Unravelling the biological and clinical features of childhood B-cell precursor acute lymphoblastic leukemia with monosomy 7

Abstract Monosomy of chromosome 7 is a common cytogenetic aberration predicting prognosis in myeloid malignancies and promoting transformation to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) in leukemia predisposing constitutional syndromes. In B-cell precursor acute lymphoblastic leukemia (BCP-ALL) the loss of chromosome 7 almost exclusively occurs in either hypodiploid karyotypes or as a secondary abnormality in Philadelphia chromosome (Ph) positive cases. Outside these contexts, the prognostic significance and biological features of ALL with monosomy 7 have not been explored. We collected data of 40 childhood patients (< 18 years of age) treated for BCP-ALL with monosomy 7 (excluding Ph-positive and hypodiploid cases) from nine countries (Germany, Austria, Italy, Netherlands, Czech Republic, Israel, Denmark).We investigated their biological and clinical characteristics, the incidence of any preexisting disorders, constitutional defects contributing to leukemia development, and long-term outcome as well as coexisting somatic aberrations.Finally, we performed single cell RNA sequencing (scRNA-seq) of four monosomy 7 B-ALL patients using the 10x Genomics Chromium controller and the Next GEM Single Cell 5' Reagent Kit protocol. Libraries were sequenced on a NovaSeqX 10B - 150bp PE/SE with 50.000 total reads per cell. Raw data were processed using Cell Ranger (v7.2.0) and Seurat (v4.4.0). Aneuploid cells were distinguished from diploid cells using the Single Cell Variational Aneuploidy analysis (SCEVAN) algorithm. Gene set enrichment analysis (GSEA) was carried out using AUCell (AUCell_1.20.0; 10.18129/B9.bioc.AUCell). We found that a prevalence of BCP-ALL with monosomy 7 among pediatric patients with ALL is <0.5% based on two consecutive Polish patient cohorts accounting for 0.4% (6/1386; ALL-IC BFM 2009) and 0.39% (5/1277; AEIOP BFM 2017), respectively. Within the entire international cohort including n=22 males and n=18 females, the median age at ALL diagnosis was 8.0 [6-12] years, the median WBC 6.9/µl (3.3-30.6), and no CNS or testicular involvement was noted. Most patients (57.5%; 23/40) were assigned to a high-risk treatment group (HRG) based on MRD level after induction therapy; three patients received HSCT in the first complete remission, five experienced a relapse, and five died. Copy number abnormalities affecting the short arm of chromosome 9 (PAX5 deletions, amplifications, CDKN2A/B deletions) and aberrations of chromosome 21 (+21; +21q22; iAMP21) recurrently co-occurred with monosomy 7 in 60% (15/25) and 30% (9/27) of patients, respectively. Interestingly, four cases carried IGH gene rearrangements resulting in three different in-frame gene fusions (IGH::DUX4, IGH::CRLF2, IGH::TRAD). In total, eight (20%) of patients showed preexisting disorders [Bloom syndrome (n=2), IKZF1-related CVID-13 (n=1), aplastic anemia with cleft palate (n=1), recurrent pancytopenia (n=1), mastocytosis (n=1), urinary tract defect (n=1), diabetes type 1 (n=1)], and three of them developed BCP-ALL as a secondary malignancy. Using scRNA-seq we generated data from 4 patients (n=1 CDKN2A/B del, PAX5 del, n=1 CDKN2A/B del, PAX5 amp, n=1 iAMP21, n=1 IGHr). Across all samples, we profiled a total of 21,740 high-quality cells, with a median of 1,866 genes per cell. We integrated our B-ALL dataset with healthy bone marrow scRNA-seq data from eight donors (Human Cell Atlas, ERP122984). In all leukemia samples, we observed four B-cell lineage clusters corresponding to distinct stages of differentiation: pro-B, pre-B, immature, and mature B-cells. Using SCEVAN, we detected and quantified aneuploid cells with reduced expression of genes encoded by chromosome 7 (score < −0.15) on average in 79% of cells across all patient samples. The main cell populations of interest were pro-B cells, being the most enriched within aneuploid monosomy 7 cells (90.6%). GSEA on Hallmark revealed the top eight downregulated pathways in aneuploid compared to diploid pro-B cells, including cell cycle (G2M checkpoint, E2F targets), OXPHOS, and DNA-repair. BCP-ALL with monosomy 7 is a rare leukemia characterized by coexisting aberrations involving chromosomes 9p, 21, and 14. Despite most patients being assigned to HRG and the overrepresentation of preexisting disorders in these patients, the overall and relapse-free survival rates of patients with monosomy 7 BCP-ALL were not substantially poorer compared to other patients allocated to HRG.