Tolfenamic acid inhibits ovarian cancer cell growth and decreases the expression of c-Met and survivin through suppressing specificity protein transcription factors
Tolfenamic acid inhibits ovarian cancer cell growth and decreases the expression of c-Met and survivin through suppressing specificity protein transcription factors
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DOI：
10.1016/j.ygyno.2011.03.014
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发表时间：
2011-07-01
影响因子：
4.7
通讯作者：
Abdelrahim, Maen
中科院分区：
文献类型：
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作者：
Basha, Riyaz;Ingersoll, Susan B.;Abdelrahim, Maen
Objective.肝细胞生长因子及其受体c-Met的异常表达与包括卵巢癌（OC）在内的多种人类恶性肿瘤的侵袭性疾病和不良预后相关。特异性蛋白（Sp）转录因子在与c-Met活化相关的信号级联中具有高度相关性。托芬那酸（TA）是一种NSAID，已知其可诱导Sp蛋白降解，Sp蛋白降解与某些癌症患者的生存率呈负相关。我们的目的是使用体外和体内模型检测TA的抗OC活性，并评估这种新化合物的抑制作用。我们开发了源自原始SKOV 3的OC亚细胞系（AF 1 -3），其对IFN α-2b的抗性更强，并且在裸鼠中比原始细胞更具致瘤性。我们使用卵巢癌细胞SKOV 3-AF 2和ES-2测试TA的抗癌活性。用DMSO（溶媒）或TA（25/50/100 μ M）处理细胞，并在24、48和72 h时测定细胞活力。处理48小时后制备细胞裂解物（50 μ M），并通过Western印迹分析评估Sp蛋白（Sp1/Sp3/Sp 4）、c-Met、存活素、Bcl 2和裂解的聚ADP-核糖聚合酶（c-PARP）的表达。使用Caspase-Glo试剂盒评估Caspase-3活性。流式细胞仪分析细胞周期分布和凋亡情况。对于体内研究，将小鼠皮下注射ES-2细胞并用载体或TA（50 mg/kg/每周三次）处理。TA对SKOV 3（AF 1 -3）和ES-2细胞的生长有明显的抑制作用。SP蛋白和c-Met的表达显著降低，表明TA可以通过降解Sp蛋白而靶向c-Met。TA显著增加凋亡分数（Annexin V阳性）、c-PARP表达和caspase-3活性。TA可显著降低Bcl 2蛋白表达，并诱导G 0/G1期细胞阻滞。体内研究表明，TA显着抑制小鼠肿瘤重量和体积。以上结果表明TA对小鼠肿瘤生长有明显的抑制作用，可抑制OC细胞增殖，诱导细胞凋亡，阻滞细胞周期，降解Sp蛋白。TA还抑制了与放射抵抗相关的suivivin的表达，提示TA除了具有抑瘤作用外，还能增强肿瘤对放射治疗的反应。这些数据清楚地表明，TA在体外和体内有效地抑制OC细胞生长。这项研究表明TA抑制OC细胞生长的潜在作用，可能增强肿瘤对放射治疗的反应，并对OC治疗有影响。(C)2011 Elsevier Inc. All rights reserved.
Objective. The aberrant expression of hepatocyte growth factor and its receptor c-Met are associated with aggressive disease and poor prognosis in a variety of human malignancies including ovarian cancer (OC). Specificity protein (Sp) transcription factors have high relevance in the signaling cascade associated with c-Met activation. Tolfenamic acid (TA), a NSAID, is known to induce the degradation of Sp proteins, which have been negatively associated with survival in some cancer patients. Our aim was to examine the anti-OC activity of TA using in vitro and in vivo models and asses the inhibitory effects of this novel compound.Methods. We developed OC sub-cell lines (AF1-3) derived from the original SKOV3 that are more resistant to IFN alpha-2b and more tumorigenic in nude mice than the original cells. We tested the anti-cancer activity ofTA using ovarian cancer cells, SKOV3-AF2 and ES-2. The cells were treated with DMSO (vehicle) or TA (25/50/100 mu M) and cell viability was measured at 24, 48, and 72 h. Cell lysates were prepared following 48 h treatment (50 mu M) and evaluated the expression of Sp proteins (Sp1/Sp3/Sp4), c-Met, survivin, Bcl2, and cleaved polyADP-ribose polymerase (c-PARP) through Western blot analysis. Caspase-3 activity was assessed with Caspase-Glo kit. Cell cycle distribution and apoptosis were analyzed using BD FACSCalibur flow cytometer. For in vivo studies mice were subcutaneously injected with ES-2 cells and treated with vehicle or TA (50 mg/kg/thrice weekly).Results. TA significantly inhibited the growth of SKOV3 (AF1-3) and ES-2 cells. The expression of Sp proteins and c-Met was significantly decreased suggesting that TA could be targeting c-Met through degradation of Sp proteins. TA greatly increased the apoptotic fraction (Annexin V positive), c-PARP expression and caspase-3 activity. TA significantly decreased Bcl2 expression and induced G0/G1 cell cycle arrest. In vivo studies revealed that TA significantly inhibited tumor weight and volume in mice. These results show that TA has a profound inhibitory effect on tumor growth in mice, reduces OC cells proliferation, induces apoptosis, cell cycle arrest and Sp proteins degradation. TA also inhabited the expression of suivivin that is associated with radiation resistance and suggests that apart from its tumor suppressant effects, TA can also enhance the tumor response to radiotherapy.Conclusions. These data clearly show that TA effectively inhibits OC cell growth in vitro and in vivo. This study represents potential role(s) of TA that suppresses OC cell growth, and may enhance tumor response to radiotherapy, and have implications in OC treatment. (C) 2011 Elsevier Inc. All rights reserved.