Selection and Evaluation of Human Recombinant Antibodies against ErbB2 Antigen for Breast Cancer Immunotherapy

Breast cancer is the most common cause of cancer-related death in women worldwide. ErBb2/HER2 breast cancer accounts for 25% - 30% of all cases of breast cancer. Approved anti-ErbB2 monoclonal antibodies, trastuzumab and pertuzumab, are currently used for the treatment of ErbB-positive breast cancer, although their clinical use is limited due to their immunogenicity. Human recombinant single-chain antibodies, which are produced by antibody engineering technologies, are new and effective antibodies in cancer immunotherapy..
To select specific single-chain variable fragments (scFvs) against 2 immunodominant ErbB2 epitopes, including trastuzumab and pertuzumab binding sites, and to evaluate their reactivity and specificity against ErbB2 epitopes..
Escherichia coli bacteria, containing a phagemid with a scFv insert segment, were used to select specific high-affinity scFvs against 2 ErbB2 epitopes, using the panning process. Polymerase chain reaction (PCR) and DNA fingerprinting were performed on the obtained clones to select the positive ones and isolate the common patterns. The selected clones were evaluated via phage Elisa in terms of reactivity and specificity to epitopes..
Single-chain antibodies, scFvI and scFvII, which are ErbB2-specific with 40% and 45% frequencies, were selected against epitopes I and II, respectively. The results of phage ELISA demonstrated a significant difference in the optical density (OD) of scFvs in reaction with the related peptides and non-peptide wells. ODs of 0.65 and 0.71 were obtained for scFvI and scFvII reactions with the corresponding peptides, whereas the ODs of non-peptide wells were 0.1 and 0.13, respectively..
Targeted cancer therapy, which acts on a specific molecule in cancer cells, minimizes the side effects of immunotherapy. Due to the unique properties of scFvs, these antibodies have been used in targeted therapy of several cancers. In this study, 2 specific scFvs were selected against 2 ErbB2 epitopes, which contained trastuzumab and pertuzumab binding sites. The results of the panning process demonstrated the selection of 2 specific scFvs (with frequencies of 40% and 45%, respectively), which significantly reacted with the corresponding epitopes in phage ELISA assay. These small, high-affinity, human antibodies, which were selected against regions containing the binding sites of 2 food and drug administration (FDA)-approved monoclonal antibodies for breast cancer immunotherapy, have the potential to be considered for breast cancer targeted therapy. However, in vitro and in vivo tests should be performed to evaluate the antitumor effects of these scFvs..