摘要:

Sequence analysis of the promoter region of the murine tyrosinase gene identified various consensus motifs including AP2 sites, cAMP and TPA response elements (CREs/TREs) and retinoic acid response element (RARE) half-sites. By linking two different promoter lengths (2.5 kb or 769 bp) to murine interleukin-2 (IL-2) cDNA we have used IL-2 production by transduced B16 cells to monitor response to inducing agents capable of acting through these elements. Aminophylline or theophylline (0.1–2 mM) added to the culture medium of transfected B16, but not 3T3, cells, increased IL-2 secretion significantly (P>0.05) in a dose-dependent fashion. This response was comparable in cells transfected with either the full length or the truncated promoter. Therefore, the cAMP responsiveness of the tyrosinase promoter probably is mediated by CREs and not AP2 sites, since the truncated promoter contains the former but not the latter regions. Retinoic acid at various concentrations (0.1–1 μM) evoked a standard increase in IL-2 production. Responses were similar for both promoter constructs, which suggests either that each RARE half-site can confer the full retinoic acid response, or that retinoic acid is mediating its effect through pathways independent of the RARE sites. TPA (2 nM–2 μM) had no effect on IL-2 production. These results demonstrate that the tyrosinase promoter can be induced by certain pharmacological agents and raise the possibility that administration of such substances may enhance expression of therapeutic genes controlled by this promoter.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

The liver metastasis formation of two human melanoma cell lines were compared in male and female SCID mice. The intrasplenic injection of both tumour lines resulted in a significantly higher number of liver metastases in male than in female mice; the incidence and weight of spleen tumours, as well as the incidence of metastases were similar. Both melanoma cell lines bound fluorescent oestradiol, progesterone and testosterone conjugates, and proved to be positive for oestrogen receptor-related protein by immunocytochemistry. These observations support the view that endocrine factors influence the progression of human melanomas. This SCID mouse model could be useful in studying the effects of hormonal manipulations on human melanoma metastases.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

We compared ganglioside profiles of animal tumours (B16 and Cloudman S91 murine melanomas, Bomirski Syrian hamster melanoma), which are widely used as models of human melanoma, and of their melanosomal fractions. A ganglioside fraction was extracted and purified and the amount of each component ganglioside was assessed by thin-layer chromatography. GM3 was the dominant ganglioside species in the murine melanomas studied. Unlike human melanomas, the GD3 expression in mouse melanomas was low. GD3 and GM3 were major gangliosides in Bomirski hamster melanoma. Alkali-labileO-acetyl-GD3, a melanoma-specific ganglioside, was detected only in Bomirski melanoma. GD2, which in human melanoma is seen as a distinct signal of tumour progression, was not found in the animal melanomas studied. Melanosomes isolated from B16 and Bomirski melanomas contained GM3 and GD3 as their major ganglioside components. These data extend the group of common antigenic determinants shared by melanosomes and cell surface of pigment cells.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

Mutations of the p53 tumour suppressor gene are common to many human malignancies. Although increased p53 expression has been observed in cutaneous malignant melanoma, mutations of the p53 gene appear to be infrequent. We ex-amined 140 benign and malignant paraffin-embedded melanocytic lesions for p53 protein expression by immunohistochemistry, using the monoclonal anti-p53 antibody DO-7 and a microwave method of antigen retrieval. Fifteen naevi and 25 melanomas were further analysed for p53 mutations within exons 5–8 of the p53 gene. DNA was extracted from paraffin sections and screening for mutations was carried out using PCR-SSCP. We demonstrated p53 protein expression in 33% of naevi (17 out of 51), 35% of primary melanomas (20 out of 58), and 70% of metastatic lesions (15 out of 21). p53 expression in benign lesions was weaker than in malignant lesions in intensity and percentage of cells staining. p53 protein expression in melanomas increased in intensity and percentage of cells staining with tumour progression. In 25% (three out of 12) of metastatic melanomas p53 mutations were detected by PCR-SSCP and increased expression of p53 protein was observed in these tumours. p53 gene mutations were not detected in any benign melanocytic lesions. We demonstrate that antigen retrieval techniques increase p53 immunoreactivity in paraffin embedded melanocytic tissues. p53 protein expression in melanomas increases with depth of tumour invasion. As p53 gene mutations occur infrequently in malignant melanoma, other mechanisms are proposed to influence p53 protein expression in melanocytic lesions.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

Immunohistochemical analysis of the N-ras p21 and the p53 proteins was carried out on formalin-fixed sections of naevi, primary melanomas and metastases from patients with sporadic melanoma (SCMM) and with hereditary melanoma (HCMM)/dysplastic naevus syndrome (DNS). Seven out of 11 (64%) common naevi and three out of nine (33%) dysplastic naevi showed increased cytoplasmic N-ras expression. No p53 immunopositivity could be recognized in any of the naevus samples. However, strong N-ras expression as well as immunopositivity for p53 was recognized among primary melanomas and metastases with significantly higher frequency among samples from patients with HCMM compared with samples from SCMM cases (for N-ras, 40% vs 10%,P< 0.01; and for p53 43% vs 17%,P< 0.05). We have earlier registered N-rascodon 61 mutations among metastases from 59% of patients with HCMM and from 24% of subjects with SCMM. A comparison of the genetic data with the immunohistochemical results showed occurrence of increased N-rasp21 expression in the presence and absence of detectable N-rasmutant alleles. Increased expression of wildtype N-rasp21 may contribute to tumorigenicity in the absence of mutational activation, at least in a subset of melanomas. Altogether, N-ras p21 alterations are registered at earlier stages than p53 alterations in melanoma development and may be of aetiological importance, whereas p53 alterations may be associated with tumour progression in the late stages. The increased frequency of phenotypic alterations registered among samples from HCMM/DNS patients may be a result of genetic instability and hypermutability and may be caused by various genetic changes such as altered gene dosage or regulatory as well as structural mutations.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

Because the resistance of melanoma to cytotoxic therapy may be due to high intracellular glutathione (GSH), we wished to monitor GSH in melanoma cells during treatment. This required an assay suitable for very small samples. So we chose analytical flow cytometry. We calibrated the flow cytometric assays against biochemically determined GSH content in a range of cultured human melanoma cell lines, then applied the assays to fine-needle tumour biopsies. Mercury orange was the first fluorochrome suitable for use in a flow cytometer with a 488 nm light source, but many technical factors influenced the results. Chloromethyl fluorescein diacetate (CMFDA) allowed assay conditions less dependent on details of cell handling. Correlations between biochemical GSH content and fluorescence intensities in cell lines were good for mercury orange and CMFDA. CMFDA is the preferred fluorochrome for estimating cellular GSH content in biopsies from human melanomas. Tumours metastatic to or beyond regional lymph nodes had significantly more cells with high GSH than primary tumours or regional recurrences.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

We have recently described a new method for measurement of tyrosinase activity in small amounts of human serum (100 μl), where the purification of tyrosinase is obtained by adsorption of the enzyme to concanavalin A sepharose. The method, which measures stereospecific dopa oxidation, was used in the winter of 1992–93 for the measurement of activity in serum obtained from 30 healthy subjects and from 10 patients with melanoma metastases. The serum tyrosinase values in the 30 subjects ranged from 0.1 to 1.0 nkatal/l, and the mean value and standard deviation was 0.4 ± 0.2 nkatal/l. In the 10 patients with melanoma, the tyrosinase serum values ranged from 1.1 to 10.6 nkatal/l, and the mean value was 3.1 nkatal/l.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

Between January 1956 and December 1990, 17 patients younger than 17 years with available pathological screens ofde novocutaneous melanoma, and with no other risk factors (xeroderma pigmentosum, giant congenital naevi, congenital melanoma or a proven family history of dysplastic naevus syndrome) were seen at the Gustave-Roussy Institute. The median age was 9 years and 9 months (range 2 years and 3 months–16 years and 9 months). The primary disease was located in the lower extremities in 10 cases, the trunk in five cases, and the upper extremities or head and neck in one case. The disease was localized for 10 patients at presentation (stage I), six had proven nodal metastasis (stage II) and one patient had nodal and breast metastases. The median thickness of the primary lesion was 2.89 mm (range 0.64–10). Five tumours were at level III on Clark's index, eight at level IV and four at level V. Six cases were classified as superficial spreading, two as unclassified radial growth, three nodular, three with Spitzoid cells, and three were unclassified. Two patients presented local recurrence with an initial unclassified melanoma, with a thickness greater than 2.5 mm. At a median follow-up time of 7 years, two patients had died from recurrent disease, and one patient had died from a second malignancy.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

We assessed the risk of cutaneous malignant melanoma associated with the presence of ovulatory abnormalities and with the use of ovulation-inducing agents (such as clomiphene citrate) in a cohort of 3,837 women evaluated at infertility clinics in Seattle, WA, between 1974 and 1985. Computer linkage with a population-based tumour registry was used to identify women diagnosed with melanoma before 1992. Data regarding infertility testing and treatment were abstracted from the infertility clinic medical records for women who developed cancer and a randomly selected subcohort. Twelve women in the cohort developed cutaneous malignant melanoma, in comparison with an expected number of 6.8 cases (standardized incidence ratio = 1.8; 95% confidence interval (CI) 0.9–3.1). Within the cohort, risk was increased among women who had used clomiphene during 12 or more menstrual cycles (relative risk = 2.2; 95% CI 0.5–10.2). All four of the women with this duration of clomiphene use who developed melanoma had ovulatory abnormalities, and three had also used human chorionic gonadotropin (HCG). No elevation in risk associated with the presence of ovulatory abnormalities was observed in the absence of at least 12 cycles of clomiphene exposure; also, there was no increased risk associated with long-term use of clomiphene among women without ovulatory abnormalities, but the number of such women was very small. Thus, it is not certain to what extent the observed increased risk of melanoma in this cohort (if not due to chance) may be attributable to the use of clomiphene or HCG, or is a reflection of some underlying hormonal abnormality for which the drug was administered.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID

摘要:

Restriction of the T cell receptor repertoire suggesting ongoing specific immune mechanisms has recently been described in melanoma tissue by several groups of investigators. The functional relevance for immunotherapy of melanoma, however, has not been established. We studied the T cell receptor repertoire in two melanoma metastases of a patient with a mixed response to immunotherapy. Expression of T cell receptor Vβ regions was determined by subgroup-specific semiquantitative RNA polymerase chain reaction (PCR). In the regressing skin lesion a restricted expression of the T cell receptor repertoire and overexpression of three Vβ subgroup genes was found; no restriction was present in the simultaneously progressing skin lesion of the same patient, compared with peripheral blood lymphocytes. Comparison of T cell receptor vβ gene expression in two metastatic lesions of a patient with simultaneously growing skin metastases, who did not receive immunotherapy, revealed only minor differences. These observations show for the first time an association between restricted T cell receptor repertoire and responsiveness of melanoma to immunotherapy and suggest a role of T cells using the overexpressed Vβ genes for the cytokine-induced tumour regression.

ISSN:0960-8931    

出版商:OVID    

年代:1995

数据来源: OVID