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Molecular Genetics, Cellular Immunology and Peptide-Chemistry for the Development on Control Tools Against Leishmaniosis

Objective

There are three main objectives:

* The study of the biological function and role in the Leishmania infectivity of two molecules encoded by developmentally regulated Leishmania genes, namely a putative nuclear growth related factor (LNP18) and an immunodominant membrane protein.
* The dissection of the cellular immune responses conferring resistance to Leishmania spp. driven pathogenic processes.
* Development of new reliable diagnostic products for early detection of leishmaniosis.

This project aims at identifying parasite factors essential for Leishmania infectivity and at dissecting cellular immune responses conferring resistance to Leishmania spp.-driven pathogenic processes. Such advances should pave the way for the development of new strategies and the rational design of future immunoprophylactic and immunotherapeutic measures to control leishmaniosis. Leishmaniosis constitutes not only a major public health problem of socio-economic importance but is also a veterinary problem in various parts of the world and especially in Mediterranean countries. Thus, there is a need for the development of new effective tools for their control. Recent advances in the field of molecular genetics, cellular immunological tools that the European partners have recently developed for characterizing and studying parasite and host factors promoting or circumventing the development of the disease. The rational approach that will be pursued is based on the use of all these tools focusing on two Leishmania molecules, a putative nuclear growth related factor (LNP18) and a B-cell immunodominant antigen (P32) that induces also T cell responses encoded by two novel genes recently identified by partners no 2 and no 4 respectively, for the elucidation of (a) their functions in pathogenesis (b) the nature of molecular and cellular effectors/regulators on which they act as sites of Leishmania delivery or spread (c) their impact on the selective activation and differentiation of the CD4+ and CD8+ T-cell subsets to drive a protective Th1-cell response and (d) their potential use as reagents for diagnosis. The immunogenic potential of the different Leishmania molecule formulations will be studied in the murine model and the recently established by partners no 3 and no 5, canine models. Such understanding will allow the design of rational approaches to combat leishmaniosis.
Expected Outcome

Following completion of the project, the results obtained will be sufficiently robust for the industrial environment and will provide new strategies and tools to help the leishmaniosis control such as: Leishmania mutants and live recombinant BCG expressing Leishmania antigens, as possible vaccine candidates, host-protective parasitic synthetic peptides and recombinant molecules as well as particulate antigens, and reliable diagnostic tools.
In particular, we propose to:

* Develop Leishmania mutants by sequential targeted gene replacement or site directed mutagenesis.
* Determine the phenotype conferred by LNP18 and P32 mutations.
* Evaluate the role of LNP18 and P32 in parasite infectivity.

The study will:

* Investigate the host-protective capability of defined Leishmania antigens. New protective antigens and two B immunodominant molecules LNP18 and P32 will be identified, cloned, sequenced and characterized, using recombinant molecules as well as synthetic peptides modelled from their respective amino-acid sequences.

* Determine ways and parameters of directing the immune system towards a Th1 type response, by examining both CD8+ and CD4+ lymphocyte responses, using vectors such as live recombinant BCG expressing Leishmania antigens e.g. LNP18 and P32, and synthetic microspheres carrying these antigens.
* Conduct protection studies in canine experimental models.

More specifically, we will evaluate the diagnostic potential of:

* recombinant LNP18 and P32
* synthetic peptides modelled from P32 and LNP18 amino acid sequences, as targets for ELISA. Data on P32 and on a LNP18 20-peptide make these molecules promising reagents for serodiagnosis.

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

INSTITUT PASTEUR
Address
25,Rue Du Docteur Roux 28
75724 Paris
France

Participants (4)

Hellenic Pasteur Institute
Greece
Address
127,Vassilissis Sofias Avenue
11521 Athens
INSTITUT AGRONOMIQUE ET VETERINAIRE HASSAN II
Morocco
Address
10101,Avenue Allal El Fassi, Rabat Institutes
10101 Rabat / Agdal
Institut Pasteur de Tunis
Tunisia
Address
13,Place Pasteur
1002 Tunis
MICROFAR - Associaçao para o Desenvolvimento de Vacinas e Produtos Imunologicos
Portugal
Address
Avenida Das Forças Armadas
1600 Lisboa