Malignant Melanocytic Tumors Research Articles

P16ink4a Expression in Benign and Malignant Melanocytic Conjunctival Lesions

Acquired conjunctival melanocytic lesions include nevi, primary acquired melanoses (PAMs), and melanomas. Conjunctival melanoma is a malignant melanocytic neoplasm with a high metastasis and mortality rate. Usually, the diagnosis can be achieved only with routine microscopic analysis, but in some cases, the samples are small or have artifacts. In these cases, complementary studies will be helpful, but currently, there are no well-understood or studied complementary methods. Objective. To analyze the immunohistochemical expression of p16 in conjunctival melanocytic lesions and to assess its potential for differentiating between benign and malignant melanocytic lesions. Methods. Immunohistochemical study against p16ink4a (p16) was performed on paraffin-embedded sections on 45 melanocytic lesions (9 melanomas, 19 nevi, and 2 PAMs with atypia and 15 without atypia). Expression was scored according to the German immunoreactive score (IRS). Results. Expression of p16 IRS differed between nevi, PAMs, and melanomas. The mean IRS for melanomas was 3.3 ± 1.8 and was lower than those for nevi (7.63 ± 3.24; P < .05), PAM with atypia (12 ± 0; P < .05), and PAM without atypia (11 ± 1.69; P < .05). Lesions with infiltration depths lower than 2 mm showed higher levels of p16. There were no differences between favorable and unfavorable locations. Conclusion. p16 Expression in conjunctival melanocytic lesions showed an expression similar to that in skin and seems to be a good marker to differentiate nevi and PAMs from melanomas. However, additional studies of larger series and follow-up are needed to confirm these findings.

A “High-Risk” Epithelioid Hemangioendothelioma Presenting as a Solitary, Ulcerated, Subcutaneous Tumor

So-called "high-risk" epithelioid hemangioendotheliomas are uncommon neoplasms that demonstrate classic histopathologic features of epithelioid hemangioendotheliomas and a size larger than 3 cm or >3 mitotic figures per 50 high power fields. These neoplasms show an increased rate of metastasis (25% of cases) and are associated with a poor 5-year survival (59%). They may pose a diagnostic challenge for dermatopathologists as they mimic metastatic carcinoma, malignant mixed tumor, melanocytic neoplasms, epithelioid sarcoma, and epithelioid angiosarcoma. High-risk epithelioid hemangioendothelioma has not been previously reported as a solitary ulcerated mass in the skin. Here, we describe one such lesion that developed as a rapidly growing ulcerative skin tumor in a 33-year-old African American man with a remote history of Burkitt lymphoma. We also review the evolution and controversies in the understanding and classification of this neoplasm.

C-kit Expression in Canine Mucosal Melanomas

The c-kit receptor is responsible for transmission of promigration signals to melanocytes; its downregulation may be involved in malignant progression of human melanocytic neoplasms. Expression of this receptor has not been examined in normal or neoplastic melanocytes from dogs. In this study, 14 benign dermal and 61 malignant mucosal melanocytic tumors were examined for c-kit (KIT) expression. Sites of the mucosal melanomas were gingiva (not further specified; n = 30), buccal gingiva (n = 6), soft palate (n = 4), hard palate (n = 5), tongue (n = 7), lip (n = 6), and conjunctiva (n = 3). Melan A was expressed in all 14 dermal melanocytomas and in 59 of 61 (96.7%) tumors from oral or conjunctival mucosa, confirming melanocytic origin. C-kit receptor expression was strong and diffuse throughout the cytoplasm in all 14 dermal melanocytomas and was identified in basilar mucosal melanocytes over submucosal neoplasms (27 of 61, 44.3%), junctional (neoplastic) melanocytes (17 of 61, 27.9%), and, less commonly, neoplastic melanocytes of the subepithelial tumors (6 of 61, 9.8%). KIT expression anywhere within the resected melanomas correlated with significantly longer survival. These results suggest that c-kit receptor expression may be altered in canine melanomas and may have potential as a prognostic indicator for mucosal melanomas.

Glucocorticoid Receptor Expression and Antiproliferative Effect of Dexamethasone on Human Melanoma Cells

Glucocorticoids, such as dexamethasone are widely used in cancer therapy and have cell type-specific pro- or antiapoptotic effects. We examined whether melanoma cells are sensitive to dexamethasone treatment. We have demonstrated for the first time that in human melanoma cell lines as well as in benign and malignant melanocytic tumors glucocorticoid receptor (GCR) is present both at mRNA and protein level. Dexamethasone applied at high doses inhibited the in vitro growth of WM983A human melanoma cells. The inhibitory effect was due to apoptosis induction. In the case of this relatively sensitive cell line dexamethasone enhanced the effect of the chemotherapeutic drug DTIC.

Primary Oral Malignant Melanoma

Malignant melanoma (MM) is a malignant neoplasm of epidermal melanocytes. Primary malignant melanoma (PMM) is rarely found in the oral cavity. It is estimated at between 0.2 and 8% of all melanomas and approximately four times more frequently in the oral mucosa of the upper jaw, usually on the palate and anterior gingiva. It is more prevalent in the older population having 50 to 80 years of age. Males are more commonly affected than females. Some rare cases of involvement of lip, mandibular gingiva and buccal mucosa were also reported previously. Here, we are reporting a rare case of MM in the oral cavity of 60-year-old male patient involving mandibular gingiva.

Optical Transfer Diagnosis of Pigmented Lesions

Optical transfer diagnosis is a novel melanoma detection system that uses morphologic-physiologic mapping. To further evaluate the potential of optical transfer diagnosis for distinguishing benign from malignant pigmented melanocytic neoplasms. Ninety-four patients with pigmented lesions suggestive of melanoma were referred for optical transfer diagnosis. After lesions were scanned with the camera, they were removed for histopathologic examination by two dermatopathologists each. From the recorded images, morphologic-physiologic maps were created with prediction models of light absorption and scattering by chromophores such as hemoglobin, keratin, and melanin at different epidermal and dermal depths. Entropy and relative entropy values derived from the morphologic-physiologic maps and a set of pure morphologic parameters were analyzed for output prediction of melanoma versus nonmelanoma. Dermoscopic images were reviewed and scored using the color, architecture, symmetry, and homogeneity (CASH) algorithm to assign a value of clinical atypia. RESULTS Of the 118 scanned and biopsied lesions (median CASH score 8), 11 were identified as melanoma or atypical melanocytic hyperplasia consistent with melanoma. For identification of melanomas, optical transfer diagnosis had a sensitivity of 100% and a specificity of 90%. This technology continues to be a promising adjunct to clinical skin cancer screening.

The potential role of vitamin D in the progression of benign and malignant melanocytic neoplasms.

Hormonally active vitamin D3, 1,25-(OH)2D3, is believed to have a role in the prevention of cancer formation and in limiting the aggressiveness of cancers that do arise. Therefore,much interest is presently being focused on 1,25-(OH)2D3 and its analogues as potential treatments for various cancers including melanoma. This article discusses the evidence in favour of a role for 1,25-(OH)2D3 in protection against the progression of melanocytic lesions and also summarizes the mechanisms by which 1,25-(OH)2D3 may act to protect against melanoma development and progression.

FISH analysis for diagnostic evaluation of challenging melanocytic lesions.

The differential diagnosis of malignant melanomas and atypical melanocytic nevi is still a diagnostic challenge. The currently accepted morphologic criteria show substantial interobserver variability, likewise immunohistochemical studies are often not able to discriminate these lesions reliably. Techniques that support diagnostic accuracy are of the greatest importance considering the growing incidence of malignant melanomas and their increase in younger patients. In this study we analyzed the feasibility of fluorescence in situ hybridization (FISH) analysis for the discrimination of malignant and benign melanocytic tumors. A panel of DNA probes was used to detect chromosomal aberrations of chromosomes 6 and 11. On a series of 5 clearly malignant and benign melanocytic tumors we confirmed the applicability of the test. Then we focused on examination of ambiguous melanocytic lesions, where atypical cells are often difficult to relocalize in the 4',6-Diamidino-2-phenylindol (DAPI)-fluorescence stain. FISH analyses were conducted on destained H&E-stained slides. By comparison of the DAPI-image with photos taken from the H&E stain, unambiguous assignment of the FISH results to the conspicuous groups of cells was possible. The results of FISH analysis were consistent with the conventional diagnosis in 11 of 14 small ambiguous lesions. Of the remaining 3 cases, 2 showed FISH-results close to the cut-off level. Comparison of FISH results on thin and thick sections revealed that the cut-off values have to be adapted for 2 microm destained sections. In conclusion, FISH analysis is a useful and applicable tool for assessment of even smallest melanocytic neoplasms, although there will remain unclear cases that cannot be solved even after additional FISH evaluation.

In vivo reflectance confocal microscopy: automated diagnostic image analysis of melanocytic skin tumours

In vivo reflectance confocal microscopy (RCM) has been shown to be a valuable imaging tool in the diagnosis of melanocytic skin tumours. However, diagnostic image analysis performed by automated systems is to date quite rare. In this study, we investigated the applicability of an automated image analysis system using a machine learning algorithm on diagnostic discrimination of benign and malignant melanocytic skin tumours in RCM. Overall, 16,269 RCM tumour images were evaluated. Image analysis was based on features of the wavelet transform. A learning set of 6147 images was used to establish a classification tree algorithm and an independent test set of 10, 122 images was applied to validate the tree model (grouping method 1). Additionally, randomly generated 'new' learning and test sets, tumour images only and different skin layers were evaluated (grouping method 2, 3 and 4). The classification tree analysis correctly classified 93.60% of the melanoma and 90.40% of the nevi images of the learning set. When the classification tree was applied to the independent test set 46.71 ± 19.97% (range 7.81-83.87%) of the tumour images in benign melanocytic skin lesions were classified as 'malignant', in contrast to 55.68 ± 14.58% (range 30.65-83.59%; t-test: P < 0.036) in malignant melanocytic skin lesions (grouping method 1). Further investigations could not improve the results significantly (grouping method 2, 3 and 4). The automated RCM image analysis procedure holds promise for further investigations. However, to date our system cannot be applied to routine skin tumour screening.

Immunohistochemical analysis of the S100A1, S100B, CD44 and Bcl-2 antigens and the rate of cell proliferation assessed by Ki-67 antibody in benign and malignant melanocytic tumours

Malignant melanomas usually have an unfavourable prognosis and poor response to chemotherapy. Deregulation of cell proliferation, programmed cell death and intercellular interactions are among several important mechanisms that might lead to malignant transformation of melanocytes and melanoma progression. The S100A1, S100B, Bcl-2 and CD44 antigens have all been described as being involved in different processes of melanoma progression. The expression of these antigens, as well as the rate of cell proliferation, was analyzed retrospectively in melanocytic tumours from 126 patients (32 males and 94 females, age ranging from 11 to 91 years). The series included benign (45 intradermal, 27 compound and eight displastic naevi) and malignant (39 primary and 14 metastatic) melanocytic tumours. The proliferating rate assessed by Ki-67 staining was lower in naevi than in melanomas, with a correlation coefficient of r = 0814. There was no overlap for rate of proliferation between benign and malignant tumours. The expression of S100A1 was low in benign melanocytic tumours and increased in malignant melanomas (r = 0.61). In contrast, a higher percentage of S100B antigen-positive cells were observed in benign melanocytic lesions than in melanomas (Pearson correlation coefficient, 0.627). In addition, positive immunostaining for S100B antigen in malignant melanomas corresponded with the areas with increased proliferating rate. The expression of Bcl-2 was lower in melanomas than in benign melanocytic tumours (r = -0.53). Bcl-2-negative areas within melanomas had an increased proliferating rate. The expression of CD44 showed a large variation both in benign and malignant melanocytic tumours. CD44 antigen expression was higher in melanomas with known metastases than in those without metastases, but this difference was not statistically significant.

Immunohistochemical Studies of Conjunctival Nevi and Melanomas

To evaluate the role of immunohistochemical methods in the diagnosis of benign and malignant conjunctival melanocytic proliferations. Retrospective immunohistopathologic study. Paraffin-embedded tissue sections from 20 conjunctival nevi and 15 invasive melanomas were immunoreacted with antibodies against cellular antigens S-100 protein, MART-1, HMB-45, CD-45, and Ki-67 nuclear proliferation protein. All nevi immunostained moderately to strongly for S-100 protein and MART-1. Results for HMB-45 were negative in the middle and lower subepithelial portions of 18 of 20 lesions; it was usually only weakly positive within the superficial junctional zone. Only 1 melanoma did not stain positively for S-100; MART-1 and HMB-45 were positive in all lesions at some level of intensity. Ki-67 positivity was restricted to the junctional zone of nevi and was diffuse in melanomas. The mean Ki-67 proliferation indices were 1.89% for the nevi and 17.3% for the melanomas. CD-45 can help to highlight lymphocytes that immunostain with Ki-67. Melanomas in situ and atypical primary acquired melanoses had more than twice the Ki-67 proliferation counts of intraepithelial junctional nevocytes (P < .001) and more intense HMB-45 cytoplasmic staining than junctional zone nevocytes. S-100 and MART-1 were not useful in separating benign from malignant lesions. Results for nevus cells beneath the junctional zone were overwhelmingly negative for HMB-45 and Ki-67. Two nevi and all melanomatous nodules were positive for HMB-45 (P < .001). A higher Ki-67 proliferation index convincingly separated melanomas from nevi (P < .001). Immunostaining for HMB-45 and Ki-67 are valuable adjuncts to careful histopathologic evaluation in assessing benign and malignant conjunctival melanocytic tumors.

Index of Suspicion

Index of Suspicion

Fluorescence in Situ Hybridization for Distinguishing Nevoid Melanomas From Mitotically Active Nevi

Nevoid melanoma may resemble benign compound or intradermal nevi by their silhouette and profile on low power. Higher power usually reveals nuclear atypia, confluence of cells, incomplete maturation and dermal mitotic activity. However, to some extent all of these features maybe seen in benign compound or intradermal nevi and no single criteria can be used to differentiate nevoid melanoma from a benign nevus. The distinction can be particularly problematic in nevi that show mitotic activity and we have noted a recent trend in diagnosis of melanocytic neoplasms with dermal mitosis as nevoid melanoma despite the presence of normal maturation in the dermis and lack of significant nuclear atypia. Therefore in this study we evaluated 10 nevoid melanomas, 4 of which resulted in metastasis and 10 mitotically active nevi with fluorescence in situ hybridization targeting key chromosomal loci previously shown to effectively discriminate been malignant and benign melanocytic neoplasms. All 10 nevoid melanomas showed copy number abnormalities by fluorescence in situ hybridization in either chromosome 6 or 11 while none of the 10 mitotically active nevi did. The results demonstrate that fluorescence in situ hybridization targeting key chromosomal loci on chromosomes 6 and 11 can be effective in discriminating nevoid melanomas from mitotically active nevi. Additionally, our study presents further evidence that dermal mitoses alone without other diagnostic features such as nuclear atypia and lack of maturation does not constitute sufficient evidence alone for a diagnosis of melanoma.

BRAF mutations in melanocytic tumors (nevi and melanomas) from organ transplant recipients

BRAF is a gene of the RAF family of kinases, frequently mutated in benign and malignant melanocytic tumors (nevi and melanomas). Organ transplant recipients are at high risk for developing various tumors, including melanocytic ones. We studied a group of 129 melanocytic tumors including various subtypes of nevi (n: 114) and melanomas (n: 15) excised from transplant (n: 63) and control (non-immunosuppressed) patients (n: 66) as to BRAF mutation status. Mutation research was performed after extraction of DNA from archival material (paraffin-embedded tissue specimens) by sequence analysis. BRAFV600E accounted for the most prevalent mutation found (94%). Melanocytic tumors from transplant patients had a lower frequency of BRAF mutations than control lesions (45.4% vs 63.5%, p<.05). The explanation for this difference is currently unknown. The possibility exists that in transplant patients, factors linked to immunosuppression (most likely immunosuppressive drugs) induce additional mutations, or activate alternative signaling pathways, which compensates for the lower rate of activating BRAF mutations in tumors developing in these patients.

Fluorescence In Situ Hybridization in the Diagnosis of Soft Tissue Neoplasms: A Review

This paper presents an overview of the role of fluorescence in situ hybridization (FISH) in the diagnosis of soft tissue neoplasms. Many soft tissue neoplasms harbor characteristic translocations or amplification of gene regions, which can be assessed by FISH, and can be used to assist in their diagnosis. We discuss the major morphologic categories in which FISH has come to be used including high-grade round cell sarcomas, spindle cell sarcomas, low-grade myxoid neoplasms, adipocytic neoplasms, and malignant melanocytic neoplasms on the basis of a recent review of soft tissue neoplasms which were analyzed by FISH. We also review the molecular alterations (translocations and amplification of gene regions), which have come to define many of these diagnostic entities and the most effective way to evaluate them with FISH with attention to potential pitfalls. Finally, we discuss the advantages and disadvantages of FISH as a technique when appraising soft tissue neoplasms.

Expression of cyclooxygenase-2 in canine uveal melanocytic neoplasms

To determine whether cyclooxygenase-2 (COX-2) is expressed in benign or malignant canine uveal melanocytic neoplasms and whether expression correlates with malignancy. Tissue sections from 71 globes; 57 with benign (n = 15), malignant (34), or mixed (8) uveal melanocytic neoplasms; 10 with nonneoplastic disease; and 4 with no abnormalities. Bleached sections from all globes and canine kidney were incubated with mouse monoclonal antibody directed against rat COX-2 protein or mouse antibody isotype control. Location, intensity, and percentage of immunolabeled cells were scored. Expression of COX-2 was detected in all but 5 globes, all of which contained neoplasms. Expression of COX-2 was detected in regions infiltrated by neoplasia in 21 globes; however, definitive labeling of tumor cells was detected in only 2 of those. In the remaining 19 globes, COX-2 expression was detected in areas also labeled in globes without disease and globes with nonneoplastic disease, especially the aqueous outflow tract and ciliary body. However, only globes with uveal malignant melanomas had detectable COX-2 expression in the iris. Expression of COX-2 was detected in the ciliary body of more globes with uveal malignant melanoma (20/34) than in those without disease (1/4), with nonneoplastic disease (4/10), or with melanocytoma (3/15) or mixed neoplasms (3/8). Canine globes with uveal melanocytic neoplasia appeared to express COX-2 in similar sites and with similar intensity as globes without neoplasia. Differentiation of benign from malignant canine uveal melanocytic neoplasms was not possible.

Intramedullary Primary Cervical Melanoma

We present an extremely rare case report of a primary intramedullary malignant melanoma arising into the cervical spinal cord. We describe and review clinical presentations, characteristic findings from imaging studies, diagnostic criteria, prognosis, and treatment of primary cervical intraspinal melanoma. A 52-year-old man was presented with numbness in 4 limbs and weakness in both upper limbs. The clinical and radiologic findings were suggestive of malignant process. To preserve the patient's neurologic functions the tumor was subtotally resected. Histopathologic examination demonstrated a malignant melanocytic tumor. Treatment of intramedullary malignant melanoma is difficult. The best treatment should be total surgical excision of the lesion, whenever possible, but removal of the tumor generally is incomplete, and radiation therapy is often recommended after the surgery. It is also important to differentiate whether melanoma is primary or metastases. It has been suggested that primary spinal melanoma exhibits slow progression and the tumor is less aggressive than the more common skin melanoma with central nervous system metastases. Diagnostic criteria and different treatment options for primary and metastatic malignant melanoma is discussed.

Reduced p16 and Increased Cyclin D1 and pRb Expression Are Correlated With Progression in Cutaneous Melanocytic Tumors

The immunohistochemical expression of cell cycle proteins p16, cyclin D1, and pRb was assessed in 112 benign and malignant melanocytic tumors and correlated with tumor progression, prognosis, and outcome. Comparing benign and malignant tumors, there were significant differences in the median score for all 3 proteins, with decreased p16 (P = .000001), increased cyclin D1 (P = .01), and increased pRb in melanomas (P = .01). There was a progressive loss of expression of p16 with progression from benign naevi to primary melanomas and to metastases. p16 was significantly decreased in primary tumors from melanoma patients who developed recurrent disease (P = .0000013). Cyclin D1 and pRb showed a progressive increase in expression from benign to malignant tumors but with relative decreases in the more advanced tumors (thick primaries and metastatic melanomas). Alterations in cell cycle proteins involved in G1/S transition are implicated in melanocytic tumor progression and have a potential role in diagnosis and prognostication.

Diagnosis of cutaneous melanocytic tumours by four-colour fluorescence in situ hybridisation

Accurate classification of primary melanocytic tumours as benign or malignant is crucial for prognostic prediction and appropriate patient management. Several chromosomal aberrations have been frequently identified in melanomas, but are absent in melanocytic naevi. We performed four-colour fluorescence in situ hybridisation (FISH) analysis of melanocytic tumours to determine the accuracy of the technique in classifying melanocytic tumours as benign or malignant. FISH was performed on paraffin-embedded tissue from 40 histologically unequivocal melanocytic tumours (10 metastatic melanomas, 10 primary melanomas and 20 benign melanocytic naevi) using the product Vysis LSI RREB1/LSI MYB/LSI CCND1/CEP 6 probes (Abbott Molecular Laboratories, USA), which is designed to detect the copy number of the RREB1 (6p25), MYB (6q23), and CCND1 (11q13) genes and FISH positivity is defined by means of a scoring algorithm. FISH distinguished the melanomas and the naevi with a sensitivity of 90% (10/10 primary melanoma cases and 8/10 metastatic melanoma cases, respectively), and a specificity of 95%. The most common abnormalities in the melanomas were increased copies of 11q (70%) and 6p (70%), followed by 6q loss relative to cep6 (50%). Fifteen of the 18 positive melanomas were positive by more than one criterion. The results of this study show that FISH, using a panel of four probes, is a sensitive and specific method of classifying benign and malignant melanocytic tumours. The four-colour FISH technique has the potential to assist in the stratification of the subgroup of melanocytic tumours which are difficult to classify using conventional histology.

Dissecting the roles of Raf- and PI3K-signalling pathways in melanoma formation and progression in a zebrafish model

Deregulated Ras signalling is implicated in most human neoplasia, exemplified by melanoma. Whereas Raf activation occurs almost ubiquitously in benign and malignant melanocytic neoplasms, implying an involvement in tumour initiation, phosphoinositide 3-kinase (PI3K) activation occurs predominantly in malignant neoplasms, implying an involvement in malignant progression. Here, we dissect the contributions of these two pathways to tumourigenesis in vivo, by modulating their activities in zebrafish melanocytes. Misexpression of oncogenic Ras (V12RAS) in founder fish induced frequent melanoma, beginning at larval stages, with concomitant activation of Raf-Mek-Erk and PI3K-Akt signalling. Misexpression of effector-domain mutants of V12RAS, or of various downstream effectors, confirmed a selective role for the Raf-Mek-Erk pathway in initiating neoplasia, but highlighted the requirement for additional Ras effector pathways for malignancy. The phenotype of animals with germ-line transmission of V12RAS resembled familial atypical mole and melanoma (FAMM) syndrome: melanocytes displayed hyperplasia, dysplasia, altered terminal differentiation and spontaneously progressed to invasive melanoma. Co-expressing a dominant-interfering form of PI3K abolished V12RAS-induced malignancy, demonstrating a direct role for PI3K signalling in the malignant progression of melanoma in vivo, and highlighting PI3K as a promising target for melanoma therapy.