Broad Spectrum Antiviral Properties of Cardiotonic Steroids Used as Potential Therapeutics for Emerging Coronavirus Infections

Cardiotonic steroids are steroid-like natural compounds known to inhibit Na+/K+-ATPase pumps. To develop a broad-spectrum antiviral drug against the emerging coronavirus infection, this study assessed the antiviral properties of these compounds. The activity of seven types of cardiotonic steroids against the MERS-CoV, SARS-CoV, and SARS-CoV-2 coronavirus varieties was analyzed using immunofluorescence antiviral assay in virus-infected cells. Bufalin, cinobufagin, and telocinobufagin showed high anti-MERS-CoV activities (IC50, 0.017~0.027 μM); bufalin showed the most potent anti-SARS-CoV and SARS-CoV-2 activity (IC50, 0.016~0.019 μM); cinobufotalin and resibufogenin showed comparatively low anti-coronavirus activity (IC50, 0.231~1.612 μM). Differentially expressed genes in Calu3 cells treated with cinobufagin, telocinobufagin, or bufalin, which had high antiviral activity during MERS-CoV infection were analyzed using QuantSeq 3′ mRNA-Seq analysis and data showed similar gene expression patterns. Furthermore, the intraperitoneal administration of 10 mg/kg/day bufalin, cinobufagin, or digitoxin induced 100% death after 1, 2, and 4 days in 5-day repeated dose toxicity studies and it indicated that bufalin had the strongest toxicity. Pharmacokinetic studies suggested that telocinobufagin, which had high anti-coronavirus activity and low toxicity, had better microsomal stability, lower CYP inhibition, and better oral bioavailability than cinobufagin. Therefore, telocinobufagin might be the most promising cardiotonic steroid as a therapeutic for emerging coronavirus infections, including COVID-19.

Cytochrome P450 enzymes: a review on drug metabolizing enzyme inhibition studies in drug discovery and development

Assessment of drug candidate's potential to inhibit cytochrome P450 (CYP) enzymes remains crucial in pharmaceutical drug discovery and development. Both direct and time-dependent inhibition of drug metabolizing CYP enzymes by the concomitant administered drug is the leading cause of drug–drug interactions (DDIs), resulting in the increased toxicity of the victim drug. In this context, pharmaceutical companies have grown increasingly diligent in limiting CYP inhibition liabilities of drug candidates in the early stages and examining risk assessments throughout the drug development process. This review discusses different strategies and decision-making processes for assessing the drug–drug interaction risks by enzyme inhibition and lays particular emphasis on in vitro study designs and interpretation of CYP inhibition data in a stage-appropriate context.

N–Skatyltryptamines—Dual 5–HT6R/D2R Ligands with Antipsychotic and Procognitive Potential

A series of N-skatyltryptamines was synthesized and their affinities for serotonin and dopamine receptors were determined. Compounds exhibited activity toward 5-HT1A, 5-HT2A, 5-HT6, and D2 receptors. Substitution patterns resulting in affinity/activity switches were identified and studied using homology modeling. Chosen hits were screened to determine their metabolism, permeability, hepatotoxicity, and CYP inhibition. Several D2 receptor antagonists with additional 5-HT6R antagonist and agonist properties were identified. The former combination resembled known antipsychotic agents, while the latter was particularly interesting due to the fact that it has not been studied before. Selective 5-HT6R antagonists have been shown previously to produce procognitive and promnesic effects in several rodent models. Administration of 5-HT6R agonists was more ambiguous—in naive animals, it did not alter memory or produce slight amnesic effects, while in rodent models of memory impairment, they ameliorated the condition just like antagonists. Using the identified hit compounds 15 and 18, we tried to sort out the difference between ligands exhibiting the D2R antagonist function combined with 5-HT6R agonism, and mixed D2/5-HT6R antagonists in murine models of psychosis.

Descriptors of Cytochrome Inhibitors and Useful Machine Learning Based Methods for the Design of Safer Drugs

Roughly 2.8% of annual hospitalizations are a result of adverse drug interactions in the United States, representing more than 245,000 hospitalizations. Drug–drug interactions commonly arise from major cytochrome P450 (CYP) inhibition. Various approaches are routinely employed in order to reduce the incidence of adverse interactions, such as altering drug dosing schemes and/or minimizing the number of drugs prescribed; however, often, a reduction in the number of medications cannot be achieved without impacting therapeutic outcomes. Nearly 80% of drugs fail in development due to pharmacokinetic issues, outlining the importance of examining cytochrome interactions during preclinical drug design. In this review, we examined the physiochemical and structural properties of small molecule inhibitors of CYPs 3A4, 2D6, 2C19, 2C9, and 1A2. Although CYP inhibitors tend to have distinct physiochemical properties and structural features, these descriptors alone are insufficient to predict major cytochrome inhibition probability and affinity. Machine learning based in silico approaches may be employed as a more robust and accurate way of predicting CYP inhibition. These various approaches are highlighted in the review.

122 Physiological effects of low level laser therapy on exercised horses

Low level laser therapy (LLLT) is a form of phototherapy which may stimulate healing of injuries and lesions from exercise. Our objective was to determine effects of LLLT before or after exercise on equine serum concentrations of cortisol, lactate, and cytochrome P450 (CYP) inhibition. Twelve quarter horses were randomly assigned to one of three groups: (A) received no laser therapy, (B) received six minutes of laser therapy before exercise, and (C) received six minutes of laser therapy after exercise. Laser (635 nm) treatment was with a class II handheld low-level laser with constant wave modulation. Horses were exercised using a walker for 30 min five days a week for three weeks, and within 30 min after exercise blood was collected on day 7, 14, and 21; blood was collected on day 0 prior to exercise as a reference point. Serum was collected, frozen; and cortisol, lactate, and CYP inhibition were determined via validated assays. Data were analyzed by ANOVA with horse as the experimental unit; an interaction (P < 0.05) between treatment group and day of study affected serum concentrations of cortisol, lactate, and CYP inhibition. For group B horses, cortisol concentrations decreased (P < 0.001) from day 14 to day 21 (31 vs 20 ± 2.9 ng/mL). Serum lactate concentrations increased (P < 0.0001) during the three weeks of training (2700, 2773, 3064, and 6061 ± 232 μM; respectively for day 0, 7, 14, and 21); whereas, inhibition of CYP decreased (P < 0.01) from day 0 to day 21. Interaction of main effects on lactate can be explained by larger increases in lactate concentrations for horses treated with LLLT during the trial; conversely, CYP inhibition decreased for horses treated with LLLT after exercise. Our results suggest that LLLT may be useful in improving performance of equine athletes.

121 Physiological effects of low level laser therapy on exercised horses

Low level laser therapy (LLLT) is a form of phototherapy, which may stimulate healing of injuries and lesions from exercise. Our objective was to determine effects of LLLT before or after exercise on equine serum concentrations of cortisol, lactate, and cytochrome P450 (CYP) inhibition. Twelve quarter horses were randomly assigned to one of three groups: (A) received no laser therapy, (B) received six minutes of laser therapy before exercise, and (C) received six minutes of laser therapy after exercise. Laser (635 nm) treatment was with a class II handheld low-level laser with constant wave modulation. Horses were exercised using a walker for 30 min five days a week for three weeks. Within 30 min after exercise blood was collected on day 7, 14, and 21; blood was collected on day 0 prior to exercise as a reference point. Serum was collected, frozen; and cortisol, lactate, and CYP inhibition were determined via validated assays. Data were analyzed by ANOVA with horse as the experimental unit; an interaction (P < 0.05) between treatment group and day of study affected serum concentrations of cortisol, lactate, and CYP inhibition. For group B horses, cortisol concentrations decreased (P < 0.001) from day 14 to day 21 (31 vs 20 ± 2.9 ng/mL). Serum lactate concentrations increased (P < 0.0001) during the three weeks of training (2700, 2773, 3064, and 6061 ± 232 μM; respectively for day 0, 7, 14, and 21); whereas, inhibition of CYP decreased (P < 0.01) from day 0 to day 21. Interaction of main effects on lactate can be explained by larger increases in lactate concentrations for horses treated with LLLT during the trial; conversely, CYP inhibition decreased for horses treated with LLLT after exercise. Our results suggest that LLLT may be useful in improving performance of equine athletes.