PO.ET05.01 · 实验与分子治疗
托芬那酸抗增殖活性的机制
Mechanism of antiproliferative activity of tolfenamic acid
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
非甾体抗炎药已被证明主要通过抑制环氧合酶（COX-2）发挥抗肿瘤活性。其中，托芬那酸（tolfenamic acid，TA）在多种癌细胞系中也展现出强效的抗癌作用，但其作用通过不依赖COX的途径实现。TA提出的作用机制涉及其靶向特异性蛋白转录因子（Sp1、Sp3和Sp4）的能力，进而下调潜在的致癌蛋白，如survivin、VEGF和c-MET。我们实验室近期的研究表明，在髓母细胞瘤细胞中，TA能够诱导DNA双链断裂（DSB）。在本研究中，我们考察了TA在多种癌细胞系（包括神经母细胞瘤、卵巢癌、乳腺癌、胰腺癌和前列腺癌）中的DNA损伤效应，并检测了DNA损伤应答通路的激活情况。用递增浓度的TA处理代表各种癌症的细胞系48小时，通过Western blot分析监测组蛋白变体H2AX的磷酸化（gammaH2AX）来评估DNA损伤。通过Western blot检测ATM/ATR和Chk1/Chk2的激活情况，研究了髓母细胞瘤细胞系中TA处理后DSB应答的激活。通过PARP-1蛋白的切割检测细胞凋亡。与我们在髓母细胞瘤中的发现一致，在所有测试的癌细胞系中，TA处理48小时后我们观察到gammaH2AX呈剂量依赖性增加。所有细胞系中切割型PARP的水平也随TA处理而升高。此外，在髓母细胞瘤细胞中，处理后24小时即可检测到gammaH2AX。我们还证明了ATM和Chk2蛋白磷酸化呈剂量依赖性增加。综上所述，这些结果表明TA处理可诱导癌细胞中的DNA DSB，从而激活ATM和Chk2这一主要的DNA损伤应答通路，最终通过激活细胞凋亡导致细胞死亡。这些结果凸显了TA的多靶点效应，以及与其他小分子或药物开发有效联合策略以实现协同治疗效果的潜力。
查看英文原文 English abstract
Non-steroidal anti-inflammatory drugs have been shown to exhibit anti-tumor activities primarily through the inhibition of cyclooxygenases (COX-2). Among them, tolfenamic acid (TA) has also demonstrated potent anti-cancer effects across multiple cancer cell lines but acts through COX independent pathways. The proposed mechanism of action of TA involves its ability to target specificity protein transcription factors (Sp1, Sp3, and Sp4) which in turn downregulate potential oncogenic proteins such as survivin, VEGF, and c-MET. Recent work from our laboratory demonstrated, in medulloblastoma cells, the ability of TA to induce DNA double-strand breaks (DSBs). In this study we investigated the DNA damaging effects of TA across multiple cancer cell lines (including neuroblastoma, ovarian, breast, pancreatic, and prostate) and examined the activation of the DNA damage response pathways. Cell lines representing various cancers were treated with increasing concentration of TA for 48h and DNA damage assessed by monitoring phosphorylation of histone variant H2AX (gammaH2AX) by Western blot analysis. Activation of the DSB response following TA treatment, in a medulloblastoma cell line was investigated by examining the activation of ATM / ATR and Chk1 / Chk2 by Western blotting. Apoptosis was detected by cleavage of PARP-1 protein. Consistent with our findings in medulloblastoma, we observed a dose dependent increase in gammaH2AX, following TA treatment for 48h, in all cancer cell lines tested. Levels of cleaved PARP also increased with TA treatment in all cell lines. Furthermore, in medulloblastoma cells, gammaH2AX was detected as early as 24h post-treatment. We also demonstrated a dose dependent increase in phosphorylation of ATM and Chk2 proteins. Taken together, these results suggest that TA treatment induces DNA DSBs in cancer cells resulting in the activation of ATM and Chk2, a major DNA damage response pathway, ultimately leading to cell death via activation of apoptosis. The results highlight TA's multi-targeted effects and the potential to develop effective combination strategies with other small molecules or drugs to achieve synergistic therapeutic outcomes.
利益披露 Disclosure
F. Rehman, None..
H. Syed, None..
U. T. Sankpal, None.