vitamin K-dependent proteins

Introduction: Growth arrest-specific gene 6 protein (GAS6) and protein S (ProS) are vitamin K-dependent proteins present in plasma with important regulatory functions in systems of response and repair to damage. They interact with receptor tyrosine kinases of the Tyro3, Axl and MerTK receptor tyrosine kinase (TAM) family, involved in apoptotic cell clearance (efferocytosis) and regulation of the innate immunity. TAM-deficient mice show spontaneous lupus-like symptoms. Here we tested the genetic profile and plasma levels of components of the system in patients with systemic lupus erythematosus (SLE), and compare them with a control healthy population. Methods: Fifty SLE patients and 50 healthy controls with matched age, gender and from the same geographic area were compared. Genetic analysis was performed in GAS6 and the TAM receptor genes on SNPs previously identified. The concentrations of GAS6, total and free ProS, and the soluble forms of the three TAM receptors (sAxl, sMerTK and sTyro3) were measured in plasma from these samples.

Low vitamin K status (indicated by undercarboxylated MGP) is associated with increased vascular calci fi cations, and these levels can be improved by effective vitamin K supplementation 28–32 It was long believed that vitamin K was only involved in forming coagulation factors (ie, maintaining haemostasis). However, other vitamin-K dependent proteins (containing γ -carboxyglutamate or Gla) are dependent on vitamin-K carboxylation for func- tionality. 33 Vitamin K acts as a cofactor in the conversion of glutamate into Gla. Gla-containing proteins (MGP and osteocalcin) regulate many anticalci fi cation and bone-forming processes in the body, which are depend- ent on vitamin K in order to be produced. Low levels of vitamin K impair activation of osteocalcin and decrease the activity of osteoblasts (cells important for building bone). 33 34 Thus, vitamin K is vital to the functionality of proteins such as osteocalcin (important for building bone), (MGP, the most potent arterial calci fi cation inhibitor known) and the growth-arrest sequence-6 protein (Gas6, involved in cell growth regulation 35

The mechanism of warfarin helps us see clearly how it inhibits further coagulation of blood but it also shows its interference in the mechanism of vitamin K. Vitamin K antagonists, also known as oralanticoagulants (OACs), are widely used for the treatment and prophylaxis of thromboebolic diseases. Short- term OAC treatment is applied often after deep venous thrombosis, while atrialfibrillation or after prosthetic heart valve implantation require long term treatment (Block, 2001). The below Figure 1 indicated; Vitamin K is decarboxylated in the process and needs to be recycled. The enzyme Vitamin K- epoxide reductase (VKORC) is essential in this cycle. It is this re-carboxylation by VKORC that is inhibited by Warfarin. Carboxylation of glutamate residues to γ-carboxyglutamates (Gla) on the N-terminal regions of vitamin K–dependent proteins requires vitamin K as a cofactor (Whitlon et al., 1978). Formation of coagulation factors II, VII, IX, and X is hence blocked by blocking this process. When the vitamin K conversion cycle is inhibited, warfarin will influence hepatic production of partially decarboxylated proteins with greatly decreased coagulant activity (Friedman et al., 1977; Malhotra et al., 1985). Carboxylation promotes the binding of phospholipid surfaces to the vitamin K–dependent coagulation factors, thereby accelerating blood coagulation (Nelsestuen, 1976).

VKD carboxylation has been associated mostly with coagulation, because it was originally observed in clotting factors (Nelsestuen, Zytkovicz, & Howard, 1974; Stenflo, Fernlund, Egan, & Roepstorff, 1974). All the VKD clotting factors require the 9–13 Glu residues at the N-terminus of the protein to be fully carboxylated in order for the protein to be functional. Defects of VKD carboxylation have long been known to cause bleeding disorders, known as combined vitamin K-dependent coagulation factors deficiency (VKCFD; Napolitano, Mariani, & Lapecorella, 2010; Prentice, 1985). With the discovery of new Gla proteins, the importance of VKD carboxylation has been extended beyond coagulation into a number of other physiological functions. For example, carboxylated matrix Gla protein (MGP) is a strong inhibitor of vascular calcification and of connective tissue mineralization, and uncarboxylated MGP has been implicated in cardiovascular diseases and other nonbleeding syndromes (Willems, Vermeer, Reutelingsperger, & Schurgers, 2014). Another VKD protein, osteocalcin (also called bone Gla protein, or BGP), is produced by osteoblasts and is important for bone formation (Ducy et al., 1996); recent studies suggest that osteocalcin also functions as a hormone affecting glucose metabolism (Mera et al., 2016). Therefore, functional study of the vitamin K cycle enzymes is not only essential for understanding blood coagulation, but it is also important for understanding many other physiological processes.

the duration of action of a single dose of racemic warfarin is 2 to 5 days. Under normal pharmacological therapy the drugs are administered to decrease the action of the clotting factors they affect by 30 to 50% 3 . Warfarin exerts its anticoagulant and antithrombotic activity by inhibiting the vitamin K-dependent carboxylation of clotting factors II, VII, IX, and X to a greater extent than the vitamin K-dependent natural anticoagulants, proteins C and S 8, 9 . Specifically, it inhibits an enzyme called the vitamin K 1 2, 3-epoxide

atherosclerotic plaque. Calcified valves including specimens with calcific aortic stenosis, calcified porcine xenograft valves, and a calcified aortic homograft valve were analyzed for Gla content, complete amino acid analysis, and tissue calcium and phosphorus levels. Normal porcine valves contained protein-bound Gla (2.0-10.6 Gla/10 4 amino acids): no Gla was present in normal valve leaflets. Furthermore, Gla levels paralleled tissue calcium content in the calcified valves. In addition, complete amino acid analysis indicated a decline in valvar collagen content plus increased acidic proteins in conjunction with valvar

The marked homology of the mammalian carboxylases sug- gests the importance of vitamin K and the synthetic machinery to generate ␥ -carboxyglutamic acid. The evolutionary conser- vation of this complex enzymatic system requiring exogenous vitamin K or a vitamin K-like cofactor and multiple enzymes for vitamin K transport and metabolism suggests a critical role for ␥ -carboxyglutamic acid that has been retained throughout phylogeny. ␥ -Carboxyglutamic acid is important for calcium binding and plays a critical functional role in generating the membrane binding properties of the vitamin K-dependent blood clotting proteins (24). The role of ␥ -carboxyglutamic acid in other mammalian proteins such as osteocalcin and matrix Gla protein remains uncertain. The role of ␥ -carboxyglutamic acid in the conotoxins also remains speculative, despite the determination of the structures of some of these peptides (21, 22, 29). We suspect that ␥ -carboxyglutamic acid plays a broader role than the highly specialized activities associated with mem- brane interaction. Perhaps, ␥ -carboxyglutamic acid is critical for function of certain housekeeping proteins that are ubiqui- tous in animal cells, although it would appear that vitamin K-dependent carboxylation is not a required step until late in embryogenesis. 1 Isolation of novel ␥ -carboxyglutamic acid-con-

Vitamin K–dependent (VKD) proteins undergo γ-glutamyl carboxylation characterized by the conversion of specific glutamic acid (Glu) residues to γ-carboxylated Glu (Gla) (reviewed in ref. 1). Carboxylation is essential for the functions of VKD proteins, which include coagulation factors like prothrombin (PT) and factors VII, IX, and X that are produced in the liver. VKD proteins are modified in the ER by the γ-glutamyl carboxylase (GGCX), which requires the vitamin K hydroquinone (KH 2 ) cofactor that is oxygenated to vitamin K 2,3-epoxide (KO) during carboxylation. KO is then recycled by VKORC1, a vitamin K oxidore- ductase (VKOR) (2, 3) that is evolutionarily conserved (4). VKORC1 recycles KO to KH 2 in 2 steps, wherein KO is first converted to vitamin K quinone (K) and then to KH 2 (5). Together, the enzymatic activities of GGCX and VKORC1 form the vitamin K cycle (1). Warfarin, an anticoagulant used by millions of people, suppresses VKD protein carboxylation by inhibiting VKORC1 (3, 6).

As with all other coagulation factors, FII, FVII, FIX, FX and the three anticoagulant proteins (PC, PS and PZ) are normally synthesized in the liver. All such factors undergo a post-translational modification of glutamate (Glu) residues into γ-carboxyglutamate (Gla) residues. Carboxylation, necessary for their normal activity in the coagulation system, involves Glu residues located in a homologous 'Gla domain' which spans approximately 45 amino acids. Gla residues enable these proteins to adapt to calcium-dependent conformation allowing their bind- ing to phospholipids [20]. The γ-carboxylation is held in the endoplasmic reticulum and is catalyzed by the enzyme GGCX which is hypothesized to bind to the pro- peptide coagulation factors at their amino terminus. Car- boxylated proteins are then transported to the Golgi for secretion and the propeptide sequence is then removed [21,22]. Vitamin K is an essential cofactor for GGCX and when a carbon dioxide is added to Glu to form Gla, the reduced form of vitamin K (vitamin K hydroquinone) is oxygenated to form vitamin K 2,3 epoxide. At this stage the enzyme VKORC is needed to regenerate the vitamin K hydroquinone, completing the so called vitamin K cycle [21] (Figure 1). Three forms of vitamin K are known: i)vitamin K1 (Phylloquinone) which is abundant in green and leafy vegetables and is produced by plants and algae; ii)vitamin K2 (Menaquinones) which is a mixture of mol- ecules produced by the microbial intestinal flora and dif- fers from K1 by unsaturated side chains of isoprenoid units with different length; iii)vitamin K3 (Menadione) which is a synthetic form and is more water-soluble.

Because anticoagulant therapy is useful for recurrent thrombosis and because this decreases antigenic levels of vitamin K-dependent plasma proteins 21-23, we used the ratio of protein C [r]

Neonatal RDSs are characterized by leakage of plasma proteins of varying sizes into the airspace, which leads to interstitial and intra-alveolar thrombin generation with subsequent fibrin deposition (5), systemic activation of clotting, complement and polymorph nuclear lymphocytes (14) . PTZ, which is a vitamin k dependent protein, proved to play a role in the prevention of coagulation (6)

We found several patient and treatment characteristics that were associated with the risk of underanticoagulation. A possible explanation for the difference in frequency of occurrence of subtherapeutic anticoagulation amongst the indication categories may be a difference in compliance: The risk was highest in patients who used anticoagulation as primary prophylaxis for venous thromboembolism and lowest in patients with mechanical heart valves, who have the highest underlying risk of thrombosis. The increased risk for patients with an arterial indication disappeared completely after adjustment for target range, indicating that the latter is the real association. One possible explanation for the higher risk of a subtherapeutic INR in high therapeutic range patients is that dosing physicians are more inclined to lower the dose when the INR is high in range in these patients than in the low therapeutic range patients. The most striking difference was between phenprocoumon and acenocoumarol. Fifty percent of patients using acenocoumarol had a subtherapeutic INR after 13 weeks compared to 51 weeks in patients using phenprocoumon. This finding is consistent with reports that longer-acting vitamin K antagonist give a more stable anticoagulation than short-acting vitamin K antagonists. 23-25

Pharmacovigilance practices were implemented at our center (Hospital de Clínicas de Porto Alegre, state of Rio Grande do Sul, Brazil) in 2002 and have since been carried out by the Clinical Pharmacy Service. The hospital’s Drug Information Center is in charge of all such activities, and conducts drug utilization studies and active case-finding of adverse drug reactions (ADRs) with the primary objective of fostering rational use of medicines. The present descriptive study, conducted from June through August 2012, included patients who were prescribed intravenous, intramuscular, or subcutaneous vitamin K. Patients as were neonates (due to routine prophylactic administration of vitamin K) [3]. Patients were identified by means of daily reports of patient prescriptions, issued by the hospital electronic medical records (EMR) system, and by means of active case-finding, which consisted of a daily review of the EMR and contact with staff physicians. The identified patients were followed until discontinuation of vitamin K therapy or until hospital discharge so as to assess the profile of vitamin K prescription at our center. Vitamin K was selected as a trigger for identification of adverse events (AEs) associated with excessive use or insufficient monitoring of therapy with oral anticoagulants, such as warfarin [4,5]. All AEs thus identified were treated as ADRs, as all therapeutic doses were individualized. Patients on oral anticoagulant therapy were monitored by means of the prothrombin time (PT), as expressed by the International Normalized Ratio (INR) (reference range 2–3); INR values >4 were flagged due to risk of bleeding and/or hemorrhage. The Naranjo et al. algorithm was used to determine the probability of a causal relationship between the prescribed medications and the suspected ADR, which was classified as definite, probable, possible, or doubtful [6]. ADRs were further classified by predictability as predictable or unpredictable, as per Thompson & Rawlins [7]; and by severity

normalize coagulation within one hour (62.2 versus 9.6%; P <0.05) [8]. The role of vitamin K in association with PCC, in maintaining the normalized coagulation over six hours has been emphasized [9]. The importance of rapid hemorrhage control has led international guidelines to recommend the use of PCC rather than FFP [10-12]. For complete VKA reversal, all guidelines recommend an infusion of PCC (at least 20 IU/kg factor IX equivalent) related to admission INR value in combination with at least 5 mg of vitamin K to rapidly achieve a post-reversal INR ≤1.5 and maintain a normal coagulation profile over six hours. So as to save time, French guidelines suggest, on the basis of a previous study, the possibility of adminis- tering a probabilistic single-regimen dose of 25 IU/kg of PCC and 10 mg of vitamin K as soon as a severe hemorrhage is diagnosed [9,10]. In all situations, post- reversal INR must be measured 30 minutes after the infusion to evaluate the efficacy of the treatment and make any necessary adjustments thereafter, and measured six hours later to control the efficacy of vitamin K [10-12]. French guidelines were published in 2008 and have become a standard of care for the management of these patients in France [10]. Treatment time frames were not specified in the international guidelines and the effect of early reversal on mortality has not been studied. At a time when new oral anticoagulant agents without specific antidotes are emerging as promising alternatives to VKAs, it appears important to better define the prognosis benefit of anti- coagulant reversal in the management of patients on oral anticoagulants with severe hemorrhage [13].

Recent evidence suggests that non-vitamin K oral anticoagulants (NOACs) are as effective as vitamin K antagonists (VKAs) such as warfarin in the prevention of stroke and systemic embolism in patients with AF with a lower risk of intracranial hemorrhage. As com- pared to warfarin, dabigatran was associated with reduced risk of ischemic stroke and systemic embo- lism as well as intracranial hemorrhage, but with a higher rate of gastrointestinal hemorrhage. 10 Apixa-

Fat-soluble vitamins include biologically active members of vitamin A (diterpenes), vitamin D (modified triterpenes), vitamin E (tocochromanols), and vitamin K (naphthoquinones) families, and related compounds, of which some act as their precursors. Vitamin A and vitamin D and the corresponding provitamins (provitamins A and provitamins D) are biosynthesised as products of the mevalonate and deoxyxylulose phosphate pathways leading to terpenoids and steroids. The biologically active compounds derived from qui- nones with phytyl or isoprenoid side-chains include members of the vitamin E and K families as well as structurally related plastoquinones and ubiqui- nones. Vitamin E, vitamin K, plastoquinones, and ubiquinones principally arise as products of the shikimate pathway that further produces aromatic

We evaluated whether a vitamin K prophylactic regimen of 1 mg vitamin K orally at birth followed by 150 μg daily during weeks 2 to 13 sufficiently prevented VKDB in breastfed infants. Our data in a high- risk group, ie, undiagnosed children with BA, show that this regimen does not successfully prevent VKDB in these children, in contrast to a regimen consisting of a single IM injection of 2 mg vitamin K at birth. This study shows that increasing the daily dose of the oral vitamin K prophylactic regimen from 25 to 150 μg fails to prevent VKDB in breastfed infants with yet undiag- nosed BA. VKDB occurred in 82% of the infants and included several cases of intracranial hemorrhage, which has been associated with serious morbidity and high mortality. 5 The

patient-driven end of the spectrum, the physician presents factual information and available options, leaving parents to decide. This would be appropriate when there is more than one option with comparable effectiveness, not the case in our vitamin K scenario. At the physician- driven end of the spectrum, care is provided without discussion, as in life-threatening situations. Our vitamin K scenario falls somewhere in the middle, which leads to the next question: When a family declines the recommended care, how far can a clinician ethically go to promote a low-risk option with clear health benefit?

Chairman Sanford: None of the mothers of the premature infants born in the hospital were given vitamin K.. Almost all the mothers of those born at home received vitamin K before delivery[r]

Vitamin K, and fresh frozen plasma returned the prothrombin time and partial thromboplastin time to normal values within 18 hours, suggesting that the infant ad severe vitamin K deficien[r]