CA1336266C - Medicaments - Google Patents

Abstract

2-[4-(4-Chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphthoquinone and its physiologically acceptable salts or other physiologically acceptable derivatives are useful in the treatment or prophylaxis of Pneumocystis carinii infections in mammals.

Description

1~36~66 .

The present invention relates to the treatment and prophylaxis of PneumocYstis carinii infections. More particularly the invention is concerned with the use of 2-[4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphthoquinone and its physiologically acceptable salts or other physiologically acceptable functional derivatives in the treatment and prophylaxis of Pneumoc~stis carinii infections, the use of said compounds for the manufacture of medicaments for the treatment and prophylaxis of P. carinii infections, and novel formulations containing said compounds.

This Application is a division of Canadian Patent Application Serial No. 608,367, filed August 15, 1989.
Pneumocystis carinii is a parasite which has a natural habitat in lung tissue. In a host with a normal immune system P.carinii is not considered to be pathogenic. However, when the immune system is defective P.carinii is liable to cause pneumonia. There is a variety of circumstances in which the immune system may be defective or deficient.
Thus, for example immune system deficiency is common in immature or premature infants (neonates). It may also result from suppression by certain drugs, which may be deliberate e.g. in certain patients receiving organ transplants, or unavoidable e.g. as a side-effect of cancer chemotherapy. Disordered growth of one or more constituent parts of the immune system, e.g. as in certain forms of cancer, may also result in immunodeficiency.

Immune deficiency may furthermore be caused by viral infections, including human immunodeficiency virus (HIV). It has been reported (Hughes, W.T.
(1987) Treatment and Prophylaxis of Pneumocystis carinii pneumonia, Parasitology Today 3(11) 332-335) that at least 60% of patients with ~, .
acquired immunodeficiency syndrome (AIDS) suffer from Pneumocystis carinii pneumonia.

In this specification the term "immunocompromised host" will be used to describe hosts with a deficient or defective immune system.

Without treatment, Pneumocystis carinii pneumonia is almost always fatal in immunocompromised hosts. The most widely used treatments for this condition are trimethoprim-sulphamethoxazole (cotrimoxaole) and pentamidine. However, both of these treatments have been reported to be only around 50-70~ effective in AIDS patients and to produce a much higher than usual incidence of adverse reactions (about 50~) (Wofsy, C.B.
Antimicrobial Agents Annual, 1986, Vol 1, p377-400). There is thus a need for new agents, especially for the prophylaxis of P.carinii pneumonia.

A wide ran~e of naphthoquinones is known in the art. Such compounds have been variously described as having antl -l~rial, anticoccidial and antitheilerial activity. Some compounds have also been described as possessing activity against external parasites. Thus, Fieser et al, J.
Amer. Chem. Soc. 1948, 70, 3156-3165 (and references cited therein) describes a large number of 2-substituted-3-hydroxy-1,4-naphtho-quinones as having anti ~l~rial activity. A number of these compounds havealso been described in U.S. Patent Specification No. 2 553 648. Further classes of 2-substituted-3-hydroxy-1,4-naphthoquinones having acti~ity as anti~l~rial, anticoccidial and/or antitheilerial agents are described in U.S. Patents Nos. 3 367 830, and 3 347 742, U.K. Patent Specification No.
1553424, and European Patent Specifications Nos. 2 228, 77551 and 77550.

European Patent Application No. 123239 discloses synergistic combinations of anti-protozoal naphthoquinones and 4-pyridinols or alkanoic esters thereof, which are said to be especially useful for the treatment or prophylaxis of malaria.

European Patent No. 123,238 discloses 2-substituted-3-hydroxy-1,4-naphthoquinones of formula (I) (CH2)n ~ ~2 (I) ~ ~ OH

wherein either Rl is hydrogen and R2 is selected from Cl 6 alkoxy, aralkoxy,Cl 6 alkyl-Cl 6 alkoxy, phenyl substituted by one or two groups selected from halogen and Cl 6 alkyl, halogen and perhalo-Cl 6 alkyl or Rl and R are both Cl 6 alkyl or phenyl, and n is zero or 1, and physiologically acceptable salts thereof. Compounds of formula (I) wherein n is zero are said to be active against the human malaria parasite Plasmodium falciparum and also against Eimeria species such as E.tenella and E.acervulina, which are causitive ore~ni! c of coccidiosis. Compounds of formula (I) where n is 1 are said to be active against protozoa of the genus Theileria, in particular T.annulata and T.parva.

It has now been found that a 2-[4-(4-chlorophenyl)cyclohexyI~-3-hydroxy-1,4-naphthoquinone is active ln vivo against Pneumocystis carinii pneumonia infections in rats. Activity has also been demonstrated in an in vitro preparation of P. carinii.

In a first aspect the present invention provides such naphthoquinone or salt or derivatives thereof for use in the treatment and/or prophylaxis of Pneumocystis carinii infections (e.g. P. carinii pneumonia) in mammals (including humans).

In another aspect the present invention provides the use of such naphthoquinone or salt or derivative thereof for the manufacture of a medicament for the treatment and/or prophylaxis of Pneumocystis carinii infections in mammals (including humans).

According to a further aspect the present invention provides a method of treating and/or preventing Pneumocystis carinii infections in a m~mm~l which comprises administering to a mammal (including a human) susceptible to infection with P.
carinii pneumonia an effective amount of such naphthoquinone, or salt or derivative thereof.

In still another aspect of the invention there is provided a pharmaceutical composition containing such naphthoquinone or a salt or derivative thereof.

In yet another aspect the invention provides the use of such naphthoquinone or a salt or derivative thereof as an anti-Pneumoc~stis carinii agent.

Prevention of P. carinii infections is particularly important in an immunocompromised host, as discussed hereinabove. In the case of immunosuppression resulting from HIV infection, prophylaxis may be required by those diagnosed as seropositive for HIV, and those with progressive generalised lymphadenopathy (PGL) or AIDS-related complex (ARC) as well as patients suffering from AIDS.

Functional derivatives of the naphthoquinone may include those in which the 3-OH is replaced by a group oCOR5 , wherein R5 is a Cl_loalkyl group, a C3_10cycloalkyl group, a Cl_loalkoxy group, or a phenyl or naphthyl group, each such R5 group being optionally substituted, e.g., by amino, mono- or di-Cl_ 4alkylamino, carboxy or hydroxy;

a group oR6 or SR6, wherein R6 is an optionally substituted Cl_l0alkyl, C3_10cycloalkyl, phenyl or naphthyl group as defined for R5; or a group NR7R8, wherein R7 and R8 each independently represent hydrogen or Cl_4alkyl, or the group NR7R8 represents a 5-7 membered saturated heterocyclic ring, which may optionally contain a further heteroatom selected from nitrogen, oxygen or sulphur;

and physiologically acceptable salts and other physiologically functional derivatives thereof.

The 3-hydroxy compound of the invention may form salts with bases, derivatives in which the 3-hydroxy is replaced by a basic amino group may form salts with acids. Suitable base salts include inorganic base salts such as alkali metal (e.g.
sodium and potassium) salts and alkaline earth metal (e.g.
calcim) saltsi organic base salts e.g. phenylethylbenzylamine, dibenzylethylenediamine, ethanolamine and diethanolamine salts; and amino acid salts e.g. lysine and arginine.
Suitable acid addition salts include those formed from hydrochloric, hydrobromic, nitric, perchloric, sulphuric, citric, tartaric, phosphoric, lactic, glutamic, oxalic, aspartic, pyruvic, acetic, succinic, fumaric, maleic, oxaloacetic, isethionic, stearic, phthalic, methanesulphonic, p-tolune sulphonic, benzenesulphonic, lactobionic and glucuronic acids.

Without wishing to be bound by theory, it is believed that derivatives of the 3-hydroxy compound in which the 3-hydroxy group is replaced by a group - oCoR5, oR6, SR6 or NR7R8 mày act as pro-drugs or bioprecursors which are converted in vivo either by the host or the parasite to the 3-hydroxy compound.
Such compounds will be referred to hereinafter as "physiologically functional derivatives". Such compounds may also however possess intrinsic biological activity.

The invention includes within its scope the use of isomers of compounds of formula (II) and mixtures of such isomers. The 3-hydroxy compound may exist in a tautomeric form in which the hydroxyl group donates its proton to one of the oxo groups and the use of such tautomeric forms is included within the scope of this invention. However, it is believed that the stable form is the 3-hydroxy compound.

The compound 2-[4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1, 4-naphthoquinone is represented by formula (V):

Cl (V) ~ OH

and physiologically acceptable salts and other physiologically functional derivatives thereof.

Thus, in one aspect the present invention provides the compound of formula (V) and physiologically acceptable salts and other physiologically functional derivatives thereof for use in the treatment and/or prophylaxis of PneumocYstis carinii infections (e.g. P. carinii pneumonia) in mammals (including humans.

- 1336~66 In another aspect the present invention provides the use of the compound of formula (V) and physiologically acceptable salts and other physiologically functional derivatives thereof for the manufacture of a medicament for the treatment and/or prophylaxis of PneumocYstis carinii infections in mammals (including humans).

According to a further aspect the present invention provides a method of treating and/or preventing PneumocYstis carinii infections which comprise administering to a mammal (including humans) susceptible to infection with P. carinii pneumonia an effective amount of the compound of formula (V), or a physiologically acceptable salt or other physiologically functional derivative thereof.

It will be appreciated that the compound of formula (V) may exist as the cis or trans isomer, that is to say that the cyclohexyl ring may be cis or trans substituted by the naphthoquinone nucleus and the chlorophenyl substituent on the cyclohexyl ring. Both cis and trans isomers and mixtures thereof in any ratio may be used in accordance with the present invention. In general when the compound is in the form of a mixture of isomers the trans isomer will be present in an -amount of about 50% or will be the pre~: In~nt isomer but the use of mixtures in which the cis isomer predominates is also included within the scope of the invention. The specific ratio of isomers may be varied as required; typical mixtures include those in which the cis/trans i;somer ratio is about 1:1,40:60 and 5:98. For use according to the present invention the trans isomer of the compound of formula (V), or a mixture of its cis and trans isomers cont~inin~ at least 95~ e.g. 99% of the trans isomer, is preferred.

Physiologically functional derivatives of the compound of formula (V) include those of formula (VI) ~ Cl i l (VI) wherein R and R each represent -O and the dotted line represents a double bond between the 2 and 3 positions of the quinone ring, in which case R13 represents a group -OCOR ; a group OR or SR ; or a group NR R , wherein R , R , R and R are as hereinbefore defined; or the dotted line represents double bonds at the 1,2 and 3,4 positions of the quinol ring and R , R and R each represents a group -OCORl , wherein R14 represents an optionally substituted Cl_lOalkyl group.

lo- 1336266 Compounds of formula (VI) are believed to be novel and form a further aspect of the present invention.

Compounds of formula (VI) have been found to exhibit activity in vitro against the parasite Plasmodium falclparum and in vivo against the parasite Plasmodium yoelii as illustrated hereinafter. These compounds may therefore be useful in the treatment and/or prophylaxis of malaria.

A preferred compound of formula (VI) is 2-acetoxy-3-[trans-4-(4-chloro-phenyl) cyclohexyl]-1,4-naphthoquinone. This compound has the advanta~e of improved water-solubility as compared with the parent compound of formula (V) .

A further preferred compound of formula (VI) is 2-[trans-4-(4-chlorophenyl) cyclohexyl]-1,3,4-triacetoxynaphthalene. This compound is colourless, in contrast to the compound (V), which is yellow, and may therefore have advantages in terms of its formulation and presentation.

Further derivatives of the compound (V) which may be used in accordance with the present invention are those of formula (VII) ~ Cl tVII) wherein X is a halogen atom, e.g. a chlorine, bromine or iodine atom, preferably a chlorine atom.

The compound of formula (VII) wherein X is chlorine has previously been described as an intermediate e.g. in the preparation of the compound of formula (I) but no biological activity has been ascribed to it. In a further aspect therefore the present invention provides a compound of formula (VII) for use as a medicament, e.g. an antiprotozoal agent, or a medicament for the treatment of Pneumocystis carinii infections.

The synthesis of compounds of formulae (I) to (VII) may be effected by methods already known and described in the chemical literature (for example the patent specifications listed hereinbefore) or by analogous methods. In particular novel compounds of formula (VI) may be prepared by the following methods which form a further aspect of this invention:

(a) reaction of a compound of formula (V) or (VII), with a compound serving to introduce the required group R13, and where appropriate the groups Rll d R12;

(b) reaction of a compound of formula (VIII):

(VIII) . .
wherein Rl is as defined above with a donor compound serving to lntroduce the 4-(4-chlorophenyl)cyclohexyl group.

With regard to process (a) compounds (VI) wherein R and R represent - O
and R13 represents a group oCOR5 may be prepared by esterification of the compound (V). Esterification may be effected in conventional manner using the appropriate acid R5CoOH or acid derivative e.g. an acid anhydride, acid chloride or an activated ester such as an alkylhaloformate e.g. an alkylchloroformate. To prepare a compound of formula (VI) wherein Rll, R
and R13 each represent a group-OCOR14,the esterification is carried out in the presence of a reducing agent, e.g. zinc.

Compounds of formula (VI) wherein R13 is a group oR6 or SR may be prepared from a compound (VII) wherein X is a halogen atom. Thus for example the group OR may be introduced by reaction with the appropriate alcohol, e.g.
methanol or ethanol in the presence of sodium, and the group SR6 may be introduced by reaction with the corresponding thiol, R SH.

Compounds of formula (VI) wherein R13 is -NR7R8 may be prepared by reduction of the corresponding compound wherein R13 is azido, e.g. using lithium aluminium hydride in tetrahydrofuran, followed where necessary and/or desired by alkylation of the resulting amino group. The azido compound may be prepared from a compound of formula (VII) wherein X is halogen, by reaction e.g. with sodium azide.

Compounds of formula (VII) may be prepared for example in an analogous manner to process (b) described below.

- 13 - 1336~66 With regard to process (b), a suitable donor compound is the corresponding cycloalkane carboxylic acid which may undergo oxidative decarboxylation.
For instance persulphate with a catalyst, such as silver ions, is convenient for the purpose, (c.f.Jacobson, N., et al., Annalen, 1972, 763, 135 and Acta Chem. Scand, 1973, 27, 3211). Conveniently ammonium persulphate can be used as the oxidising agent, and the catalyst is silver nitrate. Further details of this process are described in EPA 123238. The compound of formula (VIII) used as starting material may be prepared from the corresponding 3-halo compound using methods analogous to process (a).

Hereinafter naphthoquinones active against P.carinii, including compounds more particularly described by formulae(v) and (VI), and their physio-logically acceptable salts and other physiologically functional derivativeswill be referred to as the "naphthoquinone". It will be appreciated that the amount of the naphthoquinone required for use in the treatment or prophylaxis of P.carinii will depend inter alia on the activity of the particular compound, the route of A~' inistration, the age and weight of the patient and the severity of the condition being treated. In general, a suitable dose for A~;n;~tration to man for the treatment of P.carinii pneumonia may be in the range of O.lmg to 200mg per kilogram bodyweight per day, for example from lmg/kg to lOOmg/kg, particularly 10 to 50 mg/kg. For A~m;n;qtration by inhalation the dose may conveniently be in the range of 0.1 to 20 mg/kg/day, e.g. 0.5 to 10 mg/kg/day. It will be appreciated that for A~min;stration to neonates, lower doses may be required.

-For prophylactic treatment the naphthoquinone may also be given less frequently, e.g. as a single dose on alternate days, once or twice per week or once or twice per month. The dosage for prophylactic treatment will depend inter alia on the activity of the naphthoquinone, the frequency of administration, and, where a depot preparation or controlled release formulation is used the rate of release of the active ingredient. Thus for once-weekly ~inistration a suitable prophylactic dose could be in the range 0.05 to 100 mg/kg,e.g. 0.05 to 50 mg/kg particularly 5 to 50 mg/kg.

Suitable dosages of a compound of formula (VI) for the treatment or prophylaxis of malaria in man are also within the ranges given above for the treatment and prophylaxis of P.carinii ~neumonia.

For use according to the present invention the naphthoquinone is preferably presented as a pharmaceutical formulation.

ph~ ~ceutical formulations comprise the naphthoquinone or a physiologi-cally acceptable salt or other physiologically functional derivative thereof together with one or more pharmaceutically acceptable carriers therefor and optionally other therapeutic and/or prophylactic ingredients.
The carrier(s) must be acceptable in the sense of being compatible with the other ingredients of the formula and not deleterious to the recipient thereof.

The naphthoquinone may conveniently be presented as a pharmaceutical formulation in unit dosage form. A convenient unit dose formulation contains the naphthoquinone in an amount of from 10 mg to 3g e.g. 10 mg to lg.

Pharmaceutical formulations include those suitable for oral, topical(including dermal,buccal and sublingual),rectal or parenteral (including subcutaneous, intradermal, intramuscular and intravenous), nasal and pulmonary administration e.g. by inhalation. The formulation may, where appropriate, be conveniently presented in discrete dosage units and may be prepared by any of the methods well known in the art of pharmacy.
All methods include the step of bringing into association the naphthoqui-none with liquid carriers or finely divlded solid carriers or both andthen, if necessary, shaping the product into the desired formulation.

Pharmaceutical formulations suitable for oral A~;ni~tration wherein the carrier is a solid are most preferably presented as unit dose formulations such as boluses, capsules or tablets each cont~ining a predetermined amount of the naphthoquinone. A tablet may be made by compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable ~chin~ the naphthoquinone in a free-flowing form such as a powder or granules optionally mixed with a binder, lubricant, inert diluent, lubricating agent, surface-active agent or dispersing agent. Moulded tablets may be made by moulding an inert liquid diluent. Tablets may be optionally coated and, if uncoated, may optionally be scored. Capsules may be prepared by filling the naphthoquinone, either alone or in admixture with one or more accessory ingredients, into the capsule shells and then sealing them in the usual manner. Cachets are analogous to capsules wherein the naphthoquinone together with any accessory ingredient(s) is sealed in a rice paper envelope. The naphthoquinone compound may also be formulated as dispersible granules, which may for example be suspended in water before administration, or sprinkled on food. The granules may be packaged e.g. in a sachet. Formulations suitable for oral administration wherein the carrier is a liquid may be presented as a solution or a suspension in an aqueous liquid or a non-aqueous liquid, or as an oil-in-water liquid emulsion.

Formulations for oral administration include controlled release dosage forms e.g. tablets wherein the naphthoquinone is formulated in an appropriate release - controlling matrix, or is coated with a suitable release - controlling film. Such formulations may be particularly convenient for prophylactic use.

Pharmaceutical formulations suitable for rectal ~l' Inlqtration wherein the carrier is a solid are most preferably presented as unit dose suppositories. Suitable carriers include cocoa butter and other materials commonly used in the art. The suppositories may be conveniently formed by admixture of the naphthoquinone with the softened or melted carrier(s) followed by chilling and shaping in moulds.

Pharmaceutical formulations suitable for parenteral ~;ni qtration include sterile solutions or suspensions of the naphthoquinone in aqueous or oleaginous vehicles. Injectible preparations may be adapted for bolus in;ection or continuous infusion. Such preparations are conveniently presented in unit dose or multi-dose containers which are sealed after introduction of the formulation until required for use. Alternatively, the naphthoquinone may be in powder form which is constituted with a suitable vehicle, such as sterile, pyrogen-free water, before use.

The naphthoquinone may also be formulated as a long-acting depot prepara-tion, which may be administered by intramuscular injection or by implantation e.g. subcutaneously or intramuscularly.Depot preparations may include, for example, suitable polymeric or hydrophobic materials, or ion-e~chAnge resins. Such long-acting formulations are particularly convenient for prophylactic use.

Formulations suitable for pulmonary administration via the buccal cavity are presented such that particles con~;n;ng the naphthoquinone and desirably having a diameter in the range 0.5 to 7 microns are delivered into the bronchial tree of the recipient. Such formulations may be in the form of finely comminuted powders which may conveniently be presented in a pierceable capsule, for example of gelatin, for use in an inhalation device, or as a self-propelling formulation (also referred to as an aerosol formulation) comprising the naphthoquinone, a suitable liquid or gaseous propellant and optionally other ingredients such as a surfactant and/or a solid diluent. Suitable liquid propellants include propane and the chlorofluorocarbons, and suitable gaseous propellants include carbon dioxide. Suitable surfactants include sorbitan trioleate (which is available for example under the trademar~ "Arlacel 85 n ) ~ Polysorbate 20 and oleic acid. Self-propelling formulations may also be employed wherein the active ingredient is dispensed in the form of droplets of solution or - 18 ~

supension. The self-propelling formulation typically contains from 0.05 to 20mg/ml e.g. 0.1 to 5mg/ml of the active ingredient.

Such self-propelling formulations are analogous to those known in the art and may be prepared by established procedures. Suitably they are presented in a container provided with either a manually-operable or automatically functioning valve having the desired spray characteristics; advantageously the valve is of a metered type delivering a fixed volume, for example, 25 to 100 microlitres, upon each operation thereof.

As a further possibility the naphthoquinone may be in the form of a solution or suspension for use in an atomiser or nebuliser whereby an accelerated airstream or ultrasonic agitation is employed to produce a fine droplet mist for inhalation. Such solutions or suspensions may comprise, in addition to the naphthoquinone and solvent(s), optional ingredients such as surfactants. Suitable surfactants include those described above for self-propelling formulations. The solution or suspension typically contains from 0.05 to 20mg/ml e.g. 0.1 to 5mg/ml of the naphthoquinone.
When a suspension of the naphthoquinone is employed, this compound is preferably in finely divided form, e.g. in micronised form.

Formulations suitable for nasal ~' ini ~tration include presentations generally similar to those described above for pulmonary ~ njstration.
When dispensed such formulations should desirably have a particle diameter in the range 10 to 200 microns to enable retention in the nasal cavity;

this may be achieved by, as appropriate, use of a powder of a suitable particle size or choice of an appropriate valve. Other suitable formulations include coarse powders having a particle diameter in the range 20 to 500 microns, for administration by rapid inhalation through the nasal passage from a container held close up to the nose, and nasal drops comprising 0.2 to 5% w/v of the naphthoquinone in aqueous or oily solution or suspension.

It should be understood that in addition to the aforementioned carrier ingredients the pharmaceutical formulations for the various routes of administration described above may include, as appropriate one or more additional carrier ingredients such as diluents, buffers, flavouring agents, binders, surface active agents, thickeners, lubricants, preservatives (including anti-oxidants) and the like, and substances included for the purpose of rendering the formulation isotonic with the blood of the intended recipient.

None of the references listed hereinabove contains an invitation to ~minl.qter the compounds of formula (V) or (VI) by the nasal or pulmonary route or any suggestion that the said compounds, if ~. ini.~tered in such a manner, would be effective in the treatment of the conditions therein taught; the said disclosures likewise contain no description of any formulation suitable for administration by the nasal or pulmonary route.

Pharmaceutical formulations of the compounds of formula (V) or (VI) adapted for ~dmlnistration by the nasal or pulmonary route are believed to represent novel formulations and thus form a further feature of the present invention.

Novel compounds of formula (VI) may also be formulated in the manner described above for use in the treatment and/or prophylaxis of malaria and such formulations form a further aspect of the present invention.

The above naphthoquinones may also be used in accordance with the present invention in combination or concurrently with other therapeutic agents,'for example agents used in the treatment of immunocompromised patients, including anticancer agents such as interferons e.g. alpha-interferons;
antiviral agents such as azidothymidine (AZT,zidovudine), immunostimulants and immunodulators. The naphthoquinone may also be administered in combination with a 4-pyridinol compound, as described in EPA 123,239 e.g.
3,5-dichloro-2,6-dimethylpyridinol (meticlorpindol).

The following non-limiting examples illustrate inter alia the following aspects of the present invention:

the use of naphthoquinones in the treatment and prophylaxis of P,carinii infections;

novel pharmaceutical formulations;

novel compounds of formula (VI).

Example 1 - Preparation of compound (V) 2-~trans-4-(4-Chlorophenyl)cyclohexyll-3-hYdroxy-1 4-naphthoquinone a) 4-(4-Chlorophenyl)cyclohexane-l-carboxylic Acid Acetyl chloride (30g) and finely powdered aluminium chloride (60g) were stirred together in carbon disulphide (120 ml) and then cooled to -50C, in a C02/oxitol bath. Cyclohexene (30 g), previously cooled to -50 C, was added dropwise during 10 minutes while maint~ining the temperature of the reaction mixture at below -20C. The mixture was stirred at -50C for a further 60 minutes and the solvent then decanted to leave a gummy orange complex. A little chlorobenzene was added as the material warmed to ambient temperature; the re~inder of the chlorobenzene (total 300 ml) was then added, the so-obtained solution heated at 40C for 3 hours with stirring, poured onto a mixture of ice and concentrated hydrochloric acid and the organic layer separated, washed with 2M hydrochloric acid, 2M sodium hydroxide and water, dried over anhydrous sodium sulphate and evaporated to dryness. The product was distilled in vacuo, the fraction boiling at 140-154C (0.1 mm Hg) collected, diluted with an equal volume of petroleum ether (40-60), cooled to -6C and a continuous stream of nitrogen gss bubbled through, and the separated colourless solid recovered.

Bromine (2.8ml) was added to a solution of sodium hydroxide (6.2g) in water (42 ml) at 0C. The above-obtained substituted hexahydroacetophenone (3.lg) was dissolved in dioxan (15 ml) and the cold hypobromite solution then added, keeping the reaction mixture at below 20 C. The reaction mixture was stirred at ambient temperature for 6 hours then allowed to stand overnight. Sodium metabisulphite was added to destroy excess hypobromite, the mixture cooled and then acidified to give a colourless solid. The solid was filtered off, washed with water, dried and recrystallised from ethanol to give 4-(4-chlorophenyl)cyclohexane-1-carboxylic acid, m.p. 254-2S6C.

b) 2-~4-(4-chlorophenyl)cyclohexyll-3-chloro-1 4-naphthoquinone A mixture of 2-chloro-1,4-naphthoquinone (3.95g, 0.02 mol), 4-(4-chlorophenyl)cyclohexane-1-carboxylic acid (4.9g, 0.02 mol) and powdered silver nitrate (l.OSg, 0.0062 mol) was heated to reflux with vigorous stirring in 40 ml of acetonitrile. A solution of ammonium persulphate (12.0g, 0.0525 mol) in 50 ml of water was added dropwise over 1 hour. The mixture was refluxed for 3 hours then cooled in ice for 30 mins, after which it was filtered, and the residual sticky solid extracted twice with boiling chloroform to remove inorganic material. The chloroform was removed by evaporation to leave a yellow-brown solid (ca 2.7g). This was dissolved in 40 ml of boiling acetonitrile; a little insoluble material was removed by filtration.
On cooling, the title compound separated as yellow crystals, (550 mg) m.p. 172-175C.

- 23 ~

NMR, ~H(d6-DMS0) 8.05 (2H, mult., ~-naphth), 7.85(2H, mult., ~-naphth), 7.30 (4H, s., PhH), 3.30 (lH, br.t., CH), 2.67 (lH, b~`.t., CH), 1.2-2.4 (8H, mult., 4xCH2).

c) 2-~4-(4-chloro~henyl)cyclohexyll-3-hydroxy-1,4-naphthoguinone The product of stage (b) was suspended in 10 ml of boiling methanol and 0.55g of potassium hydroxide in 5.5 ml of water was added dropwise over 15 mins. The mixture was refluxed until a dark red solution formed, (after ca. 6 hrs) when 2 ml of concentrated hydrochloric acid was cautiously added dropwise. The mixture was cooled and filtered, and the solid residue washed thoroughly with water. The water washings were re-acidified and filtered. The combined solid residues (500 mg) mp 200-209 , were recrystallised from acetonitrile to give the title product as the trans-isomer (300 mg) m.p. 216-219C.

Example 2 2-Methoxy-3-[trans-4-(4-chlorophenyl)cyclohexyl]-1,4-naphthoquinone Sodium (0.3g, 0.013mol) was dissolved in 20ml of methanol and the compound of Example l(b) (1.5g, 0.004mol) was added. The mixture was warmed to reflux for 4 hours, then evaporated under reduced pressure. The dark red solid which remained was partitioned between water and chloroform. The chloroform layer was washed with ice cold dilute sodium hydroxide, followed by water and was then dried and evaporated to give a yellow solid (900mg).
This was recrystallised from acetonitrile to give the impure product (800mg) mpll7-120, which was further recrystallised from ethanol to give the title compound (600mg) mpl20-122 .

NMR, ~H (d6-DMSO) 8.0 (2H, mult., ~-naphth), 7.85 (2H, mult, ~-naphth) 7.35 (4H, s, PhH), 4.0 (3H, s, OMe), 3.1 (lH, br.t., CH), 2.6 (lH, br.t., CH), 1.5-2.2 (8H, m, 4xCH2).

Example 3 2-Amino-3-[trans-4-(4-chlorophenyl)cyclohexyl]-1,4-naphthoquinone a) 2-azido-3-[trans-4-(4-chlorophenyl)cyclohexyl]-1,4-naphthoquinone A solution of sodium azide (0.42g, 0.006mol) in 6ml of water was added to a suspension of the compound of Example l(b) (l.lg, 0.003mol) in 15ml of ethanol. The mixture was heated to reflux with stirring and then a further 15ml of ethanol and 6ml of water were added. Heating under reflux was continued for 4 hours followed by cooling in a refrigerator for 1 hour. The resulting yellow crystals were filtered off and washed with water and ethanol to give the impure title compound (0.9g) mpl30-135. This material was used in the next stage without further purification.

b) 2-Amino-3-[trans-4-(4-chlorophenyl)cyclohexyl]-1,4-naphthoquinone The impure product of stage (a) (0.9g) was dissolved in dry tetrahydrofuran (THF) and added dropwise to a suspension of lithium aluminium hydride (2.0g) in THF. The mixture was stirred at room temperature for 1 hour and then 2.Oml of water was added dropwise with caution. A current of air was passed through the mixture for 1 hour and then 0.7g of sodium hydroxide in 6ml of water was added. The mixture was filtered and washed with THF. The filtrate was evaporated to dryness leaving an amorphous orange material which was triturated with SVM whereupon orange crystals formed. These were filtered off, washed well with SVM and dried to yield a first crop of product (200mg) mp210-215. The reaction was repeated to give a further crop of product (300mg) mp200-210 . The two crops were combined and chromatographed, eluting with chloroform, to give the title compound (350mg) mp212-215 .

NMR, ~H (d6-DMSO) 8.0 (2H, mult., naphth) 7.8 (2H, mult., naphth), 7.35 (4H, q, PhH), 6.7 (2H, s, NH2), 2.6-3.0 (2H, mult., CH), 1.5-2.4 (8H, mult-, CH2).

Example 4 2-[trans-4-(4-Chlorophenyl)cyclohexyl]-3-(3-dimethylaminopropoxy)-1,4-naphthoquinone hydrochloride Sodium (O.lOg,4.5mmol) was dissolved in 3-dimethylamino-propan-1-ol (1.55g, 5eq) and cooled and the compound of Example l(b) (1.15g, 3mmol) was added. The mixture was stirred at room temperature for 1 hour, following which acetic acid (lml) was added and the mixture diluted with toluene - 1336266 ~
(30ml). The solution was washed with water (4 x 20ml) dried (MgS04) and evaporated in vacuo to a purple semi solid. This was taken up in acetone and a mixture of diethyl ether and hydrochloric acid added until the purple solution turned orange. The solution was evaporated in vacuo to dryness and washed several times with toluene to give a yellow/beige solid, which was recrystallised from methanol/toluene (1:99) to give the title compound as a yellow/green crystalline solid (0.92g) mpl91-194.

NMR, ~H (CDC13) 8.06 (2H, mult., ~-naphth), 7.74 (2H, mult., ~-naphth), 7.25 (4H, mult., PhH), 4.28 (2H, t., OCH2 ), 3.48 (2H, t., NCH2 ), 2.95 (6H, s., 2xMe), 3.17 (lH, br.t., CH), 2.67 (lH, br.t., CH), 2.54 (2H, mult., CH2) 1.4-2.3 (8H, mult., 4xCH2).

Example 5 2-Acetoxy-3-[trans-4-(4-chlorophenyl)cyclohexyl]-1,4-naphthoquinone The compound of Example l(c) (1.5g, 0.004mol) was suspended in acetic anhydride (3ml) and a few drops of concentrated sulphuric acid were added with vigorous stirring. A further quantity of acetic anhydride (3ml) was then added, the mixture stirred for 30 minutes and then poured into 15ml of water causing a vigorous reaction. The resulting mixture was cooled in ice and filtered to give pale yellow crystals which were washed with water and dried to give the impure product (1.6g) mpl49-152 . This was recrystallised from lOOml of SVM to give the title compound (1.3g) mpl58-160 .

NMR, ~H (d6-DMSO) 8.1 (2H, m, ~-naphth), 7.9 (2H, m, ~-naphth), 7.35 (4H, s, PhH), 3.1 (lH, br.t., CH), 2.6 (lH, br.t., CH), 2.5 (3H, s, COMe), 1.5-2.0 (8H, m, CH2).

Example 6 2-Ethoxycarbonyloxy-3-[trans-4-(4-chlorophenyl)cyclohexyl]-1,4 naphthoquinone The compound of Example l(c) (l.lg, 0.003mol) and pyridine (0.24g, 0.003mol) was stirred in 5ml of toluene and cooled in a water bath while 4ml of ethyl chloroformate was added dropwise over a period of 15 minutes.
The mixture was stirred for a further 30 minutes then poured into a mixture of ethyl acetate and water. The organic layer was separated, dried and evaporated to a yellow solid which was recrystallised from chloroform/petrol to give the impure product (850mg) mpl45-149 . This material was dissolved in chloroform, washed several times with ice cold O.lN sodium hydroxide and then water. The organic layer was dried and evaporated to give product (450mg) mpl47-9 . The reaction was repeated on the same scale, the product combined with the aforementioned material and then recrystallised from chloroform/petrol to give the title compound (1.3g) mpl53-155 .

NMR, ~H (d6-DMSO) 8.1 (2H, m, ~-naphth), 7.9 (2H, m, ~-naphth), 7.4 (4H, s, PhH), 4.3 (2H, q, OCH2), 3.1 (lH, br.t., CH), 2.6 (lH, br.t., CH), 1.5-2.0 (8H, m, CH2), 1.4 (3H, t, Me).

.

Example 7 1336266 2-[trans-4-(4-Chlorophenyl)cyclohexyl]-3-(4-dimethylaminobenzoyloxy)-1,4-naphthoquinone The compound of Example l(c) (2.0g, 5.4mmol) in dry toluene (50ml) cont~inln~ dry pyridine (0.44g, leq) was stirred at around room temperature. 4-Dimethylaminobenzoyl chloride (lg, leq) in dry toluene (25ml) was added dropwise over 15 minutes. The mixture was stirred at around room temperature for 1 hour, heated at reflux for 10 hours, left stAn~ine for 38 hours, and then refluxed for a further 7 hours. The mixture was then cooled, washed with water, sodium bicarbonate solution and again with water, dried (MgS04) and evaporated in vacuo to a yellow solid which was recrystallised from ethanol to give the title compound (1.25g) mpll7-121 (shrinks above 113).

NMR, ~H (CDC13) 8.1, 6.75 (4H, mult., ~-naphth + ~-Me2N-Ph), 7.72 (2H, mult., ~-naphth), 7.18 (4H, mult., PhH), 6.72 (2H, d, ~-Me2N-Ph), 3.18 (lH, br.t., CH), 3.14 (6H, s., 2xMe), 2.52 (lH, br.t., CH), 1.4-2.2 (8H, mult., 4xCH2 ) .

Example 8 2-[trans-4-(4-Chlorophenyl)cyclohexyl]-1,3,4-triacetoxynaphthalene The compound of Example l(c) (l.Og) and zinc dust (l.Og) was stirred at room temperature for 24 hours in acetic anhydride (6ml) with one drop TEA.

- 29 ~

The reaction mixture was filtered and added to water (50ml) and stirred for one hour. The resulting white precipitate was filtered, washed with water (4 x 20ml) and dried to give the title compound (0.4g) mpl77-179 .

NMR, ~H (d6-DMSO) 7.87 (2H, mult., ~-naphth), 7.63 (2H, mult., ~-naphth), 7.35 (4H, s., Ph.H), 2.95 (lH, br.t., CH), 2.64 (lH, br.t., CH), 2.62 ~3H, s., OAc), 2.49 (3H, s., OAc), 1.4-2.3 (8H, mult., 4xCH2).

Example 9 The following examples illustrate conventional pharmaceutical formulations which may be employed in accordance with the present invention:-A. Injectable solution A solution for intramuscular injection may be prepared by mixing:-Compound of Example 1 9.5 parts by weight Dimethyl sulphoxide 19.0 parts by weight Sorbitan monooleate 4.5 parts by weight Corn oil 67.0 parts by weight 100.O

B. Injectable solution Compound of Example 15 parts by weight N-methyl-pyrollidone48.3 parts by weight Tween ~0 2 parts by weight Span 804.7 parts by weight Miglyol 81240 parts by weight 100.O

C. Tablet Compound of Example 1- 25.0 mg Lactose BP 48.5 mg Microcrystalline Cellulose BP 10.0 mg ("Avicel*pH 101") Low-substituted Hydroxypropyl; 10.0 mg Cellulose BP ("LHPC LH-ll") Sodium Starch Glycollate BP 3.0 mg ("Explotab~') Povidone BP ("K30") 3.0 mg Magnesium Stearate BP0.5 mg 100.0 mg * ~e ~k ~ 31 -o~ Oral suspension Compound of Example 1 50 mg Avicel RC 591 75 mg Sucrose syrup 3.5 ml Methylhydroxybenzoate 5 mg Colour 0.01% w/v Cherry flavour 0.1 ~ v/v Tween*80 0.2 % v/v Water to 5 ml E. Injectable suspension Compound of Example 1 100 mg Polyvinyl pyrrolidone (PVP) 170 mg Tween*80 0.2~ v/v Methylhydroxybenzoate 0.1~ w/v Water for Injection to 3 ml F. Capsule Compound of Example 1 100 mg Starch 1500 150 mg Magnesium stearate 2.5 mg filled into a hard gelatin capsule * Trade Mark Example 10.

The following examples illustrate novel pharmaceutical formulations according to the present invention.

A. Suspensions for Nebulisation a) Compound of Example 1, sterile 1.0 mg Water for In;ections to 10.0 ml Disperse the naphthoquinone in the Water for Injections previously sterilised in a sterile container. Fill in to sterile glass ampoules, 10 ml/ampoule under aseptic conditions, and seal each ampoule by fusion of the glass.

b) The following suspension was prepared:

Compound of Example 1, micronised l.Og Polysorbate 20 0.1~ w/v Water for Injections to 10 ml The Polysorbate 20 was dispersed in the water for injections, followed by the compound of Example 1. This suspension was filled into sterile glass ampoules, lOml/ampoule under aspetic conditions, and the ampoules sealed by fusion of the glass.

B. Aerosol Formulations a) Compound of Example 1, micronised 1.0 mg Aerosol propellant to 5.0 ml Suspend the micronised naphthoquinone in the aerosol propellant.
Fill this suspension into preformed aerosol cannisters, 5 ml/cannister under pressure, through the valve orifice.

b) Compound of Example 1, micronised l.O mg Arlacel*85 0.1% w/v Aerosol propellant to 5 ml Disperse the Arlacel 85 in the aerosol propellant and then add compound of Example 1. Fill the suspension into preformed aerosol cannisters, 5ml/cannister under pressure, through the valve orifice.

C. Powder Inhalation Compound of Example 1, micronised l.O mg Lactose 29.0 mg Triturate and blend the micronised naphthoquinone with the lactose.
Fill the resulting powder blend into hard gelatin capsule shells, 30 mg per capsule.

* Trade Mark ~ 34 -D. Nasal Drops Compound of Example 1 100.0 mg Methylhydroxybenzoate 10.0 mg Water for Injections to10.0 ml Disperse the naphthoquinone and the methylhydroxybenzoate in the Water for Injections. Fill this suspension into suitable dropper bottles, 10 ml/bottle, and close by securing the dropper nozzle and bottle cap.

_ 1336266 BIOLOGICAL TEST RESULTS.

Example 11 Activity against Pneumocystis carinii Test Compounds A: 2-[trans-4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphthoquinone B: 2-[cis-4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphthoquinone C: 2-[4-(4-chlorophenyl)cyclohexyl]-3-chloro-1,4-naphthoquinone D: 2-(4-t-butylcyclohexyl)-3-hydroxy-1,4-naphthoquinone E: 2-(4-t-butylcyclohexylmethyl)-3-hydroxy-1,4-naphthoquinone a) Prophylaxis Groups of 10 rats were treated with dexamethasone to allow latent Pneumocystis carinii infection to develop. Tetracycline was also administered to protect against bacterial infections. Test compound A
was ~;ni.~tered, by gavage, from day 4 of the dexamethasone treatment, at a dose of 100 mg/kg/day. Two control groups of rats were treated with dexamethasone and tetracycline only. A further group of rats was given cotrimoxazole (trimethoprim +

sulphamethoxazole, 50 + 250 mg/kg/day, orally) in place of the test compound.

At the end of the test period the animals were sacrificed and autopsies carried out. The lungs were removed and the right lung bisected. An imprint was made onto microscope slides and stained with toluidine blue. One half of the lung was placed in formalin, processed in paraffin blocks, sectioned and stained by the Gomori methAn~in~ silver nitrate method.

After autopsy the extent of P.carinii pneumonitis was scored under coded study as none if no organism seen; l+ if P.carinii cysts seen sparsely distributed with less than one per 25 high power field (h.p.f.); 2+ if focal areas of P.carinii pneumonitis surrounded by 10 to 25 h.p.f. of normal lung and 3+ if lung diffusely and extensively involved with organisms in almost all h.p.f.s.

Results No of Early No of with No of rats with Rats deaths P.carinii PCP/
or Pneumonitis No of rats tested cannibal- None 1+ 2+ 3+
isation Test~Compound A 10 2* 8 0 0 0 0/8 Control (1) 10 2 0 1 2 5 8/8 Control (2) 10 0 0 0 2 8 10/10 * one early death, believed due to gavage, one cannibalisation.

b) Prophylaxis A further series of tests was carried out using the same general method as described above. Test compound A was administered at various dose levels, by gavage and in the diet.

The results are shown in Table 2.

~n o .,1 z o o o s ~ o o ~1 o ~. t~ a~ o ~D - "
P~, + ~.
~a~oooo~0o +
t) ~ OOOOO~1 ~100 ., I
~ ,c >t ~ +
~1 0 0 ~O O ~~1 0 :~ C ~

~ V O ~1 ~ ~ z 1 a ~ ~ , ~ ~ ~ n J ~ ~
~ ~ ~ D '~
a: h O I -~
E- O ~ L o a~ co O O O

~ ooool~7oooo o z u - ~ o x~ ~ ~~ ~ ~ ~ u _ _ o a ~D
~Oooooo.~mo~

~ ~ . .. .- - X

~ - 40 - 1 33S26 6 c) Treatment Groups of 10 rats were treated with dexamethasone and tetracycline.'for 4-6 weeks, as described in experiment (a) above. Three groups of rats were treated with Test compound A beginning after 4 weeks of immunosuppression, when Pneumocystis carinii pneumonia (PCP) had developed. Another group of rats in a parallel study was treated with Test compound A after 6 weeks of immunosuppression, when PCP infection was at an advanced stage. The results are shown in Table 3.

.~:Z; i ~ o o n o ~ o r~ I` o o~ ~~1 u o ~I ~

"
o ~ W ~ o o ~I ~ -.~ _ a +
.. ~ o ~ ~ ~ o ~I o u 1 _.

-~1 + a~
I o o o ~1~ ~1 ~1 ~1 O
~ C ~ Z
1,~ ~ a h Ul ui O oo ~ _Io oao ~o ~ ) ~

as~ qo ~
OD O a~ oI
h O ~1 0 ~
P~¦ ~ O Z ~
C V R
q~ r _ ~
O-- ~
~n o _ J ~ ~ u a) v o o o o o o u o ~ a~
:i5 '-ISl~ X
~r 3 ~ X - X

u X X X S~,~ a C _ ~ ~ t ~ ~ ._ ~ t o X ~ ~ -X ¦ R R ~ ~~ ~ ~ ~ ~ al oo o o un S~ o o ~ ~: ~ m o ~ ~ ~
O ~ f~ O ~--V 1~ CO ~ ~ C O

~ 1 0 ~d ~O O ~ R O

.,, ? ~
S~ r a aJ
X X
D ~D
i ~ ~
q~ ~
O O
u un _~ ,4 y~
- a a~
~D

_I h .l -' ~D D
~ ~ J

r ~D ~D
.. ~ ~

O ~ ~
c) n n a ~: ~
,_ ~
o o - U ~
~ n n -- ~D ~D

.,1 ,1 V D

X ~ ~
aJ
C:l d .4 _ - 43 ~

d) Treatment Groups of 15 rats were treated with dexamethasone and tetracycline for 4 weeks, as described in experiment (a) above. Test compounds (A) (E) were administered orally by stomach tube from the beginning of week 5 to the end of week 7.

In parallel with each test compound, Celacol was administered to one group of rats as a control. The results are given in Table 4.

. '~
. .
:

:~' 133626~

SCORE NO. INFECTED/
Test GROUP 0 1 2 3 4 NO. EXAMINED INFECTED
Compound (Dose/kg/day) Celacol 1 0 1 2 6 9/10 90 A 50mg/kg 1 3 3 5 0 11/12 92 75mg/kg 2 5 2 1 2 10/12 83 100mg/kg 4 7 1 1 0 9/13 69 Celacol 0 8 7 0 0 12/15 100 A 25mg/kg 3 7 4 1 0 12/15 80 50mg/kg 1 6 4 2 0 12/13 92 lOOmg/kg 4 6 2 0 0 8/12 67 Celacol 0 8 7 0 0 15/15 100 B 25mg/kg 1 8 5 1 0 14/15 93 50mg/kg 3 6 4 0 2 12/15 80 100mg/kg 2 6 2 4 0 12/14 86 ~ 45 ~ 133 626 6 Celacol 0 8 7 0 0 15/15 100 C 25mg/kg 0 5 6 3 0 14/14 100 50mg/kg 0 8 5 2 0 15/15 100 lOOmg/k~ 4 6 4 1 0 11/15 73 Celacol 0 8 2 3 2 15/15 100 D 25mg/kg 0 7 7 1 0 15/15 100 50mg/kg 1 8 4 1 0 13/14 93 lOOmg/kg 3 8 3 0 1 12/15 80 Celacol 0 8 2 3 2 15/15 100 E 25mg/kg 1 3 2 4 4 13/14 93 50mg/kg 0 2 6 4 2 14/14 100 lOOmg/kg 1 4 2 6 0 12/13 92 Example 12 Antimalarial Activity of Compounds (VI) Test methods Activity against Plasmodium Falciparum in vitro The test method was modification of that described by Desjardins et al., Antimicrob. Agents and Chemotherapy, 1979, 16, 710-718. Compounds were dissolved in ethanol at a concentration of 4.8xlO 3M and dilutions down to lxlO M were made. The drug solutions were serially diluted using RPMI
1640 medium + 10% human plasma in microtitration plates. Parasitised and fresh red blood cells were added, together with G- H-hypoxanthine, in RPMI
1640 medium + 10% human plasma and the cultures incubated for 48 hours.
Cultures were then harvested, the particulate contents collected on a glass fibre filter paper and washed copiously with water. The filter papers were dried and the radioactivity measured usin~ a scintillation counter.
Infected untreated and uninfected untreated cultures were included as controls.

The results are shown in Table 5.

Activity against Plasmodium yoelii in vivo _ - 47 -The naphthoquinone was suspended in 0.25% (w/v~ celacol in water by milling for 16-24 hours at 26 C. The suspensions were subsequently serially diluted with 0.25% (w/v) celacol in water.

At time 0, 0.1 ml of a suspension of 5xlO P.yoelii-parasitised red blood cells/ml of phosphate saline were in~ected intravenously into 15-20 g mice through a tail vein. Groups of 5 mice per treatment were dosed orally at times 6, 22, 30, 46, 54, 70 and 78 hours with 0.2 ml of the drug suspension. The compound of Example 4 was also administered intravenously.
Tail-blood smears were taken at 96 hours, stained with Giemsa and the percentage of red blood cells infected determined and compared to untreated, infected controls. Percent inhibition was correllated with dose to provide ED50 values. The results are shown in Table 5 below.

~- 1336266 TABLE S - Antimalarial activity in vitro and in vivo Compound of In vitro In vivo Example No: 50(~ ) ED50 mg/kg.
oral i.v.

lC 0.002 0.03 2 0.36 5.8 3 1.14 1.26 4 0.059 0.61 2.08 0.68 0.068 0.12 6 0.080 0.09 7 0.22 0.12

Claims (28)

1. The use of 2-[4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphthoquinone or a physiologically acceptable salt thereof for the manufacture of a medicament for the treatment or prophylaxis of Pneumocystis carinii.

2. Use of 2-[trans-4-(4-chlorophenyl)cyclo-hexyl]3-hydroxy-1,4-naphthoquinone for the manufacture of a medicament for the treatment of Pneumocystis carinii infections in mammals.

3. Use of 2-[trans-4-(4-chlorophenyl)cyclo-hexyl]-3-hydroxy-1,4-naphthoquinone for the manu-facture of a medicament for the prophylaxis of Pneumo-cystis carinii infections in mammals.

4. A pharmaceutical formulation for the treat-ment and/or prophylaxis of Pneumocystis carinii infections for nasal administration which comprises 2-[4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphtho-quinone or a physiologically acceptable salt thereof, in association with a pharmaceutically acceptable carrier therefor.

5. A formulation according to claim 4, wherein the naphthoquinone or salt is in the form of the trans isomer or a mixture of cis and trans isomers containing at least 95% of the trans isomer.

6. A pharmaceutical formulation for the treat-ment and/or prophylaxis of Pneumocystis carinii infections for pulmonary administration which com-prises 2-[4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphthoquinone or a physiologically acceptable salt thereof, in association with a pharmaceutically acceptable carrier therefor.

7. A formulation according to claim 6, wherein the naphthoquinone or salt is in the form of the trans isomer or a mixture of cis and trans isomers containing at least 95% of the trans isomer.

8. A pharmaceutical composition for the treat-ment and/or prophylaxis of Pneumocystis carinii infections comprising 2-[4-(4-chlorophenyl)-cyclo-hexyl]-3-hydroxy-1,4-naphthoquinone together with a pharmaceutically acceptable carrier therefor, adapted for nasal administration.

9. A pharmaceutical composition for the treat-ment and/or prophylaxis of Pneumocystis carinii infections comprising 2-[4-(4-chlorophenyl)-cyclo-hexyl]-3-hydroxy-1,4-naphthoquinone, together with a pharmaceutically acceptable carrier therefor, adapted for pulmonary administration.

10. A pharmaceutical formulation for the treat-ment or prophylaxis of Pneumocystis carinii infections comprising a pharmaceutically effective and acceptable amount of 2-[4-(4-chlorophenyl)-cyclohexyl]-3-hydroxy-1,4-naphthoquinone in association with a pharma-ceutically acceptable carrier therefor.

11. A pharmaceutical formulation according to claim 10, in a form for nasal administration.

12. A pharmaceutical formulation according to claim 10, in a form for pulmonary administration.

13. A pharmaceutical composition for the treat-ment and/or prophylaxis of Pneumocystis carinii infections comprising 2-[trans-4-(4-chlorophenyl)-cyclohexyl]-3-hydroxy-1,4-naphthoquinone together with a pharmaceutically acceptable carrier therefor, adapted for nasal administration.

14. A pharmaceutical composition for the treat-ment and/or prophylaxis of Pneumocystis carinii infections comprising 2-[trans-4-(4-chlorophenyl)-cyclohexyl]-3-hydroxy-1,4-naphthoquinone together with a pharmaceutically acceptable carrier therefor, adapted for pulmonary administration.

15. A pharmaceutical formulation for the treat-ment or prophylaxis of Pneumocystis carinii infections comprising a pharmaceutically effective and acceptable amount of 2-[trans-4-(4-chlorophenyl)-cyclohexyl]-3-hydroxy-1,4-naphthoquinone in association with a pharmaceutically acceptable carrier therefor.

16. A pharmaceutical formulation according to claim 15, in a form for nasal administration.

17. A pharmaceutical formulation according to claim 15, in a form for pulmonary administration.

18. An anti-Pneumocystis carinii pharmaceutical composition comprising an effective anti-Pneumocystis carinii amount of 2-[4-(4-chlorophenyl)-cyclohexyl]-3-hydroxy-1,4-naphthoquinone or a physiologically acceptable salt thereof, in association with a pharmaceutically acceptable carrier.

19. A composition according to claim 18 and comprising said naphthoquinone.

20. A composition according to claim 18 comprising said salt.

21. A composition according to claim 20 wherein said salt is a physiologically acceptable salt of trans isomer of the napthoquinone.

22. An anti-Pneumocystis carinii pharmaceutical composition comprising an effective anti-Pneumocystis carinii amount of 2-[trans-4-(4-chlorophenyl)-cyclo-hexyl]-3-hydroxy-1,4-naphthoquinone, in association with a pharmaceutically acceptable carrier.

23. A composition according to claim 18, 19, 20, 21 or 22, in a form for nasal administration.

24. A composition according to claim 18, 19, 20, 21 or 22, in a form for pulmonary administration.

25. 2-[4-(4-chlorophenyl)cyclohexyl]-3-hydroxy-1,4-naphthoquinone or a physiologically acceptable salt thereof for use in the treatment or prophylaxis of Pneumocystis carinii infections in mammals.

26. 2-[Trans-4-(4-chlorophenyl)-cyclohexyl]-3-hydroxy-1,4-naphthoquinone, for use in the treatment or prophylaxis of Pneumocystis carinii infections.

27. Use of 2-[4-(4-chlorophenyl)-cyclo-hexyl]-3-hydroxy-1,4-naphthoquinone as an anti-Pneumocystis carinii agent.

28. Use according to claim 27, wherein the naphthoquinone is in the form of the trans isomer.