GENETIC/METABOLIC EFFECT OF IRON
METABOLISM & RARE ANEMIAS
3rd Pan-European Conference on
Haemoglobinopathies & Rare Anaemias
Limassol, 24 – 26 October 2012
Clara Camaschella
Università Vita-Salute San Raffaele
IRCCS San Raffaele, Milano, Italy
DISCLOSURE
Clara Camaschella
Università Vita-Salute - IRCCS San Raffaele, Milano
NO DISCLOSURE
Iron for erythropoiesis
Daily iron needs for Hb synthesis of maturing erythroblasts: 25 mg
Advances in iron metabolism research….
Systemic iron regulation
(Hentze et al, Cell 2010)
Iron and hepcidin levels in Val Borbera individuals
(Traglia et al, J Med Genet, 2011)
Hepcidin inhibition in iron deficiency,
hypoxia and erythropoiesis expansion
Proposed inhibitors:
Epo,
Hif1-alpha,
s-HJV,
GDF15/TWSG1
TMPRSS6/Matriptase 2
(Hentze et al, Cell 2010)
1. Defects of iron absorption
IRIDA - OMIM #206200
Iron refractory iron deficiency anemia:
Autosomal recessive disorder due to TMPRSS6
(matriptase-2) mutations
Moderate anemia since childhood, severe
microcytosis
Extremely low iron and transferrin saturation
Normal serum ferritin
Inappropriately high hepcidin levels
Refractory to oral and partially refractory to iv iron
(Finberg et al, Nat Genet 2008, Sem Hematol 2009)
TMPRSS6/Matriptase-2: the hepcidin
inhibitor
Matriptase-2 is encoded by TMPRSS6 gene on chr 22
RNA expression: liver (kidney, olfactory epithelium)
Protein: 811 amino acid type II transmembrane serine
protease synthesized as an inactive zymogen
(TTPS family: enteropeptidase, hepsin,corin, matriptase 1…)
N
TM
SEA
CUB
CUB
L
L
L
SERINE PROTEASE
C
Y141C
L166fs
I212T
R774C
A605fs
G603R
K636fs
L674F
P686fs
S561X
Mask S570fs
R599X
G442R
E461fs
E486D
D521N C510S
E522K
Y335X
Y393X
Q229fs
W247fs
K253EQ
S304L R271Q
A118D
Mutations associated with IRIDA
(Silvestri et al Blood 2009
De Falco et al, Hum Mut 2010)
Hepcidin activation in IRIDA: molecular
mechanism
IRIDA
IDA
m-HJV
BMP
TMPRSS6
m-HJV
BMP
BMPR
SMADs
HEPC
 serum iron
TMPRSS6
BMPR
SMADs
HEPC
 serum iron
(Silvestri et al, Cell Met 2008;8:502-11.)
Hepcidin: the key iron regulator
Liver
Fe
hepcidin
Fe
enterocytes
macrophages
IRIDA: hematological data
Mean±SD
Hb g/dl (at presentation) 7.7±1.3
Hb g/dl (at diagnosis)
9.21±1.8
MCV fl
55.47±7.6
Transferrin saturation
5.03±2.3
%
Ferritin ng/ml
126±82
Serum hepcidin nM
257±157*
Urin. hepcidin ng/mg
creat
4113±3089
*
(Camaschella and Poggiali Curr Op Ped, 2010)
How to diagnose IRIDA
 Evidence of microcytic anemia since the first months of life
 Moderate degree of anemia, more severe in children
(increased requests)
 Familial cases (autosomal recessive)
 Discrepancy between ferritin and Tf saturation levels
 Exclude celiac disease and other absorption disorders
 (Normal/high serum hepcidin in the presence of IDA and
normal CRP)
 Refractory to oral (control dose, type of iron and compliance)
and partially refractory to iv iron
 DNA sequence of TMPRSS6 gene (common SNP excluded)
How to treat IRIDA
 Oral iron ineffective (at least two cycles)
 I.V. iron: partial or slow response
 Epo: a single case reported with positive results
(Ramsey et al, Hum Mol Genet 2009). Iron must be added
 A recent report suggest some effect of ascorbic acid:
Cau M, Galanello R, Giagu N, Melis MA. Responsiveness
to oral iron and ascorbic acid in a patient with IRIDA.
(BCMD 2011)
2. Defects of TfRC cycle
1. Defects of transferrin (the ligand)
1. Defects of TfRC are not described!
2. Defects of TfRC components: DMT1, STEAP3
(Camaschella C, Br J Haematol, in press)
Atransferrinemia (OMIM #209300)
Autosomal recessive, extremely rare
Plasma transferrin nearly absent
Severe microcytic anemia and liver iron
overload
Low urinary hepcidin levels
Responds to plasma infusions
Hpx mice
Similar phenotype
Splicing mutations of transferrin
Hepcidin low/undetectable
Hypotransferrinemia: lesson from patients
hepcidin
transferrin
100% Tf saturation
Iron-deficient
erythropoiesis
NTBI
Liver, pancreas
iron overload
Microcytic
anemia
Transferrin (and TFR cycle) are indispensable for erythropoiesis
but not for liver iron uptake (NTBI)
Hepcidin suppression by the iron-deficient erythropoiesis increases
iron absorption
DMT1 deficiency (OMIM #206100)
mk mouse and Belgrade rat
severe iron-deficient anemia due to G185R homozygous
Dmt1 mutation
Dmt1 -/- mice even more severe
Patients with homozygous or compound
heterozygous DMT1 mutations
Microcytic hypochromic anemia and liver
iron overload (less severe than
atransferrinemia)
(Iolascon et al, J Pediatr. 2008;152:136-9)
Lesson from DMT1 human mutants
DMT1 is essential in erythropoiesis
DMT1 is not essential for liver iron uptake
DMT1 is not essential for duodenal iron
absorption (alternative pathways?heme absorption?)
Increased iron absorption occurs because of
low hepcidin levels
Partial response of anemia to erythropoietin treatment
A novel type of hypochromic anemia associated
with a nonsense mutation in the STEAP3 gene
(Grandchamps et al, Blood 2011)
Differential diagnosis of iron-related
inherited anemias
Atransferrine
mia
DMT1
mutations
IRIDA
IDA
Hb
low
low
low
low
MCV
low
low
low
low
Fe
low
high
low
low
low
high
high
Tf
Low/absent
Tf sat
high
high
low
low
ferritin
high
high
normal/high
low
hepcidin
low
low
high
low
3. Defects of iron utilization:
sideroblastic anemias
Perl’s staining
Anti-MT-ferritin
(Courtesy of R. Invernizzi, Pavia)
Mitochondrial iron metabolism
Heme
(modified from Blood 105;1867-1874, 2005)
Defects of heme synthesis
X-linked sideroblastic anemia (OMIM #300751)
The commonest form
Deficiency of ALAS2 reduced heme synthesis
Affects males (rarely females) - Variable severity
Piridoxin (Vitamin B6)-responsive (some cases)
Autosomal recessive sideroblastic anemia (OMIM 301310)
Phenotype more severe than XLSA
Mutations in SLC25A38, an erythroid mitochondrial aminoacid
transporter: involved in mitochondria glycine transport (?)
Piridoxin unresponsive
(Guernsey et al, Nat Genet. 2009;41:651-3)
Defects of Fe/S clusters biogenesis
X-Linked SA with Ataxia (OMIM 301310)
A syndrome described in 1985. Few families worldwide
Mild sideroblastic anemia - Late onset of ataxia
missense mutations of ABCB7, a transporter involved in Fe/S
export from mitochondria
GLRX5 deficiency
The human counterpart of zebrafish shiraz shows sideroblastic
anemia and iron overload due to an homozygous splicing
mutation of GLRX5 (a gene of Fe/S cluster)
(Camaschella et al Blood 2007)
GLRX5-mutant patient follow up
12
2500
2000
1500
6
1000
4
haemoglobin
ferritin
500
2
20
07
La
te
20
07
20
06
Ea
rly
de
c
Ea
rly
20
04
La
te
20
04
Ea
rly
20
05
La
te
20
05
m
ay
20
03
jan
20
03
20
03
20
02
0
20
01
0
20
00
ferritin ng/ml
8
Ferritin ng/mL
Hb g/dL
Haemoglobin g/dl
10
Ye a rs
Correlation Hb/ferritin: r = -0.79
start transfusions; stop transfusions: start DFO:
stop DFO
4. Defects of iron recycling:
aceruloplasminemia
AR (OMIM #604290) - Mutations of Ceruloplasmin (Cp)
Iron overload in liver, RE cells, pancreas, basal ganglia.
Clinical triad in midlle age:
1. Diabetes
2. Neurological disease (ataxia,dementia),
3. Retinal degeneration
(Miyaijma H. in: Pagonet al eds
GeneReviews University of Washington, Seattle)
ACERULOPLASMINEMIA: pathogenesis
Acp -/- mouse
(
(
 ferroxidase activity --  cellular iron efflux !
FPN
Fe2+
liver iron overload
CP
Fe3+
Low serum Fe
Increased Fe
absorption
Mild “iron deficiency”
anemia
Aceruloplasminemia: diagnosis
Microcytic/normocytic anemia
High serum ferritin, low transferrin saturation
Low serum copper (< 10mg/dL; nv 70-125mg/dL)
(Low ferroxidase plasma activity)
Undosable Ceruloplasmin
(Cp gene mutations)
MRI of liver, pancreas and basal ganglia
(striatum thalamus and dentate nucleus)
How to recognize an atypical microcytosis
1. Refractory (or partially refractory) microcytic anemia
DMT1 deficiency: no response to i.v. iron
2. Iron parameters not congruous:
high transferrin saturation and high serum ferritin
high serum ferritin and low transferrin saturation
3. Ringed sideroblasts (any percentage)
4. Familial cases
5. (High hepcidin) TMPRSS6 mutations
(Camaschella C Br J Haematol, in press)
Inherited iron-metabolism related anemias
Disorder
Gene
Defect of iron absorption
IRIDA
TMPRSS6
Defects of iron transport/erythroid uptake
Hypotransferrinemia
TF
DMT1 mutations
DMT1
STEAP3 mutations
STEAP3
Defects of cellular iron utilization
Sideroblastic anemia
X-linked sid. anemia
ALAS2
X-linked sid. anemia/ataxia
ABCB7
AR sideroblastic anemia
SLC25A38
GLRX5
Defects of iron recycling
Aceruloplasminemia
CP
OMIM n
#206200
#209300
#206100
+301300
#30131
#205950
#604290
Camaschella C, Br J Haematol, 2012 online
E-RARE project on microcytic anemias
(ERARE-115, HMA-IRON)
Carole Beaumont (France)
Clara Camaschella (Italy)
Martina Muckenthaler (Germany)
Mayka Sanchez (Spain)
Acknowledgements
Vita-Salute University &
San Raffaele Scientific Institute
Antonella Nai, Alessia Pagani
Laura Silvestri
Alessandro Campanella
Marco Rausa
University of Naples
Achille Iolascon Luigia De Falco
University of Verona
Domenico Girelli Natascia Campostrini
Fifth Meeting of the International BioIron Society
BioIron 2013: April 14 – 18, 2013
University College London UK
www.bioiron.org