WO2003024460A1

WO2003024460A1 - Use of 7-alpha hydroxy dhea and 7-oxo dhea for treating inflammatory or functional intestinal diseases

Info

Publication number: WO2003024460A1
Application number: PCT/FR2002/003109
Authority: WO
Inventors: Michel Seman; Marc Criton
Original assignee: Laboratoires Mayoly Spindler
Priority date: 2001-09-14
Filing date: 2002-09-12
Publication date: 2003-03-27
Also published as: FR2829697A1; FR2829697B1

Abstract

The invention concerns the use of 7-hydroxylated and 7-ketonic derivatives of 3-β-hydroxylated steroid hormones for preparing a medicine for treating inflammatory or functional intestinal diseases, such as mucus colitis, ulcerative colitis and Crohn's disease.

Description

USE OF 7-ALPHA HYDROXY DHEA AND 7-OXO-DHEA FOR THE TREATMENT OF INFLAMMATORY OR FUNCTIONAL DISEASES OF THE BOWEL

The present invention relates to the use of 7-hydroxylated 5 and 7-ketone derivatives of the 3E-hydroxylated steroid hormones for the preparation of a medicament intended for treating inflammatory or functional diseases of the intestine.

Inflammatory bowel diseases, including Crohn's disease and ulcerative colitis, are pathologies of unknown etiology, the incidence of which in France and in many countries of the world is constantly increasing. 10 As a general rule, these are chronic pathologies in which flare-ups of the inflammatory process, which sometimes have a real character of severity going as far as requiring hospitalization of the patient, alternate with more or less prolonged phases of remission.

Without knowing how to treat the cause, the therapists' attitude towards these 15 diseases consists in treating the progressive relapses so as to obtain, as soon as possible, the remission, then to avoid their reappearance thanks to a treatment of interview. To do this, they mainly use three families of compounds:

- glucocorticoids such as hydrocortisone (Proctocort, Colofoam ^® ), betamethasone (Betnesol ^® ), prednisolone (Solupred ^® , Cortancyl ⁰ ,

Hydrocortancyl ^® ), tixocortol (Rectovalone ^® ) and budesonide (Entocort ^® ), which today represent the reference drugs for inflammatory bowel disease, but which are mainly used in the treatment of moderate to severe flare-ups. given their numerous meta-side effects

25 boliques, trophiques et immune;

- 5-aminosalicylic acid derivatives (5-ASA) such as sulfasalazine (Salazopyrine ^® ), mesalazine (Pentasa ^® , Rowasa ^® ) and olsalazine (Dipentum ^® ), which are rather used in the treatment of relapses weak to moderate or in relay of glucocorticoids, once the remission obtained;

30 and -. immunosuppressants such as azathioprine (Imurel ^® ) and

6-mercaptopurine (Purinethol ^® ), which are generally offered in cortico-resistant or corticosteroid-dependent forms, or to avoid surgery that would be too mutilating (in the case, for example, of large lesions of the small intestine in the Crohn's disease).

These anti-inflammatory treatments are always combined with symptomatic treatment intended to relieve the functional manifestations of these pathologies, and in particular with the administration of antispasmodics capable of reducing abdominal pain. A number of studies have shown that dehydroepi-androsterone (or 3? -Hydroxy-5-androstene-17-dione), better known as DHEA, has immunomodulatory properties (Daynes et al, Eur. J Immunol, 1990, 20: 793-802; Padgett and Loria, J. Immunol, 1994, 153: 1544-1552; Meikle et al, J. Steroid Biochem. Mol Biol, 1992, 42: 293-304), Straub and his collaborators sought to verify the extent to which this steroid hormone could be involved in the pathophysiological mechanisms involved in inflammatory bowel disease. They have thus shown that patients suffering from Crohn's disease or ulcerative colitis have serum levels of DHEA sulfate - which represents the circulating form of DHEA - significantly lower than those found in healthy subjects (Am. J. GastroenteroL, 1998,

93: 2197-2202).

These observations were later confirmed by De La Torre et al (Clin. Exp. Rheumatol, 1998, 16: 579-582).

At the same time, Poynter and Daynes (J. Biol chem., 1998, 273: 32833- 32841) have shown that DHEA is capable of inhibiting various mediators of inflammation such as the nuclear factor KB, interleukin 6 (IL-6 ) and interleukin 12 (IL-12) via the receptor activated by peroxisome proliferators (PPARα).

All of these results therefore led Andus et al. (CD ROM, Abstracts & Itenerary Planner, DDW ^® AstraZeneca, 2000) to test the effects of an oral administration of DHEA in 8 patients with an outbreak of ulcerative colitis refractory to conventional treatments. In these trials, each patient received 200 mg of DHEA once daily for 56 days, more than its usual treatment, 6 patients already on mesalazine (1.5-4 g / d), 2 on budesonide (1.5-3 g / d), 1 on azathioprine (150 mg / d) and 1 on H15 and E. coli

Nissle 1917 (100 mg / day). A significant reduction in the inflammatory thrust was obtained in 5 patients going, for 3 of them, until a remission. On the other hand, no significant effect of DHationA on inflammation was observed in the

3 other patients.

In parallel with the work carried out on DHEA, numerous studies have also focused over the last ten years on the 7-hydroxylated derivatives of

DHEA, and in particular on 7βr-hydroxy-DHEA (or 3?, 7ûr-dihydroxy-5-andro-stène-17-one) whose presence has been revealed throughout the organism, in particular at the level of the barrier organs such as the skin, liver, intestine, prostate and brain.

It is now well established that the 7-hydroxylated derivatives of DHEA, as well as those derived from other 3y # -hydroxylated steroid hormones such as pregnenolone (3? -Hydroxy-5-pregnene-20-one or PREG) and epiandrosterone (3β-hydroxy-5ûr-androstane-17-one or ISOA) exhibit marked antiglucocorticoid activity (Morfin et al, IX Internat. Congr. Hormonal Steroids, DALLAS, TX, USA, Congress Proceedings: poster D103, 1994; Morfin et al , Proceedings of the Second International Conference on Cortisol and Anti-cortisol, Las Vegas, NV, USA: p38, 1997). This activity is said to be the source of their ability to stimulate cellular immunity (Morfin and Courtray, J. Steroid Biochem. Mol. Biol, 1994, 50: 91-100; WO-A-94/08588 published on April 28 1994).

The mechanism by which 7-hydroxysteroids exert their antiglucocorticoid action has not yet been elucidated. We know that there is no cytosolic receptor for 7α-hydroxy-DHEA and that 7-hydroxysteroids do not interfere with the binding of dexamethasone to the glucocorticoid receptor. On the other hand, it would seem that these compounds are capable of decreasing the nuclear incorporation of the glucocorticoid receptor activated by dexamethasone (Morfin et al, ibid).

Anyway, the use of the 7-hydroxylated derivatives of the steroid hormones 3? -Hydroxylés was recommended, in particular in the international application WO 94/08588, in all the therapeutic indications in which an action contrary to that of glucocorticoids is likely to be sought, and in particular in the treatment of pathologies for which glucocorticoids are conventionally not recommended and those linked to side effects of these same glucocorticoids (Cushing's syndrome, adrenal tumors, disorders of the hypothalamic-pituitary-adrenal axis, diabetogenic and immunosuppressive effects - glucocorticoids, muscle atrophy, delayed healing, digestive ulcers, osteoporosis, cataracts, opportunistic infections, dysmenorrhea, ...).

Their use has also been proposed in the prevention and treatment of skin aging which is known to be favored by glucocorticoids (WO 98/40074 published September 17, 1998). Now, in the context of their work, the inventors have found that the 7-hydroxylated and 7-ketone derivatives of the 3 ^ -hydroxylated steroid hormones have an analgesic effect superior to that of DHEA, doubled by an anti-inflammatory effect less equivalent to that of the latter, these two effects being, moreover, more pronounced than those exerted by the compounds conventionally used in inflammatory bowel diseases such as sulfasalazine.

Thus, paradoxically, these derivatives, although exhibiting marked antiglucocorticoid and immunostimulatory activities, appear to present a real interest in the treatment of inflammatory bowel diseases such as Crohn's disease and ulcerative colitis, as well as that of functional intestinal diseases, and in particular irritable bowel syndrome.

A subject of the present invention is therefore the use of at least one compound corresponding to formula (I) below:

or to formula (II) below

in which :

- R _! represents a hydroxyl group, a C, -C ₂₄ organic acid ester group, linear or branched, saturated or unsaturated, a sulfuric acid or phosphoric acid ester group, a C ₂ -C ₂ carbon ether group , linear or branched, saturated or unsaturated, and optionally comprising one or more heteroatoms chosen from nitrogen, oxygen, sulfur and phosphorus, or also a C ₃ -C ₁₀₀ carbohydrate ether group, comprising optionally one or more heteroatoms chosen from nitrogen, sulfur and phosphorus; - R ₂ and R ₃ represent, when present, a hydrogen atom;

- R ₄ represents an oxygen atom or a hydroxyl group;

- Rj represents a hydrogen or oxygen atom, a hydroxyl group or a group of formula: -CH (OH) R, ₅ , = C (OH) R, ₅ , -COR,;, = CHR ₆ or - CH (R ₆ ) R ₇ in which Rg and R ₇ each represent a linear or branched, saturated or unsaturated C 1 -C ₁₀ alkyl group; provided, however, that R „R ₄ and R ₅ do not all represent a hydroxyl group; for the preparation of a medicament for the treatment of inflammatory or functional diseases of the intestine. In accordance with the invention, the compounds corresponding to formula (I) or to formula (II) are preferably used in which R represents a hydroxyl group, while R ₅ represents an oxygen atom or a group -COCH ₃ . These compounds correspond in particular to derivatives

7-hydroxylated and 7-ketones of ISOA in which case, in formula (I), R ₅ represents an oxygen atom, of PREG in which case, in formula (II), R ₅ represents a group -COCH ₃ , and DHEA in which case, in formula (II), R ₅ represents an oxygen atom. Among these derivatives, 7α-hydroxy-DHEA in which the hydroxyl group carried by the carbon atom located in position 7 of the steroid cycle is axial, and 7-oxo-DHEA (or 3 £ hydroxy-5-androstene-7 , 17 ~ dione) are particularly preferred. The medicament prepared in accordance with the invention may comprise one or more compounds of formula (I) or of formula (II), optionally in combination with another active ingredient capable of further increasing its therapeutic efficacy, and one or more pharmaceutically acceptable excipients, of which the choice will depend on the route by which one wishes to administer it and the dosage form in which it is desired to present it.

Indeed, this drug is notably capable of being administered by the three routes conventionally used to treat intestinal pathologies, namely the oral route, the rectal route which is particularly useful when these pathologies are localized in the rectum or region. rectosigmoid, and the parenteral route, especially intramuscular and intravenous, which is necessary when high doses are necessary or when it is not possible to use the oral route, for example in the case of vomiting or disorders of the consciousness.

Furthermore, by their nature both hydrophilic and lipophilic, the compounds of formula (I) and of formula (II) lend themselves to use in numerous galenical forms such as tablets, capsules, granules, solutions, suspensions, suppositories , creams, gels, ointments or mousses.

In all cases, the compound or compounds of formula (I) or of formula (II) are advantageously present in the medicament, when the latter is intended to be administered to humans, in an amount allowing the administration of a dose of compound (s) of between 1 and 50 mg / day and per kg of body weight and, preferably, of between 2 and 10 mg / day and per kg of body weight, this administration can be carried out in one or more taken daily.

The use of the 7-hydroxylated and 7-ketone derivatives of the 3 ^? - hydroxylated steroid hormones for the preparation of a medicament intended to treat inflammatory or functional pathologies of the intestine have numerous advantages. Indeed, in addition to the fact that these derivatives have truly interesting analgesic and anti-inflammatory effects, it has been shown that they are devoid of andro- gene and estrogen, unlike DHEA which is partly metabolized into sex hormones. In addition, they have excellent tolerance.

Other advantages and characteristics of the invention will appear on reading the additional description which follows and which refers to examples of demonstration of the analgesic and anti-inflammatory effects of the 7-hydroxylated and 7-ketone derivatives of the steroid hormones 3 ^ -hydroxylées. It goes without saying, however, that these examples are given solely by way of illustrations of the invention and in no way constitute a limitation thereof.

Example 1 Comparative Effectiveness of 7ûr-hydroxy-DHEA, 7-oxo-DHEA and DHEA on Abdominal Pain

The analgesic power of 7ûr-hydroxy-DHEA, 7-oxo-DHEA and DHEA was evaluated by a test conventionally used to assess the analgesic effect of a substance and which consists in researching whether this substance is suitable , when given orally, to provide protection against abdominal cramps or contortions caused in mice by intraperitoneal injection of phenylbenzoquinone.

To do this, 162 male ICR mice weighing 20 to 25 g each were randomly divided into 17 lots and, with the exception of the mice in lot 1 (30 mice) of the animals to serve as controls, received orally. : batch 2 (8 mice): 100 mg / kg of 7 "" - hydroxy-DHEA batch 3 (4 mice): 150 mg / kg of 7ûr-hydroxy-DHEA batch 4 (14 mice): 200 mg / kg of 7 ûr-hydroxy-DHEA lot 5 (4 mice): 300 mg / kg of 7ûr-hydroxy-DHEA lot 6 (8 mice): 400 mg / kg of 7 <ar-hydroxy-DHEA lot 7 (8 mice): 50 mg / kg of 7-oxo-DHEA batch 8 (12 mice): 100 mg / kg of 7-oxo-DHEA batch 9 (4 mice): 150 mg / kg of 7-oxo-DHEA batch 10 (16 mice): 200 mg / kg of 7-oxo-DHEA batch 11 (4 mice): 300 mg / kg of 7-oxo-DHEA batch 12 (12 mice): 400 mg / kg of 7-oxo-DHEA batch 13 (8 mice): 100 mg / kg DHEA . batch 14 (4 mice): 150 mg / kg of DHEA

. batch 15 (14 mice): 200 mg / kg of DHEA

. batch 16 (4 mice): 300 mg / kg of DHEA

. lot 17 (8 mice): 400 mg / kg of DHEA. 30 minutes after the administration of these treatments, 0.20 ml of a 2 to 10,000 aqueous solution of phenylbenzoquinone was injected into all the mice intraperitoneally.

The number of abdominal cramps presented by each of the mice was counted for 5 minutes from the 6 ^th minute and until the H ^th minute following the injection of phenylbenzoquinone. During this count, only the abdominal cramps characterized by the stretching of the hind legs and / or the digging of the flanks after torsion were considered positive.

The mean abdominal cramps presented by the mice were determined for each batch of mice, then, for batches 2 to 17, a percentage P of protection by the formula: P = [(mean cramps presented by the mice of the control group - average of cramps presented by the mice of the treated group) / average of cramps presented by the control group] x 100.

Figure 1 represents, in the form of curves, the evolution of this percentage of protection as obtained after treatment with 7ûr-hydroxy-DHEA (- m -), 7-oxo-DHEA (- A -) and by DHEA (- 4 -) depending on the dose administered.

This Figure shows that at equivalent doses, 7 ^ -hydroxy-DHEA and 7-oxo-DHEA systematically exhibit an analgesic effect greater than that of DHEA. The best analgesic effect is obtained with 7ûf-hydroxy-DHEA, which provides almost maximum protection (95%) against abdominal cramps at a dose of 400 mg / kg. The maximum protection obtained at the same dose is 67% for 7-oxo-DHEA and drops to 55% for DHEA. Example 2 Comparative Effectiveness of 7û-hydroxy-DHEA, DHEA and Sulfasalazine on Colon Inflammation The anti-inflammatory power of 7ûr-hydroxy-DHEA was assessed by a test intended to evaluate the effect therapeutic of an oral administration of this compound on an inflammation of the colon induced in rats by the administration intracolonic trinitrobenzene sulfonic acid (TNBS), and was compared to the therapeutic effects obtained, under the same conditions, by the oral administration of sulfasalazine on the one hand, and of DHEA on the other hand.

To do this, 72 male Wistar rats each weighing between 220 and 240 g, previously fasted for 48 hours, were divided into 8 batches and were treated orally as follows:

• lot 1 (6 rats): aqueous gel containing 1% carboxymethylcellulose (CMC) once / day from D0 to D4

• lot 2 (6 rats): 100 mg / kg of sulfasalazine 1 time per day from D0 to D4. lot 3 (10 rats): 50 mg / kg of DHEA once a day from D0 to D4

. lot 4 (10 rats): 100 mg / kg of DHEA once a day from D0 to D4

. lot 5 (10 rats): 200 mg / kg of DHEA once a day from D0 to D4

• batch 6 (10 rats): 50 mg / kg of 7ar-hydroxy-DHEA 1 time per day from D0 to D4

• batch 7 (10 rats): 100 mg / kg of 7ûr-hydroxy-DHEA 1 time per day from OJ to D4. lot 8 (10 rats): 200 mg / kg of 7ar-hydroxy-DHEA 1 time per day from D0 to D4

Sulfasalazine and 7ar-hydroxy-DHEA were administered to rats as an aqueous gel containing 1% CMC (PROLABO), while DHEA was administered as an aqueous gel containing 5% DMSO and 1% CMC. At D0 + 8 hours, the rats were anesthetized with halothane and they were administered in the colon 250 μl of a solution consisting of 50% physiological saline and 50% ethanol and containing 30 mg of TNBS, by means of a 2 mm catheter inserted into the anus over a length of 8 cm. This treatment causes clinical signs in a few days similar to those observed in Crohn's disease and ulcerative colitis.

The rats were weighed daily for the duration of the experiment.

On D4, the rats were euthanized with halothane. The colonists were removed over a length of 8 cm from the splenic elbow to the rectum, rinsed with physiological saline, cut longitudinally and examined under the microscope. Lesions were counted according to the so-called Morris score

(Morris et al, 1989, Gastroenterology, 96: 795-803) after double blind reading.

The body weights of the rats were analyzed by a Student t test. The normality and homogeneity of variance tests (Levene test), followed by parametric tests were carried out on the scores for the lesions.

Figure 2 represents, in the form of curves, the evolution of the body weight of the rats of batch 1 control (- -) and of the treated batches (batch 2: - “-; batch

3: - A -; lot 4: - π -; lot 5: - - - m - - -; lot 6: - o -; lot 7: • - - -; lot 8: - - -) as a function of time, while Figure 3 represents, in the form of histograms, the scores of the lesions obtained for these same rats.

Figure 2 shows that the body weight of the rats evolved during the test in the same way in all the batches, control or treated, while Figure 3 shows that the rats treated with sulfasalazine, DHEA and 7-hydroxy -DHEA show lesions significantly less significant than those presented by the rats of the control group.

The lowest lesion scores are obtained in rats treated with DHEA and 7α-hydroxy-DHEA, the anti-inflammatory effects of which appear to be equivalent at equivalent doses. Example 3 Comparative Effectiveness of 7-oxo-DHEA and Sulfasalazine on Colon Inflammation

The anti-inflammatory power of 7-oxo-DHEA was assessed by the same test as that described in Example 2, and was compared with that observed, under the same conditions, with treatment with sulfasalazine. 70 male Wistar rats each weighing between 220 and 240 g, previously fasted for 48 hours, were divided into 7 lots and were treated orally as follows: lot 1 (10 rats): aqueous gel containing 1% CMC 1 time / day from D0 to D4 batch 2 (10 rats): 50 mg / kg of sulfasalazine 1 time per day from D0 to D4. batch 3 (10 rats): 100 mg / kg of sulfasalazine 1 time per day from D0 to D4 batch 4 (10 rats): 200 mg / kg of sulfasalazine 1 time per day from D0 to D4 batch 5 (10 rats): 50 mg / kg of 7-oxo-DHEA 1 time per day from D0 to D4 . lot 6 (10 rats); 100 mg / kg of 7-oxo-DHEA 1 time per day from D0 to D4

. batch 7 (10 rats): 200 mg / kg of 7-oxo-DHEA 1 time per day from D0 to D4

As in Example 2, sulfasalazine was administered to the rats in the form of an aqueous gel containing 1% CMC, while 7-oxo-DHEA was administered to them in the form of an aqueous gel containing 5 % DMSO and 1% CMC.

Inflammation of the colon was induced under the same conditions as those described in example 2, except that the rats received a dose of TNBS half that of that administered in this example (15 mg versus 30 mg). Then the test was continued as described in Example 2.

Figure 4 represents, in the form of curves, the evolution of the body weight of the rats of the control batch 1 (- 4 -) and the treated batches (batch 2: - "-; batch 3: - A -; batch 4: - x -; lot 5: - * -; lot 6: - • -; lot 7: - i -) as a function of time, while Figure 5 represents, in the form of histograms, the scores of lesions obtained for these same rats.

Figure 4 shows that, there too, the body weight of the rats evolved during the test in the same way in all the batches, control or treated

As for Figure 5, it shows that the rats treated with sulfasalazine and 7-oxo-DHEA show lesions significantly less significant than those presented by the rats of the control group. The anti-inflammatory effects of sulfasalazine and 7-oxo-DHEA appear to be equivalent to the doses of 50 and 200 mg / kg per day.

On the other hand, at the dose of 100 mg / kg per day, the anti-inflammatory effects of 7-oxo-DHEA are significantly higher than those of sulfasalazine (P <0.01, Mann-Whitney test).

Claims

1. Use of at least one compound chosen from 7α-hydroxy-DHEA (3?, 7α-dihydroxy-5-andro-stène-17-one) and 7-oxo-DHEA (or 3? -Hydroxy-5 -androstene- 7,17-dione), for the preparation of a medicament for the treatment of inflammatory or functional diseases of the intestine.

2. Use according to claim 1, in which the compound is present in the medicament in an amount suitable for the administration of a dose of said compound of between 1 and 50 mg / day and per kg of body weight, in humans .

3. Use according to any one of the preceding claims, in which the compound is present in the medicament in an amount suitable for the administration of a dose of said compound of between 2 and 10 mg / day and per kg of body weight, in humans.

4. Use according to any one of the preceding claims, in which the medicament is intended to treat Crohn's disease or ulcerative colitis.

5. Use according to any one of claims 1 to 3, wherein the medicament is intended to treat irritable bowel syndrome.