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Volume 2
     

Clearance of apoptotic cells: Discovery of autoantigens and therapy for autoimmune diseases (APOCLEAR)

   
Project

QLK3-2002-02017

Cell factory area

3.1.3

EU Contribution

2,056,207 Euro

Duration

36 months

Type

RS

Starting date

01 November 2002

Keywords
autoantibodies
cell death
engulfment pathways
phagocytic activity
ABSTRACT

The major goal of this three-year project involving ten academic participants and two biotech companies is to develop novel therapeutic strategies for the treatment of human autoimmune diseases. A defective clearance of apoptotic cells consistently associates in vivo with autoimmunity. Interventions aimed to enhance the phagocytic ability of the organism may therefore limit the accumulation of apoptotic cells and break the vicious circle that maintains autoimmune responses. The project aims to define the interaction among phagocytic partners and to identify molecular interventions capable to influence this pathway, bypassing defective events or targeting defined repressor modules. Participants will share technologies and expertise to develop appropriate in vivo models.

These models will allow the in vivo validation of proposed strategies, aimed to:

  1. prevent tissue accumulation of apoptotic cells;
  2. prevent chronic inflammation and tissue damage;
  3. limit autoimmunity.
OBJECTIVES

The specific project objectives have been divided into three phases and ten work packages. Main objectives of Phase I are the identification of phagocytic partners involved in the clearance of apoptotic/necrotic cells and the characterisation of their interaction. Main objectives of Phase II are the development of novel animal models and the assessment of the in vivo consequences of the impaired clearance of dying cells, with particular attention to effects on acute and chronic inflammation and on the initiation and the maintenance of autoimmunity. Finally, the main objectives of Phase III are the identification of novel autoantigens, a valuable tool for the diagnosis and the prognostic evaluation of human autoimmunity disease and the identification of strategies to reprogram apoptotic cell clearance, directly targeting the clearance machinery characterised in Phases I and II.

DESCRIPTION OF THE WORK

To accomplish the aims of Phase I, we will identify in C. elegans novel genes involved in the clearance of apoptotic/necrotic cells. We will then proceed to define epistatic groups and mammalian orthologs. In parallel, we will identify genes involved in defective clearance of phagocytes from autoimmune patients, and study dynamics of phagocyte cell receptors during recognition and engulfment of apoptotic cells. We will eventually identify functional modules, paralleling epistatic groups defined in C. elegans, and physical interactions among phagocytic partners in mammalian phagocytes, in normal or inflammatory conditions. To accomplish the objectives of Phase II we will validate the involvement of newly identified genes in the clearance of dying cells, developing appropriate mouse models bearing selective gene deletion/inactivation or multiple gene modifications. Complementation groups will be defined in normal or inflamed tissues. To better dissect the functional interactions among partners, clearance defective animals will be crossbred with mice selectively expressing (or down-regulating) relevant genes in monocytes and inflammatory macrophages. In vivo mesangial cell killing, thymocyte development and selection of T-cell repertoire will be evaluated. To accomplish the objectives of Phase III, we will collect autoantibodies that specifically recognise epitopes generated during apoptosis and use them to identify autoantigens implicated in human autoimmune diseases.

Strategies that directly target the clearance machinery will be validated for their ability to:

  1. limit the spontaneous accumulation of cell corpses and ensuing tissue damage and autoimmunity;
  2. limit the noxious effects of acute overload of dying cells.
DELIVERABLES

Main deliverables of Phase I will be new C. elegans mutants that fail to clear apoptotic cells and optimised protocols for characterisation of genes that confer phagocytic activity. Novel genes involved in apoptotic cell clearance by phagocytes will be identified. Libraries of fluorescent-tagged phagocytic partners, of non-phagocytic mammalian cell lines expressing phagocytic partners' genes or expressing all the phagocytic partners that drive a complete engulfment pathway will be established. Mice bearing multiple gene inactivation will be developed.

Main deliverables of Phases II and III will be mice knockout for novel phagocytic partners, transgenic mice over-expressing wild-type or mutant phagocytic partners in selected tissues or cells, including phagocytes and their progeny. Adjuvants and autoantigens will be identified and validated, yielding as deliverables libraries of cell lines expressing fluorescent tagged autoantigens, diagnostic ELISA assays and preclinical protocols for therapies restoring apoptotic cell clearance.

CONSORTIUM
COORDINATOR
 
Patrizia Rovere-Querini
Immunology and Infectious Diseases
H San Raffaele Scientific Institute, DIBIT
20132 Milano, Italy
Tel: +39-02-2643 4864/4694
Fax: +39-02-2643 4786
patrizia.rovere@hsr.it
 
PARTNERS
 


John Savill, Ian Dransfield, Christopher Gregory, Chris Haslett
MRC Centre for Inflammation Research
University of Edinburgh
Edinburgh, EH8 9XD, United Kingdom
Tel: +44-131-536 2238
Fax: +44-131-536 2247
j.savill@ed.ac.uk
id@srv1.med.ed.ac.uk
chris.gregory@ed.ac.uk
c.haslett@ed.ac.uk

Giovanna Chimini
Centre d'Immunologie de Marseille Luminy Case 906
Parc Scientifique de Luminy
13288 Marseille, France
Tel: +33-491-269 404
Fax: +33-491-269 430
chimini@ciml.univ-mrs.fr

Mauro Piacentini
Cell Biology and Electron Microscopy
I.N..M.I.
I.R.C.C.S. "Lazzaro Spallanzani"
00149 Rome, Italy
Tel: +39-06 72594370
Fax: +39-06 2023500
mauro.piacentini@uniroma2.it

Laszlo Fesus
Biochemistry and Molecular Biology
University of Debrecen
Medical and Health Science Center
4012 Debrecen, Hungary
Tel: +36-52-416 432
Fax: +36-52-314 989
fesus@indi.biochem.dote.hu

Michael Hengartner
Institute of Molecular Biology
University of Zürich
8057 Zürich, Switzerland
Tel: +36-52-416 432
Fax: +36-52-314 989
michael.hengartner@molbio.unizh.ch

Catia Traversari
GenEra S.p.A.
20132 Milano, Italy
Tel: +39-02-2643 4706
Fax: +39-02-2643 4668
traversari.catia@hsr.it

Dario Soldateschi
DIESSE - Diagnostica Senese- S.p.A
53035 Monteriggioni (SI), Italy
Tel: +39-0577-587 130
Fax: +39-0577-587 130
dariosoldateschi@diesse.it

Martin Herrmann, Reinhard Voll
Medical Clinic III
Institute for Clinical Immunology and Rheumatology
91054 Erlangen, Germany
Tel: +49-9131-853 6345
Fax: +49-931-853 5776
martin.herrmann@med3.imed.uni-erlangen.de