AN IN VITRO INVESTIGATION INTO THE MECHANISM OF THE CLINICALLY RELEVANT DRUG-DRUG INTERACTION BETWEEN OMEPRAZOLE OR ESOMEPRAZOLE AND CLOPIDOGREL

Clopidogrel is a thienopyridine antiplatelet prodrug that was approved by the US FDA in 1997 and quickly supplanted ticlopidine as the primary drug therapy for reducing atherothrombotic events. It is converted to its pharmacologically active metabolite H4, which irreversibly inactivates the P2Y12 receptor on platelets, through two sequential reactions that are catalyzed mainly by CYP2C19. Common clinical practice involved the coadministration of a proton pump inhibitor (PPI, including omeprazole, esomeprazole, lansoprazole, pantoprazole, and rabeprazole) with clopidogrel to decrease the risk of upper gastrointestinal bleeding. This practice was formalized for high risk patients by the American Heart Association (and others) in 2008. By 2009, numerous publications described an unexpected decrease in clopidogrel efficacy when coadministered with PPIs, prompting both the US Food & Drug Administration (FDA) and European Medicines Agency (EMA) to issue recommendations discouraging the concomitant use of PPIs and clopidogrel. Proton pump inhibitors are also metabolized by CYP2C19. It seemed reasonable to conclude that, despite their relatively short plasma half-lives, PPIs might competitively inhibit CYP2C19, thereby reducing the efficacy of clopidogrel. In 2010, as numerous publications emerged, both regulatory agencies restricted subsequent warnings to only omeprazole and esomeprazole. The interaction between clopidogrel and PPIs, and the potential mechanisms responsible for it, continues to be a subject of much debate in 2015. This dissertation describes research that contributes to the progress made in understanding the basis for the interaction between clopidogrel and PPIs since the time of the initial regulatory statements, and in particular, why only omeprazole and esomeprazole are implicated in this drug interaction. The initial studies in this dissertation identified omeprazole (a racemic mixture of R- and S-enantiomers) and esomeprazole (the S-enantiomer) as not only competitive inhibitors, but more importantly, metabolism-dependent inhibitors (MDIs) of CYP2C19 in human liver microsomes (HLM), human hepatocytes and recombinant CYP2C19. In contrast, lansoprazole and pantoprazole did not cause metabolism-dependent inhibition (MDI) of CYP2C19. In addition to its clinical relevance, these observations are important because they underscore the importance of using a low concentration of enzyme and a short incubation time with the CYP marker substrate in order to detect MDI of CYP enzymes in vitro. In many previous studies of CYP2C19 inhibition by omeprazole or esomeprazole, the concentration of HLM was too high and/or the substrate incubation time was too long to detect MDI. The kinetic parameters for CYP2C19 inactivation by omeprazole, namely kinact and KI, were determined and used in a physiologically based pharmacokinetic (PBPK) model to predict the degree of CYP2C19 inactivation under clinical conditions. Omeprazole and esomeprazole were subsequently shown to be irreversible MDIs of CYP2C19, which explained why the decrease in clopidogrel efficacy could not be prevented in clinical studies by simply separating the doses of clopidogrel from omeprazole or esomeprazole. Subsequent studies demonstrated that, like the parent drug, two of the three major metabolites of omeprazole are also irreversible MDIs of CYP2C19. The kinetic parameters for CYP2C19 inactivation by these metabolites were determined and, along with those for omeprazole and esomeprazole, used in a mechanistic static model to predict the reduction of H4 formation from clopidogrel under clinical conditions. The model slightly overpredicted (by a factor of 2) the ability of omeprazole to block the conversion of clopidogrel to H4, its pharmacologically active metabolites, but otherwise established that inactivation of CYP2C19 is the likely mechanism for the clinical interaction between omeprazole/esomeprazole and clopidogrel. Esomeprazole and its two inhibitory metabolites, namely omeprazole sulfone and 5 O desmethylomeprazole, were subsequently determined to meet several criteria for mechanism-based inhibition (a special case of irreversible MDI). In addition, studies were initiated to test the hypothesis that the mechanism of CYP2C19 inactivation by esomeprazole and its metabolites involves the formation of a benzylic radical (on the 5,,S-methyl group) that binds covalently to the heme moiety. This hypothesis was based on the observation that the 5,,S methyl group is present on the pyridine ring of those compounds that irreversibly inactivate CYP2C19, namely omeprazole, esomeprazole, omeprazole sulfone and 5 O desmethylomeprazole, but absent from those compounds that did not inactivate CYP2C19, namely lansoprazole, pantoprazole and 5,,S-hydroxyomeprazole. Based on this hypothesis, the investigational PPI, tenatoprazole, which contains a 5,,S-methyl group, was correctly predicted to cause MDI of CYP2C19 whereas ilaprazole and rabeprazole, which lack a 5,,S-methyl group, did not cause MDI of CYP2C19. These results suggest that the investigational PPI, tenatoprazole, but not the clinically used PPIs ilaprazole or rabeprazole, may compromise the therapeutic effectiveness of clopidogrel. Finally, studies were performed in an attempt to provide direct evidence for the proposed mechanism of inactivation of CYP2C19 by esomeprazole, namely the formation of a heme adduct. The potential for the formation of a heme adduct in incubations of esomeprazole in HLM was evaluated by UHPLC analysis with UV/VIS detection and high resolution mass spectrometry (HRMS) with post-acquisition mass-defect filtering to identify heme and heme-containing adducts. Incubation of esomeprazole with NADPH-fortified HLM resulted in a substantial decrease in the amount of heme detectable by UHPLC with either UV absorbance or HRMS and appeared to show the formation of a heme adduct based on mass-defect filtering and isotopic distribution. However, the putative heme adduct was subsequently identified as a dimer of esomeprazole sulfone (a metabolite of esomeprazole formed by CYP3A4/5). Although an adduct between heme and a metabolite of esomeprazole was not ultimately identified, the potential for an unusual analytical artifact was revealed; namely, that sulfur-containing drugs can be converted to metabolites that closely resemble a heme adduct based on mass-defect filtering and isotopic distribution. In summary, this dissertation supports the hypothesis that irreversible inactivation of CYP2C19 is the mechanism by which omeprazole and esomeprazole reduce the efficacy of clopidogrel. This property is not shared by lansoprazole, pantoprazole, rabeprazole or ilaprazole. These findings support regulatory agencies¡¦ recommendations that, in order to reduce the risk of gastrointestinal bleeding, clopidogrel should not be coadministered with omeprazole or esomeprazole but should be coadministered with other PPIs that do not inactivate CYP2C19

The human gut virome: form and function

Advances in next generation sequencing technologies and the application of metagenomic approaches have fuelled an exponential increase in our understanding of the human gut microbiome. These approaches are now also illuminating features of the diverse and abundant collection of viruses (termed the virome) subsisting with the microbial ecosystems residing within the human holobiont. Here we focus on the current and emerging knowledge of the human gut virome, with a particular focus on viruses infecting bacteria (bacteriophage or phage), which are a dominant component of this viral community. We summarise current insights regarding the form and function of this ‘human gut phageome’ and highlight promising avenues for future research. In doing so we discuss the potential for phage to drive ecological functioning and evolutionary change within this important microbial ecosystem, their contribution to modulation of host-microbiome interactions and stability of the community as a whole, as well as the potential role of the phageome in human health and disease. We also consider the emerging concepts of a ‘core healthy gut phageome’ and the putative existence of ‘viral enterotypes’ and ‘viral dysbiosis’

Too Little, Too Late: How the Tidal Evolution of Hot Jupiters affects Transit Surveys of Clusters

The tidal evolution of hot Jupiters may change the efficiency of transit surveys of stellar clusters. The orbital decay that hot Jupiters suffer may result in their destruction, leaving fewer transiting planets in older clusters. We calculate the impact tidal evolution has for different assumed stellar populations, including that of 47~Tuc, a globular cluster that was the focus of an intense HST search for transits. We find that in older clusters one expects to detect fewer transiting planets by a factor of two for surveys sensitive to Jupiter-like planets in orbits out to 0.5~AU, and up to a factor of 25 for surveys sensitive to Jupiter-like planets in orbits out to 0.08~AU. Additionally, tidal evolution affects the distribution of transiting planets as a function of semi-major axis, producing larger orbital period gaps for transiting planets as the age of the cluster increases. Tidal evolution can explain the lack of detected exoplanets in 47~Tuc without invoking other mechanisms. Four open clusters residing within the {\em Kepler} fields of view have ages that span 0.4-8~Gyr--if {\em Kepler} can observe a significant number of planets in these clusters, it will provide key tests for our tidal evolution hypothesis. Finally, our results suggest that observers wishing to discover transiting planets in clusters must have sufficient accuracy to detect lower mass planets, search larger numbers of cluster members, or have longer observation windows to be confident that a significant number of transits will occur for a population of stars.Comment: 23 pages including 6 figures, accepted to Ap

Constraining Tidal Dissipation in Stars from The Destruction Rates of Exoplanets

We use the distribution of extrasolar planets in circular orbits around stars with surface convective zones detected by ground based transit searches to constrain how efficiently tides raised by the planet are dissipated on the parent star. We parameterize this efficiency as a tidal quality factor (Q*). We conclude that the population of currently known planets is inconsistent with Q*<10^7 at the 99% level. Previous studies show that values of Q* between 10^5 and 10^7 are required in order to explain the orbital circularization of main sequence low mass binary stars in clusters, suggesting that different dissipation mechanisms might be acting in the two cases, most likely due to the very different tidal forcing frequencies relative to the stellar rotation frequency occurring for star--star versus planet--star systems.Comment: 12 pages, 6 figures accepted for publication in Ap

Feasibility and Acceptability of a Mobile Technology Intervention to Support Postabortion Care in British Columbia: Phase I.

BACKGROUND: Over 30% of women in Canada undergo an abortion. Despite the prevalence of the procedure, stigma surrounding abortion in Canada leads to barriers for women to access this service. The vast majority of care is concentrated in urban settings. There is evidence to support utilization of innovative mobile and other technology solutions to empower women to safely and effectively self-manage aspects of the abortion process. This study is part 1 of a 3-phase study that utilizes user-centered design methodology to develop a digital health solution to specifically support follow-up after an induced surgical abortion. OBJECTIVE: This study aimed to (1) understand how women at 3 surgical abortion clinics in an urban center of British Columbia utilize their mobile phones to access health care information and (2) understand women's preferences of content and design of an intervention that will support follow-up care after an induced abortion, including contraceptive use. METHODS: The study design was based on development-evaluation-implementation process from Medical Research Council Framework for Complex Medical Interventions. This was a mixed-methods formative study. Women (aged 14-45 years) were recruited from 3 urban abortion facilities in British Columbia who underwent an induced abortion. Adaptation of validated surveys and using the technology acceptance model and theory of reasoned action, a cross-sectional survey was designed. Interview topics included demographic information; type of wireless device used; cell phone usage; acceptable information to include in a mobile intervention to support women's abortion care; willingness to use a mobile phone to obtain reproductive health information; optimal strategies to use a mobile intervention to support women; understand preferences for health information resources; and design qualities in a mobile intervention important for ease of use, privacy, and security. Responses to questions in the survey were summarized using descriptive statistics. Qualitative analysis was conducted with NVivo using a thematic analysis approach. This study was approved by the local ethics board. RESULTS: A waiting-room survey was completed by 50 participants, and semistructured interviews were completed with 8 participants. The average age of participants was 26 years. Furthermore, 94% (47/50) owned a smartphone, 85% (41/48) used their personal phones to go online, and 85% would use their cell phone to assist in clinical care. Qualitative analysis demonstrated that women prefer a comprehensive website that included secure email or text notifications to provide tools and resources for emotional well-being, contraceptive decision making, general sexual health, and postprocedure care. CONCLUSIONS: A community-based mixed-methods approach allowed us to understand how women use their cell phones and what women desire in a mobile intervention to support their postabortion care. The findings from this formative phase will assist in the development and testing of a mobile intervention to support follow-up care after an induced surgical abortion

Feasibility and Acceptability of a Mobile Technology Intervention to Support Postabortion Care (The FACTS Study Phase II) After Surgical Abortion: User-Centered Design.

BACKGROUND: Human-centered design is a methodology that applies an iterative participatory process that engages the end-user for whom an innovation or intervention is designed for from start to end. There is general evidence to support the use of human-centered design for development of tools to affect health behavior, but specifically for family planning provision. This study is part two of a three-phase study that uses a user-centered design methodology which uses the findings from Phase I to design, develop, and test a digital health solution to support follow-up after an induced surgical abortion. OBJECTIVE: The objectives for this study were to: (1) develop a Web-based intervention based on preferences and experiences of women who underwent an abortion as measured in the formative phase of the Feasibility and Acceptability of a Mobile Technology Intervention to Support Postabortion Care Study; (2) conduct usability testing of the intervention to determine user-friendliness and appropriateness of the intervention; and (3) finalize a beta version of the Web-based intervention for pilot testing. METHODS: The study design was based on the "development-evaluation-implementation" process from the Medical Research Council Framework for Complex Medical Interventions. This study is in Phase II of III and is based on user-centered design methodology. Phase I findings demonstrated that women engage with technology to assist in clinical care and they preferred a comprehensive website with email or text notifications to support follow-up care. In Phase II we collaborated with family planning experts and key stakeholders to synthesize evidence from Phase I. With them and a development partner we built a prototype. Usability testing was completed with 9 participants using a validated System Usability Scale. This was then used to refine the intervention for Phase III pilot study. This study was approved by the local Ethics board. RESULTS: We developed a comprehensive Web-based tool called myPostCare.ca, which includes: Post-Procedure Care, Emotional Well-Being Tool, Contraception Explorer, Sexual Health, Book an Appointment, and Other Resources. Additionally, over the course of a month after the procedure, automatic email notifications were sent to women as a form of virtual follow-up support, directing them to myPostCare.ca resources. The Web-based tool was refined based on usability testing results. CONCLUSIONS: This study demonstrated that user-centered design is a useful methodology to build programs and interventions that are women-centered, specifically for abortion care

The Roles of Tidal Evolution and Evaporative Mass Loss in the Origin of CoRoT-7 b

CoRoT-7 b is the first confirmed rocky exoplanet, but, with an orbital semi-major axis of 0.0172 AU, its origins may be unlike any rocky planet in our solar system. In this study, we consider the roles of tidal evolution and evaporative mass loss in CoRoT-7 b's history, which together have modified the planet's mass and orbit. If CoRoT-7 b has always been a rocky body, evaporation may have driven off almost half its original mass, but the mass loss may depend sensitively on the extent of tidal decay of its orbit. As tides caused CoRoT-7 b's orbit to decay, they brought the planet closer to its host star, thereby enhancing the mass loss rate. Such a large mass loss also suggests the possibility that CoRoT-7 b began as a gas giant planet and had its original atmosphere completely evaporated. In this case, we find that CoRoT-7 b's original mass probably didn't exceed 200 Earth masses (about 2/3 of a Jupiter mass). Tides raised on the host star by the planet may have significantly reduced the orbital semi-major axis, perhaps causing the planet to migrate through mean-motion resonances with the other planet in the system, CoRoT-7 c. The coupling between tidal evolution and mass loss may be important not only for CoRoT-7 b but also for other close-in exoplanets, and future studies of mass loss and orbital evolution may provide insight into the origin and fate of close-in planets, both rocky and gaseous.Comment: Accepted for publication by MNRAS on 2010 May

Genome signature-based dissection of human gut metagenomes to extract subliminal viral sequences

Bacterial viruses (bacteriophages) have a key role in shaping the development and functional outputs of host microbiomes. Although metagenomic approaches have greatly expanded our understanding of the prokaryotic virosphere, additional tools are required for the phage-oriented dissection of metagenomic data sets, and host-range affiliation of recovered sequences. Here we demonstrate the application of a genome signature-based approach to interrogate conventional whole-community metagenomes and access subliminal, phylogenetically targeted, phage sequences present within. We describe a portion of the biological dark matter extant in the human gut virome, and bring to light a population of potentially gut-specific Bacteroidales-like phage, poorly represented in existing virus like particle-derived viral metagenomes. These predominantly temperate phage were shown to encode functions of direct relevance to human health in the form of antibiotic resistance genes, and provided evidence for the existence of putative ‘viral-enterotypes’ among this fraction of the human gut virome