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Projects / Programmes source: ARIS

Inhibitors of cysteine carboxypeptidases as regulators of autoimmune and neurodegenerative processes

Research activity

Code Science Field Subfield
4.06.01  Biotechnical sciences  Biotechnology  Recombinant DNA technology 

Code Science Field
T490  Technological sciences  Biotechnology 

Code Science Field
1.07  Natural Sciences  Other natural sciences 
Keywords
Cathepsin X, Gamma-enolase,LFA-1, Integrin, Protease inhibitor, Psoriasis, Neurodegenerative diseases
Evaluation (rules)
source: COBISS
Researchers (29)
no. Code Name and surname Research area Role Period No. of publicationsNo. of publications
1.  24257  PhD Aleš Berlec  Pharmacy  Researcher  2011  217 
2.  15099  PhD Darko Černe  Pharmacy  Researcher  2012  261 
3.  15284  PhD Stanislav Gobec  Pharmacy  Researcher  2011  827 
4.  20376  Branka Grabovac    Technical associate  2011 - 2014 
5.  12688  PhD Kristina Gruden  Biotechnology  Researcher  2012 - 2014  983 
6.  01302  PhD Matjaž Jeras  Biotechnology  Researcher  2011 - 2014  360 
7.  26497  PhD Zala Jevnikar  Pharmacy  Researcher  2011  95 
8.  24402  PhD Petra Kocbek  Pharmacy  Researcher  2011  291 
9.  04648  PhD Janko Kos  Biotechnical sciences  Head  2011 - 2014  1,154 
10.  29604  PhD Bojana Kranjc Mirković  Pharmacy  Researcher  2011 - 2012  67 
11.  30881  PhD Špela Magister  Biotechnology  Researcher  2011 - 2012  24 
12.  23487  PhD Sara Mankoč Ramuš  Cardiovascular system  Researcher  2011 - 2014  73 
13.  19170  PhD Urša Pečar Fonović  Pharmacy  Researcher  2011 - 2014  144 
14.  29966  Dejan Pelko    Technical associate  2011 - 2014 
15.  36596  PhD Milica Perišić Nanut  Biotechnical sciences  Researcher  2013 - 2014  141 
16.  32035  PhD Anja Pišlar  Biotechnology  Researcher  2011 - 2014  162 
17.  26507  PhD Jure Pohleven  Biochemistry and molecular biology  Researcher  2011 - 2013  75 
18.  22657  PhD Irena Prodan Žitnik  Pharmacy  Researcher  2012  48 
19.  15813  PhD Boris Rogelj  Neurobiology  Researcher  2011 - 2014  410 
20.  31397  Matija Rojnik  Pharmacy  Researcher  2011  43 
21.  23576  PhD Jerica Sabotič  Biochemistry and molecular biology  Researcher  2011 - 2014  315 
22.  19786  Majda Sirnik    Technical associate  2012 
23.  36900  Jasna Šlenc  Pharmacy  Researcher  2014 
24.  07849  PhD Borut Štrukelj  Biochemistry and molecular biology  Researcher  2011  1,120 
25.  26198  PhD Urban Švajger  Microbiology and immunology  Researcher  2011 - 2014  202 
26.  21619  PhD Olga Vasiljeva  Oncology  Researcher  2011 - 2014  183 
27.  28905  PhD Nace Zidar  Pharmacy  Researcher  2012  215 
28.  08289  PhD Marko Živin  Neurobiology  Researcher  2011 - 2014  232 
29.  17285  Darja Žunič Kotar    Technical associate  2011 - 2014 
Organisations (4)
no. Code Research organisation City Registration number No. of publicationsNo. of publications
1.  0106  Jožef Stefan Institute  Ljubljana  5051606000  89,990 
2.  0311  Blood Transfusion Centre of Slovenia  Ljubljana  5053960  1,752 
3.  0381  University of Ljubljana, Faculty of Medicine  Ljubljana  1627066  46,179 
4.  0787  University of Ljubljana, Faculty of Pharmacy  Ljubljana  1626973  17,522 
Abstract
We propose that inhibitors of cysteine carboxypeptidase cathepsin X reduce autoimmune and neurodegenerative processes as a result of the specific role this carboxypeptidase has in regulating integrin receptor LFA-1 and g-enolase. Cathepsin X is a lysosomal cysteine protease localized predominantly in immune and neuronal cells. We discovered recently that it may regulate the activity of lymphocyte function-associated antigen-1 (LFA-1), the predominant beta-2 integrin receptor in T cells that regulates T cell signalling, cytoskeletal remodelling, migration, the growth of cellular extensions like uropods and nanotubes, and formation of the immunological synapse. Excess activity of LFA-1 leads to autoimmune diseases such as psoriasis. Because the success of treatment of these diseases with existing drugs and protocols is very limited, LFA-1 is considered as an important and promising therapeutic target for new drug development.  Several drugs have been developed to impair LFA-1 function, however, they are not appropriate for the therapy of autoimmune diseases due to insufficient selectivity against the harmful functions of LFA-1. General inhibition of LFA-1 activity may cause severe side effects, resulting in lethal infections in treated patients, as shown for efalizumab (Raptiva, Genentech), the first effective drug for treating psoriasis, which was recently withdrawn from the market. Our results show that cathepsin X enables fine regulation of LFA-1 activity by gradual degradation of the cytoplasmic tail of beta 2 chain of LFA-1, the part of the molecule that defines LFA-1 activity. In this way cathepsin X modulates the affinity of LFA-1 for some of its ligands, however, inhibition of cathepsin X does not completely impair LFA-1 activity, as is the case with efalizumab. This can be reflected in significantly reduced side effects. In neuronal cells, g-enolase is a target for cathepsin X. It is a glycolytic enzyme with additional functions in the proliferation of neuronal cells and neuritogenesis. Cathepsin X cleaves the C-terminal end of g-enolase, impairing the neurotrophic function of the latter. The cleavage prevents the binding of g-enolase to g1-syntrophin, a scaffold protein which is responsible for the translocation of g-enolase to the plasma membrane, a key step in its neurotrophic function. We recently demonstrated that inhibition of cathepsin X prevents the cleavage of g-enolase and increases the proliferation of neuronal cells and neuritogenesis, events which are very important for the treatment and prevention of neurodegenerative diseases. In this project we will use different approaches to select inhibitors of cathepsin X that are specific for the enzyme in targeted cells and non-toxic for other cells and tissues. Their selectivity and controlled release will be obtained by using delivery systems such as biodegradable polymeric nanoparticles. We will develop in vitro cell models using keratinocytes, T cells, neuronal cells and cells of microglia to resemble the pathological processes of psoriasis and neurodegenerative diseases. These models will be used to test the selected inhibitors and delivery systems. The most promising anti-cathepsin X compounds and formulations will be tested in vivo for their pharmacological properties.
Significance for science
The research in the proposed project is part of the advanced trends in science in the fields of proteolytic enzymes and processes of the immune response. Recent results obtained by our group on the regulation of integrin receptors in leukocytes, have opened up a new field of research, which has met with a wide and positive response in the scientific community, evidenced by multiple citations in journals with the highest impact factors. Further, some groups are continuing with the research, based on our results. Knowing the mechanisms of regulation of integrin receptors has enabled the basic processes in immune cells to be understood and may well explain the events leading to autoimmune diseases such as psoriasis. Our studies will define the role of cathepsin X in immune processes. This is very important if this protease is to be used as the target for the development of new drugs. Only selective inhibition of a particular protease that is truly involved in pathological processes, can lead to progress in therapy. The use of nonselective general protease inhibitors has been compromised, causing serious site effects. The therapeutic application of drugs that totally block the action of integrin receptors in immune cells is also inadequate, and some of them have had to be withdrawn from clinical practice. As an original scientific contribution we can draw attention to the newly revealed mechanism of action of cathepsin X on g-enolase and to the role of both proteins in the processes of neurodegeneration as well as the role of cathepsin X inhibitors and their ability to potentiate neurotrophic activity of g-enolase. Understanding the mechanisms of degeneration of neuronal cells and identification of molecular targets involved in these processes provides the essential basis for the development of new, effective drugs that are essential for treating neurodegenerative diseases. Within the proposed project we expect to prove that our approach of selective inhibition of integrin receptors in T cells with protease inhibitors enables fine regulation of their activity and reduces harmful site effects. The successful use of polymeric delivery systems will further limit the action of new drugs to cells that are involved in the pathological processes. Our project group will use the most recent techniques and methods that have already been introduced into laboratory practice. The collaboration with leading researchers in Slovenia and worldwide will guarantee rapid transfer of knowledge, good science and successful implementation of the project.
Significance for the country
ANG Medical treatment, health care and prevention are important elements in the socio-economic development of modern society, and every step towards better health is important. In particular, new knowledge and new drugs are essential for treating diseases where existing therapies are not effective, for example in autoimmune diseases and neurodegenerative diseases. The contribution of the proposed project was very important also for all levels of university education. It enabled students to acquire new knowledge and the project’s implementation provided opportunities for diploma and doctoral training in the fields of pharmaceutical biochemistry and biotechnology, since the members of the project team are involved in the pedagogical process at the University of Ljubljana and Postgraduate School Jožef Stefan, and as visiting professors at other universities. The results of this project will also foster our collaboration with other research groups, increasing knowledge exchange and the opportunity to obtain new European and other funds for financing our science.
Most important scientific results Annual report 2011, 2012, 2013, final report, complete report on dLib.si
Most important socioeconomically and culturally relevant results Annual report 2011, 2012, 2013, final report, complete report on dLib.si
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