Iron Trafficking and Distribution in Transfusional Overload: Insights From Comparing Diamond Blackfan Anemia with Sickle Cell Disease and Thalassemia
Authors: Porter John B. , Walter Patrick B , Neumayr Lynne D , Evans Patricia , Weyhmiller Marcela G , Harmatz Paul , Wood John C , Miller Jeffery L. , Weiss Guenter , Yuan Qing , Wang Zhiyue J , Grosse Regine , Schoennagel Bjoern P , Meerpohl Joerg , Fischer Roland , Vichinsky Elliott 
Published: October 2019

Abstract
Abstract 995

Background:
β-thalassemia major (TM) is the paradigm for chronic transfusional iron overload, in which the extra-hepatic organ failure is best described. In Sickle Cell Disease (SCD), these consequences appear later and at a lower frequency. In chronically transfused Diamond Blackfan Anemia (DBA), extra-hepatic iron overload, although less well documented, appears to occur early and at high frequency. A Multicenter Study of Iron Overload (MCSIO) aims to explore how key candidate factors affect iron distribution; including inflammation, ineffective erythropoiesis, level of iron overload, and hepcidin synthesis. Plasma non-transferrin bound iron (NTBI) could be a key mechanism by which iron is delivered to tissues and may determine the propensity for extra-hepatic iron distribution. Here we focus on how markers of ineffective erythropoiesis (IE) and erythron expansion impact iron distribution, with particular reference to NTBI and iron distribution determined by MRI.

Methods:
Iron-overloaded patients (5 TM, 5 SCD, and 5 DBA) with ferritin > 1500 g/dl or LIC > 7 mg/g dry wt, age ≥16, age 0 to 9 at initiation of transfusion and 10 to 20 years of transfusion exposure were enrolled from 3 sites in the US and Europe. 5 non-transfused healthy controls were also enrolled. A detailed medical, transfusion and chelation history were obtained with standardized MRI evaluations for hepatic, cardiac, and pituitary iron deposition. Fasting, early morning blood samples were obtained one day prior to transfusion. Chelation was held for 72 hours prior to each sample.

Results:
Results are shown in the table as median values. DBA patients had the highest NTBI prior to transfusion despite having the lowest ferritin and LIC levels. GDF15 levels were highest in TM, with similar levels in SCD and DBA. EPO levels were nearly two orders of magnitude higher in DBA than TM or SCD. DBA patients also had the highest median cardiac R2*; two patients showing values above the control range. Whereas the median pituitary R2 in DBA was not above control, two of the patients had the highest R2 values, suggesting heavy iron deposition. EPO values in DBA are nearly two orders of magnitude higher that in SCD or TM despite similar pre-transfusion Hb values. GDF15 values are approximately three times controls, while soluble transferrin receptors (sTfR) values are almost undetectable. With SCD, no patients had increased cardiac iron loading, despite median SF and LIC being the highest in this group. Surprisingly all SCD patients had pituitary R2 values above the upper limit of normal. 1 TM patient had increased cardiac R2* whereas three had increased pituitary iron. In TM, NTBI was strongly correlated with GDF15 (Pearson's Rho=0.93) but in DBA, GDF15 was inversely correlated with NTBI (-.95).

Conclusions:
High GDF15 levels have been reported in conditions associated with IE, such as TM, but not in DBA. GDF15 reputedly suppresses hepcidin synthesis, thereby increasing iron absorption and potentially NTBI levels. The increased GDF15 in DBA, while sTfr remain less than controls, suggests that erythropoietic precursors do not reach the stage where sTfr are expressed and that this occurs at a later differentiation stage than GDF15. Increasing NTBI in TM with increasing GDF15 is consistent with IE contributing to NTBI formation, but the lack of this relationship in DBA suggest another mechanism for high NTBI. As the erythron is destroyed at a pre-hemoglobinised stage in DBA, IE would not contribute directly to NTBI formation. However, the extremely high EPO levels in DBA may inhibit hepcidin synthesis, as in other conditions, thereby increasing NTBI. This in turn may account for the extra-hepatic iron distribution demonstrated by MRI in DBA. The increased pituitary iron without cardiac loading in the heavily loaded SCD patients suggests that with prolonged exposure to heavy iron overload, the pituitary iron loading may be the first indicator of extra-hepatic deposition.

Disclosures:
Porter: Novartis: Consultancy, Research Funding. Walter:Novartis: Research Funding. Harmatz:Novartis: Research Funding; Ferrokin: Research Funding. Wood:Ferrokin Biosciences: Consultancy; Shire: Consultancy; Apotex: Consultancy, Honoraria; Novartis: Honoraria, Research Funding. Vichinsky:Novartis: Consultancy, Research Funding; ApoPharma: Consultancy, Research Funding; ARUP Research lab: Research Funding.