2026-06-25 | Biophysical and Biochemical Assays for Screening Small Molecule Inhibitors Targeting Toxin-Ribosome Interactions.
Ribosome-inactivating proteins are a class of toxins that target eukaryotic ribosomes, inhibit protein synthesis, and ultimately induce cell death. Several of these toxins pose significant clinical and public health threats. Among these, ricin, derived from the castor bean plant (Ricinus communis), is a highly potent biotoxin with recognized bioterrorism potential. Other ribosome-inactivating proteins, including Shiga toxin produced by pathogenic Shigella and Escherichia coli, as well as mucoricin from Mucorales fungi, contribute to disease severity and can lead to life-threatening complications. Despite these risks, no approved therapeutics are currently available. The development of effective inhibitors depends on robust and well-defined strategies to identify and validate small molecules that disrupt toxin-ribosome interactions. Efforts to target the catalytic active site have met with limited success, largely due to its broad, shallow, and highly polar architecture, which is not conducive to high-affinity binding by drug-like molecules. In contrast, the ribosome-binding interface represents a more tractable target, as it is essential for toxin recruitment and offers more structurally defined and druggable features. Inhibitors targeting this interface can also exert allosteric effects by disrupting long-range conformational coupling between the ribosome-binding region and the active site, thereby attenuating catalytic activity without directly engaging the catalytic pocket. In this review, we compile and evaluate biophysical and biochemical assays for the discovery and characterization of small-molecule inhibitors that target toxin-ribosome interactions. We examine in vitro binding approaches, including surface plasmon resonance-based fragment screening and fluorescence anisotropy assays for ranking inhibitory activity. We further review biochemical and molecular assays that assess ribosome protection from toxin-mediated depurination, along with complementary cell-based assays that evaluate functional rescue in cellular systems. Collectively, this review consolidates current screening methodologies and highlights opportunities to refine assay strategies, thereby supporting the advancement of targeted therapeutics.
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2026-03-29 | Advancing subunit vaccine efficacy against ricin toxin with a novel DNA hydrogel adjuvant tandem assembly by CpG motifs.
The development of effective countermeasures against ricin toxin, a potent biological threat, remains a pressing public health challenge. Subunit vaccines based on the ricin toxin A subunit offer a favorable safety profile but often suffer from limited immunogenicity, necessitating advanced adjuvants to elicit protective immunity. Here, we report a novel injectable CpG-motif-based hydrogel that serves as both adjuvant and delivery vehicle for a mutated and truncated RTA antigen. This CpG-gel adjuvant, synthesized via rolling circle amplification, incorporated with guanine-rich sequences that could self-assemble into stable G-quadruplex networks to form a structurally robust scaffold with flower-like microarchitecture. Notably, the CpG-gel significantly outperformed conventional CpG 1826 by enhancing antigen persistence and promoting sustained immune activation, leading to more potent and durable humoral and cellular responses. In murine studies, the CpG-gel elicited robust high titers of antigen-specific IgG antibodies, achieved complete protection against lethal-dose RT challenge. These results demonstrated that the CpG-gel adjuvant could significantly enhances the immunogenicity of subunit vaccines, highlighting its potential for broad implications for next-generation vaccine development.
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2026-03-27 | An innovative ricin-degrading antidote based on aptamer-autophagy-tethering compound strategy.
Ricin is classified as a Category B biothreat agent due to its high toxicity and wide availability, posing a substantial threat to public security. Currently, no effective antidotes have been approved, and the development of medical countermeasures-including small-molecule inhibitors, antibodies, and vaccines-face substantial challenges. In this study, we designed innovative ricin-degrading antidotes based on the aptamer-autophagosome-tethering compound (aptamer-ATTEC) strategy. A series of aptamer-ATTEC chimeras were constructed by conjugating a high-affinity ricin aptamer with an LC3-recruiting moiety via click chemistry. The optimal compound, designated DP3-D-B0, exhibited potent anti-ricin efficacy at molecular, cellular, and animal levels. Experimental results confirmed that DP3-D-B0 mediated the formation of an intracellular ternary complex (LC3-ATTEC-ricin) that hijacks the autophagy machinery for lysosome-mediated degradation. Moreover, DP3-D-B0 was found can partially blocks ricin uptake extracellularly when administered simultaneously. This work broadens the application scope of ATTEC technology and provides a novel strategy for the targeted degradation of exogenous toxins, thus advancing the development of therapeutics against biothreat agents.
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