Genetic Iron Overload Aggravates and Pharmacological Iron Restriction Improves MDS Pathophysiology in a Preclinical Mouse Model

Background: Patients with myelodysplastic syndromes (MDS) are prone to develop iron overload as a consequence of ineffective erythropoiesis and chronic transfusion therapy. Although iron overload is a common feature in MDS, it remains unclear whether and how iron excess is detrimental for MDS pathophysiology.

Aims: This study aimed at characterizing altered iron homeostasis in a preclinical MDS mouse model, understanding the molecular mechanisms underlying iron toxicity and unraveling the potential therapeutic value of pharmacologic iron restriction in this disease condition.

Methods: To this end, we took advantage of complementary approaches and analyzed the effect of iron overload obtained through genetic activation of the iron exporter ferroportin (FPN^C326S), and iron restriction achieved through the administration of the FPN inhibitor vamifeport in NUP98-HOXD13 MDS mice, starting at 3 or 5 months of age.

Results: At steady-state MDS mice develop an iron overload phenotype, hallmarked by low hepcidin levels, elevated serum iron and transferrin saturation, non-transferrin-bound iron (NTBI) formation and tissue iron deposition. This is associated with anemia and low WBCs in the peripheral blood. FPN^C326S MDS mice show an aggravated iron phenotype, with further elevated NTBI and tissue iron accumulation and still inappropriately low hepcidin levels. Vamifeport administration in MDS mice reduced serum iron (194.8 vs 141.8 mg/dl; P<0.05) and NTBI formation (0.62 vs 0 μM; P<0.05) and prevented tissue iron loading. While iron excess in FPN^C326S MDS mice did not improve erythropoiesis and hematologic parameters in the peripheral blood, iron restriction by vamifeport significantly ameliorated anemia (Hb 9.5 vs 11.4 g/dl; RBC 5.4 vs 6.9x10⁶ cells/ml; P<0.05) and maturation of bone marrow as well as splenic RBCs in MDS mice. The improved ineffective erythropoiesis was associated with reduced oxidative stress and apoptosis in erythroid progenitors. Importantly, we observed a major role of the iron status in myeloid expansion in MDS. The number of immature myeloid blasts and myeloid progenitors in the bone marrow of MDS mice were aggravated by iron overload in FPN^C326S MDS mice and attenuated by iron restriction in vamifeport-treated MDS mice compared to control MDS animals. Myeloid bias, monitored as percentage bone marrow myeloid cells, was exacerbated by iron overload (54.3 vs 43.1% bone marrow mononuclear cells - BMNCs) and alleviated by iron restriction (67.5 vs 74.8% BMNCs; P<0.05). This was associated with increased and reduced inflammatory activation of bone marrow macrophages in FPN^C326S MDS and vamifeport-treated MDS mice, respectively. This observation demonstrates a role for NTBI in driving the reported pro-inflammatory activation of MDS monocytes/macrophages (Fuster JJ et al, Science 2017; Jaiswal S et al, NEJM 2017; Cull AH et al, Exp Hematol 2017) and suggests that this iron-driven mechanism contributes to myeloid expansion. Overall, these alterations translated into a faster and delayed transition to AML in iron-overloaded and iron-restricted MDS mice, respectively. Finally, iron overload aggravated and iron restriction alleviated ROS formation, DNA damage and pyroptotic cell death in HSPCs. By reducing the intracellular labile iron pool, vamifeport improved HSPC quality and survival, partially rescuing the stem cell pool size (0.16 vs 0.38% Lin^-BMNCs; P<0.05) in MDS mice. The ameliorated HSPC pool, reduced myeloid bias and inflammation, and attenuated ineffective erythropoiesis resulted in improved hematologic parameters for more than 3 months, and a significant survival increment in iron-restricted compared to control MDS mice (>45 days; P=0.037). We further demonstrate that a subgroup of mice can benefit from late iron restriction by vamifeport administration at 5 months of age, resulting in improved anemia and prolonged survival.

Conclusion: Our results show for the first time in a preclinical mouse model of MDS that iron excess driven by ineffective erythropoiesis has pathological implication in transfusion-independent MDS. These effects are likely aggravated by transfusions causing additional iron overload. Finally, iron restriction achieved through pharmacologic FPN inhibition by the oral FPN inhibitor vamifeport significantly improves MDS pathophysiology, uncovering the therapeutic potential of early prevention of NTBI formation in MDS.

Dürrenberger:Vifor (International) Ltd: Current Employment. Manolova:Vifor (International) Ltd: Current Employment. Vinchi:Vifor (International) Ltd: Research Funding; PharmaNutra S.p.A: Research Funding; Silence Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding.