Preclinical characterization of EMT/CSC-specific small molecule inhibitors for TNBCs

TNBC 的 EMT/CSC 特异性小分子抑制剂的临床前表征

基本信息

批准号：
9898158
负责人：
Luke Lee Lairson
金额：
$ 53.53万
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：
2017
资助国家：
美国
起止时间：
2017-04-01 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9898158
关键词：
Animal Model Biological Assay Breast Cancer Cell Breast Cancer Model Breast Cancer Patient Breast Cancer cell line Cell Cycle Cell Death Cell Differentiation process Cell Line Cells Cellular biology Chemical Agents Chemicals Clinical Trials Collaborations Cultured Tumor Cells Data Development Disease Distant Metastasis Dose Drug Combinations Drug Kinetics Drug resistance ERBB2 gene Early Diagnosis Epithelial Epithelium Estrogen receptor positive Evaluation FOXC2 gene Gene Expression Profile Generations Human In Vitro Lead Light Link Malignant Epithelial Cell Malignant Neoplasms Mammary Neoplasms Mammospheres Mass Spectrum Analysis Mediating Mediator of activation protein Medical Mesenchymal Metastatic/Recurrent Methods Modeling Molecular Molecular Biology Molecular Target Mus Neoplasm Metastasis Patients Phenotype Physiological Preclinical Testing Prevention Property Proteins Publications Reagent Recurrence Refractory Relapse Reporting Reproducibility Research Institute Resistance Rodent Role Safety Signal Transduction Snails Survival Rate Systemic Therapy TGFB1 gene Testing Therapeutic Therapeutic Agents Treatment Efficacy Tumor Biology Tumor Subtype Xenograft Model Xenograft procedure base cancer cell cancer drug resistance cancer recurrence cancer stem cell cancer therapy cell type chemotherapy cytotoxic drug development drug discovery effective therapy epithelial to mesenchymal transition high throughput screening human model improved in vitro activity in vivo inhibitor/antagonist knock-down lead candidate malignant breast neoplasm migration mortality neoplastic cell novel novel therapeutics patient population physical property pre-clinical prevent response scaffold screening small hairpin RNA small molecule small molecule inhibitor small molecule libraries standard care stem cells stem-like cell success targeted agent targeted treatment tool trait transcription factor triple-negative invasive breast carcinoma tumor tumor initiation

项目摘要

Triple negative (ER-, PR-, HER2-) breast cancers (TNBCs) lack targeted treatments, resulting in a high mortality rate. Recurrence of chemotherapy-resistant tumors as well as formation of distant metastases are both contributors to mortality within the first 3-5 years. We and others have shown that TNBCs are enriched with cancer stem-cell (CSC)- and mesenchymal features, and that the induction of an epithelial-mesenchymal- transition (EMT) promotes the de novo generation/expansion of drug-resistant CSCs. Indeed, the gene expression signature generated by induction of EMT is enriched in claudin-low breast cancers, a particular intrinsic subtype comprising nearly half of all TNBCs. We recently reported that expression of the forkhead-box transcription factor FOXC2 is uniquely induced following EMT and in CSCs, and that FOXC2 is sufficient for EMT, tumor initiation, and metastasis of TNBC cell lines and chemotherapy-resistance. Remarkably, shRNA-mediated knockdown of FOXC2 is sufficient to block induction of EMT, metastasis as well as the CSC-associated properties of tumor initiation and chemotherapy-resistance. Given these results, we reasoned that a drug discovery screen for small molecules that selectively target FOXC2-expressing CSCs will yield important chemical agents that will dramatically improve treatment of CSC-enriched TNBCs. We have performed a high-throughput small molecule screen of approximately 100,000 agents, and have identified seven candidate molecules with preferential activity against FOXC2-expressing CSCs. In this proposal, we intend to fully characterize and validate the in vitro activity of these molecules across a wide spectrum of breast cancer cell lines for their selectivity for CSCs, their ability to block induction and reverse EMT-associated tumor cell properties (including migration and invasion), and their capacity to alter CSC traits (including mammosphere formation, marker expression and colony formation). In addition, we will elucidate the relevant protein targets for these agents, by use of photo-affinity reagents followed by mass spectrometry, and also determine the functional role of these target proteins for EMT/CSC properties. Finally, we will optimize candidate molecules for in vivo delivery and acceptable pharmacokinetics, and conduct extensive preclinical testing in mice using multiple TNBC patient-derived xenografts. The results of this study will directly permit the identification of novel drugs for treating recurrent and metastatic TNBCs, and will also shed light on the molecular networks regulated by FOXC2 during its reprogramming of tumor cells towards the CSC/metastatic state. The strong collaboration between Dr. Mani and Dr. Lairson, evidenced by our extensive preliminary data, indicates a high likelihood of success. Furthermore, the extended scope of this proposal, from drug discovery using high-throughput screens, to testing in patient-derived TNBC xenografts, indicates our commitment to rapidly moving novel cancer therapeutics into clinical trials.

三重阴性（ER-，Pr-，Her2-）乳腺癌（TNBC）缺乏靶向治疗，导致较高死亡率。化学疗法耐药性肿瘤的复发以及远处转移的形成是两者都在最初的3 - 5年内导致死亡率。我们和其他人表明TNBC富含具有癌症干细胞（CSC）和间质特征，并诱导上皮 - 间质 - 过渡（EMT）促进了抗药性CSC的从头产生/扩展。确实，基因通过诱导EMT产生的表达签名富含克劳丁 - 低乳腺癌，一种特定的固有的亚型包括所有TNBC的一半。我们最近报道说，叉子盒转录因子FOXC2的表达是唯一诱导的遵循EMT和CSC，并且FOXC2足以用于EMT，肿瘤的启动和TNBC转移细胞系和化学疗法抗性。值得注意的是，shRNA介导的FOXC2的敲低足以 EMT，转移以及CSC相关的肿瘤起始特性的阻滞诱导和化学疗法抗性。鉴于这些结果，我们认为小分子的药物发现筛选有选择地靶向表达FOXC2的CSC将产生重要的化学剂改善富含CSC的TNBC的处理。我们已经进行了大约100,000个代理的高通量小分子屏幕，并具有确定了七个针对表达FOXC2的CSC的优先活性的候选分子。在这个提案，我们打算充分表征和验证这些分子在范围内的体外活性乳腺癌细胞系的CSC选择性，它们可以阻止诱导和反向的能力 EMT相关的肿瘤细胞特性（包括迁移和侵袭）及其改变CSC特征的能力（包括哺乳动物形成，标记表达和菌落形成）。此外，我们将阐明这些药物的相关蛋白质靶标，通过使用光接触试剂，然后是质谱法，并且还可以确定这些靶蛋白对于EMT/CSC特性的功能作用。最后，我们将优化用于体内递送和可接受的药代动力学的候选分子，并进行广泛的临床前使用多个TNBC患者衍生异种移植物在小鼠中进行测试。这项研究的结果将直接允许鉴定用于治疗复发和转移性的新药物 TNBC，还将阐明FOXC2在其重新编程过程中调节的分子网络朝CSC/转移状态的肿瘤细胞。 Mani博士和Lairson博士之间的强有力合作我们广泛的初步数据证明了这一点，表明成功的可能性很大。此外，延伸该提案的范围，从使用高通量屏幕的药物发现到在患者衍生的TNBC中进行测试异种移植物表明我们致力于快速将新型癌症治疗剂转移到临床试验中。