray are
defined across molecules.
Parameters shaded in yellow
require molecule specific
optimization
• For all Spero molecules the
operating parameters, basal
medium, feed type and some of
the supplement additions have
defined
• The need for additional
supplements is molecule specific
• Reasons for supplement
addition:
» Biocompatibility
» Increase in productivity
Scale
Temperature Set point 37.0 ± 0.5°C
Temperature Shift 33.0 ± 0.5°C on Day 6
DO Set point 30%
pH Set point 6.90 ± 0.1
Agitation (1-impeller) 50 rpm → 55 rpm
Air overlay 1.6 SLPM
Air Sparge 0.5 SLPM
Max. Oxygen Sparge 5 SLPM
Max. CO2 Sparge 5 SLPM
Medium CD OptiCHO + 8 mM Glutamine
Target VCD 0.50 x 106
cells/mL
Base 1M Sodium carbonate
Feed Type: LTI Feed A+B (1:1)
15% on Day 0, 10% current wv each
on Days 3, 6, and 9
Supplement 1 addition: HT Supplement 1X current wv each on Days 0 and 4
Supplement 2 addition: Cystine
Supplement 3 addition: Tyrosine
Supplement 4 addition: Soy:Yeastolate
Hydrolysate (2:3)
5g/L current wv each on Days 4 and 8
Supplement 5 addition: C1615
Harvest Add 10g/L Hydrolysate on harvest
9
11. • Vaccine candidate is monomeric gp120
• Heavily glycosylated with glycans comprising 50%
by mass
• High mannose glycans are the predominant
glycoform
• Charge profile of gp120 targets have a
heterogeneous charge profile
• Charge profile spans a broad pH range and is
molecule specific
• Molecule prone to proteolytic cleavage
1 2 3 4
Lane gp120
1 CH505TF
2 CH505w53
3 CH505w078
4 CH505w100
IEF Gel
12. • HIV env field routinely relied on lectin capture to isolate
gp120
• Some differences in glycan heterogeneity observed between
lectin and non‐lectin purified samples
• Not suitable approach for GMP manufacturing
• Implementation of a non‐affinity based capture step
• Focus on mixed mode capture for ability to fine tune selectivity
• Polishing chromatography steps rely heavily on charge
interactions
• Development supported by DHVI SPR assay and KBI octet
binding assay to ensure product activity was maintained
16. 0 1 2 3 4 5 6 7 8 9
0-600mM
NaCl gradient
• Later eluting
fractions have better
binding for CH106
• An intermediate
NaCl wash was
developed to enrich
“active” species in
eluate fraction
20. • Molecule specific challenges have led to evolution of the downstream
platform process
• Removal of intermediate urea washes
• Conversion of low pH VI to detergent inactivation
• Evaluation of alternative capture step where needed
• Despite minor optimization required, polishing unit operations, UF/DF
and viral filtration are largely unchanged from molecule to molecule
• Development and implementation of a platform downstream process
has reduced development time and streamlined tech transfer to
manufacturing
• Approach supports rapid development and manufacturing to enable
clinical entry for predicted iterative experimental Phase I trials
21. 21
Duke Human Vaccine Institute:
Dr. Barton Haynes,
Prof. Tom Denny and team
Tom VanCott, PhD and team
Dr. Mike Pensiero and team
Cell line development, Upstream &
Downstream PD, Analytical Development,
Formulation Development, cGMP
Manufacturing, QA/QC