Agonistic antibodies， which bind specifically to death receptor 5 (DR5)， can trigger apoptosis in tumor cells through the extrinsic pathway. In this present study， we describe the use of a phage display to isolate a novel fully human agonistic single chain fragment variable (scFv) antibody， which targets DR5. After five rounds of panning a large (1.2 × 10⁸ clones) phage display library on DR5， a total of over 4000 scFv clones were screened by the phage ELISA. After screening for agonism in a cell-viability assay in vitro， a novel DR5-specific scFv antibody TR2-3 was isolated， which inhibited COLO205 and MDA-MB-231 tumor cell growth without any cross-linking agents. The activity of TR2-3 in indu... More
Agonistic antibodies， which bind specifically to death receptor 5 (DR5)， can trigger apoptosis in tumor cells through the extrinsic pathway. In this present study， we describe the use of a phage display to isolate a novel fully human agonistic single chain fragment variable (scFv) antibody， which targets DR5. After five rounds of panning a large (1.2 × 10⁸ clones) phage display library on DR5， a total of over 4000 scFv clones were screened by the phage ELISA. After screening for agonism in a cell-viability assay in vitro， a novel DR5-specific scFv antibody TR2-3 was isolated， which inhibited COLO205 and MDA-MB-231 tumor cell growth without any cross-linking agents. The activity of TR2-3 in inducing apoptosis in cancer cells was evaluated by using an Annexin V-PE apoptosis detection kit in combination with flow cytometry and the Hoechst 33342 and propidium iodide double staining analysis. In addition， the activation of caspase-dependent apoptosis was evaluated by Western blot assays. The results indicated that TR2-3 induced robust apoptosis of the COLO205 and MDA-MB-231 cells in a dose-dependent and time-dependent manner， while it remarkably upregulated the cleavage of caspase-3 and caspase-8. Furthermore， TR2-3 suppressed the tumor growth significantly in the xenograft model. Taken together， these data suggest that TR2-3 exhibited potent antitumor activity both in vitro and in vivo. This work provides a novel human antibody， which might be a promising candidate for cancer therapy by targeting DR5.
