Abstract: Breast cancer is one of the most malignant tumors and is associated with high mortality rates among women. Lycium barbarum polysaccharide (LBP) is an extract from the fruits of the traditional Chinese herb, L. barbarum. LBP is a promising anticancer drug, due to its high activity and low toxicity. Although it has anticancer properties, its mechanisms of action have not been fully established. Ferroptosis, which is a novel anticancer strategy, is a cell death mechanism that relies on iron-dependent lipid reactive oxygen species (ROS) accumulation. In this study, human breast cancer cells (Michigan Cancer Foundation-7 (MCF-7) and MD Anderson-Metastatic Breast-231 (MDA-MB-231)) were treated with LBP. LBP inhibited their viability and proliferation in association with high levels of ferroptosis. Therefore, we aimed to ascertain whether LBP reduced cell viability through ferroptosis. We found that the structure and function of mitochondria, lipid peroxidation, and expression of solute carrier family 7 member 11 (SLC7A11, also known as xCT, the light-chain subunit of cystine/glutamate antiporter system X_c^-) and glutathione peroxidase 4 (GPX4) were altered by LBP. Moreover, the ferroptosis inhibitor, Ferrostatin-1 (Fer-1), rescued LBP-induced ferroptosis-associated events including reduced cell viability and glutathione (GSH) production, accumulation of intracellular free divalent iron ions and malondialdehyde (MDA), and down-regulation of the expression of xCT and GPX4. Erastin (xCT inhibitor) and RSL3 (GPX4 inhibitor) inhibited the expression of xCT and GPX4, respectively, which was lower after the co-treatment of LBP with Erastin and RSL3. These results suggest that LBP effectively prevents breast cancer cell proliferation and promotes ferroptosis via the xCT/GPX4 pathway. Therefore, LBP exhibits novel anticancer properties by triggering ferroptosis, and may be a potential therapeutic option for breast cancer.

枸杞多糖诱导乳腺癌细胞铁死亡的研究

[1]AbotalebM, KubatkaP, CaprndaM, et al., 2018. Chemotherapeutic agents for the treatment of metastatic breast cancer: an update. Biomed Pharmacother, 101:458-477.

[2]AiY, SunYN, LiuL, et al., 2021. Determination of biogenic amines in different parts of Lycium barbarum L. by HPLC with precolumn dansylation. Molecules, 26(4):1046.

[3]AmagaseH, FarnsworthNR, 2011. A review of botanical characteristics, phytochemistry, clinical relevance in efficacy and safety of Lycium barbarum fruit (Goji). Food Res Int, 44(7):1702-1717.

[4]ArziL, HoshyarR, JafarzadehN, et al., 2020. Anti-metastatic properties of a potent herbal combination in cell and mice models of triple negative breast cancer. Life Sci, 243:117245.

[5]BaiZ, PengY, YeX, et al., 2022. Autophagy and cancer treatment: four functional forms of autophagy and their therapeutic applications. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 23(2):89-101.

[6]BridgesRJ, NataleNR, PatelSA, 2012. System X_c^- cystine/glutamate antiporter: an update on molecular pharmacology and roles within the CNS. Br J Pharmacol, 165(1):20-34.

[7]ChenP, WuQB, FengJ, et al., 2020. Erianin, a novel dibenzyl compound in Dendrobium extract, inhibits lung cancer cell growth and migration via calcium/calmodulin-dependent ferroptosis. Signal Transduct Target Ther, 5:51.

[8]ChenQ, ShiRL, JiangDW, et al., 2017. Lycium barbarum polysaccharide inhibits gastric cancer cell proliferation, migration and invasion by down-regulation of MMPs and suppressing epithelial-mesenchymal transition. Int J Clin Exp Pathol, 10(7):7369-7374.

[9]CorsoCR, Mulinari Turin de Oliveira N, Moura CordeiroL, et al., 2021. Polysaccharides with antitumor effect in breast cancer: a systematic review of non-clinical studies. Nutrients, 13(6):2008.

[10]CraggGM, PezzutoJM, 2015. Natural products as a vital source for the discovery of cancer chemotherapeutic and chemopreventive agents. Med Princ Pract, 25(Suppl 2):41-59.

[11]CumaogluA, BekciH, OzturkE, et al., 2018. Goji berry fruit extracts suppress proliferation of triple-negative breast cancer cells by inhibiting EGFR-mediated ERK/MAPK and PI3K/Akt signaling pathways. Nat Prod Commun, 13(6):701-706.

[12]DengXL, LiXL, LuoS, et al., 2017. Antitumor activity of Lycium barbarum polysaccharides with different molecular weights: an in vitro and in vivo study. Food Nutr Res, 61:1399770.

[13]DengXL, LuoS, LuoX, et al., 2018. Polysaccharides from Chinese herbal Lycium barbarum induced systemic and local immune responses in H22 tumor-bearing mice. J Immunol Res, 2018:3431782.

[14]DiasDA, UrbanS, RoessnerU, 2012. A historical overview of natural products in drug discovery. Metabolites, 2(2):303-336.

[15]DixonSJ, LembergKM, LamprechtMR, et al., 2012. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell, 149(5):1060-1072.

[16]DuX, XiaoJJ, FuXF, et al., 2021. A proteomic analysis of Bcl-2 regulation of cell cycle arrest: insight into the mechanisms. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 22(10):839-855.

[17]FengL, XiaoX, LiuJ, et al., 2020. Immunomodulatory effects of Lycium barbarum polysaccharide extract and its uptake behaviors at the cellular level. Molecules, 25(6):1351.

[18]GanL, ZhangSH, YangXL, et al., 2004. Immunomodulation and antitumor activity by a polysaccharide‒protein complex from Lycium barbarum. Int Immunopharmacol, 4(4):563-569.

[19]GaoZW, DengGH, LiYJ, et al., 2020. Actinidia chinensis Planch prevents proliferation and migration of gastric cancer associated with apoptosis, ferroptosis activation and mesenchymal phenotype suppression. Biomed Pharmacother, 126:110092.

[20]García-ArandaM, RedondoM, 2019. Immunotherapy: a challenge of breast cancer treatment. Cancers (Basel), 11(12):1822.

[21]GeziciS, ŞekeroğluN, 2019. Current perspectives in the application of medicinal plants against cancer: novel therapeutic agents. Anti-Cancer Agents Med Chem, 19(1):101-111.

[22]GrecoG, CatanzaroE, FimognariC, 2021. Natural products as inducers of non-canonical cell death: a weapon against cancer. Cancers (Basel), 13(2):304.

[23]HuQH, GaoTS, ZhaoCJ, et al., 1994. The effect of active components of Lycium barbarum and garlic (LB-GO) on the synthesis of DNA and ultrastructure of U₁₄ cervix cancer cells in mice. Chin J Cancer Res, 6:266-273.

[24]IjazS, AkhtarN, KhanMS, et al., 2018. Plant derived anticancer agents: a green approach towards skin cancers. Biomed Pharmacother, 103:1643-1651.

[25]JinMM, ShiCZ, LiT, et al., 2020. Solasonine promotes ferroptosis of hepatoma carcinoma cells via glutathione peroxidase 4-induced destruction of the glutathione redox system. Biomed Pharmacother, 129:110282.

[26]KumarP, AggarwalR, 2016. An overview of triple-negative breast cancer. Arch Gynecol Obstet, 293(2):247-269.

[27]KwokSS, BuYS, LoACY, et al., 2019. A systematic review of potential therapeutic use of Lycium barbarum polysaccharides in disease. Biomed Res Int, 2019:4615745.

[28]LeeEJ, MoonGS, ChoiWS, et al., 2008. Naringin-induced p21WAF1-mediated G₁-phase cell cycle arrest via activation of the Ras/Raf/ERK signaling pathway in vascular smooth muscle cells. Food Chem Toxicol, 46(12):3800-3807.

[29]LiJ, CaoF, YinHL, HuangZJ, et al., 2020. Ferroptosis: past, present and future. Cell Death Discov, 11(2):88.

[30]LiuZX, LvXY, YangBW, et al., 2021. Tetrachlorobenzoquinone exposure triggers ferroptosis contributing to its neurotoxicity. Chemosphere, 264:128413.

[31]LouL, ChenG, ZhongB, et al., 2019. Lycium barbarum polysaccharide induced apoptosis and inhibited proliferation in infantile hemangioma endothelial cells via down-regulation of PI3K/AKT signaling pathway. Biosci Rep, 39(8):BSR20191182.

[32]LuoQ, LiZN, YanJ, et al., 2009. Lycium barbarum polysaccharides induce apoptosis in human prostate cancer cells and inhibits prostate cancer growth in a xenograft mouse model of human prostate cancer. J Med Food, 12(4):695-703.

[33]MaoF, XiaoBX, JiangZ, et al., 2011. Anticancer effect of Lycium barbarum polysaccharides on colon cancer cells involves G0/G1 phase arrest. Med Oncol, 28(1):121-126.

[34]MbavengAT, ChiGF, BonsouIN, et al., 2020. N-Acetylglycoside of oleanolic acid (aridanin) displays promising cytotoxicity towards human and animal cancer cells, inducing apoptotic, ferroptotic and necroptotic cell death. Phytomedicine, 76:153261.

[35]McDonaldER, El-DeiryWS, 2001. Checkpoint genes in cancer. Ann Med, 33(2):113-122.

[36]MiaoY, XiaoBX, JiangZ, et al., 2010. Growth inhibition and cell-cycle arrest of human gastric cancer cells by Lycium barbarum polysaccharide. Med Oncol, 27(3):785-790.

[37]NewmanDJ, CraggGM, 2020. Natural products as sources of new drugs over the nearly four decades from 01/1981 to 09/2019. J Nat Prod, 83(3):770-803.

[38]PatersonI, AndersonEA, 2005. The renaissance of natural products as drug candidates. Science, 310(5747):451-453.

[39]ShenLL, DuG, 2012. Lycium barbarum polysaccharide stimulates proliferation of MCF-7 cells by the ERK pathway. Life Sci, 91(9-10):353-357.

[40]ShinSS, HwangB, MuhammadK, et al., 2019. Nimbolide represses the proliferation, migration, and invasion of bladder carcinoma cells via Chk2-mediated G2/M phase cell cycle arrest, altered signaling pathways, and reduced transcription factors-associated MMP-9 expression. Evid-Based Complement Alternat Med, 2019:3753587.

[41]TaborS, Szostakowska-RodzosM, FabisiewiczA, et al., 2020. How to predict metastasis in luminal breast cancer? Current solutions and future prospects. Int J Mol Sci, 21(21):8415.

[42]TavsanZ, KayaliHA, 2019. Flavonoids showed anticancer effects on the ovarian cancer cells: involvement of reactive oxygen species, apoptosis, cell cycle and invasion. Biomed Pharmacother, 116:109004.

[43]WangW, LiuMX, WangY, et al., 2018. Lycium barbarum polysaccharide promotes maturation of dendritic cell via Notch signaling and strengthens dendritic cell mediated T lymphocyte cytotoxicity on colon cancer cell CT26-WT. Evid Based Complement Alternat Med, 2018:2305683.

[44]WHO, 1999. WHO Monographs on Selected Medicinal Plants. World Health Organization, Geneva.

[45]XuJ, WuKJ, JiaQJ, et al., 2020. Roles of miRNA and lncRNA in triple-negative breast cancer. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 21(9):673-689.

[46]YangWS, SriRamaratnamR, WelschME, et al., 2014. Regulation of ferroptotic cancer cell death by GPX4. Cell, 156(1-2):317-331.

[47]YangWS, KimKJ, GaschlerMM, et al., 2016. Peroxidation of polyunsaturated fatty acids by lipoxygenases drives ferroptosis. Proc Natl Acad Sci USA, 113(34):‍E4966-E4975.

[48]YuXL, LongYC, 2016. Crosstalk between cystine and glutathione is critical for the regulation of amino acid signaling pathways and ferroptosis. Sci Rep, 6:30033.

[49]ZengMH, KongQT, LiuF, et al., 2019. The anticancer activity of Lycium barbarum polysaccharide by inhibiting autophagy in human skin squamous cell carcinoma cells in vitro and in vivo. Int J Polym Sci, 2019:5065920.

[50]ZhangM, ChenHX, HuangJ, et al., 2005. Effect of Lycium barbarum polysaccharide on human hepatoma QGY7703 cells: inhibition of proliferation and induction of apoptosis. Life Sci, 76(18):2115-2124.

[51]ZhangQ, LvXL, WuT, et al., 2015. Composition of Lycium barbarum polysaccharides and their apoptosis-inducing effect on human hepatoma SMMC-7721 cells. Food Nutr Res, 59:28696.

[52]ZhangXJ, YuHY, CaiYJ, et al., 2017. Lycium barbarum polysaccharides inhibit proliferation and migration of bladder cancer cell lines BIU87 by suppressing Pi3K/AKT pathway. Oncotarget, 8(4):5936-5942.

[53]ZhaoJC, JinY, YanYM, et al., 2020. Herbal textual research on “Lycii Fructus” and “Lycii Cortex” in Chinese classical prescriptions. Mod Chin Med, 22(8):‍1269-1286 (in Chinese).

[54]ZhuCP, ZhangSH, 2006. The antitumor and immunoenhancement activity of Lycium barbarum polysaccharides in hepatoma H₂₂-bearing mice. Acta Nutr Sin, 28(2):182-183 (in Chinese).

[55]ZhuCP, ZhangSH, 2013. Lycium barbarum polysaccharide inhibits the proliferation of HeLa cells by inducing apoptosis. J Sci Food Agric, 93(1):149-156.