EP1289494A2 - Compositions de poudre - Google Patents

Compositions de poudre

Info

Publication number
EP1289494A2
EP1289494A2 EP01942075A EP01942075A EP1289494A2 EP 1289494 A2 EP1289494 A2 EP 1289494A2 EP 01942075 A EP01942075 A EP 01942075A EP 01942075 A EP01942075 A EP 01942075A EP 1289494 A2 EP1289494 A2 EP 1289494A2
Authority
EP
European Patent Office
Prior art keywords
powder
weight
salt
adjuvant
antigen
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP01942075A
Other languages
German (de)
English (en)
Inventor
Yuh-Fun Maa
Lu Zhao
Steven Joseph Prestrelski
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Powderject Vaccines Inc
Original Assignee
Powderject Vaccines Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Powderject Vaccines Inc filed Critical Powderject Vaccines Inc
Publication of EP1289494A2 publication Critical patent/EP1289494A2/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • 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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • 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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1682—Processes
    • A61K9/1694—Processes resulting in granules or microspheres of the matrix type containing more than 5% of excipient
    • 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
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A61K9/0021—Intradermal administration, e.g. through microneedle arrays, needleless injectors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • 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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1611—Inorganic compounds

Definitions

  • the invention relates to vaccine compositions. More specifically, the invention relates to vaccine compositions suitable for transdermal particle delivery from a needleless syringe system.
  • transdermal delivery provides many advantages over oral or parenteral delivery techniques, particular, transdermal delivery provides a safe, convenient and noninvasive alternative to traditional administration systems, conveniently avoiding the major problems associated with oral delivery (e.g. variable rates of absorption and metabolism, gastrointestinal irritation and/or bitter or unpleasant drug tastes) or parenteral delivery (e.g. needle pain, the risk of introducing infection to treated individuals, the risk of contamination or infection of health care workers caused by accidental needle-sticks and the disposal of used needles).
  • oral delivery e.g. variable rates of absorption and metabolism, gastrointestinal irritation and/or bitter or unpleasant drug tastes
  • parenteral delivery e.g. needle pain, the risk of introducing infection to treated individuals, the risk of contamination or infection of health care workers caused by accidental needle-sticks and the disposal of used needles.
  • transdermal delivery presents a number of its own inherent logistical problems. Passive delivery through intact skin necessarily entails the transport of molecules through a number of structurally different tissues, including the stratum corneum, the viable epidermis, the papillary dermis and the capillary walls in order for the drug to gain entry into the blood or lymph system. Transdermal delivery systems must therefore be able to overcome the various resistances presented by each type of tissue.
  • Transdermal delivery systems must therefore be able to overcome the various resistances presented by each type of tissue.
  • a number of alternatives to passive transdermal delivery have been developed. These alternatives include the use of skin penetration enhancing agents, or "permeation enhancers," to increase skin permeability, as well as non-chemical modes such as the use of iontophoresis, electroporation or ultrasound.
  • these alternative techniques often give rise to their own unique side effects such as skin irritation or sensitization. Thus, the spectrum of agents that can be safely and effectively administered using traditional transdermal delivery methods has remained limited.
  • the needleless syringe can also be used in conjunction with surgery to deliver drugs and biologies to organ surfaces, solid tumors and or to surgical cavities (e.g., tumor beds or cavities after tumor resection).
  • surgical cavities e.g., tumor beds or cavities after tumor resection.
  • any pharmaceutical agent that can be prepared in a substantially solid, particulate form can be safely and easily delivered using such devices.
  • Suitable vaccines include those comprising an antigen adsorbed into a salt adjuvant.
  • Such compositions are known in the art (see for example U.S.Patent No. 5,902,565) and are advantageous since the adjuvant enhances the immunogenicity of the vaccine.
  • An alternative method for storing adjuvant vaccine compositions is therefore required, which addresses the problems of aggregation associated with freeze-drying and which provides maximum retention of immunogenicity.
  • Prolonged storage of vaccines is essential, both for use with the novel transdermal drug delivery systems mentioned above and also for use with conventional vaccination techniques.
  • the provision of an effective alternative to freeze-drying is therefore of considerable commercial importance.
  • the vaccine be produced in a form suitable for needleless injection. Needleless injection requires the vaccine composition to be in powder form, each particle having a suitable size and strength for transdermal delivery and being capable of forming a gel on resuspension.
  • EP-B-0130619 is also concerned with the addition of stabilisers to lyophilised, or freeze-dried, vaccine preparations.
  • Lyophilised preparations of a hepatitis B vaccine comprising an inactivated purified hepatitis B virus surface antigen absorbed an aluminum gel and stabiliser are described.
  • the stabiliser is composed of at least one amino acid or salt thereof, at least one saccharide and at least one colloidal substance.
  • Very low concentrations of aluminum salt adjuvant are used, typically less than 0.1% by weight.
  • this document relates only to the hepatitis B vaccine and does not disclose a generic process, which is non-immunogen-specific. Spray-dried vaccine preparations comprising an immunogen adsorbed into an aluminum salt are disclosed in U.S. Patent No.
  • the spray freeze-drying method involves atomizing the suspended vaccine composition into liquid nitrogen. This process has two important effects: firstly, the liquid nitrogen acts as a heat transfer agent and provides rapid freezing of the suspension; and secondly, the atomisation reduces the volume of each droplet to be frozen, further increasing the freezing rate. This combined effect causes extremely rapid freezing of very small droplets of suspension and leads to the formation of smaller ice crystals in the solid.
  • the freeze concentrate regions which form during a standard freeze-drying technique are therefore significantly reduced in size. The rapid freezing of the particles, and their small size leads to powders having little or no aggregated adjuvant.
  • the present invention therefore provides simple, yet effective techniques that generate salt adjuvant-containing vaccine compositions in a powder form which is suitable for long-term storage.
  • the vaccine compositions of the invention show substantially no aggregation on reconstitution and therefore immunogenicity is substantially retained.
  • the compositions also have well-defined particle size, density and mechanical properties which collectively are suitable for powders for transdermal delivery from a needleless syringe.
  • the invention has the further, significant advantage that it is suitable for use with a wide range of vaccine compositions and may well also be applicable to other pharmaceutical compositions, in particular where similar aggregation problems are encountered.
  • the spray freeze-drying technique has been found to be entirely formulation independent within the field of adjuvant vaccine compositions.
  • the present invention provides a gel-forming free-flowing powder suitable for use as a vaccine, said powder being obtainable by spray-drying or spray freeze-drying an aqueous suspension comprising:
  • Free-flowing powder compositions suitable for vaccine use can thus be produced.
  • the compositions have well-defined particle size, density and mechanical properties which collectively are suitable for powders for transdermal delivery from a needleless syringe.
  • the invention further provides : a process for the preparation of a gel-forming free-flowing powder suitable for use as a vaccine, which process comprises spray-drying or spray freeze-drying an aqueous suspension comprising:
  • the present invention provides a powder suitable for use as a vaccine, said powder being obtainable by spray freeze-drying an aqueous suspension comprising an aluminum salt or calcium salt adjuvant having an antigen adsorbed therein.
  • the invention further provides: a process for the preparation of a powder suitable for use as a vaccine, which process comprises spray freeze-drying an aqueous suspension comprising an aluminum salt or calcium salt adjuvant having an antigen adsorbed therein; a dosage receptacle for a needleless syringe, said receptacle containing an effective amount of such a spray freeze-dried powder of the invention; a needleless syringe which is loaded with this spray freeze-dried powder of the inventic - a vaccine composition comprising a pharmaceutically acceptable carrier or diluent and the spray freeze-dried powder of the invention; and a method of vaccinating a subject, which method comprises administering to the said subject an effective amount of the spray freeze-dried powder of the invention.
  • Figure 1 shows the particle size distribution of an HBsAg adsorbed alum gel (i) before drying and (ii) after drying using a spray freeze-drying technique followed by reconstitution in water.
  • Figure 2 shows the particle size distribution of a second HBsAg adsorbed alum gel before drying and after drying via a conventional freeze drying method.
  • Figure 3 illustrates the results of an immunogenicity study using mice injected with HBsAg absorbed alum vaccine which had been dried by either spray freeze-drying (SFD) according to present invention, or using freeze-drying (FD).
  • SFD spray freeze-drying
  • FD freeze-drying
  • Figure 4 illustrates the immunogenicity of three different spray freeze-dried powders in mice immunized by either intramuscular injection using a needle or epidermal powder immunization using a powder delivery device.
  • Figure 5 illustrates the immunogenicity of spray freeze-dried diphtheria-tetanus toxoid vaccine in guinea pigs.
  • Spray freeze-dried powders of 20-38 ⁇ m and 38-53 ⁇ m in diameter were administered as a powder to the abdominal skin using a powder delivery device.
  • antigen is meant a molecule which contains one or more epitopes that will stimulate a host's immune system to make a cellular antigen-specific immune response or a humoral antibody response.
  • antigens include polypeptides including antigenic protein fragments, oligosaccharides, polysaccharides and the like.
  • the antigen can be derived from any known virus, bacterium, parasite, plant, protozoan or fungus, and can be a whole organism.
  • the term also includes tumor antigens.
  • Synthetic antigens are also included, for example polyepitopes, flanking epitopes and other recombinant or synthetically derived antigens (Bergmann et al (1993) Eur. J. Immunol. 23:2777-2781; Bergmann et al. (1996) J. Immunol. 157:3242-3249; Suhrbier, A. (1997) Immunol, and Cell Biol. 75:402-408; Gardner et al. (1998) 12 th World AIDS Conference, Geneva, Switzerland, June 28-July 3, 1998).
  • aduvants having antigen adsorbed thereon of the present invention are typically combined with one or more added materials such as carriers, vehicles, and/or excipients.
  • carriers generally refer to substantially inert materials which are nontoxic and do not interact with other components of the composition in a deleterious manner. These materials can be used to increase the amount of solids in particulate pharmaceutical compositions.
  • suitable carriers include water, silicone, gelatin, waxes, and like materials.
  • excipients include pharmaceutical grades of carbohydrates including monosaccharides, disaccharides, cyclodextrans, and polysaccharides (e.g., dextrose, sucrose, lactose, trehalose, raffmose, mannitol, sorbitol, inositol, dextrans, and maltodextrans); starch; cellulose; salts (e.g. sodium or calcium phosphates, calcium sulfate, magnesium sulfate); citric acid; tartaric acid; glycine; high molecular weight polyethylene glycols (PEG); Pluronics; surfactants; and combinations thereof.
  • carriers and/or excipients are used, they are used in amounts ranging from about 0.1 to 99 wt% of the pharmaceutical composition.
  • powder refers to a composition that consists of substantially solid particles that can be delivered transdermally using a needleless syringe device.
  • the particles that make up the powder can be characterized on the basis of a number of parameters including, but not limited to, average particle size, average particle density, particle morphology (e.g. particle aerodynamic shape and particle surface characteristics) and particle penetration energy (P.E.).
  • the average particle size of the powders according to the present invention can vary widely and is generally from 0.1 to 250 ⁇ m, for example from 10 to 100 ⁇ m and more typically from 20 to 70 ⁇ m.
  • the average particle size of the powder can be measured as a mass mean aerodynamic diameter (MMAD) using conventional techniques such as microscopic techniques (where particles are sized directly and individually rather than grouped statistically), absorption of gases, permeability or time of flight.
  • MMAD mass mean aerodynamic diameter
  • automatic particle-size counters can be used (e.g. Aerosizer Counter, Coulter Counter, HIAC Counter, or German Automatic Particle Counter) to ascertain the average particle size.
  • envelope density measurements can be used to assess the density of a powder according to the invention.
  • the envelope density of a powder of the invention is generally from 0.1 to 25 g/cm 3 , preferably from 0.8 to 1.5 g/cm 3 .
  • Envelope density information is particularly useful in characterizing the density of objects of irregular size and shape.
  • Envelope density is the mass of an object divided by its volume, where the volume includes that of its pores and small cavities but excludes interstitial space.
  • a number of methods of determining envelope density are known in the art, including wax immersion, mercury displacement, water absorption and apparent specific gravity techniques.
  • a number of suitable devices are also available for determining envelope density, for example, the GeoPycTM Model 1360, available from the Micromeritics Instrument Corp.
  • the difference between the absolute density and envelope density of a sample pharmaceutical composition provides information about the sample's percentage total porosity and specific pore volume.
  • Particle morphology particularly the aerodynamic shape of a particle
  • the particles which make up the instant powders have ji substantially spherical or at least substantially elliptical aerodynamic shape. It is also preferred that the particles have an axis ratio of 3 or less to avoid the presence of rod- or needle-shaped particles.
  • These same microscopic techniques can also be used to assess the particle surface characteristics, e.g. the amount and extent of surface voids or degree of porosity.
  • Particle penetration energies can be ascertained using a number of conventional techniques, for example a metallized film P.E. test.
  • a metallized film material e.g.
  • a 125 ⁇ m polyester film having a 350 A layer of aluminum deposited on a single side is used as a substrate into which the powder is fired from a needleless syringe (e.g. the needleless syringe described in U.S. Patent No. 5,630,796 to Bellhouse et al) at an initial velocity of about 100 to 3000 m/sec.
  • the metallized film is placed, with the metal-coated side facing upwards, on a suitable surface.
  • a needleless syringe loaded with a powder is placed with its spacer contacting the film, and then fired. Residual powder is removed from the metallized film surface using a suitable solvent. Penetration energy is then assessed using a BioRad Model GS-700 imaging densitometer to scan the metallized film, and a personal computer with a SCSI interface and loaded with MultiAnalyst software (BioRad) and Matlab software (Release 5.1, The MathWorks, Inc.) is used to assess the densitometer reading.
  • a program is used to process the densitometer scans made using either the transmittance or reflectance method of the densitometer.
  • the penetration energy of the spray-coated powders should be equivalent to, or better than that of reprocessed mannitol particles of the same size (mannitol particles that are freeze-dried, compressed, ground and sieved according to the methods of commonly owned International Publication No. WO 97/48485, incorporated herein by reference).
  • subject refers to any member of the subphylum cordata including, without limitation, humans and other primates including non-human primates such as chimpanzees and other apes and monkey species; farm animals such as cattle, sheep, pigs, goats and horses; domestic mammals such as dogs and cats; laboratory animals including rodents such as mice, rats and guinea pigs; birds, including domestic, wild and game birds such as chickens, turkeys and other gallinaceous birds, ducks, geese, and the like.
  • the term does not denote a particular age. Thus, both adult and newborn individuals are intended to be covered.
  • the methods described herein are intended for use in any of the above vertebrate species, since the immune systems of all of these vertebrates operate similarly.
  • transdermal delivery includes both transdermal ("percutaneous") and transmucosal routes of administration, i.e. delivery by passage through the skin or mucosal tissue.
  • transdermal Drug Delivery Developmental Issues and Research Initiatives, Hadgraft and Guy (eds.), Marcel Dekker, hie, (1989); Controlled Drug Delivery: Fundamentals and Applications, Robinson and Lee (eds.), Marcel Dekker Inc., (1987); and Transdermal Delivery of Drugs, Nols. 1-3, Kydonieus and Berner (eds.), CRC Press, (1987).
  • the invention is concerned with gel-forming free-flowing powders suitable for use as vaccines.
  • the powders are suitable for transdermal administration from a needleless syringe delivery system.
  • the particles which make up the powdered composition must have sufficient physical strength to withstand sudden acceleration to several times the speed of sound and the impact with, and passage through, the skin and tissue.
  • the particles are formed by spray-drying or spray freeze-drying an aqueous suspension comprising or, in some embodiments, consisting essentially of:
  • the aqueous suspension contains, as component (a), less than 1% by weight of the adjuvant having antigen adsorbed thereon.
  • the suspension contains from 0.2 or 0.3 to 0.6 or 0.75% by weight, preferably from 0.2 to 0.4% by weight, of the adjuvant onto which antigen is adsorbed.
  • the aluminum salt adjuvant is generally aluminum hydroxide or aluminum phosphate.
  • the adjuvant may be aluminum sulfate or calcium phosphate.
  • the antigen may be a viral antigen.
  • the antigen may therefore be derived from members of the families Picornaviridae (e.g. polioviruses, etc.); Caliciviridae; Togaviridae (e.g. rubella virus, dengue virus, etc.); Flaviviridae; Coronaviridae; Reoviridae; Birnaviridae; Rhabodoviridae (e.g. rabies virus, etc.); Filoviridae; Paramyxoviridae (e.g. mumps virus, measles virus, respiratory syncytial virus, etc.); Orthomyxoviridae (e.g.
  • viral antigens may be derived from papillomavirus (e.g. HPN); a herpesvirus; a hepatitis virus, e.g.
  • hepatitis A virus (HAN), hepatitis B virus (HBV), hepatitis C (HCV), the delta hepatitis virus (HDV), hepatitis E virus (HEV) or hepatitis G virus (HGV); and the tick-borne encephalitis viruses.
  • HAN hepatitis A virus
  • HBV hepatitis B virus
  • HCV hepatitis C
  • HDV delta hepatitis virus
  • HEV hepatitis E virus
  • HGV hepatitis G virus
  • tick-borne encephalitis viruses See, e.g. Virology, 3rd Edition (W.K. Joklik ed. 1988); Fundamental Virology, 2nd Edition (B. ⁇ . Fields and D.M. Knipe, eds. 1991) for a description of these viruses.
  • Bacterial antigens for use in the invention can be derived from organisms that cause diphtheria, cholera, tuberculosis, tetanus, pertussis, meningitis and other pathogenic states, including, e.g., Meningococcus A, B and C, Hemophilus influenza type B (HTB), Helicobacter pylori, Vibrio cholerae, Escherichia coli, Campylobacter, Shigella, Salmonella, Streptococcus sp, and Staphylococcus sp.
  • a combination of bacterial antigens maybe provided, for example diphtheria, pertussis and tetanus antigens.
  • Suitable pertussis antigens are pertussis toxin and/or filamentous haemagglutinin and/or pertactin, alternatively termed P69.
  • An anti-parasitic antigen may be derived from organisms causing malaria and Lyme disease.
  • Antigens for use in the present invention can be produced using a variety of methods known to those of skill in the art.
  • the antigens can be isolated directly from native sources, using standard purification techniques. Alternatively, whole killed, attenuated or inactivated bacteria, viruses, parasites or other microbes may be employed.
  • antigens can be produced recombinantly using known techniques. See, e.g., Sambrook, Fritsch & Maniatis, Molecular Cloning: A Laboratoi ⁇ Manual, Vols. I and H (D. ⁇ . Glover et. 1985).
  • Antigens for use herein may also be synthesised, based on described amino acid sequences, via chemical polymer syntheses such as solid phase peptide synthesis.
  • chemical polymer syntheses such as solid phase peptide synthesis.
  • Such methods are known to those of skill in the art. See, e.g. J.M. Stewart and J.D. Young, Solid Phase Peptide Synthesis, 2nd Ed., Pierce Chemical Co., Rockford, IL (1984) and G. Barany and R. B. Merrifield, The Peptides: Analysis, Synthesis, Biology, editors E. Gross and J. Meienhofer, Vol. 2, Academic Press, New York, (1980), pp. 3-254, for solid phase peptide synthesis techniques; and M. Bodansky, Principles of Peptide Synthesis, Springer-Verlag Berlin (1984) and E. Gross and J. Meienhofer, Eds., The Peptides: Analysis, Synthesis, Biology, supra, Vol. 1, for classical solution synthesis.
  • One or more saccharides may be present in the aqueous suspension as component (b).
  • the saccharide content is typically 1.5 to 5% by weight, preferably 2 to 4% by weight.
  • the saccharide may be a monosaccharide such as glucose, xylose, galactose, fructose, D-mannose or sorbose; a disaccharide such as lactose, maltose, saccharose, trehalose or sucrose; or a sugar alcohol such as mannitol, sorbitol, xylitol, glycerol, erytbritol or arabitol.
  • One or more amino acids or amino acid salts is present in the aqueous suspension as component (c). Any physiologically acceptable amino acid salt maybe employed.
  • the salt may be an alkali or alkaline earth metal salt such as sodium, potassium or magnesium salt.
  • the amino acid may be an acidic, neutral or basic amino acid. Suitable amino acids are glycine, alanine, glutamine, arginine, lysine and histidine. Monosodium glutamate is a suitable amino acid salt.
  • the aqueous suspension generally contains from 0.5 to 1.5% by weight, more preferably from 0.75 to 1.25% by weight, of the amino acid and/or amino acid salt.
  • the colloidal substance (d) is a divided substance incapable of passing through a semi-permeable membrane, comprised of fine particles which, in suspension or solution, fail to settle out. Suitable colloidal substances are disclosed in EP-B-0130619.
  • Component (d) may be selected from polysaccharides such as dextran or maltodextran; hydrogels such as gelatin or agarose; or proteins such as human serum albumin.
  • the substance may have a molecular weight of 500 to 80,000 or higher, for example from 1000 or 2000 to 30,000 or from 5,000 to 25,000.
  • Component (d) is generally present in the aqueous suspension in an amount of from 0.05 to 0.5% by weight, preferably from 0.07 to 0.3% by weight.
  • the adjuvant having antigen adsorbed thereon and the saccharide, amino acid or salt thereof and colloidal substance are suspended in water.
  • the aqueous suspension is spray dried or spray freeze-dried.
  • the spray-drying or spray freeze-drying conditions are selected to enable the desired particles to be produced.
  • the air inlet temperature, air outlet temperature, feed rate of the aqueous suspension, air flow rate, etc. can thus be varied as desired.
  • Any suitable spray-drier may be used.
  • the nozzle size may vary as necessary. Particular spray freeze-drying conditions are described in more detail below.
  • a gel-forming free-flowing powder can thus be provided which is suitable for use as a vaccine.
  • the proportions of the various components of the powder can be adjusting by adjusting the composition of the suspension that is spray-dried or spray freeze-dried.
  • the powder typically comprises or, in some embodiments, consists essentially of: (i) from 5 to 60%, for example from 7 to 50% such as from 10 to 30%, by weight of an aluminum salt or calcium salt adjuvant having an antigen adsorbed thereon; (ii) from 25 to 90%, for example from 30 to 80% such as from 40 to 70%, by weight of a saccharride;
  • the invention is concerned generally with powders suitable for use as vaccines that are formed by spray freeze-drying an aqueous suspension comprising an aluminum salt or calcium salt adjuvant having an antigen adsorbed therein.
  • powders are suitable for transdermal administration from a needleless syringe delivery system.
  • the particles which make up the powdered composition must have sufficient physical strength to withstand sudden acceleration of up to several times the speed of sound and the impact with, and passage through, the skin and tissue.
  • the aqueous suspension prior to spray freeze-drying, contains less than 10% by weight, for instance less than 5% weight and preferably less than 3% by weight, of the salt adjuvant having antigen adsorbed thereon.
  • the aqueous suspension typically contains at least 0.05% by weight, for instance at least 0.1% by weight or at least 0.6% by weight, of the adjuvant having antigen adsorbed thereon. More preferably, the suspension contains from 0.2 or 0.3 to 0.6%, 0.75% or 1% by weight, preferably from 0.2 to 0.4% by weight, of adjuvant onto which antigen is adsorbed. At concentrations above about 10% by weight of adjuvant salt, the aqueous suspension becomes highly viscous. This limits the ability to atomize the suspension.
  • adjuvant concentration applies to the aqueous suspension prior to spray freeze-drying.
  • the content of adjuvant salt having antigen adsorbed thereon may be as high as 50% by weight or more in the spray freeze-dried powders of the invention.
  • the adjuvant is generally an aluminum salt, for example aluminum hydroxide or aluminum phosphate.
  • the adjuvant salt may be aluminum sulfate or calcium phosphate.
  • the antigen may be a viral antigen.
  • the antigen may therefore be derived from members of the families Picornaviridae (e.g. polioviruses, etc.); Caliciviridae; Togaviridae (e.g. rubella virus, dengue virus, etc.); Flaviviridae; Coronaviridae; Reoviridae; Birnaviridae; Rhabodoviridae (e.g. rabies virus, etc.); Filoviridae; Paramyxoviridae (e.g. mumps virus, measles virus, respiratory syncytial virus, etc.); Orthomyxoviridae (e.g.
  • influenza virus types A, B and C, etc. Bunyaviridae; Arenaviridae; Retroviradae (e.g. HTLV-I; HTLV-II; HIV-1 and H ⁇ V-2); and simian immunodeficiency virus (SIN) among others.
  • Bunyaviridae Arenaviridae
  • Retroviradae e.g. HTLV-I; HTLV-II; HIV-1 and H ⁇ V-2
  • SIN simian immunodeficiency virus
  • viral antigens maybe derived from papiilomavirus (e.g. HPN); a herpesvirus; a hepatitis virus, e.g. hepatitis A virus (HAN), hepatitis B virus (HBN), hepatitis C (HCN), the delta hepatitis virus (HDN), hepatitis E virus (HEV) or hepatitis G virus (HGV); and the tick-borne encephalitis viruses.
  • a hepatitis virus e.g. hepatitis A virus (HAN), hepatitis B virus (HBN), hepatitis C (HCN), the delta hepatitis virus (HDN), hepatitis E virus (HEV) or hepatitis G virus (HGV)
  • a hepatitis virus e.g. hepatitis A virus (HAN), hepatitis B virus (HBN), hepatitis C (HCN), the delta hepati
  • Bacterial antigens for use in the invention can be derived from organisms that cause diphtheria, cholera, tuberculosis, tetanus, pertussis, meningitis and other pathogenic states, including, e.g., Meningococcus A, B and C, Hemophilus influenza type B (H B), Helicobacter pylori, Vibrio cholerae, Escherichia coli, Campylobacter, Shigella, Salmonella, Streptococcus sp, and Staphylococcus sp.
  • a combination of bacterial antigens may be provided, for example diphtheria, pertussis and tetanus antigens.
  • Suitable pertussis antigens are pertussis toxin and/or filamentous haemagglutinin and/or pertactin, alternatively termed P69.
  • An anti-parasitic antigen may be derived from organisms causing malaria and Lyme disease.
  • Antigens for use in the present invention can be produced using a variety of methods known to those of skill in the art. In particular, the antigens can be isolated directly from native sources, using standard purification techniques. Alternatively, whole killed, attenuated or inactivated bacteria, viruses, parasites or other microbes may be employed. Yet further, antigens can be produced recombinantly using known techniques. See, e.g., Sambrook, Fritsch & Maniatis, Molecular Cloning: A Laboratory Manual, Vols. I and ⁇ (D.N. Glover et. 1985).
  • Antigens for use herein may also be synthesised, based on described amino acid sequences, via chemical polymer syntheses such as solid phase peptide synthesis.
  • chemical polymer syntheses such as solid phase peptide synthesis.
  • Such methods are known to those of skill in the art. See, e.g. J.M. Stewart and J.D. Young, Solid Phase Peptide Synthesis, 2nd Ed., Pierce Chemical Co., Rockford, JL (1984) and G. Barany and R. B. Merrifield, The Peptides: Analysis, Synthesis, Biology, editors E. Gross and J. Meienhofer, Vol. 2, Academic Press, New York, (1980), pp. 3-254, for solid phase peptide synthesis techniques; and M. Bodansky, Principles of Peptide Synthesis, Springer-Verlag Berlin (1984) and E. Gross and J. Meienhofer, Eds., The Peptides: Analysis, Synthesis, Biology, supra, Vol. 1, for classical solution synthesis.
  • the aqueous suspension may consist essentially of water and adjuvant having an antigen adsorbed thereon, or further additives may be included in the suspension. Any additives may be employed provided that they are substantially non-toxic and pharmacologically inert.
  • the spray freeze-drying process has been found to be effective when applied to suspensions comprising a wide range of different additives and, as yet, the process of the invention, and therefore the powders of the invention, have been found to be entirely formulation independent.
  • the aqueous suspension comprises suitable excipients, along with protectants, solvents, salts, surfactants, buffering agents and the like.
  • suitable excipients can include free-flowing particulate solids that do not thicken or polymerize upon contact with water, which are innocuous when administered to an individual, and do not significantly interact with the pharmaceutical agent in a manner that alters its pharmaceutical activity.
  • excipients include, but are not limited to, monosaccharides such as glucose, xylose, galactose, fructose, D-mannose or sorbose, disaccharides such as lactose, maltose, saccharose, trehalose or sucrose, sugar alcohols such as marjnitol, sorbitol, xylitol, glycerol, erythritol or arabitol, polymers such as dextran, starch, cellulose or high molecular weight polyethylene glycols (PEG), amino acids or their salts, such as glycine, alanine, glutamine, arginine, lysine or histidine or their salts with alkali or alkaline earth metals such as a sodium, potassium or magnesium salts, or sodium or calcium phosphates, calcium carbonate, calcium sulfate, sodium citrate, citric acid, tartaric acid, and combinations thereof.
  • monosaccharides
  • Suitable solvents include, but are not limited to, methylene chloride, acetone, methanol, ethanol, isopropanol and water. Typically, water is used as the solvent.
  • pharmaceutically acceptable salts having molarities ranging from about 1 mM to 2M can be used.
  • Pharmaceutically acceptable salts include, for example, mineral acid salts such as hydrochlorides, hydrobromides, phosphates, sulfates, and the like; and the salts of organic acids such as acetates, propionates, malonates, benzoates, and the like.
  • Preferred excipients for use in the aqueous suspension include saccharides, amino acids or salts thereof and polymers.
  • the suspension contains one or more saccharides, such as a combination of mannitol and trehalose. Saccharides are typically present in an amount of from 0.5 to 30% by weight.
  • Typical excipient combinations include one or more saccharides and a polymer and include substantially no amino salt.
  • the total amount of excipients present in the aqueous suspension is typically from 0 to 50%, more preferably from 10 to 30%.
  • the particles of the invention are formed by first suspending the adjuvant having an antigen adsorbed therein, and any required additives, in water.
  • the aqueous suspension is then spray freeze-dried.
  • Any known technique in the art for example the methods described by Mumenthaler et al, hit. J. Pharmaceutics (1991) 72, pages 97-110 and Maa et al, Phar. Res. (1999) Vol. 16, page 249) may be used to carry out the spray freeze- drying step.
  • a typical spray freeze-drying technique involves atomising the aqueous suspension into stirred liquid nitrogen.
  • the liquid nitrogen containing frozen particles is then held at reduced temperature, for example from -60°C to -20°C, followed by vacuum drying preferably under a pressure of from 20 to 500 mT (2.666 to 66.65 Pa), and at reduced temperature such as from -50 ° C to 0°C. Drying is typically carried out in two stages, primary drying and secondary drying. Primary drying time typically ranges from 4 to 24 hours and secondary drying time typically ranges from 6 to 24 hours. The temperature may be gradually increased, whilst still under reduced pressure until room temperature is reached. This technique involves the rapid freezing of the aqueous suspension into droplets.
  • the drying step then removes the ice by sublimation without the need for high air temperatures.
  • the powder may be collected by any known technique.
  • the precise spray freeze-drying conditions used maybe selected according to the desired properties of the particles to be produced. Thus, the temperatures, pressures and other conditions may be varied as desired.
  • the powders of the invention are generally free-flowing.
  • the powders contain very little or no agglomerated adjuvant salt and are therefore capable of forming a gel on resuspension in water. Typically, substantially no precipitate forms upon resuspension.
  • a gel-like suspension without any precipitate is typically obtained. No precipitates settling out are observed after 3 hours. No precipitates may form after standing overnight, for example for 12 hours.
  • the presence of a precipitate, and the degree of agglomeration of the reconstituted gel formulation is typically assessed by the ability of the reconstituted formulation to diffract a beam of light.
  • the degree of agglomeration can also be quantitatively assessed by standard light microscopy and/or sedimentation.
  • Another suitable test for particle agglomeration can be to determine particle size before and after reconstitution using any of a number of standard particle size determination techniques, e.g. laser-based or light obscuration.
  • the particles of the invention have a size appropriate for high- velocity transdermal delivery to a subject, typically across the stratum corneum or a transmucosal membrane.
  • the mass mean aerodynamic diameter (MMAD) of the particles is from about 0.1 to 250 ⁇ m.
  • the MMAD may be from 5 to 100 ⁇ m or from 10 to 100 ⁇ m, preferably from 10 to 70 ⁇ m or from 20 to 70 ⁇ m. Generally, less than 10% by weight of the particles have a diameter which is at least 5 ⁇ m more than the MMAD or at least 5 ⁇ m less than the
  • MMAD Preferably, no more than 5% by weight of the particles have a diameter which is greater than the MMAD by 5 ⁇ m or more. Also preferably, no more than 5% by weight of the particles have a diameter which is smaller than the MMAD by 5 ⁇ m or more.
  • the particles have an envelope density of from 0.1 to 25 g/cm 3 , preferably from 0.8 to 1.5 g/cm 3 . While the shape of the individual particles may vary when viewed under a microscope, the particles are preferably substantially spherical.
  • the average ratio of the major axis:minor axis is typically from 3:1 to 1:1, for example from 2:1 to 1:1.
  • the individual particles of a powder have a substantially spherical aerodynamic shape with a substantially uniform, nonporous surface.
  • the particles will also have a particle penetration energy suitable for transdermal delivery from a needleless syringe device.
  • needleless syringe devices useful in this invention is found in the prior art, as discussed herein. These devices are referred to as needleless syringe devices and representative of these devices are the dermal PowderJect ® needleless syringe device and the oral PowderJect ® needleless syringe device (PowderJect Technologies Limited, Oxford, UK).
  • an effective amount of the powder of the invention is delivered to the subject.
  • An effective amount is that amount needed to deliver sufficient of the desired antigen to achieve vaccination. This amount will vary with the nature of the antigen and can be readily determined through clinical testing based on known activities of the antigen being delivered.
  • Needleless syringe devices for delivering particles were first described in commonly owned U.S. Patent No. 5,630,796 to Bellhouse et al, incorporated herein by reference. Although a number of specific device configurations are now available, such devices are typically provided as a pen-shaped instrument containing, in linear order moving from top to bottom, a gas cylinder, a particle cassette or package, and a supersonic nozzle with an associated silencer medium.
  • An appropriate powder in the present case, a spray-dried or spray freeze-dried powder of the invention
  • a suitable container e.g., a cassette formed by two rupturable polymer membranes that are heat-sealed to a washer-shaped spacer to form a self-contained sealed unit.
  • Membrane materials can be selected to achieve a specific mode of opening and burst pressure that dictate the conditions at which the supersonic flow is initiated, hi operation, the device is actuated to release the compressed gas from the cylinder into an expansion chamber withi the device.
  • the released gas contacts the particle cassette and, when sufficient pressure is built up, suddenly breaches the cassette membranes sweeping the particles into the supersonic nozzle for subsequent delivery.
  • the nozzle is designed to achieve a specific gas velocity and flow pattern to deliver a quantity of particles to a target surface of predefined area.
  • the silencer is used to attenuate the noise produced by the membrane rupture.
  • a second needleless syringe device for delivering particles is described in commonly owned International Publication No. WO 96/20022.
  • This delivery system also uses the energy of a compressed gas source to accelerate and deliver powdered compositions; however, it is distinguished from the system of US Patent No. 5,630,796 in its use of a shock wave instead of gas flow to accelerate the particles. More particularly, an instantaneous pressure rise provided by a shock wave generated behind a flexible dome strikes the back of the dome, causing a sudden eversion of the flexible dome in the direction of a target surface.
  • This sudden eversion catapults a powdered composition (which is located on the outside of the dome) at a sufficient velocity, thus momentum, to penetrate target tissue, e.g., oral mucosal tissue.
  • the powdered composition is released at the point of full dome eversion.
  • the dome also serves to completely contain the high- pressure gas flow, which therefore does not come into contact with the tissue. Because the gas is not released during this delivery operation, the system is inherently quiet. This design can be used in other enclosed or otherwise sensitive applications for example, to deliver particles to minimally invasive surgical sites.
  • single unit dosages or multidose containers in which a powder of the invention may be packaged prior to use, can comprise a hermetically sealed container enclosing a suitable amount of the powder that makes up a suitable dose.
  • the powder can be packaged as a sterile formulation, and the hermetically sealed container can thus be designed to preserve sterility of the formulation until use.
  • the containers can be adapted for direct use in the above-referenced needleless syringe systems.
  • Powders of the present invention can thus be packaged in individual unit dosages for delivery via a needleless syringe.
  • a "unit dosage” intends a dosage receptacle containing a therapeutically effective amount of a powder of the invention.
  • the dosage receptacle typically fits within a needleless syringe device to allow for transdermal delivery from the device.
  • Such receptacles can be capsules, foil pouches, sachets, cassettes or the like.
  • the container in which the powder is packaged can further be labeled to identify the composition and provide relevant dosage information.
  • the container can be labeled with a notice in the form prescribed by a governmental agency, for example the Food and Drug Administration, wherein the notice indicates approval by the agency under Federal law of the manufacture, use or sale of the powder contained therein for human administration.
  • optimal particle densities for use in needleless injection generally range between about 0.1 and 25 g cm 3 such as between about 0.8 and 1.7 g/cm 3 , preferably between about 0.9 and 1.5 g/cm 3 .
  • Injection velocities generally range between about 100 and 3,000 m/sec.
  • the needleless syringe systems can be provided in a preloaded condition containing a suitable dosage of the powder of the invention.
  • the loaded syringe can be packaged in a hermetically sealed container, which may further be labeled as described above.
  • test methods have been developed, or established test methods modified, in order to characterize performance of a needleless syringe device. These tests range from characterization of the powdered composition, assessment of the gas flow and particle acceleration, impact on artificial or biological targets, and measures of complete system performance. One, several or all of the following tests can thus be employed to assess the physical and functional suitability of the powder of the invention for use in a needleless syringe system.
  • PE peernetration energy
  • the film test-bed has been shown to be sensitive to particle delivery variations of all major device parameters including pressure, dose, particle size distribution and material, etc. and to be insensitive to the gas.
  • Aluminum of about 350 Angstrom thickness on a 125 ⁇ m polyester support is currently used to test devices operated at up to 60 bar.
  • Another means of assessing particle performance when delivered via a needleless syringe device is to gauge the effect of impact on a rigid polymethylimide foam (Rohacell 5 UG, density 52 kg/m 3 , Rohm Tech Inc., Maiden, MA).
  • the experimental set-up for this test is similar to that used in the metallized film test. The depth of penetration is measured using precision calipers. For each experiment a processed mannitol standard is run as comparison and all other parameters such as device pressure, particle size range, etc., are held constant. Data also show this method to be sensitive to differences in particle size and pressure.
  • a further indicator of particle performance is to test the ability of various candidate compositions to withstand the forces associated with high- velocity particle injection techniques, that is, the forces from contacting particles at rest with a sudden, high velocity gas flow, the forces resulting from particle-to-particle impact as the powder travels through the needleless syringe, and the forces resulting from particle-to-device collisions also as the powder travels through the device.
  • a simple particle attrition test has been devised which measures the change in particle size distribution between the initial composition, and the composition after having been delivered from a needleless syringe device.
  • the test is conducted by loading a particle composition into a needleless syringe as described above, and then discharging the device into a flask containing a carrier fluid in which the particular composition is not soluble (e.g., mineral oil, silicone oil, etc.).
  • a carrier fluid in which the particular composition is not soluble (e.g., mineral oil, silicone oil, etc.).
  • the carrier fluid is then collected, and particle size distribution in both the initial composition and the discharged composition is calculated using a suitable particle sizing apparatus, e.g., an AccuSizer® model 780 Optical Particle Sizer.
  • Compositions that demonstrate less than about 50%), more preferably less than about 20% reduction in mass mean diameter (as determined by the AccuSizer apparatus) after device actuation are deemed suitable for use in the needleless syringe systems described herein.
  • candidate powder compositions can be injected into dermatomed, full thickness human abdomen skin samples.
  • Replicate skin samples after injection can be placed on modified Franz diffusion cells containing 32 °C water, physiologic saline or buffer.
  • Additives such as surfactants may be used to prevent binding to diffusion cell components. Two kinds of measurements can be made to assess performance of the formulation in the skin.
  • TEWL transepidermal water loss
  • Measurement is performed at equilibrium (about 1 hour) using a Tewameter TM 210® (Courage & Khazaka, Koln, Ger) placed on the top of the diffusion cell cap that acts like a ⁇ 12 mm chimney. Larger particles and higher injection pressures generate proportionally higher TEWL values in vitro and this has been shown to correlate with results in vivo.
  • TEWL values increased from about 7 to about 27 (g/m 2 h) depending on particle size and helium gas pressure.
  • Helium injection without powder has no effect.
  • the skin barrier properties return rapidly to normal as indicated by the TEWL returning to pretreatment values in about 1 hour for most powder sizes. For the largest particles, 53-75 ⁇ m, skin samples show 50% recovery in an hour and full recovery by 24 hours.
  • the antigen component(s) of candidate powders can be collected by complete or aliquot replacement of the Franz cell receiver solution at predetermined time intervals for chemical assay using HPLC or other suitable analytical technique. Concentration data can be used to generate a delivery profile and calculate a steady state permeation rate. This technique can be used to screen formulations for early indication of antigen binding to skin, antigen dissolution, efficiency of particle penetration of stratum corneum, etc., prior to in vivo studies.
  • the particles of a powder have the following characteristics: a substantially spherical shape (e.g. an aspect ratio as close as possible to 1); a smooth surface; a suitable active loading content; less than 20% reduction in particle size using the particle attrition test; an envelope density as close as possible to the true density of the constituents (e.g. greater than about 0.8 g/ml); and a MMAD of about 20 to 70 ⁇ m with a narrow particle size distribution.
  • the compositions are typically free -flowing (e.g.
  • a powder of the invention may alternatively be used to vaccinate a subject via other routes.
  • the powder may be combined with a suitable carrier or diluent such as Water for Injections or physiologically saline.
  • a suitable carrier or diluent such as Water for Injections or physiologically saline.
  • the resulting vaccine composition is typically administered by injection, for example subcutaneously or intramuscularly.
  • an effective amount of antigen is delivered to the subject being vaccinated. Generally from 50 ng to 1 mg and more preferably from 1 ⁇ g to about 50 ⁇ g of antigen will be useful in generating an immune response. The exact amount necessary will vary depending on the age and general condition of the subject to be treated, the particular antigen or antigens selected, the site of administration and other factors. An appropriate effective amount can be readily determined by one of skill in the art. Dosage treatment may be a single dose schedule or a multiple dose schedule.
  • a multiple dose schedule is one in which a primary course of vaccination maybe with 1-10 separate doses, followed by other doses given at subsequent time intervals, chosen to maintain and/or reinforce the immune response, for example at 1-4 months for second dose and, if needed, a subsequent dose(s) after several months.
  • the dosage regimen will also, at least in part, be determined by the need of the subject and be dependent on the judgement of the practitioner. Vaccination will of course generally be effected prior to primary infection with the pathogen against which protection is desired.
  • a spray-dried immediate-release vaccine preparation was obtained according to the procedure described in US Patent No. 5,902,565.
  • a formulation containing 5%> by weight mannitol and 5% by weight aluminum phosphate (Adju-Phos) was spray dried using a bench-top spray dryer (Buchi 190).
  • the free-flowing powder that was obtained had a particle size of about 10 ⁇ m.
  • the powder was reconstituted in distilled water (1 :500 by weight). The solution failed to form a gel with the suspended particles setting in 15 minutes. By optical microscopy, the particles after reconstitution maintained their shape and size, suggesting that the alum remained coagulated and did not disintegrate.
  • Example 1 The following formulations were prepared by mixing the components listed in the Table below in 15 ml of distilled water:
  • Adju-Phos 2% by weight aluminum phosphate
  • composition of the powders obtained in relation to the solids content of the suspension subjected to spray drying was as follows:
  • Formulation 2 37.1 mg of spray-dried powder was added to 1 ml of distilled water. An off-white, grey, gel-like suspension formed. No precipitate was observed after the suspension had been allowed to stand overnight.
  • Formulation 4 29.4 mg of spray-dried powder was added to 1 ml of distilled water. A white precipitate formed after the resulting suspension had been allowed to stand overnight.
  • Example 2
  • a concentrated alum-HBsAg suspension was prepared by first washing an alum- adsorbed HBsAg vaccine obtained from Rhein Americana S.A. containing 20 ⁇ g of HBsAg (approximately 1 human dose) adsorbed on 500 ⁇ g of alum (approximately 1500 ⁇ g of aluminum hydroxide) with distilled, deionised water to remove buffer salt.
  • Alum gel was allowed to settle overnight in a 250-mL Nalgene narrow-mouth square polycarbonate bottle at 2-8°C. The supernatant (150mL) was removed and the same volume of water was added to the precipitates and mixed. This procedure was repeated for a second time.
  • lOOg of the washed alum-HBsAg formulation was weighed in a Nalgene square bottle and allowed to settle overnight at 2-8°C. After 90mL of supernatant was removed, the remaining suspension was transferred to a 50mL polypropylene centrifuge tube and centrifuged at 200 rpm for 4 minutes using a bench-top centrifuge (Allegra 6R, Beckman). The supernatant was further removed to obtain 3.369g of concentrated alum-HBsAg suspension.
  • This suspension was then mixed with 315.24mg mannitol, 81.73mg glycine, 101.91mg dextran and placebo alum gel (Al 2 O 3 at 2%) to achieve a liquid alum-HBsAg formulation having an alum concentration of 3%.
  • alum-HBsAg suspension was washed in accordance with the method described for formulation A. 20.79g of the suspension was weighed in a 50mL centrifuge tube and allowed to settle overnight at 2-8°C. After 17mL of supernatant was removed, the remaining concentrated suspension (3.572g) was mixed with 113.06mg mannitol, 47.3 lmg glycine and 23.22mg dextran to produce a liquid formulation having an alum concentration of 0.6%. The two formulations were dried using the techniques set out in Table 1 below:
  • a Dura-Stop freeze dryer (FTS System, Stone Ridge, NY) was used to freeze dry the alum-adsorbed HBsAg formulation based on the freeze-drying cycle in Table 2.
  • Each suspension solution was sprayed into liquid nitrogen stirred in a stainless steel pain using an ultrasonic atomizer (Sono Tek Corporation, Milton, NY) with a nozzle frequency of 60 kHz. Sonic energy for atomization was set at 5.0 watts.
  • Liquid feed was delivered by a MasterFlex C/L peristaltic pump at 1.5 mL/min. The pan containing frozen particles in liquid nitrogen was loaded into the Dura-lyophilizer pre-cooled to -50 °C and freeze-dried based on the condition of Table 3.
  • the lyophilized material was rendered into particulate form using a compress, grind and sieve ("C/G/S") technique. More particularly, the lyophilized material was compressed in a stainless steel dye of 13-mm in diameter (Carver Press, Wabash, IN) at a pressure of 12,000 psi for 5-10 minutes. The compressed discs were ground manually using a mortar and pestle. The ground powder was manually sieved through a stack of sieves (3-in diameter) into three size fractions, 53-75 ⁇ m, 38-53 ⁇ m, and 20-38 ⁇ m.
  • C/G/S compress, grind and sieve
  • Experiment 1 Effect of drying process on the extent of coagulation Powders 1 to 3 were reconstituted in water at a ratio of 1 :500w/w and examined using optical microscopy in accordance with standard techniques. Visual analysis of the particles was performed using an optical microscope (Model DMR, Leica, Germany) with lOx-eyepeice lens and 5x-objective lens. The system was equipped with a Polaroid camera system for image output. Optical microscopy provides a qualitative analysis of the degree of alum coagulation. In this experiment, powder 1 produced very large aggregates on reconstitution, whereas powder 2 coagulated only slightly. Powder 3 produced almost no aggregates at all.
  • the particle size of the reconstituted powders was also measured quantitatively.
  • the reconstituted powder sample was vortexed/sonicated to make a homogeneous suspension.
  • the suspension was then added to the glass container of a particle size analyzer (AccuSizer 780, Particle Sizing Systems, Santa Barbara, CA) for particle size distribution measurement.
  • the results of the measurements carried out on powders 2 and 3 both before and after spray freeze-drying are shown in Figure 1.
  • Similar comparative results for powder 1 showing particle size before and after freeze-drying are shown in Figure 2.
  • These results illustrate the similar particle size distribution of powders 2 and 3 before and after drying, demonstrating that little or no alum coagulation occurred during freeze-drying.
  • the particle size of powder 1 increases significantly after freeze- drying, indicating that significant alum coagulation has occurred.
  • Powders were reconstituted with distilled water and used to immunize Balb/C mice (female, 8 per group, 5-7 weeks old at the beginning of the study).
  • Reconstituted vaccines were administered by intraperitoneal injection using a 23 1/5 needle. Each injection administered 200 ⁇ l of solution containing 2 ⁇ g of hepatitis B surface antigen absorbed on alum. Control mice were immunized with untreated liquid hepatitis B vaccine. Following a prime (day 0) and a boost immunisation (day 28), immune responses to the hepatitis B vaccine were determined with serum collected on day 42 in an ELISA. The antibody titers were determined by comparing to reference a serum.
  • the amount of alum in the total dry mass (50% or 12%) did not affect the potency of the dry powder. Neither of the spray-freeze-dried powders had a coagulation problem. This is significant that the spray- freeze-drying formulation preserves the potency of alum salt adjuvant at a very high concentrations (3% by weight).
  • alum coagulation is associated with the potency loss of alum vaccine when freeze-dried. It is believed that the large sizes of coagulated particles, which may fail to solubilize in vivo, can not be processed by the cells of the immune system and, thus, have no potency. More importantly, the process of the invention can prepare stable dry powders with alum containing vaccine without causing coagulation. It is believed that the quick freezing in the liquid nitrogen employed in the spray-freeze-drying process is critical for preventing the coagulation, thus preserving the vaccine potency.
  • HBsAg Hepatitis B surface antigen absorbed on alum hydroxide was used as a model antigen.
  • the immunogenicity of spray-freeze-dried powders was evaluated in mice following two different routes of immunisation, intramuscular injection using a needle and epidermal powder immunisation using a needleless powder delivery device.
  • the excipients for the spray-freeze-dried formulations are shown in Table 5. In this case, the spray-freeze-dried formulations used the combination of two sugars and one polymer. There was no amino acid/salt involved.
  • mice Female, 8 per group, 5-7 weeks old at the beginning of the study
  • the study design is shown in Table 6.
  • TM intramuscular
  • EPI epidermal
  • powders were administered to the shaved abdominal skin of mice using a re-chargeable powder delivery device.
  • Control mice were immunised with untreated liquid hepatitis B vaccine by intramuscular injection.
  • the spray freeze-dried diphtheria-tetanus-toxoid vaccine was prepared under the conditions as described in Table 3 and followed by compress/grind/sieve to generate particles with mean size of 20-38 ⁇ m and 38-53 ⁇ m in diameter.
  • the formulation information is summarised in Table 7. These particles do not have coagulation problems when reconstituted in water and examined under optical microscopy (data not shown).
  • the immunogenicity of spray-freeze-dried diphtheria-tetanus-toxoid vaccine was determined in guinea pigs (Charles River). Guinea pigs (4/group) were vaccinated on days 0 and 28 by administering powders to the abdominal skin using a powder delivery device. Each animal received 0.5 mg powders containing 1.5 Lf diphtheria toxoid and 1.5 Lf tetanus toxoid absorbed on 250 ⁇ g of aluminum phosphate. Control animals were vaccinated with untreated vaccine by intramuscular injection using a 23 V2 needle.
  • Serum antibody responses to diphtheria toxoid and tetanus toxoid were measured in an ELISA using sera collected on days 42.
  • the results of the immunogenicity study are shown in Figure 5.
  • Epidermal powder immunisation with spray-freeze-dried diphtheria toxoid absorbed on alum elicited antibody responses to each of the vaccine components and the tiers are comparable to that elicited by intramuscular injection of untreated vaccine.
  • the size of the spray-freeze-dried powders did not appear to affect the immunogenicity significantly since these powders did not have coagulation problem in vivo.

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Abstract

L'invention concerne une poudre fluide formant un gel, pouvant convenir comme vaccin, préparée par séchage par pulvérisation ou par lyophilisation avec pulvérisation d'une suspension aqueuse qui contient : un antigène adsorbé sur un sel d'aluminium ou sur un adjuvant de sel de calcium, un saccharide, un acide aminé ou un sel de celui-ci, et une matière colloïdale. La poudre, destinée à des applications vaccinales, peut aussi être préparée par lyophilisation avec pulvérisation d'une suspension aqueuse d'un tel adjuvant contenant un antigène adsorbé. L'invention concerne aussi des procédés permettant de former ces compositions de poudre, ainsi que des procédés d'utilisation des compositions dans une procédure de vaccination.
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JP2004532277A (ja) * 2001-06-08 2004-10-21 パウダージェクト ワクチンズ，インコーポレーテッド 噴霧凍結乾燥組成物
EP1458362A4 (fr) * 2001-11-19 2006-01-18 Becton Dickinson Co Compositions pharmaceutiques sous forme particulaire
AU2003234090A1 (en) 2002-04-11 2003-10-27 Medimmune Vaccines, Inc. Spray freeze dry of compositions for pulmonary administration
DE10339197A1 (de) * 2003-08-22 2005-03-24 Boehringer Ingelheim Pharma Gmbh & Co. Kg Sprühgetrocknete amorphe Pulver mit geringer Restfeuchte und guter Lagerstabilität
MX2007003726A (es) * 2004-09-28 2007-06-15 Johnson & Johnson Estabilizaci??n de agentes inmunologicamente activos con adyuvantes de alumbre.
AU2006261669B2 (en) * 2005-06-27 2011-07-07 Applied Thin Films, Inc. Aluminum phosphate based microspheres
CA2624503A1 (fr) 2005-10-04 2007-04-12 Alk-Abello A/S Formulation vaccinale solide
WO2007053781A2 (fr) * 2005-11-01 2007-05-10 Novartis Ag Compositions comprenant des antigenes adsorbes par du phosphate de calcium
JP2010502747A (ja) * 2006-09-08 2010-01-28 ベクトン・ディキンソン・アンド・カンパニー ミョウバン吸着ワクチンの安定粉末製剤
EP3590503A1 (fr) 2006-10-12 2020-01-08 The University of Queensland Compositions et procédés pour moduler les réponses immunologiques
CA2681567C (fr) 2007-03-22 2016-07-19 The Regents Of The University Of Colorado, A Body Corporate Procede de preparation d'une composition de vaccin sechee liee a un adjuvant immunologiquement actif
CA2694083A1 (fr) * 2007-07-26 2009-01-29 Sanofi Pasteur Limited Compositions antigene-adjuvant et procedes
CA2716399C (fr) 2008-03-05 2020-07-21 Pierre Chouvenc Procede de stabilisation d'un adjuvant contenant une composition de vaccin
AU2009299615B2 (en) 2008-10-02 2015-07-16 Pharmathene Inc. Anthrax vaccine formulation and uses thereof
GB2499479A (en) 2010-03-31 2013-08-21 Stabilitech Ltd Stabilisation of viral particles
ES2708989T3 (es) * 2010-03-31 2019-04-12 Stabilitech Biopharma Ltd Método de conservación de adyuvantes de alumbre y vacunas potenciadas con alumbre
DK2552478T3 (en) 2010-03-31 2017-03-27 Stabilitech Ltd EXCIPIENTS FOR STABILIZING VIRUS PARTICLES
BR112013010926A2 (pt) 2010-11-01 2016-08-23 Univ Technology Sidney agentes imunomoduladores e seus usos
AU2011336413B2 (en) 2010-12-02 2015-01-22 Oncolytics Biotech Inc. Liquid viral formulations
KR102023207B1 (ko) 2010-12-02 2019-11-25 온콜리틱스 바이오테크 인코포레이티드 동결건조 바이러스 제형
CN102133396B (zh) * 2011-03-16 2013-10-16 中国人民解放军第三〇二医院 一种疫苗注射剂及其制备方法
GB201117233D0 (en) 2011-10-05 2011-11-16 Stabilitech Ltd Stabilisation of polypeptides
EP2822540B1 (fr) * 2012-03-05 2020-04-08 De Staat der Nederlanden, Vert. Door de Minister van VWS Procédés et compositions pour la stabilisation de matériaux biologiques secs
KR101441539B1 (ko) * 2013-04-01 2014-09-18 재단법인 유타 인하 디디에스 및 신의료기술개발 공동연구소 흡습성이 개선된 접착제 조성물 및 이의 제조방법
CN103641886B (zh) * 2013-11-26 2015-07-29 精晶药业股份有限公司 一种丙谷二肽的精制方法
GB201406569D0 (en) 2014-04-11 2014-05-28 Stabilitech Ltd Vaccine compositions
GB2562241B (en) 2017-05-08 2022-04-06 Stabilitech Biopharma Ltd Vaccine compositions

Family Cites Families (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
FR2505657A1 (fr) * 1981-05-13 1982-11-19 Pasteur Institut Perfectionnements apportes aux agents de stabilisation de virus vivants pour la preparation de vaccins, et vaccins stabilises contenant lesdits agents de stabilisation
HU188847B (en) * 1983-02-22 1986-05-28 Human Oltoanyagtermeloe Es Kutato Intezet,Hu Process for producing liophylized combined vaccines
JPS6013718A (ja) * 1983-07-05 1985-01-24 Chemo Sero Therapeut Res Inst Ｂ型肝炎ワクチン
EP0676088A1 (fr) * 1992-12-28 1995-10-11 YU, Shih-Chiang Cellule de memoire a semiconducteur remanente
EP0678035B9 (fr) * 1993-01-08 2006-12-13 Csl Limited Preparations vaccinales
TW404844B (en) * 1993-04-08 2000-09-11 Oxford Biosciences Ltd Needleless syringe
US5902565A (en) * 1993-12-24 1999-05-11 Csl Limited Spray dried vaccine preparation comprising aluminium adsorbed immunogens
RU2096042C1 (ru) * 1996-11-06 1997-11-20 Виктор Иванович Ситьков Способ получения поливалентной вакцины против лептоспироза животных
CN1285753A (zh) * 1997-12-02 2001-02-28 鲍德杰克特疫苗公司 透皮给药微粒疫苗组合物

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO0193829A2 *

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CA2412197A1 (fr) 2001-12-13
JP2003535119A (ja) 2003-11-25
AU7537101A (en) 2001-12-17
NZ552576A (en) 2008-06-30
AU2001275371B2 (en) 2006-12-21
IL153241A0 (en) 2003-07-06
MXPA02012039A (es) 2003-06-06
NZ523103A (en) 2005-08-26
CN1438874A (zh) 2003-08-27
WO2001093829A2 (fr) 2001-12-13
KR20030020294A (ko) 2003-03-08
WO2001093829A3 (fr) 2002-06-13
BR0111494A (pt) 2004-01-13
AU2001275371B9 (en) 2007-06-07

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