PGx testing: recommended alleles for CYP2C19 panels

Elizabeth Silverman

August 2018—After more than a year of gathering information and deliberating, members of the Association for Molecular Pathology Pharmacogenomics Working Group have issued the first in what will be a series of recommendations to standardize pharmacogenetic testing.

The group’s first set of recommendations identify a minimum set of alleles for inclusion in clinical CYP2C19 genotyping panels. Published in the May issue of the Journal of Molecular Diagnostics (Pratt VM, et al. 2018;20[3]:269–276), the recommendations were designed to complement and fill gaps in existing pharmacogenomics guidelines.

The working group, a subgroup of the AMP’s Clinical Practice Committee, came together after the 10th Genome Medicine conference of the National Human Genome Research Institute, in 2017, called for assay standardization, says group member Karen E. Weck, MD, a past chair of the CAP/American College of Medical Genetics and Genomics Biochemical and Molecular Genetics Resource Committee. An additional impetus was a survey of pharmacogenetic panels used to characterize cell lines for reference materials, conducted by the Centers for Disease Control and Prevention’s Genetic Testing Reference Material Program. It found that the variants tested by laboratories were not consistent and that no two tests that examined any of the 28 pharmacogenetic genes in the study were designed to detect the exact same set of variants. “It seemed as if there was a need for better standardization in this space,” Dr. Weck said in an AMP webinar in June, noting that Victoria Pratt, PhD, of the Department of Medical and Molecular Genetics, Indiana University School of Medicine, led the effort. “We identified where there might be heterogeneity or lack of concordance and gaps in current practice,” Dr. Weck tells CAP TODAY. She is a professor of pathology and laboratory medicine and genetics and director of the molecular genetics laboratory, University of North Carolina School of Medicine.

CYP2C19 is one of the cytochrome P450 enzymes in the liver that is important for phase one metabolism of many drugs. Together, CYP2C19, CYP2C9, CYP2D6, CYP3A4, and CYP3A5 are responsible for the metabolism of more than 80 percent of drugs prescribed today, Dr. Weck said. The cytochrome genes are also highly polymorphic and contain many variants or single nucleotide polymorphisms that can affect enzymatic activity and alter drug metabolism. In particular, the substrates of CYP2C19 include clopidogrel, voriconazole, proton pump inhibitors, SSRIs, and some tricyclic antidepressants. In addition to the normal CYP2C19*1 allele, the CYP2C19 gene can have up to 35 different alleles (named in order of discovery), many of which are associated with altered enzyme function, resulting in poor, intermediate, rapid, or ultra-rapid drug metabolism. Failure to test for an allele that alters drug metabolism results in an incorrect classification to the default CYP2C19*1 normal function allele. “If you’re not including the SNP,” Dr. Weck said, “you’re not identifying the SNP.”

Because CYP2C19 affects the metabolism of so many drugs, it is one of the most frequently tested for genes. Much of the testing has also been driven by the FDA’s 2010 black-box warning for clopidogrel that two common variant CYP2C19 alleles, CYP2C19*2 and CYP2C19*3, result in poor metabolism of clopidogrel and increase the risk of adverse cardiovascular events. Also driving CYP2C19 testing is the publication of guidelines by the Clinical Pharmacogenetics Implementation Consortium and the Dutch Pharmacogenetics Working Group for CYP2C19 for multiple drugs. “I would say that the CPIC and DPWG dosing guidelines have been extremely important in helping to recommend pharmacogenetic action based on pharmacogenetic testing,” Dr. Weck said. In her experience, “this has also produced a big uptick in the number of places that are doing pharmacogenetic testing for CYP2C19 and for other genes as well.”

She noted two caveats about pharmacogenomics. First, allele function derived from in vitro models may not directly translate to a clinical phenotype and metabolizer status. “And much of the pharmacogenetic characterization of certain allele function is done by looking at enzymatic activity in in vitro models. So we took into consideration both what’s known about functional activity in these in vitro models and what’s been shown to be clinically relevant in terms of drug response.”

Second, allele function for pharmacogenetic alleles can be substrate and/or drug concentration dependent. “So there may be certain alleles that have been shown to be very important for some drugs or certain dosages of some drugs that haven’t borne out in other cases.” This further complicates analysis of the clinical importance of pharmacogenetic alleles, she said.

The working group members first defined the attributes that a recommended must-test allele should have. Their framework for evaluation consisted of the functional status of the CYP2C19 alleles, their frequency in a multiethnic population, and whether reference materials are available. They also examined what commercial genotyping platforms are available and which alleles are on those platforms.

With respect to the variants themselves, the group reviewed the literature, testing resources, reference materials, which tests are being offered now, and the availability of CAP proficiency testing and other quality assessment programs.

Their recommendations are divided into a tier-one minimum set of alleles and a tier-two set of alleles for extended panels. To be included in tier one, a variant allele needs to meet all three of the following criteria; for tier two, at least one criterion.