A novel KMT2A–USO1 fusion gene‐induced de novo secondary acute myeloid leukaemia in a patient initially diagnosed with acute promyelocytic leukaemia

The question whether secondary acute myeloid leukaemia (sAML) originates from a newly acquired or pre-existing clone has not been resolved for decades. Most cases of sAML are generally considered as therapy-related leukaemia, especially those with chromosomal translocations at 11q23 that involve the KMT2A gene (also known as MLL). Here, we describe a 62-year-old male who was initially diagnosed with acute promyelocytic leukaemia (APL) carrying t(15;17)(q24;q21)/PML–RARa. He achieved complete remission after induction therapy with all-trans retinoic acid (ATRA), arsenic trioxide (ATO) and idarubicin (IDA). Subsequently, consolidation therapy with ATRA, IDA and cytarabine was administered for two cycles, and ATRA plus a intermediate dose of cytarabine for one cycle, followed by maintenance therapy with ATRA, ATO and methotrexate (MTX). Seventeen months after complete remission, the patient developed sAML with a t(4;11)(q21;q23) chromosomal translocation but no evidence of a PML–RARa transcript (Fig 1A and Figure S1, Table S1). During the consolidation and maintenance courses, the percentage and absolute count of peripheral monocytes in this patient increased repeatedly (Table S1). We first conducted whole genome sequencing (WGS) and RNA sequencing (RNA-seq) using bone marrow (BM) samples of APL and sAML to identify genomic and transcriptomic abnormalities, respectively. In the primary APL sample, we identified a typical PML–RARa transcript (Fig 1B, left panel). In the sAML sample, we found exon 9 of KMT2A to be fused with exon 2 of USO1 (Fig 1B, right panel), a novel KMT2A fusion partner, consistent with the t(4;11)(q21;q23) translocation detected in cytogenetic analysis (Fig 1A). USO1, also known as vesicle transport factor or P115, is a membrane tethering factor and has been reported to promote tumourigenesis through interaction with macrophage migration inhibitory factor and activation of the Erk pathway. In addition to the distinct fusion genes, somatic mutation analysis revealed no shared mutations between the APL and sAML samples (Fig 1C,D, Tables S2 and S3), suggesting that the APL and sAML clones most likely arose from distinct founding clones. Notably, two distinct forms of FLT3 mutations, i.e. FLT3– ITD and FLT3–TKD, existed in the APL and sAML samples, respectively (Fig 1C,D). To characterize the phenotype and the potential mechanisms of sAML induced by this novel fusion KMT2A–USO1, we performed single-cell RNA sequencing (scRNA-seq) using the sAML sample enriched for CD11b/CD15 double-negative progenitor cells to explore the mechanisms underlying KMT2A–USO1-induced leukaemogenesis. High-quality gene expression data were obtained from 5 088 single cells, and unsupervised clustering identified eight cell-type clusters (Fig 1E). These scRNA-seq data further revealed that KMT2A–USO1 showed similar but distinct molecular features to other KMT2A rearrangements. We found that KMT2A–USO1 also harboured the common characteristics of KMT2A fusions in significantly upregulating HOXA cluster genes (HOXA9 and HOXA10) and the HOX coregulator gene PBX3 (Fig 1F). More importantly, KMT2A–USO1 possessed several distinctive features in promoting leukaemogenesis. First, MPO, the gene generally expressing lower counts