Beáta Somlai

Vascularization of cutaneous melanoma involves vessel co-option and has clinical significance

This study was undertaken to determine the role and the fate of the peritumoural vascular plexus during the vascularization of human malignant melanoma (hMM) and in an appropriate murine melanoma model system. The prognostic significance of the vascularity of different tumour areas was also evaluated. Despite morphometry revealing several‐fold higher microvessel densities (MVDs) in the peritumoural tissue than at the centre of the tumour, the development of visceral metastases of hMM was exclusively correlated with the MVD of the tumour centre. Furthermore, the 5‐year survival of the patient group with low tumour centre MVD (<30/mm2, n=29) was 100%, compared to 1/16 patients alive with high tumour centre MVD (>30/mm2, n=16). Morphometric analysis and three‐dimensional reconstruction of vessel networks of both human and murine melanomas showed clearly that the peritumoural vascular plexus present at the melanoma base is continuously being incorporated into the growing tumour mass. Once vessels become incorporated, sprouting ceases and the proliferating endothelial cells (EC) take part only in vessel dilatation. Moreover, the immunohistochemical and ultrastructural characterization of microvessels demonstrated that the pericyte coverage of endothelial tubes was complete in all of the investigated areas, in both human and murine melanomas. Copyright

T-Cell Activation Marker Expression on Tumor-Infiltrating Lymphocytes As Prognostic Factor in Cutaneous Malignant Melanoma

The central role of T cells in antitumor immunity is well established. However, tumor progression, often seen in the presence of substantial lymphocytic infiltration, suggests that these T cells are not capable of mounting an effective immune response to control tumor growth. Evidence has accumulated that T lymphocytes infiltrating human neoplasms are functionally defective, incompletely activated, or anergic. Therefore, when characterizing the immune competent cells within lymphoid infiltrates of tumors, it is important to assess their activation state. We investigated the expression of two T-cell activation markers, interleukin 2 receptor α (CD25) and OX40 (CD134), by immunohistochemistry in primary cutaneous melanoma samples of 76 patients and analyzed it in relation to tumor stage and tumor progression (>5 years follow-up), as well as to patients’ survival. We found that the degree of infiltration by CD25+ and intratumoral OX40+ lymphocytes showed a tendency to decrease in thicker melanomas. The frequency of samples with high numbers of peritumoral CD25+ and OX40+ cells was significantly lower (P = 0.0009 and P = 0.0087, respectively) in melanomas developing distant visceral metastases, compared with nonmetastatic or lymph node metastatic tumors. For both activation markers studied, high peritumoral densities were associated with longer survival by univariate analysis (P = 0.0028 and P = 0.0255 for CD25 and OX40, respectively), whereas peritumoral OX40+ lymphocyte infiltration had an impact on survival also in multivariate analysis (P = 0.035). The results suggest that the presence of lymphocytes expressing the T-cell activation markers CD25 or OX40 shows correlation with tumor progression as well as with patients’ survival in cutaneous malignant melanoma.

Role for β3 integrins in human melanoma growth and survival

The role of αIIbβ3 integrin in regulating platelet function is well appreciated, whereas its role in tumor progression and metastasis is not. The purpose of our study was to determine a functional relevance to expression of αIIbβ3 integrin in cells derived from human solid tumors. A study of human melanoma biopsies (n = 24) showed that αIIbβ3 expression increased with tumor thickness, which is indicative of metastatic propensity. Expression of αIIbβ3 was 8% (±1.8), 33% (±10.4) and 62% (±5) in melanomas ranging in thickness from 0–1.5 mm, 1.5–4.0 mm and >4 mm, respectively; αvβ3 was equally high all categories. To determine biological function, we stably transfected αIIbβ3 into human melanoma cells that express αvβ3, but not αIIbβ3. Surface expression of αvβ3 remained unaltered between αIIbβ3 (+) and mock transfected counterparts. The αIIbβ3 (+) cells possessed increased ability to adhere, spread and migrate on fibrinogen. They had decreased ability to attach, spread and migrate on vitronectin. Immunocytochemistry showed that expression of αIIbβ3 displaced αvβ3 from focal contact points. When implanted subcutaneously into SCID mice, the αIIbβ3 (+) cells developed ∼4‐fold larger tumors when compared to their mock counterparts and the level of apoptosis was reduced within the tumors. Results suggest that co‐expression of the 2 β3 integrins, αvβ3 and αIIbβ3, in human melanoma cells enhanced cell survival and promoted growth in vivo.

FOXP3+ Cell Density in Primary Tumor Has No Prognostic Impact in Patients with Cutaneous Malignant Melanoma

Regulatory T cells (Tregs) have been implicated as inhibitors of antitumor immune reactions. However, data on the relevance of their prevalence at tumor sites in influencing disease outcome are controversial. The aim of our study was to investigate the role in tumor progression and the prognostic impact of the density of lymphocytes expressing FOXP3, a transcription factor expressed predominantly by CD4+CD25+ Tregs, in primary cutaneous melanoma. We examined the infiltration of FOXP3+ cells by immunohistochemistry in tumor samples from 97 patients and evaluated in relation to patient and tumor parameters. The degree of infiltration by FOXP3+ cells did not show correlation with the thickness of melanomas. Moreover, no associations were found with metastasis formation during the 5-year follow-up period, patient survival, or any other clinicopathologic parameters studied. These results suggest that the presence of FOXP3+ lymphocytes in primary tumors is not of prognostic importance in human cutaneous melanoma.

Association of microvessel density with infiltrating cells in human cutaneous malignant melanoma

Vascularization and host response to malignant tumors may have common molecular regulators, therefore, we analyzed the relationship between microvessel density (MVD) and tumor infiltrating cells in cutaneous malignant melanoma. Density of lymphocyte subpopulations, macrophages, dendritic cells and CD34+ microvessels was determined by immunohistochemistry in primary tumor samples from fifty-two patients with melanoma thicker than 1 mm. Intratumoral MVD did not show significant association with infiltration for any of these cell types. In the case of peritumoral reactive cell densities analyzed in the whole patient population, a positive correlation of MVD was found with CD3+ T cell density. This association was stronger in melanomas >4.0 mm and in visceral metastatic tumors. In these subgroups similar phenomenon was observed for CD8+ cells. We found significant correlation of MVD with CD68+ macrophage density only in the highest thickness category, and weak associations with B-cell and dendritic cell infiltration in visceral metastatic cases. MVD did not vary significantly in tumors categorized according to thickness, localization, ulceration or histological type. However, both intratumoral MVD and macrophage infiltration were significantly higher in male patients compared to females. The correlation of immune cell density with tumor vascularization and gender differences in vascularity and macrophage infiltration of melanoma deserve further attention.

Loss of vascular adhesion protein-1 expression in intratumoral microvessels of human skin melanoma

Intratumoral vessels are different both structurally and phenotypically, and this may have clinical significance. In this study we analysed the expression of an adhesion molecule – vascular adhesion protein-1 (VAP-1) – in melanoma-associated blood vessels in 28 primary skin melanoma cases using immunocytochemistry and immunoelectron microscopy. We have found that VAP-1 protein expression is significantly decreased in intratumoral vessels compared with peritumoral ones; this difference was independent of the tumour thickness. Loss of VAP-1 protein expression occurred in both endothelial and smooth muscle cell components. Unlike in other cancer types, the VAP-1 protein expression of intratumoral vessels did not correlate with the density of cytotoxic T-lymphocytes or dendritic cells. On the other hand, the 5-year survival of melanoma patients with low VAP-1 protein expression in intratumoral blood vessels (≤25%) was lower (26.3%) than in patients whose VAP-1 expression was higher (42.6%, P=0.0632). These results support the idea that the phenotype of intratumoral blood vessels is important in the progression of malignant melanoma.

Genotype analysis in Hungarian patients with multiple primary melanoma.

Multiple primary melanoma patients (MPMps) have better prognosis and are more prone to genetic predisposition than single melanoma patients. We aimed to compare genetic background (CDKN2A, CDK4, MITF, MC1R) of 43 Hungarian MPMps with their clinicopathological data. We observed a higher rate of synchronous first and second melanoma (MM) (49%) and a higher frequency of non‐melanoma tumor co‐occurrence (42%) than reported previously. CDKN2A mutation frequency was 4.7% (E69G, R99P). We identified a new human MC1R variant (D117G) and reported MC1R variant distributions in Hungarian MMs for the first time. The rare R163Q was exceptionally common among Hungarian MPMps, a variant otherwise frequent in Asia, but not in Europe. MC1R ‘R’ carriers showed histopathological signs of a more progressive disease than ‘r’ carriers did; however, tumor‐infiltrating lymphocytes (TILs) in their second melanomas occurred significantly more frequently. Calculating 5‐year overall survival, ‘R’ carriers showed more unfavourable prognosis (87%) than ‘r’ carriers did (95%).

Stage distribution of malignant melanomas in a Hungarian centre

INTRODUCTION Survival of patients with malignant melanoma primarily depends on tumor stage. Hungarian National Cancer Registry does not specify tumors according to TNM stages. AIM The authors aimed to survey the stage distribution of melanomas at the Department of Dermatology, Dermatooncology and Venerology, Semmelweis University. METHOD 1160 patients (558 males and 602 females, aged 60.5±16 and 57±17 years, respectively) diagnosed with cutaneous melanoma between 2004-2009 were included. RESULTS In comparison with international studies, the case distribution was favorable in stages IA and IV, i.e. the proportion of early melanomas was relatively high (IA: 43.8%), while the incidence in stage IV was low (0.4%). In stages IB-IIA the incidence was significantly lower, while in IIC, IIIA, IIIB it was higher as compared to published data from Western-Europe, Australia and the United States. CONCLUSIONS The study underlines the necessity of prevention and awareness campaigns that may result in increase of early diagnosis of melanomas.

Epidemiological survey of patients with melanoma diagnosed at the Department of Dermatology, Dermatooncology and Venerology, Semmelweis University

A melanoma túlélése elsôsorban a TNM-stádium függvénye, azonban a Nemzeti Rákregiszter a daganatokat stádiumok alapján nem részletezi. Vizsgálatunk során felmértük a Semmelweis Egyetem Bôr-, Nemikórtani és Bôronkológiai Klinikán 2004–2009 között diagnosztizált 1160 cutan melanomás beteg (558 férfi és 602 nô, átlagéletkoruk 59±16 év) stádiumait. Nagy esetszámú nemzetközi vizsgálatokkal összehasonlítva az IA és IV-es stádiumban a klinika betegeinek aránya kedvezônek bizonyult, viszonylag magas százalékban fordultak elô a korai melanomák (IA: 43,8%) és alacsony volt a késôi, IV-es stádiumú daganatok aránya (0,4%). A vizsgálat felhívja a figyelmet arra, hogy a korai diagnózis további javítása érdekében a lakosság folyamatos felvilágosítása, primer és szekunder prevenciós programok szükségesek.