Essential Role for Phospholipase C Gamma 1 (PLC-γ1) in the Survival of t(8;21) Acute Myeloid Leukemia

The t(8;21) (q22;q22) chromosomal translocation is one of the most frequent genetic alterations in acute myeloid leukemia (AML) and it remains a significant clinical problem especially for children which indicates the need for improved therapeutic strategies. Recently, we showed that peptide derived phospholipase C gamma 1 (PLC-γ1) was highly phosphorylated in pediatric t(8;21) AML. In this study, we determined PLC-γ1 phosphorylation and mRNA levels showing that PLC-γ1 expression was significantly higher in t(8;21) AML compared to other AML karyotypes and normal bone marrow (NBM) (peptide phosphorylation: p<0.01 compared to NBM, mRNA: p<0.001, compared to other AML karyotypes). This was confirmed by PLC-γ1 protein phosphorylation using primary AML samples and AML cell lines. PLC-γ1 is known to play a role in cancer progression, however, the impact of PLC-γ1 in AML is currently unknown. Therefore, we aimed to study the functional role of PLC-γ1 by investigating the cellular growth, survival and its underlying mechanism in a t(8;21) AML cell line (Kasumi-1) . ShRNA-mediated knockdown of PLC-γ1 in kasumi-1 cells significantly blocked leukemic cell growth at day 8 after transduction (p<0.05). The percentage of apoptosis in PLC-γ1 suppressed kasumi-1 cells at day 4 after transduction was two-fold higher compared to the scrambled control (p<0.01). The inhibition of cell proliferation and the induction of apoptosis upon PLC-γ1 suppression could be explained by cell cycle arrest and by increased activation of apoptotic related and cell cycle regulatory protein expressions (BAX, BCL2, p53 and Chk2). As the multidrug resistance is one of the major cause of relapse and poor prognosis in t(8;21) AML, therefore, we demonstrated, if PLC-γ1 suppression increased the sensitivity of kasumi-1 leukemia cells to cytotoxic chemotherapeutic agents (methotrexate, amsacrine and etoposide). PLC-γ1 knockdown cells at day 4 after transduction were shown to significantly reduced cell viability to the genotoxic agents, methotrexate (p<0.05, p<0.001), amsacrine (p<0.01, p<0.001) and etoposide (p<0.05, p<0.01 and p<0.001) in kasumi-1 in a dose-dependent manner. These results provide a strong rationale for the development of PLC-γ1-based therapeutic strategies for the enhancement of efficacy in t(8;21) AML treatment. Additionally, PLC-γ1 suppressed kasumi-1 cells showed significantly less proliferation upon hypoxic stress. Taken together, these results strongly support an important role for PLC-γ1 in the survival of t(8;21) AML mimicking kasumi-1 cell line and identify PLC-γ1 as a potential target for t(8;21) AML treatment.