Molnupiravir

molnupiravir (EIDD-2801) and GS-441524 were purchased from Excenen Pharmatech Co., Ltd. (China). For in vitro assays, 10 mM stocks of compounds were made by dissolving in DMSO. For in vivo studies, molnupiravir was formulated as 50 mg/ml stock in a vehicle containing 10% PEG400 (Sigma) and 2.5% Kolliphor-EL (Sigma) in water. GS-441524 was formulated as a 15 mg/ml stock in 30% PEG-400 (Sigma) in PBS containing 1%DMSO.

The antiviral drugs GS-441524, teriflunomide, ritonavir, and nirmatrelvir utilized in this study were purchased from MedChemExpress (Princeton, NJ, USA). Ruxolitinib was obtained from InvivoGen (San Diego, CA, USA), while molnupiravir was sourced from Sigma-Aldrich (St. Louis, MO, USA). The antiviral compounds were dissolved or resuspended in dimethyl sulfoxide to prepare stock solutions (5 mM) of all six drugs. The stock solutions of the drugs were stored at −20 °C for Ruxolitinib and −80 °C for GS-441524, molnupiravir, nirmatrelvir, ritonavir, and teriflunomide. The drugs were further diluted to working concentrations using EMEM medium for experimental use.

Eight-week-old Syrian Gold hamsters (72–96 g) were obtained from Janvier Labs (Saint Berthevin, France). The animals received either 150 mg/kg Molnupiravir (MedChemExpress LLC) twice daily or 20 mg/kg/day Teriflunomide (Biozol) via oral application using a feeding pipet (n = 3 per group). Teriflunomide was formulated in 20% PEG400 (Roth), 2% Tween-80 (Sigma Aldrich), 39% peanut oil (Braendle) and 39% strawberry syrup (Yo); Molnupiravir was formulated in 10% PEG400, 2% Tween-80, 44% water and 44% strawberry syrup. Molnupiravir was given twice daily with 8 h between first and second treatment each day in an application volume of 4 mL/kg. Formulations were freshly prepared before every application. At day one, PK samples were taken 0.5, 1, 2, 4, 8 and 24 h after the first dose. Compound concentrations were analyzed in whole blood using liquid chromatography-mass spectrometry. For Molnupiravir, its active metabolite, EIDD-1931/NHC, was measured.