Identifying and understanding drivers of chemoresistance in small cell lung cancer

识别和了解小细胞肺癌化疗耐药的驱动因素

基本信息

批准号：
10753857
负责人：
David MacPherson
金额：
$ 54.57万
依托单位：
FRED HUTCHINSON CANCER CENTER
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-07-01 至 2028-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10753857
关键词：
Bromodeoxyuridine Cancer Patient Cancer cell line Cell Death Chemoresistance Chromatin Cisplatin Clinical Trials Clustered Regularly Interspaced Short Palindromic Repeats Complex DNA Damage Data Etoposide Family member Gene Targeting Genes Genetic Genetic Transcription Genomics Histone H3 Human In Situ Nick-End Labeling In Vitro Kinetics Knowledge Libraries MYCL1 gene MYCN gene Malignant neoplasm of lung Modeling Molecular Mus Mutation Neurosecretory Systems Parents Pathway interactions Patients Pharmaceutical Preparations Phosphorylation Process Proliferating Proteins Proteomics RNA analysis Refractory Relapse Resistance Role SAGA System Testing Tissues cancer type chemotherapy epigenetic silencing established cell line genetic manipulation histone modification in vivo insight lentivirally transduced lung cancer cell member mutant novel overexpression patient derived xenograft model patient response prevent response screening small cell lung carcinoma tissue culture transcriptome sequencing treatment response tumor growth

项目摘要

SUMMARY Small cell lung cancer (SCLC) is an aggressive and lethal neuroendocrine lung cancer type. Most patients initially respond to chemotherapy but relapse occurs within months and genetic alterations that drive chemoresistance are poorly understood. Beyond amplification of MYC family members and epigenetic silencing of SLFN11, the field has an extremely poor understanding of genes that promote SCLC chemoresistance. We developed a novel system in which we genetically alter highly chemosensitive patient derived xenograft (PDX) models of SCLC to identify perturbations that confer resistance to cisplatin/etoposide (CIS-ETO) in vivo. Lentiviral overexpression of either MYCN or MYCL caused complete switch to chemoresistance (Grunblatt et al, 2020). To systematically identify SCLC chemoresistance drivers, we expanded use of this PDX lentiviral transduction system to perform in vivo CRISPR inactivation screens. We identified sgRNAs targeting multiple components of the SAGA (Spt- Ada-Gcn5 acetyltransferase) chromatin modifying complex as screen hits and confirmed that deleting the SAGA member USP22, a deubiquitylase, indeed confers chemoresistance in two SCLC PDX models, while return of USP22 to a USP22-null SCLC PDX model re-sensitizes to chemotherapy. Our overarching hypothesis is that suppressing the expression of USP22 and SAGA complex members drives chemoresistance in SCLC, and that transcriptional changes caused by SAGA suppression are critical. Aim 1, we will interrogate how genetically perturbing multiple SAGA complex members, including USP22 and TADA1, in PDX models alters the in vivo response to chemotherapy. Aim 2 employs genomic and proteomic approaches to develop a deep molecular understanding of the USP22-regulated genes and pathways that contribute to chemotherapy response in SCLC and uses human patient data to prioritize key SAGA targets for functional study. Decades of studying chemotherapy response in SCLC cell lines grown in vitro have provided little insight into how chemoresistance emerges, suggesting that key aspects of this process are not recapitulated under tissue culture conditions. Our novel system prioritizes the study of chemoresistance using in vivo approaches with potential to provide foundational knowledge to help prevent chemoresistance or re-sensitize chemoresistant SCLC to chemotherapy.

概括小细胞肺癌（SCLC）是一种侵袭性和致命性的神经内分泌肺癌。大多数患者最初对化疗有反应，但几个月内就会复发，并且基因改变会导致化疗耐药人们了解甚少。除了 MYC 家族成员的扩增和 SLFN11 的表观遗传沉默之外，该领域对促进 SCLC 化疗耐药的基因知之甚少。我们开发了一本小说我们对 SCLC 的高度化学敏感性患者衍生异种移植 (PDX) 模型进行基因改造，以识别体内对顺铂/依托泊苷 (CIS-ETO) 产生耐药性的扰动。慢病毒过度表达 MYCN 或 MYCL 导致完全转变为化疗耐药（Grunblatt 等人，2020）。为了系统地为了识别 SCLC 化疗耐药驱动因素，我们扩展了这种 PDX 慢病毒转导系统的使用来执行体内 CRISPR 失活筛选。我们鉴定了针对 SAGA 多个成分的 sgRNA（Spt- Ada-Gcn5 乙酰转移酶）染色质修饰复合物作为屏幕点击并确认删除 SAGA USP22 成员（一种去泛素化酶）确实在两个 SCLC PDX 模型中赋予了化学抗性，同时返回 USP22 到 USP22 无效 SCLC PDX 模型对化疗重新敏感。我们的总体假设是抑制 USP22 和 SAGA 复合体成员的表达可驱动 SCLC 的化疗耐药性，并且 SAGA 抑制引起的转录变化至关重要。目标 1，我们将探究基因如何在 PDX 模型中扰乱多个 SAGA 复合体成员（包括 USP22 和 TADA1）会改变体内对化疗的反应。目标 2 采用基因组学和蛋白质组学方法来开发深层分子了解有助于 SCLC 化疗反应的 USP22 调节基因和通路并使用人类患者数据来优先考虑功能研究的关键 SAGA 目标。数十年的学习体外培养的 SCLC 细胞系的化疗反应对化疗耐药性如何产生影响知之甚少出现，表明该过程的关键方面在组织培养条件下并未重现。我们的新系统优先使用体内方法研究化学耐药性，有可能提供帮助预防化疗耐药或使化疗耐药 SCLC 对化疗重新敏感的基础知识。