Role of UDP-Glucuronosyltransferases in UV-mediated melanomagenesis

描述(由申请人提供)：黑素细胞是保护皮肤免受紫外线辐射有害影响的关键。矛盾的是，黑素细胞是最致命的皮肤癌--黑色素瘤的前驱细胞。黑色素瘤是美国增长最快的肿瘤类型之一，在过去20年中，全球黑色素瘤病例数量翻了一番。黑色素瘤是一种极具侵袭性的肿瘤，对目前的治疗方法具有难以置信的抵抗力。如果黑色素瘤被及早发现，在肿瘤变得侵袭性之前，可以通过手术切除治愈。不幸的是，黑色素瘤的损害可能会在很长一段时间内无法识别或没有症状。因此，识别用于化学预防黑色素瘤进展的信息丰富的生物标志物和分子靶点对于控制这种致命疾病至关重要。这一建议基于一个新的观察结果，即三种UGT(UGT2B7、UGT2B10和UGT2B15)在人类黑素细胞中正常表达，并且它们的表达在黑色素瘤进展过程中丢失。由于UGT是细胞抵抗癌症形成的自然防御机制的重要组成部分，我们假设UGT表达的缺失是黑色素瘤进展所必需的。这项建议的重点是阐明在使用人类组织的黑色素瘤进展过程中，UGT是否以及何时丢失，以及这种丢失如何导致黑素细胞变成黑色素瘤。这里提供的证据表明，紫外线辐射，长期以来一直被认为是黑色素瘤的原因之一，下调了黑素细胞中UGT的表达。这一观察将被进一步表征以阐明UV-A和UV-B是否都触发了UGT表达的减少，以及这种减少是否与UGT总体活性的丧失相关。该提案还将调查UGT活性在黑色素瘤进展中的作用。由于UGT2B7、UGT2B10和UGT2B15都可以使生物活性脂质12-羟基二十碳四烯酸(12-HETE)失活，并且12-HETE水平升高与黑色素瘤的进展相关，我们推测人类黑色素瘤是通过UGT表达的缺失而由失控的12-HETE信号驱动的。为了测试这一点，UGT2B7、UGT2B10和UGT2B15将在转移性黑色素瘤细胞中稳定过表达，12-HETE水平将被监测并与细胞增殖、凋亡和侵袭性的变化相关。相反，shRNA将被用于敲除原代黑色素瘤细胞中UGT的表达，以通过检测它们的生长、避免凋亡的能力和增加侵袭力来观察这些细胞是否进展到更具侵袭性的黑色素瘤阶段。这些结果还将与12-HETE和12-HETE-葡萄糖醛酸苷水平相关。本文提出的研究结果将加强我们对黑色素瘤进展的了解，并识别新的信息丰富的生物标记物以及这种致命疾病的治疗干预的新靶点。 公共卫生相关性：该项目将研究人类黑色素瘤的进展是否由不受控制的生物活性脂质信号驱动。紫外线辐射在实现这种不受控制的生物活性脂质信号方面的作用也将被研究，UGT(负责关闭脂质信号的酶)的作用也将被研究。这些研究是第一次研究UGT在黑色素瘤进展中的作用。如果成功，这笔赠款将加强我们对黑色素瘤病因学的了解，提供黑色素瘤进展的信息生物标记物，并为这种致命疾病的治疗干预确定新的目标。

DESCRIPTION (provided by applicant): Melanocytes are essential to protecting the skin from the harmful effects of UV radiation. Paradoxically, melanocytes are the precursors of the most deadly form of skin cancer, melanoma. Melanoma is one of the fastest growing tumor types in the United States and the number of cases worldwide has doubled in the past 20 years. Melanoma is an extremely aggressive tumor and incredibly resistant to current therapies. If melanoma is detected early, before the tumor becomes invasive, it can be cured through surgical resection. Unfortunately, melanoma lesions can remain unidentifiable or asymptomatic for long periods of time. Therefore, the identification of informative biomarkers and molecular targets for the chemoprevention of melanoma progression is vital to the control of this deadly disease. This proposal builds on the novel observation that three UGTs (UGT2B7, UGT2B10 and UGT2B15) are normally expressed in human melanocytes and that their expression is lost during melanoma progression. Since the UGTs are a vital part of the cells natural defense mechanisms against cancer formation, we hypothesize that loss of UGT expression is required for melanoma progression. This proposal is focused on elucidating if and when UGTs are lost during melanoma progression in using human tissues and how that loss may cause melanocytes to become melanoma. Evidence presented here show that UV radiation, which has long been implicated in a causal role for melanoma, downregulates UGT expression in melanocytes. This observation will be further characterized to elucidate if both UV-A and UV-B triggers reduction of UGT expression and if that reduction correlates to a loss of overall UGT activity. This proposal will also investigate the role of UGT activity in melanoma progression. Since UGT2B7, UGT2B10 and UGT2B15 all can inactivate the bioactive lipid 12-hydroxyeicosatetraenoic acid (12-HETE) and elevated 12-HETE levels have been correlated to melanoma progression, we hypothesize that human melanoma is driven by uncontrolled 12-HETE signaling through loss of UGT expression. To test this, UGT2B7, UGT2B10 and UGT2B15 will be stably overexpressed in metastatic melanoma cells and the levels of 12-HETE will be monitored and correlated with alterations in cell proliferation, apoptosis and invasiveness Conversely, shRNA will be used to knockdown UGT expression in primary melanoma cells to see if those cells progress to more aggressive melanoma stages by examining their growth, ability to avoid apoptosis and increased invasiveness. These results will also be correlated to 12-HETE and 12-HETE-glucuronide levels. Results from the studies proposed here will enhance our understanding of melanoma progression and identify new informative biomarkers as well as novel targets for therapeutic intervention of this deadly disease. PUBLIC HEALTH RELEVANCE: This project will examine if human melanoma progression is driven by uncontrolled bioactive lipid signaling. The role of UV radiation in enabling this uncontrolled bioactive lipid signaling will also be examined as will the role of the UGTs (enzymes responsible for turning off lipid signaling). These studies are the first to examine UGTs in melanoma progression. If successful, this grant will enhance our understanding of melanoma etiology, provide informative biomarkers of melanoma progression and identify novel targets for therapeutic intervention of this deadly disease.