Skip to main content
SpringerLink
Log in

Demonstrating analytical similarity of a biosimilar HLX04 to bevacizumab with a panel of state-of-the-art methods and tiering of quality attributes

  • Research Paper
  • Published:
Analytical and Bioanalytical Chemistry Aims and scope Submit manuscript

Abstract

Therapeutical monoclonal antibodies are structurally and functionally complex, whereas the innovator’s manufacturing processes are proprietary. With respect to the similarity assessment, a proposed biosimilar product needs to demonstrate a side-by-side comparison between the reference product (RP) and candidate product in terms of physicochemical properties and biological activities, as well as nonclinical and clinical outcomes. Here, a comprehensive analytical similarity assessment was performed for in-depth comparison of HLX04, China-sourced Avastin® (CN-Avastin®), and Europe-sourced Avastin® (EU-Avastin®) following a tier-based quality attribute (QA) evaluation. A series of orthogonal and state-of-the-art analytical techniques were developed for the assessment. Ten lots of HLX04 were compared with 29 lots bevacizumab RP. Referred to the characterization results, HLX04 is highly similar to the RPs with respect to physicochemical properties and biological functions. In addition, HLX04 was found with similar stability and degradation behaviors upon multiple stressed conditions to bevacizumab. Minor differences were observed in glycosylation, aggregates, FcγRIIIa(F), and FcγRIIIa(V) binding activities; nevertheless, they were evaluated and demonstrated not to impact clinical outcomes. According to the reported clinical results, the totality of evidence, including the pharmacokinetic, efficacy, safety, and immunogenicity, further shows that HLX04 is similar to CN-/EU-Avastin®.

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
Fig. 8

Similar content being viewed by others

References

  1. Ferrara N, Hillan KJ, Gerber HP, Novotny W. Discovery and development of bevacizumab, an anti-VEGF antibody for treating cancer. Nat Rev Drug Discov. 2004;3(5):391–400. https://doi.org/10.1038/nrd1381.

    Article  CAS  PubMed  Google Scholar 

  2. Kramer I, Lipp HP. Bevacizumab, a humanized anti-angiogenic monoclonal antibody for the treatment of colorectal cancer. J Clin Pharm Ther. 2007;32:1–14. https://doi.org/10.1111/j.1365-2710.2007.00800.x.

    Article  CAS  PubMed  Google Scholar 

  3. U.S. Food and Drug administration (FDA). Avastin® (bevacizumab) prescribing information, Genentech, Inc., San Francisco, CA, USA 2004. https://www.accessdata.fda.gov/drugsatfda_docs/nda/2004/STN-125085_Avastin_Approv.pdf. Accessed 4 May 2023.

  4. Seo N, Polozova A, Zhang MX, Yates Z, Cao S, Li HM, Kuhns S, Maher G, McBride HJ, Liu J. Analytical and functional similarity of Amgen biosimilar ABP 215 to bevacizumab. MAbs. 2018;10(4):678–91. https://doi.org/10.1080/19420862.2018.1452580.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Bellinvia S, Edwards CJ. Explaining biosimilars and how reverse engineering plays a critical role in their development. Expert Opin Drug Discov. 2020;15(11):1–7. https://doi.org/10.1080/17460441.2020.1796627.

    Article  CAS  Google Scholar 

  6. Committee for Medicinal Products for Human Use (CHMP) of European Medicines Agency (EMA). Guideline on similar biological medicinal products (Rev.1) [Accessed 2014 Oct 23]. https://www.ema.europa.eu/en/similar-biological-medicinal-products-scientific-guideline.

  7. Committee for Medicinal Products for Human Use (CHMP) of European Medicines Agency (EMA). Guideline on similar biological medicinal products containing biotechnology-derived proteins as active substance: quality issues (revision 1) [Accessed 2014 May 22]. https://www.ema.europa.eu/en/similar-biological-medicinal-products-containing-biotechnology-derived-proteins-active-substance#current-effective-version-section.

  8. U.S. Food and Drug Administration (FDA). Guidance for industry: scientific considerations in demonstrating biosimilarity to a reference product. U.S. Department of Health and Human Services [accessed 2015 Apr]. https://www.fda.gov/regulatory-information/search-fda-guidance-documents/scientific-considerations-demonstrating-biosimilarity-reference-product.

  9. U.S. Food and Drug Administration (FDA). Guidance for industry: development of therapeutic protein biosimilars: comparative analytical assessment and other quality-related considerations. U.S. Department of Health and Human Services [accessed 2019 May]. https://www.fda.gov/search?s=development+of+therapeutic+protein+biosimilars%3A+comparative+analytical+assessment+and+other+quality-related+considerations.

  10. Alsamil AM, Giezen TJ, Egberts TC, Leufkens HG, Vulto AG, Plas MR, Gardarsdottir H. Reporting of quality attributes in scientific publications presenting biosimilarity assessments of (intended) biosimilars: a systematic literature review. Eur J Pharm Sci. 2020;154:1–10. https://doi.org/10.1016/j.ejps.2020.105501.

    Article  CAS  Google Scholar 

  11. Xu YP, Xie LQ, Zhang EH, Gao WY, Wang LL, Cao Y, Xie MH, Jiang WD, Liu S. Physicochemical and functional assessments demonstrating analytical similarity between rituximab biosimilar HLX01 and the MabThera(R). MAbs. 2019;11(3):606–20. https://doi.org/10.1080/19420862.2019.1578147.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Seo N, Huang Z, Kuhns S, Sweet H, Cao S, Wikstrom M, Liu J. Analytical and functional similarity of biosimilar ABP 798 with rituximab reference product. Biologicals. 2020;68:79–91. https://doi.org/10.1016/j.biologicals.2020.08.002.

    Article  CAS  PubMed  Google Scholar 

  13. Cerutti ML, Pesce A, Bes C, Seigelchifer M. Physicochemical and biological characterization of RTXM83, a new rituximab biosimilar. BioDrugs. 2019;33(3):307–19. https://doi.org/10.1007/s40259-019-00349-2.

    Article  CAS  PubMed  Google Scholar 

  14. Xie LQ, Zhang EH, Xu YP, Gao WY, Wang LL, Xie MH, Qin PL, Lu LH, Li SP, Shen PC, Jiang WD, Liu S. Demonstrating analytical similarity of trastuzumab biosimilar HLX02 to Herceptin® with a panel of sensitive and orthogonal methods including a novel FcγRIIIa affinity chromatography technology. BioDrugs. 2020;34(3):363–79. https://doi.org/10.1007/s40259-020-00407-0.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Lee J, Kang HA, Bae JS, Kim KD, Lee KH, Lim KJ, Choo MJ, Chang SJ. Evaluation of analytical similarity between trastuzumab biosimilar CT-P6 and reference product using statistical analyses. MAbs. 2018;10(4):547–71. https://doi.org/10.1080/19420862.2018.1440170.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Liu J, Eris T, Li C, Cao S, Kuhns S. Assessing analytical similarity of proposed Amgen biosimilar ABP 501 to adalimumab. BioDrugs. 2016;30(4):321–38. https://doi.org/10.1007/s40259-016-0184-3.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Zhang EH, Xie LQ, Qin PL, Lu LH, Xu YP, Gao WY, Wang LL, Xie MH, Jiang WD, Liu S. Quality by design-based assessment for analytical similarity of adalimumab biosimilar HLX03 to Humira®. AAPS J. 2020;22(3):1–14. https://doi.org/10.1208/s12248-020-00454-z.

    Article  CAS  Google Scholar 

  18. Yu CF, Zhang F, Xu GL, Wu G, Wang WB, Liu CY, Fu ZH, Li M, Guo S, Yu XJ, Wang L. Analytical similarity of a proposed biosimilar BVZ-BC to bevacizumab. Anal Chem. 2020;92(4):3161–70. https://doi.org/10.1021/acs.analchem.9b04871.

    Article  CAS  PubMed  Google Scholar 

  19. McClellan JE, Conlon HD, Bolt MW, Kalfayan V, Palparthy R, Rehman MI, Kirchhoff CF. The ‘totality-of-the-evidence’ approach in the development of PF-06438179/GP1111, an infliximab biosimilar, and in support of its use in all indications of the reference product. Therap Adv Gastroenterol. 2019;12:1–13. https://doi.org/10.1177/1756284819852535.

    Article  CAS  Google Scholar 

  20. Ratih R, Asmari M, Abdel-Megied AM, Elbarbry F, Deeb SE. Biosimilars: review of regulatory, manufacturing, analytical aspects and beyond. Microchem J. 2021;165:1–10. https://doi.org/10.1016/j.microc.2021.106143.

    Article  CAS  Google Scholar 

  21. Paek K, Kim G, Ahn SY, Lim JH, Jung D, Kim S, Lee JH. Assessment of the molecular mechanism of action of SB3, a trastuzumab biosimilar. BioDrugs. 2019;33(6):661–71. https://doi.org/10.1007/s40259-019-00381-2.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Fei MD, Zhang Q, Zhang L, Zhu XQ, Du CF, Zhang ZL. Development and validation of aggregates analysis method in analytical similarity assessment of HLX04 vs Avastin. J Pharm Biomed Anal. 2022;223:115121. https://doi.org/10.1016/j.jpba.2022.115121.

    Article  CAS  PubMed  Google Scholar 

  23. Ruppen I, Beydon ME, Solís C, Sacristán D, Vandenheede I, Ortiz A, Sandra K, Adhikary L. Similarity demonstrated between isolated charge variants of MB02, a biosimilar of bevacizumab, and Avastin following extended physicochemical and functional characterization. Biologicals. 2022;77:1–15. https://doi.org/10.1016/j.biologicals.2021.08.002.

    Article  CAS  PubMed  Google Scholar 

  24. Kang J, Kim SY, Vallejo D, Hageman TS, White DR, Benet A, Coghlan J, Sen KI, Ford M, Saveliev S, Tolbert TJ, Weis DD, Schwendeman SP, Ruotolo BT, Schwendeman A. Multifaceted assessment of rituximab biosimilarity: the impact of glycan microheterogeneity on Fc function. Eur J Pharm Biopharm. 2020;146:111–24. https://doi.org/10.1016/j.ejpb.2019.12.003.

    Article  CAS  PubMed  Google Scholar 

  25. Kaur T, Shukla BN, Yadav VK, Kulkarni MJ, Rao A. Comparison of glycoprofiles of rituximab versions licensed for sale in India and an analytical approach for quality assessment. J Proteomics. 2021;244:1–10. https://doi.org/10.1016/j.jprot.2021.104267.

    Article  CAS  Google Scholar 

  26. Lee JJ, Yang J, Lee C, Moon Y, Ahn S, Yang J. Demonstration of functional similarity of a biosimilar adalimumab SB5 to Humira®. Biologicals. 2019;58:7–15. https://doi.org/10.1016/j.biologicals.2018.12.002.

    Article  CAS  PubMed  Google Scholar 

  27. WHO Expert Committee on Biological Standardization. Guidelines on evaluation of biosimilars [accessed 2022 Apr 22]. https://www.who.int/publications/m/item/guidelines-on-evaluation-of-biosimilars--trs-1043--annex-3.

  28. Chow SC, Song F, Bai H. Analytical similarity assessment in biosimilar studies. AAPS J. 2016;18(3):670–7. https://doi.org/10.1208/s12248-016-9882-5.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Vandekerckhove K, Seidl A, Gutka H, Kumar M, Gratzl G, Keire D, Coffey T, Kuehne H. Rational selection, criticality assessment, and tiering of quality attributes and test methods for analytical similarity evaluation of biosimilars. AAPS J. 2018;20(68):1–9. https://doi.org/10.1208/s12248-018-0230-9.

    Article  CAS  Google Scholar 

  30. Committee for Medicinal Products for Human Use (CHMP) of European Medicines Agency (EMA). Reflection paper on statistical methodology for the comparative assessment of quality attributes in drug development. [Accessed 2021 Jul 26]. https://www.ema.europa.eu/en/statistical-methodology-comparative-assessment-quality-attributes-drug-development-scientific#current-version-section.

  31. Antes B, Amon S, Rizzi A, Wiederkum S, Kainer M, Szolar O, Fido M, Kircheis R, Nechansky A. Analysis of lysine clipping of a humanized Lewis-Y specific IgG antibody and its relation to Fc-mediated effector function. J Chromatogr B. 2007;852:250–6. https://doi.org/10.1016/j.jchromb.2007.01.024.

    Article  CAS  Google Scholar 

  32. Liu HC, Ponniah G, Zhang HM, Nowak C, Neill A, Gonzalez-Lopez N, Patel R, Cheng GL, Kita AZ, Andrien B. In vitro and in vivo modifications of recombinant and human IgG antibodies. MAbs. 2014;6(5):1145–54. https://doi.org/10.4161/mabs.29883.

    Article  PubMed  PubMed Central  Google Scholar 

  33. Hintersteiner B, Lingg N, Zhang PQ, Woen S, Hoi KM, Stranner S, Wiederkum S, Mutschlechner O, Schuster M, Loibner H, Jungbauer A. Charge heterogeneity: basic antibody charge variants with increased binding to Fc receptors. MAbs. 2016;8(8):1548–60. https://doi.org/10.1080/19420862.2016.1225642.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Qin SK, Li J, Bai YX, Shu YQ, Li W, Yin XL, Cheng Y, Sun GP, Deng YH, Zhong HJ, Li YF, Qian XP, Zhang LM, Zhang JD, Chen KH, Kang WY. Efficacy, safety, and immunogenicity of HLX04 versus reference bevacizumab in combination with XELOX or mFOLFOX6 as first-line treatment for metastatic colorectal cancer: results of a randomized, double-blind phase III study. BioDrugs. 2021;35(4):445–58. https://doi.org/10.1007/s40259-021-00484-9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Zhu XX, Qian HJ, Sun JX, Wu M, Yu C, Ding YH, Zhang XD, Chai K, Li XJ. A phase 1 randomized study compare the pharmacokinetics, safety and immunogenicity of HLX04 to reference bevacizumab sourced from the United States, the European Union, and China in healthy Chinese male volunteers. Cancer Chemother Pharmacol. 2021;88(3):465–74. https://doi.org/10.1007/s00280-021-04297-z.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Stracke J, Emrich T, Rueger P, Schlothauer T, Kling L, Knaupp A, Hertenberger H, Wolfert A, Spick C, Lau W, Drabner G, Reiff U, Koll H, Papadimitriou A. A novel approach to investigate the effect of methionine oxidation on pharmacokinetic properties of therapeutic antibodies. MAbs. 2014;6(5):1229–42. https://doi.org/10.4161/mabs.29601.

    Article  PubMed  PubMed Central  Google Scholar 

  37. Goetze AM, Liu YD, Zhang ZQ, Shah B, Lee E, Bondarenko PV, Flynn GC. High-mannose glycans on the Fc region of therapeutic IgG antibodies increase serum clearance in humans. Glycobiology. 2011;21(7):949–59. https://doi.org/10.1093/glycob/cwr027.

    Article  CAS  PubMed  Google Scholar 

  38. Thomann M, Reckermann K, Reusch D, Prasser J, Tejada ML. Fc-galactosylation modulates antibody-dependent cellular cytotoxicity of therapeutic antibodies. Mol Immunol. 2016;73:69–75. https://doi.org/10.1016/j.molimm.2016.03.002.

    Article  CAS  PubMed  Google Scholar 

  39. Hodoniczky J, Zheng YZ, James DC. Control of recombinant monoclonal antibody effector functions by Fc N-glycan remodeling in vitro. Biotechnol Prog. 2005;21:1644–52. https://doi.org/10.1021/bp050228w.

    Article  CAS  PubMed  Google Scholar 

  40. Idusogie EE, Wong PY, Presta LG, Gazzano-Santoro H, Totpal K, Ultsch M, Mulkerrinet MG. Engineered antibodies with increased activity to recruit complement. J Immunol. 2001;166(4):2571–5. https://doi.org/10.4049/jimmunol.166.4.2571.

    Article  CAS  PubMed  Google Scholar 

  41. Reusch D, Tejada ML. Fc glycans of therapeutic antibodies as critical quality attributes. Glycobiology. 2015;25(12):1325–34. https://doi.org/10.1093/glycob/cwv065.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Shields RL, Lai J, Keck R, O’Connell LY, Hong K, Meng YG, Weikert SHA, Presta LG. Lack of fucose on human IgG1 N-linked oligosaccharide improves binding to human Fcgamma RIII and antibody-dependent cellular toxicity. J Biol Chem. 2002;277(30):26733–40. https://doi.org/10.1074/jbc.M202069200.

    Article  CAS  PubMed  Google Scholar 

  43. Abès R, Teillaud JL. Impact of glycosylation on effector functions of therapeutic IgG. Pharmaceuticals. 2010;3:146–57. https://doi.org/10.3390/ph3010146.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Wang YN, Fei D, Vanderlaan M, Song A. Biological activity of bevacizumab, a humanized anti-VEGF antibody in vitro. Angiogenesis. 2004;7:335–45. https://doi.org/10.1007/s10456-004-8272-2.

    Article  CAS  PubMed  Google Scholar 

  45. Ding Y, Yu T, Sun J, Wu M, Chen Q, Qian H, Xie L, Zhang X, Liu E, Jiang W, Liu S, Luk A. A China-manufactured bevacizumab biosimilar, HLX04, matches bevacizumab sourced from China, USA and the European Union. Ann Oncol. 2018;29:ix33-34.

    Article  Google Scholar 

Download references

Acknowledgements

We gratefully thank the scientists of the Department of Analytical Science and Development, Shanghai Henlius Biologics Co., Ltd., for their assistance with experimental execution and figure preparation, as well as the Departments of Manufacturing, Quality Control, and Quality Assurance, Shanghai Henlius Biopharmaceutical Co., Ltd., for HLX04 manufacture and release.

Funding

This work was funded by Shanghai Henlius Biologics Co., Ltd.

Author information

Authors and Affiliations

  1. Department of Analytical Science and Development, Shanghai Henlius Biologics Co., Ltd, 5155 Guangfulin Road, Shanghai, 201600, China

    Lei Zhang, Lu Yu, Yanpeng Xu, Peilan Qin, Pengcheng Shen, Kemeng Liu, Mengdan Fei, Hongya Wang, Yanjing Cao, Lihong Lu, Wenyuan Gao & Zhongli Zhang

Authors
  1. Lei Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Lu Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Yanpeng Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Peilan Qin
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Pengcheng Shen
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Kemeng Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Mengdan Fei
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Hongya Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Yanjing Cao
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Lihong Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Wenyuan Gao
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Zhongli Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Conceptualization, Zhang L and Zhang ZL; acquisition and analysis, Lu Yu, Yanpeng Xu, Peilan Qin, Pengcheng Shen, Kemeng Liu, Mengdan Fei, Hongya Wang, Yanjing Cao, Lihong Lu, and Wenyuan Gao; supervision, Zhang ZL; writing, original draft preparation, Zhang L and Lu Yu; writing, review and editing, Zhang ZL. All authors have read and agreed to the published version of the manuscript. Zhang ZL is the corresponding author for the manuscript.

Corresponding author

Correspondence to Zhongli Zhang.

Ethics declarations

Conflict of interest

The authors, Lei Zhang, Lu Yu, Yanpeng Xu, Peilan Qin, Pengcheng Shen, Kemeng Liu, Mengdan Fei, Hongya Wang, Yanjing Cao, Lihong Lu, Wenyuan Gao, and Zhongli Zhang, are employees of Shanghai Henlius Biologics Co., Ltd., and declare that they have no other conflicts of interest that might be relevant to the contents of this article.

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 872 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

Zhang, L., Yu, L., Xu, Y. et al. Demonstrating analytical similarity of a biosimilar HLX04 to bevacizumab with a panel of state-of-the-art methods and tiering of quality attributes. Anal Bioanal Chem 415, 3341–3362 (2023). https://doi.org/10.1007/s00216-023-04716-5

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00216-023-04716-5

Keywords

Search

Navigation