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Int. J. Mol. Sci. 2017, 18(10), 2064; doi:10.3390/ijms18102064

A Novel Fully Human Agonistic Single Chain Fragment Variable Antibody Targeting Death Receptor 5 with Potent Antitumor Activity In Vitro and In Vivo

State Key Laboratory of Natural Medicines, School of Life Science and Technology, China Pharmaceutical University, Nanjing 210009, China
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Received: 27 July 2017 / Revised: 3 September 2017 / Accepted: 17 September 2017 / Published: 27 September 2017
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Abstract

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. View Full-Text
Keywords: death receptor 5 (DR5); human antibody; single chain fragment variable (scFv); apoptosis; phage display death receptor 5 (DR5); human antibody; single chain fragment variable (scFv); apoptosis; phage display
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Lei, G.; Xu, M.; Xu, Z.; Gu, L.; Lu, C.; Bai, Z.; Wang, Y.; Zhang, Y.; Hu, H.; Jiang, Y.; Zhao, W.; Tan, S. A Novel Fully Human Agonistic Single Chain Fragment Variable Antibody Targeting Death Receptor 5 with Potent Antitumor Activity In Vitro and In Vivo. Int. J. Mol. Sci. 2017, 18, 2064.

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