CN107303391A - The low-density lipoprotein composition of leukotrienes and the prodrug of cancer therapy drug and the prodrug - Google Patents
The low-density lipoprotein composition of leukotrienes and the prodrug of cancer therapy drug and the prodrug Download PDFInfo
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Abstract
The present invention relates to the prodrug of a kind of leukotrienes and cancer therapy drug, the preparation method of the prodrug, the prodrug low-density lipoprotein composition, and the application of the prodrug and low-density lipoprotein composition in oncotherapy.Characterized in that, the hydroxyl of linolenic carboxyl and cancer therapy drug, amino groups are covalently attached the prodrug for obtaining leukotrienes and cancer therapy drug under certain catalytic condition.The prodrug is contained to the low-density lipoprotein composition that prodrug is prepared into artificial synthesized low-density lipoprotein.The low-density lipoprotein composition of leukotrienes and the prodrug of cancer therapy drug and the prodrug can be used for the treatment of kinds of tumors, have more preferable tumor inhibitory effect compared with cancer therapy drug, and toxic side effect is substantially reduced.
Description
Technical field
Combined the present invention relates to leukotrienes with the low-density lipoprotein of the prodrug, preceding medicament preparation, prodrug of cancer therapy drug
Thing, preparation method of composition and its application in oncotherapy, belong to preparation and the application field of cancer therapy drug.
Background technology
Cancer (Cancer) is to influence one of major chronic disease of Chinese residents health, it has also become the first of Chinese residents is dead
Cause.Also, with the aggravation of the factors such as aging population, the morbidity of tumour and the death rate are still in rising trend.According to the world
The prediction of cancer research administration, if do not adopted an effective measure, Cancer in China morbidity number and death count to the year two thousand twenty and will rise to 4,000,000 people
With 3,000,000 people, the year two thousand thirty will rise to 5,000,000 people and 3,500,000 people.
Chemotherapy is the most common way of oncotherapy, but because chemotherapeutics is selective without targeting to tumor tissues,
Obvious whole body toxic side effect occurs.Therefore the therapeutic effect of chemotherapeutics is improved, mitigates the toxic side effect caused by chemotherapy
It is urgent problem to be solved in current tumor research.
Leukotrienes is a kind of n-3 classes polyunsaturated fatty acid, and the metabolism of endotrophic material has substantial connection, can subtract
The risk that few cardiovascular and cerebrovascular occurs, and with certain antitumor activity.
Low-density lipoprotein is the main carriers of internal cholesterol transport, and tumour cell is substantial amounts of due to quick increment needs
Cholesterol builds cell membrane, therefore the height expression of most tumors cell surface LDL receptor, the low density lipoprotein of medicine
Protein composition can realize the targeted therapy to tumour.
Therefore, leukotrienes and cancer therapy drug are covalently attached to form covalent prodrug, and pass through certain preparation preparation means
Contain and composition is formed in low-density lipoprotein, be conducive to improving cancer drug therapy effect, mitigate toxic side effect, reduce cancer
The treatment cost and risk of disease.
The content of the invention
The technical problems to be solved by the invention are to provide prodrug of a kind of leukotrienes and cancer therapy drug and preparation method thereof,
The prodrug is reacted by the carboxyl and the hydroxyl of cancer therapy drug, amino groups on leukotrienes under certain catalytic condition
Arrive, and the prodrug can reduce the toxic side effect of cancer therapy drug.
Another technical problem to be solved by this invention be to provide a kind of prodrug low-density lipoprotein composition and its
Preparation method, said composition can improve oncotherapy effect with targeting in tumor tissues, mitigate the secondary work of poison of chemotherapeutics
With.
In order to solve the above technical problems, the technical solution used in the present invention is：
The present invention discloses the prodrug of leukotrienes and cancer therapy drug a kind of first, the prodrug by the carboxyl on leukotrienes with
The hydroxyl of cancer therapy drug, amino groups are reacted under certain catalytic condition to be obtained.Leukotrienes includes alpha-linolenic acid or γ-Asia
Numb acid；Cancer therapy drug includes：Taxol, Docetaxel, adriamycin, Doxorubicin, cis-platinum, camptothecine, 10- hydroxy-camptothecins
Alkali, 9- HCPTs, SN38 SN-38, TPT, Irinotecan, vincaleukoblastinum, vincristine,
Eldisine, vinflunine, vinpocetine, gemcitabine, polypeptide series antineoplastic medicament (such as Goserelin, Triptorelin), ammonia
Base quinazoline ditosylate salt antineoplastic (such as Gefitinib, Tarceva, Lapatinib) or TNF, preferably Japanese yew
Alcohol and Docetaxel.
The step of present invention further discloses the preparation method of the prodrug of leukotrienes and cancer therapy drug, this method is as follows：
(1) under nitrogen protection, antineoplastic is dissolved in solvent, adds catalyst and dehydrating agent, under stirring, then
Leukotrienes is added, reaction is stirred at room temperature；
(2) above-mentioned reaction resulting material is taken, ether is added, crosses and filters out precipitation, filtrate is successively with 5% hydrochloric acid, water and chlorination
Saturated aqueous solution of sodium is respectively washed three times, collects organic phase, and drying is rotated using Rotary Evaporators, isolated by silica gel chromatographic column
Prodrug.
Wherein, in the above method solvent refer in dichloromethane, chloroform, DMF or dioxane one
Plant or its mixed solution；Catalyst is dimethylamino naphthyridine；Dehydrating agent refers to dicyclohexylcarbodiimide, 1- ethyl -3- (3-
Dimethyl aminopropyl) carbodiimide or one kind or its mixture in DIC, the reaction time is 2-24 hours.
Antineoplastic：Linolenic mol ratio is 1：0.1-10；Catalyst is 1: 0.1-10, preferably 1 with linolenic mol ratio:1；
Dehydrating agent is 1: 0.1-10, preferably 1 with leukotrienes mol ratio:2.
It is preferred that under nitrogen protection, taxol is dissolved in a certain amount of dichloromethane, catalyst dimethylamino is added
Pyridine and dehydrating agent dicyclohexylcarbodiimide, under stirring, add alpha-linolenic acid, and reaction is stirred at room temperature.Take above-mentioned reaction
Resulting material, adds ether, is filtered to remove precipitation, filtrate respectively washes three with the aqueous solution of 5% hydrochloric acid, water and sodium chloride saturation successively
Time, organic phase is collected, drying is rotated using Rotary Evaporators, passes through the isolated prodrug of silica gel chromatographic column.
The present invention further discloses the low-density lipoprotein composition of prodrug, said composition can by emulsify ultrasonic method or
Any one preparation in high-pressure stripping.Wherein emulsification ultrasonic method step is as follows：
(1) appropriate low-density lipoprotein lipotropic component and pro-drug are weighed, a small amount of organic solvent hydrotropy is added, is heated to
50-80 DEG C of melting, it is standby as oil phase.
(2) appropriate low-density lipoprotein functional polypeptide and phosphatide and the amphipathic nature material of auxiliary are weighed, appropriate injection is added
With physiological saline, 50-80 DEG C is heated to, it is standby as aqueous phase.
(3) under 400-800r/min magnetic agitation, aqueous phase is added drop-wise to organic solvent while hot and waved in most oil phase, dripped
Add after finishing, stir 5-20min, be prepared into O/W type colostrums.
(4) by the colostrum prepared rapidly use probe type ultrasonic cell pulverization instrument ultrasonic disperse, power 80-300w, when
Between 3-10min.
(5) ice-water bath cooling 5-30min solidifies preparation, crosses 0.22 μm of miillpore filter and obtains required composition.
Wherein high-pressure stripping step is as follows：
(1) appropriate low-density lipoprotein lipotropic component and pro-drug are weighed, a small amount of organic solvent hydrotropy is added, is heated to
50-80 DEG C of melting, it is standby as oil phase.
(2) appropriate low-density lipoprotein functional polypeptide and phosphatide and the amphipathic nature material of auxiliary are weighed, appropriate injection is added
With physiological saline, 50-80 DEG C is heated to, it is standby as aqueous phase.
(3) under 400-800r/min magnetic agitation, aqueous phase is added drop-wise to organic solvent while hot and waved in most oil phase, dripped
Add after finishing, stir 5-20min, be prepared into O/W type colostrums.
(4) colostrum prepared is disperseed using high pressure homogenizer.
(5) ice-water bath cooling 5-30min solidifies preparation, crosses 0.22 μm of miillpore filter and obtains required composition.
Amphipathic nature material is selected from prepared by above-mentioned preparation：Natural phospholipid, semi-synthetic phosphatide, synthetic phospholipid, the poly- second of phosphatide
Glycol conjugate, Solutol HS 15, PLURONICS F87, polyethylene glycol 1000 vitamin E succinic acid ester or it
Mixture, preferably natural phospholipid and Solutol HS 15 mixture；Organic solvent is selected from：It is ethanol, sweet
One or more in oil, propane diols, polyethylene glycol, DMA, benzyl benzyl ester, ethyl oleate, phenmethylol
Mixture, preferred alcohol.
Wherein each component percentage by weight is as follows：Drug weight percentage is 1%-15%, preferably 3%-10%；Low-density
Lipoprotein functional polypeptide percentage by weight is 5%-25%, preferably 8%-20%；Phosphatide percentage by weight is 8%-24%, preferably
18%-22%；The percentage by weight of the composition such as cholesterol component and triglycerides is 30%-75%, preferably 50%-68%.
The low-density lipoprotein composition particle diameter of prodrug prepared by the above method is in 10-100nm, favorable dispersibility, can be with
Direct injection, or it is lyophilized after lyophilized formulations are made, when using can with 5% glucose solution, physiological saline, water for injection or
Their mixture is disperseed, and reverts to drug administration by injection after the low-density lipoprotein composition of prodrug, or as needed preceding
Pharmaceutically acceptable auxiliary material, such as coemulsifier, stabilizer, pH value regulation are added in the low-density lipoprotein composition of medicine
Agent and antioxidant etc., can be prepared into capsule, oral liquid or external preparation etc. by the conventional method of pharmacy.
The low-density lipoprotein composition of aforementioned prodrugs and prodrug can apply in oncotherapy, and wherein tumour is selected from breast
Gland cancer, liver cancer, oophoroma, lung cancer, the preferably one or more of head and neck cancer or glioma, glioma and liver cancer.
In testing in vivo, glioma nude mice oxter Transplanted tumor model, the different preparation by giving have been initially set up
After find, compared with taxol listing preparation PTX, the prodrug of leukotrienes and cancer therapy drug has suitable antitumous effect, but again
Want, toxic side effect is significantly reduced in the prodrug body of leukotrienes and cancer therapy drug；Compared with PTX, the low-density lipoprotein of prodrug
White composition has stronger tumor inhibitory effect, and toxic side effect is substantially reduced in vivo.Wherein taxol, leukotrienes and anticancer
The prodrug of medicine, the low-density lipoprotein composition tumor inhibitory effect of prodrug respectively reach 51.2%, 50.0% and 72.1%.
The low-density compositions of leukotrienes of the present invention and cancer therapy drug prodrug and prodrug have advantages below：
(1) prodrug of leukotrienes and cancer therapy drug can reduce the toxic side effect of medicine in animal body, improve anticarcinogen
Envelop rate of the thing in nanometer formulation.
(2) there is more preferable tumor inhibitory effect in the low-density compositions body of prodrug and prodrug, and toxic side effect is notable
Reduction.
(3) the various Biocompatibilities of low-density lipoprotein are preferable, cost is low, and preparation technology is simple, is easy to industry
Metaplasia is produced.
Brief description of the drawings：
Fig. 1 is the composite diagram of alpha-linolenic acid and prodrugs of paclitaxel；
Fig. 2 is the mass spectrogram of alpha-linolenic acid and prodrugs of paclitaxel；
Fig. 3 is the hydrogen nuclear magnetic resonance spectrogram of alpha-linolenic acid and prodrugs of paclitaxel；
The grain size distribution that Fig. 4 is measured for the low-density lipoprotein composition of prodrug with dynamic light scattering method；
Fig. 5 is the transmission electron microscope picture of the low-density lipoprotein composition of prodrug；
Fig. 6 is taxol and alpha-linolenic acid and tumor growth curve figure of the prodrugs of paclitaxel to lotus U87 glioma nude mices；
Fig. 7 is taxol and alpha-linolenic acid and prodrugs of paclitaxel to lotus U87 glioma nude mice body weight change curves.
Fig. 8 is bent to the tumour growth of lotus U87 glioma nude mices for the low-density lipoprotein composition of taxol and prodrug
Line chart；
Fig. 9 is naked to lotus U87 gliomas for the low-density lipoprotein composition of alpha-linolenic acid and prodrugs of paclitaxel and prodrug
The tumor growth curve figure of mouse；
Figure 10 swells for the low-density lipoprotein composition of prodrug emulsion composition and prodrug to lotus U87 glioma nude mices
Knurl growth curve chart；
Figure 11 is the low-density lipoprotein composition of taxol and prodrug to lotus U87 glioma nude mice body weight change curves
Figure.
Figure 12 is Ex vivo Tumor picture of the different preparations to lotus U87 glioma nude mices.
Figure 13 is that tumor biopsy TUNEL apoptosis detects figure.
Note：Control is saline control in accompanying drawing,Preparation PTX is listed for taxol, PALA is α-Asia
The prodrug of numb acid and taxol, PALA-ME is the emulsion composition of prodrug, and PALA-sLDL is the low-density lipoprotein group of prodrug
Compound.
Embodiment
The invention will now be further described with reference to specific embodiments, advantages of the present invention and feature will be with description and
It is apparent.It should be understood that the embodiment is only exemplary, any limitation is not constituted to the scope of the present invention.This area
Technical staff should be understood that without departing from the spirit and scope of the invention can to the details of technical solution of the present invention and
Form is modified or replaced, but these modifications or substitutions each fall within protection scope of the present invention.
The synthesis of the alpha-linolenic acid of embodiment 1 and prodrugs of paclitaxel
Taxol 95mg is added in inert environments in the round-bottomed flask of 2.5ml dichloromethane, adds diformazan ammonia
Stirring reaction two hours under yl pyridines 13.6mg, dicyclohexylcarbodiimide 46mg, alpha-linolenic acid 31mg, normal temperature；Use ether
Dilution, 5% hydrochloric acid, water, saturated sodium-chloride water solution washing organic phase；Organic phase is dried with ammonium sulfate, concentrated by rotary evaporation, crosses silicon
Glue chromatography obtains product；Product is characterized with mass spectrum and proton nmr spectra respectively.
Wherein Fig. 1 is synthetic route chart.1136 and 1152 be respectively [PALA+Na in mass spectrum (Fig. 2)+] and [PALA+K+]
Molecular ion peak；It is respectively raw material taxol (PTX), alpha-linolenic acid (ALA) and product α-flax in proton nmr spectra (Fig. 3)
Acid and prodrugs of paclitaxel (PALA) hydrogen nuclear magnetic resonance spectrogram；Compared with taxol, alpha-linolenic acid and prodrugs of paclitaxel chemical shift
The corresponding characteristic peak of methyl on alpha-linolenic acid is occurred in that in 1ppm, the characteristic peak of double bond on alpha-linolenic acid is occurred in that in 5.35ppm,
This shows to successfully synthesize purer alpha-linolenic acid and prodrugs of paclitaxel.
The synthesis of the alpha-linolenic acid of embodiment 2 and Docetaxel prodrug
Docetaxel 90mg is added in inert environments in the round-bottomed flask of 2.5ml dichloromethane, adds two
Stirring reaction two hours under methylamino pyridine 13.6mg, dicyclohexylcarbodiimide 46mg, alpha-linolenic acid 31mg, normal temperature；With
5.0ml ether dilution, 5% hydrochloric acid, water, saturated sodium-chloride water solution washing organic phase；Organic phase is dried with ammonium sulfate, rotation
Inspissation contracts, and crosses silica gel chromatographic column purifying and obtains product.It is 102mg that alpha-linolenic acid, which is finally given, with Docetaxel prodrug quality,
Yield is about 85%.
The synthesis of the alpha-linolenic acid of embodiment 3 and adriamycin prodrug
Adriamycin 62mg is added in inert environments in the round-bottomed flask of 2.5ml dichloromethane, adds diformazan ammonia
Stirring reaction two hours under yl pyridines 13.6mg, dicyclohexylcarbodiimide 46mg, alpha-linolenic acid 31mg, normal temperature；Use 5.0ml
Ether dilutes, 5% hydrochloric acid, water, saturated sodium-chloride water solution washing organic phase；Organic phase is dried with ammonium sulfate, concentrated by rotary evaporation,
Cross silica gel chromatographic column purifying and obtain product.It is 74mg that alpha-linolenic acid, which is finally given, with adriamycin prodrug quality, and yield is about 80%.
The synthesis of the alpha-linolenic acid of embodiment 4 and 10-hydroxycamptothecine prodrug
10-hydroxycamptothecine 40mg is added in inert environments in the round-bottomed flask of 2.5ml dichloromethane, is added
Stirring reaction two hours under dimethylamino naphthyridine 13.6mg, dicyclohexylcarbodiimide 46mg, alpha-linolenic acid 31mg, normal temperature；
Diluted with 5.0ml ether, 5% hydrochloric acid, water, saturated sodium-chloride water solution washing organic phase；Organic phase is dried with ammonium sulfate, rotation
Inspissation contracts, and crosses silica gel chromatographic column purifying and obtains product.Finally give alpha-linolenic acid is with 10-hydroxycamptothecine prodrug quality
59mg, yield is about 83%.
The preparation of the emulsion composition of the prodrug of embodiment 5
Weigh appropriate cholesterol acid ester 35mg, olein 25mg, non-esterified cholesterol 12mg, prodrug 6mg, plus
Enter a small amount of ethanol hydrotropy, be heated to 60 ± 2 DEG C of meltings, it is standby as oil phase.Weigh appropriate soybean lecithin 35mg, polyethylene glycol 15
Hydroxy stearic acid ester 45mg, plus injection physiological saline 5ml, are heated to 60 ± 2 DEG C, standby as aqueous phase.400r/min's
Under magnetic agitation, aqueous phase is added drop-wise to organic solvent while hot and waved in most oil phase, after completion of dropping, stirred 5min, be prepared into O/W
Type colostrum.By the colostrum prepared rapidly use probe type ultrasonic cell pulverization instrument ultrasonic disperse, power 100w, time 5min,
Ultrasonic 1s intervals 1s.Ice bath 10min solidifies preparation, crosses 0.22um miillpore filters and obtains required preparation.
The emulsion composition particle diameter that prodrug is measured by dynamic light scattering is 51.80nm, and PDI is 0.102, passes through efficient liquid
It is 92.36% that phase chromatogram, which measures entrapment efficiency,.
The preparation of the emulsion composition of the prodrug of embodiment 6
Weigh appropriate cholesterol acid ester 35mg, olein 25mg, non-esterified cholesterol 12mg, prodrug 6mg, plus
Enter a small amount of ethanol hydrotropy, be heated to 60 ± 2 DEG C of meltings, it is standby as oil phase.Weigh appropriate soybean lecithin 35mg, polyethylene glycol 15
Hydroxy stearic acid ester 45mg, plus injection physiological saline 5ml, are heated to 60 ± 2 DEG C, standby as aqueous phase.400r/min's
Under magnetic agitation, aqueous phase is added drop-wise to organic solvent while hot and waved in most oil phase, after completion of dropping, stirred 5min, be prepared into O/W
Type colostrum.The colostrum prepared is disperseed rapidly with high pressure homogenizer, pressure is 200MPa, circulated 10 times.Ice bath 10min makes system
Agent solidifies, and crosses 0.22um miillpore filters and obtains required preparation.
The emulsion composition particle diameter that prodrug is measured by dynamic light scattering is 45.12nm, and PDI is 0.100, passes through efficient liquid
It is 96.09% that phase chromatogram, which measures entrapment efficiency,.
The preparation of the low-density lipoprotein composition of the prodrug of embodiment 7
Weigh appropriate cholesterol acid ester 35mg, olein 25mg, non-esterified cholesterol 12mg, prodrug 6mg, plus
Enter a small amount of ethanol hydrotropy, be heated to 60 ± 2 DEG C of meltings, it is standby as oil phase.Weigh appropriate soybean lecithin 35mg, polyethylene glycol 15
Hydroxy stearic acid ester 45mg, functional polypeptide 15mg, plus injection physiological saline 5ml, are heated to 60 ± 2 DEG C, standby as aqueous phase.
Under 400r/min magnetic agitation, aqueous phase is added drop-wise to organic solvent while hot and waved in most oil phase, after completion of dropping, stirring
5min, is prepared into O/W type colostrums.The colostrum prepared is used into rapidly probe type ultrasonic cell pulverization instrument ultrasonic disperse, power
100w, time 5min, ultrasonic 1s intervals 1s.Ice bath 10min solidifies preparation, crosses 0.22um miillpore filters and obtains required preparation.
Prodrug low-density lipoprotein composition particle diameter is measured for 66.80nm by dynamic light scattering, PDI is 0.163 (figure
4) it is 95.39%, to measure entrapment efficiency by high performance liquid chromatography.And can be observed that by transmission electron microscope prepared
Composition be spherical (Fig. 5).
The preparation of the low-density lipoprotein composition of the prodrug of embodiment 8
Weigh appropriate cholesterol acid ester 35mg, olein 30mg, non-esterified cholesterol 15mg, prodrug 10mg,
A small amount of ethanol hydrotropy is added, 60 ± 2 DEG C of meltings are heated to, it is standby as oil phase.Weigh appropriate soybean lecithin 35mg, polyethylene glycol
15 hydroxy stearic acid ester 35mg, functional polypeptide 13mg, plus injection physiological saline 5ml, are heated to 60 ± 2 DEG C, standby as aqueous phase
With.Under 400r/min magnetic agitation, aqueous phase is added drop-wise to organic solvent while hot and waved in most oil phase, after completion of dropping, is stirred
5min is mixed, O/W type colostrums are prepared into.The colostrum prepared is disperseed rapidly using high pressure homogenizer, pressure is 200MPa, circulated
10 times.Ice bath 10min solidifies preparation, crosses 0.22um miillpore filters and obtains required preparation.
Prodrug low-density lipoprotein composition particle diameter is measured in 45.32nm by dynamic light scattering, PDI is 0.113, is passed through
It is 95.18% that high performance liquid chromatography, which measures entrapment efficiency,.
The taxol of embodiment 9 and alpha-linolenic acid and drug effect in prodrugs of paclitaxel body
Oxter U87 glioma nude mice models are set up, 100mm is reached in gross tumor volume3Physiological saline is injected intravenously respectively
(Control), taxol listing preparation PTXAlpha-linolenic acid and prodrugs of paclitaxel (PALA), are administered for every three days
Once, totally three times.Measurement tumour major diameter and minor axis, draw tumor growth curve daily (with respect to knurl volume-time curve).
From experimental result (Fig. 6), compared with control group, the average tumor of PTX and alpha-linolenic acid and prodrugs of paclitaxel
Inhibiting rate is respectively 51.2% and 50.0%, and there was no significant difference between the two.
The taxol of embodiment 10 and alpha-linolenic acid and safety evaluatio in prodrugs of paclitaxel body
Oxter U87 glioma nude mice models are set up, 100mm is reached in gross tumor volume3Physiological saline is injected intravenously respectively
(Control), taxol listing preparation PTXAlpha-linolenic acid and prodrugs of paclitaxel (PALA), are administered for every three days
Once, totally three times.Daily nude mice body weight, is drawn nude mice body weight change curve, the internal security of different dosing group is weighed with this.
From experimental result (Fig. 7), compared with control group, second day body weight has apparent subtract to PTX upon administration
Gently, and in experimental endpoints body weight it is less than control group；Alpha-linolenic acid and continued weight increase, experimental endpoints after prodrugs of paclitaxel administration
When weight ratio control group it is heavier；This explanation alpha-linolenic acid is safer when being administered in vivo than taxol with prodrugs of paclitaxel.
Drug effect in the low-density lipoprotein composition body of the taxol of embodiment 11 and prodrug
Oxter U87 glioma nude mice models are set up, 100mm is reached in gross tumor volume3Physiological saline is injected intravenously respectively
(Control), taxol listing preparation PTXLow-density lipoprotein composition (PALA-sLDL, reality of prodrug
Apply the preparation of the method for example 8), it is administered once within every three days, totally three times.Measurement tumour major diameter and minor axis, draw tumor growth curve daily
(with respect to knurl volume-time curve).
From experimental result (Fig. 8), compared with control group, PTX has certain tumor growth inhibitory effect, in reality
Terminal is tested, tumor control rate is 51.2%；The low-density lipoprotein composition tumor inhibitory effect of prodrug is significantly stronger than PTX, its
The tumor control rate of experimental endpoints is 72.1%.
The alpha-linolenic acid of embodiment 12 and drug effect in the low-density lipoprotein composition body of prodrugs of paclitaxel and prodrug
Oxter U87 glioma nude mice models are set up, 100mm is reached in gross tumor volume3Physiological saline is injected intravenously respectively
(Control), alpha-linolenic acid and prodrugs of paclitaxel (PALA), (PALA-sLDL is implemented the low-density lipoprotein composition of prodrug
It is prepared by the method for example 8), it is administered once within every three days, totally three times.Measurement tumour major diameter and minor axis, draw tumor growth curve (phase daily
To knurl volume-time curve).
From experimental result (Fig. 9), compared with control group, alpha-linolenic acid with prodrugs of paclitaxel there is certain tumour to give birth to
Long inhibition, in experimental endpoints, tumor control rate is 50.0%；The low-density lipoprotein composition tumor inhibitory effect of prodrug
Alpha-linolenic acid and prodrugs of paclitaxel are significantly stronger than, the tumor control rate of its experimental endpoints is 72.1%.
Drug effect in the emulsion composition of the prodrug of embodiment 13 and the low-density lipoprotein composition body of prodrug
Oxter U87 glioma nude mice models are set up, 100mm is reached in gross tumor volume3Physiological saline is injected intravenously respectively
(Control), the emulsion composition (prepared by PALA-ME, the method for embodiment 5) of prodrug, the low-density lipoprotein composition of prodrug
(prepared by PALA-sLDL, the method for embodiment 8), is administered once for every three days, totally three times.Measurement tumour major diameter and minor axis, draw daily
Tumor growth curve (with respect to knurl volume-time curve).
From experimental result (Figure 10), compared with control group, the emulsion composition of prodrug has certain tumour growth
Inhibition, in experimental endpoints, tumor control rate is 58.8%；The low-density lipoprotein composition tumor inhibitory effect of prodrug is bright
The aobvious emulsion composition for being better than prodrug, the tumor control rate of its experimental endpoints is 72.1%.
Safety evaluatio in the low-density lipoprotein composition body of the taxol of embodiment 14 and prodrug
Oxter U87 glioma nude mice models are set up, 100mm is reached in gross tumor volume3Physiological saline is injected intravenously respectively
(Control), taxol listing preparation PTXLow-density lipoprotein composition (PALA-sLDL, reality of prodrug
Apply the preparation of the method for example 8), it is administered once within every three days, totally three times.Nude mice body weight is determined daily, draws nude mice body weight change curve, with
This weighs the internal security of different dosing group.
From experimental result (Figure 11), compared with control group, second day body weight has apparent subtract to PTX upon administration
Gently, and in experimental endpoints body weight it is less than control group, points out the toxic action of PTX；The low-density lipoprotein composition administration of prodrug
Continued weight increase afterwards, weight ratio control group is heavier during experimental endpoints；This explanation prodrug low-density lipoprotein composition compares Japanese yew
Alcohol is safer.
Ex vivo Tumor situation of the different preparations of embodiment 15 to lotus U87 glioma nude mices
Oxter U87 glioma nude mice models are set up, 100mm is reached in gross tumor volume3Physiological saline is injected intravenously respectively
(Control), taxol listing preparation PTXAlpha-linolenic acid and prodrugs of paclitaxel (PALA), the emulsion of prodrug
Composition (prepared by PALA-ME, the method for embodiment 5), low-density lipoprotein composition (PALA-sLDL, the side of embodiment 8 of prodrug
It is prepared by method), it is administered once within every three days, totally three times.Each group nude mice is put to death in experimental endpoints, solution cuts tumour and taken pictures.
Experimental result (Figure 12) understands that control group tumour is maximum in each group；PTX and alpha-linolenic acid and taxol
Prodrug has suitable tumor inhibitory effect, and Ex vivo Tumor is less than control group；The emulsion composition of prodrug and the low-density of prodrug
Lipoprotein composition Ex vivo Tumor is less than PTX and alpha-linolenic acid and prodrugs of paclitaxel group, and the low-density lipoprotein combination of prodrug
Thing Ex vivo Tumor is minimum, and illustrating the low-density lipoprotein composition of prodrug has best internal antitumor drug effect.
The TUNEL apoptosis analysis of the tumor tissues of embodiment 16
Frozen section is made in the tumour separated, uses apoptosis feelings of the TUNEL apoptosis detection kit to different dosing group
Condition is evaluated, eventually through confocal microscopy.
Experimental result (Figure 13) is understood, compared with control group, PTXBefore group and alpha-linolenic acid and taxol
The Apoptosis quantity of medicine (PALA) group substantially increases, and apoptosis situation is suitable；The emulsion composition group of prodrug and prodrug it is low
The apoptosis of tumor cells quantity of density lipoprotein composition group is more than PTXBefore group and α-linoleic acid and taxol
Medicine group, and the apoptosis-induced effect of low-density lipoprotein composition of prodrug is best.
Claims (10)
1. the prodrug of a kind of leukotrienes and cancer therapy drug, it is characterised in that leukotrienes passes through the carboxylic on leukotrienes with cancer therapy drug
The hydroxyl or amino groups of base and cancer therapy drug are covalently attached to, and the leukotrienes includes alpha-linolenic acid or gamma-Linolenic acid, described
Cancer therapy drug includes：Taxol, Docetaxel, adriamycin, Doxorubicin, cis-platinum, camptothecine, 10-hydroxycamptothecine, 9- hydroxyls
Base camptothecine, SN38 (SN-38), TPT, Irinotecan, vincaleukoblastinum, vincristine, long fields for spring sowing
Pungent, vinflunine, vinpocetine, gemcitabine, polypeptide series antineoplastic medicament (such as Goserelin, Triptorelin), amino quinoline azoles
Quinoline series antineoplastic medicament (such as Gefitinib, Tarceva, Lapatinib) or TNF, preferably taxol and many
Western taxol.
2. the preparation method of the prodrug of leukotrienes as claimed in claim 1 and cancer therapy drug, it is characterised in that the step of this method
It is rapid as follows：
(1) under nitrogen protection, antineoplastic is dissolved in solvent, adds catalyst and dehydrating agent, under stirring, add
Leukotrienes, is stirred at room temperature reaction；
(2) above-mentioned reaction resulting material is taken, ether is added, crosses and filters out precipitation, filtrate is full with 5% hydrochloric acid, water and sodium chloride successively
Respectively washed with the aqueous solution three times, collect organic phase, rotated drying using Rotary Evaporators, pass through the isolated prodrug of chromatographic column.
Wherein, in the above method solvent refer to one kind in dichloromethane, chloroform, DMF or dioxane or
Its mixed solution；Catalyst is dimethylamino naphthyridine；Dehydrating agent refers to dicyclohexylcarbodiimide, 1- ethyls -3- (3- diformazans
Base aminopropyl) carbodiimide or one kind or its mixture in DIC, the reaction time is 2-24 hours.It is anti-swollen
Tumor medicine：Linolenic mol ratio is 1:0.1-10；Catalyst is 1 with linolenic mol ratio:0.1-10, preferably 1:1；Dehydration
Agent is 1 with leukotrienes mol ratio:0.1-10, preferably 1:2.
3. prodrug as claimed in claim 1 is contained forms composition, said composition in artificial synthesized low-density lipoprotein
Or be the low-density lipoprotein composition of prodrug.
4. the preparation method of the low-density lipoprotein composition of prodrug as claimed in claim 3, it is characterised in that breast can be passed through
Change any one preparation in ultrasonic method or high-pressure stripping, concrete operation step is as follows：
(1) prodrug of appropriate cholesterol, cholesteryl ester, triglycerides, leukotrienes and cancer therapy drug is weighed, is added a small amount of organic molten
Agent hydrotropy, is heated to 50-80 DEG C of melting, standby as oil phase.
(2) appropriate low-density lipoprotein functional polypeptide and phosphatide and the amphipathic nature material of auxiliary are weighed, appropriate injection life is added
Salt solution is managed, 50-80 DEG C is heated to, it is standby as aqueous phase.
(3) under agitation, aqueous phase is added drop-wise to organic solvent while hot to wave in most oil phase, is prepared into O/W type colostrums.
(4) colostrum prepared is used into rapidly probe type ultrasonic cell pulverization instrument ultrasonic disperse, power 80-300w, time 3-
10min。
(5) or by above-mentioned colostrum disperseed using methods such as high pressure homogenizers.
(6) ice-water bath cooling 5-30min solidifies preparation, crosses 0.22 μm of miillpore filter and obtains required composition.
5. preparation method as claimed in claim 4, wherein amphipathic nature material are selected from：Natural phospholipid, semi-synthetic phosphatide, synthesis phosphorus
Fat, the polyethylene glycol conjugate of phosphatide, Solutol HS 15, PLURONICS F87, cetomacrogol 1000 vitamin
The mixture of E succinates or their mixture, preferably natural phospholipid and Solutol HS 15；Organic solvent
It is selected from：In ethanol, glycerine, propane diols, polyethylene glycol, DMA, benzyl benzyl ester, ethyl oleate, phenmethylol
One or more mixtures, preferred alcohol.
6. the low-density lipoprotein composition of prodrug as claimed in claim 3, it is characterised in that each component weight percent is such as
Under：
Drug weight percentage is 1%-15%, preferably 3%-10%；
Low-density lipoprotein functional polypeptide percentage by weight is 5%-25%, preferably 8%-20%；
Phosphatide percentage by weight is 8%-24%, preferably 18%-22%；
The percentage by weight of cholesterol component and triglycerides etc. is 30%-75%, preferably 50%-68%.
7. the low-density lipoprotein composition of prodrug as claimed in claim 3, it is characterised in that said composition average grain diameter is big
It is small in 10-100nm.
8. the low-density lipoprotein composition of prodrug as claimed in claim 3, it is characterised in that add medicine and pharmacology as needed
Upper acceptable auxiliary material, such as coemulsifier, stabilizer, pH value regulator and antioxidant, can by pharmacy routine side
Method prepares into capsule, oral liquid or external preparation etc..
9. the low-density lipoprotein composition of prodrug as claimed in claim 3, it is characterised in that can freeze and lyophilized system is made
Agent, can be disperseed when using with 5% glucose solution, physiological saline, water for injection or their mixture, form prodrug
Low-density lipoprotein composition.
10. the low density lipoprotein of the prodrug described in the prodrug and claim 3 of leukotrienes as claimed in claim 1 and cancer therapy drug
Application of the protein composition in oncotherapy, it is characterised in that the tumour is selected from breast cancer, liver cancer, oophoroma, lung cancer, head
The one or more of neck cancer or glioma, preferably glioma and liver cancer.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20080125380A1 (en) * | 1999-03-09 | 2008-05-29 | Luitpold Pharmaceuticals, Inc. | Fatty acid-anticancer conjugates and uses thereof |
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2016
- 2016-04-18 CN CN201610239404.5A patent/CN107303391A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20080125380A1 (en) * | 1999-03-09 | 2008-05-29 | Luitpold Pharmaceuticals, Inc. | Fatty acid-anticancer conjugates and uses thereof |
Non-Patent Citations (3)
| Title |
|---|
| MINA NIKANJAM ETAL: "Synthetic nano-LDL with paclitaxel oleate as a targeted drug delivery vehicle for glioblastoma multiforme", 《JOURNAL OF CONTROLLED RELEASE》 * |
| XI-YU KE ETAL: "The therapeutic efficacy of conjugated linoleic acid-paclitaxel on glioma in the rat", 《BIOMATERIALS》 * |
| 顾瀅熙等: "拟低密度脂蛋白脂质核心的制备及性质考察", 《沈阳药科大学学报》 * |
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