目的 通過網(wǎng)絡(luò)藥理學(xué)和分子對(duì)接技術(shù)分析鉤吻素甲（GEL）抗舌鱗狀細(xì)胞癌（TSCC）的靶點(diǎn)及作用機(jī)制，并通過實(shí)驗(yàn)驗(yàn)證GEL對(duì)人舌鱗癌CAL27細(xì)胞增殖、凋亡的影響及關(guān)鍵靶點(diǎn)的作用。方法 運(yùn)用網(wǎng)絡(luò)藥理學(xué)預(yù)測(cè)GEL和TSCC的共有靶點(diǎn)，繪制蛋白質(zhì)-蛋白質(zhì)相互作用（PPI）網(wǎng)絡(luò)圖，并進(jìn)行基因本體（GO）功能富集分析和京都基因與基因組百科全書（KEGG）通路富集分析，使用Autodock vina軟件進(jìn)行分子對(duì)接。通過Incucyte S3活細(xì)胞動(dòng)態(tài)分析系統(tǒng)觀察GEL作用下CAL27細(xì)胞形態(tài)，并擬合半數(shù)抑制濃度（IC₅₀）值，采用細(xì)胞增殖與活性檢測(cè)-8（CCK-8）實(shí)驗(yàn)、平板克隆實(shí)驗(yàn)、細(xì)胞周期實(shí)驗(yàn)檢測(cè)細(xì)胞增殖能力；通過Hoechst 33258染色、Rhodamine 123染色檢測(cè)細(xì)胞凋亡情況，Western blotting檢測(cè)B淋巴細(xì)胞瘤2基因相關(guān)X基因（Bax）、B淋巴細(xì)胞瘤2基因（Bcl-2）、含半胱氨酸的蛋白水解酶3基因（Caspase-3）蛋白表達(dá)。結(jié)果 篩選出藥物-疾病共有靶點(diǎn)49個(gè)，核心靶點(diǎn)為雌激素受體1（ESR1）、含半胱氨酸的蛋白水解酶3基因（CASP3）、基質(zhì)金屬蛋白酶9（MMP-9）、B淋巴細(xì)胞瘤2基因樣1（BCL2L1）、非受體酪氨酸激酶（SRC），KEGG通路富集較高的為癌癥途徑、PI3K/Akt通路等。分子對(duì)接結(jié)合良好，其中GEL與CASP3和BCL2L1有強(qiáng)結(jié)合性。體外實(shí)驗(yàn)顯示，GEL可以抑制CAL27細(xì)胞增殖并阻滯細(xì)胞周期于G₂/M期，GEL干預(yù)下CAL27細(xì)胞染色質(zhì)固縮，線粒體膜電位降低，Bax、Caspase-3蛋白表達(dá)增多，Bcl-2蛋白表達(dá)降低。結(jié)論 GEL通過多靶點(diǎn)、多途徑抑制人舌鱗癌CAL27細(xì)胞增殖并促進(jìn)其凋亡，可能與激活Bax/Bcl-2/Caspase-3通路有關(guān)。

Objective To investigate the targets and mechanism of action of GEL against tongue squamous cell carcinoma (TSCC) by network pharmacology and molecular docking techniques, and to verify the effects of GEL on the proliferation, apoptosis of human tongue squamous carcinoma CAL27 cells and the effects of key targets by experimental methods. Methods The shared targets of GEL and TSCC were predicted by network pharmacology, and the PPI network diagram was drawn. GO function and KEGG pathway enrichment analysis were performed, and molecular docking was performed using Autodock vina software. The cell morphology of CAL27 cells under GEL action was observed by Incucyte S3 live cell dynamic analysis system and the IC₅₀ value was fitted. The cell proliferation ability was detected by CCK-8 experiment, plate colony experiment and cell cycle experiment. The cell apoptosis was detected by Hoechst 33258 staining and Rhodamine 123 staining, and the expression of Bax, Bcl-2 and Caspase-3 proteins was detected by Western blotting. Results A total of 49 common targets were screened, including ESR1, CASP3, MMP-9, BCL2L1 and SRC, with the KEGG pathway enrichment being higher in cancer pathway, PI3K/Akt pathway, etc. The molecular docking was combined well, with GEL having a strong binding to CASP3 and BCL2L1. The in vitro experiment showed that GEL could inhibit the proliferation of CAL27 cells and block the cell cycle at the G₂/M phase. Under GEL intervention, the chromatin condensation of CAL27 cells was observed, the mitochondrial membrane potential was lowered, the expression of Bax, Caspase-3 proteins increased, and the expression of Bcl-2 protein decreased. Conclusion GEL can inhibit the proliferation of CAL27 cells through multiple targets and pathways, promote their apoptosis, and may be related to activating the Bax/Bcl-2/Caspase-3 pathway.

R285.5

廣西自然科學(xué)基金資助項(xiàng)目（2018GXNSFDA050009）；廣西中醫(yī)藥大學(xué)博士啟動(dòng)基金項(xiàng)目（2020BS034）；廣西國(guó)際壯醫(yī)醫(yī)院引進(jìn)人才科研啟動(dòng)基金項(xiàng)目（GZ2021RC016）