Targeting ERβ1-Positive Triple-Negative Breast Cancer: Molecular Effects of Calcitriol and 17β-Estradiol

Background Breast cancer is the most common malignancy in women. Triple-negative breast cancer (TNBC) is characterized by the absence of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2, contributing to its aggressive nature, limited treatment options, and poor prognosis. Emerging evidence highlights estrogen receptor beta 1 (ERβ1) as a potential tumor suppressor in TNBC, influencing key oncogenic pathways such as cell proliferation, survival, angiogenesis, and apoptosis. In this regard, calcitriol (active vitamin D) and 17β-estradiol have been identified as key regulators of tumor behavior. Calcitriol shows strong anti-proliferative and pro-apoptotic effects, while the ability of 17β-estradiol to modulate tumor progression through ERβ1 signaling is context-dependent. This study aims to investigate the individual and combined effects of calcitriol and 17β-estradiol in ERβ1-expressing MDA-MB-468 TNBC cells, with a focus on their role in regulation of tumor progression, angiogenesis, and apoptosis. The findings provide novel insights into the potential therapeutic utility of targeting ERβ1 in TNBC. Methodology The MDA-MB-468 TNBC cells were treated with calcitriol (1-5 µM) and/or 17β-estradiol (100-500 nM). The effect on cell viability was assayed with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. At the same time, immunoblot analysis investigated the time-dependent manner of ERβ1, epidermal growth factor receptor (EGFR), vascular epithelial growth factor (VEGF), and caspase-3. Results Both calcitriol and 17β-estradiol substantially decreased TNBC cell viability, with the highest level of cytotoxicity observed at 24 and 32 hours, respectively. The combination increased the amount of cell death. Immunoblots revealed lasting downregulation of ERβ1, EGFR, VEGF, and caspase-3 after calcitriol treatment. In comparison, 17β-estradiol demonstrated biphasic regulatory behavior for ERβ1, where ERβ1 was first downregulated, then partially recovered. The combination therapies produced more significant ERβ1 downregulation and heightened suppression of EGFR and VEGF, further enhancing their effects on TNBC progression. Conclusions This study aims to investigate the individual and combined effects of calcitriol and 17β-estradiol in ERβ1-expressing MDA-MB-468 TNBC cells, with a focus on their role in the regulation of tumor progression, angiogenesis, and apoptosis.