CN100441582C - Folid acid derivatives and their salts for preparing antitumor medicine - Google Patents
Folid acid derivatives and their salts for preparing antitumor medicine Download PDFInfo
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- CN100441582C CN100441582C CNB2006101482327A CN200610148232A CN100441582C CN 100441582 C CN100441582 C CN 100441582C CN B2006101482327 A CNB2006101482327 A CN B2006101482327A CN 200610148232 A CN200610148232 A CN 200610148232A CN 100441582 C CN100441582 C CN 100441582C
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Abstract
The present invention discloses a kind of folic acid derivatives and their salts for preparing antitumor medicine. The present invention discloses the application of these folic acid derivatives and their salts in preparing medicine for treating cancers. These folic acid derivatives and their salts have powerful tumor cell growth inhibiting activity.
Description
Technical field
The present invention relates to a class folic acid derivatives and a salt thereof, can be used for preparing the medicine for the treatment of cancer, belong to chemistry and medical technical field.
Background technology
Reduced form folic acid (FH
4) be that biosynthesizing of purine class and deoxyuridylic acid (dUMP) change into essential one carbon unit transporter in thymidylic acid (dTMP) process.In these biochemical reactions, reduced form folic acid is oxidized, is regenerated as active reduced form folic acid through Tetrahydrofolate dehydrogenase (DHFR).
Methotrexate (MTX) and similar compound are known consumingly in conjunction with the medicine of DHFR, these medicines can suppress dihydrofolate reduction and become tetrahydrofolic acid (THFA), cause dTMP and/or purine class defective, and DNA is synthetic to interrupt, and then the generation cytotoxicity, cause necrocytosis.These medicines have been developed into cancer therapy drug, and have occupied very important position as clinical medicine at present.
Pemetrexed (MTA) is the antifolic thing of many target spots of researching and developing recently, comprise DHFR by suppressing at least 3 kinds of enzymes synthetic relevant with folic acid metabolism, purine and pyrimidine, thymidylate synthase (TS), phosphoribosyl glycinamide formyl transferase (GARFT) has antitumous effect to multiple solid tumor.Pemetrexed (MTA) obtains the FDA approval U.S.'s listing, the mesothelioma of pleura patient that being used for the treatment of to perform the operation maybe should not carry out operation in April, 2004.The clinical test results of announcing in " the clinical magazine of oncology " on May 1 shows: and taxotere (Taxotere, present nonsmall-cell lung cancer standard two wires medicine) compares, LY231514 shows close patient's survival rate effect, but security is better, and side effect still less.Li Lai company submits the new drug application of LY231514 as nonsmall-cell lung cancer two wires medicine to U.S. FDA, various countries carry out clinical at tumour comprise multiple noumenal tumours such as mammary cancer, colorectal carcinoma, carcinoma of the pancreas, melanoma, bladder cancer, so its development and application prospect is boundless.
TNP-351 is a kind of DHFR inhibitor of new texture type, usually the pteridine structure in the antifolic thing is by 2,4-diamino pyrrolo-[2,3-d] pyrimidine replaces, and introduced the big trimethylene bridge of plasticity-in the 5-position, so that best conformation to be provided, hydrogen bond and hydrophobic bond interact to greatest extent with reactive site in target enzyme cavity to satisfy inhibitor.In vitro tests shows that the effect of these product anti-KB people's epidermoid cell and A549 people's non-small cell lung cancer cell is stronger more than 10 times than MTX; The in vivo test result shows that TNP-351 has drug-fast tumour cell to MTX very strong activity is also arranged, and Japan carries out clinical at present.
MTX is mainly anticancer by the folic acid antagonist effect, and stronger toxicity is arranged, and is little to the solid tumor effect, also exists the resistance of tumour cell to this medicine, therefore can not obtain satisfactory therapeutic effects.Therefore the mechanism of action that special expectation exploitation makes new advances demonstrates the cancer therapy drug of selective toxicity to cancer cells.
Summary of the invention
The object of the present invention is to provide a class folic acid derivatives and salt thereof to be used for antineoplaston, described this class folic acid derivatives and salt thereof will solve cancer therapy drug in the prior art strong toxicity,, technical problem that tumour cell to cancer therapy drug develop immunity to drugs limited to the solid tumor effect.
This class folic acid derivatives of the present invention and salt thereof are the compounds of following general formula (I) expression,
General formula (I)
R wherein
1=OH or NH
2R
2=H or CH
3Or CH
2CH
3R
3=H, or metallic cation, or ammonium ion, or organic amine positively charged ion; That is:
Work as R
1=OH, R
2=H, R
3During=H, that is: N-(4-{[(2-amino-4-hydroxy-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-methyl }-phenylformic acid)-L-L-glutamic acid,
Work as R
1=OH, R
2=CH
3, R
3During=H, that is: N-(4-{[(2-amino-4-hydroxy-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl }-phenylformic acid)-L-L-glutamic acid,
Work as R
1=OH, R
2=CH
2CH
3, R
3During=H, that is: N-(4-{[(2-amino-4-hydroxy-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl }-phenylformic acid)-L-L-glutamic acid,
Work as R
1=NH
2, R
2=H, R
3During=H, that is: N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-methyl }-phenylformic acid)-L-L-glutamic acid,
Work as R
1=NH
2, R
2=CH
3, R
3During=H, that is: N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl }-phenylformic acid)-L-L-glutamic acid,
Work as R
1=NH
2, R
2=CH
2CH
3, R
3During=H, that is: N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl }-phenylformic acid)-L-L-glutamic acid.
Further, described salt is sodium salt or sylvite or other metal cation salts that is suitable for or ammonium salt or diethyl amine salt or ethylenediamine salt or other organic amine salts that is suitable for.
The present invention also provides above-mentioned this class folic acid derivatives and the application of salt in the medicine of preparation treatment cancer thereof.
The present invention also provides a kind of this above-mentioned class folic acid derivatives and intermediate of salt thereof of preparing, and described intermediate is the compound of following general formula (II) expression,
General formula (II)
R wherein
1=OH or NH
2, R
2=H, or CH
3, or CH
2CH
3R
4Be lower alkoxy or benzyloxy or glutamate.
Further, lower alkoxy is selected from methoxyl group or oxyethyl group.
Further, glutamate is selected from dimethyl ester or diethyl ester.
Following synthetic route is with R
1=NH
2, R
2=CH
3And CH
2CH
3Be example, but be not limited only to this functional group.
Synthetic route: with methyl-formiate and chloromethyl cyanide is that raw material reaction makes 2-chloro-3-oxo-propionitrile, again with 2,4,6-Triaminopyrimidine Cheng Huanhou, with 4-aminomethyl-methyl benzoate. the hydrochloride condensation gets imine intermediate, with getting 4-{[(2 after the sodium cyanoborohydride reduction, 4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-methyl } methyl benzoate (3), behind methyl on formaldehyde and the sodium cyanoborohydride, hydrolysis, with the condensation reaction of L-glutamate diethyl ester, obtain N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino after the last hydrolysis]-methyl } phenylformic acid)-L-L-glutamic acid (7).(7) with after the diethylamine reaction get N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid two diethyl amine salt (8), (7) with after the sodium hydroxide reaction get N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid)-L-disodium glutamate salt (9).
Compound (3) with ethyl on acetaldehyde and the sodium cyanoborohydride after, hydrolysis, again with the condensation reaction of L-glutamate diethyl ester after, obtain N-(4-{[(2 after the last hydrolysis, 4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid (13).(13) with after the ammoniacal liquor reaction get N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid di-ammonium salts (14), (13) with after the potassium hydroxide reaction get N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid di-potassium (15).
Though unsymmetrical carbon is present in compound of the present invention and the intermediate thereof, unsymmetrical carbon has the absolute configuration of S (L) in the side chain of L-glutamic acid, and the absolute configuration of other asymmetric center can be the miscellany of S or R.In this case, there is diastereomer, and can separates by routine when needing and separate it with means of purification.
The present invention compares with prior art, and its technical progress is conspicuous.Used in the past anti-folic acid series antineoplastic medicament effect is more single, has grown duration of service all to produce resistance.This class folic acid derivatives of the present invention and salt thereof can have powerful inhibition growth activity to a plurality of tumour cells, be expected to overcome the resistance problem of antitumour drug deposits yields, the prospect that is advantageously applied to clinical treatment is arranged, also replenished the compound database of anti-folic acid series antineoplastic medicament simultaneously greatly.
Embodiment
This class folic acid derivatives of the present invention and salt thereof can be synthetic by several different methods, below with R
1=NH
2, R
2=CH
3And CH
2CH
3Be example, synthetic route of the present invention is described, but be not limited only to this functional group.
Step 1
2,4-diamino-5-cyano group-7 hydrogen-pyrrolo-[2,3-d] pyrimidines (1) synthetic
Reaction formula:
With sodium-acetate (13.1g 96mmol) joins in the 180ml distilled water, add again triaminopyrimidine (6.0g, 48mmol), heat temperature raising to 50 ℃, solid dissolving.Holding temperature slowly drips 2-chloro-3-oxo-propionitrile solution (40ml) at 50 ℃, and 1.5h adds.Solution becomes orange redly by colourless, becomes beige again.Continue stirring reaction 12h under this temperature, remaining THF in the pressure reducing and steaming solution is warming up to 100 ℃, back flow reaction 1h again.Have a small amount of beige solid to generate, some plate raw material reaction as can be known finishes.Stop heating, be cooled to room temperature, be evaporated to surplus less water, have a large amount of solids to separate out suction filtration.After filter cake washed with a large amount of distilled water (50mlx3), drying got grey black solid crude product 7.0g.Crude product must purer grey black product (5.0g, 60%) in methyl alcohol behind the recrystallization.Fusing point＞300 ℃ (document＞300 ℃).
1H NMR(DMSO-d
6，300MHz)δ：5.93(s，2H，2-NH
2)，6.26(s，2H，4-NH
2)，7.71(s，1H，6-CH)，11.95(s，1H，7-NH)。
EI-MS：m/z 174。
Step 2
4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-methylene)-amino]-methyl } methyl benzoate (2) synthetic
Reaction formula:
With 2, and 4-diamino-5-cyano group-7 hydrogen-pyrrolo-[2,3-d] pyrimidine (0.5g is 2.9mmol) with 4-aminomethyl-methyl benzoate. and (0.7g 3.5mmol) is dissolved into 100mlCH to hydrochloride
3Among the COOH/DMF (15%), add wet Raney's nickel 2.0g again, at the hydrogen pressure of 0.6Mpa, under 36 ℃, stirring reaction 6h.Behind the 6h, the visible raw material reaction of some plate finishes.Stopped reaction after the cooling, is poured reaction solution in one beaker into, adds the 4g silica-gel powder, suction filtration behind the mixing.Decompression steams solvent down, obtains a grey black solid crude product.Get off-white color solid phase prod (0.3g, 23.1%) after crossing column purification.Fusing point＞250 ℃.
1H NMR(DMSO-d
6，300MHz)δ：3.85(s，1H，4’COOCH
3)，4.77(s，2H，10-CH
2)，5.62(s，2H，2-NH
2)，6.58，8.95(s，2H，4-NH
2)，7.30(s，1H，6-CH)，7.45(d，2H，2’，6’-CH)，7.95(d，2H，3’，5’-CH)，8.32(s，2H，8-CH
2)，11.30(s，1H，7-NH)。
EI-MS：m/z 324。
Step 3
4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-methyl } methyl benzoate (3) synthetic
Reaction formula:
With 4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-methylene)-amino]-methyl } (0.8g 2.5mmol) joins in the 100ml methyl alcohol methyl benzoate, and (0.2g 3.5mmol), stirs under the room temperature to add sodium cyanoborohydride again.With 50% methyl alcohol/concentrated hydrochloric acid reaction solution pH is transferred to 2, solid is molten entirely.Behind the 10min, the adularescent solid generates, and continues stirring reaction 4h, and raw material reaction finishes.Stopped reaction filters, after filter cake washs with methylene dichloride (5mlx3), and oven dry.Obtain a white solid (0.57g, 70%).Fusing point 233-235 ℃.
1H NMR(DMSO-d
6，300MHz)δ：3.85(s，3H，4’-COOCH
3)，4.30(s，2H，10-CH
2)，4.43(s，2H，8-CH
2)，7.26(s，1H，6-CH)，7.33(s，1H，9-NH)，7.72(d，2H，2’，6’-CH)，7.98(d，2H，3’，5’-CH)，8.39(s，2H，2-NH
2)，9.60(s，2H，4-NH
2)，12.03(s，1H，7-NH)。
ESI-MS：m/z 327(MH+)。
Step 4
4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } methyl benzoate (4) synthetic
Reaction formula:
With 4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-methyl } (150mg 0.5mmol) joins 70ml1 to methyl benzoate, in the 4-dioxane, slowly feed formaldehyde gas (16.6mg, 0.6mmol), add again sodium cyanoborohydride (145mg, 2.3mmol), with 50% methyl alcohol/concentrated hydrochloric acid reaction solution pH is transferred to 2, stirring reaction 1h under the room temperature.Behind the 1h, the TLC demonstration still has the raw material residue, continues reaction 1h.Behind the 2h, some plate raw material reaction as can be known finishes.Stopped reaction, the evaporated under reduced pressure solvent adds 5ml water in residuum, with ammoniacal liquor reaction solution pH is transferred to 7, has a large amount of solids to separate out. filter the dry solid (0.12g, 76.7%) that gets.Fusing point 136-139 ℃.
1H NMR(DMSO-d
6，300MHz)δ：2.08(s，3H，9-N-CH
3)，3.53(s，2H，8-CH
2)，3.59(s，2H，10-CH
2)，3.84(s，3H，COOCH
3)，5.65(s，2H，2-NH
2)，6.66(s，1H，6-CH)，7.43(d，2H，2’，6’-CH)，7.92(d，2H，3’，5’-CH)，10.64(s，1H，7-NH)。
EI-MS：m/z 340。
Step 5
4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid (5) synthetic
Reaction formula:
With 4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } (120mg 0.35mmol) is dissolved in the 20ml methyl alcohol methyl benzoate, adds 1N NaOH solution 3ml, heat temperature raising to 50 ℃, stirring reaction 10h.Behind the 10h, some plate raw material reaction as can be known finishes, stopped reaction, cooling, pressure reducing and steaming methyl alcohol.In residuum, add 3ml distilled water, reaction solution pH is transferred to 7, have a large amount of white solids to generate, filter, get white solid product (83mg, 72.7%) after the oven dry with dilute hydrochloric acid.Fusing point 202-204 ℃.
1H NMR(DMSO-d
6，300MHz)δ：2.08(s，3H，9-N-CH
3)，3.57(s，2H，8-CH
2)，3.62(s，2H，10-CH
2)，5.50(s，2H，2-NH
2)，6.63(s，1H，6-CH)，7.39(d，2H，2’，6’-CH)，7.89(d，2H，3’，5’-CH)，10.53(s，1H，7-NH)。
EI-MS：m/z 326。
Step 6
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid)-L-glutamate diethyl ester (6) synthetic
Reaction formula:
With 4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } (110mg 0.34mmol) is dissolved among the 12ml DMF phenylformic acid, adds 4-methylmorpholine (40.4mg again, 0.40mmol) and 2-chloro-4,6-dimethoxy-1,3,5-triazines (70.2mg, 0.40mmol), N
2Protection, stirring reaction 2h under the room temperature.Behind the 2h, (40.4mg, 0.40mmol) (105.8mg 0.44mmol), at room temperature continues stirring reaction 3h with the L-diethyl glutamate hydrochloride to add the 4-methylmorpholine again.Behind the 3h, the some plate still has the raw material residue as can be known, continues reaction.After 2h, raw material reaction finishes, stop to stir, and the evaporated under reduced pressure solvent, residuum is crossed column purification.(eluent: methylene dichloride: methyl alcohol=40: 1-20: 1).Get transparence sclerosis product liquid (150mg, 86.3%).
1H NMR(DMSO-d
6，300MHz)δ：1.20(2t，6H，COO-CH
2C
H 3)，2.01(m，2H，β-CH
2)，2.10(s，3H，9-N-CH
3)，2.43(m，2H，γ-CH
2)，3.56(s，2H，8-CH
2)，3.65(s，2H，10-CH
2)，4.14(m，4H，COO-C
H 2CH
3)，4.42(m，1H，glu-NH-C
H)，6.88(s，1H，6-CH)，7.07(s，2H，2-NH
2)，7.40(d，2H，2’，6’-CH)，7.87(d，2H，3’-，5’-CH)，8.80(d，1H，glu-NH)，11.57(s，1H，7-NH)。
ESI-MS：m/z 512(MH+)。
Step 7
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid (7) synthetic
Reaction formula:
With N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid)-(63mg 0.12mmol) is dissolved among the 5ml THF L-glutamate diethyl ester, add 1N sodium hydroxide 1ml again, at room temperature stirring reaction 4h.Behind the 4h, the some plate still has a small amount of raw material residue as can be known, continues reaction 2h.Behind the 6h, TLC shows that raw material reaction finishes stopped reaction.Steam THF, reaction solution pH is transferred to 5, have solid to separate out with the dilute hydrochloric acid of 5N.Filtering, after the oven dry, is a beige solid crude product 6mg.After mother liquor concentrates again off-white color solid 15mg, altogether product (21mg, 38.5%).Fusing point＞250 ℃.
1H NMR(DMSO-d
6，300MHz)δ：1.98，2.05(m，2H，β-CH
2)，2.09(s，3H，9-N-CH
3)，2.35(t，2H，γ-CH
2)，3.54(s，2H，8-CH
2)，3.60(s，2H，10-CH
2)，4.40(m，1H，glu-NH-C
H)，6.03(s，2H，2-NH
2)，6.72(s，1H，6-CH)，7.38(d，2H，2’，6’-CH)，7.86(d，2H，3’，5’-CH)，8.58(d，1H，glu-NH)，10.88(s，1H，7-NH)。
13C NMR(DMSO-d
6，400MHz)δ：26.715，31.185，41.423，52.763，54.611，60.917，95.805，113.165，118.126，128.212，129.816，133.573，141.856，153.146，157.384，157.555，166.982，174.381，174.702。
HRESI-MS calcd for C
21H
26N
7O
5(MH+)456.1995，found 456.1986。
Step 8
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid two diethyl amine salt (8) synthetic
Reaction formula:
Compound 7 (33.0mg, 73 μ mol) joins in the flask, adds distilled water (2.5ml) again, diethylamine (11mg, 146 μ mol), and stirring reaction is 0.5 hour under the room temperature, boils off solvent under the decompression, and water-recrystallizing methanol obtains solid (24mg, 55%).
ESI-MS：m/z(Neg)226.5。
Step 9
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-methyl-amino]-methyl } phenylformic acid)-L-disodium glutamate salt (9) synthetic
Reaction formula:
Compound 7 (33.0mg, 73 μ mol) joins in the flask, adds distilled water (1.5ml) again, 1N NaOH solution (0.15ml), and stirring reaction is 0.5 hour under the room temperature, boils off solvent under the decompression, and water-ethyl alcohol recrystallization obtains solid (22mg, 60%).
ESI-MS：m/z 226.5(Neg)。
Step 10
4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } methyl benzoate (10) synthetic
Reaction formula:
With 4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-methyl } (50mg 0.15mmol) joins in the 13ml methyl alcohol methyl benzoate, adds 40% acetaldehyde solution (18.6mg, 0.17mmol), add again sodium cyanoborohydride (30mg, 0.45mmol), reaction solution becomes clarification, with 50% methyl alcohol/concentrated hydrochloric acid reaction solution pH is transferred to 2, stirring reaction 3h under the room temperature.Behind the 3h, some plate raw material reaction as can be known finishes.Stopped reaction, the evaporated under reduced pressure solvent adds 5ml water in residuum, with ammoniacal liquor reaction solution pH is transferred to 7, has a large amount of solids to separate out. filter, dry solid crude product 0.59g, cross behind the column purification white product (0.34g, 64%).Fusing point 104-107 ℃.
1H NMR(DMSO-d
6，300MHz)δ：1.00(t，3H，9-N-CH
2C
H 3)，2.46(m，2H，9-N-C
H 2CH
3)，3.54(s，2H，8-CH
2)，3.63(s，2H，10-CH
2)，3.84(s，3H，4’-COOCH
3)，5.48(s，2H，2-NH
2)，6.67(s，1H，6-CH)，6.85(s，2H，4-NH
2)，7.43(d，2H，2’，6’-CH)，7.90(d，2H，3’，5’-CH)，10.57(s，1H，7-NH)。
ESI-MS：m/z 377(MNa+)。
Step 11
4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid (11) synthetic
Reaction formula:
With 4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-ethyl-methyl } (100mg 0.28mmol) is dissolved among the 5ml THF methyl benzoate, adds 1N NaOH solution 1.7ml, heat temperature raising to 50 ℃, stirring reaction 7h.Behind the 7h, some plate raw material reaction as can be known finishes, stopped reaction, cooling, pressure reducing and steaming THF.In residuum, add 2ml distilled water, reaction solution pH is transferred to 7, have a large amount of white solids to generate, filter, get solid phase prod (60mg, 63%) after the oven dry with acetic acid.Fusing point 196-198 ℃ dec.
1H NMR(DMSO-d
6，300MHz)δ：1.01(t，3H，9-N-CH
2C
H 3)，2.45(m，2H，9-N-C
H 2CH
3)，3.55(s，2H，8-CH
2)，3.63(s，2H，10-CH
2)，5.53(s，2H，2-NH
2)，6.65(s，1H，6-CH)，7.10(s，2H，4-NH
2)，7.40(d，2H，2’，6’-CH)，7.89(d，2H，3’，5’-CH)，10.55(s，1H，7-NH)。
ESI-MS：m/z 341(MH+)。
Step 12
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-L-glutamate diethyl ester (12) synthetic
Reaction formula:
With 4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-amino]-ethyl-methyl } (40mg 0.12mmol) is dissolved among the 5ml DMF phenylformic acid, adds 4-methylmorpholine (13.4mg again, 0.13mmol) and 2-chloro-4,6-dimethoxy-1,3,5-triazines (20mg, 0.13mmol), N
2Protection, behind the stirring reaction 2h.2h, (13.4mg, 0.13mmol) (38.5mg 0.16mmol), at room temperature continues stirring reaction 4h with the L-diethyl glutamate hydrochloride to add the 4-methylmorpholine again under the room temperature.Behind the 4h, the some plate still has the raw material residue as can be known, continues reaction.After 6h, raw material reaction finishes, stop to stir, and the evaporated under reduced pressure solvent, residuum is crossed column purification.(eluent: methylene dichloride: methyl alcohol=30: 1-20: 1).Get transparence sclerosis product liquid (60mg, 95%).
1H NMR(DMSO-d
6，300MHz)δ：1.02(t，3H，9-N-CH
2C
H 3)，1.23(2t，6H，COO-CH
2C
H 3)，2.01(m，2H，β-CH
2)，2.32，2.72(m，2H，γ-CH
2)，2.45(m，2H，9-N-C
H 2CH
3)，3.56(s，2H，8-CH
2)，3.65(s，2H，10-CH
2)，4.14(m，4H，COO-C
H 2CH
3)，4.42(m，1H，glu-NH-C
H)，5.56(s，2H，2-NH
2)，6.65(s，1H，6-CH)，7.10(s，2H，4-NH
2)，7.38(d，2H，2’-，6’-CH)，7.82(d，2H，3’-，5’-CH)，8.68(d，1H，glu-NH)，10.58(s，1H，7-NH)。
ESI-MS：m/z 526(MH+)。
Step 13
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid (13) synthetic
Reaction formula:
With N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-(55mg 0.1mmol) is dissolved among the 5ml THF L-glutamate diethyl ester, add 1N sodium hydroxide 0.6ml again, at room temperature stirring reaction 4h.Behind the 4h, raw material reaction finishes, stopped reaction.Steam THF, reaction solution pH is transferred to 4, have solid to separate out with Glacial acetic acid.Filter, after the oven dry, get a pale solid (14mg, 30%).Fusing point＞250 ℃.
1H NMR(DMSO-d
6，300MHz)δ：1.02(t，3H，9-N-CH
2C
H 3)，1.98，2.05(m，2H，β-CH
2)，2.35(t，2H，γ-CH
2)，2.46(m，2H，9-N-C
H 2CH
3)，3.55(s，2H，8-CH
2)，3.62(s，2H，10-CH
2)，4.44(m，1H，glu-NH-C
H)，5.51(s，2H，2-NH
2)，6.65(s，1H，6-CH)，7.10(s，2H，4-NH
2)，7.38(d，2H，2’，6’-CH)，7.82(d，2H，3’，5’-CH)，8.53(d，1H，glu-NH)，10.54(s，1H，7-NH)。
13C NMR(DMSO-d
6，300MHz)δ：10.545，26.284，30.746，46.014，50.658，52.221，56.516，95.733，111.957，116.950，127.543，129.167，132.946，141.870，154.042，158.276，159.490，166.411，173.650，174.105。
ESI-MS：m/z 470(MH+)。
HRESI-MS calcd for C
22H
28N
7O
5(MH+)470.2152，found 470.2135。
Step 14
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid di-ammonium salts (14) synthetic
Reaction formula:
Compound 13 (30mg, 64 μ mol) joins in the flask, adds distilled water (1.5ml) again, 1NNH
4OH solution (0.15ml), stirring reaction is 1 hour under the room temperature, boils off solvent under the decompression, and water-ethyl alcohol recrystallization obtains solid (19.3mg, 60%).
ESI-MS：m/z(Neg)233.5。
Step 15
N-(4-{[(2,4-diamino-7H-pyrrolo-[2,3-d] pyrimidine-5-ylmethyl)-ethyl-amino]-methyl } phenylformic acid)-L-L-glutamic acid di-potassium (15) synthetic
Reaction formula:
Compound 13 (30mg, 64 μ mol) joins in the flask, adds distilled water (1.5ml) again, 1N KOH solution (0.15ml), and stirring reaction is 0.5 hour under the room temperature, boils off solvent under the decompression, and water-ethyl alcohol recrystallization obtains solid (19.5mg, 56%).
ESI-MS：m/z(Neg)233.5。
Evidence, this class folic acid derivatives of the present invention and a plurality of tumour cells of salt pair thereof have powerful inhibition growth activity, can be used to prepare antitumor drug, and are expected to overcome the resistance problem of antitumour drug deposits yields.
Example: tetrazolium (microculturetetrozolium has been selected in the anti-tumor biological test of compound 13 for use, MTT) reduction method, to human leukemia cell line (CCRF-CEM), the inhibition activity of human lung carcinoma cell line (A549) and three kinds of cell strains of mouse leukemia cell strain (L1210) is tested respectively.
The anti-tumor biological of compound 13
| Compound | CCRF-CEM， IC 50(ug/ml) | L1210， IC 50(ug/ml) | A549， IC 50(ug/ml) |
| 13 | 25.67 | 28.11 | 8.38 |
From the inhibiting rate relation of drug level and growth of tumour cell and one IC that calculates according to regular Bliss method
50, can infer as drawing a conclusion:
1, compound 13 couples of CCRF-CEM, L1210 and A549 tumour cell all have restraining effect.Especially to A549, its activity is better than MTX (IC
5082.98ug/ml) about 10 times.
2, compound 13 shows that different tumour cells is had the activity of inhibition, and the prospect that is applied to clinical treatment is preferably arranged.
Claims (5)
1. a class folic acid derivatives and a salt thereof is characterized in that: be the represented compound of following general formula,
In general formula, R
1=OH or NH
2R
2=H or CH
3Or CH
2CH
3R
3=H or metallic cation or ammonium ion or organic amine positively charged ion.
2. a class folic acid derivatives and a salt thereof as claimed in claim 1 is characterized in that: described salt is metal cation salt or ammonium salt or organic amine salt.
3. the application of described class folic acid derivatives of claim 1 and salt thereof is characterized in that being used to prepare the medicine for the treatment of cancer.
4. intermediate for preparing described class folic acid derivatives of claim 1 and salt thereof, it is characterized in that: described intermediate is the compound of following general formula,
R wherein
1=OH or NH
2, R
2=H or CH
3Or CH
2CH
3R
4Be methoxyl group, oxyethyl group, benzyloxy or glutamate.
5. intermediate as claimed in claim 4 is characterized in that described glutamate is selected from dimethyl ester or diethyl ester.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6066732A (en) * | 1998-08-21 | 2000-05-23 | The Trustees Of Princeton University | Process for the preparation of pyrrolo[2,3-d]pyrimidines |
| CN1778797A (en) * | 2004-11-25 | 2006-05-31 | 重庆医药工业研究院有限责任公司 | Production of N-(pyrrolo[2,3-d] pyrimidine-5-) acyl glusate derivative and intermediate |
| CN1840530A (en) * | 2005-03-28 | 2006-10-04 | 齐鲁制药有限公司 | Process for preparing pemetrexed |
-
2006
- 2006-12-28 CN CNB2006101482327A patent/CN100441582C/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6066732A (en) * | 1998-08-21 | 2000-05-23 | The Trustees Of Princeton University | Process for the preparation of pyrrolo[2,3-d]pyrimidines |
| CN1778797A (en) * | 2004-11-25 | 2006-05-31 | 重庆医药工业研究院有限责任公司 | Production of N-(pyrrolo[2,3-d] pyrimidine-5-) acyl glusate derivative and intermediate |
| CN1840530A (en) * | 2005-03-28 | 2006-10-04 | 齐鲁制药有限公司 | Process for preparing pemetrexed |
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| A simple and Concise Synthesis of LY231514(MTA). Edward C. Taylor et. al.Tetrahedron Letters,Vol.40 . 1999 |
| A simple and Concise Synthesis of LY231514(MTA). Edward C. Taylor et. al.Tetrahedron Letters,Vol.40. 1999 * |
| Design and Synthesis of Histidine Analogues of Folic Acidand Methotrexate as Potential Folylpolyglutamate SynthetaseInhibitors. Zhenmin Mao et. al.Journal of Medicinal Chemistry,Vol.39 No.21. 1996 |
| Design and Synthesis of Histidine Analogues of Folic Acidand Methotrexate as Potential Folylpolyglutamate SynthetaseInhibitors. Zhenmin Mao et. al.Journal of Medicinal Chemistry,Vol.39 No.21. 1996 * |
| LY231514, a Pyrrolo[2,3-d]pyrimidine-based Antifolate ThatInhibits Multiple Flate-requiring Enzymes. Chuan Shih et. al.Cancer Research,Vol.57 . 1997 |
| LY231514, a Pyrrolo[2,3-d]pyrimidine-based Antifolate ThatInhibits Multiple Flate-requiring Enzymes. Chuan Shih et. al.Cancer Research,Vol.57. 1997 * |
| 培美曲塞二钠的合成. 朱高军等.中国医药工业杂志，第36卷第12期. 2005 |
| 培美曲塞二钠的合成. 朱高军等. 中国医药工业杂志，第36卷第12期. 2005 * |
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