Intravasation Research Articles

Abstract 48: Activation of c-Met receptor signaling by carcinoma-associated fibroblasts in the tumor microenvironment

Abstract Objective: Carcinoma-associated fibroblasts (CAFs) play a key role in tumorigenesis by promoting tumor growth, invasion, and metastasis. At sites of intra-abdominal ovarian cancer metastases, more than half of the tumor microenvironment is composed of CAFs, however, the mechanism for how CAFs promote metastatic disease in ovarian cancer is not known. Because of the prominent role that the receptor tyrosine kinase c-Met plays in metastasis, we hypothesized that CAFs within the metastatic tumor microenvironment may activate c-Met signaling in ovarian cancer cells leading to tumor progression at a metastatic site. Methods: Human cancer cell lines and primary human carcinoma-associated fibroblasts isolated from patients’ ovarian cancer metastases were used in multiple biochemical assays to analyze the role of CAFs in c-Met receptor signaling. Results: Using a receptor tyrosine kinase array to analyze phosphorylation of 42 different receptor tyrosine kinases (RTKs), we found that c-Met was the only RTK that was phosphorylated in cancer cells upon co-culture with CAFs. Further analysis of the co-culture revealed that direct contact between cancer cells and CAFs was required for activation of c-Met. Interestingly, the unique ligand for c-Met, hepatocyte growth factor (HGF), could not be detected in conditioned media from CAFs by ELISA and addition of HGF neutralizing antibodies to the co-culture did not prevent activation of the receptor. Furthermore, CAFs transiently transfected with HGF siRNA were still able to activate c-Met in the human ovarian cancer cell line SKOV3ip1 suggesting that c-Met activation by CAFs is HGF-independent. This effect was not limited to ovarian cancer cell lines as co-culture of CAFs with the cervical cancer cell line, HeLa, which expresses wild-type c-Met, and the breast cancer cell line, T47D, showed the same results as the two ovarian cancer cell lines, SKOV3ip1 and OVCAR-5. Remarkably, membrane fragments isolated from CAFs were sufficient to stimulate phosphorylation of c-Met in cancer cells in an HGF-independent manner. Several candidate factors were identified in the CAF membrane proteome which can activate c-Met signaling. Conclusions: Collectively, these results suggest a potentially novel mechanism whereby CAFs activate c-Met in cancer cells through direct contact which provides an additional rationale for the therapeutic targeting of c-Met in cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 48. doi:1538-7445.AM2012-48

Activation state of stromal inflammatory cells in murine metastatic pancreatic adenocarcinoma

The histologic presence of macrophages (tumor-associated macrophages, TAMs) and neutrophils (tumor-associated neutrophils, TANs) has been linked to poor clinical outcomes for solid tumors. The exact mechanism for this association with worsened prognosis is unclear. It has been theorized that TAMs are immunomodulated to an alternatively activated state and promote tumor progression. Similarly, TANs have been shown to promote angiogenesis and tumor detachment. TAMs and TANs were characterized for activation state and production of prometastatic mediators in an immunocompetent murine model of pancreatic adenocarcinoma. Specimens from liver metastases were evaluated by immunofluorescence and immunoblotting. TAMS have upregulated expression of CD206 and CD163 markers of alternative activation, (4.14 ± 0.55-fold and 7.36 ± 1.13-fold over control, respectively, P < 0.001) but do not have increased expression of classically activated macrophage markers CCR2 and CCR5. TAMs also express oncostatin M (OSM). We found that TANs, not TAMs, predominantly produce matrix metalloproteinase-9 (MMP-9) in this metastatic tumor microenvironment, while MMP-2 production is pan-tumoral. Moreover, increased expression of VEGF colocalized with TAMs as opposed to TANs. TAMs and TANs may act as distinct effector cells, with TAMs phenotypically exhibiting alternative activation and releasing OSM and VEGF. TANs are localized at the invasive front of the metastasis, where they colocalize with MMP-9. Improved understanding of these interactions may lead to targeted therapies for pancreas adenocarcinoma.

Correlated Immunohistochemical and Cytological Assays for the Prediction of Hematogenous Dissemination of Breast Cancer

Although metastasis is a major cause of death from breast cancer, our ability to predict which tumors will metastasize is limited (American Cancer Society 2010). Proper assessment of metastatic risk and elucidating the underlying mechanisms of metastasis will help personalize therapy and may provide insight into potential therapeutic targets. Traditionally, histologic grading, staging, hormone receptors, HER2/Neu, and proliferation assays have been the gold standard on which oncologists based their treatment decisions. However, all of these are indirect measures of metastatic risk. Recent insights from intravital imaging directly address questions of mechanism and have led to a new way of using histologic and cytologic material to assess metastatic risk. This review describes the tumor microenvironment model of invasion and intravasation, as well as an emerging histopathologic application based on this model. In particular, the authors describe a new immunohistochemical approach to the assessment of metastatic risk based on the density of intravasation microenvironment sites called the tumor microenvironment of metastasis. In addition, they describe an isoform assay for the actin regulatory protein Mena using fine needle aspiration samples and the details about how these 2 assays may be applied in clinical practice in a synergistic way to assess the risk of metastasis.

Tumor Immune Escape Mechanisms that Operate During Metastasis

Immune cells actively influence, among other factors, each step of tumor development determining the chance of a cancer cell to survive in a threaten microenvironment. Antitumor immune-mediated mechanisms are activated as soon as the first cancer cell is detected and operate both during primary tumor formation and during metastasis. However, when both innate and adaptive immunity becomes impaired, tumor development occurs. In this sense, compelling evidences indicate that tumor cells employ mechanisms that circumvent or thwart the immune response to enhance their own growth. These mechanisms include the secretion of immunosuppressive factors and the induction of distinct regulatory lymphoid or myeloid cells and, as occur with the immune response, they operate both during primary tumor formation and metastasis. Interestingly, cellular and molecular mechanisms of the immune response are important components of the tumor microenvironment and have the ability to promote or suppress tumor progression depending of the context of each cell interaction. In that sense, researchers are focusing their attention in the study of the influence of the tumor microenvironment in tumor growth and metastasis to better understand cancer biology and to formulate novel therapeutic approach. This review will focus on the present knowledge about interaction between immune cells and tumors in the context of metastasis, discussing the participation of different components of innate and adaptive immune response in the process of metastasis formation and dissemination.

Tumor microenvironment in cancer treatment and metastasis

Tumor microenvironment in cancer treatment and metastasis

NFAT1 transcription factor is central in the regulation of tissue microenvironment for tumor metastasis.

Members of the nuclear factor of activated T cell (NFAT) family of transcription factors were originally described in T lymphocytes but later shown to be expressed in several immune and non-immune cell types. NFAT proteins can modulate cellular transformation intrinsically, and NFAT-deficient (NFAT1-/-) mice are indeed more susceptible to transformation than wild-type counterparts. However, the contribution of an NFAT1-/- microenvironment to tumor progression has not been studied. We have addressed this question by inoculating NFAT1-/- mice with B16F10 melanoma cells intravenously, an established model of tumor homing and growth. Surprisingly, NFAT1-/- animals sustained less tumor growth in the lungs after melanoma inoculation than wild-type counterparts. Even though melanoma cells equally colonize NFAT1-/- and wild-type lungs, tumors do not progress in the absence of NFAT1 expression. A massive mononuclear perivascular infiltrate and reduced expression of TGF-β in the absence of NFAT1 suggested a role for tumor-infiltrating immune cells and the cytokine milieu. However, these processes are independent of an IL-4-induced regulatory tumor microenvironment, since lack of this cytokine does not alter the phenotype in NFAT1-/- animals. Bone marrow chimera experiments meant to differentiate the contributions of stromal and infiltrating cells to tumor progression demonstrated that NFAT1-induced susceptibility to pulmonary tumor growth depends on NFAT1-expressing parenchyma rather than on bone marrow-derived cells. These results suggest an important role for NFAT1 in radio-resistant tumor-associated parenchyma, which is independent of the anti-tumor immune response and Th1 versus Th2 cytokine milieu established by the cancer cells, but able to promote site-specific tumor growth.

Mena deficiency delays tumor progression and decreases metastasis in polyoma middle-T transgenic mouse mammary tumors

IntroductionThe actin binding protein Mammalian enabled (Mena), has been implicated in the metastatic progression of solid tumors in humans. Mena expression level in primary tumors is correlated with metastasis in breast, cervical, colorectal and pancreatic cancers. Cells expressing high Mena levels are part of the tumor microenvironment for metastasis (TMEM), an anatomical structure that is predictive for risk of breast cancer metastasis. Previously we have shown that forced expression of Mena adenocarcinoma cells enhances invasion and metastasis in xenograft mice. Whether Mena is required for tumor progression is still unknown. Here we report the effects of Mena deficiency on tumor progression, metastasis and on normal mammary gland development.MethodsTo investigate the role of Mena in tumor progression and metastasis, Mena deficient mice were intercrossed with mice carrying a transgene expressing the polyoma middle T oncoprotein, driven by the mouse mammary tumor virus. The progeny were investigated for the effects of Mena deficiency on tumor progression via staging of primary mammary tumors and by evaluation of morbidity. Stages of metastatic progression were investigated using an in vivo invasion assay, intravital multiphoton microscopy, circulating tumor cell burden, and lung metastases. Mammary gland development was studied in whole mount mammary glands of wild type and Mena deficient mice.ResultsMena deficiency decreased morbidity and metastatic dissemination. Loss of Mena increased mammary tumor latency but had no affect on mammary tumor burden or histologic progression to carcinoma. Elimination of Mena also significantly decreased epidermal growth factor (EGF) induced in vivo invasion, in vivo motility, intravasation and metastasis. Non-tumor bearing mice deficient for Mena also showed defects in mammary gland terminal end bud formation and branching.ConclusionsDeficiency of Mena decreases metastasis by slowing tumor progression and reducing tumor cell invasion and intravasation. Mena deficiency during development causes defects in invasive processes involved in mammary gland development. These findings suggest that functional intervention targeting Mena in breast cancer patients may provide a valuable treatment option to delay tumor progression and decrease invasion and metastatic spread leading to an improved prognostic outcome.

Serotonin is a critical signalling molecule in the local and metastatic small intestinal neuroendocrine tumour microenvironment

Serotonin is a critical signalling molecule in the local and metastatic small intestinal neuroendocrine tumour microenvironment

Modeling metastasis in the mouse

Metastasis is a complex clinical and biological problem presently under intense study, and several model systems are in use to experimentally recapitulate and dissect the various steps of the metastatic process. Genetically engineered mouse models provide faithful renditions of events in tumor progression, angiogenesis, and local invasion that set the stage for metastasis, whereas engrafting of human or mouse tumor tissues into mouse hosts has been successfully exploited to investigate metastatic dissemination and colonization of distant organs. Real-time, high-resolution microscopy in live animals, and comprehensive genetic and molecular profiling are effective tools to interrogate diverse metastatic cancer cell phenotypes as well as the metastatic tumor microenvironment in different organs. By integrating the information obtained with these complementary approaches the field is currently obtaining an unprecedented level of understanding of the biology, molecular basis, and therapeutic vulnerabilities of metastasis.

Abstract 4219: Biomarkers of cancer stem cells and invasion can predict breast cancer metastasis

Abstract Background: Breast cancer mortality is largely attributable to the development of systemic, hematogenous metastatic disease. Although nearly 10-15% of patients have an aggressive form disease that metastasizes within three years after initial diagnosis, the clinical manifestations of metastatic disease can appear ten or more years later. To decrease the risk for the emergence of metastatic tumors, nearly 80% of the patients are treated with adjuvant chemotherapy. The clinical benefit is a 3-10% increase in 15-yr survival, depending upon patient age at diagnosis. Currently established prognostic criteria such as the histopathological grade of the tumor or tumor size do not successfully predict systemic metastatic potential. An enrichment of cancer stem cells in the primary tumor has been shown to be a poor prognostic marker in patients. Epithelial to Mesenchymal Transition (EMT) in breast cancer cells have been shown to give rise to cells that have cancer stem cell like properties. Both of these cells types have been shown to be highly invasive. We have identified invasion markers that can predict EMT, drug and radio resistance in breast cancer cells. In this study we are using biomarkers for cancer stem cells (CD44+/CD24−) and markers of invasion (Mena/Mena11a) to identify cancer initiating cells in Fine Needle Aspiration (FNA). We correlate these ratios with a microanatomical structure called Tumor Microenvironment of Metastasis (TMEM) comprising of a direct apposition of an endothelial cell, a cancer cell and a stromal macrophage in the breast cancer tissue. TMEM has been previously reported to strongly correlate with metastasis. Hypothesis: The percentage of CD44+/CD24− expression can be used to estimate the number of cancer stem cells in FNA samples. The ratio of Mena to Mena11a protein can be used as a measurement of tumor cell response to the microenvironment and its metastatic phenotype. Measuring the stem cell biomarkers and Mena isoforms in cells collected by FNA biopsy using FACS and combining that with the assessment of TMEM density we will be able to develop a simple, quick and inexpensive method of determining metastatic potential of an individual tumor. Methods: FNA was conducted on lumpectomy or mastectomy tissues using fine needles, quality of FNA determined by smear staining. Cells were enzymatically digested, fixed and permeabilized. They were then be subjected to flowcytometry analysis using antibodies against stem cell and invasion markers. Rest of the tumor was processed for histopathology and TMEM analysis. Statistical analysis was performed to identify correlation. Results: Our initial results show that there is a strong correlation between CD44+/CD24− cells in the FNA with TMEM scores. We also report a very high correlation with can also be predicted by the ratios of different splice variants of Mena. This method can be easily used in the clinical setting and applied to a number of different biomarkers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4219.

Abstract PL04-02: Microenvironments of carcinoma cell dissemination and metastasis: Insights derived in vivo

Abstract The in vivo invasion assay (reviewed in [1]) was developed to collect the tumor cells observed in the multiphoton microscope to migrate toward and intravasate into blood vessels [2, 3]. The in vivo invasion assay has been coupled to expression profiling of small numbers of invasive cells to reveal the identities of the genes correlated with the survival, adjuvant-resistance and chemotaxis to macrophage supplied EGF, all phenotypes of these migratory tumor cells [4-6]. These genes fall into well defined pathways and are coordinately regulated in metastatic tumor cells [6-8]. These pathways are collectively called the Invasion Signature. Multi-photon microscopy (MPM) has been used to test the importance, in invasion and metastasis, of the pathways identified in the Invasion Signature including the ZBP1 [5, 9], ROCK [10], Mena [11], cofilin [12, 13] and EGF receptor [14, 15] pathways. The results of these studies confirm that the motility pathways are synergistic to create tumor cells that have passed through EMT and are capable of chemotaxis to EGF and penetration of basement membrane barriers using invadopodia (reviewed in [16, 17]). They also lead to a new insight that the motility pathways of invasive mammary tumor cells converge through the Mena pathway making the protein Mena a potentially important marker for prognosis and therapy [11, 18]. As described above, MPM had demonstrated that invasive carcinoma cells in mouse and rat mammary tumors intravasate when associated with peri-vascular macrophages thereby identifying a tumor microenvironment of intravasation as an anatomical landmark in tumors [19, 20]. Using this multiphoton imaging-defined anatomical landmark composed of an invading carcinoma cell marked by Mena over-expression, and a peri-vascular macrophage as a guide, it was possible to define a triple stain marker for use in anatomic pathology with anti-bodies against Mena, macrophages and endothelial cells to find the same landmark in human breast tumors [21]. In humans this landmark for intravasation is called TMEM (Tumor Microenvironment of Metastasis) and its density is correlated with metastatic risk in breast cancer patients [21]. This work illustrates the power of combining multi-photon imaging at single cell resolution with mouse models of breast cancer to develop new insights into the mechanisms of human breast cancer metastasis, and new prognostic markers for clinical use.

Abstract 1441: Use of RFP and GFP transgenic nude mice to study the tumor microenvironment of liver metastasis

Abstract The tumor microenvironment (TME) is critical for tumor progression in the liver. To examine the role of the TME in liver metastasis, we developed color-coded imaging of the TME using green fluorescent protein (GFP) transgenic and red fluorescent protein (RFP) transgenic nude mice, to investigate cancer-host-cell interaction. Non-colored human colon cancer cells (HCT-116, 2×106 / 50 µl) were injected in the spleen of GFP or RFP nude mice. Fluorescence imaging of the metastatic liver tumors used the Olympus OV100 small animal imaging system. The fluorescence area of the liver was measured and liver sections were stained with hematoxylin and eosin and desmin. HCT-116 cells formed tumor colonies in the liver 28 days after cell transplantation to the spleen. In the metastatic liver tumors in RFP nude mice, RFP-expressing stroma cells derived from the host animal appeared around the edge of HCT-116 tumors. The infiltration of RFP-expressing inflammatory cells was observed in the central necrotic area of the tumors. The fluorescence of the metastatic liver tumors in the GFP nude mice was weaker than that from the RFP mice. In the GFP mouse, only non-parenchymal cells of the liver had GFP fluoresecence. The GFP non-parenchymal cells greatly increased in the liver during tumor-colony growth and surrounded the metastatic tumors. Desmin positive cells predmoninated in the non-parenchymal and increased around the metastatic tumors suggesting that fibroblasts are greatly increased around the metastatic tumors in the liver and may have a role in tumor progression. This novel imaging model of the TME will enable the identification and characterization of stromal cells involved in cancer progression in the liver. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1441.

MenaINV Isoform Expression in Human Invasive Ductal Carcinomas of the Breast Correlates with Tumor Microenvironment of Metastatsis (TMEM) Score; a Pilot Study Indicating the Potential Use of MenaINV/11a Ratio as a Predictor of Distant Metastases.

Abstract Background: Primary mammary carcinomas in rodents contain a subpopulation of invasive tumor cells that move towards perivascular macrophages and intravasate. An isoform of actin regulatory protein Mena, MenaINV is up-regulated in these invasive cells, while isoform Mena 11a is down regulated (Goswami et al 2009). The direct apposition of Mena positive tumor cells to endothelial cells and perivascular macrophages has been described as Tumor Microenvironment of Metastasis (TMEM) and its density is shown to be a predictor of metastatic outcome in human invasive ductal carcinomas of the breast (Robinson et al 2009). We questioned if MenaINV and Mena 11a isoform expression correlates with the invasiveness of breast lesions in Polyoma Middle T antigen (PyMT) mouse mammary tumors and human invasive ductal carcinomas (IDC), as well as if increased MenaINV expression is associated with high TMEM counts in human IDCs.Materials and Methods: We analyzed MenaINV and Mena 11a expression in fine needle aspiration biopsy (FNA) samples from 18 mice with PyMT mammary tumors, 3 human IDCs and 1 fibroadenoma (FA) by qRT-PCR. The expression of Mena isoforms from in situ PyMT lesions was compared with metastatic PyMT tumors and expressed as fold change. Mena isoform levels from IDC were compared to that in FA and expressed as fold change. Representative sections from IDCs were triple immunostained with anti-Mena, anti-CD 68 and anti-CD34 and independently analyzed for TMEM counts on ten x400 fields by two pathologists.Results: We observed a 7.5 fold increase in MenaINV expression and a 3 fold decrease of Mena 11a in metastatic PyMT tumors when compared to in situ lesions. The results from human samples are shown in Figure 1. The top panel shows MenaINV and Mena 11a fold change in IDCs compared to FA. The bottom panel shows TMEM counts for each IDC and MenaINV/11a ratios. Both, MenaINV expression and TMEM count varied significantly, whereas Mena 11a was exceedingly low in IDCs. In all 3 IDCs the levels of MenaINV correlated with TMEM counts.Discussion: This pilot study indicates for the first time that the expression of MenaINV and Mena 11a can be successfully analyzed in FNA biopsy samples from human breast lesions. In combination with the TMEM count, Mena isoforms can potentially become useful markers for predicting the metastatic potential of an individual breast lesion. A larger study is underway to develop a scoring system combining Mena isoform expression with the TMEM count.Figure 1: Top panel: qRT-PCR assessment of Mena splice variant expression in FNA samples from human breast lesions. Levels of MenaINV and Mena 11a in 3 IDCs were compared to those in FA. The result is expressed as fold change. Bottom panel: TMEMs on tissue sections from IDC with TMEM counts and MenaINV/11a ratios. Circles represent TMEMs. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 2140.

Metastasis: Alone or Together?

Summary Recent studies of carcinoma progression reveal the distinct routes of dissemination of discrete carcinoma cell populations and suggest that melanoma cell dissemination is linked to differentiation rather than stemness status.

Tumor Microenvironment of Metastasis (TMEM): A Novel Tissue-Based Assay for Metastatic Risk in Breast Cancer

Tumor Microenvironment of Metastasis (TMEM): A Novel Tissue-Based Assay for Metastatic Risk in Breast Cancer

Tumor Microenvironment of Metastasis in Human Breast Carcinoma: A Potential Prognostic Marker Linked to Hematogenous Dissemination

Multiphoton-based intravital imaging has shown that invasive carcinoma cells in mouse and rat mammary tumors intravasate when associated with perivascular macrophages, identifying a potential tumor microenvironment of metastasis (TMEM). We define TMEM as the tripartite arrangement of an invasive carcinoma cell, a macrophage, and an endothelial cell. The aim of this study was to determine if TMEM density in human breast carcinoma samples predicts the development of systemic, hematogenous metastases. A case-control study of 30 patients who developed metastatic breast cancer and 30 patients without metastatic disease was done. Cases were matched to controls based on currently used prognostic criteria. Paraffin-embedded primary breast cancer samples were stained using a triple immunohistochemical method allowing simultaneous identification of carcinoma cells, macrophages, and endothelial cells. Two pathologists, blinded to outcome, evaluated the number of TMEM per 20 high-power fields. No association was seen between TMEM density and tumor size or grade, lymph node metastasis, lymphovascular invasion, or hormone receptor status. TMEM density was greater in the group of patients who developed systemic metastases compared with the patients with only localized breast cancer (median, 105 versus 50, respectively; P = 0.00006). For every 10-unit increase in TMEM density, the odds ratio for systemic metastasis was 1.9 (95% confidence interval, 1.1-3.4). TMEM density predicted the development of systemic, hematogenous metastases. The ability of TMEM to predict distant metastasis was independent of lymph node status and other currently used prognosticators. Quantitation of TMEM may be a useful new prognostic marker for breast cancer patients.

Angiogenesis and intravasation of PC-3 dissemination variants

Angiogenesis and intravasation of PC-3 dissemination variants

Tumor microenvironment in cancer progression and metastasis

Tumors are unorganized organs that contain many different cell types that communicate with cancer cells. The central goal of our laboratory is to evaluate the role of these non-cancer cells in canc...

The role of tumor microenvironment in prostate cancer bone metastasis

Prostate cancer (PCa) epithelial cells require a number of factors to facilitate their establishment and growth at a distant site of metastasis. Their ability to adapt to their microenvironment, proliferate and recruit an underlying stroma is integral to the survival and growth of the metastasis. PCa predominantly metastasizes to the bone, and bone metastases are the main cause of morbidity. The bone marrow provides a permissive environment for the formation of a metastasis. In some cases, the cells may remain dormant for some time, eventually proliferating in response to an unknown "trigger." The marrow is rich in progenitor cells that differentiate into numerous cell types, producing new blood vessels, supporting fibroblasts, and an underlying extracellular matrix (ECM) that form the reactive stroma. By secreting a number of cytokines, growth factors and proteases they recruit auxiliary cells required to produce a functional stroma. These components are involved in a reciprocal interaction between the stroma and the PCa cells, allowing for the growth and survival of the tumor. Left unchecked, once a PCa tumor has established itself in the bone marrow it will eventually replace the marrow, interrupting bone homeostasis and typically promoting an osteoblastic response in the bone including osteoclastic events. The abundant deposition of new woven bone results in nerve compression, bone pain and an increase in fractures in patients with PCa bone metastases. This review will examine the tumor microenvironment, its role in facilitating tumor dissemination, growth and the resultant pathologies associated with PCa bone metastasis.

Oestrogen receptor‐β expression in melanocytic lesions

Melanomas rarely occur before puberty, have a higher death rate for males, and tend to be more invasive during pregnancy. Prior to the discovery of a second oestrogen receptor (ERbeta), studies with the initial oestrogen receptor, ERalpha, showed no obvious role for oestrogen in the pathophysiology of benign or malignant melanocytic lesions. To investigate the specific immunostaining patterns of ERalpha and ERbeta, benign nevocytic nevi, dysplastic nevi with mild, moderate and severe cytological atypia, lentigo malignas and melanomas of varying depth (Clark) and thickness (Breslow) were studied. ERbeta but not ERalpha was the predominant oestrogen receptor we found in all types of benign and malignant melanocytic lesions. The most intense ERbeta immunostaining was seen in melanocytes in dysplastic nevi with severe cytological atypia and in lentigo malignas. ERbeta expression levels also correlated with the malignant tumor microenvironment; i.e., melanocytes in proximity with keratinocytes>deeper dermal melanocytes in contact with stroma>minimally invasive melanomas>Clark Level III/IV or thick melanomas (Breslow). Discovery that ERbeta expression varies in relation to the tumor microenvironment and increasing depth of invasion suggests its possible usefulness as a surrogate marker for neoplasia and prognosis in malignant melanoma.