New cancer therapy by degradation of specific proteins

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We proposed to induce to degradation of specific protein by the modification of ubiquitin-ligase, Recently, it is reported that expression level of many proteins are regulated by not only transcription but degradation rate. Ubiquitin-porteasome system is one of the most useful system to regulate expression level of protein because of its specificity. On this unique point we try to create new ubiquitin-ligase to recognize and induce to degradate proteins which are toxic for our bodies.In this project, we used oncoprotein, Myc as model protein. Because Myc is known to bind to Max, we made several chimeric proteins, Max/Nedd4, Max/b-TrCP1, Max/CHIP. Nedd-4 is a HECT-type ubiquitin-ligase. b-TrCP1 is a F-box proteins which is a component of SCF-type ubiquitin-ligase. CHIP is a U-box type ubiquitin-ligase. It was hypothesized that Myc would be bound and ubiquitylated by these chmeric proteins.We confirmed that this artificial protein Max/CHIP bound to Myc in vitro system and also immunoprecipitation assay in vivo. The half-life of Myc reduced by overexpression of Max/CHIP protein in mammalian cells by expression vector most efficiently.Plaque assays and colony-formation assay shown that Myc-induced transformation is inhibited by Max/CHIP chimeric protein expression.Now, we observe the effect on whole bode. We implanted tumor cells which overexpressed Max/CHIP chimeric protein in Nude mouse to examine the effect of tumorigenesis.