BCR-ABL1阴性骨髓增殖性肿瘤CALR和MPL及JAK2突变表达临床研究

Expression of CALR,MPL and JAK2 mutations in BCR-ABL1-negative myeloproliterative neoplasms

目的 BCR-ABL1阴性骨髓增殖性肿瘤(myeloproliferative neoplasms,MPN)主要包括真性红细胞增多症(polycythemia vera,PV)、原发性血小板增多症(essential thrombocythaemia,ET)及原发性骨髓纤维化(primary myelofibrosis,PMF),JAK2、CALR和MPL突变在上述疾病的发病机制中有重要作用。本研究探讨CALR、MPL和JAK2突变在BCR-ABL1阴性MPN患者中的发生率,分析CALR及MPL突变阳性MPN患者的临床特点。方法应用2008年WHO造血组织和淋巴组织肿瘤分类标准,将河北医科大学第二医院2014-07-01-2017-07-01初次就诊的115例BCR-ABL1阴性MPN患者分为PV组、ET组和PMF组。采用直接测序法检测所有患者骨髓CALR、MPL及JAK2中12号外显子突变情况,采用等位基因特异性聚合酶链反应(allele specific polymerase chain reaction,AS-PCR)检测所有患者骨髓JAK2V617F突变情况,分析CALR和MPL突变阳性MPN患者的临床特征及其实验室检查特点。结果 115例BCR-ABL1阴性MPN患者中,PV 31例,其中28例(90.32%)检测到JAK2V617F突变,2例(6.45%)检测到JAK2的12号外显子突变,均未检测出CALR和MPL突变,CALR、MPL和JAK2突变均阴性PV仅1例(3.26%);ET 52例,其中33例(63.46%)检测到JAK2V617F突变,8例(15.38%)检测到CALR突变,1例(1.92%)同时检测到CALR和JAK2V617F突变,1例(1.92%)检测到MPL突变,CALR和MPL突变在JAK2V617F突变阴性ET中的发生率分别为42.11%(8/19)和5.26%(1/19),CALR、MPL和JAK2突变均阴性ET为10例(19.23%);PMF 32例,其中24例(75.00%)检测出JAK2V617F突变,5例(15.62%)检测出CALR突变,2例(6.25%)检测到MPL突变,在JAK2V617F突变阴性的PMF中CALR和MPL的突变率为分别62.50%(5/8)和25.00%(2/8),CALR、MPL和JAK2突变均阴性PMF为1例(3.13%)。115例患者中,无同时检测到JAK2、CALR和MPL突变患者,提示这3种基因突变不同时出现。ET患者中,CALR突变阳性患者危险度分层及血管事件发生率低于JAK2V617F突变阳性患者;PMF患者中,CALR突变阳性患者血小板计数高于JAK2V617F突变阳性患者,血红蛋白浓度低于JAK2V617F突变阳性患者,有更好的生存率。结论 CALR突变在JAK2/MPL阴性的MPN中有临床意义,扩大了MPN的分子学诊断范围,为MPN的诊断及治疗带来了新思路。BCR-ABL1阴性的MPN患者中,CALR突变阳性患者与JAK2V617F突变阳性患者相比,发生血管事件风险及危险度分层更低、发病年龄小,提示预后较好。

OBJECTIVE BCR-ABLl-negative myeloproliferative neoplasm（MPNs） mainly include polycythemia ver- a（PV）, essential thrombocythaemia（ET） and primary myelofibrosis（PMF）, JAK2, CALR and MPL mutations play im- portant roles in the pathogenesis of these diseases. This study aimed to investigate the mutational frequencies of CALR, MPL and JAK2 in patients with BCR-ABLl-negative MPNs, to analyze the clinical characteristics in MPN patients with CALR and MPL mutations. METHODS By using the WHO 2008 classification criteria of hematopoietic and lymphoid tissue tumor, 115 MPNs patients diagnosed in our hospital between July 2014 to July 2017 were classified as PV group, ET group and PMF group. Direct DNA sequencing samples from bone marrow was used to detect the mutations of the CALR,MPL and exon 12 of JAK2 gene in all patients, and JAK2 V617F mutations was detected by allele specific poly merase chain reaction（AS-PCR）. The clinical features of MPNs patients with CALR and MPL mutations and their labora- tory characteristics were analyzed. RESULTS In the 115 patients with gCR-ABLl-negative MPNs, there were 31 PV cases, in which 28 cases （90.32%） were detected JAK2V617F mutation, 2 cases （6.45%） were detected mutations in exon 12 of JAK2, none was detected CALR or MPL mutations, and only 1 PV case （3. 26%） was all negative for CALR, MPL and JAK2 mutations. There were 52 ET cases, in which 33 cases （63.46%） were detected JAK2V617F mutation, 8 cases （15.38%） were detected CALR mutation, and one case （1. 92%） was detected CALR and JAK2V617F muta- tions, simultaneously. One case （1. 92%） was detected MPL mutation. The incidences of CALR, MPL mutations in JAK2V617F-negative ET were 42.11%（8/19）,5.26%（1/19）, and 10 ET case （19.23%） were all negative for CALR, MPL and JAK2 mutations. There were 32 cases with PMF, in which 24 cases （75.00%） were detected JAK2V617F mu- tation, 5 cases （15.62%） were detected CALR mutation, 2 cases （6.25%） were detected MPL mutation, and in patients with JAK2V617F-negative PMF,the incidences of CALR,MPL mutation were 62.50% （5/8）, 25.00%（2/8）. Only one case （3.13%） with PMF were all negative for CALR, MPL and JAK2 mutations. None of the 115 patients was detected JAK2,CALR and MPL mutation at the same time, suggesting that the three mutations did not occur simultaneously. The risk stratification and the incidence of vascular events in ET patients with CALR mutation was lower than that with JAK2V617F mutation. The platelet count of PMF patients with CALR mutation was higher than that with JAK2V617F mutation, and the hemoglobin concentration was lower in PMF patients with CALR mutation, but with better overall survival. CONCLUSIONS CALR mutation possesses clinical significance in JAK2/MPL-negative MPNs, extends the molecular diagnostic scope of MPNs, and brings new ideas to the diagnosis and treatment of MPNs. Among tgCR-AgLl-nega- tire MPNs patients, compared with JAK2V617F mutation, patients with CAI.R mutation take on lower risk of vascular events, lower risk stratification, more young age, and better prognosis.