MYC partners with kinase-independent PIM2 at key promoters in multiple myeloma

Abstract Introduction: Multiple Myeloma (MM) is an incurable hematological malignancy characterized by abnormal plasma cell proliferation in the bone marrow. Despite recent therapeutic advances, MM remains a significant clinical challenge, largely due to the persistent problem of drug resistance. The identification of pro-survival mechanisms and regulatory networks in MM cells is therefore critical to developing new strategies to overcome resistance and improve outcomes. PIM2 is an oncogenic serine/threonine kinase whose overexpression in MM correlates with aggressive disease and poor prognosis. We previously reported a novel, kinase-independent (Ki) function of PIM2, whereby PIM2 expression is maintained via a feedback loop involving MYC and SP1 transcription factors, distinct from its canonical kinase-dependent (Kdep) activities. This Ki PIM2-MYC-SP1 complex at the PIM2 promoter represents a potentially targetable vulnerability in MM. Methods: To comprehensively investigate mechanisms of Ki PIM2 function, we generated MM1.S cell lines with FLAG-tagged wild-type (WT) and kinase-dead (KD) PIM2 overexpression. ChIP-sequencing (ChIP-seq) was performed for FLAG (PIM2), MYC, IgG, and input controls. Motif and peak analysis identified genomic loci with PIM2/MYC enrichment. ChIP-qPCR validated key promoter interactions. Gene expression was further analyzed by RT-qPCR and protein levels by Western Blot following genetic and pharmacological inhibition. Results: ChIP-seq analysis revealed robust enrichment of both FLAG-PIM2 and MYC at the PIM2 promoter in both FLAG-PIM2WT and FLAG-PIM2KD, indicating that PIM2 occupancy and complex formation with MYC at this locus are independent of its kinase activity. Genome-wide, we identified the MRPS10 promoter as another significant locus with co-enrichment of PIM2 and MYC. RT-qPCR and Western Blot confirmed that MRPS10 expression is regulated analogously to PIM2, suggesting that the PIM2-MYC-SP1 complex orchestrates a broad regulatory program extending beyond autoregulation of PIM2 itself. Conclusions: Our data suggest that PIM2, through kinase-independent mechanisms, forms a regulatory complex with MYC and SP1 at specific gene promoters, including both PIM2 and MRPS10. Disruption of this complex abrogates expression of its target genes, highlighting an expanded therapeutic window for targeting the broader PIM2-MYC-SP1 transcriptional network in MM. These findings suggest that kinase-independent PIM2-driven gene regulation is a critical node in MM pathogenesis, offering new opportunities for clinical intervention. Further studies are underway to map the full repertoire of genes governed by this complex and to develop strategies for its selective inhibition.