QUINOLINE COMPOUNDS AND PHARMACEUTICAL COMPOSITIONS AND USES THEREOF
20220220106 · 2022-07-14
Inventors
- Wu Zhong (Beijing, CN)
- Tianlong HAO (Beijing, CN)
- Ruiyuan Cao (BEIJING, CN)
- Tong CHENG (Xiamen, Fujian, CN)
- Yuexiang LI (Beijing, CN)
- Shiyong Fan (BEIJING, CN)
- Wei Li (Beijing, CN)
- Shixu WANG (Beijing, CN)
- Ningshao XIA (XIAMEN, FUJIAN, CN)
- Song Li (Beijing, CN)
Cpc classification
C07D405/04
CHEMISTRY; METALLURGY
A61P1/00
HUMAN NECESSITIES
C07D401/04
CHEMISTRY; METALLURGY
C07D409/04
CHEMISTRY; METALLURGY
C07D215/44
CHEMISTRY; METALLURGY
International classification
C07D215/44
CHEMISTRY; METALLURGY
C07D401/04
CHEMISTRY; METALLURGY
C07D405/04
CHEMISTRY; METALLURGY
C07D409/04
CHEMISTRY; METALLURGY
Abstract
Provided are quinoline derivative compounds of Formulae (I), (II) and (III) with an inhibitory effect on mTOR and applications of their pharmaceutically acceptable salts, their stereoisomers, their hydrates or their solvates in preparation of medicine for preventing and/or treating diseases caused by enteroviruses.
##STR00001##
Claims
1. A compound represented by Formula I, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof, ##STR00137## wherein: R.sub.1 is C.sub.1-10 alkyl, 3- to 14-membered cycloalkyl, C.sub.2-12 alkenyl or polyenyl, C.sub.2-12 alkenoyl or polyenoyl, 2- to 10-membered alkanoyl, 6- to 14-membered substituted or unsubstituted aryl, 3- to 14-membered substituted or unsubstituted heterocyclyl; R.sub.2 is hydrogen, or C.sub.1-10 alkyl, 3- to 14-membered cycloalkyl, C.sub.2-12 alkenyl or polyenyl, C.sub.2-12 alkenoyl or polyenoyl, 2- to 10-membered alkanoyl, 6- to 14-membered substituted or unsubstituted aryl, 3- to 14-membered substituted or unsubstituted heterocyclyl, 7- to 12-membered substituted or unsubstituted bridged ring group, amino, 6- to 14-membered substituted or unsubstituted arylimino, R.sub.3 is C.sub.1-10 alkyl, 3- to 14-membered cycloalkyl, C.sub.2-12 alkenyl or polyenyl, C.sub.2-12 alkenoyl or polyenoyl, 2- to 10-membered alkanoyl, 6- to 14-membered substituted or unsubstituted aryl, 3- to 14-membered substituted or unsubstituted heterocyclyl; R.sub.4 is hydrogen, or C.sub.1-10 alkyl, 3- to 14-membered cycloalkyl, C.sub.2-12 alkenyl or polyenyl C.sub.2-12 alkenoyl or polyenoyl, 2- to 10-membered alkanoyl, 6- to 14-membered substituted or unsubstituted aryl, 3- to 14-membered substituted or unsubstituted heterocyclyl, 7- to 12-membered substituted or unsubstituted bridged ring group, amino, 6- to 14-membered substituted or unsubstituted arylimino; R.sub.5 is C.sub.1-10 alkyl, 3- to 14-membered cycloalkyl, C.sub.2-12 alkenyl or polyenyl, C.sub.2-12 alkenoyl or polyenoyl, 2- to 10-membered alkanoyl, 6- to 14-membered substituted or unsubstituted aryl, 3- to 14-membered substituted or unsubstituted heterocyclyl; R.sub.6 is hydrogen, or C.sub.1-10 alkyl, 3- to 14-membered cycloalkyl, C.sub.2-12 alkenyl or polyenyl C.sub.2-12 alkenoyl or polyenoyl, 2- to 10-membered alkanoyl, 6- to 14-membered substituted or unsubstituted aryl, 3- to 14-membered substituted or unsubstituted heterocyclyl, 7- to 12-membered substituted or unsubstituted bridged ring group, amino, 6- to 14-membered substituted or unsubstituted arylimino.
2. (canceled)
3. (canceled)
4. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1, wherein: R.sub.1 is 5- to 6-membered cycloalkyl, 5- to 6-membered heterocyclyl, 5- to 6-membered aryl or 5- to 6-membered heteroaryl, R.sub.1 is optionally substituted by one or more R.sup.a, each R.sup.a is independently hydrogen, C.sub.1-6 alkyl, C.sub.1-6 haloalkyl, C.sub.1-6 alkoxy, alkyl-amino, hydroxy, nitro, cyano, C.sub.1-6 alkylthio, dialkyl-amino, halogen or amino; R.sub.3 is 5- to 6-membered cycloalkyl, 5- to 6-membered heterocyclyl, 5- to 6-membered aryl or 5- to 6-membered heteroaryl, R.sub.3 is optionally substituted by one or more R.sup.b, each R.sup.b is independently hydrogen, C.sub.1-6 alkyl, C.sub.1-6 haloalkyl, C.sub.1-6 alkoxy, alkyl-amino, hydroxy, nitro, cyano, C.sub.1-6 alkylthio, dialkyl-amino, halogen or amino; R.sub.5 is 5- to 6-membered cycloalkyl, 5- to 6-membered heterocyclyl, 5- to 6-membered aryl or 5- to 6-membered heteroaryl, and R.sub.5 is optionally substituted by one or more R.sup.c, each R.sup.c is independently C.sub.1-6 alkyl, C.sub.1-6 haloalkyl, C.sub.1-6 alkoxy, alkyl-amino, hydroxy, nitro, cyano, C.sub.1-6 alkylthio, dialkyl-amino, halogen or amino.
5. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1, wherein: R.sub.2 is pyridyl, phenyl, furyl, pyrazolyl, thienyl, quinolyl, R.sub.2 is optionally substituted by one or more R.sup.d, and each R.sup.d is independently C.sub.1-6 alkyl, C.sub.1-6 haloalkyl, C.sub.1-6 alkoxy, alkyl-amino, hydroxyl, nitro, cyano, C.sub.1-6 alkylthio, dialkyl-amino, halogen, amino, NH.sub.2C(O)—, R′OC(O)NH—, wherein R′ is benzyl, phenyl or C.sub.1-6 alkyl; R.sub.4 is pyridyl, phenyl, furyl, pyrazolyl, thienyl, quinolyl, vinyl, propenyl, or butenyl, R.sub.4 is optionally substituted by one or more R.sup.e, each R.sup.e is independently hydrogen, C.sub.1-6 alkyl, C.sub.1-6 haloalkyl, C.sub.1-6 alkoxy, alkyl-amino, hydroxy, nitro, cyano, C.sub.1-6 alkylthio, dialkyl-amino, halogen, amino, NH.sub.2C(O)—, NH.sub.2C(O)NH—, R′OC(O)—, wherein R′ is benzyl, phenyl, or C.sub.1-6 alkyl, R″OC(O)NH—, wherein R″ is benzyl, phenyl, or C.sub.1-6 alkyl, R″′C(O)NH—, wherein R″′ is benzyl, phenyl, or C.sub.1-6 alkyl; R.sub.6 is ##STR00138## wherein R.sup.f, R.sup.g, R.sup.h, R.sup.i are each independently hydrogen, C.sub.1-6 alkyl, C.sub.1-6 haloalkyl, C.sub.1-6 alkoxy, alkyl-amino, hydroxyl, nitro, cyano, C.sub.1-6 alkylthio, dialkyl-amino, halogen, amino, C.sub.1-6 alkyl-C(O)NH—; or R.sub.6 is pyridyl, phenyl, quinolyl, 2-oxoindolyl, pyrimidyl, isoxazolyl, 1,4-dioxa-spiro[4.5]dec-7-ene group, or pyrazolyl, R.sub.6 is optionally substituted by one or more R.sup.j, each R.sup.j is independently hydrogen, C.sub.1-6 alkyl, C.sub.1-6 alkoxy, halogen, alkyl-amino, dialkyl, 4-methyl-piperazinyl, morpholinyl, amino, R.sup.k—C(O)NH—, wherein R.sup.k— is benzyl, phenyl, p-methoxybenzyl, phenoxy or C.sub.1-6 alkyl, pyrrolidinyl, allylamino or propargylamino; or R.sub.6 is C.sub.2-6 alkenyl, R.sub.6 is optionally substituted by one or more R.sup.m, each R.sup.m is independently NH.sub.2C(O)—, NH.sub.2C(O)NH—, R.sup.n—OC(O), wherein R.sup.n is C.sub.1-6 alkyl.
6. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1, wherein R.sub.1 is; ##STR00139## R.sub.2 is ##STR00140## ##STR00141##
7. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 21, wherein R.sub.3 is ##STR00142## R.sub.4 is ##STR00143## ##STR00144##
8. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1, wherein R.sub.5 is ##STR00145## R.sub.6 is ##STR00146## ##STR00147## ##STR00148##
9. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1, wherein the compound is preferably selected from the group consisting of: (E)-6-[3-(6-amino)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-34); (E)-6-(4-amino)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-35); (E)-6-[3-(6-methyl)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-38); (E)-6-(3-aminophenyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-42); (E)-6-(4-pyridyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-04); (E)-6-(5-methoxy)pyridyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-07); (E)-6-(4-hydroxy)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-11); (E)-6-(4-fluoro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-12); (E)-6-(4-cyano)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-15); (E)-6-[4-(trifluoromethyl)phenyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-18); (E)-6-(3-ethoxy)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-16); (E)-6-(3-furyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-22); (E)-6-(2-thienyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-24); (E)-6-(3-quinolyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-25); (E)-6-(1H-4-pyrazolyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-26); (E)-6-(4-fluoro-3-methyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-32); (E)-6-(2,4-difluoro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-33); (E)-6-(3,4-difluoro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-34); (E)-6-(3-chloro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-36); (E)-6-(4-chloro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-37); (E)-6-(4-isopropyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-39); (E)-6-(4-propyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-40); (E)-6-(4-isobutyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-41); (E)-6-(4-butyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-42); (E)-6-[3-(6-fluoro)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-43); (E)-6-(4-carbamoyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-45); (E)-6-[3-(5-cyano)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-46); (E)-6-[3-(N-6-benzyloxyamido)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-7-22); 8-[3-(6-amino)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-4-32); 8-(4-carbamoyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-21); 8-[3-(6-methyl)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-23); 8-(3-furyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-25); 8-(2-thienyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-26); 8-(4-ethoxy)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-27); 8-[3-(6-fluoro)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-28); 8-(4-trifluoromethyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-29); 8-[3-(5-methoxy)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-30); 8-(3-quinolyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-32); 8-(4-chloro)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-33); 8-(4-propyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-34); 8-(4-isopropyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-35); 8-[3-(5-cyano)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-36); 8-[3-(-6-valerylamino)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (WSX-1-24); 8-[3-(-6-benzyloxyamido)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-7-23); (E)-8-(2-carbamoyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-30); (E)-8-(3-cyanopropenyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-31); (E)-8-(2-cyanovinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-32); (E)-8-(2-methoxycarbonyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-33); (E)-8-(3-ureido-propenyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-34); (E)-8-(2-tert-butoxycarbonyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-35); (E)-8-(4-ethoxycarbonyl-but-1-enyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-38); (E)-8-(2-ethoxycarbonyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (WSX-1-13); 9-[3-(6-propionylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-45); 9-(6-quinolyl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-47); 9-[3-(2-fluoro)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-48); 9-[3-(2-methyl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-49); 9-[(1H)-3-pyrazolyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-50); 9-[3-(6-fluoro)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-01); 9-[3-(2-methoxy)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-02); 9-[3-(6-methoxy)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-03); 9-[5-(2-oxo)indolyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-04) 9-[3-(6-butyrylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-05); 9-[5-(2-methoxy)pyrimidyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-06); 9-[3-(2-isobutyrylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-07); 9-[3-(6-valerylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-08); 9-[3-(6-phenylacetamide)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-10); 9-{3-[6-(4-methoxy)phenylacetylamino]pyridyl}-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-11); 9-[4-(3,5-dimethyl)isoxazolyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-12); 9-(1,4-dioxa-spiro[4.5]dec-7-en-8-yl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-13); 9-[3-(6-benzyloxyamido)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-14); 9-[3-(-6-phenoxyamido)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-15); 9-[3-(6-N,N-dimethyl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-16); 9-{3-[6-(4-methylpiperazin-1-yl)pyridyl]}-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-17); 9-[3-(6-morpholin-4-yl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-18); 9-[3-(6-amino-5-methoxy)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-19); 9-[3-(6-pyrrolidinyl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-20); 9-[3-(6-tert-butoxycarbonylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-21); 9-[3-(N-6-allylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-28); 9-[3-(6-propargylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-29); 9-(3-aminophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-11); 9-[2-(6-aminopyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-12); 9-[4-(trifluoromethyl)phenyl]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-16); 9-(3,4-dimethylphenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-17); 9-(4-tert-butylphenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-18); 9-[3-(6-methylpyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-19); 9-[3-(6-fluoropyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-20); 9-[3-(6-chloropyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-21); 9-(4-cyanophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-22); 9-(4-fluorophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-23); 9-[(4-fluoro-3-methyl)phenyl]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-24); 9-(4-chlorophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-25); 9-(4-methoxyphenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-26); 9-(N-4-acetylaminophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-27); 9-[2-(6-aminopyrazinyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-28); 9-[3-(5-aminopyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-29); (E)-9-(2-carbamoyl-vinyl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-24); (E)-9-(3-ureido-propenyl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-25); and (E)-9-(2-ethoxycarbonyl-vinyl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-26).
10. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1, wherein the pharmaceutically acceptable salt is an inorganic acid salt of the compound or an organic acid salt of the compound.
11. A pharmaceutical composition, comprising at least one compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1, and one or more pharmaceutically acceptable carriers or excipients.
12. (canceled)
13. (canceled)
14. (canceled)
15. A method for the treatment and/or prevention of a disease or condition associated with viral infection, the method comprising administering to a subject in need a therapeutically and/or prophylactically effective amount of at least one compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1.
16. A method for inhibiting the replication of an enterovirus in a mammal in need, the method comprising administering to the mammal in need a therapeutically and/or prophylactically effective amount of at least one compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 1.
17. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 4, wherein: R.sub.1 is phenyl, R.sub.1 is optionally substituted by one or more R.sup.a, each R.sup.a is independently hydrogen, trifluoromethyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, methoxy, ethoxy, propoxy, methylamino, ethylamino, dimethylamino, diethylamino, hydroxyl, nitro, cyano, methylthio, ethylthio, fluorine, chlorine, bromine, iodine, or amino; R.sub.3 is phenyl, R.sub.3 is optionally substituted with one or more R.sup.b, and each R.sup.b is independently hydrogen, trifluoromethyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, methoxy, ethoxy, propoxy, methylamino, ethylamino, dimethylamino, diethylamino, hydroxyl, nitro, cyano, methylthio, ethylthio, fluorine, chlorine, bromine, iodine, or amino; R.sub.5 is phenyl, R.sub.5 is optionally substituted by one or more R.sup.c, each R.sup.c is independently trifluoromethyl, methyl, ethyl, n-propyl, isopropyl, n-butyl, methoxy, ethoxy, propoxy, methylamino, ethylamino, dimethylamino, diethylamino, hydroxyl, nitro, cyano, methylthio, ethylthio, fluorine, chlorine, bromine, iodine, or amino.
18. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 4, wherein the alkyl-amino is C.sub.1-6 alkyl-amino, the dialkyl-amino is di(C.sub.1-6 alkyl)-amino.
19. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 5, wherein the alkyl-amino is C.sub.1-6 alkyl-amino, the dialkyl-amino is di(C.sub.1-6 alkyl)-amino.
20. The compound, a pharmaceutically acceptable salt, a stereoisomer, a hydrate or a solvate thereof according to claim 5, wherein the pharmaceutically acceptable salt is a hydrochloride, a sulfate, a phosphate, a methanesulfonate, a trifluoromethanesulfonate, an acetate, a trifluoroacetate, or a benzoate of the compound.
21. The method according to claim 15, wherein the viral infection is an infection caused by an enterovirus.
22. The method according to claim 21, wherein the enterovirus is EV71.
23. The method according to claim 21, wherein the disease or condition associated with viral infection is hand-foot-and-mouth disease.
24. The method according to claim 16, wherein the enterovirus is EV71.
Description
BRIEF DESCRIPTION OF THE DRAWINGS
[0384]
SPECIFIC MODELS FOR CARRYING OUT THE APPLICATION
[0385] The content of the present application will be described in detail through the following examples. In the present application, the following examples are used to better illustrate the application, and not used to limit the scope of the application.
Example 1
Synthesis of (E)-6-[3-(6-amino)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-34)
[0386] 80 mL microwave tube was taken, to which (E)-4-{6-bromo-4-[3-(trifluoromethylphenyl)amino]}quinoline-3-butenone (4.34 g, 10 mmol), Pd(Ph.sub.3P).sub.4 (1.16 g, 1 mmol), K.sub.2CO.sub.3 (2.76 g, 20 mmol) and 6-aminopyridineboronic acid (1.66 g, 12 mmol) were added and dissolved in a solution of 1,4-dioxane. The microwave tube was sealed with cover, placed in a microwave reactor, the temperature was set to 100° C., the reaction was carried out for 40 minutes, the microwave tube was taken out, and cooled to room temperature. The reaction solution was transferred to a separatory funnel, to which water was added, and the resulting mixture was extracted with ethyl acetate. The organic layer was washed with saturated NaCl, dried with anhydrous Na.sub.2SO.sub.4, filtered, the filtrate was dried by a rotary evaporator and purified with column chromatography (n-hexane/ethyl acetate 5:1) to obtain 3.71 g of a yellow solid, with a yield of 83%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.40 (s, 1H), 9.05 (s, 1H), 8.35 (d, J=10.6 Hz, 2H), 8.02 (d, J=9.9 Hz, 2H), 7.81 (d, J=8.5 Hz, 1H), 7.49-7.40 (m, 2H), 7.22 (d, J=7.6 Hz, 1H), 7.13 (d, J=8.1 Hz, 2H), 6.88 (d, J=16.5 Hz, 1H), 6.56 (d, J=8.6 Hz, 1H), 6.20 (s, 2H), 1.99 (d, J=11.0 Hz, 3H). MS (ESI): m/z 449.15 [M+H].sup.+. Mp 172-175° C.
Example 2
Synthesis of (E)-6-(4-amino)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-35)
[0387] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 29%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.46 (s, 1H), 9.06 (s, 1H), 8.37 (d, J=1.6 Hz, 1H), 8.03 (d, J=8.7 Hz, 1H), 7.96 (dd, J=8.7, 1.8 Hz, 1H), 7.42 (t, J=3.8 Hz, 1H), 7.38 (d, J=4.4 Hz, 1H), 7.17 (d, J=7.8 Hz, 1H), 7.13-7.07 (m, 3H), 6.92 (d, J=1.7 Hz, 1H), 6.87 (s, 1H), 6.84 (d, J=5.0 Hz, 1H), 6.59 (dd, J=8.0, 1.4 Hz, 1H), 5.18 (s, 2H), 1.97 (s, 3H). MS (ESI): m/z 448.16 [M+H].sup.+. Mp 183-185° C.
Example 3
Synthesis of (E)-6-[3-(6-methyl)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-38)
[0388] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 19%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.43 (s, 1H), 9.04 (s, 1H), 8.81 (d, J=2.0 Hz, 1H), 8.47 (s, 1H), 8.10-7.98 (m, 3H), 7.45-7.30 (m, 3H), 7.19 (d, J=7.8 Hz, 1H), 7.12 (d, J=8.9 Hz, 2H), 6.83 (d, J=16.5 Hz, 1H), 2.47 (s, 3H), 1.95 (s, 3H). MS (ESI): m/z 448.16 [M+H].sup.+. Mp 172-174° C.
Example 4
Synthesis of (E)-6-(3-aminophenyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-2-42)
[0389] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 19%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.45 (s, 1H), 9.06 (s, 1H), 8.35 (s, 1H), 8.03 (d, J=8.7 Hz, 1H), 7.96 (dd, J=8.7, 1.5 Hz, 1H), 7.40 (dd, J=15.8, 7.0 Hz, 2H), 7.18 (d, J=7.7 Hz, 1H), 7.09 (dd, J=7.9, 4.5 Hz, 3H), 6.86 (dd, J=24.7, 8.4 Hz, 3H), 6.57 (d, J=6.7 Hz, 1H), 5.18 (s, 2H), 1.97 (s, 3H). MS (ESI): m/z 448.16 [M+H].sup.+. Mp 184-186° C.
Example 5
Synthesis of (E)-6-(4-pyridyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-04)
[0390] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 21%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.51 (s, 1H), 9.08 (s, 1H), 8.69-8.61 (m, 3H), 8.18 (dd, J=8.7, 1.9 Hz, 1H), 8.09 (d, J=8.7 Hz, 1H), 7.79 (dd, J=4.6, 1.5 Hz, 2H), 7.43 (dd, J=10.2, 6.3 Hz, 1H), 7.34 (d, J=16.5 Hz, 1H), 7.21 (d, J=7.7 Hz, 1H), 7.17-7.11 (m, 2H), 6.83 (d, J=16.5 Hz, 1H), 1.94 (s, 3H). MS (ESI): m/z 434.14 [M+H].sup.+. Mp 192-195° C.
Example 6
Synthesis of (E)-6-[3-(5-methoxy)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-07)
[0391] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 20%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.49 (s, 1H), 9.10 (s, 1H), 8.56 (dd, J=25.3, 1.4 Hz, 2H), 8.33 (d, J=2.7 Hz, 1H), 8.19 (dd, J=8.7, 1.7 Hz, 1H), 8.10 (d, J=8.7 Hz, 1H), 7.72-7.64 (m, 1H), 7.52-7.36 (m, 2H), 7.24 (d, J=7.7 Hz, 1H), 7.16 (d, J=6.9 Hz, 2H), 6.88 (d, J=16.4 Hz, 1H), 3.90 (s, 3H), 2.00 (s, 3H). MS (ESI): m/z 464.15 [M+H].sup.+. Mp 164-167° C.
Example 7
Synthesis of (E)-6-(4-hydroxy)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-11)
[0392] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 22%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.68 (s, 1H), 9.47 (s, 1H), 9.07 (s, 1H), 8.38 (s, 1H), 8.12-8.00 (m, 2H), 7.62 (d, J=8.6 Hz, 2H), 7.45 (dd, J=16.1, 8.9 Hz, 2H), 7.23 (d, J=7.7 Hz, 1H), 7.15 (d, J=8.8 Hz, 2H), 6.98-6.81 (m, 3H), 2.00 (d, J=7.8 Hz, 3H). MS (ESI): m/z 449.14 [M+H].sup.+. Mp 218-222° C.
Example 8
Synthesis of (E)-6-(4-fluoro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-12)
[0393] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a green solid, with a total yield of 24%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.50 (s, 1H), 9.09 (s, 1H), 8.47 (s, 1H), 8.14-8.04 (m, 2H), 7.83 (dd, J=8.6, 5.5 Hz, 2H), 7.40 (ddd, J=26.4, 17.1, 8.5 Hz, 4H), 7.22 (d, J=7.7 Hz, 1H), 7.15 (d, J=5.9 Hz, 2H), 6.88 (d, J=16.5 Hz, 1H), 1.99 (s, 3H). MS (ESI): m/z 451.14 [M+H].sup.+. Mp 167-169° C.
Example 9
Synthesis of (E)-6-(4-cyano)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-15)
[0394] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 25%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.54 (s, 1H), 9.07 (s, 1H), 8.58 (d, J=1.5 Hz, 1H), 8.15 (dd, J=8.8, 1.8 Hz, 1H), 8.09 (d, J=8.7 Hz, 1H), 7.97 (q, J=8.6 Hz, 4H), 7.45 (t, J=7.8 Hz, 1H), 7.34 (d, J=16.5 Hz, 1H), 7.23 (d, J=7.6 Hz, 1H), 7.16 (d, J=8.0 Hz, 2H), 6.83 (d, J=16.5 Hz, 1H), 1.95 (s, 3H). MS (ESI): m/z 458.14 [M+H].sup.+. Mp 173-174° C.
Example 10
Synthesis of (E)-6-[4-(trifluoromethyl)phenyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-18)
[0395] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 23%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.53 (s, 1H), 9.11 (s, 1H), 8.59 (d, J=1.6 Hz, 1H), 8.15 (dt, J=18.4, 5.2 Hz, 2H), 8.01 (d, J=8.2 Hz, 2H), 7.86 (d, J=8.3 Hz, 2H), 7.51-7.34 (m, 2H), 7.23 (d, J=7.8 Hz, 1H), 7.16 (d, J=6.7 Hz, 2H), 6.88 (d, J=16.5 Hz, 1H), 1.98 (s, 3H). MS (ESI): m/z 501.13 [M+H].sup.+. Mp 189-191° C.
Example 11
Synthesis of (E)-6-(3-ethoxy)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-16)
[0396] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 19%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.51 (s, 1H), 9.07 (s, 1H), 8.42 (d, J=1.5 Hz, 1H), 8.08 (dt, J=16.3, 5.2 Hz, 2H), 7.71 (d, J=8.8 Hz, 2H), 7.44 (dd, J=22.5, 12.1 Hz, 2H), 7.23 (d, J=7.7 Hz, 1H), 7.15 (d, J=7.3 Hz, 2H), 7.04 (d, J=8.8 Hz, 2H), 6.87 (d, J=16.5 Hz, 1H), 4.07 (q, J=7.0 Hz, 2H), 1.99 (s, 3H), 1.34 (t, J=7.0 Hz, 3H). MS (ESI): m/z 477.17 [M+H].sup.+. Mp 184-185° C.
Example 12
Synthesis of (E)-6-(3-furyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-22)
[0397] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 23%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.37 (s, 1H), 9.04 (s, 1H), 8.37 (d, J=1.3 Hz, 1H), 8.30 (s, 1H), 8.04 (dt, J=24.3, 5.1 Hz, 2H), 7.79 (t, J=1.5 Hz, 1H), 7.47-7.35 (m, 2H), 7.22 (d, J=7.7 Hz, 1H), 7.14 (d, J=7.4 Hz, 2H), 6.98 (d, J=0.8 Hz, 1H), 6.86 (d, J=16.5 Hz, 1H), 1.98 (s, 3H). MS (ESI): m/z 423.12 (M+H].sup.+. Mp 151-154° C.
Example 13
Synthesis of (E)-6-(2-thienyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-24)
[0398] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 25%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.46 (s, 1H), 9.07 (s, 1H), 8.37 (d, J=1.8 Hz, 1H), 8.10 (dd, J=8.8, 2.0 Hz, 1H), 8.03 (d, J=8.7 Hz, 1H), 7.62 (ddd, J=6.1, 4.3, 1.0 Hz, 2H), 7.49-7.41 (m, 2H), 7.24-7.11 (m, 4H), 6.90 (d, J=16.4 Hz, 1H), 2.03 (s, 3H). MS (ESI): m/z 439.10 [M+H].sup.+. Mp 181-185° C.
Example 14
Synthesis of (E)-6-(3-quinolyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-25)
[0399] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 21%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.56 (s, 1H), 9.36 (d, J=2.1 Hz, 1H), 9.12 (s, 1H), 8.74 (d, J=7.3 Hz, 2H), 8.33 (d, J=8.6 Hz, 1H), 8.17 (d, J=8.7 Hz, 1H), 8.12-8.01 (m, 2H), 7.80 (t, J=7.7 Hz, 1H), 7.67 (t, J=7.5 Hz, 1H), 7.53-7.34 (m, 2H), 7.27 (d, J=7.8 Hz, 1H), 7.21 (d, J=1.9 Hz, 2H), 6.88 (d, J=16.4 Hz, 1H), 1.99 (s, 3H). MS (ESI): m/z 484.16 [M+H].sup.+. Mp 173-176° C.
Example 15
Synthesis of (E)-6-(1H-4-pyrazolyl)-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-26)
[0400] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 18%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 13.05 (s, 1H), 9.37 (s, 1H), 9.02 (s, 1H), 8.39 (s, 1H), 8.03 (dd, J=36.9, 8.7 Hz, 4H), 7.52-7.33 (m, 2H), 7.23 (d, J=7.6 Hz, 1H), 7.15 (d, J=8.0 Hz, 2H), 6.87 (d, J=16.5 Hz, 1H), 1.99 (s, 3H). MS (ESI): m/z 423.14 [M+H].sup.+. Mp 177-179° C.
Example 16
Synthesis of (E)-6-(4-fluoro-3-methyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-32)
[0401] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 19%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 10.54 (s, 1H), 9.07 (s, 1H), 8.74 (s, 1H), 8.24 (dd, J=8.8, 1.6 Hz, 1H), 8.13 (d, J=8.8 Hz, 1H), 7.73 (ddd, J=10.4, 8.3, 4.8 Hz, 2H), 7.57 (t, J=7.8 Hz, 1H), 7.43 (d, J=7.4 Hz, 3H), 7.31-7.23 (m, 1H), 7.17 (d, J=16.4 Hz, 1H), 6.74 (d, J=16.4 Hz, 1H), 2.32 (d, J=1.2 Hz, 3H), 1.89 (s, 3H). MS (ESI): m/z 465.15 [M+H].sup.+. Mp 209-210° C.
Example 17
Synthesis of (E)-6-(2,4-difluoro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-33)
[0402] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 20%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 10.99 (s, 1H), 9.12 (s, 1H), 8.84 (s, 1H), 8.25 (d, J=8.8 Hz, 1H), 8.17 (d, J=8.7 Hz, 1H), 7.81 (td, J=8.9, 6.7 Hz, 1H), 7.65-7.42 (m, 5H), 7.29 (td, J=8.5, 2.4 Hz, 1H), 7.06 (d, J=16.4 Hz, 1H), 6.70 (d, J=16.4 Hz, 1H), 1.85 (s, 3H). MS (ESI): m/z 469.13 [M+H].sup.+. Mp 162-164° C.
Example 18
Synthesis of (E)-6-(3,4-difluoro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-34)
[0403] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 23%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 11.42 (s, 1H), 9.13 (d, J=1.3 Hz, 1H), 9.05 (s, 1H), 8.39 (dd, J=8.9, 1.7 Hz, 1H), 8.23 (d, J=8.9 Hz, 1H), 8.14 (ddd, J=12.2, 7.7, 2.2 Hz, 1H), 7.85 (dd, J=5.8, 2.8 Hz, 1H), 7.62 (ddd, J=23.6, 9.2, 2.8 Hz, 5H), 6.92 (d, J=16.4 Hz, 1H), 6.63 (d, J=16.3 Hz, 1H), 1.79 (s, 3H). MS (ESI): m/z 469.13 [M+H].sup.+. Mp 210-213° C.
Example 19
Synthesis of (E)-6-(3-chloro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-36)
[0404] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 27%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.62 (s, 1H), 9.11 (d, J=7.8 Hz, 1H), 8.54 (d, J=1.6 Hz, 1H), 8.21-8.06 (m, 2H), 7.89-7.73 (m, 2H), 7.58-7.45 (m, 3H), 7.39 (d, J=16.5 Hz, 1H), 7.27 (d, J=7.7 Hz, 1H), 7.21 (d, J=6.3 Hz, 2H), 6.91-6.82 (m, 1H), 1.99 (s, 3H). MS (ESI): m/z 467.11 [M+H].sup.+. Mp 182-183° C.
Example 20
Synthesis of (E)-6-(4-chloro)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-37)
[0405] It was synthesized according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 28%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.95 (s, 1H), 9.07 (s, 1H), 8.64 (d, J=1.6 Hz, 1H), 8.16 (dd, J=8.8, 1.9 Hz, 1H), 8.09 (d, J=8.7 Hz, 1H), 7.90-7.81 (m, 2H), 7.61-7.53 (m, 2H), 7.49 (t, J=8.2 Hz, 1H), 7.35-7.18 (m, 4H), 6.81 (d, J=16.4 Hz, 1H), 1.93 (s, 3H). MS (ESI): m/z 467.11 [M+H].sup.+. Mp 197-199° C.
Example 21
Synthesis of (E)-6-(4-isopropyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-39)
[0406] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 26%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 10.04 (s, 1H), 9.09 (s, 1H), 8.61 (s, 1H), 8.19 (dd, J=8.8, 1.8 Hz, 1H), 8.11 (d, J=8.7 Hz, 1H), 7.74 (d, J=8.3 Hz, 2H), 7.52 (t, J=7.9 Hz, 1H), 7.33 (ddd, J=16.4, 15.2, 6.4 Hz, 6H), 6.82 (d, J=16.4 Hz, 1H), 2.96 (dt, J=13.7, 6.9 Hz, 1H), 1.95 (s, 3H), 1.24 (d, J=6.9 Hz, 6H). MS (ESI): m/z 475.19 [M+H].sup.+. Mp 143-144° C.
Example 22
Synthesis of (E)-6-(4-propyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-40)
[0407] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 25%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.58 (s, 1H), 9.08 (s, 1H), 8.47 (d, J=1.6 Hz, 1H), 8.10 (dt, J=16.6, 5.3 Hz, 2H), 7.68 (d, J=8.2 Hz, 2H), 7.50-7.35 (m, 2H), 7.29 (t, J=11.5 Hz, 2H), 7.28-7.20 (m, 1H), 7.17 (d, J=7.6 Hz, 2H), 6.87 (d, J=16.5 Hz, 1H), 2.65-2.52 (m, 2H), 1.99 (s, 3H), 1.67-1.52 (m, 2H), 0.90 (t, J=7.3 Hz, 3H). MS (ESI): m/z 475.19 [M+H].sup.+. Mp 166-169° C.
Example 23
Synthesis of (E)-6-(4-isobutyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-41)
[0408] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 21%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.64 (s, 1H), 9.08 (s, 1H), 8.50 (d, J=1.5 Hz, 1H), 8.11 (dt, J=19.8, 5.3 Hz, 2H), 7.69 (d, J=8.2 Hz, 2H), 7.47 (t, J=8.0 Hz, 1H), 7.38 (d, J=16.5 Hz, 1H), 7.26 (dd, J=10.3, 8.3 Hz, 3H), 7.18 (d, J=6.9 Hz, 2H), 6.86 (d, J=16.4 Hz, 1H), 2.49-2.45 (m, 2H), 1.97 (s, 3H), 1.91-1.81 (m, 1H), 0.86 (t, J=7.6 Hz, 6H). MS (ESI): m/z 489.21 [M+H].sup.+. Mp 187-191° C.
Example 24
Synthesis of (E)-6-(4-butyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-42)
[0409] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 25%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.60 (s, 1H), 9.04 (s, 1H), 8.45 (d, J=1.5 Hz, 1H), 8.05 (dt, J=14.6, 5.2 Hz, 2H), 7.64 (d, J=8.2 Hz, 2H), 7.43 (t, J=7.9 Hz, 1H), 7.35 (d, J=16.5 Hz, 1H), 7.26 (d, J=8.2 Hz, 2H), 7.21 (d, J=7.9 Hz, 1H), 7.15 (d, J=11.7 Hz, 2H), 6.83 (d, J=16.5 Hz, 1H), 2.57 (t, J=7.6 Hz, 2H), 1.94 (s, 3H), 1.58-1.47 (m, 2H), 1.32-1.20 (m, 2H), 0.85 (t, J=7.3 Hz, 3H). MS (ESI): m/z 489.21 [M+H].sup.+. Mp 149-150° C.
Example 25
Synthesis of (E)-6-[3-(6-fluoro)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-43)
[0410] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 21%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 10.20 (s, 1H), 9.06 (s, 1H), 8.84-8.68 (m, 2H), 8.49 (td, J=8.3, 2.7 Hz, 1H), 8.22 (dd, J=8.8, 1.8 Hz, 1H), 8.11 (d, J=8.7 Hz, 1H), 7.52 (t, J=7.8 Hz, 1H), 7.43-7.24 (m, 4H), 7.18 (d, J=16.4 Hz, 1H), 6.76 (d, J=16.4 Hz, 1H), 1.90 (d, J=3.9 Hz, 3H). MS (ESI): m/z 452.13 [M+H].sup.+. Mp 133-134° C.
Example 26
Synthesis of (E)-6-(4-carbamoyl)phenyl-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-45)
[0411] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 26%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 11.32 (s, 1H), 9.10 (d, J=29.5 Hz, 2H), 8.43 (dd, J=8.8, 1.5 Hz, 1H), 8.25 (d, J=8.8 Hz, 1H), 8.08 (d, J=30.1 Hz, 5H), 7.73-7.49 (m, 4H), 7.46 (s, 1H), 6.98 (d, J=16.3 Hz, 1H), 6.65 (d, J=16.3 Hz, 1H), 1.81 (s, 3H). MS (ESI): m/z 476.15 [M+H].sup.+. Mp 211-214° C.
Example 27
Synthesis of (E)-6-[3-(5-cyano)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-3-46)
[0412] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 22%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 11.17 (s, 1H), 9.42 (d, J=2.3 Hz, 1H), 9.18 (t, J=7.6 Hz, 1H), 9.13-8.97 (m, 2H), 8.88 (t, J=2.1 Hz, 1H), 8.48 (dd, J=8.9, 1.7 Hz, 1H), 8.22 (d, J=8.8 Hz, 1H), 7.63 (ddd, J=24.6, 18.2, 7.3 Hz, 4H), 7.01-6.86 (m, 1H), 6.62 (d, J=16.3 Hz, 1H), 1.80 (s, 3H). MS (ESI): m/z 459.14 [M+H].sup.+. Mp 227-229° C.
Example 28
Synthesis of (E)-6-[3-(N-6-benzyloxyamido)pyridyl]-4-[3-(trifluoromethyl)phenyl]aminoquinoline-3-butenone (HTL-7-22)
[0413] The synthesis was carried out according to the synthetic method of HTL-2-34 to obtain a yellow solid, with a total yield of 24%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 10.48 (s, 1H), 9.56 (s, 1H), 9.07 (d, J=11.6 Hz, 1H), 8.67 (s, 1H), 8.51 (s, 1H), 8.12 (ddd, J=25.8, 18.6, 8.7 Hz, 3H), 7.94 (d, J=8.8 Hz, 1H), 7.46-7.15 (m, 10H), 6.83 (d, J=16.4 Hz, 1H), 5.18 (d, J=11.0 Hz, 2H), 1.98-1.92 (m, 3H). MS (ESI): m/z 583.19 [M+H].sup.+. Mp 187-189° C.
Example 29
Synthesis of 8-[3-(6-amino)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-4-32)
[0414] 80 mL microwave tube was taken, to which 8-bromo-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (4.08 g, 10 mmol), Pd(Ph.sub.3P).sub.4 (1.16 g, 1 mmol), K.sub.2CO.sub.3 (2.76 g, 20 mmol) and 6-aminopyridineboronic acid (1.66 g, 12 mmol) were added and dissolved in a solution of 1,4-dioxane. The microwave tube was sealed with cover, placed in a microwave reactor, the temperature was set to 100° C., the reaction was carried out for 40 minutes, the microwave tube was taken out and cooled to room temperature. The reaction solution was transferred to a separatory funnel, diluted with water, and extracted with ethyl acetate. The organic layer was washed with saturated NaCl, dried with anhydrous Na.sub.2SO.sub.4, filtered, the filtrate was dried by a rotary evaporator, and purified with column chromatography (n-hexane/ethyl acetate 5:1) to obtain 3.59 g of a yellow solid, with a yield of 85%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.05 (s, 1H), 8.37 (s, 1H), 8.22-8.07 (m, 3H), 8.03-7.88 (m, 3H), 7.40 (d, J=8.6 Hz, 1H), 6.95 (s, 1H), 6.45 (d, J=8.6 Hz, 1H), 6.25 (s, 2H). MS (ESI): m/z 423.10 [M+H].sup.+. Mp 146-147° C.
Example 30
Synthesis of 8-(4-carbamoyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-21)
[0415] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 42%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.43 (s, 1H), 8.41 (d, J=9.2 Hz, 2H), 8.28-8.21 (m, 2H), 8.19 (d, J=7.9 Hz, 1H), 8.06 (dd, J=18.3, 10.4 Hz, 2H), 7.92 (d, J=8.3 Hz, 2H), 7.44 (d, J=8.4 Hz, 3H), 7.19 (d, J=1.7 Hz, 1H). MS (ESI): m/z 450.10 [M+H].sup.+. Mp 294-296° C.
Example 31
Synthesis of 8-[3-(6-methyl)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c] quinolin-2(1H)-one (HTL-5-23)
[0416] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 37%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.35 (s, 1H), 8.80 (d, J=2.0 Hz, 1H), 8.40-8.33 (m, 2H), 8.26 (ddd, J=15.9, 8.6, 4.5 Hz, 3H), 8.11 (d, J=8.0 Hz, 1H), 8.02 (t, J=7.9 Hz, 1H), 7.91 (d, J=8.4 Hz, 1H), 7.27 (d, J=1.8 Hz, 1H), 2.75 (s, 3H). MS (ESI): m/z 422.10 [M+H].sup.+. Mp 299-300° C.
Example 32
Synthesis of 8-(3-furyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-25)
[0417] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 44%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.06 (s, 1H), 8.35 (s, 1H), 8.18 (t, J=8.5 Hz, 2H), 8.11-8.07 (m, 2H), 8.03 (t, J=7.9 Hz, 1H), 7.97 (dd, J=8.9, 1.9 Hz, 1H), 7.75 (t, J=1.7 Hz, 1H), 6.91 (d, J=1.8 Hz, 1H), 6.16 (dd, J=1.8, 0.7 Hz, 1H). MS (ESI): m/z 397.07 [M+H].sup.+. Mp 276-277° C.
Example 33
Synthesis of 8-(2-thienyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-26)
[0418] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 41%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.08 (s, 1H), 8.37 (s, 1H), 8.20 (t, J=8.4 Hz, 2H), 8.11 (dd, J=10.2, 5.4 Hz, 2H), 8.05 (dd, J=13.3, 5.3 Hz, 1H), 7.58 (dd, J=5.1, 0.8 Hz, 1H), 7.45-7.42 (m, 1H), 7.12 (dd, J=5.0, 3.7 Hz, 1H), 7.07 (d, J=1.6 Hz, 1H). MS (ESI): m/z 413.05 [M+H].sup.+. Mp 272-274° C.
Example 34
Synthesis of 8-(4-ethoxy)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-27)
[0419] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 39%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.08 (s, 1H), 8.38 (s, 1H), 8.16 (dd, J=25.3, 8.3 Hz, 2H), 8.00 (t, J=8.1 Hz, 2H), 7.29 (d, J=8.6 Hz, 2H), 7.04 (s, 1H), 6.92 (d, J=8.6 Hz, 2H), 4.05 (q, J=6.9 Hz, 2H), 1.33 (t, J=6.9 Hz, 3H). MS (ESI): m/z 451.12 [M+H].sup.+. Mp 254-254° C.
Example 35
Synthesis of 8-[3-(6-fluoro)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-28)
[0420] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 36%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.15 (s, 1H), 8.35 (s, 1H), 8.23-8.17 (m, 3H), 8.11 (d, J=7.9 Hz, 1H), 8.06-7.96 (m, 3H), 7.26 (dd, J=8.4, 2.7 Hz, 1H), 7.11 (d, J=1.8 Hz, 1H). MS (ESI): m/z 426.08 [M+H].sup.+. Mp 262-263° C.
Example 36
Synthesis of 8-(4-trifluoromethyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-29)
[0421] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 41%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.16 (s, 1H), 8.38 (s, 1H), 8.24-8.18 (m, 2H), 8.13 (d, J=8.0 Hz, 1H), 8.07 (dd, J=8.9, 2.0 Hz, 1H), 8.00 (t, J=7.9 Hz, 1H), 7.76 (d, J=8.3 Hz, 2H), 7.57 (d, J=8.2 Hz, 2H), 7.17 (d, J=1.9 Hz, 1H). MS (ESI): m/z 475.08 [M+H].sup.+. Mp 265-266° C.
Example 37
Synthesis of 8-[3-(5-methoxy)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-30)
[0422] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 42%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.15 (s, 1H), 8.37 (s, 1H), 8.27 (d, J=2.7 Hz, 1H), 8.21 (d, J=8.9 Hz, 2H), 8.16-8.09 (m, 3H), 8.00 (t, J=7.9 Hz, 1H), 7.31-7.28 (m, 1H), 7.14 (d, J=1.8 Hz, 1H), 3.83 (s, 3H). MS (ESI): m/z 438.10 [M+H].sup.+. Mp 257-258° C.
Example 38
Synthesis of 8-(3-quinolyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-32)
[0423] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 41%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.17 (s, 1H), 8.84 (d, J=2.4 Hz, 1H), 8.43-8.39 (m, 2H), 8.28-8.22 (m, 3H), 8.18 (d, J=8.0 Hz, 1H), 8.07-8.00 (m, 2H), 7.95 (d, J=7.3 Hz, 1H), 7.82-7.77 (m, 1H), 7.70-7.66 (m, 1H), 7.28 (s, 1H). MS (ESI): m/z 458.10 [M+H].sup.+. Mp 242-244° C.
Example 39
Synthesis of 8-(4-chloro)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-33)
[0424] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 45%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ (ppm): 9.14 (d, J=4.8 Hz, 1H), 8.39 (s, 1H), 8.22-8.16 (m, 2H), 8.13 (d, J=7.9 Hz, 1H), 8.04-7.98 (m, 2H), 7.47 (d, J=8.6 Hz, 2H), 7.38 (d, J=8.6 Hz, 2H), 7.09 (d, J=1.8 Hz, 1H). MS (ESI): m/z 441.05 [M+H].sup.+. Mp 283-286° C.
Example 40
Synthesis of 8-(4-propyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-34)
[0425] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 44%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.11 (s, 1H), 8.40 (s, 1H), 8.21-8.12 (m, 3H), 8.01 (dt, J=16.1, 5.4 Hz, 2H), 7.28 (d, J=8.2 Hz, 2H), 7.21 (d, J=8.2 Hz, 2H), 7.10 (d, J=1.7 Hz, 1H), 2.59-2.53 (m, 2H), 1.58 (dd, J=15.0, 7.4 Hz, 2H), 0.89 (t, J=7.3 Hz, 3H). MS (ESI): m/z 449.14 [M+H].sup.+. Mp 220-221° C.
Example 41
Synthesis of 8-(4-isopropyl)phenyl-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-35)
[0426] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 46%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.10 (s, 1H), 8.39 (s, 1H), 8.21-8.12 (m, 3H), 8.01 (dd, J=13.6, 4.8 Hz, 2H), 7.27 (q, J=8.4 Hz, 4H), 7.11 (d, J=1.8 Hz, 1H), 2.90 (dt, J=13.7, 6.9 Hz, 1H), 1.19 (d, J=6.9 Hz, 6H). MS (ESI): m/z 449.14 [M+H].sup.+. Mp 219-220° C.
Example 42
Synthesis of 8-[3-(5-cyano)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-5-36)
[0427] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 46%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.18 (s, 1H), 9.01 (d, J=1.7 Hz, 1H), 8.76 (d, J=2.1 Hz, 1H), 8.39-8.33 (m, 2H), 8.22 (dd, J=14.9, 8.4 Hz, 2H), 8.15-8.10 (m, 2H), 8.00 (t, J=7.9 Hz, 1H), 7.18 (d, J=1.5 Hz, 1H). MS (ESI): m/z 433.36 [M+H].sup.+. Mp 262-263° C.
Example 43
Synthesis of 8-[3-(6-valerylamino)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (WSX-1-24)
[0428] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 41%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.61 (s, 1H), 9.09 (s, 1H), 8.35 (s, 1H), 8.24-8.04 (m, 5H), 8.05-7.93 (m, 2H), 7.77 (dd, J=8.7, 2.3 Hz, 1H), 7.04 (d, J=1.6 Hz, 1H), 2.36 (t, J=7.4 Hz, 2H), 1.58-1.46 (m, 2H), 1.27 (dq, J=14.5, 7.3 Hz, 2H), 0.85 (t, J=7.3 Hz, 3H). MS (ESI): m/z 507.16 [M+H].sup.+. Mp 231-232° C.
Example 44
Synthesis of 8-[3-(6-benzyloxyamido)pyridyl]-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-7-23)
[0429] The synthesis was carried out according to the synthetic method of HTL-4-32 to obtain a yellow solid, with a total yield of 36%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.49 (s, 1H), 9.09 (s, 1H), 8.35 (s, 1H), 8.26-8.05 (m, 4H), 8.07-7.94 (m, 2H), 7.85 (d, J=8.7 Hz, 1H), 7.75 (dd, J=8.8, 2.4 Hz, 1H), 7.47-7.20 (m, 5H), 7.05 (d, J=1.7 Hz, 1H), 5.16 (s, 2H). MS (ESI): m/z 557.14 [M+H].sup.+. Mp 284-286° C.
Example 45
Synthesis of (E)-8-(2-carbamoyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-30)
[0430] 8-Bromo-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (4.08 g, 10 mmol) was dissolved in DMF solution, and Pd(OAc).sub.2 (0.45 g, 2 mmol), acrylamide (1.42 g, 20 mmol), tri(o-tolyl)phosphine (1.22 g, 4 mmol) and Et.sub.3N (10.12 g, 100 mmol) were added. Under the protection of N.sub.2, the mixed solution was heated to 100° C. and reacted for 2 hours. After the completion of the reaction monitored by TLC, the reaction solution was diluted with water and extracted with ethyl acetate. The organic layer was washed with saturated NaCl and dried with anhydrous MgSO.sub.4, and then filtrated, the filtrate was dried by a rotary evaporator, and purified by column chromatography (n-hexane/ethyl acetate 5:1) to obtain 3.16 g of a yellow solid, with a yield of 79%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.12 (s, 1H), 8.29 (s, 1H), 8.14 (dd, J=16.6, 8.3 Hz, 3H), 8.01 (t, J=7.9 Hz, 1H), 7.90 (dd, J=9.0, 1.8 Hz, 1H), 7.58 (s, 1H), 7.23 (s, 1H), 7.06 (d, J=15.8 Hz, 1H), 6.94 (d, J=1.6 Hz, 1H), 6.55 (d, J=15.8 Hz, 1H). MS (ESI): m/z 400.08 [M+H].sup.+. Mp 291-292° C.
Example 46
Synthesis of (E)-8-(3-cyano-propenyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-31)
[0431] The synthesis was carried out according to the synthetic method of HTL-6-30 to obtain a yellow solid, with a yield of 77%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.09 (d, J=4.3 Hz, 1H), 8.30 (s, 1H), 8.19-8.10 (m, 2H), 8.06 (d, J=8.9 Hz, 1H), 7.99 (t, J=7.9 Hz, 1H), 7.92 (dd, J=9.0, 1.9 Hz, 1H), 6.80 (s, 1H), 6.42 (d, J=15.9 Hz, 1H), 6.18 (dt, J=15.8, 5.9 Hz, 1H), 3.54 (dd, J=5.8, 1.4 Hz, 2H). MS (ESI): m/z 396.09 [M+H].sup.+. Mp 177-179° C.
Example 47
Synthesis of (E)-8-(2-cyano-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-32)
[0432] The synthesis was carried out according to the synthetic method of HTL-6-30 to obtain a yellow solid, with a yield of 77%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.15 (d, J=3.0 Hz, 1H), 8.24 (s, 1H), 8.12 (t, J=8.1 Hz, 3H), 8.04 (dd, J=9.1, 1.7 Hz, 1H), 8.00-7.94 (m, 1H), 7.51 (d, J=16.6 Hz, 1H), 7.09 (s, 1H), 6.42 (d, J=16.6 Hz, 1H). MS (ESI): m/z 382.07 [M+H].sup.+. Mp 235-237° C.
Example 48
Synthesis of (E)-8-(2-methoxycarbonyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-33)
[0433] The synthesis was carried out according to the synthetic method of HTL-6-30 to obtain a yellow solid, with a yield of 72%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.17 (d, J=11.8 Hz, 1H), 8.31 (s, 1H), 8.15 (d, J=8.0 Hz, 2H), 8.11 (s, 2H), 8.01 (t, J=7.9 Hz, 1H), 7.37 (d, J=16.0 Hz, 1H), 7.08 (s, 1H), 6.50 (d, J=16.0 Hz, 1H), 3.71 (s, 3H). MS (ESI): m/z 415.08 [M+H].sup.+. Mp 243-245° C.
Example 49
Synthesis of (E)-8-(3-ureido-propenyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-34)
[0434] The synthesis was carried out according to the synthetic method of HTL-6-30 to obtain a yellow solid, with a yield of 75%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.05 (s, 1H), 8.29 (s, 1H), 8.17-8.09 (m, 2H), 8.04-7.95 (m, 2H), 7.85 (d, J=9.1 Hz, 1H), 6.77 (s, 1H), 6.24-6.10 (m, 3H), 5.50 (s, 2H), 3.70 (t, J=5.4 Hz, 2H). MS (ESI): m/z 429.11 [M+H].sup.+. Mp 157-160° C.
Example 50
Synthesis of (E)-8-(2-tert-butoxycarbonyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (HTL-6-35)
[0435] The synthesis was carried out according to the synthetic method of HTL-6-30 to obtain a yellow solid, with a yield of 71%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.14 (s, 1H), 8.32 (s, 1H), 8.18-8.11 (m, 2H), 8.08 (d, J=1.0 Hz, 2H), 8.01 (t, J=7.9 Hz, 1H), 7.24 (d, J=15.9 Hz, 1H), 7.01 (s, 1H), 6.37 (d, J=15.9 Hz, 1H), 1.45 (s, 9H). MS (ESI): m/z 457.13 [M+H].sup.+. Mp 186-187° C.
Example 51
Synthesis of (E)-8-(4-ethoxycarbonyl-but-1-enyl)-1-[3-(trifluoromethyl)phenyl]oxazolo [5,4-c]quinolin-2(1H)-one (HTL-6-38)
[0436] The synthesis was carried out according to the synthetic method of HTL-6-30 to obtain a yellow solid, with a yield of 73%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.06 (s, 1H), 8.29 (s, 1H), 8.13 (t, J=8.2 Hz, 2H), 8.04-7.95 (m, 2H), 7.80 (dd, J=9.0, 1.8 Hz, 1H), 6.75 (s, 1H), 6.21 (d, J=15.9 Hz, 1H), 6.11 (d, J=15.8 Hz, 1H), 4.03 (dd, J=14.2, 7.1 Hz, 2H), 2.38 (s, 4H), 1.13 (t, J=7.1 Hz, 3H). MS (ESI): m/z 457.13 [M+H].sup.+. Mp 162-164° C.
Example 52
Synthesis of (E)-8-(2-ethoxycarbonyl-vinyl)-1-[3-(trifluoromethyl)phenyl]oxazolo[5,4-c]quinolin-2(1H)-one (WSX-1-13)
[0437] The synthesis was carried out according to the synthetic method of HTL-6-30 to obtain a yellow solid, with a yield of 77%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.15 (s, 1H), 8.30 (s, 1H), 8.18-8.06 (m, 4H), 8.00 (t, J=7.8 Hz, 1H), 7.34 (d, J=16.0 Hz, 1H), 7.07 (s, 1H), 6.49 (d, J=16.0 Hz, 1H), 4.16 (dd, J=13.7, 6.6 Hz, 2H), 1.23 (t, J=7.1 Hz, 6H). MS (ESI): m/z 429.10 [M+H].sup.+. Mp 221-224° C.
Example 53
Synthesis of 9-[3-(6-propionylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-45)
[0438] 80 mL microwave tube was taken, to which 9-bromo-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (4.18 g, 10 mmol), Pd(Ph.sub.3P).sub.4 (1.16 g, 1 mmol), K.sub.2CO.sub.3 (2.76 g, 20 mmol) and 6-aminopyridineboronic acid (1.66 g, 12 mmol) were added and dissolved in a solution of 1,4-dioxane. The microwave tube was sealed with cover, and placed in a microwave reactor, the temperature was set to 100° C., the reaction was carried out for 40 minutes, the microwave tube was taken out and cooled to room temperature. The reaction solution was transferred to a separatory funnel, diluted with water, and extracted with ethyl acetate. The organic layer was washed with saturated NaCl, dried with anhydrous Na.sub.2SO.sub.4, and filtered, the filtrate was dried by a rotary evaporator and purified with column chromatography (n-hexane/ethyl acetate 5:1) to obtain 3.59 g of a yellow solid, with a yield of 84%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.63 (s, 1H), 9.17 (s, 1H), 8.34 (d, J=9.5 Hz, 1H), 8.06 (ddd, J=16.3, 11.5, 5.5 Hz, 6H), 7.95-7.82 (m, 2H), 7.48 (dd, J=8.7, 2.5 Hz, 1H), 6.98 (dd, J=22.2, 5.5 Hz, 2H), 2.42 (q, J=7.5 Hz, 2H), 1.07 (t, J=7.5 Hz, 3H). MS (ESI): m/z 489.15 [M+H].sup.+. Mp 249-250° C.
Example 54
Synthesis of 9-(6-quinolyl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-47)
[0439] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 86%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.19 (s, 1H), 8.92 (dd, J=4.1, 1.5 Hz, 1H), 8.34 (dd, J=12.5, 9.1 Hz, 2H), 8.17 (s, 4H), 7.96 (dd, J=20.0, 8.3 Hz, 2H), 7.84 (d, J=7.9 Hz, 1H), 7.70 (d, J=1.6 Hz, 1H), 7.60 (dd, J=8.3, 4.2 Hz, 1H), 7.43 (dd, J=8.8, 2.0 Hz, 1H), 7.20 (s, 1H), 6.97 (d, J=9.4 Hz, 1H). MS (ESI): m/z 468.12 (M+H].sup.+. Mp 196-197° C.
Example 55
Synthesis of 9-[3-(2-fluoro)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo [h][1,6]naphthyridine (HTL-6-48)
[0440] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 82%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.21 (s, 1H), 8.35 (d, J=9.5 Hz, 1H), 8.28-8.20 (m, 1H), 8.15 (d, J=8.7 Hz, 1H), 8.07 (s, 1H), 7.97-7.87 (m, 2H), 7.87-7.76 (m, 2H), 7.52-7.44 (m, 1H), 7.42-7.34 (m, 1H), 6.96 (dd, J=9.5, 3.8 Hz, 1H), 6.87 (d, J=1.7 Hz, 1H). MS (ESI): m/z 436.10 [M+H].sup.+. Mp 158-161° C.
Example 56
Synthesis of 9-[3-(2-methyl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-49)
[0441] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 83%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.16 (s, 1H), 8.32 (d, J=9.5 Hz, 1H), 8.25 (d, J=2.0 Hz, 1H), 8.11 (t, J=6.6 Hz, 2H), 8.03 (dd, J=8.7, 1.8 Hz, 2H), 7.90-7.78 (m, 2H), 7.42-7.29 (m, 2H), 6.95 (dd, J=16.0, 5.6 Hz, 2H), 2.50 (s, 3H). MS (ESI): m/z 432.12 [M+H].sup.+. Mp 2046-207° C.
Example 57
Synthesis of 9-[(1H)-3-pyrazolyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-50)
[0442] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 85%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 12.93 (s, 1H), 9.08 (s, 1H), 8.29 (d, J=9.5 Hz, 1H), 8.16 (d, J=7.5 Hz, 1H), 8.08 (s, 1H), 8.02 (d, J=8.3 Hz, 2H), 7.88 (t, J=7.9 Hz, 1H), 7.78 (d, J=8.0 Hz, 1H), 7.69 (s, 1H), 7.08 (s, 1H), 6.90 (d, J=9.4 Hz, 1H), 5.72 (s, 1H). MS (ESI): m/z 407.10 [M+H].sup.+. Mp 283-286° C.
Example 58
Synthesis of 9-[3-(6-fluoro)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-01)
[0443] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 80%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.15 (s, 1H), 8.31 (d, J=9.5 Hz, 1H), 8.10 (d, J=8.7 Hz, 2H), 8.05-7.96 (m, 2H), 7.92-7.82 (m, 2H), 7.80 (d, J=8.0 Hz, 1H), 7.65 (td, J=8.2, 2.7 Hz, 1H), 7.19 (dd, J=8.5, 2.8 Hz, 1H), 6.91 (dd, J=14.4, 5.6 Hz, 2H). MS (ESI): m/z 436.10 [M+H].sup.+. Mp 264-266° C.
Example 59
Synthesis of 9-[3-(2-methoxy)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-02)
[0444] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 81%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.14 (s, 1H), 8.31 (d, J=9.5 Hz, 1H), 8.13 (dd, J=4.8, 2.1 Hz, 1H), 8.07-8.01 (m, 2H), 7.91-7.84 (m, 2H), 7.83-7.71 (m, 2H), 7.01-6.88 (m, 3H), 6.75 (d, J=1.7 Hz, 1H), 3.76 (s, 3H). MS (ESI): m/z 448.12 [M+H].sup.+. Mp 272-275° C.
Example 60
Synthesis of 9-[3-(6-methoxy)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-03)
[0445] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 86%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.12 (s, 1H), 8.30 (d, J=9.5 Hz, 1H), 8.12-8.04 (m, 2H), 8.03-7.92 (m, 3H), 7.85 (t, J=7.9 Hz, 1H), 7.76 (d, J=8.0 Hz, 1H), 7.24 (dd, J=8.7, 2.6 Hz, 1H), 6.90 (dd, J=10.5, 5.6 Hz, 2H), 6.76 (dd, J=8.6, 0.4 Hz, 1H), 3.84 (s, 3H). MS (ESI): m/z 448.12 [M+H].sup.+. Mp 211-216° C.
Example 61
Synthesis of 9-[5-(2-oxo)indolyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-04)
[0446] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 79%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.49 (s, 1H), 9.11 (s, 1H), 8.29 (d, J=9.5 Hz, 1H), 8.09 (dd, J=25.1, 8.2 Hz, 3H), 8.00-7.82 (m, 2H), 7.79 (d, J=7.9 Hz, 1H), 7.11 (d, J=7.7 Hz, 1H), 6.94 (dd, J=22.7, 5.5 Hz, 2H), 6.60-6.43 (m, 2H), 3.47 (s, 2H). MS (ESI): m/z 472.12 [M+H].sup.+. Mp 281-282° C.
Example 62
Synthesis of 9-[3-(6-butyrylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-05)
[0447] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 83%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.60 (s, 1H), 9.12 (s, 1H), 8.30 (d, J=9.5 Hz, 1H), 8.08 (d, J=8.8 Hz, 3H), 8.03-7.92 (m, 3H), 7.91-7.78 (m, 2H), 7.44 (dd, J=8.7, 2.5 Hz, 1H), 6.94 (dd, J=24.4, 5.5 Hz, 2H), 2.35 (t, J=7.3 Hz, 2H), 1.65-1.50 (m, 2H), 0.87 (t, J=7.4 Hz, 3H). MS (ESI): m/z 503.16 [M+H].sup.+. Mp 279-280° C.
Example 63
Synthesis of 9-[5-(2-methoxy)pyrimidyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-06)
[0448] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 77%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.15 (s, 1H), 8.36-8.25 (m, 3H), 8.10 (d, J=8.8 Hz, 2H), 8.05-7.95 (m, 2H), 7.87 (t, J=7.9 Hz, 1H), 7.79 (d, J=8.0 Hz, 1H), 6.93 (d, J=9.4 Hz, 1H), 6.87 (d, J=1.6 Hz, 1H), 3.91 (s, 3H). MS (ESI): m/z 449.11 [M+H].sup.+. Mp 262-264° C.
Example 64
Synthesis of 9-[3-(2-isobutyrylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-07)
[0449] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 76%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.60 (s, 1H), 9.13 (s, 1H), 8.30 (d, J=9.4 Hz, 1H), 8.13-8.04 (m, 3H), 8.04-7.96 (m, 3H), 7.91-7.79 (m, 2H), 7.44 (dd, J=8.7, 2.5 Hz, 1H), 6.99 (d, J=1.5 Hz, 1H), 6.96-6.88 (m, 1H), 2.75 (dt, J=13.6, 6.8 Hz, 1H), 1.06 (d, J=6.8 Hz, 6H). MS (ESI): m/z 503.16 [M+H].sup.+. Mp 213-215° C.
Example 65
Synthesis of 9-[3-(6-valerylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-08)
[0450] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 78%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.60 (s, 1H), 9.13 (s, 1H), 8.30 (d, J=9.5 Hz, 1H), 8.08 (dd, J=8.5, 6.3 Hz, 3H), 8.03-7.93 (m, 3H), 7.92-7.79 (m, 2H), 7.45 (dd, J=8.7, 2.5 Hz, 1H), 6.99 (d, J=1.6 Hz, 1H), 6.92 (d, J=9.4 Hz, 1H), 2.38 (t, J=7.4 Hz, 2H), 1.59-1.47 (m, 2H), 1.28 (dq, J=14.6, 7.3 Hz, 2H), 0.93-0.81 (m, 3H). MS (ESI): m/z 517.18 [M+H].sup.+. Mp 274-277° C.
Example 66
Synthesis of 9-[3-(6-phenylacetylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-10)
[0451] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 72%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.90 (s, 1H), 9.13 (s, 1H), 8.30 (d, J=9.5 Hz, 1H), 8.14-7.95 (m, 6H), 7.90-7.78 (m, 2H), 7.45 (dd, J=8.7, 2.5 Hz, 1H), 7.38-7.25 (m, 4H), 7.26-7.17 (m, 1H), 6.98 (d, J=1.7 Hz, 1H), 6.92 (d, J=9.4 Hz, 1H), 3.72 (s, 2H). MS (ESI): m/z 551.16 [M+H].sup.+. Mp 143-145° C.
Example 67
Synthesis of 9-{3-[6-(4-methoxy)phenylacetylamino]pyridyl}-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-11)
[0452] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 76%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.83 (s, 1H), 9.13 (s, 1H), 8.30 (d, J=9.5 Hz, 1H), 8.14-7.94 (m, 6H), 7.90-7.77 (m, 2H), 7.45 (dd, J=8.7, 2.5 Hz, 1H), 7.24 (d, J=8.7 Hz, 2H), 6.98 (d, J=1.7 Hz, 1H), 6.92 (d, J=9.4 Hz, 1H), 6.90-6.83 (m, 2H), 3.71 (d, J=10.3 Hz, 3H), 3.63 (s, 2H). MS (ESI): m/z 581.17 [M+H].sup.+. Mp 241-243° C.
Example 68
Synthesis of 9-[4-(3,5-dimethyl)isoxazolyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-12) The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 70%. .SUP.1.H-NMR (400 MHz, DMSO-D.SUB.6.) δ 9.16 (s, 1H), 8.31 (d, J=9.5 Hz, 1H), 8.10 (d, J=8.6 Hz, 1H), 7.96 (s, 1H), 7.87 (ddd, J=8.1, 4.4, 1.1 Hz, 1H), 7.84-7.77 (m, 2H), 7.66 (dd, J=8.6, 1.8 Hz, 1H), 6.91 (d, J=9.4 Hz, 1H), 6.61 (d, J=1.6 Hz, 1H), 2.03 (s, 3H), 1.89 (s, 3H). MS (ESI): m/z 436.12 [M+H].SUP.+.. Mp 121-122° C.
Example 69
Synthesis of 9-(1,4-dioxa-spiro[4.5]dec-7-en-8-yl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-13)
[0453] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 71%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.05 (s, 1H), 8.27 (d, J=9.5 Hz, 1H), 8.08 (s, 1H), 7.97 (d, J=7.9 Hz, 1H), 7.91 (d, J=8.8 Hz, 1H), 7.86-7.75 (m, 2H), 7.68 (d, J=8.1 Hz, 1H), 6.85 (dd, J=17.0, 5.6 Hz, 2H), 5.74 (t, J=3.9 Hz, 1H), 3.95-3.78 (m, 4H), 2.24 (s, 2H), 1.94-1.77 (m, 2H), 1.62 (t, J=6.4 Hz, 2H). MS (ESI): m/z 479.15 [M+H].sup.+. Mp 210-211° C.
Example 70
Synthesis of 9-[3-(6-benzyloxyamido)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-14)
[0454] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 68%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.45 (s, 1H), 9.16 (d, J=14.3 Hz, 1H), 8.31 (d, J=9.5 Hz, 1H), 8.20-7.91 (m, 5H), 7.93-7.71 (m, 3H), 7.53-7.14 (m, 6H), 7.05-6.85 (m, 2H), 5.17 (s, 2H). MS (ESI): m/z 567.16 [M+H].sup.+. Mp 150-153° C.
Example 71
Synthesis of 9-[3-(6-phenoxyamido)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-15)
[0455] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 73%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 10.94 (s, 1H), 9.15 (d, J=10.9 Hz, 1H), 8.31 (dt, J=11.1, 5.6 Hz, 1H), 8.18-8.06 (m, 3H), 8.04-7.98 (m, 1H), 7.85 (ddd, J=14.7, 13.6, 7.7 Hz, 3H), 7.43 (ddd, J=11.5, 9.8, 5.8 Hz, 3H), 7.34-7.16 (m, 4H), 6.99 (d, J=1.9 Hz, 1H), 6.92 (dd, J=9.4, 1.3 Hz, 1H). MS (ESI): m/z 553.14 [M+H].sup.+. Mp 150-152° C.
Example 72
Synthesis of 9-[3-(6-N,N-dimethyl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-16)
[0456] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 78%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.06 (s, 1H), 8.28 (d, J=9.5 Hz, 1H), 8.09 (s, 1H), 8.02 (dd, J=8.1, 3.5 Hz, 2H), 7.96 (d, J=2.4 Hz, 1H), 7.92 (dd, J=8.7, 1.8 Hz, 1H), 7.86 (t, J=7.9 Hz, 1H), 7.76 (d, J=8.0 Hz, 1H), 7.02 (dd, J=8.9, 2.6 Hz, 1H), 6.89 (t, J=5.6 Hz, 2H), 6.53 (d, J=8.9 Hz, 1H), 3.01 (s, 6H). MS (ESI): m/z 461.15 [M+H].sup.+.
[0457] Mp 220-222° C.
Example 73
Synthesis of 9-{3-[6-(4-methylpiperazin-1-yl)pyridyl]}-2-oxo-1-[3-(trifluoromethyl) phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-17)
[0458] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 75%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.08 (s, 1H), 8.29 (d, J=9.5 Hz, 1H), 8.08 (s, 1H), 8.05-7.99 (m, 2H), 7.98-7.90 (m, 2H), 7.86 (t, J=7.9 Hz, 1H), 7.78 (d, J=8.0 Hz, 1H), 7.07 (dd, J=8.9, 2.6 Hz, 1H), 6.90 (dd, J=5.6, 3.8 Hz, 2H), 6.74 (d, J=8.9 Hz, 1H), 3.50 (dd, J=16.8, 12.0 Hz, 4H), 2.41-2.28 (m, 4H), 2.19 (s, 3H). MS (ESI): m/z 516.19 [M+H].sup.+. Mp 161-162° C.
Example 74
Synthesis of 9-[3-(6-morpholin-4-yl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-18)
[0459] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 72%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.08 (s, 1H), 8.28 (d, J=9.5 Hz, 1H), 8.08 (s, 1H), 8.02 (dd, J=8.2, 5.0 Hz, 2H), 7.99-7.90 (m, 2H), 7.85 (t, J=7.9 Hz, 1H), 7.76 (d, J=8.0 Hz, 1H), 7.10 (dd, J=8.9, 2.6 Hz, 1H), 6.90 (dd, J=5.6, 3.9 Hz, 2H), 6.75 (d, J=8.9 Hz, 1H), 3.77-3.60 (m, 4H), 3.52-3.38 (m, 4H). MS (ESI): m/z 503.16 [M+H].sup.+. Mp 286-287° C.
Example 75
Synthesis of 9-[3-(6-amino-5-methoxy)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-19)
[0460] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 81%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.07 (s, 1H), 8.28 (d, J=9.5 Hz, 1H), 8.06-7.81 (m, 6H), 7.05 (d, J=2.0 Hz, 1H), 6.97 (d, J=1.8 Hz, 1H), 6.88 (dd, J=8.5, 5.7 Hz, 2H), 6.01 (s, 2H), 3.78 (s, 3H). MS (ESI): m/z 463.13 [M+H].sup.+. Mp 233-234° C.
Example 76
Synthesis of 9-[3-(6-pyrrolidinyl)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-20)
[0461] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 77%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.05 (s, 1H), 8.27 (d, J=9.5 Hz, 1H), 8.09 (s, 1H), 8.05-7.94 (m, 3H), 7.93-7.81 (m, 2H), 7.75 (d, J=8.0 Hz, 1H), 6.98 (dd, J=8.8, 2.6 Hz, 1H), 6.87 (dd, J=10.8, 5.6 Hz, 2H), 6.32 (d, J=8.8 Hz, 1H), 3.35 (t, J=6.5 Hz, 4H), 1.91 (t, J=6.6 Hz, 4H). MS (ESI): m/z 487.17 [M+H].sup.+. Mp 252-254° C.
Example 77
Synthesis of 9-[3-(6-tert-butoxycarbonylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl) phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-21)
[0462] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 73%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.97 (s, 1H), 9.12 (s, 1H), 8.30 (d, J=9.5 Hz, 1H), 8.08 (dd, J=11.2, 2.3 Hz, 3H), 8.00 (dt, J=8.8, 4.4 Hz, 2H), 7.87 (t, J=7.8 Hz, 1H), 7.78 (dd, J=17.8, 8.4 Hz, 2H), 7.28 (dd, J=8.8, 2.5 Hz, 1H), 6.93 (dd, J=14.7, 5.6 Hz, 2H), 1.45 (s, 9H). MS (ESI): m/z 533.17 [M+H].sup.+. Mp 234-236° C.
Example 78
Synthesis of 9-[3-(6-allylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-28)
[0463] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 70%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.07 (s, 1H), 8.29 (d, J=9.7 Hz, 1H), 8.19-7.45 (m, 7H), 7.13-6.77 (m, 3H), 6.44 (d, J=8.2 Hz, 1H), 5.88 (s, 1H), 5.32-4.85 (m, 2H), 4.14 (d, J=35.1 Hz, 1H), 3.89 (s, 2H). MS (ESI): m/z 473.15 [M+H].sup.+. Mp 142-146° C.
Example 79
Synthesis of 9-[3-(6-propargylamino)pyridyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-29)
[0464] The synthesis was carried out according to the synthetic method of HTL-6-45 to obtain a yellow solid, with a yield of 68%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.14 (s, 1H), 8.31 (t, J=7.5 Hz, 1H), 8.16-8.07 (m, 2H), 8.00 (dd, J=14.2, 5.4 Hz, 1H), 7.94-7.86 (m, 1H), 7.80 (d, J=9.3 Hz, 1H), 7.71-7.59 (m, 4H), 7.42-7.32 (m, 1H), 6.98-6.87 (m, 2H), 4.19 (t, J=6.5 Hz, 2H), 4.10 (dd, J=5.5, 3.6 Hz, 1H). MS (ESI): m/z 471.14 [M+H].sup.+. Mp 139-141° C.
Example 80
Synthesis of 9-(3-aminophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-11)
[0465] 9-Bromo-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (4.18 g, 10 mmol) was dissolved in a solution of 1,4-dioxane, then Pd(dba).sub.3 (0.23 g, 0.25 mmol), CsCO.sub.3 (4.89 g, 15 mmol), 1,3-phenylenediamine (1.62 g, 15 mmol) and 4,5-bis(diphenylphosphine)-9,9-dimethylxanthene (0.14 g, 0.25 mmol) were added. Under the protection of N.sub.2, the resulting mixture was heated to 100° C. and reacted for 2 hours. When the reaction was completed, the reaction solution was diluted with water and extracted with ethyl acetate. The organic layer was washed with saturated NaCl solution, dried with anhydrous MgSO.sub.4, and filtered, the filtrate was dried by a rotary evaporator to remove solvent, and purified by column chromatography (n-hexane/ethyl acetate 5:1) to obtain 3.66 g of a yellow solid, with a yield of 82%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.86 (s, 1H), 8.24 (d, J=9.5 Hz, 1H), 7.96 (s, 1H), 7.88 (d, J=9.0 Hz, 1H), 7.80 (d, J=8.1 Hz, 1H), 7.64 (dd, J=14.4, 6.4 Hz, 2H), 7.55 (d, J=7.9 Hz, 1H), 7.31 (dd, J=9.0, 2.4 Hz, 1H), 6.91-6.82 (m, 2H), 6.44 (d, J=2.3 Hz, 1H), 6.22 (dd, J=7.9, 1.3 Hz, 1H), 6.08 (t, J=2.0 Hz, 1H), 5.89-5.77 (m, 1H), 5.16 (s, 2H). MS (ESI): m/z 447.14 [M+H].sup.+. Mp 217-218° C.
Example 81
Synthesis of 9-[2-(6-aminopyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-12)
[0466] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 81%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.90 (s, 1H), 8.49 (s, 1H), 8.24 (d, J=9.5 Hz, 1H), 7.94 (d, J=8.0 Hz, 1H), 7.87 (d, J=9.0 Hz, 1H), 7.66 (dt, J=13.4, 7.8 Hz, 3H), 7.51 (dd, J=9.0, 2.3 Hz, 1H), 7.21-7.10 (m, 2H), 6.85 (d, J=9.4 Hz, 1H), 5.91 (d, J=7.9 Hz, 1H), 5.70 (t, J=14.5 Hz, 3H). MS (ESI): m/z 448.13 [M+H].sup.+. Mp 259-261° C.
Example 82
Synthesis of 9-[4-(trifluoromethyl)phenyl]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-16)
[0467] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 83%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.98 (s, 1H), 8.71 (s, 1H), 8.26 (d, J=9.5 Hz, 1H), 7.99 (d, J=8.9 Hz, 1H), 7.86 (s, 1H), 7.69-7.59 (m, 2H), 7.52 (d, J=7.4 Hz, 1H), 7.47 (d, J=8.6 Hz, 2H), 7.39 (dd, J=9.0, 2.4 Hz, 1H), 6.87 (d, J=9.4 Hz, 1H), 6.78 (d, J=8.4 Hz, 2H), 6.71 (d, J=2.3 Hz, 1H). MS (ESI): m/z 450.11 [M+H].sup.+. Mp 151-153° C.
Example 83
Synthesis of 9-(3,4-dimethylphenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-17)
[0468] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 85%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.86 (s, 1H), 8.21 (d, J=9.5 Hz, 1H), 7.98 (s, 1H), 7.87 (d, J=9.0 Hz, 1H), 7.64 (d, J=7.7 Hz, 1H), 7.57 (s, 1H), 7.49-7.37 (m, 2H), 7.29 (dd, J=9.0, 2.4 Hz, 1H), 6.97 (d, J=8.0 Hz, 1H), 6.82 (d, J=9.4 Hz, 1H), 6.58 (d, J=2.0 Hz, 1H), 6.37 (dd, J=8.0, 2.2 Hz, 1H), 6.27 (d, J=2.3 Hz, 1H), 2.22 (d, J=20.6 Hz, 6H). MS (ESI): m/z 460.16 [M+H].sup.+. Mp 140-141° C.
Example 84
Synthesis of 9-(4-tert-butylphenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-18)
[0469] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 80%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.87 (s, 1H), 8.22 (d, J=9.5 Hz, 1H), 8.14 (s, 1H), 7.88 (d, J=9.0 Hz, 1H), 7.74-7.65 (m, 2H), 7.58 (t, J=7.8 Hz, 1H), 7.49 (d, J=7.9 Hz, 1H), 7.31 (dd, J=9.0, 2.4 Hz, 1H), 7.24-7.17 (m, 2H), 6.83 (d, J=9.4 Hz, 1H), 6.59 (d, J=8.6 Hz, 2H), 6.49 (d, J=2.3 Hz, 1H), 1.32 (d, J=7.5 Hz, 9H). MS (ESI): m/z 488.19 [M+H].sup.+. Mp 141-142° C.
Example 85
Synthesis of 9-[3-(6-methylpyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-19)
[0470] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 79%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.92 (s, 1H), 8.50 (s, 1H), 8.25 (d, J=9.5 Hz, 1H), 7.99-7.91 (m, 2H), 7.75-7.65 (m, 2H), 7.59 (t, J=7.8 Hz, 1H), 7.52 (d, J=7.8 Hz, 1H), 7.35 (dd, J=9.0, 2.4 Hz, 1H), 7.24 (d, J=8.4 Hz, 1H), 7.13 (dd, J=8.4, 2.5 Hz, 1H), 6.85 (d, J=9.4 Hz, 1H), 6.37 (d, J=2.3 Hz, 1H), 2.53 (s, 3H). MS (ESI): m/z 447.14 [M+H].sup.+. Mp 234-236° C.
Example 86
Synthesis of 9-[3-(6-fluoropyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-20)
[0471] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 83%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.90 (s, 1H), 8.34 (s, 1H), 8.23 (d, J=9.5 Hz, 1H), 7.93 (d, J=9.0 Hz, 1H), 7.71 (s, 1H), 7.59 (ddd, J=36.5, 19.9, 7.9 Hz, 4H), 7.35-7.24 (m, 2H), 7.04 (dd, J=8.7, 3.2 Hz, 1H), 6.84 (d, J=9.4 Hz, 1H), 6.32 (d, J=2.3 Hz, 1H). MS (ESI): m/z 451.11 [M+H].sup.+. Mp 232-233° C.
Example 87
Synthesis of 9-[3-(6-chloropyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-21)
[0472] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 82%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.95 (s, 1H), 8.51 (s, 1H), 8.25 (d, J=9.5 Hz, 1H), 7.96 (d, J=8.9 Hz, 1H), 7.81 (d, J=2.7 Hz, 1H), 7.77 (s, 1H), 7.69 (d, J=8.0 Hz, 1H), 7.62 (t, J=7.8 Hz, 1H), 7.55 (d, J=7.7 Hz, 1H), 7.35 (dd, J=9.0, 2.4 Hz, 1H), 7.30 (d, J=8.5 Hz, 1H), 7.11 (dd, J=8.6, 3.0 Hz, 1H), 6.86 (d, J=9.4 Hz, 1H), 6.46 (d, J=2.3 Hz, 1H). MS (ESI): m/z 467.08 [M+H].sup.+. Mp 137-138° C.
Example 88
Synthesis of 9-(4-cyanophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-22)
[0473] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 84%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.00 (s, 1H), 8.88 (s, 1H), 8.27 (d, J=9.5 Hz, 1H), 8.00 (d, J=8.9 Hz, 1H), 7.86 (s, 1H), 7.68 (dd, J=15.2, 4.4 Hz, 3H), 7.54 (d, J=8.6 Hz, 2H), 7.40 (dd, J=8.9, 2.2 Hz, 1H), 6.88 (d, J=9.4 Hz, 1H), 6.71 (dd, J=5.4, 3.1 Hz, 3H). MS (ESI): m/z 457.12 [M+H].sup.+. Mp 259-260° C.
Example 89
Synthesis of 9-(4-fluorophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-23)
[0474] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 80%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.90 (s, 1H), 8.26-8.15 (m, 2H), 7.91 (d, J=9.0 Hz, 1H), 7.70-7.62 (m, 2H), 7.57 (dd, J=18.8, 7.7 Hz, 2H), 7.31 (dd, J=9.0, 2.3 Hz, 1H), 7.05 (t, J=8.8 Hz, 2H), 6.84 (d, J=9.4 Hz, 1H), 6.75-6.65 (m, 2H), 6.37 (d, J=2.3 Hz, 1H). MS (ESI): m/z 450.12 [M+H].sup.+. Mp 176-179° C.
Example 90
Synthesis of 9-[(4-fluoro-3-methyl)phenyl]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-24)
[0475] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 78%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.84 (s, 1H), 8.21 (d, J=9.4 Hz, 1H), 8.00 (s, 1H), 7.87 (d, J=9.0 Hz, 1H), 7.67-7.57 (m, 2H), 7.54-7.42 (m, 2H), 7.26 (dd, J=9.0, 2.4 Hz, 1H), 7.02-6.93 (m, 1H), 6.81 (d, J=9.4 Hz, 1H), 6.69 (dd, J=6.8, 2.5 Hz, 1H), 6.52-6.44 (m, 1H), 6.25 (d, J=2.3 Hz, 1H), 2.21 (d, J=1.6 Hz, 3H). MS (ESI): m/z 464.13 [M+H].sup.+. Mp 210-211° C.
Example 91
Synthesis of 9-(4-chlorophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-25)
[0476] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 83%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.89 (s, 1H), 8.31 (s, 1H), 8.23 (d, J=9.4 Hz, 1H), 7.92 (d, J=8.9 Hz, 1H), 7.73 (s, 1H), 7.67 (d, J=7.9 Hz, 1H), 7.61 (t, J=7.8 Hz, 1H), 7.55 (d, J=7.7 Hz, 1H), 7.31 (dd, J=9.0, 2.4 Hz, 1H), 7.26-7.14 (m, 2H), 6.83 (d, J=9.4 Hz, 1H), 6.72-6.62 (m, 2H), 6.50 (d, J=2.3 Hz, 1H). MS (ESI): m/z 466.09 [M+H].sup.+. Mp 212-213° C.
Example 92
Synthesis of 9-(4-methoxyphenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-26)
[0477] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 84%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.83 (s, 1H), 8.19 (d, J=9.4 Hz, 1H), 7.93 (s, 1H), 7.85 (d, J=9.0 Hz, 1H), 7.68-7.55 (m, 2H), 7.55-7.46 (m, 2H), 7.26 (dd, J=9.0, 2.4 Hz, 1H), 6.82 (ddd, J=9.3, 6.1, 3.3 Hz, 3H), 6.68-6.57 (m, 2H), 6.21 (d, J=2.3 Hz, 1H), 3.81 (s, 3H). MS (ESI): m/z 462.14 [M+H].sup.+. Mp 138-140° C.
Example 93
Synthesis of 9-(4-acetylaminophenyl)imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-27)
[0478] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid with a yield of 79%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.85 (s, 1H), 8.83 (s, 1H), 8.20 (d, J=9.4 Hz, 1H), 8.02 (s, 1H), 7.86 (d, J=9.0 Hz, 1H), 7.70 (d, J=7.8 Hz, 1H), 7.62-7.45 (m, 3H), 7.42 (d, J=8.8 Hz, 2H), 7.27 (dd, J=9.0, 2.4 Hz, 1H), 6.81 (d, J=9.4 Hz, 1H), 6.61 (d, J=8.8 Hz, 2H), 6.30 (d, J=2.3 Hz, 1H), 2.09 (s, 3H). MS (ESI): m/z 489.15 [M+H].sup.+. Mp 175-177° C.
Example 94
Synthesis of 9-[2-(6-aminopyrazinyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-28)
[0479] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 72%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.55 (s, 1H), 9.30 (s, 1H), 8.36 (d, J=9.6 Hz, 1H), 8.11 (d, J=9.0 Hz, 1H), 8.03 (d, J=7.7 Hz, 1H), 7.78 (s, 1H), 7.69 (dt, J=16.2, 7.7 Hz, 2H), 7.40 (d, J=2.1 Hz, 1H), 7.32 (s, 1H), 7.19 (s, 1H), 7.02 (d, J=9.5 Hz, 1H), 6.69 (s, 2H). MS (ESI): m/z 449.13 [M+H].sup.+. Mp 192-194° C.
Example 95
Synthesis of 9-[3-(5-aminopyridyl)]imino-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-6-29)
[0480] The synthesis was carried out according to the synthetic method of HTL-6-11 to obtain a yellow solid, with a yield of 82%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 8.89 (s, 1H), 8.25 (d, J=9.5 Hz, 1H), 8.14 (s, 1H), 7.90 (d, J=8.9 Hz, 1H), 7.79 (d, J=7.7 Hz, 1H), 7.62 (dd, J=8.8, 6.9 Hz, 2H), 7.56 (d, J=2.3 Hz, 1H), 7.48 (d, J=7.9 Hz, 1H), 7.30 (dd, J=9.0, 2.4 Hz, 1H), 7.22 (d, J=2.2 Hz, 1H), 6.85 (d, J=9.4 Hz, 1H), 6.41 (t, J=2.3 Hz, 1H), 6.33 (d, J=2.3 Hz, 1H), 5.43 (s, 2H). MS (ESI): m/z 448.13 [M+H].sup.+. Mp 156-158° C.
Example 96
Synthesis of (E)-9-(2-carbamoyl-vinyl)-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-24)
[0481] 9-Bromo-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (4.18 g, 10 mmol) was dissolved in a solution of DMF, then Pd(OAc).sub.2 (0.45 g, 2 mmol), acrylamide (1.42 g, 20 mmol), tri(o-tolyl)phosphine (1.22 g, 4 mmol) and Et.sub.3N (10.12 g, 100 mmol) were added. Under the protection of N.sub.2, the mixed solution was heated to 100° C. and reacted for 2 hours. After the completion of the reaction monitored by TLC, the reaction solution was diluted with water and extracted with ethyl acetate. The organic layer was washed with saturated brine, dried with MgSO.sub.4, and filtered, and the filtrate was dried by a rotary evaporator, and purified by column chromatography (n-hexane/ethyl acetate 5:1) to obtain 3.40 g of a yellow solid, with a yield of 83%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.12 (s, 1H), 8.29 (d, J=9.5 Hz, 1H), 7.99 (ddd, J=25.9, 19.1, 7.9 Hz, 4H), 7.87-7.80 (m, 2H), 7.40 (s, 1H), 7.16 (s, 1H), 6.92 (d, J=9.4 Hz, 1H), 6.79 (d, J=15.7 Hz, 1H), 6.55 (s, 1H), 6.25 (d, J=15.7 Hz, 1H). MS (ESI): m/z 410.10 [M+H].sup.+. Mp 307-308° C.
Example 97
Synthesis of (E)-9-[3-ureido-propenyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-25)
[0482] The synthesis was carried out according to the synthetic method of HTL-7-24 to obtain a yellow solid, with a yield of 80%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.06 (s, 1H), 8.26 (dd, J=10.2, 6.3 Hz, 2H), 8.07 (s, 1H), 7.96-7.69 (m, 5H), 6.98 (d, J=11.1 Hz, 1H), 6.88 (d, J=9.4 Hz, 1H), 6.73 (d, J=1.7 Hz, 1H), 6.08 (s, 1H), 1.88 (s, 1H), 1.30 (s, 2H), 1.00 (d, J=6.1 Hz, 1H). MS (ESI): m/z 439.13 [M+H].sup.+. Mp 150-151° C.
Example 98
Synthesis of (E)-9-[2-ethoxycarbonyl-vinyl]-2-oxo-1-[3-(trifluoromethyl)phenyl]-1,2-dihydrobenzo[h][1,6]naphthyridine (HTL-7-26)
[0483] The synthesis was carried out according to the synthetic method of HTL-7-24 to obtain a yellow solid, with a yield of 81%. .sup.1H-NMR (400 MHz, DMSO-D.sub.6) δ 9.14 (s, 1H), 8.30 (d, J=9.5 Hz, 1H), 8.09 (s, 1H), 8.05-7.93 (m, 3H), 7.89 (t, J=7.9 Hz, 1H), 7.77 (d, J=8.1 Hz, 1H), 7.09 (d, J=16.0 Hz, 1H), 6.93 (d, J=9.4 Hz, 1H), 6.68 (s, 1H), 6.04 (d, J=16.0 Hz, 1H), 4.21-4.02 (m, 2H), 1.24 (t, J=7.1 Hz, 3H). MS (ESI): m/z 439.12 [M+H].sup.+. Mp 210-212° C.
[0484] Example 99: Experiment of In Vitro Anti-EV71 Activity of the Compound of the Present application
[0485] Experimental Method:
[0486] According to the mass and molecular weight of the compound to be tested, the compound to be tested was dissolved to 100 mM (mmol/L) with DMSO.
[0487] Detection of Antiviral Activity:
[0488] {circle around (1)} First, the compound to be tested was diluted to 800 μM with cell maintenance solution (DMEM+2% FBS, Gibco, catalog numbers: 11995-065, 1600-044), and three-fold gradient dilution was performed to prepare 10 concentrations in total. The diluted compound was added to a 96-well plate with white wall and transparent bottom, 50 μl per well. To both the cell control group and the virus control group, an equal volume of the cell maintenance solution was added.
[0489] {circle around (2)} EV71 virus (purchased from ATCC) was taken out from the −80° C. refrigerator and equilibrated to room temperature.
[0490] {circle around (3)} The virus seed was diluted with a virus growth solution (DMEM+2% FBS, Gibco, catalog numbers: 11995-065, 1600-044) to 100TCID.sub.50, the diluted virus seed was added to the 96-well plate of {circle around (1)}, with 50p per well. The same volume of the virus growth solution was added to the cell control group.
[0491] {circle around (4)} RD cells (purchased from ATCC) were inoculated at a concentration of 1*10.sup.5/mL into the 96-well plate of {circle around (1)}, with 100 μL per well, to reach a final volume of 200 μL per well. The final concentration of drug was 0.25 times the initial concentration.
[0492] {circle around (5)} RD cells were cultured at 37° C. for 4 days for testing.
[0493] {circle around (6)} The Buffer of CellTiter-Glo® luminescent cell viability assay reagent (Promega) was mixed with the substrate in the dark to prepare a working solution.
[0494] {circle around (7)} The culture medium in the plate was discarded, the plate was dried by patting, 100 μL of the detection reagent was added to each well, and the 96-well plate was shaken on an orbital shaker for 4 minutes to induce cell lysis. After keeping in the dark and subjecting to signal stabilization for 15 minutes, the chemiluminescence unit was detected by using MD5 microplate reader (Molecular Devices), and the plate reading program was performed according to the CellTiter-Glo preset program.
[0495] The inhibition rate-concentration curve was fitted to S-curve by using origin8.0 software to calculate the IC.sub.50 value of the compound to be tested.
[0496] Detection of Cytotoxicity:
[0497] {circle around (1)} The compound to be tested was diluted to a concentration of 400 μM with the cell maintenance solution, and 3-fold gradient dilution was carried out to obtain 10 concentrations in total.
[0498] {circle around (2)} The diluted compound was added to a 96-well plate with white wall and transparent bottom, with 100 μL per well. An equal volume of the cell maintenance solution was added to the cell control group.
[0499] {circle around (3)} RD cells were inoculated into the above 96-well plate at a concentration of 1*10.sup.5/mL, with 100 μL per well, to reach a final volume of 200 μL per well, and the final concentration of drug was 0.5 times the pretreatment concentration.
[0500] {circle around (4)} The RD cells were cultured at 37° C. for 4 days for testing.
[0501] {circle around (5)} The Buffer of CellTiter-Glo® luminescent cell viability assay reagent was mixed with the substrate in the dark to prepare a working solution.
[0502] {circle around (6)} The culture medium in the plate was discarded, the plate was dried by patting, 100 μL of the detection reagent was added to each well, and the 96-well plate was shaken on an orbital shaker for 4 minutes to induce cell lysis. After keeping in the dark and subjecting to signal stabilization for 15 minutes, the chemiluminescence unit was detected, and the plate reading program was performed according to the CellTiter-Glo preset program.
[0503] The formula for calculating the inhibition rates of drug at different dilution degrees was as follows: inhibition rate (%)=(average value of cell control group−value of experimental group)/(average value of cell control group−minimum value of experimental group)*100
[0504] Data Analysis:
[0505] The inhibition rate-concentration curve was fitted to S-curve by using origin8.0 software, and the IC.sub.50 value of the compound to be tested was calculated. The same method was used to calculate the TD.sub.50 value, and the selection index SI=TD.sub.50/IC.sub.50 was calculated according to the half maximal inhibitory concentration IC.sub.50 and the median toxic dose TD.sub.50.
[0506] The results of inhibitory activity of the compounds of Formula I, Formula II and Formula III against Enterovirus 71 (EV71) H strain were shown in the following tables:
TABLE-US-00002 TABLE 1 Inhibitory activity of compounds of Formula I against EV71 Code IC.sub.50(μM) TD.sub.50(μM) SI HTL-2-34 1.78 ± 0.03 4.99 ± 1.55 2.80 HTL-2-35 3.70 ± 0.01 5.14 ± 1.92 1.39 HTL-2-38 2.85 ± 1.20 8.71 ± 1.53 3.06 HTL-2-42 >200 >200 — HTL-3-04 >200 >200 — HTL-3-07 >200 >200 — HTL-3-11 >200 >200 — HTL-3-12 >200 >200 — HTL-3-15 >200 >200 — HTL-3-18 >200 >200 — HTL-3-16 >200 0.83 ± 0.79 — HTL-3-22 >200 1.55 ± 1.03 — HTL-3-24 >200 1.35 ± 1.16 — HTL-3-25 >200 0.68 ± 0.21 — HTL-3-26 0.85 ± 0.03 1.43 ± 1.62 1.68 HTL-3-32 >200 1.52 ± 0.87 — HTL-3-33 >200 0.72 ± 0.66 — HTL-3-34 >200 3.02 ± 0.91 — HTL-3-36 >200 4.65 ± 3.50 — HTL-3-37 >200 1.88 ± 0.46 — HTL-3-39 >200 1.75 ± 0.73 — HTL-3-40 >200 0.83 ± 0.03 — HTL-3-41 >200 0.99 ± 0.24 — HTL-3-42 >200 0.96 ± 0.15 — HTL-3-43 >200 2.23 ± 0.08 — HTL-3-45 >200 2.42 ± 0.06 — HTL-3-46 >200 3.59 ± 2.46 — HTL-7-22 >200 9.4 ± 8.0 —
[0507] The in vitro anti-EV71 activity test results of 28 compounds represented by Formula I showed that, except for HTL-2-34, HTL-2-35, HTL-2-38 and HTL-3-26 which had moderate anti-EV71 activity, other 3-butenone quinoline compounds didn't show inhibitory activity against EV71.
TABLE-US-00003 TABLE 2 Inhibitory activity of compounds of Formula II against EV71 Code IC.sub.50(μM) TD.sub.50(μM) SI HTL-4-32 5.57 ± 1.53 14.5 ± 6.4 2.60 HTL-5-21 2.00 ± 0.30 6.95 ± 1.23 3.48 HTL-5-23 4.78 ± 1.73 16.88 ± 5.46 3.53 HTL-5-25 >200 >200 — HTL-5-26 >200 >200 — HTL-5-27 >200 >200 — HTL-5-28 >200 >200 — HTL-5-29 >200 >200 — HTL-5-30 >200 68.52 ± 0.42 — HTL-5-32 >200 68.88 ± 0.13 — HTL-5-33 >200 >200 — HTL-5-34 >200 >200 — HTL-5-35 >200 69.36 ± 0.28 — HTL-5-36 >200 13.41 ± 0.18 — HTL-6-30 3.70 ± 0.21 7 ± 0.09 1.89 HTL-6-31 >200 9.20 ± 2.34 — HTL-6-32 >200 >200 HTL-6-33 >200 71.99 ± 0.78 — HTL-6-34 >200 66.64 ± 1.17 1.36 HTL-6-35 >200 76.02 ± 11.52 — HTL-6-38 >200 22.74 ± 0.01 — WSX-1-13 100 ± 2.34 73.88 ± 3.52 0.74 WSX-1-24 >200 4.27 ± 2.83 — HTL-7-23 >200 >200 —
[0508] The in vitro anti-EV71 activity test results of 24 compounds represented by Formula II showed that HTL-4-32, HTL-5-21, HTL-5-23 and HTL-6-30 showed moderate inhibitory activity against EV71, WSX-1-13 had weak inhibitory activity against EV71, and other compounds did not show inhibitory activity.
TABLE-US-00004 TABLE 3 Inhibitory activity of compounds of Formula III agaist EV71 Code IC.sub.50(μM) TD.sub.50(μM) SI HTL-6-11 0.89 7.58 ± 1.24 8.52 HTL-6-12 1.23 ± 0.22 5.19 ± 0.56 4.22 HTL-6-16 >200 7.06 ± 0.23 — HTL-6-17 >200 6.81 ± 1.03 — HTL-6-18 >200 7.04 ± 0.68 — HTL-6-19 >200 22.62 ± 2.15 — HTL-6-20 >200 59.31 ± 3.98 — HTL-6-21 >200 20.75 ± 2.11 — HTL-6-22 >200 6.95 ± 0.86 — HTL-6-23 >200 7.22 ± 0.47 — HTL-6-24 >200 7.2 ± 0.33 — HTL-6-25 >200 4.56 ± 0.26 — HTL-6-26 >200 >200 — HTL-6-27 7.14 ± 3.21 10.65 ± 0.57 1.49 HTL-6-28 3.70 ± 0.03 8.67 ± 0.97 — HTL-6-29 1.75 ± 0.41 28.65 ± 1.67 16.37 HTL-6-45 0.027 ± 0.01 0.04 ± 0.02 1.48 HTL-6-47 0.25 ± 0.23 0.03 ± 0.02 0.12 HTL-6-48 0.059 ± 0 1.23 ± 1.20 20.85 HTL-6-49 0.07 ± 0.01 0.07 ± 0.00 1.00 HTL-6-50 >200 0.17 ± 0.05 — HTL-7-01 0.07 ± 0.01 0.23 ± 0.33 3.29 HTL-7-02 >200 >200 — HTL-7-03 0.04 ± 0.01 0.13 ± 0.03 3.25 HTL-7-04 10.53 ± 0.56 10.27 ± 7.81 0.98 HTL-7-05 0.13 ± 0.06 0.75 ± 0.36 5.77 HTL-7-06 0.05 ± 0.01 0.17 ± 0.06 3.40 HTL-7-07 0.38 ± 0.39 0.46 ± 0.03 1.21 HTL-7-08 0.20 ± 0.02 0.70 ± 0.48 3.50 HTL-7-10 0.39 ± 0.18 0.74 ± 0.72 1.90 HTL-7-11 0.095 ± 0.007 0.19 ± 0.10 2.00 HTL-7-12 36.93 ± 0.41 >200 >5.42 HTL-7-13 26.34 ± 12.92 200 ± 0 7.59 HTL-7-14 23.72 ± 1.02 1.00 ± 0.81 0.04 HTL-7-15 0.09 ± 0.02 0.20 ± 0.09 2.22 HTL-7-16 0.09 ± 0.01 0.17 ± 0.06 1.89 HTL-7-17 0.10 ± 0.03 0.18 ± 0.15 1.80 HTL-7-18 1.80 ± 1.17 2.91 ± 1.16 1.62 HTL-7-19 0.045 ± 0.01 0.06 ± 0.02 1.33 HTL-7-20 0.10 ± 0.01 1.79 ± 0.67 17.90 HTL-7-21 2.08 ± 2.29 0.69 ± 0.50 0.33 HTL-7-24 0.09 ± 0.01 0.15 ± 0.01 1.67 HTL-7-25 0.41 ± 0.03 4.42 ± 0.45 10.78 HTL-7-26 0.28 ± 0.20 1.29 ± 0.57 4.61 HTL-7-28 0.09 ± 0.01 0.23 ± 0.01 2.56 HTL-7-29 0.09 ± 0.00 0.29 ± 0.03 3.22
[0509] Among the 46 compounds represented by Formula III, some of the compounds showed strong in vitro anti-EV71 activity, in which aromatic hydrocarbon-substituted compounds represented by Formula III, such as HTL-6-45, HTL-6-48, HTL-6-49, HTL-7-1, HTL-7-3, HTL-7-6, HTL-7-11, HTL-7-15, HTL-7-16, HTL-7-19, HTL-7-28, and HTL-7-29, showed similar activity to that of the positive control compound, their activities were in the same order of magnitude, and their selection index SI values were all greater than 1. Among them, the SI value of HTL-6-48 was greater than 20, and it showed high activity and relatively low toxicity. Among the imine-substituted compounds represented by Formula III, HTL-6-11, HTL-6-12, HTL-6-27, HTL-6-28 and HTL-6-29 showed moderate activity. Among the alkene-substituted compounds represented by Formula III, HTL-7-24 showed strong inhibitory activity, while HTL-7-25 and HTL-7-26 had moderate inhibitory activity on EV71.
Example 100: Experiment of In Vitro Inhibitory Activity of the Compounds of the Present Application on mTOR Kinase
[0510] Experimental Method:
[0511] Preparation of Reaction Buffers:
[0512] Basic buffer composition: 50 mM (mmol/L) HEPES (pH 7.5), 1 mM EGTA, 0.01% Tween-20, 10 mM MnCl.sub.2, 2 mM DTT (diluted from 500 mM before used).
[0513] {circle around (1)} Substrate buffer solution: 1650 μL of 2.5× substrate buffer solution was composed of 1559.6 μL of 1× basic buffer, 89.2 μL of GFP-4E-BP1 (18.5 μM stock solution, purchased from Thermo Fisher, catalog number: PV4759) and 1.2 μL of ATP (10 mM), the final concentrations were 0.4 μM GFP-4E-BP1, 3 μM ATP.
[0514] {circle around (2)} mTOR kinase buffer solution: 1650 μL of 2.5×mTOR kinase buffer solution was composed of 1640.2 μL of 1× basic buffer and 9.8 μL of mTOR (0.21 mg/mL stock solution), and the final concentration was 0.5 μg/mL.
[0515] {circle around (3)} Detection buffer solution: 3960 μL of 2× detection buffer solution was composed of 3797.1 μL of TR-FRET buffer diluent (purchased from Thermo Fisher, catalog number: PV3574), 4.5 μL of Tb-anti-p4E-BP1 antibody (3.49 μM stock solution, purchased from Thermo Fisher, catalog number: PV4757) and 158 μL of EDTA (500 mM stock solution), and the final concentrations were 2 nM Tb-anti-p4E-BP1 antibody and 10 mM EDTA.
[0516] Experimental Steps:
[0517] {circle around (1)} 20 μL of 100% DMSO solution containing 5 mM compound to be tested was added to a 96-well plate.
[0518] {circle around (2)} The compound was serially diluted 3 times in DMSO.
[0519] {circle around (3)} 1 μL of the compound in the previous step was taken, diluted with 19 μL of mTOR kinase buffer, and transferred to another 96-well plate.
[0520] {circle around (4)} 4 μL of mTOR kinase solution (purchased from Thermo Fisher, catalog number: PV4753) was added to a 384-well plate.
[0521] {circle around (5)} 2 μL of the compound in G was taken out and added to the 384-well plate with mTOR kinase, and incubated at room temperature for 15 minutes.
[0522] {circle around (6)} 4 μL of substrate solution was added to initiate the reaction.
[0523] The final concentrations of mTOR reaction solution were: 0.5 μg/mL mTOR, 0.4 μM GFP-4E-BP1, 3 μM ATP.
[0524] The final concentrations of the test compound were: 50000, 16666, 5555, 1851, 617.3, 205.8, 68.58, 22.86, 7.62, 2.54 and 0.85 nM.
[0525] The final concentration of the DMSO solution was 1%.
[0526] {circle around (7)} Incubation was performed for 60 minutes at room temperature.
[0527] {circle around (8)} 10 μL of detection buffer was added. The final concentrations were 2 nM Tb-anti-p4E-BP1 antibody and 10 mM EDTA.
[0528] {circle around (9)} Incubation was performed for 30 minutes at room temperature.
[0529] {circle around (10)} TR-FRET value was read on MD5 multi-mode plate reader (Molecular Devices). The excitation wavelength was 340 nm, the emission wavelength 1 was 495 nm, and the emission wavelength 2 was 520 nm. The ratio of 520 nm/495 nm readings was calculated as TR-FRET value.
[0530] Data Processing:
[0531] IC.sub.50 of compound was fitted by nonlinear regression equation:
Y=Bottom+(Top-Bottom)/(1+10{circumflex over ( )}((LogIC.sub.50-X)*HillSlope));
[0532] X: common logarithm value of compound concentration; Y: TR-FRET value (520 nm/495 nm).
TABLE-US-00005 TABLE 4 Inhibitory activity of some compounds on mTOR kinase Compound name IC.sub.50 (nM) HTL-2-35 7766 HTL-2-38 1013 HTL-5-21 15.05 HTL-5-23 256.80 HTL-6-30 47.57 HTL-6-34 848.60 HTL-6-11 255.30 HTL-6-12 68.16 HTL-6-45 29.24 HTL-6-47 6.17 HTL-6-48 31.46 HTL-6-49 11.94 HTL-7-01 28.76 HTL-7-02 974.20 HTL-7-03 7.25 HTL-7-04 239.90 HTL-7-05 39.03 HTL-7-06 4.82 HTL-7-07 30.17 HTL-7-08 76.60 HTL-7-10 34.79 HTL-7-11 32.51 HTL-7-12 1206 HTL-7-13 86.13 HTL-7-14 306.30 HTL-7-15 25.03 HTL-7-16 8.91 HTL-7-17 1.15 HTL-6-50 158.60 HTL-7-18 22.72 HTL-7-19 7.58 HTL-7-20 11.29 HTL-7-21 11.88
[0533] In this example, the inhibitory activity on mTOR kinase of some compounds of Formula I, Formula II and Formula III that had in vitro EV71 inhibitory activity was tested. The results showed that, similar to the experimental results of the in vitro inhibitory activity against EV71, the compounds represented by Formula I HTL-2-35 and HTL-2-38 had weak inhibitory activity on mTOR kinase; among the 4 compounds represented by Formula II, HTL-5-21 and HTL-6-30 had better inhibitory activity, while HTL-5-23 and HTL-6-34 merely showed moderate inhibitory activity; among the 27 compounds represented by Formula III, except for HTL-6-11, HTL-7-02, HTL-7-04, HTL-7-14 and HTL-6-50 which had moderate inhibitory activity, other compounds showed good enzyme inhibitory activity, in which HTL-6-47, HTL-7-03, HTL-7-06, HTL-7-16, HTL-7-17 and HTL-7-19 showed excellent inhibitory activity, IC.sub.50 of which reached nM level.
Example 101: Molecular Mechanism Experiment of the Compounds of the Present Application
[0534] In order to test the inhibitory activity of the synthesized compounds on the two mTOR complexes, the inhibitory activity experiments of mTORC1 and mTORC2 were carried out. Since mTORC1 and mTORC2 exerted their function by activating the phosphorylation of downstream substrates, the inhibitory activity of the compounds on mTORC1 and mTORC2 could be determined by detecting the phosphorylation levels of the Thr389 site of mTORC1 downstream substrate p70S6K1 and the Ser473 site of mTORC2 downstream substrate Akt.
[0535] Experimental Method:
[0536] Pretreatment of Compound:
[0537] {circle around (1)} RD cells were cultured in DMEM medium containing 10% FBS and 1×PS (penicillin and streptomycin were at concentrations 100 IU and 100 μg/mL, respectively) at 37° C. and 5% CO.sub.2.
[0538] {circle around (2)} The RD cells (5×10.sup.5 cells/2 mL medium) were inoculated into a 6-well plate and incubated at 37° C. and 5% CO.sub.2 for 24 hours.
[0539] {circle around (3)} The cells were washed once with PBS, and the cells were cultured in serum-free medium without nutrients overnight.
[0540] {circle around (4)} Formulation of compounds:
[0541] Preparation of insulin medium: Insulin was diluted in DMEM medium containing 10% FBS and 1×PS so that the final concentration of insulin was 167 nM.
[0542] Pretreatment of compound to be tested: The compound was dissolved in DMSO so that the concentration of the test compound was 20 mM, and the compound was diluted in 167 nM insulin medium to reach a concentration of 20 μM.
[0543] Preparation of rapamycin solution: Rapamycin was dissolved in DMSO to a concentration of 10 mM, and the rapamycin was diluted in 167 nM insulin medium to reach a concentration of 20 μM.
[0544] {circle around (5)} The serum-free medium in each well was removed.
[0545] {circle around (6)} 2 mL of complete medium containing DMSO was added to each well as a control carrier. The final concentration of DMSO was 0.2%.
[0546] {circle around (7)} 2 mL of 20 μM rapamycin solution and 20 μM the compound to be tested were added to the designated wells, respectively. The final concentration of DMSO was 0.2%.
[0547] {circle around (8)} The cells were incubated for 2 hours at 37° C. and 5% CO.sub.2.
[0548] Protein extraction and concentration determination:
[0549] {circle around (1)} The cells were washed once with refrigerated PBS, and then the PBS was discarded.
[0550] {circle around (2)} 150 μL of cell extraction buffer (RIPA, APPLYGEN, catalog number: C.sub.1053) was transferred into each well to lyse the cells, and then the resulting cell solution were incubated on ice for 30 minutes.
[0551] {circle around (3)} Centrifugation was performed at 14000 rpm (13000× g) for 30 minutes at 4° C.
[0552] {circle around (4)} The supernatant was transferred into a new eppendorf tube, and the cell lysate was stored at −80° C. before testing.
[0553] {circle around (5)} The protein concentration was determined by BCA method.
[0554] Preparation of Buffer Solutions:
[0555] {circle around (1)} 100× Protease inhibitor (Beyotime, catalog number: P1005): 1 mL of redistilled water was added to the protease inhibitor and stirred gently until the solid was completely dissolved.
[0556] {circle around (2)} Lysis Buffer: 2 mL of 100× protease inhibitor and 2× phosphatase inhibitor Cocktails PhosSTOP (Beyotime, catalog number: P1082) were added to 100 mL of the cell extract, and stirred gently until it was completely dissolved.
[0557] {circle around (3)} Electrophoresis Running Buffer:
[0558] 10×MOPS buffer: 52.33 g of MOPS, 30.29 g of Tris base, 10 mL of 0.5 mol/L EDTA (pH 8.5) and 5 g of SDS was dissolved in 400 mL of redistilled water, stirred to dissolve, and the pH was adjusted to 7.5, then redistilled water was added again to reach a volume of 500 mL; 1×MOPS: 100 mL of 10×MOPS was diluted with redistilled water to 1000 mL.
[0559] {circle around (4)} 1× Transfer Buffer: 100 mL of 10× transfer buffer (144 g of glucine, 30.3 g of trisbase, and distilled water were mixed to reach a volume of 1 L) and 400 mL of methanol were dissolved in 1500 mL of redistilled water, and then the redistilled water was added to reach a volume of 2000 mL.
[0560] {circle around (5)} 10×PBS Buffer (0.1M): 5 bags of PBS powder (Solarbio, catalog number: P1010) was added into 800 mL of redistilled water, stirred to dissolve, adjusted to have a pH of 7.6, and then the redistilled water was added to reach a volume of 1000 mL.
[0561] {circle around (6)} 1×PBS Buffer: 100 mL of 10×PBS buffer was diluted to 1000 mL with redistilled water.
[0562] {circle around (7)} 10% Tween-20: 20 mL of Tween-20 was added to 180 mL of redistilled water, and stirred well.
[0563] {circle around (8)} 1×PBST Buffer: 100 mL of 10×PBST buffer and 10 mL of Tween-20 were diluted to 1000 mL with re-distilled water.
[0564] {circle around (9)} Primary antibody incubation: the primary antibodies (Thermo Fisher, catalog numbers: B2H9L2 and PA5-85513) were diluted with 0.1% Tween-20 in the blocking solution (5% skimmed milk) at a ratio of 1:1000.
[0565] {circle around (10)} Secondary antibody incubation: IRDye 800CW Goat anti-Rabbit IgG (Abcam, catalog number: ab216773) was diluted with 0.1% Tween-20 in the blocking buffer at a ratio of 1:500.
[0566] Western Blot experiment:
[0567] {circle around (1)} 12 μg of total protein was added to the sample well of SDS-PAGE. Electrophoresis were performed at constant voltage of 120V until the blue marker reached the end of the gel.
[0568] {circle around (2)} At 120V, the protein on the gel was transferred to the PVDF membrane for 40 minutes by using the BIO-RAD Trans-Blot.
[0569] {circle around (3)} After transferring, the blocking was carried out with blocking buffer at room temperature for 2 hours.
[0570] {circle around (4)} The membrane was incubated with the corresponding primary antibody solution on a constant temperature shaker at 4° C. overnight.
[0571] {circle around (5)} The membrane was rinsed with 1×PBST Buffer for 3×10 min, and then incubated with the secondary antibody solution at room temperature for 1 hour.
[0572] {circle around (6)} The membrane was washed with 1×PBST Buffer for 3×10 min, and scanned and developed with Odyssey Infrared Imaging System.
[0573] The RD cells were treated with rapamycin and 33 compounds of Formula I, Formula II and Formula III at concentration of 20 μM for 2 hours, respectively, and the Western blot results were shown in
[0574] The RD cells were treated with 33 compounds of Formula I, Formula II and Formula III of the present application and 2 positive drugs to investigate their effects on the phosphorylation levels of p70 and Akt, the downstream substrates of mTORC1 and mTORC2, to evaluate the inhibitory activity of the compounds on mTORC1 and mTORC2. Through the observation of the results of the Western blot experiment, it was found that under the same detection conditions, the p70 phosphorylation expression level of cells treated with rapamycin was significantly reduced. Among the synthesized compounds, HTL-2-38, HTL-5-21, HTL-6-30, HTL-6-11, HTL-6-12, HTL-6-45, HTL-6-47, HTL-6-48, HTL-6-49, HTL-7-01, HTL-7-03, HTL-7-04, HTL-7-05, HTL-7-06, HTL-7-07, HTL-7-08, HTL-7-10, HTL-7-11, HTL-7-13, HTL-7-14, HTL-7-15, HTL-7-16, HTL-7-17, HTL-6-50, HTL-7-18, HTL-7-19, HTL-7-20 and HTL-7-21 could significantly down-regulate the p70 phosphorylation expression level, indicating that the above compounds could inhibit mTORC1. In addition, the positive control drug rapamycin could not significantly down-regulate the phosphorylation level of Akt, indicating that it could not inhibit the phosphorylation of Akt. Among the synthesized compounds, HTL-5-21, HTL-6-11, HTL-6-12, HTL-6-45, HTL-6-47, HTL-6-48, HTL-6-49, HTL-7-01, HTL-7-03, HTL-7-04, HTL-7-05, HTL-7-06, HTL-7-07, HTL-7-08, HTL-7-10, HTL-7-11, HTL-7-13, HTL-7-14, HTL-7-15, HTL-7-16, HTL-7-17, HTL-6-50, HTL-7-18, HTL-7-19, HTL-7-20 and HTL-7-21 could significantly down-regulate the phosphorylation expression level of Akt, indicating that the above compounds could inhibit mTORC2.
Example 102: Experiment of Drug Metabolism Property of the Compounds of the Present Application
[0575] The metabolic properties of drug are also important indicators for evaluating the pros and cons of the drug. In the in vitro anti-EV71 activity evaluation, some of the compounds represented by Formula III showed strong inhibitory activity against EV71. Among them, HTL-6-45, HTL-6-48, HTL-7-01, HTL-7-03, HTL-7-17 with excellent activity were selected, and their in vivo drug metabolism properties were evaluated.
[0576] Experimental Method:
[0577] Experimental animals: C.sub.57 male mice (purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.), bred for 6-8 weeks and weighing 20-30 g, 3 mice in each group of iv/po groups, a total of 6 groups.
[0578] Preparation of Sample Solutions:
[0579] {circle around (1)} Preparation of sample solution (1 mg/kg, 5 mL/kg) for injection (iv): 1 mg of the sample to be tested was dissolved in 0.5 mL DMSO, mixed by a vortex mixer, and treated ultrasonically to obtain a stock solution (2 mg/mL); 0.10 mL of the stock solution (2 mg/mL) was added to a vial, then 0.4 mL of PEG400 and 0.5 mL of water were added, and mixed by a vortex shaker, treated ultrasonically to obtain a sample solution with a concentration of 0.2 mg/mL.
[0580] {circle around (2)} Preparation of sample solution (10 mg/kg, 10 mL/kg) for oral administration (po): 1 mg of the sample to be tested was dissolved in 1 mL of “0.5% CMC/0.1% Tween-80 aqueous solution”, mixed by a vortex mixer, and treated ultrasonically to obtain a suspension of the sample to be tested with a concentration of 1 mg/mL.
[0581] Liquid Chromatography Method:
[0582] {circle around (1)} Chromatographic column: Waters XSELECT CSH C18, 2.5 μm 2.1×50 mm chromatographic column.
[0583] {circle around (2)} Mobile phase: Phase A: 5% acetonitrile in aqueous solution (0.1% formic acid); Phase B: 95% acetonitrile in aqueous solution (0.1% formic acid).
[0584] {circle around (3)} Flow rate: 0.6 mL/min.
[0585] {circle around (4)} Sampling volume: 20 μL.
[0586] For each compound, po group and iv group were set for administration. All experimental animals were fasted overnight before administration, and all administration was performed at room temperature.
[0587] Sampling time: for iv group, sampling was performed at 0.083, 0.25, 0.5, 1, 2, 4, 8 and 24 hours after injection administration (iv); for po group, sample was performed at 0.25, 0.5, 1, 2, 4, 8 and 24 hours after oral administration (po).
[0588] Sampling Method:
[0589] {circle around (1)} At each time point, about 0.03 mL of blood was collected. The blood of each sample was transferred to a plastic microcentrifuge tube containing heparin sodium anticoagulant, mixed well with the anticoagulant, and then cooled on ice and centrifuged.
[0590] {circle around (2)} The blood sample was centrifuged at 4000 rpm for 5 minutes at 4° C. to obtain plasma.
[0591] {circle around (3)} The samples was stored in a refrigerator at −75±15° C. for later test.
[0592] Analysis and Identification:
[0593] The mixed solution of acetonitrile and water (1:1) was used to dilute the stock solution to prepare a series of working solutions.
[0594] 3 μL of working solution (5, 10, 20, 50, 100, 500, 1000, 5000, 10000 ng/mL) was added to 30 μL of C57 mouse blank plasma to obtain a standard solution with a concentration of 0.5-1000 ng/mL (0.5, 1, 2, 5, 10, 50, 100, 500, 1000 ng/mL), in total of 33 μL. Four quality control samples of 1 ng/mL, 2 ng/mL, 50 ng/mL and 800 ng/mL were used to calibrate the standard curve.
[0595] {circle around (2)} The quality control samples were prepared on the day of analysis, and the method was the same as the standard solution.
[0596] {circle around (3)} 200 μL of acetonitrile containing internal standard solution was added to 33 μL of standard samples, 33 μL of quality control samples and 33 μL of unknown sample (30 μL of plasma and 3 μL of blank solution) to precipitate protein, respectively. The samples were mixed by a vortex mixer for 30 seconds and mixed well. The precipitated samples were centrifuged for 15 minutes at 4° C. and 4000 rpm.
[0597] {circle around (4)} The supernatant was pipetted quantitatively and diluted by 3 times with water, then the diluted supernatant was loaded and quantitatively analyzed by liquid chromatography-mass spectrometry technology.
[0598] {circle around (5)} Detection data were obtained, and pharmacokinetic parameters such as T.sub.1/2, Cmax, AUC, AUC, CL, Vss, F were calculated.
[0599] Through experiments, the drug metabolism data of 5 compounds of the present application were obtained, and the specific information was shown in Table 5:
TABLE-US-00006 TABLE 5 In vivo drug metabolism data of the compounds of the present application C.sub.0(ng/mL) AUC CL V.sub.ss F iv/C.sub.max T.sub.max (h .Math. ng/mL) T.sub.1/2 (h) ((mL/min)/kg) (L/kg) (%) Compd. (ng/mL) po (h) po iv/po iv/po iv iv po HTL-6-45 398/42.9 0.25 95/3.54 0.324/0.84 176 3.25 3.89 HTL-6-48 651/1290 0.25 651/133 0.840/0.99 25.6 1.59 20.2 HTL-7-01 574/298 0.5 592/69.9 1.05/2.01 27.8 2.29 12.2 HTL-7-03 596/565 0.667 668/164 1.83/1.24 24.1 3.24 24.0 HTL-7-17 364/144 0.333 168/35.7 0.718/1.58 99 3.59 21.8
[0600] Through the investigation of several drug metabolism properties, it was found that on the highest blood concentration C.sub.0/C.sub.max index, the oral C.sub.max of compound HTL-6-48 was relatively high, while the C.sub.0/C.sub.max values of several other compounds were relatively low. Comparing the peak time T.sub.max, HTL-6-45 and HTL-6-48 both reached the highest plasma concentration quickly in 0.25 hours and were absorbed relatively quickly, while the T.sub.max of HTL-7-17 was slightly lower, being 0.33 hours; the T.sub.max values of HTL-7-01 and HTL-7-03 were both above 0.5 hours. In terms of area under curve (AUC), the AUC values of HTL-6-48, HTL-7-01 and HTL-7-03 were relatively high through injection administration route, but the AUC of HTL-7-01 through oral administration was lower. In terms of half-life T.sub.1/2, except for HTL-7-03 that had relatively longer half-life through injection administration, all other compounds had relatively lower T.sub.1/2. In terms of clearance rate (CL), except for HTL-6-45 and HTL-7-17 that had relatively higher clearance rate, the other three compounds had relatively lower clearance rates, indicating that they were less liable to be cleared in the body. Comparing the apparent volume of distribution (Vss), the Vss values of HTL-6-48 and HTL-7-01 were lower, indicating that they were less liable to tend to tissue distribution, while the Vss values of HTL-6-45, HTL-7-03 and HTL-7-17 were slightly higher, indicating that they were more liable to tend to tissue distribute. As a therapeutic drug of anti-enterovirus EV71, oral bioavailability (F) was an important drug metabolism index that was focused in this text. Generally speaking, a compound having a bioavailability greater than 20% indicates it had a certain potential to be developed as drugs. Among the experimental data, the F values of HTL-6-48, HTL-7-03 and HTL-7-17 were all greater than 20%, indicating that they were candidate compounds that could possibility be further developed into drugs.
Example 103: Water Solubility Experiment of the Compounds of the Application
[0601] Water solubility is important physical and chemical property that affects the oral absorption of drugs. Therefore, we selected five compounds HTL-6-45, HTL-6-48, HTL-7-01, HTL-7-03 and HTL-7-17 which had certain potential to be developed as drugs among the compounds of the present application and tested their water solubility.
[0602] Experimental Method:
[0603] By measuring the solubility of compound in a saturated aqueous solution, the water solubility of the compound was further investigated.
[0604] Preparation of Supersaturated Solution:
[0605] {circle around (1)} About 1 mg of the compound to be tested was weighed and added to a 25 mL tube;
[0606] {circle around (2)} 5 mL of deionized water was added, shaken well, placed in a vortex shaker and mixed for 30 seconds, then treated ultrasonically for 30 seconds;
[0607] {circle around (3)} The tube was placed in a constant temperature water bath, and allowed to stand in a constant temperature water bath at 25° C. for 1 hour;
[0608] {circle around (4)} The operation of {circle around (2)} was repeated, and the standing at constant temperature of 25° C. was continued for 1 hour;
[0609] {circle around (5)} The prepared supersaturated solution supernatant was taken out and placed in a centrifuge tube, centrifuged at 15000 rpm for 5 minutes, and the upper centrifugate was taken out for later use.
[0610] Liquid Chromatography Method:
[0611] {circle around (1)} Chromatographic column: Agilent XDB Eclipse C18, 25 cm×4.6 mm×5 μm liquid chromatography column.
[0612] {circle around (2)} Mobile phase: Phase A: water (0.1% trifluoroacetic acid); Phase B: acetonitrile.
[0613] {circle around (3)} Flow rate: 1.0 mL/min.
[0614] {circle around (4)} Sampling volume: 10 μl.
[0615] Preparation of standard solution: 1.0 mg of the sample to be tested was accurately weighed, added to a 10 mL volumetric flask, dissolved in methanol as solvent, metered to a constant volume of 10 mL, the concentration of solution was calculated, and it was used as a standard solution for later use.
[0616] By using the one-point method, the chromatographic peak area of the standard solution of the sample to be tested was determined, and then the peak area of the supersaturated compound was determined, and the solubility of the compound to be tested in water was calculated accordingly.
[0617] Through experiments, the solubility values of 5 compounds of the present invention in water were measured, and the results were shown in Table 6:
TABLE-US-00007 TABLE 6 Water solubility of the compounds of the present application Compound name HTL-6-45 HTL-6-48 HTL-7-01 HTL-7-03 HTL-7-17 Water 0.39 5.88 0.42 0.61 18.64 solubility (μg/mL)
[0618] The water solubility test results showed that among the 5 compounds in the present application, HTL-6-48 and HTL-7-17 had relatively high water solubility and were compounds with high biological activity, and good drug metabolism and water solubility, and had the potential to be further developed as drugs.