FUSIONS OF AN HAEMOPHILUS PAL AS VACCINE CANDIDATES.

HAEMOPHILUS INFLUENZAE TYPE B (HIB) POLYSACCHARIDE AND POLYSACCHARIDE-PROTEIN CONJUGATE VACCINES HAVE BEEN PROVEN EFFECTIVE AGAINST HIB DISEASE. HOWEVER, CAPSULAR ANTIGEN ISOF NO VALUE AGAINST NON-TYPABLE H. INFLUENZAE (HI) DISEASE. A 16000 DALTON OUTER MEMBRANE PEPTIDOGLYCAN ASSOCIATED LIPOPROTEIN (PAL) OF HI HAS BEEN IDENTIFIED AS A POTENTIAL VACINE CANDIDATE. THIS PROTEIN HAS BEEN SHOWN TO RAISE BACTERICIDAL AND PROTECTIVE ANTIBODIES AGAINST A WIDE RANGE OF HI AND HIB STRAINS. THE GENE ENCODING THIS PROTEINHAS BEEN CLONED IN E. COLI. HIGH LEVEL PRODUCTION OF PAL INE. COLI IS TOXIC TO THE HOST CELL. THESE STUDIES ARE DESIGNED TO CREATE A MODIFIED PAL PROTEIN LACKING THE MEMBRANE SIGNAL PEPTIDE AND TO SUBCLONE 5' AND 3' ENDS, AND INTERNAL FRAGMENTS OF THE GENE LINKED IN FRAME TO THE GENE FOR THE B SUBUNIT OF E. COLI HEAT LABILE ENTEROTOXIN (LT-B). THESE CONSTRUCTS WILL BE USED TO DETERMINE IF MEMBRANE PROCESSING AND/OR FATTY ACYLATION OF PAL IS REQUIRED FOR THE GENERATION OF HIB PROTECTIVE EPITOPES; AND IF NOT, WILL ALLOW US TO IDENTIFY WHAT REGION(S) OF THE PROTEIN EXPRESS PROTECTIVE EPITOPES. PHASE II STUDIES WILL INVOLVE LARGE SCALE PURIFICATION AND TESTING OF THE MOST PROMISING CONSTRUCT AS A POTENTIAL VACCINE CANDIDATE.