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HCGB FROM CERVICAL CANCER: PEPTIDE HETEROGENITY

宫颈癌中的 HCGB：肽异质性

基本信息

批准号：
3166109
负责人：
ROLAND A PATTILLO
金额：
$ 11.64万
依托单位：
MEDICAL COLLEGE OF WISCONSIN
依托单位国家：
美国
项目类别：
财政年份：
1978
资助国家：
美国
起止时间：
1978-05-01 至 1987-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3166109
关键词：
affinity chromatography blood chemistry cervix neoplasms gel electrophoresis gel filtration chromatography gonadotropins high performance liquid chromatography human tissue radiotracer scintillation spectrometry tissue /cell culture tritium urinalysis

项目摘要

Ectopic hCG-beta secreted by DoT and CaSki human carcinoma cell lines contains two components, ectopic hCG-beta-I and ectopic hCG-beta-II. Ectopic hCG-beta-II lacks the C-terminal peptide (CTP), based on lack of recognition by antibodies against CTP, lack of susceptibility to thermolysin (which normally cleaves the CTP), and lack of binding of the desialylated derivative to Arachis hypogaea (AH) lectin (specific for Gal-GalNAc of the O-linked oligosaccharides on the CTP). Immunoreactive hCG-beta-like material has been detected in the serum or urine of 13 of 18 patients with cervical carcinoma. Conditions for large-scale roller bottle culture of CaSki cervical carcinoma cells were optimized. A method to separate ectopic hCG-beta-I from ectopic hCG-beta-II has been validated and involves differential affinity chromatography with: (1)\anti-hCG-alpha Sepharose; (2)\anti-[CTP-directed]hCG-beta-Sepharose; and (3)\anti-[core-directed]hCG-beta Sepharose. Ectopic hCG-beta-II is eluted from the third stage with 4 M guanidine\HC1, pH 3, and desalted. The purified ectopic hCG-beta-II failed to combine with standard hCG-alpha under conditions where standard hCG-beta combined with standard hCG-alpha. Evidence was obtained for the presence of additional carbohydrates on ectopic hCG-beta. DoT and CaSki culture fluids contained immunoreactive hCG-beta-like components of apparently larger size than standard hCG-beta (from pregnancy urine hCG) as demonstrated by gel filtration and sodium dodecyl sulfate-gel electrophoresis (the latter following incubation of CaSki cells with radioactive amino acids). (C)

DoT 和 CaSki 人癌细胞系分泌的异位 hCG-β 含有异位 hCG-β-I 和异位 hCG-β-II 两种成分。异位 hCG-β-II 缺乏 C 末端肽 (CTP)，基于缺乏被 CTP 抗体识别，缺乏敏感性嗜热菌蛋白酶（通常会裂解 CTP），并且缺乏与花生 (AH) 凝集素的去唾液酸化衍生物（特定于 CTP 上的 O-连接寡糖的 Gal-GalNAc）。免疫反应性 18 人中有 13 人的血清或尿液中检测到 hCG-β 样物质宫颈癌患者。大型滚瓶的条件对CaSki宫颈癌细胞的培养进行了优化。一种方法已验证将异位 hCG-β-I 与异位 hCG-β-II 分开涉及差示亲和层析： (1)\anti-hCG-alpha 琼脂糖凝胶； (2)\抗[CTP导向]hCG-β-Sepharose；和 (3)\抗[核心导向]hCG-β琼脂糖凝胶。异位 hCG-β-II 被洗脱从第三阶段开始，使用 4 M 胍\HCl，pH 3，并脱盐。这纯化的异位 hCG-β-II 未能与标准 hCG-α 结合在标准 hCG-β 与标准 hCG-α 组合的条件下。获得的证据表明存在额外的碳水化合物异位 hCG-β。 DoT 和 CaSki 培养液含有免疫反应性 hCG-β 样成分的尺寸明显大于标准 hCG-β （来自妊娠尿液 hCG）如凝胶过滤和钠所示十二烷基硫酸盐凝胶电泳（后者在孵育后 CaSki 细胞具有放射性氨基酸）。 (三)