CLONING OF ANTI-TUMOR ANTIGEN IMMUNOGLOBIN GENES

这项研究项目的主要目标是进行基因工程 免疫球蛋白基因产生用于体内定位的有用试剂 以及人类肿瘤的治疗。几个杂交瘤细胞系已经被 在这个实验室中开发的产生单抗(MAb)的 选择性地与肿瘤相关抗原反应。其中包括 癌胚抗原和肿瘤相关糖蛋白Tag-72，它 存在于多种癌症中。单抗目前正被用于 一些关于乳房、结肠和卵巢的诊断和治疗试验 癌症。在临床试验中，一种这样的单抗B72.3，是针对一种 人转移性乳腺癌的膜富集物和 识别TAG-72，已显示出被开发为诊断程序的希望 和治疗剂。小鼠单抗在体内治疗和治疗中的作用 然而，人类肿瘤的诊断是有限的，因为它们 患者的免疫原性。此问题可以通过将 小鼠抗体的恒定区与人的恒定区 抗体，使用重组DNA技术。为此，我们克隆了 重排的重链可变区和轻链可变区以及 单抗B72.3的重链和轻链基因信息的拷贝。 已构建了重链和轻链嵌合免疫球蛋白基因。 将它们分别插入适当的表达载体。稳定 通过引入这些抗体，建立了嵌合抗体产生细胞系 构建成成纤维细胞和骨髓瘤细胞系。嵌合抗体 由cdna克隆或保留的重排基因的克隆产生 亲本抗体的特异性和结合特性。

The main objective of this research project is to genetically engineer immunoglobulin genes to generate useful reagents for in vivo localization and therapy of human tumors. Several hybridoma cell lines have been developed in this laboratory that produce monoclonal antibodies (MAbs) that selectively react with tumor-associated antigens. These include the carcinoembryonic antigen and a tumor-associated glycoprotein, TAG-72, which is present in a variety of carcinomas. The MAbs are currently being used in a number of diagnostic and therapeutic trials on breast, colon, and ovarian cancers. In clinical tests, one such MAb, B72.3, which was raised against a membrane-enriched extract of a human metastatic breast carcinoma and recognizes TAG-72, has shown promise for being developed into a diagnostic and therapeutic agent. The usefulness of mouse MAbs for in vivo therapy and diagnosis of human tumors, however, is limited because of their immunogenicity in patients. This problem can be reduced by replacing the constant region of the mouse antibody with the constant region of the human antibody, using recombinant DNA techniques. To that end, we have cloned the rearranged heavy and light chain variable regions as well as the cDNA copies of the heavy and light chain gene messages of the MAb, B72.3. Chimeric heavy and light chain immunoglobulin genes have been constructed. They were inserted separately into appropriate expression vectors. Stable chimeric antibody-producing cell lines were developed by introducing these constructs into both fibroblast and myeloma cell lines. Chimeric antibody produced either from cDNA clones or clones of the rearranged genes retained specificity and binding properties of the parental antibody.