Characterization of early events in bulge McSCs and their progeny during melanoma

描述(申请人提供)：这笔赠款的目标是研究黑素细胞干细胞(McSCs)及其后代的黑色素瘤形成潜力，从而了解黑色素瘤发生的最初事件。在美国，黑色素瘤的发病率正在增加，可能是因为皮肤暴露在紫外线辐射下。目前，当黑色素瘤发生转移时，没有办法治愈它。制定早期干预的目标和早期诊断和预防的有效策略是必要的，但参与黑色素瘤发生的机制尚不清楚。在小鼠黑色素瘤模型中，我们可以在黑素细胞中诱导致癌突变。然而，小鼠的黑素细胞主要驻留在毛囊内，它们的行为在毛发周期中发生变化，因此很难区分由致癌基因突变引起的TE变化。目前，还没有研究表明这些致癌基因突变何时以及如何在体内对黑素细胞产生影响。尽管人们认为干细胞可能是肿瘤起源细胞的最佳候选者，但McSCs在黑色素瘤中的参与仍不清楚。由于McSCs位于毛囊中，其主要功能是产生毛发黑素细胞，而大多数黑色素瘤发生于毛囊间表皮，因此这一群体在黑色素瘤研究中一直处于较低水平。我们发现，毛囊间充质干细胞在UVB辐射后能在成年小鼠的皮肤上形成黑素细胞。我们推测，隆起中的McSCs可能是导致黑色素瘤的表皮黑素细胞的来源。在目标1中，我们将使用遗传小鼠模型来诱导McSCs的致癌突变(在人类黑色素瘤中经常发现的BRAF激活和PTEN缺失)。我们还将利用基因可追踪的荧光记者来追踪 突变的黑素细胞。利用这些小鼠，我们将仔细描述突变如何导致毛囊周期每个阶段隆起和表皮中黑素细胞的增殖、分化和迁移状态的变化。这些分析将首先捕捉到 在致癌基因突变驱动的黑素细胞中的初始变化，并确定不同黑素细胞群体的黑色素瘤发生率。在目标2中，我们将从不同的位置分离黑素细胞，以检测它们在免疫缺陷小鼠移植和诱导突变时产生黑色素瘤的潜力。通过分析每个特定人群中黑色素瘤的发病率，我们将确定是否存在导致黑色素瘤形成的内在差异。通过这些原位和体外研究阐明起源的细胞最终将有助于识别预防黑色素瘤发生的新策略。

DESCRIPTION (provided by applicant): The goal of this grant is to investigate the melanoma-forming potential of melanocyte stem cells (McSCs) and their progeny, thereby understanding initial events of melanomagenesis. The incidence of melanoma is increasing in the United States, presumably due to increased skin exposure to UV radiation. Currently, there is no way to cure melanoma when it becomes metastatic. Developing targets for early intervention and effective strategies for early diagnosis and prevention are necessary, however the mechanism involved in melanoma initiation is elusive. In mouse melanoma models, we can induce oncogenic mutations in melanocytes. However, melanocytes in mice primarily reside within the hair follicle and their behavior alters during the hair cycle, making it difficult to distinguish te changes caused by oncogenic mutations. Currently, no study has addressed when and how these oncogenic mutations exert its effect on melanocytes in vivo. Despite the belief that stem cells may be the best candidate for the cell of origin in tumors, the involvement of McSCs in melanoma remains unknown. Since McSCs are located in the hair follicle and their primary function is to generate hair melanocytes whereas most melanomas arise from inter-follicular epidermis, this population has been under-studied in melanoma studies. We found that follicular McSCs give rise to epidermal melanocytes in adult mice following UVB-radiation. We hypothesize that McSCs in the bulge may be the origin of epidermal melanocytes that are responsible for melanoma. In Aim 1, we will use genetic mouse models to induce oncogenic mutations (BRAF activation and PTEN loss that are often found in human melanoma) in McSCs. We will also utilize a genetically tractable fluorescent reporter to trace the fate and behavior of the mutated melanocyte. Utilizing these mice, we will carefully delineate how mutations induce change in proliferation, differentiation and migration status of melanocytes in the bulge and epidermis at each stage of the hair follicle cycle. These analyses will be the first to capture the initial alterations in melanocytes driven by oncogenic mutations and to determine the incidence of melanomagenesis from distinct populations of melanocytes. In Aim 2, we will isolate the melanocytes from distinct locations to examine their potential to produce melanoma upon transplantation and induction of the mutations in immunodeficient mice. By analyzing the incidence of melanoma in each specific population, we will determine whether there are intrinsic differences that cause melanoma formation. Elucidating the cell of origin through these in situ and ex vivo studies will ultimately help identification of novel strategies to prevent melanomagenesis.