Fetal hemoglobin induction by monomethylfumarate: relevance to prevention and treatment of sickle cell retinopathy (SR)

Abstract

Purpose: SR is a most debilitating and unfortunately common complication of sickle cell disease (SCD). Abnormal hemoglobin (HbS) polymerization in red blood cells and consequent vascular dysfunction is a primary causative factor; clinical and experimental studies support the critical involvement of other cellular and molecular factors. RPE cells were reported recently to produce Hb however this phenomenon was not evaluated in the context of retinal pathology in SCD. Here, using ARPE-19 and primary RPE cells established from HbAA (normal)- and HbSS (sickle)-expressing Townes humanized mice we (a) evaluated globin gene expression and Hb production in normal and sickle RPE, and (b) validated monomethylfumarate (MMF) as an effective inducer of γ-globin expression/fetal Hb (HbF) production in retinal and erythroid cells and therefore, as a potential therapy for preventing and treating retinal AND systemic complications of SCD.

Methods: ARPE-19 and HbAA- and HbSS- expressing primary RPE cells were treated with various concentrations of MMF, followed by analysis of α-, β- and γ-globin expression and HbF production by qPCR, western blotting, FACS and immunofluorescence. Hydroxyurea (HU), at present the only FDA-approved compound for SCD treatment, was included as a positive control. Parallel studies were performed using KU812 and primary human erythroid progenitor cells.

Results: RPE cells express α-, β- and γ-globin mRNA and consequently synthesize normal (ARPE-19 and HbAA primary RPE) and sickle (HbSS primary RPE) Hb. MMF induces, in a time- and dose-dependent manner, γ-globin expression and HbF production in all cell types evaluated, and in retina following its delivery intravitreally to live mice. HbF induction by MMF was 2-3 -fold higher than that by HU.

Conclusions: HbF-inducing therapy is at present the best strategy for preventing and treating SCD-related complications. However, little is known regarding the functional relevance of Hb production in retina by non-erythroid cells or the impact of HbF-inducing therapies in this tissue. Our present finding that MMF induces γ-globin expression and HbF production is novel and supports the possible use of this compound in SCD/SR. The fact that FDA-approved formulations in which MMF is the main bioactive ingredient are readily available enhance greatly the translational potential of this study.

Keywords: 701 retinal pigment epithelium • 636 pathobiology • 695 retinal degenerations: cell biology