ANTI-TUMOR EFFECTS OF ANTI-IDIOTYPE ANTIBODIES

在B细胞患者中观察到了实质性的抗肿瘤作用 针对该独特型的鼠源性单抗治疗淋巴瘤 细胞表面免疫球蛋白对恶性B细胞的影响。虽然这件事 这种疗法代表了治疗B细胞的一种令人兴奋的方法 恶性肿瘤，需要了解更多关于癌症的原因 这种疗法的成功或失败。在关键因素中， 需要研究的是抗独特型抗体的特征，它可能 在最终的抗肿瘤反应中发挥作用及其机制 肿瘤细胞逃脱了这些抗体的控制。我们一直专注于 确定抗体同型、表位等特征的作用 抗独特型抗体的特异性、亲和力和抗原调节能力 B细胞淋巴瘤小鼠模型38C13中的抗体。到目前为止，角色 在这个模型中已经明确地定义了抗体同种类型的 免疫选择对独特型生长的抗独特型作用 不再与治疗性抗体发生反应的变种 被发现了。此外，已确定免疫表型 逃脱肿瘤的能力可以作为抗肿瘤效果的额外标志， 分离出的变异体可以用来更准确地定义抗体 表位特异性，而转义变异克隆表达不同的 来自亲代肿瘤的轻链基因。一大批新的单抗 最近产生了抗独特型抗体，它包含 在表位特异性和结合方面存在明显差异的抗体 与亲本肿瘤及其变异体的亲和力。我们现在提议 为了更准确地定义我们新派生的抗体小组 亲和力和表位特异性。这将涉及到量化 结合常数通过Scatchard分析、结合竞争分析和 Western blotting。这些抗体的体外相对作用 靶肿瘤细胞增殖、细胞表面独特型抗原 表达(抗原调节)，抗体依赖的细胞 肿瘤细胞的细胞毒性和补体依赖的裂解 分析过了。由于体外的发现，选定的感兴趣的抗体， 将在体内进一步评估它们的抗肿瘤作用。这些 将通过分析荷瘤患者的生存时间来评估疗效。 并用流式细胞术对逃逸动物进行免疫表型分析 肿瘤。后一种分析将被用来确定 野生型肿瘤细胞与独特型变异型肿瘤细胞的比例 逃离肿瘤种群。抗体同型匹配，但不同 在抗增殖作用方面，表位特异性、亲和力和 调制能力将在这些活体研究中进行比较。最后， 肿瘤细胞逃避抗体治疗的本质将进一步 从免疫表型和分子水平进行研究。研究将会是 旨在确定独特型变异的产生过程是否 是一个持续的过程，它的频率是多少，分子是什么 在各种不同的选择出的变体之间存在关系 治疗性抗体。独特型变异分子的蛋白质研究 免疫球蛋白基因的SDS-PAGE、DNA Southern杂交分析 通过聚合酶链进行重排和核苷酸直接测序 反应技术将在变异克隆上进行。上述研究 将有望提供有关Rational的重要相关信息 淋巴瘤及其他疾病治疗中抗体的选择 恶性肿瘤，并阐明了异质性和免疫生物学。 淋巴瘤。

Substantial anti-tumor effects have been observed in patients with B-cell lymphoma treated with murine monoclonal antibodies directed at the idiotype of the cell surface immunoglobulin on malignant B-cells. Although this type of therapy represents an exciting approach to the treatment of B-cell malignancies, much needs to be learned regarding the reasons for the success or failure of such a therapy. Among the critical factors which require study are the characteristics of anti-idiotype antibodies which may play a role in the eventual anti-tumor response and the mechanisms whereby tumor cells escape control by these antibodies. We have focused on defining the role of such characteristics as antibody isotype, epitope specificity, affinity and antigenic modulation capacity using anti-idiotype antibodies in a murine model of B-cell lymphoma, 38C13. To date, the role of antibody isotype has been unambiguously defined in this model and a novel anti-idiotype effect of immunoselection for the outgrowth of idiotype variants which are no longer reactive with the therapeutic antibody has been discovered. Moreover, it has been determined that the immunophenotype of escaping tumors can serve as an additional marker of anti-tumor effects, that the isolated variants can be used to more precisely define antibody epitope specificity, and that escaping variant clones express a different light chain gene from the parental tumor. A large panel of new monoclonal anti-idiotype antibodies has recently been generated which contains antibodies with distinct differences in epitope specificity and binding affinity with respect to the parent tumor and its variants. We now propose to more precisely define our newly derived panel of antibodies with respect to affinity and epitope specificity. This will involve quantitation of binding constants by Scatchard analysis, binding competition assays, and Western blotting. The relative in vitro effects of these antibodies on proliferation of target tumor cells, cell surface idiotype antigen expression (antigenic modulation), antibody-dependent cellular cytotoxicity, and complement-dependent lysis of tumor cells will be analyzed. Selected antibodies of interest, because of in vitro findings, will be further assessed in vivo for their anti-tumor effects. These effects will be evaluated by analyzing the survival time of tumor-bearing animals and by immunophenotypic analysis by flow cytometry of escaping tumors. This latter assay will be used to determine the relative proportion of "wild type" tumor cells to idiotype variant tumor cells in escaping tumor populations. Antibodies matched for isotype but differing in anti-proliferative effects, epitope specificity, affinity, and modulation capacity will be compared in these in vivo studies. Finally, the nature of tumor cells escaping antibody therapy will be further investigated on an immunophenotypic and molecular level. Studies will be directed at determining whether the process of idiotype variant generation is an ongoing precess, what its frequency is, and what molecular relationships exist between the variants selected out by various therapeutic antibodies. Protein studies of idiotype variant molecules by SDS-PAGE, DNA analysis by Southern blotting of immunoglobulin gene rearrangements, and direct nucleotide sequencing by the polymerase chain reaction technique will be performed on variant clones. The above studies will hopefully provide important and relevant information on the rational selection of antibodies for therapy of lymphoma as well as other malignancies and shed light on the heterogeneity and immunobiology of lymphoma.