Chapter 17: Scale-Down Models for Robust Biologics Drug Product Process Development

  • Smeet DeshmukhEmail author
  • Maria O. Ogunyankin
Part of the AAPS Advances in the Pharmaceutical Sciences Series book series (AAPS, volume 35)


The objective of process development is to build robustness and demonstrate control of a manufacturing process to ensure consistent biological products within the specifications of its quality attributes. The new regulatory expectation and quality by design (QbD) principles laid out also reinforce the need for systematic process development approach and risk assessment to be done early and throughout the development. Biologics drug product manufacturing process and unit operations involved need to be well understood, characterized in terms of different stresses and critical process parameters that would impact their critical quality attributes. This chapter focuses on the approach of utilizing combination of small-scale/minipiloting tools and scale-down models (miniaturization of large-scale equipment) for process development of ready-to-use liquid drug product. The small-scale tools require minimal amount of materials to understand the sensitivity of molecules to the different stresses and quantify the limits of each of those stresses that the molecules can be exposed to during manufacturing. The scale-down models for each unit operation are discussed in detail with the suggestion of experimentation involving key process parameter variation to help define the process design space. Some relevant case studies are covered to explain the utility of the models and the resultant control strategy that can be put in place for robust drug product manufacturing.


Process risk assessment Minipiloting tools Shear stress Interfacial stress Light stress Freeze–thaw Mixing Filtration and pumping 



The authors want to thank Masano Huang, Andrew Ilott, Melissa Bentley, Kristine Rafferty, Johnathan Goldman, Mary Krause, Thiago Carvalho, Brenda Remy, and Mehrnaz Khossravi for their helpful discussion and collaboration on some of the experiments captured in the chapter.


  1. 1.
    Wang W, Singh S, Zeng DL, King K, Nema S. Antibody structure, instability, and formulation. J Pharm Sci. 2007;96(1):1–26.CrossRefGoogle Scholar
  2. 2.
    Food and Drug Administration. Lists of licensed biological products with reference product exclusivity and biosimilarity or interchangeability evaluations. In Purple Book 2019; 2019.Google Scholar
  3. 3.
    Conner J, Wuchterl D, Lopez M, Minshall B, Prusti R, Boclair D, Peterson J, Allen C. Chapter 26; The Biomanufacturing of Biotechnology Products. In: Craig Shimasaki, editor. Biotechnology Entrepreneurship. 1st ed. Elsevier, Inc., Oxford, UK. 2014;351–385Google Scholar
  4. 4.
    Harris RJ, Shire SJ, Winter C. Commercial manufacturing scale formulation and analytical characterization of therapeutic recombinant antibodies. Drug Dev Res. 2004;61:137–54.CrossRefGoogle Scholar
  5. 5.
    Wang W. Instability, stabilization and formulation of liquid protein pharmaceuticals. Int J Pharm. 1999;185:125–88.CrossRefGoogle Scholar
  6. 6.
    Rathore N, Rajan RS. Current perspectives on stability of protein drug products during formulation, fill and finish operations. Biotechnol Progress. 2008;24:504–14.CrossRefGoogle Scholar
  7. 7.
    Das N. Commercializing high concentration mAbs. BioPharm Int., 2016;29(11):47–9.Google Scholar
  8. 8.
    Shire SJ, Shahrokh Z, Liu J. Challenges in the development of high protein concentration formulations. J Pharm Sci. 2004;93:1390–402.CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Callahan DJ, Stanley B, Li Y. Control of protein particle formation during ultrafiltration/diafiltration through interfacial protection. J Pharm Sci. 2014;103(2):862–9.CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    van Reis R, Zydney A. Bioprocess membrane technology. J Membr Sci. 2007;297(1–2):16–50.CrossRefGoogle Scholar
  11. 11.
    Randolph TW, Schiltz E, Sederstrom D, Steinmann D, Mozziconacci O, Schoneich C, et al. Do not drop: mechanical shock in vials causes cavitation, protein aggregation, and particle formation. J Pharm Sci. 2015;104(2):602–11.CrossRefGoogle Scholar
  12. 12.
    Nayak A, Colandene J, Bradford V, Perkins M. Characterization of subvisible particle formation during the filling pump operation of a monoclonal antibody solution. J Pharm Sci. 2011;100(10):4198.CrossRefGoogle Scholar
  13. 13.
    Saller V, Matilainen J, Grauschopf U, Bechtold-Peters K, Mahler HC, Friess W. Particle shedding from peristaltic pump tubing in biopharmaceutical drug product manufacturing. J Pharm Sci. 2015;104(4):1440–50.CrossRefGoogle Scholar
  14. 14.
    Vandanjon L, Rossignol N, Jaouen P, Robert JM, Quéméneur F. Effects of shear on two microalgae species. Contribution of pumps and valves in tangential flow filtration systems. Biotechnology and bioengineering. Biotechnol Bioeng. 1999;63(1):1–9.CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Biddlecombe JG, Craig AV, Zhang H, Uddin S, Mulot S, Fish BC, et al. Determining antibody stability: creation of solid-liquid interfacial effects within a high shear environment. Biotechnol Prog. 2007;23:1218–22.PubMedPubMedCentralGoogle Scholar
  16. 16.
    Biddlecombe JG, Smith G, Uddin S, Mulot S, Spencer D, Gee C, et al. Factors influencing antibody stability at solid-liquid interfaces in a high shear environment. Biotechnol Prog. 2009;25(5):1499–507.CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Charm SE, Wong BL. Shear effects on enzymes. Enzym Microb Technol. 1981;32(2):111–8.CrossRefGoogle Scholar
  18. 18.
    Jasepe J, Hage SJ. Do protein molecules unfold in a simple shear flow? Biophys J. 2006;9(19):3415–24.CrossRefGoogle Scholar
  19. 19.
    Maa YF, Hsu CC. Protein denaturation by combined effect of shear and air-liquid interface. Biotechnol Bioeng. 1997;54(6):503–12.CrossRefGoogle Scholar
  20. 20.
    Thomas CR, Geer D. Effects of shear on proteins in solution. Biotechnol Lett. 2011;33(3):443–56.CrossRefGoogle Scholar
  21. 21.
    Novoselsky OY, Kolgonova LI. Cavitation model of choked flow of underheated water. Atomic Energy. 2012;112(1):1–13.CrossRefGoogle Scholar
  22. 22.
    Torisu T, Maruno T, Hamaji Y, Ohkubo T, Uchiyama S. Synergistic effect of cavitation and agitation on protein aggregation. J Pharm Sci. 2017;106(2):521–9.CrossRefGoogle Scholar
  23. 23.
    Carpenter JF, Kendrick BS, Chang BS, Manning MC, Randolph TW. Inhibition of stress-induced aggregation of protein therapeutics. Methods Enzymol. 1999;309:236–55.CrossRefGoogle Scholar
  24. 24.
    Ghadge RS, Swant SB, Joshi JB. Enzyme deactivation in a bubble column, a stirred vessel and an inclined plane. Chem Eng Sci. 2003;58(23–24):5125–34.CrossRefGoogle Scholar
  25. 25.
    Graham DE, Phillips MC. Proteins at liquid interfaces. V. Shear properties. J Colloid Interface Sci. 1980;76(1):240–50.CrossRefGoogle Scholar
  26. 26.
    Green RJ, Hopkinson I, Jones RAL. Unfolding and intermolecular association in globular proteins adsorbed at interfaces. Langmuir. 1999;15(15):5102–10.CrossRefGoogle Scholar
  27. 27.
    Koepf E, Eisele S, Schroeder R, Brezesinski G, Friess W. Notorious but not understood: how liquid-air interfacial stress triggers protein aggregation. Int J Pharm. 2018;537(1–2):202–12.CrossRefGoogle Scholar
  28. 28.
    Koepf E, Richert M, Braunschweig B, Schroeder R, Brezesinski G, Friess W. Impact of formulation pH on physicochemical protein characteristics at the liquid-air interface. Int J Pharm. 2018;541(1–2):234–45.CrossRefGoogle Scholar
  29. 29.
    Koepf E, Schroeder R, Brezesinski G, Friess W. The film tells the story: physical-chemical characteristics of IgG at the liquid-air interface. Eur J Pharm Biopharm. 2017;119:396–407.CrossRefGoogle Scholar
  30. 30.
    Ogunyankin MO, Deshmukh S, Krause M, Carvalho T, Huang M, Ilott A, Remy B, Khossravi M. Scale-down tools to assess the impact of interfacial and shear stress on biologics drug product. AAPS Pham SciTech. Submitted. 2019;20:1–9.Google Scholar
  31. 31.
    ICH QB 1996, Stability Testing: Photostability Testing of New Drug Substances and Products, 1996.
  32. 32.
    Steinmann D, Ji JA, Wang YJ, Schoneich C. Photodegradation of human growth hormone: a novel backbone cleavage between Glu-88 and Pro-89. Mol Pharm. 2013;10(7):2693–706.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Haywood J, Mozziconacci O, Allegre KM, Kerwin BA, Schoneich C. Light induced conversion of Trp to Gly and Gly hydroperoxide in IgG1. Mol Pharm. 2013;10(3):1146–50.CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Liu M, Zhang Z, Cheetham J, Ren D, Zhou ZS. Discovery and characterization of a photo-oxidative histidine-histidine cross-link in IgG1 antibody utilizing (1)(8) O-labeling and mass spectrometry. Anal Chem. 2014;86(10):4940–8.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Sreedhara A, Yin J, Joyce M, Lau K, Wecksler AT, Deperalta G, Yi L, John Wang Y, Kabakoff B, Kishore RSK. Effect of ambient light on IgG1 monoclonal antibodies during drug product processing and development. Eur J Pharm Biopharm. 2016;100:38–46.CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Tanushevski A, Rendevski S. Energy efficiency comparison between compact fluorescent lamp and common light bulb. Eur J Phys Educ. 2016;7(2):1309–7202.Google Scholar
  37. 37.
    More H. Effect of light source and UV quotient on monoclonal antibodies stability during manufacturing and storage. The Bioprocessing (Summit, Aug 2018).Google Scholar
  38. 38.
    Mallaney M, Wang S, Sreedhara A. Effect of ambient light on monoclonal antibody product quality during small-scale mammalian cell culture process in clear glass bioreactors. Biotechnol Progress. 2014;30:562–70.CrossRefGoogle Scholar
  39. 39.
    Zhou S, Schoneich C, Singh S. Biologics formulation factors affecting metal Leachables from stainless steel. AAPS Pharm Sci Tech. 2011;12:411–21.CrossRefGoogle Scholar
  40. 40.
    Markovic I. Challenges associated with extractables and/or leachables substances in therapeutic biologic protein products. Am Pharm Rev. 2006;9(6):20–7.Google Scholar
  41. 41.
    Hovorka SW, et al. Oxidative degradation of pharmaceuticals: theory, mechanisms and inhibition. J Pharm Sci. 2001;90(1):58–69.Google Scholar
  42. 42.
    Davies KJ, Delsignore ME. J Biol Chem. 1987;262(20):9908–13.PubMedPubMedCentralGoogle Scholar
  43. 43.
    Sadineni V, Chandrasekharan S, Nassar MN. Implications of trace levels of redox-active metals in drug-product formulation. Biopharm Int. 2014;27(4):30–32.Google Scholar
  44. 44.
    Singh SK. Storage consideration as part of the formulation development program for biologics. Am Pharma Rev. 2007;10(3):26–33.Google Scholar
  45. 45.
    Singh S, Kolhe P, Wang W, Nema S. Large-scale freezing of biologics. BioProcess Int. 2009;7:32–44.Google Scholar
  46. 46.
    Bhatnagar B, Bogner RH, Pikal MJ. Protein stability during freezing: separation of stresses and mechanisms of protein stabilization. Pharm Dev Technol. 2007;12:505–23.CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Lashmar UT, Vanderburgh M, Little SJ. Bulk freeze-thawing of macromolecules. BioProcess Int. 2007;5:44–54.Google Scholar
  48. 48.
    Kolhe P, Badkar A. Protein and solute distribution in drug substance containers during frozen storage and post-thawing: a tool to understand and define freezing-thawing parameters in biotechnology process development. Biotechnol Prog. 2011;27:494–504.CrossRefPubMedPubMedCentralGoogle Scholar
  49. 49.
    Kolhe P, Badkar A. Protein and solute distribution in drug substance containers during frozen storage and post-thawing: a tool to understand and define freezing-thawing parameters in biotechnology process development. Biotechnol Prog. 2011;27:494–504.CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Kolhe P, Holding E, Lary A, Chico S, Singh S. Large scale freezing of biologics: understanding protein and solute concentration changes in a Cryovessel – Part I. Biopharm Int. 2010;23:53–60.Google Scholar
  51. 51.
    Gómez G, Pikal M, Rodríguez-Hornedo N. Effect of initial buffer composition on pH changes during far-from-equilibrium freezing of sodium phosphate buffer solutions. Pharm Res. 2001;18:90–7.CrossRefGoogle Scholar
  52. 52.
    Jena S, Horn J, Suryanarayanan R, et al. Effects of excipient interactions on the state of the freeze-concentrate and protein stability. Pharm Res. 2017;34:462.Google Scholar
  53. 53.
    Desai KG, Aaron Pruett W, Martin PJ, Colandene JD, Nesta DP. Impact of manufacturing-scale freeze-thaw conditions on a mAb solution. Biopharm Int. 2017;30:30–6.Google Scholar
  54. 54.
    Jameel F, Hershenson S. Formulation and process development strategies for manufacturing biopharmaceuticals. Hoboken: Wiley; 2010.CrossRefGoogle Scholar
  55. 55.
    Sumit K, Shikha T, Deepika T, Ashish B. A quantitative approach for pharmaceutical quality by design patterns. Inveti Rapid: Pharm Anal Qual Assur. 2012;2012(4):1–8.Google Scholar
  56. 56.
    Rathore AS, Sharma C, Persad AA. Use of computational fluid dynamics as a tool for establishing process design space for mixing in a bioreactor. Biotechnol Prog. 2012;28(2):382–91.CrossRefGoogle Scholar
  57. 57.
    Vlaev SD, Georgiev D, Nikov I, Elqotbi M. The CFD approach for shear analysis of mixing reactor: verification and examples of use. J Eng Sci Technol. 2007;2(2):177–187.Google Scholar
  58. 58.
    Kim D, Stoesser T, Kim J-H. Modeling aspects of flow and solute transport simulations in water disinfection tanks. Appl Math Model. 2013;37(16–17):8039–50.CrossRefGoogle Scholar
  59. 59.
    Pillai SA, Chobisa D, Urimi D, Ravindra N. Filters and Filtration: A Review of Mechanisms That Impact Cost, Product Quality and Patient Safety. J Pharm Sci Res. 2016; 8(5):271–8.Google Scholar
  60. 60.
    Gasch J, Oertel R, Leopold CS, Knoth H. Contamination of 0.2-micrometer infusion filters by N,N-dimethylacrylamide. J Crit Care. 2010;25:172.Google Scholar
  61. 61.
    Zheng S, Smith P, Burton L, Adams M. Sensitive fluorescence-based method for the rapid determination of polysorbate-80 content in therapeutic monoclonal antibody products. Pharm Dev Technol. 2015;20(7):872–6.Google Scholar
  62. 62.
    Cromwell ME, Hilario E, Jacobson F. Protein aggregation and bioprocessing. AAPS J. 2006;8:E572–9.CrossRefPubMedPubMedCentralGoogle Scholar
  63. 63.
    Tyagi AK, Randolph TW, Dong A, Maloney KM, Hitscherich C, Carpenter JF. IgG particle formation during filling pump operation: a case study of heterogeneous nucleation on stainless steel nanoparticles. J Pharm Sci. 2009;98(1):94–104.CrossRefGoogle Scholar
  64. 64.
    Shieu W, Lamar D, Stauch OB, Maa YF. Filling of high-concentration monoclonal antibody formulations: investigating underlying mechanisms that affect precision of low-volume fill by peristaltic pump. PDA J Pharm Sci Technol. 2016;70(2):143–56.CrossRefGoogle Scholar
  65. 65.
    Hanslip S, Desai KG, Palmer M, Kemp I, Bell S, Schofield P, Varma P, Roche F, Colandene JD, Nesta DP. Syringe filling of a high-concentration mAb formulation: experimental, theoretical, and computational evaluation of filling process parameters that influence the propensity for filling needle clogging. J Pharm Sci. 2019;108(3):1130–8.CrossRefGoogle Scholar

Copyright information

© American Association of Pharmaceutical Scientists 2020

Authors and Affiliations

  1. 1.Pharmaceutical Sciences, Merck & Co. Inc.KenilworthUSA
  2. 2.Drug Product Science & Technology, Bristol-Myers Squibb, Co.New BrunswickUSA

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