General anti-hpv compositions and methods of making same

Abstract

The application discloses a general anti-HPV composition and a preparation method thereof, relates to the technical field of anti-HPV compositions, and comprises curcumin, epigallocatechin gallate, resveratrol and tanshinone II A in a molar ratio of 4:1:12.5:(1-2), and is compounded with a gel base, a solubilizing and stabilizing component to obtain a topical gel. The four-component active composition has the effects of synergistically inhibiting HPV-related cell proliferation, regulating HPV early gene expression and related signal pathways, and has good local tissue compatibility, and can be used for local prevention and treatment of different types of HPV-related skin and mucous membrane lesions.

Description

Technical Field

[0001] This invention belongs to the field of anti-HPV formulations and applications, and relates to general anti-HPV compositions and their preparation methods. Background Technology

[0002] Human papillomavirus (HPV) is a class of DNA viruses with multiple genotypes. Different HPV types exhibit different tissue tropisms and clinical manifestations. Some low-risk cutaneous HPV types can infect the skin epithelium and form verrucous external lesions; for example, plantar warts are usually associated with HPV types 1, 2, 4, 27, and 57. Genital warts, which occur in the genital area, are usually associated with low-risk mucosal HPV types, especially HPV types 6 and 11. In addition, persistent high-risk HPV infection can also involve mucosal sites such as the cervix and is associated with abnormal cervical epithelial lesions. Therefore, HPV infection is not limited to a single human site or a single clinical manifestation, but can manifest in different forms, such as cutaneous warts, external genital warts, and genital mucosal lesions, depending on the viral type and the infected tissue.

[0003] Existing HPV topical intervention products are typically formulated for specific infection sites or specific user populations. For example, topical formulations for the female reproductive tract mucosa are usually in the form of vaginal effervescent tablets, suppositories, or vaginal gels to meet the local drug delivery needs of the vagina or cervix; while for external lesions on the male external genitalia or skin surface, creams, ointments, or other topical formulations are more commonly used. Different formulations differ in application methods, local retention, mucosal adaptability, and ease of use, making it difficult for formulations designed for the female reproductive tract mucosa to be directly applied to the male external genitalia or other skin surfaces, while formulations designed for skin application may not be suitable for topical use on the reproductive tract mucosa.

[0004] On the other hand, the different infection sites and external manifestations associated with different HPV types mean that existing topical anti-HPV agents are often developed for single lesion sites or single application scenarios. This is especially true for external lesions such as plantar warts caused by cutaneous HPV, genital lesions caused by low-risk mucosal HPV, and genital mucosal lesions associated with persistent infection by high-risk HPV. Different local environments place different demands on the application, adhesion, retention, irritation, and release of active ingredients in these agents. Therefore, there is a practical need for universal topical agents that can remain stable and achieve effective contact in different local tissue environments.

[0005] Furthermore, curcumin, epigallocatechin gallate (EGCG), resveratrol, and tanshinone IIA possess different physicochemical properties, with some active ingredients exhibiting low water solubility, while EGCG demonstrates strong affinity for water. Directly mixing these multiple active ingredients to prepare a localized gel may present formulation challenges such as differences in the solubility of active ingredients, insufficient dispersion stability, and decreased uniformity of the gel system. Using only conventional mixing methods makes it difficult to simultaneously achieve stable dispersion of multiple active ingredients and satisfactory coating and adhesion properties of the final gel.

[0006] Therefore, existing technologies still require a universal topical formulation that can target different local lesions corresponding to different HPV types and is applicable to various local application scenarios such as skin, external genitalia, and genital tract mucosa. At the same time, this formulation also needs to solve the problem of stable dispersion and gelation of various anti-HPV active ingredients with different physicochemical properties in the gel system, so that the resulting formulation can take into account the combined effect of active ingredients, formulation uniformity, local application, adhesion retention, and local tissue compatibility. Summary of the Invention

[0007] The present invention aims to provide a universal anti-HPV topical gel composition and its preparation method, in order to solve the problem that existing anti-HPV topical formulations are usually designed for a single infection site or a single local application scenario, and it is difficult to meet the different local application needs of different types of HPV-related verrucous lesions, low-risk mucosal HPV-related local infections or lesions, and high-risk mucosal HPV-related epithelial abnormalities.

[0008] Meanwhile, given the differences in the physicochemical properties of curcumin or curcumin-like active components, epigallocatechin gallate, resveratrol, and tanshinone IIA, problems with dissolution, dispersion, and formulation uniformity can easily arise when multiple active components are directly mixed. Therefore, this invention further provides a gel carrier and preparation method adapted to the aforementioned quaternary active combination to improve the dispersion compatibility of multiple active components in a local gel system.

[0009] To achieve the above objectives, the present invention provides a universal anti-HPV topical gel composition, comprising a quaternary active composition, a gel matrix, a solubilizing and stabilizing system, a moisturizing system, and water.

[0010] The quaternary active composition comprises curcumin or curcumin-like active components, epigallocatechin gallate, resveratrol, and tanshinone IIA. With the molar amount of epigallocatechin gallate as 1, the molar ratio of curcumin or curcumin-like active components, epigallocatechin gallate, resveratrol, and tanshinone IIA is 4:1:12.5:(1-2).

[0011] The molar ratio described in this invention is calculated based on the actual effective content of each active component; for raw materials with insufficient purity, the actual amount of feed is calculated based on the actual content of the corresponding active ingredient in the raw material, and excipients are not included in the molar ratio of the quaternary active composition.

[0012] In a preferred embodiment, the molar ratio of the quaternary active composition is 4:1:12.5:1 or 4:1:12.5:2.

[0013] This invention, through screening of quaternary combinations, discovered that both of the aforementioned ratios can produce synergistic cell proliferation inhibition. Specifically, the combination index of the 4:1:12.5:1 combination is approximately 0.84, and the combination index of the 4:1:12.5:2 combination is approximately 0.34. The cell survival rate after treatment with the 4:1:12.5:2 combination was 36.10%, and the proliferation inhibition rate was 63.90%, indicating that the specific quaternary molar composition is not a simple superposition of the active ingredients, but rather possesses a further synergistic effect.

[0014] The gel matrix is ​​used to form a gel carrier suitable for topical application. Preferably, the gel matrix comprises one or more of carbomer, xanthan gum, or cellulose derivatives.

[0015] The solubilizing and stabilizing system is used to improve the dispersion compatibility of the quaternary active components in the formulation system, and preferably includes one or more of hydroxypropyl cyclodextrin, propylene glycol, 1,2-hexanediol, 1,2-pentanediol and PEG / PPG-17 / 6 copolymer.

[0016] The composition may further include a moisturizing system, which includes one or more of glycerin and butylene glycol.

[0017] Furthermore, the composition may also include plant-derived auxiliary components, including one or more of Centella asiatica extract, Sophora flavescens root extract, Cnidium monnieri extract, Taraxacum mongolicum extract, and Quercus acutissima root extract.

[0018] It should be noted that the quaternary active composition and the plant-derived auxiliary components are measured separately; the amount of plant extracts added is not converted into the amount of the corresponding monomeric active ingredient. In particular, matrine and matrine root extract are measured and added as different components.

[0019] The composition of this invention uses a topical gel as the main formulation, which can form a gel carrier suitable for use in different local tissue environments. Depending on the site of application, it can be used to prepare topical formulations for cutaneous HPV-related local skin lesions, low-risk mucosal HPV-related local infections or lesions, and high-risk mucosal HPV-related epithelial abnormalities.

[0020] Specifically, for applications related to cutaneous HPV, the composition can be used for local skin lesions such as verrucous lesions; for applications related to low-risk mucosal HPV, it can be used for local sites such as the external genitalia, vagina, cervix, and perianal area; and for applications related to high-risk mucosal HPV, it can be used in the preparation of local preparations for persistent HPV infection-related abnormal proliferation and cervical intraepithelial neoplasia.

[0021] The present invention also provides a method for preparing the above-mentioned general anti-HPV topical gel composition.

[0022] First, the quaternary active composition is pretreated with a solubilizing and stabilizing system to form an active component dispersion system; the active component dispersion system is mixed with an aqueous system containing a gel matrix, a moisturizing system and water, and optional plant-derived auxiliary components are added; then the pH of the mixture is adjusted and defoaming treatment is performed to obtain the general anti-HPV topical gel composition.

[0023] In a preferred embodiment, the aqueous phase system is formed by mixing a portion of purified water with the gel matrix and fully hydrating it; the active component dispersion system is formed by adding the quaternary active composition to the solubilizing and stabilizing system after heating and stirring, and then ultrasonically dispersing it; and the plant-derived auxiliary components are added to the mixed system after dispersion and filtration.

[0024] Preferably, the temperature for pretreatment of the solubilizing and stabilizing system with the quaternary active composition is 55-65°C, followed by ultrasonic treatment for 3-5 minutes; the resulting active component dispersion system is cooled to 25-45°C and then mixed with the aqueous phase system; the pH of the resulting mixture is adjusted to 5.2-5.8 and degassed for 3-5 minutes under conditions of −0.05-−0.08 MPa.

[0025] Compared with the prior art, the present invention has at least the following beneficial effects:

[0026] 1. This invention employs curcumin or curcumin-like active components, epigallocatechin gallate, resveratrol, and tanshinone IIA to construct a quaternary active combination, and determines a specific molar relationship of 4:1:12.5:(1-2) through combination screening. Experimental results show that both 4:1:12.5:1 and 4:1:12.5:2 exhibit synergistic proliferation inhibition effects, with the combination index of the 4:1:12.5:2 combination being approximately 0.34, indicating a strong synergistic effect.

[0027] 2. This invention combines the above-mentioned quaternary active combination with a local gel system. By pretreating the active components with different physicochemical properties through a solubilizing and stabilizing system, and then combining them with the gel matrix, it is beneficial to form a uniform local gel formulation, thereby improving the dispersion and compatibility problems that may occur when directly mixing multi-component active substances.

[0028] 3. This invention uses a local gel as the main formulation form, enabling the quaternary active combination to directly contact the target skin or mucous membrane tissue through local formulation, and allowing for formulation design according to different local application environments.

[0029] 4. This invention is not limited to a single HPV type or a single anatomical site, but rather constructs a local application system covering cutaneous HPV-related verrucous lesions, low-risk mucosal HPV-related local infections or lesions, and high-risk mucosal HPV-related epithelial abnormalities, thereby expanding the applicable scenarios of the same local gel technology platform. Attached Figure Description

[0030] Figure 1 The results show the synergistic effect of the quaternary active composition and the inhibition of HeLa cells;

[0031] Figure 2 The dose-dependent inhibition results of W12 cells in different HPV16 states;

[0032] Figure 3 Changes in the expression of early HPV genes E6 / E7 and p53;

[0033] Figure 4 For transcriptome differential gene and pathway enrichment analysis;

[0034] Figure 5 This is a schematic diagram of the multi-target mechanism of action of the quaternary active composition;

[0035] Figure 6 For EpiVaginal 3D vaginal tissue safety / histological results;

[0036] Figure 7 Results of the evaluation of vaginal mucosal irritation in rabbits;

[0037] Figure 8 Photos of actual wart lesions on the feet and before-and-after comparisons of use;

[0038] Figure 9 The study included the percentage of HPV test results turning negative and the rate of CIN I / LSIL lesion regression in 16 clinical application cases. Detailed Implementation

[0039] The present invention will be further described below through specific embodiments. To make the inventive objectives, technical solutions, and beneficial technical effects of the present invention clearer, the present invention will be further described in detail below with reference to the embodiments. It should be understood that the embodiments described in this specification are merely for explaining the present invention and are not intended to limit the present invention.

[0040] Unless otherwise stated, all films and reagents used in the examples are commercially available or synthesized using conventional methods and can be used directly without further processing, as are the instruments used in the examples.

[0041] Experimental reagents and consumables:

[0042] All solvents and reagents were reagent grade; water was distilled and deionized. Tanshinone IIA was purchased from Sigma (St. Louis, Missouri) and dissolved in 100% ethanol; resveratrol (99.0%; Jiaherb) and EGCG (95.3%; Taiyo Green Power Co., Ltd) were purchased from Thermo Fisher Scientific; curcumin (curcumin content ≥98%) (CAS No. 458-37-7) (Thermo Fisher Scientific, New Jersey, USA; prepared by Acros Organics for Thermo Fisher Scientific; stored at room temperature under nitrogen).

[0043] The purity of each active ingredient was determined according to the product label and quality certification documents of the actual purchased batch, and the molar ratio was calculated according to the effective content. Tanshinone IIA was dissolved in anhydrous ethanol for later use.

[0044] HeLa human cervical cancer cells were purchased from the American Type Culture Collection (ATCC) for evaluating the in vitro antiproliferative and synergistic effects of the quaternary active ingredients. The W12 series of HPV16-related cell models, including W12-20861, W12-20862, W12-20822, and W12-20850, were used to evaluate the cell proliferation inhibition effects under different HPV16 states. W12-20862 was used for early HPV gene expression and transcriptome analysis. All cells were cultured at 37°C and 5% CO2 according to their respective conditions. Cells in the logarithmic growth phase were used for experiments before the actual experiments.

[0045] Fetal bovine serum, DMEM medium, RPMI medium, penicillin-streptomycin solution, and other cell culture-related reagents were all purchased from reputable commercial suppliers.

[0046] The molar ratios of the quaternary active components are calculated based on the actual effective ingredient content and corresponding molecular weight of each raw material. Excipients are not included in the molar ratio of the quaternary active components. When the purity of the raw materials is less than 100%, the amount of material added is reverse-corrected according to the actual content.

[0047] Example 1: Preparation of a 4:1:12.5:1 quaternary anti-HPV gel

[0048] Based on 100g of gel, take 78.73g of purified water, 5.00g of butylene glycol, 5.00g of glycerol, 0.50g of 1,2-hexanediol, 0.50g of carbomer, 0.50g of triethanolamine, 0.05g of 1,2-pentanediol, 0.10g of methylparaben, 0.09g of benzalkonium bromide, 1.40g of hydroxypropyl cyclodextrin, 0.10g of betaine, and 0.50g of glyceryl glucoside; the total amount of the quaternary active components is controlled at 0.80g, of which curcumin, EGCG, resveratrol, and tanshinone IIA are added in a molar ratio of 4:1:12.5:1, and the remainder is made up to 100g with purified water.

[0049] In preparation, 70%–75% of the total water volume was slowly added to carbomer at 300–500 rpm, stirred until fully hydrated, and allowed to stand to degas. Glycerin, butylene glycol, hydroxypropyl cyclodextrin, and part of the stabilizer were added, mixed well, and then EGCG was added to obtain phase A. The remaining solubilizer and stabilizer were mixed and stirred at 55–65°C. Curcumin, resveratrol, and tanshinone IIA were added sequentially, and the mixture was sonicated for 3–5 min to form a clear solution or a stable fine dispersion, thus obtaining phase B. Phase B was cooled to 25–45°C and then slowly added to phase A, mixed at medium to low shear, and then the remaining excipients were added. The pH was adjusted to 5.2–5.8 with a 10%–20% triethanolamine aqueous solution, and the system thickened to form a homogeneous gel. Finally, the remaining purified water was added to 100 g, and the mixture was degassed at −0.05–−0.08 MPa for 3–5 min, cooled to room temperature in the dark, and the anti-HPV gel was obtained, denoted as sample G1. The above preparation process is consistent with the phase separation, ultrasonication, phase merging, pH adjustment and degassing processes recorded in the original document.

[0050] Example 2: Preparation of a 4:1:12.5:2 quaternary anti-HPV gel

[0051] Similar to Example 1, except that curcumin, EGCG, resveratrol and tanshinone IIA were added in a molar ratio of 4:1:12.5:2, the total amount of the quaternary active components was still controlled at 0.80g, and the remaining components and preparation conditions were the same as in Example 1, resulting in an anti-HPV gel, denoted as sample G2.

[0052] This molar ratio is another preferred ratio of the present invention; 4:1:12.5:1 and 4:1:12.5:2 exhibit combination indices of approximately 0.84 and 0.34, respectively, with the latter exhibiting a stronger synergistic effect.

[0053] Example 3: Anti-HPV gels with different concentrations of quaternary active ingredients

[0054] Similar to Example 1, except that the total amount of the quaternary active components was adjusted to 0.60 g, the four active components were kept in the same molar ratio of 4:1:12.5:1, and the other components and preparation conditions remained unchanged, resulting in an anti-HPV gel, denoted as sample G3.

[0055] Example 4: Anti-HPV gel containing plant extracts

[0056] Similar to Example 1, except that: 1.20 g of Sophora flavescens root extract, 1.50 g of Cnidium monnieri extract, 1.00 g of Taraxacum mongolicum extract and 1.00 g of Centella asiatica extract were further added, and the amount of purified water was reduced accordingly to make the total mass reach 100 g; the plant extracts were filtered through a 100-200 mesh sieve and then added to the system, while the other components and preparation conditions remained unchanged, to obtain an anti-HPV gel containing plant extracts, which was designated as sample G4.

[0057] It should be noted that plant extracts are auxiliary components and are not included in the molar ratio of the quaternary active composition. Centella asiatica, Sophora flavescens, Cnidium monnieri, Taraxacum mongolicum, and Quercus acutissima root are explicitly listed as optional plant extracts.

[0058] Example 5: Anti-HPV gel under optimized process conditions

[0059] Similar to Example 2, except that: in step A, stirring was performed at 400 rpm; in step B, the water bath temperature was controlled at 60°C and the mixture was sonicated for 4 min; phase B was added to phase A at 35°C, and the pH of the system was adjusted to 5.5 using a 10%–20% triethanolamine aqueous solution; finally, the mixture was degassed at −0.06 MPa for 4 min, with the other conditions remaining unchanged, to obtain an anti-HPV gel, denoted as sample G5.

[0060] Example 6: Anti-HPV gels with different solubilization systems

[0061] Similar to Example 2, except that: hydroxypropyl cyclodextrin and a portion of the cosolvent were combined to form a composite cosolvent system, and the volume ratio of the cosolvent to the stabilizer was controlled at 2 to 3:1. The remaining components and preparation conditions remained unchanged, and an anti-HPV gel was obtained, which was denoted as sample G6.

[0062] Comparative Example 1: Single EGCG Gel

[0063] Similar to Example 1, except that curcumin, resveratrol and tanshinone IIA were not added, and only EGCG with the same total amount of the quaternary active components as in Example 1 was added. The other components and preparation conditions remained unchanged, and a single EGCG gel was obtained, which was denoted as sample D1.

[0064] Comparative Example 2: Quaternary gels with molar ratios other than those of this invention

[0065] Similar to Example 1, except that the molar ratio of curcumin, EGCG, resveratrol and tanshinone IIA was adjusted to 4:1:4:1, while the other components and preparation conditions remained unchanged, resulting in a control gel, denoted as sample D2.

[0066] Comparative Example 3: Quaternary gel without pre-dispersion of active ingredients

[0067] Similar to Example 2, except that curcumin, resveratrol and tanshinone IIA were added directly to phase A without pre-dispersion in a 55-65℃ solubilizing system and ultrasonic treatment, and the other conditions remained unchanged, resulting in a control gel, which was designated as sample D3.

[0068] Comparative Example 4: Quaternary gels without hydroxypropyl cyclodextrin

[0069] Similar to Example 2, except that hydroxypropyl cyclodextrin was not added, and the mixture was made up with an equal mass of purified water. The remaining components and preparation conditions remained unchanged, and a control gel was obtained, which was designated as sample D4.

[0070] Performance Experiment 1: Synergistic Proliferation Inhibition Effect of the Quaternary Active Composition

[0071] The synergistic effect of the quaternary active ingredients was evaluated using HeLa cells. Curcumin, EGCG, resveratrol, and tanshinone IIA were prepared at different molar ratios and co-cultured with HeLa cells for 96 h. Cell viability was assessed using the EZQUANT method, and the combination index (CI) was calculated according to the intermediate-effect principle. CI < 1 indicated a synergistic effect, with lower CI indicating a more pronounced synergistic effect.

[0072] 4:1:12.5:1 Approximately 0.84 Mild collaboration 4:1:12.5:2 Approximately 0.34 Strong synergy

[0073] The results showed that both preferred molar ratios exhibited synergistic effects, with the CI of 4:1:12.5:2 being approximately 0.34. At this ratio, cell viability was 36.10%, corresponding to a proliferation inhibition rate of 63.90%. This result directly supports the selection of specific molar ratios of the quaternary active components.

[0074] Performance Experiment 2: Inhibitory Effect on Cell Proliferation in Different HPV16 States

[0075] The W12 cell model containing different HPV16 genomic states was used for evaluation, including cell subclones in different integrated and free states. Cells were treated with a quaternary active composition at concentration gradients for 120 h, cell proliferation was detected, and dose-response curves were plotted.

[0076] The results showed that the quaternary active composition exhibited dose-dependent inhibitory effects on W12 cells with different HPV16 states. The IC50 values ​​of W12-20861, W12-20862, W12-20822 and W12-20850 were 0.9, 2.7, 3.9 and 8.2 μM, respectively, indicating that the composition can act on cells with different HPV16 genomic states.

[0077] Performance Experiment 3: Detection of HPV E6 / E7 and p53 Transcription Levels

[0078] Using W12-20862 cells as a model, the cells were treated with a 2 μM quaternary active composition for 0, 6 and 24 h, respectively. GAPDH was used as an internal control, and the expression levels of HPV-E6, HPV-E7 and p53 mRNA were detected by RT-qPCR.

[0079] The results showed that HPV-E6 and HPV-E7 expression levels decreased with prolonged treatment time, while p53 increased at 24 h, indicating that the quaternary combination could simultaneously affect HPV early oncogene-related transcription and host tumor suppressor pathways.

[0080] Performance Experiment 4: Transcriptome-level Pathway Analysis

[0081] RNA sequencing was performed using HPV16-positive W12-20862 cells. After treating the cells with a 2 μM quaternary active ingredient for 24 h, RNA was extracted and used for library construction, high-throughput sequencing, and pathway enrichment analysis.

[0082] The results showed that 1245 genes were downregulated, 1376 genes were upregulated, and the remaining 12457 genes did not reach the differential expression threshold. The differentially expressed genes were mainly enriched in pathways such as p53 signaling, cell cycle, apoptosis, DNA damage response, PI3K-Akt, NF-κB, MAPK, oxidative stress, inflammation, and cellular senescence, indicating that the quaternary combination exhibits multi-pathway synergistic regulatory characteristics.

[0083] Performance Experiment 5: Three-Dimensional Evaluation of Human Vaginal Tissue Compatibility

[0084] The local tissue compatibility of the gel containing the quaternary composition was evaluated using the EpiVaginal three-dimensional human vaginal epithelial tissue model. Negative and positive toxicity controls were set up. The formulation group was exposed to 2 μM of the quaternary active composition for 24 h. After treatment, H&E staining was performed, and the release of IL-1β and IL-8 and the integrity of the tissue barrier were detected.

[0085] The results showed that the epithelial structure of the quaternary compound formulation was intact and no obvious tissue damage was observed; IL-1β and IL-8 were not significantly increased and TEER did not decrease significantly, indicating that the gel did not show obvious tissue stimulation or barrier disruption in the simulated vaginal mucosal environment.

[0086] Performance Experiment 6: Evaluation of Local Stimulation in Rabbit Vagina

[0087] A gel containing a quaternary active composition was continuously applied topically to the rabbit vagina for 7 days, with a negative control included. After the experiment, vaginal tissue was collected for H&E staining to observe the integrity of the mucosal epithelium, tissue damage, and inflammatory cell infiltration, and the vaginal mucosal irritation index was calculated.

[0088] The results showed that no obvious mucosal irritation, epithelial damage or inflammatory cell infiltration was observed in the gel treatment group, and the vaginal mucosal irritation index was 0.00, which was judged to be non-irritating.

[0089] Performance Experiment 7: Improvement of HPV Negativity and CIN I in Clinical Observation

[0090] A case series was conducted on 16 patients with persistent HPV infection. Patients used a gel containing a quaternary active compound for 3 months, followed by a 3-month follow-up period after discontinuation. HPV genotyping was performed using reverse hybridization signal amplification technology, and cytological evaluation was conducted using liquid-based thin-layer cytology combined with the Bethesda system.

[0091] The results showed that, overall, 15 out of 16 cases achieved HPV seroconversion, with a seroconversion rate of 93.75%. Among them, 9 cases were positive for high-risk HPV, with 8 achieving seroconversion, resulting in a seroconversion rate of 88.9%; and 6 cases were positive for low-risk HPV, all of whom achieved seroconversion. The 8 cases with baseline CIN I / LSIL did not show any recurrence of CIN I at the 6-month follow-up. No serious adverse events were reported during the observation period. It should be noted that this observation was a small-sample, non-randomized, uncontrolled case series, and its results are used to illustrate the feasibility of local application and clinical signals, and cannot replace randomized controlled clinical trials.

[0092] Performance Experiment 8: Case Observation of Recurrent and Refractory Plantar Warts

[0093] This observational data was derived from clinical observation case records obtained by the applicant, and included two patients with recurrent and refractory plantar warts. Both patients had a history of recurrent episodes for more than 1.5 years and had not achieved sustained improvement after previous conventional treatment. Clinical examination and histopathological examination confirmed the diagnosis of plantar warts in both patients. Further HPV genotyping was performed; Case 1 tested positive for HPV16 and HPV33, while Case 2 tested positive for HPV6, HPV18, and HPV52.

[0094] Two patients used a topical formulation containing the quaternary active composition of this invention, wherein the molar ratio of curcumin, epigallocatechin gallate, resveratrol, and tanshinone IIA in the quaternary active composition was 4:1:12.5:1. The formulation was applied topically to the plantar wart lesions once daily for six consecutive months, and changes in lesions and local tolerance before and after treatment were recorded.

[0095] The results showed that after 6 months of continuous use, the plantar wart lesions in both patients were significantly improved compared to before treatment, with no obvious local irritation or other adverse reactions observed. Photos of the foot lesions before and after treatment are available below. Figure 8 .

[0096] Performance Experiment 9: Clinical Application Case Observation

[0097] This observational data comes from clinical observation cases obtained by the applicant, including 16 cases that received topical formulations containing the quaternary active composition of this invention. In the quaternary active composition, the molar ratio of curcumin, epigallocatechin gallate, resveratrol, and tanshinone IIA is 4:1:12.5:1. The cases used the formulation at the relevant local sites once daily for 3 consecutive months, and were followed up for another 3 months after discontinuation. HPV test results and changes in CIN I / LSIL lesions were recorded before treatment, during the 3-month treatment period, and during the follow-up period.

[0098] The results showed that among the 16 cases, 15 tested negative for HPV at 3 months, a negative rate of 93.75% (15 / 16). Of these, 8 out of 9 high-risk HPV infections tested negative, a negative rate of 88.9% (8 / 9); and all 6 low-risk HPV infections tested negative, a negative rate of 100% (6 / 6). Furthermore, of the 16 cases with baseline CIN I / LSIL, no corresponding lesions were detected at the 6-month follow-up, a lesion regression rate of 100% (8 / 8). No serious adverse events were reported during the observation period, and overall local tolerability was good. The above clinical application observation results are as follows: Figure 9 As shown.

[0099] The above results indicate that the quaternary active composition of the present invention showed good HPV test seroconversion and improvement of CIN I / LSIL lesions in clinical application case observations, which can further support its application value as a topical anti-HPV composition in different HPV infections and related lesions.

[0100] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A universal anti-HPV topical gel composition, characterized in that, It includes a quaternary active composition, a gel matrix, a solubilizing and stabilizing system, a moisturizing system, and water; The quaternary active composition comprises curcumin or curcumin-like active components, epigallocatechin gallate, resveratrol, and tanshinone IIA; with the molar amount of epigallocatechin gallate as 1, the molar ratio of curcumin or curcumin-like active components, epigallocatechin gallate, resveratrol, and tanshinone IIA is 4:1:12.5:(1-2). The molar ratio is calculated based on the actual effective content of each active component, and the gel matrix is ​​used to form a gel carrier suitable for local application.

2. The universal anti-HPV topical gel composition according to claim 1, characterized in that, The curcumin-based active components include one or more of curcumin monomer, demethoxycurcumin, and bisdemethoxycurcumin; the epigallocatechin gallate, resveratrol, and tanshinone IIA are respectively converted into molar amounts according to their actual effective ingredient content.

3. The universal anti-HPV topical gel composition according to claim 1, characterized in that, The gel matrix includes one or more of carbomer, xanthan gum, or cellulose derivatives; the solubilizing and stabilizing system includes one or more of hydroxypropyl cyclodextrin, propylene glycol, 1,2-hexanediol, 1,2-pentanediol, and polyether PEG / PPG-17 / 6 copolymer.

4. The universal anti-HPV topical gel composition according to claim 1, characterized in that, It also includes plant-derived auxiliary components, which include one or more of Centella asiatica extract, Sophora flavescens root extract, Cnidium monnieri extract, Taraxacum mongolicum extract, and Quercus acutissima root extract.

5. The universal anti-HPV topical gel composition according to claim 1, characterized in that, The composition is for topical application to cutaneous HPV-related local skin lesions, including verrucous lesions.

6. The universal anti-HPV topical gel composition according to claim 1, characterized in that, The composition is for topical application to low-risk mucosal HPV-related local infections or lesions, the application sites including one or more of the external genitalia, vagina, cervix, and perianal area.

7. The universal anti-HPV topical gel composition according to claim 1, characterized in that, The composition is used for the preparation of a topical formulation of high-risk mucosal HPV-related epithelial abnormalities, including HPV-related abnormal proliferation and / or cervical intraepithelial neoplasia.

8. A method for preparing the universal anti-HPV topical gel composition of claim 1, characterized in that, Includes the following steps: 1) The quaternary active composition is pretreated with a solubilizing and stabilizing system to obtain an active component dispersion system; 2) Mix the active ingredient dispersion system with an aqueous phase system comprising a gel matrix, a moisturizing system and water, and add optional plant-derived auxiliary components; 3) Adjust the pH of the resulting mixture and perform defoaming treatment to obtain a general anti-HPV topical gel composition.

9. The method according to claim 8, characterized in that, The aqueous phase system is formed by mixing a portion of water with the gel matrix and fully hydrating the gel matrix; the active component dispersion system is formed by adding the quaternary active composition after heating and stirring the solubilizing and stabilizing system, and then ultrasonically dispersing it; the plant-derived auxiliary component is dispersed, filtered, and then mixed with the aqueous phase system and the active component dispersion system.

10. The method according to claim 9, characterized in that, The temperature for pretreatment of the solubilizing and stabilizing system with the quaternary active composition is 55-65℃, and the ultrasonic treatment time is 3-5 min. The resulting active component dispersion system is mixed with the aqueous phase system at 25-45℃. The pH of the mixed system is adjusted to 5.2-5.8, and degassed at -0.05--0.08 MPa for 3-5 min.

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