Song Yu, Zhao Qing-Li, Ogawa Ryohei, Mizukami Tatsuji, Li Yu-Mei, Cui Zheng-Guo, Saitoh Jun-Ichi, Noguchi Kyo
Division of Radiation Oncology, Department of Radiology, Faculty of Medicine, University of Toyama, Toyama 930-0194, Japan.
Department of Environmental Health, University of Fukui School of Medical Sciences, Fukui 910-1193, Japan.
Cell Signal. 2025 Aug;132:111854. doi: 10.1016/j.cellsig.2025.111854. Epub 2025 May 5.
In cancer therapeutics, natural flavonoid compounds are renowned for their diverse structures and broad biological activities, offering considerable opportunities for drug discovery. This study investigates the anticancer effects of the flavonoid 4,4'-dimethoxychalcone (DMC), focusing on its apoptotic mechanisms and therapeutic potential. Our findings reveal that DMC induces apoptosis by upregulating pro-apoptotic proteins (Bax, Bim, tBid) and downregulating anti-apoptotic proteins (Bcl-2, Mcl-1), with concurrent caspase-3 activation and PARP cleavage. This apoptotic effect is mitigated by Z-VAD-FMK, a pan-caspase inhibitor. DMC also induces mitochondrial membrane potential (MMP) loss and increases reactive oxygen species (ROS) production. Furthermore, DMC promotes endoplasmic reticulum (ER) stress, evidenced by the increased expression of p-PERK/PERK, p-IRE1/IRE1, GRP78, HSP70, ATF4, and CHOP proteins. ER stress inhibitors significantly reverse DMC-induced MMP loss, apoptosis, and upregulation of apoptosis-related proteins. Additionally, DMC activates the mitogen-activated protein kinase (MAPK) pathway, including Erk, JNK, and p38. DMC also promotes autophagosome accumulation, modulates autophagy marker proteins (LC3-II, ATG5, p62), and leads to lysosomal dysfunction-evidenced by downregulated LAMP-1 and Cathepsin D expression, lysosomal pH increase, yet unaffected LC3 and LAMP-1 co-localization. Modulating autophagy with inhibitors (3-methyladenine, 3-MA; chloroquine, CQ) or an inducer (rapamycin, Rapa) respectively enhances or reduces DMC-induced apoptosis. Treatment with 3-MA also led to a significant increase in the expression of ER stress markers CHOP and ATF4. Collectively, DMC-induced cell death is primarily due to ER stress activation and autophagic flux impairment via lysosomal dysfunction. These results suggest DMC's potential as an anticancer agent, warranting further clinical investigation.
在癌症治疗中，天然黄酮类化合物以其多样的结构和广泛的生物活性而闻名，为药物研发提供了大量机会。本研究调查了黄酮类化合物4,4'-二甲氧基查耳酮（DMC）的抗癌作用，重点关注其凋亡机制和治疗潜力。我们的研究结果表明，DMC通过上调促凋亡蛋白（Bax、Bim、tBid）和下调抗凋亡蛋白（Bcl-2、Mcl-1）诱导凋亡，同时激活半胱天冬酶-3并切割PARP。这种凋亡效应被泛半胱天冬酶抑制剂Z-VAD-FMK减轻。DMC还诱导线粒体膜电位（MMP）丧失并增加活性氧（ROS）生成。此外，DMC促进内质网（ER）应激，p-PERK/PERK、p-IRE1/IRE1、GRP78、HSP70、ATF4和CHOP蛋白表达增加证明了这一点。ER应激抑制剂显著逆转DMC诱导的MMP丧失、凋亡以及凋亡相关蛋白的上调。此外，DMC激活丝裂原活化蛋白激酶（MAPK）途径，包括Erk、JNK和p38。DMC还促进自噬体积累，调节自噬标记蛋白（LC3-II、ATG5、p62），并导致溶酶体功能障碍——LAMP-1和组织蛋白酶D表达下调、溶酶体pH值升高证明了这一点，但LC3和LAMP-1共定位未受影响。分别用抑制剂（3-甲基腺嘌呤，3-MA；氯喹，CQ）或诱导剂（雷帕霉素，Rapa）调节自噬可增强或降低DMC诱导的凋亡。用3-MA处理还导致ER应激标记物CHOP和ATF4的表达显著增加。总体而言，DMC诱导的细胞死亡主要归因于ER应激激活和通过溶酶体功能障碍导致的自噬流受损。这些结果表明DMC作为抗癌剂的潜力，值得进一步的临床研究。
