Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2018 Apr;17(4):5581-5588.
doi: 10.3892/mmr.2018.8580. Epub 2018 Feb 8.

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

Seung Taek Lim  1 , Ye Won Jeon  1 , Hongki Gwak  1 , Se Young Kim  1 , Young Jin Suh  1
Affiliations

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

Seung Taek Lim et al. Mol Med Rep. 2018 Apr.

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.

Keywords: breast neoplasm; ruxolitinib; calcitriol; synergism; apoptosis.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
The expression of VDR and JAK2 in MCF7-HER18 breast cancer cells was demonstrated by western blot analysis. VDR, vitamin D receptor; JAK, Janus kinase; HER, human epidermal growth factor receptor.
Figure 2.
Figure 2.
Cell proliferation evaluation using the BrdU assay in the presence of the indicated concentrations of ruxolitinib and/or calcitriol in MCF7-HER18 cells for 72 h. The data are presented as the mean ± standard deviation. *P<0.05. BrdU, bromodeoxyuridine; HER, human epidermal growth factor receptor.
Figure 3.
Figure 3.
Cell cycle distribution of MCF7-HER18 cells treated as indicated for 72 h. (A) Cell cycle distribution was investigated using flow cytometry and (B) the data were quantified. The data are presented as the mean ± standard deviation. *P<0.05.; HER, human epidermal growth factor receptor.
Figure 4.
Figure 4.
Flow cytometry analysis of Annexin-FITC/PI stained MCF7-HER18 cells was performed. Following exposure to the indicated treatments for 72 h. (A) B1 quadrant (Annexin−, PI+) represents necrotic cells, B2 quadrant (Annexin+, PI+) represents late apoptotic cells, B3 quadrant (Annexin−, PI−) represents viable cells, and B4 quadrant (Annexin+, PI−) represents early apoptotic cells. (B) The graph shows the proportions of early and late apoptotic cells following 72 h of treatment as indicated. The data are presented as the mean ± standard deviation. *P<0.05. FITC, fluorescein isothiocyanate; PI, propidium iodide; HER, human epidermal growth factor receptor.
Figure 5.
Figure 5.
Levels of JAK2, p-JAK2, c-Myc, cyclin-D1, caspase 3, Bcl-2, Bcl-xL, BAD and β-actin proteins in MCF7-HER18 cells were determined by western blot analysis following the indicated 72 h treatments. The data are presented as the mean ± standard deviation. *P<0.05. JAK, Janus kinase; Bcl-2, apoptosis regulator Bcl-2; Bcl-xL, Bcl-2-like protein 1; BAD, Bcl-2-associated agonist of cell death; Myc, Myc proto oncogene protein HER, human epidermal growth factor receptor.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Siegel RL, Miller KD, Jemal A. Cancer statistics, 2016. CA Cancer J Clin. 2016;66:7–30. doi: 10.3322/caac.21332. - DOI - PubMed
    1. Sorlie T, Tibshirani R, Parker J, Hastie T, Marron JS, Nobel A, Deng S, Johnsen H, Pesich R, Geisler S, et al. Repeated observation of breast tumor subtypes in independent gene expression data sets; Proc Natl Acad Sci USA; 2003; pp. 8418–8423. - DOI - PMC - PubMed
    1. Goldhirsch A, Winer EP, Coates AS, Gelber RD, Piccart-Gebhart M, Thürlimann B, Senn HJ, Panel members Personalizing the treatment of women with early breast cancer: Highlights of the St Gallen International Expert Consensus on the Primary Therapy of Early Breast Cancer 2013. Ann Oncol. 2013;24:2206–2223. doi: 10.1093/annonc/mdt303. - DOI - PMC - PubMed
    1. Russnes HG, Lingjærde OC, Børresen-Dale AL, Caldas C. Breast cancer molecular stratification: From intrinsic subtypes to integrative clusters. Am J Pathol. 2017;187:2152–2162. doi: 10.1016/j.ajpath.2017.04.022. - DOI - PubMed
    1. Voduc KD, Cheang MC, Tyldesley S, Gelmon K, Nielsen TO, Kennecke H. Breast cancer subtypes and the risk of local and regional relapse. J Clin Oncol. 2010;28:1684–1691. doi: 10.1200/JCO.2009.24.9284. - DOI - PubMed

MeSH terms