Metastatic protein network in BRAFV600E positive human thyroid cancers

BRAFV600E 阳性人类甲状腺癌的转移蛋白网络

• 批准号: 8386065
• 负责人: Carmelo Nucera
• 金额: $ 26.49万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-06 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8386065
• 关键词: Adhesions; BRAF gene; Binding; Binding Sites; Biological; Biological Assay; Biological Markers; Cell Line; Cell Proliferation; Cells; ChIP-seq; Clinical; Clinical Trials; DNA Binding; Data; Development; Disease; Disease Progression; Drug resistance; ECM receptor; Early Diagnosis; Endocrine; Extracellular Matrix; Extracellular Matrix Proteins; Focal Adhesion Kinase 1; Fostering; Gene Expression; Genes; Genetic Transcription; HMGB1 Protein; Human; Immunohistochemistry; In Vitro; Innovative Therapy; Integrin Binding; Integrins; Investigation; Link; Liquid substance; MAPK3 gene; Malignant Neoplasms; Malignant neoplasm of thyroid; Mass Spectrum Analysis; Modeling; Molecular; Mutation; N-terminal; Neck; Neoplasm Metastasis; Nuclear; Nuclear Translocation; Oral Administration; Papillary; Paraffin; Pathologic; Pathway interactions; Patient Monitoring; Patients; Phenotype; Phosphorylation; Phosphorylation Site; Phosphotransferases; Point Mutation; Process; Proteins; Proteomics; Recombinants; Recurrence; Refractory; Regulation; Residual Tumors; Residual state; Resistance; Sampling; Screening procedure; Sequence Analysis; Serum Response Element; Signal Transduction; Thrombospondin 1; Thyroid Gland; Thyroid carcinoma; Tissue Microarray; Tissues; Transcriptional Regulation; anaplastic thyroid cancer; base; cancer cell; cell motility; effective therapy; feeding; inhibitor/antagonist; insight; integrin-linked kinase; knock-down; melanoma; metastatic process; migration; mortality; mouse model; mutant; neoplastic cell; new therapeutic target; novel; outcome forecast; overexpression; prognostic; promoter; protein expression; receptor; small hairpin RNA; therapeutic target; thyroid neoplasm; transcription factor; tumor

DESCRIPTION (provided by applicant): A subset of patients with thyroid cancer has neck recurrences and metastases, is refractory to current treatments, and dies of the disease. BRAFV600E, the most frequent genetic alteration in both papillary (PTC) and anaplastic thyroid cancers (ATC), is implicated in progression from PTC to ATC. Our Gene Set Enrichment Analysis revealed that 7 of 17 gene sets up-regulated in BRAFV600E PTC were enriched in pro-metastatic extracellular matrix (ECM) proteins (e.g. thrombospondin-1 (TSP-1)) and receptors (integrins), including the integrin-linked kinases (i.e. focal adhesion kinase (FAK)) and the transcription factor (TF) HMGB1, which has been implicated in melanoma progression and metastasis. Knockdown of either TSP-1 or BRAFV600E inhibits cell proliferation, adhesion, migration/invasion, and metastasis in BRAFV600E-positive PTC and ATC cells and knockdown of TSP-1 significantly decreases levels of phospho(p)-ERK1/2, pFAK, and integrins. The novel selective BRAFV600E inhibitor PLX4720 shrinks tumor size in an orthotopic mouse model of ATC. However, persistence of the residual tumor and resumption of TSP-1, pERK1/2, and pFAK protein expression after 3 weeks of treatment suggest that metastatic thyroid cancer cells acquire resistance to BRAFV600E inhibition by up-regulating these proteins. Our objective is to identify BRAFV600E-dependent and -independent biomarkers for aggressive PTC by determining the essential signaling networks triggered by metastatic ECM proteins in BRAFV600E PTC cells. Specific Aim 1: To identify and assess potential biomarkers for PLX4720-resistant PTC by defining the molecular cascades by which TSP-1 stimulates ERK1/2 and FAK phosphorylation in human BRAFV600E PTC following inhibition of BRAFV600E. Preliminary data suggest that TSP-1 is associated with neck recurrence in BRAFV600E PTC. We will determine whether TSP-1 is a valid prognostic biomarker for PTC aggressiveness and establish an immunohistochemistry-based screening process suitable for clinical trials. To identify a larger panel of potential new prognostic biomarkers in BRAFV600E positive PTC, we plan to identify TSP-1 interactors. We will assess the correlation of some of these with TSP-1 expression and with clinico-pathological features of BRAFV600E positive PTC. To obtain a better understanding of BRAFV600E function in PTC progression, we will apply an unbiased proteomic analysis combined with functional assays to determine ECM protein interactions and intracellular signaling cascades in BRAFV600E PTC cells. Specific Aim 2: To explore whether HMGB1 is a biomarker for PTC aggressiveness. We will correlate HMGB1 expression with TSP-1 expression and clinico-pathological features of BRAFV600E PTC and explore whether HMGB1 regulates expression of TSP-1 or of TFs crucial for TSP-1 expression. The results of this study are likely to (i) identify new prognostic biomarkers of aggressive BRAFV600E positive PTC that can be assayed in biological fluids and PTC tissues to help monitor patients undergoing targeted therapies and enable earlier diagnosis of these thyroid cancers, and (ii) foster development of innovative therapies for PTC refractory to current treatments. PUBLIC HEALTH RELEVANCE: A subset of patients with thyroid cancer has recurrences and metastases, is refractory to current treatments, and dies of the disease. The V600E mutation in the BRAF gene is the most frequent genetic alteration in both papillary (PTC) and anaplastic thyroid cancers (ATC), and is implicated in progression of these diseases, for which early diagnosis and new therapies are needed. Results from this study (which will integrate clinical-pathologic and translational basic endocrine investigations) will enable routine assessment of new prognostic metastatic biomarkers that could be assayed in biological fluids and thyroid carcinoma tissues, and foster the development of innovative therapies for metastatic thyroid cancers that are refractory to current treatments.

描述（由申请人提供）：一部分甲状腺癌患者出现颈部复发和转移，对当前治疗无效，并死于该疾病。 BRAFV600E 是乳头状 (PTC) 和未分化甲状腺癌 (ATC) 中最常见的基因改变，与从 PTC 到 ATC 的进展有关。我们的基因集富集分析显示，BRAFV600E PTC 中上调的 17 个基因集中有 7 个富含促转移细胞外基质 (ECM) 蛋白（例如血小板反应蛋白-1 (TSP-1)）和受体（整合素），包括整合素-连接激酶（即粘着斑激酶 (FAK)）和 转录因子 (TF) HMGB1，与黑色素瘤的进展和转移有关。 TSP-1 或 BRAFV600E 的敲低可抑制 BRAFV600E 阳性 PTC 和 ATC 细胞中的细胞增殖、粘附、迁移/侵袭和转移，并且 TSP-1 的敲低可显着降低磷酸 (p)-ERK1/2、pFAK 和整合素。新型选择性 BRAFV600E 抑制剂 PLX4720 可缩小 ATC 原位小鼠模型中的肿瘤大小。然而，治疗3周后残留肿瘤的持续存在以及TSP-1、pERK1/2和pFAK蛋白表达的恢复表明，转移性甲状腺癌细胞通过上调这些蛋白而获得对BRAFV600E抑制的抗性。我们的目标是通过确定 BRAFV600E PTC 细胞中转移性 ECM 蛋白触发的基本信号网络来识别侵袭性 PTC 的 BRAFV600E 依赖和独立生物标志物。具体目标 1：通过定义 TSP-1 在抑制 BRAFV600E 后刺激人 BRAFV600E PTC 中 ERK1/2 和 FAK 磷酸化的分子级联，识别和评估 PLX4720 耐药 PTC 的潜在生物标志物。初步数据表明，TSP-1 与 BRAFV600E PTC 的颈部复发相关。我们将确定 TSP-1 是否是 PTC 侵袭性的有效预后生物标志物，并建立适合临床试验的基于免疫组织化学的筛选流程。为了在 BRAFV600E 阳性 PTC 中识别更多潜在的新预后生物标志物，我们计划识别 TSP-1 相互作用因子。我们将评估其中一些与 TSP-1 表达以及 BRAFV600E 阳性 PTC 临床病理特征的相关性。为了更好地了解 BRAFV600E 在 PTC 进展中的功能，我们将应用无偏蛋白质组分析与功能测定相结合，以确定 BRAFV600E PTC 细胞中的 ECM 蛋白相互作用和细胞内信号级联。具体目标 2：探讨 HMGB1 是否是 PTC 侵袭性的生物标志物。我们将把 HMGB1 表达与 TSP-1 表达和 BRAFV600E PTC 的临床病理特征相关联，并探讨 HMGB1 是否调节 TSP-1 或对 TSP-1 表达至关重要的 TF 的表达。这项研究的结果可能 (i) 确定侵袭性 BRAFV600E 阳性 PTC 的新预后生物标志物，可以在生物体液和 PTC 组织中进行检测，以帮助监测接受靶向治疗的患者并实现这些甲状腺癌的早期诊断，以及 (ii)促进对当前治疗无效的 PTC 创新疗法的开发。 公共健康相关性：一部分甲状腺癌患者会出现复发和转移，对当前治疗无效，并死于该疾病。 BRAF 基因中的 V600E 突变是乳头状 (PTC) 和未分化甲状腺癌 (ATC) 中最常见的基因改变，并且与这些疾病的进展有关，需要早期诊断和新疗法。这项研究的结果（将整合临床病理学和转化基础内分泌研究）将能够对可在生物体液和甲状腺癌组织中进行检测的新的预后转移生物标志物进行常规评估，并促进转移性甲状腺癌创新疗法的开发，目前的治疗方法难以治愈。