Super-enhancer-mediated regulation of CD38 inducibility in multiple myeloma via the MAF/TRIM47/RARα axis Free

Dysregulation of the MAF transcription factor, frequently overexpressed in multiple myeloma (MM) due to adverse translocations like t(14;16), is a well-established driver of poor prognosis. While the mechanisms are complex and involve super-enhancers (SEs), the specific downstream effectors contributing to MAF's oncogenicity remain incompletely understood. To identify critical MAF targets, we integrated genes associated with poor MM prognosis, MAF-regulated genes, and SE-associated genes, pinpointing TRIM47 as a prime candidate. Although TRIM47 is a crucial E3 ubiquitin ligase implicated in various cancers and inflammatory disorders, its functional role in MM remains undefined.

Knockdown of MAF in MM cell lines (ARP-1, KMS-11) significantly reduced both TRIM47 transcription and protein levels. Mechanistically, ChIP-seq and ATAC-seq analyses revealed that MAF binding extends beyond the TRIM47 promoter. Multiple MAF ChIP-seq peaks co-localized strongly with H3K27ac peaks within a ±10 kb window flanking the TRIM47 coding sequence (CDS), suggesting MAF-mediated SE activation drives TRIM47 expression. Functional validation using the SE inhibitor JQ1 and sgRNA-mediated targeted deletion confirmed four critical SE sites; disruption of these sites markedly diminished both MAF/H3K27ac enrichment and TRIM47 expression. To elucidate TRIM47's function, ubiquitinomics and proteomics screens identified potential interacting partners. Subsequent validation revealed a specific and robust endogenous and exogenous interaction solely between TRIM47 and RARα (Retinoic Acid Receptor Alpha), with their protein levels often showing an inverse correlation across MM cell lines. Notably, RARα activation by all-trans retinoic acid (ATRA) has been linked to enhanced efficacy of CD38-targeted immunotherapy in MM.

Using truncation mutagenesis, we mapped the interaction: RARα binds the SPRY domain of TRIM47, while TRIM47 binds RARα's ligand-binding domain (LBD). Critically, we demonstrated that TRIM47, via its RING domain, mediates K48-linked polyubiquitination of RARα specifically at lysine 134 (K134), targeting it for proteasomal degradation. Functionally, ATRA treatment induced CD38 expression more readily in MAF-knockdown or TRIM47-knockdown cells. This enhanced response was rescued by re-expressing wild-type TRIM47, but not by a RING domain-deletion mutant (TRIM47-ΔRING).

We next established that TRIM47 ablation specifically licenses ATRA-driven enhancement of CD38-targeted immunotherapy through integrated in vitro and in vivo models. Critically, co-culture experiments demonstrated that while TRIM47 knockdown alone did not alter baseline CD38-CAR-T cytotoxicity or T-cell activation profiles, ATRA pretreatment of shTRIM47 myeloma cells triggered dramatically enhanced CAR-T-mediated killing accompanied by significantly amplified IFN-γ/TNF-α secretion and robust CAR-T proliferative expansion – revealing TRIM47 targeting as a molecular prerequisite for ATRA-driven CAR-T potentiation. Paralleling these findings, in vivo studies using NSG mice engrafted with luciferase-coupled shTRIM47 myeloma cells showed comparable tumor progression and CD38-CAR-T monotherapy response relative to shCtrl controls; yet when combined with ATRA, the shTRIM47 cohort exhibited profound tumor regression and survival extension unseen in control groups. Mechanistic interrogation across both systems converged on a unified pathway: TRIM47 ablation stabilizes RARα, enabling ATRA to elevate CD38 surface expression which subsequently licenses enhanced CAR-T recognition, triggers amplified effector cytokine production, and drives clonal CAR-T-cell expansion – establishing a self-reinforcing therapeutic circuit.

In summary, our findings establish a novel MAF-TRIM47-RARα axis in MM pathogenesis: MAF orchestrates super-enhancer remodeling to drive TRIM47 expression, which subsequently targets RARα for degradation. This axis critically modulates the response to ATRA. Importantly, these results suggest that patients with t(14;16) or other high-MAF-expressing MM subtypes may exhibit diminished responsiveness to ATRA-mediated enhancement of CD38-targeted immunotherapy efficacy due to TRIM47-dependent RARα degradation. Targeting this axis represents a potential therapeutic strategy for this adverse-risk patient group.