目的:弥漫性大B细胞淋巴瘤(DLBCL)的初始治疗方法需要利妥昔单抗,环磷酰胺,阿霉素,长春新碱,和泼尼松(R-CHOP)方案。然而,40%的患者表现出欠佳的反应,有些人经历复发和难治性疾病。本研究旨在探索新的治疗策略并阐明其在DLBCL中的潜在机制。
方法:采用生物信息学技术来审查HDAC1,HDAC2,HDAC3,HDAC10,BTK,MYC,DLBCL中的TP53和BCL2基因。使用用西达本胺处理的DB和SU-DHL-4细胞进行体外实验,奥列拉布替尼,以及两者的结合。通过细胞计数试剂盒-8评估细胞活力。使用流式细胞术确定细胞凋亡和细胞周期。通过ROS和JC-1染色评估活性氧(ROS)的产生和线粒体功能。进行RNA测序和蛋白质印迹分析以阐明西达本胺和奥列拉布替尼在DLBCL细胞中联合作用的分子机制。
结果:本研究显示,西达胺与奥雷拉布替尼合用时,抗增殖作用明显增强。Compusyn软件分析显示西达本胺和奥列拉布替尼在抑制DLBCL细胞增殖方面具有协同作用,组合指数(CI)<1。这种协同作用进一步表现为增强的细胞周期停滞,凋亡诱导,细胞周期相关蛋白和抗凋亡蛋白的下调,和促凋亡蛋白的上调。此外,蛋白质印迹和RNA-Seq研究结果表明,西达胺和奥列拉布替尼联合调节PI3K/AKT/mTOR信号通路,从而促进DLBCL细胞周期阻滞和凋亡。
结论:这项研究的结果为临床使用西达本胺和奥雷拉布替尼治疗复发/难治性DLBCL提供了令人信服的理由。
OBJECTIVE: The initial therapeutic approach for diffuse large B-cell lymphoma (DLBCL) entails a rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) regimen. However, 40% of patients exhibit suboptimal responses, with some experiencing relapse and refractory conditions. This study aimed to explore novel therapeutic strategies and elucidate their underlying mechanisms in DLBCL.
METHODS: Bioinformatics techniques were employed to scrutinize correlations between the HDAC1, HDAC2, HDAC3, HDAC10, BTK, MYC, TP53, and BCL2 genes in DLBCL. In vitro experiments were conducted using DB and SU-DHL-4 cells treated with chidamide,
orelabrutinib, and a combination of both. Cell viability was assessed by cell counting kit-8. Cell apoptosis and the cell cycle were determined using flow cytometry. Reactive oxygen species (ROS) production and mitochondrial function were assessed through ROS and JC-1 staining. RNA sequencing and western blot analyses were conducted to elucidate the molecular mechanisms underlying the combined action of chidamide and
orelabrutinib in DLBCL cells.
RESULTS: This investigation revealed markedly enhanced antiproliferative effects when chidamide was combined with
orelabrutinib. Compusyn software analysis indicated a synergistic effect of chidamide and
orelabrutinib in inhibiting DLBCL cell proliferation, with a combination index (CI) < 1. This synergy further manifested as augmented cell cycle arrest, apoptosis induction, the downregulation of cell cycle-associated and antiapoptotic proteins, and the upregulation of proapoptotic proteins. Furthermore, the western blot and RNA-Seq findings suggested that combining chidamide and
orelabrutinib modulated the PI3K/AKT/mTOR signaling pathway, thereby promoting DLBCL cell cycle arrest and apoptosis.
CONCLUSIONS: The findings of this study provide a compelling justification for the clinical utilization of chidamide and orelabrutinib to treat relapsed/refractory DLBCL.