To examine whether enzalutamide enhanced palbociclib-induced cytostatic effect, we used 0.156 or 2.5 M palbociclib combined with Rabbit Polyclonal to ZAR1 various concentrations of enzalutamide in TNBC cell lines. impartial experiments performed in triplicate are shown.(TIF) pone.0189007.s002.TIF (1.2M) GUID:?9B129832-936D-4BA7-9805-0E3B5CF70D05 S2 Fig: The effects of palbociclib are attenuated by RB knockdown. MDA-MB-453 cells were transfected with siRNA against control (siCtrl) and RB1 (siRB) for 24 h, and the transfected cells were further treated with 2.5 M palbociclib for 48 h. The treated cells were analyzed by flow cytometry analysis.(TIF) pone.0189007.s003.TIF (2.4M) GUID:?A9E58DE6-3B29-4EB0-9A5C-50CED05668CD S3 Fig: AR expression could be detected in patients with TNBC and did not link to recurrence-free survival. (A) Representative tissue microarray of immunohistochemical expression of AR in TNBC samples and the events of AR expression. (B) recurrence-free survival of TNBC patients were plotted against time in month for the level of gene.(TIF) pone.0189007.s004.TIF (6.1M) GUID:?D5B1086D-7A3A-4B2C-8D1C-0DAE03A0C6D4 S4 Fig: The correlation between and expression in patients with breast cancer. The level 3 data of mRNA RSEM in breast cancer were downloaded from the TCGA and Broad GDAC Firehose data portal. The correlation between and mRNA was analyzed by Pearson correlation analysis.(TIF) pone.0189007.s005.TIF Calcipotriol (1010K) GUID:?D0E15E0F-193F-435E-939A-0E237CAF23BF Data Availability StatementAll relevant data are within the paper Abstract Objectives Triple unfavorable breast cancer (TNBC) lacks specific drug targets and remains challenging. Palbociclib, a cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitor is usually approved for metastatic estrogen receptor (ER)-positive and human epithermal growth factor 2 (HER2)-unfavorable breast cancer. The nature of cell cycle inhibition by palbociclib suggests its potential in TNBC cells. Retinoblastoma (RB, a known substrate of CDK4/6) pathway deregulation is usually a frequent occurrence in TNBC and studies have revealed that pharmacological CDK4/6 inhibition induces a cooperative cytostatic effect with doxorubicin in RB-proficient TNBC models. In addition, recent studies reported that anti-androgen therapy shows preclinical efficacy in androgen-receptor (AR)-positive TNBC cells. Here we examined the effect of palbociclib in combination with an anti-androgen enzalutamide in TNBC cells. Method MDA-MB-453, BT-549, MDA-MB-231 and MDA-MB-468 TNBC cell lines were used for studies. Protein expressions were assessed by Western blot analysis. Cytostatic effect was examined by MTT assay. Cell cycle and apoptosis were examined by flow cytometry. Results Palbociclib showed inhibitory effect in RB-proficient TNBC cells, and enzalutamide inhibited cell viability in AR-positive TNBC cells. Enzalutamide treatment could enhance the palbociclib-induced cytostatic effect in AR-positive/RB-proficient TNBC cells. In addition, palbociclib-mediated G1 arrest in AR-positive/RB-proficient TNBC cells was attenuated by RB knockdown. Conclusion Our study Calcipotriol provided a preclinical rationale in selecting patients who might have therapeutic benefit from combining CDK4/6 inhibitors with AR antagonists. Introduction Triple-negative breast cancer (TNBC) remains a challenging breast cancer subtype for its higher risk of distant recurrence, and poorer outcome after recurrence or metastasis than other types of breast malignancy [1C3]. Targeted therapy for TNBC is usually emerging in clinical trials and recent molecular profiling studies have revealed molecular heterogeneity of TNBC , highlighting the importance of obtaining biomarkers for targeted therapy guidance for TNBC. Palbociclib is usually a highly selective cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitor, which blocks the phosphorylation of retinoblastoma protein (pRB) and subsequently arrests cell cycle at G1-phase [5, 6]. Previous study showed that palbociclib in combination with hormone therapy Calcipotriol (tamoxifen) or target therapy (trastuzumab) had an effectively inhibitory effect on ER-positive and HER2-amplified breast cancer, respectively . In clinical, palbociclib in combination with letorzole (aromatase inhibitor) has been approved by the U.S. Food and Drug Administration (FDA) for the treatment of patients with ER-positive and HER2-unfavorable advanced breast malignancy [8, 9]. However, the effects of palbociclib in TNBC are not well-documented. Enzalutamide, an androgen receptor antagonist, has been approved by the FDA for the treatment of patients with metastatic prostate cancer [10, 11]. Cumulative evidences showed that enzalutamide has potent anti-tumor effects on TNBC cells, and suggested that androgen receptor (AR) might be a promising target for Calcipotriol treatment of TNBC [12C14]. However, the effect of combination palbociclib with enzalutamide in Calcipotriol TNBC cells is still unclear. In present study, we tested the combination effect of palbociclib with enzalutamide in TNBC cells. Cytostatic effects of enzalutamide, palbociclib or combined treatment and effects of treatments on AR and pRB proteins expressions were examined. Moreover, the influences on cell cycle distribution and apoptosis.