Analysis of MHC-restricted tumor rejection antigen induced by IL-12.

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Genetic effects of human-tumor cells derived peptides were investigated. Because IL-12 gene transfected tumor cells highly expressed MHC class I molecules, IL-12 gene was transfected into human tumor derived cells. However, those transfected tumor cells did not grow well. This growth inhibitory activity depends on p35 gene which was a half part of IL-12 gene. So, parental tumor cells were used in the following experiments. Mass-culture of human tumor cells was done. After cell surface peptides were collected by pH3.4 citrate-phosphate buffer solution, crude samples were filtered by specific membrane systems. Low molecular peptides (M.W.1000〜6,500 Dalton) were extracted by HPLC system. One peak among multiple peaks obtained was added to normal human cells. After incubation for several hours, m-RNAs were extracted to examine what and how many genes were up- or down-regulated in eight thousand gene tested. The expression of about two hundred genes was significantly up- or down-regulated. The up-regulated genes were associated with tumor progression, in the other hand ; the down-regulated genes were associated with repression genes against anti-tumorigenic genes. As a result, the character of this peptide was consistent with the growth activity of malignant tumor cells. The investigation hereafter will be focused on the peptides sequences by mass spectrometer. It will be also necessary for analysis of the relations of these huge gene expressions to develop the new methods for multi-dimensional unification.