Targeted Therapies in Melanoma

描述（由申请人提供）：该计划项目的长期目标是基于对与疾病相关的关键蛋白质的机制理解以及这些蛋白质的小分子抑制剂的开发，开发治疗黑色素瘤的新策略。工作组建议实现以下目标1.靶向致癌BRAF治疗黑色素瘤。致癌的BRAF突变体BRAFV 600 E已经进化为可以说是黑色素瘤中最重要的靶点。我们将使用几种方法来探索BRAF作为治疗靶点的用途：我们将：使用复杂的体外和原位体内模型，使用目前可用的对BRAF或MEK具有活性的MARK途径抑制剂（项目1）; ii.诱导针对BRAF的特异性细胞免疫，其导致小鼠黑素瘤模型中的肿瘤生长抑制（项目2）; iii.基于BRAF的晶体结构开发新一代BRAFV 600 E抑制剂（项目3）;和iv.开发新的有机抑制剂和新一代有机金属抑制剂，使用钌吡啶咔唑分子支架，以提高BRAFV 600 E的特异性和有效性（项目3和4）。我们期望通过这些生物学、免疫学、结构和化学研究的结合，开发出以BRAFV 600 E为分子靶点的黑色素瘤新疗法。2.用新一代抑制剂靶向PI 3激酶通路，并将其与BRAF抑制剂联合用于黑色素瘤治疗。基于项目1中的初步研究，我们假设PI 3a激酶通路对黑色素瘤细胞存活非常重要。项目3和4结合联合收割机结构和化学策略，以确定新一代的抑制剂PI 3 Ka/？同种型。抑制剂将开发基于上述BRAF激酶（项目4）的有机金属支架，并支持抑制剂筛选体外（项目4）和体内（项目1）和X射线晶体结构的PI 3 K？抑制剂复合物（项目3）。此外，我们将使用体外和体内模型探索新型Akt/PKB抑制剂，并研究它们如何与BRAF抑制剂协同作用（项目1和2）。3.目标GSK 3？用于诱导细胞凋亡。我们已经发现，针对GSK 3？与有机金属抑制剂（在项目4中开发的）一起使用对黑色素瘤细胞具有令人惊讶的促凋亡活性（项目1）。基于这些初步研究，我们现在将研究细胞凋亡诱导的机制，这可能是通过激活p53（项目1）。我们还将继续制备更有效和具体的有机金属GSK 3？项目4中的抑制剂用于治疗项目1中的黑素瘤细胞。大多数黑色素瘤在GSK 3？中没有突变，但p53也可以被HDM 2下调，HDM 2在黑色素瘤中经常过表达。总之，该计划项目成员之间的合作将联合收割机结合肿瘤生物学，肿瘤免疫学，生物化学，化学和结构生物学的专业知识，以产生独特的方法和试剂用于黑色素瘤治疗。

DESCRIPTION (provided by applicant): The long-term objective of this program project is to develop new strategies for the treatment of melanoma, based on a mechanistic understanding of key proteins that are associated with the disease and the development of small molecule inhibitors to these proteins. The group proposes to pursue the following goals 1. Target oncogenic BRAF for therapy of melanoma. The oncogenic BRAF mutant, BRAFV600E, has evolved as, arguably, the most important target in melanoma. We will use several approaches to explore the use of BRAF , as a target for therapy: We will: i. Use currently available inhibitors of the MARK pathways with activity for either BRAF or MEK using complex in vitro and orthotopic in vivo models (Project 1); ii. Induce specific cellular immunity against BRAF that leads to tumor growth inhibition in a melanoma model in mice (Project 2); iii. Develop a new generation of inhibitors for BRAFV600E based on the crystal structure of BRAF (Project 3); and iv. Develop new organic inhibitors and a new generation of organometallic inhibitors using a ruthenium pyridocarbazole molecular scaffold to BRAFV600E with increased specificity and efficacy (Projects 3 and 4). We expect the development of new therapies of melanoma that focus on BRAFV600E as a molecular target to result from these combined biological, immunological, structural and chemical studies. 2. Target the PI3 kinase pathway with a new generation of inhibitors and use these in combination with BRAF inhibitors for therapy of melanoma. Based on preliminary studies in project 1, we hypothesize that PI3a kinase pathways are highly important for melanoma cell survival. Projects 3 and 4 combine structural and chemical strategies to identify a novel generation of inhibitors to the PI3Ka/? isoforms. The inhibitors will be developed based on the organometallic scaffold described above for the BRAF kinases (Project 4), and supported by inhibitor screens in vitro (Project 4) and in vivo (Projects 1) and x-ray crystal structures of PI3K? inhibitor complexes (Project 3). In addition, we will explore novel Akt/PKB inhibitors using in vitro and in vivo models and investigate how they synergize with BRAF inhibitors (Projects 1 and 2). 3. Target GSK3? for induction of apoptosis. We have found that targeting GSK3? with organometallic inhibitors (developed in Project 4) has surprising apoptosis-inducing activities on melanoma cells (Project 1). Based on these preliminary studies we will now investigate the mechanisms of apoptosis induction, which may occur through activation of p53 (Project 1). We will also continue to prepare more potent and specific organometallic GSK3? inhibitors in project 4 for treatment of melanoma cells in project 1. Most melanomas are not mutated in GSK3?, but p53 can also be downregulated by HDM2, which is frequently overexpressed in melanoma. Together, the collaborations among members of the Program Project will combine expertise in tumor biology, tumor immunology, biochemistry, chemistry and structural biology to generate unique approaches and reagents for use in melanoma therapy.