Valiulienė Giedrė, Vitkevičienė Aida, Navakauskienė Rūta
Department of Molecular Cell Biology, Institute of Biochemistry, Life Sciences Center, Vilnius University, Sauletekio av. 7, LT-01257 Vilnius, Lithuania.
Eur J Pharmacol. 2020 Dec 15;889:173641. doi: 10.1016/j.ejphar.2020.173641. Epub 2020 Oct 9.
Although majority of acute promyelocytic leukemia (APL) patients achieve complete remission after the standard treatment, 5-10% of patients are shown to relapse or develop resistance to treatment. In such cases, medications that target epigenetic processes could become an appealing supplementary approach. In this study, we tested the anti-leukemic activity of histone deacetylase inhibitor Belinostat (PXD101) and histone methyltransferase inhibitor 3-Deazaneplanocin A combined with all-trans retinoic acid in APL cells NB4, promyelocytes resembling HL-60 cells and APL patients' cells. After HL-60 and NB4 cell treatment, ChIP-sequencing was performed using antibodies against hyper-acetylated histone H4. Hyper-acetylated histone H4 distribution peaks were compared in treated vs untreated HL-60 and NB4 cells. Results demonstrated that in treated HL-60 cells, the majority of peaks were distributed within the regions of proximal promoters, whereas in treated NB4 cells, hyper-acetylated histone H4 peaks were mainly localized in gene body regions. Further ChIP-seq data analysis revealed the changes in histone H4 hyper-acetylation in promoter/gene body regions of genes involved in cancer signaling pathways. In addition, quantitative gene expression analysis proved changes in various cellular pathways important for carcinogenesis. Epigenetic treatment down-regulated the expression of MTOR, LAMTOR1, WNT2B, VEGFR3, FGF2, FGFR1, TGFA, TGFB1, TGFBR1, PDGFA, PDGFRA and PDGFRB genes in NB4, HL-60 and APL patients' cells. In addition, effect of epigenetic treatment on protein expression of aforementioned signaling pathways was confirmed with mass spectrometry analysis. Taken together, these results provide supplementary insights into molecular changes that occur during epigenetic therapy application in in vitro promyelocytic leukemia cell model.
尽管大多数急性早幼粒细胞白血病（APL）患者在标准治疗后可实现完全缓解，但仍有5%-10%的患者会复发或产生治疗耐药性。在这种情况下，针对表观遗传过程的药物可能成为一种有吸引力的辅助治疗方法。在本研究中，我们测试了组蛋白去乙酰化酶抑制剂贝利司他（PXD101）和组蛋白甲基转移酶抑制剂3-去氮杂平诺菌素A与全反式维甲酸联合使用对APL细胞NB4、类似HL-60细胞的早幼粒细胞以及APL患者细胞的抗白血病活性。对HL-60和NB4细胞进行处理后，使用抗高乙酰化组蛋白H4抗体进行染色质免疫沉淀测序（ChIP-seq）。比较了处理过的和未处理的HL-60及NB4细胞中高乙酰化组蛋白H4的分布峰。结果表明，在处理过的HL-60细胞中，大多数峰分布在近端启动子区域，而在处理过的NB4细胞中，高乙酰化组蛋白H4峰主要定位在基因体区域。进一步的ChIP-seq数据分析揭示了参与癌症信号通路的基因启动子/基因体区域中组蛋白H4高乙酰化的变化。此外，定量基因表达分析证明了对致癌作用重要的各种细胞途径发生了变化。表观遗传治疗下调了NB4、HL-60和APL患者细胞中MTOR、LAMTOR1、WNT2B、VEGFR3、FGF2、FGFR1、TGFA、TGFB1、TGFBR1、PDGFA、PDGFRA和PDGFRB基因的表达。此外，通过质谱分析证实了表观遗传治疗对上述信号通路蛋白表达的影响。综上所述，这些结果为体外早幼粒细胞白血病细胞模型中表观遗传治疗应用过程中发生的分子变化提供了补充见解。
