Identification of a Midnolin-proteasome pathway impacting Multiple Myeloma cell growth

Multiple myeloma (MM), a common hematological cancer, remains incurable despite advances in treatment. Identifying new targets is essential for understanding MM biology and developing innovative therapies. While Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas9 knockout (KO) technology has identified new oncogenes, it does not address proliferation suppressor genes (PSGs), which are critical in growth suppression and often downregulated in cancer. To overcome this, we conducted a genome-wide CRISPR activation (CRISPRa) screen in a panel of MM cell lines, enabling gain of function (GOF) studies. Among the top hits impairing MM cell fitness was MIDN. We confirmed the impact of MIDN overexpression in multiple MM cell lines, defining it as a PSG in myeloma. Importantly, we observed a lower expression of MIDN in two large cohorts of newly diagnosed MM patients (n = 507 and n = 319 respectively) compared to normal plasma cells (PCs, n = 21 and n = 16 respectively). MIDN expression was also significantly lower in premalignant conditions, suggesting an early downregulation in disease progression. Furthermore, MIDN expression was significantly lower in six human MM cell lines compared to B cells from three healthy donors (HDs). MIDN encodes the protein Midnolin which mediates ubiquitination-independent protein degradation inducing the degradation of many nuclear proteins including stimulus-induced transcription factors (Fos, EGR1, NR4A1). Mass spectrometry analysis in MM cells confirmed that MIDN bound to the 19S and 20S proteasomal subunits, as well as to important myeloma nuclear factors such as IRF4. Although the mechanisms behind MIDN downregulation in MM remain unclear, our study shows that the downregulation of MIDN, mediating ubiquitination-independent protein degradation, may be a crucial event for MM development and pathogenesis, as its observed universal downregulation increases IRF4 protein expression. Further research is ongoing to explore mechanisms mediating MIDN downregulation as well as deep evaluation of the functional landscape of putative suppressor genes in MM, which may expose new vulnerabilities and provide additional therapeutic opportunities.