CN106543185B - A kind of compound for targetting ubiquitination degraded PLK1 and BRD4 albumen and its application - Google Patents
A kind of compound for targetting ubiquitination degraded PLK1 and BRD4 albumen and its application Download PDFInfo
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- CN106543185B CN106543185B CN201610991723.1A CN201610991723A CN106543185B CN 106543185 B CN106543185 B CN 106543185B CN 201610991723 A CN201610991723 A CN 201610991723A CN 106543185 B CN106543185 B CN 106543185B
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Abstract
The present invention relates to a kind of compound for targetting ubiquitination degraded PLK1 and BRD4 albumen and its application, belong to medicinal chemistry art, more particularly to a kind of compound for targetting ubiquitination degraded Polo samples kinases 1 and bromine domain protein 4, and its pharmaceutically acceptable salt, hydrate, with the drug regimen using the compound as active component, and prepare PLK1 and BRD4 albumen suppress and degradation agent and its for treat and or pre- anti-cancer in purposes.Preparation method of the present invention is simple to operate, mild condition, and gained compound is respectively provided with PLK1 and BRD4 albumen enzyme level and degrading activity, and antitumor action is notable.
Description
Technical field
The present invention relates to pharmaceutical technology field, more particularly to a kind of change for targetting ubiquitination degraded PLK1 and BRD4 albumen
Compound, and its pharmaceutically acceptable salt, hydrate and the drug regimen using the compound as active component, and preparing
PLK1 albumen and BRD4 protein inhibitors and its for treat and/or pre- preventing tumor in application.
Background technology
Ubiquitin-Proteasome Pathway (ubiquitin proteasome pathway, UPP) is Intracellular proteolysis
Main path, participate in the degraded of intracellular more than 80% protein.UPP is by ubiquitin, ubiquitin activating enzyme E1, ubiquitin binding enzyme
E2, ubiquitin ligase E3, proteasome and its substrate (protein) are formed.The process of UPP selective degradation protein is divided to two
Stage：(1) protein substrate ubiquitination：Ubiquitin molecule provides energy by APP, E2 is transferred to by E1 activation, then through E3 and specifically
Property protein substrate combine；(2) protein substrate is degraded：It can be identified by the protein molecular of ubiquitination by proteasome, and enter egg
White enzyme body is degraded into the peptide molecule of short chain.
PROTACs technologies are that target protein and intracellular E3 are furthered using a kind of difunctional small molecule, so as to cause
The degraded of target protein.PROTACs includes three parts functional structure：(1) can be with part that protein substrate is combined；(2)
Can be with part that E3 is combined；(3) two-part connects chain before.In the cell PROTACs can simultaneously with target protein and E3
With reference to making the target protein ubiquitination that can not be combined originally with E3, and then identified and degraded by proteasome.
(Angew.Chem.Int.Ed.Engl.2016,55(6),1966-1973.)
Research has shown that Thalidomide class medicine (Thalidomide, lenalidomide and pomalidomide) can be incorporated into CRBN albumen
On, CRBN can be with damage dna Binding Protein 1 (damaged DNAbinding protein 1, DDB1), Cullin-4A
(CUL4A), the regulators of cullins 1 (Roc1) combine, and form E3 ubiquitin ligase complex CRL4.The compound utilizes
Ubiquitin marks specific albumen, then hydrolyzes these albumen.(Science.2010,327(5971),1345-1350；
Nature.2014,512(7512),49-53.)
The content of the invention
Present invention aims at provide a kind of compound for targetting ubiquitination degraded PLK1 and BRD4 albumen and its application, tool
Body is compound with phthalimide fragment and preparation method thereof, and such compound is as PLK1 and BRD4 eggs
The white application suppressed with degradation agent in preventing and/or treating tumour.
The present invention relates to the compound shown in formula I and its pharmaceutically acceptable salt or hydrate：
Wherein R1, R2, R3, R4 are independently selected from H, halogen, C1-C6 alkoxies, C1-C6 alkyl；N be 1,2,3,4,
5 or 6.
Wherein preferred R1, R2, R3, it is independently selected from H, halogen, C1-C4 alkoxies；R4 is selected from C1-C4 alkyl, more
It is preferred that R4 is ethyl；
Further preferred below formula compound：
Wherein R1, R2, R3It is independently selected from H, halogen or methoxyl group；N is 2 or 3.
Unless otherwise noted, term used herein " halogen " refers to fluorine, chlorine, bromine or iodine；C1-C6 alkoxies are nails
The C5 alcoxyls of epoxide, ethyoxyl, positive propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, straight chain or straight chain
The C6 alkoxies of base, straight chain or straight chain；C1-C6 alkyl refers to methyl, ethyl, propyl group, isopropyl, butyl, isobutyl group, tertiary fourth
The C6 alkyl of the C5 alkyl of base, straight chain or straight chain, straight chain or straight chain.The present invention is further selected from：
4- (((S) -8- benzyl -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2-
(2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethoxy
Base) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- luorobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- chlorobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (3- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (2- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (the chloro- 4- luorobenzyls of 3-) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) ammonia
Base)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) second
Epoxide) ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (the bromo- 4- luorobenzyls of 3-) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) ammonia
Base)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) second
Epoxide) ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- methoxy-benzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) ammonia
Base)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) second
Epoxide) ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) second
Base) -3- methoxy benzamides.
In addition, the present invention includes pharmaceutical composition, said composition contains compound of Formula I and pharmaceutically acceptable figuration
Agent.The pharmaceutically acceptable excipient refers to any available for the diluent of drug field, adjuvant and or carrier.This
The compound of invention can be applied in combination with other active components, as long as they do not produce other detrimental effects, such as allergy is anti-
Should.
The present invention drug regimen can be configured to some formulations, wherein containing in drug field commonly use some excipient,
For example, oral formulations (such as tablet, capsule, solution or suspension)；Preparation (solution of such as injectable or the suspension of injectable
Agent, or the dried powder of injectable, adding water for injection before the injection can use immediately)；Topical formulations (such as ointment or
Solution).
Carrier for pharmaceutical composition of the present invention is available common type in drug field, including：Oral formulations
Adhesive, lubricant, disintegrant, cosolvent, diluent, stabilizer, suspending agent, pigment, flavouring etc.；Injectable formulation
Preservative, solubilizer, stabilizer etc.；The matrix of topical formulations, diluent, lubricant, preservative etc..Pharmaceutical preparation can
With by oral administration or parenteral (such as intravenous, subcutaneous, intraperitoneal or local) administration, if some drugses are in stomach condition
Under be unstable, enteric coated tablets can be configured to.
Screened by pressing down enzyme test and enzyme Degrading experiment in vitro, it has been found that this compound can suppress and degrade PLK1 and
BRD4 prolease activities.Therefore, the compounds of this invention can be used for the disease related to one expression of PLK1 and BRD4 proteinase activities
Application in disease, such as various cancers.
Screened by external activity, it has been found that the compounds of this invention has antitumor activity, therefore the compounds of this invention
Can be used for preparing treatment and or the various cancers of prevention medicine, such as breast cancer, colon cancer, prostate cancer, cancer of pancreas is non-small
Cell lung cancer, thyroid papillary carcinoma, oophoroma, melanoma or various leukaemia, particularly acute myeloid leukaemia.
The compounds of this invention can use as unique cancer therapy drug, or with one or more other antineoplastic Internet of Things
Close and use.Therapeutic alliance by each therapeutic component by simultaneously, sequentially or separating administration to realize.
The present invention is entered from thalidomide analogs on the basis of bibliography as in PROTACs with E3 ligases
The position that row combines, it had into Polo samples kinases 1 (PLK1) and bromine domain protein 4 in simultaneously from polyethylene glycol connects chain
(BRD4) structure of protease inhibitory activity, which is connected, builds PROTACs, while compound is directed to double target spots relative to single target
Point medicine can reduce the drug resistance of its antitumor activity.External PLK1 and BRD4 protease inhibiting activities, anti tumor activity in vitro
Test and external PLK1 and BRD4 albumen-degrading activity show that such targets the chemical combination of ubiquitination degraded PLK1 and BRD4 albumen
Thing (PROTACs) can degrade PLK1 and BRD4 targeting proteins, have good antitumor activity, and show excellent PLK1
With BRD4 inhibitory action.
Preparation method of the present invention is simple to operate, mild condition, and gained compound is respectively provided with PLK1 and BRD4 albumen enzyme levels
And degrading activity, antitumor action are notable.
Embodiment
Examples provided hereinafter and preparation example further elucidate and illustrated this compound and preparation method thereof.Should
Work as understanding, the scope that the scopes of following embodiments and preparation example is not limit the invention in any way.
Following synthetic route describes the preparation of the compound of Formula I of the present invention, and all raw materials are all to pass through these
Method described in route, prepared by organic chemistry filed method well-known to the ordinarily skilled artisan or it is commercially available.This hair
Bright whole final compounds are prepared by the method described in these routes or by similar method, these
Method is that organic chemistry filed is well-known to the ordinarily skilled artisan.The whole applied in these routes it is variable because
Definition in the definition of number following article or such as claim.
Reagent and condition:(a)SOCl2,MeOH；(b)NaBH(OAc)3,Na2CO3；(c)K2CO3,acetone；(d)Zn,
NH4Cl；(e)NaH,CH3I；(f)MeOH,HCl；(g)HBTU,DIPEA.
According to the compound of Formula I of the present invention, each substituent such as Summary is defined in synthetic route.
The preparation of intermediate 1
D-2- aminobutyric acid 10g, 100mL methanol is added, is then added dropwise to thionyl chloride 24.2mL, back flow reaction 2h.Subtract
Pressure is evaporated, and adds methyl tertiary butyl ether(MTBE) 100mL, has solid generation, filtration drying obtains white solid 13.52g, yield 91%.
1H-NMR(CDCl3, 400MHz):δ1.09(t,3H,CH3),2.01-2.26(m,2H,CH2),3.68(s,3H,
CH3),4.02-4.25(m,1H,CH),8.72(s,2H,NH2)。
The synthesis of intermediate 2
Intermediate 1 5g, benzaldehyde derivative 33mmol are added in dichloromethane 50mL, and triacetyl oxygen is added under ice bath
Base sodium borohydride 10.4g, 24h is reacted at room temperature, adds saturated sodium bicarbonate aqueous solution 50mL, dichloromethane extraction, merge organic phase,
Anhydrous magnesium sulfate is dried, and filtering is evaporated to obtain weak yellow liquid intermediate 2.
The preparation of intermediate 3
Intermediate 2 54mmol, K2CO34.0g is added in acetone 80mL, and the chloro- 5- nitro-pyrimidines of 2,4- bis- are added at 0 DEG C
5.6g acetone soln 20mL, 24h is reacted at room temperature, filtering, is evaporated, silica gel column chromatography (ethyl acetate：Petroleum ether 1：20) in, obtaining
Mesosome 3.
The preparation of intermediate 4
Intermediate 3 (59mmol), reduction zinc powder (28g, 7mmol), NH4Cl (9.3g, 175mmol) adds methanol 200mL
In, it is heated to reflux 24h. filterings and is evaporated, silica gel column chromatography (ethyl acetate：Petroleum ether 1：4) intermediate 4 is obtained.
The preparation of intermediate 5
Intermediate 4 (6g, 22mmol), iodomethane (3.4g, 24mmol) add DMF 50mL, 60% hydrogen are added at -15 DEG C
Change sodium (1.2g, 30mmol), react at room temperature 3h, add frozen water 100mL, ethyl acetate extraction, be evaporated, silica gel column chromatography (acetic acid
Ethyl ester：Petroleum ether 1：4) intermediate 5 is obtained.
The synthesis of intermediate 6
Intermediate 5 (2.13mg, 7.2mmol) and 4- amino -3- methoxy benzoic acids (1.88mg, 11.2mmol) add
3mL methanol 12mL water and 1.50mL concentrated hydrochloric acids, flow back 48h. evaporated under reduced pressure solvents, and methanol and Diethyl ether recrystallization obtain intermediate 6.
The preparation of intermediate 7
The fluoro- 2- of poly- second glycol diamines 0.41mmol, DIPEA105mg, 4- (2,6- dioxopiperidin -3- bases) second indoles -1,3-
Ketone 112.5mg, DMF 2mL are added, 90 DEG C of reaction 12h. are cooled to room temperature, add water 20mL, ethyl acetate extraction, extract saturation
Saline solution is cleaned, and anhydrous magnesium sulfate is dried, evaporated under reduced pressure, silica gel column chromatography (methanol：Dichloromethane 1；10) intermediate 7 is obtained.
Embodiment 1：Compound 4- (((S) -8- benzyl -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -2-
Base) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethoxies
Base) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8a) preparation
Compound 6a 0.012mmol are added in 0.250ml of DMF, then add HBTU 4.9mg, and DIPEA4.1 μ
L, intermediate 7a 0.014mmol react at room temperature 3h；Add 50mL water, acetoacetic ester extraction, extract saturated aqueous common salt cleans, anhydrous
Magnesium sulfate is dried, evaporated under reduced pressure, silica gel column chromatography (methanol：Dichloromethane 1：10) compound 8a, yield 67% are obtained.
IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.82(m,2H,CH2),2.00-2.06(m,
1H,CH)2.52-2.64(m,2H,CH×2),2.86-3.03(m,3H,CH2,CH),3.25-3.34(m,5H,CH2, CH3),
3.54–3.70(m,12H,CH2), 3.92 (s, 3H, CH3),4.29(t,1H,CH),4.43(d,1H,CH),5.05(dd,1H,
CH), 5.14 (d, 1H, CH), 7.02 (d, 1H, ArH), 7.08 (d, 1H, ArH), 7.15-7.39 (m, 6H, ArH), 7.58 (dd,
1H, ArH), 7.62 (dd, 1H, ArH), 7.87 (s, 1H, ArH), 7.94 (d, 1H, ArH)；LC-MS m/z:878.4[M+H]+。
According to the preparation method of embodiment 1, appropriate raw material, the embodiment 2-10 of system compound are selected
Embodiment 2：4- (((S) -8- (4- luorobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -2-
Base) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethoxies
Base) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8b).
Yield 60%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.84(m,2H,CH2),
2.02-2.08(m,1H,CH)2.52-2.66(m,2H,CH×2),2.86-3.04(m,3H,CH2,CH),3.28-3.36(m,
5H,CH2, CH3),3.54–3.76(m,12H,CH2), 3.92 (s, 3H, CH3),4.30(t,1H,CH),4.46(d,1H,CH),
5.06(dd,1H,CH),5.18(d,1H,CH),7.02-7.06(m,3H,ArH),7.08(d,1H,ArH),7.28-7.32(m,
2H, ArH), 7.58-7.69 (m, 3H, ArH), 7.84 (s, 1H, ArH), 7.92 (d, 1H, ArH)；LC-MS m/z:896.4[M+
H]+。
Embodiment 3:4- (((S) -8- (4- chlorobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -2-
Base) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethoxies
Base) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8c).
Yield 62%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.82(m,2H,CH2),
2.02-2.06(m,1H,CH)2.56-2.64(m,2H,CH×2),2.86-3.04(m,3H,CH2,CH),3.25-3.36(m,
5H,CH2, CH3),3.56–3.68(m,12H,CH2), 3.94 (s, 3H, CH3),4.29(t,1H,CH),4.45(d,1H,CH),
5.06(dd,1H,CH),5.17(d,1H,CH),7.02(d,1H,ArH),7.08(d,1H,ArH),7.18-7.25(m,4H,
ArH), 7.54-7.58 (m, 2H, ArH), 7.62 (dd, 1H, ArH), 7.86 (s, 1H, ArH), 7.94 (d, 1H, ArH)；LC-MS
m/z:912.3[M+H]+
Embodiment 4:4- (((S) -8- (4- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -2-
Base) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethoxies
Base) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8d)
Yield 58%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.82(m,2H,CH2),
2.00-2.06(m,1H,CH)2.52-2.64(m,2H,CH×2),2.92-3.02(m,3H,CH2,CH),3.25-3.34(m,
5H,CH2, CH3), 3.58-3.70 (m, 12H, CH2), 3.92 (s, 3H, CH3),4.32(t,1H,CH),4.43(d,1H,CH),
5.06(dd,1H,CH),5.14(d,1H,CH),7.02(d,1H,ArH),7.08(d,1H,ArH),7.31(t,1H,ArH),
7.49-7.58 (m, 5H, ArH), 7.64 (d, 1H, ArH), 7.87 (s, 1H, ArH), 7.98 (d, 1H, ArH)；LC-MS m/z:
956.3[M+H]+
Embodiment 5:4- (((S) -8- (3- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -2-
Base) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethoxies
Base) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8e)
Yield 67%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.82(m,2H,CH2),
2.02-2.08(m,1H,CH)2.54-2.64(m,2H,CH×2),2.86-3.02(m,3H,CH2,CH),3.26-3.34(m,
5H,CH2, CH3),3.54–3.72(m,12H,CH2), 3.92 (s, 3H, CH3),4.30(t,1H,CH),4.43(d,1H,CH),
5.08(dd,1H,CH),5.16(d,1H,CH),7.02(d,1H,ArH),7.08(d,1H,ArH),7.16-7.27(m,2H,
ArH), 7.34-7.39 (m, 1H, ArH), 7.48 (s, 1H, ArH), 7.53-7.56 (m, 2H, ArH), 7.61 (dd, 1H, ArH),
7.87 (s, 1H, ArH), 7.96 (d, 1H, ArH)；LC-MS m/z:956.3[M+H]+
Embodiment 6:4- (((S) -8- (2- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -2-
Base) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethoxies
Base) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8f)
Yield 70%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.79-1.84(m,2H,CH2),
2.01-2.06(m,1H,CH)2.52-2.64(m,2H,CH×2),2.86-3.05(m,3H,CH2,CH),3.26-3.34(m,
5H,CH2, CH3), 3.54-3.72 (m, 12H, CH2), 3.92 (s, 3H, CH3),4.32(t,1H,CH),4.46(d,1H,CH),
5.08(dd,1H,CH),5.16(d,1H,CH),7.02(d,1H,ArH),7.08(d,1H,ArH),7.14-7.18(m,1H,
ArH), 7.27-7.32 (m, 1H, ArH), 7.39-7.42 (m, 1H, ArH), 7.51-7.62 (m, 4H, ArH), 7.87 (s, 1H,
ArH), 7.94 (d, 1H, ArH)；LC-MS m/z:956.3[M+H]+
Embodiment 7:4- ((talk endlessly by (S) -8- (the chloro- 4- luorobenzyls of 3-) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydrochysenes
Pyridine -2- bases) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino)
Ethyoxyl) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8g)
Yield 64%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.82(m,2H,CH2),
2.00-2.06(m,1H,CH)2.52-2.64(m,2H,CH×2),2.86-3.03(m,3H,CH2,CH),3.25-3.34(m,
5H,CH2, CH3), 3.54-3.70 (m, 12H, CH2), 3.92 (s, 3H, CH3),4.30(t,1H,CH),4.43(d,1H,CH),
5.05(dd,1H,CH),5.14(d,1H,CH),7.02-7.08(m,3H,ArH),7.16-7.19(m,1H,ArH),7.34-
3.78 (m, 1H, ArH), 7.54-7.56 (m, 2H, ArH), 7.61 (dd, 1H, ArH), 7.86 (s, 1H, ArH), 7.94 (d, 1H,
ArH)；LC-MS m/z:930.3[M+H]+
Embodiment 8:4- ((talk endlessly by (S) -8- (the bromo- 4- luorobenzyls of 3-) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydrochysenes
Pyridine -2- bases) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino)
Ethyoxyl) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8h)
Yield 58%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.84(m,2H,CH2),
2.02-2.08(m,1H,CH)2.52-2.64(m,2H,CH×2),2.86-3.06(m,3H,CH2,CH),3.26-3.34(m,
5H,CH2, CH3),3.52-3.70(m,12H,CH2), 3.92 (s, 3H, CH3),4.31(t,1H,CH),4.46(d,1H,CH),
5.05(dd,1H,CH),5.17(d,1H,CH),7.03-7.08(m,3H,ArH),7.22-7.26(m,1H,ArH),7.54-
7.58 (m, 3H, ArH), 7.64 (dd, 1H, ArH), 7.87 (s, 1H, ArH), 7.94 (d, 1H, ArH)；LC-MSm/z:974.8[M
+H]+
Embodiment 9:4- (((S) -8- (4- methoxy-benzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -
2- yls) amino)-N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) second
Epoxide) ethyoxyl) ethyoxyl) ethyl) -3- methoxy benzamides (8i)
Yield 70%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.78-1.82(m,2H,CH2),
2.00-2.06(m,1H,CH)2.52-2.64(m,2H,CH×2),2.86-3.06(m,3H,CH2,CH),3.26-3.34(m,
5H,CH2, CH3),3.54-3.70(m,12H,CH2), 3.92 (s, 3H, CH3),4.29(t,1H,CH),4.43(d,1H,CH),
5.05(dd,1H,CH),5.16(d,1H,CH),6.72-6.77(d,2H,ArH),7.02(d,1H,ArH),7.08(d,1H,
ArH), 7.11-7.13 (d, 2H, ArH), 7.53-7.56 (m, 2H, ArH), 7.62 (dd, 1H, ArH), 7.84 (s, 1H, ArH),
7.92 (d, 1H, ArH)；LC-MS m/z:908.4[M+H]+
Embodiment 10:4- (((S) -8- (4- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridines -2-
Base) amino)-N- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides (8j)
Yield 63%,IH NMR(400MHz,CDCl3)δ：0.76(t,3H,CH3),1.76-1.80(m,2H,CH2),
2.02-2.08(m,1H,CH)2.52-2.64(m,2H,CH×2),2.86-3.03(m,3H,CH2,CH),3.25-3.34(m,
5H,CH2, CH3),3.54–3.70(m,8H,CH2),3.94(3H,s,O-CH3),4.29(t,1H,CH),4.43(d,1H,CH),
5.05(dd,1H,CH),5.14(d,1H,CH),7.02(d,1H,ArH),7.08(d,1H,ArH),7.31(t,1H,ArH),
7.49-7.58 (m, 5H, ArH), 7.64 (d, 1H, ArH), 7.87 (s, 1H, ArH), 7.94 (d, 1H, ArH)；LC-MS m/z:
912.3[M+H]+。
Embodiment 11：PLK1 protein inhibiting activities determine
Inhibitory activity of the compound to PLK1 albumen is determined with ELSIA methods, it is pure to sequentially add PLK1 in 96 hole microplate holes
Change albumen, the various concentrations inhibitor of 2.5 μ L DMSO dilutions, 150mM substrates, 10mM ATP, 90 μ L reaction buffers, simultaneously
Blank and Background control group are set up, is incubated at room temperature 30min.Board-washing, 100 μ L PPT-07 are added, be incubated at room temperature 30min.Board-washing,
The secondary antibody of the horseradish connection of 100 μ L anti-rabbit is added in per hole, is incubated 30min at room temperature.Board-washing, 100ul colour developings are often added in hole
Agent, 15min is incubated at room temperature.100ul reaction terminating liquids are added per hole, with exciting light 450nm, each hole fluorescence is determined and counts, meter
Calculate inhibiting rate IC50。
Embodiment 12：BRD4 protein inhibiting activity assay methods
BRD4bromodomain 1TR-FRET Assay Kit are passed through using time-resolved fluorescence (TR-FRET) method
(Cayman Chemical, USA) determines inhibitory activity of the compound to BRD4 albumen.The various concentrations that 10uL DMSO are dissolved
Inhibitor add in 384 microwell plates, add 5uL BRD4 containing 10nM albumen reaction buffer.Incubation at room temperature 15 minutes.
5uL Ac-H4 peptides are then added per hole and TR-FRET detection reagents [Anti-6His-XL665 and Streptavidin-Eu] are mixed
Liquid is closed, 1h is incubated at room temperature in dark.Measure the fluorescent emission at 620nm and 665nm after being excited under 330-350nm.Will be
Instruction of the ratio of transmitting at 665nm and 622nm as the amount of the BRD4/Ac-H4 compounds formed, calculate compound pair
The inhibitory activity IC of BRD4 albumen50。
Embodiment 13：The foundation of mtt assay measure tumor cell proliferation inhibition activity method and compound activity measure
The tumour cell (HepG2, A549, HeLa, MV4-11) to be tested in cell log growth period is pressed
Certain cell concentration is inoculated in culture plate, cultivates 24h, adds various concentrations inhibitor, cell is in 37 DEG C, 5%CO2Under the conditions of
Continue culture 48 hours, 20ul MTT solution is added per hole and continues culture 4 hours, it is dissolving crystallized with DMSO, examined with enzyme linked immunological
Survey instrument and its OD value calculating IC is determined at 570nm wavelength50。
Embodiment 14：Western-blot determines PLK1 and BRD4 protein degradation methods
Hela the or MV4-11 cells that pharmaceutical intervention is crossed are collected, and are washed 2 times with the PBS of precooling, and PMSF and RIPA is cracked
Liquid is with 1：100 ratio mixing, cell lysis 20min, 12000r/min × 20min centrifugations, takes supernatant, i.e. cell by 4 DEG C on ice
Total protein, protein content is detected with BCA standard measures, with 100 DEG C of denaturation 5min after 5 × albumen sample-loading buffer diluted protein.Albumen
It is separated by electrophoresis in SDS-PAGE, transferring film, closes 2h, 4 DEG C of overnight incubations of primary antibody.TBST washes film, secondary antibody 1:1000 are incubated 2h, change
Learn luminous rear X film developments, with the gray value of each band of Image J software analysis, inhibitor when calculating protein degradation 50%
Concentration DC50。
| Compound number | HeLa PLK1 DC50(nM) | MV4-11 BRD4 DC50(nM) |
| 8a | 83.4 | 100.5 |
| 8b | 12.3 | 32.2 |
| 8c | 56.7 | 96.8 |
| 8d | 9.6 | 13.8 |
| 8e | 126.9 | 342.7 |
| 8f | 87.6 | 131.2 |
| 8g | 21.6 | 36.8 |
| 8h | 19.3 | 24.7 |
| 8i | 129.3 | 284.2 |
| 8j | 67.9 | 94.3 |
Claims (9)
1. compound and its pharmaceutically acceptable salt shown in below formula：
Wherein R1, R2, R3It is independently selected from H, halogen or C1-C4 alkoxies；N is 2 or 3.
2. compound according to claim 1 and its pharmaceutically acceptable salt, are selected from：
4- (((S) -8- benzyl -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2- (2- (2-
(2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethyoxyl) second
Base) -3- methoxy benzamides；
4- (((S) -8- (4- luorobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2-
(2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethoxy
Base) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- chlorobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2-
(2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethoxy
Base) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2-
(2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethoxy
Base) ethyl) -3- methoxy benzamides；
4- (((S) -8- (3- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2-
(2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethoxy
Base) ethyl) -3- methoxy benzamides；
4- (((S) -8- (2- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2-
(2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethoxy
Base) ethyl) -3- methoxy benzamides；
4- (((S) -8- (the chloro- 4- luorobenzyls of 3-) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (the bromo- 4- luorobenzyls of 3-) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- methoxy-benzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino) -
N- (2- (2- (2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl)
Ethyoxyl) ethyl) -3- methoxy benzamides；
4- (((S) -8- (4- bromobenzyls) -7- ethyl -5- methyl -6- oxygen -5,6,7,8- tetrahydropteridine -2- bases) amino)-N- (2-
(2- (2- ((2- (2,6- dioxopiperidin -3- bases) -1,3- dioxy iso-indoles -4- bases) amino) ethyoxyl) ethyoxyl) ethyl) -3-
Methoxy benzamide.
3. the compound of any one and its pharmaceutically acceptable salt are being prepared and PLK1 albumen and BRD4 in claim 1-2
Application in the medicine of protein active unconventionality expression relevant disease.
4. the compound of any one and its pharmaceutically acceptable salt answering in antineoplastic is prepared in claim 1-2
With.
A kind of 5. pharmaceutical composition, it is characterised in that：Compound comprising any one in claim 1-2 and its pharmaceutically
Acceptable salt and pharmaceutically acceptable excipient.
6. pharmaceutical composition as claimed in claim 5 is being prepared and PLK1 albumen and BRD4 protein active unconventionality expression relevant diseases
Medicine in application.
7. application of the pharmaceutical composition as claimed in claim 5 in antineoplastic is prepared.
8. medicine group described in the compound and its pharmaceutically acceptable salt and claim 5 of any one in claim 1-2
Close and preparing treatment/prevention breast cancer, colon cancer, prostate cancer, cancer of pancreas, non-small cell lung cancer, thyroid papillary carcinoma, ovum
Application in the medicine of nest cancer, melanoma or leukaemia.
9. application as claimed in claim 8, wherein leukaemia is acute myeloid leukaemia.
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| CN106977584B (en) * | 2017-04-19 | 2019-12-06 | 吉林大学 | Compound for targeted ubiquitination degradation of PLK1 and BRD4 proteins and application thereof |
| EP4310496A2 (en) * | 2017-07-12 | 2024-01-24 | Dana-Farber Cancer Institute, Inc. | Compounds for tau protein degradation |
| CN109879877B (en) * | 2019-03-04 | 2021-08-10 | 吉林大学 | Compound capable of degrading PLK1 and BRD4 proteins and application thereof |
| CN110642849B (en) * | 2019-10-08 | 2021-05-28 | 中南大学湘雅医院 | Protein degradation targeting chimera and preparation method and application thereof |
| CN110845485B (en) * | 2019-12-02 | 2021-05-28 | 中南大学湘雅医院 | Protein degradation targeting chimera and preparation method and application thereof |
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| KR102337029B1 (en) | 2020-03-27 | 2021-12-09 | (주) 업테라 | PLK1 Selective Degradation Inducing Compound |
| CN112957357B (en) * | 2021-03-01 | 2022-05-27 | 中国医科大学附属第一医院 | Target KLF4 ubiquitination small molecule inhibitor and application thereof |
| KR20230004328A (en) | 2021-06-30 | 2023-01-06 | (주) 업테라 | Novel PLK1 Protein Degradation Inducing Compounds |
| CA3228601A1 (en) * | 2021-08-10 | 2023-02-16 | Soo Hee RYU | Novel plk1 degradation inducing compound |
| WO2023205701A1 (en) | 2022-04-20 | 2023-10-26 | Kumquat Biosciences Inc. | Macrocyclic heterocycles and uses thereof |
| CN115650975A (en) * | 2022-08-23 | 2023-01-31 | 四川大学华西医院 | Protein degradation targeting chimeric compound for targeted degradation of human epidermal growth factor receptor 2 and application thereof |
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