Overcoming resistance to BRAF(V600E) targeted therapies in melanoma

描述（由申请人提供）： 由于针对BRAF（V600 E）突变和免疫检查点阻断的靶向治疗，晚期黑色素瘤的治疗正在经历一场革命。然而，缓解的幅度和持久性都远非最佳（PLX 4032的中位进展约为8个月）。该PPG采用系统，全面和跨学科的方法，通过包括药物和免疫疗法在内的组合方法来阐明和克服这种治疗耐药性。我们的团队由黑色素瘤生物学的领导者以及当前关键临床试验的主要研究者组成。我们的方法建立在通过共享资源核心（核心A）使用体外培养技术和N 0 D/SCID/IL 2 RY-/-小鼠中的高效异种移植物产生成对的、患者来源的敏感性和抗性黑素瘤标本和细胞系的基础上。这些标本和相应的患者将通过一组广泛的临床和分子标准进行注释，包括候选癌基因测序、基因组拷贝数和PLX 4720 ICSO。然后，相同的配对黑色素瘤标本将进行深度测序和全基因组测序。shRNA表型筛选（项目1），以鉴定与抗性相关的获得性突变和基因依赖性（合成致死性）。命中的确认/优先级排序将建立在所有3个项目的集体知识基础上。项目2将在基因定义的小鼠黑色素瘤模型中进行耐药机制和命中验证的平行分析，包括测试来自项目1和3的候选物，以及合并对BRAF抑制剂具有获得性耐药性的肿瘤。在小鼠中鉴定的耐药机制将用于在人类研究中对候选药物进行优先排序（项目1）。项目3将研究MITF通路，其在BRAF（V600 E）抑制剂药物后拮抗细胞凋亡-提供可药用的机会-并且还研究可能由MITF通过调节黑色素瘤抗原性来调节的免疫参与。将使用基因定义的小鼠仔细检查BRAF（V600 E）拮抗剂加免疫检查点阻断的联合治疗，并在患者中进行研究。由此产生的分子和功能数据集将通过布罗德研究所的专业生物信息学和生物统计学核心（核心B）进行整合，以优先考虑和验证来自每个平台的命中，告知耐药机制，提名敏感性/耐药生物标志物，并确定临床前/临床开发的可药物漏洞。这些广泛集成的项目和核心的预期产出无法在单独的举措中复制，但将在高度协作的多学科结构中发挥作用，以加速发现治愈方法。

DESCRIPTION (provided by applicant): Treatment of advanced melanoma is undergoing a revolution due to targeted therapeutics directed at BRAF(V600E) mutations and immune checkpoint blockade. However both the magnitude and durability of responses are very far from optimal (median progression on PLX4032 of ~8 months). This PPG takes a systematic, comprehensive, and interdisciplinary approach towards elucidating and overcoming this treatment resistance with combination approaches including drugs as well as immunotherapy. Our team consists of leaders in melanoma biology as well as principal investigators of key current clinical trials. Our approach builds on generating paired, patient-derived sensitive and resistant melanoma specimens and cell lines via a Shared Resource Core (Core A) using in vitro culture techniques and highly efficient xenografts in N0D/SCID/IL2RY-/- mice. These specimens, and corresponding patients, will be annotated by an extensive set of clinical and molecular criteria including candidate oncogene sequencing, genomic copy number, and PLX4720 ICSO. The same paired melanoma specimens will then undergo deep sequencing and full-genome. shRNA phenotypic screening (Project 1), to identify acquired mutations and gene dependencies associated with resistance (synthetic lethality). Validation/prioritization of hits will build on collective knowledge from ail 3 Projects. Project 2 will carry out parallel analyses of resistance mechanisms and hit validation in genetically defined mouse melanoma models, including the testing of candidates derived from Projects 1 & 3 and the incorporation of tumors with acquired resistance to BRAF inhibitors. Resistance mechanisms identified in mice will be utilized to prioritize among candidates among the human studies (Project 1). Project 3 will examine the MITF pathway which antagonizes apoptosis after BRAF(V600E) inhibitor drugs-offering drugable opportunities-and also study immune involvement which may be regulated by MITF via modulation of melanoma antigenicity. Combination therapy with BRAF(V600E) antagonists plus immune checkpoint blockade will be scrutinized using genetically defined mice and also studied in patients. The resulting molecular and functional datasets will be integrated via a specialized Bioinformatics and Biostatistics Core at the Broad Institute (Core B) to prioritize and validate hits from each platform, inform mechanisms of resistance, nominate biomarkers of sensitivity/resistance, and identify drug-able vulnerabilities for preclinical/clinical development. The predicted output of these extensively integrated Projects and Cores could not be replicated in separate initiatives, but will function within a highy collaborative, multi-disciplinary structure to accelerate the discovery of a cure.