Genetically modified microorganism, especially phage, useful for screening for antigenic peptides recognized by T cells

• Main Authors: PIQUERAS BERNARD, LE DOUSSAL JEAN MARC, GOROCHOV GUY, DEBRE PATRICE
• Format: Patent
• Language: English; French
• Published: 19.11.1999
• Edition: 6
• Subjects: BEER; BIOCHEMISTRY; CHEMISTRY; COMPOSITIONS THEREOF; CULTURE MEDIA; ENZYMOLOGY; HUMAN NECESSITIES; HYGIENE; INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIRCHEMICAL OR PHYSICAL PROPERTIES; MEASURING; MEDICAL OR VETERINARY SCIENCE; METALLURGY; MICROBIOLOGY; MICROORGANISMS OR ENZYMES; MUTATION OR GENETIC ENGINEERING; ORGANIC CHEMISTRY; PEPTIDES; PHYSICS; PREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES; PROPAGATING, PRESERVING OR MAINTAINING MICROORGANISMS; SPIRITS; TESTING; VINEGAR; WINE

Genetically modified microorganism (especially a phage) presenting at least one complex of a peptide and a major histocompatibility complex component (P-MHC complex) on its surface is new. Independent claims are also included for the following: (1) production (M1) of the microorganism by: (a) preparing a nucleotide sequence comprising (i) a sequence coding successively for a signal peptide, an antigenic peptide, a peptide linker and one or more extracellular domains of one or more MHC components and (ii) a functional sequence; (b) cloning the nucleotide sequence in a vector suitable for expression of the sequence in a microorganism, (c) transfecting a microorganism with the vector, and (d) inducing expression of the product of the nucleotide sequence; (2) production (M2) of the microorganism by: (a) preparing a nucleotide a sequence coding successively for a signal peptide, an antigenic peptide and a linking peptide linked directly to a functional sequence; (b) as (b) - (d) above; and (c) forming the P-MHC complex by contacting the expression product with one or more independently expressed MHC components; (3) production (M3) of a soluble P-MHC complex by expressing one or more P-MHC complexes as in (1) or (2), where the functional sequence codes for a tag peptide and a stop codon, and recovering the P-MHC complex; (4) production (M4) of a P-MHC complex presented on the surface of a microorganism by expressing one or more P-MHC complexes as in (1) or (2), where the functional sequence codes for a protein permitting presentation of the P-MHC complex on the surface of the microorganism, and recovering the P-MHC complex. La présente invention concerne un procédé d'expression d'un complexe CMH-P dans un microorganisme, et plus particulièrement dans un phage.En fonction des constructions de séquences nucléotidiques et de vecteurs utilisées dans le procédé de l'invention, on obtient soit des complexes CMH-P solubles, soit des complexes CMH-P présentés à la surface de microorganismes. La présente invention concerne encore l'utilisation des complexes CMH-P solubles ou présentés à la surface de microorganismes dans des procédés de détermination de la spécificité de récepteurs de cellules T ou dans des procédés de criblage de peptides antigéniques. La présente invention concerne enfin l'utilisation des peptides antigéniques sélectionnés dans lesdits procédés de criblage pour la préparation d'une composition pharmaceutique destinée à moduler une réponse immunitaire spécifique dudit peptide.