A randomized, double‐blinded, vehicle‐controlled clinical trial of topical cryosim‐1, a synthetic TRPM8 agonist, in prurigo nodularis

Prurigo nodularis (PN) is an intensively pruritic skin disease that negatively influences quality of life. Cryosim‐1 (Intrinsic IB Spot) is a synthetic, selective transient receptor potential melastatin 8 agonist.

anti-inflammatory drugs, and biologics, have been investigated for treating PN, none have been approved by the US Food and Drug Administration. 4,5ansient receptor potential melastatin 8 (TRPM8) channels, a member of the TRP family, detect and transduce cold stimuli in the somatosensory system. 6TRPM8 channels are found in a subset of unmyelinated C-type afferents located in dorsal root ganglion and trigeminal ganglion. 7,8In addition to cool temperatures (15-28°C), natural products or synthetic compounds such as menthol and icilin can activate TRPM8. 9Topical menthol-containing gel has been used for reducing itch and pain in skin disorders, but its irritancy and short duration of action limit its general application. 10A synthetic, selective TRPM8 agonist, cryosim-1 (Intrinsic IB Spot, 1-diisopropylphosphorylheptane; Dong Wha Pharmaceutical, Seoul, South Korea), showed effectiveness in treating eczema, urticaria, and localized scalp pruritus. 11,12Cryosim-1 has several advantages: the cooling sensation lasts for more than 2 h when 1.5% cryosim-1 gel is applied to the skin, adverse reactions have not been reported, and it penetrates and shows effectiveness in the thick skin of the scalp. 11,12Based on positive findings in previous studies, we hypothesized that topical cryosim-1 gel would reduce itching in PN patients since blocking the itch sensation and stopping the itch-scratch cycle is of utmost importance for the treatment of PN.Moreover, cryosim-1 gel can penetrate the thickened, hyperkeratotic nodules that are characteristic of PN lesions.
This study aimed to investigate the safety and antipruritic efficacy of topical cryosim-1 serum in patients with PN.

| Subjects and inclusion/exclusion criteria
We included patients diagnosed with PN at the Dermatology Department of the Asan Medical Center (Seoul, Korea) between July 2022 and November 2022.The inclusion criteria of this study were as follows: (i) adults aged 20 years and older, (ii) diagnosed by a dermatologist as having PN for at least 3 months, (iii) at least 10 pruritic nodules present, (iv) pruritic nodules present in at least two locations (both legs, both arms, trunk), and (v) participants agree to cooperate and follow the schedule of the clinical trial.All participants took part voluntarily and signed written informed consent after receiving a full explanation of the study design, risks, and benefits of the whole procedure.The exclusion criteria of this study were as follows: (i) patients with severe infection; (ii) pregnant or breastfeeding women; (iii) PN associated with the use of drugs such as opioids or angiotensin-converting enzyme inhibitors; and (iv) PN related to neurotic or psychotic disorders including neurotic excoriation, compulsive disorder, and delusional parasitosis.This randomized, double-blinded, placebo-controlled clinical trial was approved by the Institutional Review Board (IRB) of the Asan Medical Center (2021-1160).This study protocol also complied with the principles of the Declaration of Helsinki and with Korea's good clinical practice guidelines.

| Study design
At the screening visit (visit 1; day 1), participants were enrolled and provided with a screening number.All participants were randomly assigned to receive cryosim-1 serum or corresponding placebo serum, which was applied three times a day to the pruritic lesions for 8 weeks (from visit 1 to visit 3).Patients were asked to complete a questionnaire and to write a daily diary about their pruritic symptoms.The patients submitted their questionnaire and diary at visit 2 (day 28 ± 3) and visit 3 (day 56 ± 3).Measurement of transepidermal water loss (TEWL), a physical examination, and an evaluation of adverse reactions were conducted at every visit.Moreover, the numerical rating scale (NRS) of pruritus was evaluated before and 2 h after serum (cryosim-1 or placebo) application at every visit.
Concomitant medications permitted during the procedure included two types of second-generation antihistamines and mild (potency less than level 4) topical steroids.Participants were not allowed to change systemic or topical drugs during the study, and rescue treatments were not allowed.The use of concomitant treatment with immunosuppressants such as cyclosporine and methotrexate, systemic corticosteroids, moderate to strong (potency greater than or equal to level 4) topical steroids, and phototherapy was prohibited during the study.

| Efficacy measures
The primary efficacy measure was the reduction in the pruritus NRS score before and 2 h after application, as assessed by the participants.Secondary measures were the NRS scores of pruritus and sleep disorder determined at the 8 week follow-up visit by participants.The patients' satisfaction with the product properties (adhesive property, oily feeling, hydrating effect, hydration duration, absorption, comfortability at application, pruritus improvement, durability of pruritus improvement) was determined with an 11-point satisfaction rating scale (0, very unsatisfied; 10, very satisfied).

| Statistical analysis
All extracted data were analyzed using the Statistical Package for the Social Sciences (version 28.0; SPSS, Chicago, IL, USA), with p-values of <0.05 considered statistically significant.The Mann-Whitney test was used to compare subgroups in the treatment and placebo groups.

| Participant characteristics
Thirty-three participants were screened and randomized to treatment or placebo.Among them, three were lost to follow-up, while 30 ( 18treatment, 12 placebo) completed the 8 weeks of the study protocol.
The average absolute eosinophil count (AEC) was 244.8/mm 3 (range 37.0-824.5/mm 3) and 3.5% (range 0.5%-8.5%) of the total white blood cell.The average total immunoglobulin E level was 1000.4IU/mL (range 14.3-5000 IU/mL), and the average LDH level was 286 IU/L.Other abnormal changes were not observed in the routine blood tests.

| The efficacy of the TRPM8 agonist for relieving pruritus and sleep disorder
The baseline mean pruritus NRS scores were 6.7 ± 0.4 and 6.1 ± 0.7 in the treatment and placebo groups, respectively (p = 0.502).The mean pruritus NRS scores at week 4 before the application were 5.9 ± 0.4 and 6.1 ± 0.7 in the treatment and placebo groups, respectively (p = 0.747), while the mean pruritus NRS scores at week 4, 2 h after the application, were 3.6 ± 0.6 and 4.3 ± 0.7 in the treatment and placebo groups, respectively (p = 0.522).At week 8, the mean pruritus NRS score before the application was 4.7 ± 0.4 in the treatment group and 6.1 ± 0.5 in the placebo group, representing a statistically significant difference (p = 0.045).Moreover, at week 8, the mean pruritus NRS score 2 h after application was 2.8 ± 0.4 in the treatment group and 4.3 ± 0.5 in the placebo group (p = 0.031) (Figure 1).
The baseline mean NRS scores for sleep disorder were 6.4 ± 0.6 and 6.1 ± 0.9 in the treatment and placebo groups, respectively (p = 0.703).At week 4, the mean NRS scores for sleep disorder were 4.2 ± 0.7 and 4.1 ± 1.0 in the treatment and placebo groups, respectively.The NRS scores for sleep disorder decreased in both the treatment and placebo groups, and the differences were not significantly significant at week 4 (p = 0.983).At week 8, the mean NRS scores for sleep disorder were 2.2 ± 0.5 for the treatment group and 4.2 ± 0.8 for the placebo group; this result was statistically significant (p = 0.031) (Figure 2).

F I G U R E 1
The numeric rating scale (NRS) scores of pruritus at baseline, week 4 before serum application, week 4 after serum application (2 h), week 8 before serum application, and week 8 after serum application (2 h) according to the treatment group (cryosim-1 or placebo).

| Changes in TEWL with the TRPM8 agonist and placebo
The baseline mean TEWL scores were 22.8 ± 5.0 and 28.2 ± 5.6 for the treatment and placebo groups, respectively (p = 0.866).The mean TEWL scores at week 4 were 21.2 ± 4.6 for the treatment group and 27.3 ± 6.2 for the placebo group (p = 0.621), while those at week 8 were 18.6 ± 4.6 for the treatment group and 26.8 ± 5.3 for the placebo group (p = 0.401).The changes in TEWL at week 4 from the baseline were 1.7 ± 2.6 for the treatment group and 0.9 ± 1.1 for the placebo group (p = 0.401), while those at week 8 from baseline were 5.1 ± 4.1 for the treatment group and − 2.3 ± 3.4 for the placebo group (p = 0.125).

| Satisfaction with product properties and safety
The mean satisfaction scales for the adhesive property of the serum were 9.0 ± 0.3 for the treatment group and 8.5 ± 0.3 for the placebo group (p = 0.145), while those for oily feeling were 0.6 ± 0.2 for the treatment group and 2.0 ± 1.0 for the placebo group (p = 0.476) (Figure 3).The other mean satisfaction scales did not differ between the two groups and were as follows: hydrating effect, 4.8 ± 0.5 for the treatment group and 5.3 ± 0.8 for the placebo group (p = 0.343); hydration duration, 4.1 ± 0.4 for the treatment group and 3.9 ± 0.8 for the placebo group (p = 0.875); absorption, 8.3 ± 0.4 for the treatment group and 8.3 ± 0.5 for the placebo group (p = 0.820); and comfortability at application, 8.3 ± 0.5 for the treatment group and 9.2 ± 0.2 for the placebo group (p = 0.242).The mean satisfaction scales for pruritus improvement were statistically significantly different between the treatment group (7.2 ± 0.6) and the placebo group (4.0 ± 0.9; p = 0.005) (Figure 3).However, the mean satisfaction scales for the durability of pruritus improvement did not show statistical significance (3.5 ± 0.5 for the treatment group, 2.9 ± 0.6 for the placebo group; p = 0.811) (Figure 3).Both placebo and treatment were well tolerated, and no adverse reactions upon application were observed during the study period.

| Treatment response to TRPM8 agonist according to baseline eosinophil and total IgE count
When analyzing the baseline laboratory results within the treatment group, we found that the patients whose AEC was higher than 300/mm 3 had better response to TRPM8 agonist at week 4.The mean pruritus NRS scores at week 4 before the application were 5.8 ± 0.6 and 6.4 ± 0.6 in the high and low AEC groups, respectively (p = 0.632), while the mean pruritus NRS scores 2 h after the application, were 1.8 ± 0.9 and 5.4 ± 0.6 in the high and low AEC groups, respectively (p = 0.013).The mean sleeping disorder NRS scores at week 4 were 2.2 ± 0.8 and 6.0 ± 0.8 in the high and low AEC groups, The numeric rating scale (NRS) scores of sleep disorder at baseline, week 4, and week 8 according to the treatment group (cryosim-1 or placebo).
respectively (p = 0.011).The mean pruritus and sleeping disorder NRS scores at the screening visit and week 8 did not show significant differences baselined on AEC.The patients whose total IgE was higher than normal range (>100 IU/mL) suffered from severe sleeping disorder (7.8 ± 0.2 for high total IgE and 6.5 ± 1.9 for normal total IgE, p = 0.032) at the screening visit.However, other mean pruritus and sleeping disorder NRS scores did not show statistically significant differences between high total IgE and normal total IgE groups at week 4 and 8.

| DISCUSS ION
PN is known for its chronic and debilitating pruritus with a wide range of underlying dermatologic, psychiatric, and systemic comorbidities. 4,13stemic corticosteroids, immunosuppressants, thalidomide, gabapentin, and antidepressants have been used in refractory cases. 14A previous retrospective study of 325 patients with chronic PN showed that only immunosuppressants and gabapentinoids were successful therapeutic agents. 14However, prolonged maintenance treatment with these drugs is frequently limited due to side effects.Moreover, no targeted therapies have been approved for patients with PN, and biologics or small molecules are not suitable for general application due to variable interindividual responses and high costs. 15Since severe itching can last for several months to years in patients with PN, safe and effective agents are needed to manage the severe itching sensation.
In this randomized, double-blind, vehicle-controlled clinical trial, we demonstrated that cryosim-1, a TRPM8 agonist, was effective for the treatment of PN.This finding was demonstrated by the NRS scores of pruritus and sleep disorder evaluated by participants at week 8. Furthermore, cryosim-1 was safe as an emollient with similar properties to existing emollients.Interestingly, we observed that the mean NRS scores of pruritus between cryosim-1 and placebo at week 4 were not statistically different, either before or after application.Similarly, the differences in mean NRS scores of sleeping disorders were not statistically significant between cryosim-1 and placebo at week 4.However, there was a significant decrease between cryosim-1 and placebo groups for both scores at week 8.The NRS of sleep disorder was significantly lower in the cryosim-1 group than in the placebo group at week 8.We believe that since PN develops after prolonged scratching and pricking of the skin, resulting in hyperkeratotic lesions as well as nerve hyperplasia, at least 8 weeks are required to stop the itch-scratch cycle and normalize the skin structure.Moreover, when pruritus NRS scores were high, cryosim-1 could not effectively relieve pruritus since it is applied topically.This notion suggests that continuous application of cryosim-1 for at least 2 months is needed for the proper management of pruritus in PN patients.
We also found that the placebo, a simple emollient, reduced pruritus NRS scores from 6.1 to 4.3 after application (2 h) at weeks 4 and 8, but the placebo did change the baseline pruritus NRS scores before application at week 4 and week 8.This finding showed that F I G U R E 3 The patient satisfaction scales with an 11-point satisfaction rating scale (0, very unsatisfied; 10, very satisfied) of the product properties, including adhesive property, oily feeling, hydrating effect, hydration duration, absorption, comfortability at application, pruritus improvement, and durability of pruritus improvement, according to the treatment group (cryosim-1 or placebo).
the emollient, oral antihistamines, and low-potency steroids (which were allowed as concomitant medications in this study) were not effective at relieving itching sensation in PN patients.In addition, we observed that cryosim-1 had no effect on TEWL, and patents were satisfied with the product properties.These results are important since xerosis can aggravate pruritus in PN patients, especially in those with barrier defects such as atopic dermatitis patients or the elderly. 16Therefore, the use of hydrating agents or additional emollients was not required when cryosim-1 was applied.
Recent research shows that TRPM8 is expressed in skin, dorsal root ganglion sensory neurons, trigeminal ganglion sensory neurons, hippocampal neurons, adipocytes, and prostate cells [17][18][19] and thus likely has roles in other fields as well as dermatologic disease.Two previous randomized studies of cryosim-1 showed that it had an antipruritic effect 2 h after application in patients with urticaria, eczema, and scalp pruritus, and the itch scale significantly decreased after 1 week compared with the baseline in patients with urticaria and scalp pruritus. 11,12Moreover, thymol ameliorated pruritus and reversed upregulated proinflammatory cytokines by activating TRPM8, resulting in alleviation of psoriasis-like skin lesions in an imiquimod-induced mouse model of psoriasis. 20In addition, TRPM8 expression is upregulated in the skin and trigeminal ganglia of a mouse model of pruritus-accompanying LL37-induced rosacea and in the epidermis of skin biopsy samples from pruritus rosacea patients; these findings suggest TRPM8 is involved in rosacea pruritus. 21Interestingly, topical application of menthol to healthy volunteers activated TRPM8 and increased cutaneous blood flow not only at the site of application but also across the dermatome. 22 this context, topical activation of peripheral TRPM8 showed neuroprotective effects and mitigated ischemic stroke in mice. 23rthermore, TRPM8 is involved in energy homeostasis, and TRPM8 agonists could be potential novel agents for treating obesity. 24e mechanism of action of TRPM8 agonists in the treatment of PN is not yet known.The itch-scratch cycle is the most important underlying behavior of PN, and cryosim-1 might reduce repetitive scratching by providing instant relief of pruritus; the drug effect is known to occur within 10 min and might normalize the skin structure.
In addition, TRPM8 agonists might reduce the skin and serum levels of the cytokines interleukin (IL)-22, IL-1β, IL-6, IL-17, and IFN-γ.IL-22 is upregulated in PN, and IL-22 together with IL-17 promotes inflammation and increases keratinocyte proliferation. 25Moreover, IL-6 is a proinflammatory cytokine known to be associated with PN. 25 The main limitation of this study is that it was designed for an 8 week treatment duration; this period is not sufficient to determine if cryosim-1 application leads to prolonged and substantial reduction of itching sensation, leading to resolution of disease.Furthermore, we could not compare the efficacy of topical cryosim-1 with other topical agents such as topical steroids or topical calcineurin inhibitors.Lastly, we allowed the use of two antihistamines and low-potency topical steroids, thus we could not directly measure the efficacy of cryosim-1 alone.However, this study design reflected the real-world practice of treating PN, which is commonly treated with multiple agents, oral antihistamines, and low-potency steroids, which together are not usually associated with severe side effects.
In conclusion, this randomized, double-blinded, placebo-controlled clinical trial showed that cryosim-1 could be a safe and effective treatment for reducing pruritus and improving sleep quality for PN patients.