Plasmodium falciparum antigens and their vaccine and diagnostic applications

Abstract

The invention concerns novel Plasmodium falciparum antigens and their vaccine and diagnostic applications. More particularly, the invention concerns immunogenic polynucleotide and polypeptide molecules, compositions comprising them, and methods for diagnosis and vaccination of malaria.

1 . An isolated or purified polynucleotide comprising a nucleotide sequence with at least 60%, preferably at least 80% and more preferably at least 95% identity with SEQ ID NO: 1 (DG747) or SEQ ID NO: 2 (DG772). 
     
     
         2 . An isolated or purified polynucleotide comprising at least 10 consecutive nucleotides identical to SEQ ID NO: 1 or SEQ ID NO: 2. 
     
     
         3 . An isolated or purified polynucleotide, which hybridizes under highly stringent conditions with a polynucleotide according to  claim 1  or  claim 2 . 
     
     
         4 . An isolated or purified polypeptide, encoded by a polynucleotide according to  claim 1 . 
     
     
         5 . An isolated or purified polypeptide comprising at least 60%, preferably at least 80% and more preferably at least 95% homology with SEQ ID NO: 3 (DG747) or SEQ ID NO: 4 (DG772). 
     
     
         6 . An isolated or purified polypeptide comprising at least 5 consecutive amino acids identical to one of the sequences selected from SEQ ID NOs: 3 to 7, and SEQ ID NO: 8. 
     
     
         7 . An isolated or purified polypeptide comprising at least 40%, preferably at least 60% more preferably at least 80% and still more preferably at least 95% identity with one of the sequences selected from SEQ ID NOs: 3 to 8, 10 and SEQ ID NO: 12. 
     
     
         8 . A recombinant or chimeric recombinant polypeptide consisting of at least one polypeptide according to  claim 4 . 
     
     
         9 . An isolated or purified antigen consisting of an antigenic fragment encoded by a polynucleotide according to  claim 1 . 
     
     
         10 . An antigenic conjugate comprising a polynucleotide according to  claim 1 , and a support on which said polynucleotide is bound. 
     
     
         11 . A conjugate according to  claim 10 , in which said support is constituted by microspheres, microparticles of latex beads, polyphosphoglycan microparticles (PGLA) or polystyrene microparticles. 
     
     
         12 . (canceled) 
     
     
         13 . Monoclonal or polyclonal antibodies specifically recognizing a polypeptide according to  claim 4 . 
     
     
         14 . Antibodies according to  claim 13 , which are humanized. 
     
     
         15 . A cloning or expression vector comprising a polynucleotide according to  claim 1 . 
     
     
         16 . A vector according to  claim 15 , in which said polynucleotide is incorporated into a site that is not essential to replication of said vector. 
     
     
         17 . A vector according to  claim 15  or  16 , which is selected from plasmids, cosmids and phages. 
     
     
         18 . A host cell comprising a vector according to  claim 15  or  16 . 
     
     
         19 . A recombinant  E. coli  cell selected from cells deposited at the CNCM on 23 May 2001 with accession numbers 1-2671 and 1-2672. 
     
     
         20 . An immunogenic composition comprising:
 any one of the following elements: a polynucleotide according to  claim 1 , a polypeptide according to  claim 4 , a conjugate according to  claim 10  or  43 ; and   a pharmaceutically acceptable vehicle.   
     
     
         21 . An immunogenic composition according to  claim 20 , consisting of at least one compound selected from alum, QS21, montanide, SBAS 2  adjuvant and incomplete Freund's adjuvant. 
     
     
         22 . An immunogenic composition according to  claim 20 , in which said polypeptide is adsorbed onto microparticles. 
     
     
         23 . An immunogenic composition according to  claim 20 , in which said polynucleotide is in the form of DNA. 
     
     
         24 . An immunogenic composition according to  claim 20 , comprising at least one epitope selected from the proteins CS, MSP-I, MSP-3, LSA-1, TRAP, STARP, SALSA, SALSA 1, SALSA II and LSA-3. 
     
     
         25 . An immunogenic composition according to  claim 20 , which can produce a cell response and/or humoral response in vivo and/or in vitro. 
     
     
         26 . An immunogenic composition according to  claim 20 , which can allow the production of γ-interferon by leukocytes from subjects immunized with irradiated sporozoites. 
     
     
         27 . An immunogenic composition according to  claim 20 , which can produce a humoral IgG response. 
     
     
         28 . An immunogenic composition according to  claim 27 , which can produce a humoral type IgG1, IgG2, IgG3 and/or IgG4 response. 
     
     
         29 . An immunogenic composition according to  claim 20 , which is capable of inducing, in vivo and in vitro, protection by a challenge infection with  Plasmodium falciparum.    
     
     
         30 . An anti-malaria vaccine comprising:
 any one of the following elements: a polynucleotide according to  claim 1 , a polypeptide according to  claim 4 , a conjugate according to  claim 10  or  43 ; and   a pharmaceutically acceptable vehicle.   
     
     
         31 . A vaccine according to  claim 30  comprising at least one epitope selected from the proteins CS, MSP-1, MSP-3, LSA-1, TRAP, STARP, SALSA, SALSA I, SALSA II and LSA-3. 
     
     
         32 . A pharmaceutical composition comprising, as the active substance, one or more polyclonal or monoclonal antibodies according to  claim 13 , in association with a pharmaceutically acceptable vehicle. 
     
     
         33 . A pharmaceutical composition according to  claim 32 , comprising at least one compound selected from alum, QS21, montanide, SBAS 2  adjuvant and incomplete Freund's adjuvant. 
     
     
         34 . (canceled) 
     
     
         35 . An in vitro method for diagnosing malaria infection, wherein the method comprises the following steps:
 a) contacting a biological tissue and/or fluid removed from an individual who is susceptible of being infected with  Plasmodium falciparum  with an antibody according to  claim 13  to allow the formation of immune complexes; and   b) detecting in vitro the presence or absence of any immune complexes formed.   
     
     
         36 . An in vitro method for diagnosing malaria infection, wherein the method comprises the following steps:
 a) contacting a biological tissue and/or fluid removed from an individual susceptible of being infected with  Plasmodium falciparum  with any one of the following elements: a polynucleotide according to  claim 1 , a polypeptide according to  claim 4 , a conjugate according to  claim 10  or  43 ; to allow the formation of immune complexes involving at least one of said elements and antibodies that may be present in said biological tissue or fluid; and   b) detecting in vitro the presence or absence of any immune complexes formed.   
     
     
         37 . A method according to  claim 35 , in which step a), the biological tissue and/or fluid is brought into contact with at least one epitope selected from CS, MSP-1, MSP-3, LSA-1, TRAP, STARP, SALSA, SALSA I, SALSA II or LSA-3. 
     
     
         38 . An in vitro malaria diagnostic kit, comprising the following elements:
 a) any one element selected from a polynucleotide according to  claim 1 , a polypeptide according to  claim 4 , a conjugate according to  claim 10  or  43 ;   b) reagents for constituting a medium suitable for a binding reaction between a test sample and at least one of the elements defined in a); and   c) reagents allowing the detection of antigen-antibody complexes produced by said binding reaction, said reagents also possibly carrying a label susceptible of being recognized by a second labelled reagent.   
     
     
         39 . An in vitro malaria diagnostic kit, comprising the following elements:
 antibodies as defined in  claim 13 ;   reagents for constituting a medium suitable for allowing a binding reaction between a test sample and at least one said antibody; and   reagents allowing the detection of antigen-antibody complexes produced by said binding reaction, said reagents also possibly carrying a label susceptible of being recognized by a second labelled reagent.   
     
     
         40 . An in vitro malaria diagnostic kit according to  claim 38 , comprising at least one peptide selected from CS, MSP-1, MSP-3, LSA-1, TRAP, STARP, SALSA, SALSA I, SALSA II and LSA-3. 
     
     
         41 . An in vitro malaria diagnostic kit according to  claim 38 , comprising a SBAS 2  adjuvant. 
     
     
         42 . An isolated or purified antigen, consisting of an antigenic fragment of a polypeptide according to  claim 4 . 
     
     
         43 . An antigenic conjugate comprising a polypeptide according to  claim 4 ; and a support on which said polypeptide is bound. 
     
     
         44 . A method for immunizing an individual, comprising administering to an individual, infected with or susceptible of being infected with malaria, a conjugate according to  claim 10  or  43 . 
     
     
         45 . A method according to  claim 36 , in which step a), the biological tissue and/or fluid is brought into contact with at least one epitope selected from CS, MSP-1, MSP-3, LSA-1, TRAP, STARP, SALSA, SALSA I, SALSA II or LSA-3. 
     
     
         46 . An in vitro malaria diagnostic kit according to  claim 39 , comprising at least one peptide molecule selected from CS, MSP-1, MSP-3, LSA-1, TRAP, STARP, SALSA, SALSA I, SALSA II and LSA-3. 
     
     
         47 . An in vitro malaria diagnostic kit according to  claim 39 , comprising a SBAS 2  adjuvant.