Elucidate the mechanism of autophagy in supporting Lkb1-deficient lung tumorigenesis and metastasis

摘要肺癌是癌症死亡的主要原因，其中非小细胞肺癌（NSCLC） 占这些案件的85%以上。KRAS是NSCLC中最常见的致癌驱动因子， 预后差，治疗选择有限。LKB 1是NSCLC中第三大最常见的突变基因。的 KRAS和LKB 1突变占NSCLC的约30%，具有增加的侵袭性， 转移的频率和对治疗剂的抗性。自噬降解蛋白质和细胞器， 使它们能够提供代谢底物，当细胞外营养物质有限时，这一功能至关重要。 虽然自噬在癌症中的作用已经被深入研究，但自噬在癌症中的确切作用， 特别是在体内，仍然是难以捉摸和有争议的。此外，靶向自噬治疗癌症通常 并没有对临床试验的进展做出重大贡献。因此，识别遗传脆弱性 因此，迫切需要对自噬抑制具有强烈敏感性的药物来改善自噬靶向- 治疗LKB 1通过激活AMP激活蛋白激酶（AMPK）调节能量稳态， 抑制分解代谢过程和上调合成代谢过程，以应对能源危机。基于早期 研究中，我们开始测试LKB 1的缺失促进癌细胞增殖，但也限制癌细胞增殖的假设。 对代谢应激的适应，这是一种可能因自噬丧失而进一步受损的性质。使用 在NSCLC的基因工程小鼠模型（GEMM）中，我们发现自噬抑制是综合性的， 在KrasG 12 D/+; Lkb 1-/-（KL）介导的肿瘤发生中是致死的;与具有完整自噬的KL肺肿瘤相反， 一个重要的自噬基因，Atg 7，显着损害肿瘤的发生和肿瘤的生长。这是 与野生型LKB 1（KrasG 12 D/+;p53-/-）肿瘤形成鲜明对比，野生型LKB 1（KrasG 12 D/+;p53-/-）肿瘤对必需的自噬不太敏感， 基因切除我们的体外研究进一步揭示了自噬调节KL所必需的脂质代谢， 癌细胞在营养饥饿中存活。这些观察结果表明，LKB 1突变易使KRAS- 驱动NSCLC的自噬抑制，LKB 1突变可作为预测性生物标志物进行探索， 精准肺癌治疗基于我们最近的发现，我们形成了我们的中心假设：自噬 通过维持Lkb 1缺陷型Kras驱动的肺肿瘤的代谢来补偿Lkb 1损失， 促进其转移。我们将通过以下具体目标来测试这一点：目标1。通过以下方式阐明机制： 其自噬调节体内脂质代谢和KL肿瘤发生。目标二。确定自噬如何 促进KL肿瘤转移。目标3：识别可能产生耐药性的代谢旁路 KL NSCLC中的自噬抑制。成功完成本提案将：（1）对角色产生新的见解 自噬在调节细胞代谢中支持KL肺肿瘤的发生和转移;（2）验证 自噬抑制是一种选择性的、强有力的治疗策略， 转移性KL NSCLC;和（3）揭示代谢旁路作为治疗抗性的潜在机制。

Abstract Lung cancer is the leading cause of cancer mortality, with non-small cell lung cancer (NSCLC) accounting for more than 85% of these cases. KRAS, the most common oncogenic driver in NSCLC, confers a poor prognosis with limited treatment options. LKB1 is the third most frequently mutated gene in NSCLC. The mutations in both KRAS and LKB1 account for about 30% of NSCLC, with increased aggressiveness, a high frequency of metastases and resistance to therapeutics. Autophagy degrades proteins and organelles and recycles them to provide metabolic substrates, a function that is critical when extracellular nutrients are limited. Although the role of autophagy in cancer has been intensively studied, the precise role of autophagy in cancer, especially in vivo, remains elusive and controversial. Moreover, targeting autophagy to treat cancer generally has not contributed significantly to the advancement of clinical trials. Therefore, identifying genetic vulnerability that renders strong sensitivity to autophagy inhibition is urgently needed to improve autophagy targeted- therapies. LKB1 regulates energy homeostasis by activating AMP-activated protein kinase (AMPK), which inhibits catabolic processes and upregulates anabolic processes, in response to energy crisis. Based on earlier studies, we began to test the hypothesis that loss of LKB1 promotes cancer cell proliferation but also restricts adaptation to metabolic stress, a property that may be further compromised by loss of autophagy. Using genetically engineered mouse models (GEMMs) of NSCLC, we found that autophagy inhibition was synthetically lethal in KrasG12D/+;Lkb1-/- (KL) mediated tumorigenesis; in contrast to KL lung tumors with intact autophagy, loss of an essential autophagy gene, Atg7, dramatically impaired both tumor initiation and tumor growth. This is in sharp contrast to wild-type LKB1 (KrasG12D/+;p53-/-) tumors that are much less sensitive to essential autophagy gene ablation. Our in vitro study further revealed that autophagy modulates lipid metabolism essential for KL cancer cells to survive nutrient starvation. These observations indicate that LKB1 mutations predispose KRAS- driven NSCLC to autophagy inhibition and that LKB1 mutations could be explored as a predictive biomarker for precision lung cancer therapy. Based on our recent findings, we form our central hypothesis: autophagy compensates for LKB1 loss by maintaining the metabolism of Lkb1-deficient Kras-driven lung tumors and promoting their metastasis. We will test this with following specific aims: Aim 1. Elucidate the mechanism by which autophagy regulates lipid metabolism and KL tumorigenesis in vivo. Aim 2. Determine how autophagy promotes KL tumor metastasis. Aim 3. Identify metabolic bypasses that potentially create resistance to autophagy inhibition in KL NSCLC. Successful completion of this proposal will: (1) yield new insights into the role of autophagy in modulating cellular metabolism in support of KL lung tumorigenesis and metastasis; (2) validate the novel concept that autophagy inhibition is a selective and powerful therapeutic strategy against primary and metastatic KL NSCLC; and (3) reveal metabolic bypass as a potential mechanism of therapy resistance.