Activation of protein phosphatase 2A in FLT3+ acute myeloid leukemia cells enhances the cytotoxicity of FLT3 tyrosine kinase inhibitors

Abstract

// Amanda M. Smith 1, 2, 3, * , Matthew D. Dun 1, 2, * , Erwin M. Lee 4 , Celeste Harrison 1, 2 , Richard Kahl 1, 2 , Hayley Flanagan 1, 2 , Nikita Panicker 1, 2 , Baratali Mashkani 1, 2, 5 , Anthony S. Don 6 , Jonathan Morris 7 , Hamish Toop 7 , Richard B. Lock 4 , Jason A. Powell 8 , Daniel Thomas 8, 9 , Mark A. Guthridge 10 , Andrew Moore 11 , Leonie K. Ashman 1, 2 , Kathryn A. Skelding 1, 2 , Anoop Enjeti 1, 2, 12 , Nicole M. Verrills 1, 2 1 School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, New South Wales, Australia 2 Hunter Medical Research Institute, Newcastle, New South Wales, Australia 3 Current address: The University of Queensland Diamantina Institute, Woolloongabba, Queensland, Australia 4 Children’s Cancer Institute Australia for Medical Research, Lowy Cancer Research Centre, UNSW, Sydney, New South Wales, Australia 5 Current address: Department of Medical Biochemistry, School of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran 6 Prince of Wales Clinical School, University of New South Wales, Sydney, New South Wales, Australia 7 School of Chemistry, University of New South Wales, Sydney, New South Wales, Australia 8 Centre for Cancer Biology, SA Pathology, Adelaide, South Australia, Australia 9 Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, California, USA 10 Department Clinical Haematology, Australian Centre for Blood Diseases, Monash University, Melbourne, Victoria, Australia 11 Translational Research Institute, The University of Queensland Diamantina Institute, Woolloongabba, Queensland, Australia 12 Calvary Mater Hospital, Newcastle, New South Wales, Australia * These authors contributed equally to this work Correspondence to: Nicole M. Verrills, email: nikki.verrills@newcastle.edu.au Keywords: PP2A, FTY720, AML, FLT3, tyrosine kinase inhibitor Received: April 07, 2016      Accepted: June 06, 2016      Published: June 18, 2016 ABSTRACT Constitutive activation of the receptor tyrosine kinase Fms-like tyrosine kinase 3 (FLT3), via co-expression of its ligand or by genetic mutation, is common in acute myeloid leukemia (AML). In this study we show that FLT3 activation inhibits the activity of the tumor suppressor, protein phosphatase 2A (PP2A). Using BaF3 cells transduced with wildtype or mutant FLT3, we show that FLT3-induced PP2A inhibition sensitizes cells to the pharmacological PP2A activators, FTY720 and AAL(S). FTY720 and AAL(S) induced cell death and inhibited colony formation of FLT3 activated cells. Furthermore, PP2A activators reduced the phosphorylation of ERK and AKT, downstream targets shared by both FLT3 and PP2A, in FLT3/ITD + BaF3 and MV4-11 cell lines. PP2A activity was lower in primary human bone marrow derived AML blasts compared to normal bone marrow, with blasts from FLT3-ITD patients displaying lower PP2A activity than WT-FLT3 blasts. Reduced PP2A activity was associated with hyperphosphorylation of the PP2A catalytic subunit, and reduced expression of PP2A structural and regulatory subunits. AML patient blasts were also sensitive to cell death induced by FTY720 and AAL(S), but these compounds had minimal effect on normal CD34+ bone marrow derived monocytes. Finally, PP2A activating compounds displayed synergistic effects when used in combination with tyrosine kinase inhibitors in FLT3-ITD + cells. A combination of Sorafenib and FTY720 was also synergistic in the presence of a protective stromal microenvironment. Thus combining a PP2A activating compound and a FLT3 inhibitor may be a novel therapeutic approach for treating AML.