Reliable low-density DNA array based on allele-specific probes for detection of 118 mutations causing familial hypercholesterolemia.
BACKGROUND
Patients with familial hypercholesterolemia (FH) have a high risk of premature cardiovascular disease (PCVD). Mutations in the LDL receptor (LDLR) gene and the R3500Q mutation in the apolipoprotein B (APOB) gene are known to cause FH, but lack of high-throughput methods makes routine genetic diagnosis difficult. The objective of this work was to develop a DNA array for large-scale identification of mutant LDLR alleles.


METHODS
We developed a low-density oligonucleotide microarray to identify 118 DNA sequence variations (117 for the LDLR gene and 1 for the APOB gene). We verified specificity and sensitivity by analyzing 1180 previously sequenced DNA samples, and conducted a blind study screening 407 Spanish patients with a clinical diagnosis of FH.


RESULTS
The DNA array confirmed the previous genotyping results in almost all cases. In the blind study, the microarray detected at least 1 mutation in 51% of the patients for whom clinical diagnosis was classified as certain according to Dutch FH-MEDPED criteria; it also identified mutations in 37% of those with a diagnosis of probable/possible FH, thus giving a definite diagnosis. Patients harboring null mutations had shorter PCVD-free survival times and higher relative risk of PCVD than patients with a missense mutation.


CONCLUSIONS
The proposed DNA array allows large-scale population screening and provides molecular information regarding mutation type and its correlation with clinical severity of FH, which can be used to develop therapeutic strategies.
Tejedor
Diego
D
Departamento de Bioquímica y Biología Molecular y Celular, Universidad de Zaragoza, Zaragoza, Spain. dtejedor@progenika.com
Castillo
Sergio
S
Mozas
Pilar
P
Jiménez
Elisa
E
López
Mónica
M
Tejedor
M Teresa
MT
Artieda
Marta
M
Alonso
Rodrigo
R
Mata
Pedro
P
Simón
Laureano
L
Martínez
Antonio
A
Pocoví
Miguel
M
D000483:Alleles; D005260:Female; D005820:Genetic Testing; D005838:Genotype; D006801:Humans; D006938:Hyperlipoproteinemia Type II / Q000175:diagnosis / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D015345:Oligonucleotide Probes; D011973:Receptors, LDL / Q000235:genetics*; D012680:Sensitivity and Specificity
D015345:Oligonucleotide Probes; D011973:Receptors, LDL