Resistance Of Breast Cancer Stem Cells Research Articles

Andrographolide Reverses Doxorubicin Resistance in Human Breast Cancer Stem Cells by Regulating Apoptotic Gene Expressions

BACKGROUND: Breast cancer stem cells (BCSCs) have been identified as playing a crucial role in therapeutic resistance. This resistance can be attributed to the anti-apoptotic protein survivin and the antioxidant MnSOD high expression. To overcome the resistance to doxorubicin (DOX), this study proposed the utilization of andrographolide (ANDRO), the primary bioactive compound in Andrographis paniculata leaves. The objective was to examine the role of andrographolide in regulating survivin, caspase-9, and caspase-3 gene expressions to reverse doxorubicin resistance in human BCSCs.METHODS: BCSCs were exposed to 0.1 µM DOX every two days or 50 µM rotenone (ROT) for 6 hours, subsequently supplemented with 0.3 mM ANDRO. Superoxide and peroxide levels were measured using DHE and DCFH-DA assay. The MnSOD, survivin, caspase-9, and caspase-3 mRNA expression levels were analyzed using qRT-PCR. Protein expressions were evaluated using Western blotting assay. MnSOD activity was determined using xanthine oxidase inhibition assay. The apoptotic cells were determined using Annexin-V/PI staining.RESULTS: This study indicated that the cytotoxic mechanisms of DOX, similar to ROT, in BCSCs were attributed to oxidative stress, as evidenced by an elevation in superoxide rather than peroxide levels, accompanied by a decrease in MnSOD activity. This study also highlighted that ANDRO reversed DOX resistance in BCSCs subjected to repeated DOX treatment by downregulating survivin and upregulating caspase-9 and caspase-3 mRNA expressions, thereby activating the intrinsic apoptotic pathway.CONCLUSION: This study provides insights into the role of ANDRO in modulating the expression of apoptotic genes, such as survivin, caspase-9, and caspase-3, to overcome DOX resistance in BCSCs.KEYWORDS: breast cancer, breast cancer stem cell, andrographolide, doxorubicin, oxidative stress, apoptosis

Abstract 3795: Investigate the mechanism of actions of flavonoid dimers in reversing doxorubicin resistance in breast cancer stem cells

Abstract Drug resistance, radioresistance and metastasis are the biggest hurdle in effective cancer treatment. Recent studies suggest that these problems might be caused by the presence of cancer stem cells (CSCs). CSCs are a small group of cells which can self-renew, remain undifferentiated, resist to cancer drugs and radiotherapy. It can be predicted that selective killing of CSCs may lead to tutor regression and may be a new therapeutic solution. Naturally occurring flavonoids are weakly active in inhibiting ABC transporter leading to drug accumulation in cancer cells. We have been working on the design, synthesis and characterisation of novel flavonoid dimers that can be used to reverse drug resistance in cancer cells. We recently discovered that a new class of triazole-linked flavonoid dimers 13_9 and 15_2 were effective in reversing drug resistance in CD44+CD24-/low MCF7 breast cancer stem cells at an effective concentration of less than 32 nM. Our studies suggested that a novel mechanism was involved which is independent of ABC transporters. To determine the mechanism behind, we have designed and synthesised a photoactivatable derivative of 13_9 (13_9X2) to identify the protein target(s). 13_9X2 contains a photoactivating functional group which can capture the target protein by forming a covalent bond under UV irradiation and a clickable functional group which can be used for pull-down and further identification using mass spectrometry. One candidate target is peroxiredoxin 1 (PRDX-1) which is a reactive oxygen species (ROS) scavenger. We hypothesise that CSCs are resistant to doxorubicin-induced ROS damage by overexpressing PRDX-1. By binding and disrupting PRDX-1, 13_9 and 15_2 reduce ROS scavenging ability of PRDX-1, resulting in the reversal of drug resistance in CSCs. This work provides new insights into the mechanisms of cell signalling regulation of cancer cell resistance in breast CSCs and provides a better understanding of how flavonoid dimers reverse drug resistance in CSCs. Additionally, this work contributes to the development of new anticancer drugs which can sensitize CSCs to current chemotherapy. Citation Format: Gege Sun, Iris L. Wong, Jiahua Cui, Zhen Liu, Bill T. Chan, Larry M. Chow. Investigate the mechanism of actions of flavonoid dimers in reversing doxorubicin resistance in breast cancer stem cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3795.

Abstract 3769: Evaluation of the expression and role of miRNAs-221 and -222 in stem cells derived from breast cancer cell line MCF-7.

Abstract Background: A small population of breast tumor cells, named tumor-initiating cells or cancer stem cells (CSCs), is highly tumorigenic when injected into immunodeficient mice. These CSCs, characterized by positive CD44 expression and low or undetectable levels of CD24 (CD44+CD24−/low), contribute to tumor metastasis as well as to drug resistance. Some miRNAs are differentially expressed in stem cells, suggesting they may play a role in stem cell regulation. Aim: This study has shown that miRNAs -221 and -222 are upregulated in breast CSCs when compared to differentiated cells. Our aim is to contribute to the better understanding of the biology of breast cancer cell line-derived CSCs, including the role of miRNAs -221 and -222 in these cells. Methods: Breast cancer cell line MCF-7 were grown as suspension cells in conditions that induce the formation of mammospheres, which are spheroid cell structures enriched with CSCs. Stem cell markers were analysed by Real Time PCR. The percentage of CD44+CD24−/low population was determined by FACS analysis. Overexpression of miRNAs -221 and -222 in mammospheres was done using lentivirus carrying plasmids of interest for overexpression of miR-221 or miR-222 and the control group were transducted by a plasmid with a random sequence. Analysis of mammosphere forming was done through mammosphere formation efficiency (MFE) assay. Placlitaxel resistance was firstly evaluated by MTT assay. Capase 3/7 is being used for apoptosis assay. Results: In this study we show that stem cell properties, such as the expression of OCT-4, NANOG and SOX2 were increased in mammosphere cultures (respectively 14-fold, 20-fold and 7-fold) and an enrichment of CD44+CD24−/low population was observed in mammosphere cultures when compared to adherent cell cultures. Furthermore, we found that miRNA-221 and miRNA-222 were upregulated in mammospheres by approximately 4-fold and 5-fold, respectively, when compared to adherent cells. Overexpression of miRNAs -221 or -222 increases the mammosphere formation efficiency (MFE) and death resistance to paclitaxel when compared to controls. Conclusions: The increased expression of miRNAs -221 and -222 in MFC-7 mammospheres could indicate that these molecules may play a role in regulatory pathways of breast CSCs. Moreover the overexpression of these miRNAs could increase mammosphere-forming capacity and confers paclitaxel resistance in breast CSCs. Citation Format: Juliana Carneiro, Margherita Iaboni, Cristina Quintavalle, Giuseppina Roscigno, Sara Martonelli, Gerolama Condorelli. Evaluation of the expression and role of miRNAs-221 and -222 in stem cells derived from breast cancer cell line MCF-7. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3769. doi:10.1158/1538-7445.AM2013-3769