PRMT5 Initiates an Epigenetic Program That Promotes Progression of Chronic Lymphocytic Leukemia

Background: Epigenetic alterations that occur as a result of arginine methyltransferase activity play a critical role in normal biological functions, and increasingly their importance has been shown in cancer initiation and progression. In particular, protein arginine methyltransferase 5 (PRMT5) is an important epigenetic regulator that influences differentiation, cell cycle progression, and many other processes. Aberrant PRMT5 activity has been implicated in tumorigenesis, however its role in the biology of chronic lymphocytic leukemia (CLL) has not been established. Despite recent improvements in therapy, CLL remains an incurable disease. While newer therapies targeting the B cell receptor pathway have shown remarkable efficacy, complete responses are not frequent. In particular, transformation of CLL to aggressive lymphoma (Richter syndrome; RS) occurs in up to 15% of patients and confers a poor prognosis; however, the mechanisms by which RS occurs are poorly understood. A better understanding of CLL progression may facilitate the development of new targeted therapies. We hypothesized that PRMT5 dysregulation initiates an epigenetic program in CLL that contributes to disease progression and potentially transformation. Moreover, we provide evidence that inhibition of PRMT5 is a promising strategy in CLL.

Methods: Eμ-PRMT5 transgenic mice were generated on the C57BL/6 background via pronuclear injection of human PRMT5 cDNA under the heavy chain promoter/enhancer. To investigate the effect of PRMT5 overexpression on CLL pathogenesis, Eμ-PRMT5 mice were crossed with Eμ-TCL1 mice, an aggressive, spontaneous model of CLL, and followed for disease onset, progression, and overall survival. Immunophenotypic analysis was performed using multicolor flow cytometry. Pharmacologic inhibition of PRMT5 was evaluated in freshly isolated CD19+-selected cells isolated from CLL patients. Kaplan-Meier curves were compared using a log-rank test.

Results: We observed that Eμ-PRMT5/TCL1 had a shorter time to leukemia onset (defined as >20% CD45+/CD5+/CD19+ in peripheral blood) compared to the Eμ-TCL1 mice. The Eμ-PRMT5/TCL1 had significantly worse overall survival compared to the Eμ-TCL1 mice. In addition to more aggressive disease, Eμ-PRMT5/TCL1 mice developed focal lymphoid tumors and lymphadenopathy in contrast to Eμ-TCL1 mice. We generated an inducible knockout CLL-like cell line using CRISPR guide RNAs targeting PRMT5 . Genomic ablation of PRMT5 led to impaired protein expression and subsequent decrease in viable cells. We next assessed the cytotoxic effect of PRMT5 inhibition in CLL using the pharmacologic agents EPZ015666 (Chan-Penebre et al. 2015 Nat Chem Biol) and an in-house developed PRMT5 inhibitor. Both compounds inhibited growth of CD19+-selected cells isolated from CLL patients as well as CLL-like cell lines (Mec-1 and HG3). Pharmacologic PRMT5 inhibition in vivo was able to efficiently modulate symmetric dimethyl arginine motifs. Studies exploring the impact of in vivo PRMT5 inhibition on disease burden and overall survival in Eμ-PRMT5/TCL1 mice are currently ongoing.

Conclusions: Our data indicate that PRMT5 facilitates CLL progression through altered epigenetic programming. PRMT5 overexpression in Eμ-TCL1 mice results in a more aggressive disease model with features of a focal lymphoid accumulation. Studies to further characterize the genomic and phenotypic features of this nodal disease are ongoing. Furthermore, our data support pharmacologic inhibition of PRMT5 as a promising therapeutic paradigm in CLL.