Protoporphyrin IX conjugated to piperazine or morpholine: Photophysical properties and photodynamic activity

Abstract

Designing photosensitizers (PS) with ideal properties for Photodynamic Therapy (PDT) remains challenging, but crucial for advancing PDT as a viable anticancer treatment. Combining porphyrin with potent anticancer agents, such as piperazine and morpholine groups, has the potential to give rise to innovative photosensitizers that exhibit heightened PDT effects in cancer cells. Herein, we present two novel molecules bearing piperazine (PpIX-Pip) or morpholine (PpIX-Morp) groups conjugated to Protoporphyrin IX (PpIX), describing their photophysical and photodynamic characteristics. Analysis of the photophysical properties revealed similar fluorescence quantum yields and lifetimes between PpIX and the conjugated molecules. The PpIX-Pip showed higher production of singlet oxygen and better interaction with HSA in comparison to PpIX-Morp. Cellular uptake assays revealed a greater incorporation rate for PpIX-Pip (37 %) compared to PpIX-Morp (29 %) in HeLa cells. Essentially, both PpIX derivatives exhibited superior uptake efficiencies compared to PpIX alone (17 %). The descending order of IC₅₀ values was PpIX > PpIX-Morp > PpIX-Pip, indicating that the introduction of piperazine and morpholine groups to PpIX elevated the toxicity in HeLa cells. When comparing IC₅₀ values obtained under irradiation to the dark control, it was observed that PpIX-Pip demonstrated higher phototoxic effect (phototherapeutic index, PI > 5), whereas PpIX-Morp exhibits medium phototoxicity towards HeLa cells (PI > 2). Among the newly studied porphyrins, PpIX-Pip displayed a higher singlet oxygen generation rate, and improved cellular uptake as well as lower value of IC₅₀ in the light. This is the first report highlighting the PDT activity of PpIX modified with the N-methylpiperazine group. Our findings open avenues for the design and development of photosensitizers linked to anticancer agent moieties, encompassing groups like piperazine and morpholine.