Pharmacological targeting of nonsense mutant TP53 and PTEN in cancer

The TP53 tumor suppressor gene encodes p53 and is inactivated by mutations in around half of all human tumors. Approximately 11% of TP53 mutations are nonsense mutations, resulting in the premature termination of translation and the production of truncated and non-functional p53 proteins. Aminoglycosides such as G418 are known to induce translational readthrough, a process in which the ribosome overcomes the stop signal introduced by a nonsense mutation and translates full-length protein. However, the clinical use of aminoglycosides is restricted due to severe side effects. We have demonstrated that combination treatments with proteasome inhibitors or compounds that disrupt the binding of p53 to the ubiquitin ligase MDM2 can synergistically enhance the levels of fulllength p53, improving the efficacy of readthrough compared to aminoglycosides alone. These combinations were proven to produce at least partially active fulllength p53, as shown by the suppression of cell growth and the induction of cell death. In parallel, chemical library screenings led to the discovery of two novel compounds, C47 and C61, showing readthrough activity and synergizing with G418 and eRF3 degraders CC-885 and CC-90009, respectively. Remarkably, C47 also exhibit readthrough activity for nonsense mutant phosphatase and tensin homolog (PTEN), expanding the scope for targeted cancer therapies. Furthermore, we have identified the 5-fluorouracil (5-FU) metabolite 5-Fluorouridine (FUr) as a potent readthrough-inducing compounds capable restoring full-length p53 expression in cells harboring nonsense mutant TP53. In vivo studies further substantiated the capability of FUr to reinstate full-length p53 expression in human tumor xenografts with TP53 R213X nonsense mutations. Finally, the first Trp53 R210X nonsense mutant knock-in mouse model has been generated. R210X corresponds to human TP53 R213X. Observations on tumor development, lifespan and other phenotypic traits in these mice provide valuable insights into the impact of TP53 nonsense mutation in a multi-organ system. These results also provide a platform for the preclinical evaluation of novel therapeutic strategies for targeting nonsense mutant TP53. In summary, these findings offer a multi-faceted approach towards understanding TP53 nonsense mutations and advancing targeted cancer therapy through pharmacological induction of translational readthrough. The discovery of novel readthrough inducing compounds, the application of combination therapy in translational readthrough, the discovery of a novel therapeutic application for 5- FU and its metabolite FUr, as well as the generation of a novel animal model collectively set the stage for the further development of personalized treatments for patients with tumors harboring nonsense mutant TP53

Synergistic Rescue of Nonsense Mutant Tumor Suppressor p53 by Combination Treatment with Aminoglycosides and Mdm2 Inhibitors

The tumor suppressor gene TP53 is inactivated by mutation in a large fraction of human tumors. Around 10% of TP53 mutations are nonsense mutations that lead to premature termination of translation and expression of truncated unstable and non-functional p53 protein. Aminoglycosides G418 (geneticin) and gentamicin have been shown to induce translational readthrough and expression of full-length p53. However, aminoglycosides have severe side effects that limit their clinical use. Here, we show that combination treatment with a proteasome inhibitor or compounds that disrupt p53-Mdm2 binding can synergistically enhance levels of full-length p53 upon aminoglycoside-induced readthrough of R213X nonsense mutant p53. Full-length p53 expressed upon combination treatment is functionally active as assessed by upregulation of p53 target genes, suppression of cell growth, and induction of cell death. Thus, our results demonstrate that combination treatment with aminoglycosides and compounds that inhibit p53 degradation is synergistic and can provide significantly improved efficacy of readthrough when compared with aminoglycosides alone. This may have implications for future cancer therapy based on reactivation of nonsense mutant TP53

Chemical regulators of epithelial plasticity reveal a nuclear receptor pathway controlling myofibroblast differentiation

Plasticity in epithelial tissues relates to processes of embryonic development, tissue fibrosis and cancer progression. Pharmacological modulation of epithelial transitions during disease progression may thus be clinically useful. Using human keratinocytes and a robotic high-content imaging platform, we screened for chemical compounds that reverse transforming growth factor beta (TGF-beta)-induced epithelial-mesenchymal transition. In addition to TGF-beta receptor kinase inhibitors, we identified small molecule epithelial plasticity modulators including a naturally occurring hydroxysterol agonist of the liver X receptors (LXRs), members of the nuclear receptor transcription factor family. Endogenous and synthetic LXR agonists tested in diverse cell models blocked alpha-smooth muscle actin expression, myofibroblast differentiation and function. Agonist-dependent LXR activity or LXR overexpression in the absence of ligand counteracted TGF-beta-mediated myofibroblast terminal differentiation and collagen contraction. The protective effect of LXR agonists against TGF-beta-induced pro-fibrotic activity raises the possibility that anti-lipidogenic therapy may be relevant in fibrotic disorders and advanced cancer