Penindolone (PND) is a novel influenza A virus dual inhibitor that blocks hemagglutinin-mediated adsorption and membrane fusion. A sensitive and specific ultra-performance liquid chromatography–tandem mass spectrometry method was developed and validated to determine PND in rat plasma. Plasma sample preparation was a simple deproteinization with acetonitrile followed by centrifugation. Chromatographic separation was performed on a C 18 column with a gradient mobile phase of acetonitrile–water containing 0.1% formic acid. Detection was carried out by electrospray ionization in negative ion multiple reaction monitoring mode. Linear detection responses were obtained for PND ranging from 1 to 1,000 ng/ml. The intra- and inter-day precision (relative standard deviations, RSD) were within 6.5%, and accuracy (relative error, RE) was within ±11.0%. The extraction recovery data for PND and internal standard (IS) were >96.0%. PND was proved to be stable during the sample storage, preparation and analytic procedures. The validated method was successfully applied to pharmacokinetic and bioavailability studies for PND in rats. The results showed the existence of twin peaks, gender difference and nonlinear pharmacokinetics for PND. In addition, two oxidation metabolites and three glucuronidation metabolites of PND were detected by ultra-high-performance liquid chromatography–high resolution mass spectrometry.
Penindolone (PND) is a novel broad-spectrum anti-Influenza A Virus (IAV) agent blocking hemagglutinin-mediated adsorption and membrane fusion. The goal of this work was to reveal the metabolic route of PND in rats. Ultra-high-performance liquid chromatography tandem high-resolution mass spectrometry (UHPLC–HRMS) was used for metabolite identification in rat bile, feces and urine after administration of PND. A total of 25 metabolites, including 9 phase I metabolites and 16 phase II metabolites, were characterized. The metabolic pathways were proposed, and metabolites were visualized via Global Natural Product Social Molecular Networking (GNPS). It was found that 65.24–80.44% of the PND presented in the formation of glucuronide conjugate products in bile, and more than 51% of prototype was excreted through feces. In in vitro metabolism of PND by rat, mouse and human liver microsomes (LMs) system, PND was discovered to be eliminated in LMs to different extents with significant species differences. The effects of chemical inhibitors of isozymes on the metabolism of PND in vitro indicated that CYP2E1/2C9/3A4 and UGT1A1/1A6/1A9 were the metabolic enzymes responsible for PND metabolism. PND metabolism in vivo could be blocked by UGTs inhibitor (ibrutinib) to a certain extent. These findings provided a basis for further research and development of PND.Keywords:penindolone;biotransformation;metabolite;liquid chromatography–mass spectrometry
Yumin Li,Lianghao Huang,Sifeng Zhu,Xiaoyao Ma,Zimin Cai,Zhao Yang,Cheng Wang,Wei Wang
Influenza A viruses (IAVs) necessitate urgently development of novel treatment approaches due to their high variability and widespread drug resistance. Penindolone (PND), distinct from anti-IAVs drugs on the market, is a promising IAVs inhibitor specifically targeting both HA1 and HA2 subunits of virus hemagglutinin. However, its clinical utility is hindered by poor water solubility and inadequate oral bioavailability. This study was for the first time to propose a practical and potent delivery approach for PND against IAVs. Inhalable PND-loaded nanoparticles (PND-NPs) were obtained through hydrophobic interactions of PND and monomethoxy poly(ethylene glycol)-poly(D, l -lactide) copolymer, realizing efficient loading of PND. The in vitro experiments demonstrated that PND-NPs exhibited lower cytotoxicity and more potent antiviral activity against IAVs than free PND (IC 50 2.5 μM vs. 42.9 μM for A/Virginia/ATCC1/2009 and 11 μM vs. 73 μM for A/Puerto Rico/8/3 (PR/8)), thus expanding the therapeutic window. In the PR/8 infected mice model, the local delivery of PND-NPs via aerosolized inhalation demonstrated that PND-NPs, superior to the effects of the clinical drug oseltamivir, effectively reduced viral load and pro-inflammatory cytokines in the lungs, and significantly prolonged the survival time of mice (twice as long as the oseltamivir treatment group). Overall, non-invasive inhalation therapy with PND-NPs holds significant promise for clinical application in the antiviral domain.
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