Small molecule probes of MYC stability and function intumorigenesis

MYC稳定性和肿瘤发生功能的小分子探针

• 批准号: 10361512
• 负责人: Sarki A. Abdulkadir
• 金额: $ 55.95万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-01 至 2026-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10361512
• 关键词: 3-Dimensional; ATAC-seq; Affect; Animal Model; Architecture; Binding; Biochemical; Biology; Cancer Model; Cell Proliferation; Cell Survival; Cell physiology; Cells; Cellular Stress Response; Cessation of life; Chemicals; Chromatin; Chromatin Structure; Combined Modality Therapy; Complement; Complex; Computer Assisted; Consensus; Coupled; DNA; DNA Binding; Dependence; Drug Kinetics; Epigenetic Process; Event; Family; Gene Expression; Genes; Genetic Transcription; Genome; Genomics; Helix-Turn-Helix Motifs; Human; In Vitro; Knowledge; MYC Family Protein; MYC gene; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of prostate; Mediating; Modeling; Oncogenes; Oncogenic; Oncoproteins; Output; Pathway interactions; Pharmaceutical Preparations; Phosphorylation; Play; Proteasome Inhibitor; Proteins; Publishing; Recurrence; Role; Series; Site; Stress; Testing; Therapeutic; Threonine; Toxicology; Transcriptional Activation; Tumorigenicity; Ubiquitin; Up-Regulation; Work; activating transcription factor; antitumor effect; base; biological adaptation to stress; c-myc Genes; cytokine; efficacy study; genetic manipulation; genome-wide; genomic locus; in silico; in vivo; inhibitor; knock-down; leukemia; member; multicatalytic endopeptidase complex; neoplastic cell; novel; novel strategies; overexpression; programs; recruit; response; screening; small molecule; small molecule inhibitor; transcription factor; transcriptome sequencing; tumor; tumor microenvironment; tumorigenesis

ABSTRACT MYC oncoproteins (including c-MYC, L-MYC and N-MYC) play critical roles in the initiation, progression and recurrence of many human malignancies. Extensive studies indicate that MYC is required to maintain tumor cell survival and proliferation. We have recently used a novel approach that combined computer-aided modeling with a rapid in vivo screen to develop a new series of direct small molecule inhibitors (MYCi’s) that show excellent selectivity, potency and tolerability in multiple MYC-driven cancer models. These compounds demonstrate a dual mechanism of action. First, direct binding of MYCi to MYC in the basic helix-loop-helix (bHLH) region disrupts complex formation with MYC which is required for MYC transcriptional activity. Secondly, binding of MYCi enhances MYC phosphorylation on threonine-58 (pT58) which promotes MYC degradation via the ubiquitin-proteasome pathway. However the key downstream effectors of these events and how they might impact cellular function are unknown. Reduction of MYC protein and enhanced pT58MYC may be expected to have profound effects on MYC family protein interactions with each other and with chromatin. In this regard, we have observed in preliminary studies that MYCi leads to selective loss of MYC at genomic loci enriched for master chromatin regulators (CTCF and FOX), suggesting disruption of the 3D architecture of the MYC-bound genome in response to MYCi. Additionally, unfolded MYC due to MYCi binding and/or enhanced MYC degradation may provoke a cellular stress response. Using unbiased ATAC-seq and RNA-seq approaches, we found that MYCi treatment activates the ATF4/CHOP stress response pathway. Importantly, activation of ATF4/CHOP by MYCi is an on-target, MYC-dependent effect. ATF4 mediates MYCi antitumor activity as ATF4 depletion partially ameliorates the antitumor effects of MYCi. Furthermore, we propose that MYCi-induced ATF4 cytokines modulate the tumor microenvironment. Activation of the ATF4 pathway by MYCi exposes potential therapeutic vulnerabilities for rational combination approaches, such as combination of MYCi with proteasome inhibitors that activates ATF4. Based on the preliminary findings, our central hypotheses is that MYCi inhibits MYC-dependent tumorigenesis by a dual-pronged mode of action. First, MYCi affects MYC family target gene expression by disrupting MYC/MAX interaction and by promoting MYC degradation. Secondly, binding of MYCi to MYC and/or MYC degradation activates an ATF4/CHOP stress response pathway that suppresses tumor cell viability. We propose the following specific aims to test these hypotheses: Aim 1). To investigate the mechanisms by which MYC inhibitor modulates MYC transcriptional activity and the epigenetic landscape. We will investigate the consequences of MYCi treatment on the recruitment of MYC, pT58MYC, and associated factors to chromatin; changes to 3D chromatin architecture; as well as the effects on MYC-driven transcriptional output in tumor cells vitro and in vivo. Aim 2). To define the mechanisms and functional consequences of ATF4/CHOP pathway activation by MYCi. We will determine mechanism of ARF4 upregulation by MYCi; define the role of MYCi- induced ATF4 in regulating target gene expression, cell viability and tumorigenicity; and assess strategies that exploit the consequences of ATF4 activation as a means of enhancing MYCi anti-tumor efficacy. These studies are significant as MYC is implicated in the majority of human cancers. The studies advance the use of MYCi as chemical probes to unmask distinct biology that complements the knowledge derived from genetic manipulations of MYC proteins. The findings will contribute to the efforts aimed at developing small molecule MYCi as potential therapeutics. Specifically, this work indicates that small-molecule MYC inhibitors have an additional anti-tumor effect due to the activation of the ATF4 pathway beyond the antitumor effects of suppressing MYC function. Finally, understanding this on-target ATF4 response provoked by small-molecule MYCi will provide rational strategies for combination therapy to enhance MYCi efficacy.

抽象的 MYC癌蛋白（包括C-MYC，L-MYC和N-MYC）在主动性，进展和 许多人类恶性肿瘤的复发。广泛的研究表明，MYC需要维持肿瘤细胞 生存和增殖。我们最近使用了一种新颖的方法，该方法将计算机辅助建模与 一个快速的体内屏幕，开发出新的直接小分子抑制剂（MYCI）的新系列，它们表现出色 多种MYC驱动的癌症模型中的选择性，效能和耐受性。这些化合物证明了 双重作用机理。首先，在基本的螺旋环螺旋（BHLH）区域中MYCI与MYC的直接结合 与MYC转录活动所必需的MYC破坏复合形成。其次，结合 Myci在三氨酸-58（PT58）上增强了MYC磷酸化，该磷酸化通过 泛素 - 蛋白酶体途径。但是，这些事件的关键下游影响以及它们如何 撞击细胞功能尚不清楚。可能预期降低MYC蛋白和增强PT58MYC 对MYC家族蛋白相互作用彼此和染色质具有深远的影响。在这方面，我们 在初步研究中观察到Myci会导致在基因组基因座的选择性损失富含的MYC 主染色质调节剂（CTCF和FOX），建议破坏MYC-BOND的3D体系结构 基因组响应myci。此外，由于Myci的绑定和/或增强了MYC，MYC展开了 降解可能会引起细胞应力反应。使用公正的ATAC-SEQ和RNA-SEQ方法，我们 发现MYCI处理激活了ATF4/CHOP应力响应途径。重要的是，激活 Myci的ATF4/CHOP是一个目标，依赖MYC的效果。 ATF4作为ATF4介导myci抗肿瘤活性 耗尽部分改善了Myci的抗肿瘤作用。此外，我们建议myci诱导的ATF4 细胞因子调节肿瘤微环境。 Myci激活ATF4途径可暴露潜力 理性组合方法的治疗脆弱性，例如Myci与蛋白酶体的组合 激活ATF4的抑制剂。基于初步发现，我们的中心假设是Myci抑制 MYC依赖性肿瘤发生通过双管造的作用方式。首先，Myci影响MYC家族目标基因 通过破坏MYC/MAX相互作用并促进MYC降解来表达。其次，myci的约束 到MYC和/或MYC降解激活ATF4/CHOP应力响应途径，抑制肿瘤细胞 生存能力。我们提出以下特定目的来检验这些假设：目标1）。调查机制 MYC抑制剂调节MYC转录活性和表观遗传景观。我们将调查 MYCI治疗对MYC，PT58MYC募集的后果以及染色质的相关因素； 更改3D染色质体系结构；以及对肿瘤细胞中MYC驱动的转录输出的影响 体内和体内。目标2）。定义ATF4/CHOP途径的机制和功能后果 Myci激活。我们将确定Myci上调ARF4的机制；定义myci-的作用 在控制靶基因表达，细胞活力和肿瘤性方面诱导了ATF4；以及评估策略 利用ATF4激活的后果，作为提高Myci抗肿瘤效率的一种手段。 这些研究很重要，因为大多数人类癌症都暗示了MYC。研究推进了 将myci用作化学问题来揭示独特的生物学，从而完成了从 MYC蛋白的遗传操作。这些发现将有助于旨在发展小的努力 分子myci作为潜在疗法。具体而言，这项工作表明小分子MYC抑制剂 由于ATF4途径的激活超出了抗肿瘤效应，具有额外的抗肿瘤效应 抑制MYC功能。最后，了解这种目标ATF4响应是由小分子引起的 Myci将提供合理疗法的合理策略，以提高Myci效率。