Iron: a global issue in hematology. Clara Camaschella, MD

Transcription

1 Iron: a global issue in hematology Clara Camaschella, MD Università Vita Salute San Raffaele e IRCCS San Raffaele - Milano Firenze, settembre 2015

2 Clara Camaschella I have nothing to disclose

3 Iron is essential in Hematology Heme Iron/Sulfur clusters Hb, cytochromes Iron deficiency anemia Anemia of chronic diseases Enzymes, IRP proteins Iron may be toxic in Hematology Fenton reaction: Fe 2+ + H 2 O 2 HO. + HO - + Fe 3+ Systemic iron overload (blood transfusions, ineffective erythropoiesis) Erythroid iron overload (sideroblastic anemia)

4 Cellular IRPs iron regulation handling ALAS2 ferritinophagy (Hentze, Galy, Muckenthaler & Camaschella, Cell 2010)

5 Iron distribution and regulation DMT1 FPN Enterocyte Fe IL-6 HEPCIDIN HEPCIDIN Senescent erythrocytes FPN Macrophage (Nemeth et al, Science 2004)

6 Systemic iron regulation: physiology (Hentze, Galy, Muckenthaler & Camaschella, Cell 2010)

7 Mechanisms of adaptation in iron deficiency anemia (Camaschella C, N Engl J Med 2015)

8 Hemochromatosis: a disorder of hepcidin liver pituitary heart skin Dietary iron Liver: fibrosis, cirrhosis, HCC Cardiac iron pancreas joints Skin pigmentation Joint disease Endocrine damage

9 Hepcidin regulation and rare disorders of the pathway liver iron circulating iron 6 Genetic iron deficiency IRIDA (Silvestri et al, Cell Met 2008; Finberg et al, Nat Genet 2008) Genetic Iron overload (Adapted from Hentze et al Cell, 2010)

10 Hepcidin suppression: iron deficiency, hypoxia and expanded erythropoiesis EPO EPO erythropoiesis? erythroid regulator(s) NTD b-thalassemia, CDA, Sideroblastic anemia. Proposed erythroid regulators: GDF15, TWSG1, shjv, HIF-1a, ERFE

11 A novel candidate erythroid regulator. * *a member of the C1q/TNF-related protein family (Kautz et al, Nat Genet 2014). a therapeutic target?

12 The Hbb th3/+ mouse: a model of thal intermedia Anemia and increased reticulocyte count RBC (10 6 /μl) Hb (g/dl) Hct (%) Retics (%) wt 9.5 ± ± ± ±0.4 Hbb th3/+ 6.6± ± ± ±0.9 Iron overload (Yang et al, PNAS 1995) Abnormal red cell morphology wt Hbb th3/+ (De Franceschi et al, Haematologica 2006) wt Hbb th3/+ (Yang et al, PNAS 1995) Low hepcidin Splenomegaly Ineffective erythropoiesis (Libani et al, Blood 2008) (Gardenghi et al, JCI 2010)

13 Genetic loss of Tmprss6 ameliorates the b-thalassemic phenotype Hbb th3/+ Tmprss6 -/- X Double mutants followed up to 6 months: Hepcidin increases and iron overload is prevented Ineffective erythropoeisis and anemia improve (Nai et al Blood 2012) TMPRSS6 gene silencing had similar effects in b-thal and HFE-hemochromatosis preclinical models (Schmidt et al, Blood 2013; Guo et al, JCO, 2013)

14 Inhibiting the hepcidin inhibitor (Camaschella C, NEJM 2013)

15 Positive effect of iron restriction in b-thalassemia Transferrin infusions (Hbb th3/+, Hbb th1/th1 ) (Li H et al, Nat Med 2010; Gelderman et al, Haematologica 2015) Use of minihepcidin (Preza et al, JCI 2011; Ramos Blood 2012; Casu et al, ASH 2014) Inhibition of the hepcidin inhibitor Tmpss6 (Schmidt et al, Blood 2013; Guo et al, JCO, 2013)

16 Tmprss6 -/- and Erfe -/- phenotype comparison Tmprss6 -/- IDA with high hepcidin Erfe -/- mouse Mild anemia (young mice) Tmprss6 -/-, Hbb thal3/+ Erfe -/-, Hbb thal3/+ Hb increase (1.5 g/dl) (Nai et al, Blood 2012) Hb unchanged (Kautz et al, Blood 2015)

17 Erythropoiesis and iron absorption in thalassemia ß-Thalassemia: ROS & Apoptosis GDF11 Decreased cell differentiation Epo EpoR Increased number of progenitor erythroid cells pjak2 pjak2 Erythroferrone GDF15 pstat5 Protection from apoptosis Cell Replication Hepcidin Increased iron absorption courtesy of Stefano Rivella

18 Implications Iron overload exacerbates ineffective erythropoiesis in β- thalassemia) Iron restriction (or manipulation of hepcidin pathway by hepcidin agonists ) may be exploited as a therapeutic tool in β-thalassemia Is iron a modifier of normal, effective erythropoiesis?

19 Why two transferrin receptors? TFR1 TFR2 Family member 1 Family member 2 Transmembrane glycoprotein Transmembrane glycoprotein: homologus to TFR1 extracellular domain IRE elements 3 UTR Binds diferric-tf at high affinity Inactivation in mouse: embryolethal ( severe iron deficiency) No IRE elements Binds diferric-tf at lower affinity than TFR1 Inactivation: in mice and humans iron overload Ubiquitous expression Restricted expression to hepatocytes, erythroid cells - EpoR interaction (Foretnikova et al, Blood 2010)

20 Sequential markers of erythroid development Erythroid Progenitors BFU-E CFU-E Terminal erythropoiesis TFR2 CD36 EPOR TFR1 (CD71) GlyA

21 Mutations of TFR2 Type 3 Hemocromatosis Autosomal recessive, rare Elevated TF saturation Hepatic iron load low hepcidin Early onset of iron overload Phlebotomy-responsive Oral iron Camaschella et al, Nat Genet 2000 Girelli et al, Haematologica 2011 Perls 250x Tfr2 KO Wild type Girelli et al, Gastroenterol 2002 Fleming et al, PNAS 2002

22 Tfr2 KO Murine models Type Phenotype Reference Tfr2 Y245X/ Y245X iron overload - low Hamp Fleming et al, 2002 Tfr2 KO iron overload - low Hamp Wallace et al, 2007 Roetto et al, 2010 Tfr2 LCKO iron overload - low Hamp Wallace et al 2007 Roetto et al, 2010 Tfr2 KO/Hfe KO severe - low Hamp iron overload Tmprss6 KO/Tfr2 KO Tmprss6 KO/Tfr2 LCKO iron deficiency - high Hamp erythrocytosis iron deficiency - high Hamp no erythrocytosis Wallace et al, 2009 Nai et al, 2014 Nai et al, 2014 (Nai et al Haematologica 2014)

23 Generation of a BM-specific Tfr2 -/- model BM Tfr2 -/- or wt wt Tfr2 -/- or wt BM (Nai, Lidonnici et al, Blood 2015)

24 Iron parameters and hepatic gene expression in Tfr2 BMKO mice (Nai, Lidonnici et al, Blood 2015)

25 Erythroid cells are increased and apoptosis decreased in the bone marrow of Tfr2 BMKO mice II III IV V (Nai, Lidonnici et al, Blood 2015)

26 Increased EPO sensitivity in Tfr2 BMKO mice TFR2 TFR2 Serum EPO ERFE I B I D

27 Tfr2 BMKO mice phenotype Erythrocytosis, increased erythroid precursors and decreased apoptosis. Normal Epo levels. Increased expression of Epo target genes in isolated erythroblasts 1. Tfr2 BMKO erythroblasts have increased Epo sensitivity 2. Deletion of erythroid Tfr2 mimics iron-deficiency

28 TFR2 links erythropoiesis and hepcidin TFR2 is stabilized on plasma membrane by the diferric transferrin ligand (Johnson et al, Mol Biol Cell 2007; Pagani et al, Haematologica 2015)

29 Does TFR2 modulate the Epo sensitivity in ID? Lack of membrane TFR2 stabilization in ID. Degradation? TFR2 releases a soluble form in iron deficiency in vitro (Pagani et al Haematologica 2015)

30 Conclusion TFR2 is a component of a novel iron-sensing mechanism that adjusts erythropoiesis according to iron availability, by modulating the erythroblast Epo sensitivity Loss of TFR2 in iron deficiency (likely by stfr2 shedding) optimizes eryhtropoiesis and increases ERFE, leading to hepcidin suppression and iron acquisition TFR2 links hepcidin and iron homeostasis to erythropoiesis TFR2 = liver sensor of circulating iron (excess) TFR2 = erythroid sensor of circulating iron (deficiency)

31 Hepcidin upregulation by inflammatory cytokines... Inflammation Activin-b IL-6 decreases iron available for pathogens HJV BMPR IL-6R TMPRSS6 P P SMAD4 SMAD1/5/8 STAT3 JAK2 P explains iron maldistribution, macrophage sequestration and iron-restricted erythropoiesis in anemia of inflammation (ACD) P P BRE BRE SRE HAMP

32 Hepcidin antagonists (modified from Fung & Nemeth Haematologica 2013) Class 1 Inhibitors of hepcidin production Class 2 Hepcidin neutralizing peptides BMP binders (shjv, heparin) BMPR inhibitors (LDN ) HJV antisense oligonucleotides Hepcidin antisense oligonucleotides Anticytokines (IL6, IL6R, TNFalpha) ESA Antihepcidin monoclonal antibodies Anticalin Spiegelmers Class 3 Interfering with hepcidin/fpn binding Anti-FPN monoclonal antibodies Thiol modifiers

33 Iron handling by tumor cells A) Normal cell Cancer cell (Torti & Torti. Nat Rev Cancer 2013) Role of hepcidin/ferroportin axis in cancer Breast cancer: Pinnix et al. Sci Transl Med Prostate cancer: Tesfay et al. Cancer Res Multiple Myeloma: Gu et al. Cancer Res Campanella et al Haematologica 2013 Bordini et al, Haematologica 2015

34 Acknowledgments Vita-Salute University and San Raffaele Scientific Institute Tiget, HSR Giuliana Ferrari M. Rosa Lidonnici Giacomo Mandelli (PRIN) - Rome Antonella Roetto, Torino Domenico Girelli, Verona Paolo Arosio, Brescia Institut Cochin, Université Paris Descartes Frédérique Verdier Patrick Mayeux Catherine Lacombe Carlos Lopez Otin, Oviedo