乙齐楚谈方（YQCTF）是一种包含八味中药的处方，该处方已在临床上被证明对非小细胞肺癌（NSCLC）患者的生活质量和生存时间具有改善的有效性。本研究旨在评估YQCTF在NSCLC小鼠模型中的治疗效果，并通过网络药理学、蛋白质组学和药效学方法进一步探索其治疗靶点。首先进行网络药理学分析，筛选出YQCTF对NSCLC的潜在活性成分和治疗靶点。制备了YQCTF的水提取物（WE）、水提取物-醇沉淀物（WEAP）和醇提取物（AE）共三种提取物，并利用超高效液相色谱-质谱/质谱（UPLC-MS/MS）分析了其化学组成，并在NSCLC小鼠模型上检测了其抗肿瘤效力。收集小鼠肿瘤组织进行蛋白质组学分析，并利用流式细胞术、免疫荧光、酶联免疫吸附法（ELISA）和Western blot进一步验证了YQCTF提取物对肿瘤微环境（TME）的免疫调节效果。 网络药理学确定了60个共同基因和许多与癌症相关的信号通路，作为YQCTF潜在治疗靶点。蛋白质-蛋白质相互作用（PPI）、基因本体论（GO）和Kyoto基因和基因组百科全书（KEGG）分析表明，YQCTF可能通过负调节与癌症相关的炎症来发挥作用。UPLC-MS/MS分析显示，YQCTF的主要成分至少包括人参皂苷、曼红素、曼红嵩、曼红嵌、鄂固醇、鄂湿固醇、鄂白鄂和鄂苷-3β-D-葡萄糖苷。所有种类的YQCTF提取物都显著抑制了肺癌异基因移植物的生长，并调节了肿瘤组织中免疫细胞的比例，即上调了T细胞的比例，促进了树突状细胞（DCs）的成熟，增加了与肿瘤相关的巨噬细胞的M1/M2比例，但降低了Treg和免疫抑制性中性粒细胞的数量。蛋白质组学发现中性粒细胞是小鼠肿瘤组织中最显著富集的与NETs形成相关的靶点，这在免疫荧光、ELISA和Western blot分析中通过中性粒细胞招募因子IL-6、HIF-1α和IL-8的下调以及NETs相关生物标志物H3cit、MPO、CD18、MMP9和ICAM-1的减少得到验证。 YQCTF抑制了小鼠NSCLC异基因移植物的进展，抑制了促癌的肿瘤相关中性粒细胞，并改善了肿瘤免疫微环境（TIME）。 版权所有 © 2023 Elsevier B.V. 发表。

Yi Qi Chu Tan Formula (YQCTF), a prescription consisting of eight traditional Chinese medicine for treating lung cancer, has been clinically proven to be effective in improving the life quality and prolonging the survival time of non-small cell lung cancer (NSCLC) patients.This study aimed to evaluate the therapeutic efficacy of YQCTF on NSCLC mice model and further explore its therapeutic targets by using network pharmacology, proteomics and pharmacodynamic methodologies.The network pharmacology analysis was firstly conducted to screen out the potential active ingredients and therapeutic targets of YQCTF against NSCLC. Three kinds of extracts, i.e. the water extract (WE), water extraction-alcohol precipitation (WEAP) and alcohol extract (AE) of YQCTF were prepared, which chemical compositions were subsequently analyzed by using ultra-high performance liquid chromatography-mass spectrometry/mass spectrometry (UPLC-MS/MS), and which anti-neoplastic efficacy was examined on NSCLC mice model. Mice tumor tissues were collected for proteomics analysis, and the immunomodulatory effects of YQCTF extracts on the tumor microenvironment (TME) were further validated by using flow cytometry, immunofluorescence, ELISA and Western blot.Network pharmacology identified 60 conjunct genes and ample cancer-related signaling pathways as potential therapeutic targets of YQCTF. Protein-protein interaction (PPI), Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that YQCTF might negatively regulate cancer-related inflammation. UPLC-MS/MS analysis showed that the main components of YQCTF include at least ginsenosides, solasodine, solamargine, solasonine, peimisine, peiminine, peimine and sipeimine-3β-D-glucosihde. All kinds of YQCTF extracts significantly inhibited the growth of lung cancer allograft and regulated the ratio of immune cells in tumor tissues, i.e. upregulated the fractions of T cells, promoted the maturation of dendritic cells (DCs), increased the M1/M2 ratio of tumor-related macrophages, but reduced the number of Tregs and immunosuppressive neutrophils. Proteomics identified neutrophils to be the most prominently enriched target linked to NETs formation in mice tumor tissue, which is verified by the downregulation of neutrophil recruiting factors involving IL-6, HIF-1α and IL-8, as well as the decreases of NETs-related biomarkers including H3cit, MPO, CD18, MMP9 and ICAM-1 in immunofluorescence, ELISA and Western blot analysis.YQCTF inhibited the progress of mice NSCLC allograft, suppressed the pro-tumorigenic tumor-associated neutrophils and improved the tumor immune microenvironment (TIME).Copyright © 2023. Published by Elsevier B.V.