O-GLYCOSYLATION OF HCG AND CANCER

这项拟议的研究有三个目标，每个目标都有自己的目标 方法学，但都有一个共同的临床目标-识别和 开发碳水化合物特异性肿瘤标记物，重点是hCG和 滋养细胞肿瘤的鉴别诊断。 1.O-连接糖单位在妊娠和肿瘤来源中的分布 HCG。在正常妊娠的尿液中，12%-14%的人 O-连接的糖单元(总共4个)是六糖结构， NeuAc-Gal-GlcNAc-(NeuAc-Gal)-GalNAc-。相比之下，试点研究表明 绒毛膜癌患者尿液中hCG的结构 受试者(2名受检者)和培养上清液中的绒毛癌细胞 大约50%的这种六糖结构。唯一发表的发现是 这种结构一直存在于癌症蛋白质上。以确认hCG含量丰富 在六糖中是绒毛膜癌的标志物，O-连接的结构 另外12-15名受试者尿液中hCG分子的糖分将 检查(方法：人绒毛膜促性腺激素免疫沉淀、β-消除/标记 与低聚糖的层析比较 标准)。为了确定六糖的丰度是否相关 随着滋养细胞疾病的恶性，尿hCG分子的恶性程度 来自良性侵袭性/部分性/完全性葡萄胎的受试者也会 接受检查。非滋养细胞肿瘤人绒毛膜促性腺激素上的O-连接糖单位， 并对其他尿液和组织中的糖蛋白进行检测。 2.滋养细胞疾病特异性检测方法的研究进展虽然 滋养细胞疾病在美国很少见(1500例妊娠中就有1例)，10倍 在几个非洲和亚洲国家发现的发病率更高。至 产生一种疾病特异性分析，即β亚单位C端肽 含六糖的绒毛膜癌hCG将被分离并 作为免疫原用于制备绵羊多克隆抗血清， 单抗接近，并作为凝集素-抗体系统的结构域。 3.糖基转移酶在正常、肿瘤和诱导性肿瘤中的活性 转化的细胞。调查通过以下途径或机制进行研究 癌症伴发蛋白糖基化的变化--糖基转移酶 活性将在滋养层外植体培养中进行检测。的效果 病毒和化学转化、生长因子等 细胞分化的诱导剂/抑制剂对酶活性的影响 在适当的情况下，活动变化的动力学方面和 将研究DNA、RNA和蛋白质合成抑制剂的影响。

There are three aims to the proposed research, each with its own methodology but all with a common clinical objective - to identify and develop carbohydrate-specific tumor markers, with emphasis on hCG and the differential diagnosis of trophoblastic neoplasms. 1. Distribution of O-linked sugar units on pregnancy- and tumor-derived hCG. On hCG in pools of urine from normal pregnancy, 12-14% of the O-linked sugar units (4 total) are a hexasaccharide structure, NeuAc-Gal-GlcNAc- (NeuAc-Gal)-GalNAc-. In contrast, pilot studies show that the structure of those on hCG in the urines of choriocarcinoma subjects (2 examined) and in the media from choriocarcinoma cells were around 50% this hexasaccharide structure. The only published findings of this structure have been on cancer proteins. To confirm that hCG abundant in hexasaccharides is a choriocarcinoma marker, the structures of O-linked sugars on hCG molecules in the urines of 12-15 additional subjects will be examined (methods: immunoprecipitation of hCG, beta-elimination/labeling of O-linked sugar units, chromatographic comparison with oligosaccharide standards). To determine if the abundance of hexasaccharide is correlated with the malignance of trophoblastic disease, that on urinary hCG molecules from subjects with the more benign invasive/partial/complete mole will also be examined. O-linked sugar units on hCG from nontrophoblastic neoplasms, and on other urinary and tissue glycoproteins will be examined. 2. Development on trophoblast disease-specific assays. Although trophoblast disease is rare in the U.S.A. (1 in 1500 pregnancies), a 10 X higher occurrence is found in several African and Asian nations. To generate a disease-specific assay, the beta subunit C-terminal peptide containing the hexasaccharides on choriocarcinoma hCG will be isolated and utilized as immunogen for generating sheep polyclonal antisera, for monoclonal antibody approaches, and as domain for a lectin-antibody system. 3. Glycosyltransferase activities in normal, neoplastic, and induced or transformed cells. To investigate the pathways or mechanisms by which changes in protein glycosylation accompany cancer, glycosyltransferase activities will be examined in trophoblast explant cultures. Effects of virus and chemical transformation, growth factors, and other inducers/suppressants of cellular differentiation on enzyme activities will be examined; where appropriate kinetic aspects of activity changes and effects of DNA, RNA and protein synthesis inhibitors will be investigated.