Identification of Core Gene Signature Associated with Synergism Between ABT-869, a FLT3 Inhibitor and SAHA, a HDAC Inhibitor in AML with FLT3-ITD Mutation.

Internal tandem duplication of fms-like tyrosine kinase 3 (FLT3-ITD) mutation occurs in about 25% of acute myeloid leukemia (AML) patients and is associated with poor prognosis. Various FLT3 inhibitors of different chemical structure are under clinical investigation for the treatment of AML patients with FLT3 mutations. In contrast to their impressive potency in cell culture systems, FLT3 inhibitors as single agents in clinical trials only induce transient reduction of peripheral blasts, but not bone marrow blasts. Combination with other small molecule drugs represents a promising strategy to improve therapeutic efficacy of FLT3 inhibitors in the clinic. Small molecule HDAC inhibitors (HDACi) have proven to be a promising new class of anticancer drugs against hematological malignancies, as well as solid tumors. The HDACi, suberoylanilide hydroxamic acid (SAHA, Vorinostat) has been examined in a combinatory fashion with other classes of anticancer agents in acute leukemias, including cyclin-dependent kinase (CDK) inhibitors (flavopiridol), HSP 90 antagonists (17-allylamino- 17-demethoxygeldanamycin, 17-AAG), and estradiol analogs (2-methoxyestradiol, 2-ME). ABT-869, a multi-targeted receptor tyrosine kinase inhibitor, inhibits FLT3 phosphorylation and signaling and now is in active clinical development. In the current study, cell proliferation and apoptosis assays revealed that combining ABT-869 with SAHA led to synergistic killing of AML cells with FLT3 mutations in conventional cell culture and human stromal cell coculture models. To elucidate the molecular mechanism of the synergistic lethality between ABT-869 and SAHA, we compared the gene expression profiles of cells from FLT3 mutated leukemia cell lines, MV4-11 and MOLM-14, treated with DMSO control, ABT-869, SAHA and combination therapy using the Affymetrix microarray platform. Here, we focused on delineating a core gene signature unique to combination therapy and common to both MV4-11 and MOLM-14 cell lines, which could be crucial for the positive cooperation we observed. The core gene signature differentially induced more than two-fold by combination therapy in both cell lines included upregulation of FRY, LMO4, IFI16, ACADSB, and S100A8, and downregulation of PTP4A3 (PRL-3), ORC1L, MND1, ZNF85, DENND3, FAM119A, ACAT2, HMGCS1, DCS2, GSPG4, CBS, TUBE1. Overexpression of PRL-3, a metastasis-associated gene, has been found in different types of solid tumors and multiple myeloma. Modulation of PRL-3 expression level using genetic approaches demonstrated that PRL-3 played an essential role in the synergism in the therapeutic effect of ABT-869 and SAHA.