Different mechanisms are involved in apoptosis induced by melanoma gangliosides on human monocyte-derived dendritic cells

Tumor escape is linked to multiple mechanisms, notably the liberation, by tumor cells, of soluble factors that inhibit the function of dendritic cells (DC). We have shown that melanoma gangliosides impair DC differentiation and induce their apoptosis. The present study was aimed to give insight into the mechanisms involved. DC apoptosis was independent of the catabolism of gangliosides since lactosylceramide did not induce cell death. Apoptosis induced by GM3 and GD3 gangliosides was not blocked by inhibitors of de novo ceramide biosynthesis, whereas the acid sphingomyelinase inhibitor desipramine only prevented apoptosis induced by GM3. Furthermore, our results suggest that DC apoptosis was triggered via caspase activation, and it was ROS dependent with GD3 ganglioside, suggesting that GM3 and GD3 induced apoptosis through different mechanisms

Production of Multiple Brain-Like Ganglioside Species Is Dispensable for Fas-Induced Apoptosis of Lymphoid Cells

Activation of an acid sphingomyelinase (aSMase) leading to a biosynthesis of GD3 disialoganglioside has been associated with Fas-induced apoptosis of lymphoid cells. The present study was undertaken to clarify the role of this enzyme in the generation of gangliosides during apoptosis triggered by Fas ligation. The issue was addressed by using aSMase-deficient and aSMase-corrected cell lines derived from Niemann-Pick disease (NPD) patients. Fas cross-linking elicited a rapid production of large amounts of complex a- and b-series species of gangliosides with a pattern and a chromatographic behavior as single bands reminiscent of brain gangliosides. The gangliosides were synthesized within the first ten minutes and completely disappeared within thirty minutes after stimulation. Noteworthy is the observation that GD3 was not the only ganglioside produced. The production of gangliosides and the onset of apoptotic hallmarks occurred similarly in both aSMase-deficient and aSMase-corrected NPD lymphoid cells, indicating that aSMase activation is not accountable for ganglioside generation. Hampering ganglioside production by inhibiting the key enzyme glucosylceramide synthase did not abrogate the apoptotic process. In addition, GM3 synthase-deficient lymphoid cells underwent Fas-induced apoptosis, suggesting that gangliosides are unlikely to play an indispensable role in transducing Fas-induced apoptosis of lymphoid cells

Induction of IgG antibodies directed to a M(r) 31,000 melanoma antigen in patients immunized with vaccinia virus melanoma oncolysates.

Pre- and postimmunization sera from eight tumor-free melanoma patients undergoing vaccinia melanoma oncolysate (VMO) therapy were used to investigate the humoral response to antigens from infected and uninfected melanoma cells and from vaccinia virus. Immunodetection on Western blots showed that all patients, in addition to reacting to several other proteins, developed IgG antibodies to a M(r) 31,000 protein antigen within 1 month of immunization. This M(r) 31,000 antigen is expressed both on VMO and on melanoma metastases in situ, disappears in primary cultures of these metastases, and is absent in extracts from vaccinia virus, from human melanoma cell lines, and from normal melanocytes, suggesting that this M(r) 31,000 protein is reexpressed following vaccinia virus infection of human melanoma cells. Periodate treatment of the blotted antigens abolished reactivity of patients' postimmunization sera with the M(r) 31,000 antigen, thus showing that this antigen is a glycoprotein and that the relevant epitope is likely to reside on its carbohydrate moiety. These anti-M(r) 31,000 IgG antibodies were absent in the sera of VMO-treated patients before immunization, absent in the serum of a normal donor hyperimmunized with vaccinia virus, and absent in normal human sera. In addition, these anti-M(r) 31,000 antibodies appeared 1 week after the first VMO injection, remained stable during the treatment, and decreased when the treatment was stopped. Such antibodies can also be demonstrated in sera of melanoma patients bearing metastases but disappeared following resection of their metastases. Thus, in melanoma patients, immunization with VMO induces an antibody response directed against a M(r) 31,000 glycoprotein likely to represent a new melanoma antigen. Further identification of this antigen could be of utmost interest for the further development of melanoma vaccines

Monoclonal antibodies to gamma-interferon treated LAN-1 cells detect modulation of ganglioside GD2 exposure on human neuroblastoma cells

A panel of 8 new Mabs have been produced against neuroblastoma cells (LAN-1) previously treated with IFN-gamma. All selected Mabs from 2 different fusions have been shown to detect epitopes on the GD2 ganglioside molecules highly expressed on all cells of neural crest origin including neuroblastoma, glioblastoma and melanoma. Our results imply that modulation of GD2 exposure on NB cells is dependent on culture conditions and moreover that IFN-gamma increases the surface expression of GD2 and thereby enhances their immunogenicity

New anti-GD2 monoclonal antibodies produced from gamma-interferon-treated neuroblastoma cells

Three monoclonal antibodies (IgG2) have been produced from hybridomas obtained by fusion of murine myeloma cells and spleen cells of mice hyperimmunized with gamma-interferon-treated neuroblastoma cells. The 3 MAbs, 7A4, 2A6 and IG8, detected an antigen present on neuroblastoma tumors and cell lines, but also on some neuro-ectoderm-derived tissues and cells. All 3 clones were shown to react with an epitope of the di-sialo-ganglioside GD2 molecules highly expressed by some neuro-ectoderm-derived tumors, mainly neuroblastoma. Whereas MAb IG8 specificity was restricted to GD2 and its o-acylated form, MAb 2A6 and 7A4 were also able to detect GD3 at high concentration of antibody as shown by TLC analysis and immunodetection. The 3 MAbs were able to lyse 100% neuroblastoma cells in the presence of rabbit or human complement. Direct binding assays with 125I-labelled MAbs showed that MAb 7A4 might be a good candidate for in vivo immunolocalization experiments. The high proportion of anti-GD2 MAbs obtained by our fusion and the increased binding of anti-GD2 MAbs on gamma-IFN-treated neuroblastoma cells suggests a modulation of the exposure and an increase in the immunogenicity of GD2 induced by gamma-IFN