Angiotropism is the histopathological correlate of pericytic mimicry and extravascular migratory metastasis (EVMM), a mechanism of melanoma spread by migration along the external surface of blood and lymphatic vessels. The frequency of angiotropism in primary cutaneous melanoma and the clinical utility of its detection remain unclear.

We investigated angiotropism in 179 primary cutaneous melanomas by hematoxylin and eosin (H&E), CD31, and S100/D240 stains.

We detected angiotropism in 31 cases (17%) by H&E. CD31 immunohistochemistry increased detection to 59 cases (33%). When lymphatic vessels were included by using S100/D240 stains, 67 cases (37%) cases were positive. Angiotropism was associated with lymphatic invasion and mitotic rate with all detection methods. There was an association with increased tumor thickness when detected by H&E and CD31. No association with sentinel lymph node status was seen. By H&E and CD31 staining, angiotropism was associated with disease progression and distant metastases by univariate, but not multivariate analysis. Overall survival was not affected by the presence of angiotropism.

Angiotropism is relatively common in primary melanoma when immunohistochemical stains are used for detection and associated with mitotic rate and intravascular lymphatic invasion. The association with disease progression and distant metastasis suggests that it represents an alternative pathway of metastasis, that is, EVMM/pericytic mimicry vs intravascular spread.

Access of melanoma cells to the cutaneous vasculature either via lymphatic invasion or angiotropism is a proposed mechanism for metastasis. Lymphatic invasion is believed to be a mechanism by which melanoma cells can disseminate to regional lymph nodes and to distant sites and may be predictive of adverse outcomes. Although it can be detected on hematoxylin- and eosin-stained sections, sensitivity is markedly improved by immunohistochemistry for lymphatic endothelial cells. Multiple studies have reported a significant association between the presence of lymphatic invasion and sentinel lymph node metastasis and survival. More recently, extravascular migratory metastasis has been suggested as another means by which melanoma cells can spread. Angiotropism, the histopathologic correlate of extravascular migratory metastasis, has also been associated with melanoma metastasis and disease recurrence. Although lymphatic invasion and angiotropism are not currently part of routine melanoma reporting, the detection of these attributes using ancillary immunohistochemical stains may be useful in therapeutic planning for patients with melanoma.

An eleven-year-old spayed female Yorkshire Terrier presented with a sublumbar mass and upon ultrasonographic examination, was revealed to have a mammary gland tumor. Black to reddish colored masses, located in the visceral peritoneum of the sublumbar region was observed on laparotomy with masectomy of the right side. In the laparotomy, we observed reddish masses multifocally located in the serosal membrane of the large intestine. Histopathologic examination of the intestinal and abdominal mass showed highly invasiveness into the muscle and metastasis of melanocytic tumor cells through the blood vessels. The mammary glands showed abnormal hyperplasia of melanocytes, destruction of the normal glands by tumor cells and infiltration of some lymphocytes in the pool of melanocytic cells. We have identified a malignant melanoma containing an angiotumoral complex in which tumor cells occupied a pericytic location along the microvessels with intravasation determined by immunohistochemistry for S100 protein and protein kinase C-α. Histologic findings in this dog lead to a diagnosis of an angiotropic metastatic malignant melanoma.

Tetraspanin protein CD151 on tumor cells supports invasion and metastasis. In the present study, we show that host animal CD151 also plays a critical role. CD151-null mice showed markedly diminished experimental lung metastasis after injection of Lewis lung carcinoma or B16F10 melanoma cells. Diminished tumor cell residence in the lungs was evident 6-24 hours after injection. Consistent with an endothelial cell deficiency, isolated CD151-null mouse lung endothelial cells showed diminished support for B16F10 adhesion and transendothelial migration, diminished B16F10-induced permeability, and diminished B16F10 adhesion to extracellular matrix deposited by CD151-null mouse lung endothelial cells. However, CD151 deletion did not affect the size of metastatic foci or subcutaneous primary B16F10 tumors, tumor aggregation, tumor clearance from the blood, or tumor-induced immune cell activation and recruitment. Therefore, the effects of host CD151 on metastasis do not involve altered local tumor growth or immune surveillance. VEGF-induced endothelial cell signaling through Src and Akt was diminished in CD151-null endothelial cells. However, deficient signaling was not accompanied by reduced endothelial permeability either in vitro (monolayer permeability assay) or in vivo (VEGF-stimulated Miles assay). In summary, diminished metastasis in CD151-null host animals may be due to impaired tumor-endothelial interactions, with underlying defects in mouse lung endothelial cell extracellular matrix production.

Transplant-related malignancies are a major contributor to morbidity and mortality in the organ-recipient population, and most often develop de novo in the immunosuppressed recipient or as recurrent malignancy after transplantation. The least common scenario, and a rare event, is a recipient malignancy derived from the donor organ. Melanoma is one of the most often reported and lethal donor-derived malignancies with a high transmission rate. Donor transmission of melanoma might be related to the biology of melanoma, with regard to tumour dormancy, late recurrence, circulating tumour cells, and the destiny of some micrometastases. Melanoma-cell dormancy explains the late recurrence that can occur after the initial treatment of melanoma, and may be relevant to our understanding and management of some melanoma micrometastasis in the sentinel node. The high incidence of circulating tumour cells in early melanoma should be considered in the context of the transmission of melanoma by apparent disease-free organ donors following removal of a primary melanoma up to 32 years before. This scenario suggests that melanoma cells can remain dormant at distant sites for decades (and possibly forever) in immunocompetent patients, only to reactivate after transplantation into an immunosuppressed recipient. Potential organ donors should be carefully screened for a history of melanoma, and excluded. The current recommendation for treatment of donor-related melanoma includes withdrawal of immunosuppression, graft rejection, and explantation of the allograft after rejection has been established. In non-renal transplant patients with life-sustaining organs, withdrawal of immunosuppression and graft rejection is not feasible, and reduction of immunosuppression or urgent retransplantation are the only possible salvage strategies. The transmission of malignancy by organ donation could be considered "nature's own experiment", but raises questions that our current understanding of the biology of melanoma cannot answer.

Significant prognostic information is available in a routine melanoma dermatopathology report. Features that are enumerated in the pathology report and that portend a potentially poorer prognosis are older age, site (acral, head, neck), male sex, increasing Breslow tumor thickness, increasing Clark's level, ulceration, increasing number of mitoses, vertical growth phase, regression, absence of a host inflammatory response, increased tumor vascularity, angiotropism, vascular invasion, neurotropism, marked atypia, and satellite metastasis.

It is a basic tenet of the sentinel lymph-node biopsy procedure that all positive sentinel lymph nodes will inevitably progress to palpable nodal recurrence if not removed. Comparison of survival is, therefore, considered permissible among patients with positive sentinel lymph nodes who undergo early lymphadenectomy with that among patients who have delayed lymphadenectomy for palpable regional node metastasis, providing that survival is calculated from the date of wide local excision of the primary tumor. Here, that fundamental assumption is contested and evidence is presented to show that a positive sentinel lymph node might have no adverse prognostic relevance in up to one-third of patients. Furthermore, in the same patients, progression to palpable nodal disease might not have occurred even if the positive sentinel node had not been removed. The term prognostic false-positivity is used to describe this phenomenon. Such patients are incorrectly up-staged, are given inaccurate prognostic information and can undergo unnecessary completion lymphadenectomy and unnecessary adjuvant therapy.

PEgr-Endostatin-EGFP plasmid was constructed to investigate its expression properties induced by ionizing irradiation and the effect of pEgr-Endostatin-EGFP gene-radiotherapy on melanoma tumor-bearing mice. The pEgr-Endostatin-EGFP plasmid was transfected into B16 cell line with liposome. The expression property of endostatin was investigated by RT-PCR and that of EGFP was detected by flow cytometry. Tumor-bearing mice were treated by the plasmid injection and 2 Gy X-irradiation of three fractions. Tumor growth was observed for 18 days after treatment. Change of tumor capillary formation was measured with histochemistry assay at the end of the experiment. The expression of GFP in B16 melanoma cells was detected after X-irradiation with 0.05-20 Gy. Time-course studies showed that the expression of GFP in B16 cells reached its peak at 8 h after irradiation with 2 Gy. The injection of pEgr-Endostatin-EGFP recombinant plasmid into the implanted B16 melanoma in C57BL/6J mice followed by local X-irradiation could significantly inhibit tumor growth with inhibition of intratumor micro-vessel density. The inhibitory effect of pEgr-Endostatin-EGFP gene-radiotherapy on the growth of B16 melanoma is correlated with the marked decrease of intratumoral vascularization. The present data point to the potential of an anti-angiogenic approach in gene-radiotherapy of cancer.

It has been suggested that performing a sentinel node biopsy (SNB) in patients with cutaneous melanoma increases the incidence of in-transit metastasis (ITM).

ITM rates for 2018 patients with primary melanomas > or =1.0 mm thick treated at a single institution between 1991 and 2000 according to 3 protocols were compared: wide local excision (WLE) only (n = 1035), WLE plus SNB (n = 754), and WLE plus elective lymph node dissection (n = 229).

The incidence of ITM for the three protocols was 4.9%, 3.6%, and 5.7%, respectively (not significant), and as a first site of recurrent disease the incidence was 2.5%, 2.4%, and 4.4%, respectively (not significant). The subset of patients who were node positive after SNB and after elective lymph node dissection also had similar ITM rates (10.8% and 7.1%, respectively; P = .11). On multivariate analysis, primary tumor thickness and patient age predicted ITM as a first recurrence, but type of treatment did not. Patients who underwent WLE only and who had a subsequent therapeutic lymph node dissection (n = 149) had an ITM rate of 24.2%, compared with 10.8% in patients with a tumor-positive sentinel node treated with immediate dissection (n = 102; P = .03).

Performing an SNB in patients with melanoma treated by WLE does not increase the incidence of ITM.

To report several samples of invasive human prostate cancer showing angiotropism, and to use human prostate cancer cells stably expressing green fluorescence protein (GFP) in in vitro and in vivo models to assess the dissemination pathway of prostate cancer cells.

Malignant melanoma and prostate carcinoma cells can migrate along anatomical structures such as nerves; previous studies showed that melanoma cells can be perivascular, on the outside of the endothelium, i.e. they are angiotropic, which suggests the hypothesis that melanoma cells also may migrate along vascular channels, termed 'extravascular migratory metastasis' (EVMM). Thus we examined histologically 10 human prostatic carcinoma specimens for the presence of angiotropism. In vitro, the PC-3 prostate cancer cells were co-cultures with capillary-like structures. In vivo, PC-3 cells were implanted on the chick chorio-allantoic membrane (CAM).

Histologically, in all 10 cases, angiotropism was detected at least focally within the tumour or at the advancing front of the tumour. In vitro, the PC-3 cells spread along the external surface of the vascular tubules; in vivo, PC-3 cells formed a cuff around some vessels a few millimetres beyond the tumour, showing angiotropism. Histopathology of the CAM confirmed the perivascular location of tumour cells and the absence of tumour cells within the vessel lumina.

The presence of angiotropic tumour cells in human invasive prostate cancers, associated with the angiotropism of GFP prostate cancer cells cultivated in vitro and in vivo in angiogenic models, raises the possibility that some prostate tumour cells may migrate along the external surface of vessels as a mechanism of spread, i.e. EVMM.

There is no information about the effect of polysaccharides from fungus, Phellinus gilvus (PG) on melanoma. The effect of PG on the proliferation and apoptosis of the B16F10 melanoma cell line was determined by a sulforhodamine B (SRB) and a sandwich enzyme-linked immunosorbent assay. The in vivo effect of PG on B16F10 melanoma cells allografted in athymic nude mice was investigated. PG decreased cell proliferation and increased cell apoptosis in a dose dependent manner in vitro. Also, PG significantly inhibits melanoma growth in mice. The PG anti-tumor effect in vivo was associated with a significant increase in the melanoma apoptosis rate. These findings support PG as a therapeutic agent against melanoma.

We have identified in malignant melanoma an angio-tumoural complex in which tumour cells occupy a pericytic location along the endothelium of microvessels without evidence of intravasation. This pericytic angiotropism of melanoma cells, without any sign of intravasation, suggests that melanoma cells may migrate along the external surface of vessels, a mechanism we have termed 'extravascular migratory metastasis' (EVMM), as distinct from intravascular dissemination.

The present study describes, for the first time, a series of 36 invasive melanoma cases ascertained for the presence of the histopathological characteristic of angiotropism.

All cutaneous melanomas (31/35) were level IV with the exception that two melanomas were level II and two level V. In all cases, angiotropism was easily observed at the advancing front of the tumour or in nearby tissue.

The study demonstrates that angiotropism of melanoma cells can be easily detected microscopically in routine tissue sections, i.e., in close proximity to microvessels (in a pericytic location). This phenomenon may prove to be important both biologically and prognostically in the routine histopathological assessment of melanoma, since we have recently shown that angiotropism could be a prognostic factor predicting risk for metastasis of melanoma. Our continued investigations to elucidate the significance of angiotropism in melanoma may help in understanding the molecular basis of metastasis and EVMM.

Previous studies have demonstrated that some tumor cells occupy a pericyte-like location in melanoma, forming angio-tumoral complexes. We hypothesized that these tumor cells are migrating along the abluminal surface of the endothelium, a mechanism termed "extravascular migratory metastasis." In the present study, we have used human and murine melanoma cells that stably express enhanced green fluorescence protein (GFP) to examine, in an ex vivo co-culture model, melanoma cell interactions with vessels that have sprouted from rat aortic rings. We also used in vivo tumor growth on the chick chorioallantoic membrane (CAM) to observe the dissemination pathway of melanoma cells. In the ex vivo rat aorta system, we observed a pericyte-like location of tumor cells that were spreading along the vascular channels. For examination of the CAM in vivo, we have used the Lugassy preparation, allowing one to obtain striking images of the relationship between fluorescent GFP cells and microvessels. Melanoma cells were found cuffing the outside of vessels around the tumor. Tumor cells were observed along the vessels several centimeters from the tumor. Confocal microscopy and histopathology confirmed the pericyte-like location of tumor cells, without any observable intravasation. The results indicate that melanoma cells can migrate along the abluminal surface of vessels. This study also demonstrates that these models can provide quantitation analysis that may prove useful in elucidating the molecular interactions involved in extravascular migratory metastasis.

Lymphatic mapping and sentinel lymphadenectomy provide a minimally invasive means of directly determining the status of the regional lymph nodes in all patients who have a primary melanoma >1 mm thick but no clinical evidence of nodal involvement. Since the histological status of the sentinel node (SN) has been shown to be the most important prognostic factor in primary melanoma patients, the World Health Organization has recently recommended that sentinel lymphadenectomy should become the new standard of care for primary melanoma patients. This paper reviews the literature with regards to developments in and the current status of SN evaluation. Developments in the histopathological versus molecular detection of melanoma nodal metastases are reviewed, with specific emphasis on the strengths, limitations and clinical significance of these techniques. Molecular evaluation of the SN offers several advantages over standard histopathological analysis. These include an improved sensitivity, the cost-effective use of multiple markers for the improvement of detection rate and prognosis, as well as being less labour-intensive and costly. Moreover, molecular analysis has the potential to allow estimation of tumour burden. We review the potential causes of technical false-negative and false-positive reverse transcription-polymerase chain reaction (RT-PCR) results and how these could be eliminated by a systematic approach consisting of (i) careful and systematic assay design, which would include efficient tissue homogenization, choice of reagents and molecular markers, primer design and the use of one-stage versus two-stage PCR; (ii) careful optimization of the RT-PCR parameters (in particular the PCR cycle number) through the use of appropriate control tissues; and (iii) aiming for high assay reproducibility and lastly by applying the necessary positive and negative controls with each batch of samples. We also review the significant improvement in patient prognosis and management that has been made possible by the development of sentinel lymphadenectomy and histopathological evaluation of the SN, and compare the clinical (predictive) value of histopathological analysis with that of RT-PCR. Although RT-PCR is able to detect additional, clinically significant SN metastases that are missed by routine histopathology, its current limitation is that it overestimates the number of patients who have clinically significant melanoma metastases. Therefore, we suggest and discuss appropriate steps that need to be taken in order to minimize these false-positives and make this molecular tool more acceptable for routine clinical use.

Patients with primary cutaneous melanocytic lesions rely not only on the knowledge, skills, and experience of their treating clinician but also on the fundamentally important input of their pathologist for accurate diagnosis and appropriate management. Free and precise communication between pathologists and surgeons is important and undoubtedly improves patient care, particularly when managing difficult or complicated cases. To provide both patient and surgeon with the necessary information they require to make the most appropriate decisions, the pathology report should include all pathologic factors that are important in determining the patient's prognosis and management. Use of a synoptic format for pathology reporting of melanomas can facilitate this. Recent studies have established that the dermal mitotic rate of a primary cutaneous melanoma is a major prognostic determinant, and have shown that its assessment and that of other important histopathologic prognostic variables are reproducible between pathologists. Sentinel node (SN) biopsy has provided a minimally invasive procedure that can accurately predict the regional node status of melanoma patients. It is well demonstrated that the use of immunohistochemical stains assists in the detection of melanoma micrometastases in SNs, although it remains unclear which is the optimal pathologic protocol for SN evaluation and whether there is a role for reverse transcriptase polymerase chain reaction (RT-PCR) in SN assessment. False negative SN biopsies may occur as a result of errors in lymphatic mapping or sentinel lymphadenectomy, or because of a deficiency in the process of histopathologic evaluation. Recent studies have shown that the likelihood of non-SN involvement when the SN is positive correlates mostly with the extent of SN involvement, in particular the tumor penetrative depth (defined as the maximum distance of melanoma cells from the inner margin of the SN capsule). It appears that assessment of the micromorphometric features of positive SNs may be useful in predicting which patients have a low probability of having metastatic tumor in non-SNs, and therefore in selecting patients who potentially may be spared a completion lymph node dissection. It is likely that future advances in our understanding of the molecular biology of melanoma will provide new insights into tumor classification, improve diagnostic accuracy and prognostic ability, and lead to the development of more precisely targeted therapies.

Cancer is the second most common cause of death among Americans, although for several age groups it ranks first. Most of these deaths are not due to the primary tumour but rather to tumour cell metastases to distant organs. There are many steps that lead to metastasis, all of which are being studied with the goal of preventing these fatalities. Normally, cells attach to the extracellular matrix to maintain tissue integrity. During cancer progression, cells become more motile and acquire invasive qualities. Tumour cells move along blood and lymph vessels or invade into them to travel to distant sites. Then, the tumour cells must attach to the vessel wall, extravasate from the vessel, invade the new tissue, proliferate, and form a secondary tumour. Angiogenesis, the formation of new blood vessels, is critical to survival of these cells at the new site and is also important for primary tumour growth and spread. Tumour cell metastasis is a complex cascade of sequential steps, each of which is not yet fully understood. Progress has been made in identifying several key activators, one of which is the extracellular matrix. A major tumour promoter is the glycoprotein laminin, which is predominantly found in the extracellular matrix produced by endothelial and epithelial cells. This review will follow the metastatic process with particular attention to the effect of laminin on tumour cells.

Although the phenomenon of tumor-to tumor metastasis is not rare, it has been reported in only eight cases with malignant melanoma as the primary tumor.

This case describes a patient with cutaneous malignant melanoma that metastasized to an angiolipoma. To our knowledge this is the second case of a malignant melanoma metastasizing to another primary cutaneous tumor and the first to do so in a lipoma.

In this report we present the clinical and histopathologic features of this special case and review the relevant literature.

The annual incidence of malignant melanoma is estimated at 10-12 per 100000 inhabitants in countries of Central Europe and the US, with more recent estimates showing a dramatic upward trend. Taurolidine (Carter/Wallace, Cranberry, NJ) is a novel, potentially effective, antitumor chemotherapeutic agent. We hypothesized that Taurolidine could inhibit the growth, induce apoptosis, affect the cell cycle and change morphology of melanoma cells. We expected this process to be different in adherent and floating subpopulations that may be reflective of solid tumors and their metastases. Analysis of MNT-1 human and B16F10 murine melanoma cells showed that at 72 h the IC(50) of Taurolidine was 25.4+/-3.3 microM for MNT-1 human melanoma cells and 30.9+/-3.6 microM for B16F10 murine melanoma cells. Taurolidine induced DNA fragmentation of melanoma cells in a dose-dependent manner. Taurolidine (75 and 100 microM) induced 52-97% Annexin-V binding (apoptosis), respectively. Evaluation of cell cycle after 72 h exposure to Taurolidine (0-100 microM) revealed that the percentage of melanoma cells in S phase increased from 27 to 40% in the adherent subpopulation and from 33 to 49% in the floating subpopulation. Phase contrast microscopy revealed a marked swelling of melanoma cells and decreasing cell numbers in adherent subpopulation starting at 24 h with 25 microM Taurolidine. Shrinkage of cells dominated at 75-100 microM Taurolidine. Using Cytospin assay in the floating population, we observed swelling of melanoma cells induced by 25-100 micro Taurolidine and appearance of giant (multinuclear) forms resulting from exposure to 75-100 micro Taurolidine. Some floating cells with normal morphology were observed with low concentrations of Taurolidine (0-25 microM). These data show that effects of Taurolidine may be different in adherent and floating subpopulations of melanoma cells. More importantly, floating subpopulations that may contain some viable melanoma cells, may be reflective of potential metastasis after treatment of solid tumors in vivo.

Relapse of melanoma after surgical treatment remains a significant clinical problem in need of novel therapies. Vitamin E succinate (VES) is a promising antitumor micronutrient. We evaluated the effect of VES on melanoma dormancy and angiogenesis.

B16F10 melanoma cells were allografted in mice. The effect of VES on melanoma dormancy was measured by monitoring tumor volume. Tumor vascularity was quantitated with CD31 immunostaining. The expression of vascular endothelial growth factor (VEGF), VEGF receptor 1, and VEGF receptor 2 in tumors was assessed by the intensity of immunostaining. VES effect on secreted VEGF protein and VEGF promoter activity was measured with enzyme-linked immunosorbent assay and transient transfection assay, respectively. Significance was determined by analysis of variance.

VES promoted melanoma dormancy (P =.0019) and inhibited melanoma angiogenesis (P <.0001). VES also significantly suppressed the expression of VEGF, VEGF receptor 1, and VEGF receptor 2 in melanoma tumors (P <.0001). Melanoma VEGF secretion (P =.0077) and melanoma VEGF promoter activity (P <.05) were significantly inhibited by VES.

VES promotes melanoma dormancy and inhibits melanoma angiogenesis. The mechanism of the VES antiangiogenesis effect involves the inhibition of VEGF gene transcription. These findings support future studies of VES in the prevention of melanoma metastasis.