LncRNA ASAP1-IT1 enhances cancer cell stemness via regulating miR-509-3p/YAP1 axis in NSCLC

Yantao Liu; Yuping Yang; Lingli Zhang; Jiaqiang Lin; Bin Li; Min Yang; Honghui Li; Kangwu Chen; Wei Zhao

doi:10.1186/s12935-021-02270-7

LncRNA ASAP1-IT1 enhances cancer cell stemness via regulating miR-509-3p/YAP1 axis in NSCLC

Cancer Cell Int. 2021 Oct 29;21(1):572. doi: 10.1186/s12935-021-02270-7.

Authors

Yantao Liu^{1

2

3}, Yuping Yang⁴, Lingli Zhang^{1

2

3}, Jiaqiang Lin⁵, Bin Li⁵, Min Yang⁵, Honghui Li⁶, Kangwu Chen⁷, Wei Zhao^{8

9}

Affiliations

¹ Department of Pharmacy, West China Second University Hospital, Sichuan University, Chengdu, China.
² Evidence-Based Pharmacy Center, West China Second University Hospital, Sichuan University, Chengdu, China.
³ Key Laboratory of Birth Defects and Related Diseases of Women and Children, Sichuan University, Ministry of Education, Chengdu, China.
⁴ Department of Respiratory Medicine, The First Affiliated Hospital of Chengdu Medical College, Chengdu, China.
⁵ School of Laboratory Medicine Chengdu Medical College, Chengdu, China.
⁶ Department of Refractive Surgery, Chengdu Aier Eye Hospital, Chengdu, China.
⁷ Department of Orthopedic Surgery, The First Affiliated Hospital of Soochow University, Suzhou, China. kangwuchen2008@hotmail.com.
⁸ School of Laboratory Medicine Chengdu Medical College, Chengdu, China. wzhao42@cityu.edu.hk.
⁹ Department of Biomedical Sciences, City University of Hong Kong, Hong Kong, China. wzhao42@cityu.edu.hk.

Abstract

Background: Non-small cell lung cancer (NSCLC) is a major cause of cancer-related death worldwide, and cancer stem cell is responsible for the poor clinical outcome of NSCLC. Previous reports indicated that long noncoding RNAs (lncRNAs) play important roles in maintaining cancer stemness, however, the underlying mechanisms remain unclear. This study investigates the role of ASAP1 Intronic Transcript 1 (ASAP1-IT1) in cancer cell stemness of NSCLC.

Methods: The expression of ASAP1-IT1, microRNA-509-3p (miR-509-3p) and apoptosis-/stemness-related genes was analyzed by qRT-PCR in NSCLC tissues, cancer cells and spheres of cancer stem cells. Knockdown of ASAP1-IT1 or overexpression of miR-509-3p in NSCLC cells by infection or transfection of respective plasmids. Sphere formation and colony formation were used to detect NSCLC stem cell-like properties and tumor growth in vitro. Luciferase reporter assays, RNA immunoprecitation (RIP) and qRT-PCR assays were used to analyze the interaction between lncRNA and miRNA. The expression of expression of regulated genes of ASAP1-IT1/miR-509-3p axis was evaluated by qRT-PCR and Western blot. The NSCLC xenograft mouse model was used to validate the role of ASAP1-IT1 in NSCLC stemness and tumor growth in vivo.

Results: ASAP1-IT1 was up-regulated in NSCLC tissues, cancer cells, and in spheres of A549-derived cancer stem cells. Downregulation of ASAP1-IT1 or overexpression of miR-509-3p significantly decreased cell colony formation and stem cell-like properties of A549-dereived stem cells with decreased expression of stem cell biomarkers SOX2, CD34, and CD133, and suppressing the expression of cell growth-related genes, Cyclin A1, Cyclin B1, and PCNA. Furthermore, knockdown of ASAP1-IT1 or overexpression of miR-509-3p repressed tumor growth in nude mice via reducing expression of tumorigenic genes. ASAP1-IT1 was found to interact with miR-509-3p. Moreover, overexpression of ASAP1-IT1 blocked the inhibition by miR-509-3p on stem cell-like properties and cell growth of A549-dereived stem cells both in vitro and in vivo. Finally, the level of YAP1 was regulated by ASAP1-IT1 and miR-509-3p.

Conclusions: YAP1-involved ASAP1-IT1/miR-509-3p axis promoted NSCLC progression by regulating cancer cell stemness, and targeting this signaling pathway could be is a promising therapeutic strategy to overcome NSCLC stemness.

Keywords: ASAP-IT1; Cancer stemness; NSCLC; YAP1; miR-509-3p.

Grants and funding

81602636/National Natural Science Foundation of China