WO2007138135A1 - Nanoparticules pour l'administration de molécules bioloquement actives - Google Patents

Nanoparticules pour l'administration de molécules bioloquement actives Download PDF

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Publication number
WO2007138135A1
WO2007138135A1 PCT/ES2007/000309 ES2007000309W WO2007138135A1 WO 2007138135 A1 WO2007138135 A1 WO 2007138135A1 ES 2007000309 W ES2007000309 W ES 2007000309W WO 2007138135 A1 WO2007138135 A1 WO 2007138135A1
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WIPO (PCT)
Prior art keywords
nanoparticles
thiamine
antigen
biologically active
biodegradable polymer
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PCT/ES2007/000309
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English (en)
Spanish (es)
Inventor
Hesham H. A. Salman
Carlos Gamazo De La Rasilla
Maite AGÜEROS BAZO
Juan Manuel Irache Garreta
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Instituto Científico Y Tecnológico De Navarra, S.A.
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Publication of WO2007138135A1 publication Critical patent/WO2007138135A1/fr

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    • 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
    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1641—Organic macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyethylene glycol, poloxamers
    • A61K9/1647—Polyesters, e.g. poly(lactide-co-glycolide)
    • 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
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5192—Processes
    • 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
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/513—Organic macromolecular compounds; Dendrimers
    • A61K9/5138—Organic macromolecular compounds; Dendrimers obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyvinyl pyrrolidone, poly(meth)acrylates
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders

Definitions

  • the invention relates to bioadhesive nanoparticles, based on a biodegradable polymer and thiamine, manufacturing processes, formulations containing them and their applications.
  • biodegradable polymeric nanoparticles have been proposed as new transport systems for drugs or biologically active molecules.
  • the most important characteristics that they offer to the biologically active molecule that they incorporate are: (i) protection against possible physical-chemical and / or enzymatic degradation (increase in their half-life in the organism); (ii) controlled release of the incorporated drug (sustained effects that reduce the number of doses); (iii) interaction / adhesion with the surface of the mucosa where absorption or drug action takes place (increased bioavailability); and (iv) in the case of vaccination, they facilitate the presentation of the incorporated antigen to the antigen presenting cells.
  • the oral route is the most convenient and popular route for drug administration.
  • bioavailability of a certain active molecule depends (i) on the characteristics of the drug molecule and its pharmaceutical form of administration, and (ii) on the physiological conditions present in the gastrointestinal tract, such as the presence of enzymes. proteolytic, peristaltic movements and presystemic metabolism.
  • the use of polymeric nanoparticles can be a good strategy to overcome some of these obstacles. In principle, these transporters have a large specific surface so that their interaction with the biological support (the gastrointestinal mucosa) is facilitated.
  • the control of drug release allows the effect of molecules with low biological half-lives to be prolonged over time.
  • nanoparticles can be captured by Peyer's plate cells and lymphoid tissue follicles. This phenomenon allows the drug to be directed towards the lymphatic route and, in the case of vaccines, to facilitate its antigen presentation.
  • conventional nanodeparticles based on biodegradable polymers have some important disadvantages with respect to their oral use: (i) some instability in gastrointestinal fluids, (ii) a low degree of intestinal absorption and (iii) a tropism or non-specific adhesion in the gastrointestinal mucosa.
  • a possible strategy to minimize these inconveniences associated with the use of conventional nanoparticles is based on the use of nanoparticles coated with certain ligands with specificity for certain mucosal receptors.
  • the main ligands proposed for the preparation of these nanoparticle-ligand conjugates have been, so far: (i) antibodies; (ii) adhesins, invasins or other bacterial proteins; (iii) carbohydrates; (iv) vitamin B 12 ; and lectins.
  • lectins appear to be the most versatile ligands. These molecules are proteins or glycoproteins, of non-immunogenic origin, capable of specifically recognizing sugars located in the glycoconjugates.
  • Another possible ligand may be thiamine.
  • Thiamine or vitamin Bl
  • the thiamine absorption process is quite complicated; although a mechanism based on the existence of a receptor (associated with a proton pump) that is located in the basolateral membranes of the jejuno and the ileum. Recently, it has been described that, in humans, the thiamine transporter is expressed throughout the entire gastrointestinal tract, although mostly in the apical membrane of the brush border tc ".
  • thiamine may be a good ligand to facilitate the binding or association of nanoparticles to the blood brain barrier and to interact with breast tumor cells.
  • thiamine may be a good ligand to facilitate the binding or association of nanoparticles to the blood brain barrier and to interact with breast tumor cells.
  • thiamine may be a good ligand to facilitate the binding or association of nanoparticles to the blood brain barrier and to interact with breast tumor cells.
  • thiamine may be a good ligand to facilitate the binding or association of nanoparticles to the blood brain barrier and to interact with breast tumor cells.
  • thiamine may be a good ligand to facilitate the binding or association of nanoparticles to the blood brain barrier and to interact with breast tumor cells.
  • thiamine may be a good ligand to facilitate the binding or association of nanoparticles to the blood brain barrier and to interact with breast tumor cells.
  • thiamine may be a good ligand to facilitate the binding or association of nanoparticles to the blood brain barrier and to interact with breast
  • particulate adjuvants in the form of emulsions, microparticles, ISCOMS or liposomes has been previously evaluated by various research groups [Singh and O ⁇ agan, Int. J. Parasitology, 33 (2003) 469-478].
  • antigen presenting cells The capture of antigens by "antigen presenting cells” is increased when they are associated with or included in polymer particles.
  • Biodegradable and biocompatible polyesters have been used in humans and animals for years as controlled antigen release systems.
  • micro- and nanoparticles are effective in inducing cellular and cytotoxic immune responses in mice.
  • oral immunization with microparticles induces potent mucosal and systemic immune responses against encapsulated antigens. This ability is a consequence of its internalization by specialized cells of the lymphoid tissue of the mucous membranes. Immunization via mucous membranes with different particulate systems has proven effective against different pathogens, such as Bordetella pertussis, Chlamidia trachomatis, Salmonella typhimurium and Brucella sp.
  • Specific allergen immunotherapy has been defined as repeated administration of allergens to patients with IgE-mediated health disorders, with the purpose of providing protection against allergic symptoms and inflammatory reactions associated with natural exposure to these allergens.
  • ThI ThI response
  • Th2 response a functional predominance of the ThI response with respect to the Th2 response
  • This modulation towards ThI is also applicable in other processes, such as control by vaccination against bacterial intracellular parasites (e.g., Brucella or Salmonellá).
  • Nanoparticles of copolymer of polyvinyl methyl ether and maleic anhydride (PVM / MA) [WO02 / 069938], optionally pegylated [ES2246694], useful as drug carriers have been described.
  • PVM / MA nanoparticles as immune stimulating substances has been described.
  • the doses should always be much higher (up to 200 times) than those normally used subcutaneously .
  • the gastrointestinal mucosa acts as a barrier very little permeable to the absorption of these macromolecules.
  • the object of the present invention is to provide nanoparticles that solve the aforementioned problems, that is, that can be administered orally and are stable and specific, that have good bioadhesive characteristics to interact with the mucous membranes, that are capable of transporting a wide group of active molecules that release the active molecule in a controlled manner.
  • these nanoparticles are intended to act as adjuvants in vaccines and immunotherapy, enhancing the immune response of antigens or allergens. It has now been found, surprisingly, that such problems can be solved by nanoparticles formed by a biodegradable polymer and thiamine.
  • nanoparticles comprising or consisting of (i) a copolymer of polyvinyl methyl ether (PVM) and maleic anhydride (MA) and (ii) thiamine, in addition to being easy to produce, provide excellent bioadhesion characteristics and size, which makes them suitable for administration through different pathways of biologically active compounds or molecules.
  • the nanoparticles provided by this invention that, optionally, contain a biologically active molecule, for example, an allergen or an antigen, have the ability to stimulate or enhance the immune response when administered to a subject, which allows its use in immunotherapy and vaccines.
  • the invention relates to nanoparticles comprising a biodegradable polymer and thiamine or its derivatives.
  • said biodegradable polymer is a copolymer of methyl vinyl ether and maleic anhydride (PVM / MA).
  • the thiamine is completely or partially covering the surface of the nanoparticles comprising said biodegradable polymer.
  • the weight ratio between thiamine and the biodegradable polymer is between 1:10 and 1: 500, preferably between 1:10 and 1: 100, more preferably around 1:40.
  • the nanoparticles provided by this invention can be used for the transport of biologically active molecules.
  • said nanoparticles can include one or more biologically active molecules, whose Chemical nature may vary within a wide range of possibilities, including, but not limited to, peptides, proteins, nucleic acids (eg, DNA, RNA, etc.), nucleosides, nucleotides, oligonucleotides, polynucleotides, etc.
  • Said biologically active molecule can be a drug or compound with therapeutic or diagnostic activity, eg, an antitumor agent, a central nervous system protector, a glucocorticoid, etc., an antigen for vaccination or an allergen for immunotherapy, among others. Therefore, the nanoparticles provided by this invention can be used in the preparation of pharmaceutical compositions.
  • the invention in another aspect, relates to a pharmaceutical composition
  • a pharmaceutical composition comprising said previously described nanoparticles and a biologically active molecule, wherein said biologically active molecule is a molecule capable of preventing, alleviating or curing a disease or a molecule with diagnostic application, together with a pharmaceutically acceptable carrier or excipient.
  • said composition is a pharmaceutical composition intended for administration orally or parenterally. If desired, said pharmaceutical composition may be in the form of a lyophilisate.
  • the invention in another aspect, relates to a vaccine or composition for immunotherapy comprising a therapeutically effective amount of the nanoparticles provided by this invention, previously described, and an antigen or allergen, together with a pharmaceutically acceptable carrier, adjuvant or excipient.
  • the invention in another aspect, relates to a product comprising, separately, a) an antigen or an allergen; and b) a composition comprising said nanoparticles based on a biodegradable polymer and thiamine, as a composition that enhances the immune response against said antigen or allergen, as a combination for simultaneous administration or sequence! to a subject, in the induction or stimulation of an immune response against said antigen or allergen in said subject.
  • the invention relates to the use of nanoparticles comprising a biodegradable copolymer and thiamine previously described in the preparation of a pharmaceutical composition for the selective stimulation of the ThI immune response, or in the preparation of a pharmaceutical composition for selective stimulation of the Th2 immune response, or in the development of a Pharmaceutical composition for balanced stimulation of Thl and Th2 immune responses.
  • the invention relates to a process for the production of nanoparticles comprising a biodegradable copolymer and thiamine previously described comprising: a) the desolvation of an organic solution comprising a biodegradable polymer, with a hydroalcoholic solution, for form the nanoparticles; b) simultaneous incubation of previously formed biodegradable polymer nanoparticles with thiamine in an aqueous solution; and c) the removal of organic solvents, whereby an aqueous suspension of nanoparticles comprising a biodegradable polymer and thiamine is obtained.
  • said nanoparticles can be stabilized by the use of crosslinking agents.
  • the concentration of the biodegradable polymer in said nanoparticles is between 0.001 and 10% w / v and that of thiamine between 0.001 and 5% w / v.
  • these can be incorporated either in the organic phase where the biodegradable polymer has previously dissolved, before desolvation, or, subsequently, in the aqueous suspension of the nanoparticles already formed so that it Produce your association.
  • FIG. 1 is a photograph of the result of a scanning electron microscopy analysis for a lyophilized sample of thiamine coated nanoparticles (T-NP).
  • Figure 2 is a diagram showing the fluorescent marker release
  • Figure 3 is formed by diagrams showing the distribution of nanoparticles in the gastrointestinal tract of laboratory animals, after oral administration of a 10 mg dose of nanoparticles labeled with RBITC.
  • Figure 3 a is a diagram showing the distribution of thiamine coated nanoparticles (T-NP);
  • Figure 3b is a diagram showing the distribution of control nanoparticles (NP);
  • Figure 3 c is a diagram showing the distribution of thiamine-coated nanoparticles (T-NP) mixed with 5 mg of free thiamine.
  • the X axis represents the different segments of the gastrointestinal tract [Stomach: Sto; portions of the small intestine: II, 12, 13, 14; blind: Ce]; the Y axis represents the adhered fraction of nanoparticles to the mucosa (mg); and the Z axis represents the time after administration (0.5, 1, 3 and 8 hours).
  • Figure 5 is a set of photographs that allow visualization of nanoparticles in normal ileum tissue (M) and in Peyer's plates (PP) by fluorescence microscopy, specifically, thiamine coated nanoparticles (T-
  • NP in the normal mucosa
  • control nanoparticles in normal mucosa
  • NP control nanoparticles in normal mucosa
  • the antibodies were quantified by ELISA starting with a 1:40 dilution of serum, followed by serial double dilutions. Immunization was performed on day 0 by single administration of either 20 ⁇ g of OVA in the different formulations subcutaneously (a, b), or 100 ⁇ g of OVA in the different formulations orally (c, d).
  • the formulations were OVA-T-NP (?), OVA-NP (?), And free OVA (?).
  • Antibody titers were determined in the animals' serum on days 0, 7, 14, 28 and 42.
  • the invention relates to nanoparticles, hereinafter nanoparticles of the invention, comprising (i) a biodegradable polymer and (ii) thiamine or a derivative thereof.
  • the nanoparticles of the invention have adequate physicochemical, bioadhesion and specificity characteristics when administered orally, which makes them into drug transport systems or antigen or allergen presenting systems of great interest.
  • the nanoparticles of the invention can prolong the residence time in the mucosa after oral administration or through another mucosa of the organism.
  • Said nanoparticles can be used to administer biologically active molecules and improve their bioavailability, including drugs with narrow absorption windows, as well as drugs originating in biotechnology and compounds or molecules used to enhance or induce immune responses in a subject.
  • nanoparticle refers to a structure formed by a biodegradable polymer modified on its surface by thiamine bonding, or a derivative thereof, which, optionally, can be crosslinked by the addition of a crosslinking agent.
  • the binding of thiamine, or a derivative thereof, to the biodegradable polymer can be a covalent bond.
  • the reaction between the biodegradable polymer and the thiamine, and, optionally, the subsequent crosslinking generates characteristic, independent and observable physical entities, the average size of which is less than 1 micrometer ( ⁇ m).
  • average size is meant the average diameter of the nanoparticle population that moves together in an aqueous medium. The average size of These systems can be measured by standard procedures known to those skilled in the art, and which are described, by way of illustration, in the experimental part that accompanies the examples described below.
  • the nanoparticles of the invention are characterized by having an average particle size of less than 1 ⁇ m, preferably having an average size between 1 and 999 nm, more preferably between 10 and 900 nm, even more preferably between 100 and 400 nm.
  • the average particle size is mainly influenced by the amount of thiamine, or its derivative, added (the greater the amount of thiamine or the derivative thereof increases the size of the nanoparticle), by the amount and molecular weight of the biodegradable polymer (the greater the size or molecular weight of the biodegradable polymer, the average size of the nanoparticle is increased), and by some parameters of the production process of said nanoparticles, such as the stirring speed and the temperature in the incubation stage with the aqueous phase It contains thiamine.
  • the nanoparticles of the invention comprise a biodegradable polymer and thiamine or a derivative thereof.
  • biodegradable refers to polymers that dissolve or degrade in a period of time that is acceptable for the desired application, in this case in vivo therapy, once they are exposed to a physiological solution. pH between 4 and 9, at a temperature between 25 ° C and 40 0 C.
  • biodegradable polymer known in the state of the art that results in the formation of nanoparticles
  • said biodegradable polymers include polyhydroxy acids, such as polylactic acid, polyglycolic acid, etc., and copolymers thereof, eg, poly (lactic-co-glycolic acid) [PLGA], etc .; polyanhydrides; polyesters; and polysaccharides, eg, chitosan, etc.
  • the molecular weight of said biodegradable polymer can vary within a wide range as long as it satisfies the established conditions of forming nanoparticles and being biodegradable.
  • the biodegradable polymer used is the copolymer of methyl vinyl ether and maleic anhydride in anhydride form (PVM / MA), commercially referred to as Gantrez® AN.
  • PVM / MA methyl vinyl ether and maleic anhydride in anhydride form
  • Gantrez® AN commercially referred to as Gantrez® AN.
  • PVM / MA has a molecular weight between 100 and 2,400 kDa, preferably between 200 and 2,000 kDa, more preferably between 180 and 250 kDa.
  • This biodegradable polymer (PVM / MA) can react with different hydrophilic substances, due to the presence of its anhydrous groups, without having to resort to the usual organic reagents (glutaraldehyde, carbodiimide derivatives, etc.) that possess an important toxicity.
  • the PVM / MA copolymer In an aqueous medium, the PVM / MA copolymer is insoluble, but its anhydride groups are hydrolyzed giving rise to carboxylic groups. The dissolution is slow and depends on the conditions in which it occurs. Due to the availability of functional groups in PVM / MA, covalent binding of molecules with nucleophilic groups, such as hydroxide or amino, takes place by simple incubation in an aqueous medium.
  • the nanoparticles of the invention comprise, in addition to the biodegradable polymer, thiamine or a derivative thereof.
  • Thiamine also called vitamin Bl, or 3 - [(4-amino-2-methyl-5-pyrimidinyl) methyl] -5- (2-hydroxyethyl) -4-methylthiazolium chloride, is a known product.
  • thiamine derivatives being understood as such, compounds structurally related to thiamine, such as their salts, among which are their addition salts, for example, their acid addition salts.
  • thiamine addition salts include thiamine hydrochloride, thiamine monophosphate chloride dihydrate; thiamine pyrophosphate; oxythiamine chloride hydrochloride; piritiamine hydrobromide, etc., all of them known compounds.
  • the nanoparticles of the invention comprise thiamine or thiamine hydrochloride.
  • the thiamine, or the derivative thereof, can be found totally or partially coating the surface of the nanoparticles comprising the biodegradable polymer.
  • the ratio thiamine (or derivative thereof): biodegradable polymer, by weight, in the nanoparticle of the invention may vary within a wide range; however, in a particular embodiment, said weight ratio is comprised between
  • nanoparticles of the invention with a ratio of approximately 0.025 mg thiamine / mg biodegradable polymer provides a very efficient adhesion (bonding) capability.
  • the nanoparticles of the invention may have a surface charge (measured by the Z potential) that varies depending on the structure of the biodegradable polymer and on the proportion thereof with respect to the thiamine or derivative thereof.
  • the contribution of the positive charge is attributed, among others, to the amino groups present in the thiamine or its derivative.
  • the magnitude of the surface charge of the nanoparticles of the invention can vary within a wide range.
  • nanoparticles of the invention have been obtained, based on PVM / MA copolymer and thiamine, with a Z potential of about -34.0 + 1.9 mV (Example 1, Table 1), while in another particular embodiment, nanoparticles of the invention (based on PVM / MA copolymer and thiamine) loaded with ovalbumin (OVA) have been obtained, with a Z potential of about -28.6 ⁇ 6.2 mV (Example 3, Table 3).
  • the nanoparticles of the invention can be obtained by conventional methods known to those skilled in the art.
  • the nanoparticles of the invention can be obtained by incubating the previously formed biodegradable polymer nanoparticles with an aqueous solution of thiamine or a derivative thereof, which makes it possible to obtain mostly biodegradable polymer nanoparticles in which the thiamine, or its derivative, is attached to the surface of the nanoparticles.
  • the biodegradable polymer present in the nanoparticles of the invention is a PVM / MA copolymer, the preparation of which is described, for example, in WO 02/069938.
  • the nanoparticles can be obtained from the biodegradable polymer by a process comprising the dissolution of said biodegradable polymer in an organic solvent (eg, acetone, etc.) and subsequent desolvation after the addition of an appropriate solvent, for example, a solvent or a mixture of solvents miscible with the solution of the biodegradable polymer, such as an ethanol-water mixture, obtaining a suspension of nanoparticles from which the organic solvents are removed by conventional methods, for example, by evaporation under reduced pressure, and, then water is added, so an aqueous suspension of nanoparticles is obtained.
  • an organic solvent eg, acetone, etc.
  • an appropriate solvent for example, a solvent or a mixture of solvents miscible with the solution of the biodegradable polymer, such as an ethanol-water mixture
  • the ratio, by volume, organic phase: hydroalcoholic solution (ethanol / water) can vary within a wide range; however, in a particular embodiment, said ratio is between 1: 1 and 1: 10 (v: v).
  • the biodegradable polymer nanoparticles are modified on their surface efficiently with thiamine or with a derivative thereof, by incubation, at room temperature, for an appropriate period of time.
  • an aqueous solution of thiamine or a derivative thereof is added to said previously obtained suspension of nanoparticles.
  • pharmaceutical grade water is used.
  • concentration of the biodegradable polymer and that of thiamine, or its derivative can vary within a wide range; however, in a particular embodiment, the concentration of the biodegradable polymer is between 0.001 and 10% w / v and that of thiamine, or between it, between 0.001 and 5% w / v.
  • the thiamine amino groups react with functional groups eventually present in the biodegradable polymer, which leads to the formation of bonds between the biodegradable polymer and thiamine.
  • the association of thiamine with the nanoparticles of the biodegradable polymer is evident due to the significant decrease in the negative surface charge of the nanoparticles (see, for example, Table 1).
  • the thiamine amino groups react with the anhydride groups of the PVM / MA copolymer, a reaction that can easily occur by simply incubating the thiamine with the aqueous suspension of the nanoparticles, which leads to bond formation.
  • the nanoparticles of the invention can be purified by conventional methods, for example, by centrifugation, ultracentrifugation, tangential filtration, or evaporation, including the use of vacuum.
  • a crosslinking agent can be added to improve the stability of the nanoparticles of the invention, as described in WO 02/069938.
  • crosslinking agents include diamine compounds, for example 1.3 diaminopropane, simple polysaccharides or saccharides, proteins and, in general, any molecule having functional groups. capable of reacting with the functional groups present in the biodegradable polymer, for example, anhydride groups present in the PVM / MA copolymer.
  • the nanoparticles of the invention can be lyophilized by conventional methods. From a pharmacological point of view, it is important to be able to have nanoparticles in lyophilized form since this improves their stability during long-term storage and conservation, in addition to reducing the volume of the product to be handled.
  • the nanoparticles of the invention can be lyophilized in the presence of a usual cryoprotectant agent such as glucose, sucrose, mannitol, trehalose, glycerol, lactose, sorbitol, polyvinyl pyrridone, etc., preferably, sucrose or mannitol; at a concentration within a wide range, preferably between 0.1% and 10% by weight.
  • the nanoparticles of the invention have a high ability to associate biologically active molecules, which makes them a drug transport system or presentation of very appropriate antigens and allergens. Therefore, in another aspect, the invention relates to a pharmaceutical composition comprising nanoparticles of the invention and at least one biologically active molecule.
  • said biologically active molecule will be inside the nanoparticle of the invention; however, it could happen that some biologically active molecule was also attached to the surface of the nanoparticle although most of said molecules will be inside (e.g., encapsulated) of the nanoparticles of the invention.
  • biologically active molecule refers to any substance used in the treatment, cure, prevention or diagnosis of a disease or that used to improve the physical and mental well-being of humans and animals.
  • said term includes both drugs and antigens and allergens.
  • the nanoparticles of the invention can incorporate one or more biologically active molecules regardless of the solubility characteristics thereof. The association capacity will therefore depend on the incorporated molecule, but, in general, said capacity is high for both hydrophilic molecules and molecules of marked hydrophobic character.
  • the pharmaceutical composition of the invention comprises nanoparticles of the invention containing one or more different drugs.
  • Illustrative, non-limiting examples of such drugs include analgesic, anti-inflammatory, antitumor, neuroprotective, antiallergic, anti-asthmatic, antibiotic agents (eg, antibacterial, antifungal, antiviral, antiparasitic, etc.), pulmonary surfactants, etc.
  • the pharmaceutical composition of the invention comprises nanoparticles of the invention containing one or more different antigens for vaccination purposes or one or more different allergens for immunotherapeutic purposes.
  • the term "antigen" refers to any substance capable of being recognized by the immune system of a subject and / or capable of inducing in a subject a humoral immune response or a cellular immune response that leads to the activation of B and / or T lymphocytes when introduced into a subject; by way of illustration, said term includes any immunogenic, native or recombinant product, obtained from a higher organism or from a microorganism, for example, a bacterium, a virus, a parasite, a protozoan, a fungus, etc., which contains one or more antigenic determinants, for example, structural components of said organisms; toxins, for example, exotoxins, etc.
  • antigen includes: "microbial” antigens, that is, microorganism antigens, including, but not limited to, viruses, bacteria, fungi and infectious parasites; said antigens include the intact microorganism as well as parts, fragments and derivatives thereof, either of natural or artificial origin, as well as synthetic or recombinant products that are identical or similar to the natural antigens of a microorganism and induce a specific immune response for that microorganism; in this sense, a compound is similar to a natural antigen of a microorganism if it induces an immune response (humoral and / or cellular) like that of the natural antigen of that microorganism; said antigens are used routinely by those skilled in the art; Y “tumor” antigens, that is, substances, for example, peptides, associated with a tumor or a cancer ("microbial" antigens, that is, substances, for example, peptides, associated with a tumor or a cancer ("microbial" antigen
  • allergen refers to a substance to which a subject is sensitive and causes an immune reaction, for example, allergenic extracts of pollens, allergenic extracts of insects, allergenic extracts of food or products.
  • food components present in saliva, tweezers or stingers of insects that induce a sensitivity reaction in a subject, components present in plants that induce a sensitivity reaction in a subject, etc., for example, protein extracts of pollens, such as grass pollen, allergic extracts of perennial Lolium, allergic extracts of olea (olive), etc .; Protein extracts of insects, such as dust mites, etc .; allergenic extracts of food components, etc.
  • any allergen can be used in the preparation of the nanoparticles loaded with allergen of the composition of the invention; however, in a particular embodiment, said allergen is ovalbumin (OVA), a protein widely used as an experimental allergenic model.
  • OVA ovalbumin
  • Illustrative, non-limiting examples of said biologically active molecules that may contain the nanoparticles of the invention include bacterial: cytoplasmic, periplasmic, cell envelope antigens (eg, inner membrane proteins, outer membrane proteins, lipopolysaccharides and mixed complexes, proteins associated to the cell wall, etc.), etc .; surface structure antigens (eg, fimbriae, glycocalyx, flagellar, etc.), including those of intracellular pathogens, such as Brucella sp., Salmonella sp., etc .; eukaryotic microorganism antigens, both soluble and superficial; viral antigens, for example, matrix, capsid, envelope, internal (including enzymatic), allergens of animal species (mites, etc.), of plants (grasses, etc.), etc.
  • cell envelope antigens eg, inner membrane proteins, outer membrane proteins, lipopolysaccharides and mixed complexes, proteins associated to the cell wall, etc.
  • said biologically active molecule is a polysaccharide, a protein, a peptide or a lipid.
  • said biologically active molecule is a nucleic acid (eg, DNA, RNA, etc.), a nucleoside, a nucleotide, an oligonucleotide, a polynucleotide, etc.
  • the nanoparticles of the invention are used to modify the distribution of the associated biologically active molecule and / or of the conventional nanoparticles when administered by a route that gives access to some mucosa of the organism (including oral, rectal, nasal, vaginal or ocular).
  • pharmaceutical compositions include any liquid composition (suspension or dispersion of the nanoparticles) for oral, oral, sublingual, topical, ocular, nasal, vaginal or parenteral administration; any composition in the form of gel, ointment, cream or balm for topical, ocular, nasal or vaginal administration; or any solid composition (tablets, capsules) for oral administration.
  • the pharmaceutical composition is administered orally.
  • said pharmaceutical composition is administered parenterally.
  • the pharmaceutical compositions described will comprise the excipients suitable for each formulation.
  • binders, disintegrants, lubricants, fillers, enteric coating, etc. will be included if necessary.
  • Oral solid formulations are prepared in conventional manner by mixing, dry or wet granulation and incorporating the nanoparticles of the invention.
  • the pharmaceutical compositions may also be adapted for parenteral administration, in the form of, for example, sterile lyophilized solutions, suspensions or products, in the appropriate dosage form; in this case, said pharmaceutical compositions will include suitable excipients, such as buffers, surfactants, etc. In any case, the excipients will be chosen based on the pharmaceutical form of administration selected.
  • the proportion of the biologically active molecule incorporated into the nanoparticle of the invention can vary within a wide range, for example, it can be up to 25% by weight with respect to the total weight of the nanoparticles. However, the appropriate proportion will depend in each case on the biologically active molecules incorporated.
  • incorporation of the biologically active molecule into the nanoparticles of the invention can be done as described in WO 02/069938, by incorporation into the solution of the biodegradable polymer before the formation of nanoparticles, or subsequently added to the aqueous suspension of the nanoparticles already formed.
  • a) Hydrophobic biologically active molecules addition in the organic phase (eg, acetone) and joint incubation / solubilization with the biodegradable polymer for a period variable time (up to 1 hour) under agitation (mechanical, magnetic or ultrasonic stirrer); and b) Biologically active bi-hydrophilic molecules: either by addition in the organic phase (eg, acetone) and joint incubation with the biodegradable polymer for a variable period of time (up to 1 hour) under stirring (mechanical, magnetic or ultrasonic agitator) until obtaining a fine suspension in the organic solvent [this procedure has been used successfully to encapsulate a model protein (ovalbumin, about 44 kDa protein); incorporation was efficient allowing high encapsulation of the model protein; or by addition in the aqueous phase to incubate with the preformed nanoparticles (this is the case used to encapsulate the
  • the nanoparticles of the invention produce a stimulatory or potentiating effect of the immune response after administration to a subject and can therefore be used as an adjuvant in vaccines or in immunotherapy
  • the nanoparticles of the invention have the ability to stimulate the two immune response pathways (ThI or Th2), so it can be used in vaccine or immunotherapeutic formulations.
  • a vaccine formulation it is generally required, depending on the pathogenic mechanisms of the organism from which the antigen (intracellular or extracellular, toxin dependent, scourge dependent, etc.), a stimulation of the ThI response ( intracellular, as in the case of Brucella, Salmonella, etc.) or Th2 response (extracellular, as in the case of Staphylococcus, Escherichia coli, enterotoxigenic bacteria, etc.).
  • ThI response intracellular, as in the case of Brucella, Salmonella, etc.
  • Th2 response extracellular, as in the case of Staphylococcus, Escherichia coli, enterotoxigenic bacteria, etc.
  • a tolerance induction is required for an immunotherapeutic formulation by the presence of the two types of response, that is, inducing ThI and Th2 responses in a balanced manner.
  • the formulations will contain nanoparticles of biodegradable polymer and thiamine, and encapsulated therein or totally or partially coating the surface thereof the antigen or
  • the invention relates to a vaccine or composition for immunotherapy comprising a therapeutically effective amount of the nanoparticles of the invention and an antigen or allergen, together with a pharmaceutically acceptable carrier or excipient.
  • the antigen or allergen may be contained within said nanoparticles and / or at least partially covering the surface of said nanoparticles.
  • the allergen or antigen present in the nanoparticles of the invention may be at least partially covering the surface of said nanoparticles and / or contained within said nanoparticles.
  • said allergen or antigen is coating all or part of the surface of said nanoparticles.
  • said vaccine or composition for immunotherapy can be found in any pharmaceutical form of administration by any appropriate route, for example, oral, rectal, nasal, sublingual, etc.
  • said vaccine or composition for immunotherapy is in a pharmaceutical form of oral administration
  • said vaccine or composition for immunotherapy is in a form Pharmaceutical administration parenterally, for example, intramuscularly (im), subcutaneously (s. ⁇ ), intravenously (iv), intraperitoneally (ip), intradermally (id), etc.
  • the dose of nanoparticles loaded with an antigen or with an allergen may vary within a wide range, for example, between about 0.01 and about 10 mg per kg of body weight, preferably, between 0.1 and 2 mg per kg of body weight.
  • the nanoparticles of the invention do not incorporate the antigen or allergen, but these are empty and are administered in combination with vaccine or immunotherapeutic compositions containing the antigen or allergen, respectively, producing a stimulatory effect of the immune response. after administration of said vaccine or immunotherapeutic composition and empty nanoparticles.
  • a further aspect of the invention is a product comprising, separately, a) an antigen or an allergen; and b) a composition comprising said nanoparticles based on a biodegradable polymer and thiamine, as a composition that enhances the immune response against said antigen or allergen, as a combination for simultaneous or sequential administration to a subject, in the induction or stimulation of a response immune against said antigen or allergen in said subject.
  • the combined administration of said vaccine or immunotherapeutic composition and of the empty nanoparticles can be carried out simultaneously or sequentially, separated in time, in any order, that is, the vaccine or immunotherapeutic composition can be administered first and, subsequently, the empty nanoparticles or vice versa.
  • said vaccine or immunotherapeutic composition and said empty nanoparticles can be administered simultaneously.
  • the vaccine or immunotherapeutic composition and the empty nanoparticles can be administered in the same composition or in different compositions.
  • PVM / MA and coated with thiamine that, optionally, contain an antigen and demonstrate the ability of said nanoparticles to specifically adhere to the gastrointestinal mucosa and, if necessary, act as an adjuvant in immunotherapy or vaccination.
  • the general method of production of thiamine coated PVM / MA nanoparticles is a variant of the general procedure described above [Arbos et al., J. Control. Relay, 83 (2002) 321-330].
  • This process comprises dissolving said copolymer in acetone followed by the addition of ethanol. A similar volume of water was added to the resulting solution, so that nanoparticles formed instantly in the medium, under the appearance of a milky suspension. Then, the organic solvents (ethanol and acetone) were removed by evaporation under reduced pressure, the particles being in a stable aqueous suspension.
  • the thiamine was incorporated into the aqueous phase that facilitates the desolvation of the polymer, leaving the reaction to act for a certain time.
  • the next step consisted of the incubation between the formed nanoparticles and the thiamine.
  • the crosslinking agent which, in this case, has been 1,3-diaminopropane (10 ⁇ g / mg polymer) can be added.
  • the purification of the nanoparticles was carried out by ultracentrifugation. Finally, the purified nanoparticles were lyophilized for long-term storage and preservation.
  • the size and zeta potential of the nanoparticles was determined in a Zetamaster (Malvern Instruments / Optilas, Spain).
  • the morphology of the nanoparticles was studied by scanning electron microscopy in a Zeiss DSM940 (Oberkochen, Germany).
  • the amount of thiamine associated with the nanoparticles was quantified by high performance liquid chromatography (HPLC), according to the procedure described by [Batifoulier et al., J. Chromatogr. B Analyt. Technol Biomed Life Sci., 816 (2005) 67-72].
  • the analysis was carried out on a model 1100 series LC chromatograph (Agilent, Waldbornn, Germany) coupled to a diode-array ultraviolet (UV) detection system.
  • the data was analyzed on a Hewlett-Packard computer using the Chem-Station G2171 program.
  • a Zobrax NH 2 reverse phase column (4.6 mm x 150 mm; 5 ⁇ m; Agilent) heated to 40 0 C was used.
  • FGS gastric medium
  • FIS simulated intestinal medium
  • the protein content [active ingredient (in this case ovalbumin)] encapsulated in the thiamine-coated nanoparticles was determined by testing with microbicinconinic acid (Micro BCA, Pierce, USA). For this, the nanoparticles were digested, at 37 ° C for 24 h, with a solution of sodium lauryl sulfate (3%) in 0.1 M NaOH. Polyacrylamide gel electrophoresis with sodium dodecyl sulfate (SDS-PAGE) was used for corroborate the content in OVA.
  • OVA-T-NP nanoparticles
  • DMF dimethylformamide
  • acetone 1: 1 v / v
  • the amount of OVA was estimated by calculating the average density of the band in the SDS-PAGE gel using the Micro Lnage ® program (Version 4.0; Olympus Optical Co., USA).
  • the calibration line was performed with OVA (0.25-2.5 ⁇ g protein / well).
  • AUC adh or area under the curve represents the fraction adhered against time (expressed as a quantity of adherent marker with respect to time) and was evaluated by the trapezoid method up to I 2 (the last sampling point), and allows quantify the intensity of the bioadhesive phenomenon.
  • T max represents the time at which maximum adhesion occurs.
  • MRT adh is the average residence time of the adhered fraction of nanoparticles and allows the relative duration of adhesive interactions to be evaluated, taking as a limit the last sampling point. The calculations were performed using the WinNonlin 1.5 program (Pharsight Corporation, USA).
  • Fluorescence microscopy studies The visualization of the distribution of thiamine coated nanoparticles was carried out by fluorescence microscopy.
  • the laboratory animals received an oral dose of 10 mg of nanoparticles labeled with RBITC.
  • the animals were sacrificed and different portions of the small intestine were removed and washed with PBS. Said portions of approximately 0.5 cm in length were treated with Sakura Tissue-Tek Oct® Compound (Sakura, Holland) and frozen in liquid nitrogen.
  • Different tissue samples were cut in sections of 5 ⁇ m in a cryostat (2800 Frigocut E, Germany), fixed to supports coated with poly-L-lysine (Sigma, Spain) and stored at - 20 0 C before viewing by microscopy of fluorescence.
  • mice (20 ⁇ 1 g) (Har ⁇ an, Spain) were divided into 6 different groups of 10 animals each.
  • the administration strategy was based on the single administration of a subcutaneous or oral dose.
  • each animal received a volume of 200 ⁇ L containing 100 ⁇ g of ovalbumin, free or encapsulated in thiamine coated nanoparticles (OVA-T-NP) or in conventional nanoparticles (OVA-NP).
  • OVA-T-NP thiamine coated nanoparticles
  • OVA-NP thiamine coated nanoparticles
  • OVA-NP thiamine coated nanoparticles
  • OVA-NP conventional nanoparticles
  • Serum anti-OVA antibodies were analyzed by ELISA with anti-IgG t and anti-IgG 2a conjugates (Sigma-Aldrich Chemie, Germany). Briefly, 96 well plates (EB, Thermo Labsystems, Vantaa, Finland) were used to perform this test, where 1 ⁇ g of ovalbumin dissolved in carbonate-bicarbonate regulatory solution (0.05 M, pH 9.6) was fixed ) at 4 ° C for 24 hours. Subsequently, the plates were blocked by incubation for 1 h at 37 ° C with 1% bovine serum albumin in a solution of 0.05% PBS-Tween 20 (PBS-T).
  • PBS-T PBS-Tween 20
  • Stool anti-OVA antibodies were analyzed by ELISA with anti-IgA conjugates.
  • OVA after coating the plates with OVA, they were blocked by adding, and incubating for 1 h at room temperature, 200 ⁇ L of a PBS-T solution containing 3% skim milk.
  • the stool extract was added to two plates and serial dilutions were made in PBS-T. Said plates were incubated for 4 h at 37 ° C. Finally, after washing, the wells were incubated with the peroxidase-labeled anti-IgA conjugate (Nordic rmmunological Labs, The Netherlands).
  • Bioadhesion data and physicochemical characteristics were compared using the Mann-Whitney U-test non-parametric test and the t-test.
  • Table 1 summarizes the main physicochemical characteristics of the nanoparticles obtained.
  • the control nanoparticles (NP) show a size close to 200 nm with a negative surface charge of -51 mV.
  • the thiamine-coated nanoparticles (T-NP) were significantly higher (close to 400 nm) and showed a significantly less negative zeta potential.
  • These nanoparticles (T-NP) showed an associated amount of thiamine of 15 ⁇ g / mg nanoparticles (Table 1).
  • Table 1 summarizes the main physicochemical characteristics of the nanoparticles obtained.
  • the control nanoparticles (NP) show a size close to 200 nm with a negative surface charge of -51 mV.
  • the thiamine-coated nanoparticles (T-NP) were significantly higher (close to 400 nm) and showed a significantly less negative zeta potential.
  • These nanoparticles (T-NP) showed an associated amount of thiamine of 15
  • the thiamine-coated nanoparticles showed a very homogeneous population of spherical particles ( Figure 1).
  • Figure 3 shows the distribution in the gastrointestinal tract of the adhered fractions of thiamine-coated nanoparticles (T-NP) ( Figure 3 a) or control nanoparticles (NP) ( Figure 3b). It can be seen that, within 30 minutes of the administration of the nanoparticle formulations, both types (NP and T-NP) showed the same ability to adhere to the mucosa of the gastrointestinal tract, mainly in the stomach area and the upper regions of the small intestine (duodenum). However, 1 h after administration of the nanoparticles, the thiamine-coated nanoparticles (T-NP) showed significantly greater bioadhesive capacity and a more homogeneous distribution than the control nanoparticles (NP).
  • the amount of thiamine-coated nanoparticles (T-NP) found adhered to the mucosa was approximately 45% of the administered dose (Figure 3).
  • the thiamine-coated nanoparticles (T-NP) showed maximum adhesion (65% of the administered dose) and an important tropism for the animal's ileum (portions 13 and 14 in Figure 3 to).
  • Bioadhesion curves were obtained by representing the total amount of nanoparticles adhered to the gastrointestinal tract as a function of time after administration (Figure 4).
  • the comparison of bioadhesion profiles for the different formulations allows us to observe that the control nanoparticles (NP) show a maximum of bioadhesion 30 minutes after their administration [Figure 4, (?)]. Subsequently, the amount of control nanoparticles (NP) attached is falling over time.
  • T-NP thiamine-coated nanoparticles
  • AUC at dh parameter that measures the intensity of bioadhesive interactions
  • NP control nanoparticles
  • TNP-COM represents the values obtained when the thiamine coated nanoparticles (T-NP) were co-administered with 5 mg of the free vitamin
  • AUC a dh (mg.h): Area under the bioadhesion curve
  • FIG. 5 shows several photographs that allow us to observe the distribution of nanoparticles in ileum samples, 2 hours after oral administration of 10 mg of nanoparticles in rats.
  • Control nanoparticles (NP) were mainly detected in the outer layer of the ileum (mucus that covers the mucosa), confirming the low affinity of these transporters for the intestinal mucosa ( Figure 5b).
  • thiamine-coated nanoparticles (T-NP) were widely distributed and showed a very large affinity for the micro villi of the intestine ( Figure 5a).
  • T-NP thiamine-coated nanoparticles
  • OVA ovalbumin
  • OVA OVA was dispersed in 2 mL of water with the help of ultrasound (Microson TM) or in an ultrasonic bath for 1 minute under cooling.
  • This suspension was added to a solution of 100 mg of the copolymer of methyl vinyl ether and maleic anhydride (PVM / MA) [Gantrez® AN 119] in 3 mL of acetone.
  • PVM / MA copolymer of methyl vinyl ether and maleic anhydride
  • 10 mL of ethanol and 10 mL of deionized water were added. The resulting mixture was allowed to homogenize for 5 minutes.
  • the nanoparticle suspension was evaporated under reduced pressure (Büchi R-144, Switzerland) until both organic solvents were removed and the final volume was adjusted with water to 10 mL.
  • the nanoparticles obtained have encapsulated OVA (OVA-NP).
  • OVA-NP encapsulated OVA
  • the resulting suspension was subjected to purification by ultracentrifugation (20 minutes at 27,000 x g). The supernatants were removed and the residue was resuspended in water or in a 5% aqueous sucrose solution. Finally, the resulting nanoparticle suspension was lyophilized, thereby keeping all its properties intact.
  • the resulting suspension was subjected to magnetic stirring for 1 hour at room temperature and purified by centrifugation at 27,000 xg for 20 minutes, collecting the supernatants to quantify the thiamine bound to the nanoparticles. Subsequently, 100 ⁇ L of a 1% v / v solution of 1,3-diaminopropane was added, subjecting the whole to magnetic stirring for 5 minutes. The resulting suspension was subjected to purification by ultracentrifugation (20 minutes at 27,000 xg). The supernatants were removed and the residue was resuspended in water or in a 5% aqueous sucrose solution. Finally, the resulting nanoparticle suspension was lyophilized, thereby keeping all its properties intact.
  • Table 3 summarizes the main physicochemical characteristics of the resulting nanoparticles (OVA-NP and OVA-T-NP).
  • OVA-T-NP formulation showed a significantly larger size (412 nm) than the control nanoparticles (280 nm).
  • the presence of OVA slightly affects the amount of thiamine attached to the surface of the nanoparticles.
  • the presence of this protein (OVA) decreased by 20% the amount of vitamin (thiamine) bound (12 ⁇ g / mg vs 15 ⁇ g / mg).
  • the amount of encapsulated OVA was determined, in this case, by densitometry of the bands of this protein in the SDS-PAGE gel, using the Microlmage® Version 4 program.
  • the amount of OVA was, for OVA-NP and OVA-T - NP, of 12.1 ⁇ 1.4 and 11.6 ⁇ 2.3 ⁇ g / mg nanoparticles, respectively.
  • Figure 6 shows the titers of IgG2a and IgGl after oral or subcutaneous immunization of Balb / C mice with a single dose of the different treatments (OVA-NP, OVA-T-NP and free protein).
  • Table 4 summarizes the values of the areas under the curves of the immune response.
  • AUCs areas under curves
  • AUCs are expressed as arbitrary units (title x time)
  • the OVA-NP nanoparticles After oral administration, the OVA-NP nanoparticles also showed a predominantly Th2 type profile (Figure 6).
  • the oral ThI response was very low (AUC of 1.5; AUQ h2 is 7 times higher than AUCa 11 , see Table 4) and, at the same time, the antibody response induced by oral administration was lower than that described after subcutaneous administration.
  • ThI-type responses may be related to the high tropism of the T-NP nanoparticles by the regions. distal of the small intestine, as well as its capture by the Peyer plates rich in antigen presenting cells ( Figures 3-5).
  • FIG. 7 shows the evolution of anti-OVA antibodies of the IgA type in the feces of mice immunized by the s.c. or oral
  • immunization with nanoparticles regardless of the route of administration, induced the production of high levels of IgA.
  • thiamine-coated nanoparticles (T-NP) induced the secretion of significantly higher levels of intestinal IgA than conventional nanoparticles.
  • this difference was 64 times greater for the OVA-T-NP nanoparticles than for the OVA-NP nanoparticles.
  • This phenomenon may be related to the effective capture of the T-NP nanoparticles by the Peyer plates of the gastrointestinal tract and their passage to the lymphocytes responsible for the synthesis and secretion at the level of IgA mucous membranes.
  • T-NP thiamine-coated nanoparticles
  • these polymeric transporters are capable of reaching Peyer's plates and potentiating a high induction of antibodies against the transported antigen.
  • the response that is generated is humoral (Th2) and cellular (ThI), which may be of interest for vaccination against numerous pathogens and for immunotherapy for the treatment of allergies.

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Abstract

L'invention concerne des nanoparticules pour l'administration de molécules biologiquement actives qui comprennent un polymère biodégradable, de préférence le copolymère de méthyl vynyl éther et anhydride maléique (PVM/MA) et thiamine. Ces nanoparticules faciles à produire et dotées d'excellentes caractéristiques de bioadhésion, de taille et de potentiel zeta sont adéquates pour l'administration de molécules actives. Le procédé d'obtention des nanoparticules s'effectue par incubation simple pendant une courte période de temps avec les nanoparticules préformées en un milieu aqueux. La molécule biologiquement active peut être ajoutée lors de la formation des nanoparticules par désolvatation ou après modification avec la thiamine. Administrées par voie orale, ces nanoparticules entrent en interaction bioadhésive avec la muqueuse gastro-intestinale qui comprend aussi le tissu lymphoïde associé aux muqueuses (plaques de Peyer), ce qui s'avère utile pour augmenter la biodisponibilité orale de différents médicaments ou pour induire des réponses immunitaires au niveau des muqueuses. Ainsi, lorsque les nanoparticules contiennent un antigène dans leur matrice, le mélange obtenu est apte à induire de fortes réponses immunitaires face à l'antigène encapsulé. Cet effet stimulant la réponse immunitaire est utile pour les vaccins et l'immunothérapie.
PCT/ES2007/000309 2006-05-26 2007-05-25 Nanoparticules pour l'administration de molécules bioloquement actives WO2007138135A1 (fr)

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US10029007B2 (en) 2011-10-05 2018-07-24 Stabilitech Biopharma Ltd Stabilisation of polypeptides
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