Identification of histological features associated with metastatic potential in thin (<1.0 mm) cutaneous melanoma with metastases. A study on behalf of the EORTC Melanoma Group. J Pathol; 197: 188–193

The original article to which this Erratum refers was published in The Journal of Pathology; 197: 188–193     

Regressing thin cutaneous malignant melanomas (< or = 1.0 mm) are associated with angiogenesis.

In previous studies, we have shown that angiogenesis is often first noted in cutaneous malignant melanomas (CMMs) under 1.0 mm in thickness. Because angiogenesis may signal a more aggressive tumor phenotype, it is important to establish the circumstances associated with onset of angiogenesis. In the present study, we have quantified tumor vascularity in a series of CMMs under 1.0 mm in thickness and either associated with or lacking histologic regression. Microvessels were identified with the lectin Ulex europaeus agglutinin I and the vessels in five fields counted within an ocular grid (area 7.84 x 10(-2) mm2) at 400 x magnification. CMMs (mean 0.48 mm) with regression had greater microvessel counts (27.2 +/- 5.1) compared with CMMs (mean 0.61 mm) without regression (mean 20.1 +/- 7.9) (P < 0.01). However, of particular interest, CMMs in the radial growth phase only and associated with regression (mean 0.40 mm) had strikingly greater vascularity (mean 28.7 +/- 6.9) versus radial growth phase CMMs (mean 0.44 mm) lacking regression (mean 16.4 +/- 6.6) (P = 0.0013). CMMs in the vertical growth phase (mean 0.81 mm) without regression had slightly less vascularity (mean 24.4 +/- 7.3) compared with vertical growth phase CMMs with regression (mean microvessels 27.2 +/- 5.1) (P = 0.1878) but significantly greater microvessels versus radial growth phase CMMs without regression (P = 0.0213). These results suggest that the onset of angiogenesis in thin CMMs is related to at least two phenomena: 1) inflammatory regression and 2) development of the vertical growth phase.     

Thin malignant melanomas (< 1.5 mm) with metastasis: a histological study and survival analysis

It is known that not all thin malignant melanomas have an excellent prognosis and that the specific features identifying the patients at risk of metastasis have not been fully elucidated. We have looked at thin malignant melanomas (less than 1.5 mm) in the East of Scotland that had proven metastasis and death, and compared the clinical and histological features with a similar group (less than 1.5 mm) that have had no further recurrence after a minimum of 6-year follow-up. We identified 26 patients with thin melanomas who had developed histologically proven metastasis and/or died following adequate surgical treatment of their primary lesion. When compared with the control group, factors found to be significantly different between the two groups and present in the group that did badly were (a) histological regression, (b) lesion size, (c) Clark level IV and (d) depth of the uninvolved dermis.     

A nationwide survey of observer variation in the diagnosis of thin cutaneous malignant melanoma including the MIN terminology. CRC Melanoma Pathology Panel.

AIM: To investigate observer variation in the diagnosis of thin cutaneous malignant melanoma and related lesions in a nationwide sample of histopathologists in the UK. METHODS: Out of a random sample of 195 pathologists, 148 (76%) participated in two circulations, the first with 20 slides and the second with 25 slides. The results were compared with those for the Cancer Research Campaign (CRC) Melanoma Pathology Panel, consisting of seven histopathologists and one dermatopathologist, which had developed and evaluated diagnostic criteria. RESULTS: In the first circulation, when no standardised diagnostic criteria were used, a fair level of agreement was achieved for an overall diagnosis using the categories benign naevi with no atypia, benign naevi with atypia and melanoma (Kappa = 0.45). This was low compared with the agreement of the panel who used agreed criteria (Kappa = 0.75). Moreover, participants in the nationwide survey were more likely to diagnose melanoma and less likely to diagnose benign naevi without atypia than the panel. In the second circulation, when diagnostic criteria and diagrams were used, there was a higher level of agreement for overall diagnosis using the categories benign, melanocytic intraepidermal neoplasia (MIN) with or without microinvasion and melanoma with vertical growth phase, and was the same as that achieved by the panel using the same criteria (Kappa = 0.68). CONCLUSIONS: As the incidence rate of thin melanomas has been increasing in the UK, it is important that standardised diagnostic criteria are used to ensure accurate reporting of incidence and correct management of patients. The use of MIN and the vertical growth phase seemed to be generally acceptable.     

Specific chromosomal defects associated with metastatic potential in K-1735 melanoma clones. Involvement of chromosomes 4 and 14.

In this study we sought to identify specific cytogenetic defects associated with the metastatic phenotypes in clones isolated from the parental K-1735 murine melanoma. All nonmetastatic clones (C-3, C-10, and C-19) exhibited trisomy of chromosomes 1, 3, 12, and 15. The only structural defect present in these clones was an interstitial deletion in a chromosome 4. In contrast, the highly metastatic clones (C-4, M-2, BB1, and X-21) exhibited trisomy of chromosomes 1, 3, 12, and 15, plus structural abnormalities of chromosomes 4 and 14, with the net result of a deletion in both. Parental K-1735 cells and clone C-16 cells, which are intermediate in their metastatic potential, had some cells with 4 and 14 alterations and others with only a deletion of chromosome 4. Clone C-16 revealed other non-clonal structural abnormalities. Our results indicate that structural anomaly of chromosome 4 and numerical alterations of certain autosomes may be associated with tumorigenic properties. In addition, structural defect in chromosome 14 is associated with high metastatic potential of K-1735 melanoma cells.     

Thick malignant melanomas (> 3 mm Breslow) with good clinical outcome: a histological study and survival analysis

Thick malignant melanomas in general tend to have a poor prognosis, but exceptions occur where there may be no further recurrence. The reasons for this difference in clinical behaviour are not fully understood. We have looked at thick malignant melanomas (greater than 3.0 mm) in the East of Scotland that have no evidence of metastasis after a minimum of 6 years follow-up and compared the clinical and histological features with a similar group, associated with histological evidence of metastasis and/or death. Both groups received similar treatment regimes. We have identified 41 patients with thick melanomas in the former group. When compared with the control group, factors found to be significantly different between the two groups were: the nature of the lower margin of the tumour; vascular invasion; and anatomical location.     

Subacute cutaneous lupus erythematosus—the annular variant. A histological and ultrastructural study of five cases

Subacute cutaneous lupus erythematosus is an uncommon, non-scarring variant of lupus erythematosus which has received scant attention in the pathology literature. Its recognition as a distinct entity is important, as, although mild systemic features may be associated, there is characteristically a relative absence of life-threatening renal or central nervous systemic involvement. Subacute cutaneous lupus erythematosus may be clinically sub-divided into the more common annular and the rarer papulosquamous (psoriasiform) variants. A clinicopathological study of five cases of the annular variant is presented with a review of the literature. We highlight the histological and ultrastructural appearances. Discriminatory features from the other variants of lupus erythematosus and additional conditions with which it may be confused, including lichen planus and acute graft-versus-host disease, are discussed.     

The prognostic significance of tumor vascularity in intermediate-thickness (0.76-4.0 mm thick) skin melanoma. A quantitative histologic study.

The vascularity of 20 primary skin melanomas was assessed histologically. These cases were selected from patients with intermediate thickness melanomas (0.76-4.0 mm thick) treated surgically to provide two groups of ten patients. One group had no evidence of recurrence with a minimum follow-up of 9 years. The second group of ten patients developed locoregional or systemic metastasis under follow-up, and seven of these patients died of disseminated melanoma. Age, sex, Breslow's tumor thickness, and Clark's level of invasion were similar in the two groups. Vascular quantitation was carried out by image analysis after vascular definition by Ulex europaeus-I agglutinin staining. The percentage vascular area at the tumor base in the recurrence group was more than twice that in the recurrence-free group. This study suggests that increased vascularity at the tumor base may have prognostic significance in intermediate thickness melanomas.     

Extent of vascularization as a prognostic indicator in thin (< 0.76 mm) malignant melanomas.

Angiogenesis in malignant melanoma (MM) was evaluated by comparing mean vessel number (MVN) in Spitz's nevi (SN), thick and thin MMs that metastasized, and thick and thin MMs with > or = 10-year survival. Vessels were identified with antibodies against factor VIII-related antigen (FVIII) and CD34 in 37 MMs (17 < or = 1.9 mm and 20 > or = 4.0 mm) with > or = 10-year follow-up and 10 SN from children (< or = 9 years old). Fields (x250) with the highest vessel density were counted by independent observers blinded to clinical outcome. There were no differences in MVN between SN versus MMs (P = 1.0), but the distribution of vessels was much more uniform in SN. Seven MM pairs (> or = 5.5 mm) and five pairs (< or = 0.75 mm) were matched by sex, age, site, stage, and primary treatment (paired t-test). In the pairs > or = 5.5 mm, there was no correlation with MVN with either metastasis or death (FVIII P = 0.98; CD34 P = 0.85). Among the thin paired lesions, high MVN (FVIII = 46, CD34 = 39) was significantly related not only to metastasis (FVIII P = 0.04, CD34 P = 0.03) but also to death (FVIII P = 0.04, CD34 P = 0.05). MVN does not separate SN versus MM nor predict outcome in thick (> or = 4.0 mm) MMs; however, high MVN (> or = 42 average) is predictive of metastasis and death in MMs < or = 0.75 mm. Larger matched studies are indicated to confirm this observation.     

<Emphasis Type="Italic">In vitro</Emphasis> assays for determining the metastatic potential of melanoma cell lines with characterized <Emphasis Type="Italic">in vivo</Emphasis> invasiveness

The metastatic potential of cancer cells is an elusive property that is indicative of the later stages of cancer progression. The ability to distinguish between poorly and highly metastatic cells is invaluable for understanding the basic biology of cancer and to develop more treatments. In this paper, we exploit a A375 melanoma cell line series (A375P, A375MA1, A375MA2) that vary in metastatic potential, to demonstrate an in vitro screening assay using polydimethylsiloxane (PDMS) microbubble well arrays that can distinguish these cell lines by their growth characteristics in including morphology, migratory potential, and clonogenic potential. These cell lines cannot be distinguished by their growth characteristics when cultured on standard tissue culture plastic or planar PDMS. Results show that the more metastatic cell lines (A375MA1, A375MA2) have a higher proliferative potential and a distinctive radial spreading growth pattern out of the microbubble well. The A375MA2 cell line also has a higher tendency to form multicellular spheroids. The ability to successfully correlate the metastatic potential of cancer cells with their growth characteristics is essential first step toward developing a high-throughput screening assay to identify aggressive tumor cells in primary samples. The capability to culture and recover aggressive cells from microbubble wells will enable identification of candidate metastatic biomarkers which has immense clinical significance.     

Expression of gp100, MART-1, tyrosinase, and S100 in paraffin-embedded primary melanomas and locoregional, lymph node, and visceral metastases: implications for diagnosis and immunotherapy. A study conducted by the EORTC Melanoma Cooperative Group

With the recent availability of novel antibodies against melanoma antigens tyrosinase and MART-1, it is important to validate their usefulness in pathology practice and in screening patients for immunotherapy treatment. In the present study conducted by the Melanoma Cooperative Group of the European Organization for Research and Treatment of Cancer (EORTC-MCG), immunohistochemical staining for gp100 (antibodies NKI-beteb and HMB-45), MART-1 (A103), tyrosinase (T311), and S100 (S100) was compared on formalin-fixed and paraffin-embedded tumour lesions from 80 patients with 130 malignant melanoma lesions, comprising 44 primary tumours, 18 locoregional metastases, 41 lymph node metastases, and 27 visceral metastases from the lung, liver, and brain. A score between 0 and 5 was allocated to each immunohistochemically stained section. These scores were evaluated in a statistical analysis. S100 was by far the most sensitive marker in all four types of lesions tested. Apart from a significantly better performance for T311 in primary melanomas compared with HMB-45, no significant differences were observed between the four remaining antigens tested. Three settings were next investigated to determine whether the expression of melanoma antigens decreases with tumour progression. First, within the primary melanomas, only NKI-beteb and A103 staining showed a nearly significant negative correlation with Clark's level of invasion and a similar tendency was observed for these antibodies with Breslow thickness. Second, when comparing primary melanoma-metastasis pairs from the same patient, lymph node metastases showed less staining with NKI-beteb, HMB-45, A103, and T311, at a level near significance. This difference was not significant when comparing the primary tumour with visceral metastases, probably due to the lower numbers of pairs. Third, regarding tumour progression from primary melanoma to locoregional, to lymph node, to visceral metastasis, a significant decrease with progression was found only for T311. The apparently stable expression of most of the melanoma antigens, and the small contribution of decreased expression in melanoma tumour progression, supports the rationale for immunotherapy based on the melanoma immunogens gp100, MART-1, and tyrosinase.     

Microelectrode study of spreading depression (SD) in frog retina--general observations of field potential associated with SD.

Using microelectrodes, the field potential change associated with SD (SDP) was recorded from frog retinas conditioned with Cl-free Ringer's. In such retinas, SDP was induced by light or chemical agents such as glutamate, aspartate and K+. The chemicals, when applied iontophoretically, produced a local graded response which eventually triggered SDP. A potential similar to the local response to chemicals was often discerned on the rising phase of SDPs produced by light or occurring spontaneously. The SDP was maximal across an innermost retinal layer 50 mum or less in thickness with the intraretinal polarity predominantly negative, indicating that the major sink of SDP is in the inner plexiform layer. The influence of SDP on the receptor potential was relatively small, but the other components completely disappeared at the beginning of SDP, recovering gradually thereafter. Concomitantly, a strong depolarization occurred in the ganglion cells. Stimulation of the optic nerve could induce SDP, but nerve impulse activity is not important for SDP because tetrodotoxin was unable to prevent SDP due to light or chemical agents.     

A retrospective histological study of 669 cases of primary cutaneous malignant melanoma in clinical stage I. The consequences of a reclassification of the original group of lentigo maligna melanomas.

A selected series of primary malignant melanoma of the skin, clinical stage I, was originally classified according to Clark's system. The consistency of this classification was tested by two Brisbane pathologists who indicated that we had misinterpreted some cases of superficial spreading malignant melanoma as lentigo maligna melanoma. We have therefore reclassified the original group of 86 lentigo maligna melanomas. This resulted in a total series of 37 (5.5%) lentigo maligna melanomas, 301 (45%) superficial spreading malignant melanomas, 194 (29%) nodular malignant melanomas (unchanged) and 137 (20.5%) unclassifiable malignant melanomas. The diagnosis of lentigo maligna melanoma was not made unless the epidermis was atrophic and dermal solar elastosis was present. The new group of lentigo maligna melanomas is dominated by cases on the head among patients over 50 years of age (especially women). This is in better agreement with other studies than our previous findings. The relationship with tumour cell type, pigmentation, mitotic count, atypia, transsectional profile, level of invasion, ulceration, vascular invasion, lymphocyte infiltration and prognosis shown by the new groups of lentigo maligna melanoma and superficial spreading malignant melanoma indicates that the cases by which the diagnosis has been changed are relatively benign. Our previous conclusions are still valid. The lentigo maligna melanoma is still the most benign type and nodular malignant melanoma still the most malignant type of melanoma. The superficial spreading malignant melanoma still represents an intermediate tumour type, although it has deviated in the benign direction.     

Constitutive expression of growth regulated oncogene (<Emphasis Type="Italic">gro</Emphasis>) in human colon carcinoma cells with different metastatic potential and its role in regulating their metastatic phenotype

The purpose of this study was to examine the expression and functional significance of the growth-regulated oncogene (gro) family in human colon carcinoma growth and metastasis. We examined constitutive expression of CXCL1 (gro-α), CXCL2 (gro-β), CXCL3 (gro-γ) and their receptor, CXCR2 in human colon carcinoma cells with different metastatic potentials. Non-metastatic and low metastatic cells expressed lower levels of CXCL1 and CXCR2 mRNA and protein as compared to high metastatic colon carcinoma cells. No difference in CXCL2 and CXCL3 mRNA expression levels was observed. Colon carcinoma cells expressing higher levels of CXCL1 exhibit increased proliferation and invasive potential. Furthermore, exogenous addition of recombinant human CXCL1 significantly enhanced the proliferation and invasiveness of colon carcinoma cells. Furthermore, treatment of KM12C cells with exogenous CXCL1 enhanced their invasiveness. Neutralizing antibody to CXCL1 in combination with antibody to CXCR2 inhibited highly metastatic KM12L4 (high CXCL1 expressor) cell proliferation. These data demonstrate that the constitutive expression of CXCL1 and its receptor CXCR2 is associated with metastatic potential and modulates colon cancer cell proliferation and an invasive phenotype.This revised version was published online in August 2005 with a corrected cover date.