Whole blood RNA offers a rapid, comprehensive approach to genetic diagnosis of cardiovascular diseases

Abstract

Purpose: Long QT Syndrome, Marfan Syndrome, hypertrophic and dilated cardiomyopathy are caused by mutations in large, multi-exon genes that are principally expressed in cardiovascular tissues. Genetic testing for these disorders is labor-intensive and expensive. We sought to develop a more rapid, comprehensive, and cost-effective approach.Methods: Paired whole blood samples were collected into tubes with or without an RNA-preserving solution, and harvested for whole blood RNA or leukocyte DNA, respectively. Large overlapping cDNA fragments from KCNQ1 and KCNH2 (Long QT Syndrome), MYBPC3 (hypertrophic and dilated cardiomyopathy), or FBN1 (Marfan Syndrome) were amplified from RNA and directly sequenced. Variants were confirmed in leukocyte DNA. Results: All 4 transcripts were amplified and sequenced from whole blood mRNA. Six known and 2 novel mutations were first identified from RNA of 10 probands, and later confirmed in genomic DNA, at considerable savings in time and cost. In one patient with MFS, RNA sequencing directly identified a splicing mutation. Results from RNA and DNA were concordant for single nucleotide polymorphisms at the same loci. Conclusion: Taking advantage of new whole blood RNA stabilization methods, we have designed a cost-effective, comprehensive method for mutation detection that should significantly facilitate clinical genetic testing in four lethal cardiovascular disorders. Genet Med 2007:9(1):23-33.

An FBN1 pseudoexon mutation in a patient with Marfan syndrome: confirmation of cryptic mutations leading to disease

“…Identification of a novel pseudoexon mutation in FBN1 confirms a role of cryptic mutations leading to the condition. In addition, these results suggest that analysis of the FBN1 message should be considered when the standard genetic diagnosis fails to identify a mutation (Miller et al 2007). Only a few cases of pseudoexon mutations have been reported to cause human genetic disorders.…”

Quantitative PCR as an Alternative in the Diagnosis of Long-QT Syndrome

“…In 2007, Miller et al [31] decided to establish peripheral whole blood to be a suitable material for isolation of mRNA for genetic testing use in the diagnosis of certain diseases of the cardiovascular system, including LQTS. Their innovative results showed that the expression profile of genes, of which mutations are responsible for the occurrence of LQTS, can be successfully examined with the use of mRNA isolated from peripheral blood.…”

“…50 mL of blood), skin biopsy, or fibroblasts culture. In addition, there was another limitation to the use of mRNA in the diagnosis of LQTS, namely, the fact that the expression of genes related to the occurrence of this disease is strictly limited to the myocardial cells and that mRNA transcripts could not be successfully extracted from lymphocytes or fibroblasts culture [31, 32]. …”

5 HT3-receptor antagonists and cardiac repolarization time in patients expressing a novel genetic target associated with baseline QTc interval abnormalities

“…Although a number of genetic targets associated with prolonged QTc intervals have been identified and studied, the utility of this information in perioperative risk reduction is limited [20][21][22][23][24]. Recently, Arking et al reported that the gene, NOS1AP (CAPON), coding for a regulator of neuronal nitric oxide synthase, constitutes a new genomic target that modulates cardiac repolarization [25].…”