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We have identified antibodies against cytoplasmic components of allogeneic uveal melanoma cells in the sera of 16 of 31 patients with proven intraocular melanoma. Similar antibodies were found in 27% of controls and in 24% of patients with uveal nevi. The antibodies in these 3 groups of subjects were absorbed by components of uveal and cutaneous melanoma cells but not by those of normal choroidal melanocytes, normal uvea and retinal pigment epithelium, dermal nevus, pigmented skin or fetal cells. We found that specificity of the antibodies was also demonstrated by absence of reactivity with normal choroidal melanocytes, loss of reactivity after immune blocking, and absence of reactivity of melanoma antigen with conjugated antihuman immunoglobulin alone. Attempted absorption of the conjugated antihuman immunoglobulin by components of uveal melanoma cells did not alter the reactivity of the conjugated antihuman immunoglobulin.

Plasma membranes were purified from tumor tissue of five malignant melanoma patients. The purified membranes were solubilized by sonication in KC1 in the presence of Triton X 100. The soluble membranes that were obtained by this procedure contained human melanoma tumor-specific antigens. The antigens were separated from the other soluble plasma membrane proteins by chromatography on affinity columns made from the patients' tumor-specific pre-absorbed IgG coupled to Sepharose 4B. Specific antigens which were eluted from these affinity columns with potassium thiocyanate were determined by crossover immunoelectrophoresis. The five tumors appear to have a similar complement of melanoma tumor-specific plasma membrane-associated antigens.

The immune response of malignant melanoma bases on evident humoral and cellular defence mechanisms of the host against his tumor. Of special interest is a defective immune response, developing in the course of the disease. Hence, any immunotherapy aims at an immunostimulation and immunoregulation. Local and systemic nonspecific immunotherapy try to raise an immune response against the tumor by stimulating unspecifically the whole immune system. On the contrary, specific immunotherapy tries to stimulate directly the defence mechanisms against the tumor by transfusion of antisera (passive immunotherapy) and sensitized cells (adoptive immunotherapy) and by immunizing the patient with tumor tissue (active immunotherapy). One of the best ways in therapy of melanoma seems to be the combination of immunotherapy with chemotherapy, as yet employed in BCG and DTIC treatment.

During studies that showed the presence of fetal antigens on the surface of human malignant melanoma tumor cells, polyvalent antisera specific for human fetal tissues of varying ages were developed. These reagents demonstrated varying patterns of expression of fetal antigens at different ages in various tissues of the human fetus. The possibility that nonneoplastic adult cells showing either maturation arrest or excessive proliferation also might express fetal antigens led to studies of human bone marrow. Although normal bone marrow cells expressed low levels of fetal antigens, large amounts were seen on bone marrow cells of patients with anemias due to iron, B12, or folic acid deficiencies, as well as on those with leukemia. Moreover, normal adult tissues adapted to long-term culture also expressed fetal antigens. After 3 weeks in organ culture adult human skin showed morphological changes similar to those seen in fetal periderm and strongly expressed fetal antigens. In addition, lymphoblasts in long-term cultured human lymphoid cell lines established from normal donors also carried surface fetal antigens. These latter antigens were shared with neoplastic B-cells (chronic lymphocytic leukemia) but not with T-cells. Their expression varied with the cell cycle. The reexpression of fetal antigens on malignant cells is thought to signal a basic derangement in the control of differentiation which is considered to be peculiar to neoplasia. However, these studies indicate that normal adult cells also may reexpress fetal antigens under circumstances unrelated to neoplasia but associated with either maturation arrest or rapid and excessive proliferation.

Antibodies directed against different components of ocular melanoma cells were identified by indirect immunofluorescence in the sera of 12 patients with histologically proven malignant melanoma of the uvea. Antibodies reacing with autologous surface membrane antigens were found only in the sera of the 5 patients with the smallest tumors. The sera from most of the patients with small, large, or extraocular tumor contained antibodies directed against autologous cytoplasmic components while about one-half of the tests for antibodies reacting with allogeneic cytoplasmic antigens were positive. The incidence of positive tests with cytoplasmic antigens, in 67 patients with clinically diagnosed benign uveal nevi and 56 other individuals not showing evidence of proliferative lesions of uveal melanocytes, was 25%.

Circulating immune complexes were detected in 62 individuals with malignant melanoma by precipitation with isolated human C1q and polyclonal rheumatoid factors. In 56 patients the C1q deviation assay showed low to moderate levels of complexes, with increased amounts with advancing stage of disease. Both heavy (greater than 19S) and intermediate (7S to 19S) varieties were present, and complexes containing tumor antigen-antibody or antibody-anti-antibody were identified. Complexes were found in the kidneys of one patient with malignancy and the nephrotic syndrome and in two further patients with melanoma in whom there were no clinical manifestations of nephrosis. Serial determinations in 51 patients showed slow cyclic variations in the levels of complexes and fluctuations in response to therapy. The coexistence of anti-antibodies, immune complex disease, and anergy in melanoma patients may indicate a deranged immune regulation consequent to chronic antigenic stimulation by the tumor.

In tumor-draining lymph nodes, humoral immunity is diverted from individually specific, cytotoxic anti-membrane AB to less specific anti-cytoplasmic AB with progression of the disease. Auto-anti-anti-bodies and suppressor cells seem to be involved in the failure of control of metastasis.

A patient with disseminated malignant melanoma developed mild renal failure with proteinuria four months following the diagnosis of his neoplasm. Renal biopsy showed a membranoproliferative glomerulonephritis. Electron microscopy revealed intramembranous dense deposits in the glomeruli as well as in tubular basement membranes and Bowman's capsule. Immunofluorescence microscopy demonstrated granular staining for C3 in these same locations and granular deposits of immunoglobulins in the glomerular capillary loops. Light and electron microscopy of the autopsy kidney obtained 5 months later revealed progression of the number, size and density of the deposits. Eluates from both autopsy and biopsy kidney contained melanoma antigen and anti-melanoma antibodies. This case illustrates the association between the dense deposits and neoplasia with the dense deposits representing immune complexes with the tumor as antigen.

Antibodies reacting with autologous and allogeneic cytoplasmic antigens and with autologous surface membrane antigens of uveal melanoma cells were demonstrated in the serum of three patients with relatively small, histologically-proven malignant melanomas of the choroid. The significance of these findings and planned applications of these investigations with regard to diagnosis, prognosis, and therapy of patients with suspected or proven ocular melanomas are discussed.

The presence of antibody in patients with malignant melanoma is well established if one examines the serum. In this report we have attempted to identify antibody within solid tumours showing that they are rarely present in any appreciable quantity on the surface of tumour cells but can be seen frequently on a number of different types of host cell within the tumours. This is discussed in the light of the role of antibody in the circulation and the possibility of antibody behaving as a blocking factor in vivo.

The interaction between immune system and growing tumor can be expressed differently at different stages of the disease. This presentation covers three facets of these reactions in melanoma patients. A. The Primary Tumor. Time-lapse cinematography, with an analysis of lymphocyte movement demonstrated positive and negative chemotaxis against tumor tissues which correlated with their histological presence or absence within the primary tumor. B. The Regional Lymph Nodes. Histological examination of regional lymph nodes showed an increase in germinal center activity and B cell number, with a decrease in sinus histiocytosis and monocyte count as the tumor progressively invaded the node. This correlated with the elution studies, wherein the antimembrane antibody decreased and the anti-cytoplasmic antibody increased during the same period of progression. C. Humoral Immunity and Metastasis. Clinical metastasis heralded the decrease of anti-membrane antibodies with a rise in anti-immunoglobulins, especially anti-idiotypic antibodies and immune complexes containing tumor-directed antibody and either antigen or anti-immunoglobulin. This triad of anti-immunoglobulin, immune complexes and anergy as seen in other diseases with persistent antigenic stimulation, results in abnormal regulation and derangement.

Biopsy specimens and sera were obtained from 103 melanoma patients. Autoantibodies were demonstrated by (1) complement-dependent cytotoxicity of autologous melanoma cells in short-term culture; (2) complement-dependent inhibition of ribonucleic acid synthesis; (3) immunofluorescent staining of the cytoplasm of killed melanoma cells and of the surface membrane of viable melanoma cells. Over one-third of the sera studied had antibodies to autologous melanoma cells. Although for technical reasons all three tests could not be performed with the cells from every melanoma, whenever multiple testing was possible there was complete concordance. The autoantibodies were virtually confined to patients in whom the disease was not widely disseminated, and over 80% of such patients had positive sera. In a limited number of patients who have been followed autoantibodies disappeared as the disease progressed to become widely disseminated. Two patients with generalized disease developed autoantibodies following inoculation by their own irradiated tumour cells.TWO TYPES OF AUTOANTIBODIES WERE RECOGNIZED: one, active against antigen(s) in the cell surface membrane, was specific for each tumour-that is, only the autologous serum reacted-and was concerned in the cytotoxic activity; the other reacted with cytoplasmic antigens which appeared to be present in most or all melanoma cells.

A case of leptomeningeal melanoma in an African child of 7 years is presented together with a survey of pigmentation in the normal African brain. There is a direct relationship between the depth of pigment of the leptomeninges and the skin in Ugandan Africans, suggesting that similar factors operate in the control of melanocytes in these two sites.

Thirteen patients have been immunized with their own irradiated melanoma cells on one or more occasions. Following these autografts tumour-specific cytotoxic antibodies were produced, the longest response lasting 14 days. This procedure had no apparent effect on the course of the disease in these patients.

The frequency and regional distribution of pigmented naevi has been studied in Ugandan Africans. An average of 11 naevi was present in each adult. The association between the distribution of naevi and that of malignant melanoma is discussed and evidence is presented that pigmented skin is able to prevent the malignant transformation of naevi and the spread of already established malignant melanoma. [ABSTRACT FROM AUTHOR]