TARGETING ENDOGENOUS ANTIBODIES TO OVARIAN CARCINOMA

描述(由申请人提供)： 人类和东半球灵长类动物自然会产生大量的 识别特定半乳糖表位的抗体，Galalal1-3GAL。 我们一直有兴趣测试这些抗Gal抗体是否可以 针对不受欢迎的癌细胞。具体地说，我们希望探索一个 将这些内源性抗体重定向至卵巢癌的可能性 过表达叶酸受体同型α(FR-α)的细胞 叶酸与半乳糖表位的化学偶联。最终的结果是 应该是靶细胞的细胞溶解。为了这个目标，现在 应用涉及叶酸-双半乳糖的全合成 免疫活性卵巢癌模型的建立及结合物的研究 老鼠。 可介导抗Gal与FR细胞结合的结合物的制备 具有高亲和力是该项目的主要化学目标。我们的战略是 向一个分子引入表位的多个拷贝，2个和4个拷贝 在它们之间的最佳距离。化学合成将是 在固相支撑体上进行。这些结合物将进行测试 FR/Gal-人鼻咽癌KB细胞促增殖能力的研究 抗Gal与细胞表面的FR结合。相互作用的特殊性 将在存在游离叶酸或游离二糖的情况下进行测试 与含有乳糖而不是半乳糖α1-3GAL的结合物一样。这个 利用流式细胞仪可以方便地表征抗体结合。 程序。 该项目的生物学目标是开发一种小鼠卵巢模型。 适合测试我们的叶酸抗肿瘤活性的癌症 在体内结合。由于正常小鼠在其体内表达半乳糖表位 组织，我们将不得不使用α-1，3-半乳糖基转移酶基因敲除(GT/KO) 老鼠。已知GT/KO小鼠与人类一样产生抗Gal抗体。我们计划 将培养中的小鼠卵巢上皮细胞转化为 在我们最近开发的一种程序下建立肿瘤形成细胞系。 然后将它们与编码全长FR-α的小鼠c DNA转染。 最后将这些GAL-/FR细胞导入健康人的腹膜 GT/KO小鼠。 我们目前对卵巢癌免疫治疗的方法是独特的，因为我们 使用的是自然产生的内源性抗体。免疫调节剂在这方面 应用都是分子量为3 kDa的小分子，呈现药动学 最有利于腹膜腔持续活动的特性也是如此 减少了潜在的副作用。

DESCRIPTION (provided by applicant): Humans and Old World primates naturally produce a significant amount of antibodies which recognize a particular galactosyl epitope, GAL alpha 1-3GAL. We have been interested in testing if these anti-Gal antibodies can be targeted to undesirable cancerous cells. Specifically we wish to explore a possibility of redirecting these endogenous antibodies to ovarian carcinoma cells which overexpress folate receptor isotype alpha (FR-alpha) by means of chemical conjugates of folic acid to the galactosyl epitope. The end result should be the cytolysis of the target cell. Towards this goal, the present application is concerned with the total synthesis of the folate-digalactose conjugates and development of an ovarian cancer model in immune competent mice. Preparation of the conjugate which can mediate anti-Gal binding to FR+ cells with high avidity is the main chemistry goal of the project. Our strategy is to introduce multiple copies, 2 and 4 copies, of the epitopes to one molecule of folic acid at an optimal distance between them. Chemical synthesis will be carried out on a solid-phase support. The conjugates will be tested with FR+/Gal- human nasopharyngeal carcinoma KB cells for their ability of promoting the anti-Gal binding to FR on the cell surface. Specificity of the interaction will be tested in the presence of free folic acid or free disaccharide as well as with a conjugate that contains lactose instead of GAL alpha 1-3GAL. The antibody binding will be conveniently characterized by means of FACS procedure. The biological goal of this project is to develop a mouse model of ovarian cancer that is suitable for testing anti-tumor activity of our folate conjugates in vivo. Since normal mice express the galactosyl epitopes in their tissue, we will have to use alpha 1,3-GALactosyltransferase-knockout (GT/KO) mice. It is known that GT/KO mice produce anti-Gal as in humans. We plan to transform the ovarian epithelial cells harvested from these mice in culture to tumor-forming cell lines following a procedure we have recently developed. They will be then transfected with murine cDNA encoding full length FR-alpha. Finally these GAL-/FR+cells will be introduced into peritoneum of healthy GT/KO mice. Our current approach to immunotherapy of ovarian cancer is unique in that we are using naturally occurring endogenous antibodies. Immune modulators in this application are all small molecules with MW < 3 kDa, rendering pharmacokinetic properties most favorable for sustained activity in peritoneal cavity as well as reduced potential side effects.