PROGNOSTIC SIGNIFICANCE AND ANALYSIS OF iNOS IN MELANOMA

目前的数据表明，黑色素瘤的发生过程是由于诱导型一氧化氮合酶(INOS)的异常结构性表达而增强的，iNOS是一种催化一氧化氮(NO)生成的酶，通常是对细胞因子的反应。黑色素瘤中反应性NO的存在是通过检测硝基酪氨酸(NT)来指示的，它是NO与蛋白质化学反应的最终产物；NT的形成被认为改变了细胞内的信号，特别是当靶标是酪氨酸激酶底物时。 人类肿瘤产生的低水平(PM)NO被认为与增加血管生成、生长、侵袭、基因组不稳定和抗凋亡有关。我们对晚期(III期)疾病患者的人类黑色素瘤样本的分析表明，肿瘤表达诱导型一氧化氮合酶的患者比没有诱导型一氧化氮合酶的患者更有可能在新辅助生物血液治疗的两年内死亡(p&lt；0.001)。具体目标1从临床到实验室，进一步测试组织标本中iNOS和NT作为原发和转移性黑色素瘤患者预后标志物的意义。 特定目标1的翻译目标是确认iNOS和NT作为黑色素瘤患者生存的新的预后标记物，并确定这两个标记物是否提供独立的预后价值。然后，重要的结果将提交给美国癌症联合委员会和黑色素瘤委员会，作为黑色素瘤的第一个分子标记物被纳入未来的分期修订中。特定目的2测试了iNOS受特定的、可识别的基因转录调控因子控制的假设，其中包括构成活性的NFKB。在正常人细胞中，NFkB的激活既可调节基础iNOS，也可调节细胞因子诱导的iNOS，但其在黑色素瘤iNOS调节中的作用尚不清楚。特异性目标3建立在iNOS激活的进一步上游机制的基础上，并测试突变的B-RAF的关联和功能作用，据报道，突变的B-RAF独立于RAS驱动MAPK。 黑色素瘤细胞。一种新的分子异源双链分析将用于检测石蜡包埋肿瘤标本中突变的B-RAF，并研究其与iNOS和结构性NFKappaB的相关性。突变的B-RAF活性的调节也在特定的目标4中被提出，作为一种调节iNOS的手段，进而调节黑色素瘤的生长。

Current data suggest that the process of melanoma tumorigenesis is enhanced by aberrant constitutive expression of the inducible form of nitric oxide synthase (iNOS), an enzyme catalyzing the generation of nitric oxide (NO), normally in response to cytokines. The presence of reactive NO in melanoma is indicated by detection of nitrotyrosine (NT), which is an end-product of the chemical reaction of NO with proteins; NT formation is known to alter intracellular signaling, especially when the target is a tyrosine kinase substrate. Production of low levels (pM) of NO by human tumors is proposed to be responsible for increased angiogenesis, growth, invasion, genomic instability and resistance to apoptosis. Our analysis of human melanoma specimens from patients with advanced (Stage III) disease indicates that patients whose tumors express iNOS are more likely to die within 2 years of neoadjuvant biohemotherapy than patients without iNOS (p<0.001). Specific Aim 1 comes from the clinic to the laboratory to further test the significance of iNOS and NT in tissue specimens as prognostic markers for primary and metastatic melanoma patients. The translational goals of Specific Aim 1 are validation of iNOS and NT as new prognostic markers for survival in melanoma patients and to determine whether either marker provides independent prognostic value. Significant results will then be submitted to American Joint Commission on Cancer, Committee on Melanoma for inclusion in future staging revisions, as the first molecular marker for melanoma to be validated. Specific Aim 2 tests the hypothesis that iNOS is controlled by specific, identifiable gene transcriptional regulators including constitutively active NFKB. NFKB activation is known to regulate basal as well as cytokine-induced iNOS in normal human cells, however its role in regulation of melanoma iNOS is unknown. Specific Aim 3 builds on further upstream mechanisms of iNOS activation and tests the association and functional role of mutated B-raf, which is reported to drive MAPK independently of Ras in melanoma cells. A novel molecular heteroduplex analysis will be employed to detect mutated B-raf in paraffin-embedded tumor specimens, and study of its association with iNOS and constitutive NFKappaB. The regulation of mutated B-raf activity is also proposed in Specific Aim 4 as a means to regulate iNOS and, by extension, melanoma growth.