Tetracycline repressor regulated oncolytic viruses

Abstract

The present invention is directed oncolytic Herpes simplex-l viruses whose replication is controlled using a tetracycline operator/repressor system. The invention also includes DNA sequences used in making the viruses and methods in which these viruses are used in the treatment of cancer patients with solid tumors.

1 . A Herpes simplex virus 1 or 2 (HSV) recombinant DNA molecule, comprising: 
 a) a first promoter sequence having a TATA element;    b) a tetracycline operator sequence comprising two op2 repressor binding sites joined by 2-20 linking nucleotides, wherein the first nucleotide in said tet operator is between 6 and 24 nucleotides 3′ to the last nucleotide in said TATA element; and    c) a gene necessary for Herpes simplex 1 or 2 (HSV) virus replication lying 3′ to said operator and operably linked to said first promoter.    
     
     
         2 . The recombinant DNA of  claim 1 , wherein said first promoter is an HSV immediate early promoter, early promoter, or late promoter.  
     
     
         3 . The recombinant DNA of  claim 2 , wherein said first promoter is an HSV immediate early promoter.  
     
     
         4 . The recombinant DNA of  claim 2 , wherein said first promoter is selected from the group consisting of: the HSV ICP4 promoter; ICP27 promoter; ICP8 promoter; UL9 promoter; gD promoter; or VP5 promoter.  
     
     
         5 . The recombinant DNA of  claim 2 , wherein said gene necessary for viral replication is a viral immediate-early, early, or late gene.  
     
     
         6 . The recombinant DNA of  claim 5 , wherein said gene necessary for viral replication is ICP27, ICP4, ICP8, UL9, gD, or VP5 and is under the control of its corresponding promoter.  
     
     
         7 . The recombinant DNA of  claim 1 , wherein said DNA has at least one sequence encoding the tet repressor under the control of a second promoter.  
     
     
         8 . The recombinant DNA of  claim 7 , wherein said second promoter is an HSV immediate early promoter, or the hCMV major immediate-early promoter.  
     
     
         9 . The recombinant DNA of  claim 7 , wherein said second promoter is the HSV ICP0 promoter or ICP4 promoter.  
     
     
         10 . The recombinant DNA of  claim 1 , further comprising a ribozyme sequence or DNA sequence that can reduce translation in the 5′ untranslated region of said gene.  
     
     
         11 . The recombinant DNA of  claim 1 , wherein said recombinant DNA is part of the HSV genome.  
     
     
         12 . The recombinant DNA of  claim 11 , wherein the HSV ICP6 gene is deleted.  
     
     
         13 . The recombinant DNA of  claim 12 , wherein the HSV ICP47 gene is deleted.  
     
     
         14 . The recombinant DNA of  claim 12 , further comprising at least one therapeutic gene that can augment tumor killing activity or anti-tumor specific immunity.  
     
     
         15 . An oncolytic HSV virus comprising as its genome, the recombinant DNA of  claim 11 .  
     
     
         16 . The oncolytic virus of  claim 15 , wherein said recombinant DNA does not include a functional HSV ICP6 gene.  
     
     
         17 . The oncolytic virus of  claim 16 , wherein said recombinant DNA does not include a functional HSV ICP47 gene.  
     
     
         18 . The oncolytic virus of  claim 15 , wherein said recombinant DNA comprises at least one therapeutic gene that can augment tumor killing activity or anti-tumor specific immunity.  
     
     
         19 . A method of treating a cancer patient having a solid tumor, comprising: 
 a) locally administering to said solid tumor the oncolytic virus of  claim 15  for a period sufficient to allow infection of tumor cells    b) administering tetracycline either systemically to said patient or locally to said tumor.    
     
     
         20 . The method of  claim 19 , wherein said oncolytic virus does not include a functional HSV ICP6 gene within its genome.  
     
     
         21 . The method of  claim 20 , wherein said oncolytic virus does not include a functional HSV ICP47 gene within its genome.  
     
     
         22 . The method of  claim 19 , wherein said oncolytic virus comprises at least one therapeutic gene that can augment tumor killing activity within its genome or anti-tumor specific immunity.  
     
     
         23 . The method of  claim 19 , wherein said tumor is a melanoma or a tumor of the lung, colon, brain, breast, prostate, pancreas, kidney, esophagus, liver, ovary, testis or stomach.