Ko Ching-Huai, Shen Shing-Chuan, Lee Tony J F, Chen Yen-Chou
Graduate Institute of Pharmaceutical Sciences, School of Pharmacy, 250 Wu-Hsing Street, Taipei, Taiwan.
Mol Cancer Ther. 2005 Feb;4(2):281-90.
Colorectal carcinoma is a leading cause of human mortality due to its high metastatic ability. Because the activation of matrix metalloproteinases (MMP) is a key factor in the metastatic process, agents with the ability to inhibit MMP activity have potential in the treatment of colorectal carcinoma. In the present study, among 36 flavonoids examined, myricetin was found to be the most potent inhibitor of MMP-2 enzyme activity in COLO 205 cells (IC50 = 7.82 micromol/L). Myricetin inhibition of MMP-2 enzyme activity was also found in the human colorectal carcinoma cell lines COLO 320HSR, COLO 320DM, HT 29, and COLO 205-X (IC50 = 11.18, 11.56, 13.25, and 23.51 micromol/L, respectively). In contrast, no inhibitory effect of MMP-2 protein expression or enzyme activity was observed in myricitrin (myricetin-3-rhamnoside)-treated cells. In 12-O-tetradecanoylphorbol-13-acetate (TPA)-stimulated COLO 205 cells, an increase in MMP-2 protein expression and enzyme activity, as well as of protein kinase C (PKC) alpha protein translocation, extracellular signal-regulated kinase (ERK) 1/2 protein phosphorylation, and c-Jun protein expression was observed. ERK inhibitor (PD98059) and PKC inhibitors (GF-109203X and H-7), but not p38 inhibitor (SB203580) or c-jun-NH2-kinase inhibitor (SP600125), significantly inhibited TPA-induced MMP-2 protein expression, with reduced ERK phosphorylation and c-Jun protein expression. Addition of myricetin but not myricitrin suppressed TPA-induced MMP-2 protein expression in COLO 205 cells by blocking the TPA-induced events, including translocation of PKCalpha from cytosol to membrane, phosphorylation of ERK1/2 protein, and induction of c-Jun protein expression. Addition of PD98059 or GF-109203X significantly enhanced the inhibitory effect of myricetin on MMP-2 enzyme activity induced by TPA. Furthermore, myricetin, but not myricitrin, suppressed TPA-induced invasion of COLO 205 cells in an in vitro invasion assay using Engelbreth-Holm-Swarm sarcoma tumor extract Matrigel-coated Transwells. Results of the present study indicate that myricetin significantly blocked both endogenous and TPA-induced MMP-2 enzyme activity by inhibiting its protein expression and enzyme activity. The blockade involved suppression of PKC translocation, ERK phosphorylation, and c-Jun protein expression.
由于其高转移能力，结直肠癌是导致人类死亡的主要原因。因为基质金属蛋白酶（MMP）的激活是转移过程中的关键因素，具有抑制MMP活性能力的药物在结直肠癌治疗中具有潜力。在本研究中，在所检测的36种黄酮类化合物中，发现杨梅素是COLO 205细胞中MMP-2酶活性最有效的抑制剂（IC50 = 7.82微摩尔/升）。在人结直肠癌细胞系COLO 320HSR、COLO 320DM、HT 29和COLO 205-X中也发现杨梅素对MMP-2酶活性有抑制作用（IC50分别为11.18、11.56、13.25和23.51微摩尔/升）。相比之下，在杨梅苷（杨梅素-3-鼠李糖苷）处理的细胞中未观察到对MMP-2蛋白表达或酶活性的抑制作用。在12-O-十四酰佛波醇-13-乙酸酯（TPA）刺激的COLO 205细胞中，观察到MMP-2蛋白表达和酶活性增加，以及蛋白激酶C（PKC）α蛋白易位、细胞外信号调节激酶（ERK）1/2蛋白磷酸化和c-Jun蛋白表达增加。ERK抑制剂（PD98059）和PKC抑制剂（GF-109203X和H-7），而不是p38抑制剂（SB203580）或c-Jun氨基末端激酶抑制剂（SP600L25），显著抑制TPA诱导的MMP-2蛋白表达，同时ERK磷酸化和c-Jun蛋白表达降低。添加杨梅素而非杨梅苷通过阻断TPA诱导的事件，包括PKCα从细胞质到细胞膜的易位、ERK1/2蛋白的磷酸化以及c-Jun蛋白表达的诱导，抑制了COLO 205细胞中TPA诱导的MMP-2蛋白表达。添加PD98059或GF-109203X显著增强了杨梅素对TPA诱导的MMP-2酶活性的抑制作用。此外，在使用Engelbreth-Holm-Swarm肉瘤肿瘤提取物包被Matrigel的Transwell体外侵袭试验中，杨梅素而非杨梅苷抑制了TPA诱导的COLO 205细胞侵袭。本研究结果表明，杨梅素通过抑制其蛋白表达和酶活性，显著阻断内源性和TPA诱导的MMP-2酶活性。这种阻断涉及抑制PKC易位、ERK磷酸化和c-Jun蛋白表达。
