Targeted protein degradation (TPD) has revolutionized drug discovery, with PROTACs leading the charge by catalytically eliminating disease-causing proteins. While conventional PROTACs degrade a single target, the complexity of diseases like cancer and neurodegeneration—marked by redundant and compensatory signaling networks—has spurred the rapid development of dual/multi-target PROTACs capable of simultaneously degrading two or more pathogenic proteins. This review systematically surveys recent advances (2023–2026) in this area, categorizing them into two classes: those targeting homologous proteins (e.g., CDKs, BCL-2/BCL-xL, HDACs, BAZ2A/BAZ2B) and those targeting distinct proteins across interconnected pathways (e.g., ERα/ARO, α-Syn/tau, BET/HDAC, PI3K/mTOR, FLT3/CHK1, CBP/BRD4, and others). For each, we critically analyze design strategies, structure–activity relationships, linker optimization, and E3 ligase selection. Despite remarkable progress—including the first dual degraders for non-kinase epigenetic regulators and protein aggregates—challenges persist in pharmacokinetics, off-target toxicity, and limited E3 ligase diversity. By consolidating key breakthroughs and practical SAR insights, this review provides a valuable resource for advancing next-generation multi-target degraders toward clinical translation for complex diseases.
Human rhinoviruses (HRVs) are major causative agents of the common cold and are strongly associated with asthma exacerbation, highlighting the need for effective antivirals. Because the viral 3 C protease (3Cpro) is essential for polyprotein processing and is highly conserved among HRV species, it represents an attractive therapeutic target. In this study, we performed structure-based virtual screening of 49,437 compounds against HRV-C 3Cpro using AutoDock Vina, followed by ADMET-based prioritization of the top hits. Docking analysis identified two lead compounds that favorably occupied the catalytic pocket and formed stabilizing interactions with key residues, including His40, Glu71, and Cys147, in comparison with the reference inhibitor rupintrivir. In 200-ns molecular dynamics simulations, both compounds stabilized the protease relative to the apo form, with compound 1 showing the lowest protein RMSD (1.21 ± 0.22 Å) and compound 2 producing the lowest Cys147 RMSF (0.48 Å). Both ligands also displayed lower ligand RMSD values than rupintrivir (compound 1: 1.49 ± 0.78 Å; compound 2: 2.01 ± 0.39 Å; rupintrivir: 4.83 ± 0.89 Å), indicating more persistent binding within the active site. MM/PBSA analysis further supported stronger effective binding for compound 1 (-23.59 ± 6.87 kcal/mol) and compound 2 (-25.25 ± 5.75 kcal/mol) than for rupintrivir (-19.43 ± 4.46 kcal/mol). Taken together, these results identify compounds 1 and 2 as promising non-covalent HRV 3Cpro inhibitor candidates for further experimental validation.
Aberrant activation of the Wnt/β-catenin signaling pathway is a hallmark of colorectal cancer, primarily driven by the interaction between β-catenin and T-cell factor 4 (Tcf4). Due to the large and shallow interface ( 4800 Å2) of this protein–protein interaction, it has historically been considered “undruggable.” However, emerging evidence highlights the presence of druggable hot spots within this interface. In this study, we employed a structure-based virtual screening approach to repurpose FDA-approved drugs targeting the β-catenin/Tcf4 interaction. Using physics-based molecular simulations and MM/GBSA binding free energy calculations, we identified Leucovorin (Folinic acid), Carbenicillin, and Ceforanide as promising candidate hit compounds with stable binding to critical hot spot residues (Asn430, Lys435, His470, Arg474, and Lys508) of β-catenin. Experimental validation using HT-29 colorectal cancer cells revealed that Carbenicillin and Ceforanide reduced the cell viability, impaired the colony formation, and exhibited superior anti-proliferative activity compared to the other candidates. These findings provide compelling evidence for the feasibility of targeting β-catenin/Tcf4 interactions using small molecules and suggest that Carbenicillin and Ceforanide may serve as promising repurposed candidates for therapeutic intervention in Wnt-driven malignancies.
An NH4I-mediated electrochemical three-component cascade reaction has been established for the preparation of trideuteromethylthiolated enaminones from primary amines, acetylacetone, and CD3SSO3Na. This simple protocol is readily applicable to gram-scale synthesis and late-stage modification of bioactive molecules. Preliminary biological screening of the prepared trideuteromethylthiolated enaminones identified compound 4l is capable of protecting cells from oxidative damage by initiating the activation of the Nrf2/HO-1 signaling pathway.
Matrix metalloproteinase−9 (MMP-9) plays a pivotal role in extracellular matrix remodelling and is strongly implicated in tumour progression, metastasis, and angiogenesis, making it an attractive therapeutic target for anticancer drug discovery. However, the development of potent and selective MMP-9 inhibitors has been significantly hampered due to the high structural homology among members of the MMP family, particularly within the catalytic domain. Structural studies revealed a non-conserved S1′ pocket of MMP-9 exhibits distinct structural variability compared to other MMP isoforms, providing an opportunity to achieve improved selectivity. To the best of our knowledge, fragment-based drug design (FBDD) approaches specifically targeting the S1′ pocket of MMP-9 have not been systematically explored. In the present study, an integrated in silico FBDD workflow was employed to identify and optimize small molecules occupying the S1′ pocket of MMP-9. Initial fragment-based virtual screening identified low-molecular-weight fragments with moderate binding affinity toward the target pocket. A promising fragment hit was subsequently optimised through a receptor cavity-guided fragment-growing strategy to extend interactions toward residues in the neighbouring S2′ pocket. Induced-fit docking analysis identified several optimized molecules with enhanced binding affinity and favourable orientation within the S1′ pocket. The stability of the protein-hit molecule complexes was further validated through 200 ns molecular dynamics simulations, which confirmed stable binding interactions for four molecules. Comparative docking and MM-GBSA analyses against MMP isoforms demonstrated a preferential affinity of hit molecules for MMP-9, suggesting a potential selectivity profile driven by ligand binding free energy and orientation in S1’ pocket. In addition, in silico ADME profiling indicated favourable pharmacokinetic properties of all the stable hit molecules, while density functional theory studies helped to gain insights into the electronic distribution and reactivity of the molecules. Collectively, this study establishes an in silico framework for the structure-based design of potentially selective MMP-9 inhibitors targeting the non-conserved S1′ pocket.
