AU722361B2 - Combinational therapeutic methods employing nitric oxide scavengers and compositions useful therefor - Google Patents

Description

i S F Ref: 415366D2

AUSTRALIA

PATENTS ACT 1990 COMPLETE SPECIICATION FOR A STANDARD PATENT

ORIGINAL

x Name and Address of Applicant: Actual Inventor(s): Address for Service: Invention Title: Medinox, Inc.

Suite E 11555 Sorrento Valley Road San Diego California 92121 UNITED STATES OF AMERICA Ching-San Lai Spruson Ferguson, Patent Attorneys Level 33 St Martins Tower, 31 Market Street Sydney, New South Wales, 2000, Australia Combinational Therapeutic Methods Employing Nitric Oxide Scavengers and Compositions Useful Therefor The following statement is a full description of this invention, including the best method of performing it known to me/us:- 5845 -1- Combinational Therapeutic Methods Employing Nitric Oxide Scavengers and Compositions Useful Therefor Field of the Invention The present invention relates to methods for directly or indirectly treating the production of species which induce the expression of nitric oxide synthase in mammals. In a particular preferred aspect, the present invention relates to methods for inactivating such species, or inhibiting the production of such species, while, at the same time, reducing nitric oxide levels, by co-administration of agents which inactivate (or inhibit the production of) such species, along with a scavenger of overproduced nitric oxide. In another preferred aspect, the present invention relates to reducing elevated nitric oxide levels associated with infectious and/or inflammatory conditions (and the treatment thereof), employing a combinational therapeutic method wherein an agent for the treatment of the infectious and/or inflammatory condition is coadministered along with a dithiocarbamate compound as a scavenger of overproduced nitric oxide. In a further preferred aspect, the present invention relates to compositions and formulations useful in the methods disclosed herein.

15 Background of the Invention In 1987, nitric oxide a gaseous free radical, was discovered in humans (see, for example, Ignarro et al., in Proc. Natl. Acad. Sci., USA 84:9265-69 (1987) and Palmer et al., in Nature 327:524-26 (1987)). As an indication of the significance of this discovery for the understanding of human physiology and pathophysiology, Science magazine selected nitric oxide as the molecule of the year in 1992.

Nitric oxide is formed from the terminal guanidino nitrogen atom of L-arginine by nitric oxide synthase (NOS; see, for example, Rodeberg et al., in Am. J. Surg. 170:292-303 (1995), and Bredt and Snyder in Ann. Rev. Biochem. 63:175-95 (1994)). Two major forms of nitric oxide synthase, constitutive and inducible enzymes, have been identified.

Under physiological conditions, a low output of -NO is produced by the constitutive, calciumdependent NOS isoform (cNOS) present in numerous cells, including endothelium and neurons. This low level of nitric oxide is involved in a variety of regulatory processes, eg., blood vessel homeostasis, neuronal communication and immune system function. On the other hand, under pathophysiological conditions, a high output of -NO is produced by the inducible, calcium-independent NOS isoform (iNOS) which is expressed in numerous cell types, including endothelial cells, smooth muscle cells and macrophages. These high levels of nitric oxide have been shown to be the aetiology of endotoxin shock. This high output of -NO further contributes to inflammation-related tissue damage, neuronal pathology, N-nitrosamine-induced carcinogenesis and mutations in human cells and bacteria via deamination reaction with DNA. Nitric oxide can therefore be seen to be a mixed blessing, being very desirable when present in small amounts, while potentially being highly detrimental when produced in excessive quantities.

Nitric oxide is a potent vasodilator (see, for example, Palmer in Arch. Surg. 128:396-401 (1993) and Radomski Moncada in Thromb. Haemos. 70:36-41 (1993). For example, in blood, -NO TR ;duced by the endothelium diffuses isotropically in all directions into adjacent tissues. As -NO Libc/03675 2 diffuses into the vascular smooth muscle, it binds to guanylate cyclase enzyme, which catalyses the production of cGMP, inducing vasodilation (see, for example, Ignarro, Ann. Rev. Toxicol. 30:535- 560 (1990); Moncada, Acta Physiol. Scand. 145:201-227 (1992); and Lowenstein and Snyder, Cell 70:705-707 (1992)). The overproduction of nitric oxide causes an extreme drop in blood pressure, resulting in insufficient tissue perfusion and organ failure, syndromes that are associated with many diseases and/or conditions septic shock, overexpression of cytokines, allograft rejection, and the like). The overproduction of nitric oxide is triggered by a number of stimuli, such as, the overproduction of inflammatory cytokines tumour necrosis factor (TNF), interleukin-1 (IL-1), interferons, endotoxin, and the like). Additionally, the overproduction of -NO has been discovered to be one of the major side-effects of cytokine therapy (see, for example, Miles et al., in Eur. J. Clin.

Invest. 24:287-290 (1994) and Hibbs et al., in J. Clin. Invest. 89:867-877 (1992)). Thus, abnormally elevated nitric oxide levels have been linked to many inflammatory and infectious diseases.

Inflammatory cytokines TNF, interleukins or interferons) and infectious agents (eg., endotoxin) induce nitric oxide overproduction by inducing transcription of the inducible nitric oxide synthase gene, leading to the production of inducible nitric oxide synthase, which in turn results in the overproduction of nitric oxide. The production of nitric oxide by the above described pathway can be disrupted in a variety of ways. Thus, for example, there have been attempts to develop monoclonal antibodies anti-endotoxin antibodies, anti-cytokine antibodies, anti-cytokine receptor antibodies, and the like) in efforts to block the -NO production pathway at the transcriptional level. Unfortunately, however, such efforts have met with very limited success (see, for example, Glauser et al., in Clin.

Infect. Dis. 18:S205-16 (1994) and St. John Dorinsky, in Chest 103:932-943 (1993)). At least one reason for the relative lack of success in the art is the fact that the production of inflammatory cytokines is short-lived (see, for example,. Wange Steinsham in Eur. J. Haematol. 50:243-249 (1993)), while overproduction of nitric oxide lasts several days, causing systemic hypotension, 25 insufficient tissue perfusion and organ failure.

Thus, for example, during endotoxemia, TNF production peaks at about 1-2 hours. Therefore, in order to be effective, anti-TNF antibodies would have to be administered at an early stage after infection. Indeed, in many clinical settings, patients are likely to already have been infected with bacteria prior to being admitted. Accordingly, such therapeutic methods have met with only limited 30 success.

S Currently, many pharmaceutical companies have turned their attention to the design and development of substrate or product analogue inhibitors of the enzyme, NOS, in efforts to treat the overproduction of -NO. However, recent data show that the inhibition of NOS is detrimental to subjects. For example, rodent studies show that inhibition of the production of nitric oxide causes intrauterine growth retardation and hind-limb disruptions in rats (see, for example, Diket et al., in Am.

J. Obstet. Gynecol. 171:1243-1250 (1994)). Furthermore, the inhibition of nitric oxide synthesis during endotoxemia has also been shown to be detrimental (see, for example, Minnard et al., in Arch. Surg.

129:142-148 (1994); Luss et al., in Biochem. Biophys. Res. Commun. 204:635-640 (1994);and Hargrecht et al., in J. Leuk. Biol. 52:390-394 (1992)). Similar results have been reported in larger animal studies, such as dogs and swine (see, for example, Statman et al., in J. Surg. Res. 57:93-98 Libc03675 (1994); Mitaka et al., Am. J. Physiol. 268 ::H2017-H2023 (1994); Robertson, et al., Arch. Surg.

129:149-156 (1994); and Henderson et al., Arch. Surg. 129:1271-1275 (1994)).

Since a variety of stimuli induce expression of nitric oxide synthase, which, in turn, leads to nitric oxide overproduction (with its attendant detrimental effects), there is a need in the art to effectively treat both the initial stimulus of nitric oxide synthase expression, and the resulting overproduction of nitric oxide, as well as overproduction of nitric oxide which may be induced (directly or indirectly) by therapeutic agents employed for the treatment of a wide variety of infectious and/or inflammatory conditions.

Brief Description of the Invention According to a first aspect, the present invention consists in a method for directly or indirectly treating the production of species which induce the expression of inducible nitric oxide synthase in a subject, said method comprising: co-administering to said subject an effective amount of a combination of at least one agent capable of directly or indirectly inactivating said species, or inhibiting production of said species, and at least one nitric oxide scavenger.

15 According to a second aspect, the present invention consists in a combination of at least one agent capable of directly or indirectly inactivating species which induce the expression of nitric oxide synthase in a subject, or inhibiting production of said species, and at least one nitric oxide scavenger when used in directly or indirectly treating the production of species which induce the expression of inducible nitric oxide synthase.

20 According to a third aspect, the present invention consists in the use of a combination of at least one agent capable of directly or indirectly inactivating species which induce the expression of nitric oxide synthase in a subject, or inhibiting production of said species, and at least one nitric oxide scavenger for the manufacture of a medicament for directly or indirectly treating the production of species which induce the expression of inducible nitric oxide synthase.

According to a fourth aspect, the present invention consists in a therapeutic method which employs an agent to inactivate materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase, comprising co-administering to a patient in need thereof a nitric oxide scavenger in combination with said agent.

According to a fifth aspect, the present invention consists in a combination of an agent to inactivate materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger when used in therapy.

According to the sixth aspect, the present invention consists in the use of a combination of an agent to inactivate materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger for the manufacture of a medicament for therapy.

According to the seventh aspect, the present invention consists in a therapeutic method which employs a therapeutic agent which, directly or indirectly, induces the expression of inducible nitric oxide synthase, comprising co-administering to a patient in need thereof a nitric oxide scavenger in combination with said therapeutic agent.

Libc/03675 3a According to an eighth aspect, the present invention consists in a combination of a therapeutic agent which, directly or indirectly, induces the expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger when used in therapy.

According to a ninth aspect, the present invention consists in the use of a combination of a therapeutic agent which, directly or indirectly, induces the expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger for the manufacture of a medicament for therapy.

According to a tenth aspect, the present invention consists in a composition comprising a combination of an agent capable of inactivating materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase and a nitric oxide scavenger in a pharmaceutically acceptable carrier therefor.

In accordance with one aspect of the present invention, combinational therapeutic methods have been developed for the in vivo inactivation or inhibition of formation (either directly or indirectly) of species which induce the expression of inducible nitric oxide synthase, as well as reducing nitric oxide levels produced as a result of -NO synthase expression. In another aspect, combinational therapeutic methods have been developed which can be employed, for example, for the treatment of infectious and/or inflammatory conditions. Thus, the effectiveness of many therapeutic agents used for the treatment of infectious and/or inflammatory conditions can be enhanced by co-administration thereof in combination with dithiocarbamate-containing nitric oxide scavenger(s).

In contrast to the inhibitory approach described in the prior art to address the problem of nitric 20 oxide overproduction (see, for example, Aisaka et al., Biochem. Biophys. Res. Commun. 60:881-886 (1989); Rees, et al., Proc. Natl. Acad. Sci. USA 86:3375-3379, (1989)); Henderson et al., in Arch.

Surg. 129:1271-1275 (1994); Hambrecht et al., in J. Leuk. Biol. 52:390-394 (1992); Luss et al., in Biochem. and Biophys. Res. Comm. 204:635-640 (1994); Robertson et al., in Arch. Surg. 129:149- :25 156 (1994); Statman et al., in J. Surg. Res. 57:93-98 (1994); and Minnard et al., in Arch. Surg.

°25 129:142-148 (1994)), in one aspect, the present invention employs a combination of inactivation (and/or inhibition) and scavenging approach whereby the stimulus of nitric oxide synthase expression is inactivated and/or expression thereof is inhibited, and overproduced nitric oxide is bound in vivo to a suitable nitric oxide scavenger. The resulting complexes render the stimulus of nitric oxide synthase expression inactive (or inhibit the production thereof), while also rendering the resulting nitric oxide harmless. The resulting complexes are eventually excreted in the urine of the host.

In another aspect, a suitable nitric oxide scavenger is co-administered along with a therapeutic agent which may promote nitric oxide formation, thereby providing a protective affect against the otherwise detrimental effects of nitric oxide overproduction.

Further in accordance with the present invention, there have been developed compositions and formulations useful for carrying out the above-described methods.

Numerous stimuli for -NO synthase are known in the art. Co-administration of agents which inactivate the stimulus of -NO synthase expression (or inhibit the production thereof), in combination with nitric oxide scavengers as described herein, provides a more effective means to treat a variety of indications than has previously been described in the art.

Libc/03675 An exemplary nitric oxide scavenger contemplated for use in the practice of the present invention is a dithiocarbamate-ferrous iron complex. This complex binds to *NO, forming a stable, water-soluble dithiocarbamate iron-NO complex having a characteristic three-line spectrum (indicative of a mononitrosyl-Fe complex) which can readily be detected at ambient temperatures by electron paramagnetic resonance (EPR) spectroscopy (See Komarov et al., in Biochem. Biophys. Res.

Commun. 195:1191-1198 (1993); and Lai and Komarov, FEBS Lett.,345:120-124, (1994)). This method of detecting 'NO in body fluids in real time has recently been described by Lai in US. 5 358 703, incorporated by reference herein in its entirety.

SThe present invention relates to combinational therapeutic methods for treating the production of species which induce the expression of nitric oxide synthase in mammals. Thus, a dual attack is mounted against a variety of stimuli which lead to the production of dangerously high in vivo levels of *NO. Combinations of agents contemplated for use in the practice of the present invention agents capable of inactivating species which induce expression of inducible nitric oxide, or agents which inhibit the production of such species, or therapeutically useful agents which also induce nitric oxide production, and nitric oxide scavengers) are administered to a host in need of such treatment. The agent capable of inactivating (or inhibiting the production of) species which induce expression of inducible nitric oxide and .NO scavengers interact with the stimulus or stimuli of nitric oxide synthase expression and in vivo produced -NO, respectively, forming a complex between said species and said agent, as well as a stable scavenger-NO complex a dithiocarbamate-metal-NO complex).

Whereas free -NO is a potent vasodilator, chelated -NO complexes *NO chelated with dithiocarbamate-iron complexes) are not. The NO-containing complex is then filtered through the kidneys, concentrated in the urine, and eventually excreted by the subject, thereby reducing in vivo *NO levels.

In another aspect, the present invention relates to reducing elevated nitric oxide levels 25 associated with infectious and/or inflammatory conditions (and the treatment thereof). In accordance with this aspect of the invention, a combinational therapeutic method is employed wherein an agent for the treatment of an infectious and/or inflammatory condition is co-administered along with a dithiocarbamate compound as a scavenger of overproduced nitric oxide.

Brief Description of the Figure 30 Figure 1 illustrates the effects of endotoxin (LPS-4 mg/kg) treatment on mean arterial pressure (MAP) with and without [(MGD)2/Fe] treatment. Bolus i.v. injection of LPS at time zero was as indicated in the Figure. Data marked by open circles are the result of bolus i.v. injection of saline, followed by 1.0mL/h of continuous saline infusion (n=11/16, note: 11 out of 16 animals died w before the end of the experiments). Data marked by closed circles are the result of [(MGD)2/Fe] infusion, O.lmmole/kg loading dose followed by 0.lmmole/kg/hr i.v. infusion (n=3/16, note: only 3 out of 16 animals died before the end of the experiments). Data points marked with an asterisk indicate the results are significantly different at p <0.05. The ratio of MGD to Fe used was 5:1 (MGD:Fe), and the dosage shown was with respect to MGD.

Libc/03675 Detailed Description of the Invention In accordance with the present invention, there are provided combinational therapeutic methods for treating a variety of conditions related to the overproduction of nitric oxide by a subject. In one aspect, the invention method comprises directly or indirectly treating the production of species which induce the expression of.inducible nitric oxide synthase in a subject. Invention methods comprise: co-administering to a subject an effective amount of a combination of at least one agent capable of directly or indirectly inactivating said species, or inhibiting production of said species, and at least one nitric oxide scavenger.

In accordance with another aspect of the present invention, combinational therapeutic methods have been developed employing an effective amount of a combination of at least one treating agent useful for the treatment of infectious and/or inflammatory conditions, and at least one dithiocarbamatecontaining nitric oxide scavenger. It has been found that the above-described combination is more effective for the treatment of infectious and/or inflammatory conditions than is the treating agent alone.

As readily recognised by those of skill in the art, a variety of agents can be used to scavenge nitric oxide. Examples of suitable agents for this purpose include non-heme iron-containing peptides or proteins, porphyrins, metalloporphyrins, dithiocarbamates, dimercaptosuccinic acid, phenanthroline, desferrioxamine, pyridoxal isonicotinoyl hydrazone (PIH), 1,2-dimethyl-3-hydroxypyrid-4-one 1,2-bis(3,5-dioxopiperazine-1-yl)propane (ICRF-187), and the like. A presently preferred class of compounds useful for such purpose is the dithiocarbamates. Dithiocarbamate-containing nitric oxide scavengers contemplated for use in the practice of the present invention include any physiologically compatible derivative of the dithiocarbamate moiety

(R)

2 Such compounds can be described with reference to the following generic structure [R1R2N-C(S)-S-]xM+ 1 2 3

(I)

25 wherein: each Ri and R2 is independently selected from a Ci up to C18 alkyl, substituted alkyl, :cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, alkylaryl, .substituted alkylaryl, arylalkyl, substituted arylalkyl, arylalkenyl, substituted arylalkenyl, arylalkynyl, substituted arylalkynyl, aroyl, substituted aroyl, acyl, substituted acyl or R 1 and R 2 can cooperate to 30 form a 6- or 7-membered ring including N, R 1 and R 2 x is 1 or 2, and M is a monovalent cation when x is 1, or M is a physiologically compatible divalent or trivalent transition metal cation when x is 2.

Presently preferred compounds having the above described generic structure are those wherein: each of R 1 and R 2 a C1 up to C12 alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl or substituted alkynyl, wherein the substituents are selected from carboxyl, oxyacyl, phenol, phenoxy, pyridinyl, pyrrolidinyl, amino, amido, hydroxy, nitro or sulfuryl, and M Fe 2 or Fe+ 3 Especially preferred compounds having the above described generic structure are those wherein: Libcd03675

R

1 a C2 up to C8 alkyl or substituted alkyl, wherein the substituents are selected from carboxyl, acetyl, pyridinyl, pyrrolidinyl, amino, amido, hydroxy or nitro, R 2 is selected from a Ci up to C6 alkyl or substituted alkyl, or R 2 can cooperate with R 1 to form a 6- or 7-membered ring including N, R 2 and

R

1 and M Fe+ 2 The presently most preferred compounds having the above-described generic structure are those wherein: R1 a C2 up to C8 alkyl or substituted alkyl, wherein the substituents are selected from carboxyl, acetyl, amido or hydroxy, R 2 a C1 up to C4 alkyl or substituted alkyl, and M Fe' 2 When R 1 and R 2 cooperate to form a 6- or 7 membered ring, the combination of R 1 and R 2 can be a variety of saturated or unsaturated 4, 5 or 6 atom bridging species selected from alkenylene or and/or -N(R)containing alkylene moieties, wherein R is hydrogen or a lower alkyl moiety.

Monovalent cations contemplated for incorporation into compounds of structure include H+, Na NH4+, tetraalkyl ammonium, and the like. Physiologically compatible divalent or trivalent transition metal cations contemplated for incorporation into the above compounds include charged forms of iron, cobalt, copper, manganese, or the like Fe+2, Fe+ 3 Co+2, Co+ 3 Cu+ 2 Mn+ 2 or Mn+ 3 In accordance with the present invention, the ratio of dithiocarbamate-species to counter-ion M can vary widely. Thus, dithiocarbamate-containing nitric oxide scavenger can be administered without any added metallic counter-ion M or a transition metal cation to dithiocarbamate-species ratio of zero), with ratios of transition metal cation to dithiocarbamate-species up to about 1:2 a 2:1 dithiocarbamate:transition metal cation complex) being suitable.

As employed herein, "substituted alkyl" comprises alkyl groups further bearing one or more substituents selected from hydroxy, alkoxy (of a lower alkyl group; wherein a lower alkyl group has about 1-4 carbon atoms), mercapto (of a lower alkyl group), cycloalkyl, substituted cycloalkyl, 25 heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, aryloxy, substituted aryloxy, halogen, trifluoromethyl, cyano, nitro, nitrone, amino, amido, acyl, oxyacyl, carboxyl, carbamate, sulfonyl, sulfonamide, sulfuryl, and the like.

As employed herein, "cycloalkyl" refers to cyclic ring-containing groups containing in the range of about 3 up to 8 carbon atoms, and "substituted cycloalkyl" refers to cycloalkyl groups further 30o bearing one or more substituents as set forth above.

As employed herein, "alkenyl" refers to straight or branched chain hydrocarbyl groups having at least one carbon-carbon double bond, and having in the range of about 2 up to 12 carbon atoms, and "substituted alkenyl" refers to alkenyl groups further bearing one or more substituents as set forth above.

35 As employed herein, "alkynyl" refers to straight or branched chain hydrocarbyl groups having at least one carbon-carbon triple bond, and having in the range of about 2 up to 12 carbon atoms, and "substituted alkynyl" refers to alkynyl groups further bearing one or more substituents as set forth above.

Libc/03675 As employed herein, "aryl" refers to aromatic groups having in the range of 6 up to 14 carbon atoms and "substituted aryl" refers to aryl groups further bearing one or more substituents as set forth above.

As employed herein, "alkylaryl" refers to alkyl substituted aryl groups and "substituted alkylaryl" refers to alkylaryl groups further bearing one or more substituents as set forth above.

As employed herein, "arylalkyl" refers to aryl substituted alkyl groups and "substituted arylalkyl" refers to arylalkyl groups further bearing one or more substituents as set forth above.

As employed herein, "arylalkenyl" refers to arylsubstituted alkenyl groups and "substituted arylalkenyl" refers to arylalkenyl groups further bearing one or more substituents as set forth above.

As employed herein, "arylalkynyl" refers to arylsubstituted alkynyl groups and "substituted arylalkynyl" refers to arylalkynyl groups further bearing one or more substituents as set forth above.

As employed herein, "aroyl" refers to arylcarbonyl species such as benzoyl and "substituted aroyl" refers to aroyl groups further bearing one or more substituents as set forth above.

As employed herein, "heterocyclic" refers to cyclic ring-containing) groups containing one or more heteroatoms N, O, S, or the like) as part of the ring structure, and having in the range of 3 up to 14 carbon atoms and "substituted heterocyclic" refers to heterocyclic groups further bearing one or more substituents as set forth above.

As employed herein, "acyl" refers to alkylcarbonyl species.

As employed herein, "halogen" refers to fluoride, chloride, bromide or iodide atoms.

Induction of expression of inducible nitric oxide synthase, and hence, overproduction of nitric oxide, is associated with a wide range of disease states and/or indications, such as, for example, septic shock, haemorrhagic shock, anaphylactic shock, toxic shock syndrome, ischaemia, cerebral ischaemia, administration of cytokines, overexpression of cytokines, ulcers, inflammatory bowel disease ulcerative colitis or Crohn's disease), diabetes, arthritis, asthma, Alzheimer's disease, 25 Parkinson's disease, multiple sclerosis, cirrhosis, allograft rejection, encephalomyelitis, meningitis, pancreatitis, peritonitis, vasculitis, lymphocytic choriomeningitis, glomerulonephritis, uveitis, ileitis, inflammation liver inflammation, renal inflammation, and the like), burn, infection (including bacterial, viral, fungal and parasitic infections), haemodialysis, chronic fatigue syndrome, stroke, cancers breast, melanoma, carcinoma, and the like), cardiopulmonary bypass, ischaemic/reperfusion injury, gastritis, adult respiratory distress syndrome, cachexia, myocarditis, autoimmune disorders, eczema, psoriasis, heart failure, heart disease, atherosclerosis, dermatitis, S..urticaria, systemic lupus erythematosis, AIDS, AIDS dementia, chronic neurodegenerative disease, chronic pain, priapism, cystic fibrosis, amyotrophic lateral sclerosis, schizophrenia, depression, premenstrual syndrome, anxiety, addiction, migraine, Huntington's disease, epilepsy, 35 neurodegenerative disorders, gastrointestinal motility disorders, obesity, hyperphagia, solid tumours neuroblastoma), malaria, haematologic cancers, myelofibrosis, lung injury, graft-versus-host disease, head injury, CNS trauma, hepatitis, renal failure, liver disease chronic hepatitis druginduced lung injury paraquat), myasthenia gravis ophthalmic diseases, and the like.

Treatment of such conditions can be carried out with a variety of reagents, such as, for example, anti cytokine antibodies, anti-cytokine receptor antibodies, anti-endotoxin antibodies, Libc/03675 bradykinin antagonists, synthetic peptide blocking bradykinin receptors, bactericidal/permeability increasing protein, antibodies to platelet activating factor, inhibitors of arachidonate metabolism, inhibitors of nitric oxide synthase enzymes, immunosuppressors, diabetic therapeutic agents, antiinflammatories, agents useful for stroke therapy, agents useful for asthma therapy, agents useful for cirrhosis therapy, anti-cancer therapeutics, anti-microbial therapeutics, anti-fungal therapeutics, antiretroviral therapeutics, agents useful for the treatment of opportunistic infections and malignancies, agents useful for the treatment of Lupus erythmatosus, agents useful for the treatment of uveitis, thrombolytic agents, antispasmodic agents, antidiarrheal agents, agents useful for the treatment of constipation, antihistamines, agents useful for the treatment of Parkinson's disease, and the like.

Such agents, employed either alone or as part of a combination of any two or more thereof, can advantageously be combined with nitric oxide scavengers as described herein, and can be used for a variety of indications, such as for example, anti-endotoxin therapy antibodies to endotoxin, antibodies to LPS-binding protein, soluble CD14 protein, bactericidal/permeability increasing protein, polymyxin B, and the like), inhibition of cytokine synthesis/release employing phosphodiesterase inhibitors, IL-4, IL- IL-13, TGF-p, corticosteroids, and the like), anti-cytokine therapy employing antibodies to TNF, soluble TNF receptors, IL-1 receptor antagonists, antibodies to IL-1 receptors, antibodies to IL-6, antibodies to interferon-y, soluble interferon-y receptors, and the like), inhibition of the coagulation cascade (and of complement activation, employing such agents as anti-Factor XII antibodies, antibodies to C5a, CI-esterase inhibitors, soluble Crl, and the like), inhibition of platelet activating factor (PAF, employing such agents as PAF receptor antagonists, and the like), inhibition of arachidonate metabolism employing agents such as cyclooxygenase inhibitors, lipoxygenase inhibitors, leukotriene inhibitors, thromboxane A2 inhibitors, prostaglandins, and the like), inhibition of nitric oxide synthase enzymes employing arginine analogues (such as L-NGmethylarginine, L-NG-nitroarginine, L-NG-aminoarginine, L-iminoethylornithine, e-N-iminoethyl-Llysine, L-NG-nitroarginine methyl ester, L-NG- hydroxyl-NG-methylarginine, L-NG-methyl

NG-

methylarginine, L-thiocitrulline, L-S-methylthiocitrulline, L-S-ethylisothiocitrulline,

S-

30 ethylisothiocitrulline, aminoguanidine, S-methyl isothiourea sulfate, and the like), heme ligands (such as 7-nitroindazole, 7,7,8,8-tetramethyl-o-quinodimethane, imidazole, 1-phenylimidazole, 2phenylimidazole, and the like), calmodulin antagonists such as chlorpromazine, W-7, and the like), and the like); immunosuppression employing one or more agents such as cyclosporin A, OKT3, FK506, 35 mycophenolate mofetil (MMF), azathioprine, corticosteroids (such as prednisone), antilymphocyte globulin, antithymocyte globulin, and the like), diabetic therapy employing one or more agents such as free pancreatic islets, encapsulated pancreatic islets, oral insulin, intravenous insulin, amylin hormone, and the like), dihydropyridine calcium channel blockers employing agents such as nifedipine, nitrendipine, Libc/03675

'-S

9 nisoldipine, and the like), acetohexamide, chlorpropamide, glyburide, glipizide, metformin, tolbutamide, tolazamide, and the like, inflammatory disease therapy employing disease-modifying agents such as antimalarials, methotrexate, sulfasalazine, mesalamine, azathioprine, 6-mercaptopurine, metronidazole, injectable and oral gold, D-penicillamine, and the like), corticosteroids, non-steroidal antiinflammatory drugs such as acetominophen, aspirin, sodium salicylate, magnesium salicylate, choline magnesium salicylate, salicylsalicylic acid, ibuprofen, naproxen, diclofenac, diflunisal, etodolac, fenoprofen calcium, fluriprofen, piroxicam, indomethacin, ketoprofen, ketorolac tromethamine, meclof enamate, meclofenamate sodium, mefenamic acid, nabumetone, oxaprozin, phenyl butyl nitrone (PBN), sulindac, tolmetin, and the like), stroke therapy employing one or more agents such as fibrinolytic agents (such as streptokinase, acylated plasminogen streptokinase complex, urokinase, tissue plasminogen activator, and the like), employing monoclonal antibodies directed against leucocyte adhesion molecules (such as intercellular adhesion molecule-i (ICAM-1), CD18, and the like), haemodilution therapy (employing modified haemoglobin solutions such as diaspirin crosslinked haemoglobin), employing growth factors (such as basic fibroblast growth factor (bFGF), transforming growth factor-beta 1 (TGF-P1), and the like), employing glutamate antagonists (such as lamotrigine, dizolcilpine maleate (MK 801), BW619C89, BW1003C87, and the like), employing NMDA antagonists (such as CGS 19755 (Selfotel), aptiganel hydrochloride, dextrorphar, d-CPPene, and the like), employing GABA agonists (such as muscimol), employing free radical scavengers (such as allopurinol, S-PBN, 21-aminosteroids, tocopherol, superoxide dismutase, dexanabinol (HU-211), selenium, carotenoids, and the like), idebenone, ticlopidine, lovastatin, citicoline, and the like), asthma therapy employing bronchodilators (such as albuterol, salmeterol, metaproternol, bitolterol, pirbuterol, terbutaline, isoproterenol, epinephrine, and the like), theophyllines (such as theophylline, aminophylline, and the like), corticosteroids (such as beclomethasone, prednisone, and the like), antimediators (such as cromolyn sodium, nedocromil sodium, and the like), and the like), :cirrhosis therapy employing diuretics (such as spironolactone), opiate antagonists (such as naloxone), cholestyramine, colchicine, colestipol, methotrexate, rifampin, ursodeoxycholic acid, and the like, anti-cancer therapy employing one or more agents such as alkylating agents (such as mechlorethamine, chlorambuccil, ifosfamide, melphalan, busulfan, carmustine, lomustine, Sprocarbazine, dacarbazine, cisplatin, carboplatin, and the like), antimetabolites (such as methotrexate, mercaptopurine, thioguanine fluorouracil, cytarabine, and the like), hormonal agents (such as testosterone propionate, fluoxymesterone, flutamide, diethylstilbestrol, ethinyl estradiol, tamoxifen, hydroxyprogesterone caproate, medroxyprogesterone, megestrol acetate and the like), adrenocorticosteroids (such as prednisone), aromatase inhibitors (such as amino glutethimide), leuprolide, goserelin acetate, biological response modifiers (such as interferon-x2a, interferon-a2b, interleukin-2, and the like), peptide hormone inhibitors (such as octreotide acetate), natural products (such as vinblastine, vincristine, vinorelbine, paclitaxel, dactinomycin, daunorubicin, idarubicin, doxorubicin, etoposide, plicamycin, mitomycin, mitoxantrone, bleomycin, hydroxyurea, mitotane, Libc/03675 fludarabine, cladribine, and the like), supportive agents (such as allopurinol, mesna, leucovorin, erythropoietin, filgrastim, sargramostim, and the like), and the like, anti-microbial therapy employing one or more agents such as celftriaxone, TMP-SMZ, penicillin, aminoglycosides, vancomycin, gentamicin, rifampin, imipenem, clindamycin, metronidazole, tetracycline, erythromycin, sulfonamide, streptomycin, ampicillin, isoniazid, pyrazinamide, ethambutol, and the like), anti-fungal therapy employing agents such as amphotericin B, griseofulvin, myastatin, flucytosine, natamycin, antifungal imidazoles clotrimazole, miconazole, ketoconazole, fluconazole, itraconazole, and the like), and the like, anti-retroviral therapy employing agents such as protease inhibitors (such as Invirase, Ritonavir, Crixivan, and the like), zidovudine, didanosine, zalcitabine, stavudine, viramune, and the like) treatment of opportunistic infections and malignancies anti-AIDS treatment, employing agents such as pentamidine, trimethoprim/sulfamethoxazole, primaquine, atovaquone, clarithromycin, clofazimine, ethambutol, rifampin, amikacin, ciprofloxacin, pyrimethamine, amphotericin B, ganciclovir, foscarnet, fluconazole, ketoconazole, acyclovir, and the like), Lupus erythymatosus therapy employing agents such as hydroxychloroquine sulfate, chloroquine sulfate, quinacrine, dapsone, isotretinoin, and the like), uveitis therapy employing agents such as corticosteroids, azathioprine, cyclosporine, and the like), thrombolytic therapy for acute myocardial infarction employing agents such as streptokinase, tissue plasminogen activator anistreplase, and the like), antispasmodic treatment employing agents such as dicyclomine, hyoscyamine, propantheline, and the like), S 25 antidiarrheal treatment employing agents such as loperamide, diphenoxylate with atropine, and the like), anticonstipation treatment employing agents such as fibre supplementation with bran, psyllium, methylcellulose, polycarbophil, cisapride, and the like), antihistamine therapy employing agents such as ethanolamines (such as 3 diphenhydramine, clemastine, and the like), ethylenediamines (such as brompheniramine, chlorpheniramine, triprolidine, and the like), phenothiazines (such as hydroxyzine), piperidines (such as terfenadine, astemizole, azatadine, cyproheptadiene, loratidine, and the like), and the like), anti-Parkinsonian therapy employing agents such as benztropine mesylate, biperiden, chlorphenoxamine, cycrimine, orphenadrine, procyclidine, trihexyphenidyl, and the like), 35 as well as other indications which involve the induction of nitric oxide synthase, as can readily be identified by those of skill in the art.

In addition, administration of many therapeutic agents can also lead to the induction of expression of inducible nitric oxide synthase, and hence, overproduction of nitric oxide. For example, nitric oxide overproduction is also associated with the following treatments, such as, for example, administration of immunosuppressants, such as glucocorticoids (methylprednisolone), myelin basic Libc/03675 11 protein 7-capaxone), anti-Fc receptor monoclonal antibodies, hydroorotate dehydrogenase inhibitor, anti-lL2 monoclonal antibodies CHI-621 and dacliximab), buspirone, castanospermine, CD-59 (complement factor inhibitor), 5-lipoxygenase inhibitor CMI-392), phosphatidic acid synthesis antagonists, ebselen, edelfosine, enlimomab, galaptin, platelet activating factor antagonists, selectin antagonists ICAM-4), interleukin-10 agonist, macrocylic lactone, methoxatone, mizoribine, OX-19, peptigen agents, PG-27, protein kinase C inhibitors, phosphodiesterase

IV

inhibitor, single chain antigen binding proteins, complement factor inhibitor, sialophorin, sirolimus, spirocyclic lactams, 5-hydroxytryptamine antagonist, anti-TCR monoclonal antibodies, CD5 gelonin, TOK-8801, and the like.

Additional treatments which lead to the overexpression of nitric oxide include administration of antimetabolite cytotoxics azathioprine, cyclophosphamide), C5a release inhibitor, benzydamine, peldesine, pentostatin, SDZ-ASM-981, thalidomide, benzoporphyrin derivatives, arachidonate antagonists halometasone, halobetasol propionate), corticosteriod (clobetasol propionate), growth hormone antagonists (octapeptide somatostatin analogue, lanreotide, angiopeptin and dermopeptin), thymopentin, and the like.

Other treatments which lead to the overexpression of nitric oxide include administration of neuroprotective agents, such as a-adrenoreceptor antagonist a-dihydroergocryptine),

NMDA

antagonists 5,6,7-tichloro-THQTQ, remacemide, 2-piperazinecarboxylic acid, Nindologlycinamide derivatives, spiro[benzo(b)thiophen-4(5H)] derivatives, CP-101606, eliprodil, dexanabinol, GV-150526, L-695902, L-701324, amantadine derivatives, dizocilpine, benzomorphan derivatives, aptiganel, (S)-a-phenyl-2-pyridine ethanamide dihyrochloride, I-aminocyclopentanecarboxylic acid, and the like), sodium channel antagonists 619C89), glycine antagonists glystasins), calcium channel antagonists 3,5-pyridinedicarboxylic acid derivatives, conopeptides, 1-piperazineethanol, thieno[2,3b]pyridine-5-carboxylic acid derivatives, NS- 3034, nilvadipine, nisoldipine, tirilazad mesylate, 2H-1-enzopyran-6-ol, nitrone spin traps, lacidipine, iomeerzine hydrochloride, lemildipine, lifarizine, CPC-304, efonidipine, F-0401, piperazine derivatives, and the like), calpain inhibitors, fibrinogen antagonists ancrod), integrin antagonists (eg., antegren), thromboxane A2 antagonist 9H-carbazole-9-propanoic acid derivatives, acid derivatives, 1-azulene sulfonic acid derivatives, and the like), brain-derived neurotropic factor, S 30 adrenergic transmitter uptake inhibitor 1-butanamine), endothelin A receptor antagonists (eg., benzenesulfonamide derivatives), GABA A receptor antagonists triazolopyrimidine derivatives, cyclohexaneacetic acid derivatives, and the like), GPIlb Ila receptor antagonists C68-22), Splatelet aggregation antagonist 2(1H)-quinolinone derivatives, 1H-pyrrole-1 -acetic acid derivatives, coumadin, and the like), Factor Xa inhibitor, CPC-211, corticotropin releasing factor agonist, thrombin inhibitor cothrombins, fraxiparine, dermatan sulfate, heparinoid, and the like), dotarizine, intracellular calcium chelators BAPTA derivatives), radical formation antagonists (eg., EPC-K1, 3-pyridinecarboxamide derivatives, superoxide dismutase, raxofelast, lubeluzole, 3Hpyrazol-3-one derivatives, kynurenic acid derivatives, homopiperazine derivatives, polynitroxyl albumin, and the like), protein kinase inhibitors 1H-1,4-diazepine), nerve growth agonist (eg., floor plate factor-5), glutamate antagonist cyclohexanepropanoic acid, riluzole, NS-409, Libc/03675 acetamide derivatives, and the like), lipid peroxidase inhibitors 2,5-cyclohexadiene-1,4dione derivatives), sigma receptor agonist cyclopropanemethanamine derivatives, SA-4503, and the like), thyrotropin releasing hormone agonist JTP-2942, L-prolinamide, posatirelin, and the like), prolyl endopeptidase inhibitor, monosialoganglioside GM1, proteolytic enzyme inhibitor (eg., nafamostat), neutrophil inhibitory factor, platelet activating factor antagonist nupafant), monoamine oxidase B inhibitor parafluoroselegiline, benzonitrile derivatives, and the like), PARS inhibitors, Angiotensin I converting enzyme inhibitor perindopril, ramipril, and the like), acetylcholine agonist pramiracetam), protein synthesis antagonist procysteine), phosphodiesterase inhibitor propentofylline), opioid kappa receptor agonist phenothiazine-2-carboxamine derivatives), complement factor inhibitor sCRI fragments), somatomedin-1, carnitine acetyltransferase stimulant acetylcarnitine), and the like.

Still further treatments which lead to the overproduction of nitric oxide include administration of T cell inhibitors, such as synthetic leucocyte antigen derived peptides, interleukin-1 receptor antagonist, MG/AnergiX, anti-CD3 monoclonal antibodies, anti-CD23 monoclonal antibodies, anti- CD28 antibodies, anti-CD2 monoclonal antibodies, CD4 antagonists, anti-E selectin antibodies, MHC inhibitors, monogens, mycophenolate mofetil, and the like.

Additional treatments which lead to overproduction of nitric oxide include administration of antimigraine agents, such as MK-462, 324C91, Phytomedicine, (S)-fluoxetine, calcium channel antagonists nimodipine/Nimotop, flunarizine, dotarizine/FI-6026, iomerizine HCLKB-2796, CPC- 304, CPC-317, and the like), a-dihydroergocryptine, 5-HT1 agonists, Sumatriptan/Imitrex, Imigran, GR-85548, 311C, GR-127607, and the like), 5-HT1:D agonists, 5-HT1A antagonists, 5-HT1B antagonists CP-93129), 5-HTiD antagonists 1H-indole-5-ethanesulfonamide derivatives, and the like), 5-HT1D receptor cloned 5-HT1D agents), 2thiophenecarboxamide, 3-piperidinamine, diclofenac potassium, dihydroergotamine DHE 450), dolasetron mesilate, dotarizine, flupirtine, histamine-H3 receptor agonist, indobufen, 1-azulenesulfonic acid derivatives, cholinesterase inhibitors, S-9977), bradykinin antagonists, nitric oxide reductase inhibitors BN-52296), nitric oxide receptor antagonists, substance P antagonists (eg., Capsaicin/Nasocap), endopeptidase inhibitors neutral endopeptidase, cloned), piperazine .derivatives, neurokinin 1 antagonists, metergoline, dopamine D2 antagonist metoclopramide lysine acetyl), enkephalinase inhibitors neutral endopeptidase), 5-HT2 antagonists

LY-

053857), 5-HT3 antagonists Dolasetron mesilate/MDL-73147, 4H-carbazol-4-one derivatives, and the like), tenosal, tolfenamic acid, cyclooxygenase inhibitors carbasalate/carbaspirin calcium, tenosal/MR-Y134, and the like), alpha adrenoreceptor antagonists arotinolol, dihydroergocryptine, and the like), opioid agonists flupirtine/D-9998), beta adrenergic antagonists S 35 propranolol), valproate semisodium, and the like.

Additional treatments which lead to the overproduction of nitric oxide include administration of antiarthritic agents, such as anti-CD4 monoclonal antibodies, phospholipase Al inhibitor, loteprednol, tobramycin, combination of loteprednol and tobramycin, salnacedin, amiprilose, anakinra, anergiX, anti-B7 antibody, anti-CD3H, anti-gp39, anti-MHC MAbs, antirheumatic peptides, AP-1 inhibitors, AR-324, purine nucleotide phosphorylase inhibitors BCX-5), bindarit, CD2 Libc/03675 antagonist BTI-322), campathlH, CD4 antagonist CE9.1, SB-210396, and the like), tumour necrosis factor antagonist p80 TNFR, rhTNFbp, peptide T, CenTNF, thalidomide, CDP-571, TBP-1, and the like), cobra venom factor, interleukin la agonist cytogenin), interleukin 2 receptor antagonist dacliximab), ICAM 1 antagonist enlimomab), interleukin 1 beta converting enzyme inhibitors ICE inhibitors), interferons thymocartin), interleukin-10 interleukin-13, interleukin 1 antagonist SR-31747, TJ-114, and the like), interleukin-2 antagonist sirolimus), phospholipase C inhibitor, neurokinin 1 antagonist L-733060), laflunimus, leflunomide, leucotriene antagonists, levamisole, LFA3TIP, macrocyclic lactone, MHC class 11 inhibitors, mizoribine, mycophenolate mofetil, NfkB inhibitors, oncolysin CD6, peldesine, pidotimod, PKC-RACK inhibitors, PNP inhibitors, reumacon, CD28 antagonist, roquinimex, RWJ-50271, subreum, T7 vector, tacrolimus, VLA antagonist TBC-772), transforming growth factor beta agonist, methionine synthase inhibitors vitamin B12 antagonist), adenosine A2 receptor agonist YT-146), antagonist zolimomab), 5-lipoxygenase inhibitor zileuton, tenidap, ABT-761, and the like), cyclooxygenase inhibitor tenoxicam, talmetacin, piroxicam cinnamate, oxaprozin, NXTHIO, ML- 3000, mofezolac, nabumetone, flurbiprofen, aceclofenac, diclofenac, dexibuprofen, and the like), metalloproteinase inhibitor XR-168, TNF convertase inhibitors, GI-155704A, AG-3340, BB-2983, and the like), nitric oxide synthase inhibitors ARL-16556), phospholipase A2 inhibitor ARL- 67974), selectin antagonist CAM inhibitors), leucotriene 84 antagonist CGS-25019C), collagenase inhibitor GR-129574A), cyclooxygenase 2 inhibitor meloxicam), thromboxane synthase inhibitor curcumin), cysteine protease inhibitor GR-373), metalloproteinase inhibitor (D-5410), lipocortins synthesis agonist rimexolone, predonisolone 21-farnesylate,

HYC-

141, deflazacort, and the like), chelating agent diacerein), elastase inhibitors, DNA directed RNA polymerase inhibitor estrogens), oxygen radical formation antagonist glucosamine sulfate), thrombin inhibitors GS-522), collagen inhibitors halofuguinone), hyaluronic acid agonist NRD-101, hylan, Dispasan, Hyalart, and the like), nitric oxide antagonists (eg., hydroxocobalamin), stromelysin inhibitors L-758354), prostaglandin El agonist misoprostol, misoprostol+diclofenac, and the like), dihydrofolate reductase inhibitor trimetrexate, MX-68, and the like), opioid antagonist nalmefene), corticotropin releasing factor antagonist NBI-103, NBI-104, and the like), proteolytic enzyme inhibitor protease nexin-1, NCY-2010, and the like), bradykinin antagonist tachykinin antagonists, NPC-17731, and the like), growth hormone antagonist octreotide), phosphodiesterase IV inhibitor PDEIV inhibitors), gelatinase inhibitor REGA-3G12), free radical scavengers SIDR-1026), prostaglandin synthase inhibitors (eg., sulfasalazine), and the like.

Additional treatments which lead to the overproduction of nitric oxide include administration of agents useful for the treatment of septic shock, such as angiogenesis inhibitors OLX-514), Sbradykinin antagonists CP-0502, NPC-17731, and the like), complement factor inhibitors C3 convertase inhibitor), C5a release inhibitors CAB-2.1), dopamine agonists dopexamine), elastase inhibitors ONO-5046), E selectin antagonists CY-1787), farnesyltransferase inhibitors RBE limonene), immunostimulants CGP-19835A, lipid A vaccine, edobacomab, nebacumab, StaphGAM, diabodies, and the like), immunosuppressants CytoTAB, Libcl03675 transcyclopentanyl purine analogues, and the like), interleukin 1 antagonists interleukin 1 receptors), interleukin 1 receptor 5 antagonists anakinra), interleukin Ib antagonists (eg., interleukin-1p), interleukin Ibeta converting enzyme inhibitors ICE-inhibitors), interleukin 8 antagonists IL-8 receptor), interleukin 13 agonists intereleukin-13), ITF-1697, lipase clearing factor inhibitors SC-59735), membrane permeability enhancers Bactericidal Permeability Increasing protein/BPI), nitric oxide antagonists hydroxocobalamin), nitric oxide synthase inhibitors L-NMMA, a-methyl-Ndelta-iminoethyl-ornithine, and the like), P2 receptor stimulants ATP analogues), phosphatidic acid synthesis antagonists lisofylline), phospholipase A2 inhibitors S-448, acylpyrrole-alkanoic acid derivatives, indoleacetic acid derivatives, and the like), platelet activating factor antagonists ABT-299, TCV-309, SM-12502, (2RS,4R)-3(2-(3pyridinyl)thiazolidin-4-oyl)indoles, UR-12670, E-5880, and the like), prostacyclin agonists (eg., taprostene), prostaglandin El agonists TLC C-53), protein kinase inhibitors SB-203580), protein kinase C inhibitors, protein synthesis antagonists procysteine), proteolytic enzyme inhibitors nafamostat), SDZ-PMX-622, selectin antagonists sulfated glycolipid cell adhesion inhibitors), thrombin inhibitors GS-522), TNF receptor-Ig, tumour necrosis factor antagonists (eg., anti-TNF MAbs, MAK-195F, TBP-1, Yeda, rhTNFbp, CDP-571, and the like), tumour necrosis factor alpha antagonists E-5531), and the like.

Still further treatments which lead to the overproduction of nitric oxide include administration of agents for the treatment of multiple sclerosis, such as 4-aminopyridine, 15+deoxyspergualin,

ACTH,

amantadine, antibody adjuvants poly-ICLC, poly-lC+poly-Llysine+carboxymethylcellulose, and the like), anti-cytokine MAb CDP- 835), anti-inflammatory agents CY-1787, CY-1503, and the like), anti-selectin MAb 5 CY-1787), anti-TCR MAb NBI-114, NBI-115, NBI-116, and the like), bacloten, bethanechol chloride, carbamazepine, carbohydrate drugs CY-1503), clonazepam, CNS and immune system function modulators NBI-106, NBI-107, and the like), cyclophosphamide, cyclosporine A, cytokines IFN-a, alfaferone, IFN-P Ib, betaseron, TGF-P2, PEG-TGF-P2, betakine, IFN-plRebif, frone, interferon-p, IFN-P, and the like), CD4+T cell inhibitors AnergiX), CD28 antagonists B7-1, B7-2, CD28, and the like), direct cytotoxicity therapies benzoporphyrin derivative FK-506, growth factors glial growth factor, GGF, nerve growth factors, TGF-P2, PEG-TGF-P2, betakine, and the like), humanised MAb anti-IFN-yMAb, smart anti-IFN-yMAb, anti-Tac antibody, smart anti-Tac antibody, and the like), humanised anti-CD4 MAb anti-CD4 MAb, centara, and the like), hydrolase stimulants castanospermine), IFN-a, IFN-y antagonists anti-IFN-yMAb, smart anti IFNy MAb, and the like), IL-2 antagonists (eg., tacrolimus, FK-506, FR-900506, Fujimycin, Prograf, IL-2 fusion toxin, DAB3891L-2, and the like), IL-4 antagonists IL-4 fusion toxin, DAB 3 8 91L-4, and the like), immune-mediated neuronal damage 35 inhibitors NBI-114, NBI-115, NBI-116, and the like), immunoglobins, immunostimulants (eg., Spoly-ICLC, edelfosine, ALP, ET-18-OCH3, ET-18-OME, NSC-24, poly-IC+poly-Llysine+carboxymethylcellulose, and the like), immunosuppressants azathioprine, Al-100 animal protein, rDNA human protein Al-101, peptide, Al-102, castanospermine, tacrolimus, FK-506, FR- 900506, Fujimycin, Prograf, anti-leukointegrin MAb, Hu23F2G, primatised anti CD4 antibody, CE9.1, Galaptin 14-1, GL14-1, Lectin-1, recombinant IML-1, linomide, roquinimex, LS-2616, transcyclo- Lbdc/03675

I

pentanyl purine analogues, MS-6044, spanidin, 15-deoxyspergualin, deoxyspurgiline, gusperimus HCL, NSC-356894, NKT-01, TCR, CD3/Ti, cyclosporine, OL-27-400, Sandlmmune, Human monogens, anti-TCR MAbs, TCAR MAbs, Monogen TM19, Monogen TM27, Monogen TM29, Monogen 5 TM31, peptigen TP12, anti-CD4 MAb, cantara, immunophilins, VX-10367, VX-10393, VX- 10428, synthetic basic copolymer of amino acids, copolymer-1, COP-1, T lymphocyte immunofusion (TIF) protein, cyclophosphamide, and the like), integrin antagonists anti-integrin monoclonal antibodies, AN-100225, AN-100226, and the like), interferon agonists poly-ICLC, poly-IC+poly-Llysine+carboxymethylcellulose, and the like), interferon-plb, isoprinosine, IV methylprednisolone, macrolides tacrolimus, FK-506, FR-900506, Fujimycin, Prograf, and the like), MAO B inhibitors selegiline, Parkinyl, and the like), methotrexate, mitoxantrone, muscle relaxants

RGH-

5002), muscarinic antagonists RGH-5002), neurosteroids NBI-106, NBI-107, and the like), octapeptides peptide oxybutinin chloride, oxygen free radical antagonists tetrandrine, biobenzylisoquinoline alkaloid, and the like), peptide agonists peptide phenoxybenzamine, phospholipase C inhibitors edelfosine, ALP, ET-18-OCH3, ET-18-OME, NSC-24, and the like), photodynamic therapies benzoporphyrin derivative plasmapheresis, platelet activating factor antagonists ginkgolide B, sN-52021, and the like), potassium channel antagonists (eg., aminodiaquine, EL-970, and the like), propranolol, prostaglandin synthase inhibitors (eg., sulfasalazine, salazosulfa-pyridine, PJ-306, SI-88, azulfidine, salazopyrin, and the like), protease antagonists ginkgolide B, BN-52021, and the like), recombinant soluble IL-1 receptors, spergualin analogues spanidin, 15-deoxyspergualin, deoxyspurgiline, gusperimus HCL, NSC- 356894, NKT-01, and the like), TCR peptide decoys NBI-114, NBI-115, NBI-116, and the like), TCR peptidomimetic decoys NBI-114, NBI-115, NBI-116, and the like), TCR peptide vaccines AI-208 (VS6.2/6.5 phenotype)), selectin antagonists lectin-1, recombinant IML- 1, and the *like), soluble TNF receptor I, TCARs TCR, CD3/Ti, peptigen TP12, and the like), TNF antagonists thalidomide, TNF inhibitors, and the like), tricyclic antidepressants, and the like.

Additional treatments which lead to the overproduction of nitric oxide include administration of organ transplantation agents, such as anti-CD25 MAbs, anti-Tac antibodies, anti-TNF MAb (eg., CDP571), apoptosin, azathioprines imuran), BCX-34, CA3, CD28, complement inhibiting factors CD59), CTLA41g, cyclosporines CsA), FK-506/rapamycin binding proteins (FKBP), glucocorticoids, humanised version of OKT3 huOKT3-185), hydroorotate dehydrogenase inhibitors Brequinar), orthoclone OKT3 IgG2a anti-T cell murine monoclonal antibody, muromonab-CD3, and the like), rapamycins AY-22989), streptomyces isolates FR-900520, FR-900523, and the like), and the like.

Additional treatments which lead to the overproduction of nitric oxide include administration of 35 agents for the treatment of systemic lupus erythematosus (SLE), such as androgen-derived steroids Org-4094), anti-CD4 humanised antibodies, anti-DNAIV-88, anti idiotypic murine MAb antiidiotypic antibody to 3E10/MAblC7), CD2 antagonists CD2), complement inhibitors (eg., recombinant MCP-based complement inhibitors), cyclosporines Sandimmune, cyclosporine analog, OG-37325, cyclosporin-G, NVal-CyA, and the like), cytokines IL-4 fusion toxin), cytokine receptor antagonists immunomodulatory cytokines), E-selectin antagonists anti-ELAM, CY- Libc103675 1787, and the like), FK506/tacrolimus Prograf), hypercalcemic agents KH-1060), IFN-y antagonists anti-IFN-y MAb, smart anti-IFN-y MAb, and the like), IL-1P converting enzyme inhibitors (ICE), IL-2 produced by E. coli celmoleukin, IL-2, TGP-3, Celeuk, and the like), immunoglobulins anti-ELAM, CY-1788, humanised CY-1787, and the like), immunostimulants thymotrinan, RGH-0205, TP3, and the like), immunosuppressants Rapamycin, AY-22989, NSC-226080, NSC-606698, anti-CD4, T-cell inhibitor, anti-tac MAb, smart anti-tac MAb, Migis (membrane immunoglobulin-isotope specific) antibodies, SM-8849, immunophilins, VX-10367, VX- 10393, VX-10428, mycophenolate mofetil, ME-MPA, RS-61444, cyclosporine, OL-27-400, Sandimmune,'IL-4 fusion toxin, trypanosomal inhibitory factor (TIF), T-cell receptor, CD3/Ti, Org- 4094, anti-TBM, CP 17193, Leflunomide/A-77-1726, ELAM-1, AnergiX, Spanidin, deoxyspurgiline, gusperimus hydrochloride, NSC-356894, NKT-O1, Roquinimex, LS-2616, linomide, LJP-394, CD-59 antigen, and the like), immunotoxins Zolimomab aritox, OrthoZyme-CD5+, XomaZyme-H65-rta, Xomazyme-CD5 Plus, and the like), intravenous immunoglobulins IVIG), integrin antagonists integrin blockers), Migis

TM

antibodies, monoclonal antibody therapeutics, murine MAb anti-SLE vaccine, MAb 3E10, and the like), primatised anti-CD4 antibodies CE9.1), protease inhibitors matrix metalloprotease (MMP) inhibitors, stromelysin, and the like), protein synthesis antagonists anti-CD6-bR, anti-T12bR, oncolysin CD6, and the like), purine nucleoside phosphorylase inhibitors BCX-25, BCX-14, and the like), selectin antagonists CY1503, Cylexin, and the like), spergualin analogues (eg., Spanidin, 15-deoxyspergualin, deoxyspurgiline, gusperimus hydrochloride, NSC-356894, NKT-01, and the like), T cell inhibitors AnergiX), tumour necrosis factor (TNF) antagonists, and the like.

Additional treatments which lead to the overproduction of nitric oxide include administration of agents for the treatment of Alzheimer's disease, such as ACh release enhancers T-588 (benzothiophene 5 derivative)), acetylcholine release stimulants DUP-996 and analogues), AMPA agonists AMAlex, Isoxazole compound series, and the like), AMPA GluR agonist (eg., IDRA-21 [7-chloro-3-methyl-3,4-dihydro-2H-1 ,2,4benzothiadiazininel), AMPA GluR antagonists (eg., S-18986 and related quinolone derivatives), anticholinesterases E-2020), Ca-antagonists (eg., NS-649, spider venom derived ICM peptides and analogues, substituted 2-aminoindanes compound series, and the like), combined anticholinesterase and muscarinic AChR antagonists PD142676), K-channel blockers Trans-R-4-(4-methoxyphenyl-methyl) cyclohexylanine and analogues, margatoxin-based functional and/or structural analogues, and the like), MI muscarinic receptor agonists Xanomeline), NMDA antagonists certain indole derivatives, (R-(R 1

,S

1 hydroxyphenyl)-p-methyl-4(phenylmenthyl)- -piperidinepropanol and analogues thereof, and the like), nicotinic AChR agonists ABT-418 [isoxazole, 3-meth-5-(1-meth-2-pyrrolidinyl)], and the like), and the like.

***Additional treatments which lead to the overproduction of nitric oxide include administration of agents for the treatment of psoriasis, such as 5-LO inhibitors Wy-50295, Wy-49232, Lonapalene, RS-43179, MK-886, L-663536, ETH-615, DUP-654, Zileuton, epocarbazolin-A, A-64077, and the like), inhibitors BF-397, Tenidap, CP-309, CP-66248, and the like), angiogenesis inhibitors platelet factor anticancer antibiotic AGM-1470, TNP-470, and the like), anti-inflammatory Libcd03675 cytochrome P450 oxidoreductase inhibitors DuP-630, DuP-983, and the like), antiproliferative compounds Zyn-Linker), arachidonic acid analogues CD581, CD554, and the like), arachidonic acid antagonists Lonopalene, RS-43179, triamcinolone acetonide with penetration enhancer Azone, betamethasone dipropionate steroid wipe, G-202, Halobetasol propionate, ultravate, s Halometasone, C-48401-Ba, Sicorten, and the like), beta-glucan receptor antagonists, betamethasone steroid wipes, calcium metabolic moderators Tacalcitol, Bonealfa, TV-02 ointment, Ro-23-6474, KH-1060, Calcipotriol, BMS-181161, BMY-30434, Dovonex, Divonex, and the like), CD4 binding inhibitors PIC 060), cell adhesion compounds CY-726, VCAM-1, ELAM-1, ICAM, and the like), cell adhesion inhibitors selectin inhibitor, GM-1930, and the like), cellular aging inhibitors Factor corticosteroids Halobetasol propionate, ultravate, Halometasone, C-48401Ba, Sicorten, and the like), cyclosporin analogues IMM-125), dihydrofolate reductase inhibitors (eg., G-301, dichlorobenzoprim, methotrexate, methotrexate in microsponge delivery system, and the like), E-selectin inhibitors ISIS 4730), endogenous active form of vitamin D3 Calcitriol, Du- 026325, and the like), fibroblast growth factor antagonists Saporin mitotoxin, Steno-Stat, and the like), fumagillin analogues AGM-1470, TNP-470, and the like), G-proteins and signal transduction compounds CPC-A), gel formulations for acne nicotinamide, N-547, Papulex, and the like), growth hormone antagonists Octreotide, Sandostatin, Lanreotide, angiopeptin, BIM-23014, Somatuline, and the like), humanised antibodies anti-CD4 antibody), hydroorotate dehydrogenase inhibitors Brequinar sodium, bipenquinate, DuP-785, and the like), ICAM-1 inhibitors ISIS 939), IL-1 and other cytokine inhibitors Septanil), IL-1 converting enzyme inhibitors, IL-1 receptor antagonists Antril), IL-2 antagonists Tacrolimus, Prograf, FK-506, and the like), IL-2 receptor-targeted fusion toxins DAB3891L-2), IL-8 receptors, immunostimulants Thymopentin, Timunox, and the like), immunosuppressants XomaZyme-CD5 Plus, cyclosporine, Sandimmune, SR-31747, anti-CD11, 18 MAb, Tacrolimus, Prograf, FK-506, FK-507, and the like), immunosuppressive agents targeting FK506 immunophilins, VX 1 0 3 6 7, VX 1 0 4 2 8, and the like), immunotoxins MAb directed against CD antigen XomaZyme-CD5 Plus), leukotriene antagonists Sch-40120, Wy-50295, Wy-49232, and the like), leukotriene B4 antagonists SC-41930, SC-50605, SC-48928, ONO-4057, LB-457, LY-255283, LY-177455, LY- 223982, LY-223980, LY-255253, and the like), leukotriene synthesis inhibitors MK-886, L- 663536, and the like), lipase clearing factor inhibitors 1-docosanol, lidakol, and the like), lipid encapsulated reducing agent Dithranol), liposomal gel Dithranol), LO inhibitors (eg., CD581, CD554, Masoprocol, Actinex, and the like), lithium succinate ointments lithium salts, ,:oo Efalith, and the like), LO/CO inhibitors P-8892, P-8977, CHX-108, FPL-62064, and the like), membrane integrity agonists lithium salts, Efalith, and the like), microtubule inhibitors (eg., S 35 Posophyliotoxin-containing compound, Psorex, and the like), octapeptide somatostatin analogues Lanreotide, angiopeptin, BIM-23014, Somatuline, and the like), oligonucleotides ISIS 4730, ISIS 3801, ISIS 1939, IL-1 inhibitors, and the like), peptide agonists octapeptide, peptide T, and the like), PKC inhibitors, phospholipase A2 compounds, pospholipase D compounds, photodynamic anticancer agents 5-aminolevulinic acid, 5-ALA, and the like), Libc103675 photodynamic therapies benzoporphyrin derivatives, synthetic chlorins, synthetic porphyrins, EF-9, and the like), photosensitiser Porfirmer sodium), PKC inhibitors Safingol, Kynac, and the like), platelet activating factor antagonists TCV-309), platelet aggregation inhibitors (eg., CPC-A), prodrug NSAIDs G-201), prostaglandin agonists eicosapentaenoic acid gammalinolenic acid combination, Efamol Marine, and the like), protein inhibitors SPC-103600, SPC- 101210, and the like), protein kinase C (PKC) inhibitors Ro-31-7549, Ro-31-8161, Ro-31-8220, and the like), protein synthesis antagonists Calcitriol, Du-026325, LG-1069, LG-1064, AGN- 190168, Namirotene, CBS-211A, and the like), purine nucleoside phosphorylase inhibitors BCX- 34), radical formation agonists benzoporphyrin derivatives), recombinant antileukoproteinases ALP-242), retinoids BMY-30123, LG-1069, LG-1064, and the like), retinoid derivatives (eg., AGN-190168), rapamycin binding proteins (FKBP) immunophilins, VX-10367, VX-10428, and the like), second generation monoaromatic retinoids Acitretin, Neotigason, and the like), soluble IL-1, IL-4 and IL-7 receptors, somatostatin analogues Octreotide, Sandostatin, and the like), steroids AGN-191743), streptomyces anulatus isolates epocarbazolin-A), superoxide dismutase EC-SOD-B), thymidylate synthase inhibitors AG-85, MPI-5002, 5-FU in biodegradable gellike matrix, 5-FU and epinephrine in biodegradable gel-like matrix, AccuSite, and the like), topical formulations P-0751, P-0802, and the like), transglutaminase inhibitors, tyrphostin EGF receptor kinase blockers AG-18, AG-555, and the like), VCAM-1 inhibitors ISIS 3801), vitamin D analogues Ro-23-6474, KH-1060, Calcipotriol, BMS-181161, BMY-30434, Dovonex, Divonex, and the like), vitamin 03 analogues Tacalcitol, Bonealfa, TV-02 ointment, and the like), vitamin D 3 derivatives 1,2-diOH-vitamin D3), and the like.

Still further treatments which lead to the overproduction of nitric oxide include administration of agents for the treatment of diabetes, such as ACE inhibitors captopril), amylin agonists and antagonists Normylin

TM

AC137, GC747, AC253, AC625, and the like), autoimmune compounds 25 Al-401),capsaicins Zostrix-HP), cell regulators protein kinase C inhibitors, Balanol, and the like), domperidones Motilium@), fluvastatins Lescol), FOX 988, fusion toxins (eg.,

DAB

3 891L-2, DAB 486 1L-2, and the like), gene therapies Transkaryotic Therapies), glucagons (eg., recombinant yeast glucagon), IL-10 compounds, iloprost, immunosuppressives tacrolimus, S Prograf, FK-506, and the like), insulin analogues AI-401, Nu-lnsulin compounds, Humulin, Iletin, 30 Humalog T M w, Lys-Pro, Amaryl, and the like), insulin-like growth factors Chiron/Ciba-Geigy compounds, Fujisawa compounds, Genentech compounds, and the like), insulinotropins (eg., PfizerlScios Nova compounds), nerve growth factors Genentech compounds), oral hypoglycemics AS-6, glimepiride, Amaryl, CL 316,243, acarbose, miglitol, recombinant yeast glucagon, GlucaGen T M NovoNorm T M glipizide, insulinotropin, CI-991/CS-045, and the like), platelet- 35 derived growth factors ZymoGenetics/NovoNordisk compounds), sulfonylureas tolbutamide, acetohexamide, tolazamide, chlorpropramide, and the like), T cell approaches anergize, AnergiXTMw, Procept compounds, T cell Sciences compounds, and the like), tolrestats (eg., Alredase®, ARI-509, and the like), and the like.

Additional treatments which lead to the overproduction of nitric oxide include the administration of agents for the treatment of stroke, such as 5-HT antagonists Piperazine derivatives), Libcl03675 reuptake inhibitors Milnacipran, Dalcipran, and the like), 5-HT 1A agonists SR-57746A,

SR-

57746, and the like), 5-HT 3 agonists SR-57227), 5-HT 4 antagonists, 5-lipoxygenase inhibitors low MW dual 5-lipoxygenase and PAF inhibitor CMI-392), ACH agonists Pramiracetam, Choline-L-alfoscerate, L-alphaglycerylphosphoryl-choline, Delecit, and the like), adenosine agonists GP-1-4683, ARA-100, arasine analogues, and the like), adenosine Al receptor agonists (eg., Azaisotere, 2-chloro-N-[4 (phenylthio)-l-piperidinyl] adenosine, 2120136, and the like), adenosine reuptake inhibitors Diphenyloxazole derivatives), adrenergic transmitter re-uptake inhibitors (eg., Bifemelane, E-0687, MCI-2016, Alnert, Celeport, and the like), aldose reductase inhibitors Spiro- 3' pyrroline derivatives), alpha antagonists Drotaverine acephyllinate, Depogen, and the like), alpha 2 agonists SNAP-5083, SNAP-5608, SNAP-5682, and the like), AMPA receptor agonists heterocyclic compound SYM-1207, heterocyclic compound SYM- 1252, and the like), AMPA receptor antagonists LY-293558, LY-215490, and the like), Ancrod/Arvin, aspirin, benzothiazoles Lubeluzole, R87926, and the like), benzodiazepine receptor antagonists 3-oxadiazolyl-1,6-naphthyridine derivatives, Tetracyclic imidazodiazepineseries imidazenil, FID-02-023, Ro-23-1412, and the like), blood substitutes, bradykinin antagonists CP-0127, Bradycor, Septicor, and the like), C5a release inhibitors (eg., protein derivative CMI-46000), calcium antagonists Lemildipine, NB-818, NPK-1886, Trimetazidine derivatives, lomerizine KP-2796, Diltiazem analog clentiazem maleate, TA-3090, and the like), calcium channel antagonists nitrendipine-like compound diperdipine, YS-201, U-92032, Diltiazem derivative, 1058, SM-6586, KP-840, F-0401, D-31-D, tetrahydronaphthalene derivatives, :i fasudil, AT-877, H-7, HA-1044, HA-1077, Eril, darodipine, dazodipine, PY-108-068, Plimo, Dihydropyridine, AE 0047, GJ-0956, Lacidipine, GR-43659, GR-43659X, GX-1048, S-312-d, S-312, S-830312, Nilvadipine, FK-235, and the like), calpain inhibitors AK-275, CX-275, and the like), carnitine palmitoyl-transferase inhibitors, carvedilol, cell adhesion molecular technology, cerebral calcium antagonist vasodilators Nimodipine, Nimotop, and the like), cholinesterase inhibitors indole and indazole derivatives, Tacrine analogues, and the like), complement factor inhibitors TK9C, protein derivative TPi6, compinact A, compinact C, Factor D inhibitors, soluble, recombinant MCP-based complement inhibitors, and the like), complement inhibitors sCRI/BRL- 55730, YM-203, and the like), coronary vasodilators Nicorandil, RP-46417, SG-75, Adancor, and the like), CPC-111, cytidyl diphosphocholine/citicholines, cytokines NBI-117), Dexanabiol, dopamine agonists, EAA receptors, endothelin antagonists SB 209670), endothelin receptor antagonists, excitatory amino acid agonists acylated polyamine analogues, N-(4hydroxyphenylpropanoyl)-spermine analogues, and the like), excitatory amino acid antagonists (eg., Tryptophan, 4,6-disubstituted stroke kynurenine derivatives, NPC-17742, CPC-701, CPC-702, and the like), glutamate antagonists Kainate quisqualate NNC-07-9202, NPC-17742, small molecule CNS-1237, NS-257, NS-072, BW-619C, CGS 19755, Riluzole, PK-26124, RP 54274, and the like), glutamate receptor antagonists Araxin compounds, Quinoxaline derivative, YM-90K, YM-900, and the like), glycine antagonists, glycine NMDA agonists 3-hydroxy-2,5-dioxo-1

H-

benz[b]azepines), glycine NMDA associated antagonists 5,6-dihydro-1H-pyrrolo[1,2,3de]quinoxaline-2,3-diones, Strychnine-insensitive glycine binding site of NMDA receptor L-687414, Libd03675 Glystasins, ACEA-2011, ACEA-3031, AC-1021, ACPC, eliprodil, and the like), growth factor antagonists non-peptide indolocarbazole neutrophic molecules, CEP-075, and the like), GPllblilla antagonists Peptide C68-22), haemorheological agents Drotaverine acephyllinate, Depogen, and the like), heparin, hydroxyl radical formation inhibitors (eg., homopiperazine derivative K-7259), hypocalcemic agents calcitonin peptide, related to hCGRP peptide), hypothermic agents/BMY-20862, ICAM-1 compounds Enlimomab), immunosuppressants small molecule compounds, NBI-117, and the like), integrin general antagonists monoclonal antibody AN-100225, monoclonal antibody AN-100226, and the like), Interleukin-1 antagonists cyclic nitrones), iron-dependent lipid peroxidation inhibitors 2- (amino-methyl) chromans), lactic acid accumulationlinhibitors small molecule CPC-211), Leukotriene B4 antagonists Ebselen, DR-3305, PZ-25, PZ-51, RP 60931, RP 61605, and the like), lipid peroxidase inhibitors Idebenone, Avan, and the like), low molecular weight small molecules, methyltransferase stimulants 4-methyl benzenesulfonate, ademetionine sulfate tosilate, FO-156, Ceritan, and the like), monoamine oxidase B inhibitors MD-280040, MD- 200243, MD-280080, Lazabemide, Ro-19-6327, and the like), MS-153, MS-424, INaI/H+Na+/Li+ exchange inhibitors Pyrazine derivatives), nadroparin Fraxiparin), nafronyllnaftidrofuryl (eg., Praxilene), nerve growth factor agonists small molecule compounds, CNTF, BDNF, 2.5S NGF, monosialoganglioside GM1, Sigen/Sygen, and the like), neuronal calcium channel blockers CPC- 304, CPC-317, and the like), neuronal differentiation compounds F-spondin), neuropeptide agonists Neurotrophic Peptide Trofexin), neutrophil inhibitory factors small molecule compounds), nitric oxide agonists hydroxy derivative N-3393, hydroxy derivative N-3398, nicorandil, Therapicon, and the like), nitric oxide antagonists, NMDA antagonists (eg., Spiroisoindoles/dizocilpine derivatives, Oxindole compound, CP-112116, LY-104658, LY-235959, FR- 115427, Sialic acid derivative, N-palmitoyl-Betaethylglycoside neuraminic acid, ND-37, Ro-01-6794, 706, Dextrorphan, Ifenprodil analogue eliprodil, SL-82.0715, Lipophilic molecules, HU-211, Remacemide, 934-423, 12495, 12859, 12942AA, Selfotel, CGS-19755, SDZ-EAA-494, CGP-40116, CGP-37849, CGP-39551, CGP-43487, and the like), NMDA antagonist-partial agonists (eg., Conantokin G peptide SYM-1010), NMDA channel blockers Aptiganel, CERESTAT, CNS 1102, and the like), NMDA receptor antagonists, NMDA receptor subtypes Kainate quisqualate NNC- 07-9202), non-competitive NMDA antagonists FPL-15896), non-ionic copolymer RheothRx, nootropic/acetylcholine agonists Oxiracetam, CT-848, Neuractiv, and the like), norepinephrine inhibitors Midalcipran), N-type calcium channel antagonists NS-626, NS-638, and the like), opioid antagonists Nalmefene, nalmetrene, JF-1, ORF-11676, Cervene, Incystene, and the like), opioid kappa receptor agonists acrylacetamide enadoline, CI-997, and the like), organoselenims Ebselen, DR-3305, PZ-25, PZ-51, RP 60931, RP 61605, and the like), oxygen scavengers Tirilazad mesylate, Lazaroids, Freedox, and the like), PA2 inhibitors (eg., Sphphospholipase A2 inhibitor), PAF antagonists nupafant, BB-2113, and the like), partial glycine NMDA agonists ACPC), peptide/GPIlblIlla antagonists Integrelin), peptidic neuronspecific calcium channel antagonists SNX-111), phosphodiesterase inhibitors Xanthine derivatives, propentofylline, Hoe-285, Hextol, and the like), phospholipase A2 inhibitors small Libc03675 21 organic molecule CEP-217), plasminogen activators r-ProUK (recombinant pro-urokinase), platelet-activating factor antagonists UK-74505), platelet adhesion inhibitors Peptide), platelet aggregation antagonists cilostazol, peptide agents, GPHb-IIIA inhibitor, TP-9201, and the like), platelet aggregation inhibitors Diaminoalkanioic acid derivatives), potassium channel agonists Nicorandil, RP-46417, SG-75, Adancor, and the like), prolyl endopeptidase

(PEP)

inhibitors JTP 4 819 protein kinase C inhibitors monosialoganglioside derivative Ligaproteolytic enzyme inhibitors Protease nexin-1, Incyte, PN-1, PN-2, Nafamostat, FUT-175, Duthan, Futhan, and the like), pyrimidine derivatives, Quinolizine derivatives KF-17329, KF- 19863, and the like), radical formation antagonists EPC-K1), recombinant tissue plasminogen activators alteplase, Activase, and the like), Schwann cell derived molecules/promoters, sigma antagonists Sigma ligand), sigma receptor antagonists tetrahyropyridinyl-isoxazolines, isoxazoles PD-144418, and the like), sodium/calcium channel modulators Lifarizine, RS-87476, and the like), sodium channel antagonists, streptokinase Streptase), substituted guanadine (eg., small molecule CNS-1237), superoxide dismutase stimulants PEG conjugated enzyme superoxide dismutase/Dismutec, PEG-SOD, and the like), thrombin inhibitors, non-peptide), thromboxane synthase inhibitors Linotroban, HN-11500, and the like), thyrotropin-releasing hormone agonists TRH agonists, Protirelin analogthymoliberin, RX-77368, and the like), ticlopidine Ticlid), TJ,8007, TRH agonists Thyrotropin releasing hormones, JTP-2942, and the like), trilazard, urokinase Abbokinase), w-conopeptide SNX-III), warfarin (eg., Coumadin), and the like.

:i Accordingly, presently preferred indications for treatment in accordance with the present invention include septic shock, ischaemia, ulcers, ulcerative colitis, diabetes, arthritis, asthma, Alzheimer's disease, Parkinson's disease, multiple sclerosis, cirrhosis or allograft rejection, and the like.

25 In accordance with a particular aspect of the present invention, the nitric oxide scavenging agent is administered in combination with one or more of the above described agents, optionally including an antibiotic gentamicin, tobramycin, amikacin, piperacillin, clindamycin, cefoxitin or vancomycin, or mixtures thereof), a vasoactive agent a catecholamine, noradrenaline, dopamine or dobutamine), or mixtures thereof. In this way, the detrimental side effects of many of the above- 30 noted pharmaceutical agents and/or the indications they are designed to address systemic hypotension) can be prevented or reduced by co-administration of a combination reagent including a nitric oxide scavenger.

SThose of skill in the art recognise that the combination of an agent capable of inactivating species which induce the expression of inducible nitric oxide (or an agent capable of inhibiting the production of such species), and nitric oxide scavengers described herein can be delivered in a variety of ways, such as, for example, orally, intravenously, subcutaneously, parenterally, rectally, by inhalation, and the like.

Since individual subjects may present a wide variation in severity of symptoms and each drug has its unique therapeutic characteristics, the precise mode of administration, dosage employed and treatment protocol for each subject is left to the discretion of the practitioner.

Libc/03675 In accordance with still another embodiment of the present invention, there are provided physiologically active composition(s) comprising a "therapeutic agent" (as described herein) and a nitric oxide scavenging compound a compound having the structure I, as described above), in a suitable vehicle rendering said composition amenable to oral delivery, transdermal delivery, Sintravenous delivery, intramuscular delivery, topical delivery, nasal delivery, and the like.

Depending on the mode of delivery employed, the above-described compositions can be delivered in a variety of pharmaceutically acceptable forms. For example, the above-described compositions can be delivered in the form of a solid, solution, emulsion, dispersion, micelle, liposome, and the like.

Pharmaceutical compositions of the present invention can be used in the form of a solid, a Solution, an emulsion, a dispersion, a micelle, a liposome, and the like, wherein the resulting composition contains one or more of each of the nitric oxide scavenging and therapeutically active compounds contemplated for use in the practice of the present invention, as active ingredients thereof, in admixture with an organic or inorganic carrier or excipient suitable for enteral or parenteral applications. The active ingredients may be compounded, for example, with the usual non-toxic, pharmaceutically acceptable carriers for tablets, pellets, capsules, suppositories, solutions, emulsions, suspensions, and any other form suitable for use. The carriers which can be used include glucose, lactose, gum acacia, gelatin, mannitol, starch paste, magnesium trisilicate, talc, corn starch, keratin, colloidal silica, potato starch, urea, medium chain length triglycerides, dextrans, and other carriers suitable for use in manufacturing preparations, in solid, semisolid, or liquid form. In addition auxiliary, stabilising, thickening and colouring agents and perfumes may be used. The active compounds (ie., "therapeutic agents" and nitric oxide scavenging compounds compounds of structure I as described herein)) are included in the pharmaceutical composition in an amount sufficient to produce the desired effect upon the target process, condition or disease.

25 Pharmaceutical compositions containing the active ingredients contemplated herein may be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. Compositions intended for oral use may be prepared according to any method known in the art for the manufacture of pharmaceutical compositions. In addition, such compositions may contain one or more agents oil° selected from a sweetening agent (such as sucrose, lactose, or saccharin), flavouring agents (such as peppermint, oil of wintergreen or cherry), colouring agents and preserving agents, and the like, in order to provide pharmaceutically elegant and palatable preparations. Tablets containing the active ingredients in admixture with non-toxic pharmaceutically acceptable excipients may also be manufactured by known methods. The excipients used may be, for example, inert diluents such as calcium carbonate, lactose, calcium phosphate, sodium phosphate, and the like; granulating and disintegrating agents such as corn starch, potato starch, alginic acid, and the like; binding agents such as gum tragacanth, corn starch, gelatin, acacia, and the like; and lubricating agents such as magnesium stearate, stearic acid, talc, and the like. The tablets may be uncoated or they may be coated by known techniques to delay disintegration and absorption in the gastrointestinal tract, thereby providing sustained action over a longer period. For example, a time delay material such as Libc/03675 glyceryl monostearate or glyceryl distearate may be employed. They may also be coated by the techniques described in the US 4 256 108; 4 160 452; and 4 265 874, to form osmotic therapeutic tablets for controlled release.

In some cases, formulations for oral use may be in the form of hard gelatin capsules wherein the active ingredients are mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate, kaolin, or the like. They may also be in the form of soft gelatin capsules wherein the active ingredients are mixed with water or an oil medium, for example, peanut oil, liquid paraffin, or olive oil.

The pharmaceutical compositions may be in the form of a sterile injectable suspension. This suspension may be formulated according to known methods using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example, as a solution in l,3-butanediol. Sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or diglycerides, fatty acids (including oleic acid), naturally occurring vegetable oils like sesame oil, coconut oil, peanut oil, cottonseed oil, etc., or synthetic fatty vehicles like ethyl oleate or the like.

Buffers, preservatives, antioxidants, and the like can be incorporated as required.

Compositions contemplated for use in the practice of the present invention may also be administered in the form of suppositories for rectal administration of the active ingredients. These compositions may be prepared by mixing the active ingredients with a suitable nonirritating excipient, such as cocoa butter, synthetic glyceride esters of polyethylene glycols (which are solid at ordinary temperatures, but liquefy and/or dissolve in the rectal cavity to release the active ingredients), and the like.

S:

Since individual subjects may present a wide variation in severity of symptoms and each active ingredient has its unique therapeutic characteristics, it is up to the practitioner to determine a subject's 25 response to treatment and vary the dosages accordingly.

Typical daily doses of nitric oxide scavengers, in general, lie within the range of from about lOp10[g up to about 100mg per kg body weight, and, preferably within the range of from 50 tg to per kg body weight and can be administered up to four times daily. The daily IV dose lies within the range of from about 1 .g to about 100mg per kg body weight, and, preferably, within the range of from 10Jg to 10mg per kg body weight.

In general, the dosage of nitric oxide scavenger employed in the practice of the present invention falls in the range of about O.01mmoles/kg body weight of the subject/hour up to about The invention will now be described in greater detail by reference to the following non-limiting examples.

Example 1 Wistar rats (male, 230-300g) were supplied by Simonson Laboratories (Gilroy, CA).

Lipopolysaccharide (LPS; S. typhosa, endotoxin) was obtained from Sigma (St. Louis, MO).

Libcd03675 N-Methyl-D-glucamine and carbon disulfide were obtained from Aldrich (Milwaukee, WI). N- Methyl-D glucamine dithiocarbamate (MGD) was synthesised by following the method of Shinobu et al. (Acta Pharmacol. Toxicol. 54:189-194 (1984)).

Example 2 As described previously (see Lai and Komarov in FEBS Lett. 345:120-124 (1994)), one [(MGD)2/Fe] complex binds to one molecule of nitric oxide to form a [(MGD)2/FeNO] complex.

Whereas free nitric oxide is a potent vasodilator, nitric oxide bound to [(MGD)2/Fe] is not. The resulting complex is then excreted from the body in the urine, thereby reducing in vivo nitric oxide levels.

The effects of [(MGD)2/Fe] treatment on the mean arterial pressure of endotoxemia in rats are shown in Figure 1. When rats were treated with lethal doses of LPS, the mean arterial pressure dropped gradually with time and reached 75mm Hg at the end of 2h. In controls, when the animals were infused with saline, their mean arterial pressure remained very low; indeed, 11 out of 16 animals died before the end of the experiments. On the other hand, when the LPS-treated animals were infused with [(MGD)2/Fe], their mean arterial pressure gradually restored to normal levels, and only 3 out of 16 animals died before the end of the experiments. Therefore, infusions of [(MGD)2/Fe] can not only restore blood pressure, but also reduces the mortality rate in endotoxin induced septic shock rats.

In summary, [(MGD)2/Fe] is potentially useful for the treatment of systemic hypotension (extreme drop in blood pressure), caused by abnormally elevated levels of nitric oxide; a condition which has been associated with many inflammatory and infectious diseases. In addition, [(MGD)2/Fe] has been shown to be safe inasmuch as the animals survived after injections of up to 1% of their body weight without apparent side effects (Lai and Komarov, supra).

Example 3 As previously described (see Komarov and Lai in Biochim. Biophys. Acta 1272:29-36 (1995)), subcutaneous administration of the [(MGD)2/Fe] complex reduced in vivo -NO levels in LPS-treated mice. Since excessive *NO production is known to induce systemic hypotension, injections of the

[(MGD)

2 /Fe] complex that reduce in vivo -NO levels should also restore blood pressure in hypotensive animals induced by LPS treatment. To test this idea, experiments Were carried out to determine the effects of administration of the [(MGD)2/Fe] complex on the blood pressure of the hypotensive rats 30 induced by LPS challenge.

Thus, male Wistar rats (230-300g) fasted overnight were anaesthetised with thiobutabarbital (Inactin, 100mg/kg, A catheter was implanted in the femoral vein for drug infusions. The femoral artery was cannulated for continuous blood pressure measurement. Rats were injected with an i.v.

bolus dose of LPS typhosa endotoxin, 4mg/kg). Two hours after LPS challenge, rats were then subjected to one of the following treatments: Control, saline infusion- 1.OmL saline i.v. injection followed by 1.0mL/h of saline infusion for [(MGD)2/Fe] (at a ratio of 2-to-0.4)-0.1mmole/kg i.v. bolus injection followed by 0.lmmole/kg infusion for Libc/03675 [(MGD)2/Fe] (at a ratio of 2-to-0.2)-0.1mmole/kg i.v. bolus injection followed by 0.lmmole/kg infusion for 2.0h, and [(MGD)2/Fe] (at a ratio of 2-to-0)-0.1mmole/kg i.v. bolus injection followed by 0.lmmole/kg infusion for The results of mean arterial pressure (MAP) measurement as a result of each of these treatments are summarised in Table 1.

Table 1 Effects of various ratios of [(MGD)2Fe] treatment on the mean arterial pressure (MAP in mmHg) in lipopolysaccharide (LPS)-induced shock rats Conditions 1 Baseline 2 (mean±SEM) 2h after LPS Exposure 2.0h after Treatment a) Control saline (n=16) 3 96±2 77±2 76±7 b) [(MGD) 2 /Fe] (2/0.4)4 (n=16) 95±3 75±2 95±3 c) [(MGD)2/Fe] 98±2 75±3 89±4 d) MGD 99±4 71±2 94±6 1 Experimental conditions were as described in the text. 2 The values of MAP prior to LPS treatment. 3 n, the number of animals in each group. 4 [(MGD)2/Fe] is defined as the ratio of [(MGD)2/Fe] to be 2 to 0.4.

The MAP of anaesthetised rats was in the range of 96 to 99mmHg. Two hours after LPS treatment, the MAP decreased to between 71 and 77mmHg, which is indicative of the onset of systemic hypotension, caused by abnormally elevated levels of nitric oxide, as also shown in Figure 1.

1i While the 2.0h saline infusion did not change the MAP, infusions of [(MGD)2/Fe] complex at various ratios, ranging from 2 to 0.4 (MGD to Fe) to 2 to 0 (MGD to Fe), restored the blood pressure to 89- 95mmHg (Table These results suggest that the i.v. infusion of MGD either with or without added iron can restore blood pressure in hypotensive rats induced by LPS challenge (Table 1).

Since MGD does not bind -NO, it is speculated that the restoration of the MAP by MGD infusion S 20 may be attributed to the MGD chelation of cellular iron released by excess -NO production, which is known to attack cellular iron-containing proteins and result in cellular iron loss during sepsis or septic shock (see, for example, Kim et al., in J. Biol. Chem. 270:5710-5713 (1995)).

This example shows that dithiocarbamate containing nitric oxide scavengers, such as MGD, either with or without added iron, are effective for the treatment of systemic hypotension, a condition which is associated with many inflammatory and/or infectious diseases.

Example 4 In order to test the efficacy of the combinational therapy of [(MGD)2/Fe] and anti-TNF antibody for treatment of LPS-induced shock, Wistar rats are anaesthetised with Ketamine/Xylazine plus 5.5mg/kg). A catheter is implanted in the femoral vein for drug administration. The femoral artery is cannulated for continuous blood pressure measurement. The animals are allowed to recover from surgery for a period of 3 days prior to experimentation. On the day of the experiment, the conscious rats are retained in restrainers and the artery line is connected to the pressure transducer for recording. Rats are injected with an i.v. bolus dose of LPS typhosa, endotoxin, 10-20mg/kg). Two hours after LPS challenge, rats are then subjected to one of the following treatments (8 animals in each group): Control, saline infusion 1.OmL saline/h of saline infusion for 6h.

Libc/03675 26

(MGD)

2 /Fe] (at a ratio of 5 to 1) 0.1 mmole/kg/h infusion for 3h, followed by saline infusion for 3h.

Anti-TNF- 7.5mg/kg/h infusion for 3h, followed by saline infusion for 3h.

Co-infusion of [(MGD) 2 /Fe] (0.1mmole/kg/h) and Anti-TNF (7.5mg/kg/h) for 3h, followed by saline infusion for 3h.

s (MGD)2/Fe] (at a ratio of 5 to 1) -0.1mmole/kg/h infusion for 3h and followed by anti-TNF infusion for 3h.

At the end of the infusion, rats are returned to their cages for observation. The 24h survival rates resulting from these various treatments are compared. Since a lethal dose of LPS is used, it is expected that all animals in control group 1 will die within 24h. Based on the results presented in Figure 1 (Example it is expected that about two thirds of the rats in the treatment group group 2, treated with [(MGD)2/Fe]) will survive after 24h. As discussed above, in endotoxemia,

TNF

production is short-lived and peaks at 1-2h. Therefore, the infusion of anti-TNF antibodies at two hours after LPS challenge as indicated in group 3 may not be able to block the induction of the inducible nitric oxide synthase gene, which results in the production of iNOS, resulting in the overproduction of nitric oxide. In group 4, the co-infusion of anti-TNF antibodies and [(MGD) 2 /Fe] is expected to produce a similar survival rate as that for group 2, employing [(MGD)2/Fe] infusion alone.

On the other hand, it is expected that the infusion of [(MGD)2/Fe] for 3 hours, followed by the infusion of anti-TNF antibodies (as done with group 5) will improve the survival rate over that in group 2, because the infusion of anti-TNF antibodies at later hours would inhibit further activation of the inducible NO synthase gene, thereby reducing the further enhancement of excessive NO production.

:i The efficacies of combinational therapy between [(MGD)2/Fe] and other therapeutic agents (such as antiendotoxin antibodies, other anti-cytokine antibodies, anticytokine receptor antibodies, and other agents, such as antibradykinin peptides, nitric oxide synthase inhibitors, and the like) can be demonstrated in a similar fashion to that described herein.

Example NO production has been shown to be increased during acute cardiac allograft rejection in rats, as evidenced by elevated urinary and plasma nitrate/nitrite levels, preceding and at the time of rejection (see, for example, Winlaw et al., in Transplantation 58:1031 (1994)). Abnormally elevated NO levels appeared to be produced by activated infiltrating host macrophages and cardiac myocytes 30 of the rejecting allograft (see, for example, Yang et al., in J. Clin. Invest. 94:714-721 (1994)).

Cyclosporin is widely used as an immunosuppressive agent to prevent allograft rejection, mainly through the inhibition of T cell activation. However, the use of cyclosporin has been associated with multiple side effects, such as, for example, nephrotoxicity, hepatotoxicity and hypertension (see, for example, Atkinson et al., in Transplantation 38:34 (1984).

Experiments were performed to evaluate the effectiveness of invention combination therapy in preventing cardiac allograft rejection in rats, employing low doses of cyclosporin and MGD/Fe. Organ donors were male Wistar-Furth (WF) strain rats weighing ~160-300g. Organ recipients were male Lewis (Lew) strain rats weighing ~210-340g. Lewis rats underwent either syngeneic Lew-Lew) or allogeneic WF-Lew) heterotropic cardiac transplantation to the abdominal aorta and vena cava by Libc/03675 27 standard microvascular surgical techniques using sodium pentobarbital anaesthesia (50mg/kg). All cardiac transplants were observed to have good contractile function, and there were no early deaths from surgical complications. Graft function was monitored by palpation through the abdominal wall twice daily. Allograft rejection was defined as the loss of palpable contractile activity, and was confirmed by direct inspection at laparotomy.

The MGD/Fe complex was prepared fresh daily by dissolving MGD in distilled water and then adding an appropriate amount of aqueous FeSO4 to create a solution with a molar ratio of MGD/Fe of -10:1. A sufficient volume of the MGD/Fe solution was prepared to allow subcutaneous injection. A stock solution of Cyclosporin A (CsA, Sandoz Pharma Ltd.) was prepared in commercially available olive oil. Animals received 2.5mg/kg intramuscularly on postoperative days one to seven. This suboptimal dose ("lowdose") of CsA was used to achieve prolongation of allograft survival, without the indefinite survival which typically results when a full dose of CsA ~10-15mg/kg/day) is administered to the rat.

Lewis rat strain recipients received a Wistar Furth allograft and one of the following treatments: single therapy with MGD-Fe (400mg/kg, sc, bid) until rejection, or low-dose cyclosporin A (CsA, 2.5mg/kg im) for seven days, or combination therapy with CsA (at the same low-dose level as used in for seven days and MGD-Fe (at the same dose level as used in for 30 days.

Body weight was used as an index of overall animal health during the study period. There was no difference in body weights at the beginning of the study, relative to the termination of the study in any of the study groups (see Table Results are expressed in the Table as mean SEM.

Table 2 SGroup Bod Weiht, g Group Preoperative Rejection 1 (Isograft) 5 289±11 305±7 2 (Allograft, No treatment) 14 264±9 251±6 3 (Allograft, MGD-Fe) 17 243±5 234+5 4 (Allograft, CsA) 18 268±4 276±3 5 (Allograft, CsA MGD-Fe) 11 267±7 297±8 Body weight taken at day 30 since the isograft does not undergo rejection.

In all of the study groups, there was a decrease in body weight of 8-14% by day 7 after 25 transplant. All groups, however, displayed a weight gain after day seven, suggesting that the initial weight loss during the study period was due to the effect of surgery upon the rats, rather than the form of treatment to which the rats were subjected. Isograft controls which received no treatment exhibited similar weight trends as compared to the various treatment groups. The increase in body weight observed in the group subjected to invention combination therapy is likely the result of increased survival in the study due to improved graft survival.

Graft survival is reported herein as the mean survival time (MST±SE) in days (see Table 3).

Libc/03675 1 28 Table 3 roup Donor Recp't n Treatment Graft survival, days (n)t MSTtt ±SEM, days LEW LEW 5 None >100

NA

WF LEW 17 None 6.5(11), 6.9±0.2 (1) 3 WF LEW 16 MGD-Fe, 400mg/kg sc bid, 11.8±0.4 until rejection 10.0 10.5 11.0(1), 11.5 12.5 13.0 13.5 14.0(1), 4 WF LEW 17 CsA, 2.5mg/kg im daily x 7 11.0 14.5+1.1 days 11.5(3), 12.0(2), 12.5(5) 13.0 19.5 (1) 22.0 (1) 23.0 (1 24.0 (1) WF LEW 11 CsA, 2.5mg/kg im daily x 7 31.0(1), 45.0±4.7 days plus MGD-Fe, 400 32.5(2), mg/kg sc bid daily x 30 days 37.0 38.0 57.0 44.5 42.0 86.0 51.5 43.0 (1) 1 number of rats tt MST mean survival time (in days) graft still functioning sacrificed due to paracardial abscess, graft still functioning Acute allograft rejection occurred in 6.9+0.2 days in untreated controls. Single drug therapy with either MGD-Fe or CsA alone significantly prolonged allograft survival, compared to untreated allografts (11.8±0.4 and 14.5±1.1 days, respectively). Combination drug therapy according to the invention, however, resulted in a dramatic prolongation 5 of graft survival 45.0±4.7 days).

MGD-Fe therapy was discontinued on day 30 posttransplant to determine whether indefinite survival >100 days) was achievable. Graft function continued beyond 30 days in all 11 rats in the group receiving invention combination therapy (see Table One of the rats in this group has a functioning allograft more than 50 days following cessation of combination therapy. All animals appeared healthy throughout the duration of the therapy, and there were no deaths.

In conclusion, modulation of nitric oxide levels, especially in combination with subtherapeutic doses of standard immunosuppressive therapy, results in a dramatic prolongation of allograft survival.

Libc/03675 29 c' While the invention has been described in detail with reference to certain preferred embodiments thereof, it will be understood that modifications and variations are within the spirit and scope of that which is described and claimed.

o *o Libc/03675

Claims (36)

1. A method for directly or indirectly treating the production of species which induce the expression of inducible nitric oxide synthase in a subject, said method comprising: co-administering to said subject an effective amount of a combination of at least one agent capable of directly or indirectly inactivating said species, or inhibiting production of said species, and at least one nitric oxide scavenger.

2. A combination of at least one agent capable of directly or indirectly inactivating species which induce the expression of nitric oxide synthase in a subject, or inhibiting production of said species, and at least one nitric oxide scavenger when used in directly or indirectly treating the production of species which induce the expression of inducible nitric oxide synthase.

3. The use of a combination of at least one agent capable of directly or indirectly inactivating species which induce the expression of nitric oxide synthase in a subject, or inhibiting production of said species, and at least one nitric oxide scavenger for the manufacture of a medicament for directly or indirectly treating the production of species which induce the expression of inducible nitric oxide synthase. S4. A method, combination or use according to any one of claims 1 to 3, wherein said species is selected from cytokines, cytokine receptors, endotoxins, platelet activating factor, bradykinin, bradykinin receptor, bacteria, coagulation factors, arachidonate metabolites or nitric oxide synthase.

5. A method, combination or use according to any one of claims 1 to 3, wherein said agent is selected from anti-endotoxin agents, inhibitors of cytokine synthesis/release, anti-cytokine agents, inhibitors of the coagulation cascade, inhibitors of complement activation, inhibitors of platelet activating factor, inhibitors of arachidonate metabolism, inhibitors of nitric oxide synthase enzymes, 5 therapeutic agents for inflammatory diseases or therapeutic agents for Crohn's disease therapy. 25 6. A method, combination or use according to any one of claims 1 to 3, wherein said agent is selected from anti-endotoxin agents, anti-cytokine agents, inhibitors of nitric oxide synthase enzymes, immunosuppressive agents or therapeutic agents for inflammatory diseases.

7. A method, combination or use according to any one of claims 1 to 3, wherein said nitric oxide scavenger is selected from the group consisting of non-heme iron-containing peptides or proteins, porphyrins, metalloporphyrins, dithiocarbamates, dimercaptosuccinic acid, phenanthroline, desferrioxamine, pyridoxal isonicotinoyl hydrazone (PIH), 1,2-dimethyl-3-hydroxypyrid-4-one (L1) and [+]-1,2-bis(3,5-dioxopiperazine-1-yl)propane (ICRF-187).

8. A method, combination or use according to any one of claims 1 to 3, wherein said nitric oxide scavenger comprises a dithiocarbamate moiety having the structure optionally associated with a physiologically compatible di- or trivalent transition metal ion, wherein structure is as follows: [R1R2N-C(S)-S]-xM+1,+2,+3 (I) wherein: each of R 1 and R 2 is independently selected from a C1 up to C18 alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, alkylaryl, f 40 a'sbstituted alkylaryl, arylalkyl, substituted arylalkyl, or R 1 and R 2 can cooperate to form a 6- or 7- Libc/03675 l p dv membered ring including N, R 1 and R 2 x is 1 or 2, and M is a monovalent cation when x is 1, or M is a physiologically compatible divalent or trivalent transition metal cation when x is 2.

9. A method, combination or use according to claim 8 wherein the ratio of transition metal ion to dithiocarbamate moiety falls in the range of zero up to about 1:2.

10. A method, combination or use according to claim 8 wherein said physiologically compatible di- or trivalent transition metal is selected from iron, cobalt, copper or manganese.

11. A method, combination or use according to any one of claims 1 to 3, wherein said combination of at least one agent, and at least one nitric oxide scavenger is delivered orally, intravenously, subcutaneously, parenterally, rectally or by inhalation.

12. A method, combination or use according to any one of claims 1 to 3, wherein said combination of at least one agent, and at least one nitric oxide scavenger is delivered in the form of a solid, solution, emulsion, dispersion, micelle or liposome.

13. A therapeutic method which employs an agent to inactivate materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase, comprising co-administering to a patient in need thereof a nitric oxide scavenger in combination with said agent.

14. A combination of an agent to inactivate materials which, directly or indirectly, induce the *ha: expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger when used in therapy. The use of a combination of an agent to inactivate materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger for the manufacture of a medicament for therapy.

16. A method, combination or use according to any one of claims 12 to 15, wherein said agent is selected from anti-endotoxin agents, inhibitors of cytokine synthesis/release, anti-cytokine S agents, inhibitors of the coagulation cascade, inhibitors of complement activation, inhibitors of platelet 25 activating factor, inhibitors of arachidonate metabolism, inhibitors of nitric oxide synthase enzymes, immunosuppressive agents, diabetic therapeutic agents, therapeutic agents for inflammatory diseases or therapeutic agents for Crohn's disease therapy. S: 17. A therapeutic method which employs a therapeutic agent which, directly or indirectly, induces the expression of inducible nitric oxide synthase, comprising co-administering to a patient in need thereof a nitric oxide scavenger in combination with said therapeutic agent.

18. A combination of a therapeutic agent which, directly or indirectly, induces the expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger when used in therapy.

19. The use of combination of a therapeutic agent which, directly or indirectly, induces the expression of inducible nitric oxide synthase, and at least one nitric oxide scavenger for the manufacture of a medicament for therapy. A method, combination or use according to any one of claims 17 to 19, wherein said therapeutic process is selected from administration of immunosuppressants, administration of antimetabolite cytotoxics, administration of neuroprotective agents, administration of T cell inhibitors, administration of antimigraine agents, administration of antiarthritic agents, administration of agents useful for the treatment of septic shock, administration of agents for the treatment of multiple Libc/03675 -32- sclerosis, administration of organ transplantation agents, administration of agents for the treatment of systemic lupus erythematosus (SLE), administration of agents for the treatment of Alzheimer's disease, administration of agents for the treatment of psoriasis, administration of agents for the treatment of diabetes, administration of agents for the treatment of stroke.

21. A composition comprising a combination of an agent capable of inactivating materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase and a nitric oxide scavenger in a pharmaceutically acceptable carrier therefor.

22. A composition according to claim 21 wherein said nitric oxide scavenger is a compound having structure wherein said compound having structure is as follows: [RiR2N-C(S)-S]-xM- 2 3 (I) wherein: each of R 1 and R 2 is independently selected from a C1 up to Ci8 alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, alkylaryl, substituted alkylaryl, arylalkyl, substituted arylalkyl or R 1 and R 2 can cooperate to form a 6- or 7- membered ring including N, R 1 and R 2 x is 1 or 2, and M is a monovalent cation when x is 1, or M is a physiologically compatible divalent or trivalent transition metal cation when x is 2. .oi 23. A composition according to claim 22 wherein M is selected from H Na NH4+ or tetraalkyl ammonium. S24 A composition according to claim 22 wherein M is selected from Fe+ 2 Fe+ 3 Co+ 2 Co+ 3 20 CU 2 Mn 2 or Mn+3. A composition according to claim 22 wherein the ratio of transition metal ion to dithiocarbamate moiety falls in the range of zero up to about 1:2. oo. 26. A composition according to claim 22 wherein: each of R 1 and R 2 a C1 up to 012 alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl or substituted alkynyl, wherein the substituents 25 are selected from carboxyl, oxyacyl, phenol, phenoxy, pyridinyl, pyrrolidinyl, amino, amido, hydroxy, nitro or sulfuryl, and M Fe 2 or Fe+ 3

27. A composition according to claim 22 wherein: R 1 a C2 up to C8 alkyl or substituted alkyl, wherein said substituents are selected from carboxyl, acetyl, pyridinyl, pyrrolidinyl, amino, amido, hydroxy or nitro, R 2 is selected from a C1 up to C6 alkyl or substituted alkyl, or R 2 can cooperate with R 1 to form a 6- or 7-membered ring including N, R 2 and R 1 and M Fe 2

28. A composition according to claim 22 wherein: R 1 a C2 up to C8 alkyl or substituted alkyl, wherein said substituents are selected from carboxyl, acetyl, amido or hydroxy, R 2 a Ci up to C4 alkyl or substituted alkyl and M Fe 2

29. A composition according to claim 22 wherein said agent is selected from anti-endotoxin agents, inhibitors of cytokine synthesis/release, anti-cytokine agents, inhibitors of the coagulation cascade, inhibitors of complement activation, inhibitors of platelet activating factor, inhibitors of arachidonate metabolism, inhibitors of nitric oxide synthase enzymes, immunosuppressive agents, diabetic therapeutic agents, therapeutic agents for inflammatory diseases or therapeutic agents for Crohn's disease therapy, anti-cytokine antibodies, anti-cytokine receptor antibodies, anti-endotoxin i b40 >tibodies, bradykinin antagonists, synthetic peptide blocking bradykinin receptors, Libc03675 S-33- bactericidal/permeability increasing protein, antibodies to platelet activating factor, or therapeutic agents for treatment of ophthalmic diseases. A composition according to claim 29 wherein said anti-endotoxin agent is selected from antibodies to endotoxin, antibodies to LPS-binding protein, soluble CD14 protein, bactericidal/permeability increasing protein or polymyxin B.

31. A composition according to claim 29 wherein said inhibitor of cytokine synthesis/release is selected from phosphodiesterase inhibitors, IL-4, IL-10, IL-13, TGF-S, aspirin, phenyl butyl nitrone or corticosteroids.

32. A composition according to claim 29 wherein said anti-cytokine agent is selected from antibodies to TNF, soluble TNF receptors, IL-1 receptor antagonists, antibodies to IL-1 receptors, antibodies to IL-6, antibodies to interferon-y or soluble interferon-y receptors.

33. A composition according to claim 29 wherein said inhibitor of the coagulation cascade is selected from anti-Factor XII antibodies, antibodies to C5a, C1-esterase inhibitors or soluble Crl.

34. A composition according to claim 29 wherein said inhibitor of platelet activating factor is a PAF receptor antagonist.

35. A composition according to claim 29 wherein said inhibitor of arachidonate metabolism is *ii selected from cyclooxygenase inhibitors, lipoxygenase inhibitors, leukotriene inhibitors, thromboxane A 2 inhibitors, or prostaglandins.

36. A composition according to claim 29 wherein said inhibitor of nitric oxide synthase 20 enzymes is selected from N-methyl-L-arginine, e-N-iminoethyl-L-lysine, aminoguanidine or S-methyl isothiourea sulfate.

37. A composition according to claim 29 wherein said immunosuppressive agent is selected from cyclosporin, OKT3, FK506, thymoglobulin or mycophenolic acid.

38. A composition according to claim 29 wherein said diabetic therapeutic agent is selected 25 from free pancreatic islets, encapsulated pancreatic islets, oral insulin, intravenous insulin, or amylin hormone.

39. A composition according to claim 29 wherein said therapeutic agent for inflammatory disease is selected from sulfasalazine, mesalamine, corticosteroids, azathioprine, 6-mercaptopurine, or metronidazole.

40. A composition according to claim 29 wherein said therapeutic agent for inflammatory disease is a dihydropyridine calcium channel blocker.

41. A composition according to claim 29 wherein said agent is selected from anti-endotoxin agents, inhibitors of cytokine synthesis/release, anti-cytokine agents, inhibitors of the coagulation cascade, inhibitors of complement activation, inhibitors of platelet activating factor, inhibitors of arachidonate metabolism, inhibitors of nitric oxide synthase enzymes, immunosuppressive agents, diabetic therapeutic agents, therapeutic agents for inflammatory diseases or therapeutic agents for Crohn's disease therapy, anti-cytokine antibodies, anti-cytokine receptor antibodies, anti-endotoxin antibodies, bradykinin antagonists, synthetic peptide blocking bradykinin receptors, bactericidal/permeability increasing protein or antibodies to platelet activating factor. Libc/03675 -34-

42. A composition according to claim 21 wherein said pharmaceutically acceptable carrier is selected from a solid, solution, emulsion, dispersion, micelle or liposome.

43. A composition according to claim 21 wherein said composition further comprises an enteric coating.

44. A composition according to claim 21 wherein said therapeutic agent for ophthalmic disease is a topical corticosteroid, an immunosuppressive agent, an antibiotic, azathioprine, ceftriaxone, drop preparations, artificial tears, topical lodoxamide, acetazolamide, pilocarpine, timolal, levobunolal, metipranolol, ganciclovir, fascarnet, methylprednisolone, prednisolone, cyclopentolate, salicylate, indomethacin, phenybutazone or dexamethazone.

45. A composition comprising a combination of an agent capable of inactivating materials which, directly or indirectly, induce the expression of inducible nitric oxide synthase and a nitric oxide scavenger, substantially as hereinbefore described with reference to any one of the examples.

46. A method for directly or indirectly treating the production of species which induce the expression of inducible nitric oxide synthase in a subject, substantially as hereinbefore described with reference to any one of the examples. **MEDINOX, INC. Patent Attorneys for the Applicant/Nominated Person 20 SPRUSON&FERGUSON 2 *o*o4 o* e g**;Ptn tony o heApiatNmntdPro :2 SR SO&oR UO Lbc/03675