Programmable protein degraders enable selective knockdown of pathogenic β-catenin subpopulations in vitro and in vivo

• Published in: bioRxiv
• Main Authors: Ye, Tina, Azmain Alamgir, Robertus, Cara, Colina, Darianna, Connor Monticello, Thomas Connor Donahue, Hong, Lauren, Vincoff, Sophia, Goel, Shrey, Fekkes, Peter, Luis Miguel Camargo, Lam, Kieu, Heyes, James, Putnam, David, Alabi, Christopher A, Chatterjee, Pranam, Delisa, Matthew P
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 11.11.2024
• Subjects: C-Terminus; Cell proliferation; Cell survival; CHIP protein; Colorectal carcinoma; Cytosol; Design; Hsp70 protein; Intracellular signalling; Malignancy; Nanoparticles; Peptides; Proteasomes; Proteins; Transcription activation; Ubiquitin-protein ligase; Wnt protein; β-Catenin
• DOI: 10.1101/2024.11.10.622803

Aberrant activation of Wnt signaling results in unregulated accumulation of cytosolic β-catenin, which subsequently enters the nucleus and promotes transcription of genes that contribute to cellular proliferation and malignancy. Here, we sought to eliminate pathogenic β-catenin from the cytosol using designer ubiquibodies (uAbs), chimeric proteins composed of an E3 ubiquitin ligase and a target-binding domain that redirect intracellular proteins to the proteasome for degradation. To accelerate uAb development, we leveraged a protein language model (pLM)-driven algorithm called SaLT&PepPr to computationally design "guide" peptides with affinity for β-catenin, which were subsequently fused to the catalytic domain of a human E3 called C-terminus of Hsp70-interacting protein (CHIP). Expression of the resulting peptide-guided uAbs in colorectal cancer cells led to the identification of several designs that significantly reduced the abnormally stable pool of free β-catenin in the cytosol and nucleus while preserving the normal membrane-associated subpopulation. This selective knockdown of pathogenic β-catenin suppressed Wnt/β-catenin signaling and impaired tumor cell survival and proliferation. Furthermore, one of the best degraders selectively β-catenin levels in livers of BALB/C mice following delivery as a lipid nanoparticle (LNP)-encapsulated mRNA. Collectively, these findings reveal the unique ability of uAbs to selectively eradicate abnormal proteins in vitro and in vivo and open the door to peptide-programmable biologic modulators of other disease-causing proteins.Competing Interest StatementM.P.D. and P.C. have financial interests in UbiquiTx, Inc. M.P.D. also has financial interests in Gauntlet, Inc. Glycobia, Inc., Resilience, Inc. and Versatope Therapeutics, Inc. M.P.D.'s and P.C.'s interests are reviewed and managed by Cornell University and Duke University, respectively, in accordance with their conflict-of-interest policies. All other authors declare no competing interests.