CN106822893A - A kind of nano liposomes with microwave effect of enhanced sensitivity and its preparation method and application - Google Patents
A kind of nano liposomes with microwave effect of enhanced sensitivity and its preparation method and application Download PDFInfo
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- A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
- A61K41/0052—Thermotherapy; Hyperthermia; Magnetic induction; Induction heating therapy
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Abstract
The invention discloses a kind of nano liposomes with microwave effect of enhanced sensitivity and its preparation technology.The nano liposomes are a diameter of 30-1000 nanometers, and the double-layer of lipoid thickness of nano liposomes is 4-6 nanometers.After antibody molecule is bonded by Electrostatic Absorption or chemistry on its surface, strengthen its active target-seeking function, make it easier for being enriched with tumor locus, reduce the damage of normal tissue in therapeutic process.Nano liposomes are applied to tumor microwave heat treatment technical field by the present invention first, and the nano liposomes have good microwave temperature rise effect, and particle diameter is than more uniform and controllable.And present invention process is simple, formation efficiency is high, easily realizes large-scale production.Further, since liposome has been applied to clinic, therefore its security is more secure compared with other nano materials, with good clinical practice potentiality.
Description
Technical field
The invention belongs to biomedicine technical field, a kind of nano liposomes with microwave effect of enhanced sensitivity and preparation method thereof and the application in tumor microwave heat treatment are more specifically designed.
Background technology
Cancer has become one of global main lethal cause of disease of threat human health, and its incidence of disease is in the trend for rising year by year.Traditional therapeutic modality of medically conventional cancer has surgery excision, chemotherapy, radiotherapy etc. at present, and these modes all can bring certain pain and toxic and side effect to patient over the course for the treatment of.
Microwave ablation is gradually received by people in recent years as a kind of emerging anti-cancer therapies.Microwave heat therapeutic is that electromagnetic wave energy is converted into heat energy, and tumour cell is killed by heating ablation.It has the lot of advantages such as simple and convenient, noninvasive or minimally invasive, accuracy is high, security is good and price is relatively cheap, with good medical application potential.However, also inevitably normal tissue can cause to damage sometimes over the course for the treatment of.In order to realize efficient, the targeted therapy of tumour, a kind of microwave enhanced sensitivity material that can apply to clinic of researcher research is needed badly.The material needs to meet following condition：1. good biocompatibility, 2. with preferable microwave effect of enhanced sensitivity, 3. can reduce the side effect that microwave heat therapeutic normal tissue is produced in the specific enrichment of tumor locus.
The United States Patent (USP) of Patent No. US005330518A and Patent No. US005861021A discloses application of the micro-wave therapeutic in oncotherapy, and its therapeutic effect is better.But liposome is applied into tumor microwave heat treatment field to have not been reported.The present invention carries the high concentration inorganic salt solution with fine microwave effect of enhanced sensitivity by the use of clinically widely used Liposome nanaparticle as carrier material, obtains a kind of nano liposomes with good microwave effect of enhanced sensitivity.Due to long-term clinical practice, verified liposome nano-carrier has good biological safety, therefore the liposome with good microwave effect of enhanced sensitivity will have wide potential applicability in clinical practice in fields such as treatments of cancer.
The content of the invention
The invention solves the problems that first technical problem be to provide a kind of nano liposomes material with microwave effect of enhanced sensitivity, the nano liposomes material has good biocompatibility, human body is had no toxic side effect, it is used directly in animal body, the effect of microwave heat therapeutic is reached, there is good clinical value in tumor microwave heat treatment field.
The invention solves the problems that second technical problem be to provide a kind of preparation method of the nano liposomes with microwave effect of enhanced sensitivity, nano liposomes pattern prepared by this method is good, size tunable.Preparation method of the present invention is simple and easy to apply, and the gentle formation efficiency of preparation condition is high, easily realizes large-scale production.
The invention solves the problems that the 3rd technical problem be to provide a kind of application scheme of the microwave enhanced sensitivity liposome in tumor microwave heat treatment.
To solve above-mentioned first technical problem, the present invention uses following proposal：
A kind of nano liposomes material with microwave effect of enhanced sensitivity, the nano liposomes material diameter is 30-1000 nanometers, and the shell thickness of the nano liposomes material is 4-6 nanometers；The nano liposomes material core has loaded the high concentration inorganic salt solution with microwave enhanced sensitivity performance.
The nano liposomes material particle size is more uniform and controllable, has excellent dispersiveness in water.
Preferably, described inorganic salts are selected from one or more in sodium chloride, potassium chloride, iron chloride, frerrous chloride, magnesium chloride, calcium chloride, ammonium sulfate, ammonium chloride, ammonium carbonate and ammonium hydrogen carbonate.
To solve above-mentioned second technical problem, the present invention uses following technical proposals：
A kind of preparation method of the nano liposomes with microwave effect of enhanced sensitivity, comprises the following steps：
(1) the salting liquid of various concentrations is prepared, and sonic oscillation is well mixed.
(2) a certain amount of phosphatide and auxiliary material are weighed, and dissolving in organic solvent, in Rotary Evaporators rotary evaporation until organic solvent is spin-dried for, is placed in vacuum drying chamber, is dried overnight afterwards.
(3) the salting liquid for preparing in advance, constant temperature aquation 30min are added.
(4) by above-mentioned solution water bath sonicator 5min, amplitude 50%~100%.
(5), by above-mentioned solution Probe Ultrasonic Searching 1-10min, super 3s stops 3s, amplitude 1%-30%.
(6) the solution that ultrasound is obtained, 30,000rpm centrifugations, 30min, the salt that removing is not wrapped up, this step is in triplicate.
(7) after being centrifuged, supernatant is abandoned, connecting some on nano liposomes surface by way of Electrostatic Absorption or chemistry are bonded afterwards has the antibody of targeting.The completely rear repeat step of question response is (6).
(8) it is last, nano liposomes obtained above are dissolved into normal saline solution, obtain final product desired nano-lipid liquid solution.
Preferably, the (1) described salting liquid of step is selected from one or more in sodium chloride, potassium chloride, iron chloride, frerrous chloride, magnesium chloride, calcium chloride, ammonium sulfate, ammonium chloride, ammonium carbonate and ammonium hydrogen carbonate.
Preferably, the (2) described organic solvent of step is selected from one or more in the polar organic solvent of ethanol, chloroform or other small molecules.
Preferably, the (7) described antibody of step is selected from one or more in folic acid, human epidermal growth factor acceptor (such as HER-2), Cetuximab, other antibody molecules.
To solve above-mentioned 3rd technical problem, the nano liposomes that will be loaded with inorganic salt solution are dispersed in physiological saline, and compound concentration scope is the dispersion liquid of 0~50mg/mL, carries out microwave irradiation treatment in vitro respectively, and its temperature change is measured with optic fiber thermometer；Or the nano liposomes of inorganic salt solution will be loaded be dispersed in physiological saline, compound concentration is the dispersion liquid of 1-20mg/mL, by tail vein injection in tumor-bearing mice body, tumor locus is enteredRow is localMicrowave irradiation, tumor temperature can be rapidly reached more than 43 DEG C, and the temperature is enough to kill tumour cell.
The present invention has advantages below：
1. the present invention is not both significantly with prior art, nano liposomes are applied to tumor microwave heat treatment technical field first in the present invention, and achieve good therapeutic effect, and it has good biocompatibility, organism is had no toxic side effect, compared to for other materials the best advantage is that at present liposome materials be applied to clinic, for solid foundation has been established in the clinical practice of the nano liposomes.
2. the nano liposomes preparation process is simple that prepared by the present invention, preparation condition is gentle, it is not necessary to special installation, and technology used is easy to grasp, size tunable, favorable reproducibility.
3. the core in the nano liposomes that prepared by the present invention is inorganic ion, under the relatively low microwave irradiation of power, inorganic ion can occur function of ion polarization, with the change of microwave field, its direction of motion frequently changes, collided with each other between ion, frictional heat, temperature can be made to be rapidly increased to 43 DEG C in the short period of time, (seeFigure 3).ReachKillThe purpose of cell carcinoma tumor ablation, has good development prospect in clinical practice.
Brief description of the drawings
With reference toAccompanying drawingSpecific embodiment of the invention is described in further detail.
Figure 1It is the transmission electron microscope of the liposome for wrapping up sodium chloride that specific embodiment 1 is obtainedFigurePiece (particle diameter is more homogeneous, mean size about 100-300nm)；
Figure 2It is the particle diameter distribution of the liposome for wrapping up sodium chloride that specific embodiment 1 is obtainedFigure (Particle diameter is more homogeneous, is consistent with the result that transmission electron microscope is obtained).
Figure 3The external microwave temperature rise effect (it can be seen that 16 DEG C are risen above in 5min, and control group only heats up about 10 DEG C, illustrates that the liposome for preparing has external microwave temperature rise effect well) of the liposome of the parcel sodium chloride obtained for specific embodiment 1.
Figure 4The cytotoxicity of the liposome of the parcel sodium chloride obtained for specific embodiment 1 is not (it can be seen that in the case where microwave irradiates, cell survival rate does not have significant change, illustrate that simple nano-particle has good biocompatibility, and in the case of microwave irradiation, with the rising of the concentration of nano-particle, cell survival rate is gradually reduced, and illustrates that nano-particle has the effect of microwave enhanced sensitivity).
Specific embodiment
For the clearer explanation present invention, with reference to preferred embodiment andAccompanying drawingThe present invention is described further.In accompanying drawingSimilar part is with identicalAccompanying drawingMark is indicated.It will be appreciated by those skilled in the art that specifically described content is illustrative and be not restrictive below, should not be limited the scope of the invention with this.
The specific embodiment for being not limited to act set forth below of technical solution of the present invention, can be any combination between various implementation methods.
Specific embodiment 1：
(1) the sodium chloride solution of 10wt% is prepared, and sonic oscillation is well mixed.
(2) it is 80~100 according to DPPC and cholesterol ratio：0~20 weighs membrane material dissolving in organic solvent, in Rotary Evaporators rotary evaporation until organic solvent is spin-dried for, is placed in vacuum drying chamber afterwards, is dried overnight.Wherein phosphatide is DPPC, phosphatidyl glycerol, DLPC, dimyristoyl phosphatidyl choline, lecithin, soybean lecithin, natural cephalin, synthesis cephalin, cuorin, DOPE, DOPC, DPPA, DPPA, DPPG, DSPG, one or more in DSPC；
(3) add the sodium chloride solution for preparing in advance as core, constant temperature aquation 30min, sodium chloride is 10 with the mass ratio of membrane material:1；
(4) by above-mentioned solution water bath sonicator 5min, amplitude 50%~100%；
(5), by ultrasound 3min under above-mentioned solution Probe Ultrasonic Searching condition of ice bath, super 3s stops 3s, amplitude 10%.
(6) the solution that ultrasound is obtained, 30,000rpm centrifugations, 30min, the salt that removing is not wrapped up, this step is in triplicate；
(7) after being centrifuged, supernatant is abandoned, connect folic acid on nano liposomes surface by way of chemistry is bonded afterwards.The completely rear repeat step of question response is (6)；
(8) it is last, nano liposomes obtained above are dissolved into normal saline solution, obtain final product desired nano-lipid liquid solution.
FromFigure 1With 2 in it can be seen that it is manufactured in the present embodiment load sodium chloride solution nano liposomes particle size be 100-300nm,Figure 2It is the transmission electron microscope photo of nano liposomes manufactured in the present embodiment.
The nano liposomes that the present embodiment is prepared are used as microwave sensitizer, and its external and internal microwave intensification experiment is as follows with tumour ablation effect：
The nano liposomes for loading sodium chloride solution are dispersed in physiological saline, microwave irradiation treatment is carried out in vitro, measure its temperature change with optic fiber thermometer, its intensification resultAs schemed 3It is shown, work well.And In vitro cell experiment resultAs schemed 4It is shown, it can be seen that in the case where microwave does not irradiate, cell survival rate does not have significant change, illustrate that simple nano-particle has good biocompatibility, and in the case of microwave irradiation, as the rising of the concentration of nano-particle, cell survival rate are gradually reduced, illustrate that nano-particle has the effect of microwave enhanced sensitivity.The experiment that heated up in animal body is as follows：The nano liposomes that sodium chloride solution will be loaded with are dispersed in physiological saline, by tail vein injection in inoculation tumor-bearing mice body, tumor locus are enteredRow is localMicrowave irradiation, tumor temperature can be rapidly reached more than 43 DEG C, and the temperature can effectively kill tumour cell；General 20 days or so tumours return to one's perfect health and skin recovers normal completely.
Specific embodiment 2~10
With embodiment 1, it be the difference is that only, the sodium chloride solution of addition is replaced with one or more in potassium chloride, iron chloride, frerrous chloride, magnesium chloride, calcium chloride, ammonium sulfate, ammonium chloride, ammonium carbonate and ammonium hydrogen carbonate respectively.Result is similar.
Specific embodiment 11~145
With embodiment 1~10, the difference is that only, the DPPC of step (2) middle addition is replaced with phosphatidyl glycerol, DLPC, dimyristoyl phosphatidyl choline, lecithin, soybean lecithin, natural cephalin, synthesis cephalin, cuorin, DOPE, DOPC, DPPA, DPPA, DPPG, DSPG, one or more in DSPC respectively, as a result similar.
Specific embodiment 146~580
With embodiment 1~145, the difference is that only, the folic acid of step (7) middle addition is replaced with one or more in human epidermal growth factor acceptor (such as HER-2), Cetuximab, other antibody molecules respectively, is as a result similar to.
Obviously; the above embodiment of the present invention is used for the purpose of clearly demonstrating example of the present invention; and it is not the restriction to embodiments of the present invention; for the those of ordinary skill in the field; can also make other changes in different forms on the basis of the above description; here all of implementation method cannot be exhaustive, it is every to belong to obvious change that technical scheme extended or change row still in protection scope of the present invention.
Claims (10)
1. a kind of nano liposomes with microwave effect of enhanced sensitivity, it is characterised in that：The nano liposomes its a diameter of 30-1000 rans, 4-6 nanometers of the nano liposomes shell thickness.
2. nano liposomes according to claim 1, it is characterised in that：The core of the nano liposomes is inorganic salts, and the Shell Materials of the nano liposomes are one or more in phosphatide, cholesterol and antibody.
3. nano liposomes according to claim 2, it is characterised in that：The inorganic salts are selected from one or more in sodium chloride, potassium chloride, iron chloride, frerrous chloride, magnesium chloride, calcium chloride, ammonium sulfate, ammonium chloride, ammonium carbonate and ammonium hydrogen carbonate.
4. nano liposomes according to claim 2, it is characterised in that：The phosphatide is selected from one or more in DPPC, phosphatidyl glycerol, DLPC, dimyristoyl phosphatidyl choline, lecithin, soybean lecithin, natural cephalin, synthesis cephalin, cuorin, DOPE, DOPC, DPPA, DPPA, DPPG, DSPG, DSPC；Antibody is one or more in folic acid, human epidermal growth factor acceptor (such as HER-2), Cetuximab, other antibody molecules；The ratio of phosphatide and cholesterol is 80~100:0~20.
5. the preparation method of a kind of nano liposomes with microwave effect of enhanced sensitivity according to claim any one of 1-4, it is characterised in that what it was realized in：
(1) the salting liquid of various concentrations is prepared, and sonic oscillation is well mixed.
(2) a certain amount of phosphatide and auxiliary material are weighed, and dissolving in organic solvent, in Rotary Evaporators rotary evaporation until organic solvent is spin-dried for, is placed in vacuum drying chamber, is dried overnight afterwards.
(3) the salting liquid for preparing in advance, constant temperature aquation 30min are added.
(4) by above-mentioned solution water bath sonicator 5min, amplitude 50%~100%.
(5), by above-mentioned solution Probe Ultrasonic Searching 1-10min, super 3s stops 3s, amplitude 1%-30%.
(6) the solution that ultrasound is obtained, 30,000rpm centrifugations, 30min, the salt that removing is not wrapped up, this step is in triplicate.
(7) after being centrifuged, supernatant is abandoned, connecting some on nano liposomes surface by way of Electrostatic Absorption or chemistry are bonded afterwards has the antibody of targeting.The completely rear repeat step of question response is (6).
(8) it is last, nano liposomes obtained above are dissolved into normal saline solution, obtain final product desired nano-lipid liquid solution.
6. the preparation method of a kind of nano liposomes with microwave effect of enhanced sensitivity according to claim 5, it is characterised in that：Described organic solvent is one or more in the polar organic solvent of ethanol, chloroform or other small molecules.
7. a kind of preparation method of the nano liposomes with microwave effect of enhanced sensitivity according to claim 5, it is characterised in that：The concentration of the salting liquid is 1%-30%；Described hydration temperature is 40-60 DEG C.
8. the preparation method of a kind of nano liposomes with microwave effect of enhanced sensitivity according to claim 5, it is characterised in that the particle diameter of nano-particle can be adjusted by changing ultrasonic power and ultrasonic time.
9. the preparation method of a kind of nano liposomes with microwave effect of enhanced sensitivity according to claim 5, it is characterized in that reaction condition is gentle, and the targeting of nano liposomes can be increased after antibody molecule in connection, largely increase nano liposomes in the enrichment of tumor locus, reduce the damage of normal tissue in therapeutic process.
10. the application of a kind of nano liposomes with microwave effect of enhanced sensitivity according to claim 1-4, it is characterised in that：The nano liposomes are dispersed in physiological saline；The nano liposomes rise to 43 DEG C -70 DEG C through microwave irradiation 1-10 minutes temperature from 25 DEG C.
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| CN107998391A (en) * | 2017-12-05 | 2018-05-08 | 中国人民解放军总医院 | A kind of microwave enhanced sensitivity composite nanometer particle and its preparation method and application |
| CN108653734A (en) * | 2018-08-28 | 2018-10-16 | 北京化工大学 | A kind of efficient up-conversion nanoparticles photosensitizer compound and the preparation method and application thereof |
| CN110227065A (en) * | 2019-03-22 | 2019-09-13 | 中山大学孙逸仙纪念医院 | Folate-targeted carries NaHCO3Liposome and its application in terms of enhancing immunotherapy of tumors effect |
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| CN110227065A (en) * | 2019-03-22 | 2019-09-13 | 中山大学孙逸仙纪念医院 | Folate-targeted carries NaHCO3Liposome and its application in terms of enhancing immunotherapy of tumors effect |
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