Jiangsu Provincial Medical Innovation Center, Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210028, China.
The Quzhou Affiliated Hospital of Wenzhou Medical University, Quzhou People's Hospital, Quzhou, 324000, China.
Cell Commun Signal. 2024 Feb 12;22(1):116. doi: 10.1186/s12964-023-01367-y.
R140Q mutation in isocitrate dehydrogenase 2 (IDH2) promotes leukemogenesis. Targeting IDH2/R140Q yields encouraging therapeutic effects in the clinical setting. However, therapeutic resistance occurs in 12% of IDH2/R140Q inhibitor treated patients. The IDH2/R140Q mutant converted TF-1 cells to proliferate in a cytokine-independent manner. This study investigated the signaling pathways involved in TF-1(R140Q) cell proliferation conversion as alternative therapeutic strategies to improve outcomes in patients with acute myeloid leukemia (AML) harboring IDH2/R140Q.
The effects of IDH2/R140Q mutation on TF-1 cell survival induced by GM-CSF withdrawal were evaluated using flow cytometry assay. The expression levels of apoptosis-related proteins, total or phosphorylated STAT3/5, ERK, and AKT in wild-type TF-1(WT) or TF-1(R140Q) cells under different conditions were evaluated using western blot analysis. Cell viability was tested using MTT assay. The mRNA expression levels of GM-CSF, IL-3, IL-6, G-CSF, leukemia inhibitory factor (LIF), oncostatin M (OSM), and IL-11 in TF-1(WT) and TF-1(R140Q) cells were quantified via RT-PCR. The secretion levels of GM-CSF, OSM, and LIF were determined using ELISA.
Our results showed that STAT3 and STAT5 exhibited aberrant constitutive phosphorylation in TF-1(R140Q) cells compared with TF-1(WT) cells. Inhibition of STAT3/5 phosphorylation suppressed the cytokine-independent proliferation of TF-1(R140Q) cells. Moreover, the autocrine GM-CSF, LIF and OSM levels increased, which is consistent with constitutive STAT5/3 activation in TF-1(R140Q) cells, as compared with TF-1(WT) cells.
The autocrine cytokines, including GM-CSF, LIF, and OSM, contribute to constitutive STAT3/5 activation in TF-1(R140Q) cells, thereby modulating IDH2/R140Q-mediated malignant proliferation in TF-1 cells. Targeting STAT3/5 phosphorylation may be a novel strategy for the treatment of AML in patients harboring the IDH2/R140Q mutation. Video Abstract.
异柠檬酸脱氢酶 2（IDH2）中的 R140Q 突变可促进白血病发生。在临床环境中，针对 IDH2/R140Q 的治疗可产生令人鼓舞的治疗效果。然而，在接受 IDH2/R140Q 抑制剂治疗的患者中，有 12%会出现治疗抵抗。IDH2/R140Q 突变体使 TF-1 细胞能够在细胞因子非依赖性方式下增殖。本研究旨在探讨 IDH2/R140Q 突变体诱导 TF-1(R140Q)细胞增殖转化涉及的信号通路，以期为携带 IDH2/R140Q 突变的急性髓系白血病（AML）患者提供新的治疗策略，改善治疗结局。
通过流式细胞术检测 GM-CSF 撤去后 IDH2/R140Q 突变对 TF-1 细胞存活的影响。采用 Western blot 分析在不同条件下野生型 TF-1(WT)或 TF-1(R140Q)细胞中凋亡相关蛋白、总 STAT3/5 及磷酸化 STAT3/5、ERK 和 AKT 的表达水平。采用 MTT 法检测细胞活力。通过 RT-PCR 定量分析 TF-1(WT)和 TF-1(R140Q)细胞中 GM-CSF、IL-3、IL-6、G-CSF、白血病抑制因子（LIF）、抑瘤素 M（OSM）和白细胞介素 11（IL-11）的 mRNA 表达水平。采用 ELISA 法测定 GM-CSF、OSM 和 LIF 的分泌水平。
研究结果表明，与 TF-1(WT)细胞相比，TF-1(R140Q)细胞中 STAT3 和 STAT5 表现出异常的组成性磷酸化。抑制 STAT3/5 磷酸化可抑制 TF-1(R140Q)细胞的细胞因子非依赖性增殖。此外，自分泌 GM-CSF、LIF 和 OSM 水平增加，与 TF-1(R140Q)细胞中组成性 STAT5/3 激活一致，而与 TF-1(WT)细胞相比。
自分泌细胞因子，包括 GM-CSF、LIF 和 OSM，有助于 TF-1(R140Q)细胞中组成性 STAT3/5 的激活，从而调节 TF-1 细胞中 IDH2/R140Q 介导的恶性增殖。靶向 STAT3/5 磷酸化可能是治疗携带 IDH2/R140Q 突变的 AML 患者的一种新策略。
