Topical dinitrochlorobenzene combined with systemic dacarbazine in the treatment of recurrent melanoma.

Fifty-nine selected patients with recurrent melanoma (37 locoregional and 22 locoregional and haematogenic) were treated with a combination of topical dinitrochlorobenzene (DNCB) and systemic dacarbazine (DTIC). The observations were analysed retrospectively. Fifteen patients (25%) (one third with distant spread) experienced a complete response, with a median duration of 10 months (range 3-210 months). Five of these patients, three of them with haematogenic disease, showed prolonged disease-free survival (85, 156, 170, 177 and 210 months). Seven patients (12%) obtained a partial response, and no response was seen in the remaining 37 patients (63%). In the presence of a severe local reaction to topical DNCB application the median survival was 12 months compared with 7 months in cases with a slight or moderate local reaction (P=0.0013). Patients showing a response to the DNCB-DTIC combination had a median survival of 18 months versus 7 months for the non-responders (P<0.0003). In view of the prolonged survival in five patients, this combined treatment schedule yields promising results in the treatment of selected melanoma patients with cutaneous recurrences, whether or not these are associated with other metastases.

Prognostic value of tuberculin and BCG immunoreactivity in stage I high-risk malignant melanoma (EORTC protocol 18781).

BACKGROUND: Since the immune system is considered to be a major determinant in the outcome of malignant melanoma, vaccination with BCG (bacillus Calmette-Guérin) or more recently with interleukins is used in this condition, although the effects of this immunotherapy are unclear. OBJECTIVE: The present study was to elucidate clinical aspects and the prognostic relevance of the cellular and humoral immune responses of melanoma patients treated with two different BCG vaccines. METHODS: A subgroup of patients from a multicenter adjuvant trial with BCG in stage I (pT3-4N0M0) high-risk melanoma was prospectively subjected to detailed analysis regarding tuberculin (PPD, purified protein derivate) skin test reactivity, local, regional and systemic reactions to BCG vaccination, PPD antibody response and disease-free survival. Patients were randomized into three arms and received either no adjuvant treatment (22 patients), BCG RIV (40 patients) or BCG Pasteur (44 patients). All patients were followed for up to 10 years (median follow-up 6 years). RESULTS: Patients treated with BCG Pasteur mounted a stronger antibody response, experienced stronger regional and systemic reactions to vaccination and converted more frequently to positive PPD skin tests, compared to controls and to patients vaccinated with BCG RIV. All BCG-treated patients who developed an antibody response had a longer disease-free interval (p = 0.05), with slightly higher significance for BCG Pasteur-treated patients (p = 0.02). CONCLUSIONS: Cellular as well as humoral immune responses to PPD and BCG thus identify stage I malignant melanoma patients with an overall better prognosis.

Effects of systemic interferon-alpha (IFN-alpha) on the antigenic phenotype of melanoma metastases. EORTC melanoma group cooperative study No. 18852.

In order to investigate the effects of in vivo treatment with interferon-alpha (IFN-alpha) on melanoma antigens, a clinical EORTC trial (No. 18852) was accompanied by an immunohistological study. Twenty patients with melanoma metastases of skin and soft tissues, eventually also of the lung, who were treated with systemic IFN-alpha, were evaluated for a comparison of metastases before (40) and during (42) treatment. Representative cryostat sections were studied immunohistologically with a panel of monoclonal antibodies against differentiation antigens (HMW-MAA, K-1-2, NKI-beteb, M-2-10-15), progression markers (transferrin receptor, ICAM-1, VLA-2), histocompatibility antigens (HLA-A, B, C, HLA-DR) and the proliferation-associated nuclear antigen Ki67. We found an overall reduction of the proliferation-associated antigen Ki-67 (p < 0.01), and an increase in expression of HLA-DR (p < 0.05) and ICAM-1 (trend) during treatment. The intensity of expression of HLA-A, B and C antigens as well as pigmentation (p < 0.01) was found to be increased. Early progression (< or = 8 weeks after onset of treatment) was associated with a lack of phenotypic changes. The data suggest an independent modulation of proliferation, pigmentation, and antigen expression by systemic treatment of metastatic melanoma with IFN-alpha.

Long-term adjuvant immunotherapy in stage I high risk malignant melanoma, comparing two BCG preparations versus non-treatment in a randomised multicentre study (EORTC Protocol 18781).

The present study reports the results of a multicentre adjuvant trial with BCG (Bacillus Calmette-Guérin) in high risk patients (Breslow thickness > or = 1.5 mm, Clark level > or = III) with malignant melanoma, after surgical removal of their primary tumour. The trial was specifically designed in order to resolve the controversy and to provide some definite answers regarding the value of adjuvant BCG treatment in stage I malignant melanoma. Patients were randomised to either BCG RIV (108 patients) or BCG Pasteur (109 patients) for 3 years or to follow-up only (110 patients). The two vaccines used had greatly divergent properties regarding their mode of preparation, their composition and their immunomodulating activities. Of the 353 randomised patients, 23 were ineligible, 3 refused participation after randomisation and 327 were evaluable for final analysis. Median follow-up time was 6 years (range 0-10 years). The log-rank test comparison showed no statistical difference between the three arms regarding time to progression (P = 0.55) and duration of survival (P = 0.82). Treatment was generally well tolerated, with no major adverse events in either treatment arm. These findings confirm data with different BCG preparations and with stage II melanoma which also demonstrated no benefit regarding patient survival and time to relapse.

Systemic therapy in disseminated melanoma.

There is no standard treatment for advanced melanoma. As long as metastases are satellites or in-transit metastases localized in a leg or arm, the prospects for curative treatment by isolation perfusion are good. But as soon as metastases have spread via the circulation, curative treatment with cytotoxic agents becomes virtually impossible. When the tumor burden is not too extensive, however, palliative treatment can be of clinical value. Some combinations of cytotoxic agents or combinations of biologic response modifiers have been shown to induce worthwhile remissions. Toxicity remains a problem, however. The advantages of the newer immunological approaches, especially with interleukin-2 (IL-2) and T-cell lymphocytes, is that treatment for a short period may result in good remissions at an early stage. Much clinical research is still needed to improve these costly approaches.

Tamoxifen as a single agent for advanced melanoma in postmenopausal women. A phase II study of the EORTC Malignant Melanoma Cooperative Group.

Between 1983 and 1989 a phase II study was carried out by the EORTC Malignant Melanoma Cooperative Group in which postmenopausal women with advanced melanoma but a good performance status received tamoxifen, 40 mg per day, as a single agent. From 12 centres, 114 patients were registered of whom 107 appeared to be eligible and 102 evaluable. Seven died of progressive disease within 4 weeks, eight others had early progressive disease (of whom seven died within 7 weeks). The response rate was 4.9%, the complete response rate 1%. Without prior chemotherapy three of 58 responded, with prior chemotherapy two of 44. Except for one of 46 patients with lung metastases who experienced a PR of these metastases, all responders were patients with slowly growing small soft tissue metastases. We conclude that, because of the few side effects, tamoxifen can be recommended for (postmenopausal) patients who have only a small number of slowly growing metastases and who are not yet candidates for treatment with toxic drugs.

Consensus on the management of melanoma of the skin in The Netherlands. Dutch Melanoma Working Party.

In 1990, the Dutch Melanoma Working Party, in cooperation with the National Organization for Quality Assurance in Hospitals, organised the second consensus conference on the management of melanoma of the skin. The following guidelines were approved: The margin of a therapeutical excision should be 1 cm for melanomas not thicker than 1.0 mm, 2 cm for a thickness of 1.1-2.0 mm, 3 cm for a thickness of 2.1-3.0 mm. No consensus was reached for tumours thicker than 3.0 mm. The conclusion of the histopathological report should state the histological type of melanoma, the thickness, the level of invasion, the presence of ulceration, regression, microsatellitosis and completeness of removal. In melanomas between 1.5 mm and 4 mm, elective lymph node dissection may be considered, but its value has not been proven. Clinically suspicious regional lymph nodes require a therapeutical lymph node dissection, solitary lymph node removal is inappropriate. Prophylactic (adjuvant) regional perfusion in primary melanoma should only be performed in the context of a clinical trial. Regional perfusion is the treatment of choice for satellitosis and/or in-transit metastases of the extremities without evidence of distant metastases. If radiotherapy is indicated, high fractionation doses are required. There is no standard therapy for distant metastases. Routine check radiographs and laboratory studies are unnecessary during the follow-up period. The follow-up period is normally 10 years.

Humoral immune response in disease-free advanced melanoma patients after vaccination with melanoma-associated gangliosides. EORTC Cooperative Melanoma Group.

Several studies have shown that melanoma-associated gangliosides are immunogenic in melanoma patients and that antibodies against them have a favorable prognostic effect. Our study aims at characterizing the humoral immune response in disease-free, advanced melanoma patients vaccinated with a total ganglioside fraction extracted from pooled metastases of human melanoma, containing as major gangliosides GM3 and GD3, and as minor ones GM2 and GD2. Prior to vaccination, all patients were made disease-free by surgical removal of skin, lymph-node or other distant metastases. Repeated vaccinations were carried out intradermally with gangliosides either in the native form in buffered solution, or in the form of liposomes. Serum samples were collected at regular intervals and assayed by ELISA for the presence of specific IgG and IgM antiganglioside antibodies. Selected samples were tested by immunostaining on thin-layer plates to specify the ganglioside species involved in the reactivity. Out of 32 evaluable patients, 17 presented a significant increase in antibody titer, mostly of the IgG isotype, which was maximal between 2 and 4 months after starting injections of gangliosides, and gradually disappeared within 1 year. No significant difference could be seen between the group of 20 patients treated with native gangliosides and the group of 12 vaccinated with the gangliosides in liposomes. All gangliosides seemed to be immunogenic, but GM2 and GD2 were somewhat more reactive. The disease-free intervals for the patients who showed an antibody response to the treatment were significantly higher (p less than 0001) than those of the non-responding group, as compared by the Kaplan-Meier method.

The EORTC Melanoma Group exchange program: evaluation of a multicenter monoclonal antibody study.

In the framework of the European Organisation for Research and Treatment of Cancer (EORTC), the Immunology and Pathology Subgroups of the Malignant Melanoma Cooperative Group undertook a large multicenter monoclonal antibody (MAb) study. Fourteen laboratories from 7 European countries tested a panel of 23 MAbs for immunohistological staining reactivity for malignant and non-malignant lesions involving the melanocytic lineage. A standardized immunoperoxidase procedure was used and the results were evaluated using a standard protocol and data evaluation form developed in collaboration with the EORTC Data Center. According to this analysis, the antibodies in the panel could be classified into 3 main groups. The first group of MAbs includes those antibodies which stained the majority (greater than 80%) of all primary tumors, irrespective of their Breslow thickness and the majority of metastatic lesions. In addition, these MAbs stained a high percentage of cells within a given lesion. Several antibodies of Group I were likewise reactive with the majority of naevoblasts and with normal melanocytes. The second group of MAbs included antibodies reacting only with a limited number of primary melanomas and metastatic lesions. Antibodies of Group II reacted only weakly, if at all, with normal melanocytes or naevocytes. The percentage of cells within a malignant lesion stained by these MAbs was always rather low. The MAb group III detected surface structures whose expression appeared to be related to tumor progression; they did not react or reacted only weakly with naevi, and they all reacted with a small number of early primary melanomas (less than 0.75 mm). The number of lesions stained increased with increasing Breslow thickness. Our study suggests that the application of a panel of well defined MAbs might be of diagnostic and prognostic value in evaluating malignant melanoma.

Initial experience with treatment of human B cell lymphoma with anti-CD19 monoclonal antibody.

Six patients with progressive B cell non-Hodgkin's lymphoma have been treated with an IgG2a mouse monoclonal antibody (mAb) against the B cell differentiation antigen CD19, with total doses varying from 225 mg to 1000 mg. Free mAb was detected in the serum after doses of 15-30 mg. After the mAb infusions the number of circulating tumour cells was temporarily reduced, but in some cases antibody-coated cells remained in the circulation for several days. mAb penetrated to extravascular tumour sites; in general higher doses were required to saturate cells in the lymph nodes than to sensitize tumour cells in the bone marrow. mAb doses of up to 250 mg were given i.v. over 4 h without major toxicity. One patient twice achieved a partial remission after two periods of mAb treatment with an 8-month interval; the second remission lasted for 9 months. One patient showed a minor response. None of the patients made antibodies against the mouse immunoglobulin. Serum immunoglobulin levels were followed as a measure of the function of the normal B cell compartment; no significant changes were seen up to 6 months after mAb treatment.

[Revised consensus melanoma of the skin]. / Herziene consensus melanoom van de huid.

A second consensus meeting on cutaneous melanoma was held on June 15, 1990. The present report concisely discusses the recommended policy concerning the diagnosis and treatment of melanoma. The changes in melanoma management in comparison with the first consensus meeting in 1984 are emphasized.

Lymph node metastasis from melanoma with an unknown primary site.

Twenty-six patients, treated surgically between 1961 and 1986 because of lymph node metastasis from melanoma with an unknown primary, were analysed. Six patients had a history of spontaneous regression of a skin lesion. Following node dissection, the overall actuarial disease-free survival rate was 49 per cent, after both 5 and 10 years. When considered as single factors, female (versus male), one lymph node involved (versus more than one node involved) and site of metastasis in the groin or axilla (versus the neck) were found to have significantly favourable effects on prognosis with 5-year survival rates of 82 per cent (25 per cent), 82 per cent (27 per cent) and 80 per cent (11 per cent) respectively. However, at multifactorial analysis only the site of cervical metastases maintained a significant influence on survival (P = 0.005). As survival in this series is comparable with, or even better than, that of adequately treated patients with lymph node metastasis from a known primary melanoma, a radical node dissection is essential also in these patients.

Aerosol application of interferon-alpha in the treatment of bronchioloalveolar carcinoma.

10 patients with locally advanced bronchioloalveolar carcinoma were treated with interferon-alpha as an inhaled aerosol. Initial doses ranged between 1 and 10 MU daily or thrice weekly and were then increased to 20 MU daily. Treatment was continued until disease progression or excessive toxicity occurred, 9 patients were evaluable for toxicity. In 1 case treatment had to be stopped after 2 weeks due to fever, fatigue and progressive dyspnoea. 2 patients developed fever, 1 had malaise, fatigue and loss of appetite and 2 had dose-dependent transient dyspnoea. According to standard criteria no tumour responses could be detected. In 6 out of 8 evaluated for response to interferon, radiological stabilisation of disease for 7-43 weeks (median 15) was observed. These results point to the feasibility of aerosol inhalation of interferon-alpha, but also to its limited antitumour activity in locally advanced bronchioloalveolar carcinoma.

Potentiation by interleukin 2 of Burkitt's lymphoma therapy with anti-pan B (anti-CD19) monoclonal antibodies in a mouse xenotransplantation model.

To study the immunotherapeutic potential of monoclonal antibodies (mAbs) directed against the human pan-B-cell antigen CD19, a xenotransplantation model was developed in which the human Burkitt's cell line Daudi is s.c. transplanted into nude mice. IgG1, IgG2b, and IgG2a isotype variants of the anti-CD19 mAb (CLB-CD19) were tested for their capacity to inhibit the growth of 10 x 10(6) Daudi cells injected s.c. into nude mice. When mAb treatment was started 30 min after the injection of tumor cells, only the IgG2a isotype of CLB-CD19 had a marked antitumor effect in vivo. If treatment with IgG2a anti-CD19 mAb alone was delayed until Day 10 after tumor injection, no therapeutic effect was observed. However, the combination of this delayed mAb treatment with recombinant interleukin 2 (rIL-2) inhibited the growth of the Daudi cells in the nude mice, while treatment with rIL-2 alone was ineffective. The results of in vitro experiments showed that peritoneal exudate cells were able to inhibit the proliferation of Daudi cells in the presence of the IgG2a isotype variant of CLB-CD19 mAb but not in the presence of the other CLB-CD19 mAb isotype variants. Fresh nude mouse spleen cells did not mediate antibody-dependent cellular cytotoxicity against CLB-CD19 mAb-sensitized Daudi cells, irrespective of the isotype used for sensitization. However, preculture of these spleen cells with rIL-2 induced antibody-dependent cellular cytotoxicity against CD19+ target cells sensitized with CLB-CD19 mAb of all isotypes. These results indicate that it is possible to enhance mAb-dependent effector systems in vivo with the lymphokine rIL-2.

Patients' and doctors' delay in the diagnosis and treatment of cutaneous melanoma.

Factors involved in patients' and doctors' delay in the diagnosis and treatment of primary or metastatic cutaneous melanoma were studied in an inquiry of 284 patients in 12 hospitals. The most important patients' delay factors were: unawareness of the danger of the earliest and to a lesser extent the late signs and symptoms in the evolution of melanoma. Age and socio-economic status were of negligible importance in this respect. When the primary melanoma was easily visible, patients noticed the first signs of the developing melanoma significantly earlier than when the primary was not visible; nevertheless visibility had no impact on tumor thickness. This again shows that the early signs were not experienced as signs that required a visit to a doctor. Patients who showed their melanoma as a secondary complaint while they visited the doctor for another condition, had a more favorable tumor thickness than those who only came for their melanoma. The most important factors in doctors' delay were lack of suspicion (often due to amelanosis), and the performance of an incisional or punch biopsy and treatment without histopathology. Melanomas discovered by chance during a physical examination while the patient was totally unaware of the lesion had a most favorable tumor thickness. These observations indicate that a better education of the general public and of the medical profession is warranted, in order to improve the prognosis of cutaneous melanoma.

The significance of conversion of skin reactivity to efficacy of bacillus Calmette-Guérin (BCG) vaccinations given immediately after radical surgery in stage II melanoma patients.

A group of 668 stage II melanoma patients was entered into a randomized prospective study aimed at evaluating the efficacy of adjuvant BCG, 5-(dimethyltriazeno)imidazole-4-carboxamide (DTIC), or a combination of the two, given immediately after radical lymph node dissection. Of these, 176 patients received BCG and 164 BCG plus DTIC. These 340 patients had histologically proven metastatic nodes and 156 had a negative skin reactivity to BCG at the beginning of treatment. The distribution of known prognostic factors (sex, age, number of positive nodes, extracapsular invasion) was balanced in the groups of patients either with initially negative or with positive skin reactivity. All patients who were initially non-reactive to BCG developed skin reactivity after 6.7 +/- 9 BCG vaccinations. Disease-free and overall survival of patients receiving BCG or BCG + DTIC with an initially negative skin reactivity to BCG was significantly (P = 7 x 10(-3) better than that observed in patients with an initial positive skin reactivity. This finding was still evident after adjustment for other known prognostic criteria (P = 0.02). It seems likely that the initial BCG skin reactivity as such marks the prognosis; however, some therapeutic effect of BCG treatment in patients having initially no skin reactivity to BCG, can not be ruled out.

Combination of cisplatin, vindesine, and dacarbazine in advanced malignant melanoma. A Phase II Study of the EORTC Malignant Melanoma Cooperative Group.

A Phase II study combining cisplatin, vindesine, and dacarbazine was performed on patients with disseminated malignant melanoma by the European Organization for Research and Treatment of Cancer (EORTC) Malignant Melanoma Cooperative Group (MMCG). The treatment consisted of intravenous administrations of dacarbazine 450 mg/m2, vindesine 3 mg/m2, and cisplatin 50 mg/m2 on day 1 and 8 of each course. Courses were repeated every 4 weeks. Treatment was discontinued in case of progression after two courses, otherwise continued until progression or for a minimum of six courses. One hundred five patients entered the trial with 92 patients being evaluable. The response rate calculated after clearance by the Extramural Review Committee shows four complete and 18 partial responses, i.e., 24%, with a median remission duration of 23 weeks. Toxicity and subjective tolerance to this regimen were moderate, requiring 140 modifications of 642 administrations (22%). Main toxicities were nausea and vomiting (95%), leucopenia (70%), alopecia (56%), peripheral neuropathy (32%), and nephrotoxicity (17%). The discussion emphasizes some particular points of interest in the management of advanced malignant melanoma.

Results of radical dissection of the groin in patients with stage II melanoma and histologically proved metastases of the iliac or obturator lymph nodes, or both.

In order to determine the value of the pelvic part of the radical dissection of the groin, the histories of 23 patients with Stage II (UICC) melanoma with histologically proved metastases of the iliac or obturator lymph nodes, or both, were studied. Histologically proved primary melanomas were found in 19 patients, 18 were localized on the leg and one on the trunk. In four patients, the primary melanoma was unknown. Seven patients had received adjuvant radiotherapy. The time between node dissection and the moment of analysis was two years for 17 patients, five years for 11 patients and ten years for ten patients. Calculated actuarially, 42 per cent of the patients were still without distant metastasis after two years and 32 per cent after five and ten years. At the time of the analysis, ten patients were alive, nine without evidence of disease and three having survived for more than five years. Remarkably, the primary tumor of all three patients was "unknown" and they had all had adjuvant radiotherapy. In pooling the data of this series with those from the literature, it appears that, of 78 patients with Stage II melanoma and deep node involvement, 12 had a disease-free survival time of more than five years after therapeutic radical groin dissection. Involvement of deep nodes does not always seem to equate with systemic disease. We think that, when there is an indication for a therapeutic groin dissection, an en bloc superficial and deep lymph node dissection is warranted.

A monoclonal antibody specific for cells of the melanocyte lineage.

A monoclonal antibody, NKI/beteb, was prepared against membranes from a human melanoma metastasis, and in immunoprecipitates of melanoma cell lysates specific 100- and 7-kd glycoproteins were found. The large glycoproteins were also present in conditioned medium of melanoma cell lines. The antigen is located on the inner side of membranes of (pre)melanosomes and premelanosomelike vesicles. The antibody reacted in the immunoperoxidase test on frozen tissue sections with 27 of 28 nevocellular nevi (15/16 common, 12/12 dysplastic), 39/39 primary melanomas (3 intraepidermal, 24 cutaneous, 12 choroidal), 56/63 melanoma metastases, and 4/4 clear-cell sarcomas (melanoma of soft tissue). With sections of formalin-fixed paraffin-embedded tissues, the reaction was less sensitive. No reactivity was detected with frozen sections of 185 other tumors, except for 1 case of non-Hodgkin's lymphoma in which macrophages were positive. With the exception of melanocytes, all frozen sections of adult tissues that were tested were negative with NKI/beteb. On the basis of its tissue distribution so far, the antigen recognized by NKI/beteb seems to be a specific and sensitive diagnostic marker for cells of the melanocyte lineage.