CN101810608B - Anti-tumor small molecule compound targeting at human phosphatidylethanolamine-bindingprotein-4 - Google Patents
Anti-tumor small molecule compound targeting at human phosphatidylethanolamine-bindingprotein-4 Download PDFInfo
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Abstract
The invention relates to an anti-tumor small molecule compound targeting at human phosphatidylethanolamine-bindingprotein-4, which is synchronized by a medicine virtual screening platform based on the combination of bioinformatics and a computer technology. The small molecule compound has the advantages of good permeability, small toxin and side effect, simple structure, easy synchronization and the like. In addition, the compound can be used together with a tumor necrosis factor (TNF-Alpha) to enhance the anti-tumor effect of the latter, so as to play the role of curing tumor. Therefore, the compound is expected to become a new anti-tumor medicine. The invention also relates to a medicine compound of the small molecule compound and purposes thereof.
Description
Technical field
The invention belongs to biotechnology and medical domain.Particularly, the present invention relates to a kind of based on bioinformatics and computer technology, the specificity obtaining through virtual screening suppresses the micromolecular compound of hPEBP4 albumen, this micromolecular compound has targeting antitumor cell hPEBP4 protein function, inhibition tumor cell growth and causes the function of apoptosis of tumor cells, thereby plays the effect for the treatment of tumor.The invention still further relates to the pharmaceutical composition that contains this micromolecular compound, and disclose the method for disease treatment by this compound medicine, the particularly purposes in the treatment of malignant tumor.
Background technology
Tumor is the second largest killer of face of mankind nowadays.Along with the development of oncomolecularbiology and the decoding of gene information, scientist has disclosed the molecule working in tumor occurs in a large number.On this basis, neoplasm targeted therapy arises at the historic moment.It is service for patients that targeted therapy moves towards clinical by the gene data of magnanimity from laboratory, becomes one of the most breathtaking field in oncology studies.Can say that recent two decades carrys out the progress the most significantly that therapeutic field of tumor obtains and is just coming into operation of multiple targeted drugs.Area of computer aided drug screening by software on protein three-dimensional structure figure directly and compound achieve a butt joint and screen, not only greatly shorten the cycle of conventional medicament research and development, and pointed strong, biological tolerance is good, onset is rapid, the advantage that cell permeability is strong is fresh combatants of neoplasm targeted therapy.
Human phosphotidylethanolabinding binding protein 4 (hPEBP4) is a new gene of recently finding.It is general high expressed in ovary, mammary gland, prostate and B lymphatic system tumor, and has no expression in normal structure.Wang Xiaojian etc. report for the first time that in 2004 the apoptosis that expression causes with breast cancer cell TNF alpha antibody excessively of this gene is relevant, the mechanism of its apoptosis inhibit is blocked relevant (the Wang X etc. of activation of downstream signal with MEK in conjunction with Raf-1 with hPEBP4, J Biol Chem, 2004,279:45855-45864).
Separately there is experiment to show that hPEBP4 may also participate in suppressing the apoptosis of the carcinoma of prostate that TRAIL causes.Tumorigenesis is not yet thoroughly illustrated so far through the exploration of more than 100 years, but the escape (" evasion of apoptosis ") to apoptotic signal has participated in the viewpoint of cell carcinogenesis and has been widely accepted (Hanahan, D. & Weinberg, R.A.Cell, 2000,100:57-70).
Since hPEBP4 is just specifically at tumor cell high expressed, and participate in the opposing of tumor to apoptotic signal, therefore may can cancel its apoptosis inhibit feature for the micromolecular compound of this molecule, and will be likely and the drug combination such as TNF-α, open up a treatment tumor Xin road.
At present, although developed the compound of some inhibition tumor growths, but people still need to develop new having, inhibition tumor cell is grown or inducing tumor cell is adjusted the active compound of dying.
Summary of the invention
Object of the present invention is just to provide a kind ofly has inhibition tumor cell growth or inducing tumor cell is adjusted the active compound of dying.Another object of the present invention is just to provide the purposes of described compound.
In a first aspect of the present invention, the purposes that acceptable salt in compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone [3-(2-(2-hydroxy-5-nitrophenyl)-5-phenyl-1H-imidazol-4-yl) phenyl) (phenyl) methanone], its pharmacy, health care conduct and learning or bromatology or ester or their mixture are provided, it adjusts the material of dying for the preparation for the treatment of tumor, inhibition tumor cell growth and/or inducing tumor cell.
In an embodiment of the invention, described material is pharmaceutical composition, Halth-care composition or food additive.
In yet another embodiment of the present invention, described compound, its pharmaceutically or in health care conduct and learning acceptable salt or ester or their mixture account for the 0.001-100wt% of described material gross weight.
In a preference, described compound, its pharmaceutically or in health care conduct and learning acceptable salt or ester or their mixture account for the 1-95wt% of described material gross weight, preferably 5-90wt%, more preferably 10-80wt%.
In another preference, described material is only containing acceptable salt or ester or their mixture in compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone, its pharmacy, health care conduct and learning or bromatology.
In another preference, described compositions contains acceptable carrier or excipient in pharmacy, health care conduct and learning or bromatology.
In another preferred embodiment of the present invention, described compositions is unit dosage forms or multi-pharmaceutics, and wherein said compound, its pharmaceutically or in health care conduct and learning the content of acceptable salt or ester or their mixture be 0.05-50000mg/ agent, preferably 0.1-10000mg/ agent, more preferably 0.5-5000mg/ agent.
In yet another embodiment of the present invention, described compositions also contains other medicine that suppresses tumor.
In a preference, the described medicine that other suppresses tumor is selected from: cause apoptosis medicine, tumor vessel inhibition medicine or immunoregulation medicament, preferably cause apoptosis medicine.
In a preference, described in cause apoptosis medicine and be selected from: tumor necrosis factor or TRAIL, preferably tumor necrosis factor, more preferably humanTNF-α.
In yet another embodiment of the present invention, the dosage form of described compositions is tablet, capsule, powder agent, granule, suspensoid or injection.
In yet another embodiment of the present invention, described tumor or tumor cell are tumor or the tumor cells of expressing human phosphotidylethanolabinding binding protein 4.
In a preference, described tumor or tumor cell are selected from: breast carcinoma, pulmonary carcinoma, gastric cancer, carcinoma of prostate, ovarian cancer, colon cancer, carcinoma of prostate, hepatocarcinoma or B lymphatic system tumor, preferably breast carcinoma, pulmonary carcinoma, gastric cancer, colon cancer, carcinoma of prostate or hepatocarcinoma, more preferably breast carcinoma.
In another preference, human phosphotidylethanolabinding binding protein 4 expressions of described tumor or tumor cell, higher than normal cell, preferably exceed 30%, more preferably exceed 50%.
In a second aspect of the present invention, a kind of compositions is provided, described compositions contains:
(1) acceptable salt or ester or their mixture in compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone, its pharmacy, health care conduct and learning or bromatology;
(2) cause apoptosis medicine; And
(3) acceptable carrier or excipient in pharmacy, health care conduct and learning or bromatology.
In a preference, the weight ratio of described component (1) and component (2) is 1: 1000 to 1000: 1, preferably 1: 500-500: 1, more preferably 1: 100-100: 1, more preferably 1: 50-50: 1.
In another preference, described component (1) accounts for the 0.001-99.9wt% of composition total weight, preferably 1-95wt%, more preferably 5-90wt%, more preferred 10-80wt%.
In yet another embodiment of the present invention, the described apoptosis medicine that causes is tumor necrosis factor.
In a preference, described compositions also contains the therapeutic agent that is selected from lower group: chemotherapeutic, as Fructus Bruceae breast, amycin, tamoxifen, 5-fluorouracil, Tegadifur, harringtonine, cytosine arabinoside, NSC-241240, paclitaxel, flutamide, ifosfamide, doxifluridine, Glass platinum, bend azoles or teniposide etc.; Tumor vessel suppresses medicine, as angiostatin, endostatin etc.; Or immunoregulation medicament is as polysaccharide-peptide, lentinan etc.
In a third aspect of the present invention, the purposes that acceptable salt in compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone, its pharmacy, health care conduct and learning or bromatology or ester or their mixture are provided, it is for the preparation of the inhibitor of human phosphotidylethanolabinding binding protein 4.
In a fourth aspect of the present invention, a kind of purposes by the mixture that in compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone, its pharmacy, health care conduct and learning or bromatology, acceptable salt or ester or their mixture and tumor necrosis factor form is provided, and it adjusts the compositions of dying for the preparation for the treatment of tumor, inhibition tumor cell growth and/or inducing tumor cell.
In another aspect of this invention, a kind of purposes of the mixture being made up of (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone and tumor necrosis factor is provided, it is characterized in that, adjust the compositions of dying for the preparation for the treatment of tumor, inhibition tumor cell growth and/or inducing tumor cell.
In another preference, described compositions is pharmaceutical composition.
In another preference, described compositions also contains and causes apoptosis medicine, is more preferably humanTNF-α.
In another preference, described medicine composition dosage form is tablet, capsule, powder agent, granule, suspensoid or injection.
In another preference, described tumor or tumor cell are tumor or the tumor cells of expressing hPEBP4 albumen.Preferably, described tumor is selected from lower group: breast carcinoma, pulmonary carcinoma, gastric cancer, colon cancer, carcinoma of prostate, hepatocarcinoma etc.; More preferably, described tumor is selected from lower group: breast carcinoma.
This aspect on the other hand in, a kind of compositions is provided, and described compositions contains pharmaceutically acceptable carrier and (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone and causes apoptosis medicine.
In another preference, the described apoptosis medicine that causes is tumor necrosis factor.
In another preference, described compositions also contains the therapeutic agent that is selected from lower group: Fructus Bruceae breast, amycin, tamoxifen, 5-fluorouracil, Tegadifur, harringtonine, cytosine arabinoside, NSC-241240, paclitaxel, flutamide, ifosfamide, doxifluridine, Glass platinum, bend azoles or teniposide, angiostatin, endostatin, polysaccharide-peptide, lentinan.
In another preference, described (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone and cause that to adjust the weight ratio between medicine of dying be 1: 1000 to 1000: 1.
This aspect on the other hand in, provide a kind of purposes of (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone, for the preparation of the inhibitor of human phosphotidylethanolabinding binding protein 4.
This aspect on the other hand in, a kind of purposes of the mixture being made up of (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone and tumor necrosis factor is provided, has adjusted the compositions of dying for the preparation for the treatment of tumor, inhibition tumor cell growth and/or inducing tumor cell.
In another preference, the present invention for tumor be the tumor of hPEBP4 abnormal expression in growth course, as breast carcinoma.
This aspect on the other hand in, a kind of method for the treatment of tumor is provided, it comprises step: give needs treat mammalian object use micromolecular compound of the present invention.
In another preference, before the method is also included in and uses micromolecular compound of the present invention, among or tumour medicine, the especially tumor necrosis factor etc. of using afterwards other cause and adjust the medicine of dying.
Other side of the present invention, due to disclosure herein, is apparent to those skilled in the art.
Accompanying drawing explanation
Below in conjunction with accompanying drawing, the present invention is further elaborated.
Fig. 1 has shown the surface plasmon resonance result of DC240016 and the external combination of hPEBP4;
Fig. 2 has shown the cell growth inhibition that DC240016 enhance TNF-α causes, wherein compound used therefor concentration is 5 μ M, and TNF-α concentration is 20ng/ml;
Fig. 3 has shown the apoptosis of DC240016 enhance TNF-α induction, and wherein compound used therefor concentration is 5 μ M, and TNF-α concentration is 20ng/ml.
The specific embodiment
The inventor, through extensive and deep research, finds that the expression of hPEBP4 in Partial tumors cell often raises, and the expression and the function that suppress hPEBP4 can the propagation of inhibition tumor cell and the oncogenic activities of inside and outside.For this reason, the inventor use software from tens thousand of candidate compounds, sifted out some may with the protein bound micromolecular compound of hPEBP4; Use subsequently the method for external combination experiment and MTT to carry out second to these compounds and take turns screening, obtain first the Compound D C240016 of enhance TNF-α to breast cancer cell line MCF-7 inhibited proliferation strongly, i.e. micromolecular compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone.Complete on this basis the present invention.
Particularly, the inventor's research shows, the selectivity high expressed of hPEBP4 in breast cancer tumour tissue, and in normal galactophore tissue, have no expression, prompting hPEBP4 may occur and grow closely related with tumor.
In L929 cell line, the apoptosis that can obviously resist TNF-α induction is expressed in crossing of hPEBP4, Promote cell's growth, and pointing out it may be a molecule with anti-apoptotic effect.
The inventor's research also finds, in the breast cancer tumour cell of high expressed hPEBP4, the expression that suppresses hPEBP4 can strengthen the breast cancer tumour cell sensitivity apoptosis-induced to TNF-α, the clonality of inhibition tumor cell.Prompting hPEBP4 is likely the action target spot for the treatment of breast carcinoma.
The inventor's research shows, anti-apoptotic effect molecule hPEBP4 in the processes such as Growth of Cells regulation and control, apoptosis, tumor generation, play an important role.Therefore, hPEBP4 probably in the diagnosis of clinical tumor and treatment as potential candidate's target.
On this basis, the inventor has screened a large amount of compounds, thereby obtain a kind of micromolecular compound that can effectively suppress tumor, described micromolecular compound is specifically at the anti-apoptosis molecule of protein level targeting---hPEBP4 albumen, biological behaviour to the tumor cell of expressing hPEBP4 is intervened, thereby the function that effectively suppresses hPEBP4, reaches antitumous effect.
Active component
As used herein, term " micromolecular compound of the present invention ", " Compound D C240016 " or " the compounds of this invention ", " (phenyl) of the present invention ketone derivatives " are used interchangeably, and all point out micromolecular compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone and pharmaceutically acceptable salt and reactive derivative.
(3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone be a kind of targeting in the lead compound of human phosphotidylethanolabinding binding protein (hPEBP4), its structural formula is as follows:
(3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone is a kind of known compound, can buy from commercial channels (as can be purchased from from Dutch Specs company), methodology of organic synthesis that also can be conventional prepares.
In the present invention, preferably " active component " refers to such micromolecular compound: described compound and human hPEBP 4 albumen can mutually combine, and described compound is combined with TNF-α can make the growth of MCF-7 Breast Cancer Cell be reduced to (i.e. reduction at least 60%) below 40%, preferably be reduced to below 38%, more preferably be reduced to below 35%, or be reduced between the scope take above numerical value as end points.
As used herein, in the present invention (phenyl) ketone derivatives used can with by pharmaceutically or the acceptable acid of physiology or the derivative salt form of alkali use.These salt include, but is not limited to: the salt forming with following mineral acid: example hydrochloric acid, sulphuric acid, nitric acid or phosphoric acid; With the salt of following organic acid formation, as acetic acid, oxalic acid, succinic acid or maleic acid; And other salt, include but not limited to: with the salt of alkali metal or alkaline-earth metal (as sodium, potassium, calcium or magnesium) formation.The particularly preferred salt of one class is sodium salt or potassium salt.
The present invention also comprises the compounds of this invention for example, existing with the form of " prodrug " of ester (carbamate) or other routine (when with this form administration, can change in vivo active part).
Micromolecular compound of the present invention can effectively suppress the function of human hPEBP 4 albumen, thus the propagation of inhibition tumor cell, the apoptosis of promotion tumor cell.Particularly, micromolecular compound of the present invention is at molecular level for human hPEBP 4 albumen, and the biological behaviour of the tumor cell to positive expression human hPEBP 4 albumen reverses.Experiment proves: (1) suppresses the propagation that the expression of hPEBP4 and function can inhibition tumor cells, promotes apoptosis of tumor cells; (2) suppress the external oncogenicity that the expression of hPEBP4 and function can inhibition tumor cells in body; (3) suppress the growth in can inhibition tumor cell body of the expression of hPEBP4 and function.
Pharmaceutical composition
The present invention also comprises the pharmaceutical composition that contains (phenyl) ketone derivatives and pharmaceutically acceptable salt or ester.(phenyl) ketone derivatives of the present invention and pharmaceutical composition thereof can be used for treating cancer, give (phenyl) ketone derivatives of the safe and effective amount of administration.
The compounds of this invention can with other therapeutic agent coupling, as cause apoptosis medicine TNF-a, TRAIL, Fructus Bruceae breast, amycin, tamoxifen, 5-fluorouracil, Tegadifur, harringtonine, cytosine arabinoside, NSC-241240, paclitaxel, flutamide, ifosfamide, doxifluridine, Glass platinum, bend azoles or teniposide etc.; Tumor vessel suppresses medicine angiostatin, endostatin etc.; Immunoregulation medicament polysaccharide-peptide, lentinan etc., the treatment of the disease that can control, alleviate or cure by inhibition hPEBP, the cancers such as such as breast carcinoma.In addition, compound of the present invention also can share with antineoplastic Chinese medicine (or its preparation).
A kind of preferred pharmaceutical composition also contains to cause adjusts die medicine, especially tumor necrosis factor, as huamn tumor necrosis factory alpha etc.
In the time that (phenyl) ketone derivatives or its pharmaceutically acceptable salt or ester are used for the treatment of tumor, it can with one or more pharmaceutically acceptable carrier or mixed with excipients, as solvent, diluent etc., thereby form pharmaceutical composition.
Liquid carrier comprises: sterilized water, Polyethylene Glycol, nonionic surfactant and edible oil (as Semen Maydis oil, Oleum Arachidis hypogaeae semen and Oleum sesami).Solid-state carrier comprises: starch, lactose, calcium hydrogen phosphate, microcrystalline Cellulose, sucrose and kaolin, as long as be applicable to the characteristic of active component and required specific administration mode.In pharmaceutical compositions, normally used adjuvant also can advantageously be included, for example flavoring agent, pigment, antiseptic and antioxidant are as vitamin E, vitamin C, 2,6 ditertiary butyl p cresol (BHT) and butylhydroxy anisole (BHA).
Conventionally, pharmaceutical composition of the present invention comprises following dosage form: oral administered dosage form: as tablet, capsule, dispersible powder, granule or suspension (suspensoid) (containing 0.05-5% suspending agent (cosolvent) according to appointment), syrup (containing 10-50% sugar according to appointment) and elixir (containing having an appointment 20-50% ethanol); Or with sterile injectable solution or suspensoid form (wait ooze medium in containing having an appointment 0.05-5% cosolvent) carry out parenteral canal drug administration.These pharmaceutical preparatioies can contain the approximately 0.001-99.9wt% mixing with carrier conventionally, preferably 0.5-99.5wt%, preferably 2.5-90wt%, the more preferably active component of 5%-60wt% (weight) ((phenyl) ketone derivatives or its pharmaceutically acceptable salt or ester), by the gross weight of compositions.
In the time of pharmaceutical compositions, conventionally, these compounds of the present invention can be formulated in to nontoxic, inertia with pharmaceutically acceptable aqueous carrier medium in, its pH is about 5-8 conventionally, preferably pH is about 6-8, although pH value can change to some extent with being formulated the character of material and disease to be treated.
The pharmaceutical composition preparing can carry out administration by conventional route, comprising (but being not limited to): in tumor, intramuscular, intraperitoneal, intravenous, subcutaneous, Intradermal, oral or topical.Preferably intravenous administration approach.
The present invention's (phenyl) ketone derivatives used also can parenteral or intraperitoneal administration.Also can in the water that is suitably mixed with surfactant (as hydroxypropyl cellulose), prepare solution or the suspension of these reactive compounds (as free alkali or pharmaceutically acceptable salt).Also can in glycerol, liquid, Polyethylene Glycol and the mixture in oil thereof, prepare dispersion liquid.Under conventional storage and service condition, in these preparations, contain antiseptic to prevent growth of microorganism.
The medicament forms that is adapted to injection comprises: aseptic aqueous solution or dispersion liquid and aseptic powder (for prepare aseptic injectable solution or dispersion liquid temporarily).In all situations, these forms must be aseptic and must be that fluid is discharged fluid to be easy to syringe.Under manufacture and condition of storage, must be stable, and must be able to prevent the pollution effect of microorganism (as antibacterial and fungus).Carrier can be solvent or disperse medium, wherein contains just like water, alcohol (as glycerol, propylene glycol and liquid polyethylene glycol), their suitable mixture and vegetable oil.
Using when (phenyl) of the present invention ketone derivatives, also can with other oncotherapy means (as radiotherapy) or other therapeutic agent (as TNF-α etc.) coupling.
The effective dose of active component used can change with the order of severity of the pattern of administration and disease to be treated.But, conventionally when the compounds of this invention every day is with about 0.01-100mg/kg the weight of animals (preferably 0.02-20mg/kg body weight, more preferably 0.1-10mg/kg body weight administration) dosage while giving, can obtain gratifying effect, preferably give with the dosage of 1-4 time every day, or with slow release form administration.For most of large mammal, the accumulated dose of every day is about 5-5000mg or higher, preferably 10-1000mg.Be applicable to dosage form for oral administration, the reactive compound that comprises the approximately 0.5-500mg mixing with solid-state or liquid pharmaceutically acceptable carrier.This dosage of scalable is to provide optimal treatment to reply.For example, by an urgent demand for the treatment of situation, can give the dosage that several times separate every day, or dosage is reduced pari passu.
From being easy to the position of preparation and administration, preferred pharmaceutical composition is fluid composition.The intravenously administrable of (phenyl) ketone derivatives is preferred.
Halth-care composition
Except pharmaceutical compositions is used for the treatment of tumor, in the present invention, also can be by acceptable salt or ester or extract in (phenyl) ketone derivatives or its health care conduct and learning for the preparation of Halth-care composition, thereby for adjuvant therapy of tumors.
In the present invention, acceptable carrier in acceptable salt or ester or extract and health care conduct and learning in (phenyl) ketone derivatives that Halth-care composition contains safe and effective amount (as 0.01-99wt%) or its health care conduct and learning.
Halth-care composition of the present invention can equally with pharmaceutical composition contain acceptable salt or ester or extract in (phenyl) ketone derivatives of same amount or its health care conduct and learning.Conventionally, in Halth-care composition, the content of (phenyl) ketone derivatives can be more lower slightly, for example, containing acceptable salt or ester in 0.01-50wt% (phenyl) ketone derivatives or its health care conduct and learning.
Halth-care composition of the present invention, can make by conventional method the dosage form of any routine, preferably tablet, oral liquid, granule and capsule preparations.
Food additive
Except pharmaceutical compositions is used for the treatment of tumor with as Halth-care composition for beyond adjuvant therapy of tumors, in the present invention, also can be by acceptable salt or ester or extract in (phenyl) ketone derivatives or its bromatology for the preparation of food additive, thereby for adding food, improve anti-tumor capacity the adjuvant therapy of tumors of object.
In the present invention, food additive can contain (phenyl) ketone derivatives or the upper acceptable salt of its bromatology or ester or the extract of safe and effective amount (as 0.01-99wt%), and acceptable carrier in bromatology.
Food additive of the present invention can equally with pharmaceutical composition or Halth-care composition contain acceptable salt or ester or extract in (phenyl) ketone derivatives of same amount or its bromatology.Conventionally, in food additive, the content of (phenyl) ketone derivatives can be lower than the content in health product, for example, containing acceptable salt or ester in 0.01-50wt% (phenyl) ketone derivatives or its bromatology.
In addition, in appropriate circumstances, it is also feasible that acceptable salt or ester or extract in (phenyl) ketone derivatives of the present invention or its bromatology are directly used as food additive, as long as they can not affect taste and/or the outward appearance of food.
Food additive of the present invention, can make by conventional method the form of any routine, such as solution, powder, syrup etc.
Major advantage of the present invention is:
(a) the present invention is directed to the designed micromolecular compound of structure of human hPEBP 4 albumen, can suppress anti-apoptosis molecule---hPEBP4 albumen by efficient targeting, thereby for treatment of cancer;
(b) micromolecular compound of the present invention can also be combined with other medicines and treatment means, for the treatment of malignant tumor.
(c) micromolecular compound of the present invention has good penetrability, and toxic and side effects is little, simple in structure, the advantage such as is easy to synthesize.
Embodiment
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment are only not used in and limit the scope of the invention for the present invention is described.The experimental technique of unreceipted actual conditions in the following example, conventionally according to normal condition, the people such as such as Sambrook, " molecular cloning: laboratory manual " (New York, publishing house of cold spring harbor laboratory, New York:Cold Spring Harbor Laboratory Press, 1989) condition described in, or the condition of advising according to manufacturer.
Unless otherwise indicated, otherwise percentage ratio and umber calculate by weight.Unless otherwise defined, the same meaning that all specialties that use in literary composition and scientific words and one skilled in the art are familiar.In addition, any method similar or impartial to described content and material all can be applicable in the present invention.The use that better implementation method described in literary composition and material only present a demonstration.
Cell line
MCF-7 cell line: ATCC:HTB-22 (purchased from ATCC), this is the breast cancer tumour cell line of a kind of positive expression hPEBP4.
The cultural method of MCF-7 cell line is as follows: cell is inoculated in containing in RPMI1640 (Invitrogen company) culture fluid of 10% calf serum to the CO that to be placed in 37 degrees Celsius, volume fraction be 5%
2cellar culture in incubator, cultivates two weeks without medicine before experiment.
The screening of embodiment 1. micromolecular compounds (DC240016)
Adopt computer assisted virtual screening method to carry out.
First,, by the method for bioinformatics, utilize computer software (described software is Modeller8.0) to set up the 3 d structure model of hPEBP4.Then, to screening purchased from the compound library (approximately 280,000 compounds) of Dutch Specs company, choose best 8700 according to marking with DOCK 4.0 softwares (U.S. Kuntz laboratory).
Continue to use Flex X software (purchased from BioSolveIT company) to dock, according to Flex X, marking is got first 600.Then, use commercially available autodock 3.05 softwares (Scripps university of U.S. Mancur Olson laboratory) to dock take the conformation of Flex X docking as initial conformation.The result of docking analyzes in conjunction with quasi-medicated property (druglikeness) and unnecessary analogue compounds and obviously irrational are removed in perusal, finally chooses 100 compounds.
Wherein, DC240016 micromolecular compound structural formula is suc as formula shown in I, i.e. (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone:
(formula I)
Embodiment 2. micromolecular compounds (DC240016) are tested with the external combination of target hPEBP4 albumen
For the DC240016 selecting (purchased from Dutch Specs company), by based on surface plasmon resonance (Surface Plasmon Resonance, SPR) Biacore 3000 instruments (Amersham company) of technology, the interaction between real-time tracking biomolecule.
Result confirms that DC240016 and hPEBP4 have combination in vitro, Figure 1 shows that the external binding curve of hPEBP4 of DC240016.
Calculate and obtain its dissociation constant R=10e-6 by analysis software.
The growth inhibited of embodiment 3. micromolecular compounds (DC240016) to tumor cell
The present embodiment has adopted conventional mtt assay.Concrete grammar is as follows:
MCF-7 cell is spread to 96 orifice plate overnight incubation with the density of 3000 cells/well.Then, micromolecular compound DC240016 with variable concentrations (1.25,2.5,5,10,25,50 μ M) separately or combine and add in hand-hole with the humanTNF-α of 20ng/ml final concentration.After 44 hours, in every hole, add MTT (5mg/ml) 10 μ l, hatch after 4 hours for 37 degrees Celsius and inhale and abandon supernatant.With after 150 μ l dmso solution purple crystals, be placed in microplate reader (Model of Bio-Rad company 680), detect the absorbance at 570nm place.
Result shows, DC240016 can be in the inhibition of the 5 remarkable enhance TNF-α of μ M final concentration (20ng/ml) on cell proliferation, and its separately application cell is not caused to significant toxic action (Fig. 2).
The detection of the enhancement effect of embodiment 4.DC240016 compound to TNF-α killing tumor cell
Breast cancer cell line MCF-7 cell is taped against in 24 orifice plates with the density in 30000/hole, hatches after 16-24 hour, add DC240016 to final concentration 5 μ M.After 4 hours, then add TNF-α to final concentration 20ng/ml or 50ng/ml.Act on after 48 hours, with detecting with flow cytometer (the Beckton Dickinson FACSCalibur of company) after fluorescent dye Rhodamine 123 (R-123) and iodate the third ingot (PI, Invitrogen company) labelling apoptotic cell.The cell of R-123/PI jack to jack adapter has represented the cell of early apoptosis, and R-123 feminine gender and the cell of the PI positive represent apoptosis and dead cell in late period.
Result is as shown in Figure 3: coupling apoptosis rate (approximately 45%) significance of TNF-α (20ng/ml) and DC240016 (5 μ M) higher than the cell (approximately 14%) with TNF-α separately.In addition, consistent with MTT result: to apply separately the apoptosis (6%) that compound does not cause significant cell.
Compound of the present invention can suppress human phosphotidylethanolabinding binding protein 4 (hPEBP4) by specificity, therefore can be used for treating the tumor of this albumen of high expressed, as breast carcinoma.The result of the above embodiments of the present invention shows, the compounds of this invention can stably suppress hPEBP4, collaborative TNF-α killing tumor cell.
All documents of mentioning in the present invention are all quoted as a reference in this application, are just quoted separately as a reference as each piece of document.In addition should be understood that those skilled in the art can make various changes or modifications the present invention after having read above-mentioned teachings of the present invention, these equivalent form of values fall within the application's appended claims limited range equally.
Claims (4)
1. the purposes by the mixture of compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone or its pharmaceutically acceptable salt and tumor necrosis factor formation, it is characterized in that, for the preparation of suppressing breast cancer cell growth or inducing breast cancer cell to adjust the compositions of dying.
2. purposes as claimed in claim 1, is characterized in that, described breast cancer cell is the breast cancer cell of expressing human phosphotidylethanolabinding binding protein 4.
3. a compositions, is characterized in that, described compositions contains:
(1) compound (3-(2-(2-hydroxyl-5-Nitrobenzol)-5-phenyl-1 hydrogen-imidazol-4 yl) phenyl) (phenyl) ketone or its pharmaceutically acceptable salt;
(2) tumor necrosis factor; And
(3) pharmaceutically acceptable carrier or excipient.
4. compositions as claimed in claim 3, is characterized in that, the dosage form of described compositions is tablet, capsule, powder agent, granule, suspensoid or injection.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101234113A (en) * | 2007-02-01 | 2008-08-06 | 中国人民解放军第二军医大学 | Anti-tumor small molecular compound targeting to phosphatidylethanolamine conjugated protein 4 of human |
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| CA2275610A1 (en) * | 1996-12-16 | 1998-06-25 | Ontogen Corporation | Modulators of proteins with phosphotyrosine recognition units |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101234113A (en) * | 2007-02-01 | 2008-08-06 | 中国人民解放军第二军医大学 | Anti-tumor small molecular compound targeting to phosphatidylethanolamine conjugated protein 4 of human |
Non-Patent Citations (2)
| Title |
|---|
| hPEBP4促进肿瘤细胞ERa介导的转录活化的研究;裘建明;《中国博士学位论文全文数据库 医药卫生科技辑》;20080915(第09期);55-57 * |
| 裘建明.hPEBP4促进肿瘤细胞ERa介导的转录活化的研究.《中国博士学位论文全文数据库 医药卫生科技辑》.2008,(第09期),55-57. |
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