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：
10360437
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 2024-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10360437
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 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.

概括由于缺乏小细胞肺癌（SCLC）的可药物致癌突变，并且未经批准的目标疗法，我们需要确定SCLC的新治疗方法。在此提案中，我们应用了整个基因组 CRISPR灭活屏幕以识别大致对SCLC必不可少的基因，或对关键遗传定义的基因子集。然后，我们确定药理学抑制是否对已识别命中的抑制也导致选择性杀害 SCLC细胞。我们应用体内系统，包括患者衍生的异种移植（PDX）和基因设计小鼠模型测试在这些遗传筛选中鉴定的主要靶标的药理抑制剂。 AIM 1将集中精力在我们已经使用CRISPR灭活屏幕上确定的途径上，对于RB/p53-Mutant至关重要 SCLC细胞。 NEDD8/RBX1途径的组成部分，控制Cullin活性和蛋白质泛素化是对于SCLC细胞必不可少的。使用MLN4924在患者中对NEDD8激活酶（NAE）的药理抑制作用衍生的异种移植（PDX）模型证实了SCLC细胞在该途径上的依赖。令人兴奋的是，我们确定了某些PDX模型中对MLN4924的特殊响应的示例。目标1：应用体内模型指导 NEDD8对SCLC子集的抑制最有可能响应。在这里，我们探讨了抑制的潜力 SCLC中的NEDD8途径。我们将将NAE抑制与顺铂 - 元素化学疗法和确定是否会产生持久的响应。一些SCLC PDX模型对NAE表现出精致的敏感性抑制;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出色的响应者。我们将确定NAE抑制是否应该是针对所有SCLC患者或关键子集，以及这种方法是否会增加对化学疗法的反应。 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 SCLC的试验。 AIM 2：应用CRISPR灭活筛选以确定SCLC的新治疗靶标。此目标将扩大我们的CRIS灭活屏幕，以包括带有关键驱动器突变的鼠标SCLC单元线超越RB/p53，包括PTEN，CREBBP和MYCL。我们的方法利用了人类 SCLC具有极高的突变负担，导致SCLC之间的广泛异质性驾驶员突变，具有定义驱动器改变的鼠标SCLC更加同质，增加了信号到噪声在识别漏洞中。我们将确定SCLC遗传定义的子集的依赖性，并将测试携带遗传改变的PDX和GEM模型中可识别的可药物靶标的抑制剂。我们利用可用的鼠标模型和派生的细胞系，以及一组遗传注释的PDX模型，在体内和ex Vivo中进行了研究，以识别和评估全新的靶向SCLC关键遗传子集的方法。