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Updated: Dec 26, 2025
An In Vitro Protocol for Evaluating MicroRNA Levels, Functions, and Associated Target Genes in Tumor Cells
Published on: May 21, 2019
miR-331-3p Suppresses Cell Proliferation in TNBC Cells by Downregulating NRP2
Mingchuan Zhao1, Mengmeng Zhang2, Zhonghua Tao1
1Department of Medical Oncology, Fudan University Shanghai Cancer Center, Fudan University School of Medicine, Shanghai, People's Republic of China.
Purpose:
Triple-negative breast cancer is characterized by fast progression with high possible for metastasis and poor survival. Dysfunction of microRNAs plays an important role in the initiation and progression of cancer. Our previous microRNA-seq data indicated the downregulation of miR-331-3p in triple-negative breast cancer tissues compared with that of the noncancer tissues. However, the function of miR-331-3p in triple-negative breast cancer remains largely unknown. Herein, the involvement of miR-331-3p in triple-negative breast cancer was investigated and the therapeutic potential of miR-331-3p was also explored.
Methods:
Real-time quantitative polymerase chain reaction was performed to detect the expression of miR-331-3p in triple-negative breast cancer tissues and cell lines. The cell proliferation was determined by the cell counting kit-8 assay. Apoptosis of triple-negative breast cancer cells was examined by annexin V/propidium iodide staining. miRDB database was used to predict the potential targets of miR-331-3p. Western blot was performed to examine the expression of the target protein.
Results:
miR-331-3p was significantly downregulated in triple-negative breast cancer tissues and cell line. Lower miR-331-3p expression was significantly correlated with the tumor size, TNM stage, and lymph node metastasis of patients with triple-negative breast cancer. Functional experiments showed that the overexpression of miR-331-3p inhibited the proliferation and increased apoptosis of triple-negative breast cancer cells. Neuropilin-2 was identified as a target of miR-331-3p, which harbored binding site of miR-331-3p in its 3'-untranslated region. Overexpression of miR-331-3p decreased the messenger RNA and protein levels of neuropilin-2 in triple-negative breast cancer cells. Restoration of neuropilin-2 partially reversed the inhibitory effects of miR-331-3p on the proliferation of triple-negative breast cancer cells.
Conclusions:
Our results demonstrated the novel function of miR-331-3p/neuropilin-2 signaling in regulating the malignant behaviors of triple-negative breast cancer cells, which suggested miR-331-3p as a potential target for the treatment of triple-negative breast cancer.
Insights
MicroRNA miR-331-3p is downregulated in triple-negative breast cancer, inhibiting proliferation and increasing apoptosis. Targeting miR-331-3p and its downstream protein, neuropilin-2, shows therapeutic potential for this aggressive cancer.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Triple-negative breast cancer (TNBC) exhibits aggressive progression, metastasis, and poor survival rates.
- MicroRNA (miRNA) dysfunction is implicated in cancer initiation and progression.
- Previous studies indicated miR-331-3p downregulation in TNBC tissues.
Purpose of the Study:
- To investigate the role of miR-331-3p in TNBC.
- To explore the therapeutic potential of miR-331-3p in TNBC.
Main Methods:
- Real-time quantitative PCR to assess miR-331-3p expression in TNBC tissues and cell lines.
- Cell Counting Kit-8 assay for proliferation and Annexin V/propidium iodide staining for apoptosis.
- Bioinformatic prediction (miRDB) and Western blot to identify and validate miR-331-3p targets.
Main Results:
- miR-331-3p was significantly downregulated in TNBC tissues and cell lines, correlating with advanced tumor stage and metastasis.
- Overexpression of miR-331-3p suppressed TNBC cell proliferation and induced apoptosis.
- Neuropilin-2 (NRP2) was identified as a direct target of miR-331-3p, with its expression reduced by miR-331-3p.
- Restoring NRP2 partially reversed the anti-proliferative effects of miR-331-3p.
Conclusions:
- miR-331-3p functions as a tumor suppressor in TNBC by targeting neuropilin-2.
- The miR-331-3p/neuropilin-2 signaling pathway regulates malignant behaviors in TNBC.
- miR-331-3p represents a promising therapeutic target for TNBC treatment.
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