Application of viscous gel in preparation of cryotherapy patch

Abstract

The invention relates to the technical field of cryotherapy, in particular to application of viscous gel in preparation of a cryotherapy patch. The invention provides an application of viscous gel in preparation of a cryotherapy patch. The viscous gel is a crystal phase in an environment of-196 DEG C or below and is a soft gel phase in an environment of human body temperature. The adhesive gel is adopted for preparing the cryotherapy patch, accurate positioning can be guaranteed, accurate control over the freezing range can be achieved, complication risks caused by gas expansion in the liquid nitrogen gasification process are avoided, due to the degradable characteristic of the adhesive gel, the adhesive gel can be naturally absorbed by tissue, and therefore the effect that the adhesive gel is not prone to damage is achieved. And in the process, a protective gel barrier is provided for a frozen wound surface range, and separation from tissues is not needed, so that secondary injury in the treatment process is reduced.

Description

Technical Field

[0001] The present invention relates to the technical field of cryotherapy, and in particular to the application of viscous gel in the preparation of cryotherapy patches. Background Art

[0002] Cryotherapy can destroy and ablate diseased tissues through pathological changes during the freezing process. Freezing causes ice crystals to form in tissues and cells, which undergo dehydration and protein denaturation, leading to damage to the tissues at the site of disease and necrosis and shedding of cells, thereby performing treatment. This therapy has been widely used in the fields of skin lesions (warts, granulomas), airway lesions (amphistorical stenosis), and cervical lesions (intraepithelial neoplasia). The current mainstream freezing technologies are: liquid nitrogen freezing and cryoprobe freezing. Liquid nitrogen freezing is to drop liquid nitrogen on the lesion for freezing, while metal cryoprobe freezing uses a freezing working gas (usually carbon dioxide CO 2 Or nitric oxide (NO) vaporization absorbs heat, causing the probe to quickly cool to -89°C, and then destroy the tissue through direct contact and repeated freezing and thawing. Although these methods have certain therapeutic effects in various clinical scenarios (such as skin warts, benign airway stenosis, and cervical intraepithelial neoplasia), they also have some obvious disadvantages: 1) The freezing range is difficult to accurately control; 2) The freezing device may adhere to the tissue, posing a risk of secondary injury or complications; 3) There is a lack of postoperative wound protection, and there is a risk of infection and poor healing. Summary of the invention

[0003] In view of this, the purpose of the present invention is to provide an application of a viscous gel in the preparation of a cryotherapy patch. The present invention uses a viscous gel to prepare a cryotherapy patch to ensure accurate positioning, to achieve precise control of the freezing range, to avoid the risk of complications caused by gas expansion during the liquid nitrogen gasification process, and due to the degradable properties of the viscous gel, it can be naturally absorbed by the tissue without separation from the tissue, thereby reducing secondary damage during the treatment process.

[0004] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0005] The present invention provides an application of a viscous gel in preparing a cryotherapy patch. The viscous gel is in a crystalline phase in an environment below -196°C and in a soft gel phase in an environment at human body temperature.

[0006] Preferably, the mass percentage of the viscous gel in the cryotherapy patch is 80% to 100%.

[0007] Preferably, the cryotherapy patch further comprises an adjuvant;

[0008] The mass percentage of the auxiliary agent in the cryotherapy patch is 1-20%.

[0009] Preferably, the auxiliary agent includes one or more of antibacterial drugs, anti-fibrosis drugs and drugs promoting epithelial regeneration.

[0010] Preferably, the antibacterial drugs include one or more of β-lactam antibiotics, aminoglycoside antibiotics, macrolide antibiotics, tetracycline antibiotics and fluoroquinolone antibiotics;

[0011] The anti-fibrosis drugs include the small molecule inhibitor RepSox and / or the drug pirfenidone;

[0012] The drug for promoting epithelial regeneration includes the small molecule inhibitor Y27632.

[0013] Preferably, the viscous gel comprises one or more of polylipoic acid hydrogel, acrylic acid / vinyl pyrrolidone copolymer succinimide ester hydrogel, lipoic acid / polyethylene glycol copolymer succinimide ester hydrogel, polyaspartic acid hydrogel, polysebacic acid glyceryl hydrogel and linear paste-like polysebacic acid glyceryl gel.

[0014] Preferably, the viscous gel comprises a linear paste-like polyglyceryl sebacate gel.

[0015] Preferably, the method for preparing the linear paste-like polyglyceryl sebacate gel comprises the following steps:

[0016] Glycerol and sebacic acid are mixed, and a first polymerization reaction is carried out under nitrogen filling conditions, and then a second polymerization reaction is carried out under vacuum conditions to obtain the linear paste-like polysebacic acid glyceride gel.

[0017] Preferably, the molar ratio of glycerol to sebacic acid is 1:(1-3).

[0018] Preferably, the temperature of the first polymerization reaction is 120-180°C and the time is 4-24h;

[0019] The temperature of the second polymerization reaction is 120-180° C. and the time is 24-48 hours.

[0020] The present invention provides an application of a viscous gel in preparing a cryotherapy patch. The viscous gel is in a crystalline phase in an environment below -196°C and in a soft gel phase in an environment at human body temperature. The present invention utilizes the property that the viscous gel is in a crystalline phase in an environment below -196°C and in a soft gel phase in an environment at human body temperature, and uses the viscous gel as a freezing medium to construct a cryotherapy patch (a cryotherapy patch that can be reversibly transformed in a temperature box). First, the constructed cryotherapy patch is placed in liquid nitrogen for freezing to achieve an ultra-low temperature phase transition and form a crystalline phase gel; after the crystalline phase gel is brought into contact with the surface of the diseased tissue, the gel achieves rapid freezing therapy by absorbing heat, and can form preliminary adhesion on the contact surface by crystallization; then, after rapid heat absorption, the viscous gel quickly softens and changes phase, transforms into a soft gel phase, and produces a strong adhesion barrier. The viscous gel can provide strong adhesion in a moist environment, form a protective barrier on the wound surface, help isolate the external environment, reduce the risk of infection to reduce secondary damage, and can also provide a strong adhesive surface by applying to the tissue surface to promote regeneration and accelerate wound repair (schematic diagram as shown in FIG. Figure 1 As shown), this protective effect not only improves the therapeutic effect, but also reduces the difficulty of postoperative management and improves the patient's postoperative quality of life. At the same time, the viscous gel of the present invention is applied to the cryotherapy patch and can be accurately positioned at the lesion site in the form of a biological patch, thereby providing a stable cold source. This method can not only achieve precise control of the freezing range and avoid the risk of complications caused by gas expansion during the liquid nitrogen gasification process, but also because of the degradable properties of the gel, it can be naturally absorbed by the tissue without separation from the tissue, thereby reducing secondary damage during the treatment process. Therefore, the above application is expected to improve the clinical efficacy of cryotherapy, especially in reducing complications and improving the therapeutic effect. It shows great potential, and can also be customized to adapt to different types and depths of lesions, so as to achieve more personalized treatment. At the same time, due to the biocompatibility and biodegradability of the viscous gel, the viscous gel that meets the above conditions becomes an ideal freezing medium, which can effectively transfer cold energy to the target area without causing additional damage, thereby achieving cryotherapy of lesions. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 This is a schematic diagram of the working principle of the cryotherapy patch of the present invention;

[0022] Figure 2 This is a physical picture of the cryo-microneedle patch described in Example 1 applied to the inner surface of the trachea for stretching, squeezing, twisting and water flushing experiments;

[0023] Figure 3 This is a physical picture of the in vitro infrared monitoring of the cryo-microneedle patch described in Example 1;

[0024] Figure 4 The phase change of the cryo-microneedle patch described in Example 1 in a rabbit body;

[0025] Figure 5 The therapeutic effect of the cryomicroneedle patch described in Example 1 on the injured part in the animal model of recurrent tracheal stenosis;

[0026] Figure 6 The gross appearance and pathological examination results of the tracheal stenosis site after 21 days of treatment with the cryomicroneedle patch described in Example 1;

[0027] Figure 7 This is a comparison chart of the therapeutic effects of the room temperature microneedle patch described in Comparative Example 1 (denoted as MN), the frozen microneedle patch described in Example 1 (denoted as Cryo-MN), and the frozen microneedle patch described in Example 2 (denoted as Cryo-RY@MN);

[0028] Figure 8 This is the degradation curve of PGS described in Example 1. DETAILED DESCRIPTION

[0029] The present invention provides an application of a viscous gel in preparing a cryotherapy patch. The viscous gel is in a crystalline phase in an environment below -196°C and in a soft gel phase in an environment at human body temperature.

[0030] The present invention does not have any special limitation on the form of the cryotherapy patch, and can adopt a form well known to those skilled in the art (such as a single-layer sheet, a multi-layer sheet, a microneedle or a sphere, etc.) according to the site of action. In an embodiment of the present invention, the cryotherapy patch can be in the form of a cryomicroneedle patch.

[0031] In the present invention, the adhesion strength of the adhesive gel is preferably >5 kPa. In the present invention, the adhesion mechanism of the adhesive gel preferably includes physical crosslinking through mechanical interlocking, electrostatic action, hydrogen bonding, ionic action, chemical crosslinking through chemical bonds or dynamic covalent crosslinking and physical and chemical synergistic adhesion mechanisms.

[0032] In the present invention, the mass percentage of the adhesive gel in the cryotherapy patch is preferably 80% to 100%, more preferably 95%.

[0033] In the present invention, the viscous gel preferably includes one or more of polylipoic acid hydrogel, acrylic acid / vinyl pyrrolidone copolymer succinimide ester hydrogel, lipoic acid / polyethylene glycol copolymer succinimide ester hydrogel, polyaspartic acid hydrogel, polysebacic acid glyceryl hydrogel and linear paste polysebacic acid glyceryl gel; more preferably includes linear paste polysebacic acid glyceryl gel (PGS); when the viscous gel is two or more of the above specific selections, the present invention does not have any special restrictions on the ratio of the above specific substances, and can be mixed in any ratio. In an embodiment of the present invention, the viscous gel can be a linear paste polysebacic acid glyceryl gel.

[0034] In the present invention, the preparation method of the linear paste-like polyglyceryl sebacate gel comprises the following steps:

[0035] Glycerol and sebacic acid are mixed, and a first polymerization reaction is carried out under nitrogen filling conditions, and then a second polymerization reaction is carried out under vacuum conditions to obtain the linear paste-like polysebacic acid glyceride gel.

[0036] In the present invention, the molar ratio of glycerol to sebacic acid is preferably 1:(1-1.5), more preferably 1:1.5. In an embodiment of the present invention, the molar ratio of glycerol to sebacic acid may be 1:1.5.

[0037] In the present invention, the nitrogen filling method is preferably to use a nitrogen blowing needle to blow nitrogen into the mixed system obtained after the mixing.

[0038] In the present invention, the temperature of the first polymerization reaction is preferably 120-140°C, more preferably 130-160°C; the time is preferably 4-24h, more preferably 10-20h; the temperature of the second polymerization reaction is preferably 120-180°C, more preferably 130-160°C; the time is preferably 12-48h, more preferably 30-40h. In an embodiment of the present invention, the temperature of the first polymerization reaction can be 140°C and the time can be 12h; the temperature of the second polymerization reaction can be 140°C and the time can be 12h.

[0039] In the present invention, the first polymerization reaction and the second polymerization reaction are preferably carried out under stirring conditions. The present invention does not have any special limitation on the stirring process, and the stirring process may be carried out using a process well known to those skilled in the art.

[0040] According to different clinical application requirements (such as biocompatibility, mechanical strength, environmental responsiveness or antibacterial properties, etc.), the present invention preferably adds corresponding additives to the viscous gel to meet the above requirements.

[0041] In the present invention, the cryotherapy patch also preferably includes an auxiliary agent; the mass percentage of the auxiliary agent in the cryotherapy patch is preferably 1-10%, more preferably 4-6%. In the present invention, the auxiliary agent preferably includes one or more of antibacterial drugs, anti-fibrosis drugs and drugs for promoting epithelial regeneration; the antibacterial drugs preferably include one or more of β-lactam antibiotics, aminoglycoside antibiotics, macrolide antibiotics, tetracycline antibiotics and fluoroquinolone antibiotics; the antifibrosis drug preferably includes the small molecule inhibitor RepSox and / or the drug pirfenidone; the drug for promoting epithelial regeneration preferably includes the small molecule inhibitor Y27632; when the auxiliary agent is two or more of the above-mentioned specific options, the present invention does not have any special restrictions on the ratio of the above-mentioned specific substances, and they can be mixed in any ratio.

[0042] In the present invention, the auxiliary agent can be sustained-released for a long time with the aid of the gel with a three-dimensional network structure in the cryotherapy patch, and the sustained-release time can be as long as 14 days.

[0043] In the present invention, the viscous gel also preferably includes one or more of natural hydrogel, polyethylene glycol hydrogel, and environmentally responsive hydrogel; the natural hydrogel preferably includes one or more of chitosan hydrogel, chitin hydrogel, collagen hydrogel and gelatin hydrogel; the environmentally responsive hydrogel preferably includes one or more of temperature-responsive PGS hydrogel, temperature-responsive isoacrylamide PNIPAM hydrogel, temperature-responsive poloxamer 407 hydrogel, pH-responsive chitosan hydrogel, glucose-responsive phenylboronic acid hydrogel, ionic strength-responsive sodium alginate hydrogel, ROS-responsive hydrogel and MMP-responsive hydrogel; when the auxiliary agent is two or more of the above-mentioned specific selections, the present invention has no special limitation on the ratio of the above-mentioned specific substances, and they can be mixed in any ratio.

[0044] The technical solutions in the present invention will be described clearly and completely below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0045] Example 1

[0046] Glycerol and sebacic acid were added to a reactor at a molar ratio of 1:1.5. The mixture was heated to 140°C and a nitrogen-bubbling needle was inserted into the obtained mixed system. After continuous stirring for 12 hours, the nitrogen was removed and the reactor was continuously evacuated using a vacuum pump. The reaction was continued for 12 hours to obtain a linear paste of polyglyceryl sebacate (PGS). The degradation curve of the PGS is shown in FIG. Figure 8 As shown by Figure 8 It can be seen that PGS can be degraded slowly and evenly in a simulated liquid environment in vitro;

[0047] The linear paste polysebacic acid glycerol is sequentially subjected to thermoplastic injection molding and low-temperature molding treatment to obtain a frozen microneedle patch (including a frozen microneedle patch with a patch thickness of 1 mm, a size of 1 cm×1 cm, and a microneedle needle length of 300 μm (for rabbit experiments) and a frozen microneedle patch with a patch thickness of 1 mm and a microneedle needle length of 600 μm (for sheep experiments));

[0048] At room temperature, the cryo-microneedle patch (size 1 cm×1 cm) was applied to the inner surface of the trachea and subjected to stretching, squeezing, twisting and water flushing experiments. Figure 2 This is a physical picture of the cryo-microneedle patch applied to the inner surface of the trachea for stretching, squeezing, twisting and water flushing experiments, by Figure 2 It can be seen that after various tests, the cryo-microneedle patch can still remain adhered to the inner surface of the trachea and can maintain adhesion in a complex environment;

[0049] The frozen microneedle patch is placed in liquid nitrogen (-196°C) to obtain a frozen microneedle patch in a crystalline phase. When used, the frozen microneedle patch in the crystalline phase is applied to the lesion site. When the frozen microneedle patch contacts the tissue, the contact surface is rapidly cooled, and water molecules and the like are rapidly crystallized, forming a simple primary adhesion. However, this simple primary adhesion cannot meet the needs of long-term treatment, so the frozen microneedle patch needs to be quickly converted into a highly adhesive soft gel state. To this end, the frozen microneedle patch in the crystalline phase is placed in an in vitro constant temperature environment of 37°C, and its temperature-time changes are detected using infrared technology. Among them, Figure 3 This is a physical picture of the in vitro infrared monitoring of the cryo-microneedle patch. Figure 3 It can be seen that in about 2 minutes, the frozen microneedle patch can reach the ambient temperature of 37°C, return to the soft gel phase, and complete the rapid phase transition. At the same time, this also shows that the soft gel can still return to the soft gel state after being placed in ultra-low temperature (-196°C), proving the reversibility of the phase transition of the frozen microneedle patch;

[0050] A frozen microneedle patch (1 cm × 1 cm) with a patch thickness of 1 mm and a microneedle length of 300 μm was placed in liquid nitrogen (-196°C) to obtain a frozen microneedle patch in the crystalline phase. The frozen microneedle patch in the crystalline phase was then applied into the rabbit trachea, and its phase change was detected again in vivo using infrared imaging. Figure 4 is the phase change of the cryo-microneedle patch in the rabbit body, Figure 4It can be seen that after the cryo-microneedle patch is placed in the rabbit body, its temperature can be observed to be extremely low, and in vivo, its temperature can be clearly observed to change rapidly, reaching 37°C within 1 minute, transforming into a soft gel state, firmly attached to the tracheal cavity surface. It can be seen that the above in vivo and in vitro verifications have shown that the cryo-microneedle patch has the characteristics of rapid and reversible phase change;

[0051] In order to evaluate the therapeutic effect of the cryomicroneedle patch, an animal model of recurrent tracheal stenosis was constructed. In the experiment, the tracheal epithelium of the animal (rabbit) was first scraped and injured to simulate the pathological process of tracheal stenosis. 14 days after the injury, the hyperplasia of the crystal stenosis site was removed by surgery, and this date was used as the starting point of the study (day 0). Finally, the cryomicroneedle patch was used to treat the injured site to evaluate its therapeutic effect: Figure 5 The therapeutic effect of the cryomicroneedle patch on the injured part in the animal model of recurrent tracheal stenosis; Figure 5 As shown, the control group that did not use the cryomicroneedle patch had obvious tracheal stenosis, severe fibrosis, and a stenosis area greater than 75%, reaching the level of grade 4 tracheal stenosis. In contrast, the degree of tracheal stenosis in the experimental group that used the cryomicroneedle patch was significantly reduced. This result shows that the cryomicroneedle patch can significantly alleviate the symptoms of recurrent airway stenosis;

[0052] In order to further verify the clinical value and application value of the cryo-microneedle patch described in the present invention, further experimental verification was carried out on large animals (sheep). In the experiment, the tracheal epithelium of the animal (sheep) was first scraped and injured to simulate the pathological process of tracheal stenosis. 14 days after the injury, the hyperplasia removal surgery of the crystal stenosis site was performed, and this date was used as the starting point of the study (day 0). Finally, the cryo-microneedle patch (1mm patch, 600μm microneedle length cryo-microneedle patch) was used to treat the injured area. Through continuous bronchoscopy and monitoring of respiratory conditions, we euthanized the experimental animals on the 21st day and obtained pathological samples of the tracheal stenosis site for examination, among which, Figure 6 The gross appearance and pathological examination results of the tracheal stenosis site 21 days after treatment with cryomicroneedle patch. Figure 6 It can be seen that the blank control group that did not use the cryomicroneedle patch showed obvious stenosis, the degree of stenosis reached grade 2, and was accompanied by complications such as infection and bleeding. In addition, the respiratory state of the animals was also unstable, and the respiratory rate increased significantly. In the experimental group treated with the cryomicroneedle patch, the recurrent stenosis was significantly controlled, and the condition of the postoperative wound was also better. Due to the presence of the viscous barrier, infection, bleeding and other conditions were significantly improved, and the wound achieved better repair. It can be seen that the cryomicroneedle patch of the present invention can not only significantly reduce recurrent airway stenosis, but also has good clinical application prospects.

[0053] Example 2

[0054] After glycerol and sebacic acid are put into a reactor at a molar ratio of 1:1.5, an auxiliary agent (including RepSox and Y27632 at a mass ratio of 1:1) with a mass percentage of 5% is added, and while heating to 140°C, a nitrogen-bubbling needle is inserted into the obtained mixed system, and stirring is continued for 12 hours, and then the nitrogen is removed, and the reactor is continuously evacuated with a vacuum pump, and the reaction is continued for 12 hours to obtain a linear paste of polysebacic acid glyceride doped with an auxiliary agent;

[0055] The preparation of the cryo-microneedle patch is described in reference to Example 1.

[0056] Comparative Example 1

[0057] Refer to Example 1, except that no low-temperature molding treatment is performed, and a room-temperature microneedle patch is obtained.

[0058] Test Case

[0059] Figure 7 The therapeutic effect diagram of the room temperature microneedle patch described in Comparative Example 1 (denoted as MN), the frozen microneedle patch described in Example 1 (denoted as Cryo-MN), and the frozen microneedle patch described in Example 2 (denoted as Cryo-RY@MN) is shown in FIG. Figure 7 It can be seen that the cryo-microneedle patch has the effects of anti-fibrosis and inhibiting airway stenosis. The addition of additives can further increase the anti-fibrosis ability and promote the structural repair of the airway.

[0060] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. The use of a viscous gel in preparing a cryotherapy patch, characterized in that: The viscous gel is in a crystalline phase in an environment below -196°C, and is in a soft gel phase in an environment at human body temperature.

2. The use according to claim 1, characterized in that The mass percentage of the adhesive gel in the cryotherapy patch is 80% to 100%.

3. The use according to claim 1, characterized in that The cryotherapy patch also includes an adjuvant; The mass percentage of the auxiliary agent in the cryotherapy patch is 1-20%.

4. The use according to claim 3, characterized in that The auxiliary agent includes one or more of antibacterial drugs, anti-fibrosis drugs and drugs promoting epithelial regeneration.

5. The use according to claim 4, characterized in that The antibacterial drugs include one or more of β-lactam antibiotics, aminoglycoside antibiotics, macrolide antibiotics, tetracycline antibiotics and fluoroquinolone antibiotics; The anti-fibrosis drugs include the small molecule inhibitor RepSox and / or the drug pirfenidone; The drug for promoting epithelial regeneration includes the small molecule inhibitor Y27632.

6. The use according to any one of claims 1 to 5, characterized in that: The viscous gel includes one or more of polylipoic acid hydrogel, acrylic acid / vinyl pyrrolidone copolymer succinimide ester hydrogel, lipoic acid / polyethylene glycol copolymer succinimide ester hydrogel, polyaspartic acid hydrogel, polysebacic acid glyceryl hydrogel and linear paste polysebacic acid glyceryl gel.

7. The use according to claim 6, characterized in that The viscous gel includes a linear, creamy polysebacylglycerol gel.

8. The use according to claim 7, characterized in that The preparation method of the linear paste-like polyglyceryl sebacate gel comprises the following steps: Glycerol and sebacic acid are mixed, and a first polymerization reaction is carried out under nitrogen filling conditions, and then a second polymerization reaction is carried out under vacuum conditions to obtain the linear paste-like polysebacic acid glyceride gel.

9. The use according to claim 8, characterized in that The molar ratio of the glycerol to sebacic acid is 1:(1-1.5).

10. The use according to claim 8 or 9, characterized in that: The temperature of the first polymerization reaction is 120-140° C. and the time is 4-24 hours; The temperature of the second polymerization reaction is 120-140° C., and the time is 12-48 hours.

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