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WO1992000098A1 - Methods of inducing immune response to aids virus - Google Patents

Methods of inducing immune response to aids virus Download PDF

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Publication number
WO1992000098A1
WO1992000098A1 PCT/EP1991/001225 EP9101225W WO9200098A1 WO 1992000098 A1 WO1992000098 A1 WO 1992000098A1 EP 9101225 W EP9101225 W EP 9101225W WO 9200098 A1 WO9200098 A1 WO 9200098A1
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Prior art keywords
composition
hiv
epitopes
peptides
recombinant
Prior art date
Application number
PCT/EP1991/001225
Other languages
French (fr)
Inventor
Daniel Zagury
Jean-Claude Imbert
Jean-Jacques Salaun
Lurhuma Zirimwamba
Original Assignee
Daniel Zagury
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Publication of WO1992000098A1 publication Critical patent/WO1992000098A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A61K39/21—Retroviridae, e.g. equine infectious anemia virus
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K40/00—Cellular immunotherapy
    • A61K40/10—Cellular immunotherapy characterised by the cell type used
    • A61K40/13—B-cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K40/00—Cellular immunotherapy
    • A61K40/20—Cellular immunotherapy characterised by the effect or the function of the cells
    • A61K40/24—Antigen-presenting cells [APC]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K40/00—Cellular immunotherapy
    • A61K40/40—Cellular immunotherapy characterised by antigens that are targeted or presented by cells of the immune system
    • A61K40/46—Viral antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55511—Organic adjuvants
    • A61K2039/55566—Emulsions, e.g. Freund's adjuvant, MF59
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K40/00
    • A61K2239/31—Indexing codes associated with cellular immunotherapy of group A61K40/00 characterized by the route of administration
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K40/00
    • A61K2239/38—Indexing codes associated with cellular immunotherapy of group A61K40/00 characterised by the dose, timing or administration schedule
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
    • C12N2710/00011—Details
    • C12N2710/24011—Poxviridae
    • C12N2710/24111—Orthopoxvirus, e.g. vaccinia virus, variola
    • C12N2710/24141—Use of virus, viral particle or viral elements as a vector
    • C12N2710/24143—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16034—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16071—Demonstrated in vivo effect
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16111—Human Immunodeficiency Virus, HIV concerning HIV env
    • C12N2740/16122—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16211—Human Immunodeficiency Virus, HIV concerning HIV gagpol
    • C12N2740/16222—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16311—Human Immunodeficiency Virus, HIV concerning HIV regulatory proteins
    • C12N2740/16322—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes

Definitions

  • This invention provides means of inducing an immune response to viral antigens as a means to protect against infection.
  • the methods of the invention also induce immune response against viral antigens in infected individuals, thereby slowing the progress of the disease.
  • HIV human immuno-deficiency virus
  • AIDS acquired immune deficiency syndrome
  • immunization against the HIV retrovirus is exemplified, such exemplification should not be considered as a limitation on the invention.
  • T 4 lymphocytes is required for viral release. These cells act as the source for expansion, and dissemination of virions in the organisms leading to AIDS.
  • the viral release which leads to death of infected cells is preceded by a stage where HIV signals are present in the cell membrane. This immunogenic stage is important as a trigger for the cellular reaction against HIV. The reaction leads to the destruction of the infected cells.
  • HIV HIV
  • rapid progress in the isolation, cloning, and sequencing of the entire genome has shown the remarkable propensity of the HIV strains to mutate, particularly within the viral envelope gene.
  • viral envelope proteins are often the target for neutralizing antibodies, this extensive variation may play an important role in the interaction between the virus and the host's immune system.
  • rapid mutation is an important means of escape from neutralizing antibodies.
  • Such mutation results in successive waves of influenza epidemics among previously infected populations.
  • For visna virus, a sheep retrovirus, the mutation rate is believed to be so rapid as to allow antibody escape during the course of a single chronic infection.
  • the CD4 binding site of HIV has been mapped to three relatively conserved regions of gpl20. Divergent isolates bind soluble CD4 and are inactivated by it, suggesting conservation of the CD4 binding site. Presumably, if neutralizing anti.bodies were directed against this site or another site responsible for a critical viral function, such antibodies would be active against numerous clinical isolates of the virus. A vaccine capable of eliciting antibodies against a broad spectrum of HIV strains is needed.
  • recombinant virus carrying HIV segments (envelope, gag, or pol).
  • HIV recombinant virus
  • An example of such a recombinant virus is the virus of Moss. Mackett, et al., J. Virol. 49, 857-864.
  • Another method of developing immune response to HIV may be accomplished by incubating autologous cells with synthetic peptides to allow peptides to form complexes with HLA antigens on the cell surface, then fixing the cells by usual methods known in the art, e.g., fixation with paraformaldehyde or glutaraldehyde (Zagury, et al, Nature. 332, pp. 728-731 (1988)).
  • compositions containing only the cell-free membranes from autologous cells that have been incubated with the appropriate peptides or have been infected with virus containing recombinant HIV nucleic acid sequences.
  • sequences to which the autologous cells are exposed are usually from the envelope, gag, or pol proteins.
  • compositions containing "cocktails" of several free peptides representing sequences found in HIV proteins can be added to oils to form an emulsion.
  • These compositions have immunogenic properties and can be administered to individuals either in conjunction with administration of preparations containing treated autologous cells or cell free membranes.
  • the peptidecontaining emulsions can also be administered alone as a means of raising an immune response in the individual
  • the emulsions are administered parenterally, intramuscularly or subcutaneously.
  • compositions of the present invention either alone or in c ⁇ njunction with a protocol that includes use of autologous cells that have been exposed in vitro to HIV antigens, or cell membranes obtained therefrom.
  • the "peptide cocktail" approach allows the use of selected amino acid sequences. These sequences may be chosen from regions of the protein that are conserved across the various strains of the organism. Additionally, peptide sequences from several stra.ins may be chosen so that a broad range of viral strains may be used to give broad group protection against HIV Vairus.
  • the peptides may be prepared by any means known in the art, such as by recombinant means or by the Merrifield process.
  • the process of preparing the peptides by synthetic means provides not only reliability, but also provides certain economic advantages.
  • Peptides should be of about 8 to 40 amino acids, with peptides of 12 to 30 amino acids preferred.
  • protein fragments having molecular weights of over 10,000 can be added to the emulsion or can be given separately from the peptides to stimulate increased antibody response.
  • adjuvants and other additives known in the art may also be added to the peptide-containing emulsions.
  • Some of the larger segments that can be used include gp160, gp41, gp 566 , gp24, reverse trans ⁇ riptase (RT), Tat protein, and protease.
  • peptides from several strains and differing locations in the HIV proteins can be used, particularly preferred sites are listed below:
  • AIDS or ARC aids-related complex patients with 150 - 400 T4 cells per mm 3 were immunized with fixed autologous B cells transformed with EBV (Epstein-Barr Virus) and infected with recombinant vaccinia expressing
  • HIV proteins env, gag, and pol on the surface of infected cells.
  • the fixed cells were given by intravenous slow drip infusion (5-7 ⁇ 10] cells), intramuscularly (10
  • the vaccinia vector used is the non-neurotropio strain Lister and the recombinant is produced by the method of Moss (sea Mackett, supra.)
  • the patients underwent biweekly physical examination. Blood samples were drawn for preparation of serum and cells for biological investigations including virus neutralizing antibody, T4 cell count, cell mediated immunity, and cell mediated cytotoxicity.
  • AIDS or ARC patients with 150 - 400 T4 cells per mm 3 received the preparation described in example 1 in conjunction with AZT (600 mg per day Other conditions were similar to those described in example 1.
  • AIDS or ARC patients with 150 - 400 T4 cells per mm 3 received treatment as described in example 1 along with discontinuous AZT at low doses (600 mg per day for 30 days every 90 days). Other conditions were similar to those described in example 1.
  • AIDS or ARC patients with 150 - 400 T4 cells per mm 3 received a mixture of synthetic HIV peptides comprising peptides which constituted immunodominant sites of env, gag, pol (peptide 342-350 according to Ratner). Immunisation protocols and patient follow up were as described in example 1.
  • Seronegativepatients were treated as described in example 5 except that the synthetic peptides administered ware protectively encapsulated as a water-in-oil emulsion. All other conditions were as described in example 5.
  • Seronegativepatients wore given synthetic peptides as In example 5 except that the peptides were administered as free peptides.
  • Seronegativepatients were given synthetic peptides representing HIV epitopes wherein the peptides were covalently linked to an immuno-enhancing moiety. All other conditions were as described in example 5.
  • Peptide derivatives having hydrophobic groups consisting of tripalmitoyl cysteine, dipalmitoyl lysine, or a non-viral peptide of alpha helix configuration were administered in accord with the protocol used to administer the peptides in example 5.
  • Peptides were administered in accord with example 5. However, the patients were also given recombinant vectors containing HIV nucleic acid sequences at separate sites in conjunction with the peptides.
  • Peptides were given in a composition containing recombinant live vectors containing HIV nucleic acid sequences mixed with the synthetic peptides which represented HIV immunodominant epitopes protectively encapsulated as a water in oil emulsion.
  • the composition was given in the manner described in example 5.
  • Montanide is a product of SEPPIC, a division of Cosmetique-Pharmacie, 70, Champs-Elysees, 75008 Paris, France, and is obtainable therefrom.

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Abstract

A safe, effective vaccine will raise antibodies against several parts of the genome of the HIV virus. A protocol for the administration of the vaccine will cause the production of an immune response to protect an individual against several strains of HIV virus. The immunotherapy protocol will cause the individual to develop protective immunity against HIV blocking viral expansion and dissemination in infected individuals. New improved immunogenic compositions are also provided.

Description

METHODS OF INDUCING IMMUNE RESPONSE TO AIDS VIRUS
SUMMARY OF THE INVENTION
This invention provides means of inducing an immune response to viral antigens as a means to protect against infection. The methods of the invention also induce immune response against viral antigens in infected individuals, thereby slowing the progress of the disease. Of particular importance is protection against human immuno-deficiency virus (HIV), the causative agent of acquired immune deficiency syndrome (AIDS). However, while immunization against the HIV retrovirus is exemplified, such exemplification should not be considered as a limitation on the invention.
BACKGROUND OF THE INVENTION
It is known that immune activation of HIV infected
T4 lymphocytes is required for viral release. These cells act as the source for expansion, and dissemination of virions in the organisms leading to AIDS. (Zagury, et al.. Science. 231, 850-853 (1986)) The viral release which leads to death of infected cells is preceded by a stage where HIV signals are present in the cell membrane. This immunogenic stage is important as a trigger for the cellular reaction against HIV. The reaction leads to the destruction of the infected cells. (Zagury, et al., Proc. Nat. Acad Sci. USA. Vol. 85. pp. 3570-3574 (1988).
Previous immunization protocols have been aimed at establishing a candidate vaccine (rV) expressing HIV proteins followed by a booster of autologous cells infected with rV which have subsequently been fixed. These lead to both humoral and cellular immune responses to HIV. (Zagury, et al., Nature 332, p. 723 (1988).)
The development of a safe and effective vaccine against the AIDS (HIV) virus has become a high priority concern of the scientific and medical community. Rapid progress in the isolation, cloning, and sequencing of the entire genome has shown the remarkable propensity of the HIV strains to mutate, particularly within the viral envelope gene. since viral envelope proteins are often the target for neutralizing antibodies, this extensive variation may play an important role in the interaction between the virus and the host's immune system. For some viruses, such as influenza, rapid mutation is an important means of escape from neutralizing antibodies. Such mutation results in successive waves of influenza epidemics among previously infected populations. For visna virus, a sheep retrovirus, the mutation rate is believed to be so rapid as to allow antibody escape during the course of a single chronic infection. If similar mutants arise in humans infected with viral infections, HIV being one example, during the course of multiple rounds of infection, it would be difficult to imagine a vaccine antigen that could keep pace with all of the possible variants. Under such circumstances it would be impossible to develop an effective vaccine.
In spite of the observed rapid mutation rate of any virus, it is possible that a virus cannot mutate at certain sites, particularly those serving essential viral functions. For example, the CD4 binding site of HIV has been mapped to three relatively conserved regions of gpl20. Divergent isolates bind soluble CD4 and are inactivated by it, suggesting conservation of the CD4 binding site. Presumably, if neutralizing anti.bodies were directed against this site or another site responsible for a critical viral function, such antibodies would be active against numerous clinical isolates of the virus. A vaccine capable of eliciting antibodies against a broad spectrum of HIV strains is needed.
Previous work using retroviral immunogenic signals in their native configuration linked to exogenous carriers and/or in water-soluble adjuvants such as alum is well known. These immunizing preparations did not, however, trigger a significant anti-viral group-specific reaction. The lack of group specific response is particularly important when the infectious agent mutates readily and/or when the species is characterized as composed of a large number of strains showing differing antigenic properties. DESCRIPTION OF THE INVENTION
It is the purpose of the present invention to provide a safe, effective vaccine that will raise antibodies against several parts of the genome of the HIV virus.
It is a further purpose of the invention to provide protocol for administration of a vaccine that causes the production of an immune response which will protect the individual against several strains of HIV virus.
It is also a purpose of the invention to provide an immunotherapy protocol that will cause the patient to develop protective immunity against HIV, which will block viral expansion and dissemination in infected individuals.
It is, additionally, a purpose of the invention to provide new, improved immunogenic compositions.
It has been possible to obtain antibodies against HIV by infecting autologous cells (patient cells) with recombinant virus (rV) carrying HIV segments (envelope, gag, or pol). An example of such a recombinant virus is the virus of Moss. Mackett, et al., J. Virol. 49, 857-864.) Another method of developing immune response to HIV may be accomplished by incubating autologous cells with synthetic peptides to allow peptides to form complexes with HLA antigens on the cell surface, then fixing the cells by usual methods known in the art, e.g., fixation with paraformaldehyde or glutaraldehyde (Zagury, et al, Nature. 332, pp. 728-731 (1988)).
It is also possible to raise an immune response by the administration of compositions containing only the cell-free membranes from autologous cells that have been incubated with the appropriate peptides or have been infected with virus containing recombinant HIV nucleic acid sequences. (See Zagury, Nature, above.) The sequences to which the autologous cells are exposed are usually from the envelope, gag, or pol proteins.
More recently, an immune response has been obtained by the administration of compositions containing "cocktails" of several free peptides representing sequences found in HIV proteins. These peptides in aqueous solution can be added to oils to form an emulsion. These compositions have immunogenic properties and can be administered to individuals either in conjunction with administration of preparations containing treated autologous cells or cell free membranes. However, the peptidecontaining emulsions can also be administered alone as a means of raising an immune response in the individual
The emulsions are administered parenterally, intramuscularly or subcutaneously.
There are several advantages in using the compositions of the present invention either alone or in cσnjunction with a protocol that includes use of autologous cells that have been exposed in vitro to HIV antigens, or cell membranes obtained therefrom. 1) The "peptide cocktail" approach allows the use of selected amino acid sequences. These sequences may be chosen from regions of the protein that are conserved across the various strains of the organism. Additionally, peptide sequences from several stra.ins may be chosen so that a broad range of viral strains may be used to give broad group protection against HIV Vairus.
The peptides may be prepared by any means known in the art, such as by recombinant means or by the Merrifield process. The process of preparing the peptides by synthetic means provides not only reliability, but also provides certain economic advantages.
Peptides should be of about 8 to 40 amino acids, with peptides of 12 to 30 amino acids preferred. In addition to the emulsion containing the peptides, protein fragments having molecular weights of over 10,000 can be added to the emulsion or can be given separately from the peptides to stimulate increased antibody response. Of course, adjuvants and other additives known in the art may also be added to the peptide-containing emulsions. Some of the larger segments that can be used include gp160, gp41, gp566, gp24, reverse transσriptase (RT), Tat protein, and protease.
While peptides from several strains and differing locations in the HIV proteins can be used, particularly preferred sites are listed below:
Y-N-K-R-K-K-I-H-I-G-P-G-R-A-F-Y-T-T-K-N-I-I-G R-I-G-P-G-R-A-F-V-T-I-G-K
Q-K-V-G-K-A-M-Y-A-F-P-I-S-G
D-M-V-E-Q-M-H-E-D-I-I-S-L-W-D-Q-S-L-K-P-C W-G-I-K-Q-L-Q-A-R-I-L-A-V-E-R-Y-L-K-D-Q
C-K-I-K-Q-I-V-K-M-W-Q-C-V-G-Q-A-I-Y
N-T-R-K-S-I-R-I-Q-R-G-P-G-R-A-F-V-T-I-G-K-I-G N-N-T-R-K-S-I-T-K-G-P-G-R-V-I-Y-A-T-G-Q-I-I-G N-N-V-R-R-S-I-S-I-G-P-G-R-A-F-R-T-R-G-K-I-I-G R-I-G-P-G-R-A
G-P-G-R-A-F-V-T-I-G-K
N-Y-T-R-K-S-V-R-I-G-F-G-Q-A-F-Y-A-T-G-D-I-I-G Q-N-T-R-Q-R-T-P-I-G-L-G-Q-S-L-Y-T-T-R-S-R-I-S N-N-T-R-R-G-I-H-F-G-P-F-Q-A-L-Y-T-T-G-I-I-V-G
EXAMPLES
Example 1
AIDS or ARC (aids-related complex) patients with 150 - 400 T4 cells per mm3 were immunized with fixed autologous B cells transformed with EBV (Epstein-Barr Virus) and infected with recombinant vaccinia expressing
HIV proteins (env, gag, and pol) on the surface of infected cells. The fixed cells were given by intravenous slow drip infusion (5-7 × 10] cells), intramuscularly (10
× l01 cells in water suspension in animal oil (squalene, Montanide 708) or subcutaneσusly.
The vaccinia vector used is the non-neurotropio strain Lister and the recombinant is produced by the method of Moss (sea Mackett, supra.) The patients underwent biweekly physical examination. Blood samples were drawn for preparation of serum and cells for biological investigations including virus neutralizing antibody, T4 cell count, cell mediated immunity, and cell mediated cytotoxicity.
Example 2
AIDS or ARC patients with 150 - 400 T4 cells per mm3 received the preparation described in example 1 in conjunction with AZT (600 mg per day Other conditions were similar to those described in example 1.
Example 3
AIDS or ARC patients with 150 - 400 T4 cells per mm3 received treatment as described in example 1 along with discontinuous AZT at low doses (600 mg per day for 30 days every 90 days). Other conditions were similar to those described in example 1.
Example 4
.Asymptomatic HIV infected individuals with >500 T4 cells per mm3 received the treatment described in example 1. Other conditions were similar to those described in example 1.
Example 5
AIDS or ARC patients with 150 - 400 T4 cells per mm3 received a mixture of synthetic HIV peptides comprising peptides which constituted immunodominant sites of env, gag, pol (peptide 342-350 according to Ratner). Immunisation protocols and patient follow up were as described in example 1.
Example 6
Patients described in example 5 were given a continuous low dose of AZT (600 mg/day). Other conditions were similar to those described in example 5.
Example 7
Patients treated as described in example 5 were given a discontinuous low dose of AZT (600 mg/day for 30 days every 90 days). Other conditions were similar to those described in example 5.
Example 8
Asymptomatic HIV infected individuals with >500 T4 cells per mm3 were treated as described in example 5. Example 9
Seronegativepatients were treated as described in example 5 except that the synthetic peptides administered ware protectively encapsulated as a water-in-oil emulsion. All other conditions were as described in example 5.
Example 10
Seronegativepatients wore given synthetic peptides as In example 5 except that the peptides were administered as free peptides.
Example 11
Seronegativepatients were given synthetic peptides representing HIV epitopes wherein the peptides were covalently linked to an immuno-enhancing moiety. All other conditions were as described in example 5.
Example 12
Peptide derivatives having hydrophobic groups consisting of tripalmitoyl cysteine, dipalmitoyl lysine, or a non-viral peptide of alpha helix configuration were administered in accord with the protocol used to administer the peptides in example 5.
Example 13
Peptides were administered in accord with example 5. However, the patients were also given recombinant vectors containing HIV nucleic acid sequences at separate sites in conjunction with the peptides.
Example 14
Peptides were given in a composition containing recombinant live vectors containing HIV nucleic acid sequences mixed with the synthetic peptides which represented HIV immunodominant epitopes protectively encapsulated as a water in oil emulsion. The composition was given in the manner described in example 5.
Materials: Montanide is a product of SEPPIC, a division of Cosmetique-Pharmacie, 70, Champs-Elysees, 75008 Paris, France, and is obtainable therefrom.

Claims

WHAT IS CLAIMED IS;
1. A composition of matter comprising peptides of 8-40 amino acids representing HTV epitopes protectively encapsulated as a water in oil emulsion.
2. The composition of claim 1, wherein the peptides are free peptides.
3. The composition of claim 1, wherein the peptides are covalently linked to an immunologically enhancing moiety.
4. The composition of claim 3, wherein the enhancing moiety is a hydrophobic segment.
5. The composition of claim 4, wherein the hydrophobic segment is selected from the group consisting of tripalmitoyl cysteine, dipalmitoyl lysine, and a nonviral peptide of alpha helix configuration.
6. The composition of claim 1, further comprising a surfactant for stabilizing the emulsion.
7. The composition of claim 1, further comprising HIV immunogenic protein or protein fragments of molecular weight over 10,000.
8. The composition of claim X, further comprising recombinant live vectors which express recombinant proteins or epitopes of HIV.
9. A composition of matter comprising autologous EBV (Epstein-Barr Virus) transformed B-cells infected with recombinant virus expressing HIV epitopes which have been fixed.
10. A composition of matter comprising autologous EBV (Epstein-Barr vixus) transformed B-cells carrying HIV epitopes at the cell surface which have been fixed.
II. A method of inducing immune response to HIV by administration of an immunogenic effective amount of the composition of claim 1.
12. The method of claim 11, wherein a recombinant live vector containing HIV nucleic acid sequences is administered mixed with a peptide-containing emulsion of the composition of claim 1.
13. The method of claim 11, wherein a recombinant vector containing HIV nucleic acid sequence is administered along with at least one peptide at a separate site.
14. The method of claim 11, used as a part of immunotherapy in AIDS and ARC patients.
15. The method of claim 12, used as part of a vaccine protocol for immunoprophylaxis in HIV infected asymptomatic individuals.
16. A method of inducing an immune response by administration of an effective amount of a composition of claim 9 or 10 as a part of immunoprophylaxis or immunotherapy for AIDS and ARC patients.
17. The method of claim 16, wherein there is administered additionally a composition of claim l.
18. The method of claim 11 or 12, which is used as part of a vaccine protocol.
19. The composition of claim 1, further comprising a surfactant to stabilize the emulsion.
20. The composition of claim 1 or 2, further comprising protein or protein fragments of molecular weight over 10, 000, such as Env gp AGO.
21. The composition of claim 1, further comprising recombinant virus which expresses recombinant proteins of HIV.
22. The method of claim 11, wherein the peptides are administered in oil subcutaneously or intramuscularly.
23. A composition comprising free peptides of 8 to 40 amino acids representing HIV epitopes, protectively encapsulated as a water-in-oil emulsion for administration to induce an immune response to HIV.
24. The composition according to claim 23, which further comprises a recombinant live vector containing RΣV nucleic acid sequences;
25. The composition according to claim 23, wherein a recombinant vector containing HIV nucleic acid sequences and at least one peptide is administered separately from said free peptides.
26. The composition according to claim 23, for use as part of immunotherapy in AIDS and ARC (Aids Related complex) patients.
27. The composition according to claim 24, for use as part of the vaccine protocol for immunoprophylaxis in HIV infected asymptomatic individuals.
28. A composition comprising autologous EBV
(Epstein-Barr Virus) transformed B cells infected with recombinant virus expressing HIV epitopes which have been fixed or EBV transformed B cells carrying HIV epitopes at the cell surface which have been fixed, for use in inducing an immune response as part of immunoprophylaxis or .immunotherapy for AIDS and ARC patients.
29. The composition of claim 28, for use with the composition of claim 23 to induce an immune response as part of immunoprophylaxis or immunotherapy for AIDS and ARC patients.
30. The composition of claims 23 and 24, for use as part of a vaccine protocol.
31. Use of a composition comprising free peptides of 8 to 40 amino acids representing HIV epitopes, protectively encapsulated as a water-in-oil emulsion, in the manufacture of a medicament for inducing an immune response to HIV.
32. The use according to claim 31, wherein the composition further comprises a recombinant live vector containing HIV nucleic acid sequences.
33. Use of a composition comprising free peptides of 8 to 40 amino acids representing .HIV epitopes, protectively encapsulated as a water-in-oil emulsion in the manufacture of a medicament for use as part of immunotherapy in AIDS and ARC patients.
34. Use of a composition comprising free peptides of 8 to 40 amino acids representing HIV epitopes protectively encapsulated as a water-in-oil emulsion and a recombinant live vector containing HIV nucleic acid sequences in the manufacture of a medicament as part of a vaccine protocol for immunoprophylaxis in HIV infected asymptomatic individuals.
35. Use of a composition comprising autologous EBV (Epstein-Barr Virus) transformed B cells infected with recombinant virus expressing HIV epitopes which have been fixed or EBV transformed B cells carrying HIV epitopes at the cell surface which have been fixed, in the manufacture of a medicament for inducing an immune response as part of immunoprophylaxis or immunotherapy for AIDS and ARC patients.
PCT/EP1991/001225 1990-06-29 1991-07-01 Methods of inducing immune response to aids virus WO1992000098A1 (en)

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WO1992022577A1 (en) * 1991-06-17 1992-12-23 Neovacs Immunogen compounds having specifically an anti-cytokine effect, method of preparation, pharmaceutical compositions and kits containing them
US7311921B2 (en) * 1995-08-25 2007-12-25 University Of Florida Research Foundation, Inc. Multi-subtype FIV vaccines

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