Antiviral modified siRNA swarms for treatment of herpes simplex virus infection

Abstract

Herpes simplex virus type 1 (HSV-1) is a common virus of humans carried by half of the global population. After the primary infection, HSV has the ability to establish life-long latency, wherefrom it can reactivate. The latent state cannot be eliminated with modern pharmaceuticals, nor is there a vaccine available, despite massive efforts. Instead, the treatment focuses on diminishing viral replication. The current treatment, however, is insufficient, as it relies almost solely on acyclovir (ACV), and its derivatives, which share their mechanism of action, making ACV-resistant infections almost untreatable. Unfortunately, such infections are rather common, as severe HSV infections require long-term prophylactic treatment to prevent recurrences, which selects for ACV-resistant variants. The lack of treatment diversity against HSV-1 infections encourages for research on novel therapies.

Previously, enzymatically synthetized swarms of small interfering (si)RNA have been established as feasible means to treat HSV infection in vitro and in vivo. They differ from regular siRNA by their enzymatic synthesis and by their substantially longer target sequence. Thus, the emergence of resistance, even during long-term prophylactic treatment, is unlikely. However, as all RNA therapy, siRNA swarms face challenges with RNA stability. Therefore, in this study, the goal was to improve the siRNA swarms by synthesizing novel anti-HSV siRNA swarms with chemical 2′-fluoro-modifications to increase RNA efficacy and stability. The modified siRNA swarms, representing modifications of each nucleotide, were first validated in vitro in cells of the nervous system. The research was continued in a highly translational cell line representing the human cornea, which we first validated for use in antiviral RNAi studies. In both cell types, the modified siRNA swarm(s) proved well tolerated and potent beyond the unmodified counterparts, with only modest effects on the host innate responses, even in the presence of viral challenge. Furthermore, all studied HSV-1 strains, including various clinical isolates, were highly sensitive to both modified and unmodified siRNA swarms, whereas their ACV sensitivity varied, proving the potential of siRNA swarms for future therapeutic use.

This study shows that incorporation of modified nucleotides to the anti-HSV siRNA swarms is advantageous, and should therefore be preferred in future studies.