NF-kB is a critical nuclear transcriptional factor that is activated in response to cellular stresses and regulates the expression of genes involved in cell proliferation and cell death. When regulated NF-kB activation is disrupted, cells undergo apoptosis. These mechanisms have been demonstrated experimentally by expressing dominant negative IkB-a in cancer cells exposed to chemotherapeutic agents or to ionizing radiation. Furthermore, inhibition of NF-kB activation results in enhanced radiation sensitivity, and our recent studies have demonstrated that a radiation inducible signaling pathway includes both, NF-kB and ATM as critical components in AT cells. We propose to better understand mechanisms by which NF-kB function may be disrupted in order to achieve enhanced tumor cell kill. The goal of this proposal is to test the hypothesis that the signaling pathway involving ATM/NF-kB/IkB offers potential molecular targets for radiation sensitization strategies for treating human cancers.
In Specific Aim 1 we propose to use anti-sense (AS) RNA or AS DNA strategies in vivo to disrupt NF-kB signaling by interfering with p65, IkB-a or IKK expression. This approach should validate potential targets for clinically translatable radiation sensitization trategies. Our experimental plan includes (1) production of AS RNA expressing vectors and AS oligodeoxynucleotides targeting functional domains of genes of interest, (2) establishment of optimal conditions for gene transfer to inhibit target molecule and (3) determination of cellular effects of target inactivation. We will use liposomal encapsulation of gene targeting agents (e.g. AS-ODN to p65) to perform preclinical experiments with a human xenograft in mice in Specific Aim 2. Tumor growth inhibition, NF-kB expression and cellular apoptosis will be measured. The completion of the proposed studies will provide additional insight into ATM/ NF-kB/IkB mediated signaling and will identify molecular targets in this pathway for radiation sensitization of cancers.
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