Tyrosinase, a copper-dependent oxidase, plays a pivotal role in melanin biosynthesis, catalyzing the conversion of tyrosine to L-3,4-dihydroxyphenylalanine (L-DOPA) and subsequently to dopaquinone, which leads to melanin formation. While melanin is essential for skin, hair, and eye coloration and provides photoprotection, abnormal tyrosinase activity is implicated in various dermatological disorders such as melasma, hyperpigmentation, freckles, and age spots. Beyond human biology, tyrosinase is also involved in enzymatic browning reactions in fruits and vegetables, affecting food quality.
Tyrosinase inhibitors are crucial for treating hyperpigmentation disorders, developing skin-whitening cosmetics, and preventing food spoilage. Commonly used inhibitors like kojic acid, arbutin, and hydroquinone are prevalent in cosmetic formulations. However, these conventional agents often suffer from limitations such as cytotoxicity, skin irritation, instability under light or heat, and variable efficacy across different skin types, underscoring a critical demand for safer, more effective, and stable alternatives.
To address this need, the study explored natural product libraries and FDA-approved drug databases as ideal sources for discovering next-generation tyrosinase inhibitors. Natural products offer unparalleled chemical diversity and structural complexity, often interacting with biological targets with high affinity. Additionally, FDA-approved drugs come with established safety and pharmacokinetic profiles, which can significantly accelerate the development pipeline for new inhibitors. The integration of advanced computational tools, specifically machine learning (ML) and molecular docking, was considered a potential solution to efficiently screen and identify promising compounds from these extensive chemical libraries, providing predictive capabilities for biological activity and detailed insights into binding interactions. This approach aimed to overcome the challenges associated with traditional experimental workflows by prioritizing candidates with high potential for potency and safety.
Methods
The study employed a four-stage screening process to identify potent tyrosinase inhibitors. First, a comprehensive library of over 500,000 compounds from natural product and FDA-approved drug databases was compiled. Next, two ligand-based machine learning models were applied: one for predicting biological activity and another for estimating specific pIC50 values. Compounds predicted to be active with a pIC50 ≥ 4.00 were then subjected to molecular docking against multi tyrosinase-like structures (including Agaricus bisporus tyrosinase and Homo sapiens tyrosinase-related protein 1) to evaluate binding affinities. Finally, the top-ranking candidates were validated through in vitro assays for tyrosinase inhibitory activity (IC50) and transdermal permeation studies.
Key Findings
• Discovery of Novel Inhibitors: Three promising compounds were identified: rhodanine-3-propionic acid, lodoxamide, and cytidine 5′-(dihydrogen phosphate).
• Superior Inhibitory Activity: All three compounds exhibited significantly higher inhibitory activity against mushroom tyrosinase compared to arbutin (IC50 = 38.37 mM) and hydroquinone (IC50 = 10.15 mM), both widely used in cosmetics.
• Potency Ranking: Rhodanine-3-propionic acid demonstrated the most potent inhibition (IC50 = 0.7349 mM), followed by lodoxamide (IC50 = 3.715 mM) and cytidine 5′-(dihydrogen phosphate) (IC50 = 6.289 mM). While less potent than kojic acid (IC50 = 0.1129 mM), their favorable safety profiles are a key advantage.
• Mechanistic Binding Insights: The identified inhibitors showed strong binding affinities through metal ion coordination (bidentate phosphate or carboxylate donors at 2.0−2.5 Å distances) and π−π interactions with histidine residues at the enzyme’s active site.
• Enhanced Skin Permeation: Transdermal permeation experiments confirmed that these compounds achieved markedly better skin permeability and retention than commercial arbutin-based formulations. Cytidine 5′-(dihydrogen phosphate) showed the highest cumulative permeation (45.34% ± 4.50%), a 3.58-fold higher rate than α-arbutin, while lodoxamide had the highest skin retention rate (32.79%).
• Favorable Safety Profiles: Their origins from food-based sources (rhodanine-3-propionic acid and cytidine 5′-(dihydrogen phosphate)) or FDA-approved drugs (lodoxamide) suggest more favorable safety profiles, potentially avoiding the irritation and photostability concerns associated with some conventional inhibitors.
For future implications, these compounds hold significant promise as next-generation agents for inhibiting melanin production in cosmetic and pharmaceutical applications. However, it is crucial to acknowledge the current reliance on surrogate tyrosinase models (e.g., Agaricus bisporus tyrosinase and Homo sapiens tyrosinase-related protein 1, and homology models) for docking and mushroom tyrosinase for in vitro assays due to the absence of a high-resolution Homo sapiens tyrosinase (HsTYR) crystal structure. Future studies should prioritize direct testing against HsTYR and evaluation in human melanocyte cultures or 3D skin models to comprehensively confirm their efficacy and safety in biologically relevant human systems, thereby facilitating their translation into practical applications.
Link to the study: https://pubs.acs.org/doi/full/10.1021/acsomega.5c04807
Figure: Stepwise workflow for screening tyrosinase inhibitors. The process includes database compilation, ML-based activity prediction, molecular docking, and in vitro validation, identifying three promising tyrosinase inhibitors.