Novel Dithiolethiones for Bladder Cancer Prevention

描述(申请人提供)：该项目致力于发现和开发用于预防膀胱癌的新型二硫代硫酮类化合物。许多证据表明，致癌物解毒第二阶段酶在预防膀胱癌方面发挥着重要作用。我们最近发现5，6-dihydrocyclopenta[c]-1，2-dithiole-3(4H)-thione(CPDT)在体内是一种非常有效的啮齿动物膀胱第二相酶的诱导剂，并假设CPDT或更有效的类似物将对膀胱癌起到高效的保护作用。该项目的第一个目的是进一步表征CPDT的第二相酶诱导特性，检测CPDT的毒性和药代动力学行为，并测试CPDT类似物以鉴定更有效的第二相酶诱导剂。这些研究将在大鼠身上进行；将测量测试化合物对三种重要的第二相酶(GST、NQO1和UGT)的影响；将合成CPDT类似物；将测量CPDT的各种毒性和药代动力学参数。目的2是确定CPDT及其更有效的类似物(在目标1中确定)诱导哪些GST和UGT亚型，这种诱导是否具有物种特异性，以及CPDT是否在细胞内代谢成新的诱导剂形式。GST和UGT的亚型被证明在预防膀胱癌中发挥重要作用，我们将使用人类和啮齿动物的膀胱上皮细胞来检测这两种亚型的反应。利用放射性标记的CPDT和LC/MS/MS研究CPDT的代谢。目的3阐明CPDT及其类似物诱导第二相酶的分子机制。我们假设Nrf2(一种转录因子)在介导这种诱导过程中起主要作用。目的4利用体外培养的膀胱上皮细胞和具有和不具有Nrf2功能的小鼠，通过DNA损伤和细胞/组织增殖的方法，评价CPDT或更有前景的类似物对膀胱癌致癌物遗传毒性的保护作用，并探讨Nrf2信号通路在介导这一作用中的作用。在AIM 4中进行的研究还将有助于确定在AIM 5中进一步实验的二硫代硫酮的合适剂量水平。目的5是利用野生型和Nrf2基因敲除小鼠，确定AIM 4中检测的二硫代硫酮在体内对化学致癌物诱导的膀胱肿瘤形成的抑制作用，以及Nrf2信号在介导这种作用中的作用。

DESCRIPTION (provided by applicant): This project is focused on identifying and developing novel dithiolethiones for the prevention of urinary bladder cancer. Many lines of evidence show that carcinogen-detoxifying Phase 2 enzymes play a major role in bladder cancer prevention. We have recently found that 5,6-dihydrocyclopenta[c]-1,2-dithiole-3(4H)-thione (CPDT) is an extremely efficacious inducer of Phase 2 enzymes in rodent bladders in vivo and hypothesize that CPDT or a more potent analog would be a highly effective protector against bladder cancer. Aim 1 of the project is to further characterize the Phase 2 enzyme-inducing property of CPDT, to examine the toxicity and pharmacokinetic behavior of CPDT, and to test CPDT analogs for identification of more potent inducers of Phase 2 enzymes. These studies will be performed in rats; the impact of the test compound on three important Phase 2 enzymes (GST, NQO1 and UGT) will be measured; CPDT analogs will be synthesized; various parameters of toxicity and pharmacokinetics of CPDT will be measured. Aim 2 is to determine which GST and UGT isoforms are induced by CPDT and its more potent analogs (identified in Aim 1), whether such induction is species specific, and if CPDT is metabolized in cells to a new inducer form. The response of the isoforms of GST and UGT which were shown to play important roles in bladder cancer prevention will be examined using bladder epithelial cells from humans and rodents. CPDT metabolism will be investigated using radiolabeled CPDT and LC/MS/MS. Aim 3 is to elucidate the molecular mechanism responsible for the induction of Phase 2 enzymes by CPDT and its analogs. We hypothesize that Nrf2 (a transcriptional factor) plays a major role in mediating such an induction. Aim 4 is to assess the protective effects of CPDT or a more promising analog against the genotoxicities of bladder carcinogens in bladder cells both in vitro and in vivo, as measured by DMA damage and cell/tissue proliferation, and to investigate the role of the Nrf2 signaling pathway in mediating such an effect, using cultured bladder epithelial cells and mice with and without Nrf2 function. Studies carried out in Aim 4 will also serve to identify the appropriate dose levels of the dithiolethione for further experimentation in Aim 5. Aim 5 is to determine the inhibitory effect of the dithiolethione examined in Aim 4 on bladder tumorigenesis induced by a chemical carcinogen in vivo and the role of Nrf2 signaling in mediating such an effect, using wild type and Nrf2 knockout mice.