Melanosomes are subcellular organelles with unique properties: they are producers and scavengers of cytotoxic quinones and are involved in free radical reactions. They behave as energy absorbing and converting devices, h...Melanosomes are subcellular organelles with unique properties: they are producers and scavengers of cytotoxic quinones and are involved in free radical reactions. They behave as energy absorbing and converting devices, have an ion-exchange capacity and show affinity to a variety of substances. Even in the absence of melanin, premelanosomes show characteristic properties due to the presence of specific proteins. The melanosome concentration in pigment tissues is high enough to offer possibilities for influencing pigment cells and for detecting and/or killing melanoma cells. The exploitation of melanosome properties in practice is reviewed.
von Stamm U, Bröcker EB, von Depka Prondzinski M
… +5 more, Ruiter DJ, Rümke P, Broding C, Carrel S, Lejeune FJ
Melanoma Res
· 1993 Jun · PMID 8104570
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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 mel...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.
Benathan M, Alvero-Jackson H, Mooy AM
… +2 more, Scaletta C, Frenk E
Melanoma Res
· 1992 Dec · PMID 1337997
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Resistance to alkylating agents has been correlated with cellular levels of reduced glutathione (GSH) and glutathione-S-transferase (GST). GSH is also involved in regulation of melanin synthesis. Therefore, we examined s...Resistance to alkylating agents has been correlated with cellular levels of reduced glutathione (GSH) and glutathione-S-transferase (GST). GSH is also involved in regulation of melanin synthesis. Therefore, we examined sensitivity to melphalan as a function of differentiation and GSH/GST levels in three human melanoma cell lines. The Me8 cell line, classified as undifferentiated on the basis of cell shape, absence of pigment, insignificant dopa oxidase activity and presence of inhibitors of dopa-melanin formation, showed the lowest GST activity among the cell lines investigated. GLL19 cells exhibited normal differentiation as indicated by the presence of dendrites, typical eumelanosomes, melanin granules and dopa oxidase activity. These cells showed the highest GSH content and the highest GST activity. The JUSO cell line showed incomplete differentiation, and its dopa oxidase and GST activities were intermediate between the Me8 and GLL19 cell lines. The sensitivity of melanoma cell lines to melphalan increased with their degree of differentiation; it was lowest for Me8, intermediate for JUSO and highest for GLL19. Dibutyryl cyclic AMP (dbcAMP) enhanced melphalan toxicity against Me8 cells. Depletion of intracellular GSH with buthionine sulphoximine (BSO) resulted in a three-fold increase in melphalan sensitivity in all three cell lines. Our results indicate that melphalan toxicity is related to cell differentiation and GSH status of melanoma cells. Based on the observed relationship between dopa oxidase, GSH/GST levels and drug toxicity, it is proposed that competition for the GSH pool between quinonoid melanin intermediates and melphalan could diminish drug conjugation and increase cytotoxicity.
Natali PG, Nicotra MR, Citro G
… +3 more, Cucco C, Bigotti A, Kantor RR
Melanoma Res
· 1992 Nov · PMID 1362663
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Intrinsic and acquired resistance to chemotherapeutic agents represents the major clinical obstacle in the control of most tumours. In vitro studies have established that multiple mechanisms, including changes in drug up...Intrinsic and acquired resistance to chemotherapeutic agents represents the major clinical obstacle in the control of most tumours. In vitro studies have established that multiple mechanisms, including changes in drug uptake and efflux and in detoxifying enzymes, are responsible for drug resistance. Among the latter, glutathione S transferases (GST) have been recognized to play a relevant role. In the present study we have evaluated GST pi immunohistochemically as well as enzymatically in benign and malignant primary and metastatic lesions of the melanocyte lineage. A parallel analysis of the multiple drug resistance (MDRI) gene product was performed in a representative number of specimens. Results of this study demonstrate that while GST pi is constitutively expressed by the melanocyte lineage, independently from the transformed stage, MDRI p-glycoprotein is detected with a significantly lower frequency. These findings clearly indicate that GST pi represents the major detoxifying metabolic pathway of the melanocyte lineage and may be responsible for the high degree of inherent resistance of malignant melanoma to available cytostatic treatments.
Results obtained using a computerized image analysis system as an aid to clinical diagnosis of melanoma are reported. The system comprises a colour television camera connected through a digitizing board to a 386 personal...Results obtained using a computerized image analysis system as an aid to clinical diagnosis of melanoma are reported. The system comprises a colour television camera connected through a digitizing board to a 386 personal computer. By means of original algorithms able to measure the shape, the colours and texture of a pigmented lesion of the skin, the system provides eight on/off indicators that are matched with the histological diagnosis to identify benign and malignant pigmented lesions. The chances that a given lesion is malignant increase with the increasing number of positive indicators. The training field of the system was constituted of images and data of 169 cutaneous lesions in 165 patients. Taking two positive indicators as the threshold between pigmented benign and malignant lesions, the efficiency of the system is 0.98, the positive predictive value is 0.45 and the negative predictive value is 0.95. These values were confirmed in a series of 44 pigmented lesions, 10 of which were melanoma, that constitute the present test series. The authors conclude that this computerized image analysis system should be regarded as a useful aid to diagnosis for a non-expert clinician. The system limit is transformation within a naevus.
Mascagna D, Ghanem G, Morandini R
… +4 more, d'Ischia M, Misuraca G, Lejeune F, Prota G
Melanoma Res
· 1992 May · PMID 1643421
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New tyrosinase-targeted compounds based on structural variants of the prototype unit 4-aminophenol have been synthesized and screened for their potential as antitumour agents against malignant melanoma. Cytotoxicity assa...New tyrosinase-targeted compounds based on structural variants of the prototype unit 4-aminophenol have been synthesized and screened for their potential as antitumour agents against malignant melanoma. Cytotoxicity assays showed that N-4-hydroxyphenylglycine (NHPG) and its alpha-methyl derivatives methylphenylglycine and dimethylphenylglycine exhibit significant antiproliferative effects on pigmented human melanoma cell lines (HBL), with inhibitory concentrations at 50% (IC50) around 80 micrograms/ml. A marked increase in cytotoxicity was observed with morpholine-containing 4-aminophenols, e.g. N-(2-morpholinoethyl)-4-aminophenol, which showed an IC50 of 20 micrograms/ml of HBL cells. Much more pronounced was the effect of the diacetoxy-derivative, DiAcMoAc, which showed an IC50 of 15 micrograms/ml on HBL cells and as low as 2 micrograms/ml on tyrosinase-containing, non-pigmented human melanoma cells (LND1), with a toxicity response of the same order of magnitude as that of melphalan. These results open interesting perspectives in the design of new targeted pro-drugs against malignant melanoma.
Previous studies on mice carrying melanoma have shown that 5-iodo-2-thiouracil (ITU) is accumulated in the tumours due to its specific incorporation into melanin during its synthesis. ITU is also selectively localized in...Previous studies on mice carrying melanoma have shown that 5-iodo-2-thiouracil (ITU) is accumulated in the tumours due to its specific incorporation into melanin during its synthesis. ITU is also selectively localized in murine melanoma metastases and in cultured human melanoma cells. Progressive formation of melanin is, however, a prerequisite for the incorporation. Four patients with disseminated melanoma were injected intravenously with 39-62 MBq [131I]TU. Blood and urine samples were gradually collected, and 3-7 days postinjection tumours were biopsied and examined by impulse counting. The patients were scanned with a gamma camera over the total body daily for 3-4 days. The radioactivity was rapidly excreted. Poor melanin pigmentation of the tumours and low proliferation rate (possibly induced by chemotherapy) decreased the uptake of radioactivity by the tumors, and no imaging was possible. One of the patients, however, had clearly progressive disease with darkly pigmented metastases which contained considerably higher levels of radioactivity than the surrounding skin. Calculations indicated that a doubling of the radioiodine dose would probably make visualization of the tumours possible.
A total of 761 melanocytic lesions were studied to elucidate the usefulness of clinical features for the diagnosis of dysplastic naevi. Characteristics associated with high (irregular border, irregular pigmentation), int...A total of 761 melanocytic lesions were studied to elucidate the usefulness of clinical features for the diagnosis of dysplastic naevi. Characteristics associated with high (irregular border, irregular pigmentation), intermediate (black coloured areas, largest diameter greater than 0.5 cm, change of size, change of colour) and low diagnostic efficiency could be defined. Combinations of criteria had high sensitivities: at least one of the following four criteria was positive in 96% of the dysplastic naevi and in all melanomas with less pronounced clinical characteristics: irregular border, irregular pigmentation, greatest diameter greater than 0.5 cm, black coloured areas. A lesion is therefore unlikely to be a dysplastic naevus or a melanoma if all these criteria are absent. When change of size and change of colour were analysed in addition to the features mentioned above a sensitivity of 0.96 was found for at least two of these six criteria. At least three of these six criteria were observed in all melanomas with less pronounced clinical characteristics. However, a rather low specificity (0.19 for at least one of four positive criteria, 0.20 for at least two of six positive criteria) indicated that dysplastic and non-dysplastic naevi cannot be clinically differentiated with acceptable certainty. With less stringent histological criteria approximately twice as high specificities were found. Specificities were about twice as high in a subgroup of patients with at least one proven dysplastic naevus besides the lesion under diagnostic consideration. This facilitates the identification of individuals at risk of developing a melanoma.
The frequency of melanoma (CMM), and of common and dysplastic naevi (CN and DN) in areas of skin chronically, intermittently and rarely exposed to UV light was investigated in 121 melanoma patients (30-50 years) and 310...The frequency of melanoma (CMM), and of common and dysplastic naevi (CN and DN) in areas of skin chronically, intermittently and rarely exposed to UV light was investigated in 121 melanoma patients (30-50 years) and 310 controls. Both cases and controls had significantly more CN in intermittently exposed areas than in areas chronically or rarely exposed. The ratio of observed to expected number of CMM was also highest in intermittently exposed skin (1.3 compared to 0.8 in chronically exposed and 0.5 in rarely exposed areas). Thus, intermittent UV exposure seems to have the most potent 'naevogenic' as well as carcinogenic effect on melanocytes. Nineteen per cent of controls and 56% of cases had naevi fulfilling the clinical criteria for DN. The distribution pattern of DN was clearly different from that of CN and does not accord with the idea that UV light is a major aetiological factor for DN. The probability of CMM significantly increased with the degree of relative clustering of CN (p less than 0.05) and of DN (p less than 0.01). This co-variation of naevi and CMM over the body surface might be the result of the local insults to the melanocyte system caused by UV light and/or to the fact that naevi are precursor lesions of CMM.
The pigmented human melanoma cell line, MM418, became demelanized when treated continuously with a nontoxic level of halistanol trisulphate (HTS), a C29 steroidal detergent isolated from a marine sponge. Nontoxic levels...The pigmented human melanoma cell line, MM418, became demelanized when treated continuously with a nontoxic level of halistanol trisulphate (HTS), a C29 steroidal detergent isolated from a marine sponge. Nontoxic levels of halistanol or of a range of anionic, cationic and neutral detergents had no such effect. Control MM418 cells varied greatly in size, appearance and pigmentation; HTS-treated cells were smaller than controls, had a uniform, generally bipolar appearance, and lacked pigment. HTS induced only minor changes in cell ultrastructure, with fewer mature melanosomes being found in treated cells. Suppression of melanin synthesis was apparent within 24 h of addition of HTS, as judged by inhibited incorporation of the false precursor, 5[125I]-2-thiouracil. Reversal of inhibition occurred within the same period after removal of HTS. Tyrosinase activity gradually decreased to 25% of the control value during a 19-day treatment with HTS, and expression of two carbohydrate-dependent tyrosinase epitopes, 5C12 and 2B7, was abolished. Expression of one other melanosomal protein and of vimentin was not affected. The results suggest that HTS inhibits maturation of tyrosinase to a form associated with melanin synthesis.
The rate of oxidation by purified mushroom tyrosinase of 30 compounds was measured by oximetry, and the tyrosinase-dependent cytotoxicity of each estimated in an in vitro assay using exposure of non-melanogenic cells to...The rate of oxidation by purified mushroom tyrosinase of 30 compounds was measured by oximetry, and the tyrosinase-dependent cytotoxicity of each estimated in an in vitro assay using exposure of non-melanogenic cells to the agents in the presence and absence of tyrosinase. Cytotoxicity was estimated by immediate inhibition of DNA synthesis; 4-hydroxyanisole was used as the reference material. Compounds that were not oxidized by tyrosinase were found to be non-toxic but there was no direct relationship between the rate of oxidation and the relative cytotoxicity of those materials that acted as substrates for the enzyme. Thioethers were found to be more cytotoxic than the corresponding phenoxyethers. This was partly due to their greater rate of oxidation by tyrosinase and, in the case of propylthiophenol, the consequence of higher effective toxicity of the lipophilic species. The optimum chain length for the side chain of the oxyethers was three saturated carbon atoms and the toxicity appeared to be influenced by the lipophilicity of the compounds, possibly reflecting the relative lipid solubility of the putative toxic ortho-quinones generated from them. The maximum tyrosinase-dependent toxicity observed was in the range 5-6 times the relative toxicity of 4-hydroxyanisole. Sulphinyl and sulphonyl derivatives were inactive. In addition to oxyethers and thioethers, esters and glycosides of oxyethers were also examined and were found to be toxic in the presence of tyrosinase when hydrolysed. The succinates were found to be oxidized and toxic in our test system, suggesting that they rapidly underwent spontaneous hydrolysis. Oximetry data suggest that slight spontaneous hydrolysis of the other compounds occurs but they were not toxic in our assay. Ring-methylated phenoxyethers were oxidized relatively slowly and were non-toxic. Fluorine-substituted phenoxyethers were oxidized slightly more rapidly and exhibited clear toxicity in our system. Sesamol was oxidized to a black pigment but was non-toxic in our assay. A water-soluble vitamin E derivative was not oxidized and was non-toxic. Allyl hydroquinone was not oxidized but exhibited significant direct toxicity.
A series of 233 consecutive primary cutaneous melanomas was histologically and clinically studied. Histologically, 53 melanomas (22.7%) were associated with naevus cells. Such a high degree of association suggests that m...A series of 233 consecutive primary cutaneous melanomas was histologically and clinically studied. Histologically, 53 melanomas (22.7%) were associated with naevus cells. Such a high degree of association suggests that melanocytic naevus may be a precursor of a large number of melanomas. Analysing the cases according to Clark's levels and Breslow's index, a decrease in the naevus-melanoma association was seen with tumour progression, suggesting that advanced tumours may overgrow pre-existing nevus cells, appearing as de novo melanomas. The comparison between histological and clinical data suggest some interpretations of the natural history of melanoma.
To assist in the distinction of melanoma from benign pigmented lesions, an imaging system was developed, comprising a frame grabber, a microcomputer, a colour video camera and flash lighting with red, green and infrared...To assist in the distinction of melanoma from benign pigmented lesions, an imaging system was developed, comprising a frame grabber, a microcomputer, a colour video camera and flash lighting with red, green and infrared filters. Over an 18-month period, video images of 70 unselected pigmented lesions for which complete diagnostic data were available, were successfully captured using the camera. Analysis software extracted features relevant to the size, colour, shape and boundary of each lesion, and these features were correlated with clinical and histological characteristics on which standard diagnoses of skin tumours are based. For discriminant analysis based on image analysis measurements, equal probabilities were assigned to three specified diagnostic groups, namely melanoma, naevi and 'other', and four of five melanomas were correctly classified when infrared data were included. However when infrared measurements were omitted, all five melanomas were correctly classified, and the overall accuracy of classification of pigmented lesions was 71%. This system holds promise as an aid in the clinical distinction of melanoma from benign pigmented skin lesions.
Malignant melanoma is not only one of the most aggressive and lethal types of neoplasm, but its incidence in the general population is currently increasing at an alarming pace. It is interesting that most melanomas retai...Malignant melanoma is not only one of the most aggressive and lethal types of neoplasm, but its incidence in the general population is currently increasing at an alarming pace. It is interesting that most melanomas retain many of their characteristics of differentiation, including a dendritic nature and the production of melanin. This review discusses the phenotypic properties of melanoma cells, including their state of differentiation, and their tumourigenic and metastatic potentials, and attempts to provide an overview of the state of current research on the interrelationships between those parameters in murine and human systems.
Cultured melanoma cells have been of great value in the study of pigment metabolism. IGR 1 human melanoma cells, established by Dr Christian Aubert, produce melanin in large quantities. These cells have been used for iso...Cultured melanoma cells have been of great value in the study of pigment metabolism. IGR 1 human melanoma cells, established by Dr Christian Aubert, produce melanin in large quantities. These cells have been used for isolation of human tyrosinase which enzyme has not previously been obtained in a pure form. IGR 1 cells contain large amounts of 5-S-cysteinyldopa which is the quantitatively most important catecholic amino acid. This review deals with the metabolism of dopa, cysteinyldopa, glutathionyldopa, cysteine and glutathione, compounds of central importance in pigment metabolism. The information available on tyrosinase, catecholic compounds and on thiols in IGR 1 melanoma cells makes these cells most suitable for further investigation of the metabolism of human melanoma cells.