Targeting Transferrin Receptor 2: A Novel "Erythropoiesis-Stimulating Approach" in a Model of Anemia of Chronic Kidney Disease
Authors: Olivari Violante , Lidonnici Maria Rosa , Pettinato Mariateresa , Silvestri Laura , Ferrari Giuliana , Camaschella Clara , Nai Antonella 
Published: February 2020

Introduction
Anemia is a common and invalidating complication of chronic kidney disease (CKD) mainly due to the impaired erythropoietin (EPO) production that parallels the progression of kidney failure. The current treatment, based on erythropoiesis-stimulating agents (ESA), is far from optimal, because of potential off-target side effects. The same holds true for HIF-stabilizers tested in phase 3 clinical trials. For this reason, the identification of agents that selectively boost erythropoiesis would be of great benefit.
Transferrin Receptor 2 (TFR2) is a protein expressed in hepatocytes, where it modulates iron homeostasis activating hepcidin production, and in erythroid cells, where it acts as a partner of erythropoietin (EPO) receptor decreasing EPO signaling. Tfr2 deletion in the liver causes iron overload due to low hepcidin levels, while its deletion in erythroid compartment enhances erythropoiesis through increased EPO sensitivity.
Aim
The study aims at exploring whether erythroid TFR2 is a potential therapeutic target in anemia of CKD, characterized by reduced EPO production and function.
Methods
CKD was induced by feeding animals an adenine-rich diet for 8 weeks. The protocol was applied to both wild-type animals, to set-up the system, and to mice with genetic bone marrow Tfr2 deletion (Tfr2BMKO). Complete blood count (CBC) and serum urea were evaluated every 2 weeks during the entire protocol. At the end of the 8 weeks animals were sacrificed and a complete phenotypic analysis of hematological parameters, renal damage and iron homeostasis was performed.
Results
Wild-type mice fed the adenine diet showed renal damage and inflammation, anemia and iron restriction, recapitulating the main features of human CKD.
Renal damage and iron restriction were similar between Tfr2BMKO and control mice, excluding a differential effect of the diet on the two genotypes. Tfr2BMKO mice maintained higher red blood cell count relative to controls for the entire timespan. Hemoglobin levels were higher in Tfr2BMKO mice for 6 weeks, while reached levels of controls at 8 weeks. Based on these findings we conclude that BM Tfr2deletion, increasing EPO sensitivity of erythroid cells, improves erythropoiesis and anemia until iron levels remains adequate.
Conclusions
We confirm that the adenine-induced murine model of CKD can be a suitable tool for the study of the pathophysiology of renal anemia and for the identification and validation of novel potential therapeutic approaches.
More importantly, our results suggest that targeting Tfr2 could become a novel approach to ameliorate anemia of CKD. Given the TFR2 restricted expression, its inactivation would enhance EPO responsiveness selectively in erythroid cells, minimizing the risk of side effects. Targeting erythroid Tfr2 might be considered a novel "erythropoiesis-stimulating approach", likely applicable to other forms of anemia due to insufficient EPO stimulation and response.

Disclosures
Camaschella: Celgene: Consultancy; Vifor Iron Core: Consultancy; Novartis: Consultancy.