Global sequencing approach for characterizing the molecular background of hereditary iron disorders.
BACKGROUND
New genetic forms of hereditary hemochromatosis (HH) or hereditary hyperferritinemia (HF) have been identified over the last few years, and abnormalities of various genes may interact in a single patient. This study aimed to develop a rapid automated method for sequencing the main genes involved.


METHODS
We used a standard 96-well microplate with a single PCR condition in an adaptation of the SCAIP (single-condition amplification with internal primer) method to sequence the HFE (hemochromatosis), HAMP (hepcidin antimicrobial peptide), HFE2/HJV [hemochromatosis type 2 (juvenile)], SLC40A1 (ferroportin), and TFR2 (transferrin receptor 2) genes, and the 5' untranslated region of the FTL (ferritin, light polypeptide) gene. To further simplify the method, we adjusted PCR conditions to avoid the use of an internal primer and applied this single-condition amplification method to 38 selected, unrelated patients. We tailored the genetic investigation according to the clinical picture, with the patients falling into 2 groups. Group 1 consisted of patients with hyperferritinemia and high transferrin saturation (TS) (classic adult and juvenile HH forms, groups 1A and 1B, respectively), and group 2 consisted of patients with hyperferritinemia and low, typical, or slightly increased TS, with or without iron overload (groups 2A and 2B, respectively).


RESULTS
With this strategy we identified single-gene and multigene abnormalities, including 6 previously undescribed abnormalities in HFE (c.794dupA), HFE2 (c.-89-4dupT), and SLC40A1 (c.262A>G, c.533G>A, c.1468G>A, and c.-59_-45del).


CONCLUSIONS
This method is a simple approach for investigating hereditary iron overload or HF and allows rapid evaluation of patients.
Cunat
Séverine
S
Laboratory of Haematology, CHU of Montpellier, Montpellier, France.
Giansily-Blaizot
Muriel
M
Bismuth
Michael
M
Blanc
François
F
Dereure
Olivier
O
Larrey
Dominique
D
Quellec
Alain Le
AL
Pouderoux
Philippe
P
Rose
Christian
C
Raingeard
Isabelle
I
Renard
Eric
E
Schved
Jean-François
JF
Aguilar-Martinez
Patricia
P
D000328:Adult; D000368:Aged; D002648:Child; D005260:Female; D005293:Ferritins / Q000097:blood; D006432:Hemochromatosis / Q000235:genetics; D006801:Humans; D019189:Iron Metabolism Disorders / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques; D009154:Mutation; D021141:Nucleic Acid Amplification Techniques; D017422:Sequence Analysis, DNA / Q000379:methods*
D005293:Ferritins