Skip to main content

Advertisement

SpringerLink
Log in

A High Threshold of Biotherapeutic Aggregate Numbers is Needed to Induce an Immunogenic Response In Vitro, In Vivo, and in the Clinic

  • Original Research Article
  • Published:
Pharmaceutical Research Aims and scope Submit manuscript

Abstract

Background and Purpose

There is concern that subvisible aggregates in biotherapeutic drug products pose a risk to patient safety. We investigated the threshold of biotherapeutic aggregates needed to induce immunogenic responses.

Methods and Results

Highly aggregated samples were tested in cell-based assays and induced cellular responses in a manner that depended on the number of particles. The threshold of immune activation varied by disease state (cancer, rheumatoid arthritis, allergy), concomitant therapies, and particle number. Compared to healthy donors, disease state patients showed an equal or lower response at the late phase (7 days), suggesting they may not have a higher risk of responding to aggregates. Xeno-het mice were used to assess the threshold of immune activation in vivo. Although highly aggregated samples (~ 1,600,000 particles/mL) induced a weak and transient immunogenic response in mice, a 100-fold dilution of this sample (~ 16,000 particles/mL) did not induce immunogenicity. To confirm this result, subvisible particles (up to ~ 18,000 particles/mL, containing aggregates and silicone oil droplets) produced under representative administration practices (created upon infusion of a drug product through an IV catheter) did not induce a response in cell-based assays or appear to increase the rate of adverse events or immunogenicity during phase 3 clinical trials.

Conclusion

The ability of biotherapeutic aggregates to elicit an immune response in vitro, in vivo, and in the clinic depends on high numbers of particles. This suggests that there is a high threshold for aggregates to induce an immunogenic response which is well beyond that seen in standard biotherapeutic drug products.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Data Availability

All data needed to evaluate the conclusions in the paper are present in the paper and/or the supplementary materials. Upon request, and subject to review and legal requirements, Amgen may provide the data that support the findings of this study.

References

  1. Wang Z, Wang G, Lu H, Li H, Tang M, Tong A. Development of therapeutic antibodies for the treatment of diseases. Mol Biomed. 2022;3(1):35.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Lu RM, Hwang YC, Liu IJ, Lee CC, Tsai HZ, Li HJ, Wu HC. Development of therapeutic antibodies for the treatment of diseases. J Biomed Sci. 2020;27(1):1.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. The Antibody Society. Therapeutic monoclonal antibodies approved or in review in the US. 2023. Available from: www.antibodysociety.org/resources/approved-antibodies. Accessed 8 Mar 2023.

  4. Kaplon H, Chenoweth A, Crescioli S, Reichert JM. Antibodies to watch in 2022. MAbs. 2022;14(1):2014296.

    Article  PubMed  PubMed Central  Google Scholar 

  5. Zhou S, Liu M, Ren F, Meng X, Yu J. The landscape of bispecific T cell engager in cancer treatment. Biomark Res. 2021;9(1):38.

    Article  PubMed  PubMed Central  Google Scholar 

  6. Goebeler ME, Bargou RC. T cell-engaging therapies - BiTEs and beyond. Nat Rev Clin Oncol. 2020;17(7):418–34.

    Article  PubMed  Google Scholar 

  7. Singh S, Kumar NK, Dwiwedi P, Charan J, Kaur R, Sidhu P, Chugh VK. Monoclonal Antibodies: A Review. Curr Clin Pharmacol. 2018;13(2):85–99.

    Article  PubMed  Google Scholar 

  8. Kotsovilis S, Andreakos E. Therapeutic human monoclonal antibodies in inflammatory diseases. Methods Mol Biol. 2014;1060:37–59.

    Article  PubMed  Google Scholar 

  9. Landolina N, Levi-Schaffer F. Monoclonal antibodies: the new magic bullets for allergy: IUPHAR Review 17. Br J Pharmacol. 2016;173(5):793–803.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Arepalli S, Kaiser PK. Pipeline therapies for neovascular age related macular degeneration. Int J Retina Vitreous. 2021;7(1):55.

    Article  PubMed  PubMed Central  Google Scholar 

  11. Charles A, Pozo-Rosich P. Targeting calcitonin gene-related peptide: a new era in migraine therapy. Lancet. 2019;394(10210):1765–74.

    Article  CAS  PubMed  Google Scholar 

  12. Chang B, Quan Q, Li Y, Qiu H, Peng J, Gu Y. Treatment of Osteoporosis, with a Focus on 2 Monoclonal Antibodies. Med Sci Monit. 2018;24:8758–66.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Ji E, Lee S. Antibody-based therapeutics for atherosclerosis and cardiovascular diseases. Int J Mol Sci. 2021;22(11):5770.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Pento JT. Monoclonal Antibodies for the Treatment of Cancer. Anticancer Res. 2017;37(11):5935–9.

    CAS  PubMed  Google Scholar 

  15. Jin S, Sun Y, Liang X, Gu X, Ning J, Xu Y, Chen S, Pan L. Emerging new therapeutic antibody derivatives for cancer treatment. Signal Transduct Target Ther. 2022;7(1):39.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Elgundi Z, Reslan M, Cruz E, Sifniotis V, Kayser V. The state-of-play and future of antibody therapeutics. Adv Drug Deliv Rev. 2017;122:2–19.

    Article  CAS  PubMed  Google Scholar 

  17. Rehder DS, Chelius D, McAuley A, Dillon TM, Xiao G, Crouse-Zeineddini J, Vardanyan L, Perico N, Mukku V, Brems DN, Matsumura M, Bondarenko PV. Isomerization of a single aspartyl residue of anti-epidermal growth factor receptor immunoglobulin gamma2 antibody highlights the role avidity plays in antibody activity. Biochemistry. 2008;47(8):2518–30.

    Article  CAS  PubMed  Google Scholar 

  18. Yan Y, Wei H, Fu Y, Jusuf S, Zeng M, Ludwig R, Krystek SR Jr, Chen G, Tao L, Das TK. Isomerization and Oxidation in the Complementarity-Determining Regions of a Monoclonal Antibody: A Study of the Modification-Structure-Function Correlations by Hydrogen-Deuterium Exchange Mass Spectrometry. Anal Chem. 2016;88(4):2041–50.

    Article  CAS  PubMed  Google Scholar 

  19. Joubert MK, Hokom M, Eakin C, Zhou L, Deshpande M, Baker MP, Goletz TJ, Kerwin BA, Chirmule N, Narhi LO, Jawa V. Highly aggregated antibody therapeutics can enhance the in vitro innate and late-stage T-cell immune responses. J Biol Chem. 2012;287(30):25266–79.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Dingman R, Balu-Iyer SV. Immunogenicity of Protein Pharmaceuticals. J Pharm Sci. 2019;108(5):1637–54.

    Article  CAS  PubMed  Google Scholar 

  21. International Conference on Harmonisation. Pharmaceutical Development Q8(R2) Current Step 4 version. International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use. 2009. Available from: https://database.ich.org/sites/default/files/Q8_R2_Guideline.pdf. Accessed 8 Mar 2023

  22. Xu Y, Wang D, Mason B, Rossomando T, Li N, Liu D, Cheung JK, Xu W, Raghava S, Katiyar A, Nowak C, Xiang T, Dong DD, Sun J, Beck A, Liu H. Structure, heterogeneity and developability assessment of therapeutic antibodies. MAbs. 2019;11(2):239–64.

    Article  CAS  PubMed  Google Scholar 

  23. Torkashvand F, Vaziri B. Main Quality Attributes of Monoclonal Antibodies and Effect of Cell Culture Components. Iran Biomed J. 2017;21(3):131–41.

    Article  PubMed  PubMed Central  Google Scholar 

  24. Rathore AS, Winkle H. Quality by design for biopharmaceuticals. Nat Biotechnol. 2009;27(1):26–34.

    Article  CAS  PubMed  Google Scholar 

  25. Yu LX. Pharmaceutical quality by design: product and process development, understanding, and control. Pharm Res. 2008;25(4):781–91.

    Article  CAS  PubMed  Google Scholar 

  26. Yu LX, Amidon G, Khan MA, Hoag SW, Polli J, Raju GK, Woodcock J. Understanding pharmaceutical quality by design. AAPS J. 2014;16(4):771–83.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Hanna-Brown MEJ, Katzenbach S, Rignall A, Gervais A, Hoffmann J, Wypych J, McLaughlin K, Borman P, Grosche O, Hamilton P, Uhlich T, Finkler C, Liebelt K. Analytical Procedure Lifecycle Management: Current Status and Opportunities. Pharm Technol. 2018;42(12):18–23.

    Google Scholar 

  28. Pharmacopeia US. Analytical procedure Lifecycle - Draft. 2022. Available from: https://www.uspnf.com/sites/default/files/usp_pdf/EN/USPNF/usp-nf-notices/gc-1220-pre-post-20210924.pdf. Accessed 8 Mar 2023.

  29. Weitzel J, Pappa H, Banik GM, Barker AR, Bladen E, Chirmule N, DeFeo J, Devine J, Emrick S, Hout TK, Levy MS, Mahlangu GN, Rellahan B, Venema J, Workman W. Understanding Quality Paradigm Shifts in the Evolving Pharmaceutical Landscape: Perspectives from the USP Quality Advisory Group. AAPS J. 2021;23(6):112.

    Article  PubMed  Google Scholar 

  30. Verch T, Campa C, Chery CC, Frenkel R, Graul T, Jaya N, Nakhle B, Springall J, Starkey J, Wypych J, Ranheim T. Analytical Quality by Design, Life Cycle Management, and Method Control. AAPS J. 2022;24(1):34.

    Article  PubMed  Google Scholar 

  31. Narhi LO, Corvari V, Ripple DC, Afonina N, Cecchini I, Defelippis MR, Garidel P, Herre A, Koulov AV, Lubiniecki T, Mahler HC, Mangiagalli P, Nesta D, Perez-Ramirez B, Polozova A, Rossi M, Schmidt R, Simler R, Singh S, Spitznagel TM, Weiskopf A, Wuchner K. Subvisible (2–100 μm) Particle Analysis During Biotherapeutic Drug Product Development: Part 1. Considerations and Strategy J Pharm Sci. 2015;104(6):1899–908.

    Article  CAS  PubMed  Google Scholar 

  32. Bansal R, Gupta S, Rathore AS. Analytical Platform for Monitoring Aggregation of Monoclonal Antibody Therapeutics. Pharm Res. 2019;36(11):152.

    Article  PubMed  Google Scholar 

  33. Singh SK, Afonina N, Awwad M, Bechtold-Peters K, Blue JT, Chou D, Cromwell M, Krause HJ, Mahler HC, Meyer BK, Narhi L, Nesta DP, Spitznagel T. An industry perspective on the monitoring of subvisible particles as a quality attribute for protein therapeutics. J Pharm Sci. 2010;99(8):3302–21.

    Article  CAS  PubMed  Google Scholar 

  34. Rosenberg AS. Effects of protein aggregates: an immunologic perspective. AAPS J. 2006;8(3):E501-507.

    Article  PubMed  PubMed Central  Google Scholar 

  35. Wang W. Protein aggregation and its inhibition in biopharmaceutics. Int J Pharm. 2005;289(1–2):1–30.

    Article  CAS  PubMed  Google Scholar 

  36. Filipe VHA, Schellekens H, Jiskoot W. Aggregation of therapeutic proteins. New Jersey: Wiley; 2010.

    Book  Google Scholar 

  37. Wang W, Singh SK, Li N, Toler MR, King KR, Nema S. Immunogenicity of protein aggregates–concerns and realities. Int J Pharm. 2012;431(1–2):1–11.

    CAS  PubMed  Google Scholar 

  38. Ratanji KD, Derrick JP, Dearman RJ, Kimber I. Immunogenicity of therapeutic proteins: influence of aggregation. J Immunotoxicol. 2014;11(2):99–109.

    Article  CAS  PubMed  Google Scholar 

  39. Moussa EM, Panchal JP, Moorthy BS, Blum JS, Joubert MK, Narhi LO, Topp EM. Immunogenicity of therapeutic protein aggregates. J Pharm Sci. 2016;105(2):417–30.

    Article  CAS  PubMed  Google Scholar 

  40. U.S. Food and Drug Administration. Inspection of injectable products for visible particulates guidance for industry draft guidance. 2021. Available from: https://www.fda.gov/media/154868/download. Accessed 8 Mar 2023.

  41. USP. Particulate Matter in Injections. USP 35; U.S. Pharmacopeial Convention. Rockville, MD; 2012. pp. 339–342.

  42. Narhi LO, Luo Q, Wypych J, Torosantucci R, Hawe A, Fujimori K, Nashed-Samuel Y, Jawa V, Joubert MK, Jiskoot W. Chemical and Biophysical Characteristics of Monoclonal Antibody Solutions Containing Aggregates Formed during Metal Catalyzed Oxidation. Pharm Res. 2017;34(12):2817–28.

    Article  CAS  PubMed  Google Scholar 

  43. Joubert MK, Luo Q, Nashed-Samuel Y, Wypych J, Narhi LO. Classification and characterization of therapeutic antibody aggregates. J Biol Chem. 2011;286(28):25118–33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Luo Q, Joubert MK, Stevenson R, Ketchem RR, Narhi LO, Wypych J. Chemical Modifications in Therapeutic Protein Aggregates Generated under Different Stress Conditions. J Biol Chem. 2011;286(28):25134–44.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Jiskoot W, Kijanka G, Randolph TW, Carpenter JF, Koulov AV, Mahler HC, Joubert MK, Jawa V, Narhi LO. Mouse Models for Assessing Protein Immunogenicity: Lessons and Challenges. J Pharm Sci. 2016;105(5):1567–75.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Bessa J, Boeckle S, Beck H, Buckel T, Schlicht S, Ebeling M, Kiialainen A, Koulov A, Boll B, Weiser T, Singer T, Rolink AG, Iglesias A. The immunogenicity of antibody aggregates in a novel transgenic mouse model. Pharm Res. 2015;32(7):2344–59.

    Article  CAS  PubMed  Google Scholar 

  47. Philo JS, Arakawa T. Mechanisms of protein aggregation. Curr Pharm Biotechnol. 2009;10(4):348–51.

    Article  CAS  PubMed  Google Scholar 

  48. Lumry R, Eyring H. Conformation Changes of Proteins. J Phys Chem. 1954;58(2):110–20.

    Article  CAS  Google Scholar 

  49. Roberts CJ. Therapeutic protein aggregation: mechanisms, design, and control. Trends Biotechnol. 2014;32(7):372–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Le Basle Y, Chennell P, Tokhadze N, Astier A, Sautou V. Physicochemical Stability of Monoclonal Antibodies: A Review. J Pharm Sci. 2020;109(1):169–90.

    Article  PubMed  Google Scholar 

  51. Vázquez-Rey M, Lang DA. Aggregates in monoclonal antibody manufacturing processes. Biotechnol Bioeng. 2011;108(7):1494–508.

    Article  PubMed  Google Scholar 

  52. Narhi LO, Schmit J, Bechtold-Peters K, Sharma D. Classification of protein aggregates. J Pharm Sci. 2012;101(2):493–8.

    Article  CAS  PubMed  Google Scholar 

  53. Das TK. Protein particulate detection issues in biotherapeutics development–current status. AAPS PharmSciTech. 2012;13(2):732–46.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Krayukhina E, Tsumoto K, Uchiyama S, Fukui K. Effects of syringe material and silicone oil lubrication on the stability of pharmaceutical proteins. J Pharm Sci. 2015;104(2):527–35.

    Article  CAS  PubMed  Google Scholar 

  55. Thirumangalathu R, Krishnan S, Ricci MS, Brems DN, Randolph TW, Carpenter JF. Silicone oil- and agitation-induced aggregation of a monoclonal antibody in aqueous solution. J Pharm Sci. 2009;98(9):3167–81.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Majumdar S, Ford BM, Mar KD, Sullivan VJ, Ulrich RG, D’Souza AJ. Evaluation of the effect of syringe surfaces on protein formulations. J Pharm Sci. 2011;100(7):2563–73.

    Article  CAS  PubMed  Google Scholar 

  57. Jones LS, Kaufmann A, Middaugh CR. Silicone oil induced aggregation of proteins. J Pharm Sci. 2005;94(4):918–27.

    Article  CAS  PubMed  Google Scholar 

  58. Ludwig DB, Carpenter JF, Hamel JB, Randolph TW. Protein adsorption and excipient effects on kinetic stability of silicone oil emulsions. J Pharm Sci. 2010;99(4):1721–33.

    Article  CAS  PubMed  Google Scholar 

  59. Mehta SB, Lewus R, Bee JS, Randolph TW, Carpenter JF. Gelation of a monoclonal antibody at the silicone oil-water interface and subsequent rupture of the interfacial gel results in aggregation and particle formation. J Pharm Sci. 2015;104(4):1282–90.

    Article  CAS  PubMed  Google Scholar 

  60. Gerhardt A, McGraw NR, Schwartz DK, Bee JS, Carpenter JF, Randolph TW. Protein aggregation and particle formation in prefilled glass syringes. J Pharm Sci. 2014;103(6):1601–12.

    Article  CAS  PubMed  Google Scholar 

  61. Kossovsky N, Heggers JP, Robson MC. Experimental demonstration of the immunogenicity of silicone-protein complexes. J Biomed Mater Res. 1987;21(9):1125–33.

    Article  CAS  PubMed  Google Scholar 

  62. Gaitonde P, Balu-Iyer SV. In vitro immunogenicity risk assessment of therapeutic proteins in preclinical setting. Methods Mol Biol. 2011;716:267–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Kraus T, Lubitz A, Schließer U, Giese C, Reuschel J, Brecht R, Engert J, Winter G. Evaluation of a 3D Human Artificial Lymph Node as Test Model for the Assessment of Immunogenicity of Protein Aggregates. J Pharm Sci. 2019;108(7):2358–66.

    Article  CAS  PubMed  Google Scholar 

  64. Telikepalli S, Shinogle HE, Thapa PS, Kim JH, Deshpande M, Jawa V, Middaugh CR, Narhi LO, Joubert MK, Volkin DB. Physical characterization and in vitro biological impact of highly aggregated antibodies separated into size-enriched populations by fluorescence-activated cell sorting. J Pharm Sci. 2015;104(5):1575–91.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. Chirmule N, Jawa V, Meibohm B. Immunogenicity to therapeutic proteins: impact on PK/PD and efficacy. Aaps j. 2012;14(2):296–302.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Ahmadi M, Bryson CJ, Cloake EA, Welch K, Filipe V, Romeijn S, Hawe A, Jiskoot W, Baker MP, Fogg MH. Small amounts of sub-visible aggregates enhance the immunogenic potential of monoclonal antibody therapeutics. Pharm Res. 2015;32(4):1383–94.

    Article  CAS  PubMed  Google Scholar 

  67. Thorlaksen C, Schultz HS, Gammelgaard SK, Jiskoot W, Hatzakis NS, Nielsen FS, Solberg H, Foderà V, Bartholdy C, Groenning M. In vitro and in vivo immunogenicity assessment of protein aggregate characteristics. Int J Pharm. 2023;631: 122490.

    Article  CAS  PubMed  Google Scholar 

  68. Moussa EM, Kotarek J, Blum JS, Marszal E, Topp EM. Physical Characterization and Innate Immunogenicity of Aggregated Intravenous Immunoglobulin (IGIV) in an In Vitro Cell-Based Model. Pharm Res. 2016;33(7):1736–51.

    Article  CAS  PubMed  Google Scholar 

  69. Joubert MK, Deshpande M, Yang J, Reynolds H, Bryson C, Fogg M, Baker MP, Herskovitz J, Goletz TJ, Zhou L, Moxness M, Flynn GC, Narhi LO, Jawa V. Use of In Vitro Assays to Assess Immunogenicity Risk of Antibody-Based Biotherapeutics. PLoS ONE. 2016;11(8): e0159328.

    Article  PubMed  PubMed Central  Google Scholar 

  70. Janeway CATP Jr, Walport M, Shlomchik MJ. Immunobiology: The Immune System in Health and Disease. New York: Garland Science; 2001.

    Google Scholar 

  71. Cyster JG, Allen CDC. B Cell Responses: Cell Interaction Dynamics and Decisions. Cell. 2019;177(3):524–40.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. Delves PJ, Roitt IM. The immune system. First of two parts. The New England journal of medicine. 2000;343(1):37–49.

    Article  CAS  PubMed  Google Scholar 

  73. Delves PJ, Roitt IM. The immune system. Second of two parts. The New England journal of medicine. 2000;343(2):108–17.

    Article  CAS  PubMed  Google Scholar 

  74. Nicholson LB. The immune system. Essays Biochem. 2016;60(3):275–301.

    Article  PubMed  PubMed Central  Google Scholar 

  75. den Haan JM, Arens R, van Zelm MC. The activation of the adaptive immune system: cross-talk between antigen-presenting cells, T cells and B cells. Immunology letters. 2014;162(2 Pt B):103–12.

    Article  Google Scholar 

  76. Pichler WJ. Adverse side-effects to biological agents. Allergy. 2006;61(8):912–20.

    Article  CAS  PubMed  Google Scholar 

  77. Fogdell-Hahn A. Antidrug Antibodies: B Cell Immunity Against Therapy. Scand J Immunol. 2015;82(3):184–90.

    Article  CAS  PubMed  Google Scholar 

  78. Krishna M, Nadler SG. Immunogenicity to Biotherapeutics - The Role of Anti-drug Immune Complexes. Front Immunol. 2016;7:21.

    Article  PubMed  PubMed Central  Google Scholar 

  79. Goins CL, Chappell CP, Shashidharamurthy R, Selvaraj P, Jacob J. Immune complex-mediated enhancement of secondary antibody responses. Journal of immunology (Baltimore, Md : 1950). 2010;184(11):6293–8.

    Article  CAS  PubMed  Google Scholar 

  80. Bi V, Jawa V, Joubert MK, Kaliyaperumal A, Eakin C, Richmond K, Pan O, Sun J, Hokom M, Goletz TJ, Wypych J, Zhou L, Kerwin BA, Narhi LO, Arora T. Development of a human antibody tolerant mouse model to assess the immunogenicity risk due to aggregated biotherapeutics. J Pharm Sci. 2013;102(10):3545–55.

    Article  CAS  PubMed  Google Scholar 

  81. Tokuda JM, Xie J, Jawa V, Hawkins JM, Ferbas J, Joh NH, Joubert MK. Use of In Vitro Human Skin Models to Assess Potential Immune Activation In Response to Biotherapeutic Attributes and Process-related Impurities. J Pharm Sci. 2022;111(4):1012–23.

    Article  CAS  PubMed  Google Scholar 

  82. Kinderman F, Yerby B, Jawa V, Joubert MK, Joh NH, Malella J, Herskovitz J, Xie J, Ferbas J, McBride HJ. Impact of Precipitation of Antibody Therapeutics After Subcutaneous Injection on Pharmacokinetics and Immunogenicity. J Pharm Sci. 2019;108(6):1953–63.

    Article  CAS  PubMed  Google Scholar 

  83. Joh NH, Thomas L, Christian TR, Verlinsky A, Jiao N, Allotta N, Jawa V, Cao S, Narhi LO, Joubert MK. Silicone Oil Particles in Prefilled Syringes With Human Monoclonal Antibody, Representative of Real-World Drug Products, Did Not Increase Immunogenicity in In Vivo and In Vitro Model Systems. J Pharm Sci. 2020;109(1):845–53.

    Article  CAS  PubMed  Google Scholar 

  84. Morgan H, Tseng SY, Gallais Y, Leineweber M, Buchmann P, Riccardi S, Nabhan M, Lo J, Gani Z, Szely N, Zhu CS, Yang M, Kiessling A, Vohr HW, Pallardy M, Aswad F, Turbica I. Evaluation of in vitro Assays to Assess the Modulation of Dendritic Cells Functions by Therapeutic Antibodies and Aggregates. Front Immunol. 2019;10:601.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  85. Boll B, Bessa J, Folzer E, Ríos Quiroz A, Schmidt R, Bulau P, Finkler C, Mahler HC, Huwyler J, Iglesias A, Koulov AV. Extensive Chemical Modifications in the Primary Protein Structure of IgG1 Subvisible Particles Are Necessary for Breaking Immune Tolerance. Mol Pharm. 2017;14(4):1292–9.

    Article  CAS  PubMed  Google Scholar 

  86. Gallucci S, Matzinger P. Danger signals: SOS to the immune system. Curr Opin Immunol. 2001;13(1):114–9.

    Article  CAS  PubMed  Google Scholar 

  87. Rombach-Riegraf V, Karle AC, Wolf B, Sorde L, Koepke S, Gottlieb S, Krieg J, Djidja MC, Baban A, Spindeldreher S, Koulov AV, Kiessling A. Aggregation of human recombinant monoclonal antibodies influences the capacity of dendritic cells to stimulate adaptive T-cell responses in vitro. PLoS ONE. 2014;9(1): e86322.

    Article  PubMed  PubMed Central  Google Scholar 

  88. Ronnelid J, Tejde A, Mathsson L, Nilsson-Ekdahl K, Nilsson B. Immune complexes from SLE sera induce IL10 production from normal peripheral blood mononuclear cells by an FcgammaRII dependent mechanism: implications for a possible vicious cycle maintaining B cell hyperactivity in SLE. Ann Rheum Dis. 2003;62(1):37–42.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  89. Haile LA, Puig M, Kelley-Baker L, Verthelyi D. Detection of innate immune response modulating impurities in therapeutic proteins. PLoS ONE. 2015;10(4): e0125078.

    Article  PubMed  PubMed Central  Google Scholar 

  90. Wullner D, Zhou L, Bramhall E, Kuck A, Goletz TJ, Swanson S, Chirmule N, Jawa V. Considerations for optimization and validation of an in vitro PBMC derived T cell assay for immunogenicity prediction of biotherapeutics. Clin Immunol. 2010;137(1):5–14.

    Article  CAS  PubMed  Google Scholar 

  91. Hermeling S, Aranha L, Damen JM, Slijper M, Schellekens H, Crommelin DJ, Jiskoot W. Structural characterization and immunogenicity in wild-type and immune tolerant mice of degraded recombinant human interferon alpha2b. Pharm Res. 2005;22(12):1997–2006.

    Article  CAS  PubMed  Google Scholar 

  92. Hermeling S, Schellekens H, Maas C, Gebbink MF, Crommelin DJ, Jiskoot W. Antibody response to aggregated human interferon alpha2b in wild-type and transgenic immune tolerant mice depends on type and level of aggregation. J Pharm Sci. 2006;95(5):1084–96.

    Article  CAS  PubMed  Google Scholar 

  93. Purohit VS, Middaugh CR, Balasubramanian SV. Influence of aggregation on immunogenicity of recombinant human Factor VIII in hemophilia A mice. J Pharm Sci. 2006;95(2):358–71.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  94. Reipert BM, van Helden PM, Schwarz HP, Hausl C. Mechanisms of action of immune tolerance induction against factor VIII in patients with congenital haemophilia A and factor VIII inhibitors. Br J Haematol. 2007;136(1):12–25.

    Article  CAS  PubMed  Google Scholar 

  95. Fradkin AH, Carpenter JF, Randolph TW. Immunogenicity of aggregates of recombinant human growth hormone in mouse models. J Pharm Sci. 2009;98(9):3247–64.

    Article  CAS  PubMed  Google Scholar 

  96. Christie M, Torres RM, Kedl RM, Randolph TW, Carpenter JF. Recombinant murine growth hormone particles are more immunogenic with intravenous than subcutaneous administration. J Pharm Sci. 2014;103(1):128–39.

    Article  CAS  PubMed  Google Scholar 

  97. Kijanka G, Bee JS, Korman SA, Wu Y, Roskos LK, Schenerman MA, Slütter B, Jiskoot W. Submicron Size Particles of a Murine Monoclonal Antibody Are More Immunogenic Than Soluble Oligomers or Micron Size Particles Upon Subcutaneous Administration in Mice. J Pharm Sci. 2018;107(11):2847–59.

    Article  CAS  PubMed  Google Scholar 

  98. Doessegger L, Banholzer ML. Clinical development methodology for infusion-related reactions with monoclonal antibodies. Clinical & translational immunology. 2015;4(7): e39.

    Article  Google Scholar 

  99. Zhang Z, Chow SY, De Guzman R, Joh NH, Joubert MK, Richardson J, Shah B, Wikstrom M, Zhou ZS, Wypych J. A Mass Spectrometric Characterization of Light-Induced Modifications in Therapeutic Proteins. J Pharm Sci. 2022;111(6):1556–64.

    Article  CAS  PubMed  Google Scholar 

  100. Yun H, Xie F, Beyl RN, Chen L, Lewis JD, Saag KG, Curtis JR. Risk of Hypersensitivity to Biologic Agents Among Medicare Patients With Rheumatoid Arthritis. Arthritis Care Res. 2017;69(10):1526–34.

    Article  CAS  Google Scholar 

  101. Gülsen A, Wedi B, Jappe U. Hypersensitivity reactions to biologics (part I): allergy as an important differential diagnosis in complex immune-derived adverse events. Allergo journal international. 2020;29(4):97–125.

    Article  PubMed  PubMed Central  Google Scholar 

  102. Gülsen A, Wedi B, Jappe U. Hypersensitivity reactions to biologics (part II): classifications and current diagnostic and treatment approaches. Allergo journal international. 2020;29(5):139–54.

    Article  Google Scholar 

  103. Strand V, Balsa A, Al-Saleh J, Barile-Fabris L, Horiuchi T, Takeuchi T, Lula S, Hawes C, Kola B, Marshall L. Immunogenicity of Biologics in Chronic Inflammatory Diseases: A Systematic Review. BioDrugs : clinical immunotherapeutics, biopharmaceuticals and gene therapy. 2017;31(4):299–316.

    Article  CAS  PubMed  Google Scholar 

  104. U.S. Food and Drug Administration. Guidance for Industry immunogenicity assessment for therapeutic protein products [Internet]. 2014. Available from: https://www.fda.gov/regulatory-information/searchfda-guidance-documents/immunogenicity-assessment-therapeutic-protein-products. Accessed 8 Mar 2023.

  105. den Engelsman J, Garidel P, Smulders R, Koll H, Smith B, Bassarab S, Seidl A, Hainzl O, Jiskoot W. Strategies for the assessment of protein aggregates in pharmaceutical biotech product development. Pharm Res. 2011;28(4):920–33.

    Article  Google Scholar 

  106. Jiao N, Barnett GV, Christian TR, Narhi LO, Joh NH, Joubert MK, Cao S. Characterization of Subvisible Particles in Biotherapeutic Prefilled Syringes: The Role of Polysorbate and Protein on the Formation of Silicone Oil and Protein Subvisible Particles After Drop Shock. J Pharm Sci. 2020;109(1):640–5.

    Article  CAS  PubMed  Google Scholar 

  107. Mahler HC, Friess W, Grauschopf U, Kiese S. Protein aggregation: pathways, induction factors and analysis. J Pharm Sci. 2009;98(9):2909–34.

    Article  CAS  PubMed  Google Scholar 

  108. Singh SK. Impact of product-related factors on immunogenicity of biotherapeutics. J Pharm Sci. 2011;100(2):354–87.

    Article  CAS  PubMed  Google Scholar 

  109. Singla A, Bansal R, Joshi V, Rathore AS. Aggregation Kinetics for IgG1-Based Monoclonal Antibody Therapeutics. Aaps j. 2016;18(3):689–702.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  110. Fesinmeyer RM, Hogan S, Saluja A, Brych SR, Kras E, Narhi LO, Brems DN, Gokarn YR. Effect of ions on agitation- and temperature-induced aggregation reactions of antibodies. Pharm Res. 2009;26(4):903–13.

    Article  CAS  PubMed  Google Scholar 

  111. Pham NB, Meng WS. Protein aggregation and immunogenicity of biotherapeutics. Int J Pharm. 2020;585: 119523.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  112. Liu L, Ammar DA, Ross LA, Mandava N, Kahook MY, Carpenter JF. Silicone oil microdroplets and protein aggregates in repackaged bevacizumab and ranibizumab: effects of long-term storage and product mishandling. Invest Ophthalmol Vis Sci. 2011;52(2):1023–34.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

The authors wish to thank Terry Goletz,Arunan Kaliyaperumal, Meghana Deshpande, Theingi Thang, Lei Zhou, and Nate Joh for useful discussions.

Funding

All work was conducted and supported by Amgen.

Author information

Author notes

  1. Martha Hokom

    Present address: Department of BioAnalytical Sciences, Genentech, Inc, South San Francisco, CA, 94080, USA

  2. Jonathan Herskovitz

    Present address: Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, 68198, USA

  3. Vibha Jawa

    Present address: Department of Pharmacometrics, Disposition & Bioanalysis, Bristol Myers Squibb, Princeton, NJ, 08543, USA

Authors and Affiliations

  1. The Department of Process Development, Amgen Inc, One Amgen Center Dr, Thousand Oaks, CA, 91320, USA

    Joseph R. Cohen, Stephen R. Brych, Siddharth Prabhu, Ahmed Elbaradei, Joshua M. Tokuda, Cathie Xiang, Xiaohong Cui, Claudia Ly, Nathan Amos, Allyson Capili, Kimya Nourbakhsh, Anthony Merlo, Julia Carreon, Jette Wypych, Linda O. Narhi & Marisa K. Joubert

  2. The Department of Biosimilars, Amgen Inc, Thousand Oaks, CA, 91320, USA

    Vivian Bi

  3. The Department of Clinical Immunology, Amgen Inc, Thousand Oaks, CA, 91320, USA

    Martha Hokom, Jonathan Herskovitz & Vibha Jawa

  4. Translational Safety and Bioanalytical Sciences, Amgen Inc, Thousand Oaks, CA, 91320, USA

    Jilin Sun & Dominador Calamba

Authors
  1. Joseph R. Cohen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Stephen R. Brych
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Siddharth Prabhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Vivian Bi
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ahmed Elbaradei
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Joshua M. Tokuda
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Cathie Xiang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Martha Hokom
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Xiaohong Cui
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Claudia Ly
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Nathan Amos
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Jilin Sun
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Dominador Calamba
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Jonathan Herskovitz
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Allyson Capili
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Kimya Nourbakhsh
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Anthony Merlo
    View author publications

    You can also search for this author in PubMed Google Scholar

  18. Julia Carreon
    View author publications

    You can also search for this author in PubMed Google Scholar

  19. Jette Wypych
    View author publications

    You can also search for this author in PubMed Google Scholar

  20. Linda O. Narhi
    View author publications

    You can also search for this author in PubMed Google Scholar

  21. Vibha Jawa
    View author publications

    You can also search for this author in PubMed Google Scholar

  22. Marisa K. Joubert
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Joseph R. Cohen or Marisa K. Joubert.

Ethics declarations

Conflict of Interest

All authors are currently, or were previously, employees, shareholders, or both employees and shareholders, of Amgen where the work was conducted.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (DOCX 52.5 KB)

Supplementary file2 (PPTX 268 KB)

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Cohen, J.R., Brych, S.R., Prabhu, S. et al. A High Threshold of Biotherapeutic Aggregate Numbers is Needed to Induce an Immunogenic Response In Vitro, In Vivo, and in the Clinic. Pharm Res 41, 651–672 (2024). https://doi.org/10.1007/s11095-024-03678-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11095-024-03678-2

Keywords

Search

Navigation