CN109793726A - Benzethonium chloride is preparing the application in the drug for preventing and treating lung cancer - Google Patents
Benzethonium chloride is preparing the application in the drug for preventing and treating lung cancer Download PDFInfo
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- CN109793726A CN109793726A CN201910189607.1A CN201910189607A CN109793726A CN 109793726 A CN109793726 A CN 109793726A CN 201910189607 A CN201910189607 A CN 201910189607A CN 109793726 A CN109793726 A CN 109793726A
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- benzethonium chloride
- lung cancer
- drug
- gefitinib
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- UREZNYTWGJKWBI-UHFFFAOYSA-M benzethonium chloride Chemical compound [Cl-].C1=CC(C(C)(C)CC(C)(C)C)=CC=C1OCCOCC[N+](C)(C)CC1=CC=CC=C1 UREZNYTWGJKWBI-UHFFFAOYSA-M 0.000 title claims abstract description 52
- 229960001950 benzethonium chloride Drugs 0.000 title claims abstract description 52
- 206010058467 Lung neoplasm malignant Diseases 0.000 title claims abstract description 50
- 239000003814 drug Substances 0.000 title claims abstract description 38
- 201000005202 lung cancer Diseases 0.000 title claims abstract description 35
- 208000020816 lung neoplasm Diseases 0.000 title claims abstract description 35
- 229940079593 drug Drugs 0.000 title claims abstract description 29
- XGALLCVXEZPNRQ-UHFFFAOYSA-N gefitinib Chemical compound C=12C=C(OCCCN3CCOCC3)C(OC)=CC2=NC=NC=1NC1=CC=C(F)C(Cl)=C1 XGALLCVXEZPNRQ-UHFFFAOYSA-N 0.000 claims abstract description 25
- 239000005411 L01XE02 - Gefitinib Substances 0.000 claims abstract description 24
- 229960002584 gefitinib Drugs 0.000 claims abstract description 24
- 201000005296 lung carcinoma Diseases 0.000 claims abstract description 15
- 230000012010 growth Effects 0.000 claims abstract description 11
- 230000035755 proliferation Effects 0.000 claims abstract description 8
- 125000001797 benzyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])* 0.000 claims abstract description 7
- 206010028980 Neoplasm Diseases 0.000 claims description 10
- 230000000118 anti-neoplastic effect Effects 0.000 claims description 6
- 238000002360 preparation method Methods 0.000 claims description 5
- 229940046044 combinations of antineoplastic agent Drugs 0.000 claims description 4
- VCOYRKXQRUGBKS-UHFFFAOYSA-N N.[Cl] Chemical compound N.[Cl] VCOYRKXQRUGBKS-UHFFFAOYSA-N 0.000 abstract description 5
- -1 benzethonium chlorides Chemical class 0.000 abstract description 4
- 230000000694 effects Effects 0.000 abstract description 4
- 238000002474 experimental method Methods 0.000 abstract description 4
- 238000011275 oncology therapy Methods 0.000 abstract description 3
- 229960003872 benzethonium Drugs 0.000 abstract description 2
- 238000001727 in vivo Methods 0.000 abstract description 2
- 239000000890 drug combination Substances 0.000 abstract 1
- 239000002574 poison Substances 0.000 abstract 1
- 231100000614 poison Toxicity 0.000 abstract 1
- 238000011580 nude mouse model Methods 0.000 description 14
- JUJBNYBVVQSIOU-UHFFFAOYSA-M sodium;4-[2-(4-iodophenyl)-3-(4-nitrophenyl)tetrazol-2-ium-5-yl]benzene-1,3-disulfonate Chemical compound [Na+].C1=CC([N+](=O)[O-])=CC=C1N1[N+](C=2C=CC(I)=CC=2)=NC(C=2C(=CC(=CC=2)S([O-])(=O)=O)S([O-])(=O)=O)=N1 JUJBNYBVVQSIOU-UHFFFAOYSA-M 0.000 description 12
- 238000011282 treatment Methods 0.000 description 9
- 241000699660 Mus musculus Species 0.000 description 8
- 230000006907 apoptotic process Effects 0.000 description 8
- 239000001963 growth medium Substances 0.000 description 8
- 238000001514 detection method Methods 0.000 description 7
- 238000000034 method Methods 0.000 description 6
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 5
- 239000003795 chemical substances by application Substances 0.000 description 5
- 238000010790 dilution Methods 0.000 description 5
- 239000012895 dilution Substances 0.000 description 5
- 239000002609 medium Substances 0.000 description 5
- 230000005740 tumor formation Effects 0.000 description 5
- 241000699666 Mus <mouse, genus> Species 0.000 description 4
- 230000002401 inhibitory effect Effects 0.000 description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 3
- 239000002246 antineoplastic agent Substances 0.000 description 3
- 229940041181 antineoplastic drug Drugs 0.000 description 3
- 230000010261 cell growth Effects 0.000 description 3
- 238000010586 diagram Methods 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 238000007920 subcutaneous administration Methods 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 108010040476 FITC-annexin A5 Proteins 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 230000005907 cancer growth Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 238000004043 dyeing Methods 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 230000005764 inhibitory process Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 108010082117 matrigel Proteins 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 238000007747 plating Methods 0.000 description 2
- XJMOSONTPMZWPB-UHFFFAOYSA-M propidium iodide Chemical compound [I-].[I-].C12=CC(N)=CC=C2C2=CC=C(N)C=C2[N+](CCC[N+](C)(CC)CC)=C1C1=CC=CC=C1 XJMOSONTPMZWPB-UHFFFAOYSA-M 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- 230000001225 therapeutic effect Effects 0.000 description 2
- 231100000331 toxic Toxicity 0.000 description 2
- 230000002588 toxic effect Effects 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 206010002091 Anaesthesia Diseases 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 208000010513 Stupor Diseases 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- DHOCGIHFPKXZJB-UHFFFAOYSA-N [Cl+].N[H] Chemical compound [Cl+].N[H] DHOCGIHFPKXZJB-UHFFFAOYSA-N 0.000 description 1
- 230000001464 adherent effect Effects 0.000 description 1
- 239000000853 adhesive Substances 0.000 description 1
- 230000001070 adhesive effect Effects 0.000 description 1
- 230000037005 anaesthesia Effects 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 239000003899 bactericide agent Substances 0.000 description 1
- 239000012148 binding buffer Substances 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
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- 238000002512 chemotherapy Methods 0.000 description 1
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- 239000000825 pharmaceutical preparation Substances 0.000 description 1
- 239000006187 pill Substances 0.000 description 1
- 238000010837 poor prognosis Methods 0.000 description 1
- 230000002980 postoperative effect Effects 0.000 description 1
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- 239000002994 raw material Substances 0.000 description 1
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Landscapes
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses benzethonium chlorides to prepare the application in the drug for preventing and treating lung cancer.The outer experiment in vivo discovery benzethonium chloride of combination can inhibit the growth of lung cancer, proliferation in the present invention, since benzethonium chloride poison has Small side effects, highly-safe, cheap advantage, therefore can be as a kind of drug for treating lung cancer.Furthermore, it has also been found that when benzyl rope chlorine ammonia and Gefitinib are used in combination, the growth of lung carcinoma cell is significantly reduced than exclusive use Gefitinib, illustrate that the addition of benzethonium chloride can increase lung carcinoma cell to the sensibility of Gefitinib, therefore, can be using benzyl rope chlorine ammonia as the sensitizer of Gefitinib, or by benzyl rope chlorine ammonia and Gefitinib drug combination, it is used for lung cancer therapy aspect.
Description
Technical field
The invention belongs to pharmaceutical technology field, in particular to benzethonium chloride is preparing answering in the drug for preventing and treating lung cancer
With.
Background technique
Lung cancer is one of most common malignant tumour, and one of most refractory disease in the world, in recent years disease incidence
Raising trend is presented.Since lung cancer and lung benign disease identify more difficulty, and lung cancer initial symptoms are unobvious, and most of patients is just
The state of an illness has developed to middle and advanced stage when examining, thus often misses its optimal treatment period, seriously affects its therapeutic effect.Lung cancer at present
Treatment means include operative treatment, radiotherapy, chemotherapy, immunization therapy, traditional Chinese medical herbal treatment etc., however cancer is to putting
The tolerance for the treatment of results in patient's postoperative recurrence and poor prognosis, and treatment difficulty increases, and clinically used anticancer drug is past
It is past with certain toxic side effect and expensive.Therefore, the safe and efficient anti-tumor drug for inhibiting lung cancer growth is found to carve
Do not allow to delay.
Benzethonium chloride (Benzethonium chloride) is food and medicine surveillance authority, the U.S. (FDA) approval for first aid
Articles are a kind of disinfection sanitizer and novel preservative, have preferable surface-active, be widely used in daily use chemicals, medicine etc.
Do anticorrosion and bactericidal agent in field.But for lung cancer, it is raw to lung carcinoma cell that there is no pertinent literatures and patent report benzethonium chloride
With inhibiting effect.
Summary of the invention
The primary purpose of the present invention is that the shortcomings that overcoming the prior art and deficiency, provide benzethonium chloride in preparation for preventing
Control the application in the drug of lung cancer.
Another object of the present invention is to provide benzethonium chlorides and Gefitinib in preparing antineoplastic combination drug
Using.
The purpose of the invention is achieved by the following technical solution: benzethonium chloride is in preparing the drug for preventing and treating lung cancer
Using.
The benzethonium chloride achievees the purpose that treat lung cancer by the growth and/or proliferation that inhibit lung cancer.
The effective concentration of the benzethonium chloride is 5~20 μM;Preferably 10~20 μM.
Benzethonium chloride inhibits the application in the growth of lung carcinoma cell and/or the drug of proliferation in preparation.
The effective concentration of the benzethonium chloride is 5~20 μM;Preferably 10~20 μM.
The lung carcinoma cell is preferably A549 cell or H1299 cell.
Benzethonium chloride and Gefitinib are preparing the application in antineoplastic combination drug.
The tumour is preferably lung cancer.
A kind of antineoplastic combined medicament, including benzethonium chloride and Gefitinib.
The effective concentration of benzethonium chloride is 5~20 μM (preferably 5 μM) in the antineoplastic combined medicament, Gefitinib
Effective concentration be 5~20 μM (preferably 5 μM).
Benzethonium chloride increases the application in drug of the lung carcinoma cell to the sensibility of Gefitinib in preparation.
Can also pharmaceutically may be used containing one or more in the drug and antineoplastic combined medicament of the prevention and treatment lung cancer
The carrier or auxiliary material of receiving;The auxiliary material is sustained release agent, excipient, filler, adhesive, wetting agent, disintegrating agent, absorbs rush
Into at least one of agent, surfactant or lubricant.
The drug can be prepared into pharmaceutical preparation using the conventional method of this field;Drug system including oral administration
Agent, such as capsule, pill, tablet, oral solution, granule, tincture.
Application of the benzethonium chloride in the sensitizer for preparing Gefitinib.
The present invention has the following advantages and effects with respect to the prior art:
1, benzethonium chloride is mainly used for disinfection sanitizer, but there is presently no relevant reports for its inhibiting effect to lung cancer
And patent.The outer experiment in vivo of combination of the present invention, the results showed that benzethonium chloride can inhibit the growth of lung cancer, proliferation, benzyl rope chlorine
Ammonium is FDA approval drug, and toxic side effect is small, highly-safe, cheap, benzethonium chloride can be dissolved with water, is administered orally,
Lung cancer therapy is assisted, a kind of new medicament selection can be provided for the clinical treatment of lung cancer, the prospect of development and utilization is good.
2, for Gefitinib as lung cancer fiest-tire medication, generate drug resistance seriously affects therapeutic effect, and in the present invention
It was found that benzyl rope chlorine ammonia can increase lung carcinoma cell to the sensibility of Gefitinib, therefore, benzethonium chloride and Gefitinib can be joined
Medicine is shared, in terms of lung cancer therapy.
Detailed description of the invention
Fig. 1 is influence diagram of the benzethonium chloride to proliferation of lung cancer cells;Wherein, figure A is the benzethonium chloride pair of various concentration
A549 Cell growth ability influences;Scheming the benzethonium chloride that B is various concentration influences the growth ability of H1299 cell.
Fig. 2 is influence diagram of the benzethonium chloride to Increase Apoptosis of Lung Cancer Cells;Wherein, figure A is the Bian rope oronain pair of various concentration
The influence of A549 and H1299 Apoptosis;Scheme the statistics knot that the Bian rope oronain that B is various concentration induces A549 apoptosis
Fruit;Scheme the statistical result that the Bian rope oronain that C is various concentration induces H1299 apoptosis.
Fig. 3 is influence diagram of the benzethonium chloride to the one-tenth knurl ability of lung carcinoma cell in nude mouse;Wherein, figure A is in nude mouse
Subcutaneous tumor;Scheme the variation that B is gross tumor volume size;Scheme the changes of weight that C is nude mice.
Fig. 4 is that benzethonium chloride increases lung carcinoma cell to the result figure of gefitinib-sensitive;Wherein, figure A is different pharmaceutical
A549 Cell growth ability is influenced;Scheming B is that different pharmaceutical influences the growth ability of H1299 cell.
Specific embodiment
Below with reference to embodiment, the present invention is described in further detail, and embodiments of the present invention are not limited thereto.
Unless stated otherwise, the present invention uses reagent, method and apparatus is the art conventional reagents, method and apparatus.Unless
It illustrates, agents useful for same and raw material of the present invention can pass through commercially available acquisition.
Embodiment 1WST-1 cytoactive detection
1. plating cells: collecting A549 and H1299 cell, (purchase is in American Type Culture Collecti (AmericanType
Culture Collection, ATCC) cell bank), cell count is carried out, is laid on 96 orifice plates according to the amount of every 3000 cells in hole
In, overnight incubation.
2. drug-treated: diluting benzethonium chloride to concentration for the treatment of (0,5,10,15,20 μM), so with DMEM complete medium
After replace original culture medium in 96 orifice plates, respectively handle 24,48 and 72h.
3.WST-1 detection: according to the dilution ratio of 1:10 (w/v), with DMEM complete medium dilution WST-1, (WST-1 is thin
Born of the same parents are proliferated detection kit), obtain the culture medium containing WST-1;Then 96 orifice plate Central Plains are replaced with the culture medium containing WST-1
In addition culture medium containing WST-1 is added in the hole without culture cell and is used as blank control, every 100 μ of hole by some culture mediums
L.96 orifice plates are put in again after being incubated for 1h in 37 DEG C of constant incubators, detect (450nm) with microplate reader read plate.Whole process is protected from light behaviour
Make.
4. result: result is as shown in Figure 1A and 1B, shows in figure, shows at the drug of concentration gradient and time gradient
Reason, the growth of lung carcinoma cell receive apparent inhibition, i.e. the benzethonium chloride proliferation that can be used for inhibiting lung carcinoma cell.
2 Apoptosis of embodiment
1. drug-treated: collect A549 and H1299 cell be laid in 6 orifice plates, making cell density is about 40%, respectively plus
Enter the DMEM medium treatment cell containing 0,10,15 and 20 μM of benzethonium chloride.
2. collecting cell: after drug-treated 48h, the cell that apoptosis occurs in supernatant is first collected, after 1100rpm is centrifuged 3min
It abandons supernatant to be collected in same centrifuge tube then 0.25% trypsin digestion of adherent cell, 1100rpm centrifugation
Supernatant is abandoned after 3min.PBS buffer solution cleaning is dyed afterwards twice.
3. dyeing: cell is resuspended with the Binding Buffer (cell apoptosis detection kit) of 500 μ L first, then often
The Annexin V-FITC that 5 μ L are added in pipe adds 5 μ L Propidium Iodide after mixing is mixed, and it is anti-to be then protected from light room temperature
Answer 15min.
4. flow cytometer detection: using C6 flow cytomery after dyeing, before upper machine, sample is filtered off with 400 mesh filter screens
Except particulate matter.Wherein Annexin V-FITC is green fluorescence, selects FL1 Air conduct measurement, and Propidium Iodide is red
Fluorescence selects FL2 FL3 Air conduct measurement.
5. result: result as indicated with 2, Fig. 2 show benzethonium chloride processing after, it will be apparent that induction of the apoptosis of cell.
3 nude mice by subcutaneous tumor formation of embodiment experiment
1. the zoopery of project strictly observes the guideline of animal care mechanism, and obtains Ji'nan University animal
The approval of the Experimental Ethical committee.
2. experimental animal used in project is that (Balb/c-nu is bought big in southern medical courses in general 6~8 week old female nude mices
Learn Experimental Animal Center), totally 15, wherein control group 5 and experimental group 10 construct nude mice by subcutaneous tumor formation model:
(1) cell is collected: being collected cell and is carried out cell count, with every nude mouse 1 × 106The amount of a A549 cell calculates
Total cell number, cell (are bought in Guangzhou Si Er Biotechnology Co., Ltd, coring) after being resuspended with PBS buffer solution with matrigel
1:1 is mixed by volume.
(2) it is inoculated with: mouse being anaesthetized and (by painless and have a pain stimulation and assess anesthesia level, determines that nude mice is in
Narcosis) after, take the above-mentioned and mixed resuspension cell of matrigel to carry out subcutaneously nude mice with 1mL syringe (25G syringe needle)
Injection.
3. cell infusion after a week, is observed the tumor formation situation of nude mouse and measured, when tumour reaches 5 × 5 size
Afterwards, start by be injected intraperitoneally and stomach-filling in a manner of be administered.3 benzethonium chloride concentration gradients are arranged altogether and (first use DMSO for this experiment
Benzethonium chloride mother liquor 50mg/mL is prepared, then is diluted to sterile water using concentration, 100 μ L are administered in every mouse every time), two kinds
Administration mode.Respectively 0mg/kg, 5mg/kg (intraperitoneal injection) and 10mg/kg (stomach-filling), are administered once for every two days by each 5.6.
In administration process, the size of nude mice weight and transplantable tumor was measured in every two days.After two weeks, nude mice is euthanized and by tumour
It takes out.
4. result: result is as shown in Figure 3.Fig. 3 A and 3B show the subendothelial tumor formation experimental result of nude mouse, the growth of tumour
Curve shows obvious inhibition of the subendothelial tumor formation of nude mouse by benzethonium chloride.Show that benzethonium chloride can inhibit the tumour of lung cancer
Growth.And Fig. 3 C is the weight statistical result of three groups of mouse after administration, shows that weight has no significant difference between three groups of mouse,
Prove the safety that uses of the benzethonium chloride as anti-tumor drug.
4 benzyl rope chlorine ammonia of embodiment increases lung carcinoma cell to the sensibility of Gefitinib
1. plating cells: collecting A549 and H1299 cell, carry out cell count, spread according to the amount of 3000, every hole cell
In 96 orifice plates, overnight incubation.
2. drug-treated: then replacing 96 to 5 μM with DMEM complete medium dilution benzethonium chloride and Gefitinib respectively
Original culture medium in orifice plate handles 72h.
3.WST-1 detection: according to the dilution ratio of 1:10 (w/v), with DMEM complete medium dilution WST-1, (WST-1 is thin
Born of the same parents are proliferated detection kit), obtain the culture medium containing WST-1;Then 96 orifice plate Central Plains are replaced with the culture medium containing WST-1
In addition culture medium containing WST-1 is added in the hole without culture cell and is used as blank control, every 100 μ of hole by some culture mediums
L.96 orifice plates are put in again after being incubated for 1h in 37 DEG C of constant incubators, detect (450nm) with microplate reader read plate.Whole process is protected from light behaviour
Make.
4. result: as shown in figure 4, Gefitinib group is used alone compared with group is used in combination, it will be apparent that inhibit lung cancer
The growth of cell illustrates that the addition of benzethonium chloride can increase lung carcinoma cell to the sensitivity of Gefitinib there are significant difference
Property.
The above embodiment is a preferred embodiment of the present invention, but embodiments of the present invention are not by above-described embodiment
Limitation, other any changes, modifications, substitutions, combinations, simplifications made without departing from the spirit and principles of the present invention,
It should be equivalent substitute mode, be included within the scope of the present invention.
Claims (10)
1. benzethonium chloride is preparing the application in the drug for preventing and treating lung cancer.
2. benzethonium chloride according to claim 1 is preparing the application in the drug for preventing and treating lung cancer, it is characterised in that:
The benzethonium chloride achievees the purpose that treat lung cancer by the growth and/or proliferation that inhibit lung cancer.
3. benzethonium chloride according to claim 1 is preparing the application in the drug for preventing and treating lung cancer, it is characterised in that:
The effective concentration of the benzethonium chloride is 5~20 μM.
4. benzethonium chloride inhibits the application in the growth of lung carcinoma cell and/or the drug of proliferation in preparation.
5. benzethonium chloride and Gefitinib are preparing the application in antineoplastic combination drug.
6. benzethonium chloride according to claim 5 and Gefitinib are preparing the application in antineoplastic combination drug,
Be characterized in that: the tumour is lung cancer.
7. a kind of antineoplastic combined medicament, it is characterised in that: including benzethonium chloride and Gefitinib.
8. antineoplastic combined medicament according to claim 7, it is characterised in that: benzyl rope in the antineoplastic combined medicament
The effective concentration of oronain is 5~20 μM, and the effective concentration of Gefitinib is 5~20 μM.
9. benzethonium chloride increases the application in drug of the lung carcinoma cell to the sensibility of Gefitinib in preparation.
10. application of the benzethonium chloride in the sensitizer for preparing Gefitinib.
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WO2008122038A1 (en) * | 2007-04-02 | 2008-10-09 | President And Fellows Of Harvard College | Regulating autophagy |
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