Continuous Processing in Pharmaceutical Manufacturing -- Contents -- List of Contributors -- Preface -- 1. Proteins Separation and Purification by Expanded Bed Adsorption and Simulated Moving Bed Technology -- 1.1 Introduction -- 1.2 Protein Capture by Expanded Bed Technology -- 1.2.1 Adsorbent Materials -- 1.2.2 Expanded Bed Adsorption/Desorption of Protein -- 1.2.3 Modeling of the Expanded Bed -- 1.3 Proteins Separation and Purification by Salt Gradient Ion Exchange SMB -- 1.3.1 Adsorption Isotherms and Kinetics of BSA and Myoglobin on Ion Exchange Resins -- 1.3.2 Salt Gradient Formation and Process Design for IE-SMB Chromatography -- 1.3.3 Separation Region of Salt Gradient IE-SMB Chromatography -- 1.3.4 Proteins Separation and Purification in Salt Gradient IE-SMB with Open Loop Configuration -- 1.4 Conclusion -- References -- 2. BioSMB Technology as an Enabler for a Fully Continuous Disposable Biomanufacturing Platform -- 2.1 Introduction -- 2.2 Integrated Continuous Bioprocessing -- 2.3 Multicolumn Chromatography -- 2.4 BioSMB Technology -- 2.5 Fully Disposable Continuous Processing -- 2.6 Case Studies -- 2.7 Regulatory Aspects -- 2.8 Conclusions -- References -- 3. Impact of Continuous Processing Techniques on Biologics Supply Chains -- 3.1 Introduction -- 3.1.1 The Biologics Industry -- 3.1.2 The Biologics Value Chain -- 3.1.3 Downstream Purification Costs -- 3.2 Chromatography Techniques Used in Downstream Purification of Biomolecules -- 3.2.1 Need for Continuous Manufacturing in Downstream Purification -- 3.2.2 The Multicolumn Countercurrent Solvent Gradient Purification Chromatography System -- 3.3 Next-Generation Biologic Products - Bispecific Monoclonal Antibodies -- 3.3.1 Major Biopharma Companies and Their Interest in Bispecific Mabs -- 3.3.2 Challenges in Purification of Bispecific Monoclonal Antibodies.

3.4 Improving the Downstream Processing of Bispecific Mabs by Introduction of MCSGP in the Value Chain -- 3.4.1 Advantages of Utilizing MCSGP Process in Bispecific Mabs Purification as Compared to Batch Chromatography -- 3.4.2 Impact of MCSGP System on Biologic Supply Chains -- 3.4.3 Impact on Patent Approval Structure of Biologic Drugs -- 3.4.3.1 For a Manufacturer Who Already has a Biologic Drug in the Market -- 3.4.3.2 For a Manufacturer Who is Developing a Biologic Drug -- 3.4.4 Impact on Big Biopharma Companies -- 3.4.5 Impact on the Chromatography Market -- 3.4.6 Limitations of the MCSGP System -- 3.5 Conclusion -- 3.6 Further Research -- Acknowledgments -- 3.A Appendix/Additional Information -- 3.A.1 Regulatory Structure for Bispecific Monoclonal Antibodies -- 3.A.1.1 Regulatory Compliance Comparison between US, EU, and Emerging Economies -- References -- 4. Integrating Continuous and Single-Use Methods to Establish a New Downstream Processing Platform for Monoclonal Antibodies -- 4.1 Introduction -- 4.2 Harvest and Clarification -- 4.2.1 The Challenge and Technology Selection -- 4.2.1.1 Centrifugation -- 4.2.1.2 Filtration -- 4.2.1.3 Impurity Precipitation -- 4.2.2 Summary -- 4.3 Capture -- 4.3.1 Background -- 4.3.1.1 Protein A Chromatography -- 4.3.2 Chromatographic Methods -- 4.3.2.1 Slurried Bed Methods -- 4.3.2.2 Continuous Chromatography -- 4.3.3 Capture Case Studies -- 4.3.3.1 Continuous Protein A Chromatography Capture Case Study -- 4.3.3.2 Effect of Clarification Method on Protein A Performance -- 4.4 Polishing -- 4.4.1 Background -- 4.4.2 Technology Selection Strategy -- 4.4.3 Complete Flow-Through Polishing Case Study -- 4.5 Cost of Goods Analysis -- 4.5.1 Methodology -- 4.5.2 Clarification -- 4.5.3 Capture -- 4.5.4 Polishing -- 4.5.5 Overall -- 4.6 Summary -- References.

5. Modeling of Protein Monomer/Aggregate Purification by Hydrophobic Interaction Chromatography: Application to Column Design and Process Optimization -- 5.1 Introduction -- 5.2 Mathematical Model -- 5.2.1 The Rate-Limiting Step in the HIC Process -- 5.2.2 Dimensional Considerations -- 5.2.2.1 Adsorption Capacity vs. Concentration of Vacant Sites (qmi vs. Cvi) -- 5.2.2.2 Concentration of Protein Adsorbed on Resin (qi vs. Cia) -- 5.2.3 Mathematical Model -- 5.3 Experimentation -- 5.3.1 Protein Solutions -- 5.3.2 Determination of Adsorption and Desorption Kinetic Constants -- 5.3.3 Column Chromatography -- 5.4 Results and Discussion -- 5.4.1 Kinetic Constants -- 5.4.2 Protein Denaturation -- 5.4.3 Model vs. Experimental Results -- 5.4.4 Applications -- 5.5 Conclusion -- Acknowledgments -- References -- 6. Continuous Animal Cell Perfusion Processes: The First Step Toward Integrated Continuous Biomanufacturing -- 6.1 Introduction -- 6.2 The Basics of Perfusion Processes -- 6.3 Cell Banking and Inoculum Development in the Context of Perfusion Processes -- 6.4 Culture Conditions -- 6.5 Cell Retention Devices -- 6.5.1 Gravitational Settlers -- 6.5.2 Centrifuges -- 6.5.3 Hydrocyclones -- 6.5.4 Acoustic (Ultrasonic) Separators -- 6.5.5 Tangential Flow-Filtration -- 6.5.6 ATF Systems -- 6.5.7 Floating Membrane Devices -- 6.5.8 Spin-Filters -- 6.5.9 Rotating Cylindrical Filters (Vortex-Flow Filters or External Spin-Filters) -- 6.5.10 Rotating Disc Filters (Controlled-Shear Filters) -- 6.6 Integrated Perfusion-Purification Processes for Continuous Biomanufacturing -- 6.7 Concluding Remarks -- References -- 7. Perfusion Process Design in a 2D Rocking Single-Use Bioreactor -- 7.1 Introduction -- 7.2 Production Costs -- 7.3 Equipment Requirements for a Single-Use Perfusion Process -- 7.4 Testing Results Single-Use Perfusion Process -- 7.5 Simplified Seeding Process.

7.6 Future Outlook -- References -- 8. Advances in the Application of Perfusion Technologies to Drosophila S2 Insects Cell Culture -- 8.1 Introduction -- 8.2 Case Study 1: Acoustic Separation -- 8.2.1 The Perfusion Setup (BioSep) -- 8.2.2 Results and Discussion -- 8.2.2.1 Development -- 8.2.2.2 Cell Count in the Bioreactor -- 8.2.2.3 Effects of BioSep Settings on Cell Loss and Viability -- 8.2.2.4 Controlling the Cell Concentration Through Bleed Rate Control -- 8.2.2.5 Effect of Total Dilution Rate on Culture Viability -- 8.2.2.6 Development of the Perfusion Rate Profile -- 8.2.2.7 Initial Testing of Robustness of Upstream Process in 1.5 l Fermentations -- 8.2.2.8 Scaling Up and Consistency in 4.5 l Fermentations -- 8.2.2.9 Process Scale-Up -- 8.2.3 Conclusions for Case Study 1 -- 8.3 Case Study 2: ATF-Based Cell Retention -- 8.3.1 ATF Technology -- 8.3.2 Methods -- 8.3.3 Results -- 8.3.3.1 Cell Counts Achieved Using Perfusion Technology -- 8.3.3.2 Effect of Feed Strategy -- 8.3.3.3 Yield Improvements Achieved Using Fed-Batch and Concentrated Perfusion -- 8.3.3.4 Protein Stability -- 8.3.4 Conclusions for Case Study 2 -- 8.4 Final Remarks -- Acknowledgments -- References -- 9. Single-Use Systems Support Continuous Bioprocessing by Perfusion Culture -- 9.1 Introduction -- 9.2 Potential Advantages in Continuous Processing -- 9.2.1 Improved Product Quality -- 9.2.2 Ease in Process Development -- 9.2.3 Improved Scalability -- 9.2.4 Increased Profitability -- 9.2.5 Sustainability -- 9.3 Challenges in Adoption of Continuous Processing -- 9.4 Continuous Biomanufacturing -- 9.5 Single-Use Systems -- 9.6 Hybrid Systems -- 9.7 Perfusion Culture -- 9.8 Single-Use in Continuous Biomanufacturing -- 9.9 Roller Bottles -- 9.10 Mechanically Agitated Suspension Reactors -- 9.11 Hollow Fiber Media Exchange -- 9.12 Packed Bed Bioreactors.

9.13 Hollow Fiber Perfusion Bioreactors -- 9.14 Continuous Flow Centrifugation -- 9.15 Acoustic Wave Separation -- 9.16 Conclusion -- References -- 10. Multicolumn Countercurrent Gradient Chromatography for the Purification of Biopharmaceuticals -- 10.1 Introduction to Multicolumn Countercurrent Chromatography -- 10.2 Introduction to Multicolumn Simulated Moving Bed (SMB) Chromatography -- 10.3 Capture Applications -- 10.3.1 Introduction -- 10.3.2 Process Principle -- 10.3.3 Application Examples -- 10.4 Polishing Applications -- 10.4.1 Introduction -- 10.4.2 Multicolumn Countercurrent Solvent Gradient Purification Principle -- 10.4.3 Multicolumn Countercurrent Solvent Gradient Purification Process Design -- 10.4.4 Multicolumn Countercurrent Solvent Gradient Purification Case Study -- 10.5 Discovery and Development Applications -- 10.6 Scale-Up of Multicolumn Countercurrent Chromatography -- 10.7 Multicolumn Countercurrent Chromatography as Replacement for Batch Chromatography Unit Operations -- 10.8 Multicolumn Countercurrent Chromatography in Continuous Manufacturing -- 10.9 Process Analytical Tools for Multicolumn Countercurrent Processes -- References -- 11. Monoclonal Antibody Continuous Processing Enabled by Single Use -- 11.1 Introduction -- 11.1.1 Single-Use Revolution to Enable Process Intensification and Continuous Processing -- 11.1.2 Principles of Continuous Multicolumn Chromatography for Biological Production (BioSMB) -- 11.2 Continuous Downstream Processing for Monoclonal Antibodies Unit Operation Development -- 11.2.1 Surge Vessels and Balancing Flows -- 11.2.2 Primary Recovery: Centrifugation and Depth Filtration -- 11.2.3 Bulk Purification: Continuous Multicolumn Chromatography - BioSMB Protein A Capture and Viral Inactivation -- 11.2.3.1 Protein A Loading Zone Optimization -- 11.2.3.2 Protein A Elution Zone Considerations.

11.2.3.3 Viral Inactivation.