Combination of the novel histone deacetylase inhibitor YCW1 and radiation induces autophagic cell death through the downregulation of BNIP3 in triple-negative breast cancer cells in vitro and in an orthotopic mouse model.
Triple-negative breast cancer (TNBC) is the most aggressive and invasive of the breast cancer subtypes. TNBC is a challenging disease that lacks targets for treatment. Histone deacetylase inhibitors (HDACi) are a group of targeted anticancer agents that enhance radiosensitivity. Bcl-2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP3) is a member of the Bcl-2 subfamily. BNIP3 is not found in normal breast tissue but is up-regulated in breast cancer. In the present study, we investigated the anti-cancer effects of a newly developed HDACi, YCW1, combined with ionizing radiation (IR) in TNBC in vitro and in an orthotopic mouse model. Furthermore, we examined the relationship between autophagy and BNIP3.



Trypan blue exclusion was used to investigate the viability of 4 T1 (a mouse TNBC cell line) and MDA-MB-231 cells (a human TNBC cell line) following combined YCW1 and IR treatment. Flow cytometry was used to determine apoptosis and autophagy. The expression levels of BNIP3, endoplasmic reticulum (ER) stress- and autophagic-related proteins were measured using western blot analysis. An orthotopic mouse model was used to investigate the in vivo effects of YCW1 and IR alone and in combination. Tumor volumes were monitored using a bioluminescence-based IVIS Imaging System 200.



We found that YCW1 significantly enhanced toxicity in 4 T1 cells compared with suberoylanilide hydroxamic acid (SAHA), which was the first HDACi approved by the Food and Drug Administration for clinical use in cancer patients. The combined treatment of YCW1 and IR enhanced cytotoxicity by inducing ER stress and increasing autophagy induction. Additionally, the combined treatment caused autophagic flux and autophagic cell death. Furthermore, the expression level of BNIP3 was significantly decreased in cells following combined treatment. The downregulation of BNIP3 led to a significant increase in autophagy and cytotoxicity. The combined anti-tumor effects of YCW1 and IR were also observed in an orthotopic mouse model; combination therapy resulted in a significant increase in autophagy and decreased tumor tissue expression of BNIP3 in the tumor tissue.



These data support the possibility of using a combination of HDACi and IR in the treatment of TNBC. Moreover, BNIP3 may be a potential target protein for TNBC treatment.
Division of Nephrology, Department of Internal Medicine, Shuang Ho Hospital, Taipei Medical University, Taipei, Taiwan.|Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan.
Hui-Wen
HW
Chiu
Department of Environmental and Occupational Health, College of Medicine, National Cheng Kung University, 138 Sheng-Li Road, Tainan, Taiwan, 704.
Ya-Ling
YL
Yeh
Institute of Basic Medical Sciences, National Cheng Kung University, Tainan, Taiwan.|Department of Pharmacology, National Cheng Kung University, Tainan, Taiwan.
Yi-Ching
YC
Wang
Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan.
Wei-Jan
WJ
Huang
Department of Radiation Oncology, Chi Mei Medical Center, Liouying, Tainan, Taiwan.|Chang Jung Christian University, Tainan, Taiwan.
Sheng-Yow
SY
Ho
National Institute of Environmental Health Sciences, National Health Research Institutes, No. 35 Keyan Road, Zhunan Town, Miaoli County, 350, Taiwan. pplin@nhri.org.tw.
Pinpin
P
Lin
Department of Environmental and Occupational Health, College of Medicine, National Cheng Kung University, 138 Sheng-Li Road, Tainan, Taiwan, 704. yjwang@mail.ncku.edu.tw.|Department of Biomedical Informatics, Asia University, Taichung, Taiwan. yjwang@mail.ncku.edu.tw.|Department of Medical Research, China Medical University Hospital, China Medical University, Taichung, Taiwan. yjwang@mail.ncku.edu.tw.
Ying-Jan
YJ
Wang
D000818:Animals; D001343:Autophagy; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects / Q000528:radiation effects; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D059248:Chemoradiotherapy / Q000379:methods*; D004149:Dioxoles / Q000008:administration & dosage* / Q000494:pharmacology; D015536:Down-Regulation; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects / Q000528:radiation effects; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000528:radiation effects; D056572:Histone Deacetylase Inhibitors / Q000008:administration & dosage* / Q000494:pharmacology; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics*; D051379:Mice; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D014230:Triazoles / Q000008:administration & dosage* / Q000494:pharmacology; D064726:Triple Negative Breast Neoplasms / Q000235:genetics / Q000628:therapy*; D023041:Xenograft Model Antitumor Assays
C090121:BNIP3 protein, human; D004149:Dioxoles; D056572:Histone Deacetylase Inhibitors; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D014230:Triazoles; C000604930:YCZ-18
Autophagy; Histone deacetylase inhibitor; Radiation; Triple-negative breast cancer