Project Summary / Abstract Our proposal focuses on the development of novel Mdm2-targeted proteolysis targeting chimeras (PROTACs) that efficiently degrade Mdm2 in p53 mutant and deficient cancer cells and are expected to target p53- independent functions of Mdm2. Our lead compounds bind with high affinity, degrade Mdm2, kill cancer cells that lack functional p53, and are efficacious in vivo. We have also demonstrated safety and tolerability in vivo, synthetic tractability, metabolic stability, and suitable in vivo exposure in mouse pharmacokinetic studies for in vivo efficacy evaluation. The tumor suppressor p53, a transcription factor, has an essential role in the prevention of human cancer. In the absence of cellular stress, the p53 protein is maintained at low levels due to its binding to Mdm2, an E3 ubiquitin ligase. However, half of all human cancers have inactivated p53 by mutation or deletion. To test if Mdm2 is required for the survival of cancer cells that lack p53 we inducibly deleted Mdm2 in primary murine p53-null lymphoma and sarcoma cells. Mdm2 loss resulted in apoptosis in vitro and in vivo in p53-null cancers, with significantly reduced tumor burden and increased survival benefit. We and others have reported that Mdm2 has p53-independent functions by binding and regulating other proteins, such as the p53 family member, p73, and Nbs1 in the Mre11-Rad50-Nbs1 DNA break repair complex. p73 has a homologous N-terminal transactivation domain to p53 which binds in the same site on Mdm2. Additionally, unlike p53, p73 is rarely inactivated in human cancers. Since Mdm2-p53 inhibitors are not responsive to tumors with inactivated p53, we pursued a PROTAC approach and provide preliminary data to confirm their ability to kill p53 mutated or deficient cancer cells. Our hypothesis is that the degradation of Mdm2 via Mdm2-targeted PROTACs will kill cancers with mutant or deleted p53 by activating the p53-independent activities of Mdm2. We will test this hypothesis with two Specific Aims. Aim 1 will focus on expanding the characterization of our lead Mdm2 targeting compounds and evaluate these in in vivo xenograft models. Aim 2 will be lead optimization of two Mdm2 PROTACs that killed p53 mutant and deleted cancers. The goal of these aims is to have a characterized, effective, and potent Mdm2 PROTAC for clinical evaluation for the treatment of p53-inactivated cancers.

Narrative The tumor suppressor p53 is essential in the prevention of human cancers, and its loss of function leads to tumor initiation and progression. Half of all human cancers have inactivated p53 by mutation or deletion, which results in aberrant aggressive growth. Our proposal focuses on the development of novel Mdm2 degraders to activate p53-independent activities of Mdm2 as an improved therapeutic approach against these aggressive tumors.