MOLECULAR MARKERS FOR PROSTATE CANCER

产品说明：前列腺癌是男性中最常见的癌症，但目前用于早期检测前列腺癌的方法不足且准确性有限。该项目的总体目标是识别前列腺肿瘤中的分子异常，以开发用于前列腺癌患者早期诊断和预后评估的新工具。这将通过监测不同类型前列腺组织中基因表达的差异来实现。差异表达的候选基因可能与肿瘤发生不直接相关，但可能有助于开发诊断标志物。我们将使用差异显示，逆转录酶PCR和基因发现阵列筛选比较mRNA丰度在5个人前列腺肿瘤细胞系。通过细胞系的RNA印迹分析确认可识别cDNA片段的表达模式。将对阳性候选物进行克隆、测序并通过BLAST分析进行鉴定。接下来，将评估鉴定的cDNA作为前列腺组织中肿瘤标志物的潜力。前瞻性标记物将用于筛选前列腺癌患者的组织样本。石蜡包埋的档案或新鲜样本将通过原位杂交进行筛选。将通过RTPCR筛选新鲜组织。将筛选未受累、结节性增生、不同级别PIN和前列腺腺癌五组组织，并对结果进行统计学分析。将寻找一两个最突出的候选cDNA片段来开发优化的诊断测试。将通过RTPCR克隆具有已知身份的潜在标记基因的全长或部分cDNA。身份未知的基因的cDNA将通过cDNA的快速扩增或通过筛选cDNA文库来克隆。克隆的cDNA将在E.大肠杆菌和更常见的蛋白质将用于产生多克隆抗体。如果确实可以使用合成肽来产生抗体，则根据表达模式，将开发使用多克隆抗体的基于ELISA的测定或更灵敏的基于RTPCR的测定。

DESCRIPTION: (adapted verbatim from the investigator's abstract) Prostatic adenocarcinoma is the most common cancer diagnosed in males, yet currently available methods for early detection of prostate cancer are inadequate and limited in accuracy. The overall goal of this project is to identify molecular abnormalities in prostate tumors to develop new tools for early diagnosis and evaluating prognosis of patients with prostate cancer. This will be accomplished by monitoring differences in gene expression in various types of prostatic tissues. Differentially expressed candidate genes may not be directly associated with tumorigenesis but may be useful for developing diagnostic markers. We will use differential display, reverse transcriptase PCR and gene discovery array filter screening to compare mRNA abundance in 5 human prostate tumor cell lines. Expression patterns of identifiable cDNA fragments will be confirmed by RNA blot analysis of cell lines. Positive candidates will be cloned, sequenced and identified by BLAST analysis. Next, identified cDNAs will be evaluated for their potential as tumor markers in prostate tissues. Prospective markers will be used to screen tissue samples from patients with prostate cancer. Paraffi- embedded archival or fresh samples will be screened by in situ hybridization. Fresh tissues will be screened by RTPCR. Five groups of tissue including uninvolved, nodular hyperplasia, different grades of PIN and prostatic adenocarcinoma will be screened, and results evaluated by statistical analysis. One or two of the most prominent candidate cDNA fragments will be pursued to develop an optimized diagnostic test. Either a full length or partial cDNA of a potential marker gene with known identity will be cloned by RTPCR. cDNA of genes with unknown identity will be cloned by rapid amplification of cDNA or by screening a cDNA library. Cloned cDNAs will be expressed in E. coli and more common proteins will be used to raise polyclonal antibodies. If indeed synthetic peptides may be used to raise antibody, depending upon the expression pattern, either an ELISA based assay using a polyclonal antibody or a more sensitive RTPCR based assay will be developed.