The FDA-Approved Oral Drug Nitazoxanide Amplifies Host Antiviral Responses and Inhibits Ebola Virus

Jasenosky, Luke D.; Cadena, Cristhian; Mire, Chad E.; Borisevich, Viktoriya; Haridas, Viraga; Ranjbar, Shahin; Nambu, Aya; Bavari, Sina; Soloveva, Veronica; Sadukhan, Supriya; Cassell, Gail H.; Geisbert, Thomas W.; Hur, Sun; Goldfeld, Anne E.

NTZ also suppressed VSV replication, and its antiviral efficacy was markedly reduced upon depletion of RIG-I in A549 cells. However, unlike what we observed for EBOV, knockdown of PKR had no effect on NTZ inhibition of VSV. Furthermore, in the absence of NTZ, we found that knockdown of PKR resulted in a significant but milder increase in VSV growth relative to the robust increase in EBOV replication we observed when PKR expression was ablated. We note that the VSV matrix protein blocks host cell type I IFN responses independently of PKR by inhibiting global mRNA synthesis and nuclear export, as well as by inhibiting global host protein synthesis (Ahmed et al., 2003; Black and Lyles, 1992; Connor and Lyles, 2005; Ferran and Lucas-Lenard, 1997; von Kobbe et al., 2000) . The VSV matrix protein shuts down host cell translation primarily by inducing dephosphorylation of the eIF4F initiation complex component eIF4E Lyles, 2002, 2005) . However, eIF2a also becomes phosphorylated at later stages of VSV infection, leading to additional negative effects on host protein synthesis, including on type I IFN production, as well as on viral protein synthesis (Connor and Lyles, 2005) . Here, we have shown that NTZ induces transcription and translation of GADD34, which is a phosphatase that reverses eIF2a phosphorylation (Novoa et al., 2001) , and has been shown to be critical for restoring translation of IFN-b and other selected cytokines in virus-infected cells (Clavarino et al., 2012a (Clavarino et al., , 2012b Dalet et al., 2017) . Given that GADD34 depletion reduces NTZ activity against VSV, our data suggest that NTZ induction of GADD34 further amplifies NTZ-augmented RIG-I signaling, providing cells with the capacity to mount an innate immune response that is sufficient to overcome VSV matrix protein inhibitory effects upon protein synthesis.