Detection of leukemia gene fusions on DNA-level through targeted Next-Generation Sequencing

Background: Targeted Next-Generation Sequencing (tNGS) is commonly used to detect genetic mutations in patients with leukemia on DNA-level, necessitating additional methods to confirm genomic rearrangements and gene fusions.Methods: A total of 357 adults diagnosed with leukemia were included in the study. The tNGS panel includes all exons from 302 genes closely associated with leukemia and 95 introns from 26 genes, facilitating the detection of gene fusion alterations within these specific regions. Additionally, the common breakpoint regions of IGH and MYC are employed for the detection of IGH or MYC rearrangements. Commercial quantitative reverse transcription polymerase chain reaction (qRT-PCR) reagents were used simultaneously to detect 45 gene fusions in RNA samples.Results: The qRT-PCR method detected a total of 102 gene fusions, encompassing 23 distinct types. The tNGS method identified gene fusions on DNA-level in 98 samples, achieving a Positive Percent Agreement (PPA) of 96.1% (98/102) when compared to the qRT-PCR method, and no false positive findings. Additionally, it revealed the presence of two gene fusions, KMT2A::ELL and KMT2A::MLLT3, which had gone undetected by qRT-PCR. The tNGS can also identify IGH or MYC gene rearrangements in patients with B-ALL, achieving a PPA of 93.8% (15/16) when compared to the FISH. Moreover, tNGS can accurately identify the specific partner genes associated with these rearrangements, facilitating a more precise analysis of the impact of mutations on prognosis.Conclusion: This study demonstrates the feasibility of employing tNGS methods to concurrently identify gene mutations and fusions (including IGH and MYC rearrangements) at the DNA level in adults with leukemia.