Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
  • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
  • A61K9/51—Nanocapsules; Nanoparticles
  • A61K9/5107—Excipients; Inactive ingredients
  • A61K9/5123—Organic compounds, e.g. fats, sugars
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/10—Dispersions; Emulsions
  • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
  • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Definitions

• the present invention aims at providing nanocapsules with a liquid lipid core loaded with active agents at this core into at least one water-soluble or water-dispersible active agent.
• Nanovesicular systems of nanocapsule or nanodroplet type whose size varies from 50 to 500 nanometers and formed of a liquid or solid core, surrounded by a hydrophobic outer membrane, are already known.
• the constituents of their membrane may be synthetic, for example of polymeric, protein or lipid nature like liposomes.
• liposomes which have a lamellar structure formed of a stack of lipid layers separated from one another by aqueous compartments always have an aqueous core.
• nano-metric structures have also already been proposed for the purpose of encapsulating active agents either in their aqueous core when the active agent is water-soluble or water-dispersible, or in their lipid layer when the active ingredient is liposoluble or lipodispersible.
• US Pat. No. 5,961,970 proposes, as an active agent, submicron-scale oil-in-water emulsions, that is to say mini-emulsions whose droplets have a hydrophobic core of lipidic nature and are stabilized. at the surface by amphiphilic and / or nonionic surfactants, like phospho lipid-type surfactants. These droplets are thus kept in suspension in an aqueous phase.
• This type of submicron emulsion is obtained from a base emulsion by subjecting it to several successive cycles of homogenization under high shear.
• US Patent 5,576,016 describes macroemulsions whose droplets are formed of a solid lipid core and which are stabilized by a phospholipid envelope.
• This phospholipid envelope has a lamellar structure formed of one or more layers of phospholipid molecules like liposomes.
• a highly hydrophobic active agent can be loaded at the core and a water-soluble active agent can be incorporated into the aqueous compartments present in the phospholipid envelope.
• an emulsion for example W / O
• a temperature which must be greater than the phase inversion temperature of the system ie the temperature at which the equilibrium between the hydrophilic and lipophilic properties of the surfactant system used is achieved.
• the emulsion is of the water-in-oil type, and during its cooling, this emulsion is reversed. at the phase inversion temperature, to become an oil-in-water type emulsion, and this having previously passed through a microemulsion state.
• This technique notably makes it possible to access an average size of the globules constituting the oily phase ranging from 0.1 to 4 ⁇ m (100 to 4000 nm).
• nanocapsules thus obtained are only proposed for the purpose of encapsulation, within their oily heart, of lipophilic or lipodispersible active agents.
• the water-dispersible active agent is advantageously a non-lipophilic active agent.
• the present invention aims precisely to propose a solution for performing this type of encapsulation.
• the present invention aims, according to a first aspect, with nanocapsules having a liquid lipid core and solid lipid shell and loaded within their lipid core with at least one water-soluble or water-dispersible active agent, said active being incorporated therein in the form of of a reverse micellar system.
• the present invention relates to a method useful for preparing nanocapsules with liquid lipid core and solid lipid shell and loaded at their lipid core into at least one water-soluble or water-dispersible active agent, said process comprising at least the steps comprising: disposing of an oil-in-water emulsion containing at least one water-soluble or water-dispersible active agent in the form of a reverse micellar system and at least one surfactant system containing at least one thermosensitive, nonionic and hydrophilic surfactant, and if necessary a lipophilic surfactant increase its temperature to a temperature T 2 greater than its phase inversion temperature (TIP) to obtain a water-in-oil emulsion followed by a decrease in temperature to a temperature T 1 , Ti ⁇ TIP ⁇ T 2 to obtain an oil-in-water emulsion again, carry out one or more temperature cycles around the phase inversion zone between Ti and T 2 , and stabilizing said system at a temperature in or near the phase inversion
• TIP phase in
• an oil-in-water emulsion containing at least one water-soluble or water-dispersible active agent in the form of an inverse micellar system proves capable of forming, when it imposes at least a phase inversion operation temperature, a microemulsion whose quenching leads to the formation of nanocapsules containing within their lipid core micelles water soluble or hydrodispersible assets.
• the progress of all the steps required to obtain the expected nanocapsules does not affect the stability of the reverse micellar system of the water-soluble or water-dispersible active.
• the expression "reverse micellar system of water-soluble or water-dispersible active” designates an architecture in which the water-soluble or water-dispersible active agents are stabilized in an oily phase by means of the molecules of the surfactant or of the surfactant system forming the micellar system containing the active.
• the water-dispersible active agent is advantageously a non-lipophilic active agent.
• Reverse micelle systems are well known to those skilled in the art and in particular exploited to carry out selective extractions of proteins or enzymes of interest.
• the choice of the surfactant system used to form the reverse micellar system must be carried out taking into account the solubility of the surfactant (s) forming it in the oily phase of the oil-in-water emulsion in which the asset is precisely intended to be formulated. This selection is clearly within the skill of those skilled in the art.
• the surfactants used to develop these reverse micelles and suitable for the invention have an HLB value of less than 10, in particular less than or equal to 6. They may indifferently belong to the families of ionic, nonionic or amphoteric surfactants.
• surfactants may be used in an active (s) / surfactant (s) weight ratio ranging from 0.01 to 0.3 and in particular from 0.05 to 0.1.
• these surfactants may be combined with co-surfactants such as, for example, phospholipids.
• co-surfactants such as, for example, phospholipids.
• phosphatidylcholines lecithin are particularly interesting.
• phospholipids suitable for the invention may be phosphatidylglycerol, phosphatidylinositol, phosphatidylserine, phosphatidic acid and phosphatidylethanolamine.
• the reverse micellar system containing the water-soluble or water-dispersible active agent to be encapsulated is prepared before it is brought into contact with the emulsion so as to stabilize the inverse micelles in the oil.
• This oily micellar suspension is then either incorporated at the beginning of the formulation before the thermal cycling (s) or else, added once the system is in its microemulsion form, that is to say in the phase inversion zone. of the emulsified system.
• a hydrophilic active agent for example sodium fluororescein crystals, is incorporated under heating at 50 ° C. and with stirring in an oily phase, for example Labrafac containing inverse micelles of a surfactant such as Span 80® (10%). w / w).
• water-soluble or water-dispersible active agents that can be encapsulated according to the invention include 5-fluorouracil, gemcitabine, doxorubicin and low molecular weight heparins.
• the nanocapsules can of course contain other water-soluble active agents adsorbed on their bark or other liposoluble active agents encapsulated in their lipid core.
• the term "adsorbed" means that the active ingredient is incorporated within the bark. This adsorption phenomenon is to be distinguished from a simple covalent bond established between a function present on said active agent and a function present on the surface of the bark of the nanocapsules.
• the active agent may especially be a pharmaceutically active compound, cosmetically active or still active in a phytosanitary or food field.
• this active ingredient is a pharmaceutically active ingredient.
• the nanocapsules of the invention are more particularly suitable for the administration of the following active principles: anti-infectives including antimycotics, antibiotics, anticancer drugs, immunosuppressants, active principles for the Central Nervous System who must pass the blood-brain barrier, such as anti-Parkinson's, analgesics and more generally the active ingredients for treating neurodegenerative diseases.
• anti-infectives including antimycotics, antibiotics, anticancer drugs, immunosuppressants, active principles for the Central Nervous System who must pass the blood-brain barrier, such as anti-Parkinson's, analgesics and more generally the active ingredients for treating neurodegenerative diseases.
• Such an active ingredient may also be of protein or peptide nature. It may also be nucleic acids such as a DNA plasmid or an interference RNA.
• the asset can also be a radiopharmaceutical. It can also be a gas or a fluid that can turn into gas.
• a microemulsion is different from a miniemulsion and a macroemulsion, such as illustrated in US Pat. Nos. 5,961,971 and 5,576,016.
• a microemulsion corresponds to a bicontinuous structuring of the material in the form of micellar structures swollen with oil or water. These micellar structures are strongly intertwined with each other and thus constitute a cohesive and stabilized homogeneous three-dimensional network. It is thus impossible to distinguish a dispersed phase from a continuous phase.
• This microemulsion is in thermodynamic equilibrium and therefore can only exist under very precise conditions of temperature, pressure and composition.
• the starting emulsion intended to form the microemulsion may comprise at least one oily fatty phase, an aqueous phase and a surfactant system comprising at least one heat-sensitive, hydrophilic and nonionic surfactant and, where appropriate, according to a preferred embodiment, a lipophilic surfactant.
• the oily fatty phase is formed of at least one liquid or semi-liquid fatty substance at ambient temperature, and in particular at least one triglyceride, a fatty acid ester, or a mixture thereof.
• the fatty acid ester may more particularly be chosen from C 8 to C 18 fatty acid esters, in particular C 8 to C 12 fatty acid esters, and in particular ethyl palmitate, ethyl oleate or myristate. ethyl, isopropyl myristate, octydodecyl myristate and mixtures thereof.
• the triglycerides used may be synthetic triglycerides or triglycerides of natural origin.
• Natural sources may include animal fats or vegetable oils eg soybean oils or sources of long chain triglycerides (LCTs).
• MCT medium chain triglycerides
• An oil to Medium chain triglycerides (MCT) is a triglyceride in which the carbohydrate chain has from 8 to 12 carbon atoms.
• Such MCT oils are commercially available.
• TCRs trade name of the industrial oilseeds company, France, for a mixture of triglycerides in which about 95% of the fatty acid chains have 8 or 10 carbon atoms.
• Myglyol ® 812 triglyceride marketed by Dynamit Nobel, Sweden for a mixture of triesters of caprylic and capric acid glycerides.
• the fatty acid units of these triglycerides may be unsaturated, monounsaturated or polyunsaturated. Mixtures of triglycerides with variable fatty acid units are also acceptable.
• HLB index or hydrophilic-lipophilic balance is as defined by C. Larpent in the K.342 Treaty of TECHNIQUES DE L'INGENIEUR.
• the triglyceride sold under the name Labrafac WL 1349 ® is particularly suitable for the invention.
• This surfactant system comprises at least one thermosensitive surfactant, hydrophilic and nonionic, which may be associated with a lipophilic surfactant.
• the surfactant system used in the claimed process is formed only of hydrophilic (s) and nonionic (s) heat-sensitive surfactant (s).
• the surfactant system used in the microemulsion may comprise one or more surfactants whose solubility in the oil increases with increasing temperature.
• the HLB of these surfactants can vary from 8 to 18, and preferably from 10 to 16, and these surfactants can be chosen from ethoxylated fatty alcohols, ethoxylated fatty acids, partial glycerides of ethoxylated fatty acids, triglycerides of polyethoxylated fatty acids, and mixtures thereof.
• ethylene oxide adducts with lauryl alcohol especially those containing from 9 to 50 oxyethylenated groups (Laureth-9 to Laureth-50 in CTFA names); adducts of ethylene oxide with behenyl alcohol, in particular those containing from 9 to 50 oxyethylenated groups (Beheneth-9 to Beheneth-50 in CTFA names); adducts of ethylene oxide with keto-stearyl alcohol (mixture of cetyl alcohol and stearyl alcohol), in particular those containing from 9 to 30 oxyethylenated groups (Ceteareth-9 to Ceteareth-30 in names) CTFA); adducts of ethylene oxide with cetyl alcohol, especially those containing from 9 to 30 oxyethylenated groups (Ceteth-9 to Ceteth- in CTFA names); adducts of ethylene oxide with stearyl alcohol, especially those containing those containing
• ethoxylated fatty acids mention may be made, for example, of ethylene oxide adducts with lauric, palmitic, stearic or behenic acids, and mixtures thereof, in particular those comprising from 9 to 50 oxyethylenated groups such as laurates of PEG-9 to PEG-50 (CTFA names: PEG-9 laurate to PEG-50 laurate); palmitates of PEG-9 to PEG-50 (in CTFA names: PEG-9 palmitate to PEG-50 palmitate); stearates from PEG-9 to PEG-50 (CTFA names: PEG-9 stearate PEG-50 stearate); palmitostearate from PEG-9 to PEG-50; behenates from PEG-9 to PEG-50 (in CTFA names: PEG-9 behenate to PEG-50 behenate); and their mixtures.
• CTFA names: PEG-9 laurate to PEG-50 laurate palmitates of PEG-9 to PEG-50 (in CTFA names: PEG-9 palmitate to
• surfactants can also be either natural compounds such as echolate phospholipids or synthetic compounds such as polysorbates which are fatty acid esters of polyethoxylated sorbitol (Tween ®), polyethylene glycol fatty acid esters eg from castor oil (Cremophor ®), polyethoxylated fatty acids, eg stearic acid (Simulsol M-53 ®), fatty alcohol ethers polyoxyethylene (Brij ®), polyoxyethylene nonphenyl ethers (Triton ® N) of hydroxylphényle polyoxyethylene ethers (Triton ® X).
• Teween ® polyethoxylated sorbitol
• Polyethylene glycol fatty acid esters eg from castor oil (Cremophor ®
• polyethoxylated fatty acids eg stearic acid
• Simulsol M-53 ® fatty alcohol ethers polyoxyethylene
• Brij ® polyoxyethylene nonphen
• It can especially be a 2-hydroxystearate polyethylene glycol and in particular that marketed under the name Solutol ® HS 15 by BASF (Germany).
• the surfactant system selected according to the invention advantageously comprises, besides the heat-sensitive, hydrophilic and non-ionic surfactant, at least one lipophilic surfactant.
• the lipophilic surfactant is more particularly based on phospholipids which are advantageous in view of their biocompatible nature.
• phosphatidylcholines are particularly interesting.
• phospholipids may be phosphatidylglycerol, phosphatidylinositol, phosphatidylserine, phosphatidic acid and phosphatidylethanolamine.
• Phospholipid derivatives can be isolated from natural sources or synthetically prepared.
• EPICURON 120 ® (Lukas Meyer, Germany) which is a mixture of about 70% phosphatidylcholine, 12% phosphatidylethanolamine and about 15% other phospholipids;
• - VO VOTINE 160 ® (Lukas Meyer, Germany) which is a mixture comprising about 60% phosphatidylcholine, 18% phosphatidylethanolamine and 12% other phospholipids,
• the lipophilic surfactant is a lecithin whose proportion of phosphatidylcholine ranges from 40 to 80% by weight.
• Lipoid S75-3 Lipoid S75-3 (Lipoid ® GmbH, Germany). This is soy lecithin. The latter contains about 69% phosphatidylcholine and 9% phosphatidylethanolamine. This component is the only solid component at 37 ° and at room temperature in the formulation.
• the ratio of liquid fatty substance / lipid surfactant (s) may vary from 1 to 15, preferably from 1.5 to 13, more preferably from 3 to 8.
• the particle size decreases when the proportion of hydrophilic surfactant increases and when the proportion of surfactants (hydrophilic and optionally lipophilic) increases.
• the surfactant causes a decrease in the interfacial tension and thus a stabilization of the system which promotes the obtaining of small particles.
• the aqueous phase of the microemulsion may also advantageously contain 1 to 4% of a particularly inorganic salt, such as sodium chloride.
• a particularly inorganic salt such as sodium chloride.
• the microemulsion advantageously contains from 1 to 3% of lipophilic surfactant (s), from 5 to 15% of hydrophilic surfactant (s), from 5 to 15% of an oily phase, from 64 to 89% of an aqueous phase (the percentages are expressed by weight relative to the total weight of the microemulsion).
• a microemulsion of the invention may be formed of at least one fatty acid triglyceride and a 2-hydroxysterate derivative of polyethylene glycol, and optionally a lecithin.
• the fatty phase is a fatty acid triglyceride
• the lipophilic surfactant is a lecithin
• the hydrophilic surfactant is a derivative of 2-hydroxystearate polyethylene glycol and in particular Solutol ® HS 15.
• the emulsion considered according to the invention is converted into a microemulsion according to the phase inversion technique, in particular by temperature change.
• This phase inversion in temperature is advantageously caused by imposing at least one cycle of rise and fall in temperature to the emulsion.
• the temperature of the phase inversion zone tends to decrease as and when the temperature cycles imposed. This phenomenon is precisely advantageous when the asset that it is desired to encapsulate or adsorb is a temperature-sensitive asset. Under such conditions, it is preferred to introduce the asset at the time of a temperature-compatible cycle.
• TIP phase inversion temperature
• phase inversion zone between Ti and T 2 it is advantageous to carry out one or more temperature cycles around the phase inversion zone between Ti and T 2 , until a translucent suspension is observed, which corresponds to the formation of a microemulsion.
• the system is then stabilized at a temperature which corresponds to the structure of the system in the expected microemulsion.
• the phase inversion between the oil / water emulsion and the water / oil emulsion results in a decrease in conductivity as the temperature increases until it vanishes.
• Ti is a temperature at which the conductivity is at least 90 - 95% of the conductivity measured at 20 0 C and T 2 is the temperature at which the conductivity vanishes and the water-in-oil emulsion is formed.
• TIP phase inversion temperature
• a microemulsion In the formation zone of a microemulsion (translucent mixture), the hydrophilic and hydrophobic interactions are balanced because the tendency of the surfactant system is to form both direct micelles and inverse micelles.
• W / O emulsion opaque white mixture
• the surfactant promotes the formation of a water-in-oil emulsion.
• the emulsion becomes an O / W emulsion.
• the number of cycles applied to the microemulsion depends on the amount of energy required to form the nanocapsules.
• the mixture is homogenized, for example by means of a Rayneri 350 rpm, and heated by progressively increasing the temperature by means of a water bath to a temperature greater than or equal to the T2 phase inversion temperature, that is to say until a transparent or translucent phase is obtained (zone of microemulsion or lamellar phase) then of a more viscous white phase which indicates the obtaining of the inverse emulsion (E / H).
• the heating is then stopped and the stirring maintained until it returns to ambient temperature, via the phase inversion temperature T1, that is to say the temperature at which the expected microemulsion is formed.
• T1 phase inversion temperature
• the microemulsion having formed subsequently undergoes quenching according to the invention.
• This step intended to form the nanocapsules according to the invention consists in an abrupt cooling (or quenching) of the microemulsion at a temperature conducive to the solidification of the interfacial films comprising the microemulsion, advantageously at a temperature much lower than T1 under magnetic stirring.
• This quenching can be carried out by diluting the medium 3 to 10 times with deionized water at 20 ° C +/- 1 ° C., which is thrown into the fine microemulsion.
• the nanocapsules obtained are stirred for 5 minutes.
• the organization of the system in the form of nanocapsules after soaking is reflected visually by a change of appearance of the initial system which changes from opaque white to translucent white with Tyndall effect (bluish tints). This change occurs at a temperature below the TIP. This temperature is generally between 6 to 15 0 C below the TIP.
• nanocapsules loaded at their lipid core into at least one water-soluble active agent are obtained.
• nanocapsules is to be distinguished from nanospheres.
• Nanocapsules are understood to mean particles consisting of a liquid or semi-liquid core at ambient temperature, coated with a solid film at ambient temperature, as opposed to nanospheres which are matrix particles, ie whose whole mass is solid. .
• the nanospheres contain a pharmaceutically active principle, it is finely dispersed in the solid matrix.
• the nanocapsules obtained according to the invention have a mean size of less than 150 nm, preferably less than 100 nm, more preferably less than 50 nm. These sizes can be determined by photon correlation spectroscopy, scanning electron microscopy, transmission electron microscopy in cryoscopic mode.
• the thickness of the solid film or bark is advantageously between 2 and 10 nm. It is equal to about one-tenth of the diameter of the particles. This thickness can be calculated by mass balance, or visualized by transmission electron microscopy. Negative shading or else by transmission electron microscopy in cryoscopic mode.
• the nanocapsules of the invention are colloidal lipid particles.
• the polydispersity index of the nanocapsules of the invention is advantageously between 5 and 15%. This index is determined on the size histogram obtained by photon correlation spectroscopy method.
• the nanocapsules are each composed of a substantially liquid or semi-liquid lipid core at room temperature, coated with a substantially lipidic bark solid at room temperature.
• the expression "essentially lipidic" means that the core and the bark forming the nanocapsules according to the invention consist of more than 50% by weight, in particular more than 75% by weight, in particular more than 80% by weight, or even more than 90%, more particularly more than 95% of their respective weight, or even all of one or more lipid compounds (hydrophobic).
• a nanocapsule according to the invention may comprise a bark formed of at least one solid lipophilic surfactant at room temperature.
• nanocapsules according to the invention are particularly advantageous as an asset formulation vehicle.
• these nanocapsules may be useful for the manufacture of phytosanitary or pharmaceutical compositions.
• the present invention also provides compositions containing nanocapsules according to the invention.
• ambient temperature designates a temperature ranging from 18 to 25 ° C.
• Example 1 Preparation of nanocapsules whose lipid core is loaded into a water-soluble active agent
• sodium fluorescein crystals (10 mg) are incorporated under heating at 50 ° C. and with stirring in Labrafac (3 g) containing inverse micelles of Span 80 (0.6 g) (20% w / w). .
• micellar suspension After homogenization of this micellar suspension, 0.25 ml are introduced into the preceding emulsion.
• the whole is placed under magnetic stirring. Heating is applied until a temperature of 85 ° C. is reached. With magnetic stirring, the system is allowed to cool down to a temperature of 60 ° C. These thermal cycles (between 85 ° C. and 60 ° C.) are carried out. three times in order to obtain better and better structured microemulsions. The system is then maintained in its microemulsion form by stabilizing it at a temperature included in (or in the near vicinity) of the Phase Inversion Zone, in this case 65 ° C.
• the nanocapsules are finalized by dipping in cold water (5 0 C).
• the nanocapsules are separated from the medium by centrifugation.

Landscapes

• Health & Medical Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• Veterinary Medicine (AREA)
• Engineering & Computer Science (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Medicinal Chemistry (AREA)
• Pharmacology & Pharmacy (AREA)
• Public Health (AREA)
• Animal Behavior & Ethology (AREA)
• General Health & Medical Sciences (AREA)
• Epidemiology (AREA)
• Nanotechnology (AREA)
• Physics & Mathematics (AREA)
• Molecular Biology (AREA)
• Dispersion Chemistry (AREA)
• Biomedical Technology (AREA)
• Biophysics (AREA)
• Optics & Photonics (AREA)
• Chemical Kinetics & Catalysis (AREA)
• General Chemical & Material Sciences (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Organic Chemistry (AREA)
• Manufacturing Of Micro-Capsules (AREA)
• Medicinal Preparation (AREA)

Applications Claiming Priority (2)

Publications (1)

Family

ID=39046733

Family Applications (1)

Country Status (5)

Families Citing this family (11)

Citations (2)

Family Cites Families (3)

• 2007
  • 2007-06-11 FR FR0755652A patent/FR2916973B1/fr active Active
• 2008
  • 2008-06-11 EP EP08805977A patent/EP2167053A2/de not_active Withdrawn
  • 2008-06-11 CA CA2690471A patent/CA2690471A1/fr not_active Abandoned
  • 2008-06-11 WO PCT/FR2008/051042 patent/WO2009001019A2/fr active Application Filing
  • 2008-06-11 US US12/663,937 patent/US9333180B2/en not_active Expired - Fee Related

Patent Citations (2)

Also Published As

Similar Documents

Legal Events