Exploring the Ca(2+) signaling and cytotoxicity induced by the alantolactone in breast cancer cells and its potential implications in treatment using the Ca(2+) chelating agent BAPTA-AM.

Alantolactone, a bioactive sesquiterpene lactone derived from the roots of Inula helenium (elecampane), has garnered attention in biomedical and pharmacological research for its diverse therapeutic properties, including anticancer, anti-inflammatory, antimicrobial, and antioxidant activities. Despite its well-documented bioactivity, the effects of alantolactone on calcium ion (Ca(2+)) signaling and the underlying mechanisms in human breast cancer cells remain poorly understood. This study explored how alantolactone influences intracellular Ca(2+) levels ([Ca(2+)](i)), cell viability, and the role of Ca(2+)-dependent pathways in T-47D human breast cancer cells. Specifically, it examined the relationship between Ca(2+) signaling and cytotoxicity in cells exposed to alantolactone, with or without the Ca(2+) chelator BAPTA-AM. The findings reveal that alantolactone (25-75 μM) increases [Ca(2+)](i) in a concentration-dependent manner, while concentrations of 25-100 μM induce cytotoxicity, an effect that can be reversed by BAPTA-AM pre-treatment. Removing extracellular Ca(2+) significantly inhibits Ca(2+) influx, and both SKF96365 and 2-APB, modulators of store-operated Ca(2+) channels, block the alantolactone-induced Ca(2+) entry. Additionally, in a Ca(2+)-free environment, thapsigargin, an inhibitor of the endoplasmic reticulum Ca(2+) pump, suppresses the alantolactone-induced rise in [Ca(2+)](i), while alantolactone reduces the [Ca(2+)](i) increase triggered by thapsigargin. Moreover, inhibiting phospholipase C (PLC) with U73122 abolishes the alantolactone-induced [Ca(2+)](i) elevation. These results suggest that alantolactone-induced cell death in T-47D cells is Ca(2+)-dependent, involving Ca(2+) entry via store-operated channels and Ca(2+) release from the endoplasmic reticulum, with PLC playing a pivotal role. Importantly, the ability of BAPTA-AM to reverse alantolactone's cytotoxic effects highlights its potential therapeutic significance in breast cancer research.