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Title: Expert Network to study of the genetic origin and molecular pathogenesis of
corneal dystrophies
Integrated international research to study molecular pathogenesis of corneal dystrophies and its
genetic origin
ACRONYM: CORNEAGEN
Quality of Life
ID= 1155582
UNAME= NJ1155582N1
PASSWORD= NJ1155582N1
Memorandum of the 1st meeting in Budapest January 12, 2002
Participating Organizations:
CS 01 University of Debrecen, Faculty of Medicine Department of Ophthalmology,
Hungary
Short name: UDFMDO
Scientific coordination
Prof. Andras Berta, M.D.
Dept. of Ophthalmol.,
Medical and Health Sciences Center,
University of Debrecen,
Nagyerdei krt. 98., PO box: 30.
H-4012 DEBRECEN, HUNGARY
Tel.: +36 30 229 5889
Tel./Fax.: +36 52 415 816
Berta, Andra [email protected]
Modis, Laszlo [email protected]
User Name is : CORNEAGEN2
Password is : shu407
CO 02 VARIMED Ltd., Hungary (manager)
Short name: VARIMED
Project and database management
Vari, Sandor [email protected]
VARIMED Ltd.
Budapest, Szalanci u 5, H-1124
Telephone: +36-1-487-0429
Fax: +36-1-487-0430
AC03 VITAMIB SARL. France
Short name: VITAMIB
eForms, electronic database
Xavier Fabre
[email protected]
User Name is : CORNEAGEN10
Password is : dze530
CR04 University of Erlangen, Department of Ophthalmology, Germany
Short name: UEDO
Naumann G.O.H., MD
Professor and Chairman, Department of Ophthalmology
Friedrich-Alexander-Universitat, Erlangen-Nürnberg,
Schwabachanlage 6 (Kopfklinikum)
D-91054 ERLANGEN/GERMANY
Tel. (09131) 8534477
FAX (09131) 8536435
[email protected]
Dr. Berthold Seitz
Fax (49) 9131 853 4436
[email protected]
User Name is : CORNEAGEN4
Password is : mnae459
CR05 Community Hospital Århus, Department of Ophthalmology, Denmark
Short name: COHADOP
Professor Dr. Niels Ehlers
Community Hospital Århus
Norrebrogade 44
DK-8000 Arhus C
Phone: (45) 8949 3237
Fax: (45) 8949 3230
[email protected]
User Name is : CORNEAGEN5
Password is : khe378
CR06 University of Helsinki , Central Hospital, Finland
Short Name: UHCH
Genome typing
Haartmaninkatu 4C, FIN-00029, HYKS, Finland.
Tervo, Timo, [email protected]
Juha Holopainen [email protected]
Tiina Alitalo [email protected]
User Name is : CORNEAGEN7
Password is : plo259
CR07 Hadassah University Hospital, Department of Ophthalmology, Israel
Short name: HUHDO
Professor Dr. J. Frucht-Pery
Hadassah University Hospital,
Department of Ophthalmology, Israel
P.O. Box 12000,
91120 Jerusalem, Israel,
Fax: (972) 2 6428896
[email protected]
User Name is : CORNEAGEN8
Password is : hmo2001
CR08 Lublin University School Of Medicine (Medical Academy), Ophthalmology
& 1st Eye Hospital
Short name: LUSMOEH
Professor Zbigniew Zagórski, M.D., Professor and Chairman
Tadeusz Krwawicz Chair of Ophthalmology & 1st Eye Hospital
Lublin University School Of Medicine (Medical Academy)
ul. Chmielna 1, 20-079 Lublin, Poland,
Tel./Fax: (48) 81-5324827
[email protected]
User Name is: CORNEAGEN11
Password is: shou434
CR09 Slovak Postgradual Academy of Medicine, Ophthalmic Department
Short name: SPAOP
Prof Dr. Andrej Cernak
Ophthalmic Department
Slovak Postgradual Academy of Medicine
St. Cyril and Method Hospital
Antolská 11, 851 07 Bratislava, Slovakia
Tel: (421) 76867 2039
Fax: (421) 76381 2145
[email protected]
User Name is : CORNEAGEN12
Password is : shou721
CR10 Charles University, Department of Ophthalmology, Lexum Laser Excimer
Centrum
Short name: CUDO-LEXUM
Doc. MUDr. Martin Filipec CSc.
Lexum Laser Excimer Centrum
Valčikova 1950/3
Praha 8 Liben
Tel/Fax: (420) 026884240
Czech Republic
[email protected]
User Name is : CORNEAGEN13
Password is : kle232
CR10 Cedars-Sinai Medical Center, Ophthalmology Research Department of
Surgery
Existing NIH Grant is the matching activities and fund
Short name: CSMCORD
Burns & Allen Research Institute
8700 Beverly Blvd.
Los Angeles, CA 90048-750
Kenney, Cristina [email protected]
Nesburn, Antony [email protected]
Cc: Daniel Oshiro [email protected]
User Name is : CORNEAGEN9
Password is : bli165
List of the representatives attended of the meeting:
Prof. Andras Berta, MD
Dr. Laszlo Modis, MD
Dr. Tiina Alitalo, PhD
Dr. Sandor G. Vari, MD
Agenda;
FORMATION OF CONSORTIUM
1.
Participants of the meeting agreed to form the CORNEAGEN Consortium, simce
the Scientific Coordinator of the project received confirmation of interest from the
partners listed above.
2.
Partners agreed to make a registration on the CORDIS site as a potential EU
project (see summary for CORDIS)
3.
Partners agreed to cancel the submission of a proposal on February 8, 2002 QoL
14. Support for research infrastructures.
4.
Partners agreed to focus on the EU Sixth Framework Expert Networks and
organize such a network until November 2002
5.
Next meeting will be organized in April or May, 2002 to finalize the WPs, the
role and contribution of the partners.
Summary for Cordis WEB:
The scientific co-ordinator of the project began to investigate the molecular pathogenesis
of type I lattice (Haab-Dimmer) dystrophy, which is one of the 5q31 related corneal
dystrophies in 1995. The purpose of their former studies was the identification of the
amyloid precursor in LCDI and its biochemical characterization. They also carried out
immunohistochemical investigations in scarring and keratoconus corneas to better
understand the role of mutations in the cornea. In their biochemical investigations, they
showed a special protein in the LCDI corneas, the N-terminal sequence of which showed
100% identity with the mutated protein. They also showed the presence of mutated
protein in the corneal deposits by immunohistochemistry. In scarring corneas, they found
increased beta IG-H3 (kerato-epithelin) level at the sites of scarring. In keratoconus
corneas the immunohistochemically detectable beta IG-H3 amount was less than normal,
however, it increased when scarring was also present in the specimens. Partners plan to
bring together a collaborative group or called thematic network of investigators to further
study the molecular genetic basis, immunohistological and biochemical characteristics of
5q31 related (and possibly other) corneal dystrophies.
1. Investigation of the DNA mutations occurring in the subjected region
2. Investigation of the effect of these mutations on the turnover of beta IG-H3 (keratoepithelin) protein and the development of the protein deposits.
3.
Definition of potential treatment modalities.
Possible inhibition of enzymes involved into the pathological degradation products.
Possible replenishment of missing enzymes for dissolution of pathological deposits.
Key words:
0150 Databases, Database Management, Data Mining
0152 Decision Support Tools
0217 European Integration
0263 Genetics
0264 Genomes, Genomics
Project Goals:
The scientific co-ordinator of the project in 1995 began to investigate the molecular
pathogenesis of type I lattice (Haab-Dimmer) dystrophy, which is one of the BIGH3
(5q31) related corneal dystrophies. The purpose of their former studies was the
identification of the amyloid precursor in LCDI and its biochemical characterization.
They also carried out immunohistochemical investigations in scarring and keratoconus
corneas to better understand the role of BIGH3 (kerato-epithelin) protein in the cornea. In
the biochemical investigations, they showed a 42 kD protein in the LCDI corneas, the Nterminal sequence of which showed 100% identity with BIGH3. They also showed the
presence of BIGH3 protein in the corneal deposits by immunohistochemistry. In scarring
corneas, they found increased BIGH3 level at the sites of scarring.
The immunohistochemically detectable BIGH3 amount was less than normal in
keratoconus corneas, however, it increased when scarring was also present in the
specimens. In the future, they would like to continue their investigations in two main
directions:
1. Investigation of the DNA mutations occurring in Central Europe
3. Investigation of the effect of these mutations on the turnover of BIGH3 protein and
the development of amyloid mutations.
Ad 1.: At present, at least eighteen different mutations are known in the BIGH3 gene, and
lattice type IIIA and IV dystrophies were shown to be BIGH3 related dystrophies as well.
The mutations occurring in Central-Eastern Europe have not been studied yet. First, we
would like to analyse the mutations occurring in Hungary, which are most probably
similar to those occurring in other parts of Europe. We began to Collect DNA samples
from the dystrophic patients and their family members. We plan to analyse these samples
by PCR amplification, followed by direct automated DNA sequencing. After finding the
specific mutations, simpler tests, such as PCR-RFLP or PCR-SSCP may be used.
Ad 2.: Investigation of the changes in protein metabolism caused by the mutations is a
much more complicated task. The corneal button excised during keratoplasty is too small
for extensive biochemical studies and does not allow in vivo investigations. To overcome
this problem, we plan to establish immortalized epithelial cell lines from dystrophic and
normal corneas. The immortalization is done with viral or plasmid vectors, containing
SV40 viral sequences. The establishment of such immortalized cell lines is going on at
present in our laboratory. We expect to establish immortalized epithelial cell lines from
both normal and dystrophic corneas in one or two years. These immortalized cell lines
seem to be suitable model systems for the investigation of the pathogenesis of these
dystrophies. They will be helpful in determining the proteolytic steps involved in the
metabolism of BIGH3 and identifying the site and the enzymes of proteolysis. On the
long term, these studies may help the development of drugs being able to inhibit the
accumulation of abnormal protein end-products.
Partners plan to bring together a collaborative group called “thematic network” of
investigators to further study the molecular genetic basis, immunohistological and
biochemical characteristics of corneal dystrophies, with special emphasis on the beta IGH3 (kerato-epithelin) related dystrophies and keratoconus. This project description may
be fitted to the study of other corneal dystrophies (macular, Meesmann, etc.) as well, if
other partners have more intention or experience in the investigation of these diseases
Workplan:
WP01 Project Coordination and Management
Project Management:
The participating partners agreed that the time until February 08, 2002 is too short, the
Consortium could not meet the challenges of the “Quality of Life and Management of
Living Resources Programme, Support for Research Infrastructures”. Partners will go for
the next meeting if we have positive answers from most of the partners no later then
February 15, 2002, and based on the information provided by the partners of the
Consortium we are planning the next meeting in April or May 2002.
WP02 Sample collection
Blood
all patients diagnosed with dystrophy and available family members
Do you have in-house or institutional collaborators to extract
DNA from these samples?
.
Workplan:
The investigation of DNA mutations in the 5q31 chromosome region
Aim: to better understand the role of BIGH-3 in the following corneal dystrophies:
1.
2.
3.
4.
5.
lattice types I, III, IIIa and IV
Groenouw I
Avellino
Reis-Bucklers
since some earlier data showed that beta IG-H3 and related proteins may be
involved in the pathogenesis of keratoconus we plan to further investigate the
relationship of keratoconus and beta IG-H3.
Cornea
all patients who went through keratoplasty
The samples should be cut into two parts:
one for tissue culture
one for frozen section histopathology (Deep frozen –70C to section
for immunohistochemical staining) and/or biochemical
investigations
Mutation detection:
Automated DNA sequencing for the patient.
Can you have the DNA extracted from the blood sample sequenced with
an automated DNA sequencer?
Simplified tests, such as PCR-RFLP or PCR-SSCP for mapping the family
(Benefit to investigate penetrance and expressivety of gene.)
Are these simple tests available to study of family members in
your institution?
Tissue culture of dystrophic corneal cells:
Can you establish immortalized epithelial cell lines from
dystrophic and normal corneas?
A.
Phenotype characterization by complex biochemical methods:
Identification of enzymes involved in the degradation process and detection of
defects in the process
B.
Biochemical analyses of the pathological deposits (amyloid, hyalin
ect.):
Identification of abnormal molecular products using protein sequencing and
linking the abnormality to the morphological and functional defect
Development of a European electronic database for genotypes and phenotypes from
eForm for mutation (HTML and XML)
eForm for clinical and histological data
Correlation analysis of European Database (genotypes and phenotypes):
Potential treatment modalities:
Possible inhibition of enzymes involved in the pathological degradation products.
Possible replenishment of missing enzymes for dissolution of pathological deposits.