) Many human carcinoma cell lines are regulate in their proliferation through autocrine growth regulation involving growth factors, e.g. epidermal growth factor (EGF) or transforming GF-alpha (TGF-alpha), and members of the EGF receptor (EGFR) receptor tyrosine kinases (RTK), Ionizing radiation (IR) in the therapeutic dose range activates EGFR and with repeated exposure induces up-regulation of EGFR, identifying EGFR as early and late IR response gene. A number of downstream components of EGFR-dependent signal transduction pathways are activated by IR including phospholipase C, stimulation of p42/44 MAPkinase, and transcriptional up-regulation of c- Fos. The significance of this activation is indicated by the fact that single or repeated IR exposures can induce a measurable proliferation response in mammary (MCC) and squamous carcinoma cells (SCC), As IR- induced proliferation responses to repeated IR exposures have been implicated in counteracting the toxic effects of IR disruption of this response should result in improve radiotherapeutic outcomes. Disruption of the IR-induced proliferation through genetic modulation of EGFR expression is the overall objective of the proposed experimentation and will accomplished through the following specific aims: (1) Generation of additional MCC and SCC cells expressing inducible mitogenesis-deficient (MUT) or antisense (AS) EGFR and their characterization with respect to growth and expression profile of EGF/TGF-alpha and other erbB RTK; (2) Mechanistic studies on the effects of EGFR-MUT/-AS expression on IR- induced proliferation and stress pathways and their manipulation through Adenovirus (Ad)-mediated transient transfections in order to shift he balance of proliferation and stress responses with one endpoint of apoptosis; (3) Correlation of studies under Specific Aim 2 with overall cellular responses of IR-induced proliferation, clonogenic survival, and apoptotic cell death; (4) As a first step toward the intervention of the IR-induced proliferation responses through transient transfections of EGFR-MUT/-AS, proven under Specific Aim 1-3, using an AD infection system including characterization of those with respect to signal transduction and overall cellular responses.
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