title: The anti-cancer potency and mechanism of a novel tumor-activated fused toxin, DLM.
abstract: Melittin, which acts as a membrane-disrupting lytic peptide, is not only cytotoxic to tumors, but also vital to normal cells. Melittin had low toxicity when coupled with target peptides. Despite significant research development with the fused toxin, a new fused toxin is needed which has a cleavable linker such that the fused toxin can release melittin after protease cleavage on the tumor cell surface. We describe a novel fused toxin, composed of disintegrin, uPA (urokinase-type plasminogen activator)-cleavable linker, and melittin. Disintegrin is a single strand peptide (73 aa) isolated from Gloydius Ussuriensis venom. The RGD (Arg-Gly-Asp) site of disintegrin dominates its interaction with integrins on the surface of the tumor cells. uPA is over-expressed and plays an important role in tumor cell invasiveness and metastatic progression. The DLM (disintegrin-linker-melittin) linker is uPA-cleavable, enabling DLM to release melittin. We compared binding activity of our synthesized disintegrin with native disintegrin and report that DLM had less binding activity than the native form. uPA-cleavage was evaluated in vitro and the uPA-cleavable linker released melittin. Treating tumors expressing uPA with DLM enhanced tumor cell killing as well as reduced toxicity to erythrocytes and other non-cancerous normal cells. The mechanism behind DLM tumor cell killing was tested using a DNA ladder assay, fluorescent microscopy, flow cytometry, and transmission electron microscopy. Data revealed tumor cell necrosis as the mechanism of cell death, and the fused DLM toxin with an uPA-cleavable linker enhanced tumor selectivity and killing ability.
authors: Department of Biomedicine, Institute for Regeneration Medicine, Jilin University, Changchun 130021, China. sundjjl@126.com.
authors: Dejun
authors: D
authors: Sun
authors: Department of Biomedicine, Institute for Regeneration Medicine, Jilin University, Changchun 130021, China. sunmn1987@hotmail.com.
authors: Miaonan
authors: M
authors: Sun
authors: Department of Biochemistry, School of Basic Medicine, Jilin Medical College, Jilin 130000, China. zhuwenhejl@163.com.
authors: Wenhe
authors: W
authors: Zhu
authors: Department of Biomedicine, Institute for Regeneration Medicine, Jilin University, Changchun 130021, China. wzdfjfn@126.com.
authors: Zhiding
authors: Z
authors: Wang
authors: Department of Biomedicine, Institute for Regeneration Medicine, Jilin University, Changchun 130021, China. liyfjlu1989@126.com.
authors: Yuefei
authors: Y
authors: Li
authors: School of Pharmaceutical Sciences, Jilin University, Changchun 130021, China. ma_jie@jlu.edu.cn.
authors: Jie
authors: J
authors: Ma
mesh_terms: D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology* / Q000633:toxicity; D001792:Blood Platelets / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D019483:Disintegrins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology* / Q000633:toxicity; D004912:Erythrocytes / Q000187:drug effects; D005434:Flow Cytometry; D057809:HEK293 Cells; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008555:Melitten / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology* / Q000633:toxicity; D008856:Microscopy, Fluorescence; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism; D015539:Platelet Activation / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology* / Q000633:toxicity; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D019483:Disintegrins; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D008555:Melitten; C047981:arginyl-glycyl-aspartic acid; D014568:Urokinase-Type Plasminogen Activator