Enrichment of phage display library against breast cancer cells for isolation of anti-HER2 camelid single domain antibodies

Breast cancer cells can be hidden from immune cells by shedding tumor markers such as HER2. The increased soluble HER2 (sHER2) concentrations are associated with the outcome of HER2-positive breast cancer and sensitivity to trastuzumab treatment. To this end, camelid single domain antibodies (VHH) with unique properties and binding ability are very striking targeting agents to detect sHER2. By panning on HER2 negative and positive cells, an immune camel library was enriched against HER2 antigen. Affinity and specificity of four selected VHHs to HER2, detection of sHER2 and binding of selected VHHs to HER2 on breast cancer cells were investigated by ELISA. After cell panning and ELISA, four VHHs that recognized HER2 antigen better than negative control were identified. High affinity HER2 specific VHHs (1012-1013 M-1) were able to detect sHER2 (2 µg/ml). When the mixture of VHH and sMUC1 was added to HER2 coated-well, the VHH bound to HER2. RR4 and RR6 VHHs showed the highest binding to SKBR3 (HER2 expressing cell) (2 ± 0.33 and 2.5 ± 0.15, respectively) compared to HER2 negative cell (0.38 ± 0.17 and 0.4 ± 0.12, respectively). The HER2 status of a primary tumor and responses to treatment can be evaluated by measuring sHER2 as a biomarker by HER2 specific VHHs.