CN113018428A - Preparation method of universal injectable microwave sensitization immune gel - Google Patents
Preparation method of universal injectable microwave sensitization immune gel Download PDFInfo
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- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/39—Medicinal preparations containing antigens or antibodies characterised by the immunostimulating additives, e.g. chemical adjuvants
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- A61B18/18—Surgical instruments, devices or methods for transferring non-mechanical forms of energy to or from the body by applying electromagnetic radiation, e.g. microwaves
- A61B18/1815—Surgical instruments, devices or methods for transferring non-mechanical forms of energy to or from the body by applying electromagnetic radiation, e.g. microwaves using microwaves
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- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
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Abstract
A preparation method of universal injectable microwave sensitization immune gel comprises the following steps: (1) stirring and dissolving sodium alginate powder in water to obtain a solution A; (2) adding an immunostimulant into the solution A obtained in the step (1) and continuously stirring at room temperature to obtain a solution B; (3) slowly dropwise adding CaCl into the solution B obtained in the step (2)2Dropwise adding the aqueous solution while stirring to obtain uniform and stable immune gel; (4) the final product was stored at 4 ℃. Preparation of the ImmunogelThe method has simple steps and mild conditions, can be used after mixing, and is convenient for large-scale commercial production. The obtained gel has good biological safety, injectability, microwave sensitization capability and obvious immune activation capability, can be used for universal microwave ablation-immune combined anti-tumor treatment, and has good application value and clinical transformation prospect.
Description
Technical Field
The invention relates to the field of preparation and application of hydrogel, in particular to a preparation method of immune gel with microwave thermal therapy sensitization, immune activation and high-efficiency anti-tumor treatment effects.
Background
The percutaneous microwave ablation is a local tumor ablation treatment method widely used in clinic, and mainly generates an electromagnetic field through a microwave electrode inserted into a tumor tissue to drive polar molecules in the tumor tissue to continuously rub to generate heat, so that the coagulation necrosis of the tumor tissue is promoted, and the purpose of ablating the tumor is finally achieved. Compared with the traditional surgical treatment, the microwave ablation has the remarkable advantages of minimal invasion, high efficiency, few complications and the like. Nevertheless, there is still a risk of tumor progression and recurrence after microwave ablation in tumor patients. Research shows that the local and distant 3-year recurrence rate of tumor patients after microwave ablation is 10-13% and 27-59% respectively. Therefore, how to effectively inhibit the tumor recurrence and metastasis after microwave ablation becomes a difficult problem to be solved urgently in clinic.
Immunotherapy, a therapeutic approach that utilizes the autoimmune system to kill tumor cells, has shown great potential in preventing tumor recurrence and distant metastasis. In fact, during the microwave ablation process, a large amount of cell debris (containing abundant tumor-associated antigens) is released per se, and various inflammatory cells such as dendritic cells, natural killer cells, macrophages, T lymphocytes and the like are recruited. This provides a natural and excellent opportunity for immune cells of the body to recognize, present tumor antigens and exert tumor vaccine-like functions. However, due to the inherent characteristics of low antigen presentation efficiency, weak immune stimulation signal, and immune tolerance of tumor immune microenvironment, the anti-tumor immune response induced by microwave ablation is far from sufficient to effectively prevent tumor recurrence and distant metastasis. Therefore, the combination of immunotherapy and microwave ablation is expected to solve the troublesome problems of local tumor recurrence and distant metastasis after microwave ablation in clinical work, thereby improving the prognosis of patients.
Currently, various local and systemic immunotherapy strategies are applied for combined anti-tumor therapy after microwave ablation, such as immune checkpoint blockade therapy, cytokine therapy, immune cell therapy, vaccine therapy, and immunoadjuvant therapy. However, each of these current immunotherapeutic strategies faces several challenges, such as the need for elaborate encapsulation carriers, complex cellular engineering, worrying biosafety, and unsatisfactory universality. Taking the most popular immune checkpoint blockade therapy as an example, the therapy has already achieved remarkable clinical therapeutic effects in the treatment of various tumors (such as non-small cell lung cancer, malignant melanoma, etc.) by targeting cell surface molecules such as CTLA-4, PD-1, and PD-L1. Nevertheless, immune checkpoint blockade therapy still suffers from inevitable drawbacks such as effective, severe systemic side effects, tedious gene screening procedures only on a fraction of patients or tumor types, and therefore only a small fraction of patients are eligible for this therapy. Recently, in situ tumor vaccine therapy using tumor itself as antigen source and immune adjuvant has attracted more and more attention, especially in the fields of optical-immune combination therapy and radiotherapy-immune combination therapy. The immune adjuvant is a special immune stimulating compound, can improve the quality of anti-tumor immune response by promoting antigen presentation, amplifying an activation signal and reducing immune tolerance, and has more universal applicability. Since each tumor has a unique set of tumor-associated antigens and tumor neoantigens, immune adjuvant-mediated in situ vaccine therapy can take full advantage of all tumor-associated antigens after microwave ablation without the need to identify and select specific vaccine antigens. Therefore, the immune adjuvant and microwave ablation can effectively eliminate primary tumor, and can fully utilize all released tumor antigens to carry out in-situ immune activation to generate highly specific anti-tumor immune response. However, many immunological adjuvants are hydrophobic molecules (e.g., FDA approved imiquimod R837, etc.), requiring careful construction of the vector to increase its bioavailability. Therefore, the development of a simple, universal and clinically promising delivery strategy for in situ high-dose administration and long-time retention of hydrophobic immunoadjuvants is a critical problem to be solved in the current microwave ablation-immune combined anti-tumor treatment strategy.
Disclosure of Invention
The invention aims to solve the problems of complex cell engineering, need of elaborately designing an encapsulation carrier, limitation of application range and the like in the existing microwave thermotherapy-immunotherapy process, and provides a preparation method of a universal injectable microwave sensitization immune gel.
Sodium alginate is a natural biological macromolecule separated from brown algae, and the sodium alginate molecules are crosslinked in the presence of calcium ions or magnesium ions, so that a porous mesh hydrogel structure with good biocompatibility is quickly formed, and the sodium alginate is widely used in the fields of wound healing, drug delivery, tissue engineering and the like. Due to the abundant ions in the gel and the high-efficiency water locking capacity, the sodium alginate hydrogel has excellent microwave sensitization capacity, is favorable for improving the targeting property of microwave ablation, and reduces the thermal damage to surrounding normal tissues. In addition, due to the unique process of changing the ionic crosslinking into glue, the sodium alginate hydrogel can efficiently load various hydrophilic/hydrophobic drugs, and is very suitable for constructing injectable immune gels loaded with various immunostimulants.
The method for constructing the injectable microwave sensitization immune gel by utilizing the sodium alginate-calcium ion hydrogel to load the immunostimulant has the remarkable advantages of simple steps, low cost, high loading efficiency, biological friendliness, large-scale production and the like. The immune gel prepared by the method can promote the high-efficiency microwave ablation of primary tumors, enhance the specific anti-tumor immunity of organisms after microwave ablation, and obviously inhibit tumor recurrence and distant tumor metastasis. The method provides a simple and universal approach for individual minimally invasive tumor treatment, and has huge clinical transformation potential.
The technical scheme of the invention is as follows:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) stirring sodium alginate powder in water in a container until the sodium alginate powder is fully dissolved, and stirring at room temperature to obtain a sodium alginate solution A with the concentration of 5-20 mg/mL;
2) adding an immunostimulant into the solution A obtained in the step 1) to a final concentration of 0.01-2 mg/mL, and continuously stirring at room temperature to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2The aqueous solution is stirred while being dripped, and uniform and stable immune gel can be obtained after dripping;
4) the resulting immunogels were stored at 4 ℃.
The sodium alginate and CaCl2The concentration ratio of the sodium alginate to the CaCl is adjusted and determined according to the requirement of the immune gel colloid on fluidity in practical application, and the fluidity of the immune gel colloid is determined along with the sodium alginate and the CaCl2The concentration ratio is increased, and the concentration ratio is specifically 10: 0.5-10: 1.5.
The immunostimulant refers to biological molecules or drugs capable of effectively stimulating the immune system of the body, including but not limited to imiquimod (R837), resiquimod (R848), CpG oligodeoxynucleotide (CpG-ODN), Lipopolysaccharide (LPS) and 1-methyltryptophan (1-MT). There are two methods of adding immunostimulants:
The invention has the advantages and effects that:
the preparation method of the immune gel provided by the invention has the remarkable advantages of simple steps, low cost, high loading efficiency, biological friendliness, large-scale production and the like. The preparation process of the immune gel is green and environment-friendly due to the characteristic that sodium alginate gel is changed from solid to gel, namely the immune gel is mixed to be used without additional purification, 100 percent high-efficiency loading of various hydrophilic/hydrophobic immune stimulants is realized, and meanwhile, the large-scale commercial production is facilitated. The injectable immune gel prepared by the method has good microwave sensitization capability and tumor in-situ immune activation capability. Under the microwave stimulation, the immune gel can not only efficiently ablate primary tumor tissues, but also fully utilize tumor-related antigens released after microwave ablation, strongly stimulate an organism to generate specific anti-tumor immunity, and finally effectively inhibit tumor recurrence and distant metastasis. The gel is not limited to specific types of tumors in the using process, has good microwave thermotherapy-immune combination treatment effect on most of solid tumors, and has good universality. In addition, the components involved in the preparation and application of such immunocolloids are FDA approved for use. In a word, the preparation method of the immune gel provided by the invention has simple steps, is green and environment-friendly, is convenient for large-scale production, has high-efficiency microwave thermotherapy-immune combined anti-tumor treatment effect, and has larger development potential and clinical transformation prospect in the tumor treatment field.
Drawings
FIG. 1 is a state diagram of an injectable microwave sensitization immune gel.
FIG. 2 is a scanning electron microscope image of the injectable microwave sensitization immune gel.
FIG. 3 is the microwave temperature rise curve of injectable microwave sensitization immune gel.
FIG. 4 is a diagram of the in vitro immune activation effect of the injectable microwave sensitization immune gel.
Detailed Description
Example 1:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) in a container, stirring 100mg of sodium alginate powder in 5mL of water until the sodium alginate powder is fully dissolved, and stirring at room temperature for 1h to obtain a sodium alginate solution A (20 mg/mL);
2) adding 10mg of immune stimulant imiquimod (R837) powder (the final concentration is 2mg/mL) into the solution A obtained in the step 1), and continuously stirring at room temperature for 1h to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2Aqueous solution (2mg/mL, 5mL, sodium alginate with CaCl2The concentration ratio is 10:1), stirring while dripping, and obtaining uniform and stable immune gel after dripping;
4) the resulting immunogels were stored at 4 ℃.
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, scanning electron microscope observation, microwave temperature rise monitoring and in-vitro immune activation capability evaluation.
As a result: FIG. 1 is a macroscopic picture of an injectable microwave-sensitized immune gel, which is uniform and stable; FIG. 2 is a scanning electron microscope image of an injectable microwave sensitization immune gel, which shows the porous network structure inside the gel; FIG. 3 is a microwave temperature rise curve diagram of injectable microwave sensitization immune gel, under the microwave stimulation, the immune gel is obviously heated, the heating amplitude is obviously higher than that of a pure water solution group, and the excellent microwave sensitization capability is shown; FIG. 4 is a diagram of the in vitro immune activation effect of an injectable microwave-sensitized immune gel, which can effectively stimulate the expression of dendritic cell surface molecules and promote immune activation and antigen presentation.
Example 2:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) in a container, 50mg of sodium alginate powder is stirred in 5mL of water until the sodium alginate powder is fully dissolved, and after stirring for 1h at room temperature, a sodium alginate solution A (10mg/mL) is obtained;
2) adding 10mg of immune stimulant imiquimod (R837) powder (the final concentration is 2mg/mL) into the solution A obtained in the step 1), and continuously stirring at room temperature for 1h to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2Aqueous solution (1mg/mL, 5mL, sodium alginate with CaCl2The concentration ratio is 10:1), stirring while dripping, and obtaining uniform and stable immune gel after dripping;
4) the resulting immunogels were stored at 4 ℃.
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Example 3:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) in a container, stirring 100mg of sodium alginate powder in 5mL of water until the sodium alginate powder is fully dissolved, and stirring at room temperature for 1h to obtain a sodium alginate solution A (20 mg/mL);
2) adding 0.05mg of immune stimulant imiquimod (R837) powder (the final concentration is 0.01mg/mL) into the solution A obtained in the step 1), and continuously stirring at room temperature for 1h to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2Aqueous solution (2mg/mL, 5mL, sodium alginate with CaCl2The concentration ratio is 10:1), stirring while dripping, and obtaining uniform and stable immune gel after dripping;
4) the resulting immunogels were stored at 4 ℃.
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Example 4:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) in a container, stirring 100mg of sodium alginate powder in 5mL of water until the sodium alginate powder is fully dissolved, and stirring at room temperature for 1h to obtain a sodium alginate solution A (20 mg/mL);
2) adding 10mg of immune stimulant imiquimod (R837) powder (the final concentration is 2mg/mL) into the solution A obtained in the step 1), and continuously stirring at room temperature for 1h to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2Aqueous solution (1mg/mL, 5mL, sodium alginate with CaCl2Concentration ratio of 10:0.5), stirring while dripping, and obtaining uniform and stable immune gel after dripping;
4) the resulting immunogels were stored at 4 ℃.
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Example 5:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) in a container, stirring 100mg of sodium alginate powder in 5mL of water until the sodium alginate powder is fully dissolved, and stirring at room temperature for 1h to obtain a sodium alginate solution A (20 mg/mL);
2) adding 10mg of immune stimulant imiquimod (R837) powder (the final concentration is 2mg/mL) into the solution A obtained in the step 1), and continuously stirring at room temperature for 1h to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2Aqueous solution (3mg/mL, 5mL, sodium alginate with CaCl2Concentration ratio of 10:1.5), stirring while dripping, and obtaining uniform and stable immune gel after dripping;
4) the resulting immunogels were stored at 4 ℃.
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Example 6:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) in a container, stirring 100mg of sodium alginate powder in 5mL of water until the sodium alginate powder is fully dissolved, and stirring at room temperature for 1h to obtain a sodium alginate solution A (20 mg/mL);
2) dissolving 10mg of imiquimod (R837) as immunostimulant in CaCl2Aqueous solution (2mg/mL, 5mL, sodium alginate with CaCl2The concentration ratio is 10:1, the final concentration of R837 is 2mg/mL), and the solution is stirred for 1h at room temperature to obtain a solution B;
3) slowly dripping the solution B obtained in the step 2) into the sodium alginate solution A, stirring while dripping, and obtaining uniform and stable immune gel after dripping;
4) the resulting immunogels were stored at 4 ℃.
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Example 7:
a preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) in a container, stirring 100mg of sodium alginate powder in 5mL of water until the sodium alginate powder is fully dissolved, and stirring at room temperature for 1h to obtain a sodium alginate solution A (20 mg/mL);
2) adding 10mg of immunostimulant 1-methyltryptophan (1-MT) powder (final concentration is 2mg/mL) into the solution A obtained in the step 1), and continuously stirring at room temperature for 1h to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2Aqueous solution (2mg/mL, 5mL, sodium alginate with CaCl2The concentration ratio is 10:1), stirring while dripping, and obtaining uniform and stable immune gel after dripping;
4) the resulting immunogels were stored at 4 ℃.
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Example 8:
the preparation process and method of one kind of universal injectable microwave sensitizing immune gel is the same as that in example 1 except that the immunostimulant R837 is re-quimod (R848).
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Example 9:
the preparation process and method of one kind of universal injectable microwave sensitizing immune gel is the same as that in example 1 except that the immunostimulant R837 is changed into CpG oligodeoxynucleotide (CpG-ODN).
The injectable microwave sensitization immune gel prepared in the embodiment is used for carrying out macroscopic observation, observation under a scanning electron microscope, microwave heating monitoring and in-vitro immune activation capability evaluation, and the detection result is similar to that of the example 1.
Claims (5)
1. A preparation method of universal injectable microwave sensitization immune gel comprises the following steps:
1) stirring sodium alginate powder in water in a container until the sodium alginate powder is fully dissolved, and stirring at room temperature to obtain a sodium alginate solution A with the concentration of 5-20 mg/mL;
2) adding an immunostimulant into the solution A obtained in the step 1) to a final concentration of 0.01-2 mg/mL, and continuously stirring at room temperature to obtain a solution B;
3) slowly dripping CaCl into the solution B obtained in the step 2)2The aqueous solution is stirred while being dripped, and uniform and stable immune gel can be obtained after dripping;
4) the resulting immunogels were stored at 4 ℃.
2. The method for preparing the universal injectable microwave sensitization immune gel according to the claim 1, which is characterized in that: the immunostimulant in step 2) refers to a biological molecule or drug capable of effectively stimulating the immune system of the body, including but not limited to imiquimod (R837), resiquimod (R848), CpG oligodeoxynucleotide (CpG-ODN), Lipopolysaccharide (LPS), and 1-methyltryptophan (1-MT).
3. The method for preparing the universal injectable microwave sensitization immune gel according to the claim 1, which is characterized in that: the fluidity of the immune gel colloid can be changed along with the sodium alginate and CaCl2The concentration ratio is increased, and the optimal sodium alginate and CaCl can be determined by adjusting the requirement on the fluidity of the immune gel colloid in practical application2The concentration ratio of (a).
4. The method for preparing the universal injectable microwave sensitization immune gel according to the claim 3, which is characterized in that: the sodium alginate and CaCl2The concentration ratio of (A) to (B) is 10: 0.5-10: 1.5.
5. The method for preparing the universal injectable microwave-sensitized immune gel according to any one of claims 1 to 4, wherein the method comprises the following steps: there are two methods of adding immunostimulants: method 1, directly dispersing immunostimulant in sodium alginate solution A, and then dripping CaCl2The solution forms a gel; method 2, dispersing immunostimulant in CaCl2And then the gel is dropwise added into the sodium alginate solution A to form gel.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113398266A (en) * | 2021-07-06 | 2021-09-17 | 苏州百迈生物医药有限公司 | Microwave ablation conformal agent and application thereof |
| CN114699364A (en) * | 2022-03-29 | 2022-07-05 | 山东大学 | Alginate injectable hydrogel and preparation method and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104069494A (en) * | 2014-07-15 | 2014-10-01 | 中国科学院理化技术研究所 | Sodium alginate microcapsule for tumor microwave hyperthermia technology, and preparation method and application thereof |
| CN111375064A (en) * | 2018-12-29 | 2020-07-07 | 苏州百迈生物医药有限公司 | Pharmaceutical composition for chemotherapy and immunization combination treatment |
-
2021
- 2021-03-15 CN CN202110274020.8A patent/CN113018428A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104069494A (en) * | 2014-07-15 | 2014-10-01 | 中国科学院理化技术研究所 | Sodium alginate microcapsule for tumor microwave hyperthermia technology, and preparation method and application thereof |
| CN111375064A (en) * | 2018-12-29 | 2020-07-07 | 苏州百迈生物医药有限公司 | Pharmaceutical composition for chemotherapy and immunization combination treatment |
Non-Patent Citations (3)
| Title |
|---|
| HAIYAN PAN等: "In Situ Fabrication of Intelligent Photothermal Indocyanine Green−Alginate Hydrogel for Localized Tumor Ablation", 《ACS APPLIED MATERIALS & INTERFACE》 * |
| XIAOHAN等: "In situ thermal ablation of tumors in combination with nano-adjuvant and immune checkpoint blockade to inhibit cancer metastasis and recurrence", 《BIOMATERIALS》 * |
| 高春梅等: "海藻酸钠水凝胶的制备及其在药物释放中的应用", 《化学进展》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113398266A (en) * | 2021-07-06 | 2021-09-17 | 苏州百迈生物医药有限公司 | Microwave ablation conformal agent and application thereof |
| CN114699364A (en) * | 2022-03-29 | 2022-07-05 | 山东大学 | Alginate injectable hydrogel and preparation method and application thereof |
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Application publication date: 20210625 |