Two new water-soluble PAH-substituted (PAH - polyaromatic hydrocarbon) naphtho[1,2-d] [1,3]oxazole dyes 1–2 were synthesized and investigated for intramolecular charge transfer (ICT) behavior, aggregation, and ability to fluorescent staining and photosensitization on living cancer cells of HeLa and Vero cultures. To make clear this behavior a detailed analysis of the relationship between the structure and photophysical properties 1–2 were carried out based on experimental and computation data. As a result of the staining experiment, there was a diffuse distribution in the cytoplasm of 2-(9-anthracenyl)-containing naphthoxazole 1 as a hydrophilic fluorophore. Whereas 2-(1-pyrenyl)-derivative 2 showed accumulation in the endoplasmic reticulum, as well as additional distribution in lipid bilayers. Photodynamic activity of type II was confirmed for the cationic exogenous photosensitizer 2: the IC50 value in the case of HeLa cell culture was 0.50 μM. The radical mechanism of photodynamic activity was determined by the EPR method.