This thesis combines five published research papers investigating methods of improving the safety and control of oral anticoagulation, with the use of computer assisted dosage and the standardisation of the International Normalised Ratio (INR). The INR is a conventional measurement derived from the time it takes blood of a patient to form a clot and is used to monitor the effects of widely used oral anticoagulants such as warfarin for the prevention of stroke and other related disorders. The first paper investigates whether the use of computer-assisted programs was as safe and effective as medical staff manual dosage in the prevention of bleeding or thrombotic complications during oral anticoagulant treatment. This was an international multi-centre randomised study conducted by the European Action on Anticoagulation (EAA) investigating the clinical benefit of two computer programs, PARMA 5 (Italy) and DAWN AC (UK). Composite clinical events were reduced by 7.6% using computer programs, though not achieving statistical significance (p=0.1), showing computer programs to be not dissimilar to medical staff dosage. The second paper recommends guidelines for screening safety and effectiveness of other marketed computer programs based on the results of the EAA study. A process for a candidate computer program to achieve non-inferiority relative to the medical staff dosage arm from the EAA study is explained. The third paper introduces a modified approach to the 'Direct INR' method for the standardisation of INR termed the 'Prothrombin Time/INR Line' (PT/INR). This was directly compared to the local International Sensitivity Index (ISI) calibration procedure originally approved by the World Health Organisation and later by the United States Food and Drug Administration (FDA). Using manually certified lyophilised plasmas tested by specialist centres, the PT/INR Line using a set of 5 calibrant plasmas to establish a fitted line to estimate local INR was shown to be as effective as the FDA procedure. The fourth paper investigates the PT/INR Line further by using simulated sets of calibrant plasmas across the therapeutic range of 2.0-4.5 INR and determining the PT/INR Line. Local INR of five validation plasmas, certified by 3 centres using the manual PT technique, was determined using the estimated PT/INR Lines and compared with local ISI calibration. Using 4 or 5 calibrant plasmas to determine the PT/INR Line was shown to be as accurate as local ISI calibrations for reliable local INR.The fifth and final paper assessed INR variability and control in oral anticoagulant therapy using a method termed the Variance Growth Rate (VGR), and compared its predictive ability of adverse events with the Time in Target INR range (TIR), the conventional method used in evaluating the quality of oral anticoagulant therapy. The VGR method was shown to be a better predictor of adverse bleeding or thrombotic episodes in the short term period prior to an event (3 and 6 months) compared with TIR.

Resumo: A vitamina K, descoberta em 1929, tem reconhecida função na coagulação sangüínea. Esta vitamina age como co-fator na reação de carboxilação para biossíntese, após tradução, de resíduos de ácido gama carboxi-glutâmico (Gla) em limitado número de proteínas. A reação de carboxilação é passo necessário para a formação de fatores e proteínas da coagulação (fatores II, VII, IX, X, proteínas C e S) e proteínas envolvidas no metabolismo ósseo, como osteocalcina e proteína Gla da matriz. As drogas anticoagulantes orais são usadas na prevenção primária e secundária de fenômenos tromboembólicos e funcionam reduzindo a atividade plasmática das proteínas de coagulação vitamina K-dependentes. As causas da instabilidade da terapia anticoagulante são múltiplas e incluem: baixa adesão ao tratamento, interação droga-droga, resistência herdada à varfarina e alterações dietéticas. O presente trabalho versa sobre a influência do estado nutricional relacionado à vitamina K sobre a terapia de anticoagulação oral, em pacientes portadores de doença vascular. Levantou-se a hipótese de que o consumo de dieta com teor inadequado de vitamina K (elevado ou reduzido), assim como as concentrações plasmáticas da vitamina, podem estar associados a dificuldades na promoção da anticoagulação ou favorecer o estado de hipocoagulabilidade. Objetivou-se ainda a realização de avaliação do estado nutricional relacionado à vitamina K do grupo estudado, identificar as principais fontes dietéticas de vitamina K e investigar o efeito de diversas variáveis (chamadas variáveis descritivas) sobre a ingestão e concentrações plasmáticas de vitamina K. Para atender aos objetivos propostos, o trabalho incluiu dois estudos: transversal e longitudinal. Do estudo transversal, constam a aplicação de protocolo, que contém: os dados... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Vitamin K, which was discovered in the decade of 1930, has a recognized function in blood coagulation. The vitamin acts as a cofactor in posttranslational conversion of specific glutamyl residues to gammacarboxyglutamic acid (GLA) residues in limited numbers of proteins. The carboxylation reaction is a necessary step to synthesis of factors and anticoagulant proteins (factors II , VII, IX, X, proteins C and S) and proteins involved in bone metabolism, such as osteocalcin and matrix GLA-protein. The oral anticoagulant drugs have been used in the primary and secondary prevention of thromboembolic disorders and act reducing the plasmatic activity of blood coagulation proteins. Causes of the unstable anticoagulant therapy are multiple and include poor compliance, drug-drug interaction, inherited warfarin resistance and dietary changes. The present study examined the influence of vitamin K nutritional status on the warfarin anticoagulation therapy in vascular disease patients. We tested the hypothesis that inadequate diet intake of vitamin K (high or low), and the vitamin plasmatic concentrations are related to a disturbances of the coagulation leading to hypercoagulability or favoring hypocoagulability condition. The aim of the study was to assess the vitamin K nutritional status of the group, identify important food sources and investigate the effect of some variables on the dietary vitamin K intake and the vitamin plasmatic concentration. The work included a crosssectional and longitudinal study. The cross-sectional study consisted of clinical history and examination, a 24-hour recall method for assessment of recent food intake of vitamin K and laboratory methods for biochemical assessment, including coagulation test and body composition techniques. For the longitudinal study, a food frequency questionnaire was applied in order to... (Complete abstract click electronic access below) / Orientador: Sérgio Alberto Rupp de Paiva / Coorientador: Álvaro Oscar Campana / Doutor

Phylloquinone is a natural analog of vitamin K that has been shown to both inhibit cancer cell growth and induce apoptosis in several cancer cell lines. This study examined these effects in a non-Hodgkin lymphoma cell line, known as U937. Cell growth inhibition and apoptosis were assessed through the quantification of cell density and area, following treatment with several concentrations of phylloquinone. In addition, apoptosis was detected and quantified using immunofluorescent markers of apoptosis (i.e. annexin V, APO-BrdU). Treatment with phylloquinone resulted in reduced overall cell density, increased overall cell area, and an increased frequency of apoptosis in U937 cells. Increasing both phylloquinone concentration and treatment time enhanced these effects. These results are significant because they document the anti-cancer effects of this analog of vitamin K, as well as provide insight into the morphological changes that occur during apoptosis in U937 cells.

Les vitamines D et K sont des micronutriments liposolubles qui participent au bon fonctionnement de l’organisme. Elles jouent des rôles clés dans la prévention de trouble de l'hémostase et de la coagulation, des pathologies osseuses, métaboliques et cardiovasculaires. Cependant, même si ces vitamines sont apportées en quantités suffisantes par notre alimentation, leurs effets bénéfiques sont étroitement conditionnés par leur biodisponibilité. Or, mieux connaitre les mécanismes d’absorption permettrait d’appréhender leur biodisponibilité. Nous avons tout d’abord montré que l’absorption de la vitamine K implique des transporteurs du cholestérol, SR-B1 et CD36. Nous avons également montré que l’entérocyte est non seulement capable d’effluer les vitamines D et K néo-absorbées mais également d’excréter ces vitamines du compartiment sanguin vers la lumière intestinale. Ce phénomène bien connu pour le cholestérol (excrétion transintestinale du cholestérol) implique des transporteurs communs, dont ABCB1 et ABCG5/G8. Dans un second temps, dans le cadre de la relance de la consommation des légumineuses, nous avons mis en évidence que la présence de légumineuses dans un repas limite la biodisponibilité de ces vitamines. En effet, les fibres, phytates, saponines et tanins diminuent leur bioaccessibilité et/ou leur captage. La méthode de cuisson des légumineuses, en affectant leur composition nutritionnelle, peut moduler l’incorporation des vitamines D et K au sein des micelles mixtes et donc affecter leur biodisponibilité. Ces données soulignent ainsi le fait que les légumineuses doivent être cuites de manière appropriée et consommés dans des repas riches en micronutriments. / Vitamin D and K are fat-soluble micronutrients that participate to the proper functioning of the organism. They are essential to prevent bleeding, bone, metabolic and cardiovascular disorders. However, even if those vitamins are provided in sufficient quantities in our diet, their health effects are closely linked to their bioavailability. A better knowledge of their absorption mechanisms would help to optimize their bioavailability.Firstly, we showed that vitamin K absorption involves the cholesterol transporters SR-B1 and CD36. We also showed that enterocytes can not only efflux newly absorbed vitamins D and K but also excrete vitamin D and K from the blood compartment to the intestinal lumen. This phenomenon of transintestinal excretioninvolves the cholesterol membrane transporters ABCB1 and ABCG5/G8.Secondly, we showed that the presence of pulses within a meal limits vitamin D and K bioavailability. Indeed, fibers, phytates, saponins and tannins can decrease bioaccessibility and/or uptake of vitamin K. By modulating the nutritional profile of pulses, the cooking method can impact on fat-soluble vitamin transfer to mixed micelles, and in turn affect their bioavailability. These data suggest that pulses must be cooked in an appropriate manner and consumed in micronutrient-rich meals.Keywords: vitamin D, vitamin K, bioaccessibility, intestinal absorption, pulses.

Background:Compounded vitamin K oral liquids may be useful in some patient populations, or when an appropriate solid dosage form is not available. While vitamin K oral liquid is typically prepared with sterile water for injection (SWFI), other compounding agents may be more palatable.Objective:To evaluate stability of compounded vitamin K liquids in SWFI, Ora-Sweet, simple syrup, cherry syrup, and SyrPalta stored in amber plastic oral syringes.Methods:Five types of compounded vitamin K liquids were prepared in triplicate—Ora-Sweet, simple syrup, cherry syrup, SyrPalta, and SWFI without flavoring; aliquoted into amber plastic oral syringes; and stored in a laboratory refrigerator (4.9°C to 5.4°C). On study days, 3 syringes from each batch were removed, diluted to assay concentration, and compared with a freshly prepared US Pharmacopeia reference solution. The samples and reference were analyzed using a previously validated high-performance liquid chromatography–ultraviolet method. Product stability was defined as 90% to 110% labeled amount. Results were further compared using a 2-way ANOVA (analysis of variance; P = .05) with post hoc Tukey’s correction for multiple comparisons.Results:Vitamin K in SWFI, SyrPalta, and cherry syrup was stable for 21 days, 7 days, and 24 hours, respectively, under refrigeration in amber plastic oral syringes. Vitamin K in Ora-Sweet and simple syrup demonstrated high within-day variability and low potency. Statistically significant differences were detected between the SWFI formulation and all other vehicles.Conclusion:Vitamin K in SWFI is appropriate for longer-term storage of unit-dosed vitamin K; however, SyrPalta and cherry syrup may be used for short-term storage or immediate administration of vitamin K.

Vitamin K has a negligible effect on bone mineral density (BMD) and a large but uncertain effect on fractures. The three studies in the thesis explored uncertainty about the effect of vitamin K on fractures using the methods of economic evaluation and Bayesian meta-analysis. In study 1, a Markov probabilistic microsimulation model was developed for a hypothetical cohort of 50-year-old postmenopausal women without osteoporosis. This was a fracture incidence-based model, populated with data from the literature. It was used to examine the cost-effectiveness of two supplementation strategies over a lifetime horizon. We compared vitamin K2 (or vitamin K1) concurrent with vitamin D3 and calcium versus vitamin D3 and calcium alone. Study 2 included a systematic review, and classical and Bayesian univariate meta-analyses to determine the efficacies of the K vitamins on BMD or fractures in current and future trials. Study 3 used Bayesian bivariate random-effects meta-analysis to jointly model the treatment effects on two correlated bone outcomes. We compared the estimates from the univariate and bivariate meta-analyses and explored how these results would change the conclusions of the cost-effectiveness analysis. The strategies including vitamin K were highly cost-effective at willingness-to-pay of $50,000/QALY (quality-adjusted life year); however, the results were most sensitive to changes in the efficacy of vitamin K. The univariate meta-analyses showed large uncertainties in the anti-fracture effects of vitamin K2 in current and future trials. The bivariate 95% credible intervals were considerably narrower than those from the univariate meta-analyses. Using future odds ratios from the bivariate meta-analyses, vitamin K2 cost more than $100,000/QALY while vitamin K1 was cost-saving. Our analyses found substantial uncertainty around the estimates of the vitamin K effect on fractures. We recommend against routine use of vitamin K for fracture prevention. Bayesian bivariate meta-analysis accounts for all available information and should be considered when the treatment effects are measured on two correlated outcomes.

Vitamin K has a negligible effect on bone mineral density (BMD) and a large but uncertain effect on fractures. The three studies in the thesis explored uncertainty about the effect of vitamin K on fractures using the methods of economic evaluation and Bayesian meta-analysis. In study 1, a Markov probabilistic microsimulation model was developed for a hypothetical cohort of 50-year-old postmenopausal women without osteoporosis. This was a fracture incidence-based model, populated with data from the literature. It was used to examine the cost-effectiveness of two supplementation strategies over a lifetime horizon. We compared vitamin K2 (or vitamin K1) concurrent with vitamin D3 and calcium versus vitamin D3 and calcium alone. Study 2 included a systematic review, and classical and Bayesian univariate meta-analyses to determine the efficacies of the K vitamins on BMD or fractures in current and future trials. Study 3 used Bayesian bivariate random-effects meta-analysis to jointly model the treatment effects on two correlated bone outcomes. We compared the estimates from the univariate and bivariate meta-analyses and explored how these results would change the conclusions of the cost-effectiveness analysis. The strategies including vitamin K were highly cost-effective at willingness-to-pay of $50,000/QALY (quality-adjusted life year); however, the results were most sensitive to changes in the efficacy of vitamin K. The univariate meta-analyses showed large uncertainties in the anti-fracture effects of vitamin K2 in current and future trials. The bivariate 95% credible intervals were considerably narrower than those from the univariate meta-analyses. Using future odds ratios from the bivariate meta-analyses, vitamin K2 cost more than $100,000/QALY while vitamin K1 was cost-saving. Our analyses found substantial uncertainty around the estimates of the vitamin K effect on fractures. We recommend against routine use of vitamin K for fracture prevention. Bayesian bivariate meta-analysis accounts for all available information and should be considered when the treatment effects are measured on two correlated outcomes.

La vitamine K fait l’objet d’un intérêt croissant en regard du rôle qu’elle peut jouer dans la santé humaine hormis celui bien établi dans la coagulation sanguine. De plus en plus d’études expérimentales lui confèrent des fonctions dans le système nerveux central, particulièrement dans la synthèse des sphingolipides, l’activation de la protéine vitamine K-dépendante Gas6 et la protection contre les dommages oxydatifs. Toutefois, il demeure beaucoup moins bien établi si la perturbation de ces fonctions peut conduire à des déficits cognitifs. L’objectif principal de cette thèse est de vérifier l’hypothèse selon laquelle le statut vitaminique K des personnes âgées en santé est un déterminant de la performance cognitive. En vue de la réalisation de cet objectif, une meilleure compréhension des indicateurs du statut vitaminique K s’avérait nécessaire. Chacune des études présentées vise donc un objectif spécifique : 1) évaluer le nombre de rappels alimentaires de 24 heures non consécutifs nécessaire pour mesurer l’apport habituel de vitamine K des personnes âgées; 2) évaluer la valeur d’une seule mesure de la concentration sérique de vitamine K comme marqueur de l’exposition à long terme; et 3) examiner l’association entre le statut vitaminique K et la performance cognitive des personnes âgées en santé de la cohorte québécoise NuAge. Trois dimensions cognitives ont été évaluées soient la mémoire épisodique verbale et non-verbale, les fonctions exécutives et la vitesse de traitement de l’information. Cette thèse présente la première étude appuyant l’hypothèse d’un rôle de la vitamine K dans la cognition chez les personnes âgées. Spécifiquement, la concentration sérique de vitamine K a été associée positivement à la performance en mémoire épisodique verbale, et plus particulièrement au processus de consolidation de la trace mnésique. En accord avec les travaux chez l’animal et l’action de la protéine Gas6 dans l’hippocampe, un rôle spécifique de la vitamine K à l’étape de consolidation est biologiquement plausible. Aucune association significative n’a été observée avec les fonctions exécutives et la vitesse de traitement de l’information. Parallèlement, il a été démontré qu’une mesure unique de la concentration sérique de vitamine K constitue une mesure adéquate de l’exposition à long terme à la vitamine K. De même, il a été établi que six à 13 rappels alimentaires de 24 heures sont nécessaires pour estimer précisément l’apport de vitamine K des personnes âgées en santé. Collectivement, les résultats de ces deux études fournissent des informations précieuses aux chercheurs permettant une meilleure interprétation des études existantes et une meilleure planification des études futures. Les résultats de cette thèse constituent une avancée importante dans la compréhension du rôle potentiel de la vitamine K dans le système nerveux central et renforce la nécessité qu’elle soit considérée en tant que facteur nutritionnel du vieillissement cognitif, en particulier chez les personnes traitées par un antagoniste de la vitamine K. / There is an increasing interest in the potential roles of vitamin K in human health aside from the one well established in blood clotting. Notably, a growing number of experimental studies show that vitamin K has biological functions in the central nervous system, particularly in the synthesis of sphingolipids, the activation of the vitamin K-dependent protein Gas6, and the prevention of oxidative damage. However, it remains uncertain whether the disruption of these functions can lead to cognitive deficits. The main objective of this thesis is to test the hypothesis that vitamin K status is a determinant of cognitive performance in healthy older adults. In order to achieve this goal, a better understanding of vitamin K status indicators was needed. Accordingly, three studies are presented each aiming at a specific objective: 1) estimate the number of non-consecutive 24-hour dietary recalls needed to assess usual vitamin K intake; 2) determine whether a single measurement of serum vitamin K concentration is an adequate indicator of long-term vitamin K exposure; and 3) examine the association between vitamin K status and cognitive performance in healthy older adults from the Québec NuAge Study. Three cognitive domains were assessed namely verbal and non-verbal episodic memory, executive functions, and speed of processing. This thesis presents the first study providing support for a role of vitamin K in cognition in older adults. Specifically, serum vitamin K concentration was positively associated with performance in verbal episodic memory, particularly in the consolidation process of the memory trace. Consistent with rodent studies and the action of the protein Gas6 in the hippocampus, a specific role of vitamin K in memory consolidation is biologically plausible. No significant association was observed with executive functions and speed of processing. Additionally, it was established that a single measurement of serum vitamin K concentration is adequate for assessing long-term vitamin K exposure in healthy older adults. Similarly, it was determined that six to 13 24-hour dietary recalls are needed to accurately assess usual vitamin K intake. Collectively, both studies provide valuable information to researchers for better interpretation of existing studies and planning future ones. Results from this thesis constitute an important step in the understanding of the potential role of vitamin K in the central nervous system, emphasizing the need to consider this micronutrient as a nutritional factor of cognitive aging, especially among those treated with vitamin K antagonists.

Background: Vitamin-K antagonists (VKA) are a class of oral anticoagulant medications used to prevent blood clots. The anticoagulant intensity of VKAs is measured with a blood test known as the International Normalized Ratio (INR). Traditionally, international guidelines have recommended INR tests every 4 weeks for all patients. However, adherence to these guidelines has never been investigated in real world settings. The objectives of this study were to describe adherence to INR testing in Saskatchewan among patients receiving VKA medications, and to identify predictors of optimal adherence. Methods: This was a retrospective cohort study of VKA users in Saskatchewan captured in the administrative data between 2003 and 2010. Physician claims for anticoagulation monitoring were used as a proxy for INR testing. Adherence to INR testing was measured using the Continuous, Multiple-Interval Measure of Medication Gaps (CMG). Individuals were considered adherent if adherence by the CMG was at least 80%. Hierarchical (random effects) logistic regression models were developed to identify important predictors of optimal INR monitoring. Individual physician identification was considered a random effect in these models. The dependent variable was the achievement of optimal adherence, defined as ≥80% adherence to the 4-week test interval. Results: Among 17,388 VKA users, 42% resided in rural areas and virtually all (99%) were monitored by a general practitioner. During a median follow-up of 514 days, 50% of patients exhibited at least 74% adherence to INR testing if a 4-week interval was used as the reference standard. However, the estimated median adherence increased dramatically to 98% when the benchmark for optimal testing was lengthened to every 12 weeks. The most prominent risk factors for poor adherence to INR monitoring appeared to be rural residence (rural vs. urban OR 0.55, 95% CI 0.47-0.64 among subjects age ≥75 years) and duration of VKA therapy (≥731 vs. 35-90 days OR 0.04, 95% CI 0.03-0.05). Discussion: Adherence to INR testing appeared to be acceptable for most VKA-treated patients in Saskatchewan. However, this data indicated that adherence might be more problematic in the subgroup of rural residents. Possible explanations include reduced access to testing facilities or the shortage of physicians in rural areas. Further research is required to understand if poor access is the underlying cause of non-adherence to INR testing in the rural population.

Thesis (Ph. D.)--University of Washington, 2005. / Vita. Includes bibliographical references (p. 149-157).