Molecular mechanisms of regulation of cellular processes related to some human diseaes

Molecular mechanisms implicated in regulation of cellular processes are crucial for maintainance of homeostasis, defence against environmental or endogenous stressors  and damage repair. We will combine basic research on isolated enzymes, model organisms, cell lines and human tissues with applied research and clinical studies with a goal of better understanding of molecular mechanisms that regulate these cellular processes in health and disease. The translational aspect of our programme is to find new potential diagnostic and prognostic biomarkers, identify and investigate novel targets for treatment and pharmacogenomic markers of treatment response that will support individualized medicine. Isolated and naturally occuring enzymes represent potential novel treatment targets. We will develop methods for high throughput evaluation of putative inhibitors of cholinesterases (AChE and BChE), that will be applicable on purified recombinant enzymes and enzymes naturally occuring in sera from healthy individuals and AD patients. Exo- and endogenous delivery systems such as cell-penetrating peptides (CPP) and exosomes will be studied. As penetration of CPP perturbes the membrane and this leads to side effects, we will study interactions of CPPs from the pepfect family with model membranes in terms of membrane structure and transported cargo (siRNA) to develop safer CPPs. Exosomes area unique endogenous delivery system of proteins, lipids and microRNAs and are extensively studied as biomarkers for several diseases. We will study exosomes and miRNA in CSF and plasma samples to assess the clinical relevance of exosomes as biomarkers of encephalopathy in AIDS and of neurodegeneration in Alzheimer's disease. Eukaryotic organisms isolated from hypersaline environments H.werneckii (extremely adaptable), W.ichthiophaga (obligate halophile) and A.pullulans  (moderately halotolerant) are our model organisms to study signal transduction pathways involved in stress response.  We have recently obtained whole genome sequences of these organisms. Comparative studies of the available genomes, functional studies with knock out strains and development of appropriate mathematical models will yield deeper insights into regulatory networks and adaptive strategies of these organisms to stressful conditions. Endometrial cancer and endometriosis are associated with disturbed estrogen and progesterone metabolism and signalling.  Detailed investigation of these processes in model cell lines and respective tissues will lead to potential novel drug targets and novel biomarkers applicable for early diagnosis. Pharmacogenetic studies of rheumatoid arthritis (RA), childhood ALL and mesothelioma will be continued by building respective clinical-pharmacogenetic models, their validation in prospective studies and independent cohorts and the incorporation of the relevant ones into clinically usefull decision-making algorithms that will support individualized medicine.

Designed as a continuation of the current programme, the proposed programme keeps the core of the most relevant topics, where our results already made a major scentific  impact and upgrades it with up-to-date methodologies and new research topics that are of high scientific relevance and will lead to better understanding of molecular processes in health and disease. Better understanding of enzymes catalytic and structure-function properties and developed high throughput methods for evaluation of a large number of putative inhibitors may have important toxicological or clinical implications. The studies of cell penetrating peptides have already resulted in a successful patent. Exosome and miRNA research represents a new research topic that can be extended both to basic research of different molecular processes and applied in translational research for better uderstanding of disease processes. Genomic, transcriptomic, proteomic and pharmacogenomic approaches will contribute to better understanding of basic mechanisms implicated in molecular and cellular processes on several levels.  In collaboration of several research groups and institutions, genomes of H. werneckii , A. pullulans and W. ichthyophaga were recently sequenced and our study is the first to use these genome/transcriptome data for comparative analysis of signal transduction pathways in stresfull conditions. The knockouts of halophillic fungi that we plan to prepare have not been described so far. The research of hormone dependent disesase and cancer will expand the knowledge in the field of molecular endocrinology, especially with regards to novel mechanisms of steroid hormones action and contribute to identification of novel drug targets and development of non-invasive assays for early diagnosis and individualized treatment. Research will be performed in collaboration with the leading groups in the field of molecular endocrinology of hormone-dependent diseases, and will transfer some state-of-the-art methodologies to our laboratories. In the field of biomarker research and pharmacogenetics, our expertise and results support individualized medicine and may establish a basis for a future university spin off company which may provide new employment opportunities for Slovenian postdoctoral researchers. In summary, the set course of our reserch will lead to better understanding of basic molecular processes and identification of new targets for new approaches to treatment and novel molecular biomarkers that may serve as diagnostic and prognostic factors for personalized medicine. Besides high scientific output per unit FTEhigh number of full professors (7) in the programme group, a large number of completed PhD, MSc and diploma theses and six patent applications resulting from several research topics in the last five years indicate that the continuation of our research has good potential to lead to original and relevant findings.

The fact that 7 researchers in the programme group are full professors, speaks for our outstanding achievements in terms of research excellence, and teaching and professional quality. We lead several core and elective courses in the field of biochemistry and molecular biology at the undergraduate and post-graduate studies and mentored 18 PhD and 3 MSc theses and comentored 6 PhD theses  in the last 5 years.  We also mentored 28 and comentored 17 graduation theses. Considering the fact that students at the Faculty of Medicine do not write graduation theses, these numbers show that we also actively participate in teaching at other faculties. Thus we significantly contribute to the quality of the undergraduate and graduate education at the University of Ljubljana, as well as to the development of the field of biochemistry and molecular biology in Slovenia. When taking into account our full-time teaching load, our programme group achieved outstanding results per FTE unit. In addition to the development of higher education, the program has a significant impact on the international recognition of Slovenia. As the studied halophilic organisms were first isolated in Sečovlje salterns, Slovenian researchers in this field achieved unprecedented international recognition, which is very important for the promotion of Slovenia. The results of studies on the genome and / or transcriptome of these organisms will undoubtedly lead to new patent applications and may also have a great importance for the economy, because organisms with enhanced tolerance to salt are interesting for bioremediation, biotechnology and agronomy. Our programme will also have a big impact on health care. Translation of basic research to clinical applications constituts an important part of our programme. In the course of our clinical studies in the field of hormone-related disorders, cancer and rheumatoid arthritis, we have identified potential new potential diagnostic and prognostic markers, new targets for treatment as well as pharmacogenetic markers of response to treatment. When supported by replication and multicenter studies, our research will support the transition from the current treatments, based on the characteristics of the total patient population, to the predictive, preventive and personalized medicine. In addition to more effective treatment, which means big savings for health care system, also patients' quality of life of will significantly improve.