Modeling del(17p) in multiple myeloma for molecular and functional characterization

Abstract Multiple myeloma (MM) remains a challenging hematological malignancy, with outcomes varying widely due to genetic heterogeneity. Among the cytogenetic abnormalities, deletion of the short arm of chromosome 17 (del(17p)) stands out as an important high-risk feature influencing prognosis. Del17p often involves the loss of the TP53 gene at 17p13.1 locus, a key tumor suppressor, however the impact of loss of multiple other genes on 17p on tumor cell behavior is still unclear. Evidence from studies indicates that the size of the deletion and the number of affected genes correlate with worse outcomes, highlighting a multifactorial impact that extends beyond TP53. In this study, using Molecular Alteration of Chromosomes with Engineered Tandem Elements (MACHETE) method, we induced del17p by creating a 21-Mb deletion of the short arm of chromosome 17 in the wild-type AMO1 cell line. This was followed by single-cell clone selection and expansion. The successful deletion of the 17p arm was confirmed in three single-cell expanded cultures through ultra-low pass sequencing and fluorescence in situ hybridization (FISH) for 17p, with deep whole-genome sequencing (WGS) further validating the 17p deletion breakpoint. Transcriptomic analysis revealed that 83% of genes located on the 17p arm were significantly downregulated (padj < 0.05) compared to parental cells. We next observed that del17p cells exhibit comparable proliferation rates to parental cells but demonstrate enhanced clonogenic potential, increased invasion capacity, and greater resistance to proteasome inhibitors. To determine which gene drives the phenotypic effects of del17p, we analyzed transcriptomic data from del17p patient cells across three independent datasets to identify genes on the 17p locus that are consistently downregulated in patients with del17p MM. By combining these findings with transcriptomic data from our isogenic line, we pinpointed a core group of 14 genes within the 17p locus that are consistently downregulated in del17p cells and reside within the minimal deleted region. Notably, reduced expression of SAT2 stood out as being specifically linked to worse patient outcomes. Functional studies further demonstrated that reintroducing SAT2 into 17p-deleted cell lines counteracts the enhanced clonogenic potential driven by 17p loss, suggesting that SAT2 plays a critical role in modulating the aggressive behavior of del17p cells. In conclusion, our induced del17p cell line using MACHETE technique provides ability to attain new insights into the molecular mechanisms underlying del17p-associated phenotypes and identifies SAT2 as one of the key genes within the 17p locus that contributes to the phenotypic effects of del17p.