Automated Interlaboratory Comparison of Therapeutic Drug Monitoring Data and Its Use for Evaluation of Published Therapeutic Reference Ranges.

Therapeutic drug monitoring is a tool for optimising the pharmacological treatment of diseases where the therapeutic effect is difficult to measure or monitor. Therapeutic reference ranges and dose-effect relation are the main requirements for this drug titration tool. Defining and updating therapeutic reference ranges are difficult, and there is no standardised method for the calculation and clinical qualification of these. The study presents a basic model for validating and selecting routine laboratory data. The programmed algorithm was applied on data sets of antidepressants and antipsychotics from three public hospitals in Denmark. Therapeutic analytical ranges were compared with the published therapeutic reference ranges by the Arbeitsgemeinschaft für Neuropsychopharmakologie und Pharmakopsychiatrie (AGNP) and in additional literature. For most of the drugs, the calculated therapeutic analytical ranges showed good concordance between the laboratories and to published therapeutic reference ranges. The exceptions were flupentixol, haloperidol, paroxetine, perphenazine, and venlafaxine + o-desmethyl-venlafaxine (total plasma concentration), where the range was considerably higher for the laboratory data, while the calculated range of desipramine, sertraline, ziprasidone, and zuclopenthixol was considerably lower. In most cases, we identified additional literature supporting our data, highlighting the need of a critical re-examination of current therapeutic reference ranges in Denmark. An automated approach can aid in the evaluation of current and future therapeutic reference ranges by providing additional information based on big data from multiple laboratories.

Simple and Robust Detection of CYP2D6 Gene Deletions and Duplications Using CYP2D8P as Reference.

Genotyping of the CYP2D6 gene is the most commonly applied pharmacogenetic test globally. Significant economic interests have led to the development of a plurality of assays, available for almost any genotyping platform or DNA detection chemistry. Of all the genetic variants, copy number variations are particular difficult to detect by polymerase chain reaction. Here, we present two simple novel approaches for the identification of samples carrying either deletions or duplications of the CYP2D6 gene; by relative quantification using a singleplex 5'nuclease real-time PCR assay, and by high-resolution melting of PCR products. These methods make use of universal primers, targeting both the CYP2D6 and the reference gene CYP2D8P, which is necessary for the analysis. The assays were validated against a reference method using a large set of samples. The singleplex nature of the 5'nuclease real-time PCR ensures that the primers anneal with equal affinity to both the sequence of the CYP2D6 and the reference gene. This facilitates robust identification of gene deletions and duplications based on the cycle threshold value. In contrast, the high-resolution melting assay is an end-point PCR, where the identification relies on variations between the amount of product generated from each of the two genes.

Pharmacogenetic testing revisited: 5' nuclease real-time polymerase chain reaction test panels for genotyping CYP2D6 and CYP2C19.

Due to their involvement in the metabolization of commonly prescribed psychopharmaceutical drugs, the cytochrome oxidase genes CYP2D6 and CYP2C19 are extensive targets for pharmacogenetic testing. The existence of common allelic variants allows the prediction of a metabolic phenotype based on a genotype result, hereby supplying a clinical tool for optimizing prescription and minimizing adverse effects. In this study, we present the development of two 5' nuclease real-time polymerase chain reaction (PCR) test panels, capable of detecting eight of the most clinically relevant alleles of the CYP2D6 gene (*2, *3, *4, *6, *9, *10, 17, *41) and the three most common nonfunctional alleles of CYP2C19 (*2, *3, *4). The assays have been thoroughly validated using a large collection of reference samples, by parallel testing and by DNA sequencing. The reanalysis of reference samples provided the calculation of the frequency of the CYP2D6*4K allele in a population, not previously reported. Furthermore, original test results from CYP2D6*41, generated based on the presence of the 2850T and the lack of the -1584G single-nucleotide polymorphism (SNP), were compared with genotyping based on the current acknowledged founder SNP 2988G of this allele. These results indicate that up to 17.7% of the patients originally tested as carriers of the CYP2D6*41 allele may have had an incorrect phenotypic result assigned. The two 5' nuclease real-time PCR test panels have subsequently been optimized for use in the clinical laboratory, using a standard real-time PCR instrument and software.

Marine mimivirus relatives are probably large algal viruses.

BACKGROUND: Acanthamoeba polyphaga mimivirus is the largest known ds-DNA virus and its 1.2 Mb-genome sequence has revealed many unique features. Mimivirus occupies an independent lineage among eukaryotic viruses and its known hosts include only species from the Acanthamoeba genus. The existence of mimivirus relatives was first suggested by the analysis of the Sargasso Sea metagenomic data. RESULTS: We now further demonstrate the presence of numerous "mimivirus-like" sequences using a larger marine metagenomic data set. We also show that the DNA polymerase sequences from three algal viruses (CeV01, PpV01, PoV01) infecting different marine algal species (Chrysochromulina ericina, Phaeocystis pouchetii, Pyramimonas orientalis) are very closely related to their homolog in mimivirus. CONCLUSION: Our results suggest that the numerous mimivirus-related sequences identified in marine environments are likely to originate from diverse large DNA viruses infecting phytoplankton. Micro-algae thus constitute a new category of potential hosts in which to look for new species of Mimiviridae.