Role of Glucocorticoid Receptor in Normal and Malignant Germinal Center B Cells

Diffuse large B cell lymphoma (DLBCL), the most common lymphoma type, remains incurable in ~30% of patients. We found that super-enhancers (SEs) are hypermutated by Activation-Induced Deaminase (AID)-mediated aberrant somatic hypermutation in >90% of DLBCL cases, indicating a prominent role in this disease pathogenesis (Bal et al. Nature 2022). Hypermutated SEs (over 80 recurrently targeted; on average, 12/case) are predominantly linked to genes encoding regulators of B cell development and well-known lymphoma oncogenes, such as BCL6, BCL2, and CXCR4. Among these, we showed that hypermutation of the SEs linked to BCL2 and CXCR4 abrogates the binding of the glucocorticoid receptor (GR), a transcription factor encoded by NR3C1. As a result, BCL2 and CXCR4 evade GR-mediated transcriptional repression, leading to their deregulated expression in germinal center (GC) B cells. Given the observation that the NR3C1 gene is genetically inactivated in a small percentage of DLBCL cases and considering the significant role of glucocorticoids in DLBCL treatment, we set out to comprehensively investigate the function of the GR in both normal and malignant GC B cells. We found that GR is detected in the nucleus of most GC B cells, suggesting that it is active as a transcription factor. Similarly, 32/32 DLBCL primary cases analyzed display GR nuclear expression, irrespective of molecular subtype. Analysis of a conditional GC-specific Nr3c1 knock-out (KO) mouse model showed that GCs form normally in the absence of Nr3c1. However, the transcriptional profile of Nr3c1-KO GC B cells was significantly altered compared to that of wild-type (WT) cells. By integrating the GR binding profile obtained in normal human GC B cells with the transcripts differentially expressed in Nr3c1-KO GC B cells, we identified significantly differentially expressed genes that are directly modulated by GR during the GC reaction. Gene-set enrichment analysis showed that GR controls several signaling pathways relevant to GC biology, suggesting a modulatory role on the BCR, MAPK, mTOR and apoptotic pathways, among others (q<0.05). In addition, GR targets display a significant overlap with the BCL6 transcriptional network, suggesting a cooperative role in GC B cell development. Notably, GR was shown to positively regulate programs involved in plasma cell differentiation, while repressing targets associated with memory B cell development. Consistently, loss of Nr3c1 impairs plasma cell differentiation in vivo and ex-vivo, in part due to the significantly reduced induction of Irf4 and Blimp1 protein. Finally, we identified several NR3C1 binding sites that are recurrently targeted by somatic mutations in DLBCL (2-10% of cases), suggesting that genetic impairment of GR activity is heterogeneous and extends beyond CXCR4 and BCL2. Together, these data support a role for the GR pathway in GC physiology. Recurrent inactivation of the NR3C1 gene or part of its transcriptional regulatory program may contribute to DLBCL pathogenesis, with implications for presently unknown specific roles of glucocorticoids in the therapeutic regimens for DLBCL.