The invention provides methods for treating an individual having a retinoid responsive disorder. In one embodiment, a method involves administering to the individual an effective amount of a selective CYP26B inhibitor, the selective CYP26B inhibitor having at least 10-fold selectivity for CYP26B relative to CYP26A. In another embodiment, a method involves administering to the individual an effective amount of a selective CYP26A inhibitor, the selective CYP26A inhibitor having a chemical formula set forth in the specification. The invention further provides screening methods for identifying a selective CYP26A inhibitor or selective CYP26B inhibitor.

Description of the Invention

Retinoids play important roles in regulating gene expression during embryonic development and in the maintenance of adult epithelial tissues. The amount of a retinoid, such as retinoic acid (RA), present in the body at a given time is regulated, in part, by cellular metabolism. The cytochrome P450 molecules CYP26A and CYP26B, also known as P450RAI-1 and P450RAI-2, respectively, metabolize RA into more polar hydroxylated and oxidized derivatives, thus reducing the level of this retinoid in the body. By blocking the RA-destroying activity of CYP26A or CYP26B, the amount of natural or administered RA in a cell can be maintained at a beneficial level. Therefore, an inhibitor of CYP26A or CYP26B can be used to beneficially maintain or increase a level of a retinoid in an individual, either alone or in conjunction with retinoid treatment.

Administration of certain CYP26A inhibitors to mammals has been observed to cause a significant increase in endogenous retinoic acid levels (see, for example, U.S. Pat. Nos. 6,531,599 and 6,495,552). For example, treatment of human patients with the CYP26A inhibitor liarozole results in beneficial effects similar to those observed upon treatment with retinoids, such as amelioration of psoriasis (Kuijpers, et al., British Journal of Dermatology 139:380-389 (1998)).

As herein below, selective inhibitors of CYP26A having at least 10-fold selectivity for CYP26A relative to CYP26B have been identified and are referenced as Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29 (see Original Patent). A variety of exemplary selective CYP26A inhibitors and selective CYP26B inhibitors are shown in Table 4 (see Original Patent). The selective CYP26A inhibitors shown in Table 4 have selectivity that ranges from at least 10-fold to at least 888-fold selectivity for CYP26A relative to CYP26B. The selective CYP26B inhibitors shown in Table 4 have selectivity that ranges from at least 10-fold to at least 83-fold selectivity for CYP26B relative to CYP26A. Although the selective CYP26A inhibitors disclosed herein are structurally unrelated to previously described CYP26A inhibitors, these compounds can provide similar beneficial effects by reducing destruction of endogenous or administered retinoids, including, without limitation, retinoic acid and retinoic acid analogs. Based on the identification of selective inhibitors of CYP26A and CYP26B, the present invention provides therapeutic methods that involve selective inhibition of CYP26A or CYP26B. The methods are useful for beneficially treating an individual having any of a variety of retinoid responsive disorders disclosed herein below or otherwise known in the art.

As disclosed herein in Example I, compounds that selectively inhibit CYP26A or CYP26B activity were demonstrated to be effective in reducing sebocyte differentiation in an animal model of acne development. In this animal model, which involves observation of sebaceous gland differentiation in hamster flank organ, retinoids and retinoic acid receptor agonists blocked differentiation, which is akin to blocking acne development in a mammal such as a human. Moreover, treatment with a selective CYP26A inhibitor or a selective CYP26B inhibitor reduced sebocyte differentiation as effectively as ACCUTANE.TM., one of the most widely used commercially available retinoic acid acne medications. Thus, in one embodiment, the present invention provides a method of treating a retinoid responsive disorder by administering an effective amount of a selective CYP26A inhibitor having any of Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29 or a selective CYP26B inhibitor that has at least 10-fold selectivity for CYP26B relative to CYP26A.

As used herein, the term "retinoid responsive disorder" means a condition or disease that normally has at least one symptom that is improved, alleviated, delayed in onset, or prevented upon administration of retinoic acid (RA) or a synthetic retinoid having RA activity. A variety of retinoid responsive disorders are well known to those skilled in the art and include, without limitation, cancers such as skin cancer, oral cancer and Kaposi's sarcoma; skin disorders such as acne, psoriasis and eczema; and multiple other disorders of diverse etiology, including but not limited to emphysema and Alzheimer's disease.

The ability of RA or a synthetic retinoid having RA activity to improve, alleviate, delay onset of, or prevent at least one symptom of a condition or disease, if not known, can be determined using well known methods, including those described herein below. A synthetic retinoid used for such a determination can be any of a variety of experimental or clinically used retinoids. Exemplary synthetic retinoids currently in clinical use include acitretin, isotretinoin, tretinoin, tazarotene and adapalene.

A variety of retinoid responsive skin disorders can be treated according to a method of the invention. Such skin disorders include, without limitation, inflammatory and non-inflammatory acne, psoriasis, eczema, atopic dermatitis, Pityriasis rubra pilaris, multiple basal cell carcinomasactinic keratoses, arsenic keratoses, ichthyoses and other keratinization and hyperproliferative disorders of the skin, Darriers disease, lichen planus, glucocorticoid damage (steroid atrophy), microbial infection of the skin, excessive pigmentation of the skin, and photodamage of the skin.

In several embodiments, the invention provides methods for treating acne that involve administering a selective CYP26A inhibitor or a selective CYP26B inhibitor. Acne is a common disease of the pilosebaceous glands, and is characterized by comedones, papules, pustules, inflamed nodules, superficial pus-filled cysts, and (in extreme cases) canalizing and deep, inflamed, sometimes purulent sacs.

Multiple categories or types of acne have been defined in the art based on the severity of the condition. For example, "superficial acne" is characterized by blackheads (open comedones) or whiteheads (closed comedones), inflamed papules, pustules, and superficial cysts, with large cysts occurring occasionally. As another example, "deep acne" is similarly characterized except with deep inflamed nodules and pus-filled cysts, which often rupture and become abscesses. Other categories of acne include "acne vulgaris," which is the most common form of acne; "acne conglobata," which is acne that covers the back, chest, and buttocks with pustules and nodules that often connect under the skin; "acne fulminans," which is an extreme form of acne conglobata that involves a sudden onset of pustules and nodules, infected nodules, fever, joint pain, and possible loss of weight or appetite; "acne medicamentosa," which is caused by a drug; "comedonal acne," which is acne characterized by whiteheads and blackheads without other forms of skin lesions, and "cystic acne," which occurs when the infected contents of a pustule or pimple erupt beneath the skin, rather than on the surface. It is understood that a method of the invention can be used to beneficially treat an individual having any form of mild, moderate or severe acne including, but not limited to, those discussed above.

In a further embodiment, the invention provides a method for treating an individual having a proliferative disorder that involves administering to the individual an effective amount of a selective CYP26B inhibitor that has at least 10-fold selectivity for CYP26B relative to CYP26A, or an effective amount of a selective CYP26A inhibitor, which is represented by any of Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29. As used herein, the term "proliferative disorder" means a disease or abnormal condition that results in unwanted or abnormal cell growth, viability or proliferation. Proliferative disorders include diseases such as cancer, in which the cells are neoplastically transformed, and diseases resulting from overgrowth of normal cells. For example, cell proliferative disorders include diseases associated with the overgrowth of connective tissues, such as various fibrotic diseases, including scleroderma, arthritis, alcoholic liver cirrhosis, keloid, and hypertropic scarring; vascular proliferative disorders, such as atherosclerosis; benign tumors, and the abnormal proliferation of cells mediating autoimmune disease. As used herein, the term "cancer" means a class of diseases characterized by the uncontrolled growth of aberrant cells, including all known cancers, and neoplastic conditions, whether characterized as malignant, benign, soft tissue or solid tumor. Specific cancers that can be treated using a method of the invention include, without limitation, cancers of the skin, breast, eye, prostate, cervix, uterus, colon, bladder, esophagus, stomach, lung, larynx, oral cavity, blood and lymphatic system, metaplasias, dysplasias, neoplasias, leukoplakias, papillomas of the mucous membranes and Kaposi's sarcoma.

A compound that beneficially maintains or increases a level of a retinoid in the body by selective inhibition of CYP26A or CYP26B can be used to treat a variety of cancers. For example, increasing the amount of retinoid in the body of cancer patients has been a successful strategy for treating a variety of cancers. For example, retinoids have been used to reduce or prevent oral, skin and head and neck cancers in individuals at risk for these tumors (see, for example, Bollag et al., Ann. Oncol. 3:513-526 (1992); Chiesa et al., Eur. J. Cancer B. Oral Oncol. 28:97-102 (1992); Costa et al., Cancer Res. 54:Supp. 7, 2032-2037 (1994)). Retinoids have also been used to treat squamous cell carcinoma of the cervix and the skin (see, for example, Verma, Cancer Res. 47:5097-5101 (1987); Lippman et al., J. Natl Cancer Inst. 84:235-241 (1992); Lippman et al., J. Natl Cancer Inst. 84:241-245 (1992)) and Kaposi's sarcoma (see, for example, Bonhomme, et al., Ann. Oncol. 2:234-235 (1991)), and have found significant use in the therapy of acute promyelocytic leukemia (see, for example, Huang et al., Blood 72:567-572 (1988); Castaigne et al., Blood 76:1704-1709 (1990); Lo Coco et al., Blood 77:1657-1659 (1991); Warrell et al., N. Engl. J. Med 324:1385-1393 (1991); and Chomienne et al., FASEB J. 10: 1025-1030 (1996)). Therefore, a compound that beneficially maintains or increases a level of a retinoid in the body by selective inhibition of CYP26A or CYP26B can be used to treat a variety of cancers using a method of the invention.

In another embodiment, the invention provides a method for treating an individual having a retinoid responsive neurological disorder that involves administering to the individual an effective amount of a selective CYP26B inhibitor that has at least 10-fold selectivity for CYP26B relative to CYP26A, or an effective amount of a selective CYP26A inhibitor represented by any of Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29. Because a selective CYP26A inhibitor or selective CYP26B inhibitor can beneficially maintain or increase a level of a retinoid in a nervous system tissue of an individual, such an inhibitor can be used for treating a variety of neurological disorders that respond beneficially to retinoids. Both CYP26A and CYP26B are highly expressed in the brain, and multiple lines of evidence indicate that retinoids such as RA have important roles in normal neurological function and neurological disease. For example, retinaldehyde, dehydrogenase, the enzyme that forms retinoic acid from retinaldehyde has 1.5 to 2-fold higher activity in the hippocampus and parietal cortex of Alzheimer's diseased brains than in normal controls (Conner and Sidell, Mol Chem Neuropathol 30(3):239-52 (1997)). In addition, retinoid hypofunction and impaired retinoid transport have been indicated to be contributing factors in Alzheimer's Disease (Goodman and Pardee, Proc. Natl. Acad. Sci. USA 4; 100(5):2901-5 (2003)). Exemplary retinoid responsive neurological disorders that can be treated with a selective CYP26A or selective CYP26B inhibitor according to a method to the invention, include, yet are not limited to, Alzheimer's Disease, schizophrenia, Parkinson's disease, anxiety, depression, drug addiction, disorders of cognition, emesis, eating disorders, attention deficit-hyperactivity disorder, Tourette's Syndrome, Huntington's disease, tardive dyskinesia, Lesch-Nyhan disease, Rett syndrome or any neurological disorder that is retinoid responsive.

In a further embodiment, the invention provides a method for treating an individual having a retinoid responsive inflammatory disorder or autoimmune disorder by administering to the individual an effective amount of a selective CYP26B inhibitor that has at least 10-fold selectivity for CYP26B relative to CYP26A, or an effective amount of a selective CYP26A inhibitor represented by any of Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29. A method of the invention can be used to treat any of a variety of inflammatory disorders, including, without limitation, those resulting from injury; infection by a bacteria, virus, fungus or other pathogen; autoimmune disorders; and other abnormal conditions. Exemplary retinoid responsive inflammatory disorders that can be treated using a method of the invention include, without limitation, inflammatory skin disorders, for example, psoriasis; inflammatory gastrointestinal disorders, for example, ileitis, irritable bowel syndrome, ulcerative colitis and Crohn's disease; autoimmune disorders such as rheumatoid and other forms of arthritis; organ transplant rejection; and any inflammatory or autoimmune disorder that is retinoid responsive.

In another embodiment, the invention provides a method for treating an individual having a retinoid responsive ocular disorder that involves administering to the individual an effective amount of a selective CYP26B inhibitor that has at least 10-fold selectivity for CYP26B relative to CYP26A, or an effective amount of a selective CYP26A inhibitor represented by any of Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29. A method of the invention can be used for treating a variety of ocular disorders, including, without limitation, diabetic retinopathy; macular edema such as macular edema associated with diabetes mellitus or other conditions; retinal degeneration such as age-related macular degeneration or retinitis pigmentosa; inflammatory disorders of the retina; vascular occlusive conditions of the retina such as retinal vein occlusions or branch or central retinal artery occlusions; retinopathy of prematurity; retinopathy associated with blood disorders such as sickle cell anemia; damage following retinal detachment; damage or insult due to vitrectomy surgery or retinal surgery; and other retinal damage including therapeutic damage such as that resulting from laser treatment of the retina, for example, pan-retinal photocoagulation for diabetic retinopathy or photodynamic therapy of the retina, for example, for age-related macular degeneration; genetic and acquired optic neuropathies such as optic neuropathies characterized primarily by loss of central vision, for example, Leber's hereditary optic neuropathy (LHON), autosomal dominant optic atrophy (Kjer disease) and other optic neuropathies such as those involving mitochondrial defects, aberrant dynamin-related proteins or inappropriate apoptosis, proliferative vitreoretinopathy (PVR), retinal detachment, dry eye, as well as any ocular disorder that is retinoid responsive. In reference to ocular disorders see, for example, Carelli et al., Neurochem. Intl. 40:573-584 (2002); and Olichon et al., J. Biol. Chem. 278:7743-7746 (2003).

In yet another embodiment, the invention provides a method for treating an individual having a pulmonary disorder that involves administering to the individual an effective amount of a selective CYP26B inhibitor that has at least 10-fold selectivity for CYP26B relative to CYP26A, or an effective amount of a selective CYP26A inhibitor represented by any of Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29. Retinoid treatment has been used to successfully treat, for example, emphysema, which is a pulmonary disorder resulting from progressive destruction of alveolar septae that was considered irreversible until it was shown that retinoid acid administration can reverse anatomic and physiologic signs of emphysema in a rat model (Massaro and Massaro, Nature Medicine 3:675-7 (1997)). A method of the invention can be used to treat emphysema or another pulmonary disorder that is retinoid responsive. Non-limiting examples of pulmonary disorders include, but are not limited to, obstructive pulmonary disorders such as emphysema, chronic bronchitis, bronchial asthma, bronchiectasis and cystic fibrosis; and restrictive pulmonary disorders such as interstitial fibrosis, pulmonary edema, adult respiratory distress syndrome, rheumatoid spondylitis and pleural effusion.

It is understood that increasing the level of a retinoid in a tissue of an individual can have beneficial effects in individuals having a variety of other retinoid responsive disorders, including cardiovascular disorders such as, without limitation, diseases associated with lipid metabolism including dyslipidemias and post-angioplasty restenosis; and diabetes. Thus, the methods can be useful for treating a variety of retinoid responsive disorders including, but not limited to, skin disorders, autoimmune disorders, inflammatory disorders, proliferative disorders, neurological disorders, ocular disorders and pulmonary disorders.

By specific mention of the above categories of retinoid responsive disorders, those skilled in the art will understand that such terms include all classes and types of these disorders. For example, the term "skin disorder" is intended to include any skin disorder having a symptom that is improved, alleviated, delayed in onset or prevented upon administration of RA or a synthetic retinoid having RA activity; and likewise for other classes of disorders such as proliferative disorders, pulmonary disorders, autoimmune disorders, inflammatory disorders, neurological disorders and ocular disorders. Those skilled in the art will know how to appropriately assess whether a disorder has at least one symptom that is improved, alleviated, delayed in onset or prevented upon administration of RA or a synthetic retinoid having RA activity. Therefore, the methods of the invention are applicable to known retinoid responsive disorders as well as disorders determined to be retinoid responsive, for example, in an animal model that corresponding to a particular disorder.

As disclosed herein below, CYP26A and CYP26B have distinct expression levels in various tissues in humans and other mammals. Thus, a selective CYP26A inhibitor or selective CYP26B inhibitor can be used to inhibit the activity of a particular CYP26, such as CYP26A, for example, in a selected tissue without substantially altering the activity of CYP26B in that tissue, and visa versa. A selective CYP26A inhibitor or selective CYP26B inhibitor also can be used to target inhibition of a CYP26 in a particular tissue, if desired. It is also understood that a selective CYP26A inhibitor or selective CYP26B inhibitor can be beneficially used without targeting particular tissues for CYP26 inhibition.

In humans, CYP26A expression has been observed in liver, brain and placenta (Ray et al. J. Biol. Chem. 272:18702-18708 (1997)); CYP26B expression has been observed in brain (White et al. Proc. Natl. Acad. Sci. USA 97:6403-6408 (2000)) as well as in kidney, lung, spleen, fetal spleen, skeletal muscle, thymus, peripheral blood leukocyte, lymph node, bone, stomach, placenta, duodenum, small intestine, and pituitary gland (PCT/CA00/01493). It is understood that either or both CYP26A and CYP26B can be present in other tissues in humans or other mammals. For example, as shown in Example I, both CYP26A and CYP26B are expressed in the skin of hamsters.

It is understood that a selective CYP26A or selective CYP26B inhibitor can be useful for treating a retinoid responsive disorder associated with a tissue in which either CYP26A or CYP26B is expressed. For example, an individual having a cancer in a tissue in which CYP26A is expressed can be treated using a selective CYP26A inhibitor, while an individual having a cancer in a tissue in which CYP26B is expressed can be treated using a selective CYP26B inhibitor. Thus, the expression of CYP26A or CYP26B in a particular tissue can be used to assess whether treatment of a particular retinoid responsive disorder with a selective CYP26A or selective CYP26B inhibitor is appropriate. It is understood that a selective CYP26A or selective CYP26B inhibitor can be useful for treating a retinoid responsive disorder even when the target tissue contains a low level of expression of either CYP26A or CYP26B. Methods for determining expression levels of CYP26A or CYP26B mRNA or protein are well known to those skilled in the art and are described, for example, in Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, New York (1992) and Ausubel et al., Current Protocols in Molecular Biology, John Wiley and Sons, Baltimore, Md. (1998).

Selective CYP26A and CYP26B Inhibitors

A. Functional Characteristics

The methods of the invention involve administering a selective CYP26A inhibitor or selective CYP26B inhibitor. As used herein, the term "selective CYP26A inhibitor" means a compound that reduces CYP26A expression or activity at least 10-fold more than any reduction affected by the inhibitor on the expression or activity of CYP26B. A selective CYP26A inhibitor can have, for example, at least 10-fold selectivity for CYP26A relative to CYP26B, at least 20-fold, at least 40-fold, at least 80-fold, at least 100-fold, at least 500-fold, at least 800-fold, or at least 1000-fold selectivity for CYP26A relative to CYP26B. As used herein, a "CYP26B inhibitor" means that reduces CYP26B expression or activity at least 10-fold more than any reduction affected by the inhibitor on the expression or activity of CYP26A. A selective CYP26B inhibitor can have, for example, at least 10-fold selectivity for CYP26A relative to CYP26B, at least 20-fold, at least 40-fold, at least 80-fold, at least 100-fold, at least 500-fold, at least 800-fold, or at least 1000-fold selectivity for CYP26B relative to CYP26A. Exemplary small molecule selective CYP26A inhibitors are disclosed herein as Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29; exemplary small molecule selective CYP26B inhibitors are disclosed herein as Formulas 5, 15 and 30 through 32. A selective CYP26A or selective CYP26B inhibitor also can be, for example, a protein, peptide, peptidomimetic, ribozyme, nucleic acid molecule or oligonucleotide, oligosaccharide, or small molecule or combination thereof.

A selective CYP26A inhibitor or selective CYP26B inhibitor useful in a method of the invention can act by any mechanism, and can, for example, alter the catalytic action of the specified enzyme and consequently reduce, or in some cases, stop catalysis. A selective CYP26A inhibitor or selective CYP26B inhibitor can therefore be a competitive, noncompetitive, or uncompetitive inhibitor of CYP26A or CYP26B, respectively, and further can function in a reversible or irreversible manner. It is understood that a selective CYP26A inhibitor or selective CYP26B inhibitor also can act indirectly, such as by reducing or down-regulating mRNA or protein expression of CYP26A or CYP26B, respectively.

A selective CYP26A inhibitor or selective CYP26B useful in a method of the invention does not bind to a retinoic acid receptor (RAR), or subunit thereof.

B. Structural Characteristics

As disclosed herein, a variety of structurally unrelated compounds having at least 10-fold selectivity for CYP26A relative to CYP26B can be selective CYP26A inhibitors; and a variety of structures unrelated compounds having at least 10-fold selectivity for CYP26B relative to CYP26A, can be selective CYP26B inhibitors and therefore can be useful in a method of the invention. A selective CYP26A inhibitor can be, for example, an organic molecule represented by any of Formulas 1 through 4, 6 through 14, 16, 17 and 18 through 29 below, or a pharmaceutically acceptable salt, ester, amide, steroisomer or racemic mixture thereof. A selective CYP26B can be, for example, an organic molecule represented by any of Formulas 5, 15 and 30 through 32, below, or a pharmaceutically acceptable salt, ester, amide, steroisomer or racemic mixture thereof.

As disclosed herein in Table 4 (see Original Patent), identified CYP26A inhibitors have selectivities of about 10-fold, about 40-fold, about 60-fold, about 80-fold, about 200-fold, about 300-fold, and about 900-fold; the identified CYP26B inhibitors have selectivities of about 10 fold, such as about 25-fold, about 50-fold and about 80-fold. It is understood that these and other selective CYP26A inhibitors and selective CYP26B inhibitors can be useful for treating a retinoid responsive disorder according to a method of the invention.
 

Claim 1 of 12 Claims

1. A method for treating an individual having a retinoid responsive skin disorder selected from the group consisting of acne, psoriasis, eczema, atopic dermatitis, Pityriasis rubra pilaris, multiple basal cell carcinomasactinic keratoses, arsenic keratoses, ichthyoses, Darriers disease, lichen planus, glucocorticoid damage, microbial infection of the skin, excessive pigmentation of the skin, and photodamage of the skin, comprising administering to said individual an effective amount of a selective CYP26A inhibitor having a formula selected from -- see Original Patent.

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