Project 1: Targeting autophagy for the treatment of KRAS-mutant PDAC

项目1：摘要 自噬是癌细胞回收有缺陷的细胞器和大分子的自我降解过程。 作为一种营养来源来支持他们日益增长的新陈代谢需求。自噬被提升了，并且对 KRAS突变的胰腺导管腺癌(PDAC)的致瘤性生长，为 自噬抑制剂羟基氯喹治疗PDAC的临床评价令人失望的是，当使用 作为单一疗法和标准护理的结合，HCQ对PDAC的临床疗效有限。 我们最近确定，用关键的KRAS效应通路的抑制剂RAF-1治疗PDAC。 MEK-ERK丝裂原活化蛋白激酶级联，意外导致自噬进一步升高， 使PDAC严重依赖自噬，并对自噬抑制高度敏感。我们决定 抑制ERK会损害其他关键的代谢过程(糖酵解、线粒体功能)，从而导致 自噬的补偿性上调。我们的发现，连同基本上相同的结论 另一项独立的联合发表的研究，导致我们启动了两项临床试验，评估了两项批准的 MEK抑制剂(曲美替尼、比尼美替尼)联合盐酸氟喹诺酮治疗PDAC。因为我们最近的研究表明 ERK抑制剂在PDAC中将具有更好的活性，这促使我们启动了II期临床试验 ERK抑制剂LY3214996联合HCQ治疗转移性PDAC(目标1)。虽然很早 对这种组合的同情心护理利用的观察支持了这一点的显著临床影响 结合起来，我们的初步研究(目标2和3)支持我们的前提，即我们可以改进这种疗法。 目的2研究是基于我们应用的2500个基因的可药物基因组CRISPR-Cas9遗传缺失- 功能筛选以确定调节HCQ抗肿瘤活性的基因。已识别的命中结果要么增强了 或降低HCQ生长抑制活性代表HCQ抗性的候选组合或生物标志物， 分别进行了分析。反应的生物标记物然后可以应用于在Aim 1临床中收集的肿瘤活检组织 试行评估。Aim 3研究涉及使用525种肿瘤药物组的化学文库筛选的应用 目的：寻找能增强HCQ细胞毒作用的联合制剂。一起，从AIMS 2中产生的组合 然后，3项研究将被推进到目标4项研究，在那里我们将应用PDAC有机化合物或原位小鼠 肿瘤模型，以确定未来临床评估的组合。因为我们已经发现ERK MAPK 抑制导致肿瘤相关基因表达的变化，从而提高抗肿瘤免疫 响应，我们还将评估我们的组合对肿瘤细胞因子表达和对肿瘤- 相关的免疫细胞。总而言之，我们的研究将开发针对自噬的新的联合疗法 用于治疗KRAS突变的PDAC。

PROJECT 1: ABSTRACT Autophagy is a self-degradation process whereby cancer cells recycle defective organelles and macromolecules as a nutrient source to support their increased metabolic needs. Autophagy is elevated and essential for the tumorigenic growth of KRAS-mutant pancreatic ductal adenocarcinoma (PDAC), providing the rationale for clinical evaluation of the autophagy inhibitor hydroxychloroquine (HCQ) for PDAC. Disappointingly, when used as monotherapy in combination with standard of care, HCQ has shown limited to no clinical efficacy for PDAC. We recently determined that the treatment of PDAC with inhibitors of the key KRAS effector pathway, the RAF- MEK-ERK mitogenic activated protein kinase cascade, unexpectedly caused further elevation of autophagy, rendering PDAC acutely dependent on autophagy, and hypersensitive to autophagy inhibition. We determined that ERK inhibition impaired other critical metabolic processes (glycolysis, mitochondrial function) that then led to compensatory upregulation of autophagy. Our findings, together with essentially identical conclusions by another independent co-published study, has led to our initiation of two clinical trials evaluating two approved MEK inhibitors (trametinib, binimetinib) in combination with HCQ for PDAC. Since our recent studies suggest that ERK inhibitors will have superior activity in PDAC, this has prompted our initiation of a phase II clinical trial with the ERK inhibitor LY3214996 in combination with HCQ for metastatic PDAC (Aim 1). While early observations from compassionate care utilization of this combination support a significant clinical impact for this combination, our preliminary studies (Aims 2 and 3) support our premise that we can improve upon this therapy. Aim 2 studies are based on our application of a 2,500-gene druggable genome CRISPR-Cas9 genetic-loss-of- function screen to identify genes that modulate HCQ anti-tumor activity. The identified hits that either enhance or reduce HCQ growth inhibitor activity represent candidate combinations or biomarkers for HCQ resistance, respectively. The biomarkers for response can then be applied to tumor biopsies collected in the Aim 1 clinical trial evaluation. Aim 3 studies involve our application of a chemical library screen using a 525-oncology drug set to identify combinations that enhance the cytotoxicity of HCQ. Together, combinations that arise from Aims 2 and 3 studies will then be advanced to Aim 4 studies, where we will apply PDAC organoid or orthotopic mouse tumor models to identify combinations for future clinical evaluation. Since we have found that ERK MAPK inhibition causes tumor-associated gene expression changes that can lead to an improved anti-tumor immune response, we will also evaluate the impact of our combinations on tumor cytokine expression and on tumor- associated immune cells. In summary, our studies will develop novel combination therapies to target autophagy for the treatment of KRAS-mutant PDAC.