Multiplex ARMS analysis to detect 13 common mutations in familial hypercholesterolaemia

Abstract

DNA analysis and mutation identification is useful for the diagnosis of familial hypercholesterolaemia (FH), particularly in the young and in other situations where clinical diagnosis may be difficult, and enables unambiguous identification of at-risk relatives. Mutation screening of the whole of the three FH-causing genes is costly and time consuming. We have tested the specificity and sensitivity of a recently developed multiplex amplification refractory mutation system assay of 11 low-density lipoprotein receptor gene (LDLR) mutations, one APOB (p.R3527Q) and one PCSK9 (p.D374Y) mutation in 400 patients attending 10 UK lipid clinics. The kit detected a mutation in 54 (14%) patients, and a complete screen of the LDLR gene using single-stranded conformation polymorphism/denaturing high performance liquid chromatography identified 59 different mutations (11 novel) in an additional 87 patients, for an overall detection rate of 35%. The kit correctly identified 38% of all detected mutations by the full screen, with no false-positive or false-negative results. In the patients with a clinical diagnosis of definite FH, the overall detection rate was higher (54/110 = 49%), with the kit detecting 52% of the full-screen mutations. Results can be obtained within a week of sample receipt, and the high detection rate and good specificity make this a useful initial DNA diagnostic test for UK patients.

Extraction of Sequence Conservation Features for the Prioritization of Candidate Single Amino Acid Polymorphisms

“…All of these SAAPs belong to gene LDLR and rank top 140 among 13736 SAAPs (less than 1.00%). These results are just in accordance with some previously published research works [28][29][30], which point out that FH results from defective lowdensity lipoprotein receptor (LDLR) activity, mainly due to LDLR gene defects.…”

The use of next-generation sequencing in clinical diagnosis of familial hypercholesterolemia

“…10,11 During the past few years, high-throughput next-generation sequencing (NGS)-based methods have become available for DNA analysis. They have not only proven successful in new disease gene identification, but following the availability of economic "benchtop" sequencers, they have become more easily applicable to targeted diagnostic sequencing.…”

Elucigene FH20 and LIPOchip for the diagnosis of familial hypercholesterolaemia: a systematic review and economic evaluation.

“…The sensitivity of the kit was found to be 30% for patients with a clinical diagnosis of possible FH, 52% for patients with definite FH and 38% for those with a clinical diagnosis of definite or possible FH (Table 10). Mutation-level analysis The previous report 54 to the study by Taylor and colleagues 37 reported that there were no false-positive and no false-negative results from the Elucigene FH13 kit for detection of FH-causing mutations in patients.…”

“…Hooper and colleagues 36 reported exon-by-exon sequencing of the LDLR gene, whereas Yarram 38 reported sequencing of all 18 LDLR exons and the promoter region. 54 were included because all study participants received both Elucigene FH20 and also a reference standard of sequencing of the LDLR gene, unlike the Taylor and colleagues 2010 report, 37 in which only test-negatives on Elucigene FH20 went on to receive CGA.…”

“…A previous report 54 to Taylor and colleagues 37 provided information on an earlier version of Elucigene (FH13). In this report the FH13 kit was validated against a reference standard of sequencing the LDLR gene in a patient population in which all patients were clinically diagnosed with definite or possible FH based on the Simon Broome criteria and all received testing with the kit and sequencing of the LDLR gene.…”