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  • A61K2039/58—Medicinal preparations containing antigens or antibodies raising an immune response against a target which is not the antigen used for immunisation
  • A61K2039/585—Medicinal preparations containing antigens or antibodies raising an immune response against a target which is not the antigen used for immunisation wherein the target is cancer
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  • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
  • A61K9/167—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction with an outer layer or coating comprising drug; with chemically bound drugs or non-active substances on their surface
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  • C12N2710/16634—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
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  • C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
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  • C12N2760/16134—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
  • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
  • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
  • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

• the invention relates to a method of eliciting a T cell response against a T cell epitope of interest (EOI).
• the method comprises the administration of a NOI which encodes the EOI.
• NOI which encodes the EOI.
• at least 2, 4, 6, 10 or 20 or more (up to and including, for example, 40) different NOI's may be administered, wherein each of the NOI's encode the same epitope.
• the same NOI may be administered.
• a protein comprising the EOI it is understood that at least 2, 4, 10 or more (up to an including, for example 20) different proteins may be administered which comprise the epitope.
• the same protein which comprises the epitope
• the present invention provides a method of eliciting an enhanced cellular mediated immune (CMI) response against at least one target antigen (TA) in a host mammalian subject; wherein the method comprises administering a nucleotide sequence of interest (NOI) encoding one or more epitopes of interest (EOI) of the TA at least twice to the host mammalian subject; wherein the intervals between each NOI administrations ranges from about 48 hours to about 144 hours; and wherein the method is effective to provide an enhanced CMI response against the or each expressed EOI in the host mammalian subject.
• CMI cellular mediated immune
• TA target antigen
• NOI nucleotide sequence of interest
• EOI epitopes of interest
• humoral immune response refers to an immune response mediated by antibody molecules.
• the antibodies generated by humoral immunity are primarily effective against extracellular infectious agents.
• the term "cell mediated immune (CMI) response” is one mediated by T- lymphocytes and/or other white blood cells.
• the CMI immune mechanisms are generally more effective against intracellular infections and disease because the CMI mechanisms prime T cells in a way that, when a TA appears at a later date, memory T cells are activated to result in a CMI response that destroys target cells that have the corresponding TA or a portion thereof on their cell surfaces, and thereby the infecting pathogen.
• the CMI response is focused on the destruction of the source of infection mediated by either effector cells that destroy infected cells of the host by direct cell-to-cell contact and/or by the release of molecules, such as cytokines, that possess anti-viral activity.
• the CMI response which is characterised by a specific T lymphocyte cellular response, is crucial to produce resistance to diseases caused by cancer, viruses, pathogenic and other intracellular microorganisms.
• eliciting a response is often discussed in terms of eliciting a CMI response.
• the eliciting of a CMI response is understood to include eliciting of a T cell response.
• the response which is elicited by the method of the invention may be an "enhanced" response.
• An “enhanced” response may be said to occur if the response elicited by the method of the invention is more than the response which is elicited in a control method, where in the control method the same amount of NOI (and if applicable also the same amount of protein) has been administered as was administered in the method of the invention.
• a control method may for example consist of administration of the NOI (and if applicable also protein) in a single administration.
• the control method may consist of administration of the NOI (and if applicable also protein) in two separate administrations which are 28 days apart.
• the term "enhancing a T -cell response" encompasses improvements in all aspects of the T-cell response which include but are not limited to a stimulation and/or augmentation and/or potentiation and/or up-regulation of the magnitude and/or duration, and/or quality of the T-cell response to a repeatedly administered NOI encoding an EOI of a target antigen.
• the T-cell response may be enhanced by either enhancing the activation and/or production and/or distribution and/or proliferation of the induced T-cells and/or longevity of the T-cell response to T-cell inducing/modulating NOIs encoding EOIs from a TA.
• the enhancement of the T-cell response in a host subject may be associated with the enhancement and/or modulation of the Thl immune response in the host subject.
• target antigen means an immunogenic peptide or protein of interest comprising one or more epitopes capable of inducing a CMI response to an infectious pathogen such as but not limited to a bacteria, viruses, fungi, yeast, parasites and other microorganisms capable of infecting mammalian species.
• the target antigen can include but is not limited to an auto-antigen, a self-antigen, a cross-reacting antigen, an alloantigen, a tolerogen, an allergen, a hapten, an immunogen or parts thereof as well as any combinations thereof.
• the EOI may be from any of the types of antigens or proteins mentioned herein or from any of the specific antigens or proteins mentioned herein. EPITOPE
• Peptide length the peptide should be at least 8 or 9 amino acids long to fit into the MHC class I complex and at least 8-25, such at least as 13-25 amino acids long to fit into a class II MHC complex. This length is a minimum for the peptide to bind to the MHC complex. It is preferred for the peptides to be longer than these lengths because cells may cut peptides.
• the peptide should contain an appropriate anchor motif which will enable it to bind to the various class I or class II molecules with high enough specificity to generate an immune response (See Bocchia, M.
• the method of the present invention is generally applicable to enhancing the CMI response against NOIs encoding EOIs from any source (for example from a pathogen), including those from a wide variety of infectious agents such as viruses or parasites.
• the EOI may be derived from pathogenic agents derived from tumour cells which multiply unrestrictedly in an organism and may thus lead to pathological growths. Examples of such pathogenic agents are described in Davis, B.D. et al (Microbiology, 3rd ed., Harper International Edition).
• the EOI of the TA may contain one or more T cell epitopes.
• T cell epitope refers generally to those features of a peptide structure which are capable of inducing a T cell response.
• T cell epitopes comprise linear peptide determinants that assume extended conformations within the peptide-binding cleft of MHC molecules (Unanue et al. (1987) Science 236: 551-557).
• a T cell epitope is generally a peptide having at least about 3-5 amino acid residues, and preferably at least 5-10 or more amino acid residues, such as 8 to 25 amino acid residues.
• the genetic adjuvant is cholera toxin (CT), enterotoxigenic E. Coli heat-labile toxin (LT), or a derivative, subunit, or fragment of CT or LT which retains adjuvanticity.
• CT cholera toxin
• LT enterotoxigenic E. Coli heat-labile toxin
• the genetic adjuvant is LT.
• the genetic adjuvant may be CTB or LTB.
• a viral promoter is used to drive expression from the NOI.
• the promoter may be a Cytomegalovirus (CMV) promoter.
• CMV Cytomegalovirus
• a preferred promoter element (particularly in the case where the NOI encodes an HIV antigen) is the CMV immediate early (IE) promoter devoid of intron A, but including exon 1.
• IE CMV immediate early
• the NOI or protein is administered via the skin, for example by a transdermal route. While it is believed that any accepted mode and route of immunization can be employed and nevertheless achieve some advantages in accordance herewith, the Examples below demonstrate particular advantages with transdermal NOI administration. In this regard, and without being bound by theory, it is believed that transdermal administration is preferred because it more efficiently activates the cell mediated arm of the immune system.
• an appropriate dose for prophylactic or therapeutic use in an adult of the human species is from about 1 pg to about 5 mg, preferably from about 10 pg to about 1 mg, most preferably from about 25 pg to about 500 pg.
• Particle-mediated delivery techniques have been compared to other types of NOI administration, and found markedly superior. Fynan et al. (1995) Int. J. Immunopharmacology 17:79-83, Fynan et al. (1993) Proc. Natl. Acad. Sci. USA 90:11478-11482, and Raz et al. (1994) Proc. Natl. Acad. Sci. USA 91:9519- 9523.
• substantially homologous or homologous also refers to sequences showing complete identity to the specified DNA or polypeptide sequence.
• DNA sequences that are substantially homologous or homologous can be identified in a Southern hybridization experiment under, for example, stringent conditions, as defined for that particular system.
• stringent hybridization conditions can include 50% formamide, 5x Denhardt's Solution, 5x SSC, 0.1% SDS and 100 pg/ml denatured salmon sperm DNA and the washing conditions can include 2x SSC, 0.1% SDS at 37 C followed by lx SSC, 0.1% SDS at 68 C. Defining appropriate hybridization conditions is within the skill of the art.
• Such nodes are physically located in the proximity of the site of NOI administration or in the area draining the site of administration, and also included are those draining nodes that are physically at a greater distance from the administration site.
• a second immunisation that comprises at least one administration to the subject of (a) a NOI encoding the T cell epitope, or (b) a protein comprising the T cell epitope, wherein the time between - the first administration of the first immunisation, and - the first administration of the second immunisation, is from 21 to 365 days, wherein the assay comprises carrying out the method on a mammalian subject and then determining the level of activated or memory T cells specific to the epitope in the subject.
• Figure 6A shows IFN- ⁇ ELISPOT data obtained from domestic pigs following the first cluster immunization.
• Figure 6B shows the average area of erythema present at the site of antigen administration in pigs (4 animals) immunized with pPJV7630.
• Figure 14 (i) to (viii) provides the Feature Maps of Key Plasmids in the Construction of pPJV7563.
• any particular plasmid vector or other DNA vector or viral vector is not a limiting factor in this invention and other DNA or viral vectors may be substituted for those disclosed herein upon a reading of the present disclosure. It is well within the skill of the artisan to choose particular promoter/regulatory sequences and operably link those promoter/regulatory sequences to a DNA sequence encoding a desired antigen. Such technology is well known in the art and is described, for example, in Sambrook (ibid), and Ausubel (ibid).
• NOIs comprising EOIs were coated onto gold particles in order to provide exemplary compositions according the present invention.
• the coated particles were administered to animal subjects, and the ability of the compositions to elicit antigen specific T cell and/or antibody responses was assessed.
• HBsAg vector plasmid was constructed as follows. To generate the HbsAg coding region, the pAM6 construct (obtained from the American Type Culture Collection "ATCC") was cut with Neol and treated with mung bean nuclease to remove the start codon of the X-antigen. The resultant D ⁇ A was then cut with BamRI and treated with T4 D ⁇ A polymerase to blunt-end the D ⁇ A and creates an HBsAg expression cassette. The HBsAg expression cassette is present in the 1.2 kB fragment. The plasmid construct pPJV7077 (Schmaljohn et al. (1997) J. Virol.
• cells from na ⁇ ve animals were used to bring the total to lxl 0 6 .
• Cells were incubated overnight in a tissue culture incubator in the presence of the peptide as described above. The plates were then washed 2 times with PBS and 1 time with distilled water. This was followed by 3 washes with PBS.
• a biotinylated anti IFN- ⁇ monoclonal antibody (Pharmingen) was added to the plate (50 ⁇ l of a 1 ⁇ g/ml solution in PBS) and incubated for 2 hr at RT.
• the plasmid encoding HIV-1 gpl20 was then constructed as follows.
• the pCIA-Env/T plasmid encodes a truncated form of HIV-1 gpl60, and is identical to the pC-Env construct except that the env coding sequences are truncated at the HindUI site at nucleotide position 8188. This results in a truncated gpl60 translation product with the truncation point lying 128 amino acid residues downstream of the gpl20/gp41 processing site.
• DNA antigen plasmid With respect to the construction of the DNA antigen plasmid, standard PCR techniques were used to construct the plasmid.
• the standard PCR conditions that were used for the construction of the vector were as follows: lx PCR core buffer with 1.5 mM MgCl 2 (Promega Corp., Madison, Wl); 0.400 ⁇ M of each primer; 200 ⁇ M of each dNTP (USB Inc., Cleveland, OH); 2.5 ⁇ g Taq polymerase (Promega Corp., Madison, Wl); 1.0 ng template DNA; water to 100 ⁇ l; and a mineral oil (Aldrich Chemical Inc., Milwaukee Wl) overlay.
• the purpose of this experiment was to vary the time interval between DNA administrations of a single gene plasmid.
• TC-1 cells were obtained from Johns Hopkins School of Medicine. Cells were expanded in culture for a minimum number of passages and vials were then frozen and stored in liquid N 2 until use. Cells were prepared for injection in PBS. Anesthetized mice were injected subcutaneously with between 2 X IO 4 to 2 X IO 5 cells in 50 to 100 ul on the shaved right flank. Tumors were measured Mon., Wed., and Fri. beginning at day 7. Two diameter measurements at right angles were taken and multiplied to produce a square area. The health of the animals was also monitored. Mice were euthanized if tumors grew to greater than 120 mm 2 , if tumors appeared necrotic, or mice appeared moribund.
• mice were sacrificed one week after last immunization. Spleens were removed aseptically and single cell suspensions were prepared. Cells were plated at 1 X IO 6 or 5 X 10 5 cells per well in BD (-IFN ELISPOT kits according to manufacturers instructions. Peptides specific for E7 CD4 and CD8 and E6 CD8 were added at a final concentration of 10 uM. Media wells contained equivalent amounts of DMSO as wells containing peptide. Specific spots were calculated by subtracting media spots from those induced in the presence of peptide.
• mice were given 1,2,3 or 4shots of pPJV7630 (using 4 day intervals between shots) and then cells from the lymph nodes examined 8 days later. Results are shown in Figure 30. With increasing number of shots, increases in the size of nodes was apparent. Weights and cell numbers in lymph nodes measured 8 days after completion of vaccine delivery were increased over na ⁇ ve mice when more than 1 shot was given with 3 shots giving the highest values.

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• 2004
  • 2004-10-12 EP EP04794672A patent/EP1675596A4/en not_active Withdrawn
  • 2004-10-12 WO PCT/US2004/033391 patent/WO2005035779A2/en not_active Ceased
  • 2004-10-12 MX MXPA06003977A patent/MXPA06003977A/es not_active Application Discontinuation
  • 2004-10-12 KR KR1020067009022A patent/KR20060123138A/ko not_active Ceased
  • 2004-10-12 SG SG200806935-3A patent/SG146662A1/en unknown
  • 2004-10-12 JP JP2006534429A patent/JP2007508319A/ja not_active Withdrawn
  • 2004-10-12 EA EA200600738A patent/EA012066B1/ru not_active IP Right Cessation
  • 2004-10-12 BR BRPI0415202-6A patent/BRPI0415202A/pt not_active IP Right Cessation
  • 2004-10-12 AU AU2004280630A patent/AU2004280630A1/en not_active Abandoned
  • 2004-10-12 US US10/574,922 patent/US20070026015A1/en not_active Abandoned
  • 2004-10-12 NZ NZ547042A patent/NZ547042A/en unknown
  • 2004-10-12 CA CA002542295A patent/CA2542295A1/en not_active Abandoned
  • 2004-10-12 CN CNA2004800364591A patent/CN1889963A/zh active Pending
• 2006
  • 2006-04-06 IL IL174849A patent/IL174849A0/en unknown
  • 2006-05-09 NO NO20062081A patent/NO20062081L/no not_active Application Discontinuation

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