The present invention relates to novel therapeutic agents, and in particular to [l]benzopyrano[4, 3 -_] - pyrazoles, to processes for their preparation, to pharmaceutical compositions containing them and to their therapeutic activity as imrunomodulatory agents.

European Patent Application 354693 relates to compounds of structure A

wherein R± represents hydrogen or together with R ₂ represents a bond; R ₂ together wi .h either one of R-^ and

R ₃ represents a bond; R3 together with either one of R ₂ and R ₄ represents a bond; R ₄ represents hydrogen or together with R3 represents a bond; R5 represents hydrogen or methyl; Rg represents hydrogen, halo, a C ₂ _g alkanoyl group, a C ₂ _ ₆ alkoxycarbonyl group, a C _j . alkylthio group, a C-^.g alkylsulphinyl group, a C _1-6 alkylsulphonyl group, carbamoyl, carboxy, or R ₅ and Rg together with the carbon atom to which they are attached represent a cyclopropyl group; R ₇ represents hydrogen, halo, trifluoromethyl, methoxy, a C _ alkyl group, a ^c l-6 alkyithio group or a C-^.g alkylsulphinyl group; Rg

represents hydrogen, halo or trifluoromethyl; g and R- _j _ _Q / which may be the same or different, each represent halo; or Rg represents hydrogen and R _T _Q represents hydrogen, halo, trifluoromethyl, hydroxy, nitro, a C ₂ _g alkanoyloxy group, a ^_g alkyl group or a C^_g alkoxy group, which have immunomodulatory activity.

The present invention provides novel compounds of formula I

wherein R^ represents hydrogen or methyl;

R ₂ represents hydrogen, C ₂ _g alkoxycarbonyl or C ₂ _g alkanoyl;

R ₃ represents hydrogen, halo, trifluoromethyl, alkyl, C- _j ^ alkoxy or SK)),^;

R ₄ represents hydrogen, halo, trifluoromethyl, . alkyl, Cτ__ alkoxy or SCO^Y- _j ^

or R and R ₄ , together with the carbon atoms to which they are attached, represent a fused benz ring which may be substituted by one or more groups selected from _ alkyl, C_- _β alkoxy, SCO^Y-^, halo or trifluoromethyl;

R ₅ represents hydrogen, halo, trifluoromethyl, C- _j __g alkyl, C^. alkoxy, hydroxy or C ₂ _g alkanoyloxy;

Y-_ represents C ₁ _g alkyl; m is 0, 1 or 2;

or pharmaceutically acceptable salts thereof, which have immunomodulatory activity.

Compounds of formula I may be considered to possess therapeutic advantages over compounds known in the art.

It will be understood that a group containing a chain of 3 or more carbon atoms may be straight or branched, for example propyl includes ri-propyl and isopropyl, and butyl includes ri-butyl, sec-butyl, isobutyl and tert-butyl. The length of the carbon chain is specified for certain substituents, for example C _g alkoxycarbonyl refers to an alkoxycarbonyl group having from two to six carbon atoms. The term "halo" covers fluoro, chloro or bro o.

In preferred compounds of formula I, R _] _ represents hydrogen.

Preferably R ₂ represents hydrogen, C ₂ _ ₄ alkanoyl

(for example acetyl δr propanoyl) or C ₂ _g alkoxycarbonyl

(for example methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl or butoxycarbonyl) . Most preferably R ₂ represents hydrogen, acetyl or C ₂ _ ₄ alkoxycarbonyl. Particularly preferred are compounds in which R ₂ represents hydrogen, methoxycarbonyl, ethoxycarbonyl or propoxycarbonyl (namely n_-propoxycarbonyl and isopropoxycarbonyl) , especially hydrogen or ethoxycarbonyl.

Suitably R3 and R ₄ , which may be the same or different, represent hydrogen, halo, trifluoromethyl,

^c _l -6 ^alkvl - preferably C _1-Λ alkyl (for example methyl, ethyl or propyl), C _] __ _ζ alkylthio (for example methyl hio or ethylthio) , ^c ι-6 alkylsulphinyl (for example methylsulphinyl or ethylsulphinyl) or C- _j __g alkoxy (for example methoxy, ethoxy or propoxy) .

In preferred compounds of formula I, R represents hydrogen, halo, trifluoromethyl or C- _j __ alkylthio. More preferably R3 represents hydrogen, fluoro, chloro, bromo or trifluoromethyl, especially hydrogen.

In preferred compounds of formula I, R ₄ represents hydrogen, halo, trifluoromethyl or C- _j __g alkylthio. More preferably R _Λ represents hydrogen, fluoro, chloro, bromo or trifluoromethyl, especially hydrogen or trifluoro¬ methyl, and most preferably trifluoromethyl.

In further preferred compounds of formula I, R3 and R ₄ , together with the carbon atoms to which they are attached, are joined to form a fused benz ring which may be substituted by one or more groups selected from Cτ__ ₄ alkyl (for example methyl, ethyl or propyl) , C- _j __ ₄ alkoxy (for example methoxy, ethoxy or propoxy) , halo or trifluoromethyl. Preferably the ring is unsubstituted or has a single substituent. The substitution may occur at any of positions 4-,5-,6- and/or 7- of the benzothiazole group.

The substituent R5 may be located at any position on the benz ring, namely in position 6-, 7-, 8- or 9- of the benz ring. Accordingly the substituents R5 specified herein are to be considered as named at each of these positions-

Suitably R5 represents hydrogen, fluoro, trifluoromethyl, hydroxy, C ₂ _g alkanoyloxy, C^.g alkyl (especially in position 8- or 9- of the benz ring) , or ^c l-6 ^l k°xy (especially in position 9- of the benz

ring) . As stated above R^ may be located at the 6- position, especially for the substituents 6-fluoro, 6- hydroxy or 6-methoxy. The 7-position may also bear a substituent such as 7-fluoro or 7-hydroxy. In preferred compounds of formula I, R5 is ocated at position 8- or 9- of the benz ring and represents hydrogen, fluoro, trifluoromethyl, hydroxy, C ₂ _g alkanoyloxy (for example acetoxy, propanoyloxy, butanoyloxy or pentanoyloxy) , C- _j __ ₄ alkyl (for example methyl, ethyl or propyl)- or C- _j __ ₄ alkoxy (for example methoxy or ethoxy) , more preferably hydrogen, fluoro, hydroxy or ₂ _ alkanoyloxy. Most preferably R5 represents hydrogen, fluoro, hydroxy or

-2-4 alkanoyloxy . In particularly preferred compounds

R 5 represents hydrogen, fluoro or C ₂ _ ₄ alkanoyloxy in position 8- of the benz ring or hydrogen, hydroxy or ^c 2-4 alkanoyloxy in position 9- of the benz ring.

Especially preferred are compounds in which R ₅ represents hydrogen, 8-fluoro, 9-hydroxy or 9-acetoxy.

Preferred classes of compounds of formula I are those represented by formula Ila

or lib

lib

wherein R-_, R ₂ , R3, R and R5 are as hereinbefore defined. In compounds of formula Ila, more preferably R5 represents hydrogen, halo (for example fluoro or chloro) , hydroxy or ^c 2-6 alkanoyloxy, especially hydrogen or fluoro. In compounds of formula lib, more preferably R5 represents hydrogen, hydroxy, -^_ alkoxy or C _g alkanoyloxy, especially hydrogen, hydroxy or ^c 2-4 alkanoyloxy.

In compounds of formula Ila and lib, Rτ_ preferably represents hydrogen and R ₂ preferably represents hydrogen or ^c 2-6 alkoxycarbonyl (for example methoxycarbonyl, ethoxycarbonyl or propoxycarbonyl) . In such compounds R preferably represents hydrogen and R ₄ preferably represents halo or trifluoromethyl. More preferably R represents hydrogen and R ₄ represents trifluoromethyl.

Further preferred classes of compounds are those represented by formula Ilia

or Illb

llb

wherein R _j , R ₂ and R5 are as hereinbefore defined, Rg represents i-6 alkyl, ^c ±-ξ _> alkoxy, halo or trifluoromethyl and q is 0, 1 or 2, more preferably 0 or 1, most preferably 0. In compounds of formula Ilia, preferably Rι_ represents hydrogen, R ₂ represents hydrogen or C ₂ _ alkoxycarbonyl and R5 represents hydrogen, hydroxy, fluoro or C ₂ _g alkanoyloxy and q is 0. In compounds of formula Illb, preferably R- _] _ represents hydrogen, R ₂ represents hydrogen or C ₂ _g alkoxycarbonyl and R5 represents hydrogen, hydroxy or C ₂ _g alkanoyloxy and q is 0. In compounds of formula Ilia, most preferably R ₅ represents hydrogen or fluoro. In compounds of formula Illb, most preferably R5 represents hydrogen, hydroxy or C ₂ _ alkanoyloxy.

Particularly preferred compounds of formula I are the compounds listed in Table A, including hydrates or solvates.

Compounds of formula I may contain one or more chiral centres and exist in different optically active forms. When compounds of formula I contain one chiral centre the compounds exist in two enantiomeric forms and the present invention includes both enantiomers and mixtures of enantiomers. The enantiomers may be resolved by methods known to those skilled in the art, for example by formation of diastereoisomeric salts which may be separated, for example, by crystallisation; formation of diastereoisomeric derivatives or complexes which may be separated, for example, by crystallisation, gas-liquid or liquid chromatography; selective derivatisation of one enantiomer by reaction with an enantiomer-specific reagent, for example enzymatic oxidation or reduction; or gas-liquid or liquid chromatography in a chiral environment, for example on a chiral support, for example silica with a bound chiral ligand or in the presence of a chiral solvent. It will be appreciated that where the active moiety is

transformed by the separation procedures described above, a further step is required to convert the product to the active moiety. Alternatively, specific enantiomers may be synthesised by asymmetric synthesis using optically active reagents, substrates, catalysts or solvents, or by converting one enantiomer into the other by asymmetric transformation.

When compounds of formula I contain more _. than one chiral centre, the compounds may exist in diastereoisomeric forms. The present invention includes each diastereoisomer and mixtures of the diastereoisomers. The diastereoisomers may be separated by methods known to those skilled in the art, for example by crystallisation or liquid chromatography.

Certain compounds of formula I may exist in different tautomeric forms or as different geometric isomers.

Some compounds of formula I are bases and may form salts with inorganic or organic acids, for example hydrochloric acid or hydrobromic acid. Other compounds of formula I are acidic and may form salts with inorganic or organic bases, for example sodium hydroxide. It will be appreciated that such salts, provided they are pharmaceutically acceptable, may be used in therapy in place of the corresponding compounds of formula I. Such salts may be prepared for example by reacting the compound of formula I with a suitable acid or base in a conventional manner.

Certain compounds of formula I may exist in more than one crystal form and the present invention includes each crystal form and mixtures thereof.

Certain compounds of formula I may also exist in the form of solvates, for example hydrates, and the

present invention includes each solvate and mixtures thereof.

The present invention also includes pharmaceutical compositions containing a therapeutically effective amount of a compound of formula I together with a pharmaceutically acceptable diluent or carrier.

As used hereinafter, the term "active compound" denotes a [l]benzopyrano[4,3-cJpyrazole of formula I. In therapeutic use, the active compound may be administered orally, rectally, parenterally or topically, preferably orally or topically. Thus the therapeutic compositions, of the present invention take the form of any of the known pharmaceutical compositions ^' for oral, rectal, parenteral or topical administration. Pharmaceutically acceptable carriers suitable for use in such compositions are well known in the art of pharmacy. The compositions of the invention may contain 0.1-90% by weight of active compound. The compositions of the invention are generally prepared in unit dosage form. The excipients used in the preparation of these compositions are the excipients known in the pharmacist's art.

Compositions for oral administration are preferred compositions of the invention and these are known pharmaceutical forms for such administration, for example tablets, capsules, syrups and aqueous or oily suspensions. Tablets may be prepared by mixing the active compound with an inert diluent such as lactose or calcium phosphate in the presence of disintegrating agents, for example maize starch, and lubricating agents, for example magnesium stearate, and tableting the mixture by known methods. The tablets may be formulated in a manner known to those skilled in the art so as to give a sustained release of the compounds of the present invention. Such tablets may, if desired, be

provided with enteric coatings by known methods, for example by the use of cellulose acetate phthalate. Similarly, capsules, for example hard or soft gelatin capsules, containing the active compound with or without added excipients, may be prepared by conventional means and, if desired, provided with enteric coatings in a known manner. The tablets and capsules may conveniently each contain 0.1 to 500 mg of the active compound. Other compositions for oral administration include, for example, aqueous suspensions containing the active compound in an aqueous medium in the presence of a non¬ toxic suspending agent such as sodium carboxymethy- lcellulose, and oily suspensions containing a compound of the present invention in a suitable vegetable oil, for example arachis oil.

Compositions for topical administration are also preferred compositions of the invention. The pharmaceutically active compound may be dispersed in a pharmaceutically acceptable cream, ointment or gel. A suitable cream may be prepared by incorporating the active compound in a topical vehicle such as petrolatum and/or light liquid paraffin, dispersed in an aqueous medium using surfactants. An ointment may be prepared by mixing the active compound with a topical vehicle such as a mineral oil, petrolatum and/or a wax e.g. paraffin wax or beeswax. A gel may be prepared by mixing the active compound with a topical vehicle comprising a gelling agent e.g. basified Carbomer BP, in the presence of water. Topically administrable compositions may also comprise a matrix in which the pharmaceutically active compounds of the present invention are dispersed so that the compounds are held in contact with the skin in order to administer the compounds transder ally. A suitable transdermal composition may be prepared by mixing the pharmaceutically active compound with a topical vehicle, such as described above, together with a potential

transdermal accelerant such as dimethyl sulphoxide or propylene glycol.

Compositions of the invention suitable for rectal administration are known pharmaceutical forms for such administration, for example suppositories with semi- synthetic glycerides or polyethylene glycol bases.

Compositions of the invention suitable for parenteral administration are known pharmaceutical forms for such administration, for example sterile suspensions in aqueous and oily media or sterile solutions in a suitable solvent.

In some formulations it may be beneficial to use the compounds of the present invention in the form of particles of very small size, for example as obtained by fluid energy milling.

In the compositions of the present invention the active compound may, if desired, be associated with other compatible pharmacologically active ingredients.

The compounds of formula I are indicated for- use as immunomodulatory agents, and are generally immunosuppressants, but some compounds, in certain disease states, may exhibit immunostimulant activity.

The compounds according to the invention are useful in the treatment of diseases resulting from an aberrant immune reaction. Thus the pharmaceutical compositions containing a therapeutically effective amount of a compound of formula I may be used to treat diseases with an immunological association for example tissue rejection, such as kidney rejection; autoimmune diseases, such as rheumatoid arthritis and systemic lupus erythematosus; cutaneous disorders, such as contact sensitivity, eczema and psoriasis; and neoplasia, such as melanoma.

In such treatment the amount of the compound of formula I administered per day will be such as to give a therapeutic effect and is generally in the range 0.1 to 2000 mg, preferably 1 to 500 mg.

Accordingly, in another aspect, the present invention also includes a method of treating diseases with an immunological association, comprising the administration of a therapeutically effective amount of a compound of formula I.

The therapeutic activity of compounds of formula I has been demonstrated by means of tests on standard laboratory animals. Such tests include, for example, the oral and parenteral administration of the compounds to BALB/c mice. Thus, compounds of formula I are useful as immunomodulatory agents. Whilst the precise amount of active compound administered will depend on 'a number of factors, for example the age of the patient, the severity of the condition and the past medical history and always lie ^' s within the sound discretion of the administering physician, a suitable dose for oral administration to mammals, including humans, is generally within the range 0.01-40 mg/kg/day, more usually 0.2-25 mg/kg/day given in single or divided doses. For parenteral administration, a suitable dose is generally withi ^' the range 0.001-4.0 mg/kg/day, more usually 0.005-1 mg/kg/day given in single or divided doses or by continuous infusion. A suitable preparation for topical administration generally contains the active ingredient within the range 0.01-20% by weight, more usually 0.05-5% by weight. Oral administration is preferred.

Processes for the preparation of compounds of formula I will now be described. These processes form a further aspect of the present invention.

Compounds of formula I may be prepared by oxidising compounds of formula IV

or tautomers thereof, for example by reaction with chloranil.

Compounds of formula I may also be prepared by reacting compounds of formula V

or tautomers thereof, with compounds of formula VI

ι ₂ HCR _g R- _Q V I

in which Rg represents (OQ) ₂ and R ₁₀ represents OQ or NQ' ₂ ; or Rg represents (SQ) ₂ and R _1Q represents SQ or NQ' ₂ ; or Rg represents =NH and R-J_Q represents OQ or SQ; or Rg represents =0 and R^g represents a leaving group for example an optionally substituted 1-imidazolyl group, in which Q and Q' represent C- _j __ ₄ alkyl or benzyl, for example by heating at 50-200°C.

Compounds of formula I in which R- _] _ represents hydrogen and R represents C ₂ _g alkanoyl may also be

prepared by reacting compounds of formula V, or tautomers thereof, with compounds of formula Vila

or tautomers thereof, in which R-^ represents C-^.ς alkyl, R ₁₂ and R^ may be the same or different, and each represent C- _j __g alkyl or benzyl, for example by heating in an organic liquid for example xylene at a temperature between 50-200°C.

Compounds of formula I in which R ₂ represents C ₂ _g alkanoyl may also be prepared by reacting compounds of formula V, or tautomers thereof, with compounds of formula Vllb

or tautomers thereo ^' f, in which R-J_-J_ represents C±- _$ alkyl, R- _j _ ₂ and R^ ^ma y be the same or different and each represents C^_ alkyl or benzyl, for example by heating in an organic liquid, for example xylene at a temperature between 50 and 250°C.

Compounds of formula I in which R5 represents C ₂ _g alkanoyloxy may also be prepared by acylating compounds of formula I in which R5 represents hydroxy by reaction with an acylating agent. The acylation reaction may be carried out by reacting the hydroxy compound of formula I with an acyl halide e.g. R ₁₄ C0C1 or an acid anhydride

(R- _[ _ ₄ CO) ₂ 0 in which Rη_ ₄ represents alkyl in the presence of a base at a temperature in the range -10°C to 40°C. The acylation reaction may also be carried out by reacting the compound of formula I with a carboxylic acid R ₁₄ COOH in the presence of a dehydrating agent, for example dicyclohexylcarbodiimide, preferably in the presence of a base e.g. pyridine. Compounds of formula I in which R^ represents hydroxy may be prepared by reacting compounds of formula I in which Rς represents _] __ alkoxy with a Lewis acid, for example aluminium chloride or boron tribromide.

Compounds of formula I may also be prepared by reacting compounds of formula VIII

in which R ₁₅ represents hydrogen, or tautomers thereof, or in which R ₁₅ represents a group COR ₁₇ wherein R ₁₇ represents hydrogen, optionally substituted C _1- alkyl or benzyl and R ₁₆ represents COCHR ₁ R ₂ , with a base e.g. piperidine in a suitable solvent e.g ethanol.

Compounds of formula I which are represented by formulae Ila/b and Illa/b may be prepared as described with reference to the preparation of compounds of formula I above.

Compounds of formula IV may be prepared by reducing compounds of formula I for example by reaction with sodium borohydride.

Compounds of formula IV may also be prepared by reacting compounds of formula IX

in which R- _j _ represents COOR g or carbamoyl and Rτ_ represents C- _j __ ₄ alkyl or benzyl with a hydrazine of formula X

for example, by heating at 50-250°C for example in acetic acid or in an inert organic liquid containing an acid catalyst, e.g. xylene containing p-toluene- sulphonic acid.

Compounds of formula V may be prepared by reacting compounds of formula XI

in which R ₂₀ represents hydrogen, with a hydrazine of formula X, for example by heating at 50-200°C in an organic liquid for example toluene. Preferably the

Compounds of formula V in which R^ represents hydroxy1 may also be prepared by reacting compounds of formula V in which R^ represents Cι__g alkoxy with a

Lewis acid, for example aluminium chloride or boron tribromide.

Compounds of formula VI in which R represents (0Q) and R-J_ _Q represents OQ may be prepared for example a) by reacting compounds of formula Rι_R CH-CX in which X is halo with a sodium alkoxide of formula NaOQ in which Q is CI _^ alkyl or benzyl, or b) by reacting compounds of formula R- _j _R ₂ CH-CN with an alcohol of formula QOH in the presence of an anhydrous acid, for example hydrogen chloride, to give compounds of formula R ₁ R CH-C(=NH)OQ as their acid salts, e.g. hydrochloride salts, which are then reacted with further alcohol of formula QOH.

Compounds of formula VI in which Rg represents (SQ) ₂ and R ₁₀ represents SQ may be prepared for example from compounds of formula Rτ_R ₂ CH-COCl by reaction with thiols of formula QSH in which Q represents C _1-4 alkyl or benzyl in the presence of a Lewis acid, for example zinc chloride.

Other compounds of formula VI and compounds of formula Vila may be prepared by methods known to those skilled in the art.

Compounds of formula Vllb or tautomers thereof may be prepared by the acylation of compounds of formula XII

by reaction with an acyl chloride R ₁₁ -C0C1, for example in the presence of pyridine in an inert solvent at a temperature in the range -10°C to 50°C.

Compounds of formula VIII in which R-^ represents OR ₁₇ and R- _j _g represents COCHR-^R;-) may be prepared by acylation of compounds of formula VIII in which R^ represents COR ₁₇ and R- _j _g represents hydrogen, for example by reaction with an acid anhydride of formula (R ₁ R ₂ CHCO) ₂ 0 or an acid halide e.g. of formula R-^CHCOCl.

Compounds of formula VIII in which R^c- represents COR- _j _ ₇ and R- _j^ represents hydrogen may be prepared by the acylation of compounds of formula V for example by reaction with an acid anhydride of formula (R^CO^O in the presence of a salt (e.g. the sodium salt) of the corresponding acid (e.g. at ambient temperature for 2.5 hours) .

Compounds of formula VIII in which R^g and R- _j _ are identical and represent COCHR- _j _R ₂ , may be prepared by acylation of compounds of formula V for example by using an acid anhydride of formula (R- _j jR ₂ CHCO) 0 in the presence of a salt (e.g. the sodium salt) of the corresponding acid (e.g. at ambient temperature for 19 hours or under reflux for 0.5 hours) .

Compounds of formula VIII in which R^g represents COCHR R2 and R- _] _5 represents hydrogen, or tautomers thereof, may be prepared by reacting a compound of

formula VIII in which R-^ _ζ represents COR ₁₇ and R- _j _g represents COCHR- _] ^ with a base e.g. piperidine in a suitable solvent e.g. ethanol.

Compounds of formula IX in which R-τ_g represents COOR- _| _g may be prepared by alkylating compounds of formula XIII

for example by reaction with lithium dialkyl copper.

Compounds of formula IX in which R^g represents carbamoyl may be prepared from compounds of formula IX in which R- _] _g represents cyano by methods known to those skilled in the art, for example by reacting with methanol saturated with hydrogen chloride and then water.

Compounds of formula X may be made by methods known to those skilled in the art. One method is the reaction of compounds of formula XIV

S //

B.NHNHC x IV

NH

in which B represents hydrogen or an acyl group for example C ₂ _g alkanoyl such as acetyl, with compounds of formula XV

0 II R ₄ CCHR ₃ X X V

in which X represents halo. The above reaction yields compounds of formula X as the hydrohalide salt, which may be basified to give compounds of formula X.

Compounds of formula XI may be prepared by reacting compounds of formula XVI

in which R ₂ - _j _ represents hydrogen with malonic acid in the presence of an acid chloride e.g. phosphoryl chloride and a Lewis acid e.g. zinc chloride.

Compounds of formula XI may also be prepared by reacting compounds of formula XVI in which R _2] _ represents a group COR ₂₂ in which R ₂₂ represents alkyl, with a base, for example sodium hydride, followed by treatment with a dialkyl carbonate of formula (QO) ₂ CO in which Q represents Cχ- alkyl or benzyl, e.g. dimethyl carbonate.

Compounds of formula XII to XVI may be prepared by methods known to those skilled in the art.

Certain intermediate compounds of formulae IV, V, VIII, X and salts thereof are believed to be novel compounds. All novel compounds herein are claimed as a further aspect of the invention.

The invention is illustrated by the following non- limitative Examples. In the Examples parts and percentages are by weight and compositions of mixed solvents are given by volume. Characterisation was by elemental analysis and one or more of the following spectroscopic techniques: nuclear magnetic resonance, infra-red and mass spectroscopy.

Example 1

a) A stirred mixture of 4-hydroxycoumarin (6.5 g) and 2-hydrazinobenzothiazole (10.0 g) in dry toluene (69 ml) was heated under reflux for 3.7 hours, with removal of the water formed in the reaction. The solid obtained on cooling was collected by filtration and digested into boiling industrial methylated spirit followed by hot filtration. On cooling the filtrate, the solid formed was collected by filtration to give l-(2- benzothiazolyl) -3- (2-hydroxyphenyl) -2-pyrazolin-5-one, m.p. 193-196°C. ^'

b) A stirred mixture of 1- (2-benzothiazolyl) -3- (2- hydroxyphenyl)-2-pyrazolin-5-one (1.0 g) and triethyl orthoacetate (1.7 ml) was heated at 130-135°C for 10 minutes. On cooling, ether was added and the solid formed was collected by filtration to give 2- (2- benzothiazolyl) -4-methyl [l]benzopyrano[4, 3-c_]pyrazol- 3(2H)-one, m.p. 239-241°C (with shrinkage at 223°C) .

Example 2

A stirred mixture of 1- (2-benzothiazolyl) -3- (2- hydroxyphenyl)-2-pyrazolin-5-one, (1.0 g) (Example la) and triethyl ortho(2-ethoxycarbonyl)acetate (2.3 g) was heated at 130-135°C for 15 minutes. On cooling, ether was added and the solid formed was collected by filtration to give ethyl 2- (2-benzothiazolyl)-3-oxo-2,3-

dihydro[l]benzopyrano[4,3-c]pyrazole-4-acetate, m.p. 185-189°C.

Example 3

a) A stirred mixture of 6-fluoro-4-hydroxycoumarin (10 g) and 2-hydrazinobenzothiazole (6.9 g) in dry toluene (100 ml) was heated under reflux for 0.5 hours, with removal of the water formed in the reaction. The solid obtained on cooling was collected by filtration, digested with dichloromethane and filtered. The filtrate was extracted with aqueous sodium hydroxide solution (5M) and the extract was acidified with dilute hydrochloric acid. The resulting solid was collected by filtration, washed with water and isopropanol, and recrystallised from ethyl acetate to give l-(2-benzo- thiazolyl) -3-(5-fluoro-2-hydroxyphenyl) -2-pyrazolin-5- one, m.p. 221-222°C.

b) A stirred mixture of 1-(2-benzothiazolyl) -3- (5- fluoro-2-hydroxyphenyl) -2-pyrazolin-5-one (0.8 g) and triethyl orthoacetate (7.2 ml) was heated at 140°C for 5 minutes. On cooling, the solid obtained was collected by filtration and washed with ether and then hot ethyl acetate. Recrystallisation from dichloromethane/ methanol gave 2- (2-benzothiazolyl) -8-fluoro-4- methyl[l]benzopyranot4,3-c pyrazol-3 (211)-one, m.p. 278- 280°C (with decomposition) .

Example 4

a) A stirred mixture of 4,5-dihydroxycoumarin (7.2 g) and 2-hydrazinobenzothiazole (10 g) in dry toluene (69 ml) and p-toluenesulphonic acid (0.2 g) was heated under reflux for 5.5 hours, with removal of the water formed in the reaction. The solid obtained on cooling was collected by filtration and digested with boiling industrial methylated spirit. After hot filtration, the

solid collected was partitioned between aqueous sodium hydroxide solution (5M) and ethyl acetate. The aqueous phase was acidified with concentrated hydrochloric acid and was digested with a mixture of boiling methanol and dichloromethane and the mixture hot filtered to give 1- (2-benzothiazolyl) -3- (2, 6-dihydroxyphenyl) -2-pyrazolin- 5-one, m.p. 264-266°C (with decomposition) .

b) A stirred mixture of 1- (2-benzothiazolyl) -3- (2, 6- dihydroxyphenyl) -2-pyrazolin-5-one (1.3 g) and triethyl orthoacetate (2.2 ml) was heated at 130-135°C for 15 minutes. A further portion of triethyl orthoacetate

(2.2 ml) was added to the reaction mixture and stirring was continued at 130-135°C for a further 15 minutes. On cooling, ether was added to the reaction mixture. After filtration the solid collected was suspended in boiling methanol and then dichloromethane was added. The mixture was heated under reflux and then hot filtered. The filtrate was concentrated and the solid product obtained was collected by filtration to give 2- (2- benzothiazolyl) -9-hydroxy-4-methyl [1]benzopyrano[4, 3- c.]pyrazol-3 (2H.) -one, m.p. 283-286°C (with decomposition) .

Example 5

a) A stirred mixture of 1-(2-benzothiazolyl)-3-(2, 6- dihydroxyphenyl)-2-pyrazolin-5-one (1.6 g) (Example 4a) in acetic anhydride (10 ml) was treated with sodium acetate (1.8 g) . The reaction mixture was stirred at ambient temperature for 16 hours. A further portion of sodium acetate (0.9 g) was added and stirring was continued for a further 24 hours. The reaction mixture was then added to water (9 volumes) /petroleum ether (1 volume; bp 60-80°C) and the solid product obtained on standing was collected by filtration to give l-(2- benzothiazolyl) -3- (2, 6-diacetoxyphenyl) pyrazol-5-yl acetate, m.p.- 159-162°C.

b) A stirred mixture of 1-(2-benzothiarolyl) -3-(2,6- diacetoxyphenyl)pyra ^* _oi-5-yl acetate (1.9 g) in ethanol

(8 ml) was treated with piperidine (0.4 ml) and heated under reflux for 3 minutes. On cooling, the solid collected by filtration was purified by chromatography on Florisil(R) using dichloromethane and then 5% methanol in dichloromethane as the mobile phase to give, after trituration with ether, 2-(2-benzothiazolyl) -4- methyl-3-oxo-2,3-dihydro[1]benzopyrano[4,3-<c]pyrazol-9- yl acetate, m.p. 246-251°C.

Example 6

a) A stirred mixture of 1-acetyithiosemicarbazide (5 g) , 3-bromo-l,l,l-trifluoropropan-2-one (7.2 g) and ethanol (50 ml) was heated under reflux for 3 hours. The mixture was evaporated to dryness, triturated with acetonitrile and filtered to give 4-trifluoromethyl-2- thiazolylhydrazine as the hydrobromide salt. The solid collected was dissolved in water and basified to pH 12 with aqueous ammonium hydroxide solution. After warming at 90°C to give a solution, followed by cooling, the solid obtained was 4-trifluoromethyl-2- thiazolylhydrazine, m.p. 118-119°C.

b) A mixture of 4-trifluoromethyl-2-thiazolyl¬ hydrazine (2.1 g) , p-toluenesulphonic acid (0.2 g) and 4-hydroxycoumarin (0.9 g) in xylene (50 ml) was heated under reflux for 6 hours, with removal of water formed in the reaction. After cooling and filtration, the filtrate was evaporated to dryness and the solid obtained was triturated with toluene. The mixture was then filtered and the solid collected was triturated with hydrochloric acid (5M) and washed with water to give 3- (2-hydroxyphenyl) -1- (4-trifluoromethyl-2- thiazolyl)-2-pyrazolin-5-one, m.p. 210-211°C.

c) A stirred mixture of 3- (2-hydroxyphenyl) -1- (4- trifluoromethyl-2-thiazolyl) -2-pyrazolin-5-one (3.0 g) and triethyl orthoacetate (10 ml) was heated at 140°C for 40 minutes. On cooling, the reaction was quenched with ethyl acetate and the mixture filtered. The solid product collected was 4-methyl-2- (4-trifluoromethyl-2- thiazolyl) [l]benzopyrano[4, 3-c_] -pyrazol-3 (2H) -one, m.p. 274-276°C.

Example 7

A stirred mixture of 3- (2-hydroxyphenyl) -1- (4- trifluoromethyl-2-thiazolyl) -2-pyrazolin-5-one (3.0 g) (Example 6b) and triethyl ortho(2-ethoxycarbonyl) - acetate (6.0 g) was heated at 140°C for 0.5 hours. On cooling, the reaction mixture was quenched with ethyl acetate and the solid formed was collected by filtration to give ethyl 3-oxo-2-(4-trifluoromethyl-2-thiazolyl) - 2,3-dihydro[1]benzopyrano[4,3 -_]pyrazole-4-acetate, m.p. 172-173°C.

Example 8

In the preparation of capsules, 10 parts by weight of active compound and 240 parts by weight of lactose are de-aggregated and blended. The mixture is filled into hard gelatin capsules, each capsule containing 10 mg active compound.

Example 9

In the preparation of capsules, 50 parts by weight of active compound, 300 parts by weight of lactose and 3 parts by weight of magnesium stearate are de-aggregated and blended. The mixture is filled into hard gelatin capsules, each capsule containing 50 mg of active ingredient.

Example 10

Tablets are prepared from the following ingredients.

Parts bv weight Active compound 10

Lactose 190

Maize starch 22.

Polyvinylpyrrolidone 10

Magnesium stearate 3

- The active compound, the lactose and some of the starch are de-aggregated, blended and the resulting mixture is granulated with a solution of the polyvinylpyrrolidone in ethanol. The dry granulate is blended with magnesium stearate and the rest of the starch. The mixture is then compressed in a tableting machine to give tablets containing:

a) 10 mg b) 100 mg c) 500 mg

of active compound.

Example 11

Tablets are prepared by the method of Example 10. The tablets are enteric coated in a conventional manner using a solution of 20% cellulose acetate phthalate and 3% diethyl phthalate in ethanol-.dichloromethane (1:1) .

Example 12

In the preparation of suppositories, 100 parts by weight of active compound is incorporated in 1300 parts by weight of semi-synthetic glycerides as the

suppository base and the mixture formed into suppositories each containing 100 mg Of active ingredient.

Example 13

In the preparation of ointments the active compound is incorporated into the base by thorough homogenization until the drug is evenly distributed. The ointment is packed into 10 g amber jars with screw-capped lined lids.

Active compound 0.1 g

White soft paraffin to 10 g

The compounds of the invention are immuno¬ modulatory agents, especially immunosuppressants and may show therapeutic activity at a dose of 200 mg/kg or lower. Preferred compounds of the invention show activity at 50 mg/kg or lower. The therapeutic activity of the preferred compounds of the present invention has been demonstrated by a cutaneous hypersensitivity test (CH test) in which the compounds are administered parenterally to BALB/c mice. This test was carried out in the following way.

Female BALB/c mice, weight range 16-24 g, were used in groups of eight. The abdomen of each mouse was shaved and 20 μl of a solution of a sensitising agent, 5% w/v 4-ethoxymethylene-2-phenyl-2-oxazolin-5-one (oxazolone) in acetonerolive oil (3:1 by volume), was applied to the shaved area. Immediately after sensitisation, the test compound in one of the dosages listed below was injected intraperitoneally as a suspension in 1.5% v/v sorbitan esters, under the trade name Tween 80, in sterile water (100 μl) . 100 μl of the same suspension was injected likewise every 24 hours for a further 7 days. The dosages used were selected from

the following values: 50, 30, 10, 3, 1, 0.3, 0.1, 0.03 or 0.01 mg/kg.

Two groups of at least eight BALB/c mice were used as a control simultaneously with each test in a similar manner to that described above except that no test compound was included in the daily injections.

On the seventh day after sensitisation, 10 μl of a solution of 1% w/v oxazolone in acetone: olive oil (3:1 by volume) was applied to one ear (the challenged ear) of each of the test mice and the control mice. (A more potent challenge dose of 1.5% w/v oxazolone in acetone:olive oil was employed in a few cases) . After 24 hours the thickness of the challenged ear and the thickness of the non-challenged ear of the same animal were measured with an engineer's screw gauge micrometer. The difference in thickness between the challenged ear and the non-challenged ear in each animal is a measure of the response of that animal to oxazolone. A comparison between the response of mice treated with the test compound and mice treated with the control indicates the effectiveness of the test compound as an immunomodulatory agent. The compounds were considered to be active at a particular dose if a 20% or greater reduction in ear swelling, which was statistically significant (p <0.05) according to Dunnett's test, between treated and control groups was obtained in at least two out of three CH tests, (or, where more than three tests have been carried out, a majority of the tests) at that dose (see for example Int. Arch. Allergy, 38, p246-259 (1970)) .

Each of the compounds of formula I illustrated in

Table A below was active at 50 mg/kg in at least two out of three tests at 50 mg/kg .unless indicated otherwise

(see Notes following the Table) . The minimum effective dose (i.e the lowest dose of those used at which the

compound is active as defined above) for each compound is given in Table A. The Example (Ex) number or numbers listed adjacent to each compound indicates the process or processes illustrating the preparation of that compound in the Examples.

Table A

Ex Compound Name Minimum

Effective

Dose

(mg/kg)

2-(2-benzothiazolyl) -4-methyl[1]benzo¬ 3 pyrano[4,3-cjpyrazol-3 (2H) -one

Ethyl 2- (2-ben∑othiazolyl) -3-oxo- 50 2,3-dihydro[1]ben∑opyrano[4,3-c.] - pyrazole-4-acetate

2- (2-benzothiazolyl) -8-fluoro-4- «_3 ^< methyl[1]benzopyrano[4,3-c_]pyrazol- 3(2H)-one

2-(2-benzothiazolyl) -9-hydroxy-4- ≤3- methyl[1]benzopyrano[4,3-c pyrazol- 3(2H)-one

2-(2-benzothiazolyl) -4-methyl-3-oxo- ≤3- 2,3-dihydro[1]benzopyrano[4,3-cj- pyrazol-9-yl acetate

4-methyl-2- (4-trifluoromethyl-2- ^3 thiazoly1) [1]benzopyrano[4,3-c - pyrazol-3 (2jH) -one

7 Ethyl 3-oxo-2-(4-trifluoromethyl-2- : thiazolyl) -2, 3-dihydro[1]benzopyrano- [4,3-c.]pyrazole-4-acetate

NOTES: * Active in each of 2 tests at 3mg/kg

The compounds of the present invention also show activity in a variety of other in-vivo screens, which show the utility of the compounds as immunomodulants, particularly in suppressing the immune response. Administration of the compounds has been carried out orally or parenterally. Some compounds have been found to be active in a test which determines their effects on humoral immunity by assaying the sera collected at the end of the oxazolone induced cutaneous hypersensitivity test described above (CH test) for changes in the amount of anti-oxazolone antibody produced, and a Graft versus Host test similar to that used by Smith S R, Terminelli C, Kipilman C T and Smith Y. , J. Immunopharmacology 1981;3(2),133-170.

For example, the compounds prepared in the following Examples were also found to be active in the above-described antibody test after parenteral administration at the doses described in Table A. A compound was deemed to be active if, at a dose of 50 mg/kg or lower, it caused a decrease in the relative serum anti-oxazolone antibody concentration determined by an enzyme linked immunosorbent assay (ELISA) by a factor of 0.5 or greater calculated by the following formula:-

O.D. (C _T ) - O.D. (T-, )

where O.D. (Cτ_) is the optical density of the control serum at a dilution of 1/128

O.D. (C ₂ ) is the optical density of the control serum at a dilution of 1/256

O.D. (T^) is the optical density of the test serum at a dilution of 1/128

The control and test sera were diluted with phosphate buffered saline (pH 7.3) containing 0.05% v/v Tween 20 (trade name) .

Activity of compounds in above test:

Table B

Minimum Effective

Examples Dose (mo/kσl

1 <:3 2 <3

2 <:3

4 <3

5 <:3

6 .3 7 «c3