CN104353074A - Method for killing tumor cells through combination of gold-mediated near-infrared light heat effect and autophagy inhibitor - Google Patents
Method for killing tumor cells through combination of gold-mediated near-infrared light heat effect and autophagy inhibitor Download PDFInfo
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Abstract
The invention provides a method for killing tumor cells through combination of a gold-mediated near-infrared light heat effect and an autophagy inhibitor. Compared with the prior art, the use quantity of gold nanoparticles can be reduced due to introduction of the autophagy inhibitor, and a god tumor cell killing effect can be obtained by a low dose of laser, so that the injury risk of laser and gold nanoparticles on normal cell tissue is reduced.
Description
Technical field
The present invention relates to the near infrared light heat effect of gold mediation and combining, for killing tumor cells of autophagy inhibitor.
Background technology
Cancer is undergone mutation owing to controlling the related gene of cell cycle, and genetic instability accumulation formed occur in various crowd, a kind of persistent ailment being difficult to cure on age level and different human body organ, the health of the serious threat mankind and life.Also there is no comparatively effective diagnosis and treatment means at present, thoroughly can cure cancer.
The means of current Therapeutic cancer are mainly: surgical operation direct removal of lesions place tumor, radiotherapy, the combined therapy of chemotherapy or above several treatment means.But, adopt operative treatment, be difficult to all tumor tissues to excise, for being difficult to the position of exercising tumor resection, or non-physical tumor, operative treatment will become in a helpless situation hopeless.Chemicotherapy is limited to cannot distinguish tumor cell and normal cell, can produce larger toxic and side effects, finally likely cause the toxicity that cancer patient is larger to normal cell.
Nanotechnology has unique advantage in the diagnosis and treatment of cancer.Diagnosis and treatment nano material has the yardstick similar with nucleic acid, polypeptide and protein and other, nano material is made very easily to enter tissue and cell interior through interstice, by blood capillary, even penetrate blood brain barrier, hide the biological defensive system of body, be convenient to biodegradation or absorption; The larger specific surface area of nano-particle, provides the site of functional group or bioactive molecule construction of GPI-anchored mGM, processedly can be modified into polyfunctional nano-particle; Nanostructured characteristic (as porous, hollow, multilamellar etc.), makes nano-particle have higher medicine and loads with ability, and conveying and the medicament slow release of being convenient to medicine control; Nano grain surface can modify PEG equimolecular, can optionally optionally be assembled at tumor locus by " infiltration-be detained enhancement effect " and is detained, strengthen the treatment effect of tumor.
The tumor photo-thermal treatment of nano material mediation is the nano-particle entering inside tumor cells by selectivity is a kind of method that amount of localized heat kills and wounds cancerous cell by the power conversion of light.This kind of oncotherapy means, can by means of the selective aggregation of high-efficiency low-toxicity nano-particle at tumor locus, and in vitro by non-intervention type laser direct irradiation tumor stove, the photo-thermal realizing tumor kills and wounds.Have toxic and side effects little, the features such as targeted therapy is effective, its relevant clinical front and clinical research is tested in the ascendant.
Near-infrared laser has stronger deep tissues penetration capacity, and in addition, the optical absorption characteristics of gold nano grain near infrared region and the hypotoxicity of himself, make the photo-thermal therapy scheme of gold mediation become one of ideal platform of tumor photo-thermal treatment
Autophagy is a kind of cell self degradation mechanism of cell, and comprise the initial of autophagic vacuole, parcel treats degradation material, the autophagic vacuole finally formed and lysosome fusion, the complete procedure of degraded inclusions.Autophagy is relevant with pathological process to the important physiology of human body
Regulation and control autophagy, can the growth of Tumor suppression, strengthens chemotherapeutics, nano anti-cancer reagent to the fragmentation effect of cancerous cell.Autophagy related gene Becline 1, at the normal expression of cell, can lead oncogenic generation.Cancer occur early stage, the reduction of cell autophagy level, can cause the continuous proliferation of cancerous cell, this illustrate autophagy be suppress cancer.Such as silver nano-grain, itself can as antitumor and anticancer agent, the autophagy effect suppressing silver nano-grain to cause, and can strengthen its fragmentation effect to tumor, reduce the dosage of nanometer formulation, reduce toxic and side effects.The nano-particle such as C60, MnO also by regulating cell autophagy effect, can strengthen chemotherapeutics killing and wounding cancerous cell, reducing the using dosage of chemotherapeutics.
Summary of the invention
The object of the invention is to the photo-thermal effect by combining gold mediation and autophagy inhibitor, the photo-thermal effect improving gold mediation, to the fragmentation effect of cancerous cell, reduces the using dosage of gold nano grain and laser.
First aspect of the present invention provides a kind of nano-particle and autophagy inhibitor for the preparation of based on the medicine of photo-thermal effect killing tumor cells or the purposes of test kit.
In a preferred embodiment, described nano-particle is selected from gold nano grain, poly-dopamine nano-particle, SWCN, Graphene or its combination, preferred gold nano grain, described gold nano grain comprises cage type gold nano grain, spherical gold nano grain, bar-shaped gold nano grain, starlike gold nano grain.
In a preferred embodiment, described tumor is cervical cancer, breast carcinoma or glioma.
In a preferred embodiment, described autophagy inhibitor is selected from 3-MA, Wortmannin (wortmannin), siRNA, shRNA of autophagy related gene or its combination.
In a preferred embodiment, described autophagy related gene comprises Atg5 [NCBI Reference Sequence:NM_058484.5], Becline-1 [mRNA NCBI Reference Sequence:NM_003766.3; MRNA NCBI Reference Sequence:NM_019584.3], Atg7 [mRNA NCBI Reference Sequence:NM_001253717.1; MRNA NCBI Reference Sequence:NM_001136031.2].
Second aspect of the present invention provides the medicine of photo-thermal killing tumor cells or the purposes of test kit of autophagy inhibitor for the preparation of strengthening nanoparticle mediated.
In a preferred embodiment, described nano-particle is selected from gold nano grain, poly-dopamine nano-particle, SWCN, Graphene or its combination, preferred gold nano grain, described gold nano grain comprises cage type gold nano grain, spherical gold nano grain, bar-shaped gold nano grain, starlike gold nano grain.
In a preferred embodiment, described tumor is cervical cancer, breast carcinoma or glioma.
In a preferred embodiment, described autophagy inhibitor is selected from 3-MA, Wortmannin, siRNA, shRNA of autophagy related gene or its combination.
In a preferred embodiment, described autophagy related gene comprises Atg5, Becline-1, Atg7.
The present invention compared with prior art has the following advantages:
1, the introducing of autophagy inhibitor, can reduce the consumption of gold nano grain
2, use and can obtain good tumor cytotoxicity effect compared with the laser of low dosage, thus reduce laser to the risk worked the mischief of normal cell tissue.
Accompanying drawing explanation
Transmission electron microscope (TEM) result of the gold nano grain of Fig. 1 synthesis;
The photothermal deformation curve of the gold nano cage of Fig. 2 synthesis, Au: gold nano grain; PVP: polyvinyl pyrrolidone;
Fig. 3 cage type gold nano grain on the impact of cell viability, Con: blank, Au: gold nano grain; PVP: polyvinyl pyrrolidone;
Fig. 4 shows the photo-thermal effect of gold mediation with the increase of gold nano grain consumption, strengthens the fragmentation effect to cancerous cell, Au: gold nano grain; NIR: near-infrared laser;
The irradiation time that Fig. 5 shows increases laser can increase effect Con: the blank Au: gold nano grain of gold nano grain killing tumor cell; NIR: near-infrared laser;
Fig. 6 gold nano grain, independent near-infrared laser, and near-infrared laser associating gold nano grain process GFP-LC3/HeLa cell is on the impact of autophagy associated protein LC3, wherein A is fluorescence micrograph, B is protein imprinted result and corresponding Image J software analysis result, Con: blank Au: gold nano grain; NIR: near-infrared laser; Wort: wortmannin; 3-MA:3-methyladenine; Fig. 7 is in HeLa cell, and autophagy inhibitor Wortmannin can significantly suppress gold absorption near-infrared laser to cause cell LC3-II excessively to accumulate;
Fig. 8 autophagy inhibitor 3-MA, Wortmannin strengthen the effect that the golden photo-thermal mediated kills and wounds human cervical carcinoma cell HeLa and mouse mastopathy cell 4T1, and Con: blank Au: gold nano grain; NIR: near-infrared laser, Wort: wortmannin, 3-MA:3-methyladenine;
After the expression of the reticent ATG5 of Fig. 9 Atg 5siRNA, the photo-thermal that significantly can strengthen gold mediation kills and wounds the effect of human cervical carcinoma cell HeLa.NIR: near-infrared laser
Detailed description of the invention
Preparation gold nano cage, poly-dopamine nano-particle chemical reagent used is all purchased from sigma; Base SWCN Short-SWCNT (-COOH, 1-2nm, >90%) and Graphene are purchased from Nanjing Xian Feng nanometer; Microtubule related light 3 (LC3) plasmid is received in N.Mizushima (Tokyo, Japan) [Man, N.; Chen, Y.; Zheng, F.; Zhou, W.; Wen, L.P., Induction of genuine autophagy by cationic lipids in mammalian cells.Autophagy 2010,6 (4), 449-54.]; 3-methyladenine (3-MA, 08592), trehalose (Tre, T9531), Hoechst 33342 (B2261), propidium iodide (PI, P4864) and chloroquine (Chloroquine, CQ, C6628) buy from sigma company; Tetrazolium bromide (MTT, TB0799) is the purchase of raw work biology from Shanghai; Wortmannin (Wortmmanin, Wort, s1952) is purchased from green skies biotechnology; Cell culture related reagent is purchased from Invitrogen; LC3 antibody (NB100-2220) is purchased from Novus (Littleton, CO), and green fluorescent protein antibody GFP (sc-101536) is purchased from Santa Cruz Biotechnology; Glyceraldehyde phosphate dehydrogenase (GAPDH) antibody (MAB374) is purchased from Millipore; With two anti-(W4021) of HRP against murine and two anti-(W4011) of anti-rabbit purchased from Promega (Wisconsin, USA); Enhanced chemiluminescence test kit is purchased from Biological Industries (Kibbutz Beit Haemek, Israel); Geneticin/G418 (Gibco, 11811-031) is dissolved in the deionized water of sterilizing, is prepared into the liquid storage of 100mg/mL, and freezing is preserved.
Embodiment 1: the preparation and characterization of gold nano cage
The synthesis preparation of silver nanocubes:
100mL ethylene glycol joins in the round-bottomed flask of 250mL, puts into the oil bath preheating 1 hour of 150 DEG C subsequently, and magnetic agitation.NaHS (NaHS) ethylene glycol solution of 1.2mL, 3mM joins in reaction solution; After 2 minutes, 10mL, 3mM hydrochloric acid (HCl) ethylene glycol solution joins in reaction solution; Add 25mL subsequently, the polyvinylpyrrolidone (PVP, molecular weight ~ 55000) of 20mg/mL; After 2 minutes, the Silver Trifluoroacetate (CF3COOAg) of 8mL, 282mM.Reaction carries out about 20 minutes, by ultraviolet-visible absorption spectroscopy instrument determination reaction solution absorption spectrum at about 440nm (silver nanocubes of corresponding about 40nm), reaction flask is inserted rapidly cessation reaction in ice-water bath.Gained silver nanocubes acetone is washed once, uses deionized water centrifuge washing subsequently 2 times, is suspended in the aqueous solution of 15mL stand-by subsequently.
The preparation of gold nano cube cage:
20mL deionized water is joined in 100mL reaction flask, add 1mL silver nanocubes suspension subsequently, then the chlorauric acid solution of 1mM is added drop-wise in reaction solution with the speed of 1mL/min, until when absorption spectrum Einstein shift is to 800nm, stops dripping gold chloride.To excess chlorination sodium solution be added in reaction solution and spend the night, the silver chloride generated with solubilizing reaction, centrifuge washing 3 times subsequently.
As shown in Figure 1, the gold nano grain of the result display synthesis of transmission electron microscope (transmition electron microscopy, TEM) is cage type, and cube structure, size is 40-60nm.
Embodiment 2: the photothermal deformation curve of gold nano cage
By the DMEM of 400 μ L, PBS, containing 100 μ g/mL polyvinylpyrrolidone (Polyvinyl Pyrrolidone, PVP) 400 μ L PBS and respectively containing 0.5,1,2,2.5, the PBS of 400 μ L systems of the gold nano cage of 3,4,5 μ g/mL, be placed in transparent glass round bottom pipe, bottom laser instrument (Changchun Xin Chenye Photoelectric Technology Co., Ltd.) exposure tube of use MDL-III-808nm-2.5w, output is 1w, now the non-butt coupling optical fiber of laser instrument.And use the temperature of thermocouple the real time measure liquid in pipe.As shown in Figure 2, result shows, gold nano grain has comparatively superior photothermal deformation ability.
Embodiment 3: cell culture and cell viability experiment
Human cervical cancer cell lines HeLa cell and mouse mastopathy cell 4T1 are bought by ATCC and obtain.Microtubule related light 3 (LC3) plasmid is received in N.Mizushima (Tokyo, Japan).Based on HeLa cell [Zhang, the Q. of this GFP-LC3 plasmid construction stable transfection GFP-LC3; Yang, W.J.; Man, N.; Zheng, F.; Shen, Y.Y.; Sun, K.J.; Li, Y.; Wen, L.P., Autophagy-mediated chemosensitization in cancer cells by fullerene C60nanocrystal.Autophagy 2009,5 (8), 1107-1117.] Human cervical cancer cell lines HeLa cell, it is 37 DEG C, CO that the HeLa cell (GFP-LC3/HeLa) of mouse mastopathy cell 4T1 and stably express GFP-LC3 is cultivated in temperature
2concentration is in the incubator of 5%, containing the penicillin/streptomycin of 100units/mL and the amphotericin B of 2.5 μ g/mL in DMEM culture medium (hyclone containing 10%).GFP-LC3/HeLa cell, uses G418 to carry out maintenance growth a period of time before the use.Follow sterile working's principle, in superclean bench, complete cell culture, go down to posterity, the operations such as process.Rifle head, EP pipe, vial etc. used in aseptic experiment operating process test consumptive material all in 121 DEG C, 60 minutes autoclave sterilizations.
The preparation of cell culture phosphate buffer used (PBS): accurately take sodium chloride (NaCl) 8g, potassium dihydrogen phosphate (KH
2pO
4) 0.27g, sodium hydrogen phosphate (Na
2hPO 4) 1.42g, potassium chloride (KCl) the 0.2g Millipore ultra-pure water that adds about 900mL is stirred to dissolving, and pH is adjusted to 7.4 by concentrated hydrochloric acid, and last standardize solution cumulative volume 1L, properly preserves in room temperature after 121 DEG C of autoclavings.
HeLa cell divides in 96 holes, and cell seeding density is 10,000 cells/well, the culture medium system of every hole 100 μ L.After cell attachment 20-24 hour, the gold nano grain of 3.12,6.25,12.5,25,50,100,200 μ g/mL is used to process cell respectively 24 hours.Every hole adds the MTT solution (5mg/mL prepares with the PBS of pH=7.4) of 10 μ L, continues to hatch 4 hours.Carefully suck the medium supernatant in hole, every hole adds the DMSO solution of 150 μ L, after first Can crystallization is fully dissolved, select 570nm wavelength, microplate reader measures the absorbance value of each culture hole, record result also, after removing Background absorbance, draws the growth curve of cell, calculates cell viability.
As shown in Figure 3, cage type gold nano grain has less cytotoxicity, in 100 μ g/mL and following concentration, has no significant effect (cell viability is greater than 80%) the vigor of cell.
Embodiment 4: the photo-thermal of gold nano grain to tumor cell kills and wounds
HeLa is through trypsinization, and DMEM culture medium is resuspended, plants in 24 porocyte culture plates, treats that adherence rate reaches about 90%, carry out follow-up experiment.
Dosage and the irradiation time of near-infrared laser remain unchanged, and add variable concentrations (0,25,50,75,100 μ g/mL) gold nano grain, after hatching 6h altogether with cell, trypsin digestion cell, 1000rpm, 5min centrifugal collecting cell, and cell is resuspended in the DMEM culture medium of 500 μ L, be transferred in 48 orifice plates, carry out laser treatment with irradiation 5min.Then, cell is suitably diluted, plant 96 porocyte culture plates respectively.After 16 hours, mtt assay is used to detect the change of cell viability
The photo-thermal effect that Fig. 4 shows gold mediation along with the increase of the consumption of gold nano grain, can strengthen the fragmentation effect to cancerous cell.Cell viability experimental result shows, and when the concentration of gold nano grain is 50 μ g/mL, can kill and wound the cell of about about 30%.
Keep the dosage of gold nano grain constant, increase the irradiation time of laser.Use 50 μ g/mL gold nano grain pretreatment cells after 6 hours, respectively 5min is irradiated to cell, 7.5min, 10min, detect the fragmentation effect to cancerous cell.Fig. 5 shows, and the irradiation time increasing laser can increase the effect of gold nano grain killing tumor cell.
Embodiment 5: gold nano grain, independent near-infrared laser, and the generation of near-infrared laser associating gold nano grain Induces Autophagy in HeLa and GFP-LC3/HeLa cell
HeLa is through trypsinization, DMEM culture medium is resuspended, plants in 24 porocyte culture plates, treats that adherence rate reaches about 90%, add the gold nano grain of variable concentrations 50 μ g/mL, after hatching 6h altogether with cell, trypsin digestion cell, 1000rpm, 5min centrifugal collecting cell, and cell is resuspended in the DMEM culture medium of 500 μ L, be transferred in 48 orifice plates, carry out laser treatment with irradiation 5min.Then, cell is suitably diluted, plant in 24 porocyte culture plates, every hole 100,000 cell, continue cultivation 16 hours.Trypsinization, collecting cell, carries out protein imprinted experiment, detects the generation of autophagy.
In Fig. 6, GFP-LC3/HeLa is stable transfection and the HeLa cell line of the LC3 albumen of continuous expression GFP label.When autophagy occurs, GFP-LC3-I will be processed to GFP-LC3-II, is gathered on autophagic vacuole, and under fluorescence microscope, we are easy to observe the film bubble spline structure (vesicle diameter is about 300-900nm) with green fluorescence.Through the gold nano grain of 50 μ g/mL, independent near-infrared laser, and near-infrared laser associating gold nano grain process GFP-LC3/HeLa cell, the change that the inner GFP point-like of fluorescence microscope (Olympus IX71) observation of cell is assembled, experimental result as Fig. 6 A shows, and gold nano grain and near-infrared laser coprocessing significantly can strengthen the generation that cell interior point-like is assembled.
Autophagy associated protein LC3 is the marker protein of autophagy, when cell autophagy level is lower, and being distributed in cytosol of LC3 albumen disperse.LC3 albumen is now LC3 protein I type.But when autophagy is induced, the LC3 albumen be distributed in cytosol will be sheared and be processed into membrane-bound LC3-II, is distributed on the film of autophagic vacuole.By the conversion of western blotting technical Analysis LC3 protein I type to II type, we can conclude the generation of autophagy to a certain extent.As the protein imprinted result of Fig. 6 B and the result display of Image J software analysis, gold nano grain and near-infrared laser coprocessing significantly can promote autophagy associated protein LC3 from I type to the conversion of II type.
Embodiment 6: the photo-thermal killing tumor cell of gold mediation and autophagy inhibitor conbined usage
HeLa is through trypsinization, DMEM culture medium is resuspended, plantation is in 24 porocyte culture plates, when adherence rate reaches about 85%, add or do not add autophagy inhibitor 3-MA (3-MA) or wortmannin (Wortmannin), after pretreatment 1h, add 50 μ g/mL gold nano grains, continue process cell 6 hours.Trypsin digestion cell, 1000rpm, 5min centrifugal collecting cell, and cell is resuspended in or without in 500 μ L DMEM culture medium of autophagy inhibitor, is transferred in 48 orifice plates, carries out laser treatment with irradiation 5min.Then, use and have or without the DMEM culture medium that autophagy suppresses, cell is suitably diluted, plant in 96 porocyte culture plates or 24 porocyte culture plates respectively, continue cultivation 16 hours.The two dye of Hochest3342/PI, the ratio of statistics cell death.Wherein, use the step of the cell death situation after Hochest3342/PI two dye detection photo-thermal therapy as follows: emigrated cells culture medium, uses the hochest3342 pigmented cells core of 10 μ g/mL; Dead cell, cell membrane penetration sexually revises, can be painted by the propidium iodide (PI) of 10 μ g/mL, fluorescence microscope (Olympus IX71) is used to take pictures, in the statistics visual field, PI positive cell and the painted cell of hoechst33342, the ratio of the former with the latter is multiplied by 100%, is mortality rate.Trypsinization, collecting cell, carries out Western blot experiment, detects 3-MA and wortmannin to the inhibitory action of autophagy.
As shown in Figure 7, in HeLa cell, autophagy inhibitor Wortmannin can significantly suppress gold absorption near-infrared laser to cause cell LC3-II excessively to accumulate.In GFP-LC3HeLa cell, autophagy inhibitor 3-MA can suppress the accumulation of GFP-LC3II, the release of the free GFP in T suppression cell inside, the level of T suppression cell autophagy.
As shown in Figure 8, autophagy inhibitor 3-MA, Wortmannin itself does not have toxicity to cell, and can pass through T suppression cell autophagy, and the photo-thermal strengthening gold mediation kills and wounds the effect of human cervical carcinoma cell HeLa and mouse mastopathy cell 4T1.
Embodiment 7: the photo-thermal killing tumor cell of gold mediation and the conbined usage of autophagy related gene (Atg5) siRNA
Atg5 siRNAs
(four kinds of Atg5siRNA wait mass mixing thing, its sequence is respectively: Sense 5'-GGA AUA UCC UGC AGA AGA ATT-3'(SEQ ID No:1), Anti-sense 5'-UUC UUC UGC AGG AUA UUC CTT-3'(SEQ ID No:2); Sense 5'-CAU CUG AGC UAC CCG GAU ATT-3'(SEQ ID No:3), Anti-sense 5'-UAU CCG GGU AGC UCA GAU GTT-3'(SEQ ID No:4); Sense 5'-GAC AAG AAG ACA UUA GUG ATT-3'(SEQ ID No:5), Anti-sense 5'-UCA CUA AUG UCU UCU UGU CTT-3'(SEQ ID No:6); Sense 5'-CAA UUG GUU UGC UAU UUG ATT-3'(SEQ ID No:7), Anti-sense 5'-UCA AAU AGC AAA CCA AUU GTT-3'(SEQ ID No:8)), negative control siRNA sequence is Sense 5'-UUC UCC GAA CGU GUC ACG UTT-3'(SEQ ID No:9), Antisense 5'-ACG UGA CAC GUU CGG AGA ATT-3'(SEQ ID No:10)) by the lucky agate biosynthesis in Shanghai.Cell is assigned in 24 orifice plates, use lipo2000 (Invitrogen, 11668-027) transfection negative control siRNA and Atg5siRNAs.After 36 hours, add or do not add 50 μ g/mL gold nano grains, continue process cell 6 hours.Trypsin digestion cell, 1000rpm, 5min centrifugal collecting cell, and cell is resuspended in 500 μ L DMEM culture medium, be transferred in 48 orifice plates, carry out laser treatment with irradiation 5min, or room temperature places 5min.
Then, use DMEM culture medium suitably to be diluted by cell, plant in 96 porocyte culture plates or 24 porocyte culture plates respectively, continue cultivation 16 hours.The situation of cell death before and after the cell death detection method check processing of use embodiment 6.Experimental result is shown in accompanying drawing 9.Gold nano grain and near-infrared laser do not have cytotoxicity to cell, but when both are united can killing tumor cell, Atg5siRNAs silence is fallen after ATG5 expresses, and the photo-thermal that can strengthen gold mediation further kills and wounds the effect of human cervical carcinoma cell HeLa.
Laser instrument model involved in the present invention is MDL-III-808-2.5 (Changchun Xin Chenye Photoelectric Technology Co., Ltd.), coupling core diameter 400 μm, long 1 meter, the optical fiber of SMA905 interface, when laser treatment with irradiation is carried out to cell, laser output power is 2.5W, fiber-optic output distance Tissue Culture Plate bottom 5 centimetres.
Claims (10)
1. nano-particle and autophagy inhibitor are for the preparation of based on the medicine of photo-thermal effect killing tumor cells or the purposes of test kit.
2. purposes according to claim 1, described nano-particle is selected from gold nano grain, poly-dopamine nano-particle, SWCN, Graphene or its combination, preferred gold nano grain, described gold nano grain comprises cage type gold nano grain, spherical gold nano grain, bar-shaped gold nano grain, starlike gold nano grain.
3. purposes according to claim 1, described tumor is cervical cancer, breast carcinoma or glioma.
4. purposes according to claim 1, described autophagy inhibitor is selected from 3-MA, Wortmannin, siRNA, shRNA of autophagy related gene or its combination.
5. purposes according to claim 4, described autophagy related gene comprises Atg5, Becline-1, Atg7.
6. autophagy inhibitor is for the preparation of the medicine of photo-thermal killing tumor cells or the purposes of test kit that strengthen nanoparticle mediated.
7. purposes according to claim 6, described nano-particle is selected from gold nano grain, poly-dopamine nano-particle, SWCN, Graphene or its combination, preferred gold nano grain, described gold nano grain comprises cage type gold nano grain, spherical gold nano grain, bar-shaped gold nano grain, starlike gold nano grain.
8. purposes according to claim 6, described tumor is cervical cancer, breast carcinoma or glioma.
9. purposes according to claim 6, described autophagy inhibitor is selected from 3-MA, Wortmannin, siRNA, shRNA of autophagy related gene or its combination.
10. purposes according to claim 9, described autophagy related gene comprises Atg5, Becline-1, Atg7.
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| CN107050052A (en) * | 2017-06-09 | 2017-08-18 | 中国科学院上海应用物理研究所 | A kind of autophagy blocking system based on nano material and preparation method thereof, and the application in arsenical pharmaceutical therapeutic entity knurl |
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