A screen for Fli-1 transcriptional modulators identifies PKC agonists that induce erythroid to megakaryocytic differentiation and suppress leukemogenesis

Abstract

The ETS-related transcription factor Fli-1 affects many developmental programs including erythroid and megakaryocytic differentiation, and is frequently de-regulated in cancer. Fli-1 was initially isolated following retrovirus insertional mutagenesis screens for leukemic initiator genes, and accordingly, inhibition of this transcription factor can suppress leukemia through induction of erythroid differentiation. To search for modulators of Fli-1, we hereby performed repurposing drug screens with compounds isolated from Chinese medicinal plants. We identified agents that can transcriptionally activate or inhibit a Fli-1 reporter. Remarkably, agents that increased Fli-1 transcriptional activity conferred a strong anti-cancer activity upon Fli-1-expressing leukemic cells in culture. As opposed to drugs that suppress Fli1 activity and lead to erythroid differentiation, growth suppression by these new Fli-1 transactivating compounds involved erythroid to megakaryocytic conversion (EMC). The identified compounds are structurally related to diterpene family of small molecules, which are known agonists of protein kinase C (PKC). In accordance, these PKC agonists (PKCAs) induced PKC phosphorylation leading to activation of the mitogen-activated protein kinase (MAPK) pathway, increased cell attachment and EMC, whereas pharmacological inhibition of PKC or MAPK diminished the effect of our PKCAs. Moreover, in a mouse model of leukemia initiated by Fli-1 activation, the PKCA compounds exhibited strong anti-cancer activity, which was accompanied by increased presence of CD41/CD61 positive megakaryocytic cells in leukemic spleens. Thus, PKC agonists offer a novel approach to combat Fli-1-induced leukemia, and possibly other cancers,by inducing EMC in part through over-activation of the PKC-MAPK-Fli-1 pathway.

Novel flavagline‐like compounds with potent Fli‐1 inhibitory activity suppress diverse types of leukemia

“…HEL cells undergoing erythroid differentiation with A1544 and A1545 compounds associated with Fli‐1 downregulation and upregulation of EKLF , SHIP 1, GATA 1, and p27 kip1 . In contrast, TPA blocks erythroid differentiation and promotes megakaryocytic maturation through activation of Fli‐1(↑) and downstream gene p21 cip1 and lineage markers CD 41/ CD 61, as described .…”

“…The MEK inhibitor U0126 blocked Fli-1 and the phosphorylation of eIF4E and ERK. This suppression was not In contrast, TPA blocks erythroid differentiation and promotes megakaryocytic maturation through activation of Fli-1(↑) and downstream gene p21 cip1 and lineage markers CD41/CD61, as described [27].…”

“…Fli-1 high or low expression is shown to play a critical role in commitment of MEP to become megakaryocytes or erythrocytes, respectively [10,52]. We previously showed that Fli-1 agonist compounds induced a megakaryocytic differentiation program in erythroleukemic cells by upregulating megakaryocytic-specific target genes, leading to leukemic inhibition [27]. Data in this study also revealed that drug-mediated Fli-1 inhibition resulted in a strong anti-leukemic effect, through induction of erythroid differentiation.…”

“…These studies led to identification of several small molecules/compounds targeting both DNA‐ and RNA‐binding activity of Fli‐1 . Our group has recently identified a class of Fli‐1 agonists that exhibit strong anticancer activity in vitro and in vivo . We demonstrated that these Fli‐1 agonists do not block erythroid differentiation, while they promote megakaryocytic differentiation of erythroleukemic progenitors, resulting in strong anticancer activity in vivo .…”

“…Our group has recently identified a class of Fli-1 agonists that exhibit strong anticancer activity in vitro and in vivo [27]. We demonstrated that these Fli-1 agonists do not block erythroid differentiation, while they promote megakaryocytic differentiation of erythroleukemic progenitors, resulting in strong anticancer activity in vivo [27]. In the current study, through screening compounds isolated from medicinal plants in China, we identified two novel inhibitors 4 0 -demethoxy-3 0 ,4 0methylenedioxyrocaglaol and rocaglaol with flavaglinelike structures that suppressed Fli-1 and EWS-FLI1 transactivation activity.…”