MiR-103a-3p Promotes Tumorigenesis of Breast Cancer by Targeting ETNK1

Abstract

Background: We aimed to elucidate the molecular mechanism of miR-103a-3p regulating breast cancer progression.

Methods: Firstly, clinical tissues was obtained from 2019-2023 at Yancheng Third People's Hospital, Yancheng, China. miR-103a-3p or ETNK1 expression in clinical tissues or breast cancer cell lines was analyzed with qRTPCR. MDA-MB-231 cells were performed with miR-103a-3p inhibitor or mimic, and OE-ETNK1. The proliferation and apoptosis ability were detected by CCK-8 and TUNEL assay. The xenograft models were established by inoculating transfected MDA-MB-231 cells to BALB/c mice.

Results: miR-103a-3p showed an overexpression and was related to poor prognosis in breast cancer. miR-103a-3p-deprived MDA-MB-231 cells displayed weaker levels of cell proliferation and higher rates of apoptosis. In contrast, ETNK1 was downregulated in breast cancer and proved to be a downstream target of miR-103a-3p. Xenograft models subjected to either miR-103a-3p antagomir treatment or ETNK1-knockdown resulted in tumor growth suppression.

Conclusion: miR-103a-3p might promote breast cancer progression by inhibiting ETNK1.

Keywords: Apoptosis; Breast cancer; MiR-103a-3p; Proliferation.

Fig. 1:

miR-103a-3p is highly overexpressed in breast cancer and leads to poor prognosis (A) The expression of miR-374b-5p, let-7i-5p, miR-651-5p, and miR-103a-3p in 5 pairs of breast cancer clinical specimens was quantified by qRT-PCR. (B) TCGA database was used to analyze miR-103a-3p level in breast cancer. (C) miR-103a-3p expression in 16 pairs of clinical tissues was detected by qRT-PCR. (D) TCGA database showed relationship between miR-103a-3p and survival rate of breast cancer patients. ns, not significant; ***, P < 0.001

Fig. 2:

miR-103a-3p regulates proliferation and apoptosis of MDA-MB-231 cells. (A) Transfection efficiency of miR-103a-3p was detected by qRT-PCR. (B) CCK-8 and (C) Colony formation assay were carried out for the detection of cell proliferation ability. (D) TUNEL staining was carried out to detect the apoptosis level in each group. **, P < 0.01; ***, P< 0.001

Fig. 3:

miR-103a-3p targets binding ETNK1. (A) The expression of ETNK1 in different cell lines was quantified by qRT-PCR. (B) TCGA database was used for the analyzing of the relationship between ETNK1 and survival rate of breast cancer patients. (C) The binding sites of miR-103a-3p and ETNK1 were determined by using Targetscan website. (D) Dual-luciferase reporter assay verified the binding relationship between ETNK1 and miR-103a-3p. (E) QRT-PCR and (F) western blot were performed for the detection of the mRNA and protein expression of ETNK1 in each group. *, P < 0.05; **, P < 0.01; ***, P < 0.001

Fig. 4:

miR-103a-3p regulates proliferation and apoptosis of breast cancer cells through ETNK1. MDA-MB-231 cells were transfected with mimic NC+OE NC, miR-103a-3p mimic+OE NC, miR-103a-3p mimic+OE-ETNK1. (A) The expression of miR-103a-3p and ETNK1 was quantified by qRT-PCR. (B) CCK-8 and (C) colony formation assay were performed for the detection of the cell proliferation. (D) TUINEL staining was used to detect the apoptosis level. **, P < 0.01; ***, P < 0.001

Fig. 5:

miR-103a-3p regulates proliferation and apoptosis of mouse breast cancer through ETNK1. We used stable transfected MDA-MB-231 cells to construct breast cancer xenografts in nude mice, the group name of which was: antagomir NC + sh-NC, miR-103a-3p antagomir + sh-NC, miR-103a-3p antagomir+sh-ETNK1. (A) The mouse tumor was dissected and photographed. (B) Tumor volume and (C) weight were measured. (D) QRTPCR was performed to qualify the related factors in tumor tissues. (E) ETNK1 expression was analyzed by IHC. (F) The apoptosis level was detected by TUNEL staining. ***, P < 0.001