Performance characteristics of the first Food and Drug Administration (FDA)-cleared digital droplet PCR (ddPCR) assay for BCR::ABL1 monitoring in chronic myelogenous leukemia.

Chronic myelogenous leukemia (CML) is a hematopoietic stem cell malignancy that accounts for 15-20% of all cases of leukemia. CML is caused by a translocation between chromosomes 9 and 22 which creates an abnormal fusion gene, BCR::ABL1. The amount of BCR::ABL1 transcript RNA is a marker of disease progression and the effectiveness of tyrosine kinase inhibitor (TKI) treatment.

Higher Adalimumab Levels Are Associated with Histologic and Endoscopic Remission in Patients with Crohn's Disease and Ulcerative Colitis.

Background: Optimal levels of adalimumab (ADA) have not been defined according to the ultimate goal of inflammatory bowel disease treatment--histologic and/or endoscopic healing. The aim of this study was to assess the relationship between random serum ADA levels and histologic and endoscopic healing in patients with inflammatory bowel disease.

Methods: This was a cross-sectional study including 66 patients receiving maintenance therapy with ADA for Crohn's disease or ulcerative colitis.

Antibodies to adalimumab are associated with future inflammation in Crohn's patients receiving maintenance adalimumab therapy: a post hoc analysis of the Karmiris trial.

Introduction: Data on immunogenicity to adalimumab (ADL) therapy in patients with IBD is limited. We performed additional analyses on the Karmiris cohort using the homogeneous mobility shift assay (HMSA) focusing on the inter-relationship of serum ADL concentration, antibodies-to-adalimumab (ATA), inflammatory markers and sustained response.

Methods: 536 prospectively collected serum samples were available for analysis of ADL concentration and ATA using HMSA.

Antibodies to infliximab are associated with lower infliximab levels and increased likelihood of surgery in pediatric IBD.

Background: Adult studies suggest antibodies to infliximab (ATI) correlate with loss of response in inflammatory bowel disease but pediatric data are limited.

Methods: We conducted a cross-sectional study of trough infliximab levels and ATI in 134 pediatric and young adult patients receiving infliximab. At the time serum was obtained demographics, disease phenotype, duration of infliximab therapy, use of combination therapy (methotrexate or 6-mercaptopurine with infliximab), and surgery were recorded.

Concentrations of 6-thioguanine nucleotide correlate with trough levels of infliximab in patients with inflammatory bowel disease on combination therapy.

Background & Aims: In patients with inflammatory bowel diseases, the combination of infliximab and thiopurines (such as 6-thioguanine) is more effective treatment than monotherapy. We assessed the correlation between serum levels of 6-thioguanine (6-TGN) and infliximab levels or antibodies to infliximab (ATI).

Methods: We performed a cross-sectional study of 72 patients receiving maintenance therapy with infliximab and a thiopurine for inflammatory bowel disease at the Crohn's and Colitis Center of the University of Miami, FL.

Long-term outcome of patients with Crohn's disease who discontinued infliximab therapy upon clinical remission.

Background & Aims: There are limited data on the effects of discontinuing infliximab therapy for Crohn's disease (CD). We investigated the long-term outcome of patients with CD who discontinued infliximab while in clinical remission, and searched for prognostic markers of continued remission after infliximab cessation.

Methods: We performed a retrospective, single-center study of 100 patients with CD who discontinued infliximab upon achieving clinical remission; 84 patients continued immunomodulator therapy.

The relationship between infliximab concentrations, antibodies to infliximab and disease activity in Crohn's disease.

Objective: Although low infliximab trough concentrations and antibodies to infliximab (ATI) are associated with poor outcomes in patients with Crohn's disease (CD), the clinical relevance of ATI in patients with adequate infliximab concentrations is uncertain. We evaluated this question using an assay sensitive for identification of ATI in the presence of infliximab.

Design: In an observational study, 1487 trough serum samples from 483 patients with CD who participated in four clinical studies of maintenance infliximab therapy were analysed using a fluid phase mobility shift assay.

Early trough levels and antibodies to infliximab predict safety and success of reinitiation of infliximab therapy.

Background & Aims: Few agents are available for the treatment of inflammatory bowel diseases, and patients frequently become unresponsive to biologics. We investigated the feasibility of reinitiating infliximab therapy for patients who previously received only episodic therapy with, lost response to, or had infusion reactions to infliximab. We also aimed to identify factors associated with the success and safety of restarting infliximab, such as antibodies to infliximab and trough levels of the drug.

Monitoring of adalimumab and antibodies-to-adalimumab levels in patient serum by the homogeneous mobility shift assay.

This report describes the analytical validation and application of the homogeneous mobility shift assay (HMSA) method for the measurement of adalimumab and human antibodies-to-adalimumab (ATA) in serum samples from patients who have lost response to adalimumab treatment. Validation of the ATA- and the adalimumab-HMSA revealed a lower limit of detection to be 0.026 U/mL for ATA and 0.

Antibody response to infliximab and its impact on pharmacokinetics can be transient.

Objectives: Infliximab (IFX) is successfully used in the treatment of inflammatory bowel diseases but some patients generate antibodies to IFX (ATI) jeopardizing the pharmacokinetics of the drug. Little is known about the factors influencing ATI formation and whether or not this immune reaction is permanent. Our aim was to investigate the kinetics of ATI formation and drug levels in relation to inflammatory markers and the clinical evolution of the patients.

Development and validation of a homogeneous mobility shift assay for the measurement of infliximab and antibodies-to-infliximab levels in patient serum.

Antibody-based drugs such as infliximab (IFX) are effective for the treatment of inflammatory bowel disease (IBD) and other immune-mediated disorders. The development of antibodies against these drugs may result in unfavorable consequences, including the loss of drug efficacy, hypersensitivity reactions, and other adverse events. Therefore, accurate monitoring of serum drug and anti-drug antibody levels should be an important part of therapy for patients being treated with an antibody-based drug.

The homotetrameric phosphoseryl-tRNA synthetase from Methanosarcina mazei exhibits half-of-the-sites activity.

Synthesis of cysteinyl-tRNA(Cys) in methanogenic archaea proceeds by a two-step pathway in which tRNA(Cys) is first aminoacylated with phosphoserine by phosphoseryl-tRNA synthetase (SepRS). Characterization of SepRS from the mesophile Methanosarcina mazei by gel filtration and nondenaturing mass spectrometry shows that the native enzyme exists as an alpha4 tetramer when expressed at high levels in Escherichia coli. However, active site titrations monitored by ATP/PP(i) burst kinetics, together with analysis of tRNA binding stoichiometry by fluorescence spectroscopy, show that the tetrameric enzyme binds two tRNAs and that only two of the four chemically equivalent subunits catalyze formation of phosphoseryl adenylate.

Redundant synthesis of cysteinyl-tRNACys in Methanosarcina mazei.

A subset of methanogenic archaea synthesize the cysteinyl-tRNA(Cys) (Cys-tRNA(Cys)) needed for protein synthesis using both a canonical cysteinyl-tRNA synthetase (CysRS) as well as a set of two enzymes that operate via a separate indirect pathway. In the indirect route, phosphoseryl-tRNA(Cys) (Sep-tRNA(Cys)) is first synthesized by phosphoseryl-tRNA synthetase (SepRS), and this misacylated intermediate is then converted to Cys-tRNA(Cys) by Sep-tRNA:Cys-tRNA synthase (SepCysS) via a pyridoxal phosphate-dependent mechanism. Here, we explore the function of all three enzymes in the mesophilic methanogen Methanosarcina mazei.

Kinetic quality control of anticodon recognition by a eukaryotic aminoacyl-tRNA synthetase.

Aminoacyl-tRNA synthetases are an ancient class of enzymes responsible for the matching of amino acids with anticodon sequences of tRNAs. Eukaryotic tRNA synthetases are often larger than their bacterial counterparts, and several mammalian enzymes use the additional domains to facilitate assembly into a multi-synthetase complex. Human cysteinyl-tRNA synthetase (CysRS) does not associate with the multi-synthetase complex, yet contains a eukaryotic-specific C-terminal extension that follows the tRNA anticodon-binding domain.

Shape-selective RNA recognition by cysteinyl-tRNA synthetase.

The crystal structure of Escherichia coli cysteinyl-tRNA synthetase (CysRS) bound to tRNA(Cys) at a resolution of 2.3 A reveals base-specific and shape-selective interactions across an extensive protein-RNA recognition interface. The complex contains a mixed alpha/beta C-terminal domain, which is disordered in the unliganded enzyme.