Pharmacokinetics and Efficacy of a Potential Smallpox Therapeutic, Brincidofovir, in a Lethal Monkeypox Virus Animal Model

Abstract

Smallpox, caused by Variola virus (VARV), was eradicated in 1980; however, VARV bioterrorist threats still exist, necessitating readily available therapeutics. Current preparedness activities recognize the importance of oral antivirals and recommend therapeutics with different mechanisms of action. Monkeypox virus (MPXV) is closely related to VARV, causing a highly similar clinical human disease, and can be used as a surrogate for smallpox antiviral testing. The prairie dog MPXV model has been characterized and used to study the efficacy of antipoxvirus therapeutics, including recently approved TPOXX (tecovirimat). Brincidofovir (BCV; CMX001) has shown antiviral activity against double-stranded DNA viruses, including poxviruses. To determine the exposure of BCV following oral administration to prairie dogs, a pharmacokinetics (PK) study was performed. Analysis of BCV plasma concentrations indicated variability, conceivably due to the outbred nature of the animals. To determine BCV efficacy in the MPXV prairie dog model, groups of animals were intranasally challenged with 9 × 10⁵ plaque-forming units (PFU; 90% lethal dose [LD₉₀]) of MPXV on inoculation day 0 (ID0). Animals were divided into groups based on the first day of BCV treatment relative to inoculation day (ID-1, ID0, or ID1). A trend in efficacy was noted dependent upon treatment initiation (57% on ID-1, 43% on ID0, and 29% on ID1) but was lower than demonstrated in other animal models. Analysis of the PK data indicated that BCV plasma exposure (maximum concentration [C_max]) and the time of the last quantifiable concentration (AUC_last) were lower than in other animal models administered the same doses, indicating that suboptimal BCV exposure may explain the lower protective effect on survival.IMPORTANCE Preparedness activities against highly transmissible viruses with high mortality rates have been highlighted during the ongoing coronavirus disease 2019 (COVID-19) pandemic. Smallpox, caused by variola virus (VARV) infection, is highly transmissible, with an estimated 30% mortality. Through an intensive vaccination campaign, smallpox was declared eradicated in 1980, and routine smallpox vaccination of individuals ceased. Today's current population has little/no immunity against VARV. If smallpox were to reemerge, the worldwide results would be devastating. Recent FDA approval of one smallpox antiviral (tecovirimat) was a successful step in biothreat preparedness; however, orthopoxviruses can become resistant to treatment, suggesting the need for multiple therapeutics. Our paper details the efficacy of the investigational smallpox drug brincidofovir in a monkeypox virus (MPXV) animal model. Since brincidofovir has not been tested in vivo against smallpox, studies with the related virus MPXV are critical in understanding whether it would be protective in the event of a smallpox outbreak.

Keywords: animal models; experimental therapeutics; monkeypox; smallpox; virology.

FIG 1

Animals (n = 16) were dosed with a three-dose (q48h) regimen of 20 (n = 8) or 5 (n = 8) mg/kg BCV via oral gavage, so that BCV pharmacokinetics could be determined following single (a, b) and repeat (c, d) dosing. An increase in concentrations of BCV with increased dose is seen. BCV plasma concentrations were generally below the limit of quantitation (1 ng/ml; BLQ values were not plotted) by 24 h following the 5-mg/kg dose (c) and by 48 h following the 20-mg/kg BCV dose (d). Data for female animals (F) are represented by filled blue symbols, and data for male animals (M) are represented by open red symbols. *, two animals (ID numbers 12054 and 13070) mistakenly received 20 mg/kg instead of 5 mg at the 48-h drug administration point.

FIG 2

Survival curve for groups of MPXV-challenged prairie dogs. Animals were treated with brincidofovir starting 1 day preinfection (–1), on the day of infection (day 0), or 1 day postinfection (day 1) with MPXV. Placebo (vehicle-only) animals were challenged with MPXV and treated with the vehicle to serve as positive controls. Differences in mortality rates and day of euthanasia among all groups (day −1, day 0, day +1, vehicle only) were not statistically significant (P = 0.56 and P = 0.61, respectively). There was no significant difference in survivorship between the treatment timing groups and vehicle only group (P = 0.42). A survival trend was observed, with more animals surviving the sooner brincidofovir treatment began.

FIG 3

Comparison of immune responses (days 11 and 14) between groups of MPXV-challenged prairie dogs, using ELISA. Optical density minus the cutoff value (OD–COV) levels were compared between treatment groups (preinfection [day −1], day of infection [day 0], and 1 day postinfection [day 1] groups and the placebo group [vehicle]) using the Wilcoxon rank sum test, as data were not normally distributed.

FIG 4

Viable virus was detected in necropsy samples from MPXV-challenged prairie dogs. Animals were treated with BCV starting 1 day preinfection (day −1) (a), on the day of infection (day 0) (b), or 1 day postinfection (day 1) (c). (d) Placebo (vehicle-only) animals were challenged with MPXV and treated with the vehicle to serve as positive controls. * indicates that one or more of those samples had the cell culture monolayer destroyed or that plaques were present but below the limit of detection (LOD) for this assay. † indicates that the animal died before the study end (day 25 p.i.). SM Ln, SM LN, or Sm Ln, submandibular lymph node; Sm. Int., small intestine; Mes LN, mesenteric lymph node.

FIG 5

Viral titration comparison of treatment groups on day 14 p.i. One animal from each group (vehicle, 1 day postinfection [TX], day of infection [day 0], or preinfection [day −1]) succumbed on day 14. Viral titer results are shown as PFU per gram of tissue for comparison between the animals. TX, treatment; SM LN, submandibular lymph node; Sm. Int., small intestine; Mes LN, mesenteric lymph node.