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Updated: May 13, 2026
Isolation and Functional Assessment of Human Breast Cancer Stem Cells from Cell and Tissue Samples
Published on: October 2, 2020
ShRNA targeting Notch1 sensitizes breast cancer stem cell to paclitaxel
1Department of Pathology, Dalian Medical University, Dalian 116044, PR China.
Abstract:
Breast cancer is currently the most lethal gynecologic malignancy in many countries, and paclitaxel is a cornerstone in the treatment of this malignancy. Unfortunately, the efficacy of paclitaxel is limited due to the development of drug resistance. Evidence has suggested that cancer stem cells (CSCs) are involved in resistance to various forms of therapies, including chemotherapy. However, the interaction between paclitaxel resistance and CSCs and its underlying mechanisms have not been previously explored. In this study, we confirmed that paclitaxel enriched breast CSCs (CD44+/CD24-) in a dose-dependent manner in MCF-7 human breast cancer cell line. We then demonstrated that Notch1 was overexpressed in breast CSCs isolated from paclitaxel-treated MCF-7 cells compared to non-CSCs. The short hairpin RNA (shRNA) mediated knock-down of Notch1 inhibited MCF-7 cell proliferation and induced cell apoptosis. The anti-apoptosis protein NF-κB was decreased significantly when treated with shRNA-Notch1, and this effect was sharply improved by combination with paclitaxel. Paclitaxel decreased CD44+/CD24- cell population in MCF-7 cells and reduced the size and number of primary mammospheres after down-regulating the Notch1. Furthermore, shRNA-Notch1 inhibited the growth of tumor xenografts in nude mice noticeably. RT-PCR and Western blotting analysis showed that the expressions of ALDH1, NICD, Hes-1 and the drug transporter ABCG2 were decreased both in vitro and in vivo. These results suggest that Notch1 might play a critical role in the resistance to paclitaxel, and targeting Notch1 may have important clinical applications in cancer therapy.
Insights
Paclitaxel resistance in breast cancer is linked to cancer stem cells (CSCs). Targeting Notch1, a protein overexpressed in CSCs, may overcome this resistance and improve treatment efficacy.
Area of Science:
- Oncology
- Cell Biology
- Molecular Medicine
Background:
- Breast cancer remains a leading cause of cancer-related deaths globally.
- Paclitaxel is a key chemotherapy agent, but its effectiveness is hampered by drug resistance.
- Cancer stem cells (CSCs) are implicated in therapeutic resistance, yet their role in paclitaxel resistance is not well understood.
Purpose of the Study:
- To investigate the role of cancer stem cells (CSCs) in paclitaxel resistance in breast cancer.
- To explore the underlying mechanisms, specifically focusing on the Notch1 signaling pathway.
- To evaluate the therapeutic potential of targeting Notch1 in overcoming paclitaxel resistance.
Main Methods:
- Enrichment of breast CSCs (CD44+/CD24-) in paclitaxel-treated MCF-7 cells.
- Assessment of Notch1 expression in CSCs versus non-CSCs.
- Knockdown of Notch1 using short hairpin RNA (shRNA) and evaluation of its effects on cell proliferation, apoptosis, and CSC markers.
- In vivo studies using tumor xenografts in nude mice.
- Analysis of gene and protein expression (RT-PCR, Western blotting) for key markers including ALDH1, NICD, Hes-1, and ABCG2.
Main Results:
- Paclitaxel treatment dose-dependently enriched breast CSCs (CD44+/CD24-) in MCF-7 cells.
- Notch1 was significantly overexpressed in paclitaxel-induced CSCs.
- Notch1 knockdown inhibited cell proliferation, induced apoptosis, and decreased NF-κB activity.
- Down-regulating Notch1 reduced CSC populations, mammosphere formation, and tumor xenograft growth.
- Expressions of ALDH1, NICD, Hes-1, and ABCG2 were decreased by Notch1 knockdown both in vitro and in vivo.
Conclusions:
- Notch1 plays a crucial role in paclitaxel resistance in breast cancer by promoting CSC enrichment and survival.
- Targeting the Notch1 pathway presents a promising strategy to overcome paclitaxel resistance.
- Inhibition of Notch1 may enhance the efficacy of paclitaxel chemotherapy in breast cancer treatment.
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