Identifying Events and Genetic Regulators of Melanoma Progression

描述(由申请人提供)：本提案的目标是检查黑色素瘤形成的事件和遗传缺陷。黑色素瘤是最具侵袭性和致命性的皮肤癌。大多数良性痣和黑色素瘤都有激活BRAF丝氨酸/苏氨酸激酶的突变，这表明BRAF激活是肿瘤发生中重要但不充分的一步。P53通路的改变也与黑色素瘤的形成有关。虽然许多功能都归因于BRAF和P53，但人们对这些功能中的哪一个在将正常黑素细胞转化为黑色素瘤细胞方面发挥了重要作用却知之甚少。此外，导致这一过程的其他遗传缺陷尚未确定。表达人致癌BRAF的斑马鱼品系已经被创造出来，并且是p53的突变株。这些菌株可靠地发展为黑色素瘤，在组织病理学上与人类相似。初步结果表明，这些突变体中的双核、多倍体黑素细胞可能会导致肿瘤。在目标1中，将检查黑色素瘤形成的事件。斑马鱼中的转基因方法和对培养中的哺乳动物细胞的长期成像将被用来确定BRAF导致双核的机制。细胞分选结合移植将被用来确定双核细胞是否会导致黑色素瘤。在目标2中，有助于黑色素瘤形成的遗传缺陷将被识别和表征。在初步研究中，Kit受体酪氨酸激酶被确认为黑色素瘤发生和侵袭的抑制因子。将评估其抑制黑色素瘤所需的试剂盒的活性。为了识别更多的黑色素瘤遗传缺陷，将使用阵列比较基因组杂交(ACGH)来表征这些肿瘤中反复出现的染色体获得和丢失。来自人类和斑马鱼黑色素瘤的aCGH数据将被用来确定候选癌基因，这些基因将在一种新的高通量测试中进行测试，以确定对黑色素瘤形成的贡献。这些研究可以确定疾病的诊断和预后指标以及癌症治疗的治疗目标。

DESCRIPTION (provided by applicant): The goals of this proposal are to examine the events and genetic defects that underlie melanoma formation. Melanoma is the most aggressive and lethal form of skin cancer. Most benign nevi and melanomas have mutations activating the BRAF serine/threonine kinase, suggesting that BRAF activation is an important but insufficient step in tumorigenesis. p53 pathway alterations are also implicated in melanoma formation. While many functions have been ascribed to BRAF and p53, little is known about which if any of these functions is important in transforming normal melanocytes into melanoma cells. Furthermore, additional genetic defects that contribute this process have yet to be defined. Zebrafish strains that express human oncogenic BRAF and are mutant for p53 have been created. These strains reliably develop melanomas that are histopathologically similar to those in humans. Preliminary results indicate that binucleate, polyploid melanocytes in these mutants may give rise to tumors. In Aim 1, the events of melanoma formation will be examined. Transgenic approaches in zebrafish and long-term imaging of mammalian cells in culture will be used to determine the mechanism by which BRAF causes binuclearity. Cell sorting coupled with transplantation will be employed to determine whether binucleate cells can give rise to melanomas. In Aim 2, genetic defects that contribute to melanoma formation will be identified and characterized. In preliminary studies, the Kit receptor tyrosine kinase was identified as a suppressor of melanoma onset and invasion. Activities of Kit required for its melanoma suppression will be assessed. To identify additional melanoma genetic defects, array comparative genomic hybridization (aCGH) will be used to characterize recurrent chromosomal gains and losses in these tumors. aCGH data from human and zebrafish melanomas will be used to identify candidate oncogenes that will be tested in a novel, high-throughput assay for contribution to melanoma formation. These studies may identify diagnostic and prognostic indicators of disease as well as therapeutic targets for cancer treatment.