Abstract
Abstract Snake venoms contain various biologically active proteins including C-type lectins and -like proteins (CTLs), phospholipases, proteinases and metalloproteinases, that could target diverse cellular processes. Among these active components, CTLs have demonstrated antithrombotic effects through their platelet agglutination and coagulation activities. In our study, we have detected anti-angiogenesis activity from the venom of Deinagkistrodon acutus snake. The partial purified fraction of snake venom with anti-angiogenesis activity was isolated through a stepwise separation in anion-exchange and cation-exchange chromatography. At the same time, the monoclonal antibody was generated and used for further protein purification by affinity chromatography. This highly purified protein was named as ZK002. By MALDI-TOF mass spectrometry determination, the mass of ZK002 is about 30KD. By LC-MS/MS analysis, protein sequence of highly purified ZK002 was determined and compared with the Deinagkistrodon acutus snake protein database. ZK002 is a heterodimer of CTLs consisting of alpha and beta peptide chains. Using In vitro chicken embryo chorioallantoic membrane assay and human umbilical vein endothelial cell (HUVEC) tube formation assay, it showed that ZK002 exhibited an anti-angiogenesis effect. Moreover, ZK002 also demonstrated an anti-cancer effect as shown in the ApcMin/+ mouse model for colorectal cancer. Administration of ZK002 significantly suppressed the total number and the size of tumors and showed no observable adverse effect. To further elucidate the anti-angiogenesis functions of ZK002, we deduced the coding DNA sequences from the peptide sequences of the alpha- and beta-chains, then subcloned individual gene sequence into yeast and mammalian expression vectors. Results showed that the recombinant vectors express secreted peptides with the expected molecular weight. Transient transfection of the alpha- and beta-recombinant vectors individually or in combination inhibit tube formation in HUVEC cells, suggesting that both alpha- and beta-chains possess anti-angiogenesis activity. The success of cloning and expression of recombinant ZK002 would greatly facilitate the future development of ZK002 as anti-angiogenesis and anti-cancer drugs. (This work is supported by the Innovation and Technology Fund (UIM208) of Hong Kong Government) Citation Format: Shan Yu, Xiangrong Dai, William C.S. Tai, Roy C.Y. Choi, Chaofan Liang, Benjamin X.y. Li, Karl W.K. Tsim, Wendy W.L. Hsiao. Anti-angiogenesis and anti-cancer activities of ZK002, a C-type lectin-like protein isolated from snake venom. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5077. doi:10.1158/1538-7445.AM2013-5077
Published Version
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