Recent advancements in targeted protein knockdown technologies-emerging paradigms for targeted therapy

Abstract

A generalized therapeutic strategy for various disease conditions, including cancer, is to deplete or inactivate harmful protein targets. Various forms of protein or gene silencing molecules, e.g., small molecule inhibitors, RNA interference (RNAi), and microRNAs (miRNAs) have been used against druggable targets. Over the past few years, targeted protein degradation (TPD) approaches have been developed for direct degradation of candidate proteins. Among the TPD approaches, proteolysis targeting chimeras (PROTACs) have emerged as one of the most promising approaches for the selective elimination of proteins via the ubiquitin-proteasome system. Other than PROTACs, TPD methods with potential therapeutic use include intrabody-mediated protein knockdown and tripartite motif-21 (TRIM-21) mediated TRIM-Away. In this review, protein knockdown approaches, their modes of action, and their advantages over conventional gene knockdown approaches are summarized. In cancers, disease-associated protein functions are often executed by specific post-translational modifications (PTMs). The role of TRIM-Away is highlighted in the direct knockdown of PTM forms of target proteins. Moreover, the application challenges and the prospective clinical use of TPD approaches in various diseases are also discussed.

Keywords: Protein knockdown; cancer; intrabodies; post-translational modification; proteolysis targeting chimeras; tripartite motif-21.

Figure 1

Systematic representation of PROTAC-mediated protein knockdown. The first step is the synthesis of PROTAC molecule, which includes a ligand for the protein of interest (POI) and ubiquitin E3 enzyme. In the next step, the PROTAC molecule binds with POI and E3 ligase to form a ternary complex, which eventually leads to polyubiquitination and subsequent degradation of POI via the ubiquitination pathway. Created with BioRender.com

Figure 2

Timeline of advancement in PROTAC technology. MetAP2 [17]; FKBP12 [18, 19]; PI3K, FRS2α [20]; BCL-XL [25]. SGK3: glycose synthase kinase-3; VPS34: vascular protein sorting-34 [26, 27]; AR: androgen receptor; E2F: E2 transcription factor; RBPs: ribosome binding proteins [28]; P62: protein 62 [29]

Figure 3

Schematic representation for TRIM mediated protein knockdown of membranous protein HER2 and HER3, and specific PTM forms of cytosolic protein STAT3. 1. Transfection of mAbs specific for targeted protein in breast cancer-TRIM-21 overexpressing cells; 2. formation of TRIM-21-antibody-protein complex; 3. proteasomal degradation of targeted proteins; 4. knockdown of HER2, HER3 and STAT3 protein leads to inactivation of the downstream signaling respectively. P: phosphorylation