Lack of pro-arrhythmic potential for the novel soluble guanylate cyclase stimulator vericiguat: results from preclinical studies

Introduction Vericiguat is an orally-administered soluble guanylate cyclase stimulator, developed for the treatment of symptomatic chronic heart failure (HF) in adult patients who have had a previous decompensation event. At the maximum therapeutic dose of 10 mg once daily in patients with HF, the protein-unbound plasma concentrations of vericiguat and its major pharmacologically inactive N-glucuronide metabolite M-1 are approximately 18 nmol/l and 43 nmol/l, respectively. As part of an integrated risk assessment, vericiguat and its M-1 metabolite were characterised electrophysiologically in vivo and in vitro. This was performed according to the International Council for Harmonisation standard S7B guideline and to recent related “best practice” revisions (draft ICH E14/S7B Q&A), being adopted as a result of the Comprehensive In Vitro Proarrhythmia Assay (CIPA) initiative. Purpose To assess the potential for a proarrhythmic risk from vericiguat and its M-1 metabolite in a series of preclinical studies. Methods The potential for proarrhythmic risk was investigated in conscious telemetered dogs and in a series of in vitro electrophysiological studies, including mechanistic ion channel studies, using both generally accepted and CIPA voltage-clamp protocols under conditions simulating normal and diseased physiological states. The ion channels studied were hERG, hNav1.5, hCav1.2, hKvLQT1/minK and hKv4.3. Transfected human embryonic kidney cell lines were used for the hERG, hNav1.5 and hKvLQT1/minK studies; transfected Chinese hamster ovary cell lines were used for the hCav1.2 and hKv4.3 studies. Results In dogs, administration of vericiguat as single oral doses was associated with dose-dependent decreases in arterial blood pressure (consistent with its mode of action) and compensatory increases in heart rate (Table 1). Heart rate-corrected QT (QTc) intervals were not prolonged by vericiguat to a clinically meaningful extent. Neither vericiguat nor its M-1 metabolite inhibited cardiac ion channels (hERG, hNav1.5, hCav1.2, hKvLQT1/minK and hKv4.3) at exposure multiples of >150-fold (Table 2). Conclusion There was no preclinical evidence of proarrhythmic risk from the in vitro (simulating normal and diseased physiological states) and in vivo assessment of vericiguat or its major N-glucuronide metabolite M-1. This integrated risk assessment of non-clinical data supports the conclusion that administration of vericiguat 10 mg once daily in humans is not associated with meaningful QTc prolongation. FUNDunding Acknowledgement Type of funding sources: Private company. Main funding source(s): Funding for this research was provided by Bayer AG, Berlin, Germany and Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA.

Pregnancy-Related Hormones Increase UGT1A1-Mediated Labetalol Metabolism in Human Hepatocytes

Pregnancy-related hormones (PRH) are recognized as important regulators of hepatic cytochrome P450 enzyme expression and function. However, the impact of PRH on the hepatic expression and function of uridine diphosphate glucuronosyltransferases (UGTs) remains unclear. Using primary human hepatocytes, we evaluated the effect of PRH exposure on mRNA levels and protein concentrations of UGT1A1, UGT2B7, and other key UGT enzymes, and on the metabolism of labetalol (a UGT1A1 and UGT2B7 substrate commonly prescribed to treat hypertensive disorders of pregnancy). Sandwich-cultured human hepatocytes (SCHH) from female donors were exposed to the PRH estradiol, estriol, estetrol, progesterone, and cortisol individually or in combination. We quantified protein concentrations of UGT1A1, UGT2B7, and four additional UGT1A isoforms in SCHH membrane fractions and evaluated the metabolism of labetalol to its glucuronide metabolites in SCHH. PRH exposure increased mRNA levels and protein concentrations of UGT1A1 and UGT1A4 in SCHH. PRH exposure also significantly increased labetalol metabolism to its UGT1A1-derived glucuronide metabolite in a concentration-dependent manner, which positively correlated with PRH-induced changes in UGT1A1 protein concentrations. In contrast, PRH did not alter UGT2B7 mRNA levels or protein concentrations in SCHH, and formation of the UGT2B7-derived labetalol glucuronide metabolite was decreased following PRH exposure. Our findings demonstrate that PRH alter expression and function of UGT proteins in an isoform-specific manner and increase UGT1A1-mediated labetalol metabolism in human hepatocytes by inducing UGT1A1 protein concentrations. These results provide mechanistic insight into the increases in labetalol clearance observed in pregnant individuals.

An Investigation on Glucuronidation Metabolite Identification, Isozyme Contribution, and Species Differences of GL-V9 In Vitro and In Vivo

GL-V9 is a prominent derivative of wogonin with a wide therapeutic spectrum and potent anti-tumor activity. The metabolism characteristics of GL-V9 remain unclear. This study aimed to clarify the metabolic pathway of GL-V9 and investigate the generation of its glucuronidation metabolites in vitro and in vivo. HPLC-UV-TripleTOF was used to identify metabolites. The main metabolite that we found was chemically synthesized and the synthetic metabolite was utilized as standard substance for the subsequent metabolism studies of GL-V9, including enzyme kinetics in liver microsomes of five different species and reaction phenotyping metabolism using 12 recombinant human UDP-glucuronosyltransferase (UGT) isoforms. Results indicated that the glucuronidation reaction occurred at C5-OH group, and 5-O-glucuronide GL-V9 is the only glucuronide metabolite and major phase II metabolite of GL-V9. Among 12 recombinant human UGTs, rUGT1A9 showed the strongest catalytic capacity for the glucuronidation reaction of GL-V9. rUGT1A7 and rUGT1A8 were also involved in the glucuronidation metabolism. Km of rUGT1A7-1A9 was 3.25 ± 0.29, 13.92 ± 1.05, and 4.72 ± 0.28 μM, respectively. In conclusion, 5-O-glucuronide GL-V9 is the dominant phase II metabolite of GL-V9 in vivo and in vitro, whose formation rate and efficiency are closely related to isoform-specific metabolism profiles and the distribution of UGTs in different tissues of different species.

Bioavailability and In Vivo Antioxidant Activity of a Standardized Polyphenol Mixture Extracted from Brown Propolis

Several lines of evidence demonstrate the antioxidant, anti-inflammatory and antimicrobial activities of propolis, mostly ascribed to its polyphenol content. However, little is known regarding the bioavailability of propolis in acute and prolonged settings of oral administration. In this study, we first determined the content of the main polyphenols in a brown propolis extract obtained using a patented extraction method (Multi Dinamic Extraction—M.E.D.) by RP-HPLC-UV-PDA-MSn analysis, followed by the bioavailability of galangin and chrysin, the most abundant polyphenols in the mixture (7.8% and 7.5% respectively), following acute (single bolus of 500 mg/kg containing about 3.65 mg of the polyphenol mixture) and prolonged (100, 250 and 500 mg/kg body for 30 days) oral administration in 30 male 8 weeks old C57BL/6 wild-type mice. In the acute setting, blood was taken at 30 s and 5, 10, 15, 20, 25, 30, 45, 60 and 120 min following the oral bolus. In the prolonged setting, blood samples were obtained after 10, 20 or 30 days of administration. At the end of treatment, expression of antioxidant enzymes (superoxyde dismutase, SOD-1; catalase, CAT; glutathione peroxidase, GSS) was evaluated in liver tissue. Following both acute and prolonged administration, neither galangin nor chrysin were detectable in the plasma of mice, whereas the glucuronide metabolite of galangine was detectable 5 min after acute administration. At the end of the prolonged treatment SOD-1 was found to have increased significantly, unlike CAT and GSS. Overall, these data suggest that oral administration of whole brown propolis extract is followed by rapid absorption and metabolization of galangin followed by adaptations of the antioxidant first line defense system.