PB1652 ABERRANT EZH2 HIGH EXPRESSION IN THE PATIENTS WITH ACUTE LEUKEMIA AND EFFECT OF A NOVEL EZH2 INHIBITOR GSK126 ON CELL PROLIFERATION AND APOPTOSIS OF THE MALIGNANT CELLS
Authors: Z. Ge, Y. Gao, Q. Zhang, Q. Han, J. Zi, C. Song
Journal: HemaSphere (2019)

Background: Enhancer of Zeste homolog 2 (EZH2) is an inevitable subunit of polycomb repressive complex 2 (PRC2), and a critical enzyme for the trimethylation of lysine 27 of histone H3 (H3K27me3). EZH2 mutations and aberrant expression are implicated in tumorigenesis and correlate with poor prognosis in tumors. GSK126, a highly selective inhibitor of EZH2 exerts its anti-tumor effect by inducing cell proliferation arrest and apoptosis. However, the GSK126 anti-tumor effect is still not well determined in acute leukemia. Aims: To examine EZH2 expression and mutation and investigate the effect of EZH2 inhibitor GSK126 on cell proliferation and apoptosis in adult acute leukemia. Methods: Bone marrow samples from 169 ALL patients (124 B-ALL, 42 T-ALL, and 3 B-/T-ALL), 32 AML and 37 control were included. PCR amplification of EZH2 exons combining Sanger sequencing was performed in 146 ALL and 32 AML samples for mutation screening. The RT-qPCR was conducted to measure EZH2 expression in 126 available ALL and 37 control samples. Patients were allocated in a high or low EZH2 expression cohort (3rd-4thquartile vs.1st-2nd quartiles) by SPSS 20.0. The CCK-8 assay and Annexin V/PI staining were performed to measure the cell proliferation and apoptosis, respectively in the leukemia cell lines. Results: Three EZH2 exonic mutations in 2 (2.7%) of 146 ALL and 1 mutant in 1 (3.1%) of 32 AML samples were detected in our cohort. Two mutants (K466T and T467fs∗ > 3) in T-ALL and one mutant (E404D) in AML are our identified novel mutants. The two ALL patients with EZH2 mutations showed multiple relapses, poor outcome, and failure of new drug treatment. EZH2 mRNA level is significantly higher in ALL patients compared to 37 normal controls (P < 0.001). The two EZH2 mutated patients showed EZH2high expression. Patients with EZH2high expression harbored a higher median percentage of lymphoblast counts in both BM and peripheral blood (89% vs. 79%, P = 0.024; 84.2% vs. 48.6%, P = 0.043) and higher frequency of splenomegaly and lymphadenopathy than those with EZH2low expression. The relapsed and refractory rate was much higher in the EZH2high group than the EZH2low group (65.0% vs. 44.8%, P = 0.015). Moreover compared to control group (DMSO), GSK126 treatment with doses ranged from 5 μM to 25 μM showed significant dose- and time-dependent cell proliferation suppression in CEM T-ALL cells (P = 0.000), U937 AML cells (P = 0.000), while only the dose ranged 15 μM to 30 μM showed significant dose- and time-dependent cell proliferation suppression in Raji lymphoma cells (P = 0.000). The IC50 of GSK126 on CEM, U937, Raji cells were (13.46 ± 0.83) μmol/L, (11.65 ± 1.02) μmol/L, (15.00 ± 0.19) μmol/L, respectively. The GSK126 (8,12, & 16 μM) treatment significantly promoted the apoptosis of CEM and U937 cells compared with control group (DMSO), respectively(P = 0.000); while only 16 μM GSK126 treatment showed the significant effect on the apoptosis of Raji cells. Furthermore, GSK126 treatment significantly suppressed the expression of EZH2, and also its downstream effector Bcl-2 and Bcl-xL in the cells. Summary/Conclusion: EZH2 is highly expressed in adult leukemia and the EZH2 mutations count partly on the aberrant high expression, which reveals that EZH2 may play important roles in acute leukemia progression. EZH2 inhibitor GSK126 induces cell proliferation arrest and apoptosis in the malignant cell lines, which indicate the potential to incorporate the EZH2 inhibitor into the clinical trial in the therapy of acute leukemia.