Ishikawa J, Xu H J, Hu S X, Yandell D W, Maeda S, Kamidono S, Benedict W F, Takahashi R
Center for Biotechnology, Baylor College of Medicine, The Woodlands, Texas 77381.
Cancer Res. 1991 Oct 15;51(20):5736-43.
The retinoblastoma (RB) gene was the first tumor suppressor gene isolated and its inactivation is associated with the pathogenesis of several types of human cancer. In this study, we investigated the involvement of the RB gene in bladder and renal cell carcinomas by determining the loss of heterozygosity (LOH) at the RB locus and by DNA, RNA, and protein analysis of the RB gene. Whenever possible, the latter included Western blotting and immunohistochemical staining of the RB protein. In bladder carcinoma, 2 of the 8 cell lines we studied had an inactivated RB gene; one cell line lacked RB expression without a gross RB deletion, whereas the other cell line expressed only the underphosphorylated form of the RB protein. None of 16 low-grade noninvasive bladder carcinomas showed an alteration in RB protein by direct Western blot analysis, whereas 2 of 14 high-grade, invasive tumors had no RB protein as measured by both Western blotting and immunohistochemical staining. This suggests that the loss of RB function may be more important in the progression of bladder cancer than in its initiation, although more extensive studies are required. LOH within the RB locus was observed in 5 of 27 informative cases of primary bladder, ureter, or renal pelvis carcinoma. However, none of the 5 cases with LOH at the RB locus had a functional loss of RB protein expression. In renal cell carcinoma, one of the 12 cell lines had a gross homozygous deletion of the RB gene, and 2 of 32 primary tumors were negative for RB protein expression. LOH at the RB locus also was found in only 2 of 30 informative cases, one of which lacked RB expression. These results are the first to demonstrate the involvement of RB inactivation in the development of advanced primary bladder carcinoma and suggest that RB loss could have a role in certain renal cell carcinomas. Our data, however, show no correlation between LOH at the RB locus in bladder cancer and actual inactivation of the RB gene at the protein level. This may suggest that there is a second tumor suppressor or recessive cancer gene on chromosome 13 in bladder cancer and/or that the mechanism of RB inactivation in bladder cancer frequently involves independent mutations of each RB allele.
视网膜母细胞瘤（RB）基因是首个被分离出来的肿瘤抑制基因，其失活与多种人类癌症的发病机制相关。在本研究中，我们通过确定RB基因座处杂合性缺失（LOH）以及对RB基因进行DNA、RNA和蛋白质分析，来研究RB基因在膀胱癌和肾细胞癌中的作用。只要有可能，后者包括对RB蛋白进行蛋白质印迹法和免疫组织化学染色。在膀胱癌中，我们研究的8个细胞系中有2个的RB基因失活；一个细胞系没有明显的RB基因缺失但缺乏RB表达，而另一个细胞系仅表达RB蛋白的低磷酸化形式。16例低级别非侵袭性膀胱癌经直接蛋白质印迹法分析均未显示RB蛋白有改变，而14例高级别侵袭性肿瘤中有2例经蛋白质印迹法和免疫组织化学染色检测均无RB蛋白。这表明RB功能丧失在膀胱癌进展中可能比在其起始阶段更重要，尽管还需要更广泛的研究。在27例原发性膀胱、输尿管或肾盂癌的信息性病例中有5例观察到RB基因座内存在LOH。然而，这5例RB基因座处有LOH的病例中，没有一例RB蛋白表达出现功能丧失。在肾细胞癌中，12个细胞系中有1个存在RB基因的纯合大片段缺失，32例原发性肿瘤中有2例RB蛋白表达为阴性。在30例信息性病例中只有2例在RB基因座处发现LOH，其中1例缺乏RB表达。这些结果首次证明了RB失活参与晚期原发性膀胱癌的发生发展，并提示RB缺失可能在某些肾细胞癌中起作用。然而，我们的数据显示膀胱癌中RB基因座处的LOH与蛋白质水平上RB基因的实际失活之间没有相关性。这可能表明膀胱癌13号染色体上存在第二个肿瘤抑制基因或隐性癌基因，和/或膀胱癌中RB失活机制经常涉及每个RB等位基因的独立突变。
