Highlights of the 12th Japan Bioanalysis Forum Symposium.

Approximately 300 people associated with pharmaceutical industries, contractors, academic institutions and regulatory authorities attended the 12th Japan Bioanalysis Forum Symposium. The webinar was conducted from 9 to 11 March 2021. The theme of the symposium was 'for the next generation', and the event provided 'an opportunity for young researchers in bioanalysis (including students)' and 'an opportunity to discuss new frontiers of bioanalysis'. The speakers focused on hot topics of bioanalysis, including biomarker analysis, patient centric sampling, virtual clinical trials, gene therapy, cancer genome medicine and therapeutic middle molecules. The symposium presented a platform for the discussion of the prospects and challenges facing bioanalysts working in the field of pharmacokinetics. This report presents the key issues discussed.

Analytical method validation for biomarkers as a drug development tool: points to consider.

Biomarkers are an important drug developmental tool. Assessment of quantitative analytical methods of biomarkers is not included in any regulatory documents in Japan. Use of biomarkers in clinical evaluations and supporting the post-marketing evaluation of drug efficacy and/or adverse reactions requires assessment and full validation of analytical methods for these biomarkers. The Biomarker Analytical Method Validation Study Group is a research group in Japan comprising industry and regulatory experts. Group members discussed and prepared this 'points to consider document' covering measurements of endogenous metabolites/peptides/proteins by ligand binding assays and chromatographic methods with or without mass spectrometry. We hope this document contributes to the global harmonization of biomarker assay validation.

Comparison of the Pharmacokinetics of RIPK1 Inhibitor GSK2982772 in Healthy Western and Japanese Subjects.

BACKGROUND AND OBJECTIVES: GSK2982772 is an oral small-molecule RIPK1 inhibitor with potential therapeutic efficacy in immune-mediated inflammatory diseases (IMIDs). An inter-ethnic comparison of GSK2982772 pharmacokinetics was conducted based on data from Western (Study 1) and Japanese subjects (Study 2). METHODS: Both studies were single-centre, randomised, double-blind, placebo-controlled studies with objectives to assess the safety and characterise the pharmacokinetics of GSK2982772. Western subjects in Study 1 (NCT03305419), Part A (N = 15), were randomly assigned to receive 120 mg three times daily (TID), 240 mg TID, or 360 mg twice daily (BID) doses of GSK2982772, or placebo (TID or BID) for 1 day. Part B subjects (N = 47) received GSK2982772 120 mg TID, 240 mg TID, or placebo TID for 14 days. Japanese subjects in Study 2 (N = 13) (NCT03590613) were randomly assigned to receive TID doses of GSK2982772 60, 120, 240 mg TID or placebo TID for 1 day. RESULTS: GSK2982772 was well tolerated and adverse events were generally mild. Maximum observed plasma drug concentration (Cmax), time to reach Cmax (Tmax), area under the plasma drug concentration versus time curve after the first GSK2982772 dose (AUC(0-7)) of 120 and 240 mg, and (AUC(0-24)) values for the 120 and 240 mg TID doses over a single day were similar in Japanese and Western subjects. CONCLUSIONS: The pharmacokinetics and tolerability of GSK2982772 were similar between Western and Japanese subjects, justifying inclusion of Japanese subjects in future global clinical studies to assess the therapeutic potential of RIPK1 inhibition for the treatment of IMIDs. Clinical Trials: NCT03305419 and NCT03590613 available from http://www.clinicaltrials.gov .

Evaluation of the Pharmacokinetics, Safety, and Tolerability of a Single Oral Dose of Pyrimethamine in Healthy Male Subjects of Japanese and European Ancestry.

The pharmacokinetics of pyrimethamine have been evaluated in various populations but have not been reported in subjects of Japanese ancestry following administration as a single-agent tablet. Furthermore, although pyrimethamine pharmacokinetics after a single dose of the single-agent tablet studied in Western countries have been reported, these studies are old, and the ancestry of the subjects was not specified. Consequently, this study investigated the pharmacokinetics and safety of a single oral 50-mg dose of pyrimethamine in healthy male subjects of Japanese and European ancestry. Seven subjects of each ancestry group were administered pyrimethamine, along with calcium folinate. After absorption, pyrimethamine was eliminated, with a mean half-life of 122.8 hours in Japanese subjects and 99.5 hours in European subjects. The mean Cmax and AUC0-t were 433.8 ng/mL and 59.63 µg·h/mL in Japanese subjects and 372.7 ng/mL and 42.83 µg·h/mL in European subjects. No safety concerns were reported during the study. Although pyrimethamine exposure was slightly higher in subjects of Japanese than of European ancestry, a considerable overlap in the range of parameter values was observed. Considering the range of pyrimethamine exposure reported previously, difference in exposure observed in this study would not be considered of note.

The 10th Japan Bioanalysis Forum Symposium conference: a report.

The Japan Bioanalysis Forum Symposium was held on 12-14 February 2019 (Yokohama, Japan), in celebration of its 10th anniversary, and over 370 participants from pharmaceutical industries, contractors, academia and regulatory authorities from home and abroad came together in Yokohama. The 3-day symposium particularly aimed to foster collaboration with the scientists surrounding bioanalysts, according to the theme 'Open to the Public.' The symposium also included a broad range of pioneering programs, such as lectures by speakers from DMPK/metabolomics fields, discussions of future bioanalysis and poster presentations by publicly offered presenters as well as the regular ones we had organized. This report summarizes the major topics as a conference report.

Evaluation of the Safety, Tolerability, and Pharmacokinetics of GSK2269557 (Nemiralisib) Administered Via Dry Powder Inhaler to Healthy Japanese Subjects.

The aim of the study was to assess the safety, tolerability, and pharmacokinetics of single and repeat doses of nemiralisib administered via a dry powder inhaler to healthy Japanese subjects. This was a single-center, double-blind, randomized, placebo-controlled, parallel, single- and repeat-ascending-dose study. Thirty-six healthy Japanese male subjects were randomized to receive either 1 dose strength of nemiralisib or placebo. The study consisted of a screening period, a single-dose session (session 1), a repeat-dose session (session 2), a 10-day washout period between the sessions, and then a follow-up visit 10 ± 1 days after the last dose of session 2. No serious adverse events were reported. No clinically significant abnormalities were found in clinical laboratory results, vital signs, or spirometry results. Generally, exposure (maximum observed plasma concentration [Cmax ] and area under the concentration-time curve [AUC]) increased with dose in an approximately proportional manner. Plasma Tmax was achieved rapidly at approximately 0.08 hours, and the terminal elimination half-life (T1/2 ) was approximately 40 hours. Tmax and T1/2 did not change between days or doses in the single- and repeat-dose sessions. Following 10 daily doses of 200, 500, and 700 µg nemiralisib, accumulation was observed, and the ratios (session 2, day 10:session 1) for Ro(AUC0-24 ) and R(Cmax ) were 2.4-3.0 and 1.5-1.7, respectively. Steady state was achieved by 6-7 days, based on trough observed plasma drug concentration (Ctrough ) values.

Evaluation of the Safety, Tolerability, Pharmacokinetics, and Pharmacodynamics of a Single Intravenous Dose of Miridesap in Healthy Japanese Subjects.

This phase 1 study characterized the safety, tolerability, pharmacokinetics, and pharmacodynamics of miridesap (GSK2315698) following an intravenous (IV) infusion in healthy Japanese men. Subjects in Cohort 1 received 1-hour IV infusions of 10, 20, and 40 mg of miridesap or placebo, and subjects in Cohort 2 received a 15-hour IV infusion of 20 mg/h of miridesap or placebo. No treatment-related adverse events were reported. No new safety signals were identified for either vital signs or clinical laboratory parameters. A dose-dependent increase was observed in miridesap exposure (area under the concentration-time curve and maximum observed drug concentration) in the 10 to 40 mg/h dose range after a 1-hour IV infusion of miridesap. Rapid depletion of circulating serum amyloid P component was observed after the initiation of miridesap infusion. Serum amyloid P component concentrations fell in a dose-dependent manner following administration of miridesap.

The ninth Japan Bioanalysis Forum symposium.

The ninth Japan Bioanalysis Forum symposium took place at tower hall Funabori, Tokyo, Japan, between 6 and 8 February, 2018. Bioanalytical scientists from the pharmaceutical industry, CROs, academia and regulatory bodies had many meaningful and relevant discussions on current topics of interest in bioanalysis. The 3-day symposium featured updated perspectives and experiences on regulated bioanalysis of small and large molecules, biomarker measurement and assessment of immunogenicity, as well as new areas of bioanalytical validation such as quantitative polymerase chain reaction(qPCR) and flow cytometry. There were over 260 participants from six countries, with 23 oral and 11 poster presentations, including the outcomes of Japan Bioanalysis Forum discussion groups. This report summarizes the major discussion topics from the conference.

Sample Management: Recommendation for Best Practices and Harmonization from the Global Bioanalysis Consortium Harmonization Team.

The importance of appropriate sample management in regulated bioanalysis is undeniable for clinical and non-clinical study support due to the fact that if the samples are compromised at any stage prior to analysis, the study results may be affected. Health authority regulations do not contain specific guidance on sample management; therefore, as part of the Global Bioanalysis Consortium (GBC), the A5 team was established to discuss sample management requirements and to put forward recommendations. The recommendations from the team concern the entire life span of the sample and include the following: 1. Sampling procedures should be described in the protocol or within the laboratory manual. This information should include the volume of the sample to be collected, the required anticoagulant, light sensitivity, collection and storage containers, and labeling with a unique identifier. 2. The correct procedures for processing and then storing the samples after collection at the clinical/non-clinical testing site and during shipment are also very important to ensure the analyte(s) stability and should be documented. 3. Chain of custody for the samples must be maintained throughout the complete life span of each sample. This is typically maintained via paper and electronic data systems, including Laboratory Information Management Systems (LIMS) where available. 4. Pre- and post-analysis storage location and conditions must also be clearly defined at the analytical laboratory. The storage temperature of the samples must be traceable and controlled by monitoring and warning alerts. The team suggests moving away from using temperatures and to adopt standard terminology of "room temperature," "refrigerator," "freezer," and "ultra-freezer" that have defined and industry-wide accepted temperature ranges. 5. At the end of the study, documentation of the samples' disposal is required.

6th Japan Bioanalysis Forum Symposium: challenge of regulated bioanalysis.

Biomarker measurement and LC-MS analysis of large molecule pharmaceuticals (LM-MS) are becoming increasingly important in pharmaceutical development, although they are not included in the existing bioanalytical method validation guidelines in Japan. The 6th Japan Bioanalysis Forum symposium presented challenges regarding biomarkers and LM-MS, as well as the current circumstances for regulated bioanalysis, by inviting speakers from Japan, the USA, European Union and Asia-pacific. Japan Bioanalysis Forum discussion groups also presented their outcomes and openly discussed these with the attendees - over 200 dedicated individuals from industry, regulatory agencies and academia. The symposium successfully reinforced the idea that fit-for-purpose approaches are necessary, and that science should drive any judgments and actions throughout drug development.

Safety, tolerability, pharmacokinetics, and pharmacodynamics of losmapimod in healthy Japanese volunteers.

This phase 1 study characterized the safety, tolerability, pharmacokinetics, and pharmacodynamics of losmapimod and its metabolite GSK198602 following single and repeat doses of oral losmapimod in healthy Japanese volunteers. Subjects (n = 41) received single oral doses of losmapimod (2.5, 7.5, 20 mg) or matching placebo on 3 separate days (n = 20) or losmapimod 7.5 mg or matching placebo twice daily for 14 days (n = 21). Assessments included maximum observed plasma concentration (Cmax ), time to Cmax (Tmax ), apparent terminal-phase half-life (t1/ )2 , area under the curve (AUC), and change in C-reactive protein and phosphorylated heat shock protein 27 levels. No serious adverse events occurred during the study, and there were no safety concerns regarding clinical laboratory parameters, 12-lead electrocardiogram, or vital signs. The losmapimod Tmax was 3-4 hours, and the mean t1/2 was approximately 7.9-9.0 hours, with no appreciable difference in Tmax and apparent clearance following oral dosing between dosing regimens. Single and repeat oral doses of losmapimod were well tolerated in healthy Japanese volunteers. The Tmax of GSK198602 was similar to and t1/2 was slightly longer than those of losmapimod. Approximate dose-proportional increases in exposure to losmapimod and GSK198602 were observed in AUC with single-dose administration. Repeat-dose trough concentrations reached steady state within 2 days, with an observed accumulation ratio of 1.56 and 1.91 for losmapimod and GSK198602, respectively.

A highly sensitive LC-MS/MS method capable of simultaneously quantitating celiprolol and atenolol in human plasma for a cassette cold-microdosing study.

A highly sensitive simultaneous quantitative method for a cassette cold-microdosing study on celiprolol and atenolol was developed with liquid chromatography-tandem mass spectrometry. The method utilizes a combination of solid-phase extraction (SPE) with strong cation exchange (SCX) cartridge columns and reversed-phase chromatography with an ODS analytical column. SCX-SPE cartridge columns (100 mg sorbent) were used for a selective extraction of celiprolol, atenolol and metoprolol (internal standard) from 500 µL of human plasma samples. Turbo-ion spray at positive mode was employed for the ionization of the drug compounds. Quantitation was performed on a triple quadrupole mass spectrometer by selected reaction monitoring with the transitions of m/z 380 to m/z 251 for celiprolol and m/z 267 to m/z 145 for atenolol. Separation of analytes was achieved on an ODS column (100 mm length × 2.1 mm id, 3 µm) by a gradient elution with 10 mM formic acid and methanol by varying their proportion at a flow rate of 0.2 mL/min. The method was validated in the range of 1-250 pg/mL for celiprolol and 2.5-250 pg/mL for atenolol and was successfully applied to the elucidation of pharmacokinetic profiling in a cold cassette microdosing study of the ß-blockers.

Importance of serum concentration of adefovir for Lamivudine-adefovir combination therapy in patients with lamivudine-resistant chronic hepatitis B.

Lamivudine (LMV)-adefovir pivoxil (ADV) combination therapy suppresses the replication of LMV-resistant hepatitis B virus (HBV), although its efficacy in suppressing HBV varies among patients. This study analyzed the clinical, virological, and pharmaceutical factors that influence the effect of the combination therapy. Patients negative for hepatitis B virus e antigen (HBeAg) and with low HBV DNA titers immediately prior to the combination therapy effectively cleared serum HBV DNA (P=0.0348 and P=0.0310, respectively). The maximum concentration of ADV in serum (ADV Cmax) was higher in patients who showed HBV DNA clearance (P=0.0392), and the cumulative clearance rates of HBV DNA were significantly higher in patients with ADV Cmax equal to or greater than 24 ng/ml (P=0.0284). HBeAg negativity and lower HBV DNA at the start of the combination therapy and higher ADV Cmax were found to be independent factors for serum HBV DNA clearance. Serum creatinine increased significantly during the combination therapy, and the ADV Cmax was higher in patients with low creatinine clearance rates. In conclusion, higher serum concentrations of ADV are associated with a good response to therapy based on clearance of HBV DNA in serum. However, care should be taken to prevent worsening of renal function due to high ADV serum concentrations.