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的第六大常见癌症。黑色素瘤的早期检测和手术切除仍然是黑色素瘤治愈性治疗的主要手段。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鉴定容易获得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将是非常有价值的。基于血液的生物标志物具有优于基于组织的生物标志物的几个优点。外周全血代表全身过程，包括免疫应答和细胞通信。鉴于黑色素瘤中存在肿瘤抑制和肿瘤促进免疫应答和炎症，并鉴于血液中存在循环肿瘤细胞、内皮细胞和骨髓源性细胞和前体细胞，我们假设使用来自黑色素瘤患者的外周血细胞和高通量qRT-PCR基因表达方法的血液生物标志物将为黑色素瘤提供新的生物标志物。虽然黑色素瘤组织和黑色素瘤细胞的遗传特征已被广泛研究，但尚未进行代表血液循环中肿瘤相关环境的黑色素瘤的基于血液的生物标志物研究。该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，我们已经鉴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集合。在这项提案中，我们将应用生物统计学方法来生成特定目标1中的签名模型，并将分析血液组分以确定负责特定目标2中候选基因变化的细胞亚群。我们的提案将提供第一个分析恶性黑色素瘤血液遗传生物标志物的综合研究。在未来，我们将把本提案中确定的特征模型应用到更广泛的黑色素瘤患者群体中以验证我们的数据，应用到其他癌症（如结肠癌、肺癌）和炎症性疾病（如类风湿性关节炎）人群中以验证和改进特异性，并应用到特定的黑色素瘤患者中以进一步回答有关特定生物标志物的问题。这些研究将在未来的R 01资助机会中提出。此外，该方法可能用于诊断各种疾病，预测预后和治疗反应以及风险评估，因此，在样品制备和数据分析方面的拟议研究将受到广泛关注。开发分析方法来检测和鉴定容易获得的体液（例如外周血）中的生物标志物对于癌症患者的早期诊断和管理将是非常有价值的。我们假设使用来自黑色素瘤患者的外周血细胞和高通量qRT-PCR基因表达方法的基于血液的生物标志物将为黑色素瘤提供新的生物标志物。我们已经确定了恶性黑色素瘤患者血液中的癌症组织或疾病血液中先前未描述的独特基因组，并将对基因应用生物统计学方法以生成恶性黑色素瘤的签名模型。

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.