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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：
10641667
负责人：
David MacPherson
金额：
$ 50.63万
依托单位：
FRED HUTCHINSON CANCER 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 2024-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10641667
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 Epithelium 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 Proteins RBX1 gene Signal Transduction System TP53 gene Testing Therapeutic Tumor Volume Work cancer initiation cancer type chemotherapy driver mutation drug sensitivity druggable target efficacy testing exceptional responders gene function improved in vivo in vivo Model inhibitor knock-down lung cancer cell mouse model mutant neuroendocrine differentiation new therapeutic target novel novel strategies novel therapeutic intervention novel therapeutics overexpression patient derived xenograft model pharmacologic programs response screening small cell lung carcinoma 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.

总结由于小细胞肺癌（SCLC）缺乏可药物治疗的致癌突变，因此，我们需要确定SCLC的新治疗方法。在这个建议中，我们应用全基因组 CRISPR失活筛选以鉴定广泛地对SCLC或关键遗传定义的SCLC至关重要的基因。子集然后，我们确定对识别出的命中的药理学抑制是否也导致选择性杀伤。 SCLC细胞。我们应用体内系统，包括患者来源的异种移植物（PDX）和基因工程小鼠模型来测试这些遗传筛选中确定的关键靶点的药理学抑制剂。目标1将重点我们已经使用CRISPR失活筛选确定了Rb/p53突变所必需的途径， SCLC细胞。NEDD8/RBX1通路的组成部分，控制Cullin活性和蛋白质泛素化，对SCLC细胞至关重要。MLN 4924对患者NEDD 8活化酶（NAE）的药理学抑制作用衍生的异种移植（PDX）模型证实了SCLC细胞对该途径的依赖。令人兴奋的是，我们发现某些PDX模型中MLN 4924的异常反应示例。目的1：应用体内模型指导 NEDD8抑制对SCLC亚群最有可能有反应。在这里，我们探索抑制 SCLC中的NEDD8通路。我们将联合收割机NAE抑制与顺铂-依托泊苷化疗联合，确定是否产生持久的反应。一些SCLC PDX模型对NAE表现出极高的灵敏度抑制;我们将确定超级响应PDX模型的潜在机制和遗传特征，可以用来帮助预测特殊反应者。我们将确定NAE抑制是否应该针对所有SCLC患者，或关键子集，以及这种方法是否增加了对化疗的反应。 MLN4924已经在其他癌症类型的临床试验中进行了测试，这项工作可能会迅速导致临床在SCLC中进行试验。目的2：应用CRISPR失活筛选来确定SCLC的新治疗靶点。这一目标将扩展我们的CRISPR失活筛选，以包括具有关键驱动突变的小鼠SCLC细胞系除Rb/p53外，还包括Pten、Crebbp和Mycl。我们的方法利用了这样一个事实， SCLC具有极高的突变负担，导致具有特定突变的SCLC之间存在广泛的异质性。驱动突变，具有定义的驱动改变的小鼠SCLC更同质，增加信噪比来识别漏洞。我们将确定遗传定义的SCLC子集的依赖性，并将测试在PDX和GEM模型中识别的可药用靶点的抑制剂，其具有确定的遗传改变。我们利用可用的小鼠模型和衍生的细胞系，沿着一系列遗传注释的PDX模型，在体内和离体研究，以确定和评估靶向SCLC关键遗传子集的全新方法。