A device combination and method of use for long acting sustained release treatment of HPV infection in the female vagina

Abstract

The present application relates to the technical field of HPV repair, in particular to a long-acting sustained-release device combination for treating female vaginal HPV infection and a use method thereof, which provides a safe, low-irritation and convenient-to-use drug delivery mode, and comprises a vaginal drug delivery device A and a device B used in cooperation with each other, the magnetic therapy of the vaginal drug delivery device A can promote capillary dilation and improve microcirculation, the IgY in the drug sustained-release layer of the vaginal drug delivery device A can be stably kept in the gel at a high titer, the alginate sulfate-carboxymethyl chitosan composite microemulsion makes the drug more easily gathered at the infection site, the magnetic therapy and the sustained-release drug interact with each other, promote the absorption and dissipation of inflammation, and produce a good repair environment, the functional powder dressing of the subsequent vaginal drug delivery device B is convenient for mild absorption of the exudate on the surface of the vagina after the magnetic therapy and the drug sustained-release treatment without causing discomfort, and the functional powder dressing releases various ions at a certain rate, further repairing damaged cells and tissues.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of HPV infection, in particular to a device combination and use method for long-acting sustained-release treatment of female vaginal HPV infection. BACKGROUND

[0002] HPV (Human papillomavirus, human papillomavirus) is a class of double-stranded DNA viruses with epithelium tropism, which is composed of nucleic acid and capsid protein. HPV virus has epithelium tropism, and can replicate in large quantities in the epithelial basal cells of epidermis, genital organs, oral mucosa and other parts after infecting the host. When the body's immunity is reduced, it can randomly integrate into the host genome, and then silence the E2 gene, thereby reducing its inhibition of E6 and E7 genes, making E6 and E7 proteins overexpressed, supplemented by the expression of E5 protein, thereby inducing infected cells to proliferate indefinitely or even cancer. According to epidemiological statistics, almost all cases of cervical cancer are caused by HPV infection. Although most HPV infections can be cleared by the body's own immune system, in some cases, HPV infection can become persistent. As can be seen, the prevention and control of HPV virus is of great significance to the physiological health of women.

[0003] However, in the prior art, the prevention and treatment of HPV virus mainly relies on vaccines or interferons, and the commonly used vaginal external drugs for HPV prevention and repair have poor effect. Chinese Invention Application CN202210211404.X discloses an anti-HPV virus yolk immunoglobulin gel and a preparation method thereof. The anti-HPV virus yolk immunoglobulin gel comprises the following raw materials: anti-HPV yolk immunoglobulin, alginate, alkali gelatin, carbomer, neutralizing agent, preservative, polyethylene glycol, glycerol and purified water. The preparation method of the anti-HPV virus yolk immunoglobulin gel comprises: taking the above-mentioned raw materials and mixing them uniformly to obtain a mixed solution; and emulsifying the mixed solution to obtain the target gel. However, this technology still has limitations: 1. Yolk immunoglobulin is a protein antibody, which is unstable in structure and prone to denaturation and failure. In actual use, the addition of the technology to the gel reduces the antibody titer and affects the use effect. 2. The drug has a short action time, resulting in a long treatment cycle. 3. The drug is not convenient to administer and is easy to contaminate clothes. 4. It cannot repair the vaginal microecology and vaginal mucosa. SUMMARY

[0004] The purpose of the present application is to overcome the deficiencies in the prior art, and to provide a device combination and use method for long-acting sustained-release treatment of female vaginal HPV infection. The device is safe, low in irritation, and convenient to use. IgY can be stably maintained at a high titer in the gel, the drug has a long action time, and it can repair the vaginal microecology and vaginal mucosa.

[0005] The device combination for long-acting sustained-release treatment of female vaginal HPV infection of the present application comprises a vaginal administration device A and a vaginal administration device B used in cooperation,

[0006] The vaginal administration device A comprises a traction rod, one end of the traction rod is connected with a silica gel sheet, and the other end is connected with an ellipsoidal sealing tube, a permanent magnet is arranged in the sealing tube 6, the outer surface of the sealing tube is coated with a drug sustained-release gel layer, and the outer surface of the drug sustained-release gel layer is coated with release paper,

[0007] The preparation method of the drug sustained-release gel layer comprises the following steps:

[0008] Step A1, preparation of a pectin solution, 30-50 parts by weight of pectin is weighed and added to 300-500 parts by weight of deionized water, and magnetic stirring is performed until the pectin is completely dissolved;

[0009] Step A2, preparation of a drug-pectin mixed solution, 30-60 parts by weight of anti-HPV IgY antibody recombinant human interferon alpha-2b composite nanoliposomes, 50-100 parts by weight of alginate sulfate-carboxymethyl chitosan nanomicelles, 20-50 parts by weight of beta-glucan, and 10-30 parts by weight of nanosilver antibacterial agent are weighed and added to the pectin solution, and magnetic stirring is performed for 10-20 min to obtain the drug-pectin mixed solution;

[0010] Step A3, preparation of a PVA solution, 60-100 parts by weight of polyvinyl alcohol is added to 300-500 parts by weight of deionized water, and stirring is performed at 90°C for 30 min, and then the stirring is continued to cool to room temperature to obtain the PVA solution;

[0011] Step A4, the drug-pectin mixed solution is mixed with the PVA solution, and stirring is performed at room temperature for 10-20 min to uniformly mix them, the mixed solution is poured into a mold, the mold containing the mixed solution is placed in a freezer at -22°C for 8H, and then the mold is removed after being restored to room temperature to obtain the gel sustained-release layer of the vaginal administration device A;

[0012] The vaginal administration device B comprises a plastic storage bottle, a bottle cap, and a drug delivery tube, the plastic storage bottle contains a functional powder dressing, the bottle opening of the plastic storage bottle is provided with external threads, the bottle cap and the drug delivery tube are both provided with internal threads, and the wall of the drug delivery tube is provided with a drug outlet,

[0013] The preparation method of the functional powder dressing is as follows:

[0014] Step B1, 100-200 parts by weight of bioactive glass, 500-700 parts by weight of oligoisomaltose, and 100-200 parts by weight of propolis extract are weighed and placed in a ball mill, and mixed and ground for 20-40 min;

[0015] Step B2: Weigh 5-20 parts by weight of mucosal repair factor and 1-5 parts by weight of vaginal probiotic freeze-dried powder, mix them with the mixed powder obtained in the first step, and stir for 10-20 minutes to obtain the functional powder dressing of vaginal drug delivery device B.

[0016] Preferably, the method for preparing the anti-HPV IgY antibody recombinant human interferon α-2b complex nanoliposomes in step A2 comprises the following steps:

[0017] The first step is to prepare PBS buffer with a pH value of 5-7;

[0018] In the second step, 10-20 parts by weight of anti-HPV egg yolk antibody and 10-20 parts by weight of recombinant human α-2b interferon are dissolved in 400-500 parts by weight of PBS solution to be used as the aqueous phase;

[0019] The third step is to weigh 300-400 parts by weight of ether and 400-600 parts by weight of chloroform, and stir them to form a mixed solution;

[0020] Step 4: weigh 250-280 parts by weight of egg yolk lecithin and 20-50 parts by weight of cholesterol, add them to the mixed solution in step 3, slowly stir to dissolve them, and use them as the organic phase;

[0021] Step 5: slowly inject the aqueous phase from step 2 into the organic phase from step 4, and stir ultrasonically using an ultrasonic stirrer to obtain a mixed liquid;

[0022] Step 6: The mixed liquid in step 5 is subjected to vacuum rotary evaporation at 35-45° C. to remove ether and chloroform;

[0023] In the seventh step, 3000-4000 parts by weight of PBS buffer and 100-200 parts by weight of Tween-60 are added to dissolve the material in the rotary evaporator in step 6, and ultrasonic stirring is continued to completely dissolve the material to obtain anti-HPV egg yolk antibody recombinant human α-2b interferon complex nanoliposomes.

[0024] Preferably, the method for preparing the alginate sulfate-carboxymethyl chitosan microemulsion in step A2 comprises the following steps:

[0025] The first step is to prepare the oil phase by weighing 10-30 parts by weight of lithospermum officinale oil and 20-40 parts of camellia oil, and stirring them with a magnetic stirrer at 500 rpm for 10-20 minutes to obtain the oil phase;

[0026] Second step, water phase preparation, 5-10 parts by weight of carboxymethyl chitosan and 10-20 parts by weight of alginate sulfate are weighed and added into 700-900 parts by weight of deionized water, and the material is completely dissolved by magnetic stirring at a speed of 500 r / min for 50-70 min, to obtain the water phase;

[0027] Third step, mixed surfactant preparation, 80-100 parts by weight of anhydrous ethanol and 80-100 parts by weight of polysorbate 80 are weighed and stirred at a speed of 500 r / min for 10-20 min, to obtain the mixed surfactant;

[0028] Fourth step, oil phase surfactant mixture preparation, the oil phase and the mixed surfactant are mixed, and stirred at a speed of 500 r / min for 10-20 min, to obtain the oil phase surfactant mixture;

[0029] Fifth step, the water phase is stirred at a speed of 500 r / min for 10-20 min, and the oil phase surfactant mixture is added dropwise in the stirring process, and after the dropwise addition is completed, the stirring is continued at the same speed for 20-40 min, to obtain the alginate sulfate-carboxymethyl chitosan composite nanomicroemulsion.

[0030] Preferably, the traction rod is made of hard material, and the sealing tube is made of soft material.

[0031] Preferably, the plastic storage bottle is an elastic plastic storage bottle.

[0032] A method for using a device combination for long-acting and sustained-release treatment of female vaginal HPV infection, which comprises the following steps:

[0033] Step (1) remove the release paper of the vaginal administration device A, insert the sealing tube coated with the drug sustained-release gel layer on the outer surface into the vagina and stay for 2-8 hours;

[0034] Step (2) remove the vaginal administration device A, connect the plastic storage bottle and the administration tube of the vaginal administration device B, insert the administration tube into the vagina, and squeeze the plastic storage bottle, so that the functional powder dressing is sprayed from the drug outlet into the vagina.

[0035] Preferably, it is used every night before going to sleep.

[0036] Anti-HPV IgY antibody recombinant human interferon alpha-2b complex liposome, IgY antibody specificity binds to HPV virus to prevent viral invasion and kill HPV virus, recombinant human interferon alpha-2b has broad-spectrum antiviral effect and multiple immunomodulatory effect, can improve the phagocytic activity of macrophages and enhance the specific cytotoxicity of lymphocytes to target cells, promote and maintain the immune surveillance, immune protection and immune self-stabilizing function of the body, and the complex liposome can greatly enhance the stability of the protein polypeptide drug, so that it can maintain high titer for a long time, and can better enter the vaginal mucosa to kill deep viruses.

[0037] Alginate sulfate-carboxymethyl chitosan microemulsion, alginate sulfate can specifically kill HPV virus, thereby effectively treating female vaginal HPV infection, and carboxymethyl chitosan has good antibacterial performance and can inhibit pathogenic microorganisms in the female vagina, the combination of the two can enhance the stability of the drug, the binding force of the drug to the vaginal mucosa, and the efficacy of the drug.

[0038] Beta-glucan has the effects of anticancer, antibacterial, antiviral, antifungal, antiparasitic, and promoting wound healing.

[0039] Nanosilver antibacterial agent, broad-spectrum antibacterial can kill various vaginal pathogenic microorganisms.

[0040] The drug slow-release gel layer keeps the vagina at a suitable humidity and pH, and produces a good modified environment, which can relieve the irritation and instantaneous sharp pain caused by the rapid increase of local pH when the bioactive glass in the functional powder dressing contacts the vagina, and provides good comfort. Under a moist environment, the cell migration speed is accelerated, which can promote healing, and the patient's comfort is high.

[0041] Strong magnetic ferriboron permanent magnet continuously generates a 3D magnetic field, which can promote capillary dilation, improve microcirculation, promote absorption and dissipation of inflammation, analgesia and spasmolysis, increase red blood cell volume to improve blood and oxygen supply in the body, thereby promoting metabolism and accelerating inflammation absorption and repair of damaged mucosa.

[0042] Bioactive glass is a kind of material that can repair, replace and regenerate tissues in the body, and has the function of forming a bond between tissues and materials. The degradation products of bioactive glass can promote the generation of growth factors, promote cell proliferation, enhance osteoblast gene expression and bone tissue growth, and effectively promote the repair of damaged mucosa.

[0043] Mucosal repair factor is an active protein polypeptide substance that can strongly promote the proliferation and growth of human cells, rapidly repair damaged mucosa, and restore the body's own immunity.

[0044] Oligoisomaltose is a natural prebiotic that can promote the growth of lactobacillus and is used to improve the environment of female vagina and regulate the balance of vaginal flora.

[0045] Propolis extract can inhibit and kill various anaerobes and pathogenic fungi and clean the vagina without any irritation and discomfort, and is safe to use, and can treat cervical erosion, erosive cervical ectropion, endometritis, vaginitis, trichomonas vaginitis and other diseases, and has a remarkable effect on promoting the healing of vaginal wounds after gynecological surgery.

[0046] Vaginal probiotics are mainly live lactobacillus, which can supplement a large number of healing bacteria to the vagina, regulate the balance of vaginal flora, establish a defense barrier of probiotic film, restore the self-cleaning function of the vagina, and effectively improve the immunity of the vagina.

[0047] The present application provides a safe, low-irritating, convenient-to-use HPV infection treatment administration mode, which comprises a vaginal administration device A and a vaginal administration device B used in cooperation, and is used at different times, can play a synergistic effect and enhance the treatment effect. The magnetic therapy of the vaginal administration device A can promote capillary dilation and improve microcirculation, the IgY in the drug sustained-release layer of the vaginal administration device A can be stably maintained at a high titer in the gel, the alginate sulfate-carboxymethyl chitosan composite microemulsion makes the drug more easily gathered in the infected site, the magnetic therapy and the sustained-release drug interact with each other, promote the absorption and dissipation of inflammation, the vagina is treated and repaired to a certain extent, and a good repair environment is generated, which is convenient for the functional powder dressing of the subsequent vaginal administration device B. The functional powder dressing gently absorbs the exudate on the surface of the vagina after the magnetic therapy and drug sustained-release treatment without causing discomfort, and releases various ions at a certain rate to further repair damaged cells and tissues in a gentle and comfortable condition. BRIEF DESCRIPTION OF DRAWINGS

[0048] Figure 1 is a structural schematic view of the vaginal administration device A of the present application;

[0049] Figure 2 is a structural schematic view of the vaginal administration device B of the present application.

[0050] Reference signs:

[0051] Figure 1 In the vaginal administration device A, 1 is a silica gel sheet 1, 2 is a traction rod, 3 is release paper, 4 is a drug sustained-release gel layer, 5 is a permanent magnet, and 6 is a sealed tube.

[0052] Figure 2 In the vaginal administration device B, 1' is a plastic storage bottle, 2' is a bottle cap, 3' is a drug delivery tube, and 4' is a drug outlet hole. DETAILED DESCRIPTION

[0053] The application will be further described in conjunction with the following examples and drawings.

[0054] Example 1

[0055] A device combination for long-acting sustained-release treatment of female vaginal HPV infection, comprising a vaginal administration device A and a vaginal administration device B used in conjunction, the vaginal administration device A comprising a traction rod 2, one end of the traction rod 2 being connected with a silica gel sheet 1, the other end being connected with an ellipsoidal sealing tube 6, three permanent magnets 5 of different orientations being arranged inside the sealing tube 6, the outer surface of the sealing tube 6 being coated with a drug sustained-release gel layer 4, the outer surface of the drug sustained-release gel layer 4 being coated with release paper 3, the traction rod 2 being made of hard material, and the sealing tube 6 being made of soft material, polyethylene material being used.

[0056] The preparation method of the drug sustained-release gel layer 4 comprises the following steps:

[0057] Step A1, preparation of pectin solution, 30 parts by weight of pectin is weighed and added to 300 parts by weight of deionized water, and magnetic stirring is performed until the pectin is completely dissolved;

[0058] Step A2, preparation of drug-pectin mixed solution, 30 parts by weight of anti-HPV IgY antibody recombinant human interferon alpha-2b composite nanoliposomes, 50 parts by weight of alginate sulfate-carboxymethyl chitosan nanomicelles, 20 parts by weight of β-glucan, and 10 parts by weight of nanosilver antibacterial agent are weighed and added to the above-mentioned pectin solution, and magnetic stirring is performed for 10 min to obtain the drug-pectin mixed solution;

[0059] Step A3, preparation of PVA solution, 60 parts by weight of polyvinyl alcohol is added to 300 parts by weight of deionized water, and heating stirring is performed at 90℃ for 30 min, and then stirring is continued to cool to room temperature to obtain the PVA solution;

[0060] Step A4, mixing of the drug-pectin mixed solution and the PVA solution, stirring is performed at room temperature for 10 min to make them uniformly mixed, the mixed solution is poured into a mold, the mold containing the mixed solution is placed in a freezer at-22℃ for 8H, and then it is restored to room temperature, the mold is removed, and the gel sustained-release layer of the vaginal administration device A is obtained;

[0061] The vaginal administration device B comprises a plastic storage bottle 1', a bottle cap 2', and a drug administration tube 3', the plastic storage bottle 1' contains functional powder dressing, the bottle opening of the plastic storage bottle 1' is provided with external threads, the bottle cap 2' and the drug administration tube 3' are both provided with internal threads, and the tube wall of the drug administration tube 3' is provided with a drug outlet hole 4', the plastic storage bottle 1' is an elastic plastic storage bottle 1',

[0062] The preparation method of the functional powder dressing is as follows:

[0063] Step B1, take 100 parts by weight of bioactive glass, 500 parts by weight of oligoisomaltose and 100 parts by weight of propolis extract, put into a ball mill, mix and grind for 20 min;

[0064] Step B2, take 5 parts by weight of mucosal repair factor and 1 part by weight of freeze-dried powder of vaginal probiotics, mix with the mixed powder obtained in the first step, stir for 10 min, and obtain the functional powder dressing of the vaginal administration device B.

[0065] The preparation method of the anti-HPV IgY antibody recombinant human interferon α-2b complex nanoliposome in step A2 comprises the following steps:

[0066] First step, prepare PBS buffer with pH value of 5;

[0067] Second step, dissolve 10 parts by weight of anti-HPV egg yolk antibody and 10 parts by weight of recombinant human α-2b interferon in 400 parts by weight of PBS solution as the aqueous phase;

[0068] Third step, weigh 300 parts by weight of diethyl ether and 400 parts by weight of chloroform, stir uniformly to form a mixed solution;

[0069] Fourth step, weigh 250 parts by weight of egg yolk lecithin and 20 parts by weight of cholesterol, add to the mixed solution of the third step, slowly stir to dissolve, as the organic phase;

[0070] Fifth step, slowly inject the aqueous phase of the second step into the organic phase of the fourth step, use an ultrasonic stirrer to ultrasonically stir to obtain a mixed liquid;

[0071] Sixth step, the mixed liquid of the fifth step is subjected to vacuum rotary evaporation at 35°C to remove diethyl ether and chloroform;

[0072] Seventh step, add 3000 parts by weight of PBS buffer and 100 parts by weight of Tween-60 to dissolve the material in the rotary evaporator of step six, continue to ultrasonically stir to completely dissolve the material, and obtain the anti-HPV egg yolk antibody recombinant human α-2b interferon complex nanoliposome.

[0073] The preparation method of alginate sulfate-carboxymethyl chitosan microemulsion in step A2 comprises the following steps:

[0074] First step, oil phase preparation, take 10 parts by weight of radix lithospermi oil and 20 parts by weight of camellia oil, 500 r / min magnetic stirring for 10 min, and obtain the oil phase;

[0075] Second step, water phase preparation, 5 parts by weight of carboxymethyl chitosan and 10 parts by weight of alginate sulfate are weighed into 700 parts by weight of deionized water, and the material is completely dissolved by magnetic stirring at a speed of 500 r / min for 50 min, and the water phase is obtained;

[0076] Third step, preparation of mixed surfactant, 80 parts by weight of anhydrous ethanol and 80 parts by weight of polysorbate 80 are weighed, and magnetic stirring is carried out at a speed of 500 r / min for 10 min, and the mixed surfactant is obtained;

[0077] Fourth step, preparation of oil phase surfactant mixture, the oil phase and the mixed surfactant are mixed, and magnetic stirring is carried out at a speed of 500 r / min for 10 min, and the oil phase surfactant mixture is obtained;

[0078] Fifth step, the water phase is stirred at a speed of 500 r / min for 10 min, and the oil phase surfactant mixture is added dropwise into the stirring solution, and after the addition is completed, the stirring is continued at the same speed for 20 min, and the alginate sulfate-carboxymethyl chitosan composite nano microemulsion is obtained.

[0079] A method for using a device combination for long-acting sustained-release treatment of female vaginal HPV infection, comprising the following steps:

[0080] Step (1) tear off the release paper 3 of the vaginal administration device A, and insert the sealed tube 6 coated with the drug sustained-release gel layer 4 on the outer surface into the vagina and stay for 2 hours;

[0081] Step (2) pull out the vaginal administration device A, connect the plastic storage bottle 1' and the administration tube 3' of the vaginal administration device B, insert the administration tube 3' into the vagina, and squeeze the plastic storage bottle 1' to spray the functional powder dressing from the drug outlet hole 4' into the vagina.

[0082] Use every night before going to sleep. Example 2

[0083] A device combination for long-acting sustained-release treatment of female vaginal HPV infection, comprising a vaginal administration device A and a vaginal administration device B used in cooperation, the vaginal administration device A comprising a traction rod 2, one end of the traction rod 2 being connected with a silica gel sheet 1, and the other end being connected with an ellipsoidal sealed tube 6, the sealed tube 6 being internally provided with three permanent magnets 5 with different orientations, the outer surface of the sealed tube 6 being coated with a drug sustained-release gel layer 4, and the outer surface of the drug sustained-release gel layer 4 being coated with a release paper 3, the traction rod 2 being made of a hard material, and the sealed tube 6 being made of a soft material and being made of polyethylene material.

[0084] The preparation method of the drug sustained-release gel layer 4 comprises the following steps:

[0085] Step A1, preparation of pectin solution, 40 parts by weight of pectin was weighed and added to 400 parts by weight of deionized water, and magnetically stirred until the pectin was completely dissolved;

[0086] Step A2, preparation of drug pectin mixed solution, 40 parts by weight of anti-HPV IgY antibody recombinant human interferon alpha-2b complex nanoliposome, 80 parts by weight of alginate sulfate-carboxymethyl chitosan nanomicroemulsion, 30 parts by weight of beta-glucan and 20 parts by weight of nanosilver antibacterial agent were added to the above pectin solution, and magnetically stirred for 15 min to obtain the drug pectin mixed solution;

[0087] Step A3, preparation of PVA solution, 80 parts by weight of polyvinyl alcohol was added to 400 parts by weight of deionized water, heated and stirred at 90℃ for 30 min, and then continuously stirred and cooled to room temperature to obtain the PVA solution;

[0088] Step A4, mixing the drug pectin mixed solution with the PVA solution, stirring at room temperature for 15 min to make them uniformly mixed, pouring the mixed solution into a mold, placing the mold containing the mixed solution in a freezer at-22℃ for 8H, restoring to room temperature, and removing the mold to obtain the gel sustained-release layer of the vaginal administration device A;

[0089] The vaginal administration device B comprises a plastic storage bottle 1', a bottle cap 2' and a administration tube 3', the plastic storage bottle 1' contains functional powder dressing, the bottle mouth of the plastic storage bottle 1' is provided with external threads, the bottle cap 2' and the administration tube 3' are both provided with internal threads, the tube wall of the administration tube 3' is provided with a medicine outlet hole 4', and the plastic storage bottle 1' is an elastic plastic storage bottle 1'.

[0090] The preparation method of the functional powder dressing is as follows:

[0091] Step B1, 150 parts by weight of bioactive glass, 600 parts by weight of oligoisomaltose and 150 parts by weight of propolis extract were placed in a ball mill and mixed and ground for 30 min;

[0092] Step B2, 10 parts by weight of mucosal repair factor and 3 parts by weight of vaginal probiotic freeze-dried powder were mixed with the mixed powder obtained in the first step, and stirred for 15 min to obtain the functional powder dressing of the vaginal administration device B.

[0093] The preparation method of the anti-HPV IgY antibody recombinant human interferon alpha-2b complex nanoliposome in step A2 comprises the following steps:

[0094] First step, prepare PBS buffer with pH value of 6;

[0095] Second step, 15 parts by weight of the anti-HPV yolk antibody and 15 parts by weight of the recombinant human α-2b interferon are dissolved in 450 parts by weight of the PBS solution as the aqueous phase;

[0096] Third step, 350 parts by weight of the ether and 500 parts by weight of the chloroform are weighed and stirred uniformly into a mixed solution;

[0097] Fourth step, 260 parts by weight of the egg yolk lecithin and 30 parts by weight of the cholesterol are weighed and added into the mixed solution of the third step, and are slowly stirred to be dissolved as the organic phase;

[0098] Fifth step, the aqueous phase of the second step is slowly injected into the organic phase of the fourth step, and an ultrasonic stirrer is used for ultrasonic stirring to obtain a mixed liquid;

[0099] Sixth step, the mixed liquid of the fifth step is subjected to vacuum rotary evaporation at 40°C to remove the ether and the chloroform;

[0100] Seventh step, 3500 parts by weight of the PBS buffer and 150 parts by weight of the Tween-60 are added to dissolve the material in the rotary evaporator of the sixth step, and the material is completely dissolved by continuing ultrasonic stirring to obtain the anti-HPV yolk antibody recombinant human α-2b interferon complex nanoliposome.

[0101] The preparation method of the alginate sulfate-carboxymethyl chitosan microemulsion in the step A2, which comprises the following steps:

[0102] First step, oil phase preparation, 20 parts by weight of the borage oil and 30 parts by weight of the camellia oil are weighed and stirred at 500 r / min for 15 min to obtain the oil phase;

[0103] Second step, water phase preparation, 8 parts by weight of the carboxymethyl chitosan and 15 parts by weight of the alginate sulfate are weighed and added into 800 parts by weight of the deionized water, and the material is completely dissolved by stirring at 500 r / min for 60 min to obtain the water phase;

[0104] Third step, mixed surfactant preparation, 90 parts by weight of the anhydrous ethanol and 90 parts by weight of the polysorbate 80 are weighed and stirred at 500 r / min for 15 min to obtain the mixed surfactant;

[0105] Fourth step, oil phase surfactant mixture preparation, the oil phase and the mixed surfactant are mixed, and the mixture is stirred at 500 r / min for 15 min to obtain the oil phase surfactant mixture;

[0106] Fifth step, the above water phase 500 r / min magnetic stirring 15 min, the stirring speed is not changed, in the stirring dropwise adding the above oil phase surfactant mixture, dropwise adding is finished, under the same speed stirring 30 min, namely the alginate sulfate-carboxymethyl chitosan composite nano microemulsion is obtained.

[0107] A method of using a device combination for long-acting sustained-release treatment of HPV infection in the vagina of a female, comprising the following steps:

[0108] Step (1) tear off the release paper 3 of the vaginal administration device A, and insert the sealed tube 6 coated with the drug sustained-release gel layer 4 on the outer surface into the vagina and stay for 4 hours;

[0109] Step (2) pull out the vaginal administration device A, and then connect the plastic storage bottle 1' and the administration tube 3' of the vaginal administration device B, insert the administration tube 3' into the vagina, and squeeze the plastic storage bottle 1', so that the functional powder dressing is sprayed from the drug outlet hole 4' into the vagina.

[0110] Use every night before going to sleep.

[0111] Example 3

[0112] A device combination for long-acting sustained-release treatment of HPV infection in the vagina of a female, comprising a vaginal administration device A and a vaginal administration device B used in cooperation, wherein the vaginal administration device A comprises a traction rod 2, one end of the traction rod 2 is connected with a silica gel sheet 1, and the other end is connected with an ellipsoidal sealed tube 6, the sealed tube 6 is internally provided with three permanent magnets 5 with different orientations, the outer surface of the sealed tube 6 is coated with a drug sustained-release gel layer 4, the outer surface of the drug sustained-release gel layer 4 is coated with a release paper 3, the traction rod 2 is made of a hard material, and the sealed tube 6 is made of a soft material and is made of polyethylene material.

[0113] The preparation method of the drug sustained-release gel layer 4 comprises the following steps:

[0114] Step A1, preparation of a pectin solution, weigh 50 parts by weight of pectin and add to 500 parts by weight of deionized water, and magnetically stir until the pectin is completely dissolved;

[0115] Step A2, preparation of a drug-pectin mixed solution, weigh 60 parts by weight of anti-HPV IgY antibody recombinant human interferon α-2b composite nano-liposome, 100 parts by weight of alginate sulfate-carboxymethyl chitosan nano-microemulsion, 50 parts by weight of β-glucan, and 30 parts by weight of nano-silver antibacterial agent, and add to the above pectin solution, and magnetically stir for 20 min to obtain the drug-pectin mixed solution;

[0116] Step A3, preparation of PVA solution, 100 parts by weight of polyvinyl alcohol, 500 parts by weight of deionized water, heated stirring 30 min at 90℃, continue to cool to room temperature, stirring, PVA solution is obtained;

[0117] Step A4, the drug pectin mixture and PVA solution, stirring at room temperature for 20 min, make it mixed evenly, the mixture is poured into the mold, the mold with the mixture is placed in-22℃ freezing 8H, restore to room temperature, remove the mold, the gel sustained release layer 4 of the vaginal administration device A is obtained;

[0118] The vaginal administration device B comprises a plastic storage bottle 1', a bottle cap 2' and a administration tube 3', the plastic storage bottle 1' contains functional powder dressing, the bottle mouth of the plastic storage bottle 1' is provided with external threads, the bottle cap 2' and the administration tube 3' are provided with internal threads, the wall of the administration tube 3' is provided with a medicine outlet hole 4', and the plastic storage bottle 1' is an elastic plastic storage bottle 1'.

[0119] The preparation method of the functional powder dressing is as follows:

[0120] Step B1, 200 parts by weight of bioactive glass, 700 parts by weight of oligoisomaltose and 200 parts by weight of propolis extract are weighed and put into a ball mill for mixing and grinding for 40 min;

[0121] Step B2, 20 parts by weight of mucosal repair factor and 5 parts by weight of vaginal probiotic freeze-dried powder are weighed and mixed with the mixed powder obtained in the first step, and stirring for 20 min, the functional powder dressing of the vaginal administration device B is obtained.

[0122] The preparation method of the anti-HPV IgY antibody recombinant human interferon alpha-2b complex nano-liposome in step A2 comprises the following steps:

[0123] First step, preparation of PBS buffer solution with pH value of 7;

[0124] Second step, 10-20 parts by weight of anti-HPV egg yolk antibody and 20 parts by weight of recombinant human alpha-2b interferon are dissolved in 500 parts by weight of PBS solution as the water phase;

[0125] Third step, 400 parts by weight of diethyl ether and 600 parts by weight of chloroform are weighed and stirred uniformly into a mixed solution;

[0126] Fourth step, 280 parts by weight of egg yolk lecithin and 50 parts by weight of cholesterol are weighed and added to the mixed solution of the third step, and slowly stirred to dissolve, as the organic phase;

[0127] Fifth step, the water phase of the second step is slowly injected into the organic phase of the fourth step, and an ultrasonic stirrer is used for ultrasonic stirring to obtain a mixed liquid;

[0128] Step 6: The mixed liquid in step 5 is subjected to vacuum rotary evaporation at 45° C. to remove ether and chloroform;

[0129] In the seventh step, 4000 parts by weight of PBS buffer and 200 parts by weight of Tween-60 were added to dissolve the material in the rotary evaporator in step six, and ultrasonic stirring was continued to completely dissolve the material to obtain anti-HPV egg yolk antibody recombinant human α-2b interferon complex nanoliposomes.

[0130] The method for preparing alginate sulfate-carboxymethyl chitosan microemulsion in step A2 comprises the following steps:

[0131] The first step is to prepare the oil phase by weighing 30 parts by weight of lithospermum officinale oil and 40 parts of camellia oil, and stirring them with a magnetic stirrer at 500 rpm for 20 minutes to obtain the oil phase;

[0132] The second step is to prepare the aqueous phase by weighing 10 parts by weight of carboxymethyl chitosan and 20 parts by weight of alginate sulfate, adding them to 900 parts by weight of deionized water, and magnetically stirring at a speed of 500 rpm for 70 minutes until the materials are completely dissolved to obtain the aqueous phase;

[0133] The third step is to prepare a mixed surfactant by weighing 100 parts by weight of anhydrous ethanol and 100 parts by weight of polysorbate 80, and magnetically stirring at 500 rpm for 20 minutes to obtain a mixed surfactant;

[0134] The fourth step is to prepare the oil phase surfactant mixture, wherein the oil phase and the mixed surfactant are mixed, and magnetic stirring is performed at 500 rpm for 20 minutes to obtain the oil phase surfactant mixture;

[0135] The fifth step is to magnetically stir the aqueous phase at 500 r / min for 20 min, keep the stirring speed constant, and add the oil phase surfactant mixture dropwise during stirring. After the addition is complete, stir at the same speed for 40 min to obtain alginate sulfate-carboxymethyl chitosan composite nano-microemulsion.

[0136] A method for using a device combination for long-acting sustained-release treatment of HPV infection in female vagina, comprising the following steps:

[0137] Step (1) tear off the release paper 3 of the vaginal drug delivery device A, and insert the sealed tube 6 coated with the drug sustained-release gel layer 4 into the vagina for 8 hours;

[0138] Step (2) extract the vaginal medication device A, connect the plastic storage bottle 1' and medication tube 3' of the vaginal medication device B, insert the medication tube 3' into the vagina, squeeze the plastic storage bottle 1', and spray the functional powder dressing from the medication outlet 4' into the vagina.

[0139] Used before going to bed every night.

[0140] Performance test of final product.

[0141] Drug release performance

[0142] Test group, IgY antibody recombinant human interferon α-2b complex nano-liposome drug release gel was prepared according to the preparation process of the drug release gel layer of device A of Example 2 of the present application.

[0143] Control group, the carbomer gel was swelled overnight, and triethanolamine was added dropwise in stirring, and then IgY antibody recombinant human interferon α-2b complex nano-liposome of Example 2 of the present application was added, and stirring was performed to obtain the gel.

[0144] 10 g of the above-mentioned drug release gel and 10 g of the carbomer gel were respectively put into beakers containing 200 ml of simulated body fluid, and were left to stand for extraction. After 1H, 2H, 4H, 6H, 8H and 16H, 150 ml of the upper extraction liquid was carefully drawn, and 150 ml of simulated body fluid was added to the extraction beaker at the same time. The drug release performance was evaluated by detecting the protein content of the extracted liquid. The detection results are shown in Table 1:

[0145] .

[0146] Result analysis: the control group has dissolved most of the drug components in the first two hours, and has basically completely dissolved after 4H; the test group (drug release gel of the present application) can slowly and uniformly dissolve the drug within 12H, and the extraction liquid can maintain a moderate drug concentration for a long time. The drug release gel of the present application has good drug release performance, and can significantly prolong the drug action time, and at the same time, avoid the stimulation caused by high drug concentration in a short time.

[0147] Protection performance of antibody recombinant human interferon α-2b complex liposome on antibody titer

[0148] Test group, IgY antibody recombinant human interferon α-2b complex nano-liposome drug release gel was prepared according to the preparation process of the drug release gel layer of device A of Example 2 of the present application.

[0149] Control group, the carbomer was swelled overnight, and triethanolamine was added dropwise in stirring, and then IgY antibody and recombinant human interferon α-2b of Example 2 were respectively added, and stirring was performed to obtain the interferon complex gel of the IgY antibody group.

[0150] The test group and the control group gels are stored under the same conditions, and samples are taken at 0 months, 6 months, 12 months, 18 months, 24 months, 30 months and 36 months, respectively. The samples are extracted for 24 hours, and the protective performance of the complex liposome on the antibody titer is evaluated by detecting the antibody titer of the extract. The detection results are shown in Table 2:

[0151] .

[0152] Result analysis: IgY antibody is an immunoglobulin, and the protein is prone to structural changes, denaturation, decomposition and other adverse effects in a common storage environment, resulting in a decrease in antibody titer. The control group has already lost titer during preparation, and the antibody titer decreases significantly during storage. After 18 months, the antibody titer is basically undetectable. The nanoliposome can encapsulate the IgY antibody in a protective film, significantly delaying the denaturation and decomposition of the protein that affect the antibody titer. The antibody in the test group can still maintain a high level after 30 months of storage. The IgY antibody recombinant human interferon alpha-2b complex liposome of the application has excellent protective performance on the antibody titer.

[0153] 1.3 Treatment effect

[0154] 1.3.1 HPV virus negative conversion time and negative conversion rate

[0155] Select 60 female patients with positive HPV-DNA detection of cervical exfoliative cells, and divide them into two groups. The test group has 30 cases, and the application is used for treatment, once a day, before going to bed at night, using device A of the application, and then taking out device A after sleeping, and then spraying the powder dressing into the vagina by using device B. The control group has 30 cases, and recombinant human interferon alpha-2b gel is used for treatment, once a day before going to bed at night. The cervical exfoliative cells HPV-DNA detection of the patients in the two groups is rechecked on the 7th day, the 14th day, the 21st day, the 28th day and the 35th day. The results are shown in Table 3:

[0156] .

[0157] Result analysis: As can be seen from the above table, the average negative conversion time of the test group has a significant advantage compared with the control group, and the final negative conversion rate is also significantly higher than that of the control group. The application has the advantages of short negative conversion time and high negative conversion rate in treating female vaginal HPV virus infection.

[0158] Bacterial vaginosis and vaginal microecological mucosal repair performance

[0159] Select 120 cases of bacterial vaginitis patients, average into two groups. 60 cases in the test group, using the application for treatment, once a day, before going to sleep, using the device A of the application, after sleeping, the device A is extracted, and then the powder dressing is sprayed into the vagina by using the device B, and the treatment is stopped after cure. 60 cases in the control group, using carbomer vaginal tampon gel for treatment, once a day, and the treatment is stopped after cure. The patients in the two groups are reexamined on the 7th day, the 14th day, the 21st day, the 28th day and the 35th day, the improvement of symptoms is effective, and the disappearance of symptoms is cured, and the statistical results are shown in table 4. Follow-up is carried out at 1 month, 3 months, 6 months, 9 months and 12 months after cure, and the recurrence of vaginitis is understood, and the results are recorded in table 5.

[0160] .

[0161] Result analysis: the statistical results in table 4 show that the application has excellent curative effect on bacterial vaginitis, fast onset speed and short cure time.

[0162] .

[0163] Result analysis: the statistical results in table 5 show that the application has excellent curative effect on bacterial vaginitis, fast onset speed and short cure time.

[0164] Experiment summary:

[0165] The drug sustained-release gel layer of the application can slowly and uniformly release the treatment drug, prolong the drug treatment time, reduce the stimulation of the drug to the vagina and shorten the treatment time.

[0166] The IgY antibody recombinant human interferon alpha-2b complex liposome can effectively protect the stability of IgY in the preparation, maintain high titer for a long time, and ensure excellent treatment effect.

[0167] The application improves the stability of alginate sulfate and the binding force with the vaginal mucosa by preparing alginate sulfate-carboxymethyl chitosan composite microemulsion, and alginate sulfate is more easily gathered in the infection site, so that the killing and inhibiting effect of alginate sulfate on HPV virus is enhanced.

[0168] The application can better and faster kill HPV virus by using a plurality of effective components, and cooperating with 3D magnetic therapy, and the conversion is fast and the conversion rate is high.

[0169] The application can effectively repair the vaginal flora balance and maintain a good vaginal microecology by the treatment of device A and the repair of device B while treating the infection and inflammation. The bioactive glass, mucosal repair factor, oligoisomaltose, propolis extract and vaginal probiotics in device B can repair the vaginal microecology, and cooperate with 3D magnetic therapy to accelerate the repair of vaginal mucosa, restore the self-purification ability of the vagina, improve the vaginal immunity, significantly reduce the recurrence of vaginal inflammation and reduce the occurrence of vaginal diseases.

[0170] Finally, it should be noted that the above examples are only used to illustrate the technical solutions of the present application, and are not a limitation on the scope of protection of the present application. Although the present application has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present application can be modified or replaced by equivalents without departing from the essence and scope of the technical solutions of the present application.

Claims

1. A device combination for treating female vaginal HPV infection with a long-acting sustained-release agent, characterized by: It includes a vaginal drug delivery device A and a vaginal drug delivery device B that are used in conjunction with each other. Vaginal drug delivery device A includes a traction rod, one end of which is connected to a silicone sheet, and the other end is connected to an ellipsoidal sealing tube, a permanent magnet is provided inside the sealing tube, and the outer surface of the sealing tube is covered with a drug-releasing gel layer, and the outer surface of the drug-releasing gel layer is covered with release paper. The preparation method of the drug sustained-release gel layer is as follows: Step A1, preparing a pectin solution: weigh 30-50 parts by weight of pectin, add it to 300-500 parts by weight of deionized water, and stir magnetically until the pectin is completely dissolved; Step A2, preparing a drug-pectin mixed solution, weighing 30-60 parts by weight of anti-HPV egg yolk antibody recombinant human interferon α-2b complex nanoliposomes, 50-100 parts by weight of alginate sulfate-carboxymethyl chitosan composite microemulsion, 20-50 parts by weight of β-glucan and 10-30 parts by weight of nanosilver antibacterial agent, adding the mixture to the above pectin solution, and magnetically stirring for 10-20 minutes to obtain the drug-pectin mixed solution; Step A3, preparing a PVA solution: add 300-500 parts by weight of deionized water to 60-100 parts by weight of polyvinyl alcohol, heat and stir at 90° C. for 30 minutes, continue stirring and cool to room temperature to obtain a PVA solution; Step A4: Mix the drug-pectin mixture with the PVA solution, stir at room temperature for 10-20 minutes to mix evenly, pour the mixture into a mold, freeze the mold containing the mixture at -22°C for 8 hours, return to room temperature, and remove the mold to obtain the gel sustained-release layer of the vaginal drug delivery device A; Vaginal drug delivery device B includes a plastic storage bottle, a bottle cap, and a drug delivery tube. The plastic storage bottle contains a functional powder dressing. The bottle mouth of the plastic storage bottle is provided with an external thread. The bottle cap and the drug delivery tube are both provided with an internal thread. The wall of the drug delivery tube is provided with a drug delivery hole. The preparation method of the functional powder dressing is: Step B1, weighing 100-200 parts by weight of bioactive glass, 500-700 parts by weight of isomaltooligosaccharide, and 100-200 parts by weight of propolis extract, placing them in a ball mill, and mixing and grinding them for 20-40 minutes to obtain a mixed powder; Step B2: weigh 5-20 parts by weight of mucosal repair factor and 1-5 parts by weight of vaginal probiotic freeze-dried powder, mix them with the mixed powder, and stir for 10-20 minutes to obtain the functional powder dressing of vaginal drug delivery device B. The method for preparing the alginate sulfate-carboxymethyl chitosan composite microemulsion in step A2 comprises the following steps: The first step is to prepare the oil phase by weighing 10-30 parts by weight of lithospermum officinale oil and 20-40 parts of camellia oil, and stirring them with a magnetic stirrer at 500 rpm for 10-20 minutes to obtain the oil phase; The second step is to prepare the aqueous phase by weighing 5-10 parts by weight of carboxymethyl chitosan and 10-20 parts by weight of alginate sulfate, adding them to 700-900 parts by weight of deionized water, and stirring them magnetically at a speed of 500 rpm for 50-70 minutes until the materials are completely dissolved to obtain the aqueous phase; The third step is to prepare a mixed surfactant by weighing 80-100 parts by weight of anhydrous ethanol and 80-100 parts by weight of polysorbate 80, and magnetically stirring at 500 rpm for 10-20 minutes to obtain a mixed surfactant; The fourth step is to prepare the oil phase surfactant mixture, mixing the above oil phase and mixed surfactant, and magnetically stirring at 500 rpm for 10-20 minutes to obtain the oil phase surfactant mixture; Step 5: The aqueous phase is magnetically stirred at 500 r / min for 10-20 min, and the stirring speed remains unchanged. The oil phase surfactant mixture is added dropwise during stirring. After the addition is completed, the mixture is stirred at the same speed for 20-40 min to obtain alginate sulfate-carboxymethyl chitosan composite microemulsion.

2. The device combination for treating female vaginal HPV infection according to claim 1, characterized in that: The preparation method of the anti-HPV egg yolk antibody recombinant human interferon α-2b composite nanoliposome in step A2 is The following steps are involved: The first step is to prepare PBS buffer with a pH value of 5-7; In the second step, 10-20 parts by weight of anti-HPV egg yolk antibody and 10-20 parts by weight of recombinant human α-2b interferon are dissolved in 400-500 parts by weight of PBS solution to be used as the aqueous phase; The third step is to weigh 300-400 parts by weight of ether and 400-600 parts by weight of chloroform, and stir them to form a mixed solution; Step 4: weigh 250-280 parts by weight of egg yolk lecithin and 20-50 parts by weight of cholesterol, add them to the mixed solution in step 3, slowly stir to dissolve them, and use them as the organic phase; Step 5: slowly inject the aqueous phase from step 2 into the organic phase from step 4, and stir ultrasonically using an ultrasonic stirrer to obtain a mixed liquid; Step 6: The mixed liquid in step 5 is subjected to vacuum rotary evaporation at 35-45° C. to remove ether and chloroform; In the seventh step, 3000-4000 parts by weight of PBS buffer and 100-200 parts by weight of Tween-60 are added to dissolve the material in the rotary evaporator in step 6, and ultrasonic stirring is continued to completely dissolve the material to obtain anti-HPV egg yolk antibody recombinant human α-2b interferon complex nanoliposomes.

3. The device combination for treating female vaginal HPV infection according to claim 1, characterized in that: The traction rod is made of a hard material, and the sealing tube is made of a soft material.

4. The device combination for treating female vaginal HPV infection according to claim 1, characterized in that: The plastic storage bottle is an elastic plastic storage bottle.

5. Use of the long-acting sustained-release device combination for treating female vaginal HPV infection as described in any one of claims 1 to 4 in the preparation of a medical device product for treating vaginal HPV infection.

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