A role for AKT2 in BRAFV600E melanoma initiation, progression, and response to therapy

AKT2 在 BRAFV600E 黑色素瘤发生、进展和治疗反应中的作用

基本信息

批准号：
9320005
负责人：
Siobhan K. Mcree
金额：
$ 3.58万
依托单位：
TUFTS UNIVERSITY BOSTON
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-07-16 至 2018-05-15
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9320005
关键词：
AKT Signaling Pathway AKT inhibition AKT1 gene AKT2 gene AKT3 gene Ablation Aggressive behavior Allografting Antibodies Automobile Driving BRAF gene Cell Line Cells Classification Clinical Trials Data Diagnostic Disease Disease Progression Dose-Limiting Drug Targeting Drug resistance Early Diagnosis Extravasation Family Future Gene Amplification Genes Genetic Genomics Goals Human In Vitro Injection of therapeutic agent Investigation MAP Kinase Signaling Pathways Malignant Neoplasms Mass Spectrum Analysis Mediating Mediator of activation protein Melanoma Cell Metabolism Metastatic Melanoma Mitogen-Activated Protein Kinases Morphology Mus Mutation Neoplasm Metastasis Null Lymphocytes Oncogenic Outcome PI3K/AKT Pathway interactions Pharmaceutical Preparations Phenotype Phosphoproteins Phosphotransferases Play Process Protein Isoforms Protein-Serine-Threonine Kinases Proto-Oncogene Proteins c-akt Recurrence Resistance Role Skin Cancer Specificity Testing Therapeutic Therapeutic Intervention Toxic effect Treatment Protocols Tumor Burden Ursidae Family Work cancer cell cell growth cell motility epithelial to mesenchymal transition experimental study improved in vivo inhibitor/antagonist kinase inhibitor knock-down melanocyte melanoma migration mouse model mutant novel novel therapeutic intervention novel therapeutics outcome forecast resistance mechanism response targeted treatment therapeutic candidate treatment response tumor

项目摘要

Project Summary Despite recent advances, melanoma remains the deadliest form of skin cancer, and new therapeutic strategies are urgently needed. The MAP kinase (MAPK) and PI3K/AKT pathways are central to both disease progression and chemoresistance in melanoma, with dual targeting approaches yielding promising results that would benefit from increased target specificity and selectivity. The AKT family of serine/threonine kinases comprises three highly homologous and functionally distinct isoforms (AKT1, AKT2, and AKT3) that play unique roles in melanoma but have not yet been leveraged for isoform-specific targeting. AKT2 is an attractive candidate for therapeutic intervention, given that AKT2 mutations and AKT2 gene amplification are known mechanisms of adaptive resistance to current targeted therapies such as the BRAF inhibitor (BRAFi) Vemurafenib. We hypothesize that AKT2 is involved in melanoma invasion and metastasis, and acts as a mediator of the BRAFi response. We further suggest that testing AKT2 isoform-specific targeting in combination with BRAF inhibitors is an anti-melanoma strategy that may avoid serious dose limiting toxicities commonly associated with pan-AKT inhibition. This work will establish the contribution of AKT2 to BRAF-mutant melanoma progression and metastasis, using both human melanoma cell lines and mouse models. In Aim1, we will investigate the effect of AKT2 knockdown on human melanoma cell migration and invasion, and whether this differs in the presence of the BRAFi Vemurafenib. Further, we will identify AKT2-specific substrates that influence the BRAFi response by immunoprecipitating phosphoproteins reacting with AKT phospho-substrate-specific antibodies in cells treated with Vemurafenib with or without AKT2-specific knockdown. Immunoprecipitates will be analyzed by mass spectrometry, prioritized and validated using AKT2 wild-type and knockdown cells to determine if manipulation of any substrate alters cell sensitivity to BRAF inhibition. In Aim 2, we will investigate the contribution of genetic AKT2 loss in a BRAF-mutant melanoma prone mouse model, and perform metastasis seeding experiments by intra-cardiac injection of AKT2 null cell lines, as well as drug studies in allograft tumors. We will investigate the effect of co-targeting BRAF and AKT2 with specific inhibitors, as a proof-of- principle for future studies. With AKT inhibitors in clinical trials for melanoma, our findings could yield novel therapeutic strategies and increase the efficiency of existing therapies to improve treatment options and outcome for this devastating disease.

项目概要尽管最近取得了进展，黑色素瘤仍然是最致命的皮肤癌形式，新的治疗方法迫切需要制定战略。 MAP 激酶 (MAPK) 和 PI3K/AKT 通路是这两种疾病的核心黑色素瘤的进展和化疗耐药性，双靶向方法产生了有希望的结果将受益于提高的目标特异性和选择性。丝氨酸/苏氨酸激酶 AKT 家族包含三种高度同源且功能不同的亚型（AKT1、AKT2 和 AKT3）在黑色素瘤中发挥独特作用，但尚未用于异构体特异性靶向。 AKT2 是一个有吸引力的鉴于 AKT2 突变和 AKT2 基因扩增是已知的，因此是治疗干预的候选者对当前靶向治疗（例如 BRAF 抑制剂（BRAFi））的适应性耐药机制维莫非尼。我们假设 AKT2 参与黑色素瘤的侵袭和转移，并充当 BRAFi 反应的中介者。我们进一步建议测试 AKT2 亚型特异性靶向与 BRAF 抑制剂联合使用是一种抗黑色素瘤策略，可以避免严重的剂量限制性毒性通常与泛 AKT 抑制相关。这项工作将确定 AKT2 对 BRAF 突变黑色素瘤进展的贡献以及转移，使用人类黑色素瘤细胞系和小鼠模型。在目标 1 中，我们将研究以下效果： AKT2 敲低对人黑色素瘤细胞迁移和侵袭的影响，以及这在存在 AKT2 的情况下是否有所不同 BRAFi 维莫非尼。此外，我们将确定影响 BRAFi 反应的 AKT2 特异性底物通过免疫沉淀磷蛋白与细胞中 AKT 磷酸底物特异性抗体反应用 Vemurafenib 治疗，伴或不伴 AKT2 特异性敲低。免疫沉淀物将通过以下方式进行分析质谱法，使用 AKT2 野生型和敲低细胞进行优先排序和验证，以确定是否任何底物的操作都会改变细胞对 BRAF 抑制的敏感性。在目标 2 中，我们将调查遗传性 AKT2 缺失对 BRAF 突变黑色素瘤易感小鼠模型的影响，并进行转移通过心脏内注射 AKT2 无效细胞系进行播种实验，以及同种异体移植中的药物研究肿瘤。我们将研究用特定抑制剂共同靶向 BRAF 和 AKT2 的效果，作为证据- 今后学习的原则。随着 AKT 抑制剂进入黑色素瘤临床试验，我们的发现可能会产生新的成果治疗策略并提高现有疗法的效率，以改善治疗选择和这种毁灭性疾病的结果。