Research Report 2010 2011 - Helmholtz-Zentrum für ...

112 SCIENTIFIC REPORTS | Infection and Immunity | Pharmaceutical Research
05.3 Chemical Biology of Infectious Disease
PROJECT LEADERS | Dr. Ronald Frank | Department of Chemical Biology | rfr@helmholtz-hzi.de
Dr. Florenz Sasse | Department of Chemical Biology | fsa@helmholtz-hzi.de
PROJECT MEMBERS | Jihad Al-Qudsi | Ulrike Beutling | Randi Diestel | Dr. Michelle Fountain | Dr. Raimo Franke |
Dr. Bernd Hofer | Denis Koska | Michael Mrosek | Dr. Irina Nickeleit | Dr. Mahtab Nourbakhsh | Dr. Marc Reboll |
Saad Shaaban | Galina Sergeev | Dennis Schwab | Dr. Dr. Werner Tegge | Dr. Peter Washausen | Marina Wöhl
The aim of the project is to elucidate the molecular mechanisms
of infection processes by interfering small molecules.
Special assay systems and screening techniques are developed
to select bioactive substances out of chemical libraries
whose mode of action will be further analysed. These
analyses should lead to new antibiotics, chemotherapeutics,
and immune modulators. The knowledge about their mode
of actions will open new ways of therapy. Within the framework
of the „Chemical Pipeline“, we built an infrastructure
that allows a rapid and effective search in miniaturised
assay systems. Our library of about 90,000 samples includes
a unique collection of natural compounds isolated from
myxobacteria (see 05.1), substances from collaboration
partners, purchased compound collections, and substances
that were synthesised by us. The infrastructure is also at
the disposal of external scientists and to other applications
via the German ChemBioNet (www.chembionet.de).
Phenotypes characterise the mode of action There is
no easy way to elucidate the mode of action of a biologically
active compound. Each biologically active compound
changes the metabolic and signalling pathways within the
Biologically active substances induce phenotypical changes in
treated cells which are characteristic for their mode of action.
This is shown for chivosazol A. The green actin filament structures
of the treated cells in B clearly differ from the control cells
in A. In C we see a hierarchical cluster analysis of substances
from myxobacteria. D gives a detail which shows that chivosazol
A clusters together with a second sample that was given “blind”,
a clear proof of principle.
complex biochemical network of a target cell. This can lead
to striking changes in the morphology of the cell and the
cell organelles or in the protein pattern. These phenotypical
changes can be visualised via antibodies or specific dyes.
Comparing phenotypes is one way to get hints about the
mode of action. Now we have strengthened this method
by automated microscopy and statistical analysis of large
amounts of data. By automated microscopy we can take a lot
of pictures of treated cells, evaluate them by software-based
image analysis, and form a profile of the biologically active
compound. By comparing the profiles of known and unknown
compounds we get hints about the mode of action of
the new substance. This is done by software-based statistical
methods, e.g., hierarchical cluster analysis. Substances,
which have the same mode of action, group together then.
In the meantime we have also got useful hints concerning
the mode of action of new substances. First follow-up
experiments confirmed the first hints.
Inhibitors of the adherence of Staphylococcus aureus to
human epithelial cells In the context of the BMBF funded
consortium “SkinStaph” a high-throughput amenable adhesion
assay was established, which provides the possibility
to search for new inhibitors of the interaction between the
pathogenic bacterium Staphylococcus aureus and human
epithelial cells (see Fig. page 109). Highly reliable data is
obtained by a newly developed method that employs an
automated microscope. Epithelial cells are incubated with
the bacteria in the presence of potentially interfering compounds,
followed by the removal of non-adherent bacteria
and the staining of the cells. The nuclei of the epithelial
cells are stained with a DNA specific dye (DAPI) and the
bacteria are labelled with specific fluorescent antibodies.
Several pictures for every compound are taken with the
auto mated microscope. With a software module the nuclei
and the bacteria are automatically quantified and set into
relation. This allows the identification of substances that
reduce the adhesion of the bacteria. After screening more
than 4,000 individual substances and mixtures, several
promising compounds were identified. These drug candidates
are currently investigated with primary human nasal
cells and their toxicity is determined with cell-based assays.
Diestel, R., Irschik, H., Jansen, R., Khalil, M.W., Reichenbach, H., & Sasse, F. (2009)
Chivosazole A and F, cytostatic macrolides from Myxobacteria, interfere with actin.
ChemBioChem 10, 2900-2903.