All-trans retinoic acid enhances, and a pan-RAR antagonist counteracts, the stem cell promoting activity of EVI1 in acute myeloid leukemia

Abstract

Ecotropic virus integration site 1 (EVI1), whose overexpression characterizes a particularly aggressive subtype of acute myeloid leukemia (AML), enhanced anti-leukemic activities of all-trans retinoic acid (atRA) in cell lines and patient samples. However, the drivers of leukemia formation, therapy resistance, and relapse are leukemic stem cells (LSCs), whose properties were hardly reflected in these experimental setups. The present study was designed to address the effects of, and interactions between, EVI1 and retinoids in AML LSCs. We report that Evi1 reduced the maturation of leukemic cells and promoted the abundance, quiescence, and activity of LSCs in an MLL-AF9-driven mouse model of AML. atRA further augmented these effects in an Evi1 dependent manner. EVI1 also strongly enhanced atRA regulated gene transcription in LSC enriched cells. One of their jointly regulated targets, Notch4, was an important mediator of their effects on leukemic stemness. In vitro exposure of leukemic cells to a pan-RAR antagonist caused effects opposite to those of atRA. In vivo antagonist treatment delayed leukemogenesis and reduced LSC abundance, quiescence, and activity in Evi1high AML. Key results were confirmed in human myeloid cell lines retaining some stem cell characteristics as well as in primary human AML samples. In summary, our study is the first to report the importance of EVI1 for key properties of AML LSCs. Furthermore, it shows that atRA enhances, and a pan-RAR antagonist counteracts, the effects of EVI1 on AML stemness, thus raising the possibility of using RAR antagonists in the therapy of EVI1high AML.

Evi1 Counteracts Anti-Leukemic and Stem Cell Inhibitory Effects of All-Trans Retinoic Acid on Flt3-ITD/Npm1c-Driven Acute Myeloid Leukemia Cells

“…These effects were counteracted by experimental expression of Evi1 . These data confirm the earlier noted genetic and molecular complexity of the responsiveness of AML LSCs to atRA [ 27 , 37 ].…”

“…In AML1-ETO -expressing murine BM cells, atRA increased serial replating ability and led to the formation of larger and more immature colonies [ 21 ], suggesting that atRA promoted LSC activity on this genetic background. Similarly, atRA promoted leukemic stemness in an MLL-AF9 -driven mouse model of AML in a manner dependent on the expression of Evi1 [ 37 ]. MLL rearrangements are frequently associated with EVI1 overexpression in human AML [ 45 , 46 ], and this is reflected in the corresponding mouse model [ 37 , 47 ].…”

“…Similarly, atRA promoted leukemic stemness in an MLL-AF9 -driven mouse model of AML in a manner dependent on the expression of Evi1 [ 37 ]. MLL rearrangements are frequently associated with EVI1 overexpression in human AML [ 45 , 46 ], and this is reflected in the corresponding mouse model [ 37 , 47 ]. In BM cells from MLL-AF9 leukemic mice, atRA promoted the abundance and quiescence of an immunophenotypically defined LSC-enriched cell population and enhanced the activity of LSCs as determined by serial replating and in vivo limiting dilution assays.…”

“…In BM cells from MLL-AF9 leukemic mice, atRA promoted the abundance and quiescence of an immunophenotypically defined LSC-enriched cell population and enhanced the activity of LSCs as determined by serial replating and in vivo limiting dilution assays. These effects of atRA were abolished by knock-down of Evi1 [ 37 ]. Correspondingly, an RAR antagonist decreased LSC-related properties in an Evi1 -dependent manner in vitro and in vivo, and prolonged survival of both of primary treated and secondary recipient mice with Evi1 high , MLL-AF9 -driven AML.…”

“…By contrast, atRA promoted serial replating ability—considered as a readout of stem cell activity—in AML1-ETO -expressing murine BM cells [ 21 ]. Similarly, in an MLL-AF9 -driven mouse model of AML, atRA augmented stem cell abundance, quiescence, and activity in a manner that was dependent on the expression of Evi1 [ 37 ]. EVI1 and atRA also collaborated to promote stem cell-related properties in human AML cell lines and primary samples [ 37 ].…”