The GOF Mutant p53 Beacon System

项目总结

Project Summary We have developed a system in which Gain-Of-Function (GOF) mutant p53 makes living cells fluoresce and luminesce (GOF Mutant p53 Beacon System). This powerful tool allows us to instantly assess the presence or absence of mutant p53 in living cells and is not affected by wild-type p53. It also provides a means to assess activity and properties for a large variety of mutant p53 alleles in cell lines. We propose to test the utility of the Beacon System for assessing small molecule mutant p53 inhibitors and wild-type p53 reactivators. We will adapt the Beacon System for use in high throughput screening of inhibitors such that when mutant p53 is inhibited, the cells will fluoresce and luminesce. The utility of the system for identifying cellular modulators of mutant p53 will also be investigated by adapting the system so that when mutant p53 is inhibited, the cells become resistant to puromycin. The ultimate phase of developing this tool is to make transgenic mice in which cells that acquire p53 mutations fluoresce and luminesce. This advance should allow us to visualize any tumor growth in the living mouse as cells acquire p53 mutations and as the tumor cells grow, evolve, invade, and metastasize and establish glowing cell populations at new locations. The mouse system will also be adapted such that cells that acquire p53 mutations will express a cell surface streptavidin tag making it easy to purify the cells at various stages of cancer development to study cellular properties, stem-cell properties, genetic events, and the evolution of these events at early to late stages of cancer. Development of this system should create opportunities for a simple and rapid screening for cancer therapeutics that target mutant p53, one of the most prevalent cancer-causing dominant defects in all cancers. A mouse in which the tumor glows in the living mouse will provide a powerful real-time analysis of cancer progression. This tool will advance our knowledge of how tumors grow and tumor cells migrate, and how they respond to therapeutics.