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.

Vitamin D3 and calcium cosupplementation alleviates cadmium hepatotoxicity in the rat: Enhanced antioxidative and anti‐inflammatory actions by remodeling cellular calcium pathways

“…[56] These controversial results could be related to the interactions between VD 3 and the B-cell lymphoma 2 (BCL2) protein, which regulates the ER Ca 2+ release by inhibiting ITPR1 and RyR1. [57,58] Whilst VD 3 promoted Ca 2+ -dependent apoptosis in cancer cells by inhibiting BCL2, [58] the hormone promoted BCL2 during oxidative stress and inflammatory conditions, thereby preserved cell viability by constraining [Ca 2+ ] I release through ITPR1 and RyR1. [57,59] An alternative explanation for the VD 3 inhibitory effects on SOCs could also be related to the observed increase in the S100A1 protein, which alleviated myocardial infarction in animals by inhibiting [Ca 2+ ] I release through RYR1.…”

Inhibition of tumor growth and angiogenesis of tamoxifen‑resistant breast cancer cells by ruxolitinib, a selective JAK2 inhibitor

“…We believe that the functional role of JAK2, which can be inhibited by ruxolitinib, is not mainly involved in the growth of TAMR-MCF7 cells. Lim et al (21) recently demonstrated that ruxolitinib and calcitriol possessed synergistic anticancer effect in MCF-7 cells. Although ruxolitinib showed marginal inhibition rate in cell growth of MCF-7 cells, synergistic inhibition of cell proliferation was observed by ruxolitinib/calcitriol combination.…”

“…In preclinical studies, ruxolitinib was shown to inhibit the proliferation of JAK2 V617F -positive Ba/F3 cells showing a constitutively active mutant form of JAK2, and to alleviate MPN symptoms in JAK2 V617F -transgenic mice (20). Moreover, involvement of JAK2 in cell proliferation was experimentally verified in various cancer cell lines, lung cancer cells (H661, H1975, H1563, ADOR, NSCLC1), breast cancer cells (MCF-7, MCF-7-HER18, SUM149, BT474, BT549, SKBR3) and glioblastoma cells (GBM6, GBM12) (21)(22)(23). However, the therapeutic effects of ruxolitinib on chemo-resistant breast cancer cells, and especially TAM-resistant cells, remain obscure.…”

“…Ruxolitinib directly inhibits JAK1 and JAK2 kinases, with IC 50 values of 3.3 and 2.8 nM, respectively (20). Ruxolitinib reduced protein levels of phosphorylated JAK2 and subsequently inhibited protein expression of its downstream target genes, c-Myc, cyclin D1 and Bcl-2 (21). In preclinical studies, ruxolitinib was shown to inhibit the proliferation of JAK2 V617F -positive Ba/F3 cells showing a constitutively active mutant form of JAK2, and to alleviate MPN symptoms in JAK2 V617F -transgenic mice (20).…”

Targeting the JAK/STAT pathway in solid tumors

“…The JAK1/2-selective inhibitor ruxolitinib is FDA approved for the treatment of polycythemia vera, myelofibrosis, and graft versus host disease, and it has been shown to decrease STAT3 activation in preclinical models of several solid tumors [18,22,65] . Ruxolitinib inhibited STAT3 activation and decreased cell growth in breast cancer [66,67] , NSCLC [68] , HNC [69] , esophageal cancer [70] , bladder cancer [71] , HCC [72] , cervical cancer [73] , and colorectal cancer [74,75] cell lines. In pancreatic cancer cells, ruxolitinib treatment was also shown to decrease expression of pro-angiogenic genes and impede epithelial-to-mesenchymal transition [76,77] .…”