Comparative Evaluation of Human serum Albumin Conjugation and Micellization for the Delivery of Chlorin-based Photosensitizers

In this study, conjugation with human serum albumin (HSA) was explored as an alternative to micellar formulations for improving aqueous solubility and modulating the delivery profiles of chlorin-based photosensitizers (PSs) intended for photodynamic therapy (PDT).

A comparative analysis was performed for chlorin e6, HSA conjugates of chlorin derivatives and their unconjugated micellar formulations using in vitro assays on the A431 carcinoma cell line. Intracellular accumulation kinetics were quantified by flow cytometry at multiple incubation times, and photoinduced cytotoxicity was evaluated following 2 h, 4 h, and 8 h incubation in the presence and absence of a washout step.

Chlorin e6 demonstrated relatively slow intracellular uptake and correspondingly lower photoinduced cytotoxicity. In contrast, the non-conjugated PSs exhibited rapid and pronounced accumulation, accompanied by high levels of phototoxicity. HSA-conjugated PSs displayed a distinctly different uptake profile: intracellular accumulation increased gradually and continuously over time, yielding cytotoxicity values that approached those observed with unconjugated PSs at longer incubation periods. This behavior reflects the altered transport mechanisms imposed by albumin binding and suggests sustained cellular delivery rather than rapid saturation.

Micellar PS formulations showed the highest uptake rates and phototoxicity in vitro; however, their non-specific endocytic internalization may limit their biological selectivity in vivo. Although HSA conjugation resulted in comparatively slower uptake in vitro, albumin is an endogenous long-circulating carrier capable of passive tumor targeting and receptor-mediated cellular uptake, suggesting potential advantages for in vivo biodistribution, reduced systemic toxicity, and improved tumor selectivity compared to micellar systems.

These findings highlight that HSA conjugation provides a tunable delivery strategy that balances solubility enhancement and photodynamic efficacy, thereby representing a promising alternative to micellar formulations for PDT applications.

This work was carried out with the support of the Ministry of Science and Higher Education of the Russian Federation within the framework of the state assignment, research topic project № FSFZ-2025-0020.