目的 探究非諾貝特抗腎透明細(xì)胞癌（ccRCC）的作用及潛在機(jī)制。方法 通過在線數(shù)據(jù)庫Swiss Target Prediction、Pharm Mapper、Target Net、Gene Cards、OMIM、TTD等分析非諾貝特、ccRCC和凋亡相關(guān)靶點(diǎn)，STRING數(shù)據(jù)庫構(gòu)建蛋白質(zhì)-蛋白質(zhì)相互作用（PPI）網(wǎng)絡(luò)并通過Cytoscape進(jìn)行可視化，采用Metascapep平臺進(jìn)行基因本體（GO）和京都基因與基因組百科全書（KEGG）富集分析，通過Auto Dock Tools和PyMOL軟件進(jìn)行分子對接。采用CCK-8法檢測非諾貝特對人腎透明細(xì)胞癌786-O細(xì)胞株、人腎皮質(zhì)近曲小管上皮細(xì)胞株（HK-2）活力的影響；劃痕實(shí)驗(yàn)、克隆形成實(shí)驗(yàn)和流式細(xì)胞術(shù)分別檢測非諾貝特（50、75、100 μmol· L^-1）對786-O細(xì)胞遷移、增殖、細(xì)胞周期的影響；Hoechst33258染色和流式細(xì)胞術(shù)檢測非諾貝特對786-O細(xì)胞凋亡的影響；Western bloting檢測非諾貝特對786-O細(xì)胞信號轉(zhuǎn)導(dǎo)與轉(zhuǎn)錄激活因子3（STAT3）、p-STAT3、B細(xì)胞淋巴瘤2蛋白（Bcl-2）、Bcl-2關(guān)聯(lián)X蛋白（Bax）、半胱氨酸天冬氨酸蛋白酶-3（Caspase-3）蛋白表達(dá)的影響。結(jié)果 網(wǎng)絡(luò)藥理學(xué)分析獲得非諾貝特、ccRCC和凋亡共同靶點(diǎn)108個，按照網(wǎng)絡(luò)拓?fù)浞治鼋Y(jié)果中的度（degree）值篩選出主要核心靶點(diǎn)為STAT3、EGFR、MMP-9、PPARG、RELA、BCL2L；KEGG富集分析表明JAK2-STAT3信號通路在抗ccRCC中發(fā)揮重要作用；分子對接結(jié)果顯示非諾貝特與主要核心靶點(diǎn)STAT3、EGFR、MMP9、PPARG、RELA、BCL2L具有良好的結(jié)合活性。體外實(shí)驗(yàn)結(jié)果表明，與對照組比較，非諾貝特呈濃度和時間相關(guān)性抑制786-O和HK-2細(xì)胞的生長（P＜0.01、0.001），且對786-O細(xì)胞活力的抑制作用強(qiáng)于HK-2細(xì)胞；可將786-O細(xì)胞周期阻滯在G₁期（P＜0.001）；顯著抑制786-O細(xì)胞集落形成、遷移能力（P＜0.01、0.001）；顯著誘導(dǎo)786-O細(xì)胞凋亡（P＜0.001）；顯著下調(diào)p-STAT和Bcl-2、上調(diào)Bax和Caspase-3蛋白表達(dá)（P＜0.05）。結(jié)論 非諾貝特可能通過下調(diào)STAT3抑制ccRCC細(xì)胞增殖遷移，誘導(dǎo)凋亡，進(jìn)而發(fā)揮抗ccRCC的作用。

Objective To explore the role and potential mechanism of fenofibrate in the treatment of renal clear cell carcinoma (ccRCC). Methods The online databases such as Swiss Target Prediction, Pharm Mapper, Target Net, Gene Cards, OMIM, TTD were used to analyze the targets of fenofibrate, ccRCC and apoptosis. The STRING database was applied to construct a proteinprotein interaction (PPI) network and visualized using Cytoscape. Molecular function (GO) and pathway (KEGG) enrichment analysis was performed using the Metascapep platform, and Auto Dock Tools and PyMOL software were used for molecular docking. The CCK-8 method was used to detect the effect of fenofibrate on the viability of human renal clear cell carcinoma 786-O cell line and human renal cortical proximal tubular epithelial cell line (HK-2). Scratch assay, clone formation assay, and flow cytometry were used to detect the effects of fenofibrate (50, 75, and 100 μmol·L^-1) on the migration, proliferation, and cell cycle of 786-O cells, respectively. Hoechst33258 staining and flow cytometry were used to detect the effect of fenofibrate on the apoptosis level of 786-O cells. Western blotting was used to detect the effects of fenofibrate on the expression of signal transduction and transcription activator 3 (STAT3), p-STAT3, B-cell lymphoma 2 protein (Bcl-2), Bcl-2 associated X protein (Bax), and Caspase-3 protein in 786-O cells. Results Network pharmacology analysis identified 108 common targets of fenofibrate, ccRCC and apoptosis. Based on the degree values in network topology analysis, the main core targets were selected as STAT3, EGFR, MMP-9, PPARG, RELA and BCL2L. KEGG enrichment analysis revealed that the JAK2-STAT3 signaling pathway played an important role in anticcRCC. The molecular docking results showed that fenofibrate had good binding activity with the main core targets STAT3, EGFR, MMP9, PPARG, RELA and BCL2L. The in vitro experimental results showed that compared with the control group, fenofibrate inhibited the growth of 786-O and HK-2 cells in a concentration - and time-dependent manner (P <0.01, 0.001), and had a stronger inhibitory effect on the viability of 786-O cells than HK-2 cells, 786-O cell cycle could be arrested in G₁ phase (P <0.001), significantly inhibited the colony formation and migration ability of 786-O cells (P <0.01, 0.001), significantly induced apoptosis in 786-O cells (P <0.001), significantly downregulated p-STAT and Bcl-2, upregulated Bax and Caspase-3 protein expression (P <0.05). Conclusion Fenofibrate inhibited the proliferation and migration of ccRCC cells, induced apoptosis, and exerted anti-ccRCC effect by downregulation of STAT3.

R965

國家自然科學(xué)基金資助項(xiàng)目（82274159）;湖南省自然科學(xué)基金科藥聯(lián)合基金項(xiàng)目（2022JJ80088）;湖南省衛(wèi)生健康委員會重點(diǎn)指導(dǎo)課題（202213055529）;湖南省自然科學(xué)基金青年項(xiàng)目（2023JJ40485）;湖南省教育廳優(yōu)秀青年項(xiàng)目（23B0387、22B0355）;中藥粉體與創(chuàng)新藥物省部共建國家重點(diǎn)實(shí)驗(yàn)室培育基地開放基金項(xiàng)目（23PTKF1016）;藥學(xué)一流學(xué)科開放基金項(xiàng)目（研究生創(chuàng)新性實(shí)驗(yàn)計(jì)劃重點(diǎn)支持項(xiàng)目，2021YX07）;湖南省藥學(xué)一流學(xué)科資助項(xiàng)目