Yang Jun-Jun, Chen Hui, Zheng Xiao-Qun, Li Hai-Ying, Wu Jian-Bo, Tang Li-Yuan, Gao Shen-Meng
Department of Laboratory Medicine, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, China E-mail :
Asian Pac J Cancer Prev. 2015;16(6):2219-25. doi: 10.7314/apjcp.2015.16.6.2219.
SHP1 negatively regulates the Janus kinase 2/signal transducer and activator of transcription (JAK2/STAT) signaling pathway, which is constitutively activated in myeloproliferative neoplasms (MPNs) and leukemia. Promoter hypermethylation resulting in epigenetic inactivation of SHP1 has been reported in myelomas, leukemias and other cancers. However, whether SHP1 hypermethylation occurs in MPNs, especially in Chinese patients, has remained unclear. Here, we report that aberrant hypermethylation of SHP1 was observed in several leukemic cell lines and bone marrow mononuclear cells from MPN patients. About 51 of 118 (43.2%) MPN patients including 23 of 50 (46%) polycythaemia vera patients, 20 of 50 (40%) essential thrombocythaemia and 8 of 18 (44.4%) idiopathic myelofibrosis showed hypermethylation by methylation-specific polymerase chain reaction. However, SHP1 methylation was not measured in 20 healthy volunteers. Hypermethylation of SHP1 was found in MPN patients with both positive (34/81, 42%) and negative (17/37, 45.9%) JAK2V617F mutation. The levels of SHP1 mRNA were significantly lower in hypermethylated samples than unmethylated samples, suggesting SHP1 may be epigenetically inactivated in MPN patients. Furthermore, treatment with 5-aza-2'-deoxycytidine (AZA) in K562 cells showing hypermethylation of SHP1 led to progressive demethylation of SHP1, with consequently increased reexpression of SHP1. Meanwhile, phosphorylated JAK2 and STAT3 were progressively reduced. Finally, AZA increased the expression of SHP1 in primary MPN cells with hypermethylation of SHP1. Therefore, our data suggest that epigenetic inactivation of SHP1 contributes to the constitutive activation of JAK2/STAT signaling. Restoration of SHP1 expression by AZA may contribute to clinical treatment for MPN patients.
SHP1对Janus激酶2/信号转导及转录激活因子（JAK2/STAT）信号通路起负向调节作用，该信号通路在骨髓增殖性肿瘤（MPN）和白血病中呈组成性激活。在骨髓瘤、白血病及其他癌症中，已报道启动子高甲基化导致SHP1表观遗传失活。然而，SHP1高甲基化是否发生在MPN中，尤其是中国患者中，仍不清楚。在此，我们报告在几种白血病细胞系及MPN患者的骨髓单个核细胞中观察到SHP1异常高甲基化。118例MPN患者中有51例（43.2%）出现高甲基化，其中真性红细胞增多症患者50例中有23例（46%）、原发性血小板增多症患者50例中有20例（40%）、原发性骨髓纤维化患者18例中有8例（44.4%），采用甲基化特异性聚合酶链反应检测发现高甲基化。然而，未对20名健康志愿者检测SHP1甲基化情况。在JAK2V617F突变阳性（占81例中的34例，42%）和阴性（占37例中的17例，45.9%）的MPN患者中均发现SHP1高甲基化。高甲基化样本中SHP1 mRNA水平显著低于未甲基化样本，提示MPN患者中SHP1可能发生表观遗传失活。此外，在显示SHP1高甲基化的K562细胞中用5-氮杂-2'-脱氧胞苷（AZA）处理导致SHP1逐渐去甲基化，进而SHP1重新表达增加。同时，磷酸化JAK2和STAT3逐渐减少。最后，AZA使SHP1高甲基化的原发性MPN细胞中SHP1表达增加。因此，我们的数据表明SHP1的表观遗传失活导致JAK2/STAT信号通路的组成性激活。AZA恢复SHP1表达可能有助于MPN患者的临床治疗。
