Rivaroxaban and other novel oral anticoagulants: pharmacokinetics in healthy subjects, specific patient populations and relevance of coagulation monitoring

Unlike traditional anticoagulants, the more recently developed agents rivaroxaban, dabigatran and apixaban target specific factors in the coagulation cascade to attenuate thrombosis. Rivaroxaban and apixaban directly inhibit Factor Xa, whereas dabigatran directly inhibits thrombin. All three drugs exhibit predictable pharmacokinetic and pharmacodynamic characteristics that allow for fixed oral doses in a variety of settings. The population pharmacokinetics of rivaroxaban, and also dabigatran, have been evaluated in a series of models using patient data from phase II and III clinical studies. These models point towards a consistent pharmacokinetic and pharmacodynamic profile, even when extreme demographic factors are taken into account, meaning that doses rarely need to be adjusted. The exception is in certain patients with renal impairment, for whom pharmacokinetic modelling provided the rationale for reduced doses as part of some regimens. Although not routinely required, the ability to measure plasma concentrations of these agents could be advantageous in emergency situations, such as overdose. Specific pharmacokinetic and pharmacodynamic characteristics must be taken into account when selecting an appropriate assay for monitoring. The anti-Factor Xa chromogenic assays now available are likely to provide the most appropriate means of determining plasma concentrations of rivaroxaban and apixaban, and specific assays for dabigatran are in development

The effect of rivaroxaban on biomarkers in blood and plasma: a review of preclinical and clinical evidence

AbstractRivaroxaban is a direct, oral factor Xa inhibitor that is used for the prevention and treatment of various thromboembolic disorders. Several preclinical and clinical studies have utilized specific molecules as biomarkers to investigate the potential role of rivaroxaban beyond its anticoagulant activity and across a range of biological processes. The aim of this review is to summarize the existing evidence regarding the use of blood-based biomarkers to characterize the effects of rivaroxaban on coagulation and other pathways, including platelet activation, inflammation and endothelial effects. After a literature search using PubMed, almost 100 preclinical and clinical studies were identified that investigated the effects of rivaroxaban using molecular biomarkers. In agreement with the preclinical data, clinical studies reported a trend for reduction in the blood concentrations of D-dimers, thrombin–antithrombin complex and prothrombin fragment 1 + 2 following treatment with rivaroxaban in both healthy individuals and those with various chronic conditions. Preclinical and also some clinical studies have also reported a potential impact of rivaroxaban on the concentrations of platelet activation biomarkers (von Willebrand factor, P-selectin and thrombomodulin), endothelial activation biomarkers (matrix metalloproteinase-9, intercellular adhesion molecule-1 and vascular cell adhesion molecule-1) and inflammation biomarkers (interleukin-6, tumor necrosis factor-α and monocyte chemoattractant protein-1). Based on the results of biomarker studies, molecular biomarkers can be used in addition to traditional coagulation assays to increase the understanding of the anticoagulation effects of rivaroxaban. Moreover, there is preliminary evidence to suggest that rivaroxaban may have an impact on the biological pathways of platelet activation, endothelial activation and inflammation; however, owing to paucity of clinical data to investigate the trends reported in preclinical studies, further investigation is required to clarify these observations.</jats:p

A High-Sensitivity, Medium-Density, and Target Amplification–Free Planar Waveguide Microarray System for Gene Expression Analysis of Formalin-Fixed and Paraffin-Embedded Tissue

Abstract Background: Many microarray platforms and their associated assay chemistries do not work properly with RNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissue samples, a feature that severely hampers the use of microarrays in oncology applications, for which FFPE tissue is the routine specimen. Furthermore, the limited sensitivity of most microarray platforms requires time-consuming and costly amplification reactions of the target RNA, which negatively affects clinical laboratory work flow. Methods: We developed an approach for sensitively and reliably measuring mRNA abundances in FFPE tissue samples. This approach involves automated RNA extractions, direct hybridization of extracted RNA to immobilized capture probes, antibody-mediated labeling, and readout with an instrument applying the principle of planar waveguides (PWG). A 14-gene multiplex assay conducted with RNA isolated from 20 FFPE blocks was correlated to an analysis of the same with reverse-transcription quantitative real-time PCR (RT-qPCR). Results: The assay sensitivity for gene expression analysis obtained for the PWG microarray platform was &amp;lt;10 fmol/L, eliminating the need for target preamplification. We observed a correlation coefficient of 0.87 to state-of-the-art RT-qPCR technology with RNA isolated from FFPE tissue, despite a compressed dynamic range for the PWG system (a 2.9-log dynamic range for PWG in our test system vs 5.0 logs for RT-qPCR). The precision of the PWG platform was comparable to RT-qPCR (Pearson correlation coefficient of 0.9851 for PWG vs 0.9896 for RT-qPCR) for technical replicates. Conclusions: The presented PWG platform demonstrated excellent sensitivity and precision and is especially well suited for any application for which fast, simple, and robust multiplex assays of RNA in FFPE tissue are required.</jats:p

Receptor Editing in a Transgenic Mouse Model: Site, Efficiency, and Role in B Cell Tolerance and Antibody Diversification

AbstractMice carrying transgenic rearranged V region genes in their IgH and Igκ loci to encode an autoreactive specificity direct the emerging autoreactive progenitors into a pre-B cell compartment, in which their receptors are edited by secondary Vκ-Jκ rearrangements and RS recombination. Editing is an efficient process, because the mutant mice generate normal numbers of B cells. In a similar nonautoreactive transgenic strain, neither a pre-B cell compartment nor receptor editing was seen. Thus, the pre-B cell compartment may have evolved to edit the receptors of autoreactive cells and later been generally exploited for efficient antibody diversification through the invention of the pre-B cell receptor, mimicking an autoreactive antibody to direct the bulk of the progenitors into that compartment

Pharmacodynamics, Pharmacokinetics and Safety of Bay 1093884, an Antibody Directed Against Human TFPI, in Patients with Factor VIII or IX Deficiency (With and Without Inhibitors): A Phase 1 Study

Abstract Background: BAY 1093884 is a fully human monoclonal antibody directed against the K1 and K2 domains of human tissue factor pathway inhibitor (TFPI). TFPI regulates the initiation of coagulation and is important for regulation of normal hemostatic response. In factor (F)VIII and FIX deficiency (hemophilia A [HA] and B [HB], respectively), lack of adequate amplification (FXa generation) in tandem with intact regulation contributes to reduced clotting. Inhibition of TFPI has the potential to normalize coagulation in hemophilia even in the absence of FVIII or FIX and it is expected to result in normalization of thrombin generation and consequently improvement in clot formation. TFPI is an attractive therapeutic target as it is unaffected by FVIII or FIX inhibitors and can be targeted in both HA and HB. A potential advantage of targeting TFPI is that downstream mechanisms that protect from excessive coagulation (including antithrombin), remain intact. Methods: BAY 1093884 was evaluated in a multicenter, open-label study in patients with severe HA or HB, with or without inhibitors. Safety, pharmacokinetics (PK) and pharmacodynamics (PD) were assessed after single intravenous (IV) (0.3 and 1 mg/kg) and subcutaneous (SC) (1, 3, and 6 mg/kg) doses. A multiple-dose (MD) cohort received BAY 1093884 150 mg SC once weekly for 6 weeks. Total and free TFPI protein levels and TFPI activity were measured. Thrombin generation assay (TGA) parameters were measured using a calibrated automated thrombogram. Clot formation was assessed using rotational thromboelastometry (ROTEM). A mechanistic physiology-based PK/PD (PBPK/PD) model was developed based on single-dose PK data observed in the study, and used to determine dosing in the MD cohort. Data for HA, HB, Japanese and inhibitor patient subgroups were analyzed separately. Results: Thirty-two patients were enrolled; 30 with HA and 2 with HB, of which 4 (3 HA, 1 HB) were included in the MD cohort. At least 4 patients were treated in each administration method/dose group. Clot formation using ROTEM was assessed in 6 patients. Total and free TFPI-related parameters showed dose-dependent effects, with the concentrations being below the lower limit of quantification for longer duration at higher doses. Improvements in TGA parameters were observed following both IV and SC administration; duration of improvement was longer at higher doses. BAY 1093884 led to shortening of the clotting time in all 6 patients tested. Dose-dependent increases in plasma concentration exceeding dose proportionality were seen following IV and SC administration, consistent with expectations of target-mediated drug disposition for BAY 1093884. Moderate accumulation was observed after MD. Limited information on subgroups (patients with inhibitors [n=4], patients with HB [n=2] and Japanese origin [n=1]) suggests that the PK and PD data for these subgroups were consistent with those from non-inhibitor HA patients. The PBPK/PD model adequately described the concentration time course and total TFPI following single-dose IV and SC administration of BAY 1093884; the PK and PD data from the MD cohort were as predicted by the model. The model also supported that the PK/PD of BAY 1093884 are independent of inhibitor status. No serious adverse events, deaths, or adverse events leading to discontinuation were observed. Fibrinogen and platelet counts remained within normal range at most visits and no meaningful changes in levels of antithrombin, protein C, or FV were seen. The mechanisms for anticoagulation and fibrinolysis were intact. Conclusions: Overall, BAY 1093884 gave no safety concerns in the dose range evaluated. Results suggest that BAY 1093884 shifts hemostatic balance towards greater thrombin generation and clot formation as intended. PK/PD characteristics were similar for inhibitor and HB patients as compared with HA patients, potentially supporting use in all of these patient types. The PBPK/PD model developed can be used for dose selection in future studies. Treatment efficacy will be evaluated in a future study. Disclosures Chowdary: Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Swedish Orphan Biovitrum AB (Sobi): Honoraria; Baxalta (Shire): Honoraria, Membership on an entity's Board of Directors or advisory committees; Novo Nordisk: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Freeline: Consultancy; CSL Behring: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Biogen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bayer: Honoraria. Lissitchkov:Novo Nordisk: Other: Investigator fees as a participant of the clinical trial. Willmann:Bayer: Employment. Schwers:Bayer: Employment. Michaels:Bayer: Employment. Shah:Bayer: Employment. </jats:sec

B Cell Development under the Condition of Allelic Inclusion

AbstractMice whose IgH alleles are engineered to encode two distinct antibody heavy (H) chains generate a normal-sized B cell compartment in which most cells stably express the two heavy chains. This demonstrates that “toxicity” of bi-allelic H chain expression and cell- autonomous mechanisms of silencing in-frame IgH gene rearrangements do not significantly contribute to allelic exclusion at the IgH locus. Notwithstanding, the stability of the various engineered IgH loci during B cell development in the bone marrow differed substantially from each other