LIQUID CRYSTAL FORMS OF CYCLOSPORIN

Abstract: This invention relates to novel, liquid crystal forms of the cyclic peptide cyclosporin and to novel powder formulations of cyclosporin prepared using this novel liquid crystal form of the drug. Methods for preparing and using these formulations are also provided. In particular, the present invention relates to dispersible spray dried particles of cyclosporin suitable for pulmonary delivery.

Description: LIQUID CRYSTAL FORMS OF CYCLOSPORIN FIELD OF THE INVENTION This invention relates to a liquid crystal form of the cyclic peptide cyclospoπn and to powder formulations of cyclospoπn prepared using this novel liquid crystal form of the drug. Methods for prepaπng and using these formulations are also provided. In particular, the present invention relates to dispersible spray dπed particles of cyclospoπn suitable tor pulmonary delivery BACKGROUND OF THE INVENTION The cyclospoπns are a group of non-polar ohgopeptides with immunosuppressant activity. Cyclospoπn A. also known as cyclospoπne. is the major known cyclospoπn. with the structure of cyclospoπns B through I also being known (The Merck Index. Twelfth Edition. 464-465 (1996)). A number of synthetic cyclospoπn analogs have been prepared. (Id.) Cyclospoπn A is an orally active lmmunosuppressive drug that has been used for immune suppression since the mιd-1980's (Guzman et al.. J. of Pharm Sci. 82:5) 496-506 (1993)). It has become the mainstay of organ transplant therapy as prophylaxis against organ rejection. The oπginal cyclosporme product for this use. Sandimmune by Sandoz. is formulated in corn oil and designed for oral delivery, however, bioavailabihty from the gastrointestinal tract tends to be low and somewhat erratic. (Id.) Recently. Sandoz has begun marketing an improved proprietary oral formulation (Neoral) that is claimed to be more reliable than the oπginal (Med. Ad. News, Feb 1996, 7-10). Cyclospoπn A causes kidney and liver toxicity at high doses when delivered orally and tolerabihty must always be monitored along with clinical assessments of rejection (Physician ' s Desk Reference. 52 nd Ed. 1891-1901 ( 1998)) 1 In order to avoid the complications associated with oral delivery, and. in particular, to prevent lung transplant rejection, it may be desirable to deliver cyclospoπn directly to the lungs. In fact, nebulized cyclospoπn A appears to be efficacious in preventing lung transplant rejection using aerosolized liquid ethanol and polyethylene glycol cyclospoπne formulations (Burckart. et al.. Inhalation Delivery of Therapeutic Peptides and Proteins. Marcel Dekker, NY, pp 281-299 ( 1997)). Nebulized cyclspoπn A also appears to lower oral cortico-steroid dependency in asthma (Morley, et al., Ciclosponn Form for Pulmonary Administration, European Patent Application No 92104426.9 (1992)). Liposomal cyclospoπne has also been administered as an aerosol using a nebulizer ( Waldrep, et al., Cyclosporin A Liposome Aerosol: Particle Size and Calculated Respiratorv Deposition, Intl. J Pharm. 97:205- 212 (1993)). The aim of such formulations has been to decrease toxicity compared to conventional oral formulations and to provide an alternative to nebulized solutions containing cosol vents. Nebulized solution delivery of cyclospoπn suffers from limited drug solubility in aqueous based vehicles. Further, there are safety concerns surrounding nebuhzation of organic vehicles Delivery of nebulized solutions and suspensions both suffer from low drug delivery efficiency from commercial nebulizers. Aerosohzation of cyclospoπn A (CsA) with MDI's would involve a solution of CsA in propellant(s) (chlorofluorocarbon or non-chlorofluorocarbon propellants) or the use of finely divided CsA suspended in propellant(s). Poor drug delivery efficiency and low drug-carrying payload capacity make MDI's an inconvenient means of aerosol delivery for human dosing regimens that may require 1 mg to 20 mg of CsA delivered per day to the lung. In view of the difficulty of dehveπng a solution of cyclospoπn by inhalation, it may be desirable to deliver cyclospoπn as a dry powder. The ability to deliver pharmaceutical compositions as dry powders, however, is problematic in certain respects. The dosage of many pharmaceutical compositions is often cπtical, so it is desirable that dry powder delivery systems be able to accurately, precisely and reliably deliver the intended amount of drug. It is also essential that dry powders for pulmonary delivery be readily dispersible in order to assure adequate distπbution and systemic absorption. Because CsA can cause gingivitis, it is important that oropharyngeal deposition be minimized. SUMMARY OF THE INVENTION The present invention provides a novel liquid crystal form of cyclospoπn not previously known. This novel form is thermotropic liquid crystal cyclospoπn It has unexpectedly been found that spray drying organic solutions containing cyclospoπn, in particular cyclospoπn A (CsA), under specific conditions, results in this novel form of cyclospoπn. Spray drying of organic solutions containing cyclospoπn produces powders where the paniculate cyclospoπn exhibits a lack of 3-dιmensιonal (3-d) order as determined by powder X-ray diffraction (PXRD) and also exhibits 2-d order when analyzed by small angle X-ray scatteπng (SAXS). Further, it exhibits a phase change from solid to liquid over a narrow temperature range with a step-wise change in heat capacity, i.e.. a glass transition-like melt. This form of cyclospoπn is liquid crystal cyclospoπn. The process conditions for spray drying may be vaπed within certain limits to achieve very narrow particle size distπbutions that make the powders especially suitable for efficient delivery by oral inhalation. These powders have high delivery efficiency when aerosolized with a dry powder inhaler and have demonstrated physical, chemical, and aerosol stability over prolonged periods of high temperature and high humidity In one aspect the invention provides liquid crystal cyclospoπn. In particular, the liquid crystal form of cyclospoπn A is provided. In another aspect the invention provides dispersible powder formulations of liquid crystal cyclospoπn for pulmonary delivery In particular, cyclospoπn-based 3 dispersible powder formulations which are spray dπed from cyclospoπn and. optionally, excipient. in a solvent are provided, as are methods for making these formulations. Spray dπed cyclospoπn A powders are specifically provided. 5 In a further aspect the invention provides methods for treating a subject suffering from or subject to a condition which may be alleviated or prevented by the administration of cyclospoπn compπsing administeπng the dispersible powder cyclospoπn formulations descπbed above. In particular, methods to alleviate or prevent lung diseases or conditions which affect the lung are provided. Cyclospoπn may be 10 used as an anti-inflammatory, immunosuppressive or anti-asthmatic agent. BRIEF DESCRIPTION OF THE DRAWINGS Figures 1A through 1C illustrate the difference in X-ray powder diffraction 15 patterns between two of the known crystalline forms of cyclospoπne (non-solvated orthorhombic (Figure 1 A) and dihydrate tetragonal (Figure IB)) and the novel thermotropic liquid crystal form of cyclospoπne provided by the present invention (Figure 1C). ->. 0 Figure 2A illustrates a representative open pan differential scanning caloπmetry (DSC) tracing and Figure 2B illustrates a representative closed pan DSC tracing for the thermotropic liquid crystal form of cyclospoπne of the present invention Figure 3 presents the small angle X-ray scatteπng data for orthorhombic and 25 tetragonal crystalline cyclospoπne and for spray dπed liquid crystal cyclospoπne at 10, 80 and 150°C. Figure 4 presents the dielectπc analysis (DEA) of a thermotropic liquid crystal CsA formulation according to the present invention. 30 Figures 5A. 5B and 5C illustrate HPLC analysis of spray dπed cyclospoπn A at accelerated storage conditions of 110°C for 196 hours. 140°C for 50 hours and 210°C for 10 minutes, respectively. Figures 6A. 6B and 6C demonstrate that spray dried cyclospoπn A powder stored for 10 months at 40°C and 75% relative humidity that showed no appreciable degradation based on HPLC analysis. Figure 7 demonstrates that spray dπed cyclospoπn A powders stored at room temperature for 15 months that showed no appreciable degradation based on HPLC analysis. DETAILED DESCRIPTION OF THE INVENTION The present invention is based at least in part on the discovery of a novel, thermotropic liquid crystal form of cyclospoπn, particularly cyclospoπn A This liquid crystal form of cyclospoπn may be formulated as a dispersible powder by spray drying from organic solvents. The cyclospoπn-based compositions are suitable for pulmonary delivery due to their dispersibihty characteπstics The compositions of the invention are readily aerosolized and presented to the deep lung of a host when delivered by a dry powder inhaler. The powder formulations of the present invention retain stability, are readily dispersible for pulmonary delivery and allow for unit dose packaging The invention consists, in part, of compositions compπsing cyclospoπn in dispersible powder formulations. The use of particles of a certain size range allows for delivery of cyclosporin to the alveolar area of the lungs (i.e , to the deep lung) Optionally, the powder formulations of the present invention may contain stabilizers and excipients such as buffer salts, sugars, tonicifiers, preservatives and anti-oxidants The compositions of the present invention are useful in pulmonary dry powder drug delivery systems, including but not limited to those disclosed in U.S. Patent No. 5.458.135 and International Patent Publication WO96/09085. The solid state forms of CsA that have previously been reported are descπbed in Table 1. Table 1. Solid State Forms of Cyclosporin A *- Crystal System Solvation Melting Range Space Group Unit Cell Dimensions Reference (°C) (A) a ) c tetragonal dihydrate 140- 150 P4, 13.837 13.387 41.242 1 ,2 tetragonal non-solvaled III " P4, nr nr n 3 orthorhombic di-(di-isopropyl ether) - 150 P2|2,2, 12.5 22.9 23.4 1 orthorhombic non-solvated 180- 195 P2|2,2, 12.7 15.7 36.3 1 ,3 orthorhombic monohydrate III " P2,2,2, 12.5 22.9 28.4 4 amorphous n/a n/a n/a n/a n/a n/a 3 nr = not reported n/a = not applicable o H O W e References 1 Giron. et al.. Orthorhombic Ciclosporin Crystals, UK Patent Application No. 8829556 3 ( 1988). 2 Loosli. et al.. The Conformation of C\closporιn A in the Crystal and in Solution, Helvetica Chemica Acta, 68:682-704 ( 1985) 3 Matha, et al.. 77ze Story of the Czech Cxclosporin A, 28 pp. 4 Knott. et al., Neutron Structure of the Immunosuppressant Cvclosponn-A, Acta Cryst., 046:1528-1533 ( 1990). Tetragonal and orthorhombic crystal forms of CsA exhibit high melting points and have characteristic sharp diffraction peaks when analyzed by PXRD and are birefπngent to polaπzed light. Amorphous mateπals. unlike liquid crystals, have no peaks by SAXS and are not birefπngent to polaπzed light Liquid crystals show a distinct melt over a narrow temperature range unlike amorphous glasses which show no such melt. An amorphous form of CsA has been mentioned without report of its physicochemical properties (Morley, et al., European Patent Application No. 92104426.9 ( 1992)). We have discovered that CsA powders prepared by organic solvent spray drying can be designed such that they are not of the orthorhombic or tetragonal crystal forms. Indeed, no sharp diffraction peaks indicative of 3-dιmensιonal order are observed by PXRD of the spray dπed powders (Figure 1C) However, the spray dπed powders do exhibit 2-dιmensιonal order when analyzed by SAXS, indicative of a liquid crystal form (Figure 3) Furthermore, the CsA powders prepared by organic solvent spray drying also demonstrated a distinct endotherm between 0-75°C when analyzed by differential scanning caloπmetry (DSC) at 107mιn in an open pan (Figure 2A). Since the physical nature of the powders does not change (not a melt) through that temperature range and because the endotherm is absent in the closed pan DSC trace (Figure 2B). the transition is believed to be due to solvent evaporation. A melt occurs beginning at about 120°C when analyzed by hotstage microscopy at 2°/mιnute. The DSC thermogram at the same heating rate shows a step-wise change in heat capacity (Cp) (a Tg-hke transition) at that temperature. Typically, 3-d ordered crystals do not exhibit a step-wise Cp change going from solid to liquid. Furthermore, dielectric analysis is consistent with DSC results and confirms that the transition observed at ~120°C is a second order transition, as it is demonstrated by the observed frequency dependency A. Definitions As used herein the following terms have the following meanings: The terms "dispersibihty" or "dispersible" mean a dry powder having a moisture and/or residual solvent content of less than about 10% by weight (%w), usually below about 5%w and preferably less than about 3%w and often less than about l%w; a particle size of between 0.1 and 15 μm, often between 0.2 μm and 10 μm, usually about 0.4 to 5 μm mass median diameter (MMD), preferably about 1 to 4 μm MMD and most preferably 1 to 2 μm MMD. a delivered dose of greater than about 30%. usually greater than about 40%, preferably greater than about 50% and most preferably greater than about 60%; and an aerosol particle size distribution of about 1-5 μm mass median aerodynamic diameter (MMAD), usually about 1.5-4.5 μm MMAD and preferably about 1.5-4.0 μm MMAD, or with at least about 40% (preferably at least about 50%) of the particles less than about 3.3 μm in diameter. The term "cyclospoπn" means any of the group of non-polar cyclic ohgopeptides with immunosuppressant activity and includes known cyclospoπns A through I. In particular, this term includes cyclospoπn A, also known as cyclospoπne. Synthetically produced, naturally-deπved or puπfied and recombinantly produced moieties are included, as are analogs, deπvatives. agonists, antagonists and pharmaceutically acceptable salts ot any of these. The term also includes cyclospoπns which have D- amino acids, modified, deπvatized or non-naturally occurπng ammo acids in the D- or L- configuration and or peptomimetic or prodrug units as part of their structure. A thermotropic liquid crystal is a state of matter distinct from the amorphous and 3-dιmensιonal crystalline states and characteπzed by the existence of long range order in one (nematic) or two (smectic) dimensions in the absence of solvent. An amorphous phase lacks long-range order and a 3-dιmensιonal crystalline phase contains 3- dimensional long-range order. The term "powder" means a composition that consists of finely dispersed solid particles that are free flowing and capable of being readily dispersed in an inhalation device and subsequently inhaled by a subject so that the particles reach the spaces of the deep lung to permit deposition in the alveoli. Thus, the powder is said to be "respirable." The terms "pharmaceutical excipient" or "additive" mean compounds which stabilize cyclospoπn and/or improve powder aerosol performance and stability The types of excipients useful in the present invention include buffer salts, sugars, tonicifters, preservatives and anti-oxidants, and the like. The term "physically stable" or physical stability" intends a composition that does not show a change in phase over time. The term "chemically stable" or "chemical stability" means that the composition shows less than 10% and, preferably, less than 5% total degradation in 2 years at room temperature at the storage conditions specified. "Aerosol stability" means that the aerosol composition shows no statistical change in delivered dose efficiency with time. 10 The term ''subject" includes any human or animal species in need of cyclospoπn for treatment or prophylaxis of conditions for which pulmonary delivery of cyclospoπn would be efficacious. B. Compositions- The present invention is drawn to liquid crystal forms of cyclospoπn and to dispersible cyclospoπn-containing powder compositions suitable for pulmonary delivery formed by spray drying from organic solvents. The dispersible powder compositions compπse a therapeutically effective amount of cyclospoπn. optionally in combination with a pharmaceutically acceptable earner or excipient. Spray drying is a process that utilizes high temperatures and high-earner gas flow rates to rapidly evaporate solvents from an atomized solution that contains dissolved solutes. The solvent evaporates to leave a solid particle. The size of the particle is dependent upon the conditions of the spray drying process (e.g., solids content of solution, pressure of atomization gas, design of atomizer nozzle and design of cyclone collector). Control of particle size and particle size distπbution is important for efficient inhalation delivery to the airways and the deep lung. The mass median diameter (MMD) of the particles should preferably be between 1 and 2 μm, with 100% of the particles less than 15 μm. It can also be difficult to control particle size and particle size distπbution in compositions produced by spray drying. For pulmonary delivery it is cπtical that the average particle size be maintained in a respirable range and that the amount of the composition compπsing particles outside the target size range be minimized. Moreover, it can sometimes be difficult to achieve a desired low residual solvent and/or moisture content required for physical and chemical stability m the final paniculate product, particularly in an economic manner. Useful methods are disclosed, for example, in 11 International Patent Application No. PCT/US97/07779, the disclosure ot which is incorporated herein by reference in its entirety Cyclospoπn. including cyclospoπn A (CsA) is very hydrophobic and is practically insoluble (<6 μg/mL) in aqueous vehicles. Organic solvents with boiling points less than 200°C, preferably less than 150°C, and most preferably less than 100°C should be used to obtain powders with low residual solvent. The preferred solvents for making spray dπed cyclospoπn (including CsA) powders include, but are not limited to, ethanol, acetone, acetomtπle. methanol, isopropanol. and methylene chloπde. either alone or in combination or in cosolvent systems. Pharmaceutically acceptable protic solvents with low dielectric constants are more preferred (e.g., ethanol is preferred over methanol) Solvent mixtures with less than 50%, preferably less than 25%, and most preferably with 10% or less water by volume may also be employed for spray drying cyclospoπn. Use of water in the solvent mixture allows the incorporation of water- soluble excipients into the CsA particles, however non-aqueous systems are preferred. Water-soluble excipients useful in the present invention include, but are not limited to, buffer salts (e.g., citnc acid/sodium citrate), natural and synthetic sugars as bulking agents (e.g., lactose, mannitol), tonicifiers (e.g., sodium chloride), and preservatives and anti- oxidants (e.g., ascorbic acid/sodium ascorbate). Pharmaceutical excipients and/or additives generally useful in the present invention include suitable pH adjusters or buffers such as organic salts prepared from organic acids and bases, such as sodium citrate, glycine, sodium tartrate. sodium lactate, tromethamine and the like. Proteins (e.g., HSA. recombinant human albumin (rHA), gelatin and casein), peptides (e.g., aspartame) and ammo acids (e.g., alanine, glycine. arginme, glutamic acid and aspartic acid) which improve dispersibihty of the powder may be useful. Carbohydrates/sugars and alditols are also useful. Suitable carbohydrate/sugar 12 compounds include sucrose, trehalose, lactose, raffinose, and the like. Suitable alditols include mannitol and pyranosyl sorbitol and the like. Polymeπc excipients/additives include polyvinylpyrrohdones (PVP), Ficolls, soluble hydroxy ethyl starch, dextrates and the like of high molecular weight. Also useful are small amounts of pharmaceutically acceptable surfactants such as Tweens, chelators such as EDTA and inorganic acids and bases such as sodium phosphate and the like. Other suitable pharmaceutical excipients and/or additives include those disclosed in Remington, Pharmaceutical Sciences 18th ed. (1990), the disclosure of which is incorporated herein by reference. The temperatures employed for drying the atomized solution droplets may range from 20 to 300°C, preferably 30 to 150°C, and most preferably 40 to 120°C. These temperatures are expressed as the outlet temperature of the earner gas. Specifically, the outlet temperature is the temperature of the gas at the outlet of the drying chamber pπor to entry into the cyclone and collector. Correspondingly higher temperatures are required at the point of atomization to achieve the recommended outlet temperatures. The spray drying process may include maintaining the powder an additional peπod of time at a given temperature after the completion of cyclospoπn solution feed through the system (i.e., secondary drying). This secondary drying may be used to reduce any residual solvent left in the powder. A droplet size of about 4 to about 8 μm diameter is preferred in order to achieve optimal powder characteπstics. Such a droplet size may be achieved, for example, by using the atomization method descπbed in International Patent Application No PCT/US97/07779. Unless otherwise specified, atomization methods that result in droplet sizes of 4-8 μm were used in the examples that follow. The mass median diameter (MMD) of the powders prepared by organic spray drying were measured by centπfugal sedimentation with a Hoπba CAPA-700 Particle 13 Size Analyzer A powder sample was dispersed in a vehicle of Sedisperse W-l 1 (Micromeπtics. Norcross. GA) which was pre-saturated with cyclospoπn A and filtered pπor to the addition of the powder sample. The particle size ranged from about 0.7 to about 2.4 MMD. The size of the CsA particles was confirmed by scanning electron microscopy; the particles were also found to be generally spheπcal in shape, i.e., from smooth spheres to dimpled, raisin- ke or wπnkled. The aerosol performance charactenstics of the powders were evaluated using the Inhale Therapeutic System's aerosol device. The device includes an aerosol chamber and employs a volume of compressed air to disperse the powder from an aluminum foil blister package. The delivered dose efficiency (DDE) for each powder was defined as the percentage of the nominal dose contained within a blister package that exited the mouthpiece of the aerosol device and was captured on a filter through which a vacuum was drawn (30 L/min) for 2.5 seconds following device actuation. The filter was weighed before and after actuation of the device to determine the mass of powder delivered past the mouthpiece. The particle size distπbution of the aerosolized powders was determined using an Andersen cascade impactor through which a vacuum (28.3 L/min) was pulled for 2 5 seconds. The residual solvent and/or moisture content of the powder particles of the present invention is usually below about 10% by weight, preferably below about 5%w and more preferably below about 3%w Such low solvent and/or moisture content powders are generally physically and chemically stable duπng storage at room temperature and are readily dispersible in an inhalation device to form an aerosol Stability studies of spray dned cyclospoπn formulations of the present invention were performed, and showed that these compositions retained aerosol and physical stability. In particular, the DDE of a cyclosporin powder spray dπed from ethanol at 14 70°C without secondary drying was measured immediately after preparation and found to be 48.4%. The powder was then stored at room temperature for 10 months. The DDE was again measured and found to be 49.5%. indicating that the powder retained aerosol stability. In another stability test, the DDE of cyclospoπn powders spray dned from ethanol at 70°C without secondary drying was measured immediately after preparation and found to be about 72% The powder was then stored at the accelerated conditions of 40°C and 75% relative humidity (RH). DDE was measured after 8 weeks and again after 15 weeks of storage under these conditions Results showed that the DDE remained approximately the same, i.e.. about 75% at 8 weeks and about 74% at 15 weeks. A DSC scan of this powder formulation done immediately after preparation showed a melt at 118.61°C, while a DSC scan of the formulation done after 15 weeks at 40°C and 75% relative humidity showed a melt at 119.00°C. These results indicate that no physical change of the powder occurred over this peπod and the powder retained aerosol stability. The amount of cyclospoπn which constitutes a therapeutically effective amount will vary in the composition depending on the biological activity of the cyclospoπn employed and the amount needed in the unit dosage form. The condition to be treated or prevented will also determine the amount of cyclosponn required, as will the subject to which the composition is being administered. The compositions compπse at least about 40% by weight cyclospoπn in the formulation, preferably between about 70% to about 100% and most preferably about 90% to about 100%. The amount of excipients and pharmaceutically acceptable additives may be from about 0-60%, preferably from about 0-30% and most preferably from about 0-10% by weight. The compositions of the present invention will often be packaged as unit doses where a therapeutically effective amount of the cyclospoπn composition is present in a 15 unit dose receptacle, such as a blister pack, gelatin capsule, or the like, so long as a moisture barrier is provided. The cyclospoπn-based dry powder compositions of the present invention may be produced by spray drying solutions or slurπes of the cyclosponn and, optionally, excipients, in a non-aqueous solvent under conditions to provide a respirable dry powder. Solvents may include ethanol. acetone, acetomtπle, methanol and isopropanol, which may be readily dried. Further, the cyclospoπn-based dry powder compositions may also be produced by evaporative drying, freeze-drying, quench from a melt, precipitation including super-critical fluid precipitation. C. Characteπzation: It was found that, by spray drying cyclosporin from organic solvents, a thermotropic liquid crystal form of cyclosponn is formed. In particular, charactenzation of this cyclosponn form using polanzed light microscopy, showed that it was birefπngent, indicating that it was a non-amorphous form of cyclospoπn. Similarly, SAXS analysis showed the presence of sharp peaks (Figure 3), a charactenstic of non- amorphous matenals. Further charactenzation of this novel form of cyclospoπn by powder X-ray diffraction disclosed no sharp diffraction peaks which would have indicated a 3- dimensional order such as that found in crystalline structures, indicating that this was not a 3-dimensιonal crystalline form of cyclospoπn. Figures 1A through 1C show X-ray powder diffraction patterns of two crystalline forms of cyclosponn (tetragonal and orthorhombic) and our novel spray dried form. Hotstage microscopy, DSC, DEA, and SAXS were used to charactenze the novel form of cyclosporin. Hotstage microscopy of this material showed a distinct melting 16 point which is charactenstic of both crystalline and liquid crystalline matenals. DSC showed a step-wise heat capacity change at the melt temperature, charactenstic of liquid crystalline matenals. Figure 2A shows a DSC (heating rate at 10°C/mιn) of the liquid crystal cyclosponn of the present invention, indicating a melting point beginning at 122°C and which can be between 115°C and 125°C When analyzed by DEA, such transition was found to be frequency dependent, suggesting it is indeed a second order transition and not a true melt. The liquid crystal state was confirmed by SAXS. which showed sharp diffraction peaks at low diffraction angles, as charactenstic of 2- dimensional order in liquid crystals Figure 3 shows SAXS for spray dned cyclospoπn The material remains a liquid crystal below the melt (at 10°C and 80°C) and above the melt (at 150°C). D. Pulmonary Cyclosponn Pulmonary cyclosponn is useful for the treatment of asthma and lung transplants but has the potential for use in many other indications as well Pulmonary cyclosponn may be useful to treat sarcoidosis. Obhterative bronchiohtis (OB), the pulmonary pathology that occurs in lung transplant rejection, also occurs in heart and bone marrow rejection, thus there is the potential for inhaled cyclosponn to be of use in other transplant therapies in conjunction with oral immunosuppressants Chronic inflammatory lung disease, chronic obstructive pulmonary disease, emphysema, pnmary and secondary pulmonary hypertension, cystic fibrosis. lung infections or idiopathic pulmonary fibrosis (IPF) are other pulmonary diseases that may respond to inhaled cyclosponn, since they appear to be caused or exacerbated by an overly reactive immune system. Further, cyclosponn may be useful to treat pulmonary complications associated with autoimmune diseases such as rheumatoid arthritis. The advantage of pulmonary delivery of cyclospoπn for lung disease or conditions which affect the lungs is that the total body burden of drug can be reduced, which reduces or eliminates systemic side effects 17 According to the current invention, cyclospoπn may be delivered directly to the deep lung in dry powder form using a dry powder delivery device. A significant requirement for such dry powder delivery devices is efficiency. The delivered dose must be relatively high to reduce the number of breaths required to achieve a total dosage. The ability to achieve adequate dispersion is a significant technical challenge that requires in part that each unit dosage of the powder composition be readily and reliably dispersible. Certain pulmonary delivery devices, such as those disclosed in U.S. Patent No. 5,458,135 and International Patent Publication WO96/09085 (the disclosures of which are incorporated herein by reference) are useful for pulmonary delivery of dry powder drugs. The disclosure of each of the publications, patents or patent applications herein mentioned is hereby incorporated by reference in its entirety to the same extent as if the language of each individual publication, patent and patent application was specifically and individually incorporated by reference. DISCLOSURE OF THE EXAMPLES OF THE INVENTION The following examples are not intended to limit the scope of the invention in any manner. Mateπals and Methods: In general the following mateπals and methods were used in the examples that follow unless otherwise indicated. Matenals: Cyciosponn A, GMP grade, was obtained as a powder crystallized from acetone (melting point 148-150°C) from Poll Industna Chimica, S.p.A. 18 Sample Storage: Spray dπed powders were stored under dry atmosphere (RH <5%). Physical Methods: Powder particle size distπbution The particle size distπbution (PSD) of the spray dπed powder samples was measured with a Hoπba CAPA-700 centnfugal sedimentation particle size analyzer. A powder sample was dispersed in a vehicle of Sedisperse W- 11 (Micromeπtics, Norcross, GA) which was pre-saturated with cyclospoπn A and filtered pnor to the addition of the powder sample. Approximately 5 mg of powder was suspended in about 5 ml of the Sedisperse and sonicated 5-10 minutes in a Lab Supplies ultra-sonicator before analysis. The instrument was configured to measure a particle size range of 0.4 to 10 μm in diameter using a particle density of about 1 2 g/cm\ Powder X-ray diffraction (PXRD) The PXRD was performed on a Siemens D-500 X-ray Diffractor. The sample was measured at 3 mιn (0.8 sec/0.04° step) and 0.57step, 1 sec/step respectively The scan was run continuously from 2° to 40° in 2Θ. Small angle X-ray scatteπng (SAXS) SAXS analysis was performed on a Rigaku 12KW diffractometer equipped with a Kratky camera and a Braun 10-cm position sensitive detector, using a copper X-ray source of 1.542 A at a scanning rate of 0.127mιn in the range of 0 to 2.2° in 2Θ. 19 Method to test birefπngency Birefringence was tested using a polaπzed light microscope. Nikon Optiphot 2- Pol. equipped with a Hamamatsu camera and controller C2400. Pictures were pnnted on a graphic printer UP890MD. The powders, either dry or immersed in Sedisperse W-11 or water, were examined under 20X, 40X and 60X objectives, and photomicrographs were taken through normal and cross polanzed light. Method for hotstage microscopy Hotstage microscopy was performed on a Nikon Optiphot 2-Pol equipped with a Mettler Toledo FP82HT Hotstage and a Hamamatsu camera and controller C2400. Photomicrographs were pnnted on a graphic pπnter UP890MD. The slide was placed in the hotstage and a representative field was found using the 40X objective. Samples were heated at a rate of 2°C/mιn from room temperature to past the melting point. Pictures were taken when changes were visuallv observed. Method for TGA-Residual solvent Samples were analyzed by Oneida Research Services, Inc. The TGA was performed on a Omnitherm 1500. Samples were heated from 30-200°C/mιn under a nitrogen atmosphere with a flow rate of 30 ml/min. The weight loss due to drying was measured and is presented as % weight lost. Method for TGA-Decomposition Samples were analyzed by Oneida Research Services, Inc. The TGA was performed on a Omnitherm 1500 TGA. Samples were heated from 40-600°C at 107mιn under a nitrogen atmosphere with a flow rate of 30 ml/min. 20 Differential scanning calonmetry (DSC) The DSC scans were performed on a TA Instrument model 2920 Modulated DSC with a TA refπgerated cooling system (RCS) unit, and pure helium gas with a flow rate of ~120 cm mιn. The cell flow rate was set at about 40 cm Vmin. The scans were performed at 10°C/mιn non-modulated, with an equilibration temperature of -30°C for 15-30 min, followed by heating to about 200-225°C Open and closed aluminum pans were filled with about 2 mg-6 mg of powder (Figures 2A and 2B, respectively) Dielectπc analysis (DEA) The DEA scan was performed on a TA Instrument 2970 Dielectnc Analyzer using liquid nitrogen to cool the sample to the initial temperature. The powder (40-45 mg) was pressed into a pellet. Vτ inch in diameter and approximately 0 3 mm thick, using a Carver press for 40 seconds at 1 ton. Two thin layers of Teflon 25 μm and 7/16 inch in diameter, were inserted into the pellet die to alleviate adhesion to the die faces. Duπng the measurement, the pellet was surrounded by a silicone gasket, ID 9/16 inch, OD 1 1/16 inches thickness, 0.35 mm. to help maintain thickness dunng the measurement The pellet was also sandwiched between two layers of Teflon, 25/xm thick, to remove possible contnbutions from ionic conductivity Electrodes were fabπcated from TA Instruments ' sensors for sputter coated samples and gold foil, 25μm thick, one disk 7/16 inch in diameter and the other 6/8 inch in diameter, so that only the sample contacted the foil electrodes. Expenments were performed using a heating rate of 2°C/mιn from -40 to 200°C and using the following frequencies 1 Hz, 10 Hz, 100 Hz, 1,000 Hz. 10,000 Hz, 100,000 Hz. The permitivities measured with this method were in arbitrary units since the data was not adjusted for the gold foil sensors; however, this does not impact the interpretation of the results (Figure 4) 21 Aerosol Methods: Delivered dose assay The delivered dose assay was performed to determine the efficiency and reproducibi ty of pulmonary delivery of the dispersible powder cyclospoπn compositions. The aerosol performance characteπstics of the powders were evaluated using the Inhale Therapeutic System's aerosol device, similar to devices disclosed in WO96/09085. The device includes an aerosol chamber and employs a volume of compressed air to disperse the powder from an aluminum foil blister package. The delivered dose efficiency (DDE) for each powder was defined as the percentage of the nominal dose contained within a blister package that exited the mouthpiece of the aerosol device and was captured on a glass fiber filter (Gelman, 47 mm diameter) through which a vacuum was drawn (30 L/min) for 2 5 seconds following device actuation. Delivered dose efficiency was calculated by dividing the mass of the powder collected on the filter by the mass of the powder in the blister pack. Each result was the average of 5-10 replicate measurements. Aerosol particle size distπbution The aerosol particle size distnbution was obtained using an eight stage cascade impactor (Graseby Andersen. Smyrna. Georgia). The impactor air-flow was set to pull a vacuum of 28.3 L/min. the calibrated flow-rate for the instrument, for 2.5 seconds. For each run. the blister packs filled with approximately 5 mg of powder were dispersed from the inhaler. The particle size was determined by weighing the powder on the impactor plates and evaluating the results on a log-probability plot. Both the mass median aerodynamic diameter (MMAD) and the mass fraction less than 5 μm were determined from the log-probability plot " >? Chemical Stability Method: The stability-indicating HPLC method that was used is descπbed in Ohyai, et al.. Kinetics of Acid-Catalvzed Degradation of Cxclosporin A and its Analogs in Aqueous Solutions, Peptide and Protein Res. 43:239-247 ( 1994). The method was run according to that descnbed, however the mobile phase ratio was adjusted slightly to obtain the retention times for cyclosponn A and iso-cyclospoπn A. Example 1: Cyclosporin A Spray Dried From Ethanol At 70°C With No Secondary Drying Solution Preparation 1.5 g of cyclosponn A was dissolved in 50 mL of ethanol (200 proof, USP, NF grade from Spectrum). Spray Drying A dry powder compnsed of cyclospoπn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer using a nitrogen atmosphere containing less than 5% oxygen (with N 2 atm <5% ON with the following parameters: Outlet Temperature 70°C Inlet Temperature 100°C Feed Rate 5 mL/min Atomizer Flow Rate 13 t/min Secondary Drying None Powder Charactenzation PXRD of the powder showed that it exhibited a double halo with two maxima at 2Θ equal to 8.5° and 17° (Figure IC). The absence of sharp peaks in the diffractogram show that the matenal is not a 3-dιmensιonal crystal. Polaπzed light microscopy showed 23 the particles to be birefπngent. The SEM images appear to be rounded and highly wrinkled particles. There was no solid state change in the formulation due to the increase in relative humidity as shown by the DSC's of the powders exposed to 0% relative humidity and 75% relative humidity. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 69°C. The Tg-like endotherm, which is a melt, appeared on the scan at 122°C onset temperature. Hotstage microscopy showed the melt to be in the range of 138-140°C. The MMD of the powder sample was determined to be 1.6 μm. with 96.5% less than 5.2 μm. The DEA showed a frequency-dependent change in the permitivity indicative of a second order transition, in the same temperature range as determined by DSC. where the change in heat capacity started around 125°C. At the temperature range that the large endotherm was seen in the DSC scan (~20-70°C), there was no change in the permitivity, suggesting that the endotherm is not due to a phase change but rather to solvent evaporation. Aerosol Charactenzation The delivered dose efficiency (DDE) of the above spray dned cyclospoπn A powder was determined to be 79%±4.2% (n=10). The mass median aerodynamic diameter (MMAD) was determined to be 2.81 μm. with 85% less than 5 μm. Chemical Stability HPLC analysis showed degradation products of the spray dπed cyclospoπn A under stressed conditions. Figure 5A shows a sample stored at 110°C for 196 hours, Figure 5B shows a sample stored at 140°C for 50 hours and Figure 5C shows a sample stored at 210°C for 10 minutes. The degradation products for the liquid crystal cyclosponn are different than for other reported forms of cyclosponn. (Ohyai, et al., Kinetics of Acid-Catalyzed Degradation ofCxclosporin A and its Analogs in Aqueous 24 Solution, Peptide and Protein Res. 43 239-247 ( 1994) and Ohyai, et al.. Kinetics and Mechanism of Isomenzation of Cyclosponn A, Pharm Res. 9(5):617-622 (1992)) Example 2: Cyclosporin A Spray Dried From Acetone At 88°C With Secondary Drying Solution Preparation 1.5 g of cyclosponn A was dissolved in 50 mL of acetone (HPLC grade from J T Baker) Spray Drying A dry powder compnsed of cyclospoπn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% 0 2 with the following parameters: Outlet Temperature 88°C Inlet Temperature 118°C Feed Rate 5 mL/min Atomizer Flow Rate 13 t/min Secondary Drying 85°C/5 mm Once the solution was consumed, the outlet temperature was maintained at 85°C for 5 mm by slowly decreasing the inlet temperature to provide secondary drying Powder Charactenzation PXRD of the powder showed that it exhibited a double halo with two maxima at 2Θ equal to 8.5° and 17° Polanzed light microscopy showed the particles to be birefπngent. TGA analysis of the powder showed the powder to have 0.1% by weight of residual solvent and a decomposition temperature range of 347-421°C. The SEM image of the powder showed the particles to be rounded, with slight dimples. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 58°C. The Tg-like endotherm, which is a melt, appeared on the scan at 121°C onset temperature. The MMD of the powder sample was determined to be 1.19 μm, with 95.8% less than 5.2 μm. Aerosol Charactenzation The DDE of the above spray dned cyclosponn A powder was determined to be 59%±9% (n=10). The MMAD was determined to be 2.0 μm with 84% less than 5 μm. Example 3: Cyclosporin A Spray Dried From Ethanol At 85°C With Secondary Drying Solution Preparation 1.5 g of cyclosponn A was dissolved in 50 mL of ethanol (200 proof, USP, NF grade from Spectrum). Spray Drying A dry powder compnsed of cyclospoπn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% 0 2 with the following parameters: Outlet Temperature 85°C Inlet Temperature 120°C Feed Rate 5 mL/min Atomizer Flow Rate 13 t min Secondary Drying 85°C/5 mm Once the solution was consumed, the outlet temperature was maintained at 85°C for 5 mm by slowly decreasing the inlet temperature to provide secondary drying. 26 Powder Charactenzation PXRD of the powder showed that it exhibited a double halo with two maxima at 2Θ equal to 8.5° and 17° Polaπzed light microscopy showed the particles to be birefπngent. TGA analysis of the powder showed the powder to have 0 3% by weight of residual solvent and a decomposition temperature range of 348-425°C. The SEM image of the powder showed the particles were raisin-like. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 62°C. The Tg-like endotherm. which is a melt, appeared on the scan at 122°C onset temperature. The MMD of the powder sample was determined to be 1 27 μm. with 100% less than 5 2 μm Aerosol Charactenzation The DDE of the above spray dned cyclospoπn A powder was determined to be 71.4%±6% (n=10). The MMAD was determined to be 2.8 μm with 86% less than 5 μm. Chemical Stability HPLC analysis showed no appreciable degradation from samples of the spray dned cyclosponn A powder stored at 40°C and 75% relative humidity tor 10 months (see Figures 6A, 6B and 6C) The powder is considered to be chemically stable over the time course of the study. Figure 6 A is a chromatogram of the mobile phase. Figure 6B is a chromatogram of the bulk cyclosponn A m the mobile phase with a 25 μg load Figure 6C is a chromatogram of the spray dπed cyclospoπn A aged at 40°C and 75% relative humidity for 10 months reconstituted in the mobile phase. 27 Example 4: Cyclosporin A Spray Dried From Acetonitrile At 101°C With Secondary Drying Solution Preparation 1.5 g of cyclospoπn A was dissolved in 50 mL of acetonitπle (HPLC grade from Burdick and Jackson). Sprav Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% O2 with the following parameters: Outlet Temperature 101°C Inlet Temperature 141°C Feed Rate 5 mL/min Atomizer Flow Rate 15 lit/min Secondary Drying 100°C/5 min Once the solution was consumed, the outlet temperature was maintained at 100°C for 5 min by slowly decreasing the inlet temperature to provide secondary drying. Powder Charactenzation PXRD of the powder showed that it exhibited a double halo with two maxima at 2Θ equal to 8.5° and 17° Polarized light microscopy showed the particles to be birefnngent. The SEM image of the powder showed the particles to be slightly dimpled. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 69°C. The Tg-like endotherm, which is a melt, appeared on the scan at 121°C onset temperature. The MMD of the powder sample was determined to be 1.99 μm. with 99% less than 5.2 μm. 28 Aerosol Charactenzation The DDE of the above spray dπed cyclospoπn A powder was determined to be 69.9%±7% (n=10). The MMAD was determined to be 1.9 μm with 83% less than 5 μm. Example 5: Cyclosporin A Spray Dried From Acetone At 102-103°C With Secondary Drying Solution Preparation 1.5 g of cyclosponn A was dissolved in 50 mL of acetone (HPLC grade from J.T. Baker). Sprav Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% O2 with the following parameters: Outlet Temperature 102-103°C Inlet Temperature 140°C Feed Rate 5 mL/min Atomizer Flow Rate 15 lit/min Secondary Drying 101°C/5 min Once the solution was consumed, the outlet temperature was maintained at 101°C for 5 min by slowly decreasing the inlet temperature to provide secondary drying. Powder Charactenzation Polanzed light microscopy showed the particles to be birefnngent. The SEM image of the powder showed that the particles looked round. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 69°C. The Tg-like endotherm, which is a melt, appeared on the scan at 123°C onset temperature. 29 The MMD of the powder sample was determined to be 1.20 μm with 87.0% <5.2 μm. Aerosol Charactenzation The DDE of the above spray dned cyclosponn A powder was determined to be 63.3%±7% (n=10). The MMAD was determined to be 1.8 μm with 80% less than 5 μm. Example 6: 90% Cyclosporin A:10% Citrate Spray Dried From Ethanol At 85°C With Secondary Drying Solution Preparation 1.35 g of cyclosponn A was dissolved in 50 mL of ethanol (200 proof, USP, NF grade from Spectrum). 150 mg of Sodium Citrate from Sigma Chemicals was dissolved in 2.5 mL of de-ionized water. The ethanol/cyclosponn A solution was added to the Sodium Citrate/water solution and swirled rapidly The resulting suspension was processed in the Spray Dryer. Sprav Drying A dry powder compnsed of cyclospoπn A and sodium citrate (90.10) was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% 0 2 with the following parameters: Outlet Temperature 85°C Inlet Temperature 120°C Feed Rate 5 mL/min Atomizer Flow Rate 13 lit/min Secondary Drying 85°C/5 min 30 Once the solution was consumed, the outlet temperature was maintained at 85°C for 5 min by slowly decreasing the inlet temperature to provide secondary drying. Powder Charactenzation PXRD of the powder showed that it exhibited a double halo with two maxima at 2Θ equal to 8.5° and 17°. The sharp diffraction peaks which stood out above the double halo corresponded to a sodium citrate dihydrate scan. Polaπzed light microscopy showed the particles to be birefπngent. TGA analysis of the powder showed the powder to have 1.3% by weight of residual solvent and a decomposition temperature range of 288-395°C. The SEM image of the powder showed the particles appeared raisin-like and some facetted individual citrate crystals were observed. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 59°C. The Tg-like endotherm, which is a melt, appeared on the scan at 117°C onset temperature. The MMD of the powder sample was determined to be 1.1 lμm, with 96.7% less than 5.2 μm. Aerosol Charactenzation The DDE of the above spray dned cyclosponn A:cιtrate (90: 10) powder was determined to be 65.9%±5% (n=10). The MMAD was determined to be 3.2 μm with 78% less than 5 μm. Example 7: Cyclosporin A Spray Dried From Ethanol At 85°C Without Secondary Drying Solution Preparation 1.5 g of cyclospoπn A was dissolved in 50 mL of ethanol (200 proof, USP, NF grade from Spectrum). 31 Sprav Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% 0 2 with the following parameters: Outlet Temperature 85°C Inlet Temperature 120°C Feed Rate 5 mL/min Atomizer Flow Rate 13 ht min Secondary Drying None Powder Charactenzation PXRD of the powder showed that it exhibited a double halo with two maxima at 2Θ equal to 8.5° and 17°. Polarized light microscopy showed the particles to be birefnngent. TGA analysis of the powder showed the powder to have 0.7% by weight of residual solvent and a decomposition temperature range of 347-428°C. The SEM image of the powder showed that the particles looked raisin-like. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 65°C. The Tg-like endotherm, which is a melt, appeared on the scan at 122°C onset temperature. The MMD of the powder sample was determined to be 0.9 μm, with 97 6% less than 5.2 μm. Aerosol Charactenzation The DDE of the above spray dπed cyclosponn A powder was determined to be 70.8%±3% (n=10). The MMAD was determined to be 2.7 μm with 85% less than 5 μm. 32 Example 8: Cyclosporin A Spray Dried From Ethanol At 101°C With No Secondary Drying Solution Preparation 1.0 g of cyclospoπn A was dissolved in 33 mL of ethanol (HPLC grade). Sprav Drying A dry powder compnsed of cyclospoπn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 3 atm <5% 0 2 with the following parameters: Outlet Temperature lore Inlet Temperature 138°C Feed Rate 5 mL/min Atomizer Flow Rate 14.5 lit/min Secondary Drying None Powder Charactenzation Polanzed light microscopy showed the particles to be birefnngent. The SEM images showed that the particles were slightly dimpled. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 68°C. The Tg-like endotherm, which is a melt, appeared on the scan at 122°C onset temperature. The MMD of the powder sample was determined to be 2.3 μm, with 86.6% less than 5.2 μm. The X-ray diffraction pattern of the powder showed a halo with two broad maxima at 2Θ equal to -8.5° and -18.8°. The small angle X-ray diffraction pattern of the powder showed a peak at 20 equal to 0.2°, indicative of 2-dιmensιonal order. Aerosol Charactenzation The DDE of the above spray dned cyclospoπn A powder was determined to be 64%±6.9% (n=7). The MMAD was determined to be 2.59 μm with 75% less than 5 μm. 33 Example 9: Cyclosporin A Spray Dried From Acetone At 49°C With No Secondary Drying Solution Preparation 1.0 g of cyclosponn A was dissolved in 33 mL of acetone (HPLC grade). Sprav Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% 0 2 with the following parameters: Outlet Temperature 49°C Inlet Temperature 60°C Feed Rate 5 mL/min Atomizer Flow Rate 14.5 lit/min Secondary Drying None Powder Charactenzation Polaπzed light microscopy showed the particles to be birefnngent. The SEM images showed that the particles were round and dimpled. The MMD of the powder sample was determined to be 3.5 μm, with 70.3% less than 5.2 μm. Aerosol Charactenzation The DDE of the above spray dned cyclosponn A powder was determined to be 52.4%±2.1% (n=7). The MMAD was determined to be 2.30 μm with 63.6% less than 5 μm. 34 Example 10: Cyclosporin A Spray Dried From Acetone At 100°C With No Secondary Drying Solution Preparation 1.0 g of cyclospoπn A was dissolved in 33 mL of acetone (HPLC grade). Spray Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% O with the following parameters: Outlet Temperature 100°C Inlet Temperature 135°C Feed Rate 5 mL/min Atomizer Flow Rate 14.5 lit/min Secondary Drying None Powder Charactenzation Polanzed light microscopy showed the particles to be birefπngent. The SEM images showed that the particles were round and non-dimpled. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 66°C. The Tg-like endotherm, which is a melt, appeared on the scan at 120°C onset temperature. The MMD of the powder sample was determined to be 2.75 μm. with 76.3% less than 5.2 μm. The X-ray diffraction pattern of the powder showed a halo with a broad maxima at 2Θ equal to -7 8°. The small angle X-ray diffraction pattern of the powder showed a peak at 2Θ equal to 0.2°, indicative of 2-dιmensιonal order. 35 Aerosol Charactenzation The delivered dose efficiency (DDE) of the above spray dπed cyclospoπn A powder was determined to be 57.3%±3.42% (n=7) The mass median aerodynamic diameter (MMAD) was determined to be 2.1 μm with 70% less than 5 μm. Example 11: Cyclosporin A Spray Dried From Isopropanol At 77°C, No Secondary Drying Solution Preparation 1.0 g of cyclosponn A was dissolved in 33 mL of isopropyl alcohol (HPLC grade). Sprav Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N atm <5% 0 2 with the following parameters: Outlet Temperature 77°C Inlet Temperature 105°C Feed Rate 5 mL/min Atomizer Flow Rate 14.5 lit/mm Secondary Drying None Powder Charactenzation Polaπzed light microscopy showed the particles to be birefnngent. The SEM images showed that the particles were round and slightly dimpled. The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 66°C. The Tg-like endotherm, which is a melt, appeared on the scan at 122°C onset temperature. The MMD of the powder sample was determined to be 2.22 μm, with 85.7% less than 5.2 μm. The X-ray diffraction pattern of the powder showed a halo with 36 a broad maxima at 20 equal to -7 5° The small angle X-ray diffraction pattern of the powder showed a peak at 20 equal to 0 1°, indicative of 2-dιmensιonal order. Aerosol Charactenzation The DDE of the above spray dned cyclosponn A powder was determined to be 69.2%±2.42% (n=7). The MMAD was determined to be 3.8 μm with 97 8% less than 5 μm. Example 12: Cyclosporin A Spray Dried From Isopropanol At 104°C, No Secondary Drying Solution Preparation 1.0 g of cyclospoπn A was dissolved in 33 mL of isopropyl alcohol (HPLC grade). Sprav Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% O2 with the following parameters: Outlet Temperature 104°C Inlet Temperature 145°C Feed Rate 5 mL/min Atomizer Flow Rate 14.5 lit/min Secondary Drying None Powder Charactenzation Polanzed light microscopy showed the particles to be birefπngent The SEM images showed that the particles were round and non-dimpled The DSC scan showed a large endotherm ranging from about 20°C to about 70°C with a peak maximum at 66°C 37 The Tg-like endotherm. which is a melt, appeared on the scan at 121°C onset temperature. The MMD of the powder sample was determined to be 2.36 μm. with 89.9% less than 5.2 μm. The X-ray diffraction pattern of the powder showed a halo with two broad maxima at 29 equal to -8 9° and -19°. The small angle X-ray diffraction pattern of the powder showed a peak at 29 equal to 0.1°, indicative of 2-dιmensιonal order. Aerosol Charactenzation The DDE of the above spray dned cyclosponn A powder was determined to be 67.1%±2.85% (n=7). The MMAD was determined to be 2.7 μm with 76.8% less than 5 μm. Example 13: Cyclosporin A Spray Dried From Methanol At 63°C With No Secondary Drying Solution Preparation 1.0 g of cyclosponn A was dissolved in 33 mL of methanol (HPLC grade). Sprav Drying A dry powder compnsed of cyclosponn A was produced by spray drying the organic solution using a Buchi B-190 Laboratory Spray Dryer with N 2 atm <5% 0 2 with the following parameters: Outlet Temperature 63°C Inlet Temperature 88°C Feed Rate 5 mL/min Atomizer Flow Rate 14.5 lit min Secondary Drying None 38 Powder Charactenzation Polanzed light microscopy showed the particles to be birefπngent. The SEM images showed that the particles were very dimpled. The MMD of the powder sample was determined to be 2.37 μm. with 90.1% less than 5.2 μm. The DSC scan showed a large endotherm ranging from about 20-70°C with a peak maximum at 52°C. The Tg- like endotherm, which is a melt, appeared on the scan at 119°C onset temperature. Aerosol Characterization The DDE of the above spray dried cyclosponn A powder was determined to be 67.2%±3.43% (n=6). The MMAD was determined to be 2.5 μm with 80.7% less than 5 μm. Example 14: The method of Example 1 was followed except that the solution was atomized using a standard, commercially available, Buchi nozzle. The mass mean diameter (MMD) of the droplets using this nozzle was between 7 and 15 μm. Chemical Stability HPLC analysis showed no appreciable degradation from the samples of the spray dried cyclospoπn A powder. Figure 7 is a chromatogram of the spray dried sample reconstituted in a mobile phase after storage for 15 months at room temperature. The powder was determined to be chemically stable over the time course of the study. 39 Example 15: Processing of Spray Dried Cyclosporin A Powder Formulations Spray dπed cyclospoπn A powders were produced from vanous solvents using a vanety of spray drying temperatures with or without secondary drying. Secondary drying appeared to have no effect on particle properties. Measurement of residual solvent showed that little solvent was left in the particles. Lower levels of residual solvent are preferred to minimize any possible lung lrntation caused by solvent The percent yield, defined as the weight of CsA-containing powder recovered in the collector of the spray dryer divided by the weight of CsA (and any excipient) in the solution which was spray dried (times 100%), ranged from 22% to 78%. Yields of at least about 20% are preferred, with higher yields being generally more preferred, so long as other powder characteπstics such as MMD and DDE are acceptable. The fine particle traction (%). defined as DDE times the % <5 μm. ranged from 33.3 to 67 7%. Powders with a tine particle fraction of at least about 25% are preferred. Several batches of CsA powders were spray dned from ethanol at 70°C without secondary drying. The results of these batches is presented in Table 2. 40 Table 2. CsA Powders Spray Dried from Ethanol at 70°C with No Secondary Drying Trial Yield MMD DDE % <5 μm Fine % μm % Particle Fraction 1 70.0 0.8 71.4 85.6 61.1 2 66.3 1.6 78.6 84.4 66.4 3 59.5 1.7 76.3 78.0 59 5 Modification of the above-descnbed modes of carrying out the vanous embodiments of this invention will be apparent to those skilled in the art following the teachings of this invention as set forth herein. The examples descnbed above are not limiting, but are merely exemplary of this invention, the scope of which is defined by the following claims. The disclosure of each publication, patent or patent application mentioned in this specification is hereby incorporated by reference to the same extent as if each individual publication, patent or patent application were specifically and individually indicated to be incorporated by reference. 41

Claims: It is Claimed: 1. Cyclosponn in liquid crystal form. 2. The cyclosponn of Claim 1 wherein the cyclosponn is cyclosponn A. 3. The cyclosponn of Claim 1 which shows sharp peaks by small angle X-ray scatteπng. 4. The cyclospoπn of Claim 1 which is in dispersible powder form. 5. The cyclosponn of Claim 1 for use as an immunosuppressive, anti- lnflammatory, or anti-asthmatic agent. 6. The cyclospoπn of Claim 1 which is prepared by spray drying from a solvent. 7. A composition for pulmonary delivery compnsing liquid crystal cyclospoπn in respirable powder particles. 8. The composition of Claim 7 wherein the cyclospoπn is cyclosponn A. 9. The composition of Claim 7 which is dispersible. 10. The composition of Claim 7 which further compnses a pharmaceutically acceptable excipient or earner. 11. The composition of Claim 7 which is prepared by spray drying. 42 12. The composition of Claim 7 wherein the cyclosponn comprises at least about 40% by weight of the composition. 13. The composition of Claim 7 wherein the particles in the powder have a particle size range between 0.1 and 15 μm MMD. 14. The composition of Claim 7 wherein the particles have an MMAD of less than about 5 μm. 15. The composition of Claim 7 which has a delivered dose efficiency of at least about 30%. 16. A method for preparing the composition of Claim 7 compnsing: a) mixing cyclosponn with a solvent to form a solution or suspension; and b) spray drying the mixture formed in step a) under conditions which provide a respirable powder. 17. The method of Claim 16 further compnsing the step of adding a pharmaceutically acceptable excipient or earner pnor to spray drying. 18. The method of Claim 17 wherein said solvent compnses a solution of less than 50% water. 19. The method of Claim 16 wherein the solvent is selected from the group consisting of ethanol, acetone, acetonitπle, isopropanol and methanol. 43 20. A method of treating or preventing a condition in a subject which may be prevented or alleviated by cyclosponn. the method compπsing pulmonary administration of a therapeutically effective amount of the composition of Claim 7 to a subject susceptible to or suffenng from the condition. 21 The method of Claim 20 wherein the condition is selected from the group consisting of asthma, transplant rejection, sarcoidosis, chronic inflammatory lung disease, chronic obstructive pulmonary disease, emphysema, pnmary and secondary pulmonary hypertension, cystic fibrosis. lung infections, rheumatoid arthπtis and ldiopathic pulmonary fibrosis. 44

Inventors: BENNETT DAVID B, CABOT KIRSTEN M, FOSTER LINDA C, LECHUGA-BALLESTEROS DAVID, PATTON JOHN S, TAN TRIXIE K, BENNETT, DAVID, B., CABOT, KIRSTEN, M., FOSTER, LINDA, C., LECHUGA-BALLESTEROS, DAVID, PATTON, JOHN, S., TAN, TRIXIE, K.
Applicants: INHALE THERAPEUTIC SYST, INHALE THERAPEUTIC SYSTEMS, INC.
Application year: 1999
Main classification: A61K38
IPC codes: A61K9, A61K38, A61P11, A61P29, A61P37