Regulation of melanogenesis via ubiquitin-proteasome system and autophagy by 3,3,5-trimethylcyclohexyl succinate dimethylamide and tranexamic acid.

Background: Excessive melanogenesis in skin melanocytes results in hyperpigmentation disorders.

Objective: This study investigates the combined effects and mechanisms of 3,3,5-trimethylcyclohexanol succinate dimethylamide (335, a succinic acid derivative) and tranexamic acid (TXA) on melanogenesis.

Methods: Skin penetration of 335 was assessed via Raman spectroscopy. Interactions between 335 and TXA were predicted by molecular docking. The anti-melanogenic effect was evaluated by measuring melanin content in human keratinocytes (HaCaT) and mouse melanoma cells (B16). Transcriptome sequencing was performed to identify key pathways by which 335 regulates melanogenesis. Melanosomes were isolated from human melanoma cells (MNT-1) and co-cultured with keratinocytes to validate the specific mechanisms. A 3D melanin skin model was established to evaluate the anti-pigmentation effect of 335 and TXA by analyzing apparent chroma, lightness (L*), and melanin content and distribution.

Results: The skin penetration of 335 was enhanced by the addition of TXA, and the two compounds exhibited hydrogen bonding and hydrophobic interactions. The combination of 335 and TXA (1:10) significantly reduced the melanin content in B16 cells compared to individual treatments. Transcriptomics revealed 335 modulates the ubiquitin-proteasome system (UPS) and autophagy. Experimental validation confirmed that 335 and TXA combination inhibited melanin production by promoting ubiquitination degradation of tyrosinase and autophagic degradation of melanosomes. In the 3D skin model, the combination enhanced skin brightness (apparent chroma and L* value) and reduced melanin deposition.

Conclusion: The combination of 335 and TXA inhibits melanogenesis by UPS-mediated tyrosinase degradation and autophagy-driven melanosome degradation, offering a promising strategy for hyperpigmentation treatment.