Patent Application: US-201213423040-A

Abstract:
the invention is directed to a compound of formula , wherein r is selected from the group consisting of hydrogen atom and unsubstituted or substituted phenyl group ; r 1 is selected from the group consisting of hydrogen atom and unsubstituted or substituted phenyl group ; x is selected from the group consisting of hydrogen atom or halogen atom ; and y is selected from the group consisting of hydrogen atom , halogen atom , c 1 - c 4 alkyl group , nitro group , and — ocf 3 group , as well as for its use in therapy , preferably for the treatment of cancer , and to a related pharmaceutical composition , the use of the compound for the manufacture of a medicament for the respective medical indication , and a method of synthesis of the compounds of the invention .

Description:
the general procedures for the preparation of the target derivatives of isatin is described in schemes 1 and 2 . the target compounds can be synthesized via the reaction of the appropriate isatin with hydrazine hydrate to get the corresponding ( z )- 3 - hydrazinyl - indene - 1 - h — or 1 - phenyl - indolin - 2 - one ( 2a - c ) [ 24 ], scheme 1 . 2a - c can be achieved by conventional method or microwave , assisted method ( mwi ). target compounds can also be obtained by conventional method or mwi through coupling the appropriate isatin derivatives with 2a - c as illustrated by scheme 2 . the synthesized compounds were purified by flash chromatography and crystallized from ethanol . the structures were confirmed by spectroscopic methods of analyses . structures of these targets are given in table 1 . 2 . 1 . conventional method : a mixture of isatins 1a - c ( 1 mmol ) and hydrazine hydrate ( 99 %, 0 . 055 g , 1 . 1 mmol ) in absolute methanol ( 25 ml ) was refluxed for 1 h , and then cooled to room temperature . the precipitate of hydrazones was filtered and dried . the crude product was recrystallized from etoh / dmf to give hydrazones 2a - c in 69 - 77 % yield . 2 . 2 . microwave method : the appropriate isatins 1a - c ( 1 mmol ) and hydrazine hydrate ( 99 %, 0 . 055 g , 1 . 1 mmol ) in absolute methanol ( 10 ml ) were placed in the tube of microwave reactor and irradiated at 90 ° c . for 1 min . the temperature of the reaction mixture was adjusted by the computer of the microwave device . then left to cool , the resulting residue was recrystallised from etoh / dmf to afford the corresponding hydrazones 2a - c in 85 - 90 % yield . 3 . 1 . conventional method : a mixture of hydrazones 2a - c ( 1 mmol ) and isatins 1a - h ( 1 mmol ) in ethanol ( 25 ml ) was refluxed for 4 - 6 h , and then cooled to room temperature . the precipitate was filtered and dried . the crude product was recrystallized from etoh / dmf to obtain compounds 3 - 23 in 66 - 89 % yield . 3 . 2 . microwave method : a solution of hydrazones 2a - c ( 1 mmole ) and isatins 1a - h ( 1 mmole ) in ethanol ( 15 ml ) were prepared . few drops of glacial acetic acid were added and whole reaction mixture was irradiated under microwave irradiation at 90 ° c . for 7 minutes . the reaction mixture was cooled . the solid that separated on cooling was filtered , washed with cold ethanol , dried and recrystallised from etoh / dmf . ir ( kbr ) v 3361 - 3199 ( nh , nh 2 ), 1687 ( c ═ o ), 1609 ( c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 87 ( d , 1h , j = 7 . 0 hz , arh ), 6 . 97 ( t , 1h , j = 6 . 5 hz , arh ), 7 . 16 ( t , 1h , j = 6 . 5 hz , arh ), 7 . 37 ( d , 1h , j = 7 . 0 hz , arh ), 9 . 57 ( d , 1h , j = 14 . 0 hz , d 2 o exch ., amino h ), 10 . 56 ( d , 1h , j = 14 . 0 hz , d 2 o exch ., - amino h ), 10 . 71 ( s , d 2 o exch ., 1h , nh ); 13 c nmr δ 109 . 93 , 117 . 43 , 121 . 32 , 126 . 17 , 127 . 0 , 162 . 75 ; ms m / z (%) 161 ( m + , 39 . 7 ), 103 . 7 ( 64 . 3 ), 46 . 8 ( 100 ). ir ( kbr ) v 3365 - 3153 ( nh , nh 2 ), 1682 ( c ═ o ), 1585 ( c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 85 ( m , 1h , arh ), 6 . 97 ( m , 1h , arh ), 7 . 15 ( d , 1h , j = 7 . 5 hz , arh ), 9 . 81 ( d , 1h , j = 15 . 0 hz , d 2 o exch ., amino h ), 10 . 65 ( d , 1h , j = 15 . 0 hz , d 2 o exch ., amino h ), 10 . 72 ( s , d 2 o exch ., 1h , nh ); 13 c nmr δ 104 . 30 ( 2 j f - c = 25 . 3 hz ), 110 . 74 ( 3 j f - c = 8 . 3 hz ), 113 . 10 ( 2 j f - c = 24 . 2 hz ), 123 . 59 ( 3 j f - c = 9 . 2 hz ), 125 . 65 , 134 . 69 , 158 . 10 ( 1 j f - c = 235 . 3 hz ), 162 . 95 ; ms m / z (%) 179 ( m + , 11 . 8 ), 61 . 9 ( 55 . 7 ), 40 . 1 ( 100 ). ir ( kbr ) v 3375 - 3208 ( nh 2 ), 1674 ( c ═ o ), 1592 ( c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 82 ( m , 1h , arh ), 7 . 11 ( m , 1h , arh ), 7 . 20 ( m , 1h , arh ), 7 . 43 - 7 . 53 ( m , 4h , arh ), 7 . 59 ( m , 2h , arh ), 9 . 90 ( d , 1h , j = 15 . 0 hz , d 2 o exch ., amino h ), 10 . 61 ( d , 1h , j = 14 . 5 hz , d 2 o exch ., amino h ); 13 c nmr δ 109 . 06 , 117 . 51 , 122 . 52 , 124 . 63 , 126 . 63 , 126 . 98 , 127 . 86 , 129 . 44 , 129 . 51 , 133 . 79 , 139 . 44 , 159 . 98 ; ms m / z (%) 237 . 1 ( m + , 100 ), 192 ( 60 . 1 ), 51 ( 93 . 6 ). ir ( kbr ) v 3276 ( 2nh ), 1722 ( 2c ═ o ), 1615 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 92 ( d , 2h , j = 7 . 5 hz , arh ), 7 . 02 ( t , 1h , j = 7 . 5 hz , arh ), 7 . 43 ( t , 1h , j = 7 . 5 hz , arh ), 7 . 51 ( d , 1h , j = 7 . 5 hz , arh ), 11 . 02 ( s , 2h , 2nh ); 13 c nmr δ 111 . 09 , 115 . 75 , 122 . 53 , 128 . 17 , 134 . 39 , 144 . 70 , 145 . 16 , 163 . 39 ; ms m / z (%) 290 . 5 ( m + , 6 . 6 ), 46 ( 74 . 9 ), 40 . 1 ( 100 ). ir ( kbr ) v 3448 ( nh ), 1734 ( 2c ═ o ), 1606 ( 2c ═ n ) cm − 1 ; ms m / z (%) 266 . 1 ( m + , 3 . 5 ), 40 . 1 ( 100 ). ir ( kbr ) v 3420 - 3284 ( 2nh ), 1722 ( 2c ═ o ), 1616 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 92 - 7 . 53 ( m , 7h , arh ), 11 . 02 ( s , d 2 o exch ., 2h , 2nh ); 13 c nmr δ 111 . 09 , 112 . 12 ( 3 j f - c = 7 . 4 hz ), 115 . 0 ( 2 j f - c = 25 . 7 hz ), 115 . 77 , 116 . 20 ( 3 j f - c = 8 . 6 hz ), 120 . 77 ( 2 j f - c = 23 . 7 hz ), 122 . 52 , 128 . 48 , 134 . 59 , 141 . 56 , 144 . 70 , 145 . 16 , 145 . 44 , 145 . 64 , 157 . 50 ( 1 j f - c = 263 . 0 hz ), 163 . 39 , 163 . 44 ; ms m / z (%) 308 . 2 ( m + , 5 . 0 ), 46 ( 100 ). ir ( kbr ) v 3420 - 3244 ( 2nh ), 1736 ( 2c ═ o ), 1617 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 86 - 7 . 58 ( m , 7h , arh ), 11 . 03 ( s , d 2 o exch ., 2h , 2nh ); ms m / z (%) 325 . 2 ( m ′+ 1 , 6 . 8 ), 324 . 4 ( m + , 15 ), 78 ( 100 ). ir ( kbr ) v 3239 ( 2nh ), 1735 ( 2c ═ o ), 1612 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 88 - 7 . 66 ( m , 7h , arh ), 11 . 02 ( s , d 2 o exch ., 1h , nh ), 11 . 15 ( s , d 2 o exch ., 1h , nh ); 13 c nmr δ 111 . 11 , 113 . 05 , 113 . 64 , 115 . 79 , 117 . 48 , 122 . 52 , 125 . 05 , 128 . 62 , 130 . 34 , 134 . 62 , 136 . 52 , 144 . 35 , 144 . 76 , 145 . 43 , 145 . 91 , 163 . 06 , 163 . 43 ; ms m / z (%) 369 ( m + , 19 . 0 ), 40 . 1 ( 100 ). ir ( kbr ) v 3286 ( 2nh ), 1723 ( 2c ═ o ), 1615 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 2 . 21 ( s , 3h , ch 3 ), 6 . 82 - 7 . 53 ( m , 7h , arh ), 11 . 01 ( s , d 2 o exch ., 2h , 2nh ); 13 c nmr δ 20 . 53 , 110 . 87 , 111 . 08 , 115 . 75 , 122 . 53 , 128 . 17 , 128 . 48 , 131 . 42 , 134 . 39 , 134 . 74 , 142 . 94 , 144 . 70 , 144 . 81 , 145 . 16 ; ms m / z (%) 303 . 9 ( m + , 4 . 0 ), 40 . 1 ( 100 ). ir ( kbr ) v 3447 ( 2nh ), 1731 ( 2c ═ o ), 1617 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 93 - 8 . 36 ( m , 7h , arh ), 11 . 06 ( s , d 2 o exch ., 1h , nh ), 11 . 69 ( s , d 2 o exch ., 1h , nh ); ms m / z (%) 335 ( m + , 9 . 7 ), 47 . 8 ( 100 ). ir ( kbr ) v 3446 - 3245 ( 2nh ), 1740 ( 2c ═ o ), 1617 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 92 - 7 . 56 ( m , 7h , arh ), 11 . 02 ( s , d 2 o exch ., 1h , nh ), 11 . 20 ( s , d 2 o exch ., 1h , nh ); 13 c nmr δ 111 . 11 , 112 . 18 , 115 . 72 , 116 . 42 , 121 . 12 , 122 . 54 , 127 . 20 , 128 . 67 , 134 . 69 , 142 . 95 , 144 . 26 , 145 . 15 , 145 . 49 , 146 . 00 , 159 . 38 , 163 . 36 , 163 . 43 ; ms m / z (%) 374 ( m + , 7 . 9 ), 44 . 9 ( 100 ). ir ( kbr ) v 1731 ( 2c ═ o ), 1605 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 82 - 7 . 78 ( m , 18h , arh ); 13 c nmr δ 110 . 03 , 115 . 47 , 123 . 53 , 126 . 92 , 128 . 48 , 128 . 62 , 129 . 67 , 133 . 48 , 134 . 38 , 144 . 11 , 146 . 18 , 161 . 53 ; ms m / z (%) 442 . 2 ( m + , 3 . 3 ), 64 ( 100 ). ir ( kbr ) v 3282 ( nh ), 1735 ( 2c ═ o ), 1608 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 82 - 7 . 67 ( m , 12h , arh ), 11 . 07 ( s , d 2 o exch ., 1h , nh ); ms m / z (%) 384 . 2 ( m + , 3 . 5 ), 48 ( 100 ). ir ( kbr ) v 3448 ( nh ), 1736 ( 2c ═ o ), 1609 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 83 - 7 . 63 ( m , 12h , arh ), 11 . 18 ( s , d 2 o exch ., 1h , nh ); ms m / z (%) 400 . 1 ( m + , 5 . 1 ), 63 ( 100 ). ir ( kbr ) v 3448 ( nh ), 1734 ( 2c ═ o ), 1608 ( 2c ═ n ) cm − 1 ; ms m / z (%) 444 . 7 ( m + , 1 . 8 ), 43 . 8 ( 100 ). ir ( kbr ) v 3447 ( nh ), 1736 ( 2c ═ o ), 1609 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 2 . 20 ( s , 3h , ch 3 ), 6 . 83 - 7 . 63 ( m , 12h , arh ), 11 . 18 ( s , d 2 o exch ., 1h , nh ); ms m / z (%) 379 . 9 ( m + , 7 . 4 ), 62 . 9 ( 100 ). ir ( kbr ) v 3447 ( nh ), 1740 ( 2c ═ o ), 1609 ( 2c ═ n ) cm − 1 ; ms m / z (%) 411 . 1 ( m + , 1 . 7 ), 45 . 8 ( 100 ). ir ( kbr ) v 3236 ( nh ), 1747 ( 2c ═ o ), 1608 ( 2c ═ n ) cm − 1 ; ms m / z (%) 449 . 6 ( m + , 2 . 7 ), 49 . 9 ( 100 ). ir ( kbr ) v 3252 ( 2nh ), 1739 ( 2c ═ o ), 1625 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 93 - 7 . 33 ( m , 6h , arh ), 11 . 04 ( s , d 2 o exch ., 2h , 2nh ); 13 c nmr δ112 . 12 ( 3 j f - c = 7 . 2 hz ), 115 . 20 ( 2 j f - c = 25 . 5 hz ), 116 . 24 ( 3 j f - c = 9 . 2 hz ), 120 . 90 ( 2 j f - c = 23 . 8 hz ), 141 . 82 , 145 . 93 , 157 . 50 ( 1 j f - c = 238 . 0 hz ), 163 . 49 ; ms m / z (%) 326 ( m + , 11 ), 44 . 9 ( 100 ). ir ( kbr ) v 3245 ( 2nh ), 1736 ( 2c ═ o ), 1618 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 94 - 7 . 54 ( m , 6h , arh ), 11 . 04 ( s , d 2 o exch ., 1h , nh ), 11 . 14 ( s , d 2 o exch ., 1h , nh ); 13 c nmr δ 112 . 08 ( 3 j f - c = 7 . 5 hz ), 112 . 60 , 115 . 35 ( 2 j f - c = 25 . 7 hz ), 116 . 26 ( 3 j f - c = 8 . 9 hz ), 117 . 05 , 120 . 90 ( 2 j f - c = 23 . 9 hz ), 126 . 03 , 127 . 82 , 133 . 89 , 141 . 81 , 144 . 24 , 145 . 59 , 146 . 13 , 157 . 50 ( 1 j f - c = 237 . 9 hz ), 163 . 24 , 163 . 50 ; ms m / z (%) 342 ( m + , 17 . 9 ), 63 ( 100 ). ir ( kbr ) v 3245 ( 2nh ), 1735 ( 2c ═ o ), 1616 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 89 - 7 . 66 ( m , 6h , arh ), 11 . 04 ( s , d 2 o exch ., 1h , nh ), 11 . 14 ( s , d 2 o exch ., 1h , nh ); 13 c nmr δ 112 . 14 ( 3 j f - c = 7 . 0 hz ), 113 . 06 , 113 . 61 , 115 . 37 ( 2 j f - c = 25 . 3 hz ), 116 . 24 ( 3 j f - c = 8 . 9 hz ), 117 . 54 , 120 . 90 ( 2 j f - c = 23 . 7 hz ), 130 . 57 , 136 . 69 , 141 . 81 , 144 . 61 , 145 . 50 , 146 . 18 , 157 . 50 ( 1 j f - c = 238 . 0 hz ), 163 . 11 , 163 . 50 ; ms m / z (%) 387 ( m + , 8 . 5 ), 46 . 9 ( 100 ). ir ( kbr ) v 3392 - 3186 ( 2nh ), 1735 ( 2c ═ o ), 1623 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 2 . 22 ( s , 3h , ch 3 ), 6 . 81 - 7 . 33 ( m , 6h , arh ), 10 . 91 ( s , d 2 o exch ., 1h , nh ), 11 . 02 ( s , d 2 o exch ., 1h , nh ); ms m / z (%) 322 . 3 ( m + , 6 . 9 ), 40 . 1 ( 100 ). ir ( kbr ) v 3248 ( 2nh ), 1737 ( 2c ═ o ), 1624 ( 2c ═ n ) cm − 1 ; 1 h nmr ( dmso - d 6 ) δ 6 . 93 - 8 . 35 ( m , 6h , arh ), 11 . 08 ( s , d 2 o exch ., 1h , nh ), 11 . 71 ( s , d 2 o exch ., 1h , nh ); ms m / z (%) 353 . 5 ( m + , 12 . 4 ), 63 ( 100 ). ir ( kbr ) v 3246 ( 2nh ), 1735 ( 2c ═ o ), 1624 ( 2c ═ n ) cm − 1 ; ms m / z (%) 392 . 4 ( m + , 8 . 9 ), 62 . 9 ( 100 ). the cytotoxicity of the prepared compounds was evaluated at laboratory of cell biology , national cancer institute , national institutes of health , bethesda , md . 20892 , group in biomolecular , usa , using the following protocol : kb - 3 - 1 cells ( a hela deriviative ) and its mdr derivative ( kb - v1 ) were grown as previously described [ 24 ]. cell survival was measured by the mtt ( 3 -( 4 , 5 - dimethylthiazol - 2 - yl )- 2 , 5 - diphenyltetrazolium bromide ) assay as previously described [ 24 ]. briefly , cells were seeded in 100 μl of growth medium at a density of 5000 cells / well in 96 - well plates and allowed to establish for 24 h , at which time serially diluted drugs were added in an additional 100 μl of growth medium . cells were then incubated for 72 h at 37 ° c . in humidified 5 % co 2 , at which time the growth medium was drawn and replaced with mtt in imdm growth medium and incubated for 4 h . the mtt solution was then drawn from the wells , and 100 μl of acidified ethanol solution was added to each well and after 15 min absorption at 560 nm was measured . ic 50 cytotoxicity values were determined as the drug concentration that reduced the absorbance to 50 % of that in untreated control wells and derived from at least three separate experiments · hours , treated with the specified compound or vehicle ( 0 . 1 % dmso final ) control , and incubated at 37 ° c . an additional 72 hours . the effect of treatment on cell viability was determined using the luminescent cell titer glo assay ( promega ). results are given in table 2 . variable and promising activity and selectivity revealed by the synthesized compounds against mdr cells , results are given in table 2 . accordingly the synthesized bis - isatin derivatives are potential candidates for treatment mdr cancer . a the mdr1 selectivity ( rr ) is calculated as the ratio of a compound &# 39 ; 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