QUATERNARY AMINE COMPOUNDS WITH ISOPROPYLMETHYLPHENOL ESTER MOIETIES AS ANTIVIRALS, ANTIBACTERIALS AND ANTIMYCOTICS

Abstract

Disclosed are compounds having the following formula (I):Formula (I) wherein R is an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −2, or R is a quaternary amine having the following formula (Ia):Formula (Ia) wherein R.sub.a and R.sub.b are each an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −3; R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5 and R.sub.6 are each independently selected from C.sub.1-10 alkyl and H; E has the following Formula (Ib), wherein the ester oxygen is bonded to the aromatic ring of each E at the same position, the position being position 2 or 3; and wherein R.sub.E is H or a halide.

##STR00001##

Claims

1. A compound having the following formula: ##STR00053## wherein R is an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −2, or R is a quaternary amine having the following formula: ##STR00054## wherein R.sub.a and R.sub.b are each an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −3; R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5 and R.sub.6 are each independently selected from C.sub.1-10 alkyl and H; E has the following formula ##STR00055## wherein the ester oxygen is bonded to the aromatic ring of each E at the same position, the position being position 2 or 3; and wherein R.sub.E is H or a halide.

2. A compound according to claim 1, having the following formula: ##STR00056##

3. A compound according to claim 1, wherein R is a saturated linear alkylene chain having between 8 and 16 carbon atoms, for example 10 carbon atoms.

4. A compound according to claim 1, wherein R.sub.a and R.sub.b are each a saturated linear alkylene chain having between 8 and 16 carbon atoms, for example 10 carbon atoms.

5. A compound according to claim 1, wherein the one or more anions are selected from chloride anions and bromide anions.

6. A compound according to claim 1, wherein R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5 and R.sub.6 are each methyl.

7. A compound according to claim 1, having the following formulae: ##STR00057##

8. A compound having the following formula: ##STR00058## wherein R is an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −2, or R is a quaternary amine having the following formula: ##STR00059## wherein R.sub.a and R.sub.b are each an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −3; R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5 and R.sub.6 are each independently selected from C.sub.1-10 alkyl and H; E has the following formula ##STR00060## wherein the ester oxygen is bonded to the aromatic ring of each E at the same position, the position being 2 or 3; wherein R.sub.E is H or a halide and wherein X is 2, 3, 4 or 6.

9. A compound according to claim 8 having the following formula: ##STR00061##

10. A compound according to claim 8 or 9, wherein R is a saturated linear alkylene chain having between 8 and 16 carbon atoms, for example 10 carbon atoms.

11. A compound according to claim 8 or 9, wherein R.sub.a and R.sub.b are each a saturated linear alkylene chain having between 8 and 16 carbon atoms, for example 10 carbon atoms.

12. A compound according to claim 8 or 9, wherein the one or more anions are selected from chloride anions and bromide anions.

13. A compound according to claim 8 or 9 having the following formula: ##STR00062## wherein X is 3 or 6.

14. A pharmaceutical composition comprising the compound of any of claims 1-9.

15. A compound according to any one of claims 1 to 9 for use as a medicament.

16. A compound according to any one of claims 1 to 9 for use in the treatment of fungal infections and/or bacterial infections.

17. A compound according to any one of claims 1 to 9 for use in the treatment of Herpes virus and/or Human Papilloma virus.

18. A process for producing the compound according to claim 1 comprising: i) reacting a compound having the formula (XV) ##STR00063## wherein the alcohol group is bonded to the benzene ring in position 2 or 3, and R.sub.E is H or a halide, with R.sub.7CH.sub.2COCl to form a compound having the formula (XVI): ##STR00064## wherein R.sub.7 is chlorine or bromine, E is ##STR00065## wherein the ester oxygen is bonded to the aromatic ring of E at position 2 or 3, and wherein R.sub.E is H or a halide; and ii) reacting 1 molar equivalents of the compound having the formula (XVII) ##STR00066## with 2 molar equivalent of the compound having formula (XVI) to form a compound having the formula ##STR00067## wherein R is an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −2; or R is a quaternary amine having the following formula: ##STR00068## wherein R.sub.a and R.sub.b are each an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −3; E has the following formula ##STR00069## wherein the ester oxygen is bonded to the aromatic ring of each E at the same position, the position being position 2 or 3 and wherein R.sub.E is H or a halide; R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5 and R.sub.6 are each independently selected from C.sub.1-10 alkyl and H.

19. The process according to claim 18, wherein the reactions of step II is carried out in acetonitrile as solvent.

20. A process for the production of a compound according to claim 8 comprising reacting the compound having formula (I) ##STR00070## with bromine to form a compound having formula (VIII) ##STR00071## wherein R is an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −2, or R is a quaternary amine having the following formula: ##STR00072## wherein R.sub.a and R.sub.b are each an alkylene chain having between 8 and 20 carbon atoms, and A is one or more anions having a total charge of −3; R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5 and R.sub.6 are each independently selected from C.sub.1-10 alkyl and H; E has the following formula ##STR00073## wherein the ester oxygen is bonded to the aromatic ring of each E at the same position, the position being 2 or 3; wherein R.sub.E is H or a halide; and wherein X is 2, 3, 4 or 6.

Description

Example 1

Synthesis of the compound of formula (IV)

[0110] ##STR00038##

[0111] In a first step, 1,10-dibromodecane is reacted with 2 molar equivalents of dimethylamine to form 1,10-Bis(dimethylamino)decane. The reaction takes place at 4-5° C. in a suitable solvent such as benzene, diethyl ether or dimethylamine and is followed by a step of acid extraction followed by alkaline treatment and extraction with diethyl ether. The extracted fractions are dried over magnesium sulphate and then purified by vacuum distillation.

[0112] In a second step, Carvacrol (2-Methyl-5-(1-methylethyl)-phenol) is reacted with chloroacetyl chloride. The reaction is carried out at −10° C. for 1 hour and then stirred at room temperature for 12 hours. The reaction mixture is then washed with acid, followed by treatment with sodium bicarbonate and then water. The organic layer is dried over sodium sulphate, filtered and the solvent is removed under vacuum.

[0113] In a third step, 2 molar equivalents of the compound formed in the second step is reacted with one molar equivalent of 1,10-Bis(dimethylamino)decane to form the compound of formula (IV). The reaction in the third step is carried out using an appropriate solvent such as acetonitrile and the reaction mixture is stirred for 24 hours.

[0114] It will be appreciated that further purification and separation steps may also be included in the process, for example between each of the above steps and also after the process is complete to purify the final compound having formula (IV).

[0115] Separation steps may include steps of performing column chromatography, low pressure liquid chromatography, high performance liquid chromatography and the like. Purification steps may include standard purification processes known in the art, for example, filtration, evaporation, liquid-liquid extraction, crystallisation, adsorption, recrystallization, chromatography, distillation and the like.

[0116] The compound of formula (IV) has the following systematic name, formula and molecular weight:

[0117] Systematic name:N1,N10-bis(2-(5-isopropyl-2-methylphenoxy)−2-oxoethyl)-N1,N1,N10,N10-tetramethyldecane-1,10-diaminium dichloride

[0118] Formula:C.sub.38H.sub.62N.sub.2O.sub.4.sup.+2 2Cl.sup.−

[0119] Molecular weight: 681.81

##STR00039##

Example 2

Synthesis of the Compound of Formula (V)

[0120] ##STR00040##

[0121] In a first step, 1,10-dibromodecane is reacted with 2 molar equivalents of dimethylamine to form 1,10-Bis(dimethylamino)decane. The reaction takes place at 4-5° C. in benzene in a suitable solvent such as benzene, diethyl ether or dimethylamine and is followed by a step of acid extraction followed by alkaline treatment and extraction with diethyl ether. The extracted fractions are dried over magnesium sulphate and then purified by vacuum distillation.

[0122] In a second step, thymol (2-isopropyl-5-methylphenol) is reacted with chloroacetyl chloride. The reaction is carried out at −10° C. for 1 hour and then stirred at room temperature for 12 hours. The reaction mixture is then washed with acid, followed by treatment with sodium bicarbonate and then water. The organic layer is dried over sodium sulphate, filtered and the solvent is removed under vacuum.

[0123] In a third step, 2 molar equivalents of the compound formed in the second step is reacted with one molar equivalent of 1,10-Bis(dimethylamino)decane to form the compound of formula (V). The reaction in the third step is carried out using an appropriate solvent such as acetonitrile and the reaction mixture is stirred for 24 hours.

[0124] It will be appreciated that further purification and separation steps may also be included in the process, for example between each of the above steps and also after the process is complete to purify the final compound having formula (V).

[0125] Separation steps may include steps of performing column chromatography, low pressure liquid chromatography, high performance liquid chromatography and the like. Purification steps may include standard purification processes known in the art, for example, filtration, evaporation, liquid-liquid extraction, crystallisation, adsorption, recrystallization, chromatography, distillation and the like.

[0126] Systematic Name:N1,N10-bis(2-(2-isopropyl-5-methylphenoxy)−2-oxoethyl)-N1,N1,N10,N10-tetramethyldecane-1,10-diaminium dichloride

[0127] Formula:C.sub.38H.sub.62N.sub.2O.sub.4.sup.+22Cl.sup.−

[0128] Molecular Weight: 681.81

##STR00041##

Example 3

Synthesis of the Compound of Formula (VI)

[0129] ##STR00042##

[0130] In a first step, 2 molar equivalents of 1,10-Dibromodecane are reacted with 3 molar equivalents of dimethylamine. The reaction takes place at 4-5° C. in a suitable solvent such as benzene or diethyl ether and is followed by a step of acid extraction followed by alkaline treatment and extraction with diethyl ether. The extracted fractions are dried over magnesium sulphate and then purified by vacuum distillation.

[0131] In a second step, carvacrol (2-Methyl-5-(1-methylethyl)-phenol) is reacted with chloroacetyl chloride. The reaction is carried out at −10° C. for 1 hour and then stirred at room temperature for 12 hours. The reaction mixture is then washed with acid, followed by treatment with sodium bicarbonate and then water. The organic layer is dried over sodium sulphate, filtered and the solvent is removed under vacuum.

[0132] In a third step, one molar equivalent of the compound formed in step 1 is reacted with the 2 molar equivalents of the compound formed in step 2 to form the compound having formula (VI). The reaction in the third step is carried out using an appropriate solvent such as acetonitrile and the reaction mixture is stirred for 24 hours.

[0133] It will be appreciated that further purification and separation steps may also be included in the process, for example between each of the above steps and also after the process is complete to purify the final compound having formula (VI).

[0134] Separation steps may include steps of performing column chromatography, low pressure liquid chromatography, high performance liquid chromatography and the like. Purification steps may include standard purification processes known in the art, for example, filtration, evaporation, liquid-liquid extraction, crystallisation, adsorption, recrystallization, chromatography, distillation and the like.

[0135] Systematic Name:N1-(2-(5-isopropyl-2-methylphenoxy)−2-oxoethyl)-N10-(10-((2-(5-isopropyl-2-methylphenoxy)-2-oxoethyl)dimethylammonio)decyl)-N1,N1,N10,N10-tetramethyldecane-1,10-diaminium bromide dichloride

[0136] Formula:C.sub.50H.sub.88N.sub.3O.sub.4.sup.+3Br2Cl.sup.−

[0137] Molecular Weight: 946.06

##STR00043##

Example 4

Synthesis of the Compound of Formula (VII)

[0138] ##STR00044##

[0139] In a first step, 2 molar equivalents of 1,10-Dibromodecane are reacted with 3 molar equivalents of dimethylamine The reaction takes place at 4-5° C. in a suitable solvent such as benzene or diethyl ether and is followed by a step of acid extraction followed by alkaline treatment and extraction with diethyl ether. The extracted fractions are dried over magnesium sulphate and then purified by vacuum distillation.

[0140] In a second step, thymol (2-isopropyl-5-methylphenol) is reacted with chloroacetyl chloride. The reaction is carried out at −10° C. for 1 hour and then stirred at room temperature for 12 hours. The reaction mixture is then washed with acid, followed by treatment with sodium bicarbonate and then water. The organic layer is dried over sodium sulphate, filtered and the solvent is removed under vacuum.

[0141] In a third step, one molar equivalent of the compound formed in step 1 is reacted with the 2 molar equivalents of the compound formed in step 2 to form the compound having formula (VII). The reaction in the third step is carried out using an appropriate solvent such as acetonitrile and the reaction mixture is stirred for 24 hours.

[0142] It will be appreciated that further purification and separation steps may also be included in the process, for example between each of the above steps and also after the process is complete to purify the final compound having formula (VII).

[0143] Separation steps may include steps of performing column chromatography, low pressure liquid chromatography, high performance liquid chromatography and the like. Purification steps may include standard purification processes known in the art, for example, filtration, evaporation, liquid-liquid extraction, crystallisation, adsorption, recrystallization, chromatography, distillation and the like.

[0144] Systematic Name:N1-(2-(2-isopropyl-5-methylphenoxy)−2-oxoethyl)-N10-(10-((2-(2-isopropyl-5-methylphenoxy)-2-oxoethyl)dimethylammonio)decyl)-N1,N1,N10,N10-tetramethyldecane-1,10-diaminium bromide dichloride

[0145] Formula:C.sub.50H.sub.88N.sub.3O.sub.4.sup.+3Br2Cl.sup.−

[0146] Molecular Weight: 946.06

##STR00045##

[0147] It will be appreciated that to synthesis the Bromine complexes defined by formula XI to XIV, the compounds of formula IV to VII are treated with bromine to form the complexes of formulas XI to XIV. It will be appreciated that further purification and separation steps may also be included in this process.

Example 5

[0148] Minimum Inhibitory Concentration (MIC) Determination by Broth Micro Dilution

[0149] Equipment [0150] McFarland standard 0.5 [0151] Falcon round-bottom 5 ml tubes [0152] Disposable loops (1 μl and 10 μl) [0153] Pipettes tips (0.5 μl-1000 μl) [0154] Tissue Culture plate, 96 well [0155] Multichannel micropipettes (5-50 μl),(30-300 μl) [0156] Disposable Petri dishes [0157] Disposable reagent reservoirs

[0158] Media [0159] Sterile normal saline [0160] TSB (Tryptic Soy Broth) [0161] TSA (Tryptic Soy Agar) [0162] 0,02% resazurin sterile solution

[0163] Bacterial strains [0164] Staphylococcus aureus ATCC-6538 [0165] Escherichia coli A TCC-8739

[0166] Procedure

[0167] Standardisation of Inoculum

[0168] From a pure o/n culture, material from at least 3-4 colonies was chosen and suspended totally in 2 ml saline in tubes. The suspensions were mixed. The inoculums were standardized according to the scale of 0.5 MacFarland (dilution factor 1:10, sterile saline solution) to obtain the final concentration of 1.5×10.sup.4 cfu/ml. The inoculums were 30 mixed. The suspension was used for inoculation within 15 minutes.

[0169] Inoculation and Incubation

[0170] Each sample was dissolved in an appropriate solvent (water/96% ethanol) to a concentration of 5000 μg/ml. Using the method of serial dilutions, the concentration of the initial solution was adjusted to 625 μg/ml for gram negative bacteria and 78 μg/ml for gram positive bacteria. For fungi, the working concentration was 1250 μg/ml.

[0171] The tests were performed in 96 well plates, in which each well received 100 μl of sterile TSB.

[0172] 100 μl of the initial concentration of the sample was added to the first well. Serial dilutions were made by transferring 100 μl from each previous well to the subsequent one (dilution factor 1:1). In each well, 20 μl of the inoculum is added. Each well had a final volume of 120 μl.

[0173] The microbial growth, sterility of the medium and the solvent were controlled simultaneously. The plates were incubated at 37±1° C., for 24 h for bacteria and at 34±1° C., for 48 h for yeasts. The inhibition of growth of bacteria was revealed by the addition of resazurin sterile solution (20 μl, 0.02%, w/v) and re-incubation for 3 hours. Minimum inhibitory concentration (MIC), which is defined as the lowest concentration of sample capable of inhibiting the growth of microorganisms, was determined by the permanence of blue coloration in the wells. A change of color from blue to red (due to the reduction of dye) indicated bacterial growth. The wells that showed no apparent growth were selected to evaluate the MIC, which was determined by the absence of microbial growth on plates containing TSB.

[0174] The Results of these Experiments are Set Out Below in the Following Tables.

[0175] Staphylococcus aureus ATCC 6538 4.5×10.sup.3 Cfu/Ml

TABLE-US-00001 TABLE 1 Compound MIC/μg/mL Comparative compound 1 1.95 Formula VII 0.98

[0176] Escherichia coli ATCC-8739 6.5×10.sup.3 Cfu/ml

TABLE-US-00002 TABLE 2 Compound MIC/μg/mL Comparative compound 1 7.8 Formula VII 3.9

TABLE-US-00003 TABLE 3 Compound Structure Comparative Compound 1 [00046] Formula VII [00047]

Example 6

[0177] MIC Determination by Broth Micro Dilution

[0178] Equipment [0179] McFarland standard 0.5 [0180] Falcon round-bottom 5 ml tubes [0181] Disposable loops (1 μl and 10 μl) [0182] Pipettes tips (0.5 μl-1000 μl) [0183] Tissue Culture plate, 96well [0184] Multichannel micropipettes (5-50 μl),(30-300 μl) [0185] Disposable Petri dishes [0186] Disposable reagent reservoirs

[0187] Media [0188] Sterile normal saline [0189] TSB (Tryptic Soy Broth) [0190] TSA (Tryptic Soy Agar) [0191] 0,02% resazurin sterile solution

[0192] Bacterial strains [0193] Staphylococcus aureus ATCC-6538 [0194] Escherichia coli A TCC-8739 [0195] Pseudomonas aeruginosa ATCC-9027 [0196] Candida albicans ATCC-10231 [0197] Candida albicans NCTC-885-653 [0198] Saccharomyces cerevisiae ATCC-9763 [0199] Salmonella enterica serovar Typhimurium ATCC 14028 [0200] Saccharomyces cerevisiae ATCC-2601

[0201] Procedure

[0202] Standardisation of Inoculum

[0203] From a pure o/n culture, material from at least 3-4 colonies was chosen. The material was suspended totally in 2 ml saline in tubes and mixed.

[0204] The inoculums were standardized according to the scale of 0.5 MacFarland (dilution factor 1:10, sterile saline solution) to obtain the final concentration of 1.5×10.sup.4cfu/ml. The inoculums were mixed. The suspension was used for inoculation within 15 minutes.

[0205] Inoculation and Incubation

[0206] Each sample was dissolved in an appropriate solvent (water/96% ethanol) to a concentration of 5000 μg/ml. Using the method of serial dilutions, the concentration of the initial solution was adjusted to 625 μg/ml for gram negative bacteria and 78 μg/ml for gram positive bacteria. For fungi the working concentration was 1250 μg/ml.

[0207] The tests were performed in 96 well plates, in which each well received 100 μl of sterile TSB. 100 μl of the initial concentration was added to the first well. Serial dilutions were made by transferring 100 μl from each previous well to the subsequent one (dilution factor 1:1). In each well 20 μl of the inoculum was added. Each well had a final volume of 120 μl. The microbial growth, sterility of the medium and the solvent were controlled simultaneously. The plates were incubated at 37 t 1° C., for 24 hours for bacteria and at 34±1° C., for 48 hours for yeasts. The inhibition of growth of bacteria was revealed by the addition of resazurin sterile solution (20 μl, 0.02%, w/v) and re-incubation for 3 h.

[0208] MIC, which is defined as the lowest concentration of sample capable of inhibiting the growth of microorganisms, was determined by the permanence of blue coloration in the wells. A change of color from blue to red (due to the reduction of dye) indicated the bacterial growth. The wells that showed no apparent growth were selected to evaluate the MIC, which was determined by the absence of microbial growth on plates containing TSB.

[0209] The Results of these Experiments are Set Out in the Following Tables.

[0210] Escherichia coli ATCC-8739 1.7×10.sup.4 Cfu/ml

TABLE-US-00004 TABLE 4 Compound MIC/μg/mL Comparative compound 2 7.8 Formula V 3.9 Formula IV 3.9 Vancomycin 78

[0211] Salmonella enterica serovar Typhimurium ATCC 14028 1.3×10.sup.4 Cfu/ml

TABLE-US-00005 TABLE 5 Compound MIC/μg/mL Comparative compound 2 7.8 Formula IV 3.9 Vancomycin 78

[0212] Escherichia coli ATCC-8739 6.6×10.sup.3 Cfu/ml

TABLE-US-00006 TABLE 6 Compound MIC/μg/mL Comparative compound 2 7.8 Formula V 3.9

[0213] Pseudomonas aeruginosa ATCC-9027 1.9×10.sup.4 Cfu/ml

TABLE-US-00007 TABLE 7 Compound MIC/μg/mL Comparative compound 2 7.8 Formula V 3.9

[0214] Escherichia coli ATCC-8739 4.1×10.sup.3 Cfu/ml

TABLE-US-00008 TABLE 8 Compound MIC/μg/mL Comparative compound 2 3.9 Formula V 1.95 Formula IV 1.95

[0215] Pseudomonas aeruginosa ATCC-9027 4.7×10.sup.3 Cfu/ml

TABLE-US-00009 TABLE 9 Compound MIC/μg/mL Comparative 1.95 compound 2 Formula IV 0.98

TABLE-US-00010 TABLE 10 MIC/μg/mL Comparative compound 2 Formula V Formula IV Candida 15.6 3.9-7.8 3.9-7.8 albicans ATCC 10231

TABLE-US-00011 TABLE 11 Comparative compound 2 Formula V Candida 15.6 3.9-7.8 albicans NCTC 885-653

TABLE-US-00012 TABLE 12 Compound MIC/μg/mL Comparative 1.95 compound 2 Formula V 0.98

[0216] Escherichia coli ATCC-8739 6.5×10.sup.3 Cfu/ml

TABLE-US-00013 TABLE 13 Compound MIC/μg/mL Comparative 7.8 compound 2 Formula V 3.9 Comparative 7.8 compound 3 Formula XI 3.9

[0217] Salmonella enterica serovar Typhimurium ATCC 14028 (log 4 conditions)

TABLE-US-00014 TABLE 14 Compound MIC/mKg/mL Comparative compound 2 7.8 Formula IV 3.9

[0218] Escherichia coli ATCC-8739 (log 4 conditions)

TABLE-US-00015 TABLE 15 Compound MIC/μg/mL Comparative compound 2 7.8 Formula V 3.9 Formula IV 3.9

[0219] Pseudomonas aeruginosa ATCC-9027 (Log 4 conditions)

TABLE-US-00016 TABLE 16 Compound MIC/μg/mL Comparative compound 3 7.8 Formula XI 3.9

[0220] The means of MIC for different isomers (μg/ml) were tested against various fungal strains that were clinical isolate resistant or sensitive to Fluconazole and Voriconazole (resistance indicated for more than 20 fungal generations, cultivated in the presence of Fluconazole or Voriconazole).

[0221] MIC values for Voriconazole and Fluconazole for all tested strains were 255 μg/ml and 511 μg/ml respectively.

[0222] Candida albicans ATCC-10231 Voriconazole resistant (20 G)

TABLE-US-00017 TABLE 17 Compound MIC/μg/mL Comparative 7.8 compound 2 Formula V 3.9

[0223] Candida albicans ATCC-10231 Fluconazole resistant (20 G)

TABLE-US-00018 Compound MIC/μg/mL Comparative 15.6 compound 2 Formula V 7.8

[0224] Candida albicans NTCT-885-653 Voriconazole resistant (20 G)

TABLE-US-00019 TABLE 18 Compound MIC/μg/mL Comparative 1.95 compound 2 Formula V 0.98

[0225] Saccharomyces cerevisiae ATCC-9763 Fluconazole resistant (20 G)

TABLE-US-00020 TABLE 19 Compound MIC/μg/mL Comparative 3.9 compound 2 Formula IV 0.98

[0226] Saccharomyces cerevisiae ATCC-2601 Fluconazole resistant (20 G)

TABLE-US-00021 TABLE 20 Compound MIC/μg/mL Comparative 1.95 compound 2 Formula IV 0.98 Formula V 0.98

Example 7

[0227] Antiviral Activity

[0228] MRC5 cells were seeded in 96 well plates at a density of 20,000 cells/well and cultured at 37° C. under 5% CO.sub.2 overnight. The next day, a medium containing serially diluted compounds and the virus were added to the wells. The resulting cultures were incubated at 37° C. and under 5% CO.sub.2. Four days post infection, the medium was replenished. Seven days post infection, fluorescence intensity was determined using Acumen Cellista. Antiviral activity of the compounds was calculated based on the inhibition of expression of GFP at each concentration normalized by the cell control. Cytotoxicity of the compounds was assessed under the same conditions, but without virus infection, in parallel. Cell viability was measured with CCK8 following the manufacturer's manual.

[0229] Antiviral activity and cytotoxicity of the compounds was expressed as % Inhibition and % Viability, respectively, and calculated with the formulas below:

Inhibition (%)=100−(Raw data CPD−AverageCC)/(AverageVC−AverageCC)*100

Viability (%)=(Raw data CPD−AverageMC)/(AverageCC−AverageMC)*100

[0230] Raw data CPD indicates the values of the compound-treated wells; Average VC, Average CC and Average MC indicate the average values of the virus control, cell control and medium control wells, respectively.

[0231] EC50 and CC50 values were calculated using the GraphPad Prism software.

TABLE-US-00022 TABLE 21 Compound treatment Detection Virus Cell line duration reagent US3-6-EGFP- MRC5 7/Fluorometry GFP, CCK8 HCMV-AD169

[0232] The results of the experiments are set out below in table 22.

TABLE-US-00023 TABLE 22 EC.sub.50 CC.sub.50 SI Compound (μg/ml) (μg/ml) (CC.sub.50/EC.sub.50) Comparative 41.47 >120.00 >2.89 compound 2 Formula IV 32.75 >120.00 >3.66 Comparative 18.94 23.45 1.24 compound 3 Formula XI 15.76 32.92 2.09

TABLE-US-00024 TABLE 23 Compound Structure Comparative compound 2 [00048] Formula V [00049] Formula IV [00050] Comparative compound 3 [00051] Formula XI [00052]