Oxidative Pentose Cycle in Hypoxic Cancer Cell Response

描述（由申请人提供）：缺氧在癌细胞抵抗DNA损伤剂中的作用已经在体外和体内得到了清楚的证明。一些临床研究也证明了缺氧肿瘤的治疗效果很差。DNA修复、修复蛋白Kf和细胞内谷胱甘肽解毒在细胞对DNA损伤拓扑异构酶II抑制剂和γ辐射的反应中起主要作用。虽然这些因子的功能可以被功能性硫醇的氧化抑制，但细胞通过氧化戊糖磷酸循环（OPPC）产生NADPH来抵抗硫醇氧化还原变化。例如，羟乙基二硫醚（HEDS）是一种独特的无毒硫醇氧化剂，它不能有效地用OPPC修饰正常CHO细胞中的细胞硫醇氧化还原。我们的初步研究首次表明，OPPC活性受损的CHO突变体是heds介导的DNA损伤剂致敏的理想候选者。OPPC的独特特性消除了hads介导的硫醇氧化还原修饰和DNA损伤敏化，将被用于敏化缺氧的癌细胞，而不影响葡萄糖精通的正常细胞。葡萄糖剥夺在一些实体瘤的缺氧细胞中很常见，因为血管化差和代谢活性高。

DESCRIPTION (provided by applicant): The role of hypoxia in the resistance of cancer cells to DNA damaging agents has been clearly demonstrated in in vitro and in vivo. Several clinical studies have also demonstrated poor therapeutic outcome for hypoxic tumors. DNA repair, repair protein Kf, and intracellular GSH detoxification play a major role in cellular response to DNA damaging topoisomerase II inhibitors and gamma radiation. Although the functions of these factors can be inhibited by oxidation of functional thiols, cells resist thiol redox changes by producing NADPH through oxidative pentose phosphate cycle (OPPC). For example hydroxyethyldisulfide (HEDS), a unique non-toxic thiol oxidant, is not effective in modifying the cellular thiol redox in normal CHO cells with OPPC. Our preliminary studies indicated, for the first time, that CHO mutants impaired with OPPC activity are the ideal candidates for HEDS-mediated sensitization to DNA damaging agents. This unique property of OPPC, which eliminates HEDS-mediated thiol redox modification and DNA damage sensitization, will be exploited to sensitize hypoxic cancer cells deprived of glucose, a substrate for OPPC, without affecting glucose proficient normal cells. Glucose deprivation is common in hypoxic cells of several solid tumors because of poor vascularization and higher metabolic activity. We hypothesize that glucose/OPPC depleted hypoxic cancer cells are susceptible to HEDS-mediated redox [Protein thiol (PSH) & glutathione (GSH)] modification and sensitization to "and tumor and DNA damaging" Topo II inhibitors and gamma radiation through multiple mechanisms that include inhibition of DNA repair, DNA repair protein function, GSH detoxification, and anti-apoptotic factors. We will test our hypothesis in hypoxic human cancer cells using biochemical and molecular approaches. First, we will determine the application of thiol redox modification in the sensitization of glucose deprived hypoxic cancer cells to topo II inhibitors and gamma radiation. Additional studies will also be conducted to test the efficacy of HEDS in low glucose hypoxic tumor in in vivo. Second, using siRNA and antibody technologies, we will determine whether specific inhibitors of OPPC can also induce HEDS-mediated redox modification, and subsequent sensitization of hypoxic cancer cells to DNA damaging agents. Finally, we will determine the mechanisms of redox modification- mediated hypoxic sensitization by determining its effect on DNA double strand breaks, DNA repair protein Kf, GSH detoxification and pro-apoptotic factors.