Retinoic acid (RA) supports cellular growth and differentiation and has been shown to attenuate or completely reverse the malignant phenotype for many cell lines.

Retinoic acid is also implicated in cell proliferation in skin conditions. Synthetic retinoids have shown promise in the treatment of oral leukoplakia and head and neck cancer, although their use may be associated with significant toxicity. Retinoids have also been used topically and orally for the treatment of skin diseases, i. e. actinic keratoses (precursors to squamous cell carcinoma), nonmelanoma skin cancer, acne vulgaris, psoriasis and disorders of keratinisation.

More recently, the benefit of retinoids to photodamaged skin has led to their use in anti- wrinkle cosmetic preparations.

Vitamin A (retinol) is oxidised through retinal by dehydrogenases in the cytoplasm of target cells in low yield to all trans-retinoic acid (RA). RA is at least 100 fold more active than retinol and is considered to account for its biological action. RA has a short half life (c. 1 hour) and therefore the potency of RA is reduced when administered systemically due to metabolism by human liver and intestine cytochrome P450s to the inactive 4-hydroxy- RA and thence by dehydrogenases to the partially active 4-keto-RA and inactive polar metabolites. The specific P450s (P450-RA) responsible for 4-hydroxylation of RA in the human liver have not been characterised, but several reconstituted P450s CYP1A2/2B6/2C8/2D6/2E1/3A4, can catalyse the reaction. Repeated exogenous administration of RA leads to a lowering of RA levels due to induction of the metabolising enzymes. A drug which can prolong and intensify the action of endogenous RA on the epidermal cell by inhibiting P450-RA metabolising enzymes would have potential as a clinical agent in the treatment of certain skin conditions.

The imidazoles, ketoconazole and liarozole, have been reported as inhibitors of RA-metabolising enzymes whilst being studied as inhibitors of 17a-hydroxylase: 17,20-lyase (P450 17a) as agents for the treatment of androgen-dependent prostatic cancer by lowering testosterone levels. Ketoconazole lacks specificity towards P450 17 and inhibits several other cytochrome P450 enzymes on the steroidogenic pathway of androgen synthesis and has a poor pharmacokinetic profile.

Liarozole (R75251), a related compound, also inhibits testicular (but not adrenal) P450 17a and similarly lowers testosterone levels in human volunteers, but its effect on androgen-independent carcinoma has been partially attributed to inhibition of P450-RA with an associated increase in RA levels (ketoconazole is also an inhibitor of this enzyme). This view has been confirmed by experiments showing that RA metabolism in epidermal cells is inhibited (IC50-2ptM) and also that endogenous RA levels are increased and the elimination rate from the plasma of injected RA is reduced. Liarozole has also been in trials for the treatment of severe psoriasis (oral) and acne (topical and oral) and it seems likely that this action is due to inhibition of epidermal P450-RA.

RA-metabolising enzyme inhibitors intended for topical use for skin diseases should show little unwanted systemic effects due to the involvement of other enzymes, e. g. steroidogenic P450 enzymes such as P450 17a, CSCC, 21-hydroxylase. This objective should be achievable due to either poor sub-dermal penetration or, should this occur, low dose effects due to plasma dilution of the small load of drug placed on a restricted area of skin.

It is therefore the aim of the present invention to provide a P450-RA inhibitor of low activity towards other steroidogenic P450 enzymes and therefore low inherent toxicity to alleviate some of the aforementioned problems.

We have now discovered compounds that are inherently less active inhibitors of P450 17a, the main target of liarozole and ketoconazole, and yet are inhibitors of retinoic acid metabolism with potency comparable with that of ketoconazole. This selectivity for inhibition of retinoic acid metabolism is useful in the treatment of the skin conditions mentioned above.

According to the present invention there is provided a class of pyrrolidine diones suitable for use as retinoic acid metabolising enzyme inhibitors, which pyrrolidine diones are represented by the general formula (I): in which R'is hydrogen or an alkyl, cycloalkyl or aralkyl group; each of R'and R'is independently hydrogen or an alkyl, cycloalkyl or aryl group; and each of x and y is independently zero or one; wherein at least one of R', W and R3 has at least five carbon atoms.

W and R3 can, of course, be the same or different. It is preferred that at least one of Rl, R2 and le is a hydrogen atom; when R'is an aralkyl group, it is preferably benzyl (HSC6-CH2-). When any of R', R2 and R3 is an alkyl group, it preferably has 1 to 7 carbon atoms; when the relevant substituent is cycloalkyl it preferably has 5 or 6 carbon atoms.

The compounds according to the invention have low activity against other P450 enzymes involved in cortisol production, which is indicative of low toxicity should systemic absorption occur. The compounds according to the invention also have a short half life in the rat, [about lh] further emphasising their suitability for topical application. The compounds are racemates and it might be expected that one of the resolved enantiomers would possess most of the activity in accordance with general findings for inhibition of P450s.

The following Table 1 lists exemplary compounds according to the present invention, together with their respective values for inhibition of retinoic acid metabolism (according to the protocol described herein under the heading"In vitro inhibition studies").

This shows that the compounds according to the invention have potency comparable to that of ketoconazole: Table 1. Inhibition of P450-RA Inhibition(100µM)Compound% 1 2 3 4 5 6 Ketoconazole The pyrrolidine diones according to the present invention are suitable for use as retinoic acid metabolism inhibitors. Pharmaceutically acceptable salts of the pyrrolidine diones of the present invention (such as chloride, citrate or sulphate salts) may also be suitable for use as retinoic acid metabolism inhibitors. The present invention therefore preferably further comprises a pharmaceutical formulation comprising such a pyrrolidine dione (or such a salt), together with a pharmaceutically acceptable carrier, diluent or excipient therefor.

According to the present invention, the preferred route of administration of the formulation is topical. A formulation suitable for topical use may contain the free base of a pyrrolidine dione according to the invention. Furthermore, it is preferred that the formulation suitable for topical use contains a pharmaceutically acceptable carrier, diluent or excipient which is substantially non-aqueous in nature. The formulation may be in the form of a gel, foam, salve, emollient, cream or ointment, and may, in some cases, contain paraffin wax.

The pyrrolidine dione (or salt thereof) can be used for the treatment of dermatological conditions in (i) a dose form designed for topical administration in a dose range of 0.5-10% (w/v) with a concentration of 1 to 3% being preferred, and (ii) a dose form designed for oral administration in a dose range of 50-SOOmg daily with a daily dose of 150 to 250mg being preferred.

Alternatively, the pharmaceutical formulation according to the invention may be in a form suitable for oral administration. The formulation suitable for oral administration may comprise a suitable pharmaceutically acceptable salt, or free base, of the pyrrolidine dione according to the invention. In this case, the diluent, carrier or excipient may comprise, for example, lactose, microcrystalline cellulose and/or calcium phosphate dihydrate.

A preferred compound is compound 6, in which the 3-position of the pyrrolidine dione is substituted with the 1- (4-aminophenyl)-2-phenylethyl group.

According to another embodiment of the present invention, a pyrrolidine dione according to the invention (or salt thereof) may also be used in the treatment of various dermatological conditions. These dermatological conditions may include ichthyosis, acne, psoriasis, wrinkles or photodamaged skin. Furthermore, the pyrrolidine diones of the present invention (or salts thereof) may be used in anti-ageing preparations and in preparations to protect the skin from the effects of radiation damage caused, for example, by radiation therapy of tumours and to alleviate such damage.

Table 2. In vitro activity profiles of (I), aminoglutethimide and ketoconazole P450AROMP45017αP450C21 ICso (µM) IC50(µM) % inhibition % inhibition (at 200 yM) (at 100 µM) Aminoglutethimide 24.50 53.3 23.9 7.6 (I) 0.35 410 25.5 19.2 Ketoconazole ND ND ND 95.7 ND = not done Exemplary methods for the preparation of compounds according to the present invention, and their biological screening, will now be described.

METHODS In Vitro Inhibition Studies 1. P450-RA Sets of tubes, in triplicate, with a total volume of 400µl containing (11,12-3H) retinoic acid lOnM (10µl of 400 nM stock), unlabelled retinoic acid in methanol (10µl of 120jnM stock to give 3, uM), inhibitor (8µl of 5mM ethanol stock to give 100, uM concentration in final assay volume), phosphate buffer 50mM (pH=7.4, 312µl), NADPH solution (50AI of 16mg/ml) were prepared, and the tubes vortexed and preheated in a water bath for 4 min.

The enzyme reaction was initiated by addition of rat liver microsomes (10ttl of 10mg/ml) and the mixture incubated at 37°C for 25 min. The enzyme action was arrested by addition of 100il of 1 % formic acid and the tubes were placed in ice for 5 min. Then 3ml of ethyl acetate containing 0.02% butylated hydroxy anisole was added and the tubes vortexed for 10 s. The tubes were then left for another 5 min at room temperature and the organic layer (2ml) was removed from each tube and transferred to another set of tubes. The ethyl acetate extracts were evaporated using a univap centrifuge connected to a vacuum pump and a multitrap at-80°C.

After 60 min the tubes were removed and the residue was reconstituted in absolute methanol (1001tel) and 50y1 was injected into a HPLC machine equipped with a 10µm C18 yBonda pack column (3.9x300mm, Millipore) using acetonitrile: water (containing 1 % ammonium acetate) 75: 25 as mobile phase and connected to a mixer (Reeve model 9702) and a radioactive detector (Reeve model 9701). The results were analysed by an on line computer system. The scintillation fluid used was a mixture of 50: 50 Hisafe 3 and methanol.

Percentage inhibition was calculated from the conversion rate of the samples containing inhibitors to that of control samples which contained absolute ethanol instead of inhibitor solution. Ketoconazole was used as a standard.

Due to the sensitivity of retinoic acid all the above assays were carried out in a dark room equipped with yellow light.

2. P450 17a The incubation mixture consisting of test compound (100 yM, final concentration) in ethanol (10µl), NADPH generating system (50, u1) and [1,2,6,7-3H] 17a- hydroxyprogesterone (1, uM final concentration, 10/cl) made up to a final volume of 0.5ml with phosphate buffer (50, uM, pH 7.4) was warmed in a shaking water bath at 37°C for 5 min and the reaction was started by addition of the rat testis microsome suspension (0.12 mg/mg final protein concentration). After incubation at 37°C for 30 min, the experiment was terminated by the addition of diethyl ether (3 ml). Following extraction, the ether layer was evaporated to dryness and the extract, reconstituted in acetone (50AI) was chromatographed on plastic- backed silica gel plates using chloroform-cyclohexane-ethyl acetate-methanol (80: 10: 10: 4 v/v).

Location of labelled steroids on the TLC plates was assessed by adding unlabelled steroid on the same plate.

The relevant spots, located under uv light (254 nm) on the TLC plates, were transferred to vials containing acetone (lml) and scintillation fluid (2ml), mixed thoroughly and counted in a liquid scintillation counter. A control was run containing ethanol without inhibitor. Enzyme activity was measured as conversion of substrate to products (androstenedione and testosterone).

3. P450scc (cholesterol side chain cleavage enzyme) Incubation mixture (0.5ml) in phosphate buffer (50mM, pH 7.4) containing a range of concentrations of test compound in ethanol (20µl) NADPH generating system (50µl) and [26, (27)-3H] 25-hydroxycholesterol and 25-hydroxycholesterol (0.412 µM final concentration; 50µl) were warmed in a shaking water bath at 37°C for 5 min and the reaction was started by addition of the mitochondrial fraction of bovine adrenal cortex (0.2mg/ml protein final concentration, 10µl). After incubation at 37°C for 15 min, aliquots (2501tu) were added separately to tubes containing NaOH (1 M, O. lml) and chloroform (lml). The mixture was vortexed and then aliquots (3001tu) from the aqueous layer were added to tubes containing alumina (0.25g) and water (750AI). The tubes were centrifuged for 20 min at 3000 rpm and then aliquots of the supernatant (5001tu) were removed and added to instagel scintillation fluid (3 ml) and counted for 1 min. A control was run containing ethanol without inhibitor. The ICso values for AG and I were obtained from graphical plots of the percentage remaining enzyme activity versus Log10 [inhibitor].

4. P450c21 (21-hydroxylase) The incubation mixture contained test compound (20OItM final concentration) in ethanol (20µl), NADPH generating system (50µl) and [4-l4C] progesterone (2.12yM final concentration, 10µl) and was made up to a final volume of 0.5ml with phosphate buffer (50mM, pH 7.4). It was warmed in a shaking water bath at 37°C for 5 min and the reaction was started by addition of rat adrenal microsomes (10, ul, final protein concentration 0.045mg/ml). The experiment was terminated after 15 min by addition of diethyl ether (3ml).

Following the extraction into the diethyl ether (2 x 3ml) the ether layer was evaporated to dryness and the residue was reconstituted in acetone (50µl) for chromatography on plastic- backed silica gel plates, using chloroform-ethyl acetate (80: 20 v/v). Location of labelled steroid on the TLC plates was assessed by adding unlabelled steroid on the same plate. The spots located under uv light (254 nm) were transferred to vials containing acetone (lml) and scintillation fluid (2ml), mixed thoroughly and counted in a liquid scintillation counter. A control was run containing ethanol without inhibitor.

5. P450AROM (aromatase) Incubation (0.5ml) in phosphate buffer (50mM, pH 7.4) containing the NADPH generating system (50 1), a range of concentrations of inhibitor in ethanol (101tu) and [lp_ 3 H] androstenedione (0. 4µM final concentration, 10µl) were warmed to 37°C for 5 min and the reaction was started by addition of the placental microsomal protein (0.2mg/ml, 10µl). After incubation for 5 min aliquots (300µl) were removed and added separately to a cold mixture of 1 % (w/v) activated charcoal (900µl) and mercuric chloride (lmM, 300/il). The mixture was vortexed, allowed to stand for 20 min and then centrifuged at 3000 rpm at 4°C for 20 min.

Aliquots 300µl) from the supernatant were dispersed in Instagel (2ml) and counted for 1 min.

A control was run containing ethanol without inhibitor. The ICso values were obtained from graphical plots of the percentage remaining enzyme activity vs Log10 inhibitor concentration.

Synthesis General Methods for the Preparation of Compound 1, 2 and 4 W Fy N143 C-CT42 C CHZ CNCO COOHCOOL H R-X RetluxAcetone K, C03 O, N Fuming Nitric Acid <1 -40'C R R O Pd/C Ho 8 f-+ If-+ HCI gas C. gas I ! R I R R 3-Phenylpyrrolidine-2,5-dione (7) Phenylsuccinic acid (33g, 0.165mol) was dissolved in 0.88 ammonia (45ml) and heated on a metal bath (180-200°C) until a white solid was formed. Further heating to 200°C resulted in the formation of a brown viscous liquid which solidifed on trituration with ether to give pale yellow crystals. Recrystallisation (ethanol) gave white crystals (25g. 75.5%), m. p. = 89- 91°C (from methanol). v., 3200, 3100 (NH); 1770,1710 (C=O), 1600 (Ph) cmi'. 8 (CDCl3), 9.4 (1H, br, N-H), 7.34 (5H, m, Ph-H), 4.04 (1H, dd, JAx= 5Hz and Jx = lOHz, CHxCHHB), 3.20 (1H, dd, JxB=9Hz and JAB = 8Hz, CHXCHAHB), 2.77 (1H, dd, JAX = 5Hz and JBA = 18 Hz, <BR> <BR> <BR> <BR> CHxCHAHB).<BR> <BR> <BR> <BR> <BR> <BR> <BR> <P> I-Octvl-3-phenvlpvrrolidine-2. 5-dione (8) A mixture of 3-phenylpyrrolidine-2,5-dione (7) (4g, 0.0228 mol), anhydrous K2CO3 (9.5g, 0.0684 mol), iodo octane (6.5g, 0.0273 mol) and acetone (50 ml) was refluxed for 3h. The reaction was monitored by TLC using the solvent system petroleum ether (60-80°C) ethyl acetate (2: 1) until the reaction was complete. The powder bed was removed by filtration and the filtrate collected and concentrated under vacum to leave a brown oil. The crude oil was dissolved in ethyl acetate and passed down a column (30 mm x 400 mm) of silica gel (0.063-0.200 mm). The unreacted iodo-octane was washed out using light petroleum ether (40-60°C) and 1- octyl-3-phenylpyrrolidine-2,5-dione (8) was eluted using the solvent system petroleum ether (40- 60°C/ethyl acetate (2: 1). Evaporation of the solvent gave (8) as a yellow oil (4.5 g, 68.5%).

(Found: C, 75.36; H, 8.62; N, 4.89. C18H25O2N requires C, 75.22; H, 8.77; N, 4.78%. vu,, 2940, 2860 (CH); 1780,1700 (C=O); 1600 (Ph) cri'. # (CDCl3) 7.25 (5H, m, Ph-H), 3.92 (lH, dd, JAX = 5Hz and JBx = 9Hz, CHx CHAHB), 3.5 (2H, t, J = 8Hz, N-CH2), 3.08 (lH, dd, JXB = 9Hz and J, = 18 Hz, CH, CHAHB), 2.65 (1H, dd, JXA = 5Hz and JBA = 18 Hz, CH, CHAHB), 1.55-0. 85 (15H, m,(CH2)6-CH3).

3-(4'-Nitrophenyl)-1-octylpyrrolidine-2,5-dione(9) In a three-necked flask, fitted with a mechanical stirrer was placed 1-octyl-3- phenylpyrrolidine-2,5-dione (8) (4g, 0.0139 mol) and the contents of the flask were cooled in a petroleum ether (60-80°C)/cardice bath to-40°C. Cooled (-40°C) fuming nitric acid (25 ml) was then added at such a rate that the temperature was maintained between-40° and-30°C.

After lh the cooling bath was removed and the temperature allowed to rise to-10°C. The resulting mixture was poured with vigorous stirring into ice/water (200 ml), and extracted with chloroform (2 x 100 ml). The combined organic layers were washed with sodium bicarbonate solution (1 x 100 ml), water (1 x 100 ml), dried (MgSO4) and the solvent removed under reduced pressure to leave 3-(4'-nitrophenyl)-1-octyl pyrrolidine-2,5-dione (9) as a yellow oil (4.3g; 93%).

(Found: C, 64.78; H, 7.39; N, 8.33. C18H24O4N2 requires C, 65.04; H, 7.28; N, 8.43%). v. 2940-2870 (CH); 1780,1700 (C=O); 1530,1350 (NO2) cm-1. #(CDCl3) 8.12 (2H, d, J = 9Hz, Ph-H), 7.46 (2H, d, J = 9Hz, Ph-H), 4.10 (1H, dd, JAX= 5Hz and JBX = 9Hz CHX CHAHB), 3.50 (2H, t, J = 6Hz, N-CH2), 3.25 (1H, dd, JxB= 9Hz and JAB = 18 Hz, CHX CHAHB), 2.75 (1H, dd, <BR> <BR> <BR> JXA = 5Hz and JBA = 18 Hz, CHX CHA HB), 1.60-0.8 (15H, m, (CH2) 6-CH3).<BR> <BR> <BR> <BR> <BR> <BR> <P>3- (4'-Aminophenyl)-1-octyl pyrrolidine-2.5-dione HO) A mixture of 3- (4'-nitrophenyl)-1-octyl pyrrolidine-2,5-dione (9) (4.0g, 0.01204 mol) and palladium on charcoal catalyst (10%, 0.4g) in absolute ethanol (50 ml) was shaken with hydrogen gas at atmospheric pressure until the uptake of hydrogen was complete (820 cc). The charcoal was then filtered off and the solvent removed under reduced pressure, to leave 3- (4'- aminophenyl)-1-octyl pyrrolidine-2,5-dione (10) as a yellow oil 55%).

(Found: C, 70.88; H, 8,. 74; N, 9.55. C18H26O2N2 requires C, 71.49; H, 8.67; N, 9.26%). v.,,,,,, 3380 (NH); 2940-2860 (C-H); 1780,1700 (C = 0); 1620 (Ph) cm-1. #(CDCl3), 6.95 (2H, d, J=9Hz=Ph-H), 6. 65 (2H, d, J = 9Hz, Ph-H) 3.85 (1H, dd, JAx = 5Hz and JBX = 10 Hz, C_x CHAHB), 3.55 (4H, m, N-CH2, NH2), 3.15 (1H, dd, JXB= 8Hz and JAB = 18 Hz, CHX CHAHB), 2.75 (1H, dd, JXA = 4Hz and JBA = 18 Hz, CHx CHA HB), 1.60-0.8 (15 H, m, (CH2)6-CH3).

3- 4'-Aminophenyl-1-octyl pvrrolidine-2.5-dione hydrochloride (11) 3-(4'-Aminophenyl)-1-octyl pyrrolidine-2,5-dione (10) (2.0g, 0.0066 mol) was dissolved in anhydrous ether and dry hydrogen chloride gas was passed through. The white precipitate formed was filtered off and recrystallised (ethanol) to give white crystals of 3- (4'- aminophenyl)-1-octyl pyrrolidine-2,5-dione hydrochloride (11) (1.0g, 44.6%) m. p. = 198-200°C.

(Found: C, 63.56; H, 7.94; N, 8.30. C18H27O2N2Cl requires C, H, 8.03; N, 8.26%). V..

(KBr) 2980,2920 (C-H), 2610 (NH+3, 1780,1740 (C=O), 1620 Ph cri'. 8 (DMSO) 7.34 (4H, s, Ph-H), 4.21 (1H, dd, JAX = 5Hz and JBX = 9Hz, CHx CHAT$), 3.38 (2H, t, J = 7Hz, N-CH2); 3.18 (1H, dd, JXB = 9Hz, and JAB = 18 Hz, CHX CHAHB); 2.72 (1H, dd, JXA = 5Hz and JBA = 18Hz, CHx CHAHB), 1.58-0.85 (15H, m, (CH2) 6-CH3).

Method 2 on Fumin Nitric acid _ CT-t"-CHz o -40 C H HZ COOT-1 COOL COOL Reflux CH: COCI Toluene 02N N (R_NHz I Toluene Cl CH CH-CH2 COOHCO-NH-R RefluxCH3COCI Toluene HyN H2N I H _ 02Not A '. O N O RI R R Ov N O GARS Gaz R R 2- (4'-Nitrophenyn-butan-1. 4-dioic add (11) In a three-necked flask, fitted with a mechanical stirrer, phenyl succinic acid (lOg, 0.0515 mol) was added portionwise to fuming nitric acid (50 ml) at-40°C (acetone/cardice bath) at such a rate as to maintain the temperatue at-40°C. After Ih the resulting mixture was poured with vigorous stirring into ice/water (300 ml) and the white precipitate formed was filtered off to <BR> <BR> <BR> <BR> give 2- (4'-nitrophenyl) butan-1,4-dioic acid (11) (6.8 g, 68%) m. p. 212-216°C. v.. 2920 (CH2); 1728 (C=O); 1610 (Ph-H); 1532,1361 (NO2) cm-1. #(CDCl3) 8.22 (2H, d, J = 9Hz, Ph-H), 7.56 (2H, d, J = 9Hz, Ph-H), 4.20 (1H, dd, JBX = 9Hz and JAx =6Hz, C_x CHA HB), 3.23 (1H, dd, JAB = 18 Hz and JxB = 9Hz, CHX CHA HB), 2.70, dd, JBA = 18 Hz and JXA = 6Hz, CHX CHA HB).

2-(4'-Nitrophenyl butanedioic anydride (12) A mixture of 2- (4'-nitrophenyl) butan-1,4-dioic acid (11) (3.7 g, 0.0154 mol) toluene (50 ml) and acetyl chloride (6.0 g; 0.0774 mol; 5.5 ml) was placed in a 200 ml flask and refluxed until a clear solution was obtained. The toluene was then removed under pressure to leave 2- (4'-nitrophenyl) butanedioic anhydride (12) as a yellow oil (3.4 g; 99%). vmatx 1879,1800 (C=O); 1600 (Ph); 1530,1350 (NO2) cm~'.

4-Cyclohexylamino-2- (4'-nitrophenyl)-4-oxo-butanoic acid (13) 2- (4'-nitrophenyl) butanedioic anhydride (12) (3.4 g, 0.0153 mol) was suspended in toluene (40 ml) and cyclohexylamine (3.0 g, 0.0307 mol; 3.5 ml) was added dropwise. The mixture was allowed to stand at room temperature for 30 min and the purple solution obtained was acidified with hydrochloric acid (2M). The organic layer was then washed with water (2 x 50 ml), dried (MgSO4) and the solvent removed under reduced pressure to leave a yellow solid.

The solid was recrystallised (ethanol) to give the butanoic acid (13) (3.8 g, 77.5%) m. p. 165°C (Found: C, 60.55; H, 6.54: N, 8.62. C16H20O5N2 required C, 59.99; H, 6.29; N, 8.75%). v.,,,, 3350 (NH); 2940 (CH); 1760,1650 (C=O); 1530,1350 (NO2)cm-1. #(DMso) 8.12 (2H, d, J = 9Hz, Ph-H), 7.6 (1H, d, J = 8Hz, NH), 7.5 (2H, d, J = 9Hz, Ph-H), 4.13 (1H, dd, J = = 6Hz and JBX=8 Hz, CHX CHAHB), 2.95 (1H, dd, J = 8Hz and JAB = 18 Hz, CHX CHAHB), 2. 55 (1H, dd, JXA = 6Hz and JBA = 18 Hz, CHx CHAHB), 2.2-1.2 (11H, m, N-cyclohexyl).

1-Cvclohexy4'-nitrophenyl) pyrrolidine-2.5-dione (14) A mixture of 4-cyclohexylamino-2- (4'-nitrophenyl)-4-oxo-butanoic acid (13) (3.6 g; 0.0112 mol), toluene (40 ml) and acetyl chloride (4.4 g; 0.056 mol; 3.9 ml) was refluxed for 3h at 90°C until a clear solution was obtained. The solvent was then removed under reduced pressure to give a brown oil which was dissolved in ethanol and decolourised using charcoal. The yellow filtrate was concentrated under vacuum to leave a yellow oil which solidified on standing at room temperature. Upon washing with cold ether, white crystals were obtained of 1- cyclohexyl-3- (4'-nitrophenyl) pyrrolidine-2,5-dione (14), (1.4 g; 41.2%) m. p. 92.5-94.5°C (Found C, 63.53; H, 5.87; N, 9.27. C, 6H, S04N2 requires C, 63.56; H, 6.00; N, 9.27%). <BR> <BR> <BR> <BR> v. 2940 (C-H); 1780,1700 (C=O); 1610 (Ph); 1530,1350 (NO2) cm~'. # (DMSO) 8.15 (2H, d, J = 9Hz, Ph-H), 7.55 (2H, d, J = 9Hz, Ph-H), 4.28 (1H, dd, JAx = 5Hz and JBX = 9Hz, CHx CHAHB), 3.2 (1H, dd, JxB = 9Hz and JAB = 18 Hz, CHX CHA HB), 2.75 (1H, dd, JXA = 5 Hz and JBA= 18 Hz, CHX CHAHB), 2.2-1. 0 (11H, m, N-cyclohexyl).

1-Cyclohexyl-3-(4'-aminophenyl)pyrrolidine-2,5-dione(1) A mixture of 1-cyclohexyl-3- (4'-nitrophenyl) pyrrolidine-2,5-dione (14) (1.2 g; 0.0039 mol) and palladium charcoal catalyst (10%, 0.12 g) in absolute ethanol (50 ml) was shaken with hydrogen gas at atmospheric pressure until the uptake of hydrogen was complete. The charcoal was then filtered off (No 4 sinter with celite filter aid) and the solvent removed under reduced pressure to leave 1-cyclohexyl-3- (4'-aminophenyl) pyrrolidine-2-, 5-dione (1) (1 g; 92.6%) as a brown viscous oil.

(Found C, 68.26; H, 7.62; N, 9.94. C16H2002N2+IH20 requires C, 68.30, H, 7.40, N, 9.94%). vmax 3460, 3370 (NH2) ; 2940,2860 (C-H); 1770,1700 (C=O), 1630 (Ph) cari'. 8 (DMSO) 6.95 (2H, d, J = 9Hz, Ph-H), 6.65 (2H, d, J = 9Hz, Ph-H), 3.9 (1H, dd, JAX x=5Hz and JBX = lOHz, CHXCHAHB), 3 18 (1H, dd, JXB = 9Hz and JAB = 18 Hz, CHX CHA HB) 2.65 (1H, dd, JXA = 5Hz and JBA = 18 Hz, CHx CHA HB), 1.8-1.2 (11H, m, N-cyclohexyl). l-Cyc!ohexyt-3-(4'-aminopheny!)pyrrolidine-2.5-dionehydrocMo ride(15) 1-Cyclohexyl-3-(4'-aminophenyl)(4'-aminophenyl) pyrrolidine-2,5-dione (1) (1.0 g) was dissolved in anhydrous ether and hydrogen chloride gas was passed through the solution. The yellow precipitate formed was filtered off and dried at the pump. Recrystallisation (ethanol) gave white crystals (0.5 g, 50%), m. p. 240-242°C.

(Found C 63.09; H 7.06; N, 8.99. C16H21O2N2Cl requires C, 62.23; H 6.85, N, 9.07%. vmax 3350 (NH3+); 2900 (C-H); 1770,1690 (C=O) cm-1. 8 (DMSO) 7.55 (2H, d, J = 9Hz, Ph-H), 7.30 (2H, d, J= 9Hz, Ph-_), 4.15 (1H, dd, J, = 5Hz and JBX = 9Hz, CH CHAHB), 3.25(1H, dd, JXB=9Hz and J. = 18Hz, CHXCHAHB), 2.73 (1H, dd, JXA = 5Hz and JBA = 18 Hz, CHXCHAHB), 2.1-1.1 (11H, m, N-cyclohexyl).