Limba Românã
 

Project no. 41001/2007 - PNCD II

Title The Identification and Characterization of Brain Cancer Biomarkers Expressed in Serum and Cerebrospinal Fluid Through Advanced Glicomic Methods Based on Mass Spectrometry.
   
Acronym  BRAIN_CAN_GLICOMICS
   
Project abstract

 Brain cancers are a group of severe diseases, usually detected through costly methods, based on imagistic, applied in an advanced stage of the disease and with a reserved prognostic. The most practical alternative is the early detection of tumors in a stage in wich surgical resection is still helpfull, and which should be based on a routine scan of biomarkers before symptoms appear. Therefore, the development of simple techniques and protocols to allow the translation of laboratory research in clinical usefull applications is highly recommended. We propose an interdisciplinary project within PNCD II framework using advanced techniques for mass spectrometry, nanotechnologies and medicine focused on gangliosides and glycosaminoglycans glycomic expressed in brain cancer patients' body fluids. In this project advanced, automatized, chip based high sensitivity methods of MS will be developed to establish the changes in the expression pattern of gangliosides and glycosaminoglycans as biomarkers in early diagnosis, evaluation and discovery of tumors of the brain. High sensitivity analisys through mass spectrometry will include also the identification and characterization of biomarkers through MS fragmenting in multiple stages. Data will be collected, compared and processed about the specific composition of gangliosides and glycosaminoglycans, epimerisation, sequencing, sulphatation patterns and different structure and composition changes as well as the concentration in serum and cerebrospinal fluid of patients diagnosed with cancer compared to healthy subjects. The main preliminary condition of this study is the integration of life science mass spectrometry with microfluids technology within a partnership of excellence in complementary research areas. We are expecting that this integrative system will find a general applicability as a fast, accurate, noninvasive method able for early diagnostic of brain cancer, evaluation of agressivity and treatment response aiming an improvement in the EU health system and outside it.

   
Objectives
The aim of the present project is the development of new glycomic and glycoproteomic methods based on high performance mass spectrometry combined with robotized infusion methods and chip/microfluids technology in order to detect early stage brain cancer biomarkers and to monitor the evolution of the tumor. The developed methods are based on innovative microfluid/MS technologies which will be conceived, tested and for the first time applied to the analisys of glycoconjugates from body fluids like blood and cerebrospinal fluid.
   
Coordinating Institution University of Medicine and Pharmacy Carol Davila Bucharest
   
Project Partners University of Medicine and Pharmacy Carol Davila - Bucharest

National Institute for Research&Development in Electrochemistry and Condensed Matter - Timisoara

University Aurel Vlaicu - Arad

University of Medicine and Pharmacy Victor Babes - Timisoara
   
Contracting Authority  National Centre for Programme Management
   
Project Timespan  36 months
   
Project Phases

Phase I / 2007: The Evaluation of Actual Progress in Brain Cancer Diagnostic Through Imagistic Methods (15.12.2007)

Phase II/2008: Comparative Evaluation of Actual Trends in Brain Cancer Research Through "omics" Methods. Identification/Selection of Cases (Healthy Patients and Donors). (30.10.2008)

Phase III/2009: Obtaining and Preparing of Serum and Cerebrospinal Fluid (CSF) Probes. Isolation and Purification of Complex Glycosaminoglycans(GAG) Mixtures from CSF (30.05.2009)

Phase IV/2009: Isolation and purification of Complex Mixtures of Gangliosides (GG) from Serum and CSF (30.11.2009)

Phase V/2010: Screening and Fragmentation of GAG and GG through chip-nanoelectrospray QTOF MS, HCT MSn and HPLC HCT MS/MS (14.09.2010)

   
Project Director Contact Coordinates

Prof. univ. Leon Zagrean
email: lzagrean @ univermed-cdgm.ro
8 Eroii Sanitari ave., 050474 Bucharest, Romania

   
Results

ISI Published Papers

1. Glycosylation analysis of glycoproteins and proteoglycans using capillary electrophoresis-mass spectrometry strategies
Sabine Amon, Alina D. Zamfir, Andreas Rizzi
Electrophoresis 2008, 29, 2485–2507, DOI 10.1002/elps.200800105 (ISI)

2. High-Throughput Analysis of Gangliosides in Defined Regions of Fetal Brain by Fully Automated Chip-Based Nanoelectrospray Ionization Multistage Mass Spectrometry.
Alina Serb, Catalin Schiopu, Corina Flangea, Željka Vukelic, Eugen Sisu, Nicolae Dinca, Leon Zagrean and Alina D. Zamfir
Eur. J. Mass Spectrom. 15, 541-553, 2009 (ISI)

3. Synthesis and structural characterization of amino-functionalized polysaccharides
I. Sisu, V. Udrescu, C. Flangea, S. Tudor, N. Dinca, L. Rusnac, A.D. Zamfir, E.Sisu
Cent. Eur. J. Chem. 7, 66-73, 2009 (ISI)

4. Fully automated chip-based negative mode nanoelectrospray mass spectrometry of
fructooligosaccharides produced by heterologously expressed levansucrase from Pseudomonas syringae pv. tomato DC3000
T.Visnapuu, A. D. Zamfir, C. Mosoarca, M. D. Stanescu, T.Alamäe
Rapid Commun. Mass Spectrom. 23, 1337–1346, 2009 (ISI, IF=3.05)

5. Analysis of novel over- and undersulfated glycosaminoglycan sequences by enzyme
cleavage and multiple stage mass spectrometry
A.D. Zamfir, C. Flangea, A. Serb, E.Sisu, N. Dinca, P. Bruckner, D. G. Seidler
Proteomics 9, 1-10, 2009 (ISI, IF=5.87)

6. Top-down glycolipidomics: fragmentation analysis of ganglioside oligosaccharide core and
ceramide moiety by chip-nanoelectrospray collision-induced dissociation MS2-MS6
A. Serb, C. Schiopu, C. Flangea, E. Sisu, A.D. Zamfir. J. Mass Spectrom. 2009 (ISI, IF=3.21)

7. Discrimination of GalNAc (4S/6S) sulfation sites in chondroitin sulfate disaccharides by
chip-based nano-electrospray multistage mass spectrometry
C. Flangea, A. Serb, C. Schiopu, S. Tudor, E. Sisu, D. G. Seidler, A. D. Zamfir
Cent. Eur. J. Chem. 2009 (ISI)

8. Qualitative and quantitative analysis of gallic acid in Alchemilla vulgaris, Allium ursinum, Acorus calamus and Solidago virga-aurea by chip-electrospray ionization mass spectrometry and high performance liquid chromatography
Dumitru Condrat, Cristina Mosoarca, Alina D. Zamfir, Florin Crisan, Maria R. Szabo and Alfa X. Lupea
Central European Journal of Chemistry, vol. 3, no. 3, June 2010 (ISI)


Scientific Presentations

1.Screening si Fragmentare Comparativa a Gangliozidelor din Creierul Fetal Uman prin Metode Avansate de Spectrometrie de Masa
Alina F. Serb, Eugen Sisu, Corina Flangea, Alina D. Zamfir, Leon Zagrean,
Conferinta Diaspora in Cercetarea Stiintifica Romaneasca, Bucuresti, 17-19 septembrie 2008

2.Sistem Informatic de Operare pentru Interpretarea Spectrelor de Masa a Gangliozidelor din Creierul Uman
Catalin Schiopu, Eugen Sisu, Sorin Nadaban, Carmen Fifor, Alina Zamfir, Conferinta Diaspora in Cercetarea Stiintifica Romaneasca, Bucuresti, 17-19 septembrie 2008

3. Determination and structural characterization of anencephaly-associated gangliosides by advanced mass spectrometry based on chip electrospray ionization and multistage sequencing. A. D., Zamfir, C. Mosoarca, C.Flangea, Z. Vukelic., E. Sisu
6th Forum of European Neuroscience, 12-16 July 2008 Geneva, Switzerland; FENS Abstr., vol.4, 083.29, 2008

4. Comparative assay on ganglioside differential expression in defined regions of uman brain by modern mass spectrometric methods.
A. F., Serb, C. Flangea, Z.Vukelic, E.Sisu , L.Zagrean, A. Zamfir
6th Forum of European Neuroscience, 12-16 July 2008 Geneva, Switzerland; FENS Abstr., vol.4, 075.11, 2008

5. Gangliozidele prezinta o inalta specificitate filogenetica in creierul fetal uman, demonstrata prin investigatii de spectrometrie de masa nanoESI bazate pe chip
A.F.Serb, E.Sisu, C.Schiopu, A.D. Zamfir, L.Zagrean
Al IV-lea Simpozion al Societatii Nationale de Neurostiinte, 30-31 mai 2008, Bucuresti, Romania;Abstr.Vol. (ISBN: 978-973-708-323-4), p.42

6. Investigation of glycosaminoglycan sulfation pattern by high performance chip electrospray multistage mass spectrometry
C.Flangea, A.F. Serb, S.Tudor, I.B.Pancan, E.Sisu, D.G.Seidler, A.D.Zamfir
International Symposium Research and Education in Innovation Era, 2nd Edition, November 20th - 21st , 2008

7. Computer software for the interpretation brain ganglioside mass spectra
C.Schiopu, E.Sisu, Ž.Vukelic, N.Dinca, S.Nadaban, C.Fifor, A.D.Zamfir
International Symposium Research and Education in Innovation Era, 2nd Edition, November 20th - 21st, 2008

8. Profiling of gangliosides expressed in human hemangioma by fully-automated chip-based nanoelectrospray tandem mass spectrometry
A.Serb, C.Schiopu, C.Flangea, E.Sisu, S.Tudor, I.B.Pancan,Ž. Vukelic, A.D. Zamfir
International Symposium Research and Education in Innovation Era, 2nd Edition, November 20th - 21st, 2008

9. Potentials of a fully automated chip-based nanoelectrospray ionization high capacity trap mass spectrometry in neuroglycolipidomics: characterization of ganglioside composition
specificity in selected fetal human brain regions
Ž.Vukelic, D. Marincic, A.Serb, A. D. Zamfir
Invited lecture: Austrian Mass Spectrometry Forum, Feb. 17-19, 2009, Vienna, Austria

10. Novel saccharide sequence analysis of glycosaminoglycans by enzyme cleavage and multistage MS: unusual exuronic acid sulfation in decorin
C. Flangea, C. Schiopu, E. Sisu, N. Dinca, D. G. Seidler, A. D. Zamfir
Oral presentation: Austrian Mass Spectrometry Forum, Feb. 17-19, 2009, Vienna, Austria

11. A new Mass Spectrometry Society – Perspectives and plans
A.D. Zamfir
Invited lecture: Anniversary Conference, 20 years of Analytical Chemistry in Konstanz,
March 5-7, 2009, Konstanz, Germany

12. Chip-based nanoelectrospray multistage mass spectrometry in neuroglycolipidomics: high-throughput analysis of gangliosides in defined regions of fetal brain
A.F. Serb, C. Schiopu, D. Marincic, Ž. Vukelic, A.D. Zamfir
Oral presentation: 42nd German Society for Mass Spectrometry Conference, Konstanz,
Germany, March 8-11, 2009

13. Investigation of novel under- and over-sulfated regions in decorin glycosaminoglycans by electrospray ionization collision-induced dissociation multistage mass spectrometry
C. Flangea, E. Sisu, N. Dinca, D.G. Seidler, A. D. Zamfir
Poster presentation: 42nd German Society for Mass Spectrometry Conference, Konstanz,
Germany, March 8-11, 2009

14. A novel procedure for derivatization of long chain polysaccharides to enhance chip-based nanoelectrospray tandem mass spectrometric analysis
C. Flangea, I.Sisu, N. Dinca, E. Sisu, A. D. Zamfir
Poster presentation: 42nd German Society for Mass Spectrometry Conference, Konstanz,
Germany, March 8-11, 2009

15. Chip-Based nanoelectrospray high capacity ion trap mass spectrometry of gangliosides from precentral vs. postcentral gyrus of fetal human brain
D.Marincic, Ž.Vukelic, C. Schiopu, M. Kos, A. D. Zamfir
Poster presentation: 42nd German Society for Mass Spectrometry Conference, Konstanz,
Germany, March 8-11, 2009

16. Screening and sequencing of fructooligosaccharides produced by heterologously expressed levansucrase from Pseudomonas syringae pv. tomato DC3000 by fully automated chip based mass spectrometry
C. Mosoarca, T. Visnapuu, T. Alamäe, A. D. Zamfir
Poster presentation: 42nd German Society for Mass Spectrometry Conference, Konstanz,
Germany, March 8-11, 2009

17. High-throughput mass spectrometric determination of ganglioside expression and structure in human fetal cerebellum
R. M. Ghiulai, C. Mosoarca, C. Novaconi, D. Marincic, Ž. Vukelic, A. D. Zamfir
Poster presentation: 42nd German Society for Mass Spectrometry Conference, Konstanz,
Germany, March 8-11, 2009

18. Chip-based mass spectrometry for glycomics and glycoproteomics
A. D. Zamfir
Invited lecture: University of Konstanz, May 13, 2009

19. Analysis of brain chondroitin/dermatan sulfate glycosaminoglycans by fully automated chip-based nanoelectrospray multistage mass spectrometry
C. Flangea, E. Sisu, D. G. Seidler, A. D. Zamfir
Poster presentation: 57th American Society for Mass Spectrometry Conference on Mass Spectrometry and Allied Topics, May 31-June 6, 2009, Philadelphia, USA

20. Compositional and structural analysis of gangliosides in human cerebrospinal fluid by chip-based nanoelectrospray ionization tandem mass spectrometry
A. Serb, C. Schiopu, D. Marincic, Ž. Vukelic, A. D. Zamfir
Poster presentation: 57th American Society for Mass Spectrometry Conference on Mass Spectrometry and Allied Topics, May 31-June 6, 2009, Philadelphia, USA

21. Combining fully-automated chip-nanoelectrospray ion trap mass spectrometry and GanglioSoft 1.2 computer software for identification of human hemangioma gangliosides
C.Schiopu, A. Serb, E. Sisu, Ž. Vukelic, A. D. Zamfir
Poster presentation: 57th American Society for Mass Spectrometry Conference on Mass
Spectrometry and Allied Topics, May 31-June 6, 2009, Philadelphia, USA