Immune selection in murine tumors. Ph.d thesis.APMIS Suppl 2003; (106):1-46AS
It must be assumed that all tumor cells produce proteins which do not belong to a normal cell. These are called tumor-associated or tumor-specific antigens. In the classic immune surveillance theory it is believed that the cellular immune defense (the T-cell system) continuously discovers and eliminates newly arisen tumor cells which express such tumor-specific antigens. Since then it has been shown that one of the preconditions for the T-cell system to be able to recognize antigens is that they are presented by MHC class I histocompatibility antigens. There is a continual processing and presentation of all intracellular proteins in a cell. Thus, a tumor cell which produces an abnormal protein will also present this and thereby expose itself to being killed by cytotoxic T cells. The antigens are presented in the form of short peptides (8-9 aminoacids), which arise as a result of controlled degradation of the original proteins. The peptides thus formed are transported by specialised molecules in the so-called endogenous antigen processing and presentation pathway, and are eventually bound to and presented by MHC class I molecules. It has been shown that many tumors express less MHC class I on their surface compared to the normal tissue from which they have arisen, and also that patients with reduced immune function have an increased incidence of certain forms of cancer. It is therefore widely believed that a low MHC class I level contributes to the ability of tumor cells to avoid the T-cell-mediated immune defense. The aim of the present research project was to confirm the existence of a T-cell-mediated immune selection in primary tumors. Another of its goals was to elucidate the extent to which tumor cells with low MHC class I expression showed poor ability to present antigen, and whether the reason for this could be found in one or more of the molecular systems which participate in antigen processing and presentation. By using the chemical carcinogen 3-methylcholanthrene a total of 144 tumors were induced in immunologically normal and T-cell defective mice, respectively. It was assumed that tumors induced in normal mice would be immune selected, whilst this would not be the case for tumors from T-cell defective mice. This enabled us to work with a tumor-material where the two populations only differed in that the one part had undergone selection by a T-cell system and the other had not. Tumor induction time turned out to be shorter in immune defective than in normal mice, and the tumor frequency was higher, which might be due to the fact that in normal mice tumor growth was inhibited and in certain cases stopped by the T cells. On transplantation of the uncloned cell lines which were established from the primary tumors to immunologically normal congenic recipients, we were able to show that most of the tumors which originated from mice with a functional T-cell system, and which must therefore be assumed to have undergone selection in the primary tumor host, were not immunogenic and were therefore accepted. On the other hand, most tumors which originated from T-cell-defective mice were rejected as a sign that immunogenic tumor cells, assumed to have expressed tumor antigen, had not been eliminated in the primary tumor host. Still, we found that the ability of tumor cells to induce an immune response on transplantation was not reflected in their MHC class I expression. Both tumor lines from immunodeficient and normal mice had highly varying MHC class I levels, and contrary to expectations the highest levels were seen in tumor lines from immunologically normal mice. At the same time we found that the expression levels for the three different MHC class I molecules were the same in the individual tumor lines, which might indicate that the three genes are syn-regulated. The MHC class I mRNA content in tumors from normal mice was generally concordant with the surface level of MHC protein. Among the tumor lines from immunodeficient mice, on the other hand, we found several where there was no such agreement, which was taken to indicate that tumor cells with deviant MHC class I gene transcription had not been eliminated, in contrast to in the immunocompetent tumor hosts. The ability of tumor cells to present antigen was investigated by infecting cells with virus and thereafter assessing their ability to function as target cells for virus-specific T cells in a cytotoxic test. Their ability to do this varied considerably, but showed a correlation with their MHC class I expression. Among the transplanted tumor lines that were not able to present viral antigen, the majority were accepted, while most of the tumor lines which were rejected on transplantation possessed the ability to present virus. Closer analysis of the composition of proteasomes, heat shock protein content and TAP molecule function, which are all involved in the antigen processing system, did not immediately reveal any defects. Treatment with interferon gamma, which is known to upregulate the transcription of MHC class I and a number of other proteins which are involved in antigen presentation, showed that by far the majority of the tumor lines were able to respond normally. This was also true for the tumor lines which had deviant MHC class I gene transcription and the cells which showed poor ability to present viral antigen. We did find, however, three cell lines which did not respond to interferon gamma, and they all had defective interferon gamma-signaling, not because they did not express the interferon-receptor on the surface, but possibly on account of their lacking phosphorylation of an intracellular signal molecule, Stat1.