Understanding and Overcoming Resistance to BRAF/MEK Kinase Inhibitors in Melanoma

了解并克服黑色素瘤对 BRAF/MEK 激酶抑制剂的耐药性

基本信息

批准号：
10307107
负责人：
Meenhard F Herlyn
金额：
$ 49.38万
依托单位：
UNIVERSITY OF PENNSYLVANIA
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：
2019-12-13 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10307107
关键词：
Affect BRAF gene Bar Codes Biological Biology CRISPR/Cas technology Categories Cell Line Cells Combined Modality Therapy Complement DNA Development Distant Drug resistance Epigenetic Process Evolution Gene Expression Genes Growth Human Image Immunotherapy In Vitro Individual Libraries MAP2K1 gene MAP3K1 gene MEKs Melanoma Cell Minor Modeling Molecular Molecular Target Mutation Nature Organ Pathway interactions Patients Pattern Pharmaceutical Preparations Pharmacotherapy Phenotype Population RNA RNA analysis Relapse Resistance Resistance Process Resistance development Signal Pathway Specimen System Testing Therapeutic Time Transitional Cell Neoplasm Treatment outcome Tumor Cell Line Validation base cancer cell cancer drug resistance cellular transduction genome-wide human model in vivo in vivo Model inhibitor inhibitor therapy kinase inhibitor melanoma mouse model mutant neoplastic cell novel novel strategies novel therapeutic intervention novel therapeutics patient derived xenograft model preclinical study rational design relapse patients repository resistance mechanism response single-cell RNA sequencing targeted treatment three dimensional cell culture transcriptomics treatment response treatment strategy tumor tumor heterogeneity

项目摘要

The vast majority (>80%) of patients with BRAFV600E/K melanomas initially respond to highly specific BRAFV600E/K inhibitors as monotherapy or BRAF/MEK inhibitor combination therapy, but at varying levels. Nearly all patients relapse after three months to two years, despite that all of their tumor cells are carrying the BRAF mutations. In this application, we focus on the biological dynamics of small populations within BRAFV600E/K melanomas that are a priori resistant or rapidly adapt upon treatment and drive eventual relapse. In the first aim we will follow the dynamics of mutant BRAFV600E/K melanoma cells treated with BRAF and MEK inhibitor combinations. We have identified a slow-cycling subpopulation that is dynamically changing as the cells survive the initial treatment, then persist and finally regrow with acquired resistance to a variety of drugs. We will follow the cells in in vitro and in vivo models of human melanoma using bar codes and single cell RNA analyses. These studies will not only determine whether minor subpopulation(s) evolve under drug treatment stochastically or hierarchically, but also will provide us with information on their pre-existence and/or whether malignant cell plasticity is the major reason for intra-tumor heterogeneity to drug responses. In the second aim, we are testing the hypothesis that intrinsically resistant subpopulations of cells present new therapeutic opportunities that, if targeted, will specifically inhibit the onset of resistance. Our preliminary studies using CRISPR/Cas9 have allowed us to functionally dissect the pathways that form these rare subpopulations and, separately, the pathways that allow those subpopulations to reprogram. We will validate hits in 3D culture systems and in in vivo mouse models. We expect that targeting separate aspects of the resistance process in a rationally designed way (in vitro and then in preclinical studies) will enable the reduction of resistance onset upon BRAF/MEK inhibitor therapy.

BRAFV600E/K黑色素瘤的绝大多数（> 80％）最初对特定的BRAFV600E/K抑制剂作为单药治疗或BRAF/MEK抑制剂组合疗法，但是在不同的水平。三个月至两年后，几乎所有患者都复发了，尽管它们的肿瘤细胞携带BRAF突变。在此应用程序中，我们专注于 BRAFV600E/K黑色素瘤中小种群的生物动力学是先验的耐药或迅速适应治疗并驱动最终复发。在第一个目标中，我们将遵循BRAF和MEK处理的突变体BRAFV600E/K黑色素瘤细胞的动力学抑制剂组合。我们已经确定了一个慢节奏亚群，该亚群是动态的随着细胞的生存而变化，然后持续存在并最终重生对多种药物的抗性。我们将跟随人类体外和体内模型的细胞黑色素瘤使用条形码和单细胞RNA分析。这些研究不仅会决定次要亚群是在药物治疗下以随机或等级的方式进化，但是还将为我们提供有关其先前和/或是否恶性细胞的信息可塑性是对药物反应的肿瘤内异质性的主要原因。在第二个目标中我们正在检验以下假设，即细胞内本质上具有抗性亚群如果目标有针对性，则可以特别抑制抗药性的发作。我们的使用CRISPR/CAS9的初步研究使我们能够在功能上剖析途径这形成了这些罕见的亚群，并分别是允许这些的途径子群以重新编程。我们将验证3D培养系统和体内小鼠的命中型号。我们希望以合理的方式针对抵抗过程的各个方面设计的方式（在体外，然后在临床前研究中）将使电阻降低 BRAF/MEK抑制剂疗法发作。