Nrf2, autophagy, and arsenic carcinogenesis

 描述（由申请人提供）：长期暴露于无机砷是一个全球性的公共卫生问题，与肺癌、皮肤癌和膀胱癌的风险增加有关。在这些恶性肿瘤中，砷诱发的肺癌死亡率最高.尽管多年的研究和与其暴露相关的健康影响的严重性，但砷的确切致癌机制尚未完全阐明。我们以前曾报道过，环境相关的低剂量砷阻断自噬，导致Nrf 2的长期激活，Nrf 2是适应性抗氧化和促生存反应的主要协调者。具体而言，砷诱导的Nrf 2活化不是通过典型的活性氧（ROS）传感机制发生的，而是通过自噬依赖的非典型机制发生的。自噬是降解受损细胞器和蛋白质聚集体的大量降解途径。自噬阻断可导致有缺陷的线粒体的积累和过量的ROS产生，这导致DNA突变。众所周知，Nrf 2的激活可以保护细胞免受ROS应激，这可以使砷暴露的细胞存活并维持驱动恶性转化的基因突变。因此，我们推测砷诱导的自噬阻断导致的Nrf 2的长期激活对于砷介导的恶性转化是必不可少的。我们的假设是支持最近的几个出版物表明，在自噬缺陷的小鼠长期Nrf 2激活的致瘤作用被取消的同时Nrf 2基因敲除。此外，早期的研究发现，许多癌细胞中Nrf 2的组成性水平很高，这有利于它们的增殖和化疗抗性，这种效应被称为Nrf 2的“黑暗面”。 我们已经产生了大量的数据表明砷通过抑制自噬体-溶酶体融合步骤来阻断自噬。三个SNARE介导融合：自噬体外膜上的Stx 17通过SNAP 29与溶酶体膜上的VAMP 8相互作用。我们相信，这些蛋白质中的任何一种的基因消融都应该防止自噬体与溶酶体的融合，并且这种消融的效果应该模拟砷介导的p62依赖性Nrf 2上调的效果。为了更好地理解砷的致癌性，我们提出：目的1：阐明砷阻断自噬体-溶酶体融合的详细分子机制。 目的2：确定自噬失调和Nrf 2激活延长是否是恶性转化所必需的。目的3：测试细胞系的致瘤性并将其与Nrf 2激活延长相关联。 影响：详细和彻底了解的分子事件，导致长期的Nrf 2激活砷诱导的致癌作用，将被证明是非常有价值的预防和治疗策略的生成，以及在识别生物标志物，为人群的风险。

 DESCRIPTION (provided by applicant): Chronic exposure to inorganic arsenic is a worldwide public health problem that has been associated to increased risks of developing cancers of the lung, skin, and bladder. Among these malignancies, arsenic- induced lung cancer presents with the highest mortality rate. The precise carcinogenic mechanism of arsenic has not yet been fully elucidated despite many years of research and the severity of the health effects associated to its exposure. We have previously reported that environmentally relevant, low doses of arsenic block autophagy, which resulted in prolonged activation of Nrf2, the main orchestrator of the adaptive antioxidant and pro-survival response. Specifically, arsenic-induced Nrf2 activation does not occur through the canonical, reactive oxygen species (ROS)-sensing mechanism but through the autophagy-dependent, non- canonical mechanism. Autophagy is a bulk degradation pathway that degrades damaged organelles and protein aggregates. Autophagy blockage can result in accumulation of defective mitochondria and excessive ROS production, which cause DNA mutations. Nrf2 activation is well known for its cellular protection against ROS stress, which can enable arsenic-exposed cells to survive and sustain gene mutations that drive malignant transformation. Therefore, we hypothesize that prolonged activation of Nrf2 resulting from arsenic-induced autophagy blockage is essential for the arsenic-mediated malignant transformation. Our hypothesis is supported by several recent publications demonstrating that the tumorigenic effect of prolonged Nrf2 activation in autophagy-deficient mice was abolished by concurrent Nrf2 knockout. Moreover, earlier studies have found high constitutive levels of Nrf2 in many cancer cells, which favor their proliferation and chemo resistance, an effect known as the "dark side" of Nrf2. We have generated substantial amounts of data indicating that arsenic blocks autophagy by inhibiting the autophagosome-lysosome fusion step. Three SNAREs mediate the fusion: Stx17 on the outer membrane of the autophagosome interacts through SNAP29 with VAMP8 that resides on the lysosome membrane. We believe that genetic ablation of any of these proteins should prevent the fusion of the autophagosome with the lysosome, and the effects of this ablation should mimic the effects of arsenic- mediated p62-dependent Nrf2 up regulation. To better understand arsenic carcinogenicity, we propose: Aim 1: Elucidate the detailed molecular mechanism by which arsenic blocks autophagosome- lysosome fusion. Aim 2: Determine if autophagy dysregulation and prolonged Nrf2 activation are essential for malignant transformation. Aim 3: Test the tumorigenicity of cell lines and correlate it with prolonged Nrf2 activation. Impact: A detailed and thorough understanding of the molecular events leading to the prolonged Nrf2 activation in arsenic-induced carcinogenesis will prove extremely valuable in the generation of preventive and therapeutic strategies, as well as in the identification of biomarkers, for the populations at risk.