Epidemiology of invasive cutaneous melanoma.

Data are presented on the current incidence of melanoma with recent and predicted future trends illustrating a likely continuing increase in incidence. Risk factors for developing melanoma are discussed, including current known melanoma susceptibility genes. Phenotypic markers of high-risk subjects include high counts of benign melanocytic naevi. Other risk factors considered include exposure to natural and artificial ultraviolet radiation, the effect of female sex hormones, socioeconomic status, occupation, exposure to pesticides and ingestion of therapeutic drugs including immunosuppressives and non-steroidal anti-inflammatory drugs. Aids to earlier diagnosis are considered, including public education, screening and use of equipment such as the dermatoscope. Finally, the current pattern of survival and mortality is described.

Utility of adjuvant systemic therapy in melanoma.

The lack of effective drugs in stage IV melanoma has impacted the effectiveness of adjuvant therapies in stage II/III disease. To date, chemotherapy, immunostimulants and vaccines have been used with minimal success. Interferon (IFN) has shown an effect on relapse-free survival (RFS) in several clinical trials; however, without a clinically significant effect on overall survival (OS). A recently conducted meta-analysis demonstrated prolongation of disease-free survival (DFS) in 7% and OS benefit in 3% of IFN-treated patients when compared with observation-only patients. There were no clear differences for the dose and duration of treatment observed. Observation is still an appropriate control arm in adjuvant clinical trials. Regional differences exist in Europe in the adjuvant use of IFN. In Northwest Europe, IFN is infrequently prescribed. In Central and Mediterranean Europe, dermatologists commonly prescribe low-dose IFN therapy for AJCC stage II and III disease. High-dose IFN regimens are not commonly used. The population of patients that may benefit from IFN needs to be further characterised, potentially by finding biomarkers that can predict response. Such studies are ongoing.

An evidence base for reconsidering current follow-up guidelines for patients with cutaneous melanoma less than 0.5 mm thick at diagnosis.

BACKGROUND: Despite current guidelines, there is uncertainty about the required duration and frequency of follow-up visits for patients with invasive primary cutaneous melanoma < 0.5 mm thick. OBJECTIVES: To review patients with invasive melanoma thinner than 0.5 mm followed for at least 5 years to provide an evidence base for considering modification of guidelines. METHODS: A retrospective review of 430 patients diagnosed in the west of Scotland during 1992-2001 with melanoma < 0.5 mm was carried out. Recurrence, deaths from melanoma and second primary melanomas were all identified. RESULTS: From 1992 to 2001, 430 melanomas < 0.5 mm thick at diagnosis were diagnosed out of a total of 3036 primary cutaneous melanomas. To date there have been 593 deaths from melanoma (19%) in the whole group. Five of these deaths were reported in patients with melanomas < 0.5 mm, but on pathological review two were thicker than 0.5 mm at diagnosis (1.5 and > 3 mm), and the remaining three patients all developed thicker second primary melanomas (2.7, 12.0 and 19.0 mm) with a recurrence pattern and timing indicating that these thicker primaries were the cause of death. Fourteen further patients developed a second primary melanoma, and 13 are currently alive and disease free, one dying of other causes. CONCLUSIONS: Our data indicate that recurrence and subsequent death from melanomas < 0.5 mm is a very rare event, and that quarterly follow-up for 3 years will yield very few events. Modification of current guideline recommendations are suggested to include a period of patient education concentrating on recognition of second primary tumours followed by rapid access to an expert opinion if required.

Quantitation of dendritic cells in normal and abnormal human epidermis using monoclonal antibodies directed against Ia and HTA antigens.

Two monoclonal antibodies, NAI 34 and DA6 231, have been used as surface markers to count epidermal Langerhans cells in normal skin, allergic contact dermatitis, and mycosis fungoides of the plaque and poikilodermatous varieties. The antibodies recognized the HTA (human thymocyte antigen) and Ia antigens, respectively. In all situations the numbers of cells labeled by the two antibodies differed. In normal skin 75% of dendritic cells were labeled by both antibodies and 25% were labeled by NAI 34 alone. In contact dermatitis there is an increase in dendritic cells labeled by both antibodies and this pattern is also seen in untreated plaque stage mycosis fungoides. In poikilodermatous mycosis fungoides, striking staining of the epidermal keratinocytes with DA6 231 is seen with no dendritic cells labeled. Dendritic cells are, however, clearly seen with NAI 34 staining. Numbers of dendritic cells labeled by both antibodies fall during PUVA therapy.

CDKN2A germline mutations in U.K. patients with familial melanoma and multiple primary melanomas.

We report six of 16 U.K. melanoma families and two of 17 patients with multiple primary melanomas and a negative family history who have between them four different functionally damaging mutations of the CDKN2A (p16) gene: an Arg 24 Pro substitution in exon 1 in one family, a stop codon at codon 44 of exon 1 in one family, and a Met 53 Ile substitution in exon 2 in four families. One multiple primary melanoma patient also has the Met 53 Ile mutation and a second has a G-T substitution at the IVS2 + 1 splice donor site. Our data together with other recent publications from France and the U.S.A. indicate that screening melanoma kindreds with only two affected family members for CDKN2A mutations is justified.

The phototumorigenic potential of broad-band (270-350 nm) and narrow-band (311-313 nm) phototherapy sources cannot be predicted by their edematogenic potential in hairless mouse skin.

The new Philips TL01 narrow-band (311-313 nm) and conventional broad-band (e.g., Philips TL12; 270-350 nm) sources are effective for psoriasis phototherapy, for which treatment regimens are based on a predetermined minimal erythema dose. TL01 phototherapy treatment times are approximately half those with TL12 for psoriasis, whereas the cumulative exposure doses at clearing are similar. We compared the phototumorigenic potential of TL01 and TL12 radiation in mouse skin. Groups of albino Skh-1 hairless mice were exposed for 5 d/week at three dose levels. At each dose level, TL12 and TL01 doses were equally edematogenic. At each dose level, TL01 radiation was significantly more effective at producing first tumors of 1 mm in diameter and multiple tumors. At the lower two dose levels, TL01 radiation produced a significantly greater proportion of squamous cell carcinomas. This study demonstrates that TL01 radiation is more phototumorigenic than TL12 radiation at equally edematogenically weighted doses. This is in contrast with previous reports that edema production by polychromatic sources is predictive of their phototumorigenic effect in Skh-1 mice. The absolute cumulative TL12 dose needed to induce tumors was much less than that for TL01 radiation. The possibility of increased tumor risk with TL01 phototherapy should be considered but must be balanced against the high phototherapeutic efficacy of this source, short treatment times, and the low cumulative doses necessary for clearing of psoriasis.

Localization of elevated transforming growth factor-alpha in psoriatic epidermis.

Biopsies of involved and uninvolved skin from five patients with plaque psoriasis and normal skin from four healthy volunteers were investigated for steady-state quantities of TGF-alpha RNA and protein by in situ hybridization and immunohistochemistry. Increased levels of TGF-alpha RNA were found only in the high-level keratinocytes of involved psoriatic skin (p less than 0.001). Elevated levels of TGF-alpha protein were seen in both the high-level and basal layers of involved psoriatic skin compared to uninvolved psoriatic and normal control skin. Elevated TGF-alpha gene expression is thus implicated in the hyperproliferation of keratinocytes and possibly the altered maturation pattern seen in psoriasis.

Retinoic acid-induced inhibition of metastatic melanoma cell lung colonization and adhesion to endothelium and subendothelial extracellular matrix.

The effect of pretreatment of metastatic B16 melanoma cells with 10(-6) M all trans-retinoic acid resulted in a significant inhibition of lung colonization following injection of 10(5) cells into the tail vein of syngeneic C57BL mice. Adhesion of melanoma cells to vascular endothelial cell monolayers, and subendothelial extracellular matrix was also inhibited by pretreatment with retinoic acid, as was tumour cell aggregation following seeding of pretreated cells on to 0.5% agar. Release of 35SO4 from radiolabelled subendothelial extracellular matrix by melanoma cells was essentially unaltered by retinoic acid pretreatment, as was the release of radiolabel from [3H]proline-labelled matrix, while plasminogen activator activity was enhanced in retinoic-acid-treated cells. These observed changes in adhesive properties may be responsible, at least in part, for the retinoic-acid-induced inhibition of lung colonization.

Incidence, risk factors and prevention of melanoma.

The incidence of cutaneous malignant melanoma steadily increased between 1940 and 1990 in both sexes. However, this increase appears to have peaked in females in Scotland. Excessive exposure to ultraviolet radiation among Caucasians is the main etiologic factor implicated in the incidence of melanoma. Mortality due to melanoma also increased between 1940 and 1990, but the rate of increase is less than that of the incidence of melanoma. This may be due to earlier diagnosis and treatment of melanoma as a result of public education campaigns. Independent risk factors for developing melanoma include the presence of benign melanocytic naevi (moles), the development of lentigines or freckles, three or more dysplastic naevi and a history of three or more severe sunburns that resulted in peeling or blistering. Approximately 2% of melanoma patients have a family history of the disease and research into potential melanoma susceptibility genes is ongoing. Primary prevention campaigns, initiated mainly in Australia, are aimed at encouraging sensible sun exposure. Secondary prevention campaigns are directed at preventing death from melanoma by encouraging early diagnosis and treatment. Additional prospective studies are needed to determine if the incidence of melanoma has peaked and whether the recent trends observed in females will also occur in males.

Melanoma cell-derived factor stimulation of fibroblast glycosaminoglycan synthesis--the role of platelet-derived growth factor.

The hyaluronan-rich matrix surrounding many tumours may facilitate tumour growth, invasion and angiogenesis, with the majority of this hyaluronan apparently being synthesised by normal fibroblasts, stimulated to do so by tumour cell-derived factors. Melanoma cell-conditioned medium (CM) stimulates up to a 6-fold increase in fibroblast glycosaminoglycan (GAG) synthesis, with the active factors being present in tumour CM ultrafiltration fractions > 30 kDa and < 1 kDa. These fractions are poorly active individually, but when recombined, the activity is substantially greater than the additive effect. The objective of this study was to identify the factors present in the ultrafiltration fraction > 30 kDa that produce a greater than additive effect with the fraction < 1 kDa in stimulating the incorporation of 3H glucosamine into fibroblast GAGs. A number of factors including basic fibroblast growth factor (bFGF), interleukin (IL)-1 beta, pleiotrophin, platelet-derived growth factor (PDGF), transforming growth factor-beta (TGF-beta), tumour necrosis factor-alpha (TNF-alpha) and vascular endothelial growth factor (VEGF) failed to stimulate any significant increase in GAG synthesis, but when added to the < 1 kDa tumour CM fraction, both PDGF and to a lesser extent, bFGF, exhibited potent stimulating activities. Neutralising antibodies to PDGF and bFGF added to the melanoma CM decreased the fibroblast GAG-stimulating activity by 29% and 40%, respectively, in C8161 melanoma CM and by 47% and 45%, respectively, in Hs294T melanoma CM. The activities of PDGF-AA and PDGF-BB isoforms were indistinguishable, suggesting the PDGF-alpha receptor plays a role in the GAG-stimulatory response. Western analysis following treatment with PDGF, bFGF or melanoma CM revealed banding patterns for PDGF and melanoma CM that were similar. Immunoprecipitation of the PDGF-alpha receptor revealed it to be phosphorylated in fibroblasts treated with PDGF and melanoma CM, but not with control fibroblast CM. These studies suggest that PDGF plays an important role in the GAG-stimulating activity of the melanoma CM, but requires the presence of an as yet unidentified novel low molecular weight factor for full activity.

Stage II melanoma in the west of Scotland, 1976-1985: prognostic factors for survival.

The outcome of 142 patients undergoing therapeutic lymphadenectomy for clinical stage II malignant melanoma was retrospectively assessed. 5 year survival was 26%, and survival was not altered in the 25 patients who received two courses of adjuvant combination chemotherapy after lymphadenectomy. On univariate analysis, the most significant determinants of survival were the number of malignant nodes removed at lymphadenectomy (P = 0.00004), the age of the patient (P = 0.009) and the disease-free interval between primary and stage II disease (P = 0.01). The following features were not significantly related to survival: sex, site, histogenetic type of primary tumour, tumour thickness and level of invasion. The number of malignant lymph-nodes was confirmed on multivariate analysis as the single most useful and significant predictor of survival, with the patient's age providing an additional significant contribution. In future adjuvant trials in stage II melanoma after therapeutic lymphadenectomy, patients should be stratified for both age and number of malignant nodes.

A comparison of dosimetric methods in isolated limb perfusion with melphalan for malignant melanoma of the lower extremity.

The three dosimetric schedules currently used in isolated limb perfusion with melphalan for malignant melanoma of the lower limb were compared in a series of 51 patients. The doses prescribed by each of the three methods (based on total body weight (TBW), limb tissue volume (LTV) and total blood volume in the perfusion circuit (TBV)) were calculated for all patients and were then compared using Wilcoxon's signed-rank test. This revealed that the method based on TBV consistently prescribed much lower doses of drug than either of the other two methods. Pharmacokinetic profiles of melphalan obtained by HPLC analysis of blood samples during the procedure also showed that the method did not reliably predict the concentration of melphalan achieved in the perfused limb. The dosimetric schedule based on LTV prescribed slightly higher doses than that based on TBW. However, the technique is more difficult to practise due to the problems of measuring the limb volume by immersion. We conclude that the dosimetric schedule based on TBW is the most appropriate by virtue of its simplicity, the high doses of melphalan which it prescribes, and the well-controlled toxicity which it produces.

A single centre's 10 year experience with isolated limb perfusion in the treatment of recurrent malignant melanoma of the limb.

The aim of this study was to assess whether isolated limb perfusion can be performed safely and whether it offers improved disease-free survival for patients with limb malignant melanoma. Between August 1983 and July 1993, 103 patients (78 female, 25 male) with recurrent limb melanoma were treated by isolated limb perfusion (ILP) in Glasgow, U.K. The mean age of the patients was 62 years; 95 had leg recurrence, 8 had arm recurrence. The mean time from original diagnosis to ILP was 48 months (range 1-290). 102 iliac, 5 femoral, 7 popliteal and 8 axillary perfusions were performed. All patients had stage II (local recurrence within 3 cm of primary site) or stage III (regional metastases; tissues excluding nodes, nodes or combination) disease according to the MD Anderson Cancer Centre Staging System. At a mean follow-up of 30.7 months, 68 patients had died of recurrent disease (mean time to death 22.5 months). The 2 and 5 year survival of the group was 50 and 26%, respectively and disease-free survival was 23 and 12%, respectively. At first perfusion, 76% of patients showed complete response and 23% showed partial response. With repeat perfusion, 47% showed complete response and 53% had partial response. In conclusion, ILP is safe and has an acceptable morbidity. It achieves highly satisfactory local disease control but long-term survival is the exception.

Melphalan concentration and distribution in the tissues of tumour-bearing limbs treated by isolated limb perfusion.

Levels of melphalan (L-phenylalanine mustard) were measured in the tissues of tumour-bearing limbs treated by isolated limb perfusion (ILP). 41 samples of melanoma tissue, normal fat and skin were excised from 15 patients during ILP. A high performance liquid chromatography assay was used to measure melphalan concentrations. Levels of melphalan were higher in tumour than in fat (P < 0.01, Wilcoxon signed-ranks test), and not significantly different from levels in adjacent skin. In 2 cases there was significant regional toxicity in the treated limb, but this was not related to the levels of melphalan measured in the tissues of the limb. It is encouraging that the concentrations of melphalan which were achieved in large necrotic nodules by ILP were similar to those in well-perfused normal skin.

Validity of the histopathological criteria used for diagnosing dysplastic naevi. An interobserver study by the pathology subgroup of the EORTC Malignant Melanoma Cooperative Group.

Ten (dermato)pathologists studied 50 cutaneous melanocytic lesions including common naevocellular naevi, dysplastic naevi (DN), melanomas in situ and invasive primary melanomas, with emphasis on the histological criteria of DN. Using a standardised form, 20 defined histopathological features were scored (semi)quantitatively. Concordance of diagnosis, efficacy and reproducibility of features were investigated. DN were distinguished well from the other entities (mean Po 0.87). Agreement on the degree of atypia of DN was low. The reproducibility of the scoring was best for the following features: irregular nests, lymphohistiocytic infiltrate, marked junctional proliferation and large nuclei. The overall values of these features to discriminate between DN and non-DN were better than for the other features studied. Using the presence of at least three of the four features as a condition for the diagnosis of DN, values for sensitivity, specificity and positive and negative predictive values were 0.86, 0.91, 0.96 and 0.73, respectively. On the basis of the results these features seem best suited as histological criteria for the diagnosis of DN.

Histopathologic diagnosis of dysplastic nevi: concordance among pathologists convened by the World Health Organization Melanoma Programme.

Dysplastic nevi are an important indicator of risk of cutaneous malignant melanoma. The study of and, particularly, international communication regarding this group of lesions have been hindered by a lack of precision in diagnosis. In an effort to broaden understanding, a panel of pathologists agreed upon a set of criteria for the diagnosis of dysplastic melanocytic nevi. Two major and four minor criteria were defined. The major criteria are (1) basilar proliferation of atypical nevomelanocytes (extending at least three rete ridges or "pegs" beyond any dermal nevo-cellular component), and (2) organization of this proliferation in a lentiginous or epithelioid-cell pattern. Minor criteria are (1) the presence of lamellar fibrosis or concentric eosinophilic fibrosis, (2) neovascularization, (3) inflammatory response, and (4) fusion of rete ridges. Diagnosis required presence of both major criteria and at least two minor criteria. One hundred fourteen histologic specimens of benign acquired nevi, dysplastic nevi, and radial-growth-phase melanomas were examined by the members of this panel; their diagnoses were compared to determine degree of concordance. The established criteria yielded 92% mean concordance overall.

Comparative study of three methods of plastic embedding in diagnostic dermatopathology.

Three methods of plastic embedding were assessed for their value in routine dermatopathology. The J B 4, Taab embedding, and Taab transmit techniques were compared for quality, convenience, and safety. The Taab transmit method was the most satisfactory method, being both rapid and simple. An additional advantage is that the resin can be removed and the immunoperoxidase technique carried out on thin sections.

Use of NK1 C3 monoclonal antibody in the assessment of benign and malignant melanocytic lesions.

The monoclonal antibody NK1 C3, synthesised by the Netherlands Cancer Institute, has been used to assess its value in the diagnosis of melanocytic lesions. The antigen recognised by this antibody is not denatured by formalin fixation, with the result that the antibody can be used for retrospective studies on conventionally processed material. Positive results were obtained in primary melanoma (18/18), secondary melanoma (21/21), junctional and compound naevi (32/32), intradermal naevi (9/12), congenital naevi (3/3), so called dysplastic naevi (13/13), blue naevi (5/5), and Spitz tumours (3/14). Non-melanocytic tumours were tested for comparison. The results showed relative but not complete specificity of the antibody for melanocytic tumours, with positive results only in breast and prostate tumours (2/6 and 2/5 respectively). Negative results were obtained with basal and squamous cell carcinoma, appendage tumours, neural tumours, and apudomas. The staining pattern of NK1 C3 was compared with that of antibodies to S100 protein and to neurone specific enolase. Compared with S100 protein NK1 C3 gave stronger staining of a higher percentage of cells in the 12 specimens in which a direct comparison was made. Antibody raised against neurone specific enolase in sheep gave very poor results with heavy background staining. We suggest that NK1 C3 is a useful addition to the battery of monoclonal antibodies of value to the diagnostic histopathologist.

Proposed alternative terminology and subclassification of so called "dysplastic naevi".

The term "dysplastic" melanocytic naevus has recently been used to describe pigmented naevi with unusual histological and clinical features. There is currently no clear clinical or pathological definition of the term, and this has led to a lack of comparability of material described in reports on these lesions. As a result of careful histological study and a clinicopathological correlation of 100 naevi, we suggest that three distinct groups of histopathological features distinguish so called dysplastic naevi from banal melanocytic naevi. These are architectural atypia, cytological atypia, and a host response. Description of each of these features in routine reports and in published series in place of the loose use of the term "dysplastic" would enable comparisons to be made between series of melanocytic lesions reported from different centres. In the course of this study we observed a considerably increased incidence of naevus type giant cells in the dermal portion of the atypical naevi. These giant cells should not be confused with possibly premalignant cytological atypia.