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

Yankai Zhang, Alireza Paikari, Pavel Sumazin, Carly C. Ginter Summarell, Jacy R. Crosby, Eric Boerwinkle, Mitchell J. Weiss and Vivien A. Sheehan

Key points

  • Functional studies in human primary erythroid progenitor cells support a role for FOXO3 in γ-globin regulation.

  • Metformin treatment of human primary erythroid progenitor cells increases fetal hemoglobin in a partially FOXO3-dependent manner.

Abstract

Induction of red blood cell fetal hemoglobin (HbF, α2γ2) ameliorates the pathophysiology of sickle cell disease (SCD) by reducing the concentration of sickle hemoglobin (HbS, α2βS2) to inhibit its polymerization. Hydroxyurea (HU), the only FDA-approved drug for SCD, acts in part by inducing HbF, but is not fully effective, reflecting the need for new therapies. Whole exome sequence analysis of rare genetic variants in SCD patients identified FOXO3 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 (HSPCs) induced to undergo erythroid differentiation. FOXO3 gene silencing reduced γ-globin RNA levels and HbF levels in erythroblasts, while 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 non-erythroid cells, caused dose-related, FOXO3-dependent, increases in both percentage (%) HbF protein and the fraction of HbF-immunostaining cells (F-cells). Combined HU and metformin treatment induced HbF additively and reversed the erythroid maturation arrested caused by HU treatment alone. HbF induction in erythroid precursors by metformin 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.

  • Submitted November 15, 2017.
  • Accepted May 25, 2018.