β-Glucuronidase Inhibition in Drug Development: Emerging Strategies for Mitigating Drug-Induced Toxicity and Enhancing Therapeutic Outcomes

β-glucuronidase (βG) is a critical enzyme involved in the hydrolysis of glucuronide conjugates, significantly influencing drug metabolism, detoxification processes, and enterohepatic circulation. Although essential for maintaining physiological homeostasis, dysregulated βG activity has been implicated in diverse pathological conditions, including drug-induced toxicity, inflammation, and hormone-dependent cancers. Specifically, microbial βG expressed by gut microbiota can reactivate glucuronide-conjugated drugs, leading to adverse reactions through increased drug toxicity and reduced therapeutic efficacy. Consequently, inhibition of βG has emerged as an attractive therapeutic approach to reduce chemotherapy-induced toxicity, gastrointestinal complications, and metabolic disorders. This review systematically examines recent progress in the discovery, characterization, and optimization of βG inhibitors, focusing on natural products, synthetic molecules, and microbiome-targeted agents. Structure–activity relationship analyses reveal crucial functional groups and chemical modifications necessary for enhancing inhibitor potency, selectivity, and bioavailability. In addition, contemporary advances in βG inhibitor evaluation through enzyme kinetics, molecular docking simulations, high-throughput screening, and preclinical animal models are discussed, alongside essential pharmacokinetic parameters, including absorption, distribution, metabolism, excretion, and potential drug-drug interactions. Furthermore, emerging approaches such as microbiome modulation, CRISPR-based enzyme engineering, and combination therapies are explored. Despite promising preclinical outcomes, significant challenges remain regarding clinical translation, such as selectivity, bioavailability, and regulatory compliance. Ultimately, this review highlights future opportunities in precision medicine, emphasizing personalized βG inhibitor development to optimize therapeutic safety and effectiveness across various disease states.