WO2012153130A1 - Salts of 4 -ethyl-1-(2 - phenylethyl) - 8 -phenoxy - 2, 3 - dihydro - 1h - pyrrolo [3, 2 -c] quinoline and their use for treating infections - Google Patents

Abstract

The present invention relates to certain acid addition salts of 4-methyl-1-(2-phenylethyI)-8- phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]-quinoline, including the methane sulfonic acid addition salt thereof, to a process for the preparation of such compounds, and to the use of such compounds for the treatment of microbial infections.

Description

SALTS OF 4 - ETHYL - 1 - ( 2 - PHENYLETHYL) - 8 - PHENOXY -2,3 - DIHYDRO - 1H - PYRROLO [3 , 2 -C] QUINOLINE AND THEIR USE FOR TREATING INFECTIONS

The present invention relates to certain acid addition salts of 4-methyl-1-(2-phenylethyl)-8- phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline, including the methane sulphonic acid addition salt, to a process for the preparation of such compounds, and to the use of such compounds for the treatment of microbial infections.

The synthesis and antimicrobial activity of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro- 1 H-pyrrolo[3,2-c]-quinoline is disclosed in International Patent Application, Publication Number WO2007/054693. This application also discloses the hydrochloride acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline. International Patent Application, Publication Number WO2008/056151 discloses topical pharmaceutical compositions comprising a variety of pyrrolo[3,2-c] quinoline derivatives including 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline and pharmaceutically acceptable derivatives thereof.

Pharmaceutically active ingredients (APIs) with one or more basic centres may be prepared and formulated as acid addition salts. The preparation of such salt forms may improve the physical and/or pharmaceutical properties of the basic API. However, it is generally not possible to predict which salt forms of a given API will possess advantages relative to the API in free base form, or to other salt forms of the same API, prior to preparation and characterisation of each individual salt form.

It has now surprisingly been discovered that certain acid addition salts of 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline, including the methane sulphonic acid addition salt, possesses improved aqueous solubility compared to the corresponding free base, are thermally stable, non-hygroscopic and readily susceptible to large-scale synthesis. These unexpected properties render the acid addition salts of the present invention particularly suitable for development as APIs.

Thus, according to a first embodiment of the invention, there is provided an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid, sulphuric acid and tartaric acid addition salts thereof, i.e.: 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hydrobromide;

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline methane sulphonate (or mesylate);

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrro!o[3,2-c]-quinoline p-toluene sulphonate (or tosylate);

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline succinate and hemi-succinate;

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline sulphate; and

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline tartrate and hemi-tartrate.

In a preferred embodiment of the invention, the acid addition salt of 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline is selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid and sulphuric acid addition salts thereof. These salts have been shown to possess particularly good thermal stability and purity profiles.

In an alternative preferred embodiment of the invention, the acid addition salt of 4-methyl-1- (2-phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]-quinoline is selected from the group consisting of the methane sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof. These salts have been shown to possess improved aqueous solubility compared to the corresponding free base and the hydrochloride acid addition salt.

In a more preferred embodiment of the invention, the acid addition salt of 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline is selected from the group consisting of the methane sulphonic acid, succinic acid (preferably hemi-succinate) and tartaric acid (preferably hemi-tartrate) addition salts thereof.

In the most preferred embodiment of the invention, the acid addition salt is the methane sulfonic acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline. According to a further embodiment of the invention, there is provided a pharmaceutical composition comprising an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3- dihydro-1H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi- succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts, and a pharmaceutically acceptable carrier. Such a composition may be used for treatment of a microbial infection, in particular for killing multiplying and/or clinically latent microorganisms associated with such an infection.

In a further embodiment of the invention, there is provided the use of an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts, for the treatment of a microbial infection, in particular for killing multiplying and/or clinically latent microorganisms associated with such an infection. In still a further embodiment of the invention, there is provided a method of treating a microbial infection, in particular killing multiplying and/or clinically latent microorganisms associated with such an infection, which comprises administering to a mammal, including man, an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts.

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline has the following chemical structure:

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline free base and the hydrochloride salt thereof may be prepared according to the methods disclosed in WO2007/054693 and WO2008/056151 , the contents of which are incorporated herein by reference as if the publication was specifically and fully set forth herein.

The acid addition salts of the present invention may be prepared by conventional methods in the art, for example as described in Berge, S. M. et al., J. Pharm. Sci., 1977, 66(1), 1 - 19; Stahl, P.H. and Wermuth, C.G., Handbook of Pharmaceutical Salts: Properties, Selection and Use, 2011 , 2^nd Edition, Wiley-VCH, and references cited therein.

By way of example, a representative process for the preparation of 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline methane sulphonate is given in the scheme below.

HCI

(5)

(i) EtOAC, MeOH, 40% K₂C0₃

(ii) MsOH

(6) where, EtOAc - ethyl acetate; MeOH - methanol; and MsOH - methane sulphonic acid The reagents 4-phenoxyaniline (compound (1)) and 2-acetylbutyrolactone (compound (2)) are commercially available, for example from Sigma-Aldrich.

Representative ¹H NMR, XRPD and DSC data for 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3- dihydro-1 H-pyrrolo[3,2-c]-quinoIine methane sulphonate prepared according to the method of Example 2 are reproduced herein in Figures 1, 2 and 3 respectively.

A representative ¹H NMR spectrum for 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro- 1 H-pyrrolo[3,2-c]-quinoline hemi-succinate is reproduced in Figure 4.

Representative H NMR and XRPD spectra for 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3- dihydro-1 H-pyrrolo[3,2-c]-quinoline p-toluene sulphonate are reproduced in Figures 5 and 6 respectively.

According to a preferred embodiment of the present invention, there is provided 4-methyl-1- (2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline methane sulphonate characterised by at least one of the following:

(i) a ¹H NMR spectrum (270MHz, CDCI₃) with peaks at 8.37 (1 H, d, J = 9.2 Hz), 7.45-7.55 (1 H, m), 7.35-7.45 (3H, m), 7.1-7.3 (4H, m), 6.92-7.1 (4H, m), 3.7-3.9 (4H, m), 3.02 (2H, t, J = 9.6 Hz), 2.88 (3H, s and 2H, t, 7.2Hz) and 2.60 (3H, s); and/or

(ii) a H NMR spectrum (270MHz, DR-DMSO) with peaks at 7.86 (1 H, d, J=9.2Hz), 7.68 (1H, dd, J=2.7Hz, 9.4Hz), 7.58 (1 H, d, 2.2Hz), 7.38-7.51 (2H, m), 7.08-7.28 (6H, m), 6.97-7.06 (2H, m), 3.7-4.1 (4H, m), 3.08 (2H, t, J=9.6Hz), 2.88 (2H, t, 7.4Hz), 2.46 (3H, s), 2.30 (3H, s); and/or

(iii) an XRPD pattern with peaks at 8.33, 9.33, 11.92, 14.52, 14.99, 16.59, 16.79, 17.07, 17.23, 17.37, 18.32, 18.86, 19.46 ±0.2 ⁰ 2Θ; and/or

(iv) a melting point of from about 178 to about 181 °C. In a further preferred embodiment of the invention, there is provided 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]-quinoline methane sulphonate having a ¹H NMR spectrum substantially as depicted in Figure 1 herein.

In still a further preferred embodiment of the invention, there is provided 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]-quinoline methane sulphonate having an XRPD pattern substantially as depicted in Figure 2 herein. In still a further preferred embodiment of the invention, there is provided 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline methane sulphonate having a DSC trace substantially as depicted in Figure 3 herein.

According to a further embodiment of the present invention, there is provided a process for preparing an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts, which process comprises: dissolving or suspending 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline free base in a suitable solvent; treating the resulting solution or suspension with an acid selected from the group consisting of hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid, sulphuric acid or tartaric acid; and

(iii) optionally isolating the compound so formed.

According to a further embodiment of the present invention, there is provided a process for preparing an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts, which process comprises:

(i) dissolving or suspending an acid addition salt of 4-methyl-1-(2-phenylethyl)-8- phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]-quinoline, preferably the hydrochloride salt thereof, in a suitable solvent; (ii) treating the resulting mixture with a suitable base;

(iii) treating the resulting solution or suspension with an acid selected from the group consisting of hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid, sulphuric acid or tartaric acid; and

(iv) optionally isolating the compound so formed. Suitable reaction solvents include esters, alcohols and mixtures thereof. Preferably, the solvent is de-gassed before use. Examples of suitable esters include methyl acetate, ethyl acetate, n-propyl acetate, /so-propyl acetate, n-butyl acetate, /so-butyl acetate, fert-butyl acetate and sec-butyl acetate, and mixtures thereof. Examples of suitable alcohols include methanol, ethanol, n-propanol, /so-propanol, n-butanol, /so-butanol, ferf-butanol and sec- butanol, and mixtures thereof. Preferably, the solvent is a mixture of ethyl acetate and methanol. More preferably, the ratio of ethyl acetate to methanol is from 5:1 to 10:1 , most preferably 10:1.

Suitable bases include alkali and alkaline earth metal carbonates, alkali metal hydroxides and alkali metal bicarbonates. Examples of suitable alkali and alkaline earth metal carbonates include magnesium carbonate, calcium carbonate, lithium carbonate, sodium carbonate and potassium carbonate. Examples of suitable alkali metal hydroxides include magnesium hydroxide and calcium hydroxide. Examples of suitable alkali and alkaline earth metal bicarbonates include magnesium bicarbonate, calcium bicarbonate and sodium bicarbonate.

Suitably, the ratio of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline (free base) to acid employed is from 1 :1 to 1 :5 equivalents, such as 1 :1, 1 :1.5, 1 :2, 1 :3, 1 :4 or 1 :5 equivalents. Preferably, the ratio is 1 :1.

Both succinic and tartaric acid possess two ionising sites and have been found to be capable of forming "hemi" salts with 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline. 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro- H-pyrrolo[3,2- c]-quinoline hemi-succinate and hemi-tartrate may be prepared using a free base to acid ratio of about 1 :0.5 equivalents, such as 1 :0.4, 1 :0.5 or 1 :0.6 equivalents.

The acid addition salts of the present invention may be isolated from solution using conventional means, for example by filtration, followed by washing and/or drying.

The acid addition salts of the present invention possess antimicrobial activity and may in particular be used to kill multiplying and/or clinically latent microorganisms associated with microbial infections. In a preferred embodiment of the invention, there is provided the use of an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]- quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts, to kill clinically latent microorganisms associated with a microbial infection. As used herein, "kill" means a loss of viability as assessed by a lack of metabolic activity.

As used herein, "clinically latent microorganism" means a microorganism that is metabolically active but has a growth rate that is below the threshold of infectious disease expression. The threshold of infectious disease expression refers to the growth rate threshold below which symptoms of infectious disease in a host are absent.

The metabolic activity of clinically latent microorganisms can be determined by several methods known to those skilled in the art; for example, by measuring mRNA levels in the microorganisms or by determining their rate of uridine uptake. In this respect, clinically latent microorganisms, when compared to microorganisms under logarithmic growth conditions (in vitro or in vivo), possess reduced but still significant levels of:

(I) mRNA (e.g. from 0.0001 to 50%, such as from 1 to 30, 5 to 25 or 10 to 20%, of the level of mRNA); and/or

(II) uridine (e.g. [³H]uridine) uptake (e.g. from 0.0005 to 50%, such as from 1 to 40, 15 to 35 or 20 to 30% of the level of [³H]uridine uptake). Clinically latent microorganisms typically possess a number of identifiable characteristics. For example, they may be viable but non-culturable; i.e. they cannot typically be detected by standard culture techniques, but are detectable and quantifiable by techniques such as broth dilution counting, microscopy, or molecular techniques such as polymerase chain reaction. In addition, clinically latent microorganisms are phenotypically tolerant, and as such are sensitive (in log phase) to the biostatic effects of conventional antimicrobial agents (i.e. microorganisms for which the minimum inhibitory concentration (MIC) of a conventional antimicrobial is substantially unchanged); but possess drastically decreased susceptibility to drug-induced killing (e.g. microorganisms for which, with any given conventional antimicrobial agent, the ratio of minimum microbiocidal concentration (e.g. minimum bactericidal concentration, MBC) to MIC is 10 or more).

As used herein, the term "microorganisms" means fungi and bacteria. References herein to "microbiaf, "antimicrobiaf and " antimicrobial^' shall be interpreted accordingly. For example, the term "microbiaf means fungal or bacterial, and "microbial infection" means any fungal or bacterial infection. As used herein, the term "bacteria" (and derivatives thereof, such as "microbial infection") includes, but is not limited to, references to organisms (or infections due to organisms) of the following classes and specific types: Gram-positive cocci, such as Staphylococci (e.g. Staph, aureus, Staph, epidermidis, Staph, saprophytics, Staph, auricularis, Staph, capitis capitis, Staph, c. ureolyticus, Staph, caprae, Staph, cohnii cohnii, Staph, c. urealyticus, Staph, equorum, Staph, gallinarum, Staph, haemolyticus, Staph, hominis hominis, Staph, h. novobiosepticius, Staph, hyicus, Staph, intermedius, Staph, lugdunensis, Staph, pasteuri, Staph, saccharolyticus, Staph, schleiferi schleiferi, Staph, s. coagulans, Staph, sciuri, Staph, simulans, Staph, warneri and Staph, xylosus);

Streptococci (e.g.beta-haemolytic, pyogenic streptococci (such as Strept. agalactiae, Strept. cam^'s, Strept. dysgalactiae dysgalactiae, Strept. dysgalactiae equisimilis, Strept. equi equi,

Strept. equi zooepidemicus, Strept. iniae, Strept. porcinus and Strept. pyogenes), microaerophilic, pyogenic streptococci (Streptococcus "milleri", such as Strept anginosus,

Strept. constellatus constellatus, Strept constellatus pharyngidis and Strept. intermedius), oral streptococci of the "mitis" (alpha-haemolytic - Streptococcus "viridans", such as Strept. mitis, Strept. oralis, Strept sanguinis, Strept. cristatus, Strept gordonii and Strept. parasanguinis), "salivarius" (non-haemolytic, such as Strept. salivarius and Strept. vestibularis) and "mutans" (tooth-surface streptococci, such as Strept criceti, Strept. mutans,

Strept. ratti and Strept. sobrinus) groups, Strept. acidominimus, Strept. bovis, Strept faecalis,

Strept equinus, Strept. pneumoniae and Strept. suis, or Streptococci alternatively classified as Group A, B, C, D, E, G, L, P, U or V Streptococcus);

Gram-negative cocci, such as Neisseria gonorrhoeae, Neisseria meningitidis, Neisseria cinerea, Neisseria elongata, Neisseria flavescens, Neisseria lactamica, Neisseria mucosa, Neisseria sicca, Neisseria subflava and Neisseria weaveri;

Bacillaceae, such as Bacillus anthracis, Bacillus subtilis, Bacillus thuringiensis, Bacillus stearothermophilus and Bacillus cereus;

Enterobacteriaceae, such as Escherichia coli, Enterobacter (e.g. Enterobacter aerogenes, Enterobacter agglomerans and Enterobacter cloacae), Citrobacter (such as Citrob. freundii and Citrob. divernis), Hafnia (e.g. Hafnia alvei), Erwinia (e.g. Erwinia persicinus), Morganella morganii, Salmonella (Salmonella enterica and Salmonella typhi), Shigella (e.g. Shigella dysenteriae, Shigella flexneri, Shigella boydii and Shigella sonnei), Klebsiella (e.g. Klebs. pneumoniae, Klebs. oxytoca, Klebs. ornitholytica, Klebs. planticola, Klebs. ozaenae, Klebs. terrigena, Klebs. granulomatis (Calymmatobacterium granulomatis) and Klebs. rhinoscleromatis), Proteus (e.g. Pr. mirabilis, Pr. rettgeri and Pr. vulgaris), Providencia (e.g. Providencia alcalifaciens, Providencia rettgeri and Providencia stuartii), Serratia (e.g. Serratia marcescens and Serratia liquifaciens), and Yersinia (e.g. Yersinia enterocolitica, Yersinia pestis and Yersinia pseudotuberculosis);

Enterococci (e.g. Enterococcus avium, Enterococcus casseliflavus, Enterococcus cecorum, Enterococcus dispar, Enterococcus durans, Enterococcus faecalis, Enterococcus faecium, Enterococcus flavescens, Enterococcus gallinarum, Enterococcus hirae, Enterococcus malodoratus, Enterococcus mundtii, Enterococcus pseudoavium, Enterococcus raffinosus and Enterococcus solitarius);

Helicobacter (e.g. Helicobacter pylori, Helicobacter cinaedi and Helicobacter fennelliae); Acinetobacter (e.g. A. baumanii, A. calcoaceticus, A. haemolyticus, A. johnsonii, A. junii, A. Iwoffi and A. radioresistens);

Pseudomonas (e.g. Ps. aeruginosa, Ps. maltophilia {Stenotrophomonas maltophilia), Ps. alcaligenes, Ps. chlororaphis, Ps. fluorescens, Ps. luteola. Ps. mendocina, Ps. monteilii, Ps. oryzihabitans, Ps. pertocinogena, Ps. pseudalcaligenes, Ps. putida and Ps. stutzeri);

Bacteriodes fragilis;

Peptococcus (e.g. Peptococcus niger);

Peptostreptococcus;

Clostridium (e.g. C. perfringens, C. difficile, C. botulinum, C. tetani, C. absonum, C. argentinense, C. baratii, C. bifermentans, C. beijerinckii, C. butyricum, C. cadaveris, C. carnis, C. celatum, C. clostridioforme, C. cochlearium, C. cocleatum, C. fallax, C. ghonii, C. glycolicum, C. haemolyticum, C. hastiforme, C. histolyticu , C. indolis, C. innocuum, C. irregulare, C. leptum, C. limosum, C. malenominatum, C. novyi, C. oroticum, C. paraputrificum, C. piliforme, C. putrefasciens, C. ramosu , C. septicum, C. sordelii, C. sphenoides, C. sporogenes, C. subterminale, C. symbiosum and C. tertium);

Mycoplasma (e.g. M. pneumoniae, M. hominis, M. genitalium and M. urealyticum);

Mycobacteria (e.g. Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium fortuitum, Mycobacterium marinum, Mycobacterium kansasii, Mycobacterium chelonae, Mycobacterium abscessus, Mycobacterium leprae, Mycobacterium smegmitis, Mycobacterium africanum, Mycobacterium alvei, Mycobacterium asiaticum, Mycobacterium aurum, Mycobacterium bohemicum, Mycobacterium bovis, Mycobacterium branded, Mycobacterium brumae, Mycobacterium celatum, Mycobacterium chubense, Mycobacterium confluentis, Mycobacterium conspicuum, Mycobacterium cookii, Mycobacterium flavescens, Mycobacterium gadium, Mycobacterium gastri, Mycobacterium genavense, Mycobacterium gordonae, Mycobacterium goodii, Mycobacterium haemophilum, Mycobacterium hassicum, Mycobacterium intracellulare, Mycobacterium interjectum, Mycobacterium heidelberense, Mycobacterium lentiflavum, Mycobacterium malmoense, Mycobacterium microgenicum, Mycobacterium microti, Mycobacterium mucogenicum, Mycobacterium neoaurum, Mycobacterium nonchromogenicum, Mycobacterium peregrinum, Mycobacterium phlei, Mycobacterium scrofulaceum, Mycobacterium shimoidei, Mycobacterium simiae, Mycobacterium szulgai, Mycobacterium terrae, Mycobacterium thermoresistabile, Mycobacterium triplex, Mycobacterium triviale, Mycobacterium tusciae, Mycobacterium ulcerans, Mycobacterium vaccae, Mycobacterium wolinskyi and Mycobacterium xenopi); Haemophilus (e.g. Haemophilus influenzae, Haemophilus ducreyi, Haemophilus aegyptius, Haemophilus parainfluenzae, Haemophilus haemolyticus and Haemophilus parahaemolyticus);

Actinobacillus (e.g. Actinobacillus actinomycetemcomitans, Actinobacillus equuli, Actinobacillus hominis, Actinobacillus lignieresii, Actinobacillus suis and Actinobacillus ureae);

Actinomyces (e.g. Actinomyces israelii);

Brucella (e.g. Brucella abortus, Brucella canis, Brucella melintensis and Brucella suis);

Campylobacter (e.g. Campylobacter jejuni, Campylobacter coli, Campylobacter lari and Campylobacter fetus);

Listeria monocytogenes;

Vibrio (e.g. Vibrio cholerae and Vibrio parahaemolyticus, Vibrio alginolyticus, Vibrio carchariae, Vibrio fluvialis. Vibrio furnissii, Vibrio hollisae, Vibrio metschnikovii, Vibrio mimicus and Vibrio vulnificus);

Erysipelothrix rhusopathiae;

Corynebacteriaceae (e.g. Corynebacterium diphtheriae, Corynebacterium jeikeum and Corynebacterium urealyticum);

Spirochaetaceae, such as Borrelia (e.g. Borrelia recurrentis, Borrelia burgdorferi, Borrelia afzelii, Borrelia andersonii, Borrelia bissettii, Borrelia garinii, Borrelia japonica, Borrelia lusitaniae, Borrelia tanukii, Borrelia turdi, Borrelia valaisiana, Borrelia caucasica, Borrelia crocidurae, Borrelia duttoni, Borrelia graingeri, Borrelia hermsii, Borrelia hispanica, Borrelia latyschewii, Borrelia mazzottii, Borrelia parkeri, Borrelia persica, Borrelia turicatae and Borrelia venezuelensis) and Treponema (Treponema pallidum ssp. pallidum, Treponema pallidum ssp. endemicum, Treponema pallidum ssp. pertenue and Treponema carateum); Pasteurella (e.g. Pasteurella aerogenes, Pasteurella bettyae, Pasteurella canis, Pasteurella dagmatis, Pasteurella gallinarum, Pasteurella haemolytica, Pasteurella multocida multocida, Pasteurella multocida gallicida, Pasteurella multocida septica, Pasteurella pneumotropica and Pasteurella stomatis);

Bordetella (e.g. Bordetella bronchiseptica, Bordetella hinzii, Bordetella holmseii, Bordetella parapertussis, Bordetella pertussis and Bordetella trematum); Nocardiaceae, such as Nocardia (e.g. Nocardia asteroides and Nocardia brasiliensis);

Rickettsia (e.g. Ricksettsii or Coxiella burnetii);

Legionella (e.g. Legionalla anisa, Legionalla birminghamensis, Legionalla bozemanii,

Legionalla cincinnatiensis, Legionalla dumoffii, Legionalla fee!eii, Legionalla gormanii, Legionalla hackeliae, Legionalla israelensis, Legionalla jordanis, Legionalla lansingensis,

Legionalla longbeachae, Legionalla maceachernii, Legionalla micdadei, Legionalla oakridgensis, Legionalla pneumophila, Legionalla sainthelensi, Legionalla tucsonensis and

Legionalla wadsworthii);

Moraxella catarrhalis;

Cyclospora cayetanensis;

Entamoeba histolytica;

Giardia lamblia;

Trichomonas vaginalis;

Toxoplasma gondii;

Stenotrophomonas maltophilia ;

Burkholderia cepacia, Burkholderia mallei and Burkholderia pseudomallei;

Francisella tularensis;

Gardnerella (e.g. Gardneralla vaginalis and Gardneralla mobiluncus);

Streptobacillus moniliformis;

Flavobacteriaceae, such as Capnocytophaga (e.g. Capnocytophaga canimorsus,

Capnocytophaga cynodegmi, Capnocytophaga gingivalis, Capnocytophaga granulosa,

Capnocytophaga haemolytica, Capnocytophaga ochracea and Capnocytophaga sputigena);

Bartonella (Bartonella bacilliformis, Bartonella clarridgeiae, Bartonella elizabethae, Bartonella henselae, Bartonella quintana and Bartonella vinsonii arupensis);

Leptospira (e.g. Leptospira biflexa, Leptospira borgpetersenii, Leptospira inadai, Leptospira interrogans, Leptospira kirschneri, Leptospira noguchii, Leptospira santarosai and Leptospira weilii);

Spirillium (e.g. Spirillum minus);

Baceteroides (e.g. Bacteroides caccae, Bacteroides capillosus, Bacteroides coagulans, Bacteroides distasonis, Bacteroides eggerthii, Bacteroides forsythus, Bacteroides fragilis, Bacteroides merdae, Bacteroides ovatus, Bacteroides putredinis, Bacteroides pyogenes, Bacteroides splanchinicus, Bacteroides stercoris, Bacteroides tectus, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides ureolyticus and Bacteroides vulgatus); Prevotella (e.g. Prevotella bivia, Prevotella buccae, Prevotella corporis, Prevotella dentalis (Mitsuokella dentalis), Prevotella denticola, Prevotella disiens, Prevotella enoeca, Prevotella heparinolytica, Prevotella intermedia, Prevotella loeschii, Prevotella melaninogenica, Prevotella nigrescens, Prevotella oralis, Prevotella oris, Prevotella oulora, Prevotella tannerae, Prevotella venoralis and Prevotella zoogleoformans);

Porphyromonas (e.g. Porphyromonas asaccharolytica, Porphyromonas cangingivalis, Porphyromonas canoris, Porphyromonas cansulci, Porphyromonas catoniae, Porphyromonas circumdentaria, Porphyromonas crevioricanis, Porphyromonas endodontalis, Porphyromonas gingivalis, Porphyromonas gingivicanis, Porphyromonas levli and Porphyromonas macacae);

Fusobacterium (e.g. F. gonadiaformans, F. mortiferum, F. naviforme, F. necrogenes, F. necrophorum necrophorum, F. necrophorum fundiliforme, F. nucleatum nucleatum, F. nucleatum fusiforme, F. nucleatum polymorphum, F. nucleatum vincentii, F. periodonticum, F. russii, F. ulcerans and F. varium);

Chlamydia (e.g. Chlamydia trachomatis);

Cryptosporidium (e.g. C. parvum, C. hominis, C. canis, C. felis, C. meleagridis and C. muris); Chlamydophila (e.g. Chlamydophila abortus (Chlamydia psittaci), Chlamydophila pneumoniae {Chlamydia pneumoniae) and Chlamydophila psittaci {Chlamydia psittaci));

Leuconostoc (e.g. Leuconostoc citreum, Leuconostoc cremoris, Leuconostoc dextranicum, Leuconostoc lactis, Leuconostoc mesenteroides and Leuconostoc pseudomesenteroides); Gemella (e.g. Gemella bergeri, Gemella haemolysans, Gemella morbillorum and Gemella sanguinis); and

Ureaplasma (e.g. Ureaplasma parvum and Ureaplasma urealyticum).

As used herein, the term "fungf (and derivatives thereof, such as "fungal infection") includes, but is not limited to, references to organisms (or infections due to organisms) of the following classes and specific types:

Absidia (e.g. Absidia corymbifera);

Ajeilomyces (e.g. Ajellomyces capsulatus and Ajellomyces dermatitidis);

Arthroderma (e.g. Arthroderma benhamiae, Arthroderma fulvum, Arthroderma gypseum,

Arthroderma incurvatum, Arthroderma otae and Arthroderma vanbreuseghemii);

Aspergillus (e.g. Aspergillus flavus, Aspergillus fumigatus and Aspergillus niger);

Blastomyces (e.g. Blastomyces dermatitidis);

Candida (e.g. Candida albicans, Candida glabrata, Candida guilliermondii, Candida krusei,

Candida parapsilosis, Candida tropicalis and Candida pelliculosa);

Cladophialophora (e.g. Cladophialophora carrionii);

Coccidioides (e.g. Coccidioides immitis and Coccidioides posadasii);

Cryptococcus (e.g. Cryptococcus neoformans);

Cunninghamella (e.g. Cunninghamella sp.) Epidermophyton (e.g. Epidermophyton floccosurn);

Exophiala (e.g. Exophiaia dermatitidis);

Filobasidiella (e.g. Filobasidiella neoformans);

Fonsecaea (e.g. Fonsecaea pedrosoi);

Fusarium (e.g. Fusarium solani);

Geotrichum (e.g. Geotrichum candidum);

Histoplasma (e.g. Histoplasma capsulatum);

Hortaea (e.g. Hortaea werneckii);

Issatschenkia (e.g. Issatschenkia orientalis);

Madurella (e.g. Madurella grisae);

Malassezia (e.g. Malassezia furfur, Malassezia giobosa, Malassezia obtusa, Malassezia pachydermatis, Malassezia restricta, Malassezia slooffiae and Malassezia sympodialis);

Microsporum (e.g. Microsporum canis, Microsporum fulvum and Microsporum gypseum);

Microsporidia;

Mucor (e.g. Mucor circinelloides);

Nectria (e.g. Nectria haematococca);

Paecilomyces (e.g. Paecilomyces variotii);

Paracoccidioides (e.g. Paracoccidioides brasiliensis);

Penicillium (e.g. Penicillium marneffei);

P/cft/a (e.g. P/ch/a anomala and P/cft/a guilliermondii);

Pneumocystis (e.g. Pneumocystis jiroveci (Pneumocystis carinii));

Pseudallescheria (e.g. Pseudallescheria boydii);

Rhizopus (e.g. Rhizopus oryzae);

Rhodotorula (e.g. Rhodotorula rubra);

Scedosporium (e.g. Scedosporium apiospermum);

Schizophyllum (e.g. Schizophyllum commune);

Sporothrix (e.g. Sporothrix schenckii);

Trichophyton (e.g. Trichophyton mentagrophytes, Trichophyton rubrum, Trichophyton verrucosum and Trichophyton violaceum); and

Trichosporon (e.g. Trichosporon asahii, Trichosporon cutaneum, Trichosporon inkin and Trichosporon mucoides).

Particular bacteria that may be treated using the acid addition salts of the present invention include:

Staphylococci, such as Staph, aureus (either Methicillin-sensitive (i.e. MSSA) or Methicillin- resistant (i.e. MRSA)) and Staph, epidermidis; Streptococci, such as Strept. agalactiae and Strept. pyogenes;

Bacillaceae, such as Bacillus anthracis;

Enterobacteriaceae, such as Escherichia coli, Klebsiella (e.g. Klebs. pneumoniae and Klebs. oxytoca) and Proteus (e.g. Pr. mirabilis, Pr. rettgeri and Pr. vulgaris);

Haemophilis influenzae;

Enterococci, such as Enterococcus faecalis and Enterococcus faecium; and

Mycobacteria, such as Mycobacterium tuberculosis.

Preferably, the bacterium is Staph. Aureus; either MSSA or MRSA. In a preferred embodiment of the invention there is provided the use of an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof, for nasal decolonisation of MSSA or MRSA, preferably MRSA.

Particular fungi that may be treated with the acid addition salts of the present invention include Aspergillus fumigatus, Candida albicans, Cryptococcus neoformans, Histoplasma capsulatum and Pneumocystis jiroveci.

Particular conditions which may be treated using the acid addition salts of the present invention include tuberculosis (e.g. pulmonary tuberculosis, non-pulmonary tuberculosis (such as tuberculosis lymph glands, genito-urinary tuberculosis, tuberculosis of bone and joints, tuberculosis meningitis) and miliary tuberculosis), anthrax, abscesses, acne vulgaris, actinomycosis, asthma, bacilliary dysentry, bacterial conjunctivitis, bacterial keratitis, bacterial vaginosis, botulism, Buruli ulcer, bone and joint infections, bronchitis (acute or chronic), brucellosis, burn wounds, cat scratch fever, cellulitis, chancroid, cholangitis, cholecystitis, cutaneous diphtheria, cystic fibrosis, cystitis, diffuse panbronchiolitis, diphtheria, dental caries, diseases of the upper respiratory tract, eczema, empymea, endocarditis, endometritis, enteric fever, enteritis, epididymitis, epiglottitis, erysipelis, erysipelas, erysipeloid, erythrasma, eye infections, furuncles, gardnerella vaginitis, gastrointestinal infections (gastroenteritis), genital infections, gingivitis, gonorrhoea, granuloma inguinale, Haverhill fever, infected burns, infections following dental operations, infections in the oral region, infections associated with prostheses, intraabdominal abscesses, Legionnaire's disease, leprosy, leptospirosis, listeriosis, liver abscesses, Lyme disease, lymphogranuloma venerium, mastitis, mastoiditis, meningitis and infections of the nervous system, mycetoma, nocardiosis (e.g. Madura foot), non-specific urethritis, opthalmia (e.g. opthalmia neonatorum), osteomyelitis, otitis (e.g. otitis externa and otitis media), orchitis, pancreatitis, paronychia, pelveoperitonitis, peritonitis, peritonitis with appendicitis, pharyngitis, phlegmons, pinta, plague, pleural effusion, pneumonia, postoperative wound infections, postoperative gas gangrene, prostatitis, pseudo-membranous colitis, psittacosis, pulmonary emphysema, pyelonephritis, pyoderma (e.g. impetigo), Q fever, rat-bite fever, reticulosis, ricin poisoning, Ritter's disease, salmonellosis, salpingitis, septic arthritis, septic infections, septicameia, sinusitis, skin infections (e.g. skin granulomas, impetigo, folliculitis and furunculosis), syphilis, systemic infections, tonsillitis, toxic shock syndrome, trachoma, tularaemia, typhoid, typhus (e.g. epidemic typhus, murine typhus, scrub typhus and spotted fever), urethritis, wound infections, yaws, aspergillosis, candidiasis (e.g. oropharyngeal candidiasis, vaginal candidiasis or balanitis), cryptococcosis, favus, histoplasmosis, intertrigo, mucormycosis, tinea (e.g. tinea corporis, tinea capitis, tinea cruris, tinea pedis and tinea unguium), onychomycosis, pityriasis versicolor, ringworm and sporotrichosis; or infections with MSSA, MRSA, Staph, epidermidis, Strept. agalactiae, Strept pyogenes, Escherichia coli, Klebs. pneumoniae, Klebs. oxytoca, Pr. mirabilis, Pr. rettgeri, Pr. vulgaris, Haemophilis influenzae, Enterococcus faecalis and Enterococcus faecium.

It will be appreciated that references herein to "treatment" extend to prophylaxis as well as the treatment of established diseases or symptoms.

The acid addition salts of the present invention may be administered alone or in combination with another antimicrobial compound. In one embodiment of the invention there is provided a combination comprising an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3- dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi- succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof, and another antimicrobial agent.

In a further embodiment, the invention provides the use of an acid addition salt of 4-methyl-1- (2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi- tartrate) addition salts thereof, in combination with another antimicrobial agent for the treatment of a microbial infection, preferably for killing clinically latent microorganisms associated with a microbial infection. As used herein, the term "in combination with" covers both separate and sequential administration of an antimicrobial agent and an anesthetic agent. When the agents are administered sequentially, either the antimicrobial agent or the anesthetic agent may be administered first. When administration is simultaneous, the agents may be administered either in the same or a different pharmaceutical composition. Adjunctive therapy, i.e. where one agent is used as a primary treatment and the other agent is used to assist that primary treatment, is also an embodiment of the present invention.

According to a further embodiment of the invention, there is provided a product comprising an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]- quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof and another antimicrobial agent as a combined preparation for simultaneous, separate or sequential use in the treatment of a microbial infection. Suitable antimicrobial compounds for use in combination with the acid addition salts of the present invention include one or more compounds selected from the following:

(1 ) β-Lactams, including:

(i) penicillins, such as

(I) benzylpenicillin, procaine benzylpenicillin, phenoxy-methylpenicillin, methicillin, propicillin, epicillin, cyclacillin, hetacillin, 6-aminopenicillanic acid, penicillic acid, penicillanic acid sulphone (sulbactam), penicillin G, penicillin V, phenethicillin, phenoxymethylpenicillinic acid, azlocillin, carbenicillin, cloxacillin, D-(-)-penicillamine, dicloxacillin, nafcillin and oxacillin,

(II) penicillinase-resistant penicillins (e.g. flucloxacillin),

(III) broad-spectrum penicillins (e.g. ampicillin, amoxicillin, metampicillin and bacampicillin),

(IV) antipseudomonal penicillins (e.g. carboxypenicillins such as ticarcillin or ureidopenicillins such as piperacillin),

(V) mecillinams (e.g. pivmecillinam), or

(VI) combinations of any two or more of the agents mentioned at (I) to (V) above, or combinations of any of the agents mentioned at (I) to (V) above with a β-lactamase inhibitor such as tazobactam or, particularly, clavulanic acid (which acid is optionally in metal salt form, e.g. in salt form with an alkali metal such as sodium or, particularly, potassium); (ii) cephalosporins, such as cefaclor, cefadroxil, cefalexin (cephalexin), cefcapene, cefcapene pivoxil, cefdinir, cefditoren, cefditoren pivoxil, cefixime, cefotaxime, cefpirome, cefpodoxime, cefpodoxime proxetil, cefprozil, cefradine, ceftazidime, cefteram, cefteram pivoxil, ceftriaxone, cefuroxime, cefuroxime axetil, cephaloridine, cephacetrile, cephamandole, cephaloglycine, ceftobiprole, PPI-0903 (TAK-599), 7-aminocephalosporanic acid, 7-aminodes- acetoxycephalosporanic acid, cefamandole, cefazolin, cefmetazole, cefoperazone, cefsulodin, cephalosporin C zinc salt, cephalothin, cephapirin; and

(iii) other β-lactams, such as monobactams (e.g. aztreonam), carbapenems (e.g. imipenem (optionally in combination with a renal enzyme inhibitor such as cilastatin), meropenem, ertapenem, doripenem (S-4661) and RO4908463 (CS-023)), penems (e.g. faropenem) and 1-oxa-P-lactams (e.g. moxalactam).

Tetracyclines, such as tetracycline, demeclocycline, doxycycline, lymecycline, minocycline, oxytetracycline, chlortetracycline, meclocycline and methacycline, as well as glycylcyclines (e.g. tigecycline).

Aminoglycosides, such as amikacin, gentamicin, netilmicin, neomycin, streptomycin, tobramycin, amastatin, butirosin, butirosin A, daunorubicin, dibekacin, dihydrostreptomycin, G 418, hygromycin B, kanamycin B, kanamycin, kirromycin, paromomycin, ribostamycin, sisomicin, spectinomycin, streptozocin and thiostrepton.

(i) Macrolides, such as azithromycin, clarithromycin, erythromycin, roxithromycin, spiramycin, amphotericins B (e.g. amphotericin B), bafilomycins (e.g. bafilomycin A1), brefeldins (e.g. brefeldin A), concanamycins (e.g. concanamycin A), filipin complex, josamycin, mepartricin, midecamycin, nonactin, nystatin, oleandomycin, oligomycins (e.g. oligomycin A, oligomycin B and oligomycin C), pimaricin, rifampicin, rifamycin, rosamicin, tylosin, virginiamycin and fosfomycin.

(ii) Ketolides such as telithromycin and cethromycin (ABT-773).

(iii) Lincosamines, such as lincomycin.

Clindamycin and clindamycin 2-phosphate.

Phenicols, such as chloramphenicol and thiamphenicol.

Steroids, such as fusidic acid (optionally in metal salt form, e.g. in salt form with an alkali metal such as sodium).

Glycopeptides such as vancomycin, teicoplanin, bleomycin, phleomycin, ristomycin, telavancin, dalbavancin and oritavancin. (9) Oxazolidinones, such as linezolid and AZD2563.

(10) Streptogramins, such as quinupristin and dalfopristin, or a combination thereof.

(11) (i) Peptides, such as polymyxins (e.g. colistin and polymyxin B), lysostaphin, duramycin, actinomycins (e.g. actinomycin C and actinomycin D), actinonin, 7- aminoactinomycin D, antimycin A, antipain, bacitracin, cyclosporin A, echinomycin, gramicidins (e.g. gramicidin A and gramicidin C), myxothiazol, nisin, paracelsin, valinomycin and viomycin.

(ii) Lipopeptides, such as daptomycin.

(iii) Lipoglycopeptides, such as ramoplanin.

(12) Sulfonamides, such as sulfamethoxazole, sulfadiazine, sulfaquinoxaline, sulfathiazole (which latter two agents are optionally in metal salt form, e.g. in salt form with an alkali metal such as sodium), succinylsulfathiazole, sulfadimethoxine, sulfaguanidine, sulfamethazine, sulfamonomethoxine, sulfanilamide and sulfasalazine.

(13) Trimethoprim, optionally in combination with a sulfonamide, such as sulfamethoxazole (e.g. the combination co-trimoxazole).

(14) Antituberculous drugs, such as isoniazid, rifampicin, rifabutin, pyrazinamide, ethambutol, streptomycin, amikacin, capreomycin, kanamycin, quinolones (e.g. those at (q) below), para-aminosalicylic acid, cycloserine and ethionamide.

(15) Antileprotic drugs, such as dapsone, rifampicin and clofazimine.

(16) (i) Nitroimidazoles, such as metronidazole and tinidazole.

(ii) Nitrofurans, such as nitrofurantoin.

(17) Quinolones, such as nalidixic acid, norfloxacin, ciprofloxacin, ofloxacin, levofloxacin, moxifloxacin, gatifloxacin, gemifloxacin, garenoxacin, DX-619, WCK 771 (the arginine salt of S-(-)-nadifloxacin), 8-quinolinol, cinoxacin, enrofloxacin, flumequine, lomefloxacin, oxolinic acid and pipemidic acid.

(18) Amino acid derivatives, such as azaserine, bestatin, D-cycloserine, 1 ,10- phenanthroline, 6-diazo-5-oxo-L-norleucine and L-alanyl-L-1-aminoethyl-phosphonic acid.

(19) Aureolic acids, such as chromomycin A3, mithramycin A and mitomycin C.

(20) Benzochinoides, such as herbimycin A.

(21) Coumarin-glycosides, such as novobiocin.

(22) Diphenyl ether derivatives, such as irgasan.

(23) Epipolythiodixopiperazines, such as gliotoxin from Gliocladium fimbriatum.

(24) Fatty acid derivatives, such as cerulenin.

(25) Glucosamines, such as 1-deoxymannojirimycin, 1-deoxynojirimycin and /V-methyl-1- deoxynojirimycin. (26) Indole derivatives, such as staurosporine.

(27) Diaminopyrimidines, such as iclaprim (AR-100).

(28) Macrolactams, such as ascomycin.

(29) Taxoids, such as paclitaxel.

(30) Statins, such as mevastatin.

(31 ) Polyphenolic acids, such as (+)-usnic acid.

(32) Polyethers, such as lasalocid A, lonomycin A, monensin, nigericin and salinomycin.

(33) Picolinic acid derivatives, such as fusaric acid.

(34) Peptidyl nucleosides, such as blasticidine S, nikkomycin, nourseothricin and puromycin.

(35) Nucleosides, such as adenine 9- -D-arabinofuranoside, 5-azacytidine, cordycepin, formycin A, tubercidin and tunicamycin.

(36) Pleuromutilins, such as GSK-565154, GSK-275833 and tiamulin.

(37) Peptide deformylase inhibitors, such as LBM415 (NVP PDF-713) and BB 83698 (38) Antibacterial agents for the skin, such as fucidin, benzamycin, clindamycin, erythromycin, tetracycline, silver sulfadiazine, chlortetracycline, metronidazole, mupirocin, framycitin, gramicidin, neomycin sulfate, polymyxins (e.g. polymixin B) and gentamycin.

(39) Miscellaneous agents, such as methenamine (hexamine), doxorubicin, piericidin A, stigmatellin, actidione, anisomycin, apramycin, coumermycin A1 , L(+)-lactic acid, cytochalasins (e.g. cytochalasin B and cytochalasin D), emetine and ionomycin.

(40) Antiseptic agents, such as chlorhexidine, phenol derivatives (e.g. thymol and triclosan), quarternary ammonium compounds (e.g. benzalkonium chloride, cetylpyridinium chloride, benzethonium chloride, cetrimonium bromide, cetrimonium chloride and cetrimonium stearate), octenidine dihydrochloride, and terpenes (e.g. terpinen-4-ol).

In a further embodiment of the invention, there is provided the use of an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof, as a sterilising agent or as a preservative.

Alternatively, the acid addition salts of the present invention may be employed in methods of sterilisation or preservation, such as: (i) a method of sterilising an object, the method comprising applying to said an acid addition salt of object 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi- succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof; or

(ii) a method of preserving an inorganic or, preferably, an organic material, said method comprising contacting, combining or mixing said material with an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof. In relation to the method described at (i) above, the object is preferably other than a human or animal body. Further, the materials that may be preserved according to the method described at (ii) above include polymers, lubricants, paints, fibres, leather, paper, foodstuffs, water and aqueous mixtures and solutions. Further, when used as a sterilising agent, the compound of the invention may be used either alone or in combination with a conventional sterilising agent. The term "conventional sterilising agent', when used herein, includes references to alcohols (e.g. industrial methylated spirits or ethanol), sodium chloride, thymol, chlorhexidine, cationic surfactants (e.g. cetrimide), iodine (optionally combined with povidone), phenolics (e.g. triclosan), oxidants (e.g. hydrogen peroxide, potassium permanganate or sodium hypochlorite) and any one or more of the conventional antimicrobial agents described above.

The acid addition salts of the present invention may be administered as the raw material but the active ingredients are preferably provided in the form of pharmaceutical compositions. The active ingredients may be used either as separate formulations or as a single combined formulation. When combined in the same formulation it will be appreciated that the two compounds must be stable and compatible with each other and the other components of the formulation. Formulations of the invention include those suitable for oral, parenteral (including subcutaneous e.g. by injection or by depot tablet, intradermal, intrathecal, intramuscular e.g. by depot and intravenous), rectal and topical (including dermal, buccal and sublingual) or in a form suitable for administration by inhalation or insufflation administration. The most suitable route of administration may depend upon the condition and disorder of the patient. Preferably, the compositions of the invention are preferably formulated for oral or topical administration, most preferably for topical administration. In one embodiment of the invention, there is provided a topical composition comprising an acid addition salt of 4- methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi- tartrate) addition salts thereof, for intra-nasal application.

The formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy e.g. as described in "Remington: The Science and Practice of Pharmacy", Lippincott Williams and Wilkins, 21^st Edition, (2005). Suitable methods include the step of bringing into association to active ingredients with a carrier which constitutes one or more excipients. In general, formulations are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers or finely divided solid carriers or both and then, if necessary, shaping the product into the desired formulation. It will be appreciated that when the two active ingredients are administered independently, each may be administered by a different means.

When formulated with excipients, the active ingredients may be present in a concentration from 0.1 to 99.5% (such as from 0.5 to 95%) by weight of the total mixture; conveniently from 30 to 95% for tablets and capsules and 0.01 to 50% (such as from 3 to 50%) for liquid preparations. A suitable concentration for the acid addition salts of the present invention is from 0.1 to 5% (w/v) of the total mixture.

Formulations suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets (e.g. chewable tablets in particular for paediatric administration), each containing a predetermined amount of active ingredient; as powder or granules; as a solution or suspension in an aqueous liquid or non-aqueous liquid; or as an oil-in-water liquid emulsion or water-in-oil liquid emulsion. The active ingredients may also be presented a bolus, electuary or paste. A tablet may be made by compression or moulding, optionally with one or more excipients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with other conventional excipients such as binding agents (e.g. syrup, acacia, gelatin, sorbitol, tragacanth, mucilage of starch, polyvinylpyrrolidone and/or hydroxymethyl cellulose), fillers (e.g. lactose, sugar, microcrystalline cellulose, maize-starch, calcium phosphate and/or sorbitol), lubricants (e.g. magnesium stearate, stearic acid, talc, polyethylene glycol and/or silica), disintegrants (e.g. potato starch, croscarmellose sodium and/or sodium starch glycolate) and wetting agents (e.g. sodium lauryl sulphate). Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered active ingredient with an inert liquid diluent. The tablets may be optionally coated or scored and may be formulated so as to provide controlled release (e.g. delayed, sustained, or pulsed release, or a combination of immediate release and controlled release) of the active ingredients. Alternatively, the active ingredients may be incorporated into oral liquid preparations such as aqueous or oily suspensions, solutions, emulsions, syrups or elixirs. Formulations containing the active ingredients may also be presented as a dry product for constitution with water or another suitable vehicle before use. Such liquid preparations may contain conventional additives such as suspending agents (e.g. sorbitol syrup, methyl cellulose, glucose/sugar syrup, gelatin, hydroxymethyl cellulose, carboxymethyl cellulose, aluminium stearate gel and/or hydrogenated edible fats), emulsifying agents (e.g. lecithin, sorbitan mono-oleate and/or acacia), non-aqueous vehicles (e.g. edible oils, such as almond oil, fractionated coconut oil, oily esters, propylene glycol and/or ethyl alcohol), and preservatives (e.g. methyl or propyl p-hydroxybenzoates and/or sorbic acid).

Topical compositions, which are useful for treating disorders of the skin or of membranes accessible by digitation (such as membrane of the mouth, vagina, cervix, anus and rectum), include creams, ointments, lotions, sprays, gels and sterile aqueous solutions or suspensions. As such, topical compositions include those in which the active ingredients are dissolved or dispersed in a dermatological vehicle known in the art (e.g. aqueous or nonaqueous gels, ointments, water-in-oil or oil-in-water emulsions). Constituents of such vehicles may comprise water, aqueous buffer solutions, non-aqueous solvents (such as ethanol, isopropanol, benzyl alcohol, 2-(2-ethoxyethoxy)ethanol, propylene glycol, propylene glycol monolaurate, glycofurol or glycerol), oils (e.g. a mineral oil such as a liquid paraffin, natural or synthetic triglycerides such as Miglyol™, or silicone oils such as dimethicone). Depending, inter alia, upon the nature of the formulation as well as its intended use and site of application, the dermatological vehicle employed may contain one or more components selected from the following list: a solubilising agent or solvent (e.g. a β-cyclodextrin, such as hydroxypropyl β-cyclodextrin, or an alcohol or polyol such as ethanol, propylene glycol or glycerol); a thickening agent (e.g. hydroxymethyl cellulose, hydroxypropyl cellulose, carboxymethyl cellulose or carbomer); a gelling agent (e.g. a polyoxyethylene- polyoxypropylene copolymer); a preservative (e.g. benzyl alcohol, benzalkonium chloride, chlorhexidine, chlorbutol, a benzoate, potassium sorbate or EDTA or salt thereof); and pH buffering agent(s) (e.g. a mixture of dihydrogen phosphate and hydrogen phosphate salts, or a mixture of citric acid and a hydrogen phosphate salt). Topical formulations may also be formulated as a transdermal patch.

Methods of producing topical pharmaceutical compositions such as creams, ointments, lotions, sprays and sterile aqueous solutions or suspensions are well known in the art. Suitable methods of preparing topical pharmaceutical compositions are described, e.g. in WO9510999, US6974585, WO2006048747, as well as in documents cited in any of these references.

Topical pharmaceutical compositions according to the present invention may be used to treat a variety of skin or membrane disorders, such as infections of the skin or membranes (e.g. infections of nasal membranes, axilla, groin, perineum, rectum, dermatitic skin, skin ulcers, and sites of insertion of medical equipment such as i.v. needles, catheters and tracheostomy or feeding tubes) with any of the bacteria, fungi described above, (e.g. any of the Staphylococci, Streptococci, Mycobacteria or Pseudomonas organisms mentioned hereinbefore, such as S. aureus (e.g. Methicillin resistant S. aureus (MRSA))).

Particular bacterial conditions that may be treated by topical pharmaceutical compositions of the present invention also include the skin- and membrane-related conditions disclosed hereinbefore, as well as: acne vulgaris; rosacea (including erythematotelangiectatic rosacea, papulopustular rosacea, phymatous rosacea and ocular rosacea); erysipelas; erythrasma; ecthyma; ecthyma gangrenosum; impetigo; paronychia; cellulitis; folliculitis (including hot tub folliculitis); furunculosis; carbunculosis; staphylococcal scalded skin syndrome; surgical scarlet fever; streptococcal peri-anal disease; streptococcal toxic shock syndr ome; pitted keratolysis; trichomycosis axillaris; pyoderma; external canal ear infections; green nail syndrome; spirochetes; necrotizing fasciitis; Mycobacterial skin infections (such as lupus vulgaris, scrofuloderma, warty tuberculosis, tuberculides, erythema nodosum, erythema induratum, cutaneous manifestations of tuberculoid leprosy or lepromatous leprosy, erythema nodosum leprosum, cutaneous M. kansasii, M. malmoense, M. szulgai, M. simiae, M. gordonae, M. haemophilum, M. avium, M. intracellulare, M. chelonae (including M. abscessus) or M. fortuitum infections, swimming pool (or fish tank) granuloma, lymphadenitis and Buruli ulcer (Bairnsdale ulcer, Searles' ulcer, Kakerifu ulcer or Toro ulcer)); as well as infected eczma, burns, abrasions and skin wounds.

Particular fungal conditions that may be treated by topical pharmaceutical compositions of the present invention also include include the skin- and membrane-related conditions disclosed hereinbefore, as well as: candidiasis; sporotrichosis; ringworm (e.g. tinea pedis, tinea cruris, tinea capitis, tinea unguium or tinea corporis); tinea versicolor; and infections with Trichophyton, Microsporum, Epidermophyton or Pityrosporum ovale fungi.

Compositions for use according to the invention may be presented in a pack or dispenser device which may contain one or more unit dosage forms containing the active ingredients. The pack may, e.g. comprise metal or plastic foil, such as a blister pack. Where the compositions are intended for administration as two separate compositions these may be presented in the form of a twin pack.

Pharmaceutical compositions may also be prescribed to the patient in "patient packs" containing the whole course of treatment in a single package, usually a blister pack. Patient packs have an advantage over traditional prescriptions, where a pharmacist divides a patients' supply of a pharmaceutical from a bulk supply, in that the patient always has access to the package insert contained in the patient pack, normally missing in traditional prescriptions. The inclusion of the package insert has been shown to improve patient compliance with the physician's instructions.

The administration of the combination of the invention by means of a single patient pack, or patients packs of each composition, including a package insert directing the patient to the correct use of the invention is a desirable feature of this invention.

According to a further embodiment of the present invention there is provided a patient pack comprising an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi-tartrate) addition salts thereof, and an information insert containing directions on the use of the combination of the invention. In another embodiment of the invention, there is provided a double pack comprising in association for separate administration, an antimicrobial agent, preferably having biological activity against clinically latent microorganisms, and an acid addition salt of 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid (preferably hemi-succinate), sulphuric acid and tartaric acid (preferably hemi- tartrate) addition salts thereof. The amount of active ingredients required for use in treatment will vary with the nature of the condition being treated and the age and condition of the patient, and will ultimately be at the discretion of the attendant physician or veterinarian. In general however, doses employed for adult human treatment will typically be in the range of 0.02 to 5000 mg per day, preferably 1 to 1500 mg per day. The desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, e.g. as two, three, four or more sub- does per day.

Experimental Methods

1 H NMR

NMR spectra were collected on a Bruker 270 MHz instrument equipped with an auto-sampler and controlled by a DRX400 console. Automated experiments were acquired using Delta NMR Processing & Control Software version 4.3. Samples were prepared in d6-DMSO, unless otherwise stated. Analysis was carried out using (ACD/Specmanager 7.11). XRPD

X-Ray Powder Diffraction patterns were collected on a PANalytical diffractometer using Cu Ka radiation (45kV, 40mA), .Θ-Θ goniometer, focusing mirror, divergence slit (1/2"), soller slits at both incident and divergent beam (4mm) and a PIXcel detector. The software used for data collection was X'Pert Data Collector, version 2.2f and the data was presented using X'Pert Data Viewer, version 1.2d.

Samples were run under ambient conditions and were analysed by transmission foil XRPD, using the powder as received. Approximately 2-5 mg of the sample was mounted on a 96 position sample plate supported on a polyimide (Kapton, 12.7 μηη thickness) film. Data was collected in the range 3 - 40° 2Θ with a continuous scan (speed of 0.146 ⁰ 20/s). DSC

DSC data was collected on a PerkinElmer Pyris 4000 DSC. The instrument was verified for energy and temperature calibration using certified indium. A predefined amount in milligrams (mg) of the sample was placed in a pin holed aluminium pan and typically heated at 20° C.min^"1 from 30°C to 350° C, or varied as experimentation dictated. The instrument control and data analysis was Pyris Software v9.0.1.0174.

HPLC

System: Agilent 1100/1200 series liquid chromatograph or equivalent

Column: Zorbax Eclipse XDB-C18, 4.6 x 150 mm,

5pm particle size (ex Agilent; #993967-902)

Mobile Phase: A - 0.01 M Ammonium Acetate Buffer (pH 8.0)

B - Acetonitrile

Flow Rate: 1.0 ml/min

Injection Volume: 10 μΙ

Detection: UV @ 254 nm

Column Temperature: 40 °C

Post Run: 5 min

Gradient: Time (min) %A %B

0 95 5

5 95 5

15 5 95

25 5 95

27 95 5

Gravimetric Vapour Sorption (GVS)

Sorption isotherms were obtained using a Hiden Isochema moisture sorption analyser (model IGAsorp), controlled by IGAsorp Systems Software V6.50.48. The sample was maintained at a constant temperature (25 °C) by the instrument controls. The humidity was controlled by mixing streams of dry and wet nitrogen, with a total flow of 250 ml. min^"1. The instrument was verified for relative humidity content by measuring three calibrated Rotronic salt solutions (10 - 50 - 88%). The weight change of the sample was monitored as a function of humidity by a microbalance (accuracy +/- 0.005 mg). A defined amount of sample was placed in a tarred mesh stainless steel basket under ambient conditions. A full experimental cycle consisted of two scans (sorption and desorption) at a constant temperature (25 °C) and 10 % RH intervals over a 10 - 90 % range (90 minutes for each humidity level). This type of experiment should demonstrate the ability of samples studied to absorb moisture (or not) over a set of well determined humidity ranges. The following non-limiting examples illustrate the present invention. Example 1

Synthesis of 4-methyl-1 -(2-phenylethyI)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane sulphonate

Potassium carbonate (1638 g) was dissolved in water (2453 mL) at room temperature. Ethyl acetate (4095 mL) and methanol (410 mL) were added and the solvents were degassed. 4- methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hydrochloride (450 g) was added and the reaction mixture stirred until two clear phases were achieved (-20 minutes). The phases were separated and the organic phase was dried over magnesium sulfate filtered and placed under nitrogen. Methane sulfonic acid (70.0 mL) was added whilst maintaining the temperature <30°C and the resulting reaction mixture was stirred at room temperature overnight. The solid was filtered, washed with ethyl acetate (3 x 630mL) and pulled dry. Further drying of the solid in a vacuum oven at 40°C yielded the desired product (457.6g). H NMR (270MHz, CDCI₃): 8.37 (1 H, d, J=9.2Hz), 7.45-7.55 (1 H, m), 7.35-7.45 (3H, m), 7.1-7.3 (4H, m), 6.92-7.1 (4H, m), 3.7-3.9 (4H, m), 3.02 (2H, t, J=9.6Hz), 2.88 (3H, s and 2H, t, 7.2Hz), 2.60 (3H, s).

Example 2

Synthesis of 4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane suiphonate

4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hydrochloride (30 g) was suspended in ethyl acetate (450 mL) and saturated aqueous K₂C0₃ (300 mL) added. The mixture was stirred vigorously until all of the solid had dissolved (1-2 hours), then the layers were separated and the aqueous layer extracted with ethyl acetate (100 mL). The combined organics were dried (MgS0₄), filtered, and concentrated in vacuo to give 4- methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline free base (26.3 g).

The free base was dissolved/suspended in ethyl acetate (250 mL), then a solution of methane sulfonic acid (7.3 g, 76 mmol) in methanol (76 mL) was added and the reaction mixture stirred at room temperature. After approximately 1 hour a suspension began to form. The mixture was stirred overnight, filtered, and the solid washed with ethyl acetate (25 mL), sucked dry then dried in vacuo at 50 °C overnight to give the desired product {22.6 g) as a pale yellow solid. Purity by HPLC: 99.72%. MS: 381.4 (M+).

Example 3

Synthesis of 4-methyM -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane sulphonate

K₂C0₃ (1638 g) was dissolved in water (2453 mL) at room temperature. Ethyl acetate (4095 mL) and methanol (410 ml_) were added and the solvents were degassed. 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hydrochloride (450 g) was added and the reaction stirred until 2 clear phases were achieved (~20 minutes). The phases were separated and the organic phase was dried over MgS0₄, filtered and placed under nitrogen. Methane sulfonic acid (70.0 mL) was added whilst maintaining the temperature <30°C and the resulting reaction stirred at room temperature overnight. The solid was filtered, washed with ethyl acetate (3 x 630 mL) and pulled dry. Further drying of the solid in a vacuum oven at 40°C yielded the desired product (457.6g).

Example 4

Synthesis of 4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline hemi-succinate

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline free base (500 mg) was dissolved in ethyl acetate (2.5 mL) and THF (0.5 mL) then a solution of succinic acid (85 mg) in methanol (0.73 mL) was added and the reaction mixture stirred at room temperature. After approximately 1 hour a suspension began to form. The mixture was stirred overnight then filtered and the solid washed with ethyl acetate (1 mL), sucked dry then dried in vacuo at 50 °C overnight to give the desired product (363 mg) as a pale yellow solid. Purity by HPLC: 99.39%. Melting point by DSC: 151 °C (peak); 149 °C (onset). MS: 381.0 (M+). A representative ¹H NMR spectrum for 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3- dihydro-1 H-pyrrolo[3,2-c]-quinoline hemi-succinate is reproduced in Figure 4.

Example 5

Synthesis of 4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline p-toluene sulphonate

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline p-toluene sulphonate was prepared by mixing 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H- pyrrolo[3,2-c]-quinoline (0.6 g) with p-toluene sulphonic acid (1.75 ml / 1 M) in 5 volumes of ethyl acetate using an analogous method to that described in Example 4. Melting point by DSC: 176 °C (onset). Representative ¹H NMR and XRPD spectra for 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]-quinoline p-toluene sulphonate are reproduced in Figures 5 and 6 respectively.

Example 6

Synthesis of 4-methyl-1 -(2-phenyIethyI)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline hydrobromide

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hydrobromide was prepared by mixing 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline (0.66 g) with hydrobromic acid (1.90 ml / 1 M) in 5 volumes of iso-propyl alcohol using an analogous method to that described in Example 4. Melting point by DSC: 237 °C (main peak); 212 ^QC (small endotherm). Example 7

Synthesis of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]- quinoline sulphate

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline sulphate was prepared by mixing 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline (0.64 g) with sulphuric acid (1.79 ml / 1 M) in 5 volumes of iso-propyl alcohol using an analogous method to that described in Example 4. Melting point by DSC: 285 °C (main peak); 150 °C (small endotherm).

Example 8

Synthesis of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]- quinoline hemi-tartrate

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hemi-tartrate was prepared by mixing 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline (100 mg) with tartaric acid (0.6 eq. / 1 M) in 5 volumes of ethyl acetate using an analogous method to that described in Example 4. Melting point by DSC: 181 °C (main peak); 108 °C (small endotherm). A representative ¹H NMR spectrum for 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hemi-tartrate is reproduced in Figure 7. Solubility Studies A solubility study of various acid addition salts of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3- dihydro-1 H-pyrrolo[3,2-c]-quinoline was undertaken using HPLC grade water in order to rank the salts identified. A 20 mg sample of the relevant 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline salt was added to 1 mL of HPLC grade water at 25 °C and stirred at a constant rate for 15 minutes. If all of the material dissolved within this period, further salt was added in 20 mg portions up to a total of 140 mg. The resulting mixtures were stirred at a constant rate overnight at 25 °C and those containing solid were filtered. The filtrates were analysed for solubility by HPLC. The results are shown in the table below.

^*A saturated suspension was not formed within the bounds of this experiment. A solubility study of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]- quinoline methane sulphonate was undertaken at various pHs and at different saline (NaCI) concentrations. The aim of the study was to determine the solubility of the methane sulphonate salt under conditions similar to those experienced in the nasal cavity in vivo. A 50 mg sample of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane sulphonate was added to 1 mL of buffer (see Table 1 for details) at 25 °C and stirred for 15 minutes. If all the material had dissolved further salt was added in 50 mg portions up to a total of 200 mg. The resulting mixtures were stirred overnight at 25 °C then those containing solid were filtered. The filtrates were analysed for solubility.

The results are shown in the table below, with pH measured on final suspensions and solubility assessed by HPLC.

These data show that there was a significant variation in solubility with pH and a significant decrease in solubility when the NaCI concentration was increased from 0.1 to 1%. A second series of solubility studies were subsequently carried out on the methane sulphonate salt. Measurement of the solubility at 37 °C showed a significant increase compared to 25 °C, i.e. 228 mg/mL compared to 120 mg/mL

Measurement of solubility at increasing pH was also measured. A 500 mg sample of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane sulphonate was added to 1 mL of distilled water at 25 °C and stirred for 15 minutes. The pH was adjusted to the appropriate value by the addition of aqueous NaOH (0.001 to 0.1 M), the resulting mixtures were stirred overnight at 25 °C then those containing solid were filtered. The filtrates were analysed for solubility.

The results from this series are summarised in the table below, with pH measured on final suspensions and solubility assessed by HPLC.

In most cases the material had dissolved after an overnight stir out. In the cases where solid remained (pH 4-5), the solubility remained very high across the range with no return of free base as a suspension.

Following on from these results a further solubility study was carried out, in increasing concentrations of saline, adjusted to pH 6 at 33 °C.

A 500 mg sample of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane sulphonate was added to 0.75 mL of the appropriate saline solution at 33 °C and stirred for 15 minutes. The pH was adjusted to 6 by the addition of aqueous NaOH (0.1 M), the resulting mixtures were stirred overnight at 25 °C and those containing solid were filtered. The filtrates were analysed for solubility. The results are summarised in the table below.

0.9% Suspension No Change 98.8

1.1 % Suspension No Change 31.4

These results show a broad correlation between the amount of NaCI added to the mixture and the solubility obtained, with a significant decrease once the level of NaCI added exceeds 0.9%. The solubility measurements are not entirely linear which may be accounted for by the difficulties encountered with assessing the pH of thick suspensions on a small scale. However a clear trend is shown.

Stability Studies

1 % and 5% solutions of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane sulphonate were prepared in de-ionised water. These solutions were then stored exposed to air and natural light for 2 weeks. After two weeks the samples were re-analysed and the 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]- quinoline methane sulphonate was found to be stable with no increase in the key oxidised impurity. The results are summarised in the table below.

Hyqroscopicitv Studies

The hydrobromide, methane sulphonate, sulphate and p-toluene sulphonate salts of 4- methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline were analysed by GVS according to the general protocol described herein to monitor their ability to adsorb moisture. 4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline hydrochloride was employed as a reference material. None of the salts tested were found to be hygroscopic.

Biological Data

In vitro efficacy test of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H- pyrrolo[3,2-c]-quinoline free base, hydrochloride and methane sulphonate salts against stationary phase Staphylococcus aureus Bacterial strain

Staphylococcus aureus (Oxford); Gram positive; Reference strain Growth of bacteria

Log phase growth of methicilin sensitive Staphylococcus aureus (MSSA) was carried out as follows:

Bacteria were grown in 10 ml of nutrient broth (No. 2, (Oxoid)) overnight at 37°C with continuous shaking at 120 rpm. The overnight cultures were diluted 1000 X with iso-sensitest broth. The cultures were incubated at 37°C with shaking for 1-2 hours to reach log CFU 6. Viability of the bacteria was estimated by colony forming unit (CFU) counts. From serial 10- fold dilutions of the experimental cultures, 100 μΙ samples were added to triplicate plates of nutrient agar plates (Oxoid) or blood agar plates (Oxoid). CFUs were counted after incubation of the plates at 37°C for 24 hours.

Bacterial cultures: 10 μΙ of overnight cultures was added to 10 ml of fresh iso-sensitest broth to make the inoculation to 10⁶ CFU/ml. 290 μΙ of the cell suspension was added to each well of the 96 well plate, which was incubated at 37°C for 24 hours. 300 μΙ of iso-sensitest broth without bacterial cells were added to wells of the plate as a no bacterial control.

MIC determination: The optical density of the bacterial cells was read at 405 nm using a plate reader (Bio TEK). The MIC concentration was determined as the lowest concentration of drug which inhibits the bacterial growth.

Compounds and Preparation

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline methane sulphonate was dissolved in H₂0 at 100 mg/ml. The compound was further diluted with water to final concentrations from 32 to 0.5 Mg/ml for MIC and stationary phase kill.

4-methyl-1-(2-phenyletby|)-8-ph methane sulphonate hydrochloride was dissolved in DMSO to 10 mg/ml.

4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline free base was dissolved in DMSO to 10 mg/ml. The compound was further diluted with DMSO to final concentrations from 32 to 0.5 pg/ml. Results (1 ) Methane sulphonate (mesylate) salt

(2) Hydrochloride salt and free base

Conclusion

The methane sulphonate salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline was found to have the same MIC (8 Mg/ml) as the free base and the hydrochloride salt.

Claims

Claims

1. An acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid, sulphuric acid and tartaric acid addition salts thereof.

2. 4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline

methane sulphonate.

3. 4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline

methane sulphonate according to claim 2 characterised by at least one of the following:

(i) a ¹H NMR spectrum (270MHz, CDCI₂) with peaks at 8.37 (1H, d, J = 9.2 Hz), 7.45-7.55 (1 H, m), 7.35-7.45 (3H, m), 7.1-7.3 (4H, m), 6.92-7.1 (4H, m), 3.7-3.9 (4H, m), 3.02 (2H, t, J = 9.6 Hz), 2.88 (3H, s and 2H, t, 7.2Hz) and 2.60 (3H, s); and/or

(ii) a ¹H NMR spectrum (270MHz, D_fi-DMSO) with peaks at 7.86 (1 H, d, J=9.2Hz), 7.68 (1 H, dd, J=2.7Hz, 9.4Hz), 7.58 (1 H, d, 2.2Hz), 7.38-7.51 (2H, m), 7.08-7.28 (6H, m), 6.97-7.06 (2H, m), 3.7-4.1 (4H, m), 3.08 (2H, t, J=9.6Hz), 2.88 (2H, t, 7.4Hz), 2.46 (3H, s), 2.30 (3H, s); and/or

(iii) an XRPD pattern with peaks at 8.33, 9.33, 11.92, 14.52, 14.99, 16.59, 16.79, 17.07, 17.23, 17.37, 18.32, 18.86, 19.46 ±0.2 ° 2Θ; and/or

(iv) a melting point of from about 178 to about 181 °C.

4. A pharmaceutical composition comprising an acid addition salt of 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1H-pyrrolo[3,2-c]-quinoline according to claim 1 and a pharmaceutically acceptable carrier.

5. A pharmaceutical composition according to claim 4 for topical administration.

6. A process for preparing an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy- 2,3-dihydro-1H-pyrrolof3,2-c]-quinoline according to claim 1 , which process comprises:

(i) dissolving or suspending 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3- dihydro-1 H-pyrrolo[3,2-c]-quinoline free base in a suitable solvent;

(ii) treating the resulting solution or suspension with an acid selected from the group consisting of hydrobromic acid, methane sulphonic acid, p- toluene sulphonic acid, succinic acid, sulphuric acid or tartaric acid; and

(iii) optionally isolating the compound so formed.

7. A process for preparing an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy- 2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline according to claim 1 , which process comprises:

(i) dissolving or suspending an acid addition salt of 4-methyl-1-(2- phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline, preferably the hydrochloride salt thereof, in a suitable solvent;

(ii) treating the resulting mixture with a suitable base;

(iii) treating the resulting solution or suspension with an acid selected from the group consisting of hydrobromic acid, methane sulphonic acid, p- toluene sulphonic acid, succinic acid, sulphuric acid or tartaric acid; and

(iv) optionally isolating the compound so formed.

8. Use of an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline according to claim 1 for the treatment of an microbial infection.

9. Use according to claim 8, for killing multiplying and/or clinically latent microorganisms associated with a microbial infection.

10. A method of treating a microbial infection, which comprises administering to a mammal, including man, an acid addition salt of 4-methyl-1-(2-phenylethyl)-8- phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline according to claim 1.

11. Use of an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline according to claim 1 for the treatment of tuberculosis (pulmonary tuberculosis, non-pulmonary tuberculosis (such as tuberculosis lymph glands, genito-urinary tuberculosis, tuberculosis of bone and joints, tuberculosis meningitis) and miliary tuberculosis), anthrax, abscesses, acne vulgaris, actinomycosis, asthma, bacilliary dysentry, bacterial conjunctivitis, bacterial keratitis, bacterial vaginosis, botulism, Buruli ulcer, bone and joint infections, bronchitis (acute or chronic), brucellosis, burn wounds, cat scratch fever, cellulitis, chancroid, cholangitis, cholecystitis, cutaneous diphtheria, cystic fibrosis, cystitis, diffuse panbronchiolitis, diphtheria, dental caries, diseases of the upper respiratory tract, eczema, empymea, endocarditis, endometritis, enteric fever, enteritis, epididymitis, epiglottitis, erysipelis, erysipelas, erysipeloid, erythrasma, eye infections, furuncles, gardnerella vaginitis, gastrointestinal infections (gastroenteritis), genital infections, gingivitis, gonorrhoea, granuloma inguinale, Haverhill fever, infected burns, infections following dental operations, infections in the oral region, infections associated with prostheses, intraabdominal abscesses, Legionnaire's disease, leprosy, leptospirosis, listeriosis, liver abscesses, Lyme disease, lymphogranuloma venerium, mastitis, mastoiditis, meningitis and infections of the nervous system, mycetoma, nocardiosis (Madura foot), non-specific urethritis, opthalmia (opthalmia neonatorum), osteomyelitis, otitis (otitis externa and otitis media), orchitis, pancreatitis, paronychia, pelveoperitonitis, peritonitis, peritonitis with appendicitis, pharyngitis, phlegmons, pinta, plague, pleural effusion, pneumonia, postoperative wound infections, postoperative gas gangrene, prostatitis, pseudo-membranous colitis, psittacosis, pulmonary emphysema, pyelonephritis, pyoderma (impetigo), Q fever, rat-bite fever, reticulosis, ricin poisoning, Ritter's disease, salmonellosis, salpingitis, septic arthritis, septic infections, septicameia, sinusitis, skin infections (skin granulomas, impetigo, folliculitis and furunculosis), syphilis, systemic infections, tonsillitis, toxic shock syndrome, trachoma, tularaemia, typhoid, typhus (epidemic typhus, murine typhus, scrub typhus and spotted fever), urethritis, wound infections, yaws, aspergillosis, candidiasis (oropharyngeal candidiasis, vaginal candidiasis or balanitis), cryptococcosis, favus, histoplasmosis, intertrigo, mucormycosis, tinea (tinea corporis, tinea capitis, tinea cruris, tinea pedis and tinea unguium), onychomycosis, pityriasis versicolor, ringworm and sporotrichosis; or infections with MSSA, MRSA, Staph. epidermidis, Strept. agalactiae, Strept. pyogenes, Escherichia coli, Klebs. pneumoniae, Klebs. oxytoca, Pr. mirabilis, Pr. rettgeri, Pr. vulgaris, Haemophilis influenzae, Enterococcus faecalis and Enterococcus faecium.

12. Use of an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline according to claim 1 for nasal decolonisation of MSSA or MRSA, preferably MRSA.

13. Use of an acid addition salt of 4-methyl-1-(2-phenylet yl)-8-phenoxy-2,3-dihydro-1 H- pyrrolo[3,2-c]-quinoline according to claim 1 as a sterilising agent or as a preservative.

14. An acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy-2,3-dihydro-1H- pyrrolo[3,2-c]-quinoline selected from the group consisting of the hydrobromic acid, methane sulphonic acid, p-toluene sulphonic acid, succinic acid, sulphuric acid and tartaric acid addition salts thereof for the treatment of microbial infections.

15. 4-methyl-1 -(2-phenylethyl)-8-phenoxy-2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline

methane sulphonate for the treatment of microbial infections.

16. A product comprising an acid addition salt of 4-methyl-1-(2-phenylethyl)-8-phenoxy- 2,3-dihydro-1 H-pyrrolo[3,2-c]-quinoline according to claim 1 and another antimicrobial agent as a combined preparation for simultaneous, separate or sequential use in the treatment of a microbial infection.