The invention relates to an applicator comprising dissolvable microneedles, wherein the microneedles comprise hardened hyaluronic acid (HA) and trichloroacetic acid (TCA), to a process for preparing such an applicator, and to trichloroacetic acid for use in the treatment of warts, corns, or callus, wherein the use involves application of an applicator according to the invention onto warts, corns, callus, or skin tags.

Background art

Skin lesions such as warts, corns and calluses, are hardened areas on the skin. They are a form of hyperkeratosis, i.e. thickening of the outermost layer of the skin, often associated with the presence of abnormally high keratin levels. Warts are typically small, rough, hard growths that are similar in colour to the rest of the skin. Warts are caused by infection with a type of human papillomavirus (HPV). The virus is believed to enter the body through skin that has been damaged slightly. A number of types exist, including common warts, plantar warts, and filiform warts. Warts are very common, with most people being infected at some point in their lives. Without treatment, most types of warts resolve in months to years. In individuals who are otherwise healthy, they do not typically result in significant problems, although they may be painful on certain places of the body, such as on the bottom of the feet. Corns and calluses are thick, hardened layers of skin that develop when skin tries to protect itself against friction and pressure. They most often develop on the feet and toes, or hands and fingers. Corns are smaller than calluses and have a hard centre surrounded by inflamed skin. They tend to develop on feet parts that do not bear weight, such as the tops and sides of toes, and can be painful when pressed. Calluses are rarely painful and develop on the soles of feet, especially under the heels or balls and on hand palms. All of these skin lesions have in common that they are generally isolated patches of thickened skin and are cosmetically uninviting.

A common treatment for warts is cryotherapy, which involves freezing the wart, generally with liquid nitrogen, creating a blister between the wart and the epidermal layer after which the wart and the surrounding dead skin fall off. Although cryogenic kits have become available which can be used at home, treatment is usually performed by a medical doctor. A more commonly used at home therapy is keratolysis. This involves the peeling away of dead surface skin cells with keratolytic chemicals such as salicylic acid, formic acid, lactic acid, trichloroacetic acid, monochloroacetic acid, etc., which in varying concentrations are available in over-the-counter products. Treatment with these chemicals often has to take place by carefully applying a solution of the chemical onto the wart, thereby taking the necessary precautions to prevent application of the chemicals to healthy tissue. Treatment by this method often requires several months of (daily) treatment in order for the wart tissue to disappear. One reason for this long treatment required, is that the chemicals are only applied on top of the skin lesion, and cannot sufficiently enter into the core of the lesion (due to the hardened skin) for a more effective action.

More recently, microneedle patches have been developed. These consist of a patch with micrometer sized needles to be applied on the skin. By applying the patch onto the skin, the microneedles provide tiny punctures into the skin, which punctures provide a route for active ingredients to enter the skin for a more effective treatment. The active ingredients may be subsequently applied, or they may even be loaded into the microneedle itself. In the latter case, the microneedle may for example be fabricated from a hard biodegradable polymer which is mixed with the active ingredient. During application onto the skin, the microneedle degrades and releases the active ingredient. Such a microneedle patch is for example disclosed in IN201721017985A. In particular, IN201721017985A discloses a polymeric microneedle patch intended for the treatment of hyperkeratotic conditions like warts, corns, calluses, keloids, psoriasis, acne and eczema, comprising polymeric microneedles such as polyvinylchloride or cellulose and one or more keratolytic agents such as salicylic acid.

Description of the invention

It is an objective of the present invention to provide a more effective method for the at home treatment of warts and other skin lesions such as corns, calluses and skin tags or at least to provide a useful alternative. Thereto, the present invention provides an applicator comprising microneedles, preferably dissolvable microneedles, wherein the microneedles preferably comprise hardened hyaluronic acid (HA) and preferably a combination of hardened HA and a compound chosen from the group consisting of salicylic acid, formic acid, monochloroacetic acid (MCA), trichloroacetic acid (TCA), most preferably TCA. The term dissolvable indicates that the microneedles can degrade in a human body, i.e. the matrix material, e.g. polymer of the microneedle will degrade (dissolve) in the body by depolymerizing into monomers and/or oligomers which can be processed further by the body. The applicator is preferably used for the treatment of a wart, corn, callus, or skin tag. Thus, in an aspect, the invention provides for a compound chosen from the group consisting of salicylic acid, formic acid, monochloroacetic acid (MCA), trichloroacetic acid, preferably TCA for use in the treatment of a skin lesion such as a wart, corn, callus or skin tag, wherein the use involves application of an applicator comprising dissolvable microneedles preferably comprising hardened hyaluronic acid (HA) and a compound chosen from the group consisting of salicylic acid, formic acid, monochloroacetic acid (MCA), trichloroacetic acid (TCA), most preferably trichloroacetic acid (TCA), onto said skin lesion. The applicator of the invention is preferably a medical applicator, i.e. suitable for medical use. The present disclosure however also encompasses a two step method for treating said skin lesion, wherein in a first step the skin lesion is perforated by using an applicator comprising microneedles, e.g. metal microneedles or microneedles as taught herein, and wherein in a second step a compound chosen from the group consisting of salicylic acid, formic acid, monochloroacetic acid (MCA), trichloroacetic acid (TCA), preferably trichloroacetic acid (TCA), is applied onto said skin lesion, for example in an aqueous solution or gel.

In an embodiment, the use according to the present invention can be seen as cosmetic use.

Salicylic acid is often the agent of choice for treating warts. A less frequently used alternative is TCA. TCA is a powerful irritant that works by hydrolyzing the cellular proteins, which leads to inflammation and cell death. TCA causes destruction of warts by chemical coagulation of cellular proteins. As TCA is a stronger acid than salicylic acid, it may prefer safety precautions while applying the solution to prevent necrosis of tissue adjacent to warts.

Microneedling has become popular for the treatment of wrinkles. By treatment of the skin with microneedles, i.e. needles with a length of between 1 and 1000 pm, such as between 100 and 1000 pm, between 200 and 1000 pm, orbetween 300 and 1000 pm, skin cells are induced to produce collagen, which diminishes wrinkles. Furthermore, hyaluronic acid is a popular skin care ingredient as it increases the moisture content of the skin. When applied after microneedling, the compound chosen from the group consisting of salicylic acid, formic acid, monochloroacetic acid (MCA), trichloroacetic acid, preferably TCA can penetrate the skin more deeply for an enhanced moisturizing effect. Both measures can advantageously be combined by crosslinking hyaluronic acid to form hardened HA microneedles, i.e. to form solid HA microneedles. This advantageously combines the effects of microneedling with the improved moisture providing properties of hyaluronic acid provided more deeply into the skin. The microneedles can be applied by a patch, which is then left on the skin for a long enough time to allow biodegradation of the crosslinked hyaluronic acid in the skin, after which the patch may be removed. Use of such a patch is therefore very user friendly.

The present inventors have found that the combination of TCA with hyaluronic acid microneedles is surprisingly desirable for the treatment of skin lesions such as warts, corns and callus. A synergistic effect can be observed, as the effect of TCA in combination with the perforation by hyaluronic acid microneedles is greater than what can be expected based on a mere juxtaposition of both measures. This is explained in the experimental section.

The microneedles according to the present invention may be produced in a process of preparing an applicator, the process comprising: a) providing a microneedle template (mold), which comprises a cavity and a plurality of cavities for microneedles; b) loading of a (matrix) material, preferably comprising hyaluronic acid (e.g. in aqueous solution, e.g. between 0.1 and 0.5 g TCA 1 1 water), and optionally an active ingredient, into the cavity and cavities for microneedles of the microneedle template, wherein the active pharmaceutical ingredient is preferably TCA; c) optionally subjecting the loaded microneedle template to a centrifuging force (e.g. between 2000-5000 rpm for 1-10 minutes) such that the (matrix) material and the active ingredient fill up the microneedle cavities; d) drying of the filled-up microneedle template, optionally by applying heat and/or reduced pressure.

Alternatively, a process for producing the microneedles in a process for preparing an applicator may comprise the steps of: i. dissolving hyaluronic acid in a suitable solvent, preferably water, ii. pouring the mixture in a suitable complimentary microneedle mold and optionally applying a reduced pressure and/or heat until the microneedles are dry, thereby forming a microneedle array, iii. removing the microneedle array from the mold, iv. optionally cutting the microneedle array into a desired shape and size, v. providing application means to the array, wherein in step i) TCA is also dissolved in the solvent (e.g. between 0.1 and 0.5 g TCA 1 1 water) and/or after step iii) or iv) the microneedles are at least partially coated with (an aqueous solution or gel comprising) TCA.

The microneedle mold may for example be prepared as disclosed in LIS2018078498. To make a mold for casting the microneedles, a template is initially prepared from a suitable material such as silicon or a metal such as steel or titanium or a polymer such as polycarbonate or polymethacrylic acid. The template comprises a plurality of microneedles which have a size and shape corresponding to the desired size and shape of the microneedles of the applicator, i.e. typically either conical or pyramidal with a tapering tip. The mold may then be made by casting a liquid polymer material such as polydimethylsiloxane over the microneedle template. When the material is dried and cured the mold comprises microdepressions that retain the negative shape of the microneedles on which it is cast.

The length and number of microdepressions (microcavities) in the mold determine the length and number of the microneedles in the final applicator. The number of microneedles per unit area may vary widely, typically between 2 and 100 microneedles per cm ² , but is more often in the range of 5-75 microneedles, more usually 10-50 microneedles, preferably 15-30 microneedles such as 20-25 microneedles per cm ² .

For example, the microneedle array to be applied may be between 0.5 - 2.5 cm ² . Then the number of microneedles would typically be between 1 - 250. If the microneedle array to be applied is between 1 - 2 cm ² , then the number of microneedles would typically be between 2 and 200.

In total, each applicator may contain from 2 to 1000 microneedles, preferably from 400 to 800 microneedles, or from 25 to 250 microneedles.

Similarly, the length of the microneedles may vary, but they should preferably be of sufficient length to penetrate into the skin lesion. The microneedles may therefore have a length of 100-1500 pm, or from 100-500 pm, or from 300-1000 pm, e.g. 500-700 pm, or about 300 or 600 pm. Such a length of the microneedles generally ensures that the microneedle penetrates the skin lesion deep enough for the active ingredients to be optimally effective. The shape of the microneedles is also determined by the shape of the microdepressions in the mold and may be conical or pyramidal in shape and with a sharp tip. The microneedles may comprise longitudinally extending ridges to facilitate the insertion of the microneedles into the skin.

The aspect ratio of the microneedle (the length to width at the base of the microneedle) may vary according to the method used to produce the mold. A typical aspect ratio is 3:2, i.e. the length is 1.5 times greater than the width at the base. Another aspect ratio is 10:1. Preferably the aspect ratio is between 3:2 and 10:1. Preferably, the microneedles have a base width of between 70 - 150 pm, preferably of 80 - 120 pm, such as about 100 pm.

Microneedle arrays according to the invention may then be produced by filling the microdepressions in the mould with mixed hyaluronic acid and solvent, and optionally TCA. The solvent may be any suitable solvent for dissolving hyaluronic acid and optionally TCA, i.e. when both HA and TCA are to be dissolved in step i), then the solvent preferably should dissolve both HA and TCA, or if only HA is to be dissolved, then the solvent is only required to dissolve HA. A suitable solvent is water. The solvent is then removed, e.g. by drying in a desiccator under vacuum and/or in a drying oven at an appropriate temperature, e.g. at room temperature or above 30, 40, 50, 60, 80, 100 °C.

The dried microneedles may then be removed from the mold. This may for example be done by applying removal means such as adhesive layer on top of the mold and applying pressure to ensure good contact between the adhesive layer and the base of the microneedles followed by pulling the microneedles out of the mold. If the adhesive layer is medical tape, and the tape is wider than the size of the array in at least two opposite directions, then steps iii. and iv. may be combined, such that the tape that is used for removal at the same time forms the application means. The parts of the tape that are wider than the array may then be used for sticking the array the site of application, e.g. on the body.

If the mold was made in a larger size, individual arrays of an appropriate size may be prepared by cutting the array into pieces of a desired shape and size. Application means may then further be applied. For example, the array of microneedles may be applied to an application device, such as any suitable means for applying the array to the site of interest, e.g. a patch (flexible sheet) preferably comprising an adhesive layer, a (plastic) handle, cartridge, or a housing having a substantially planar application side (comprising the array) and an opposed top side, i.e. allowing to apply the array to a site of interest, e.g. by a user.

Finally, a suitable backing layer or membrane may be applied to protect the microneedle array, and optionally or alternatively, e.g. in the case where the applicator is a patch, said backing layer may protect the sticky part of the patch in order to retain adhesive properties until use of the patch.

In an embodiment, the microneedles may be impregnated with TCA, i.e. comprise TCA. In this case, TCA is suitably present in step i. of the process according to the invention, i.e. in step i) TCA is also dissolved in the solvent. In the process, TCA is then preferably present in the solution in a ratio of between 30:70 w:w and 70:30 w:w, between 40:60 w:w and 60:40 w:w, or 10:90 w:w with respect to hyaluronic acid (i.e. TCA 10, HA 90)

Additionally or alternatively, in step iii) the microneedles are at least partially coated with a liquid (e.g. aqueous solution) or gel comprising TCA. Thus, preferably, the microneedles are at least partially coated with a liquid (e.g. aqueous solution) or gel comprising TCA. The liquid (e.g. aqueous solution) or gel may be applied e.g. by immersion. The TCA concentration in the liquid (e.g. aqueous solution) or gel is preferably between 30-80 w/v%, 40-60 w/v% or 40 - 80 w/v%, more preferably between 50 - 80 w/v%. Higher concentrations are more desirable for treating the skin lesions than lower concentrations. . The entire outer surface of the microneedles may be coated with the liquid, e.g. aqueous solution or gel. Alternatively, only the tip of the microneedles may be coated, e.g. up to 10%, 20%, 30%, 40% or 50% of the length of the microneedles may be coated.

As according to the present disclosure, instead of TCA as used herein, a compound chosen from the group consisting of salicylic acid, formic acid, monochloroacetic acid (MCA) may be used. Said compound, or preferably TCA, may be applied in an amount of for example between 0.01 to 0.1 mg TCA per microneedle Further, instead of the use of hyaluronic acid as taught herein, any C1-C14 organic acid that is suitable for forming hardened microneedles may be used, for example caprylic acid. Preferably the applicator according to the present invention is applied in one or more cycles, wherein a cycle consists of daily application for 3 or 4 days, followed by a rest period of 3 or 4 days. Most preferable a cycle consists of daily application for 4 days followed by a rest period of 3 days.

Brief description of the figures

Fig. 1a is a side view of a mold filled with HA microneedles

Fig. 1b is a side view of a mold filled with HA microneedles and tape

Fig. 1c is a side view of the tape with HA microneedles

Fig. 1d is a side view of tape with microneedles and backing membrane

Fig. 1e is a bottom view of a patch

Fig. 1f is a bottom view of another embodiment of a patch

Fig. 2a is a side view of a cut microneedle array with medical tape

Fig. 2b is a side view of a cut microneedle array with medical tape and backing membrane

Fig 2c is a bottom view of an embodiment of a patch with medical tape.

Detailed description of the figures

The invention will now be further explained with reference to the figures. Fig.1a discloses a mold 1 filled with HA microneedles 2. In Fig. 1b removal means 3, e.g. tape, has been applied to the mold 1 for removing the microneedles. The tape sticks to the microneedle array, i.e. to the base portions of the microneedles, with portion 4. Furthermore, the tape is wider than the array and the extending portion 8 may later be used to stick the microneedle array to the body. Fig. 1d further shows backing membrane 7 for protecting the microneedles and the extending portion 8 (drawings are not to scale). Fig 1e is a bottom view of the patch of Fig 1c. If the adhesive tape is suitable for application to the body, such as medical tape, and the tape extends from the array in all directions, then the tape that is used for removal of the array from the mold at the same time forms the applicating means. The extending portion 8 of the tape is then used for sticking the array to the body. Fig 1f is an alternative embodiment of a patch. Such a patch may for example be formed if the adhesive tape extends from the array in only two opposite directions.

Alternatively, individual arrays of an appropriate size may be prepared by cutting the removal means 3 with the microneedle array into pieces of a desired shape and size. Applicating means 5, which may for example medical tape, may then further be applied, see Fig. 2a. In Fig 2b, a backing membrane 7 has been applied. Fig 2c shows a bottom view of the embodiments of Figs. 2a and 2b, with sticking portion 6 of application means 5 for adhesion to the body. Example

Microneedle patches were prepared by using a microneedle template (mold) with a larger cavity and a plurality of cavities for microneedles, loading of a (matrix) material and an active ingredient into the cavity and cavities for microneedles of the microneedle template, then subjecting the loaded microneedle template to a centrifuging force such that the (matrix) material and the active ingredient fill up the microneedle cavities, and finally drying of the filled-up microneedle template. The patches were used in a patient study.

Method of data collection: Data is collected from a number of patients with warts on the feet, with a maximum of 3 warts/patient. All the patients who or whose parents/legal guardians (in case of minors) are psychologically able to understand the study related information and to give an informed consent are included.

Detailed information on the subject age, sex, occupations and duration of the lesions are taken. History of trauma at the site of lesions are taken. Details of any previous treatment and also if on any immunosuppressive therapy are considered. The dimensions of the warts (size), and the exact location of the warts are recorded. Furthermore, a thorough systemic examination is done in each case to exclude the existence of any other disease along with the warts. Digital photography is taken at the initiation of the study. The below criteria for inclusion and exclusion are used.

Inclusion criteria are:

1. Patients with warts (plantar warts) on the feet.

2. Patient age above 8 years.

3. Both sexes.

4. Patients willing to come for regular follow up.

5. Patients with 3 or less than 3 warts.

6. Patients with at least one selected wart on the foot not older than 1 year and not previously treated and having a diameter of 3 to 6 mm.

Exclusion criteria are:

1. Age below 8 years.

2. Patients not willing to come for regular follow up.

3. Patients who are HIV positive or immunosuppressive due to other causes.

4. Patients with more than 3 warts.

5. Warts on other parts of the body. 6. Any other significant dermatological problem on the tested area, according to the investigator’s judgement.

7. Pregnancy, lactation and patient planning for pregnancy.

8. Patients with a known allergy to any of the product components.

9. Patients that have previously treated the wart.

The patients are divided into 6 groups, which are each subjected to a different treatment regimen. Each regimen is conducted for a maximum of 4 treatment cycles.

Regimen 1 :

A commercially available salicylic acid (SA) based anti-wart solution is applied daily. One week of treatment is equal to one cycle.

Regimen 2:

A commercially available TCA based anti-wart gel is applied twice a day for 4 days, followed by 4 days of rest, which is one cycle.

Regimen 3:

A patch with poly(lactic acid) microneedles coated with a 50% SA gel is applied 1 time a day for 4 days, followed by 4 days of rest. This cycle can be repeated up to max 4 times.

At application the patch is pressed for 3 minutes. The patch is allowed to stay on until next application or until it falls off.

Regimen 4:

A patch with poly(lactic acid) microneedles coated with a 50% TCA gel is applied 1 time a day for 4 days, followed by 4 days of rest. This cycle can be repeated up to max 4 times.

At application the patch is pressed for 3 minutes. The patch is allowed to stay on until next application or until it falls off.

Regimen 5:

A patch with hyaluronic acid microneedles coated with a 50% SA gel is applied 1 time a day for 4 days, followed by 4 days of rest. This cycle can be repeated up to max 4 times.

At application the patch is pressed for 3 minutes. The patch is allowed to stay on until next application or until it falls off. Regimen 6:

A patch with hyaluronic acid microneedles coated with a 50% TCA gel is applied 1 time a day for 4 days, followed by 4 days of rest. This cycle can be repeated up to max 4 times.

At application the patch is pressed for 3 minutes. The patch is allowed to stay on until next application or until it falls off.

Regimen 7:

A patch with hyaluronic acid microneedles coated with a 60% TCA gel is applied 1 time a day for 4 days, followed by 4 days of rest. This cycle can be repeated up to max 4 times.

At application the patch is pressed for 3 minutes. The patch is allowed to stay on until next application or until it falls off.

Regimen 8:

A patch with hyaluronic acid microneedles coated with a 50% TCA gel is applied 1 time a day for 4 days, followed by 3 days of rest. This cycle can be repeated up to max 4 times.

At application the patch is pressed for 3 minutes. The patch is allowed to stay on until next application or until it falls off.

Treatment is continued for the maximum period specified above or till the disappearance of the wart, whichever is earlier and it will be considered as a failure if there are no desirable results after the maximum number of treatment cycles have been finalized. Success of the treatment is judged by the disappearance or improvement of the wart tissue without any serious side effects.

All the patients are followed up at least after each treatment cycle. The response to therapy, the appearance of new lesions and the presence of secondary infections is noted, and the investigator decides if the treatment is continued, if multiple cycles are required. In case of secondary infection, the treatment is discontinued and treated for infection.

In patients who have unforeseen and serious side effects as decided by the investigator, treatment is discontinued and considered as failure. Patients in whom there is recurrence or failure of treatment will be subjected to other modalities of treatment.

On the day of final visit the patients’ opinion about quality of life based questions mentioned below related to the product are recorded, with a response of 4 equaling a high satisfaction and the lowest score being 1. Question/Score 1 2 3 4

A I was satisfied with the product B I would recommend this product

C I found this product easy to use

D I found this product effective

The results are summarized in a satisfaction score ranging from poor (1) to excellent (4).

Results of the study are summarized in Table 1 below.

TABLE 1.

Poor = no result

Fair = partial improvement after 4 cycles

Good = complete resolution after 4 cycles

Excellent = complete resolution after < 4 cycles

As can be seen from the results, commercially available products have the lowest performance, the resulting being only fair. Microneedles other than HA combined with either SA of TCA perform better, even as HA microneedles combined with SA. However, surprisingly the best results are achieved when HA microneedles are combined with TCA.

A higher concentration of TCA in the gel as well as an alternative treatment regimen with a shorter rest period both surprisingly result in a better patient satisfaction.

It is expected that the abovementioned putative effects can be effected in larger cohorts.