Mechanism-based Optimization of Tirapazamine Analogs

该项目有三大目标，每一个目标都与该项目的总体目标有关，即开发针对肿瘤缺氧细胞的新疗法。进一步整合所有三个目标的主题是使用多细胞层（MCL）培养来量化药物及其活性代谢物的血管外运输。第一个目标是开发一种改进的低氧选择性细胞毒素替拉帕嗪（TPZ）的类似物，该药物目前处于III期临床试验中。该方法基于我们对TPZ的空间分辨药代动力学/药效学（PK/PD）模型的见解，该模型明确考虑了血管外运输。该模型表明，通过增加血管外扩散系数、优化代谢速率和改善代谢中导致细胞毒性的部分（lambda），可以获得较大的活性增益。我们将使用PK/PD模型通过筛选来指导化合物的进展，并将评估模型中每个成分的药物构效关系（例如组织中的扩散系数）。我们的方法将测试特定的假设：(i) DNA靶向可用于提高细胞毒性效力和lambda，以及（ii）增加TPZ自由基的细胞内扩散范围是增加lambda的进一步策略。第二个目标是描述肿瘤坏死和缺氧区重组梭菌表达的酶激活前药所产生的旁观者效应。这将涉及鉴定由大肠杆菌硝基还原酶、β -葡萄糖醛酸酶和β -半乳糖苷酶激活前药产生的活性代谢物，并量化活性物种在mcl中的血管外运输和由此产生的旁观者死亡。第三个目标是确定从筛选针对hif -1 α上调的新药中，哪些初始“命中”具有足够好的血管外运输特性，以保证进一步开发。

This Project has three broad goals, each of which relate to the overall objective of this Program to develop novel therapies for hypoxic cells in tumors. A further integrating theme across all three goals is use of multicellular layer (MCL) cultures to quantify extravascular transport of drugs and their active metabolites. The first goal is to develop an improved analog of the hypoxia-selective cytotoxin tirapazamine (TPZ), which is currently in phase III clinical trial. The approach is based on insights from our spatially-resolved pharmacokinetic/pharmacodynamic (PK/PD) model for TPZ, which explicitly takes extravascular transport into account. This model demonstrates that large gains in activity could be achieved by increasing extravascular diffusion coefficients, optimizing rates of metabolism, and improving the fraction of metabolism that contributes to cytotoxicity (lambda). We will use the PK/PD model to guide advancement of compounds through screening, and will evaluate drug structure-activity relationships for each component of the model (e.g. diffusion coefficient in tissue). Our approach will test the specific hypotheses that (i) DNA targeting can be used to improve the cytotoxic potency and lambda, and (ii) increasing the intracellular diffusion range of the TPZ radical is a further strategy for increasing lambda. The second goal is to characterize bystander effects resulting from activation of prodrugs by enzymes expressed by recombinant clostridia in necrotic and hypoxic regions of tumors. This will involve the identification of active metabolites resulting from activation of prodrugs by E. coli nitroreductase, beta-glucuronidase and beta-galactosidase, and to quantify extravascular transport of the active species, and the resulting bystander killing, in MCLs. The third goal is to identify which of the initial "hits" from a screen for novel drugs targeting HIF-1alpha upregulation have good enough extravascular transport properties to warrant further development.