Compounds for use as medicaments

Abstract

Compounds for use in the treatment of Clostridium difficile infection A compound of formula (I), or a pharmaceutically acceptable salt, solvate or prodrug thereof, for use as medicament, wherein each

Description

Field of the Invention Described herein are compounds for use as medicaments, as well as compositions 5 comprising said compounds. Background of the Invention Bacterial infections are prevalent around the world. Of particular concern are infections that are picked up by patients whilst in a healthcare environment. These are io typically known as nosocomial infections, or healthcare-associated infections. Many nosocomial infections are becoming more difficult to treat due to increasing resistance by the bacteria to conventional antibiotics. For example, Clostridioid.es difficile (C. difficile) is a nosocomial pathogen that causes 15 significant mortality and morbidity globally. C. difficile primarily affects patients who have been treated with broad spectrum antibiotics for an unrelated condition, resulting in damage to the gut microbiome and a resultant loss of colonisation resistance to C. difficile, with patients who receive longer courses of therapy being at greater risk than those receiving short courses. 20 C. difficile produces 4-methylphenol (p-cresol) by the fermentation of 4-hydroxyphenylalanine (p-tyrosine) via the intermediate 4-hydroxyphenylacetic acid (p-hydroxyphenylacetic acid (p-HPA)) or by the conversion of exogenous 4-hydroxyphenylactetic acid. Both pathways utilise the action of HpdBCA decarboxylase, 25 which is encoded by the hpdBCA operon. 4-methylphenol selectively inhibits growth of Gram-negative bacteria of the Gammaproteobacteria class, including Escherichia coli, Proteus mirabillis and Klebsiella oxytoca. The production of 4-methylphenol by C. difficile in the human gut may, therefore, reduce gut microbial diversity and allow for the proliferation of C. difficile, which can result in sustained or recurrent C. difficile 30 infections. Less than 1% of the currently sequenced bacteria in the gut microbiome encode the decarboxylase required to make 4-methylphenol from p-HPA. Other bacteria are known to produce 4-methylphenol. Whilst treatment of C. difficile is effective with metronidazole, vancomycin or 35 fidaxomicin, a major feature of C. difficile infection (CDI) is the high proportion of patients (20-30%) who suffer from recurrence (often multiple recurrences) either as a 16 05 25 result of reinfection or relapse. Furthermore, existing antibiotics may have a negative impact on the healthy protective microbiome, which is important in preventing diseases such as cancer and diabetes, as well as conditions that may be linked to the health of the microbiome, such as autism spectrum disorders, for example. 5 Therefore, treatments that prevent the recurrence of C. difficile infections and do not have significant effects on the protective microbiome are in high demand. Summary of the Invention io According to a first aspect of the invention, there is provided a compound of Formula (I): Ri R2 Q^R3 Formula (I) or a pharmaceutically acceptable salt or solvate thereof, for use as a medicament, wherein: 0 15 Q is HO , each of R1 and R2 together represent =0 and R3 is -OH; 0 Q is HO , each of R1 and R2 is -H and R3 is -C(0)NH2, -CH20H, -CN or -CO2CH3; 0 Q is HO ? each of R1 and R2 is -CH3 and R3 is -C02H; or & 20 Q is H3O , each of R1 and R2 is -H and R3 is -C02H. The compound is selected from the group consisting of: 16 05 25 5 The compound is for use in the treatment of a bacterial infection. In some embodiments, the compound is for use in the inhibition of 4-methylphenol production by bacteria. 10 The compound is for use in the treatment of Clostridioid.es difficile infection. According to a second aspect of the invention, a pharmaceutically acceptable salt or solvate of a compound of the first aspect is provided. 15 According to a third aspect of the invention, a pharmaceutical composition comprising a compound of the first aspect, or a pharmaceutically acceptable salt or solvate of the second aspect, and a pharmaceutically acceptable excipient, is provided. Brief Description of the Drawings 20 Figures la to 5b are growth curves for C. difficile 63oAerm and E. coli with compounds disclosed herein; Figure 6 shows viable cell data, determined by colony forming unit (CFU), for C. difficile, a mutant of C. difficile and its complemented strain when grown with E. coli; Figures 7 to 13 show CFU data for C. difficile and E. coli when treated with the 25 compounds described herein; and Figures 14 and 15 are HPLC data. Detailed Description of the Invention A first aspect of the invention provides a compound of Formula (I): Ri R2 Q^R3 Formula I or a pharmaceutically acceptable salt or solvate thereof, for use as a medicament, 5 wherein: Q is HO , each of R1 and R2 together represent =0 and R3is -OH; 16 05 25 Q is HO , each of R1 and R2 is -H and R3 is -C(0)NH2, -CH20H, -CN or -CO2CH3; 10 Q is HO ? each of R1 and R2 is -CH3 and R3 is -C02H; or Q is H3O , each of R1 and R2 is -H and R3 is -C02H. It has been found that these compounds inhibit the growth of bacteria. The compounds 15 are for the inhibition of the growth of the bacterium C. difficile. The compounds are for use in the treatment of C. difficile infections. An “alkyl” substituent group or an alkyl moiety in a substituent group may be linear (i.e. straight-chained) or branched. Examples of alkyl groups / moieties include 20 methylene (-CH2-), methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl, t-butyl and n-pentyl groups / moieties. Unless stated otherwise, the term “alkyl” does not include “cycloalkyl”. Typically an alkyl group is a Ci-Ci2 alkyl group. More typically an alkyl 16 05 25 group is a Ci-Ce alkyl group. An “alkylene” group is similarly defined as a divalent alkyl group. An “alkenyl” substituent group or an alkenyl moiety in a substituent group refers to an 5 unsaturated alkyl group or moiety having one or more carbon-carbon double bonds. Examples of alkenyl groups / moieties include ethenyl, propenyl, 1-butenyl, 2-butenyl, 1-pentenyl, 1-hexenyl, 1,3-butadienyl, 1,3-pentadienyl, 1,4-pentadienyl and 1,4-hexadienyl groups / moieties. Unless stated otherwise, the term “alkenyl” does not include “cycloalkenyl”. Typically an alkenyl group is a C2-C12 alkenyl group. More 10 typically an alkenyl group is a C2-C6 alkenyl group. An “alkenylene” group is similarly defined as a divalent alkenyl group. An “alkynyl” substituent group or an alkynyl moiety in a substituent group refers to an unsaturated alkyl group or moiety having one or more carbon-carbon triple bonds. 15 Examples of alkynyl groups / moieties include ethynyl, propargyl, but-2-ynyl and but-2-ynyl. Typically an alkynyl group is a C2-C12 alkynyl group. More typically an alkynyl group is a C2-C6 alkynyl group. An “alkynylene” group is similarly defined as a divalent alkynyl group. 20 A “cyclic” substituent group or a cyclic moiety in a substituent group refers to any hydrocarbyl ring, wherein the hydrocarbyl ring may be saturated or unsaturated (including aromatic) and may include one or more heteroatoms, e.g. N, O or S, in its carbon skeleton. Examples of cyclic groups include cycloalkyl, cycloalkenyl, heterocyclic, aryl and heteroaryl groups as discussed below. A cyclic group may be 25 monocyclic, bicyclic (e.g. bridged, fused or spiro), or polycyclic. Typically, a cyclic group is a 3- to 12-membered cyclic group, which means it contains from 3 to 12 ring atoms. More typically, a cyclic group is a 3- to 7-membered monocyclic group, which means it contains from 3 to 7 ring atoms. 30 A “heterocyclic” substituent group or a heterocyclic moiety in a substituent group refers to a cyclic group or moiety including one or more carbon atoms and one or more (such as one, two, three or four) heteroatoms, e.g. N, O or S, in the ring structure. Examples of heterocyclic groups include heteroaryl groups as discussed below and non-aromatic heterocyclic groups such as azetidinyl, azetinyl, tetrahydrofuranyl, pyrrolidinyl, 35 tetrahydrothiophenyl, tetrahydropyranyl, piperidinyl, piperazinyl, morpholinyl, 16 05 25 thiomorpholinyl, oxetanyl, thietanyl, pyrazolidinyl, imidazolidinyl, dioxolanyl, oxathiolanyl, thianyl and dioxanyl groups. A “cycloalkyl” substituent group or a cycloalkyl moiety in a substituent group refers to a 5 saturated hydrocarbyl ring containing, for example, from 3 to 7 carbon atoms, examples of which include cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. Unless stated otherwise, a cycloalkyl substituent group or moiety may include monocyclic, bicyclic or polycyclic hydrocarbyl rings. 10 A “cycloalkenyl” substituent group or a cycloalkenyl moiety in a substituent group refers to a non-aromatic unsaturated hydrocarbyl ring having one or more carboncarbon double bonds and containing, for example, from 3 to 7 carbon atoms, examples of which include cyclopent-i-en-i-yl, cyclohex-i-en-i-yl and cyclohex-i,3-dien-i-yl. Unless stated otherwise, a cycloalkenyl substituent group or moiety may include 15 monocyclic, bicyclic or polycyclic hydrocarbyl rings. An “aryl” substituent group or an aryl moiety in a substituent group refers to an aromatic hydrocarbyl ring. The term “aryl” includes monocyclic aromatic hydrocarbons and polycyclic fused ring aromatic hydrocarbons wherein all of the fused 20 ring systems (excluding any ring systems which are part of or formed by optional substituents) are aromatic. Examples of aryl groups / moieties include phenyl, naphthyl, anthracenyl and phenanthrenyl. Unless stated otherwise, the term “aryl” does not include “heteroaryl”. 25 The term “halo” includes fluoro, chloro, bromo and iodo. Unless stated otherwise, where a group is prefixed by the term “halo”, such as a haloalkyl or halomethyl group, it is to be understood that the group in question is substituted with one or more halo groups independently selected from fluoro, chloro, 30 bromo and iodo. Typically, the maximum number of halo substituents is limited only by the number of hydrogen atoms available for substitution on the corresponding group without the halo prefix. For example, a halomethyl group may contain one, two or three halo substituents. A haloethyl or halophenyl group may contain one, two, three, four or five halo substituents. Similarly, unless stated otherwise, where a group is prefixed by a 35 specific halo group, it is to be understood that the group in question is substituted with one or more of the specific halo groups. For example, the term “fluoromethyl” refers to a methyl group substituted with one, two or three fluoro groups. 16 05 25 Unless stated otherwise, where a group is said to be “halo-substituted”, it is to be 5 understood that the group in question is substituted with one or more halo groups independently selected from fluoro, chloro, bromo and iodo. Typically, the maximum number of halo substituents is limited only by the number of hydrogen atoms available for substitution on the group said to be halo-substituted. For example, a halo-substituted methyl group may contain one, two or three halo substituents. A halo- 10 substituted ethyl or halo-substituted phenyl group may contain one, two, three, four or five halo substituents. Unless stated otherwise, any reference to an element is to be considered a reference to all isotopes of that element. Thus, for example, unless stated otherwise any reference to 15 hydrogen is considered to encompass all isotopes of hydrogen including deuterium and tritium. Q is a substituted 6-membered cyclic group. 20 The cyclic group is heterocyclic. A “heteroaryl” substituent group or a heteroaryl moiety in a substituent group refers to an aromatic heterocyclic group or moiety. The term “heteroaryl” includes monocyclic aromatic heterocycles and polycyclic fused ring aromatic heterocycles wherein all of the 25 fused ring systems (excluding any ring systems which are part of or formed by optional substituents) are aromatic. Examples of heteroaryl groups / moieties include the following: O On O On On nO O G G G G G G G 30 wherein G = O, S or NH. 16 05 25 In the context of the present specification, unless otherwise stated, a Cx-Cy group is defined as a group containing from n to n carbon atoms. For example, a Ci-C4 alkyl group is defined as an alkyl group containing from 1 to 4 carbon atoms. Optional substituents and moieties are not taken into account when calculating the total number 5 of carbon atoms in the parent group substituted with the optional substituents and / or containing the optional moieties. For the avoidance of doubt, replacement heteroatoms, e.g. N, O or S, are to be counted as carbon atoms when calculating the number of carbon atoms in a Cx-Cy group. For example, a morpholinyl group is to be considered a Ce heterocyclic group, not a C4 heterocyclic group. 10 For the purposes of the present specification, where it is stated that a first atom or group is “directly attached” to a second atom or group it is to be understood that the first atom or group is covalently bonded to the second atom or group with no intervening atom(s) or groups being present. So, for example, for the group 15 (C=O)N(CH3)2, the carbon atom of each methyl group is directly attached to the nitrogen atom and the carbon atom of the carbonyl group is directly attached to the nitrogen atom, but the carbon atom of the carbonyl group is not directly attached to the carbon atom of either methyl group. 0' 0' HO H3C 30 R1 and R2 are the same or together represent =0. The compound has the following formula: 16 05 25 The compounds described herein have been found to reduce the proliferation of C. difficile when in the presence of other bacteria, such as E. coli. Without wishing to be bound by theory, it is believed that compounds described herein inhibit the production io of 4-methylphenol by C. difficile, which is toxic to bacteria, such as E. coli. By inhibiting the production of 4-methylphenol, E. coli can effectively compete against C. difficile and gut microbial diversity may be preserved. A second aspect of the invention provides a pharmaceutically acceptable salt or solvate 15 of any compound of the first aspect of the invention. The compounds of the present invention can be used both in their free base form and their acid addition salt form. For the purposes of this invention, a “salt” of a compound of the present invention includes an acid addition salt. Acid addition salts are 20 preferably pharmaceutically acceptable, non-toxic addition salts with suitable acids, including but not limited to inorganic acids such as hydrohalogenic acids (for example, hydrofluoric, hydrochloric, hydrobromic or hydroiodic acid) or other inorganic acids (for example, nitric, perchloric, sulfuric or phosphoric acid); or organic acids such as organic carboxylic acids (for example, propionic, butyric, glycolic, lactic, mandelic, 25 citric, acetic, benzoic, salicylic, succinic, malic or hydroxysuccinic, tartaric, fumaric, 16 05 25 maleic, hydroxymaleic, mucic or galactaric, gluconic, pantothenic or pamoic acid), organic sulfonic acids (for example, methanesulfonic, trifluoromethanesulfonic, ethanesulfonic, 2-hydroxyethanesulfonic, benzenesulfonic, toluene-p-sulfonic, naphthalene-2-sulfonic or camphorsulfonic acid) or amino acids (for example, 5 ornithinic, glutamic or aspartic acid). The acid addition salt may be a mono-, di-, tri- or multi-acid addition salt. A preferred salt is a hydrohalogenic, sulfuric, phosphoric or organic acid addition salt. A preferred salt is a hydrochloric acid addition salt. The compounds of the present invention can also be used both, in their free acid form io and their salt form. For the purposes of this invention, a “salt” of a compound of the present invention includes one formed between a protic acid functionality (such as a carboxylic acid group) of a compound of the present invention and a suitable cation. Suitable cations include, but are not limited to lithium, sodium, potassium, magnesium, calcium and ammonium. The salt may be a mono-, di-, tri- or multi-salt. 15 Preferably the salt is a mono- or di-lithium, sodium, potassium, magnesium, calcium or ammonium salt. More preferably the salt is a mono- or di-sodium salt or a mono- or dipotassium salt. Preferably any salt is a pharmaceutically acceptable non-toxic salt. However, in 20 addition to pharmaceutically acceptable salts, other salts are included in the present invention, since they have potential to serve as intermediates in the purification or preparation of other, for example, pharmaceutically acceptable salts, or are useful for identification, characterisation or purification of the free acid or base. 25 The compounds and / or salts of the present invention may be anhydrous or in the form of a hydrate (e.g. a hemihydrate, monohydrate, dihydrate or trihydrate) or other solvate. Such solvates may be formed with common organic solvents, including but not limited to, alcoholic solvents e.g. methanol, ethanol or isopropanol. 30 The compounds, salts and solvates of the present invention may contain any stable isotope including, but not limited to 12C, 13C, JH, 2H (D), 160,170,180 and any radioisotope including, but not limited to "C, 14C, 3H (T), 130. The compounds, salts and solvates of the present invention may be in any polymorphic 35 or amorphous form. 16 05 25 The compounds of the first aspect are for use in the treatment of a bacterial infection. A fourth aspect of the invention provides a compound of the first aspect for use in the inhibition of 4-methylphenol production by bacteria. 5 The compounds of the first aspect are for use in the treatment of Clostridioid.es difficile infection. A sixth aspect of the invention provides a pharmaceutical composition comprising a 10 compound of the first aspect, or a pharmaceutically acceptable salt or solvate of the second aspect of the invention, and a pharmaceutically acceptable excipient. Conventional procedures for the selection and preparation of suitable pharmaceutical formulations are described in, for example, “Aulton’s Pharmaceutics - The Design and 15 Manufacture of Medicines”, M. E. Aulton and K. M. G. Taylor, Churchill Livingstone Elsevier, 4th Ed., 2013. Pharmaceutically acceptable excipients including adjuvants, diluents or carriers that maybe used in the pharmaceutical compositions of the invention are those 20 conventionally employed in the field of pharmaceutical formulation, and include, but are not limited to, sugars, sugar alcohols, starches, ion exchangers, alumina, aluminium stearate, lecithin, serum proteins such as human serum albumin, buffer substances such as phosphates, glycerine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes such as 25 protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat. 30 In the pharmaceutical composition may additionally comprise one or more further active agents. The pharmaceutical composition may be provided as a part of a kit of parts, wherein 35 the kit of parts comprises the pharmaceutical composition of the fourth aspect of the invention and one or more further pharmaceutical compositions, wherein the one or 16 05 25 more further pharmaceutical compositions each comprise a pharmaceutically acceptable excipient and one or more further active agents. The term “treatment” as used herein refers equally to curative therapy, and 5 ameliorating or palliative therapy. The term includes obtaining beneficial or desired physiological results, which may or may not be established clinically. Beneficial or desired clinical results include, but are not limited to, the alleviation of symptoms, the prevention of symptoms, the diminishment of extent of disease, the stabilisation (i.e., not worsening) of a condition, the delay or slowing of progression / worsening of a io condition / symptoms, the amelioration or palliation of the condition / symptoms, and remission (whether partial or total), whether detectable or undetectable. The term “palliation”, and variations thereof, as used herein, means that the extent and / or undesirable manifestations of a physiological condition or symptom are lessened and / or time course of the progression is slowed or lengthened, as compared to not 15 administering a compound, salt, solvate, prodrug or pharmaceutical composition of the present invention. The term “prevention” as used herein in relation to a disease, disorder or condition, relates to prophylactic or preventative therapy, as well as therapy to reduce the risk of developing the disease, disorder or condition. The term “prevention” includes both the avoidance of occurrence of the disease, disorder or 20 condition, and the delay in onset of the disease, disorder or condition. Experimental The suitability of the following compounds, EXi to EX8, for reducing the proliferation of the bacteria C. difficile, and thus the treatment of C. difficile infections, was 25 investigated. 16 05 25 EX2 Toxicity to C. difficile Compounds EXi to EX5 were separately tested to determine their toxicity to C. difficile 5 and E. coli. Each compound was dissolved to 6.6 mM in minimal media (MM) with 1% DMSO based on the MM described by Cartman et al. in Appln. Environ. Microbiol,, 2010,76, 1103 and the concentration was matched in a 1:1 ratio of the compound to the substrate 10 p-HPA. To determine growth rates of C. difficile and E. coli in the presence of compounds EXi to EX5, three colonies of C. difficile or three colonies of E. coli were used to inoculate a primary culture of 10 mL MM in 50 cm3 vented tissue culture flasks (Thermo 15 Scientific). Following overnight incubation on a shaking platform at 50 rpm, the primary culture was back diluted in MM to OD59O nm 0.5 using a Fisherbrand™ Digital Colorimeter model 4.5 (Fisher Scientific). 1 ml of the OD59O nm 0.5 culture was inoculated into 10 ml of MM in 50 cm3 vented tissue culture flasks with and without 4-hydroxyphenylacetic acid and the relevant compound (EXi to EX5). 20 OD59o nm readings were determined every hour for eight hours using a Fisherbrand™ Digital Colorimeter model 4.5 (Fisher Scientific). 16 05 25 Growth curves denoting growth over an 8-hour time period for C. difficile and E. coli in MM with the presence of 4-hydroxyphenylacetic acid and inhibitors EXi to EX5, each at 6.6 mM. Absorbance was measured at OD59Onm. *£<0.05, **p<o.oi, samples represent a minimum of three biological replicates graphed in GraphPad prism (vq). 5 Raw absorbance was normalised using a blank MM control. The growth curves are shown in Figures la to 5b. The data show that EXi inhibited the growth of E. coli and had no effect on the growth 10 of C. difficile. EX2 was identified as being toxic to both E. coli and C. difficile. EX3 inhibited the growth of C. difficile when both EX3 and p-HPA were present and inhibited the growth of E. coli. EX4 had no effect on the growth of C. difficile. EX5 had no effect on the growth of either C. difficile or E. coli. 15 Co-Culture fmutated C. difficile') In order to evaluate whether reducing 4-methylphenol production by C. difficile would result in a reduction in the growth of C. difficile, co-cultures comprising C. difficile, a 4-methylphenol deficient mutant of C. difficile and E. coli were developed and used as a surrogate readout of 4-methylphenol production. 20 Under anaerobic conditions, individual overnight cultures of C. difficile and an intestinal competitor, E. coli, were grown in 10 ml of BHI + 0.05% (w / v) L-cysteine supplemented with 100 ng / ml or 250 ng / ml anhydrotetracycline (to induce expression of hpdBCA expressed in trans in the complement strain). Individual monocultures 25 were normalised to a starting optical density (ODeoo) of 0.5, and were inoculated 1 / 10 into BHIS broth supplemented with 0.05% L-cysteine (w / v) and either 0.1, 0.2 or 0.3%p-HPA (w / v) (6.6 mM, 13.1 mM and 19.7 mM, respectively), these monocultures were grown until C. difficile reached an OD 0.5-0.6 (~7 hours). The competitor was back diluted to an ODeoo 0.5 and inoculated 1:10 into the C. difficile culture, to create a 30 competitive co-culture. These co-cultures were grown anaerobically, shaking (50 rpm) for 24 hours and were plated onto both BHIS non-selective plates and BHIS with D-cycloserine (250 mg / L) and cefoxitin (8 mg / L) (CC) plates. CFU counts of both C. difficile and the competitor were determined by serial dilutions plated in triplicate and an average of the three technical replicates was used to determine total 35 CFU. Each experiment was performed in triplicate and linear regression analysis was 16 05 25 performed in Statai5 on the log™ of the CFU, statistically significant differences were observed p<o.05. The results are shown in Figure 6. The data show that E. coli is sensitive to 4- 5 methylphenol and so reduced 4-methylphenol production by C. difficile corresponds to an increase in the growth of E. coli. If C. difficile is mutated to prevent 4-methylphenol production, the viability of C. difficile relative to E. coli is decreased. Co-Cultures fCompounds EXi to EX7) 10 Compounds EXi to EX7 were tested to determine their ability to reduce the growth of C. difficile relative to E. coli. C. difficile 63dAerm and E. coli were grown overnight in MM. C. difficile was back diluted to an OD59O nm of 0.2.18 pl of the back dilution was added to 1.8 ml of the test 15 conditions: MM, MM + 6.6 mMp-HPA, MM + 6.6 mM inhibitor, and MM + 6.6 mMp-HPA + 6.6 mM inhibitor, in a 24 well plate. Each inhibitor was dissolved in DMSO such that the final concentration in the 24 well plate was 1% v / v. C. difficile was grown for 8 hours before E. coli underwent back dilution to an OD59O nm of 0.2 and 18 pl was inoculated into the C. difficile wells to give the co-culture. The co-culture was grown for 20 14 hours. The proportion of the co-cultures made up by C. difficile and E. coli were determined by colony-forming units per millilitre (CFU / ml) assays with co-cultures plated on to BHIS plates in duplicate with selective media for each species. C. difficile was selected for using cycloserine (250 mg / 1) and cefoxitin (8 mg / 1) whilst E. coli was selected for using vancomycin (4 mg / 1). CFU serial ten-fold dilutions of the co-culture 25 were made in PBS with dilutions to io-6. CFUs were counted on the following day. CFU data is presented using CFU percentages which were calculated by dividing the number of CFUs for each species by the total number of CFUs for both E. coli and C. difficile and multiplying by too. Each experiment was performed in a minimum of three independent replicates. Data was analysed by linear regression carried using Stataiy. 30 The results are shown in Figures 7 to 13. The data show that each of compound EXi to EX5 is able to inhibit the growth of C. difficile. Based on this, these compounds would be expected to be effective in 35 maintaining gut microbial diversity and preventing relapse of C. difficile infection. Compounds EX6 and EX7 were also able to inhibit the growth of C. difficile with little or no negative impact on the growth of E. coli. Moreover, all of the compounds had minimal effect on the proportions of the co-cultures when p-HPA was not present. This supports that the effect was as a result of 4-methylphenol production inhibition because 4-methylphenol is only produced in the presence of p-HPA or from tyrosine 5 fermentation to p-HPA. HPLC (Monocultures) C. difficile monocultures were established in the presence of the following compounds EXi, EX3, EX5 and EX8. 16 05 25 Samples of 63oAerm were collected following growth in minimal media with 1% DMSO v / v for 8 hours with 1.5 mg / ml p-HPA with matched (6.6 mM) concentrations of compounds EXi, EX3, EX5 and EX8. At the time of sample collection, the OD59O nm of the culture was measured. Samples were immediately filter sterilised using 0.22 pm 15 filters, then frozen at -80 °C prior to HPLC analysis. The filter-sterilized samples were transferred to HPLC vials and analysed immediately by injecting onto the HPLC column. Separations were achieved by utilizing an Acclaim 120 (Thermofisher), C18,5 pm (4.6 x 150 mm), with the mobile phase consisting of 20 ammonium formate (10 mM, pH 2.7) and menthol (v / v; 50:50) at a flow rate of 1400 pl / min. p-HPA andp-cresol were detected by the diode array detector (PDA; DAD 3000) set at 280 nm. Peak identity was confirmed by measuring the retention time of commercially available p-HPA andp-cresol, and determination of absorbance spectra was performed using the DAD. A calibration curve of each compound was generated by 25 Chromeleon (Dionex software) using known amounts of the reference standards (0-5 mg / ml) dissolved in media and injected onto the column, and the amount ofp-HPA andp-cresol in the samples was determined. Samples from three independent biological replicates were analysed compared to media controls and standard curves. The limit of detection forp-HPA andp-cresol are 0.001 and 0.0005 mg / ml, 30 respectively. The p-cresol concentration was normalised to growth by using the OD59O nm measured at the time of sample collection. 16 05 25 The results are shown in Figure 14. The HPLC data demonstrate that the production of 4-methylphenol by C. difficile is 5 reduced in the presence of compounds EXi, EX3, EX5 and EX8. The reduced 4-methylphenol production is indicative that these compounds are capable of inhibiting the production of 4-methylphenol by C. difficile. By inhibiting the production of 4-methylphenol, other gut bacteria, such as E. coli, would be expected to be able to effectively compete against C. difficile. 10 HPLC (Co-cultures) A C. difficile 63oAerm and E. coli co-culture was established (as per the methods previously described) in the presence of compound EX3. 15 Following co-culture, CFU assays were undertaken to determine relative survival of C. diff and E.coli. The remaining sample was filter sterilised (0.22 pm filters), then frozen at -80 °C prior to the HPLC analysis. HPLC analysis was carried out as described above (HPLC monocultures) with the exception of there being no normalisation of the 4-methylphenol concentration to OD59O nm. 20 HO EX3 The results are shown in Figure 15. 25 The HPLC co-culture data demonstrate that the production of 4-methylphenol by C. difficile is reduced in the presence of compound EX3 and E. coli.

Claims

1. A compound selected from the group consisting of:16 05 25or a pharmaceutically acceptable salt or solvate thereof, for use as a medicament, wherein the medicament is for use in the treatment of Clostridioid.es difficile (C. difficile) infection.

2. A compound as claimed in claim 1, wherein the medicament is for the inhibition of 4-methylphenol production by bacteria.

3. A pharmaceutically acceptable salt or solvate of a compound as claimed in either claim 1 or claim 2.

4. A pharmaceutical composition comprising a compound as claimed in either claim 1 or claim 2, or a pharmaceutically acceptable salt or solvate as claimed in claim 3, and a pharmaceutically acceptable excipient.

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