Non-oncogene addiction in cancer cells

癌细胞中的非癌基因成瘾

• 批准号: 10702519
• 负责人: Ji Luo
• 金额: $ 47.35万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10702519
• 关键词: Antineoplastic Agents; Autophagocytosis; Biological Models; CHD1 gene; Cancer Biology; Categories; Cells; Chromosomal Stability; DNA Damage; Dependence; Development; Drug Targeting; Equilibrium; Evolution; Genes; Genetic; Genetic Screening; Goals; Growth; Immunologic Surveillance; Inflammatory; KRAS oncogenesis; KRAS2 gene; Knowledge; Ligase; Malignant Neoplasms; Mass Spectrum Analysis; Metabolic stress; Mitotic; Molecular; Mutate; Neoplasm Metastasis; Normal Cell; Oncogenes; Oncogenic; Outcome; Pathway interactions; Phenotype; Play; Pre-Clinical Model; Proteins; Publishing; RAF1 gene; RNA Interference; RNA Splicing; RNA interference screen; Role; Signal Transduction; Spliceosomes; Stress; Sumoylation Pathway; Therapeutic; Work; base; biological adaptation to stress; cancer cell; cancer genome; cancer therapy; combinatorial; effective therapy; experience; inhibitor; malignant state; mutant; novel therapeutics; oncogene addiction; proteotoxicity; small molecule; targeted treatment; tumor

BACKGROUND. We define NOA as increased dependency of cancer cells on stress-response pathways for survival. The malignant state of the cancer cell is associated with a unique set of oncogenic stress phenotypes involving DNA damage, metabolic stress, proteotoxic stress, inflammatory microenvironment, and immune surveillance. Oncogenic stress renders cancer cells more dependent on stress-response pathways for survival and consequently, more sensitive to the disruption of these stress-response pathways. In contrast, normal cells in the body do no experience oncogenic stress, therefore normal cells are much less sensitive to the perturbation of stress-response pathways. This fundamental difference in cancer vs. normal cell's dependency on stress-response pathways forms the theoretical basis of NOA. NOA is distinct from oncogene dependency because stress response pathway genes are rarely mutated in the cancer genome. NOA represents a broad category of synthetic lethal and collateral dependency mechanisms in cancer cells, and targeting NOA could offer orthogonal therapeutic approaches to current targeted treatment of tumors. OBJECTIVES: The objectives of this project are 1.) Identify NOA genes and genetic pathways in cancer cells using genetic screen and hypothesis-based approaches; 2) Investigate the molecular mechanism that underly a NOA phenomenon and understand its essentiality to the oncogenic state; and 3) Explore the therapeutic implication of NOA as potential cancer drug targets. We will carry out these objectives primarily using cancer cells harboring the KRAS oncogene as our model system as KRAS mutant tumors have generally lacked effective therapies. MAJOR ACTIVITIES, SIGNIFICANT RESULTS AND KEY OUTCOMES. 1.) NOA of KRAS mutant cells to the RNA splicing factor ERH. Through a synthetic lethal RNAi screen in KRAS mutant cells, we have identified the ERH gene as a synthetic lethal partner in KRAS mutant cells. ERH is an evolutionarily conserved protein with poorly understood function. Using mass-spectrometry, we identified ERH associates with the spliceosome protein SNRPD3 and is therefore a component of the RNA splicing machinery. We showed that ERH is required for the proper splicing and expression of a subset of mitotic genes including CENPE that are critical for maintaining chromosomal stability in KRAS mutant cells. This work defined a new function for the ERH protein uncovered a previously unknown NOA in KRAS mutant cells to the RNA splicing machinery. This work suggests that selective perturbation of RNA splicing to disrupt the balance of mitotic proteins could be a potential approach to target KRAS mutant cancer cells. This work has been completed and published. 2.) NOA of KRAS mutant cells to the SUMO pathway for transformed growth. Through a synthetic lethal RNAi screen in KRAS mutant cells, we have identified the SUMO pathway, particularly the SUMO E2 ligase UBE2, to play an important role in supporting the viability and transformation growth of KRAS mutant cancer cells. Using mass-spectrometry, we identified multiple proteins whose SUMOylation are disrupted in a KRAS-dependent manner. Using gene rescue approaches, we showed that several SUMO target proteins, including KAP1, CHD1 and EIF3L, are critical for the viability of KRAS mutant cells under anchorage independent conditions. This work identifies a new role of the SUMO pathway in KRAS-driven oncogenesis and suggests the SUMO pathway as a potential drug target for KRAS mutant tumors. This work has been published. We are currently exploring the effect of small-molecule SUMO inhibitors in preclinical models of KRAS mutant cancer. 3) NOA of KRAS mutant cells to autophagy. Through a combinatorial RNAi analysis of the gene dependency landscape in KRAS mutant cells, we identified critical OA and NOA that components in the Ras signaling network and in stress-response pathways, respectively. We found CRAF/RAF1 as the major onco-effector of mutant KRAS and the autophagy E1 ligase ATG7 as a major NOA. This work identifies the autophagy pathway as a co-target with CRAF as a rational combination for KRAS mutant cells. This work has been published. We are currently investigating the mechanism by which autophagy contributes to the growth and survival of KRAS mutant tumors.

背景资料。我们将NOA定义为癌细胞生存对应激反应途径的依赖性增加。癌细胞的恶性状态与一系列独特的致癌应激表型有关，包括DNA损伤、代谢应激、蛋白毒性应激、炎性微环境和免疫监视。致癌应激使癌细胞更加依赖应激反应通路生存，因此对这些应激反应通路的破坏更加敏感。相比之下，体内的正常细胞不经历致癌应激，因此正常细胞对应激反应通路的扰动不那么敏感。癌症和正常细胞对应激反应通路的依赖性的这种根本差异形成了NOA的理论基础。NOA不同于癌基因依赖，因为应激反应途径基因在癌症基因组中很少发生突变。NOA代表了癌细胞中合成的致死和侧枝依赖机制的广泛类别，靶向NOA可以为当前的肿瘤靶向治疗提供正交的治疗方法。目标：本项目的目标是1.)通过基因筛选和基于假设的方法确定癌细胞中的NOA基因和遗传途径；2)研究NOA现象背后的分子机制，了解其对肿瘤状态的重要性；以及3)探索NOA作为潜在的癌症药物靶点的治疗意义。我们将主要使用携带KRAS癌基因的癌细胞作为我们的模型系统来实现这些目标，因为KRAS突变肿瘤通常缺乏有效的治疗方法。重大活动、重大成果、关键成果。1)KRAS突变细胞对RNA剪接因子ERH的NOA作用。通过在KRAS突变细胞中进行合成致死RNAi筛选，我们已经确定erh基因是KRAS突变细胞中的合成致死伙伴。ERH是一种进化上保守的蛋白质，其功能知之甚少。利用质谱仪，我们确定erh与剪接体蛋白SNRPD3有关，因此是RNA剪接机制的一个组成部分。我们发现ERH是包括CENPE在内的一组有丝分裂基因的正确剪接和表达所必需的，CENPE对维持KRAS突变细胞的染色体稳定至关重要。这项工作为ERH蛋白定义了一种新的功能，该蛋白在KRAS突变细胞中发现了一个以前未知的NoA到RNA剪接机制。这项工作表明，选择性干扰RNA剪接来破坏有丝分裂蛋白的平衡可能是一种潜在的靶向KRAS突变癌细胞的方法。这部作品已经完成并出版。2.)NOA对KRAS突变细胞向相扑途径转化生长的影响。通过在KRAS突变细胞中合成致死RNAi筛选，我们已经确定了相扑途径，特别是相扑E2连接酶UBE2，在支持KRAS突变癌细胞的存活和转化生长中发挥重要作用。使用质谱仪，我们鉴定了多个蛋白质，它们的SUMO化以KRAS依赖的方式被破坏。利用基因拯救方法，我们证明了KAP1、CHD1和EIF3L等几种SUMO靶蛋白对KRAS突变细胞在锚定非依赖条件下的生存至关重要。这项工作确定了相扑通路在KRAS驱动的肿瘤发生中的新作用，并暗示相扑通路可能成为治疗KRAS突变肿瘤的潜在药物靶点。这部作品已经出版了。我们目前正在探索小分子相扑抑制剂在KRAS突变癌症临床前模型中的作用。3)KRAS突变细胞对自噬的NOA反应。通过对KRAS突变细胞中基因依赖格局的组合RNAi分析，我们分别确定了RAS信号网络和胁迫反应途径中关键的OA和NOA组分。我们发现CRAF/RAF1是突变的KRAS的主要癌效应因子，自噬的E1连接酶ATG7是主要的NOA。这项工作发现自噬途径是一个共同的靶点，CRAF是KRAS突变细胞的合理组合。这部作品已经出版了。我们目前正在研究自噬促进KRAS突变肿瘤生长和存活的机制。