Short hairpin-loop-structured oligodeoxynucleotides reduce HSV-1 replication

Alexander Falkenhagen,Karin Moelling,Jochen Heinrich

doi:10.1186/1743-422x-6-43

Alexander Falkenhagen, Karin Moelling + Show 1 more

Open Access

https://doi.org/10.1186/1743-422x-6-43

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Abstract

The Herpes simplex virus (HSV) is known as an infectious agent and widespread in the human population. The symptoms of HSV infections can range from mild to life threatening, especially in immune-compromised individuals. HSV infections are commonly treated with the guanosine analogue Aciclovir, but reports of resistance are increasing. Efforts are made to establish single-stranded antisense oligodeoxynucleotides (as) and small interfering ribonucleic acids (siRNAs) for antiviral treatment. Recently, another class of short interfering nucleic acids, partially double-stranded hairpin loop-structured 54 mer oligodeoxynucleotides (ODNs), was shown to allow hydrolysis of HIV RNA by binding to the viral RNA. This leads to a substrate for the viral RNase H. To assess the potential of such ODNs for inhibition of HSV-1 replication, five partially double-stranded ODNs were designed based on the sequences of known siRNAs against HSV-1 with antiviral activity. Three of them are directed against early and two against leaky late genes. Primary human lung fibroblasts, MRC-5, and African green monkey kidney cells, Vero, were transfected with ODNs and subsequently infected. The effect on HSV-1 replication was determined by analyzing the virus titer in cell culture supernatants by quantitative PCR and plaque assays. An inhibitory effect was observed with all five selected ODNs, with two cases showing statistical significance in both cell types. The observed effect was sequence-specific and dose dependent. In one case the ODN was more efficient than a previously described siRNA directed against the same target site in the mRNA of UL5, a component of the helicase/primase complex. HSV-1 virions and ODNs can be applied simultaneously without transfection reagent, but at a 50-fold higher concentration to Vero cells with similar efficiencies. The results underline the potential of partially double-stranded hairpin loop-structured ODNs as antiviral agents.

Highlights

The Herpes simplex virus (HSV), a member of the family Herpesviridae, is a large DNA virus with a high prevalence in the human population
HSV genes are expressed in a highly regulated cascade during productive infection [1]. They are grouped in immediate early (IE), early (E), leakylate (LL) and late genes (L) according to the time point of expression after infection
After 24 h the cell culture supernatants were harvested and HSV DNA levels were determined by quantitative PCR using primers and a probe targeting the HSV type 1 (HSV-1) glycoprotein G [15]

Summary

Introduction

The Herpes simplex virus (HSV), a member of the family Herpesviridae, is a large DNA virus with a high prevalence in the human population. After 24 h the cell culture supernatants were harvested and HSV DNA levels were determined by quantitative PCR (qPCR) using primers and a probe targeting the HSV-1 glycoprotein G [15]. The ODNs against HSV-1 reduced the viral titer by 60 to 80% in comparison to infected cells incubated with PBS (data not shown).

Results

Conclusion