Human cytomegalovirus DNA constructs and uses therefor

Abstract

Novel DNA molecules for in vitro and in vivo expression of HCMV gB, gB transmembrane-deleted derivatives, pp65, pp150, and IE-exon-4 proteins are described. Preferably, the molecules are plasmids. Also described are methods of using these DNA molecules to induce immune responses to HCMV, and the use of a plasmid of the invention to prime immune responses to HCMV vaccines.

What is claimed is:  
     
         1 . A DNA molecule which is non-replicating in mammals and comprises a sequence encoding a human cytomegalovirus antigen, 
 wherein the sequence is operably linked to regulatory sequences for expressing the antigen in mammals and wherein the antigen elicits an immune response in the mammal.    
     
     
         2 . The DNA molecule according to  claim 1 , which is a plasmid.  
     
     
         3 . The DNA molecule according to  claim 1 , wherein said antigen is selected from the group consisting of: 
 (a) gB;    (b) a gB derivative lacking at least the transmembrane domain;    (c) pp65;    (d) pp150;    (e) immediate-early exon-4; and    (f) combinations of (a)-(e).    
     
     
         4 . The DNA molecule according to  claim 3 , which comprises a sequence encoding the gB and the pp65 antigens.  
     
     
         5 . The DNA molecule according to  claim 3 , which comprises a sequence encoding the gB derivative and a sequence encoding the pp65 antigen.  
     
     
         6 . A pTet-gB DNA plasmid, said plasmid comprising the human cytomegalovirus (HCMV) gB gene and a tetracycline regulatable HCMV-immediate early promoter, said promoter controlling the expression of gB.  
     
     
         7 . A pΔRC/CMV DNA plasmid, said plasmid comprising the human cytomegalovirus (HCMV) gB gene and capable of expressing gB.  
     
     
         8 . A pΔRC-gB 680  plasmid, said plasmid comprising the portion of the human cytomegalovirus (HCMV) gene encoding the N-terminal 680 amino acids of the gB protein (gB 1-680 ) and capable of expressing gB 1-680 .  
     
     
         9 . A pΔRC-pp150 plasmid, said plasmid comprising the human cytomegalovirus (HCMV) gene encoding the HCMV pp150 tegument protein and capable of expressing pp150.  
     
     
         10 . A pΔRC-exon-4 plasmid, said plasmid comprising the portion of the human cytomegalovirus (HCMV) gene encoding HCMV immediate-early (IE)-exon-4 and capable of expressing IE-exon-4.  
     
     
         11 . An immunogenic composition comprising a carrier and a DNA molecule according to  claim 1 .  
     
     
         12 . A pCBgBΔtm plasmid, said plasmid comprising the human cytomegalovirus (HCMV) gB gene containing a deletion in the transmembrane domain.  
     
     
         13 . An immunogenic composition comprising a carrier and a DNA molecule according to  claim 1 .  
     
     
         14 . The immunogenic composition according to  claim 13 , wherein the DNA molecule is selected from the group consisting of: 
 (a) pΔRC-gB;    (b) pTet-gB;    (c) pΔRC-pp65;    (d) pΔRC-gB 680 ;    (e) pΔRC-pp150;    (f) pCBgB;    (g) pCBgBΔtm; and    (h) pΔRC-exon-4.    
     
     
         15 . The immunogenic composition according to  claim 13 , comprising two or more DNA molecules.  
     
     
         16 . The immunogenic composition according to  claim 14 , comprising a first DNA molecule which comprises a sequence encoding the gB antigen or a gB derivative, and a second DNA molecule which comprises a sequence encoding the pp65 antigen.  
     
     
         17 . The immunogenic composition according to  claim 13 , wherein the carrier is selected from the group consisting of saline and isotonic water.  
     
     
         18 . A method of inducing human cytomegalovirus-specific (HCMV) immune responses in an animal, comprising the step of administering to said animal an effective amount of a first immunogenic composition according to  claim 13 .  
     
     
         19 . The method according to  claim 18 , wherein the composition comprises pTet-gB and pΔRC-pp65.  
     
     
         20 . The method according to  claim 18 , further comprising the step of administering a second immunogenic composition to said animal, said second immunogenic composition comprising a plasmid selected from the group consisting of: 
 (a) pΔRC-gB;    (b) pTet-gB;    (c) pΔRC-pp65;    (d) pΔRC-gB 680 ;    (e) pΔRC-pp150;    (f) pCBgB;    (g) pCBgBΔtm; and    (h) pΔRC-IE-Exon-4.    
     
     
         21 . The method according to  claim 18 , wherein said second immunogenic composition is administered between about 2 to about 15 weeks following administration of said first immunogenic composition.  
     
     
         22 . A method of priming immune responses to a selected human cytomegalovirus immunogenic composition, comprising the steps of: 
 administering a first immunogenic composition according to  claim 13  and administering the selected human cytomegalovirus immunogenic composition.    
     
     
         23 . The method according to  claim 22 , wherein the first immunogenic composition is administered between about 4 and 15 weeks prior to administration of the selected immunogenic composition.  
     
     
         24 . The method according to  claim 22 , wherein the first immunogenic composition comprises pTet-gB.  
     
     
         25 . The method according to  claim 24 , wherein pTet-gB is administered in an amount between about 50 μg to about 160 μg.  
     
     
         26 . The method according to  claim 22 , wherein the selected immunogenic composition comprises an immunogen selected from the group consisting of a recombinant virus comprising an HCMV immunogen, an HCMV protein, and HCMV virions.  
     
     
         27 . The method according to  claim 26 , wherein the HCMV protein is gB.  
     
     
         28 . The method according to  claim 26 , wherein the recombinant virus is selected from the group consisting of Ad5.gb and Ad5-IE-exon-4.  
     
     
         29 . A DNA molecule which is non-replicating in mammals and comprises a sequence encoding a human cytomegalovirus antigen selected from the group consisting of: 
 (a) pp65;    (b) pp150;    (c) immediate-early exon-4;    (d) gB and an antigen of (a) to (c);    (e) a gB derivative lacking at least the transmembrane domain and an antigen of (a) to (c); and    (f) a combination of antigens (a) to (c).    
     
     
         30 . An immunogenic composition comprising a carrier and at least two DNA molecules, wherein said DNA molecules are selected from the group consisting of: 
 (a) gB;    (b) a gB derivative lacking at least the transmembrane domain;    (c) pp65;    (d) pp150; and    (e) immediate-early exon-4.