Employing CRISPR inactivation screening and in vivo models towards improving treatments for SCLC

利用 CRISPR 失活筛选和体内模型来改善 SCLC 的治疗

基本信息

批准号：
10092980
负责人：
David MacPherson
金额：
$ 51.66万
依托单位：
FRED HUTCHINSON CANCER RESEARCH CENTER
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-03-08 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10092980
关键词：
Biology Cancer Model Cancer Patient Cancer cell line Cell Death Cell Line Cisplatin Clinical Trials Clustered Regularly Interspaced Short Palindromic Repeats Cullin Proteins Dependence Enzyme Inhibition Enzymes Epithelial Essential Genes Etoposide Exhibits Gene Deletion Genes Genetic Genetic Models Genetic Screening Genetic Transcription Genetically Engineered Mouse Genotype Goals Heterogeneity Human Lead Link Lung MYCL1 gene Modeling Molecular Mus Mutate Mutation Neuroendocrine Tumors Noise Oncogenic Pathway interactions Patients Pharmacology Proteins RBX1 gene Signal Transduction System TP53 gene Testing Therapeutic Tumor Cell Line Tumor Volume Work base cancer type chemotherapy driver mutation drug sensitivity druggable target efficacy testing exceptional responders gene function improved in vivo in vivo Model inhibitor/antagonist knock-down lung cancer cell lung small cell carcinoma mouse model mutant neuroendocrine differentiation new therapeutic target novel novel strategies novel therapeutics overexpression patient derived xenograft model programs response screening small molecule inhibitor targeted treatment tumor whole genome

项目摘要

SUMMARY With a lack of druggable oncogenic mutations in small cell lung cancer (SCLC) and no approved targeted therapies, we need to identify new treatment approaches for SCLC. In this proposal, we apply whole genome CRISPR inactivation screens to identify genes that are essential for SCLC broadly, or for key genetically defined subsets. We then determine whether pharmacologic inhibition of identified hits also results in selective killing of SCLC cells. We apply in vivo systems including patient derived xenograft (PDX) and genetically engineered mouse models to test pharmacologic inhibitors of key targets identified in these genetic screens. Aim 1 will focus on a pathway that we already identified using CRISPR inactivation screens to be essential for Rb/p53-mutant SCLC cells. Components of a NEDD8/RBX1 pathway, controlling Cullin activity and protein ubiquitylation, were essential for SCLC cells. Pharmacologic inhibition of NEDD8 activating enzyme (NAE) using MLN4924 in patient derived xenograft (PDX) models confirmed reliance of SCLC cells on this pathway. Excitingly, we identified examples of exceptional responses to MLN4924 in some PDX models. Aim 1: Apply in vivo models to direct NEDD8 inhibition to subsets of SCLC most likely to respond. Here, we explore the potential for inhibiting the NEDD8 pathway in SCLC. We will combine NAE inhibition with cisplatin-etoposide chemotherapy and determine whether durable responses result. Some SCLC PDX models exhibited exquisite sensitivity to NAE inhibition; we will identify underlying mechanisms and genetic features of super-responding PDX models that could be used to help predict exceptional responders. We will determine whether NAE inhibition should be directed to all SCLC patients, or to key subsets, and whether this approach augments response to chemotherapy. MLN4924 is already being tested in clinical trials for other cancer types and this work could rapidly lead to clinical trials in SCLC. Aim 2: Apply CRISPR inactivation screens to identify new therapeutic targets for SCLC. This Aim will extend our CRISPR inactivation screens to include mouse SCLC cell lines with key driver mutations beyond Rb/p53, including Pten, Crebbp and Mycl. Our approach takes advantage of the fact that while human SCLC has an extremely high mutational burden, leading to extensive heterogeneity among SCLC with a given driver mutation, mouse SCLC with defined driver alterations are more homogenous, increasing signal to noise in identifying vulnerabilities. We will identify dependencies in genetically-defined subsets of SCLC and will test inhibitors of identified druggable targets in PDX and GEM models that harbor defined genetic alterations. We leverage available mouse models and derived cell lines, along with a bank of genetically annotated PDX models, studied in vivo and ex vivo, to identify and evaluate completely new ways to target key genetic subsets of SCLC.

概括小细胞肺癌 (SCLC) 缺乏可药物治疗的致癌突变，且没有获批的靶向药物疗法，我们需要找到 SCLC 的新治疗方法。在这个提案中，我们应用了全基因组 CRISPR 失活筛选可识别广泛的 SCLC 所必需的基因，或关键基因定义的基因子集。然后我们确定对已识别的命中的药物抑制是否也会导致选择性杀死小细胞肺癌细胞。我们应用体内系统，包括患者来源的异种移植 (PDX) 和基因工程小鼠模型来测试这些基因筛选中确定的关键目标的药理学抑制剂。目标1将聚焦我们已经使用 CRISPR 失活筛选确定了一条对于 Rb/p53 突变体至关重要的途径小细胞肺癌细胞。控制 Cullin 活性和蛋白质泛素化的 NEDD8/RBX1 通路的组成部分是对于 SCLC 细胞至关重要。使用 MLN4924 对患者的 NEDD8 激活酶 (NAE) 进行药理学抑制衍生异种移植（PDX）模型证实了 SCLC 细胞对该途径的依赖。令人兴奋的是，我们发现一些 PDX 模型中对 MLN4924 的异常响应的示例。目标1：应用体内模型直接 NEDD8 抑制最有可能对 SCLC 亚型产生反应。在这里，我们探索抑制的潜力 SCLC 中的 NEDD8 通路。我们将 NAE 抑制与顺铂依托泊苷化疗相结合，确定是否产生持久的反应。一些 SCLC PDX 模型对 NAE 表现出极高的敏感性抑制;我们将确定超级反应 PDX 模型的潜在机制和遗传特征，可用于帮助预测特殊响应者。我们将确定是否应该抑制 NAE 针对所有 SCLC 患者或关键亚群，以及这种方法是否增强对化疗的反应。 MLN4924 已经在其他癌症类型的临床试验中进行测试，这项工作可能会迅速导致临床试验 SCLC 试验。目标 2：应用 CRISPR 失活筛选来确定 SCLC 的新治疗靶点。该目标将扩展我们的 CRISPR 失活筛选，以包括具有关键驱动突变的小鼠 SCLC 细胞系超出 Rb/p53，包括 Pten、Crebbp 和 Mycl。我们的方法利用了这样一个事实：虽然人类 SCLC 具有极高的突变负担，导致特定 SCLC 之间存在广泛的异质性驱动突变，具有明确驱动改变的小鼠 SCLC 更加同质，信噪比增加在识别漏洞时。我们将确定 SCLC 基因定义子集中的依赖性，并测试 PDX 和 GEM 模型中已确定的可药物靶标的抑制剂，这些模型包含明确的遗传改变。我们利用可用的小鼠模型和衍生细胞系，以及一系列基因注释的 PDX 模型，体内和离体研究，以确定和评估针对 SCLC 关键遗传亚群的全新方法。