The mechanism of bifonazole-induced [Ca²⁺]i rises and non-Ca²⁺-triggered cell death in PC3 human prostate cancer cells

Bifonazole is an antifungal drug widely used for treating skin diseases. The effect of bifonazole on physiology of cancer cells is unclear. The effect of bifonazole on cytosolic free Ca²⁺ concentrations ([Ca²⁺]i) and viability in PC3 human prostate cancer cells was explored. The Ca²⁺-sensitive fluorescent dye, fura-2, was applied to measure [Ca²⁺]i. Bifonazole at concentrations of 5-30M induced a [Ca²⁺]i rise in a concentration-dependent manner. The response was reduced by 50% by removing extracellular Ca²⁺. Bifonazole-evoked [Ca²⁺]i rise was not altered by nifedipine, econazole, SK&F96365 and protein kinase C activator, but was inhibited by 75% by GF109203X, a protein kinase C inhibitor. In Ca²⁺-free medium, treatment with the endoplasmic reticulum Ca²⁺ pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) nearly abolished bifonazole-evoked [Ca²⁺]i rise. Conversely, treatment with bifonazole abolished BHQ-evoked [Ca²⁺]i rise. Inhibition of phospholipase C with U73122 abolished bifonazole-induced [Ca²⁺]i rise. At 30-100M, bifonazole decreased cell viability concentration-dependently, which was not reversed by chelating cytosolic Ca²⁺ with 1,2-bis(2-aminophenoxy)ethane-N,N,N″,N′-tetraacetic acid/acetoxy methyl. Annexin V/propidium iodide staining data suggest that bifonazole (30-100M) induced apoptosis concentration-dependently. Together, in PC3 human prostate cancer cells, bifonazole induced [Ca²⁺]i rises by inducing phospholipase C- and protein kinase C-dependent Ca²⁺ release from the endoplasmic reticulum and Ca²⁺ influx via non-store-operated pathways. Bifonazole induced cell death that might involve apoptosis.