Cancer Biology Program, Fox Chase Cancer Center, Philadelphia, Pennsylvania.
Genomics Facility, Fox Chase Cancer Center, Philadelphia, Pennsylvania.
Clin Cancer Res. 2021 Feb 15;27(4):1200-1213. doi: 10.1158/1078-0432.CCR-18-3683. Epub 2020 Nov 17.
Receptor-interacting protein kinase 3 (RIPK3) phosphorylates effector molecule MLKL to trigger necroptosis. Although RIPK3 loss is seen in several human cancers, its role in malignant mesothelioma is unknown. This study aimed to determine whether RIPK3 functions as a potential tumor suppressor to limit development of malignant mesothelioma.
RIPK3 expression was examined in 66 malignant mesothelioma tumors and cell lines. Promoter methylation and siRNA studies were performed to assess the mode of silencing in RIPK3-deficient malignant mesothelioma cells. Restoration of RIPK3 expression in RIPK3-negative malignant mesothelioma cells, either by treatment with 5-aza-2'-deoxycytidine or lentiviral expression of cDNA, was performed to assess effects on cell viability, necrosis, and chemosensitization.
Loss of RIPK3 expression was observed in 42/66 (63%) primary malignant mesotheliomas and malignant mesothelioma cell lines, and RT-PCR analysis demonstrated that downregulation occurs at the transcriptional level, consistent with epigenetic silencing. RIPK3-negative malignant mesothelioma cells treated with 5-aza-2'-deoxycytidine resulted in reexpression of RIPK3 and chemosensitization. Ectopic expression of RIPK3 also resulted in chemosensitization and led to necroptosis, the latter demonstrated by phosphorylation of downstream target MLKL and confirmed by rescue experiments. Mining of expression and survival outcomes among patients with malignant mesothelioma available from The Cancer Genome Atlas repository revealed that promoter methylation of is associated with reduced expression and poor prognosis.
These data suggest that RIPK3 acts as a tumor suppressor in malignant mesothelioma by triggering necroptosis and that epigenetic silencing of by DNA methylation impairs necroptosis and contributes to chemoresistance and poor survival in this incurable disease.
受体相互作用蛋白激酶 3（RIPK3）磷酸化效应分子 MLKL 以引发坏死性凋亡。尽管在几种人类癌症中观察到 RIPK3 的缺失，但它在恶性间皮瘤中的作用尚不清楚。本研究旨在确定 RIPK3 是否作为一种潜在的肿瘤抑制因子发挥作用，以限制恶性间皮瘤的发展。
检查了 66 例恶性间皮瘤肿瘤和细胞系中的 RIPK3 表达。进行启动子甲基化和 siRNA 研究，以评估 RIPK3 缺陷型恶性间皮瘤细胞中沉默的模式。通过用 5-氮杂-2'-脱氧胞苷处理或慢病毒表达 cDNA，恢复 RIPK3 阴性恶性间皮瘤细胞中的 RIPK3 表达，以评估对细胞活力、坏死和化疗增敏的影响。
在 66 例原发性恶性间皮瘤和恶性间皮瘤细胞系中观察到 RIPK3 表达缺失，RT-PCR 分析表明下调发生在转录水平，与表观遗传沉默一致。用 5-氮杂-2'-脱氧胞苷处理的 RIPK3 阴性恶性间皮瘤细胞导致 RIPK3 的重新表达和化疗增敏。RIPK3 的异位表达也导致化疗增敏，并导致坏死性凋亡，这可以通过下游靶标 MLKL 的磷酸化来证明，并通过挽救实验来证实。从癌症基因组图谱存储库中挖掘到的恶性间皮瘤患者的表达和生存结果表明， 的启动子甲基化与 表达降低和预后不良相关。
这些数据表明，RIPK3 在恶性间皮瘤中作为肿瘤抑制因子发挥作用，通过触发坏死性凋亡，而 DNA 甲基化对 的表观遗传沉默会损害坏死性凋亡，并导致这种无法治愈的疾病中的化疗耐药性和预后不良。
