End-to-End Integrated Continuous Manufacture of Monoclonal Antibodies

Transcription

1 End-to-End Integrated Continuous Manufacture of Monoclonal Antibodies Fabian Steinebach, Daniel Karst, Massimo Morbidelli, ETH Zurich 11/17/216 1

2 Continuous Integrated Production Opinion: Higher demands in product quality is the main driver Production Capture VI Polishing Perfusion Culture capturesmb MCSGP Advantageous: Stable operation operation More uniform product quality Lean process with smaller equipment Ease in modulation Daniel Karst

3 Perfusion: Concept and Benefits Continuous feed of nutrients/ removal of inhibitory metabolites (LAC, AMM) Cell retention device to obtain high cell concentrations Advantages: - prolonged culture at higher VCD 11/17/216 3

4 Product Residence Time Continuous Protein fraction / (-) Perfusion Fed-batch Fed-Batch Time / (days) Probability for protein modification Advantages: - prolonged culture at higher VCD - short residence time favorable for unstable proteins - steady state improved/constant product quality/homogeneity - Small equipment footprint 11/17/216 4

5 Reactor Control for stable operation Bleed A 8 1 Feed Xv Harvest fixed to 1 RV/day VCD / (1 6 cells/ml) Cell density & viability Viability / (%) gravimetric feedback i / (d ) All results for an IgG monoclonal antibody Daniel Karst

6 Continuous Capture Process Improve process performance by counter-current loading

7 Continuous Multi-Column Capture Concept: Adsorption on several columns Single column can be loaded beyond DBC Breakthrough adsorbed on 2 nd column Benefits: Better resin utilization Less buffer consumption Higher productivity 11/17/216 7

8 Process Modeling and Simulation a. Adsorption Molecular level 2 Adsorption sites for each Protein A molecule b. Mass transport Resin particle level Core shrinkage with moving boundary due to adsorption c. Mass balance Column level Integration of mass balance results in breakthrough curve 11/17/216 8

9 Parameter Estimation Model calibration with breakthrough curves i. Different mab concentrations from different cell concentrations ii. Different linear velocities due to different perfusion rates.2 c Feed =.2 mg/ml.35.3 c Feed =.43 mg/ml.6.5 c Feed =.77 mg/ml Concentration [mg/ml] cm/h 36 cm/h 458 cm/h Concentration [mg/ml] cm/h 36 cm/h 458 cm/h Concentration [mg/ml] cm/h 36 cm/h 458 cm/h Volume [ml] Volume [ml] Volume [ml] Model-based design of continuous capture process 11/17/216 9

10 Effect of Cycle Length on Process Performance Yield (%) Capacity utilization (%) Yield Capacity utilization Buffer consumption Cycle length (min) Buffer consumption (L/g) Cycle length limited by product loss For a given feed concentration there is an optimal cycle length Process needs to be adapted accordingly

11 Comparison of Capture Process Performances c Feed =1.5 g/l Batch 1 cm CaptureSMB 1cm 3-C PCC 1cm 4-C PCC 1cm Increased capacity utilization more Product per column lifetime decreased Buffer utilization higher product concentration Simulation results D. Baur, et al., Biotech. J., /17/216 11

12 Integration of USP and DSP Integration of capture step - Less product storage - increased product quality - stable and robust process Need adaptation to process changes - cell aging - resin aging - disturbances in the environment/ flows

13 Setup for Continuous Integrated Process Perfusion Harvest rate: 1RV/day Capture Columns: 2 x 2 ml Cycle time adjusted to feed concentration A VCD / (x1 6 cells/ml) B Flow rate / (1/day) Time / (days) 1.4 Harvest 1.2 Bleed Time / (days) Viability / (%) 11/17/216 13

14 Process Control c feed / (mg/ml) mio/ml transition 6 mio/ml 4 mio/ml Operation points 99% 99.9% 95% < 95% t cycle / (min) USP Flow rate Titer Model DSP Column Size Switch times A B Harvest concentration / (g/l) t cycle / (min) at-line offline Time / (days) Performance Purity Yield Product conc. Productivity C Productivity / (mg/ V col /cycle) Time / (days) Time / (days) Yield / (%)

15 Product Quality at Steady State Process performance: better economic performance at higher viable cell density A 1 B 5 C 7 D Media Buffer Productivity Yield Product Quality: Varying product quality between each steady state 11/17/ Consumption / (L/gmAb ) Productivity / (mg/ml/h) Overall yield / (%) 4 2 Consumption / (L/gmAb ) F x1 6 cells/ml 4 x1 6 cells/ml 6 x1 6 cells/ml F Relative amount / (%) Relative amount / (%) Glycoform / (%) 2 CV-2 CV-1 CV- CV+1 CV+2 Oligomer Dimer Monomer Fragments M5+FA1 FA2 FA2G1 FA2G2 FA2G2S1

16 Continuous Polishing Product polishing Removal of product & process related impurities

17 MCSGP Polishing: Multicolumn Process Goal separate 3 components Implementation Apply gradient Alternate between interconnected and batch states I1 1 /B1 1 /I2 1 B2 1 I1 2 B1 2 I2 2 B2 2 Recycle overlapping regions of chromatogram high yield take out pure fractions high purity All columns do same operations over one cycle concentration c F c E c D c C c B c A c Load Load F Wash Elute W Recycle W/P Elute P Recycle P/S Clean column W P S CV A CV B CV C CV D CV E Column Volume

18 MCSGP Polishing: Multicolumn Process Goal separate 3 components Implementation Apply gradient Alternate between interconnected and batch states Recycle overlapping regions of chromatogram high yield take out pure fractions high purity All columns do same operations over one cycle

19 Acknowledgement The USP Team Ernesto Scibona Vania Bertrand Moritz Wolf Veronika Schneider Anna Pechlaner Dr. Hervé Broly Dr. Matthieu Stettler Xavier Le Saout Dr. Jonathan Solacroup and the USP team DSP Team Nicole Ulmer Daniel Baur Lara Decker Fabian Feidl

20 Thanks for your attention