Synergistic anticancer effects of ruxolitinib and calcitriol in estrogen receptor-positive, human epidermal growth factor receptor 2-positive breast cancer cells

Abstract

The Janus kinase (JAK)1 and JAK2 inhibitor, ruxolitinib, and the active form of Vitamin D (calcitriol) were previously reported to possess anticancer effects in breast cancer. The present study investigated the combined effects of ruxolitinib and calcitriol on an estrogen receptor (ER)-positive, human epidermal growth factor receptor 2 (HER2)-positive, breast cancer cell line. The ER and HER2-positive MCF7-HER18 breast cancer cell line was used to investigate the combination effect of ruxolitinib and calcitriol. A bromodeoxyuridine (BrdU) assay was used to investigate cell growth inhibition. The synergism of this combination therapy was examined using the Chou-Talalay method. Cell cycle analysis was performed by flow cytometry, and apoptosis was evaluated by flow cytometry following Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI) staining. Alterations in protein expression levels were analyzed by western blotting. The BrdU assay indicated that combination treatment using ruxolitinib and calcitriol produced a synergistic anti-proliferative effect in MCF7-HER18 breast cancer cells. Annexin V-FITC/PI staining and cell cycle analysis identified a synergistic increase in apoptosis and sub-G1 arrest in the presence of ruxolitinib and calcitriol. Western blot analysis revealed that these synergistic effects of ruxolitinib and calcitriol were associated with reduced protein levels of JAK2, phosphorylated JAK2, c-Myc proto oncogene protein, cyclin-D1, apoptosis regulator Bcl-2 and Bcl-2-like protein 1, and with increased levels of caspase-3 and Bcl-2-associated agonist of cell death proteins. The results of the present study demonstrated the synergistic anticancer effects of ruxolitinib and calcitriol in ER and HER2-positive MCF7-HER18 breast cancer cells. Based on these findings, ruxolitinib and calcitriol may have potential as a combination therapy for patients with ER and HER2-positive breast cancer.