GB2365426A

GB2365426A - Bactericidal benzamide derivatives

Info

Publication number: GB2365426A
Application number: GB0018887A
Authority: GB
Inventors: Michael G Thomas; Nigel M Allanson
Original assignee: Pantherix Ltd
Current assignee: Pantherix Ltd
Priority date: 2000-08-01
Filing date: 2000-08-01
Publication date: 2002-02-20
Also published as: GB0018887D0

Abstract

Compounds useful for the treatment of bacterial infection, many of which are new, are of formulae 1 and 2 <EMI ID=1.1 HE=133 WI=143 LX=400 LY=657 TI=CF> <PC>wherein ring A is optionally substituted by one or more groups R1 independently chosen from H, C<SB>1-6</SB> alkyl, C<SB>2-6</SB> alkenyl, C<SB>1-4</SB> alkylaryl, C<SB>1-4</SB> alkylCO<SB>2</SB>R4, -C0<SB>2</SB>R4, -CONR4R5, -CN, <SB>-</SB>CF<SB>3</SB>, -S0<SB>2</SB>R4, phenyl, substituted phenyl, halogen, hydroxyl, C<SB>1-4</SB> alkoxy, -NH<SB>2</SB>, -NHR4, -NR4R5, NO<SB>2</SB> and -SR4; and R4 and R5 are independently H, C<SB>1-6</SB> alkyl, C<SB>2-6</SB> alkenyl, C<SB>1-4</SB> alkylaryl or C<SB>1-4</SB> alkylCO<SB>2</SB>C<SB>1-4</SB> alkyl;<BR> ```ring B is optionally substituted by one or more groups R2 independently defined as for R1; and<BR> ```Ring C is optionally substituted by one or more groups R3 independently defined as for R1. Subject to certain disclaimers, the compound of formulae 1 and 2 are also claimed.

Description

<Desc/Clms Page number 1> COMPOUNDS AND THEIR THERAPEUTIC USE Field of the invention This invention relates to compounds and their therapeutic use. Background of the invention Several chemical classes of compound are known that possess considerable antibacterial activity, and these have proven of immense value in the treatment of bacterial diseases and infection. They include, among others, the penicillins, the cephalosporins, the aminoglycoside antibiotics, vancomycin analogues and the sulfonamide drugs. This last class of compounds has bacteriostatic properties mediated by its action on the bacterial pathway to folic acid. Compounds of similar structure to those of the invention have been described before as antihelminthic or antitrichocephaliasis agents, and secondly, as PGE2 agonists or antagonists. However there are no reports of antibacterial activity. For this reason, the antibacterial property of such compounds could not have been predicted and is therefore novel and inventive.

SU 1794941 Al discloses the synthesis and antihelminthic properties of compound A

Med. Parazitol. Parazit. Bolezni (1), pg 28-30, 1997 discloses the synthesis and use of compound B for the treatment of the parasitic infections fasciolliasis and intestinal nernatodiasis in sheep.

Med. Parazitol. Parazit. Bolezni (6) pg 52-3, 1991 discloses preclinical trial data on compound C to treat opisthorchiasis and trichocephaliasis in animals

EP 0 947 500 Al discloses compound D as a member of a group of carboxylic acid compounds having PGE2 agonist or antagonist activity.

The carboxyl function on ring C apparently is essential for PGE2 activity. N-(2,4-Dichlorophenyl)-2-(4-chloro-3-nitrobenzenesulfonylamino)-5-chlorobenzamide is available from Specs & Biospecs N.V., Fleminglean, The Netherlands. No utility is ascribed to this compound.

Summary of the invention Compounds of the invention are nitrophenylsulfonamides of Formulae 1 and 2.

In Formulae 1 and 2, ring A may be substituted by one or more groups R1 which may be independently chosen from: H, C1-6 alkyl, C2-6 alkenyl, C1-4 alkylaryl, C1-4 alkyIC02R4, -C02R4, -CONR4R5, -CN, -CF3, - S02R4, phenyl, substituted phenyl, halogen, hydroxyl, C1-4 alkoxy, -NH2, -NHR4, -NR4R5, N02 and -SR4.

Wherein R4 and R5 are independently defined as: H, C1-6 alkyl, C2-6 alkenyl, C1-4 alkyiaryl or C1-4 alkyIC02Cl-4alkyl.

Ring B may be substituted by one or more groups R2 which may be independently selected from: H, C1-6 alkyl, C2.6 alkenyl, C1-4 alkylaryl, C14 alkyIC02R4, -C02R4, -CONR4R5, -CN, -CF3, - S02R4, phenyl, substituted phenyl, halogen, hydroxyl, C14 alkoxy, -NH2, -NHR4, -NR4R5, N02 and -SR4.

Wherein R4 and R5 are independently defined as: H, C1-6 alkyl, C2-6 alkenyl, C14 alkylaryl or C14 alkylC02C14alkyl.

Ring C may be substituted by one or more groups R3 which may be independently selected from: H, Cj-6 alkyl, C2.6 alkenyl, C14 alkylaryl, -CONR4R5, -CN, -CF3, -S02R4, phenyl, substituted phenyl, halogen, hydroxyl, C1-4 alkoxy, -NH2, -NHR4, -NR4R5, N02 and -SR4.

Wherein R4 and R5 are independently defined as: H, Cl-r, alkYl, C2-6 alkenyl, C14 alkylaryl Compounds of the invention include salts thereof,

Compounds of the invention, most or all of which are new, have therapeutic utility. The compounds are useful as active ingredients in medicines, for the treatment of bacterial infection in man and animals.

Description of the Invention Certain compounds of this invention are preferred. In particular, see the subclaims.

The term "Cl-6 alkyl" as used herein refers to straight and branched chain alkyl groups having up to 6 C atoms. Examples are methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl and tert- butyl. "Alkyl" may have the same meaning. "Halogen" means F, Cl, Br or 1. "Alkoxy" means C1.6 alkyl-O-. "Heterocyclyi" means a saturated, unsaturated or aromatic ring of 5 to 8 atoms containing one or more heteroatoms such as 0, S or N, and which may be bonded via any C or ring atom.

Compounds of the invention may be prepared by procedures that are generally known. A general synthetic procedure is shown in Scheme 1. The method comprises the reaction of a compound of formula 3 with a compound of formula 4 to give an intermediate of formula 5, wherein R1 and R2 are as previously defined, X is a halogen atom and R4 is a hydroxyl or a protecting group such as alkoxy or functionalised alkoxy. The reaction may be performed at a temperature of -800C to 2500C preferably in presence of a base and an inert organic solvent. Suitable reaction conditions are the use of 4-N,N-dimethylaminopyridine in pyridine or toluene at 600C or lithium hexamethy1disilazide in anhydrous tetrahydrofuran at OOC.

If R4 is a protecting group such as an alkoxy group, the next step is hydrolyse the ester to a carboxylic acid (R4 = hydroxyl) using known deprotection methods such as an aqueous solution of a base.

The intermediate of formula 5 is then reacted with an amine of formula 6 under dehydrating conditions to give the compounds of formulae 1 and 2. In formula 5, R4 is a hydroxyl group and in formula 6, R3 is as previously defined. The reaction can preferably be carried out in an inert organic solvent at a temperature of -20'C to 1200C, in the presence of a dehydrating agent such as HATU, HBTU, DIC or DCC. Alternatively, the R4 group of formula 5 can be converted from a hydroxyl group to a leaving group such as a halogen atom. Suitable reagents for this conversion include thionyl chloride or phosphorus oxychloride. The acid halide is then reacted with the amine of formula 6 in the presence of a base and solvent to give compounds of formulae I and 2.

Compounds of formula 1 a as defined in claim 3 may be synthesised from the corresponding compounds of formula 1 a wherein R1 = halogen, by heating them with an appropriate thiol, amine, alkoxide, hydroxide or potassium cyanide in an inert solvent and base at between 250C and 150()C.

Any other compounds of the invention may be prepared in analogous manner. Suitable starting materials are available or can be prepared by one of ordinary skill in the art. Compounds of formula 1, 1 a or 2 may contain one or more chiral centres and exist in optically active forms. When a compound of formula 1, 1 a or 2 or a salt thereof contains a single chiral centre (for example sec-butyl) it may exist in two enantiomeric forms. The present invention includes individual enantiomers and mixtures of these enantiomers. The enantiomers may be obtained by methods known to those skilled in the art. Such methods typically include resolution via formation of diastereomeric salts or complexes which may be separated, for example, by crystallisation; resolution via formation of diastereomeric derivatives or complexes which may be separated, for example, by crystallisation, gas-liquid or liquid chromatography; selective reaction with one enantiomer by reaction with an enantiomer- specific reagent, for example, enzymatic esterification, oxidation or reduction, followed by separation of the modified and unmodified enantiomers; or gas-liquid or liquid chromatography on a chiral support such as silica with a bound chiral ligand or in the presence of a chiral solvent. It will be appreciated that where the desired enantiomer is converted into another chemical entity by one of the separation processes described above, at least one further step will subsequently be required to liberate the desired enantiomeric form. Alternatively, specific enantiomers may be synthesised by asymmetric synthesis using optically active reagents, substrates, catalysts or solvents, or by converting one enantiomer into another by asymmetric transformation.

When a compound of formulae 1, 1 a or 2 or a salt thereof contains more than one chiral centre it may exist in diastereomeric forms. The diastereomeric pairs may be separated by methods known to those skilled in the art, for example, chromatography or crystallisation and the individual enantiomers within each pair may be separated as described above. The present invention includes each diastereomer of compounds of fon-nulae 1, 1 a or 2 and mixtures thereof.

Some compounds of formulae 1, 1 a or 2 may exist in the form of solvates, for example, hydrates, which also fall within the scope of the present invention.

The compounds of formulae 1, 1 a or 2 may form organic or inorganic salts, for example, the compounds of formulae 1, 1 a or 2 may form addition salts with inorganic or organic acids, e.g. hydrochloric acid, hydrobromic acid, furnaric acid, tartaric acid, citric acid, sulfuric acid, hydiodic acid, maleic acid acetic acid, succinic acid, benzoic acid, pamoic acid, palmitic acid, dodecanoic acid and acidic amino-acids such as glutamic acid. Such compounds of fon-nulae 1, 1 a or 2 may form base addition salts, for example, with alkali metal hydroxides e.g. sodium hydroxide, with amino-acids e.g. lysine or arginine or with organic bases e.g. meglumine. It will be appreciated that such salts, provided that they are pharmaceutically acceptable may be used in therapy in place of compounds of formulae 1, 1 a or 2. Such salts are prepared by reacting the compound of formulae 1, 1 a or 2 with a suitable acid or base in a conventional manner. Such salts may also exist in the form of solvates, for example, hydrates. The present invention includes each salt and any solvate thereof.

Certain compounds of formulae 1, 1 a or 2 or salts thereof may exist in more than one crystal form and the present invention includes each crystal form and mixtures thereof.

The present invention also provides pharmaceutical compositions containing a therapeutically effective amount of a compound of formulae 1, 1 a or 2 including pharrnaceutically acceptable salts thereof together with a pharmaceutically acceptable diluent or carrier As used hereinafter, the term "active compound" denotes a compound of forrnulae 1, 1 a or 2 including pharmaceutically acceptable salts thereof. In therapeutic use, the active compound may be administered orally, rectally, parenterally, topically, ocularly, aurally, nasally, intravaginally or to the buccal cavity, to give a local and/or systemic effect. Thus the therapeutic compositions of the present invention may take the form of any of the known pharmaceutical compositions for such methods of administration. The compositions may be formulated in a manner known to those skilled in the art so as to give a controlled release, for

example rapid release or sustained release, of the compounds of the present invention. Pharmaceutically acceptable carriers suitable for use in such compositions are well known in the art of pharmacy. The compositions of the invention may contain 0.1-99% by weight of active compound. The compositions of the invention are generally prepared in unit dosage form. Preferably the unit dosage of active ingredient is 1-500 mg. 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 there are known pharmaceutical forms for such administration, for example tablets, capsules, granules, syrups and aqueous or oily suspensions.

Tablets may be prepared from a mixture of the active compound with fillers such as lactose or calcium phosphate, disintegrating agents, for example maize starch, lubricating agents, for example magnesium stearate, binders for example microcrystalline cellulose or polyvinyl pyrrolidone and other optional ingredients known in the art to permit tableting the mixture by known methods. The tablets may, if desired, be coated using known methods and excipients which may include enteric coating using for example hydroxypropylmethylcellulose phthalate. 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 known methods and if desired, provided with enteric coatings in a known manner. The tablets and capsules may conveniently each contain 0.1 to 1000 mg (for example 10 mg, 50 mg, 100 mg, 200 mg, 400 mg, 600 mg, or 800 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 carboxymethy1cellu lose, and oily suspensions containing a compound of the present invention in a suitable vegetable oil, for example sunflower oil.

The active compound may be formulated into granules with or without additional excipients. The granules may be ingested directly by the patient or they may be added to a suitable liquid carrier (for example water) before ingestion. The granules may contain disintegrants (for example a pharmaceutically acceptable effervescent couple formed from an acid and a carbonate or bicarbonate salt) to facilitate dispersion in the liquid medium.

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 transdermally. 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 accelerate 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 hard fat, 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 or sterile solutions in a suitable solvent.

Compositions of the invention suitable for inhalation via the mouth and/or the nose are the known pharmaceutical forms for such administration, for example aerosols, nebulised solutions or powders. Metered dose systems, known to those skilled in the art, may be used. Compositions suitable for application to the buccal cavity include slow dissolving tablets, troches, chewing gum, gels, pastes, powders, mouthwashes or rinses.

The compounds of the present invention may also be administered by continuous infusion either from an external source, for example by intravenous infusion, or from a source of the compound placed within the body, internal sources include implanted reservoirs containing the compound to be infused which is continuously released for example by osmosis and implants which may be a) liquid such as an oily solution or suspension of the compound to be infused for example in the form of a very sparingly water-soluble derivative such as a dodecanoate salt or b) solid in the form of an implanted support for example of a synthetic resin of waxy material for the compound to be infused. The support may be a single body containing all the compound or a series of several bodies each containing part of the compound to be delivered.

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. The following Examples illustrate the invention.

1. 2-(41-chloro-31-nitrobenzene sulfonylamino)-benzoic acid

To anthranilic acid (343 mg, 2.5 mmol) in 1 M aqueous sodium carbonate (5 mL) was added 4-chloro-3-nitrobenzenesulfonyl chloride (507 mg, 2.5 mmol) and the mixture heated at 60 OC until the sulfonyl chloride dissolved and then stirred for a further 15 h at room temperature. The reaction mixture was poured into water and acidified with 2 N HCI(aq), The precipitate was filtered off and dried under vacuum in a desiccator to give 2-(4'-chloro-3'-nftrobenzene sulfonylamino)-benzoic acid as a white solid (461 mg, 52%).

1H nmr (40OMHZ) d6 DMSO 8 11.3 (1 H, br s, -S02NH), 8.52(l H, d), 8.10 (1 H, dd), 7.99 (1 H, d) 7.91 (1 H, dd), 7.57 (1 H, dd), 7.47 (1 H, d), 7.21 (2H, dd). m/z 355 (M-H) and 357 (M+H). 2. 2-(41-chloro-3'-nitrobenzene sulfonylamino)-5-chlorobenzoic acid

To a stirred solution of sulfonamide (1) (1.2 g, 3.4 mmol) in acetic acid (20 mL) at 70 OC was added sulfuryl chloride (0.7 mL, 8.7 mmol) and the reaction mixture was heated at 95 OC. After 2 h a second addition of sulfuryl chloride (0.7 mL, 8.7 mmol) was made. After a further 2 h the solution was cooled to room temperature and poured into water (100 mL). The resultant precipitate was collected and dried to give 2-(4'-chloro-3'-nitrobenzene sulfonylamino)-5- chlorobenzoic acid as a pale yellow solid (1.2 g, 3.1 mmol, 91 %).

'H nmr (40OMHz) d6 DIVISO 8 8.51 (1 H, d), 8.07 (1 H, dd) 7.97 (1 H, d), 7.85 (1 H, d), 7.63 (1 H, dd), 7.48 (1 H, d). m/z 389, 391, and 393 ( all M-H) 3. N-Phenyl 2441-chloro-3'-nitrobenzene sulfonylamino)-benzamide

Carboxylic acid (1) (100 mg, 0.28 mmol) was heated under reflux in thionyl chloride (5 mQ for 2 h before removal of the excess reagent by distillation. The residue was taken up in DMF (5 mQ and treated with a solution of aniline (26 jil-, 0.28 mmol) and triethylamine ((47 4L, 0.28 mmol) in DMF (5mQ. The solution was heated under reflux for 2 h then stirred for an additional 16 h at room temperature. The solvent was evaporated and the residue recrystallised from acetic acid-water. Column chromatography on silica, using CH2CI2 as an eluant, gave the desired compound as a pale yellow solid (24 mg, 0.056 mmol, 20%).

1H nmr (40OMHz) CDC13 8 8.08 (1 H, d) 7.95 (1 H, dd), 7.79 (1 H, dd), 7.63 (1 H, br s), 7.61 - 7.57 (3H, m), 7.51 - 7.40 (4H, m) 7.31 (1 H, m) 7.25 (1 H, m). m/z 430 and 432 (both M-H)" 4. N-(411-Chlorophenyl) 2-(41-chloro-3'-nitrobenzene sulfonylamino)-benzamide

Carboxylic acid (1) (100 mg, 0.28 mmol) was heated under reflux in thionyl chloride (5 mQ for 1 h before removal of the excess reagent by distillation. The residue was taken up in CH2CI2 (5 mQ and treated with 4-chloroaniline (36 mg, 0.28 mmol) and triethylamine ((47 #LL, 0.28 mmol). The solution was stirred for 16 h at room temperature. The solvent was evaporated and the residue was chromatographed on silica, eluant (9:1) CH2CI2 - petrol ether to give the desired compound as a solid (13 mg, 0.056 mmol, 20%).

1H nmr (40OMHz) CDC13 8 7.96 (1 H, dd) 7.87 (1 H, dd), 7.70 (1 H, dd), 7.52 - 7.42 (3H, m) 7.37-7.28 (4H, AB system), 7.22 (1 H, m). m/z 464, 466 and 468 ( all M-H)-.

S. N-Phenyl 2-(W-chloro-X-nitrobenzene sulfonylamino)-5-chlorobenzamide

Carboxylic acid (2) (100 mg, 0.26 mmol) was heated under reflux in thionyl chloride (5 mL) for 2 h before removal of the excess reagent by distillation. The residue was taken up in DMF (5 mQ and treated with a solution of aniline (23 #LL, 0.26 mmol) and triethylamine (39 #LL, 0.26 mmol) in DMF (5mL). The solution was heated at 65 OC for 2 h then stirred for an additional 16 h at room temperature. The solvent was evaporated and the residue was chromatographed on silica using CH2Cl2 to give the desired compound as a solid (40 mg, 0.086 mmol, 33%).

1 H nmr (40OMHz) CDC13 8 10.2 (1 H, S, S02NH), 8.03 (1 H, d) 7.85 (1 H, dd), 7.66 (1 H, d), 7.50 (1 H, br s), 7.46 - 7.32 (6H, m), 7.17 (1 H, rn). m/z 464, 466 and 468 ( all M-H)- 6. N-(411-Chlorophenyl) 2-(41-chloro-3'-nitrobenzene sulfonylarnino)-5- chlorobenzamide

Carboxylic acid (2) (1 g, 2.6 mmol) was heated under reflux in thionyl chloride (5 mQ for 1 h 30 min before removal of the excess reagent by distillation. The residue was taken up in (CH2CI)2 (5 mQ and treated with a solution of 4-chloroaniline (328 mg, 2.6 mmol) and triethylamine (390 gL, 2.6 mmol) in 1,2-dichloroethane (5 mQ. The solution was heated under reflux for 2 h then stirred for 16 h at room temperature. The solvent was evaporated

and the residue was chromatographed on silica using CH2CI2 to give the desired compound as a solid (600 mg, 46%) 1H nmr (40OMHz) CDC13 6 10-15 (1 H, s, S02NH) 8. 10 (1 H, d) 7.85 (1 H, dd), 7.65 (1 H, dd), 7.59 (1 H, br s) 7.46-7.44 (4H, m), 7.37 - 7.29 (41-1, m). m/z 500 and 502 ( both M-H)-.

7. N-(411-Chlorophenyl) 2-(41-dimethylamino-3-nitrobenzene sulfonylamino)-5- chlorobenzamide

A solution of compound (6) (200 mg, 0.41 mmol), N, N-diiso pro pylethylamine (190 l.LL, 1.10 mmol and glycine (93 mg, 1.23 mmol) in DMF (5 ml-) was heated under reflux at 120 OC for 22 h. The reaction mixture was diluted with CI-12CI2 (30 mQ and washed with H20 (3 x 30 mQ and 2N aqueous HCI (20 mQ then dried over Na2SO4 and evaporated to give a solid (220 mg). Chromatography on silica using 5% MeOH in CH2CI2 gave the desired product as a yellow solid (140 mg, 0.27mmol, 67%).

1 H nmr (40OMHz) CDC13 6 9.84 (1 H, S, S02NH) 7.95 (1 H, s) 7.72 (21-1, dd), 7.60 (11 H, s), 7.55 (1 H, d), 7.50 (1 H, dd), 7.39 (11 H, d), 6.83 (1 H, d), 2.85 (61-1, s, N(CH3)2). m/z 507 and 509 ( M- H)-. 531 (M+Na)+.

8. N-(211,411-Dichlorophenyl) 2-(41-chloro-31-nitrobenzene sulfonylamino)-S- chlorobenzamide

Purchased from SPECS & BioSPECS B.V., Fleminglaan, The Netherlands.

Procedure for determininci Minimum Inhibitorv Concentrations (MIC)s and Minimum Bactericidal Concentrations (MBC)s on bacteria Reference: NCCLS 1987. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically - 4t" edition, approved standard M7-A4, pl 1-14, NCCLS, Villanova, Pa. And NCCLS, 1987. Methods for determining bactericidal activity of antimicrobial agents. Proposed guideline. Standards 26-P. NCCLS, Villanova, Pa.

A Dilution of compounds Stocks of compounds were prepared to a concentration of 10 mM in DMSO. The solutions were diluted 1:5 in Mueller-Hinton Broth (MHB) for 2 mM.

B Preparation of 96-well Microtitre Plate 1 . Add 0.1 mL of Mueller-Hinton Broth (MHB) to each of the 96 wells.

2. Add 0.1 mL of test compound solution (at twice the desired highest concentration) to column 1 of the plate.

3. Mix then remove 0.1 mL and add to column 2 4. Repeat the process across the plate to column 11, then discard the final 0. 1 mL. Column 12 is the growth control with no antimicrobial agent.

5. Add 5 #Ll- of the inoculum (see below) to each well and incubate at 350C overnight.

C Preparation of the inoculum The inoculum is prepared by making a direct broth or saline suspension of isolated colonies from an 18-24 h blood agar plate. These are prepared as follows: I . Suspend 3-5 colonies in 5 mL MHB then vortex for 15 seconds.

2. Adjust the suspension to 0.5 McFarland turbidity standard (approximately 1-2 x 108 cfu/mL) 3. Dilute the above 1: 10 in broth or saline.

4. Add 5 #il- to each well and incubate overnight. D MIC colony counts These are done on inoculum suspensions periodically to ensure that the final inoculum concentration routinely approximates to 5 X 105 cfu/mL. 0.01 mL is removed from the growth control well immediately after inoculation and added to the 10 mL 0.9% saline. 0.1 mL of the suspension is then spread over an agar medium. After incubation, the presence of 50 colonies would indicate an inoculum density of 5 x 105 cfu/mL. E IVIBC method 1 . Mix tubes without visible growth 2. Subculture a defined volume from these tubes to a blood agar plate. 3. Incubate overnight at 350C 4. Count the number of colonies from the original inoculum. Determine a colony count that represents 0.01 % of the original inoculum (99.99% reduction).

5. Count colonies from MBC plates. Any number equal to or less than the determined colony count is considered as 99.99% kill or a bactericidal result.

Results of MIC and IVIBC determinations Compounds have shown activity against a well known Oxford laboratory strain of S. aureus [NCTC 6571 (ATCC 9144) Heatley N.G. 1944. Antibiotic assay and sensitivity test organism. Biochem J. 38,51]. Compound (6) also has shown against a wider range of gram positive bacteria including both laboratory strains and drug resistant clinical isolates.

Claims

CLAIMS 1 . Use of a compound for the manufacture of a medicament for the treatment of bacterial infection, wherein the compound is of formula 1

wherein ring A is optionally substituted by one or more groups R1 independently chosen from H, CI-6 alkyi, C2-6 alkenyl, CI-4 alkylaryl, CI-4 alkyIC02R4, -C02R4, -CONR4R5, -CN, - CF3, -S02R4, phenyl, substituted phenyl, halogen, hydroxyl, C1-4 alkoxy, -NH2, -NHR4, - NR4R5, N02 and -SR4; and R4 and R5 are independently H, C1.6 alkyl, C2-r, alkenyl, CI-4 alkylaryl or C14 alkylC02C1-4 alkyl; ring B is optionally substituted by one or more groups R2 independently defined as for R1; and Ring C is optionally substituted by one or more groups R3 independently defined as for R1.
2. Use of a compound for the manufacture of a medicament for the treatment of bacterial infection, wherein the compound is of formula 2

wherein rings A, B, C are each optionally substituted as defined in claim 1.
3. The use according to claim 1, wherein compound is of formula la

where R1 is -CI, -OH, OR4, NH2, NHR4 or NR4R5 in which R4 and R5 are as defined in claim 1; and R2 and R3 are independently selected from H, Cl, Br, F and methyl.
4. The use of claim 1, wherein the compound is N-Phenyl 2-(4-chloro-3-nitrobenzenesulfonylamino)benzamide; N-(4-Chlorophenyl) 2-(4-chloro-3-nftrobenzenesulfonylamino)benzamide; N-Phenyl 2-(4-chloro-3-nftrobenzenesulfonylamino)-5-chlorobenzamide; N-(4-Chlorophenyl) 2-(4-chloro-3-nftrobenzenesulfonylamino)-5-chlorobenzamide; or N-(4-Chlorophenyl) 2-(4-dimethylamino-3-nitrobenzenesulfonylamino)-5- chlorobenzamide; or N-(2,4-Dichlorophenyl)-2-(4-chloro-3-nitrobenzenesulfonylamino)-5-chlorobenzamide.
5. A compound of forrnula 1 as defined in claim 1, independent of use, excepting N-(4- chlorophenyl)-2-(4-chloro-3-nitrobenzenesulfonylamino)-5-chlorobenzamide, N-(4- chlorophenyl)-2-(4-chloro-3-nftrobenzenesulfonamino)-5-bromobenzamide and N-(2,5- dichlorophenyl)-2-(4-chloro-3-nitrobenzenesuffonylamino)-5-bromobenzamide.

<Desc/Clms Page number 11>
6. A compound of formula 2 as defined in claim 2, independent of use, excepting 4-[2-(4- nitrobenzenesulfonylamino)-5-chlorobenzamino]benzoic acid.
7. A compound according to claim 5 which is recited in claim 4.
8. A phan-naceutical composition comprising as an active ingredient a compound according to any of claims 5 to 7 together with a carrier or diluent.
9. Use of a compound according to any of claims 5 to 7 for the manufacture of a medicament for the treatment of a bacterial infection.
10. The use of any of claims Ito 4 and 9, wherein the infection is caused by a gram positive organism.
11. The use of claim 10, wherein the organism is Staphlococcus aureus.