Background/Objectives:Chronic lymphocytic leukemia (CLL) remains an incurable B-cell malignancy. B-CLL cells exhibit an extended lifespan in part due to the activation of survival pathways such as NF-kB. A crosstalk between NF-kB and GSK-3β pathways has been reported. NF-kB has also been identified as a primary target of the NEDD8-activating enzyme inhibitor MLN4924. Our objective was to investigate potential synergies of MLN4924 with other NF-kB-targeting agents for the treatment of CLL and elucidate the mechanisms of action underlying this pathway regulation.Methods:To assess the cytotoxic efficacy of the combined ex vivo treatment with CHIR-99021 and MLN4924, we employed 7-AAD staining and XTT viability assays on primary samples from CLL patients. Subsequently, we conducted various analyses to identify the molecular mechanisms underlying the cytotoxic effects of this combination.Results:We discovered a discrepancy between the mRNA and protein levels of IkBɑ and provided evidence of translational control over its expression. This observation may explain why, unlike other cell types, B-CLL cells did not activate NF-kB signaling following inhibition of GSK-3ß. Furthermore, we describe a synergistic effect between a specific GSK-3ß inhibitor, CHIR-99021/Laduviglusib, and the NEDD8-activating enzyme inhibitor MLN4924/Pevonedistat, at doses that only slightly affect healthy B cell viability ex vivo. We investigated the molecular basis of this co-induction of cell death by analyzing the alterations in apoptosis-related gene expression. We found that the combinational treatment enhances a reduction in BCL2 mRNA expression levels, providing an alternative approach for BCL-2 inhibition in CLL that could have therapeutic implications for the treatment of refractory CLL cases.Conclusions:our findings revealed a unique interaction between GSK-3ß and NF-kB pathways in CLL and their regulation of BCL2 expression.
背景/目的：慢性淋巴细胞白血病（CLL）仍是一种无法治愈的B细胞恶性肿瘤。B-CLL细胞寿命延长部分归因于NF-kB等生存通路的激活。已有研究报道NF-kB与GSK-3β通路存在交互作用，且NF-kB被确认为NEDD8活化酶抑制剂MLN4924的主要作用靶点。本研究旨在探讨MLN4924与其他靶向NF-kB药物联合治疗CLL的潜在协同效应，并阐明该通路调控的作用机制。 方法：为评估CHIR-99021与MLN4924联合离体治疗的细胞毒性效应，我们对CLL患者原代样本进行7-AAD染色和XTT活力检测。随后通过多种分析手段探究该联合治疗方案产生细胞毒性作用的分子机制。 结果：研究发现IkBɑ的mRNA与蛋白表达水平存在差异，并证实其表达受翻译水平调控。这一发现可能解释为何与其他细胞类型不同，B-CLL细胞在GSK-3ß抑制后未激活NF-kB信号通路。此外，我们观察到特异性GSK-3ß抑制剂CHIR-99021/Laduviglusib与NEDD8活化酶抑制剂MLN4924/Pevonedistat在仅轻微影响健康B细胞离体存活率的剂量下产生协同效应。通过分析凋亡相关基因表达变化，我们探讨了该联合方案诱导细胞死亡的分子基础，发现联合治疗能增强BCL2 mRNA表达水平的降低，这为CLL的BCL-2抑制提供了新策略，可能对难治性CLL病例的治疗具有临床意义。 结论：本研究揭示了CLL中GSK-3ß与NF-kB通路独特的相互作用及其对BCL2表达的调控机制。
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