Patent Application: US-201214369904-A

Abstract:
the present invention relates to a novel 2 - aminothiazole derivative , n - thiazole - 2 - il )- 3 - propane amide and to a use thereof as an anti - cancer drug . it is verified that the compound of the present invention has highly superior cancer cell - specific cytotoxic activity and in - vivo anti - cancer activity for inhibiting the growth of tumors in an animal model for cancer disease , and therefore can be developed as an anti - cancer candidate substance .

Description:
hereinafter , the present invention will be described in detail with reference to examples . it is clear to those skilled in the art that examples are intended to specifically describe the present invention , and do not limit the scope of the present invention according to the gist of the present invention . 3 -( 2 - furyl ) propionic acid ( 3 . 85 mmoles ), n , n ′- dicyclohexyl carbodiimide ( 3 . 85 mmoles ), and hydroxybenzotriazole ( 0 . 385 mmoles ) were dissolved in dry methylene chloride ( mc ) ( 40 ml ), followed by stirring for 1 . 5 hours at room temperature under nitrogen . 2 - amino - 5 - bromothiazole monohydrobromide ( 3 . 85 mmoles ) was added thereto , and then the reaction mixture was stirred for two days at room temperature . reaction completion was checked by thin layer chromatography ( tlc ). after reaction completion , the reaction mixture was quenched with water . the reaction mixture was separated using mc and distilled water . the separated organic layer was dehydrated using anhydrous mgso 4 . the reaction mixture was concentrated under reduced pressure to remove a solvent , dissolved in a small amount of mc , and then separated by flash column chromatography . finally , 0 . 772 g ( 66 . 67 %) of n -( 5 - bromo - 2 - thiazolyl )- 2 - furanpropanamide was produced as light yellow crystals . 1 h nmr ( 300 mhz , cdcl 3 ) δ10 . 53 ( s , 1h ), 7 . 34 - 7 . 32 ( d , 2h ), 6 . 28 ( t , 1h ), 6 . 08 ( d , 1h ), 3 . 11 ( m , 2h ), 2 . 82 ( m , 2h ). n -( 5 - bromo - 2 - thiazolyl )- 2 - propanamide ( 600 mg , 1 . 99 mmoles ), 4 - fluorophenyl boric acid ( 0 . 42 g , 2 . 99 mmoles ), and sodium carbonate anhydride ( 1 . 05 g , 9 . 95 mmoles ) were dissolved in dimethyl fluoride ( dmf , 18 ml ) and h 2 o ( 0 . 6 ml ), followed by stirring for 1 . 5 hours at room temperature . after stirring , the reaction mixture was maintained for 5 minutes under nitrogen . tetrakis ( triphenylphosphine )- palladium ( 0 . 68 g , 0 . 588 mmoles ) was added thereto , and then maintained at 80 ° c . in an oil bath , followed by stirring overnight . reaction completion was checked by tlc . after reaction completion , the reaction mixture was quenched with distilled water . the reaction mixture was separated using mc and distilled water . the separated organic layer was dehydrated using anhydrous mgso 4 . the reaction mixture was concentrated under reduced pressure to remove a solvent , dissolved in a small amount of mc , and then separated by flash column chromatography . the purified product was recrystallized by hexane and ethylacetate ( hex - ea ) to obtain 16 . 8 mg ( 2 . 7 %) of a white crystalline compound . the synthesized compound , n -( 5 -( 4 - fluorophenyl ) thiazol - 2 - yl )- 3 -( furan - 2 - yl ) propanamide was named hf - 154 , and used for the measurement of the anti - cancer activity in the following examples . 1 h nmr ( 300 mhz , cdcl 3 ) δ10 . 68 ( s , 1h ), 7 . 52 ( m , 2h ), 7 . 31 ( d , 1h ), 7 . 09 ( t , 2h ), 6 . 29 ( t , 1h ), 6 . 10 ( d , 1h ), 3 . 14 ( t , 2h ), 2 . 87 ( t , 2h ). the prostate cancer cell line , pc3 was obtained from korean cell line bank . after adding rosewell park memorial institute ( rpmi ) 1640 cell culture medium ( lm001 - 01 , welgen ) containing 10 % fetal bovine serum ( fbs ), 100 units / ml of penicillin and 100 μg / ml of streptomycin to a 100 φ dish , cells were inoculated on the dish and cultured in a co 2 incubator ( vs - 9000c , vision ) in 5 % co 2 / 95 % humidified air atmosphere at 37 ° c . when the dish was filled with cells by 80 %, the cells were observed with a microscope and subcultured using phosphate buffered saline ( pbs ) and a 0 . 05 % trypsin - ethylene diamine tetraacetic acid ( edta ) solution . ( 2 ) measurement of cytotoxicity activity against cancer cells through mtt analysis toxicity to cells was analyzed using 3 -[ 4 , 5 - dimethylthiazol - 2 - yl ]- 2 , 5 - diphenyl - tetra zolium bromide ( mtt ) analysis . as cancer cells to be measured , 8 . 0 × 10 3 cells / ml was aliquoted on a 96 - well culture dish and then the cells were cultured for 24 hours . the next day , the synthesized compound was dissolved in a medium according to each concentration , and diluted in a medium according to each concentration . the medium in which the compound was dissolved according to each concentration was added to a culture dish at 1 ml , dimethyl sulfoxide ( dmso ) was added to a control well so as to have the same volume as the added compound . after the addition , the cells were cultured in an incubator in 5 % co 2 at 37 ° c ., and then survival capability of cells was measured at 24 hours and 48 hours after the culture . after the medium was removed , the cells were washed twice using pbs , then 200 μl of a solution containing 1 mg / ml of mtt ( sigma - aldrich chemical co ., usa ) was added thereto , and then the cells were further cultured for 3 hours in a co 2 incubator at 37 ° c . the 96 - well culture dish was centrifuged for 10 minutes at 2000 rpm to remove the mtt solution . dmso was added to each well at 200 μl and shaken for 15 minutes to dissolve the produced purple crystals . the solution was added to the 96 - well culture dish , and then absorbance was measured at 540 nm . relative absorbance was calculated using absorbance according to concentration of drugs divided by absorbance of a control group ( drug - untreated group ). as an experiment result by mtt analysis , it could be found that the compound according to the present invention , hf - 154 has an ic 50 value of about 0 . 1 nm in a prostate cancer cell line , pc3 , and a very superior anti - cancer activity for prostate cancer ( see fig1 and table 1 ). further , a cytotoxicity effect of the compound , hf - 154 , was measured against each of lung cancer cells ( a549 ), skin cancer cells ( sk - mel2 ), colorectal cancer cells ( hct15 ), ovarian cancer cells ( sk - ov - 3 ) and breast cancer cells ( mcf7 ), and a result is shown in table 1 . in addition to the prostate cancer cells , it was found that ovarian cancer cells ( sk - ov - 3 ) and breast cancer cells ( mcr7 ) have ic 50 values of about 0 . 5 nm and 0 . 3 nm , respectively , lung cancer cells ( a549 ), skin cancer cells ( sk - mel2 ), colorectal cancer cells ( hct15 ) each have an ic 50 value of about 1 to 5 nm , and therefore the compound has superior anti - cancer efficacy to other cancer cells as well as prostate cancer cells . the anti - cancer activity of the compound according to the present invention , hf - 154 was measured in in - vivo using mice in which a prostate cancer cell line , pc3 , was transplanted . an experimental group included a group of two mice to which an excipient ( vehicle ) was administered and a group of four mice to which the compound according to the present invention , hf - 154 , was administered . the compound according to the present invention , hf - 154 , was administered to each mouse in an amount of 2 mg / kg / day ( 100 μl i . v .). first , after administration for four consecutive days , administration was stopped , and tumor growth was compared at 7 days after the stoppage . as shown in fig2 and 3 , it could be found that the size of the tumor of cancer cells transplanted to the mice to which the compound according to the present invention , hf - 154 , was administered was significantly reduced , compared to the excipient administration group . the present invention relates to a novel 2 - aminothiazole derivative , n -( 5 -( 4 - fluorophenyl ) thiazol - 2 - yl )- 3 -( furan - 2 - yl ) propanamide , and use thereof as an anti - cancer drug . the compound according to the present invention is found to have a very superior specific cytotoxicity activity against cancer cells and an in - vivo anti - cancer activity which inhibits growth of a tumor in an animal model in which cancer cells are transplanted , and hence may be developed as a candidate anti - cancer compound . it is clear to those skilled in the art that the specific portion of the present invention was described in detail above , in which the specific technique is only one embodiment and does not limit the scope of the present . therefore , the substantial scope of the present invention will be defined by the appended claims and their equivalents . 2 . foley p l and shuler m l : biotechnol bioeng 105 : 26 - 36 , 2010 . 3 . kim , k . s . et al . j . med . chem . 2002 , 45 , 3905 . 4 . misra , r . n . et al . j . med . chem . 2004 , 47 , 1719 . 5 . borzilleri , r . m . et al . j . med . chem . 2006 , 49 , 3766 . 6 . kim k s , kimball s d , misra r n , et al . : j med chem 45 : 3905 - 3927 , 2002 . 7 . misra r n , xiao h y , kim k s , et al . : j med chem 47 : 1719 - 1728 , 2004 . 8 . jung f h , pasquet g , lambert - van der brempt c , et al . : j med chem 49 : 955 - 970 , 2006 . 9 . borzilleri r m , bhide r s , banish j c , et al . : j med chem 49 : 1 3766 - 3769 , 2006 . 10 . kuramoto m , sakata y , terai k , et al . : org biomol chem 6 : 2772 - 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