HALTING THE NAD+-DEPENDENT DEACETYLASE SIRT6 UNVEILS SPLICEOSOME DEREGULATION AS EXPLOITABLE VULNERABILITY FOR AGGRESSIVE MYELOMA

Epigenetic defects play a major role in tumors, by promoting aggressiveness, drug resistance and disease progression. Targeting enzymes responsible for these changes holds promise but the knowledge-gap and lack of selective inhibitors severely hampers their clinical use. Recently, we found that Multiple Myeloma (MM) cells exhibit high level of NAD+-dependent deacetylase SIRT6 as an adaptive response to their intrinsic genomic instability. Here we focus on splicing-related events resulting as the most enriched pathways among MM patients with high SIRT6 levels. CoMMpass dataset analysis confirmed the prognostic role of SIRT6 in MM, with its presence correlating with poor outcome and aggressiveness. Transcriptome analysis of SIRT6-depleted MM cells indicated spliceosome perturbation as critical: chemical and genetic approaches resulted in enhanced anti-MM activity of spliceosome modulator Sudamycin D6 (SD6) thus supporting RNA splicing deregulation as vulnerability for these cells. Spliceosome is a therapeutic weakness for MYC-driven cancer, as such we found that while c-MYC oncogene loss shows resistance, its re-expression enhances efficacy of tested strategy. A direct role for SIRT6 on splicing factors genes was also demonstrated using ChIP-seq analyses. Overall, the dysregulated spliceosome activity triggered by epigenetic machinery blocking represents a novel therapeutic strategy for poor-prognosis subset of MM patients overexpressing c-MYC The HMCL were purchased from ATCC or DSMZ. All cell lines were Mycoplasma-free and routinely tested for it. Cells were cultured in RPMI-1640 medium containing 10% fetal bovine serum (FBS; GIBCO), 4mM glutamine, 100 U ml−1 penicillin, and 100 μg ml−1 streptomycin (GIBCO). 293T were cultured in DMEM high glucose containing 10% FBS (GIBCO), 4 mM glutamine, 100 U ml−1 penicillin, and 100 μg ml−1 streptomycin (GIBCO). RNA samples from NCI-H929 and MM1.S cells scramble (SCR) and silenced for SIRT6 (shSIRT6) were obtained in duplicate for each condition and processed using WT PLUS Reagent Kit, according to manifacturer’s protocol (Thermo Fisher). mRNA-transcriptome profiling was assessed using GeneChip™ Human Gene 2.0 ST Array (Thermo Fisher).