Explore the vast range of titles available.

Immunoaffinity–mass spectrometry (IA-MS) is an emerging analytical genre with several advantages for profiling and determination of protein biomarkers. Because IA-MS combines affinity capture, analogous to ligand binding assays (LBAs), with mass spectrometry (MS) detection, this platform is often described using the term hybrid methods. The purpose of this report is to provide an overview of the principles of IA-MS and to demonstrate, through application, the unique power and potential of this technology. By combining target immunoaffinity enrichment with the use of stable isotope-labeled internal standards and MS detection, IA-MS achieves high sensitivity while providing unparalleled specificity for the quantification of protein biomarkers in fluids and tissues. In recent years, significant uptake of IA-MS has occurred in the pharmaceutical industry, particularly in the early stages of clinical development, enabling biomarker measurement previously considered unattainable. By comparison, IA-MS adoption by CLIA laboratories has occurred more slowly. Current barriers to IA-MS use and opportunities for expanded adoption are discussed. The path forward involves identifying applications for which IA-MS is the best option compared with LBA or MS technologies alone. IA-MS will continue to benefit from advances in reagent generation, more sensitive and higher throughput MS technologies, and continued growth in use by the broader analytical community. Collectively, the pursuit of these opportunities will secure expanded long-term use of IA-MS for clinical applications.

In the SELECT trial, disease activity was reduced in patients with multiple sclerosis who received daclizumab high-yield process (HYP) for 52 weeks. The primary aim of the SELECTION extension study was to assess the safety and immunogenicity of extended treatment with daclizumab HYP. A multicentre, randomised, double-blind, 52-week extension trial was done in 74 centres in the Czech Republic, Germany, Hungary, India, Poland, Russia, Ukraine, and the UK between Feb 13, 2009, and Oct 3, 2012. Eligible patients were aged 18–55 years, had relapsing-remitting multiple sclerosis, and had completed the SELECT study. Patients who received placebo in SELECT were randomly assigned (1:1) to receive 150 mg or 300 mg subcutaneous daclizumab HYP every 4 weeks for 52 weeks (treatment initiation group); those who had received daclizumab HYP were randomly assigned (1:1) to continue their present dose with (washout and re-initiation group) or without (continuous treatment group) a washout period of 20 weeks. All randomisation was done with a centralised, interactive voice-response system. Patients and personnel were masked to treatment assignment, except for the site pharmacist who prepared the study drug but had no interaction with patients. The primary endpoints were the safety and immunogenicity of daclizumab HYP. Analyses were by intention to treat. This study is registered with ClinicalTrials.gov, number NCT00870740. 517 (91%) of 567 patients who completed the SELECT trial entered SELECTION, of whom 170 were in the treatment initiation group, 173 in the continuous treatment group, and 174 in the washout and re-initiation group. 11 patients in the treatment initiation group (6%), 13 in the continuous treatment group (8%), and ten in the washout and re-initiation group (6%) had any serious adverse event other than relapse of multiple sclerosis. One patient in the washout and re-initiation group (300 mg daclizumab HYP) died because of autoimmune hepatitis; a contributory role of daclizumab HYP could not be excluded. Seven patients tested positive for neutralising antidrug antibodies: one (1%) of 128 for whom data were available in the continuous treatment group (this patient also tested positive at SELECTION baseline), four (2%) in the treatment initiation group, and two (2%) of 129 in the washout and re-initiation group. Adverse events and immunogenicity were not increased in the second year of continuous treatment with daclizumab HYP or during treatment washout and re-initiation. These results support further assessment of daclizumab HYP for relapsing-remitting multiple sclerosis. Biogen Idec and AbbVie Biotherapeutics.

Persistent upregulation of signaling by cytokine tumor necrosis factor–like weak inducer of apoptosis (TWEAK) through its receptor fibroblast growth factor–inducible molecule-14 (Fn14) promotes chronic inflammation and tissue destruction. The aim of this study was to explore the safety and tolerability of the TWEAK-blocking monoclonal antibody BIIB023 and determine its pharmacokinetics and effects on TWEAK pathway pharmacodynamic markers in rheumatoid arthritis (RA). Phase I, first-in-human, 2-part, multicenter, double-blind, dose-escalation study. Patients were randomized to a single dose of BIIB023 (0.03–20 mg/kg) (n = 38) or placebo (n = 15) as an add-on to methotrexate. Three open-label cohorts of RA patients taking background disease-modifying antirheumatic drugs and stable tumor necrosis factor (TNF) inhibitor therapy (n = 12) received a single-dose of BIIB023 of 2, 10, or 20 mg/kg and were assessed over 70 days. The incidence of treatment-emergent adverse events for the BIIB023 monotherapy cohorts and open-label cohorts of BIIB023 as add-on therapy to TNF inhibitors compared with placebo were 47% and 50% versus 33%, respectively. Serum exposure to BIIB023 increased in a dose-dependent manner from 0.03 to 20 mg/kg, but not in direct proportion to dose level. After administration, the time course of BIIB023 serum concentration was multiphasic and showed expedited elimination when levels decreased to < 10 µg/mL. Serum-soluble TWEAK levels were suppressed at all dose levels by 6 hours post-dose and recovered to baseline between days 7 and 28. A trend toward downward modulation of serum biomarkers of inflammatory response was suggested in monocyte chemoattractant protein 1, inducible protein 10, macrophage inflammatory protein 1β, and tissue inhibitor of metalloproteinase 1 in the BIIB023 group versus placebo. Single-dose BIIB023 showed a favorable safety and tolerability profile in RA. Suppression of serum-soluble TWEAK for ≤ 28 days was observed and downward trends in serum biomarkers suggested.

This conference report summarized a full-day workshop, “best practices for the development and fit-for-purpose validation of biomarker methods,” which was held prior to the American Association of Pharmaceutical Scientists (AAPS) PharmSci360 Congress, San Antonio, TX, November 2019. The purpose of the workshop was to bring together thought leaders in biomarker assay development in order to identify which assay parameters and key statistical measures need to be considered when developing a biomarker assay. A diverse group of more than 40 scientists participated in the workshop. The workshop and subsequent working dinner stimulated robust discussion. While a consensus on best practices was not achieved, some common themes and major points to consider for biomarker assay development have been identified and agreed on. The focus of this conference report is to summarize the presentations and discussions which occurred at the workshop. Biomarker assay validation is a complex and an evolving area with discussions ongoing.

The 2019 13 Workshop on Recent Issues in Bioanalysis (WRIB) took place in New Orleans, LA, USA on April 1–5, 2019 with an attendance of over 1000 representatives from pharmaceutical/biopharmaceutical companies, biotechnology companies, contract research organizations and regulatory agencies worldwide. WRIB was once again a 5-day, week-long event – a full immersion week of bioanalysis, biomarkers, immunogenicity and gene therapy. As usual, it was specifically designed to facilitate sharing, reviewing, discussing and agreeing on approaches to address the most current issues of interest including both small- and large-molecule bioanalysis involving LCMS, hybrid LBA/LCMS, LBA cell-based/flow cytometry assays and qPCR approaches. This 2019 White Paper encompasses recommendations emerging from the extensive discussions held during the workshop and is aimed to provide the bioanalytical community with key information and practical solutions on topics and issues addressed, in an effort to enable advances in scientific excellence, improved quality and better regulatory compliance. Due to its length, the 2019 edition of this comprehensive White Paper has been divided into three parts for editorial reasons. This publication (Part 3) covers New Insights in Biomarker Assay Validation, Current & Effective Strategies for Critical Reagent Management, Flow Cytometry Validation in Drug Discovery & Development & CLSI H62, Interpretation of the 2019 FDA Immunogenicity Guidance and Gene Therapy Bioanalytical Challenges. Part 1 (Innovation in Small Molecules and Oligonucleotides & Mass Spectrometry Method Development Strategies for Large Molecule Bioanalysis) and Part 2 (Recommendations on the 2018 FDA BMV Guidance, 2019 ICH M10 BMV Draft Guideline and regulatory agencies' input on bioanalysis, biomarkers, immunogenicity and gene therapy) are published in volume 11 of , issues 22 and 23 (2019), respectively.

Myostatin (MSTN) is an attractive therapeutic target for the treatment of muscle degeneration-related diseases and is being evaluated as a target engagement biomarker. A sensitive 2D-LC–MS/MS assay was developed to quantify MSTN in different animal species. Sample preparation involved SDS denaturation of serum proteins followed by tryptic digestion and peptide enrichment by SPE. The assay was validated with LLOQ of 2.5 ng/ml in rat and monkey serum. The precision was within 13.7%, and the bias was within ±12.6% for all quality control samples in authentic matrices. This new assay was successfully applied to measure MSTN in mouse, rat, monkey and human serum. The total MSTN in rat and monkey serum was elevated following administration of an MSTN inhibitor.

Develop LC-MS/MS-based assays to measure total and free complement C5 in cynomolgus monkey serum as a target engagement biomarker for pharmacokinetic/pharmacodynamic correlation study. The C5-specific signature peptide derived from pellet digestion of serum proteins with and without prior immunodepletion of the drug-bound C5 by protein A beads was quantified to assess free and total C5 levels, respectively. Conditions for immunodepletion by protein A were optimized to ensure complete depletion of IgGs (and drug-bound C5). The effect of sample dilution on drug-target dissociation and thus free C5 measurement was evaluated by applying a mathematical simulation. The procedure described here allows for the assessment of protein target engagement, aiding in pharmacokinetic/pharmacodynamic correlation analysis and human dose projection.

Immunologic responses to anti-PD-1 therapy in melanoma patients occur rapidly with pharmacodynamic T cell responses detectable in blood by 3 weeks. It is unclear, however, whether these early blood-based observations translate to the tumor microenvironment. We conducted a study of neoadjuvant/adjuvant anti-PD-1 therapy in stage III/IV melanoma. We hypothesized that immune reinvigoration in the tumor would be detectable at 3 weeks and that this response would correlate with disease-free survival. We identified a rapid and potent anti-tumor response, with 8 of 27 patients experiencing a complete or major pathological response after a single dose of anti-PD-1, all of whom remain disease free. These rapid pathologic and clinical responses were associated with accumulation of exhausted CD8 T cells in the tumor at 3 weeks, with reinvigoration in the blood observed as early as 1 week. Transcriptional analysis demonstrated a pretreatment immune signature (neoadjuvant response signature) that was associated with clinical benefit. In contrast, patients with disease recurrence displayed mechanisms of resistance including immune suppression, mutational escape, and/or tumor evolution. Neoadjuvant anti-PD-1 treatment is effective in high-risk resectable stage III/IV melanoma. Pathological response and immunological analyses after a single neoadjuvant dose can be used to predict clinical outcome and to dissect underlying mechanisms in checkpoint blockade. Neoadjuvant PD-1 blockade in patients with resectable melanoma followed by adjuvant maintenance results in early immunological effects driving clinical benefit and reveals transcriptional and genomic mechanisms of response.