Target discovery and combination therapy in KRAS mutant cancer

背景KRAS癌基因是肺的重要部分中的驱动突变， 胰腺癌和结肠直肠癌。近年来，靶向肿瘤的抑制剂取得了突破性进展， KRAS-G12 C突变体是几十年来第一个Ras靶向疗法。对于其他KRAS突变型肿瘤， 没有有效的靶向治疗。对KRAS抑制剂的耐药性迅速发展， 癌症患者，以及可以预防或延迟耐药性的药物组合， 延长患者反应的持久性。除了Ras，激酶抑制剂， Ras效应子途径已被开发用于靶向MAPK和PI 3 K途径。但这些 这些药物在临床试验中尚未显示出对KRAS突变型肿瘤的疗效。到目前为止， 靶向治疗主要集中在阻断OA，而较少关注 阻断KRAS突变细胞中的NOA。共同靶向NOA和OA可能会导致更好的 具有正交治疗模式的药物组合，改善的治疗窗，和 在KRAS突变型肿瘤中更持久的反应。目标. 1)使用功能基因组筛选 确定致癌基因成瘾（OA）和非致癌基因成瘾（NGC）引起的遗传依赖性。 成瘾（NOA）在KRAS突变癌细胞; 2）评估的机制，选择性 KRAS突变细胞的靶向治疗，以及导致耐药性的机制;以及3） 确定正交靶向KRAS中OA和NOA不同方面的药物组合 突变细胞以实现更好的治疗窗口。主要活动、重大业绩及 关键成果。1)基因筛选以识别KRAS突变癌症的功能漏洞 细胞使用合并的CRISPR/Cas9基因KO文库，我们已经进行了功能丧失性敲除。 在不同培养条件下对KRAS突变癌细胞进行筛选。我们已经确定了一 一个候选的、可药用的基因，对KRAS突变体的锚定非依赖性生长至关重要 细胞，我们目前正在研究这种遗传依赖性的机制， 探索这个目标的小分子抑制剂。2)Ras耐药机制 途径抑制剂。使用不同的KRAS突变癌细胞系，包括结肠直肠癌， 胰腺，肺和多发性骨髓瘤细胞，我们已经评估了遗传和 有助于对KRAS抑制剂和MEK耐药的转录后机制 抑制剂的我们目前正在阐明如何逆转这些耐药机制， 在临床前模型中改善治疗反应。3)新药鉴定 KRAS突变型癌症的联合治疗。使用高通量药物筛选和假设驱动 方法，我们正在评估涉及靶向Ras OA的抑制剂的药物组合， 阻断致癌Ras信号传导和靶向NOA通路的抑制剂， KRAS突变癌细胞中的应激。我们对比了KRAS中药物组合的毒性， 突变癌细胞与正常上皮细胞中的突变癌细胞对比以评估治疗窗， 我们使用小鼠异种移植和自体肿瘤模型来评估药物组合的 在体内对KRAS驱动的肿瘤的功效。

BACKGROUND. The KRAS oncogene is a driver mutation in a significant fraction of lung, pancreatic and colorectal cancers. Recently breakthroughs with inhibitors targeting the KRAS-G12C mutant is the first Ras targeted therapies in decades. For other KRAS mutant tumors, there are no effective targeted therapies. Resistance to KRAS inhibitors develop rapidly in cancer patients, and drug combination that can prevent or delay drug resistant are needed to extend the durability of response in patients. In addition to Ras, kinases inhibitors against Ras effector pathways have been developed to target the MAPK and PI3K pathways. However, these agents have not demonstrated efficacy in clinical trials against KRAS mutant tumors. Thus far, targeted therapies have primarily focused on blocking OA, and less attention have been devoted to blocking NOA in KRAS mutant cells. Co-targeting NOA and OA together could lead to better drug combinations with orthogonal therapeutic modalities, improved therapeutic window, and more durable response in KRAS mutant tumors. OBJECTIVES. 1) Using functional genomic screens to identify genetic dependencies resulting from oncogene addiction (OA) and non-oncogene addiction (NOA) in KRAS mutant cancer cells; 2) Evaluating the mechanism, selectivity of targeted therapies in KRAS mutant cells, and the mechanisms leading to drug resistance; and 3) Identify drug combinations that orthogonally target distinct aspects of OA and NOA in KRAS mutant cells to achieve better therapeutic window. MAJOR ACTIVITIES, SIGNIFICANT RESULTS AND KEY OUTCOMES. 1) Genetic screens to identify functional vulnerabilities in KRAS mutant cancer cells. Using pooled CRISPR/Cas9 gene KO libraries, we have carried out loss-of-function screens in KRAS mutant cancer cells under different culture conditions. We have identified a candidate, druggable gene that is critical for the anchorage-independent growth of KRAS mutant cells, and we are currently investigating the mechanism of this genetic dependency and exploring small molecule inhibitors of this target. 2) Mechanisms of drug resistant to Ras pathway inhibitors. Using different KRAS mutant cancer cell lines including colorectal, pancreatic, lung, and multiple myeloma cells, we have evaluated genetic and post-transcriptional mechanisms that contribute to resistance to KRAS inhibitors and MEK inhibitors. We are currently elucidating how these resistance mechanisms can be reversed to improve therapeutic response in pre-clinical models. 3) Identification of novel drug combinations for KRAS mutant cancer. Using high-throughput drug screen and hypothesis-driven approach, we are evaluating drug combinations that involve inhibitors targeting Ras OA to block oncogenic Ras signaling and inhibitors targeting NOA pathways to exacerbate oncogenic stress in KRAS mutant cancer cells. We contrast the toxicity of drug combinations in KRAS mutant cancer cells against that in normal epithelial cells to assess the therapeutic window, and we use mouse xenograft and autochthonous tumor models to evaluate the drug combination's efficacy against KRAS-driven tumors in vivo.