INDIUM COMPLEXES OF SUBSTITUTED 8-HYDROXY-QUINOLINES, PHARMACEUTICAL COMPOSITIONS CONTAINING THEM AND THEIR USE AS ANTIBACTERIAL AGENTS

Abstract: Complexes of indium with ring substituted derivatives of 8-hydroxyquinoline having a halogen substituent at one or both of the 5- and 7-positions and/or a halogenated alkyl group substituent at the 2-position are of value as phar­ maceuticals, particularly as antibacterial agents.

Description: IMPROVEMENTS IN OR RELATING TO ANTIBACTERIAL AGENTS This invention relates to compounds having pharmaceutical activity, particularly as antibacterial agents. 8-Hydroxyquinoline and certain derivatives thereof have been used as pharmaceuticals, particularly by virtue of their antibacterial action, and especially against gram-positive bacteria. This gram-positive anti-bacterial activity is reported to require 2+ 2+ and Fe3+, and 3+ the presence of one of the cation Cu , Fe and Fe3 , and it is believed that the active antibacterial moiety is a complex formed between such a metal cation and the 8-hydroxyquinoline or its derivative CSelective Toxicity by Adrien Albert (Methuen), Fourth Edition, 1968, pages 331 to 3433. 8-Hydroxyquinoline and derivatives thereof have also been employed for the treatment of parasitic and fungal infections. Usually, however, 8-hydroxyquinoline and its derivatives have been limited to topical use, for example for the treatment of dermatoses, although some oral use has occurred in the case of parasitic infections, for example for the treatment of amoebiasis, since parenteral use of the compounds has been ruled out due to their inactivation by serum. Furthermore, the use of the compounds as antibiotics has hitherto largely been limited to the treatment o gram-positive infections. Despite this somewhat restricted p-cture of the value of 8-hydroxyquinoline derivatives as pharmaceuticals and the reported 2+ 2- 3+ necessity for the presence of Cu , Fe or Fe3 in order for their gram-positive activity to be expressed, I have investigated the activity of complexes of certain 8-=ydroxyquinoline derivatives with a particular metal other than iron or copper and found that these novel complexes show considerable promise as antibacterial agents without the limitations referred to above. Not only do the complexes show a markedly higher antibacterial activity than the corresponding metal-free 8-hydroxyquinoline compounds, but they show good levels of activity against gram-negative bacteria and, moreover, have shown activity when given parenterally in vivo. Accordingly, the present invention comprises a complex with indium of a ring substituted derivative of 8-hydroxyquinoline which has a halogen substituent at one or both of the 5- and 7-positions and/or a halogenated alkyl group substituent at the 2-position. Preferred complexes according to the present invention contain one atom of indium in combination with two molecules of the 8-hydroxyquinoline derivative. In such a complex, the trivalent indium cation will be only partially neutralised by the anions derived from the 8-hydroxy group of each molecule of the derivative through the reaction EMI2.1 which occurs on formation of the complex, and an additional monovalent physiologically acceptable anion is required to establish neutrality. Such an anion may, for example, be a halide ion such as the chloride ion or may be any other such physiologically acceptable anion, for example one selected with a particular view to enhancing the aqueous solubility of the complex and being derived from an acid such as methane sulphonic acid, isethionic acid, etc. The present invention does not exclude complexes containing a proportion of indium:8-hydroxyquinoline derivative other than 1:2 and spectrometric studies of the stoichiometry of the reaction of In 3+ with various 8-hydroxv- quinoline derivatives have indicated the formation in solution cf various other complexes, for example 1:1 complexes which require a divalent or two monovalent physiologically acceptable anions to establish neutrality, although in contrast to the situation with 8-hydroxyquinoline itself, 1:3 complexes which have the advantage of being neutral as such without the presence of an additional anion, appear to be more difficult to form. It is believed, however, that the 1:2 complexes are in general the most stable form, although this may not always be the case so that with 7-iodo8-hydroxyquinoline 5-sulphonic acid, for example, it appears that whilst this compound will form a 1:2 complex, a 1:5 complex may be the most stable form. It is usually preferred therefore to use either a solid 1:2 complex or a solution containing a complex formed from one molar equivalent of In and 2 molar equivalents of the 8-hydroxyquinoline derivative. Among the halogens comprising a halogen substituent, iodo and especially chloro groups are of somewhat greater interest, although fluoro and especially bromo groups are also of interest. Halogen substitution at the 5-position, either as a 5,7-dihalogenated derivative or as a 5-monohalogenated derivative is often of most interest. In the case of the halogenated alkyl group substituents, fluorinated groups are of particular interest and the halogenated alkyl group is preferably a lower alkyl (C1 to C4) group, for example a halogenated methyl group such as a trifluoromethyl group. The 8-hydroxyquinoline derivative may contain two or more different halogen atoms, and may contain both a 5- and/or 7-halogen substituent and a 2-halogenated alkyl group substituent, although halogen substitution is in general of greater interest. Other types of substituent may also be present, either at one of the 5or 7-positions and/or at other ring carbon atom positions, including the 2-position, although it is possible that substitution adjacent to the nitrogen atom may inhibit the formation of the usual form of complex and result instead in the formation of an alternative form of complex, for example containing a 1:6 proportion of indium: 8-hydroxyquinoline derivative, which may possess a lower level of activity. For this reason, 2-substituted 8-hydroxyquinoline derivatives, including those having a halogenated alkyl group substituent at this position, are of somewhat lesser interest in the context of the present invention than derivatives not containing such a substituent. Additional substituents may include one or more lower alkyl groups such as methyl, for example a 2-methyl group, but groups of particular interest are those of an ionic nature which confer a greater degree of water solubility on the derivative, although it should be appreciated that the presence of an additional substituent may also reduce activity, for example as discussed above. Such ionic groups include carboxylic and sulphonic acid groups in either the free acid form (-C02H and -SO3H) or in the form of physiologically acceptable salts such as those formed with the alkali metals, for example sodium, or with an ammonium or substituted ammonium cation. Other opportunities for enhancing water solubility through salt formation are somewhat limited since the hydroxy group of each 8-hydroxyquinoline derivative molecule in the complex is usually involved in complex formation and involvement of the nitrogen lone pair of electrons in salt formation can adversely affect complex formation. Examples of specific complexes according to the present invention are the indium complexes of 5-chloro-8-hydroxyquinoline, 7-chloro-8-hydroxyquinoline, 5,7-dichloro-8-hydroxyquinoline, 5-chloro-7-iodo-8-hydroxyquinoline, 5,7-diiodo-8-hydroxyquinoline, 5,7-dibromo-8-hydroxyquinoline, 7-bromo-5-chloro-8-hydroxyquinoline, and 7-iodo-8-hydroxyquinoline 5-sulphonic acid and physiologically acceptable salts thereof. The indium complexes of the 2-methyl derivatives of these compounds, for example 5,7-dichloro-2-methyl8-hydroxyquinoline, may also be specifically mentioned, as may the indium complexes of 2-trifluoromethyl-8-hydroxyquinoline and of the analogues of the 2-methyl compounds referred to above in which the 2-methyl group is replaced by a 2-trifluoromethyl group. The complexes are conveniently prepared by reaction in a suitable mutual solvent of the 8-hydroxyquinoline derivative and a suitable indium salt, for example an indium halide and particularly indium chloride, usually as the trihydrate, InCl3.3H20, in appropriate proportions, for example a 1:2 molar proportion of salt: derivative. Where the complex contains an additional anion to achieve neutrality, the indium salt may itself provide this anion, although it is possible if desired to insert an alternative anion subsequent to the initial formation of a complex. Suitable solvents for the reaction of derivative and indium salt include organic solvents such as alcohols and ketones, indium chloride being of interest in this respect for its significant level of solubility in organic solvents. Alternatively, aqueous solvent mixtures may be used so that in one procedure, for example, the indium salt in dilute aqueous hydrochloric acid is mixed with a solution of the 8-hydroxyquinoline derivative in ethanol, or in dimethylsulphoxide. Reaction to form the complex usually takes place quite rapidly at room temperature so that a period of 30 minutes, for example, is usually fully sufficient. Depending upon the solvent used, the particular reactants and their concentration, precipitation may occur from the reaction solution, for example consequent upon dilution of a solution in ethanol or dimethylsulphoxide with water, or, alternatively, the solvent may be removed in vacuo to leave the complex. In some cases the complex may be obtainable in crystalline form. Synthetic routes to a variety of 5- and/or 7-halogenated 8-hydroxyquinolines are described in the literature. 2-Trifluoromethyl-8-hydroxyquinoline may be prepared by the procedure of March et al, Journal of Medicinal Chemistry, 1973, 16, 337-342 and other derivatives containing a 2-trifluoromethyl group by an analogous procedure. It will be appreciated that, in general, modifications of the procedures described in the art for the preparation of the 8-hydroxyquinoline derivatives and of the procedures described above for the preparation of the indium complexes may be used, if desired, as will be apparent to those skilled in the art. The formulation of complexes according to the present invention for use as a pharmaceutical for both human and animal administration may be effected by a variety of methods, but usually involves the use of a physiologically acceptable diluent or carrier. The complex may, for instance, be applied as an aqueous or oily solu=ion, suspension or emulsion for parenteral administration, the compcsition therefore preferably being sterile and pyrogen-free. The complex may also be compounded for oral administration, more usually in the presence of conventional solid carrier materials such as starch lactose, dextrin and magnesium stearate. Alternative formulations are as aerosols, suppositories, cachets and, for localised treatment, as suitable creams, lotions or drops. The compositions may conveniently be formulated in unit dosage form, i.e. in the form of discrete portions each containing a unit dose, or a multiple or sub-multiple of a unit dose. Dosage levels may, however, vary quite considerably according to the particular complex and also the type of treatment in which they are used but, as a general guide it may be stated that a suitable regimen, particularly as regards systemic treatments, often involves daily doses at a level of 1 to 20 mg/kg, for example 2.5 or 5 to 10 mg/kg, extending over a suitable period, for example 5-10 days, and repeated, often after a rest period, as required. It will be appreciated, however, that in some circumstances higher or lower dose levels than this may be appropriate. Compositions containing complexes according to the present invention are of interest for the treatment of a range of bacterial infections. Thus, the complexes are of interest for their activity against gram-positive bacteria, in particular those belonging to the genera Staphylococcus and Strephtococcus. In vitro experiments with Staphylococcus aureus and Staphylococcus epidermidis have shown a markedly higher level of activity for the indium complexes than for the corresponding metal free derivative of 8-hydroxyquinolone. Moreover, the complexes show very worthwhile activity against gram-negative bacteria, especially aerobic rather than anaerobic bacteria, for example various Enterobacteriaceae such as Escherichia, Klebsiella, Aerobacter and Salmonella. It will be appreciated, however, that the complexes according to the present invention, and specific complexes in particular, may be of more interest in the treatment of infections caused by some of these bacteria than by some others. Thus, for example, we have found that in general the level of activity of the complexes against Klebsiella pneumoniae, Escherichia coli and Salmonella typhinurium is often greater than against Pseudomonas aeruginosa. In addition to systemic use, including parental administration for which the indium complexes, unlike the metal-free compounds, are suitable, the complexes are also of interest for localised action, for example through topical application to the skin for the treatment of gram-negative and especially gram-positive bacterial infections, for example for the treatment of seborrheic dermatitis, the higher activity of the indium complexes giving them a marked advantage as compared with the metal-free compounds in this area also. As indicated previously, formulations for topical administration include creams, lotions etc. for use, for example of formulations for topical administration include a formed material, for example in the form of a patch, etc., which may be attached to the skin at a particular place on the body and thereby provide a dose of the indium complex at that location by release from the material. An alternative use, of particular relevance in the context of gram-negative bacterial infections and where treatment may be effected at the required site through oral administration, is the treatment of infections of the gut, for example the treatment of gastroenteritis caused by E. coli, typhoid fever, cholera caused by Vibrio cholerae and dysentery caused by Shigella. Although the compounds of the present invention are of particular interest for the treatment of bacterial infections, they are also of some interest for the treatment of fungal and especially parasitic infections, being used in a broadly similar manner to that described above for the treatment of bacterial infections. The invention is illustrated by the following Examples. EXAMPLES EXAMPLE 1: PREPARATION OF INDIUM COMPLEXES 3+ 1:2 complexes are prepared of lr with the 8-hydroxyquinoline derivatives: 5-chloro-8-hydroxyquincline; 5,7-dichloro-8-hydroxyquinoline; 5-chloro-7-iodo-8-hydroxycainoline; and 5, 7-d iiod o-8- hydroxyquinoline. In each case india= chloride trihydrate (InC13.3H20; 0.137 g, 0.5 m.mole) an¯ 1 m.mole of the 8-hydroxyquinoline derivative are dissolved together in dimethylsulphoxide (20 ml) to give a 0.5 mM solution of he complex, the formula of which is illustrated below in respect of the second of the 8-hydroxyquinoline derivatives named above. EMI7.1 EXAMPLE 2: PREPARATION OF INDIUM COMPLEX OF 5,7-DICHLORO-8 HYDROXYQUINOLINE Indium chloride trihydrate (1.37 g, 5.0 m.mole) and 5,7-dichloro-8-hydroxyquinoline (2.14 g, 10 m.mole) are dissolved together in ethanol (200 ml) and the solution is concentrated to dryness on a rotary evaporator. The resulting yellow glass is recrystallised by solution in a small volume of ethanol followed by the careful addition of water to give the indium complex of 5,7-dichloro-8-hydroxyquinoline having the formula shown in Example 1 as a bright yellow powder with a melting point above 3400C. The complex has a low solubility in water but is readily soluble in a wide range of organic solvents. EXAMPLE 3: IN VITRO ACTIVITY OF INDIUM COMPLEXES Experiments were carried out to find the minimal inhibitory concentrations (MIC) for solutions of various indium complexes against Staphylococcus aureus (human isolate), Escherichia coli (018; colV ) and Klebsiella pneumoniae and to compare these with the MIC values for the corresponding metal-free compounds. The complexes were prepared in solution as described in Example 1 except that in the case of the complex of 7-iodo-8-hydroxyquinoline 5-sulphonic acid, 5 molar equivalents of metal-free compound were reacted with the indium chloride trihydrate and the solution was prepared in water. The solutions of the metal-free compounds were prepared in the same solvent as used for the equivalent complex. The culture medium used consisted of a 9:1 mixture of trypticase soy broth and 6% w/v aqueous sodium bicarbonate solution. 4 The medium was bulk inoculated with approximately 10 test bacteria per ml and 5.0 ml aliquots were then dispensed into sterile 20 ml bottles to which 0.05 ml of the test solution had been added. The test solutions were obtained from the original solution of the indium complex or metal-free compound by dilution to give a suitable range of concentrations in wM. Control samples, to which either no test solution was added or to which the solvent of the test solution only was added, were also prepared. The bottles were incubated on a rotary shaker for 18 hours at 370C in an atomsphere of 5% C02 in air (higher MIC values being obtained if the sodium bicarbonate and the C02 were omitted). For each compound tested the samples at the various concentrations were studied to determine the lowest concentration of the compound at which no growth could be, seen. The MIC values thus obtained are shown in Table 1 were it will be seen that the values are in each case lower for the indium complex as compared with the metalfree compound. (The controls showed that the solvents had no obvious effect on bacterial growth). Table 1 EMI9.1 <tb> <SEP> Compound <SEP> MIC <SEP> Values <SEP> (pM) <SEP> <tb> <SEP> S. <SEP> aureus <SEP> E. <SEP> coli <SEP> R. <SEP> pneumoniae <tb> 5, <SEP> 7-d <SEP> ichloro-8- <SEP> <tb> hydroxyquinoline <SEP> 6.3 <SEP> 100 <SEP> 100 <tb> Indium <SEP> complex <SEP> 3.2 <SEP> 1.6 <SEP> 12.5 <tb> 5-Chloro-8-kydroxyquinoline <SEP> 18.8 <SEP> 38.5 <SEP> 38.5 <tb> Indium <SEP> complex <SEP> 1.6 <SEP> 6.3 <SEP> 25 <tb> 7-Iodo-8-hyizoxyquinoline <SEP> <tb> 5-sulphonic <SEP> acid <SEP> > 500 <SEP> > 500 <SEP> > 500 <tb> Indium <SEP> complex <SEP> 50 <SEP> > 100 <SEP> > 100 <tb> Generally similar results have been obtained with Staphylococcus epidermis (human isolate) as for S. aureus. EXAMPLE 4: IN VIVO TESTS OF INDIUM COMPLEX OF 5, 7-DICHLORO-8- HYDROXY QUINOLINE Experiments were carried out to compare the activity in vivo against E. coli (018, colV+), K. pneumoniae and S. aureus (human isolate) of the indium complex of 5,7-dichloro-8-hydroxyquinoline with that of the metal-free compound. The solutions for administration were prepared as follows: (1) 2 equivalents of 5,7-dichloro-8-hydroxyquinoline were dissolved in a 15 mM solution of InCi3. 3H20 in ethanol containing 1% v/v of glycerol. A 1.0 ml aliquot of this solution was dried in vacuo using a sterile universal container and the resultant residue was treated with 15 ml of sterile 0.15 M aqueous sodium chloride containing 1% w/v of sodium bicarbonate and the solid dispersed by sonication. A 0.2 ml dose of the final preparation corresponds to a dose of 4.6 mg/kg in a 25 g mouse. (2) 6.4 mg of 5,7-dichloro-8-hydroxyquinoline were moistened by the addition of 0.05 ml of dimethyl sulphoxide and to the moistened material was added 15 ml of sterile 0.15 M aqueous sodium chloride containing 1% w/v of sodium bicarbonate, the solid then being dispersed by sonication. A 0.2 ml dose of the final preparation corresponds to a dose of 3.4 mg/kg in a 25 g mouse. Groups of 5 male or female Parks mice weighing from 23-25 g were infected by intraperitoneal injection of one of 8.0 x 106 E. coli, 3.5 x 10 K. pneumoniae and 7.0 x 10 S. aureus, the organisms -n each case bing suspended in 0.2 ml of 10% v/v trypticase soy broth in 0.15 M aqueous sodium chloride, three groups of zice being infected with each organism. For each organism, sne of the three groups of infected mice was treated by intraperiteneal injection with 0.2 ml of a suspension of the indium comrlex whilst a second group was similarly treated with 0.2 ml of E suspension of the metal-free compound. The treatment was given twice per day for 1 day in the case of E. coli infection, twice per cay for 4 days in the case of K. pneumoniae infection, and twice per day for 2 days in the case of the S. aureus infection. The third group of mice in each case received no treatment. The results obtained in terms of the number of deaths at 7 days and the percentage of deaths in the group are shown in Table 2 where it will be seen that in each case the indium complex showed a much greater anti-bacterial effect than the metal-free compound. Table 2 EMI11.1 <tb> <SEP> Compound <SEP> Organism <SEP> Deaths <SEP> at <SEP> Deaths <tb> <SEP> 7 <SEP> days <SEP> X <tb> Control <SEP> 5 <SEP> 100 <tb> 5,7-Dichloro-8-hydroxy <tb> quinoline <SEP> E. <SEP> coli <SEP> 5 <SEP> 100 <tb> Indium <SEP> complex <SEP> 0 <SEP> 0 <tb> Control <SEP> 5 <SEP> 100 <tb> 5, <SEP> 7-Dichloro-8-hydroxy- <SEP> <tb> quinoline <SEP> K. <SEP> pneumoniae <SEP> 5 <SEP> 100 <tb> Indium <SEP> complex <SEP> 0 <SEP> 0 <tb> Control <SEP> 4 <SEP> 80 <tb> 5, <SEP> 7-Dichloro-8-hydroxy- <SEP> <tb> quinoline <SEP> S. <SEP> aureus <SEP> 4 <SEP> 80 <tb> Indium <SEP> complex <SEP> 2 <SEP> 40 <tb>

Claims: CLAIMS 1. A complex with indium of a ring substituted derivative of 8-hydroxyquinoline having a halogen substituent at one or both of the 5 and 7-positions and/or a halogenated alkyl group substituent at the 2-position. 2. A complex with indium of a ring substituted derivative of 8-hydroxyquinoline having a halogen substituent at one or both of the 5- and 7-positions. 3. A complex according to Claim 2, which has a halogen substituent at the 5-position or the 7-position, or halogen substituents at the 5- and 7-positions, said substituent or substituents being selected from chloro, iodo and bromo. 4. A complex according to Claim 3, in which the 8-hydroxyquinoline derivative is substituted only at the 5- and/or 7-positions. 5. A complex according to Claim 4, in which the 8-hydroxyquinoline derivative is selected from 5-chloro-8-hydroxyquinoline, 7-chloro-8hydroxyquinoline, 5,7-dichloro-8-hydroxyquinoline, 5-chloro-7-iodo8-hydroxyquinoline, 5,7-dfiodo-8-hydroxyquinoline, 5,7-dibromo-8hydroquinoline, 7-bromo-5-chloro-8-hydroxyquinoline, and 7-iodo-8hydroxyquinoline 5-sulphoric acid and physiologically acceptable salts thereof. 6. A complex according to Claim 1, which has a fluorinated lower alkyl group substituent at the 2-position. 7. A complex according to Claim 1, which has a trifluoromethyl substituent at the 2-posit=on. 8. A complex according to Claim 7, in which the 8-hydroxyquinoline derivative is 2-trifluoromethyl-8-hydroxyquinoline. 9. A complex according to any of Claims 1 to 8, which is a neutral complex containing a 1:2 molar proportion of In3 :8-hydroxy- quinoline derivative. 10. A complex according to Claim 9 being a 1:2 indium complex of 5-chloro-7-iodo-8-hydroxyquinoline, 5,7-dibromo-8-hydroxyquinoline, 5,7-diiodo-8-hydroxyquinoline or 7-iodo-8-hydroxyquinoline 5-sulphonic acid. 11. A complex according to Claim 9, being a 1:2 indium complex of 5, 7-dichloro-8-hydroxyquinoline. 12. A process for the preparation of an indium complex of a ring substituted derivative of 8-hydroxyquinoline having a halogen substituent at one or both of the 5- and 7-positions and/or a halogenated alkyl group substituent at one or both of the 2-position which which comprises reacting said derivative with In ions. 3+ 13. A process according to Claim 12, in which the In ions are provided by an indium trihalide. 14. A pharmaceutical composition which comprises an indium complex of a ring substituted derivative of 8-hydroxyquinoline having a halogen substituent at one or both of the 5- and 7-positions and/or a halogenated alkyl group substituent at the 2-position together with a physiologically acceptable diluent or carrier. 15. A pharmaceutical composition according to Claim 14, in a form suitable for topical application. 16. A pharmaceutical composition according to Claim 15, being an ointment or shampoo. 17. A pharmaceutical composition according to Claim 15, being a formed material for attachment to the skin and incorporating the indium complex. 18 A complex with indium of a ring substituted derivative of 8--ydroxyquinoline having a halogen substituent at one or both of the 5- and 7-positions and/or a halogenated alkyl group substituent at the 2-position for use in the treatment of bacterial infections.

Inventors: ROGERS, HENRY JOSEPH
Applicants: NATIONAL RESEARCH DEVELOPMENT CORPORATION
Application year: 1983
Main classification: C07D215
IPC codes: C07F5, C07D215