CN101817829A - 4-substituted anilino-podophyllotoxine derivative and application - Google Patents
4-substituted anilino-podophyllotoxine derivative and application Download PDFInfo
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Abstract
The invention provides a 4-substituted anilino-podophyllotoxine derivative which refers to a 4'-O-demethyl-4-deoxy-4-substituted anilino-podophyllotoxine derivative. A series of compounds with anti-cancer activity is synthetized by taking the traditional drug etoposide as a lead compound by structural transformation; primary in vitro screening and in vivo antitumor tests show that the compounds have better anti-tumor activity, wherein when the compound GL3 improves the anti-tumor action, the toxicity of the compound GL3 is obviously reduced, and thereby, the compound GL3 can be used for preparing anti-tumor drugs which has higher antitumor activity and lower toxicity and has effect on multi-drug drug resistant tumors. The structural general formula of the 4-substituted anilino-podophyllotoxine derivative is shown in the specification.
Description
Technical field
The invention belongs to compound and purposes, relate generally to the preparation of 4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative, and the application in the preparation antitumor drug.
Background technology
Etoposide and teniposide are the antitumor drugs that uses clinically at present, and they all are the semi-synthetic derivatives of 4 '-O-demethyl epipodophyllotoxin.4 '-O-demethyl epipodophyllotoxin analog derivative can destroy the 26S Proteasome Structure and Function of DNA by suppressing the activity of DNA topoisomerase II, thus the performance antineoplastic action.
Although two above-mentioned compounds use for many years clinically, but still have poorly water-soluble, bone marrow depression is easy to generate problems such as resistance, thereby has limited its application.Therefore, in order to overcome above-mentioned shortcoming, seek the more podophyllotoxin analogue of high-efficiency low-toxicity, the investigator has carried out a series of structure of modification work to podophyllotoxin.
On the basis of the structure of summing up podophyllotoxin and activity relationship, people such as Claude Monneret in 1999 have proposed the structure activity relationship of podophyllotoxin analogue: a dioxolane is optimal selection; E ring rotate freely with 2 α, 3 beta comfigurations be active necessary; The anti-tumor activity of 4 beta comfigurations is apparently higher than 4 α configurations; 4 is effective decorating site, and the Etoposide glucosides partly is beneficial to the inhibition activity of raising compound to the DNA topoisomerase II with amino being substituted with, and causes the DNA chain break simultaneously.It should be noted that some 4 beta-aromatic amine podophyllotoxin derivatives not only have cytotoxic activity to the tumour cell of some Etoposide tolerances, can also reduce the expression or proteic mistake of reduction multidrug-resisting (MDR) of DNA topoisomerase II and express.
Summary of the invention
The purpose of this invention is to provide a class 4-substituted anilino-podophyllotoxine derivative, mainly refer to 4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative, have following general structural formula (I):
Wherein
X represents-(CH
2)
n-, n be 0～2 or-CH=CH-;
R
1, R
2, R
3, R
4, R
5Identical or different, R
1, R
2, R
3, R
4, R
5For hydrogen atom, halogen atom, hydroxyl, nitro, contain 1 to 6 carbon atom straight or branched alkyl or contain the alkoxyl group of the straight or branched of 1 to 3 carbon atom;
Another purpose of the present invention provides the application of 4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative in the preparation antitumor drug.Preliminary in-vitro screening finds that they to various tumor cell strains, comprise 95D, A549, KB, KB/VCR etc. all have and suppress proliferation function preferably, demonstrate the cytotoxic activity stronger than Etoposide, compound GL2 wherein, the effect of GL3 is particularly outstanding, to the IC of above-mentioned cell strain
50Value is between 0.42～16.43 μ M.Further the anti-tumor in vivo experiment is found, compound GL3 is under heavy dose of administration condition of 100mg/kg, inhibitory rate to 58.1%, tumour inhibiting rate than positive control medicine cis-platinum is high by 13.7%, this result shows that this compounds in the inhibiting while of improving tumour, can reduce its toxicity well.Restraining effect to the high transitivity lung cancer of people 95D Nude Mice is found the inhibitory rate to 57.3% of compound GL3.
Characteristics of the present invention are to utilize existing antitumor drug Etoposide as lead compound, pass through structure of modification, synthetic a series of compounds with anti-tumor activity, in the hope of find that anti-tumor activity is higher, toxicity is lower, to multidrug resistance tumour effective antitumour medicine.Press down the knurl experiment in preliminary in-vitro screening and the body and show that these compounds have better antitumor activity, wherein compound GL3 is when improving tumor inhibition effect, and its toxicity also obviously reduces.
Embodiment
The present invention is further described in conjunction with the embodiments.Following embodiment is that explanation is of the present invention, rather than limits the present invention by any way.
Embodiment 1 synthetic general formula of the present invention is:
Embodiment 2: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-to the methoxy benzamide base) anilino] epipodophyllotoxin (GL1)
The first step:(reference literature method: Francesco Troisi et al.Tetrahedron Letters 2007,48,7986-7989)
With 5.05g tert-Butyl dicarbonate (Boc
2O, 23mmol) be dissolved in the dioxane, under condition of ice bath, (Compound I in dioxane solution 46.3mmol), at room temperature stirs after dripping and spends the night to be added dropwise to the 5g Ursol D, then with the reaction soln concentrating under reduced pressure, with the remaining resistates of silica gel chromatography purifying (eluent: ethyl acetate/petroleum ether=7/3), get Compound I I, yield: 92.7%; Yellow solid, fusing point: 108.9-113.7 ℃.
Second step:
Under nitrogen protection, (compound III 8.5mmol) is dissolved in the 50mL anhydrous acetonitrile with 5.11g sodium iodide (34mmol) with the 5g Etoposide; In addition exsiccant 4.3mL trimethylchlorosilane is dissolved in the 30mL anhydrous acetonitrile, under condition of ice bath, slowly be added drop-wise to this solution in the reaction solution, after dropwising, stirring at room 1 hour, obtain intermediate 4 '-O-demethyl-4-deoxidation-4-iodo epipodophyllotoxin (compound IV), need not purifying, directly carry out the next step.
The 3rd step:
In the reaction solution of compound IV, add 5.1g Carbon Dioxide barium (34mmol), stirred 10 minutes, add anhydrous triethylamine reaction solution is transferred to alkalescence, add 2.12g Compound I I (10.2mmol), stirring at room 12～15 hours; With the reaction solution suction filtration, filtrate decompression concentrates, the resistates that usefulness silica gel chromatography purifying is left (eluent: ethyl acetate/petroleum ether=1/1), get compound V, yield: 85.7%, yellow solid, fusing point: 136.1～138.6 ℃,
1H NMR (500MHz, CDCl
3) δ 7.20-7.21 (m, 2H), 6.74 (s, 1H), 6.51 (s, 1H), 6.48-6.50 (d, J=9.0Hz, 2H), 6.36 (ws, 1H), 6.33 (s, 2H), 5.95and 5.97 (d, J=10.5Hz, 2H), 5.47 (ws, 1H), 4.61 (m, 1H), 4.57-4.58 (d, J=5.0Hz, 1H), 4.34-4.37 (t, J=8.0Hz, 1H), 3.96-4.00 (m, 1H), 3.79 (s, 6H), 3.13-3.16 (dd, J=5.0Hz, 1H), 2.94-3.01 (m, 1H).
The 4th step:
3.1g compound V (5.24mmol) is dissolved in the 30mL anhydrous ethyl acetate solution, exsiccant HCl gas is fed in the reaction solution, reacting below 0 ℃ 3 hours; With the reaction solution suction filtration, the solid that obtains is a compound VI, yield: 95.7%, and yellow solid, fusing point: 183.3～186.3 ℃,
1H NMR (500MHz, DMSO) δ 9.93 (s, 1H), 7.11-7.13 (m, 2H), 6.76-6.77 (m, 3H), 6.54 (s, 1H), 6.25 (m, 2H), 6.33 (s, 2H), 5.96and 5.99 (d, J=10.5Hz, 2H), 4.85 (m, 1H), 4.50-4.51 (d, J=5.0Hz, 1H), 4.33-4.35 (m, 2H), 3.64 (s, 6H), 3.27-3.28 (dd, J=5.0Hz, 1H), 2.96-3.03 (m, 1H).
The 5th step:
110mg compound VI (0.18mmol) is dissolved in the 5mL anhydrous methylene chloride solution, add triethylamine solution is transferred to alkalescence, stir moments later, adding 36mg I-hydroxybenzotriazole (HOBt, 0.27mmol), 28mg anisic acid (0.18mmol), and under ice bath, add 38mg 1-(3-dimethylamino-propyl)-3-ethyl-carbodiimide hydrochloride (EDCHCl, 0.20mmol), stir after 5 minutes room temperature reaction 5-8 hour; Reaction solution is poured in the water into extraction, organic phase anhydrous sodium sulfate drying; With the organic phase concentrating under reduced pressure, with the remaining resistates of silica gel chromatography (eluent: ethyl acetate/petroleum ether=1/1), obtain GL1, output 65mg, yield: 56.7%, yellow solid, fusing point: 143.8-145.7 ℃,
1HNMR (500MHz, CDCl
3) δ 7.82 (d, J=8.5Hz, 2H, 2 " ', 6 " '-H), 7.65 (s, 1H ,-CONH-), 7.44 (d, J=8.0Hz, 2H, 3 "; 5 "-H), 6.96 (d, J=8.5Hz, 3 " ', 5 " '-H), 6.76 (s, 1H, 5-H), 6.55 (d, J=9.0Hz, 2H, 2 "; 6 "-H), 6.52 (s, 1H, 8-H), 6.34 (s, 2H, 2 ', 6 '-H), 5.95and 5.97 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.69 (m, 1H, 4-H), 4.61 (d, J=5.0Hz, 1H, 1-H), 4.39 (t, J=8.0Hz, 1H, 11-H), 4.02 (t, J=8.0Hz, 1H, 11-H), 3.91 (s, 3H ,-OCH
3), 3.83 (s, 6H, 3 ', 5 '-OCH
3), 3.16 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 3.04 (m, 1H, 3-H) .ESI-MS:m/z[M+H]
+625.
Embodiment 3: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-to the fluorobenzoyl amido) anilino] epipodophyllotoxin (GL2)
Operating process such as embodiment 1 just replace anisic acid with parafluorobenzoic acid.Obtain yellow solid, yield: 57%, fusing point: 157.2-158.9 ℃,
1HNMR (500MHz, CDCl
3) δ 7.86 (m, 2H, 2 " ', 6 " '-H), 7.66 (s, 1H,-CONH-), 7.44 (d, J=8.5Hz, 2H, 3 ", 5 "-H), 7.14 (t, J=8.5Hz, 3 " ', 5 " '-H), 6.76 (s, 1H, 5-H), 6.55 (d, J=9.0Hz, 2H, 2 ", 6 "-H), 6.52 (s, 1H, 8-H), 6.34 (s, 2H, 2 ', 6 '-H), 5.95and 5.97 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.66 (m, 1H, 4-H), 4.59 (d, J=5.5Hz, 1H, 1-H), 4.36 (t, J=8.0Hz, 1H, 11-H), 3.98 (t, J=8.0Hz, 1H, 11-H), 3.85 (br, 1H ,-CONH), 3.79 (s, 6H, 3 ', 5 '-OCH
3), 3.13 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 3.00 (m, 1H, 3-H).
Embodiment 4: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-to the acetoxyl group benzoylamino) anilino] epipodophyllotoxin (GL3)
Operating process such as embodiment 1 just use acetoxy-benzoic acid is replaced anisic acid.Obtain yellow solid, yield: 28.13%, fusing point: 182.2-184.5 ℃,
1H NMR (500MHz, CDCl
3) δ 7.87 (d, J=8.5Hz, 2H, 2 " ', 6 " '-H), 7.71 (s, 1H ,-CONH-), 7.44 (d, J=8.5Hz, 2H, 3 "; 5 "-H), 7.20 (d, J=8.5Hz, 3 " ', 5 " '-H), 6.76 (s, 1H, 5-H), 6.55 (d, J=9.0Hz, 2H, 2 "; 6 "-H), 6.53 (s, 1H, 8-H), 6.34 (s, 2H, 2 ', 6 '-H), 5.95and 5.97 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.65 (m, 1H, 4-H), 4.58 (d, J=4.5Hz, 1H, 1-H), 4.36 (t, J=8.0Hz, 1H, 11-H), 3.98 (t, J=8.0Hz, 1H, 11-H), 3.85 (br, 1H ,-CONH), 3.79 (s, 6H, 3 ', 5 '-OCH
3), 3.12 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 2.98 (m, 1H, 3-H) .2.33 (s, 3H ,-COCH
3) .ESI-MS:m/z[M+H]
+653.
Embodiment 5: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-p-nitrophenyl formamido-) anilino] epipodophyllotoxin (GL4)
Operating process such as embodiment 1 just replace anisic acid with p-nitrobenzoic acid.Obtain orange solids, yield: 43.57%, fusing point: 230.1-231.7 ℃,
1H NMR (500MHz, CDCl
3) δ 8.34 (d, J=8.5Hz, 2H, 2 " ', 6 " '-H), 8.03 (d, J=8.5Hz, 2H, 3 " ', 5 " '-H), 7.70 (s, 1H ,-CONH-), 7.47 (d, J=8.0Hz, 2H, 3 ", 5 "-H), 6.77 (s, 1H, 5-H), 6.58 (d, J=9.0Hz, 2H, 2 "; 6 "-H), 6.54 (s, 1H, 8-H), 6.34 (s, 2H, 2 ', 6 '-H), 5.97and 5.98 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.68 (m, 1H, 4-H), 4.61 (d, J=5.0Hz, 1H, 1-H), 4.37 (t, J=8.0Hz, 1H, 11-H), 3.99 (t, J=8.0Hz, 11-H), 3.87 (br, 1H ,-CONH-), 3.79 (s, 6H, 3 ', 5 '-OCH
3), 3.13 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 3.00 (m, 1H, 3-H).
Embodiment 6: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-hydrocinnamamide base) anilino] epipodophyllotoxin (GL5)
Operating process such as embodiment 1 just replace anisic acid with phenylpropionic acid.Obtain yellow solid, yield: 60.63%, fusing point: 156.8-158.5 ℃,
1H NMR (500MHz, CDCl
3) δ 8.15 (d, J=9.0Hz, 2H, 3 " ', 6 " '-H), 7.41 (d, J=9.0Hz, 2H, 4 " ', 5 " '-H), 6.91 (s, 1H,-CONH-), 6.73 (s, 1H, 5-H), 6.53 (s, 1H, 8-H), 6.48 (d, J=8.5Hz, 2H; 2 ", 6 "-H), 6.33 (s, 2H, 2 '; 6 '-H), 5.95and 5.97 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.62 (m, 1H, 4-H), 4.59 (d, J=5.0Hz, 1H, 1-H), 4.34 (t, J=8.0Hz, 1H, 11-H), 3.95 (t, J=8.0Hz, 11-H), 3.79 (s, 6H, 3 ', 5 '-OCH
3), 3.15 (t, J=7.5Hz, 2H ,-NHCOCH
2CH
2-Ph), 3.11 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 2.99 (m, 1H, 3-H) .2.66 (t, J=7.5Hz, 2H ,-NHCOCH
2CH
2-Ph).
Embodiment 7: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-ortho-nitrophenyl propionamido-) anilino] epipodophyllotoxin (GL6)
Operating process such as embodiment 1 just replace anisic acid with the ortho-nitrophenyl propionic acid.Obtain brown solid, yield: 31.90%, fusing point: 158.3-159.2 ℃,
1H NMR (500MHz, CDCl
3) δ 8.15 (d, J=9.0Hz, 2H, 3 " ', 6 " '-H), 7.41 (d, J=9.0Hz, 2H, 4 " ', 5 " '-H), 6.91 (s, 1H,-CONH-), 6.73 (s, 1H, 5-H), 6.53 (s, 1H, 8-H), 6.48 (d, J=8.5Hz, 2H; 2 ", 6 "-H), 6.33 (s, 2H, 2 '; 6 '-H), 5.95and 5.97 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.62 (m, 1H, 4-H), 4.59 (d, J=5.0Hz, 1H, 1-H), 4.34 (t, J=8.0Hz, 1H, 11-H), 3.95 (t, J=8.0Hz, 11-H), 3.79 (s, 6H, 3 ', 5 '-OCH
3), 3.15 (t, J=7.5Hz, 2H ,-NHCOCH
2CH
2-Ph), 3.11 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 2.99 (m, 1H, 3-H) .2.66 (t, J=7.5Hz, 2H ,-NHCOCH
2CH
2-Ph).
Embodiment 8: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-phenylallene amide group) anilino] epipodophyllotoxin (GL7)
Operating process such as embodiment 1 just replace anisic acid with TRANSCINNAMIC ACID.Obtain yellow solid, yield: 29.84%, fusing point: 153.7-155.2 ℃,
1H NMR (500MHz, CDCl
3) δ 7.73 (m, 1H ,-CH=CH), 7.54 (d, J=5.0Hz, 2H, 2 " '; 6 " '-H), 7.47 (d, J=5.5Hz, 2H, 3 ", 5 "-H), 7.39 (m, 3H, 3 " ', 4 " ', 5 " '-H), 7.15 (s, 1H ,-CH=CH); 6.77 (s, 1H, 5-H), 6.57 (s, 1H, 8-H), 6.54 (m; J=4.0Hz, 2H, 2 ", 6 "-H), 6.35 (s, 2H; 2 ', 6 '-H), 5.97and 5.98 (ABq, J=1.0Hz, 2H ,-OCH
2O-), 4.66 (m, 1H, 4-H), 4.60 (d, J=4.5Hz, 1H, 1-H), 4.37 (t, J=8.0Hz, 1H, 11-H), 4.00 (t, J=8.0Hz, 11-H), 3.80 (s, 6H, 3 ', 5 '-OCH
3), 3.13 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 3.01 (m, 1H, 3-H).
Embodiment 9: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-to the anisole propionamido-) anilino] epipodophyllotoxin (GL8)
Operating process such as embodiment 1 just replace anisic acid with 4-Methoxyhydrocinnamic acid.Obtain pale solid, yield: 71.78%, fusing point: 149.4-151.2 ℃,
1H NMR (500MHz, CDCl
3) δ 7.24 (m, 2H, 2 " ', 6 " '-H), 7.16 (d, J=8.5Hz, 2H, 3 ", 5 "-H), 6.84 (m, 3H, 3 "; 4 ", 5 "-H), 6.74 (s, 1H, 5-H), 6.52 (s; 1H, 8-H), 6.48 (d, J=8.0Hz, 2H; 2 ", 6 "-H), 6.33 (s, 2H, 2 '; 6 '-H), 5.95and 5.97 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.62 (m, 1H, 4-H), 4.59 (d, J=5.0Hz, 1H, 1-H), 4.34 (t, J=8.0Hz, 1H, 11-H), 3.96 (t, J=8.0Hz, 11-H), 3.79 (m, 6H, 3 ', 5 '-OCH
3), 3.11 (dd, J=5.0Hz, 14.0Hz, 1H, 2-H), 3.00 (m, 1H, 3-H), 2.98 (t, J=7.5Hz, 2H ,-NHCO-CH
2CH
2-Ph), 2.59 (t, J=7.5Hz, 2H ,-NHCO-CH
2CH
2-Ph).
Embodiment 10: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-to the brombenzamide base) anilino] epipodophyllotoxin (GL9)
Operating process such as embodiment 1 just replace anisic acid with parabromobenzoic acid.Obtain the purple solid, yield: 46.77%, fusing point: 160.9-162.6 ℃,
1H NMR (500MHz, CDCl
3) δ 8.06 (d, J=8.5Hz, 2H, 2 " ', 6 " '-H), 7.74 (d, J=8.5Hz, 2H, 3 " ', 5 " '-H), 7.69 (s, 1H,-CONH-), 7.62 (m, 2H, 2 ", 6 "-H), 7.45 (d, J=8.5Hz, 2H, 3 ", 5 "-H), 6.78 (s, 1H, 8-H), 6.56 (m, 3H, 5-H, 2 ', 6 '-H), 5.97and 5.99 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.67 (m, 2H, 4-H, 1-H), 4.43 (m, 1H, 11-H), 3.87 (m, 1H, 11-H), 3.70 (s, 6H, 3 ', 5 '-OCH
3), 3.19 (m, 1H, 2-H), 3.02 (m, 1H, 3-H).
Embodiment 11: the preparation of compound 4 '-O-demethyl-4-deoxidation-[4-(4 "-to chloro-benzoyl amino) anilino] epipodophyllotoxin (GL10)
Operating process such as embodiment 1 just replace anisic acid with Chlorodracylic acid.Obtain the purple solid, yield: 14.26%, fusing point: 165.6-167.2 ℃,
1H NMR (500MHz, CDCl
3) δ 8.01 (d, J=9.0Hz, 2H, 2 " ', 6 " '-H), 7.72 (d, J=8.5Hz, 2H, 3 " ', 5 " '-H), 7.65 (s, 1H,-CONH-), 7.52 (m, 2H, 2 ", 6 "-H), 7.41 (d, J=8.5Hz, 2H, 3 ", 5 "-H), 6.75 (m, 1H, 8-H), 6.51 (m, 3H, 5-H, 2 ', 6 '-H), 5.97and 5.99 (AB q, J=1.0Hz, 2H ,-OCH
2O-), 4.61 (m, 2H, 4-H, 1-H), 4.37 (m, 1H, 11-H), 3.80 (m, 1H, 11-H), 3.68 (s, 6H, 3 ', 5 '-OCH
3), 3.12 (m, 1H, 2-H), 3.00 (m, 1H, 3-H).
Embodiment 12:4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative is to the vitro inhibition effect of different tumour cells
1. experiment material
Cell strain: the high transitivity lung cancer cell line of people 95D, human lung carcinoma cell line A549, human oral epidermal carcinoma cell strain KB and persister KB/VCR thereof.
Substratum: HG-DMEM substratum or RPMI1640 substratum contain 10% calf serum or foetal calf serum.
Medicine and preparation: medicine is 4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative, and medicine is dissolved in DMSO, and dilution obtains 5 concentration by a certain percentage.
2. experimental technique
With the above-mentioned tumour cell that is in logarithmic phase, with 2 * 10
4Individual/ml is inoculated in 96 well culture plates, and every hole adds cell suspension 200 μ l, after cultivating 24h, adds the soup 1 μ l of above-mentioned preparation respectively, and each concentration is established 3 multiple holes.Cell is at 37 ℃, 5%CO
2After hatching 72 hours in the incubator, adding concentration is the MTT solution 20 μ l of 2.5mg/ml, continues to cultivate 4 hours.Supernatant liquor is removed in suction, adds 100 μ l DMSO and shakes up, and with the OD value in microplate reader each hole of mensuration under the 570nm wavelength, the calculation formula of cell inhibitory rate is:
Cell inhibitory rate %=(control group OD value-medication group OD value)/control cells OD value * 100% obtains IC with the Bliss method
50
3. experimental result
10 representative compounds of table 1. synthetic are to the IC of different tumor cell lines
50(μ M) value
4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative effect was measured IC respectively after 72 hours
50Value.The result shows: 1) 10 compounds all have certain inhibition activity to the high transitivity lung cancer cell line of people 95D, human lung carcinoma cell line A549, human oral epidermal carcinoma cell strain KB and human oral epidermal carcinoma drug-resistant cell strain KB/VCR; 2) 10 compounds have the cytotoxic activity stronger than Etoposide, its IC to the high transitivity lung cancer cell line of people 95D
50Value is between 0.41～6.56 μ M, and KB/VCR has the cytotoxic activity stronger than Etoposide, its IC to human oral epidermal carcinoma drug-resistant cell strain
50Value shows overriding resistance activity preferably between 2.15～16.43 μ M; 3) compound GL2, GL3 is tried tumor cell line to four kinds all the cytotoxic activity stronger than Etoposide, to the IC of above-mentioned cell
50Value is between 0.42～16.43 μ M.
Embodiment 13:4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative GL3 is to the restraining effect of S180 osteosarcoma mouse
1. experiment material
Laboratory animal: SPF level ICR kind mouse, male, 24, body weight 25 ± 2g.Company limited provides by the Si Laike laboratory animal.Laboratory animal production licence: SCXK (Shanghai) 2007-0008.Laboratory animal occupancy permit number: SCXK (Zhejiang) 2007-0098.
The animal rearing condition: meet the requirement of SPF level animal facility, 20～25 ℃ of temperature, humidity 40～70%, each 12h of illumination light and shade, rate of ventilation is 10～20 times/hour, freely drink water (urban drinking water), illumination replaces 12h/12h for light and shade round the clock.
2. experimental technique
2.1 inoculation S180 cell
With 5 * 10
6Individual S180 cell abdominal injection ICR mouse, mouse ascites was extracted on the 7th day in the inoculation back, and being diluted to cell count with stroke-physiological saline solution is 1 * 10
6The tumor cell suspension of individual/mL, every mouse is in right fore oxter inoculation 0.2mL, and next day, random packet began administration.
2.2 grouping and administration
24 mouse of inoculation S180 knurl strain are divided into 3 groups at random, are respectively negative control group, GL3 100mg/kg, cisplatin 2mg/kg, and wherein the negative control treated animal is 10, and 8 of cisplatin treated animals, GL3 treated animal are 6.Negative control group is irritated stomach and is given solvent 0.1ml/10g, GL3 100mg/kg group mouse gives the soup 0.1ml/10g of corresponding dosage, the cisplatin group is given 0.2mg/ml soup 0.1ml/10g, GL3 is in inoculation back the 1st, drug withdrawal after 2,4 administrations, the 9th day execution mouse, get the knurl piece and weigh, calculate the tumor suppression percentage.
3. experimental result
Compared with the control, body weight slightly descends, and tangible tumor-inhibiting action is arranged after the GL3 group 100mg/kg group administration 3 times.The results are shown in Table 2.Compound GL3 is under heavy dose of administration condition of 100mg/kg, inhibitory rate to 58.1%, tumour inhibiting rate than positive control medicine cis-platinum is high by 13.7%, and this result shows that this compound in the inhibiting while of improving tumour, can reduce its toxicity well.
The influence that table 2.GL3 is heavy to S180 tumor-bearing mice knurl
*Compare with control group p＜0.01
Embodiment 14:4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative GL3 is to the restraining effect of the high transitivity lung cancer of people 95D Nude Mice
1. laboratory animal
Nude mouse: the Balb/c strain, female, 20,4～5 ages in week, body weight 17 ± 1g.Provide credit number by Shanghai Slac Experimental Animal Co., Ltd.: SCXK (Shanghai) 2007-0005.
2. experimental technique
2.1 inoculated tumour cell
The 95D lung carcinoma cell with contain 10% foetal calf serum 1640 in 37 ℃, 5%CO
2The conventional cultivation in the incubator, after going down to posterity, when treating that cell reaches aequum, the digestion collecting cell.With 1 * 10
7The high transitivity lung cancer cell line of individual 95D people is injected into oxter, nude mouse left side, treats that tumor growth is to 100-200mm
3After, the animal random packet is begun administration.
2.2 grouping and administering mode
Inoculate in advance the 95D nude mouse from 20 and to select 10 consistent nude mouses of tumor growth to be divided into 2 groups at random.1) Control group, 5; 2) GL350mg/kg group, 5.The GL350mg/kg group is irritated stomach every other day and is given 5mg/ml GL3 solution 0.1ml/10g, successive administration 20 days.On every Mondays, three, five weigh and measure gross tumor volume, put to death nude mouse in the 20th day, weigh, it is long-pending to measure the knurl block, calculates relative tumour volume (RTV), tumour appreciation rate (T/C) and tumor suppression percentage relatively, does the statistics detection.
3. experimental result
The administration of GL3 group is after 5 days, and body weight change is obvious, and at the 7th day animal dead is arranged, after this whether administration by the state decision of animal, in whole process, only the 1st, 3, administration in 10,12,14,17 and 19 days.Behind the animals administer 20 days, it is 9.3 ± 1.5 that Control organizes relative knurl volume (RTV), and GL350mg/kg group RTV value is 4.3 ± 0.6, and corresponding tumour appreciation rate (T/C) relatively is 46.3% (seeing Table 3).Administration is cutd open after 20 days and is got the knurl piece extremely and weigh, and GL3 50mg/kg group Nude Mice weight in average is 0.84 ± 0.10g, and its tumour inhibiting rate (%) is 57.3.GL3 50mg/kg group mice-transplanted tumor weight in average has been compared obvious decline with control group, and significant difference is arranged, and sees Table 4.This result shows that this compound has the obvious suppression effect to the high transitivity lung cancer of people 95D Nude Mice.
Table 3.GL3 is to the experimental therapy effect of people's lung cancer 95D Nude Mice
D1: administration first day.*: P＜0.05, * * *: P＜0.001, vs solvent control group on the same day, student-t check.
Table 4.GL3 is to the influence of people's lung cancer 95D Nude Mice tumor weight
*: P＜0.05, vs solvent control group on the same day, student-t check.
Claims (7)
1. 4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative is characterized in that having following general structure (I):
Wherein:
X is-(CH
2)
n-, n be 0～2 or-CH=CH-;
R
1, R
2, R
3, R
4, R
5Identical or different, R
1, R
2, R
3, R
4, R
5For hydrogen atom, halogen atom, hydroxyl, nitro, contain 1 to 6 carbon atom straight or branched alkyl or contain the alkoxyl group of the straight or branched of 1 to 3 carbon atom.
2. 4 '-O-demethyl according to claim 1-4-deoxidation-4-substituted anilino-podophyllotoxine derivative is characterized in that described X is-(CH
2)
n-, n is 0, R
1=R
2=R
4=R
5Be hydrogen, R
3Be fluorine, chlorine, bromine, methoxyl group or nitro, cyano group or acetoxyl group.
3. 4 '-O-demethyl according to claim 1-4-deoxidation-4-substituted anilino-podophyllotoxine derivative is characterized in that described X is-(CH
2)
n-, n is 0, R
2=R
3=R
4=R
5Be hydrogen, R
1Be fluorine, chlorine, bromine, methoxyl group, nitro, cyano group or acetoxyl group.
4. 4 '-O-demethyl according to claim 1-4-deoxidation-4-substituted anilino-podophyllotoxine derivative is characterized in that described X is-(CH
2)
n-, n is 1, R
1=R
2=R
4=R
5Be hydrogen, R
3Be fluorine, chlorine, bromine, methoxyl group, nitro, cyano group or acetoxyl group.
5. 4 '-O-demethyl according to claim 1-4-deoxidation-4-substituted anilino-podophyllotoxine derivative is characterized in that described X is-(CH
2)
n-, n is 2, R
2=R
3=R
4=R
5Be hydrogen, R
1Be fluorine, chlorine, bromine, methoxyl group or nitro, cyano group or acetoxyl group.
6. 4 '-O-demethyl according to claim 1-4-deoxidation-4-substituted anilino-podophyllotoxine derivative is characterized in that described X is-CH=CH-R
1=R
2=R
3=R
4=R
5Be hydrogen.
7. according to the application of arbitrary described 4 '-O-demethyl-4-deoxidation-4-substituted anilino-podophyllotoxine derivative in the claim 1～6 in the preparation antitumor drug.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102532152A (en) * | 2010-12-27 | 2012-07-04 | 中国医学科学院药物研究所 | 4'-demethylepipodophyllotoxin compounds and use thereof as anticancer agent |
| CN102603761A (en) * | 2012-02-02 | 2012-07-25 | 华东师范大学 | Etoposide compound with hydroxamic acid structure and preparation method and usage thereof |
| CN104945409A (en) * | 2015-05-15 | 2015-09-30 | 东华大学 | Prodrug of compound podophyllotoxin PPT with anti-tumour activity and preparation method thereof |
| CN115197099A (en) * | 2022-05-30 | 2022-10-18 | 安徽昊帆生物有限公司 | Preparation method of N-Boc-1,4-phenylenediamine |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101074233A (en) * | 2007-06-22 | 2007-11-21 | 浙江大学 | 4'-demethylpodoph-yllotoxin derivative, its production and use |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101074233A (en) * | 2007-06-22 | 2007-11-21 | 浙江大学 | 4'-demethylpodoph-yllotoxin derivative, its production and use |
Non-Patent Citations (1)
| Title |
|---|
| 《药学学报》 20001231 陈再新等 4- -酯取代-4-脱氧-4-去甲表鬼臼毒素衍生物的合成及其抗肿瘤活性 353-357 1-7 第35卷, 第5期 2 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102532152A (en) * | 2010-12-27 | 2012-07-04 | 中国医学科学院药物研究所 | 4'-demethylepipodophyllotoxin compounds and use thereof as anticancer agent |
| CN102603761A (en) * | 2012-02-02 | 2012-07-25 | 华东师范大学 | Etoposide compound with hydroxamic acid structure and preparation method and usage thereof |
| CN102603761B (en) * | 2012-02-02 | 2013-12-25 | 华东师范大学 | Etoposide compound with hydroxamic acid structure and preparation method and usage thereof |
| CN104945409A (en) * | 2015-05-15 | 2015-09-30 | 东华大学 | Prodrug of compound podophyllotoxin PPT with anti-tumour activity and preparation method thereof |
| CN115197099A (en) * | 2022-05-30 | 2022-10-18 | 安徽昊帆生物有限公司 | Preparation method of N-Boc-1,4-phenylenediamine |
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