Verigene^® CFTR and CFTR PolyT Nucleic Acid Tests (IVD)

Background

The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene, located on the long arm of human chromosome 7 (7q31), transports chloride ions (Cl-) out of certain types of epithelial cells in the mucosal lining of the lungs, liver, pancreas, digestive tract, reproductive tract, and skin. The movement of Cl- out of these epithelial cells by the CFTR protein creates an electric gradient that in turn moves positively charged sodium ions (Na+) in the same direction. This movement reduces the water potential outside the cells drawing along water by osmosis. The movement of water is important for maintaining a fluid layer of mucous on the surface of these epithedial cells. Certain mutations in the CFTR gene change the structure of the protein, inhibiting its ability to transport Cl-.¹

The most common clinical presentation of mutations in the CFTR gene is the disease Cystic Fibrosis (CF). Cystic Fibrosis is a hereditary disorder characterized by lung congestion and infection and malabsorption of nutrients by the pancreas. The most severe symptom of CF is viscous mucous in the lungs, which is dire ctly linked to pulmonary disease. Other symptoms include chronic sinusitis, nasal polyps, liver disease, pancreatitis, and in some instances, male infertility via congenital bilateral absence of the vas deferens. The overall average life expectancy of CF patients is approximately 30 years. There are ~ 1,000 new CF cases diagnosed each year in the US and about 30,000 children and adults living with CF in the United States.²

Over 1,300 mutations have been identified in the CFTR gene. Of the known mutations, the vast majority occur at frequencies of <0.1% or represent private mutations. The carrier frequency for CFTR mutations varies across populations, but is highest in Ashkenazi Jewish and Non-Hispanic Caucasians (Table 1).³

In 2001 and again in 2004, a working group for CF screening from the American College of Medical Genetics (ACMG) issued a policy for CFTR genetic testing.^4,5 The working group recommended a panel of 23 mutations to be tested as part of a routine population carrier screening. The revised panel from the 2004 ACMG Policy includes:

• 23 core mutations
• Three exon 10 variants and
• Intron 8 polymorphisms (5T/7T/9T) but only as a reflex when R117H is detected during carrier screening. (Figure 1)

Nanosphere offers two Test Cartridges for the detection of mutations in the CFTR gene: a core panel Test Cartridge and a reflex PolyT Test Cartridge (see Table 2).

Intended Use

The Verigene^® CFTR and Verigene^® CFTR PolyT Nucleic Acid Tests are qualitative in vitro diagnostic devices used to genotype a panel of mutations and variants in the cystic fibrosis transmembrane conductance regulator (CFTR) gene in genomic DNA isolated from human peripheral whole blood specimens.The panel includes mutations and variants recommended by the 2004 American College of Medical Genetics (ACMG) and the 2005 American College of Obstetricians and Gynecologists (ACOG). The Verigene^® CFTR Nucleic Acid Test provides information intended to be used for carrier testing in adults of reproductive age and in confirmatory diagnostic testing of newborns and children.

These tests are not indicated for use in fetal diagnostic or pre-implantation testing and not indicated for standalone diagnostic purposes and the results should be used in conjunction with other available laboratory and clinicalinformation. Both tests are intended to be used on the Verigene^® System.

Clinical Study Highlights

Percent agreement of the Verigene^® CFTR Nucleic Acid Test as compared to bi-directional DNA sequencing was assessed in a methods comparison study using 234 samples distributed across three clinical test sites. The samples were comprised of retrospective and prospective samples of mixed genotype that included all mutations and variants evaluated by the test. In a methods comparison study, percent agreement for all genotypes was 100%. Percent agreement of the CFTR PolyT Nucleic Acid Test as compared to bidirectional DNA sequencing was assessed using DNA extracted from 40 prospective whole blood samples distributed across three clinical test sites. Percent agreement for all genotypes was 95%.

Reproducibility of the Verigene^® CFTR and CFTR PolyT Nucleic Acid Tests was assessed at each of the three clinical test sites using a panel of 24 reproducibility test samples. Reflex PolyT testing was conducted where appropriate. At each site, two operators each ran the panel of 24 reproducibility samples once per day for 5 non-consecutive days for a total of 240 samples per site. Samples consisted of DNA extracted from leukocyte-depleted whole blood spiked with cell lines with specific CFTR mutations. The test call rate for both the CFTR and CFTR PolyT tests across the three clinical test sites was 94% with 100% agreement to sample genotype as determined by bi-directional DNA sequencing.

Literature Cited

1. Rowe SM, Miller S, Sorcher ET. NEJM 2005; 352): 1992-2001
2. Cystic Fibrosis Foundation Website 2007: http://nanosphere.orbisdesign.net/www.cff.org
3. Amos T, Foldman GL, Monaghan K. et al. American College of Medical Genetics Technical Standards and Guidelines for CFTR Mutation Testing, 2006 Edition; http://nanosphere.orbisdesign.net/www.acmg.net
4. Grody WW, Cutting GR, Klinger KW, et al. Genet Med 2001; 3): 149-154
5. Watson MS, Cutting GR, Desnick RT, et al. Genet Med 2004; 6): 387-39

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