Nanogels containing bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules

Abstract

Novel nanogels containing one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules and methods for their manufacture. Nanogel-coated biomaterial implants, medical devices or bioprostheses and methods for their manufacture.

Description

[Technical Field]

[0001] The present invention relates to nanogels comprising bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, in particular derivatives of vitamin E. The invention also relates to medical devices, biomaterial implants or bioprostheses coated with the nanogels, to methods for making such nanogels and to methods for coating medical devices, biomaterial implants or bioprostheses, in particular catheters.

[0002] Most medical devices, biomaterial implants, or bioprostheses pose biocompatibility problems. Importantly, implantation of a foreign body into the vascular system activates the contact pathway of coagulation, which can lead to thrombotic complications. For example, the surface roughness of a medical device, biomaterial implant, or bioprosthesis is an important factor affecting thrombogenicity.

[0003] Medical devices, biomaterial implants or bioprostheses can also become infected, and treatment of such infections generally requires the administration of antibiotics that target the causative bacteria.

[0004] Furthermore, the rise in antibiotic resistance has increased the demand for antibiotics that exhibit both antibacterial and anti-antibiotic effects, a demand that cannot be met by conventional one-target, one-molecule approaches, necessitating other approaches such as multi-target antibiotics.

[0005] Nanogels are known in the art. WO 2018 / 122318 A1 describes nanogels composed of a first hydrophilic polymer or copolymer having catechol groups crosslinked with a second hydrophilic polymer having one or more reactive moieties, the nanogel also containing a bioactive molecule, a therapeutic molecule or a drug.

[0006] Such nanogels can be directly immobilized or attached to the surface of medical devices, biomaterial implants, or bioprostheses to release bioactive molecules, therapeutic molecules, or drugs. However, a challenge in using such nanogels is the loading of water-insoluble or hydrophobic bioactive molecules, therapeutic molecules, or drugs, which leads to irregular release kinetic profiles and lower pharmacological efficacy.

[0007] Furthermore, after release is initiated, such nanogels release only low levels of hydrophobic bioactive molecules, therapeutic molecules or drugs, therefore, immediate pharmacological effects cannot be achieved quickly, and bacterial adhesion to medical devices, biomaterial implants or bioprostheses cannot be completely prevented. Summary of the Invention

[0008] The present inventors have discovered improved nanogels comprising one or more bioactive molecules, therapeutic molecules or drugs together with an amphiphilic molecule, preferably a vitamin E derivative or most preferably an amphiphilic cyclodextrin; the nanogels exhibit improved antibacterial efficacy and anti-antibiotic resistance.

[0009] The present invention surprisingly and advantageously provides more efficient loading of hydrophobic bioactive molecules, therapeutic molecules, or drugs in nanogels and a steady, gradual, and continuous release over several weeks. Hydrophobic bioactive molecules, therapeutic molecules, or drugs can be combined with hydrophilic bioactive molecules, therapeutic molecules, or drugs in such nanogels and released simultaneously. Furthermore, nanogels containing bioactive molecules, therapeutic molecules, or drugs advantageously remain stable over time in liquid suspension by maintaining their structural integrity and uniform distribution in a dispersed state, particularly in aqueous media.

[0010] Thus, the nanogels of the present invention are for use in the treatment or prevention of bacterial infections, particularly for topical administration to a host mammal.

[0011] The use of such novel nanogels in coatings for medical devices, biomaterial implants or bioprostheses, particularly catheters, also provides improved medical devices, biomaterial implants or bioprostheses.

[0012] Medical devices, biomaterial implants or bioprostheses coated with the nanogels of the present invention advantageously provide a more homogeneous, hydrophilic and smooth surface and content uniformity, leading to more reproducible release kinetics of hydrophobic bioactive molecules, therapeutic molecules or drugs from the coated medical device, biomaterial implant or bioprosthesis and subsequent improved pharmacological efficacy.

[0013] At the same time, medical devices, biomaterial implants or bioprostheses coated with the nanogels of the present invention advantageously reduce cell adhesion to the surface of the medical device, biomaterial implant or bioprosthesis, especially when the nanogel comprises a combination of a bacteriostatic agent and an antivirulence or bactericidal agent. DETAILED DESCRIPTION OF THE INVENTION

[0014] The present invention relates to novel nanogels, which are three-dimensional crosslinked particles with submicron particle size that offer larger cargo spaces that can be used to incorporate bioactive molecules, therapeutic molecules or drugs encapsulated by amphiphilic molecules, particularly vitamin E derivatives or amphiphilic cyclodextrins.

[0015] Such nanogels for use in the treatment and prevention of bacterial infections, particularly bacterial pathogenicity, are particularly useful for topical administration to a host mammal in need of such treatment.

[0016] Such nanogels can also be fixed or attached onto the surface of any biomaterial or medical device, be it metallic or polymeric, or onto bioprostheses, thereby helping to reduce or prevent infection and improve the biocompatibility and hemocompatibility of temporarily or permanently implanted materials, maintaining their functionality and increasing their durability.

[0017] According to a first aspect, the present invention provides a compound of formula (1) [ka] [wherein x is an integer greater than 1, preferably 1 to 100] It is composed of poly(methacrylamide) having a quinoline group represented by the formula: A nanogel, wherein the poly(methacrylamide) is crosslinked with a polymer having primary or secondary amine groups; the nanogel comprises one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules; The nanogel is provided. The amphiphilic molecule is preferably a vitamin E derivative, more preferably D-α-tocopheryl polyethylene glycol succinate. The amphiphilic molecule can also be an amphiphilic cyclodextrin, preferably hydroxypropyl-β-cyclodextrin.

[0018] Alternatively, the present invention also provides a compound represented by formula (6): [ka] [wherein n is an integer greater than 1, preferably 1 to 100] It is composed of poly(vinylquinoline) represented by A nanogel, wherein the poly(vinylquinoline) is crosslinked with a polymer having primary or secondary amine groups; the nanogel comprises one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules; The nanogel is provided. The amphiphilic molecule is preferably a vitamin E derivative, more preferably D-α-tocopheryl polyethylene glycol succinate. The amphiphilic molecule can also be an amphiphilic cyclodextrin, preferably hydroxypropyl-β-cyclodextrin.

[0019] Alternatively, the present invention also provides nanogels composed of a combination of both quinoline polymers or copolymers composed of poly(methacrylamide) and poly(vinylquinoline). The present invention also extends to any polymer or copolymer containing quinone groups.

[0020] Poly(methacrylamide) having quinoline groups represented by formula (1) can be obtained by oxidation of the catechol group (also known as benzene 1,2-diol group) of P(mDOPA) represented by formula (2). The oxidation is preferably carried out in an aqueous medium under basic conditions at a pH greater than 10, preferably between 10 and 12. [ka]

[0021] The poly(vinylquinoline) represented by formula (6) can be similarly prepared by the reaction of the poly(vinylquinoline) represented by formula (7) [ka] [wherein n is an integer greater than 1, preferably 1 to 100] It is obtained by oxidation of the catechol group (also known as the benzene 1,2 diol group) of

[0022] The oxidation is preferably carried out in an aqueous medium under basic conditions at a pH above 10, preferably between 10 and 12.

[0023] Amphiphilic molecules are molecules that have both hydrophilic and lipophilic properties. When dispersed in water, amphiphilic molecules have the property of self-assembling, for example, into micelles. They can be surfactants such as sodium dodecyl sulfate, 1-octanol, cocamidopropyl betaine, benzalkonium chloride, phospholipids, cholesterol, glycolipids, fatty acids, vitamin-polyether copolymers, such as vitamin E-polyoxyalkylene copolymers, particularly D-α-tocopheryl polyethylene glycol succinate (TPGS). Amphiphilic molecules can also be amphiphilic cyclodextrins obtained by grafting hydrocarbon chains onto the hydroxyl groups of cyclodextrins, such as hydroxypropyl-β-cyclodextrin or sulfobutylether-β-cyclodextrin. Amphiphilic cyclodextrins can incorporate hydrophobic biomolecules, therapeutic molecules, or drugs into their hydrophobic cavities.

[0024] For clarity, amphiphilic molecules have organic moieties as well as hydrophilic groups, and do not include molecules that are biologically active molecules, therapeutic molecules, or drugs as defined in the present invention. For example, vancomycin, minocycline, or ticagrelor are not considered amphiphilic molecules.

[0025] The polymer with primary or secondary amine groups can be polyallylamine, polyvinylamine, polyvinylamide, polyvinyl alcohol, poly(meth)acrylate, poly(meth)acrylamide, polyurethane, polyethylene glycol (PEG), polyelectrolytes (cationic, anionic or zwitterionic) with reactive groups that are primary or secondary amines.

[0026] The polymer with primary or secondary amine groups can be a natural or synthetic polymer with primary or secondary amine functional groups, such as polyvinylamine, chitosan or a protein.

[0027] In a preferred embodiment, the polymer having primary amine groups may comprise a polyallylamine, such as poly-(allylamine hydrochloride), also known as PAH, shown below, where p is an integer greater than 1, preferably p is 10 to 300: [ka]

[0028] The bioactive molecule, therapeutic molecule or drug can be an antibiotic, an anti-biofilm agent, an anti-platelet agent, an anti-coagulant, an anti-thrombotic agent and an anti-calcification agent.

[0029] Bioactive agents (also called bioactive molecules) can be molecules derived from plants, seeds, fungi, animals, humans, or microorganisms, or can be synthetically produced. They can include any substance that is desired to be delivered to a molecule, cell, tissue, or organ to regulate or otherwise alter molecular or cellular function, including therapeutic effects. Bioactive agents include, but are not limited to, pharmaceutically active compounds or diagnostic compounds.Bioactive molecules or compounds include nucleotides (aptamers, RNAi, antisense oligonucleotides), peptides, oligopeptides, proteins, apoproteins, glycoproteins, antigens and antibodies or antibody fragments thereof, receptors and other membrane proteins, retro-inverso oligopeptides, protein analogs in which at least one non-peptide bond replaces a peptide bond, enzymes, coenzymes, enzyme inhibitors, amino acids and their derivatives, hormones, lipids, phospholipids, liposomes, ricin or lysine. toxins, such as aflatoxin, digoxin, xanthotoxin, rubratoxin; analgesics, such as aspirin, ibuprofen and acetaminophen; bronchodilators, such as theophylline and albuterol; beta-blockers, such as propranolol, metoprolol, atenolol, labetolol, timolol, penbutolol and pindolol; antibacterial agents, such as the above and ciprofloxacin, cinoxacin and norfloxacin; antihypertensives, such as clonidine, methyldopa, prazolam, Zosyn, verapamil, nifedipine, aptopril and enalapril; cardiovascular drugs, such as antiarrhythmics, cardiac glycosides, antianginals and vasodilators; central nervous system drugs, such as stimulants, psychotropic drugs, antimanic drugs and antidepressants; antiviral drugs; antihistamines, such as chlorphenylmine and brompheniramine; anticancer drugs, such as chemotherapeutic drugs, e.g. chlorambucil, carboplatin, busulfan derivatives, doxorubicin, etoposide, topotecan (TPT); tranquilizers, e.g. diazepam, cordiazepine antidepressants, such as fluoxetine, amitriptyline, nortriptyline, and imipramine; H-2 antagonists, such as nizatidine, cimetidine, famotidine, and ranitidine; anticonvulsants; antinausea drugs; prostaglandins; muscle relaxants; anti-inflammatory substances; stimulants; decongestants; antiemetics; diuretics; antispasmodics; antiasthmatics; antiparkinsonian drugs; expectorants; antitussives; mucolytics; vitamins; and minerals and nutritional additives.Other molecules include nucleotides; oligonucleotides; polynucleotides; and art-recognized biologically functional analogs and derivatives, including, for example, methylated polynucleotides and nucleotide analogs with phosphorothioate linkages; plasmids, cosmids, artificial chromosomes, other nucleic acid vectors; those that are substantially complementary to at least one endogenous nucleic acid or that contain sequences of the opposite sense to at least a portion of a selected viral or retroviral genome; promoters; enhancers; inhibitors; and other ligands for regulating gene transcription and translation.

[0030] The bioactive agent can be an anti-infective, such as an antibiotic, e.g., amikacin, gentamicin, kanamycin, neomycin, netilmicin, tobramycin, paromomycin, streptomycin, spectinomycin, geldanamycin, herbimycin, rifaximin, loracarbef, ertapenem, dolupenem, imipenem / cilastatin, meropenem, cefadroxil, cefazolin, cephalothin, cephalexin, cefaclor, cefamandole, cefoxitin, cefoproxil, cefuroxime, or cefixime. , cefdinir, cedophytren, cefoperazone, cefotaxime, cefpodoxime, ceftazidime, ceftibuten, ceftizoxime, ceftriaxone, cefepime, ceftaroline fosamil, ceftobiprole, teicoplanin, vancomycin, telavancin, divervancin, oritavancin, clindamycin, lincomycin, daptomycin, azithromycin, clarithromycin, dirithromycin, erythromycin, roxithromycin, troleandomycin, telithromycin, Spiramycin, aztreonam, furazolidone, nitrofurantoin, linezolid, amoxicillin, ampicillin, piperacillin, ticarcillin, bacitracin, colistin, polymyxin B, ciprofloxacin, enoxacin, gatifloxacin, gemifloxacin, levofloxacin, lomefloxacin, moxifloxacin, nalidixic acid, norfloxacin, ofloxacin, mafenide, sulfacetamide, sulfazine, silver sulfazine, sulfadimethoxine, sulfamethoxazole These include, but are not limited to, benzodiazepine, benzocaine, benzodiazepine, benzophenone, benzocaine, benzodiazepine ...

[0031] Antibiotics can be either bacteriostatic, inhibiting growth without killing, or bactericidal, killing. The following antibiotics from the list above are generally considered bacteriostatic: minocycline (especially for S. aureus), tetracyclines, macrolides, clindamycin, linezolid, chloramphenicol, chlorhexidine, or alexidine (when used at low levels), or combinations thereof, while other antibiotics from the list above are almost always bactericidal.

[0032] Antibiotics can also be antivirulence agents when provided at low levels.

[0033] Virulence factors are molecules produced by pathogens that enable colonization, immune evasion, and damage to host cells. Antivirulence agents target pathogen virulence factors instead of killing or halting their growth, thereby neutralizing infectious pathogens. In contrast to bactericidal antibiotics, which create resistance, antivirulence agents do not create selective pressure for resistance. Antivirulence agents interfere with the interaction of pathogens, particularly bacteria, with host mammals, thereby reducing damage to the host and impairing the bacterial ability to cause disease. Antivirulence agents can inhibit the production of bacterial toxins or prevent their attachment to tissues.

[0034] Anti-biofilm agents include, but are not limited to, naturally occurring peptides such as human cathelicidin LL-37 or the bovine peptide indolicidin, or synthetic peptides such as 1018, natural compounds having a 2-aminoimidazole moiety, 2-aminoimidazole inhibitors, benzimidazole analogs, indole-triazo-amide analogs, plant-derived biofilm inhibitors such as emodin, phloretin, casbane diterpenes, resveratrol and its oligomers, sulfur derivatives, brominated furanone analogs, bromopyrrole alkaloids, skyramycin and (-)-ageloxime D structures, cembranoids, N-acylhomoserine lactone analogs, carolactone, molecules that prevent the formation of amyloid-like fibers, fatty acids, nitric oxide donors, ionic liquids such as 1-alkyl-3-methylimidazolium chlorides, 1-alkylquinolinium bromides, all of which may be used in combination with conventional antibiotics.

[0035] Antiplatelet agents include irreversible cyclooxygenase inhibitors, such as aspirin and triflusal (Disglend), adenosine diphosphate (ADP) receptor inhibitors, such as clopidogrel (Plavix), prasugrel (Effient), ticagrelor (Brilik and Brilinta), ticlopidine (Ticlid), phosphodiesterase inhibitors, such as cilostazol (Pletal), protease-activated receptor 1 (PAR-1) antagonists, such as vorapaxar (Zontivity), glycoprotein These include, but are not limited to, IIB / IIIA inhibitors (intravenous administration only), such as abciximab (ReoPro), eptifibatide (Integrilin), tirofiban (Aggrastat), adenosine reuptake inhibitors, such as dipyridamole (Persantin), thromboxane inhibitors, thromboxane synthase inhibitors and thromboxane receptor antagonists, such as terutroban, glycoprotein VI inhibitors, such as revecept, glycoprotein Ib inhibitors, and von Willebrand factor inhibitors.

[0036] Anticoagulants include, but are not limited to, acenocoumarol, coumatetralyl, dicoumarol, ethyl biscoumaceate, phenprocoumon, warfarin, chlorindione, dypgenadione, phenindione, cyclomarol, bemiparin, certoparin, ardeparin, dalteparin, enoxaparin, nadroparin, parnaparin, reviparin, dabigatran, apixaban, betrixaban, darexaban, edoxaban, otamixaban, rivaroxaban, alteplase, danaparoid, tinzaparin, and fondaparinux.

[0037] Thrombolytic agents include, but are not limited to, alteplase, reteplase, tenecteplase, saruplase, urokinase, anistreplase, monteplase, streptokinase, ancrod, brinase, and fibrinolysin.

[0038] Anti-calcification agents include, but are not limited to, bisphosphonates, aluminum salts, glutaraldehyde, aminooleic acid, and metalloproteinase inhibitors.

[0039] In a preferred embodiment, the nanogel according to the invention comprises an antibiotic and / or an antiplatelet drug.

[0040] In another preferred embodiment, the nanogel according to the invention comprises an antimicrobial agent, more preferably a combination of a bacteriostatic agent and an antivirulence agent, that inhibits bacterial growth and adhesion when the nanogel is coated on a surface.

[0041] Advantageously, nanogels containing such a combination of bacteriostatic and antivirulence agents slow bacterial growth as measured by metabolic rate and have increased anti-adhesion properties when the nanogels are coated onto medical devices, biomaterial implants or bioprostheses. Preferably, the molar ratio of bacteriostatic agent to antivirulence agent is 1:1 to 1:10, more preferably 1:2.

[0042] In a more preferred embodiment, the bacteriostatic agent is minocycline.

[0043] In another preferred embodiment, the bacteriostatic agent is chlorhexidine.

[0044] In a preferred embodiment, the biologically active molecule, therapeutic molecule or drug has the formula (3): [ka] [In the formula, R 1 C optionally substituted with one or more halogen atoms 3-5 alkyl; R 2 is a phenyl group optionally substituted with one or more halogen atoms; R 3 and R 4 are both hydroxyl; R is OH or XOH, and X is CH, OCH, CH, or a bond. or a pharmaceutically acceptable salt or solvate thereof, or a solvate of such a salt, provided that when X is CH or a bond, then R 1 is not propyl; X is CH2 and R 1 is CH2CH2CF3, butyl or pentyl, R 2 The phenyl group in must be substituted with fluorine; X is OCH2CH2 and R 1 When is propyl, R 2 The phenyl group must be substituted with fluorine.

[0045] The triazolo(4,4-d)-pyrimidine derivatives of formula (3) advantageously have antiplatelet activity, but also have antibacterial activity. They are particularly useful for reducing or preventing infection of medical devices, biomaterial implants, or bioprostheses that come into contact with blood when inserted or implanted into a mammalian host, and for preventing thrombosis. Thrombosis actually promotes infection of medical devices, biomaterial implants, or bioprostheses that come into contact with blood. The mammalian host can be a human patient or an animal.

[0046] In a most preferred embodiment, the triazolo(4,4-d)-pyrimidine derivative is (1S,2S,3R,5S)-3-[7-[(1R,2S)-2-(3,4-difluorophenyl)cyclopropylamino]-5-(propylthio)-3H-[1,2,3]-triazolo[4,5-d]pyrimidin-3-yl]-5-(2-hydroxyethoxy)-1,2-cyclopentanediol), also known as triafluosyl.

[0047] In another most preferred embodiment, the triazolo(4,4-d)-pyrimidine derivative is (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol, also known as fluometacyl, or Fluometacyl®, and has the formula (5): [ka] It is shown as follows.

[0048] In another preferred embodiment, the biologically active molecule, therapeutic molecule or drug is represented by formula (4): [ka] [In the formula, X 1 and X 2 are independently N, CH, CR 8 and R8 is C 1-6 Alkyl, C 2-6 Alkenyl or C 2-6 alkynyl; provided that X 1 or X 2 If one of the is equal to N, the remaining X 1 or X 2 , CH, CR 8 More selected; -Y- is -O- or -S-; R 11 and R 12 independently, C 1-6 -Alkyl, C 2-6 -Alkenyl, C 2-6 -alkynyl, C 3-6 -Cycloalkyl, aryl, aryl-C 1-6 -alkyl, wherein the alkyl or cycloalkyl moiety may be mono- or polysubstituted with OH or halogen, and the aryl moiety may be halogen, —C 1-6 Alkyl, -C 1-6 Alkoxy, -OH, -NO2, -CN, -NH2, -NHR 8 , -N(R 8 )2-COOH, -COOR 8 , -CONH2, -CONHR 8 , -CON(R 8 ) 2、 -SO2NH2, -SO2NHR 8 or -SO2N(R 8 ) may be mono- or polysubstituted by 2; R 13 , R 14 , R 15 , R 16 and R 17 are independently H, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, -OH, -NO2, -CN, -NH2, -NHR 8 , -N(R 8 )2-COOH, -COOR 8 , -CONH2, -CONHR 8 , -CON(R 8 ) 2、 -SO2NH2, -SO2NHR8 or -SO2N(R 8 )2] or an optical isomer thereof, a racemic mixture thereof, a pharmaceutically acceptable acid addition salt, a pharmaceutically acceptable metal salt or alkylated ammonium salt, or a prodrug thereof.

[0049] The pyrimidine derivatives of formula (4) advantageously have antibacterial effects. They are particularly useful for reducing or preventing infection of medical devices, biomaterial implants, or bioprostheses when inserted or implanted into a mammalian host. The mammalian host may be a human patient or an animal.

[0050] In another preferred embodiment, the biologically active molecule, therapeutic molecule or drug is a bisguanide, preferably chlorhexidine (hereinafter also referred to as chlorhexidine chloride).

[0051] As used herein, biguanide or HN(C(NH)NH) refers to a compound of formula (7): [ka] Bisguanides can be chlorhexidine, alexidine, and polyhexylbiguanide. As used herein, chlorhexidine refers to chlorhexidine base (8), also known as chlorhexidine chloride. [ka] However, it may also refer to chlorhexidine salts such as chlorhexidine diphosphanilate, chlorhexidine digluconate, chlorhexidine diacetate, chlorhexidine dinitrate, chlorhexidine dihydrochloride, chlorhexidine dichloride, chlorhexidine acetate, chlorhexidine dipropionate, chlorhexidine maleate, chlorhexidine succinate, chlorhexidine thiosulfate, chlorhexidine diacid phosphate, chlorhexidine malate, chlorhexidine dibenzoate, chlorhexidine diisophthalate, chlorhexidine dilaurate, chlorhexidine distearate, etc.

[0052] Alexidine, as used herein, refers to alexidine base, but can also refer to alexidine hydrochloride, alexidine dihydrochloride, alexidine monoacetate, alexidine diacetate, alexidine gluconate, alexidine digluconate, and mixtures thereof.

[0053] A bioactive agent, therapeutic molecule or drug is dispersed in a solvent along with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin, more preferably hydroxypropyl-β-cyclodextrin, and then added to the resulting dispersion of poly(methacrylamide) or poly(vinylquinoline) bearing quinoline groups.

[0054] The solvent can be any solvent that contains OH group.The solvent should not contain NH and / or -SH bond to avoid interaction with catechol group.The solvent can be, for example, water, alcohol, such as methanol, ethanol, butanol, propanol, etc., or a combination thereof.

[0055] The vitamin E derivative may be any copolymer obtained by esterifying vitamin-E (also called α-tocopherol succinate) with an acid ester, such as vitamin E acetate, or with a linear or branched polyether, such as a polyoxyalkylene, for example polyoxyethylene, polyoxypropylene, polyoxypropylene-polyoxyethylene copolymer, polyethylene glycol, polypropylene glycol, and the like.

[0056] The polyalkylene glycol has a molecular weight of 500-2000, preferably 750-1000, and most preferably 1000.

[0057] In a preferred embodiment, the vitamin E derivative is D-α-tocopheryl polyethylene glycol succinate (TPGS) represented by formula A. TPGS is a copolymer obtained by esterification of vitamin E (also called α-tocopherol succinate) with polyethylene glycol (PEG750 or PEG1000). [ka]

[0058] Amphiphilic molecules, particularly vitamin E derivatives, and more specifically TPGS copolymers, efficiently form micelles in solvents such as water or aqueous solutions containing 0-60% alcohol, e.g., ethanol. TPGS encapsulates hydrophobic bioactive agents, therapeutic molecules, or drugs, increasing their loading into the nanogel and also their efficacy and bioavailability through continuous release from within the nanogel over several weeks, preferably over 10 days.

[0059] TPGS copolymers efficiently form micelles in aqueous solutions with hydrophobic bioactive agent molecules, therapeutic molecules, or drugs when mixed at a TPGS:(bioactive agent, therapeutic molecule, or drug) ratio of 1:1 w / w to 5:1 w / w, preferably 2:1 w / w.

[0060] The hydrophilic portion of the TPGS copolymer, polyethylene glycol (PEG), forms the corona of the micelle, while the hydrophobic portion, tocopherol succinate, forms the core of the micelle. The hydrophobic core of the micelle can solubilize poorly soluble or insoluble drugs and partially protect bioactive agents, therapeutic molecules, or drugs from the aqueous environment. TPGS molecules encapsulate hydrophobic bioactive agents, therapeutic molecules, or drugs, contributing to their better stability when inserted into nanogels.

[0061] The micelles obtained by such encapsulation of bioactive agents, therapeutic molecules or drugs have an average particle size in the range of 10-100 nm, preferably 10 nm.

[0062] An amphiphilic molecule, preferably TPGS, combined with a bioactive agent, therapeutic molecule or drug, is encapsulated in the micelles and entrapped in the nanogel.

[0063] The nanogels comprise micelles of amphiphilic molecules, preferably TPGS, together with bioactive agents, therapeutic molecules, or drugs. The nanoparticles have a diameter of less than 1000 nm, e.g., about 100 nm to 300 nm. For example, nanogels can have a diameter of less than about 500 nm, less than about 300 nm, less than about 200 nm, or less than about 150 nm. In certain embodiments, nanogels of the present invention have a diameter of about 150 nm to about 250 nm. In certain embodiments, nanogels of the present invention have a diameter of about 100 to about 250 nm.

[0064] Nanogels may also contain amphiphilic molecules as amphiphilic cyclodextrin (Cy) structures or vesicles that incorporate hydrophobic bioactive agents, therapeutic molecules or drugs into the Cy hydrophobic cavities.

[0065] When micelles or vesicles are loaded into nanogels, amphiphilic molecules, preferably TGPS, more preferably hydroxypropyl-β-cyclodextrin, surprisingly reduce bacterial adhesion to the coated medical device, biomaterial implant or bioprosthesis and do not interfere with the pharmacological effect of the bioactive agent, therapeutic molecule or drug, as is possible in a solvent mixture.

[0066] Nanogels according to the invention can advantageously be loaded with higher levels of bioactive molecules, therapeutic molecules or drugs, and therefore also advantageously allow for longer release in contact with cells, tissues or organs when coated onto medical devices, biomaterial implants or bioprostheses.

[0067] The nanogels according to the invention can advantageously be loaded with both hydrophilic and hydrophobic bioactive agents, therapeutic molecules or drugs, preferably with a molar ratio of hydrophilic to hydrophobic bioactive agents, therapeutic molecules or drugs of 1:0 to 1:1, preferably 1:0.5.

[0068] The nanogels of the present invention maintain their structural integrity and prevent aggregation or degradation over time. Stable nanogels maintain their dispersion state, ensuring uniform distribution and optimal performance. The stable nanogel structure enhances the controlled release of drugs.

[0069] According to a second aspect, the present invention provides a method for producing a nanogel comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, said nanogel being obtained by one of both the following methods, depending on whether each bioactive molecule, therapeutic molecule or drug is loaded separately or simultaneously with the amphiphilic molecules into the nanogel: a) When each bioactive molecule, therapeutic molecule or drug is loaded separately with an amphiphilic molecule, the method comprises the following steps in order: i) Equation (1) [ka] [wherein x is an integer greater than 1, preferably 1 to 100] mixing a poly(methacrylamide) having quinoline groups represented by the formula: ii) adding a solution of a polymer having primary or secondary amine groups to the mixture obtained in step i) to obtain a crosslinked nanogel containing one bioactive molecule, therapeutic molecule or drug together with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin; iii) repeating steps i) and ii) for each additional bioactive molecule, therapeutic molecule, or drug; iv) mixing the crosslinked nanogels obtained in step iii) to obtain a nanogel containing one or more biomolecules, therapeutic molecules or drugs; b) When the bioactive molecule, therapeutic molecule or drug is loaded simultaneously with the amphiphilic molecule, the method comprises the following steps in order: i) Equation (1) [ka] [wherein x is an integer greater than 1, preferably 1 to 100] mixing a poly(methacrylamide) having quinoline groups represented by the formula: ii) adding a solution of a polymer having primary or secondary amine groups to the mixture obtained in step i) to obtain a crosslinked nanogel containing one or more bioactive molecules, therapeutic molecules or drugs together with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin.

[0070] In the first step (i), the formation of micelles or vesicles occurs immediately after adding a solution of poly(methacrylamide) having quinoline groups represented by formula (1) to a mixture of one or more biologically active molecules, therapeutic molecules, or drugs and amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins, in a solvent. The solvent can be water, alcohol, or a combination thereof. The solvent is preferably ethanol. The solution of poly(methacrylamide) having quinoline groups can be water, alcohol, or a combination thereof, but is preferably water.

[0071] The addition is carried out at room temperature under stirring.

[0072] In the second step (ii), a polymer bearing primary or secondary amine groups is reacted with a poly(methacrylamide) bearing quinoline groups, as shown in formula (1), via a quinoline-amine reaction, to obtain nanogels containing one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins, in a solvent.

[0073] Alternatively, the present invention provides a method for making a nanogel comprising a bioactive molecule, a therapeutic molecule or a drug together with an amphiphilic molecule, the nanogel being obtained by one of both of the following methods, depending on whether each bioactive molecule, therapeutic molecule or drug is loaded separately or simultaneously with the amphiphilic molecule into the nanogel: a) When each bioactive molecule, therapeutic molecule or drug is loaded separately with an amphiphilic molecule, the method comprises the following steps in order: i) mixing a poly(vinyl)quinoline group of formula (6) with only one biologically active molecule, therapeutic molecule or drug together with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin; ii) adding a solution of a polymer having primary or secondary amine groups to the mixture obtained in step i) to obtain a crosslinked nanogel containing one bioactive molecule, therapeutic molecule or drug together with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin; iii) repeating steps i) and ii) for each additional bioactive molecule, therapeutic molecule, or drug; iv) mixing the crosslinked nanogels obtained in step iii) to obtain a nanogel containing two or more biomolecules, therapeutic molecules or drugs; b) When a bioactive molecule, a therapeutic molecule or a drug is loaded with an amphiphilic molecule, the method comprises the following steps in order: i) Equation (6) [ka] [wherein n is an integer greater than 1, preferably 1 to 100] mixing a poly(vinyl)quinoline of the formula: ii) adding a solution of a polymer having primary or secondary amine groups to the mixture obtained in step i) to obtain a crosslinked nanogel containing one or more bioactive molecules, therapeutic molecules or drugs together with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin.

[0074] In the first step (i), micelle or vesicle formation and encapsulation with amphiphilic molecules occurs immediately after adding a solution of poly(vinyl)quinoline of formula (6) to a mixture of one or more bioactive molecules, therapeutic molecules or drugs and amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins, in a solvent.

[0075] The solvent can be water or an alcohol or a combination thereof, but is preferably ethanol.

[0076] The solution of poly(vinylquinoline) can be water or alcohol or a combination thereof, but is preferably water.

[0077] The addition is carried out at room temperature with stirring.

[0078] In the second step (ii), a polymer bearing primary or secondary amine groups is reacted with poly(vinylquinoline) of formula (6) via quinoline-amine interactions to obtain nanogels containing one or more bioactive molecules, therapeutic molecules or drugs encapsulated with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins.

[0079] In a preferred embodiment, the vitamin E derivative is D-α-tocophenyl polyethylene glycol succinate.

[0080] In another preferred embodiment, the amphiphilic cyclodextrin is hydroxypropyl-β-cyclodextrin.

[0081] In another preferred embodiment, the polymer having primary or secondary amine groups comprises polyallylamine, preferably poly-(allylamine hydrochloride), also known as PAH, shown below, where p is an integer greater than 1, preferably p is 10 to 300, and most preferably p is 160: [ka]

[0082] The solvent can be any solvent that contains an OH group and therefore at least one hydrogen that is prone to interact in a hydrogen bond, such as water, an alcohol, such as methanol, ethanol, butanol, propanol, etc., or a combination thereof.

[0083] In a preferred embodiment, the solvent is an alcohol, preferably ethanol.

[0084] In a preferred embodiment, the biologically active molecule, therapeutic molecule, or drug is a triazolo(4,4-d)-pyrimidine derivative of formula (3), and most preferably, the triazolo(4,4-d)-pyrimidine derivative is (1S,2S,3R,5S)-3-[7-[(1R,2S)-2-(3,4-difluorophenyl)cyclopropylamino]-5-(propylthio)-3H-[1,2,3]-triazolo[4,5-d]pyrimidine, also known as triafluosyl. The compound is (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol or Fluometacyl®, also known as (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol or Fluometacyl®.

[0085] In another preferred embodiment, the bioactive molecule, therapeutic molecule or drug is a bisguanide, most preferably chlorhexidine.

[0086] In another further preferred embodiment, the triazolo(4,4-d)-pyrimidine derivative is combined with a bacteriostatic antibiotic, preferably minocycline. The triazolo(4,4-d)-pyrimidine derivative is preferably triafluosyl or fluometacyl or Fluometacyl®.

[0087] In another further preferred embodiment, a triazolo(4,4-d)-pyrimidine derivative is combined with a bisguanide, most preferably chlorhexidine.

[0088] According to a third aspect, the present invention provides a biomaterial implant, medical device or bioprosthesis, the surface or part of which comprises a compound of formula (1) [ka] [wherein x is an integer greater than 1, preferably 1 to 100] It is composed of poly(methacrylamide) having a quinoline group represented by the formula: A nanogel, wherein the poly(methacrylamide) is crosslinked with a polymer having primary or secondary amine groups; the nanogel comprises one or more bioactive molecules, therapeutic molecules or drugs together with an amphiphilic molecule, preferably a vitamin E derivative, more preferably D-α-tocophenyl polyethylene glycol succinate; A nanogel-coated biomaterial implant, medical device or bioprosthesis is provided.

[0089] Alternatively, the present invention relates to a biomaterial implant, medical device or bioprosthesis, the surface or a portion thereof comprising a compound of formula (6): [ka] [wherein n is an integer greater than 1, preferably 1 to 100] It is composed of poly(vinyl)quinoline represented by the formula: A nanogel, wherein the poly(vinyl)quinoline is crosslinked with a polymer having primary or secondary amine groups; the nanogel comprises one or more bioactive molecules, therapeutic molecules or drugs together with an amphiphilic molecule, preferably a vitamin E derivative, more preferably D-α-tocophenyl polyethylene glycol succinate; A nanogel-coated biomaterial implant, medical device or bioprosthesis is provided.

[0090] Alternatively, the present invention provides a biomaterial implant, medical device or bioprosthesis, the surface of which, or a portion thereof, is coated with a nanogel composed of quinoline-containing poly(methacrylamide) and polyvinylquinoline or a copolymer thereof.

[0091] The present invention also extends to any biomaterial implant, medical device or bioprosthesis coated with a nanogel composed of a polymer or copolymer containing quinoline groups.

[0092] Biomaterial implants, medical devices or bioprostheses coated with nanogels according to the invention advantageously provide a more homogeneous, hydrophilic and smooth surface.

[0093] Such a more homogeneous, hydrophilic and smooth surface of a nanogel-coated biomaterial implant, medical device or bioprosthesis advantageously reduces injury and reduces thrombogenicity when inserted into a human patient or animal.

[0094] Also, biomaterial implants, medical devices or bioprostheses coated with nanogels comprising bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules advantageously provide more reproducible release kinetics of the hydrophobic bioactive molecules, therapeutic molecules or drugs from the coated nanogel and from the medical device, biomaterial implant or bioprosthesis, resulting in better pharmacological efficiency of the bioactive molecules, therapeutic molecules or drugs, and in particular preventing infection and thrombotic complications.

[0095] Furthermore, biomaterial implants, medical devices or bioprostheses coated with nanogels according to the present invention also advantageously provide a reduction in bacterial adhesion on their surface.

[0096] When micelles containing one or more bioactive molecules, therapeutic molecules, or drugs encapsulated with amphiphilic molecules, preferably vitamin E derivatives, more preferably D-α-tocophenyl polyethylene glycol succinate, are loaded into nanogels coated on the surface of biomaterial implants, medical devices, or bioprostheses, bacterial adhesion on the surface of the biomaterial implants, medical devices, or bioprostheses is reduced. This anti-adhesion effect is synergistically enhanced when a bacteriostatic agent is present in the nanogel.

[0097] Furthermore, the amphiphilic molecules in the nanogel coating, especially TPGS, do not interfere with the pharmacological effect of bioactive agents, therapeutic molecules or drugs, as can happen when used in solution.

[0098] The biomaterial implant can be any implantable foreign material for clinical use in a host mammal, such as for artificial joints, pacemakers, implantable cardioverter defibrillators, intravascular or urinary catheters or catheters containing materials, stents including coronary stents, mechanical and biological bioprosthetic heart valves, intraocular lenses, dental implants, etc.

[0099] A medical device can be, but is not limited to, any device, tool, instrument, implant, etc. related to the medical field or the practice of human or veterinary medicine, or intended for use in preventing or treating disease. Medical devices can include all natural and synthetic materials, as well as both fibrous and non-fibrous materials. For example, materials can be composed of metal, plastic, paper, glass, ceramic, textile, rubber, polymer, composite, or any other material or combination of materials. Exemplary medical devices include, but are not limited to, any type of catheter; cannula; needle; stent of any size, shape, or configuration; coil of any size, shape, or configuration; contact lens; intrauterine device (IUDS); peristaltic pump chamber; endotracheal tube; gastrointestinal feeding tube; arteriovenous shunt; condom; oxygenator and kidney membrane; gloves; pacemaker lead; wound dressing; metal pins, plates, and screws; metal hip prosthesis; artificial knee; and gel. In one embodiment, the nanogel of the present invention can be used to coat a catheter to prevent bacterial infection.

[0100] A bioprosthesis can be a prosthesis constructed from biological materials, including, but not limited to, heart valves, pericardium, vascular grafts, bladder prostheses, tendon prostheses, hernia patches, surgical meshes, and skin substitutes.

[0101] In one embodiment, the nanogels of the present invention can be used to coat bioprosthetic heart valves, such as decellularized porcine heart valves or bovine pericardium, to prevent bacterial infection and thrombosis.

[0102] The coated biomaterial implants, medical devices or bioprostheses may be used in a human or animal host for diagnosis, to prevent or treat disease, or for medical intervention.

[0103] In a preferred embodiment, the polymer having primary or secondary amine groups is poly-(allylamine hydrochloride) represented by formula (2), where p is an integer greater than 1, preferably 10-300.

[0104] In another preferred embodiment, the biologically active molecule, therapeutic molecule or drug is a triazolo(4,4-d)-pyrimidine derivative of formula (3); preferably (1S,2S,3R,5S)-3-[7-[(1R,2S)-2-(3,4-difluorophenyl)cyclopropylamino]-5-(propylthio)-3H-[1,2,3]-triazolo[4,5-d]pyrimidin-3-yl]-5-(2-hydroxyethoxy)-1,2-cyclo, also known as triafluosyl pentanediol) or ticagrelor; or (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol, also known as fluometacyl, or Fluometacyl®, which has both antibacterial and antiplatelet properties.

[0105] In another preferred embodiment, the bioactive molecule, therapeutic molecule or drug is a bisguanide, more preferably chlorhexidine.

[0106] In another preferred embodiment, the triazolo(4,4-d)-pyrimidine derivative is combined with a bacteriostatic antibiotic, preferably minocycline. Most preferably, the triazolo(4,4-d)-pyrimidine derivative is triafluocyl or fluometacyl or Fluometacyl®.

[0107] In another preferred embodiment, a triazolo(4,4-d)-pyrimidine derivative is combined with a bisguanide, more preferably chlorhexidine.

[0108] The nanogels according to the present invention can be immobilized or attached to the surface of a biomaterial implant, medical device, or bioprosthesis using various physical or chemical methods known in the art, such as electrografting, layer-by-layer deposition, spin-coating, spraying, or simply dipping the biomaterial implant, medical device, or bioprosthesis into a solution containing a quinoline-containing poly(methacrylamide) of formula (1) mixed with a mixture of a bioactive molecule, therapeutic molecule, or drug and an amphiphilic molecule, preferably a vitamin E derivative.

[0109] Alternatively, the surface of the biomaterial implant, medical device or bioprosthesis can be immersed in a solution of poly(vinylquinoline) mixed with one or more bioactive molecules, therapeutic molecules or drugs and amphiphilic molecules, preferably a vitamin E derivative.

[0110] Bioactive molecules, therapeutic molecules, or drugs are loaded into the nanogel in an encapsulated form with an amphiphilic molecule, preferably TPGS, and gradually and continuously released over time from the biomaterial implant, medical device, or bioprosthesis into a mammalian host, which may be a human patient or an animal, for several weeks, preferably two weeks.

[0111] When micelles are loaded into nanogels, amphiphilic molecules, especially TGPS, surprisingly reduce bacterial adhesion on the surface and do not interfere with the pharmacological effect of bioactive agents, therapeutic molecules or drugs.

[0112] Such anti-adhesion effect is synergistically enhanced when a bacteriostatic agent is present in the nanogel.

[0113] In a fourth aspect, the present invention provides a method for producing a medical device, biomaterial implant or bioprosthesis having a nanogel-coated surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins, said method comprising the following steps in order: ci) optionally immersing the surface to be coated in a dopamine buffer solution; cii) immersing the surface coated in step ci) in a solution of a polymer having primary or secondary amine groups; and ciii) immersing the coated surface obtained in step cii) in a liquid suspension of a crosslinked nanogel comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; said crosslinked nanogel being obtained in liquid suspension, preferably in water, by one of both methods of the invention; and civ) drying the crosslinked coated surface obtained in step ciii) to obtain a coated crosslinked nanogel surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; cv) optionally repeating steps cii) to civ) to obtain a surface covered with several layers of crosslinked nanogels containing one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins. The present invention provides a method comprising:

[0114] Alternatively, the present invention also provides a method for producing a medical device, biomaterial implant or bioprosthesis having a crosslinked monolayer or optionally multilayer coated surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins, said method comprising the following steps in order: di) optionally immersing the surface to be coated in a dopamine buffer solution; dii) immersing the surface coated in step di) in a solution of a polymer having primary or secondary amine groups; and diii) The coated surface obtained in step dii) is treated with a solution of the formula (1) [ka] [wherein x is an integer greater than 1, preferably 1 to 100] and one or more biologically active molecules, therapeutic molecules or drugs, together with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin; div) drying the cross-linked coated surface obtained in step diiii) to obtain a coated cross-linked monolayer surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; dv) optionally repeating steps dii) to div) to obtain a surface covered with crosslinked multilayers comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins. The present invention provides a method comprising:

[0115] In optional steps ci) and di), the medical device, biomaterial implant, or bioprosthesis is first immersed in a buffer solution, particularly a Tris buffer solution containing dopamine, to strongly anchor a first polymer layer to the surface of the medical device, biomaterial implant, or bioprosthesis. A primer coating of polydopamine PDA is then produced on the surface of the medical device by polymerization of dopamine molecules bearing a 4-(2-aminoethyl)benzene-1,2-diol motif.

[0116] In steps cii) and dii), the polymer having primary or secondary amine groups is preferably a PAH, and the covalent grafting of the PAH onto the primer coating occurs by amine / quinone reaction and / or Schiff base formation at room temperature.

[0117] In step diiii), the pre-coated surface obtained in dii) is immersed in a solution containing poly(methacrylamide) having quinoline groups of formula (1) mixed with one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins. The covalent grafting of the polymer layer is preferably achieved by the reaction of the primary or secondary amines of the PAH monolayer with the quinolone groups of formula (1) [ka] [wherein x is an integer greater than 1, preferably 1 to 100] This is carried out by the same reaction between the quinoline groups of poly(methacrylamide) and / or Schiff base formation.

[0118] In step ciii), the pre-coated surface obtained in step cii) is immersed in a liquid suspension of cross-linked nanogels comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; the liquid suspension is preferably water.

[0119] In steps cii) to cv), covalent grafting with a polymer, preferably a PAH, having primary or secondary amine groups occurs between the layer having quinone groups coated onto the medical device, biomaterial implant or bioprosthesis, resulting in a crosslinked nanogel coating on the medical device, biomaterial implant or bioprosthesis.

[0120] In steps dii) to dv), covalent grafting with a polymer, preferably a PAH, having primary or secondary amine groups occurs between the monolayers coated onto the medical device, biomaterial implant or bioprosthesis, resulting in a crosslinked multilayer coating on the medical device, biomaterial implant or bioprosthesis.

[0121] Similar to poly(vinylquinoline), the present invention also provides a method for producing a medical device, biomaterial implant or bioprosthesis having a nanogel-coated surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins, the method comprising the following steps in order: ei) optionally immersing the surface to be coated in a dopamine buffer solution; eii) immersing the surface coated in step ei) in a solution of a polymer having primary or secondary amine groups; and eiii) immersing the coated surface obtained in step eii) in a liquid suspension of a crosslinked nanogel comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins, said crosslinked nanogel being obtained by one of both methods of the invention in liquid suspension, preferably in water; eiv) drying the cross-linked coated surface obtained in step eiii) to obtain a coated cross-linked nanogel surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; ev) optionally repeating steps eii) to eiv) to obtain a surface covered with several layers of crosslinked nanogels comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins. The present invention provides a method comprising:

[0122] Alternatively, the present invention also provides a method for producing a medical device, biomaterial implant or bioprosthesis having an optionally multi-layer coated surface, the method comprising the steps of: fi) optionally immersing the surface to be coated in a dopamine buffer solution; fii) immersing the surface coated in step fi) in a solution of a polymer having primary or secondary amine groups; and fiii) The coated surface obtained in step fii) is treated with a solution of the formula (6) [ka] [wherein n is an integer greater than 1, preferably 1 to 100] and one or more biologically active molecules, therapeutic molecules or drugs together with an amphiphilic molecule, preferably a vitamin E derivative or an amphiphilic cyclodextrin; fiv) drying the cross-linked coated surface obtained in step fiii) to obtain a cross-linked monolayer coated surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; fv) optionally repeating steps fii) to fiv) to obtain a crosslinked multi-coated surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives. The present invention provides a method comprising:

[0123] In optional steps ei) and fi), the medical device, biomaterial implant, or bioprosthesis is first immersed in a buffer solution, particularly a Tris buffer solution containing dopamine (DOPA), to strongly fix a first layer of coating to the surface of the medical device, biomaterial implant, or bioprosthesis. A primer coating of PDA is then produced on the surface of the medical device by polymerization of dopamine bearing a 4-(2-aminoethyl)benzene-1,2-diol motif.

[0124] In steps eii) and fii), the polymer having primary or secondary amine groups is a PAH, and covalent grafting of the PAH onto the primer coating occurs by amine / quinone reaction and / or Schiff base formation at room temperature.

[0125] In step fiii), the pre-coated surface obtained in fiii) is immersed in a solution containing poly(vinylquinoline) of formula (6) mixed with one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins. Covalent grafting of the polymer layer was achieved by covalent reaction and / or Schiff base formation between the primary amines of the PAH monolayer and the quinone groups of poly(vinylquinoline) of formula (6).

[0126] In steps eii) to ev), covalent grafting with a polymer having primary or secondary amine groups, preferably a PAH, occurs between the nanogel layers coated onto the medical device, biomaterial implant or bioprosthesis, resulting in a crosslinked nanogel coating on the medical device, biomaterial implant or bioprosthesis.

[0127] In steps fii) to fv), covalent grafting with a polymer, preferably a PAH, having primary or secondary amine groups occurs between the monolayers coated onto the medical device, biomaterial implant or bioprosthesis, resulting in a crosslinked multilayer coating on the medical device, biomaterial implant or bioprosthesis.

[0128] Alternatively, the present invention also provides a method for producing a medical device, biomaterial implant, or bioprosthesis having a coated surface comprising a nanogel composed of poly(methacrylamide) having a quinoline group represented by formula (1) and polyvinylquinoline having formula (6), or a copolymer thereof.

[0129] The present invention also extends to a method for producing a biomaterial implant, medical device or bioprosthesis having a surface coated with a nanogel composed of a polymer or copolymer having quinoline groups.

[0130] The resulting nanogel may contain one or more, preferably two or more, bioactive molecules, therapeutic molecules and / or drugs. Bioactive molecules may include antibiotics and / or antiplatelet agents.

[0131] The method of the present invention can be used without the need for a primer coating step, in which case the coating adhesion is based on the adhesive properties of the free quinone groups present on the nanogel surface, which are sufficient to coat and immobilize the nanogels according to the invention on the surface of medical devices, biomaterial implants or bioprostheses.

[0132] A medical device, biomaterial implant, or bioprosthesis having a surface coated with two or more layers of crosslinked nanogel can be fabricated by repeating steps cii) and ciii) or eii) and eiii) of the above method. Medical devices, biomaterial implants, or bioprostheses containing two, three, four, five, or more layers of nanogel can be fabricated.

[0133] In a preferred embodiment, the biologically active molecule, therapeutic molecule or drug is a triazolo(4,4-d)-pyrimidine derivative of formula (3); preferably (1S,2S,3R,5S)-3-[7-[(1R,2S)-2-(3,4-difluorophenyl)cyclopropylamino]-5-(propylthio)-3H-[1,2,3]-triazolo[4,5-d]pyrimidin-3-yl]-5-(2-hydroxyethoxy)-1,2-cyclopentadiene, also known as triafluosyl. or (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol or Fluometacyl®, also known as fluometacyl, which has both antibacterial and antiplatelet properties.

[0134] In another preferred embodiment, the triazolo(4,4-d)-pyrimidine derivative is combined with a bacteriostatic antibiotic, preferably minocycline. The triazolo(4,4-d)-pyrimidine derivative is preferably triafluosyl or fluometacyl or Fluometacyl®.

[0135] In another preferred embodiment, the bioactive molecule, therapeutic molecule or drug is a bisguanide, most preferably chlorhexidine.

[0136] The method of the present invention can be used to coat only a portion of the surface of a medical device, biomaterial implant, or bioprosthesis, or substantially the entire or the entire surface of a medical device, biomaterial implant, or bioprosthesis.

[0137] The present invention also provides a coated medical device, biomaterial implant or bioprosthesis according to the present invention or produced by a method of the present invention for use in preventing or reducing infection when the medical device, biomaterial implant or bioprosthesis is implanted in a mammal, which may be a human patient or an animal.

[0138] In a fifth aspect, the present invention further provides a pharmaceutical composition comprising a nanogel according to the present invention or produced by a method of the present invention for use in preventing or reducing infection when applied by topical administration to a host mammal, which may be a human patient or an animal.

[0139] In a preferred embodiment, a pharmaceutical composition comprising a nanogel according to the invention is applied to animals, preferably dogs, sheep or cattle, for the treatment of skin diseases caused by bacterial infections, such as mastitis or pyoderma.

[0140] The pharmaceutical composition comprising the nanogel according to the invention is preferably a gel, but may also be a physiologically compatible liquid composition.

[0141] Pharmaceutical compositions may contain amphiphilic molecules, as well as biologically active molecules, therapeutic molecules, or drugs, auxiliary substances, preservatives, solvents, and / or viscosity-adjusting agents. Solvents include, for example, water, saline, or any other physiological solution, ethanol, glycerol, oils such as vegetable oils, or mixtures thereof. Viscosity-adjusting agents include, for example, carboxymethylcellulose.

[0142] In a preferred embodiment, the bioactive molecule, therapeutic molecule or drug is a bacteriostatic agent, preferably minocycline or chlorhexidine, in combination with an anti-infective or antiseptic or antivirulence agent.

[0143] In a most preferred embodiment, the biologically active molecule, therapeutic molecule or drug is triafluosyl.

[0144] In a sixth aspect, the present invention also provides the use of a nanogel according to the invention or produced by a method of the invention for inhibiting bacterial adhesion to the surface of a medical device, in particular the surface of a catheter.

[0145] A method of inhibiting bacterial adhesion on a surface may include the steps of: i) optionally immersing the surface to which bacterial adhesion is to be inhibited in a buffered solution of dopamine; ii) immersing the surface optionally coated in step i) in a solution of a polymer having primary or secondary amine groups; iii) immersing the surface obtained in step ii) in a solution of poly(methacrylamide) having quinoline groups of formula (1) mixed with one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; iv) drying the cross-linked coated surface obtained in step iii) to obtain a coated cross-linked monolayer surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins.

[0146] Alternatively, a method of inhibiting bacterial adhesion on a surface may comprise the steps of: i) immersing a surface to which bacterial adhesion is to be inhibited in a buffered solution of dopamine; ii) immersing the surface optionally coated in step i) in a solution of a polymer having primary or secondary amine groups; iii) immersing the surface obtained in step ii) in a solution of poly(vinylquinoline) of formula (6) mixed with one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; iv) drying the cross-linked coated surface obtained in step iii) to obtain a coated cross-linked monolayer surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins.

[0147] Alternatively, a method of inhibiting bacterial adhesion on a surface may comprise the steps of: i) optionally immersing the surface to which bacterial adhesion is to be inhibited in a buffered solution of dopamine; ii) immersing the surface coated in step i) in a solution of a polymer having primary or secondary amine groups; and iii) immersing the coated surface obtained in step ii) in a liquid suspension of a crosslinked nanogel comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; said crosslinked nanogel being obtained in liquid suspension, preferably in water, by one of both methods of the invention; and iv) drying the cross-linked coated surface obtained in step iii) to obtain a coated cross-linked nanogel surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins; v) Optionally, repeating steps cii) to civ) to obtain a surface covered with several layers of crosslinked nanogels comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives or amphiphilic cyclodextrins.

[0148] The invention will now be further described, by way of non-limiting examples only, with reference to the following figures and experimental examples. [Brief explanation of the drawings]

[0149] [Figure 1] FIG. 1 shows the dynamic size analysis (dynamic light scattering, DLS) of nanogel diameters in aqueous suspensions of NTTs containing triafluosil and TPGS according to the present invention compared to nanogel NTs containing only triafluosil according to WO2018 / 122318. [Figure 2] Figure 2 shows nanogel diameters in aqueous suspensions containing triafluosyl according to the invention alone (A) or in the presence of TPGS (B). Solid line: DLS analysis performed 1 hour after nanogel formation. Dashed line: DLS analysis performed 24 hours later. [Figure 3] FIG. 3 shows the surface topography of a polyurethane (PU) catheter (NTT) coated with nanogel according to the present invention compared with the surface topography of a PU catheter (NT) coated with nanogel according to WO2018 / 122318 by SEM analysis. [Figure 4] FIG. 4 shows the release kinetics of triafluosyl from nanogels according to the present invention (NTT) compared with the release kinetics of triafluosyl from nanogels according to WO2018 / 122318 (NT). [Figure 5] Figure 5 shows nanogel diameters in aqueous suspensions containing triafluosyl, TPGS, and minocycline. Solid line: DLS analysis performed 1 hour after nanogel formation. Dashed line: DLS analysis performed 24 hours later. [Figure 6] Figure 6 shows real-time microcalorimetry of S. aureus BAA-1556 metabolic activity, expressed as heat flow. The solid line represents control bacteria in vehicle (0.6% ethanol); the thick dashed line represents bacteria treated with 10 μg / mL triafluosil; the thick dotted line represents bacteria treated with 20 μg / mL triafluosil; the thin dashed line represents bacteria treated with both triafluosil and TPGS (10 μg / mL:10 μg / mL); and the thin dotted line represents bacteria treated with both triafluosil and TPGS (20 μg / mL:20 μg / mL). [Figure 7] Figure 7 shows real-time microcalorimetry of S. aureus BAA-1556 metabolic activity upon catheter attachment, expressed as heat flow. Solid line: control uncoated PU catheter; dashed line: PU catheter coated with triafluosyl-loaded nanogel; dotted line: PU catheter coated with triafluosyl-TPGS-loaded nanogel. [Figure 8]Figure 8 shows real-time microcalorimetry of bacterial metabolic activity during catheter attachment, expressed as heat flow. A: Baseline growth of S. aureus BAA-1556 (MRSA) at a dilution of 10-6 (no catheter section); B: Control, uncoated catheter section; C: NGM, a catheter section coated with five nanogel layers and grafted with PEG, loaded with 0.5 mg / mL of minocycline added to the last layer of nanogel (t=0 day); D: NTM, a catheter section coated with five nanogel layers and grafted with PEG, loaded with 0.05 mg / mL of ticagrelor, followed by 0.5 mg / mL of minocycline added to the last layer of nanogel (t=0 day); E: NTTM, a catheter section loaded with a combination of Triafluocyl® / TPGS (0.1 / 0.2 mg / mL, w / w), followed by 0.5 mg / mL of minocycline added to the last layer of nanogel (t=0 day). F, NGM, a catheter section coated with five layers of nanogel loaded with 0.5 mg / mL minocycline in the last layer of nanogel and grafted with PEG (t=20 days); G, NTM, a catheter section coated with five layers of nanogel loaded with 0.05 mg / mL Triafluocyl® followed by 0.5 mg / mL minocycline in the last layer of nanogel and grafted with PEG (t=20 days); H, NTTM, a catheter section coated with five layers of nanogel loaded with a combination of Triafluocyl® / TPGS (0.1 / 0.2 mg / mL, w / w) followed by 0.5 mg / mL minocycline in the last layer of nanogel (t=20 days). [Figure 9] Figure 9 shows the DLS analysis of nanogel diameter in aqueous suspension with chlorhexidine chloride (Chc) and TPGS as solid lines and without TPGS as dashed lines. [Figure 10] Figure 10 shows the DLS analysis of nanogel diameter in aqueous suspension containing triafluosyl, chlorhexidine (Chc) and TPGS as solid lines and without TPGS as dashed lines. [Figure 11]FIG. 11 shows photographs of nanogels containing Fluometacyl®, Chc and TPGS (NFCcT), as well as nanogels without TPGS (NFCc), showing the complete precipitation of NFCc after 48 hours. [Figure 12] Figure 12 shows real-time microcalorimetry of bacterial metabolic activity upon catheter attachment expressed as heat flow. Curve A is control P. aeruginosa 10-7 cell growth; curve B is nanogel-coated PU catheter (NFCc) loaded with Fluometacyl® and chlorhexidine chloride; curve C is nanogel-coated PU catheter (NFCcT) loaded with Fluometacyl® and chlorhexidine chloride in the presence of TPGS. [Figure 13] Figure 13 shows real-time microcalorimetry of P. aeruginosa metabolic activity upon catheter attachment, expressed as heat flow. Curve A: nanogel-coated PU catheter loaded with Fluometacyl® and chlorhexidine chloride (NFCc); Curve B: bacteria on nanogel-coated PU catheter loaded with Fluometacyl® and immersed in chlorhexidine chloride solution (1 mg / mL) (NF / Cc); Curve C: nanogel-coated PU catheter loaded with Fluometacyl® and chlorhexidine chloride in the presence of TPGS (NFCcT). [Figure 14] FIG. 14 shows DLS analysis of nanogel diameter in aqueous suspension for nanogels containing a mixture of fluometacil, chlorhexidine, and polymyxin B (NFCccP). [Figure 15] FIG. 15 shows DLS analysis of nanogel diameter in aqueous suspension for nanogels containing fluometacil and cyclodextrin (NFCy) before (solid line) and after (dashed line) addition of PAH. [Figure 16] FIG. 16 shows the stability study of nanogel diameter in aqueous suspension for NF, NFT, and NFCy containing various concentrations of fluometacil and stabilizer (TPGS or cyclodextrin) using DLS analysis. [Figure 17]Figure 17 shows real-time microcalorimetry of bacterial metabolic activity expressed as heat flow. Curve A is cell growth with control S. aureus MRSA 10-6; curve B is cells with Fluometacyl® (10 μg / mL); curve C is cells with Fluometacyl® (10 μg / mL) and hydroxypropyl-β-cyclodextrin (42 μg / mL); curve D is cells with Fluometacyl (20 μg / mL); curve E is cells with Fluometacyl® (20 μg / mL) and hydroxypropyl-β-cyclodextrin (42 μg / mL). [Figure 18] Figure 18 shows real-time microcalorimetry of bacterial metabolic activity upon catheter attachment expressed as heat flow. Curve A: control S. aureus MRSA 10-6 cell growth; Curve B: bacteria on nanogel-coated PU catheters loaded with Fluometacyl® and TPGS (0.2 / 0.4 mg / mL, w / w); Curve C: bacteria on nanogel-coated PU catheters loaded with Fluometacyl® and hydroxypropyl-β-cyclodextrin (0.2 / 15 mg / mL, w / w). [Example]

[0150] Below is a reference table summarizing the various nanogel compositions obtained according to the invention in the following examples. NT: Nanogel according to WO2018 / 122318A1 NTT: Nanogels containing triafluosyl and TPGS NG: Nanogels without bioactive molecules, therapeutic molecules or drugs NT: Nanogel containing triafluosyl NTTM: Nanogels containing triafluocyl, minocycline and TPGS NCcT: Nanogel containing chlorhexidine chloride and TPGS NFCcT: Nanogel containing Fluometacyl®, chlorhexidine chloride and TPGS NFCc: Fluometacyl®, a nanogel containing chlorhexidine chloride NFCy: Nanogels containing fluometacil and hydroxypropyl-β-cyclodextrin NF: Nanogel containing fluometasil NFT: Nanogels containing fluometasil and TPGS NFCccP: A nanogel containing fluometacil, chlorhexidine, and polymyxin B

[0151] Example 1: Preparation of nanogels according to the invention containing P(mDOPA) triafluosyl and TPGS (also referred to as NTT) The nanogel preparation is similar to that disclosed in WO2018 / 122318A1, except for drug loading in the presence of additional amphiphilic molecules.

[0152] The nanogels are prepared in a liquid solution, and after crosslinking, the nanogels remain suspended in the liquid solution.

[0153] 1.1 Oxidation of PmDOPA to Pox(mDOPA) A homopolymer of 3,4-dihydroxy-L-phenylalanine (P(mDOPA)) bearing methacrylamide was synthesized according to Faure & al in Adv Funct. Mater. 2012;22:5271-5282 and oxidized in aqueous medium under basic conditions for 12 h to obtain water-soluble Pox(mDOPA). The oxidized catechol moiety of Pox(mDOPA) is required for covalent interaction of PAHs via amine / quinoline reaction and / or Schiff base formation at room temperature, resulting in the production of stable crosslinked nanogels in aqueous suspension.

[0154] P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was added slowly to raise the pH to above 10 and promote oxidation of the catechol group of P(mDOPA).

[0155] 1.2 Preparation of Triafluosyl / TPGS Mixture in Ethanol Triafluosyl (also known as ticagrelor, provided by Polpharma) and TPGS (provided by MedChemExpress LLC) were dissolved separately in ethanol to prepare a 3.33 mg / mL stock solution. 160 μL of triafluosyl solution and 160 μL of TPGS solution were mixed under magnetic stirring (300 rpm), and then concentrated to approximately 160 μL under vacuum at room temperature.

[0156] 1.3 Preparation of nanogels containing triafluosyl / TPGS and PAHs Pox(mDOPA) (5 mL, 0.5 mg / mL) was added to the concentrated mixture obtained in 1.2 under magnetic stirring (300 rpm). After homogenization at room temperature for 1 hour, a pH 10 PAH aqueous solution (0.5 mL, 0.5 mg / mL) (Sigma-Aldrich) was slowly added to the mixture. The solution was reacted at room temperature for 1 hour under vigorous magnetic stirring (500 rpm).

[0157] Nanogels with diameters ranging from 100 nm to 350 nm were observed in aqueous suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0158] Example 2: Preparation of nanogels according to the invention containing polyvinyl(quinoline), triafluosyl, and TPGS 2.1 Oxidation of polyvinylcatechol (polyvinylquinoline) Similar to P(mDOPA), poly(vinylcatechol) provided by Polykey was oxidized in aqueous media under basic conditions for 12 hours to obtain water-soluble polyvinylquinoline. The oxidized catechol moiety of poly(vinylcatechol) is required for covalent interaction with PAHs via amine / quinoline reaction and / or Schiff base formation at room temperature, resulting in the production of stable crosslinked nanogels in aqueous suspension.

[0159] Poly(vinylcatechol) (10 mg) was dissolved in distilled water (20 mL), and NaOH (0.1 M) was added slowly to raise the pH above 10 for 12 h and promote the oxidation of the catechol groups of poly(vinylcatechol).

[0160] 2.2 Preparation of Triafluosyl / TPGS Mixture in Ethanol Triafluosyl (also known as tikagrelor) and TPGS were dissolved separately in ethanol to prepare a 3.33 mg / mL stock solution of triafluosyl and TPGS. 160 μL of triafluosyl solution and 160 μL of TPGS solution were mixed under magnetic stirring (300 rpm), and then concentrated to approximately 160 μL under vacuum at room temperature.

[0161] 2.3 Preparation of nanogels containing triafluosyl / TPGS and PAHs Poly(vinylquinoline) (5 mL, 0.5 mg / mL) was added to the concentrated mixture obtained in 2.2 under magnetic stirring (300 rpm). After homogenization for 1 hour, a pH 10 aqueous PAH solution (0.5 mL, 0.5 mg / mL) was slowly added to the mixture. The solution was reacted at room temperature for 1 hour under vigorous magnetic stirring (500 rpm).

[0162] Nanogels with diameters ranging from 100 nm to 350 nm were observed in aqueous suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0163] Example 3: Fabrication of a medical device with nanogels according to the present invention A polyurethane catheter (provided by Carfill) (5 Fr intravascular grade polyurethane tubing) was coated according to the method of the present invention.

[0164] Step 1: A catheter (4.5 cm long) was immersed in Aldrich's dopamine tris buffer solution pH=7.4 (0.2 g / L) for 5 hours.

[0165] Step 2: After rinsing twice with 5 mL of water, the modified catheter substrate was immersed in an aqueous PAH solution (pH>10) for 1 hour and rinsed twice with 5 mL of water.

[0166] Step 3: The modified catheter substrate obtained in step 2 was immersed in an aqueous solution of bioactive molecule-loaded nanogel prepared according to Example 1 or 2 for 18 hours and rinsed twice with 5 mL of water.

[0167] Steps 2 to 3 were repeated to build a multilayer assembly of nanogels on the surface of the coated device (five times to obtain a five-layer assembly of cross-linked nanogels).

[0168] Example 4: Comparison of nanogel NTT according to the present invention with nanogel NT of WO2018 / 122318A1 For comparison with the nanogels of the present invention, nanogels with and without triafluosyl are prepared according to WO2018 / 122318A1.

[0169] Preparation of triafluosyl-free nanogels (NG): P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was slowly added to raise the pH above 10 and promote oxidation of the catechol group of P(mDOPA). After overnight incubation at room temperature, a pH 10 aqueous solution of PHA (0.5 mL, 0.5 g / L) was slowly added to the Pox(mDOPA) solution. The solution was reacted for 1 h at room temperature under vigorous stirring (500 rpm, magnetic stirrer). Nanogels with diameters ranging from 100 nm to 250 nm were observed in the aqueous suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0170] Preparation of nanogels containing triafluosyl (NT): P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was slowly added to raise the pH above 10 and promote oxidation of the catechol group of P(mDOPA). After overnight incubation at room temperature, 0.16 mL of triafluosyl solution (3.33 mg / mL in DMSO) was added dropwise to the P(mDOPA) solution under stirring (300 rpm, magnetic stirrer) at room temperature. After 1 hour of homogenization, a pH 10 PAH aqueous solution (0.5 mL, 0.5 g / L) was slowly added to the mixture. The solution was reacted for 1 hour at room temperature under vigorous stirring (500 rpm, magnetic stirrer). Nanogel NTs with diameters ranging from 120 nm to 750 nm were observed in the aqueous suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0171] 4.1 Particle size and stability The diameter size of the nanogel particles according to the invention is compared with that obtained for the nanogels (NT) of WO2018 / 122318A1 with and without triafluosyl.

[0172] Compared with the diameters of nanogels without triafluosyl and TPGS (ranging from 100 nm to 250 nm) and nanogels containing only triafluosyl (ranging from 120 nm to 750 nm), the diameters of the nanogels of the present invention (ranging from 100 nm to 350 nm) are clearly intermediate between nanogel particles alone and nanogel particles containing triafluosyl. The nanogels of the present invention also exhibit a lower size distribution, as shown in Figure 1.

[0173] Figure 2 shows an increase in the size of the nanogel NTs after 24 h (dashed line), indicating the formation and precipitation of nanogel aggregates, which was not observed in the NTT nanogels, which remained stable after 24 h.

[0174] Therefore, the nanogels according to the present invention are more stable than those disclosed in WO2018 / 122318A1.

[0175] 4.2 Surface Topography As can be seen in Figure 3, SEM analysis of the coated catheters showed clear differences between the NT and NTT coatings. A more homogeneous and smooth surface was observed in the NTT coating, while large particle agglomerations present in the NT coating were observed, which increased the surface roughness.

[0176] 4.3 Surface hydrophilicity Contact angle measurements are a useful method for determining surface hydrophilicity. Table 1 shows contact angle analysis on glass coverslips (CS) (provided by WTR). CS was coated with polydopamine (CS-PDA). One-, three-, and five-layer NTTs (CS-NTT 1-layer, 3-layer, and 5-layer) were compared with a five-layer nanogel NT layer (CS-NT 5-layer PEG) according to WO2018 / 122318A1. Similar to WO2018 / 122318A1, PEG (MW 2000 g / mol) was covalently bonded onto the nanogel coating. Contact angles were measured after 30 seconds.

[0177] The static contact angle was clearly reduced in the presence of nanogel layers (NT or NTT) compared to PDA (primer polydopamine). The coated surface with five layers of NTT (nanogel with TPGS) showed a much lower water contact angle (18.3°) than the coated surface with the same number of NT layers (35.5°), indicating that the presence of PEG chains (1000 g / mol) in the TPGS molecules significantly improved the surface hydrophilicity even after the deposition of only one layer of NTT (25.3°). [Table 1]

[0178] 4.3 Release kinetics In vitro drug release from the coated catheter surface was quantified at predetermined time points up to 10 days. UV spectrophotometry was used to analyze the effect of the presence of TPGS on drug release kinetics, and drug release profiles were generated for each catheter from the data obtained in the experiments (Figure 4).

[0179] The presence of precipitates in the NT coating was found to significantly affect the drug release kinetics of the coated catheters. Drug release data showed that both coatings exhibited gradual and continuous release of the drug. However, significant variability was observed for the NT coating, indicating a lack of homogeneity. Furthermore, the release kinetics of triafluosyl with NT was much faster than with NTT, resulting in a shortened release duration, which negatively impacted the duration of the coating's pharmacological activity.

[0180] 4.4 Triafluosyl content in coated catheters Six polyurethane 4.5 cm long catheters (provided by Carfill) (5 Fr intravascular grade polyurethane tubing) were coated with NTT or NT according to the method of the present invention (NTT) or WO2018 / 122318A1 (NT).

[0181] The triafluosyl content in the coated catheter (4.5 cm) was determined by HPLC on a Waters Acquity UPLC System consisting of a quaternary solvent delivery system, an adjustable injection volume injector, a temperature-controlled autosampler, a column thermostat, and a photodiode array detector. The assay method was performed according to the ticagrelor monograph (European Pharmacopoeia 10.4). Briefly, the analytical column was an XBridge Phenyl, 150 x 4.6 mm, 3 μm (Waters) with a guard column, Security Guard Phenyl, 3 x 4 mm (Phenomenex). A 50 μL injection volume was used at a flow rate of 1.0 mL / min at 40 °C (column temperature). Mobile phase A: phosphate buffer pH 3.0-water-acetonitrile (1:89:10 v / v / v). Mobile phase B: phosphate buffer pH 3.0-water-acetonitrile (1:29:70 v / v / v). The detection wavelength was 300 nm.

[0182] The data presented in Table 2 show that a two-fold increase in the amount of triafluosil was obtained from the NTT-coated catheters compared to the NT-coated catheters. [Table 2]

[0183] Example 5: Preparation of nanogels according to the present invention (also referred to as NTTM) containing P(mDOPA) triafluosyl, minocycline and TPGS 5.1 Oxidation of PmDOPA → Pox(mDOPA). A homopolymer of 3,4-dihydroxy-L-phenylalanine (P(mDOPA)) with methacrylamide is oxidized in aqueous medium under basic conditions for 12 h to obtain water-soluble Pox(mDOPA). The oxidized catechol moiety of Pox(mDOPA) is required for covalent interaction of PAHs via amine / quinoline reaction and / or Schiff base formation at room temperature, and consequently, for the production of stable crosslinked nanogels in aqueous suspension.

[0184] P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was added slowly to Pox(mDOPA) to raise the pH to above 10 and promote the oxidation of the catechol group of P(mDOPA).

[0185] 5.2 Preparation of Triafluocyl / Minocycline / TPGS Mixture in Ethanol Triafluosyl (also known as ticagrelor), minocycline, and TPGS were dissolved separately in ethanol to prepare a 3.33 mg / mL stock solution. 160 μL of triafluosyl solution, 320 μL of TPGS solution, and 320 μL of minocycline solution were mixed under stirring (300 rpm), and then concentrated to approximately 160 μL under vacuum at room temperature.

[0186] 5.3 Preparation of nanogels containing triafluosyl / minocycline / TPGS and PAHs Pox(mDOPA) (5 mL, 0.5 mg / mL) was added to the concentrated mixture obtained in 5.2 under stirring. After homogenization at room temperature for 1 hour, an aqueous PAH solution (0.5 mL, 0.5 mg / mL) at pH 10 was slowly added to the mixture. The solution was reacted at room temperature under vigorous stirring (500 rpm) for 1 hour.

[0187] Stable nanogels with diameters ranging from 150 nm to 350 nm were observed in aqueous suspension by dynamic light scattering (Malvern) ( Figure 5 ).

[0188] The NTTM nanogels, like the NTT nanogels, were stable for up to 24 hours after formation. In contrast, the NT nanogels according to WO2018 / 122318A1 were not stable (Table 3). [Table 3]

[0189] On the other hand, when triafluosyl is released from nanogels prepared according to the method of the present invention and thus encapsulated by TPGS, surprisingly, a less inhibitory effect of TPGS is observed, as shown in the examples below.

[0190] Example 6 Effect of TPGS on the inhibitory activity of triafluosyl against bacterial growth S. aureus (ATCC BAA-1556, MRSA) was grown overnight in TSB (tryptic soy broth) medium and then cultured at 1 × 10 in fresh TSB. 4The bacterial suspensions were diluted 2-fold. Subsequently, 300 μL aliquots of the diluted bacterial suspensions were supplemented with 10 μg / mL and 20 μg / mL triafluosil (Polpharma), either alone or in a 1:1 w / w mixture with TPGS (MedChemExpress LLC, Europe). Both triafluosil and TPGS were prepared in absolute ethanol as described in the previous examples and refrigerated as master stocks in absolute ethanol. Triafluosil, TPGS, or ethanol as a vehicle (0.6% final concentration) was added to the bacterial suspensions to obtain concentrations (10 μg / mL and 20 μg / mL), followed by brief vortexing. Bacterial aliquots were then distributed into dedicated non-activated inserts in 48-well plates and grown for 24 hours under static conditions at 37°C using Calscreener technology. Cell growth and metabolic activity were measured in real time as described in https: / / cordis.europa.eu / project / id / 784514.

[0191] Real-time microcalorimetry of bacterial metabolic activity, expressed as heat flow, is shown for cell growth in TSB medium in Figure 6. The solid line represents control bacteria in vehicle (0.6% ethanol); the thick dashed line represents bacteria treated with 10 μg / mL triafluosil; the thick dotted line represents bacteria treated with 20 μg / mL triafluosil; the thin dashed line represents bacteria treated with both triafluosil and TPGS (10 μg / mL:10 μg / mL); and the thin dotted line represents bacteria treated with both triafluosil and TPGS (20 μg / mL:20 μg / mL).

[0192] Triafluosil, also known as ticagrelor, significantly reduced the growth of S. aureus at a concentration of 10 μg / mL, but at a higher dose of 20 μg / mL, bacterial growth was completely inhibited. Surprisingly, the addition of an equivalent amount (w / w) of TPGS to triafluosil significantly reduced the inhibitory effect of triafluosil on bacterial growth.

[0193] Example 7 Testing the bacterial adhesion prevention properties of nanogel coatings of triafluosyl alone and in combination with TPGS against S. aureus S. aureus (ATCC BAA-1556, MRSA) was grown overnight in TSB (tryptic soy broth) medium and then cultured at 1 × 10 in fresh TSB. 6 The bacterial suspension was diluted 2-fold. Subsequently, 1000 μL aliquots of the diluted bacterial suspension were added to two PU catheter segments (0.5 cm long) coated with nanogels loaded with either triafluosyl (0.5 mg / mL) or triafluosyl-TPGS (0.5 mg / mL w / w). Uncoated PU catheters served as controls. The nanogel coatings were prepared according to the previously described procedures in Examples 1 and 4. The bacterial solution was incubated with the catheters at 37°C and 220 rpm for 30 minutes. The catheters were then washed twice with saline (0.9% NaCl) and placed in dedicated non-activated inserts of a 48-well plate containing 300 mL of fresh TSB medium and grown for 24 hours under static conditions at 37°C using a Calscreener.

[0194] Figure 7 shows real-time microcalorimetry of bacterial metabolic activity during catheter attachment, expressed as heat flow. The solid line represents a control uncoated PU catheter; the dashed line represents a PU catheter coated with triafluosyl-loaded nanogels; and the dotted line represents a PU catheter coated with triafluosyl-TPGS-loaded nanogels.

[0195] Nanogel coating with triafluosyl can increase the anti-adhesive properties of PU catheters, which is seen as a delay in peak metabolic rate. The presence of TPGS in the coating further shifted the peak of bacterial growth, indicating less bacterial adhesion on the catheter surface.

[0196] Example 8. Combination of Triafluosyl-TPGS and the Antibacterial Agent Minocycline Confers Long-Term Anti-Adhesion Properties Against S. aureus to Nanogel (NTTM) Coatings S. aureus (ATCC BAA-1556, MRSA) was grown overnight in TSB (tryptic soy broth) medium and then cultured at 1 × 10 in fresh TSB. 6Subsequently, 1000 μL aliquots of the diluted bacterial suspension were applied to two catheter segments (0.5 cm long) coated with nanogels loaded with either triafluosyl (0.05 mg / mL, NT™) or triafluosyl-TPGS (0.1 / 0.2 mg / mL w / w, NT™) and minocycline as a bacteriostatic agent.

[0197] Preparation of nanogel solution containing Pox (mDOPA) and PAH (NG) A pH 10 aqueous solution of PAH (0.5 mL, 0.5 mg / mL) was slowly added to Pox(mDOPA) (5 mL, 0.5 mg / mL) under magnetic stirring (300 rpm). The solution was reacted at room temperature for 1 h under vigorous stirring (500 rpm).

[0198] Nanogels (NGs) with diameters ranging from 100 nm to 300 nm were observed in aqueous suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0199] Nanogel coating of catheters was performed according to the procedure previously described in Example 3. Minocycline (0.5 mg / mL) was added to the last layer of nanogel, either NG, NT, or NTT, resulting in the formation of the following variants: NGM, NTM, or NTTM, respectively. Uncoated catheter segments served as controls. To observe the longevity of the anti-adhesion activity of the tested coatings, two sets of coated catheter segments were tested: "washed" (long-term drug release assay) and "unwashed" (not incubated in buffer, time = 0 days) after 20 days of incubation in PBS / DMSO 2%.

[0200] To test the anti-adhesion properties of the nanogel coating, the bacterial solution was incubated with the catheters for 30 min at 37 °C and 220 rpm. Subsequently, the catheters were washed twice with saline (0.9% NaCl) and placed in dedicated non-activated inserts of 48-well plates containing 300 mL of fresh TSB medium and grown for 24 h under static conditions at 37 °C using a Calscreener. Microcalorimetry readouts were obtained.

[0201] Figure 8 shows real-time microcalorimetry of bacterial metabolic activity during catheter attachment, expressed as heat flow. A is 10 -6 Baseline growth of S. aureus (MRSA) at dilutions of 0.5 mg / mL (no catheter section); B, control uncoated catheter section; C, PEG-grafted catheter section coated with five layers of nanogel loaded with NGM and 0.5 mg / mL minocycline in the last layer of nanogel (t=0 day); D, PEG-grafted catheter section coated with five layers of nanogel loaded with NTM and 0.05 mg / mL triafluosil followed by 0.5 mg / mL minocycline in the last layer of nanogel (t=0 day); E, NTTM and a combination of triafluosil / TPGS (0.1 / 0.2 mg / mL, w / w) followed by 0.5 mg / mL minocycline in the last layer of nanogel. F: A catheter section coated with five layers of nanogel (t = 0 day); F: A catheter section coated with five layers of nanogel (NGM) loaded with 0.5 mg / mL minocycline in the last layer of nanogel and grafted with PEG (t = 20 days); G: A catheter section coated with five layers of nanogel (NTM) loaded with 0.05 mg / mL triafluosil followed by 0.5 mg / mL minocycline in the last layer of nanogel and grafted with PEG (t = 20 days); H: A catheter section coated with five layers of nanogel (NTTM) loaded with a combination of triafluosil / TPGS (0.1 / 0.2 mg / mL, w / w) followed by 0.5 mg / mL minocycline in the last layer of nanogel (t = 20 days).

[0202] The introduction of minocycline into the coating completely reduces bacterial adhesion to the catheters NGM, NTM and NTTM, as seen as background levels at the beginning of the drug release study (day 0, plots C, D, E).

[0203] The anti-adhesion effect of the coated catheters, especially NTM and NTTM (plots G and H), can be sustained for at least 20 days (the drug release period in this experimental setup). There is a clear synergistic effect for triafluosyl and minocycline (NTM, NTTM) compared to the non-triafluosyl coated NGM. This effect is enhanced in the presence of TPGS (NTTM), as seen by a shift and decrease in the microcalorimetry signal.

[0204] The use of a combination of triafluosyl and TPGS offers the possibility to increase the amount of triafluosyl loaded, resulting in a stronger synergy between triafluosyl and minocycline (NTTM vs. NTM). This clearly demonstrates the superiority of NTTM coating over NTM, as indicated by a more pronounced and sustained anti-adhesion effect in the long term (more than 20 days).

[0205] Example 9: Preparation of a nanogel according to the present invention containing P(mDOPA), chlorhexidine chloride, and TPGS (Nanogel NCcT) The preparation of the nanogels is similar to that described in Example 1, except for the drug loading, which uses chlorhexidine chloride as the active pharmaceutical ingredient and TPGS as the amphiphilic molecule.

[0206] 9.1 Oxidation of PmDOPA to Pox(mDOPA) A homopolymer of 3,4-dihydroxy-L-phenylalanine (P(mDOPA)) with methacrylamide is oxidized in aqueous medium under basic conditions for 12 h to obtain water-soluble Pox(mDOPA). The oxidized catechol moiety of Pox(mDOPA) is necessary for the covalent interaction of PAHs via amine / quinoline reaction and / or Schiff base formation at room temperature, and consequently, for the fabrication of stable crosslinked nanogels.

[0207] P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was added slowly to raise the pH to above 10 and promote oxidation of the catechol group of P(mDOPA).

[0208] 9.2 Preparation of Chlorhexidine Chloride / TPGS Mixture in DMSO Chlorhexidine chloride (Chc) and TPGS were dissolved separately in DMSO to prepare 10 mg / mL stock solutions of Chc and TPGS. 115 μL of Chc solution and 234 μL of TPGS solution were mixed under stirring (300 rpm) at room temperature for 10 minutes.

[0209] 9.3 Preparation of nanogels containing chlorhexidine chloride / TPGS and PAHs Pox(mDOPA) (5 mL, 0.5 mg / mL) was added to the mixture obtained in step 2 under stirring (300 rpm). After homogenization at room temperature for 1 hour, an aqueous PAH solution (0.5 mL, 0.5 mg / mL) at pH 10 was slowly added to the mixture. The solution was reacted at room temperature under vigorous stirring (500 rpm) for 1 hour.

[0210] Nanogels (NCcT) with diameters ranging from 100 nm to 350 nm were observed in liquid suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0211] Comparison of nanogel NCcT according to the present invention with nanogel NCc without TPGS: particle size and stability The diameter size of the nanogel particles according to Example 9 of the present invention is compared with that obtained without TPGS (NCc).

[0212] Preparation of nanogels containing chlorhexidine chloride (NCc): P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was slowly added to raise the pH above 10 and promote the oxidation of the catechol group of P(mDOPA). After overnight incubation at room temperature, 0.110 mL of Chc solution (10 mg / mL in DMSO) was added dropwise to the Pox(mDOPA) solution under stirring (300 rpm) at room temperature. After 1 hour of homogenization, a pH 10 aqueous PAH solution (0.5 mL, 0.5 g / L) was slowly added to the mixture. The solution was reacted under vigorous stirring (500 rpm) at room temperature for 1 h. Larger nanogels (NCs) with diameters exceeding 500 nm were observed in the mixed solution by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0213] Nanogels loaded with Chc in the presence of TPGS (NCcT) exhibit smaller diameters ranging from 100 nm to 350 nm and have a smaller size distribution compared to the diameter of nanogels without TPGS (NCc), as shown in Figure 9.

[0214] Example 10: Preparation of a nanogel according to the invention (nanogel NFCcT) containing P(mDOPA), Fluometacyl®, chlorhexidine chloride and TPGS The nanogel preparation is similar to that described in Example 1, except for drug loading, which uses Fluometacyl® (Fluo) and chlorhexidine chloride, and TPGS as the amphiphilic molecule.

[0215] 10.1 Oxidation of PmDOPA to Pox(mDOPA) A homopolymer of 3,4-dihydroxy-L-phenylalanine (P(mDOPA)) with methacrylamide is oxidized in aqueous medium under basic conditions for 12 h to obtain water-soluble Pox(mDOPA). The oxidized catechol moiety of Pox(mDOPA) is required for covalent interaction of PAHs via amine / quinoline reaction and / or Schiff base formation at room temperature, and consequently, for the production of stable crosslinked nanogels in aqueous medium.

[0216] P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was added slowly to raise the pH to above 10 and promote oxidation of the catechol group of P(mDOPA).

[0217] 10.2 Preparation of Fluometacyl®, Chlorhexidine Chloride / TPGS Mixture in DMSO Chlorhexidine chloride (Chc) and TPGS were dissolved separately in DMSO to prepare 10 mg / mL stock solutions of Fluo, Chc, and TPGS. 115 μL of Fluometacyl® solution, 115 μL of Chc solution, and 234 μL of TPGS solution were mixed under stirring (300 rpm) at room temperature for 10 minutes.

[0218] 10.3 Preparation of nanogels containing Fluo, Chc / TPGS, and PAHs Pox(mDOPA) (5 mL, 0.5 mg / mL) was added to the mixed solution obtained in 12.2 under stirring (300 rpm). After homogenization at room temperature for 1 hour, an aqueous solution of PAH (0.5 mL, 0.5 mg / mL) at pH 10 was slowly added to the mixture, and the solution was reacted at room temperature for 1 hour under vigorous stirring (500 rpm).

[0219] Nanogels (NFCcT) with diameters ranging from 100 nm to 350 nm were observed in the mixed solution by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0220] Comparison of the nanogel NFCcT according to the present invention with the nanogel NFCc without TPGS The diameter size of nanogels according to Example 10 of the present invention is compared with nanogels loaded with Fluo and Chc but not with TPGS (NFCc).

[0221] Preparation of chlorhexidine chloride-free nanogel (NFCc): P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was slowly added to raise the pH above 10 and promote oxidation of the catechol group of P(mDOPA). After overnight incubation at room temperature, 0.11 mL of Chc solution (10 mg / mL in DMSO) was added, followed by 0.11 mL of Fluo solution (10 mg / mL in DMSO) to the Pox(mDOPA) solution under stirring (300 rpm) at room temperature. After 1 hour of homogenization, a pH 10 aqueous PAH solution (0.5 mL, 0.5 g / L) was slowly added to the mixture. The solution was allowed to react for 1 hour at room temperature under vigorous stirring (500 rpm). Large nanogels (NFCc) with diameters exceeding 500 nm were observed in the mixture by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0222] Nanogels loaded with Fluometacyl® and Chc in the presence of TPGS (NFCcT) exhibited smaller particle diameters ranging from 100 nm to 350 nm and had a narrower size distribution compared to the nanogels without TPGS (NFCc) diameter, as shown in Figure 10.

[0223] After 24 h, an increase in the size of NFCc was observed, indicating the formation and precipitation of nanogel aggregates, which was not observed for NFCcT, which remained stable after 24 h (Figure 11).

[0224] Example 11: The anti-adhesion properties of nanogel-coated catheters loaded with Fluometacyl® and chlorhexidine in the presence or absence of TPGS were tested against Pseudomonas aeruginosa. 11.1 Preparation of test samples for microcalorimetry P. aeruginosa (ATCC 15442) was grown overnight in TSB (tryptic soy broth) medium and then cultured at 1 × 10 in fresh TSB. 7The diluted bacterial suspension was diluted 2:1.5 times. Subsequently, a 1000 mL aliquot of the diluted bacterial suspension was added to a 2PU catheter segment (0.5 cm long) coated with five layers of nanogel loaded with Fluometacyl® (0.2 mg / mL) and chlorhexidine chloride (0.2 mg / mL) either with or without TPGS (0.4 mg / mL). The nanogel coatings were prepared and loaded with the respective bioactive molecules according to the following procedure.

[0225] The bacterial solution was incubated with the catheters for 30 minutes at 37°C and 220 rpm. The catheters were then washed twice with saline (0.9% NaCl) and placed in dedicated, non-activated inserts of 48-well plates containing 300 mL of fresh TSB medium and grown for 24 hours under static conditions at 37°C using a Calscreener.

[0226] 11.2 Preparation of Nanogels (NFCc and NFCcT) Containing Pox (mDOPA) and PAH and Drug-Loaded (Fluometacyl® and Chlorhexidine) A pH 10 PAH aqueous solution (0.5 mL, 0.5 mg / mL) was slowly added to Pox(mDOPA) (5 mL, 0.5 mg / mL) pre-incubated with Fluometacyl® (0.2 mg / mL) and chlorhexidine chloride (CHLXc, 0.2 mg / mL) with or without TPGS (0.4 mg / mL) for 10 min under magnetic stirring (300 rpm). The solution was reacted at room temperature for 1 h under vigorous stirring (500 rpm).

[0227] Nanogels with diameters ranging from 100 nm to 300 nm were observed in aqueous suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0228] Test drugs were stored frozen as the following stock solutions: Fluometacyl® (3.3 mg / mL stock in DMSO, prepared according to Eur J Med Chem 2020 Dec 15:208:112767), chlorhexidine chloride (3.3 mg / mL stock in DMSO, Merck), TPGS (3.3 mg / mL stock in DMSO, Merck).

[0229] Figure 12 shows real-time microcalorimetry of bacterial metabolic activity upon catheter attachment expressed as heat flow. Curve A is the control P. aeruginosa 10 -7 Curve B is nanogel-coated PU catheter (NFCc) loaded with Fluometacyl® and chlorhexidine chloride; curve C is nanogel-coated PU catheter (NFCcT) loaded with Fluometacyl® and chlorhexidine chloride in the presence of TPGS.

[0230] Nanogel coatings containing fluometacil® and chlorhexidine chloride in the presence of TPGS confer enhanced anti-adhesion properties to PU catheters compared to coatings lacking TPGS. This is seen as a shift in the time from the population adhering to the catheter during initial incubation to the peak of bacterial metabolic activity. In other words, fewer bacteria adhere to the catheter, and metabolic activity appears later.

[0231] TPGS, as an amphiphilic molecule, enhances the entrapment of hydrophobic and hydrophilic antimicrobial molecules, such as Fluometacyl® and chlorhexidine, into the nanogel coating.

[0232] Example 12: Nanogel-coated catheters loaded with Fluometacyl® and chlorhexidine exhibit stronger anti-adhesion properties against Pseudomonas aeruginosa in the presence of TPGS 12.1 Preparation of test samples for microcalorimetry P. aeruginosa (ATCC 15442) was grown overnight in TSB (tryptic soy broth) medium and then cultured at 1 × 10 in fresh TSB. 7 The diluted bacterial suspension was diluted 2x. Subsequently, a 1000 mL aliquot of the diluted bacterial suspension was added to a 2PU catheter segment (0.5 cm long) coated with five nanogel layers loaded with Fluometacyl® (0.2 mg / mL) and immersed in aqueous chlorhexidine chloride (1 mg / mL) for 1 hour, or to a 2PU catheter segment (0.5 cm long) coated with five nanogel layers loaded with Fluometacyl® (0.2 mg / mL) and chlorhexidine chloride (0.2 mg / mL) in the presence or absence of TPGS (0.4 mg / mL). The nanogel coatings were prepared according to the following procedure, and the respective bioactive molecules were added to them.

[0233] The bacterial solution was incubated with the catheters for 30 minutes at 37°C and 220 rpm. The catheters were then washed twice with saline (0.9% NaCl) and placed in dedicated, non-activated inserts of 48-well plates containing 300 mL of fresh TSB medium and grown for 24 hours under static conditions at 37°C using a Calscreener.

[0234] 12.2 Preparation of drug-loaded nanogels (NFCcT) containing Pox (mDOPA) and PAH A pH 10 PAH aqueous solution (0.5 mL, 0.5 mg / mL) was slowly added to Pox(mDOPA) (5 mL, 0.5 mg / mL) pre-incubated with Fluometacyl® (0.2 mg / mL) and chlorhexidine chloride (Cc, 0.2 mg / mL) for 10 min with or without TPGS (0.4 mg / mL) under magnetic stirring (300 rpm). The solution was reacted for 1 h at room temperature under vigorous stirring (500 rpm). Alternatively, instead of loading the nanogels with both Fluometacyl® and chlorhexidine chloride, nanogel-coated catheters containing Fluometacyl® were immersed in an aqueous Chc solution (1 mg / mL) under stirring (orbital shaker) for 1 h to load chlorhexidine separately from Fluometacyl®.

[0235] Nanogels with diameters ranging from 100 nm to 300 nm were observed in aqueous suspension by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0236] Test drugs were stored frozen as the following stock solutions: Fluometacyl® (3.3 mg / mL stock in DMSO, prepared according to Eur J Med Chem 2020 Dec 15:208:112767), chlorhexidine chloride (3.3 mg / mL stock in DMSO, Merck), and TPGS (3.3 mg / mL stock in DMSO, Merck). For the soaking procedure, fresh aqueous solutions of 1 mg / mL were prepared from the powder.

[0237] Figure 13 shows real-time microcalorimetry of bacterial metabolic activity upon catheter attachment, expressed as heat flow. Curve A: nanogel-coated PU catheter loaded with Fluometacyl® and chlorhexidine chloride (NFCc); Curve B: bacteria on nanogel-coated PU catheter loaded with Fluometacyl® and immersed in chlorhexidine chloride solution (1 mg / mL) (NF / Cc); Curve C: nanogel-coated PU catheter loaded with Fluometacyl® and chlorhexidine chloride in the presence of TPGS (NFCcT).

[0238] In the presence of TPGS, nanogel coatings loaded with Fluometacyl® and chlorhexidine conferred the strongest anti-adhesion properties, as seen by a forward-shifted peak metabolic signal. Immersion in chlorhexidine conferred intermediate properties compared to the approach of simultaneously loading Fluometacyl® and chlorhexidine. This supports the positive effect of TPGS for loading bioactive molecules into nanogel coatings.

[0239] Example 13 Stabilization of Fluometacyl®, Chlorhexidine, Polymyxin B Mixtures with and without TPGS The preparation of nanogels is similar to that described above for NF and NFT, except that a mixture of Fluometacyl®, chlorhexidine and polymyxin B is used, maintaining the same concentration (0.2 mg / mL).

[0240] Stabilization of different active pharmaceutical ingredients or drugs can be problematic due to interactions that may exist between molecules. This example clearly demonstrates the added value of the presence of TPGS to enhance nanogel stability in liquid suspension. NFCcP nanogels containing three different drugs or APIs without TPGS produce large aggregates with extremely low stability. However, in the presence of TPGS, NFCcPT nanogels exhibit better stability, and DLS analysis shows that the nanogel diameter is approximately micron-sized in liquid suspension. In Figure 14, a mixture of nanogels NFT, NFCcT, and NPT exhibits nanogel diameters ranging from 100 nm to 350 nm, which has higher stability in liquid suspension.

[0241] Example 14 Preparation of a nanogel (NFCy) according to the invention containing P(mDOPA) Fluometacyl® and hydroxypropyl-β-cyclodextrin The preparation of nanogels is similar to that described above, except that hydroxypropyl-β-cyclodextrin (hereinafter also referred to as cyclodextrin) is used instead of TPGS.

[0242] 14.1 Oxidation of PmDOPA to Pox(mDOPA) A homopolymer of 3,4-dihydroxy-L-phenylalanine (P(mDOPA)) with methacrylamide is oxidized in aqueous medium under basic conditions for 12 h to obtain water-soluble Pox(mDOPA). The oxidized catechol moiety of Pox(mDOPA) is required for covalent interaction of PAHs via amine / quinoline reaction and / or Schiff base formation at room temperature, and consequently, for the fabrication of stable crosslinked nanogels in liquid dispersions.

[0243] P(mDOPA) (2.5 mg) was dissolved in distilled water (5 mL), and NaOH (0.1 M) was added slowly to raise the pH to above 10 and promote oxidation of the catechol group of P(mDOPA).

[0244] 14.2 Preparation of Fluometacyl® / Hydroxypropyl-β-Cyclodextrin Mixture in DMSO 75 μL of hydroxypropyl-β-cyclodextrin solution (provided by Merck) 40% (400 mg / mL in H2O) and 60 μL of fluometacil solution (3.3 mg / mL in DMSO) were mixed under magnetic stirring (300 rpm).

[0245] 14.3 Preparation of Nanogels Containing Fluometacyl® / Hydroxypropyl-β-Cyclodextrin and PAHs Pox(mDOPA) (0.87 mL, 0.5 mg / mL) was added to the mixture obtained in 14.2 under magnetic stirring (300 rpm). DLS analysis (Figure 15) shows the presence of species smaller than 10 nm, which characterizes cyclodextrin molecules. After 1 h of homogenization at room temperature, a pH 10 aqueous PAH solution (0.087 mL, 0.5 mg / mL) was slowly added to the mixture. The solution was allowed to react for 1 h at room temperature under vigorous magnetic stirring (500 rpm). Figure 15 shows the nanogels NFCy (dashed line) with diameters ranging from 100 nm to 350 nm in the liquid suspension, as observed by dynamic light scattering (Zetasizer Advance Pro, Malvern) compared to the solution before the addition of PAH (in the absence of nanogels) (solid line).

[0246] Example 15: Comparison of the stability of nanogels NF, NFT and NFCy according to the invention containing various concentrations of Fluometacyl® and TPGS or hydroxypropyl-β-cyclodextrin The preparation of nanogels was similar to that described above for NF, NFT, and NFCy, except that various concentrations of Fluometacyl® (0.2, 0.4, and 0.8 mg / mL) were used, maintaining the same ratios of amphiphilic molecules (Fluo:TPGS 1:2 and Fluo:cyclodextrin 1:75).

[0247] The size and polydispersity of the different nanogels are monitored by DLS on days 0, 1, and 4. It can be clearly seen that increasing the concentration of fluometacil increases the size and polydispersity of the nanogel NFs and induces the formation of precipitates. However, in the presence of the amphiphilic molecules TPGS and hydroxypropyl-β-cyclodextrin, which are considered as stabilizers, the nanogels NFT and NFCy show a higher degree of stability.

[0248] Example 16 Hydroxypropyl-β-cyclodextrin, in contrast to TPGS, does not affect the antibacterial efficacy of Fluometacyl® in liquid medium tested against Staphylococcus aureus 16.1 Preparation of test samples for microcalorimetry S. aureus (MRSA, ATCC 6538) was grown overnight in TSB (tryptic soy broth) medium and then cultured at 1 × 10 in fresh TSB. 7 The diluted bacterial suspension was diluted 1:1.5 times. Subsequently, 1000 mL aliquots of the diluted bacterial suspension were supplemented with Fluometacyl® (10 or 20 μg / mL) or a mixture of Fluometacyl® (10 or 20 μg / mL) and hydroxypropyl-β-cyclodextrin (42 μg / mL). Subsequently, 300 μL of the suspension was placed in a dedicated non-activated insert of a 48-well plate and grown for 24 hours under static conditions at 37°C using a Calscreener.

[0249] 16.2 Preparation of Stock Solutions Fluometacyl® (prepared according to Eur J Med Chem 2020 Dec 15:208:112767) was kept frozen as the following stock solution of 3.3 mg / mL in EtOH: Hydroxypropyl-β-cyclodextrin (Merck) was kept at room temperature as a powder, and the appropriate stock solution in HO was prepared fresh before use.

[0250] Figure 17 shows real-time microcalorimetry of bacterial metabolic activity expressed as heat flow. Curve A is the control S. aureus MRSA10-6 Curve A is cells with Fluometacyl® (10 μg / mL); Curve C is cells with Fluometacyl® (10 μg / mL) and hydroxypropyl-β-cyclodextrin (42 μg / mL); Curve D is cells with Fluometacyl® (20 μg / mL); Curve E is cells with Fluometacyl® (20 μg / mL) and hydroxypropyl-β-cyclodextrin (42 μg / mL). In solution, hydroxypropyl-β-cyclodextrin did not impair the antibacterial activity of Fluometacyl® compared to TPGS (previous example).

[0251] Example 17: Anti-adhesion properties of Fluometacyl®-loaded nanogel-coated catheters in the presence of TPGS or hydroxypropyl-β-cyclodextrin tested against Staphylococcus aureus 17.1 Preparation of test samples for microcalorimetry S. aureus (MRSA, ATCC 6538) was grown overnight in TSB (tryptic soy broth) medium and then cultured at 1 × 10 in fresh TSB. 7 The diluted bacterial suspension was diluted 2:1.5 times. Subsequently, a 1000 mL aliquot of the diluted bacterial suspension was added to a 2PU catheter segment (0.5 cm long) coated with five layers of nanogels loaded with Fluometacyl® (0.2 mg / mL) and TPGS (0.4 mg / mL) or Fluometacyl® (0.2 mg / mL) and hydroxypropyl-β-cyclodextrin (15 mg / mL). The nanogel coatings were prepared and loaded with the respective bioactive molecules according to the following procedure.

[0252] The bacterial suspension was incubated with the catheters for 30 minutes at 37°C and 220 rpm. The catheters were then washed twice with saline (0.9% NaCl) and placed in dedicated non-activated inserts of 48-well plates containing 300 mL of fresh TSB medium and grown for 24 hours under static conditions at 37°C using a Calscreener.

[0253] 17.2 Preparation of drug-loaded nanogels containing Pox (mDOPA) and PAH (NG) A pH 10 aqueous PAH solution (0.5 mL, 0.5 mg / mL) was slowly added under magnetic stirring (300 rpm) to 5 mL of Pox(mDOPA) (0.5 mg / mL) pre-incubated for 10 min with either Fluometacyl® (0.2 mg / mL) and TPGS (0.4 mg / mL) or Fluometacyl® (0.2 mg / mL) and hydroxypropyl-β-cyclodextrin (15 mg / mL). The solution was reacted for 1 h at room temperature under vigorous stirring (500 rpm).

[0254] Nanogels with diameters ranging from 100 nm to 300 nm were observed by dynamic light scattering (Zetasizer Advance Pro, Malvern).

[0255] Test drugs were stored frozen as the following stock solutions: Fluometacyl® (3.3 mg / mL stock in DMSO, prepared according to Eur J Med Chem 2020 Dec 15:208:112767), TPGS (3.3 mg / mL stock in DMSO, Merck). Hydroxypropyl-β-cyclodextrin (Merck) was kept at room temperature as a powder, and appropriate stock solutions in HO were prepared fresh before use.

[0256] Figure 18 shows real-time microcalorimetry of bacterial metabolic activity upon catheter attachment expressed as heat flow. Curve A is the control S. aureus MRSA 10 -6 Curve B: bacteria on nanogel-coated PU catheter loaded with Fluometacyl® and TPGS (0.2 / 0.4, w / w); Curve C: bacteria on nanogel-coated PU catheter loaded with Fluometacyl® and hydroxypropyl-β-cyclodextrin (0.2 / 15, w / w).

[0257] The nanogel coating containing Fluometacyl® and hydroxypropyl-β-cyclodextrin confers enhanced anti-adhesion properties to PU catheters compared to the coating containing TPGS. This is seen as a decrease in signal and a shift in the time to peak metabolic activity of the bacteria grown from the population attached to the catheter during the initial incubation. In other words, fewer bacteria adhere to the catheter section, and metabolic activity appears later.

[0258] Hydroxypropyl-β-cyclodextrin enhances entrapment of Fluometacyl® into the nanogel coating compared to TPGS.

Claims

1. Formula (1) 【Chemistry 1】 wherein x is an integer greater than 1, preferably 1 to 100. and / or Formula (6) 【Chemistry 2】 wherein n is an integer greater than 1, preferably n is 1 to 100. It is composed of poly(vinylquinoline) represented by the formula: A nanogel crosslinked with a polymer having primary or secondary amine groups; A nanogel, wherein the nanogel comprises one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules.

2. 2. The nanogel according to claim 1, wherein the amphiphilic molecule is a vitamin E derivative, preferably D-α-tocophenyl polyethylene glycol succinate.

3. 2. The nanogel according to claim 1, wherein the amphiphilic molecule is an amphiphilic cyclodextrin, preferably hydroxypropyl-β-cyclodextrin.

4. The polymer having primary or secondary amine groups is represented by the formula (2): 【Transformation 3】 wherein p is an integer greater than 1, and preferably p is 10 to 300. The nanogel according to any one of claims 1 to 3, which is poly-(allylamine hydrochloride) represented by the formula:

5. The biologically active molecule, therapeutic molecule or drug is represented by the formula (3): 【Chemistry 4】 [In the formula, R 1 C optionally substituted with one or more halogen atoms 3-5 alkyl; R 2 is a phenyl group optionally substituted with one or more halogen atoms; R 3 and R 4 are both hydroxyl; R is OH or XOH, and X is CH 2 , OCH 2 CH 2 or a bond] or a pharmaceutically acceptable salt or solvate thereof, or a solvate of such a salt, provided that X is CH 2 or bond, R 1 is not propyl; X is CH 2 and R 1 is CH 2 CH 2 CF 3 , butyl, or pentyl; 2 The phenyl group in must be substituted with fluorine; 2 CH 2 and R 1 When is propyl, R 2 The phenyl group in must be substituted with fluorine; Preferably, the triazolo(4,4-d)-pyrimidine derivative is (1S,2S,3R,5S)-3-[7-[(1R,2S)-2-(3,4-difluorophenyl)cyclopropylamino]-5-(propylthio)-3H-[1,2,3]-triazolo[4,5-d]pyrimidin-3-yl]-5-(2-hydroxyethoxy)-1,2-cyclopentanediol), also known as triafluosyl; or fluometa 5. The nanogel of any one of claims 1 to 4, which is (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol, also known as sil, or Fluometacyl®, or a combination thereof.

6. The biologically active molecule, therapeutic molecule or drug is represented by the formula (4): 【Transformation 5】 [In the formula, X 1 and X 2 are independently N, CH, CR 8 and R 8 is C 1-6 Alkyl, C 2-6 Alkenyl or C 2-6 alkynyl; 1 or X 2 If one of the is equal to N, the remaining X 1 or X 2 , CH, CR 8 Selected from: -Y- is -O- or -S-; R 11 and R 12 independently, C 1-6 -Alkyl, C 2-6 -Alkenyl, C 2-6 -alkynyl, C 3-6 -Cycloalkyl, aryl, aryl-C 1-6 -alkyl, wherein the alkyl or cycloalkyl moiety may be mono- or polysubstituted with OH or halogen, and the aryl moiety may be halogen, —C 1-6 Alkyl, -C 1-6 Alkoxy, -OH, -NO 2 , -CN, -NH 2 , -NHR 8 , -N(R 8 ) 2 -COOH, -COOR 8 , -CONH 2 , -CONHR 8 , -CON(R 8 ) 2、 -SO 2 NH 2 , -SO 2 NHR 8 or -SO 2 N(R 8 ) 2 may be mono- or polysubstituted with; R 13 , R 14 , R 15 , R 16 and R 17 are independently H, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, -OH, -NO 2 , -CN, -NH 2 , -NHR 8 , -N(R 8 ) 2 -COOH, -COOR 8 , -CONH 2 , -CONHR 8 , -CON(R 8 ) 2、 -SO 2 NH 2 , -SO 2 NHR 8 or -SO 2 N(R 8 ) 2 is] The nanogel according to any one of claims 1 to 4, which is a pyrimidine derivative represented by the formula:

7. 3. The nanogel of claim 2, wherein the ratio of the biologically active molecule, therapeutic molecule or drug to D-α-tocophenyl polyethylene glycol succinate is 1:1 w / w to 1:5 w / w.

8. 4. The nanogel of claim 3, wherein the ratio of bioactive molecule, therapeutic molecule or drug to hydroxypropyl-β-cyclodextrin is from 1:1 w / w to 1:200 w / w.

9. 9. The nanogel according to any one of claims 1 to 8, wherein the one or more biologically active molecules, therapeutic molecules or drugs are bacteriostatic agents in combination with antivirulence or bactericidal agents, preferably minocycline or chlorhexidine.

10. The nanogel of any one of claims 1 to 9, wherein the bioactive molecule, therapeutic molecule or drug is fluometacyl or a combination of Fluometacyl® and chlorhexidine.

11. The nanogel of any one of claims 1 to 10, wherein the bioactive molecule, therapeutic molecule or drug is fluometacyl or a combination of Fluometacyl® with chlorhexidine and polymyxin B.

12. A nanogel according to any one of claims 1 to 11 for use in the treatment or prevention of bacterial infections.

13. A nanogel according to any one of claims 1 to 12 for use in topical administration.

14. A biomaterial implant, medical device or bioprosthesis, the surface of which or part of which is coated with a nanogel according to any one of claims 1 to 11.

15. The biomaterial implant, medical device according to claim 14, wherein the medical device is a catheter.

16. 1. A method for making a nanogel comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, the nanogel being obtained by one of the following methods, depending on whether each bioactive molecule, therapeutic molecule or drug is loaded separately or simultaneously with the amphiphilic molecules into the nanogel: a) When each bioactive molecule, therapeutic molecule or drug is loaded separately with an amphiphilic molecule, the method comprises the following steps in order: i) Equation (1) 【Transformation 6】 wherein x is an integer greater than 1, preferably 1 to 100. and / or a poly(methacrylamide) having a quinoline group represented by formula (6): 【Transformation 7】 wherein n is an integer greater than 1, preferably n is 1 to 100. mixing a poly(vinylquinoline) of the formula: with only one bioactive molecule, therapeutic molecule or drug together with an amphiphilic molecule; ii) adding a solution of a polymer having primary or secondary amine groups to the mixture obtained in step i) to obtain a crosslinked nanogel containing one bioactive molecule, therapeutic molecule or drug together with amphiphilic molecules; iii) repeating steps i) and ii) for each additional biologically active molecule, therapeutic molecule, or drug; iv) mixing the crosslinked nanogels obtained in step iii) to obtain a nanogel containing two or more biomolecules, therapeutic molecules or drugs together with amphiphilic molecules; b) When the bioactive molecule, therapeutic molecule or drug is loaded simultaneously with the amphiphilic molecule, the method comprises the following steps in order: i) Equation (1) 【Transformation 8】 wherein x is an integer greater than 1, preferably 1 to 100. and / or a poly(methacrylamide) having a quinoline group represented by formula (6): 【Chemistry 9】 wherein n is an integer greater than 1, preferably n is 1 to 100; mixing a poly(vinylquinoline) of the formula: with one or more bioactive molecules, therapeutic molecules or drugs together with an amphiphilic molecule; ii) adding a solution of a polymer having primary or secondary amine groups to the mixture obtained in step i) to obtain a nanogel containing one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules.

17. 17. The method of claim 16, wherein the amphiphilic molecule is a vitamin E derivative, preferably D-α-tocophenyl polyethylene glycol succinate.

18. 17. The method of claim 16, wherein the amphiphilic molecule is an amphiphilic cyclodextrin, preferably hydroxypropyl-β-cyclodextrin.

19. 17. The method of claim 16, wherein the ratio of the biologically active molecule, therapeutic molecule or drug to D-α-tocophenyl polyethylene glycol succinate is 1:1 w / w to 1:5 w / w.

20. 17. The method of claim 16, wherein the ratio of the biologically active molecule, therapeutic molecule or drug to hydroxypropyl-β-cyclodextrin is 1:1 to 1:

200.

21. The one or more biologically active molecules, therapeutic molecules, or drugs are triazolo(4,4-d)-pyrimidine derivatives of formula (3); preferably, the triazolo(4,4-d)-pyrimidine derivative is (1S,2S,3R,5S)-3-[7-[(1R,2S)-2-(3,4-difluorophenyl)cyclopropylamino]-5-(propylthio)-3H-[1,2,3]-triazolo[4,5-d]pyrimidin-3-yl]-5-(2-hydroxyethyl ... or (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol, also known as Fluometacyl, or Fluometacyl®, or a combination thereof.

22. The biologically active molecule, therapeutic molecule or drug is represented by the formula (4): 【Chemistry 10】 [In the formula, X 1 and X 2 are independently N, CH, CR 8 and R 8 is C 1-6 Alkyl, C 2-6 Alkenyl or C 2-6 alkynyl; 1 or X 2 If one of the is equal to N, the remaining X 1 or X 2 , CH, CR 8 Selected from: -Y- is -O- or -S-; R 11 and R 12 independently, C 1-6 -Alkyl, C 2-6 -Alkenyl, C 2-6 -alkynyl, C 3-6 -Cycloalkyl, aryl, aryl-C 1-6 -alkyl, wherein the alkyl or cycloalkyl moiety may be mono- or polysubstituted with OH or halogen, and the aryl moiety may be halogen, —C 1-6 Alkyl, -C 1-6 Alkoxy, -OH, -NO 2 , -CN, -NH 2 , -NHR 8 , -N(R 8 ) 2 -COOH, -COOR 8 , -CONH 2 , -CONHR 8 , -CON(R 8 ) 2、 -SO 2 NH 2 , -SO 2 NHR 8 or -SO 2 N(R 8 ) 2 may be mono- or polysubstituted with; R 13 , R 14 , R 15 , R 16 and R 17 are independently H, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, -OH, -NO 2 , -CN, -NH 2 , -NHR 8 , -N(R 8 ) 2 -COOH, -COOR 8 , -CONH 2 , -CONHR 8 , -CON(R 8 ) 2、 -SO 2 NH 2 , -SO 2 NHR 8 or -SO 2 N(R 8 ) 2 is] or an optical isomer thereof, a racemic mixture thereof, a pharmaceutically acceptable acid addition salt, a pharmaceutically acceptable metal salt or alkylated ammonium salt, or a prodrug thereof.

23. 21. The method of any one of claims 16 to 20, wherein the one or more biologically active molecules, therapeutic molecules or drugs is a bacteriostatic agent, preferably minocycline or chlorhexidine, in combination with an antivirulence agent or bactericidal agent.

24. 1. A method for producing a medical device, biomaterial implant or bioprosthesis having a nanogel-coated surface comprising one or more bioactive molecules, therapeutic molecules or drugs in combination with amphiphilic molecules, the method comprising the following steps in order: ci) optionally immersing the surface to be coated in a dopamine buffer solution; cii) immersing the surface coated in step ci) in a solution of a polymer having primary or secondary amine groups; and ciii) immersing the coated surface obtained in step cii) in a liquid suspension of crosslinked nanogels comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules; and civ) drying the crosslinked coated surface obtained in step ciii) to obtain a coated crosslinked nanogel surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules; cv) optionally repeating steps cii) to civ) to obtain a surface covered with several layers of crosslinked nanogels containing one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules. A method comprising:

25. 1. A method for producing a medical device, biomaterial implant or bioprosthesis having a coated surface, the coating comprising the following sequence of steps: i) optionally immersing the surface to be coated in a dopamine buffer solution; ii) immersing the surface coated with dopamine in step i) in a solution of a polymer having primary or secondary amine groups; and iii) The coated surface obtained in step ii) is treated with a compound represented by the formula (1) 【Chemistry 11】 wherein x is an integer greater than 1, preferably 1 to 100. and / or a poly(methacrylamide) having a quinoline group represented by formula (6): 【Chemistry 12】 wherein n is an integer greater than 1, preferably n is 1 to 100. soaking a mixture of poly(vinylquinoline) and one or more bioactive molecules, therapeutic molecules, or drugs together with amphiphilic molecules; iv) drying the cross-linked coated surface obtained in step iii) to obtain a coated cross-linked monolayer surface comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules; v) Optionally, repeating steps ii) to iv) to obtain a surface covered with several layers of cross-linked multilayers comprising one or more bioactive molecules, therapeutic molecules or drugs together with amphiphilic molecules, preferably vitamin E derivatives. The method obtained is:

26. 26. The method according to claim 24 or 25, wherein the amphiphilic molecule is a vitamin E derivative, preferably D-α-tocophenyl polyethylene glycol succinate.

27. 26. The method according to claim 24 or 25, wherein the amphiphilic molecule is an amphiphilic cyclodextrin, preferably hydroxypropyl-β-cyclodextrin.

28. 28. The method according to any one of claims 24 to 27, wherein the one or more biologically active molecules, therapeutic molecules or drugs is a bacteriostatic agent, preferably minocycline or chlorhexidine, in combination with a triazolo(4,4-d)-pyrimidine derivative or pyrimidine derivative.

29. The one or more biologically active molecules, therapeutic molecules, or drugs may be (1S,2S,3R,5S)-3-[7-[(1R,2S)-2-(3,4-difluorophenyl)cyclopropylamino]-5-(propylthio)-3H-[1,2,3]-triazolo[4,5-d]pyrimidin-3-yl]-5-(2-hydroxyethoxy)-1,2-cyclopentanediol), also known as triafluosyl; or fluometasil 28. The method of any one of claims 24 to 27, wherein the compound is (1S,2R,3S,4R)-4-[7-[[(1R,2S)-2-(3,4-difluorophenyl)-cyclopropyl]amino]-5-(propylthio)-3H-1,2,3-triazolo[4,5-d]pyrimidin-3-yl]-1,2,3-cyclopentanetriol, also known as Fluometacyl®, or Fluometacyl®, or a combination thereof.

30. A pharmaceutical composition comprising a nanogel according to any one of claims 1 to 11 for use in the treatment or prevention of bacterial pathogenicity in a host mammal by topical administration.

31. Use of a nanogel according to any one of claims 1 to 11 as an inhibitor of cell adhesion on the surface of a biomaterial implant, medical device or bioprosthesis.

32. 32. The use according to claim 31 , wherein the biomaterial implant, medical device or bioprosthesis is a catheter.

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