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After discovering that the HI-virus is responsible for AIDS, over 20 different drugs were approved for therapeutic use against this disease. Therefore, there are more drugs available against AIDS than against all other virus diseases. The first HIV-protease inhibitor, a member of the A2 family of aspartic proteases, became available in HAART (Highly Active Antiretroviral Therapy) in 1995 and resulted in a great increase in quality of life of HIV-infected patients. Nonetheless, HAART achieves less than optimal results in many cases because poor bioavailability and toxicity result in serious side effects and resistance of the HI-virus develops fast. So it remains of great interest to develop new inhibitor classes that are based on non-peptidic scaffolds and feature different binding modes.
Members of the A1 family of aspartic proteases are promising drug targets, too. However, until now only the renin inhibitor Aliskiren received approval for therapeutic use. Despite great efforts to develop inhibitors against BACE1 or Cathepsin D, no active substance could reach the market yet.
This thesis describes how novel classes of substances were designed for the inhibition of the A1 and A2 families of aspartic proteases. Endothiapepsin was chosen as a representative of the A1 family and HIV-protease as a representative of the A2 family. Five different inhibitor classes based on new scaffolds were computer-based designed, synthesized and analyzed with respect to their affinities to the selected enzymes. Cyclic urea compounds, piperidine-diesters, three-armed piperidines, open-chained primary amines and hydroxylamines were developed. It was possible to optimize at least one inhibitor of each class, except for the cyclic urea compounds, to reach submircomolar affinity ranges.
The cyclic urea inhibitor class achieved low micromolar affinities against endothiapepsin. This is a highly satisfying result, considering that these compounds are comparatively small and offer only two substituents to address the specificity pockets.
The piperidine-diesters were found to be promising scaffolds for the inhibition of A1 family aspartic-proteases, too. Two of the compounds developed in this series showed affinities of about 750 nM against endothiapepsin, although there were again only two residues to occupy the specificity pockets. It seems feasible to further increase these affinities by selected optimization steps.
No significant improvement could be observed for the three-armed piperidine derivatives. In a series of eleven synthesized compounds, nine achieved affinities at least in the single-digit micromolar range against endothiapepsin. For HIV-protease, the most active compound is an indazol-derivative with 14 M.
The open-chained, primary amines showed very good affinities against endothiapepsin. In this series, the most potent inhibitor shows a Ki-value of 550 nM against endothiapepsin. The other compounds showed good to moderate affinities as well, whereas the affinities against HIV-protease were considerably lower.
The four substance classes mentioned before showed significantly better affinities against endothiapepsin than against HIV-protease. In contrast, the hydroxylamine-class represents a new scaffold that showed better affinities to HIV-protease. It is therefore the most promising class for the design of HIV-protease-inhibitors. By variation of the substitution pattern, it was possible to identify two submicromolar inhibitors against the HIV-protease in this series. Trifluormethyl-derivative achieved an affinity of 140 nM, which is a very good result for an initial series with no previous knowledge about the binding mode. A reproducible structure-activity relationship could be determined for the hydroxylamine-derivatives, which allows a prediction about a possible binding mode.
As part of this work, a one-pot method to convert alcohols into the corresponding amines in a Mitsunobu-Staudinger-reaction using DPPA as an azide source avoiding the isolation of the azide intermediate was developed. In addition, a study concerning the usage of the detergent Trition X-100 in fluorescence-based binding-assays was conducted. Depending on concentration, Triton X-100 can have negative effects on assay results.