摘要
目的:建立一种敏感性高且能定量分析ABL1激酶区突变的检测方法,为慢性髓系白血病(CML)的早期诊断和治疗提供有力支持。方法:收集35例经Sanger测序法检测为阴性的CML患者剩余标本。ABL1激酶区突变检测采用上海源奇生物医药科技有限公司的突变检测试剂盒采用^(△△)Ct值法分析突变率通过计算突变率并除以融合基因表达量,得出ABL1激酶区的相对突变率。结果:在35例经Sanger测序法检测为阴性患者中重新采用荧光定量PCR法检测ABL1激酶区突变,有7例出现T315I突变2例T315A突变2例Y253H突变,1例E255K突变,其相对突变率在0.1%-19.42%之间,均属于低频突变,超出了Sanger测序法的检测范围。随后应用该方法对126例CML患者进行ABL1突变检测与Sanger测序法相比,该方法的阳性率提高,显示出更高的敏感性。针对患者的突变情况进行相应的临床治疗调整后,观察到患者BCR-ABL1融合基因表达显著性下降或转为阴性。结论:与Sanger测序法相比,荧光定量PCR法在检测ABL1激酶区突变方面具有更高敏感性,能早期筛选出低频ABL1激酶区突变,并进行相对定量分析,该方法具有较好的临床应用前景。
Objective:To establish a highly sensitive and quantitative detection method for ABLI kinase region mutations,provide strong support for the early diagnosis and treatment of chronic myeloid leukemia(CML).Methods:Sampele from 35 CML patients who were initially tested negative for ABLI kinase region mutations by Sanger sequencing were collected.The ABLI kinase region mutation was detected by the fluorescence quantitative detection kit of Shanghai Yuanqi Biopharmaceutical Technology Co.,Ltd.The mutation rate was analyzed by ^(△△)Ct value method.The relative mutation rate of the final ABL1 kinase region was determined by dividing the mutation rate by the expression level of the fusion gene.Results:Among the 35 CML patients initially tested negative for ABLI mutations by the Sanger sequencing method,7 cases of T315I mutation,2 cases of T315A mutation,2 cases of Y253H mutation,and 1 cases of E255K mutation after detection of the new method.The relative mutation rates range from 0.1%to 19.42%,which could not be detected by Sanger sequencing method.Subsequently,this method was used to detect the ABLI mutation in 126 CML patients,and the positive rate exceeded that of the Sanger sequencing method.The BCR-ABLI gene expression significantly reduced or negative after adjusting treatment strategy based on the mutation situation.Conclusion:Compared with Sanger sequencing,fluorescence quantitative PCR has higher sensitivity and can screen for low-frequency ABLI kinase mutations in the early stage.Moreover,it can also perform relative quantitative analysis,so the method has good clinical application prospects for detecting ABLl mutation.
作者
程焕臣
李思
王殿智
刘宇
贡铁军
马军
CHENG Huan-Chen;LI Si;WANG Dian-Zhi;LIU Yu;GONG Tie-Jun;MA Jun(Institute of Harbin Hematology&Oncology,Harbin First Hospital,Harbin 150010,Heilongjiang Province,China)
出处
《中国实验血液学杂志》
CAS
CSCD
北大核心
2024年第5期1377-1380,共4页
Journal of Experimental Hematology
基金
黑龙江省中医药局项目(ZHY2022-006)
黑龙江省自然科学基金(LH2019C038)。
关键词
荧光定量PCR
基因
ABL1
耐药
CML
fluorescence quantitative PCR
gene
ABL1
drug resistance
chronic myeloid leukemia
