Project 3: Characterization of Beta-Catenin Degraders for the Treatment of Colorectal Cancer

PROJECT SUMMARY/ABSTRACT: Project 3 The long-term objective of this study is to investigate how a new class of compounds, β-catenin degraders, can be exploited to target Wnt/β-catenin (henceforth Wnt)-driven colorectal cancers (CRCs). Aberrant activation of the Wnt pathway drives the initiation and progression of nearly all CRCs. To date, no drugs inhibiting the Wnt pathway have been FDA-approved due to a lack of druggable targets that can bypass common Wnt pathway mutations in CRC. In a conceptual breakthrough, using an NMR-based fragment screen followed by X-ray guided medicinal chemistry with support from NCI’s Chemical Biology Consortium, we have discovered small molecules that bind with nanomolar affinity to β-catenin, the central regulator of the Wnt pathway. These compounds were used to create chimeric small molecule-based PROTACs that degrade β-catenin in CRC cells with pathway- activating mutations. We have shown that β-catenin degradation occurs via the ubiquitin-proteasome pathway and that the PROTAC degraders can inhibit the expression of Wnt target genes. Furthermore, this class of compounds inhibits the growth of both CRC cells and spheroids from a β-catenin mutant CRC cell line. These highly potent PROTACs also degraded β-catenin and inhibited Wnt pathway signaling in vivo, using CRC cell line xenografts in nude mice. We thus demonstrate that β-catenin PROTACs may provide a new means for controlling aberrant Wnt signaling and lay a foundation for the discovery of a novel treatment for nearly all CRCs. This project aims to use cutting-edge sophisticated in vitro, ex vivo, and in vivo approaches to better understand the effects of β-catenin degradation in CRC, thereby enabling their clinical translation. We propose to characterize the mechanism of action of the β-catenin PROTACs in CRC, determine the therapeutic window and potential toxicities associated with β-catenin degradation, and provide proof of principle for the effectiveness of β-catenin PROTACs in CRC patient-derived organoids (PDOs), CRC cell line xenografts, and a stem cell-driven mouse model of colorectal neoplasia. We propose three specific aims: 1) test the sensitivity of CRC cell monolayers and spheroids to β-catenin PROTACs; 2) determine the extent of PROTAC-induced degradation of E-cadherin-associated β-catenin; and 3) examine the effects of PROTACs in CRC cell line xenografts and Lrig1CreERT2;Apcflox/+ mice. At the end of this funding period, we will have fully characterized high-quality β-catenin PROTAC drug candidates, enabling their progression to IND-enabling studies.