核型正常髓系肿瘤患者的基因突变研究

Characterizing the genomic landscape in myeloid malignancies with normal karyotype

目的:研究正常核型的髓系肿瘤患者的基因突变谱系。方法:回顾性检测2013年6月—2017年8月于南京医科大学附属常州第二人民医院及苏州大学附属第一医院门诊及住院治疗的正常核型的原发急性髓系白血病(acute myelogenous leukemia,AML)及骨髓增生异常综合征(myelodysplastic syndrome,MDS)患者共102例。采用第二代基因测序技术检测49种靶基因;采用Sanger测序法检测FLT3-ITD、NPM1基因12号外显子、CALR基因9号外显子及CEBPA基因的TAD、BZIP功能结构域突变发生情况。结果:(1)82例AML患者中,总突变发生率为98.8%(81/82),其中≥3个基因突变共存发生率为52.4%(43/82)。突变检出率最高为NPM1(35.4%,29/82),其他突变率>10%的基因依次为FLT3-ITD(25.6%,21/82)、CEBPA双突变(24.4%,20/82)、DNMT3A(19.5%,16/82)、TET2(18.3%,15/82)、NRAS(13.4%,11/82)、RUNX1(11.0%,9/82)、CSF3R(11.0%,9/82)。(2)20例MDS患者的总突变率为90%(18/20),其中≥3个基因突变共存发生率为55.0%(11/20)。突变检出率最高为RUNX1(35.0%,7/20),其他突变依次为ASXL1(25.0%,5/20)、SF3B1(15.0%,3/20)、BCOR(15.0%,3/20)、FLT3-TKD(15.0%,3/20)。(3)AML患者总的基因突变率与MDS患者相似(P=0.097)。AML患者中,NPM1、CEBPA双突变在AML中的发生率明显高于MDS患者(P<0.05)。MDS患者基因突变主要为RUNX1、ASXL1、SF3B1及BCOR为主,发生率明显高于AML患者(P<0.05)。(4)功能归类后显示,DNA甲基化调节基因及酪氨酸激酶受体基因突变在AML中的发生率明显高于MDS患者(P<0.05),而组蛋白修饰基因及RNA剪接因子突变在MDS中的发生率明显高于AML(P<0.05)。结论:正常核型的髓系肿瘤患者体内存在多个不同功能基因突变组合的亚克隆,原发AML与MDS在基因突变谱上有很大不同。

Objective:This study aims to characterize the gene mutation in myeloid malignancies with normal karyotype by exploring multiple gene mutations.Methods:Tatal 102 acute myelogenous leukemia(AML)with normal karyotype and myelodysplastic syndrome(MDS)paticnts were retrospectively analyzed in Changzhou Second Affiliated Hospital of Nanjing Medical Univcrsity and the First Affiliated Hospital of Suzhou University.Targeted second generation sequencing were performed using a custom-designed 49-gene panel.CALR,FLT3 internal tandem duplication(FLT3-ITD),NPM1 and CEBPA mutation were detected by Sanger sequencing.Results:(1)Together,gene mutations accounted for a considerable frequency of 98.8%AML patients.Coexistence of≥3 mutations was identified in 52.4%patients.The most commonly mutated gene was NPM1(35.4%),followed by FLT3-ITD(25.6%),CEBPA double mutations(24.4%),DNMT3 A(19.5%),TET2(18.3%),NRAS(13.4%),RUNX1(11.0%)and CSF3R(11.0%)mutations.(2)The gene mutations were present in 90%MDS patients.Coexistence of≥3 mutations was in 55.0%patients.The most commonly mutated gene was RUNX1(35.0%),followed by ASXL1(25.0%),SF3B1(15.0%),FLT3-TKD(15.0%)and BCOR(15.0%)mutations.(3)The incidence of mutation was similar in AML patients and MDS patients(P=0.097).The mutation rate of each of NPM1 and CEBPA double mutations was higher in patients with AML(P<0.05),while RUNX1,ASXL,SF3B1 and BCOR mutations were identified more frequently in MDS patients,both differences were significant(P<0.05).(4)Gene aberrations involved in DNA methylation(DNMT3A,TET2,IDH1/2)and receptors/kinases(FLT3-ITD,FLT3-TKD,JAK1,JAK2,JAK3,c-KIT,PDGFRA,PDGFRB,MPL,CSF3R,NOTCH1,IL7R)significantly predominated in AML while post-translational chromatin modification(EZH2,ASXL1/2,SETD2)and RNA splicing(SRSF2,SF3B1,ZRSR2,U2AF1)significantly predominated in MDS(P<0.05).Conclusion:Different functional mutation combinations revealed multiple sub-clones in myeloid malignancies with normal karyotype.The genomic landscape of AML patients was different from MDS patients.