Forskolin Sensitizes Human Acute Myeloid Leukemia Cells to H3K27me2/3 Demethylases GSKJ4 Inhibitor via Protein Kinase A

Abstract

Acute myeloid leukemia (AML) is an aggressive hematological malignancy occurring very often in older adults, with poor prognosis depending on both rapid disease progression and drug resistance occurrence. Therefore, new therapeutic approaches are demanded. Epigenetic marks play a relevant role in AML. GSKJ4 is a novel inhibitor of the histone demethylases JMJD3 and UTX. To note GSKJ4 has been recently shown to act as a potent small molecule inhibitor of the proliferation in many cancer cell types. On the other hand, forskolin, a natural cAMP raising compound, used for a long time in traditional medicine and considered safe also in recent studies, is emerging as a very interesting molecule for possible use in cancer therapy. Here, we investigate the effects of forskolin on the sensitivity of human leukemia U937 cells to GSKJ4 through flow cytometry-based assays (cell-cycle progression and cell death), cell number counting, and immunoblotting experiments. We provide evidence that forskolin markedly potentiates GSKJ4-induced antiproliferative effects by apoptotic cell death induction, accompanied by a dramatic BCL2 protein down-regulation as well as caspase 3 activation and PARP protein cleavage. Comparable effects are observed with the phosphodiesterase inhibitor IBMX and 8-Br-cAMP analogous, but not by using 8-pCPT-2′-O-Me-cAMP Epac activator. Moreover, the forskolin-induced enhancement of sensitivity to GSKJ4 is counteracted by pre-treatment with Protein Kinase A (PKA) inhibitors. Altogether, our data strongly suggest that forskolin sensitizes U937 cells to GSKJ4 inhibitor via a cAMP/PKA-mediated mechanism. Our findings provide initial evidence of anticancer activity induced by forskolin/GSKJ4 combination in leukemia cells and underline the potential for use of forskolin and GSKJ4 in the development of innovative and effective therapeutic approaches for AML treatment.

The KDM Inhibitor GSKJ4 Triggers CREB Downregulation via a Protein Kinase A and Proteasome-Dependent Mechanism in Human Acute Myeloid Leukemia Cells

“…More recently, Li et al (25) have highlighted the therapeutic potential of GSKJ4 also for AML. According to the latter findings, we also provided evidence in which GSKJ4 causes growth inhibition in myeloid leukemia cells (30).…”

“…As JMJD3 and UTX demethylase inhibitor, we have shown that GSKJ4 increases H3K27me3 status in AML cells. Moreover, GSKJ4 has recently been shown to counteract cell proliferation in different tumor types, including AML (16)(17)(18)(19)(20)(21)(22)(23)(24)(25)30).…”

GPRC5A facilitates cell proliferation through cell cycle regulation and correlates with bone metastasis in prostate cancer

“…19 The small molecule forskolin can promote CREB phosphorylation. 40 Here, both forskolin treatment and GPRC5A KO suppressed proliferation of prostate cancer cells ( Fig. 3), suggesting that the cAMP-dependent pathway has negative regulatory effects on prostate cancer cell proliferation.…”

“…Therefore, GPRC5A may form a negative feedback loop in which the phosphorylation of CREB is suppressed . The small molecule forskolin can promote CREB phosphorylation . Here, both forskolin treatment and GPRC5A KO suppressed proliferation of prostate cancer cells (Fig.…”

The Functions of the Demethylase JMJD3 in Cancer

“…The latter protein is phosphorylated on Ser-133 by a protein kinase A (PKA) before being directed to the proteasome [ 73 ]. Moreover, it seems that the association of GSK-J4 with a natural cAMP raising compound (forskolin) potentiates the anti-proliferative effects of GSK-J4 via PKA in some AML cells (U937 cells), leading to B-cell lymphoma 2 protein (BCL2) downregulation and caspase 3 activation [ 74 ].…”