Molecular Requirements for Recombinant Cytotoxins Efficacy

在绝大多数高级别胶质瘤(HGG)中发现白介素13(IL13)的限制性受体(R)。 病人。IL13的HGG相关受体被分子鉴定为IL13RA2，它属于 属于肿瘤抗原家族，称为癌症/睾丸肿瘤抗原(CTA)。CTA提供了高度特异性 癌症的分子靶向/识别。此外，基于IL13的细菌毒素，假单胞菌外毒素 含有A(PE)的重组细胞毒素被证明是治疗前脑胶质瘤的有效药物。 临床评估，第一代这些细胞毒素正在进行第三阶段临床试验。这个结构-- IL13的功能关系分析及其最近揭示的溶液结构证明了α-螺旋 D似乎在IL13与其HGG相关受体的结合中起着关键作用。现建议： 产生新的高度特异和高效的基于IL13的细胞毒素，将结合(I)新的 关于IL13结构-功能关系的知识，(Ii)关于胶质瘤细胞生物学的新信息，以及(Iii) 以前在临床环境中使用细胞毒素的经验。因此，这些新的细胞毒素将是 由基因工程形式的IL13和另一种细菌毒素白喉的衍生物组成 毒素，DT390。这些细胞毒素设计背后的主要思想是生产新的细胞毒素， 允许IL13的结合区，即D-螺旋，自由地被IL13的HGG相关受体获得 并消除细胞因子上与其正常生理受体相互作用的部位 在许多重要器官，包括中枢神经系统。IL13将被改造成具有DT毒素 部分远离其与IL13R；x2的结合部位，这是PE不能完全实现的。此外，IL13 α-螺旋D突变体是IL13RA2的超级激动剂，已经被鉴定出来，它们将用于 细胞毒素的优化设计。细胞毒素的限制性结合位点数和一个 胞内蛋白酶激活的细菌毒素(Furin)在细胞毒素对肿瘤细胞的杀伤中起着重要作用。 因此，将研究其表达水平的调节，并将其表达水平与 新型IL13-DT细胞毒素的功效。基于DT390-IL13突变体的新型细胞毒素将在体外和 为了在活体内演示分子设计如何在生物学背景下发生变化 靶标的表达和毒素的激活部分反映在它们的抗肿瘤效果上。这是意料之中的 收集到的新信息将对进一步的重组抗癌分子设计具有不可估量的价值 细胞毒素。

A restricted receptor (R) for interleukin 13 (IL13) was found in a vast majority of high-grade gliomas (HGG) patients. The HGG-associated receptor for IL13 was identified molecularly to be the IL13Ra2, and it belongs to a family of tumor antigens, termed cancer/testis tumor antigens (CTA). CTA provide high specificity for molecular targeting/recognition of cancer. Furthermore, IL13-based bacterial toxin, Pseudomonas exotoxin A (PE)-containing recombinant cytotoxins were documented to be the potent anti-glioma agents in pre- clinical evaluation, and the first generation of these cytotoxins is in Phase III clinical trials. The structure- function relationship analysis of IL13 and its recently revealed solution structure documented that alpha-helix D appears to play pivotal role in the binding of IL13to its HGG-associated receptor. It is proposed to generate novel highly specific and highly efficacious IL13-based cytotoxins that will incorporate (i) new knowledge on IL13 structure-function relationship, (ii) new information on glioma cell biology, and (iii) previous experience with the use of cytotoxins in a clinical setting. Thus, these new cytotoxins will be composed of genetically engineered forms of IL13and a derivative of another bacterial toxin, Diphtheria toxin, DT390. The principal idea behind the design of these cytotoxins is to produce novel cytotoxins that allow the binding region of IL13, the D-helix, to be freely available to the HGG-associated receptor forIL13 and to eliminate the site on the cytokine that interacts with its normal physiological receptor that is expressed in many vital organs, including the central nervous system. IL13will be engineered to have the DT toxin moiety remote form its binding site to the IL13R<x2, which cannot be fully achieved with PE. Moreover, IL13 alpha-helix D mutants, super agonists of the IL13Ra2, have been identified and they will be used in the design of optimized cytotoxins. The number of the restricted binding sites for the cytotoxins and an intracellular protease activating bacterial toxins (furin) play major roles in the cytotoxins' tumor cell killing. Thus, the regulation of their expression levels will be examined and the expression levels correlated with the novel IL13-DT cytotoxins efficacy. The novel DT390-IL13 mutant-based cytotoxins will be tested in vitro and in vivo in order to demonstrate how the changes in the molecular design in the context of biological expression of the target and toxin's activating moiety are reflected in their anti-tumor efficacy. It is expected that the new information gathered will be invaluable in further molecular design of recombinant anti-cancer cytotoxins.