Identification and validation of tumor biomarkers for urogenital cancer using proteomic analysis of cell culture supernatant

通过比较低侵袭性人膀胱癌细胞系和高侵袭性人膀胱癌细胞系的分泌蛋白质谱，寻找与膀胱癌侵袭相关蛋白质的候选蛋白质。用鸟枪式蛋白质组学方法对细胞培养上清液中分离的蛋白质进行鉴定。我们发现CXCL1调节膀胱癌细胞的侵袭能力。该趋化因子可能是侵袭性膀胱癌尿液生物标志物的潜在候选者，也是预防肿瘤侵袭的潜在治疗靶点。为鉴定和验证新的肾癌血清肿瘤标志物，我们建立了一种新的无血清培养上清液蛋白分析方法，并比较了稳定表达野生型pVHL和空表达载体的786-O肾细胞癌(VHL-/-)亚克隆的蛋白质组。利用该方法，我们鉴定了三种不同的VHL调节分泌蛋白(PAI-1、IGFBP-3和Clusterin)。慢性肾细胞癌患者血清IGFBP-3水平明显高于正常对照组。另一方面，我们利用新的多维鸟枪蛋白质组学技术，对4例散发性透明细胞肾癌患者肾切除术前后的血清蛋白质组进行了初步分析。共有6个蛋白质被鉴定为候选标记物，在手术前检测到，但在手术后消失。与癌旁正常组织相比，其中2例在肿瘤组织中呈双阳性表达。现在我们对这些候选蛋白质进行进一步的验证。

New proteomic approach for the identification of proteins that are potentially involved in the tumor invasion of bladder cancer.We searched for the candidate proteins by comparing the profiles of secreted proteins among the poorly invasive human bladder carcinoma cell line and the highly invasive cell line. The proteins isolated from cell culture supernatants were identified by shotgun proteomics. We found that CXCL1 modulates the invasive abilities of bladder cancer cells and. this chemokine maybe a potential candidate of urinary biomarker for invasive bladder cancer and a possible therapeutic target for preventing tumor invasion.New proteomic approach for the identification and validation of novel serum tumor makers for renal cell carcinoma.We established a new method fir analyzing the secreted proteins in cell culture supernatants by using serum-free media and compared the proteome among 786-O RCC (VHL-/-) subclones stably transfected with either a plasmid encoding wild-type pVHL or an empty expression plasmid. By using this method, we identified three different VHL regulated secreted proteins (PAI-1,IGFBP-3 and clusterin). IGFBP-3 is elevated significantly in the serum from patients with ccRCC comparing with that from normal controls. On the other hand, we had done the preliminary analysis of "serum proteome" of 4 patients with sporadic clear cell RCC before and after the nephrectomy by using novel multidimensional shotgun proteomics. A total of 6 proteins were identified as candidate markers that were detected before the operation, but disappeared after the procedure. Two of them were biddy expressed in tumor specimens comparing with that of normal adjacent normal tissue. Now we perform further validation of these candidate proteins.