Correcting β-thalassemia by combined therapies that restrict iron and modulate erythropoietin activity

Abstract

ß-thalassemia intermedia is a disorder characterized by ineffective erythropoiesis (IE), anemia, splenomegaly and systemic iron overload. Novel approaches are being explored based on the modulation of pathways that reduce iron absorption (i.e. using hepcidin activators like Tmprss6-antisense oligonucleotides (ASO)) or increase erythropoiesis (by erythropoietin (EPO) administration or by modulating the ability of transferrin receptor 2 (Tfr2) to control red blood cell (RBC) synthesis). Targeting Tmprss6 mRNA by Tmprss6-ASO was proven to be effective in improving the IE and splenomegaly by inducing iron restriction. However we postulated that combinatorial strategies might be superior to single therapies. Here we combined Tmprss6-ASO with EPO administration or removal of a single Tfr2 allele in the bone marrow of animals affected by ß-thalassemia intermedia (Hbbth3/+). EPO administration alone or removal of a single Tfr2 allele increased hemoglobin levels and RBCs. However, EPO or Tfr2 single allele deletion alone, respectively, exacerbated or did not improve the splenomegaly in ß-thalassemic mice. To overcome this issue, we postulated that some level of iron restriction (by targeting Tmprss6) would improve the splenomegaly while preserving the beneficial effects on RBC production mediated by EPO or Tfr2 deletion. While administration of Tmprss6-ASO alone improved the anemia, combination of Tmprss6-ASO+EPO or Tmprss6-ASO+Tfr2 single allele deletion showed significantly higher hemoglobin levels as well as reduction of splenomegaly. In conclusion, our results clearly indicate that these combinatorial approaches are superior to single treatments in ameliorating the IE and anemia in ß-thalassemia and could provide guidance to translate some of these approaches into viable therapies.

Global loss of Tfr2 with concomitant induced iron deficiency greatly ameliorates the phenotype of a murine thalassemia intermedia model

“…We do not know if this phenotypic improvement would be sustained over a longer treatment period. Of note, work by Casu et al published during the preparation of this manuscript demonstrate not only a comparable amelioration of the Hbb th3/+ hematological phenotype, but also a similar decrease in splenomegaly using Tmprss6‐ASO with removal of a single Tfr2 allele in the bone marrow of Hbb th3/+ mice 35 …”

“…Because we employed the Tfr2 Y245X/Y245X genetic null model we cannot determine if the enhanced effectiveness of the Tmprss6 siRNA treatment requires a loss of functional Tfr2 in the liver, bone marrow, or both tissues. However, it was recently shown by others that loss of functional Tfr2 in bone marrow alone, along with pharmacological iron restriction, is able to ameliorate the Hbb th3/+ phenotype 35 . It may be of interest to confirm that loss of Tfr2 specifically in hepatocytes does not enhance effective erythropoiesis in a thalassemic model under mild iron restriction.…”

Transferrin Receptors in Erythropoiesis

“…In 2018, Artuso et al found that the bone-marrow specific deletion of Tfr2 moderately lessens the symptoms of β-thalassemia in their Hbb th3/+ mouse model, when combined with an iron restricted diet [ 87 ]. More recently, a study from Casu et al combined Tfr2 knockout with iron depletion by targeting the negative regulator of Hepcidin ( Tmprss6 ) which shows superior correction of anemia in Hbb th3/+ mice [ 88 ]. In both studies, the number of RBCs and the hemoglobin level are increased and iron deposits in the liver, spleen, and kidney are decreased.…”

“…As iron deprivation seems to ameliorate the symptoms, this is an alternate explanation to consider. These studies in the context of β-thalassemia intermedia have been done with the same mouse models [ 87 , 88 ]. However, varying results have been obtained with Tfr2 knockout depending on the mouse model before [ 10 , 59 , 61 , 62 ], probably partly due to differing basal level of iron [ 63 ].…”

Iron Through the Prism of Haematology

“…For example, administration of short peptides that mimic the activity of hepcidin have improved ineffective erythropoiesis, anaemia and iron overload in thalassaemic mice 44 . This effect can also be achieved in thalassaemic mice by targeting Tmprss6 mRNA with Tmprss6 antisense oligonucleotides alone or combined with other modalities 45 . Clinical trials have started where exogenous hepcidin mimetics, administered to humans, have been shown to lower transferrin saturation, 46 although the effect in thalassaemia and MDS is not yet known.…”