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Functional genomics of complex regulatory networks from yeast to human: cross-talk of sterol homeostasis and drug metabolism

Final Report Summary - STEROLTALK (Functional genomics of complex regulatory networks from yeast to human: cross-talk of sterol homeostasis and drug metabolism)

The vision of STEROLTALK was to combine dedicated functional genomic tools, three model organisms and in silico modelling, for the discovery of new drug targets and novel chemical entities for treatment of hyperlipidemias, and to establish the drug-drug interactions. This has allowed STEROLTALK to importantly contribute to deciphering the multi-level response of cholesterol homeostasis to known drugs (statins) and candidate drugs (LK-compounds). Most of research has been performed in human primary hepatocytes and in wild type or nuclear receptor knockout mouse livers, and in recombinant yeast strains that express key human enzymes of cholesterol synthesis. In addition to commercially available tools we have developed original STEROLTALK tools for functional genomic studies of the hypolipidemic drugs. Tools include a dedicated STEROLTALK microarray in the human and mouse version that has been used successfully in drug-perturbation studies of the human primary hepatocytes and in mouse experiments. The STEROLTALK antibody set includes over 20 evaluated antibodies for the human drug metabolizing cytochromes P450 (CYPs) and antipeptide antibodies against enzymes of the cholesterol synthesis, majority of which are novel and are not available commercially. The STEROLTALK recombinant yeast strains that overexpress human enzymes of the cholesterol synthesis pathway were not only proven to represent original hypolipidemic drug screening tools, but are also a resource for the recombinant human cholesterogenic enzymes that can be used for kinetic studies or applied as epitopes for the new antibody production. Analytical (microarray and RT-PCR results) and sterol metabolome data and a limited set of the available western analyses results have been joined with the literature data to build the in silico model of cholesterol synthesis. This dynamical model was able to reproduce the experimentally verified action of statins, the well-known drugs of cholesterol synthesis, as well as the two novel potential drugs from the LK series of Lek Pharmaceuticals.

The STEROLTALK goal to build an in silico model that would predict the action of the novel hypolipidemic drugs, has thus fully been reached. Additionally, the Byesian approach to construction of gene regulatory networks has led to the prediction of another transcriptional regulator of the cholesterol synthesis, which has been proven experimentally. Furthermore, proteomic tools (two-dimensional (2D) gel electrophoresis and nLC-MS) have been established, optimised and successfully used for the analysis of human hepatocytes. In addition to already known players, proteins not yet known to be involved in STEROLTALK relevant regulation processes were found to be differentially regulated. Last but not least, the STEROLTALK database served as a share point for the exchange of experimental protocols and samples with unique Steroltalk coded signatures. The database served as well as a repository for the transcriptome and RT-PCR data, applying the Base 2.7.2. software that includes the MIAME standard of the minimal required information about a microarray experiment.

It is important to note that the formal end of the EU funding of the STEROLTALK does not mean an end to the developed STEROLTALK tools, models and samples. The University of Ljubljana, where the scientific coordination took place, will continue to maintain and upgrade the STEROLTALK database in collaboration with Crea, d.o.o. P10 of this project. The database will remain available to the STEROLTALK partners without additional cost for one year and will be continued afterwards with the shared cost contribution.

Additionally, the model of the successful sharepoint within an international consortium, for experiment, protocol and sample exchange, will be used as a template in planning research under the current and future Seventh Framework Programme (FP7) systems biology calls. Also other STEROLTALK tools will remain alive in further research - the idea of a dedicated microarray that, in combination with commercial pan-genomic arrays, leads to a much quicker hypothesis formulation; the collection of STEROLTALK antibodies and the heterologous expression of STEROLTALK proteins in the yeast, will be very helpful in further research and can as well find commercial application. Additionally, research samples, such as mouse livers, human and mouse RNAs and other isolates, remain available for additional analyses within the broader scientific community. These valuable research samples from well controlled STEROLTALK experiments can contribute to a quicker solving of the questions regarding the cholesterol synthesis and drug metabolism networks that remained un-answered after the three-year STEROLTALK period.

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