CN102397280B - Application of 2 alpha-hydroxy protopanoxadiol medicine - Google Patents
Application of 2 alpha-hydroxy protopanoxadiol medicine Download PDFInfo
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- CN102397280B CN102397280B CN 201110362152 CN201110362152A CN102397280B CN 102397280 B CN102397280 B CN 102397280B CN 201110362152 CN201110362152 CN 201110362152 CN 201110362152 A CN201110362152 A CN 201110362152A CN 102397280 B CN102397280 B CN 102397280B
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Abstract
The invention belongs to the fields of chemical industries and medicines, and relates to the application of 2 alpha-hydroxy protopanoxadiol in preparation of antitumor medicines. Antitumor cells comprise leukemia or pancreatic cancers. The 2 alpha-hydroxy protopanoxadiol belongs to a natural product and has certain clinical application value. A small molecular compound is used as a new antitumor medicine or an auxiliary component of the new antitumor medicine for development, has an obvious antitumor effect, is environment-friendly and provides a new method for treating and healing tumors.
Description
Technical field
The invention belongs to chemical field and field of medicaments, relate to the application of 2 Alpha-hydroxy protopanoxadiols in the preparation antitumor drug.
Background technology
2 Alpha-hydroxy protopanoxadiols, structural formula is as follows.
1998, Cui Jian-Fang etc. utilizes alkali excision method to prepare the 2 Alpha-hydroxy protopanoxadiols (chemical compound 6 in the table 1, Alkaline cleavage of gypenosides and characterization of dammarane-type aglycons by gas chromatography-mass spectrometry.Phytochemical Analysis.9 (3), 128-133).1999, Cui etc. have reported the method (chemical compound 7 of isolation identification 2 Alpha-hydroxy protopanoxadiols from Herb Gynostemmae Pentaphylli, Gynostemma pentaphyllum:identification of major sapogenins and differentiation from Panax species.European Jo urnal of Pharmaceutical Sciences.8 (3), 187-191).But, about the function of 2 Alpha-hydroxy protopanoxadiols, especially clear and definite medicinal function, present research and report are very few.
2 Alpha-hydroxy protopanoxadiols are natural products, have certain clinical use value.Along with people to the going deep into of the chemistry and biology of this compounds research, its molecular mechanism of action will progressively clear and definite, this will further promote chemical constitution modification and the structure activity study of this compounds, and help to improve the medical value of this compounds.
Summary of the invention
The purpose of this invention is to provide the application of 2 Alpha-hydroxy protopanoxadiols in the preparation antitumor drug.
The invention provides the application of 2 Alpha-hydroxy protopanoxadiols in the preparation antitumor drug.Described antitumor drug can be anti-cancer of pancreas medicine or anti-leukemia medicine.This antitumor drug can be injection or tablet.
The invention provides a kind of method that suppresses tumor cell in vitro propagation, 2 Alpha-hydroxy protopanoxadiols are added in the culture fluid of tumor cell.Described tumor cell can be liver blood cancerous cell or pancreatic cancer cell.The hepatoma carcinoma cell that adopts in one embodiment of the present of invention is QGY.Generally speaking, the final concentration that adds 2 Alpha-hydroxy protopanoxadiols is 1-100 μ M.For example, 1-5 μ M, 1-10 μ M, 1-20 μ M, 1-50 μ M, 5-10 μ M, 5-20 μ M, 5-50 μ M, 5-100 μ M, etc.
The present invention also provides a kind of antitumor drug, and the active component of described antitumor drug is 2 Alpha-hydroxy protopanoxadiols.Described tumor can be cancer of pancreas or leukemia.
Micromolecular compound 2 Alpha-hydroxy protopanoxadiols of the present invention are available from Chinese Academy of Sciences institute of materia medica, and high performance liquid chromatography is surveyed purity, HPLC 〉=98%.
Micromolecular compound of the present invention also can adopt the preparation method preparation of various routines.For example, adopt the method for artificial chemosynthesis.
Utilize micromolecular compound of the present invention, by various conventional screening techniques, can filter out with 2 Alpha-hydroxy protopanoxadiols interactional material takes place, as receptor, inhibitor or antagonist etc.
The present invention and inhibitor, antagonist etc. when when (administration) used in treatment, can provide different effects.Usually, but these materials are formulated in nontoxic, inertia with pharmaceutically acceptable aqueous carrier medium in, wherein pH is about 5-8 usually, preferably pH is about 6-8, although pH value can change to some extent with being formulated Substance Properties and disease to be treated.The pharmaceutical composition for preparing can carry out administration by conventional route, comprising (but being not limited to): intramuscular, intraperitoneal, subcutaneous, Intradermal or topical.
Be example with 2 Alpha-hydroxy protopanoxadiols of the present invention, can be with itself and suitable pharmaceutically acceptable carrier coupling.This class pharmaceutical composition contains chemical compound and pharmaceutically acceptable carrier or the excipient for the treatment of effective dose.This class carrier comprises (but being not limited to): saline, buffer, glucose, water, glycerol, ethanol and combination thereof.Pharmaceutical preparation should be complementary with administering mode.2 Alpha-hydroxy protopanoxadiols of the present invention can be made into the injection form, for example are prepared by conventional method with normal saline or the aqueous solution that contains glucose and other adjuvant.Pharmaceutical composition such as tablet and capsule can be prepared by conventional method.Pharmaceutical composition such as injection, solution, tablet and capsule should be made under aseptic condition.The dosage of active component is the treatment effective dose, for example every day about 1 microgram/kg body weight-Yue 5 mg/kg body weight.In addition, 2 Alpha-hydroxy protopanoxadiols of the present invention also can use with the other treatment agent.
When 2 Alpha-hydroxy protopanoxadiols of the present invention are used as medicine, 2 Alpha-hydroxy protopanoxadiols for the treatment of effective dose can be applied to mammal, wherein should treat effective dose usually at least about 10 micrograms/kg body weight, and in most of the cases be no more than about 8 mg/kg body weight, preferably this dosage is about 10 micrograms/kg body weight-Yue 1 mg/kg body weight.Certainly, concrete dosage also should be considered factors such as route of administration, patient health situation, and these all are within the skilled practitioners skill.
The invention provides the application of 2 Alpha-hydroxy protopanoxadiols in the preparation antitumor drug.2 Alpha-hydroxy protopanoxadiols are natural products, can obviously suppress the propagation of tumor cell.Micromolecular compound of the present invention is developed as new antitumor drug or its auxiliary element, and tumor killing effect is obvious, environmental protection, and will and cure tumor for treatment provides a kind of new approach and means.
The specific embodiment
Experimental technique:
1. cell recovery
1) from liquid nitrogen container, takes out frozen pipe, directly drop in 37 ℃ of warm water, and shake frequently and make it melt as early as possible.
2) from 37 ℃ of water-baths, take out frozen pipe, with suction pipe sucking-off cell suspension, inject centrifuge tube and add culture fluid more than 10 times, mix the back low-speed centrifugal, abandon supernatant, repeat again to wash once with culture fluid.
3) with after the suitable dilution of culture fluid, the inoculated and cultured bottle is placed on 37 ℃ of incubators and leaves standstill cultivation, changes culture fluid next day, continues to cultivate.Go down to posterity when being cultured to finite concentration.The PANC-1 cell culture is in containing the DMEM high glucose medium of 10% hyclone, and the k562 cell culture contains 100U/ml penicillin and 100 μ g/ml streptomycins in the culture medium in containing 1640 culture medium of 10% hyclone.
2. passage is cultivated
The situation of observation of cell growth every day goes down to posterity when converging when cell grows to about 90% in culture bottle, goes down to posterity once every 2-4 days approximately.One bottle goes down to posterity into three bottles, or a 25cm
2Go down to posterity in a 75cm
2Culture bottle in.Method:
1) with 1 * phosphate buffer washed cell once.
2) add the digestion of 2-3ml trypsinization liquid, place 37 ℃ of incubator numbers minute.Pat Tissue Culture Flask with hands, make cell separation.
3) suitable culture medium with the Gibico hyclone that contains 10-15% stops trypsinization.Cell is sub-packed in the new culture bottle, continues to cultivate.
3. cell cryopreservation
1) get the cell tryptase protease digestion that is cultured to exponential phase, be collected in the centrifuge tube and counting, centrifugal.
2) reject trypsin and old culture fluid add the frozen culture fluid (containing 10%DMSO, 40%DMEM and 50%Gibico hyclone) that configures, and the ultimate density of cell is 0.5-1 * 10 in the cryopreserving liquid
7/ ml.Blow and beat gently with suction pipe and to make cell even, divide in the aseptic frozen pipe of packing into then, every pipe adds 1-1.5ml.
3) frozen pipe is put into freezing storing box and put-80 ℃ of quick-freezings, move in the liquid nitrogen container after 5 hours and preserve.
4. medicine is prepared:
2 Alpha-hydroxy protopanoxadiols are dissolved in DMSO (dimethyl sulfoxide), and the mother solution that is mixed with 100mM or 50mM is standby.
Embodiment 1MTS method is measured 2 Alpha-hydroxy protopanoxadiols to the growth inhibited effect of pancreatic cancer cell
3000/hole of PANC-1 cell (available from Chinese Academy of Sciences's cell bank) is seeded to 96 orifice plates, cultivate and made it adherent back adding 2 Alpha-hydroxy protopanoxadiols (available from Shanghai Pharmaceutical Inst., Chinese Academy of Sciences) in 24 hours, if 6 Concentraton gradient, each concentration are established 3 multiple holes.Cell is at 37 ℃, 5%CO
2Cultivate after 72 hours under the condition, outwell culture fluid, measure cell survival rate with MTS test kit (Promega company).
Method of testing is: cell is washed one time with serum-free medium, added the MTS chromophoric solution (adding 2ml solution 1 and 100 μ l solution 2 in the 10ml serum-free medium, fully mixings) for preparing in advance according to the amount of 100 μ l/well.Do not have the hole of cell to be made as this bottom outlet with one, absorb in order to the bias light of calibration solution.Cell is put into cell culture incubator to be continued to cultivate 2～4 hours, read absorbance value (reference wavelength 630-700nm with microplate reader then, measure wavelength 490nm), calculate cell survival rate, to measure hole absorbance value/control wells absorbance value as the numerical value of cell survival rate.According to cell survival rate, calculate 2 Alpha-hydroxy protopanoxadiols to the IC50 value of PANC-1 cell.
IC50 refers to be suppressed a half
InhibitorConcentration.Here be the PANC-1 cell quantity and be the concentration of a half 2 Alpha-hydroxy protopanoxadiols of contrast.The calculating of IC50 generally need be measured the dosage effect more than 5, obtains function calculation by curve fitting again and gets.
The result: 2 Alpha-hydroxy protopanoxadiols are 664 μ M to the IC50 value of PANC-1 cell.
Embodiment 22 Alpha-hydroxy protopanoxadiols are to human leukemia cell's growth inhibited effect
Utilize cck-8 test kit (Japanese colleague's chemistry institute) to detect.
Step:
1) k562 cell (available from ATCC) is planted in 96 orifice plates uniformly, every porocyte number is 10000.
2) treat adherent, the back dosing of spending the night, (2 Alpha-hydroxy protopanoxadiol concentration are respectively 50,16.67,5.56,1.85,0.62 μ M, and each concentration has 3 multiple holes in dosing.
3) cultivate 48 hours, complete medium is replaced to the mixture (10: 1) of serum-free medium and CCK8, hatched 2 hours in 37 ℃ of incubators.
4) being to measure wavelength with 450nm, is the contrast wavelength with 650nm, measures reading on microplate reader.
The result: 2 Alpha-hydroxy protopanoxadiols are 4.77 μ M to the IC50 value of k562 cell.
Claims (6)
1.2 the application of Alpha-hydroxy protopanoxadiol in the anti-cancer of pancreas of preparation or leukemia medicament.
2. application as claimed in claim 1 is characterized in that, described medicine is anti-cancer of pancreas medicine.
3. application as claimed in claim 1 is characterized in that, described medicine is anti-leukemia medicine.
4. application as claimed in claim 1 is characterized in that, described medicine is injection or tablet.
5. a method that suppresses tumor cell in vitro propagation is characterized in that, 2 Alpha-hydroxy protopanoxadiols is added in pancreatic cancer cell or leukaemia's the culture fluid.
6. method as claimed in claim 5 is characterized in that, the final concentration that adds 2 Alpha-hydroxy protopanoxadiols is 1-100 μ M.
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| Application Number | Priority Date | Filing Date | Title |
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| CN 201110362152 CN102397280B (en) | 2011-11-15 | 2011-11-15 | Application of 2 alpha-hydroxy protopanoxadiol medicine |
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| CN 201110362152 CN102397280B (en) | 2011-11-15 | 2011-11-15 | Application of 2 alpha-hydroxy protopanoxadiol medicine |
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| CN102397280B true CN102397280B (en) | 2013-07-10 |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104208074B (en) * | 2013-06-01 | 2017-10-10 | 复旦大学 | Purposes of the 2 α hydroxyl protopanoxadiols in tumor multi-drug resistance reversal agents are prepared |
| CN104586860A (en) * | 2013-11-01 | 2015-05-06 | 中国科学院上海药物研究所 | Use of dammarane-type triterpene derivatives |
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