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The effects of Thymidine phsphorylase on tumor angiogenesis and on the sensitivity to anti-tumor drugs

胸苷磷酸化酶对肿瘤血管生成及抗肿瘤药物敏感性的影响

基本信息

批准号：
06670172
负责人：
HARAGUCHI Misako
金额：
$ 1.28万
依托单位：
Kagoshima University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06670172/
关键词：
Tymidine phosphorylase Angiogenesis The sensitivity to anti-tumor drugs Chick embryo chorioallantoic membrane assay Chemtaxis 鶏卵漿尿膜法

项目摘要

Human thymidine phsphorylase (dThdPase) is thought to be indentical to an angiogenic factor, Platelet-derived endothelial cell growth factor (PD-ECGF) .However, the whole amino acid sequence of dThdPase is still unknown. N-termina amino acid sequencing of dThdPase isolated human placenta gave the sequence Ac-AALMTPGTGAPPAPG.Comparison with the sequence predicted from the PD-ECGFcDNA reveals that residues 2-16 of dThdPase are indentical to that of PD-ECGF.The primary translational product of dThdPase would be processed one amino acid from the translation-initiating methionine residue and Ala-2 acetylated.dThdPase has angiogenic activity. To investigate whether the dThdpase activity of PD-ECGF/dThdpase is indispensable to its angiogenic activity, three PD-ECGF/dThdpase mutant which have no dThdpase activity were made by site-derected mutagenesis. The lysates of COS cells transfected with the wild-type PD-ECGF/dThdpase cDNA had angiogenic activity, but those transfected with the mutant PD-ECGF/dThdpase did not. An inhibitor of dThdpase, 6-amino-5-chlorouracil, inhibited the angiogenic activity of purified dThdPase. These findings indicate that dThdpase activity is indispensable to angiogenic activity of PD-ECGF/dThdpase.Human KB epidermal carcinoma cells transfected with PD-ECGF cDNA expressed a 55-kDa protein that was detected with anti-dThdPase antibody and the cell lysate had dThdPase activity. The sensitivity of the transfected cells to 5'-deoxy-5-fluorouridine was higher than that of untransfected KB cells. These results demonstrate that dThdPase is involved in the activation of these anticancer agents.

人胸腺嘧啶核苷磷酸酶(DThdPase)与血管生成因子--血小板衍化内皮细胞生长因子(PD-ECGF)密切相关，但其氨基酸全序列尚不清楚。对分离的人胎盘dThdPase进行N-末端氨基酸测序，得到了Ac-AALMTPGTGAPPAPG序列。与PD-ECGFcDNA预测的序列比较，dThdPase的2-16个残基与PD-ECGF相同。dThdPase的初级翻译产物将被翻译起始的蛋氨酸残基和Ala-2乙酰化处理成一个氨基酸。dThdPase具有血管生成活性。为了研究pD-ECGF/dThdpase的dThdpase活性是否对其血管生成活性是必不可少的，通过定点突变获得了三个不具有dThdpase活性的pd-ECGF/dThdpase突变体。野生型PD-ECGF/dThdpase基因转染的COS细胞裂解产物具有血管生成活性，而突变型PD-ECGF/dThdpase基因转染的COS细胞裂解产物不具有血管生成活性。DThdpase的抑制剂6-氨基-5-氯尿嘧啶可抑制纯化的dThdPase的血管生成活性。这些结果表明，dThdpase活性对PD-ECGF/dThdPase的血管生成活性是不可或缺的。转导PD-ECGF基因的人KB表皮癌细胞表达一种55 kDa的蛋白，抗dThdPase抗体检测到该蛋白，细胞裂解物具有dThdPase活性。转染组KB细胞对5‘-脱氧-5-氟尿嘧啶的敏感性高于未转染组。这些结果表明dThdPase参与了这些抗癌药物的激活。