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Bitopertin, a selective oral GLYT1 inhibitor, improves anemia in a mouse model of β-thalassemia
Alessandro Matte, … , Carlo Brugnara, Lucia De Franceschi
Alessandro Matte, … , Carlo Brugnara, Lucia De Franceschi
Published October 8, 2019
Citation Information: JCI Insight. 2019;4(22):e130111. https://doi.org/10.1172/jci.insight.130111.
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Research Article Hematology

Bitopertin, a selective oral GLYT1 inhibitor, improves anemia in a mouse model of β-thalassemia

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Abstract

Anemia of β-thalassemia is caused by ineffective erythropoiesis and reduced red cell survival. Several lines of evidence indicate that iron/heme restriction is a potential therapeutic strategy for the disease. Glycine is a key initial substrate for heme and globin synthesis. We provide evidence that bitopertin, a glycine transport inhibitor administered orally, improves anemia, reduces hemolysis, diminishes ineffective erythropoiesis, and increases red cell survival in a mouse model of β-thalassemia (Hbbth3/+ mice). Bitopertin ameliorates erythroid oxidant damage, as indicated by a reduction in membrane-associated free α-globin chain aggregates, in reactive oxygen species cellular content, in membrane-bound hemichromes, and in heme-regulated inhibitor activation and eIF2α phosphorylation. The improvement of β-thalassemic ineffective erythropoiesis is associated with diminished mTOR activation and Rab5, Lamp1, and p62 accumulation, indicating an improved autophagy. Bitopertin also upregulates liver hepcidin and diminishes liver iron overload. The hematologic improvements achieved by bitopertin are blunted by the concomitant administration of the iron chelator deferiprone, suggesting that an excessive restriction of iron availability might negate the beneficial effects of bitopertin. These data provide important and clinically relevant insights into glycine restriction and reduced heme synthesis strategies for the treatment of β-thalassemia.

Authors

Alessandro Matte, Enrica Federti, Michael Winter, Annette Koerner, Anja Harmeier, Norman Mazer, Tomas Tomka, Maria Luisa Di Paolo, Luigia De Falco, Immacolata Andolfo, Elisabetta Beneduce, Achille Iolascon, Alejandra Macias-Garcia, Jane-Jane Chen, Anne Janin, Christhophe Lebouef, Franco Turrini, Carlo Brugnara, Lucia De Franceschi

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Figure 5

Bitopertin beneficially affects in vitro β-thalassemic erythropoiesis, with reduction of protoporphyrin IX, as a marker of heme restriction.

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Bitopertin beneficially affects in vitro β-thalassemic erythropoiesis, w...
(A) Cell proliferation of β-thalassemic erythroid precursors derived by in vitro liquid culture of CD34+ cells isolated from peripheral blood of β-thal subjects with or without bitopertin 10 nM (n = 3). Arrows indicate when bitopertin 10 nM was added to the culture medium (see Supplemental Figure 6B for dose-response data on cell viability in normal erythropoiesis). Data are presented as mean ± SD; *P < 0.05 compared with vehicle-treated β-thal cells; Wilcoxon’s signed-rank test for multiple comparisons. (B) Effect of bitopertin (10 nM) on maturation pattern of β-thalassemic erythroid precursors from a single patient in triplicate. Representative flow cytometric plot and box plots of the differentiation pattern of β-thalassemic erythroid precursors at 14 days of culture with or without bitopertin (10 nM). The following surface markers were used: CD36, glycophorin-A, and CD71. This allows for the identification of the following homogenous cell populations: pro-erythroblasts (Pro-E), basophilic erythroblasts corresponding to intermediate erythroblasts (Int-E), and polychromatic and orthochromatic erythroblasts as late erythroblasts (Late-E) (see also refs. 4, 44). Data are shown as median and minimum/maximum (n = 4); *P < 0.05 compared with vehicle-treated β-thalassemic cells; Wilcoxon’s signed-rank test for multiple comparisons. (C) Effect of bitopertin on the index of erythroid maturation of β-thalassemic cells from 4 separate individuals at 9 and 14 days of cell culture. *P < 0.05 compared with vehicle-treated β-thalassemic cells; Mann-Whitney U test with multiple-comparisons correction. (D) Effect of bitopertin (10 nM) on the amount of annexin V+ cells during β-thalassemic erythroid maturation from 4 separate individuals at 9 and 14 days of cell culture. *P < 0.05 compared with vehicle-treated β-thalassemic cells, Mann-Whitney U test with multiple-comparison correction. (E) Intracellular content of protoporphyrin IX in healthy and β-thalassemic cells (4.5 × 106 cells) after 14 days of cell cultures with and without bitopertin (10 nM). Data are presented as median and minimum/maximum (n = 4) from each group; *P < 0.05 compared with vehicle; Mann-Whitney U test with multiple-comparisons correction.

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