WO2011000889A1 - Topical vaccine formulations and methods of treating drug addiction using same - Google Patents

Topical vaccine formulations and methods of treating drug addiction using same Download PDF

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Publication number
WO2011000889A1
WO2011000889A1 PCT/EP2010/059318 EP2010059318W WO2011000889A1 WO 2011000889 A1 WO2011000889 A1 WO 2011000889A1 EP 2010059318 W EP2010059318 W EP 2010059318W WO 2011000889 A1 WO2011000889 A1 WO 2011000889A1
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Prior art keywords
hapten
carrier
topical
drug
topical vaccine
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PCT/EP2010/059318
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French (fr)
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William Henry
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William Henry
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Publication of WO2011000889A1 publication Critical patent/WO2011000889A1/en

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    • 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
    • A61K39/0013—Therapeutic immunisation against small organic molecules, e.g. cocaine, nicotine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A61K47/6415—Toxins or lectins, e.g. clostridial toxins or Pseudomonas exotoxins
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A61K47/646—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent the entire peptide or protein drug conjugate elicits an immune response, e.g. conjugate vaccines
    • 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
    • A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
    • A61K2039/541—Mucosal route
    • 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
    • A61K2039/55555—Liposomes; Vesicles, e.g. nanoparticles; Spheres, e.g. nanospheres; Polymers
    • 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
    • A61K2039/60—Medicinal preparations containing antigens or antibodies characteristics by the carrier linked to the antigen
    • A61K2039/6031—Proteins
    • A61K2039/6037—Bacterial toxins, e.g. diphteria toxoid [DT], tetanus toxoid [TT]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/70—Web, sheet or filament bases ; Films; Fibres of the matrix type containing drug
    • A61K9/7023—Transdermal patches and similar drug-containing composite devices, e.g. cataplasms
    • A61K9/703—Transdermal patches and similar drug-containing composite devices, e.g. cataplasms characterised by shape or structure; Details concerning release liner or backing; Refillable patches; User-activated patches
    • A61K9/7038—Transdermal patches of the drug-in-adhesive type, i.e. comprising drug in the skin-adhesive layer
    • A61K9/7046—Transdermal patches of the drug-in-adhesive type, i.e. comprising drug in the skin-adhesive layer the adhesive comprising macromolecular compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/70—Web, sheet or filament bases ; Films; Fibres of the matrix type containing drug
    • A61K9/7023—Transdermal patches and similar drug-containing composite devices, e.g. cataplasms
    • A61K9/703—Transdermal patches and similar drug-containing composite devices, e.g. cataplasms characterised by shape or structure; Details concerning release liner or backing; Refillable patches; User-activated patches
    • A61K9/7038—Transdermal patches of the drug-in-adhesive type, i.e. comprising drug in the skin-adhesive layer
    • A61K9/7046—Transdermal patches of the drug-in-adhesive type, i.e. comprising drug in the skin-adhesive layer the adhesive comprising macromolecular compounds
    • A61K9/7053—Transdermal patches of the drug-in-adhesive type, i.e. comprising drug in the skin-adhesive layer the adhesive comprising macromolecular compounds obtained by reactions only involving carbon to carbon unsaturated bonds, e.g. polyvinyl, polyisobutylene, polystyrene
    • A61K9/7061—Polyacrylates

Definitions

  • topical vaccine formulations comprising a hapten-carrier conjugate, a lipid, and a surfactant; methods of preparing same; and methods of using same for the treatment of drug abuse.
  • Hapten-carrier conjugates are currently in clinical trials as therapeutics for drug addiction.
  • a composition containing the hapten-carrier conjugate e.g.. a cocaine-bacterial toxin conjugate or a nictonic-bacterial toxin conjugate
  • antibodies specific to the drug of addiction arc elicited.
  • the therapeutic composition is a cocaine-carrier conjugate
  • treatment induces an anti-cocaine antibody response which reduces cocaine in the bloodstream or mucosal tissue of the subject, thereby reducing the physiologically addictive effects of the cocaine.
  • Treatment with a nicotine-carrier conjugate elicits an analogous effect towards nicotine, and reduces the individual's gratificiation from the use of nicotine.
  • topical vaccine formulations which may be used for the treatment of, for example, drug abuse.
  • the topical vaccine formulations provided herein comprise a hapten-carrier conjugate, a lipid, and a surfactant, optionally in a
  • the lipid is a phospholipid.
  • the surfactant is a non-ionic surfactant.
  • the hapten-carrier conjugate in the topical vaccine formulation is capable of eliciting an immune response in the form of anti-hapten antibodies within the subject which upon subsequent exposure to the drug neutralizes the hapten and diminishes or eliminates the pharmacological effects associated therewith.
  • the hapten is a drug or drug derivative or drug metabolite, preferably a drug of addiction, for example cocaine or nicotine.
  • the topical vaccine formulation can be used as a therapeutic for treating drug addiction, for example a cocaine or nicotine addiction, by vaccination of a subject with the topical vaccine formulation comprising a hapten-carrier conjugate, a lipid, and a surfactant.
  • the hapten-carrier conjugate is a cocaine-protein or nicotine-protein conjugate.
  • the hapten-carrier conjugate of the topical vaccine formulation is derived from at least one hapten and at least one T cell epitope-containing carrier, which conjugate is capable of stimulating the production of anti-hapten antibodies.
  • the carrier is a bacterial toxin, for example cholera toxin B.
  • the topical vaccine formulation comprises a therapeutically effective amount of hapten-carrier conjugate, for example from about 0.1 to about 50 mg of the hapten-carrier conjugate per g of the formulation.
  • the topical vaccine formulation comprises about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 8 mg, about 10 mg, about 12 mg, about 14 mg, about 16 mg, about 18 mg, or about 20 mg of the hapten-carrier conjugate per g of the formulation.
  • the topical vaccine formulation is a patch, for example, for transdermal delivery or transcutaneous delivery.
  • the hapten-carrier conjugate elicits an immune anti-hapten antibodies within the subject which upon subsequent exposure to the drug neutralizes the drug and diminishes or eliminates the pharmacological effects associated therewith.
  • the hapten-carrier conjugate is derived from a hapten that is a drug or drug derivative or drug metabolite, preferably a drug of addiction, for example cocaine or nicotine.
  • the topical vaccine formulation is N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoe
  • the topical vaccine formulation comprises from about 0.1 to about 50 mg of the hapten-carrier conjugate. In some embodiments of the method, the composition comprises about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 8 mg, about 10 mg, about 12 mg, about 14 mg, about 16 mg, about 18 mg, or about 20 mg of the hapten-carrier conjugate per g of the formulation.
  • the topical vaccine formulations provided herein are useful for the application, administration and/or transport of the hapten-carrier conjugate into and through barriers and constrictions, such as the skin of a subject (e.g., humans).
  • the topical vaccine formulations pro ⁇ ided herein form vesicles or other extended surface aggregates (ESAs), wherein the vesicular preparations have improved permeation capability through the semi-permeable barriers, such as skin.
  • ESAs extended surface aggregates
  • the topical vaccine formulation provided herein are able to form vesicles characterized by their deformability and/or adaptability.
  • the vesicles' deformability and/or adaptability allow the vesicles to penetrate the pores of the skin and deliver the hapten-carrier conjugate to the subject in an amount sufficient to elicit an immune response.
  • the adaptability or deformability of the vesicles may be determined by the ability of the vesicles to penetrate a barrier with pores having an average pore diameter at least 50% smaller than the average vesicle diameter before the penetration.
  • the topical vaccine formulations provided herein are used to deliver the hapten-carrier conjugate non-invasively.
  • the topical vaccine formulations provided herein may be administered in place of other vaccine formulations, including intravenous vaccines.
  • the term '"'subject refers to an animal, including, but not limited to, a primate (e.g. , human), cow, sheep, goat, pig, horse, dog, cat, rabbit, rat, or mouse.
  • a primate e.g. , human
  • the terms '"'subject ' "' and '"'patient'”' are used interchangeably herein in reference, for example, to a mammalian subject, such as a human subject.
  • the term '"'treat,”" “"treating,”” or '"'treatment of” means that the severity of a subject's condition is reduced or at least partially improved or ameliorated and/or that some alleviation, mitigation or decrease in at least one clinical symptom is achieved and/or there is an inhibition or delay in the progression of the condition and/or delay in the progression of the onset of disease or illness.
  • the term ""treat,”" '"'treating,'-" or “”treatment of” also means managing the disease state.
  • compositions provided herein denotes that a formulation does not result in an unacceptable level of irritation in the subject to whom the formulation is administered. Preferably such level will be sufficiently low to provide a formulation suitable for approval by regulatory authorities.
  • the term '""sufficient amount,”” “"amount effective to,”” or an “"amount sufficient to”” achieve a particular result refers to an amount of a hapten-carrier conjugate that is effective to produce a desired effect, which is optionally a therapeutic effect (i.e., by administration of a therapeutically effective amount).
  • a therapeutic effect i.e., by administration of a therapeutically effective amount.
  • “”therapeutically effective”” amount is an amount that provides some alleviation, mitigation, and/or decrease in at least one clinical symptom.
  • Clinical symptoms associated with the disorder that can be treated by the methods provided herein are well-known to those skilled in the art. Further, those skilled in the art will appreciate that the therapeutic effects need not be complete or curative, as long as some benefit is provided to the subject. For example, a ""sufficient amount'"' or “"an amount sufficient to"" can be an amount that is effective to treat a disease.
  • the term “"about”” means a range surrounding a particular numeral value which includes that which would be expected to result from normal experimental error in making a measurement.
  • the term “"about”” when used in connection with a particular numerical value means ⁇ 1%, ⁇ 2%, ⁇ 3%, ⁇ 4%, ⁇ 5%, ⁇ 10%, ⁇ 15%, or ⁇ 20% of the numerical value.
  • alkyl refers to a linear or branched saturated monovalent hydrocarbon radical, wherein the alkyl may optionally be substituted with one or more substituents Q as described herein.
  • alkyl also encompasses both linear and branched alkyl, unless otherwise specified.
  • the alkyl is a linear saturated monovalent hydrocarbon radical that has 1 to 20 (Ci-20), 1 to 15 (C 1-15), 1 to 12 (Cj- 12 ), 1 to 10 (Ci- 10 ), or 1 to 6 (Ci-6) carbon atoms, or a branched saturated monovalent hydrocarbon radical of 3 to 20 (C 3 . 20 ), 3 to 15 (Cs-is), 3 to 12 (C3.12), 3 Io 10 (C 3 -1 0 ), or 3 to 6 (Cj -6 ) carbon atoms.
  • linear C] -6 and branched C 3-6 alkyl groups are also referred as '"lower alkyl.”
  • alkyl groups include, but are not limited to, methyl, ethyl, propyl (including all isomeric forms), n-propyl, isopropyl, butyl (including all isomeric forms), n-butyl, isobutyl, sec-butyl, t-butyl, pentyl (including all isomeric forms), and hexyl (including all isomeric forms).
  • alkyl refers to a linear saturated
  • the term '"'aryP' refers to a monocyclic aromatic group and/or multicyclic monovalent aromatic group that contain at least one aromatic hydrocarbon ring.
  • the ary has from 6 to 20 (C6-20X from 6 to 15 ( €5-15), or from 6 to 10 (CVi 0) ring atoms.
  • aryl groups include, but are not limited to, phenyl, naphthyl, fluorenyl, azulenyl, anthryl, phenanthryl, pyrenyl, biphenyl, and terphenyl.
  • Aryl also refers to bicyclic or tricyclic carbon rings, where one of the rings is aromatic and the others of which may be saturated, partially unsaturated, or aromatic, for example, dihydronaphthyl, indenyl, indanyl. or tetrahydronaphthyl (tetralinyl).
  • aryl may also be optionally substituted with one or more substituents Q as described herein.
  • heteroaryl refers to a monocyclic aromatic group and/or multicyclic aromatic group that contain at least one aromatic ring, wherein at least one aromatic ring contains one or more heteroatoms independently selected from O, S, and N.
  • Each ring of a heteroaryl group can contain one or two O atoms, one or two S atoms, and/or one to four N atoms, provided that the total number of heteroatoms in each ring is four or less and each ring contains at least one carbon atom.
  • the heteroaryl may be attached to the main structure at any heteroatom or carbon atom which results in the creation of a stable compound. In certain embodiments, the heteroaryl has from 5 to 20, from 5 to 15, or from 5 to 10 ring atoms.
  • Examples of monocyclic heteroaryl groups include, but are not limited to, pyrrolyl, pyrazolyl, pyrazolinyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, thiadiazolyl, isothiazolyl, furanyl, thienyl, oxadiazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, and triazinyl.
  • bicyclic heteroaryl groups include, but are not limited to, indolyl, benzothiazolyl, benzoxazolyl, benzothienyl, quinolinyl, tetrahydroisoquinolinyl, isoquinolinyl,
  • benzimidazolyl benzopyranyl, indolizinyl, benzofuranyl, isobenzofuranyl, chromonyl, coumarinyl.
  • tricyclic heteroaryl groups include, but are not limited to, carbazoiyl, benzindolyl, phenanthrollinyl, acridinyl, phenanthridinyl, and xanthenyl.
  • heteroaryl may also be optionally substituted with one or more substituents Z as described herein.
  • alkenoyl refers to ⁇ C(O)-alkenyl.
  • ⁇ "alkenyl refers to a linear or branched monovalent hydrocarbon radical, which contains one or more, in one embodiment, one to five, carbon-carbon double bonds.
  • the alkenyl may be optionally substituted with one or more substituents Z as described herein.
  • alkenyl also embraces radicals having ""cis”” and “"/raw”" configurations, or alternatively, '""Z'"' and "'"E”-' configurations, as appreciated by those of ordinary skill in the art.
  • alkenyl' encompasses both linear and branched alkenyl, unless otherwise specified.
  • C 2-6 alkenyl refers to a linear unsaturated monovalent hydrocarbon radical of 2 to 6 carbon atoms or a branched unsaturated
  • the alkenyl is a linear monovalent hydrocarbon radical of 2 to 30 (C 2-3 o), 2 to 24 (C 2-2 O, 2 to 20 ( €2-20), 2 to 15 (C2- 15 ), 2 to 12 (C 2- I 2 ), 2 to 10 (C 2 -] 0), or 2 to 6 (C 2-6 ) carbon atoms, or a branched monovalent hydrocarbon radical of 3 to 30 (C 3-3O ), 3 to 24 (C 3-24 ), 3 to 20 (C 3-2 o), 3 to 15 (C 3 .
  • alkenyl groups include, but are not limited to, ethenyl, propen-1-yl, propcn-2-yl, allyl, butenyl, and 4- methylbutenyl.
  • the alkenoyl is mono-alkenoyl, which contains one carbon-carbon double bond.
  • the alkenoyl is di-alkenoyl, which contains two carbon-carbon double bonds.
  • the alkenoyl is poly- alkenoyl, which contains more than two carbon-carbon double bonds.
  • heterocyclyl refers to a monocyclic non- aromatic ring system and/or multicyclic ring system that contains at least one non-aromatic ring, wherein one or more of the non-aromatic ring atoms are heteroatoms independently selected from O, S, or N; and the remaining ring atoms are carbon atoms.
  • the heterocyclyl or heterocyclic group has from 3 to 20, from 3 to 15, from 3 to 10, from 3 to 8, from 4 to 7, or from 5 to 6 ring atoms.
  • the heterocyclyl is a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which may include a fused or bridged ring system, and in which the nitrogen or sulfur atoms may be optionally oxidized, the nitrogen atoms may be optionally quaternized, and some rings may be partially or fully saturated, or aromatic.
  • the heterocyclyl may be attached to the main structure at any heteroatom or carbon atom which results in the creation of a stable compound.
  • heterocyclic radicals include, but are not limited to, acridinyl, azepinyl, benzimidazolyl, benzindolyl, benzoisoxazolyl, benzisoxazinyl, benzodioxanyl, benzodioxolyl, benzofuranonyl, benzofuranyl, benzonaphthofuranyl, benzopyranonyl, benzopyranyl, benzotetrahydrofuranyl, benzotetrahydrothienyl, benzothiadiazolyl.
  • benzothiazolyl benzothiophenyl, benzotriazolyl, benzothiopyranyl, benzoxazinyl, benzoxazolyl, benzothiazolyl, ⁇ -carbolinyl, carbazolyl, chromanyl, chromonyl, cinnolinyl, coumarinyl, decahydroisoquinolinyl, dibenzofuranyl, dihydrobenzisothiazinyl,
  • phenanthridinyl phenathrolinyl, phenarsazinyl, phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, piperazinyl, piperidinyl, 4-piperidonyl, pteridinyl, purinyl, pyra/inyl, pyrazolidinyl, pyrazolyl, pyridazinyl, pyridinyl, pyridopyridinyl, pyrimidinyl, pyrrolidinyl, pyrrolinyl, pyrrolyl, quinazolinyl, quinolinyl, quinoxalinyl, quinuclidinyl, tctrahydrofuryl, tetrahydrofuranyl, tetrahydroisoquinolinyl, tetrahydropyranyl, tetrahydrothienyl, tetra
  • halogen refers to fluorine, chlorine, bromine, and/or iodine.
  • Optionally substituted is intended to mean that a group, including alkyl, alkenyl, alkynyl, cycloalkyl, aryl, aralkyl, heteroaryl, and heterocyclyl, may be substituted with one or more substituents Z, in one embodiment, one, two, three or four substituents Z, where each Z is independently selected from the group consisting of cyano, halo, oxo, nitro, C 1-6 alkyl, halo- C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 cycloalkyl, C 6- ⁇ aryl, C 7 - I4 aralkyl, heteroaryl, heterocyclyl, -C(O)R 6 , -C(O)OR 6 , -C(O)NR f R g , - C(NR e )NR f R g , -OR C ,
  • optically active and “"enantiomerically active” refer to a collection of molecules, which has an enantiomeric excess of no less than about 50%, no less than about 70%, no less than about 80%, no less than about 90%, no less than about 91%, no less than about 92%, no less than about 93%, no less than about 94%, no less than about 95%, no less than about 96%, no less than about 97%, no less than about 98%, no less than about 99%, no less than about 99.5%, or no less than about 99.8%.
  • the prefixes R and S are used to denote the absolute configuration of the molecule about its chiral centcr(s).
  • the (+) and (-) are used to denote the optical rotation of the compound, that is, the direction in which a plane of polarized light is rotated by the optically active compound.
  • the (-) prefix indicates that the compound is levorotatory, that is, the compound rotates the plane of polarized light to the left or counterclockwise.
  • the (+) prefix indicates that the compound is dextrorotatory, that is, the compound rotates the plane of polarized light to the right or clockwise.
  • the sign of optical rotation, (+) and (-) is not related to the absolute configuration of the molecule, R and S.
  • solvate refers to a compound provided herein or a salt thereof, which further includes a stoichiometric or non-stoichiometric amount of solvent bound by non-covalent intermolecular forces. Where the solvent is water, the solvate is a hydrate.
  • the formulations provided herein comprise a hapten-carrier conjugate, a lipid, preferably a phospholipid, a surfactant, preferably a nonionic surfactant, and an aqueous solution, having a pH ranging from 3.5 to 9.0, preferably from 4 to 7.5.
  • the formulations may optionally contain buffers, antioxidants, preservatives, microbicides. antimicrobials, emollients, co-solvents, and/or thickeners.
  • the formulations provided herein form vesicles or other extended surface aggregates (ESAs), wherein the vesicular preparations have improved permeation capability through the semi-permeable barriers, such as skin and/or nails.
  • ESAs extended surface aggregates
  • the vesicles or extended surface aggregates provided herein comprise of a hapten carrier conjugate, a lipid, and one or more membrane destabilizing agents, such as surfactants.
  • the hapten-carrier conjugates for use in the topical vaccine formulations of the disclosure can be any hapten-carrier conjugate that elicits an appropriate immune response to the free hapten.
  • the hapten-carrier conjugates are those described in U.S. Patent Application Publication No. 2005/0124061 (published June 9, 2005), herein incorporated by reference in its entirety.
  • the carrier is a bacterial toxin is from a gram-negative bacteria. In some embodiments, the carrier is a bacterial toxin is from a gram-positive bacteria. In some embodiments, the gram-negative bacteria is Escherichia coli. In some embodiments, the bacteria is the gram-positive bacillus, staphylococcus, streptococcus, streptomyces, or mollicutes (mycoplasma). In some embodiments, the carrier is a bacterial exotoxin. In some embodiments, the carrier is a bacterial toxin that has been modified to reduce its toxicity. Bacterial toxins for use in accordance with the disclosure include, but are not limited to, cholera toxin, preferably CTB (including recombinant CTB (rCTB)),
  • Escherichia coli toxins such as heat-labile enterotoxin (LT), heat-stable exotoxin (ST), cytotoxic necrotizing factor (CNF), cytolethal distending toxin (CLDT), or enteroaggregative E.
  • LT heat-labile enterotoxin
  • ST heat-stable exotoxin
  • CNF cytotoxic necrotizing factor
  • CLDT cytolethal distending toxin
  • enteroaggregative E enteroaggregative E.
  • EAST heat-stable toxin
  • Dtx diphtheria toxin
  • tetanus toxin shiga toxins
  • botulinum toxin staphylococci toxins, such as Staphylococcus aureus alpha toxin, Exfoliatin B or leukocidin, staphylococcal toxic shock syndrome toxin (TSST-I), staphylococcus enterotoxins or exfoliative toxins
  • streptococci toxins such as pneumolysin of Streptococcus pneumoniae, streptolysin O, Erythrogenic toxin (streptococcal pyrogenic exotoxin (SPE)), and other pyrogenic toxins of Streptococcus pyogenes
  • clostridial toxins such as toxin A/toxin B of Clostridium difficile, Iota family, C2 family (toxins C and D), or C3 toxins, or neurotoxins A-G of Clos
  • a bacterial ADP-ribosylating exotoxin is preferably used, such as, for example, cholera toxin, diphtheria toxin, pertussis toxin, Pseudomonas exotoxin A, or E. coli LT.
  • the catalytic subunit (usually, the '"-A"" subunit) of the bacterial ADP- ribosylating exotoxin is used as carrier.
  • the receptor-binding subunit of the bacterial ADP-ribosylating exotoxin (usually, the ""B"" subunit) is preferred.
  • both subunits or a fragment or fragments thereof are used as a carrier.
  • a bacterial pore-forming toxin is used, such as perfringiolysin O, hemolysin, listeriolysin, anthrax EF, alpha toxin, pneumolysin, streptolysin O, or leukocidin.
  • the carrier is a pyrog ⁇ nic exotoxin, or a modified form thereof, such as staphylococcal enterotoxins serotypes A-E, G, and H, group A streptococcal pyrogenic exotoxins A-C, staphylococcal exfoliatin toxin, and staphylococcal TSST-I .
  • Particularly useful bacterial toxin carriers include any bacterial toxin with the ability to enhance a mucosal immune response, for example, CTB or any toxin in the E. coli heat-labile enteroloxin (LTB) family of bacterial toxins.
  • the bacterial toxin carrier of the instant invention contains at least one T cell epitope which is capable of stimulating the T cells of the subject, which in turn help the B cells initiate and maintain sustained antibody production to portions of the entire conjugate, including the hapten portion.
  • T cell epitope which is capable of stimulating the T cells of the subject, which in turn help the B cells initiate and maintain sustained antibody production to portions of the entire conjugate, including the hapten portion.
  • the carrier since a carrier is selected because it is immunogenic, a strong immune response to the vaccine in a diverse patient population is expected.
  • the carrier like the hapten, must be sufficiently foreign to elicit a strong immune response to the vaccine.
  • a conservative, but not essential, approach is to use a carrier to which most patients have not been exposed to avoid the phenomenon of carrier-induced epitope suppression.
  • Vaccines which utilize carrier proteins to which patients are already immune are commercially available.
  • carriers containing a large number of lysines are particularly suitable for conjugation according to the methods of the instant invention. In certain embodiments, therefore, the bacterial toxin carriers of the invention are modified so that their immunogenic properties are enhanced.
  • Cholera toxin is the enterotoxin produced by Vibrio cholerae and consists of five identical B subunits with each subunit having a molecular weight of 1 1.6 KDa (103 amino acids) and one A subunit of 27.2 KDa (230 amino acids) (Finkelstein (1988) Immunochem. MoI. Gen. Anal. Bac. Path. 85-102).
  • the binding subunit, CTB binds to ganglioside GMl on the cell surface (Sixma et al. (1991) Nature 351 :371-375; Orlandi et al. (1993) J. Biol. Chem. 268: 17038-17044).
  • CTA is the enzymatic subunit which enters the cell and catalyzes ADP- ribosylation of a G protein, constitutively activating adenylate cyclase (Finkelstein (1988) Immunochem, MoI. Gen. Anal. Bac. Path. pp. 85-102). In the absence of the A subunit, cholera toxin is not toxic.
  • CTB is the bacterial toxin carrier.
  • CTB is a highly immunogenic protein subunit capable of stimulating strong systemic and mucosal antibody responses (Lycke (1992) J. Immunol. 150:4810-4821 ; Holmgren et al. (1994) Am. J. Trop. Med. Ilyg. 50:42-54; Silbart et al. (1988) J. Immun. Meth. 109:103-1 12; Katz et al. (1993) Infection Immun. 61 : 1964-1971).
  • IgA and IgG anti-hapten response is highly desirable in blocking, for example, cocaine or other substances for which is immunity is desired that are administered nasally or by inhalation, and in blocking nicotine or other substances that are absorbed in the mouth and lungs.
  • CTB has already been shown to be safe for human use in clinical trials for cholera vaccines (Holmgren et al., supra; Jertborn et al. (1994) Vaccine 32: 1078-1082; '"'The Jordan Report, Accelerated Development of Vaccines"" 1993., NIAID, 1993). It is a discovery of this invention that hapten-carrier conjugates comprising CTB have even greater immunogenicity when CTB contains a signal peptide..
  • CTBs generated according to these methods, and the methods presented herein, may be modified so that it contains a signal peptide as described herein.
  • CTB is produced in Vibrio cholerae strain 213. See, for example, International Patent Application Publication No. WO 2005/042749, published May 12. 2005, which is incorporated by reference herein in its entirety.
  • CTB is produced in E, coli (see, e.g., U.S. Patent
  • Amounts of recombinant CTB have been expressed and purified amounts which, once optimized, are produced in large fermentation batches.
  • Processes for expressing and purifying recombinant protein are known in the art, for example, U.S. Patent Application Ser. No. 07/807,529.
  • CTB may be purified by affinity chromatography (Tayot et al. (1981) Eur. J. Biochem. 1 13:249-258), conjugated to cocaine or nicotine derivatives, and the conjugate may then be further purified.
  • the purified CTB and the resulting conjugate are analyzed for purity and for maintenance of the pentameric structure of CTB.
  • Techniques include SDS-PAGE, native PAGE, gel filtration chromatography, Western blotting, direct and GMl -capture ELISA, and competition ELISA with biotinylated CTB.
  • Level of haptenation is measured by mass spectrometry, reverse phase HPLC and by analysis of the increase in UV absorbance resulting from the presence of the hapten. Both the solubility and the stability of the conjugate are optimized in preparation for full-scale formulation. Details of some of these analyses are given in the Examples.
  • the pentameric structure of CTB is a preferred carrier for practice of the present invention, and GMl binding is an effective assay to determine that the pentameric form of CTB is present, the present invention is not limited to the use of the pentameric form of CTB.
  • Other forms of CTB are contemplated (e.g., monomer, dimer, etc.) that may be manipulated for use in the invention. If a carrier other than the pentameric form of CTB is utilized, then one skilled in the art would use an appropriate assay to determine the presence and activity of the required carrier, e.g., the use of GMl binding to determine the presence of the pentameric form of CTB).
  • CTB Another useful CTB for use as a carrier is cholera toxin which provides improved mucosal response over CTB. It has been reported that the enzymatieally active A subunit adjuvant enhances activity (Liang et al. (1988) J. Immunol. 141 :1495-1501; Wilson et al. (1993) Vaccine 1 1 : 113-118; Snider et al. (1994) 1 Immunol. 153:647).
  • a ""hapten" for the purposes of this disclosure is a low-molecular- weight organic compound that reacts specifically with an antibody and which is incapable of inciting an immune response by itself but is immunogenic when complexed to a T cell epitope- containing carrier (i.e., as a hapten-carrier conjugate).
  • the hapten is poorly immunogenic by itself.
  • the hapten is characterized as the specificity- determining portion of the hapten-carrier conjugate, that is, it is capable of reacting with an antibody specific to the hapten in its free state. In some embodiments, in a non-immunized subject, there is an absence of formation of antibodies to the hapten.
  • the hapten in a non-immunized subject, there may be a low level of antibodies to the hapten, or a level of antibodies to the hapten for which an increase in the immune response to the hapten is desired.
  • the hapten is a drug, an analog of a portion of the drug, or a drug derivative.
  • the hapten is an addictive drug, analog of a portion of an addictive drug, or addictive drug derivative.
  • the addictive drug is nicotine or cocaine.
  • the immunogenic composition when initially administered will give rise to a ""desired measurable outcome.
  • the desired measurable outcome is the production of a high titer of anti-hapten antibodies (approximately 0.1 mg/ml to 1 mg/ml or greater of specific antibody in the serum).
  • manipulation of the dosage regimen suitable for the individual gives and maintains a sustained desired therapeutic effect.
  • the ""desired therapeutic effect” is the neutralization of a sufficient fraction of free hapten to reduce or eliminate the pharmacological effects of the hapten (e.g., nicotine or cocaine) within a therapeutically acceptable time frame by anti- hapten antibodies specific for the hapten upon a subsequent exposure to the hapten.
  • the hapten e.g., nicotine or cocaine
  • Determining the therapeutically acceptable time frames for how long it takes to get a sufficient antibody response to a given hapten and how-long is that antibody response is maintained thereto are achieved by those skilled in the art by assessing the characteristics of the subject to be immunized, hapten (e.g., drug of abuse) to be neutralized, as well as the mode of administration. Using this and other immunization protocols as a model, one skilled in that art would expect the immunity or the period of protection to last several months, up to more than one year.
  • hapten e.g., drug of abuse
  • the hapten- carrier conjugate can be prepared by modifying the hapten sufficiently to render it capable of being conjugated or joined to a carrier while maintaining enough of the structure so that it is recognized as free state hapten (for example, as free cocaine or nicotine). It is preferable that a vaccinated individual has antibodies which recognize free hapten (e g., cocaine or nicotine). Radioimmunoassay and competition ELISA assay experiments can measure antibody titers to free hapten. Antibodies of interest are hapten- specific antibodies and, for example, cocaine-specific antibodies or nicotine- specific antibodies. It will be understood to one of ordinary skill in the art that the principles and methods used described herein may be extended from this disclosure to a wide range of hapten-carrier conjugates useful in the treatment of a variety of diseases, conditions, or drug addictions and toxic responses.
  • the hapten is a drug, such as, for example, hallucinogens, for example mescaline and LSD; Cannabinoids, for example THC; Stimulants, for example amphetamines, cocaine, phenmetrazine, methylphenidate; Nicotine; Depressants, for example, nonbarbiturates (e.g.
  • hapten-carrier conjugates for use in the topical vaccine formulations can be found in U.S. Patent Application Publication No. 2005/0124061
  • Haptens are generally conjugated to a carrier by means of a chemical linker, the length and nature of which is such that the hapten is displaced a sufficient distance from the carrier domain to allow its optimal recognition by the antibodies initially raised against it.
  • the length of the linker can be varied by, for example, modifying the number Of-CH 2 groups in a given linker. The following is a non-exhaustive list of linkers usable in the hapten- carrier conjugates described herein:
  • n is an integer from 1 to about 20, preferable from about 3 to about 6; Y is selected S, O, or Nil; and Q is the carrier.
  • the chemical and/or conceptual precursor to the hapten carrier conjugate will be an analog of the hapten having one of the above-mentioned linkers connected thereto, wherein Q is H, OH, CH 3 , OCH 3 , COOH, a halogen, an activated ester or esters (such as 2-nitro-4-sulfophenyl ester or N- oxysuccinimidyl ester), a group or groups reactive toward the carrier (such as a mixed anhydride, acyl halide, acyl azide, alkyl halide, N-maleimide, imino ester, isocyanate, and isothiocyanate), or another linker identified by its ""CJ'"' reference number.
  • Q is H, OH, CH 3 , OCH 3 , COOH, a halogen, an activated ester or esters (such as 2-nitro-4-sulfophenyl ester or N- oxysuccinimidyl ester),
  • a T cell epitope containing carrier may be modified by methods known to those skilled in the art to facilitate conjugation to the hapten (e.g., by thiolation), for example with 2-iminothiolane (Traut's reagent) or by succinylation, etc.
  • “-'BSA”” is bovine serum albumin;
  • ""DC” is dicyclohexylcarbodiimide;
  • ""DMF” is N 9 N- dimethylformamide;
  • ""EDTA” is ethylenediamine tetraacetic acid, disodium salt;
  • ""HATU” is O-(7-azabenzotriazol-1-yl)-l ,l ,3,3-tetramethyluronium hexafluorophosphate;
  • ""NMM” is N-methylmorpholine;
  • ""HBTU” is 2-(1H-benzotriazole-1-yl)-l, 1,3,3- tetramethyluronium hexafluorophosphate;
  • ""TNTU” is 2-(5
  • the hapten-carrier conjugate is a conjugate of nicotine or nicotine metabolites.
  • chemical precursors to a nicotine carrier conjugate can be synthesized by selectively alkylating the pyridine nitrogen of (S)-(-)- nicotine in anhydrous methanol, with ethyl 3-bromobutyrate, 5-bromovaleric acid, 6- bromohexanoic acid or 8-bromooctanoic acid. The resultant chemical precursor can then be conjugated to a carrier protein usin, for example, HATU.
  • succinylated nornicotine is the chemical precursor to the conjugate.
  • a ""lipid” is any substance, which has properties like or similar to those of a fat. As a rule, it has an extended apolar group (the “"chain”", X) and generally also a water-soluble, polar hydrophilic part, the ""head”” group (Y) and has the basic Formula II:
  • n is equal to or larger than zero.
  • all amphfphilic substances including, but not limited to glycerides, glycerophospholipids, glycerophosphinolipids, glycerophosphonolipids, sulfolipids, sphingolipids, isoprenoid lipids, steroids or sterols and carbohydrate-containing lipids can generally be referred to as lipids, and are included as such in this disclosure.
  • a list of relevant lipids and lipid related definitions is provided in EP 0 475 160 Al (see, e.g. p. 4, 1. 8 to p. 6. 1. 3) and U.S. Patent No. 6,165,500 (see. e.g., col. 6, 1. 10 to col. 7. 1. 58), each incorporated herein by reference in their entirety.
  • a phospholipid is, for example, a compound of Formula II:
  • R 1 CH 2 - CHR 2 - CR 3 H- O— PHO 2 -O-R 4 (II) wherein R 1 and R 2 cannot both be hydrogen, OH or a Cj-C 3 alkyl group, and typically are independently, an aliphatic chain, most often derived from a fatty acid or a fatty alcohol; R J generally is a hydrogen.
  • the OH-group of the phosphate is a hydroxyl radical or hydroxyl anion (i.e., hydroxide) form, dependent on degree of the group ionization.
  • R 4 may be a proton or a short-chain alkyl group, substituted by a tri-short-chain alkylammonium group, such as a trimethylammonium group, or an amino-substituted short-chain alkyl group, such as 2-trimethylammonium ethyl group (cholinyl) or 2-dimethylammonium short alkyl group.
  • a sphingophospholipid is, for example, a compound of Formula IIB:
  • R 1 is a fatty-acid attached via an amide bond to the nitrogen of the sphingosine and R 4 has the meanings given under Formula II.
  • a lipid preferably is a substance of formulae II or IIB, wherein R 1 and/or R 2 are acyl or alkyl, n-hydroxyacyl or n-hydroxyalkyl, but may also be branched, with one or more methyl groups attached at almost any point of the chain; usually, the methyl group is near the end of the chain (iso or anteiso).
  • the radicals R 1 and R 2 may moreover either be saturated or unsaturated (mono-, di- or poly-unsaturated).
  • R 3 is hydrogen and R 4 is 2-trimethylammonium ethyl (the latter corresponds to the phosphatidyl choline head group), 2-dimethylammonium ethyl, 2 -methyl ammonium ethyl or 2-aminoethyl (corresponding to the phosphatidyl ethanolamine head group).
  • R 4 may also be a proton (giving phosphatidic acid), a serine (giving phosphatidylserine), a glycerol (giving phosphatidylglycerol), an inositol (giving phosphatidylinositol), or an alkylamine group (giving phosphatidylethanolamine in case of an ethylamine), if one chooses to use a naturally occurring glycerophosphoUpid. Otherwise, any other sufficiently polar phosphate ester, such that will form a lipid bilayer, may be considered as well for making the formulations of the disclosure.
  • Table 1 lists preferred phospholipids in accordance with the disclosure.
  • the preferred lipids in context of this disclosure are uncharged and form stable, well hydrated bilayers; phosphatidylcholines, phosphatidylethanolamine, and sphingomyelins are the most prominent representatives of such lipids. Any of those can have chains as listed in the Table 1 , the ones forming fluid phase bilayers, in which lipid chains are in disordered state, being preferred.
  • Different negatively charged, i.e., anionic, lipids can also be incorporated into vesicular lipid bilayers to modify the (cationic) drug loading into or release from the resulting lipid aggregates.
  • Attractive examples of such charged lipids are phosphatidylglycerols, phosphatidylinositols and, somewhat less preferred, phosphatide acid (and its alkyl ester) or phosphatidylserine. It will be realized by anyone skilled in the art that it is less commendable to make vesicles just from the charged lipids than to use them in a combination with electro- neutral bilayer component(s).
  • buffer composition and/or pH care must be selected so as to ensure the desired degree of lipid head-group ionization and/or the desired degree of electrostatic interaction between the, oppositely, charged drug and lipid molecules.
  • the charged bilayer lipid components can in principle have any of the chains listed in the Table 1.
  • the chains forming fluid phase lipid bilayers are clearly preferred, however, both due to vesicle adaptability increasing role of increasing fatty chain fluidity and due to better ability of lipids in fluid phase to mix with each other, and with drugs.
  • the fatty acid- or fatty alcohol-derived chain of a lipid is typically selected amongst the basic aliphatic chain types given in the following tables:
  • Suitable fatty residues can furthermore be branched, for example, can contain a methyl group in an iso or anteiso position of the fatty acid chain, or else closer to the chain middle, as in 10- ⁇ -methyloctadecanoic acid or tuberculostearic chain.
  • Relatively important amongst branched fatty acids are also isoprenoids, many of which are derived from
  • 3,7,11 ,15-tetramethylhexadec-trans-2-en-1-ol the aliphatic alcohol moiety of chlorophyll.
  • Examples include 5.9,13,17-tetramethyloctadecanoic acid and especially 3,7,1 1,15- tetramethylhexadecanoic (phytanic) and 2,6,10,14-tetramethylpentadecanoic (pristanic) acids.
  • a good source of 4,8,12-trimethyltridecanoic acid are marine organisms. Combination of double bonds and side chains on a fatty residue are also possible.
  • suitable fatty residues may carry one or a few oxy- or cyclic groups, especially in the middle or towards the end of a chain.
  • alicyclic fatty acids are those comprising a cyclopropane (and sometimes cyclopropene) ring, but cyclohexyl and cycloheptyl rings can also be found and might be useful for purposes of this disclosure.
  • 2-(D)-Hydroxy fatty acids are more ubiquitous than alicyclic fatty acids, and are also important constituents of sphingolipids.
  • unsaturation, branching or any other kind of derivatization of a fatty acid is best compatible with the intention of present disclosure of the site of such modification is in the middle or terminal part of a fatty acid chain.
  • the cis- unsaturated fatty acids are also more preferable than trans-unsata ⁇ ated fatty acids and the fatty radicals with fewer double bonds are preferred over those with multiple double bonds, due to oxidation sensitivity of the latter.
  • symmetric chain lipids are generally better suited than asymmetric chain lipids.
  • a preferred lipid of the Formula II is, for example, a natural phosphatidylcholine, which used to be called lecithin. It can be obtained from egg (rich in palmitic, Cj 6 o, and oleic, Cis i, but also comprising stearic. C] 8 o > palmitoleic, C) 6 i, linolenic, C] 8 2, and arachidonic, C20 4 . radicals), soybean (rich in unsaturated Ci 8 chains, but also containing some palmitic radical, amongst a few others), coconut (rich in saturated chains), olives (rich in
  • saffron safflower
  • sunflowers rich in n-6 linoleic acid
  • linseed rich in n-3 linolenic acid
  • whale fat rich in monounsaturated n-3 chains
  • primrose or stimula rich in n-3 chains.
  • natural phosphatidyl ethanolamines used to be called cephalins
  • sphingomyelins of biological origin are typically prepared from eggs or brain tissue.
  • phosphatidylserines also typically originate from brain material whereas phosphatidylglycerol is preferentially extracted from bacteria, such as E. CoIi, or else prepared by way of transphosphatidylation, using phospholipase D, starting with a natural phosphatidylcholine.
  • bacteria such as E. CoIi
  • phospholipase D phospholipase D
  • the preferred phosphatidic acid is either extracted from any of the mentioned sources or prepared using phospholipase D from a suitable phosphatidylcholine.
  • R 4 in Formula II corresponds to 2- trimethylammonium ethyl
  • R 1 and R 2 are aliphatic chains, as defined in the preceding paragraph with 12 to 30 carbon atoms, preferentially with 14 to 22 carbon atoms, and even more preferred with 16 to 20 carbon atoms, under the proviso that the chains must be chosen so as to ensure that the resulting ESAs comprise fluid lipid bilayers. This typically means use of relatively short saturated and of relatively longer unsaturated chains.
  • R 4 in Formula HB corresponds to 2-trimethylammonium ethyl
  • R 1 is an aliphatic chain, as defined in the preceding paragraph, with 10 to 20 carbon atoms, preferentially with 10 to 14 carbon atoms per fully saturated chain and with 16-20 carbon atoms per unsaturated chain.
  • Synthetic phosphatidyl ethanolamines (R 4 is 2-aminoefhyl), synthetic phosphatidic acids (R 1 is a proton) or its ester (R 4 corresponds, for example, to a short-chain alkyl, such as methyl or ethyl), synthetic phosphatidyl serines (R 4 is L- or D-serine), or synthetic
  • phosphatidyl (poly)alcohols such as phosphatidyl inositol, phosphatidyl glycerol (R 4 is L- or D-glycerol) are preferred as lipids, wherein R 1 and R 2 are fatty residues of identical or moderately different type and length, especially such as given in the corresponding tables given before in the text.
  • R 1 can represent alkenyl and R 2 identical hydroxyalkyl groups, such as tetradecylhydroxy or hexadecylhydroxy. for example, in ditetradecyl or dihexadecylphosphatidyl choline or efhanolamine, R 1 can represent alkenyl and R 2
  • hydroxyacyl such as a plasmalogen (R 4 trimethylammonium ethyl), or R 1 can be acyl, such as lauryl, myristoyl or palmitoyl and R 2 can represent hydroxy as, for example, in natural or synthetic lysophosphatidyl cholines or lysophosphatidyl glycerols or lysophosphatidyl ethanolamines, such as 1 -myristoyl or 1-palmitoyllysophosphatidyl choline or -phosphatidyl ethanolamine; frequently, R J represents hydrogen.
  • a lipid of Formula HB is also a suitable lipid within the sense of this disclosure.
  • n l
  • R 1 is an alkenyl group
  • R 2 is an acylamido group
  • R 3 is hydrogen
  • R 4 represents 2-trimethylammonium ethyl (choline group).
  • a lipid is known under the name of sphingomyelin.
  • Suitable lipids furthermore are a lysophosphatidyl choline analog, such as 1 - lauroyl-1,3-dihydroxypropane-3-phosphoryl choline, a monoglyceride, such as monoolein or monomyristin, a cerebroside, ceramide polyhexoside, sulfatide, sphingoplasmalogcn, a ganglioside or a glyceride, which does not contain a free or esterified phosphoryl or phosphono or phosphino group in the 3 position.
  • a lysophosphatidyl choline analog such as 1 - lauroyl-1,3-dihydroxypropane-3-phosphoryl choline
  • a monoglyceride such as monoolein or monomyristin
  • a cerebroside such as monoolein or monomyristin
  • ceramide polyhexoside such as a cerebroside, ceramide polyhex
  • a glyceride is diacyl glyceride or 1 -alkenyl- l-hydroxy-2-acyl glyceride with any acyl or alkenyl groups, wherein the 3-hydroxy group is etherified by one of the carbohydrate groups named, for example, by a galactosyl group such as a monogalactosyl glycerin.
  • Lipids with desirable head or chain group properties can also be formed by biochemical means, for example, by means of phospholipases (such as phospholilpase Al, A2, B, C and, in particular, D), desaturases, elongases, acyl transferases, etc., from natural or synthetic precursors.
  • phospholipases such as phospholilpase Al, A2, B, C and, in particular, D
  • desaturases elongases
  • acyl transferases etc.
  • a suitable lipid is any lipid, which is contained in biological membranes and can be extracted with the help of apolar organic solvents, such as chloroform.
  • lipids also include, for example, steroids, such as estradiol, or sterols, such as cholesterol, beta-sitosterol, desmosterol, 7-keto-cholesterol or beta-cholestanol, fat-soluble vitamins, such as retinoids, vitamins, such as vitamin Al or A2, vitamin E, vitamin K, such as vitamin Kl or K2 or vitamin Dl or D3, etc.
  • the less soluble amphiphilic components comprise or preferably comprise a synthetic lipid, such as myristoleoyl, palmitolcoyl, petroselinyl, petroselaidyl, oleoyl, elaidyl, cis- or trans-vaccenoyl, linolyl, linolenyl, linolaidyl, octadecatetraenoyl, gondoyl,
  • a synthetic lipid such as myristoleoyl, palmitolcoyl, petroselinyl, petroselaidyl, oleoyl, elaidyl, cis- or trans-vaccenoyl, linolyl, linolenyl, linolaidyl, octadecatetraenoyl, gondoyl,
  • docosadienoyl docosalrienoyl, docosatetraenoyl, lauroyl, tridecanoyl, myristoyl,
  • the more soluble amphiphilic components(s) is/are frequently derived from the less soluble components listed above and, to increase the solubility, substituted and/or complexed and/or associated with a butanoyl, pentanoyl, hexanoyl, heptanoyl, octanoyl, nonanoyl, decanoyl or undecanoyl substituent or several, mutually independent, selected substituents or with a different material for improving the solubility.
  • a further suitable lipid is a diacyl- or dialkyl-glycerophosphoetha- nolamine azo polyethoxylene derivative, a didecanoylphosphatidyl choline or a
  • the amount of lipid in the formulation is from about 1% to about 10%, about 1% to about 4%, about 4% to about 7% or about 7% to about 10% by weight.
  • the lipid is a phospholipid.
  • the phospholipid is a phosphatidylcholine.
  • surfactant' has its usual meaning.
  • a list of relevant surfactants and surfactant related definitions is provided in EP 0 475 160 Al (see, e.g., p. 6, 1. 5 to p.14. 1.17) and U.S. Pat. No, 6,165,500 (see, e.g., col. 7, 1. 60 to col. 19, 1. 64), each herein incorporated by reference in their entirety, and in appropriate surfactant or pharmaceutical Handbooks, such as Handbook of Industrial Surfactants or US Pharmacopoeia, Pharm. Eu.
  • the surfactants are those described in Tables 1-18 of U.S. Patent Application Publication No.
  • the list includes ionized long-chain fatty acids or long chain fatty alcohols, long chain fatty ammonium salts, such as alkyl- or alkenoyl-trimethyl-, -dimethyl- and -methyl- ammonium salts, alkyl- or alkenoyl-sulphate salts, long fatty chain dimethyl-aminoxides, such as alkyl- or alkenoyl-dimethyl-aminoxides, long fatty chain, for example alkanoyl, dimethyl-aminoxides and especially dodecyl dimethyl-aminoxide, long fatty chain, for example alkyl-N-methylglucamide- s and alkanoyl-N-methylglucamides, such as MEGA-8, MEGA-9 and MEGA- 10, N-long fatty chain-N,N-dimethyl glycines, for example N-alkyl- N, N-dimethyl glycines, 3-(long fatty chain-dimethylam
  • polyethylen-glycol- acylphenyl ethers especially nonaethylen-glycol-octyl- phenyl ether, polyethylene-long fatty chain-ethers, especially polyethylene-acyl ethers, such as nonaethylen-decyl ether, nonaethylen-dodecyl ether or octaethylene-dodecyl ether, polyethyleneglycol-isoacyl ethers, such as octaethyleneglycol-isotridecyl ether, polyethyleneglycol-sorbitane-long fatty chain esters, for example polyethyleneglycol-sorbitane-acyl esters and especially polyoxyethylene- monolaurate (e.g.
  • polysorbate 20 or Tween 20 polyoxyethylene-sorbitan-monooleate (e.g. polysorbate 80 or Tween 80), polyoxyethylene-sorbitan-monolauroleylate, polyoxyethylene - sorbitan-monopetroselinate, polyoxyethylene -sorbitan— monoelaidate, polyoxyethylene - sorbitan-myristoleylate, polyoxyethylene -sorbitan-palmitoleinylate, polyoxyethylene- sorbitan-p- etroselinylate, polyhydroxyethylcne-long fatty chain ethers, for example polyhydroxyethylene-acyl ethers, such as polyhydroxyethylene-lauryl ethers,
  • polyhydroxyethylene-myristoyl ethers polyhydroxyethylene-cetylst- earyl, polyhyd roxyethylene-palmityl ethers, polyhyd roxyethylene-oleoyl ethers, polyhydroxyethylene- palmitoleoyl ethers, polyhydroxyethylene-lino- leyl, polyhydroxyethylen-4, or 6, or 8, or 10, or 12-lauryl, miristoyl, palmitoyl, palmitoleyl, oleoyl or linoeyl ethers (Brij series), or in the corresponding esters, polyhydroxyethylen-laurate, -myristate, -palmitate, -stearatc or -oleate, especially polyhydroxyethylen-8-stearate (Myrj 45) and polyhydroxyethylen-8-oleate, polyethoxylated castor oil 40 (Cremophor EL), sorbitane-mono
  • lysophospholipids n-octadecylene-glycerophosphatidic acid, octadecylene- phosphorylglycerol, octadecylene-phosphorylserine, n-long fatty chain-glycero-phosphatidic acids, such as n-acyl-glycero-phosphatidic acids, especially lauryl glycero-phosphatidic acids, oleoyl-glycero-phosphatidic acid, n-long fatty chain-phosphorylglyccrol, such as n- acyl-phosphorylglycerol, especially lauryl-, myristoyl-, oleoyl- or palmitoeloyl- phosphorylglycerol.
  • n-long fatty chain-phosphoryl serine such as n-acyl-phosphorylserine, especially lauryl-, myristoyl-, oleoyl- or palmitoeloyl-phosphorylserine, n-tetradecyl-glycero- phosphatidic acid, n-tetradecyl-phosphorylglycerol, n-tetradecyl-phosphorylserine, corresponding-, elaidoyl-, vaccenyl-lysophospholipids, corresponding short-chain
  • Surfactant chains are typically chosen to be in a fluid state or at least to be compatible with the maintenance of fluid-chain state in carrier aggregates.
  • Table 5 lists preferred surfactants in accordance with the disclosure.
  • the surfactant is a nonionic surfactant.
  • the surfactant may be present in the formulation in about 1% to about 10%, about 1% to about 4%, about 4% to about 7% or about 7% to about 10% by weight.
  • the nonionic surfactant is selected from the group consisting of: polyoxyethylene sorbitans (polysobate surfactants), polyhydroxyethylene stearates or polyhydroxyethylene laurylethcrs (Brij surfactants).
  • the surfactant is a polyoxyethylene-sorbitan- monooieate (e.g. polysorbate 80 or Tween 80).
  • the polysorbate can have any chain with 12 to 20 carbon atoms.
  • the polysorbate is fluid in the formulation, which may contain one or more double bonds, branching, or cyclo-groups.
  • the topical vaccine formulations contain 1 to 10% by weight, 1 to 15 % by weight, 1 to 20% by weight, or 1 to 30% of a hapten-carrier conjugate. In some embodiments, the topical vaccine formulations contain 1 to 10% by weight, 1 to 15 % by weight. 1 to 20% by weight, or 1 to 30% by weight of the lipid. In some embodiments, the topical vaccine formulations contain 1 to 10% by weight, 1 to 15 % by weight, 1 to 20% by weight, 1 to 30% by weight, 1 to 40%) by weight, or 1 to 50% by weight of the surfactant.
  • the topical vaccine formulations may have a range of lipid to surfactant ratios, f he ratios may be expressed in terms of molar terms (mol lipid /mol surfactant).
  • the molar ratio of lipid to surfactant in the topical vaccine formulations is from about 1 :2 to about 10: 1.
  • the ratio is from about 1 : 1 to about 2:1, from about 2: 1 to about 3:1, from about 3 : 1 to about 4: 1, from about 4: 1 to about 5 : 1 , or from about 5:1 to about 10: 1.
  • the lipid to surfactant ratio is about 1.0, about 1.25, about 1.5, about 1.75, about 2.0, about 2.5, about 3.0, or about 4.0.
  • the topical vaccine formulations may have varying ratios of the hapten-carrier conjugate to lipid.
  • the ratios may be expressed in terms of molar ratios (mol hapten-carrier conjugate /mol lipid).
  • the molar ratio of the hapten-carrier conjugate to lipid in the topical vaccine formulations is from about 1 :50 to about 50: 1, from about 1 :25 to about 25: 1, from about 1 :10 to about 10:1, from about 1 :5 to about 5:1, from about 1 :50 to about 50:1 , or from about 0.2:1 to about 2:1.
  • the ratio is from about 0.2:1 to about 0.7:1 , from about 0.7:1 to about 1.2: 1 , from about 1.2:1 to about 1 ,7:1, or from about 1.7:1 to about 2:1.
  • the topical vaccine formulations may also have varying amounts of total amount of the following three components: the hapten-carrier conjugate, lipid and surfactant combined (TA).
  • the TA amount may be stated in terms of weight percent of the total composition.
  • the TA is from about 1% to about 40%, about 5% to about 30%, about 7.5% to about 15%, about 5% to about 10%. about 10% to about 20%, or about 20% to about 30%.
  • the TA is 8%, 9%, 10%. 15%, or 20%.
  • the topical vaccine formulations optionally contain one or more of the following ingredients: co-solvents, chelators, buffers, antioxidants, preservatives, mierobicides, emollients, humectants, lubricants, and thickeners. Preferred amounts of optional components are described in Table 7.
  • the topical vaccine formulations include a buffer to adjust the pH of the aqueous solution to a range from pH 3.5 to pH 9.5, pH 4 to pH 7.5. or pH 4 to pH 6.5.
  • buffers include, but are not limited to, acetate buffers, lactate buffers, phosphate buffers, and propionate buffers.
  • the buffer concentration is in the range between about 1 mM and about 1 M, about 1 mM and about 100 mJVl, or about 1 mM and about 10 mM.
  • the topical vaccine formulations are formulated in aqueous media.
  • the formulations may be formulated with or without co- solvents, such as lower alcohols.
  • a ""microbicide”” or “"antimicrobial”” agent is commonly added to reduce the bacterial count in pharmaceutical formulations.
  • Some examples of mierobicides arc short chain alcohols, including ethyl and isopropyl alcohol, chlorbutanol, benzyl alcohol, chlorbenzyl alcohol, dichlorbenzylalcohol, hexachlorophene; phenolic compounds, such as cresol, 4-chloro-m-cresoI, p-chloro-m-xylenol, dichlorophene, hexachlorophene, povidon- iodine; parabenes, especially alkyl-parabenes, such as methyl-, ethyl-, propyl-, or butyl- paraben, benzyl paraben; acids, such as sorbic acid, benzoic acid and their salts; quaternary ammonium compounds, such as alkonium salts, e.g., a bromine
  • Examples of '"antioxidants” are butylated hydroxyanisol (BHA), butylated hydroxytoluene (BHT) and di-tert-butylphenol (LY178002, LY256548, HWA-131, BF-389, CI-986, PD- 127443, E-5119, BI-L-239XX, etc.), tertiary butylhydroquinone (TBHQ), propyl gallate (PG), l-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ); aromatic amines
  • BHA butylated hydroxyanisol
  • BHT butylated hydroxytoluene
  • LY178002 di-tert-butylphenol
  • LY256548 LY256548, HWA-131, BF-389, CI-986, PD- 127443, E-5119, BI-L-239XX, etc.
  • TBHQ tertiary but
  • phenols and phenolic acids include tocopherols (alpha, beta, gamma, delta) and their derivatives, such as tocopheryl-acylate (e.g., -acetate, - laurate, myristate, -palmitate, -oleate, -linoleate, etc., or any other suitable tocopheryl- lipoate), tocopheryl-polyoxyethyle ⁇ e-succinate; trolox and corresponding amide and thiocarboxamide analogues; ascorbic acid and its salts, isoascorbate, (2 or 3 or 6)-o
  • oxidized compounds such as sodium bisulphite, sodium metabisulphite, thiourea
  • chelating agents such as ethylene glycol-bis-(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EDTA), ethylenedioxy-diethylene- dinitrilo-tetraacetic acid (GDTA), desferral
  • miscellaneous endogenous defence systems such as transferrin, lactoferrin, ferritin, cearuloplasmin, haptoglobion, heamopexin, albumin, glucose, ubiquinol-10
  • enzymatic antioxidants such as superoxide dismutase and metal complexes with a similar activity, including catalase, glutathione peroxidase, and less complex molecules, such as beta-carotene, bilirubin, uric acid
  • flavonoids flavones, flavonols, flavonones,
  • thickickeners are used to increase the viscosity of pharmaceutical formulations to and may be selected from selected from pharmaceutically acceptable hydrophilic polymers, such as partially etherified cellulose derivatives, comprising carboxymethyl-, hydroxyethyl-, hydroxypropyl-, hydroxypropylmethyl- or methyl-cellulose; completely synthetic hydrophilic polymers comprising polyacrylates, polymethacrylates,
  • ollients include, but are not limited to, glycerine, propylene glycol, sorbitol, and mannitol.
  • the topical vaccine formulations further comprise a polar liquid medium.
  • the topical vaccine formulations may be administered in an aqueous medium.
  • the topical vaccine formulations provided herein may be in the form of a solution, suspension, emulsion, cream, lotion, ointment, gel, spray, film forming solution, lacquer or a patch soaked with the formulation.
  • Table 7 lists preferred excipients for the formulation.
  • formulations provided herein may form vesicles or ESAs characterized by their adaptability, deformability, or penetrability.
  • vesicle or aggregate ""adaptability" which governs the ""tolerable surface curvature”
  • the vesicles provided herein may be characterized by their ability to adjust the aggregates * shape and properties to the anisotropic stress caused by pore crossing.
  • Sufficient adaptability implies that a vesicle or an aggregate can sustain different unidirectional forces or stress, such as one caused by pressure, without extensive fragmentation, which defines a ""stable”"' aggregate.
  • a “"barrier” in the context of this disclosure is (as in, for example, EP 0 475 160 and WO 98/17255) a body with through-extending narrow pores, such narrow pores having a radius which is at least 25% smaller than the radius of the ESAs (considered as spherical) before said ESAs permeate through such pores.
  • lipid vesicles made from any common phosphatidylcholine in the gel lamellar phase or else from any biological phosphatidylcholine/cholesterol 1/1 mol/mol mixture or else comparably large oil droplets, all having the specified relative diameter, are three examples for such non-adaptable aggregates.
  • the term """stable" means that the tested aggregates do not change their diameter spontaneously or under the transport related mechanical stress (e.g. during passage through a semipermeable barrier) unacceptably, which most often means only to a pharmaceutically acceptable degree. A 20-40% change is normally considered acceptable; the halving or doubling of aggregate diameter is borderline and a greater change in diameter is typically unacceptable. Alternatively and very conveniently, the change in aggregate diameter resulting from pore crossing under pressure is used to assess system stability; the same criteria are then applied as for ""narrow”" pores, mutatis mutandis. To obtain the correct value for aggregate diameter change, a correction for flux/vortex effects may be necessary. These procedures are described in greater detail in the publications of the applicant in Cevc el al., Biochim. Biophys. Acta 2002; 1564:21-30.
  • Non-destructing passage of ultradeformable, mixed lipid aggregates through narrow pores in a semi-permeable barrier is thus diagnostic of high aggregate adaptability. If pore radius is two times smaller than the average aggregate radius the aggregate must change its shape and surface-to- volume ratio at least 100% to pass without fragmentation through the barrier.
  • An easy and reversible change in aggregate shape inevitably implies high aggregate deformability and requires large surface-to-volume ratio adaptation.
  • a change in surface-to- volume ratio per se implies: a) high volume compressibility, e.g. in the case of compact droplets containing material other than, and immiscible with, the suspending fluid; b) high aggregate membrane permeability, e.g. in the case of vesicles that are free to exchange fluid between inner and outer vesicle volume.
  • Also provided herein are methods of administering a pharmaceutical composition comprising a hapten-carrier conjugate, a lipid, and a surfactant.
  • the hapten-carrier conjugate in the topical vaccine formulations elicits an immune response in a cell, tissue, organ, and/or subject or patient when administered,
  • the terms "''subject'"' or ""patient”" are used
  • the terms ""subject"'" and '""subjects” refer to an animal (e.g. , birds, reptiles, and mammals), preferably a mammal including a non-primate (e.g., a camel, donkey, zebra, cow, pig, horse, goat, sheep, cat, dog, rat, and mouse) and a primate (e.g., a monkey, chimpanzee, and a human), and most preferably a human.
  • the subject or patient has a drug addiction.
  • the subject or patient is at risk for developing or re-developing a drug addiction.
  • the hapten-carrier conjugate in the topical vaccine formulations induce an immune response from the adaptive immune system, such as a B cell response and/or a T cell response.
  • the immune response induced by the hapten-carrier conjugate in the topical vaccine formulations is an antibody response.
  • the hapten-carrier conjugate in the topical vaccine formulations induces a humoral immune response, such as an interferon response and/or an interleukin response, e.g. , an interleukin-4 response.
  • the hapten- carrier conjugate in the topical vaccine formulations induces one or more types of immune response but not another immune response.
  • the hapten-carrier conjugate induces a combination of immune responses.
  • the hapten-carrier conjugate in the topical vaccine formulations can induce a robust IFN response which has other biological consequences in vivo, affording protection against subsequent diseases or conditions or concurrent diseases or conditions.
  • the hapten-carrier conjugate in the topical vaccine formulations can induce a robust TNF ⁇ or interleukin response which has other biological consequences in vivo, affording protection against subsequent diseases or conditions or concurrent diseases or conditions.
  • the topical vaccine formulation comprising the hapten- carrier conjugate may be administered to a na ⁇ ve subject, i.e., a subject that does not have a disease, condition, drug addiction, or has not been and is not currently infected with an infectious agent.
  • the topical vaccine formulation comprising the hapten-carrier conjugate may be administered to a na ⁇ ve subject that is predisposed to acquiring such disease, condition, drug addiction, or infection.
  • the topical vaccine formulation comprising the hapten-carrier conjugate may also be
  • the topical vaccine formulation is administered as a patch.
  • Patches contemplated for use with the topical vaccine formulations of the invention include those disclosed in the following references, herein incorporated by reference in their entirety: """Transcutaneous immunization with heat-labile enterotoxin: development of a needle-free vaccine patch,"" Expert Rev Vaccines 2007;6(5): 809-819; ""Transcutaneous immunization with the heat-labile toxin (LT) of enterotoxigenic Escherichia coli (ETEC): protective efficacy in a double-blind, placebo-controlled challenge study,-' * ' Vaccine 2007; 25(18):3684- 3691 ; Immunization using the skin: new approaches,-'" In: Moingeon P (Ed.), Vaccines, Frontiers in Design and Development, Horizon Scientific Press, Ltd., Wymondham, UK, pp.
  • the patch further comprises one or more antipuritics, adhcsives, solvents, co-solvents, plasticizers, polymeric matrices, stabilizers, thickeners, preservatives, and the like.
  • topical antipruritics effective in reducing itching during transdermal nicotine delivery include bisabolol or ⁇ (-)bisabolol, oil of chamomile, chamazulene, allantoin, D-panthenol glycyrrhetenic acid, corticosteroids, antihistamines, and combinations thereof.
  • the antipruritic is a combination of 18- ⁇ -glycyrrhetenic acid and D-panthenol (i.e., ANTIPHLOGISTICUM "ARO").
  • Examples of pharmaceutically acceptable pressure sensitive adhesives include acrylic, silicone, vinyl acetate and synthetic or natural rubber adhesives as well as other adhesives useful in transdermal drug delivery.
  • the adhesives may be used alone or in combination to prepare an adhesive drug matrix or may be applied to the skin-contacting surface of a polymeric matrix or reservoir patch to adhere said patch to the skin.
  • Examples of adhesives are acrylic adhesives such as RA 2484, RA 2333, RA 2397, R 363 and R 362 from Monsanto Co.
  • Other acrylic adhesives such as Durotak, manufactured by Morton Thiokol, Inc., and Neocryl XA5210bby Polyvinyl Chemicals, Ltd. may be utilized.
  • Vinyl acetate adhesives include Flexcryl-1614, 1617, 1618 and 1625 from Air Products. Numerous silicone based adhesives may be used, such as Q72929, Q27406, X72920 and 355, each manufactured by Dow-Corning. Natural and synthetic rubbers include polyisobutylenes, neoprenes, polybutadienes and polyisoprenes. [0113] Polymeric matrix-forming agents include pharmaceutically acceptable polymers such as polyvinyl alcohol, polyvinylpyrolidones, gelatin and partially hydro lyzed polyvinyl alcohols.
  • solvents include water and polar organic solvents, such as ethanol, and mixtures thereof.
  • the patched comprise co-solvents, for example, mineral oil, silicone-based liquids, and low molecular weight polyisobutylenes.
  • preservatives, antioxidants and chelating agents examples include butylated hydroxyanisole (BHA), butylatcd hydroxytoluene (BHT), sodium metabisulfate, ⁇ -tocopherol, maleic acid, ethylenediaminetetraacetic acid (EDTA), and cysteine hydrochloride.
  • BHA butylated hydroxyanisole
  • BHT butylatcd hydroxytoluene
  • sodium metabisulfate sodium metabisulfate
  • ⁇ -tocopherol maleic acid
  • EDTA ethylenediaminetetraacetic acid
  • cysteine hydrochloride examples include butylated hydroxyanisole (BHA), butylatcd hydroxytoluene (BHT), sodium metabisulfate, ⁇ -tocopherol, maleic acid, ethylenediaminetetraacetic acid (EDTA), and cysteine hydrochloride.
  • Components useful for imparting the desired wear and pharmacokinetic characteristics to the patch include, for example, polymeric matrix-forming materials added to facilitate curing of the adhesives, for example Aerotex Resin 3730 (American Cyanamid), and a thickener added to adjust the viscosity of the polymer mixture to the desired viscosity for coating on a backing material.
  • the thickener can be an acrylic polymer thickener such as AMSCO 6038 A (Unocal), methyl cellulose and hydroxypropylmethyl cellulose.
  • Plasticizers may be added to impart softness and flexibility to the adhesive, a typical plasticizer being glycerin.
  • a rate-controlling membrane may be included to effect controlled release of hap ten-carrier conjugate from the patch.
  • Materials suitable for rate-controlling membranes include cthylene-vinyl acetate (EVA) copolymer membranes (e.g. 1-20% vinyl acetate), polyvinylalcohol (PVA) gels and silicone films.
  • the patch comprises a) a backing layer which is impermeable to the components of the adhesive layer b), b) an adhesive layer capable of releasing the hapten-carrier conjugate and consisting of a permeable polymeric material which is compatible with the skin and contains at least one topical vaccine formulation which is capable of permeation across the skin, a combination of eucalyptol having a purity of at least 70% with an additional flux enhancer and further optional pharmaceutical excipients, and c) a protective release liner which can be peeled from the adhesive contact layer b).
  • the backing layer comprises a material or a combination of materials which must be impermeable to the components of the adhesive layer, especially to liquid components of the formulation.
  • the backing layer may be made of high- or low- density polymers such as polyethylene, polypropylene, polyvinyl chloride, polyethylene terephthalate or also cellulose acetate or vinyl acetate/vinyl chloride copolymers and combinations thereof, preferably of composite films.
  • the adhesive layer b) is located between the backing layer a) and the release liner c) and consists of a polymeric material which is compatible with the skin and is in the form of a matrix which contains the topical vaccine formulation and, for enhancing penetration through the skin, a combination of eucalyptol and an additional flux enhancer and further optional excipients.
  • Suitable polymeric materials are permeable to the topical vaccine formulation as well as to the combination of eucalyptol and the additional flux enhancer and, besides being compatible to the skin, have sufficient adhesiveness that the therapeutic system remains attached to the skin for at least one day and can subsequently be removed without exerting additional force. Further, suitable polymeric materials are dimensionally stable, despite the solvent properties of eucalyptol and the chemically pure 1 ,8-cineol.
  • silicone rubber for example linear polysiloxanes in which the silicon atoms in the siloxane chain are substituted by two identical or different alkyl groups such as methyl or ethyl groups, aryl groups, for example phenyl groups, alkcnyl groups such as vinyl or allyl groups, alkylaryl groups such as tolyl or xylyl groups, or alkyl groups, for example benzyl groups, and the terminal silicon atoms are substituted by three of the above organic radicals.
  • the preparation of these silicones is disclosed in U.S. Pat. Nos.
  • Silicones which are vulcanisable at room temperature are preferred.
  • Further suitable polymeric materials are hydrophilic polymers of monoesters of unsaturated carboxylic acids such as acrylic acid or methacrylic acid, for example the polyhydroxyethylacrylates or
  • polyhydroxyethylmethacrylates thereof the preparation of which is disclosed in U.S. Pat. Nos. 2,976,576 and 3,220,960, as well as copolymers of water-soluble aliphatic or cyclic vinyl amides, for example poly-N-vinylmethylacetamide, poly-N-vinylethy ⁇ acetamide, poly- N-vinyl-methylpropionamide, poly-N-vinylethylpropionamide, poly-N-vinylmethyl- isobutyramide, poly-N-vinyl-2-pyrrolidone, poly-N-vmyl-2-piperidone, poly-N-vinyl- .epsilon.-caprolactam, poly-N-vinyl-5-methyl-2-pyrrolidone or poly-N-vinyl-3-methyl-2- pyrrolidone, preferably poly-N-vinylpyrrolidone having an average molecular weight of about 10,000 to 360,000, with water-soluble poly
  • Preferred polymers are natural or synthetic rubber, for example polyisoprene, 1 ,4-butadiene polymer or polyisobutylene and mixtures thereof. Particularly preferred polymers are mixtures of different polyisobutylenes, with molecular weight ranges from about 1.0 x IQ 3 to about 5.0 x 10 4 , or from about 1.0 x 10 3 to about 5.0 x 10 6 .
  • the term "eucalyptol having a degree of purity of at least 70%'' includes formulations containing more than 70% to 100% of 1,8-cineol.
  • formulations containing about 70-95% of 1,8-cineol are also designated as eucalyptus oils, whereas the term “eucalyptol” is used for formulations containing more than 95% of 1 ,8-cineol.
  • Eucalyptus oils are terpentine-containing essential oils which contain eucalyptol or 1,8-cineol as main constituent (more than 70%) and which can be isolated from leaves, roots or the bark of eucalyptus plants of the species Eucalyptus globulus (common eucalyptus tree), Eucalyptus maculata, Eucalyptus cladocalyx or Eucalyptus sideroxylon.
  • this eucalyptus oil can be further processed to chemically pure 1 ,8-cineol (more than 99% pure).
  • the adhesive layer b) preferably contains this chemically pure 1,8-cineol together with an additional flux enhancer.
  • Flux enhancers also known in the literature as penetration enhancers, accelerators or sorption promoters, have the property of permitting the passage of drugs through the skin that are in themselves impermeable to the skin, especially the stratum corneum which functions as a barrier, and of enhancing the permeability of drugs (e.g., hapten-carrier conjugates) and so make possible the permeation and absorption of therapeutic amounts of the drug to be delivered.
  • the use of a combination of eucalyptol, preferably in the form of chemically pure 1,8-cineol, together with an additional flux enhancer, may substantially increases the amount released and the rate of release and the corresponding absorption (or rate of absorption) through the skin.
  • additional flux enhancers are monohydric saturated or unsaturated aliphatic, cycloaliphatic or aromatic alcohols of 1 to 12 carbon atoms, for example ethanol, isopropanol, n-hexanol or cyclohexanol; aliphatic, cycloaliphatic or aromatic hydrocarbons of 5 to 12 carbon atoms, for example hexane, cyclohexane, isopropyl benzene and the like; cycloaliphatic or aromatic aldehydes and ketones of 4 to 10 carbon atoms, for example cyclohexanone; amides such as acetamide, N,N-diethyl-m- toluamide, N,N-di-lower alkylacetamide such as N,N-dimethylacetamide or N 5 N- diethylacetamide, dimethyl propionamide; Cio-C 2 o alkanoylamides, for example N 5 N- dimethyllauroy
  • the flux enhancer is a combination of chemically pure 1 ,8- cineol with N-methyl-2-pyrrolidone, for example a combination of 5 to 9.5 parts by weight of 1,8-cineol and 5 to 0.5 parts by weight (based on 10 parts by weight) of N-methyl-2- pyrrolidone.
  • such combinations have the advantage that they can be processed with the polymeric material of the adhesive layer b), for example polyisobutylene mixtures, to homogeneous matrix systems of sufficient adhesiveness which are capable of drug release.
  • the flux enhancer is combination of about 9 parts of chemically pure 1,8-cineol and 1 part by weight of N-methyl-2-pyrrolidone is especially preferred.
  • the adhesive layer may optionally further comprise one or more of water, isotonic aqueous saline solution, dextrose in water or saline solution, liquid glyceryl triesters with low molecular fatty acids, lower alkanols, natural oils such as corn oil, groundnut oil, sesame oil, castor oil and the condensates thereof with ethylene oxide and the like, hydrocarbons such as mineral oil of pharmaceutical quality, silicones, emulsifiers such as monoglycerides or diglycerides of fatty acids, phosphatidic acid derivatives such as lecithin or cephalin, polyalkyene glycols such as polyethylene glycol, aqueous phases to which a swelling agent such as sodium carboxymethyl cellulose has been added, sodium alginate, polyvinylpyrrolidone and the like, to which dispersants or emulsifiers such as lecithin may also be added, polyoxyethylene and the like.
  • water isotonic aqueous s
  • the patch comprises a pressure-sensitive adhesive composition suitable as a matrix for controlled release of the topical vaccine formulation therefrom comprising a blend of a rubber-based pressure-sensitive adhesive and a soluble polyvinylpyrrolidone (PVP).
  • PVP polyvinylpyrrolidone
  • the term '"polyvinylpyrrolidone," or 'TVP refers to a polymer, either a homopolymer or copolymer, containing N-vinylpyrrolidone as the monomeric unit.
  • Typical PVP polymers are homopolymeric PVPs and the copolymer vinyl acetate vinylpyrrolidone.
  • the homopolymeric PVPs are known to the pharmaceutical industry under a variety of designations including Povidone, Polyvidone, Polyvidonum, Polyvidonum solubile, and PoIy(I -vinyl-2-pyrrolidone).
  • the copolymer vinyl acetate vinylpyrrolidone is known to the pharmaceutical industry as Copolyvidon, Copolyvidone, and Copolyvidonum.
  • the term '"soluble PVP-' means that the PVP is soluble in water and generally is not substantially cross-linked, and has a molecular weight of less than about 2,000,000.
  • the rubber-based pressure-sensitive adhesive is polysiloxane.
  • Polysiloxane is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 9% to about 97%, from about 8% to about 97%, or from about 14% to about 94% of the total weight of the polymer blend, by weight of the total pressure-sensitive adhesive composition.
  • Suitable polysiloxanes include silicone pressure-sensitive adhesives which are based on two major components: a polymer, or gum, and a tackifying resin.
  • the polysiloxane adhesive is usually prepared by cross-linking the gum, typically a high molecular weight polydiorganosiloxane.
  • Suitable silicone pressure-sensitive adhesives are commercially available and include the silicone adhesives sold under the trademarks BIO-PSA X7-3027, BIO-PSA X7- 4503, BIO-PSA X7-4603, BIO-PSA X7-4301 , BIO-PSA X7-4303, BIO-PSA X7-4919, BIO- PSA X7-2685, and BIO-PSA X7-3122 by Dow Corning Corporation, Medical Products, Midland, Mich.
  • pressure-sensitive adhesive composition comprises a ternary blend comprising a rubber-based pressure-sensitive adhesive, a polyacrylate polymer, and a soluble PVP, wherein the rubber-based pressure-sensitive adhesive is a polysiloxane.
  • Polysiloxane is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 9% to about 97% by weight of the total pressure-sensitive adhesive composition, while the polyacrylate polymer is preferably present in an amount ranging from about 5% to about 85%.
  • the ratio of the polyacrylate polymer to the rubber-based pressure-sensitive adhesive is from about 2:98 to about 96:4, and more preferably from about 2:98 to about 86: 14 by weight.
  • blends comprising a rubber-based pressure-sensitive adhesive and a polyacrylate polymer, wherein the rubber-based pressure-sensitive adhesive is a polysiloxane.
  • Polysiloxane is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 9% to about 97% by weight of the total pressure-sensitive adhesive composition, while the polyacrylate polymer is preferably present in an amount ranging from about 5% to about 85%.
  • the ratio of the polyacrylate polymer to the rubber-based pressure-sensitive adhesive is from about 2:98 to about 96:4, and more preferably from about 2:98 to about 86: 14 by weight.
  • soluble PVP is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 1% to about 20% by weight of the total pressure-sensitive adhesive composition.
  • the pressure-sensitive adhesive compositions may further optionally comprise enhancers, fillers, co-solvents, and excipients.
  • a multiple polymer adhesive system comprises a blend of 14-94% by weight of a rubber-based pressure-sensitive adhesive, 5-85% by weight of a polyacrylate polymer, and 2-10% by weight of a soluble PVP, and the multiple polymer adhesive system comprises about 50-99% by weight of the dermal adhesive composition.
  • This multiple polymer adhesive system is combined with a topical vaccine formulation in the amount of 0.1-50% by weight of the total dermal adhesive composition.
  • Optional additives, such as co-solvent for the drug (up to 30% by weight) and enhancers (up to 20% by weight) may be included in the dermal adhesive composition.
  • the topical vaccine formulation does not result in complete protection or cure (i.e., from drug addiction), but results in a lower level of addiction compared to an untreated subject.
  • Benefits include, but are not limited to, reduced severity of symptoms of the disease or condition and a reduction in the duration of the disease or condition
  • the topical vaccine formulation is used to protect against a disease or condition (e.g., an infection or drug addiction) in na ⁇ ve subjects.
  • the prophylactic and/or therapeutic effect of the topical vaccine formulation are based, in part, upon achieving or inducing an immune response (e.g., a humoral immune response or adaptive immune response).
  • the immunogenic compositions induce a detectable serum titer of an antibody against an antigen or hapten in either the subject or an animal model thereof (e.g. mouse, rat, pig, goat, sheep or canine model).
  • the serum titer of an antibody can be determined using techniques known to one of skill in the art. e.g., immunoassays such as ELISAs.
  • the antibodies generated by administering an immunogenic composition of the invention are neutralizing antibodies.
  • administration of the topical vaccine formulation to a subject or animal model thereof results in a serum titer of about 1 ⁇ g/ml, about 2 ⁇ g/ml, about 5 ⁇ g/ml, about 6 ⁇ g/ml, about 10 ⁇ g/ml, about 15 ⁇ g/ml, about 20 ⁇ g/ml, about 25 ⁇ g/ml , about 50 ⁇ g/ml, about 75 ⁇ g/ml, about 100 ⁇ g/ml, about 125 ⁇ g/ml, about 150 ⁇ g/ml, about 175 ⁇ g/ml, about 200 ⁇ g/ml, about 225 ⁇ g/ml, about 250 ⁇ g/ml, about 275 ⁇ g/ml, about 300 ⁇ g/ml, about 325 ⁇ g/ml, about 350 ⁇ g/ml, about 375 ⁇ g/ml, or more of an antibody that specifically binds to the hapten or anti
  • the serum titer is 100 ⁇ g/ml or more.
  • administration of the topical vaccine formulation results in a plasma titer of 100 ⁇ g/ml to 1 mg/ml or more, preferably more than about 500 ⁇ g/ml.
  • the immune response may be determined in the subject or in a animal model, which response is then correlated or extrapolated to a predicted response in the subject, e.g., a human or livestock, such as a pig, sheep, goat, or cow.
  • a method for preventing, treating, managing, or ameliorating at least one disease or condition (e.g., a drug addiction or viral infection) in a subject comprising administering to said subject a first dose of an effective amount of the topical vaccine formulation.
  • the dose of the topical vaccine formulation administered to the subject or animal model is about 10-20 ⁇ ig.
  • the dose of the topical vaccine formulation administered to the subject or animal model is about 75-100 ⁇ g.
  • the dose of the topical vaccine formulation administered to the subject or animal model is about 500-1000 ⁇ g.
  • the amount of the topical vaccine formulation which will be effective in the treatment, prevention and/or amelioration of a particular disease or condition will depend on the nature of the disease, and can be determined by standard clinical techniques. In addition, in vitro assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed in the topical vaccine formulation will also depend on, for example, the seriousness of the disease or disorder, and should be decided according to the judgment of the practitioner and each subject's circumstances. Effective doses may be extrapolated from dose response curves derived from in vitro or animal model test systems.
  • the topical vaccine formulation is administered to a subject in combination with one or more other therapies for the prevention or treatment of at least one disease or condition.
  • the therapies e.g., prophylactic or therapeutic agents
  • the therapies are administered less than 5 minutes apart, less than 30 minutes apart, 1 hour apart, at about 1 hour apart, at about 1 to about 2 hours apart, at about 2 hours to about 3 hours apart, at about 3 hours to about 4 hours apart, at about 4 hours to about 5 hours apart, at about 5 hours to about 6 hours apart, at about 6 hours to about 7 hours apart, at about 7 hours to about 8 hours apart, at about 8 hours to about 9 hours apart, at about 9 hours to about 10 hours apart, at about 10 hours to about 1 1 hours apart, at about 1 1 hours to about 12 hours apart, at about 12 hours to 18 hours apart, 18 hours to 24 hours apart, 24 hours to 36 hours apart, 36 hours to 48 hours apart, 48 hours to 52 hours apart, 52 hours to 60 hours apart, 60 hours to 72 hours apart, 72 hours to 84 hours apart,
  • the topical vaccine formulations are useful in the treatment or prevention of a disease, condition, or infection.
  • the condition is a drug addiction or drug allergy.
  • the drug of addiction is nicotine.
  • the drug of addiction is cocaine.
  • the topical vaccine formulations are specific for a hapten compared to cellular components and/or compared to the carrier.
  • the topical vaccine formulations exhibit low cytotoxicity in eukaryotic cells, preferably mammalian cells.
  • the topical vaccine formulation reduces or inhibits a drug addiction, dependence, or allergy.
  • the immunogenic composition eliminates or reduces the drug addiction or drug dependence by 75%, 80%, 85%, 90%, 95%, 98%, 99%, 75-99.5%, 85-99.5%, or 90-99.8% in a subject as determined by an assay described herein or known to one of skill in the art.
  • the topical vaccine formulations are useful in methods of preventing, treating and/or managing drug addiction or drug dependence.
  • the topical vaccine formulations is useful in preventing, treating and/or managing a disease or condition that exhibits resistance to other treatments.
  • the topical vaccine formulation inhibits or reduces the circulation in the bloodstream a drug or antigen against which it is targeted by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85% as measured by a standard assay known to one of skill in the art, or an assay described herein.
  • the topical vaccine formulation inhibits or reduces the penetration of the drug or antigen against which it is targeted from one organ, tissue or cell to another organ, tissue or cell as measured using a standard assay known to one of skill in the art, or an assay described herein.
  • the topical vaccine formulation inhibits or reduces the ability of a drug, such as cocaine or nicotine, to enter the brain by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85% as measured using a standard assay known to one of skill in the art, or an assay described herein.
  • a drug such as cocaine or nicotine
  • the present invention provides methods of preventing, treating and/or managing a disease or condition, such as a drug addiction, said methods comprising administering to a subject in need thereof one or more topical vaccine formulations.
  • the invention provides a method of preventing, treating/and or managing a cocaine addiction or a nicotine addiction.
  • the invention also provides methods of preventing, treating and/or managing a disease or condition, said methods comprising administering to a subject in need thereof one or more topical vaccine formulations, and one or more other therapies (e.g., prophylactic or therapeutic agents).
  • the other therapies are currently being used, have been used or are known to be useful in the prevention, treatment and/or management of the disease or condition.
  • Non-limiting examples of such prophylactic or therapeutic methods are provided below.
  • the combination therapies of the invention can be administered sequentially or concurrently.
  • the combination therapies comprise administering the topical vaccine formulation at least one other therapy which has the same mechanism of action.
  • the combination therapies comprise administering the topical vaccine formulation and at least one other therapy which has a different mechanism of action than the compound.
  • the combination therapies improve the prophylactic and/or therapeutic effect of an immunogenic composition of the invention by functioning together with immunogenic composition to have an additive or synergistic effect.
  • the combination therapies reduce the side effects associated with each therapy taken alone.
  • the topical vaccine formulation further comprises the prophylactic or therapeutic agents of the combination therapies.
  • the administration of the topical vaccine formulation effects administration of both the hapten-carrier conjugate and the prophylactic or therapeutic agents.
  • the prophylactic or therapeutic agents of the combination therapies can be administered concurrently to a subject in separate formulations.
  • the prophylactic or therapeutic agents may be administered to a subject by the same or different routes of administration.
  • the topical vaccine is administered to a subject, preferably a human, to prevent, treat and/or manage a drug addiction.
  • a subject preferably a human
  • the other prophylactic or therapeutic agents are currently being used, have been used or are known to be useful in the prevention, treatment and/or management of the drug addiction or a symptom or condition related to it, e.g., a psychiatric or psychological condition.
  • the topical vaccine formulation may be used as any line of therapy, e.g., a first, second, third, fourth or fifth line therapy, for a disease or condition.
  • the subject administered the topical vaccine formulation has not received a prior therapy.
  • the subject administered the topical vaccine formulation has received a prior therapy.
  • the subject administered the topical vaccine formulation has administered the topical vaccine
  • a method for treating and/or managing a disease or condition e.g., a drug addiction, in a subject refractory to conventional therapies for such a condition, the methods comprising administering to said subject a dose of a prophylactically or therapeutically effective amount of an immunogenic composition of the invention.
  • synthetic drugs peptides (including cyclic peptides), polypeptides, proteins, nucleic acids (e.g., DNA and RNA nucleotides including, but not limited to, antisense nucleotide sequences, triple helices, RNAi, and nucleotide sequences encoding biologically active proteins, polypeptides
  • agents include, but are not limited to, immunomodulatory agents (e.g., interferon), anti-inflammatory agents (e.g.. adrenocorticoids, corticosteroids (e.g., beclomethasone, budesonide, flunisolide, fluticasone, triamcinolone, methlyprednisolone, prednisolone, prednisone, hydrocortisone), glucocorticoids, steroids, and non-steroidal anti-inflammatory drugs (e.g., aspirin, ibuprofen, diclofenac, and COX-2 inhibitors), pain relievers, anti-psychotics.
  • immunomodulatory agents e.g., interferon
  • anti-inflammatory agents e.g.. adrenocorticoids, corticosteroids (e.g., beclomethasone, budesonide, flunisolide, fluticasone, triamcinolone, methlyprednisol
  • anti-depressants anti-anxiety drugs, anti- epileptics, leukotreine antagonists (e.g., montelukast, methyl xanthines, zafirlukast, and zileuton), beta2-agonists (e.g., albuterol, biterol, fenoterol, isoetharie, metaproterenol, pirbuterol, salbutamol, terbutalin formoterol, salmeterol, and salbutamol terbutaline), anticholinergic agents (e.g., ipratropium bromide and oxitropium bromide), sulphasalazine, penicillamine, dapsone, antihistamines, anti-malarial agents (e.g., hydroxychloroquine), antiviral agents (e.g., nucleoside analogs (e.g., zidovudine, acyclovir, gangcyclovir, vidarabine,
  • Any therapy which is known to be useful, or which has been used or is currently being used for the prevention, management, and/or treatment of the disease or condition, or a disease or condition associated with the disease or condition, can be used in combination with the topical vaccine formulation. See, e.g., Gilman et al., Goodman and Gilman's: The Pharmacological Basis of Therapeutics, 10th ed., McGraw-Hill, New York, 2001 ; The Merck Manual of Diagnosis and Therapy, Berkow, M.D. et al.
  • therapies e.g., prophylactic or therapeutic agents which have been or are currently being used for preventing, treating and/or managing various diseases or conditions.
  • antibacterial agents including antibiotics
  • topical vaccine formulation include, but are not limited to, aminoglycoside antibiotics, glycopeptides, amphenicol antibiotics, ansamycin antibiotics, cephalosporins, cephamycins oxazolidinones, penicillins, quinolones, streptogamins, tetracyciins, and analogs thereof.
  • the topical vaccine formulation is used in combination with other protein synthesis inhibitors, including but not limited to, streptomycin, neomycin, erythromycin, carbomycin, and spiramycin.
  • the antibacterial agent is selected from the group consisting of ampicillin, amoxicillin, ciprofloxacin, gentamycin, kanamycin, neomycin, penicillin G, streptomycin, sulfanilamide, and vancomycin.
  • the antibacterial agent is selected from the group consisting of azithromycin, cefonicid, cefotetan, cephalothin, cephamycin, chlortetracycline, clarithromycin, clindamycin, cycloserine, dalfopristin.
  • doxycycline erythromycin, linezolid, mupirocin, oxytetracycline, quinupristin, rifampin, spectinomycin, and trimethoprim.
  • amphenicol antibiotics e.g., azidamfenicol, chloramphenicol, florfenicol, and
  • ansamycin antibiotics e.g., rifamide and rifampin
  • carbacephems e.g., loracarbef
  • carbapenems e.g., biapenem and imipenem
  • cephalosporins e.g., cefaclor, cefadroxil, cefamandole, cefatrizine, cefazedone, cefozopran, cefpimizole, cefpiramide, and cefpirome
  • cephamycins e.g., cefbuperazone, cefmetazole, and cefminox
  • folic acid analogs e.g., trimethoprim
  • glycopeptides e.g., vancomycin
  • lincosamides e.g., clindamycin, and lincomycin
  • macrolides e.g., azithromycin, carbomycin, clarithomycin, dirithromycin,
  • Additional examples include cycloserine, mupirocin, tuberin amphomycin, bacitracin, capreomycin, colistin, enduracidin, enviomycin, and 2,4 diaminopyrimidines (e.g., brodimoprim).
  • antiviral agents that can be used in combination with the antibacterial agent is selected from the group consisting of ampicillin, amoxicillin, ciprofloxacin, gentamycin, kanamycin, neomycin, penicillin G, streptomycin, sulfanilamide, and vancomycin.
  • the antibacterial agent is selected from the group consisting of azithromycin, cefonicid, cefotetan, cephalothin, cephamycin, chlortetracycline, clarithromycin, clindamycin, cycloserine, dalfopristin, doxycycline, erythromycin, linezolid, mupirocin, oxytetracycline, quinupristin, rifampin, spectinomycin, and trimethoprim.
  • Antiviral agents include, but are not limited to, non-nucleoside reverse transcriptase inhibitors, nucleoside reverse transcriptase inhibitors, protease inhibitors, and fusion inhibitors,
  • the antiviral agent is selected from the group consisting of amantadine, oseltamivir phosphate, rimantadine, and zanamivir.
  • the antiviral agent is a non-nucleoside reverse transcriptase inhibitor selected from the group consisting of delavirdine, efavirenz, and nevirapine.
  • the antiviral agent is a nucleoside reverse transcriptase inhibitor selected from the group consisting of abacavir, didanosine, emtricitabine, emtricitabine, lamivudine, stavudine, tenofovir DF, zalcitabine, and zidovudine.
  • the antiviral agent is a protease inhibitor selected from the group consisting of amprenavir, atazanavir, fosamprenav, indinavir, lopinavir, nelfinavir, ritonavir, and saquinavir.
  • the antiviral agent is a fusion inhibitor such as enfuvirtide.
  • nucleoside reverse transcriptase inhibitors e.g., AZT, ddT, ddC, 3TC, d4T
  • non-nucleoside reverse transcriptase inhibitors e.g., dclavird
  • anti-viral agents include but are not limited to acemannan; acyclovir; acyclovir sodium; adefovir; alovudine: alvircept sudotox; amantadine hydrochloride
  • cipamfylline cipamfylline
  • cytarabine hydrochloride delavirdine mesylate
  • desciclovir didanosine
  • disoxaril edoxudine; enviradene; enviroxime; famciclovir; famotine hydrochloride;
  • fiacitabine fialuridine; fosarilate; foscamet sodium; fosfonet sodium; ganciclovir; ganciclovir sodium; idoxuridine; kethoxal; lamivudine; lobucavir; memotine hydrochloride; methisazone; nevirapine; oseltamivir phosphate (TAMIFLUTM); penciclovir; pirodavir; ribavirin;
  • rimantadine hydrochloride FLUMADINETM
  • saquinavir mesylate somantadine hydrochloride
  • sorivudine sorivudine
  • statolon sorivudine
  • stavudine tilorone hydrochloride
  • trifluridine rimantadine hydrochloride
  • valacyclovir hydrochloride vidarabine; vidarabine phosphate; vidarabine sodium phosphate; viroxime; zalcitabine; zanamivir (RELENZATM); zidovudine; and zinviroxime.
  • the amount of topical vaccine formulation, and the amount of hapten-carrier conjugate within the topical vaccine formulation, that will be effective in the prevention, treatment and/or management of a disease or condition can be determined by standard clinical techniques. In vitro or in vivo assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed will also depend, e.g., on the route of administration, the type of disease or condition to be treated, and the seriousness of the disease or condition, and should be decided according to the judgment of the practitioner and each patient's or subject's circumstances.
  • Exemplary doses include milligram or microgram amounts per kilogram of subject or sample weight (e.g., about 1 microgram per kilogram to about 500 milligrams per kilogram, about 5 micrograms per kilogram to about 100 milligrams per kilogram, or about 1 microgram per kilogram to about 50 micrograms per kilogram).
  • a daily dose is at least 50 mg. 75 mg, 100 mg, 150 mg, 250 mg, 500 mg, 750 mg, or at least 1 S-
  • the dosage is a concentration of 0.01 to 5000 mM, 1 to 300 mM, 10 to 100 mM and 10 mM to 1 M. In another embodiment, the dosage is a
  • the dosage is a concentration of 0.01 to 5000 mM, 1 to 300 mM, 10 to 100 mM and 10 mM to 1 M. In another embodiment, the dosage is a
  • the dosage is 0.25 ⁇ g/kg or more, preferably 0.5 ⁇ g/kg or more, 1 ⁇ g/kg or more, 2 ⁇ g/kg or more, 3 ⁇ g/kg or more, 4 ⁇ g/kg or more, 5 ⁇ g/kg or more, 6 ⁇ g/kg or more, 7 ⁇ g/kg or more, 8 ⁇ g/kg or more, 9 ⁇ g/kg or more, or 10 ⁇ g/kg or more.
  • 25 ⁇ g/kg or more preferably 50 ⁇ g/kg or more, 100 ⁇ g/kg or more, 250 ⁇ g/kg or more, 500 ⁇ g/kg or more, 1 mg/kg or more, 5 mg/kg or more, 6 mg/kg or more, 7 mg/kg or more, 8 mg/kg or more, 9 mg/kg or more, or 10 mg/kg or more of a patient's body weight.
  • the dosage is a unit dose of 10-20 ⁇ g, 20-50 ⁇ g, 50-75 ⁇ g, 75-100 ⁇ g, 100-200 ⁇ g, 200-300 ⁇ g, 300-400 ⁇ g, 400-500 ⁇ g, 500-600 ⁇ g, 600-700 ⁇ g, 700- 800 ⁇ g, 800-900 ⁇ g, or 900-1000 ⁇ g, or more.
  • the dosage is a unit dose of 5 mg, preferably 10 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg. 350 mg, 400 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg or more.
  • the dosage is a unit dose that ranges from about 5 mg to about 100 mg, preferably about 100 mg to about 200 ⁇ g, about 150 mg to about 300 mg, about 150 mg to about 400 mg, 250 ⁇ g to about 500 mg, about 500 mg to about 800 mg, about 500 mg to about 1000 mg, or about 5 mg to about 1000 mg.
  • a subject is administered one or more doses of a prophylactically or therapeutically effective amount of the topical vaccine formulation, wherein the prophylactically or therapeutically effective amount is not the same for each dose, in another embodiment, a subject is administered one or more doses of a
  • prophylactically or therapeutically effective amount of the topical vaccine formulation wherein the dose of a prophylactically or therapeutically effective amount administered to said subject is increased by, e.g., 0.01 ⁇ g/kg, 0.02 ⁇ g/kg, 0.04 ⁇ g/kg, 0.05 ⁇ g/kg, 0.06 ⁇ g/kg, 0.08 ⁇ g/kg, 0.1 ⁇ g/kg, 0.2 ⁇ g/kg, 0.25 ⁇ g/kg, 0.5 ⁇ g/kg, 0.75 ⁇ g/kg, 1 ⁇ g/kg, 1.5 ⁇ g/kg, 2 ⁇ g/kg, 4 ⁇ g/kg, 5 ⁇ g/kg, 10 ⁇ g/kg, 15 ⁇ g/kg, 20 ⁇ g/kg, 25 ⁇ g/kg, 30 ⁇ g/kg, 35 ⁇ g/kg, 40 ⁇ g/kg.
  • a subject is administered one or more doses of a prophylactic ally or therapeutically effective amount of the topical vaccine formulation, wherein the dose is decreased by. e.g., 0.01 ⁇ g/kg, 0.02 ⁇ g/kg, 0.04 ⁇ g/kg, 0.05 ⁇ g/kg, 0.06 ⁇ g/kg, 0.08 ⁇ g/kg, 0.1 ⁇ g/kg, 0.2 ⁇ g/kg, 0.25 ⁇ g/kg, 0.5 ⁇ g/kg, 0.75 ⁇ g/kg, 1 ⁇ g/kg, 1.5 ⁇ g/kg, 2 ⁇ g/kg, 4 ⁇ g/kg, 5 ⁇ g/kg, 10 ⁇ g/kg, 15 ⁇ g/kg, 20 ⁇ g/kg, 25 ⁇ g/kg, 30 ⁇ g/kg, 35 ⁇ g/kg, 40 ⁇ g/kg, 45 ⁇ g/kg, or 50 ⁇ g/kg
  • a subject is administered one or more doses of an effective amount of the topical vaccine formulation, wherein the effective amount inhibits or reduces the level or hapten in the body or circulating antigen by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85%.
  • a subject is administered one or more doses of an effective amount the topical vaccine formulation, wherein the effective amount inhibits or reduces the severity of the disease or condition, e.g., a drug addiction, by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85%.
  • the effective amount inhibits or reduces the severity of the disease or condition, e.g., a drug addiction, by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75%
  • a subject is administered one or more doses of an effective amount of the topical vaccine formulation, wherein the effective amount inhibits or reduces nicotine or cocaine addiction by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%. at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%. or up to at least 85%.
  • the dosages of prophylactic or therapeutic agents other than the topical vaccine formulation which have been or are currently being used for the prevention, treatment and/or management of the disease or condition or a disease or condition related to it can be determined using references available to a clinician such as, e.g., the Physicians' Desk Reference (55th ed. 2001), Preferably, dosages lower than those which have been or are currently being used to prevent, treat and/or manage the disease or condition are utilized in combination with the topical vaccine formulation.
  • the topical vaccine formulation can be used as a
  • the drug-conjugates or immunogenic compositions comprising them may be administered to a mammal prior to any exposure to the drag to generate anti-drug antibodies.
  • the generated anti-drug antibodies would be present in the mammal to bind to any drug introduced subsequent to the administration of the topical vaccine formulation, and therefore minimize or prevent the chance of becoming addicted to the drug.
  • the topical vaccine formulation can be used in a subject as vaccines.
  • the topical vaccine formulations comprising at least one hapten-carrier conjugate are capable of eliciting the production of a sufficiently high titer of antibodies specific to the hapten such that upon subsequent challenge with the hapten said antibodies are capable of reducing the addictive properties of the drug.
  • the immune response to the topical vaccine formulation is the formation of both anti-hapten and anti-carrier antibodies. The therapeutic level is reached when a sufficient amount of the anti-drug specific antibodies are elicited and maintained to mount a neutralizing attack on drug introduced after vaccination.
  • the therapeutic regimens can allow for sufficient time for production of antibodies after initial vaccination and any boosting.
  • the antibody titers remain sufficiently high to provide an effective response upon subsequent exposure to the drug for about two months to about one year or more, or at least three months.
  • the topical vaccine formulation optionally includes adjuvants.
  • the topical vaccine formulation comprises a hapten-carrier conjugate wherein the hapten is nicotine or a nicotine derivative, which can be used to immunize mammals, particularly humans, to elicit anti- nicotine antibodies capable of binding free drug and preventing transit of the drug to the reward system in the brain thereby abrogating addictive drug-taking behavior (e.g., smoking cigarettes).
  • addictive drug-taking behavior e.g., smoking cigarettes.
  • nicotine binds to the alpha-subunit of the nicotinic acetylcholine receptors in the brain which results in an increase in dopamine release. It is thought that increased numbers of nicotinic acetylcholine receptors in the brain enhance the physiological dependence of nicotine.
  • anti-nicotine antibodies would presumably limit the distribution of nicotine across the blood-brain barrier to the brain, thus reducing its pharmacological effects.
  • Antibody intervention in the case of nicotine may have some advantages over cocaine. For example, there is some level of standardization with nicotine delivery; that is, each cigarette contains on average 9 mg of nicotine of which 1-3 mg are effectively dispensed during smoking. Additionally, the peak plasma concentration of nicotine is 25-50 ng/ml which is significantly lower than that of cocaine (0.3-1 ⁇ g/ml). This should provide an ideal opportunity for intervention with moderately high affinity antibodies.
  • initial vaccination with the topical vaccine formulations creates high titers of hapten-specific antibodies in vivo.
  • Periodic tests of the vaccinated subjects plasma are useful to determine individual effective doses. Titer levels are increased and maintained through periodic boosting. It is anticipated that this therapeutic will be used in combination with current drug rehabilitation programs, including counseling.
  • the therapeutic compositions of the present invention may be aimed at a single drug or several drugs simultaneously or in succession and may be used in combination with other therapies.
  • the topical vaccine formulation can be used without adverse interactions in combination with conventional pharmacological approaches and ""short term"" passive immunization to enhance the overall effect of therapy.
  • the hapten-carrier conjugate which is prepared by coupling one or more hapten molecules to a T cell epitope containing carrier to obtain a hapten-carrier conjugate capable of stimulating T cells
  • the chosen molecule is similar to those molecules which are endogenous to the individual, antibodies raised against such a molecule could cross-react with many different molecules in the body giving an undesired effect.
  • the drug to be selected as the hapten drug/haptenj must be sufficiently foreign and of a sufficient size so as to avoid eliciting antibodies to molecules commonly found inside a human body.
  • alcohol for example, would not be suitable for the therapeutic of the instant invention.
  • the antibodies raised against the therapeutic composition are highly specific and of a sufficient quantity to neutralize the drug either in the blood stream or in the mucosa or both.
  • the drugs which are suitable for therapeutic composition are, for example, hallucinogens, for example mescaline and LSD; Cannabinoids, for example THC; Stimulants, for example amphetamines, cocaine, phenmetrazine, methylphenidate; Nicotine; Depressants, for example, nonbarbiturates (e.g. bromides, chloral hydrate etc.), methaqualone.
  • kits comprising one or more containers filled with the topical vaccine formulation.
  • the kits can be used in the above-described methods.
  • the kits can be used for the prevention, treatment, and/or management of a disease or condition, e.g., a drug addiction (e.g., to cocaine or nicotine).
  • the kit comprises a topical vaccine formulation or an antibody raised in response to the formulation, in one or more containers.
  • a kit comprises a topical vaccine formulation or an antibody raised in response to the formulation, in one or more containers, and one or more other prophylactic or therapeutic agents, in one or more other containers.
  • the kit further comprises instructions for using the the topical vaccine formulation or the antibody raised in response to the formulation, as well as an explanation of side effects of the topical vaccine formulation or the antibody raised in response to the formulation, and dosage information for a particular route of administration.
  • Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration. 5.
  • topical vaccine formulations may be prepared by the following general procedure:
  • the organic phase is produced by weighing the lipid, the surfactant, and any additional lipophilic excipients into suitable containers followed by mixing these components into anoptically isotropic phase which appears as a clear solution, where the hapten-carrier conjugate is as described above, and is preferably a nicotine-carrier conjugate or a cocaine- carrier conjugate. During mixing, the organic phase will be heated up to a temperature of about 5 to about 60 °C.
  • the aqueous phase is prepared by weighing the non-lipophilic components, including the hapten-carrier conjugate, and water, which serves as solvent, into suitable containers and then mixing these components into a clear solution. During mixing, the temperature will be elevated to about 5 to about 60 °C.
  • the isotropic organic phase and the clear aqueous phase are combined under stirring in a suitable vessel. Before and during the combination the temperature of both phases must be kept between about 5 to about 60 °C or between about 35 and about 45 °C.
  • the resulting intermediate is homogenised mechanically at a temperature of about 5 to about 60 °C, e.g., about 40 °C. Before starting homogenisation, the pressure in the production vessel is lowered to - 0.08 MPa. The desired average carrier size is typically reached after 10 minutes of homogenisation.
  • Table 8 describes the amount of surfactant, lipid, hapten-carrier conjugate, and other excipients for some topical vaccine formulations provided herein.
  • the amount of the hapten-carrier conjugate, lipid, surfactant, and other excipients is described in terms of the percent total in the formulation.
  • Hapten-Carrier Conjugate 0.1 - 50 mg/g (nicotine- and cocaine-, which specific amounts besides 1 mg/g?)
  • Surfactant 1 to 10% by weight (Sphingosylphosphorylcholine, phosphatidyl choline, sphingomyelin, phosphatidyl glycerol, any others? Specific amounts?)
  • Lipid 1 to 10% by weight (T ween 80, Brij 98, Cremophor, Span 20, any others? Specific amounts?)
  • Buffer pH 3 - 9.5 (acetate, phosphate, lactate)
  • Antimicrobials 0 - 10 mg/g (benzyl alcohol + methyl parabene, each alone, others? E.g., benzalkonium chloride?)
  • Antioxidants 0 - 10 mg/g (include any here?)
  • Emollients 0 - 50 mg/g (glycerol, mannitol, others? Specific amounts besides 6.25 g/mg?)
  • the topical vaccine formulation comprises 1 mg/g TA- NIC as the hapten-carrier conjugate, 64.516 mg/g sphingosylphosphorylcholine, 35.484 mg/g Tween 80 , 6.25 mg/g mannitol, 5.25 mg/g benzyl alcohol, 2.5 mg/g methyl parabene. 3 mg/g EDT ⁇ , 1 1.25 mM phosphate buffer at pH 6.3, 30 mg/g ethanol, and 851.774 mg/g water.

Abstract

Provided herein are topical hapten-carrier conjugate formulations comprising a hapten-carrier conjugate, a lipid, and a surfactant, and uses thereof for the treatment of drug addiction.

Description

TOPICAL VACCINE FORMULATIONS AND METHODS OF TREATING DRUG
ADDICTION USING SAME
[0001] This application claims the benefit of U.S. Provisional Patent Application No. 61/221 ,812, filed June 30, 2009, herein incorporated by reference in its entirety.
1. FIELD
[0002] Provided herein are topical vaccine formulations comprising a hapten-carrier conjugate, a lipid, and a surfactant; methods of preparing same; and methods of using same for the treatment of drug abuse.
2. BACKGROUND
[0003] Hapten-carrier conjugates are currently in clinical trials as therapeutics for drug addiction. In these ongoing trials, when a composition containing the hapten-carrier conjugate, e.g.. a cocaine-bacterial toxin conjugate or a nictonic-bacterial toxin conjugate, is administered to a an addicted individual, antibodies specific to the drug of addiction arc elicited. For example, when the therapeutic composition is a cocaine-carrier conjugate, treatment induces an anti-cocaine antibody response which reduces cocaine in the bloodstream or mucosal tissue of the subject, thereby reducing the physiologically addictive effects of the cocaine. Treatment with a nicotine-carrier conjugate elicits an analogous effect towards nicotine, and reduces the individual's gratificiation from the use of nicotine.
[0004] The potential success of these clinical trials exposes a need for formulations which allow for facile administration of these hapten-carrier conjugates to enhance patient compliance.
3. SUMMARY OF THE DISCLOSURE
[0005] Provided herein are topical vaccine formulations which may be used for the treatment of, for example, drug abuse. The topical vaccine formulations provided herein comprise a hapten-carrier conjugate, a lipid, and a surfactant, optionally in a
pharmaceutically acceptable carrier. In some embodiments, the lipid is a phospholipid. In some embodiments, the surfactant is a non-ionic surfactant. The hapten-carrier conjugate in the topical vaccine formulation is capable of eliciting an immune response in the form of anti-hapten antibodies within the subject which upon subsequent exposure to the drug neutralizes the hapten and diminishes or eliminates the pharmacological effects associated therewith. In some embodiments, the hapten is a drug or drug derivative or drug metabolite, preferably a drug of addiction, for example cocaine or nicotine. In some embodiments, the topical vaccine formulation can be used as a therapeutic for treating drug addiction, for example a cocaine or nicotine addiction, by vaccination of a subject with the topical vaccine formulation comprising a hapten-carrier conjugate, a lipid, and a surfactant. In some embodiments, the hapten-carrier conjugate is a cocaine-protein or nicotine-protein conjugate.
[0006] In some embodiments, the hapten-carrier conjugate of the topical vaccine formulation is derived from at least one hapten and at least one T cell epitope-containing carrier, which conjugate is capable of stimulating the production of anti-hapten antibodies. In some embodiments, the carrier is a bacterial toxin, for example cholera toxin B.
[0007] In some embodiments, the topical vaccine formulation comprises a therapeutically effective amount of hapten-carrier conjugate, for example from about 0.1 to about 50 mg of the hapten-carrier conjugate per g of the formulation. In some embodiments, the topical vaccine formulation comprises about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 8 mg, about 10 mg, about 12 mg, about 14 mg, about 16 mg, about 18 mg, or about 20 mg of the hapten-carrier conjugate per g of the formulation.
[0008] In some embodiments, the topical vaccine formulation is a patch, for example, for transdermal delivery or transcutaneous delivery.
[0009] Also provided herein are methods of treating, for example, drug abuse, comprising topically administering to a subject a topical vaccine formulation comprising a
therapeutically effective amount of hapten-carrier conjugate, a lipid, and a surfactant. In some embodiments of the method, the hapten-carrier conjugate elicits an immune anti-hapten antibodies within the subject which upon subsequent exposure to the drug neutralizes the drug and diminishes or eliminates the pharmacological effects associated therewith. In some embodiments of the method, the hapten-carrier conjugate is derived from a hapten that is a drug or drug derivative or drug metabolite, preferably a drug of addiction, for example cocaine or nicotine.
[0010] In some embodiments of the method, the topical vaccine formulation is
administered regularly, for example, once or twice daily. In some embodiments, the topical vaccine formulation is administered for at least three weeks. [0011] In some embodiments of the method, the topical vaccine formulation comprises from about 0.1 to about 50 mg of the hapten-carrier conjugate. In some embodiments of the method, the composition comprises about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 8 mg, about 10 mg, about 12 mg, about 14 mg, about 16 mg, about 18 mg, or about 20 mg of the hapten-carrier conjugate per g of the formulation.
[0012] In some embodiments, the topical vaccine formulations provided herein are useful for the application, administration and/or transport of the hapten-carrier conjugate into and through barriers and constrictions, such as the skin of a subject (e.g., humans).
[0013] In some embodiments, the topical vaccine formulations pro\ ided herein form vesicles or other extended surface aggregates (ESAs), wherein the vesicular preparations have improved permeation capability through the semi-permeable barriers, such as skin. While not to be limited to any mechanism of action, the topical vaccine formulation provided herein are able to form vesicles characterized by their deformability and/or adaptability. The vesicles' deformability and/or adaptability allow the vesicles to penetrate the pores of the skin and deliver the hapten-carrier conjugate to the subject in an amount sufficient to elicit an immune response. The adaptability or deformability of the vesicles may be determined by the ability of the vesicles to penetrate a barrier with pores having an average pore diameter at least 50% smaller than the average vesicle diameter before the penetration.
[0014] In some embodiments, the topical vaccine formulations provided herein are used to deliver the hapten-carrier conjugate non-invasively. The topical vaccine formulations provided herein may be administered in place of other vaccine formulations, including intravenous vaccines.
4. DETAILED DESCRIPTION OF THE DISCLOSURE
[0015] To facilitate understanding of the disclosure set forth herein, a number of terms are defined below.
[0016] Generally, the nomenclature used herein and the laboratory procedures in organic chemistry, medicinal chemistry, and pharmacology described herein are those well known and commonly employed in the art. Unless defined otherwise, all technical and scientific terms used herein generally have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. [0017] The term '"'subject"" refers to an animal, including, but not limited to, a primate (e.g. , human), cow, sheep, goat, pig, horse, dog, cat, rabbit, rat, or mouse. The terms '"'subject'"' and '"'patient'"' are used interchangeably herein in reference, for example, to a mammalian subject, such as a human subject.
[0018] The term '"'treat,"" ""treating,"" or '"'treatment of" means that the severity of a subject's condition is reduced or at least partially improved or ameliorated and/or that some alleviation, mitigation or decrease in at least one clinical symptom is achieved and/or there is an inhibition or delay in the progression of the condition and/or delay in the progression of the onset of disease or illness. The term ""treat,"" '"'treating,'-" or ""treatment of" also means managing the disease state.
[0019] The term ""pharmaceutically acceptable"" when used in reference to the formulations provided herein denotes that a formulation does not result in an unacceptable level of irritation in the subject to whom the formulation is administered. Preferably such level will be sufficiently low to provide a formulation suitable for approval by regulatory authorities.
[0020] The term '""sufficient amount,"" ""amount effective to,"" or an ""amount sufficient to"" achieve a particular result refers to an amount of a hapten-carrier conjugate that is effective to produce a desired effect, which is optionally a therapeutic effect (i.e., by administration of a therapeutically effective amount). Alternatively stated, a
""therapeutically effective"" amount is an amount that provides some alleviation, mitigation, and/or decrease in at least one clinical symptom. Clinical symptoms associated with the disorder that can be treated by the methods provided herein are well-known to those skilled in the art. Further, those skilled in the art will appreciate that the therapeutic effects need not be complete or curative, as long as some benefit is provided to the subject. For example, a ""sufficient amount'"' or ""an amount sufficient to"" can be an amount that is effective to treat a disease.
[0021] As used herein with respect to numerical values, the term ""about"" means a range surrounding a particular numeral value which includes that which would be expected to result from normal experimental error in making a measurement. For example, in certain embodiments, the term ""about"" when used in connection with a particular numerical value means ±1%, ±2%, ±3%, ±4%, ±5%, ±10%, ±15%, or ±20% of the numerical value. [0022] The term ""alkyl"" refers to a linear or branched saturated monovalent hydrocarbon radical, wherein the alkyl may optionally be substituted with one or more substituents Q as described herein. The term ""alkyl"" also encompasses both linear and branched alkyl, unless otherwise specified. In certain embodiments, the alkyl is a linear saturated monovalent hydrocarbon radical that has 1 to 20 (Ci-20), 1 to 15 (C 1-15), 1 to 12 (Cj- 12), 1 to 10 (Ci-10), or 1 to 6 (Ci-6) carbon atoms, or a branched saturated monovalent hydrocarbon radical of 3 to 20 (C3.20), 3 to 15 (Cs-is), 3 to 12 (C3.12), 3 Io 10 (C3-10), or 3 to 6 (Cj-6) carbon atoms. As used herein, linear C]-6 and branched C3-6 alkyl groups are also referred as '"lower alkyl."" Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl (including all isomeric forms), n-propyl, isopropyl, butyl (including all isomeric forms), n-butyl, isobutyl, sec-butyl, t-butyl, pentyl (including all isomeric forms), and hexyl (including all isomeric forms). For example,
Figure imgf000006_0001
alkyl refers to a linear saturated
monovalent hydrocarbon radical of 1 to 6 carbon atoms or a branched saturated monovalent hydrocarbon radical of 3 to 6 carbon atoms.
[0023] The term '"'aryP'" refers to a monocyclic aromatic group and/or multicyclic monovalent aromatic group that contain at least one aromatic hydrocarbon ring. In certain embodiments, the ary] has from 6 to 20 (C6-20X from 6 to 15 (€5-15), or from 6 to 10 (CVi 0) ring atoms. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, fluorenyl, azulenyl, anthryl, phenanthryl, pyrenyl, biphenyl, and terphenyl. Aryl also refers to bicyclic or tricyclic carbon rings, where one of the rings is aromatic and the others of which may be saturated, partially unsaturated, or aromatic, for example, dihydronaphthyl, indenyl, indanyl. or tetrahydronaphthyl (tetralinyl). In certain embodiments, aryl may also be optionally substituted with one or more substituents Q as described herein.
[0024] The term ""heteroaryl"" refers to a monocyclic aromatic group and/or multicyclic aromatic group that contain at least one aromatic ring, wherein at least one aromatic ring contains one or more heteroatoms independently selected from O, S, and N. Each ring of a heteroaryl group can contain one or two O atoms, one or two S atoms, and/or one to four N atoms, provided that the total number of heteroatoms in each ring is four or less and each ring contains at least one carbon atom. The heteroaryl may be attached to the main structure at any heteroatom or carbon atom which results in the creation of a stable compound. In certain embodiments, the heteroaryl has from 5 to 20, from 5 to 15, or from 5 to 10 ring atoms.
Examples of monocyclic heteroaryl groups include, but are not limited to, pyrrolyl, pyrazolyl, pyrazolinyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, thiadiazolyl, isothiazolyl, furanyl, thienyl, oxadiazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, and triazinyl. Examples of bicyclic heteroaryl groups include, but are not limited to, indolyl, benzothiazolyl, benzoxazolyl, benzothienyl, quinolinyl, tetrahydroisoquinolinyl, isoquinolinyl,
benzimidazolyl, benzopyranyl, indolizinyl, benzofuranyl, isobenzofuranyl, chromonyl, coumarinyl. cinnolinyl, quinoxalinyl, indazolyl, purinyl, pyrrolopyridinyl, furopyridinyl, thienopyridinyl, dihydroisoindolyl, and telrahydroquinolinyl. Examples of tricyclic heteroaryl groups include, but are not limited to, carbazoiyl, benzindolyl, phenanthrollinyl, acridinyl, phenanthridinyl, and xanthenyl. In certain embodiments, heteroaryl may also be optionally substituted with one or more substituents Z as described herein.
[0025] The term '""alkenoyl"" as used herein refers to ~C(O)-alkenyl. The term ■""alkenyl"" refers to a linear or branched monovalent hydrocarbon radical, which contains one or more, in one embodiment, one to five, carbon-carbon double bonds. The alkenyl may be optionally substituted with one or more substituents Z as described herein. The term ""alkenyl"'" also embraces radicals having ""cis"" and ""/raw"" configurations, or alternatively, '""Z'"' and "'"E"-' configurations, as appreciated by those of ordinary skill in the art. As used herein, the term ""alkenyl'"' encompasses both linear and branched alkenyl, unless otherwise specified. For example, C2-6 alkenyl refers to a linear unsaturated monovalent hydrocarbon radical of 2 to 6 carbon atoms or a branched unsaturated
monovalent hydrocarbon radical of 3 to 6 carbon atoms. In certain embodiments, the alkenyl is a linear monovalent hydrocarbon radical of 2 to 30 (C2-3o), 2 to 24 (C2-2O, 2 to 20 (€2-20), 2 to 15 (C2-15), 2 to 12 (C2-I2), 2 to 10 (C2-] 0), or 2 to 6 (C2-6) carbon atoms, or a branched monovalent hydrocarbon radical of 3 to 30 (C3-3O), 3 to 24 (C3-24), 3 to 20 (C3-2o), 3 to 15 (C3. 15), 3 to 12 (C3-I2), 3 to 10 (C3-Io), or 3 to 6 (C3-6) carbon atoms. Examples of alkenyl groups include, but are not limited to, ethenyl, propen-1-yl, propcn-2-yl, allyl, butenyl, and 4- methylbutenyl. In certain embodiments, the alkenoyl is mono-alkenoyl, which contains one carbon-carbon double bond. In certain embodiments, the alkenoyl is di-alkenoyl, which contains two carbon-carbon double bonds. In certain embodiments, the alkenoyl is poly- alkenoyl, which contains more than two carbon-carbon double bonds.
[0026] The term ""heterocyclyl"" or ""heterocyclic'""' refers to a monocyclic non- aromatic ring system and/or multicyclic ring system that contains at least one non-aromatic ring, wherein one or more of the non-aromatic ring atoms are heteroatoms independently selected from O, S, or N; and the remaining ring atoms are carbon atoms. In certain embodiments, the heterocyclyl or heterocyclic group has from 3 to 20, from 3 to 15, from 3 to 10, from 3 to 8, from 4 to 7, or from 5 to 6 ring atoms. In certain embodiments, the heterocyclyl is a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which may include a fused or bridged ring system, and in which the nitrogen or sulfur atoms may be optionally oxidized, the nitrogen atoms may be optionally quaternized, and some rings may be partially or fully saturated, or aromatic. The heterocyclyl may be attached to the main structure at any heteroatom or carbon atom which results in the creation of a stable compound. Examples of such heterocyclic radicals include, but are not limited to, acridinyl, azepinyl, benzimidazolyl, benzindolyl, benzoisoxazolyl, benzisoxazinyl, benzodioxanyl, benzodioxolyl, benzofuranonyl, benzofuranyl, benzonaphthofuranyl, benzopyranonyl, benzopyranyl, benzotetrahydrofuranyl, benzotetrahydrothienyl, benzothiadiazolyl.
benzothiazolyl, benzothiophenyl, benzotriazolyl, benzothiopyranyl, benzoxazinyl, benzoxazolyl, benzothiazolyl, β-carbolinyl, carbazolyl, chromanyl, chromonyl, cinnolinyl, coumarinyl, decahydroisoquinolinyl, dibenzofuranyl, dihydrobenzisothiazinyl,
dihydrobenzisoxazinyl, dihydrofuryl, dihydropyranyl, dioxolanyl, dihydropyrazinyl, dihydropyridinyl, dihydropyrazolyl, dihydropyrimidinyl, dihydropyrrolyl, dioxolanyl, 1,4- dithianyl, furanonyl, furanyl, imidazolidinyl, imidazolinyl, imidazolyl, imidazopyridinyl, imidazothiazolyl, indazolyl, indolinyl, indolizinyl, indolyl, isobenzotetrahydrofuranyl, isobenzotetrahydrothienyl, isobenzothienyl, isochromanyl, isocoumarinyl, isoindolinyl, isoindolyl, isoquinolinyl, isothiazolidinyl, isothiazolyl, isoxazolidinyl, isoxazolyl, morpholinyl. naphthyridinyl, octahydroindolyl, octahydroisoindolyl, oxadiazolyl, oxazolidinonyl, oxazolidinyl, oxazolopyridinyl, oxazolyl, oxiranyl, perimidinyl,
phenanthridinyl, phenathrolinyl, phenarsazinyl, phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, piperazinyl, piperidinyl, 4-piperidonyl, pteridinyl, purinyl, pyra/inyl, pyrazolidinyl, pyrazolyl, pyridazinyl, pyridinyl, pyridopyridinyl, pyrimidinyl, pyrrolidinyl, pyrrolinyl, pyrrolyl, quinazolinyl, quinolinyl, quinoxalinyl, quinuclidinyl, tctrahydrofuryl, tetrahydrofuranyl, tetrahydroisoquinolinyl, tetrahydropyranyl, tetrahydrothienyl, tetrazolyl, thiadiazolopyrimidinyl, thiadiazolyl, thiamorpholinyl, thiazolidinyl, thiazolyl, thienyl, triazinyl, triazolyl, and 1,3,5-trithianyl. In certain embodiments, heterocyclic may also be optionally substituted with one or more substituents Z as described herein.
[0027] The term ""halogen"", ""halide"" or -'"halo"" refers to fluorine, chlorine, bromine, and/or iodine.
[0028] The term ""Optionally substituted"" is intended to mean that a group, including alkyl, alkenyl, alkynyl, cycloalkyl, aryl, aralkyl, heteroaryl, and heterocyclyl, may be substituted with one or more substituents Z, in one embodiment, one, two, three or four substituents Z, where each Z is independently selected from the group consisting of cyano, halo, oxo, nitro, C1-6 alkyl, halo- C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 cycloalkyl, C6-^ aryl, C7-I4 aralkyl, heteroaryl, heterocyclyl, -C(O)R6, -C(O)OR6, -C(O)NRfRg, - C(NRe)NRfRg, -ORC, -OC(O)R0, -OC(O)OR6, -OC(O)NRfRg, -0C(=NRe)NRfRg, - OS(O)RC, -OS(O)2R6, -OS(O)NRfRg, -OS(O)2NRfRs, -NRfR8, -NReC(O)Rf, -NReC(0)0Rf, -NReC(0)NRfRg, -NReC(=NRh)NRrRg, -NReS(0)Rf, -NReS(0)2Rf, -NR6S(0)NRfRg, - NReS(0)2NRfRg, -SRe, -S(O)Re, -S(O)2R6, and -S(O)2NRfRg, wherein each Re, Rf, Rg, and R is independently hydrogen, Ci.6 alkyl, C2-6 alkenyl, C2_g alkynyl, C3-7 cycloalkyl, C6-I4 aryl. C7- I 4 aralkyl, heteroaryl, or heterocyclyl; or R and R8 together with the N atom to which they are attached form heterocyclyl.
[0029] The terms ""optically active"" and ""enantiomerically active"" refer to a collection of molecules, which has an enantiomeric excess of no less than about 50%, no less than about 70%, no less than about 80%, no less than about 90%, no less than about 91%, no less than about 92%, no less than about 93%, no less than about 94%, no less than about 95%, no less than about 96%, no less than about 97%, no less than about 98%, no less than about 99%, no less than about 99.5%, or no less than about 99.8%.
[0030] In describing an optically active compound, the prefixes R and S are used to denote the absolute configuration of the molecule about its chiral centcr(s). The (+) and (-) are used to denote the optical rotation of the compound, that is, the direction in which a plane of polarized light is rotated by the optically active compound. The (-) prefix indicates that the compound is levorotatory, that is, the compound rotates the plane of polarized light to the left or counterclockwise. The (+) prefix indicates that the compound is dextrorotatory, that is, the compound rotates the plane of polarized light to the right or clockwise. However, the sign of optical rotation, (+) and (-), is not related to the absolute configuration of the molecule, R and S.
[0031] The term ""solvate"" refers to a compound provided herein or a salt thereof, which further includes a stoichiometric or non-stoichiometric amount of solvent bound by non-covalent intermolecular forces. Where the solvent is water, the solvate is a hydrate.
[0032] The formulations provided herein comprise a hapten-carrier conjugate, a lipid, preferably a phospholipid, a surfactant, preferably a nonionic surfactant, and an aqueous solution, having a pH ranging from 3.5 to 9.0, preferably from 4 to 7.5. The formulations may optionally contain buffers, antioxidants, preservatives, microbicides. antimicrobials, emollients, co-solvents, and/or thickeners.
[0033] While not to be limited by any mechanism of action, the formulations provided herein form vesicles or other extended surface aggregates (ESAs), wherein the vesicular preparations have improved permeation capability through the semi-permeable barriers, such as skin and/or nails. The vesicles or extended surface aggregates provided herein comprise of a hapten carrier conjugate, a lipid, and one or more membrane destabilizing agents, such as surfactants.
4.1. HAPTEN-CARRIER CONJUGATE
[0034] The hapten-carrier conjugates for use in the topical vaccine formulations of the disclosure can be any hapten-carrier conjugate that elicits an appropriate immune response to the free hapten. In some embodiments, the hapten-carrier conjugates are those described in U.S. Patent Application Publication No. 2005/0124061 (published June 9, 2005), herein incorporated by reference in its entirety.
4.1.1 Carriers for use in hapten-carrier conjugates
[0035] In some embodiments, the carrier is a bacterial toxin is from a gram-negative bacteria. In some embodiments, the carrier is a bacterial toxin is from a gram-positive bacteria. In some embodiments, the gram-negative bacteria is Escherichia coli. In some embodiments, the bacteria is the gram-positive bacillus, staphylococcus, streptococcus, streptomyces, or mollicutes (mycoplasma). In some embodiments, the carrier is a bacterial exotoxin. In some embodiments, the carrier is a bacterial toxin that has been modified to reduce its toxicity. Bacterial toxins for use in accordance with the disclosure include, but are not limited to, cholera toxin, preferably CTB (including recombinant CTB (rCTB)),
Escherichia coli toxins such as heat-labile enterotoxin (LT), heat-stable exotoxin (ST), cytotoxic necrotizing factor (CNF), cytolethal distending toxin (CLDT), or enteroaggregative E. coli heat-stable toxin (EAST), diphtheria toxin (Dtx), tetanus toxin, shiga toxins, botulinum toxin, staphylococci toxins, such as Staphylococcus aureus alpha toxin, Exfoliatin B or leukocidin, staphylococcal toxic shock syndrome toxin (TSST-I), staphylococcus enterotoxins or exfoliative toxins, streptococci toxins, such as pneumolysin of Streptococcus pneumoniae, streptolysin O, Erythrogenic toxin (streptococcal pyrogenic exotoxin (SPE)), and other pyrogenic toxins of Streptococcus pyogenes, clostridial toxins, such as toxin A/toxin B of Clostridium difficile, Iota family, C2 family (toxins C and D), or C3 toxins, or neurotoxins A-G of Clostridium bolulinum, alpha toxin, beta-2 toxin, or perfringiolysin O of Clostridium per fringens (Perfringens enterotoxin), enterotoxin of Barter oides fragilis, Aeromonas hydrophila/mrolysin, filamentous hemagglutinin (FHA) of Bordatella pertussis, Clostridium or Bacillus binary toxins, streptokinase, the adenylate cyclase toxin of Bordatella pertussis (pertussis AC; ""'pertussis toxin"") or its dermonecrotic toxin, Bacillus anthracis edema factor (EF), anthrax toxin (lethal factor (LF)), hemolysin of Escherichia coli, listeriolysin of Listeria monocytogenes, and Pseudomonas exotoxin (exotoxin A). In some embodiments, a bacterial ADP-ribosylating exotoxin is preferably used, such as, for example, cholera toxin, diphtheria toxin, pertussis toxin, Pseudomonas exotoxin A, or E. coli LT. In some embodiments, the catalytic subunit (usually, the '"-A"" subunit) of the bacterial ADP- ribosylating exotoxin is used as carrier. In other embodiments, the receptor-binding subunit of the bacterial ADP-ribosylating exotoxin (usually, the ""B"" subunit) is preferred. In yet other embodiments, both subunits or a fragment or fragments thereof are used as a carrier. In some embodiments, a bacterial pore-forming toxin is used, such as perfringiolysin O, hemolysin, listeriolysin, anthrax EF, alpha toxin, pneumolysin, streptolysin O, or leukocidin. In some embodiments, the carrier is a pyrogεnic exotoxin, or a modified form thereof, such as staphylococcal enterotoxins serotypes A-E, G, and H, group A streptococcal pyrogenic exotoxins A-C, staphylococcal exfoliatin toxin, and staphylococcal TSST-I . Particularly useful bacterial toxin carriers include any bacterial toxin with the ability to enhance a mucosal immune response, for example, CTB or any toxin in the E. coli heat-labile enteroloxin (LTB) family of bacterial toxins.
[0036] In some embodiments, the bacterial toxin carrier of the instant invention contains at least one T cell epitope which is capable of stimulating the T cells of the subject, which in turn help the B cells initiate and maintain sustained antibody production to portions of the entire conjugate, including the hapten portion. Thus, since a carrier is selected because it is immunogenic, a strong immune response to the vaccine in a diverse patient population is expected. In preferred embodiments, the carrier, like the hapten, must be sufficiently foreign to elicit a strong immune response to the vaccine. A conservative, but not essential, approach is to use a carrier to which most patients have not been exposed to avoid the phenomenon of carrier-induced epitope suppression. However, even if carrier-induced epitope suppression does occur, it is manageable as it has been overcome by dose changes (DiJohn et al. (1989) Lancet 1415-1418) and other protocol changes (Etlinger et al. (1990) Science 249:423-425), including the use of CTB (Stok et al. (1994) Vaccine 12:521-526). Vaccines which utilize carrier proteins to which patients are already immune are commercially available. Still further, carriers containing a large number of lysines are particularly suitable for conjugation according to the methods of the instant invention. In certain embodiments, therefore, the bacterial toxin carriers of the invention are modified so that their immunogenic properties are enhanced.
4.1.1.1 CTB as a carrier
[0037] Cholera toxin is the enterotoxin produced by Vibrio cholerae and consists of five identical B subunits with each subunit having a molecular weight of 1 1.6 KDa (103 amino acids) and one A subunit of 27.2 KDa (230 amino acids) (Finkelstein (1988) Immunochem. MoI. Gen. Anal. Bac. Path. 85-102). The binding subunit, CTB, binds to ganglioside GMl on the cell surface (Sixma et al. (1991) Nature 351 :371-375; Orlandi et al. (1993) J. Biol. Chem. 268: 17038-17044). CTA is the enzymatic subunit which enters the cell and catalyzes ADP- ribosylation of a G protein, constitutively activating adenylate cyclase (Finkelstein (1988) Immunochem, MoI. Gen. Anal. Bac. Path. pp. 85-102). In the absence of the A subunit, cholera toxin is not toxic.
[0038] In preferred embodiments, CTB is the bacterial toxin carrier. CTB is a highly immunogenic protein subunit capable of stimulating strong systemic and mucosal antibody responses (Lycke (1992) J. Immunol. 150:4810-4821 ; Holmgren et al. (1994) Am. J. Trop. Med. Ilyg. 50:42-54; Silbart et al. (1988) J. Immun. Meth. 109:103-1 12; Katz et al. (1993) Infection Immun. 61 : 1964-1971). This combined IgA and IgG anti-hapten response is highly desirable in blocking, for example, cocaine or other substances for which is immunity is desired that are administered nasally or by inhalation, and in blocking nicotine or other substances that are absorbed in the mouth and lungs. In addition, CTB has already been shown to be safe for human use in clinical trials for cholera vaccines (Holmgren et al., supra; Jertborn et al. (1994) Vaccine 32: 1078-1082; '"'The Jordan Report, Accelerated Development of Vaccines"" 1993., NIAID, 1993). It is a discovery of this invention that hapten-carrier conjugates comprising CTB have even greater immunogenicity when CTB contains a signal peptide..
4,1.1.2 CTB preparation
[0039] Methods of making and using CTB as a carrier for incorporation into toxin-carrier conjugates are known. For example, see, U.S. Patent Application Publication No. 2005- 0124061, published June 9, 2005; U.S. Patent No. 5,876,727, issued March 2, 1999; and U.S. Patent No. 5,760,184; each of which is incorporated herein by reference in its entirety. CTBs generated according to these methods, and the methods presented herein, may be modified so that it contains a signal peptide as described herein.
[0040] In a preferred embodiment, CTB is produced in Vibrio cholerae strain 213. See, for example, International Patent Application Publication No. WO 2005/042749, published May 12. 2005, which is incorporated by reference herein in its entirety.
[0041] In other embodiments, CTB is produced in E, coli (see, e.g., U.S. Patent
Application Publication No. 2005-0124061, incorporated herein by reference in its entirety).
[0042] Amounts of recombinant CTB have been expressed and purified amounts which, once optimized, are produced in large fermentation batches. Processes for expressing and purifying recombinant protein are known in the art, for example, U.S. Patent Application Ser. No. 07/807,529. For example, CTB may be purified by affinity chromatography (Tayot et al. (1981) Eur. J. Biochem. 1 13:249-258), conjugated to cocaine or nicotine derivatives, and the conjugate may then be further purified. The purified CTB and the resulting conjugate are analyzed for purity and for maintenance of the pentameric structure of CTB. Techniques include SDS-PAGE, native PAGE, gel filtration chromatography, Western blotting, direct and GMl -capture ELISA, and competition ELISA with biotinylated CTB. Level of haptenation is measured by mass spectrometry, reverse phase HPLC and by analysis of the increase in UV absorbance resulting from the presence of the hapten. Both the solubility and the stability of the conjugate are optimized in preparation for full-scale formulation. Details of some of these analyses are given in the Examples.
[0043] Although the pentameric structure of CTB is a preferred carrier for practice of the present invention, and GMl binding is an effective assay to determine that the pentameric form of CTB is present, the present invention is not limited to the use of the pentameric form of CTB. Other forms of CTB are contemplated (e.g., monomer, dimer, etc.) that may be manipulated for use in the invention. If a carrier other than the pentameric form of CTB is utilized, then one skilled in the art would use an appropriate assay to determine the presence and activity of the required carrier, e.g., the use of GMl binding to determine the presence of the pentameric form of CTB).
[0044] Another useful CTB for use as a carrier is cholera toxin which provides improved mucosal response over CTB. It has been reported that the enzymatieally active A subunit adjuvant enhances activity (Liang et al. (1988) J. Immunol. 141 :1495-1501; Wilson et al. (1993) Vaccine 1 1 : 113-118; Snider et al. (1994) 1 Immunol. 153:647).
4.1.2 Haptens for use in hapten-carrier conjugates
[0045] A ""hapten"" for the purposes of this disclosure is a low-molecular- weight organic compound that reacts specifically with an antibody and which is incapable of inciting an immune response by itself but is immunogenic when complexed to a T cell epitope- containing carrier (i.e., as a hapten-carrier conjugate). In some embodiments, the hapten is poorly immunogenic by itself. Further, the hapten is characterized as the specificity- determining portion of the hapten-carrier conjugate, that is, it is capable of reacting with an antibody specific to the hapten in its free state. In some embodiments, in a non-immunized subject, there is an absence of formation of antibodies to the hapten. In some embodiments, in a non-immunized subject, there may be a low level of antibodies to the hapten, or a level of antibodies to the hapten for which an increase in the immune response to the hapten is desired. In some embodiments, the hapten is a drug, an analog of a portion of the drug, or a drug derivative. In some embodiments, the hapten is an addictive drug, analog of a portion of an addictive drug, or addictive drug derivative. In some embodiments, the addictive drug is nicotine or cocaine.
[0046] The immunogenic composition, or, in some embodiments, the vaccine, when initially administered will give rise to a ""desired measurable outcome."" Initially, the desired measurable outcome is the production of a high titer of anti-hapten antibodies (approximately 0.1 mg/ml to 1 mg/ml or greater of specific antibody in the serum). However, manipulation of the dosage regimen suitable for the individual gives and maintains a sustained desired therapeutic effect. The ""desired therapeutic effect"" is the neutralization of a sufficient fraction of free hapten to reduce or eliminate the pharmacological effects of the hapten (e.g., nicotine or cocaine) within a therapeutically acceptable time frame by anti- hapten antibodies specific for the hapten upon a subsequent exposure to the hapten.
Determining the therapeutically acceptable time frames for how long it takes to get a sufficient antibody response to a given hapten and how-long is that antibody response is maintained thereto are achieved by those skilled in the art by assessing the characteristics of the subject to be immunized, hapten (e.g., drug of abuse) to be neutralized, as well as the mode of administration. Using this and other immunization protocols as a model, one skilled in that art would expect the immunity or the period of protection to last several months, up to more than one year. [0047] In some embodiments, the hapten- carrier conjugate can be prepared by modifying the hapten sufficiently to render it capable of being conjugated or joined to a carrier while maintaining enough of the structure so that it is recognized as free state hapten (for example, as free cocaine or nicotine). It is preferable that a vaccinated individual has antibodies which recognize free hapten (e g., cocaine or nicotine). Radioimmunoassay and competition ELISA assay experiments can measure antibody titers to free hapten. Antibodies of interest are hapten- specific antibodies and, for example, cocaine-specific antibodies or nicotine- specific antibodies. It will be understood to one of ordinary skill in the art that the principles and methods used described herein may be extended from this disclosure to a wide range of hapten-carrier conjugates useful in the treatment of a variety of diseases, conditions, or drug addictions and toxic responses.
[0048] In some embodiments, the hapten is a drug, such as, for example, hallucinogens, for example mescaline and LSD; Cannabinoids, for example THC; Stimulants, for example amphetamines, cocaine, phenmetrazine, methylphenidate; Nicotine; Depressants, for example, nonbarbiturates (e.g. bromides, chloral hydrate etc.), methaqualonc, barbiturates, diazepam, flurazepam, phencyclidine, and fluoxetine; Opium and its derivatives, for example, heroin, methadone, morphine, meperidine, codeine, pentazocine, and propoxyphene; and '"'Designer drugs""' such as ""'ecstasy.
4.1.3 Preparation of hapten-carrier conjugates
[0049] In some embodiments, hapten-carrier conjugates for use in the topical vaccine formulations can be found in U.S. Patent Application Publication No. 2005/0124061
(published June 9, 2005), herein incorporated by reference in its entirety. It will be appreciated by one of ordinary skill in the art that the methods described in U.S. Patent Application Publication No. 2005/0124061 are readily applicable to a variety of haptens and a variety of carriers. By way of example, some of the methods are discussed hereinbelow.
[0050] Haptens are generally conjugated to a carrier by means of a chemical linker, the length and nature of which is such that the hapten is displaced a sufficient distance from the carrier domain to allow its optimal recognition by the antibodies initially raised against it. The length of the linker can be varied by, for example, modifying the number Of-CH2 groups in a given linker. The following is a non-exhaustive list of linkers usable in the hapten- carrier conjugates described herein:
CJ O Q CJ l (CH2)nQ
CJ 1.1 CO2Q
CJ 1.2 COQ
CJ 1.3 OCH3
CJ 2 OCO(CH2)nQ
CJ 2.1 OCOCH-Q
CJ 2.2 OCOCH(O)CH2
CJ 2.3 OCO(CHi)nCH(O)CH2
CJ 3 C0(CH2)nC0Q
CJ 3.1 CO(CI I2)nCNQ
CJ 4 0C0(CH2)nC0Q
CJ 4.1 OCO(CH2)nCNQ
CJ 5 CH2OCO(CH2)nCOQ
CJ 5.1 CH2OCO(CH2)nCNQ
CJ 6 CONI 1(CI I2)nQ
CJ 7 Y(CH2)nQ
CJ 7.1 CH2Y(CH2)nQ
CJ 8 OCOCH(OH)CH2Q
CJ 8.1 OCO(CHz)nCH(OH)CH2Q
CJ 9 OCOC6H5
CJ 10
Figure imgf000016_0001
, wherein Q' is a modified
protein; and
CJ 11 YCO(CH2X1COQ,
where n is an integer from 1 to about 20, preferable from about 3 to about 6; Y is selected S, O, or Nil; and Q is the carrier.
[0051] It will be understood to one of ordinary skill in the art that the chemical and/or conceptual precursor to the hapten carrier conjugate will be an analog of the hapten having one of the above-mentioned linkers connected thereto, wherein Q is H, OH, CH3, OCH3, COOH, a halogen, an activated ester or esters (such as 2-nitro-4-sulfophenyl ester or N- oxysuccinimidyl ester), a group or groups reactive toward the carrier (such as a mixed anhydride, acyl halide, acyl azide, alkyl halide, N-maleimide, imino ester, isocyanate, and isothiocyanate), or another linker identified by its ""CJ'"' reference number.
[0052] A T cell epitope containing carrier may be modified by methods known to those skilled in the art to facilitate conjugation to the hapten (e.g., by thiolation), for example with 2-iminothiolane (Traut's reagent) or by succinylation, etc.
[0053] Abbreviations of commercially obtainable compounds used herein include:
"-'BSA"" is bovine serum albumin; ""DC"" is dicyclohexylcarbodiimide; ""DMF"" is N9N- dimethylformamide; ""EDC"" or ""EDAC'"1 is N-ethyl-N'-(3-(dimethylamino) propyl) carbodiimide hydrochloride; ""EDTA"" is ethylenediamine tetraacetic acid, disodium salt; ""HATU"" is O-(7-azabenzotriazol-1-yl)-l ,l ,3,3-tetramethyluronium hexafluorophosphate; ""NMM"" is N-methylmorpholine; ""HBTU"" is 2-(1H-benzotriazole-1-yl)-l, 1,3,3- tetramethyluronium hexafluorophosphate; ""TNTU"" is 2-(5-norbornene-2,3- dicarboximido)-l,l ,3,3-tetramethyluronium tetrafluoroborate; ""PyBroP®"" is bromo-tris- pyrrolidino-phosphonium hexafluorophosphate; and ""HOBt"" is N-hydroxybenzotriazole.
[0054] IUPAC nomenclature for several named compounds are: 3β-(benzoyloxy)-8- azabicyclo[3.2.1]octane-2β-carboxylic acid methyl ester for ""norcocaine;"" 3β- (benzoyloxy)-8-methyl-8-azabicyclo[3.2.1]octane-2β -carboxylic acid for ""benzoyl ecognine;"" 3β-(benzoyloxy)-8-methyl-8-azabicyclo[3.2.1]octane-2β-carboxylic acid methyl ester for ""cocaine;"" 3β-(hydroxy)-8-methyl-8-azabicyclo[3.2.1]octane-2β-carboxylic acid methyl ester for ""ecgonine methyl ester;"" 1 -methyl-2-(2-pyridyl)pyrrolidine for
""nicotine;"" and N-methyl-2-(3-pyridyl)-5-pyrrolidmone for ""cotinine.""
[0055] In some embodiments, the hapten-carrier conjugate is a conjugate of nicotine or nicotine metabolites. In some embodiments, chemical precursors to a nicotine carrier conjugate can be synthesized by selectively alkylating the pyridine nitrogen of (S)-(-)- nicotine in anhydrous methanol, with ethyl 3-bromobutyrate, 5-bromovaleric acid, 6- bromohexanoic acid or 8-bromooctanoic acid. The resultant chemical precursor can then be conjugated to a carrier protein usin, for example, HATU.
[0056] In some embodiments, succinylated nornicotine is the chemical precursor to the conjugate.
4.2. LIPID
[0057] In the sense of this disclosure, a ""lipid"" is any substance, which has properties like or similar to those of a fat. As a rule, it has an extended apolar group (the ""chain"", X) and generally also a water-soluble, polar hydrophilic part, the ""head"" group (Y) and has the basic Formula II:
X-Yn (I) wherein n is equal to or larger than zero.
[0058] Lipids with n=0 are referred to as apolar lipids and lipids with n>l are referred to as polar lipids. In this sense, all amphfphilic substances, including, but not limited to glycerides, glycerophospholipids, glycerophosphinolipids, glycerophosphonolipids, sulfolipids, sphingolipids, isoprenoid lipids, steroids or sterols and carbohydrate-containing lipids can generally be referred to as lipids, and are included as such in this disclosure. A list of relevant lipids and lipid related definitions is provided in EP 0 475 160 Al (see, e.g. p. 4, 1. 8 to p. 6. 1. 3) and U.S. Patent No. 6,165,500 (see. e.g., col. 6, 1. 10 to col. 7. 1. 58), each incorporated herein by reference in their entirety.
[0059] A phospholipid is, for example, a compound of Formula II:
R1— CH2- CHR2- CR3H- O— PHO2-O-R4 (II) wherein R1 and R2 cannot both be hydrogen, OH or a Cj-C3 alkyl group, and typically are independently, an aliphatic chain, most often derived from a fatty acid or a fatty alcohol; RJ generally is a hydrogen.
[0060] The OH-group of the phosphate is a hydroxyl radical or hydroxyl anion (i.e., hydroxide) form, dependent on degree of the group ionization. Furthermore, R4 may be a proton or a short-chain alkyl group, substituted by a tri-short-chain alkylammonium group, such as a trimethylammonium group, or an amino-substituted short-chain alkyl group, such as 2-trimethylammonium ethyl group (cholinyl) or 2-dimethylammonium short alkyl group.
[0061] A sphingophospholipid is, for example, a compound of Formula IIB:
R1—Sphingosine— O— PHO2-O-R4 (IIB)
wherein R1 is a fatty-acid attached via an amide bond to the nitrogen of the sphingosine and R4 has the meanings given under Formula II.
[0062] A lipid preferably is a substance of formulae II or IIB, wherein R1 and/or R2 are acyl or alkyl, n-hydroxyacyl or n-hydroxyalkyl, but may also be branched, with one or more methyl groups attached at almost any point of the chain; usually, the methyl group is near the end of the chain (iso or anteiso). The radicals R1 and R2 may moreover either be saturated or unsaturated (mono-, di- or poly-unsaturated). R3 is hydrogen and R4 is 2-trimethylammonium ethyl (the latter corresponds to the phosphatidyl choline head group), 2-dimethylammonium ethyl, 2 -methyl ammonium ethyl or 2-aminoethyl (corresponding to the phosphatidyl ethanolamine head group). R4 may also be a proton (giving phosphatidic acid), a serine (giving phosphatidylserine), a glycerol (giving phosphatidylglycerol), an inositol (giving phosphatidylinositol), or an alkylamine group (giving phosphatidylethanolamine in case of an ethylamine), if one chooses to use a naturally occurring glycerophosphoUpid. Otherwise, any other sufficiently polar phosphate ester, such that will form a lipid bilayer, may be considered as well for making the formulations of the disclosure.
[0063] Table 1 lists preferred phospholipids in accordance with the disclosure.
Figure imgf000020_0001
[0064] The preferred lipids in context of this disclosure are uncharged and form stable, well hydrated bilayers; phosphatidylcholines, phosphatidylethanolamine, and sphingomyelins are the most prominent representatives of such lipids. Any of those can have chains as listed in the Table 1 , the ones forming fluid phase bilayers, in which lipid chains are in disordered state, being preferred.
[0065] Different negatively charged, i.e., anionic, lipids can also be incorporated into vesicular lipid bilayers to modify the (cationic) drug loading into or release from the resulting lipid aggregates. Attractive examples of such charged lipids are phosphatidylglycerols, phosphatidylinositols and, somewhat less preferred, phosphatide acid (and its alkyl ester) or phosphatidylserine. It will be realized by anyone skilled in the art that it is less commendable to make vesicles just from the charged lipids than to use them in a combination with electro- neutral bilayer component(s). In case of using charged lipids, buffer composition and/or pH care must selected so as to ensure the desired degree of lipid head-group ionization and/or the desired degree of electrostatic interaction between the, oppositely, charged drug and lipid molecules. Moreover, as with neutral lipids, the charged bilayer lipid components can in principle have any of the chains listed in the Table 1. The chains forming fluid phase lipid bilayers are clearly preferred, however, both due to vesicle adaptability increasing role of increasing fatty chain fluidity and due to better ability of lipids in fluid phase to mix with each other, and with drugs.
[0066] The fatty acid- or fatty alcohol-derived chain of a lipid is typically selected amongst the basic aliphatic chain types given in the following tables:
Figure imgf000021_0001
Figure imgf000022_0001
[0067] Other double bond combinations or positions are possible as well.
[0068] Suitable fatty residues can furthermore be branched, for example, can contain a methyl group in an iso or anteiso position of the fatty acid chain, or else closer to the chain middle, as in 10-Λ-methyloctadecanoic acid or tuberculostearic chain. Relatively important amongst branched fatty acids are also isoprenoids, many of which are derived from
3,7,11 ,15-tetramethylhexadec-trans-2-en-1-ol, the aliphatic alcohol moiety of chlorophyll. Examples include 5.9,13,17-tetramethyloctadecanoic acid and especially 3,7,1 1,15- tetramethylhexadecanoic (phytanic) and 2,6,10,14-tetramethylpentadecanoic (pristanic) acids. A good source of 4,8,12-trimethyltridecanoic acid are marine organisms. Combination of double bonds and side chains on a fatty residue are also possible.
[0069] Alternatively, suitable fatty residues may carry one or a few oxy- or cyclic groups, especially in the middle or towards the end of a chain. The most prominent amongst the later, alicyclic fatty acids, are those comprising a cyclopropane (and sometimes cyclopropene) ring, but cyclohexyl and cycloheptyl rings can also be found and might be useful for purposes of this disclosure. 2-(D)-Hydroxy fatty acids are more ubiquitous than alicyclic fatty acids, and are also important constituents of sphingolipids. Also interesting are 15-hydroxy- hexadecanoic and 17-hydroxy-octadecanoic acids, and maybe 9-hydroxy-octadeca-^rα«.v- 10,frαrø-12-dienoic (dimorphecolic) and 13-hydroxy-octadeca-c«-9,/Tαrø-l l-dienoic
(coriolic) acid. Arguably the most prominent hydroxyl-fatty acid in current pharmaceutical use is ricinoleic acid, (D-(-)12-hydroxy-octadec-c«-9-enoic acid, which comprises up to 90% of castor oil, which is also often used in hydrogcnated form. Epoxy-, methoxy-, and furanoid-fatty acids are of only limited practical interest in the context of this disclosure,
[0070] Generally speaking, unsaturation, branching or any other kind of derivatization of a fatty acid is best compatible with the intention of present disclosure of the site of such modification is in the middle or terminal part of a fatty acid chain. The cis- unsaturated fatty acids are also more preferable than trans-unsataτated fatty acids and the fatty radicals with fewer double bonds are preferred over those with multiple double bonds, due to oxidation sensitivity of the latter. Moreover, symmetric chain lipids are generally better suited than asymmetric chain lipids.
[0071] A preferred lipid of the Formula II is, for example, a natural phosphatidylcholine, which used to be called lecithin. It can be obtained from egg (rich in palmitic, Cj6 o, and oleic, Cis i, but also comprising stearic. C]8 o> palmitoleic, C)6 i, linolenic, C]82, and arachidonic, C204. radicals), soybean (rich in unsaturated Ci8 chains, but also containing some palmitic radical, amongst a few others), coconut (rich in saturated chains), olives (rich in
monounsaturated chains), saffron (safflower) and sunflowers (rich in n-6 linoleic acid), linseed (rich in n-3 linolenic acid), from whale fat (rich in monounsaturated n-3 chains), from primrose or primula (rich in n-3 chains). Preferred, natural phosphatidyl ethanolamines (used to be called cephalins) frequently originate from egg or soybeans. Preferred sphingomyelins of biological origin are typically prepared from eggs or brain tissue. Preferred
phosphatidylserines also typically originate from brain material whereas phosphatidylglycerol is preferentially extracted from bacteria, such as E. CoIi, or else prepared by way of transphosphatidylation, using phospholipase D, starting with a natural phosphatidylcholine. The preferably used phosphatidylinositols are isolated from commercial soybean
phospholipids or bovine liver extracts. The preferred phosphatidic acid is either extracted from any of the mentioned sources or prepared using phospholipase D from a suitable phosphatidylcholine.
[0072] Furthermore, synthetic phosphatidyl cholines (R4 in Formula II corresponds to 2- trimethylammonium ethyl), and R1 and R2 are aliphatic chains, as defined in the preceding paragraph with 12 to 30 carbon atoms, preferentially with 14 to 22 carbon atoms, and even more preferred with 16 to 20 carbon atoms, under the proviso that the chains must be chosen so as to ensure that the resulting ESAs comprise fluid lipid bilayers. This typically means use of relatively short saturated and of relatively longer unsaturated chains. Synthetic
sphingomyelins (R4 in Formula HB corresponds to 2-trimethylammonium ethyl), and R1 is an aliphatic chain, as defined in the preceding paragraph, with 10 to 20 carbon atoms, preferentially with 10 to 14 carbon atoms per fully saturated chain and with 16-20 carbon atoms per unsaturated chain.
[0073] Synthetic phosphatidyl ethanolamines (R4 is 2-aminoefhyl), synthetic phosphatidic acids (R1 is a proton) or its ester (R4 corresponds, for example, to a short-chain alkyl, such as methyl or ethyl), synthetic phosphatidyl serines (R4 is L- or D-serine), or synthetic
phosphatidyl (poly)alcohols, such as phosphatidyl inositol, phosphatidyl glycerol (R4 is L- or D-glycerol) are preferred as lipids, wherein R1 and R2 are fatty residues of identical or moderately different type and length, especially such as given in the corresponding tables given before in the text. Moreover. R1 can represent alkenyl and R2 identical hydroxyalkyl groups, such as tetradecylhydroxy or hexadecylhydroxy. for example, in ditetradecyl or dihexadecylphosphatidyl choline or efhanolamine, R1 can represent alkenyl and R2
hydroxyacyl, such as a plasmalogen (R4 trimethylammonium ethyl), or R1 can be acyl, such as lauryl, myristoyl or palmitoyl and R2 can represent hydroxy as, for example, in natural or synthetic lysophosphatidyl cholines or lysophosphatidyl glycerols or lysophosphatidyl ethanolamines, such as 1 -myristoyl or 1-palmitoyllysophosphatidyl choline or -phosphatidyl ethanolamine; frequently, RJ represents hydrogen. [0074] A lipid of Formula HB is also a suitable lipid within the sense of this disclosure. In Formula HB, n=l, R1 is an alkenyl group, R2 is an acylamido group, R3 is hydrogen and R4 represents 2-trimethylammonium ethyl (choline group). Such a lipid is known under the name of sphingomyelin.
[0075] Suitable lipids furthermore are a lysophosphatidyl choline analog, such as 1 - lauroyl-1,3-dihydroxypropane-3-phosphoryl choline, a monoglyceride, such as monoolein or monomyristin, a cerebroside, ceramide polyhexoside, sulfatide, sphingoplasmalogcn, a ganglioside or a glyceride, which does not contain a free or esterified phosphoryl or phosphono or phosphino group in the 3 position. An example of such a glyceride is diacyl glyceride or 1 -alkenyl- l-hydroxy-2-acyl glyceride with any acyl or alkenyl groups, wherein the 3-hydroxy group is etherified by one of the carbohydrate groups named, for example, by a galactosyl group such as a monogalactosyl glycerin.
[0076] Lipids with desirable head or chain group properties can also be formed by biochemical means, for example, by means of phospholipases (such as phospholilpase Al, A2, B, C and, in particular, D), desaturases, elongases, acyl transferases, etc., from natural or synthetic precursors.
[0077] Furthermore, a suitable lipid is any lipid, which is contained in biological membranes and can be extracted with the help of apolar organic solvents, such as chloroform. Aside from the lipids already mentioned, such lipids also include, for example, steroids, such as estradiol, or sterols, such as cholesterol, beta-sitosterol, desmosterol, 7-keto-cholesterol or beta-cholestanol, fat-soluble vitamins, such as retinoids, vitamins, such as vitamin Al or A2, vitamin E, vitamin K, such as vitamin Kl or K2 or vitamin Dl or D3, etc.
[0078] The less soluble amphiphilic components comprise or preferably comprise a synthetic lipid, such as myristoleoyl, palmitolcoyl, petroselinyl, petroselaidyl, oleoyl, elaidyl, cis- or trans-vaccenoyl, linolyl, linolenyl, linolaidyl, octadecatetraenoyl, gondoyl,
eicosaenoyl, eicosadienoyl, eicosatrienoyl, arachidoyl, cis- or trans-docosaenoyl,
docosadienoyl, docosalrienoyl, docosatetraenoyl, lauroyl, tridecanoyl, myristoyl,
pentadecanoyl, palmitoyl, heptadecanoyl, stearoyl or nonadccanoyl, glycerophospholipid or corresponding derivatives with branched chains or a corresponding dialkyl or sphingosin derivative, glycolipid or other diacyl or dialkyl lipid.
[0079] The more soluble amphiphilic components(s) is/are frequently derived from the less soluble components listed above and, to increase the solubility, substituted and/or complexed and/or associated with a butanoyl, pentanoyl, hexanoyl, heptanoyl, octanoyl, nonanoyl, decanoyl or undecanoyl substituent or several, mutually independent, selected substituents or with a different material for improving the solubility.
[0080] A further suitable lipid is a diacyl- or dialkyl-glycerophosphoetha- nolamine azo polyethoxylene derivative, a didecanoylphosphatidyl choline or a
diacylphosphoolligomaltobionarnide.
[0081] In certain embodiments, the amount of lipid in the formulation is from about 1% to about 10%, about 1% to about 4%, about 4% to about 7% or about 7% to about 10% by weight. In a specific embodiment, the lipid is a phospholipid. In another specific embodiment, the phospholipid is a phosphatidylcholine.
4.3. SURFACTANT
[0082] The term ""'surfactant'"" has its usual meaning. A list of relevant surfactants and surfactant related definitions is provided in EP 0 475 160 Al (see, e.g., p. 6, 1. 5 to p.14. 1.17) and U.S. Pat. No, 6,165,500 (see, e.g., col. 7, 1. 60 to col. 19, 1. 64), each herein incorporated by reference in their entirety, and in appropriate surfactant or pharmaceutical Handbooks, such as Handbook of Industrial Surfactants or US Pharmacopoeia, Pharm. Eu. In some embodiments, the surfactants are those described in Tables 1-18 of U.S. Patent Application Publication No. 2002/0012680 Al, published January 31, 2002, the disclosure of which is herein incorporated by reference in its entirety. The following list therefore only offers a selection, which is by no means complete or exclusive, of several surfactant classes that are particularly common or useful in conjunction with present patent application. Preferred surfactants to be used in accordance with the disclosure include those with an IILB greater than 12. The list includes ionized long-chain fatty acids or long chain fatty alcohols, long chain fatty ammonium salts, such as alkyl- or alkenoyl-trimethyl-, -dimethyl- and -methyl- ammonium salts, alkyl- or alkenoyl-sulphate salts, long fatty chain dimethyl-aminoxides, such as alkyl- or alkenoyl-dimethyl-aminoxides, long fatty chain, for example alkanoyl, dimethyl-aminoxides and especially dodecyl dimethyl-aminoxide, long fatty chain, for example alkyl-N-methylglucamide- s and alkanoyl-N-methylglucamides, such as MEGA-8, MEGA-9 and MEGA- 10, N-long fatty chain-N,N-dimethyl glycines, for example N-alkyl- N, N-dimethyl glycines, 3-(long fatty chain-dimethylammonio)-alkane- sulphonates, for example 3-(acyidimethylammonio)-alkanesulphonates, long fatty chain derivatives of sulphosuccinate salts, such as bis(2-ethylalkyl) sulphosuccinate salts, long fatty chain- sulphobetaines, for example acyl-sulphobetaines, long fatty chain betaines, such as
EMPIGEN BB or ZWITTERGENT-3-16, -3-14, -3-12, -3-10, or -3-8, or polyethylen-glycol- acylphenyl ethers, especially nonaethylen-glycol-octyl- phenyl ether, polyethylene-long fatty chain-ethers, especially polyethylene-acyl ethers, such as nonaethylen-decyl ether, nonaethylen-dodecyl ether or octaethylene-dodecyl ether, polyethyleneglycol-isoacyl ethers, such as octaethyleneglycol-isotridecyl ether, polyethyleneglycol-sorbitane-long fatty chain esters, for example polyethyleneglycol-sorbitane-acyl esters and especially polyoxyethylene- monolaurate (e.g. polysorbate 20 or Tween 20), polyoxyethylene-sorbitan-monooleate (e.g. polysorbate 80 or Tween 80), polyoxyethylene-sorbitan-monolauroleylate, polyoxyethylene - sorbitan-monopetroselinate, polyoxyethylene -sorbitan— monoelaidate, polyoxyethylene - sorbitan-myristoleylate, polyoxyethylene -sorbitan-palmitoleinylate, polyoxyethylene- sorbitan-p- etroselinylate, polyhydroxyethylcne-long fatty chain ethers, for example polyhydroxyethylene-acyl ethers, such as polyhydroxyethylene-lauryl ethers,
polyhydroxyethylene-myristoyl ethers, polyhydroxyethylene-cetylst- earyl, polyhyd roxyethylene-palmityl ethers, polyhyd roxyethylene-oleoyl ethers, polyhydroxyethylene- palmitoleoyl ethers, polyhydroxyethylene-lino- leyl, polyhydroxyethylen-4, or 6, or 8, or 10, or 12-lauryl, miristoyl, palmitoyl, palmitoleyl, oleoyl or linoeyl ethers (Brij series), or in the corresponding esters, polyhydroxyethylen-laurate, -myristate, -palmitate, -stearatc or -oleate, especially polyhydroxyethylen-8-stearate (Myrj 45) and polyhydroxyethylen-8-oleate, polyethoxylated castor oil 40 (Cremophor EL), sorbitane-mono long fatty chain, for example alkylate (Arlacel or Span series), especially as sorbitane-monolaurate (Arlacel 20, Span 20), long fatty chain, for example acyl-N-methylglucamides, alkanoyl-N-methylglucamides, especially decanoyl-N-methylglucamide, dodecanoyl-N-methylglucamide, long fatty chain sulphates, for example alkyl-sulphates, alkyl sulphate salts, such as lauryl-sulphate (SDS), oleoyl-sulphate; long fatty chain thioglucosides, such as alkylthioglucosides and especially heptyl-, octyl- and nonyl-beta-D-thioglucopyranoside; long fatty chain derivatives of various carbohydrates, such as pentoses, hexoses and disaccharides, especially alkyl-glucosides and maltosides, such as hexyl-, heptyl-, octyl-, nonyl- and decyl-beta-D-glucopyranoside or D- maltopyranoside; further a salt, especially a sodium salt, of cholate, deoxycholate, glycocholate, glycodeoxycholate, taurodeoxycholate, taurocholate, a fatty acid salt, especially oleate. εlaidate, linoleate, laurate, or myristate, most often in sodium form,
lysophospholipids, n-octadecylene-glycerophosphatidic acid, octadecylene- phosphorylglycerol, octadecylene-phosphorylserine, n-long fatty chain-glycero-phosphatidic acids, such as n-acyl-glycero-phosphatidic acids, especially lauryl glycero-phosphatidic acids, oleoyl-glycero-phosphatidic acid, n-long fatty chain-phosphorylglyccrol, such as n- acyl-phosphorylglycerol, especially lauryl-, myristoyl-, oleoyl- or palmitoeloyl- phosphorylglycerol. n-long fatty chain-phosphoryl serine, such as n-acyl-phosphorylserine, especially lauryl-, myristoyl-, oleoyl- or palmitoeloyl-phosphorylserine, n-tetradecyl-glycero- phosphatidic acid, n-tetradecyl-phosphorylglycerol, n-tetradecyl-phosphorylserine, corresponding-, elaidoyl-, vaccenyl-lysophospholipids, corresponding short-chain
phospholipids, as well as all surface active and thus membrane destabilising polypeptides. Surfactant chains are typically chosen to be in a fluid state or at least to be compatible with the maintenance of fluid-chain state in carrier aggregates.
[0083] Table 5 lists preferred surfactants in accordance with the disclosure.
Figure imgf000029_0001
Figure imgf000030_0001
[0084] In certain embodiments, the surfactant is a nonionic surfactant. The surfactant may be present in the formulation in about 1% to about 10%, about 1% to about 4%, about 4% to about 7% or about 7% to about 10% by weight. In certain embodiments, the nonionic surfactant is selected from the group consisting of: polyoxyethylene sorbitans (polysobate surfactants), polyhydroxyethylene stearates or polyhydroxyethylene laurylethcrs (Brij surfactants). In a specific embodiment, the surfactant is a polyoxyethylene-sorbitan- monooieate (e.g. polysorbate 80 or Tween 80). In certain embodiments, the polysorbate can have any chain with 12 to 20 carbon atoms. In certain embodiments, the polysorbate is fluid in the formulation, which may contain one or more double bonds, branching, or cyclo-groups.
4.4. FORMULATIONS
[0085] In some embodiments, the topical vaccine formulations contain 1 to 10% by weight, 1 to 15 % by weight, 1 to 20% by weight, or 1 to 30% of a hapten-carrier conjugate. In some embodiments, the topical vaccine formulations contain 1 to 10% by weight, 1 to 15 % by weight. 1 to 20% by weight, or 1 to 30% by weight of the lipid. In some embodiments, the topical vaccine formulations contain 1 to 10% by weight, 1 to 15 % by weight, 1 to 20% by weight, 1 to 30% by weight, 1 to 40%) by weight, or 1 to 50% by weight of the surfactant.
[0086] The topical vaccine formulations may have a range of lipid to surfactant ratios, f he ratios may be expressed in terms of molar terms (mol lipid /mol surfactant). In some embodiments, the molar ratio of lipid to surfactant in the topical vaccine formulations is from about 1 :2 to about 10: 1. In some embodiments, the ratio is from about 1 : 1 to about 2:1, from about 2: 1 to about 3:1, from about 3 : 1 to about 4: 1, from about 4: 1 to about 5 : 1 , or from about 5:1 to about 10: 1. In specific embodiments, the lipid to surfactant ratio is about 1.0, about 1.25, about 1.5, about 1.75, about 2.0, about 2.5, about 3.0, or about 4.0.
[0087] The topical vaccine formulations may have varying ratios of the hapten-carrier conjugate to lipid. The ratios may be expressed in terms of molar ratios (mol hapten-carrier conjugate /mol lipid). In some embodiments, the molar ratio of the hapten-carrier conjugate to lipid in the topical vaccine formulations is from about 1 :50 to about 50: 1, from about 1 :25 to about 25: 1, from about 1 :10 to about 10:1, from about 1 :5 to about 5:1, from about 1 :50 to about 50:1 , or from about 0.2:1 to about 2:1. In certain embodiments, the ratio is from about 0.2:1 to about 0.7:1 , from about 0.7:1 to about 1.2: 1 , from about 1.2:1 to about 1 ,7:1, or from about 1.7:1 to about 2:1. [0088] The topical vaccine formulations may also have varying amounts of total amount of the following three components: the hapten-carrier conjugate, lipid and surfactant combined (TA). The TA amount may be stated in terms of weight percent of the total composition. In some embodiments, the TA is from about 1% to about 40%, about 5% to about 30%, about 7.5% to about 15%, about 5% to about 10%. about 10% to about 20%, or about 20% to about 30%. In specific embodiments, the TA is 8%, 9%, 10%. 15%, or 20%.
[0089] Selected ranges for total lipid amounts, lipid/surfactant ratios (mol/mol) and the hapten-carrier conjugate /surfactant ratios (mol/mol) for the topical vaccine formulations are described in Table 6 below:
Figure imgf000032_0001
Figure imgf000033_0001
[0090] In some embodiments, the topical vaccine formulations optionally contain one or more of the following ingredients: co-solvents, chelators, buffers, antioxidants, preservatives, mierobicides, emollients, humectants, lubricants, and thickeners. Preferred amounts of optional components are described in Table 7.
[0091] In some embodiments, the topical vaccine formulations include a buffer to adjust the pH of the aqueous solution to a range from pH 3.5 to pH 9.5, pH 4 to pH 7.5. or pH 4 to pH 6.5. Examples of buffers include, but are not limited to, acetate buffers, lactate buffers, phosphate buffers, and propionate buffers. In some embodiments, the buffer concentration is in the range between about 1 mM and about 1 M, about 1 mM and about 100 mJVl, or about 1 mM and about 10 mM.
[0092] In some embodiments, the topical vaccine formulations are formulated in aqueous media. In some embodiments, the formulations may be formulated with or without co- solvents, such as lower alcohols.
[0093] A ""microbicide"" or ""antimicrobial"" agent is commonly added to reduce the bacterial count in pharmaceutical formulations. Some examples of mierobicides arc short chain alcohols, including ethyl and isopropyl alcohol, chlorbutanol, benzyl alcohol, chlorbenzyl alcohol, dichlorbenzylalcohol, hexachlorophene; phenolic compounds, such as cresol, 4-chloro-m-cresoI, p-chloro-m-xylenol, dichlorophene, hexachlorophene, povidon- iodine; parabenes, especially alkyl-parabenes, such as methyl-, ethyl-, propyl-, or butyl- paraben, benzyl paraben; acids, such as sorbic acid, benzoic acid and their salts; quaternary ammonium compounds, such as alkonium salts, e.g., a bromide, benzalkonium salts, such as a chloride or a bromide, cetrimonium salts, e.g., a bromide, phenoalkecinium salts, such as phenododecinium bromide, cetylpyridinium chloride and other salts; furthermore, mercurial compounds, such as phenylmercuric acetate, borate, or nitrate, thiomersal, chlorhexidine or its gluconate, or any antibiotically active compounds of biological origin, or any suitable mixture thereof. [0094] Examples of '"antioxidants"" are butylated hydroxyanisol (BHA), butylated hydroxytoluene (BHT) and di-tert-butylphenol (LY178002, LY256548, HWA-131, BF-389, CI-986, PD- 127443, E-5119, BI-L-239XX, etc.), tertiary butylhydroquinone (TBHQ), propyl gallate (PG), l-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ); aromatic amines
(diphenylamine, p-alkylthio-o-anisidine, ethylenediamine derivatives, carbazol,
tetrahydroindenoindol); phenols and phenolic acids (guaiacol, hydroquinone, vanillin, gallic acids and their esters, protocatechuic acid, quinic acid, syringic acid, ellagic acid, salicylic acid, nordihydroguaiaretic acid (NDGA), eugenol); tocopherols (including tocopherols (alpha, beta, gamma, delta) and their derivatives, such as tocopheryl-acylate (e.g., -acetate, - laurate, myristate, -palmitate, -oleate, -linoleate, etc., or any other suitable tocopheryl- lipoate), tocopheryl-polyoxyethyleπe-succinate; trolox and corresponding amide and thiocarboxamide analogues; ascorbic acid and its salts, isoascorbate, (2 or 3 or 6)-o- alkylascorbic acids, ascorbyl esters (e.g., 6-o-lauroyl, myristoyl, palmitoyl-, oleoyl, or linoleoyl-L-ascorbic acid, etc.). Also useful are the preferentially oxidized compounds, such as sodium bisulphite, sodium metabisulphite, thiourea; chelating agents, such as ethylene glycol-bis-(2-aminoethyl)-N,N,N',N'-tetraacetic acid (EDTA), ethylenedioxy-diethylene- dinitrilo-tetraacetic acid (GDTA), desferral; miscellaneous endogenous defence systems, such as transferrin, lactoferrin, ferritin, cearuloplasmin, haptoglobion, heamopexin, albumin, glucose, ubiquinol-10); enzymatic antioxidants, such as superoxide dismutase and metal complexes with a similar activity, including catalase, glutathione peroxidase, and less complex molecules, such as beta-carotene, bilirubin, uric acid; flavonoids (flavones, flavonols, flavonones, flavanonals, chacones, anthocyanins), N-acetylcysteine, mesna, glutathione, thiohistidine derivatives, triazoles; tannines, cinnamic acid, hydroxycinnamatic acids and their esters (coumaric acids and esters, caffeic acid and their esters, ferulic acid, (iso-) chlorogenic acid, sinapic acid); spice extracts (e.g., from clove, cinnamon, sage, rosemary, mace, oregano, allspice, nutmeg); carnosic acid, carnosol, carsolic acid: rosmarinic acid, rosmaridiphenol, gentisic acid, ferulic acid; oat flour extracts, such as avenanthramide 1 and 2; thioethers, dithioethers, sulphoxides, tetralkylthiuram disulphides; phytic acid, steroid derivatives (e.g.. U74006F); tryptophan metabolites (e.g., 3-hydroxykynurenine, 3- hydroxyanthranilic acid), and organochalcogenides,
[0095] ""Thickeners"'" are used to increase the viscosity of pharmaceutical formulations to and may be selected from selected from pharmaceutically acceptable hydrophilic polymers, such as partially etherified cellulose derivatives, comprising carboxymethyl-, hydroxyethyl-, hydroxypropyl-, hydroxypropylmethyl- or methyl-cellulose; completely synthetic hydrophilic polymers comprising polyacrylates, polymethacrylates,
poly(hydroxyethyl)-, poly(hydroxypropyl)-, poly(hydroxypropylmethyl)methacrylate, polyacrylonitrile, methallyl-sulphonate, polyethylenes, polyoxyethylenes, polyethylene glycols, polyethylene glycol-lactide, polyethylene glycol-diacrylate, polyvinylpyrrolidone, polyvinyl alcohols, poly(propylmethacrylamide), poly(propylene fumarate-co-ethylene glycol), poloxamers, polyaspartamide, (hydrazine cross-linked) hyaluronic acid, silicone; natural gums comprising alginates, carrageenan, guar-gum, gelatine, tragacanth, (amidated) pectin, xanthan, chitosan collagen, agarose; mixtures and further derivatives or co-polymers thereof and/or other pharmaceutically, or at least biologically, acceptable polymers.
[0096] Examples of ""emollients"" include, but are not limited to, glycerine, propylene glycol, sorbitol, and mannitol.
[0097] In some embodiments, the topical vaccine formulations further comprise a polar liquid medium. In some embodiments, the topical vaccine formulations may be administered in an aqueous medium. In some embodiments, the topical vaccine formulations provided herein may be in the form of a solution, suspension, emulsion, cream, lotion, ointment, gel, spray, film forming solution, lacquer or a patch soaked with the formulation.
[0098] Table 7 lists preferred excipients for the formulation.
Figure imgf000036_0001
4.4. VESICULAR FORMULATIONS
[0099] While not to be limited to any mechanism of action or any theory, the
formulations provided herein may form vesicles or ESAs characterized by their adaptability, deformability, or penetrability.
[0100] The term vesicle or aggregate ""adaptability"" which governs the ""tolerable surface curvature"" is defined as the ability of a given vesicle or aggregate to change easily its properties, such as shape, elongation ratio, and surface to volume ratio. The vesicles provided herein may be characterized by their ability to adjust the aggregates* shape and properties to the anisotropic stress caused by pore crossing. Sufficient adaptability implies that a vesicle or an aggregate can sustain different unidirectional forces or stress, such as one caused by pressure, without extensive fragmentation, which defines a ""stable""' aggregate. If an aggregate passes through a barrier fulfilling this condition the terms ""adaptability"" and (shape) ""deformability"" plus ""permeability"" are essentially equivalent. A ""barrier"" in the context of this disclosure is (as in, for example, EP 0 475 160 and WO 98/17255) a body with through-extending narrow pores, such narrow pores having a radius which is at least 25% smaller than the radius of the ESAs (considered as spherical) before said ESAs permeate through such pores.
[0101] The term "'"narrow"" used in connection with a pore implies that the pore radius is significantly, typically at least 25%, smaller than the radius of the entity tested with regard to its ability to cross the pore. The necessary difference typically should be greater for the narrower pores. Using 25% limit is therefore quite suitable for >150 nm diameter whereas >100% difference requirement is more appropriate for the smaller systems, e.g., with <50 nm diameter. For diameters around 20 nm, aggregate diameter difference of at least 200% is often required.
[0102] The term ""semipermeable"" used in connection with a barrier implies that a solution can cross transbarrier openings whereas a suspension of non-adaptable aggregates (large enough for the above definition of ""narrow"" pores to apply) cannot. Conventional lipid vesicles (liposomes) made from any common phosphatidylcholine in the gel lamellar phase or else from any biological phosphatidylcholine/cholesterol 1/1 mol/mol mixture or else comparably large oil droplets, all having the specified relative diameter, are three examples for such non-adaptable aggregates. [0103] The term """stable"" means that the tested aggregates do not change their diameter spontaneously or under the transport related mechanical stress (e.g. during passage through a semipermeable barrier) unacceptably, which most often means only to a pharmaceutically acceptable degree. A 20-40% change is normally considered acceptable; the halving or doubling of aggregate diameter is borderline and a greater change in diameter is typically unacceptable. Alternatively and very conveniently, the change in aggregate diameter resulting from pore crossing under pressure is used to assess system stability; the same criteria are then applied as for ""narrow"" pores, mutatis mutandis. To obtain the correct value for aggregate diameter change, a correction for flux/vortex effects may be necessary. These procedures are described in greater detail in the publications of the applicant in Cevc el al., Biochim. Biophys. Acta 2002; 1564:21-30.
[0104] Non-destructing passage of ultradeformable, mixed lipid aggregates through narrow pores in a semi-permeable barrier is thus diagnostic of high aggregate adaptability. If pore radius is two times smaller than the average aggregate radius the aggregate must change its shape and surface-to- volume ratio at least 100% to pass without fragmentation through the barrier. An easy and reversible change in aggregate shape inevitably implies high aggregate deformability and requires large surface-to-volume ratio adaptation. A change in surface-to- volume ratio per se implies: a) high volume compressibility, e.g. in the case of compact droplets containing material other than, and immiscible with, the suspending fluid; b) high aggregate membrane permeability, e.g. in the case of vesicles that are free to exchange fluid between inner and outer vesicle volume.
4-5 METHODS QF ADMINISTRATION
[0105] Also provided herein are methods of administering a pharmaceutical composition comprising a hapten-carrier conjugate, a lipid, and a surfactant.
4.5.1 Immunogenicity of the hapten-carrier conjugate
[0106] In some embodiments, the hapten-carrier conjugate in the topical vaccine formulations elicits an immune response in a cell, tissue, organ, and/or subject or patient when administered, As used herein, the terms "''subject'"' or ""patient"" are used
interchangeably. As used herein, the terms ""subject"'" and '""subjects"" refer to an animal (e.g. , birds, reptiles, and mammals), preferably a mammal including a non-primate (e.g., a camel, donkey, zebra, cow, pig, horse, goat, sheep, cat, dog, rat, and mouse) and a primate (e.g., a monkey, chimpanzee, and a human), and most preferably a human. In some embodiments, the subject or patient has a drug addiction. In some embodiments, the subject or patient is at risk for developing or re-developing a drug addiction.
[0107] In some embodiments of the invention, the hapten-carrier conjugate in the topical vaccine formulations induce an immune response from the adaptive immune system, such as a B cell response and/or a T cell response. In some embodiments, the immune response induced by the hapten-carrier conjugate in the topical vaccine formulations is an antibody response. In some embodiments, the hapten-carrier conjugate in the topical vaccine formulations induces a humoral immune response, such as an interferon response and/or an interleukin response, e.g. , an interleukin-4 response. In some embodiments, the hapten- carrier conjugate in the topical vaccine formulations induces one or more types of immune response but not another immune response. In certain embodiments, the hapten-carrier conjugate induces a combination of immune responses. Moreover, in some embodiments, the hapten-carrier conjugate in the topical vaccine formulations can induce a robust IFN response which has other biological consequences in vivo, affording protection against subsequent diseases or conditions or concurrent diseases or conditions. In some embodiments, the hapten-carrier conjugate in the topical vaccine formulations can induce a robust TNFα or interleukin response which has other biological consequences in vivo, affording protection against subsequent diseases or conditions or concurrent diseases or conditions.
4.5.2 Administration to a patient
[0108] In some embodiments, the topical vaccine formulation comprising the hapten- carrier conjugate may be administered to a naϊve subject, i.e., a subject that does not have a disease, condition, drug addiction, or has not been and is not currently infected with an infectious agent. In some embodiments, the topical vaccine formulation comprising the hapten-carrier conjugate may be administered to a naϊve subject that is predisposed to acquiring such disease, condition, drug addiction, or infection. In some embodiments, the topical vaccine formulation comprising the hapten-carrier conjugate may also be
administered to a subject that has and/or has had a disease, condition, drug addiction, or infection.
[0109] In some embodiments, the topical vaccine formulation is administered as a patch. Patches contemplated for use with the topical vaccine formulations of the invention include those disclosed in the following references, herein incorporated by reference in their entirety: """Transcutaneous immunization with heat-labile enterotoxin: development of a needle-free vaccine patch,"" Expert Rev Vaccines 2007;6(5): 809-819; ""Transcutaneous immunization with the heat-labile toxin (LT) of enterotoxigenic Escherichia coli (ETEC): protective efficacy in a double-blind, placebo-controlled challenge study,-'*' Vaccine 2007; 25(18):3684- 3691 ; Immunization using the skin: new approaches,-'" In: Moingeon P (Ed.), Vaccines, Frontiers in Design and Development, Horizon Scientific Press, Ltd., Wymondham, UK, pp. 81-104, 2005; ""Dose sparing for influenza vaccination using intradermal immunization,"" N Engl J Med 2004:351(22):2295-2301 ; ""'Transcutaneous immunization: a human vaccine delivery strategy using a patch,"" Nat M?d 2000;6(12):1403-1406; US 2007/0009542 Al (Levin et al.); US 2007/0088248 Al (Glenn et al); US 2003/0125278 Al (Tang et al.); US 2002/0193729 Al (Cormier et al.); WO 00/44349 (IDEA AG); U.S. Patent No. 4,908,213 (Govil et al.); U.S. Patent No. 5,079,008 (Sinnreich et al.); and U.S. Patent No. 5,656,286 (Miranda et al.).
[0110] In certain embodiments, the patch further comprises one or more antipuritics, adhcsives, solvents, co-solvents, plasticizers, polymeric matrices, stabilizers, thickeners, preservatives, and the like.
[0111] Examples of topical antipruritics effective in reducing itching during transdermal nicotine delivery include bisabolol or α(-)bisabolol, oil of chamomile, chamazulene, allantoin, D-panthenol glycyrrhetenic acid, corticosteroids, antihistamines, and combinations thereof. In one embodiment, the antipruritic is a combination of 18-β-glycyrrhetenic acid and D-panthenol (i.e., ANTIPHLOGISTICUM "ARO").
[0112] Examples of pharmaceutically acceptable pressure sensitive adhesives include acrylic, silicone, vinyl acetate and synthetic or natural rubber adhesives as well as other adhesives useful in transdermal drug delivery. The adhesives may be used alone or in combination to prepare an adhesive drug matrix or may be applied to the skin-contacting surface of a polymeric matrix or reservoir patch to adhere said patch to the skin. Examples of adhesives are acrylic adhesives such as RA 2484, RA 2333, RA 2397, R 363 and R 362 from Monsanto Co. Other acrylic adhesives, such as Durotak, manufactured by Morton Thiokol, Inc., and Neocryl XA5210bby Polyvinyl Chemicals, Ltd. may be utilized. Vinyl acetate adhesives include Flexcryl-1614, 1617, 1618 and 1625 from Air Products. Numerous silicone based adhesives may be used, such as Q72929, Q27406, X72920 and 355, each manufactured by Dow-Corning. Natural and synthetic rubbers include polyisobutylenes, neoprenes, polybutadienes and polyisoprenes. [0113] Polymeric matrix-forming agents include pharmaceutically acceptable polymers such as polyvinyl alcohol, polyvinylpyrolidones, gelatin and partially hydro lyzed polyvinyl alcohols.
[0114] Examples of solvents include water and polar organic solvents, such as ethanol, and mixtures thereof. In some embodiment, the patched comprise co-solvents, for example, mineral oil, silicone-based liquids, and low molecular weight polyisobutylenes.
[0115] Examples of preservatives, antioxidants and chelating agents that can be included in the transdermal patch include butylated hydroxyanisole (BHA), butylatcd hydroxytoluene (BHT), sodium metabisulfate, α-tocopherol, maleic acid, ethylenediaminetetraacetic acid (EDTA), and cysteine hydrochloride.
[0116] Components useful for imparting the desired wear and pharmacokinetic characteristics to the patch include, for example, polymeric matrix-forming materials added to facilitate curing of the adhesives, for example Aerotex Resin 3730 (American Cyanamid), and a thickener added to adjust the viscosity of the polymer mixture to the desired viscosity for coating on a backing material. The thickener can be an acrylic polymer thickener such as AMSCO 6038 A (Unocal), methyl cellulose and hydroxypropylmethyl cellulose. Plasticizers may be added to impart softness and flexibility to the adhesive, a typical plasticizer being glycerin. Stabiliziers, added to prevent degradation by heat and light and to improve aging characteristics, include polyvinylpyrolidone.
[0117] A rate-controlling membrane may be included to effect controlled release of hap ten-carrier conjugate from the patch. Materials suitable for rate-controlling membranes include cthylene-vinyl acetate (EVA) copolymer membranes (e.g. 1-20% vinyl acetate), polyvinylalcohol (PVA) gels and silicone films.
[0118] In certain embodiments, the patch comprises a) a backing layer which is impermeable to the components of the adhesive layer b), b) an adhesive layer capable of releasing the hapten-carrier conjugate and consisting of a permeable polymeric material which is compatible with the skin and contains at least one topical vaccine formulation which is capable of permeation across the skin, a combination of eucalyptol having a purity of at least 70% with an additional flux enhancer and further optional pharmaceutical excipients, and c) a protective release liner which can be peeled from the adhesive contact layer b).
[0119] In these embodiments, the backing layer comprises a material or a combination of materials which must be impermeable to the components of the adhesive layer, especially to liquid components of the formulation. The backing layer may be made of high- or low- density polymers such as polyethylene, polypropylene, polyvinyl chloride, polyethylene terephthalate or also cellulose acetate or vinyl acetate/vinyl chloride copolymers and combinations thereof, preferably of composite films.
[0120] In these embodiments, the adhesive layer b) is located between the backing layer a) and the release liner c) and consists of a polymeric material which is compatible with the skin and is in the form of a matrix which contains the topical vaccine formulation and, for enhancing penetration through the skin, a combination of eucalyptol and an additional flux enhancer and further optional excipients.
]0121J Suitable polymeric materials are permeable to the topical vaccine formulation as well as to the combination of eucalyptol and the additional flux enhancer and, besides being compatible to the skin, have sufficient adhesiveness that the therapeutic system remains attached to the skin for at least one day and can subsequently be removed without exerting additional force. Further, suitable polymeric materials are dimensionally stable, despite the solvent properties of eucalyptol and the chemically pure 1 ,8-cineol. Examples of such polymeric materials are silicone rubber (silicones), for example linear polysiloxanes in which the silicon atoms in the siloxane chain are substituted by two identical or different alkyl groups such as methyl or ethyl groups, aryl groups, for example phenyl groups, alkcnyl groups such as vinyl or allyl groups, alkylaryl groups such as tolyl or xylyl groups, or alkyl groups, for example benzyl groups, and the terminal silicon atoms are substituted by three of the above organic radicals. The preparation of these silicones is disclosed in U.S. Pat. Nos. 2,541,137, 2.723,966, 2,863,846, 2,890,188, 2,927,907, 3,002,951 and 3,035,016. Silicones which are vulcanisable at room temperature are preferred. Further suitable polymeric materials are hydrophilic polymers of monoesters of unsaturated carboxylic acids such as acrylic acid or methacrylic acid, for example the polyhydroxyethylacrylates or
polyhydroxyethylmethacrylates thereof, the preparation of which is disclosed in U.S. Pat. Nos. 2,976,576 and 3,220,960, as well as copolymers of water-soluble aliphatic or cyclic vinyl amides, for example poly-N-vinylmethylacetamide, poly-N-vinylethyϊacetamide, poly- N-vinyl-methylpropionamide, poly-N-vinylethylpropionamide, poly-N-vinylmethyl- isobutyramide, poly-N-vinyl-2-pyrrolidone, poly-N-vmyl-2-piperidone, poly-N-vinyl- .epsilon.-caprolactam, poly-N-vinyl-5-methyl-2-pyrrolidone or poly-N-vinyl-3-methyl-2- pyrrolidone, preferably poly-N-vinylpyrrolidone having an average molecular weight of about 10,000 to 360,000, with water-soluble polyvinyl acetate or polyvinyl alcohol of different acetate content, for example polyvinyl acetate having a molecular weight of about 5,000 to 400,000 or polyvinyl alcohol having a degree of hydrolysis of about 85-98% and a degree of polymerization of about 500 to 2,500. Preferred polymers are natural or synthetic rubber, for example polyisoprene, 1 ,4-butadiene polymer or polyisobutylene and mixtures thereof. Particularly preferred polymers are mixtures of different polyisobutylenes, with molecular weight ranges from about 1.0 x IQ3 to about 5.0 x 104, or from about 1.0 x 103 to about 5.0 x 106.
[0122] In these embodiments, the term "eucalyptol having a degree of purity of at least 70%'' includes formulations containing more than 70% to 100% of 1,8-cineol. In various pharmacopeias, formulations containing about 70-95% of 1,8-cineol are also designated as eucalyptus oils, whereas the term "eucalyptol" is used for formulations containing more than 95% of 1 ,8-cineol. Eucalyptus oils are terpentine-containing essential oils which contain eucalyptol or 1,8-cineol as main constituent (more than 70%) and which can be isolated from leaves, roots or the bark of eucalyptus plants of the species Eucalyptus globulus (common eucalyptus tree), Eucalyptus maculata, Eucalyptus cladocalyx or Eucalyptus sideroxylon. By means of conventional purification methods, for example rectification, this eucalyptus oil can be further processed to chemically pure 1 ,8-cineol (more than 99% pure). The adhesive layer b) preferably contains this chemically pure 1,8-cineol together with an additional flux enhancer.
[0123] Flux enhancers, also known in the literature as penetration enhancers, accelerators or sorption promoters, have the property of permitting the passage of drugs through the skin that are in themselves impermeable to the skin, especially the stratum corneum which functions as a barrier, and of enhancing the permeability of drugs (e.g., hapten-carrier conjugates) and so make possible the permeation and absorption of therapeutic amounts of the drug to be delivered. The use of a combination of eucalyptol, preferably in the form of chemically pure 1,8-cineol, together with an additional flux enhancer, may substantially increases the amount released and the rate of release and the corresponding absorption (or rate of absorption) through the skin. Examples of additional flux enhancers are monohydric saturated or unsaturated aliphatic, cycloaliphatic or aromatic alcohols of 1 to 12 carbon atoms, for example ethanol, isopropanol, n-hexanol or cyclohexanol; aliphatic, cycloaliphatic or aromatic hydrocarbons of 5 to 12 carbon atoms, for example hexane, cyclohexane, isopropyl benzene and the like; cycloaliphatic or aromatic aldehydes and ketones of 4 to 10 carbon atoms, for example cyclohexanone; amides such as acetamide, N,N-diethyl-m- toluamide, N,N-di-lower alkylacetamide such as N,N-dimethylacetamide or N5N- diethylacetamide, dimethyl propionamide; Cio-C2o alkanoylamides, for example N5N- dimethyllauroylamide, 1-n- Cιo-C2o alkylazacycloheptan-2-one, for example 1-n- dodecylazacycloheptan-2-one (Azone.RTM., Nelson), or N-2-hydroxyethylacetamide; N- alkyl-substituted cyclic amides, for example N-methyl-2-pyrrolidone; and also carriers and/or penetration enhancers such as aliphatic, cycloaliphatic and aromatic esters, for example isopropyl myristate, N,N-di-lower alkylsulfoxide, unsaturated oils, halogenated or nitrated aliphatic, cycloaliphatic hydrocarbons, salicylates, polyalkylene glycol silicates and mixtures thereof. In one embodiment, the flux enhancer is a combination of chemically pure 1 ,8- cineol with N-methyl-2-pyrrolidone, for example a combination of 5 to 9.5 parts by weight of 1,8-cineol and 5 to 0.5 parts by weight (based on 10 parts by weight) of N-methyl-2- pyrrolidone. Compared with pure N-methyl-2-pyrrolidone or 1,8-cineol, such combinations have the advantage that they can be processed with the polymeric material of the adhesive layer b), for example polyisobutylene mixtures, to homogeneous matrix systems of sufficient adhesiveness which are capable of drug release. In a particular embodiment, the flux enhancer is combination of about 9 parts of chemically pure 1,8-cineol and 1 part by weight of N-methyl-2-pyrrolidone is especially preferred.
[0124] In these embodiment, the adhesive layer may optionally further comprise one or more of water, isotonic aqueous saline solution, dextrose in water or saline solution, liquid glyceryl triesters with low molecular fatty acids, lower alkanols, natural oils such as corn oil, groundnut oil, sesame oil, castor oil and the condensates thereof with ethylene oxide and the like, hydrocarbons such as mineral oil of pharmaceutical quality, silicones, emulsifiers such as monoglycerides or diglycerides of fatty acids, phosphatidic acid derivatives such as lecithin or cephalin, polyalkyene glycols such as polyethylene glycol, aqueous phases to which a swelling agent such as sodium carboxymethyl cellulose has been added, sodium alginate, polyvinylpyrrolidone and the like, to which dispersants or emulsifiers such as lecithin may also be added, polyoxyethylene and the like. These excipients may contain further additives such as preservatives, stabilisers, wetting agents, emulsifiers and the like.
[0125] In other embodiments, the patch comprises a pressure-sensitive adhesive composition suitable as a matrix for controlled release of the topical vaccine formulation therefrom comprising a blend of a rubber-based pressure-sensitive adhesive and a soluble polyvinylpyrrolidone (PVP). [0126] In these embodiments, the term '"polyvinylpyrrolidone," or 'TVP" refers to a polymer, either a homopolymer or copolymer, containing N-vinylpyrrolidone as the monomeric unit. Typical PVP polymers are homopolymeric PVPs and the copolymer vinyl acetate vinylpyrrolidone. The homopolymeric PVPs are known to the pharmaceutical industry under a variety of designations including Povidone, Polyvidone, Polyvidonum, Polyvidonum solubile, and PoIy(I -vinyl-2-pyrrolidone). The copolymer vinyl acetate vinylpyrrolidone is known to the pharmaceutical industry as Copolyvidon, Copolyvidone, and Copolyvidonum. In these embodiments, the term '"soluble PVP-' means that the PVP is soluble in water and generally is not substantially cross-linked, and has a molecular weight of less than about 2,000,000.
[0127] In certain of these embodiments, the rubber-based pressure-sensitive adhesive is polysiloxane. Polysiloxane is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 9% to about 97%, from about 8% to about 97%, or from about 14% to about 94% of the total weight of the polymer blend, by weight of the total pressure-sensitive adhesive composition. Suitable polysiloxanes include silicone pressure-sensitive adhesives which are based on two major components: a polymer, or gum, and a tackifying resin. The polysiloxane adhesive is usually prepared by cross-linking the gum, typically a high molecular weight polydiorganosiloxane. with the resin, to produce a three-dimensional silicate structure, via a condensation reaction in an appropriate organic solvent. The ratio of resin to polymer is the most important factor which can be adjusted in order to modify the physical properties of polysiloxane adhesives. Sobieski, et al., "Silicone Pressure Sensitive Adhesives," Handbook of Pressure-Sensitive Adhesive Technology, 2nd cd., pp. 508-517 (D. Satas, ed.), Van Nostrand Reinhold, N. Y. (1989). Further details and examples of silicone pressure sensitive adhesives which are useful in the practice of this invention are described in the following U.S. Patent Nos. 4,591,622; 4,584,355; 4,585,836; and 4,655,767. Suitable silicone pressure-sensitive adhesives are commercially available and include the silicone adhesives sold under the trademarks BIO-PSA X7-3027, BIO-PSA X7- 4503, BIO-PSA X7-4603, BIO-PSA X7-4301 , BIO-PSA X7-4303, BIO-PSA X7-4919, BIO- PSA X7-2685, and BIO-PSA X7-3122 by Dow Corning Corporation, Medical Products, Midland, Mich.
[0128] In other embodiments, pressure-sensitive adhesive composition comprises a ternary blend comprising a rubber-based pressure-sensitive adhesive, a polyacrylate polymer, and a soluble PVP, wherein the rubber-based pressure-sensitive adhesive is a polysiloxane. Polysiloxane is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 9% to about 97% by weight of the total pressure-sensitive adhesive composition, while the polyacrylate polymer is preferably present in an amount ranging from about 5% to about 85%. In one embodiment, the ratio of the polyacrylate polymer to the rubber-based pressure-sensitive adhesive is from about 2:98 to about 96:4, and more preferably from about 2:98 to about 86: 14 by weight. Other embodiments include blends comprising a rubber-based pressure-sensitive adhesive and a polyacrylate polymer, wherein the rubber-based pressure-sensitive adhesive is a polysiloxane. Polysiloxane is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 9% to about 97% by weight of the total pressure-sensitive adhesive composition, while the polyacrylate polymer is preferably present in an amount ranging from about 5% to about 85%. Preferably, the ratio of the polyacrylate polymer to the rubber-based pressure-sensitive adhesive is from about 2:98 to about 96:4, and more preferably from about 2:98 to about 86: 14 by weight. In both binary and ternary blends, soluble PVP is preferably present in the pressure-sensitive adhesive composition in an amount ranging from about 1% to about 20% by weight of the total pressure-sensitive adhesive composition. The pressure-sensitive adhesive compositions may further optionally comprise enhancers, fillers, co-solvents, and excipients.
[0129] In a particular embodiment, a multiple polymer adhesive system comprises a blend of 14-94% by weight of a rubber-based pressure-sensitive adhesive, 5-85% by weight of a polyacrylate polymer, and 2-10% by weight of a soluble PVP, and the multiple polymer adhesive system comprises about 50-99% by weight of the dermal adhesive composition. This multiple polymer adhesive system is combined with a topical vaccine formulation in the amount of 0.1-50% by weight of the total dermal adhesive composition. Optional additives, such as co-solvent for the drug (up to 30% by weight) and enhancers (up to 20% by weight) may be included in the dermal adhesive composition.
[0130] In certain embodiments, the topical vaccine formulation does not result in complete protection or cure (i.e., from drug addiction), but results in a lower level of addiction compared to an untreated subject. Benefits include, but are not limited to, reduced severity of symptoms of the disease or condition and a reduction in the duration of the disease or condition
[0131] In certain embodiments, the topical vaccine formulation is used to protect against a disease or condition (e.g., an infection or drug addiction) in naϊve subjects. [0132] The prophylactic and/or therapeutic effect of the topical vaccine formulation are based, in part, upon achieving or inducing an immune response (e.g., a humoral immune response or adaptive immune response). In one aspect, the immunogenic compositions induce a detectable serum titer of an antibody against an antigen or hapten in either the subject or an animal model thereof (e.g. mouse, rat, pig, goat, sheep or canine model). The serum titer of an antibody can be determined using techniques known to one of skill in the art. e.g., immunoassays such as ELISAs. In a specific embodiment, the antibodies generated by administering an immunogenic composition of the invention are neutralizing antibodies.
[0133] In one embodiment, administration of the topical vaccine formulation to a subject or animal model thereof results in a serum titer of about 1 μg/ml, about 2 μg/ml, about 5 μg/ml, about 6 μg/ml, about 10 μg/ml, about 15 μg/ml, about 20 μg/ml, about 25 μg/ml , about 50 μg/ml, about 75 μg/ml, about 100 μg/ml, about 125 μg/ml, about 150 μg/ml, about 175 μg/ml, about 200 μg/ml, about 225 μg/ml, about 250 μg/ml, about 275 μg/ml, about 300 μg/ml, about 325 μg/ml, about 350 μg/ml, about 375 μg/ml, or more of an antibody that specifically binds to the hapten or antigen. In some embodiments, the serum titer is 100 μg/ml or more. In some embodiments, administration of the topical vaccine formulation results in a plasma titer of 100 μg/ml to 1 mg/ml or more, preferably more than about 500 μg/ml. The immune response may be determined in the subject or in a animal model, which response is then correlated or extrapolated to a predicted response in the subject, e.g., a human or livestock, such as a pig, sheep, goat, or cow.
4,5.3 Methods of use
[0134] In some embodiments, a method for preventing, treating, managing, or ameliorating at least one disease or condition (e.g., a drug addiction or viral infection) in a subject is provided, the method comprising administering to said subject a first dose of an effective amount of the topical vaccine formulation. In some embodiments, the dose of the topical vaccine formulation administered to the subject or animal model is about 10-20 μig. In some embodiments, the dose of the topical vaccine formulation administered to the subject or animal model is about 75-100 μg. In some embodiments, the dose of the topical vaccine formulation administered to the subject or animal model is about 500-1000 μg.
[0135] The amount of the topical vaccine formulation which will be effective in the treatment, prevention and/or amelioration of a particular disease or condition will depend on the nature of the disease, and can be determined by standard clinical techniques. In addition, in vitro assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed in the topical vaccine formulation will also depend on, for example, the seriousness of the disease or disorder, and should be decided according to the judgment of the practitioner and each subject's circumstances. Effective doses may be extrapolated from dose response curves derived from in vitro or animal model test systems.
[0136] In various embodiments, the topical vaccine formulation is administered to a subject in combination with one or more other therapies for the prevention or treatment of at least one disease or condition. In certain embodiments, the therapies (e.g., prophylactic or therapeutic agents) are administered less than 5 minutes apart, less than 30 minutes apart, 1 hour apart, at about 1 hour apart, at about 1 to about 2 hours apart, at about 2 hours to about 3 hours apart, at about 3 hours to about 4 hours apart, at about 4 hours to about 5 hours apart, at about 5 hours to about 6 hours apart, at about 6 hours to about 7 hours apart, at about 7 hours to about 8 hours apart, at about 8 hours to about 9 hours apart, at about 9 hours to about 10 hours apart, at about 10 hours to about 1 1 hours apart, at about 1 1 hours to about 12 hours apart, at about 12 hours to 18 hours apart, 18 hours to 24 hours apart, 24 hours to 36 hours apart, 36 hours to 48 hours apart, 48 hours to 52 hours apart, 52 hours to 60 hours apart, 60 hours to 72 hours apart, 72 hours to 84 hours apart, 84 hours to 96 hours apart, or 96 hours to 120 hours part. In preferred embodiments, two or more therapies are administered within the same patient or subject visit. Non-limiting examples of agents that can be administered in combination with an immunogenic composition of the invention or an antibody generated by the composition of the invention are found below.
[0137] In some embodiments the topical vaccine formulations are useful in the treatment or prevention of a disease, condition, or infection. In certain embodiments, the condition is a drug addiction or drug allergy. In certain specific embodiments, the drug of addiction is nicotine. In other embodiments, the drug of addiction is cocaine. In some embodiments, the topical vaccine formulations are specific for a hapten compared to cellular components and/or compared to the carrier. In another embodiment, the topical vaccine formulations exhibit low cytotoxicity in eukaryotic cells, preferably mammalian cells.
[0138] In some embodiments, the topical vaccine formulation reduces or inhibits a drug addiction, dependence, or allergy. In some embodiment, the immunogenic composition eliminates or reduces the drug addiction or drug dependence by 75%, 80%, 85%, 90%, 95%, 98%, 99%, 75-99.5%, 85-99.5%, or 90-99.8% in a subject as determined by an assay described herein or known to one of skill in the art. Accordingly, the topical vaccine formulations are useful in methods of preventing, treating and/or managing drug addiction or drug dependence. In some embodiment, the topical vaccine formulations is useful in preventing, treating and/or managing a disease or condition that exhibits resistance to other treatments.
[0139] In certain embodiments, the topical vaccine formulation inhibits or reduces the circulation in the bloodstream a drug or antigen against which it is targeted by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85% as measured by a standard assay known to one of skill in the art, or an assay described herein.
[0140] In some embodiments, the topical vaccine formulation inhibits or reduces the penetration of the drug or antigen against which it is targeted from one organ, tissue or cell to another organ, tissue or cell as measured using a standard assay known to one of skill in the art, or an assay described herein. In some embodiments, the topical vaccine formulation inhibits or reduces the ability of a drug, such as cocaine or nicotine, to enter the brain by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85% as measured using a standard assay known to one of skill in the art, or an assay described herein.
4.5.4 Prophylactic and Therapeutic Methods
[0141] The present invention provides methods of preventing, treating and/or managing a disease or condition, such as a drug addiction, said methods comprising administering to a subject in need thereof one or more topical vaccine formulations. In one embodiment, the invention provides a method of preventing, treating/and or managing a cocaine addiction or a nicotine addiction.
[0142] The invention also provides methods of preventing, treating and/or managing a disease or condition, said methods comprising administering to a subject in need thereof one or more topical vaccine formulations, and one or more other therapies (e.g., prophylactic or therapeutic agents). In some embodiments, the other therapies are currently being used, have been used or are known to be useful in the prevention, treatment and/or management of the disease or condition. Non-limiting examples of such prophylactic or therapeutic methods are provided below.
[0143] The combination therapies of the invention can be administered sequentially or concurrently. In one embodiment, the combination therapies comprise administering the topical vaccine formulation at least one other therapy which has the same mechanism of action. In another embodiment, the combination therapies comprise administering the topical vaccine formulation and at least one other therapy which has a different mechanism of action than the compound.
[0144] In a specific embodiment, the combination therapies improve the prophylactic and/or therapeutic effect of an immunogenic composition of the invention by functioning together with immunogenic composition to have an additive or synergistic effect. In another embodiment, the combination therapies reduce the side effects associated with each therapy taken alone.
[0145] In some embodiments, the topical vaccine formulation further comprises the prophylactic or therapeutic agents of the combination therapies. Thus, in some embodiments of the combination therapy, the administration of the topical vaccine formulation effects administration of both the hapten-carrier conjugate and the prophylactic or therapeutic agents. Alternatively, the prophylactic or therapeutic agents of the combination therapies can be administered concurrently to a subject in separate formulations. The prophylactic or therapeutic agents may be administered to a subject by the same or different routes of administration.
[0146] In some embodiments, the topical vaccine is administered to a subject, preferably a human, to prevent, treat and/or manage a drug addiction. In a specific embodiment, the other prophylactic or therapeutic agents are currently being used, have been used or are known to be useful in the prevention, treatment and/or management of the drug addiction or a symptom or condition related to it, e.g., a psychiatric or psychological condition.
[0147] The topical vaccine formulation may be used as any line of therapy, e.g., a first, second, third, fourth or fifth line therapy, for a disease or condition. In some embodiments, the subject administered the topical vaccine formulation has not received a prior therapy. In other embodiments, the subject administered the topical vaccine formulation has received a prior therapy. In some embodiments, the subject administered the topical vaccine
formulation was refractory to a prior therapy or experienced adverse side effects to a prior therapy or a prior therapy was discontinued due to unacceptable levels of toxicity to the subject,
[0148] In some embodiment, a method is provided for treating and/or managing a disease or condition, e.g., a drug addiction, in a subject refractory to conventional therapies for such a condition, the methods comprising administering to said subject a dose of a prophylactically or therapeutically effective amount of an immunogenic composition of the invention.
4.5.5. Agents Useful in Combination with Formulations
[0149] Therapeutic or prophylactic agents that can be used in combination with the topical vaccine formulation for the prevention, treatment and/or management of a disease or condition include, but are not limited to, small molecules, synthetic drugs, peptides (including cyclic peptides), polypeptides, proteins, nucleic acids (e.g., DNA and RNA nucleotides including, but not limited to, antisense nucleotide sequences, triple helices, RNAi, and nucleotide sequences encoding biologically active proteins, polypeptides or peptides), antibodies, synthetic or natural inorganic molecules, mimetic agents, and synthetic or natural organic molecules. Specific examples of such agents include, but are not limited to, immunomodulatory agents (e.g., interferon), anti-inflammatory agents (e.g.. adrenocorticoids, corticosteroids (e.g., beclomethasone, budesonide, flunisolide, fluticasone, triamcinolone, methlyprednisolone, prednisolone, prednisone, hydrocortisone), glucocorticoids, steroids, and non-steroidal anti-inflammatory drugs (e.g., aspirin, ibuprofen, diclofenac, and COX-2 inhibitors), pain relievers, anti-psychotics. anti-depressants, anti-anxiety drugs, anti- epileptics, leukotreine antagonists (e.g., montelukast, methyl xanthines, zafirlukast, and zileuton), beta2-agonists (e.g., albuterol, biterol, fenoterol, isoetharie, metaproterenol, pirbuterol, salbutamol, terbutalin formoterol, salmeterol, and salbutamol terbutaline), anticholinergic agents (e.g., ipratropium bromide and oxitropium bromide), sulphasalazine, penicillamine, dapsone, antihistamines, anti-malarial agents (e.g., hydroxychloroquine), antiviral agents (e.g., nucleoside analogs (e.g., zidovudine, acyclovir, gangcyclovir, vidarabine, idoxuridine, trifluridine, and ribavirin), foscarnet, amantadine, rimantadine, saquinavir, indinavir, ritonavir, and AZT) and antibiotics (e.g., dactinomycin (formerly actinomycin), bleomycin, erythomycin, penicillin, mithramycin, and anthramycin (AMC)).
[0150] Any therapy which is known to be useful, or which has been used or is currently being used for the prevention, management, and/or treatment of the disease or condition, or a disease or condition associated with the disease or condition, can be used in combination with the topical vaccine formulation. See, e.g., Gilman et al., Goodman and Gilman's: The Pharmacological Basis of Therapeutics, 10th ed., McGraw-Hill, New York, 2001 ; The Merck Manual of Diagnosis and Therapy, Berkow, M.D. et al. (eds.), 17th Ed., Merck Sharp & Dohmc Research Laboratories, Rahway, NJ, 1999; Cecil Textbook of Medicine, 20th Ed., Bennett and Plum (eds.), W.B. Saunders, Philadelphia, 1996 for information regarding therapies (e.g., prophylactic or therapeutic agents) which have been or are currently being used for preventing, treating and/or managing various diseases or conditions.
[0151] In some embodiment, antibacterial agents, including antibiotics, that can be used in combination with the topical vaccine formulation include, but are not limited to, aminoglycoside antibiotics, glycopeptides, amphenicol antibiotics, ansamycin antibiotics, cephalosporins, cephamycins oxazolidinones, penicillins, quinolones, streptogamins, tetracyciins, and analogs thereof.
[0152] In some embodiments, the topical vaccine formulation is used in combination with other protein synthesis inhibitors, including but not limited to, streptomycin, neomycin, erythromycin, carbomycin, and spiramycin.
[0153] In one embodiment, the antibacterial agent is selected from the group consisting of ampicillin, amoxicillin, ciprofloxacin, gentamycin, kanamycin, neomycin, penicillin G, streptomycin, sulfanilamide, and vancomycin. In another embodiment, the antibacterial agent is selected from the group consisting of azithromycin, cefonicid, cefotetan, cephalothin, cephamycin, chlortetracycline, clarithromycin, clindamycin, cycloserine, dalfopristin.
doxycycline, erythromycin, linezolid, mupirocin, oxytetracycline, quinupristin, rifampin, spectinomycin, and trimethoprim.
[0154] Additional, non-limiting examples of antibacterial agents for use in combination with the topical vaccine formulations include the following: aminoglycoside antibiotics (e.g., apramycin, arbekacin, bambermycins, butirosin, dibekacin, neomycin, neomycin,
undecylenate, netilmicin, paromomycin, ribostamycin, sisomicin, and spectinomycin), amphenicol antibiotics (e.g., azidamfenicol, chloramphenicol, florfenicol, and
thiamphenicol), ansamycin antibiotics (e.g., rifamide and rifampin), carbacephems (e.g., loracarbef), carbapenems (e.g., biapenem and imipenem), cephalosporins (e.g., cefaclor, cefadroxil, cefamandole, cefatrizine, cefazedone, cefozopran, cefpimizole, cefpiramide, and cefpirome), cephamycins (e.g., cefbuperazone, cefmetazole, and cefminox), folic acid analogs (e.g., trimethoprim), glycopeptides (e.g., vancomycin), lincosamides (e.g., clindamycin, and lincomycin), macrolides (e.g., azithromycin, carbomycin, clarithomycin, dirithromycin, erythromycin, and erythromycin acistrate), monobactams (e.g., aztreonam, carumonam, and tigemonam), nitrofurans (e.g., furaltadone, and furazolium chloride), oxacephems (e.g., flomoxef, and moxalactam), oxazolidinones (e.g., linezolid), penicillins (e.g., amdinocillin, amdinocillin pivoxil, amoxicillin, bacampicillin, benzylpenicillinic acid, benzylpenicillin sodium, epicillin, fenbenicillin, floxacillin, penamecillin, penethamate hydriodide, penicillin o benethamine, penicillin 0, penicillin V, penicillin V benzathine, penicillin V hydrabamine, penimepicycline, and phencihicillin potassium), quinolones and analogs thereof (e.g., cinoxacin, ciprofloxacin, clinafloxacin, flumequine, grepagloxacin, levofloxacin, and moxifloxacin), streptogramins (e.g., quinupristin and dalfopristin), sulfonamides (e.g., acetyl sulfamethoxypyrazine, benzylsulfamide, noprylsulfamide, phthalylsulfacctamide, sulfachrysoidine, and sulfacytine), sulfones (e.g., diathymosulfone, glucosulfonc sodium, and solasulfone), and tetracyclines (e.g., apicycline, chlortetracycline, clomocycline, and demeclocycline). Additional examples include cycloserine, mupirocin, tuberin amphomycin, bacitracin, capreomycin, colistin, enduracidin, enviomycin, and 2,4 diaminopyrimidines (e.g., brodimoprim).
[0155] m some embodiments, antiviral agents that can be used in combination with In one embodiment, the antibacterial agent is selected from the group consisting of ampicillin, amoxicillin, ciprofloxacin, gentamycin, kanamycin, neomycin, penicillin G, streptomycin, sulfanilamide, and vancomycin. In another embodiment, the antibacterial agent is selected from the group consisting of azithromycin, cefonicid, cefotetan, cephalothin, cephamycin, chlortetracycline, clarithromycin, clindamycin, cycloserine, dalfopristin, doxycycline, erythromycin, linezolid, mupirocin, oxytetracycline, quinupristin, rifampin, spectinomycin, and trimethoprim.
[0156] Antiviral agents include, but are not limited to, non-nucleoside reverse transcriptase inhibitors, nucleoside reverse transcriptase inhibitors, protease inhibitors, and fusion inhibitors, In one embodiment, the antiviral agent is selected from the group consisting of amantadine, oseltamivir phosphate, rimantadine, and zanamivir. In another embodiment, the antiviral agent is a non-nucleoside reverse transcriptase inhibitor selected from the group consisting of delavirdine, efavirenz, and nevirapine. In another embodiment, the antiviral agent is a nucleoside reverse transcriptase inhibitor selected from the group consisting of abacavir, didanosine, emtricitabine, emtricitabine, lamivudine, stavudine, tenofovir DF, zalcitabine, and zidovudine. In another embodiment, the antiviral agent is a protease inhibitor selected from the group consisting of amprenavir, atazanavir, fosamprenav, indinavir, lopinavir, nelfinavir, ritonavir, and saquinavir. In another embodiment, the antiviral agent is a fusion inhibitor such as enfuvirtide. Additional, non-limiting examples of antiviral agents for use in combination with an immunogenic composition of the invention include the following: rifampicin, nucleoside reverse transcriptase inhibitors (e.g., AZT, ddT, ddC, 3TC, d4T), non-nucleoside reverse transcriptase inhibitors (e.g., dclavirdine efavirenz, nevirapine), protease inhibitors (e.g., aprenavir, indinavir, ritonavir, and saquinavir), idoxuridine, cidofovir, acyclovir, ganciclovir, zanamivir, amantadine, and palivizumab.
Other examples of anti-viral agents include but are not limited to acemannan; acyclovir; acyclovir sodium; adefovir; alovudine: alvircept sudotox; amantadine hydrochloride
(SYMMETRELTM); aranotin; arildone; atevirdine mesylate; avridine; cidofovir;
cipamfylline; cytarabine hydrochloride; delavirdine mesylate; desciclovir; didanosine;
disoxaril; edoxudine; enviradene; enviroxime; famciclovir; famotine hydrochloride;
fiacitabine; fialuridine; fosarilate; foscamet sodium; fosfonet sodium; ganciclovir; ganciclovir sodium; idoxuridine; kethoxal; lamivudine; lobucavir; memotine hydrochloride; methisazone; nevirapine; oseltamivir phosphate (TAMIFLUTM); penciclovir; pirodavir; ribavirin;
rimantadine hydrochloride (FLUMADINETM); saquinavir mesylate; somantadine hydrochloride; sorivudine; statolon; stavudine; tilorone hydrochloride; trifluridine;
valacyclovir hydrochloride; vidarabine; vidarabine phosphate; vidarabine sodium phosphate; viroxime; zalcitabine; zanamivir (RELENZATM); zidovudine; and zinviroxime.
4.5.6. Dosages and Frequency
[0157] The amount of topical vaccine formulation, and the amount of hapten-carrier conjugate within the topical vaccine formulation, that will be effective in the prevention, treatment and/or management of a disease or condition can be determined by standard clinical techniques. In vitro or in vivo assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed will also depend, e.g., on the route of administration, the type of disease or condition to be treated, and the seriousness of the disease or condition, and should be decided according to the judgment of the practitioner and each patient's or subject's circumstances.
[0158] Exemplary doses include milligram or microgram amounts per kilogram of subject or sample weight (e.g., about 1 microgram per kilogram to about 500 milligrams per kilogram, about 5 micrograms per kilogram to about 100 milligrams per kilogram, or about 1 microgram per kilogram to about 50 micrograms per kilogram). In some embodiments, a daily dose is at least 50 mg. 75 mg, 100 mg, 150 mg, 250 mg, 500 mg, 750 mg, or at least 1 S-
[0159] In some embodiments, the dosage is a concentration of 0.01 to 5000 mM, 1 to 300 mM, 10 to 100 mM and 10 mM to 1 M. In another embodiment, the dosage is a
concentration of at least 5 μM, at least 10 μM, at least 50 μM, at least 100 μM, at least 500 μM, at least 1 mM, at least 5 mM, at least 10 mM, at least 50 mM, at least 100 mM, or at least 500 mM.
[0160] In one embodiment, the dosage is a concentration of 0.01 to 5000 mM, 1 to 300 mM, 10 to 100 mM and 10 mM to 1 M. In another embodiment, the dosage is a
concentration of at least 5 μM, at least 10 μM, at least 50 μM, at least 100 μM, at least 500 μM, at least 1 mM, at least 5 mM, at least 10 mM, at least 50 mM, at least 100 mM, or at least 500 mM. In a specific embodiment, the dosage is 0.25 μg/kg or more, preferably 0.5 μg/kg or more, 1 μg/kg or more, 2 μg/kg or more, 3 μg/kg or more, 4 μg/kg or more, 5 μg/kg or more, 6 μg/kg or more, 7 μg/kg or more, 8 μg/kg or more, 9 μg/kg or more, or 10 μg/kg or more. 25 μg/kg or more, preferably 50 μg/kg or more, 100 μg/kg or more, 250 μg/kg or more, 500 μg/kg or more, 1 mg/kg or more, 5 mg/kg or more, 6 mg/kg or more, 7 mg/kg or more, 8 mg/kg or more, 9 mg/kg or more, or 10 mg/kg or more of a patient's body weight.
[0161] In another embodiment, the dosage is a unit dose of 10-20 μg, 20-50 μg, 50-75 μg, 75-100 μg, 100-200 μg, 200-300 μg, 300-400 μg, 400-500 μg, 500-600 μg, 600-700 μg, 700- 800 μg, 800-900 μg, or 900-1000 μg, or more. In some embodiments, the dosage is a unit dose of 5 mg, preferably 10 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg. 350 mg, 400 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg or more. In another embodiment, the dosage is a unit dose that ranges from about 5 mg to about 100 mg, preferably about 100 mg to about 200 μg, about 150 mg to about 300 mg, about 150 mg to about 400 mg, 250 μg to about 500 mg, about 500 mg to about 800 mg, about 500 mg to about 1000 mg, or about 5 mg to about 1000 mg.
[0162] In another embodiment, a subject is administered one or more doses of a prophylactically or therapeutically effective amount of the topical vaccine formulation, wherein the prophylactically or therapeutically effective amount is not the same for each dose, in another embodiment, a subject is administered one or more doses of a
prophylactically or therapeutically effective amount of the topical vaccine formulation, wherein the dose of a prophylactically or therapeutically effective amount administered to said subject is increased by, e.g., 0.01 μg/kg, 0.02 μg/kg, 0.04 μg/kg, 0.05 μg/kg, 0.06 μg/kg, 0.08 μg/kg, 0.1 μg/kg, 0.2 μg/kg, 0.25 μg/kg, 0.5 μg/kg, 0.75 μg/kg, 1 μg/kg, 1.5 μg/kg, 2 μg/kg, 4 μg/kg, 5 μg/kg, 10 μg/kg, 15 μg/kg, 20 μg/kg, 25 μg/kg, 30 μg/kg, 35 μg/kg, 40 μg/kg. 45 μg/kg, or 50 μg/kg, as treatment progresses. In another embodiment, a subject is administered one or more doses of a prophylactic ally or therapeutically effective amount of the topical vaccine formulation, wherein the dose is decreased by. e.g., 0.01 μg/kg, 0.02 μg/kg, 0.04 μg/kg, 0.05 μg/kg, 0.06 μg/kg, 0.08 μg/kg, 0.1 μg/kg, 0.2 μg/kg, 0.25 μg/kg, 0.5 μg/kg, 0.75 μg/kg, 1 μg/kg, 1.5 μg/kg, 2 μg/kg, 4 μg/kg, 5 μg/kg, 10 μg/kg, 15 μg/kg, 20 μg/kg, 25 μg/kg, 30 μg/kg, 35 μg/kg, 40 μg/kg, 45 μg/kg, or 50 μg/kg. as treatment progresses.
[0163] In some embodiments, a subject is administered one or more doses of an effective amount of the topical vaccine formulation, wherein the effective amount inhibits or reduces the level or hapten in the body or circulating antigen by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85%. In other embodiments, a subject is administered one or more doses of an effective amount the topical vaccine formulation, wherein the effective amount inhibits or reduces the severity of the disease or condition, e.g., a drug addiction, by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%, at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%, or up to at least 85%.
[0164] In other embodiments, a subject is administered one or more doses of an effective amount of the topical vaccine formulation, wherein the effective amount inhibits or reduces nicotine or cocaine addiction by at least 20% to 25%, preferably at least 25% to 30%, at least 30% to 35%. at least 35% to 40%, at least 40% to 45%, at least 45% to 50%, at least 50% to 55%, at least 55% to 60%, at least 60% to 65%, at least 65% to 70%, at least 70% to 75%, at least 75% to 80%. or up to at least 85%.
[0165] The dosages of prophylactic or therapeutic agents other than the topical vaccine formulation which have been or are currently being used for the prevention, treatment and/or management of the disease or condition or a disease or condition related to it can be determined using references available to a clinician such as, e.g., the Physicians' Desk Reference (55th ed. 2001), Preferably, dosages lower than those which have been or are currently being used to prevent, treat and/or manage the disease or condition are utilized in combination with the topical vaccine formulation.
[0166] The above-described administration schedules are provided for illustrative purposes only and should not be considered limiting. A person of ordinary skill in the art will readily understand that all doses are within the scope of the invention.
4.5.7. Methods Of Inducing an Immune Response to a Drug of Addiction
[0167] In some embodiments, the topical vaccine formulation can be used as a
therapeutic, to treat a drug addiction or drug dependence, or as a prophylactic. In prophylactic use embodiments, the drug-conjugates or immunogenic compositions comprising them may be administered to a mammal prior to any exposure to the drag to generate anti-drug antibodies. The generated anti-drug antibodies would be present in the mammal to bind to any drug introduced subsequent to the administration of the topical vaccine formulation, and therefore minimize or prevent the chance of becoming addicted to the drug.
[0168] In some embodiments, the topical vaccine formulation can be used in a subject as vaccines. In such embodiments, the topical vaccine formulations comprising at least one hapten-carrier conjugate are capable of eliciting the production of a sufficiently high titer of antibodies specific to the hapten such that upon subsequent challenge with the hapten said antibodies are capable of reducing the addictive properties of the drug. Without being limited in any way by theory, it is believed that the immune response to the topical vaccine formulation is the formation of both anti-hapten and anti-carrier antibodies. The therapeutic level is reached when a sufficient amount of the anti-drug specific antibodies are elicited and maintained to mount a neutralizing attack on drug introduced after vaccination. The therapeutic regimens can allow for sufficient time for production of antibodies after initial vaccination and any boosting. In some embodiments, the antibody titers remain sufficiently high to provide an effective response upon subsequent exposure to the drug for about two months to about one year or more, or at least three months. The topical vaccine formulation optionally includes adjuvants.
[0169] In embodiments used in the treatment of nicotine addiction, the topical vaccine formulation comprises a hapten-carrier conjugate wherein the hapten is nicotine or a nicotine derivative, which can be used to immunize mammals, particularly humans, to elicit anti- nicotine antibodies capable of binding free drug and preventing transit of the drug to the reward system in the brain thereby abrogating addictive drug-taking behavior (e.g., smoking cigarettes). Without being limited by theory in any way, it is believed that nicotine binds to the alpha-subunit of the nicotinic acetylcholine receptors in the brain which results in an increase in dopamine release. It is thought that increased numbers of nicotinic acetylcholine receptors in the brain enhance the physiological dependence of nicotine. As discussed above in relation to cocaine, anti-nicotine antibodies would presumably limit the distribution of nicotine across the blood-brain barrier to the brain, thus reducing its pharmacological effects. Antibody intervention in the case of nicotine, however, may have some advantages over cocaine. For example, there is some level of standardization with nicotine delivery; that is, each cigarette contains on average 9 mg of nicotine of which 1-3 mg are effectively dispensed during smoking. Additionally, the peak plasma concentration of nicotine is 25-50 ng/ml which is significantly lower than that of cocaine (0.3-1 μg/ml). This should provide an ideal opportunity for intervention with moderately high affinity antibodies.
[0170] In some embodiments, initial vaccination with the topical vaccine formulations creates high titers of hapten-specific antibodies in vivo. Periodic tests of the vaccinated subjects plasma are useful to determine individual effective doses. Titer levels are increased and maintained through periodic boosting. It is anticipated that this therapeutic will be used in combination with current drug rehabilitation programs, including counseling. Further, the therapeutic compositions of the present invention may be aimed at a single drug or several drugs simultaneously or in succession and may be used in combination with other therapies. For example, in some embodiments, the topical vaccine formulation can be used without adverse interactions in combination with conventional pharmacological approaches and ""short term"" passive immunization to enhance the overall effect of therapy.
[0171] Without being limited by theory in any way, it is believed that the hapten-carrier conjugate, which is prepared by coupling one or more hapten molecules to a T cell epitope containing carrier to obtain a hapten-carrier conjugate capable of stimulating T cells
(immunogenic), leads to T cell proliferation and a characteristic release of mediators which activate relevant B cells and stimulate specific antibody production.
[0172] In selecting a drug suitable for conjugation according to the instant invention, one skilled in the art would select drug with properties likely to elicit high antibody titers.
However, if the chosen molecule is similar to those molecules which are endogenous to the individual, antibodies raised against such a molecule could cross-react with many different molecules in the body giving an undesired effect. Thus, the drug to be selected as the hapten (drug/haptenj must be sufficiently foreign and of a sufficient size so as to avoid eliciting antibodies to molecules commonly found inside a human body. For these reasons, alcohol, for example, would not be suitable for the therapeutic of the instant invention. The antibodies raised against the therapeutic composition are highly specific and of a sufficient quantity to neutralize the drug either in the blood stream or in the mucosa or both. Without limiting the invention, the drugs which are suitable for therapeutic composition (not in order of importance) are, for example, hallucinogens, for example mescaline and LSD; Cannabinoids, for example THC; Stimulants, for example amphetamines, cocaine, phenmetrazine, methylphenidate; Nicotine; Depressants, for example, nonbarbiturates (e.g. bromides, chloral hydrate etc.), methaqualone. barbiturates, diazepam, flurazepam, phencyclidine, and fluoxetine; Opium and its derivatives, for example, heroin, methadone, morphine, meperidine, codeine, pentazocine, and propoxyphene; and "'"Designer drugs'"' such as ""ecstasy"".
4.6. KITS
[0173] In some embodiments is provided a pharmaceutical pack or kit comprising one or more containers filled with the topical vaccine formulation. The kits can be used in the above-described methods. In particular, the kits can be used for the prevention, treatment, and/or management of a disease or condition, e.g., a drug addiction (e.g., to cocaine or nicotine).
[0174] In some embodiment, the kit comprises a topical vaccine formulation or an antibody raised in response to the formulation, in one or more containers. In another embodiment, a kit comprises a topical vaccine formulation or an antibody raised in response to the formulation, in one or more containers, and one or more other prophylactic or therapeutic agents, in one or more other containers. In a particular embodiment, the kit further comprises instructions for using the the topical vaccine formulation or the antibody raised in response to the formulation, as well as an explanation of side effects of the topical vaccine formulation or the antibody raised in response to the formulation, and dosage information for a particular route of administration. Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration. 5. EXAMPLES
5.1 Example 1: Topical Vaccine Formulations
[0175] The topical vaccine formulations may be prepared by the following general procedure:
1. Organic phase production, which contains all lipophilic excipients
The organic phase is produced by weighing the lipid, the surfactant, and any additional lipophilic excipients into suitable containers followed by mixing these components into anoptically isotropic phase which appears as a clear solution, where the hapten-carrier conjugate is as described above, and is preferably a nicotine-carrier conjugate or a cocaine- carrier conjugate. During mixing, the organic phase will be heated up to a temperature of about 5 to about 60 °C.
2. Aqueous phase production
The aqueous phase is prepared by weighing the non-lipophilic components, including the hapten-carrier conjugate, and water, which serves as solvent, into suitable containers and then mixing these components into a clear solution. During mixing, the temperature will be elevated to about 5 to about 60 °C.
3. Production of a concentrated intermediate by combination of both phases
The isotropic organic phase and the clear aqueous phase are combined under stirring in a suitable vessel. Before and during the combination the temperature of both phases must be kept between about 5 to about 60 °C or between about 35 and about 45 °C. The resulting intermediate is homogenised mechanically at a temperature of about 5 to about 60 °C, e.g., about 40 °C. Before starting homogenisation, the pressure in the production vessel is lowered to - 0.08 MPa. The desired average carrier size is typically reached after 10 minutes of homogenisation.
Three process parameters must be controlled carefully during the production of the concentrated intermediate: temperature, homogeniser circulation velocity, and overall processing time.
4. Production of the final bulk product by mixing the concentrated intermediate with dilution buffer. The concentrated intermediate is diluted with the dilution buffer to the intended final concentration. The mixture is carefully stirred in the mixing vessel at 20 °C to homogeneity.
[0176] Table 8 describes the amount of surfactant, lipid, hapten-carrier conjugate, and other excipients for some topical vaccine formulations provided herein. The amount of the hapten-carrier conjugate, lipid, surfactant, and other excipients is described in terms of the percent total in the formulation.
[0177] Hapten-Carrier Conjugate: 0.1 - 50 mg/g (nicotine- and cocaine-, which specific amounts besides 1 mg/g?)
Surfactant: 1 to 10% by weight (Sphingosylphosphorylcholine, phosphatidyl choline, sphingomyelin, phosphatidyl glycerol, any others? Specific amounts?)
Lipid: 1 to 10% by weight (T ween 80, Brij 98, Cremophor, Span 20, any others? Specific amounts?)
Buffer: pH 3 - 9.5 (acetate, phosphate, lactate)
Antimicrobials: 0 - 10 mg/g (benzyl alcohol + methyl parabene, each alone, others? E.g., benzalkonium chloride?)
Antioxidants: 0 - 10 mg/g (include any here?)
Emollients: 0 - 50 mg/g (glycerol, mannitol, others? Specific amounts besides 6.25 g/mg?)
Chelators: 0 - 25 mg/g (EDTA, others?)
Other: 0 - 50 mg/g (ethanol, others?)
[0178] In a particular embodiment, the topical vaccine formulation comprises 1 mg/g TA- NIC as the hapten-carrier conjugate, 64.516 mg/g sphingosylphosphorylcholine, 35.484 mg/g Tween 80 , 6.25 mg/g mannitol, 5.25 mg/g benzyl alcohol, 2.5 mg/g methyl parabene. 3 mg/g EDTΛ, 1 1.25 mM phosphate buffer at pH 6.3, 30 mg/g ethanol, and 851.774 mg/g water.
Figure imgf000062_0001
Figure imgf000063_0001
Figure imgf000064_0001
Figure imgf000065_0001
Figure imgf000066_0001
Figure imgf000067_0001
Figure imgf000068_0001
Figure imgf000069_0001
Figure imgf000070_0001

Claims

WHAT IS CLAIMED IS:
1. An immunogenic composition comprising nicotine conjugated to a carrier, a lipid and a non-ionic surfactant.
2. The immunogenic composition of claim 1 , wherein the carrier is a bacterial toxin.
3. The immunogenic composition of claim 2, wherein the bacterial toxin is derived from cholera toxin B, pseudomonas exotoxin, diphtheria toxin, tetanus toxoid, shiga toxin, and pertussis toxin.
4. The immunogenic composition of claim 1 formulated for topical delivery.
5. The immunogenic composition of claim 1 formulated for transmucosal delivery.
6. The immunogenic composition of claim 1 wherein the nicotine conjugate is present at a dose from 3.0 to 50 mg/g.
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