FR2797398A1 - MICROPARTICLES FOR PULMONARY ADMINISTRATION - Google Patents

Abstract

The invention concerns a biocompatible microparticle designed to be inhaled comprising at least an active principle and at least a layer coating said active particle which is the outer layer of said microparticle, said outer layer comprising at least a coating agent. The invention is characterised in that said microparticle has a mean diameter ranging between 1 mu m and 30 mu m, an apparent density ranging between 0.02 g/cm<3> and 0.8 g/cm<3> and it is obtainable by a method comprising essential steps which consist in bringing together a coating agent and an active principle and introducing a supercritical fluid, under agitation in a closed reactor.

Description

The present invention relates to the field of microparticles intended to be administered by the pulmonary route.

A bibliographic study has shown that a great deal of research relating to this technology has been carried out. Aerosols for the delivery of therapeutic agents into the respiratory tract have been described, for example (Adjei, A. and Garren, J. Pharm. Res., 7: 565-569 (1990); and Zanen, P. and Lamm, JWJ Int. J. Pharm., 114: 111-115 (1995)). The respiratory tract includes the upper respiratory tract which includes the larynx and oropharynx, and the lower respiratory tract including the trachea which continues in bifurcations: the bronchi and bronchioles. The terminal bronchioles then divide into respiratory bronchioles which lead to the ultimate zone of the respiratory system, the pulmonary alveoli also called the deep lung (Gonds, I. Aerosols for delivery of therapeutic and diagnostic agents to the respiratory tract, in Critical Reviews in Therapeutic Drug Carrier Systems, 6: 273-313 (1990)). The deep lung or alveoli are the primary target of therapeutic inhalation aerosols intended for the systemic route. Aerosols intended to be inhaled have previously been used for the treatment of local pulmonary disorders such as asthma and cystic fibrosis (Anderson et al., Am. Rev. Respir. Dis., 140: 1317-1324 (1989)) . In addition, they can be used for the systemic release of peptides and proteins (Patton and Platz, Advanced Drug Delivery Reviews, 8: 179-196 (1992)). However, a number of difficulties are encountered when it is desired to apply the drug release via the pulmonary route to the release of macromolecules. Among these difficulties are the denaturation of the protein during nebulization, a significant loss of the level of drugs inhaled in the oropharynx (which often exceeds 80%), poor control of the deposition zone, poor reproducibility therapeutic results due to variations in respiratory models, too rapid absorption of drugs generating local toxic effects, and phagocytosis by macrophages of the lung.

The human lung can quickly eliminate or degrade hydrolyzable products deposited in the form of aerosols, this phenomenon generally takes place over a period of between a few minutes and a few hours. In the upper pulmonary tract, the ciliated epithelium contributes to the phenomenon of mucociliary escalator by which particles are carried from the pulmonary tract to the mouth (Pavia, D. Lung Mucociliary Clearance, in Aerosols and the Lung Clinical and Experimental Aspects, Clarke, SW and Pavia, D., Eds., Butterworths, London, 1984; Anderson et al., Am. Rev. Respir. Dis., 140 1317-1324 (1989)). In the deep lung, alveolar macrophages are able to phagocytize particles as soon as they are deposited.

Local and systemic inhalation therapies generally allow a controlled and relatively slow release of the active principle (Gonds, I., Physico-chemical principles in aerosol delivery, in Topics in Pharmaceutical Sciences 1991, DJA Crommelin and KK Midha, Eds., Stuttgart: Medpharm Scientific Publishers, pp. 95-117 (1992)). The slow release of the therapeutic aerosol can prolong the residence time of the drug delivered in the pulmonary tract or in the acini and decrease the rate of entry of the drugs into the blood stream. Thus, patient tolerance is increased by reducing the frequency of administration (Langer, R., Science, 249: 1527-1533 (1990); and Gonds, I. Aerosols for delivery of therapeutic and diagnostic agents to the respiratory tract, in Critical Reviews in Therapeutic Drug Carrier Systems 6: 273-313 (1990)). Among the drawbacks represented by the dry powder formulations, there is the fact that the powders of ultra-fine particles exhibit generally poor flow and nebulization properties, leading to the production of aerosol fractions which are allowed in the spray. respiratory system relatively slowly, these fractions of the inhaled aerosol usually settle in the mouth and throat (Gonds, I., in Topics in Pharmaceutical Sciences 1991, D. Crommelin and K. Midha, Editors, Stuttgart: Medpharm Scientific Publishers, 95-117 (1992)). The main problem encountered with most aerosols is particle aggregation generated by inter-particle interactions such as hydrophobic, electrostatic and capillary interactions. Effective dry powder inhalation therapy for both immediate and sustained release of therapeutic agents, both topically and systemically, requires the use of a powder with minimal aggregation that avoids or less than suspending the natural clearance mechanisms of the lung until the moment when the active principle is released. There is currently a demand for improved inhalation aerosols for pulmonary delivery of therapeutic agents. Likewise there is presently a need for drug carriers which are capable of delivering the drug in an effective amount into the pulmonary tract or into the alveolar areas of the lungs. In addition, there is also a need for drug carriers which can be used as inhalation aerosols which are biodegradable and which permit the controlled release of drugs into the pulmonary tract and the alveolar area of the lungs, as well. There is a demand for particles for pulmonary drug delivery which exhibit improved nebulization properties. This research tends to show that it is difficult to prepare microparticles which meet the criteria imposed on them by their applications under effective conditions.

In order to be sufficiently effective, these microparticles must not be damaged during administration, when they pass in nebulized form. The bioavailability of these microparticles must reach a sufficiently high value, and the bioavailability of the microparticles of the prior art generally does not exceed 50%, because of a low rate of deposition of the microparticles in the alveolar pulmonary regions.

In addition, in order to maintain their effectiveness during pulmonary administration, the microparticles, once deposited in the alveoli, must be sufficiently stable in the mucosa of the surface of these alveoli.

Thus it may prove to be advantageous to prepare microparticles with immediate or delayed release, locally or systemically, however these microparticles generally have an outer layer whose thickness relative to the diameter of said particle is not negligible.

The microparticles according to the invention consist of a core containing the active material coated with a membrane of coating agent deposited by the supercritical fluid technique. This particular structure distinguishes them from the microparticles of the prior art which are matrix microspheres obtained by techniques of emulsion-evaporation of solvent, extraction of solvent by aqueous phases or nebulization-drying of an organic solution.

Consequently, the present invention relates to biocompatible microparticles intended to be inhaled comprising at least one active principle and at least one layer coating this active principle which is the outer layer of said microparticles, said outer layer containing at least one coating agent, characterized in that that said microparticles are uniformly coated, have an average diameter of between 1 μm and 30 μm, an apparent density of between 0.02 glcm3 and 0.8 glcm3 and that they can be obtained according to a process comprising the essential steps which are the bringing together of a coating agent with an active principle and the introduction of a supercritical fluid. These microparticles do not agglomerate when they are administered, and may possibly allow a prolonged release of the active principle. The microparticles according to the invention exhibit a bioavailability greater than 60% and preferably greater than 80% by virtue of an improvement in the rate of deposition of the particles in the alveolar pulmonary zones.

It has thus been demonstrated that the implementation of a process for preparing microparticles by a technique known as supercritical fluid using, as coating agent, carefully chosen biocompatible materials makes it possible to obtain microparticles of size controlled and which have a surface condition such that said microparticles do not agglomerate and are deposited in the alveolar pulmonary areas.

The biocompatible microparticles intended for inhalation according to the invention have an outer layer of uniform thickness comprising a coating agent which prevents the aggregation of these particles with one another. They are obtained by a process allowing a coating which matches the surface of these microparticles in these asperities. The quality of this coating is essentially due to the supercritical fluid technique.

Said method comprises two essential steps which are the placing in the presence of a coating agent with an active principle and the introduction of a supercritical fluid in order to ensure the coacervation of the coating agent. It is clear from the remainder of the description that these two steps are not necessarily carried out in the order announced. The first process for preparing the microparticles according to the invention differs from the second process in that the coating agent is at no time in solution in the fluid in the liquid or supercritical state.

In fact, a first covering of the process according to the invention comprises the following steps - suspending an active principle in a solution of at least one substantially polar coating agent in an organic solvent, said active principle being insoluble in the solvent. organic, said substantially polar coating agent being insoluble in a fluid in the supercritical state, said organic solvent being soluble in a fluid in the supercritical state, - bringing the suspension into contact with a fluid in the supercritical state, so as to to desolvate in a controlled manner the substantially polar coating agent and ensure its coacervation, - substantially extract the solvent by means of a fluid in the supercritical state and evacuate the SC fluid / solvent mixture, - recover the microparticles. The fluid used for the implementation of this first process is preferably liquid CO 2 or in the supercritical state (SC fluid).

The organic solvent used for the covering of this first process is generally chosen from the group consisting of ketones, alcohols and esters. The supercritical fluid is brought into contact with the suspension of active principle containing the coating agent in solution, either by introducing the supercritical fluid into an autoclave already containing the suspension, or by injecting the suspension into an autoclave containing the supercritical fluid. .

When the supercritical fluid employed is CO 2, it is possible to use CO 2 in liquid form or directly CO 2 in the supercritical state. According to another variant, the suspension can also be brought into contact with liquid CO 2 which will then pass to the supercritical state by increasing the pressure and / or the temperature in the autoclave in order to extract the solvent.

When one chooses to use the liquid COZ variant, the temperature is preferably chosen between 20 and 50 C and the pressure between 50 and 150 105 Pa. When the supercritical COZ variant is used, the temperature is generally chosen between 35 and 60 C, preferably between 35 and 50 C, and the pressure between 80 and 250 105 Pa, preferably between 100 and 220 105 Pa. The mass of organic solvent introduced into the autoclave represents at least 3%, preferably between 3 , 5% and 25% of the mass of the supercritical fluid or liquid used to cause the desolvation of the coating agent. The microparticles obtained by the implementation of this first process have an outer layer which is practically free from solvent, the amount of solvent in the outer layer is in fact less than 500 ppm. In addition, the microparticles thus obtained exhibit a perfectly smooth surface state (absence of roughness).

The coating agents which can be used for the implementation of this first process are more particularly the biodegradable (co) polymers of α-hydroxycarboxylic acids, in particular the homopolymers and copolymers of lactic and glycolic acids, and more particularly PLA (Poly-L- lactide) and PLGA (Poly-Lactic-co-Glycolic-Acid), biocompatible polymers of polyethylene glycol type, polyethylene oxides, block copolymers of polyethylene oxide-polypropylene oxide type, and polysaccharides. The implementation of the second method according to the invention consists in placing an active principle in suspension in a supercritical fluid containing at least one coating agent dissolved therein and then in modifying the pressure and / or temperature conditions of the medium to ensuring the coacervation of the particles, by precipitation of the coating agent around the particles of active principle, that is to say ensuring the coacervation of the particles by physicochemical modification of the medium.

The coating agents which can be used for the implementation of this second process are more particularly phospholipids such as in particular phosphatidylcholine (PC), phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), dipalmitoyl-phosphatidylcholine (DPPC), dioleyl -phophatidylethanolamine (DOPE), dioleyl-phosphatidylcholine (DOPC), dimyristoyl-phosphatidylglycerol (DMPG), fatty acid esters such as isopropyl myristate, ethyl palmitate. Biodegradable or bioerodible polymers soluble in a supercritical fluid can also be used in this second process.

The coacervation (or aggregation) of a coating agent is caused by physicochemical modification of a medium containing an active substance in suspension in a solution of coating agent in a solvent. The supercritical fluid preferentially used is supercritical CO2 (C02SC), the initial operating conditions typical of this second process will be around 31 to 80 C and the pressures from 75 to 250 105 Pa, although values can be used. higher of one or the other of the two parameters or both, provided of course that the higher values have no harmful or degradation effect on the active principle being coated, nor on the coating agents.

Moreover, one can also choose other fluids commonly used as supercritical fluid, the document E.M. Phillips and V.J. Stella, lnt. <I> J. </I> Pharm, 94, 1 to 10, 1993 - Rapid Expansion From Supercritical Solutions: Application To Pharmaceutical Processes specifies the conditions of use of a certain number of these fluids in their supercritical state. This second process involves the suspension in an autoclave of an insoluble active principle, including the supercritical fluid, then the introduction into this autoclave of the coating agent which is in the form of a solute in the supercritical fluid.

The pressure and / or the temperature are then modified so as to decrease the solubility of the coating agent in the fluid. Thus the affinity of the coating agent for the active principle increases in such a way that this coating is adsorbed around the active principle. Once this coating agent has been deposited on the active principle, the autoclave is depressurized and the microparticles are recovered. To implement this second process, the active principle to be coated is placed in an autoclave equipped with a stirrer, then the system is pressurized by introducing into the autoclave a fluid brought under supercritical conditions. Finally, the coating agent (s) is introduced into the autoclave, then the temperature and / or the pressure inside the autoclave is modified in a controlled and regulated manner so as to gradually reduce the solubility of the coating agent (s). . When the solubility of this or these coating agent (s) in the supercritical fluid decreases, they precipitate) and the affinity of these agents for the surface of the active principle leads to their adsorption on this surface. A variant of this process consists in placing the coating agent in the autoclave before introducing the active principle therein or alternatively by simultaneously introducing therein the active principle and then a fluid capable of passing into the supercritical state. Pressurizing the autoclave to produce a supercritical fluid state will then cause the coating agent to dissolve in said supercritical fluid.

The coating agent is thus deposited in such a way that this agent faithfully matches the surface of the active principle. The active principle can be in the form of a liquid which can thus form an emulsion in the supercritical fluid, of preformed solid particles, and in particular of microparticles optionally already coated, for example with mono- or disaccharides. The stirring speeds can vary between 200 and 400 revolutions / min for solid particles and between 600 and 1000 revolutions / min when the active principle is a liquid. Such agitation ensures the suspension of the active principle in the supercritical fluid when the latter is introduced. The supercritical conditions are ensured by a modification of the temperature and / or the pressure inside the autoclave. Thus, the temperature of the autoclave is between 35 and 80 C, preferably between 35 and 45 C and the pressure is between 100 and 250 105 Pa and preferably between 180 and 220 105 Pa. The coating agent is introduced. in the autoclave at the same time as the supercritical fluid or after the introduction into the autoclave of the supercritical fluid. In any case, to ensure good solubilization of the coating agent in the supercritical fluid, the system is kept in equilibrium with stirring, the adequate concentration of active principle and of coating agent is established as a function of the desired microparticle and one is established. leave this equilibrium under agitation for one hour. The temperature and the pressure are then modulated at a sufficiently slow rate to completely transfer the coating agent (s) from the supercritical fluid to the surface of the active principle and the system is depressurized to isolate the microparticles which are removed from the autoclave.

The microparticles according to the present invention have a diameter of between 1 Nm and 30 μm, preferably between 2 μm and 15 Nm, and even more preferably between 3 Nm and 8 Nm and an apparent density of between 0.02 glcm3 and 0.8 glcm3 and preferably between 0.05 glcm3 and 0.4 g / cm3.

The active principle / coating agent mass ratio of these microparticles is between 95/5 and 5/95. In the case of controlled release microparticles, the amount of active principle is low compared to the coating agent, the active principle / coating agent mass ratio is then between 5I95 and 20/80, on the contrary in the case where the coating is intended to stabilize the particle, in particular when the microparticle is immediate release, the active principle / coating agent mass ratio is generally between 95/5 and 70/30 and preferably between <B> 9515 </B> and 80I20 . The coating agents for the microparticles according to the invention are - biodegradable (co) polymers of α-hydroxycarboxylic acids, in particular homopolymers and copolymers of lactic and glycolic acids, and more particularly PLA (Poly-L-lactide) and PLGA ( Poly-Lactic-co-Glycolic-Acid), biocompatible polymers of polyethylene glycol type, polyethylene oxides, block copolymers of polyethylene oxides-polypropylene oxides type, and polysaccharides, or a mixture of at least two compounds chosen from the biocompaptible and biodegradable (co) polymers presented above and such as they exhibit suitable solubilities, - phospholipids such as in particular phosphatidylcholine (PC), phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), dipalmitoyl-phosphatidylcholine (DPPC), dioleylphophatidylethanolamine (DOPE), dioleyl-phosphatidylcholine (DOPC), dimyristoyl-phosphatidylglycerol (DMPG), esters fatty acids such as isopropyl myristate, ethyl palmitate, biodegradable or bioerodible polymers soluble in a supercritical fluid or a mixture of at least two compounds chosen from phospholipids and fatty acid esters shown below above and such as they exhibit suitable solubilities. Said active principle may be in the form of a liquid, a solid powder or an inert porous solid particle comprising on its surface an active principle.

The active principles used are chosen from a wide variety of therapeutic and prophylactic compounds. They are more particularly chosen from proteins and peptides such as insulin, calcitonin, analogues of the LH-RH hormone, polysaccharides such as heparin, anti-asthmatics such as budesonide, beclometasone dipropionate and its active metabolite beclometasone 17-monopropionate, beta-estradiol hormones, testosterone, bronchodilators such as albuterol, cytotoxic agents, corticosteroids, antigens, DNA fragments ... The examples which follow illustrate the invention without limiting its scope. <U> Example 1 </U> This example illustrates the first method of implementing the invention.

80 mg of PLGA are dissolved in 80 ml of ethyl acetate. 400 mg of micronized insulin is suspended in the solution thus obtained and the suspension is placed in an autoclave with a capacity of 1.5 I. In a first step, the pressure is increased to 100 105 Pa by introducing the liquid CO 2 while remaining at a constant temperature of 28 C. The CO 2 in the liquid state mixes with the suspension thus making it possible to wet the insulin, and also making it possible to ensure the gradual precipitation of the coating agent.

The CO 2 is passed to the supercritical state by gradually increasing the pressure to 200 105 Pa. The temperature is jointly maintained at 40 ° C. Thus the ethyl acetate is extracted. These conditions are maintained for 15 minutes, then the CO 2 / ethyl acetate mixture is discharged by decompressing to 75 105 Pa in a separator. The ethyl acetate is recovered in this separator and the CO 2 in the supercritical state returns to a tank.

The ethyl acetate is recovered and the successive cycles of introduction of liquid CO 2, of passing to the supercritical state and of evacuation of CO 2 + ethyl acetate are repeated until complete elimination of the acetate. ethyl.

The decompression is necessarily carried out by the gas phase so as not to reconcentrate the coating agent in the remaining ethyl acetate. After the decompression phase, the operation can be repeated several times by reintroducing C02 in order to find a pressure of 200 105 Pa and a temperature of 40 C. Finally, the depressurization is carried out and the C02 + solvent mixture is extracted then fresh C02 is reintroduced which is brought to the supercritical state in order to completely extract the solvent. The temperature in this case is generally between 35 and 45 C and the pressure between 180 and 220 105 Pa. 460 mg of non-aggregated microparticles with an average size of 3 μm and comprising 87% by weight of insulin are thus obtained, which exhibit improved nebulization properties. <B> <U> Example 2 </U> </B> This example illustrates the second method of implementing the invention.

1.3 g of dipalmitoyl-phosphatidylcholine (DPPC) are placed in a sealed bag formed from porous filter paper (porosity 2 μm), said bag then being attached to the shaft of the agitator placed in an autoclave of capacity 1, 5 I.

Then 3.0 g of beclometasone dipropionate is added in the form of a free powder prepared by atomization. The autoclave is sealed, the mixture is stirred at 430 rpm and then the interior of the autoclave is pressurized by adding CO 2. When the autoclave is pressurized, the temperature of the autoclave is increased to 50 C. The pressure of the autoclave is thus 220 105 Pa. The CO 2 is then in supercritical form.

The system is then allowed to equilibrate for one hour.

The coating agent initially inside the bag thus dissolves in the supercritical CO 2 and forms a homogeneous solution in the autoclave. The temperature of the autoclave is then slowly reduced to 27 C at linear speed for a period of 38 minutes starting from 50 C. The phase in suspension in supercritical CO 2 is thus transformed into a mixture of liquid and gaseous CO 2, the particles of active principle being in suspension in the liquid CO2. By then depressurizing to atmospheric pressure, particles of beclometasone dipropionate coated with a uniform layer of DPPC are obtained. 3.7 g of non-aggregated beclometasone dipropionate microparticles with a mean diameter of 5 μm coated with a continuous layer of DPPC are thus obtained, which exhibit improved nebulization properties.

Claims (1)

<B><U>REVENDICATIONS</U> </B> <B> 1. </B> Biocompatible microparticle intended to be inhaled comprising at least one active principle and at least one layer coating this active principle which is the outer layer of said microparticle, said outer layer containing at least one coating agent, characterized in that said microparticle is uniformly coated, has an average diameter of between 1 μm and 30 Nm, an apparent density of between 0.02 g / cm3 and 0, 8 g / cm3 and that it can be obtained according to a process comprising the essential steps which are the placing in the presence of a coating agent with an active principle and the introduction of a supercritical fluid. <B> 2. </B> Microparticles according to claim 1, characterized in that they have an average diameter of between 2 μm and 15 Nm, and even more preferably between 3 Nm and 8 Nm and a bulk density of between between 0.05 glcm3 and 0.4 glcm3. 3. Microparticle according to claim 1 or 2 capable of being obtained by a process comprising the following steps - suspending an active principle in a solution of at least one substantially polar coating agent in an organic solvent, said active principle being insoluble. in the organic solvent, said substantially polar coating agent being insoluble in a fluid in the supercritical state, said organic solvent being soluble in a fluid in the supercritical state, - bringing the suspension into contact with a fluid in the supercritical state, so as to desolvate in a controlled manner the substantially polar coating agent and ensure its coacervation, - substantially extracting the solvent by means of a fluid in the supercritical state and removing the SC fluid / solvent mixture, - recovering the microparticles. <B> 4. </B> Microparticle according to claim 1 or 2, obtainable by a process which consists in placing an active principle in suspension in a supercritical fluid containing at least one coating agent dissolved therein then to ensure the coacervation of the particles, by physicochemical modification of the medium. <B> 5. </B> Microparticle according to claim 3, characterized in that the coating agent is chosen from the group formed by biodegradable (co) polymers of α-hydroxycarboxylic acids, in particular homopolymers and copolymers of lactic acids and glycolics, and more particularly PLA (Poly-L-lactide) and PLGA (Poly-Lactic-co-Glycolic-Acid), biocompatible polymers of polyethylene glycol type, polyethylene oxides, block copolymers of polyoxide type ethylene, propylene polyoxides and polysaccharides. 6. Microparticle according to claim 4, characterized in that the coating agent is chosen from the group formed by phospholipids such as in particular phosphatidylcholine (PC), phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), dipalmitoyl-phosphatidylcholine (DPPC), dioleyl-phophatidylethanolamine (DOPE), dioleyl-phosphatidylcholine (DOPC), dimyristoyl-phosphatidylglycerol (DMPG), fatty acid esters such as isopropyl myristate, ethyl palmitate, polymers biodegradable or bioerodible soluble in a supercritical fluid. 7. Microparticle according to one of claims 1 to 6, characterized in that the active principle is chosen from the group formed by proteins and peptides such as insulin, calcitonin, analogues of the LH-RH hormone. , polysaccharides such as heparin, anti-asthmatics such as budesonide, beclometasone dipropionate and its active metabolite beclometasone 17-monopropionate, beta-estradiol hormones, testosterone, bronchodilators such as albuterol, cytotoxic agents, corticosteroids, antigens, fragments of A. D. N. 8. Microparticle according to one of claims 1 to 7, characterized in that the weight ratio of active principle / coating agent of this particle is between 95I5 and 5I95. 9. Microparticle according to claim 7, characterized in that the microparticle is for immediate release and that the active principle / coating agent mass ratio of this particle is between 95I5 and <B> 80120. </B> 10. Process for the preparation of microparticles intended to be inhaled and comprising the following steps - suspending an active principle in a solution of at least one substantially polar coating agent in an organic solvent, said active principle being insoluble in the organic solvent, said substantially polar coating agent being insoluble in a fluid in the supercritical state, said organic solvent being soluble in a fluid in the supercritical state, - bringing the suspension into contact with a fluid in the supercritical state, so as to desolvate the coating agent in a controlled manner substantially polar and ensure its coacervation, - substantially extract the solvent by means of a fluid in the supercritical state and evacuate r the SC fluid / solvent mixture, - recover the microparticles. <B> 11. </B> Process for preparing microparticles intended to be inhaled, which consists in suspending an active principle in a supercritical fluid containing at least one coating agent dissolved therein and then ensuring the coacervation of the particles, by physico-chemical modification of the medium.