Targeting dysregulated transcriptome as therapy for post-myeloproliferative neoplasm (MPN) sAML

靶向转录组失调作为骨髓增殖后肿瘤 (MPN) sAML 的治疗方法

基本信息

批准号：
10595080
负责人：
KAPIL BHALLA
金额：
$ 46.63万
依托单位：
UNIVERSITY OF TX MD ANDERSON CAN CTR
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-13 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10595080
关键词：
ATAC-seq Acetylation Apoptotic Attenuated BCL2 gene BCL2L1 gene Binding Binding Sites Bromodomain Bromodomains and extra-terminal domain inhibitor C-terminal CDK9 Protein Kinase Catalytic Domain Cell Culture Techniques Cell Line Cell Survival ChIP-seq Chromatin Chromatin Remodeling Factor Clinical Cyclin-Dependent Kinase Inhibitor Cyclin-Dependent Kinases DNA Polymerase II DNA Sequence Alteration DNMT3a Dependence Disease Progression EZH2 gene Elements Enhancers Enzymes Epigenetic Process Evaluation Exhibits Family member Genes Genetic Transcription Growth Hematopoietic stem cells Histones In Vitro Induction of Apoptosis JAK1 gene JAK2 gene Janus kinase Lysine MCL1 gene Mediating Mediator Messenger RNA Mutation Myelofibrosis Myeloproliferative disease NF-kappa B Neoplasm Metastasis Oncogenes Outcome PIM1 gene Patients Phenotype Phosphorylation Positive Transcriptional Elongation Factor B Predisposition Productivity Protac Protein Family Proteins RELA gene RNA RUNX1 gene Reader Refractory Repression Resistance SRSF2 gene STAT3 gene Secondary acute myeloid leukemia Serine Signal Transduction TP53 gene Techniques Testing Therapeutic Time Transcript Xenograft Model antagonist c-myc Genes calreticulin cell growth chemotherapy cyclin T1 epigenetic regulation epigenome genomic profiles improved in vivo inhibitor mimetics negative elongation factor novel pre-clinical preclinical efficacy promoter protein expression recruit resistance mechanism stem cells transcription factor transcriptome transcriptome sequencing

项目摘要

Myeloproliferative neoplasms with myelofibrosis (MPN-MF) exhibit constitutive activity of JAK-STAT signaling due to mutations in JAK2, c-MPL or calreticulin genes. Additional mutations in chromatin/transcriptional modifiers (epimutations) induce transformation to AML (sAML) in up to 20% of patients with MPN-MF. Lack of significant activity of the JAK1 & 2 inhibitor (JAKi) ruxolitinib and of AML chemotherapy highlights the need to develop and test novel agents and combinations that would improve clinical outcome in patients with post- MPN sAML. Genetic alterations and dysregulated epigenome produce the dysregulated transcriptome responsible for the transformed phenotype and therapy-refractoriness in post-MPN sAML blast progenitor cells (BPCs). This dysregulated transcriptome is dependent on ‘chromatin-reader’ BET (bromodomain and extra-terminal) proteins (BETPs), e.g., BRD4, and on its interactor pTEFb (positive transcription elongation factor b), both recruited to super-enhancers and promoters of actively-transcribed oncogenes. Cyclin dependent kinase 9 (CDK9), the catalytic subunit of pTEFb, phosphorylates RNA pol II (RNAP2), promoting RNAP2-mediated mRNA transcript elongation of oncogenes essential for growth and survival of post-MPN sAML BPCs. However, effects of BETP-CDK9 axis inhibition on active super-enhancers/enhancers and promoters with resulting impact on the dysregulated transcriptome and survival have not been elucidated in patient-derived (PD) sAML BPCs. Additionally, epigenetic mechanisms of resistance to CDK9 or BETP inhibitor (CDK9i or BETi) treatment and their therapeutic abrogation in post-MPN sAML BPCs need evaluation. Our preliminary studies demonstrate that CDK9i or BETP-antagonist (BETi and BETP- PROTACs) treatment induces apoptosis of post-MPN sAML BPCs, which is associated with repression of sAML-relevant oncogenes, e.g., c-MYC, STAT3/5, NFkB, Bcl-xL and MCL-1. We hypothesize that BETP- antagonist and CDK9i-based combinations will repress the dysregulated transcriptome and oncogenes, and with JAKi or BCL2/Bcl-xL inhibitor co-treatment, synergistically induce in vitro and in vivo lethality in PD, post- MPN sAML BPCs. Specific aims of these studies are: Aim 1: To elucidate the effects of BETP-PROTAC and CDK9i on active super-enhancers/enhancers (by ATAC-Seq and ChIP-Seq), mRNA transcriptome (by RNA-Seq) and on protein expressions (by CyTOF), as well as determine their pre-clinical efficacy against genetically-profiled, cultured cell lines and PD, post-MPN sAML BPCs. Aim 2: To determine lethal activity of BETP-PROTAC and CDK9i-based combinations against JAKi-sensitive and JAKi-persister/resistant sAML BPCs, utilizing in vitro cell cultures and in vivo xenograft models. Aim 3: To elucidate the dysregulated epigenome and transcriptome as well as susceptibility to BETP-PROTAC-based combinations in BETi- or CDK9i-persister/resistant post-MPN sAML BPCs.

骨髓增生性肿瘤，具有骨髓纤维化（MPN-MF）暴露的JAK-STAT信号的本构活性由于JAK2，C-MPL或钙网蛋白基因的突变。染色质/转录的其他突变在多达20％的MPN-MF患者中，修饰剂（Epimutation）会影响向AML（SAML）转化。缺乏 JAK1和2抑制剂（JAKI）鲁u列尼和AML化学疗法的显着活性突出了需要开发和测试新颖的药物和组合，可以改善后临床结果 MPN SAML。遗传改变和表观遗传组失调会导致转录组失调负责MPN后SAML BLAST祖细胞中转化的表型和治疗 - 反对性细胞（BPC）。该失调的转录组取决于“染色质阅读器”下注（溴化域和末端）蛋白质（BETP），例如BRD4及其相互作用的PTEFB（阳性转录伸长因子b），均被招募到主动转录的癌基因的超级增强剂和启动子。细胞周期蛋白依赖性激酶9（CDK9），PTEFB的催化亚基，磷酸化RNA POL II（RNAP2），促进 RNAP2介导的癌基因的mRNA转录伸长对MPN的生长和存活至关重要 SAML BPC。但是，Betp-CDK9轴抑制对主动超增强剂/增强剂的影响促进者对转录组失调和生存的影响尚未阐明患者来源（PD）SAML BPC。另外，对CDK9或BETP的耐药性的表观遗传机制 MPN SAML BPC的抑制剂（CDK9I或BETI）治疗及其治疗杂种评估。我们的初步研究表明，CDK9I或Betp-Antagonist（Beti和Betp- Protacs）治疗诱导MPN SAML BPC的凋亡，这与表达有关 SAML相关的癌基因，例如C-Myc，STAT3/5，NFKB，BCL-XL和MCL-1。我们假设betp- 基于拮抗剂和基于CDK9I的组合将反映失调的转录组和癌基因，以及与Jaki或Bcl2/Bcl-XL抑制剂共同处理，在PD中协同诱导体外和体内致死性 MPN SAML BPC。这些研究的具体目的是：目标1：阐明Betp-Protac的影响和CDK9I在主动超增强剂/增强剂（通过ATAC-SEQ和CHIP-SEQ），mRNA转录组（通过 RNA-seq）和蛋白质表达式（通过cytof），并确定其针对临床前的效率 MPN SAML BPC遗传培养的细胞系和PD。目标2：确定致命活动基于Betp-Protac和CDK9I对Jaki敏感和Jaki-Persister/抗性SAML的组合 BPC，使用体外细胞培养和体内特征模型。目标3：阐明失调表观基因组和转录组以及对基于Betp-Protac的组合的敏感性 CDK9I-Persister/抗性后MPN SAML BPC。