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Supplementary Figures S1-9 and Tables S1-3. Supplementary Figure S1 - Effects of knockdown of NUAK2 on melanoma cells and specificity of the p-AKT antibody. Supplementary Figure S2 - Immunohistochemical analyses of p-Akt and NUAK2 expression in clinical specimens. Supplementary Figure S3 - Kaplan-Meier curves for overall survival of Acral and Non-CSD melanoma patients. Supplementary Figure S4 - Knockdown of NUAK2 induces apoptosis in SM2-1 melanoma cells. Supplementary Figure S5 - Immunohistochemistry showing the expression of p21 following inhibition of the PI3K pathway by LY294002. Supplementary Figure S6 - Schematic diagram of the regulation of the cell cycle machinery by NUAK2 and PI3K pathways. Supplementary Figure S7 - Knockdown of CDK2 by siCDK2 in C32 and mel18 melanoma cells. Supplementary Figure S8 - Immunohistochemical analyses of the expression of CDK2 in clinical specimens. Supplementary Figure S9 - Expression of NUAK2, PTEN, p-Akt, Akt and CDK2, and cell proliferation in various melanoma cell lines treated with Roscovitine. Supplementary Table S1 - Clinical parameters and expressions of CDK2, p-Akt and NUAK2 of 56 acral melanomas and 35 non-CSD melanomas with survival information. Supplementary Table S2 - Genomic status of NUAK2 and PTEN gene in each cell line. Supplementary Table S3 - Expression of p27 and CDK2 in primary melanomas with or without over-expression of NUAK2 and p-Akt.

Summary Wheat-dependent exercise-induced anaphylaxis (WDEIA) is a life-threatening food allergy triggered by wheat in combination with the second factor such as exercise. The identification of potential genetic risk factors for this allergy might help high-risk individuals before consuming wheat-containing food. We aimed to identify genetic variants associated with WDEIA. A genome-wide association study was conducted in a discovery set of 77 individuals with WDEIA and 924 control subjects via three genetic models. The associations were confirmed in a replication set of 91 affected individuals and 435 control individuals. Summary statistics from the combined set were analyzed by meta-analysis with a random-effect model. In the discovery set, a locus on chromosome 6, rs9277630, was associated with WDEIA in the dominant model (OR = 3.95 [95% CI, 2.31−6.73], p = 7.87 × 10−8). The HLA-DPB1∗02:01:02 allele displayed the most significant association with WDEIA (OR = 4.51 [95% CI, 2.66−7.63], p = 2.28 × 10−9), as determined via HLA imputation following targeted sequencing. The association of the allele with WDEIA was confirmed in replication samples (OR = 3.82 [95% CI, 2.33−6.26], p = 3.03 × 10−8). A meta-analysis performed in the combined set revealed that the HLA-DPB1∗02:01:02 allele was significantly associated with an increased risk of WDEIA (OR = 4.13 [95% CI, 2.89−5.93], p = 1.06 × 10−14). Individuals carrying the HLA-DPB1∗02:01:02 allele have a significantly increased risk of WDEIA. Further validation of these findings in independent multiethnic cohorts is needed.

The prevalence of primary axillary hyperhidrosis in Japan is 5.75% (males, 6.60%; females, 4.72%) in the population aged 5–64 years. No study on comprehensively evaluated direct medical costs, hygiene product costs, and productivity loss in axillary hyperhidrosis patients has been published in Japan. The aim of this study was to estimate the cost of illness for axillary hyperhidrosis in Japan by conducting a nationwide insurance claims database analysis and a cross‐sectional Web‐based survey. Among patients diagnosed with primary axillary hyperhidrosis at least once between November 2012 and October 2019, health insurance receipt data of 1447 patients were analyzed. A cross‐sectional Web‐based survey was conducted on 321 patients aged 16–59 years with axillary hyperhidrosis to calculate hygiene product costs and productivity loss using a Work Productivity and Activity Impairment questionnaire. Furthermore, nationwide estimation was performed for the hygiene product costs and productivity loss based on the number of patients estimated from the prevalence. The annual direct medical costs per axillary hyperhidrosis patient were ¥91 491 in 2016, ¥93 155 in 2017, and ¥75 036 in 2018. In all of these years, botulinum toxin type A injection accounted for approximately 90% of the total costs. The annual total cost of hygiene products per axillary hyperhidrosis patient was ¥9325. The overall work impairment (%) of working patients with axillary hyperhidrosis was 30.52%, and its monthly productivity loss was ¥120 593/patient. The activity impairment (%) of full‐time housewives with axillary hyperhidrosis was 49.05% and its monthly productivity loss was ¥176 368/patient. The annual hygiene product cost based on the nationwide estimation was ¥24.5 billion and the monthly productivity loss was ¥312 billion. The significant cost associated with axillary hyperhidrosis was clarified. If out‐of‐pocket expenses for treatments not covered by health insurance are included in the estimation, the cost will further increase.

Scrofuloderma is one of the cutaneous manifestations of tuberculosis and usually occurs when underlying tuberculosis such as lymphadenitis directly involves the skin. However, the diagnosis of scrofuloderma without other apparent clinical manifestations of tuberculosis is sometimes challenging. A 27-year-old male from Bangladesh presented with a dome-shaped tumor on his right clavicle. MRI showed a high-density area that expanded from the lymph nodes between the internal jugular vein and the common carotid artery into the tumor on his right clavicle. Bacterial examinations of cultures from the tumor detected Mycobacterium tuberculosis. Further examinations for pulmonary tuberculosis including chest X-ray and computed tomography did not detect any lesions suggestive of pulmonary tuberculosis. From those examinations, the diagnosis of scrofuloderma was made. We emphasize the importance of recognizing a variety of clinical manifestations of tuberculosis including scrofuloderma due to the recent increase of immigrants from developing countries.

Sensitization to galactose-α-1,3-galactose (α-Gal) leads to the development of α-Gal syndrome, which includes red meat allergy and cetuximab-induced anaphylaxis. Since tick bites represent the main cause of α-Gal sensitization, it was speculated that sensitization to α-Gal occurs throughout Japan. However, few cohort studies have investigated α-Gal sensitization in Japan. Therefore, we aimed to elucidate the subclinical sensitization rate to α-Gal in Japan. Sera were obtained from 300 participants without food or cetuximab allergy at Shimane University Hospital (Shimane prefecture), Tokyo Medical and Dental University Hospital (Tokyo metropolis), and Tohoku University Hospital (Miyagi prefecture). ImmunoCAP-bovine thyroglobulin (BTG), ImmunoCAP-beef, and IgE immunoblotting with cetuximab were performed to detect α-Gal-specific IgE. Clinical information was collected from participants using a questionnaire. The overall positivity rate of ImmunoCAP-BTG was 4.0% without significant inter-institute differences, whereas that for ImmunoCAP-beef was 9.7% with a significant inter-institute difference. Tokyo Medical and Dental University Hospital (19.0%) had the highest positivity rate. The positivity rate based on cetuximab IgE immunoblotting was 2.7%, without any significant inter-institute differences. The overall positivity rate for both ImmunoCAP-BTG and cetuximab immunoblotting was 2.0%, with a significant inter-institute difference; 5.0% of Shimane University Hospital was the highest. Two cases showed sensitization against the non-α-Gal epitope of cetuximab. The overall positivity rate for both ImmunoCAP-beef and cetuximab immunoblotting was 1.3%, without significant inter-institute differences. Male sex was associated with positive beef-specific IgE. The prevalence of subclinical sensitization to α-Gal is estimated at 2.0%-4.0% in Japan and may be higher in rural areas, supporting an association between tick bites and α-Gal sensitization. In contrast, the prevalence of subclinical sensitization to beef is 9.7% in Japan and is highest in Tokyo Metropolis, suggesting the presence of another IgE-binding epitope apart from α-Gal and another sensitization route in the sensitization to beef IgE.

Interleukin (IL)-31 is a type 2 cytokine involved in the itch sensation in atopic dermatitis (AD). The cellular origins of IL-31 are generally considered to be Th2 cells. Macrophages are also a candidate as cellular sources of IL-31.To determine the expression of IL-31 by macrophages and to elucidate the productive mechanisms and contributions to itch in AD skin lesions.Expression of IL-31 by macrophages, expressions of thymic stromal lymphopoietin (TSLP) and periostin, and presence of infiltrating basophils in human AD lesions were examined through immunofluorescent staining, and correlations were assessed. Furthermore, mechanisms of inducing IL-31-expressing macrophages were analyzed in an MC903-induced murine model for AD in vivo and in mouse peritoneal macrophages ex vivo.A significant population of IL-31A network comprising IL-31-expressing macrophages, TSLP, periostin, and basophils plays a significant role in AD itch.

A phase 3 study was conducted to verify the efficacy and safety of 5% sofpironium bromide (BBI‐4000) gel (hereinafter referred to as sofpironium) administrated for 6 weeks in Japanese patients with primary axillary hyperhidrosis. The primary efficacy end‐point was the proportion of patients who satisfied both criteria of a Hyperhidrosis Disease Severity Score (HDSS) of 1 or 2 at the end of 6‐week treatment and a 50% or more reduction in total gravimetric weight of sweat at the end of treatment relative to baseline. A total of 281 patients were randomized to receive 5% sofpironium (141 patients) or vehicle (140 patients), and all patients were included in the full analysis set (FAS). In the FAS, 70.1% of patients were female, and the median age was 35.0 years. The proportion of patients who achieved the primary efficacy end‐point was 53.9% in the sofpironium group and 36.4% in the vehicle group, with a statistically significant difference of 17.5% (95% confidence interval, 6.02–28.93) between these two groups (P = 0.003). The incidence of adverse events was 44.0% in the sofpironium group and 30.7% in the vehicle group, and the incidence of adverse drug reactions was 16.3% in the sofpironium group and 5.0% in the vehicle group. Reported adverse events were generally mild or moderate in severity. In the sofpironium group, common events (incidence, ≥5%) were nasopharyngitis (14.2%) and dermatitis/erythema at the application site (8.5%/5.7%), with no serious adverse events reported. This study demonstrated the efficacy and safety of 5% sofpironium.

Optical coherence tomography (OCT) is a high-resolution tomographic imaging technique that uses optical interference. OCT has enabled the non-invasive three-dimensional analysis of individual acrosyringia in the stratum corneum in human skin. However, no report on the measurement of sweating by OCT using clinical data from humans has been published to date. Twenty patients with hyperhidrosis and twenty healthy subjects were included in this study. Imaging of acrosyringia in the stratum corneum using OCT and measurement of the sweat rate using the ventilated capsule method were performed simultaneously. The hand grip exercise of the right hand was used as a load to induce sweating, and the left fingertip was measured before and after the exercise load. Five acrosyringia were extracted from each OCT image, and their volumes were calculated. The mean volume of each acrosyringium was divided by the thickness of the stratum corneum to calculate the mean cross-sectional area of the acrosyringium. Furthermore, the number of sweat droplets on the skin surface was measured. The mean cross-sectional area of acrosyringia after the load increased both in patients with hyperhidrosis and in healthy subjects (P < 0.001). The mean cross-sectional area of acrosyringia of patients with hyperhidrosis was larger than that of healthy subjects (P < 0.001). The mean cross-sectional area of acrosyringia and the sweat rate showed a positive correlation before and after the load (r = 0.88 to 0.91). The number of droplets also increased after the load (P < 0.001), and the number of droplets in patients with hyperhidrosis was higher than in healthy subjects (P < 0.001). Our study has shown that acrosyringia in the stratum corneum increase in proportion to the sweat rate. OCT is a rigorous and valuable method that can measure and quantify sweating in the body without being an invasive procedure.

Acquired idiopathic generalized anhidrosis is a rare disease with unknown etiology. Sudden loss of sweating function adversely affects young patients’ quality of life. Although systemic corticosteroid therapy is the most frequently reported treatment for the disease, its effectiveness is controversial because of the risk of recurrence. To assist clinical decision‐making regarding whether to use steroids, we investigated the treatment responsiveness and recurrence rates in patients undergoing steroid pulse therapy and explored factors affecting these rates. We retrospectively collected data of 124 patients who received steroid pulse therapy to calculate the rate of responsiveness to the therapy. We also conducted a time‐to‐event analysis in a cohort of 57 patients who responded to steroid pulse therapy to estimate the recurrence rate after the therapy. As a result, the response and recurrence rates were 73% and 48%, respectively. Recurrence occurred within 1 year in most patients. The overall effectiveness of steroid pulse therapy was estimated to be 57% considering the recurrence rate. A delay from onset to treatment and younger age appeared to be negative factors for effectiveness. Moreover, we found a significant seasonal effect on both treatment and recurrence: autumn was the worst season for acquired idiopathic generalized anhidrosis in Japan. Our study revealed that steroid pulse therapy can be expected to be effective in half of treated patients. We recommend starting the therapy promptly after the diagnosis; however, it is also worth considering the season for treatment planning.

Background One of the hallmarks of bullous pemphigoid (BP) is moderate to severe chronic itch. Managing this is difficult because little is known about the mechanisms of itch in BP. Objective We sought to elucidate the pathophysiologic mechanisms of itch in BP. Methods The expression of itch mediators in lesions of 24 patients with BP and 6 healthy individuals were examined through immunofluorescence staining. Furthermore, the expression of itch mediators and itch severity was correlated. Results Itch severity was correlated with eosinophils, substance P, neurokinin 1R, interleukin (IL) 31 receptor A, oncostatin M receptor-β, IL-13, periostin, and basophils. There was also a trend between itch severity and IL-31 expression. Most of the cells expressing IL-31 or neurokinin 1R were identified as eosinophils. Intraepidermal nerve fiber density was decreased. Other itch mediators, including mast cells, IL-4, thymic stromal lymphopoietin, transient receptor potential vanilloid 1 and ankyrin 1, and protease activated receptor 2 were not significantly correlated with itch severity. Limitations The relatively small sample size, the examination of protein expression exclusively through immunofluorescent analysis, and lack of functional assays in patients are the limitations. Conclusions Multiple factors are involved in BP-associated itch, including eosinophils, substance P, neurokinin 1R, IL-31, IL-31 receptor A, oncostatin M receptor-β, IL-13, periostin, and basophils. They could be useful therapeutic targets.

Introduction Our previous clinical studies have demonstrated the short-term efficacy and safety of the sirolimus gel for patients with tuberous sclerosis complex (TSC). However, long-term clinical evidence is lacking. Our objective was to assess the safety and efficacy of long-term treatment with the sirolimus gel for the skin lesions of TSC patients. Methods We conducted a multicenter, open-label, uncontrolled clinical trial in 94 Japanese patients with TSC. Patients applied the 0.2% sirolimus gel on their face or head twice daily for > 52 weeks (maximum 136 weeks for safety). The safety endpoints were the rate of adverse event (AE)-caused discontinuation (primary endpoint) and the incidence of AEs. The efficacy endpoint was the response rate of angiofibromas, cephalic plaques, and hypomelanotic macules. Results Among 94 enrolled patients (mean age, 21 years; range 3–53 years), the rate of AE-caused discontinuation was 2.1% (2/94 patients). Although application site irritation and dry skin occurred relatively frequently, none of the drug-related AEs were serious; most of the drug-related AEs resolved rapidly. The major drug-related AEs (≥ 5% in incidence) were application site irritation (30.9%), dry skin (27.7%), acne (20.2%), eye irritation (8.5%), pruritus (8.5%), erythema (7.4%), dermatitis acneiform (6.4%), and dermatitis contact (5.3%). The response rates of angiofibromas, cephalic plaques, and hypomelanotic macules were 78.2% [95% confidence interval (CI) 68.0–86.3%], 66.7% (95% CI 51.1–80.0%), and 72.2% (95% CI 46.5–90.3%), respectively. Conclusions The gel was well tolerated for a long time by patients with TSC involving facial skin lesions and continued to be effective. Trial Registration ClinicalTrials.gov identifier: NCT02634931.

Background NUAK2 is a critical gene that participates in the carcinogenesis of various types of cancers including melanomas. However, the expression patterns of NUAK2 in normal skin and in various types of skin tumors have not been fully elucidated to date. Objectives To elucidate the distribution and localization of NUAK2 expression in normal skin, and characterize the expression patterns of NUAK2 and YAP in various types of skin tumors. Methods In this study, we characterized the expression of NUAK2 in tissues by developing a novel NUAK2-specific monoclonal antibody and using that to determine NUAK2 expression patterns in normal skin and in 155 cases of various types of skin tumors, including extramammary Paget’s disease (EMPD), squamous cell carcinoma (SCC), Bowen’s disease (BD), actinic keratosis (AK), basal cell carcinoma (BCC) and angiosarcoma (AS). Further, we analyzed the expression patterns of YAP and p-Akt in those tumors. Results Our analyses revealed that NUAK2 is expressed at high frequencies in EMPD, SCC, BD, AK, BCC and AS. The expression of p-Akt was positively correlated with tumor size in EMPD (P = 0.001). Importantly, the expression of NUAK2 was significantly correlated with YAP in SCC (P = 0.012) and in BD (P = 0.009). Conclusions Our results suggest that the YAP-NUAK2 axis has critical importance in the tumorigenesis of SCC and BD, and that therapeutic modalities targeting the YAP-NUAK2 axis may be an effective approach against skin tumors including SCC and BD.

Anhidrotic/hypohidrotic ectodermal dysplasia (A/HED) is a congenital disorder characterized by anhidrosis/hypohidrosis and inadequate hair and dental dysplasia. Large-scale case studies of patients with A/HED have already been conducted overseas, while there has been no large-scale study, but only a few case reports in Japan. Furthermore, an epidemiological study of this disease has not been conducted in Japan to date. The purpose of this study was to investigate the clinical characteristics of A/HED patients, the status of genetic aberrations and complications of A/HED in Japan. Initially, we conducted a physician-initiated questionnaire survey of A/HED patients who visited medical institutions across Japan to investigate their backgrounds, clinical symptoms, genotypes, diagnostic methods and complications of A/HED. We also investigated the presence or absence of various allergic diseases (atopic dermatitis-like skin manifestations, bronchial asthma and food allergies). Questionnaires were also obtained from 26 patients with ectodermal dysplasia (ED) who visited four medical institutions. We compared the incidence of allergic diseases in healthy controls in a similar study to that of patients. Twenty-four of those patients were considered to have A/HED, of which 18 had a confirmed genetic diagnosis and were genotyped. All patients had anhidrosis or hypohidrosis, hair and dental dysplasia, and unique facial appearance; 23 patients had several cutaneous manifestations and seven patients had periorbital pigmentation. In addition, there was a significantly higher incidence of atopic dermatitis-like cutaneous manifestations, bronchial asthma and food allergies in the A/HED patients than in healthy controls. We report the results from a questionnaire survey of 24 patients with A/HED. This is the first report of a large number of A/HED patients in Japan. This study clarifies the status of clinical diagnosis and genetic testing of A/HED patients in Japan, as well as the characteristics of their skin symptoms and allergic complications.

The mechanisms underlying itch are not fully understood. Physicians usually encounter difficulty controlling itch in generalized pruritus. Since only a small percentage of patients with generalized pruritus respond to antihistamines (H1 receptor antagonists), a variety of itch mediators and mechanisms other than histaminergic signals are considered to be involved in itch for these non-responsive patients. In 2012, we created guidelines for generalized pruritus. Those guidelines have been updated and revised to make some of the definitions, diagnostic terms, and classifications more applicable to daily clinical practice. Cutaneous pruritus as designated in these guidelines is a disease characterized by itch without an observable rash. Generalized pruritus (without skin inflammation) is defined as the presence of itch over a wide area, and not localized to a specific part of the body. This entity includes idiopathic pruritus, pruritus in the elderly, symptomatic pruritus, pregnancy-associated pruritus, drug-induced pruritus, and psychogenic pruritus. Localized pruritus (without skin inflammation) represents fixed itch localized to a specific part of the body, and includes anogenital pruritus, scalp pruritus, notalgia paresthetica, and brachioradial pruritus. These guidelines outline the current concepts and specify the diagnostic methods/treatments for cutaneous pruritus.

Prurigo is a treatment-resistant skin disease characterized by multiple isolated papules/nodules that cause severe itch. Prurigo papules/nodules occur either as primary lesions or as secondary lesions due to persistent scratching. The fundamental concepts and classifications of prurigo have not been sufficiently established, and considerable confusion remains regarding this topic. Clinical guidelines for chronic prurigo in Japan were published in 2012 in an attempt to reduce confusion regarding the concepts of prurigo and to standardize laboratory tests and treatments. However, the diagnostic terms for prurigo and associated concepts have changed over time, and new forms of treatment are under development. We have, thus, updated and revised the guidelines to classify prurigo based on clinical forms and causes, and disease name classifications based on the clinical form have been further simplified, such as prurigo nodularis, prurigo chronica multiformis, and prurigo (not otherwise specified). Expressions for acute, subacute, and chronic forms are not used. These guidelines outline the current concepts and specify treatments for prurigo.

Recently, we found a novel function of the lactic acid bacterium Tetragenococcus halophilus derived from miso, a fermented soy paste, that induces interleukin (IL)-22 production in B cells preferentially. IL-22 plays a critical role in barrier functions in the gut and skin. We further screened other bacteria species, namely, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Weissella, Pediococcus, and Bacillus, in addition to Tetragenococcus and found that some of them possessed robust IL-22-inducible function in B cells in vitro. This process resulted in the augmented expression of activation markers CD86 and CD69 on B and T cells, respectively. However, these observations were not correlated with IL-22 production. We isolated Bacillus coagulans sc-09 from miso and determined it to be the best strain to induce robust IL-22 production in B cells. Furthermore, feeding B. coagulans sc-09 to mice augmented the barrier function of the skin regardless of gut microbiota.

Objectives Prurigo is a treatment‐resistant inflammatory disease of unknown etiology. Persistent and severe itch is a major and important clinical symptom, but pathological mechanisms and/or actual mediators for itch in prurigo remain to be defined. Methods We investigated blood levels of β‐endorphin, dynorphin A, and autotaxin in adult patients with prurigo (n = 18), including prurigo nodularis, prurigo chronica multiformis, and other forms. Patients with hemodialysis, hepatic dysfunction, and pregnancy were excluded. Immunofluorescence staining for β‐endorphin and autotaxin in lesional skin was also performed. In addition, β‐endorphin and autotaxin synthesis of cultured epidermal keratinocytes in response to several cytokine stimulation was assessed in vitro. Results Blood levels of β‐endorphin were higher in patients with prurigo than in control subjects (n = 20), while dynorphin A levels were comparable in the two groups. Blood autotaxin levels also were increased in patients with prurigo compared to control subjects. On the one hand, epidermal expression of β‐endorphin was increased in skin lesions of prurigo chronica multiformis, but not in prurigo nodularis. On the other hand, enhanced expression of autotaxin was observed in the epidermis in both prurigo nodularis and prurigo chronica multiformis. In vitro studies showed that β‐endorphin‐expressing epidermal keratinocytes increased following stimulation with TNF‐α, interleukin‐31, and/or IFN‐γ. Autotaxin was also detected in the cytoplasm of cultured epidermal keratinocytes. The fraction of autotaxin‐expressing cells increased when stimulated with TNF‐α and/or interleukin‐31. Conclusions Imbalance of opioid receptor signals and/or the autotaxin‐lysophosphatidic acid axis may contribute, at least in part, to the pathogenesis and intractable pruritus in some of the patients with prurigo.

Objectives Chronic prurigo is a reactive skin disease marked by multiple pruriginous lesions such as papules, nodules, and erythema. We previously showed that basophil infiltration into the pruriginous lesions and basophil activation in blood are frequently observed. Therefore, basophils may be involved in the pathogenesis of chronic prurigo. In this study, we examined the efficacy of anti‐immunoglobulin E (IgE) therapy with omalizumab, which blocks basophil activation, in patients with prurigo. Methods Seven patients with chronic prurigo (prurigo chronica multiformis [PCM]: five patients; prurigo nodularis [PN]: two patients) and one patient with prurigo subacuta (PS) were enrolled. The infiltration of basophils into skin lesions was assessed by immunohistochemical analyses using a monoclonal antibody. All patients were treated three times with 300 mg omalizumab every 4 weeks, regardless of serum levels of total IgE. Skin symptoms were assessed using our unique skin scores before and after treatment with omalizumab. The efficacy of the treatment was determined according to the reduction ratio of skin scores as follows: high (ratio > 70%), moderate (70% ≥ ratio > 40%), mild (40% ≥ ratio > 10%), or none (10% ≥ ratio). Results All five patients with PCM were improved: The efficacy was high for three patients, moderate for one patient, and mild for one patient. In addition, the pruriginous lesions were improved mildly in the patient with PS, but not in those with PN. Conclusions Anti‐IgE antibody therapy with omalizumab may be effective for PCM but not for the other types of chronic prurigo.