BRAFV600E and VEGFR2 synergize to trigger papillary thyroid cancer progression

BRAFV600E 和 VEGFR2 协同触发甲状腺乳头状癌进展

• 批准号: 8728483
• 负责人: Carmelo Nucera
• 金额: $ 36.11万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8728483
• 关键词: Acetyl-CoA Carboxylase; Autocrine Communication; Automobile Driving; BAY 54-9085; Biological Markers; Cell Adhesion; Cell Survival; Cell-Cell Adhesion; Cells; ChIP-seq; Clinical Trials; Coculture Techniques; Collagen; Combined Modality Therapy; Data; Development; Dose; Drug resistance; Early Diagnosis; Endothelial Cells; Extracellular Matrix; Extracellular Matrix Proteins; Genes; Goals; Human; Implant; In Vitro; Integrins; Link; Luciferases; MAPK3 gene; Malignant neoplasm of lung; Metabolic; Metabolism; Modeling; Mus; Mutation; Neck; Neoplasm Metastasis; Oral; Papillary thyroid carcinoma; Paracrine Communication; Pathway interactions; Patients; Pericytes; Pharmaceutical Preparations; Pharmacotherapy; Phosphorylation; Phosphotransferases; Play; Process; Prognostic Marker; Property; Proteins; Proteomics; Recurrence; Refractory; Reporting; Research; Residual Tumors; Residual state; Resistance; Role; SCID Mice; Sampling; Signal Pathway; Signal Transduction; Specimen; Survival Rate; System; Testing; Thyroid Gland; Transcription factor genes; Treatment Efficacy; Up-Regulation; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2; anaplastic thyroid cancer; angiogenesis; autocrine; base; cancer cell; cancer therapy; cell motility; density; drug development; improved; in vitro Assay; inhibitor/antagonist; insight; lymph nodes; melanoma; migration; mortality; mouse model; novel; outcome forecast; paracrine; pre-clinical; protein expression; public health relevance; receptor; resistance mechanism; response; screening; small hairpin RNA; standard care; therapy resistant; tumor; tumor growth; tumor microenvironment; tumor progression

DESCRIPTION (provided by applicant): Papillary thyroid cancer (PTC) typically has a favorable prognosis; however, patients with PTC carrying the BRAFV600E mutation show resistance to radioiodine treatment and have high rates of metastases and low survival rates. Inhibitors of BRAFV600E, including vemurafenib, have been tested recently as treatments for BRAFV600E-melanomas, with very promising results. However, development of secondary tumor resistance to these compounds has been reported. Another potential player in PTC aggressiveness is vascular endothelial growth factor receptor 2 (VEGFR2), a crucial regulator of angiogenesis. Sorafenib, which targets VEGFR2, was only partially effective against metastatic PTC, suggesting secondary resistance mechanisms to this drug as well. Our preliminary results show that metastatic primary human BRAFV600E positive PTC cells in vitro were resistant to high doses of either vemurafenib or sorafenib alone, but combined treatment with both drugs decreased cell viability by greater than 90%. BRAFV600E up-regulates VEGFR2 and pERK1/2 in PTC cells, triggering recruitment of endothelial cells and pericytes. We also observed upregulation of ACAC¿ (metabolic gene) suggesting that BRAFV600E harnesses a metabolic response that promotes PTC metastasis, a novel mechanism. Our preliminary data also indicate that VEGFR2 is associated with neck recurrence in BRAFV600E PTC. Our objective is to determine how BRAFV600E and VEGFR2 fit into the autocrine and paracrine signaling pathways that control PTC cell progression. Our central hypothesis is that BRAFV600E and VEGFR2 pathways synergize in PTC cells, increasing secretion of extracellular matrix (ECM) proteins and leading to increased PTC cell adhesion, migration/invasion, and angiogenesis. We propose a secondary pathway in which the BRAFV600E/ERK1/2 signaling cascade triggers hyper-expression of VEGFR2 in PTC cells. The alternate VEGFR2 pathway synergizes with residual BRAFV600E activity in the presence of vemurafenib, conferring secondary drug resistance in PTC cells. We will test this by manipulating VEGFR2 and BRAFV600E function and assessing tumorogenic properties in our novel models: a 3D co-culture system that recapitulates the PTC microenvironment, an orthotopic mouse with primary human PTC cells with heterozygous BRAFWT/V600E, and ChIp-seq analysis. Aim 1: To elucidate the autocrine and paracrine pathways by which BRAFV600E and VEGFR2 promote PTC aggressiveness. These studies will delineate VEGFR2-interacting ECM proteins that enhance ERK1/2 phosphorylation in BRAFV600E PTC and provide new insight into vemurafenib and sorafenib secondary drug resistance. Aim 2: To assess whether VEGFR2 protein expression is a long-term prognostic biomarker for PTC aggressiveness in BRAFV600E PTC. Aim 3: To assess efficacy of anti-BRAFV600E and anti-VEGFR2 combined therapy in a pre-clinical mouse model of human heterozygous BRAFWT/V600E PTC. These proposed studies are highly likely to provide evidence supporting clinical trials of combined targeted therapy for BRAFV600E-PTC and identify novel targets for drug development and biomarkers for aggressive PTC.

描述（由申请人提供）：甲状腺乳头状癌（PTC）通常预后良好;然而，携带BRAFV 600 E突变的PTC患者对放射性碘治疗表现出耐药性，转移率高，生存率低。BRAFV 600 E的抑制剂，包括维罗非尼，最近已经被测试作为BRAFV 600 E-黑色素瘤的治疗，具有非常有希望的结果。然而，已经报道了继发性肿瘤对这些化合物的耐药性的发展。PTC侵袭性的另一个潜在参与者是血管内皮生长因子受体2（VEGFR 2），它是血管生成的关键调节因子。靶向VEGFR 2的索拉非尼对转移性PTC仅部分有效，这也表明了对该药物的继发性耐药机制。我们的初步结果表明，转移性原代人BRAFV 600 E阳性PTC细胞在体外对高剂量的单独的威罗菲尼或索拉菲尼具有抗性，但用两种药物联合治疗使细胞活力降低超过90%。BRAFV 600 E上调PTC细胞中的VEGFR 2和pERK 1/2，触发内皮细胞和周细胞的募集。我们还观察到ACAC？（代谢基因）的上调，表明BRAFV 600 E利用促进PTC转移的代谢反应，这是一种新的机制。我们的初步数据还表明VEGFR 2与BRAFV 600 E PTC的颈部复发相关。我们的目标是确定BRAFV 600 E和VEGFR 2如何适应控制PTC细胞进展的自分泌和旁分泌信号通路。我们的中心假设是BRAFV 600 E和VEGFR 2通路在PTC细胞中协同作用，增加细胞外基质（ECM）蛋白的分泌，并导致PTC细胞粘附、迁移/侵袭和血管生成增加。我们提出了一个次要途径，其中BRAFV 600 E/ERK 1/2信号级联触发PTC细胞中VEGFR 2的高表达。在存在维罗非尼的情况下，替代的VEGFR 2途径与残余的BRAFV 600 E活性协同作用，从而在PTC细胞中赋予继发性耐药性。我们将通过操纵VEGFR 2和BRAFV 600 E功能并评估我们新模型中的致瘤特性来测试这一点：重现PTC微环境的3D共培养系统，具有杂合BRAFWT/V600 E的原代人PTC细胞的原位小鼠，以及ChIp-seq分析。目的1：阐明BRAFV 600 E和VEGFR 2促进PTC侵袭性的自分泌和旁分泌途径。这些研究将描述增强BRAFV 600 E PTC中ERK 1/2磷酸化的VEGFR 2相互作用ECM蛋白，并为vemurafenib和sorafenib继发性耐药性提供新的见解。目的2：评估VEGFR 2蛋白表达是否是BRAFV 600 E PTC中PTC侵袭性的长期预后生物标志物。目的3：评估抗BRAFV 600 E和抗VEGFR 2组合疗法在人杂合BRAFWT/V600 E PTC的临床前小鼠模型中的功效。这些拟议的研究极有可能为BRAFV 600 E-PTC联合靶向治疗的临床试验提供支持证据，并确定药物开发的新靶点和侵袭性PTC的生物标志物。