The nuclear matrix is important for defining nuclear structure and regulating metabolic processes such as DNA and RNA synthesis, and possibly DNA repair. We propose to determine whether the nuclear matrix is a critical structure involved in the manifestation of thermal damage and heat-radiosensitization. We will test the hypotheses that alterations in nuclear matrix proteins are related to heat-induced alterations in (or recovery of) nuclear matrix structure are correlated with cytotoxicity. Using antibodies against specific nucleic matrix proteins, we will quantitate heat-induced changes in nuclear lamin and interior matrix proteins, we will quantitate heat-induced changes in nuclear lamina nd interior matrix proteins, we will quantitate heat-induced changes in nuclear lamin and interior matrix proteins in situ using flow cytometry to determine the relative abundance of these proteins throughout the cell cycle. Structural changes in fixed cells will be evaluated with confocal microscopy and correlated with change in chromatin distribution and survival. Since changes ina the nuclear matrix might be expected to contribute to inhibition of DNA strand break repair observed during heat- radiosensitization, we will also determine temporal and spatial patterns of DNA repair in heated, irradiated cells. Flow cytometric quantitation of changes in nuclear matrix proteins may have clinical relevance as a predictive assay for response to anticancer treatments such as hyperthermia, since our preliminary data suggests that heat causes solubilization of lamin B in apoptotic an necrotic cells within hours after heating. Solubilization of nuclear matrix proteins could prove to be a rapid alternative to colony-forming assays for predicting heat sensitivity. Elucidation of the mechanisms of action of hyperthermia, alone or combined with radiation, may lead to optimization of cancer treatments and the preferential eradication of tumor cells.
