CN114887071B - Spleen-targeting nano delivery carrier - Google Patents
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
- A61K47/542—Carboxylic acids, e.g. a fatty acid or an amino acid
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- A61K39/0005—Vertebrate antigens
- A61K39/0011—Cancer antigens
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- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/51—Nanocapsules; Nanoparticles
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- A61K9/5123—Organic compounds, e.g. fats, sugars
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/51—Nanocapsules; Nanoparticles
- A61K9/5107—Excipients; Inactive ingredients
- A61K9/513—Organic macromolecular compounds; Dendrimers
- A61K9/5146—Organic macromolecular compounds; Dendrimers obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyethylene glycol, polyamines, polyanhydrides
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
- A61K2039/53—DNA (RNA) vaccination
Abstract
The application discloses a spleen-targeted nano delivery carrier, which is an artificially modified lipid nanoparticle, wherein the lipid nanoparticle adopts basic carrier materials including but not limited to: dlin-MC3-DMA, DSPC, chol, DMG-PEG; the delivery vehicle further comprises a functional fatty acid for modifying the lipid nanoparticle, wherein the functional fatty acid has the following biological and/or pharmaceutical properties: spleen targeting; the object body is self-owned; reducing the ability/potential of the metabolic cells of the target body to ingest the delivery vehicle. The application adopts stearic acid to modify nano lipid plasmid, thus improving spleen targeting of nano delivery carrier. The nano delivery carrier provided by the application has good profile, is nearly spherical, has good storage stability, can be suitable for targeted drug delivery systems of drugs (chemical drugs and biological drugs), and has important practical significance in scientific research and medical practicality.
Description
Technical Field
The application relates to the technical field of molecular medicaments and preparation thereof, in particular to a delivery carrier of a targeted medicament and related technologies.
Background
Lipid nanoparticles are generally composed of ionizable lipids, helper lipids, cholesterol, and polyethylene glycol modified lipids, among others. Lipid nanoparticles were primarily used initially for siRNA delivery. Compared with adenovirus-related virus, lentivirus and other virus vectors, lipid nanoparticle and other non-virus vectors have better safety. siRNA drugs using lipid nanoparticles as carriers were approved by the FDA for marketing in 2018, and the administration mode thereof is intravenous drip, which suggests the safety and effectiveness of the lipid nanoparticles as nucleic acid drug delivery carriers. At present, lipid nanoparticles have become the most commonly used mRNA vaccine delivery vectors, and in the development of novel coronavirus vaccines, mRNA vaccines using lipid nanoparticles as vectors have been marketed in batches for the control of novel coronaviruses. Despite the late onset, mRNA lipid nanoparticle vaccines for tumor immunotherapy are rapidly developed and many studies have entered the clinical trial stage. The lipid nanoparticle can protect mRNA from degradation by RNase in and out of a receptor, and improve the delivery of mRNA in vivo and the expression of the mRNA in antigen presenting cells such as DCs and the like. The lipid nanoparticle loaded with mRNA is mainly expressed in liver after intravenous injection, and the insufficient expression of mRNA in immune organs such as spleen limits the immunocompetence of the mRNA tumor vaccine, which suggests that the liposome still needs to be further modified as an mRNA vaccine delivery carrier.
Spleen is the largest peripheral immune organ of the body, has hematopoietic, blood storage and filtration effects, and is also the site for generating immune response upon antigen stimulation. After entering spleen, antigen can be taken up by antigen presenting cell and presented to T cell to induce T cell activation and proliferation to generate sensitized T lymphocyte. The lymph node is used as a peripheral immune organ, has the functions of filtering and removing foreign matters, can treat foreign matter antigens to generate immune response, and is applied to the field of tumor immunotherapy. However, lymph nodes have a low ability to clear cancer cells and are the locus of an immune response against antigens from the lymph fluid. Unlike lymph nodes, the spleen is the site where an immune response is generated against antigens in the blood, and about 90% of circulating blood passes through the spleen, which biological properties make it possible to deliver intravenous tumor antigens to the spleen by using targeted delivery techniques, by inducing a rapid anti-tumor immune response, exerting a highly potent anti-tumor effect.
In recent years, researchers have begun to attempt to bring antigens into the spleen to exert immune anti-tumor effects. There are researches reports that researchers deliver antigen mRNA to spleen to exert the anti-tumor activity of the vaccine by simply adjusting the proportion of the cationic liposome to the mRNA, but no related reports exist later, which are probably that the toxicity of the cationic liposome and the immune anti-tumor effect of the vaccine in clinical experiments are poor. There are also researchers that deliver DNA to B cells in the spleen to exert prophylactic immune anti-tumor activity. In addition, researchers have exerted immune anti-tumor effects by adjusting the particle size of the nano-delivery vehicle to deliver protein/polypeptide antigens to the spleen, although there is some anti-tumor effect, this strategy is not efficient in delivering antigens to the spleen, and the intensity of the cytotoxic T lymphocyte response is not clear.
Disclosure of Invention
The technical problem to be solved by the application is to provide a spleen-targeted nano delivery carrier, which improves the expression of active pharmaceutical ingredients such as mRNA and the like in the spleen by brand-new design and adjustment of lipid nano particles, and safely and efficiently realizes spleen-targeted administration of drugs/nutrients.
In order to solve the technical problems, the technical scheme adopted by the application is as follows.
A spleen-targeting nano-delivery vehicle which is an artificially modified lipid nanoparticle, wherein the lipid nanoparticle adopts basic carrier materials including but not limited to: dlin-MC3-DMA, DSPC, chol, DMG-PEG; the delivery vehicle further comprises a functional fatty acid for modifying the lipid nanoparticle, wherein the functional fatty acid has the following biological and/or pharmaceutical properties: spleen targeting (targeting); target organism self-sex (safety); reducing the ability/potential (efficiency) of the target body to metabolize cells to ingest the delivery vehicle.
As a preferred embodiment of the present application, the functional fatty acid is Stearic Acid (SA) and/or its analogues and derivatives.
As a preferred embodiment of the present application, the delivery vehicle is prepared by precipitation, wherein the molar content of the functional fatty acid is 10% -80%, preferably 60% -80%.
As a preferred embodiment of the present application, the base carrier material used for the delivery vehicle is a combination of Dlin-MC3-DMA, DSPC, chol and DMG-PEG.
As a preferred technical scheme of the application, the delivery carrier is prepared by adopting a precipitation method, wherein the components are in parts by mole: 13-20 parts of Dlin-MC3-DMA, 2.5-4.1 parts of DSPC, 10-15.6 parts of Cholesterol, 0.3-0.7 part of DMG-PEG and 63-70 parts of SA.
As a preferred technical scheme of the application, the delivery carrier is prepared by adopting a precipitation method, wherein the components are in parts by mole: 15-18 parts of Dlin-MC3-DMA, 3.0-3.6 parts of DSPC, 12-13.6 parts of Cholesterol, 0.45-0.55 part of DMG-PEG and 65-68 parts of SA.
As a preferred technical scheme of the application, the delivery carrier is prepared by adopting a precipitation method, wherein the components are in parts by mole: 16.7 parts of Dlin-MC3-DMA, 3.3 parts of DSPC, 12.8 parts of Cholesterol, 0.50 part of DMG-PEG and 66.7 parts of SA.
As a preferred embodiment of the present application, the delivery vehicle is used for delivering a medicament, antigen, immunomodulator, other active ingredient, single ingredient or any combination of ingredients targeting the spleen.
As a preferred embodiment of the application, the delivery vehicle targets dendritic cells within the spleen.
As a preferable embodiment of the present application, the target body is a mammal including a human body.
The beneficial effects of adopting above-mentioned technical scheme to produce lie in:
the spleen targeting nano delivery carrier is prepared by modifying the nano lipid plasmid with stearic acid, so that the spleen targeting of the nano delivery carrier is improved. The functional material fatty acid of the nano delivery carrier is derived from natural products, has low preparation cost and is beneficial to industrial production and application. The nano delivery carrier provided by the application has good profile, is nearly spherical, has good storage stability, can be suitable for targeted drug delivery systems of drugs (chemical drugs and biological drugs), and has important practical significance in scientific research and medical practicality.
Drawings
FIG. 1 is a schematic in vivo effect of spleen targeted drugs.
FIG. 2 is a schematic representation of in vivo spleen targeting results for the nanodelivery vehicle of the application and other control nanodelivery vehicles.
Fig. 3 is a data graph of reduced liver profile of delivery system after stearic acid modification, from which liver profile and expression before and after modification can be directly seen.
FIG. 4 is a graph showing hepatocyte uptake results of the nano-delivery vehicle of the present application and other control nano-delivery vehicles.
Fig. 5 is a graph showing the storage stability test results of the nano-delivery vehicle of the present application.
FIG. 6 is a schematic diagram of the microscopic morphology and particle size and potential detection results of the nano-delivery vehicle of the present application.
Detailed Description
The following examples illustrate the application in detail. The raw materials and the equipment used by the application are conventional commercial products, and can be directly obtained through market purchase.
In the following description of embodiments, for purposes of explanation and not limitation, specific details are set forth, such as particular system architectures, techniques, etc. in order to provide a thorough understanding of the embodiments of the application. It will be apparent, however, to one skilled in the art that the present application may be practiced in other embodiments that depart from these specific details.
It should be understood that the terms "comprises" and/or "comprising," when used in this specification and the appended claims, specify the presence of stated features, integers, steps, operations, elements, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, and/or groups thereof.
It should also be understood that the term "and/or" as used in the present specification and the appended claims refers to any and all possible combinations of one or more of the associated listed items, and includes such combinations.
Furthermore, the terms "first," "second," "third," and the like in the description of the present specification and in the appended claims, are used for distinguishing between descriptions and not necessarily for indicating or implying a relative importance.
Reference in the specification to "one embodiment" or "some embodiments" or the like means that a particular feature, structure, or characteristic described in connection with the embodiment is included in one or more embodiments of the application. Thus, appearances of the phrases "in one embodiment," "in some embodiments," "in other embodiments," and the like in the specification are not necessarily all referring to the same embodiment, but mean "one or more but not all embodiments" unless expressly specified otherwise. The terms "comprising," "including," "having," and variations thereof mean "including but not limited to," unless expressly specified otherwise.
Example 1 overview of the in vivo route of action of spleen-targeted drugs (e.g., mRNA nanovaccines)
Referring to fig. 1, the effect of spleen targeting mRNA nano vaccine in vivo to exert tumor immunotherapy effect is schematically shown. The stearic acid modified lipid nanoparticle mRNA novel nano vaccine delivery system can target and deliver the loaded antigen mRNA to dendritic cells of antigen presenting cells in spleen for specific expression after intravenous injection, and the mRNA novel nano vaccine delivery system stimulates activation of the antigen presenting cells while improving mRNA expression, promotes the antigen presenting cells to present antigen molecules to T cells, proliferates and differentiates the T cells, generates antigen specific effector T cells, kills tumor cells by the effector T cells, and finally plays a role in powerful immunity and anti-tumor.
Example 2 preparation of nanodelivery vehicle
The specific method for preparing the targeted spleen nano delivery vector in the embodiment is as follows: respectively measuring Dlin-MC3-DMA, DSPC, CHO-HP, DMG-PEG2000 or Stearic Acid (SA) stock solutions with corresponding prescription amounts, uniformly mixing, adding absolute ethyl alcohol with corresponding volumes for standby, dissolving mRNA with the prescription amounts in carbonate buffer solution with certain volumes, rapidly and uniformly mixing the ethanol solution with the lipid dissolved and the carbonate buffer solution with the mRNA according to the volume ratio of 1:3, incubating at room temperature for about 30min, and removing the ethyl alcohol by an ultrafiltration method. (the preparation method of the common nano-delivery vehicle is similar to the preparation method of the targeting spleen nano-delivery vehicle described above.)
Firstly, designing nanometer delivery carriers modified by stearic acid with different contents. The amounts of each component in the nano-delivery vehicle are shown in table 1 below.
Table 1 prescription composition of stearic acid modified nano delivery vehicles with different contents
Based on the above study, nanodelivery vehicles modified with different levels of stearic acid were further designed. The amounts of each component in the optimized nanodelivery vehicle are shown in table 2 below.
Table 2 prescription composition of optimized stearic acid modified nano-delivery vehicles
Example 3 spleen targeting of nanodelivery vehicles
In vivo distribution studies were performed using 6-7 week old normal female C57BL6/C mice, 3 mice per group. The LUC-mRNA was used in an amount of 4. Mu.g/mouse by intravenous injection. The mice were free to drink water after injection. 6h after intravenous injection, luciferin substrate. Mu.L was intraperitoneally injected. After 7min, the mice are euthanized, organs such as liver, spleen and lung are separated, and the expression of stearic acid modified nano delivery vectors with different contents in the parts such as liver, spleen and lung after intravenous injection is examined.
The test results are shown in FIG. 2. Figure 2 shows in vivo mRNA delivery behavior of the novel delivery system for stearic acid modified lipid nanoparticle mRNA at varying levels. In the figure, A is a representative picture of the expression of main organs in IVT LUC-mRNA with a formula of A, B, C and D are quantitative statistical results of expression signals of liver, spleen and lung respectively, and E is the proportion of the expression signals of the liver, spleen and lung in the total expression signals. F is a representative picture of the expression of main organs in IVT LUC-mRNA with a formula of B, G, H and I are quantitative statistical results of expression signals of livers, spleens and lungs respectively, and J is the proportion of the expression signals of livers, spleens and lungs in the total expression signals.
From graphs A-E, the results of in vivo targeting studies of the novel delivery system for stearic acid modified lipid nanoparticle mRNA with different contents according to the formula A show that: lipid nanoparticles without fatty acid modification mainly deliver mRNA to liver for expression, and in addition, spleen also has a certain mRNA expression; the addition of fatty acid can change the in vivo mRNA delivery behavior of the lipid nanoparticle, and is characterized in that the addition of fatty acid can reduce the expression of mRNA in the liver and increase the relative enrichment of mRNA in the liver to a certain extent; although the addition of fatty acids did not completely reduce mRNA liver delivery behavior of lipid nanoparticles, the above data suggests that our fatty acid addition may reduce mRNA expression in the liver, and stearic acid modified lipid nanoparticles may have the potential to deliver mRNA to spleen expression, worthy of further investigation.
From figures F-I, the results of in vivo targeting studies of the novel mRNA delivery system for stearic acid modified lipid nanoparticles of formulation B showed: through systematic optimization of stearic acid addition in a lipid nanoparticle mRNA delivery system, the stearic acid modified lipid nanoparticle can remarkably reduce mRNA liver delivery behavior of the lipid nanoparticle, and specifically deliver mRNA to spleen for expression; the above data suggest that our idea of constructing novel delivery systems for spleen-targeted lipid nanoparticle mRNA based on liver fatty acid metabolism strategies is viable.
Lipid nanoparticles without stearic acid modification mainly deliver mRNA to liver for expression, and in addition, spleen has a certain mRNA expression; the addition of stearic acid can change the in vivo mRNA delivery behavior of the lipid nanoparticle, and is particularly characterized in that the addition of fatty acid can reduce the expression of mRNA in the liver and increase the relative enrichment of mRNA in the liver to a certain extent. The in vivo targeting study test results of the delivery system prepared by the preferred embodiment show that: the modified lipid nanoparticle can remarkably reduce the mRNA liver delivery behavior of the lipid nanoparticle when the stearic acid proportion is 66.7%, and can specifically deliver mRNA to spleen for expression, and the delivery system can also realize targeted delivery of mRNA to spleen for expression when the stearic acid proportion is further increased to 75%, so that the expression level of mRNA encoding protein is remarkably reduced.
Example 4 hepatocyte uptake characteristics of nanodelivery vehicles
First, cy 5-labeled spleen-targeted nano delivery vehicles (SLNPs) and Cy 5-labeled normal nano delivery vehicles (LNPs) were prepared by replacing part of cholesterol with Cy 5-labeled cholesterol, intravenous injection was performed at a dose of Cy 5-labeled cholesterol of 20 μg/mouse, euthanasia was performed on the mice after 2 hours, major organs such as liver and spleen of the mice were isolated, photographing was performed using a small animal biopsy imager, and then single cell suspensions were prepared using a milling method. Subsequent correlation detection is performed.
mu.L of single cell suspension (the number of cells per flow tube is about 2X 106) was added to each flow tube, 1. Mu.L of Anti-mouse CD16/32 was added to each flow tube, incubated at 4℃for 10min after homogenization, 1. Mu.L of APC-R700-Anti-mouse CD45, FITC-Anti-mouse-CD3, BV510-Anti-mouse B220, PE-Anti-mouse-CD11B and PercP-Cy5.5-Anti-mouse-CD11c were added to the corresponding flow tubes, and stained for 45min at 4℃in the absence of light, after reaching incubation time, cells were washed 3 times with sterile PBS, resuspended in 0.3mL of sterile PBS for flow detection analysis, and uptake of Cy 5-labeled Cy nanodelivery vehicles (SLNPs) and 5-labeled common nanodelivery vehicles (LNPs) for each cell subset in the spleen were examined.
The specific detection results are shown in figures 3-4. It can be seen that the uptake of stearic acid by hepatic parenchymal cells (CD 45-) is significantly reduced compared to the common delivery systems, in particular, that the cells significantly reduced by the novel delivery system of stearic acid-uptake modified lipid nanoparticle mRNA are mainly non-immune cells (hepatic parenchymal cells, CD 45-) and secondarily B cells and macrophages. Considering that the proportion of non-immune cells in the liver, mainly CD45-, is less than 10%, the stearic acid modified lipid rice delivery system significantly reduces the mediation of parenchymal cells compared with the common lipid nanoparticle mRNA novel delivery system. The details are as follows.
FIG. 3 shows in vivo IVT LUC-mRNA expression and distribution results of Cy 5-labeled stearic acid modified lipid nanoparticle mRNA novel delivery systems loaded with IVT LUC-mRNA. In the figure, A is the expression situation of IVT LUC-mRNA in each organ in the body, B is the statistical result of the expression of IVT LUC-mRNA in each organ in the body, C is the distribution situation of a Cy 5-marked stearic acid modified lipid nanoparticle mRNA novel delivery system in each organ in the body, and D is the statistical result of the distribution situation of the Cy 5-marked stearic acid modified lipid nanoparticle mRNA novel delivery system in each organ in the body.
According to the results of fig. 3: cy 5-labeled stearic acid-modified lipid nanoparticle mRNA novel delivery system capable of specifically delivering IVT LUC-mRNA to spleen expression after intravenous injection into mice; while the distribution results show that the novel delivery system of the Cy 5-marked stearic acid modified lipid nanoparticle mRNA is mainly distributed in the liver, and notably, the distribution of the novel delivery system of the stearic acid modified lipid nanoparticle mRNA in the liver is obviously reduced compared with that of the novel delivery system of the common lipid nanoparticle mRNA, so that the novel delivery system of the stearic acid modified lipid nanoparticle mRNA with the targeting of the spleen dendritic cells is suggested to be feasible to design based on the fatty acid metabolism characteristics of the liver.
FIG. 4 shows the results of flow cytometry detection of Cy 5-labeled stearic acid modified lipid nanoparticle mRNA novel delivery systems and common lipid nanoparticle mRNA delivery systems taken up by liver immune cells (CD45+) and non-immune cells (CD 45-). In the figure, A is a strategy for detecting different cell subsets in the liver by flow cytometry, and B and C are representative flow dot plots and statistical results of a novel delivery system of Cy 5-labeled stearic acid modified lipid nanoparticle mRNA and a delivery system of common lipid nanoparticle mRNA taken up by non-immune cell subsets in the liver respectively. D and E are representative flow plots and statistics, respectively, of a novel delivery system for Cy 5-labeled stearic acid-modified lipid nanoparticle mRNA and a common lipid nanoparticle mRNA delivery system taken up by immune cell subsets in the liver.
According to the results of fig. 4: the novel delivery system of the stearic acid modified lipid nanoparticle mRNA is obviously reduced compared with the novel delivery system of the common lipid nanoparticle mRNA, particularly, cells which are obviously reduced by taking the novel delivery system of the stearic acid modified lipid nanoparticle mRNA are mainly non-immune cells, and the proportion of CD45+ immune cells in the liver is considered to be less than 10 percent, so that the novel delivery system of the stearic acid modified lipid nanoparticle mRNA is obviously reduced compared with the novel delivery system of the common lipid nanoparticle mRNA and is mediated by liver parenchymal cells. The experimental results of fig. 4 directly demonstrate that it is feasible to design a novel delivery system for spleen dendritic cell targeted stearic acid modified lipid nanoparticle mRNA based on liver fatty acid metabolism herein.
Example 5 storage stability of nanodelivery vehicles
The newly prepared spleen-targeted nano delivery vector is divided into a plurality of parts on average, stored at the temperature of 4 ℃, sampled on days 0, 7, 15 and 30 respectively, and the expression activity of the spleen-targeted nano delivery vector is studied by using a small animal living body imager by adopting 6-7 week old normal female C57BL6/C mice.
Each time point was divided into PBS group and spleen-targeted nano-delivery vehicle group, 2 mice per group. After 6h intravenous injection into mice, luciferin substrate μl was intraperitoneally injected. The main organs such as the liver, spleen and lung of the mice are separated, and the storage stability of the mice is studied by examining the expression activity of nano delivery vectors targeting the spleen at different storage time points.
The results are shown in FIG. 5. FIG. 5 shows in vivo expression activity of the novel mRNA delivery system stored at 4 ℃. According to fig. 5, when the novel delivery system for stearic acid modified lipid nanoparticle mRNA with spleen targeting is placed for 30 days at 4 ℃, the spleen targeting does not change at all, and meanwhile, the effect of delivering mRNA to spleen expression is not reduced, so that the novel delivery system for stearic acid modified lipid nanoparticle mRNA has good storage stability at 4 ℃, and the in vivo expression behavior of the novel delivery system for mRNA can be maintained. This result would facilitate in vivo application of the spleen-targeted stearic acid modified lipid nanoparticle mRNA novel delivery system.
EXAMPLE 6 microcosmic morphology of nanodelivery vehicle
Diluting the spleen-targeted nano delivery carrier (B5) colloidal solution, then dripping the diluted solution onto a special copper mesh, standing for about 5min, removing the redundant colloidal solution, dripping 2% (w/v) phosphotungstic acid dye liquor for 5min, removing the redundant dye liquor, standing and drying, and then observing the microscopic morphology of the target nano delivery carrier by using a projection electron microscope and photographing.
The results are shown in FIG. 6-A. Fig. 6A shows the microscopic morphology of the novel delivery system for spleen-targeted mRNA. According to FIG. 6A, the novel mRNA delivery system has a particle size range of about 100nm, is spherical, has no blocking, and has uniform particle size distribution.
Example 7 particle size and potential detection of nanodelivery vehicles
A volume of spleen-targeted nano-delivery vehicle (B5) was measured, diluted to concentration with purified water, and particle size and surface potential of the spleen-targeted nano-delivery vehicle were measured in a laser particle size analyzer.
The results are shown in FIGS. 6-B and 6-C. Figure 6 shows the particle size and potential B for the novel delivery system of spleen-targeted mRNA particle size and C for surface potential. According to FIG. 6, the particle size of the novel mRNA delivery system was about 150nm (B), and the potential was about-10 mV (C).
In the foregoing embodiments, the descriptions of the embodiments are emphasized, and in part, not described or illustrated in any particular embodiment, reference is made to the related descriptions of other embodiments.
The above embodiments are only for illustrating the technical solution of the present application, and not for limiting the same; although the application has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit and scope of the technical solutions of the embodiments of the present application, and are intended to be included in the scope of the present application.
Claims (3)
1. A spleen-targeting nano-delivery vehicle which is an artificially modified lipid nanoparticle, characterized in that:
the base carrier material adopted by the lipid nanoparticle comprises Dlin-MC3-DMA, DSPC, cholesterol, DMG-PEG;
the lipid nanoparticle is prepared by adopting a precipitation method, wherein the mole parts of the components are as follows: 16.7 parts of Dlin-MC3-DMA, 3.3 parts of DSPC, 12.8 parts of Cholesterol, 0.50 part of DMG-PEG and 66.7 parts of stearic acid.
2. The spleen-targeted nano-delivery vehicle of claim 1, wherein: the delivery vehicle is used to deliver a drug, antigen, or combination thereof that targets the spleen.
3. The spleen-targeted nano-delivery vehicle of claim 1, wherein: the delivery vehicle targets dendritic cells within the spleen.
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