A multicenter comparison study on the detection of BCR-ABL tyrosine kinase domain point mutation

目的 调查不同医院BCR-ABL酪氨酸激酶区点突变检测的准确性和一致性.方法 由6家医院各制备10份来自于酪氨酸激酶抑制剂耐药的BCR-ABL(+)患者骨髓或外周血样本的cDNA,每份分装成6管派发,每家医院按照自身的操作程序检测共60份比对样本的BCR-ABL激酶区点突变,由北京大学人民医院结合测序图谱进行结果比对分析.结果 37份(61.7％)比对样本的碱基及相应氨基酸突变类型6家医院报告均一致.53份样本可以确定突变类型,共涵盖23种;1份样本没有突变;6份样本由于室间图谱差异较大无法明确结果.突变比例低造成12份样本结果不一致,二者明显相关(P=0.008);测序图谱特殊及扩增区域未覆盖各造成1份样本结果不一致;3份样本发生扩增失败;7份样本发生检测或报告错误.80.6％的结果明确的样本室间报告突变比例差值在20％以内.标本内参弱与检测失败和室间测序图谱差异大均相关.结论 通过样本比对能够发现临床常规检测存在的问题,促进检测水平的提高.突变比例低是导致室间突变报告不一致的主要因素.Objective To investigate the accuracy and consistency of the detection of BCR-ABL tyrosine kinase domain point mutation among different laboratories.Methods Every one of 6 laboratories prepared 10 cDNA samples from tyrosine kinase inhibitors resistant BCR-ABL (P210 or P190) positive patients' bone marrow or peripheral blood.Each cDNA sample was divided into 6 aliquots and delivered to the laboratories.All 6 laboratories tested BCR-ABL point mutations of 60 samples according to their own protocols.Peking University People' s Hospital analyzed the comparison results based on both the reports and sequencing chromatogram from all laboratories.Results All laboratories reported the same nucleotide and corresponding amino acid mutations in 37 samples (61.7％).Of 60 samples,53 had confirmed mutation types,and a total of 23 types were included;1 had no mutation;mutation types of 6 samples could not be determined because of the big differences among chromatograms from different laboratories.Low percentages of mutants were significantly related to results inconsistency (P=0.008).Inconsistent result of one sample was caused by the unique chromatogram of the mutant L248V,and one by the non-coverage amplification of PCR product from different laboratories.Amplification was failed in 3 samples.Testing or sequencing mistakes occurred in 7 samples.The differences in the mutant percentages among laboratories were less than 20％ in the 80.6％ of samples with confirmed results.Low internal control gene copies (ABL＜10 000) were significantly related to both failed amplification and big differences among chromatograms from different laboratories (P=0.005 and ＜0.001,respectively).Conclusion Problems in the clinical routine detection of BCR-ABL point mutation could be exposed and improvement could be achieved by sample exchange and comparison.Low percentage of mutant is the main reason which causes the discrepancy of BCR-ABL point mutation results among different laboratories.PubMed中文核心期刊要目总览(PKU)中国科技核心期刊(ISTIC)中国科学引文数据库(CSCD)11902-9053