Currently, doctors consider patients to be in complete remission if a pathologist is unable to find any malignant cells in a small sample of the bone marrow. However, Dr. Bejar explained that, by using NGS, pathologists can sequence the blood of the bone marrow to look for mutations associated with residual disease. This would detect evidence of “measurable residual disease†that might be missed in the bone marrow samples.
Ensuring Reliability and Accuracy
Before NGS becomes a routine part of patient care, the results it generates need to be trustworthy and accurate. There are hundreds of laboratories and companies that conduct NGS, but, as Julie Eggington, MS, PhD, CEO of the Center of Genomic Interpretation, told ASH Clinical News, not all are created equal.
While Dr. Eggington said that any lab performing NGS must be Clinical Laboratory Improvement Amendments (CLIA)–accredited, “a CLIA accreditation is basically pointless for NGS tests.†The U.S. government legislated CLIA accreditation in 1988, long before NGS was even imagined.
CLIA accreditation establishes quality standards for laboratory testing performed on specimens from humans, such as blood, body fluid, and tissue. However, she noted, in the context of labs performing NGS, the certification mainly demonstrates that a lab isn’t using expired chemicals for tests, and not that interpretation of results is correct.
In April 2019, the Clinical Laboratory Improvement Advisory Committee, a group managed by the Centers for Disease Control and Prevention, convened a workgroup on NGS to consider recommendations for assuring the quality of NGS-based testing in clinical laboratory settings.3 The group outlined challenges in performing NGS test validations and quality control and assurance, but has not issued a set of NGS-specific standards for accreditation.
The College of American Pathologists (CAP) and the Association for Molecular Pathology (AMP) published a set of five worksheets that “translate regulatory specifications into concrete instructions that guide the user through the entire life cycle of an NGS test.â€4 The guidance focuses on variant detection in the setting of inherited disease and help labs document NGS protocols, but Dr. Eggington said even these aren’t able to keep pace with the progress being made in NGS, especially in the context of malignancy.
“While choosing a CAP-accredited lab is better than choosing a lab that only has CLIA accreditation, it still doesn’t mean that the test is worthwhile,†she explained. “I can generate protocols and put them in binders and pass inspection to ensure that I can reproduce the results, but it doesn’t mean that my results or interpretations are accurate.â€
Laboratories might try to overcorrect for lack of quality by putting as much information in the NGS test report as they can. “Some doctors really like that because just about every patient case they send in will come back with a recommendation,†said Dr. Eggington. “The problem is that the majority of the recommendations will be useless, containing false positives and dead leads. It is a waste of time for the patients and a waste of money for the health care system.â€
Even if two separate laboratories both seem to be high-quality, they may report conflicting results. Dr. Pfeifer provided this example: Several years ago, his team sequenced a patient whose care team had requested a second opinion. Even though he considered the first lab that conducted the NGS to be a high-quality lab, his lab still found a mutation that the first one didn’t.
“The previous lab was running a very narrow hotspot-based test,†he explained. “Although we had a hotspot-based assay, our target area was a little bigger. So, the variant we found was in a part of the DNA that we tested and they didn’t.â€
The two tests were virtually identical from a technical and an analytic point of view, Dr. Pfeifer continued, “it’s just that, because the tests were designed differently, they produced two different sets of information.â€
Quantifying Quality
Laboratories may test for anywhere from one or two gene mutations in certain parts of the genome to hundreds of genes, depending on the clinical question that needs to be answered. “We require our clinical colleagues to tell us what they want us to do,†said Dr. Pfeifer. “At last count, there are about 8 billion base pairs in the human genome. That’s an awful lot. Somebody has to tell the sequencing laboratory what to look for.â€