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Next-gen sequencing settling in, making its mark

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William Check, PhD

November 2015—Resource heavy, reimbursement challenged. Next-generation sequencing has its difficulties, but its value to patient care is without question. For many laboratories today, it’s a test sent out, but as for so many other tests, it won’t always be.

Dr. Pfeifer

Dr. Pfeifer

“As techniques evolve, platforms become more user-friendly, and solid bioinformatic pipelines become available, it will not be too long before labs at large hospitals or large health systems are faced with the question of whether the economics are at the point to bring testing in-house,” says John Pfeifer, MD, PhD, vice chair for clinical affairs in pathology and immunology and professor of pathology and immunology and of obstetrics and gynecology at Washington University School of Medicine.
Until then, a handful of early adopters share their experiences and a few tips.

Dr. Fernandes

Dr. Fernandes

At Weill Cornell College of Medicine, the laboratory of Helen Fernandes, PhD, moved from single-gene tests to NGS panels for oncology specimens in September 2014. (One single-gene test, for EGFR, was retained for its rapid turnaround time.) In addition to helping guide therapy, she and colleagues are detecting mutations in specimens that help enroll patients into clinical trials. They are using a 50-gene panel for routine clinical care, one that took four months to validate and four to five months to earn final approval from the New York State Department of Health.

They use the AmpliSeq panel on the Ion Torrent for solid tumors. For hematologic neoplasms, the laboratory is validating a hematology panel on the Illumina MiSeq, which, Dr. Fernandes says, “is an easier platform to use and less labor-intensive.”

She describes two cases in which the technology had a positive impact. First, in a patient with cholangiocarcinoma, NGS identified a specific variant in the IDH1 gene that was a therapeutic target in ongoing clinical trials. In a second case, NGS findings facilitated the classification of thyroid carcinoma. “The pathologist sent two tumor nodules for molecular analysis. NGS detected a BRAF mutation in one thyroid nodule, which classified it as papillary, and an NRAS mutation in the other nodule, which qualified it as a follicular variant.” NGS showed that the two nodules harbored different driver mutations and supported the histopathological findings.

The technology lends itself to several observations, Dr. Fernandes says. “Over the last few months we have identified about 20 specimens from patients where we tested two different lung adenocarcinoma biopsies from the same lobe or different lobe. Interestingly, tumors in different lobes of the same lung or contralateral lungs had distinct genomic profiles. That complements the histopathological findings and has the potential to identify independent primaries.”

Dr. Fernandes’ laboratory has performed a second type of comparison with NGS: between an FNA (cytology) specimen and the corresponding surgical resection from the same patient. Of the 17 or 18 lung cancer cases they have studied so far, the same genomic variant was identified in both specimen types in 90 percent of the cases. “Cytologic specimens [cell blocks] are adequate for NGS and give you the same information as the surgical resection specimen 90 percent of the time,” she says.

In collaboration with Mark Rubin, MD, director of the Institute for Precision Medicine and the team at Cornell, Dr. Fernandes has assisted in the validation of a whole exome sequencing test for cancer that captures 80 to 90 percent of the exome. They submitted it eight months ago and are awaiting approval from the New York State Department of Health. “It is run on the Illumina HiSeq because it needs deeper coverage and higher throughput,” she says.

The molecular pathology laboratory at Weill Cornell is currently validating a commercial NGS test that looks at DNA and RNA from more than 140 genes simultaneously. To illustrate the utility of this type of panel, Dr. Fernandes points to lung cancer’s two types of important mutations—DNA variants in EGFR and gene fusions in ALK, ROS, and RET genes. “Right now, when we get a sample from a patient with lung adenocarcinoma, we interrogate the DNA variants on our 50-gene panel and send tissue to the cytogenetic laboratory for testing of gene fusions. Using a single NGS assay that can detect and identify DNA variants and RNA fusions simultaneously provides for a comprehensive testing approach.”

These advanced tests will be far more costly, Dr. Fernandes says, adding, “A major concern is getting reimbursed for NGS testing. We really need to get reimbursed for the 50-gene panel.” She expects it to happen soon.

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