Abstract 5154: Proteolysis and gene silencing targeting chimera (PROGENTAC)

Abstract Introduction: In biological systems, nucleic acids mirror computational networks, integrating digital genetic codes with analog molecular interactions to achieve precise, context-dependent control of life processes. Harnessing this property provides an opportunity to design programmable therapeutics with exceptional specificity and minimal toxicity. Building on this inspiration, we devised a method for the conditional activation of a dual-functional nucleic acid construct comprising gene-silencing antisense oligonucleotide (ASO) and protein degrader, a DNA-based proteolysis-targeting chimera. We have coined the term “Proteolysis Gene Silencing Targeting Chimera” (PROGENTAC) to describe this technology. This dual functionality of conditional PROGENTAC offers several advantages, such as enhanced specificity and combating multidrug-resistant in complex diseases1. The molecular design of PROGENTAC consists of three elements: 1) DNA-templated, spatially controlled protein-degrading chimera (DTAC) platform2; 2) ASO for targeted gene modulation; and 3) rationally proposed conditional nucleic acid-based molecular framework that regulate PROGENTAC activity3 in response to disease-specific intracellular cues or chemical trigger. Experimental Procedures: In house solid phase oligo synthesis was utilized to construct both DTAC and modified ASO constructs. In house organic synthesis was utilized to modify small molecule-based inhibitors. Electrophoretic mobility shift assay (EMSA) was utilized to visualize conditional activation of PROGENTAC in cell-free based media. Fluorescence resonance energy transfer (FRET) assay was utilized to evaluate kinetics of PROGENTAC activation. Additional experiments were conducted including, Cell treatments, Western blots, Confocal imaging, Flow Cytometry, Proteomics, and Cytotoxicity assays. Results and Summary: Three cancer cell lines U251, A549 and AU565 showed excellent activity of DTAC that targets Cyclin D1-CDK4/6 complex protein with efficacy of DC50 of 20-100 nM range. For a proof of concept, we successfully constructed GFP protein silencing ASO construct with various nucleic acid modifications such as 2’-OMe, locked nucleic acid (LNA), and phosphorothioate (PS backbone). The optimal chemically modified potent ASO construct was successfully integrated with DTAC in PROGENTAC molecular framework. PROGENTAC construct was evaluated for structural stability and conditional activation to release both protein degrading DTAC and gene silencing ASO modalities. In the conference we will further talk about extended cellular results and therapeutic properties of PROGENTAC. Citation: 1) Dagogo-Jack, I. & Shaw, A. T. Nat Rev Clin Oncol 15, 81-94 (2018); 2) Zheng, L. et al. J. Am. Chem. Soc. 2025, 147, 33, 29742-29755; 3) Yan, H., Zhang, X., Shen, Z. et al. Nature 415, 62-65 (2002). Citation Format: Abhay Prasad, Rong Zheng, Deeksha Satyabola, Yang Xu, Yichen Yan, Hao Yan. Proteolysis and gene silencing targeting chimera (PROGENTAC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 5154.