Understanding intrinsic resistance to direct KRAS inhibition in colorectal cancers

了解结直肠癌对直接 KRAS 抑制的内在抵抗

• 批准号: 10346971
• 负责人: John D Gordan
• 金额: $ 52.64万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10346971
• 关键词: AXIN2 gene; BRAF gene; CRISPR interference; CTNNB1 gene; Cancer Biology; Cancer Etiology; Cancer Model; Cell Culture Techniques; Cell Line; Cell Proliferation; Cells; Cessation of life; Cholangiocarcinoma; Clinical Data; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Colorectal; Colorectal Cancer; Complex; Data; Dependence; Development; Down-Regulation; Drug Combinations; Drug resistance; Engineering; Exposure to; Feedback; Gene Expression; Genes; Genetic Transcription; Genomic approach; Human; In Vitro; Individual; Inferior; KRAS oncogenesis; KRAS2 gene; LGR5 gene; Laboratories; Link; Malignant Neoplasms; Malignant neoplasm of lung; Maps; Mediator of activation protein; Mitogen-Activated Protein Kinases; Modeling; Mutation; Non-Small-Cell Lung Carcinoma; Oncogenes; Oncogenic; Organoids; Outcome; Output; PIK3CA gene; Pancreatic Ductal Adenocarcinoma; Pathway interactions; Patients; Pharmaceutical Preparations; Phosphotransferases; Porcupines; Proteomics; Recurrence; Refractory; Regimen; Regulation; Resistance; Role; Signal Transduction; Skin; Tankyrase; Testing; Tissues; Toxic effect; Translations; Tumor Cell Line; United States; WNT Signaling Pathway; Work; base; cancer cell; cancer genomics; colon cancer cell line; colon cancer patients; colorectal cancer treatment; functional genomics; genomic data; improved; in vivo; inhibitor; interest; knock-down; melanoma; metastatic colorectal; mutant; new technology; novel strategies; novel therapeutics; patient derived xenograft model; patient population; programs; resistance mechanism; response; small molecule; small molecule inhibitor; systemic toxicity; targeted agent; targeted treatment; tool; transcription factor; treatment strategy; tumor; tumor growth

PROJECT SUMMARY/ABSTRACT Metastatic colorectal cancers (CRC) are the second leading cause of cancer death in the US. While advances in targeted therapies have transformed the treatment of many cancers, CRC has proven largely refractory to this approach. Thus, while agents targeting BRAFV600E and the KRASG12C mutation have dramatically improved the treatment of lung cancer and melanoma, they have only shown limited impact in CRC. CTNNB1 transcription is upregulated in >75% of CRC via APC inactivation and other mutations. As CTNNB1 is a common mediator of drug resistance and has been shown to be sufficient to maintain CRC proliferation, we hypothesize that it is a key mediator of intrinsic resistance to KRAS inhibition in CRC. Although multiple agents target CTNNB1 regulation via the WNT pathway, these have proven too toxic for human use to date. Thus, we have used proteomics to map the signaling response to KRASG12C inhibition in CRC cell lines and kinome-wide knockdown to identify kinases whose suppression synergizes with KRASG12C inhibition. By integrating these two approaches, we were able to uncover several kinases that function as signaling links between KRAS and CTNNB1, and whose inhibition synergizes with direct KRAS inhibition to reduce CTNNB1 target gene expression. As KRASG12C inhibitors do not impact normal KRAS signaling, this exciting preliminary data suggests that we may be able to preferentially downregulate CTNNB1 in tumors without systemic toxicity. We will build on this key preliminary data in this project: In Aim 1, we will expand our analysis of the kinase response to KRASG12C inhibition to additional CRC cell lines and the assess the impact of key kinases on CTNNB1 transcription. In Aim 2, we will use CRISPR in patient-derived xenografts (PDX) to circumvent the limitations of available small molecules to validate the role of CTNNB1 in APC-mutant CRC PDX. We will further use CRISPR or small molecules (when available) to test kinases already found to modulate CTNNB1 or emerging from Aim 1 in CRC treatment models and to determine their role in in vivo CRC biology. Finally, in Aim 3 we will develop KRASG12C CRC organoid and cell line models with mutations in PIK3CA, a common CRC mutation that co-occurs with KRAS mutations and is likely to cause resistance to KRASG12C inhibitors, but for which there are no models currently available. These tools will allow us to stratify the impact of PIK3CA mutation on our current treatment strategies and to optimize a regimen engineered specifically for this combination of mutations. This rigorous study of KRAS-driven signaling in CRC leverages new small molecules and robust quantitative approaches to unmask links between KRAS and the mechanisms that support CRC after KRAS inhibition. Uncovering the basis of resistance to direct KRAS inhibition in CRC will yield rational combination strategies primed for translation into clinical trials.

项目总结/摘要 转移性结直肠癌（CRC）是美国癌症死亡的第二大原因。虽然进步 靶向治疗已经改变了许多癌症的治疗，CRC已被证明在很大程度上对此难治。 approach.因此，尽管靶向BRAFV 600 E和KRASG 12 C突变的药剂显著改善了BRAFV 600 E的表达，但其在治疗中的作用仍然是显著的。 尽管它们用于肺癌和黑色素瘤的治疗，但它们在CRC中仅显示出有限的影响。 在>75%的CRC中，通过APC失活和其他突变，CTNNB 1转录上调。由于CTNNB 1是 一种常见的耐药性介质，并已被证明足以维持CRC增殖，我们 假设它是CRC中对KRAS抑制的内在抗性的关键介质。虽然多个代理商 通过WNT途径靶向CTNNB 1调节，迄今为止这些已被证明对人类使用毒性太大。 因此，我们已经使用蛋白质组学来绘制CRC细胞系中对KRASG 12 C抑制的信号应答， 全激酶组敲低以鉴定其抑制与KRASG 12 C抑制协同的激酶。通过 结合这两种方法，我们能够发现几种作为信号联系的激酶 在KRAS和CTNNB 1之间，并且其抑制与直接KRAS抑制协同作用以减少CTNNB 1 靶基因表达。由于KRASG 12 C抑制剂不影响正常的KRAS信号传导，因此这一令人兴奋的初步研究 数据表明我们可能能够优先下调肿瘤中的CTNNB 1而没有全身毒性。 在本项目中，我们将以这些关键的初步数据为基础：在目标1中，我们将扩展我们对激酶的分析， KRASG 12 C抑制对其他CRC细胞系的反应，并评估关键激酶对 CTNNB 1转录。在目标2中，我们将在患者来源的异种移植物（PDX）中使用CRISPR来规避CRISPR。 可用小分子的局限性，以验证CTNNB 1在APC突变型CRC PDX中的作用。我们将进一步 使用CRISPR或小分子（如果可用）来测试已经发现调节CTNNB 1或新出现的激酶 从目标1中的CRC治疗模型，并确定其在体内CRC生物学中的作用。最后，在目标3中， 开发具有PIK 3CA突变的KRASG 12 C CRC类器官和细胞系模型，PIK 3CA是一种常见的CRC突变， 与KRAS突变共同发生，可能导致对KRASG 12 C抑制剂的耐药性，但 目前没有可用的模型。这些工具将使我们能够对PIK 3CA突变对我们目前的研究的影响进行分层。 治疗策略，并优化专门针对这种突变组合设计的方案。 这项对CRC中KRAS驱动的信号传导的严格研究利用了新的小分子和稳健的定量分析。 揭示KRAS与KRAS抑制后支持CRC的机制之间联系的方法。 揭示CRC中直接KRAS抑制耐药的基础将产生合理的组合策略 已经准备好进行临床试验了