Construction, screening and selection of specific nanobodies against apoptosis-inducing epitope of NTR-DR5 domain using phage display technique (Article Uncorrected Proof)

One of the most promising strategies in cancer therapy is to induce apoptotic pathway. For this purpose، several constructed agonists of Death Receptor 5 (DR5) are in clinical development. Extrinsic and intrinsic apoptosis pathways of various cancer cells are primarily induced through the activation of the proapoptotic DR5. The extracellular domain of DR5 is comprised of several functional domains، among them the cysteine-rich domains (CRDs) play a critical role in TRAIL-DR5-mediated apoptosis. It has recently been shown that the binding of agonistic monoclonal antibody to another N-terminal domain of DR5 could mediate its activation and apoptosis induction. Variable domains derived from heavy chain antibodies (hcAb) called VHHs or nanobodies are robust، efficient and smallest antigen binding fragments. These unique features of VHHs make them potential therapeutic and diagnostic candidates. In the present study، using phage display technology، a library containing VHH genes was generated of an immunized camel with hapten-peptide 1ITQQDLAPQQRA12 and used to isolate the binders of this peptide. Through screening the phage library، three binders with high binding ability to desired epitope in the NTR region were obtained. Considering to the key role of this epitope in apoptosis inducing، these selected binders could be potential candidates to trigger apoptosis in various cancer cells.