Establishment of a cost-effective method to detect FLT-ITD and D835 mutations in acute myeloid leukemia patients in the Taiwanese population

Objective: The FMS-related tyrosine kinase 3 (FLT-3) gene is a hematopoietic growth factor receptor gene, an independent negative prognostic factor, which affects the proliferation and differentiation of stem cells or hematopoietic progenitor cells. Patients with FLT-3 gene mutations have a worse prognosis and responsiveness to chemotherapy than those without these mutations. Our study aims to establish a conventional detection method for FLT3-ITD and D835 mutations in patients with acute myeloid leukemia (AML). Materials and methods: In this study, we recruited 100 patients with AML. Primers were designed to distinguish between wild-type FLT-3, FLT3-ITD, and D835 variants. Methods using a polymerase chain reaction (PCR)-Agilent 2100 Bioanalyzer, PCR-ABI PRISM 3100 Genetic Analyzer, and PCR-agarose gel electrophoresis were compared. Results: A high-accuracy, easily operated, low-cost technique to detect the FLT-3 variation with 99.9% specificity was established in this study. The PCR platform, the Agilent 2100 Bioanalyzer (plus DNA 1000 LabChip kit) chip analysis platform, and the ABI PRISM 3100 Genetic Analyzer (plus GeneScan-500 size standard) short tandem repeat (STR) fluorescence analysis platform were used in different experimental comparisons. The ABI PRISM 3100 Genetic Analyzer (plus GeneScan-500 size standard) STR fluorescence analysis platform was the most suitable method to detect FLT3 variants. This method has a high degree of sensitivity, accuracy, and a specificity of 99.9%. Conclusion: AML with the homozygous mutated FLT-3 may have a worse cure rate than AML with heterozygous mutation. This mutation is not related to drug resistance, but is a factor in a high risk of relapse; it is also related to unfavorable overall survival. Our designed detection methods should provide key information to develop personalized medicine for AML patients.