Ho M K, Geist C, Murray J, Kato K, DeLellis R, Wolfe H, Carney W, Rabin H, Durda P
Medical Products Department, E.I. Du Pont de Nemours & Co., North Billerica, Massachusetts 01862.
Cancer Res. 1988 Sep 1;48(17):4969-75.
Using conventional murine hybridoma technology, we have produced a monoclonal antibody (MAb), 89E5, which recognizes two keratin-like polypeptides (Mr 53,000 and 45,000), which are preferentially expressed by epithelial tumors. In addition to detection of tumor cells by immunohistochemistry, MAb 89E5 was able to localize to tumor xenografts in nude mice after iodination of its F(ab')2 fragments. To develop potentially less immunogenic antibodies to antigens defined by MAb 89E5, studies were performed to produce a human counterpart to the mouse MAb. The mouse 89E5 MAb was used to purify the 89E5 polypeptides from tumor cell lines. The partially purified 89E5 antigen was then used to sensitize human splenic lymphocytes in vitro. Immortalization of the sensitized cells by cell fusion resulted in a human IgM MAb, PA1, which showed the same reactivity pattern on a panel of cell lines as did the mouse MAb 89E5. Immunofluorescent studies showed that both 89E5 and PA1 had staining patterns on epithelial cells indicative of antibodies to cytokeratin. Furthermore, PA1 immunoprecipitated two polypeptides (Mr 53,000 and 45,000) which comigrated with the 89E5 polypeptides. Competitive binding assays showed that the PA1 MAb and 89E5 MAb recognized closely associated epitopes. As with the 89E5 MAb, PA1 was reactive with tumor tissues in immunohistochemical studies. These studies indicate that the PA1 MAb is a human counterpart of the mouse 89E5 MAb. Direct comparison of human MAb and mouse MAb against the same antigen could yield valuable information on the efficacy of using human MAb in vivo.
利用传统的小鼠杂交瘤技术，我们制备了一种单克隆抗体（MAb）89E5，它能识别两种角蛋白样多肽（分子量分别为53,000和45,000），这两种多肽在上皮肿瘤中优先表达。除了通过免疫组织化学检测肿瘤细胞外，MAb 89E5的F(ab')2片段碘化后能够在裸鼠体内定位到肿瘤异种移植瘤上。为了开发针对由MAb 89E5定义的抗原的潜在免疫原性较低的抗体，开展了相关研究以制备小鼠MAb的人源对应物。小鼠89E5 MAb用于从肿瘤细胞系中纯化89E5多肽。然后用部分纯化的89E5抗原在体外使人类脾淋巴细胞致敏。通过细胞融合使致敏细胞永生化，得到了一种人IgM MAb PA1，它在一组细胞系上显示出与小鼠MAb 89E5相同的反应模式。免疫荧光研究表明，89E5和PA1在上皮细胞上的染色模式均表明它们是针对细胞角蛋白的抗体。此外，PA1免疫沉淀了两种与89E5多肽迁移率相同的多肽（分子量分别为53,000和45,000）。竞争性结合试验表明，PA1 MAb和89E5 MAb识别紧密相关的表位。与89E5 MAb一样，PA1在免疫组织化学研究中与肿瘤组织反应。这些研究表明，PA1 MAb是小鼠89E5 MAb的人源对应物。直接比较针对相同抗原的人源MAb和小鼠MAb可能会产生关于在体内使用人源MAb的疗效的有价值信息。
