The long-term objective of this work is to determine if the constitutive activation of the AP-1 transcriptional complex is one mechanism for how tumor cells gain resistance to the cytotoxicity of hyperthermic radiosensitization. In addition, we will determine if the upstream signaling factor, p38, is a target to both inhibit the AP-1 transcriptional complex and inhibit the resistance to hyperthermic radiosensitization. This mechanistic information is potentially important to improving tumor cell responses to heat-induced radiosensitization, which is currently being evaluated as a therapeutic modality. Preliminary results suggest that: (1) heat shock activates AP-1 DNA-binding activity via a mechanism involving the redox sensitive signaling factor Ref-1 in HeLa and NIH 3T3 cells; (2) OC-14 peroxide resistant cells overexpress c-Fos, c-Jun, and exhibit elevated AP-1 DNA activity when compared to their parent cell line, HA-1; (3) OC-14 cells are resistant to the cytotoxicity from hyperthermic radiosensitization; (4) indomethacin inhibits both the increase in AP-1 DNA-binding activity and the resistance to hyperthermic radiosensitization in OC-14 cells; and (5) a specific chemical that inhibits p38, SB20358 abolishes the effect of indomethacin and restores the increase in AP-1 DNA-binding activity and the resistance to hyperthermic radiosensitization in OC-14 cells. There is considerable evidence that transient activation of specific cytoplasmic signaling and nuclear transcription factors may play a protective role in the tumor cell response to therapeutic modalities. Thus, the proposed work will test the hypothesis that one mechanism for resistance to hyperthermic radiosensitization involves AP-1 transcription complex activation. In addition, it will be determined if p38 is an upstream signaling factor involved in both the activation of AP-1 and resistance to hyperthermic radiosensitization.
The specific aims are: 1) determine if two human H202 resistance tumor cell lines are also resistant to hyperthermic radiosensitization and if the AP-1 complex is also constitutively activated, similar to OC-14 cells; 2) determine if AP-1 constitutive activity, using permanent transfected HeLa cell lines that overexpress c-Fos/c-Jun wild-type and dominant positive proteins, results in a phenotype similar to OC-14 cells; 3) determine if the mechanism of indomethacin-induced inhibition of AP-1 DNA-binding activity is due to inhibition of gene expression, mRNA stabilization, protein degradation, or alteration of the activity of the basic DNA-binding domain of c-fos/c-jun, 4) determine if mitogen-activating kinase p38 is a target molecule for indomethacin induced inhibition of AP-1 activity and a target for inhibition of the resistance of OC-14 cells to hyperthermic radiosensitization using either a dominant negative p38 vector or a constitutive dominant positive upstream activator of p38, MKK6b.
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| Locke, J; Zeug, A; Thompson Jr, D et al. (2005) Localized versus regional hyperthermia: comparison of xenotransplants treated with a small animal ultrasound system and waterbath limb immersion. Int J Hyperthermia 21:271-81 |
