CN104193790A - Formyl adenine compound - Google Patents

Abstract

The invention relates to a formyl adenine compound, and in particular relates to a novel compound (which is represented by formula I) and salt thereof. An experiment shows that growth of tumor cells can be suppressed, and the compound is used for preparing a tumor resistant medicament; the compound can be reacted with inorganic acids, such as phosphoric acid, nitric acid and sulfuric acid, and organic acids, such as formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, glycine, valine, maleic acid, fumaric acid and succinic acid, to generate salt. The compound has a better effect of suppressing the growth of the tumor cells.

Description

A kind of formyl adenine compound

Technical field

The present invention relates to a kind of adenine compound with inhibition tumor cell growth.

Background technology

Tumour is the serious disease of harm humans health, and the preventing and controlling of tumour are the emphasis in medical research field always.At present, due to the problems such as environmental pollution of bringing in industrial development, the mankind's living environment quality constantly declines, and causes the sickness rate of tumor disease and lethality rate constantly to rise.Radiotherapy, chemotherapy are to live at present in treating the Main Means of tumour.But chemotherapy, radiotherapy, having suppressed when cancer cells is grown also to have suppressed Normocellular growth, have reduced immunity of organisms, cause new complication.The specifics for the treatment of tumor disease can not be satisfactory, and clinical cytotoxic drug selectivity used is not high at present, causes, to Normocellular pernicious killing and wounding, having limited its application.

The present invention relates to a kind of new adenine compound, through external, anti-tumor in vivo, experimental results show that to there is good activity.

Clofarex (clofarabine) is a kind of novel ucleosides anti-cancer agent, be applied to now treating leukemia of children and lymphoma, because bioavailability is lower, the form of clinical employing drug administration by injection, we carry out a series of structural modification for Clofarex, final discoverable type (I) compound has good bioavailability, by bioavailability test-results, show that the bioavailability of formula (I) compound is Clofarex 8 times, absolute bioavailability reaches 48.1%, can utilize the form of therapy tumour of oral administration.

disclosure of the invention

The invention provides a kind of antineoplastic compound, is a kind of new adenine derivative and ester and salt, is specially formula (I) compound or its pharmacy acceptable salt:

The mineral acids such as above-claimed cpd and sulfuric acid, phosphoric acid, sulfonic acid, hydrochloric acid, with glycine, L-Ala, Methionin, arginine, Serine, phenylalanine, proline(Pro), tyrosine, aspartic acid, L-glutamic acid, Histidine, leucine, methionine(Met), Threonine, Pyrrolidonecarboxylic acid, tryptophane or α-amino-isovaleric acid, dextrocamphoric acid, oxysuccinic acid, citric acid, toxilic acid, succsinic acid, oxalic acid, pentanedioic acid, oxalic acid, lactic acid or propanedioic acid; The organic acid reactions such as pamoic acid, hydroxynaphthoic acid, gentisinic acid, Whitfield's ointment, oxyacetic acid, amygdalic acid, lactic acid, 4-acetaminobenzoic acid or nicotinic acid obtain salt.

This compound can be obtained by reaction scheme below.

Specific embodiment:

Embodiment 1: the preparation of compound:

1, compound 4 is synthetic:

Accurately take 94.5 grams of raw materials 2 and 243 grams of raw materials 3 join in 500.0mL ethylene dichloride, at 100 ℃, stirring reaction is 16 hours, stopped reaction, reaction solution is cooled to room temperature, adds the ammoniacal liquor 300mL of 2.0N, stir 20 minutes, stratification, separates organic layer, with distilled water wash (200mL * 3), after underpressure distillation, obtain 222.9 and digest compound 4, productive rate is 70.1%.HNMR(400Hz，DMSO)：8.68(s，1H)，6.22(d，J＝3.2Hz，1H)，4.05-4.01(d，J＝3.6Hz，2H)，3.92-3.90(m，1H)，3.85-3.82(m，1H)，3.78-3.81(m，1H)，1.81(m，6H)，1.01(d，J＝4.0Hz，36H)；MS(m/z)：636.5。

2, compound 5 is synthetic:

Under room temperature, by 95.4, digest compound 4 and join in the round-bottomed flask of 1000mL, add Virahol and the 45.0mL strong aqua (concentration is about 25%) of 200.0mL, load onto prolong and nitrogen ball, at 80 ℃, react and spend the night.Stopped reaction, reaction solution is cooled to room temperature, and underpressure distillation is digested compound 5 except obtaining 85.3 after desolventizing, and productive rate is 92.3%.HNMR(400Hz，DMSO)：8.68(s，1H)，6.22(d，J＝3.2Hz，1H)，4.05-4.01(d，J＝3.6Hz，2H)，3.92-3.90(m，1H)，3.85-3.82(m，1H)，3.78-3.81(m，1H)，1.81(m，6H)，1.01(d，J＝4.0Hz，36H)；MS(m/z)：617.7。

3, compound 6 is synthetic:

Under room temperature, accurately take 61.6 and digest compound 5 and 15.0 grams of triethylamines join in the methylene dichloride of 200.0mL, stirring and dissolving is complete.18 grams of chloroformyl n-pentyl esters are dissolved with 50.0mL methylene dichloride, be then slowly added drop-wise in above-mentioned reaction solution, dropwise rear continuation reaction 5 hours.Stopped reaction, adds 100.0mL distilled water to reaction solution, stirs 10 minutes, separates organic layer, and the acetic acid solution washing (50.0mL * 3) with 1.0N, separates organic layer, and underpressure distillation is digested compound 6 except obtaining 53.5 after desolventizing, and productive rate is 73.3%.HNMR(400Hz，DMSO)：8.68(s，1H)，8.01(s，1H)，6.22(d，J＝3.2Hz，1H)，4.09-4.07(t，J＝3.6Hz，2H)，4.05-4.01(d，J＝3.6Hz，2H)，3.92-3.90(m，1H)，3.85-3.82(m，1H)，3.78-3.81(m，1H)，1.81(m，6H)，1.58-1.56(m，2H)，1.33-1.29(m，4H)，1.05(t，J＝4.4Hz，3H)，1.01(d，J＝4.0Hz，36H)；MS(m/z)：731.6。

4, compound 1 is synthetic:

Digest 6,7.0 grams of Methanaminium, N,N,N-trimethyl-, fluorides of compound and 4.5 grams of Glacial acetic acid join in the DMF of 100.0mL by 36.5, under room temperature, stirring reaction spends the night.Stopped reaction, to reaction solution, add 500.0mL distilled water, stir 10 minutes, then be extracted with ethyl acetate (100.0mL * 3), merge organic phase, distilled water wash for organic phase (50.0mL * 3), separate organic layer, anhydrous sodium sulfate drying, underpressure distillation takes the crude product that obtains compound 1 after solvent, to crude product, add 50.0mL ethyl acetate, heating for dissolving, then slowly add sherwood oil until there be muddy appearance in solution, at this moment, stop heating, naturally cool to room temperature, then move in ice-water bath cooling 1 hour, suction filtration, filter cake rinses (10.0mL * 3) with ether, after vacuum-drying, obtain 11.6 grams of pure compounds 1, productive rate is 55.8%.HNMR(400Hz，DMSO)：8.68(s，1H)，8.01(s，1H)，6.22(d.J＝3.2Hz，1H)，4.09-4.07(t，J＝3.6Hz，2H)，3.92-3.90(m，1H)，3.85-3.82(m，1H)，3.78-3.81(m，1H)，3.75-4.72(m，2H)1.58-1.56(m，2H)，1.33-1.29(m，4H)，1.05(t，J＝4.4Hz，3H)；MS(m/z)：418.2。

Embodiment 2: the mensuration of compound 1 bioavailability in embodiment 1

1 material

1.1 medicines and reagent C LM (in embodiment 1, compound 1), measure its massfraction >=97.0% (by dry product) through HPLC method.Methyl alcohol is chromatographically pure, J.T Baker company; Commercially available Wahaha Pure Water, CL (Clofarex).

1.2 instrument Shimadzu high performance liquid chromatographs, Japanese Shimadzu company; XW-80A type eddy mixer, Industrial Co., Ltd. of upper Nereid section; PROINO whizzer, U.S. Kendro Lab Prod GmbH; PK514BP ultrasonic cleaning machine, German Bandel company; AX2 plum Teller-Tuo benefit AX205Delta Range electronic balance, Switzerland Mei Tele company.

1.3 animal male Wistar rats, weight 300～400g.

2 methods and result

2.1 administration and sample collecting

Water is can't help in the last late fasting of rat experiment.CLM is dissolved in physiological saline and is mixed with suspension, ig administration, dosage 400mg/kg.Separately CLM is dissolved in containing in the physiological saline of 2% dimethyl sulfoxide (DMSO) (DMSO) to iv administration, dosage 160mg/kg (2.5mL/kg).Before administration and after administration 5,10,20,30,60,90,120,240,360,480min jugular sinus gets blood 0.5mL, anticoagulant heparin, the centrifugal 5min of 10800r/min, gets supernatant liquor, cryopreservation is standby.

Suspension and the DMSO normal saline solution of preparation CL same as above.

2.2 reference substance solution preparations

CLM major part after the oral administration of rat becomes CL, with the form of CL, exists, and therefore selects CL product in contrast.Precision takes CL reference substance 10.00mg and puts in 10mL measuring bottle, is mixed with the CL storing solution of 1.00g/L with methyl alcohol, and 4 ℃ of refrigerators keep in Dark Place standby.With front with methyl alcohol be diluted to 0.1,0.25,0.5,1.0,2.5,5.0,10.0,25.0mg/L series reference substance solution is standby.

2.3 plasma samples are processed

The accurate plasma sample 100 μ L that draw, put in 1.5mL centrifuge tube, add 1mol/L hydrochloric acid soln 15 μ L, vortex vibration 0.5min, add vinyl acetic monomer-normal hexane (3: 1) extraction liquid 900 μ L and Virahol 10 μ L, vortex vibration 2min, 5000r/min low-temperature centrifugation 5min, get supernatant liquor, 37.5 ℃ of water-baths, nitrogen dries up, and residue redissolves by 100 μ L moving phases, vortex 2min, sample introduction 10 μ L.Plasma sample need be processed on hypothermia operation table.

2.4 chromatographic condition methods: adopt Shim-packVP-ODS (5 μ m, 4.6mm * 150mm) chromatographic column, moving phase is that acetonitrile ammonium acetate solution (is got ammonium acetate 0.31g, be dissolved in water and be diluted to 1000mL, add again formic acid 1.0mL) (10: 90), flow velocity 1.0mL/min, sample size 20 μ L, detect wavelength 262nm.(Clofarex is good in sample size 0.4～242.5 μ g/mL scope internal linear relation, and regression equation is A=23147C+12468 (r=0.9999, n=5), and average average recovery is that 99.83%, RSD is 0.42%.)

2.5 sample determinations under above-mentioned chromatographic condition, the about 10.5min of the retention time of CL, in blood plasma, endogenous impurity is substantially noiseless to the mensuration of CL.Draw respectively " 2.2 " lower serial reference substance solution 0.5mL, add blank plasma 0.5mL, by " 2.3 " lower method operation, by " 2.4 ", measure, record respectively peak area, take CL peak area as ordinate zou (Y), AB concentration is that X-coordinate (X) carries out linear regression, the regression equation that obtains typical curve is Y=25.89X-0.524, r2=0.9997, and B is good in 5～50 μ g/mL linear relationships.The RSD of the precision test of the method, the stability test in 3h, circulation ratio test is respectively 0.63%, 0.78%, 1.19%; The rate of recovery is that 98.33%, RSD is 1.22%, and result is good, up to specification.

2.6 data analyses are the Plasma Concentration in rat body-time data DAS2.0 pharmacokinetics routine processes by gained CL, calculates its pharmacokinetic parameters (with sx ± expression) and absolute bioavailability in rat body.

The pharmacokinetic studies rat ig of 2.7CLM and CL different way of administration and iv give after CLM, CL, Plasma Concentration-time curve of CL is analyzed through DAS2.0 software, calculate the pharmacokinetic parameters (table 1) of CL in rat body after CLM and CL ig and iv administration, and calculate absolute bioavailability (Fab) according to formula.

Fab=(AUC ig administration * iv dosage)/(AUC iv administration * ig dosage)

After CLM ig administration, the absolute bioavailability of CL in rat body is that after 47.98%, CL ig administration, the absolute bioavailability of CL in rat body is 5.89% as calculated.In embodiment 1, the bioavailability of compound 1 (CLM) is much larger than Clofarex (CL)

Pharmacokinetic parameters in rat body after table 1 administration

Parameter	Unit	IV administration (CLM)	Ig administration (CLM)	IV administration (CL)	Ig administration (CL)
						T max	h	0.08±0.17	1.56±0.33	0.11±0.05	1.12±0.33
T 1/2	h	5.83±0.44	5.5±0.21	5.33±0.20	5.45±0.32
						AUC	mg*h/L	36.67±5.31	44.76±4.13	35.67±3.31	5.91±1.13

Note: CLM exists with CL prodrug forms, for ease of comparing, and the T of CLM _1/2to be converted into the T after CL _1/2statement

Compound 1 anti-tumor activity test in embodiment 3 embodiment 1

Experiment material:

Medicine and reagent: CLM (compound 1 in embodiment 1), white powder

Cyclophosphamide for injection, Hengrui Medicine Co., Ltd., Jiangsu Prov.'s product.

Animal and knurl strain: SPF level Kunming kind small white mouse, body weight 18～22g, mouse H22 liver cancer

Instrument: CO ₂incubator (Forma3110, USA), Bechtop (BCN-1360, east, Harbin connection), inverted microscope (Nikon).

H22 liver cancer is got the ascites preservation of going down to posterity after Kunming mouse intraperitoneal inoculation.Get the H22 liver cancer tumor-bearing mice that ascites goes down to posterity the 10th, de-cervical vertebra is put to death mouse, and sterilization skin of abdomen, draws oyster white ascites with asepsis injector, and it is 1 * 107 cell/ml that the injection physiological saline of take is adjusted tumour cell concentration.With cotton ball soaked in alcohol sterilization Kunming mouse right side armpit skin, in the above-mentioned tumor cell suspension 0.2ml of subcutaneous vaccination, conventional raising.

Grouping and administration: 50 of tumor-bearing mices, by body weight, be divided at random 5 groups, 10 every group, be respectively model group, endoxan group, CLM 10,30,90mg/kg group.Each organizes mouse by dosage shown in table 2 and mode administration, and endoxan group only gives endoxan one time in abdominal cavity in lotus knurl second day, and CLM organizes equal every day of gastric infusion 1 time, continuous 10 days.Administration volume 20ml/kg body weight.After last administration 24 hours, de-cervical vertebra was put to death mouse, weighs, and strips tumor tissue and weighs, and calculates tumour inhibiting rate.

Result with represent significant difference comparison between organizing with t check.

Result shows, the continuous gastric infusion of CLM of 30mg/kg 10 days is inhibited to the growth of mice transplanted tumor H22 liver cancer.

The restraining effect of table 2CLM to mouse H22 liver cancer growth

With model group comparison: ^*, P < 0.05; ^*, P < 0.01.

Claims (2)

1. suc as formula the compound of structure shown in (I):

2. the purposes of compound in preparing medicine for treating tumor thing described in claim 1.