Development and characterization of DNAzyme candidates demonstrating significant efficiency against human rhinoviruses

Publication date: Available online 14 August 2018

Source: Journal of Allergy and Clinical Immunology

Author(s): Daniel P. Potaczek, Sebastian D. Unger, Nan Zhang, Styliani Taka, Sven Michel, Nesibe Akdağ, Feng Lan, Markus Helfer, Christoph Hudemann, Markus Eickmann, Chrysanthi Skevaki, Spyridon Megremis, Anne Sadewasser, Bilal Alashkar Alhamwe, Fahd Alhamdan, Mübeccel Akdis, Michael R. Edwards, Sebastian L. Johnston, Cezmi A. Akdis, Stephan Becker

Abstract

Background

Infections with human rhinoviruses (RVs) are responsible for millions of common cold episodes and the majority of asthma exacerbations, especially in childhood. No drugs specifically targeting RVs are available.

Objective

To identify specific anti-rhinoviral molecules based on DNAzyme technology as candidates to a clinical study.

Methods

A total of 226 candidate DNAzymes were designed against two regions of RV RNA genome identified to be sufficiently highly conserved between virus strains, i.e. 5'-untranslated (5'-UTR) and cis-acting replication element (CRE), by use of three test strains, RVA1, -A16, and -A29. All DNAzymes were screened for their cleavage efficiency against in vitro-expressed viral RNA. Those showing any catalytic activity were subjected to bioinformatic analysis of their reverse complementarity to 322 published rhinoviral genomic sequences. Further molecular optimization was conducted for most promising candidates. Cytotoxic and off-target effects were excluded in HEK293-cell-based systems. Antiviral efficiency was analyzed in infected human bronchial BEAS-2B cells and ex vivo-cultured human sinonasal tissue.

Results

Screening phase generated DNAzymes characterized by either good catalytic activity or by high RV strain coverage but no single molecule represented a satisfactory combination of those two features. Modifications in length of the binding domains of two lead candidates, Dua-01(-L12R9) and Dua-02(-L10R11), improved their cleavage efficiency to an excellent level with no loss in eminent strain coverage (about 98%). Both DNAzymes showed highly favorable cytotoxic/off-target profiles. Subsequent testing of Dua-01-L12R9 in BEAS-2B cells and sinonasal tissue demonstrated its significant antiviral efficiency.

Conclusions

Effective and specific management of RV infections with Dua-01-L12R9 might be useful in preventing asthma exacerbations, which should be verified by clinical trials.

Graphical abstract

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