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SNP Test Aims to ID Patients Who May Spontaneously Clear HCV


NEW YORK (GenomeWeb) – Hepatitis C viral infection can lead to life-threatening liver complications and can be quite costly to treat. Scientists recently discovered that particular genotypes are associated with spontaneous clearance of the virus, which could enable better allocation of resources in some settings.

To aid in this cause, researchers at the University of New South Wales in Australia have now created an assay for these genotypes incorporating PCR and high-resolution melt analysis that they hope will allow for better clinical management of HCV, particularly in low-resource environments.

Published in the Journal of Molecular Diagnostics, the assay detects SNPs in interferon lambda 3 (IFNL3) and interferon lambda 4 (IFNL4) and was optimized on a number of sample types including plasma, serum, whole blood, buffy coat, and dried blood spots.

A series of GWAS investigations about six years ago originally linked chronic HCV, treatment failure, and viral clearance with SNPs in the IL28B locus encoding the antiviral cytokine interferon lambda, Francois Lamoury, co-author on the study and scientist at the Viral Hepatitis Clinical Research Program at UNSW, told GenomeWeb in an email.

These included a flurry of studies published in Nature, Nature Genetics, and Gastroenterology, Lamoury said. A further Nature paper focused on the relationship between IFNL and spontaneous clearance of HCV, and a study in Hepatology in 2014 assessed this phenomenon with a large global cohort.

The relationship was also supported by work published recently in Nature Genetics distinguishing the IFNL4 gene, highlighting that "variation across the IFN-lambda region is a key determinant of the host response to HCV," according to an accompanying review

Other groups have deployed TaqMan assays and GWAS methods to detect this variation, Lamoury said, adding that his group previously used Sanger sequencing.

The HRM analysis uses newer fluorescent DNA dyes, such as Syto9, which saturate double strand DNA, he said. After PCR amplification, a melting curve analysis is performed with gene scanning to detect any change in the profile of the melting curve. 

The test is now being used on patients. "After developing our HRM assay, we completed significant validation to get an accreditation from the Australian National Association of Testing Authorities to perform this assay for clinical management at St. Vincent's Hospital in Sydney, Australia," Lamoury said.

Lamoury's group uses the Roche LightCycler 480 for the assay but other platforms would work as well. "Many recent quantitative PCR instruments allow HRM analysis, although sometimes a software upgrade might be needed," he noted. 

The assay could also potentially be used for HCV companion diagnosis, although the patient usually bears the cost of testing at this time.

"This is one reason why we developed an inexpensive alternative," Lamoury said. And while the IFNL genotyping may be less relevant in the context of highly effective directly acting antivirals (DAAs), "interferon-containing therapy will continue to be standard of care in resource limited settings and therefore [the test] might continue to inform HCV care," he said.

He further added that a recent study in Nature Communications suggested IFNL genotype may also be an etiology-independent predictor of liver fibrosis and therefore the HRM assay may also "be relevant to help manage those with untreated chronic HCV and other chronic liver diseases."

The group considered patenting the assay, but decided instead to "publish and provide the method as an affordable tool for other researchers and resource-limited settings." 

There are currently seven in vitro molecular diagnostics cleared by the US Food and Drug Administration to detect HCV infections, and firms such as Cepheid, Beckman Coulter, GenMark Diagnostics, and Janssen Diagnostics have tests in various stages of development.

But there are no approved companion diagnostics to guide HCV treatment once a patient is diagnosed, although GenomeWeb reported that one of the 2009 Nature studies was funded by Schering-Plough, a maker of an interferon-based HCV drug that owns the IP around one of the SNPs and was since acquired by Merck. A researcher who worked on that study with Schering-Plough described it to GenomeWeb as a "watershed moment" in pharmacogenomics.

Lamoury's study also suggested the SNP test could potentially "identify those likely to clear naturally and in whom treatment could be delayed, or help prioritize DAA treatment to those less likely to respond to interferon-containing regimens." Phrased thusly, the SNP test determines which patients may not need to buy the expensive drug treatments, but the assay could also theoretically be used to select patients most likely to respond to drug treatment, perhaps providing a lift to clinical trials.