Despite advances in surgery, radiation, and
chemotherapy, novel
therapeutics are needed for
head and neck cancer treatment. The objective of this current study was to evaluate
alexidine dihydrochloride as a novel compound lead for head and
neck cancers. Using a tetrazolium-based assay, the dose required to reduce cell viability by 50% (ED50) was found to be approximately 1.8 micromol/L in FaDu (human hypopharyngeal squamous
cancer) and approximately 2.6 micromol/L in C666-1 (human undifferentiated
nasopharyngeal cancer) cells. In contrast, the ED50 values were much higher in untransformed cells, specifically at approximately 8.8 micromol/L in GM05757 (primary normal human fibroblast), approximately 8.9 micromol/L in HNEpC (primary normal human nasal epithelial), and approximately 19.6 micromol/L in NIH/3T3 (mouse embryonic fibroblast) cells.
Alexidine dihydrochloride did not interfere with the activities of
cisplatin,
5-fluorouracil, or radiation, and interacted in a less-than-additive manner.
DNA content analyses and
Hoechst 33342 staining revealed that this compound induced apoptosis.
Alexidine dihydrochloride-induced mitochondrial damage was visualized using transmission electron microscopy. Mitochondrial membrane potential (DeltaPsiM) depolarization was detectable after only 3 hours of treatment, and was followed by cytosolic Ca2+ increase along with loss of membrane integrity/cell death.
Caspase-2 and
caspase-9 activities were detectable at 12 hours,
caspase-8 at 24 hours, and
caspase-3 at 48 hours. FaDu cell clonogenic survival was reduced to < 5% with 1 micromol/L
alexidine dihydrochloride, and, correspondingly, this compound decreased the in vivo
tumor-forming potential of FaDu cells. Thus, we have identified
alexidine dihydrochloride as the first bisbiguanide compound with anticancer specificity.