Clinical versus Histological Assessment of Basal Cell Carcinoma Subtype and Thickness of Tumours Selected for Photodynamic Therapy.

Photodynamic therapy is an approved treatment for primary, superficial, and small nodular basal cell carcinomas with a thickness of < 2 mm located on low-risk sites. Histologically verified basal cell carcinomas clinically assessed as suited for photodynamic therapy were included. The study aimed to investigate the agreement between clinical and histological assessments of basal cell carcinoma subtypes and thickness of tumours selected for photodynamic therapy with histopathological evaluation as a reference. A total of 343 tumours were included. The agreement between clinical and histological diagnosis of basal cell carcinoma subtype was 72% (p < 0.001). Clinical assessment of subtype had a sensitivity of 93% and specificity of 55% for superficial tumours and a sensitivity of 55% and specificity of 85% for nodular tumours. The mean ± SD thickness values by clinical and histological assessments were 0.95 ± 0.53 and 0.86 ± 0.75. The difference of 0.09 mm was statistically significant (p = 0.017), but not considered to be clinically relevant, although the differences between specific subgroups could be relevant. Among basal cell carcinomas clinically diagnosed as superficial, 91% were histologically consistent with the current photodynamic therapy criteria. The main results suggest that histopathological evaluation should precede photodynamic therapy to ensure selection of suitable basal cell carcinomas. In selected cases, the clinical diagnosis alone may be adequate before proceeding with photodynamic therapy.

New, simplified versus standard photodynamic therapy (PDT) regimen for superficial and nodular basal cell carcinoma (BCC): A single-blind, non-inferiority, randomised controlled multicentre study.

BACKGROUND: Topical photodynamic therapy (PDT) is an approved and widely used treatment for low-risk basal cell carcinoma (BCC), comprising two sessions with an interval of 1 week. Simplification of the treatment course can be cost-effective, easier to organize, and cause less discomfort for the patients. METHODS AND FINDINGS: We performed an investigator-initiated, single-blind, non-inferiority, randomized controlled multicentre study with the objective of investigating whether a simpler and more flexible PDT regimen was not >10% less effective than the standard double PDT in the treatment of primary, superficial, and nodular ≤2 mm-thick BCC and evaluate the cosmetic outcome. With a non-inferiority margin of 0.1 and an expected probability complete response of 0.85, 190 tumours were required in each group. Histologically verified BCCs from seven centres in Norway were randomly assigned (1:1) to either receive a new regimen of single PDT with one possible re-treatment of non-complete responding tumours, or the standard regimen. The primary endpoint was the number of tumours with complete response or treatment failure at 36 months of follow-up, assessed by investigators blinded to the treatment regimen. Intention-to-treat and per-protocol analyses were performed. The cosmetic outcome was recorded. The study was registered with ClinicalTrials.gov, NCT-01482104, and EudraCT, 2011-004797-28. A total of 402 BCCs in 246 patients were included; 209 tumours assigned to the new and 193 to the standard regimen. After 36 months, there were 61 treatment failures with the new and 34 failures with the standard regimen. Complete response rate was 69.5% in the new and 81.1% in the standard treatment group. The difference was 11.6% (upper 97.5% CI 20.3), i.e. > than the non-inferiority margin of 10%. Cosmetic outcomes were excellent or good in 92% and 89% following the new and standard regimens, respectively. CONCLUSIONS: Single PDT with possible re-treatment of primary, superficial, and nodular ≤ 2-mm-thick BCC was significantly less effective than the approved standard double treatment. The cosmetic outcome was favorable and comparable between the two treatment groups.

Expression of ß-Catenin, E-Cadherin, and α-Smooth Muscle Actin in Basal Cell Carcinoma Before Photodynamic Therapy in Non-recurrent and Recurrent Tumors: Exploring the Ability of Predicting Photodynamic Therapy Outcome.

Photodynamic therapy (PDT) is an effective and cosmetically beneficial treatment of low-risk basal cell carcinomas (BCCs). To optimize PDT response, it is important to correctly select tumors. We sought to find markers that could identify such tumors beyond contributions from clinical and histological examination. Studies have shown that ß-catenin, E-cadherin, and α-smooth muscle actin (SMA) expression can indicate BCC aggressiveness/BCC invasiveness. We wanted to use these markers in an explorative study to investigate whether they were differently expressed among non-recurring compared with recurring BCCs, to evaluate their ability of predicting PDT outcome. Fifty-two BCCs were stained with antibodies against ß-catenin, E-cadherin, and α-SMA, and evaluated using immunoreactive score (IRS), subcellular localization, and stromal protein expression. Results showed that IRS of E-cadherin was significantly different among recurring compared with non-recurring BCCs and with area under a receiver operating characteristic curve of 0.71 (95% confidence interval: 0.56-0.86, p=0.025). Stromal ß-catenin expression significantly increased among recurring BCCs. Some recurring BCCs had intense expression in the deep invading tumor edge. In conclusion, E-cadherin, and stromal and deep edge ß-catenin expression were most prominent in BCCs that recurred post-PDT, suggesting they could potentially predict PDT outcome. Further studies are needed to investigate whether these results are of clinical value.

Basosquamous Basal Cell Carcinoma with Bone Marrow Metastasis.

Basal cell carcinoma (BCC) is the most common cancer in Caucasians. It is slow growing and rarely metastasizes. If left untreated over time, invasive growth can occur. We present a patient case with a primary BCC located in the right sub-mammary area, with extensive metastases to the skeleton and bone marrow. Histopathological examination of the tumour showed BCC with a diverse growth pattern. There were no signs of local metastases. Surgery was successfully performed. Three months post-surgery the patient developed normocytic anaemia and elevated inflammation markers. [18F]FDG PET/CT showed extensive FDG uptake in the entire skeleton and bone marrow. Biopsy confirmed the infiltration of BCC with similar histopathological features as the primary tumour. Prognosis of metastasized BCC is poor and, therefore, long-term follow-up of patients with risk factors is of importance.

Prognostic value of uPAR expression and angiogenesis in primary and metastatic melanoma.

Angiogenesis is important for the progression of cutaneous melanoma. Here, we analyzed the prognostic impact of the angiogenic factor urokinase plasminogen activator resecptor (uPAR), vascular proliferation index (VPI) and tumor necrosis as a measure of hypoxia in a patient series of nodular melanomas (n = 255) and matched loco-regional metastases (n = 78). Expression of uPAR was determined by immunohistochemistry and VPI was assessed by dual immunohistochemistry using Factor-VIII/Ki67 staining. Necrosis was recorded based on HE-slides. As novel findings, high uPAR expression and high VPI were associated with each other, and with increased tumor thickness, presence of tumor necrosis, tumor ulceration, increased mitotic count and reduced cancer specific survival in primary melanoma. In matched cases, VPI was decreased in metastases, whereas the frequency of necrosis was increased. Our findings demonstrate for the first time the impact on melanoma specific survival of uPAR expression and VPI in primary tumors, and of increased necrosis as an indicator of tumor hypoxia in loco-regional metastases. These findings support the importance of tumor angiogenesis in melanoma aggressiveness, and suggest uPAR as an indicator of vascular proliferation and a potential biomarker in melanoma.

Nestin expression is associated with aggressive cutaneous melanoma of the nodular type.

The intermediate filament nestin, a neural stem-cell marker, is reported to be expressed more strongly in melanomas compared with benign melanocytic lesions, and increasingly expressed in advanced melanoma stages. However, the prognostic impact of nestin on melanoma has not been well elucidated. The aim of the present study was to evaluate the prognostic influence of nestin expression in cutaneous melanoma in comparison with standard clinico-pathologic variables. In a large series of nodular cutaneous melanoma (n=348), nestin expression was assessed by immunohistochemistry using tissue microarray (TMA) sections. For comparison, nestin staining in corresponding metastases as well as in superficial spreading melanomas and benign nevi was also examined. Nestin was expressed to varying degrees in a majority of nodular melanomas (92%), and was significantly associated with increased tumor thickness, high mitotic count, and the presence of ulceration and tumor necrosis. Also, expression was stronger in the nodular type than in superficial spreading melanomas and benign nevi, but without significant difference when compared with matched metastases from the former. Importantly, strong expression of nestin was significantly associated with reduced survival in multivariate analysis. In conclusion, increased nestin expression was associated with aggressive melanoma features, with independent prognostic impact on multivariate survival analysis when compared with clinico-pathologic factors.

Malignant melanoma--diagnosis, treatment and follow-up in Norway.

BACKGROUND: The incidence of malignant melanoma in Norway is among the highest in the world and rising, with approximately 1 500 persons receiving the diagnosis annually. Correct surgical primary treatment cures 80-90%, while 10-20% experience relapses. The treatment of a metastatic malignant melanoma has changed considerably in the last 1-2 years as a result of clinical experience with new drugs. The current publication provides an updated overview of the treatment of malignant melanoma in Norway. METHOD: The article is based on a search in PubMed and on the authors' own research and clinical experience. RESULTS: After several decades with almost no change in the treatment of malignant melanoma, we have seen a positive development over the past couple of years. New treatment methods for malignant melanoma with distant spreading metastases have yielded favourable results in selected patients and are currently established in cancer departments in Norway. INTERPRETATION: Rapid and correct primary treatment is curing most patients with malignant melanoma. New drugs offer hope for selected patient groups with metastatic disease. Several new types of targeted treatment are being tested in clinical studies in Norway and elsewhere in the world.

Melanoma prone families with CDK4 germline mutation: phenotypic profile and associations with MC1R variants.

BACKGROUND: CDKN2A and CDK4 are high risk susceptibility genes for cutaneous malignant melanoma. Melanoma families with CDKN2A germline mutations have been extensively characterised, whereas CDK4 families are rare and lack a systematic investigation of their phenotype. METHODS: All known families with CDK4 germline mutations (n=17) were recruited for the study by contacting the authors of published papers or by requests via the Melanoma Genetics Consortium (GenoMEL). Phenotypic data related to primary melanoma and pigmentation characteristics were collected. The CDK4 exon 2 and the complete coding region of the MC1R gene were sequenced. RESULTS: Eleven families carried the CDK4 R24H mutation whereas six families had the R24C mutation. The total number of subjects with verified melanoma was 103, with a median age at first melanoma diagnosis of 39 years. Forty-three (41.7%) subjects had developed multiple primary melanomas (MPM). A CDK4 mutation was found in 89 (including 62 melanoma cases) of 209 tested subjects. CDK4 positive family members (both melanoma cases and unaffected subjects) were more likely to have clinically atypical nevi than CDK4 negative family members (p<0.001). MPM subjects had a higher frequency of MC1R red hair colour variants compared with subjects with one tumour (p=0.010). CONCLUSION: Our study shows that families with CDK4 germline mutations cannot be distinguished phenotypically from CDKN2A melanoma families, which are characterised by early onset of disease, increased occurrence of clinically atypical nevi, and development of MPM. In a clinical setting, the CDK4 gene should therefore always be examined when a melanoma family tests negative for CDKN2A mutation.

Tumor necrosis is a prognostic factor in thick cutaneous melanoma.

The significance of tumor necrosis in cutaneous melanoma has not been well elucidated. The purpose of this study was to explore the prognostic impact of necrosis in comparison with other known clinicopathologic factors in these tumors. Initially, 457 consecutive cases of nodular cutaneous melanoma (1981 to 2008) were included in this series. Tumor necrosis was assessed on hematoxylin and eosin-stained sections and was recorded as significant when an area of at least a quarter of a high-power field (×400; 0.07 mm) was occupied by necrotic cells and as sparse when clusters of at least 5 necrotic cells were observed. Tumor necrosis (26% of the cases) was associated with increased tumor thickness, high mitotic count, presence of tumor ulceration, and decreased survival. Stratified analyses (univariate and multivariate) with standard microscopic variables indicated the strongest prognostic influence of necrosis in tumors thicker than 4 mm. Notably, in the stratum of pT4 tumors, presence of necrosis was a stronger prognostic predictor than was ulceration. Tumor necrosis was a significant prognostic indicator providing additional information to established predictors of patient outcome in this series of nodular cutaneous melanoma, predominantly among thick tumors (>4 mm). Presence of necrosis was a stronger indicator for worse outcome compared with ulceration in pT4 tumors.

Suppression of heat shock protein 27 induces long-term dormancy in human breast cancer.

The mechanisms underlying tumor dormancy have been elusive and not well characterized. We recently published an experimental model for the study of human tumor dormancy and the role of angiogenesis, and reported that the angiogenic switch was preceded by a local increase in VEGF-A and basic fibroblast growth factor. In this breast cancer xenograft model (MDA-MB-436 cells), analysis of differentially expressed genes revealed that heat shock protein 27 (HSP27) was significantly up-regulated in angiogenic cells compared with nonangiogenic cells. The effect of HSP27 down-regulation was further evaluated in cell lines, mouse models, and clinical datasets of human patients with breast cancer and melanoma. Stable down-regulation of HSP27 in angiogenic tumor cells was followed by long-term tumor dormancy in vivo. Strikingly, only 4 of 30 HSP27 knockdown xenograft tumors initiated rapid growth after day 70, in correlation with a regain of HSP27 protein expression. Significantly, no tumors escaped from dormancy without HSP27 expression. Down-regulation of HSP27 was associated with reduced endothelial cell proliferation and decreased secretion of VEGF-A, VEGF-C, and basic fibroblast growth factor. Conversely, overexpression of HSP27 in nonangiogenic cells resulted in expansive tumor growth in vivo. By clinical validation, strong HSP27 protein expression was associated with markers of aggressive tumors and decreased survival in patients with breast cancer and melanoma. An HSP27-associated gene expression signature was related to molecular subgroups and survival in breast cancer. Our findings suggest a role for HSP27 in the balance between tumor dormancy and tumor progression, mediated by tumor-vascular interactions. Targeting HSP27 might offer a useful strategy in cancer treatment.

Prognostic importance of the mitotic marker phosphohistone H3 in cutaneous nodular melanoma.

Mitotic count is a known prognostic predictor in cutaneous melanoma, and is included in the current American Joint Committee on Cancer tumor-node-metastasis (TNM) staging system. The mitotic marker phosphohistone H3 (PHH3) is considered to facilitate counting of mitosis, and the purpose of this study was to evaluate the prognostic significance and strength of PHH3 in comparison with standard mitotic counting in cutaneous malignant melanoma. A total of 457 consecutive cases of nodular cutaneous melanoma were initially included in this series. The mitotic count was assessed on hematoxylin and eosin sections, and PHH3 was then examined by immunohistochemistry on standard sections of paraffin-embedded tumor tissue. Both the mitotic count and the number of PHH3-stained mitotic figures were recorded in a minimum area of 1 mm(2). Increased mitotic count and PHH3 value were both associated with unfavorable features like tumor thickness and presence of ulceration. Univariate survival analysis showed a highly significant prognostic impact of mitotic count and PHH3, whereas multivariate analysis indicated PHH3 to be a stronger prognostic indicator than mitotic count. Assessment of mitotic activity by PHH3 immunostaining might have important practical advantages, and should be further studied to consider a place in routine examination of all cutaneous melanomas.

Ki-67 expression is superior to mitotic count and novel proliferation markers PHH3, MCM4 and mitosin as a prognostic factor in thick cutaneous melanoma.

BACKGROUND: Tumor cell proliferation is a predictor of survival in cutaneous melanoma. The aim of the present study was to evaluate the prognostic impact of mitotic count, Ki-67 expression and novel proliferation markers phosphohistone H3 (PHH3), minichromosome maintenance protein 4 (MCM4) and mitosin, and to compare the results with histopathological variables. METHODS: 202 consecutive cases of nodular cutaneous melanoma were initially included. Mitotic count (mitosis per mm2) was assessed on H&E sections, and Ki-67 expression was estimated by immunohistochemistry on standard sections. PHH3, MCM4 and mitosin were examined by staining of tissue microarrays (TMA) sections. RESULTS: Increased mitotic count and elevated Ki-67 expression were strongly associated with increased tumor thickness, presence of ulceration and tumor necrosis. Furthermore, high mitotic count and elevated Ki-67 expression were also associated with Clark's level of invasion and presence of vascular invasion. High expression of PHH3 and MCM4 was correlated with high mitotic count, elevated Ki-67 expression and tumor ulceration, and increased PHH3 frequencies were associated with tumor thickness and presence of tumor necrosis. Univariate analyses showed a worse outcome in cases with elevated Ki-67 expression and high mitotic count, whereas PHH3, MCM4 and mitosin were not significant. Tumor cell proliferation by Ki-67 had significant prognostic impact by multivariate analysis. CONCLUSIONS: Ki-67 was a stronger and more robust prognostic indicator than mitotic count in this series of nodular melanoma. PHH3, MCM4 and mitosin did not predict patient survival.

Tumor necrosis is associated with increased alphavbeta3 integrin expression and poor prognosis in nodular cutaneous melanomas.

BACKGROUND: Tumor necrosis and apoptotic activity are considered important in cancer progression, but these features have not been much studied in melanomas. Our hypothesis was that rapid growth in cutaneous melanomas of the vertical growth phase might lead to tissue hypoxia, alterations in apoptotic activity and tumor necrosis. We proposed that these tumor characteristics might be associated with changes in expression of cell adhesion proteins leading to increased invasive capacity and reduced patient survival. METHODS: A well characterized series of nodular melanoma (originally 202 cases) and other benign and malignant melanocytic tumors (109 cases) were examined for the presence of necrosis, apoptotic activity (TUNEL assay), immunohistochemical expression of hypoxia markers (HIF-1 alpha, CAIX, TNF-alpha, Apaf-1) and cell adhesion proteins (alphavbeta3 integrin, CD44/HCAM and osteopontin). We hypothesized that tumor hypoxia and necrosis might be associated with increased invasiveness in melanoma through alterations of tumor cell adhesion proteins. RESULTS: Necrosis was present in 29% of nodular melanomas and was associated with increased tumor thickness, tumor ulceration, vascular invasion, higher tumor proliferation and apoptotic index, increased expression of alphavbeta3 integrin and poor patient outcome by multivariate analysis. Tumor cell apoptosis did also correlate with reduced patient survival. Expression of TNF-alpha and Apaf-1 was significantly associated with tumor thickness, and osteopontin expression correlated with increased tumor cell proliferation (Ki-67). CONCLUSION: Tumor necrosis and apoptotic activity are important features of melanoma progression and prognosis, at least partly through alterations in cell adhesion molecules such as increased alphavbeta3 integrin expression, revealing potentially important targets for new therapeutic approaches to be further explored.

Loss of BMI-1 expression is associated with clinical progress of malignant melanoma.

BMI-1 is a member of the Polycomb group of genes (PcGs) and is involved in embryonic gene regulation and maintenance of adult stem cells. It has been suggested that BMI-1 protein is important in cell cycle regulation, since both p16/INK4a and p14/ARF are downstream BMI-1 targets. BMI-1 has been implicated in the development and progression of several malignancies, but its role in melanocytic tumors of the skin is unknown. In the present study, using immunohistochemistry on 178 benign and malignant melanocytic lesions and two different antibodies, BMI-1 expression was reduced in melanomas compared with benign nevi. In established melanomas, loss of BMI-1 expression was associated with features of aggressive tumors, such as increased tumor cell proliferation, presence of necrosis and increased expression of both N-cadherin and beta3-integrin, indicating a more invasive and mesenchymal phenotype. Low BMI-1 expression was associated with low p14 and CDK4 but not with p16 expression. Low levels of BMI-1 expression were also significantly associated with decreased patient survival.

Mutation analysis of the EGFR-NRAS-BRAF pathway in melanomas from black Africans and other subgroups of cutaneous melanoma.

Earlier studies have shown frequent mutations in the BRAF and NRAS genes in cutaneous melanoma, but these alterations have not been examined in the rare category of melanoma from black Africans. Moreover, the frequency of epidermal growth factor receptor (EGFR) mutations in melanocytic tumors is not known. We therefore examined 165 benign and malignant melanocytic lesions (including 118 invasive melanomas and 18 metastases collected as consecutive cases from various time periods and from two different pathology departments; the 51 nodular melanomas were randomly selected from a larger, consecutive, population-based series of nodular melanomas) with respect to alterations in the EGFR, BRAF and NRAS genes. Mutations in EGFR (exons 18-21) were not detected. EGFR protein expression was observed in a subgroup of melanomas, but without associations with clinicopathologic phenotype or prognosis. Cytoplasmic EGFR expression was, however, significantly increased from benign nevi to melanomas. Mutations in BRAF and NRAS were detected in superficial melanoma (25 and 29%, respectively), nodular melanoma (29 and 28%, respectively) and lentigo maligna melanoma (15 and 16%, respectively). In a series of melanomas from black Africans (n=26), only two BRAF mutations (8%) were found, both being different from the common T1799A substitution. Moreover, melanomas from black Africans exhibited mutations in NRAS exon 1 only (12%), whereas NRAS exon 2 mutations were predominant in melanomas from Caucasians. Thus, the frequencies of BRAF and NRAS mutations were particularly low in melanomas from black Africans, supporting a different pathogenesis of these tumors.

Five-year follow-up of a randomized, prospective trial of topical methyl aminolevulinate photodynamic therapy vs surgery for nodular basal cell carcinoma.

OBJECTIVE: To compare 5-year lesion recurrence rates in primary nodular basal cell carcinoma treated with topical methyl aminolevulinate photodynamic therapy (PDT) or simple excision surgery. DESIGN: Prospective, randomized, multicenter study. SETTING: University hospital dermatology departments. PATIENTS: A total of 97 patients, 50 with 53 lesions treated with methyl aminolevulinate PDT and 47 with 52 lesions treated by excision surgery, were included in the per protocol analysis. Of the lesions treated with methyl aminolevulinate PDT and surgery, 49 and 52, respectively, showed complete clinical response at 3 months after treatment and were observed for long-term outcome evaluation. INTERVENTIONS: Topical methyl aminolevulinate cream, 160 mg/g, applied for 3 hours before illumination (75 J/cm(2) of red light at 570 to 670 nm) on 2 or 4 occasions (12 [23%] of 53 lesions); or excision surgery. MAIN OUTCOME MEASURES: Histologically confirmed lesion recurrence, sustained lesion complete response rate (time-to-event analysis), and investigator assessment of cosmetic outcome, 5 years after the last treatment. RESULTS: At 5 years, recurrence was documented in 7 (14%) of 49 lesions (95% confidence interval [CI], 6%-27%) treated with methyl aminolevulinate PDT vs 2 (4%) of 52 lesions (95% CI, 1%-13%) treated with excision surgery (P = .09). Estimated sustained lesion complete response rates were 76% (95% CI, 59%-87%) and 96% (95% CI, 84%-99%), respectively (P = .01). More patients treated with methyl aminolevulinate PDT than surgery had an excellent or good cosmetic outcome: 27 (87%) of 31 patients (95% CI, 70%-96%) vs 19 (54%) of 35 patients (95% CI, 37%-71%) (P = .007). CONCLUSIONS: Long-term follow-up indicates superior efficacy of surgery to methyl aminolevulinate PDT in nodular basal cell carcinoma. However, methyl aminolevulinate PDT is also an effective treatment for this indication and exhibits a more favorable cosmetic outcome.

EZH2 expression is associated with high proliferation rate and aggressive tumor subgroups in cutaneous melanoma and cancers of the endometrium, prostate, and breast.

PURPOSE: EZH2 is a member of the polycomb group of genes and important in cell cycle regulation. Increased expression of EZH2 has been associated previously with invasive growth and aggressive clinical behavior in prostate and breast cancer, but the relationship with tumor cell proliferation has not been examined in human tumors. The purpose of this study was to validate previous findings in a population-based setting, also including tumors that have not been studied previously. PATIENTS AND METHODS: In our study of nearly 700 patients, we examined EZH2 expression and its association with tumor cell proliferation and other tumor markers, clinical features, and prognosis in cutaneous melanoma and cancers of the endometrium, prostate, and breast. RESULTS: Strong EZH2 expression was associated with increased tumor cell proliferation in all four cancer types. Associations were also found between EZH2 and important clinicopathologic variables. EZH2 expression showed significant prognostic impact in melanoma, prostate, and endometrial carcinoma in univariate survival analyses, and revealed independent prognostic importance in carcinoma of the endometrium and prostate. CONCLUSION Our findings point at EZH2 as a novel and independent prognostic marker in endometrial cancer, and validate previous findings on prostate and breast cancer. Further, EZH2 expression was associated with features of aggressive cutaneous melanoma. The fact that EZH2 might identify increased tumor cell proliferation and aggressive subgroups in several cancers may be of practical interest because the polycomb group proteins have been suggested as candidates for targeted therapy. EZH2 expression should, therefore, be further examined as a possible predictive factor.

Importance of P-cadherin, beta-catenin, and Wnt5a/frizzled for progression of melanocytic tumors and prognosis in cutaneous melanoma.

PURPOSE: It has been proposed that melanoma cells shift from E-cadherin to N-cadherin expression during tumor development, and recent gene profiling has shown increased expression of Wnt5a/Frizzled in aggressive melanomas possibly by interactions with beta-catenin. We therefore wanted to investigate the role of cadherin subtypes, beta-catenin, and Wnt5a/Frizzled in melanocytic tumors, with focus on prognosis in nodular melanomas. EXPERIMENTAL DESIGN: The immunohistochemical expression of E-cadherin, N-cadherin, P-cadherin, beta-catenin, and Wnt5a/Frizzled was examined using tissue microarrays of 312 melanocytic tumors. RESULTS: Cytoplasmic expression of P-cadherin was associated with increasing tumor thickness (P=0.005) and level of invasion (P=0.019), whereas membranous staining was associated with thinner (P=0.012) and more superficial (P=0.018) tumors. Increased cytoplasmic P-cadherin was associated with reduced survival (P=0.047). Lack of nuclear beta-catenin expression was related to increased tumor thickness (P=0.002) and poor patient survival in univariate (P=0.0072) and multivariate (P=0.004) analyses. Membranous expression of N-cadherin was significantly increased from primary tumors to metastatic lesions, whereas E-cadherin staining tended to be decreased. Wnt5a and its receptor Frizzled were highly coexpressed, and nuclear expression of both markers was significantly reduced from benign nevi to melanomas, with a shift from nuclear to cytoplasmic expression in malignant tumors. In addition, Wnt5a expression was significantly associated with nuclear beta-catenin expression. CONCLUSIONS: Alterations in the expression and subcellular localization of cell adhesion markers are important in the development and progression of melanocytic tumors, and strong cytoplasmic P-cadherin expression and loss of nuclear beta-catenin staining were associated with aggressive melanoma behavior and reduced patient survival.

BRAF and NRAS mutations are frequent in nodular melanoma but are not associated with tumor cell proliferation or patient survival.

Previous studies have shown frequent mutations in the BRAF (V-raf murine sarcoma viral oncogene homolog B1) or NRAS (neuroblastoma RAS viral [V-ras] oncogene homolog) genes in cutaneous melanoma, but the relationship between these alterations and tumor cell proliferation has not been examined in human melanoma. In our study of 51 primary nodular melanomas and 18 paired metastases, we found mutations in BRAF (codon 600, previously denoted 599) in 15 primary tumors (29%) and eight metastases (44%). The figures for NRAS mutations were 27% and 22%, respectively. Mutations in BRAF and NRAS genes were mutually exclusive in all but one case, and were maintained from primary tumors through their metastases. Mutations, however, were not associated with tumor cell proliferation by Ki-67 expression, tumor thickness, microvessel density, or vascular invasion, and there were no differences in patient survival. Although BRAF and NRAS mutations are likely to be important for the initiation and maintenance of some melanomas, other factors might be more significant for proliferation and prognosis in subgroups of aggressive melanoma.