DNA expression vectors and methods of use

Abstract

The present invention relates to novel plasmid constructs useful for the delivery of DNA vaccines. The present invention provides novel plasmids having a transcription cassette capable of directing the expression of a vaccine nucleic acid insert encoding immunogens derived from any pathogen, including fungi, bacteria and viruses. The present invention, however, is particularly useful for inducing in a patient an immune response against pathogenic viruses such as HIV, measles or influenza. Immunodeficiency virus vaccine inserts of the present invention express non-infectious HIV virus-like particles (VLP) bearing multiple viral epitopes. VLPs allow presentation of the epitopes to multiple histocompatability types, thereby reducing the possibility of the targeted virus escaping the immune response. Also described are methods for immunizing a patient by delivery of a novel plasmid of the present invention to the patient for expression of the vaccine insert therein. Optionally, the immunization protocol may include a booster vaccination that may be a live vector vaccine such as a recombinant pox virus or modified vaccinia Arbora vector. The booster live vaccine vector includes a transcription cassette expressing the same vaccine insert as the primary immunizing vector.

What is claimed is:  
     
         1 . A vector comprising: 
 a termination sequence encoding for the lambda T0 terminator;    a prokaryotic origin of replication;    a selectable marker gene; and    a eukaryotic transcription cassette comprising a vaccine insert encoding one or more immunogens derived from a pathogen.    
     
     
         2 . The vector of  claim 1 , wherein the pathogen is a viral pathogen.  
     
     
         3 . The vector of  claim 1 , wherein the viral pathogen is selected from the group consisting of HIV, measles, influenza, polio and rubella.  
     
     
         4 . The vector of  claim 1 , wherein the one or more immunogens is selected from the group consisting of HIV Gag, HIV gp120 HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza hemagglutinin, mutants thereof, and subsequences thereof.  
     
     
         5 . The vector of  claim 1 , wherein the one or more immunogens are further selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, and HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         6 . The vector according to  claim 1 , wherein the vaccine insert encoding one or more immunogen or immunogens further comprises at least one C3d gene.  
     
     
         7 . A method of immunizing or treating a patient, comprising the step of administering a therapeutically effective amount of a physiologically acceptable composition, comprising the vector of  claim 1 .  
     
     
         8 . The method of  claim 7 , further comprising the step of subsequently administering a therapeutically effective amount of a composition comprising a recombinant pox virus vector expressing one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza hemagglutinin, mutants thereof, and subsequences thereof.  
     
     
         9 . The method of  claim 7 , wherein the one or more immunogens is selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         10 . The method of  claim 7 , further comprising subsequently administering a therapeutically effective amount of a recombinant pox virus vector expressing one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza hemagglutinin, mutants thereof, and subsequences thereof.  
     
     
         11 . The method of  claim 7 , wherein the vector expresses one or more immunogens selected from the group consisting of HIV Gag, HIV gp120. HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         12 . A vector comprising the DNA sequence SEQ ID NO: 1.  
     
     
         13 . The vector of  claim 12 , wherein the vector further comprises a vaccine insert encoding one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza selected hemagglutinin, influenza transmembrane hemagglutinin mutants thereof, and subsequences thereof; and optionally at least one C3d gene.  
     
     
         14 . The vector of  claim 12 , wherein the one or more immunogens is selected from the group consisting of HIV Gag, HIV gp12O, HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         15 . A vector comprising the DNA sequence SEQ ID NO: 2.  
     
     
         16 . The vector of  claim 15 , wherein the vector further comprises a vaccine insert encoding one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza selected hemagglutinin, influenza transmembrane hemagglutinin mutants thereof, and subsequences thereof; and optionally at least one C3d gene.  
     
     
         17 . The vector of  claim 15 , wherein the one or more immunogens is selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         18 . A vector comprising the DNA sequence SEQ ID NO: 3.  
     
     
         19 . The vector of  claim 18 , wherein the vector further comprises a vaccine insert encoding one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza selected hemagglutinin, influenza transmembrane hemagglutinin mutants thereof, and subsequences thereof, and optionally at least one C3d gene.  
     
     
         20 . The vector of  claim 18 , wherein the one or more immunogens is selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         21 . A vector comprising the DNA sequence SEQ ID NO: 4.  
     
     
         22 . The vector of  claim 21 , wherein the vector further comprises a vaccine insert encoding one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza selected hemagglutinin, influenza transmembrane hemagglutinin mutants thereof, and subsequences thereof, and optionally at least one C3d gene.  
     
     
         23 . The vector of  claim 21 , wherein the one or more immunogens is selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         24 . A vector comprising the DNA sequence SEQ ID NO: 5.  
     
     
         25 . The vector of  claim 24 , wherein the vector further comprises a vaccine insert encoding one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza selected hemagglutinin, influenza transmembrane hemagglutinin mutants thereof, and subsequences thereof; and optionally at least one C3d gene.  
     
     
         26 . The vector of  claim 24 , wherein the one or more immunogens is selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         27 . A method of immunizing or treating a patient comprising administering a therapeutically effective amount of a physiologically acceptable composition comprising a vector wherein the vector is a nucleic acid comprising a sequence selected from SEQ ID NOS: 1, 2, 3, 4 and 5, and wherein administration of the therapeutically effective amount of the composition is by an intramuscular or intradermal route.  
     
     
         28 . The method of  claim 27 , wherein the vector expresses one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza hemagglutinin, mutants thereof, and subsequences thereof; and optionally at least one C3d gene.  
     
     
         29 . The method of  claim 27 , wherein the vector expresses one or more immunogens selected from the group consisting of HIV Gag, HIV gp120. HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         30 . The method of  claim 27 , further comprising subsequently administering a therapeutically effective amount of a recombinant pox virus vector expressing one or more immunogens selected from the group consisting of HIV Gag, HIV gp120, HIV Pol, HIV Env, HIV VLP, measles fusion protein, measles hemagglutinin, measles nucleoprotein, influenza hemagglutinin, mutants thereof, and subsequences thereof.  
     
     
         31 . The method of  claim 27 , wherein the vector expresses one or more immunogens selected from the group consisting of HIV Gag, HIV gp120. HIV Pol, HIV Env, HIV VLP, mutants thereof, and subsequences thereof.  
     
     
         32 . A method of immunizing or treating a patient in need thereof comprising administering a therapeutically effective amount of a composition comprising the vector of  claim 29 .