Department of Molecular Genetics, Faculty of Biology and Environmental Protection, University of Lodz, Pomorska 141/143, 90-236 Lodz, Poland.
Int J Mol Sci. 2024 Sep 24;25(19):10257. doi: 10.3390/ijms251910257.
Drug resistance is one of the major obstacles to the clinical use of doxorubicin, an extensively used chemotherapeutic drug to treat various cancers, including leukemia. Inhibition of the nuclear factor erythroid 2-related factor 2 (NRF2) seems a promising strategy to reverse chemoresistance in cancer cells. NRF2 is a transcription factor that regulates both antioxidant defense and drug detoxification mechanisms. In this study, we investigated the potential of three inhibitors of NRF2-K67, retinoic acid and ML-385-to overcome doxorubicin resistance in promyelocytic leukemia HL-60 cells. For this purpose, low-dose doxorubicin was used to establish doxorubicin-resistant HL-60/DR cells. The expression of NRF2 and its main repressor, Kelch-like ECH-associated protein 1 (KEAP1), at mRNA and protein levels was examined. HL-60/DR cells overexpressed NRF2 at mRNA and protein levels and down-regulated KEAP1 protein compared to drug-sensitive HL-60 cells. The effects of NRF2 inhibitors on doxorubicin-resistant HL-60/DR cell viability, apoptosis, and intracellular reactive oxygen species (ROS) levels were analyzed. We observed that NRF2 inhibitors significantly sensitized doxorubicin-resistant HL-60/DR cells to doxorubicin, which was associated with increased intracellular ROS levels and the expression of , suggesting the participation of the mitochondrial-dependent apoptosis pathway. Furthermore, ML-385 inhibitor was used to study the expression of NRF2-KEAP1 pathway genes. NRF2 gene and protein expression remained unchanged; however, we noted the down-regulation of KEAP1 protein upon ML-385 treatment. Additionally, the expression of NRF2-regulated antioxidant and detoxification genes including , , and was maintained upon ML-385 treatment. In conclusion, our results demonstrated that all the studied inhibitors, namely K67, retinoic acid, and ML-385, increased the efficacy of doxorubicin in doxorubicin-resistant HL-60/DR cells, and suggested a potential strategy of combination therapy using NRF2 inhibitors and doxorubicin in overcoming doxorubicin resistance in leukemia.
耐药性是多柔比星（一种广泛用于治疗包括白血病在内的各种癌症的化疗药物）临床应用的主要障碍之一。抑制核因子红细胞 2 相关因子 2（NRF2）似乎是逆转癌细胞化疗耐药性的一种有前途的策略。NRF2 是一种转录因子，可调节抗氧化防御和药物解毒机制。在这项研究中，我们研究了三种 NRF2 抑制剂（K67、维甲酸和 ML-385）在 HL-60 早幼粒细胞白血病细胞中克服多柔比星耐药性的潜力。为此，我们使用低剂量多柔比星建立了多柔比星耐药 HL-60/DR 细胞。检测了 NRF2 及其主要抑制因子 Kelch 样 ECH 相关蛋白 1（KEAP1）在 mRNA 和蛋白水平的表达。与药物敏感 HL-60 细胞相比，HL-60/DR 细胞在 mRNA 和蛋白水平上过度表达 NRF2，下调 KEAP1 蛋白。分析了 NRF2 抑制剂对多柔比星耐药 HL-60/DR 细胞活力、凋亡和细胞内活性氧（ROS）水平的影响。我们观察到 NRF2 抑制剂显著增强了多柔比星耐药 HL-60/DR 细胞对多柔比星的敏感性，这与细胞内 ROS 水平的增加和的表达有关，提示参与了线粒体依赖性凋亡途径。此外，还使用 ML-385 抑制剂研究了 NRF2-KEAP1 通路基因的表达。NRF2 基因和蛋白表达保持不变；然而，我们注意到 ML-385 处理后 KEAP1 蛋白下调。此外，ML-385 处理后，NRF2 调节的抗氧化和解毒基因包括、、和的表达得以维持。总之，我们的研究结果表明，所有研究的抑制剂，即 K67、维甲酸和 ML-385，均增强了多柔比星在多柔比星耐药 HL-60/DR 细胞中的疗效，并为使用 NRF2 抑制剂和多柔比星克服白血病多柔比星耐药性的联合治疗提供了一种潜在策略。
