Targeting the PI3K/AKT Pathway using Pan-AKT Degraders

Abstract

The serine/threonine protein kinase AKT, a downstream effector of phosphoinositide 3-kinase (PI3K), is a critical regulator of numerous cellular processes including proliferation, survival, metabolism, and apoptosis. Genetic aberrations in PI3K pathway components are prevalent across cancer types, resulting in AKT hyperactivation. Thus, AKT represents a rational and appealing therapeutic target. The AKT family of kinases is comprised of three highly homologous isoforms, AKT1, AKT2, and AKT3. Several ATP-competitive, allosteric, and covalent pan-AKT inhibitors have been developed, but thus far have displayed limited therapeutic benefit in clinical trials, due to lack of efficacy at tolerable concentrations and development of resistance. To effectively target hyperactive AKT signaling for cancer treatment, new modalities are needed that address these challenges. Recently, targeted protein degradation has been developed as an alternative strategy to inhibition. This thesis reports the development and extensive characterization of two heterobifunctional degraders that selectively deplete AKT1, AKT2, and AKT3 in vitro and in vivo. These compounds, INY-03-041 and INY-05-040, comprise the catalytic AKT inhibitor GDC-0068 (ipatasertib) chemically conjugated to ligands for Cereblon or Von Hippel Lindau (VHL) ubiquitin ligases. Both degraders demonstrated improved selectivity and potency compared to GDC-0068. AKT degradation was rapid and unexpectedly long-lasting, even after compound washout, and maintained suppression of downstream signaling. AKT degraders demonstrated more potent anti-proliferative effects than catalytic AKT inhibition in cancer cells in vitro and induced degradation of AKT in vivo. AKT degraders were further used to investigate AKT biology with the goal of delineating non-kinase dependent roles of AKT. Using a multi-omics approach to examine the metabolic and transcriptional consequences of AKT degradation versus inhibition, we identified and validated activation of stress kinase signaling as a cellular response specific to AKT degradation. Degrader-specific induction of stress kinases was validated by immunoblotting, and inhibition of the stress kinase JNK rescued AKT degrader-specific cell death. Together, these data support the use of AKT degraders as novel tools to decode the pleiotropic mechanisms that govern AKT signaling in human cancer, and as potential cancer therapeutics.