Entwicklung von DNAzymen gegen Humane Rhinoviren: Design und in vitro Testung

Asthma bronchiale ist ein heterogenes Krankheitsbild aus wiederkehrender Ob-struktion der unteren Atemwege, bedingt durch Hyperreagibilität der Atemwegs-muskulatur, Hypersekretion und entzündlicher Ödembildung in der Schleimhaut, das weltweit über 300 Millionen Menschen betrifft. Oft beginnt die Kra...

Full description

Saved in:
Bibliographic Details
Main Author: Unger, Sebastian
Contributors: Renz, Harald (Prof. Dr. med.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2018
Online Access:PDF Full Text
Tags: Add Tag
No Tags, Be the first to tag this record!

Asthma is a heterogeneous syndrome of recurrent obstruction in the lower airways caused by hyperreactivity of the airway muscles, hypersecretion and in¬flammatory edema in the mucous membrane, which affects over 300 million pe¬ople worldwide. The disease often starts at a young age and affects all age groups as a chronic disease without curative therapy. The exact mechanisms and reasons for development have been the subject of research for many years and have as¬sured the interaction of genetic disposition and environmental influences as the most important factors. With constantly high disease rates in industrialized na¬tions and increasing prevalence in developing countries, asthma is associated with high costs for healthcare systems worldwide. High proportions of costs and burden for the affected patients are due to asthma exacerbations, with associated need for emergency treatment and hospital admissions, most often triggered by infection with human rhinoviruses. So far there is no established therapy against these pathogens of the common cold. With their genome consisting of a positive single strand RNA molecule, they offer a potential target for antisense molecules, in particular for DNA enzymes, which have increasingly been used successfully in research and clinical studies in recent years. In this work, DNA enzymes were developed against sections within the 5`-untranslated region and the CRE element of three representative viruses and tested for in vitro activity. The molecules were able to specifically cleave the viral RNA. By creating an in silico database it was in a second step possible to check the remaining, not directly tested virus types for the presence of the found cleavage sites. This revealed the problem of highly active DNA enzymes for individual virus types without broad coverage of the whole on the one hand, and DNA enzymes with broad coverage of almost all virus types but low activity on the other. The secondary structure of the virus genome was identified as the cause for dif¬fering activity and coverage between virus types. In a next step the cleavage ac¬tivity could be optimised by modifying the DNA enzyme binding domain length. Likewise, it has been possible to select promising DNA enzymes that are able to attack over 98 percent of known virus type isolates with excellent cleavage acti¬vity. These molecules can now be used and tested in further in vivo experiments to provide a targeted cure for cold viruses and the asthma exacerbations they cause.