Implementation of preemptive DNA sequence-based pharmacogenomics testing across a large academic medical center: The Mayo-Baylor RIGHT 10K Study.

PURPOSE: The Mayo-Baylor RIGHT 10K Study enabled preemptive, sequence-based pharmacogenomics (PGx)-driven drug prescribing practices in routine clinical care within a large cohort. We also generated the tools and resources necessary for clinical PGx implementation and identified challenges that need to be overcome. Furthermore, we measured the frequency of both common genetic variation for which clinical guidelines already exist and rare variation that could be detected by DNA sequencing, rather than genotyping. METHODS: Targeted oligonucleotide-capture sequencing of 77 pharmacogenes was performed using DNA from 10,077 consented Mayo Clinic Biobank volunteers. The resulting predicted drug response-related phenotypes for 13 genes, including CYP2D6 and HLA, affecting 21 drug-gene pairs, were deposited preemptively in the Mayo electronic health record. RESULTS: For the 13 pharmacogenes of interest, the genomes of 79% of participants carried clinically actionable variants in 3 or more genes, and DNA sequencing identified an average of 3.3 additional conservatively predicted deleterious variants that would not have been evident using genotyping. CONCLUSION: Implementation of preemptive rather than reactive and sequence-based rather than genotype-based PGx prescribing revealed nearly universal patient applicability and required integrated institution-wide resources to fully realize individualized drug therapy and to show more efficient use of health care resources.

Pharmacogenomics testing in patients with liver transplant and potential impact on prospective management.

Aim: Pharmacogenomics (PGx) tests are performed on whole-blood or saliva specimens. In patients with a transplanted liver, PGx results may be discordant with hepatic drug metabolizing enzyme activity. We evaluate the incidence and impact of PGx testing in liver transplant recipients, detail potential errors and describe clinical decision support (CDS) solution implemented. Materials & methods: A retrospective cohort study of liver transplant recipients at Mayo Clinic who underwent PGx testing between 1 January 1996 and 7 October 2019 were characterized. Impact of a CDS solution was evaluated. Results: There were 129 PGx tests in 117 patients. PGx testing incidence increased before (per year incidence rate ratio = 1.45, 95% CI: 1.20-1.74, p < 0.001) and after transplant (incidence rate ratio = 1.48, 95% CI: 1.27-1.72, p < 0.001). Three erroneous PGx tests were avoided 6 months following CDS implementation. Conclusion: Incidence of PGx testing in liver transplant recipients is increasing, leading to erroneous therapeutic decisions. CDS interventions and education are needed to prevent errors.

Increased risk of hospitalization for ultrarapid metabolizers of cytochrome P450 2D6.

BACKGROUND: Cytochrome P450 2D6 (CYP2D6) is responsible for the metabolism of clinically used drugs and other environmental exposures, but it is unclear whether the CYP2D6 phenotype is associated with adverse health outcomes. The aim was to determine the association of CYP2D6 phenotype with the risk of hospitalization or an emergency department (ED) visit among a group of primary care patients. METHODS: In this study, 929 adult patients underwent CYP2D6 testing. The primary outcome was risk of hospitalization or an ED visit from January 2005 through September 2014. CYP2D6 genotypes were interpreted as 1 of 7 clinical phenotypes, from ultrarapid to poor metabolizer, and patients with the extensive metabolizer phenotype were used as the reference group. The hazard ratios (HRs) and 95% confidence intervals (CIs) were estimated for finding the association of CYP2D6 phenotypes with the risk of hospitalization or an ED visit by using Cox proportional hazard models and adjusting for age and sex. RESULTS: The median age was 49 years (interquartile range, 46-52 years); 74% of patients had 3 or fewer chronic conditions, 285 had at least 1 hospitalization, and 496 had at least 1 ED visit. The risk of hospitalization was higher among patients who were ultrarapid metabolizers compared to extensive metabolizers (47% vs 30%; HR, 1.69; 95% CI, 1.11-2.57), as was the risk of an ED visit (62% vs 49%; HR, 1.50; 95% CI, 1.05-2.14). For poor metabolizers compared to extensive metabolizers, there was no difference in the risk of hospitalization (HR, 0.95; 95% CI, 0.58-1.56), but there was an increase in the risk of an ED visit (HR, 1.38; 95% CI, 0.96-1.98) (the difference was not statistically significant). CONCLUSION: We found an increased risk of hospitalization or an ED visit among ultrarapid compared to extensive CYP2D6 metabolizers. Further research identifying the mechanisms of the association and ultimate clinical utility is warranted.

Analysis of compound heterozygous CYP2C19 genotypes to determine cis and trans configurations.

BACKGROUND: Through allele specific PCR we studied 220 CYP2C19 compound heterozygous samples, of unknown ethnicity, to determine the haplotype for each of the variations within a sample. MATERIALS & METHODS: The genotypes assessed were: 180 *2 and *17 samples (100% in trans); 20 *2 and *11 samples (100% in cis); ten *4 and *17 samples (50% of the samples were *1/*4B and 50% *4A/*17); six *2, *11 and *17 samples (100% showed *2 and *11 in cis, and *17 in trans); two *2, *4 and *17 samples (100% *4B with *2 in trans); one sample with *17 and *34 (these were in trans); and one sample that contained *2, *17, c.463G>T (p.E155X; *17 and c.463G>T were in cis, with *2 in trans). RESULTS & CONCLUSION: In our study, we observed a different frequency for the *4B allele (when a sample contains both *4 and *17); and identified *17 occurring in cis with a novel nonsense allele. Accurately assessing a patient's genotype, including assignment of a haplotype, can be important when making a phenotype prediction.

Associations between serotonin transporter gene polymorphisms and heat pain perception in adults with chronic pain.

BACKGROUND: The triallelic serotonin transporter gene linked polymorphic region (5-HTTLPR) has been associated with alterations in thermal pain perception. The primary aim of this study was to investigate the associations between heat pain (HP) perception and the triallelic 5-HTTLPR in a large cohort of adults with chronic pain. METHODS: The cohort included 277 adults with chronic pain who met inclusion criteria, and were consecutively admitted to an outpatient pain rehabilitation program from March 2009 through March 2010. Individuals were genotyped for the triallelic 5-HTTLPR (including rs25531) and categorized as high, intermediate, or low expressors of the serotonin transporter. Standardized measures of HP perception were obtained using a validated quantitative sensory test method of levels. RESULTS: The distribution of the high, intermediate, and low expressing genotypes was 61 (22%), 149 (54%) and 67 (24%), respectively. The Hardy-Weinberg P-value was 0.204 which indicated no departure from equilibrium. A significant effect of genotype was observed for values of HP threshold (P = 0.029). Individual group comparisons showed that values of HP threshold were significantly greater in the intermediate compared to the high expressing group (P = 0.009) but not the low expressing group (P > 0.1). In a multiple variable linear regression model, the intermediate group (P = 0.034) and male sex (P = 0.021) were associated with significantly greater values of HP 0.5, but no significant genotype-by-sex interaction effect was observed. CONCLUSIONS: In this study that involved adults with chronic pain, the intermediate triallelic 5-HTTLPR expressing group, but not the low expressing group, was associated with greater HP thresholds compared to the high expressing group.

CYP2D6*11 and challenges in clinical genotyping of the highly polymorphic CYP2D6 gene.

CYP2D6 is genotyped clinically for prediction of response to tamoxifen, psychotropic drugs and other medications. Phenotype prediction is dependent upon accurate genotyping. The CYP Allele Nomenclature Committee maintains the allelic nomenclature for CYP2D6; however, in some cases, the list of polymorphisms associated with a given allele is incomplete. Clinical laboratories and in vitro diagnostic manufacturers rely upon this nomenclature, in addition to the literature, to infer allelic function and haplotypes and when they design CYP2D6-testing platforms. This article provides more complete sequencing data for the CYP2D6*11 allele and describes the difficulties encountered in genotyping CYP2D6 when incomplete data are available. The CYP Allele Nomenclature Committee should provide clear information about the completeness of the original data used to define each allele.

Frequency of undetected CYP2D6 hybrid genes in clinical samples: impact on phenotype prediction.

Cytochrome P450 2D6 (CYP2D6) is highly polymorphic. CYP2D6-2D7 hybrid genes can be present in samples containing CYP2D6*4 and CYP2D6*10 alleles. CYP2D7-2D6 hybrid genes can be present in samples with duplication signals and in samples with homozygous genotyping results. The frequency of hybrid genes in clinical samples is unknown. We evaluated 1390 samples for undetected hybrid genes by polymerase chain reaction (PCR) amplification, PCR fragment analysis, TaqMan copy number assays, DNA sequencing, and allele-specific primer extension assay. Of 508 CYP2D6*4-containing samples, 109 (21.5%) harbored CYP2D6*68 + *4-like, whereas 9 (1.8%) harbored CYP2D6*4N + *4-like. Of 209 CYP2D6*10-containing samples, 44 (21.1%) were found to have CYP2D6*36 + *10. Of 332 homozygous samples, 4 (1.2%) harbored a single CYP2D7-2D6 hybrid, and of 341 samples with duplication signals, 25 (7.3%) harbored an undetected CYP2D7-2D6 hybrid. Phenotype before and after accurate genotyping was predicted using a method in clinical use. The presence of hybrid genes had no effect on the phenotype prediction of CYP2D6*4- and CYP2D6*10-containing samples. Four of four (100%) homozygous samples containing a CYP2D7-2D6 gene had a change in predicted phenotype, and 23 of 25 (92%) samples with a duplication signal and a CYP2D7-2D6 gene had a change in predicted phenotype. Four novel genes were identified (CYP2D6*13A1 variants 1 and 2, CYP2D6*13G1, and CYP2D6*13G2), and two novel hybrid tandem structures consisting of CYP2D6*13B + *68×2 + *4-like and CYP2D6*13A1 variant 2 + *1×N were observed.

Narcolepsy: immunological aspects.

Narcolepsy with cataplexy is a debilitating sleep disorder with an estimated prevalence of about 0.05%. Narcolepsy is caused by a selective loss of hypocretin (orexin) producing neurons in the perifornical hypothalamus. Based on the very strong association with the HLA subtype DQB1*0602, it is currently hypothesized narcolepsy is caused by an autoimmune-mediated process directed at the hypocretin neurons. So far however, studies focusing on general markers of (auto)immune activation, as well as humoral immunity against the hypocretin system have not yielded consistent results supporting this hypothesis.

Narcolepsy: a review of evidence for autoimmune diathesis.

Sporadic narcolepsy with cataplexy is a disabling disease that is strongly associated with the major histocompatibility class II allele HLA DQB1*0602 and is characterized by profound reduction in the cerebrospinal fluid (CSF) concentration of hypocretin 1 levels. This article provides a comprehensive review of the evidence that neurologic autoimmunity is the pathogenic basis of narcolepsy with cataplexy. Despite this evidence, specific antibody markers for narcolepsy have been elusive. Clinical trials using intravenous immunoglobulin infusions in recent onset narcolepsy with cataplexy have led to improvement in cataplexy in some patients. Future research must focus on elucidation of immune markers and early ameliorative treatments for narcolepsy.

Genome projects and gene therapy: gateways to next generation biological weapons.

Genomic and gene therapy research promise important gains into the treatment of human, animal, and plant diseases. However, there is a military aspect to this research that must be recognized. This research enables a new form of biological warfare named genomic warfare. This is the first peer-reviewed scientific article to discuss this threat in-depth. This article advocates that we begin to deal with this threat. Despite that the United States and many signatories of previous bioweapons treaties have agreed not to use biological weapons, mankind has a track record of using all of the weapons at its disposal. This article has a review of the literature and a basic overview of genomic research and gene therapy, which is followed by a discussion of how this therapy can be weaponized. How genomic warfare weapons might be deployed, how deployment may be detected, and the policies and research that would reduce this threat will be described. It is the aim of this article to clearly articulate that this risk exists and to encourage public health, scientific, political, and military leaders to take action to deal with the risk.

The voltage-gated calcium channel gamma subunits: a review of the literature.

Members of the voltage-gated calcium channel y subunit gene family (Cacng), have been rapidly discovered since the discovery of the identification of the mouse gamma2 gene (Cacng2) and its association with the stargazer mutant mouse line. The fact that this mutant mouse line exists has allowed researchers to gain insights into the function of the gamma2 subunit. For example, stargazer mice have elevated levels of neuropeptide Y production, very low cerebellar brain derived neurotrophic factor production, and diminished cerebellar GABAA alpha6 and beta3 production. Study of this mutant mouse line has also revealed that the gamma2 subunit is involved in AMPA receptor trafficking and targeting to the synaptic membrane. For the most part, the effect of gamma2 subunits on the electrophysiology of voltage-gated calcium channels is to downregulate calcium channel activity by causing a hyperpolarizing shift in the inactivation curve. This finding and the association of these subunits with AMPA receptor trafficking has led some researchers to question the actual role of the gamma subunits. This article reviews the discovery, cellular localization, tissue distribution, and function of the eight members of the Cacng family.

Search for neuron-specific and nonneuron-specific antibodies in narcoleptic patients with and without HLA DQB1*0602.

STUDY OBJECTIVES: Narcolepsy is strongly associated with the presence of HLA DQB1*0602. This and other evidence suggests that human narcolepsy is an autoimmune disease. This is in distinction to that found in canine models where hypocretin receptor 2 mutations are etiologic. We decided to test for the presence of several neuron-specific and organ-specific autoantibodies to see if they were present in HLA DQB1*0602-associated or cataplexy-associated narcolepsy or could serve as a serologic marker of the illness. DESIGN: We tested for N-type and P/Q-type voltage-gated calcium-channel antibodies, neuronal nicotinic acetylcholine receptor alpha3 subunit, acetylcholine receptor-binding antibodies, striated muscle antibodies, Type 1 Purkinje cell cytoplasmic antibodies, types 1 and 2 antineuronal nuclear antibodies and amphiphysin antibodies, GAD-65 antibody, and thyroid microsomal and thyroglobulin antibodies in the serum of 43 patients with or without cataplexy, 41 with known HLA status. SETTING: Narcoleptic subjects were recruited from the Mayo Sleep Disorders Center. PARTICIPANTS: N/A. INTERVENTIONS: N/A. MEASUREMENTS AND RESULTS: No antibody test yielded significantly positive results for the group as a whole or for subgroups of patients with cataplexy or positive HLA DQB1*0602 status. CONCLUSIONS: These results do not support the hypothesis that narcolepsy is an autoimmune disease. However, it is possible that the autoimmune attack is very selective and does not involve the epitopes measured in this study. Recent findings that the hypocretin neurotransmission system is involved in animal models of narcolepsy should lead to research to look for antibodies directed against components of the hypocretin neurotransmission system in narcolepsy.