3D bioprinted melanoma constructs reveal delivery-dependent efficacy of phytochemical-gold nanoparticle formulations.

This study investigates the efficacy of phytochemical nanoformulations-specifically curcumin and thymoquinone (TQ) delivered via gold nanoparticles (AuNPs)-against melanoma A375 cells in both 2D and 3D bioprinted gelatin-alginate scaffolds. Phytochemicals such as curcumin, TQ, epigallocatechin gallate, and betulin exhibit multi-target anticancer effects, but their clinical translation is limited by poor solubility, rapid metabolism, and low tumor penetration. We compared free phytochemicals, AuNP co-administration, and AuNP-phytochemical conjugates, assessing their effects on viability, ROS generation, and mitochondrial membrane potential over time. In 2D cultures, all agents exhibited dose-dependent cytotoxicity, with curcumin and TQ proving to be the most potent. However, in 3D scaffolds mimicking tumor microenvironments, only AuNP-phytochemical conjugates sustained mitochondrial and redox stress, overcoming adaptation barriers and providing durable suppression of melanoma viability. Free and co-administered agents displayed metabolic rebound and limited efficacy due to diffusion constraints and extracellular matrix-driven resistance. Unlike prior studies that investigate either free phytochemicals or nanoparticle delivery in isolation, this work integrates 2D and 3D bioprinted A375 melanoma constructs to directly compare free, co-administered, and gold-nanoparticle-conjugated phytochemicals. We show that only conjugated AuNP-phytochemical formulations sustain mitochondrial and redox stress long enough to overcome 3D adaptation barriers, establishing a delivery-strategy-dependent framework for preclinical evaluation of phytochemical nanomedicines.