MACG1, a mouse monoclonal antibody detecting a monosialoganglioside expressed in tumor-infiltrating macrophages.

MacG1 is a mouse monoclonal antibody (mAb) directed against a ganglioside, which is differentially expressed by macrophages infiltrating malignant melanomas and benign melanocytic lesions. mAb MacG1 was obtained by immunization with liposomes containing a mixture of gangliosides extracted from malignant melanoma. The antibody was selected for binding to melanoma gangliosides and for reactivity with frozen tissue sections of malignant melanoma. mAb MacG1 showed reactivity in 25 of 46 melanomas examined but in only 1 of 51 nevi tested. The mAb did not react with melanoma cells but did with cytoplasmic granules and deposits associated with large dispersed cells, which were also found in some nonmelanomatous tumors and in some lymphoid tissues. Using mAbs directed against differentiation antigens these cells were identified as macrophages. In nearly all reactive tissues MacG1-positive macrophages accounted for a minority of the total macrophages. The difference in reactivity between malignant melanomas and nevi could not be explained by the variable numbers of total macrophages in these lesions. It is suggested that mAb MacG1 may define a functionally distinct subpopulation of tumor-infiltrating macrophages. Staining of cells other than macrophages was observed in some normal and malignant neural tissues. MacG1 bound to a monosialoganglioside extracted from melanoma and reacted only with NeuAc alpha 2-3Gal beta 1-4Glc-Cer when tested with a panel of ganglioside standards.

Immunohistochemical localization of ganglioside GD3 in human malignant melanoma, epithelial tumors, and normal tissues.

The specific tissue distribution of melanoma-associated ganglioside II3-alpha-N-acetylneuraminosyl-alpha 2----8-N-acetylneuraminosyllactosylceramide (GD3) was studied on 175 cryopreserved, unfixed human tissue sections with R-24 mouse monoclonal antibody by indirect immunoperoxidase staining. A striking specificity of monoclonal antibody R-24 for malignant melanoma tissues was established. Ganglioside GD3 was detected in all 21 tissue sections of 21 patients with primary melanoma and in all 37 probes of 24 patients with metastatic malignant melanoma. The majority of tumor cells in the samples of primary malignant melanoma expressed GD3; however, GD3 expression was more heterogeneous in samples of metastatic lesions even in different metastases of the same patient. Of 11 nevi, 9 reacted with monoclonal antibody R-24, while melanocytes in the basal layer of normal skin stained only weakly and irregularly. None of the 32 normal and 12 fetal human tissue types were R-24 positive, but a strong cytoplasmic staining was observed with single cells in the dermis and in the interstitial tissue of the gastrointestinal tract, in the interlobular septa of the thymus, and in other distinct locations. Only two malignant carcinoid tumors of 38 nonmelanomatous tumors tested reacted with monoclonal antibody R-24.

Expression of the c-src protooncogene in human skin tumors.

Retroviral oncogenes are genetic elements, the expression of which is responsible for the transformed phenotype of cells. These genes are derived from normal cellular DNA sequences called cellular protooncogenes, which are present in all human cells and seem to have potential transforming ability in tumors of nonviral origin, since it is possible that they undergo structural alterations and/or changes in their expression. Human skin tumors were analyzed in this study with respect to the expression of the c-src protooncogene, the cellular homologue of the Rous sarcoma virus transforming gene, by measuring the enzymatic activity of its gene product, the pp60c-src kinase activity. Tyrosine-specific kinase activity was detected in all skin tumors tested. The expression pattern of the c-src gene product in the melanomas tested was differential and varying kinase levels in different metastases from the same patient were detected. The elevation of kinase activity as compared to normal skin ranged from about 4- to 20-fold.

Presence of cytoadhesins (IIb-IIIa-like glycoproteins) on human metastatic melanomas but not on benign melanocytes.

Glycoproteins IIb and IIIa, a heterodimer complex, play a vital role in blood platelet aggregation and are members of a wide family of membrane receptors known as integrins or cytoadhesins. Cellular interaction to extracellular matrix (ECM) adhesive proteins is mediated by integrins. Certain tumor cells are known to interact with ECM and blood platelets in the process of metastasis. However, it is not known if tumor cells, compared with their normal counterparts, acquire IIb-IIIa-like receptors to help them in their metastatic spread. In this study, monoclonal antibodies directed against the IIb-IIIa platelet glycoprotein complex were used on frozen biopsies of normal and various tumor tissues to detect the presence of these integrins. These studies demonstrate the presence of IIb-IIIa-like glycoproteins on the cells of metastatic malignant melanoma but not on benign melanocytes and rarely on other tumors. The presence of integrins on melanomas may help explain their propensity for frequent metastasis.

A monoclonal antibody against nerve growth factor receptor. Immunohistochemical analysis of normal and neoplastic human tissue.

The expression of human nerve growth factor (NGF) receptor in tumors and normal tissue was investigated with the use of a monoclonal antibody recently developed against that protein. This antibody, NGFR5, reacted strongly with 100% of 25 nerve sheath tumors. Eight of nine pheochromocytomas and three of three paragangliomas also had positive results, but the immunoreactivity was restricted to the sustentacular cell population. Within cells of melanocytic lineage, there was no immunostaining of melanocytes in normal epidermis, whereas 13 of 14 benign nevi had positive results, primarily involving spindled nevocytic structures within the dermis. NGF receptor was scarcely expressed in human melanoma; 9 of 19 melanomas had positive results, but immunoreactivity was generally restricted to rare cells within the larger tumor cell population. Among nonneurogenic mesenchymal tumors, results were generally negative: 0 of 5 chondrosarcomas, 0 of 6 malignant fibrous histiocytomas, 0 of 3 meningiomas, and 1 of 8 leiomyosarcomas were immunoreactive. Carcinomas were variable in immunoreactivity: 12 of 16 squamous cell carcinomas had positive results, whereas adenocarcinomas demonstrated focal, basal epithelial immunoreactivity and neuroendocrine tumors generally had negative results. Among normal tissues, in addition to expected neural immunostaining, NGFR 5 reacted positively with several nonneural cell types, including lymphoidal follicular dendritic cells, myoepithelial cells, vascular adventitia, and basal epithelium of oral mucosa and hair follicles. Antibodies to NGF receptor may play a role in the identification of benign and malignant soft tissue lesions.

Type IV collagen and laminin staining patterns in benign and malignant cutaneous lesions.

The presence of both laminin and type IV collagen was sought at the dermo-epidermal junction and in the dermis adjacent to benign melanocytic naevi of the junctional, compound, and intradermal types; dysplastic naevi; and both primary and secondary melanoma. In all, 154 lesions were studied, using antibodies to laminin and type IV collagen and an indirect immunoperoxidase technique. The staining patterns seen with the two antibodies were virtually identical, although that of laminin was generally fainter. Breaks in and thinning of the normally continuous line of type IV collagen and laminin at the dermo-epidermal junction were seen in association with the junctional activity of benign naevi, and in malignant melanomas in association with invasive tumour cells. Both benign and malignant cells of the melanocyte series showed relatively light pericellular staining around individual cells and clusters of cells in the papillary dermis. This staining pattern was much stronger in the deeper reticular dermis. It is concluded that the pattern of staining of these two antibodies and in particular the presence of breaks in type IV collagen and laminin at the dermo-epidermal junction are not specific for either benign or malignant melanocytic lesions and cannot be used as a diagnostic marker of invasive malignancy.

Estrogen and progesterone receptors in melanocytic lesions. Occurrence in patients with dysplastic nevus syndrome.

Estrogen and progesterone binding studies in a series of 22 melanocytic lesions from 14 patients with the dysplastic nevus syndrome were done using a fluorescent estrogen and progesterone binding technique. Melanocytic lesions from these patients, including primary cutaneous melanomas, dysplastic nevi, and benign nevi, contained large numbers of estrogen and progesterone binding cells. Comparison is made to a series of control intradermal nevi that had little or no detectable estrogen or progesterone binding. Increased hormonal binding, and possibly responsiveness, is seen in melanomas, melanoma precursor lesions such as dysplastic nevi and congenital nevi, as well as benign nevi from patients with the dysplastic nevus syndrome.

Analysis of epidermal fibrous proteins in inflammatory linear verrucous epidermal nevus.

A baby had inflammatory linear verrucous epidermal nevus (ILVEN). Scale from the lesion was analyzed by sodium lauryl sulfate polyacrylamide gel electrophoresis. It showed a pattern different from psoriatic scale. This procedure was useful in distinguishing the lesion (or ILVEN) from psoriasis, the diagnosis that had been made earlier.

Immunohistochemical study of S-100 protein and neuron specific enolase (NSE) in melanocytes and the related tumors.

Normal human skin, malignant melanoma, nevocellular nevus, blue nevus, nevus of Ota and mongolian spot were immunohistochemically investigated on the localization of S-100 protein and neuron specific enolase (NSE). Tissues were fixed with buffered-formalin, processed with routine procedure and examined by the ABC technique. All cases of malignant melanoma and nevocellular nevus showed a relatively high amount of S-100 protein, but NSE was scantly demonstrated on about the half cases of these tumors. Blue nevus, nevus of Ota and mongolian spot revealed the presence of a small amount of S-100 protein and NSE on the half cases. Normal melanocytes were devoid of S-100 protein and NSE. Our results suggest that S-100 protein is the useful marker for diagnosis of malignant melanoma, and immunoreactive intensity for S-100 protein represents the differentiation of neural crest derived melanogenic cells and tumors.

Incidence of estrogen receptor in benign nevi and human malignant melanoma.

This study was performed as an initial step to evaluate the possible effects of steroid hormones on benign nevi. Cytoplasmic receptors for estrogen were found in nine (41%) of 21 benign nevi from melanoma patients. Ten (37%) of 27 melanoma tumor tissues also had receptor for estrogen. Benign nevi from 17 normal persons had no estrogen receptor activity. The similar incidence of estrogen receptor in benign nevi from melanoma patients and in melanoma tissue itself, compared with the absence of receptor in benign nevi from a normal population, suggest that steroid hormones such as estrogen may represent a molecular alteration associated with a potential pathophysiologic transformation of a benign nevus to its malignant counterpart.

[Ratio of cyclo-3':5'-adenosinemonophosphate to cyclo-3':5'-guanosine-monophosphate in human tumor tissue]. / Verhältnis von cyclo-3':5'-adenosinmonophosphat zu cyclo-3':5'-guanosinmonophosphat in menschlichen Tumorgeweben.

In human tumor tissues of different degrees of differentiation--nevus-cell-nevus, basalioma, malign melanoma--the cAMP and cGMP content was determined and compared with the corresponding normal values. It is demonstrated that the quotient of the cAMP to the cGMP values is of importance rather than the latter values for themselves. For the benign tumor, this quotient differs only slightly from that of the adjacent normal, sound tissue. On the other hand, for the two malign tumors a drastic decrease of the quotient as compared to that of the normal tissue was found to occur.

Proton NMR and fast-atom bombardment mass spectrometry analysis of the melanoma-associated ganglioside 9-O-acetyl-GD3.

A glycolipid antigen, detected by a monoclonal antibody (ME 311) obtained by immunizing mice with a human metastatic melanoma cell line (WM 46), was isolated and structurally characterized. Using immunostaining on thin-layer chromatograms for monitoring, 1.0 mg of a pure alkali-labile disialoganglioside was obtained from 23 g of packed melanoma cells (WM 164). Fractionation of the lipid extract was done on DEAE-Sepharose columns into total disialogangliosides which were repeatedly separated by high-pressure liquid chromatography. On mild alkaline treatment, the ganglioside was converted to a slower migrating species identical with a ganglioside GD3 isolated from the same source (Neu5Ac alpha 2----8Neu5Ac alpha 2----3Gal beta 1----4Glc beta 1----1-cer-amide) and specifically detected by monoclonal antibody R24. Comparison of the two gangliosides by fast-atom bombardment mass spectrometry (revealing an acetyl group on the terminal sialic acid on the alkali-labile species) and by 1H NMR (indicating the position of the acetyl group) suggested the following structure: Neu5,9Ac2 alpha 2----8Neu5Ac alpha 2----3Gal beta 1----4Glc beta 1----1-ceramide. This is identical with a ganglioside proposed earlier to exist in melanoma cells (Cheresh, D. A., Varki, A. P., Varki, N. M., Stallcup, W. B., Levine, J., and Reisfeld, R. A. (1984) J. Biol. Chem. 259, 7453-7459). Immunostaining with ME 311 antibody of cell extracts on thin-layer chromatography chromatograms revealed only this ganglioside in the melanoma cells, while normal human brain was negative. However, in one of the total ganglioside extracts tested for presence of binding with antibody ME 311, three gangliosides were found to bind. No evidence was obtained for the presence of the antigenic epitope in mucins or glycoproteins of the melanoma cells.

The polymorphic apocrine nevus: a study of a unique tumor including carcinoembryonic antigen staining.

The apocrine nevus is a rare tumor. We report a 32-year-old man with a neck nodule, histologically an apocrine nevus displaying mature and immature apocrine structures in a distinctive pattern. The carcinoembryonic antigen staining was positive intracellularly only in the smaller luminal structures. We have named this unique tumor a polymorphic apocrine nevus.

Detection of S-100 protein as an aid to the identification of melanocytic tumors.

We sought S-100 protein in sections prepared from formalin-fixed paraffin-embedded tumor and normal tissues by an immunoperoxidase technique with a rabbit anti-bovine S-100 protein antiserum as first antibody and aminoethyl carbazole as developer. S-100 protein was demonstrated in 56 of 56 cutaneous melanomas, regardless of their primary or metastatic status, and in 35 of 35 nevocytic nevi. The amount of S-100 protein was independent of the degree of melanization of the tumors. Only 1 of 51 non-neural non-melanocytic tumors tested contained S-100 protein. The technique is valuable in confirming the melanocytic nature of primary or metastatic tumors and in detecting small foci of metastatic melanoma in lymph nodes and other tissues. It also seems applicable to research on melanocytic anomalies.

Are acquired nevi oestrogen-dependent tumours?

Twelve benign acquired nevi were studied for the possible specific binding of the ER D5 monoclonal antibody. The ER D5 monoclonal antibody identifies the p29 protein, which is found in the cytoplasm of oestrogen-sensitive cells and which is present in a higher concentration than nuclear oestrogen receptors. No staining was seen in nevoid cells, and the results are taken to support the hypothesis that acquired nevi are not oestrogen-dependent tumours.

Fibrous papule: a tumor of fibrohistiocytic cells that contain factor XIIIa.

Factor XIIIa, a blood coagulation factor, has been found in a variety of cell types, including dendritic reticulum cells and fibroblast-like mesenchymal cells. We hypothesized that fibrous papule, a lesion of uncertain histogenesis, was composed of dermal stellate cells and in this report demonstrate that this neoplasm consists of cells that contain this factor. Nevus cells do not contain factor XIIIa.

Melanoma cell heterogeneity. A study of two monoclonal antibodies compared with S-100 protein in paraffin sections.

Fifty-six formalin, Bouin's, and Carnoy's fixed, paraffin-embedded malignant melanomas (21 primary, 35 secondary), were studied by avidin-biotin complex immunohistochemistry using monoclonal antibodies (MoAb) HMB-45 and B1.1, comparing reactivity with polyclonal anti-S-100 protein. B1.1 (anti-CEA MoAb) was expressed in a minor percentage of cells of the invasive component of some primary melanomas, and weak to moderately in scattered metastic melanoma cells. MoAb HMB-45 prepared against melanocytic tumors reacted with over 90% of all tumors studied, being weakly reactive in one, and nonreactive in four metastases. This antibody stained some primary melanomas and their dysplastic nevus components in a heterogeneous manner, but was largely nonreactive in deep dermal nevus cells that were in association with invasive melanoma, enabling recognition of the deepest penetration of melanoma cells in the dermal nevus component. MoAb HMB-45 appears specific for melanoma cells, with no cross-reactivity with nonnevomelanocytic malignant tumors (unlike polyclonal anti-S-100 protein). MoAb HMB-45 is more sensitive in detecting malignant melanoma cell heterogeneity than anti-S-100 protein.