HCGB FROM CERVICAL CANCER: PEPTIDE HETEROGENITY

宫颈癌中的 HCGB：肽异质性

• 批准号: 3166107
• 负责人: ROBERT O HUSSA
• 金额: $ 9.96万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 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/3166107
• 关键词: affinity chromatography; blood chemistry; cervix neoplasms; gel electrophoresis; gel filtration chromatography; high performance liquid chromatography; human tissue; radiotracer; scintillation spectrometry; tissue /cell culture; urinalysis

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 含有两种成分，异位hCG-β-I和异位hCG-β-II。 异位hCG-β-II缺乏C-末端肽（CTP），基于缺乏 CTP抗体的识别，缺乏对 嗜热菌蛋白酶（其通常切割CTP），以及缺乏与CTP的结合。 花生（Arachis hypogaea）凝集素的去唾液酸化衍生物（对 CTP上O-连接寡糖的Gal-GalNAc）。 免疫反应 在18名患者中的13名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 CaSki宫颈癌细胞的培养进行了优化。 的方法 已证实异位hCG-β-I与异位hCG-β-II分开， 涉及使用以下物质的差示亲和层析：（1）\抗-hCG-α （2）抗-[CTP导向的] hCG-β-琼脂糖;和 (3)抗-[核心导向] hCG-β琼脂糖凝胶。 洗脱异位hCG-β-II 从第三阶段用4 M胍\HCl，pH 3洗脱，并脱盐。 的 纯化的异位hCG-β-II不能与标准hCG-α联合收割机结合 在标准hCG-β与标准hCG-α结合的条件下。 获得的证据表明， 异位hCG-β DoT和CaSki培养液含有免疫反应性 hCG-β样成分明显大于标准hCG-β (from妊娠尿hCG），如凝胶过滤和钠 十二烷基硫酸盐凝胶电泳（后者孵育后， 具有放射性氨基酸的CaSki细胞）。 （丙）