The invention generally relates to a method for pulmonary delivery of therapeutic, prophylactic and diagnostic agents to a patient wherein the agent is released in a sustained fashion, and to particles suitable for use in the method. In particular, the invention relates to a method for the pulmonary delivery of a therapeutic, prophylactic or diagnostic agent comprising administering to the respiratory tract of a patient in need of treatment, prophylaxis or diagnosis an effective amount of particles comprising a polycationic complexing agent which is complexed with a therapeutic, prophylactic or diagnostic agent or any combination thereof having a charge capable of complexing with the polycationic complexing agent upon association with the bioactive agent. The particles can further comprise a pharmaceutically acceptable carrier. The amount of polycationic complexing agent present in the particles is an amount sufficient to sustain the release of diagnostic, therapeutic or prophylactic agent from the particles. For example, the amount of complexing agent present can be at about 5% weight/weight (w/w) or more of the total weight of the complexing agent and therapeutic, diagnostic or prophylactic agent. Release of the agent from the administered particles occurs in a sustained fashion.

DETAILED DESCRIPTION OF THE INVENTION

A description of preferred embodiments of the invention follows.

Therapeutic, prophylactic or diagnostic agents, can also be referred to herein as "bioactive agents", "medicaments" or "drugs".

The invention relates to a method for the pulmonary delivery of therapeutic, prophylactic and diagnostic agents comprising administering to the respiratory tract of a patient in need of treatment, prophylaxis or diagnosis an effective amount of particles comprising a polycationic complexing agent which is complexed with a therapeutic, prophylactic or diagnostic agent or any combination thereof having a charge which permits complexation with the polycationic complexing agent upon association with the bioactive agent. The particles can further comprise a pharmaceutically acceptable carrier. The amount of polycationic complexing agent present in the particles is an amount sufficient to sustain the release of therapeutic, prophylactic or a diagnostic agent from the particles. For example, the amount of complexing agent present in the particles can be about 5% weight/weight (w/w) or more of the total weight of the complexing agent and the therapeutic, prophylactic or diagnostic agent. Release of the agent from the administered particles occurs in a sustained fashion. In a particular embodiment, the particles can be in the form of a dry powder.

The particles of the invention release bioactive agent in a sustained fashion. As such, the particles possess sustained release properties. "Sustained release", as that term is used herein, refers to a release of active agent in which the period of release of an effective level of agent is longer than that seen with the same bioactive agent which is not complexed with a polycationic complexing agent, prior to administration. In addition, a sustained release can also refer to a reduction in the burst of agent typically seen in the first two hours following administration, and more preferably in the first hour, often referred to as the "initial burst". In a preferred embodiment, the sustained release is characterized by both the period of release being longer in addition to a decreased initial burst. For example, a sustained release of insulin can be a release showing elevated serum levels of insulin at least 4 hours post administration, such as about 6 hours or more.

"Pulmonary delivery", as that term is used herein refers to delivery to the respiratory tract. The "respiratory tract", as defined herein, encompasses the upper airways, including the oropharynx and larynx, followed by the lower airways, which include the trachea followed by bifurcations into the bronchi and bronchioli (e.g., terminal and respiratory). The upper and lower airways are called the conducting airways. The terminal bronchioli then divide into respiratory bronchioli which then lead to the ultimate respiratory zone, namely, the alveoli, or deep lung. The deep lung, or alveoli, are typically the desired target of inhaled therapeutic formulations for systemic drug delivery.

Complexation of the polycationic complexing agent with the therapeutic, prophylactic or diagnostic agent can result from ionic complexation, salt bridge formation, charge-charge interaction or a combination thereof.

In a particular embodiment, complexation of the therapeutic, prophylactic or diagnostic agent and the polycationic complexing agent can be a result of ionic complexation or bonding.

The particles suitable for use in the method can comprise a therapeutic, prophylactic or diagnostic agent which is complexed with a polycationic complexing agent wherein the charge of the bioactive agent is such that it is able to undergo complexation with the polycationic complexing agent upon association, prior to administration.

For example, the particles suitable for pulmonary delivery can comprise a therapeutic, prophylactic or diagnostic agent which possesses an overall net negative charge at the time of complexation with the polycationic complexing agent. For example, the agent can be insulin and the polycationic complexing agent can be protamine.

"Pulmonary pH range", as that term is used herein, refers to the pH range which can be encountered in the lung of a patient. Typically, in humans, this range of pH is from about 6.4 to about 7.0, such as from 6.4 to about 6.7. pH values of the airway lining fluid (ALF) have been reported in "Comparative Biology of the Normal Lung", CRC Press, (1991) by R. A. Parent and range from 6.44 to 6.74).

The term polycationic complexing agent, as used herein, refers to an agent which has two or more cationic sites and is capable of complexing, for example, by ionic complexation with an active agent of opposite charge. Suitable polycationic complexing agents include, but are not limited to, protamine, spermine, spermidine, chitosan and a polycationic polyamino acid. A polycationic polyamino acid can be a homopolymer of a cationic amino acid such as polylysine or polyarginine or a random copolymer of cationic and non-cationic amino acids with the cationic amino acid present in an amount sufficient to impart cationic change characteristics to the random copolymer. For example, a polycationic polyamino acid such as polylysine or polyarginine is a polycationic polyamino acid homopolymer. Such, homopolymers can be commercially obtained in varying molecular weight ranges. For example, polylysine can be purchased from Sigma in molecular weights ranging from 1000 to about 300,000. Further, random copolymers containing lysine or arginine in sufficient amounts to render the resulting copolymer cationic can also be purchased from Sigma. Examples of such random copolymers include, but are not limited to, polylysine-alanine at ratios of 1:1, 2:1 or 3:1 and molecular weight ranges from 20,000 to 50,000. The amount of polycationic complexing agent present in the particles is an amount sufficient to sustain the release of therapeutic, prophylactic or diagnostic agent from the particles. For example, the amount of complexing agent present in the particles can be about 5% weight/weight (w/w) or more of the total weight of the complexing agent and the therapeutic prophylactic or diagnostic agent.

The interaction, for example, complexation of the polycationic complexing agent with the bioactive agent of opposite charge can be achieved by associating, for example, mixing the bioactive agent in a suitable aqueous solvent or cosolvent with at least one suitable polycationic complexing agent under pH conditions suitable for complexation of the polycationic complexing agent and the bioactive agent. Typically, the polycationic-complexed active agent will be in the form of a precipitate. Preferably, the precipitated polycationic-complexed active agent remains in the solid state throughout the process used to obtain the final particles for administration. In a preferred embodiment, the bioactive agent is complexed with protamine. Most preferably, the protamine is complexed to insulin.

Suitable pH conditions to obtain complexation of a polycationic complexing agent with a bioactive agent can be determined based on the pKa of the bioactive agent and the charge characteristics of the polycationic complexing agent. That is, the pH of the system wherein complexation takes place should be adjusted based on the pKa of the active agent and the charge characteristics of the polycationic complexing agent in order to impart a negative charge on the active agent and polycationic characteristics to the complexing agent. Suitable pH conditions are typically achieved through use of an aqueous buffer system as the solvent (e.g., citrate, phosphate, acetate, etc.). Adjustment to the desired pH can be achieved with addition of an acid or base as appropriate. Suitable solvents are those in which the bioactive agent and the polycationic complexing agent are each at least slightly soluble. For example, sodium citrate, acetate, and phosphate buffers.

For example, employing a protein as the active agent, the agent may be mixed with the polycationic complexing agent in a buffer system wherein the protein has a negative charge. Specifically, insulin, for example, may be mixed with the desired polycationic complexing agent in an aqueous buffer system (e.g. citrate, phosphate, acetate, etc.), the pH of the resultant solution then can be adjusted to a desired value using an appropriate base solution (e.g., 1 N NaOH). That is, the pH of the insulin and polycationic complexing agent mixture can be adjusted to about pH 6.7. At this pH insulin molecules have a net negative charge (pI is about 5.5) and the complexing agent should be positively charged resulting in complexation of the polycationic complexing agent to the insulin typically achieving a precipitate of the polycationic complexed insulin.

The polycation complexed bioactive agent can then, if desired, be mixed with a pharmaceutically acceptable carrier. Typically, the solution containing the precipitated polycationic complexed biologically active agent is mixed with a solution of the pharmaceutically acceptable carrier. Suitable pharmaceutically acceptable carriers and appropriate solvent systems for use with same are provided in detail below. The solvent is then removed from the resulting mixture. Solvent removal techniques include, for example, lyophilization, evaporation and spray drying. Spray drying of the resulting mixture is a preferred method of preparing the particles of the invention. Specific spray drying processes are discussed in detail below. It is preferred that the solid polycationic complexed biologically active agent remains in solid form throughout the processing of the final particles in the method described herein administered.

The total amount of polycationic complexing agent present in the particles of the invention is an amount sufficient to sustain the release of therapeutic, prophylactic or diagnostic agent from the particles. For example, the amount of complexing agent present in the particles can be about 5% weight/weight (w/w) or more of the total weight of the complexing agent and the therapeutic, prophylactic or diagnostic agent, such as, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, etc. For example, the ratio of polycationic complexing agent to bioactive agent present in the combined weight of the complexing and active agent of the particles of the invention can be about 5% w/w or more and range from about 5% w/w to about 10% w/w, or from about 10% w/w to about 30% w/w etc. It is understood that the upper limit of polycationic complexing agent present depends upon the tolerance of the formulation by the recipient. For example, the formulation can have the polycationic complexing agent present at about 50% or more by weight of the total weight of the complexing agent and the therapeutic, prophylactic or diagnostic agent.

The particles of the invention, can when desired, further comprise a multivalent metal cation. "Multivalent metal cation" as that term is used herein, refers to metal cations which possess a valency of +2 or more. The multivalent metal cation can be chosen to have a charge opposite to that of the active agent when the multivalent metal cation and active agent are associated. Combinations of multivalent metal cation can be used.

Suitable multivalent metal cations include, but are not limited to, biocompatible multivalent metal cations.

It is understood that the multivalent metal cations suitable for complexation with an active agent of opposite charge can be any of the transition state metals of the periodic table, and the non-transition state metals, for example, calcium (Ca), zinc (Zn), cadmium (Cd), mercury (Hg), strontium (Sr), and barium (Ba). Divalent metal cations are preferred, such as, Zn(II), Ca(II), Cu(II), Mg(II), Ni(II), Co(II), Fe(II), Ag(II), Mn(II) or Cd(II).

The metal cation can be complexed with the bioactive agent using the conditions described above for complexation with the polycationic complexing agent. The amount of multivalent metal cation includes both multivalent metal cation which is complexed with the biologically active agent, as well as any multivalent metal cation which is present but not complexed with the biologically active agent. For example, the multivalent metal cation which is not associated with the active agent can be present, for example, as the metal cation of a metal cation-containing component, such as a multivalent metal cation-containing salt.

Suitable multivalent metal cation-containing components include, but are not limited to, salts having the multivalent metal cations described above and a suitable pharmaceutically acceptable counterion. The counterion can be, for example, chloride, bromide, citrate, tartrate, lactate, methanesulfonate, acetate, sulfonate formate, maleate, fumarate, malate, succinate, malonate, sulfate, phosphate, hydrosulfate, pyruvate, mucate, benzoate, glucuronate, oxalate, ascorbate, the conjugate base of a fatty acid (e.g., oleate, laurate, myristate, stearate, arachidate, behenate, arachidonate) and combinations thereof.

The particles of the invention can, when desired, further comprise a pharmaceutically acceptable carrier. Suitable pharmaceutically acceptable carriers can be chosen, for example, based on achieving particles having the desired characteristics for inhalation to the area of the respiratory tract where delivery is needed and therapeutic action is achieved. Pharmaceutically acceptable carriers suitable for use in the invention include, but are not limited to, phospholipids, sugars and polysaccharides, such as maltodextrin.

In a preferred embodiment of the invention, the pharmaceutically acceptable carrier of the particles is a phospholipid . Examples of suitable phospholipids include, among others, phosphatidic acids, phosphatidylcholines, phosphatidylalkanolamines such as a phosphatidylethanolamines, phosphatidylglycerols, phosphatidylserines, phosphatidylinositols and combinations thereof.

Specific examples of phospholipids include, 1,2-diacyl-sn-glycero-3-phosphocholine and a 1,2-diacyl-sn-glycero-3-phosphoalkanolamine phospholipids. Suitable examples of 1,2-diacyl-sn-glycero-3-phosphocholine phospholipids include, but are not limited to, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dilaureoyl-sn-3-glycero-phosphocholine (DLPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC).

Suitable examples of 1,2-diacyl-sn-glycero-3-phosphoalkanolamine phospholipids include, but are not limited to, 1,2-dipalmitoyl-sn-glycero-3-ethanolamine(DPPE), 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine(DMPE), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine(DSPE), 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine (DLPE), and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE).

Other classes of phospholipids suitable for use in the invention as a pharmaceutically acceptable carrier include 1,2-diacyl-sn-glycero-3-alkylphosphocholines and 1,2-diacyl-sn-glycero-3-alkylphosphoalkanolamines.

Specific examples of 1,2-diacyl-sn-glycero-3-alkylphosphocholine phospholipids include, but are not limited to, 1,2-dipalmitoyl-sn-glycero-3-ethylphosphocholine(DPePC), 1,2-dimyristoyl-sn-glycero-3-ethylphosphocholine(DMePC), 1,2-distearoyl-sn-glycero-3-ethylphosphocholine(DSePC), 1,2-dilauroyl-sn-glycero-3-ethylphosphocholine (DLePC), and 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine(DOePC).

Specific examples of 1,2-diacyl-sn-glycero-3-alkylphosphoalkanolamines include, but are not limited to 1,2-dipalmitoyl-sn-glycero-3-ethylethanolamine(DPePE), 1,2-dimyristoyl-sn-glycero-3-ethylphosphoethanolamine(DMePE), 1,2-distearoyl-sn-glycero-3-ethylphosphoethanolamine(DSePE), 1,2-dilauroyl-sn-glycero-3-ethylphosphoethanolamine (DLePE), and 1,2-dioleoyl-sn-glycero-3-ethylphosphoethanolamine (DOePE).

Other phospholipids are known to those skilled in the art and are described in U.S. patent application Ser. No. 09/752,109 entitled "Particles for Inhalation Having Sustained Release Properties" filed on Dec. 29, 2000 and U.S. patent application Ser. No. 09/752,106 entitled "Particles for Inhalation Having Sustained release Poperties" filed on Dec. 29, 2000 the contents of all of which are incorporated herein in their entirety. In a preferred embodiment, the phospholipids are endogenous to the lung.

The phospholipid, can be present in the particles in an amount ranging from about 0 to about 90 weight %. More commonly it can be present in the particles in an amount ranging from about 10 to about 60 weight %.

In another embodiment of the invention, the phospholipids or combinations thereof are selected to impart sustained release properties to the highly dispersible particles. The phase transition temperature of a specific phospholipid can be below, around or above the physiological body temperature of a patient. Preferred phase transition temperatures range from 30.degree. C. to 50.degree. C., (e.g., within .+-.10 degrees of the normal body temperature of patient). By selecting phospholipids or combinations of phospholipids according to their phase transition temperature, the particles can be tailored to have sustained release properties. For example, by administering particles which include a phospholipid or combination of phospholipids which have a phase transition temperature higher than the patient's body temperature, the release of active agent can be slowed down. On the other hand, rapid release can be obtained by including in the particles phospholipids having lower transition temperatures. Particles having sustained release properties and methods of modulating release of a biologically active agent are described in U.S. patent application Ser. No. 09/644,736 entitled Modulation of Release From Dry Powder Formulations by Controlling Matrix Transition, filed on Aug. 23, 2000, the entire contents of which are incorporated herein by reference.

Therapeutic, prophylactic or diagnostic agents, can also be referred to herein as "bioactive agents", "medicaments" or "drugs". It is understood that one or more bioactive agents can be present in the particles of the invention. Hydrophilic as well as hydrophobic agents can be used. The agent must be capable of possessing a charge which allows it to undergo complexation with the polycationic complexing agent.

The amount of bioactive agent present in the particles of the invention can be from about 0.1 weight % to about 95 weight %. For example, from about 1 to about 50%, such as from about 5 to about 30%. Particles in which the drug is distributed throughout a particle are preferred.

Suitable bioactive agents include agents which can act locally, systemically or a combination thereof. The term "bioactive agent," as used herein, is an agent, or its pharmaceutically acceptable salt, which when released in vivo, possesses the desired biological activity, for example therapeutic, diagnostic and/or prophylactic properties in vivo.

Examples of bioactive agent include, but are not limited to, proteins and peptides, polysaccharides and other sugars, lipids, and DNA and RNA nucleic acid sequences having therapeutic, prophylactic or diagnostic activities. Agents with a wide range of molecular weight can be used.

The agents can have a variety of biological activities, such as vasoactive agents, neuroactive agents, hormones, anticoagulants, immunomodulating agents, cytotoxic agents, prophylactic agents, diagnostic agents, antibiotics, antivirals, antisense, antigens, antineoplastic agents and antibodies.

Proteins, include complete proteins, muteins and active fragments thereof, such as insulin, immunoglobulins, antibodies, cytokines (e.g., lymphokines, monokines, chemokines), interleukins, interferons (.beta.-IFN, .alpha.-IFN and .gamma.-IFN), erythropoietin, somatostatin, nucleases, tumor necrosis factor, colony stimulating factors, enzymes (e.g. superoxide dismutase, tissue plasminogen activator), tumor suppressors, blood proteins, hormones and hormone analogs (e.g., growth hormone, such as human growth hormone (hGH)), adrenocorticotropic hormone and luteinizing hormone releasing hormone (LHRH)), vaccines (e.g., tumoral, bacterial and viral antigens), antigens, blood coagulation factors; growth factors; granulocyte colony-stimulating factor (G-CSF); peptides include parathyroid hormone-related peptide, protein inhibitors, protein antagonists, and protein agonists, calcitonin; nucleic acids include, for example, antisense molecules, oligonucleotides, and ribozymes. Polysaccharides, such as heparin, can also be administered.

Bioactive agents for local delivery within the lung, include agents such as those for the treatment of asthma, chronic obstructive pulmonary disease (COPD), emphysema, or cystic fibrosis. For example, genes for the treatment of diseases such as cystic fibrosis can be administered, as can beta agonists, steroids, anticholinergics, and leukotriene modifiers for asthma.

Nucleic acid sequences include genes, oligonucleotides, antisense molecules which can, for instance, bind to complementary DNA to inhibit transcription, and ribozymes.

The particles can further comprise a carboxylic acid which is distinct from the polycation complexed biologically active agent. In one embodiment, the carboxylic acid includes at least two carboxyl groups. Carboxylic acids include the salts thereof as well as combinations of two or more carboxylic acids and/or salts thereof. In a preferred embodiment, the carboxylic acid is a hydrophilic carboxylic acid or salt thereof. Suitable carboxylic acids include but are not limited to hydroxydicarboxylic acids, hydroxytricarboxylic acids and the like. Citric acid and citrates, such as, for example sodium citrate, are preferred. Combinations or mixtures of carboxylic acids and/or their salts also can be employed.

The carboxylic acid can be present in the particles in an amount ranging from about 0 to about 80% weight. Preferably, the carboxylic acid can be present in the particles in an amount of about 10 to about 20%.

The particles suitable for use in the invention can further comprise an amino acid. In a preferred embodiment the amino acid is hydrophobic. Suitable naturally occurring hydrophobic amino acids, include but are not limited to, leucine, isoleucine, alanine, valine, phenylalanine, glycine and tryptophan. Combinations of hydrophobic amino acids can also be employed. Non-naturally occurring amino acids include, for example, beta-amino acids. Both D, L configurations and racemic mixtures of hydrophobic amino acids can be employed. Suitable hydrophobic amino acids can also include amino acid derivatives or analogs. As used herein, an amino acid analog includes the D or L configuration of an amino acid having the following formula: --NH--CHR--CO--, wherein R is an aliphatic group, a substituted aliphatic group, a benzyl group, a substituted benzyl group, an aromatic group or a substituted aromatic group and wherein R does not correspond to the side chain of a naturally-occurring amino acid. As used herein, aliphatic groups include straight chained, branched or cyclic C1 C8 hydrocarbons which are completely saturated, which contain one or two heteroatoms such as nitrogen, oxygen or sulfur and/or which contain one or more units of unsaturation. Aromatic or aryl groups include carbocyclic aromatic groups such as phenyl and naphthyl and heterocyclic aromatic groups such as imidazolyl, indolyl, thienyl, furanyl, pyridyl, pyranyl, oxazolyl, benzothienyl, benzofuranyl, quinolinyl, isoquinolinyl and acridintyl.

Suitable substituents on an aliphatic, aromatic or benzyl group include --OH, halogen (--Br, --Cl, --I and --F) --O(aliphatic, substituted aliphatic, benzyl, substituted benzyl, aryl or substituted aryl group), --CN, --NO.sub.2, --COOH, --NH.sub.2, --NH(aliphatic group, substituted aliphatic, benzyl, substituted benzyl, aryl or substituted aryl group), --N(aliphatic group, substituted aliphatic, benzyl, substituted benzyl, aryl or substituted aryl group).sub.2, --COO(aliphatic group, substituted aliphatic, benzyl, substituted benzyl, aryl or substituted aryl group), --CONH.sub.2, --CONH(aliphatic, substituted aliphatic group, benzyl, substituted benzyl, aryl or substituted aryl group)), --SH, --S(aliphatic, substituted aliphatic, benzyl, substituted benzyl, aromatic or substituted aromatic group) and --NH-- C(.dbd.NH)--NH.sub.2. A substituted benzylic or aromatic group can also have an aliphatic or substituted aliphatic group as a substituent. A substituted aliphatic group can also have a benzyl, substituted benzyl, aryl or substituted aryl group as a substituent. A substituted aliphatic, substituted aromatic or substituted benzyl group can have one or more substituents. Modifying an amino acid substituent can increase, for example, the lipophilicity or hydrophobicity of natural amino acids which are hydrophilic.

A number of the suitable amino acids, amino acid analogs and salts thereof can be obtained commercially. Others can be synthesized by methods known in the art. Synthetic techniques are described, for example, in Green and Wuts, "Protecting Groups in Organic Synthesis", John Wiley and Sons, Chapters 5 and 7, 1991.

Hydrophobicity is generally defined with respect to the partition of an amino acid between a nonpolar solvent and water. Hydrophobic amino acids are those acids which show a preference for the nonpolar solvent. Relative hydrophobicity of amino acids can be expressed on a hydrophobicity scale on which glycine has the value 0.5. On such a scale, amino acids which have a preference for water have values below 0.5 and those that have a preference for nonpolar solvents have a value above 0.5. As used herein, the term hydrophobic amino acid refers to an amino acid that, on the hydrophobicity scale has a value greater or equal to 0.5, in other words, has a tendency to partition in the nonpolar acid which is at least equal to that of glycine.

Examples of amino acids which can be employed include, but are not limited to: glycine, proline, alanine, cysteine, methionine, valine, leucine, tyrosine, isoleucine, phenylalanine, tryptophan. Preferred hydrophobic amino acids include leucine, isoleucine, alanine, valine, phenylalanine, glycine and tryptophan. Combinations of hydrophobic amino acids can also be employed. Furthermore, combinations of hydrophobic and hydrophilic (preferentially partitioning in water) amino acids, where the overall combination is hydrophobic, can also be employed. Combinations of one or more amino acids can also be employed.

The amino acid can be present in the particles of the invention in an amount of from about 0% to about 60 weight %. Preferably, the amino acid can be present in the particles in an amount ranging from about 5 to about 30 weight %. The salt of a hydrophobic amino acid can be present in the particles of the invention in an amount of from about 0% to about 60 weight %. Preferably, the amino acid salt is present in the particles in an amount ranging from about 5 to about 30 weight %. Methods of forming and delivering particles which include an amino acid are described in U.S. patent application Ser. No. 09/382,959, filed on Aug. 25, 1999, entitled "Use of Simple Amino Acids to Form Porous Particles During Spray Drying" the entire teaching of which is incorporated herein by reference.

In a further embodiment, the particles can also include other excipients such as, for example, buffer salts, dextran, polysaccharides, lactose, trehalose, cyclodextrins, proteins, peptides, polypeptides, fatty acids, fatty acid esters, inorganic compounds, phosphates.

In one embodiment of the invention, the particles can further comprise polymers. Biocompatible or biodegradable polymers are preferred. Such polymers are described, for example, in U.S. Pat. No. 5,874,064, issued on Feb. 23, 1999 to Edwards et al., the teachings of which are incorporated herein by reference in their entirety.

In yet another embodiment, the particles include a surfactant other than the phospholipids described above. As used herein, the term "surfactant" refers to any agent which preferentially absorbs to an interface between two immiscible phases, such as the interface between water and an organic polymer solution, a water/air interface or organic solvent/air interface. Surfactants generally possess a hydrophilic moiety and a lipophilic moiety, such that, upon absorbing to particles, they tend to present moieties to the external environment that do not attract similarly-coated particles, thus reducing particle agglomeration. Surfactants may also promote absorption of a therapeutic or diagnostic agent and increase bioavailability of the agent.

Suitable surfactants which can be employed in fabricating the particles of the invention include but are not limited to hexadecanol; fatty alcohols such as polyethylene glycol (PEG); polyoxyethylene-9-lauryl ether; a surface active fatty acid, such as palmitic acid or oleic acid; glycocholate; surfactin; a poloxamer; a sorbitan fatty acid ester such as sorbitan trioleate (Span 85); Tween 80 and tyloxapol.

The surfactant can be present in the particles in an amount ranging from about 0 to about 5 weight %. Preferably, it can be present in the particles in an amount ranging from about 0.1 to about 1.0 weight %.

It is understood that when the particles include a carboxylic acid, an amino acid, a surfactant or any combination thereof, interaction between these components of the particle and the polycationic complexing agent can occur.

The particles, also referred to herein as powder, can be in the form of a dry powder suitable for inhalation. In a particular embodiment, the particles can have a tap density of less than about 0.4 g/cm.sup.3. Particles which have a tap density of less than about 0.4 g/cm.sup.3 are referred to herein as "aerodynamically light particles". More preferred are particles having a tap density less than about 0.1 g/cm.sup.3.

Aerodynamically light particles have a preferred size, e.g., a volume median geometric diameter (VMGD) of at least about 5 microns (.mu.m). In one embodiment, the VMGD is from about 5 .mu.m to about 30 .mu.m. In another embodiment of the invention, the particles have a VMGD ranging from about 9 .mu.m to about 30 .mu.m. In other embodiments, the particles have a median diameter, mass median diameter (MMD), a mass median envelope diameter (MMED) or a mass median geometric diameter (MMGD) of at least 5 .mu.m, for example from about 5 .mu.m to about 30 .mu.m.

Aerodynamically light particles preferably have "mass median aerodynamic diameter" (MMAD), also referred to herein as "aerodynamic diameter", between about 1 .mu.m and about 5 .mu.m. In one embodiment of the invention, the MMAD is between about 1 .mu.m and about 3 .mu.m. In another embodiment, the MMAD is between about 3 .mu.m and about 5 .mu.m.

In another embodiment of the invention, the particles have an envelope mass density, also referred to herein as "mass density" of less than about 0.4 g/cm.sup.3. The envelope mass density of an isotropic particle is defined as the mass of the particle divided by the minimum sphere envelope volume within which it can be enclosed.

Tap density can be measured by using instruments known to those skilled in the art such as the Dual Platform Microprocessor Controlled Tap Density Tester (Vankel, N.C.) or a GeoPyc.TM. instrument (Micrometrics Instrument Corp., Norcross, Ga. 30093). Tap density is a standard measure of the envelope mass density. Tap density can be determined using the method of USP Bulk Density and Tapped Density, United States Pharmacopia convention, Rockville, Md., 10.sup.th Supplement, 4950 4951, 1999. Features which can contribute to low tap density include irregular surface texture and porous structure.

The diameter of the particles, for example, their VMGD, can be measured using an electrical zone sensing instrument such as a Multisizer IIe, (Coulter Electronic, Luton, Beds, England), or a laser diffraction instrument (for example Helos, manufactured by Sympatec, Princeton, N.J.). Other instruments for measuring particle diameter are well known in the art. The diameter of particles in a sample will range depending upon factors such as particle composition and methods of synthesis. The distribution of size of particles in a sample can be selected to permit optimal deposition within targeted sites within the respiratory tract.

Experimentally, aerodynamic diameter can be determined by employing a gravitational settling method, whereby the time for an ensemble of particles to settle a certain distance is used to determine the aerodynamic diameter of the particles. An indirect method for measuring the mass median aerodynamic diameter (MMAD) is the multi-stage liquid impinger (MSLI). Specific instruments which can be employed to determine aerodynamic diameters include those known under the name of Aerosizer.TM. (TSI, Inc., Amherst, Mass.) or under the name of Anderson Cascade Impactor (Anderson Inst., Sunyra, Ga.).

The aerodynamic diameter, d.sub.aer, can be calculated from the equation:

.times. .rho. ##EQU00001## where d.sub.g is the geometric diameter, for example the MMGD and .rho. is the powder density.

Particles which have a tap density less than about 0.4 g/cm.sup.3, median diameters of at least about 5 .mu.m, and an aerodynamic diameter of between about 1 .mu.m and about 5 .mu.m, preferably between about 1 .mu.m and about 3 .mu.m, are more capable of escaping inertial and gravitational deposition in the oropharyngeal region, and are targeted to the airways or the deep lung. The use of larger, more porous particles is advantageous since they are able to aerosolize more efficiently than smaller, denser aerosol particles such as those currently used for inhalation therapies.

In comparison to smaller particles the larger aerodynamically light particles, preferably having a VMGD of at least about 5 .mu.m, also can potentially more successfully avoid phagocytic engulfment by alveolar macrophages and clearance from the lungs, due to size exclusion of the particles from the phagocytes' cytosolic space. Phagocytosis of particles by alveolar macrophages diminishes precipitously as particle diameter increases beyond about 3 .mu.m. Kawaguchi, H., et al., Biomaterials 7: 61 66 (1986); Krenis, L. J. and Strauss, B., Proc. Soc. Exp. Med., 107: 748 750 (1961); and Rudt, S. and Muller, R. H., J. Contr. Rel., 22: 263 272 (1992). For particles of statistically isotropic shape, such as spheres with rough surfaces, the particle envelope volume is approximately equivalent to the volume of cytosolic space required within a macrophage for complete particle phagocytosis.

The particles may be fabricated with the appropriate material, surface roughness, diameter and tap density for localized delivery to selected regions of the respiratory tract such as the deep lung or upper or central airways. For example, higher density or larger particles may be used for upper airway delivery, or a mixture of varying sized particles in a sample, provided with the same or different therapeutic agent may be administered to target different regions of the lung in one administration. Particles having an aerodynamic diameter ranging from about 3 to about 5 .mu.m are preferred for delivery to the central and upper airways. Particles having an aerodynamic diameter ranging from about 1 to about 3 .mu.m are preferred for delivery to the deep lung.

Inertial impaction and gravitational settling of aerosols are predominant deposition mechanisms in the airways and acini of the lungs during normal breathing conditions. Edwards, D. A., J. Aerosol Sci., 26: 293 317 (1995). The importance of both deposition mechanisms increases in proportion to the mass of aerosols and not to particle (or envelope) volume. Since the site of aerosol deposition in the lungs is determined by the mass of the aerosol (at least for particles of mean aerodynamic diameter greater than approximately 1 .mu.m), diminishing the tap density by increasing particle surface irregularities and particle porosity permits the delivery of larger particle envelope volumes into the lungs, all other physical parameters being equal.

The low tap density particles have a small aerodynamic diameter in comparison to the actual envelope sphere diameter. The aerodynamic diameter, d.sub.aer, is related to the envelope sphere diameter, d (Gonda, I., "Physico-chemical principles in aerosol delivery," in Topics in Pharmaceutical Sciences 1991 (eds. D. J. A. Crommelin and K. K. Midha), pp. 95 117, Stuttgart: Medpharm Scientific Publishers, 1992)), by the formula:

.times. .rho. ##EQU00002## where the envelope mass .rho. is in units of g/cm.sup.3. Maximal deposition of monodispersed aerosol particles in the alveolar region of the human lung (.about.60%) occurs for an aerodynamic diameter of approximately d.sub.aer=3 .mu.m. Heyder, J. et al., J. Aerosol Sci., 17: 811 825 (1986). Due to their small envelope mass density, the actual diameter d of aerodynamically light particles comprising a monodisperse inhaled powder that will exhibit maximum deep-lung deposition is:

.times..times. .rho..mu..times..times..function..times..times..rho.<.times..times..ti- mes..times. ##EQU00003## where d is always greater than 3 .mu.m. For example, aerodynamically light particles that display an envelope mass density, .rho.=0.1 g/cm.sup.3, will exhibit a maximum deposition for particles having envelope diameters as large as 9.5 .mu.m. The increased particle size diminishes interparticle adhesion forces. Visser, J., Powder Technology, 58: 1 10. Thus, large particle size increases efficiency of aerosolization to the deep lung for particles of low envelope mass density, in addition to contributing to lower phagocytic losses.

The aerodyanamic diameter can be calculated to provide for maximum deposition within the lungs, previously achieved by the use of very small particles of less than about five microns in diameter, preferably between about one and about three microns, which are then subject to phagocytosis. Selection of particles which have a larger diameter, but which are sufficiently light (hence the characterization "aerodynamically light"), results in an equivalent delivery to the lungs, but the larger size particles are not phagocytosed. Improved delivery can be obtained by using particles with a rough or uneven surface relative to those with a smooth surface.

Suitable particles can be fabricated or separated, for example by filtration or centrifugation, to provide a particle sample with a preselected size distribution. For example, greater than about 30%, 50%, 70%, or 80% of the particles in a sample can have a diameter within a selected range of at least about 5 .mu.m. The selected range within which a certain percentage of the particles must fall may be for example, between about 5 and about 30 .mu.m, or optimally between about 5 and about 15 .mu.m. In one preferred embodiment, at least a portion of the particles have a diameter between about 6 and about 11 .mu.m. Optionally, the particle sample also can be fabricated wherein at least about 90%, or optionally about 95% or about 99%, have a diameter within the selected range. The presence of the higher proportion of the aerodynamically light, larger diameter particles in the particle sample enhances the delivery of therapeutic or diagnostic agents incorporated therein to the deep lung. Large diameter particles generally mean particles having a median geometric diameter of at least about 5 .mu.m.

The particles can be prepared by spray drying. For example, a spray drying mixture, also referred to herein as "feed solution" or "feed mixture", which includes the bioactive agent in association with a polycationic complexing agent, for example, complexed and a pharmaceutically acceptable carrier are fed to a spray dryer.

For example, complexation of the polycationic complexing agent with the bioactive agent of opposite charge can be achieved by mixing the bioactive agent in a suitable aqueous solvent with at least one suitable polycationic complexing agent under pH conditions suitable for forming a complex of the polycationic complexing agent and bioactive agent. Typically, the polycation-complexed active agent will be in the form of a precipitate. Preferably, the precipitated polycation-complexed active agent remains in the solid state throughout the process used to obtain the final particles for administration. In a prefered embodiment, the bioactive agent is complexed with protamine. Most preferably, the protamine is complexed to insulin.

Suitable pH conditions to form a polycation complexed bioactive agent can be determined based on the pKa of the bioactive agent. That is, the pH of the system wherein complexation takes place should be adjusted based on the pKa of the active agent in order to impart a negative charge on the active agent. Suitable pH conditions are typically achieved through use of an aqueous buffer system as the solvent (e.g., citrate, phosphate, acetate, etc.). Adjustment to the desired pH can be achieved with addition of an acid or base as appropriate. Suitable solvents are those in which the bioactive agent and the polycationic complexing are each at least slightly soluble. For example, sodium citrate, acetate, and phosphate buffer systems.

The polycation complexed bioactive agent can, if desired, be further mixed with a pharmaceutically acceptable carrier, as described above or immediately processed into particles for administration without a pharmaceutically acceptable carrier. Suitable organic solvents can be used to form a solution of the pharmaceutically acceptable carrier. Alternatively an aqueous solvent can be used to solubilize the carrier or a combination of aqueous and organic solvent can be employed. Suitable organic solvents include, but are not limited to, alcohols for example, ethanol, methanol, propanol, isopropanol, butanols, and others. Other organic solvents include, but are not limited to, perfluorocarbons, dichloromethane, chloroform, ether, ethyl acetate, methyl tert-butyl ether and others. Aqueous solvents that can be present in the feed mixture include water and buffered solutions. Both organic and aqueous solvents can be present in the spray-drying mixture fed to the spray dryer. In one embodiment, an ethanol water solvent is preferred with the ethanol:water ratio ranging from about 50:50 to about 90:10. The mixture can have a neutral, acidic or alkaline pH. Optionally, a pH buffer can be included. Preferably, the pH can range from about 3 to about 10.

The total amount of solvent or solvents being employed in the mixture being spray dried generally is greater than 99 weight percent. The amount of solids (drug, charged lipid and other ingredients) present in the mixture being spray dried generally is less than about 1.0 weight percent. Preferably, the amount of solids in the mixture being spray dried ranges from about 0.05% to about 0.5% by weight.

Using a mixture which includes an organic and an aqueous solvent in the spray drying process allows for the combination of hydrophilic and hydrophobic components, while not requiring the formation of liposomes or other structures or complexes to facilitate solubilization of the combination of such components within the particles.

Suitable spray-drying techniques are described, for example, by K. Masters in "Spray Drying Handbook", John Wiley & Sons, New York, 1984. Generally, during spray-drying, heat from a hot gas such as heated air or nitrogen is used to evaporate the solvent from droplets formed by atomizing a continuous liquid feed. Other spray-drying techniques are well known to those skilled in the art. In a preferred embodiment, a rotary atomizer is employed. An example of a suitable spray dryer using rotary atomization includes the Mobile Minor spray dryer, manufactured by Niro, Inc., Denmark. The hot gas can be, for example, air, nitrogen or argon.

Preferably, the particles of the invention are obtained by spray drying using an inlet temperature between about 100.degree. C. and about 400.degree. C. and an outlet temperature between about 50.degree. C. and about 130.degree. C.

The spray dried particles can be fabricated with a rough surface texture to reduce particle agglomeration and improve flowability of the powder. The spray-dried particle can be fabricated with features which enhance aerosolization via dry powder inhaler devices, and lead to lower deposition in the mouth, throat and inhaler device.

The particles of the invention can be employed in compositions suitable for drug delivery via the pulmonary system. For example, such compositions can include the polycation-complexed biologically active agent and a pharmaceutically acceptable carrier for administration to a patient, preferably for administration via inhalation. The particles can be co-delivered with larger carrier particles, not including a therapeutic agent, the latter possessing mass median diameters for example in the range between about 50 .mu.m and about 100 .mu.m. The particles can be administered alone or in any appropriate pharmaceutically acceptable vehicle, such as a liquid, for example saline, or a powder, for administration to the respiratory system.

Particles including a medicament, for example one or more of the drugs listed above, are administered to the respiratory tract of a patient in need of treatment, prophylaxis or diagnosis. Administration of particles to the respiratory system can be by means such as known in the art. For example, particles are delivered from an inhalation device. In a preferred embodiment, particles are administered via a dry powder inhaler (DPI). Metered-dose-inhalers (MDI) or instillation techniques also can be employed.

Various suitable devices and methods of inhalation which can be used to administer particles to a patient's respiratory tract are known in the art. For example, suitable inhalers are described in U.S. Pat. No. 4,069,819, issued Aug. 5, 1976 to Valentini, et al., U.S. Pat. No. 4,995,385 issued Feb. 26, 1991 to Valentini, et al., and U.S. Pat. No. 5,997,848 issued Dec. 7, 1999 to Patton, et al. Other examples include, but are not limited to, the SPINHALER.RTM. (Fisons, Loughborough, U.K.), ROTAHALER.RTM. (Glaxo-Wellcome, Research Triangle Technology Park, N.C.), FLOWCAPS.RTM. (Hovione, Loures, Portugal), INHALATOR.RTM. (Boehringer-Ingelheim, Germany), and the AEROLIZER.RTM. (Novartis, Switzerland), the DISKHALER.RTM. (Glaxo-Wellcome, RTP, NC) and others, such as known to those skilled in the art. Preferably, the particles are administered as a dry powder via a dry powder inhaler, such as those described in U.S. patent application entitled "Inhalation Device and Method", filed Apr. 16, 2001, application Ser. No. 09/835,302 by Edwards, et al.

Preferably, particles administered to the respiratory tract travel through the upper airways (oropharynx and larynx), the lower airways which include the trachea followed by bifurcations into the bronchi and bronchioli and through the terminal bronchioli which in turn divide into respiratory bronchioli leading then to the ultimate respiratory zone, the alveoli or the deep lung. In a preferred embodiment of the invention, most of the mass of particles deposits in the deep lung. In another embodiment of the invention, delivery is primarily to the central airways. Delivery to the upper airways can also be obtained.

In one embodiment of the invention, delivery to the pulmonary system of particles is in a single, breath-actuated step, as described in U.S. patent application, High Efficient Delivery of a Large Therapeutic Mass Aerosol, application Ser. No. 09/591,307, filed Jun. 9, 2000, which is incorporated herein by reference in its entirety. In another embodiment of the invention, at least 50% of the mass of the particles stored in the inhaler receptacle is delivered to a subject's respiratory system in a single, breath-activated step. In a further embodiment, at least 5 milligrams and preferably at least 10 milligrams of a medicament is delivered by administering, in a single breath, to a subject's respiratory tract particles enclosed in the receptacle. Amounts as high as 15, 20, 25, 30, 35, 40 and 50 milligrams can be delivered.

As used herein, the term "effective amount" means the amount needed to achieve the desired therapeutic or diagnostic effect or efficacy. The actual effective amounts of drug can vary according to the specific drug or combination thereof being utilized, the particular composition formulated, the mode of administration, and the age, weight, condition of the patient, and severity of the symptoms or condition being treated. Dosages for a particular patient can be determined by one of ordinary skill in the art using conventional considerations, (e.g. by means of an appropriate, conventional pharmacological protocol). For example, effective amounts of albuterol sulfate range from about 100 micrograms (.mu.g) to about 10 milligrams (mg).

Aerosol dosage, formulations and delivery systems also may be selected for a particular therapeutic application, as described, for example, in Gonda, I. "Aerosols for delivery of therapeutic and diagnostic agents to the respiratory tract," in Critical Reviews in Therapeutic Drug Carrier Systems, 6: 273 313, 1990; and in Moren, "Aerosol dosage forms and formulations," in: Aerosols in Medicine. Principles, Diagnosis and Therapy, Moren, et al., Eds, Esevier, Amsterdam, 1985.
 

Claim 1 of 48 Claims

1. A method of delivery to the pulmonary system comprising: administering to the respiratory tract of a patient in need of treatment, prophylaxis or diagnosis an effective amount of a dry powder having a tap density of less than about 0.1 g/cm.sup.3 and comprising a polycationic complexing agent which is complexed with a therapeutic, prophylactic or diagnostic agent wherein, the amount of polycationic complexing agent present in the particles is about 5% weight/weight or more of the total weight of the complexing agent and therapeutic, diagnostic or prophylactic agent and wherein release of the agent is sustained.

____________________________________________
If you want to learn more about this patent, please go directly to the U.S. Patent and Trademark Office Web site to access the full patent.