An atypical RUNX1::ETV6::RUNX1 fusion in a pediatric patient with precursor B-cell acute lymphoblastic leukemia

ETV6::RUNX1 fusion is one of the most common genetic alterations among children with precursor B-cell acute lymphoblastic leukemia (B-ALL). This fusion generally results from a t(12;21), which fuses the 5' region of ETV6 to the 3' region of RUNX1. Here we report a novel atypical fusion in a 2-year-old male patient with B-ALL. Chromosome, FISH, and microarray were first performed to define a genetic subtype. Chromosome analysis showed three copies of chromosome 21. FISH results revealed ETV6::RUNX1 fusion. However, the signal pattern was atypical with the two fusion signals sitting on two different chromosome 21s, a RUNX1 signal on the third chromosome 21, and two ETV6 signals on two chromosome 12s. Microarray did not readily resolve the FISH signal pattern but confirmed gain of the entire chromosome 21 without any imbalances and showed two losses affecting exon 2 and exons 6-8 of ETV6. Subsequently, optical genome mapping (OGM) identified the fusion created by excision and insertion of ETV6 exons 3-5 into intron 2 of RUNX1. Long-read DNA and RNA sequencing confirmed the fusion identified by OGM and resolved the fusion breakpoints. The identified novel RUNX1::ETV6::RUNX1 fusion contains all important elements reported in the classical ETV6::RUNX1 fusion and is assumed to be oncogenic. Further, transcriptome profiling demonstrated that this case clusters with other typical ETV6::RUNX1 fusion B-ALL cases, suggesting similar functional behavior from this atypical fusion. The patient was treated using a standard risk ALL protocol and is in remission for two years. This report demonstrates how comprehensive genomic profiling with OGM and long-read sequencing can help resolve atypical findings from conventional methods, enable proper sub-classification and potentially inform risk assessment in relapsed or drug-resistant disease.