Title:  Methods for producing an immune response against HIV-1

United States Patent:  6,511,845

Issued:  January 28, 2003

Inventors:  Davis; Alan R. (4411 Oak Forest Dr., Missouri City, TX 77459); Hung; Paul P. (506 Ramblewood Dr., Bryn Mawr, PA 19010); Lubeck; Michael D. (2265 Spangler Rd., York, PA 17402); Natuk; Robert J. (63 Vones La., Raritan, NJ 08869); Chanda; Pranab K. (30 Zaitz Farm Rd., West Windsor, NJ 08550); Murthy; Shridhara C. S. (4677 Erin Ct., Ann Arbor, MI 48105); Lee; Shaw-Guang L. (155 S. Spring Mill Rd., Villanova, PA 19085)

Appl. No.:  618360

Filed:  July 18, 2000

This invention provides a method of protecting a primate from an infectious organism by stimulating the production of antibodies or cell mediated immunity to the infectious organism which comprises administering to said primate intranasally, intramuscularly, or subcutaneously, live recombinant adenoviruses in which the virion structural protein is unchanged from that in the native adenovirus from which the recombinant adenovirus is produced, and which contain the gene coding for the antigen corresponding to said antibodies or inducing said cell mediated immunity. Preferably, the infectious organism is HIV and the primate is a human.

SUMMARY OF THE INVENTION

This invention provides a method of producing antibodies or cell mediated immunity to an infectious organism in a warm blooded mammal which comprises administering to said warm blooded mammal intranasally, intramuscularly, or subcutaneously, live recombinant adenoviruses in which the virion structural protein is unchanged from that in the native adenovirus from which the recombinant adenovirus is produced, and which contain the gene coding for the antigen corresponding to said antibodies or inducing said cell mediated immunity. The warm blooded mammal is preferably a primate, most preferably a human.

In its preferred embodiments, this invention provides a method of producing antibodies to human immunodeficiency virus (HIV-1), hepatitis B, hepatitis C, human papilloma virus, respiratory syncytial virus, rotavirus, or parainfluenza virus in a warm blooded mammal which comprises administering to said warm blooded mammal intranasally, intramuscularly, or subcutaneously, live recombinant adenoviruses in which the virion structural protein is unchanged from that in the native adenovirus from which the recombinant adenovirus is produced and which contain the gene coding for, respectively, human immunodeficiency virus, hepatitis B, hepatitis C, human papilloma virus, respiratory syncytial virus, rotavirus, or parainfluenza virus.

This invention also provides composition for producing antibodies or cell mediated immunity to an infectious organism in a warm blooded mammal, comprising live recombinant adenoviruses in which the virion structural protein is unchanged from that in the native adenovirus from which the recombinant adenovirus is produced, and which contain the gene coding for the antigen corresponding to said antibodies or inducing said cell mediated immunity, said composition being formulated in an intranasal, intramuscular, or subcutaneous dosage form.

Although this specification specifically refers to adenovirus of types 4, 5, or 7, live, infectious adenovirus of any type may be employed in this invention. Additionally, while the specification specifically refers to adenoviruses having an early region 3 (E3) deletion, adenoviruses which are attenuated, contain a temperature sensitive lesion, or a E1 deletion may also be used as a vector. Similarly, although specific reference has been made to vaccines producing antibodies to HIV, hepatitis B, hepatitis C, human papilloma virus, respiratory syncytial virus, rotavirus, or parainfluenza virus, our invention provides vaccines against any infectious agent containing an antigen to which a warm-blooded animal will produce antibodies or cell mediated immunity, and which antigen is coded for by a gene composed of up to about 3000 base pairs. Thus, for example, included within the scope of the invention are immunization against such diseases as influenza, hepatitis A, cholera, E. coli, pertussis, diphtheria, tetanus, shigellosis, gonorrhea, mycoplasma pneumonia, and the like.

In one embodiment, the method of treatment includes administering the recombinant adenovirus both prophylactically to an HIV-1 susceptible mammal and as immunotherapy following detection of HIV in said mammal. Regimens containing the following recombinant adenoviruses were used to produce the anti-HIV responses.

In a preferred embodiment, the method is a method of protecting a primate against HIV-1 infection comprising intranasal or intramuscular administration to said primate of an intranasal or intramuscular dosage of a recombinant adenovirus having a deletion in the E3 gene and an expression cassette containing a major late promoter, a tripartite leader sequence, part or all of the HIV-1 gp160 sequence and a polyadenylation signal sequence. Preferably the primate is a human. The expression cassette is inserted into the recombinant adenovirus between the E4 promoter and the inverted terminal repeat. Optionally the intranasal or intramuscular administration of recombinant adenovirus is followed by one or more intranasal or intramuscluar booster administrations of the recombinant adenovirus. The recombinant adenovirus is a serotype 4, 5 or 7 serotype adenovirus and optionally the expression cassette additionally comprises part of all of the coding sequence for the HIV-1 rev gene inserted in frame after the HIV-1 gp160 sequence and before the polyadenylation signal sequence. The HIV-1 gp160 sequence can be from the MN strain gp160 sequence or the LAV strain gp160 sequence. In an alternative embodiment, the HIV-1 gp160 sequence is replaced by a sequence encoding the gag-pro region of HV-1. In either embodiment, when the initial administration is followed by one or more intranasal or intramuscular booster administrations of the recombinant adenovirus, the last booster administration may be followed by an intramuscular injection of at least one booster immunization with an HIV-1 subunit antigen preparation, preferably containing an HIV-1 gag and/or env polypeptide sequence. For intranasal administration, the intranasal dosage administered is in the range of 1.times.10⁷ pfu of virus and for intramuscular administration, the intramuscular dosage administered is in the range of 1.times.10⁷ to 2.times.10⁹ pfu of virus. The intranasal booster is administered in a dosage in the range of 1.times.10⁷ to 1.times.10⁸ pfu of virus and the intramuscular booster is administered in a dosage in the range of 1.times.10¹⁰ to 8.times.10¹⁰ pfu of virus. When a subunit antigen booster is employed, the subunit antigen preparation contains between 200 .mu.g and 0.5 mg of HIV-1 polypeptide.

Referring to the above table Ad4, Ad5, and Ad7 refer to human adenoviruses types 4, 5, and 7 respectively in which the E3 region has been deleted. Env refers to the HIV envelope glycoprotein (gp 160) gene. Gag refers to the HIV gag/pro gene. Rev refers to the HIV regulatory gene. Hrev refers to an altered version of the rev gene where the nucleotide sequences were changed without changing the amino acid sequence employing codons that were frequently used in human genes.  Tpl refers to the upstream adenovirus tripartite leader sequence with an intervening sequence between the first and second leaders that are positioned in front of the recombinant genes. The constructs designated Ad7-env, Ad7-gag, Ad7-gag-1, Ad4-env, Ad4-gag, Ad4-gag-1, Ad5-env, and Ad5-gag contain either the gag or the env gene from the LAV strain of HIV and the constructs Ad7-env_MN, Ad4-env_MN, and Ad5-env_MN contain the env gene from the MN strain of HIV. The recombinant adenoviruses made from the LAV and MN strains of HIV-1 are illustrative of recombinant adenoviruses covered by this invention. This invention also covers recombinant adenoviruses which include the env and/or gag genes from other strains of HIV-1.

Both the Ad-env and Ad-env_MN adenoviruses were shown to replicate in human A549 cells and expressed recombinant env antigen in vitro demonstrating their capability of generating cell mediated, humoral, and secretory immunity in a mammal.

As described in detail below, intranasal administration of Ad-HIV recombinant viruses to naive chimpanzees resulted in both priming and boosting of both humoral and cell-mediated immune responses directed at HIV recombinant antigens. The recombinant adenoviruses administered to chimpanzees were shown to produce antibodies to the env and gag proteins of HIV. IgG antibodies specific for HIV were observed in nasal, saliva, and vaginal secretions following administration of the recombinant adenoviruses and IgA antibodies specific for HIV were observed in nasal and saliva secretions. The first set of recombinant viruses (Ad7) appeared to be shed the longest period of time and induce the best anti-Ad antibody response. The results also showed that administration of Ad-HIV vaccines by the intranasal route was superior to administration of enteric-coated recombinant viruses by the oral route.

Optimum immune responses directed at HIV antigens required primary infection one booster immunization with a heterotypic recombinant Ad-HIV to elicit strong anti-HIV binding antibodies. Intranasal administration of the Ad-HIV viruses effectively primed chimpanzees to respond with high titered neutralizing antibodies to HIV-1 following subsequent HIV-1 subunit protein booster immunization.

Claim 1 of 24 Claims

What is claimed is:

1. A method of producing an immune response against HIV-1 infection in a human comprising intranasal administration to said human an intranasal dosage of a recombinant adenovirus having a deletion in the E3 gene and an expression cassette containing a major late promoter, a tripartite leader sequence, part or all of the HIV-1 gp160 sequence and a polyadenylation signal sequence, said cassette being inserted into said recombinant adenovirus between the E4 promoter and the inverted terminal repeat of said recombinant adenovirus, wherein said intranasal administration of the recombinant adenovirus is followed by one or more intranasal or intramuscular booster administrations of said recombinant adenovirus.
 

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