陆颖颖;黄枭;罗子宸;祁明媛;单进军;张雯;狄留庆;
LU Ying-ying;HUANG Xiao;LUO Zi-chen;QI Ming-yuan;SHAN Jin-jun;ZHANG Wen;DI Liu-qing;School of Pharmacy, Nanjing University of Chinese Medicine;Jiangsu Engineering Research Center for Efficient Delivery System of TCM;Jiangsu Key Laboratory of Pediatric Respiratory Disease, Institute of Pediatrics, Nanjing University of Chinese Medicine;Hunan Yandi Biological Engineering Co., Ltd.;

• 1:南京中医药大学药学院
• 2:江苏省中药高效给药系统工程技术研究中心
• 3:南京中医药大学中医儿科学研究所江苏省儿童呼吸疾病(中医药)重点实验室
• 4:湖南炎帝生物工程有限公司

摘要(Abstract)：

采用血清非靶向代谢组学技术研究蛹虫草抗非小细胞肺癌药效作用的潜在机制。以Balb/c裸鼠构建人肺癌荷瘤裸鼠移植瘤模型，比较模型组与顺铂组，冬虫夏草低、中、高剂量组，蛹虫草低、中、高剂量组中移植瘤裸鼠的肿瘤体积、肿瘤质量及抑瘤率，以评价蛹虫草对肺癌生长的影响。收集移植瘤裸鼠血清样本，使用气相色谱-质谱(gas chromatography-mass spectrometry, GC-MS)仪器进行检测，运用SIMCA 13.0软件开展血清代谢轮廓比较，采用MetaboAnalyst 5.0探究相关代谢通路。药效学数据表明，与模型组相比，蛹虫草高剂量组和顺铂组裸鼠的肿瘤体积与肿瘤组织质量显著减小(P<0.05或P<0.01)。代谢组学结果显示，蛹虫草组与模型组裸鼠血清的代谢轮廓具有明显差异，其内源性代谢物含量受不同程度的调整，初步鉴定出42个差异代谢物及7条代谢通路。综上，蛹虫草能够显著抑制人肺癌移植瘤裸鼠肿瘤的生长，其潜在作用机制可能与影响氨酰-tRNA合成，谷氨酰胺和谷氨酸代谢，丙氨酸、天冬氨酸和谷氨酸代谢，乙醛酸和二羧酸代谢，苯丙氨酸、酪氨酸和色氨酸生物合成，精氨酸生物合成及氮代谢等通路密切相关。该研究从代谢物角度阐释了蛹虫草抗非小细胞肺癌的作用机制，为蛹虫草抗肿瘤研究及临床应用奠定基础。
This study investigated the potential mechanism of Cordyceps militaris(CM) against non-small cell lung cancer(NSCLC) based on serum untargeted metabolomics. Specifically, Balb/c nude mice were used to generate the human lung cancer A549 xenograft mouse model. The tumor volume, tumor weight, and tumor inhibition rate in mice in the model, cisplatin, Cordyceps(low-, medium-, and high-dose), and CM(low-, medium-, and high-dose) groups were compared to evaluate the influence of CM on lung cancer. Gas chromatography-mass spectrometry(GC-MS) was used for the analysis of mouse serum, SIMCA 13.0 for the compa-rison of metabolic profiles, and MetaboAnalyst 5.0 for the analysis of metabolic pathways. According to the pharmacodynamic data, the tumor volume and tumor weight of mice in high-dose CM group and cisplatin group decreased as compared with those in the model group(P<0.05 or P<0.01). The results of serum metabolomics showed that the metabolic profiles of the model group were significantly different from those of the high-dose CM group, and the content of endogenous metabolites was adjusted to different degrees. A total of 42 differential metabolites and 7 differential metabolic pathways were identified. In conclusion, CM could significantly inhibit the tumor growth of lung cancer xenograft mice. The mechanism is the likelihood that it influences the aminoacyl-tRNA biosynthesis, the metabolism of D-glutamine and D-glutamate, metabolism of alanine, aspartate, and glutamate, metabolism of glyoxylate and dicarboxylic acid, biosynthesis of phenylalanine, tyrosine, and tryptophan, arginine biosynthesis as well as nitrogen metabolism. This study elucidated the underlying mechanism of CM against NSCLC from the point of metabolites. The results would lay a foundation for the anticancer research and clinical application of CM.

关键词(KeyWords)： 蛹虫草;冬虫夏草;肿瘤;非小细胞肺癌;血清代谢组学
Cordyceps militaris;Cordyceps;tumor;non-small cell lung cancer;serum metabolomics

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金项目(82074024,82104688);; 江苏省自然科学基金项目(BK20190800);; 江苏省高等学校自然科学研究项目(19KJB360004);; 南京中医药大学自然科学基金项目(XPT82104688)

作者(Authors): 陆颖颖;黄枭;罗子宸;祁明媛;单进军;张雯;狄留庆;
LU Ying-ying;HUANG Xiao;LUO Zi-chen;QI Ming-yuan;SHAN Jin-jun;ZHANG Wen;DI Liu-qing;School of Pharmacy, Nanjing University of Chinese Medicine;Jiangsu Engineering Research Center for Efficient Delivery System of TCM;Jiangsu Key Laboratory of Pediatric Respiratory Disease, Institute of Pediatrics, Nanjing University of Chinese Medicine;Hunan Yandi Biological Engineering Co., Ltd.;

DOI: 10.19540/j.cnki.cjcmm.20220613.702

参考文献(References)：

• [1] RITTMEYER A,BARLESI F,WATERKAMP D,et al.Atezolizumab versus docetaxel in patients with previously treated non-small-cell lung cancer (OAK):a phase 3,open-label,multicentre randomised controlled trial[J].Lancet,2017,389 (10066):255.
• [2] SIEGEL R L,MILLER K D,FUCHS H E,et al.Cancer Statistics,2021[J].CA Cancer J Clin,2021,71 (1):7.
• [3] HWANG J H,PARK S J,KO W G,et al.Cordycepin induces human lung cancer cell apoptosis by inhibiting nitric oxide mediated ERK/Slug signaling pathway[J].Am J Cancer Res,2017,7 (3):417.
• [4] LIU X C,ZHU Z Y,LIU Y L,et al.Comparisons of the anti-tumor activity of polysaccharides from fermented mycelia and cultivated fruiting bodies of Cordyceps militaris in vitro[J].Int J Biol Macromol,2019,130:307.
• [5] ZHANG J,WEN C,DUAN Y,et al.Advance in Cordyceps militaris (Linn) Link polysaccharides:isolation,structure,and bioactivities:a review[J].Int J Biol Macromol,2019,132,906.
• [6] YOON S Y,PARK S J,PARK Y J,et al.The anticancer pro-perties of Cordycepin and their underlying mechanisms[J].Int J Mol Sci,2018,19 (10):3027.
• [7] HAN E S,OH J Y,PARK H J,et al.Cordyceps militaris extract suppresses dextran sodium sulfate-induced acute colitis in mice and production of inflammatory mediators from macrophages and mast cells[J].J Ethnopharmacol,2011,134 (3):703.
• [8] 都兴范，李军，米锐，等.蛹虫草和冬虫夏草主要活性成分含量比较[J].食用菌，2010,32 (6):61.
• [9] 中华人民共和国卫生部.关于批准蛹虫草为新资源食品的公告[EB/OL].(2009-03-18)[2022-03-15].http://www.nhc.gov.cn/sps/s7891/200903/3b1e7201d2924d94899ea6f896a7d765.shtml.
• [10] 中共中央国务院印发《“健康中国2030”规划纲要》[J].中华人民共和国国务院公报，2016(32):5.
• [11] 吕尚，杨世林，饶毅，等.代谢组学及其相关技术在中药研发领域的应用进展[J].中国中药杂志，2018,43(21):4182.
• [12] SONG J,WANG Y,TENG M,et al.Cordyceps militaris induces tumor cell death via the caspase-dependent mitochondrial pathway in HepG2 and MCF-7 cells[J].Mol Med Rep,2016,13 (6):5132.
• [13] BIZARRO A,FERREIRA I C,SOKOVI■,et al.Cordyceps militaris (L.) link fruiting body reduces the growth of a non-small cell lung cancer cell line by increasing cellular levels of p53 and p21[J].Molecules,2015,20 (8):13927.
• [14] 钱文娟，杨瑞，谢彤，等.蒲地蓝消炎口服液治疗甲型H1N1流感病毒性肺炎的GC-MS代谢组学研究[J].中草药，2018,49 (10):2258.
• [15] 彭琳秀，谢彤，单进军，等.基于气相色谱-质谱联用的抗结核药致肾损伤代谢组学及谷胱甘肽治疗作用研究[J].分析化学，2020,48 (9):1160.
• [16] 袁嘉瑞，汪春飞，宋捷，等.淫羊藿醇提物抑制肺癌生长及调节免疫作用研究[J].中国中药杂志，2016,41(1):112.
• [17] HO P C,LIU P S.Metabolic communication in tumors:a new layer of immunoregulation for immune evasion[J].J Immunother Cancer,2016,4:4.
• [18] SCHMIDT D R,PATEL R,KIRSCH D G,et al.Metabolomics in cancer research and emerging applications in clinical oncology[J].CA Cancer J Clin,2021,71 (4):333.
• [19] HOSIOS A M,HECHT V C,DANAI L V,et al.Amino acids rather than glucose account for the majority of cell mass in proli-ferating mammalian cells[J].Dev Cell,2016,36 (5):540.
• [20] ZHAO J S,SHI S,QU H Y,et al.Glutamine synthetase licenses APC/C-mediated mitotic progression to drive cell growth[J].Nat Metab,2022,4(2):239.
• [21] CAPPELLETTI P,TALLARITA E,RABATTONI V,et al.Proline oxidase controls proline,glutamate,and glutamine cellular concentrations in a U87 glioblastoma cell line[J].PLoS ONE,2018,13 (4):e0196283.
• [22] VAZQUEZ A,KAMPHORST J J,MARKERT E K,et al.Can-cer metabolism at a glance[J].J Cell Sci,2016,129 (18):3367.
• [23] LIBERTI M V,LOCASALE J W.The Warburg effect:how does it benefit cancer cells?[J].Trends Biochem Sci,2016,41 (3):287.
• [24] METALLO C M,GAMEIRO P A,BELL E L,et al.Reductive glutamine metabolism by IDH1 mediates lipogenesis under hypo-xia[J].Nature,2011,481 (7381):380.
• [25] REINFELD B I,MADDEN M Z,WOLF M M,et al.Cell-programmed nutrient partitioning in the tumor microenvironment[J].Nature,2021,593 (7858):282.
• [26] MOHAMED A,DENG X,KHURI F R,et al.Altered glutamine metabolism and therapeutic opportunities for lung cancer[J].Clin Lung Cancer,2014,15 (1):7.
• [27] 李静阳，向丽，孟宪丽.中药调控谷氨酰胺代谢重编程在治疗肺癌中的潜力[J].中国药理学与毒理学杂志，2021,35 (10):743.
• [28] ALTMAN B J,STINE Z E,DANG C V,From Krebs to clinic:glutamine metabolism to cancer therapy[J].Nat Rev Cancer,2016,16 (12):773.
• [29] RAJENDRAN V,KALITA P,SHUKLA H,et al.Aminoacyl-tRNA synthetases:structure,function,and drug discovery[J].Int J Biol Macromol,2018,111:400.
• [30] LONG X,SONG K,HU H,et al.Long non-coding RNA GAS5 inhibits DDP-resistance and tumor progression of epithelial ova-rian cancer via GAS5-E2F4-PARP1-MAPK axis[J].J Exp Clin Cancer Res,2019,38 (1):345.
• [31] VANDER H M G,DEBERARDINIS R J.Understanding the intersections between metabolism and cancer biology[J].Cell,2017,168 (4):657.
• [32] GARCIA-BERMUDEZ J,BAUDRIER L,LA K,et al.Aspartate is a limiting metabolite for cancer cell proliferation under hypoxia and in tumors[J].Nat Cell Biol,2018,20 (7):775.
• [33] 吕雅鑫，陈龙，蒋义鑫，等.淫羊藿苷抑制非小细胞肺癌生长的代谢组学研究[J].上海中医药杂志，2021,55(6):84.