Enhanced Cell Uptake of Rhodamine B for Photodynamic Therapy under Hypoxic Conditions Using Sepia Melanin Nanoparticles

Photodynamic therapy (PDT) is a promising light-triggered therapy for medical conditions, including solid tumors, and it is already being used in clinical settings. PDT relies on the use of a photosensitizer that generates cytotoxic oxygen species upon photoexcitation. Because of their high molar extinction coefficient, Rhodamine (Rh) dyes are promising candidates as photosensitizers. However, the low solubility and poor cell uptake of Rh dyes hamper their use in this context. Moreover, because PDT relies on the availability of molecular oxygen, PDT formulations should effectively function both at normal oxygen levels and in hypoxic conditions. We herein address these limitations by proposing the use of natural melanin nanoparticles (MNPs) as an intracellular nanocarrier for RhB isothiocyanate (RhB). MNPs derived from the ink of Sepia officinalis are affordable and offer innate biocompatibility, making them promising platforms for drug delivery. In this study, we loaded them with RhB (19% loading efficiency), thus enhancing the aqueous stability of the dye and promoting its cellular internalization. Our results demonstrate that RhB-MNPs exhibit reduced intrinsic cytotoxicity yet are highly cytotoxic when photoexcited, decreasing cell viability below 60% and 50% in SaOs-2 and U-87 cell lines, respectively. Unlike in most reported PDT approaches, this effect is of equal magnitude under atmospheric O 2 levels and in hypoxic conditions. The PDT effect was further characterized by quantifying ROS production and unveiling the ROS generation mechanism. This research represents a significant advancement in PDT nanomedicine, offering possibilities for enhancing therapeutic efficacy in cancer treatment.