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  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
  • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
  • C07D405/06—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P13/00—Drugs for disorders of the urinary system
  • A61P13/02—Drugs for disorders of the urinary system of urine or of the urinary tract, e.g. urine acidifiers
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P15/00—Drugs for genital or sexual disorders; Contraceptives
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P25/00—Drugs for disorders of the nervous system
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/12—Antivirals
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P37/00—Drugs for immunological or allergic disorders
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P37/00—Drugs for immunological or allergic disorders
  • A61P37/08—Antiallergic agents
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D303/00—Compounds containing three-membered rings having one oxygen atom as the only ring hetero atom
  • C07D303/02—Compounds containing oxirane rings
  • C07D303/36—Compounds containing oxirane rings with hydrocarbon radicals, substituted by nitrogen atoms
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D473/00—Heterocyclic compounds containing purine ring systems
  • C07D473/02—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6
  • C07D473/04—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms
  • C07D473/06—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms with radicals containing only hydrogen and carbon atoms, attached in position 1 or 3
  • C07D473/10—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms with radicals containing only hydrogen and carbon atoms, attached in position 1 or 3 with methyl radicals in positions 3 and 7, e.g. theobromine

Definitions

• the invention relates to a class of epoxide-containing therapeutic compounds that act as drugs on the cellular and biochemical level to modulate cellular responses to noxious, proinflammatory stimuli. More specifically, the inventive compounds have at least one epoxide group on a side chain bonded to a core moiety.
• Pentoxifylline (l-(5-oxohexyl)-3,7-dimethylxanthine), abbreviated PTX, is a xanthine derivative which has seen widespread medical use for the increase of blood flow.
• PTX is disclosed in U.S. Patents 3,422,307 and 3,737,433. Metabolites of PTX were summarized in Davis et al., Applied Environment Microbiol. 48:327, 1984. A metabolite of PTX is l-(5- hydroxyhexyl)-3,7-dimethylxanthine, designated Ml. Ml was also disclosed as increasing cerebral blood flow in U.S. Patents 4,515,795 and 4,576,947.
• U.S. Patents 4,833,146 and 5,039,666 disclose use of tertiary alcohol analogs of xanthine for enhancing cerebral blood flow.
• U.S. Patent 4,636,507 describes an ability of PTX and Ml, to stimulate chemotaxis in polymorphonuclear leukocytes in response to a stimulator of chemotaxis.
• PTX and related tertiary alcohol substituted xanthines inhibit activity of certain cytokines to affect chemotaxis (U.S. Patent 4,965,271 and U.S. Patent 5,096,906).
• Administration of PTX and GM-CSF decrease tumor necrosis factor (TNF) levels in patients undergoing allogeneic bone marrow transplant (Bianco et al., Blood 76: Supplement 1 (522A), 1990). Reduction in assayable levels of TNF was accompanied by reduction in bone marrow transplant-related complications. However, in normal volunteers, TNF levels were higher among PTX recipients. Therefore, elevated levels of TNF are not the primary cause of such complications.
• TNF tumor necrosis factor
• inventive com rounds and compositions are suitable for normal routes of therapeutic administration and permit effective dosages to be provided.
• the invention is directed to epoxide-containing alkyl side chains bonded to a core moiety useful in modulating cellular response to external or in situ primary stimuli, as well as to specific modes of administration of such compounds in effective amounts.
• inventive compounds comprise epoxide-substituted alkyl side chain (R) bonded to a core moiety, comprising:
• n is an integer from about 4 to about 16 and j is an integer from about 0 to about 12, including resolved enantiomers and/or diastereomers, salts, solvates, hydrates and mixtures thereof.
• n is an integer from about 4 to about 12, more preferably from about 4 to about 10.
• J is preferably an integer from about 0 to about 3.
• the alkyl groups may also be substituted by a hydroxyl, halo or dimethylamino group and/or interrupted by an oxygen atom, H or alkyl (1-4C).
• the core moiety may be a heterocyclic or a non-heterocyclic moiety.
• a non- heterocyclic moiety is, for example, an amino acid (one or two), an hydroxyl group, a carboxyl group, a sulfoxide group, a sulfonate group, a phosphate group, an amide, an amine, a ketone, a simple ionic functional group, a terminal hydrogen or halogen atom.
• Exemplary core moiety amino acids may include one or more of the following: alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine and valine.
• the core moiety may preferably be a dipeptide comprising two amino acids selected from the foregoing exemplary list
• Exemplary halogen atoms include bromine, chlorine, fluorine and iodine.
• heterocyclic core moieties may be substituted or unsubstituted, and preferably may include, but are not limited to, substituted or unsubstituted phthalimide, homophthalimide, quinazolidinedione, quinazoline, xanthine, glutarimide, piperidine, piperidone, 7- valerolactam, cyclohexane, cyclohexene, benzene, uracil, thymine, uracil fused to napthalene, ortho-phenol, imidazole amide, pyrrole amide, benzamide, tetrahydrophthalimide or succinimide.
• Preferred heterocyclic core moieties may contain at least one ring nitrogen atom, the R side chain being bonded to a ring nitrogen.
• the heterocyclic moiety may be xanthine, phthalimide, thymine, alkyl-substituted (Cl-6) thymine, uracil, alkyl-substituted (Cl-6) uracil, glutarimide, 2,5,4'-trihalobenzophenone, 1,4- trihalomethylbenzamide (preferably the halogen groups are selected from chloro, bromo, iodo and fluoro) and resorcinol.
• the heterocyclic core moiety may be 1 ,7- methylxanthine, 8-amino-3-me_hy_xan_hine, 7-methylhypoxanthine, dimethyldihydroxypyrazolo[4,3-d]pyrimidine, methylpyrrolo[2,3-d]pyrimidine, 5- and 6- substituted uracils, 6-aminouracil, 2,4-dioxohexahydro-l,3,5-triazine, methylbarbituric acid, isocarbostyril, 1,2,3,4-tetrahydroisoquinolin, 2-hydroxypyridine, 3,3-dimethylflutarimide, 1,3-dihydroxynapthalene, 1,3-cyclopentanedione, 2-pyrrole amide, 3-pyrrole amide, 1- pyrrole amide and substituted benzamides.
• Preferable compounds of the invention having a xanthine core moiety, may include, but are not limited to, compounds having a single epoxide-substituted alkyl side chain (R) at position 7 of the xanthine nucleus.
• the invention includes a method for modulating an immune response or a cellular response to external or in situ primary stimuli comprising administering an effective amount of an inventive compound.
• the inventive compounds have been found to inhibit a specific phospholipid-based pathway that amplifies a signal within a cell. This pathway tends to be activated in response to noxious or inflammatory stimuli.
• the inventive compounds also decrease proliferation of tumor cells in response to an activated oncogene; stimulate hematopoiesis in the presence of agents which inhibit hematopoiesis, such as chemotherapeutic agents; suppress the activation of T-cells in the presence of antigen and the secretion of antibodies by B-cells in the presence of antigen; suppress the activation of macrophage or endothelial cells by endotoxins, tumor necrosis factor (TNF), interleukin-1 (IL-1) or granulocyte macrophage colony stimulating factor (GM-CSF); enhance the resistance of mesenchymal cells to TNF; inhibit the proliferation of smooth muscle cells, endothelial cells, fibroblasts and other cell types in response to growth factors, such as platelet derived growth factor (PDGF), PDGF-AA, PDGF-BB, fibroblast growth factor (FGF), epidermal growth factor (EGF), etc.; inhibit the activation of T-cells and viral replication in response to human immunodeficiency virus; inhibit
• inflammation cells block release of TNF and IL-1 in various cell types in response to inflammatory stimuli, block activation and proliferation of lymphoc y es and other cell types to IL-1 and interleukin-2 (IL-2), and the like, including the clinical manifestations of these cellular and biochemical events.
• IL-2 interleukin-2
• the inventive compounds in vitro, block IL-1 signal transduction through the Type 1 receptor as shown, for example, by preventing IL-1 and IL-1 plus PDGF (platelet derived growth factor) induction of proliferation of smooth muscle and kidney mesengial cells; 2) suppress regulation of adhesion molecules as shown, for example, by blocking NCAM in endothelial cells of CD 18 in neutrophils; 3) inhibit T ⁇ F, LPS and IL-1 induced metalloproteases (an inflammation model); 4) block LPS, T ⁇ F or IL-1 induced cellular activation (for prevention and treatment of septic shock); 5) suppress T cell and B cell antigen activation by cross-linking CD3 complex; 6) inhibit mast cell activation by IgE; and 7) suppress malignant phenotype in transformed cells and tumor cell lines.
• PDGF platelet derived growth factor
• the inventive compounds inhibit signal transduction mediated through the Type I IL-1 receptor, and are therefore considered as IL-1 antagonists.
• Dinarello and Wolff "The Role of Interleukin-1 in Disease," N. Engl. J. Me ⁇ 328, 106 (Jan. 14, 1993), describe the role of IL-1 as "an important rapid and direct determinant of disease.”
• IL-1 acts directly on the blood vessels to induce vasodilatation through the rapid production of platelet activating factor and nitric oxide, whereas in autoimmune disease it acts by stimulating other cells to produce cytokines or enzymes that then act on the target tissue.” Ibid.
• the article describes a group of diseases mediated by IL-1, including many of the foregoing diseases.
• the invention is directed to a pharmaceutical composition
• a pharmaceutical composition comprising an inventive compound and an effective amount of an agent which reduces the activity of the enzyme P-450, such as a quinolone, to increase the pharmacokinetic half-life of an inventive compound.
• Figure 1 shows a mixed lymphocyte reaction of three inventive compounds CT1605 ( ⁇ -(5,6-oxidohexyl) glutarimide), CT1808 (N3-(5,6-oxidohexyl)-Nl-methyluracil), and CT1906 (N ⁇ -(5,6-oxidohexyl) N ⁇ -methylthymine).
• the mixed lymphocyte reaction shows a proliferative response of PBMC (peripheral blood mononuclear cells) to allogeneic stimulation determined in a two-way mixed lymphocyte reaction.
• PBMC peripheral blood mononuclear cells
• Each of the inventive compounds tested was effective (and more potent than PTX although not shown on this graph) in this immune modulating activity assay procedure.
• Figure 2 shows a comparison of three dose levels of CT1808 and CT1906 and no drug control to inhibit thymocyte proliferation.
• the thymocytes were obtained from normal female Balb/C mice and stimulated with Concanavalin A (Con A) and/or interleukin-1 alpha (L-l ⁇ ). Drugs were added to the cell cultures two hours before activation with Con A and/or IL-l ⁇ . As shown in Figure 2, both drugs inhibited thymocyte proliferation is a dose- dependent fashion.
• Figure 3 shows a comparison of CT1605 and CT1808 on inhibition of B-cell proliferation.
• a Ramos C-cell tumor line was treated with 250 ⁇ M CT1808 or CT1605 for one hour prior to stimulation of proliferation with anti-mu antibody or phorbol myristic acid (PMA, 5 nM).
• PMA phorbol myristic acid
• FIG. 4 shows a comparison of CT1605, CT1808 and CT1906 on PDGF-induced (platelet derived growth factor) proliferation of human stromal cells.
• Human stromal cells were starved in serum-free media for 24 hours and then stimulated with 50 ng ml of PDGF- BB. The drugs were added at various indicated concentrations one hour prior to PDGF stimulation.
• Tritiated thymidine was added for 24 hrs at the time of PDGF stimulation to measure cellular proliferation. Background counts were approximately 5% of control levels. All three drugs inhibited PDGF-induced stimulation in a dose response fashion.
• Figure 5 shows the effect of CT1605, CT1808 and CT1906 to inhibit adhesion of U937 cells to activated human umbilical vein endothelial cells (HUNEC).
• HUNEC cells were activated with 20 ng/ml of T ⁇ F for 12 hrs. Drug was added to each culture (except for controls) one hour prior to adding T ⁇ F.
• Figure 6 shows the effects of CT1605, CT1808 and CT1906 to inhibit cell surface expression of VC AM in human umbilical vein endothelial cells (HUNEC).
• HUNEC human umbilical vein endothelial cells
• the HUNEC cells were stimulated with 20 ng/ml T ⁇ F- ⁇ for 20 hrs and then stained for immunofluorescence using a monoclonal antibody recognizing NCAM, followed by a goat anti-mouse antibody conjugated to phycoerythrin. The cells were analyzed for antibody binding using flow cytometry.
• Figure 6 shows an analysis of mean relative fluorescence intensity of 10,000 cells, analyzed by flow cytometry. The mean fluorescence levels were decreased by all three drugs from control levels (T ⁇ F treatment, no drug). Detailed Description of the Invention
• the invention is directed to a defined genus of inventive compounds which can control cellular behavior by a particular phase of a secondary messenger pathway system (Bursten et al., J. Biol. Chern. 266:20732, 1991).
• the second messengers are lipids or phospholipids and use the following abbreviations:
• PE phosphatidyl ethanolamine
• LPA lysophosphatidic acid
• DAG diacylglycerol
• LPLD lysophospholipase-D
• LPAAT lysophosphatidic acid acyl transferase
• PAPH phosphatidic acid phosphohydrolase
• PLA2 phospholipase A-2.
• PAA phosphoarachidonic acid
• PLA-2 phospholipase A2
• PC phosphatidyl choline
• PA, cyclic pathway PAA, LPA, PA and DAG intermediates substituted with L-saturated, 2-linoleoyl or l,2-dileolyl/l,2-sn-dilinoleoyl at the indicated sn- 1 and sn-2 positions.
• Classical PI Pathway PI, DAG, PA intermediates substituted with 1-stearoyl, 2- arachidonoyl fatty acyl side chains. ' -'
• PLD-generated PA PE, PC, LPA, PA and DAG intermediates substituted with, e.g., 1,2-sn-dioleoyl-, 1-alkyl, 2-linoleoyl-, and 1-alkyl, 2-docosahexaneoyl-side chains.
• Lysophosphatidic acid transferase effects the synthesis of phosphatidic acid (PA) from lysophosphatidic acid (LPA) by incorporation of an acyl group from acyl CoA. Hydrolysis of the phosphate moiety by PA phosphohydrolase (PAPH) results in the formation of DAG.
• PA phosphatidic acid
• PAPH PA phosphohydrolase
• the compounds of the invention include inhibitors of subspecies of LPAAT in PAPH enzymes with substrate specificity for intermediates with 1,2-dmnsaturated and 1- alkyl, 2-unsaturated subspecies.
• One representative example of such an inhibitor is PTX.
• PTX blocks PAPH in a specific activation pathway that does not involve PI but rather derives from a PA that is largely composed of 1,2-diunsaturated and l-alkyl,2-unsaturated subspecies. This was shown, for example, by the demonstration that human mesangial cells stimulated with TNF produce DAG from PI and regenerate PI in the absence and the presence of PTX.
• inventive compounds comprise epoxide-substituted alkyl side chain (R) bonded to a core moiety, comprising:
• n is an integer from about 4 to about 16 and j is an integer from about 0 to about 12, including resolved enantiomers and/or diastereomers, salts, solvates, hydrates and mixtures thereof.
• n is an integer from about 4 to about 12, more preferably from about 4 to about 10.
• J is preferably an integer from about 0 to about 3.
• the alkyl groups may also be substituted by a hydroxyl, halo or dimethylamino group and/or interrupted by an oxygen atom, H or alkyl (1-4C).
• the core moiety may be a heterocyclic or a non-heterocyclic moiety.
• a non- heterocyclic moiety is, for example, an amino acid (one or two), an hydroxyl group, a carboxyl group, a sulfoxide group, a sulfonate group, a phosphate group, an amide, an amine, a ketone, a simple ionic functional group, a terminal hjfdrogen or halogen atom.
• Exemplary core moiety amino acids may include one or more of the following: alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine and valine.
• the core moiety may preferably be a dipeptide comprising two amino acids selected from the foregoing exemplary list.
• Exemplary halogen atoms include, but are not limited to, bromine, chlorine, fluorine and iodine.
• heterocyclic core moieties may be substituted or unsubstituted, and preferably may include, but are not limited to, substituted or unsubstituted phthalimide, homophthalimide, quinazolidinedione, quinazoline, xanthine, glutarimide, piperidine, piperidone, 7- valerolactam, cyclohexane, cyclohexene, benzene, uracil, thymine, uracil fused to napthalene, ortho-phenol, imidazole amide, pyrrole amide, benzamide, tetrahydrophthalimide or succinimide.
• Preferred heterocyclic core moieties may contain at least one ring nitrogen atom, the R side chain being bonded to a ring nitrogen.
• the heterocyclic moiety may be xanthine, phthalimide, thymine, alkyl-substituted (Cl-6) thymine, uracil, alkyl-substituted (Cl-6) uracil, glutarimide, 2,5,4'-trihalobenzophenone, 1,4- trihalomethylbenzamide (preferably the halogen groups are selected from chloro, bromo, iodo and fluoro) and resorcinol.
• the heterocyclic core moiety may be 1,7- methylxanthine, 8-amino-3-methylxanthine, 7-methylhypoxanthine, dimethyldihydroxypyrazolo[4,3-d]pyrimidine, methylpyrrolo[2,3-d]pyrimidine, 5- and 6- substituted uracils, 6-aminouracil, 2,4-dioxohexahydro-l,3,5-triazine, methylbarbituric acid, isocarbostyril, 1,2,3,4-tetrahydroisoquinolin, 2-hydroxypyridine, 3,3-dimethylflutarimide, 1,3-dihydroxynapthalene, 1,3-cyclopentanedione, 2-pyrrole amide, 3-pyrrole amide, 1- pyrrole amide and substituted benzamides.
• Preferable compounds of the invention having a xanthine core moiety, may include, but are not limited to, compounds having a single epoxide-substituted alkyl side chain (R) at position 1 of the xanthine nucleus.
• inventive compounds may be provided as enantiomeric or diastereomeric mixtures or in resolved or partially resolved forms. Standard procedures are used for resolution of optical isomers. It is contemplated that the different enantiomeric variants (e.g., stereoisomers and chiral forms) of the epoxide-containing compounds will have different drug activities, based upon their differential ability to inhibit PAPH and LPAAT.
• An optical isomer substantially free of the corresponding enantiomer and/or diastereomers has at least about 85% relevant optical isomer, preferably at least about 95% relevant optical isomer and especially at least about 99% or higher relevant optical isomer, but most preferably where the amount of other optical forms is undetectable.
• the invention further comprises a pharmaceutical composition comprising one or a plurality of inventive compounds and a pharmaceutically acceptable carrier or excipient.
• the cells to be treated with an inventive compound or inventive pharmaceutical composition may either be contacted with the compound of the invention in in vitro culture, in an extracorporeal treatment, or by administering the compound of the invention or pharmaceutical composition thereof to a subject whose cells are to be treated.
• Illustrative compounds of the invention include both racemic mixture and R and S enantiomers of the following compounds (designated accordingly as R and S) shown in Table
• CT1103 N-(5,6-Oxidohexyl)phthalimide CT1105 N-(8,9-Oxidonony_)phtha_imide CT1109 N-(10,l l-Oxidoundecyl)phthalimide CT1114 N-(10,l l-Oxidoundecyl)homophthalimide CT1206 1 -(5 ,6-Oxidohexyl)-3-methylbenzoyleneurea CT1301 N-(5,6-Oxidohexylamido)glutaric acid, methyl ester CT1409 l-(8,9-Oxidononyl)-3-methyl-7-methylpivaloylx__nth_ne CT1410 1 -(5 ,6-Oxidononyl)-3-methyl-7-methylpivaloylxanthine CT1412 1-(1 l,10-Oxidounde
• the compounds of the invention provide a mechanism to maintain homeostasis in cells contacted by primary stimuli through mitigating the effects of these primary stimuli on the secondary signaling pathways invoked within seconds of the primary stimulus.
• inventive pharmaceutical compositions comprising an effective amount of at least one of the inventive compounds or a pharmaceutically acceptable salt, hydrate or solvate thereof and at least one pharmaceutically acceptable excipient or carrier.
• the inventive pharmaceutical compositions are useful for: 1) protecting and treating endotoxic shock and sepsis induced by gram positive or negative bacteria; 2) inhibiting, treating or preventing tumor cell growth, such as cancer; 3) stimulating hematopoiesis inhibited by cytoreductive therapies (e.g., chemotherapy or radiotherapy); 4) treating or preventing autoimmune diseases, such as insulin dependent diabetes mellitus (IDDM), arthritis (including rheumatoid arthritis), multiple scherosis, Alzheimers disease, glomerular nephritis, Graves disease, and atheroschlerosis; 5) treating or preventing male pattern baldness by stimulation of hair growth through reversal of an apoptotic process; 6) preventing hair loss caused by cytoreductive therapies; 7) preventing the symptoms of ARDS (acute respiratory distress syndrome) caused by trauma; 8) treating or preventing
• IDDM insulin dependent diabetes mellitus
• arthritis including rheumatoid arthritis
• multiple scherosis Alzheimers disease
• TNF tumor necrosis factor
• rheumatoid arthritis rheumatoid spondylitis
• osteoarthritis gouty arthritis and other arthritic conditions
• sepsis septic shock, endotoxic shock, gram negative sepsis, toxic shock syndrome, adult respiratory distress syndrome, cerebral malaria, chronic pulmonary inflammatory disease, silicosis, pulmonary sarcoidosis, bone resorption diseases, reperfusion injury, graft versus host reaction, allograft rejections, fever, myalgias due to infection such as influenza, cachexia secondary to infection, AIDS or malignancy, AIDS, other viral infections (e.g., CMV, influenza, adenovirus, herpes family), keloid formation, scar tissue formation, Crohn's disease, ulcerative colitis, or pyresis.
• CMV CMV
• influenza adenovirus
• herpes family keloid formation
• scar tissue formation Crohn's disease
• inventive compounds or pharmaceutically acceptable salts thereof can be used in the manufacture of a medicament for the prophylactic or therapeutic treatment of any disease state in a human or other mammal, which is exacerbated or signaled 1) through the specific phospholipid-based messenger pathway that amplifies signals within a cell, and 2) by excessive or unregulated-production of messenger inflammatory cytokines such as TNF or IL-1.
• TNF messenger signaling there are several disease states in which excessive or unregulated monocyte/macrophage TNF production exacerbates or causes the disease. These include, for example, neurodegenerative diseases such as Alzheimers disease, endotoxemia or toxic shock syndrome (Tracey et al., Nature 330:662, 1987 and Hinshaw et al., Circ.
• inventive compounds may be used topically in the treatment of prophylaxis of topical disease states mediated or exacerbated by excessive TNF or IL-1, such as viral infections (herpes or viral conjunctivitis), psoriasis, fungal or yeast infections
• the inventive compounds may preferably be used to stimulate hematopoiesis, prevent and treat septic shock, treat acute and chronic inflammatory disease, treat or prevent an autoimmune disease, treat a fungal or yeast infection, and stimulate hair growth (when applied topically, as confirmed in in vivo results on nude mice).
• the inventive compounds are useful as an adjuvant to inhibit toxic side effects of drugs. These side effects include, for example, side effects of: 1) interleukin-2 (EL-2); 2) cyclosporin A and FK506; and 3) amphotericin B.
• inventive compounds also inhibit antigen-induced T cell activation, like cyclosporin or FK506, but, unlike cyclosporin or FK506, do not: 1) prevent generation of ⁇ K and LAK cells; 2) suppress IL-2 release from T cells; or 3) suppress IL-8 release.
• Metalloproteases mediate tissue damage such as glomerular diseases of the kidney, joint destruction in arthritis, and lung destruction in emphysema, and play a role in tumor metastases.
• Three examples of metalloproteases include a 92 kD type V gelatinase induced by T ⁇ F, IL-1 and PDGF plus bFGF, a 72 kD type IV collagenase that is usually constitutive and induced by TNF or IL-1, and a stromelysin/PUMP-1 induced by TNF and IL-1.
• the inventive compounds can inhibit TNF or IL-1 induction of the 92 kD type V gelatinase inducable metalloprotease.
• inventive compounds can reduce PUMP-1 activity induced by 100 U/ml of IL-1. Accordingly, the inventive compounds prevent induction of certain metalloproteases induced by IL-1 or TNF and are not involved with constitutively produced proteases (e.g., 72 kD type IV collagenase) involved in normal tissue remodeling.
• constitutively produced proteases e.g., 72 kD type IV collagenase
• the compounds of the invention are used in connection with patients undergoing bone marrow transplantation (BMT), regardless of whether the BMT is matched allogeneic, mismatched allogeneic, or autologous.
• BMT bone marrow transplantation
• Patients receiving autologous transplants are aided by treatment with compounds of the invention even though they do not necessarily need to be administered immunosuppressive agents, since they do not develop graft-versus- host disease (GVHD).
• GVHD graft-versus- host disease
• the toxic effect of the chemotherapy or radiation therapy used in connection with the disease, in response to which the transplantation has been performed constitutes a negative stimulus with regard to the patients' cells.
• the inventive compounds are able to increase the percentage of patients who survive.
• the percentage of fatalities within the first 100 days that is considered acceptable is 15-20% for "good risk” patients and 30-40% for "high risk”. These fatalities are due to the direct effects of high doses of chemo/radiation.
• GVHD contributes to the death rate in allogeneic marrow recipients.
• a tumor burden a hormone-related disorder, a neurological disorder, an autoimmune disease, inflammation, restenosis, hypertension, unwanted immune response, viral infection, nephritis, mucositis, and various allergic responses.
• Prevention of allergic responses include prevention of acute allergic response and thus moderation or prevention of rhinorrhea, serious drainage, diffuse tissue edema, and generalized pruritus.
• Other symptoms of chronic allergic response include, as well as the foregoing, dizziness, diarrhea, tissue hyperemia, and lacrimal swelling with localized lymphocyte infiltration.
• Allergic reactions are also associated with leukotriene release and the distal effects thereof, including asthmatic symptoms including development of airway obstruction, a decrease in FEVl, changes in vital capacity, and extensive mucus production.
• Suitable subjects for the administration of compounds of the invention include patients being administered toxic agents for the treatment of tumors, such as chemotherapeutic agents or irradiation therapy, as well as treatment with biological response modifiers such as
• tumor suppressing cells such as lymphokine activated killer cells (LAK) and tumor- infiltrating lymphocytes (TIL cells); patients suffering from neoplasias generally, whether or not otherwise treated including acute and chronic myelogenous leukemia, hairy cell leukemia, lymphomas, megakaryocytic leukemia, and the like; disease states caused by bacterial, fungal, protozoal, or viral infection; patients exhibiting unwanted smooth muscle cell proliferation in the form of, for example, restenosis, such as patients undergoing cardiac surgery; patients who are afflicted with autoimmune diseases, thus requiring deactivation of T and B cells, and patients who have neurological disorders.
• LAK lymphokine activated killer cells
• TIL cells tumor- infiltrating lymphocytes
• the compounds of the invention further are able to decrease the enhanced levels of a relevant PA and DAG resulting from stimulation of synaptosomes with acetylcholine and/or epinephrine. This suggests that the effects of the compounds of the invention are to both enhance the release of inhibitory neural transmitters such as dopamine, and to modulate the distal "slow current" effects of such neurotransmitters.
• the drugs of the invention are also useful to raise the seizure threshold, to stabilize synapses against neurotoxins such as strichnine, to potentiate the effect of anti- Parkinson drugs such as L-dopa, to potentiate the effects of soporific compounds, to relieve motion disorders resulting from administration of tranquilizers, and to diminish or prevent neuron overfiring associated with progressive neural death following cerebral vascular events such as stroke.
• the compounds of the invention are useful in the treatment of norepinephrine-deficient depression and depressions associated with the release of endogenous glucocorticoids, to prevent the toxicity to the central nervous system of dexamethasone or methylprednisolone, and to treat chronic pain without addiction to the drug.
• the compounds of the invention are useful in the treatment of children with learning and attention deficits and generally improve memory in subjects with organic deficits, including Alzheimer's patients.
• a particularly preferred regimen for use in treating leukemia is 4-50 mg/kg body weight. It is to be understood, however, that for any particular subject, specific dosage regimens should be adjusted to the individual's need and to the professional judgment of the person administering or supervising the administration of the inventive compounds.
• NIH3T3-D5C3 cells can be used to compare effects of an inventive compound alone or in combination with a P-450 inhibitor by comparing transformation phenotype control, incubation with an inventive compound, and coincubation of an inventive compound with the P-450 enzyme inhibitor.
• Compounds that inhibit P-450 include, for example, (mg range daily dosage) propranolol (20-100), metaprolol (20-100); verapamil (100-400), diltiazem (100-400), nifedipine (60-100); cimetidine (400-2,400); ciprofloxacin (500-2000), enoxacin (500- 2,000), norfloxacin (500-2000), ofloxacin (500-2,000), pefloxacin (500-2,000); erythromycin (100-1,000), troleandomycin (100-1,000); ketoconizole (100-2,000), thiabenzadole (100- 1,000); isoniazid (100-1000); mexiletine (100-1,000); and dexamethasone (1-100 mg).
• the compounds of the invention and a P-450 inhibitor can be administered individually or in a single composition.
• a suitable formulation will depend on the nature of the disorder to be treated, the nature of the medicament chosen, and the judgment of the attending physician.
• the inventive compounds are formulated either for injection or oral administration, although other modes of administration such as transmucosal or transdermal routes may be employed. Suitable formulations for these compounds can be found, for example, in Remington's Pharmaceutical Sciences (latest edition), Mack Publishing Company, Easton, PA.
• the level of dosage can be appreciably diminished by coadministration of a P-450 inhibitor, such as a quinolone.
• a P-450 inhibitor such as a quinolone.
• a strong synergistic effect may be obtained with such a quinolone.
• PTX means pentoxifylline.
• This example illustrates a synthesis of N-(5,6-Oxidohexyl)phthalimide (CT1103).
• 1- bromo-5-hexene (6.52 g, 40 mmol) was added to a potassium phthalimide (7.4 g, 40 mmol) suspension in 50 mL of dimethyl sulfoxide and stirred overnight. After 12 hours of stirring at room temperature, the reaction was poured into a separatory funnel containing 300 mL of water and extracted with dichloromethane (5 X 200 mL). The organic extracts were combined, washed with water (100 mL) and brine (100 mL), dried over anhydrous magnesium sulfate and concentrated under reduced pressure.
• This example illustrates a synthesis of N-(9,8-Oxidononyl)phthalimide (CT1105).
• 1- bromo-8-nonene (8.2 g, 40 mmol) was added to a suspension of potassium phthalimide (7.4 g, 40 mmol) in 50 mL of dimethyl sulfoxide and stirred overnight. After 12 hours of stirring at room temperature, the reaction product was poured into a separatory funnel containing 300 mL of water and extracted with dichloromethane (5 X 200 mL). Organic extracts were combined, washed with water (100 mL) and brine (100 mL), dried over anhydrous magnesium sulfate and concentrated under reduced pressure.
• N-(10,11-Oxidoundecyl)phthalimide (CT1109).
• the mixture was then poured into water (80 mL) and extracted with ethyl acetate (3 x 70 mL). The combined extracted organic portions were washed with water (3 x 100 mL), dried with magnesium sulfate and evaporated, resulting in a cream-colored solid.
• Purification by column chromatography (using ethyl acetate/hexane) yielded (5.50g, 86%) N-(10- Undecenyl)phthalimide (CT-1107) as a white solid.
• N-(10-Undecenyl)phthalimide (4.97 g, 16.6 mmol), prepared as discribed above, 4-methylmorpholine-N-oxide (8.62 mL, 60% by wt in water, 50.0 mmol) and potassium osmate dihydrate (58 mg, 0.16 mmol) in 100 mL acetone/water (1:1 by wt) was stirred for 16 hours. Water (100 mL)'and sodium sulfite (10 g) were added and the resulting solution stirred for an additional hour.
• N-(10,11-Dihydroxyundecyl)phthalimide (2.35 g, 7.10 mmol) was stirred for 3 hours with HBr (6.90 mL of a 30% solution in acetic acid, 21.3 mmol). The mixture was then added over 10 minutes to a solution of water (50 mL), ice (25 g) and NaHCO3 (15 g) and stirred for an additional 30 min. The resulting reaction product was extracted with dichloromethane (3 x 70 mL) and the combined organic phase was dried using magnesium sulfate and evaporated, yielding a residue of N-(10-acetoxy-ll-bromoundecyl)phthalimide.
• this crude product was treated in methanol (10 mL) with a solution of sodium methoxide 1 (prepared from sodium — 0.23 g, 10.0 mmol — and 10 mL methanol). After 60 minutes the reaction mixture containing treated crude product was added to water (30 mL) and extracted with dichloromethane (100 mL ' , 2 x 50 mL). The extracted organic portions were combined, dried and evaporated, yielding 2.00 g (89% yield) N-(10,11-Oxidoundecyl)phthalimide (CT1109) as a white solid.
• CT1109 N-(10,11-Oxidoundecyl)phthalimide
• This example illustrates a synthesis of N-( 10,11 -Oxidoundecyl)homophthalimide (CT1114).
• a mixture of homophthalic acid (54.0 g; 0.3 mole) and finely powdered urea (19.82 g; 0.33 mole) were heated to 175-185 ⁇ C until no more ammonia evolves, evidenced by pH paper.
• the crude product was refluxed with methanol (500 mL) and homophthalimide isolated by filteration (29g; 60%).
• Sodium hydride(95%) (576 mg, 24 mmol) was added to a solution of homophthalimide (3.2 g, 20 mmol) in anhydrous dimethylsulfoxide (75 mL).
• This example illustrates a synthesis of l-(5,6-Oxidohexyl)-3-methylbenzoyleneurea (CT1206).
• Sodium metal 0.071 g, 3.1 mmol
• methanol 3.1 mL
• l-(5-Acetoxy-6-bromohexyl)-3- methylbenzoyleneurea (1.17 g, 2.9 mmol) was dissolved into methanol ( 25 mL) and added to the sodium methoxide solution over 5 minutes. After stirring for 1 hour, 50 mL of water were added to the solution.
• This example illustrates a synthesis of 3-Methyl-7-methylpivaloyl-l-(8,9- oxidononyl)xanthine (CT1409).
• a mixture of 3-methylxanthine (Aldrich, 1.00 g, 6.0 mmol), sodium hydride (145 mg, 6.0 mmol) and dimethyl sulfoxide (20 mL) was stirred until homogeneous (0.5 hours).
• Chloromethylpivalate (865mL, 904 mg, 6.0 mmol) was added and the reaction stirred for 18 hours.
• the reaction mixture was poured into water (70 mL) and then extracted with 25% ethanol/dichloromethane (4 X 60 mL).
• This example illustrates a synthesis of l-(8,9-Oxidohexyl)-3-methyl-7- methylpivaloylxanthine (CT1410).
• a mixture of 3-methylxanthine (Aldrich, 1.00 g, 6.0 mmol), sodium hydride (145 mg, 6.0 mmol) and dimethyl sulfoxide (20 mL) was stirred until homogeneous (0.5 hours).
• Chloromethylpivalate (865mL, 904 ipg, 6.0 mmol) was added and the reaction stirred for 18 hours.
• the reaction mixture was poured into water (70 mL) and then extracted with 25% ethanol dichloromethane (4 X 60 mL).
• Example 8 This example illustrates a synthesis of 1-(1 l,10-Oxidoundecanyl)-3-methyl-7- methylpivalylxanthine (CT1412).
• Chloromethylpivalate (865mL, 904 mg, 6.0 mmol) was added and the reaction stirred for 18 hours.
• the reaction mixture was poured into water (70 mL) and then extracted with 25% ethanol/dichloromethane (4 X 60 mL).
• the crude product obtained was further purified by flash chromatography over silica gel using 50% hexane/ethyl acetate eluant to yield 1.05 g (73.8% yield) l-(10-undecenyl)-7-methylpivaloyl-3-methylxanthine (CT1403).
• This example illustrates a synthesis of l-(l l,10-Oxidoundecanyl)-3-methylxanthine (CT1413).
• the reaction mixture was quenched with saturated ammonium chloride solution (5 mL) and extracted with 20% ethanol/dichloromethane (3x30 mL).
• This example illustrates a synthesis of 7-(l l,10-Oxidondecyl)-l,3-dimethylxanthine (CT-1423).
• Sodium hydride(95%) 0.575 g, 24 mmol was added to a solution of theophylline (3.6 g, 20 mmol) in dimethylsulfoxide (100 mL).
• l-bromoundec-10-ene (4.66 g, 20 mmol) was added and stirred for 12 hours at room temperature.
• the reaction mixture was then poured into a separatory funnel containing water (300 mL) and extracted with dichloromethane (5 X 100 mL).
• This example illustrates a synthesis of 7-(l l,10-Oxidondecyl)-l-methyl-2,4- dioxotetrahydropteridine (CT1426).
• l-Methy_-4,5-diaminouracil (13.6 g; 59.4 mole) was suspended in water (150 mL) and converted to its hydrochloride by drop- wise addition of concentrated hydrochloric acid unitl the solution is strongly acidic.
• Glyoxan sodiumbisulphite (20.4 g; 71.8 mmol) was then added and the reaction mixture refluxed for 30 minutes.
• the reaction mixture was then poured into a separatory funnel containing water (300 mL) and extracted with ethyl acetate (5 X 100 mL). The organic extracts were combined, washed with water (100 mL) and brine (100 mL), dried over anhydrous magnesium sulfate and concentrated under reduced pressure.
• the crude product obtained was further purified by flash chromatography over silica gel using 50% hexane/ethyl acetate eluant to yield 5 g (94% yield) of 3-(10-undecenyl)-l-methyl-2,4-dioxotetrahydropteridine (CT1421).
• Example 12 This example illustrates a synthesis of l-Methyl-3-(5,6-oxidohexyl)xanthine
• This example illustrates a synthesis of l-(6,7- _.-Oxidononyl)-3,7-dimethylxanthine (CT-1509).
• This example illustrates a synthesis of l-(5,6-Oxidohexyl)-3,7-dimethylxanthine (CT1541).
• CTL1541 l-(5,6-Oxidohexyl)-3,7-dimethylxanthine
• the solution was treated with water (200 mL) and extracted with dichloromethane (3 X 80 mL).
• This example illustrates a synthesis of l-(8,9-Oxidooctyl)-3,7-dimethy_xanthine (CT1553).
• sodium hydride 580 mg, 24.2 mmol
• dimethylsulfoxide 100 mL
• 8-bromo-l-octene 3.96 g, 22 mmol
• the mixture was then poured into water (200 mL) and extracted with dichloromethane (3 X 50 mL).
• This example illustrates a synthesis of l-(4,5-Oxohexyl)-3,7-d_methylxanthine (CT1555).
• CTL1555 l-(4,5-Oxohexyl)-3,7-d_methylxanthine
• This example illustrates a synthesis of l-(9,10-Oxidodecyl)-3,7-dimethylxanthine (CT1565).
• CTD5 9-decene-l-ol
• dichloromethane 100 mL
• methanesulfonyl chloride 2.20 g, 1.5 mL, 19.2 mmol
• triethylamine 2.91 g, 28.8 mmol
• This example illustrates a synthesis of 3,7-dimethyl-l-(6,7-tr_-7-_;-ox_donony_)xanthine (CT-1569).
• a mixture of 6-ri_:-nonen-l-ol (TCI, 990 mg, 7.0 mmol) and thiophenol (60 mg) was heated at 105-110°C under argon for 4 hours to give 6-nonen-l-ol 872 mg, 88% yield) with a 4:1 trans:cis isomer ratio.
• the mixture was stirred with methanesulfonyl chloride (694 mg, 6.1 mmol) in dichloromethane (20 mL) at 0°C.
• Triethylamine (925 mg, 9.2 mg) was added dropwise and stirring continued for 1 hour.
• the reaction mixture was added to an aqueous saturated solution of sodium bicarbonate (10 mL) and the layers were separated.
• the aqueous layer was extracted with dichloromethane (2 x 15 mL).
• the combined organic layers were washed with a 5% solution of hydrogen chloride (10 mL), water (10 mL), and an aqueous saturated solution of sodium chloride (10 mL) and then dried over sodium sulfate.
• the solvent was removed under vacuum to give the mesylate, which was used in the next step without purification.
• This example illustrates a synthesis of l-(6,7-Oxidoheptyl)-3,7-dimethylxanthine (CT- 1586).
• 6-hepten-l-ol (6.00 g, 52.6 mmol) in dichloromethane (120 mL) at 0° C was added methanesulfonyl chloride (6.07 g, 4.0 mL , 53.0 mmol), followed by triethylamine (7.79 g, 77.0 mmol). After stirring for 10 minutes at 0°C, the reaction was allowed to warm to 25°C and then stirred for 2 hours.
• This example illustrates a synthesis of l-(3-(R)-methyl-7-methyl-6,7-oxidooctyl)-3,7- dimethylxanthine (CT1588R).
• Sodium hydride(95%) (631 mg, 25 mmol) was added to a solution of theobromine (4.14g, 23 mmol) in dimethylsulfoxide (75 mL).
• (R)(-)Citronellyl bromide 5.0 g, 22.8 mmol
• Example 22 This example illustrates a synthesis of l-(3-(S)-methyl-7-methyl-6,7-oxidooctyl)-3,7- dimethylxanthine (CT1588S).
• the crude product obtained was further purified by flash chromatography over silica gel using 30% petroleum ether/ethyl acetate eluant to yield 5.7 g (80% yield) l-(3-(S)-methyl-7-methyloct-6-enyl)-3,7- dimethylxanthine (CT1596S) as an yellow oil.
• This example illustrates a synthesis of l-(4,5-Oxipentyl)-3,7- dimethylxanthine.
• Sodium hydride (95%) (1.38 g, 55 mmol) was added to a solution of theobromine (9.0 g, 50 mmol) in dimethylsulfoxide (300 mL).
• l-bromo-4-pentene (1 7.45 g, 50 mmol) was added.
• the reaction was poured into a separatory funnel containing 1 L of water and extracted with dichloromethane (5 X 200 mL).
• This example illustrates a synthesis of l-(10,l l-Oxidoundecanyl)-3,7- dimethylxanthine (CT1594).
• Sodium hydride(95%) (1.26 g, 50 mmol) was added to a solution of theobromine (7.2g, 40 mmol) in dimethylsulfoxide (300 mL).
• undecenylmesylate (7.95 g, 30 mmol) was added and stirred for 12 hours at room temperature.
• the reaction was warmed to 70-80°C and stirred for 4 hours.
• the reaction mixture was then poured into a separatory funnel containing 1 L of water and extracted with dichloromethane (5 X 200 mL).
• This example illustrates a synthesis of l-(5,6-Oxidohexyl)glutarimide (CT-1605M).
• Sodium hydride (425 mg, 17.7 mmol) was added to a solution of glutarimide (2.00 g, 7.7 mmol) in dimethyl sulfoxide (40 mL).
• 6-bromo-l-hexene (2.90 g, 17.7 mmol) is added.
• the reaction was poured into a separatory funnel containing 100 mL water and extracted with dichlormethane (4 X 50 mL).
• This example illustrates a synthesis of N-(8,9-Oxidononyl)glutarimide (CT1606).
• Sodium hydride (1.02 g, 44 mmol) was added to a solution of glutarimide (5.00 g, 44 mmol) in dimethyl sulfoxide (150 mL).
• 9-bromo-l-nonene (9.02 g, 44 mmol) was added.
• the reaction was poured into a separatory funnel containing 100 mL water and extracted with dichlormethane (3 X 70 mL).
• This example illustrates a synthesis of N-(11,10-Oxidoundecyl)glutarimide (CT1611).
• Sodium hydride (95%) (168 mg, 7 mmol) was added to a solution of glutarimide (565.6 mg, 5 mmol) in dimethyl sulfoxide (15 mL).
• l-bromundec-10-ene (1.165 g, 5 mmol) was added and stirred for 12 hours at room temperature.
• the reaction mixture was then poured into a separatory funnel containing 100 mL of water and extracted with dichlormethane (5 X 75 mL).
• This example illustrates a synthesis of N-( 10,11 -Oxidoundecyl)-2-piperidone (CT1618).
• a mixture of potassium hydroxide (1.55 g, 25 mmol, pellets ground in mortar and pestle) and tetrabutylammonium bromide (1.61 g, 5.0 mmol) was stirred in dry tetrahydrofuran (10 mL).
• a solution of d-valerolactam (2.5 g, 25 mmol) and l-bromo-10- undecene (Lancaster, 5.9 g, 25 mmol) in tetrahydrofuran (15 mL) was added by syringe pump over 1 hour.
• This example illustrates a synthesis of l-Methyl-3-(8,9-oxidononyl)uracil (CT1804).
• Sodium hydride (365 mg, 16 mmol) was added to a stirring solution of 1-methyluracil (2.00 g, 16 mmol) in dimethyl sulfoxide (40 mL).
• 6-bromo-l-nonene (3.26 g, 16 mmol) was added and the mixture stirred for 3 days.
• the reaction was then poured into water (50 mL) and extracted with dichloromethane (3 X 60 mL).
• This example illustrates a synthesis of 3-(5,6-oxidohexyl)-2-methyldihydrouracil (CT1820).
• Sodium hydride (288 mg, 12 mmol) was added to a solution of N- methylhydrouracil (1.54 g, 12 mmol) and l-bromo-5-hexene (1.63g, 10 mmol) in 20 mL of dimethyl sulfoxide at room temperature and stirred for 12 hours.
• the reaction mixture was then quenched with water (80 mL) and extracted with dichloromethane (3x100 mL).
• the combined organic extract was washed with saturated aqueous salt solution solution (50 mL), dried over anhydrous magnesium sulfate and concentrated under reduced pressure.
• This example illustrates a synthesis of 3-(10,l l-Oxidoundecanyl)-l- methylhydrouracil (CT-1822).
• Sodium hydride (288 mg, 12 mmol) was added to a solution of N-methylhydrouracil (1.54 g, 12 mmol) and l-bromo-10-undecene (2.33 g, 10 mmol) in 20 mL of dimethyl sulfoxide at room temperature and stirred for 12 hours.
• the reaction mixture was then quenched with water (80 mL) and extracted with dichloromethane (3x100 mL).
• Example 34 This example illustrates a synthesis of 3-(5,6-Oxidohexyl)-l-methylthymine
• This example illustrates a synthesis of l-Methyl-3-(8,9-oxidononyl)thymine (CT1910).
• Sodium hydride (343 mg, 14 mmol) was added to a stirring solution of 1- methylthymine (2.00 g, 14 mmol) in dimethylsulfoxide (40 mL).
• 9- bromo-1-nonene (2.93 g, 14 mmol) was added and the mixture stirred for 20 hours.
• the reaction was poured into 40 mL water and extracted with dichloromethane (3 X 50 mL). The organic layers were combined, washed with water (40 mL), saturated aqueous salt solution (20 mL), and dried over sodium sulfate.
• the solvent was evaporated to yield 2.76 g (73% yield) l-Methyl-3-(8-nonenyl)thymine (CT1917) as a colorless oil which solidified upon standing.
• This example illustrates a synthesis of 3-(l l,10-Oxidoundecyl)-l-methylthymine (CT1932).
• Sodium hydride (95%) (168 mg, 7 mmol) was added to a solution of 1- methylthymine (700.5 mg, 5 mmol) in dimethylsulfoxide (15 mL).
• l-bromundec-10-ene (1.165 g, 5 mmol) was added and stirred for 12 hours at room temperature.
• the reaction mixture was then poured into a separatory funnel containing 100 mL of water and extracted with dichloromethane (5 X 75 mL).
• Example 37 This example illustrates a synthesis of l-(3,4-Oxidobutyl)-3,7-dimethy_xanthine
• Example 38 This example illustrates a synthesis of 1-(1 l,12-Oxidododecyl)-3,7 -dimethylxanthine
• This example illustrates a synthesis of l-(9,10-Oxidoctadecyl)-3,7-dimethylxanthine (CT2541).
• Triphenylphosphine (5.24 g; 20 mmol) was added in portions to a solution of oleyl alcohol (5.37 g; 20 mmol) and carbontetrabromide (6.63 g; 20 mmol) in 400 mL of dichloromethane and stirred for an hour at room temperature. The solvent was removed under reduced pressure and the residue extracted with hexane (3 X 200 mL). Further purification was done by flash chromatography over silica gel using hexane as eluant to yield 5.82 g (88% yield) of l-bromo-9-octadecene.
• the crude product obtained was further purified by flash chromatography over silica gel using a 30% acetone/petroleum ether eluant to yield 0.44 g (34 % yield) of )l-(9-octadecenyl)-3,7- dimethylxanthine (CT2539).
• This example illustrates a synthesis of l-(4-(S)-Methyl-7,8-oxido-8-methylnonyl)- 3,7 -dimethylxanthine (CT2548S).
• C2548S 3,7 -dimethylxanthine
• reaction mixture was extracted with dichloromethane (2 x 15 mL), and the combined organic phases were dried using magnesium sulfate and the solvent evaporated to yield a residue of l-(4-(S)-methyl-7- acetoxy-8-bromo-8-methylnonyl)-3,7-dimethylxanthine.
• This example illustrates a synthesis of l-(4-(R)-Methyl-7,8-oxido-8-methylnon)-3,7- dimethylxanthine (CT2548R).
• C2548R l-(4-(R)-Methyl-7,8-oxido-8-methylnon)-3,7- dimethylxanthine
• reaction mixture was extracted with dichloromethane (3 x 30 ml) using dried magnesium sulfate and the solvent evaporated to yield 0.51 g (97% yield) l-(4-(R)-Methyl-7,8- dihydroxy-8-methylnonyl)-3,7-dimethylxanthine (CT2537R) as a colorless oil.
• 1 -(4-(R)-Methyl-7 , -dihydroxy- 8-methylnonyl)-3 ,7-dimethylxanthine (0.29 g, 0.80 mmol) was stirred with hydrogen bromide (1.00 L of a 30% solution in acetic acid, 2.40 mmol) for 4 hours.
• This example illustrates a synthesis of l-(3,7-Dimethyl-2,3,6,7-dioxidoctyl)-3,7- dimethylxanthine (CT2552).
• Sodium hydride (95%) (0.28 g, 12 mmol) was added to a solution of theobromine (2.16 g, 12 mmol) in dimethylsulfoxide (50 mL).
• geranyl bromide (2.17 g, 10 mmol) was added.
• the reaction mixture was warmed to 60°C for 3 hours and then poured into a separatory funnel containing 150 mL of water and extracted with dichloromethane (5 X 75 mL).
• This example illustrates a synthesis of l-(12,13-Oxidotridecyl)-3,7-dimethylx__nthine (CT2562).
• CCT2562 l-(12,13-Oxidotridecyl)-3,7-dimethylx__nthine
• This example illustrates a synthesis of l-(7,8-cw-Oxidodecyl)-3,7-dimethylxanthine (CT2563).
• 7-c/5 ⁇ Decenal Johnson Matthey Catalog Co., lO.OOg, 6.5 mmol
• ethanol 100 mL
• Ammonium chloride solution (sat., 60 mL) was added with water (50 mL) and the mixture was extracted with dichloromethane (3 x 100 mL).
• This example illustrates a synthesis of l-(13,14-Oxidotetradecyl)-3,7- dimethylxanthine (CT3503).
• CTC3503 l-(13,14-Oxidotetradecyl)-3,7- dimethylxanthine
• reaction mixture was extracted with dichloromethane (3 x 100 mL) and the organic phase dried (magnesium sulfate) and evaporated to yield 2.10 g (96% yield) l-(13,14-Dihydroxytetradecyl)-3,7- dimethylxanthine as a white solid.
• l-(13,14-Dihydroxytetradecyl)-3,7-dimethylxanthine (0.80 g, 1.96 mmol) was stirred with HBr (1.94 mL of a 30% solution in acetic acid, 5.88 mmol) for 2 hours.
• This example illustrates a synthesis of l-(16,17-Oxidoheptadecyl)-3,7- dimethylxanthine (CT-3516).
• CTL l-(16,17-Oxidoheptadecyl)-3,7- dimethylxanthine
• This example illustrates a synthesis of N-(5,6-Oxidohexylamido)glutaric acid, methyl ester (CT1301).
• Sodium hydride (425 mg, 17.7 mmol) was added to a solution of glutarimide (2.00 g, 7.7 mmol) in dimethyl sulfoxide (40 mL).
• 6-bromo-l-hexene (2.90 g, 17.7 mmol) was added.
• the reaction was poured into a separatory funnel containing 100 mL water and extracted with dichlormethane (4 X 50 mL).
• nude mice This example illustrates the effects of CT1541 as hair growth stimulant in nude mice.
• nu/nu mice In a procedure similar to that used in a commercial model for predicting human hair-growing ability of minoxidil, nu/nu (nude) mice were painted twice daily for 16 days on the right flank with CT1541 using sterile applicators. Researches handled the price under a laminar flow hood with applicator, wearing face mask and sterile gloves.
• one mouse was sacrificed by cervical dislocation and skin biopsies taken from the treated areas of the shoulder/flank and the non-treated area of the dorsal pelvis (rump). Specimens were placed in 10% buffered formalin solution.
• a microscopic analysis of the skin biopsies confirmed that follicles in the treated areas had hair shafts which sometimes exit to the surface. There were mild accumulations of mixed inflammatory cells in the dermis. In contrast, hair follicles from untreated skin biopsies were smaller/shorter and less often extend into the subcutis. Hair shafts were rarely seen. A few mixed inflammatory cells were in the dermis in the untreated areas as well.
• the treated sections had more normal appearing hair follicles than the untreated sections. In addition, numerous hair shafts were seen exiting follicles in the treated sections.
• This example illustrates the effects of CT1105, CT1114, CT1413, CT1439, CT1594, CT2518, CT2548R, CT2548S, CT2562 and CT3503 as effective inhibitors of IL-2 induced proliferation of thymocytes.
• Single cell suspensions of thymus gland cells obtained from 4-6 week old mice were prepared. Two-hundred thousand cells were plated into individual wells of flat-bottom 96-well plates in RPMI-1640-10% FCS medium. The invention compounds were then added to the wells at varying concentrations and the cells incubated for 1-2 hours at 37°C.
• ConA Concanavalin A
• IL2 Interleukin-2
• This example illustrates the effects of CT1114, CT1413, CT1560, CT1565, CT1594, CT2518, CT2548R, CT2548S and CT3503 as effective inhibitors of normal human bone marrow stromal cells (MSC) proliferation in response to Platelet Derived Growth Factor BB (PDGF B) and IL-l ⁇ (50 and 10 ng/ml, respectively).
• MSC Platelet Derived Growth Factor BB
• IL-l ⁇ 50 and 10 ng/ml, respectively.
• MSC Mesenchymal stem cells
• inventive compounds were added to the cells at the appropriate concentration and then the cells again incubated for 1 hour.
• PDGF B -and IL-l ⁇ were added to the cells along with 3H-thymidine and the cells again incubated for an additional 24 hours. After 24 hours of incubation, the cells were then harvested to assess incorporation of
• Example 51 This example illustrates the effects of CT1605, CT1808 and CT1906 as immune modulators in a mixed lymphocyte reaction.
• Figure 1 shows the effects of three inventive compounds CT1605 (N-(5,6-oxidohexyl) glutarimide), CT1808 (N -(5,6oxidohexyl)-N 1 - methyluracil), and CT1906 (N3-(5,6-oxidohexyl) N* -methylthymine.
• the mixed lymphocyte reaction shows a proliferative response of PBMC (peripheral blood mononuclear cells) to allogeneic stimulation determined in a two-way mixed lymphocyte reaction.
• PBMC peripheral blood mononuclear cells
• This example illustrates a comparison of three dose levels of CT1808 and CT1906 and no drug control to inhibit thymocyte proliferation.
• the thymocytes were obtained from normal female Balb/C mice and stimulated with Concanavalin A (Con A) and/or IL-l ⁇ (interleukin-1 alpha).
• Con A Concanavalin A
• IL-l ⁇ interleukin-1 alpha
• the thymuses were dissociated and plated into 96-well plates at a density of 2 x 10 ⁇ cells/well. Dilutions of Con A and/or IL-l ⁇ were added to the wells and the cells were incubated for 4 days at 37°C. Drugs were added to the cell cultures two hours before activation with Con A and/or IL-l ⁇ .
• This example illustrates a comparison of CT1605 and CT1808 for inhibition of B-cell proliferation.
• a Ramos C-cell tumor line was treated with 250 ⁇ M CT1808 or CT1605 for one hour prior to stimulation of proliferation with anti-mu antibody or PMA (5 nM).
• PMA anti-mu antibody
• proliferation was measured with tritiated thymidine.
• both CT1605 and CT1808 inhibited proliferation in this model.
• This example illustrates a comparison of CT1605, CT1808 and CT1906 on PDGF- induced (platelet derived growth factor) proliferation of human stromal cells.
• Human stromal cells were starved in serum-free media for 24 hours and then stimulated with 50 ng/ml of PDGF-BB. The drugs were added at various indicated concentrations one hour prior to PDGF stimulation. Tritiated thymidine was added for 24 hrs at the time of PDGF stimulation to measure cellular proliferation. Background counts were approximately 5% of control levels. As shown in Figure 4, all three drugs inhibited PDGF-induced stimulation in a dose response fashion.
• HUVEC cells 4000/well, seeded 72 hrs in advance
• TNF human umbilical vein endothelial cells
• Drug was added to each culture (except for controls) one hour prior to adding TNF.
• Cell adhesion was determined by measuring fluorescence on a fluorescence plate reader. As illustrated in Figure 5, all three drugs showed a decrease in cell adhesion caused in a dose dependent fashion
• This example illustrates the effects of CT1605, CT1808 and CT1906 to inhibit cell surface expression of VCAM in human umbilical vein endothelial cells (HUVEC).
• the HUVEC cells were stimulated with 20 ng/ml TNF- ⁇ for 20 hrs and then stained for immunofluorescence using a monoclonal antibody recognizing VCAM, followed by a goat anti-mouse antibody conjugated to phycoerythrin. The cells were analyzed for antibody binding using flow cytometry.
• Figure 6 shows an analysis of mean relative fluorescence intensity of 10,000 cells, analyzed by flow cytometry. The mean fluorescence levels were decreased by all three drugs from control levels (TNF treatment, no drug).

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