United States Patent:   6,024,977

Assignee:  Oregon Health Sciences University (Portland, OR)

Filed:  September 3, 1997

This invention herein describes a method of facilitating the entry of drugs into cells and tissues at physiologically protected sites at pharmicokinetically useful levels and also a method of targeting drugs to specific organelles within the cell. This polar lipid/drug conjugate targeting invention embodies an advance over other drug targeting methods known in the prior art, because the invention provides drug concentrations in such physiologically protected sites that can reach therapeutically-effective levels after administration of systemic levels much lower than are currently administered to achieve a therapeutic dose. This technology is appropriate for use with psychotropic, neurotropic and neurological drugs, agents and compounds, for rapid and efficient introduction of such agents across the blood-brain barrier. Further, the invention provides means for retention and prolonged enzymatic release of psychotropic, neurotropic and neurological drugs, agents and compounds comprising the conjugates of the invention, in the brain and central nervous system.

SUMMARY OF THE INVENTION

The present invention is directed to an improved method for delivering biologically-active compounds, particularly drugs including preferably psychotropic, neurotropic and neurologically-acting drugs, to physiologically protected sites in an animal in vivo. This delivery system achieves specific delivery of such biologically-active compounds through conjugating the compounds with a polar lipid carrier. This invention has the specific advantage of facilitating the entry of such compounds into cells and tissues protected by such physiological barriers as the blood-brain barrier via a polar lipid carrier, achieving effective intracellular concentration of such compounds more efficiently and with more specificity than conventional delivery systems.

The invention provides compositions of matter comprising a biologically-active compound covalently linked to a polar lipid carrier molecule. Preferred embodiments also comprise a spacer molecule having two linker functional groups, wherein the spacer has a first end and a second end and wherein the lipid is attached to the first end of the spacer through a first linker functional group and the biologically-active compound is attached to the second end of the spacer through a second linker functional group. In preferred embodiments, the biologically-active compound is a drug, most preferably a psychotropic, neurotropic or neurologically-acting drug or agent, or an antioxidant. Preferred polar lipids include but are not limited to acyl- and acylated carnitine, sphingosine, ceramide, phosphatidyl choline, phosphatidyl glycerol, phosphatidyl ethanolamine, phosphatidyl inositol, phosphatidyl serine, cardiolipin and phosphatidic acid. Preferred biologically-active compounds include neurotropic agents such as L-dopa, hydroxytryptamine and metabolites thereof; amantadine, benztropine, bromocryptine, diphenhydramine, levadopa (a particularly preferred embodiment) and combinations thereof (e.g., with carbidopa as provided as Sinemet.RTM.); pergolid, trihexphenidyl, ethosuximide, valproic acid, carbamazepine (e.g., Tegretol.RTM.) and, in a particularly preferred embodiment, the 10- or 11-hydroxy analogues of carbamazepine; primidone, gabapentin in a particularly preferred embodiment; lamotrigine in a particularly preferred embodiment; felbamate, paramethadione and trimethadione; phenothiazines, thioxanthemes and related compounds; clozapine, haldoperidol, loxapine (Loxitane.RTM.), benzodiazapene antidepressants of the norepinephrine reuptake inhibitor type; monoamine oxidase inhibitors, and antioxidants such as carotenes, glutathione and N-acetylcysteine. Pharmaceutical compositions comprising the drug/polar lipid conjugates of the invention are also provided.

The invention also provides compositions of matter comprising a biologically-active compound covalently linked to a lipid, most preferably a polar lipid, carrier molecule via a spacer molecule wherein the spacer allows the biologically-active compound to act without being released at an intracellular site. In these embodiments of the invention, the first linker functional group attached to the first end of the spacer is characterized as "strong" and the second linker functional group attached to the second end of the spacer is characterized as "weak", with reference to the propensity of the covalent bonds between each end of the spacer molecule to be broken.

In other embodiments of the compositions of matter of the invention, the spacer allows the facilitated hydrolytic release of the biologically-active compound at an intracellular site. Other embodiments of the spacer facilitate the enzymatic release of the biologically-active compound at an intracellular site. In particularly preferred embodiments, the spacer functional group is hydrolyzed by an enzymatic activity found in brain tissue, including neuronal, glial and other brain cell types, preferably an esterase and most preferably an esterase having a differential expression and activity profile in the appropriate target cell type. In additional preferred embodiments, specific release of biologically-active compounds is achieved by enzymatic or chemical release of the biologically-active compound by extracellular cleavage of a cleavable linker moiety via an enzymatic activity specific for brain tissue, with resulting specific uptake of the released psychotropic, neurotropic or neurological agent by the appropriate cell in said tissue.

In another embodiment of this aspect of the invention, the spacer molecule is a peptide of formula (amino acid).sub.n, wherein n is an integer between 2 and 25, preferably wherein the peptide comprises a polymer of one or more amino acids.

In other embodiments of the compositions of matter of the invention, the biologically-active compound of the invention has a first functional linker group, and a lipid, most preferably a polar lipid, carrier has a second functional linker group, and the compound is covalently linked directly to the lipid carrier by a chemical bond between the first and second functional linker groups. In preferred embodiments, each of the first and second functional linker groups is a hydroxyl group, a primary or secondary amino group, a phosphate group or substituted derivatives thereof or a carboxylic acid group.

In another aspect of the invention is provided compositions of matter comprising a drug, most preferably an a psychotropic, neurotropic or neurological drug or agent, covalently linked to a polar lipid carrier molecule. Preferred embodiments also comprise a spacer molecule having two linker functional groups, wherein the spacer has a first end and a second end and wherein the lipid is attached to the first end of the spacer through a first linker functional group and the drug is attached to the second end of the spacer through a second linker functional group. Preferred embodiments of the invention are provided wherein the drug is a psychotropic, neurotropic or neurological drug or agent. Preferred polar lipids include but are not limited to acyl- and acylated carnitine, sphingosine, ceramide, phosphatidyl choline, phosphatidyl glycerol, phosphatidyl ethanolamine, phosphatidyl inositol, phosphatidyl serine, cardiolipin and phosphatidic acid. Preferred psychotropic, neurotropic or neurological drugs or agents comprising the conjugates of the invention include L-dopa, hydroxytryptamine and metabolites thereof; amantadine, benztropine, bromocryptine, diphenhydramine, levadopa (a particularly preferred embodiment) and combinations thereof (e.g., with carbidopa as provided as Sinemet.RTM.); pergolid, trihexphenidyl, ethosuximide, valproic acid, carbamazepine (e.g., Tegretol.RTM.) and, in a particularly preferred embodiment, the 10- or 11-hydroxy analogues of carbamazepine; primidone, gabapentin in a particularly preferred embodiment; lamotrigine in a particularly preferred embodiment; felbamate, paramethadione and trimethadione; phenothiazines, thioxanthemes and related compounds; clozapine, haldoperidol, loxapine (Loxitane.RTM.), benzodiazapene antidepressants of the norepinephrine reuptake inhibitor type; monoamine oxidase inhibitors, and antioxidants such as carotenes, glutathione and N-acetylcysteine. Pharmaceutical compositions comprising the drug/polar lipid conjugates of the invention are also provided.

The invention also provides compositions of matter comprising a psychotropic, neurotropic or neurological drug or agent, covalently linked to a polar lipid carrier molecule via a spacer molecule, wherein the spacer allows the drug to act without being released at an intracellular site. In these embodiments of the invention, the first linker functional group attached to the first end of the spacer is characterized as "strong" and the second linker functional group attached to the second end of the spacer is characterized as "weak", with reference to the propensity of the covalent bonds between each end of the spacer molecule to be broken.

In other embodiments of the compositions of matter of the invention, the spacer allows the facilitated hydrolytic release of a psychotropic, neurotropic or neurological drug or agent at an intracellular site. Other embodiments of the spacer facilitate the enzymatic release of the psychotropic, neurotropic or neurological drug or agent of the invention at an intracellular site. In particularly preferred embodiments, the spacer functional group is hydrolyzed by an enzymatic activity found in a physiologically-protected site, such as the brain and central nervous system and more particularly including neuronal, glial and other brain cell types, wherein said enzymatic activity is preferably an esterase and most preferably an esterase having a differential expression and activity profile in different tissue cell types. In additional preferred embodiments, specific release of the psychotropic, neurotropic or neurological drug or agent of the invention is achieved by enzymatic or chemical release of these drugs by extracellular cleavage of a cleavable linker moiety via an enzymatic activity specific for, for example, brain tissue, followed by specific uptake of the released psychotropic, neurotropic or neurological drug or agent by the appropriate cell in said tissue.

In another embodiment of this aspect of the invention, the spacer molecule is a peptide of formula (amino acid).sub.n, wherein n is an integer between 2 and 25, preferably wherein the peptide comprises a polymer of one or more amino acids.

In still further embodiments of the compositions of matter of the invention are provided psychotropic, neurotropic or neurological drugs or agents having a first functional linker group, and a polar lipid carrier having a second functional linker group, wherein the drug is covalently linked directly to the polar lipid carrier by a chemical bond between the first and second functional linker groups. In preferred embodiments, each of the first and second functional linker groups is a hydroxyl group, a primary or secondary amino group, a phosphate group or substituted derivatives thereof or a carboxylic acid group. Preferred psychotropic, neurotropic or neurological drugs or agents comprising the conjugates of the invention include L-dopa, hydroxytryptamine and metabolites thereof; amantadine, benztropine, bromocryptine, diphenhydramine, levadopa (a particularly preferred embodiment) and combinations thereof (e.g., with carbidopa as provided as Sinemet.RTM.); pergolid, trihexphenidyl, ethosuximide, valproic acid, carbamazepine (e.g., Tegretol.RTM.) and, in a particularly preferred embodiment, the 10- or 11-hydroxy analogues of carbamazepine; primidone, gabapentin in a particularly preferred embodiment; lamotrigine in a particularly preferred embodiment; felbamate, paramethadione and trimethadione; phenothiazines, thioxanthemes and related compounds; clozapine, haldoperidol, loxapine (Loxitane.RTM.), benzodiazapene antidepressants of the norepinephrine reuptake inhibitor type; monoamine oxidase inhibitors, and antioxidants such as carotenes, glutathione and N-acetylcysteine. Preferred polar lipids include but are not limited to acyl- and acylated carnitine, sphingosine, ceramide, phosphatidyl choline, phosphatidyl glycerol, phosphatidyl ethanolarnine, phosphatidyl inositol, phosphatidyl serine, cardiolipin and phosphatidic acid. Pharmaceutical compositions comprising the drug/polar lipid conjugates of the invention are also provided.

Preferred embodiments of this aspect of the invention include compositions of matter that are polar lipid conjugates of anticonvulsive agents, antiparkinsonian drugs, alkaloids, catecholamines including dopamine analogues and derivatives, muscarinic receptor agonists and antagonists, cholinergic receptor agonists and antagonists, calcium channel blockers, .gamma.-aminobutyric acid (GABA) receptor agonists, antagonists, and uptake inhibitors and enhancers; phenothiazines, thioxanthemes and related compounds; clozapine, haldoperidol, loxapine (Loxitane.RTM.), benzodiazapene antidepressants of the norepinephrine reuptake inhibitor type; monoamine oxidase inhibitors; antidepressants and antimanic agents, antioxidants and other compounds that mitigate the effects of reactive oxygen species (for the treatment of Alzheimer's disease, Parkinson's disease, or other neurodegenerative conditions such as ataxia telangiectasia and amyelolaterosclerosis (ALS)).

As disclosed herein, the invention comprehends a polar lipid-drug conjugate wherein the polar lipid selectively promotes association with and transit across certain physiological barriers to protected tissue sites, thereby facilitating delivery of drugs and other pharmaceutical agents to such physiologically restricted or protected sites. In embodiments comprising a spacer moiety, the spacer component of the conjugates of the invention will preferably act to specifically release the drug from the lipid at the target site; prevent the non-specific release from the drug from the lipid in the systemic circulation or in hepatic, renal or other inappropriate cells, tissue or organs; target the conjugate to a specific cell or cell type within the protected tissue; prevent interaction and/or uptake of the drug by hematopoietic, ocular, hepatic or renal tissues; or perform other functions to maximize the effectiveness of the drug.

This type of conjugate has numerous advantages. The drug-lipid conjugates of the invention provide delivery of a variety of psychotropic, neurotropic and neurological drugs and agents to physiologically restricted or protected sites in vivo at concentrations and pharmicokinetic rates not heretofore attainable. A benefit of this advantage is the achievement of therapeutic indices of agents in such protected sites whereby the agent is useful for achieving a desired therapeutic goal. Another benefit is decreased hepatic toxicity, hematopoietic suppression (such as thrombocytopenia, leukopenia, aplastic anemia, leukocytosis, eosinophilia, pancytopenia, agranulocytosis), reduced systemic metabolism, degradation and toxicity, reduced hepatic clearance, reduced systemic adverse drug interactions, and generally reduced side effects due to the achievement of a lower, therapeutically-effective dose as the result of surmounting the physiological barrier. These biological effects can also result in simplified dosage schedules, particularly for drugs with short systemic half-lives.

In addition, the lipid/drug conjugates promote the intracellular entry of a variety of potentially useful drugs at pharmokinetic rates not currently attainable. The range of targeted cell types is not limited per se by particular, limited biological properties of the cell (such as the number and type of specific receptor molecules expressed on the cell surface). In contrast to traditional attempts to simply target drugs to specific cells, the conjugates of the invention can also target drugs to specific intracellular organelles and other intracellular compartments. In certain preferred embodiment, the conjugates of the invention incorporate a variable spacer region that may allow pharmacologically-relevant rates of drug release from polar lipid carrier molecules to be engineered into the compositions of the invention, thereby increasing their clinical efficacy and usefulness. Thus, time-dependent drug release and specific drug release in cells expressing the appropriate degradative enzymes are a unique possibility using the drug-lipid conjugates of the invention.

In particular, felicitous design of the psychotropic, neurotropic/neurological drug/spacer/polar lipid conjugate can provide an in vivo reservoir of time-dependent drug release in the physiologically protected tissue, resulting in specific delivery of therapeutic amounts to such tissues using a reduced dosage regime to minimize non-specific, systemic and deleterious side effects. In such formulations, the amount and activity of the psychotropic, neurotropic or neurological drug can be modulated by release via cleavage, preferably hydrolytic cleavage, of the spacer moiety, most preferably by an enzymatic activity in the protected tissue (e.g., brain) that has a differential pattern of expression or activity in different cell types in said tissue. The conjugates of the invention can also be combined with other drug delivery approaches to further increase specificity and to take advantage of useful advances in the art.

Claim 1 of 28 Claims

1. A method for treating a pathological condition or disease state in cells, tissues or organs in an animal, the method comprising the step of administering to the animal a pharmaceutical composition comprising a psychotropic, neurotropic or neurological drug, a polar lipid carrier, two linker functional groups and a spacer, wherein the spacer has a first end and a second end and wherein the polar lipid is attached to the first end of the spacer through a first linker functional group and the drug is attached to the second end of the spacer through a second linker functional group, in an acceptable carrier or formulation and in an amount sufficient to alleviate the pathological condition or disease state in the animal.

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