Cell Killing effect heparin-binding EGF-like growth factor

细胞杀伤作用肝素结合EGF样生长因子

• 批准号: 07670553
• 负责人: ONO Minoru
• 金额: $ 1.54万
• 依托单位: ASAHIKAWA MEDICAL COLLEGE
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07670553/
• 关键词: HBEGF; Heparin; Tunror; Hepatoma; tokim; growth; chemotherapy; Pseudomonas; heparan sulfate proteoglycan

Heparin-binding EGF-like growth factor (HB-EGF) is a potent mitogen for smooth muscle cell, fibroblast, and it also stimulates hepatocyte proliferation. We generated several chimeric toxins by fusing the cDNA sequence of HB-EGF and the mutant of Pseudomonas exotoxin, PE^<4E>KDEL (PE) that lacks the binding abiliy to specific receptor. HB-EGF-PE was generated by fusing the DNA fragment encoding the full length mature HB-EGF polypeptide to the N-terminus of PE^<4E>KDEL,while HB-PE was generated by fusing the 45 N-terminal heparin-binding sequence to PE^<4E>KDEL.HB-EGF-PE was capable of binding both to EGF receptor and heparan sulfate proteoglycans (HSPGs), whereas HB-PE was capable of binding only to HSPGs on the target cells. Human hepatoma cells, SK-Hep1, Hep-G2 and PLC/PRF/5 were killed in a very low concentration of HB-EGF-PE with the ID50 of 0.1 to 0.5 ng/ml. HB-PE could also killed SK-Hep1 with the ID50 of 50 ng/ml, whereas it was resistant to PE.Both exogenous EGF and heparin inhibited the cytotoxicity of HB-EGF-PE.These results indicated the existence of two alternative pathways for the internalization of the chimeric toxins defined by two different targets, EGFR and HSPGs.

肝素结合EGF样生长因子（HB-EGF）是平滑肌细胞，成纤维细胞的有效有丝分裂原，并且还刺激了肝细胞增殖。我们通过融合Hb-EGF的cDNA序列和假单胞菌毒素的突变体，PE^<4e> kdel（PE）产生了几种嵌合毒素，该突变体缺乏与特定受体的结合。 HB-EGF-PE是通过融合DNA片段将编码全长的成熟HB-EGF多肽与PE^<4e> kDel的N末端进行融合而产生的，而HB-PE则是通过将45 N末端肝素结合序列融合到PE^<4e> kdel.hb-egf-par的45 n末端肝素结合序列来产生的，而ED均可受体的能力。蛋白聚糖（HSPG），而HB-PE仅与靶细胞上的HSPG结合。人肝癌细胞，SK-HEP1，HEP-G2和PLC/PRF/5在非常低的HB-EGF-PE中杀死，ID50为0.1至0.5 ng/ml。 HB-PE还可以用50 ng/ml的ID50杀死SK-HEP1，而对PE的外源性EGF和肝素的耐药性抑制了HB-EGF-PE的细胞毒性。这些结果表明存在两个不同目标的内在化的两种交替途径，由两个不同的目标定义了两个不同的目标。

项目成果

Mizukami Y,Ohhira M,Matsumoto A,Murazumi Y,Murazumi K,Ohta H,Ohhira Ma, Ono M,Miyake T,Maekawa I,Kohgo Y.: "Primary biliary cirrhosis associated with idiopatic thrombocytopenic purpura." J Gastroenterol. 31. 284-288 (1996)

Mizukami Y、Ohhira M、Matsumoto A、Murazumi Y、Murazumi K、Ohta H、Ohhira Ma、Ono M、Miyake T、Maekawa I、Kohgo Y.：“与特发性血小板减少性紫癜相关的原发性胆汁性肝硬化。”

Ono, M: "Cell killing effect of heparin-bindingEGF-like growth factor Pseudomonas exotoxin on human hepatoma cells." International Hepatology Communications. 6. 79-84 (1996)

Ono, M：“肝素结合 EGF 样生长因子假单胞菌外毒素对人肝癌细胞的细胞杀伤作用。”

Ohhira, M: "Changes in free radical-meyabolizing enzymes and lipid peroxides in the liver of Long-Evans with cinnamon-like coat color rats." J Gastroenterology. 30. 619-623 (1995)

Ohhira, M：“具有肉桂样皮毛颜色的朗-埃文斯大鼠肝脏中自由基代谢酶和脂质过氧化物的变化。”

Mizukami, Y: "Primary biliary cirrhosis asociated with idiopatic thrombocytopenic purpura." J Gastroenterol. 31. 284-288 (1996)

Mizukami，Y：“与特发性血小板减少性紫癜相关的原发性胆汁性肝硬化。”

Ohhira M.Matsumoto A,Ohhira Ma, Murazumi K,Ohta H,Ono M,Kohgo Y.: "Paraumbilical vein Aneuysm Case reports." Angiology. 47. 517-521 (1996)

Ohhira M.Matsumoto A、Ohhira Ma、Murazumi K、Ohta H、Ono M、Kohgo Y.：“脐旁静脉动脉瘤病例报告”。