Brilacidin, a Non-Peptide Defensin-Mimetic Molecule, Inhibits SARS-CoV-2 Infection by Blocking Viral Entry.

Brilacidin (PMX-30063), a non-peptide defensin-mimetic small molecule, inhibits SARS-CoV-2 viral infection but the anti-viral mechanism is not defined. Here we determined its effect on the specific step of the viral life cycle. Brilacidin blocked SARS-CoV-2 infection but had no effect after viral entry. Brilacidin inhibited pseudotyped SARS-CoV-2 viruses expressing spike proteins from the P.1 Brazil strain and the B.1.1.7 UK strain. Brilacidin affected viral attachment in hACE2-dependent and independent manners depending on the concentrations. The inhibitory effect on viral entry was not mediated through blocking the binding of either the spike receptor-binding domain or the spike S1 protein to hACE2 proteins. Taken together, brilacidin inhibits SARS-CoV-2 infection by blocking viral entry and is active against SARS-CoV-2 variants.

Brilacidin Demonstrates Inhibition of SARS-CoV-2 in Cell Culture.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the newly emergent causative agent of coronavirus disease-19 (COVID-19), has resulted in more than two million deaths worldwide since it was first detected in 2019. There is a critical global need for therapeutic intervention strategies that can be deployed to safely treat COVID-19 disease and reduce associated morbidity and mortality. Increasing evidence shows that both natural and synthetic antimicrobial peptides (AMPs), also referred to as Host Defense Proteins/Peptides (HDPs), can inhibit SARS-CoV-2, paving the way for the potential clinical use of these molecules as therapeutic options. In this manuscript, we describe the potent antiviral activity exerted by brilacidin-a de novo designed synthetic small molecule that captures the biological properties of HDPs-on SARS-CoV-2 in a human lung cell line (Calu-3) and a monkey cell line (Vero). These data suggest that SARS-CoV-2 inhibition in these cell culture models is likely to be a result of the impact of brilacidin on viral entry and its disruption of viral integrity. Brilacidin demonstrated synergistic antiviral activity when combined with remdesivir. Collectively, our data demonstrate that brilacidin exerts potent inhibition of SARS-CoV-2 against different strains of the virus in cell culture.

Pharmacokinetics of sildenafil after single oral doses in healthy male subjects: absolute bioavailability, food effects and dose proportionality.

AIMS: To determine the absolute bioavailability, dose proportionality and the effects of food on the pharmacokinetics of single oral doses of sildenafil citrate. METHODS: Three open-label, randomized crossover studies were conducted in healthy male subjects. Absolute bioavailability was determined by comparing pharmacokinetic data after administration of single oral and intravenous 50-mg doses of sildenafil (n=12 subjects). Food effects were examined by comparing pharmacokinetic data for sildenafil and its primary circulating metabolite, UK-103,320, after administration of a single oral 100-mg dose in the fasted and fed states (n=34 subjects). Dose proportionality was assessed from pharmacokinetic data obtained after administration of four single oral doses of sildenafil (25, 50, 100 and 200 mg) to 32 subjects. The safety and tolerability of sildenafil were also assessed in all of these studies. RESULTS: The calculated absolute oral bioavailability of sildenafil was 41% (90% CI: 36--47). Food slowed the rate of absorption, delaying mean tmax by approximately 1 h and reducing Cmax by 29% (90% CI: 19--38). Systemic exposure, as assessed by the mean area under the plasma concentration--time curve (AUC), was reduced by 11% (90% CI: 6--16). These food effects were not considered to be of clinical significance. There was statistical evidence of nonproportionality in Cmax and AUC over the dose range 25--200 mg. However the degree of nonproportionality was small, with predicted increases in Cmax and AUC of 2.2- and 2.1-fold, respectively, for a doubling in dose, and was thought to be clinically nonsignificant. Sildenafil was well tolerated in the three studies; the majority of adverse events were mild and transient. CONCLUSIONS: Sildenafil had a mean absolute bioavailability of 41%. Food caused small reductions in the rate and extent of systemic exposure; these reductions are unlikely to be of clinical significance. Across the dose range of 25--200 mg, systemic exposure increased in a slightly greater than dose-proportional manner.

The effects of steady-state erythromycin and azithromycin on the pharmacokinetics of sildenafil in healthy volunteers.

AIMS: Sildenafil, an effective oral treatment for erectile dysfunction, is predominantly metabolized by the cytochrome P450 isozyme 3A4, which is inhibited by a number of the macrolide antibiotics. Therefore, two placebo-controlled, parallel-group studies were conducted to evaluate the effects of multiple doses of erythromycin and azithromycin on the pharmacokinetics, safety and tolerability of a single oral 100-mg dose of sildenafil. METHODS: In the erythromycin interaction study, 26 male volunteers (18--45 years of age) received open-label sildenafil 100 mg on day 1. Half received blinded erythromycin (500 mg) twice daily on days 2--6, and the other half received placebo. On day 6, all subjects received a second 100-mg dose of sildenafil. In the azithromycin interaction study, 24 male volunteers (19--33 years of age) received open-label 100 mg sildenafil on day 1. Half then received blinded azithromycin (500 mg) once daily on days 2--4, and the other half received placebo. On day 4, all subjects received another 100-mg dose of sildenafil. In both studies, blood samples were collected on the first and last study day for the analysis of plasma concentrations of sildenafil and its primary metabolite, UK-103,320. RESULTS: Repeated dosing with erythromycin caused statistically significant increases in the AUC and Cmax of sildenafil (2.8-fold and 2.6-fold, respectively) but had no effect on Tmax, kel or t1/2. A statistically significant 1.4-fold increase in the AUC of UK-103,320 was also observed, as well as a significant decrease in kel, resulting in an increase of about 1 h in t1/2. In contrast, repeated dosing with azithromycin caused no significant change in any pharmacokinetic parameter of either sildenafil or UK-103,320. Erythromycin, azithromycin and sildenafil were well tolerated; adverse events were mild and transient. No subject withdrew from either trial for any reason related to study drug. CONCLUSIONS: These results indicate that erythromycin modifies the pharmacokinetics of sildenafil by inhibiting its CYP3A4-mediated first-pass metabolism. Given these data, a lower starting dose of sildenafil (25 mg) may be considered for patients receiving erythromycin or other potent CYP3A4 inhibitors. Azithromycin did not affect the pharmacokinetics of sildenafil; therefore, no adjustment in dosage is necessary for patients receiving these drugs concomitantly.

The effects of sildenafil on human sperm function in healthy volunteers.

AIMS: This double-blind, randomized, four-period, two-way crossover study was conducted to evaluate the acute effects of oral sildenafil (100-mg single dose) on sperm motility, count, density, morphology and vitality as well as ejaculate volume and viscosity in healthy male subjects. The concentrations of sildenafil and its primary circulating metabolite UK-103,320 were measured in ejaculate and compared with those in plasma. The study also included assessments of safety and tolerability. METHODS: A total of 17 healthy male volunteers aged 19--34 years were randomized to receive a single 100-mg dose of sildenafil for two periods and a single dose of placebo for two periods, with each period separated by a minimum of 5--7 days. Sperm and ejaculate properties were evaluated from semen samples taken at screening and 1.5 h after dose. An additional semen sample was collected 4 h after dose, and drug and metabolite concentrations were measured in this sample and the sample taken 1.5 h after dose for comparison with plasma concentrations. Blood samples were collected before each dose and 0.25, 0.5, 1, 2, 3, 4 and 6 h after dose for measurement of sildenafil and metabolite concentrations. RESULTS: Sildenafil had no statistically significant effect on sperm motility, count or density; the percentage of abnormal sperm forms; or the percentage of living sperm. It also did not affect ejaculate volume or viscosity. All measures were within normal ranges. Sildenafil distributed into the semen rapidly, resulting in significant correlations between concentrations of sildenafil in the semen and total (R2=0.588) or free (R2=0.454) plasma concentrations (P<0.0001). Total semen concentrations of sildenafil were 18% of total plasma concentrations. UK-103,320 appeared to distribute more slowly from the plasma into the semen, resulting in a lack of correlation between semen and plasma concentrations. The amount of sildenafil and UK-103,320 in the ejaculate was small (< 2x10(-4)% of the administered dose at 1.5 h). Sildenafil was well tolerated; no patient withdrew from the study due to adverse events attributed to sildenafil. CONCLUSIONS: These results indicate that a single 100-mg oral dose of sildenafil does not have an adverse effect on sperm function or ejaculate quality.