The c-terminal to lish (CTLH) E3 ligase complex modulates myeloma glucocorticoid sensitivity by regulating nfkb signaling Free

Background: Glucocorticoids remain a cornerstone of frontline therapy for multiple myeloma (MM), yet resistance to these agents—particularly dexamethasone—poses a major barrier to sustained clinical responses. Although dexamethasone promotes apoptosis through glucocorticoid receptor (GR)-mediated transcriptional activation, the molecular networks that modulate this response in MM are incompletely understood.

Methods: To systematically identify genetic determinants of glucocorticoid (GC) sensitivity, we conducted genome-wide CRISPR-Cas9 loss-of-function screens in KMS11 and LP1 t(4;14) MM cell lines exposed to dexamethasone. Key resistance and sensitization nodes were validated using targeted gene disruption and functional assays.

Results: Our screening results confirmed, as predicted, that canonical glucocorticoid receptor (GR) signaling components—such as NR3C1 (GR) and its co-chaperone PTGES3—are essential for dexamethasone-induced apoptosis. We also identified NFKBIA and NFKBIB, members of the NF-κB inhibitor family, as key mediators of glucocorticoid cytotoxicity; disruption of the genes encoding these proteins conferred resistance to glucocorticoids. In contrast, chemical inhibitors of IκB kinase (IKK)—which block IκB degradation and thereby suppress NF-κB activity—synergized with glucocorticoids to induce cytotoxicity in MM cells. Intriguingly, genetic depletion of FKBP5, a feedback regulator of GR signaling, sensitized MM cells to dexamethasone, suggesting a potential therapeutic vulnerability. Supporting this, both a small-molecule inhibitor of FKBP5 and a PROTAC-based degrader enhanced GC-mediated apoptosis. Most notably, we identified the C-terminal to LisH (CTLH) E3 ubiquitin ligase complex as a novel driver of dexamethasone resistance. Although disruption of individual CTLH components—such as MAEA and WDR26—had only a modest effect on MM cell viability, it significantly enhanced GC-induced cytotoxicity. Analysis of genomic data from the MMRF CoMMpass study revealed that high expression of various CTLH subunits correlates with poor patient survival. Given that nearly all patients in this study receive glucocorticoids as part of standard therapy, this association implies that elevated CTLH activity contributes to GC tolerance. Mechanistically, inhibition of CTLH activity did not affect GR protein levels, nuclear translocation, or transcriptional output. However, it led to a striking increase in GC-mediated IκB induction, thereby suppressing NF-κB signaling and amplifying GR-driven apoptotic pathways. Notably, this enhanced IκB induction was not accompanied by changes in mRNA expression, suggesting that the CTLH complex facilitates IκB degradation. This previously unrecognized function of the CTLH complex reveals a druggable axis of glucocorticoid resistance in multiple myeloma.Conclusion: Our findings position the CTLH complex as a central modulator of glucocorticoid resistance in MM. Targeting CTLH-mediated degradation pathways—particularly in combination with next-generation GR agonists—may restore dexamethasone sensitivity while minimizing systemic toxicity. This work lays the foundation for innovative, mechanism-informed combination therapies in relapsed/refractory MM.