DESIGN AND USE OF CD7-SPECIFIC RECOMBINANT IMMUNOTOXINS

本申请的长期目标是开发新的 用于人骨髓治疗的重组免疫毒素 移植和体内治疗人白血病， 淋巴瘤 重组免疫毒素包括一类新的 被设计为选择性细胞毒性的药物。 他们是 融合蛋白，其中抗原结合（可变）区 抗体与强效催化毒素相连 基因 工程免疫毒素有几种关于功效的假设 重组免疫毒素和化学连接免疫毒素的区别。 对泛T细胞抗原CD 7具有特异性的重组免疫毒素将 作为一种模型，免疫毒素可能被证明是有效的， T-ALL的治疗 在第一个具体目标中，可变区 由CD 7表达的基因将被用作模型系统。 CD 7是 几乎在所有T细胞急性淋巴细胞白血病（T-ALL）上表达 因此这些免疫毒素可能被证明是有效的治疗 T-ALL。 在第一个具体目标中，通过以下方式表达可变区基因： 将CD 7特异性杂交瘤与蓖麻毒素A链（RTA）连接， 基因以不同的方式来确定哪个编码最有效 免疫毒素 这些化学缀合至RTA的效力。 的 体内比较将使用T-ALL的SCID小鼠模型。 内化、药代动力学和生物分布研究将 检查哪些因素影响重组体的体内功效 和化学免疫缀合物。 在第二个具体目标重组中， 含有假单胞菌外毒素变体的CD 7特异性免疫毒素 将构建并测定A（PE 38 KDEL）。 施行本 实验的目的是确定RTA和PE 38 KDEL在细胞毒性方面是否不同 当针对CD 7时。 第三个具体目标针对的是一个问题 在RTA免疫毒素的所有临床试验中遇到：剂量限制 由内皮细胞损伤引起的毒性。 假设RTA 单独使用可通过受体介导的内吞作用杀死内皮细胞。 将产生RTA的突变体并筛选酶活性 不再结合和杀死内皮细胞的变体。 的 这些突变体的鉴定应该允许构建更多的 治疗有效的免疫毒素。

The long-term objective of this application is to develop novel recombinant immunotoxins for therapeutic use in human bone marrow transplantation and for the in vivo treatment of human leukemias and lymphomas. Recombinant immunotoxins comprise a new class of pharmacologic agents designed to be selectively cytotoxic. They are fusion proteins in which the antigen-binding (variable) regions of antibodies are linked to potent, catalytic toxins. Genetically engineered immunotoxins have several hypotheses concerning the efficacies of recombinant versus chemically-linked immunotoxins. Recombinant immunotoxins specific for the pan -T cell antigen CD7 will be used as a model immunotoxins may prove to be effective in the treatment of T -ALL. In the first specific aim, the variable region genes expressed by a CD7 will be used as a model system. CD7 is expressed on virtually all T cell acute lymphoblastic leukemias (T-ALL) and so these immunotoxins may prove to be effective in the treatment of T-ALL. In the first specific aim the variable region genes expressed by a CD7-specific hybridoma will be linked to the ricin toxin A chain (RTA) gene in different ways to determine which encodes the most efficacious immunotoxin. The potency of the these chemically conjugated to RTA. The in vivo comparisons will employ SCID mouse models for T-ALL. Internalization, pharmacokinetic, and biodistribution studies will examine what factors influence the in vivo efficacy of the recombinant and chemical immunoconjugates. In the second specific aim recombinant CD7-specific immunotoxins containing a variant of Pseudomonas exotoxin A (PE38KDEL) will be constructed and assayed. The purpose of these experiments is to determine if RTA and PE38KDEL differ in cytotoxicity when targeted to CD7. The third specific aim addresses a problem encountered in all clinical trials of RTA-immunotoxins: dose-limiting toxicity caused by endothelial cell damage. The hypothesis is that RTA alone can kill endothelial cells via receptor-mediated endocytosis. Mutants of RTA will be generated and screened for enzymatically active variants that no longer bind and kill endothelial cells. The identification of these mutants should allow for the construction of more therapeutically effective immunotoxins.