The movement of DNA from the cytoplasm to the nucleus remains one of the major barriers to efficient gene transfer and expression. Without localization of DNA to the nucleus, no transcription or """"""""gene therapy"""""""" can take place. Based on our studies on intracellular trafficking of nanoparticles, we have recently discovered surface modified NPs that are taken up by cells via non-endocytic pathway, have significantly greater intracellular uptake and retention than unmodified NPs, and more importantly, these nanoparticles target to the nucleus. Our objective is to test the hypothesis that the direct nuclear delivery of modified nanoparticles would deliver the encapsulated DNA directly to the nucleus that would also enhance the level of gene expression. Our goal is to investigate the molecular mechanism of nuclear localization of modified nanoparticles and determine their efficacy as a non-viral gene expression vector. To test our hypothesis, we will use wild-type (wf)-p53 gene, a tumor suppressor gene as a model therapeutic gene, and cancer as a model pathologic condition. We propose that the nanoparticle-mediated sustained expression of wf-p53 gene in the tumor tissue would lead effective regression of tumor via induction of cell apoptosis and/or over expression of anti-angiogenic factors.
The specific aims of the proposal are: i) To investigate the formulation determinants that are critical to enhancing and sustaining the level of gene expression; ii) To investigate the molecular mechanism of nuclear-localization of modified nanoparticles; iii) To determine the therapeutic efficacy of wf-p53 gene encapsulated in modified nanoparticles in prostate cancer cell line; iv) To study the pharmacokinetics of biodistribution of modified nanoparticles via intravenous administration to achieve tumor-specific gene delivery; and v) To evaluate the efficacy of modified nanoparticles using wf-p53 gene in a murine model of prostate cancer. We anticipate that these nanoparticles can be used as an effective non-viral sustained gene expression system. Efficient nuclear delivery of nanoparticles could have far reaching implications, especially for delivery of drugs and proteins directly to the nucleus, both for therapeutic purposes and as well as to study their cellular and biochemical functions.
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