Eco-Friendly, Simple, Fast, and Sensitive UPLC-MS/MS Method for Determination of Pexidartinib in Plasma and Its Application to Metabolic Stability

Pexidartinib is the first drug approved by the U.S. Food and Drug Administration specifically to treat the rare joint tumor tenosynovial giant cell tumor. In the current study, a validated, selective, and sensitive UPLC-MS/MS assay was developed for the quantitative determination of pexidartinib in plasma samples using gifitinib as an internal standard (IS). Pexidartinib and IS were extracted by liquid-liquid extraction using methyl tert-butyl ether and separated on an acquity BEH C18 column kept at 40 °C using a mobile phase of 0.1% formic acid in acetonitrile: 0.1% formic acid in de-ionized water (70:30). The flow rate was 0.25 mL/min. Multiple reaction monitoring (MRM) was operated in electrospray (ESI)-positive mode at the ion transition of 418.06 > 165.0 for the analyte and 447.09 > 128.0 for the IS. FDA guidance for bioanalytical method validation was followed in method validation. The linearity of the established UPLC-MS/MS assay ranged from 0.5 to 1000 ng/mL with r > 0.999 with a limit of quantitation of 0.5 ng/mL. Moreover, the metabolic stability of pexidartinib in liver microsomes was estimated.

Design, Synthesis, and Antidepressant Activity Study of Novel Aryl Piperazines Targeting Both 5-HT1A and Sigma-1 Receptors

A total of 20 novel aryl piperazine derivatives were designed and synthesized, and their structures were confirmed by mass spectrometry and nuclear magnetic resonance analyses. Their 5-HT1A and sigma-1 receptor affinities were determined, and six of them showed high affinities (K i < 20 nmol/L) to both 5-HT1A and sigma-1 targets. Then, metabolic stability (T 1/2) tests of six compounds in rat and human liver microsomes were performed. Our data indicated that compound 27 has both high affinity for 5-HT1A and sigma-1 receptors (5-HT1A: K i = 0.44 nmol/L; sigma-1: K i = 0.27 nmol/L), and good metabolic stability (T 1/2 values are 21.7 and 24.6 minutes, respectively). Interestingly, results from the forced swimming test, mouse tail suspension test, and preliminary pharmacokinetic test suggested the marked antidepressant activity, good pharmacokinetic characteristics, and low toxicity of compound 27 in the two models. In conclusion, compound 27 has great value of further study as an active molecule of antidepressant drugs.

In vitro Metabolic Stability of Drugs and Applications of LC-MS in Metabolite Profiling

Metabolic stability of a compound is an important factor to be considered during the early stages of drug discovery. If the compound has poor metabolic stability, it never becomes a drug even though it has promising pharmacological characteristics. For example, a drug is quickly metabolized in the body; it does not have sufficient in vivo exposure levels and leads to the production of toxic, non-active or active metabolites. A drug is slowly metabolized in the body it could remain longer periods in the body and lead to unwanted adverse reactions, toxicity or may cause drug interactions. Metabolic stability assay is performed to understand the susceptibility of the compound to undergo biotransformation in the body. Intrinsic clearance of the compound is measured by metabolic stability assays. Different in vitro test systems including liver microsomes, hepatocytes, S9 fractions, cytosol, recombinant expressed enzymes, and cell lines are used to investigate the metabolic stability of drugs. Metabolite profiling is a vital part of the drug discovery process and LC–MS plays a vital role. The development of high-resolution (HR) MS technologies with improved mass accuracy, in conjunction with novel data processing techniques, has significantly improved the metabolite detection and identification process. HR-MS based data acquisition (ion intensity-dependent acquisition, accurate-mass inclusion list-dependent acquisition, isotope pattern-dependent acquisition, pseudo neutral loss-dependent acquisition, and mass defect-dependent acquisition) and data mining techniques (extracted ion chromatogram, product ion filter, mass defect filter, isotope pattern filter, neutral loss filter, background subtraction, and control sample comparison) facilitate the drug metabolite identification process.

The Use of Nanomedicine to Target Signaling by the PAK Kinases for Disease Treatment

P21-activated kinases (PAKs) are serine/threonine kinases involved in the regulation of cell survival, proliferation, inhibition of apoptosis, and the regulation of cell morphology. Some members of the PAK family are highly expressed in several types of cancer, and they have also been implicated in several other medical disorders. They are thus considered to be good targets for treatment of cancer and other diseases. Although there are several inhibitors of the PAKs, the utility of some of these inhibitors is reduced for several reasons, including limited metabolic stability. One way to overcome this problem is the use of nanoparticles, which have the potential to increase drug delivery. The overall goals of this review are to describe the roles for PAK kinases in cell signaling and disease, and to describe how the use of nanomedicine is a promising new method for administering PAK inhibitors for the purpose of disease treatment and research. We discuss some of the basic mechanisms behind nanomedicine technology, and we then describe how these techniques are being used to package and deliver PAK inhibitors.

Tackling the opioid crisis - development of kappa-opioid receptor peptide agonists for safer analgesic therapy

The kappa opioid receptor (KOPr) has exceptional potential as an analgesic target, seemingly devoid of the many peripheral side-effects of Mu receptors. Kappa-selective, small molecule pharmaceutical agents have been developed, but centrally mediated side effects have the limited their clinical translation. Here, we modify an active endogenous Dynorphin peptide with the aim of improving drug-likeness and developing safer KOPr agonists for clinical use. Using rational, iterative design and modern peptide chemistry, we developed a series of potent, selective and metabolically stable peptides from Dynorphin 1-7. Peptides were assessed for cAMP-modulation against Kappa, Mu and Delta opioid receptors, metabolic stability, KOPr specificity and binding, and interrogated for in vitro desensitisation and pERK signalling capability. Finally, lead peptides were evaluated for efficacy in Freund’s complete adjuvant rat model of inflammatory nociception. A library of 70 peptides was synthesised and assessed for pharmacological and metabolic stability factors. At least 10 peptide candidates showed low nanomolar activity (˂50 nM) in a cAMP assay, specificity for KORr, and plasma half-life >60 min, with 6 candidates also stable in trypsin. None of the selected peptides showed pERK activity, with a bias towards cAMP signalling. In vivo, KA305 and KA311 showed anti-nociception opioid receptor-specific activity comparable to morphine and U50 844. These highly potent and metabolically stable peptides are promising opioid analgesic leads for clinical translation. Since they are biased peptide KOPr agonists, it is plausible they lack many of the most significant side effects, such as tolerance, addiction, sedation and euphoria/dysphoria, common to opioid analgesics.

The Metabolic Stability of Antimicrobial Peptides IK8 in Plasma and Liver S9

In the face of mounting global antibiotic resistance, which has become a critical clinical problem, antimicrobial peptides (AMPs) have received considerable interest as new therapeutics with the efficacy for the treatment of multidrug-resistant (MDR) infections due to their novel mechanism. However, certain inherent shortcomings such as instability seriously limit their systemic applications in the clinic. In this study, we intend to clarify the connection between three configurations of IK8 and their stability in plasma and liver S9 of various species by confirming the metabolites. The structural information of these metabolites was scanned and identified using HPLC and Q-TOF, respectively. The results found that IK8-D exhibits superior stability, compared with IK8-2D and IK8-L in plasma and liver S9 incubation, which indicated D- type amino acids could significantly increase the stability of antimicrobial peptides.

Radiolabeled Bombesin Analogs

The gastrin-releasing peptide receptor (GRPR) is expressed in high numbers in a variety of human tumors, including the frequently occurring prostate and breast cancers, and therefore provides the rationale for directing diagnostic or therapeutic radionuclides on cancer lesions after administration of anti-GRPR peptide analogs. This concept has been initially explored with analogs of the frog 14-peptide bombesin, suitably modified at the N-terminus with a number of radiometal chelates. Radiotracers that were selected for clinical testing revealed inherent problems associated with these GRPR agonists, related to low metabolic stability, unfavorable abdominal accumulation, and adverse effects. A shift toward GRPR antagonists soon followed, with safer analogs becoming available, whereby, metabolic stability and background clearance issues were gradually improved. Clinical testing of three main major antagonist types led to promising outcomes, but at the same time brought to light several limitations of this concept, partly related to the variation of GRPR expression levels across cancer types, stages, previous treatments, and other factors. Currently, these parameters are being rigorously addressed by cell biologists, chemists, nuclear medicine physicians, and other discipline practitioners in a common effort to make available more effective and safe state-of-the-art molecular tools to combat GRPR-positive tumors. In the present review, we present the background, current status, and future perspectives of this endeavor.