Program: Oral and Poster Abstracts
Session: 616. Acute Myeloid Leukemia: Novel Therapy, excluding Transplantation: Poster I
In the current study, we show that the pro-differentiation effects of ATRA are markedly potentiated when combined with agents that target PI3K/AKT/mTOR signalling. In AML cell lines and primary patient samples, we observed additive pro-differentiation effects when ATRA was combined with inhibitors of PI3K (ZSTK474) and mTOR complex proteins (Torin-1, WYE-125132). However, when combined with the bromodomain inhibitor NVP-BEZ235, a dual inhibitor of PI3K and mTOR, we observed synergistic induction of CD11b by FACS analysis. Combination studies revealed loss of cell viability, cell cycle arrest in G1 phase, and impaired clonogenic survival, which was more prominent for ATRA combination treatments than with any agent used alone (Figure 1). To assess the role of c-Myc in mediating these effects, we measured c-Myc protein levels and PI3K/AKt/mTOR pathway markers at different time-points following treatment with ATRA alone and in combination with the inhibitors described above (Figure 2). Our findings suggest that ATRA alone quickly down-regulates c-Myc (within 6 hours) through transcriptional repression. Disruption of the PI3K/AKT/mTOR pathway further down-regulates c-Myc (within 3 hours) through destabilization and enhanced degradation. ATRA combined with NVP-BEZ235 produced maximal c-Myc suppression, and led to more cell kill than any other combination tested. Detailed analysis of changes in the transcriptome in MV-411 cells following treatment with ATRA and NVP-BEZ235 revealed that both agents act jointly on the regulation of the same biological pathways and processes, but regulate different sets of genes within these pathways. Updated mechanism based studies will be presented.
In conclusion, suppression of c-Myc levels through disruption of PI3K/AKT/mTOR signalling augments the anti-leukemic effects of ATRA. These data support the clinical investigation of ATRA combined with rapalogs or bromodomain inhibitors.
Figure 1 - Combination treatment with PI3K/mTORC inhibitors and ATRA decreases cell viability in AML cells. MV4-11 cells were treated as indicated with combinations of BEZ (1µM), WYE (1µM) or ZSTK (2.5µM) and ATRA (0.1 µM). Number of cells was determined by CellTiter-Glo¨ luminescent cell viability assay (Promega). Data were analyzed by one-way ANOVA (P < 0.0001) followed by TukeyÕs post-hoc test. * P < 0.05, ** P < 0.01, *** P < 0.001,**** P < 0.0001.
Figure 2 – Reduced expression of MYC protein by inhibition of the PI3K/AKT/mTORC pathways. Immunoblotting/quantification of MYC protein levels in MV4-11 cells following treatment with combinations of WYE (1µM), BEZ (1µM), ZSTK (2.5µM) and ATRA (0.1 µM).
Disclosures: No relevant conflicts of interest to declare.
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