Development of a sensitive LC-MS/MS assay to support human microdose study for an oral agonist of the GLP-1 receptor.

The purpose of this work was to determine the feasibility of supporting a clinical microdose study for PF-06882961 (danuglipron), an oral small molecule agonist of the GLP-1 receptor, by LC-MS/MS. Statistical instrument parameter optimization using response surface methodology was employed to develop a LC-MS/MS method for the analyte, PF-06882961. An LC-MS/MS method was developed and validated to support a proof of concept microdose pharmacokinetics preclinical study in monkeys, administered PF-06882961 (0.

Inception and development of a LC-MS/MS assay for the multiplexed quantitation of nine human drug transporter biomarkers.

It has become common practice to assess solute carrier transporter (SLC)-mediated drug-drug interactions (DDIs) by quantitating various individual endogenous compounds as biomarkers in human plasma and urine. The goal of this work was to develop biomarker multiplex assays that could be utilized during first in human studies to support the simultaneous assessment of clinical DDI risk across various SLCs. Hydrophilic interaction chromatography-MS/MS methods were developed, and validations were performed.

LC-MS/MS method for the quantitation of OCT1 biomarker isobutyrylcarnitine in human plasma with clinical sample analysis.

Isobutyrylcarnitine (IBC) is a possible biomarker for hepatic OCT1, as IBC plasma concentrations are reduced when OCT1 is inhibited. An accessible, characterized assay is needed to quantitate IBC in human plasma. A triple quadrupole MS surrogate matrix assay for the quantitation of IBC was characterized to support a first-in-human study.

Development and implementation of urinary transporter biomarkers to facilitate assessment of drug-drug interaction.

Novel urinary biomarker evaluation approaches to support inhibition assessment for renal transporters (e.g., OCT2, multidrug and toxin extrusion proteins [MATEs]).

A Multiplexed HILIC-MS/HRMS Assay for the Assessment of Transporter Inhibition Biomarkers in Phase I Clinical Trials: Isobutyryl-Carnitine as an Organic Cation Transporter (OCT1) Biomarker.

There is a growing interest in using endogenous compounds as drug transporter biomarkers to facilitate drug-drug interaction (DDI) risk assessment in early phase I clinical trials. Compared to other drug transporters, however, no valid biomarker for hepatic organic cation transporter (OCT) 1 has been described to date. The present work represents the first report of an endogenous compound, isobutyryl-l-carnitine (IBC), as a potential clinical OCT1 biomarker for DDI assessment.

Evaluation of OptiFlow™-MS/MS for bioanalysis of pharmaceutical drugs and metabolites.

Microflow tandem mass spectrometry-based methods have been proposed as options to improve sensitivity and selectivity while improving sample utility and solvent consumption. Here, we evaluate a newly introduced microflow source, OptiFlow™, for quantitative performance. We performed a comparison of the OptiFlow and IonDrive™ sources, respectively, on the same triple quadrupole mass spectrometer.

Bioanalysis of Targeted Nanoparticles in Monkey Plasma via LC-MS/MS.

This work represents the first reporting of a comprehensive bioanalytical GLP methodology detailing the mass spectrometric quantitation of PF-05212384 dosed as a targeted polymeric encapsulated nanoparticle (PF-07034663) to monkeys. Polymeric nanoparticles are a type of drug formulation that enables the sustained release of an active therapeutic agent (payload) for targeted delivery to specific sites of action such as cancer cells. Through the careful design and engineering of the nanoparticle formulation, it is possible to improve the biodistribution and safety of a given therapeutic payload in circulation.

LC-MS/MS assay for N-methylnicotinamide in humans, an endogenous probe for renal transporters.

Background: N-methylnicotinamide (1-NMN) has been proposed as a potential clinical biomarker to assess drug-drug interactions involving organic cation transporters (OCT2) and multidrug and toxin extrusion protein transporters.

Results: A hydrophilic interaction liquid chromatography-MS/MS assay, to quantify 1-NMN, in human plasma and urine is reported.

Materials & Methods: A hydrophilic interaction chromatography (HILIC)-tandem mass spectrometry (MS/MS) assay to quantify 1-NMN in human plasma and urine is reported.

A fully automated and validated human plasma LC-MS/MS assay for endogenous OATP biomarkers coproporphyrin-I and coproporphyrin-III.

Aim: A validated LC-MS/MS assay for the quantitation of coproporphyrin-I and -III (CP-I, CP-III) in human plasma has been developed to understand the utility of both as possible endogenous biomarkers for organic anion-transporting polypeptides (OATP)-mediated drug-drug interactions (DDIs).

Materials And Methods:  Human plasma extracts were analyzed for CP-I and CP-III using a Sciex API 6500+ mass spectrometer. Results: The assay was utilized for plasma samples from a clinical DDI study involving a new chemical entity that presented as an OATP inhibitor in vitro.

A sensitive method for the quantitation of the peptide-based glucagon-like peptide-1 receptor agonist liraglutide in plasma using microfluidics chromatography tandem MS.

Aim: An LC-MS/MS assay for the quantitation of liraglutide, a peptide-based injectable glucagon-like peptide-1 receptor agonist, has been developed as a convenient alternative to the enzyme-linked immunosorbent assay, and used to characterize liraglutide pharmacokinetics in cynomolgus monkeys.

Results: Assay calibration curves exhibited a linear dynamic range of 10-5000 ng/ml and correlation coefficient ≥0.98.

A highly selective and sensitive LC-MS/HRMS assay for quantifying coproporphyrins as organic anion-transporting peptide biomarkers.

Aim: Coproporphyrin-I (CP-I) and coproporphyrin-III (CP-III) in plasma and urine have been proposed as biomarkers for assessing drug-drug interactions involving hepatic drug transporters such as organic anion-transporting peptides (OATP), 1B1 and 1B3. Materials & methods: Plasma and urine extracts were analyzed for CP-I/CP-III using a TripleTOF API6600 mass spectrometer. Results: Previously unreported, CP-I/CP-III doubly charged ions (m/z 328.

Small molecule specific run acceptance, specific assay operation, and chromatographic run quality assessment: recommendation for best practices and harmonization from the global bioanalysis consortium harmonization teams.

Consensus practices and regulatory guidance for liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) assays of small molecules are more aligned globally than for any of the other bioanalytical techniques addressed by the Global Bioanalysis Consortium. The three Global Bioanalysis Consortium Harmonization Teams provide recommendations and best practices for areas not yet addressed fully by guidances and consensus for small molecule bioanalysis. Recommendations from all three teams are combined in this report for chromatographic run quality, validation, and sample analysis run acceptance.

Utilization of hydrophilic-interaction LC to minimize matrix effects caused by phospholipids.

Background: In bioanalysis, phospholipids may affect the precision and accuracy of LC-MS/MS methods and compromise the quality of the results, especially when samples in complex biomatrices are extracted by protein precipitation techniques.

Results: It was found that the retentive behavior of both common pharmaceuticals and physiologically relevant phospholipids under bare silica hydrophilic-interaction LC (HILIC) is more predictable than under reversed-phase conditions. In particular, the retention time of phospholipids was not significantly affected by varying the salt and acid modifiers in the mobile phases, but common pharmaceuticals can be shifted away from these phospholipid interferences through mobile phase modifiers.

Application of the dried spot sampling technique for rat cerebrospinal fluid sample collection and analysis.

Dried blood spotting (DBS) sample collection is gaining favor in the pharmaceutical industry due to benefits that include reduced animal usage and easier sample shipment and storage when compared to traditional plasma collection/analysis. The applicability of the DBS card to alternate, limited-volume, matrices has not been as fully characterized as their use with whole blood. In this paper we explored the application of the DBS sample collection technique to rat cerebrospinal fluid (CSF).

Development and validation of a direct enantiomeric separation of pregabalin to support isolated perfused rat kidney studies.

Pregabalin (Lyrica) is the first compound approved to treat the neural pain associated with fibromyalgia. Pregabalin is the S-enantiomer of a gamma-amino acid analogue and chiral separation from its R-enantiomer must be achieved to support metabolic studies. The direct chiral separation of pregabalin from its R-enantiomer has been developed and HPLC/MS/MS assays have been validated to support isolated perfused rat kidney studies.

Toxicity and toxicokinetics of the cyclin-dependent kinase inhibitor AG-024322 in cynomolgus monkeys following intravenous infusion.

Purpose: Cyclin-dependent kinases (CDKs) play a significant role in the control of cell-cycle progression and exhibit aberrant regulation in various neoplastic diseases. AG-024322 is a potent inhibitor of CDK1, CDK2, and CDK4 that produces cell-cycle arrest and antitumor activity in preclinical models. This study evaluated the toxicity of AG-024322 when given by intravenous (IV) infusion to cynomolgus monkeys, including reversibility of effects.