Prevention of COX-2 Derived DNA and Histone Modifications in Cancer

 描述（由申请方提供）：将在拟定的研究中检查环加氧酶（考克斯）的高反应性代谢产物促进COX依赖性癌症生长的假设。一致的证据表明，COX有助于恶性肿瘤的演变，它包括流行病学研究和对照临床试验以及动物模型和细胞研究的结果。诱导型考克斯-2亚型在许多上皮细胞肿瘤中表达，包括结肠、肺、乳腺和胰腺的上皮细胞肿瘤。COX的产物是前列腺素H2（PGH 2），其经历细胞特异性酶促生物转化为前列腺素D2、E2、F2 a、I2和血栓烷A2。目前所有关于COX导致癌症的机制的研究都涉及这些酶促衍生的前列腺素的作用。然而，PGH 2也经历非酶促重排，产生高反应性的β-酮醛、左旋甘草定（LG）E2和D2，占总产物的20%。LG几乎瞬间反应形成伯胺的共价加合物。我们已经证明LGE 2与DNA中的脱氧胞苷碱基形成共价加合物。的加合物的稳定形式已被表征为吡咯，我们已经开发了一个高效液相色谱/串联质谱分析，采用稳定同位素稀释。这允许在两种癌细胞系中鉴定LG-DNA加合物的考克斯-2依赖性形成。这种庞大的COX衍生的共价加合物的形成表明，它可能会引起诱变和/或染色体不稳定性。除了DNA的这种修饰，LG与赖氨酸的ε-胺反应形成具有内酰胺结构的稳定加合物。考克斯-2衍生的LG-组蛋白加合物也已在人肺癌细胞系中鉴定，提高了驱动基因或肿瘤抑制基因转录的表观遗传改变的可能性。我们已经开发了一种有效的LG清除剂，乙基水杨胺，它可以防止DNA和组蛋白形成LG加合物，但不抑制COX。它还清除其他活性羰基，如丙二醛，这是环氧合酶和脂质过氧化的产物。 拟议的研究将采用乙基水杨胺作为工具，以解决DNA和组蛋白的修饰反应性羰基，如LG和丙二醛有助于考克斯-2依赖性癌症的生长的假设。将确定体内癌症中这些DNA和组蛋白修饰的存在以及乙基水杨胺对这些修饰的减少，以验证药理学干预的疗效。考克斯-2与仅在美国每年导致> 200，000人死亡的高度恶性癌症相关。因此，了解它如何有助于恶性肿瘤的演变是重要的。

 DESCRIPTION (provided by applicant): The hypothesis that highly reactive metabolites of the cyclooxygenase (COX) enzymes contribute to the COX-dependent growth of cancers will be examined in the proposed research. A concerted body of evidence indicates that the COXs contribute to the evolution of malignant neoplasms, and it includes findings from epidemiological studies and controlled clinical trials as well as investigations in animal models and cells. The inducible COX-2 isoform is expressed in many epithelial cell tumors, including those of the colon, lung, breast, and pancreas. The product of the COXs is prostaglandin H2 (PGH2) which undergoes cell specific enzymatic biotransformation to the prostaglandins D2, E2, F2a, I2, and thromboxane A2. All current research on the mechanism by which the COXs contribute to cancer addresses the actions of these enzymatically derived prostanoids. However, PGH2 also undergoes non-enzymatic rearrangement yielding the highly reactive -ketoaldehydes, levuglandins (LG) E2 and D2 as 20% of the total products. The LGs react almost instantaneously to form covalent adducts of primary amines. We have demonstrated that LGE2 forms a covalent adduct with deoxycytidine bases in DNA. The stable form of the adduct has been characterized as a pyrrole, and we have developed an HPLC/tandem mass spectrometric analysis for it, employing stable isotope dilution. This has permitted identification of COX-2 dependent formation of the LG-DNA adduct in two cancer cell lines. The formation of this bulky COX-derived covalent adduct suggests that it could engender mutagenesis and/or chromosomal instability. In addition to this modification of DNA, LG reacts with the epsilon-amine of lysine to form stable adducts with a lactam structure. COX-2 derived LG-histone adducts also have been identified in a human lung cancer cell line, raising the possibility of epigenetic alteration of the transcription of driver genes or tumor suppressor genes. We have developed a potent scavenger of LGs, ethyl-salicylamine, that prevents formation of LG adducts of DNA and histones but does not inhibit the COXs. It also scavenges other reactive carbonyls such as malondialdehyde which is a product of the cyclooxygenases and of lipid peroxidation. The proposed research will employ ethyl-salicylamine as a tool with which to address the hypothesis that modification of DNA and histone by reactive carbonyls such as LG and malondialdehyde contribute to the growth of COX-2 dependent cancers. The presence of these DNA and histone modifications in in vivo cancers as well as reduction of these modifications by ethyl-salicylamine will be determined to verify the efficacy of the pharmacologic intervention. COX-2 is associated with highly malignant cancers that result in >200,000 deaths annually in the US alone. Accordingly, understanding how it contributes to the evolution of malignancy is important.