12th Congress of the European Hematology ... - Haematologica

12 th Congress of the European Hematology Association
D gene as responsible for PCT, showing a high molecular heterogeneity.
Few data are presently available on the Italian population. In a previous
work, we have identified 18 different molecular defects among 22
unrelated Italian PCT patients. Aims. The aim of this study was to identify
the molecular defect in the URO-D gene in fifteen new families
with typical clinical and biochemical signs of PCT. Methods. The promoters,
the entire coding region and the intron-exon boundaries of URO-D
gene has been amplified by polymerase chain reaction and submitted to
direct automated sequencing. Results. The plasma peak at 620 nm was
positive in all subjects. PCR analysis and automated direct sequencing
identified seven molecular defects in URO-D gene, three of which are
new finding. In this study we analyzed a total of 22 subjects, 20 of which
carry a mutation. Table 1 summarized the mutations identified. Summary.
These results allowed the identification of three novel URO-D mutations.
In a previous work, we have identified other thirteen new molecular
defects for a total of sixteen different mutations restricted to the Italian
population; the 243-245 del CAT and the 1107G>A are the most
common. So far our group identified a total of twenty one different
mutations among Italian patients. This study confirmed the high heterogeneity
of molecular abnormalities responsible for PCT phenotype and
the presence of clusters of mutations in particular geographic areas.
Table 1. Mutations identified in the italian population. The mutations identified
by our group are indicated with*.
0781
FERROPORTIN DISEASE OR TYPE IV HEMOCHROMATOSIS: SHOWING A
HETEROGENEOUS PHENOTYPICAL EXPRESSION
M.A. Molina, 1 J.A. García, 2 R. Pérez, 2 M.T. Gallego, 2 M.J. Giménez, 3
C. Avivar, 2 A. Remacha, 4 A. Altés, 5 M. Baiget4 1 Empresa Pública Hospital de Poniente, EL EJIDO. ALMERÍA; 2 Hospital de
Poniente, EL EJIDO, ALMERÍA; 3 Transfusion Center, ALMERÍA; 4 Hospital
de Sant Pau, BARCELONA; 5 Hospital de l'Esperit Sant, STA. COLOMA DE
GAMENET,. BARCELONA, Spain
Introduction. Hereditary hemochromatosis (HH) is the most prevalent
form of iron overload in caucasians. This term comprises a group of disorders
that lead to abnormalities in iron homeostasis, leading to
increased intestinal absorption of iron and tissue deposition. If not treated,
the accumulated iron can result in tissue damage and lesions such as
liver cirrhosis, diabetes mellitus, arthropathy, myocardiopathy,
endocrine disorders and hepatocellular hepatocarcinoma. A wide majority
of clinical cases of HH are associated to homozygosity from the
C282Y mutation of the HFE gene (type I hemochromatosis), although
some other mutations of this gene can also lead to iron overload. A
recently discovered molecule (in the year 2000), Ferroportin 1, FPN1, is
the only known exporter of cellular iron. In 2001, some families with
atypical autosomal dominant HH were reported, in whom missense
mutations in the FPN1 gene were found (SLC40A1). This disorder has
been named type IV hemochromatosis, or Ferroportin disease, and its
clinical presentation seems to be heterogeneous. Objectives. To describe
the phenotypic expression in members of a family suffering from type
292 | haematologica/the hematology journal | 2007; 92(s1)
IV HH. Among them, in 2005, a novel mutation resulting in a single
nucleotide substitution (c.263G>C) in exon 3 of the FPN1 gene was
found. Results. From 1998 up to the present, 6 members of this family
have been included in a weekly or fortnightly phlebotomy program.
There were four males and two females . No symptoms were present,
except arthralgia in the oldest man. They began phlebotomies at the
ages of 56,50,46,22 for the males and 24 and 19 for the females. A slightly
low platelet count was presented in four cases (115×10 3 /L; 141×10 3 /L;
113×10 3 /L and 140×10 3 /L). The glucose level was normal in all of them
and also the liver enzymes, except the ALT and GGT levels, which were
slightly higher in the oldest man. All of their ferritin levels were elevated:
9,075 ng/mL; 2,830 ng/mL; 5,291 ng/mL; 1,870 ng/mL in the males,
and 1,524 ng/mL and 724 ng/mL in the females. Their transferrin saturation
was 91%; 43.4%; 71.2%; 63.7%; 41% and 60% respectively. Liver
biopsies were done on the four males with Grade IV iron overload in
the oldest man and Grade III in the rest. Fibrosis was present in three
cases but none of them presented a cirrhosis state. Two of the male cases,
in the biweekly phlebotomy program, showed a tendency to anemia.
Several other members of this family, who have rejected periodical phlebotomy
therapy, show extremely high serum ferritin levels but no related
symptoms. Conclusions. 1. Ferroportin disease, or type IV hereditary
hemochromatosis, must be suspected in cases of familial asymptomatic
iron overload that present at early ages, even in women, with disproportionately
low transferrin saturations considering the greatly elevated ferritin
levels. 2. A standard biweekly phlebotomy program may cause a
tendency to anemia and therefore may not be well tolerated. 3. The
response to phlebotomies is heterogeneous, even between members of
the same family with the same molecular lesion. 4. Prospective population
studies are required to define the most appropriate therapy in this
type of hemochromatosis, in view of the apparently benign nature of
this particular iron overload.
0782
IRON REGULATING GENES AND CHELATION
M.-L. Hatzinicolaou, D. Palaiologou, G. Papanikolaou, A. Kattamis
University of Athens, ATHENS, Greece
Background. Iron-chelating therapy has been used in clinical practice
for over 20 years and has contributed in a significant decrease in mortality
and morbidity of patients with transfusion-dependent anemias
and chronic iron overload. Many aspects of the mechanisms of action
of chelating agents have yet to be elucidated. So far, no studies have
identified the intracellular iron pathways affected by both chelators and
little is known on their effects on gene expression and translational modulation.
Aims. The aims of the present study were to evaluate the effects
of iron chelation on the expression of iron-regulating genes. Methods. To
study the effects of iron chelation on the expression of iron-related
genes, normal and iron loaded human hepatic cells HepG2, were treated
with the iron chelating agents, desferrioxamine (DFO) and deferasirox
(Exjade, ICL670), which were provided in pure form by Novartis Pharma
AG, Switzerland. The cells were initially cultured with either ferric
ammonium citrate (FAC) 50 µm or normal medium. After 24 hours DFO
or deferasirox was added in the cultures for further 12-hour period. The
transcriptional expression of the iron-related genes, namely DMT1 (divalent
metal transporter 1), TfR1 (transferin receptor 1), HAMP (hepcidin)
and Ireg1 (ferroportin), was studied using quantitative Real-Time PCR
(qRT-PCR). Statistic analysis was performed using Student’s paired t-test.
Results. Expression of both iron importers DMT1 and TfR1 is induced
by both DFO and deferasirox, indicating effective depletion of intracellular
iron with chelation. Regulation is most probably mediated via the
IRE/IRP system and stabilization of mRNA transcripts. Ireg1 mRNA
expression was increased by DFO and greater by deferasirox mainly in
non-iron loaded cells. Only deferasirox resulted in mild up-regulation of
ferroportin’s transcription in iron-loaded cells. Hepcidin transcription
was up-regulated in low iron conditions following chelation. The induction
of HAMP mRNA expression was more pronounced in iron-loaded
hepatic cells, especially with deferasirox. Conclusions. We conclude that
DFO and deferasirox modulate the transcription of iron-regulating genes
in the human hepatic cells HepG2. The effects on gene expression
depend on the initial iron status of the cell. A more pronounced response
was observed with deferasirox compared to DFO, suggesting that
deferasirox could prove to be more effective in mobilizing intracellular
iron and in controlling iron overload. Further studies are required to fully
understand the underlying mechanisms of DFO and deferasirox mode
of action and to explore the possibility that chelation therapy affects iron
homeostasis via alternative pathways, like the modulation of gene translation
and/or transcription.