Developing a protocol for profiling blood markers of melanoma

描述（由申请人提供）：恶性黑色素瘤是美国男性和女性第六大常见癌症。早期发现和手术切除黑色素瘤仍然是根治性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具有挑战性。因此，开发分析方法来检测和识别易获取的体液（如外周血）中的生物标志物，对于癌症患者的早期诊断和管理具有很高的价值。与基于组织的生物标志物相比，基于血液的生物标志物有几个优势。外周全血代表包括免疫反应和细胞通讯在内的系统过程。鉴于黑色素瘤中存在肿瘤抑制和肿瘤促进免疫反应和炎症，并且血液中存在循环肿瘤细胞、内皮细胞、骨髓来源细胞和前体细胞，我们假设使用黑色素瘤患者外周血细胞的血液生物标志物和高通量qRT-PCR基因表达方法将为黑色素瘤提供新的生物标志物。尽管黑素瘤组织和黑素瘤细胞的遗传特征已经被广泛研究，但代表血液循环中肿瘤相关环境的黑素瘤基于血液的生物标志物研究尚未进行。本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志物分析方案。在使用微阵列和qRT-PCR分析的初步实验中，我们已经确定了黑色素瘤患者血液中差异表达的独特基因，甚至来自I/II期黑色素瘤患者。我们发现的基因是以前在癌症组织或疾病血液中没有描述过的独特组合。在本提案中，我们将应用生物统计学方法在Specific Aim 1中生成签名模型，并将分析血液组分以确定在Specific Aim 2中负责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，分析恶性黑色素瘤的血液遗传生物标志物。在未来，我们将在更广泛的黑色素瘤患者中应用本提案中确定的签名模型来验证我们的数据，在其他癌症（如结肠癌，肺癌）和炎症性疾病（如类风湿关节炎）的人群中验证和完善特异性，并在特定的黑色素瘤患者中进一步回答特定生物标志物的问题。这些研究将在未来的R01资助机会中提出。此外，该方法对各种疾病的诊断、预后和治疗反应的预测以及风险评估都有潜在的用处，因此在样品制备和数据分析方面的拟议研究将引起广泛的兴趣。开发分析方法来检测和识别易获取的体液（如外周血）中的生物标志物，对于癌症患者的早期诊断和管理具有很高的价值。我们假设使用黑色素瘤患者外周血细胞的血液生物标志物和高通量qRT-PCR基因表达方法将为黑色素瘤提供新的生物标志物。我们已经确定了以前未在恶性黑色素瘤患者的癌症组织或疾病血液中描述的独特基因集，并将应用生物统计学方法对这些基因进行研究，以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.