Potentially useful markers for desmoplastic melanoma: an analysis of KBA.62, p-AKT, and ezrin

Desmoplastic melanoma (DM) represents only a small portion of newly diagnosed melanoma cases but can be a significant diagnostic challenge for dermatopathologists. Immunohistochemical stains are often useful to confirm the diagnosis; however, DM is notorious for not expressing most of the melanocytic markers other than S100. Recent studies have suggested several new markers, which may be promising in the diagnosis of DM. KBA.62 is a relatively new antimelanoma monoclonal antibody, which has not been well characterized in DM. Ezrin and p-Akt are additional markers, which have been shown to be involved in cell survival and proliferation. We collected 12 cases of DM and 18 cases of other lesions that could be included in the differential diagnosis. The H&E sections were reviewed, and immunohistochemical stains for KBA.62, p-AKT, and Ezrin were performed. Seventy-five percent of the DM cases (9 of 12) demonstrated positive staining (>5% of tumor cells staining) with KBA.62, with an average of 39% of cells staining. One hundred percent of the DM cases (12 of 12) demonstrated positive staining with Ezrin, with an average of 49% of tumor cells staining. Seventy-five percent of the DM cases (9 of 12) demonstrated positive staining with p-Akt, with an average of 49% of cells staining. KBA.62 and Ezrin demonstrated statistically significant increased staining of DM cases compared with the other lesions (P = 0.05 and P = 0.007, respectively), although it was not useful in distinguishing DM versus malignant peripheral nerve sheath tumor, whereas p-Akt showed no significant differences in staining between DM and the other cases. These findings suggest that KBA.62 may be a useful marker in confirming the diagnosis of DM.     

Follow-up of melanocytic skin lesions with digital epiluminescence microscopy: patterns of modifications observed in early melanoma, atypical nevi, and common nevi

BACKGROUND: Digital epiluminescence microscopy (DELM) has been reported to be a useful technique for the follow-up of melanocytic nevi. One of the promises of this technique is to identify modifications over time that indicate impending or incipient malignancy and to facilitate surveillance of melanocytic skin lesions, particularly in patients with multiple clinically atypical nevi. OBJECTIVE: Our purpose was to report on patterns of modifications over time observed in benign melanocytic skin lesions and melanoma. METHODS: A total of 1862 sequentially recorded DELM images of melanocytic lesions from 202 patients (mean age, 36.1 years; 54.0% female patients) with multiple clinically atypical nevi were included in the analysis. The median follow-up interval was 12. 6 months. Melanocytic lesions with substantial modifications over time (enlargement, changes in shape, regression, color changes or appearance of ELM structures known to be associated with melanoma) were excised and referred to histopathologic examination. RESULTS: A total of 75 melanocytic skin lesions (4.0%) from 52 patients (mean age, 33.3 years; 63.5% female patients) showed substantial modifications over time and were excised and referred to histopathologic examination. Eight changing lesions were histologically diagnosed as early melanomas. These lesions frequently showed focal enlargement associated with a change in shape as well as appearance of ELM structures that are known to be associated with melanoma. In contrast, the majority of benign changing lesions (common and atypical nevi) showed symmetric enlargement without substantial structural ELM changes. Six of the 8 patients in whom melanoma developed were unaware of the fact that the lesion had changed over time. CONCLUSION: We demonstrate that follow-up of melanocytic lesions with DELM helps to identify patterns of morphologic modifications typical for early melanoma. DELM may therefore serve as a useful tool to improve the surveillance of patients with multiple atypical nevi.     

干细胞标志物CD133、nestin和CD44在恶性黑素瘤皮损中的表达及意义

目的 探讨干细胞标志物CD133、nestin和CD44在恶性黑素瘤中的表达及意义.方法 采用免疫组化SP法检测3种标记物在30例恶性黑素瘤及30例皮内痣皮损中的表达.结果 CD133、nestin和CD44在恶性黑素瘤组织中的阳性表达率分别是53.33%,80.00%和20.00%,在皮内痣分别是23.33%,53.33%和0,两组各标志物阳性率差异比较均有统计学意义(P＜0.05).CD133、nestin和CD44在恶性黑素瘤组织中阳性细胞率平均值分别为2.98%±5.62%,34.92%±34.89%和1.28%±3.26%,在皮内痣分别为0.10%±0.21%,7.26%±13.13%和0,两组各标志物差异比较均有统计学意义(P＜0.05).两组中阳性细胞率与患者性别、年龄、病程比较,差异无统计学意义(P＞0.05).结论 干细胞标志物CD133、nestin和CD44在恶性黑素瘤中高表达,在皮内痣中低表达或不表达,提示恶性黑素瘤中可能存在肿瘤干细胞.     

Digital dermoscopy analysis for the differentiation of atypical nevi and early melanoma: a new quantitative semiology

OBJECTIVES: To use a digital dermoscopy analyzer with a series of "borderline" pigmentary skin lesions (ie, clinically atypical nevi and early melanoma) to find correlation between the studied variables and to determine their discriminating power with respect to histological diagnosis. DESIGN: A total of 147 pigmentary skin lesions were histologically examined by 3 experienced dermatopathologists and identified as nevi (n = 90) and melanomas (n = 57). The system evaluated 36 variables to be studied as possible discriminant variables, grouped into 4 categories: geometries, colors, textures, and islands of color. SETTING: University medical department. PATIENTS: A sample of patients with excised pigmentary skin lesions (nevi and melanomas). MAIN OUTCOME MEASURES: Sensitivity, specificity, and accuracy of the model for evaluating "borderline" pigmentary skin lesions. RESULTS: After multivariate stepwise discriminant analysis, only 13 variables were selected to compute the canonical discriminant function. CONCLUSION: The present method made it possible to determine which objective variables are important for distinguishing atypical benign pigmentary skin lesions and early melanoma.     

Risks and benefits of sequential imaging of melanocytic skin lesions in patients with multiple atypical nevi.

OBJECTIVE: To evaluate the utility of sequential imaging of melanocytic skin lesions. DESIGN: With the use of a computerized test environment, digital images of 80 melanocytic skin lesions (including 10 early melanomas) were presented to 24 dermatologists with different levels of experience in 3 sessions. The 3 sessions were designed to simulate the decision-making process (1) without the possibility of follow-up, (2) with the possibility of follow-up, and (3) after presentation of follow-up images. MAIN OUTCOME MEASURES: Diagnostic performance in terms of sensitivity, specificity, accuracy, treatment threshold, and utility. RESULTS: The possibility of follow-up increased the treatment threshold in all groups of dermatologists compared with decision making without the possibility of follow-up. The increase of the treatment threshold was accompanied by a loss of sensitivity and a gain in specificity. The overall diagnostic accuracy remained unchanged. After presentation of follow-up images, the diagnostic accuracy improved significantly. The sensitivity improved for all readers, but the specificity improved only for the most experienced readers. The utility of sequential imaging depended on the compliance of patients with follow-up. Under the assumption that all patients are compliant with follow-up, the utility of sequential imaging was superior to decision making without follow-up over a broad range of benefit-risk ratios. CONCLUSIONS: Sequential imaging of melanocytic skin lesions is a useful procedure for patients with multiple atypical nevi. Uncritical use of sequential imaging cannot be recommended, because the utility of this technique depends on the experience in the interpretation of follow-up images and on the patient's compliance with follow-up.     

microRNA在人皮肤黑素瘤与色素痣中共同差异表达的研究

目的 筛选人皮肤黑素瘤组织中表达的已知hsa-microRNA。方法 采用SBC microRNA芯片技术,将6例人皮肤黑素瘤组织总RNA分别与9例健康人色素痣总RNA混合物对比,筛选已知的468条hsa-microRNA表达情况。用qPCR分别检测这6例人皮肤黑素瘤组织中各候选hsa-microRNA的表达。将芯片和qPCR两种方法检测结果一致的候选hsa-microRNA确定为有意义的共同差异表达hsa-microRNA。结果 2倍以上差异表达miRNA占所检测miRNA的12.18% ～ 86.33%,5倍以上差异表达miRNA占1.28% ～ 19.02%,10倍以上差异表达miRNA占0.43% ～ 5.34%。 此6例人皮肤黑素瘤中miRNA-21 明显上调表达,miRNA-320和miRNA-494明显下调表达。结论 人皮肤黑素瘤组织中miRNA-21明显上调表达,miRNA-320和miRNA-494明显下调表达。     

Increased intraepidermal melanocyte frequency and size in dysplastic melanocytic nevi and cutaneous melanoma. A comparative quantitative study of dysplastic melanocytic nevi, superficial spreading melanoma, nevocellular nevi, and solar lentigines

Dysplastic melanocytic nevi (DMN) are distinguished histologically by a hyperplasia of variably atypical intraepidermal melanocytes in a lentiginous epidermal pattern. In order to further characterize the intraepidermal melanocytes of DMN, 4 representative specimens each of DMN, acquired nevocellular nevi (NCN), solar lentigines (SL), and superficial spreading melanoma (SSM) were selected on the basis of predetermined criteria, confirmed in a blind histologic assessment, and compared in a quantitative morphologic study using 6 micron-thick hematoxylin and eosin stained sections of L-dihydroxyphenylalanine (dopa) preincubated vertical tissue slices of lesion and adjacent normal skin. The average melanocyte frequency, expressed as the percent of dopa-reactive perikarya among 600 consecutive basal unit cells, was significantly greater in DMN (60 +/- 23%) than in NCN (18 +/- 3%), SL (25 +/- 7%), and adjacent skin (14 +/- 3%), but similar to that in SSM (71 +/- 11%). The average mean diameter of 200 consecutive epidermal basal unit melanocytes was significantly larger in DMN (11 +/- 2 microns) than in NCN (7 +/- 0.4 microns), SL (6 +/- 0.1 microns), and adjacent skin (6 +/- 0.4 microns), but significantly smaller than in SSM (16 +/- 3 microns). The observed similarities of intraepidermal melanocytes in selected DMN and SSM, as well as distinct differences from melanocytes in selected NCN and SL, support the hypothesis that some varieties of DMN may represent potential precursors of cutaneous melanoma.     

Histochemical determinations of copper, zinc, and iron in pigmented nevi and melanoma.

Histochemical determinations of copper, zinc, and iron in intradermal pigmented nevi and melanomas revealed the presence of copper and iron in melanoma but not in nevi. Zinc was not detected in either melanomas or nevi. However, melanin was removed from the tissues prior to staining; therefore, it is possible that zinc was also removed by the procedure. Although the function of copper and iron in the melanoma cell is not known, they may be components of abnormal enzymes.     

Comprehensive analysis of 112 melanocytic skin lesions demonstrates microsatellite instability in melanomas and dysplastic nevi, but not in benign nevi

INTRODUCTION: the length of DNA repetitive sequences (microsatellite instability (MSI)) represent distinct tumorigenic pathways associated with several familial and sporadic tumors. MATERIAL AND METHODS: To investigate the prevalence and frequency of MSI in melanocytic lesions, the polymerase chain reaction (PCR)-based microsatellite assay was used to examine formalin-fixed, paraffin-embedded tissues of 30 benign melanocytic nevi, 60 melanocytic dysplastic nevi (MDN), and 22 primary vertical growth phase cutaneous malignant melanomas (CMM). Twenty-four microsatellite markers at the 1p, 2p, 3p, 4q and 9p chromosomal regions were used. RESULTS: MSI was found at 1p and 9p in MDN and CMM but not in benign melanocytic nevi. The overall prevalence of MSI was 17/60 (28%) in MDN and 7/22 (31%) in CMM. The frequency of MSI ranged from 2/24 (9%) to 4/24 (17%) and was most commonly found at D9S162. There was a statistically significant correlation between degree of atypia and frequency of MSI (p<0.001) in MDN. There were two MSI banding patterns: band shifts and additional bands. CONCLUSIONS: The data presented revealed the presence of low-frequency MSI (MSI-L) at the 1p and 9p regions in both MDN and CMM. Whether the MSI-L pattern reflects a defect in mismatch repair genes is still to be determined.     

Malignant melanoma arising from nevi, p53, p16, and Bcl-2: expression in benign versus malignant components

BACKGROUND: The etiology of malignant melanoma has been the subject of much study. Tumour suppressor genes p53 and p16 and the antiapoptotic, Bcl-2, are implicated in the development and progression of malignant melanoma. OBJECTIVE: To compare the expression of p53, p16, and the Bcl-2 genes in both benign and malignant components of malignant melanoma arising from pre-existing nevocellular nevi. METHODS: Twenty cases of malignant melanoma arising from pre-existing nevi were selected and studied by immunohistochemistry for the expression of p53 (D07) CDK4I/MTS-1/INK <4, which detects both wild and mutant type, p16 CDK4I/MTS-1/INK <4, and Bcl-2 using an avidin-biotin technique on formalin-fixed, paraffin-embedded sections. RESULTS: Fifteen cases demonstrated p53 immunoreactivity in the malignant components ranging from 5 to 20% with no expression being seen in the benign components. The p16 gene showed strong nuclear reactivity in the benign components of 14 cases and weak reaction in 6 cases; the malignant components expressed weak nuclear staining in 18 cases and cytoplasmic staining in all cases. The Bcl-2 gene was expressed strongly to moderately in benign components and weakly in malignant components of nine cases, and was negative in 11 cases. CONCLUSION: Immunostaining for p53 is expressed only in the malignant component, whereas p16 and Bcl-2 showed decreased staining and a different pattern in malignant compared with benign components. These findings suggest that expression and alterations in the subcellular localization of the cell cycle regulators may contribute to the mechanism of tumourigenesis.     

Tissue microarray analysis of RANKL in cutaneous lupus erythematosus and psoriasis

Recently, it was discovered that the receptor activator of nuclear factor κB (RANK)/RANK ligand (RANKL) is part of an important signal transduction pathway for tissue homoeostasis. Therefore, we were interested in investigating RANKL expression in the epidermis of skin lesions from patients with different subtypes of cutaneous lupus erythematosus (CLE) and psoriasis as well as normal healthy donors. Using the tissue microarray technique, skin biopsy specimens were evaluated by immunohistochemistry. RANKL showed a significantly increased expression in the epidermis of skin biopsy specimens from patients with psoriasis (median: 4, range: 0-5) compared to patients with CLE (median: 0, range: 0-4) (P<0.001). No significant differences in epidermal RANKL expression between the CLE subtypes were detected. These data show a different expression of RANKL in the epidermis of skin lesions from patients with CLE compared to those with psoriasis suggesting that RANKL might play an important role in the pathogenesis of the disease.     

CD10, p63 and CD99 expression in the differential diagnosis of atypical fibroxanthoma,spindle cell squamous cell carcinoma and desmoplastic melanoma

Background: Atypical fibroxanthoma (AFX) is a pleomorphic spindle cell lesion of the skin; it is considered in the differential diagnosis with spindle cell malignant melanoma (MM) and sarcomatoid carcinoma/spindle cell squamous cell carcinoma (SCC). An optimum approach has yet to fully emerge with respect to the immunohistochemical discrimination of these lesions. Methods: Departmental archives from 1978 onwards were searched for clinicopathologically confirmed cases of AFX, MM and SCC. Immunostains for CD10, CD99 and p63 were performed in each case. Scored staining results were analyzed using Fisher's Exact Test. Results: Twenty‐seven of 31 cases of AFX were positive for CD10, as compared with 3 of 22 SCCs and 0 of 20 MMs. CD10 positivity was preferentially associated with the diagnosis of AFX (p < 0.001). p63 reactivity was observed in 15/22 cases of SCCs, 5/31 AFXs and 1/20 MMs. CD99 reactivity was observed in 3/31 cases of AFX, 2/22 SCCs and 3/20 MMs. Conclusion: CD10 positivity is relatively specific in this context for the diagnosis of AFX. Its utility is enhanced when only strong, diffuse membranocytoplasmic staining is considered as a positive result. In contrast to prior reports, p63 was not found to be highly sensitive for SCC. Similarly, CD99 showed no preferential staining of any single diagnostic group of lesions. Kanner WA, Brill LB, Patterson JW, Wick MR. CD10, p63 and CD99 expression in the differential diagnosis of atypical fibroxanthoma, spindle cell squamous cell carcinoma and desmoplastic melanoma.     

Precursors of malignant melanoma. Problems in computing the risk of malignant melanoma arising in dysplastic and congenital nevocytic nevi

It has recently been shown that both dysplastic and congenital nevi are precursors to malignant melanoma. These findings are based upon mathematical models that show an increased risk of the nevi evolving into melanoma over random choice. However, problems exist with these models that may invalidate their results. The recommendation to remove dysplastic and congenital nevi prophylactically based upon models such as these is premature.     

Nucleolar organizer regions in malignant melanoma and melanocytic nevi. Comparison of two counting methods

Using a silver staining technique, we studied nucleolar organizer regions (AgNOR) in paraffin sections of junctional nevi, compound nevi, intradermal nevi, blue nevi, dysplastic nevi, Spitz nevi, lentigo maligna, malignant melanomas in nevus, superficial spreading melanomas, and nodular melanomas. Two methods of counting black dots within nuclei were employed. One method was to count the discrete black dots within the nuclei, including the tiny black dots seen within the nucleolus; the second method did not take into account the subsidiary cluster of tiny black dots seen within the nucleolus, instead treating these dots as a single structure. Whichever method we used, a significant difference was found between the pooled mean AgNOR numbers for benign and malignant lesions. We found an overlap, however, between benign, in particular Spitz and dysplastic nevi, and malignant lesions when considering individual counts of AgNOR using both methods. We conclude that studying AgNOR does not seem to be a useful technique to differentiate Spitz and dysplastic nevi from malignant melanomas.     

Upregulation of S100P,receptor for advanced glycation end products and ezrin in malignant melanoma

S100P is a member of the S100 family. Increased levels of S100P have been documented in various malignancies. Binding of extracellular S100P to receptor for advanced glycation end products (RAGE) or coupling of intracellular S100P with a cytoskeletal protein, ezrin, play a crucial role in tumor growth, invasion and metastasis. However, little is known about the expression of S100P, RAGE and ezrin in malignant melanoma. We immunostained these three molecules in 20 primary and 20 metastatic melanomas. Samples of 20 benign nevus pigmentosus and 10 of normal skin were tested as controls. The expression levels (percentage of positively stained cells) of S100P, RAGE and ezrin were significantly higher in melanomas than in nevus pigmentosus. Moreover, slightly but significantly higher expression levels were observed in metastatic than in primary melanomas. Significant positive correlations were evident between the expression levels of S100P and RAGE, S100P and ezrin, and RAGE and ezrin, respectively. In conclusion, the coordinate upregulation of S100P, RAGE and ezrin may possibly facilitate malignant transformation of melanoma.     

Evaluation of tumour-infiltrating CD4+CD25+FOXP3+ regulatory T cells in human cutaneous benign and atypical naevi, melanomas and melanoma metastases

BACKGROUND: CD4+CD25+FOXP3+ regulatory T cells (Tregs) are thought to induce immunotolerance in melanoma. They have not yet been investigated in the entire spectrum of melanocytic cutaneous lesions within a tumour site. OBJECTIVES: To evaluate CD4+CD25+FOXP3+ Tregs among tumour-infiltrating lymphocytes in cutaneous melanocytic lesions. METHODS: We analysed 128 lesions (10 benign junctional common naevi, 10 benign compound common naevi, 10 compound Spitz naevi, 10 junctional atypical naevi, 20 compound atypical naevi, 20 radial growth phase melanomas, 30 vertical growth phase melanomas and 18 melanoma metastases). Tregs were identified by CD25-FOXP3 double immunostains. RESULTS: This study indicates that CD4+/CD25+FOXP3+ Tregs are present in all groups of lesions. Junctional atypical naevi, compound atypical naevi and radial growth phase melanomas showed the highest percentages of CD4+CD25+FOXP3+ Tregs (junctional atypical naevi vs. junctional common naevi, compound common naevi, compound Spitz naevi, melanoma metastases: P < 0.0001; junctional atypical naevi vs. vertical growth phase melanomas: P = 0.001; compound atypical naevi vs. junctional common naevi, compound common naevi: P < 0.0001; compound atypical naevi vs. compound Spitz naevi, melanoma metastases: P = 0.002; compound atypical naevi vs. vertical growth phase melanomas: P = 0.02; radial growth phase melanomas vs. junctional common naevi, compound common naevi, compound Spitz naevi, melanoma metastases: P < 0.0001; radial growth phase melanomas vs. vertical growth phase melanomas: P = 0.008). CONCLUSIONS: The strong prevalence of CD25+FOXP3+ Tregs both in junctional and compound atypical naevi and radial growth phase melanomas, suggests that they induce immunotolerance early during melanoma genesis, favouring melanoma growth. Their evaluation within a tumour site could be useful for prognostic and therapeutic purposes.     

Antigenic phenotype of pigment cell lesions: a study of melanoma, its precursor lesions and heterogeneity in metastatic melanoma

Expression of surface antigens on cells comprising malignant, dysplastic, and benign pigment cell lesions has been evaluated in situ using an avidin-biotin immunohistologic technique. We have demonstrated expression of HLA-DR, beta 2-microglobulin, and 2 melanoma-associated antigens (MAA) in a range of benign and malignant lesion, and have confirmed the close association of HLA-DR and MAA on malignant lesions. In addition, 5 subcutaneous lesions removed at the same time from one individual were evaluated. Clinical and histologic appearance differed among lesions although the antigenic patterns were similar.     

Comparison of macromelanosomes and autophagic giant melanosome complexes in nevocellular nevi, lentigo simplex and malignant melanoma

This study compared the fine structure of macromelanosomes with that of giant melanosome complexes formed through melanosomal autophagocytosis in nevocytes and melanocytes of nevocellular nevi, lentigo simplex and malignant melanoma. While macromelanosomes were found only on rare occasions in these pigmentary disorders [2 of 79 nevocellular nevi (2 junctional nevi), 3 of 12 lentigo simplex and 2 of 93 malignant melanoma], the giant autophagic melanosome complexes were always present, indicating that macromelanosomes are not synthesized simply through melanosomal autophagocytosis. Although both macromelanosomes and giant melanosome complexes exhibited acid phosphatase activity similar to melanosomes, they showed many different ultrastructural features. Characteristically, macromelanosomes contained numerous vesiculoglobular bodies, whereas these bodies were absent in giant melanosome complexes. In those tissues where the presence of macromelanosomes had been ruled out by light microscopy, none of the giant melanosome complexes revealed ultrastructural features indicative of macromelanosomes. Various phases of melanosomal degradation were seen, indicating that they were not simply end-products of lysosomal degradation of melanosomes. It was thought that the key process in the development of macromelanosomes was the accumulation of vesiculoglobular bodies.