Modulation by interferons of HLA antigen, high-molecular-weight melanoma associated antigen, and intercellular adhesion molecule 1 expression by cultured melanoma cells with different metastatic potential.

The effect of leukocyte (IFN-alpha), fibroblast (IFN-beta), and immune (IFN-gamma) interferon and/or mezerein on the expression of HLA antigens and melanoma-associated antigens by the melanoma cell line MeWo and its metastatic variant MeM 50-10 was investigated, since this information may contribute to our understanding of the molecular mechanism(s) underlying the metastatic process and of the role of cell differentiation and growth suppression in the antigenic changes induced by interferon (IFN). The three types of IFN had no effect on the expression of high-molecular-weight melanoma-associated antigen, but enhanced that of HLA Class 1 antigens and of intercellular adhesion molecule 1 on MeWo and MeM 50-10 cells. The enhancing effect of IFN-gamma was more marked than that of IFN-alpha and IFN-beta. Furthermore IFN-gamma enhanced the expression of intercellular adhesion molecule 1 by MeM 50-10 cells more than by MeWo cells. IFN-beta was shown for the first time to induce HLA Class II antigens; the effect of IFN-beta, like that of IFN-gamma, is differential on the two cell lines and on the gene products of the HLA-D region. Like IFN-gamma, IFN-beta induced only HLA-DR antigens on MeM 50-10 cells. The results of Northern blot analysis with HLA-DR beta, -DQ beta, and -DP beta probes suggest that the differential modulation of the gene products of the HLA-D region by IFN-beta and IFN-gamma reflects transcriptional and posttranscriptional events. The differential susceptibility to modulation by IFN-beta and IFN-gamma of HLA Class II antigens on MeWo and MeM 50-10 cells is an intrinsic property of each cell line, since only small differences were detected in the number and/or affinity of receptors on the two cell lines. Furthermore, the lack of marked effects of mezerein on the antigen-modulating activity of the three types of IFN, in spite of an enhancement of their differentiating activity, suggests that the changes in the antigenic profile induced by IFN do not represent a differentiation-related phenomenon.

Subpopulations of human T lymphocytes in patients with Hodgkin's disease before and after treatment.

Circulating T gamma and T mu cells enumerated in the peripheral blood of 51 untreated Hodgkin's disease (HD) patients and 66 treated HD patients tested after few months to more than 3 years of remission, brought about by radiation and/or chemotherapy. 53 age matched normal healthy individuals were studied as controls. Untreated HD patients showed significant increase in T gamma cells (p less than 0.001) and decrease in T mu cells (p less than 0.001) when compared to normal donors. The abnormal percentages of T cell subsets did not correlate with the severity of the disease. A progressive partial restoration in the proportion of these two subsets of T lymphocytes was seen in the disease free condition. However, even after 3 years of remission the recovery was not equivalent to controls. There was no correlation between the recovery of T gamma and T mu cells with the modality of treatment.

Distribution of subpopulations of T lymphocytes in Hodgkin's disease.

In this paper, two aspects of T cell subsets are investigated. Splenic mononuclear cells (MNC) of untreated Hodgkin's disease (HD) patients have been investigated for proportion of immunoregulatory T gamma and T mu cells from 8 uninvolved spleens and 13 involved splenic tissues from HD patients with splenic involvement. As controls, two normal spleens from accident cases were used. It was found that irrespective of the involvement of splenic tissue in the disease process, the HD spleens showed lower percentage of T gamma cells (23.77 +/- 1.03) and higher sequestration of T mu cells (31.6 +/- 2.13) compared to normal spleens (42.5% and 13% resp.). However, there was no significant difference in the total T cell percentages of HD and normal spleens (49.1 +/- 1.13% and 52% resp.). The results therefore indicated the possibility of abnormal sequestration and traffic of T cell subsets in HD. We have also reported here a comparison between T cell subsets from the PBMNC of treated and untreated HD patients and normal healthy donors as assessed by the FcR markers and monoclonal antibodies of Leu series. It was found that the abnormality in T cell subsets could be demonstrated by FcR markers, while Leu 2a and Leu 3a reactivities did not differ in HD and normal PBMNC. The subset proportions identified by two tests did not tally with each other.

Modulation by cytokines of HLA antigens, intercellular adhesion molecule 1 and high molecular weight melanoma associated antigen expression and of immune lysis of clones derived from the melanoma cell line MeM 50-10.

Clones were isolated from the cultured human melanoma cell line MeM 50-10, which metastasizes in nude mice with a pattern similar to that in patients with melanoma. Analysis with monoclonal antibodies detected heterogeneity among the clones in the expression of HLA class I antigens, HLA class II antigens, intercellular adhesion molecule 1 and high molecular weight melanoma associated antigen. The clones MeM A16 and MeM A18 were also shown to display differential susceptibility to modulation by immune interferon (IFN-gamma) and/or tumor necrosis factor (TNF-alpha) of the expression of the four types of antigens analyzed. In spite of differences in the antigenic profile, the two clones did not differ in their susceptibility to lysis by lymphokine-activated killer (LAK) cells and by anti-HLA-A2 cytotoxic T cells. The increase in the expression of HLA class I antigens induced by IFN-gamma and/or TNF-alpha on the two clones was associated with an increased susceptibility to lysis by anti-HLA-A2 cytotoxic T cells. Because of the metastasizing properties of cultured melanoma cells MeM 50-10, the clones we have isolated, with their distinct antigenic profile and differential susceptibility to modulation by cytokines, may represent useful models to investigate the role of distinct antigenic structures in the metastatic process.

Differential modulation by tumor necrosis factor and immune interferon of HLA class-II antigens expressed by melanoma cells.

Tumor necrosis factor (TNF-alpha) was compared to immune interferon (IFN-gamma) for its ability to modulate the expression and shedding of HLA antigens, of intercellular adhesion molecule I (ICAM I) and of high-molecular-weight melanoma-associated antigen (HMW MAA) by a panel of melanoma cell lines. The latter included the melanoma cell line MeWo and its metastatic variant MeM 50-10, which display differential susceptibility to modulation of HLA class-II antigens by IFN-gamma and the cell lines SK-MEL-93-DX-2 and SK-MEL-93-DX-3, which originated from anatomically distinct metastases in patient DX. TNF-alpha enhanced the expression of HLA class-I antigens on all 7 melanoma cell lines tested, although to a lower extent than IFN-gamma and the combination of IFN-gamma and TNF-alpha. TNF-alpha displayed a differential effect on the expression of HLA class-II antigens by the 7 melanoma cell lines: it enhanced it on 3 out of the 4 cell lines with constitutive expression of HLA class-II antigens and induced them on 1 of 3 cell lines without detectable expression of these antigens. The effects of IFN-gamma were different since it enhanced HLA class-II antigens on the 4 cell lines with constitutive expression of these antigens and induced them on 2 out of the remaining 3 lines. Interestingly, both TNF-alpha and IFN-gamma enhanced the expression of HLA class-II antigens by SK-MEL-93-DX-3 cells. On the other hand only TNF-alpha induced the expression of HLA class-II antigens by MeWo cells and only IFN-gamma induced such expression by MeM 50-10 cells and by SK-MEL-93-DX-2 cells. The effect of the combination of TNF-alpha and IFN-gamma was similar to that of the individual cytokines. Both TNF-alpha and IFN-gamma displayed a differential effect on the expression of the gene products of the HLA-D region by the melanoma cell lines. Northern blot analysis with HLA-DR beta-, DQ beta- and DP beta-specific probes suggests that the modulation of HLA class-II antigens by both cytokines reflects transcriptional and post-transcriptional events. TNF-alpha enhanced the expression of ICAM-I on all the melanoma cell lines, although to a lower extent than IFN-gamma and the combination of IFN-gamma and TNF-alpha. Lastly, neither TNF-alpha nor IFN-gamma displayed a marked effect on the expression of HMW-MAA by the melanoma cell lines tested.

Functional activities of enriched Fc receptor bearing T lymphocytes in Hodgkin's disease.

Our earlier studies have demonstrated that the peripheral blood mononuclear cells (PBMNC) of patients with Hodgkin's disease (HD) have reduced percentage of T mu cells and increased percentage of T gamma cells, while the splenic MNC showed the reversed proportions. In this paper, the functional abilities of subsets with these phenotypes have been investigated. T gamma and T mu cells were enriched from theophylline-sensitive and -resistant populations of lymphocytes obtained from PBMNC and splenic MNC of untreated HD patients and normal healthy donors. The effect of addition of these enriched populations on PHA and mixed leukocyte culture (MLC) responses of corresponding autologous MNC has been studied. The results showed that both PHA and MLC responses of normal as well as of HD lymphocytes could be suppressed by T gamma cells, while T mu cells failed to show an appreciable helper activity. The T gamma population in HD, therefore, appears to maintain suppressor function, thereby influencing the peripheral blood T cell responses.

Effect of administration of BCG, levamisole and irradiated leukemic cells on immune status and remission status in chronic myelogenous leukemia.

The immunomodulating effects of bacillus Calmette-Guérin (BCG) and levamisole with and without irradiated autologous leukemic cells were tested in patients with chronic myeloid leukemia (CML) after control of white cell count with busulfan. The response has been evaluated with respect to remission period and in vivo and in vitro immunological studies comprising delayed cutaneous hypersensitivity response to recall antigens and dinitrochlorobenzene, T cell and its subsets percentages, T-cell response to phytohemagglutinin, and to leukemia cell extracts by blastogenesis and leukocyte migration inhibition. Patients receiving BCG or levamisole did show marginal prolongation of remission, however immune parameters failed to show any improvement. In contrast, improvements in specific and nonspecific immune parameters were observed in patients receiving BCG or levamisole along with irradiated leukemic cells, however, concomitant clinical benefit was not obtained. Development of a better immunotherapeutic approach appears essential for immunomodulation in CML.

Immunosuppressive activity of T cell clones generated from human T cells stimulated with autologous TPHA cells.

T cell clones were generated from human T cells stimulated with autologous phytohemagglutinin (PHA)-activated T (TPHA) cells. Characterization of three T cell clones originated from donor SF and one from donor JM showed that they proliferated when stimulated with autologous TPHA cells, non-T cells, and peripheral blood mononuclear cells, but did not proliferate when stimulated with allogeneic TPHA cells, non-T cells, and mononuclear cells, with autologous and allogeneic resting T cells, and with PHA. These results in conjunction with the blocking of the proliferation by anti-histocompatibility leukocyte antigen class II monoclonal antibodies indicate that these class II antigens are involved in the proliferation of T cell clones stimulated with autologous lymphoid cells. The four T cell clones are cytotoxic neither to autologous lymphoid cells nor to a panel of cultured human cell lines. The four T cell clones display immunosuppressive activity, since they inhibit the proliferation of autologous and allogeneic cells stimulated with antigens and mitogens and the secretion of immunoglobulin by B cells stimulated with pokeweed mitogen in presence of T cells. Furthermore, the four T cell clones display differential inhibitory activity on the proliferation of cultured human cell lines. The immunosuppressive activity is species-specific, since the T cell clones do not inhibit the proliferation of murine cells. The suppression is mediated by a factor(s) with an apparent m.w. of 13,000 to 16,000. The suppressor activity is labile at alkaline pH and is lost following incubation with pronase (100 U/ml) for 30 min at 37 degrees C.

Differential induction by immune interferon of the gene products of the HLA-D region on the melanoma cell line MeWo and its metastatic variant MeM 50-10.

This study has shown for the first time an association between the metastatic properties of two autologous melanoma cell lines and their susceptibility to induction of HLA class II Ag by IFN-gamma. After in vitro incubation with IFN-gamma the melanoma cell line MeWo did not acquire reactivity with anti-HLA class II antibodies, whereas its metastatic variant MeM 50-10 did. The differential susceptibility to induction of HLA class II Ag on the two cell lines cannot be accounted for by either differences in the number and affinity of IFN-gamma receptors or in the sensitivity to IFN-gamma, but most likely reflects an intrinsic property of each cell line. Serologic and immunochemical investigations with anti HLA-DR, DQ, and DP mAb have indicated that only HLA-DR Ag are induced by IFN-gamma on MeM 50-10 cells. Northern blot analysis with HLA-DR beta, DQ beta, and DP beta probes suggest that different mechanisms underlie the differential susceptibility to induction by IFN-gamma of the gene products of the HLA-D region. The regulatory mechanism(s) that control the expression of HLA class II Ag appear to be different from those controlling the expression of the melanoma-associated Ag tested, inasmuch as the modulation of the latter by IFN-gamma did not differ on the two melanoma cell lines.