Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K16/00—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
  • C07K16/16—Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies against material from plants
• • 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
  • A61K39/00—Medicinal preparations containing antigens or antibodies
  • A61K39/385—Haptens or antigens, bound to carriers
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P25/00—Drugs for disorders of the nervous system
  • A61P25/30—Drugs for disorders of the nervous system for treating abuse or dependence
  • A61P25/34—Tobacco-abuse
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
  • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
  • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K2317/00—Immunoglobulins specific features
  • C07K2317/90—Immunoglobulins specific features characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
  • C07K2317/92—Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value

Definitions

• Nicotine plays a central role in precipitating addiction to smoking tobacco.
• a nicotine vaccine stimulates the immune system to identify nicotine as a foreign antigen, eliciting antibodies that alter nicotine pharmacokinetics.
• Anti- nicotine antibodies reduce the concentration of free nicotine in the blood and prevent it from entering the central nervous system. Blocking the activation of brain reward systems can facilitate extinction of the addictive behavior, leading to better smoking cessation outcomes.
• a clinically approved nicotine vaccine would be a complementary addition to the available tools, which, when leveraged appropriately, could afford significantly better rates of sustained smoking abstinence.
• NicVAX ® represents the most clinically advanced nicotine vaccine to date, having progressed all the way through Phase 111.(8-1 1) It was safe and well tolerated, but was effective for only a fraction of clinical trial participants. (12, 13) Nevertheless, given the huge promise of a clinically approved nicotine vaccine, research continues unmitigated. Many design and formulation aspects have been scrutinized in recent years to furnish something better then NicVAX ® . Efforts include boosting immunogenicity through the use of newer adjuvants(14- 17), improving practicality through alternative routes of administration(18), and adopting multivalent strategies( 19-22) to increase anti-nicotine antibody binding capacity.
• the invention is directed, in various embodiments, to haptens suitable for use in raising antibodies to (-)-nicotine; to antigens comprising the haptens; to antibodies prepared using the antigens comprising the haptens; to vaccines comprising the anti-nicotine antigens; and to methods of treatment of tobacco habituation or addiction in patients comprising the use of anti-nicotine vaccines / antigens.
• the haptens in various embodiments, have sufficient structural similarity to the alkaloid (-)-nicotine, the major drug component of tobacco (Nicotiana), such that antibodies raised, e.g., in a human patient, against antigens comprising the haptens of the invention also react with (-)-nicotine, and thus the antigens can be suitable for administration to patients for raising anti-nicotine antibodies by means of the patient's immune system.
• Such antibodies can serve to bind nicotine, such as from smoked or chewed forms of tobacco, and block the drug effect on the patient that serves to reinforce the drug addiction in the patient.
• the antibodies generated by use of the antigenic vaccines of the invention can be selective for (-)-nicotine (i.e., (S)-nicotine).
• the invention provides a hapten of formula (-)- 3'-AmNic (-)-3'-AmNic; a conjugated hapten comprising (-)-3'-AmNic wherein the (-)-3'-AmNic hapten is covalently bonded via a linker to a carrier protein; an antigen for
• administering comprising administering an effective amount of the conjugated hapten or an antigenic mixture comprising the conjugated hapten to a patient suffering from the addiction or habituation, such that antibodies are produced in the patient having affinity for (-)-nicotine.
• the invention provides a (-)-nicotine hapten of formula (-)-N4N
• a conjugated hapten comprising (-)-N4N wherein the (-)-N4N hapten is covalently bonded via a linker to a carrier protein; an antigen for administration to a patient, comprising the conjugated hapten and optionally adjuvant(s); an antiserum produced in a patient comprising antibodies having immunological affinity for (-)-nicotine; and a method of treatment of nicotine or tobacco addiction or habituation comprising administering an effective amount of the conjugated hapten or an antigenic mixture comprising the conjugated hapten to a patient suffering from the addiction or habituation, such that antibodies are produced in the patient having affinity for (-)-nicotine.
• Figure 1 shows a synthetic protocol for the preparation of (-)-3'-AmNic and (+)- 3'-AmNic, which were then conjugated via a succinate linker to tetanus toxin ("TT") and evaluated for their specificity towards (-)-nicotine.
• TT tetanus toxin
• Figure 2 shows a synthetic protocol for the preparation and conjugation of (-)- N4N.
• FIG 4 shows binding affinity curves for radioimmunoassays (RIA) (-)-3'- AmNic and (+)-3'-AmNic.
• hapten-carrier conjugates and antigen compositions specifically and selectively target only (-)-nicotine; thus, in accordance with the invention, a vaccine according to embodiments described herein efficiently elicit antibodies capable of sequestering only (-)-nicotine.
• the notion that antibodies can enantiodifferentiate was first appreciated by Landsteiner nearly a century ago(28, 29) and continues to be exploited to this day.
• Such work includes enantioselective catalytic antibodies (30-32) and stereospecific mAb to nicotine(33) and cocaine.
• hybridomas were selected using (S)-(-)-[3H]nicotine, thereby optimizing for antibodies specific for the naturally occurring isomer.
• the present invention exploits a capacity of antibodies to enantiodifferentiate in developing vaccines for nicotine.
• hapten as used in the present invention is a low-molecular weight organic compound that, by itself, is incapable of eliciting an immune response. However, it will elicit an immune response once attached to a carrier molecule. According to some embodiments, the hapten is attached to the carrier via a linker.
• a hapten of the present invention is a nicotine derivative that is ?raTO-3'-aminomethylnicotine (3'AmNic).
• the hapten is a single enantiomer of ?ra « -3'-aminomethylnicotine, specifically (-)-3'-AmNic. Both enantiomers are shown below:
• the hapten is (S)-3-(3-(l-Methylpyrrolidin-2- yl)pyridin-4-yl)propan- 1 -amine, (-)-N4N, as shown below:
• methylene C3 ⁇ 4 attached to C4- position
• a heteroatom e.g., oxygen
• both steric and stereoelectronic considerations are made, with the goal of eliciting a better anti-nicotine antibody response (higher titers or concentrations, higher affinity and specificity, superior functional antagonism of nicotine's pharmacokinetics and pharmacodynamics) by virtue of this novel nicotine hapten design.
• a nicotine hapten is directly attached to a carrier with or without a linker.
• a single nicotine hapten can be attached to each available amine group on a carrier protein.
• General methods for directly conjugating haptens to carrier proteins, using a homobifunctional or a heterobifunctional cross-linker are well known in the art, for example, by G. T. Hermanson in Bioconjugate Techniques, Academic Press (1996) and Dick and Beurret in Conjugate Vaccines. Contribu. Microbiol. Immunol., Karger, Basal (1989) vol. 10, 48- 114.
• Direct conjugation using bifunctional crosslinkers generally results in a molar ratio of hapten to protein being limited by the number of functional groups available on the protein for the specific conjugation chemistry.
• a carrier protein possessing n number of lysine moieties theoretically presents n+ ⁇ primary amines (including the terminal amino) available for reaction with a linker carboxyl group.
• direct conjugation gives rise to formation of n+ ⁇ amido bonds, i.e., a maximum of n+ ⁇ haptens attached.
• conjugated hapten density can depend upon concentration of the reactants used to conjugate the nicotine hapten to the carrier protein, and the nature of the carrier protein.
• hapten density (molar ratio of conjugated hapten to protein carrier) can range from about 10 to about 70, from about 20 to about 60, and from about 30 to about 50.
• the haptent is then conjugated to a carrier protein which will be used to raise antibodies to the nicotine carrier conjugate.
• the carrier protein of the present invention generally is any suitable immunogenic protein or polypeptide.
• An "immunogenic" molecule is one that is capable of eliciting an immune response.
• the carrier protein is a T-cell epitope.
• the "carrier protein” is a multi-antigenic peptide (MAP), which is a branched peptide.
• MAP multi-antigenic peptide
• hapten density and valency are maximized because of multiple branched amino acid residues.
• amino acids that can be used to form a MAP include, but are not limited to, lysine.
• a carrier protein comprises a molecule containing at least one T cell epitope which is capable of stimulating the T cells of the subject, which subsequently induces B cells to produce antibodies against the entire hapten-carrier conjugate molecule.
• epitope as used herein includes any determinant on an antigen that is responsible for its specific interaction with an antibody molecule. Epitopic determinants usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and have specific three dimensional structural characteristics as well as specific charge characteristics.
• a carrier protein is selected based upon its ability to elicit a strong immunogenic response so that a diverse population of patients can be treated by the inventive hapten-carrier conjugates.
• the carrier protein must be sufficiently foreign to elicit a strong immune response to the vaccine.
• the carrier protein in this regard is a large molecule capable of imparting immunogenicity to a covalently- linked hapten.
• the carrier is one that is used in the preparation of therapeutic conjugate vaccines, such as a number of toxins of pathogenic bacteria and their toxoids. Examples include diphtheria and tetanus toxins and their medically acceptable
• the carrier is bovine serum albumin (BSA) or keyhole limpet hemocyanin (KLH), both of which are commonly used as carriers in the development of conjugate vaccines when experimenting with animals.
• the protein is flagellar filament structural protein (FliC).
• the carrier is a protein that is antigenically similar to bacterial toxins, often referred to as a cross-reacting materials (CRM).
• CCM cross-reacting materials
• nicotine hapten-carrier conjugates of the present invention are prepared by reacting one or more haptens with a carrier protein to yield a hapten carrier conjugate.
• a variety of functional groups are used to facilitate the linking or conjugation of a carrier to a hapten of the present invention. These include functional moieties such as carboxylic acids, anhydrides, mixed anhydrides, acyl halides, acyl azides, alkyl halides, N-maleimides, imino esters, isocyanates, amines, thiols, and isothiocyanates and others that are capable of forming a covalent bond with a reactive group of a protein molecule.
• functional moieties such as carboxylic acids, anhydrides, mixed anhydrides, acyl halides, acyl azides, alkyl halides, N-maleimides, imino esters, isocyanates, amines, thiols, and isothiocyanates and others that are capable of forming a covalent bond with a reactive group of a protein molecule.
• the reactive group is the amino group of a lysine residue or a thiol group on a carrier protein or a modified carrier protein molecule which, when reacted, results in amide, amine, thioether, amidine urea or thiourea bond formation.
• Other suitable activating groups and conjugation techniques are well known in the art (Wong, Chemistry of Protein Conjugation and Cross-Linking, CRC Press, Inc. (1991); Hermanson, BIOCONJUGATE TECHNIQUES, Academic Press: 1996; and Dick and Beurret in Conjugate Vaccines. Contribu. Microbiol. Immunol., Karger, Basal (1989) vol. 10, 48-1 14.)
• the linker is a linear moiety for conjugation of haptens to carrier proteins.
• the linker is a succinyl moiety.
• ADH adipic acid dihydrazide
• the antiserum of the present invention comprises antibodies that are produced in response to an antigen, which itself comprises a hapten-carrier conjugate as described herein.
• an antigen which itself comprises a hapten-carrier conjugate as described herein.
• techniques for making monoclonal antibodies are well-known in the art.
• monoclonal antibodies can be obtained by injecting mice with a composition comprising the nicotine hapten-carrier conjugate, subsequently verifying the presence of antibody production by removing a serum sample, removing the spleen to obtain B-lymphocytes, fusing the B-lymphocytes with myeloma cells to produce hybridomas, cloning the hybridomas, selecting positive clones which produce antibodies to the hapten-carrier conjugate, culturing the clones that produce antibodies to the antigen, and isolating the antibodies from the hybridoma cultures.
• monoclonal antibodies can be isolated and purified from hybridoma cultures by a variety of well-established techniques.
• the techniques include affinity chromatography with Protein-A Sepharose, size-exclusion chromatography, and ion-exchange chromatography (Coligan at pages 2.7.1- 2.7.12 and pages 2.9.1-2.9.3; Baines et al., "Purification of Immunoglobulin G (IgG),” in METHODS IN MOLECULAR BIOLOGY, VOL. 10, pages 79-104 (The Humana Press, Inc. 1992).
• polyclonal antibodies also are well-known in the art.
• an animal is injected with immunogenic material and then antibody rich serum is collected which contains therein a mixture of antibodies that are directed against numerous epitopes of the immunogen that was injected.
• Suitable host mammals for the production of antibodies include, but are not limited to, humans, rats, mice, rabbits, and goats.
• fragments also can be utilized.
• the fragments are produced by methods that include digestion with enzymes such as pepsin or papain and/or cleavage of disulfide bonds by chemical reduction.
• antibody fragments encompassed by the present invention can be synthesized using an automated peptide synthesizer such as those supplied commercially by Applied Biosystems, Multiple Peptide Systems and others, or they may be produced manually, using techniques well known in the art (Geysen et al., J. Immunol. Methods 102: 259 (1978)). Direct determination of the amino acid sequences of the variable regions of the heavy and light chains of the monoclonal antibodies according to the invention can be carried out using conventional techniques.
• a fragment according to some embodiments of the present invention is an Fv fragment.
• An Fv fragment of an antibody is made up of the variable region of the heavy chain (Vh) of an antibody and the variable region of the light chain of an antibody (VI). Proteolytic cleavage of an antibody can produce double chain Fv fragments in which the Vh and VI regions remain non- covalently associated and retain antigen binding capacity.
• Fv fragments also include recombinant single chain antibody molecules in which the light and heavy chain variable regions are connected by a peptide linker (Skerra, et al. Science, 240, 1038-41 (1988)).
• Antibody fragments according to other embodiments of invention include Fab, Fab', F(ab)2, and F(ab')2, which lack the Fc fragment of an intact antibody.
• the present invention provides therapeutic methods and uses for preventing nicotine from crossing the blood brain barrier.
• administration of a nicotine hapten-carrier conjugate to a patient generates antibodies against nicotine in the bloodstream of the patient.
• anti-nicotine antibodies generated in a suitable host mammal and outside the body of the patient to be treated can be administered to a patient. If the patient smokes, the nicotine in his blood will be bound by the circulating anti-nicotine antibodies, preventing the nicotine from reaching the brain. Therefore, the antibodies prevent the physiological and psychological effects of nicotine that originate in the brain. Because the smoker will experience a lessening or cessation of these effects, he/she will lose the desire to smoke. The same therapeutic effects result if a patient uses smokeless tobacco, after being immunized with a nicotine hapten-carrier conjugate of the invention. Additionally, the conjugates and antibodies of the invention exert their effects by affecting the ability of nicotine to stimulate the peripheral nervous system.
• the conjugates of the invention are suitable for treating and preventing nicotine addiction.
• a nicotine-carrier conjugate of the invention is administered to a patient suffering from nicotine addiction.
• patients at risk for developing nicotine addiction, such as teenagers are treated with a conjugate according to the invention.
• Direct administration of the conjugate to a patient is called "active immunization.”
• a vaccine composition of the present invention comprises at least one nicotine hapten-carrier conjugate in an amount sufficient to elicit an immune response thereto.
• the nicotine hapten carrier conjugate is capable of remaining in vivo at a concentration sufficient to be active against subsequent intake of nicotine.
• Initial vaccination with the nicotine hapten carrier conjugate of the present invention creates high titers of antibodies that are specific to nicotine.
• the therapeutically effective amount of a conjugate which is administered to a patient in need of treatment for nicotine addiction is readily determined by the skilled artisan. Suitable dosage ranges are 1-1000 ⁇ g/dose. It generally takes a patient one to several weeks to generate antibodies against a foreign antigen.
• the production of antibodies in a patient's blood can be monitored by using techniques that are well-known to the skilled artisan, such as ELISA, radioimmunoassay (RIA), and Western blotting methods.
• Therapeutic effectiveness also can be monitored by assessing various physical effects of nicotine, such as blood pressure.
• the inventive nicotine hapten-carrier conjugates can be processed to afford a composition that is administered to a patient.
• modes of administration include but are not limited to intranasal, intratracheal, oral, dermal, transmucosal subcutaneous injection and intravenous injection.
• the skilled artisan will recognize that the initial injection may be followed by subsequent administration of one or more "boosters" of conjugate.
• the booster increases the production of antibodies against the nicotine hapten-carrier conjugate of the invention.
• the vaccine or antiserum compositions of the present invention comprises at least one adjuvant.
• the adjuvant is selected so that the effect of the carrier protein is not inhibited.
• Adjuvants those which are physiologically acceptable to humans; these include, but are not limited to, alum, QS-21, saponin and MPLA (monophosphoryl lipid A).
• the vaccine compositions according to other embodiments optionally comprise one or more pharmaceutically acceptable excipients.
• the excipients include one or more of sterile water, salt solutions such as saline, sodium phosphate, sodium chloride, alcohol, gum arabic, vegetable oils, benzyl alcohols, polyethylene glycol, gelatin, mannitol, carbohydrates, magnesium stearate, viscous paraffin, fatty acid esters, hydroxy methyl cellulose and buffers. Any additional excipients known to the skilled artisan are useful in the present invention.
• the hapten-carrier conjugates of the present invention are incorporated into a pharmaceutical composition for administering to a patient in need of treatment or prevention of nicotine addiction.
• a pharmaceutical composition for administering to a patient in need of treatment or prevention of nicotine addiction.
• the composition containing the hapten-carrier conjugate is to be used for injection, for instance, the hapten- carrier conjugate is solubilized in an aqueous, saline solution at a pharmaceutically acceptable pH.
• an injectable suspension of the hapten-carrier conjugate is possible to use.
• the composition can contain optional components to ensure purity, enhance bioavailability and/or increase penetration.
• the vaccine composition optionally contains at least one auxiliary agent, such as dispersion media, coatings, microspheres, liposomes, microcapsules, lipids, surfactants, lubricants, preservatives and stabilizers. Any additional auxiliary agents known to the skilled artisan are useful in the present invention. Also useful herein are any agents which act to synergize the effect of the present vaccine composition.
• the pharmaceutical composition of the present invention is sterile and is sufficiently stable to withstand storage, distribution, and use. Additionally, the composition may contain additional components in order to protect the composition from infestation with, and growth of, microorganisms.
• the composition is manufactured in the form of a lyophilized powder that is reconstituted by a pharmaceutically acceptable diluent just prior to administration.
• Methods of preparing sterile injectable solutions are well known to the skilled artisan and include, but are not limited to, vacuum drying, freeze- drying, and spin drying. These techniques yield a powder of the active ingredient along with any additional excipient incorporated into the pre-mix.
• Passive immunization comprises administration of or exposure to a polyclonal antibody or monoclonal antibody which has been raised in response to a nicotine hapten carrier conjugate of the invention.
• Such antibodies can be generated in animals or humans.
• Antibodies raised in response to a nicotine conjugate of the invention can be administered to prevent addiction to nicotine.
• such antibodies can be administered to people considered to be at risk for developing addiction to nicotine, such as teenagers.
• Antibodies also are suitable for treating a patient addicted to nicotine.
• the antibodies bind nicotine in the blood, and prevent nicotine from crossing the blood brain barrier.
• antibodies raised by administration of the inventive hapten-carrier conjugate have a molecular weight range of from about 150 kDa to about 1,000 kDa.
• a therapeutically effective amount of a therapeutic antibody of the invention which is administered to a patient in need of treatment for nicotine addiction is readily determined by the skilled artisan. Suitable dosage ranges are 1- 1000 ⁇ g/dose.
• a therapeutic composition according to some embodiments of the present invention comprises at least one antibody produced in response to a nicotine-carrier conjugate of the invention.
• the compositions optionally contain one or more pharmaceutically acceptable excipients.
• Useful excipients include sterile water, salt solutions such as saline, sodium phosphate, sodium chloride, alcohol, gum arabic, vegetable oils, benzyl alcohols, polyethylene glycol, gelatin, mannitol, carbohydrates, magnesium stearate, viscous paraffin, fatty acid esters, hydroxy methyl cellulose and buffers. Any additional excipients known to the skilled artisan are useful in the present invention.
• the antibodies of the present invention in order to be administered to a patient in need of treatment or prevention of nicotine addiction, are incorporated into a pharmaceutical composition.
• the composition comprising an antibody can be formulated in an aqueous, saline solution at a pharmaceutically acceptable pH for injection. However, it is possible to use an injectable suspension of the antibody.
• the composition contains optional components to ensure purity, enhance bioavailability and/or increase penetration.
• a pharmaceutical composition comprising an antibody of the present invention is sterile and is sufficiently stable to withstand storage, distribution, and use. Additionally, the composition optionally contains additional components in order to protect the composition from infestation with, and growth of, microorganisms.
• Methods of preparing sterile injectable solutions are well known to the skilled artisan and include, but are not limited to, vacuum drying, freeze-drying, and spin drying. These techniques yield a powder of the active ingredient along with any additional excipient incorporated into the pre- mix.
• High-resolution mass spectra were recorded on an Agilent LC/MSD TOF mass spectrometer by electrospray ionization time-of-flight reflectron experiments.
• IR spectra were recorded on a Thermo Scientific Nicolet 380 FTIR spectrometer.
• CpG ODN cytosine-phosphorothioate-guanine oligodeoxynucleotide
• Vaccines prepared in this manner contained 50 ⁇ g of conjugate, 50 ⁇ g of CpG, and 20 ⁇ ⁇ of Alhydrogel per 100 ⁇ ⁇ of complete formulation.
• racemic ?ra « -3'-aminomethylnicotine (3'-AmNic, ( ⁇ )-2) was prepared from commercially available racemic iran -cotininecarboxylic acid (( ⁇ )-l).
• SFC chiral supercritical fluid chromatography
• Example 2(A) and 2(B) four hapten-protein conjugates ((-)-N4N-SucFliC, (-)-N4N-SucTT, (-)-3'-AmNic-SucFliC, and (-)- 3'-AmNic-SucTT) were prepared and then evaluated as vaccine immunogens for eliciting anti-nicotine antibodies. Each conjugate was formulated with
• ELISA and cross-reactive ELISA were carried out using either (-)- or (+)-3'-AmNicSucBSA, prepared in a manner analogous to the TT conjugates described above.
• plasma samples were run against their respective haptens: rat plasma from the (-)-3'-AmNicSucTT group was assayed on (-)-3'- AmNicSucBSA plates, and rat plasma from the (+)-3'-AmNicSucTT group was assayed on (+)-3'-AmNicSucBSA plates.
• rat plasma from the (-)-3'-AmNicSucTT group was assayed on (+)-3'- AmNicSucBSA plates
• rat plasma from the (+)-3'-AmNicSucTT group was assayed on (-)-3'-AmNicSucBSA plates.
• n 12 per group, mid-point titers
• Radioimmunoassay provides a means for determining the average binding affinity and average antibody concentration for a soluble ligand.
• the ligand is soluble and free to associate/dissociate in the analysis milieu, it offers significant advantage over ELISA, in which the ligand is immobilized on the plate surface, not to mention conjugated to a carrier protein (e.g., BSA).
• a carrier protein e.g., BSA
• the equilibrium environment simulated in an RIA experiment much more closely mimics that of free nicotine distributed in the blood and brain during tobacco use. It behooves researchers in the field to routinely incorporate RIA to evaluate the immunogenic efficacy of drug of abuse vaccine
• Nicotine-specific plasma antibody binding affinities and antibody concentrations were determined by competitive (RIA) using an adaptation of the procedure described by Muller.(46) First, the plasma dilution that binds -50% of 3H-labeled nicotine was determined. Then, the affinity constant was calculated by competition with unlabeled nicotine. Because plasma samples were pooled for each vaccine group described in Example 5 above, the measured affinity constants are average affinities for each group.
• the (-)-group gave rise to superior ELISA titers and RIA antibody concentrations with a roughly 4-fold difference observed throughout.
• the aim is to optimize protein design for a given ligand target ("antibody efficiency”); the inverse is widely applied in medicinal chemistry: optimizing a ligand design for a given protein target (e.g., "ligand efficiency”).
• the ratio (X) is a means for assessing antibody efficiency and, in turn, vaccine efficacy.
• the purpose of this example is to demonstrate the antagonism of nicotine vaccines, as described above, against nicotine by measuring the latency of mouse paw withdrawal from a hot plate.
• mice Four test groups of mice were immunized with the four vaccines, respectively, and according to the immunization schedule as described above in Example 5.
• a fifth and vaccine -naive group of mice received saline injections during the immunization schedule. All five groups of mice were then administered nicotine and then assayed on a hot plate to measure the latency in paw withdrawal from the hot plate is measured.
• %MPE (test - baseline)/(cutoff - baseline) x 100, where "test” is the latency to respond after treatment; “baseline” is the latency to respond prior to treatment; and “cutoff is the preset time at which the test was ended in the absence of a response.
• a baseline measure was obtained for each animal in the five groups prior to the immunization schedule above.
• Nabi Biopharmaceuticals announces results of first NicVAX phase III clinical trial: smoking cessation immunotherapy failed to meet primary endpoint. Nabi Biopharmaceuticals: Rockville, MD, July 18, 201 1.
• Nabi Biopharmaceuticals announces results of second NicVAX phase III clinical trial: smoking cessation immunotherapy failed to meet primary endpoint. Nabi Biopharmaceuticals: Rockville, MD, November 7, 201 1.
• tSVP Synthetic Vaccine Particle
• Immunogenicity of individual vaccine components in a bivalent nicotine vaccine differ according to vaccine formulation and administration conditions.

Landscapes

• Chemical & Material Sciences (AREA)
• Health & Medical Sciences (AREA)
• Organic Chemistry (AREA)
• Life Sciences & Earth Sciences (AREA)
• General Health & Medical Sciences (AREA)
• Medicinal Chemistry (AREA)
• Immunology (AREA)
• Animal Behavior & Ethology (AREA)
• Pharmacology & Pharmacy (AREA)
• Veterinary Medicine (AREA)
• Public Health (AREA)
• Botany (AREA)
• Biochemistry (AREA)
• Proteomics, Peptides & Aminoacids (AREA)
• Molecular Biology (AREA)
• Genetics & Genomics (AREA)
• Biophysics (AREA)
• Microbiology (AREA)
• Epidemiology (AREA)
• Mycology (AREA)
• Addiction (AREA)
• Biomedical Technology (AREA)
• Psychiatry (AREA)
• Engineering & Computer Science (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Neurology (AREA)
• General Chemical & Material Sciences (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Neurosurgery (AREA)
• Chemical Kinetics & Catalysis (AREA)
• Peptides Or Proteins (AREA)
• Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
• Plural Heterocyclic Compounds (AREA)
• Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Priority Applications (10)

Applications Claiming Priority (2)

Related Child Applications (2)

Publications (1)

Family

ID=54554631

Family Applications (1)

Country Status (8)

Cited By (1)

Families Citing this family (1)

Citations (4)

Family Cites Families (5)

• 2015
  • 2015-05-19 US US15/311,322 patent/US20170107275A1/en not_active Abandoned
  • 2015-05-19 KR KR1020167035358A patent/KR102435658B1/ko active Active
  • 2015-05-19 JP JP2016568391A patent/JP6564396B2/ja not_active Expired - Fee Related
  • 2015-05-19 CN CN201910630434.2A patent/CN110452217B/zh not_active Expired - Fee Related
  • 2015-05-19 WO PCT/US2015/031583 patent/WO2015179403A1/en not_active Ceased
  • 2015-05-19 AU AU2015264325A patent/AU2015264325B2/en not_active Expired - Fee Related
  • 2015-05-19 EP EP15796375.2A patent/EP3145920B1/en active Active
  • 2015-05-19 CA CA2949667A patent/CA2949667C/en active Active
  • 2015-05-19 CN CN201580026158.9A patent/CN106458971A/zh active Pending
• 2017
  • 2017-11-21 US US15/819,283 patent/US20180086820A1/en not_active Abandoned
• 2018
  • 2018-12-10 US US16/215,296 patent/US10487141B2/en not_active Expired - Fee Related

Patent Citations (4)

Non-Patent Citations (1)

Also Published As

Similar Documents

Legal Events