USC-087 protects Syrian hamsters against lethal challenge with human species C adenoviruses

Abstract

Human adenoviruses (AdV) cause generally mild infections of the respiratory and GI tracts as well as some other tissues. However, AdV can cause serious infection in severely immunosuppressed individuals, especially pediatric patients undergoing allogeneic hematopoietic stem cell transplantation, where mortality rates are up to 80% with disseminated disease. Despite the seriousness of AdV disease, there are no drugs approved specifically to treat AdV infections. We report here that USC-087, an N-alkyl tyrosinamide phosphonate ester prodrug of HPMPA, the adenine analog of cidofovir, is highly effective against multiple AdV types in cell culture. USC-087 is also effective against AdV-C6 in our immunosuppressed permissive Syrian hamster model. In this model, hamsters are immunosuppressed by treatment with high dose cyclophosphamide. Injection of AdV-C6 (or AdV-C5) intravenously leads to a disseminated infection that resembles the disease seen in humans, including death. We have tested the efficacy of orally-administered USC-087 against the median lethal dose of intravenously administered AdV-C6. USC-087 completely prevented or significantly decreased mortality when administered up to 4 days post challenge. USC-087 also prevented or significantly decreased liver damage caused by AdV-C6 infection, and suppressed virus replication even when administered 4 days post challenge. These results imply that USC-087 is a promising candidate for drug development against HAdV infections.

Figure 1. Structures (A) and in vitro anti-HAdV activity (B) of CDV, HPMPA and USC-087

Antiviral activity (against HAdV-C5) and cytotoxicity were evaluated in 3–5 separate experiments and EC₅₀ values (open symbols) and CC₅₀ values (solid symbols) are shown with horizontal bars representing the mean and standard deviation of the data.

Figure 2. USC-087 administered prophylactically reverses Ad5-induced mortality and morbidity, mitigates Ad5-induced liver damage, and decreases virus burden in the liver of immunosuppressed female Syrian hamsters

Ad5-infected hamsters were administered with vehicle or 10 mg/kg of USC-087 p.o. q.d., or with 20 mg/kg of CDV i.p. 3 times weekly; starting 1 day before challenge. A. Diagram of the experimental design. B. Survival. Ad5-Vehicle v. all other groups p=0.0161 (Log rank). C. Serum alanine aminotransferase levels. For this and similar subsequent figures, each symbol represents the value from an individual animal; the horizontal bar signifies the mean. D. Virus burden in the liver. NQ: not quantifiable (below the quantification threshold of 5×10⁴ TCID₅₀/g); *: p<0.05, **: p<0.01

Figure 3. Prophylactically administered USC-087 is more effective in counteracting the effects of disseminated Ad6 infection in female Syrian hamsters than equimolar doses of CDV

USC-087 was administered p.o. starting at one day prior to infection and continued daily for the course of the experiment. CDV was administered i.p. starting with an induction dose one day before virus challenge and continued with 3 weekly injections for the duration of the experiment. A. Diagram of the experimental design. B. Survival. Ad6-Vehicle v. Ad6-USC-087 1 mg/kg p=0.0047; Ad6-Vehicle v. Ad6-USC-087 3 mg/kg or Ad6-USC-087 10 mg/kg p<0.0001; Ad6-Vehicle v. Ad6-CDV 0.8 mg/kg p=0.4505; Ad6-Vehicle v. Ad6-CDV 2.4 mg/kg p=0.0178; Ad6-Vehicle v. Ad6-CDV 8 mg/kg p=0.0001; Ad6-Vehicle v. Ad6-CDV 8 mg/kg p<0.0001 (Log rank); C. Serum alanine transaminase levels. Empty symbols in this and subsequent graphs indicate that the sample was collected from a moribund animal sacrificed ahead of schedule. D. Virus burden in the liver. *: p<0.05, **: p<0.01; for the purposes of statistical calculations, a value of 5×10⁴ TCID₅₀/g was assumed for all not detectable and not quantifiable samples; NQ: not quantifiable; ND: not detectable

Figure 4. USC-087 reduces mortality and morbidity even when administered 3 or 4 days after virus challenge

CP-treated hamsters were infected i.v. with Ad6 and left untreated or treated with USC-087 at 10 mg/kg p.o. q.d. Treatment with the drug started at 1 day before or 1, 2, 3, or 4 days after virus challenge. A. Diagram of the experimental design. B. Survival. n=15. Ad6-Vehicle v. all Ad6-USC-087 p<0.05 (Log rank) C. Serum ALT levels at 7 days post challenge n=5 to 9. *: p<0.05 D. Virus burden in the liver at 7 days post challenge. Empty symbols represent data collected from moribund animals. **: p<0.01, ***: p<0.001; for the purposes of statistical calculations, a value of 5×10⁴ TCID₅₀/g was assumed for all not detectable and not quantifiable samples), NQ: not quantifiable, ND: not detectable. E. Mean body weight changes for the Vehicle-Vehicle and the Vehicle-USC-087 groups. p<0.0001 (Two-Way ANOVA).