SPIROCYCLIC ARYL PHOSPHORUS OXIDE AND ARYL PHOSPHORUS SULFIDE
20170129909 ยท 2017-05-11
Assignee
Inventors
- Zhaozhong Ding (Shanghai, CN)
- Minghui Zhang (Jinan, CN)
- Shuhui Chen (Shanghai, CN)
- Xile LIU (Shanghai, CN)
- Yidong Zhu (Jinan, CN)
- Chuanwen Fan (Jinan, CN)
- Baoping Zhao (Shanghai, CN)
- Long Zhang (Jinan, CN)
- Yingying YANG (Jinan, CN)
- Qingmei ZHENG (Jinan, CN)
- Shansong ZHENG (Jinan, CN)
- Haiwen WAN (Shanghai, CN)
- Jinqing Hu (Shanghai, CN)
Cpc classification
C07F9/65615
CHEMISTRY; METALLURGY
A61K45/06
HUMAN NECESSITIES
A61K31/675
HUMAN NECESSITIES
C07F9/6558
CHEMISTRY; METALLURGY
C07F9/65583
CHEMISTRY; METALLURGY
A61P35/00
HUMAN NECESSITIES
International classification
C07F9/6558
CHEMISTRY; METALLURGY
C07F9/6561
CHEMISTRY; METALLURGY
A61K31/675
HUMAN NECESSITIES
Abstract
Disclosed is a spirocyclic aryl phosphorus oxide or sulfide as an ALK inhibitor, particularly, a compound represented by formula (I) as an ALK inhibitor or pharmaceutically acceptable salt thereof.
##STR00001##
Claims
1. A compound represented by formula (I) or pharmaceutically acceptable salt thereof, ##STR00208## wherein, T.sub.1 is selected from N or C(R.sub.01); T.sub.2 is selected from N(R.sub.01), O, S(O).sub.2 or CH(NR.sub.01R.sub.02); R.sub.01 or R.sub.02 is separately and independently selected from H, or C.sub.1-6 alkyl, C.sub.1-6 heteroalkyl, C.sub.3-6 cycloalkyl-(CH.sub.2).sub.0-3- or C.sub.3-6 heterocyclohydrocarbyl-(CH.sub.2).sub.0-3 which is optionally substituted by 1, 2 or 3 halogen, hydroxyl and/or cyano; wherein the hetero is 1, 2 or 3 group(s) seleted from O, S, N, S(O).sub.2 and/or S(O); optionally, R.sub.01 and R.sub.02 on T.sub.2 are together linked to the same N atom to form a 3-6 membered ring, the ring contains 1, 2 or 3 heteroatom(s), the heteroatom is selected from O, S or N; each of D.sub.1-D.sub.4 is separately and independently selected from (CR.sub.1R.sub.2).sub.1-3, O, S, C(O), S(O).sub.2 or S(O); D is selected from N(R.sub.01), O or S; W is selected from O, S, N(CN) or N(OMe); R.sub.3 is selected from R.sub.03, OR.sub.03 or SR.sub.03; R.sub.03 is selected from C.sub.1-4 alkyl, C.sub.1-4 haloalkyl or C.sub.3-5 cycloalkyl-(CH.sub.2).sub.0-3; Z is selected from N or C(R.sub.4); X.sub.1 is selected from C(R.sub.x1) or N; X.sub.2 is selected from C(R.sub.x2) or N; R.sub.x1, R.sub.x2, R.sub.1, R.sub.2 and R.sub.4 are separately and independently selected from H, F, Cl, Br, I, CN, OH, SH, NH.sub.2, or C.sub.1-6 alkyl, C.sub.1-6 heteroalkyl, C.sub.3-6 cyclohydrocarbyl-(CH.sub.3-6 or C.sub.3-6 heterocyclohydrocarbyl-(CH.sub.2).sub.0-3 which is optionally substituted by 1, 2 or 3 halogen, hydroxyl, and/or cyano; wherein the hetero represents 1, 2 or 3 heteratom(s) which is selected from O, S or N; R.sub.p1 and R.sub.p2 are separately and independently selected from H, C.sub.1-4 alkyl or C.sub.1-4 haloalkyl; optionally, R.sub.p1 and R.sub.p2 are together linked to the same P group to form a 5-6 membered ring, the ring contains 1, 2 or 3 heteratom(s), the heteroatom is selected from O, S, N or P; and optionally, the positions of Z and ##STR00209## can be interchanged.
2. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, wherein R.sub.01 and R.sub.02 are separately and independently selected from H, CH.sub.3, CD.sub.3, CF.sub.3, CHF.sub.2, CH.sub.2CH.sub.3, CH(CH.sub.3).sub.2, CH.sub.2CF.sub.3, CH.sub.2CH.sub.2CF.sub.3, CH.sub.2CH.sub.2F, CH.sub.2CH.sub.2S(O).sub.2CH.sub.3, CH.sub.2CH.sub.2CN, ##STR00210## CH.sub.2CH(OH)(CH.sub.3).sub.2, CH.sub.2CH(F)(CH.sub.3).sub.2 or CH.sub.2CH.sub.2F.
3. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, wherein R.sub.03 is selected from CH.sub.3, CD.sub.3, CF.sub.3, CHF.sub.2, CH.sub.2CH.sub.3, CH(CH.sub.3).sub.2, CH.sub.2CF.sub.3, CH.sub.2CH.sub.2CF.sub.3, CH.sub.2CH.sub.2F or ##STR00211##
4. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, wherein R.sub.p1 and R.sub.p2 are separately and independently selected from H, CH.sub.3, CD.sub.3, CF.sub.3, CHF.sub.2, CH.sub.2CH.sub.3, CH(CH.sub.3).sub.2, CH.sub.2CF.sub.3, CH.sub.2CH.sub.2CF.sub.3 or CH.sub.2CH.sub.2F.
5. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, wherein R.sub.x1, R.sub.x2, R.sub.1, R.sub.2 and R.sub.4 are separately and independently selected from H, F, Cl, Br, I, CH.sub.3, CD.sub.3, CF.sub.3, CHF.sub.2, CH.sub.2CH.sub.3, CH(CH.sub.3).sub.2, CH.sub.2CF.sub.3, CH.sub.2CH.sub.2CF.sub.3, CH.sub.2CH.sub.2F, CH.sub.2CH.sub.2S(O).sub.2CH.sub.3, CH.sub.2CH.sub.2CN, ##STR00212## CH.sub.2CH(OH)(CH.sub.3).sub.2, CH.sub.2CH(F)(CH.sub.3).sub.2 or CH.sub.2CH.sub.2F.
6. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, wherein NR.sub.01R.sub.02 on T.sub.2 is selected from NHCH.sub.3, N(CH.sub.3).sub.2, N(CH.sub.2CH.sub.3).sub.2, ##STR00213##
7. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, wherein D, D.sub.1-4, or T.sub.2 is separately and independently selected from NH,NMe- or O; D.sub.1-4 or T.sub.2 can also be selected from CH(NCH.sub.3CH.sub.3).
8. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, wherein the spiro moiety ##STR00214## is selected from ##STR00215##
9. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, which is represented by formula (II): ##STR00216## wherein each variable is defined as that in any one of claims 1-8.
10. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, which is represented by formula (III): ##STR00217## wherein each variable is defined as that in any one of claims 1-8.
11. The compound or pharmaceutically acceptable salt thereof as defined in claim 1, which is selected from the group consisting of ##STR00218## ##STR00219## ##STR00220## ##STR00221## ##STR00222## ##STR00223## ##STR00224## ##STR00225## ##STR00226##
12. A process for preparing the compound represented by formula (I) as defined in claim 1, wherein T.sub.1 represents N, T.sub.2 represents NH, the preparation route of which was shown as scheme A or C: ##STR00227## ##STR00228## ##STR00229## ##STR00230## wherein PG is an amino-protecting group, preferably BOC, Bn and Cbz, other variables are defined as any one of preceding claims.
13. The process for preparing the compound as defined in claim 12, wherein the process for preparing A5 is shown as scheme B: ##STR00231##
14. A use of the compound or pharmaceutically acceptable salt thereof as defined in any one of claims 1 to 11 in manufacturing a medicament for the treatment of cancer which is related to ALK and/or EGFR and mutation thereof, or in combined therapy with ROS1, BRAF, c-MET, HER2, KRAS/MEK, PIK3CA, FDFR, DDR2 and/or VEGFR inhibitors for treating cancer, or in combined therapy with cytotoxin for treating cancer.
Description
DETAILED DESCRIPTION OF THE INVENTION
[0100] To describe the present invention in more detail, the following examples are illustrated. However, the scope of the present invention is not limited thereto.
Route A
[0101] ##STR00033##
EXAMPLE 1
(2-((5-chloro-2-((2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)phenyl)amino)pyrimidin-4-yl) amino)phenyl)dimethylphosphine oxide
[0102] ##STR00034##
EXAMPLE 1A
[0103] ##STR00035##
[0104] The mixture of 1-bromine-4-methoxy benzene (8.15 g, 43.6 mmol), potassium vinyltrifluoroborate (7.08 g, 52.8 mmol), Pd(dppf)Cl.sub.2 (1.6 g, 2.20 mmol) and cesium carbonate (28.7 g, 88.1 mmol) in 1,4-dioxane (120 mL) and water (25 mL) was heated to 110 C., stirred for 16 hours. TLC showed the reaction was complete, the reaction mixture was filtered, the filtrate was concentrated to give crude product; the crude product was purified by column chromatography (PE) to give the title compound (yellow oil, 3.45 g, yield 59%).
EXAMPLE 1B
2,2-dichloro-3-(4-methoxyphenyl)cyclobutanone
[0105] ##STR00036##
[0106] The mixture of Example 1A (3.45 g, 9.33 mmol) and copper-zinc reagent (3.18 g, 64.3 mmol) in anhydrous THF (25.0 mL) was heated to reflux, then phosphorus oxychloride (7.78 g, 50.7 mmol) and a solution of 2,2,2-trichloro-acetyl chloride (8.75 g, 48.1 mmol) in anhydrous THF (25.0 mL) were added sequentially. After 1 hour, the reaction mixture was stirred for 12 hours under reflux. TLC (PE) showed that the reaction was complete, the mixture was filtered, the filtrate was concentrated in vacuum to give brown oil compound (6.0 g, crude product), which was used directly to the next reaction.
EXAMPLE 1C
3-(4-methoxyphenyl)cyclobutanone
[0107] ##STR00037##
[0108] Under stirring, to a mixture of zinc powder (6.4 g, 98.4 mmol) in HOAc (25.0 mL) was added a solution of Example 1B (6.0 g, 24.6 mmol) in HOAc (25.0 mL). The reaction mixture was heated to 70 C., and stirred for 4 hours. The reaction mixture was filtered and concentrated to dry to give the crude product, which was purified by pre-HPLC to give a compound as light yellow oil (1.02 g, yield 24%). .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.23 (dd, J=6.8, 1.6 Hz, 2H), 6.91 (dd, J=9.6, 2.8 Hz, 2H), 3.82 (s, 3H), 3.70-3.55 (m, 1H), 3.54-3.40 (m, 2H), 3.70-3.55 (m, 2H).
EXAMPLE 1D
Methyl 2-(3-(4-methoxyphenyl)cyclobutylidene)acetate
[0109] ##STR00038##
[0110] At 0 C., to a suspension of the sodium hydride (278 mg, 6.96 mmol, 60%) in anhydrous THF (20.0 mL) was added methyl-2-(diethoxy phosphoryl) ethyl acetate (1.46 g, 6.96 mmol), the reaction mixture was stirred for 0.5 hours at 0 C. Then a solution of Example1C (1.02 g, 5.80 mmol) in anhydrous THF (10 mL) was dropwise added to the reaction mixture. The reaction mixture was heated to 23 C., and stirred for 16 h at 23 C. TLC (PE:ethyl acetate=10:1) showed the reaction was complete. The reaction mixture was cooled to 0 C., H.sub.2O (20 mL) was slowly added to quench the reaction, and the mixture was extracted with EtOAc (20 mL). The organic layer was washed respectively with H.sub.2O (10 mL) and brine (10 mL) and then dried over anhydrous sodium sulphate and concentrated to dry to give the crude product, which was purified by column chromatography (PE:ethyl acetate=10:1) to give the title compound (0.99 g, 73%) as colorless oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.21 (d, J=8.8 Hz, 2H), 6.90 (d, J=8.8 Hz, 2H), 5.80-5.68 (m, 1H), 3.83 (s, 3H), 3.73 (s, 3H), 3.58-3.48 (m, 2H), 3.33-3.16 (m, 2H), 2.98-2.90 (m, 1H).
EXAMPLE 1E
Methyl 2-(3-(4-methoxyphenyl)-1-(nitromethyl)cyclobutyl)acetate
[0111] ##STR00039##
[0112] To a mixture of Example 1D (0.99 g, 4.27 mmol) in anhydrous THF (25.0 mL) were added nitromethane (521 mg, 8.54 mmol) and TBAF (1.67 g, 6.41 mmol). Then the reaction mixture was heated to 70 C. and stirred for 16 hours. After cooled to 18 C., the mixture was concentrated to dry to give the crude product, which was purified by column chromatography (PE:ethyl acetate=9:1) to give the title compound (1.15 g, yield 92%) as light yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.18-7.08 (m, 2H), 6.94-6.88 (m, 2H), 4.89 (s, 1H), 4.71 (s, 1H), 3.82 (s, 3H), 3.75 (s, 1.5H), 3.71 (s, 1.5H), 3.65-3.47 (m, 1H), 2.90 (s, 1H), 2.75 (s, 1H), 2.65-2.57 (m, 1H), 2.52-2.44 (m, 1H), 2.32-2.26 (m, 1H), 2.20-2.10 (m, 1H).
EXAMPLE 1F
2-(4-Methoxyphenyl)-6-azaspiro[3.4]octan-7-one
[0113] ##STR00040##
[0114] At 18 C. and under nitrogen gas atomsphere, to a mixture of Example 1E (1.15 g, 3.92 mmol) in methanol (15.0 mL) was added Raney Ni (0.20 g). After heated to 50 C., the reaction mixture was stirred for 16 hours under hydrogen gas atmosphere (pressure: 50 psi). LCMS showed that the reaction was complete. The reaction mixture was filtered through diatomite, the filtrate was concentrated in vacuum to give the title compound (0.77 g, crude product) as a white solid. LCMS (ESI) (10-80CD): m/z: 232.2 [M+1].
EXAMPLE 1G
2-(4-Methoxyphenyl)-6-methyl-6-azaspiro[3.4]octan-7-one
[0115] ##STR00041##
[0116] At 0 C., to a mixture of NaH (60 mg, 1.5 mmol, 60%) in anhydrous DMF (5.0 mL) was added Example 1F (230 mg, 1.0 mmol); the reaction mixture was stirred for 1 hour under 0 C. Then Mel (213 mg, 1.5 mmol, dissolved in 5 mL anhydrous THF) was dropwise added to the above mixture. The mixture was heated to 16 C. and stirred for 16 hours. LCMS showed that the reaction was complete, the reaction was quenched with H.sub.2O (20 mL), and extracted with ethyl acetate (20 mL). The organic layer was washed with H.sub.2O (10 mL) and brine (10 mL) respectively, dried over anhydrous sodium sulfate and concentrated to dry to give the title compound (0.24 g, crude product) as yellow oil. LCMS (ESI) (5-95AB): m/z: 246.3 [M+1].
EXAMPLE 1H
2-(4-Methoxy-3-nitrophenyl)-6-methyl-6-azaspiro[3.4]octan-7-one
[0117] ##STR00042##
[0118] A solution of Example 1G (240 mg, 1.0 mmol) in acetic anhydride (5.0 mL) was cooled to 0 C., concentrated nitric acid (1.0 mL) was slowly added into the reaction mixture, and then the reaction mixture was heated to 18 C. and stirred for 2 hrs. LCMS showed the reaction was complete. The mixture filtered, the filtrate was concentrated in vacuum to give the title compound (202 mg, crude) as yellow oil. LCMS (ESI) (5-95AB): m/z: 291.2 1[M+1].
EXAMPLE 1I
2-Methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline
[0119] ##STR00043##
[0120] At 0 C., to a solution of Example 1H (202 mg, 0.70 mmol) in THF (10.0 mL) was added LAH (106 mg, 2.80 mmol); the reaction mixture was heated to 70 C. and stirred for 12 hrs. LCMS showed that the reaction was complete, the reaction mixture was cooled to 0 C., ethyl acetate (20.0 mL) was dropwise added to quench the reaction, followed by adding H.sub.2O (2.0 mL), the mixture was filtered, the filtrate was concentrated in vacuum to give the title compound (120 mg, crude) as brown oil. LCMS (ESI) (10-80CD): m/z: 247.2 [M+1].
EXAMPLE 1J
Dimethylphosphine Oxide
[0121] ##STR00044##
[0122] A solution of MeMgBr (1.30 mol, 434.46 mL) in THF (800 mL) was cooled to 0 C. under N.sub.2, diethyl phosphate (60.0 g, 434.46 mmol, dissolved in 40 mL THF) was dropwise added into the reaction mixture over 2 hrs and the internal temperature was maintained below 0 C. After the addition, the reaction mixture was heated to 20 C. and stirred for 14 hrs. A solution of potassium carbonate (177 g, dissolved in 250 mL H.sub.2O) was add to the reaction mixture to quench the reaction. White solid precipitated, filtered, the cake was washed with ethanol (100 mL), the filtrate was concentrated, and the solid precipitated during the concentration was filtered. Toluene (200 mL) was added to the filtrate, and concentrated to dry to remove H.sub.2O, thereby obtaining the title compound (30.11 g, 385.78 mmol, yeild 88.79%) as colorless thick oil. .sup.1HNMR (400 MHz, CDCl.sub.3): , 7.74-7.67 (m, 0.5H), 6.57-6.52 (m, 0.5H), 1.56 (d, J=3.6 Hz, 3H), 1.53 (d, J=3.6 Hz, 3H).
EXAMPLE 1K
(2-Aminophenyl)dimethylphosphine oxide
[0123] ##STR00045##
[0124] 2-Iodoaniline (12.50 g, 57.07 mmol), Example 1J (5.35 g, 68.49 mmol), K.sub.3PO.sub.4 (14.54 g, 68.49 mmol), Xantphos (660.44 mg, 1.14 mmol) and palladium acetate (256.26 mg, 1.14 mmol) were add into DMF (80 mL), the reaction mixture was heated to 100 C. and stirred for 16 hrs under N.sub.2. LCMS (DCM:methanol=10:1) showed that the reaction was complete. The mixture was filtered and concentrated, the residue was diluted with aq.HCl (1N, 80 mL), followed by adjusting pH to 2, the resultant mixture was filtered. The filtrate was extracted with DCM (100 mL2), the aqueous phase was separated, adjusted pH to about 9 with aq.Na.sub.2HCO.sub.3, and then extracted with DCM (200 mL2). The organic layer was dried over anhydrous sodium sulfate and concentrated to dry. The crude product was purified by recrystallization (PE:ethyl acetate=5:1) to give the title compound (6.00 g, 35.47 mmol, yield 62.15%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.20 (t, J=7.6 Hz, 1H), 7.04 (dd, J=13.6, 7.6 Hz, 1H), 6.69-6.58 (m, 2H), 5.35 (br s, 2H), 1.75 (s, 3H), 1.71 (s, 3H). LCMS (ESI) (10-80CD): m/z: 170.1 [M+1].
EXAMPLE 1L
(2-((2,5-Dichloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0125] ##STR00046##
[0126] At 16 C., to a mixture of Example 1K (2.50 g, 14.8 mmol) and 2,4,5-trichloropyrimidine (2.85 g, 15.5 mmol) in DMF (20 mL), was added DIPEA (3.82 g, 29.6 mmol). The reaction mixture was heated to 70 C. and stirred for 16 hrs. TLC showed that the reaction was complete. The reaction mixture was diluted with H.sub.2O (50 mL), and extracted with EtOAc (40 mL3). The combined organic layer was washed with brine (20 mL2), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum. The crude was recrystallized in ethanol to give the title compound (3.20 g, 10.1 mmol, yeild 68.4%) as white solid. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.50 (dd, T=8.0, 4.0 Hz, 1H), 8.35-8.28 (m, 1H), 7.69-7.59 (m, 2H), 7.36-7.28 (m, 1H), 1.91 (s, 3H), 1.88 (s, 3H). LCMS (ESI) (5-95AB): mh: 315.9 [M+1].
EXAMPLE 1M
(2-((5-Chloro-2-((2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0127] ##STR00047##
[0128] To a mixture of Example 1I (120 mg, 0.49 mmol), Example 1L (154 mg, 0.49 mmol) in tert-butanol (5.0 mL) was added MeSO.sub.3H (141 mg, 1.47 mmol); the reaction mixture was heated to 90 C. and stirred for 12 hrs under N.sub.2. LCMS showed that the reaction was complete, the mixture was concentrated in vacuum to give the crude, which was purified by pre-HPLC to give the title compound (52.2 mg, yeild 20%) as brown oil. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.45 (s, 1H), 8.31-8.27 (m, 1H), 8.10 (s, 1H), 7.82 (s, 1H), 7.70-7.64 (m, 1H), 7.57-7.51 (m, 1H), 7.34-7.30 (m, 1H), 6.95-6.89 (m, 2H), 3.86 (s, 3H), 3.53 (s, 1H), 3.44 (s, 1H), 3.35-3.32 (m, 1.5H), 3.28-3.22 (m, 1.5H), 2.96 (s, 1.5H), 2.88 (s, 1.5H), 2.44-2.39 (m, 1H), 2.36-2.27 (m, 2H), 2.21-2.14 (m, 2H), 2.12-2.08 (m, 1H), 1.87 (s, 3H), 1.84 (s, 3H). LCMS (ESI) (5-95AB): m/z: 526.2 [M+1].
EXAMPLE 2
2-(3-((5-Chloro-4-((2-(dimethylphosphoryl)phenyl)amino)pyrimidin-2-yl)amino)-4-methoxyphenyl)-6-methyl-6-azaspiro[3.4]octan-7-one
[0129] ##STR00048##
EXAMPLE 2A
2-(3-Amino-4-methoxyphenyl)-6-methyl-6-azaspiro[3.4]octan-7-one
[0130] ##STR00049##
[0131] Under Ar gas atomsphere, to a solution of Example 1H (701 mg, 2.42 mmol) in MeOH (5.0 mL) was added Pd/C (150 mg); the mixture was reacted under H.sub.2 (pressure:15 psi) for 16 hrs at 22 C. LCMS showed the reaction was complete, the reaction mixture was filtered through diatomite. The filtrate was concentrated in vacuum thereby giving the title compound (596 mg, crude) as white solid. LCMS (ESI) (10-80CD): m/z: 261.2 [M+1].
EXAMPLE 2B
2-(3-((5-Chloro-4-((2-(dimethylphosphoryl)phenyl)amino)pyrimidin-2-yl)amino)-4-methoxyphenyl)-6-methyl-6-azaspiro[3.4]octan-7-one
[0132] ##STR00050##
[0133] The process for this Example is the same as that for Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced by 2-(3-amino-4-methoxyphenyl)-6-methyl-6-azaspiro[3.4]octan-7-one. The title compound was obtained as brown oil (yeild 32%). LCMS (ESI) (5-95AB): m/z: 540.1 [M+1].
Route B
[0134] ##STR00051## ##STR00052##
EXAMPLE 3
(2-((5-Chloro-2-((2-methoxy-4-(2,8-diazaspiro[4.5]decan-8-yl)phenyl)amino)pyrimidin-4-ylamino)phenyl)dimethylphosphine oxide
[0135] ##STR00053##
EXAMPLE 3A
tert-Butyl 4-(2-ethoxy-2-oxoethylidene)piperidine-1-carboxylate
[0136] ##STR00054##
[0137] At 0 C., to a solution of ethyl 2-(diethoxyphosphoryl) acetate (6.18 g, 27.6 mmol) in anhydrous THF (100 mL) was added NaH (1.2 g, 30.1 mmol, 60%) in portions slowly, the mixture was stirred for 1 hr at 0 C., then tert-butyl-4-oxopiperidine-1-formate (5 g, 25.1 mmol) was added to the reaction mixture slowly. The reaction mixture was stirred for 12 hrs under 0 C. TLC (PE:ethyl acetate=3:1) showed that the reaction was complete. The reaction was quenched by H.sub.2O (50 mL), extracted with EtOAc (100 mL), the organic layer was dried over anhydrous sodium sulfate and concentrated to dry to give the title compound (4.95 g, yield 73%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 5.73 (s, 1H), 4.23-4.14 (m, 2H), 3.53-3.47 (m, 4H), 2.96-2.93 (m, 2H), 2.31-2.28 (m, 2H), 1.48 (s, 9H), 1.28-1.32 (m, 3H).
EXAMPLE 3B
tert-Butyl 4-(2-ethoxy-2-oxoethyl)-4-(nitromethyl)piperidine-1-carboxylate
[0138] ##STR00055##
[0139] At 30 C., to a reaction mixture of Example 3A(5 g, 18.6 mmol) in MeCN (100 mL) were added DBU (5.62 g, 37.0 mmol) and nitromethane (2.3 g, 37 mmol), the reaction mixture was heated to 80 C. and stirred for 12 hrs, TLC(PE:ethyl acetate=3:1) showed that the reaction was complete. The reaction mixture was concentrated to remove the solvent; the residue was diluted with EtOAc (50 mL), and sequentially washed with H.sub.2O (30 mL), saturated NaHCO.sub.3 (20 mL) and brine (20 mL); the organic layer was dried over anhydrous sodium sulfate, concentrated to give brown oil, which was purified by column chromatograph (PE:ethyl acetate=10:1, 8:1) to give the title compound (3.2 g, yield 52.4%) as colorless oil. .sup.1H NMR (400 MHz, CDCl3): , 4.76 (s, 2H), 4.24-4.16 (q, J=7.2 Hz, 2H), 3.57-3.52 (m, 2H), 3.44-3.39 (m, 2H), 2.64 (s, 2H), 1.68-1.63(m, 4H), 1.48 (s, 9H), 1.30 (t, J=7.2 Hz, 3H). LCMS (ESI) (5-95AB): m/z: 353.1 [M+Na.sup.+].
EXAMPLE 3C
Ethyl tert-butyl 3-oxo-2,8-diazaspiro[4.5]decane-8-carboxylate
[0140] ##STR00056##
[0141] A solution of Example 3B (1.5 g, 4.54 mmol) in methanol (50 mL) was stirred for 10 minutes, under Ar gas atomsphere, was added Raney Ni (150 mg, 10%). The reaction mixture was heated to 50 C. and stirred for 12 hrs under H.sub.2 (pressure: 50 psi). TLC (PE:ethyl acetate=10:1) showed that the reaction was complete. The reaction mixture was filtered and concentrated to give the title compound (920 mg, yeild 80%) as white solid. LCMS (ESI) (5-95AB): m/z: 509.4 [2M+1].
EXAMPLE 3D
2,8-diazaspiro[4.5]decan-3-one
[0142] ##STR00057##
[0143] A mixture of Example 3C (920 mg, 3.62 mmol) in DCM/TFA (5 mL/5 mL) was stirred for 12 hrs at 30 C., TLC (PE:ethyl acetate=1:1) showed the reaction was complete. The reaction mixture was concentrated in vacuum to obtain crude, the crude was dissolved in H.sub.2O (30 mL), adjusted pH to 10 with a solution of sodium hydroxide (1N); the mixture was extracted with DCM (15 mL4). The combined organic layer was dried over anhydrous sodium sulfate and concentrated to give the title compound (550 mg, yield 98%) as yellow oil.
EXAMPLE 3E
8-(3-Methoxy-4-nitrophenyl)-2,8-diazaspiro[4.5]decan-3-one
[0144] ##STR00058##
[0145] To a solution of Example 3D (550 mg, 3.57 mmol) in DMF(15 mL), were added potassium carbonate (1.5 g, 10.7 mmol) and 4-fluoro-2-methoxy-1-nitrobenzene (855 mg, 5.0 mmol); the reaction mixture was heated to 90 C. and stirred for 12 hrs. TLC (DCM:methanol=20:1) showed that the reaction was complete. The reaction mixture was diluted with EtOAc (50 mL), sequentially washed with H.sub.2O (20 mL), sat. NaHCO.sub.3 (20 mL), brine (20 mL), the organic layer was dried over anhydrous Na.sub.2SO.sub.4, filtered and concentrated to give brown oil, the crude was purified by column chromatography (DCM:methanol=20:1) to give the title compound (1.02 g, yield 95%) as yellow solid. .sup.1H NMR (400 MHz, DMSO-d6): , 7.89 (d, J=9.6 Hz, 1H), 7.59 (s, 1H), 6.60 (dd, J=2.4 Hz, 9.6 Hz, 1H), 6.51 (d, J=2.4 Hz, 1H), 3.90 (s, 3H), 3.59-3.56 (m, 2H), 3.44-3.42 (m, 2H), 3.17 (d, J=5.2 Hz 2H), 3.10 (s, 2H), 1.64-1.61 (t, J=5.6 Hz, 4H). LCMS (ESI) (5-95AB): miz: 306.3 [M+1].
EXAMPLE 3F
8-(4-Amino-3-methoxyphenyl)-2,8-diazaspiro[4.5]decan-3-one
[0146] ##STR00059##
[0147] At 30 C., to a mixture of Example 3E (500 mg, 1.64 mmol) in EtOH (10 mL) and H.sub.2O (10 mL), were added ferrous powder (367 mg, 6.56 mmol) and NH.sub.4Cl (260 mg, 4.92 mmol). The reaction mixture was heated to 80 C. and stirred for 5 hrs, TLC (DCM:methanol=20:1) showed that the reaction was complete. The reaction mixture filtered, the filtrate was concentrated in vacuum to give title compound (450 mg, yield 99%) as black oil.
EXAMPLE 3G
2-Methoxy-4-(2,8-diazaspiro[4.5]decan-8-yl)aniline
[0148] ##STR00060##
[0149] A solution of Example 3F (400 mg, 1.45 mmol) in anhydrous THF (10 mL) was stirred for 0.5 hrs at 0 C., LiAlH.sub.4 (58 mg, 1.53 mmol) was added in portions to the above mixture. After addition, the mixture was heated to 80 C. and reacted for 12 hrs. TLC (DCM:methanol=6:1) showed that the reaction was complete. The reaction mixture was quenched with H.sub.2O(10 mL), and extracted with DCM (10 mL3), the organic layer was separated, dried and concentrated to give the crude as black oil, the crude was purified by pre-HPLC to give the title compound (80 mg, yield 20%) as white solid.
EXAMPLE 3H
(2-((5-Chloro-2-((2-methoxy-4-(2,8-diazaspiro[4.5]decan-8-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0150] ##STR00061##
[0151] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 2-methoxy-4-(2,8-diazaspiro[4.5]decan-8-yl)aniline. The title compound was obtained as white solid (yield 20%). LCMS (ESI) (5-95AB): m/z: 541.1 [M+1].
EXAMPLE 4
(2-((5-Chloro-2-((2-methoxy-4-(2,7-diazaspiro[3.5]nonan-7-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0152] ##STR00062##
EXAMPLE 4A
tert-butyl 7-(4-methoxy-3-nitrophenyl)-2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester
[0153] ##STR00063##
[0154] A mixture of tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylate (100 mg, 0.44 mmol), 4-fluoro-2-methoxy-1-nitrobenzene (113 mg, 0.66 mmol) and potassium carbonate (152 mg, 1.1 mmol) in DMSO (3 mL) was heated to 90 C. and stirred for 12 hrs. LCMS showed that the reaction was complete, to the reaction mixture was added H.sub.2O (20 mL), and extracted with DCM (25 mL2). The organic layer was washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated to give the crude. Tthe crude was purified by pre-TLC (PE:ethyl acetate=1:1) to give the title compound (150 mg, yield 90%) as yellow oil. LCMS (ESI) (5-95AB): m/z: 378.1 [M+1].
Example 4B
tert-butyl 7-(3-amino-4-methoxyphenyl)-2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester
[0155] ##STR00064##
[0156] This Example was prepared according to the process as described in Example 3F, 8-(3-methoxy-4-nitrophenyl)-2,8-diazaspiro[4.5]decan-3-one was replaced by tert-butyl 7-(4-methoxy-3-nitrophenyl)-2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester to give the title compound as green oil (crude), which was used directly to the next step. LCMS (ESI) (5-95AB): m/z: 348.2 [M+1].
EXAMPLE 4C
(2-((5-Chloro-2-((2-methoxy-4-(2,7-diazaspiro[3.5]nonan-7-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0157] ##STR00065##
[0158] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with tert-butyl 7-(3-amino-4-methoxyphenyl)-2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester to give the title compound as colorless oil (yield 16%). LCMS (ESI) (5-95AB): m/z: 527.1 [M+1].
EXAMPLE 5
(2-((5-Chloro-2-((2-methoxy-4-(2-methyl-2,7-diazaspiro[3.5]nonan-7-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0159] ##STR00066##
EXAMPLE 5A
1-tert-butyl 4-methyl 4-(chloromethyl)piperidine-1,4-dicarboxylate
[0160] ##STR00067##
[0161] Under atmosphere of N.sub.2, at 78 C., to a solution of 1-tert-butyl-4-methylpiperidine-1,4-dicarboxylate (4.60 g, 18.91 mmol, 1.00 eq.) in THF(100 mL), was added LDA (2M, 18.91 mL) dropwise. After addition, the mixture was stirred for 2 hrs at 78 C. under atmosphere of N.sub.2. At 78 C., chloroiodomethane (10 g, 56.72 mmol) was dropwise added through a syringe into the reaction mixture, then the resultant solution was heated slowly to 20 C. and stirred for 12 hrs. The reaction mixture was quenched with aq. NH4Cl, and ethyl acetate and H.sub.2O were added into the mixture. The organic layer was separated and concentrated, the crude was purified by HPLC (PE:ethyl acetate=100:1 to 25:1) to give the title compound (2.39 g, 8.19 mmol, yield 43.32%) as yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): 3.86 (br. s., 2H) 3.76 (s, 3H) 3.59 (br. s., 2H) 3.00 (br. s., 2H), 2.16 (d, J=13.2 Hz, 2H) 1.45 (m, 9H).
EXAMPLE 5B
tert-Butyl 4-(chloromethyl)-4-(hydroxymethyl)piperidine-1-carboxylic acid tert-butyl ester
[0162] ##STR00068##
[0163] A solution of Example 5A (14.4 g, 49.35 mmol) in anhydrous THF (150 mL) was cooled to 0 C., LAH (2.25 g, 59.22 mmol) was added in portions to the solution, the solution was stirred for 25 minutes at 0 C. TLC showed that the reaction was complete, the mixture was quenched with H.sub.2O (2.25 mL) at 0-10 C., then sodium hydroxide solution (1N, 2.25 mL) was added, the mixture was filtered; the cake was washed with ethyl acetate (30 mL2), the filtrate was washed with H.sub.2O (150 mL) and brine (150 mL), dried over anhydrous sodium sulfate, concentrated to give the title compound (9.79 g, 37.12 mmol, 75.21% yield) as light yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): 3.69-3.56 (m, 4H), 3.47-3.37 (m, 4H), 1.60-1.51 (m, 4H), 1.50-1.43 (m, 9H).
EXAMPLE 5C
tert-Butyl 4-(chloromethyl)-4-formylpiperidine-1-carboxylic acid tert-butyl ester
[0164] ##STR00069##
[0165] At 65 C., dimethyl sulfoxide (6.57 g, 84.15 mmol) in DCM (20 mL) was added to a solution of oxalyl chloride (5.34 g, 42.07 mmol) in DCM (90 mL). Then, Example 5B (9.79 g, 37.12 mmol) in DCM (20 mL) was dropwise added into the above mixture, and the internal temperature was kept below 60 C. The reaction mixture was stirred for 15 minutes at 65 C. to 60 C. At 60 C., triethylamine (18.77 g, 185.49 mmol) was added to the reaction mixture. After the addition, the reaction mixture was heated to rt (20 C.), quenched by saturated aq. Na.sub.2HCO.sub.3 (50 mL). The organic layer was separated, and washed with brine (30 mL), dried over anhydrous sodium sulfate, and purified by column chromatography (DCM:methanol=50:1 to 10:1) to give the title compound (10.50 g, crude) as light yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): 9.59 (s, 1H), 3.92-3.67 (m, 2H), 3.62 (s, 2H), 3.10 (t, J=10.4 Hz, 2H), 2.08 (dt, J=13.6, 4.0 Hz, 2H), 1.60-1.52 (m, 2H), 1.47 (s, 9H).
Example 5D
tert-Butyl 2-methyl-2,7-diazaspiro[3.5]nonane-7-carboxylic acid ethyl ester
[0166] ##STR00070##
[0167] At 15 C., to a mixture of Example 5C (10.50 g, 40.12 mmol) and methylamine hydrochloride (10.51 g, 155.65 mmol) in methanol (100 mL), was added sodium cyanoborohydride (14.50 g, 230.7 mmol), then the mixture was stirred for 16 hrs at 100 C. TLC showed that the reaction was complete. The mixture was concentrated to dry, the resultant residue was diluted by ethyl acetate (100 mL), washed with H.sub.2O (100 mL) and brine (100 mL), dried over anhydrous sodium sulfate and concentrated. The crude was purified by column chromatography (DCM:methanol=100:1 to 10:1) to give the title compound (4.20 g, 17.48 mmol, 43.56% yield) as yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): 3.97 (br. s., 4H), 3.42-3.36 (m, 4H), 2.99 (s, 3H), 1.94-1.84 (m, 4H), 1.45 (s, 9H).
EXAMPLE 5E
2-Methyl-2,7-diazaspiro[3.5]nonane
[0168] ##STR00071##
[0169] At 15 C., a solution of Example 5D (4.20 g, 17.48 mmol) in HCl/methanol (30 mL) was stirred for 0.5 hrs. TLC showed that the reaction was complete. The mixture was concentrated to give the title compound (3.62 g, hydrochloride) as white solid. .sup.1H NMR (400 MHz, CD.sub.3OD): 4.24 (d, J=15.6 Hz, 2H), 4.00 (d, J=15.2 Hz, 2H), 3.29-3.15 (m, 4H), 2.99 (s, 3H), 2.32-2.07 (m, 4H).
EXAMPLE 5F
7-(3-Methoxy-4-nitrophenyl)-2-methyl-2,7-diazaspiro[3.5]nonane
[0170] ##STR00072##
[0171] This Example was prepared according to the process as described in Example 4A, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester was replaced by 2-methyl-2,7-diazaspiro[3.5]nonane to give the title compound as yellow solid, yield 92%. LCMS (ESI) (10-80CD): m/z: 292.2 [M+1].
EXAMPLE 5G
2-Methoxy-4-(2-methyl-2,7-diazaspiro[3.5]nonan-7-yl)aniline
[0172] ##STR00073##
[0173] At 15 C., to a mixture of Example 5F (1.20 g, 4.12 mmol), NH.sub.4Cl (1.40 g, 26.17 mmol) in methanol (20 mL) and DCM (2 mL), zinc powder (2.00 g, 30.59 mmol) was added, then the reaction mixture was stirred for 10 minutes at 35 C. TLC showed that the reaction was complete. The mixture was filtered, the filtrate was concentrated. The resultant residue was dissolved in saturated potassium carbonate solution (20 mL), and extracted with a mixture of DCM and methanol (DCM:methanol=20:1, 20 mL*2). The combined organic layers were dried over anhydrous sodium sulfate and concentrated to give the title compound (920 mg, 3.52 mmol, 85.44% yield) as dark green oil. .sup.1H NMR (400 MHz, CDCl.sub.3): 6.64 (d, J=8.4 Hz, 1H), 6.53 (d, J=2.4 Hz, 1H), 6.42 (dd, J=8.4, 2.4 Hz, 1H), 3.85 (s, 3H), 3.07 (s, 4H), 2.97-2.92 (m, 4H), 2.37 (s, 3H), 1.95-1.85 (m, 4H).
EXAMPLE 5H
(2-((5-Chloro-2-((2-methoxy-4-(2-methyl-2,7-diazaspiro[3.5]nonan-7-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0174] ##STR00074##
[0175] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 2-methoxy-4-(2-methyl-2,7-diazaspiro[3.5]nonan-7-yl)aniline to give the title compound as yellow solid, yield 23%. .sup.1H NMR (400 MHz, CD.sub.3OD): 8.22 (s, 2H), 7.87-7.61 (m, 3H), 7.57-7.45 (m, 1H), 7.38 (br. s., 1H), 7.12 (d, J=8.0 Hz, 1H), 4.34 (d, J=10.4 Hz, 2H), 4.06 (d, J=11.2 Hz, 2H), 3.99 (s, 3H), 3.74-3.49 (m, 4H), 3.03 (s, 3H), 2.51-2.30 (m, 4H), 1.89 (d, J=13.6 Hz, 6H); LCMS (ESI) (0-60AB):m/z: 541.2 [M+1].
EXAMPLE 6
(2-((5-Chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0176] ##STR00075##
EXAMPLE 6A
tert-Butyl 6-(4-methoxy-3-nitrophenyl)-2,6-diazaspiro[3.4]octane-2-carboxylic acid tert-butyl ester
[0177] ##STR00076##
[0178] This Example was prepared according to the process as described in Example 4A, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester was replaced with tert-butyl 2,6-diazaspiro[3.4]octane-2-carboxylic acid tert-carboxylic ester to give the title compound as colorless oil, yield 92%. LCMS (ESI) (5-95AB): m/z: 364.1 [M+1].
EXAMPLE 6B
tert-Butyl 6-(3-amino-4-methoxyphenyl)-2,6-diazaspiro[3.4]octane-2-carboxylic acid tert-butyl ester
[0179] ##STR00077##
[0180] This Example was prepared according to the process as described in Example 3F, 8-(3-methoxy-4-nitrophenyl)-2,8-diazaspiro[4.5]decan-3-one was replaced with tert-butyl 6-(4-methoxy-3-nitrophenyl)-2,6-diazaspiro[3.4]octane-2-carboxylic acid tert-butyl ester to give the title compound as green oil (crude). LCMS (ESI) (5-95AB): m/z: 334.2 [M+1].
EXAMPLE 6C
[0181] (2-((5-Chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
##STR00078##
[0182] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced by tert-butyl 6-(3-amino-4-methoxyphenyl)-2,6-diazaspiro[3.4]octane-2-carboxylic acid tert-butyl ester to give the title compound as colorless oil, yield 19%. LCMS (ESI) (5-95AB): m/z: 513.0 [M+1].
EXAMPLE 7
(2-((5-Chloro-2-((2-methoxy-4-(2-methyl-2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0183] ##STR00079##
[0184] To a solution of Example 6C (50 mg, 0.039 mmol) in THF (5 mL) was added aq. formaldehyde (9.5 mg, 37%, 0.117 mmol); the reaction mixture was stir for 30 minutes at 16 C., then into the reaction mixture was added sodium triacetoxyborohydride (25 mg,0.117 mmol), stirred for 12 hrs at 16 C. LCMS showed that the reaction was complete. The reaction mixture was filtered, purified by pre-HPLC to give the title compound (9.6 mg, yield 46%) as white solid. LCMS (ESI) (5-95AB): m/z: 527.2 [M+1].
EXAMPLE 8
(2-((5-Chloro-2-((2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0185] ##STR00080##
EXAMPLE 8A
9-Benzyl-2,4-dioxo-3,9-diazaspiro[5.5]undecane-1,5-dicarbonitrile
[0186] ##STR00081##
[0187] At 5-8 C., ammonium acetate (2.04 g, 26.42 mmol, 0.10 eq) was added to cyanacetate (90 g, 796 mmol, 3.00 eq.) in methanol (100 mL); then 1-benzyl-4-piperidone (50 g, 0.264 mol) was add into the reaction mixture; at 10 C., aq. ammonia (46.3 g, 370 mmol, 1.40 eq) was added into the reaction mixture, the mixture was stirred for 1 hrs at 0-5 C. The reaction mixture was warmed to 20 C. (rt.), and stirred for 20 hrs. LCMS showed that the product was generated. H.sub.2O (100 mL) was added to the mixture, and heated to 55 C. The conc. hydrochloric acid (12M) was added to adjust pH to 4 while temperature was kept no more than 70 C. The reaction was cooled to 10 C., stirred for 30 minutes and filtered. The cake was washed with H.sub.2O, dried in the air to give the title compound (66 g, 77% yield) as white solid. LCMS (ESI) (0-30AB): m/z: 323.0 [M+1].
EXAMPLE 8B
1-Benzyl-3,7-diazaspiro[bicyclo[3.3.1]nonane-9,4-piperidine]-2,4,6,8-tetraone
[0188] ##STR00082##
[0189] A mixture of Example 8A (1.00 g, 3.10 mmol, 1.00 eq) in sulfuric acid (88%, 4 mL) was stirred for 4 hrs at 60 C. To the reaction mixture was added H.sub.2O (1.4 mL), heated to 100 C. and stirred for 1 hr. To the reaction mixture was added H.sub.2O (5 mL) in again, then cooled to 10 C., stirred for 30 minutes at 10 C., and then filtered. The cake was washed with cool H.sub.2O (5 mL) and dried to give the title compound (1.11 g, crude) as white solid. .sup.1H NMR (400MHz, CDCl.sub.3): 11.87 (s, 2H), 9.56 (br. s., 1H), 7.47 (s, 5H), 4.34 (d, J=4.4 Hz, 2H), 3.75 (br. s., 1H), 3.22 (br. s., 4H), 1.88 (br. s., 4H).
EXAMPLE 8C
9-Benzyl-3,9-diazaspiro[5.5]undecane-2,4-dione
[0190] ##STR00083##
[0191] At 15 C., to a flask containing Example 8B (1.10 g, 3.22 mmol, 1.00 eq) was added aq. NaOH (5N,5 mL), the mixture was heated to 70 C. and stirred 4 hrs. The mixture was cooled to 45 C., conc. hydrochloric acid (12 N, 1.5 mL) was slowly added until pH of the solution reached around 7. The mixture was further heated to70-75 C., then conc. HCl (12 N,1 mL) was dropwise added to control the rate of the release of carbon dioxide until pH reached around 3-4. The mixture was still further heated to 70-75 C. and reacted for 1 hr. The resultant suspension was cooled to 10 C. and stirred for 0.5 hrs. The solid was filtered and washed with H.sub.2O (25 mL). The solid was dried to give the title compound (380 mg, 1.40 mmol, yield for 2 steps 45%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): 10.89 (s, 1H), 10.60 (br. s, 1H), 7.57 (br. s., 2H), 7.44 (br. s., 3H), 4.28 (br. s., 2H), 3.11 (br. s., 4H), 2.76 (br. s., 2H), 2.42 (br. s., 2H), 2.00-1.48 (m, 4H).
EXAMPLE 8D
3-Benzyl-3,9-diazaspiro[5.5]undecane
[0192] ##STR00084##
[0193] At 0-10 C., to a solution of Example 8C (10.6 g, 38.92 mmol, 1.00 eq) in THF (120 mL) was added LAH (5.17 g, 136.22 mmol, 3.50 eq), then the mixture was stirred for 3 hrs at 65 C. TLC showed that the reaction was complete. The mixture was cooled to 10 C., and quenched with H.sub.2O (5.2 mL), aq. sodium hydroxide (1N, 5.2 mL) was then added. The mixture was filtered, the filtrate was concentrated to give the title compound (7.40 g, 30.28 mmol, 77.81% yield) as light yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): 7.34-7.27 (m, 5H), 3.52 (s, 2H), 2.88-2.73 (m, 4H), 2.46-2.35 (m, 4H), 1.58-1.51 (m, 4H), 1.48-1.39 (m, 4H).
EXAMPLE 8E
tert-Butyl 3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0194] ##STR00085##
[0195] To a mixture of Example 8D (500 mg, 2.05 mmol) and Boc.sub.2O (450 mg, 2.06 mmol) in methanol was added triethylamine (311 mg, 3.08 mmol), then stirred for 16 hrs at 20-30 C. The reaction mixture was concentrated, the residue was diluted with ethyl acetate (20 mL), and washed with H.sub.2O (15 mL2) and brine (20 mL), the organic layer was dried over anhydrous sodium sulfate, concentrated. The crude intermediate was dissolved in ethanol (15 mL) and acetic acid (2 mL), Pd(OH).sub.2/C (0.1 g) was then added and the mixture was reacted under H.sub.2 stomsphere (50 Psi) for 20 hrs. The mixture was filtered, the filtrate was concentrated to give the acetate of the title compound (320 mg, 1.26 mmol, 61.37% yield).
EXAMPLE 8F
tert-Butyl 9-(3-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0196] ##STR00086##
[0197] This Example was prepared according to the process as described in Example 4A, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester was replaced with tert-butyl 3,9-diazaspiro[5.5]undecane-3-carboxylic acid tert-butyl ester to give the title compound as yellow oil, yield 62%. .sup.1H NMR (400 MHz, CD.sub.3OD): , 7.95 (d, J=9.2 Hz, 1H), 6.55 (dd, J=9.6, 2.4 Hz, 1H), 6.48 (d, J=2.4 Hz, 1H), 3.95 (s, 3H), 3.56-3.41 (m, 8H), 1.72-1.65 (m, 4H), 1.58-1.50 (m, 4H), 1.48 (s, 9H).
EXAMPLE 8G
tert-Butyl 9-(4-amino-3-methoxyphenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0198] ##STR00087##
[0199] This Example was prepared according to the process as described in Example 3F, 8-(3-methoxy-4-nitrophenyl)-2,8-diazaspiro[4.5]decan-3-one was replaced by tert-butyl 9-(3-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate to give the title compound as brown oil, yield 36%. LCMS (ESI) (5-95AB): m/z: 376.2 [M+1].
EXAMPLE 8H
(2-((5-Chloro-2-((2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0200] ##STR00088##
[0201] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replace with tert-butyl 9-(4-amino-3-methoxyphenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester to give the title compound as yellow solid, yield 21%. LCMS (ESI) (5-95AB): m/z: 555.2 [M+1].
EXAMPLE 9
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0202] ##STR00089##
[0203] This Example was prepared according to the process as described in Example 7, take the place of (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((5-chloro-2-((2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as yellow solid, yield 57%. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.28 (s, 1H), 8.12 (br. s., 1H), 7.81-7.68 (m, 3H), 7.65 (d, J=2.0 Hz, 1H), 7.56-7.49 (m, 1H), 7.37 (d, J=8.8 Hz, 1H), 4.02 (s, 3H), 3.74 (br. s., 4H), 3.47 (d, J=12.8 Hz, 2H), 3.24 (t, J=12.8 Hz, 2H), 2.93 (s, 3H), 2.42-1.97 (m, 5H), 1.93-1.75 (m, 9H). LCMS (ESI) (0-60AB): miz: 569.2 [M+1].
EXAMPLE 10
(2-((5-Chloro-2-((4-(9-isopropyl-3,9-diazaspiro[5.5]undecan-3-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0204] ##STR00090##
[0205] A mixture of Example 8C (100 mg, 0.18 mmol), sodium triacetoxyborohydride (114 mg, 0.54 mmol), acetic acid (21.6 mg, 0.36 mmol) and acetone (20.9 mg, 0.36 mmol) in THF (5.0 mL) was stirred for 16 hrs at 18 C. LCMS showed that the reaction was complete, and the mixture was concentrated in vacuum to remove the solvent to give the crude, the crude was purified by pre-HPLC to give the title compound (50 mg, yield 47%) as brown solid. LCMS (ESI) (10-80AB): m/z: 597.3 [M+1].
EXAMPLE 11
(2-((5-Chloro-2-((2-(difluoromethoxy)-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0206] ##STR00091##
EXAMPLE 11A
5-Fluoro-2-nitrophenol
[0207] ##STR00092##
[0208] At 0 C., to a stirred solution of 4-fluoro-2-methoxy nitrobenzene (3 g, 17.53 mmol) in DCM (30 mL) was dropwise added boron tribromide. The reaction was stirred for 1.5 hrs at 0 C. TLC (PE: ethyl acetate=10:1) showed that 4-fluoro-2-methoxy nitrobenzene disappeared. The solution was slowly added into ice H.sub.2O (100 mL), and extracted with DCM (50 mL3). The organic layer was dried and concentrated to give the title compound (2.5 g, yield 90.8%) as yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 10.80 (s, 1H), 8.16 (dd, J=9.6, 5.6 Hz, 1H), 6.84 (dd, J=9.6, 2.4 Hz, 1H), 6.77-6.67 (m, 1H).
EXAMPLE 11B
2-(Difluoromethoxy)-4-fluoro-1-nitrobenzene
[0209] ##STR00093##
[0210] Under continuing stirring, to a solution of Example 11A (2.0 g, 12.73 mmol) in DMF (20 mL) were added ClCF.sub.2COONa (6.9 g, 44.56 mmol) and sodium carbonate (1.62 g, 15.28 mmol). The mixture was heated to 90 C. and stirred for 16 hrs. TLC (PE:ethyl acetate=10:1) showed that 5-fluoro-2-nitro phenol disappeared. The reaction mixture was diluted with ethyl acetate (100 mL), and washed with H.sub.2O (20 mL2). The organic layer was dried and concentrated to give the crude, the crude was purified by silica gel column chromatography (PE:ethyl acetate=10:1) to give the title compound (1.4 g, yield 53.1%) as yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 8.07-7.98 (m, 1H), 7.17-7.05 (m, 2H), 6.65 (t, J=72.0 Hz, 1H).
EXAMPLE 11C
tert-Butyl 9-(3-(difluoromethoxy)-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0211] ##STR00094##
[0212] This Example was prepared according to the process as described in Example 4A, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester was replaced with tert-butyl 3,9-diazaspiro[5.5]undecane-3-carboxylic acid tert-butyl ester, and 4-fluoro-2-methoxyl-1-nitrobenzene with 2-(difluoromethoxy)-4-fluoro-1-nitrobenzene to give the title compound as yellow oil, yield 77%.
EXAMPLE 11D
tert-Butyl 9-(4-amino-3-(difluoromethoxy)phenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0213] ##STR00095##
[0214] This Example was prepared according to the process as described in Example 3F, 8-(3-methoxy-4-nitrophenyl)-2,8-diazaspiro[4.5]decan-3-one was replaced with tert-butyl 9-(3-(difluoromethoxy)-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid ethyl ester to give the title compound as green solid, yield 95%. LCMS (ESI) (5-95AB): m/z: 412.2 [M+1].
EXAMPLE 11E
(2-((5-Chloro-2-((2-(difluoromethoxy)-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0215] ##STR00096##
[0216] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with tert-butyl 9-(4-amino-3-(difluoromethoxy)phenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester to give the title compound as white solid, yield 22%. LCMS (ESI) (0-60AB): m/z: 591.2 [M+1].
EXAMPLE 11F
(2-((5-Chloro-2-((2-(difluoromethoxy)-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenypamino)pyrimidin-4-yl)amino)phenyl)dimethylphosphine oxide
[0217] ##STR00097##
[0218] This Example was prepared according to the process as described in Example 7, (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((5-chloro-2-((2-(difluoromethoxy)-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as colorless oil, yield 22%. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.31 (s, 1H), 8.12 (m, 1H), 7.86-7.84 (m, 1H), 7.79-7.65 (m, 3H), 7.62-7.55 (m, 1H), 7.54-7.48 (m, 1H), 7.08 (t, J=72.0 Hz, 1H), 3.75-3.62 (m, 4H), 3.52-3.45 (m, 2H), 3.29-3.21 (m, 2H), 2.94 (s, 3H), 2.34-2.13 (m, 4H), 2.11-1.98 (m, 2H), 1.89 (d, J=13.6 Hz, 6H), 1.88-1.79 (m, 2H). LCMS (ESI) (5-95 AB): m/z: 605.2 [M+1].
EXAMPLE 12
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine sulfide
[0219] ##STR00098##
EXAMPLE 12A
(2-Aminophenyl)dimethyl sulfide
[0220] ##STR00099##
[0221] (2-Aminophenyl)dimethyl phosphine oxide (1.00 g, 5.91 mmol) and Lawesson's reagent (4.78 g, 11.82 mmol) was placed in toluene (60 mL), the reaction mixture was heated to 110 C. and stirred for 4 hrs. TLC (PE:ethyl acetate=3:1) showed that the reaction was complete. The mixture was filtered and concentrated, the crude was purified by pre-HPLC (basic) to give the title compound (600 mg, 3.24 mmol, yield 54.81%) as colorless oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.31-7.27 (m, 1H), 7.22-7.15 (m, 1H), 6.81-6.75 (m, 1H), 6.68-6.60 (m, 1H), 2.08 (s, 3H), 2.05 (s, 3H)
EXAMPLE 12B
(2-((2,5-Dichloropyrimidin-4-yl)amino)phenyl)dimethyl sulfide
[0222] ##STR00100##
[0223] Example 12A (50.0 mg, 269.94 umol), 2,4,5-trichloropyrimidine (148.54 mg, 809.82 umol) and potassium carbonate (111.93 mg, 809.82 umol) were placed in DMF (3 mL), the mixture was stirred for 12 hrs at 60 C. TLC (PE:ethyl acetate=3:1) showed that the product was generated. To the reaction mixture was added H.sub.2O (10 mL), and extracted with ethyl acetate (10 mL3). The combined organic layers were dried and concentrated to obtain yellow oil. The oil was separated and purified by pre-TLC (PE:ethyl acetate=3:1) to give the title compound (25.00 mg, 75.2612 mol, 27.88% yield) as yellow solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 10.12 (br. s., 1H), 8.28 (s, 1H), 8.10 (dd, J=8.0, 4.8 Hz, 1H), 7.61 (dd, J=8.0, 8.0 Hz, 1H), 7.46 (dd, J=13.8, 7.6 Hz, 1H), 7.33-7.28 (m, 1H), 2.07 (s, 3H), 2.04 (s, 3H).
EXAMPLE 12C
3-(3-Methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane
[0224] ##STR00101##
[0225] A solution of Example 8A (0.7 g,1.73 mmol) in trifluoroacetic acid (4 mL) and DCM (4 mL) was stirred for 1 hr at 16 C. LCMS showed that the reaction was complete. To the mixture was added aq. sodium carbonate (50 mL), and extracted with DCM (50 mL2). The organic layer was dried over anhydrous sodium sulfate, and filtered and concentrated to give the title compound (0.7 g, crude) as yellow solid. LCMS (ESI) (0-60AB): m/z: 306.0 [M+1].
EXAMPLE 12D
3-(3-Methoxy-4-nitrophenyl)-9-methyl-3,9-diazaspiro[5.5]undecane
[0226] ##STR00102##
[0227] At 16 C., to a solution of Example 12C (0.7 g, 2.3 mmol) in THF (10 mL) was added aq. formaldehyde (207 mg, 6.9 mmol, 37%), and stirred for 0.5 hrs at 16 C. Sodium triacetoxyborohydride (1.5 g, 6.9 mmol) was added, the mixture was stirred for 12 hrs at 16 C. LCMS showed that the reaction was complete. The mixture was diluted with DCM (60 mL), filtered and concentrated to give the title compound (0.6 g, yield 82%) as yellow oil. LCMS (ESI) (0-60AB): m/z: 320.2 [M+1].
EXAMPLE 12E
2-Methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline
[0228] ##STR00103##
[0229] To a solution of Example 12D (0.6 g, 1.9 mmol) in EtOH/H.sub.2O(12 mL) were added Fe powder (1.1 g, 18.8 mmol) and NH.sub.4Cl (1.1 g, 18.8 mmol). The mixture was stirred for 2 hrs at 80 C. TLC (DCM:methanol=6:1) showed that the reaction was complete. The reaction mixture was filtered and concentrated. The resultant residue was purified by pre-TLC (DCM:methanol=6:1) to give the title compound (400 mg, yield 74%) as green solid. .sup.1H NMR (400 MHz, CD.sub.3OD): , 6.72 (d, J=8.4 Hz, 1H), 6.64 (d, J=2.0 Hz, 1H), 6.51 (d, J=8.4 Hz, 1H), 3.86 (s, 3H), 3.08-2.96 (m, 4H), 2.58-2.45 (m, 4H), 2.32 (s, 3H), 1.75-1.52 (m, 8H).
EXAMPLE 12F
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl sulfide
[0230] ##STR00104##
[0231] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline, and (2-((2,5-dichloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((2,5-dichloropyrimidin-4-yl)amino)phenyl)dimethyl sulfide to give the title compound as brown solid, yield 36%. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.35 (s, 1H), 7.89 (dd, J=13.6, 8.0 Hz, 1H), 7.75-7.80 (m, 2H), 7.71 (d, J=8.8 Hz, 1H), 7.57-7.66 (m, 2H), 7.22 (d, J=8.4 Hz, 1H), 4.02 (s, 3H), 3.69 (br. s., 4H), 3.47 (d, J=12.8 Hz, 2H), 3.18-3.29 (m, 2H), 2.92 (s, 3H), 2.19-2.38 (m, 2H), 2.15-1.94 (m, 10H), 1.75-1.91 (m, 2H). LCMS (ESI) (0-60AB): m/z: 585.2 [M+1].
##STR00105##
EXAMPLE 13
(2-((5-Chloro-2-((4-(2-(dimethylamino)-7-azaspiro[3.5]nonan-7-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0232] ##STR00106##
EXAMPLE 13A
tert-Butyl 2-(dimethylamino)-7-azaspiro[3.5]nonane-7-carboxylic acid ethyl ester
[0233] ##STR00107##
[0234] At rt., under N.sub.2 atomsphere, to a mixture of tert-butyl-2-oxo-7-azaspiro[3.5]nonane-7-carboxylic acid ethyl ester (200 mg, 0.835 mmol) in MeOH (10 mL) were added dimethylamine hydrochloride (340.73 mg, 4.18 mmol) and triethylamine (507.41 mg, 5.01 mmol). The reaction mixture was stirred for 30 minutes at rt., then sodium triacetoxyborohydride (531.38 mg, 2.51 mmol) was added. The reaction mixture was stirred for 6 hrs at 30 C. LCMS showed that the reaction was complete. The mixture was concentrated in vacuum to give the residue, the residue was diluted with H.sub.2O (30 mL) and extracted with DCM (50 mL2). The organic layer was washed with brine (50 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum to give the title compound (200 mg, yield 89.16%) as white solid. LCMS (ESI) (5-95AB): m/z: 269.3 [M+1].
EXAMPLE 13B
N,N-Dimethyl-7-azaspiro[3.5]nonan-2-amine
[0235] ##STR00108##
[0236] A mixture of Example 13A (400.00 mg, 1.49 mmol) in TFA (10 mL) was stirred for 12 hrs at 30 C. TLC showed that the reaction was complete. The mixture was concentrated in vacuum to give the title compound (251 mg, crude) as yellow oil.
EXAMPLE 13C
7-(3-Methoxy-4-nitrophenyl)-N,N-dimethyl-7-azaspiro[3.5]nonan-2-amine
[0237] ##STR00109##
[0238] Under N.sub.2 stomsphere, to a solution of Example 13B (251.00 mg, 1.49 mmol) and 4-fluoro-2-methoxyl-1-nitrobenzene (305.98 mg, 1.79 mmol) in dimethyl sulfoxide (10 mL) was added potassium carbonate (617.80 mg, 4.47 mmol), the mixture was stirred for 12 hrs at 90 C. TLC showed that the reaction was complete. The mixture was poured into H.sub.2O (25 mL) and extracted with DCM (50 mL2). The organic layer was washed with brine (50 ml2), dried over anhydrous sodium sulfate, filtered and concentrated to give the residue, the residue was purified by silica gel column chromatography (DCM:methanol=80:1, 60:1) to give the title compound (310 mg, yield 65.14%) as yellow solid. LCMS (ESI) (5-95AB): m/z: 320.3 [M+1].
EXAMPLE 13D
7-(4-Amino-3-methoxyphenyl)-N,N-dimethyl-7-azaspiro[3.5]nonan-2-amine
[0239] ##STR00110##
[0240] To a solution of Example 13C (310 mg, 0.97 mmol) in methanol (10 mL) were added Zn powder (317.33 mg, 4.85 mmol) and NH.sub.4Cl (207.66 mg, 3.88 mmol). The suspension was stirred for 0.5 hrs at 30 C. TLC showed that the starting material was completely consumed. The reaction mixture was filtered, concentrated to give the title compound (185 mg, yield 65.86%) as white solid.
EXAMPLE 13E
(2-((5-Chloro-2-((4-(2-(dimethylamino)-7-azaspiro[3.5]nonan-7-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0241] ##STR00111##
[0242] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 7-(4-amino-3-methoxyphenyl)-N,N-dimethyl-7-azaspiro[3.5]nonan-2-amine to give the title compound as white solid, yield 16%. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.29 (s, 1H), 8.14 (br. s, 1H), 7.68-7.82 (m, 3H), 7.60 (d, J=2.0 Hz, 1H), 7.54 (t, J=6.8 Hz, 1H), 7.31 (dd, J=8.8, 2.0 Hz, 1H), 4.04 (s, 3H), 3.85 (t, J=8.4 Hz, 1H), 3.65-3.77 (m, 4H), 2.86 (s, 6H), 2.60 (br. s., 2H), 2.22-2.34 (m, 6H), 1.85-1.92 (m, 6H). LCMS (ESI) (5-95AB): m/z: 569.2 [M+1].
EXAMPLE 14
(2-((5-Chloro-2-((4-(9-(dimethylamino)-3-azaspiro[5.5]undecan-3-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0243] ##STR00112##
EXAMPLE 14A
tert-butyl 10-(dimethylamino)-3-azaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0244] ##STR00113##
[0245] This Example was prepared according to the process as described in Example 13A, tert-butyl-2-oxo-7-azaspiro[3.5]nonane-7-carboxylic acid ethyl ester was replaced with tert-butyl 9-oxo-3-azaspiro[5.5]undecane-3-carboxylic acid methyl ester to give the title compound as yellow oil, yield 79.37%.
EXAMPLE 14B
N,N-Dimethyl-3-azaspiro[5.5]undecan-9-amine
[0246] ##STR00114##
[0247] This Example was prepared according to the process as described in Example 13B, tert-butyl 2-(dimethylamino)-7-azaspiro[3.5]nonane-7-carboxylic acid ethyl ester was replaced with tert-butyl 10-(dimethylamino)-3-azaspiro[5.5]undecane-3-carboxylic acid methyl ester to give the title compound as yellow oil (crude), which was used directly in the next step without purification.
EXAMPLE 14C
3-(3-Methoxy-4-nitrophenyl)-N,N-dimethyl-3-azaspiro[5.5]undecan-9-amine
[0248] ##STR00115##
[0249] This Example was prepared according to the process as described in Example 13C, N,N-dimethyl-7-azaspiro[3.5]nonan-2-amine was replaced with N,N-dimethyl-3-azaspiro[5.5]undecan-9-amine to give the title compound as dark solid, yield 55%. LCMS (ESI) (5-95AB): m/z: 348.1[M+1].
EXAMPLE 14D
3-(4-Amino-3-methoxyphenyl)-N,N-dimethyl-3-azaspiro[5.5]undecan-9-amine
[0250] ##STR00116##
[0251] This Example was prepared according to the process as described in Example 13D, 7-(3-methoxy-4-nitrophenyl)-N,N-dimethyl-7-azaspiro[3.5]nonan-2-amine was replaced with 3-(3-methoxy-4-nitrophenyl)-N,N-dimethyl-3-azaspiro[5.5]undecan-9-amine to give the title compound as dark solid, yield 82%. LCMS (ESI) (5-95AB): nth: 318.2[M+1].
EXAMPLE 14E
(2-((5-Chloro-2-((4-(9-(dimethylamino)-3-azaspiro[5.5]undecan-3-yl)-2-methoxyphenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0252] ##STR00117##
[0253] This Example was prepared according to the process as described in Example 1M, take the place of 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 3-(4-amino-3-methoxyphenyl)-N,N-dimethyl-3-azaspiro[5.5]undecan-9-amine to give the title compound as yellow solid, yield 11%. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.29 (s, 1H), 8.11-8.20 (m, 1H), 7.70-7.80 (m, 3H), 7.64 (d, J=2.8 Hz, 1H), 7.52-7.56 (m, 1H), 7.36 (dd, J=8.8, 2.0 Hz, 1H), 4.04 (s, 3H), 3.58-3.86 (m, 4H), 3.26-3.29 (m, 1H), 2.91 (s, 6H), 1.97-2.22 (m 8H), 1.91 (s, 3H), 1.88 (s, 3H), 1.73-1.82 (m, 2H), 1.44-1.56 (m, 2H). LCMS (ESI)(0-60AB): m/z: 597.3[M+1].
##STR00118##
EXAMPLE 15
(2-((2-((2-Methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-5-(trifluoromethyl)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0254] ##STR00119##
EXAMPLE 15A
(2-((2-Chloro-5-(trifluoromethyl)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0255] ##STR00120##
[0256] This Example was prepared according to the process as described in Example 1L, 2,4,5-trichloropyrimidine was replaced with 2,4-dichloro-5-(trifluoromethyl)pyrimidine to give the title compound as white solid, yield 23%. LCMS (ESI) (5-95AB): m/z: 350.0 [M+1].
EXAMPLE 15B
2-Methoxy-5 (6-methyl-6-diazaspiro[3.4]octa-2-yl)aniline and 2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline
[0257] ##STR00121##
[0258] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 2-methoxy-4-(9-methyl-3-1,9-diazaspiro[5.5]undecan-3-yl)aniline, and (2-((2,5-dichloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide with (2-((2-chloro-5-(trifluoromethyl)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as colorless oil, yield 20%. LCMS (ESI) (5-95AB): m/z: 603.3 [M+1].
EXAMPLE 16
(2-((4-((2-Methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-1,3,5-triazin-2-yl)amino)phenyl)dimethyl phosphine oxide
[0259] ##STR00122##
EXAMPLE 16A
(2-((4-Chloro-1,3,5-triazin-2-yl)amino)phenyl)dimethyl phosphine oxide
[0260] ##STR00123##
[0261] To a solution of 2,4-dichloro-1,3,5-triazine (50 mg, 0.33 mmol) in MeCN (3.0 mL), were added DIPEA (52 mg, 0.40 mmol) and (2-aminophenyl) dimethyl phosphine oxide (85 mg, 0.50 mmol). The reaction mixture was stirred for 2 hrs at 20 C. TLC (DCM:methanol=20:1) showed that the reaction was complete. The mixture was concentrated, the resultant residue was diluted with H.sub.2O (10 mL), then acidified with aq. HCl solution (1N) to adjust pH to 7, and extracted with DCM (20 mL2). The organic layer was dried over anhydrous sodium sulfate and concentrated to give the title compound (50 mg, yield 53.05%) as yellow oil. LCMS (ESI) (0-30AB): m/z: 282.9 [M+1].
EXAMPLE 16B
tert-Butyl 9-(4-((4-((2-(dimethylphosphoryl)phenyl)amino)-1,3,5-triazin-2-yl)amino)-3-methoxyphenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0262] ##STR00124##
[0263] To a solution of Example 16A (40 mg, 0.14 mmol) in MeCN (3.0 mL) was added DIPEA (27 mg, 0.21 mmol) and Example 8B (53 mg, 0.14 mmol). The mixture was stirred for 2 hrs at 20 C. TLC (DCM:methanol=20:1) showed that the reaction was complete. The mixture was concentrated, the resultant residue was diluted with H.sub.2O (10 mL), then adjusted pH to 7 with aq. HCl solution (1N), and extracted with DCM (20 mL2). The organic layer was dried and concentrated to give the title compound (50 mg, yield 56.8%) as yellow oil. LCMS (ESI) (5-95AB): m/z: 622.3 [M+1].
EXAMPLE 16C
(2-((4-((2-Methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-1,3,5-triazin-2-yl)amino)phenyl)dimethyl phosphine oxide
[0264] ##STR00125##
[0265] Example 16B (50 mg,0.08 mmol) was dissolved in TFA (3.0 mL), the reaction mixture was stirred for 2 hrs at 20 C. The solution was dissolved in DCM (5 mL) and concentrated to dry to give the title compound (40 mg, crude) as yellow oil.
EXAMPLE 16D
(2-((4-((2-Methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-1,3,5-triazin-2-yl)amino)phenyl)dimethyl phosphine oxide
[0266] ##STR00126##
[0267] This Example was prepared according to the process as described in Example 7, (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((4-((2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-1,3,5-triazin-2-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as white solid, yield 15%. LCMS (ESI) (0-60AB): m/z: 536.3 [M+1].
EXAMPLE 17
(2-((2-(amino)-7H-pyrrolo[2,3-D](2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0268] ##STR00127##
EXAMPLE 17A
(2-((2-Chloro-7-((2-(trimethylsilypethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0269] ##STR00128##
[0270] At 0 C., to a solution of Example 1K (300 mg, 1.77 mmol) in DMF (5.0 mL) was added NaH (106 mg, 2.66 mmol), then 2,4-dichloro-7-((2-(trimethylsilypethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidine (677 mg, 2.13 mmol) was added in portions into the reaction mixture, the reaction mixture was stirred for 5 hrs at 20 C. TLC (methanol:DCM=20:1) showed that the reaction was complete. The mixture was diluted with EtOAc (20 mL), and washed with H.sub.2O (10 mL). The organic layer was dried and concentrated. The crude was purified by pre-TLC (DCM:methanol=10:1) to give the title compound (270 mg, yield 33.8%) as yellow oil. LCMS (ESI) (5-95AB): m/z: 451.2 [M+1].
EXAMPLE 17B
(2-((2-((2-Methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-7-((2-(trimethylsilypethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0271] ##STR00129##
[0272] To a solution of Example 17A (187 mg, 0.41 mmol) in dioxane (5.0 mL) were added Example 12E (100 mg, 0.34 mmol), Xphos (32 mg, 0.07 mmol), Pd.sub.2(dba).sub.3 (32 mg, 0.03 mmol) and potassium carbonate (95 mg, 0.69 mmol). The reaction mixture was stirred for 16 hrs at 90 C. TLC (PE:ethyl acetate=3:1) showed that reaction was complete. The suspension was diluted with ethyl acetate (20 mL) and filtered. The organic layer was washed with H.sub.2O (50 mL) then dried, concentrated. The crude was purified by pre-TLC (DCM:methanol=10:1) to give the title compound (100 mg, yield 41.1%) as yellow oil. LCMS (ESI) (5-95AB): nilz: 704.2 [M+1].
EXAMPLE 17C
(2-((2-(amino)-7H-pyrrolo[2,3-D](2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl) phenyl)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0273] ##STR00130##
[0274] Example 17B (100 mg, 0.14 mmol) was dissolved in TFA (3.0 mL), the reaction mixture was stirred for 30minutes at 20 C. TLC (DCM:methanol=10:1) showed that the starting material disappeared. The reaction mixture was diluted with DCM (10 mL) and concentrated to give the intermediate (59 mg, crude); the crude intermediate was dissolved in methanol (5.0 mL), then NaOH was added to adjust pH to 12, and the mixture was stirred for 30minutes at 20 C. LCMS showed that the product was generated. The reaction mixture was neutralized with aq. HCl solution (1N), and purified by pre-HPLC to give the title compound (10.58 mg, 12.98%) as white solid. LCMS (ESI) (0-60AB): adz: 574.3 [M+1].
##STR00131##
EXAMPLE 18
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)(methyl)amino)phenyl)dimethyl phosphine oxide
[0275] ##STR00132##
EXAMPLE 18A
(2-((2,5-Dichloropyrimidin-4-yl)(methyl)amino)phenyl)dimethyl phosphine oxide
[0276] ##STR00133##
[0277] To a solution of Example 1L (100 mg, 0.32 mmol), K.sub.2CO.sub.3 (87.44 mg, 0.63 mmol) in MeCN (2 mL) was added Mel (0.1 mL, 1.58 mmol). The reaction mixture was stirred for 8 hrs at 70 C. TLC (DCM:methanol=20:1) showed that the reaction was complete. To the mixture was added ethyl acetate (50 mL) and washed with brine (30 mL), dried and concentratedto give yellow oil. The residue was purified by pre-TLC (DCM:methanol=20:1) to give the title compound (50 mg, yield 47.88%) as yellow oil. .sup.1H NMR (400 MHz, CDCl3): , 8.05 (s, 1H), 7.90-7.80 (m, 1H), 7.60-7.48 (m, 2H), 7.14-7.20 (m, 1H), 3.55 (s, 3H), 1.79 (dd, J=19.2, 12.8 Hz, 6H).
EXAMPLE 18B
(2-((5-Chloro-2-((2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)(methyl)amino)phenyl)dimethyl phosphine oxide
[0278] ##STR00134##
[0279] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with tert-butyl 3,9-diazaspiro[5.5]undecan-3-carboxylic acid methyl ester, and (2-((2,5-dichloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide with (2((2,5-dichloropyrimidin-4-yl)(methyl)amino)phenyl)dimethyl phosphine oxide to give the title compound as yellow oil, yield 33%. LCMS (ESI) (0-60AB): m/z: 569.2 [M+1].
EXAMPLE 18C
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)(methyl)amino)phenyl)dimethyl phosphine oxide
[0280] ##STR00135##
[0281] This Example was prepared according to the process as described in Example 7, (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((5-chloro-2-((2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)(methyl)amino)phenyl)dimethyl phosphine oxide to give the title compound as white solid, yield 32%. LCMS (ESI) (0-60AB): miz: 583.2 [M+1].
EXAMPLE 19
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)oxy)phenyl)dimethyl phosphine oxide
[0282] ##STR00136##
[0283] A mixture of Example 12E (60 mg, 0.2 mmol), (2-((2,5-dichloropyrimidin-4-yl)oxy)phenyl) dimethyl phosphine oxide (66 mg, 0.2 mmol), Xantphos (12 mg, 0.02 mmol), palladium acetate (5 mg, 0.02 mmol), CsCO.sub.3(205 mg, 0.6 mmol) in dioxane (10 mL) was heated to 100 C. under N.sub.2 atomphere and stirred for 12 hrs. LCMS showed that the reaction was complete. The reaction mixture was concentrated, and purified by pre-HPLC to give the title compound (10.78 mg, yield 9%) as white solid. LCMS (ESI) (5-95AB): m/z: 570.2 [M+1].
##STR00137##
EXAMPLE 20
(2-((5-Chloro-2-((5-fluoro-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0284] ##STR00138##
EXAMPLE 20A
1-Bromo-2-fluoro-5-methoxy-4-nitrobenzene
[0285] ##STR00139##
[0286] To a solution of 1-bromino-2,5-difluoro-4-nitrobenzene (5 g, 21.01 mmol) in methanol (100 mL), was added NaOMe(4.56 g, 84.43 mmol). The mixture was stirred for 12 hrs at 60 C. TLC (PE:ethyl acetate=6:1) showed that the reaction was complete. The reaction solution was cooled to it, concentrated in vacuum. The residue was diluted with H.sub.2O (30 mL), and extracted with ethyl acetate (50 mL2). The organic layer was washed with saturated brine (20 mL2), dried over anhydrous sodium sulfate, then filtered and concentrated. The residue was purified by silica gel column chromatography (PE:ethyl acetate=50:1, 30:1, 20:1) to give the title compound (3.86 g, 15.44 mmol, 73.48% yield) as yellow solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.72 (d, J=7.6 Hz, 1H), 7.31 (d, J=5.6 Hz), 3.96 (s, 3H).
EXAMPLE 20B
tert-Butyl 9-(2-fluoro-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0287] ##STR00140##
[0288] To a solution of Example 20A (120 mg, 0.48 mmol) in dioxane (10 mL), were added tert-butyl-3,9-diazaspiro[5.5]undecane-3-carboxylic acid tert-butyl ester (139.59 mg, 0.48 mmol) Pd.sub.2(dba).sub.3 (43.95 mg, 0.048 mmol), Xantphos (22.85 mg, 0.048 mmol) and CsCO.sub.3 (469.14 mg, 1.44 mmol); the reaction mixture was stirred for 12 hrs at 100 C. TLC (PE:ethyl acetate=2:1) showed that reaction was complete. The mixture was diluted with EtOAc (20 mL), washed sequentially with H.sub.2O (20 mL) and brine (20 mL), dried over anhydrous Na.sub.2SO.sub.4, filtered and concentrated to give black oil, which was separated and purified by column chromatography (PE:ethyl acetate=30:1, 20:1) to give the title compound (148 mg, yield 72.82%) as yellow solid. LCMS (ESI) (5-95AB): m/z: 424.3 [M+1].
EXAMPLE 20C
tert-Butyl 9-(2-fluoro-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester
[0289] ##STR00141##
[0290] This Example was prepared according to the process as described in Example 12C, take tert-butyl 9-(3-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid was replaced with tert-butyl 9-(2-fluoro-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid ethyl ester to give the title compound as yellow oil (crude), which was used directly in the next step.
EXAMPLE 20D
3-(2-Fluoro-5-methoxy-4-nitrophenyl)-9-methyl-3,9-diazaspiro[5.5]undecane
[0291] ##STR00142##
[0292] This Example was prepared according to the process as described in Example 12D, 3-(3-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane was replaced with tert-butyl 9-(2-fluoro-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid methyl ester to give the title compound as yellow solid (crude), which was used directly in the next step without purification. LCMS (ESI) (5-95AB): m/z: 338.2 [M+1].
EXAMPLE 20E
5-Fluoro-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline
[0293] ##STR00143##
[0294] Under N.sub.2 stomsphere, to a solution of Example 20D (90 mg, 0.267 mmol) in MeOH (10 mL), were added Zn powder (87.21 mg, 1.33 mmol) and NH.sub.4Cl (57.07 mg, 1.07 mmol). The reaction mixture was stirred for 0.5 hrs at 25 C. TLC (DCM:methanol=10:1) showed that the starting material disappeared. The reaction mixture was filtered, concentrated to give the title compound (80 mg, crude) as yellow solid.
EXAMPLE 20F
(2-((5-Chloro-2-((5-fluoro-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0295] ##STR00144##
[0296] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 5-fluoro-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline to give the title compound as white solid, yield 20%. .sup.1H NMR (400 MHz, CD.sub.3OD) , 8.34 (s, 1H), 8.10 (br. s., 1H), 7.69-7.86 (m, 3H) 7.51-7.59 (m, 1H), 7.42 (d, J=6.8 Hz, 1H), 4.02 (s, 3H), 3.58-3.69 (m, 4H) 3.47 (d, J=12.8 Hz, 2H), 3.24 (t, J=12.8 Hz, 2H), 2.93 (s, 3H), 2.14-2.23 (m, 4H) 1.96 (br. s., 2H), 1.90 (d, J=13.2 Hz, 6H), 1.76-1.86 (m, 2H). LCMS (ESI) (5-95AB): mh: 587.3 [M+1].
EXAMPLE 21
(2-((2-((5-Bromo-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-5-chloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0297] ##STR00145##
Compound 21
EXAMPLE 21A
1-Bromo-2-fluoro-4-methoxy-5-nitrobenzene
[0298] ##STR00146##
[0299] At 0 C., to a solution of 1-bromo-2-fluoro-4-methoxybenzene (5.00 g, 24.39 mmol) in sulfuric acid (20.0 mL), was added potassium nitrate (2.47 g, 24.39 mmol) in portions, the reaction mixture was stirred for 0.5 hrs at 0 C. TLC (PE:ethyl acetate=3:1) showed that the reaction was complete. The reaction mixture was added into ice water (50.0 mL) and quenched, extracted with EtOAc (100 mL2). The organic layer was dried over anhydrous sodium sulfate and filtered, concentrated to dry in vacuum to give the title compound (5.55 g, yield 90%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 8.20 (d. J=6.8 Hz, 1H), 6.91 (d, J=10 Hz, 1H), 3.99 (s, 3H).
EXAMPLE 21B
tert-Butyl 9-(2-bromo-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate
[0300] ##STR00147##
[0301] This Example was prepared according to the process as described in Example 4A, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester was replaced with tert-butyl 3,9-diazaspiro[5.5]undecane-3-carboxylic acid tert-butyl ester, and4-fluoro-2-methoxy-1-nitrobenzene with 1-bromo-2-fluoro-4-methoxy-5-nitrobenzene to give the title compound as yellow oil, yield 92%. LCMS (ESI) (0-60AB): mh: 484.2 [M+1].
EXAMPLE 21C
3-(2-Bromo-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane
[0302] ##STR00148##
[0303] This Example was prepared according to the process as described in Example 12C, 9-(3-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylic acid was replaced with tert-butyl 9-(2-bromo-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate to give the title compound as yellow solid (crude), which was used directly in the next step. LCMS (ESI) (0-60AB): mh: 384.1 [M+1].
EXAMPLE 21D
3-(2-Bromo-5-methoxy-4-nitrophenyl)-9-methyl-3,9-diazaspiro[5.5]undecane
[0304] ##STR00149##
[0305] This Example was prepared according to the process as described in Example 12D, 3-(3-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane was replaced with 3-(2-bromo-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane to give the title compound as yellow oil, yeild 48%. LCMS (ESI) (0-60AB): mh: 398.1 [M+1].
EXAMPLE 21E
5-Bromo-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline
[0306] ##STR00150##
[0307] This Example was prepared according to the process as described in Example 13D, 7-(3-methoxy-4-nitrophenyl)-N,N-dimethyl-7-diazaspiro[3.5]nona-2-amine was replaced with 3-(2-bromo-5-methoxy-4-nitrophenyl)-9-methyl-3,9-diazaspiro[5.5]undecane to give the title compound as green solid, yield 72%. LCMS (ESI) (10-80CD): m/z: 368.2 [M+1].
EXAMPLE 21F
(2-((2-((5-Bromo-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)-5-chloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0308] ##STR00151##
[0309] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 5-bromo-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline to give The title compound as colorless oil, yield 12%. LCMS (ESI) (5-95AB): m/z: 649.1 [M+1]
EXAMPLE 22
(2-((5-Chloro-2-((5-chloro-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0310] ##STR00152##
EXAMPLE 22A
1-Chloro-2-fluoro-4-methoxybenzene
[0311] ##STR00153##
[0312] To a solution of 4-chloro-3-fluorophenol (500 mg, 3.41 mmol) in acetone (10 mL) were added potassium carbonate (942.59 mg, 6.82 mmol) and Mel (1.94 g, 13.64 mmol). The reaction mixture was stirred for 16 hrs at 60 C. TLC (PE:ethyl acetate=20:1) showed that the reaction was complete. The mixture was concentrated to remove the solvent, then to the residue was added H.sub.2O (20 mL) and extracted with ethyl acetate (20 mL3). The organic layer was dried and concentrated to give the title compound (470 mg, 2.93 mmol, 85.84% yield) as colorless oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.24-7.31 (m, 2H), 6.71 (dd, J=10.8, 2.8 Hz, 1H), 6.65 (dt, J=8.8, 1.4 Hz, 1H), 3.80 (s, 3H).
EXAMPLE 22B
1-Chloro-2-fluoro-4-methoxy-5-nitrobenzene
[0313] ##STR00154##
[0314] This Example was prepared according to the process as described in Example 21A, 1-bromo-2-fluoro-4-methoxybenzene was replaced with 1-chloro-2-fluoro-4-methoxybenzene to give the title compound as yellow oil, yield 51%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 8.06 (d, J=7.6 Hz, 1H), 6.92 (d, J=10.4 Hz, 1H), 3.98 (s, 3H).
EXAMPLE 22C
tert-butyl 9-(2-chloro-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate
[0315] ##STR00155##
[0316] This Example was prepared according to the process as described in Example 4A, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester was replaced with tert-butyl 3,9-diazaspiro[5.5]undecane-3-carboxylate, and take 4-fluoro-2-methoxyl-1-nitrobenzene with 1-chloro-2-fluoro-4-methoxy-5-nitrobenzene to give the title compound as yellow oil, yield 93%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 8.04 (s, 1H), 6.57 (s, 1H), 3.96 (s, 3H), 3.38-3.47 (m, 4H), 3.16 (br. s., 4H), 1.69-1.73 (m, 4H), 1.53 (br. s., 4H), 1.47 (s, 9H).
EXAMPLE 22D
tert-Butyl 9-(4-amino-2-chloro-5-methoxyphenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate
[0317] ##STR00156##
[0318] This Example was prepared according to the process as described in Example 13D, 7-(3-methoxy-4-nitrophenyl)-N,N-dimethyl-7-azaspiro[3.5]nonane-2-amine was replaced with tert-butyl 9-(2-chloro-5-methoxy-4-nitrophenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate to give the title compound as brown oil, yield 60%. LCMS (ESI) (10-80CD):m/z: 310.2 [M100+1].
EXAMPLE 22E
(2-((5-Chloro-2-((5-chloro-2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyridin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0319] ##STR00157##
[0320] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with tert-butyl 9-(4-amino-2-chloro-5-methoxyphenyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate to give the title compound as brown oil, yield 35%. LCMS (ESI) (5-95AB): m/z: 589.2 [M+1].
EXAMPLE 22F
(2-((5-Chloro-2-((5-chloro-2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0321] ##STR00158##
[0322] This Example was prepared according to the process as described in Example 7, (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((5-chloro-2-((5-chloro-2-methoxy-4-(3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyridin-4-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as white solid, yield 40%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 8.35 (s, 1H), 8.06 (dd, J=7.6, 3.6 Hz, 1H), 7.97 (s, 1H), 7.80 (dd, J=13.2, 7.2 Hz, 1H), 7.73 (t, J=8.0 Hz, 1H), 7.58 (s, 1H), 7.50-7.57 (m, 1H), 4.06 (s, 3H), 3.74-3.78 (m, 4H), 3.44-3.47 (m, 2H), 3.24 (t, J=12.4 Hz, 2H), 2.92 (s, 3H), 2.15-2.30 (m, 4H), 2.01 (br. s., 2H), 1.90 (s, 3H), 1.87 (s, 3H), 1.77-1.86 (m, 2H). LCMS (ESI) (5-95AB):m/z: 603.2 [M+1].
EXAMPLE 23
(2-((5-Chloro-2-((2-methoxy-5-methyl-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0323] ##STR00159##
EXAMPLE 23A
tert-butyl 9-methyl-3,9-diazaspiro[5.5]undecane-3-carboxylate
[0324] ##STR00160##
[0325] At 0 C., to a solution of tert-butyl 3,9-diazaspiro[5.5]undecan-3-carboxylate (120 mg, 0.47 mmol) in methanol (5 mL) was added 37% formaldehyde solution (71 mg, 2.36 mmol), the reaction mixture was stirred for 0.5 hrs at 16 C. Then sodium triacetoxyborohydride (299 mg, 1.42 mmol) was add into the reaction mixture, and stirred for 12 hrs at 16 C. LCMS showed that the reaction was complete. The mixture was concentrated to remove the solvent, the obtained residue was diluted with H.sub.2O (10 mL) and extracted with DCM (60 mL), the organic layer was dried over anhydrous sodium sulfate, filtered and concentrated to give the title compound as colorless oil (100 mg, yield 79%).
EXAMPLE 23B
3-Methyl-3,9-diazaspiro[5.5]undecane
[0326] ##STR00161##
[0327] A solution of Example 23A (100 mg, 0.37 mmol) in trifluoroacetic acid (2 mL) and DCM (2 mL) was stirred for 1 hr at 16 C. LCMS showed that the reaction was complete. The mixture was alkalized by Na.sub.2CO.sub.3 solution (50 mL) and extracted with DCM (50 mL2). The organic layer was dried over sodium sulfate, filtered and concentrated to give the title compound as brown oil (70 mg).
EXAMPLE 23C
3-(5-Methoxy-2-methyl-4-nitrophenyl)-9-methyl-3,9-diazaspiro[5.5]undecane
[0328] ##STR00162##
[0329] This Example was prepared according to the process as described in Example 4A, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylic acid methyl ester was replaced with 3-methyl-3,9-diazaspiro[5.5]undecane, and 4-fluoro-2-methoxyl-1-nitrobenzene with 1-fluoro-5-methoxy-2-methyl-4-nitrobenzene to give the title compound as yellow oil, yield 40%. LCMS (ESI) (0-60AB):m/z: 334.2 [M+1].
EXAMPLE 23D
2-Methoxy-5-methyl-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline
[0330] ##STR00163##
[0331] This Example was prepared according to the process as described in Example 13D, 7-(3-methoxy-4-nitrophenyl)-N,N-dimethyl-7-azaspiro[3.5]nonane-2-amine was replaced with 3-(5-methoxy-2-methyl-4-nitrophenyl)-9-methyl-3,9-diazaspiro[5.5]undecane to give the title compound as brown solid, yield 9%. LCMS (ESI) (10-80CD): m/z: 304.3 [M+1].
EXAMPLE 23E
(2-((5-Chloro-2-((2-methoxy-5-methyl-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0332] ##STR00164##
[0333] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 2-methoxy-5-methyl-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline to give the title compound as white solid, yield 19%. .sup.1H NMR (400 MHz, CD3OD): , 8.28 (s, 1H), 8.17 (br.s, 1H), 7.75-7.84 (m, 1H), 7.62-7.74 (m, 2H), 7.48-7.57 (m, 1H), 7.42 (s, 1H), 4.04 (s, 3H), 3.56-3.84 (m, 4H), 3.42-3.54 (m, 2H), 3.18-3.27 (m, 2H), 2.94 (s, 3H), 2.21-2.44 (m, 6H), 2.01-2.13 (m, 2H), 1.76-1.95 (m, 9H). LCMS (ESI) (5-95AB): m/z: 583.3 [M+1].
EXAMPLE 24
(2-((5-Chloro-2-((4-methoxy-6-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)pyridin-3-yl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0334] ##STR00165##
EXAMPLE 24A
3-(4-Methoxy-5-nitropyridin-2-yl)-9-methyl-3,9-diazaspiro[5.5]undecane
[0335] ##STR00166##
[0336] Under N.sub.2 atomsphere, a solution of Example 23B (70 mg, 0.42 mmol), 2-chloro-4-methoxyl-5-nitropyridine (94 mg, 0.5 mmol), Pd.sub.2(dba).sub.3 (38 mg, 0.04 mmol), Xantphos (24 mg, 0.04 mmol) and CsCO.sub.3 (271 mg, 0.83 mmol) in dioxane (5 mL) was stirred for 12 hrs at 90 C. LCMS showed that the reaction was complete. The reaction mixture was diluted with H.sub.2O (40 mL) and extracted with DCM (50 mL2). The organic layer was washed with brine (30 mL2), dried over anhydrous sodium sulfate, filtered and concentrated. The obtained residue was purified by silica gel column chromatography (PE:ethyl acetate=4:1, 3:1) to give the title compound (50 mg, yield 37.5%) as yellow oil.
EXAMPLE 24B
4-Methoxy-6-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)pyridin-3-amine
[0337] ##STR00167##
[0338] This Example was prepared according to the process as described in Example 13D, 7-(3-methoxy-4-nitrophenyl)-N,N-dimethyl-7-azaspiro[3.5]nonane-2-amine was replaced with 3-(4-methoxy-5-nitropyridin-2-yl)-9-methyl-3,9-diazaspiro[5.5]undecane to give the title compound as brown solid, yield 88%.
EXAMPLE 24C
(2-((5-Chloro-2-((4-methoxy-6-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)pyridin-3-yl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0339] ##STR00168##
[0340] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 4-methoxy-6-(9-methyl-3,9-diaz aspiro[5.5]undecan-3-yl)pyridin-3-amine to give the title compound as white solid, yield 15%. LCMS (ESI) (10-80CD): m/z: 570.2 [M+1].
##STR00169##
EXAMPLE 25
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)-3-fluorophenyl)dimethyl phosphine oxide
[0341] ##STR00170##
EXAMPLE 25A
(2-Amino-3-fluorophenyl)dimethyl phosphine oxide
[0342] ##STR00171##
[0343] At 15 C., to a mixture of 2-bromo-6-fluoro-aniline (1.00 g, 5.26 mmol) and dimethyl phosphine oxide (451.84 mg, 5.79 mmol) in H.sub.2O (20 mL) were added potassium carbonate (2.91 g, 21.05 mmol) and Pd/C (150 mg), the reaction mixture was stirred and heated under microwave irradiation for 3 hrs at 160 C. TLC (PE:ethyl acetate=10:1) showed that the reaction was complete. The reaction mixture was extracted with DCM (20 mL4). The combined organic layer was filtered, the filtrate was concentrated and separated by silica gel column chromatography (DCM:methanol=1:0 to 10:1) to give the title compound (100 mg, yield 10.16%) as yellow solid. .sup.1H NMR (400 MHz, CDCl.sub.3): 7.12-7.03 (m, 1H), 6.86 (dd, J=8.0, 13.2 Hz, 1H), 6.68-6.58 (m, 1H), 5.52 (br. s., 2H), 1.79 (d, J=13.2 Hz, 6H).
EXAMPLE 25B
(2-((2,5-Dichloropyrimidin-4-yl)amino)-3-fluorophenyl)dimethyl phosphine oxide
[0344] ##STR00172##
[0345] At 0 C., to a mixture of Example 25A (100 mg, 0.534 mmol) in DMF (2 mL) was added NaH (53.60 mg, 1.34 mmol, 60%), the reaction mixture was stirred for 0.5 hrs at 0 C., 2,4,5-trichloropyrimidine (196 mg, 1.07 mmol) was added to the reaction mixture, the reaction mixture was stirred for 16 hrs at 15 C. LCMS showed that the reaction was complete. To the reaction mixture was added H.sub.2O (30 mL) and extracted with ethyl acetate (15 mL3). The combined organic layer was washed with brine (20 mL) and concentrated, the residue was purified by silica gel column chromatography (DCM:methanol=1:0 to 10:1) to give the title compound (150 mg, yield 84.02%) as brown oil. .sup.1H NMR (400 MHz, CDCl.sub.3): 8.28 (s, 1H), 7.53-7.33 (m, 3H), 7.28-7.17 (m, 1H), 5.54 (br. s., 2H), 1.85 (d, J=13.2 Hz, 6H). LCMS (ESI) (10-80CD): m/z: 334.0 [M+1].
EXAMPLE 25C
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)-3-fluorophenyl)dimethyl phosphine oxide
[0346] ##STR00173##
[0347] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline, and (2-((2,5-dichloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide with (2-((2,5-dichloropyrimidin-4-yl)amino)-3-fluorophenyl)dimethyl phosphine oxide to give the title compound as brown solid, yield 17%. .sup.1H NMR (400MHz, CDCl.sub.3): 8.36 (s, 1H), 7.84-7.73 (m, 2H), 7.71-7.62 (m, 2H), 7.55-7.49 (m, 2H), 7.16-7.09 (m, 1H), 4.02 (s, 3H), 3.80-3.60 (m, 4H), 3.54-3.50 (m, 2H), 3.29-3.20 (m, 2H), 2.94 (s, 3H), 2.47-1.90 (m, 6H), 1.83 (d, J=13.6 Hz, 8H). LCMS (ESI) (0-60AB): m/z: 587.3 [M+1].
EXAMPLE 26
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-fluorophenyl)dimethyl phosphine oxide
[0348] ##STR00174##
EXAMPLE 26A
(2-Amino-5-fluorophenyl)dimethyl phosphine oxide
[0349] ##STR00175##
[0350] This Example was prepared according to the process as described in Example 1K, 2-iodoaniline was replaced by 4-fluoro-2-iodoaniline to give the title compound as brown solid, yield 47%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 6.96 (m, 1H), 6.77 (m, 1H), 6.60 (m, 1H), 5.21 (br s, 2H), 1.72-1.81 (d, J=22.8 Hz, 6H). LCMS (ESI) (10-80CD): m/z: 188.1 [M+1].
EXAMPLE 26B
(2-((2,5-Dichloropyrimidin-4-yl)amino)-5-fluorophenyl)dimethyl phosphine oxide
[0351] ##STR00176##
[0352] This Example was prepared according to the process as described in Example 1L, (2-aminophenyl)dimethyl phosphine oxide was replaced with (2-amino-5-fluorophenyl)dimethyl phosphine oxide to give the title compound as white solid, yield 50%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 11.34 (br. s., 1H), 8.66 (m, 1H), 8.19-8.28 (m, 1H), 7.35-7.28 (m, 1H), 6.98 (m, 1H) 1.89-1.83 (d, J=13.2HZ, 6H). LCMS (ESI) (5-95AB): m/z: 334.1 [M+1].
EXAMPLE 26C
(2-((5-Chloro-2-((2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)phenyl)amino)pyrimidin-4-yl)amino)-5-fluorophenyl)dimethyl phosphine oxide
[0353] ##STR00177##
[0354] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 2-methoxy-4-(9-methyl-3,9-diazaspiro[5.5]undecan-3-yl)aniline, and (2-((2,5-dichloropyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide with (2-((2,5-dichloropyrimidin-4-yl)amino)-5-fluorophenyl)dimethyl phosphine oxide to give the title compound as white solid, yield 10%. LCMS (ESI) (5-95AB): mlz: 587.2 [M+1].
##STR00178##
EXAMPLE 27
(2-((5-Chloro-2-((2-methoxy-4-(3-methyl-3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0355] ##STR00179##
EXAMPLE 27A
tert-Butyl 4-formylpiperidine-1-carboxylate
[0356] ##STR00180##
[0357] At 70 C., under N.sub.2 atomsphere, to a solution of dimethyl sulfoxide (4.37 g, 46.52 mmol) in DCM (25 mL) was dropwise added oxalyl chloride (5.9 g, 46.52 mmol) in DCM (75 mL); then at 70 C., a solution of tert-butyl 4-(hydroxymethyl)piperidine-1-carboxylate (5 g, 23.26 mmol) in DCM (40 mL) was dropwise added into the above mixture. The reaction mixture was stirred for 15 minutes at 70 C., triethylamine (11.76 g, 116.3 mmol) was dropwise added into the reaction mixture. After addition, the reaction mixture was stirred for 1 hr at 70 C., and heated to 15 C. The reaction mixture was poured into H.sub.2O, and extracted with DCM (200 mL). The organic layer was washed with Na.sub.2HCO.sub.3 solution (100 mL), dried over anhydrous sodium sulfate, filtered and concentrated to dry. The obtained oil was purified by silica gel column chromatography (PE:ethyl acetate=10:1 to 3:1) to give the title compound (2.4 g, yield 48%) as yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 9.68 (s, 1H), 3.98-3.92 (m, 2H), 2.96-2.91 (m, 2H), 2.45-2.42 (m, 1H), 1.89-1.60 (m, 2H), 1.60-1.54 (m, 2H), 1.47 (s, 9H).
EXAMPLE 27B
tert-Butyl 9-oxo-3-azaspiro[5.5]undec-7-ene-3-carboxylate
[0358] ##STR00181##
[0359] Butyl-3-ene-2-ketone (0.658 g, 9.39 mmol) was add to a solution of Example 27A (2 g, 9.39 mmol) in THF (100 mL). The reaction mixture was cooled to 5 C., a solution of KOH-ethanol (3mol/L, 1.57 mL, 4.7 mmol) was dropwise added into the reaction mixture in 5 minutes. The reaction mixture was heated to 15 C. and stirred for 16 hrs. PE (10 mL) was added into the reaction mixture, and the mixture was washed with brine (100 mL). The organic layer was concentrated to give the crude. The crude was purified by silica gel column chromatography (PE:ethyl acetate=10:1 to 2:1) to give the title compound (1.12 g, yield 45%) as yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 6.82 (d, J=10 Hz, 1H), 5.97 (d, J=10 Hz, 1H), 3.57-3.56 (m, 2H), 2.50-2.47 (m, 2H), 2.01-1.97 (m, 2H), 1.67-1.65 (m, 2H), 1.61-1.59 (m, 2H), 1.49 (s, 9H).
EXAMPLE 27C
tert-Butyl 9-oxo-3-azaspiro[5.5]undecane-3-carboxylate
[0360] ##STR00182##
[0361] Pd/C (200 mg, 1.88 mmol) was added to a solution of Example 27B (5.00 g, 18.84 mmol) in methanol (100 mL). The suspension was vacuated and replaced by H.sub.2. The reaction mixture was kept at 10-25 C. under H.sub.2 (18 psi) atmosphere and stirred for 5 hrs. The reaction mixture was filtered, the filtrate was concentrated. The crude was purified by silica gel column chromatography (PE:EA=2:1) to give the title compound (4.78 g, 17.88 mmol, yield 94.9%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 3.47-3.44 (m, 4H), 2.38-2.35 (m, 4H), 1.81-1.77 (m, 4H), 1.58-1.56 (m, 4H), 1.49 (s, 9H).
EXAMPLE 27D
tert-Butyl 9-(((trifluoromethypsulfonypoxy)-3-azaspiro[5.5]undec-8-ene-3-carboxylate
[0362] ##STR00183##
[0363] At 78 C., under N.sub.2 atomsphere, lithium diisopropylamide (2.5M, 1.22 mL, 9 mmol) was dropwise added into a solution of Example 27C (2 g, 7.5 mmol) in THF (20 mL). After addition, the reaction mixture was stirred for 2 hrs. 1,1,1-Trifluoro-N-phenyl-N-((trifluoromethyl) sulfonyl)methanesulfonamide (2.67 g, 7.48 mmol, dissolved in 5 mL THF) was dropwise added into the reaction mixture, and the mixture was stirred for 1.5 hrs at 78 C. The mixture was heated to 10 C. and stirred for 2.5 hrs, quenched by NH.sub.4Cl solution (30 mL), and extracted with ethyl acetate (50 mL2). The organic layer was washed by brine (50 mL), dried over Na.sub.2SO.sub.4 and concentrated to give the residue. The residue was purified by silica gel column chromatography (PE:ethyl acetate=5:1) to give the title compound (2.3 g, yield as 77%) as brown oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 5.71 (t, J=4.0 Hz, 1H), 3.51-3.45 (m, 2H), 3.39-3.32 (m, 2H), 2.37-2.35 (m, 2H), 2.16-2.09 (m, 2H), 1.71-1.67 (m, 2H), 1.48 (s, 9H).1.45-1.42 (m, 4H).
EXAMPLE 27E
4-Bromo-2-methoxy aniline
[0364] ##STR00184##
[0365] NBS (4.34 g, 24.36 mmol) was added into a solution of 2-methoxyl aniline (3 g, 24.36 mmol) in MeCN (30 mL), the reaction mixture was stirred for 15 minutes at 15 C. To the reaction mixture was added aq. sodium sulfite (40 mL) to quench the reaction, the mixture was extracted with ethyl acetate (50 ml); the organic layer was dried over Na.sub.2SO.sub.4, concentrated to give the crude, the crude was purified by silica gel column chromatography (PE:ethyl acetate=10:1) to give the title compound (2.7 g, yield 55%) as yellow solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 6.93-6.91 (m, 2H), 6.62-6.59 (dd, J=1.6, 2.0 Hz, 1H), 3.86 (s, 1H).
EXAMPLE 27F
2-Methoxy-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline
[0366] ##STR00185##
[0367] Bis(pinacolato)diboron (628.4 mg, 2.47 mmol), Pd[P(C.sub.6H.sub.5).sub.3].sub.4 (150 mg, 0.13 mmol) and potassium acetate (485 mg, 4.95 mmol) were added into a solution of Example 27E (500 mg, 2.47 mmol) in DMSO (5 mL), the reaction mixture was stirred for 40 minutes at 150 C. under microwave irradiation. The reaction mixture was dilute by ethyl acetate (50 mL) and H.sub.2O (30 mL). The organic layer was separated, dried and concentrated to give the crude, the crude was purified by silica gel column chromatography (PE:ethyl acetate=10:1) to give the title compound (160 mg, yield 26%) as yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.31 (d, J=8.0 Hz, 1H), 7.22 (s, 1H), 6.71 (d, J=8.0 Hz, 1H), 3.91 (s, 3H), 1.35 (s, 12H).
EXAMPLE 27G
tert-Butyl 9-(4-amino-3-methoxyphenyl)-3-azaspiro[5.5]undec-8-ene-3-carboxylate
[0368] ##STR00186##
[0369] Pd(dppf)Cl.sub.2 (15 mg, 0.075 mmol) and potassium carbonate (204 mg, 1.5 mmol) were added into a solution of Example 27D (300 mg, 0.75 mmol), Example 27F (187 mg, 0.75 mmol) in dioxane (10 mL), the reaction mixture was stirred for 16 hrs at 110 C. The reaction mixture was filtered through diatomite, the filtrate was concentrated to give the crude, the crude was purified by silica gel column chromatography (PE:ethyl acetate=3:1) to give the title compound (110 mg, yield 39%) as yellow oil. LCMS (ESI)(5-95AB): m/z: 373.2 [M+1].
EXAMPLE 27H
tert-Butyl 9-(4-amino-3-methoxyphenyl)-3-azaspiro[5.5]undecane-3-carboxylate
[0370] ##STR00187##
[0371] Under N.sub.2 atomsphere, Pd/C (10 mg, 10%) was added into a solution of Example 27G (110 mg, 0.3 mmol) in methanol (5 mL), the reaction mixture was stirred for 5 hrs under H.sub.2 (15 PSI) at 16 C. The reaction mixture was filtered through diatomite and concentrated in vacuum to give the crude title compound (130 mg) as yellow oil. LCMS (ESI) (5-95AB): m/z: 319.1 [M56+1].
EXAMPLE 27I
(2-((5-Chloro-2-((2-methoxy-4-(3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0372] ##STR00188##
[0373] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with tert-butyl 9-(4-amino-3-methoxyphenyl)-3-azaspiro[5.5]undecane-3-carboxylate to give the title compound as colorless oil, yield 15%. LCMS (ESI) (5-95AB): m/z: 554.2[M+1].
EXAMPLE 27J
(2-((5-Chloro-2-((2-methoxy-4-(3-methyl-3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0374] ##STR00189##
[0375] This Example was prepared according to the process as described in Example 7, (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((5-chloro-2-((2-methoxy-4-(3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyridin-4-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as yellow solid, yield 27%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 8.31-8.15 (m, 2H), 7.75-7.70 (m, 1H), 7.57 (s, 1H), 7.45-7.37 (m, 2H), 7.06 (s, 1H), 6.91 (s, 1H), 3.90 (s, 3H), 3.44-3.36 (m, 2H), 3.22-3.19 (m, 2H), 2.91 (s, 3H), 2.66-2.64 (m 1H), 2.44-1.93 (m, 2H), 1.93 (s, 3H), 1.92-1.88 (d, J=13.6 Hz, 6H),1.89 (s, 3H), 1.84-1.60 (m 9H), 1.33-1.31 (m, 1H). LCMS (ESI) (5-95AB): m/z: 568.2 [M+1].
EXAMPLE 28
(2-((5-Chloro-2-((2-(difluoromethoxy)-4-(3-methyl-3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0376] ##STR00190##
EXAMPLE 28A
5-Chloro-2-nitrophenol
[0377] ##STR00191##
[0378] This Example was prepared according to the process as described in Example 11A, 4-fluoro-2-methoxy-1-nitrobenzene was replaced with 4-chloro-2-methoxy-1-nitrobenzene to give the title compound as yellow oil, yield 93%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 10.67 (s, 2H), 8.07 (d, J=8.8 Hz, 1H), 7.20 (d, J=2.4 Hz, 1H), 6.99 (dd, J=9.2,2.0 Hz, 1H).
EXAMPLE 28B
4-Chloro-2-(difluoromethoxy)-1-nitrobenzene
[0379] ##STR00192##
[0380] This Example was prepared according to the process as described in Example 11B, 5-fluoro-2-nitrophenol was replaced with 5-chloro-2-nitrophenol to give the title compound as yellow oil, yield 78%. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.92 (d, J=8.8 Hz, 1H), 7.42-7.37 (m, 2H), 6.64 (t, J=72.4 Hz, 1H).
EXAMPLE 28C
2-(3-(Difluoromethoxy)-4-nitrophenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane
[0381] ##STR00193##
[0382] Bis(pinacolato)diboron (262 mg, 1.00 mmol), Pd(dppf)Cl.sub.2 (73.17 mg, 0.1 mmol), Ph.sub.3P (262.29 mg, 1.00 mmol) and potassium acetate (196.28 mg, 2.00 mmol) were added into a solution of Example 28C (223.56 mg, 1.00 mmol) in dioxane (10 mL), the reaction mixture ventilated and stirred for 16 hrs at 100 C. TLC (PE:ethyl acetate=10:1) showed that the starting material disappeared completely. The reaction mixture was poured into H.sub.2O (30 mL) and extracted with ethyl acetate (25 mL2). The organic layer was washed with brine (30 mL), dried over anhydrous MgSO.sub.4, concentrated in vacuum to give the residue which was purified by silica gel column chromatography (PE:ethyl acetate=30:1, 20:1) to give the title compound (300 mg, yield 95.21%) as yellow solid.
EXAMPLE 28D
tert-Butyl 9-(3-(difluoromethoxy)-4-nitrophenyl)-3-azaspiro[5.5]undec-8-ene-3-carboxylate
[0383] ##STR00194##
[0384] Under N.sub.2 atomsphere, Pd(dppf)Cl.sub.2 (116 mg, 0.16 mmol) and Na.sub.2CO.sub.3 (503 mg, 4.75 mmol) were added into a mixture of Example 27D (948 mg, 2.37 mmol), Example 28C (500 mg, 1.58 mmol) in dioxane (4 mL) and H.sub.2O (2 mL), the reaction mixture was stirred for 16 hrs at 110 C. The reaction mixture was filtered through diatomite, concentrated to give the crude, the crude was purified by silica gel column chromatography (PE:ethyl acetate=5:1) to give the title compound (602 mg, yield 58%) as yellow solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.92 (d, J=8.8 Hz, 1H), 7.38-7.34 (m, 2H), 6.82-6.28 (m, 2H), 3.54-3.47 (m, 2H), 3.38-3.33 (m, 2H),2.41 (s,2H), 2.18 (d, J=9.6 Hz, 1H), 1.70-1.65 (m,2H), 1.46-1.42 (m,5H).
EXAMPLE 28E
tert-Butyl 9-(4-amino-3-(difluoromethoxy)phenyl)-3-azaspiro[5.5]undecane-3-carboxylate
[0385] ##STR00195##
[0386] Pd/C(20 mg) was added into a solution of Example 28D (200 mg, 0.46 mmol) in THF (10 mL); the reaction mixture was stirred for 5 hrs under H.sub.2 atomsphere (50 psi) at 16 C. The reaction mixture was filtered through diatomite and concentrated to give the title compound (150 mg, yield 80%) as brown solid. LCMS (ESI) (10-80CD): m/z: 355.2 [M+156].
EXAMPLE 28F
(2-((5-Chloro-2-((2-(difluoromethoxy)-4-(3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0387] ##STR00196##
[0388] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with tert-butyl 9-(4-amino-3-(difluoromethoxy)phenyl)-3-azaspiro[5.5]undecane-3-carboxylate to give the title compound as brown oil, yield 37.2%.
EXAMPLE 28G
(2-((5-Chloro-2-((2-(difluoromethoxy)-4-(3-methyl-3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0389] ##STR00197##
[0390] This Example was prepared according to the process as described in Example 7, (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((5-chloro-2-((2-(difluoromethoxy)-4-(3-azaspiro[5.5]undecan-9-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as white solid, yield 24%. LCMS (ESI) (5-95AB): m/z: 604.2 [M+1].
EXAMPLE 29
(2-(5-Chloro-2-((5-fluoro-2-methoxy-4-(7-methyl-7-azaspiro[3.5]nonan-2-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0391] ##STR00198##
EXAMPLE 29A
Benzyl 4-methylenepiperidine-1-carboxylate
[0392] ##STR00199##
[0393] Under N.sub.2 atomsphere, at rt., tert-butyl 4-oxo-piperidine-1-carboxylate (9.00 g, 45.17 mmol) was added into a mixture of methyl triphenyl phosphonium bromide (16.14 g, 45.17 mmol) and potassium tert-butoxide (5.58 g, 49.69 mmol) in THF (350 mL) once at a time. The reaction mixture was stirred for 3 hrs at 16 C. TLC showed that the reaction was complete. The residue was poured into H.sub.2O. The aqueous phase was extracted with ethyl acetate (200 mL3). The combined organic layer was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by silica gel column chromatography (PE:ethyl acetate=10:1, 5:1) to give the title compound (8.40 g, yield 80.4%) as colorless solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.41-7.34 (m, 5H), 5.17 (s 2H), 4.78 (s, 2H), 3.55-3.52 (m, 4H), 2.24-2.21(m, 4H).
EXAMPLE 29B
[0394] Benzyl 1,1-dichloro-2-oxo-7-azaspiro[3.5]nonane-7-carboxylate
##STR00200##
[0395] At 15-20 C., 2,2,2-trichloroacetic chloride (23.58 g, 129.70 mmol) was dropwise added into a mixture of Example 29A (6.00 g, 25.94 mmol) and ZnCu (30.10 g, 233.46 mmol) in THF (300 mL), after 0.5 hrs, the reaction mixture was stirred for 16 hrs at 20-30 C. The reaction solution was quenched by ice-water slowly, then extracted with ethyl acetate (250 mL3). The combined organic layer was washed with brine (100 mL2), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by silica gel column chromatography (PE:ethyl acetate=10:1, 5:1) to give the title compound (3.81 g, yield 42.81%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.42-7.36 (m, 5H), 5.17 (s 2H), 4.21-4.17 (m, 2H), 3.13 (s 2H), 3.05-2.98 (m, 2H), 2.02-1.96 (m, 2H), 1.87-1.82 (m, 2H).
EXAMPLE 29C
Benzyl 2-oxo-7-azaspiro[3.5]nonane-7-carboxylate
[0396] ##STR00201##
[0397] Zn powder (1.72 g, 26.3 lmmol) and NH.sub.4Cl (4.69 g, 87.70 mmol) were added into a mixture of Example 29B (3.00 g, 8.77 mmol) in methanol (50 mL). The reaction mixture was stirred for 0.5 hrs at 10-20 C. The reaction mixture was poured into H.sub.2O (50 mL), and stirred for 20minutes. The aqueous phase was extracted with ethyl acetate (50 mL3). The combined organic layer was washed with sat. brine (30 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by silica gel column chromatography (PE:ethyl acetate=30:1, 20:1) to give the title compound (2.12 g, yield 8.44%) as yellow solid. .sup.1H NMR (400 MHz, CDCl.sub.3): , 7.40-7.35 (m, 5H), 5.16 (s 2H), 3.54-3.51 (m, 4H), 2.85 (s, 4H), 1.76-1.74 (m, 4H).
EXAMPLE 29D
Benzyl 2-(2-fluoro-5-methoxyphenyl)-2-hydroxy-7-azaspiro[3.5]nonane-7-carboxylate
[0398] ##STR00202##
[0399] Under N.sub.2 atomsphere at 60 C., n-butyllithium (249.83 mg, 3.90 mmol) was added into a mixture of 2-bromo-1-fluoro-4-methoxyl-benzene (400 mg,1.95 mmol) in THF (15 mL). The mixture was stirred for 1 hr at 60 C. Example 29C (399.75 mg, 1.46 mmol) was added into the reaction mixture over 20 minutes at 60 C. The reaction mixture was stirred for 2 hrs at 60 C., the reaction mixture was poured into H.sub.2O (50 mL), and stirred for 20minutes. The aqueous phase was extracted with ethyl acetate (50 mL3), the combined organic layer was washed with brine (50 mL2), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by silica gel column chromatography (PE:ethyl acetate=10:1, 1:1) to give the title compound (340 mg, yield 43.65%) as colorless oil.
EXAMPLE 29E
Benzyl 2-(2-fluoro-5-methoxyphenyl)-7-azaspiro[3.5]nonane-7-carboxylate
[0400] ##STR00203##
[0401] Under N.sub.2 atomsphere, at rt., triethylsilane (221.24 mg, 1.90 mmol) was added into a mixture of Example 29D (380 mg, 0.95 lmmol) in DCM (10 mL). The reaction mixture was stirred for 1 hr at 18 C. The reaction mixture was poured into H.sub.2O (20 mL), and stirred for 20 minutes. The aqueous phase was extracted with ethyl acetate (30 mL3). The combined organic layer was washed with sat. brine (20 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by silica gel column chromatography (PE:ethyl acetate=10:1, 3:1) to give the title compound (160 mg, yield 43.86%) as yellow oil. LCMS(ESI)(5-95AB): m/z: 384.2[M+1].
EXAMPLE 29F
Benzyl 2-(2-fluoro-5-methoxy-4-nitrophenyl)-7-azaspiro[3.5]nonane-7-carboxylate
[0402] ##STR00204##
[0403] At 0-10 C., nitric acid (268.28 mg, 4.17 mmol) was added into a mixture of Example 29E (160 mg, 0.417 mmol) in AcOH(4 mL) once at a time. The reaction mixture was stirred for 2 hrs at 18 C. The mixture was poured into ice H.sub.2O (20 mL), and stirred for 20 minutes. The aqueous phase was extracted with ethyl acetate (40 mL3). The combined organic layer was washed with sat. brine (20 mL), dried over anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by silica gel column chromatography (PE:ethyl acetate=2:1) to give the title compound (145 mg, yield 81.10%) as yellow oil. LCMS(ESI)(5-95AB): m/z: 451.1 [M+Na].
EXAMPLE 29G
5-Fluoro-2-methoxy-4-(7-azaspiro[3.5]nonan-2-yl)aniline
[0404] ##STR00205##
[0405] Under N.sub.2 atomsphere, Pd/C (40 mg, 10%) was added into a mixture of Example 29F (148.00 mg, 345.43 umol) in THF (10 mL). The reaction solution was vacuumized and exchanged with H.sub.2, the reaction mixture was stirred for 3 hrs under H.sub.2 (16 psi) at 10-25 C. The reaction mixture was filtered, the filtrate was concentrated. The crude was purified by silica gel column chromatography (DCM:methanol=20:1) to give the title compound (50 mg, yield 54.76%) as yellow oil. LCMS(ESI)(5-95AB): m/z: 265.2 [M+1].
EXAMPLE 29H
(2-((5-Chloro-2-((5-fluoro-2-methoxy-4-(7-azaspiro[3.5]nonan-2-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0406] ##STR00206##
[0407] This Example was prepared according to the process as described in Example 1M, 2-methoxy-5-(6-methyl-6-azaspiro[3.4]octan-2-yl)aniline was replaced with 5-fluoro-2-methoxy-4-(7-azaspiro[3.5]nonan-2-yl)aniline to give the title compound as yellow oil, yield 27.21%. LCMS(ESI)(5-95AB): m/z: 544.2 [M+1].
EXAMPLE 29
(2-((5-Chloro-2-((5-fluoro-2-methoxy-4-(7-methyl-7-azaspiro[3.5]nonan-2-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide
[0408] ##STR00207##
[0409] This Example was prepared according to the process as described in Example 7, (2-((5-chloro-2-((2-methoxy-4-(2,6-diazaspiro[3.4]octan-6-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide was replaced with (2-((5-chloro-2-((5-fluoro-2-methoxy-4-(7-azaspiro[3.5]nonan-2-yl)phenyl)amino)pyrimidin-4-yl)amino)phenyl)dimethyl phosphine oxide to give the title compound as yellow solid, yield 42%. .sup.1H NMR (400 MHz, CD.sub.3OD): , 8.28 (s, 1H), 8.25-8.19 (m, 1H), 7.79-7.73 (m, 1H), 7.63-7.61(m, 1H), 7.50-7.41(m, 2H), 6.98 (d, J=6.8, 1H), 3.93(s, 3H), 3.81-3.72(m, 1H), 3.52-3.39 (m, 2H), 3.17-3.11 (m, 1H), 3.01 (m,1H), 2.89(s, 3H), 2.56-2.51(m 1H), 2.33-2.30(m, 2H), 2.19-2.06(m, 2H), 1.91-1.88(m, 8H). LCMS(ESI)(5-95AB): m/z: 558.2 [M+1].
[0410] The Biochemical Assay
[0411] The Experimental Materials
[0412] Enzyme: ALK wild-type, ALK C1156Y and ALK L1196M supplied by Carna Biosciences (Japan), EGFR T790M/L858R supplied by Life technology (Madison, Wis.).
[0413] HTRF kit: supplied by Cis-Bio International, with Eu-marked TK1 antibody, XL665- and biotin-labelled polypeptide substrates.
[0414] Testing instrument: Envision (PerkinElmer).
[0415] The Experimental Method
[0416] The compound to be tested is diluted at a gradient by 3 times thereby giving 11 gradients starting from 1 m to 0.017 nM for the final concentration.
[0417] 10 l wild-type ALK enzyme reaction mixture system: 0.5 nM wild-type ALK, 1 M biotin-TK1 peptide, 30 M ATP. Reaction buffer: 50 mM Hepes (pH7.5), 10 mM MgCl2, 0.01 mM NaV3VO4. The reaction plate was white Proxiplate 384-Plus plate (PerkinElmer), reaction was reacted at room temperature for 90 minutes.
[0418] 10 l ALK C1156Y enzyme reaction mixture system: 0.15 nM ALK C1156Y, 1 M biotin-TK1 peptide, 30 M ATP. Reaction buffer: 50 mM Hepes (pH7.5), 10 mM MgCl.sub.2, 0.01 mM NaV3VO4. The reaction plate was white Proxiplate 384-Plus plate (PerkinElmer). Reaction was reacted at room temperature for 60 minutes.
[0419] 10 l ALK L1196M enzyme reaction mixture system: 0.15 nM ALK L1196M, 1 M biotin-TK1 peptide, 30 M ATP. Reaction buffer: 50 mM Hepes (pH7.5), 10 mM MgCl2, 0.01 mM NaV3VO4. The reaction plate was white Proxip at 384:-Plus plate (PerkinElmer), reaction was reacted at room temperature for 60 minutes.
[0420] 10 l EGER T790M/L858R enzyme reaction mixture system: 0.08 nM EGER T790M/L858R, 1 M biotin-TK1 peptide, 20 M ATP. Reaction buffer: 50 mM Hepes (pH7.5), 10 mM MgCl2, 0.01 mM NaV3VO4. The reaction plate was white Proxiplate 384-Plus plate (PerkinElmer). Reaction was reacted at room temperature for 60 minutes.
[0421] Detecting reaction: 10 l testing reagent was added to the reaction plate, the final concentration of Antibody was 2 nM, XL665 was 62.5 nM. Incubation was carried out for 60 minutes at room temperature. Detection was by Envision.
[0422] Data Analysis
[0423] Data were transformed into inhibition rate (%) according to the following formula: (MinRatio)/(MaxMin)*100%. IC.sub.50 was obtained according to 4 parameters curve fitting (Model 205 in XLFIT5, iDBS).
[0424] Cell Assay
[0425] The Experimental Materials
[0426] RPMI1640, fetal bovine serum, penicillin/streptomycin solution are purchased from Life Technology (Madison, Wis.). Cell Titer-Glo luminescent Cell viability reagents was supplied by Promega (Madison, Wis.). Karpas299 cell line was supplied by European Collection of cell Cultures (ECACC). The instrument: Envision (PerkinElmer).
[0427] The Experimental Method
[0428] A 384-well plate, 2500 Karpas-299 cells per well were seeded, 45 l volume, which was incubated overnight in the CO, incubator at 37 C. The compounds to be tested are (leaned by 3 times at a gredient, thereby obtaining 10 gredients starting from 2.5 mM to 2.5 M of the concentration, reduplicating double holes. 49 l Medium per hole was added into the middle plate. 1 l Compound was transferred to the middle plate from the plate where compound was diluted at a gradient and fully mixed. 5 l Liquid was further transferred to the cell plate from the the middle plate. Cells was further incubated for 72 hours in the CO, incubator. After 72 hours, 25 l testing reagent was added. Incubation was carried out for 10 minutes at room temperature, then detected by Envision.
[0429] Data Analysis
[0430] Data were transformed into inhibition rate (%) according to the following formula: (Max-Sample)/(Max-Min)*100%. IC.sub.50 was obtained according to 4 parameters curve fitting (Model 205 in XLFIT5, iDBS).
[0431] ALK enzymatic inhibition IC.sub.50, ALK L1196M enzymatic inhibition IC.sub.50, ALK C1156Y enzymatic inhibition IC.sub.50, EGFR T790M/L858R enzymatic inhibition IC.sub.50, and ALK IC.sub.50 of Karpas-299 cells of the compounds in the invention were listed in the Table below. Compounds with IC.sub.50 between 1 nM to 100 nM were designated as +++; compounds with IC.sub.50 between 101 nM to 1000 nM were designated as ++, and compounds with IC.sub.50 above 1000 nM were designated as +.
TABLE-US-00001 ALK ALK ALK EGFR IC.sub.50 L1196M IC.sub.50 C1156Y IC.sub.50 T790M/L858R IC.sub.50 Karpas-299 Cell Compound (nM) (nM) (nM) (nM) ALK IC.sub.50 (nM) 1 +++ ++ ++ ++ 2 ++ ++ ++ ++ 3 +++ +++ +++ +++ +++ 4 +++ +++ +++ +++ +++ 5 +++ +++ +++ +++ +++ 6 +++ +++ +++ +++ ++ 7 +++ ++ ++ ++ + 8 +++ +++ +++ +++ +++ 9 +++ +++ +++ +++ +++ 10 +++ +++ +++ +++ +++ 11 +++ +++ +++ +++ +++ 12 +++ +++ +++ +++ +++ 13 +++ +++ +++ +++ +++ 14 +++ +++ +++ +++ +++ 15 + ++ + + 16 ++ ++ ++ +++ 17 +++ +++ +++ +++ +++ 18 + + + + + 19 + + + + + 20 +++ +++ +++ +++ +++ 21 +++ +++ +++ +++ +++ 22 +++ +++ +++ +++ +++ 23 +++ +++ +++ +++ +++ 24 +++ +++ +++ ++ 25 +++ +++ +++ ++ 26 +++ +++ +++ +++ +++ 27 +++ +++ +++ +++ +++ 28 +++ +++ +++ +++ +++ 29 +++ +++ +++ +++ +++
[0432] In Vivo Efficacy Study
[0433] The in vivo efficacy data below show that the compounds of the present invention have unexpected antitumor activity and reduced tumor size against both wild type LU-01-0015 xenotransplantation (PDX) model (BALB/c nude mice) source of lung cancer patients and LU-01-0319 Crizotinib resistance model (BALB/c nude mice) over the reference compound AP26113. For example, in the model of LU-01-0015, after 16 days dating from compound 9, 12, 22, 27 etc being administered (25 mg/kg), the tumor volume was reduced to 34-50 nim from 277 mm.sup.3, but AP26133 only to 119 mm.sup.3.
[0434] 1. In vivo efficacy experiment on BALB/c nude mice by subcutaneously implanting LU-01-0015 xenotransplantation (PDX) from lung cancer patients
[0435] BALB/c nude mice, female. 6 to 8 weeks, about 18 to 22 g bodyweight, was kept in a special environment without pathogens, and a single ventilated cage (5 mice per cage). Beddings and water in all the cages were disinfected before use. All the animal had free access to standard authorized commercially available lab diet. 80 mice were supplied by Shanghai BK Laboratory Animal Co., LTD for study. Each mouse was subcutaneously implanted tumor tissue (20-30 mm.sup.3) at right flank for growth of tumor. When the average tumor volume reached about 250-300 mm.sup.3, the assay started. Test compounds were delivered orally everyday, 25 mg/kg, Tumor size was measured with two-dimensional caliper every 3 days, size was recorded by mm.sup.3, and calculated by the following formula: V=V=0.5ab2, where a and b are respectively the long diameter and short diameter length of the tumor. The antitumor efficacy was determined by dividing the average increased tumor volume in compound-treated animal by the average increased tumor volume in untreated animal.
TABLE-US-00002 Tumor volume (mm.sup.3) Example Compound 0 day 3 days 6 days 9 days 12 days 16 days 19 days 23 days Vehicle 277 432 506 581 637 815 942 968 Crizotinib 275 297 288 311 340 413 419 439 AP26113 278 207 174 162 142 119 122 119 27 277 158 82 68 49 34 41 35 12 276 199 97 90 58 49 42 32 22 277 234 137 90 68 46 54 41 26 278 235 127 84 55 46 59 53 5 277 259 214 172 117 104 123 124 9 275 158 108 82 58 48 46 43
[0436] 2. In vivo anti-tumor efficacy experiment on Crizotinib resistant BALB/c nude mice by subcutaneously implanting LU-01-0319 xenotransplantation (PDX) tumor
[0437] LU-01-0319 xenotransplantation tumor models initially obtained from clinical sample through surgical resection were implanted into nude mice, which was defined as a batch of P0 (LU-01-0319-P0). The next batch was defined as P1 (LU-01-0319-P1) which was implanted tumors from P0. FP3 was regained from P2, the next batch was defined as FP4 which was implanted tumors from FP3. After about 2 to 3 weeks, tumor size reached about 300 mm.sup.3, and the tumor-bearing mice were treated with Crizotinib. The continuous increasing tumors were defined as LU-01-0319 antitumor models. BALB/c nude mice, female, 6 to 8 weeks, about 18 to 22 g bodyweight, 75 mice were used for study, supplied by Shanghai BK Laboratory Animal Co., LTD. Tumour biopsies (about 30 mm.sup.3) of LU-01-0319R FP6 were implanted subcutaneously on right side of each mouse for growth of tumor. About 2 to 3 weeks dating from implanting, tumor size reached about 300 mm.sup.3, the turnor-bearing mice were treated with Crizotinib (10/25/50/75 mg/kg). Dose of Crizotinib was appropriately varied depending on the tumor size. When average tumor volume reached about 500 mm.sup.3, the assay began. Once a day the compounds to be tested were orally administered. Tumor size were measured twice a week with a caliper at two dimensions, and calculated with the following formula: V=0.5ab2, where a and b are respectively long diameter and short diameter of the tumor. The antitumor efficacy was determined by dividing the average increased tumor volume in compound-treated animal by the average increased tumor volume in untreated animal.
TABLE-US-00003 Tumor volume (mm.sup.3) Example Compound 0 day 3 days 7 days 10 days 13 days 17 days 19 days Vehicle 502 813 948 1236 1382 1807 1934 Crizotinib*** 502 708 828 1002 1116 1415 1488 AP26113* 501 656 690 898 1096 1415 1499 AP26113** 502 646 604 593 504 568 508 9* 502 722 859 1101 1218 1441 1521 9** 502 557 416 261 196 133 102 27* 503 689 777 1003 1177 1490 1602 27** 502 650 752 743 705 756 692 *dosage 3 mg/Kg; **dosage 10 mg/Kg; ***25 mg/Kg
[0438] 3. In vivo efficacy experiment on BALB/c nude mice by subcutaneously implanting with LU-01-0319 xenotransplantation (PDX) from lung cancer patients
[0439] BALB/c nude mice, female, 6 to 8 weeks, about 18 to 22 g bodyweight, wss kept in a special environment without pathogens, and a single ventilated cage (5 mice per cage). Beddings and water in all the cages were disinfected before use. All the animal had free access to standard authorized commercially available lab diet. 80 mice were supplied by Shanghai BK Laboratory Animal Co., LTD for study. Each mouse was subcutaneously implanted tumor tissue (20-30 mm.sup.3) at right flank for growth of tumor. When the average tumor volume reached about 250-300 mm.sup.3, the assay started. Test compounds were delivered orally everyday, 10 mg/kg. Tumor size was measured with two-dimensional caliper every 3 days, size was recorded by mm.sup.3, and calculated by the following formula: V=V=0.5ab2, where a and b are respectively the long diameter and short diameter length of the tumor. The antitumor efficacy was determined by dividing the average increased tumor volume in compound-treated animal by the average increased tumor volume in untreated animal.
TABLE-US-00004 Tumor volume (mm.sup.3) Compound 0 days 2 days 6 days 9 days 13 days 16 days 20 days 23 days Vehicle 372 480 559 715 939 1121 1357 1732 LDK378 373 397 366 350 425 369 329 268 AP26113 371 437 448 485 512 492 457 391 9 371 359 369 301 305 291 203 172
[0440] ALK inhibitors of the present invention can be used in the treatment of various cancers including anaplastic large cell lymphoma, non-small cell lung cancer, diffuse large B cell lymphoma, inflammatory myofibroblastic tumor, neuroblastoma, thyroid anaplastic carcinoma and rhabdomyosarcoma. ALK inhibitors can be used as a therapy alone or in combination with other chemotherapy agents.