Kinome-guided Targeting of Cooperative Dependencies in BRAF and KRAS Mutated Colorectal Cancer

转移性结直肠癌（mCRC）是美国癌症相关死亡的第二大原因。大致 一半的mCRC携带BRAF或KRAS突变，与预后较差和治疗较少相关 选项.靶向含有BRAF和KRAS的信号级联的组分的临床试验结果 都没什么吸引力这些发现表明，mCRC不是由一个独特的途径驱动或支配的， 一个致癌基因。本提案的目的是检验一个假设，即一组关键的 上调的平行通路作为合作依赖性发挥作用， BRAF和KRAS突变的mCRC。实现这一目标需要新的资源和技术。 因此，拟议的研究将配对患者代表性标本，包括肿瘤样本和患者- 衍生的异种移植物（PDX）模型，由共同PI，Atreya博士建立，具有新的高通量激酶活性 地图（HT-KAM）平台开发的共同PI，博士Coppé。HT-KAM使用肽库作为组合 直接测量激酶活性并揭示激酶可操作漏洞的传感器 途径。目的1和2将关注BRAF（V600 E）突变的mCRC。目标1的起始材料是 BRAF突变的mCRC PDX肿瘤用相应患者在临床试验中接受的方案治疗， 或BRAF抑制剂与靶向正交作用模式的药剂组合，通过调查CRC细胞发现， 线路与HT-KAM的试点版本。目的1：研究BRAF的保守激酶依赖性通路 突变的PDX肿瘤激酶签名将通过HT-KAM的极大扩展版本建立， 使用PhosphoAtlas的蓝图计算集成到功能网络的层次结构中， Coppé博士制造的磷酸反应组这将揭示哪些途径和哪些特定的依赖性是 最保守的。目标2将测试新的组合策略来治疗BRAF突变的mCRC，从 初步研究和PDX肿瘤的激酶组谱。两到四种药物的战略可能性，针对不同的 将从候选目标中确定优先顺序。治疗假设将在细胞系中进行测试， 将进行协同分析。最有希望的组合将在BRAF突变的 mCRC PDX模型。最佳疗效的方案将在预期的翻译中进行优化 回到病人。目标3将关注KRAS（G12）突变的mCRC。初步数据预测， 驱动BRAF和KRAS突变的mCRC的能力将不同。KRAS（G12）突变的mCRC提供了 有机会直接评估手术标本，并开发相应的PDX模型。相同的 如针对BRAF突变的mCRC所描述的迭代工作流程将被应用于发现和靶向合作的 KRAS（G12）突变的mCRC的依赖性。该项目的长期目标是：1）提高 通过专门针对BRAF或KRAS mCRC的治疗策略实现患者生存;以及2）建立 一个可推广到任何肿瘤类型的变革性发现管道。

Metastatic colorectal cancer (mCRC) is the second leading cause of cancer-related death in the US. Roughly half of mCRCs harbor a mutation in BRAF or KRAS, associated with worse prognosis and fewer treatment options. Results of clinical trials targeting components of the signaling cascades containing BRAF and KRAS have been underwhelming. These findings signify that mCRC is not driven by a unique pathway or dominated by a single oncogene. The objective of this proposal is to test the hypothesis that a critical set of upregulated, parallel pathways function as cooperative dependencies, driving therapeutic resistance in BRAF and KRAS mutated mCRC. New resources and technologies are required to accomplish this objective. Thus, the proposed research will pair patient-representative specimens, including tumor samples and patient- derived xenograft (PDX) models established by co-PI, Dr. Atreya, with a novel high-throughput kinase-activity mapping (HT-KAM) platform developed by co-PI, Dr. Coppé. HT-KAM uses peptide libraries as combinatorial sensors to directly measure the activity of kinase enzymes and reveal actionable vulnerabilities of kinase pathways. Aims 1 and 2 will focus on BRAF(V600E) mutated mCRC. The starting materials for Aim 1 are BRAF mutated mCRC PDX tumors treated with regimens received by corresponding patients on clinical trials, or a BRAF inhibitor combined with agents targeting orthogonal modes of action, found by surveying CRC cell lines with a pilot version of HT-KAM. Aim 1 will identify the conserved kinase-dependent pathways of BRAF mutated PDX tumors. Kinase signatures will be established via a vastly expanded version of HT-KAM, and computationally integrated into hierarchies of functional networks using the PhosphoAtlas blueprint of the phospho-reactome built by Dr. Coppé. This will reveal which pathways and which specific dependencies are most conserved. Aim 2 will test new combinatorial strategies to treat BRAF mutated mCRC, identified from pilot studies and the kinome profiles of PDX tumors. Two- to four-drug strategic possibilities, directed at distinct pathways, will be prioritized from candidate targets. Treatment hypotheses will be tested in cell lines and synergy analysis will be performed. The most promising combinations will then be tested in BRAF mutated mCRC PDX models. The regimens with the greatest efficacy will be optimized in anticipation of translation back to patients. Aim 3 will focus on KRAS(G12) mutated mCRC. Preliminary data predicts that the functional susceptibilities driving BRAF and KRAS mutated mCRC will differ. KRAS(G12) mutated mCRC offers the opportunity to directly evaluate surgical specimens, and to develop corresponding PDX models. The same iterative workflow as described for BRAF mutated mCRC will be applied to discover and target the cooperative dependencies of KRAS(G12) mutated mCRC. The long-term goals of this project are to are: 1) to improve the survival of patients via therapeutic strategies specifically tailored to BRAF or KRAS mCRC; and 2) to establish a transformative discovery pipeline that is generalizable to any tumor type.