Project 3: Mechanisms and therapeutic targeting of NRAS in melanoma

摘要 关于RAS亚型在功能上的不同及其影响，还有很多需要了解 每个RAS蛋白上的特定突变。NRAS突变黑色素瘤代表着两种严重的未得到满足的需求 有效治疗方案的条款，也是阐明这些功能的绝佳机会 不同之处。虽然KRAS是癌症中突变的主要RAS亚型，但NRAS是 在恶性黑色素瘤中，主要的RAS亚型发生突变，而61密码子突变在恶性黑色素瘤中很少见 KRAS，他们在NRAS中占主导地位。项目2最近证明了NRAS Q61R而不是G12D可以 促进Ink4a缺陷小鼠黑色素瘤的形成。这些具有挑衅性的发现表明，NRAS Q61R和 G12D必须激活不同的效应器，和/或以不同的方式激活效应器。鉴于优惠待遇 NRAS Q61突变在黑色素瘤中的发生，我们建议进行研究以阐明其信号机制 这区分了Q61和G12突变的NRAS在驱动这些癌症中的作用。我们假设 它的结构、生化和生物学特性与更常见的G12突变体不同 在肺癌、结直肠癌和胰腺癌中发现了KRAS蛋白。我们还发现了一种意想不到的 NRAS突变型黑色素瘤对KRAS和HRAS WT形式的要求。我们将调查效应器 由不同的NRAS突变体驱动的信号机制，定义了WT亚型的要求，以及 利用候选方法和不偏不倚的方法来确定可能导致新组合的目标 有效治疗NRAS驱动的黑色素瘤的治疗方法。为了实现我们的目标，我们提出了三个目标。在……里面 目标1，我们将阐明细胞NRAS下游已知和未知的效应信号通路 密码子61处的突变与密码子12处的突变。这一目标将与项目2密切合作，该项目 将重点研究同一组NRAS突变体的结构和生化差异。项目1和项目4将 还要检查不同RAS中相同突变的一些细胞和致瘤表型 异构体。在目标2中，我们将表征WT RAS亚型的需求，并确定功能 效应器和信号网络的差异，以及转化后的生长特性。在《目标3》中，我们将 确定克服黑色素瘤对突变型NRAS上瘾的相同机制是否也能克服 依赖于WT RAS亚型。要做到这一点，我们首先要审问YAP，他被认为有能力拯救 癌症中突变KRAS的成瘾，接下来我们将进行一项新的功能基因筛查以确定 以不偏不倚的方式建立非YAP机制。总的来说，我们的研究将阐明更好地理解 被忽略的NRAS亚型，表征了不同NRAS突变之间的功能差异，确定了 WT和突变的NRAS驱动的效应器信号通路和网络之间的关系，并识别 NRAS驱动的黑色素瘤的新治疗选择的新方向。

ABSTRACT Much remains to be learned about how RAS isoforms differ functionally from each other, and about the impact of specific mutations on each RAS protein. NRAS-mutant melanoma represents both a critical unmet need in terms of efficacious therapeutic options and also an outstanding opportunity to elucidate these functional differences. Although KRAS is the predominant RAS isoform mutated in cancers overall, NRAS is the predominant RAS isoform mutated in malignant melanoma, and whereas mutations at codon 61 are rare in KRAS, they predominate in NRAS. Project 2 has recently demonstrated that Nras Q61R but not G12D could drive melanoma formation in Ink4a-deficient mice. These provocative findings indicate that NRAS Q61R and G12D must activate distinct effectors, and/or activate effectors in a distinct manner. Given the preferential occurrence of NRAS Q61 mutations in melanoma, we propose studies to elucidate the signaling mechanisms that distinguish the roles of Q61- versus G12-mutant NRAS in driving these cancers. We hypothesize that there are structural, biochemical and biological properties distinct from those of the more common G12-mutant KRAS proteins found in lung, colorectal and pancreatic carcinomas. We have also identified an unexpected requirement for both KRAS and HRAS WT forms in NRAS-mutant melanomas. We will investigate effector signaling mechanisms driven by different NRAS mutants, define the requirements for WT isoforms, and leverage both candidate- and unbiased methodologies to identify targets that can lead to novel combination therapies for effective treatment of NRAS-driven melanomas. To address our goals, we propose three aims. In Aim 1, we will elucidate known and unknown effector signaling pathways downstream of cellular NRAS mutated at codon 61 versus codon 12. This Aim will be performed in close collaboration with Project 2, which will focus on structural and biochemical differences in the same panel of NRAS mutants. Projects 1 and 4 will also examine some of the cellular and tumorigenic phenotypes of the same mutations in different RAS isoforms. In Aim 2, we will characterize the requirement for WT RAS isoforms, and determine functional differences in effectors and signaling networks, as well as in transformed growth properties. In Aim 3, we will determine whether the same mechanisms that overcome melanoma addiction to mutant NRAS also overcome dependency on WT RAS isoforms. To do this, we will first interrogate YAP, known to be capable of rescuing addiction to mutant KRAS in carcinomas, and we will next perform a novel functional genetic screen to identify non-YAP mechanisms in an unbiased manner. Collectively, our studies will elucidate a better understanding of the neglected NRAS isoform, characterize functional distinctions among different NRAS mutations, determine relationships between WT and mutant NRAS-driven effector signaling pathways and networks, and identify new directions for novel therapeutic options in NRAS-driven melanoma.