MXPA02007187A - A medicinal aerosol formulation. - Google Patents

Abstract

A medicinal formulation is disclosed. The formulation comprises: a therapeutic amount of a protein or peptide medicament, a fluid for containing said medicament having a molecular size ranging from 1 K Dalton to about 150 K Daltons, a fluid carrier for containing the medicament, and a stabilizer selected from an amino acid, a derivative thereof or a mixture of the foregoing.

Description

MEDICINAL FORMULATION IN AEROSOL This application claims the priority of the provisional application of E.U.A. Serial No. 60 / 177,987, filed January 25, 2000, which is incorporated herein by reference.

BACKGROUND OF THE INVENTION Field of the Invention This invention relates to a medicinal aerosol formulation, and more particularly, to a medicinal aerosol formulation comprising a protective colloid stabilizer.

Description of Related Art Delivery of drugs to the lung through inhalation is an important means of treating a variety of conditions, including common local conditions such as fibrosis, pneumonia, bronchial asthma and chronic obstructive pulmonary disease and some systemic conditions including management of pain, immune deficiency, hormonal therapy, erotropolesis, diabetes, etc. Steroids, β2 agonists, anticholinergic agents, proteins and polypeptides are among the drugs that are administered to the lung for lM? L? éÉátt * ****** J¿- *? ai i i mfliit if nur * 8 * 1- A-ts-1 purposes, These drugs are commonly administered to the lung in the form of an aerosol of particles of respirable size (less than about 10 um in diameter), In order to ensure the appropriate particle size in the aerosol, the particles can be prepared in size Breathable and then incorporated into a colloidal dispersion containing either a propellant, such as a pressure metered dose inhaler (MDI), or air such as is the case with a dry powder inhaler (DPI). Alternatively, the formulations can be prepared in the form of an emulsion or solution in order to avoid the problem of appropriate particle size in the formulation. The formulations in solution, however, must be supplied in a manner that produces particles or droplets of respirable size. For MDI preparations, once prepared, the aerosol formulation is filled into an aerosol can equipped with a metered dose valve. In the hands of the patient, the formulation is delivered through an actuator adapted to direct the dose from the valve to the patient. It is important that an aerosol formulation be stable so that the dose delivered, discharged from the metered dose valve, is reproducible. Rapid formation of cream, sedimentation, or flocculation TÜÍ, AÉé-ÉÉlÉiÍfiiriit1r < l ?? ipi iiiiÉ '.. ^ - • - --.-., ....- .., .. ^^^ 3 after agitation, they are common sources of non-reproduction of doses in suspension formulations. This is especially the case when a binary aerosol formulation containing only medically and propellant, v.gr ,, 1, 1, 1, 2-tetrafluoroethane, is employed or when said formulation contains small amounts of surfactant as well. Adhesion of the valve can also cause inability to replicate doses. In order to overcome these problems, MDI aerosol formulations often contain surfactants, which serve as suspension aids to stabilize the suspension for a sufficient time to allow dosing reproducible. Certain surfactants also function as lubricants to lubricate the valve to ensure smooth operation. A myriad of materials are known and described for use as dispersion aids in aerosol formulations. The proper quality of materials, however, depends on the particular drug and the propellant or type of propellant used in the formulation. It is sometimes difficult to dissolve sufficient amounts of conventional surfactants of hydrofluorocarbon (HFC) propellants such as HFC-134a and HFC-227. The cosolvents, such as ethanol. have been used to overcome this problem, as described in the U.S. Patent. No, 5,225,183. An alternative approach that avoids cosolvents involves materials that are soluble or homogeneously dispersible in hydrofluorocarbon propellants and are said to be effective surfactants and dispersion aids in an aerosol formulation. Among these materials are certain fluorinated surfactants and certain polyethoxy surfactants. Drugs that are relatively small molecules are much more predictable in terms of their aerosol formulation characteristics than macromolecules. The macromolecules, such as peptides or proteins, ranging in molecular size from about 1K Dalton to about 150K Dalton in molecular size are highly unpredictable and present unique problems in forming aerosol formulations thereof that are stable and provide reproducible dosing. Most of the peptide and protein drugs, such as hormones, eg, insulin, amylin, etc., enzymes, antinfective, are very variable in their amino acid composition and three-dimensional structure. Consequently, its superficial activity is highly variable, and importantly, it is not yetÍti ??.?. Áí? .. ** f * ik4. * > * J ...... available a model that explains the differences in surface activity of protein based on its most basic and structural properties, such as molecular weight, adsorption capacity, solubility, division coefficient and isoelectric pH. Hemoglobin, for example, has a much higher affinity for solid surfaces than albumin, however, the molecular weights of these two proteins are very similar. Fundamentally, the diversity in surface activity of peptides and proteins originates in the linear amino acid sequence that uniquely characterizes each type of protein. The amino acid side chains often vary dramatically in that some carry no charge at any pH, however they exhibit considerable polar character (serene, threonine). Other amino acids are ionizable and vary from regularly acidic (aspartic and glutamic acid are completely negatively charged at the physiological pH of 7.4) to basic functionalities, such as the imidazole group in histidine (which carries a partial positive charge at pH 7.4), and even more basic amino groups in lysine and arginine that carry full positive charges at a pH of 7.4. Another group of amino acids, somewhat similar to hydrocarbon in character, generally appears to have a much lower solubility profile in water (tryptophan. -feet phenylalanine, isoleucine, etc) than many of the other amino acids found in biological systems It is to be noted that the hydrophobicity of these amino acids that water rejection varies greatly with their specific structure in the protein. For example, the side chain of simple methyl group in alanma contributes only 0.5 kcal / mol to the free energy of water transfer to an organic phase, while the indo group! of double ring in tpptofán contributes 0 3.4 kcal. The variety of amino acid side chains, together with the many different types of chemical interactions that result in solution and on surfaces, should be expected to have a considerable impact on the stability of the aerosol formulation as well as transport of these peptide thioterapic agents and protein through biological membranes The diverse character of amino acid side chains, together with the completeness of various combinations of amino acids present in each particular protein, means that the physicochemical properties of proteins, their intermolecular as well as intramolecular reactivity , and also their ability to interact with surfaces must be highly variable Due to their large size, and correspondingly due to the Large numbers of charged amino acid side chains, proteins have many charges distributed through their outer surface This could lead to very large variations in the aerosol formulation and lung admission stability of these compounds. The polypeptide and protein drugs also generally have multiple ionization sites and, therefore, frequently demonstrate pH-dependent solubility profiles. Importantly, the hydrophilic nature of these compounds provides excellent conditions for high aqueous solubility. Consequently, the majority of the Protein and protein drugs exhibit extremely low lipid solubility characteristics, the latter being possible as a reason why the dispersions of these drugs in hydrofluorocarbon propellants would be physically and chemically stable through a wide range of storage conditions. An aerosol drug formulation comprising peptide and protein drugs in carrier or formulation medium within which they are virtually insoluble, is needed to reduce the hydrolytic and chemical deactivation usually typical of aqueous solutions. The combination of a large surface area, thin absorption carrier, and extensive vasculature it constitutes a favorable absorbent environment for proteins and peptides when delivered by the pulmonary route. Studies show that intratracheal (i.t.) peptide administration is rapid and quantifiable; however, the resulting distribution is often located in central airways. Administration by aerosol, for example, depending on the particle size distribution, can be used to provide more uniform distribution with increased alveolar penetration. The absorption of drug from the airways depends on the deposition site, the method of drug delivery, the type of solute presentation and composition of the formulation. Therefore, the formulation and device characteristics will have a dramatic impact on the regimen and extension of peptide absorption from the lung. Studies show that the absorption regimes after aerosol delivery of small molecular weight compounds can be approximately twice the i.t. What is desired is to present peptides and proteins as hydrophobic dispersions through a multi-dose inhalation device ("pMDI") in order to have greater penetration of the drug particles into the peripheral lung wherein absorption must be significantly greater than for the drug deposited centrally as in the case with aqueous instillations. The amount that can be absorbed from the lung's insulin into the bloodstream has been demonstrated by a number of scientists. A 1990 journal article [Lung, supplement pp 677-684] showed from multiple studies that aerosolized insulin delivered to the lung provided a half-life of 15-25 minutes, but the results were highly variable. Comprehensible studies have shown that aerosolized insulin delivered peripherally to the lung of mice produced a bioavailability of more than 50.7 percent in contrast to 5 to 6 percent biodispomption seen in liquid insulin dripped into the central airways. These studies, therefore, support the contention that aerosolized insulin must be delivered peripherally to the lung for maximum efficiency and that the inadvertent central arrangement of inhaled aerosolized insulin will produce an effect ten times lower than desired. These ten-fold dosage variations are clearly unacceptable if aerosolized insulin should become an effective means of treating diabetes. Thus, there is a need for effective, high-precision aerosol devices to achieve the tolerances required to subject insulin to its contents Yes _ # t Út'- iiliüálfi 'r r humans. This concept to achieve the tolerances required to aerosolize insulin in diabetes management would also apply to amilma and glucagon, hormones associated with insulin in the regulation of plasma glucose concentration, which until now, must be administered by subcutaneous injection ( sc). Dry powder presentations of peptide and protein drugs have unique opportunities in formulations, which do not occur in liquid presentations such as pMDIs and nebulized solutions. The dry powder aerosols of peptide and protein drugs, due to the improved solid state stability, are attractive from the formulation point of view since much of the undesirable solution and interactive effects of liquid state are circumvented. In this regard, reference is made to Rubsamen et al, U.S. Patent No. 5,672,581 and Patton et al., In the U.S. Patent. No. 5,775,320. Both approaches of Rubsamen and Patton are therapeutically feasible even when their complexity and expected inherent costs limit their applicability in the management of a chronic disease such as diabetes mellitus. In this way, it is a problem to use a complicated, expensive device such as the portable nebulizer, electronically based to deliver Routinely hipollicic to patients who need them. It is also a problem to use a large, bulky, hard-to-clean dry powder aerosol device such as the Patton device to deliver hypoglycemics to the body through the lung. Thus, the primary objective in formulating a peptide or protein drug as a dry powder inhalation aerosol (DPI) is to allow the drug, and in some cases, added excipients, to form an aerocolloid that is chemically and physically stable and it can remain in suspension until the drug particle reaches the alveolar or other absorption sites. Once in the absorption site, the drug particles must be trapped efficiently at the deposition site, rapidly dissolve in the epithelial lining fluids, and rapidly absorbed through the biomembrane, thus limiting the possible deactivation by enzymes of metabolization in the airways. Spray drying is a process used to prepare drug particles for drug formulations. Spray drying is a single-stage process that transforms a solution or suspension into a fine powder. Generally, spray drying produces particles fca * ^ *? * t ^? IM? JU * l¡H? hj ** l? d * !? spherical, which are often hollow, resulting in a powder with low volume density compared to the initial material. The powder characteristics of spray-dried materials (ie, particle size distribution, volume density, porosity, moisture content, dispersibility, etc.) are generally good in many aspects, but the particles manufactured by this process show low flow characteristics. In addition, a requirement for heating during particle formation by this process makes spray drying less desirable for heat-sensitive compounds such as peptide and protein drugs. Thus, it is a problem that most dry powder aerosols demonstrate adhesion and low fluidity through the equipment of the device to the extent that the accuracy of dose delivery becomes a problem for the patient. Another problem associated with the formulations of peptide and protein as dry powder aerosols is that of packing the material as agglomerates in such a device that during the aerosolization, the agglomerates are broken, and the individual particles are released before entry into the airways. The preparation of robust agglomerates of particles of micron or submicron size is a reasonably direct task that is can be achieved by conventional granulation, with or without polymeric binders. However, the requirement that upon entering the airways the agglomerates must be broken into primary particles, it probably eliminates a simple conventional approach to granulation since the microparticle forces could be too large to allow simple, efficient and rapid deagglomeration. total adhesive between two different particles or total cohesive force between two similar particles can be considered as being constituted by a sum of one or more attractive forces Many of these forces are known to be responsible for the formation of adhesive units between dry powder and particles of excipient in formulations Therefore, the goal of any manipulation of forces between particles will be to produce agglomerates of between say, 50 and 200 um in diameter that are robust enough to support the flow, storage and packing in the delivery device, but that they can deagglomerate quickly and completely ect by the shear stresses in the inspired air stream This problem, which is very common in the aerosol formulations of the protein and the peptide, can be completely avoided in liquid formulations within which the drug is insoluble. tJj¡, ÉJÉ¿-t ^ í ». ^ - ^ -.-- s - * - j-tf ftl * 'tt-rflliff"' "i r r * 1 * ~ ^ ** - * - - It presents as a colloidal dispersion, and is sterically protected against self-association. Therefore, a desire is to formulate peptide and protein drugs as flocculated colloids, loose in non-aqueous media, such as hydrofluorocarbons, which break quickly and easily into discrete particles during aerolization to the airways Additionally, it is desired to present peptide and protein drugs in formulation systems within which the drug particles are perpetually in random motion, thereby eliminating aggregate formation of the individual drug particles. Other non-injectable diabetes therapies have been proposed, some demonstrating that a biotherapeutic response could be produced following nasal administration of insulin when formulated with detergents and other membrane penetrants, as indicated in Moses et al. Diabetes, Vol 32, November 1983; and Salzman et al., New England Journal of Medicine, Vol, 312, No 17. Variability between significant subject and irritation of nasal membranes to varying degrees is observed Since diabetes is a chronic disease that must be continuously treated by the patient. administration of insulin, and since mucosal irritation tends to increase with the a * M ..- .. ^^? ^ jj ^? * l * m? * ji ^ fo. * ¡*, i Repeated exposures to membrane penetration enhancers, efforts to develop an insulin administered nasally in a non-invasive manner have not been commercialized. Consequently, a safe, reproducible, effective, non-invasive delivery means for drug and protein via the lung as pMDIs is desired and needed.

SUMMARY OF THE INVENTION It has surprisingly been found that novel and stable medicinal aerosol formulations of macromolecular drugs can be obtained without the use of cosolvents, such as ethanol, or surfactants, such as sorbitan trioleate which are added to an aerosol formulation. primary of small molecule drugs. Stable medicinal aerosol formulations are obtained by the use of a protective colloid stabilizer.

DETAILED DESCRIPTION OF THE INVENTION This application makes reference to the application of E.U.A. Serial No. 09 / 158,369 filed September 2, 1998, which is hereby incorporated by reference in its entirety. This invention involves a formulation of stable suspension spray suitable for delivery under pressure comprising (1) a medicament or macromolecular drug in particles, (2) an appropriate propellant, and (3) an appropriate stabilizer. An appropriate drug or macromolecular drug is one that is suitable for administration by inhalation, the inhalation being used for oral and nasal inhalation therapy. A colloidal, stable dispersion of a drug in a fluid, v gr. , air, hydrocarbon gases, chlorofluorocarbon (CFC) propellants and non-CFC propellants, such as tetrafluoroethane (HFA-134a) and heptaf luoropropane (HFA-227) are described. A stabilizer of a polyionic species, such as an amino acid and a small molecule peptide, as inactive formulation components that trigger the loss of adhesive bond strength between the drug particles is employed. An electret or sterically stabilized aerocolloid particles of the selected drugs is formed in this way. Electretes are the electrostatic equivalent of permanent magnets but may be susceptible to rupture in the presence of moisture, such as that present in air or in ambient humidity conditions of the respiratory tract. Consequently, the present invention is applies to dry powder aerosols, portable nebulizer systems, as well as pressurized metered dose inhaler formulations. The resulting aerocolloid is chemically and physically stable and can remain in suspension until the selected drug or drug particles reach the alveolar or other sites in a patient's airway., e.g., human, animal, being treated Once in the absorption site, the drug particles should be trapped efficiently at the deposition site as a result of moisture in the environment, dissolve rapidly in the fluids of epithelial lining, and they are quickly absorbed through the patient's biomembranes, limiting in this way the possible deactivation by enzymes of metabolism in the airways. An appropriate medicament to which the present invention is directed is one that forms a stable hydrophobic dispersion, suitable for delivery to a patient, e.g., human or animal. Typically, the medicament includes a polypeptide, polypeptide, or biotherapeutic protein ranging from 0.5 K Dalton to 150 K Dalton in molecular size. In particular, the protein peptide, polypeptide or biotherapeutic drug includes diabetic aids; insulins and analogues insulin; amylin; glucagon; surfactants; immunomodulation peptides such as cytokines, chemokines, lymphokines, interleukins such as taxol, interleukin-1, interleukin-2, and inter-ferons; erythroproteins; thrombolytics and heparins; anti proteases, antitrypsins and amiloride; rhDNase; antibiotics and other anti-infectives, hormones and growth factors such as parathyroid hormones, LH-RH analogues and GnRH; nucleic acids; DDAVP; calcitonins; cyclosporin; ribavirin; enzymes; heparins; hematopoietic factors, cyclosporins; vaccines; immunoglobulins; vasoactive polypeptides; antisense agents; genes, oligonucleotides, and nucleotide analogues. The term diabetic help includes natural, synthetic, semisynthetic and recombinant drugs such as activin, glucagon, insulin, somatostatin, proinsulin, amylin, and the like. The term "insulin" will be construed as encompassing natural extracted human insulin, recombinantly produced human insulin, insulin extracted from bovine and / or porcine sources, recombinantly produced porcine and bovine insulin and mixtures of any of these insulin products. The term is intended to encompass the polypeptide normally used in the treatment of diabetics in a substantially purified form, but encompasses the use of the term in its commercially available pharmaceutical form, which includes additional excipients. The insulin is preferably produced recombinantly and can be dehydrated (completely dry) or in solution. The terms "insulin analog", "monomeric insulin" and the like are also used interchangeably herein and are intended to encompass any form of "insulin" as defined above, wherein one or more of the amino acids within the polypeptide chain has been replaced with an alternative amino acid and / or wherein one or more of the amino acids have been omitted and wherein one or more additional amino acids has been added to the polypeptide chain or amino acid sequences that act as insulin at decrease blood glucose levels. In general, the "insulin analogs" of the present invention include "insulin lispro analogs," as described in US Patent No. 5,547,929, incorporated herein by reference in its entirety, insulin analogs including insulin LysPro and insulin humáloga, and other "super insulin analogues", where the ability of the insulin analog to affect serum glucose levels is improved ki.¿ .. a ^ taj-Hm * t fi-p-JF * - "-t« - • ---- J-fc ----- * i »J-ja * jtf¡M-aa -ia »-------.» - «- substantially in comparison with conventional insulin as well as heptoselective insulin analogues that are more active in the liver than in adipose tissue. Preferred analogs are monomeric insulin analogs, which are insulin-like compounds used for the same general purpose as insulin such as insulin lispro, ie, compounds that are administered to reduce blood glucose levels. The term "amylin" includes natural human amylin, bovine amylin, porcine, rat, mouse, as well as synthetic, semisynthetic or recombinant amylin or amylin analogues including pramlmtide and other amylin agonists as described in the US Patent No. 5,686,411, and US Patent No. 5,854,215, both of which are hereby incorporated by reference in their entirety. The term "immunomodulation proteins" includes cytokines, kinokines, lymphokine complement components, accessory and adhesion molecules of the immune system and its receptors of human or non-human animal specificity Useful examples include GM-CSF, IL-2, IL-12 OX40, OX40L (gp34), lymphotactin, CD40, CD0OL. Useful examples include interleukins for example interleukins 1 to 15, ? J.i Ml * .tAHfah. - ..... ^ JaU ^ fca interferon alpha, beta or gamma, tumor necrosis factor, granulocyte-macrophage colony stimulation factor (GM-CSF), macrophage colony stimulus factor (M-CSF), granulocyte colony stimulus factor (G-CSF), chemokines such as neutrophil activation protein (NAP), macrophage chemoattractant and activation factor (MCAF), RANTES, macrophage inflammatory peptides MlP-la and MlP-lb, complement components and their receptors, or an accessory molecule such as B7.1, B7.2, ICAM-1,2 or 3, and cytokine receptors, OX40 and ligand-OX40 (gp34) are additional useful examples of immunomodulatory proteins. Immunomodulatory proteins for various purposes may be of human or non-human animal specificity and may be represented for the present purposes, as the case may be and as may be convenient, by extracellular domains and other fragments with the binding activity of the proteins that occur naturally, and muteins thereof, and their fusion proteins with other polypeptide sequences, e.g., with immunoglobulin heavy chain constant domains. When nucleotide sequences encoding more than one immunomodulation protein are inserted, for example, they may comprise more than one cytokine or a combination of cytokines and accessory / adhesion molecules.

The term "interferon" or "IFN" as used herein means the family of specific proteins in highly homologous species that inhibit viral replication and cell proliferation and modulated immune response. Interferons are grouped into three classes based on their cellular origin and antigenicity, alpha-inter-feron (leukocytes), beta-interferon (fibroblasts) and gamma-interferon (immunocompetent cells). The recombinant and analogous forms of each group have been developed and are commercially available. The subtypes in each group are based on antigenic / structural characteristics. At least 24 alpha interferons (grouped in subtypes A through H) having different amino acid sequences have been identified by isolating and encoding DNA in sequence these porptides. See also Viscomi, 1996 Biotherapy 19 '59 -86, the contents of which are incorporated by reference herein in their entirety The terms "alpha-interferon", "alpha inte ferone", "alpha-interferon", "leukocyte interferon" "and IFN are used interchangeably herein to describe members of this group Both naturally occurring and recombinant alpha interferons, including consensus interferon such as that described in US Patent No. 4,897,471, Í, _-_-._ * _ ij - i -A_.il. _- ,. *. * - .. --S -a.

The content of which is incorporated herein by reference in its entirety, can be used in the practice of the invention. Human leukocyte interferon prepared in this manner contains a mixture of human leukocyte interferons having different amino acid sequences. The purified natural human alpha interferons and mixtures thereof which can be used in the practice of the invention include, but are not limited to, interferon alfa-nl (Ins) Welfferong available from Glaxo available from Sumitomo, Japan, interferon alfa-nl (Ins) -Wellcome Ltd., London, Great Britain; and interferon alfa-n3 Alferon RTM available from Purdue Fredepck Co, Norwalk, Conn The term "erythropoietin" is applied to isolated synthetic, semisynthetic, recombinant, natural, human, monkey, or other animal or microbiological polypeptide products having part or all the primary structural conformation (ie, continuous sequence of amino acid residues) and one or more of the biological properties (v.gr, immunological properties and biological activity in vivo and in vitro) of erotropoietin that occurs naturally, including allelic variants of the same These polypeptides are also uniquely characterized as being the prokaryotic host expression product or | ¿^ J j § ¿> * ...- .._ .. »*.-_« - »> , ...-. --- - * ------ < - «faith --- * aj-a eukaryotic (v. g., by bacterial, yeast and mammalian cells in culture) of exogenous DNA sequences obtained by genomic or cDNA cloning or by gene synthesis. Products of microbial expression in vertebrate cells (v. gr, mammal or bird) they can also be characterized by freedom of association with human proteins or other contaminants that can be associated with erythropoietin in their natural mammalian cell environment or in extracellular fluids such as plasma or urine. The typical yeast products (e.g., Saccaromyces cerevisiae) or prokaryote (e.g., E. coli) host cells are free from association with any mammalian proteins. Depending on the host employed, the polypeptides of the invention can be glycosylated with mammalian or other eukaryotic carbohydrates or may be non-glycosylated The polypeptides of the invention may also include an initial methionine amino acid residue (in the -1 position). The novel glycoprotein products of the invention include those having primary structural conformation sufficiently capable of duplication of that of an erythropoietin occurring naturally (v. g., human) to allow possession of one or more of the biological properties thereof and having a composition of average carbohydrate that differs from that of erythropoietin that occurs naturally (v. gr., human). The terms "heparins" and "thrombolytics" include anticoagulation factors such as heparin, low molecular weight heparin, tissue plasminogen activator (TPA), urokinase (Abbokinase) and other factors used to control clots. The terms "anti proteases" and "protease inhibitors" are used interchangeably and are applied to synthetic, semisynthetic, recombinant, naturally occurring or non-naturally occurring, soluble or immobilized agents reactive with receptors, or act as antibodies enzymes or nucleic acids . These include receptors that modulate a humoral immune response, receptors that modulate an immune cell response (e.g., T-cell receptors) and receptors that modulate a neurological response (e.g., glutamate receptor, glycine receptor, acid gamma-aminobutyric acid (receptor (GABA)) These include cytokine receptors (involved in arthritis, septic shock, transplant rejection, autoimmune disease and inflammatory diseases), the main histocompatibility receptors (NHC) Class I and II associated with presentation of antigen to cytotoxic T-cell receptors and / or T-cell receptor receptor Í? S l. ? .i *? * - t .i. - ^ - -.- - - - - - - -. - - ~.-.-- > «- ~. ~ ..- ...- * > ^ AA.l lj (involved in autoimmune diseases) and the thrombin receptor (involved in cardiovascular disease coagulation). The list also includes antibodies that recognize autoantigens such as those antibodies implicated in autoimmune disorders and antibodies that recognize viral antigens (v. Gr, HIV, herpes simplex virus) and / or microbes. The terms "hormones" and "growth factors" include hormone that releases hormones such as growth hormone, thyroid hormone, thyroid-releasing hormone (TRH), gonadotropin-releasing hormone (GnRH), leuteinization hormone, leutemization hormone-releasing hormone (LHRH, including superagonists and antagonists such as leuotrplide , deltirelix, gosorelin, nafarelin, danazol, etc.) from natural, human, porcine, bovine, ovine, synthetic, semi-synthetic or recombinant sources. These also include somatostatin analogues such as octreotide (Sandostatin). Other agents in this category of biotherapeutics include medications for uterine contraction (v gr, oxytocma), diuresis (v. G., Vasopressin), neutropenia (v.gr, GCSF), respiratory disorders (e.g., superoxide dismutase), RDS (e.g., surfactants, optionally including apoproteins), and the like Í.Á 1AÁ W-i. -.... t ¿t-L- i-? -. "» ...., J. a ..._. t > . ^, iriÉ.Íifc, t The term "enzymes" includes recombinant deoxipbonuclease such as DNAse (Genentech) from Corporation, proteases (e.g., serum proteases such as trypsin and thrombin), polymerases (vgr, RNA polymerases, DNA polymerases), transcpptases and Reverse coughs, enzymes involved in arthritis, osteoporosis, inflammatory diseases, diabetes, allergies, organ transplant rejection, oncogene activation (eg, dihydrofolate reductase), signal transduction, auto-cycle regulation, transcription, replication of DNA and repair. The term "nucleic acids" includes any segment of DNA or RNA that contains naturally occurring or non-naturally occurring nucleosides, or other proteinoid agents capable of binding specifically to other nucleic acids or oligonucleotides through complementary hydrogen bonding and are also capable of agglutination. to non-nucleic acid ligands In this regard, reference is made to Bock, L, et al, Nature 355: 564-566 (1992) reporting the inhibition of thrombin-catalyzed conversion of fibpnogen to fibrin using aptamer DNA. Biological molecules for which the lead molecules can be synthesized and selected in accordance with the invention include, but are not limited to, agonists and antagonists for cell membrane receptors, neurotransmitters, toxins and venoms, viral epitopes, hormones, opiates, steroids, peptides, enzyme substrates and inhibitors, cofactors, drugs, lectins, sugars, oligonucleotides, nucleic acids , oligosaccharides, lipids, proteins, and analogs of any of the above molecules. The term "analogue" refers to a molecule, which shares a common functional activity with the molecule that is considered to be analogous and typically shares common structural features as well. The term "recombinant" refers to any type of cloned biotherapeutical expressed in prokaryotic cells or a molecule made by genetic engineering, or combinatorial release of molecules that can be further processed to another state to form a second combinatorial library, especially molecules that contain protecting groups that improve the physio-chemical, pharmacological and clinical safety of the biotherapeutic agent. The term "vaccines" refers to therapeutic compositions for stimulating humoral and cellular immune responses, either isolated, or through a cell »K? - * • faith - * ^ - "- * ..? * L- .., .., .. • * - - ------ • - -,.« * *? Í? ? presenting antigen, such as an activated dendritic cell, which is capable of activating T-cells to produce a multivalent cellular immune response against a selected antigen. The cell having a potent antigen is stimulated by exposing the cell in vitro to a polypeptide complex. The polypeptide complex may comprise a dendritic cell binding protein and a polypeptide antigen, but preferably, the polypeptide antigen is either a tissue-specific tumor antigen or a gene-oncogene product. However, it is observed that other antigens, such as viral antigens can be used in said combination to produce immunostimulatory responses. In another preferred embodiment, the dendritic cell binding protein that is part of the immunostimulatory polypeptide complex is GM-CSF. In a further preferred embodiment, the polypeptide antigen that is part of the complex is the tumor specific antigen prostatic acid phosphatase. In still other preferred embodiments, the polypeptide antigen can be any of the oncogene product peptide antigens. The polypeptide complex may also contain, between the dendritic cell binding protein and the polypeptide antigen, a binding peptide. The polypeptide complex can | tt t, a _, .- l.J. i -...- .. I i.I. jfc-a ..- ..-.-...- ü a, «& JLá .. i comprising a dendritic cell binding protein covalently linked to a polypeptide antigen, said polypeptide complex being preferably formed from a dendritic cell binding protein, preferably GM-CSF, and a polypeptide antigen, The polypeptide antigen is preferably a tissue-specific tumor antigen such as prostatic acid phosphatase (PAP), or an oncogene product, such as Her2, p21RAS, and p53, however, other modalities, such as viral antigens, are also within the scope of The invention The term "immunoglobulins" encompasses polypeptide oligonucleotides involved in host defense mechanisms such as coding and decoding by one or more gene vectors, conjugating various nucleic acid binding moieties in host defense cells, or coupling expressed vectors to help in the treatment of a human or animal subject. The drugs included in this class of polypeptides include IgG, IgE, IgM, IgD, either individually or in combination with one another. For purposes of the formulations of this invention, which are intended for inhalation into the lungs, the drug or drug is micronized preferably by which an amount Therapeutically effective or fraction (eg, ninety percent or more) of the drug is in particles. Typically, the particles have a diameter of less than about 10 microns, and preferably less than about 5 microns, so that the particles can be inhaled into the respiratory tract and / or lungs. The drug or drug selected is present in the formulations of the invention in a therapeutically effective amount, ie, an amount such that the drug can be administered as a dispersion, aerosol, oral or nasal inhalation, and causes its desired therapeutic effect, typically preferred with one dose, or through several doses. The drug is typically administered as an aerosol of a conventional valve, eg, a metered dose valve, through an aerosol adapter is also known as an actuator. The term "amount" as used herein refers to an amount or a concentration as appropriate to the context. The amount of a drug that constitutes a therapeutically effective amount varies in accordance with factors such as the potency of the particular drug, the route of administration of the formulation, and the mechanical system used to administer the formulation. An amount Therapeutically effective of a particular drug can be selected by those of ordinary experience in the field with due consideration to said factors. Generally, a therapeutically effective amount will be from about 0.001 parts by weight to about 5 parts by step based on 100 parts by weight of the selected fluid or propellant. A suitable fluid includes air, a hydrocarbon such as n-butane, propane, isopentane, etc., or a propellant. A suitable propellant is any fluorocarbon, vg, a 1-6 hydrogen containing f luorocarbon (such as CHF2CHF3, CF3CH2F, CH2F2CH3 and CF3CHFCF3), a perfluorocarbon, v gr. , a perfluorocarbon of 1-4 carbons, (such as CF3CF3, CF3CF2CF3); or any mixture of the above, having a sufficient vapor pressure to make them effective as propellants. Some typical suitable propellants include conventional chlorofluorocarbon (CFC) propellants such as propellants 11, 12 and 114 or a mixture thereof. Non-CFC propellants such as 1, 1, 1, 2-tetraf luoroethane (Propellant 134a), 1, 1, 1, 2, 3, 3, 3-heptaf luoropropane (Propellant 227) or a mixture thereof are preferred The fluid or propellant of preference is present in an amount sufficient to drive a plurality of doses ^ ij ^ ... ^ A1.1 i 1 .. selected from a spray can when it is used. An appropriate stabilizer is selected. An appropriate stabilizer includes (1) an amino acid selected from (a) a monoamino carboxylic acid of the formula, H2N-R-COOH (I), (b) a monoamino dicarboxylic acid of the formula H2N-R (COOH) 2 (II) ) and (c) a diamino monocarboxylic acid of the formula (H2N) 2R COOH (III), wherein R is a straight or branched alkyl radical of 1 to 22 carbon atoms, which may be mono or poly-substituted with such moieties as sulfide (-S-), oxide (-0-), hydroxyl (-OH), amide (-NH), sulfate (-S04); aril of the formula wherein X is hydrogen, halogen (F, Cl, BR, I), alkyl of 1 to 6 carbon atoms, alkoxy of 1 to 6 carbon atoms, hydroxy and nitro; a heterocyclic, such as thienyl, furyl, pyranyl, imidazolyl, pyrrolyl, tizolyl, oxazolyl, pyridyl, and pipmidinyl compounds; (2) a derivative of the amino acid selected from (a) acid addition salts of the mo group, obtained from inorganic acids, such as hydrochloric, hydrobromic acids.

I? i-a? *. *. h **? .L *. *. * _- "" - '> --- »* --h-JHii..1.1 sulfuric, nitric, phosphoric and perchloric, as well as organic acids, such as tartaric, citric, acetic, succinic, maleic, oxalic acids; (b) amides of the carboxylic acid group, e.g., glutamine, di-peptides, e.g., salts and esters of oxidized and non-oxidized L-cysteinylglycine, gamma-L-glutamyl-L-cysteine, N-acetyl-L-cysteine-glycine, either conjugated, unconjugated or polymeric forms of L-Gly-L-Glu and L-Val-L-Thr, L-aspartyl-L-phenylalanine, muramyl dipeptides, nutrients such as L-tyrosyl-L-tyrosine, L- alani 1-L-tyrosine, L-arginyl-L-tyrosine, L-tyrosyl-L-arginine, N-Cbz-L-Leu-L-Leu-OCH? its salts or esters, glycyl-glycine, N-acetyl-L-aspartate-L-glutamate (NAAG), etc.; and triproptides, v.gr, oxidized and non-oxidized gamma-L-glutamyl-L-cysteinylglycine; muramyl tripeptides, etc. (c) esters of the carboxylic acid group obtained from straight or branched chain aliphatic alcohols of 1 to 6 carbon atoms, v gr. , L-aspartyl-L-phenylalanine methyl ester (Aspartame (R)), (3) an ether of any of the foregoing; (4) a hydrate or hemihydrate of any of the foregoing, and (5) a mixture of the amino acid and the amino acid derivative. The appropriate amino acids of the formula I include glycine, alanine, valine, leucma, isoleucine, leucylalanine, methionine, threonine, isovaline, y-? ¡.á x. 35 phenylalanine, tyrosine, serine, cysteine, N-acetyl-L-cysteine, histidine, tryptophan, proline, and hydroxyproline, e.g., trans-4-hydroxy proline. The compounds of the formula II include, aspartic acid, and glutamic acid, the compounds of the formula (III) include arginine, glutamine, lysine, hydroxylysine, ornithine, asparagine and citrulline. A fluid or aerosol formulation preferably comprises a protective colloid stabilizer in an amount effective to stabilize the formulation relative to an identical formulation that does not contain the stabilizer, so that the drug does not sediment, acme or flocculate after agitation so quickly as to prevent reproducible dosing of the drug. The reproducible dosage can be achieved if the formulation retains a substantially uniform drug concentration for about fifteen seconds at about five minutes after agitation. For optimal functional and therapeutic performance of the aerosol formulation, either as a dry powder or as an aerosol suspension, the stabilizer is present either as a coarse carrier (e.g., 20-90 um) or as a powder finely micronized ^ < _ 10 um in diameter. In any case, drug dosimetry 'Í A.? ? A, .. L, KÁ.? ., ..J ^ - ,.: ,, .. _____________ti ____________ ..... Ja-feiti. i.

Reproducible is obtained without the need to qualify the patient's inspiration maneuver. Consequently, excellent dose uniformity is obtained at inflows of up to 2 liters, or at inspiratory flow rates of as low as 15 liters per minute at approximately 90 liters per minute. The particular amount of stabilizer constituting an effective amount depends on the stabilizer. Particularly, the particular propellant, and the particular drug used in the formulation Therefore it is not practical to list specific effective amounts for use with specific formulations of the invention, but such amounts can be readily determined by those experienced in the art with the Due consideration of the factors discussed above. Generally, however, the stabilizer may be present in a formulation in an amount of about 0.001 parts per million to about 200,000 parts per million, more preferably about 1 part per million to about 10,000 parts per million, so more preferably from about 10 parts per million to about 5,000 parts per million of the total formulation. It has surprisingly been found that The formulation of the invention is stable without the need to employ a cosolvent, such as ethanol, or surfactants. However, additional components, such as conventional lubricants or surfactants, cosolvents, ethanol, etc., may also be present in an aerosol formulation of the invention in appropriate amounts readily determined by those experienced in the field. to the US Patent No. 5,225,183, which is incorporated by reference herein in its entirety. Generally, the formulations of the invention can be prepared by combining (i) a drug in an amount sufficient to provide a plurality of therapeutically effective doses (n) the stabilizer in an amount effective to stabilize each of the formulations; (iii) the fluid or propellant in an amount sufficient to drive a plurality of doses, v gr. , from an aerosol canister, and (iv) any additional optional components, v gr, ethanol as a cosolvent, and disperse the components. The components can be dispersed using a conventional mixer or homogenizer, by stirring, or by ultrasonic energy. The components can also be dispersed using a ball mill or microfilmizer Volume formulations can be transferred to smaller individual aerosol vials using valve to valve transfer methods, pressure filling, or using conventional cold filling methods. A stabilizer used in a suspension aerosol formulation is not required to be soluble in the propellant. Those which are not sufficiently soluble can be coated on the drug particles in an appropriate amount and the coated particles can then be incorporated into a formulation as described above. Aerosol cans equipped with conventional valves, preferably metered-dose valves, can be used to deliver the formulations of the invention. It has been found, however, that the selection of valve assemblies suitable for use with the aerosol formulations depends on the particular stabilizer and other adjuvants used (if any), on the propellant, and on the particular drug being used. Conventional neoprene and buna valve rubbers used in metered dose valves to deliver conventional CFC formulations frequently have less than optimum valve delivery characteristics and ease of operation when used with formulations containing HFC-134a or HFC-227 Therefore, certain formulations of the invention are preferably delivered through a valve assembly wherein the diaphragm is made of a nitrile rubber such as DB-218 (American Gasket and Rubber, Schiller Park, 111.) or an EPDM rubber such as VistalonMR (Exxon), Royalene "* (UniRoyal, bunaEP (Bayer), Diaphragms made by extrusion injection molding or compression molding of a thermoplastic elastomeric material are also suitable. such as FLEXOMERMR GERS 1085 NT fine polyol (Union Carbide) Conventional aerosol cans, coated or uncoated, anodized or non-anodized, eg, those made of aluminum, glass, stainless steel, polyethylene terephthalate, and coated cans With epon, epoxy, etc., they can be used to contain a formulation of the invention, Conventional nebulizer systems can be used with the formulations of this invention, as well as aerosols and n powder The formulation of the invention can be delivered to the respiratory tract and / or lung by oral inhalation in order to effect bronchodilation or in order to treat a condition susceptible to treatment by inhalation, v. gr, asthma, lung disease Chronic Obstructive The formulations of the invention may also be delivered by nasal inhalation in order to treat, v.gr, allergic rhinitis, rhinitis, (local) or (systom) -diabetes, or may be delivered through topical administration (v.gr , oral) in order to treat, eg, angina or local infection ,: i-Á? A. ±. I- -.

Claims (8)

1. - A medicinal formulation, comprising: (a) a therapeutically effective amount of a protein or porptide medicament having a molecular size ranging from about 1K Dalton to about 150K Daltons; (b) a fluid carrier for containing the medicament; and (c) a stabilizer selected from an amino acid, a derivative thereof, or a mixture thereof.

2. The formulation according to claim 1, wherein the medicament is selected from the group consisting of an insulin, an insulin analogue, an amylin, an immunornodulatory protein, an interleukin, an inteferon, a cryopoietin, a heparin, a thrombolytic, an antitrypsin, an antiprotease, a hormone, a growth factor, an enzyme, a nucleic acid, an immunoglobulin, an antibiotic, an antiinfective, a calcitonin, a hematopoiotic factor, a vaccine, a vasoactive peptide, an antisense agent, an oligonucleotide, DNase, a cyclosprine, ribavirin, or a mixture of any of the above documents. f, i .... k. - i. i, üil-la _ * - »__ A., nor I1? * l II ti-Milíil 'I llJühlTTT i ** •! * - * •'

3. - The formulation according to claim 1, wherein the medicament is selected from the group consisting of an insulin, an insulin analog, an amylin, glucagon, LH-RH, deltirex, leuprolide, gosorelin, nafarelin, octreotide, somatostatin, a calcitonin, parathyroid hormone, TRH, growth hormone-releasing hormone, G-CSF, G-SF, a cytokine, rhDNAse, a heparin, an antibiotic, albumin, ovalbumin, ammoloride, DDAVP, VIP, a cyclosporin, a eri tropoietin, one inferred, IgG, IgE, IgM, IgA, IgD, an interleukin, IRAP, papain, peroxidase, serratio peptidase, catalase, a-1-antitrypsin, a gene; a vector, an amiloride, a rhDNAse, an oligonucleotide, ribavipna, or a mixture of any of the above medicaments

4. The formulation according to claim 1, wherein the stabilizer is selected from the group consisting of twenty amino acids existing, any mixture of them, and any derivative of the above.

5. The formulation according to claim 1, wherein the stabilizer is selected from the group consisting of (1) a di-peptide selected from the group consisting of a salt and an ester of oxidized L-cysteinylglycine. and not rusted, ta - * - Aj¿á? . * 8 * fc «.at-aa. > -J --- .. ^. .... .. -, __ »- gamma-L-glutamyl-L-cysteine, N-acetyl-L-cystma-gli ciña; (2) a conjugated, unconjugated or polymeric form of L-Gly-L-Glu and L-Val-L-Thr; (3) L-aspartyl-L-phenylalanine; (4) a muramyl dipeptide; (5) a nutrient selected from the group consisting of L-tyrosyl-L-tyrosine, L-alanyl-L-tyrosine, L-arginyl-L-tyrosine, L-tyrosyl-L-argmin, N-Cbz-L -Leu-L-Leu-OCH and salts or esters of the previous ones; (6) glycyl glycine; (7) N-acetyl-L-asparate-L-glutamate, (NAAG), (8) a tripeptide selected from the group consisting of an oxidized and a non-oxidized form of gamma-L-glutamyl-L-cysteinylglycine or a muramyl tripeptide; and (9) a mixture of any of the above stabilizers.

6. The formulation according to claim 1, wherein the fluid carrier is a propellant selected from the group consisting of 1,1,1,2-tetrafluoroethane, 1, 1, 2, 3, 3, 3-heptafluoropropane or a mixture thereof.

7. The formulation according to claim 1, wherein the fluid carrier is a hydrocarbon propellant selected from the group consisting of n-butane, propane, isopentane or a mixture thereof. Í. ? .li? .t .. ^ ^ á? L ^ ??? ? á a u ?? á0k - * .. - -, -aA --------.--- ___, ___- & --------- ?.

8. - The formulation according to claim 1, further including a cosolvent 9 - The formulation according to claim 8, wherein the cosolvent comprises ethanol. 10. The formulation according to claim 1, wherein the stabilizer is present in an effective amount to prevent sedimentation, creaming or flocculation of the formulation for a time sufficient to allow reproducible dosing of the drug after agitation of the formulation. 11 - The formulation in accordance with Claim 10, wherein the stabilizer is present in an amount ranging from about 0.001 parts per million to about 200,000 parts per million of the total weight of the formulation 12. A method for preparing a stable, medicinal aerosol formulation. according to claim 1, comprising (a) combining (i) the medicament in an amount sufficient to provide a plurality of therapeutically effective doses, (n) the fluid carrier in an amount sufficient to propel a plurality of the therapeutically effective doses. , and (m) the iXi ?, .Í -Á? ..: stabilizer in an effective amount to stabilize the formulation; and (b) disperse components (i), (ii) and (ii) 13. The method according to claim 12, wherein the medicinal aerosol formulation further comprises combining in step (a) a cosolvent and in step (b) dispersing the components (i), (ii) ), (üi) with the cosolvent 14.- A method for treating in a human or an animal a condition capable of treatment by oral or nasal inhalation, comprising, administering a formulation according to claim 1 to the human or animal by inhalation oral or nasal 15. A formulation according to claim 1 in an aerosol can equipped with a metered dose valve. 16. A method for stabilizing a suspension aerosol formulation, comprising a propellant and a protein or peptide medicament, comprising: incorporating into the formulation a stabilizer selected from the group consisting of an appropriate amino acid, a derivative thereof , or any mixture of the above, in an amount that is effective HBt S i l .i, -; -. ,. "--.--? .. .. - - .. to J». -.-- tJt-a .---. to prevent sedimentation creaming or flocculation of the formulation for a time sufficient to allow reproducible dosing of the drug after agitation of the formulation 17 - A metered dose inhaler containing a medicinal aerosol formulation the formulation comprising (a) a protein or peptide medicament in a therapeutically effective amount (b) a propellant and (c) an appropriate stabilizer selected from an amino acid, an amino acid derivative or a mixture thereof, present in an amount sufficient to stabilize the formulation to prevent sedimentation creaming or flocculation for a time sufficient to allow reproducible dosing of the drug after shaking of the formulation 18 - The metered dose inhaler according to claim 17 wherein the stabilizer is selected from the group consisting of twenty amino acids existing any mixture of any of the above and any derivative of the above