Genotyping Performance between Saliva and Blood-Derived Genomic DNAs on the DMET Array: A Comparison

The Affymetrix Drug Metabolism Enzymes and Transporters (DMET) microarray is the first assay to offer a large representation of SNPs conferring genetic diversity across known pharmacokinetic markers. As a convenient and painless alternative to blood, saliva samples have been reported to work well for genotyping on the high density SNP arrays, but no reports to date have examined this application for saliva-derived DNA on the DMET platform. Genomic DNA extractions from saliva samples produced an ample quantity of genomic DNA for DMET arrays, however when human amplifiable DNA was measured, it was determined that a large percentage of this DNA was from bacteria or fungi. A mean of 37.3% human amplifiable DNA was determined for saliva-derived DNAs, which results in a significant decrease in the genotyping call rate (88.8%) when compared with blood-derived DNAs (99.1%). More interestingly, the percentage of human amplifiable DNA correlated with a higher genotyping call rate, and almost all samples with more than 31.3% human DNA produced a genotyping call rate of at least 96%. SNP genotyping results for saliva derived DNA (n = 39) illustrated a 98.7% concordance when compared with blood DNA. In conclusion, when compared with blood DNA and tested on the DMET array, saliva-derived DNA provided adequate genotyping quality with a significant lower number of SNP calls. Saliva-derived DNA does perform very well if it contains greater than 31.3% human amplifiable DNA

Feasibility of pharmacy-initiated pharmacogenetic screening for CYP2D6 and CYP2C19

PURPOSE: Our purpose was to investigate the feasibility of pharmacy-initiated pharmacogenetic (PGt) screening in primary care with respect to patient willingness to participate, quality of DNA collection with saliva kits, genotyping, and dispensing data retrieved from the pharmacy. METHODS: Polypharmacy patients aged >60 years who used at least one drug with Anatomical Therapeutic Chemical (ATC) code N06AA01-N06AX19 (antidepressants), A02BC01-A02BC05 (proton-pump inhibitors), N05AA01-N05AH04 (antipsychotics), or C07AB02 (metoprolol) in the preceding 2 years were randomly selected. DNA was collected with saliva kits and genotyped for CYP2D6 and CYP2C19 with the AmpliChip. Pharmacy dispensing records were retrieved and screened for drugs interacting with the patient's CYP2D6 and CYP2C19 genotype by using the evidence-based PGt guidelines from the Dutch Pharmacogenetics Working Group. RESULTS: Out of the 93 invited patients, 54 (58.1%) provided informed consent. Nine saliva samples (16.7%) contained too little DNA. Call rates for CYP2D6 and CYP2C19 were 93.3% and 100%, respectively. Frequencies of genotype-predicted phenotype were 2.4%, 38.1%, 54.8%, and 4.8% for CYP2D6 poor metabolizers (PM), intermediate metabolizers (IM), extensive metabolizers (EM), and ultrarapid metabolizers (UM) respectively. For CYP2C19 genotype-predicted phenotype, frequencies were 2.2%, 15.6%, and 82.2% for PM, IM, and EM, respectively. CONCLUSIONS: This study shows that pharmacy-initiated PGt screening is feasible for a primary care setting

Do nuclear DNA and dental nonmetric data produce similar reconstructions of regional population history? An example from modern coastal Kenya

This study investigates whether variants in dental morphology and nuclear DNA provide similar patterns of intergroup affinity among regional populations using biological distance (biodistance) estimates. Many biodistance studies of archaeological populations use skeletal variants in lieu of ancient DNA, based on the widely accepted assumption of a strong correlation between phenetic- and genetic-based affinities. Within studies of dental morphology, this assumption has been well supported by research on a global scale but remains unconfirmed at a more geographically restricted scale. Paired genetic (42 microsatellite loci) and dental (nine crown morphology traits) data were collected from 295 individuals among four contemporary Kenyan populations, two of which are known ethnically as “Swahili” and two as “Taita;” all have welldocumented population histories. The results indicate that biodistances based on genetic data are correlated with those obtained from dental morphology. Specifically, both distance matrices indicate that the closest affinities are between population samples within each ethnic group. Both also identify greater divergence among samples from the different ethnic groups. However, for this particular study the genetic data may provide finer resolution at detecting overall among-population relationships

Extrahepatic cytochrome P450s - Relation to cancer susceptibility

Cytochrome P450 (CYP) enzymes in extrahepatic tissues may play important roles in regulating the capacity of individual cells to metabolize environmental carcinogens and hormones. In this thesis, we characterized the cytochrome P450 profile of the human normal breast and investigated whether polymorphisms in the genes encoding the estrogen metabolizing enzymes CYP1131 and Catechol-O-methyltransferase (COMT) were associated with breast or endometrial cancer risk. We also identified, cloned and characterized the novel extrahepatic cytochrome P450 2S1 (CYP2S1) enzyme with a possible role in metabolism of xenobiotics. Detected levels of CYP enzymes in extrahepatic tissues are generally lower than in the liver and thus difficult to measure. By partial purification of CYPs from breast reduction samples, we spectrally quantified the level in the human normal breast to be approximately 1000-fold lower than in the human liver. We characterized the CYP expression pattern by RTPCR and Western blot in 15 samples and found enzymes involved in the metabolism of environmental carcinogens, steroid hormones and drugs. A large interindividual variation in the expression of CYP1A1, 2A6, 2B6, 2D6 and 3A were shown whereas CYP1B1, 2C, 2E 1, 4A and 19 was present in most samples. Molecular epidemiological studies on the association between polymorphic variants of enzymes involved in estrogen biosynthesis and metabolism and hormonal cancer risk are published with rapidly increasing frequency. However, reported data are generally inconsistent, partly due to small sample sizes. We investigated the association between CYP1B1 and COMT genotypes, respectively, and breast and endometrial cancer (only CYP1B1) in a large population-based case-control study on Swedish postmenopausal women. We genotyped approximately 1 500 breast cancer cases, 690 endometrial cancer cases and 1 500 controls and calculated odds ratios and 95 percent confidence intervals from conditional logistic regression models. We found no overall association between CYP1B1 or COMT genotype and breast cancer risk. Stratified analyses indicated that the CYP1B1*3/*3 genotype may be of importance in conjunction with menopausal hormone use and that the low activity allele of COMT appeared associated with an increased risk for lobular breast cancer. CYP1B1 genotype had no effect on endometrial cancer risk, neither in overall nor in subgroup analyses. The CYP2S1 gene was identified through homology searches with known CYP sequences against the EST database and the high throughput genomic sequences database. The gene was found to be located together with the genes of subfamily CYP2A, CYP2B and CYP2F on chromosome 19. The predicted 504 amino acid sequence displayed 38-49 percent identity with other CYP2 family members and contained the typical structural CYP characteristics; the conserved cystein, the proline rich region and the N-terminal hydrophobic stretch. CYP2S1 mRNA was shown to be highly expressed in lung, trachea, and stomach. The transcript was detected in the liver but at lower levels. A rabbit peptide antiserum against the C-terminus of CYP2S1 recognized a single band in human lung with an apparent molecular weight of 50 kD. Subcellular localization and immunocytochemistry revealed CYP2S1 to be a microsomal protein. High expression in respiratory and gastrointestinal tract indicates a role for this enzyme in extrahepatic: xenobiotic metabolism. Unpublished observations from our lab show that small aromatic hydrocarbons appear to be substrates for CYP2S1. In summary, our work have contributed to the understanding of the presence of CYPs in extrahepatic tissues, particularly breast and lung, and evaluated a possible role for genetic polymorphisms in estrogen metabolism genes in relation to hormonal cancer susceptibility