Feasible and Rapid Screening of IDH1/2 and FLT3-TKD2 Mutations by High-Resolution Melting for Patients with Acute Myeloid Leukemia.

Background

In recent years, numerous recurrently mutated genes have been identified in acute myeloid leukemia (AML), some of which, such as FLT3 and IDH1/2, serve as therapeutic targets, offering new treatment options. Rapid mutational analysis is crucial for timely and optimal therapy selection. This study aims to develop and validate a rapid, cost-effective, and sensitive screening method for detecting IDH1, IDH2, and FLT3-TKD2 mutations using polymerase chain reaction (PCR) and high-resolution melting curve analysis (HRM).

Methods

A PCR-HRM assay was developed to simultaneously detect mutations in IDH1, IDH2, and FLT3-TKD2. The method was applied to a cohort of 1363 AML patients, and its performance, including turnaround time, was evaluated through comparison with next-generation sequencing (NGS) results.

Results

The PCR-HRM method demonstrated a positive percent agreement of 98%, 98%, and 92% for IDH1, IDH2, and FLT3-TKD2, respectively, and a negative percent agreement of 100% for all three genes compared to NGS. No false positives were observed, and false negatives were detected in less than 1% of cases, mostly in FLT3-TKD2, all occurring below the established limit of detection. The turnaround time and cost of PCR-HRM were significantly lower than those of NGS.

Conclusions

This method offers a highly sensitive, specific, and time-efficient approach for the simultaneous detection of IDH1, IDH2, and FLT3-TKD2 mutations in AML patients. Its rapid turnaround time and cost-effectiveness make it a valuable tool for routine clinical screening, facilitating timely and targeted treatment decisions.