Peripheral Blood Molecular Staging of Breast Cancer

描述(申请人提供)：乳腺癌仍然是美国女性中最常见的癌症，估计占所有女性新发癌症病例的30%。目前乳腺癌的分期方法是侵入性的、昂贵的、缺乏敏感性的，限制了临床医生为个别患者量身定做治疗方法的能力。乳腺癌过度表达基因的鉴定和分子生物学的最新进展为建立更敏感、更特异、更具成本效益的乳腺癌分期方法提供了机会。最近开发了两种新技术，显著提高了检测外周血中乳腺癌相关基因过度表达的能力：一种新的多孔屏障密度梯度离心法和实时RT-PCR技术。这项研究的假设是，乳腺癌患者外周血中乳腺癌细胞的实时RT-PCR检测与临床结果相关。在具体目标1，目标1中，我们将建立确定阳性分子检测结果的标准。为了确定标记物阳性的严格阈值，我们将从一组没有恶性肿瘤证据的健康志愿者队列中获取外周血样本。我们将确定候选分子标记在这些标本中的背景表达。我们还将确定候选标记物的背景表达是否独立于基线患者特征，包括年龄(50对50)、种族(白人与非白人)和良性乳腺疾病的存在(存在与不存在)。在目标2中，我们将设计和验证内部阳性对照基因片段模拟物，作为分子检测的附加对照。在具体目标2中，我们将在圣路易斯华盛顿大学和南卡罗来纳医科大学的92名受试者参加的双盲前瞻性队列研究中，确定循环乳腺癌细胞分子检测的临床相关性。基本假设是，与外周血液中没有乳腺癌分子证据的IV期乳腺癌患者相比，在外周血液中有乳腺癌分子证据的IV期乳腺癌患者开始新的系统治疗方案将经历减少的一年无进展和总存活率。次要目标将解决在治疗开始后对乳腺癌细胞进行分子检测的重要性。 分子遗传学的进步给医学带来了一场革命；成功开发出一种检测乳腺癌患者外周血中乳腺癌细胞的分子检测方法，可能会对那些患有这种疾病的不幸女性起到同样的作用。

DESCRIPTION (provided by applicant): Breast cancer remains the most common cancer among American women, representing an estimated 30% of all new cancer cases in women. The current staging modalities for breast cancer are invasive, expensive, and lack sensitivity, limiting the ability of clinicians to tailor therapies to individual patients. The identification of genes overexpressed in breast cancer combined with recent advances in molecular biology provide the opportunity to establish more sensitive, specific, and potentially cost-effective ways of staging breast cancer. Two new technologies have recently been developed that significantly enhance the ability to detect breast cancer-associated gene overexpression in the peripheral blood: a novel porous barrier density gradient centrifugation technique, and real-time RT-PCR. The hypothesis of the proposed research is that realtime RT-PCR detection of breast cancer cells in the peripheral blood of breast cancer patients is associated with clinical outcome. In Specific Aim 1, Objective 1 we will establish criteria for defining positive molecular test results. To establish rigorous threshold values for marker positivity, we will obtain peripheral blood samples from a cohort of healthy volunteer subjects with no evidence of malignancy. We will determine the background expression of candidate molecular markers in these specimens. We will also determine if the background expression of candidate markers is independent of baseline patient characteristics including age (>50 versus =50), race (White versus Not White) and the presence of benign breast disease (Present versus Absent). In Objective 2, we will design and validate internal positive control gene fragment mimetics to be used as an additional control for the molecular assays. In Specific Aim 2, we will define the clinical relevance of molecular detection of circulating breast cancer cells in a double-blinded prospective cohort study involving 92 subjects at Washington University in Saint Louis and the Medical University of South Carolina. The primary hypothesis is that Stage IV breast cancer patients initiating a new regimen of systemic therapy with molecular evidence of breast cancer in the peripheral blood will experience a decreased 1-year progression-free and overall survival compared to Stage IV breast cancer patients with no molecular evidence of breast cancer in the peripheral blood. Secondary objectives will address the significance of molecular detection of breast cancer cells after the initiation of therapy. Advances in molecular genetics have brought a revolution to medicine; the successful development of a molecular assay for the detection of breast cancer cells in the peripheral blood of breast cancer patients is likely to do the same for those unfortunate women afflicted with this disease.