ASMT Regulates Tumor Metastasis Through the Circadian Clock System in Triple-Negative Breast Cancer

FenFen Xie; LiLi Wang; YaJing Liu; ZhenBang Liu; ZuoYang Zhang; Jing Pei; ZhengSheng Wu; MuXin Zhai; YunXia Cao

doi:10.3389/fonc.2020.537247

ASMT Regulates Tumor Metastasis Through the Circadian Clock System in Triple-Negative Breast Cancer

Front Oncol. 2020 Oct 21:10:537247. doi: 10.3389/fonc.2020.537247. eCollection 2020.

Authors

FenFen Xie^{1

2

3

4

5

6}, LiLi Wang⁷, YaJing Liu^{1

2

3

4

5}, ZhenBang Liu⁸, ZuoYang Zhang⁹, Jing Pei¹⁰, ZhengSheng Wu⁹, MuXin Zhai¹¹, YunXia Cao^{1

2

3

4

5}

Affiliations

¹ Reproductive Medicine Center, Department of Obstetrics and Gynecology, The First Affiliated Hospital of Anhui Medical University, Hefei, China.
² NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract (Anhui Medical University), Hefei, China.
³ Key Laboratory of Population Health Across Life Cycle (Anhui Medical University), Ministry of Education of the People's Republic of China, Hefei, China.
⁴ Anhui Province Key Laboratory of Reproductive Health and Genetics, Hefei, China.
⁵ Biopreservation and Artificial Organs, Anhui Provincial Engineering Research Center, Anhui Medical University, Hefei, China.
⁶ Department of Histology and Embryology, Anhui Medical University, Hefei, China.
⁷ School of Biomedical Sciences, The University of Queensland, Brisbane, QLD, Australia.
⁸ School of Life Sciences, University of Science and Technology of China, Hefei, China.
⁹ Department of Pathology, Anhui Medical University, Hefei, China.
¹⁰ Department of Breast Surgery, The First Affiliated Hospital of Anhui Medical University, Hefei, China.
¹¹ First Clinical Medical College, Anhui Medical University, Hefei, China.

Abstract

Objective: Triple-negative (PR^-, ER^-, HER-2^-) breast cancer (TNBC) is regarded as more aggressive and more likely to recur after medical care. Emerging evidence has demonstrated that the circadian clock system regulates cell-signaling pathways critical to cancer cell proliferation, survival and metastasis, meaning that it could be a good candidate for TNBC treatment. As such, the aim of the current study was to examine the molecular mechanism by which the circadian clock system contributes to cancer progression in TNBC.

Methods: Cancer cells and primary breast cancer tissues were immunostained for the measurement of circadian clock proteins (CLOCK, BMAL1 and PER1) and acetylserotonin methyltransferase (ASMT). The association between ASMT and clock proteins was assessed using siRNA and Western blot. Transwell assays were used to detect cancer cell migration and invasion while MTT assays were utilized to evaluate cell proliferation.

Results: Circadian clock proteins (CLOCK, BMAL1, and PER1) and ASMT expression were higher in TNBC and triple positive breast cancer (TPBC) compared with para-carcinoma tissues (PCTs). Intriguingly, there was an obvious correlation between circadian clock proteins and ASMT expression in both TPBC and TNBC. Similarly, circadian clock proteins and ASMT were expressed to a greater extent in BT-474 (triple-positive) cells than in MDA-MB-231 (triple-negative) cells. The inhibition of ASMT reduced circadian clock protein levels in both breast cancer cell lines. Further analysis showed that the expression levels of ASMT and circadian clock proteins did not correlate with clinical parameters such as age, tumor size, histologic grade and CK5/6, but increased significantly with lymphatic invasion in TNBC. In agreement with this finding, knockdown of ASMT significantly leads to reductions in migration and invasion in MDA-MB-231 cells. However, over-expression of CLOCK reversed the decreases seen in ASMT inhibited cells.

Conclusion: Our study suggests that ASMT regulates the circadian clock system in breast cancer and inhibition of ASMT reduces the invasiveness of triple-negative breast cancer cells by downregulating clock protein in a certain extent, indicating the potential value of ASMT as a drug target for TNBC treatment.

Keywords: acetylserotonin methyltransferase; circadian clock system; invasion; migration; triple negative breast cancer.