Deciphering epigenetically-regulated pathways to improve targeted therapy for invasion and metastasis in head and neck cancer

破译表观遗传调控途径以改善头颈癌侵袭和转移的靶向治疗

基本信息

批准号：
10650527
负责人：
Robi D Mitra
金额：
$ 60.8万
依托单位：
WASHINGTON UNIVERSITY
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：
2023-03-15 至 2027-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10650527
关键词：
Anabolism Binding Bromodomains and extra-terminal domain inhibitor CRISPR interference Cancer Etiology Cell Line Cells Cholesterol Cholesterol Homeostasis Cisplatin Clinical Trials Data Disease Drug Kinetics Drug Targeting Enhancers Epigenetic Process Epithelium FDA approved Fatty Acids Gene Expression Gene Targeting Genes Genetic Transcription Goals Head and Neck Cancer Head and Neck Squamous Cell Carcinoma Human Hybrids In Vitro Invaded Lead Learning Length Longevity Malignant Neoplasms Mass Spectrum Analysis Mesenchymal Metabolic Pathway Metabolism Modeling Molecular Morbidity - disease rate Neoplasm Metastasis Oncology Operative Surgical Procedures Organoids Outcome Pathway interactions Patients Pharmaceutical Preparations Phenotype Pre-Clinical Model Process Prognosis Proteins Radiation Radiation therapy Recurrence Role Smoking History Technology Testing Treatment Failure adverse outcome attributable mortality candidate selection chemotherapy cholesterol biosynthesis drug development drug testing experimental study gene network gene synthesis genome-wide analysis improved in vivo in vivo Model inhibitor insight knock-down lymph nodes metabolomics mevalonate mortality neoplastic cell oxidation prevent protein biomarkers response single cell analysis single cell sequencing small molecule small molecule inhibitor targeted treatment therapeutic target transcriptome sequencing treatment response tumor tumor heterogeneity tumor xenograft

项目摘要

PROJECT SUMMARY Head and neck squamous cell carcinoma (HNSCC) is the sixth leading cause of cancer-related mortality, with most deaths attributable to metastasis and treatment failure. Unfortunately, our understanding of the pathways that underlie invasion and metastasis is incomplete, but some hints were uncovered by our recent single cell sequencing analyses which revealed that many HNSCC tumors contained cells that were neither fully epithelial nor fully mesenchymal, but were in a unique, hybrid epithelial-mesenchymal (hybrid-EM) state. The presence of hybrid-EM cells was more predictive of poor response than smoking history, and functional studies established hybrid-EM as a key driver of invasion and metastasis. Together, these observations establish hybrid-EM as a central pathway in HNSCC progression. However, hybrid-EM marker proteins are not druggable, so it remains challenging to therapeutically target this state. Cell states like hybrid-EM are regulated by a variety of mechanisms, but super-enhancers, in particular, have been identified as essential for maintaining cell identity. We therefore targeted super-enhancers in HNSCC using the BET inhibitor JQ1 and observed a reduction in HNSCC invasion of ~2-fold and a suppression of hybrid-EM. This result was encouraging, but effect size was modest due to the indirect and non-specific mode of action of JQ1. We hypothesized that if we could elucidate genes regulated by super-enhancers in HNSCC, we could identify molecular pathways that are more directly and specifically involved with invasion and metastasis and uncover more potent targets. Our initial experiments found that super-enhancers regulate cholesterol biosynthesis genes in hybrid-EM cells and that their knockdown (KD) robustly disrupts hybrid-EM phenotypes, with a >2000-fold and >8000-fold reduction in invasion upon KD of two mevalonate synthesis genes. Importantly, the protein products of many of the cholesterol genes we identified can be inhibited by commercially available small molecules. Indeed, we found that statins, which inhibit cholesterol biosynthesis, potently inhibit invasion (>10-fold reduction). These data provide strong support for our original hypothesis. We now propose to use CRISPRi to systematically investigate the role of cholesterol metabolism in invasion in HNSCC using sophisticated in vitro and in vivo models (Aim 1). In Aim 2, we will evaluate 9 small molecule inhibitors of cholesterol biosynthesis in cell lines, patient-derived organoids (PDOs), and in vivo pre-clinical models. Finally, in Aim 3, we will identify the full regulatory network of super-enhancers that control hybrid-EM, specifically focusing on Brd2-4 which are inhibited by JQ1. For these experiments, we will use single cell approaches to account for tumor heterogeneity and thereby uncover additional pathways beyond cholesterol biosynthesis that direct invasion and metastasis. Together, these experiments will provide critical insights into how super-enhancers regulate hybrid-EM, while exploring the possibility that by targeting the super-enhancer-regulated cholesterol biosynthesis pathway using existing FDA-approved drugs, we could circumvent a length drug development process and move quickly into human trials.

项目摘要头颈部鳞状细胞癌（HNSCC）是癌症相关死亡率的第六个主要原因，大多数死亡归因于转移和治疗失败。不幸的是，我们对路径的理解这是入侵和转移的基础是不完整的，但是我们最近的单个单元发现了一些提示测序分析表明，许多HNSCC肿瘤都包含既不完全上皮的细胞也不是完全间质的，但处于独特的杂化上皮 - 间质（杂化 - EM）状态。存在杂种 - EM细胞的预测比吸烟史更可预测反应，并且建立了功能研究杂种EM是入侵和转移的主要驱动力。这些观察结果共同将杂种EM作为一个 HNSCC进展中的中心途径。但是，混合-EM标记蛋白不可吸毒，因此仍然存在具有挑战性的治疗目标。诸如Hybrid-Em之类的细胞状态受各种调节机制，但尤其是超增强剂，已被确定为维持细胞身份必不可少的。因此，我们使用BET抑制剂JQ1瞄准了HNSCC中的超级增强剂，并观察到降低 HNSCC入侵约2倍，并抑制混合EM。这个结果令人鼓舞，但效果大小是由于JQ1的间接和非特异性作用模式，适度。我们假设，如果我们能阐明由HNSCC中超级增强剂调节的基因，我们可以鉴定出更直接的分子途径并特别参与入侵和转移，并发现更多有效的靶标。我们的最初实验发现超级增强剂调节杂交EM细胞中的胆固醇生物合成基因，并敲低（kd）强劲地破坏杂种 - EM表型，在KD上的入侵降低了2000倍和> 8000倍两个甲谷酸合成基因。重要的是，许多胆固醇基因的蛋白质产物我们可以通过市售的小分子抑制所识别。确实，我们发现他汀类药物抑制了胆固醇生物合成，有效抑制侵袭（降低> 10倍）。这些数据为我们的大力支持原始假设。现在，我们建议使用CRISPRI系统地研究胆固醇的作用使用复杂的体外和体内模型中HNSCC入侵的代谢（AIM 1）。在AIM 2中，我们将评估9个细胞系中胆固醇生物合成的小分子抑制剂，患者衍生的类器官（PDOS），和体内临床前模型。最后，在AIM 3中，我们将确定超级增强剂的完整监管网络该控制杂种 - 专门针对JQ1抑制的BRD2-4。对于这些实验，我们将使用单细胞方法来解释肿瘤异质性，从而发现其他途径除了直接侵袭和转移的胆固醇生物合成之外。这些实验将共同提供对超级增强剂如何调节混合动力EM的批判性见解，同时探索通过针对的可能性使用现有FDA批准的药物，超促脑调节的胆固醇生物合成途径，我们可以规避长期的药物开发过程，并迅速进入人类试验。