GB2183630A - Cephalosporin antibiotics - Google Patents

Abstract

Cephalosporin antibiotics of the general formula (I> <IMAGE> wherein R<1> represents an isoquinolinium group or a 2,3-cyclopentenopyridinium group, and non-toxic salts and non-toxic metabolically labile esters thereof are disclosed. Processes for their preparation and pharmaceutical compositions containing them are also described.

Description

SPECIFICATION "Cephalosporin antibiotics" This invention relates to improvements in or relating to cephalosporins. More particularly it relates to new cephalosporin compounds and derivatives thereof having valuable antibiotic activity.

The cephalosporin compounds in this specification are named with reference to "cepham" after J. Amer.

Chem. Soc., 1962, 84, 3400, the term "cephem" referring to the basic cepham structure with one double bond.

Cephalosporin antibiotics are widely used in the treatment of diseases caused by pathogenic bacteria in human beings and animals, and are especially useful in the treatment of diseases caused by bacteria which are resistant to other antibiotics such as penicillin compounds, and in the treatment of penicillin-sensitive patients.

In many instances it is desirable to employ a cephalosporin antibiotic which exhibits activity against both Gram-positive and Gram-negative micro-organisms, and a significant amount of research has been directed to the development of various types of broad spectrum cephalosporin antibiotics.

Thus, for example, in our British Patent Specification No. 1399086, we describe a novel class of cephalosporin antibiotics containing a 7ss-(x-etherified oxyimino)-acylamido group, the oxyimino group having the syn configuration. This class of antibiotic compounds is characterised by high antibacterial activity against a range of Gram-positive and Gram-negative organisms coupled with particularly high stability to ss-lactamases produced by various Gram-negative organisms.

The discovery of this class of compounds has stimulated further research in the same area in attempts to find compounds which have improved properties, for example against particular classes of organisms, especially Gram-negative organisms. This interest is reflected in the very large numbers of patent applications which have been filed relating to cephalosporin antibiotics having particular oxyimino etherifying groups in combination with particular substituents both on the 7ss-acylamido side chain and at the 3-position of the cephalosporin nucleus.

In British Patent Specification No. 1604971 a wide variety of cephalosporin antibiotics are generally disclosed in which the 713-position side-chain may be selected from, inter alia, a 2-(2-aminothiazol-4-yl)-2-(etherified oxyimino)acetamido group, in which the etherifying group, amongst very many possible meanings, may be an alkyl group (e.g. methyl) substituted by a halogen atom, although there is no specific exemplification of compounds having such a group and the preferred etherifying group is stated to be an unsubstituted methyl group. Halogen atoms which, when present, are preferred are stated to be chlorine and bromine atoms.The 3-position group may also be selected from a very large number of alternatives and a possible 3-substituent is an optionally substituted isoquinoliniummethyl group, although again there is no specific exemplification of the preperation of compounds having such a 3-substituent.

European Patent Application No. 111935 generically defines cephalosporin compounds in which the 713-position side chain may be selected from, inter alia, a 2- (2-aminothiazol -4-yl) -2- (etherified oxyimino)acetamido group in which the etherifying group may be chosen from a large number of possibilities, including alkyl groups which may carry, inter alia, one or more halogen atoms. According to the generic definition, the 3-position group of the cephalosporin nucleus may inter alia be isoquinoliniummethyl. However, in the compounds specifically exemplified, only difluoromethyl and 2,2,2-trifluoroethyl groups are found as examples of haloalkyl oxime groups.

European Patent No. 135142 generically defines cephalosporins in which the 713-position side chain maybe selected from inter alia, a 2-(2-aminothiazol-4-yl)-2-(etherified oxyimino)acetamido group in which the etherifying group may be chosen from a large number of possibilities, including alkyl groups which may carry, inter alia, one or more halogen atoms. The monofluoromethyl group is mentioned by way of illustration but such groups do not feature in the specific examples. According to the generic definition, the 3-position group of the cephalosporin nucleus may, inter alia, be an isoquinoliniummethyl group or a cyclopentenopyridiniummethyl group.

We have now discovered that by the selection of a (Z)-2-(2-aminothiazol-4-yl)-2-(etherified oxyimino)acetamido group at the 713-position in combination with either an isoquinoliniummethyl or a 2,3-cyclopentenopyridiniummethyl group at the 3-position, and also by the selection of a monofluoromethoxyimino group as the etherified oxyimino grouping, cephalosporin compounds having a particularly advantageous profile of activity (described in more detail below) against a wide range of commonly encountered pathogenic organisms may be obtained.

Accordingly, we provide cephalosporin antibiotics of the general formula (I)

wherein R' represents an isoquinolinium group or a 2,3-cyclopentenopyridinium group, and non-toxic salts and non-toxic metabolically labile esters thereof.

The compounds according to the invention are syn isomers. The syn isomeric form is defined by the configuration of the -O.CH2F group with respect to the carboxamido group. In this specification, the syn configuration is denoted structurally as

It will be understood that since the compounds according to the invention are geometric isomers, some admixture with the corresponding anti isomer may occur.

The invention also includes within its scope the solvates (especially the hydrates) of the compounds of formula (I) and of their non-toxic salts. It also includes within its scope non-toxic salts and solvates of metabolically labile esters of the compounds of the formula (I). It will be appreciated that the solvates should be pharmacologically acceptable.

The compounds according to the present invention may exist in tautomeric forms (for example in respect of the 2-aminothiazolyl group) and it will be understood that such tautomeric forms, e.g. the 2-iminothiazolinyl form, are included within the scope of the invention.

The compounds according to the invention exhibit broad spectrum antibiotic activity both in vitro and in vivo.

They have high activity against both Gram-positive and Gram-negative organisms, including many ss-lactamase producing strains. The compound also possess high stability to ss-lactamases produced by a range of Gram-negative and Gram-positive organisms.

Compounds according to the invention have been found to exhibit high activity against strains (including penicillinase-producing strains) of Gram-positive bacteria such as Staphylococcus aureus, Staphylococcus epidermidis and Streptococcus species. This is coupled with high activity against various members of the Enterobacteriaceae (e.g. strains of Escherichia coli, Klebsiella pneumoniae, Citrobacter diversus, Enterobacter cloacae, Serratia marcescens, Proteus mfrabllis and indole-positive Proteus organisms such as Proteus vulgaris, Proteus morganii and Providence species), strains of Haemophllus influenzae, and Acinetobacter calcoaceticus as well as good activity against Pseudomonas species.This combination of high activity against Gram-positive organisms with high activity against Gram-negative organisms possessed by the compounds of the invention is unusual and particularly advantageous.

The compound of formula (I) according to the invention wherein R' represents an isoquinolinium group has been found to be particularly active in vivo (mouse protection test) against Escherichia coli.

The compound of formula (I) according to the invention wherein R' represents a 2,3-cyclopentenopyridinium group has been found to be particularly active in vitro against strains of Enterobacter cloacae.

Non-toxic salt derivatives which may be formed by reaction of the carboxyl group present in the compounds of formula (I) include inorganic base salts such as alkali metal salts (e.g. sodium and potasium salts) and alkaiine earth metal salts (e.g. calcium salts); amino acid salts (e.g. lysine and arginine salts); organic base salts (e.g.

procaine, phenylethylbenzylamine, dibenzylethylenediamine, ethanolamine, diethanolamine and N-methylglucosamine salts). Other non-toxic salt derivatives include acid addition salts, e.g. formed with hydrochloric, hydrobromic, sulphuric, nitric, phosphoric, formic and trifluoroacetic acids. The salts may also be in the form of resinates formed with, for example, a polystyrene resin or cross-linked polystyrene divinylbenzene copolymer resin containing amino or quaternary amino groups or sulphonic acid groups, or with a resin containing carboxyl groups, e.g. a polyacrylic acid resin. Soluble base salts (e.g. alkali metal salts such as the sodium salt) of the compounds of the formula (I) may be used in therapeutic applications because of the rapid distribution of such salts in the body upon administration. Where, however, insoluble salts of compounds (I) are desired in a particular application, e.g. for use in depot preparations, such salts may be formed in conventional manner, for example with appropriate organic amines.

Non-toxic metabolically labile ester derivatives which may be formed by esterification of the carboxyl group in the parent compound of formula (I) include acyloxyalkyl esters, e.g. lower alkanoyloxy-methyl or -ethyl esters such as acetoxy-methyl or -ethyl or pivaloyloxymethyl esters. In addition to the above ester derivatives, the present invention includes within its scope the compounds of formula (I) in the form of other physiologically acceptable equivalents, i.e. physiologically acceptable compounds which, like the metabolically labile esters, are converted in vivo into the parent antibiotic compound of formula (I).

These and other salt and ester derivatives such as the salts with toluene-p-sulphonic and methanesulphonic acids or the esters with t-butyl or diphenylmethyl esterifying groups may be employed as intermediates in the preparation and/or purification of the present compounds of formula (I), for example in the processes described below.

It will be appreciated that the compounds of the invention are usually present in the form of a betaine containing a positively-charged 3-substituent and a carboxylate group, and therefore esters and salts of compounds of formula (I) with bases will be associated with an anion A0 to balance the positive charge on the 3-substituent. Such an anion will also be non-toxic and may be derived from any of the acids described above which will form non-toxic salt derivatives.

The compounds of the invention may be used for treating a variety of diseases caused by pathogenic bacteria in human beings and animals, such as respiratory tract infections and urinary tract infections.

According to another embodiment of the invention we provide a process for the preparation of an antibiotic compound of general formula (I) as herein-before defined or a non-toxic salt or non-toxic metabolically labile ester thereof which comprises forming a compound of formula (I')

(wherein R' is as defined previously;B is -S- or -SO- (x- or ss-); R2 is an amino or protected amino group; and the dotted line bridging the 2-, 3- and 4-positions indicates that the compound is a ceph-2-em or ceph-3-em compound) or a salt thereof, or a corresponding compound having a group of formula -COOR3 at the 4-position [where R3 is a hydrogen atom or a carboxyl blocking group such as, for example, the residue of an ester-forming aliphatic or araliphatic alcohol or of an ester-forming phenol, silanol or stannanol (the said alcohol, phenol, silanol or stannanol preferably containing from 1 to 20 carbon atoms)] and having an associated anion E0 such as, for example a halide (e.g. chloride or bromide) or trifluoroacetate anion, by (A) acylating a compound of the formula (II)

(wherein R', B and the dotted line are as defined above) or a salt, e.g. an acid addition salt (formed with, for example, a mineral acid such as hydrochloric, hydrobromic, sulphuric, nitric or phosphoric acid or an organic acid such as methanesulphonic or toluene-p-sulphonic acid) or an N-silyl derivative thereof, or a corresponding compound having a group of formula -CO(YR3 at the 4-position (where R3 is as defined above) and having an associated anion EB as defined above with an acid of formula (III)

(Wherin R2 is as defined above) or a salt thereof or with an acylating agent corresponding thereto; or (B) reacting a compound of formual (IV)

(wherein R2, B and the dotted line are as hereinbefore defined; R3 represents hydrogen or a carboxyl blocking group; and X is a replaceable residue of a nucleophile, e.g. an acetoxy or dichloroacetoxy group or a chlorine, bromine or iodine atom) or a salt thereof, with isoquinoline or 2,3-cyclopentenopyridine; whereafter, if necessary and/or desired in each instance, any of the following reactions, in any appropriate sequence, are carried out:i) conversion of a A2-isomer into the desired A3-isomer, ii) reduction to the compound wherein B is -SO- to form a compound wherein B is -S-, iii) conversion of a carboxyl group into a non-toxic metabolically labile ester function, iv) formation of a non-toxic salt function, and v) removal of any carboxyl blocking and/or N-protecting groups.

The above reactions i) to v) may be carried out in conventional manner.

In the above-described process (A), the starting material of formula (Il) is preferably a compound wherein B is -S- and the dotted line represents a ceph-3-em compound.

Acylating agents which may be employed in the preparation of compounds of formula (I) include acid halides, particularly acid chlorides or bromides. Such acylating agents may be prepared by reacting an acid (III) or a salt thereof with a halogenating agent e.g. phosphorus pentachloride, thinoyl chloride or oxalyl chlpride.

Acylations employing acid halides may be effected in aqueous and non-aqueous reaction media, conveniently at temperatures of from -50 to + 50 C, preferably -40 to + 30 C, if desired in the presence of an acid binding agent. Suitable reaction media include aqueous ketones such as aqueous acetone, aqueous alcohols such as aqueous ethanol, esters such as ethyl acetate, halogenated hydrocarbons such as methylene chloride, amides such as dimethylacetamide, nitriles such as acetonitrile, or mixtures of two or more such solvents.Suitable acid binding agents include tertiary amines (e.g. triethylamine or dimethylaniline), inorganic bases (e.g. calcium carbonate or sodium bicarbonate), and oxiranes such as lower 1 ,2-alkylene oxides (e.g. ethylene oxide or propylene oxide) which bind hydrogen halide liberated in the acylation reaction.

Acids of formula (III) may themseleves be used as acylating agents in the preparation of compounds of formula (I). Acylations employing acids (III) are desirably conducted in the presence of a condensing agent, for example a carbodiimide such as N,N'-dicyclohexyicarbodiimide or N-ethyl-N'-y-dimethylaminoprnpylcar- bodiimide; a carbonyl compound such as carbonyldiimidazole; an isoxazolium salt such as N-ethyl-5phenylisoxazolium perchlorate; or N -ethoxycarbonyl-2-ethoxy-1, 2-dihydroquinoline.

Acylation may also be effected with other amide-forming derivatives of acids or formula (III) such as for example, an activated ester, a symmetrical an hydride or a mixed an hydride (e.g. formed with pivalic acid or with a haloformate, such as a lower alkylhaloformate). Mixed anhydrides amy also be formed with phosphorus acids (for example phosphoric or phosphorous acids), sulphuric acid or aliphatic or aromatic sulphonic acids (for example toluene-p-sulphonic acid). An activated ester may conveniently be formed in situ using, for example, 1 -hydroxybenzotriazole in the presence of a condensing agent as set out above. Alternatively, the activated ester may be preformed.

Acylation reactions involving the free acids or their above-mentioned amide-forming derivatives are desirably effected in an an hydros reaction medium, e.g. methylene chloride, tetrahydrofuran, dimethylformamide or acetonitrille.

An alterrnative method of activation is, for example, by reacting an acid of formula (III) with a solution or suspension preformed by adding a carbonyl halide, in particular oxalyl chloride or phosgene, or a phosphoryl halide such as phosphorus oxychloride to a solvent such as halogenated hydrocarbon, for example methylene chloride, containing a lower acyl tertiary amide such as N,N-dimethylformamide. The activated form of the the acid of formula (III) may then be reacted with a 7-amino compound of formula (Il) in a suitable solvent or mixture of solvents for example alcohols such as an alkanol, e.g. ethanol or industrial methylated spirits; halogenated hydrocarbons, e.g. dichloromethane; ethers, e.g. tetrahydrofuran or dioxan; esters, e.g. ethyl acetate; ketones, e.g. acetone; amides, e.g. N,N-dimethylacetarnide; acetonitrile; water and mixtures thereof. The acylation reaction may conveniently be effected at temperatures of from - 50"C to + 50 C, preferably - 40 to + 30"C, if desired in the presence of an acid binding agent, for example as described above (e.g. triethylamine, dimethylaniline or sodium bicarbonate).

If desired, the above acylation reactions amy be carried out in the presence of a catalyst such as 4-dimethylaminopyridine.

The acids of formula (III) and acylating agents corresponding thereto may, if desired, be prepared and employed in the form of their acid addition salts. Thus for example, acid chlorides may conveniently be employed as their hydrochloride salts, and acid bromides as their hydrobromide salts.

Isoquinoline or 2,3-cyclopentenopyridine may act as a nucleophile to displace a wide variety of substituents X from a cephalosporin of formula (IV). To some extent the facility of the displacement is related to the pK, of the acid HX from which the substituent is derived. Thus atoms or groups X derived from strong acids tend, in general, to be more easily displaced than atoms or groups derived from weaker acids.

The displacement of X by the nucleophile may conveniently be effected by maintaining the reactants in solution or suspension. The reaction is advantageously effected using from 1 to 10 moles of the nucleophile.

Nucleophilic displacement reactions may convieniently be carried out on those compounds of formula (IV) wherein the substituent X is a halogen atom or an acyloxy group, for example as discussed below.

,Acyloxy groups Compounds of formula (IV) wherein X is an acetoxy group are convenient starting materials for use in the nucleophilic displacement reaction. Alternative starting materials in this class include compounds of formula (IV) in which X is the residue of a substituted acetic acid e.g. chloroacetic acid, dichloroacetic acid and trifluoroacetic acid.

Displacement reactions on compounds (IV) possessing X substituents of this class, particularly in the case where X is an acetoxy group, may be facilitated by the presence in the reaction medium of iodide or thiocyanate ions. Reactions of this type are descibed in more detail in British Patent Specifications Nos. 1132621 and 1171603.

The substituent X may also be derived from formic acid, a haloformic acid such as chloroformic acid, or a carbamic acid.

When using a compound of formula (IV) in which X represents an acetoxy or substituted acetoxy group, it is generally desirable that the group R3in formual (IV) should be a hydrogen atom and that B should represent S. In this case, the reaction is advantageously effected in an aqueous medium, preferably at a pH of 5 to 8, particularly 5.5 to 7.

The above-descibed process employing compounds of formula (IV) in which X is the residue of a substituted acetic acid may be carried out as described in British Patent Specification No. 1241657.

When using compounds of formula (IV) in which X is an acetoxy group, the reaction is conveniently effected at a temperature of 30"C to 11 0 C, preferably 50"C to 80"C.

Halogens Compounds of formula (IV) in which X is a chlorine, bromine or iodine atom can also be conveniently used as starting materials in the nucleophilic displacement reaction. When using compounds of formula (IV) in this case, B may represent -SO- and R3 may represent a carboxyl blocking group. The reaction is conveniently effected in a non-aqueous medium which preferably comprises one or more organic solvents, advantageously of a polar nature such as ethers, e.g. dioxan or tetrahydrofuran; halogenated hydrocarbons, e.g. dichloromethane, esters, e.g. ethyl acetate; amides, e.g. formamide and N,N-dimethylformamide; and ketones e.g. acetone. In certain cases the nucleophile itself may be the solvent. Other suitable organic solvents are described in more detail in British Patent Specification No. 1326531.The reaction medium should be neither extremely acidic nor extremely basic. In the case of reactions carried out on compounds of formula (IV) in which R3 is a carboxyl blocking group the product will be formed as the corresponding halide salt which may, if desired, be subjected to one or more ion exchange reactions to obtain a salt having the desired anion.

When using compounds or formula (IV) in which X is a halogen atom as described above, the reaction is conveniently effected at a temperature of - 1 0 C to + 50"C, preferably + 1 0 C to + 30 C.

The reaction product may be separated from the reaction mixture, which may contain, for example, unchanged cephalosporin starting material and other substances, by a variety of processes including recrystallisation, ionophoresis, column chromatography and use of ion-exchangers (for example by chromatography on ion-exchange resins) or macroreticular resins.

A A2-cephalosporin ester derivative obtained in accordance with the process of the invention may be converted into the corresponding desired A3-derivative by, for example, treatment of the A2-ester with a base, such as pyridine or triethylamine.

A ceph-2-em reaction product may also be oxidised to yield the corresponding ceph-3-em oxide, for example by reaction with a peracid, e.g. peracetic or m-chloroperbenzoic acid; the resulting sulphoxide may subsequently be reduced as described hereinafter to yield the corresponding desired ceph-3-em sulphide.

Where a compound is obtained in which B is -SO- this may be converted into the corresponding sulphide by, for example, reduction of the corresponding acyloxysulphonium or alkoxysulphonium salt prepared in situ by reaction with e.g. acetyl chloride in the case of an acetoxysulphonium salt, reduction being effected by, for example, sodium dithionite or by iodide ion as in solution of potassium iodide in a solvent e.g. acetic acid, acetone, tetrahydrofuran, dioxan, dimethylformamide or dimethylacetamide. The reaction may be effected at a temperature of from -20 C to + 50"C.

Metabolically labile ester derivates of the compounds of formula (I) may be prepared by reacting a compound of formula (I) or a salt or protected derivative thereof with the appropriate esterifying agent such as an acyloxyalkyl halide (e.g. iodide) conveniently in an inert organic solvent such as dimethylformide or acetone, followed, where necessary, by removal of any protecting groups.

Base salts of the compounds of formula (I) may be formed by reacting an acid of formula (I) with an appropriate base. Thus, for example, sodium or potassium salts may be prepared using the respective 2-ethylhexanoate or hydrogen carbonate salt. Acid addition salts may be prepared by reacting a compound of formula (I) or a metabolically labile ester derivative thereof with the appropriate acid.

Where a compound of formula (I) is obtained as a mixture of isomers, the syn isomer may be obtained by, for example, conventional methods such as crystallisation or chromatography.

For use as starting materials for the preparation of compounds of general formula (I) according to the invention, compounds of general formula (III) and acid halides and anhydrides corresponding thereto in their syn isomeric form or in the form of mixtures of the syn isomers and the corresponding anti isomers containing at least 90% of the syn isomer are preferably used.

Acids of formula (III) and their derivatives may be prepared by etherification of a compound of formula (V)

(wherein R2 is as hereinbefore defined and R4 represents hydrogen Or a carboxyl blocking group) or a salt thereof, by selective reaction with a compound of general formula (VI) T.CH2F (Vl) (wherein T is chloro, bromo or iodo; sulphate; or sulphonate, such as tosylate), followed by removal of any carboxyl blocking group R4. Separation of isomers may be effected either before or after such etherification. The etherification reaction is conveniently carried out in the presence of a base, e.g. potassium carbonate or sodium hydride, and is preferably conducted in an organic solvent, for example dimethylsulphoxide, a cyclic ether such as tetrahydro-furan of dioxan, or an N,N-disubstituted amide such as dimethylformide.Under these conditions the configuration of the oxyimino group is substantially unchanged by the etherification reaction. When the compound of the formula (V) is employed in the form of a free acid or a salt with a base, the etherification reaction is generally carried out in the presence of a strong base, e.g., potassium t-butoxide, sufficient base being added to form a dianion. Furthermore, the reaction should be effected in the presence of a base if an acid addition salt of a compound of formula (V) is used the amount of the base being sufficient to netralise rapidly the acid in question.

Acids of formula (III) may also be prepared by reaction of a compound of formula (VII)

(wherein R2 and R4 are as hereinbefore defined) with a compound of formula (Vlil) H2N.O.CH2F (veil) followed by removal of any carboxyl blocking group R4, and where necessary the separation of syn and anti isomers.

The acids of formula (III) may be converted into the corresponding acid halides and anhydrides and acid addition salts by conventional methods, for example as described hereinabove.

Where X is a chlorine, bromine or iodine atom in formula (IV), ceph-3-em starting compounds may be prepared in conventional manner, e.g. by halogenation of a 7ss-protected amino-3-methylceph-3-em-4-carboxylic acid ester 113-oxide, removal of the 7p-protecting group, acylation of the resulting 7amino compound to form the desired 7ss-acylamido group, e.g. in an analogous manner to process (A) above, followed by reduction of the 113-oxide group later in the sequence. This is described in British Patent No. 1326531.The corresponding ceph-2-em compounds may be prepared by the method of Dutch published Patent Application No. 6902013 by reaction of a 3-methylceph-2-em compound with N-bromosuccinimide to yield the corresponding 3bromomethylceph-2-em compound.

Where X in formula (IV) is an acetoxy group, such starting materials may be prepared, for example, by acylation of 7-aminocephalosporanic acid, e.g. in an analogous manner to process (A) above. Compounds of formula (IV) in which X represents other acyloxy groups can be prepared by acylation of the corresponding 3-hydroxymethyl compounds which may be prepared for example by hydrolysis of the appropriate 3-acetoxymethyl compounds, e.g. as described for example in British Patent Specification Nos. 1474519 and 1531212.

The starting materials of formula (II) may also be prepared in conventional manner, for example, by nucleophilic displacement of the corresponding 3-acetoxymethyl compound with the appropriate nucleophile, e.g. as described in British Patent Specification No. 1028563, or by the method described in British Patent Specification No. 2052490A.

A further method for the preparation of the starting materials of formula (II) comprises deprotecting a corresponding protected 7amino compound in conventional manner, e.g. using PCI5.

It should be appreciated that in some of the above transformations it may be necessary to protect any sensitive groups in the molecule of the compound in question to avoid undesirable side reactions. Examples of suitable protecting groups are given in "Protective Groups in Organic Synthesis" by Theodora W. Greene (John Wiley and Sons, 1981). For example, during any of the reaction sequences referred to above it may be necessary to protect the NH2 group of the aminothiazolyl moiety, for example by tritylation, acylation (e.g. chloroacetylation or formylation), protonation or other conventional method.The protecting group may thereafter be removed in any convenient way which does not cause breakdown of the desired compound, e.g. in the case of a trityl group by using an optionally halogenated carboxylic acid, e.g. acetic acid, formic acid, chloroacetic acid or trifluoroacetic acid or using a mineral acid, e.g. hydrochloric acid or mixtures of such acids, preferably in the presence of a protic solvent such as water, or, in the case of a chloroacetyl group, by treatment with thiourea.

Carboxyl blocking groups used in the preparation of compounds of formula (I) or in the preparation of neecessary starting materials are desirably groups which may readily be split off at a suitable stage in the reaction sequence, conveniently at the last stage. It may, however, be convenient in some instances to employ non-toxic metabolically labile carboxyl blocking groups such as acyloxy-methyl or -ethyl groups (e.g. acetoxy-methyl or-ethyl or pivaloyloxymeethyl) and retain these in the final product to give an appropriate ester derivative of a compound of formula (I).

Suitable carboxyl blocking groups are well known in the art, a list of representative blocked carboxyl groups being included in British Patent No. 1399086. Preferred blocked carboxyl groups include aryl lower alkoxycarbonyl groups such as p-methoxybenzyloxycarbonyl, p-nitrobenzyloxycarbonyl and diphenylmethoxycarbonyl; lower alkoxycarbonyl groups such as t-butoxycarbonyl; and lower haloalkoxycarbonyl groups such as 2,2,2-trichloroethoxycarbonyl. The carboxyl blocking group may subsequently be removed by any of the appropriate methods disclosed in the literature; thus, for example, acid or base catalysed hydrolysis is applicable in many cases, as are enzymically-catalysed hydrolyses.

The antibiotic compounds of the invention may be formulated for administration in any convenient way, by analogy with other antibiotics and the invention thereof includes within its scope pharmaceutical compositions comprising an antibiotic compound in accordance with the invention adapted for use in human or veterinary medicine. Such compositions may be presented for use in conventional manner with the aid of any necessary pharmaceutical carriers or excipients.

The antibiotic compounds according to the invention may, for example, be formulated for injection and may be presented in unit dose form, in ampoules, or in multi-dose containers, if necessary with an added preservative.

The compositions may also take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilising and/or dispersing agents. Alternatively the active ingredient may be in powder form for reconstitution with a suitable vehicle, e.g. sterile, pyrogen-free water, before use.

If desired, such powder formulations may contain an appropriate non-toxic base in order to improve the water-solubility of the active ingredient and/or to ensure that when the powder is reconstituted with water, the pH of the resulting aqueous formulation is physiologically acceptable. Alternatively the base may be present in the water with which the powder is reconstituted. The base may be, for example, an inorganic base such as sodium carbonate, sodium bicarbonate or sodium acetate, or an organic base such as lysine or lysine acetate.

The antibiotic compounds may also be formulated as suppositories e.g. containing conventional suppository bases such as cocoa butter or other glycerides.

Compositions for veterinary medicine may, for example, be formulated as intramammary preparations in either long acting or quick-release bases.

The compositions may contain from 0.1% upwards, e.g. 0.1-99% of the active material, depending on the method of administration. When the compositions comprise dosage units, each unit will preferably contain 100-3000 mg of the active ingredient e.g. 200-2000mg. The daily dosage for adult human treatment will preferably range from 200-12000 mg. e.g. 1000-9000 mg per day, depending inter alia on the nature of the infection and the route and frequency of administration. In general, intravenous or intramuscular administration will be employed, for example using 400 to 6000 mg per day e.g.. 500 to 4000 mg per day of the active ingredient in adult human treatment. In treating Pseudomonas infections higher daily doses may be required.

It will be appreciated that in some circumstances, for example, in the treatment of neonates, smaller dosage units and daily dosage units may be desirable.

The antibiotic compounds according to the invention may be administered in combination with other therapeutic agents such as antibiotics, for example penicillins or other cephalosporins.

The following Examples illustrates the invention. All temperatures are in C ; DMSO is dimethylsulphoxide; EtOH is ethanol.

INTERMEDIATE 1 Ethyl (Z) -2-fluoromethoxyimino-2- (triphenylmethyl-aminothiazol-4-yl) acetate Ethyl (Z) -2-hydroxyimino-2- (2-triphenylmethyl-aminothiazol-4-yl) acetate, hydrochloride salt (8.7g) was stirred with potassium carbonate (1 5.35g) in dimethyl sulphoxide (30ml) under nitrogen at 21 C.

Bromofluoromethane (ca 39) was added. The nitrogen flow was stopped and the stirring continued for two hours. The mixture was poured into an ice-water mixture with stirring and the solid was collected by filtration and washed with water. The solid was dissolved in methylene chloride and the organic layer was separated and dried with magnesium sulphate. Evaporation gave a foam. This was dissolved in methylene chloride and pre-absorbed onto Kieselgel 60 (50g). This was added to the top of a column of similar silica (1 25g) set up in 10% ethyl acetate in cyclohexane. The column was eluted successively with 10%, 20% and 33% ethyl acetate in cyclohexane.After combination of appropriate fraction, evaporation gave the title compound (8.06g) as a foam; max (EtOH) 302nm (E1cm 1%92), #infl include 227.5nm (E1cm 1%546) and 259nm (E1cm 1%221), vmax (CHBr3) 3400 (NH), 1739 (ester) and 1522cm-1 (C=N).

INTERMEDIATE 2 (Z) -2-Fluoromethoxyimino-2- (2-triphenylmethylamino-thiazol-4 yl) acetic acid.

Intermediate 1 (7.8g) was stirred under reflux with sodium hydroxide (0.83g) in ethanol (50ml) and water (1 Oml) for 15 minutes. The mixture was cooled and the crystalline precipitate was collected by filtration and washed with ethanol and ether and dried. This solid was partitioned between methylene chloride (80ml) and water (40ml) with vigorous stirring and 88% orthophosphoric acid (2ml) was added. Solid remained and this was collected by filtration. This solid was suspended in tetrahydrofuran (75ml) and 2M hydrochloric acid (8ml) was added when a solution formed. Evaporation reduced the volume of solution by one half and methylene chloride (50ml) was added.The aqueous layer was extracted with more methylene chloride and the combined organic layers were washed with water, dried with magnesium sulphate and evaporated to a solid, the title compound (4.82g) ; #infl include 224nm (E1cm 1%564), 254.5nm (E1cm 1%) and 260nm (E1cm 1% 205) ;# (d6DMSO) 1.02 (s; NH), 2.64 (s; phenyl protons) 2.91 (s; thiazole 5-H), and 4.29 (d, J 56 Hz; CH2F).

EXAMPLE 1 a) (6R, 7R,2 'Z) - 7-[2-Fluoromethoxyimino -2- (2-tritylaminothiazol-4-yl) acetamidolj-3- (11- (2,3- cyclopentenopyridinium) -methyl)ceph-3-em-4-carboxylate Oxalyl chloride (0.15ml) was added to a solution of DMF (0.2ml) in methylene chloride (10ml) with stirring under nitrogen at -20'C. After ten minutes with ice-water cooling, the mixture was recooled to - 20 C and (Z) -2- Fluoromethoxyimino-2- (2-tritylaminothiazol-4-yl) acetic acid (740mg) was added.The solution was stirred with ice-water cooling for ten minutes before recooling to - 20 C and the solution was added to a stirred solution of (6R,7R)-7-amino-3-(1-(2,3-cyclopentenopyridinium)methyl)ceph-3-em-carboxylate (720mg) in industrial methylated spirits (12ml) containing water (3ml) and triethylamine (1.2ml) at ca 5 . The solution was allowed to warm to 21 over 30 minutes. The solution was poured into water containing sodium chloride and the product was extracted with methylene chloride (twice). The combined organic layers were washed with water and dried (sodium sulphate) and evaporated. The residue was triturated with ethyl acetate to give a solid (850mg). This was dissolved in methylene chloride and loaded onto a column of Sorbsil U30 (209) set up in acetone.The column was eluted with 9:1, acetone:water, and then 3:1, acetone:water which brought off the product. The acetone was evaporated and the aqueous residue was extracted with methylene chloride. After drying with sodium sulphate, evaporation and trituration with diethyl ether gave the title compound (480mg), #max (Ethanol) 267nm (E1cm 1% 213), #infl include 225nm (E1cm 1%447), 262.5nm (E1cm 1%208), 227 nm (E1cm 1% 2298nm (E1cm 1%91), vmax (Nujol) 3700 to 2500 (NH, NH+), 1778 (ss-lactam, 1679 and 1529 (amide) and 1620cm-1 (COO-).

b) (6R,7R,2 'Z)-7-[2- (2-Aminothiazol-4-yl) -2-FluoromethoxyimiPoacetamido]-3- (1- (2,3-cyclopen- tenopyridinium)-methyl)ceph-3-em-4-carboxylate The above protracted substance (420mg) was dissolved in formic acid (3ml) with stirring and water (1.5ml) was added. After 1.75 hours at 21", the mixture was filtrated and the filter-cake was leached with 2:1, formic acid: water. The combined filtrates were evaporated and the residue triturated with acetone.The precipitate was collected by filtration, washed with acetone and dried to give the title compound (200mg), #max (pH buffer) 223.5nm (E1cm 1%339), and 259.5nm (E1cm 1%259) #infl 265.5nm (E1cm 1%258) and 302nm (E1cm 1% 104), vmax (Nujol) 3700 to 2200 (NH2, NH, NH+), 1776 (ss-lactam), 1671 and 1539 (amide) and 1620cm-1 (COO-).

EXAMPLE 2 a) (6R, 7R,2 'Z)-7-[2-Fluoromethoxyimino-2- (tritylaminothiazol-4- yl) acetamidoj- 3- (1 - isoquinollniummethyl) - ceph-3-em-4-carboxylate Oxalyl chloride (0.1 5ml) was added to a solution of DMF (0.2ml) in methylene chloride (10ml) with stirring under nitrogen at -20' and the mixture was stirred with ice-water cooling for ten minutes before recooling to -20 , (Z)-2-Fluoromethoxyimino-2-(2-tritylaminothiazol-4-yl) acetic acid (750mg) was added and the solution was stirred with ice-water cooling for ten minutes before recooling to -20 .This solution was added to a solution of (6R,7R)-7-amino-3-(1-isoquinoliniummetyl) ceph-3-em-4-carboxylate (680mg) in industrial methylated spirits (1 2ml) and water (3ml) containing triethylamine (1.4ml) at 21". The solution was allowed to warm to 21 over 30 minutes and then poured into water. Two drops of 2N hydrochloric acid were added and the mixture was extracted with methylene chloride twice and the combined organic layers were then washed with water twice and dried with sodium sulphate.Evaporation and trituration with diethyl ethyl and ethyl acetate gave a solid (1.05g). A portion (890mg) was triturated with ethanol (50ml) and the residue dried to give the title compound (590mg). #max (ethanol) 318.5nm (E1cm 1% 76), #infl include 229.5nm (E1cm 1%612, 259.5nm (E1cm 1% 223), 266nm (E1cm 1%218) and 304nm (E1cm 1%98), vmax (Nujol) 3400 (NH), 1776 (ss-lactam), 1672 and 1524 (amide) and 16118cm-1 (COO-).

b) (6R, 7R,2'Z)-7-[2-Fluoromethoxyimino-2-(2-aminothiazol-4-yl)acetamido]-3-(1-isoquinoliniummethyl)cephh-3-em-4-carboxylatee The above protected intermediate (520mg) was dissolved in formic acid (3.5ml) and water (1.5ml) was added with stirring. After 30 minutes at 21", the mixture was filtered and the filter-cake was leached with formic acid -water mixture (7:3). The combined filtrates were evaporated and the residue was triturated with acetone.

precipitate was collected by filtration, washed with acetone and dried to give the title compound (250mg), #infl (pH 6 buffer) 233nm (E 1% 820), 254nm (E 1% 319), 275nm (E 1% 234), 289nm (E 1% 175) and 323nm 1cm 1cm 1cm 1cm (E 1% 96), vmax (nuiol 3300 (NH) 1778 (ss-lactam) 1670 and 1530 (amide) and 1610-1 (COO-) 1cm -

Claims (1)

1. Cephalosporin antibiotics of the general formula (I)

wherein R1 represents an isoquinolinium group or a 2,3-cyclopentenopyridinium group, and non-toxic salts and non toxic metabolically labile esters thereof.