Molecular Mechanisms of a Male-Specific Positive Feedback Loop in Liver Cancer

肝细胞癌 (HCC) 是一种致命的癌症，影响的退伍军人比普通美国人还要多 人口。危险因素包括肝炎病毒感染、肥胖、衰老、酗酒/吸毒、接触有毒物质以及 遗传/表观遗传倾向，共同促进基因突变、细胞增殖和信号传导 通路功能障碍，导致癌症。值得注意的是，男性患 HCC 的几率是女性的 3 至 6 倍。 肝癌中男性占主导地位一直是医学领域的一个长期未解之谜。男性性激素 雄激素及其受体雄激素受体 (AR) 会加剧致癌过程，包括促进 肝炎病毒复制、癌基因和信号通路激活以及抑制肿瘤抑制因子 活动，从而在肝癌中发挥男性偏好。因此，抗雄激素引起了人们的极大兴趣 作为临床领域 HCC 治疗的疗法。重要的是，我们检测到了 本构活跃的 AR 变体，例如AR-V7，存在于选定的 HCC 标本中。由于这些 AR 变体非常重要 它们在前列腺癌转移进展中的作用，在 HCC 中的检测表明它们也可以发挥作用 肝癌中的致癌功能。此外，我们还发现了男性特有的正反馈循环， 男性特有的致癌基因 TSPY 相互作用并放大靶基因的 AR 和 AR-V7 反式激活， 分别以配体依赖和独立的方式包括其自身的基因，从而潜在地影响 抗雄激素疗法对 HCC 患者的有效性。 TSPY 是性腺母细胞瘤基因座的基因 (GBY) 位于 Y 染色体上。它是一种细胞周期调节剂，其功能障碍会促进细胞增殖和 肿瘤发生。 TSPY阳性HCC患者的生存率比阴性患者差，而核 AR/AR-V7 的位置表明预后不良。因此，了解这种雄性的作用机制 AR/AR-V7 和位于 Y 的 TSPY 基因之间在肝癌发生中的特异性正反馈环将 为诊断、预后和临床试验的转化应用提供科学基础 抗雄激素作为治疗 HCC 的药物。 该项目重点研究男性特异性正反馈回路的协同和致癌功能 AR/AR-V7 和 TSPY 在肝癌发生中的 3 个具体目标。一、TSPY、AR的表达模式 配对 HCC 肿瘤/非肿瘤 RNA 样本和病理标本中的 AR-V7 和 AR-V7 将被分析 证实这种男性特有的正反馈循环的存在，并评估它们的表达是否 模式，包括细胞学位置，可用作患者的诊断和预后特征。 其次，将研究Y定位的TSPY的AR和AR-V7转录激活机制 使用流体动力学转染策略的肝癌细胞和小鼠肝脏。三、TSPY的致癌功能， AR 和 AR-V7 在 HCC 中的作用将通过对整只小鼠肝脏的稳定流体动力学转染进行研究。他们的 将在正常条件和致瘤倾向下评估诱导肝肿瘤发生的能力。 关于它们在肝癌发生中的协同作用机制的全球观点将得到表征 通过转录组和顺反组分析差异基因表达、靶基因偏好和 与细胞生长、增殖相关的癌基因、肿瘤抑制因子和信号通路的失调 和肿瘤发生。我们将使用由此产生的动物模型来评估抗雄激素药物作为治疗方法 肝癌治疗。拟议的研究将为男性的分子机制提供重要的见解。 性激素受体 AR/AR-V7 及其与 Y 染色体癌基因 TSPY 的协同作用 肝癌发生中男性特异性正反馈回路。该项目的成功将为 为 AR/AR-V7 和基于 TSPY 的诊断、预后和开发的转化应用奠定了基础 HCC 临床管理的治疗策略；并将极大有利于人们的医疗保健服务 退伍军人易患或患有肝癌。

Hepatocellular carcinoma (HCC) is a deadly cancer that affects more Veterans than the general American population. Risk factors include hepatitis virus infection, obesity, aging, alcohol/drug abuses, toxic exposure, and genetic/epigenetic predisposition, which collectively promote genetic mutations, cell proliferation and signaling pathway dysfunctions, leading to cancer. Significantly, men are affected 3 to 6 times higher than women in HCC. The male predominance in HCC has been a long-standing enigma in the medical field. The male sex hormone androgen and its receptor, androgen receptor (AR), exacerbate the oncogenic processes, including promotion of hepatitis viral replication, activation of oncogenes and signaling pathways, and repressing tumor suppressor activities, thereby exerting male preference in liver cancer. Hence, there are significant interests in antiandrogens as therapeutics in treatments of HCC in the clinical field. Importantly, we have detected the expression of constitutively active AR variants, e.g. AR-V7, in selected HCC specimens. Since these AR variants play important roles in metastatic advances in prostate cancer, their detection in HCC suggests that they could also exert oncogenic functions in liver cancer. Furthermore, we have identified a male-specific positive feedback loop, in which a male-specific oncogene TSPY interacts and amplifies AR and AR-V7 transactivation of target genes, including its own gene, in ligand-dependent and independent manners respectively, thereby potentially affecting the effectiveness of antiandrogen therapeutics for HCC patients. TSPY is the gene for the gonadoblastoma locus (GBY) on the Y chromosome. It is a cell cycle regulator, dysfunction of which promotes cell proliferation and oncogenesis. TSPY-positive HCC patients have worse survival rate than the negative ones while nuclear locations of AR/AR-V7 signify poor prognosis. Hence, understanding the mechanisms of actions of this male- specific positive feedback loop between AR/AR-V7 and the Y-located TSPY gene in hepatocarcinogenesis will provide the scientific foundation for translational applications in diagnosis, prognosis and clinical trials of antiandrogens as therapeutics in treatments of HCC. The project focuses on the synergistic and oncogenic functions of the male-specific positive feedback loop of AR/AR-V7 and TSPY in hepatocarcinogenesis under 3 specific aims. First, the expression patterns of TSPY, AR and AR-V7 in paired HCC tumor/non-tumor RNA samples and pathological specimens will be analyzed to substantiate the existence of such male-specific positive feedback loop and to evaluate if their expression patterns, including cytological locations, can be used as diagnostic and prognostic signatures for the patients. Second, the mechanisms of AR and AR-V7 transcriptional activation of the Y-located TSPY will be studied in HCC cells and mouse livers using hydrodynamic transfection strategy. Third, the oncogenic functions of TSPY, AR, and AR-V7 in HCC will be studied by stable hydrodynamic transfection to the livers of whole mice. Their ability to induce hepatic oncogenesis will be evaluated under normal conditions and tumorigenic predisposition. The global views on the mechanisms of their synergistic actions in hepatocarcinogenesis will be characterized with transcriptome and cistrome analyses in terms of differential gene expression, target gene preferences, and dysregulation of oncogenes, tumor suppressors and signaling pathways associated with cell growth, proliferation and tumorigenesis. We will use the resulting animal models to evaluate antiandrogen drugs as therapeutics in HCC treatments. The proposed research will provide critical insights on the molecular mechanisms of the male sex hormone receptor AR/AR-V7 and synergistic actions with the Y chromosome-located oncogene TSPY in the male-specific positive feedback loop in hepatocarcinogenesis. Success of the project will provide the scientific basis for translational applications in development of AR/AR-V7 and TSPY-based diagnosis, prognosis and therapeutic strategies in clinical management of HCC; and will greatly benefit the health care delivery to the Veterans predisposed to or suffering from liver cancer.