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Basal cell carcinoma is the most common type of skin cancer, and surgical excision with clear margins is the standard of care. Surgical margins are determined based on risk factors (high or low risk) for recurrence according to the National Comprehensive Cancer Network and Japanese basal cell carcinoma guidelines. The clarity of the clinical tumor border (well-defined or poorly defined) is considered a risk factor, and significant discrepancies in the judgment of clinical tumor borders among dermato-oncologists may occur. Therefore, we analyzed the dermato-oncologists' concordance in judging the clinical tumor border of basal cell carcinoma. Forty-seven dermato-oncologists (experts: 37; young trainees: 10) participated in this study. The datasets of clinical and dermoscopic photographs of 79 Japanese cases of head and neck basal cell carcinoma were used to determine the concordance in the judgment of clinical tumor border. The probability of the border that was selected more often was used to calculate the rater agreement rate for each dataset. Correct judgment was defined as a more frequently selected border, and the concordance rate of clarity of clinical tumor border for each dermato-oncologist was calculated based on the definition of the correct judgment. A median concordance rate of 85% or higher for all dermato-oncologists was predefined as an acceptable rate for clinical use. Of the 79 datasets, rater agreement rates were 80-100%, 60-79%, and 51-59% for 55, 19, and five datasets, respectively. The median concordance rate for all dermato-oncologists was 86% (interquartile range: 82-89%). There was no significant difference in the concordance rate between the experts and the trainees (median, 87% vs. 85.5%; p = 0.58). The concordance rates of dermato-oncologists for all datasets were relatively high and acceptable for clinical use.

Objective: Malignant melanomas of the vulva (VuM) and vagina (VaM) represent a unique subgroup of rare malignant melanomas with critical biological properties and treatment differing from that of other cancers. In Japan, adequate surveys on these have not been performed. The objective of this study was to elucidate the clinicopathological demographics and the outcomes of VuM and VaM in Japan.Methods: This retrospective observational study included women with invasive VuM or VaM, identified from older medical records in Japan. Clinical data were collected and the Kaplan-Meier method was used to analyze progression-free survival (PFS) and overall survival (OS). Univariate and multivariate regression models were used to identify factors significantly related to survival.Results: A total of 217 patients were identified: 109 (50.2%) with VuM and 108 (49.8%) with VaM. The median PFS was 16.8 months in patients with VuM (95% confidence interval [CI] 23.1-87.7) and 15.6 months in patients with VaM (95% CI 8.4-12.6). The median OS was 43.9 months (95% CI 60-138) and 31.1 months (95% CI 24.8-45.3) in patients with VuM and VaM, respectively. Multivariate analysis showed that a >III American Joint Committee on Cancer (AJCC) disease stage (hazard ratio [HR] = 2.063; 95% CI = 0.995-4.278) was associated with poorer PFS, and unknown surgical margin was the only independent factor influencing OS (HR = 2.188; 95% CI = 1.203-3.977).Conclusions: The overall outcomes of VuM and VaM remain poor in Japan. The AJCC stage and the surgical margin are significant predictors of survival.

Background The American Joint Committee on Cancer (AJCC) 8th Edition Cancer Staging System was implemented in 2018; however, it has not been validated in an Asian melanoma population. Objective The purpose of this study was to validate the new system using a cohort of Japanese melanoma patients. Methods The AJCC 7th and 8th Editions were used for TNM classification of patients in a database established by the Japanese Melanoma Study Group. Patient data with sufficient information to be applicable to the AJCC 8th staging were selected. The Kaplan–Meier method was used to estimate disease-specific survival and relapse-free survival. Results In total, data for 3097 patients were analyzed. The 5-year disease-specific survival according to the 7th and 8th Edition staging system were as follows: IA = 98.5%/97.9%; IB = 95.4%/96.2%; IIA = 94.2%/94.1%; IIB = 84.6%/84.4%; IIC = 72.2%/72.2%; IIIA = 76.2%/87.5%; IIIB = 60.7%/72.6%; IIIC = 42.0%/55.3% and IIID = none/26.0%. The 5-year relapse-free survival according to the 7th and 8th Edition staging was as follows: IA = 94.5%/92.7%; IB = 85.4%/85.3%; IIA = 80.1%/79.4%; IIB = 71.4%/70.6%; IIC = 56.8%/55.7%; IIIA = 56.8%/69.4%; IIIB = 42.6%/56.8%; IIIC = 20.0%/33.3% and IIID = none/6.5%. Conclusion The results show that new staging system could efficiently classify our Japanese melanoma cohort. Although there was no difference in Stage I and II disease between the 7th and 8th Edition systems, we should be careful in managing Stage III disease since the survival curves of the 8th Edition staging were completely different from the 7th Edition. Moreover, our results indicate that adjuvant therapies for Stage IIB and IIC should be developed, since the relapse-free survival for these stages were equivalent to Stage IIIA and IIIB, respectively.

Background The incidence of melanoma among those of an Asian ethnicity is lower than in Caucasians; few large-scale Asian studies that include follow-up data have been reported. Objectives To investigate the clinical characteristics of Japanese patients with melanoma and to evaluate the prognostic factors. Methods Detailed patient information was collected from the database of Japanese Melanoma Study Group of the Japanese Skin Cancer Society. The American Joint Committee on Cancer seventh Edition system was used for TNM classification. The Kaplan-Meier method and Cox proportional hazards model were used to estimate the impact of clinical and histological parameters on disease-specific survival in patients with invasive melanoma. Results In total, 4594 patients were included in this analysis. The most common clinical type was acral lentiginous melanoma (40.4%) followed by superficial spreading melanoma (20.5%), nodular melanoma (10.0%), mucosal melanoma (9.5%), and lentigo maligna melanoma (8.1%). The 5-year disease-specific survival for each stage was as follows: IA = 98.0%, IB = 93.9%, IIA = 94.8%, IIB = 82.4%, IIC = 71.8%, IIIA = 75.0%, IIIB = 61.3%, IIIC = 41.7%, and IV = 17.7%. Although multivariate analysis showed that clinical classifications were not associated with survival across all stages, acral type was an independent poor prognostic factor in stage IIIA. Conclusions Our study revealed the characteristics of melanoma in the Japanese population. The 5-year disease-specific survival of each stage showed a similar trend to that of Caucasians. While clinical classification was not associated with survival in any stages, acral type was associated with poor survival in stage IIIA. Our result might indicate the aggressiveness of acral type in certain populations.

Melanoma is one of the most lethal and malignant cancers and its incidence is increasing worldwide, and Japan is not an exception. Although there are numerous therapeutic options for melanoma, the prognosis is still poor once it has metastasized. The main concern after removal of a primary melanoma is whether it has metastasized, and early detection of metastatic melanoma would be effective in improving the prognosis of patients. Thus, it is very important to identify reliable methods to detect metastases as early as possible. Although many prognostic biomarkers (mainly for metastases) of melanoma have been reported, there are very few effective for an early diagnosis. Serum and urinary biomarkers for melanoma diagnosis have especially received great interest because of the relative ease of sample collection and handling. Several serum and urinary biomarkers appear to have significant potential both as prognostic indicators and as targets for future therapeutic methods, but still there are no efficient serum and urinary biomarkers for early detection, accurate diagnosis and prognosis, efficient monitoring of the disease and reliable prediction of survival and recurrence. Levels of 5-S-cysteinyldopa (5SCD) in the serum or urine as biomarkers of melanoma have been found to be significantly elevated earlier and to reflect melanoma progression better than physical examinations, laboratory tests and imaging techniques, such as scintigraphy and echography. With recent developments in the treatment of melanoma, studies reporting combinations of 5SCD levels and new applications for the treatment of melanoma are gradually increasing. This review summarizes the usefulness of 5SCD, the most widely used and well-known melanoma marker in the serum and urine, compares 5SCD and other useful markers, and finally its application to other fields.

Most patients with extramammary Paget’s disease (EMPD) show a good prognosis; however, some patients develop fatal metastases. Early detection is important for improving prognosis, due to the difficulties associated with the treatment of distant EMPD metastases. Several studies have emphasized the importance of the invasion level of the primary lesion for predicting the presence of metastasis, and deeper invasion or increased thickness is correlated with poorer prognosis. Vascular tumor invasion of the primary lesion can also predict the risk of metastasis. Lymph node metastasis is a strong indicator for poor prognosis, and the number of lymph node metastases affects patient outcome, in that there is a significant difference in survival between patients with zero or one lymph node metastasis and those with more than two lymph node metastases. Serum markers may be able to predict the presence of systemic metastases, and carcinoembryonic antigen and cytokeratin 19 fragment 21-1 reflect disease progression and may be clinically valuable. Although several genetic alterations have been determined for EMPD, factors determining prognosis should be further explored.

Background The importance of the genetic background of melanoma cells to the individual susceptibility to treatment has become apparent. In Caucasians, BRAF mutations are frequently detected in lesions on the skin of younger patients compared to NRAS and KIT mutations. However, clinical and pathological characteristics associated with BRAF , NRAS and KIT mutations have not been fully evaluated in East Asians. Objective To clarify clinical and pathological characteristics associated with BRAF , NRAS and KIT mutations in Japanese melanoma patients. Methods Clinical data were retrospectively collected from 11 hospitals in Japan. BRAF , NRAS and KIT mutations were evaluated with polymerase chain reaction and Sanger sequencing. The relationships between these gene mutations and pathological and clinical findings were analyzed. Results The number of cases examined was 171 (primary: 135, metastases: 11, paired: 25), and all were Japanese patients. The detection rates of BRAF , NRAS and KIT mutations were 30.4%, 12.3% and 12.9%, respectively. Compared with the wild type, the presence of BRAF mutations was significantly associated with younger age (median, 50.0 years vs. 70.0 years, p BRAF mutation was frequently detected in the lesions of the scalp (80%; 4/5), trunk (72.0%; 18/25), extremities (56.7%; 17/30) and neck (44.4%; 4/9), and the least prevalent were the face (22.2%; 2/9), nail (12.5%; 3/24), palm or sole (8.9%; 4/45) and mucosa (0%). NRAS mutations were prevalent in the face (33.3%) and palm or sole (20.0%), and the median age of these patients was 70.5 years. A KIT mutation was observed in the nail apparatus (25%), palm or sole (15.6%) and mucosa (18.2%). The median age of the patients with a KIT mutation was 63.0 years. Heterogeneity of mutations between primary and metastatic lesions was detected in six of 25 cases (24%). Solar elastosis was identified in 12 of 71 cases (15.3%), among which four cases harbored BRAF V600E (2 cases), BRAF V600K , NRAS Q61K or NRAS Q61L , respectively. Conclusion Some clinical characteristics associated with BRAF , NRAS and KIT mutations were observed in Japanese patients, and we observed both similarities to and differences from those of Caucasians. Our findings could provide useful information in efforts to clarify the tumor genesis of malignant melanomas.

Muir-Torre syndrome (MTS) is a familial cancer syndrome characterized by a predisposition to keratoacanthoma (KA) and sebaceous tumors. Although MTS and hereditary non-polyposis colorectal cancer (HNPCC) share the same genetic alterations in mismatch repair (MMR) genes, the other skin lesions in MTS or HNPCC have been only rarely reported. We report a family with an MSH2 mutation c.1126_1127delTT (p.Leu376Thrfs*12). A 46-year-old male proband developed KA with sebaceous differentiation, colon cancer and gastric cancer, and fulfilled the diagnostic criteria for MTS. His 80-year-old mother, diagnosed with HNPCC, presented with multiple gastrointestinal tract cancers, Bowen's disease and actinic keratosis. Immunostaining revealed attenuated MSH2 protein expression in KA, as well as in Bowen's disease and actinic keratosis lesions. These findings suggest that MMR gene abnormality is also critical in the development of benign or malignant cutaneous tumors such as actinic keratosis and Bowen's disease in MTS/HNPCC patients.

To investigate the trends of antimicrobial resistance in pathogens isolated from skin and soft-tissue infections (SSTI) at dermatology departments in Japan, a Japanese surveillance committee conducted the first nationwide survey in 2013. Three main organisms were collected from SSTI at 30 dermatology departments in medical centers and 10 dermatology clinics. A total of 860 strains – 579 of Staphylococcus aureus , 240 of coagulase-negative Staphylococci , and 41 of Streptococcus pyogenes – were collected and shipped to a central laboratory for antimicrobial susceptibility testing. The patient profiles were also studied. Among all 579 strains of S. aureus , 141 (24.4%) were methicillin-resistant (MRSA). Among 97 Staphylococcus epidermidis strains, 54 (55.7%) were methicillin-resistant (MRSE). MRSA and MRSE were more frequently isolated from inpatients than from outpatients. Furthermore, these methicillin-resistant strains were also isolated more frequently from patients with histories of taking antibiotics within 4 weeks and hospitalization within 1 year compared to those without. However, there were no significant differences in MIC values and susceptibility patterns of the MRSA strains between patients with a history of hospitalization within 1 year and those without. Therefore, most of the isolated MRSA cases at dermatology departments are not healthcare-acquired, but community-acquired MRSA. S. pyogenes strains were susceptible to most antibiotics except macrolides. The information in this study is not only important in terms of local public health but will also contribute to an understanding of epidemic clones of pathogens from SSTI.

9529 Background: Anti-PD-1 antibodies are used worldwide for patients (pts) with advanced melanoma. Clinical trials have demonstrated its efficacy and safety for nonacral cutaneous melanoma (NACM) in controlled settings. Since acral melanoma (AM) is epidemiologically and molecularly distinct from NACM, data on the real-world efficacy of anti-PD1 antibodies in advanced AM is still lacking. Thus, we aim to analyze the real-world efficacy and safety of anti-PD-1 antibodies in advanced AM. Methods: We retrospectively reviewed clinical records of advanced AM treated in any line with an anti-PD1 antibody at 21 Japanese institutions. Clinical response was assessed by (Response Evaluation Criteria in Solid Tumors) RECIST criteria. Survival was estimated using Kaplan-Meier analysis. Toxicity was assessed according to CTCAE 4.0. Results: A total of 193 pts (median age, 71 years) with advanced AM (subungual, 70; palm and sole, 123) were included in the study. Anti-PD-1 antibody was used as first-line therapy in 143 pts (74.1%). Nivolumab was used in 168 pts and pembrolizumab in 25 pts. Base-line lactate dehydrogenase (LDH) was within normal level in 102 pts (52.8%). The objective response rate (ORR) of all pts was 16.5% (complete response 3.1%, partial response 13.5%), and median overall survival (OS) was 18.1 months. Normal LDH level was significantly associated with better prognosis than abnormal level (median OS 24.9 vs 10.7 months; P < 0.001). Although baseline demographics and characteristics were almost similar between subungual group and palm and sole group, ORR was significantly lower in the subungual group than in palm and sole group (6/70 pts [8.6%] vs 26/123 pts [21.1%]; P = 0.026); median OS was significantly poorer as well (12.8 vs 22.3 months; P = 0.031). Immune-related adverse events of grades 3 to 5 occurred in 27 pts (14.0%). One patient (0.5%) died of grade 5 myasthenia gravis. Conclusions: Real-world efficacy of anti-PD-1 antibodies in AM pts is limited. Notably, pts with subungual melanoma showed poorer response and survival, making them strong candidates for further research of efficacy of anti-CTLA-4 and anti-PD-1 combination therapy.

Cowden syndrome (CS) is a rare inherited disorder characterized by multiple hamartomas and an increased risk of carcinomas, such as thyroid and breast cancers [1]. Characteristic mucocutaneous manifestations include multiple facial trichilemmomas, oral mucosal papillomatoses and acral keratosis. A mutation in the phosphatase and tensin homolog (PTEN) gene has been identified as the cause of CS [2]. The present report describes a case of extramammary Paget's disease (EMPD) which developed in a Japanese [...]

Although nivolumab is associated with a significant improvement in overall survival and progression-free survival, only 20 to 40% of patients experience long-term benefit. It is therefore of great interest to identify a predictive marker of clinical benefit for nivolumab. To address this issue, the frequencies of CD4+ T cell subsets (Treg, Th1, Th2, Th9, Th17 and Th22), CD8+ T cells, and serum cytokine levels (IFNγ, IL-4, IL-9, IL-10, TGF-β) were assessed in 46 patients with melanoma. Eighteen patients responded to nivolumab, and the other 28 patients did not. An early increase in Th9 cell counts during the treatment with nivolumab was associated with an improved clinical response. Before the first nivolumab infusion, the responders displayed elevated serum concentrations of TGF-β compared to non-responders. Th9 induction by IL-4 and TGF-β was enhanced by PD-1/PD-L1 blockade in vitro. The role of IL-9 in disease progression was further assessed using a murine melanoma model. In vivo IL-9 blockade promoted melanoma progression in mice using an autochthonous mouse melanoma model, and the cytotoxic ability of murine melanoma-specific CD8+ T cells was enhanced in the presence of IL-9 in vitro. These findings suggest that Th9 cells, which produce IL-9, play an important role in the successful treatment of melanoma patients with nivolumab. Th9 cells therefore represent a valid biomarker to be further developed in the setting of anti-PD-1 therapy.

Although extramammary Paget disease (EMPD) usually appears as carcinoma in situ, it sometimes becomes invasive (iEMPD) and fatal. However, a TNM staging system for iEMPD has yet to be established.The aim of this study was to establish a TNM staging system for iEMPD.We retrospectively collected iEMPD patients treated at 12 institutes in Japan. Factors reported to be associated with survival such as distant metastasis, lymph node (LN) metastasis, and primary tumor status were evaluated using the log-rank test.We enrolled 301 iEMPD patients, of whom 114 had remote metastases (49 had both distant and LN metastasis; 2, distant metastasis only; and 63, LN metastasis only) and the remaining 187 patients had no remote metastasis. Distant metastasis (M1) showed worse survival (P0.00001). In the analysis of the 250 patients without distant metastasis, LN metastasis also showed worse survival (P0.00001). Among the patients with LN metastasis, 2 or more LN metastases (N2) showed worse survival than did single LN metastasis (N1, P=0.02). Lastly, in the analysis of the 187 patients without metastasis, tumor thickness of over 4mm or lymphovascular invasion showed worse survival (T2, P0.05 and P0.001, respectively). Patients with neither of these features were defined as T1. From these results, we propose this TNM staging system: stage I, T1N0M0; stage II, T2N0M0; stage IIIa, anyTN1M0; stage IIIb, anyTN2M0; stage IV, anyTanyNM1. Other than stages II and IIIa, each stage had a statistically distinct survival curve.We propose a TNM staging system for EMPD using simple factors for classification that could provide important prognostic information in managing EMPD. However, accumulation of more patient data and further revision of the system are required.

The benefit of a sentinel lymph node (SLN) biopsy and adjuvant therapy for patients with thick (>4 mm) melanoma has not been well studied in the Asian population. We examined the benefit of an SLN biopsy and adjuvant therapy on prognosis in Japanese patients with thick melanoma. A review of the melanoma database collected from 26 institutions in Japan identified 291 patients with thick melanoma between 2005 and 2010. Univariate and multivariate analyses were performed to evaluate the factors predictive of the overall survival (OS) and the disease-free survival (DFS). Of the 242 patients with thick melanoma who underwent an SLN biopsy, the results for 96 (40%) were positive. On multivariate analysis, increased Breslow thickness (relative risk, 1.11; 95% confidence interval, 1.05-1.17; P=0.0002) and SLN metastasis (2.14; 1.04-4.43; P=0.040) were associated with a poor OS. Increased Breslow thickness (1.11; 1.04-1.18; P =0.0018), ulceration (3.11; 1.25-7.72; P=0.014), satellitosis (3.89; 1.62-9.31; P=0.0023), and SLN metastasis (2.24; 1.16-4.36; P=0.017) were associated with DFS. Adjuvant chemotherapy had no impact on either OS or DFS. Adjuvant use of a monthly dermal injection of interferon-β (IFN-β) was associated with a improvement in both OS (0.34; 0.17-0.67; P=0.0022) and DFS (0.42; 0.20-0.86; P=0.018). An SLN biopsy provided useful prognostic information and the adjuvant use of IFN-β improved both OS and DFS in Japanese patients with thick melanoma. These results were consistent with those of previous studies carried out on a white population. Therefore, we suggest that an SLN biopsy and adjuvant IFN should be considered for patients with thick melanoma irrespective of the Breslow thickness or ethnicity.

Objective To identify the 140-kd autoantigen recognized by anti-155/140 autoantibodies that are associated with adult cancer-associated dermatomyositis (DM) and juvenile DM and to determine the clinical relevance of anti-155/140 antibodies in a large cohort. Methods Sera from 456 DM patients were assessed for the presence of anti-155/140 antibodies by immunoprecipitation using K562 cell extracts as substrate. Using immunoprecipitation and Western blotting, we then examined whether anti-155/140–positive sera recognized transcription intermediary factor 1α (TIF-1α), TIF-1β, and TIF-1γ. The clinical associations of antigen reactivity were also evaluated. Results Anti-155/140–positive sera reacted with 140-kd TIF-1α in addition to 155-kd TIF-1γ. Among sera from 456 DM patients, 52 were reactive with both TIF-1α and TIF-1γ, while another 25 were reactive with TIF-1γ alone. Additionally, 7 were reactive with TIF-1β. Malignancy was more frequently found in adult patients with both anti–TIF-1α and anti–TIF-1γ antibodies than in those with anti–TIF-1γ antibodies alone (73% versus 50%; P < 0.05). In addition to juvenile DM patients and middle-aged and older DM patients with high percentages of malignancy, 8 “young adult” DM patients without malignancy had these autoantibodies. Conclusion Anti-155/140 antibodies target TIF-1 family proteins, TIF-1α and TIF-1β, in addition to TIF-1γ. Since TIF-1 proteins have significant roles in oncogenesis, these antibodies may be produced during misdirected antitumor immunity.

Various microscopic classifications of metastatic sentinel lymph nodes (SLN) have been reported along with predictors of additional lymph node positivity and their correlations with the prognosis. The purpose of this study was to re-evaluate these classifications in the Japanese population. We selected the following three classifications, based on the procedural simplicity of the measurements: maximum diameter (maximum diameter of the largest tumor lesion in the SLN; 1.0 mm), invasion depth (depth of tumor invasion measured from the capsule in the SLN; SI £0.3 mm, SII >0.3 to £1.0 mm, SIII >1.0 mm), and microanatomic location (microanatomic location of the tumor deposits within the SLN; ‘‘subcapsular’’, ‘‘parenchymal’’, ‘‘combined’’, ‘‘multifocal’’, ‘‘extensive’’). A retrospective study, using prescribed survey forms, was carried out. Among the 450 patients, including the 149 cases with SLN metastasis, an additional lymph node positivity rate of 0% could be predicted only in patients with a maximum diameter category of less than 0.1 mm. As compared with that in the SLN metastasis-negative cases, however, the prognosis was poorer in cases with SLN metastasis, even those with lesions falling under the maximum diameter category of less than 0.1 mm, invasion depth category of SI (£0.3 mm) and microanatomic location category of subcapsular. The prognosis is particularly poor for the microanatomic location category of extensive, which should thus be regarded as a macrometastasis. A prospective study with standardized procedures, including pathological evaluation, is needed in order to confirm our conclusion.

頭部血管肉腫は極めて予後の悪い腫瘍であるが, 最近では新しい治療法により予後に若干の改善がみられるようになっている。治療法と予後との関連を検討するため1995年～2005年の11年間に金沢大学皮膚科で経験した12例 (男性8例, 女性4例) の頭部血管肉腫について解析した。12例中9例は広範囲切除 (マージンの平均2.5cm) を含む集学的治療を行った。手術を行わなかった2例を含む4例にrIL-2を使用した。6例で電子線の術後照射を, 2例では再発病変に対し放射線治療を行った。ドセタキセルによる化学療法 (weekly TXT) を3例の肺転移巣の治療に, 5例で術後補助療法として行った。局所再発は5例にみられた。7例に肺転移が出現し, 12例中7例が原病死した。術後にweekly TXT療法を行った群は行わなかった群に比べ有意に生存期間が長かった。

乳房外パジェット病の診療ガイドラインは, エビデンスレベルの高い報告がないため推奨グレードが低いものにならざるを得ない。従って推奨度は治療の現状とエキスパートオピニオンも参考に決定されており, ガイドラインの質向上のためには今後の研究に期待される。実際の診療においてはガイドラインの推奨度だけでなくその根拠となるエビデンスをも吟味して, 最終的には主治医の判断が尊重されるべきである。

Axillary lymph node dissection (ALND) has been recommended to include levels I–III for melanoma patients who have evidence of metastasis in the axillary sentinel lymph node (SLN). The extent of the subsequent axillary dissection is in debate. The objective of this study was to determine the frequency of metastasis of level III nodes in addition to that of level II nodes in this setting. A multi-institutional retrospective study was undertaken in 14 melanoma treatment centers in Japan. Between 2007 and 2012, 69 patients with involved axillary SLNs underwent a subsequent ALND and 55 underwent level I and II dissections. Level III metastatic nodes, which is our primary endpoint, were seen in only 1 patient (1.5 %). The level II metastatic rate was 4.4 %. Our study sample size was small, but melanoma patients with positive SLN rarely had level III disease, suggesting that level III dissection may be unnecessary. We also found that level II metastasis was not so frequent. More evidence is needed to standardize the extent of ALND and to identify the patients who would have the most benefit with undergoing level II dissection for positive axillary SLNs.

In invasive extramammary Paget's disease (EMPD), distant metastases may develop and the condition may become fatal; however, no standardized treatment has been established. Although based on only a few cases, several chemotherapy regimens were reported to be promising. We conducted a multicenter, retrospective study to evaluate the efficacy of docetaxel for metastatic EMPD. We retrospectively collected data on 18 metastatic EMPD patients treated using docetaxel from 1998 to 2012 in 12 institutes in Japan. The following clinical data were collected: tumor response, time to progression, overall survival and adverse effects. Of those, three patients treated combined with S-1, one patient treated with weekly schedule and one patient treated combined with radiotherapy were excluded from the further analysis. All 13 patients received monthly docetaxel as the first-line treatment. The average number of treatment cycles was 9.1. Among the 12 patients with a confirmed response, seven (58%) showed a partial response, three (25%) stable disease and two (17%) progressive disease. The disease control rate (partial response + stable disease) was as high as 83%. The time to progression and median overall survival were 7.1 and 16.6 months, respectively. The 1-year overall survival rate determined by the Kaplan-Meier method was 75.0%. All adverse effects were manageable and no treatment-related deaths were observed. The high disease control rate and overall survival shown by this study suggest that first-line use of docetaxel may be a promising treatment for metastatic EMPD. A prospective clinical trial is required to confirm our results.

Background Although extramammary Paget's disease (EMPD) mostly presents as intraepithelial carcinoma, we sometimes encounter patients with invasive EMPD (iEMPD) who have lymph node metastasis and may develop distant metastasis. Although sentinel lymph node biopsy (SLNB) is widely accepted for various cancers, there is no large study that has assessed its role in iEMPD. Objective The main objective of this study is to assess the role of SLNB in patients with iEMPD. Materials and methods We retrospectively collected data on 151 iEMPD patients treated from 1998 to 2012 in 11 institutes in Japan. All 151 patients received curative surgery for their primary tumor and none of them had distant metastasis. SLNB was performed on the 107 patients without lymphadenopathy to determine their LN status. The 44 other patients with lymphadenopathy underwent one of the following procedures to determine their LN status: SLNB in 22 cases, immediate LN dissection in 21, and LN biopsy in 1. Results Compared to those without lymphadenopathy, patients with lymphadenopathy had advanced primary tumors (nodule in the primary tumor, thicker tumor, deeper invasion level, and lymphovascular invasion). The rate of LN metastasis in patients with lymphadenopathy was 80%, compared to 15% in patients without lymphadenopathy who underwent SLNB. Compared to those with negative SLN, patients with positive SLN had advanced primary tumors (nodule in the primary tumor, deeper invasion level, and lymphovascular invasion). Multivariate analysis revealed that dermal invasion (odds ratio 5.8, p = 0.04) and lymphovascular invasion (odds ratio 18.0, p = 0.0023) were independent factors associated with SLN positivity. Notably, there was no difference in survival between patients with or without SLN metastasis ( p = 0.71). On the other hand, patients with lymphadenopathy showed worse survival than those with positive SLN ( p = 0.045). Conclusion Clinical lymphadenopathy was strongly correlated with pathological LN metastasis and also associated with worse survival than absence of lymphadenopathy. The rate of occult LN metastasis detected by SLNB was 15%. Survival was not affected by SLN status even when an advanced primary tumor was present in patients with positive SLN. Our results raise the possibility that SLNB and subsequent LN dissection improved the survival of patients with early stage lymphatic spread. Our study indicates that SLNB should be considered for iEMPD if lymphadenopathy is not apparent.

To investigate whether the frequency of the BRAFV600E (V-raf murine sarcoma virus oncogene homolog B1) mutation in melanocytic nevi is associated with sun exposure patterns, we examined 120 acquired melanocytic nevi excised from various anatomic sites, including glabrous skin, as well as 62 congenital nevi. We used a new mutation detection system based on the shifted termination assay, called Mutector, which was able to detect only 5% of heterozygous mutant cells within the samples. We detected the mutation in 105/120 (87.5%) acquired nevi and 43/62 (69.4%) congenital nevi. Notably, we found the mutation in 35/43 (81.4%) acquired nevi excised from glabrous skin and genitalia. These results strongly suggest that UV light is not necessarily required for the acquisition of the BRAFV600E mutation, and suggest that non-mutagenic effects of UV light to melanocytes may be more important in the nevogenesis. Additionally, we showed heterogeneous distribution of BRAF-mutated cells within the lesions of small congenital nevi by a combination of laser microdissection and direct sequencing. Finally, we found low frequency of BRAFV600E mutation (6/20, 30.0%) in medium-sized congenital nevi. Most of these nevi with wild-type BRAF had neroblastoma ras viral oncogene homolog mutations (9/14, 64.3%), suggesting different pathogenesis of medium-sized congenital nevi from acquired nevi and small congenital nevi.

Although there is lymphatic flow into the popliteal fossa from a skin tumor located in the lower leg, popliteal metastasis is extremely rare. Recently, sentinel lymph nodes outside traditional nodal basins have been identified. This study investigated the incidence of sentinel nodes in the popliteal region and the indication for biopsy.Fourteen patients with various skin cancers involving the lower extremities (nine melanomas, four squamous cell carcinomas, and one sweat gland carcinoma) underwent lymphoscintigraphy and excision with sentinel lymph node biopsy.In all 14 patients, hot spots showed accumulation in the groin region. Five of 14 patients (36%) demonstrated popliteal sentinel nodes in addition to the inguinal nodes. Three of five popliteal sentinel nodes were histologically studied. A patient with acral melanoma demonstrated micrometastasis of melanoma cells in a popliteal node but not in the groin node.This study demonstrates that sentinel lymph nodes located in the popliteal fossa are frequently detected by lymphoscintigraphy and that biopsy should be performed if popliteal nodes are identified.

Background: Distinguishing basal cell carcinoma (BCC) from other benign and malignant skin tumors is sometimes a difficult task for the pathologists. Because the activation of hedgehog signals and the up-regulation of its critical transcriptional factor Gli1 are well documented in BCC, a molecular technique measuring Gli1 transcripts may aide the diagnosis. Methods: Gli1 transcript levels were measured by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR) using RNA extracted from formalin-fixed, paraffin-embedded tissues of 68 cases of various skin tumors. Hematoxylin and eosin-stained pathology slides were independently reviewed by three expert dermatopathologists. Results: The histological diagnoses were unambiguous in 53 tumors. The tumors included BCC (21), squamous cell carcinoma (13), seborrheic keratoses (8), trichoepithelioma (5), eccrine poroma/porocarcinoma (4), and sebaceous epithelioma/carcinoma (2). In these unambiguous cases, all BCC and trichoepithelioma tumors showed high expression of Gli1mRNA, while the expression was virtually absent in other tumors. The diagnosis was discordant among three pathologists in the remaining 15 tumors. Histological diagnoses included BCC, BCC with sebaceous differentiation, sebaceoma/sebaceous epithelioma, trichoblastoma, trichoepithelioma, basaloid follicular harmartoma, basosquamous carcinoma, etc. Six of them showed high Gli1 transcript levels. Conclusions: Quantification of Gli1 transcripts by RT-PCR is helpful in discriminating BCC and trichoepithelioma from other skin tumors.

Background: Apoptotic pathway aberrations are reported as important tumor progression factors in melanoma. Objective: Effect of soluble Fas ligand (sFasL) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on short-term cultured melanoma cell viability from different stages of melanoma. Results: Recombinant human FasL reduced viability after 18 h in a dose-dependent manner in 4 of 5 cell cultures from primary tumors and 1 of 9 cell cultures from metastatic melanoma (67.5 vs. 96.4%, p = 0.007). DNA fragmentation on flow cytometry confirmed apoptosis. Incubation with TRAIL had no effect on melanoma cell viability. Immunohistochemistry showed Fas in 3 of 4 primary and in 6 of 7 metastatic lesions, no FasL in primary lesions, and FasL in 5 of 7 metastatic lesions. Conclusion: Melanoma short-term cell cultures from primary tumors show decreased viability under FasL, but not TRAIL stimulation rather than short-term cell cultures derived from metastases.

最近16年間に当科で治療した乳房外Paget病55例について, 臨床的統計および予後の検討を行った。平均年齢は70.8歳, 男女比は2.9対1, 発生部位は外陰部が50例, 肛囲が3例, 下腹部1例, 腋窩4例であった。他臓器癌を合併したものは12.7%で, 表皮内癌は49%であった。リンパ節転移は18%に認められた。局所再発は7%に生じた。原病死は7例, 13%であった。原発巣に結節があるもの, 組織学的にT3以上のものは予後が不良であった。

Background. Surgical treatment of a recessive dystrophic epidermolysis bullosa patient is very difficult for both the surgeon and the anesthetist because of the fragility of the skin and abnormal nature of the tumor bed. Objective. We report a case of 54-year-old Japanese recessive dystrophic epidermolysis bullosa patient with squamous cell carcinoma (SCC) of the lateral malleolus. Methods. A tumor measuring 5.0 × 5.5 cm was surgically excised. The defect was then reconstructed by full-thickness skin grafting. To avoid airway complications, general anesthesia was administered using a face mask. Because the regional lymph nodes were swollen before surgery, the patient underwent sentinel lymph node biopsy. Results. The patient remains well with no sign of recurrence or metastasis 7 months after surgery. Conclusion. To preserve activities of daily living, surgery should be performed for squamous cell carcinomas arising in recessive dystrophic epidermolysis bullosa patients.

Sentinel lymph node (SLN) biopsies have widely been used for the detection of occult LN metastasis of malignant melanoma (MM). In addition to conventional biomarkers, we assessed the diagnostic and prognostic significance of melanoma-initiating cell (MIC) markers in SLNs of MM. We examined the expressions of gp100, MART-1 and tyrosinase mRNA for routine diagnosis and those of ABCB5, CD133, nestin, KDM5B, NGFR and RANK mRNA as MIC markers. The presence of micrometastasis was confirmed immunohistochemically using antibodies to S-100, HMB-45, MART-1, and tyrosinase. Discordance between immunohistochemical and molecular data was observed in 14 of 70 (20.0%) patients, among whom five (7.1%) were positive for only molecular markers;two of these five patients tested positive for micrometastasis by repeated immunohistochemical stainings. The quantitative expression levels of gp100, MART-1, and tyrosinase mRNA were significantly higher in the metastatic LNs;the cut-off values remain to be elucidated. ABCB5 mRNA expression was detected more frequently in the metastatic SLNs (p＜0.05) and in the group of patients with recurrence. To make a definite diagnosis of metastasis, we still need a combination of immunohistochemical and molecular probes. ABCB5 might be a suitable molecular marker for the detection of melanoma-initiating cells in SLNs.

We present a sporadic case of a Japanese female infant with ectodermal dysplasia, complete cleft lip and palate, severe skin erosions at birth and recurrent scalp infection. She had typical clinical features of ankyloblepharon, ectodermal defects and cleft lip and palate (AEC) syndrome without ankyloblepharon. Histological and immunohistochemical analyses showed reduced granular cell layers and aberrant expression of the p63 protein in the suprabasal keratinocytes. Mutation analysis of the exon 13 of p63 gene revealed a heterozygous in-frame 3-bp insert (c. 538-539 ins TTC) encoding additional amino acid residues (F). This is the first report of sterile alpha motif domain mutation except for single nucleotide transitions.

One of the principal applications of tumour markers is the early detection of recurrent disease in the follow-up of patients. In the study described here, we compared the usefulness of two serum markers for melanoma, 5-S-cysteinyldopa (5-S-CD) and melanoma inhibitory activity (MIA), in the monitoring of postsurgical melanoma patients. A total of 154 serum samples taken from 45 melanoma patients, who underwent surgery of the primary tumour and were under periodical follow-up for 13 to 180 months, were analysed. Serum MIA measurements were performed using an enzyme-linked immunosorbent assay (ELISA), and 5-S-CD levels were determined using high performance liquid chromatography (HPLC). In 72 serum samples taken from a group of 31 non-progressive patients with a median follow-up of 48.5 months, false positive rates of both markers were equally low, being 6.9% (five out of 72) for 5-S-CD and 8.3% (six out of 72) for MIA. In contrast, the sensitivity of MIA at the time point of the first clinical relapse in 14 progressive patients was significantly greater than that of 5-S-CD (0.64 [nine out of 14] versus 0.21 [three out of 14]; P < 0.05). Furthermore, seven patients showed abnormal serum MIA values 4-53 months prior to the clinical detection of metastasis, and the elevated levels were often noted on multiple occasions. These results show that MIA was superior to 5-S-CD in monitoring melanoma patients under periodical follow-up after the primary surgery. Repeated elevation of serum MIA levels may predict the presence of clinically undetectable occult metastases, which warrants further prospective investigations to assess the prognostic significance of serum MIA levels in postsurgical melanoma patients.

5-S-Cysteinyldopa (5-S-CD) has been used as a biochemical marker of melanoma progression. In this study, we measured serum levels of 5-S-CD in 2648 samples taken from 218 patients in order to evaluate the usefulness of this parameter in following melanoma progression and prognosis. 5-S-CD levels were significantly elevated above the upper limit of the normal range (10 nmol/l) in stage IV melanoma patients. The sensitivity of elevated serum 5-S-CD levels in detecting distant metastasis was 73%, while the specificity was 98% and the positive predictive value 94%. The sensitivity was improved to 77% when cases of amelanotic melanoma were excluded. Patients without metastases had elevated 5-S-CD values in 5% of the 1480 serum samples. Changes in serum 5-S-CD levels were followed during disease progression until the end stage in 49 patients. In 33% of the patients, elevation of serum 5-S-CD levels preceded clinical detection of visceral metastases, and in 37% elevation of 5-S-CD levels occurred at the same time as visceral metastasis. Patients with elevated 5-S-CD levels before or after surgical treatment had significantly shorter survival times than those with normal levels. These results show that the level of 5-S-CD in the serum is a sensitive and specific marker in predicting distant metastases. Elevated serum levels of 5-S-CD, before or after surgical treatment, is associated with a poor prognosis.

Patients with primary cutaneous melanoma underwent sentinel node (SN) mapping and biopsy at 25 facilities in Japan by the combination of radiocolloid with gamma probe and dye. Technetium-99m ((99m)Tc)-tin colloid, (99m)Tc-phytate, 2% patent blue violet (PBV) and 0.4% indigo carmine were used as tracers. In some hospitals, 0.5% fluorescent indocyanine green, which allows visualization of the SN with an infrared camera, was concomitantly used and examined. A total of 673 patients were enrolled, and 562 cases were eligible. The detection rates of SN were 95.5% (147/154) with the combination of tin colloid and PBV, 98.9% (368/372) with the combination of phytate and PBV, and 97.2% (35/36) with the combination of tin colloid or phytate and indigo carmine. SN was not detected in 12 cases by the combination method, and the primary tumor was in the head and neck in six of those 12 cases. In eight of 526 cases (1.5%), SN was detected by PBV but not by radiocolloid. There were 13 cases (2.5%) in which SN was detected by radiocolloid but not by PBV. In 18 of 36 cases (50%), SN was detected by radiocolloid but not by indigo carmine. Concomitantly used fluorescent indocyanine green detected SN in all of 67 cases. Interference with transcutaneous oximetry by PVB was observed in some cases, although it caused no clinical trouble. Allergic reactions were not reported with any of the tracers. (99m)Tc-tin colloid, (99m)Tc-phytate, PBV and indocyanine green are useful tracers for SN mapping.

We herein describe the case of a 68-year-old man who developed overt diabetes mellitus following the topical administration of dexamethasone 0.1%-containing ointment over a five-month period to treat oral lichen planus. The topical dexamethasone therapy was discontinued gradually, and the patient was subsequently treated with insulin for one month without clinical signs of overt adrenal insufficiency. An oral glucose tolerance test revealed impaired glucose tolerance after the treatment. The potential for the deterioration of glucose metabolism must be considered when patients with impaired glucose tolerance are treated with relatively low doses of topical corticosteroid ointment on the oral mucosa, even for short periods.

Nevus sebaceous is a congenital malformation of the skin within which a number of neoplasms showing adnexal differentiation may arise. Recently, deletions in the patched gene region were reported in nevus sebaceous and constitutive activation of the patched-hedgehog signaling pathway was implicated in the development of tumors arising within nevus sebaceous. To substantiate further a role of the patched-hedgehog signaling pathway in secondary tumors arising within nevus sebaceous, we examined 11 nevus sebaceous associated with secondary tumors for loss of heterozygosity of the patched gene region by microsatellite polymerase chain reaction and patched mRNA expression by in situ hybridization. Unexpectedly, however, none of the tumors (including eight trichoblastomas) and nevus sebaceous lesions showed loss of heterozygosity at any polymorphic loci close to the patched gene. Further more, none of the nevus sebaceous lesions and secondary tumors gave detectable signals for patched mRNA. In contrast, four of 11 sporadic basal cell carcinomas, that were examined for comparison, showed loss of heterozygosity at the patched gene locus (p0.05), and moderate to strong signals for patched mRNA was observed in all seven basal cell carcinoma tumors examined (p0.0001). Additional investigation by reverse transcription-polymerase chain reaction in four basal cell carcinomas and two nevus sebaceous tumors also showed the expression of Gli-1, another target gene in the patched-hedgehog signaling pathway, in all the basal cell carcinomas samples but not in any of the nevus sebaceous tumors examined. The findings in this study do not support the view that the deregulation of the patched-hedgehog signaling pathway is involved in the pathogenesis of nevus sebaceous and associated tumors, and show that, although morphologically similar, trichoblastomas and basal cell carcinomas have a different molecular pathogenesis.

In mouse the melanocortin 5 receptor is known to regulate sebaceous gland function. To clarify its role in man, we have studied melanocortin 5 receptor expression in skin, and allelic variation at the melanocortin 5 receptor locus in diverse human populations and candidate disease groups. Melanocortin 5 receptor protein and mRNA expression were studied by immunohistochemistry and reverse transcriptase polymerase chain reaction. Melanocortin 5 receptor mRNA was detected in normal skin and cultured keratinocytes but not in cultured fibroblasts or melanocytes. Immunohistochemistry revealed melanocortin 5 receptor immunoreactivity in the epithelium and appendages, including the sebaceous gland, eccrine glands, and apocrine glands, as well as low level expression in the interfollciular epidermis. In order to screen for genetic diversity in the melanocortin 5 receptor that might be useful for allelic association studies we sequenced the entire melanocortin 5 receptor coding region in a range of human populations. One nonsynonymous change (Phe209Leu) and four synonymous changes (Ala81Ala, Asp108Asp, Ser125Ser, and Thr248Thr) were identified. Similar results were found in each of the populations except for the Inuit in which only the Asp108Asp variant was seen. The apparent "global distribution" of melanocortin 5 receptor variants may indicate that they are old in evolutionary terms. Variation of melanocortin 5 receptor was examined in patients with acne (n = 21), hidradenitis supprativa (n = 4), and sebaceous gland lesions comprising sebaceous nevi, adenomas, and hyperplasia (n = 13). No additional mutations were found. In order to determine the functional status of the Phe209Leu change, increase in cAMP in response to stimulation with alpha-melanocyte-stimulating hormone was measured in HEK-293 cells transfected with either wild-type or the Phe209Leu variant. The variant melanocortin 5 receptor was shown to act in a concentration-dependent manner, which did not differ from that of wild type. We have therefore found no evidence of a causative role for melanocortin 5 receptor in sebaceous gland dysfunction, and in the absence of any association between variation at the locus and disease group, the pathophysiologic role of the melanocortin 5 receptor in man requires further study.

We report our experience of sentinel lymph node biopsy in malignant melanoma cousing patent blue V dye, preoperative lymphoscintigrapy and handheld gamma probe. Patients underwent preoperative lymphoscintigraphy with99mTc-labeled stannous phytate a day before operation. The patients subsequently underwent intraoperative lymphatic mapping and selective lymph node dissection with patent blue dye and radiopharmaceutical agent. A handheld gamma probe determined the radioactive counts over the blue stainned lymph node and the other nodes mapped by lymphoscintigraphy. The single injection of99mTc-labeled stannous phytate was effective in both preoperative lymphoscintigraphy and handheld gamma probeguided intraoperative determination of sentinel lymph nodes. The dissected lymph nodes were assessed by both intraoperative frozen section investigation and formaline-fixed section stained with hematoxylin and eosin and HMB-45. We failed to detect micrometastases by frozen section in 3 of 3 positive sentinel lymph nodes from 2 patients.Although the addition of radiopharmaceutical agents serves as a useful adjunct, the blue dye technique remains the criterion standard for selective lymph node dissection. The frozen section analysis is not recommended for determination of subsequent therapeutic lymph node dissection. [Skin Cancer (Japan) 2001; 16: 205-210]

Growth dependence with androgen (Nishi J Natl Cancer Inst 84: 519, 1992) and frequent expression of structurally unaltered androgen receptor (Fujimoto Lab Invest 80: 1465, 2000) in Extramammary Paget's disease (EMPD) may provide arational basis for hormone therapy. We report the first attempt of total androgen blockade (TAB) therapy in a 78-year-old man with advanced EMPD. The TAB therapy, which is widely used for prostate cancer, consisted. of oral administration of chlormadinone acetate 100mg daily and subcutaneous injection of leuprorelin acetate 3.75mg every month. Although skin metastases on his lower abdomen gradually progressed, para-trachial lymphadenopathy disappeared and para-aortic lymph nodes decreased in size by more than 60%. He died of the disease one year after starting the TAB therapy. Future clinical trial is required to establish the role of TAB therapy in the management of advanced stage EMPD patients. [Skin Cancer (Japan) 2001; 16: 247-250]

Extramammary Paget's disease (EMPD) is an uncommon neoplasm of the skin that shows differentiation to an apocrine sweat gland. Although we previously showed that erbB-2 overexpression may play a part in the progression of EMPD, molecular genetic defects underlying the development of EMPD are poorly understood. In the study described here, we examined androgen receptor expression and gene alterations in 30 cases of EMPD without internal malignancy. Immunohistochemistry revealed that 24 of 30 (80%) cases of EMPD variably expressed nuclear androgen receptor. Semi-quantitation of receptor content by scoring immunostained sections showed no difference between in situ (n = 17) and invasive (n = 13) EMPD tumors. Androgen receptor expression was also observed in four of six lymph node metastases. In these lymph nodes, expression of androgen receptor mRNA was confirmed by reverse transcriptase-polymerase chain reaction. Direct sequencing of exon 2 through exon 8, which encodes DNA- and hormone-binding domains of the androgen receptor gene, revealed no mutation in any of the 10 advanced stage tumors. Neither amplification nor deletion of the androgen receptor gene locus was detected by dual color fluorescence in situ hybridization analysis in 14 tumors. The present findings showing frequent expression of structurally unaltered androgen receptor in an advanced stage of EMPD may provide a rational basis for hormone therapy, which is widely used in the treatment of metastatic prostate cancer and androgen receptor-positive breast cancer recurrence.

乳房外パジェット病においてリンパ節転移の有無は予後を規定する重要な因子となる。自験例の解析により原発巣の浸潤レベルがリンパ節転移の程度や予後と相関することが示唆された。今後は多施設共同研究によりリンパ節転移と予後の関連をさらに細かく解析し, 転移陽性例における治療指針を示すことが必要であろう。

AIMS: To evaluate immunohistochemistry and reverse transcriptase polymerase chain reaction (RT-PCR) for melanoma associated antigens (MAA) in detecting occult melanoma cells in lymph nodes which were missed on routine pathology. METHODS: Occult melanoma cells were sought in 436 lymph nodes from 32 patients with cutaneous melanoma of the lower extremities by immunohistochemistry using the melanoma specific antibody HMB-45. The detection sensitivity of routine histology, immunohistochemistry, and RT-PCR was also compared in 23 lymph node samples from six patients. RESULTS: Immunohistochemistry showed that 15 of 24 patients (62.5%) who had no detectable metastasis by routine histology had at least one lymph node containing HMB-45 positive cells, mostly seen singly in the medullary sinus. No difference was found in known clinicopathological prognostic factors and recurrence rates between the two groups of patients with and without occult HMB-45 positive cells. RT-PCR analyses showed that the nested PCR for tyrosinase was more sensitive than a combination of single round PCR for five different MAA, including tyrosinase, MART-1/Melan A, Pmel-17, TRP-1, and TRP-2, detecting tyrosinase mRNA in six nodes which were negative by HMB-45 immunohistochemistry. CONCLUSIONS: Immunohistochemistry + RT-PCR is more sensitive than routine histology in detecting occult melanoma cells in lymph nodes. The nested PCR for tyrosinase should be used in future studies investigating the prognostic significance of such lymph node micrometastases.