Old drugs with new tricks: Efficacy of fluoroquinolones to suppress replication of flaviviruses

Antiviral therapies are urgently needed to treat infections with flaviviruses such as Zika (ZIKV) and dengue (DENV) virus. Repurposing FDA-approved compounds could provide the fastest route to alleviate the burden of flaviviral diseases. In this study, three fluoroquinolones, enoxacin, difloxacin and ciprofloxacin, curtailed replication of flaviviruses ZIKV, DENV, Langat (LGTV) and Modoc (MODV) in HEK-293 cells at low micromolar concentrations. Time-of-addition assays revealed that enoxacin suppressed ZIKV replication when added at 6 hours post-infection, suggesting inhibition of an intermediate step in the virus life cycle, whereas ciprofloxacin and difloxacin had a wider window of efficacy of 2, 6, and 8 hours post-infection for difloxacin and 2 to 8 hours post-infection for ciprofloxacin. The efficacy of enoxacin to suppress ZIKV replication in 5-week-old A129 mice was evaluated in two experiments. First, mice were infected with 1×105 plaque-forming units (pfu) ZIKV FSS13025 (n=20) or PBS (n=11) on day 0 and subsets were treated with enoxacin at 10mg/kg or 15mg/kg or diluent orally twice daily on days 1-5. Treated and control mice did not differ in weight change or virus titer in serum or brain. Mice treated with enoxacin showed a significant, 5-fold decrease in ZIKV titer in testes relative to controls. Second, mice were infected with 1×102 pfu ZIKV (n=13) or PBS (n=13) on day 0 and subsets were treated with 15mg/kg oral enoxacin or diluent twice daily on days 0 (pre-treatment) and 1-5. Mice treated with enoxacin showed a significant, 2.5-fold decrease in ZIKV titer in testes relative to controls, while weight and viral load in the serum, brain, and liver did not differ between treated and control mice. Enoxacin efficacy in cultured murine Sertoli cells was not enhanced compared to efficacy in HEK-293 cells. ZIKV can be sexually transmitted, so reduction of titer in the testes by enoxacin should be further investigated. Author Summary Flaviviruses such as Zika and dengue virus pose a significant threat to public health worldwide, and there are currently no antiviral therapies to treat any flaviviral infection. Repurposing FDA-approved drugs as anti-flaviviral therapies can accelerate clinical use. We demonstrated that fluoroquinolone antibiotics exhibit anti-flaviviral efficacy, suppressing flavivirus replication in cultured human cells. Additionally, we found that the fluoroquinolone enoxacin suppressed Zika virus replication in mouse testes. While Zika virus is primarily transmitted via mosquitoes, the virus also undergoes sexual transmission. The importance of sexual transmission for the overall epidemiology of the virus remains unclear; nonetheless all routes of potential transmission to pregnant women are of concern as fetal infection in utero can have devastating effects. Thus, our data indicate that fluoroquinolones hold promise for treatment of flaviviral infections, particularly infection of the testes by Zika virus, and that this class of drugs warrants further study. ### Competing Interest Statement The authors have declared no competing interest.