Comparison of methods for assessing the immunogenicity of biological drugs based on monoclonal antibodies using the example of pembrolizumab

Valeriia S. Chernyshenko; Чернышенко В. С.; Natalia M. Faustova; Фаустова Н. М.; Vera M. Kosman; Косман В. М.; Marina V. Karlina; Карлина М. В.; Anna Yu. Romanenko; Романенко А. Ю.; Marina N. Makarova; Макарова М. Н.; Valery G. Makarov; Макаров В. Г.; Anastasia A. Dmitrieva; Дмитриева А. А.; Elena V. Shipaeva; Шипаева Е. В.; Mikhail G. Chernobrovkin; Чернобровкин М. Г.; Victoria A. Postnikova; Постникова В. А.; Olga V. Filon; Филон О. В.; Mikhail Yu. Samsonov; Самсонов М. Ю.; Vasiliy G. Ignatiev; Игнатьев В. Г.

doi:10.17650/1726-9784-2026-25-1-71-83

Comparison of methods for assessing the immunogenicity of biological drugs based on monoclonal antibodies using the example of pembrolizumab

Authors: Chernyshenko V.S.¹, Faustova N.M.¹, Kosman V.M.¹, Karlina M.V.¹, Romanenko A.Y.¹, Makarova M.N.¹, Makarov V.G.¹, Dmitrieva A.A.², Shipaeva E.V.², Chernobrovkin M.G.³, Postnikova V.A.³, Filon O.V.², Samsonov M.Y.², Ignatiev V.G.²
Affiliations:
1. Research-and-manufacturing company “HOME OF PHARMACY”
2. Joint-Stock Company “R-Pharm”
3. Limited Liability Company “Research Lab”
Issue: Vol 25, No 1 (2026)
Pages: 71-83
Section: ORIGINAL REPORTS
Published: 30.04.2026
URL: https://bioterapevt.abvpress.ru/jour/article/view/1557
DOI: https://doi.org/10.17650/1726-9784-2026-25-1-71-83
ID: 1557

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Abstract

Background. To ensure the safety and effectiveness of biological drugs, it is critically important to evaluate their immunogenicity. The most commonly used enzyme-linked immunosorbent assay (ELISA) has a number of limitations, so it is important to understand the possibilities of alternative approaches, for example, the biolayer interferometry (BSI) method.

Aim. Development, validation and comparison of two methods for screening assay in the context of assessing the immunogenicity of biological drugs based on monoclonal antibodies using the example of pembrolizumab with multiple intravenous administration to Macaca fascicularis.

Materials and methods. Immunogenicity was evaluated as part of a comparative study of toxicity and local tolerance of two drugs with International Nonproprietary Name pembrolizumab following repeated administration. Macaca fascicularis (3 groups of 3 males and 3 females in each) on days 1, 8, 15 and 22 of the experiment received intravenously a control substance (buffer solution), test drug (RPH-075, JSC R-Pharm, Russia) and reference drug (Keytruda®, MSD International GmbH, Ireland) at a dose 30 mg/kg. Serum samples, were collected pre-dose and on study days 15, 22, 29, and were tested for assessment of binding antibodies to pembrolizumab using two methods, bridging ELISA and BSI.

Results. During the development and testing of methods for binding antibodies (BAb) to pembrolizumab quantification in monkey’s blood serum of by the bridge ELISA and BSI assays, it was revealed that the bridge ELISA has greater sensitivity (12.3 and 419.7 ng/mL, respectively). Determination of BAb level to pembrolizumab by ELISA revealed immunogenicity in 4 out of 12 animals; when analyzing samples by BSI, an increasing of BAb concentration was detected in only one animal.

Conclusion. Bridging ELISA is more sensitive than BSI making it the preferred method for assessing of drugs with unknown or suspected low immunogenicity.

Keywords

immunogenicity, pembrolizumab, bridging enzyme-linked immunosorbent assay, biolayer interferometry, anti-drug antibody, binding antibodies, Macaca fascicularis

References

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Comparison of methods for assessing the immunogenicity of biological drugs based on monoclonal antibodies using the example of pembrolizumab

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Abstract

Keywords

About the authors

References

Supplementary files