Metformin induces FOXO3-dependent fetal hemoglobin production in human primary erythroid cells

Abstract

Induction of red blood cell (RBC) fetal hemoglobin (HbF; α2γ2) ameliorates the pathophysiology of sickle cell disease (SCD) by reducing the concentration of sickle hemoglobin (HbS; α2β2) to inhibit its polymerization. Hydroxyurea (HU), the only US Food and Drug Administration (FDA)-approved drug for SCD, acts in part by inducing HbF; however, it is not fully effective, reflecting the need for new therapies. Whole-exome sequence analysis of rare genetic variants in SCD patients identified as a candidate regulator of RBC HbF. We validated these genomic findings through loss- and gain-of-function studies in normal human CD34 hematopoietic stem and progenitor cells induced to undergo erythroid differentiation. gene silencing reduced γ-globin RNA levels and HbF levels in erythroblasts, whereas overexpression of FOXO3 produced the opposite effect. Moreover, treatment of primary CD34 cell-derived erythroid cultures with metformin, an FDA-approved drug known to enhance FOXO3 activity in nonerythroid cells, caused dose-related FOXO3-dependent increases in the percentage of HbF protein and the fraction of HbF-immunostaining cells (F cells). Combined HU and metformin treatment induced HbF additively and reversed the arrest in erythroid maturation caused by HU treatment alone. HbF induction by metformin in erythroid precursors was dependent on FOXO3 expression and did not alter expression of BCL11A, MYB, or KLF1. Collectively, our data implicate FOXO3 as a positive regulator of γ-globin expression and identify metformin as a potential therapeutic agent for SCD.

Spermidine Suppresses Inflammatory DC Function by Activating the FOXO3 Pathway and Counteracts Autoimmunity

“…Under conditions of high glucose, AKT phosphorylates FOXO3 at three conserved residues, Thr32, Ser253, and Ser315. This leads to FOXO3 binding to 14-3-3, exposure of the nuclear localization signal, transporting to the cytosol and being degraded, causing reduced expression of FOXO3 target genes (Zhang et al, 2018). When ATP is low, FOXO3 is phosphorylated by AMPK at Ser413 and exhibits increased activity (Lee et al, 2013;Zhang et al, 2018).…”

“…This leads to FOXO3 binding to 14-3-3, exposure of the nuclear localization signal, transporting to the cytosol and being degraded, causing reduced expression of FOXO3 target genes (Zhang et al, 2018). When ATP is low, FOXO3 is phosphorylated by AMPK at Ser413 and exhibits increased activity (Lee et al, 2013;Zhang et al, 2018). Foxo3a-deficient DCs express higher levels of proinflammatory cytokines (Dejean et al, 2009;Lee et al, 2013;Litvak et al, 2012;Thompson et al, 2015;Zhang et al, 2018).…”

“…When ATP is low, FOXO3 is phosphorylated by AMPK at Ser413 and exhibits increased activity (Lee et al, 2013;Zhang et al, 2018). Foxo3a-deficient DCs express higher levels of proinflammatory cytokines (Dejean et al, 2009;Lee et al, 2013;Litvak et al, 2012;Thompson et al, 2015;Zhang et al, 2018). For these reasons, we hypothesized that FOXO3 might be the critical transcriptional factor in the process.…”

Stability of fetal hemoglobin levels in patients receiving metformin therapy

“…Recently, in-vitro studies using progenitor-derived erythroid cells from normal individuals or sickle cell disease (SCD) patients, have identified metformin, an oral hypoglycemic drug, as a promising agent to increase fetal hemoglobin (HbF) levels [1][2][3] . The mechanisms through which metformin increases HbF involve Forkhead box O transcription factor-3 (FOXO3) and adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathway, which is key in controlling HbF regulation 3 .…”

“…The mechanisms through which metformin increases HbF involve Forkhead box O transcription factor-3 (FOXO3) and adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathway, which is key in controlling HbF regulation 3 . In addition, these in-vitro experiments have demonstrated that induction of HbF with metformin is somehow superior to hydroxyurea and combination of the two agents show additive effects 2 . In light of these findings, a pilot study with metformin in patients with hemoglobinopathies is presently enrolling (NCT02981329, ClinicalTrials.…”