Lipidome-based rapid diagnosis with machine learning for detection of TGF-ß signalling activated area in head and neck cancer.

BACKGROUND: Several pro-oncogenic signals, including transforming growth factor beta (TGF-ß) signalling from tumour microenvironment, generate intratumoural phenotypic heterogeneity and result in tumour progression and treatment failure. However, the precise diagnosis for tumour areas containing subclones with cytokine-induced malignant properties remains clinically challenging. METHODS: We established a rapid diagnostic system based on the combination of probe electrospray ionisation-mass spectrometry (PESI-MS) and machine learning without the aid of immunohistological and biochemical procedures to identify tumour areas with heterogeneous TGF-ß signalling status in head and neck squamous cell carcinoma (HNSCC). A total of 240 and 90 mass spectra were obtained from TGF-ß-unstimulated and -stimulated HNSCC cells, respectively, by PESI-MS and were used for the construction of a diagnostic system based on lipidome. RESULTS: This discriminant algorithm achieved 98.79% accuracy in discrimination of TGF-ß1-stimulated cells from untreated cells. In clinical human HNSCC tissues, this approach achieved determination of tumour areas with activated TGF-ß signalling as efficiently as a conventional histopathological assessment using phosphorylated-SMAD2 staining. Furthermore, several altered peaks on mass spectra were identified as phosphatidylcholine species in TGF-ß-stimulated HNSCC cells. CONCLUSIONS: This diagnostic system combined with PESI-MS and machine learning encourages us to clinically diagnose intratumoural phenotypic heterogeneity induced by TGF-ß.

Japanese guidelines for allergic rhinitis 2020.

Like asthma and atopic dermatitis, allergic rhinitis is an allergic disease, but of the three, it is the only type I allergic disease. Allergic rhinitis includes pollinosis, which is intractable and reduces quality of life (QOL) when it becomes severe. A guideline is needed to understand allergic rhinitis and to use this knowledge to develop a treatment plan. In Japan, the first guideline was prepared after a symposium held by the Japanese Society of Allergology in 1993. The current 8th edition was published in 2016, and is widely used today. To incorporate evidence based medicine (EBM) introduced from abroad, the most recent collection of evidence/literature was supplemented to the Practical Guideline for the Management of Allergic Rhinitis in Japan 2016. The revised guideline includes assessment of diagnosis/treatment and prescriptions for children and pregnant women, for broad clinical applications. An evidence-based step-by-step strategy for treatment is also described. In addition, the QOL concept and cost benefit analyses are also addressed. Along with Allergic Rhinitis and its Impact of Asthma (ARIA), this guideline is widely used for various clinical purposes, such as measures for patients with sinusitis, childhood allergic rhinitis, oral allergy syndrome, and anaphylaxis and for pregnant women. A Q&A section regarding allergic rhinitis in Japan was added to the end of this guideline.

Safety profile and immunological response of dual sublingual immunotherapy with house dust mite tablet and Japanese cedar pollen tablet.

BACKGROUND: There have been no studies of dual administration of sublingual immunotherapy (SLIT) tablets for perennial and seasonal allergic rhinitis. This trial (JapicCTI-184014) was conducted to investigate the safety profile and immunological response during dual therapy with SQ house dust mite (HDM) and Japanese cedar pollen (JCP) SLIT tablets. METHODS: This was a multicenter, open-label, randomized trial of 109 Japanese patients with coexisting HDM and JCP allergic rhinitis who had positive tests for HDM- and JCP specific IgE (≥0.7 kU/L). Patients were allocated to receive HDM (N = 54) or JCP (N = 55) SLIT tablets alone for 4 weeks followed by 8 weeks of dual therapy with both SLIT tablets administered within 5 min of each other. Adverse events (AEs), adverse drug reactions (ADRs), and serum IgE and IgG4 specific for HDM (Dermatophagoides farinae, Dermatophagoides pteronyssinus) and JCP were recorded. RESULTS: The percentage of subjects with AEs and ADRs was similar between the two groups and between the two periods of monotherapy and dual therapy. Most AEs and ADRs were mild in severity, and no serious events were observed. The most common ADRs were local events in the oral cavity. Levels of IgE and IgG4 specific for HDM (D. farinae, D. pteronyssinus) and JCP were increased after treatment with HDM and JCP SLIT tablets, respectively. CONCLUSIONS: Dual therapy with both SLIT tablets administered within 5 min after 4 weeks of monotherapy with HDM or JCP tablet was well tolerated and induced the expected immunological responses.

Efficacy of house dust mite sublingual tablet in the treatment of allergic rhinoconjunctivitis: A randomized trial in a pediatric population.

BACKGROUND: The efficacy and safety of 300 index of reactivity (IR) tablets of house dust mite (HDM) allergen extracts in Japanese pediatric (5-16 years old) patients with allergic rhinitis (AR) were assessed in a double-blind, randomized, placebo-controlled study (JAPIC CTI-152981). METHODS: Patients were randomized 1:1 to HDM sublingual tablets or placebo once daily for 52 weeks. The primary end-point was average adjusted symptom score (AASS; average of daily Rhinitis Total Symptom Scores, comprising sneezing, rhinorrhea, nasal congestion, and nasal pruritus, adjusted for rescue medication use), analyzed during Weeks 48-52 (mixed-effects model for repeated measures). RESULTS: Of 438 patients randomized, 403 (92%; 193 active, 210 placebo) completed the study. AASS (least-squares [LS] mean [SE]) during Weeks 48-52 was significantly (P = 0.0005) lower in the active group compared with placebo (6.32 [0.20] vs 7.27 [0.19]; relative LS mean difference, -13%). Immunological responses (IgE and IgG4 antibodies specific to antigens of two HDM species) were significantly greater in the active group compared with placebo (P < 0.0001). Almost all patients experienced mild or moderate adverse events (AEs). The most common treatment-related AEs were oral pruritus, mouth edema, throat irritation, and ear pruritus. One patient experienced serious pseudocroup (subglottic laryngitis) that recovered. There were no deaths or anaphylaxis requiring the use of injectable adrenaline. CONCLUSIONS: The HDM tablet significantly improved symptoms of HDM-induced perennial AR and was associated with a significant immunological response. The safety profile in pediatric patients was consistent with that in adults, with no new safety concerns.

Establishment of enzyme-linked immunosorbent facilitated antigen binding as a biomarker assay for Japanese cedar pollen allergy immunotherapy.

BACKGROUND: Clinical efficacy of allergen-specific Immunotherapy (AIT) towards Japanese cedar (JC) pollen allergy is firmly established but JC pollen-specific biomarker assays are lacking. Treatment-related increase of allergen-specific antibodies is a robust biomarker of successful AIT. Allergen-specific non-IgE antibodies are believed to reduce the effects of allergen exposure by competing with IgE for allergen binding, and in-vitro assays quantifying the effects of AIT-induced IgE-blocking antibodies are advantageous. A cell-free enzyme-linked immunosorbent facilitated antigen binding (ELIFAB) assay of JC pollen was established. METHODS: Serum IgE-allergen complexes were captured by immobilized recombinant CD23, and allergen-IgE-CD23 complexes were detected by a biotin-conjugated anti-human IgE antibody. Sera from JC pollen-allergic subjects without or with subcutaneous immunotherapy (SCIT) with JC pollen extract were used (n = 11/group). RESULTS: Optimal assay conditions were established at 20 µg/mL CD23 and 0.3 µg/mL JC pollen extract, and the dependency on CD23 and IgE was verified. The data show that the JC pollen ELIFAB assay is fit for purpose and demonstrates that the IgE-blocking activity is significantly increased in the JC pollen SCIT group compared with the non-treated group. CONCLUSION: The JC pollen ELIFAB assay represents a simple, cell-free biomarker assay for monitoring the development of IgE-blocking antibody activity during JC pollen AIT.

Comparison of the Allergenic Potency of House Dust Extract and House Dust Mite Allergen Extract for Subcutaneous Allergen Immunotherapy.

Subcutaneous allergen immunotherapy (SCIT) with non-standardized house dust (HD) extracts has been used in Japan since 1963 for house dust mite (HDM)-allergic patients. Since the potencies of HD extracts are unknown, the allergenic potency of HD extracts was examined by comparing with a standardized HDM allergen extracts. The major allergen content of HDM in the extracts was measured using a sandwich enzyme-linked immunosorbent assay (ELISA). The immunoglobulin E (IgE) inhibitory activities of the extracts were measured by a competitive ELISA. The extract concentrations giving 50% inhibition of IgE binding (log10 IC50) were determined from dose-response curves and defined as inhibitory activities. A linear regression line was constructed from the log10 IC50 values of the standardized HDM extract to interpolate the relative potency of the HD extract with strength of 1 : 10 w/v (HD 1 : 10). The amounts of major allergens (Der f 1, Der p 1 and Der 2) were 116.3 µg/mL in the HDM allergen extract (100000 Japanese Allergy Units [JAU]/mL) and 0.77 µg/mL in the HD 1 : 10. The inhibitory activity (log10 IC50 values) of HD 1 : 10 was 2.389 ± 0.078, indicating the allergenic potency was between 200 and 2000 JAU/mL. Based on regression analysis (R2 >0.99), the allergenic potency of HD 1 : 10 was estimated to be 842 ± 128 JAU/mL. The present study determined the major allergen content of HD extract, which contributes to its allergenic potency. The allergenic potency of HD 1 : 10 was ca. 100-fold less than that of HDM allergen extract.

Efficacy and safety of SQ house dust mite sublingual immunotherapy-tablet in Japanese children.

BACKGROUND: The SQ house dust mite (HDM) sublingual immunotherapy (SLIT)-tablet (TO-203, Torii, Japan/ALK, Denmark) treatment has been effective against respiratory allergic diseases in patients aged ≥12 years during European, Japanese, and North American trials. This trial was conducted to investigate the efficacy and safety of this treatment in Japanese children (5-17 years) with moderate-to-severe HDM allergic rhinitis (AR). METHODS: In this randomized, double-blind, placebo-controlled trial, 458 Japanese children were randomly assigned to a daily SQ HDM SLIT-tablet [10 000 Japanese Allergy Unit (JAU), equivalent to 6 SQ-HDM in Europe and the US] or placebo (1:1) treatment for 1 year. Inclusion required an AR symptom score of ≥7 on at least 7 days during a 14-day run-in period while symptomatic treatment was withdrawn. The primary endpoint was the total combined rhinitis score (TCRS) comprising AR symptom and medication scores during the last 8 weeks of the treatment period. RESULTS: The analysis of primary endpoint demonstrated statistically significant absolute reduction in TCRS of 1.22 with a relative difference of 23% (95% confidence interval, 14% to 31%) in the 10 000 JAU compared with placebo. Predefined stratified analyses revealed the same degree of efficacy of 1.11 (P = 0.002), 21% (8% to 32%) and 1.36 (P = 0.001), 26% (11% to 38%), respectively, in pediatric (5-11 years) and adolescent subjects (12-17 years). The treatment was well tolerated by both pediatric and adolescent subjects. CONCLUSION: This trial, for the first time, demonstrated the efficacy and safety of the HDM SLIT-tablet in pediatric patients with moderate-to-severe HDM AR (JapicCTI-152953).

Efficacy and safety of the SQ house dust mite sublingual immunotherapy tablet in Japanese adults and adolescents with house dust mite-induced allergic rhinitis.

BACKGROUND: The SQ house dust mite (HDM) sublingual immunotherapy (SLIT) tablet has been approved in 11 European countries and Japan for patients with HDM-induced respiratory allergic disease. OBJECTIVE: This trial was conducted to confirm the efficacy and safety of the SQ HDM SLIT tablet in Japanese patients with moderate-to-severe HDM-induced allergic rhinitis (AR). METHODS: The trial was a randomized, double-blind, placebo-controlled trial including 946 Japanese adults and adolescents (12-64 years). Subjects were randomly assigned to daily treatment with the SQ HDM SLIT tablet at a dose of 10,000 Japanese allergy units (JAU) or 20,000 JAU or to placebo (1:1:1). The primary end point was the total combined rhinitis score (TCRS), which is composed of AR symptom and medication scores during the efficacy evaluation period. Symptom and medication scores of AR and conjunctivitis, rhinitis quality of life, and symptom-free and symptom-severe days were evaluated as secondary end points. RESULTS: Analysis of the primary end point demonstrated statistically significant reductions in TCRSs of 1.15 (22%, P < .001) in the 10,000-JAU group and 0.99 (19%, P < .001) in the 20,000-JAU group compared with the placebo group. The statistically significant treatment effect was evident from 12 weeks of treatment onward. All secondary end points, except AR medication score, were statistically significant in favor of active treatment compared with placebo. Post hoc analysis of TCRSs in adolescents showed the same efficacy as in adults (P < .05). The treatment was well tolerated by both adults and adolescents. CONCLUSION: The trial confirmed the efficacy and safety profile of the SQ HDM SLIT tablet in Japanese adult and adolescent patients with moderate-to-severe HDM-induced AR. These data support the robust efficacy and safety profile of previously reported European data.

Current status of sublingual immunotherapy for allergic rhinitis in Japan.

Japanese cedar pollen (JCP) and house dust mite (HDM) are two major allergens that cause allergic rhinitis (AR) in Japan and the prevalence of AR is increasing. Pharmacothearpy is a commonly used treatment, but the level of patient satisfaction is very low. Allergen immunotherapy (AIT) is the only therapeutic modality that provides not only symptom relief but also quality of life improvement that leads to a high rate of satisfaction. In particular, sublingual immunotherapy (SLIT) is a safe and effective treatment for AR. Here we introduce a large-scale double-blind, placebo-controlled trial of SLIT in Japanese patients using JCP droplets or HDM tablets conducted in Japan. The immediate future of SLIT in Japan is also discussed.

Long-term safety of subcutaneous immunotherapy with TO-204 in Japanese patients with house dust mite-induced allergic rhinitis and allergic bronchial asthma: Multicenter, open label clinical trial.

BACKGROUND: To evaluate the long-term safety of subcutaneous immunotherapy with TO-204, a standardized house dust mite (HDM) allergen extracts, we conducted a multicenter, open label clinical trial. METHODS: Japanese patients aged 5-65 years were eligible for the study, if they had HDM-induced allergic rhinitis (AR), allergic bronchial asthma (BA), or both. TO-204 was administered in a dose titration scheme, and the maintenance dose was determined according to the predefined criteria. The treatment period was 52 weeks, and patients who were willing to continue the treatment received TO-204 beyond 52 weeks. This clinical trial is registered at the Japan Pharmaceutical Information Center (Japic CTI-121900). RESULTS: Between July 2012 and May 2015, 44 patients (28 with AR and 16 with allergic BA) were enrolled into the study. All patients were included in the analysis. The duration of treatment ranged from 23 to 142 weeks and the median maintenance dose was 200 Japanese allergy units (JAU). Adverse events occurred in 22 patients (50%). The most common adverse event was local reactions related to the injection sites. Four patients experienced anaphylactic reactions when they were treated with the dose of 500 JAU. Two patients experienced anaphylactic shock with the doses of 1000 JAU at onset. These 6 patients could continue the study with dose reduction. CONCLUSIONS: Safety profile of TO-204 was acceptable in Japanese patients with HDM-induced AR or allergic BA. Higher doses should be administered carefully, because the risk of anaphylaxis increased at doses of 500 or 1000 JAU.

Pooled efficacy and safety data for house dust mite sublingual immunotherapy tablets in adolescents.

BACKGROUND: House dust mite (HDM) respiratory allergy is a common and burdensome disease in children and adolescents. There are few HDM allergy immunotherapy trials in children with perennial allergic rhinitis. This post hoc analysis used pooled data to evaluate efficacy and safety of the SQ HDM sublingual immunotherapy (SLIT) tablet in adolescents (12-17 years). METHODS: In two double-blind, placebo-controlled trials conducted in North America and Japan, respectively, subjects aged 12+ years with HDM allergic rhinitis were randomized to up to 1 year of treatment. The primary end-point in both trials was the average total combined rhinitis score (TCRS) during the last 8 weeks of treatment in the active group compared with placebo. Data from subjects aged 12-17 years were pooled (N=395). RESULTS: In the pooled adolescent subpopulation, average TCRS improved 22% with 12 SQ HDM vs placebo (absolute treatment difference of 1.04; P<.01). Rhinitis daily symptom score (DSS), conjunctivitis DSS and rhinitis daily medication score (DMS) were also significantly improved vs placebo in the pooled adolescent subpopulation (all P<.05). There were no new safety signals for adolescents. The frequency of adverse events was similar in adolescents and adults with the majority being mild application site-related events. CONCLUSIONS: Treatment with 12 SQ HDM appears to be effective and well tolerated in adolescents with HDM allergic rhinitis.

Japanese guidelines for allergic rhinitis 2017.

Like asthma and atopic dermatitis, allergic rhinitis is an allergic disease, but of the three, it is the only type I allergic disease. Allergic rhinitis includes pollinosis, which is intractable and reduces quality of life (QOL) when it becomes severe. A guideline is needed to understand allergic rhinitis and to use this knowledge to develop a treatment plan. In Japan, the first guideline was prepared after a symposium held by the Japanese Society of Allergology in 1993. The current 8th edition was published in 2016, and is widely used today. To incorporate evidence based medicine (EBM) introduced from abroad, the most recent collection of evidence/literature was supplemented to the Practical Guideline for the Management of Allergic Rhinitis in Japan 2016. The revised guideline includes assessment of diagnosis/treatment and prescriptions for children and pregnant women, for broad clinical applications. An evidence-based step-by-step strategy for treatment is also described. In addition, the QOL concept and cost benefit analyses are also addressed. Along with Allergic Rhinitis and its Impact of Asthma (ARIA), this guideline is widely used for various clinical purposes, such as measures for patients with sinusitis, childhood allergic rhinitis, oral allergy syndrome, and anaphylaxis and for pregnant women. A Q&A section regarding allergic rhinitis in Japan was added to the end of this guideline.

Complementary and alternative medicine for allergic rhinitis in Japan.

BACKGROUND: Complementary and alternative medicine (CAM) is extensively used in patients with allergic diseases worldwide. The purpose of this study was to investigate the actual situation of CAM practice in the treatment of allergic rhinitis. METHODS: We distributed questionnaires to otolaryngologists at 114 facilities in Japan. The subjects who participated in this study included children <16 years of age and adults ≥16 years of age diagnosed with allergic rhinitis by otolaryngologists. The survey was performed in the period from September 2007 to August 2009. Furthermore, we performed the same investigation out of the hospital setting, such as during general health examinations. All questionnaires were returned to Chiba University and analyzed. RESULTS: The proportions of patients who had ever experimented with CAM in the hospital survey were 7.1% (225/3170) and 19.2% (1416/7363) of children and adults, respectively. Approximately 36.2% of the adult patients thought that the treatments were effective. The main reasons for CAM use were safety, convenience and low price. However, the group who spent more than $1000 on CAM felt more dissatisfaction and anxiety related to treatment at the hospital. The situation of CAM practice was not consistent and was instead influenced by the backgrounds of the subjects. CONCLUSIONS: Many patients who receive CAM report feeling that the effects of treatment provided by hospitals are insufficient and have concerns about the side effects of such treatments. Information regarding standard treatments, as described in the guidelines, should become widely known and diffused, and strong communication with patients should be considered.

Questionnaire about Snoring and Daytime Sleepiness Due to Springtime Hay Fever.

Bothersome symptoms of hay fever impair not only patients' quality of life but also their labor productivity and learning efficiency. Excessive daytime sleepiness (EDS) caused by hay fever is thought to be one of the reasons for these impairments. The purpose of this study was to investigate the relationship between the severity of springtime hay fever and EDS by using a questionnaire. The questionnaire included information about age, sex, height, weight, severity of hay fever, treatment for hay fever, smoking and alcohol consumption habit, history of drug use for sleeping, existence of snoring, and Japanese version of the Epworth Sleepiness Scale. After excluding responses containing insufficient data, responses from 1,734 patients were considered as eligible. By performing logistic regression analysis, we analyzed the effect of the aforementioned parameters on the comorbidity of EDS and snoring. The odds ratio (OR) to comorbid EDS was significantly higher in the moderate and severe hay fever groups than in the asymptomatic hay fever group (moderate: OR=1.76, p=0.014, severe: OR=2.53, p<0.001). Also, OR to comorbid snoring was significantly higher in the severe hay fever group than in the asymptomatic hay fever group (severe: OR=1.90, p=0.001).

The Efficacy of Sublingual Immunotherapy for Allergic Rhinitis May Vary with the Time of Day.

BACKGROUND: Sublingual immunotherapy (SLIT) is a safe and effective treatment for allergic rhinitis (AR). However, many issues regarding SLIT remain to be resolved, including the optimal timing of administration. This study investigated the effect of time of day on SLIT efficacy with the goal of optimizing the therapeutic outcome. METHODS: We performed prophylactic SLIT at different times of day (10 a.m. or 10 p.m.) in 2 mouse models of AR: an ovalbumin (OVA)-induced AR model and Cry j 1-induced AR model, and compared the effects. RESULTS: In the OVA-induced AR model, mice sublingually receiving OVA at 10 a.m. exhibited a greater decrease in total and OVA-specific IgE levels than mice treated at 10 p.m. In addition, mice treated at 10 a.m. exhibited reductions in OVA-specific IL-4, IL-10, and IL-13 production by splenocytes relative to mice treated at 10 p.m. Furthermore, we observed a more efficient capture of sublingually administered OVA in submandibular lymph nodes at 10 a.m. than at 10 p.m. in mice. Similar results were observed in the Cry j 1-induced AR model using Japanese cedar pollen extract for SLIT. CONCLUSIONS: Given the allergen-specific antibody and T cell responses, we suggest that SLIT may be more effective in the resting phase than in the active phase (note that mice are nocturnal animals). Thus, we propose that a chronotherapeutic approach should be considered for SLIT to maximize its effectiveness.

Epithelial splicing regulatory proteins 1 (ESRP1) and 2 (ESRP2) suppress cancer cell motility via different mechanisms.

ESRP1 (epithelial splicing regulatory protein 1) and ESRP2 regulate alternative splicing events associated with epithelial phenotypes of cells, and both are down-regulated during the epithelial-mesenchymal transition. However, little is known about their expression and functions during carcinogenesis. In this study, we found that expression of both ESRP1 and ESRP2 is plastic: during oral squamous cell carcinogenesis, these proteins are up-regulated relative to their levels in normal epithelium but down-regulated in invasive fronts. Importantly, ESRP1 and ESRP2 are re-expressed in the lymph nodes, where carcinoma cells metastasize and colonize. In head and neck carcinoma cell lines, ESRP1 and ESRP2 suppress cancer cell motility through distinct mechanisms: knockdown of ESRP1 affects the dynamics of the actin cytoskeleton through induction of Rac1b, whereas knockdown of ESRP2 attenuates cell-cell adhesion through increased expression of epithelial-mesenchymal transition-associated transcription factors. Down-regulation of ESRP1 and ESRP2 is thus closely associated with a motile phenotype of cancer cells.

Use of Sato's curved laryngoscope and an insulated-tip knife for endoscopic incisional therapy of esophageal web.

We experienced two cases of esophageal web accompanying severe stricture that were treated by endoscopic incisions with an insulated-tip knife (IT-knife). With attention paid to the mucosa at the stricture, the lesion was incised with an IT-knife without complications. Sato's curved laryngoscope was used even in cervical esophageal lesions and an excellent field was secured.

Japanese Society of Allergology task force report on standardization of house dust mite allergen vaccines - secondary publication.

BACKGROUND: In the 1990s, the Japanese Society of Allergology (JSA) standardized Japanese cedar pollen allergen vaccines. In the present study, the task force for house dust mite (HDM) allergen standardization of the Committee for Allergens and Immunotherapy of JSA reports the standardization of HDM allergen vaccines in Japan. METHODS: In vivo allergenic potency was determined by intradermal testing of 51 Japanese adults with positive serum specific IgE to HDM allergens. In vitro total IgE binding potency was analyzed by competition ELISA using a pooled serum, with sera obtained from 10 allergic patients. The amounts of HDM group 1 (Der 1) and group 2 major allergens in eight HDM allergen extracts were measured by sandwich ELISAs. Correlation between the in vitro total IgE binding potency and major allergen levels was analyzed. RESULTS: We selected a JSA reference HDM extract and determined its in vivo allergenic potency. The in vitro total IgE binding potency significantly correlated with Der 1 content, group 2 allergen content, and their combined amount, indicating that measurement of major allergen contents can be used as a surrogate in vitro assay. CONCLUSIONS: The task force determined the in vivo allergenic potency (100,000 JAU/ml) and Der 1 content (38.5 µg/ml) of the JSA reference HDM extract, selected the measurement of Der 1 content as the surrogate in vitro assay, and decided that manufacturers can label a HDM allergen extract as having a titer of 100,000 JAU/ml if it contains 22.2-66.7 µg/ml of Der 1.

Muscarinic receptor binding of the novel radioligand, [3h]imidafenacin in the human bladder and parotid gland.

The aim of the current study was to demonstrate highly specific and direct binding activity of tritium ([(3)H])-labeled imidafenacin for labeling muscarinic receptors in human bladder and parotid gland. Specific binding of [(3)H]imidafenacin in human tissues was saturable, reversible, and of high affinity. The Kd value for specific [(3)H]imidafenacin binding in the human bladder was approximately 3 times higher than that in the parotid gland. Unlabeled imidafenacin as well as the clinically used antimuscarinic agents, oxybutynin, tolterodine, and solifenacin, competed with [(3)H]imidafenacin for binding sites in the human bladder and parotid gland in a concentrationdependent manner, which indicated pharmacological specificity of [(3)H]imidafenacin binding sites. The Ki for imidafenacin in the human bladder approximately corresponded to pharmacological potency for the competitive blockade of carbachol-induced contractions of bladder, indicating a close correlation between binding affinity of imidafenacin to bladder muscarinic receptors and its pharmacological effects in the bladder. In conclusion, the current study is the first to provide direct evidence to demonstrate that imidafenacin bound muscarinic receptors in the human bladder, supporting its clinical relevance as a therapeutic agent for overactive bladder. [(3)H]Imidafenacin may also be a useful radioligand for labeling the M3 subtype of muscarinic receptors with higher selectivity than other radioligands.