非小细胞肺癌（NSCLC）是最常见的肺癌类型，发病率很高。本研究旨在揭示小檗碱的抗NSCLC机制并确定新的治疗靶点。小檗碱相关靶点来自SuperPred、SwissTargetPrediction和GeneCards。 NSCLC 相关目标是从 GeneCards 和 DisGeNET 收集的。 GEO 数据库、UCSC Xena 和 limma 鉴定了差异表达基因 (DEG)。使用 clusterProfiler 进行 GO 和 KEGG 分析。自噬相关基因和转录因子分别从HADb和KnockTF收集。 STRING 和 Cytoscape 用于 PPI 网络分析。使用 CIBERSORT 评估 NSCLC 中的免疫细胞浸润，并评估其与自噬相关靶点的相关性。使用 PyMOL 和 AutoDock 进行分子对接。采用qRT-PCR和CCK-8法进行体外验证。获得了小檗碱相关靶点、NSCLC相关靶点和DEGs的30个交叉靶点。 GO和KEGG分析显示，交叉靶标主要涉及氧化应激、粘着斑和细胞-基质连接，以及AGE-RAGE、松弛素、FoxO和雌激素信号通路。值得注意的是，CAPN1、IKBKB 和 SIRT2 被确定为最重要的自噬相关靶点，并获得了 21 个相应的转录因子。 PPI 网络分析表明 CAPN1、IKBKB 和 SIRT2 与其他 50 个基因相互作用。 B细胞幼稚、T细胞CD8、T细胞CD4幼稚、滤泡辅助T细胞和单核细胞等50种免疫细胞类型与NSCLC发病机制有关，其中CAPN1、IKBKB和SIRT2与免疫细胞相关。分子对接显示小檗碱与 CAPN1、IKBKB 和 SIRT2 具有良好的结合活性。体外检测显示 NSCLC 细胞中 CAPN1、IKBKB 和 SIRT2 的表达低于正常细胞。值得注意的是，小檗碱抑制NSCLC细胞的活力并升高CAPN1、IKBKB和SIRT2的表达。小檗碱可能主要通过靶向CAPN1、IKBKB和SIRT2来治疗NSCLC。

Non-small cell lung cancer (NSCLC) is the most common lung cancer type with high incidence. This study aimed to reveal the anti-NSCLC mechanisms of berberine and identify novel therapeutic targets.Berberine-related targets were acquired from SuperPred, SwissTargetPrediction, and GeneCards. NSCLC-re-lated targets were collected from GeneCards and DisGeNET. Differentially expressed genes (DEGs) were identified GEO database, UCSC Xena, and limma. GO and KEGG analyses were performed using clusterProfiler. Autophagy-related genes and transcriptional factors were collected from HADb and KnockTF, respectively. STRING and Cytoscape were used for PPI network analysis. Immune cell infiltration in NSCLC was assessed using CIBERSORT, and its correlation with autophagy-related targets was evaluated. Molecular docking was conducted using PyMOL and AutoDock. qRT-PCR and CCK-8 assay was used for in vitro verification.Thirty intersecting targets of berberine-related targets, NSCLC-related targets, and DEGs were obtained. GO and KEGG analyses revealed that the intersecting targets were mainly implicated in oxidative stress, focal adhesion, and cell-substrate junction, as well as AGE-RAGE, relaxin, FoxO, and estrogen signaling pathways. Significantly, CAPN1, IKBKB, and SIRT2 were identified as the foremost autophagy-related targets, and 21 corresponding transcriptional factors were obtained. PPI network analysis showed that CAPN1, IKBKB, and SIRT2 interacted with 50 other genes. Fifty immune cell types, such as B cells naive, T cells CD8, T cells CD4 naive, T cells follicular helper, and monocytes, were implicated in NSCLC pathogenesis, and CAPN1, IKBKB, and SIRT2 were related to immune cells. Molecular docking revealed the favorable binding activity of berberine with CAPN1, IKBKB, and SIRT2. In vitro assays showed lower CAPN1, IKBKB, and SIRT2 expression in NSCLC cells than that in normal cells. Notably, berberine inhibited the viability and elevated CAPN1, IKBKB, and SIRT2 expression in NSCLC cells.Berberine might treat NSCLC mainly by targeting CAPN1, IKBKB, and SIRT2.