信号衔接蛋白p66Shc抑制肺癌转移机制的研究

Anoikis (detachment-induced apoptosis) resistance and aberrant autophagy (or macroautophagy), which are two main characteristics of lung cancer cells, promote the survival and give rise to metastasis. However, the mechanisms of anoikis and autophagy regulation in cancer cells which sustain oncogenic alterations remain largely elusive. We have previously shown that the adaptor protein p66Shc is a focal adhesion-associated protein and mediates anoikis through a RhoA-dependent mechanosensory test, thus plays an important role in preventing solid cancer metastasis via k-Ras-dependent control of proliferation and survival. In our on-going project of p66Shc function during epithelial-to-mesenchymal transition, EMT, we also identified the interaction between p66Shc and the leukemia-associated RhoGEF, LARG. This interaction may regulate the downstream activation of RhoA. Along with our preliminary data demonstrating that p66Shc significantly correlates with lung cancer patients' survival time, we propose here that reactivation of p66Shc and enhanced autophagy pathway might be a promising strategy to inhibit lung cancer metastasis. To test this hypothesis, we will use clinical lung cancer samples, lung cancer cell lines as well as mouse model to investigate the crosstalk between anoikis resistance and autophagy pathways mediated by p66Shc in vitro and in vivo. Based on our experimental data, the regulatory pathways of anoikis resistance and autophagic cell death will be elucidated and evaluated in clinical lung cancer samples as new candidates for metastasis treatment. Hopefully we may gain significant insight into the anoikis resistance and autophagy-related molecular events associated with lung cancer metastasis and add to means of assessing reactivation of p66Shc and enhanced autophagy for prevention of lung cancer metastasis. Also, a better understanding the role of p66Shc underlying anoikis and autophagic cell death including the determination of the upstream regulators and downstream effectors of p66Shc functional pathways may lead to the development of novel anti-tumor strategies for metastasis.

失巢凋亡抵抗和细胞自噬途径的变异是肺癌细胞获得转移的两个重要特性，其维持的分子机制不清楚。我们已证实信号衔接蛋白p66Shc通过激活RhoA，抑制k-Ras活性介导失巢凋亡，在实验动物水平抑制肺癌细胞转移；在研究p66Shc调控EMT过程中我们发现p66Shc与RhoGEF LARG相互作用调控细胞自噬途径，并且肺癌临床样品中p66Shc表达缺失显著影响患者存活，提示作为关键调控蛋白p66Shc介导细胞失巢凋亡和细胞自噬性死亡途径。本课题期望在现有的实验基础上来阐明肺癌细胞在转移过程中p66Shc协调失巢凋亡和细胞自噬性死亡新的调控和维持机制，依据实验结果描绘p66Shc调控失巢凋亡和细胞自噬性死亡的信号途径，并在临床肺癌组织样品中加以验证，同时探讨基于p66Shc生物学效应而进行的抑制肺癌细胞转移治疗的可行性。研究结果将有助于了解肺癌细胞发生发展的调控途径，丰富对肺癌转移分子机制的认识。

失巢凋亡抵抗（anoikis resistance）和细胞自噬(macroautophagy or autophagy)途径的变异是肺癌细胞获得转移的两个重要特性，其维持的分子机制不清楚。在过去的四年里，我们证实了介导失巢凋亡的信号衔接蛋白p66Shc通过p115-RhoGEF 和GEF-H1激活RhoA，而且细胞自噬的流量增加；与此相反，在不表达的p66Shc肺癌细胞中，容易产生失巢凋亡抵抗，并通过激活ZEB1介导EMT，细胞自噬流量减缓；结合我们过去的实验结果，我们认为作为抑制实体肿瘤转移的关键调控蛋白，p66Shc介导细胞失巢凋亡和细胞自噬性死亡途径；同时，在研究p66Shc的转录调控机制中，我们也发现来源于淋巴细胞的Aiolos等一系列转录因子促进肺癌细胞的转移，而Aiolos可以特异性地抑制包括p66Shc在内的抑制转移的基因的表达，增强了转移能力，故抑制Aiolos调控的途径成为一个潜在的药物作用靶点，比如用反应停特异地促进Aiolos的降解，增加p66Shc介导细胞失巢凋亡，从而抑制转移。此外，由于意识到细胞自噬对转移的调控作用，我们在细胞水平研究了转录调控因子NUPR1（nuclear protein 1）对细胞自噬的影响，基于获得的实验数据，得到了基金委的另一项面上项目的资助，初步的实验结果显示：对于依赖细胞自噬存活的肿瘤细胞而言，在自噬溶酶体阶段进行干预可能更不利于肿瘤细胞的存活，详细的分子机制还在研究之中，期望研究结果将丰富对肺癌转移分子机制的认识，为进一步了解肿瘤细胞发生发展的自噬调控途径提供帮助。

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