Developing a protocol for profiling blood markers of melanoma

描述(申请人提供)：恶性黑色素瘤是美国男性和女性的第六大常见癌症。黑色素瘤的早期发现和手术切除仍然是治疗黑色素瘤的主要方法。然而，检测早期黑色素瘤和预测预后往往是具有挑战性的。因此，发展分析方法来检测和识别外周血等容易获得的体液中的生物标志物将对癌症患者的早期诊断和治疗具有非常重要的价值。基于血液的生物标记物比基于组织的生物标记物有几个优势。外周全血代表包括免疫反应和细胞通讯在内的全身过程。鉴于黑色素瘤中存在抑制肿瘤和促进肿瘤的免疫反应和炎症反应，血液中存在循环中的肿瘤细胞、内皮细胞、骨髓源性细胞和前体细胞，我们假设使用黑色素瘤患者外周血细胞的血液生物标记物和高通量qRT-PCR基因表达方法将为黑色素瘤提供新的生物标记物。虽然黑色素瘤组织和黑色素瘤细胞的遗传特征已经得到了广泛的研究，但代表血液循环中肿瘤相关环境的黑色素瘤的血液生物标志物研究还没有进行。这项提案的目标是开发一种使用高通量基因表达分析的黑色素瘤血液生物标记物分析方案。在使用微阵列和qRT-PCR分析的初步实验中，我们已经在黑色素瘤患者的血液中发现了差异表达的独特基因，甚至在I/II期黑色素瘤患者中也是如此。我们发现的这些基因是以前在癌症组织或疾病血液中未曾描述过的独特基因。在这个提案中，我们将应用生物统计学方法来生成特定目标1的特征模型，并将分析血液部分以确定特定目标2候选基因变化的细胞亚群。我们的提案将提供第一个全面的研究，分析恶性黑色素瘤基于血液的遗传生物标记物。未来，我们将把这项提案中确定的标志性模型应用于更广泛的黑色素瘤患者池中以验证我们的数据，应用于其他癌症(如结肠癌、肺癌)和炎症性疾病(如类风湿性关节炎)的人群以验证和完善特异性，并应用于特定的黑色素瘤患者以进一步回答关于特定生物标记物的问题。这些研究将在未来的R01资助机会中提出。此外，该方法学可能有助于广泛疾病的诊断、预后和治疗反应的预测以及风险评估，因此拟议的样品制备和数据分析研究将引起广泛的兴趣。项目简介开发分析方法以检测和识别外周血等容易获得的体液中的生物标记物，将对癌症患者的早期诊断和管理具有极高的价值。我们假设，使用黑色素瘤患者外周血细胞的血液生物标记物和高通量qRT-PCR基因表达方法将为黑色素瘤提供新的生物标记物。我们已经确定了以前在恶性黑色素瘤患者血液中的癌症组织或疾病血液中没有描述的独特的基因集，并将对这些基因应用生物统计学方法来生成恶性黑色素瘤的特征模型。

DESCRIPTION (provided by applicant): Malignant melanoma is the 6th most common cancer for men and women in the USA. Early detection and surgical resection of melanoma remains the mainstay of curative treatment of melanoma. However, detecting early melanoma and predicting prognosis is often challenging. Therefore, developing analytical methodologies to detect and identify biomarkers in easily accessible body fluid such as peripheral blood would be highly valuable for the early diagnosis and management of cancer patients. Blood-based biomarkers have several advantages over tissue-based biomarkers. Peripheral whole blood represents systemic processes including immune responses and cellular communications. Given the presence of tumor-inhibiting and tumor-promoting immune responses and inflammation in melanoma, and given the presence of circulating tumor cells, endothelial cells, and bone-marrow-derived cells and precursor cells in the blood, we hypothesize that blood-based biomarkers using peripheral blood cells from melanoma patients and high-throughput qRT-PCR gene expression methodology will provide new biomarkers for melanoma. Although genetic signatures of melanoma tissues and melanoma cells have been extensively studied, blood-based biomarker studies of melanoma that represent tumor-associated environment in the blood circulation have not been conducted. The goal of this proposal is to develop a protocol for blood biomarker profiling of melanoma using high- throughput gene expression analysis. In the preliminary experiments using microarray and qRT-PCR analyses, we have identified differentially expressed unique genes in the blood of melanoma patients, even from stage I/II melanoma patients. The genes we discovered are unique sets not previously described in cancer tissues or disease blood. In this proposal, we will apply biostatistical methodologies to generate signature models in Specific Aim 1, and will analyze fractions of blood to identify the cellular subsets responsible for the changes in candidate genes in Specific Aim 2. Our proposal will provide the first comprehensive study that analyzes blood-based genetic biomarkers of malignant melanoma. In the future, we will apply the signature models identified in this proposal into a more-extensive pool of melanoma patients to verify our data, into a population of other cancers (such as colon, lung cancers) and inflammatory diseases (such as rheumatoid arthritis) to validate and refine the specificity, and into a specific melanoma patients to further answer questions on specific biomarkers. These studies will be proposed in future R01 funding opportunities. Furthermore, the methodology is potentially useful for the diagnosis of a wide range of diseases, prediction of prognosis and treatment response, and risk assessment, thus the proposed studies in sample preparation and data analysis will be of wide interest. PROJECT NARRATIVE developing analytical methodologies to detect and identify biomarkers in easily accessible body fluid such as peripheral blood would be highly valuable for the early diagnosis and management of cancer patients. We hypothesize that blood-based biomarkers using peripheral blood cells from melanoma patients and high-throughput qRT-PCR gene expression methodology will provide new biomarkers for melanoma. We have identified unique sets of genes not previously described in cancer tissues or disease blood in the blood of malignant melanoma patients, and will apply biostatistical methodologies to the genes to generate signature models for malignant melanoma.