Thesis (M.Sc.) - Cairo University - Faculty of Pharmacy - Department of Microbiology and Immunology

Immunogenicity is a major challenge in drug development and patient care. Clinicians and regulators are familiar with immunogenicity concerns of therapeutics, growth factors, and enzyme replacements. Unlike small molecule therapeutics, the biological therapeutic agents are likely to trigger undesirable immunogenic responses against themselves upon their administration. This imparts a problem that has to be considered upon judging their risk-benefit ratio.The development of monoclonal antibodies (mAbs) counts as one of the major medical steps forward, opening up endless possibilities for research, diagnosis, and treatment of a wide range of various diseases and disorders. Development of both humanized and fully human mAbs was expected to be non-immunogenic, that allow repeated administration without any anti-drug antibodies (ADA) responses. Unfortunately, these expectations were proven to be unrealistic. They fail to completely eliminate mAb immunogenicity and ADA formation; they have reduced the extreme immunogenicity associated with murine origin mAbs, but still, they have shown to induce antibodies that sometimes have an impact on clinical outcomes. Accurate ADA detection is necessary, to provide sufficient information for patient monitoring and clinical intervention. However, assays of ADAs that are developed against mAbs have more challenges as both the analytes and antigens (mAbs) are antibodies. In this study, we tested the immunogenicity developed in patients sera due to the use of trastuzumab and that developed in laboratory animals injected with this recombinant humanized IgG1 monoclonal antibody(trastuzumab). The trastuzumab immunogenicity was assessed in blood samples using an in vitro approach and in laboratory animalsusingin vivo approach. The in vitro approach depended on both detections of anti-trastuzumab antibody (Ab) levels in patients' serum samples withdrawn at different points during the trastuzumab treatment course; and measurement of neutralizing activity of these ADAs using MTT cytotoxicity assay against MCF-7 cell line. The detection of anti-trastuzumab Abs was carried out using the affinity capture elution (ACE) assay methodin both approaches. In vitro analysis of patients' sera for antibodies developed against trastuzumab revealed that this monoclonal antibody has low immunogenicity. Only 1% of samples showed high levels of anti-trastuzumab antibodies which might affect the treatment course. In vivo immunogenicity testing in mice showed also low immunogenicity of trastuzumab that could support the relevant clinical dataapplied in this study

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