ASCORBATE-INDUCED RADIOSENSITIZATION IN PANCREATIC CANCER

描述（由申请人提供）：胰腺腺癌是美国癌症死亡的第四大原因，并且发病率正在上升；预后仍然令人沮丧。我们建议研究一种全新的方法，使用药物抗坏血酸作为辅助放疗，治疗胰腺癌。静脉注射抗坏血酸（即抗坏血酸、维生素C），而不是口服抗坏血酸，会产生高血浆浓度，其浓度在对肿瘤细胞具有细胞毒性的范围内。我们小组的初步研究表明，抗坏血酸可诱导胰腺癌细胞的氧化应激和细胞毒性；这种细胞毒性似乎在肿瘤细胞中比在正常细胞中更大。我们已经确定，抗坏血酸是过氧化氢（H2O2）在体外和体内通过其自氧化传递的前药。我们之前资助的R21基金CA137230“抗坏血酸诱导胰腺癌细胞毒性的机制”的研究表明，抗坏血酸在人类可达到的剂量下，通过h2o2介导的机制降低了所有胰腺癌细胞系的活力和增殖。目前的应用扩展了这项临床前工作，以确定该策略是否可以有效地用作胰腺癌放射治疗的辅助手段。如果胰腺癌细胞（相对于正常细胞）更容易受到抗坏血酸诱导的细胞毒性的影响，因为金属离子催化的抗坏血酸自氧化作用增加，导致H2O2的产生增加，那么使用抗坏血酸作为佐剂将有望使胰腺肿瘤细胞对标准治疗（如电离辐射，增加活性氧水平）敏感。我们将使用生物化学/分子生物学技术来确定抗坏血酸诱导的放射致敏和氧化应激在体外和体内的相对易感性；我们将使用FLT-PET（氟标记胸腺正电子发射断层扫描）作为一种非侵入性的体内细胞增殖指标。本建议的重点是使用补充方法（高剂量抗坏血酸）改善标准抗癌治疗（电离辐射）在治疗胰腺癌中的治疗比例。目前的提议将在体外和体内验证金属离子催化抗坏血酸自氧化产生H2O2诱导人类胰腺癌细胞放射增敏的假设。我们将用以下两个具体目标来检验我们的假设：1。通过金属离子催化抗坏血酸自氧化产生H2O2的通量，确定抗坏血酸是否选择性地诱导人胰腺癌细胞（相对于正常细胞）的放射致敏。2. 确定抗坏血酸是否选择性地诱导人胰腺癌细胞在体内的放射致敏。如果我们能够严格证明抗坏血酸介导的放射致敏作用在人类胰腺癌细胞中诱导了优先氧化应激和随后的细胞毒性，那么这一应用的结果将为合理设计一种对NCCAM的RFA PA-09-167高度敏感的联合癌症治疗模式提供基础，该模式名为“癌症治疗补充方法的发展项目”。

DESCRIPTION (provided by applicant): Adenocarcinoma of the pancreas is the fourth leading cause of cancer death in the United States and is increasing in incidence; the prognosis remains dismal. We propose to investigate an entirely new approach, using pharmacological ascorbate as an adjuvant to radio-therapy, to treat pancreas cancer. Intravenous ascorbate (i.e., ascorbic acid, vitamin C), but not oral ascorbate, produces high plasma concentrations, which are in the range that are cytotoxic to tumor cells. Preliminary studies from our group have demonstrated that ascorbate induces oxidative stress and cytotoxicity in pancreatic cancer cells; this cytotoxicity appears to be greater in tumor vs. normal cells. We have firmly established that pharmacological ascorbate is a pro-drug for delivery of hydrogen peroxide (H2O2) in vitro and in vivo via its autoxidation. Studies from our previously funded R21 grant CA137230 "Mechanisms of ascorbate-induced cytotoxicity in pancreatic cancer" have demonstrated that ascorbate, in doses achievable in humans, decreases viability and proliferation in all pancreatic cancer cell lines examined, via a H2O2-mediated mechanism. The current application extends this preclinical work to determine if this strategy can be used effectively as an adjuvant in radiotherapy for pancreatic cancer. If pancreatic cancer cells (relative to normal cells) are more susceptible to ascorbate-induced cytotoxicity due to increased metal ion catalyzed ascorbate autoxidation leading to increased production of H2O2, then using ascorbate as an adjuvant would be expected to sensitize pancreatic tumor cells to standard therapies such as ionizing radiation that increase levels of reactive oxygen species. We will use biochemistry/molecular biology techniques to determine relative susceptibility to ascorbate-induced radiosensitization and oxidative stress in vitro and in vivo; we will employ FLT-PET (fluoro-labeled thymidine positron emission tomography) as a non- invasive in vivo index of cell proliferation. This proposal focuses on improvement of the therapeutic ratio of a standard anti-cancer therapy (ionizing radiation) using a complementary approach (high dose ascorbate), in the treatment of pancreatic cancer. The current proposal will test the hypothesis that metal ion catalyzed H2O2 generation from the auto-oxidation of ascorbate induces radiosensitization of human pancreatic cancer cells in vitro and in vivo. We will test our hypotheses with the following two Specific Aims: 1. Determine if ascorbate selectively induces radio-sensitization of human pancreatic cancer cells (vs. normal cells) in vitro via the flux of H2O2 generated from metal ion catalyzed auto-oxidation of ascorbate. 2. Determine if ascorbate selectively induces radio-sensitization of human pancreatic cancer cells in vivo. If we can rigorously demonstrate that the radiosensitization mediated by pharmacological ascorbate induces preferential oxidative stress and subsequent cytotoxicity in human pancreatic cancer cells, then the results of this application will provide a foundation for the rational design of a combined modality cancer therapy that is highly responsive to the RFA PA-09-167 from NCCAM entitled "Developmental Projects in Complementary Approaches to Cancer Care".