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  • 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
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  • C12N7/00—Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof
• • 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
  • A61K2039/58—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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  • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
  • C12N2710/00011—Details
  • C12N2710/20011—Papillomaviridae
  • C12N2710/20022—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
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  • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
  • C12N2710/00011—Details
  • C12N2710/20011—Papillomaviridae
  • C12N2710/20034—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
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  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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  • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
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  • C12N2710/20011—Papillomaviridae
  • C12N2710/20061—Methods of inactivation or attenuation
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  • C12N2710/00011—Details
  • C12N2710/20011—Papillomaviridae
  • C12N2710/20071—Demonstrated in vivo effect

Definitions

• Mucosal carcinomas including mucosal squamous cell carcinoma (mSCC) constitute a common type of cancer affecting the upper aerodigestive tract, anus and cervix 1 .
• High-risk human papillomavirus (HPV) is the cause of a subset of head and neck cancer (HNC), anal and cervical cancers.
• vaccines based on commensal HPVs are beneficial in protecting against mucosal cancers.
• compositions described herein for use in a method of treating, or reducing the risk of developing, mucosal cancer in a subject.
• the subject has an increased risk of developing mucosal cancer or is immunocompromised, e.g., as a result of an acquired immunodeficiency, primary immunodeficiency, or an organ transplant.
• the commensal human papilloma viruses are low risk a-HPV, P-HPV, y-HPV, and/or p-HPV strains, e.g., the commensal human papilloma viruses are P-HPV and/or y-HPV strains listed in Table A.
• the plurality of antigenic peptides comprises peptides derived from one or more El, E2, E4, E5, E6 or E7 proteins.
• the plurality of antigenic peptides comprises peptides derived from proteins from a plurality of commensal human papilloma viruses.
• compositions comprise at least 200 peptides each having a unique sequences, e.g., comprising a plurality of peptides for each unique sequence.
• the T cell adjuvant comprises one or more of nanoparticles that enhance T cell response; poly-ICLC (carboxymethylcellulose, polyinosinic-polycytidylic acid, and poly-L-lysine double-stranded RNA), Imiquimods, CpG oligodeoxynuceotides and formulations (IC31, QB 10), AS04 (aluminium salt formulated with 3-O-desacyl-4'-monophosphoryl lipid A (MPL)), AS01 (MPL and the saponin QS-21), MPLA, STING agonists, other TLR agonists, Candida albicans Skin Test Antigen (Candin), GM-CSF, Fms-like tyrosine kinase-3 ligand (Flt3L), and/or IFA (Incomplete Freund’s adjuvant).
• poly-ICLC carboxymethylcellulose, polyinosinic-polycytidylic acid, and poly-L-lysine
• the T cell adjuvant comprises topical imiquimod and/or topical 5 -fluorouracil and/or topical calcipotriene (calcipotriol), e.g., in combination with 5 -fluorouracil.
• the mucosal cancer is a cancer of the oral and sinonasal mucosa, optionally mucosal squamous cell carcinoma (mSCC), e.g., of the tongue; head and neck cancer (HNC); a cancer of the mucosa of the upper respiratory tract; or a cancer of the genitourinary tract, optionally anal cancer, cervical cancer, or a cancer of the vulva, vestibule, vagina, perineum, or perianal tissue.
• mSCC mucosal squamous cell carcinoma
• HNC head and neck cancer
• a cancer of the mucosa of the upper respiratory tract or a cancer of the genitourinary tract, optionally anal cancer, cervical cancer, or a cancer of the vulva, vestibule, vagina, perineum, or perianal tissue.
• FIGs. 1A-C Papillomavirus colonization protects against carcinogen- induced tongue cancer.
• FIG. 1 A Time to tumor onset on MmuP V 1 - versus sham- infected tongue of immunocompetent p53 +/ " mice after treatment with DMBA/4NQO carcinogenesis protocol (log-rank test).
• FIG. IB Survival ofMmuPVl- versus sham-infected immunocompetent p53 +/ " mice after treatment with DMBA/4NQO carcinogenesis protocol (log-rank test). Note that the burden of oral lesions induced by carcinogens is the cause of weight loss and mortality in the animals.
• FIG. 1 A Time to tumor onset on MmuP V 1 - versus sham- infected tongue of immunocompetent p53 +/ " mice after treatment with DMBA/4NQO carcinogenesis protocol (log-rank test).
• FIG. 1 B Survival ofMmuPVl- versus sham
• FIGs. 2A-B Papillomavirus colonization protects against the expansion of carcinogen-induced mutant Trp53 clones in tongue epithelium.
• FIG. 1A The size of mutant Trp53 clones in MmuPVl- versus sham-infected tongue of immunocompetent p53 +/ " mice after treatment with DMBA/4NQO carcinogenesis protocol (Mann Whitney U test).
• FIG. IB Representative p53-stained images of MmuPVl- (top) and sham-infected (bottom) tongue treated with DMBA/4NQO. Red arrow points to the mutant Trp53 clones in MmuPVl - and sham-infected tongue.
• High-risk HPV vaccines that are directed at humeral immunity reduce the risk of mSCC by blocking a-HPV infection.
• an effective modality to prevent the large majority of HNCs caused by smoking and mSCCs in individuals who are already infected with high-risk HPVs is lacking.
• Treatments for mSCC include surgery and radiation for localized disease, and chemotherapy, targeted therapy and immunotherapy for unresectable and metastatic cancers.
• the treatments for mSCC represents a rising public health challenge. Therefore, new technologies to prevent and treat mSCC are urgently needed.
• High-risk a-HPVs are known to induce oropharyngeal mSCCs 2 ; however, carcinogen-induced HNCs constitute more than 70% of mSCCs that affect the upper aerodigestive tract 3 .
• low-risk commensal HPVs are ubiquitously found in the upper aerodigestive tract, anal canal and reproductive tract 4,5 . Although they do not make any oncogenic contribution to mSCC development, the impact of immunity against commensal HPVs on mSCC development is unexplored.
• MmuPVl skin colonization of immunocompetent mice had no oncogenic effect but instead protected the skin from chemical as well as UV skin carcinogenesis in a CD8 + T cell-dependent manner.
• the adoptive transfer of memory T cells from skin draining lymph nodes of the immune mice rendered immunity to wart-bearing mice and provided them with protection against skin carcinogenesis 6 .
• commensal HPV vaccines can also be used for cancer prevention and therapy in mucosal sites, as papillomavirus colonization of the oral cavity protected immunocompetent hosts from mSCC.
• compositions that can be used to induce a T cell-based immune response against P-HPVs, thereby reducing the risk that the subject will develop mucosal cancer.
• the vaccines induce T cell immunity against commensal viruses that have already infected the tissue, with the goal not to prevent or eliminate the infection but rather to use of the virus presence in all cells to boost the detection of early cancerous clones and their elimination by T cells.
• Current high-risk HPV vaccines for cervical and head and neck cancer prevention are meant to prevent infection in the first place and have minimal efficacy in individuals already infected with the virus.
• Antigenic peptides are meant to prevent infection in the first place and have minimal efficacy in individuals already infected with the virus.
• the present compositions include a plurality of antigenic peptides derived from (i.e., comprising a fragment of, i.e., consecutive amino acids from) proteins, e.g., El, E2, E6, or E7 proteins, from commensal human
• compositions do not include peptides derived from HPV types that are associated with cancer, e.g., high-risk HPVs such as HPV16 or 18. See, e.g., Ma et al., J Virol. 2014 May; 88(9): 4786-4797; Doorbar et al., Rev Med Virol.
• the peptides can be derived from any antigenic protein in the virus; in some embodiments, the peptides are derived from an El, E2, E4, E5, E6 or E7 protein. Sequences for these proteins in a number of commensal strains are provided. In some embodiments, at least 50 or more, 100, 150, 200, 250, 300, 350, 400, 450, 500, or more different peptides (i.e., peptides having different sequences) are included in the compositions. In some embodiments, at least 50 or more, 100, 150, 200, 250, 300, 350, 400, 450, 500, or more different peptides from each virus strain are included in the compositions, and peptide from two or more virus strains are included.
• the peptides are of a length that is optimized for MHCI/MHCII presentation, e.g., 9-30 amino acids, e.g., 12-25, 12-18, 12-16, 13-16, 14-16, or 15 amino acids.
• the sequences of the peptides can be synthetic long overlapping peptides, e.g., identified, e.g., bioinformatically to predict antigenicity and/or generated using a moving window of overlapping peptides to cover the entire protein, e.g., 15 amino acid peptides with 10 amino acid overlap (similar to the “gene walk” methods used to identify optimal antisense oligonucleotides).
• overlapping synthetic long peptides are used (Zom et al., Cancer Immunol Res. 2014 Aug;2(8):756-64).
• the compositions can include a plurality of peptides derived from one or more (e.g., a plurality of) different virus strains.
• the peptides are preferably synthetic peptides; methods for synthesizing peptides are known in the art, including solution-phase techniques and solid-phase peptide synthesis (SPPS). See, e.g., Petrou and Sarigiannis, Ch.
• the present compositions can include a plurality of proteins, e.g., virus-like particles containing of El, E2, E6, or E7 proteins from commensal human papilloma viruses, e.g., low risk a-HPV, P-HPV, y-HPV and/or p- HPV strains such as those listed in Table A (see, e.g., Yang et al., Virus Res 231, 148- 165 (2017); Hancock et al., Therapeutic HPV vaccines. Best Pract Res Clin Obstet Gynaecol 47, 59-72 (Feb. 2018); Joh et al., Exp Mol Pathol. ;93(3):416-21 (2012)).
• Nucleic acid-based vaccines e.g., virus-like particles containing of El, E2, E6, or E7 proteins from commensal human papilloma viruses, e.g., low risk a-HPV, P-HPV, y-HP
• the present compositions can include a plurality of DNA plasmids and/or RNA replicons that contain nucleotide sequences to express proteins or antigenic peptides derived from (i.e., comprising a fragment of, i.e., consecutive amino acids from) proteins, e.g., El, E2, E6, or E7 proteins, from commensal human papilloma viruses, e.g., low risk a-HPV, P-HPV, y-HPV and/or p- HPV strains such as those listed in Table A (see, e.g., Yang et al., Virus Res 231, 148- 165 (2017); Hancock et al., Therapeutic HPV vaccines. Best Pract Res Clin Obstet Gynaecol 47, 59-72 (2018)).
• the present compositions can include a plurality of viral vectors that are engineered to express proteins or antigenic peptides derived from (i.e., comprising a fragment of, i.e., consecutive amino acids from) proteins, e.g., El, E2, E6, or E7 proteins, from commensal human papilloma viruses, e.g., low risk a-HPV, P-HPV, y-HPV and/or p-HPV strains such as those listed in Table A (see, e.g., Yang et al., Virus Res 231, 148-165 (2017); Hancock et al., Therapeutic HPV vaccines. Best Pract Res Clin Obstet Gynaecol 47, 59-72 (2018)).
• proteins or antigenic peptides derived from (i.e., comprising a fragment of, i.e., consecutive amino acids from) proteins, e.g., El, E2, E6, or E7 proteins, from commensal human
• Viral vectors for use in the present methods and compositions include recombinant retroviruses, adenovirus, adeno-associated virus, alphavirus, and lentivirus.
• a preferred viral vector system useful for delivery of nucleic acids in the present methods is the adeno-associated virus (AAV).
• AAV is a tiny non-enveloped virus having a 25 nm capsid. No disease is known or has been shown to be associated with the wild type virus.
• AAV has a single-stranded DNA (ssDNA) genome.
• ssDNA single-stranded DNA
• AAV has been shown to exhibit long-term episomal transgene expression, and AAV has demonstrated excellent transgene expression in numerous tissues including the brain, particularly in neurons.
• Vectors containing as little as 300 base pairs of AAV can be packaged and can integrate. Space for exogenous DNA is limited to about 4.7 kb.
• An AAV vector such as that described in Tratschin et al., Mol. Cell. Biol.
• 5:3251-3260 (1985) can be used to introduce DNA into cells.
• a variety of nucleic acids have been introduced into different cell types using AAV vectors (see for example Hermonat et al., Proc. Natl. Acad. Sci. USA 81 :6466-6470 (1984); Tratschin et al., Mol. Cell. Biol. 4:2072-2081 (1985); Wondisford et al., Mol. Endocrinol. 2:32-39 (1988); Tratschin et al., J. Virol. 51 :611-619 (1984); and Flotte et al., J. Biol. Chem. 268:3781-3790 (1993).
• AAV9 has been shown to efficiently cross the blood-brain barrier.
• AAV capsid can be genetically engineered to increase transduction efficient and selectivity, e.g., biotinylated AAV vectors, directed molecular evolution, self-complementary AAV genomes and so on.
• AAV1 is used.
• retrovirus vectors and adeno-associated virus vectors can be used as a recombinant gene delivery system for the transfer of exogenous genes in vivo, particularly into humans. These vectors provide efficient delivery of genes into cells, and the transferred nucleic acids are stably integrated into the chromosomal DNA of the host.
• packaging cells which produce only replication-defective retroviruses has increased the utility of retroviruses for gene therapy, and defective retroviruses are characterized for use in gene transfer for gene therapy purposes (for a review see Miller, Blood 76:271 (1990)).
• a replication defective retrovirus can be packaged into virions, which can be used to infect a target cell through the use of a helper virus by standard techniques. Protocols for producing recombinant retroviruses and for infecting cells in vitro or in vivo with such viruses can be found in Ausubel, et al., eds., Current Protocols in Molecular Biology, Greene Publishing Associates, (1989), Sections 9.10-9.14, and other standard laboratory manuals. Examples of suitable retroviruses include pLJ, pZIP, pWE and pEM which are known to those skilled in the art. Examples of suitable packaging virus lines for preparing both ecotropic and amphotropic retroviral systems include 'PCrip, 'PCre, TU and v FAm.
• Retroviruses have been used to introduce a variety of genes into many different cell types, including epithelial cells, in vitro and/or in vivo (see for example Eglitis, et al. (1985) Science 230: 1395-1398; Danos and Mulligan (1988) Proc. Natl. Acad. Sci. USA 85:6460-6464; Wilson et al. (1988) Proc. Natl. Acad. Sci. USA 85:3014-3018; Armentano et al. (1990) Proc. Natl. Acad. Sci. USA 87:6141-6145; Huber et al. (1991) Proc. Natl. Acad. Sci.
• adenovirus-derived vectors The genome of an adenovirus can be manipulated, such that it encodes and expresses a gene product of interest but is inactivated in terms of its ability to replicate in a normal lytic viral life cycle. See, for example, Berkner et al., BioTechniques 6:616 (1988); Rosenfeld et al., Science 252:431-434 (1991); and Rosenfeld et al., Cell 68: 143-155 (1992).
• adenoviral vectors derived from the adenovirus strain Ad type 5 dl324 or other strains of adenovirus are known to those skilled in the art.
• Recombinant adenoviruses can be advantageous in certain circumstances, in that they are not capable of infecting nondividing cells and can be used to infect a wide variety of cell types, including epithelial cells (Rosenfeld et al., (1992) supra).
• the virus particle is relatively stable and amenable to purification and concentration, and as above, can be modified so as to affect the spectrum of infectivity.
• introduced adenoviral DNA (and foreign DNA contained therein) is not integrated into the genome of a host cell but remains episomal, thereby avoiding potential problems that can occur as a result of insertional mutagenesis in situ, where introduced DNA becomes integrated into the host genome (e.g., retroviral DNA).
• the carrying capacity of the adenoviral genome for foreign DNA is large (up to 8 kilobases) relative to other gene delivery vectors (Berkner et al., supra; Haj-Ahmand and Graham, J. Virol. 57:267 (1986).
• Alphaviruses can also be used. Alphaviruses are enveloped single stranded RNA viruses that have a broad host range, and when used in gene therapy protocols alphaviruses can provide high-level transient gene expression. Exemplary alphaviruses include the Semliki Forest virus (SFV), Sindbis virus (SIN) and Venezuelan Equine Encephalitis (VEE) virus, all of which have been genetically engineered to provide efficient replication-deficient and -competent expression vectors. Alphaviruses exhibit significant neurotropism, and so are useful for CNS- related diseases. See, e.g., Lundstrom, Viruses. 2009 Jun; 1(1): 13-25; Lundstrom, Viruses.
• a live commensal HPV vaccine strategy can be used to optimally boost antiviral T cell immunity in the mucosa to prevent cancer development and treat early mucosal cancers or precancerous lesions with active virus, which include adenocarcinoma in situ, intraepithelial lesions of the vagina, penis and anus; squamous intraepithelial lesions (SILs); cervical intraepithelial neoplasia (CIN); and dysplasia and carcinoma in situ.
• Active virus include adenocarcinoma in situ, intraepithelial lesions of the vagina, penis and anus; squamous intraepithelial lesions (SILs); cervical intraepithelial neoplasia (CIN); and dysplasia and carcinoma in situ.
• SILs squamous intraepithelial lesions
• CIN cervical intraepithelial neoplasia
• Dyplasia and carcinoma in situ Platforms to generate and expand
• compositions can also include an adjuvant to increase T cell response.
• an adjuvant to increase T cell response can be included, e.g., as described in Stano et al., Vaccine (2012) 30:7541-6 and Swaminathan et al., Vaccine (2016) 34: 110-9. See also Panagioti et al., Front. Immunol., 16 February 2018; doi.org/10.3389/fimmu.2018.00276.
• an adjuvant comprising poly-ICLC (carboxymethylcellulose, polyinosinic-poly cytidylic acid, and poly-L-lysine double-stranded RNA), Imiquimod, Resiquimod (R-848), CpG oligodeoxynuceotides and formulations (IC31, QB 10), AS04 (aluminium salt formulated with 3-O-desacyl-4'-monophosphoryl lipid A (MPL)), AS01 (MPL and the saponin QS-21), MPLA, STING agonists, other TLR agonists, GM-CSF, Fms-like tyrosine kinase-3 ligand (Flt3L), and/or IF A (Incomplete Freund’s adjuvant) can also be used.
• ICLC carboxymethylcellulose, polyinosinic-poly cytidylic acid, and poly-L-lysine double-stranded RNA
• Imiquimod
• topical imiquimod and/or topical 5 -fluorouracil and/or topical calcipotriene (calcipotriol) in combination with 5 -fluorouracil could serve as adjuvants for the vaccine (this would be particularly applicable in subjects with pre- malignant lesions, who are commonly treated with these topical agents). See, e.g., Khong and Willem, Journal for ImmunoTherapy of Cancer 4:56 (2016); Coffman et al., Immunity.
• compositions typically include a pharmaceutically acceptable carrier.
• pharmaceutically acceptable carrier includes saline, solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration.
• compositions are typically formulated to be compatible with its intended route of administration.
• routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, intratumoral, intramuscular or subcutaneous administration.
• solutions or suspensions used for parenteral, intradermal, intramuscular, or subcutaneous application can include the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide.
• the parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.
• compositions suitable for injectable use can include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion.
• suitable carriers include physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, NJ) or phosphate buffered saline (PBS).
• the composition must be sterile and should be fluid to the extent that easy syringability exists. It should be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi.
• the carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyetheylene glycol, and the like), and suitable mixtures thereof.
• the proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants.
• Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like.
• isotonic agents for example, sugars, polyalcohols such as mannitol, sorbitol, sodium chloride in the composition.
• Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent that delays absorption, for example, aluminum monostearate and gelatin.
• Sterile injectable solutions can be prepared by incorporating the active compound in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filtered sterilization.
• dispersions are prepared by incorporating the active compound into a sterile vehicle, which contains a basic dispersion medium and the required other ingredients from those enumerated above.
• the preferred methods of preparation are vacuum drying and freeze-drying, which yield a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.
• the therapeutic compounds are prepared with carriers that will protect the therapeutic compounds against rapid elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems.
• a controlled release formulation including implants and microencapsulated delivery systems.
• Biodegradable, biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid.
• Such formulations can be prepared using standard techniques, or obtained commercially, e.g., from Alza Corporation and Nova Pharmaceuticals, Inc.
• Liposomal suspensions (including liposomes targeted to selected cells with monoclonal antibodies to cellular antigens) can also be used as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in U.S. Patent No. 4,522,811.
• compositions can be included in a container, pack, or dispenser together with instructions for administration.
• the vaccine compositions described herein can be used to boost immunity against mucosal cancer in immunocompetent subjects, as well as immunosuppressed or immunocompromised patients who have reduced T cell immunity against P-HPVs and are prone to developing multiple mucosal warts and cancers (loaded with virus) with poor prognosis.
• the subjects do not have cancer (e.g., do not have mucosal cancer).
• the subjects are at high risk (i.e., have a risk that is above that of the general population) of developing mucosal cancer.
• the subject may have a family history of cancer, a personal history of smoking, alcohol use, ionizing radiation, occupational exposures (e.g., acid mists), diet exposure (e.g., salted fish), precancerous lesions, relevant sexual history, or a family history or personal history of mucosal cancer.
• the subjects have, or have a history of, recurrent respiratory papillomatosis.
• the subject may be immunosuppressed, e.g., due to an organ transplant, an acquired immunodeficiency, e.g., HIV/AIDS, or primary human immunodefi ci ency .
• Subjects who can be treated using the present methods include mammals, e.g., human and non-human veterinary subjects.
• the present compositions can be used to induce anti-cancer immunity, to reduce the risk of developing mucosal cancer, i.e., a cancer of a mucosal tissue.
• Mucosal cancers include cancers of the oral and sinonasal mucosa including mucosal squamous cell carcinoma (mSCC); head and neck cancer (HNC); cancers of the mucosa of the upper respiratory tract; and cancers of the genitourinary tract, including anal cancer, cervical cancer, and cancers of the vulva, vestibule, vagina, perineum and perianal.
• the methods include administering one or more doses of the vaccine compositions described herein to a subject, e.g., a subject in need thereof.
• compositions are administered in an effective amount.
• An “effective amount” is an amount sufficient to effect beneficial or desired results.
• an effective amount is one that achieves a desired therapeutic effect, e.g., an amount necessary to treat a disease, or to reduce risk of development of disease or disease symptoms (also referred to as a therapeutically effective amount or a prophylactically effective amount, respectively).
• An effective amount can be administered in one or more administrations, applications or dosages.
• a therapeutically effective amount of a therapeutic compound i.e., an effective dosage) depends on the therapeutic compounds selected.
• the compositions can be administered one from one or more times per day to one or more times per week; including once every other day.
• treatment of a subject with a therapeutically effective amount of the therapeutic compounds described herein can include a single treatment or a series of treatments.
• the methods can include administering a first dose, followed by a second dose at a later time (e.g., a “booster” dose), e.g., at 1, 2, 4, 6, 8, 12, 18, 24, or 52 weeks later.
• Dosage, toxicity and therapeutic efficacy of the therapeutic compositions can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective in 50% of the population).
• the dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD50/ED50.
• Compositions that exhibit high therapeutic indices are preferred. While compositions that exhibit toxic side effects may be used, care should be taken to minimize and reduce side effects.
• the data obtained from cell culture assays and animal studies can be used in formulating a range of dosage for use in humans.
• the dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity.
• the dosage may vary within this range depending upon the dosage form employed and the route of administration utilized.
• the therapeutically effective dose can be estimated initially from cell culture assays.
• a dose may be formulated in animal models. Such information can be used to more accurately determine useful doses in humans.
• the methods can also include administration of one or more other treatments known in the art for mucosal cancer, e.g., in subjects who have mucosal cancer, or treatment to reduce the risk of developing mucosal cancer.
• a combination treatment with the compositions described herein plus treatments for benign or precancerous lesions e.g., surgical treatments (e.g., excision (e.g., loop electrosurgical excision procedure (LEEP) or large loop electrosurgical excision of the transformation zone (LLETZ) for cervical lesions, or cold-steel surgery, laser excision and laser evaporation for vaginal lesions) or destruction (e.g., with carbon dioxide vaporization, cryotherapy, electrocauterization, or cold (thermo) coagulation)); photodynamic therapy; topical or systemic medications (e.g., podophyllin resin, podophyllotoxin, organic acids, such as salicylic acid, trichloroacetic acid and bich
• these agents boost antigen presentation (innate signals) while the present compositions boost antigen recognition by T cells.
• surgical treatments e.g., excision (e.g., loop electrosurgical excision procedure (LEEP) or large loop electrosurgical excision of the transformation zone (LLETZ) for cervical lesions, or cold-steel surgery, laser excision and laser evaporation for vaginal lesions) or destruction (e.g., with carbon dioxide vaporization, cryotherapy, electrocauterization, or cold (thermo) coagulation)); radiation therapy; photodynamic therapy; topical or systemic medications (e.g., podophyllin resin, podophyllotoxin, organic acids, such as salicylic acid, trichloroacetic acid and bichloroacetic acid, 5- fluorouracil, topical imiquimod, calcipotriene plus 5-fluorouracil, bleomycin, IFN-a, or cidofovir); or chemotherapy (e.
• excision e.g., loop electro
• mice were anesthetized with isoflurane followed by micro-aberrations over 0.2-0.3 cm 2 area on the base of the tongue using an 18-gauge sterile needle.
• Purified virus inoculum from MmuPVl - induced muzzle warts of B6.Cg-Foxnl nu/nu mice was applied onto the scarified tongue and spread homogeneously. The same viral inoculum was used for all infected mice.
• mice were subjected to DMBA/4NQO carcinogenesis protocol that we have optimized to induce carcinogenesis on the tongue of p53 +/ " mice on the C57BL/6 background (Figs. 1 A-C) 7 .
• Animals’ tongues were treated with a single dose of 50 pg 7,12-Dimethylbenz(a)anthracene (DMBA) in 50 pL of sesame oil applied to the tongue followed by 0.5% 4-nitroquinoline 1 -oxide (4NQO) treatment in drinking water for 20 weeks.
• DMBA 7,12-Dimethylbenz(a)anthracene
• 4NQO 4-nitroquinoline 1 -oxide
• FIGs. 2A-B show the effect of papillomavirus tongue colonization on reducing the size of malignant clones (marked as Trp53 mutant clones) in mice tongue, which indicates the protective impact of papillomavirus colonization against early malignant transformation in mucosal epithelia.
• Cancer Stat Facts Common Cancer Sites. 2. Agalliu I, Gapstur S, Chen Z, Wang T, Anderson RL, Teras L, Kreimer AR, Hayes RB, Freedman ND, Burk RD. Associations of Oral alpha-, beta-, and gamma-Human Papillomavirus Types With Risk of Incident Head and Neck Cancer. JAMA Oncol 2016: 2; 599-606

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