Matin S F, Rackley R R, Sadhukhan P C, Kim M S, Novick A C, Bandyopadhyay S K
Department of Urology, Lerner Research Institute, The Cleveland Clinic Foundation, Ohio 44195, USA.
Cancer Res. 2001 Mar 1;61(5):2261-6.
The limited success of IFN-alpha therapy for clinical treatment of transitional cell carcinoma (TCC) has prompted us to investigate the responsiveness of TCC lines to IFN-alpha. The response to IFN-alpha in terms of 561 gene induction, an IFN-stimulated response element-containing IFN-alpha/beta-inducible gene, and IFN-stimulated gene factor 3 (ISGF3) formation was normal in primary human urothelial cells. We tested the antiproliferative effects of IFN-alpha in three TCC lines as a measure of IFN-alpha responsiveness, and variable patterns of growth inhibition were observed in three TCC lines. More than 90% growth inhibition was noted in TCCSUP cells, whereas only 40% and 10% inhibition by IFN-alpha was observed in 5637 and HT1197 cells, respectively. IFN-alpha treatment formed extremely low levels of ISGF3 in electrophoretic mobility shift assays in these later two relatively insensitive cells. In addition, expression of the 561 gene was significantly reduced in these two TCC lines by Northern blots. We have further identified a low expression level of Tyk2 in HT1197 cells compared with two other TCCs. This suggests that an extremely low ISGF3 level after IFN-alpha treatment may be due to low Tyk2 expression or other unidentified defects. In 5637 cells, p48 protein expression was undetectable. This undetectable p48 expression is not due to a deletion in the coding region because the correct size protein is detected following IFN-gamma treatment. Consequently, the ISGF3 complex formation and 561 gene induction were restored by IFN-gamma pretreatment plus IFN-alpha treatment. Introduction of p48 expressing plasmid into 5637 cells was sufficient to form the ISGF3 complex by IFN-alpha treatment, suggesting the defect lies in the expression of p48 protein in 5637 cells. Detailed mechanistic understanding of the action of IFNs in bladder cancer cell lines may explain the abrogated therapeutic response of IFN-alpha in the clinical treatment of TCCs.
α干扰素治疗移行细胞癌（TCC）的临床效果有限，促使我们研究TCC细胞系对α干扰素的反应性。在原代人尿路上皮细胞中，就561基因诱导（一种含干扰素刺激反应元件的α/β干扰素诱导基因）以及干扰素刺激基因因子3（ISGF3）形成而言，对α干扰素的反应是正常的。我们测试了α干扰素在三种TCC细胞系中的抗增殖作用，以此作为α干扰素反应性的指标，在三种TCC细胞系中观察到了不同的生长抑制模式。在TCCSUP细胞中观察到超过90%的生长抑制，而在5637和HT1197细胞中，α干扰素分别仅导致40%和10%的抑制。在电泳迁移率变动分析中，在这两种相对不敏感的细胞中，α干扰素处理形成的ISGF3水平极低。此外，通过Northern印迹法发现在这两种TCC细胞系中561基因的表达显著降低。我们进一步发现，与其他两种TCC细胞相比，HT1197细胞中Tyk2的表达水平较低。这表明α干扰素处理后ISGF3水平极低可能是由于Tyk2表达低或其他未确定的缺陷。在5637细胞中，未检测到p48蛋白表达。这种未检测到的p48表达并非由于编码区缺失，因为在γ干扰素处理后可检测到正确大小的蛋白。因此，γ干扰素预处理加α干扰素处理可恢复ISGF3复合物形成和561基因诱导。将表达p48的质粒导入5637细胞足以通过α干扰素处理形成ISGF3复合物，这表明缺陷在于5637细胞中p48蛋白的表达。对干扰素在膀胱癌细胞系中作用的详细机制理解可能解释α干扰素在TCC临床治疗中治疗反应不佳的原因。
