Validation of Two-Tiered Enzyme-Linked Immunosorbent Assay for the Detection of Anti-HeberProt-P® Antibodies

Rosales Ileana,Izquierdo Mabel,Hernández Carlos,Delgado Magali,Carpio Emilio,Valdivia Onel,Pérez Enrique,Pérez-Bernal Maylin

doi:10.35840/2633-8912/2414

Abstract

Heberprot-P<sup>®</sup> is a therapeutic product approved in Cuba for the treatment of diabetic foot ulcer. In spite of its ample clinical use, its immunogenicity in patients has not been evaluated. Regulatory agencies have developed guidelines for the validation of immunoassays designed at the detection of anti-drug antibodies to biologicals. The aim on this work was to validate a method for the detection of binding antibodies in human serum to recombinant human epidermal growth factor (rhEGF), the active ingredient of Heberprot-P<sup>®</sup>.

Highlights

As a result of the revolution in genetic engineering and biotechnology, a new group of drugs has emerged: Biopharmaceuticals; known as therapeutic proteins or biologicals
As minimum required dilution (MRD) was chosen 1:100, since in 90% of studied samples the background signal reduction was less than 100%
To determine recovery in this enzyme-linked immunosorbent assay (ELISA), dilution curves of anti-EGF antibodies was achieved by serial dilutions of positive control, using two types of diluents: Sample matrices at MRD or the dilution buffer (DB)

Summary

Introduction

As a result of the revolution in genetic engineering and biotechnology, a new group of drugs has emerged: Biopharmaceuticals; known as therapeutic proteins or biologicals. The clinical effects of these immune responses in patients range from no measurable (clinically benign) to extremely harmful [1] Even when both arms (humoral and cellular) of immune response can be activated by these drugs, the presence of anti-drug antibodies (ADA) has been of special concern. Taking this potential risk into consideration, regulatory agencies request that immunogenicity must be assessed as part of the approval process of therapeutic proteins and this information should be included in the in the prescribing information as a subsection of the “adverse reaction” section entitled Immunogenicity [1]. Regarding detection of ADA, regulatory agencies have developed statements and guidelines advising the proper methods and type of assays for the evaluation of unwanted immunogenicity, highlighting measurement and characterization of antibodies [2,3]

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Conclusion