Diogo M. L. Santos

The overexpressed epidermal growth factor receptor (EGFR) in lung, breast, and colorectal cancers is associated with an increased risk of mortality. Therefore, targeted photodynamic therapy (PDT) has emerged as an alternative for cancer treatment. To enable EGFR-directed PDT, this study reports the synthesis of a bifunctional nanoprobe (NP-BF) based on gold nanoparticles (AuNPs) functionalized with the monoclonal antibody cetuximab (CX) and the photosensitizer chlorin e6 (Ce6) using carbodiimide chemistry. The resulting NP-BF showed spectroscopic evidence of successful conjugation through amide bond formation, preservation of the structural features of cetuximab, and retention of the photophysical properties of Ce6 after functionalization. In addition, the nanosystem remained colloidally stable, exhibiting a zeta potential of - 40.7 mV and an increase in hydrodynamic diameter of approximately 9 nm, consistent with antibody incorporation. A cellular uptake assay using the MDA-MB-468 cell line revealed internalization with maximum uptake in 60 min, corresponding to approximately 1.32 × 10⁵ nanoprobes per cell. PDT assays demonstrated a cytotoxic response with cell viability dropped below 10% at concentrations ≥ 1.62 μmol L^-1 (≈ 0.97 µg mL^-1 CE6 equivalent) under 660 nm irradiation, while non-irradiated samples preserved viability above 90%, confirming negligible dark toxicity. These results demonstrate the successful synthesis and functionalization of NP-BF and its effective photodynamic activity in cancer cells.