Mitochondria‐Targeting Photocytotoxic Ferrocenyl Conjugates of N ‐Alkylpyridinium Salts

Abstract

Ferrocenyl (Fc) conjugates (1–3) of alkylpyridinium cations (E)-N-alkyl-4-[2-(ferrocenyl)vinyl]pyridinium bromide (alkyl = n-butyl in 1, N,N,N-triethylbutan-1-aminium bromide in 2, and n-butyltriphenylphosphonium bromide in 3) were prepared and characterized, and their photocytotoxicities and cellular uptakes in HeLa cancer and 3T3 normal cells were studied. The species with a 4-methoxyphenyl moiety (4) instead of Fc was used as a control. The triphenylphosphonium-appended 3 was designed for specific delivery into the mitochondria of the cells. Compounds 1–3 showed metal-to-ligand charge-transfer bands at λ ≈ 550 nm in phosphate buffered saline (PBS). The Fc+/Fc and pyridinium core redox couples were observed at 0.75 and –1.2 V versus a saturated calomel electrode (SCE) in CH2Cl2/0.1 M (nBu4N)ClO4. Conjugate 3 showed a significantly higher photocytotoxicity in HeLa cancer cells [IC50 = (1.3 ± 0.2) μM] than in normal 3T3 cells [IC50 = (27.5 ± 1.5) μM] in visible light (400–700 nm). The positive role of the Fc moiety in 3 was evident from the inactive nature of 4. A JC-1 dye (5,5,6,6-tetrachloro-1,1,3,3-tetraethylbenzimidazolylcarbocyanine iodide) assay showed that 3 targets the mitochondria and induces apoptosis by the mitochondrial intrinsic pathway caused by reactive oxygen species (ROS). Annexin/propidium iodide studies showed that 3 induces apoptotic cell death in visible light by ROS generation, as evidenced from dichlorofluorescein diacetate assay. Compounds 1–3 exhibit DNA photocleavage activity through the formation of hydroxyl radicals.