Epigenetic therapy with 3-deazaneplanocin A and gemcitabine for pancreatic cancer

描述（由申请人提供）：尽管吉西他滨对早期胰腺肿瘤具有卓越的细胞毒性，但由于肿瘤细胞的侵袭性生长特性和凋亡抗性，吉西他滨在晚期肿瘤中实际上无效。文献证据表明，表观遗传改变（如组蛋白和DNA超甲基化）可能在沉默肿瘤抑制基因中发挥重要作用，使肿瘤细胞克隆扩增并抵抗细胞毒性。我们的长期目标是改善胰腺癌的化疗管理。这项R03基金申请的总体目标是测试表观遗传沉默的肿瘤抑制基因的再激活可以增强胰腺癌的化学致敏性的概念。具体来说，我们的中心假设是一种基于机制的结构优化的新型组蛋白去甲基化剂，3'deazaneplanocin-A (DZnep)，与吉西他滨联合将在胰腺癌细胞中产生优越的细胞毒性反应。这一假设是基于我们的初步数据，该数据显示DZnep在培养的胰腺癌细胞中转运依赖性地增强了吉西他滨的化学致敏性。该研究的基本原理是，合成去甲基化药物改善吉西他滨化疗敏感性的证据将为继续研究表观遗传治疗胰腺癌的潜力提供实验基础。中心假设将通过追求三个具体目标来检验。特异性目的1将确定胰腺癌中DZnep的细胞转运和激活机制。目的1的工作假设是亲水性DZnep的去甲基化和化学致敏能力将受到载体介导的转运过程的速率限制。这一假设是基于我们的初步数据，表明当核苷运输活性被抑制时，dznep诱导的反应显著减少。特异性目标2将在体外生成和测试一系列DZnep酰基衍生物。目的2的工作假设是，n4取代的脂肪酸酰胺衍生物会增加DZnep的亲脂性，并允许它绕过细胞内激活的运输要求。我们的假设是基于我们的初步研究，表明另一种核苷（曲沙他滨）的酰基前药绕过了运输要求。特异性目的3将确定DZnep前药-吉西他滨联合用药的体内抗癌效果。我们的工作假设是，DZnep前药与吉西他滨联合治疗携带胰腺癌异种移植的小鼠，与单独使用吉西他滨治疗相比，将显示出更好的化疗效果。这一假设是基于细胞培养中DZnep和吉西他滨联合方案获得的初步体外结果的外推。这些研究的贡献将是显著的，因为成功证明了一种优越的表观遗传化疗方案可能取代效果较差的吉西他滨单药治疗实践。这项工作具有创新性，因为它首次提出了一种基于核苷的表观遗传化疗治疗胰腺癌的可能性。

DESCRIPTION (provided by applicant): Despite superior cytotoxicity against early-stage pancreatic tumors, gemcitabine is practically ineffective in late-stage tumors due to the invasive growth characteristics and apoptotic resistance of tumor cells. Evidence in literature suggest that epigenetic alterations (e.g. histone and DNA hypermethylation) may play vital roles in silencing tumor suppressor genes, allowing tumor cells to clonally expand and resist cytotoxicity. Our long-term goal is to improve the chemotherapeutic management of pancreatic cancer. The overall objective of this R03 grant application is to test the concept that the reactivation of epigenetically silenced tumor suppressor genes can augment chemosensitization in pancreatic cancer. Specifically, it is our central hypothesis that a mechanism-based structure optimization of a novel histone-demethylation agent, 3'deazaneplanocin-A (DZnep), in combination with gemcitabine will enact a superior cytotoxic response in pancreatic cancer cells. This hypothesis is based on our preliminary data that show transport-dependent augmentation of gemcitabine chemosensitization by DZnep in cultured pancreatic cancer cells. The rationale underlying the proposed research is that proof of synthetic demethylating agents improving gemcitabine chemosensitivity will provide an experimental basis to continue investigating the potential of epigenetic therapy for pancreatic cancer. The central hypothesis will be tested by pursuing three specific aims. Specific aim 1 will determine the cellular transport and activation mechanisms of DZnep in pancreatic cancer. The working hypothesis for aim 1 is that the demethylating and chemosensitizing abilities of hydrophilic DZnep will be rate-limited by a carrier-mediated transport process. This hypothesis is based on our preliminary data that show a significant reduction in DZnep-induced responses when nucleoside transport activity was inhibited. Specific aim 2 will generate and test a battery of DZnep acyl derivatives in vitro. The working hypothesis for aim 2 is that N4-substituted fatty acid amide derivatives will increase the lipophilicity of DZnep and allow it to bypass the transport requirement for intracellular activation. Our hypothesis is based on our preliminary studies showing the acyl prodrugs of another nucleoside (troxacitabine) bypassing the transport requirement. Specific aim 3 will determine the in vivo anti-cancer efficacies of DZnep prodrug-gemcitabine combinations. Our working hypothesis is that the combined treatment of a DZnep prodrug with gemcitabine in mice carrying pancreatic cancer xenografts will exhibit superior chemotherapeutic efficacy compared with gemcitabine treatment alone. This hypothesis is based on the extrapolation of preliminary in vitro results obtained with DZnep and gemcitabine combination schedules in cell culture. The contribution from these studies will be significant because successful demonstration of a superior epigenetic-chemotherapeutic regimen is likely to replace the less efficacious gemcitabine monotherapy practice. The proposed work is innovative because it addresses for the first time the possibility of a nucleoside-based epigenetic-chemotherapeutic therapy in pancreatic cancer. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because pancreatic cancer is a dire problem in the US with an estimated death of about 35,000 people annually. Since gemcitabine monotherapy is not very effective, epigenetic agents in combination with standard chemotherapeutic care will act as a superior treatment regimen necessary for improving survival in pancreatic cancer patients.