The Polycomb group protein Ezh2 catalyzes the Histone H3 lysine-27 trimethylation (H3K27me3) within the Polycomb Repressive Complex 2 (PRC2). PRC2 exerts a critical control over the expression of a large set of target genes controlling important biological functions, including cell proliferation, differentiation and stem cell self-renewal.
Aberrant Ezh2 function is commonly observed in several cancer types and is due to deregulated enzymatic activity and/or expression of the Polycomb protein. Studies in preclinical models have started to reveal the importance of Ezh2 in B cell lymphomagenesis. In contrast, little is known about the effects of Ezh2 deregulated function/constitutive expression in B cell tumor maintenance and progression.
The present study addresses this issue taking advantage of a MYC-driven mouse lymphoma model, featuring high Ezh2 expression as a result of malignant B cell transformation. Conditional, genetic inactivation of Ezh2 methyltransferase activity in aggressive primary Burkitt-like mouse B cell lymphomas led to the identification of two classes of tumors, differentially responding to the loss of Polycomb function. In type-1 lymphomas, Ezh2 inactivation impaired clonal tumor growth starting from single lymphoma cells. Instead, type-2 lymphomas were largely resistant to the loss of Ezh2 catalytic function, giving rise to a substantial number of Ezh2 mutant clones. Transcriptome analyses allowed the identification of a molecular signature discriminating type-1 from type-2 lymphomas, including genes controlling cell cycle progression, DNA replication and cell survival, which were more expressed in type-2 tumors. These results correlated with a more aggressive behavior of type-2 lymphomas when transplantated into immunoproficient hosts.
The growth of rare Ezh2 mutant subclones, established from type-1 lymphomas, was impaired by the treatment with an Ezh1/2 small molecule inhibitor, identifying the Ezh2 paralog, Ezh1, as a determinant of resistance of tumor cells to Ezh2 inactivation. Ezh2 inhibition led to genome wide loss of H3K27me3, which was comparable between lymphoma types. However, while the loss of H3K27me3 at target genes in type-1 lymphomas failed to alter their expression, in type-2 lymphomas Ezh2 targets were in most cases deregulated following the loss of the histone mark. Based on these results, we propose that Ezh2 mutant subclones from type-1 lymphomas select an H3K27me3-independent mechanism to ensure correct regulation of Ezh2 target genes, which is needed for tumor growth. We also find that residual H3K27me3 is deposited at the promoter of new genes by a non-canonical PRC2/Ezh1, in Ezh2 mutant subclones from type-1 lymphomas. This activity alters the expression of target genes contributing to tumor growth.
We finally report the isolation of clonal variants from type-1 lymphomas that acquire secondary resistance to pharmacological Ezh1/2 inhibition. The latter tumors (together with type-2 lymphomas) will be instrumental to unravel the genetic bases of resistance of MYC-driven lymphomas to PRC2 inhibition.
Anti-Ezh2 inhibitors are currently being tested in phase-1 and -2 clinical trials for the treatment of both solid and blood cancers including B cell lymphomas. Our studies highlight the importance of understanding in more detail the mechanisms of action of Ezh2/PRC2 in tumors, in order to identify those that may benefit from anti-Ezh2 therapies. Our results also provide evidence for mechanisms of lymphoma resistance to Ezh2 inhibition and suggest strategies to circumvent such resistance
