Abstract. Cutaneous melanoma is an aggressive cancer and its onset and growth are associated, through direct and indirect interactions, with the cancer microenvironment. The microenvironment comprises a dynamic complex of numerous types of cells (due to histogenesis) that constantly interact with each other through multiple cytokines and signaling proteins. Macrophages are one of the most thoroughly studied pleiotropic cells of the immune system. One of their major cytophysiological functions is their involvement in phagocytosis. Previous studies examining the microenvironment of melanomas and tumor-associated macrophages have revealed that they are involved in all stages of melanomagenesis. In the case of cancer initiation, they form an inflammatory microenvironment and then suppress the anticancer activity of the immune system, stimulate angiogenesis, enhance migration and invasion of the cancer cells, and ultimately contribute to the metastatic process. The present review provides a detailed overview on the function of macrophages in the melanoma microenvironment. IntroductionMelanomas are a rare but aggressive cutaneous type of cancer in humans (1). At the dissemination stage in a majority of cases, the disease is resistant to treatment with cytostatics and radiotherapy (1). Therefore, the identification of novel molecular mechanisms involved in the melanomagenesis process and tumor progression have enabled the production of targeted therapies that yield notable effects (1). The basis for melanomagenesis is the accumulation of genetic disorders in the melanocyte (the most frequent ones include the following mutations: B-Raf proto-oncogene, serine/threonine kinase, N-Ras proto-oncogene, GTPase and phosphatase and tensin homolog) (1). However, only the interaction between microenvironment elements and genetic changes in the melanocyte result in the ultimate transformation of a dysplastic melanocyte into a melanoma cell, and at further stages result in the local invasion and dissemination of the primary lesion (1). It is the microenvironment that is one of the key elements of cancer formation and is being studied at present.A melanoma microenvironment is a markedly heterogenic population of cells that involves fibroblasts, macrophages, lymphocytes, other immune system cells, adipocytes and cells that form the structural elements of cutaneous blood vessels sunk in the extracellular matrix (2). The aforementioned complex network of cellular associations are constantly interacting through direct contact and active protein substances including secretory proteins (e.g., metalloproteinases or

The presence of ulceration has been considered as one of the most important primary tumor characteristics of cutaneous malignant melanoma (CMM) for predicting patient outcome. Yet recently, scientific attention has been drawn towards another microscopic feature of primary tumors, the mitotic rate (MR). The present study aimed to examine the relationship between the presence of ulceration and the mitotic rate and clinicopathological characteristics and melanoma patient survival, and to discuss the results in the context of AJCC melanoma staging recommendations. Tissue samples were obtained from 104 patients treated for CMM. In classical H&E staining, the mitotic rate and the presence of ulceration were evaluated. Non-mitogenic tumors were defined as having 0 mitoses/mm2, low mitogenic potential, 1-2 mitoses/mm2 and highly mitogenic tumors, ≥3 mitoses/mm2. In the entire group of 104 patients, a high mitotic rate (hMR) and ulceration were highly negative prognostic factors, and indicated considerably shorter overall survival, cancer-specific overall survival and disease-free survival. Notably, hMR appeared to have a statistically significant negative impact on survival in early melanomas in both the pT1 (P=0.001) and pT2 subgroups (P=0.006). Kaplan‑Meier analysis of the remaining subsets (pT3 and pT4) did not reveal any important differences in the 5-year survival with regard to MR values. The presence of ulceration also had a prognostic significance for early melanomas, but only for pT1 tumors (P=0.05). Multivariate analysis confirmed that hMR was strongly associated with an unfavorable prognosis. Ulceration had no prognostic significance in the Cox proportional hazards model. Considering the biology of melanoma, hMR seems to be a more reliable parameter than the presence of ulceration. The value of MR categorizes melanomas into tumors with low or high proliferative potential, thus giving direct information concerning their capacity to infiltrate deeper layers of the dermis and, potentially, to generate regional lymph node and distant metastases.

Epithelial-mesenchymal transition (EMT) is a biological process that drives polarized, immotile epithelial cells to undergo multiple biochemical changes to acquire a mesenchymal cell phenotype. The characteristic features of EMT are cell apolarity, loss of cellular adhesion, reduced expression of E-cadherin and increased migratory capacity, as well as invasiveness. EMT is a physiological process that is essential for normal embryonic development. Additionally, abnormal activation of EMT contributes to some human pathologies such as tissue fibrosis, cancer cell invasion and metastasis. In both situations, the basic molecular mechanisms are similar, but lead to different effects depending on cell type and biological conditions of the environment. TGF-β is a multifunctional cytokine that controls proliferation, differentiation and other functions in many cell types. It has been found that neoplastic development converts TGF-β into an oncogenic cytokine. It activates various molecular processes, which are engaged in EMT initiation. All that makes TGF-β a key regulator of EMT.

Objectives Merkel cell carcinoma is a rare but very aggressive cutaneous tumor. We evaluated the prognostic potential of B-cell markers (terminal deoxynucleotidyl transferase [TdT], PAX5, CD117), follicular stem cell markers (CK15, CK19), p63, p53, RB, and Merkel cell polyomavirus (MCPyV; CM2B4) in 136 primary cutaneous Merkel cell carcinomas. Methods Clinical, histopathologic, and immunohistochemical analyses were performed. The results were correlated with patient outcomes by Fisher exact test, log-rank tests, and Cox multivariate models. Results By Fisher exact test, although TdT significantly correlated with both lack of progression (P = .0087) and alive status (P = .0056), MCPyV status correlated only with alive status (P = .031). In univariate analyses, TdT, MCPyV, and RB significantly correlated with improved overall survival, whereas p63 and CK15 correlated with worse overall survival. However, in multivariate analyses, only TdT expression remained as an independent predictor of improved overall survival, Merkel cell carcinoma-specific survival, and progression-free survival. By linear regression analyses, significant correlations between MCPyV vs TdT, PAX5, and CD117 were observed. Conclusions TdT expression is a potential marker of better survival in Merkel cell carcinoma. Expression of B-cell markers is associated with MCPyV, suggesting that clonal viral integration might play a role in the expression of these markers.

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(1) Background: Poly(ADP-ribose) polymerase 1) (PARP1) is a pleiotropic enzyme involved in several cellular processes, e.g., DNA damage repair, regulation of mitosis, and immune response. Little is known about the role of PARP1 in melanoma development and progression. We aimed to investigate the prognostic significance of PARP1 expression in cutaneous melanoma through evaluation of mRNA and protein levels of PARP1 in normal melanocytes and melanoma cell lines, as well as in patients’ tissue material from surgical resections. (2) Methods: An in vitro model was based on two types of normal human melanocytes (HEMn-DP and HEMn-LP) and four melanoma cell lines (A375, WM1341D, Hs294T, and WM9). PARP1 mRNA gene expression was estimated using real-time polymerase chain reaction (RT-PCR), whereas the protein level of PARP1 was evaluated by fluorescence confocal microscopy and then confirmed by Western Blotting analysis. The expression of PARP1 was also assessed by immunohistochemistry in formalin-fixed paraffin-embedded tissues of 128 primary cutaneous melanoma patients and correlated with follow-up and clinicopathologic features. (3) Results: The in vitro study showed that melanoma cells exhibited significantly higher PARP1 expression at mRNA and protein levels than normal melanocytes. High PARP1 expression was also associated with the invasiveness of tumor cells. Elevated nuclear PARP1 expression in patients without nodal metastases strongly correlated with significantly shorter disease-free survival (p = 0.0015) and revealed a trend with shorter cancer-specific overall survival (p = 0.05). High PARP1 immunoreactivity in the lymph node-negative group of patients was significantly associated with higher Breslow tumor thickness, presence of ulceration, and a higher mitotic index (p = 0.0016, p = 0.023, and p < 0.001, respectively). In patients with nodal metastases, high PARP1 expression significantly correlated with the presence of microsatellitosis (p = 0.034), but we did not confirm the prognostic significance of PARP1 expression in these patients. In the entire analyzed group of patients (with and without nodal metastases at the time of diagnosis), PARP1 expression was associated with a high mitotic index (p = 0.001) and the presence of ulceration (p = 0.036). Moreover, in patients with elevated PARP1 expression, melanoma was more frequently located in the skin of the head and neck region (p = 0.015). In multivariate analysis, high PARP1 expression was an independent unfavorable prognosticator in lymph node-negative cutaneous melanoma patients. (4) Conclusions: In vitro molecular biology approaches demonstrated enhanced PARP1 expression in cutaneous melanoma. These results were confirmed by the immunohistochemical study with clinical parameter analysis, which showed that a high level of PARP1 correlated with unfavorable clinical outcome. These observations raise the potential role of PARP1 inhibitor-based therapy in cutaneous melanoma.

Background: Merkel cell carcinomas of unknown primary (MCC-UPs) are defined as deep-seated tumors without an associated cutaneous tumor. Although the distinction has important clinical implications, it remains unclear whether these tumors represent primary tumors of lymph nodes or metastatic cutaneous primaries. Methods: We compared the immunohistochemical profiles of four groups of MCCs (Merkel cell polyomavirus (MCPyV)-positive UP, MCPyV-negative UP, MCPyV-positive known primary (KP), and MCPyV-negative KP) using B-cell and pre-B-cell markers, cell cycle regulating proteins, follicular stem cell markers, and immune markers, and performed next generation and Sanger sequencing. Results: Virus-positive and virus-negative MCC-UPs exhibited an immunoprofile similar to virus-positive and virus-negative primary cutaneous MCCs, respectively. MCC-UP tumors (both virus-positive and -negative) were immunogenic with similar or even higher tumoral PD-L1 expression and intratumoral CD8 and FoxP3 infiltrates in comparison to MCPyV-positive cutaneous tumors. In addition, similar to primary cutaneous MCCs, MCPyV-negative MCC-UPs exhibited UV signatures and frequent high tumor mutational burdens, whereas few molecular alterations were noted in MCPyV-positive MCC-UPs. Conclusions: Our results showed distinct UV-signatures in MCPyV-negative tumors and high immunogenicity in MCPyV-positive tumors. Although additional studies are warranted for the MCPyV-positive cases, our findings are supportive of a cutaneous metastatic origin for MCPyV-negative MCC-UP tumors.

The most important result of our study was that we confirmed the strict correlation between SPARC and NCAD expression and clinicopathological parameters related with melanoma progression, which is a specific clinical equivalent of the molecular mechanisms of epithelial-mesenchymal transition process and confirms its key role in the disease outcome.