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

• 批准号: 10133464
• 负责人: Luke Lee Lairson
• 金额: $ 48.37万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2022-12-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10133464
• 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; Chemoresistance; Clinical Trials; Collaborations; Cultured Tumor Cells; Data; Development; Disease; Distant Metastasis; Dose; Drug Combinations; Drug Kinetics; Drug resistance; ERBB2 gene; Early Diagnosis; Epithelial; 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; Patient-derived xenograft models of breast cancer; 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; Tumor Biology; Tumor Subtype; Xenograft procedure; base; cancer cell; cancer drug resistance; cancer recurrence; cancer stem cell; cancer therapy; cell type; 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 derived xenograft model; patient population; physical property; pre-clinical; prevent; response; scaffold; screening; small hairpin RNA; small molecule; small molecule inhibitor; small molecule libraries; standard care; stem cell model; stem cells; stem-like cell; success; targeted agent; targeted treatment; therapeutically effective; tool; trait; transcription factor; triple-negative invasive breast carcinoma; tumor; tumor initiation

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.

三阴性（ER-、PR-、HER2-）乳腺癌（TNBC）缺乏针对性治疗，导致高发病率 死亡率。化疗耐药肿瘤的复发以及远处转移的形成 两者都是导致头 3-5 年内死亡的因素。我们和其他人已经证明 TNBC 正在丰富 具有癌症干细胞（CSC）和间充质特征，并且诱导上皮-间充质- 转变（EMT）促进耐药CSC的从头生成/扩增。确实，基因 由 EMT 诱导产生的表达特征在密蛋白低乳腺癌中丰富，这是一种特殊的乳腺癌。 内在亚型占所有 TNBC 的近一半。 我们最近报道叉头盒转录因子 FOXC2 的表达是被独特诱导的 EMT 后和 CSC 中，FOXC2 足以促进 TNBC 的 EMT、肿瘤发生和转移 细胞系和化疗耐药性。值得注意的是，shRNA 介导的 FOXC2 敲低足以 阻断 EMT 诱导、转移以及肿瘤发生和转移的 CSC 相关特性 化疗耐药。鉴于这些结果，我们推断小分子药物发现筛选 选择性靶向表达 FOXC2 的 CSC 将产生重要的化学制剂，这些化学制剂将显着 改善富含 CSC 的 TNBC 的治疗。 我们对大约 100,000 种药物进行了高通量小分子筛选，并获得了 确定了七种对表达 FOXC2 的 CSC 具有优先活性的候选分子。在这个 根据提议，我们打算在广泛的范围内全面表征和验证这些分子的体外活性 乳腺癌细胞系谱系对 CSC 的选择性、阻断诱导和逆转的能力 EMT 相关肿瘤细胞特性（包括迁移和侵袭）及其改变 CSC 特征的能力 （包括乳腺球形成、标记表达和集落形成）。此外，我们将阐明 通过使用光亲和试剂，然后进行质谱分析，确定这些试剂的相关蛋白质靶标，以及 还确定这些靶蛋白对 EMT/CSC 特性的功能作用。最后我们来优化一下 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研究 使用多个 TNBC 患者来源的异种移植物在小鼠中进行测试。 这项研究的结果将直接确定治疗复发性和转移性癌症的新药 TNBC，还将揭示 FOXC2 在重编程过程中调控的分子网络 肿瘤细胞走向CSC/转移状态。 Mani 博士和 Lairson 博士之间的密切合作， 我们广泛的初步数据证明，成功的可能性很高。此外，扩展的 该提案的范围从使用高通量筛选的药物发现到患者来源的 TNBC 测试 异种移植，表明我们致力于将新型癌症疗法快速推进临床试验。