WO1999012547A1

WO1999012547A1 - Treatment of inflammatory conditions

Info

Publication number: WO1999012547A1
Application number: PCT/GB1998/002660
Authority: WO
Inventors: David Blake; Declan Naughton; Ian Stratford; Ged Adams
Original assignee: Theramark Limited; Adams, Margaret
Priority date: 1997-09-08
Filing date: 1998-09-08
Publication date: 1999-03-18
Also published as: AU9082798A; AU9082698A; WO1999012548A1

Abstract

Treatment of arthritis, perodontitis or Crohn's disease is effected by use of a bioreductive compound, e.g, a 2-nitroimidazole.

Description

TREATMENT OF INFLAMMATORY CONDITIONS

The present invention relates to the treatment of inflammatory conditions. In particular, the invention relates to a new medical use for bioreductive compounds in the treatment of rheumatoid arthritis, Crohn's disease and peridontitis where hypoxia and/or ischemia are implicated.

Rheumatoid arthritis is a common chronic systemic inflammatory disease which predominantly affects the synovial joints. Early rheumatoid arthritis is characterised primarily by inflammation of the synovium. However, as the disease progresses the patient may suffer destruction of cartilage and bone. As yet, there is no curative treatment for rheumatoid arthritis and management of the condition is aimed at alleviating pain and improving or maintaining joint function. This may be accomplished through physiotherapy as well as the use of drugs.

Many different drugs have been suggested for use in treating rheumatoid arthritis. In mild cases an analgesic alone may be all that is required. However, most patients require the additional anti-inflammatory effect provided by a non-steroidal anti-inflammatory drug (NSAID). Although NSAIDs provide symptomatic relief they do not suppress the rate of cartilage erosion or alter the course of the disease and additional treatment is often necessary to try and achieve this. Drugs used in this way are known as second-line agents and include the antimalarials, sulphasalazine, penicillamine and cyclosporin.

Corticosteroids such as dexamethasone have also been suggested for use in the treatment of rheumatoid arthritis. Although systemic corticosteroids can suppress the symptoms of the disease, their clinical use is limited by adverse side effects.

There thus exists a need for any new treatment which may help to relieve or improve symptoms associated with rheumatoid arthritis, in particular which may provide a cure for the disease. As long ago as 1982, metronidazole was proposed for use in treating rheumatoid arthritis (S. Afr. Med. Journal, 1 May 1982, pages 648-649). (Metronidazole is a member of the 5-nitroimidazole group of drugs with widespread clinical use as an antibiotic and antiparasitic agent). Administration of high doses of metronidazole to patients with active rheumatoid disease was found to cause an initial exacerbation of the inflammatory condition, even in previously unaffected tissues. This was also accompanied by influenza-like symptoms such as pyrexia, profuse sweating and headache (Herxheimer reaction). However, it was suggested that after a period of from several weeks to several months the initial tissue reaction died down and the clinical manifestations of the disease gradually disappeared. The suggestion was that metronidazole was responsible for killing a pathogenic free-living bacterium present in the rheumatoid lesions. However, no clinical or laboratory data were published to substantiate these findings.

A much more recent double blind study carried out by The Centre for Rheumatic Diseases at Glasgow Royal Infirmary in the UK failed to identify a useful role for metronidazole in the treatment of rheumatoid arthritis (Annals of the Rheumatic Diseases 5 .: 758-760, 1992). In this study, metronidazole did not have any disease modifying properties and was found to be unacceptably toxic. We now believe that the results of this later study are in error.

US-A-4 218 449 discloses the use of 4- and 5-nitroimidazoles in the treatment of rheumatoid arthritis and related collagen and auto-immune (rheumatoid) diseases and suggests that it is the antiprotozoal activity of the compounds which render them useful for treating these conditions.

Whilst it is known that rheumatoid arthritis is associated with chronic synovial inflammation and poor perfusion of the synovial tissues, we have now surprisingly found that in patients suffering from rheumatoid arthritis the synovial tissues are in many cases profoundly hypoxic (pO₂ < 12 mm Hg) and contain high levels of reductases. Whilst not wishing to be bound by theoretical considerations, it is believed that in patients suffering from rheumatoid arthritis there are pockets of cells in the synovium which are hypoxic and that it is these cells which are primarily responsible for the inflammation associated with the disease.

Hypoxic tissues are also believed to be present in chronic peridontitis, a condition associated with sever inflammation of the peridontium. Examples of prior treatments for peridontitis include administration metronidazole (a 4-nitroimidazole) (see for example J. Clin. Periodontol., vol 18, pages 177-181 (1991).

A further condition in which hypoxic tissue is present in Crohn's disease. Examples of prior treatments of this condition have included use of metronidazole (a 4-nitromidazole) (see for example Postgrad. Med., vol 74, pages 155-157 (1983)) and ornidazole (a 5-nitroimidazole) (see for example Schweiz. Med. Wochenschr., vol. 108, pages 1075-1077, and Am. J. Gastroenterol, vol. 83, pages 892-893 (1988).

The latter article states that the authors do not have information concerning the possible mechanisms of action of the drug but propose that action against bacteria may be a possible mechanism.

WO-A-98 35701 (Theramark Ltd) disclose conjugates for treatment of hypoxic conditions and comprising a bioreductive carrier moiety having linked thereto at least one therapeutic agent such that at a hypoxic site the carrier undergoes bioreduction with elimination of the therapeutic agent.

We now propose a new treatment for those suffering from arthritis, in particular rheumatoid arthritis, peridontitis and Crohn's disease which exploits the presence of hypoxia in the tissues. Thus, it has now been found that bioreductive compounds capable of killing hypoxic cells are particularly effective in relieving the symptoms of inflammatory conditions associated with hypoxia and/or ischemia and accordingly are of benefit in the treatment of arthritic conditions, e.g. rheumatoid arthritis, peridontitis and Crohn's disease. Thus, viewed from one aspect the invention provides the use of a bioreductive compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of arthritis, Crohn's disease or peridontitis with the proviso that the bioreductive compound is other than a 4- or 5-nitroimidazole and is other than a conjugate comprising a bioreductive carrier moiety having at least one therapeutic agent linked thereto.

The invention is useful in the treatment of rheumatoid or osteoarthritis, especially rheumatoid arthritis.

In another aspect the invention provides a method of treatment of arthritis, e.g. rheumatoid arthritis, peridontitis or Crohn's disease in the human or non-human animal body comprising administering to said body a bioreductive compound, or a pharmaceutically acceptable salt thereof with the proviso that the bioreductive compound is other than a 4- or 5-nitroimidazole and is other than a conjugate comprising a bioreductive carrier moiety having at least one therapeutic agent linked thereto.

Bioreductive compounds are a class of compound requiring metabolic reduction to generate cytotoxic metabolites. This process is facilitated by the presence of appropriate reductases and the lower oxygen conditions present in hypoxic or ischemic tissue compared with normal (normoxic) tissue. As a result of this specificity, a number of bioreductive compounds capable of producing cytotoxic metabolites under hypoxic conditions have been proposed for use in combination with radiotherapy treatment of tumors in which the presence of hypxoia has been demonstrated.

A large number of bioreductive compounds are known to act as potent alkylating agents after undergoing reduction in vivo. Examples of know bioreductive alkylating agents include compounds such as activated enamines, vinylogous quinone methides, simple quinone methides and α-methylene lactones or lactams. Bioactivation of such compounds produces species which are electron deficient and which are capable of covalent binding to a nucleophilic centre on a biomolecule, such as DNA.

Thus, bioreductive compounds act as a substrate for cellular reductases and are capable of reductive metabolism to generate a compound more toxic than its parent. Reductive metabolism means the addition of electrons to the compound with the addition of electrons effectively activating the compound. Most bioreductive compounds that have been developed for use in the treatment of tumors exhibit an optimum "trapping" potential when hypoxia is profound (pO₂ <12 mm Hg) and this is believed to form the basis for their selectivity for hypoxic as opposed to normal tissues.

Bioreductive compounds are known to be substantially stable in an oxygenated environment. However, in a hypoxic or ischemic environment, such compounds are converted into cytotoxic species. Whilst not wishing to be bound by theoretical considerations, we now believe that these species are effective in killing hypoxic cells considered to be responsible for the severe inflammation associated with arthritis, in particular rheumatoid arthritis.

As used herein, the term "bioreductive compound" is intended to define any molecule which is reduced in the presence of reducing enzymes or reductases to generate a cytotoxic metabolite. For example, a bioreductive compound may be any comparatively non-reactive compound which in the presence of reductases is converted into a more reactive, cytotoxic species. The common feature shared by the bioreductive compounds for use in the method of the invention is that enzyme- mediated reductive activation is a prerequisite for their toxicity.

The bioreductive compounds for use in the method of the invention are capable of targeting tissues having an enhanced reductase activity. This is believed to be a consequence of hypoxic metabolism and/or reduced oxygenation of such tissues.. Preferred bioreductive compounds for use in accordance with the invention are those which have the ability to penetrate poorly perfused tissues and which only undergo bioreduction in a hypoxic and/or ischemic environment.

A large number of bioreductive compounds of diverse structure are know to be effective in killing mammalian hypoxic cells and are described in the literature. Such compounds include the quinones, aromatic nitro compounds, N-oxides, activated enamines, vinylogous quinone methides, simple quinone methides and α-methylene lactones or lactams. All such compounds and their derivatives are considered to be within the scope of the present invention, as are their pharmaceutically acceptable salts, including both organic and inorganic salts, e.g. with alkali and alkaline earth metals, ammonium, ethanolamine. diethanolamine and meglumine, chloride, hydrogen carbonate, phosphate, sulphate and acetate counterions. Suitable pharmaceutically acceptable salts are well described in the pharmaceutical literature.

Preferred bioreductive compounds for use in the invention are compounds having a one-electron reduction potential of -250m V to -450mV, more preferably - 300mV to - 00mV, even more preferably -320mV to -380mV, e.g. about -350mV and include the quinones, aromatic nitro compounds and N-oxides.

The initial reductive step in the activation of aromatic nitro compounds is believed to be a one-electron process to give the nitro radical anion (RNO₂ ^'). This can be reoxidised back to the parent nitro compound by oxygen. However, under hypoxic conditions this can undergo further reduction to give cytotoxic metabolites such as the nitroso or hydroxylamine derivatives. These are capable of reacting with DNA and are thereby toxic.

Aromatic nitro compounds which may be used in the invention include such compounds having a nitro group bonded to, for example, a benzene, imidazole, pyrrole, triazole. tetrazole, thiazole, oxazole, benzothiazole, oxazole, benzoxazole, furan, thioquinol, thiophene or indole ring. Particularly preferred aromatic nitro compounds are nitroimidazoles and their derivatives, in particular the 2- nitroimidazoles. Preferred nitrimidazoles include benznidazole, etanidazole, misonidazole and flexnidazole. Of these, benznidazole is particularly preferred for use in the invention.

Aromatic N-oxides are another class of bioreductive compound selectively toxic to hypoxic cells. Such compounds require one-electron reduction to express toxicity. It is believed to be the N-oxide radical anion which causes toxicity by interaction directly with cellular macromolecules. It is thought that hydrogen is abstracted from bases in DNA giving rise to extensive single- and double-stranded breaks.

An example of an aromatic N-oxide preferred for use in the method of the invention is the benzo-triazene-di-N-oxide, tirapazamine.

Quinones are also an important class of bioreductive compound suitable for use in the method of the invention. This class of compounds include the mitomycins and their analogues. Agents such as mitomycin C (MMC) and E09 are rendered toxic following activation by one- or two-electron reductions and are particularly suitable for use in the invention. This class of compound also includes the napthoquinones, indoloquinones. quinolino quinones and their derivatives. The electron deficient quinone nucleus in such compounds readily undergoes reduction in vivo to form the corresponding electron rich hydroquinone which in turn is capable of functioning as an alkylating agent.

Reductases known to be involved in the activation of bioreductive compounds include DT diaphorase, cytochrome P450, NADPH-dependent cytochrome P450 reductase and xanthine oxidase. The ease of reduction of any given bioreductive compound will depend upon its ability to act as a substrate for the intracellular reducatses and the expression levels of such enzymes within the particular cell type. The choice of bioreductive compound for use in the invention will thus depend upon the type of enzymes present at the site of inflammation. Indeed, it may be useful to determine the relative enzyme activities in the inflamed tissues of individual patients before starting treatment.

The usefulness of hypoxia-directed bioreductive agents will thus be enhanced by identification of appropriate tissues for treatment. This may be based on a predictive assay for hypoxia and/or identifying the presence of enzymes at the site of inflammation which are important for the bioactivation of drugs or conversely their detoxification.

P450 reductase has been shown to be important in the one-electron reduction of 2-nitroimidazoles, e.g. benznidazole, to the nitro radical anion. For tirapazamine it is the one-electron reduced species which is toxic and this can be formed by both P450 reductase and by P450 itself. Where high levels of DT-diaphorase are present this may be exploited by the use of quinone-based bioreductives since these are known to be activated by this enzyme. Indoloquinones are particularly good substrates for DT diaphorase, an enzyme commonly found in most tissues. One bioreductive which has one of the greatest specificities for activation by DT- diaphorase is the indoloquinone E09.

In connection with the treatment of arthritic, the bioreductive compounds for use in the im'ention may be used in combination with other conventional treatment for arthritis, e.g. rheumatoid arthritis, such treatment being capable of targeting the surviving aerobic cell population. Thus, the bioreductive compounds may be used in combination with a second medicament effective in treating or in relieving the symptoms associated with arthritis, preferably a medicament effective in treating or relieving the symptom of rheumatoid arthritis. In this regard, the bioreductive compounds may conveniently be used in combination with a non-steroidal anti- inflammatory agent (NSAID), a corticosteroid or glucocorticoid, an alkylating agent, an antimalarial, a gold compound, penicillamine, sulphasalazine, methotrexate or azathioprine. The bioreductive may be administered as a combined preparation with the second medicament. Alternatively, the second medicament may be administered separately, prior to, during or subsequent to administration of the bioreductive compound.

Thus, viewed from a further aspect the invention provides a pharmaceutical composition comprising a bioreductive compound, or a pharmaceutically acceptable salt thereof., together with at least one compound effective in treating or in relieving the symptoms associated with arthritis, and at least one pharmaceutically acceptable carrier or excipient. Preferred bioreductives for use in accordance with this aspect of the invention include not only those outlined above, but also 4- and 5-nitroimidazoles, e.g. metronidazole. Metronidazole and nimorazole are especially preferred.

Viewed from a yet further aspect the invention provides a pack containing a bioreductive compound, or a pharmaceutically acceptable salt thereof, and separately at least one compound effective in treating or in relieving the symptoms associated with arthritis, for simultaneous, separate or sequential administration in the treatment of arthritis, e.g. rheumatoid arthritis.

Viewed from a yet still further aspect the invention provides the use of a bioreductive compound, or a pharmaceutically acceptable salt thereof, together with at least one compound effective in treating or in relieving the symptoms associated with arthritis, in the manufacture of medicaments for simultaneous, separate or sequential administration in the treatment of arthritis, e.g. rheumatoid arthritis.

In another aspect the invention provides a method of treatment of rheumatoid arthritis in the human or non-human animal body comprising administering to said body a bioreductive compound, or a pharmaceutically acceptable salt thereof, together with at least one compound effective in treating or in relieving the symptoms associated with arthritis, e.g. rheumatoid arthritis.

Examples of antimalarials suitable for use in this aspect of the invention include chloroquine and hydroxychloroquine. Examples of suitable gold compounds include auranofin and sodium aurothiomalate. Examples of suitable alkylating agents include chlorambucil and cyclophosphamide. Examples of glucocorticoids for use in this aspect of the invention include methylprednisolone, triamcinolone and hexacetonide. Corticosteroids suitable for use in combination with the bioreductive compound include dexamethsaone.

So far, we have focused primarily on eliminating the hypoxic cell fraction in a heterogeneous cell population using bioreductive compounds. However, there may also be some benefit in combining the use of bioreductive compounds with a treatment which renders all cells responsible for the inflammation hypoxic, i.e. which renders all cells susceptible to bioreductive drug activity.

Thus viewed from another aspect the invention provides the use of a bioreductive compound or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of inflammatory conditions arthritis, peridontitis and Crohn's disease in which hypoxia is artificially induced.

Hypoxia may be induced in inflammatory tissues by the use of vasoactive agents capable of manipulating blood flow to the site of inflammation. Drugs which alter the oxygen affinity of haemoglobin thereby decreasing oxygen availability, and methods of inducing damage to the vasculature of the inflamed tissues may also be used. Examples of agents suitable for inducing or enhancing the hypoxic status of the target cell of tissue include (i) inhibitors of nitric oxide synthease which serve to reduce blood supply and (ii) left shift agents, e.g. BW 589C, which serve to modify the oxygen affinity of haemoglobin and cause a left shift in the oxygen-haemoglobin dissociation curve.

The bioreductive compounds for use in the method of the invention are in general well-known and readily available. Alternatively, these may be synthesised in accordance with conventional synthesis techniques known to those skilled in the art. Techniques for the synthesis of quinones, in particular indoloquinones are described for example in J. Org. Chem. 50:4276-4281, 1985. Techniques for the synthesis of nitroimidazoles and their derivatives, in particular nimorazole, are described in US-A- 3399193. Techniques which may be used in the synthesis of other nitroimidazole derivatives are described in US-A-3458528.

The bioreductive compounds for use in the invention are preferably formulated prior to administration together with at least one pharmaceutically acceptable carrier, diluent or excipient.

The active ingredient in such compositions may comprise from about 0.1% to about 99% by weight of the formulation. By "pharmaceutically acceptable" is meant that the ingredient must be compatible with other ingredients of the compositions as well as physiologically acceptable to the patient.

Pharmaceutical compositions for use according to the present invention may be formulated in a conventional manner using readily available pharmaceutical or veterinary aids. Thus the active ingredient may be incorporated, optionally together with other active substances, with one or more conventional carriers, diluents and/or excipients. to produce conventional galenic preparations such as tablets, pills, powders, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, aerosols, soft and hard gelatin capsules, suppositories, sterile injectable solutions, sterile packaged powders, and the like.

Examples of suitable carriers, excipients, and diluents are lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water syrup, water, water/ethanol, water/glycol, water/polyethylene, glycol, propylene glycol, methyl cellulose, methylhydroxybenzoates, propyl hydroxybenzoates, talc, magnesium stearate, mineral oil or fatty substances such as hard fat or suitable mixtures thereof. The compositions may additionally include lubricating agents, wetting agents, emulsifying agents, suspending agents, preserving agents, sweetening agents, flavouring agents, and the like. The formulations may be formulated so as to provide quick, sustained or delayed release of the active ingredient after administration to the patient by use of procedures well known in the art. The compositions are preferably formulated in a unit dosage form, e.g. with each dosage containing from about 0.1 to about 500mg of the active ingredient.

The precise dosage of the active ingredient and the length of the treatment will depend upon a number of factors including the age and weight of the patient, the specific condition being treated and its severity, and the route of administration. In general, an effective dose will be of the order of from about 0.01 mg/kg to about 20 mg/kg bodyweight per day, e.g. from about 0.05 to about 10 mg/kg per day, administered one or more times daily. Thus, an appropriate dose for an adult may be from 10 to 100 mg per day, e.g. 20 to 50 mg per day.

Administration may be by any suitable method known in the art, including for example oral, parenteral (e.g. intramuscular, subcutaneous, intraperitoneal or intravenous), rectal or topical administration.

The present invention will now be further illustrated by way of the following non-limiting examples and accompanying drawings in which Fig. 1 and 2 illustrate the results of Example 1.

Example 1

A rat model of inflammation was used to investigate the effect of misonidazole in treating or preventing inflammation.

1. Ninety male Wistar rats weighting approximately 190 g were grouped into 18 cages of n = 5 and allowed to acclimatise for a week.

2. In each rat, an air pouch was induced by injecting subcutaneously 20ml of sterile air into the dorsum. The air pouches were reinflated after two days with another 20ml of air. 3. About 4 days later, when the air pouches had stabilised, they were challenged with 1ml of 2% carageenan solution to induce an inflammatory response. This was day 0 of the experiment.

4. Misonidazole at concentrations of lmg, 5mg, lOmg and 20mg was then administered for the only time, into the air pouch of the rats, on day 0. There were 20 rats assigned to each concentration of the drug (n=80).

5. After day 1 (i.e. 24 hours after the injection of drug ), 5 rats (1 cage) for each concentration of the drug (total 4 cages) were sacrificed. This was repeated for days 2, 3 and 6 until all the cages had been sacrificed.

6. Two cages (10 rats) were used as a control whereby the air pouch was administered with 2 mis of saline instead of the drugs. These rats were sacrificed at the same time points as above on days 1, 2, 3 and 6.

Following sacrifice exudate was aspirated from inside the pouch and its volume measured. The pouch was dissected out and weighed in toto. Tissue blocks were then cut randomly from the pouch wall and fixed either in formal saline and made into wax blocks or frozen in liquid nitrogen.

Thin sections were cut from both the wax blocks and frozen tissue and stained as below prior to quantitative assessment by microscopy with image analysis.

2. Histological staining of pouch wall tissue to quantify cellular responses

A standard staining technique (Haematoxylin and Eosin) differentiates cell nuclei and cytoplasm allowing assessment of changes in the structure of wax embedded tissue. 2.1. Procedure for staining with Hematoxylin and Eosin

dewax in histoclear x2 rehydrate in 100% Industrial Methylated Spirit (Ims) x2 rehydrate in 70% IMS place submerged in running tap water for 60 seconds place in Hematoxylin or 10 min place submerged in running tap water for 5 minutes place in eosin for 5 minutes place submerged in running tap water for 2 minutes rehydrate in 70% IMS rehydrate in 100% IMS x2 mount

2.2. Quantitative analysis of H & E Sections

A scoring system as disclosed by Ragno et al (Annals of the Rheumatic Diseases; 54 - pages 59-65) was used. Blind assessments of tissue sections were graded on a scale of 0-3 according to the presence and severity of the following parameters:

• Pouch proliferation

proliferation of the pouch membrane is diagnostic an inflammatory response

• Pyknotic nuclei

=> the presence of these nucleic staining intensely with haematoxylin indicate dead/dying cells

Because these data tend to follow an exponential distribution they were transformed to make them fit into a more normal distribution using the following formula, where x is the mean score for each section:

SUBSTΓΓUTE SHEET (RULE 26) X (V + ι)-ι

The transformed data were used to plot Fig 1 (pouch proliferation) and Fig 2 (pyknotic nuclei).

Table 1 below shows P values comparing proliferation in control tissue and with various doses of Misonidazole.

Table 1

• From the results in Fig 1 and Table 1. it can be seen that the control shows increasing pouch proliferation from days one to six.

• All doses of misonidazole reduce the levels of proliferation by day six (Fig. 1) though the values are not significant (Table 1).

• All misonidazole doses inhibit proliferation on day three, significantly so with 5mg (Table 1).

• 20mg misonidazole shows significant proliferation inhibition on day 2 (Table 1). The lowest (lmg) dose is also significantly inhibitory at this time point and the 5 mg dose nearly so. Table 2 below shows P values comparing pyknotic nuclei in control tissue and following various doses of Misonidazole.

• From Fig 2 and Table 2, it can be seen that there are few damaged cells in the control.

• All concentrations of misonidazole induce an increase in the number of pyknotic nuclei (Fig. 4). This is significant for all doses on day 1 and, in particular, day 2, the time at which the pouch is most hypoxic.

3. Conclusions

1. Misonidazole is able to target and kill hypoxic cells during the inflammatory response, as indicated by significant increases in pyknotic index.

2. The bioreductive drug significantly inhibits proliferation of the pouch. Misonidazole is most effective on days two and three when the pouch is hypoxic.

Example 2

A tablet is prepared using the following ingredients: Quantity

(mg per tablet)

Nimorazole 60

(available from Pharmacia Upjohn)

Starch 45

Microcrystalline cellulose 35

Polyvinylpyrrolidone

(as 10% solution in water) 4

Sodium carboxymethyl starch 4.5

Magnesium stearate 0.5

Talc 1

Total 150mg

The active ingredient, starch and cellulose are sieved and mixed thoroughly. The aqueous solution containing polyvinylpyrrolidone is then mixed with the resultant powder, and the mixture is then passed through a sieve to form a granulate. The remaining ingredients are sieved and then added to the granulate. After mixing these are compressed on a tablet machine to yield tablets each containing 60mg active ingredient.

Example 3

Capsules are prepared using the following ingredients:

Quantity

(mg per capsule)

Nimorazole 250

(available from Pharmacia Upjohn)

Starch, dried 200

Magnesium stearate 10

Total 460mg

The above ingredients are mixed and filled into hard gelatin capsules in 460mg quantities.

Claims

1. The use of a bioreductive compound, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for use in the treatment of arthritis, Crohn's disease or peridontitis with the proviso that the bioreductive compound is other than a 4- or 5-nitroimidazole and is other than a conjugate comprising a bioreductive carrier moiety having at least one therapeutic agent linked thereto.

2. Use as claimed in claim 1 in the treatment of rheumatoid or osteoarthritis.

3. Use as claimed in claim 1 or 2 wherein the bioreductive compound is capable of targeting tissues having an enhanced reductase activity.

4. Use as claimed in any preceding claim wherein the bioreductive compound is a quinone, aromatic nitro compound, N-oxide, activated enamine, vinylogous quinone methide, simple quinone methide, ╬▒-methylene lactone, lactam, or pharmaceutically acceptable salt thereof.

5. Use as claimed in claim 6 wherein the aromatic nitro compound is a nitroimidazole or derivative thereof.

6. Use as claimed in claim 5 wherein the aromatic nitro compound is a 2- nitroimidazole.

7. Use as claimed in claim 4 wherein the nitroimidazole is metronidazole, benznidazole, tinidazole, ornidazole, misonidazole, secnidazole, carnidazole, nimorazole, flexnidazole, panidazole or satranidazole.

8. Use as claimed in claim 4 wherein the aromatic N-oxide is tirapazamine.

9. Use as claimed in claim 4 wherein the quinone is a mitomycin or mitomycin analogue, a napthoquinone, an indoloquinone, a quinolino quinone or derivative thereof.

10. Use as claimed in claim 9 wherein the quinone is mitomycin C (MMC) or E09.

11. A method of treatment of arthritis, Crohn's disease or peridontitis in the human or non-human animal body, said method comprising administering to said body an effective amount of a bioreductive compound, or a pharmaceutically acceptable salt thereof, with the proviso that the bioreductive compound is other than a 4- or 5-nitroimidazole and is other than a conjugate comprising a bioreductive carrier moiety having at least one therapeutic agent linked thereto.

12. A pharmaceutical composition comprising a bioreductive compound, or a pharmaceutically acceptable salt thereof, together with a second medicament effective in treating or relieving the symptoms associated with an inflammatory condition, and at least one pharmaceutically acceptable carrier or excipient.

13. A pack containing a bioreductive compound, or a pharmaceutically acceptable salt thereof, and separately a second medicament effective in treating or relieving the symptoms associated with an arthritis, Crohn's disease or peridontitis for simultaneous, separate or sequential administration.

14. Use of a bioreductive compound, or a pharmaceutically acceptable salt thereof, together with a second medicament effective in treating or relieving the symptoms associated with an inflammatory condition, in the manufacture of medicaments for simultaneous, separate or sequential administration in the treatment of arthritis, Crohn^" s disease or peridontitis.

15. A composition, pack or use as claimed in any one of claims 12 to 14 wherein the bioreductive compound is as defined in any one of claims 1 to 10.

16. A composition, pack or use as claimed in any one of claims 12 to 15 wherein the second medicament is a non-steroidal anti-inflammatory agent (NSAID), a corticosteroid or glucocorticoid, an alkylating agent, an antimalarial, a gold compound, penicillamine, sulphasalazine, methotrexate or azathioprine.