The oral protease inhibitor (PF-07321332) protects Syrian hamsters against infection with SARS-CoV-2 variants of concern

Abstract

There is an urgent need for potent and selective antivirals against SARS-CoV-2. Pfizer developed PF-07321332 (PF-332), a potent inhibitor of the viral main protease (Mpro, 3CLpro) that can be dosed orally and that is in clinical development. We here report that PF-332 exerts equipotent in vitro activity against the four SARS-CoV-2 variants of concerns (VoC) and that it can completely arrest replication of the alpha variant in primary human airway epithelial cells grown at the air-liquid interface. Treatment of Syrian Golden hamsters with PF-332 (250 mg/kg, twice daily) completely protected the animals against intranasal infection with the beta (B.1.351) and delta (B.1.617.2) SARS-CoV-2 variants. Moreover, treatment of SARS-CoV-2 (B.1.617.2) infected animals with PF-332 completely prevented transmission to untreated co-housed sentinels.

Fig. 1. Antiviral activity of PF-332 in human airway epithelial cells infected with SARS-CoV-2 B1.1.7.

Human airway epithelial cells, fully differentiated in an air-liquid culture system, were treated with compound at the basal site starting 1 h before infection with SARS-CoV-2 B.1.1.7 (alpha variant). Infection was done at the apical site. On a day 2 and b day 4 post viral RNA in culture supernatant was quantified. Each drug-treated condition is from 3 independent cultures. The infected/untreated control is from 4 independent cultures and the uninfected/untreated control from 2 independent cultures. Data in a, b represented as mean ± SD. Asterisks in a, b indicate the statistical significance between treated samples and the infected-untreated control. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (one-way ANOVA). Pi post-infection. Source data are provided as a Source Data file.

Fig. 2. In vivo efficacy of PF-332 against Beta SARS-CoV-2 (B.1.351) variant in Syrian hamsters.

a Design of the study. b Viral RNA levels in the lungs of control (vehicle-treated) and PF-332-treated (at 125 or 250 mg/kg, BID) SARS-CoV-2−infected hamsters at day 4 post-infection. Individual data and median values (indicated by bars) are presented and are expressed as log₁₀ SARS-CoV-2 RNA copies per mg lung tissue. Data were analyzed with the Mann–Whitney U test (two-sided). ***P = 0.0007, ****P < 0.0001. c Infectious viral loads in the lungs of control (vehicle-treated) and PF-332-treated SARS-CoV-2-infected hamsters at day 4 pi (expressed as log₁₀ TCID₅₀ per mg lung tissue). Individual data and median values (indicated by bars) are presented. Data were analyzed with the Mann–Whitney U test (two-sided). *P = 0.034, ****P < 0.0001. d Weight change at day 4 pi in percentage, normalized to the body weight at the day of infection. Bars represent means ± SD. Data were analyzed with the Mann–Whitney U test (two-sided). **P = 0.0031, ***P = 0.0003. e Cumulative severity score from H&E stained slides of lungs from control (vehicle-treated) and PF-332-treated hamsters. Individual data and median values (indicated by bars) are presented and the dotted line represents the median score of untreated non-infected hamsters. Data were analyzed with the Mann–Whitney U test (two-sided). ****P < 0.0001. All data b–e are from two independent experiments with n = 12 for vehicle and 250 mg/kg BID groups and n = 10 for 125 mg/kg BID group. PF-332 = PF-07321332. Source data are provided as a Source Data file.

Fig. 3. The effect of treatment with PF-332 on the transmission of the Delta variant to untreated sentinel hamsters.

a Design of the study. b Viral RNA levels in the lungs of control (vehicle-treated), PF-332-treated (250 mg/kg, BID) SARS-CoV-2-infected index hamsters (closed circles), and non-infected, non-treated contact hamsters (open circles) at day 3 and 4 post-infection (pi), respectively, are expressed as log₁₀ SARS-CoV-2 RNA copies per mg lung tissue. Individual data and median values (indicated by bars) are presented. c Infectious viral loads in the lungs of control (vehicle-treated), PF-332-treated SARS-CoV-2−infected index hamsters, and non-infected, non-treated contact hamsters at day 3 and 4 pi, respectively, are expressed as log₁₀ TCID₅₀ per mg lung tissue. Individual data and median values (indicated by bars) are presented. Data in b, c were analyzed with the Mann–Whitney U test (two-sided). **P = 0.0022. PF-322 = PF-07321332. The data in b, c are from a single experiment and with 6 animals per group. Source data are provided as a Source Data file.