Table1_Dual COX-2/5-LOX inhibitors from Zanthoxylum simulans inhibit gastric cancer cells by cross-mediating thyroid, estrogen, and oxytocin signaling pathways.XLSX

Cyclooxygenase 2 (COX-2) and 5-lipoxygenase (5-LOX) are overexpressed in gastric cancer cells, the dual inhibitors of which exhibit potential against metastasis and invasion with fewer side effects. To discover inhibitors targeting COX-2 and 5-LOX, we conducted ultrafiltration and enrichment calculation to screen candidates in quaternary alkaloids (QAs) from Zanthoxylum simulans through LC and LC-Q-TOF. For intensive peaks, peaks 19 (berberine) and 21 (chelerythrine) were observed as the most potent dual candidates and showed selective affinity to 5-LOX over COX-2. Peak 19 showed an enrichment at 4.36 for COX-2 and 22.81 for 5-LOX, while peak 21 showed an enrichment at 7.81 for COX-2 and 24.49 for 5-LOX. Molecular docking results revealed chelerythrine as a better dual inhibitor, showing time- and dose-dependent anti-proliferation against AGS cells. Bio-informatics strategies, such as Gene Expression Omnibus (GEO), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG), suggested that hormone pathways in gastric cancer cells might be mediated by chelerythrine. Further reviews and summaries helped outline the mechanisms by which COX-2/5-LOX inhibitors might promote apoptosis in gastric cancer cells via estrogen, thyroid, and oxytocin signaling pathways. Chelerythrine was also added to gastric cancer cells to verify the regulation of these three signaling pathways. As a result, significant calling back of thyroid-stimulating hormone receptor (TSHR), thyroid hormone α3 (TRα3), and thyroid hormone receptor β1 (TRβ1) and suppressing estrogen receptor α36 (ER-α36)–Src could benefit the anti-proliferation of chelerythrine. However, it was disappointing that regulation of estrogen receptor α66 (ER-α66), estrogen receptor β (ER-β), and oxytocin receptor (OTR) contributed inversely negative effects on anti-gastric cancer cells. At present, the integrative study not only revealed chelerythrine as the most potent dual COX-2/5-LOX inhibitor from QAs but also generally highlighted that comprehensive regulation of the estrogen, thyroid, and oxytocin pathway should be noted once gastric cancer cells were treated with inflammatory inhibitors.

环氧合酶2（Cyclooxygenase 2, COX-2）与5-脂氧合酶（5-lipoxygenase, 5-LOX）在胃癌细胞中呈过表达状态，其双重抑制剂具备抗转移、抗侵袭的活性潜力，且副作用更低。为筛选靶向COX-2与5-LOX的抑制剂，本研究借助液相色谱（LC）与液相色谱-四极杆飞行时间质谱（LC-Q-TOF）技术，对野花椒（Zanthoxylum simulans）提取物中的季胺类生物碱（quaternary alkaloids, QAs）组分开展超滤富集与评分计算，以筛选候选化合物。针对信号强度较高的色谱峰，峰19（小檗碱，berberine）与峰21（白屈菜红碱，chelerythrine）被鉴定为活性最强的双重靶点候选化合物，且相较于COX-2，二者对5-LOX展现出更优的选择性结合亲和力。峰19对COX-2的富集倍数为4.36，对5-LOX的富集倍数为22.81；峰21对COX-2的富集倍数为7.81，对5-LOX的富集倍数为24.49。分子对接结果显示，白屈菜红碱是更为优异的双重靶点抑制剂，其对AGS胃癌细胞的抗增殖活性呈时间依赖性与剂量依赖性特征。生物信息学（bio-informatics）分析策略涵盖基因表达综合数据库（Gene Expression Omnibus, GEO）、基因本体（Gene Ontology, GO）及京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG），分析结果提示胃癌细胞内的激素信号通路可能受白屈菜红碱调控。后续的文献梳理与汇总分析，明确了COX-2/5-LOX抑制剂通过雌激素、甲状腺激素及催产素信号通路诱导胃癌细胞凋亡的潜在机制。本研究进一步将白屈菜红碱作用于胃癌细胞，以验证上述三条信号通路的调控作用。实验结果表明，促甲状腺激素受体（thyroid-stimulating hormone receptor, TSHR）、甲状腺激素α3（thyroid hormone α3, TRα3）与甲状腺激素受体β1（thyroid hormone receptor β1, TRβ1）的显著恢复表达，以及雌激素受体α36（estrogen receptor α36, ER-α36）-Src信号轴的抑制，可增强白屈菜红碱的抗增殖效应。但令人遗憾的是，雌激素受体α66（ER-α66）、雌激素受体β（ER-β）与催产素受体（OTR）的调控却对胃癌细胞的抗增殖治疗产生了反向不利影响。综上，本整合研究不仅从季胺类生物碱组分中筛选得到活性最强的COX-2/5-LOX双重抑制剂白屈菜红碱，同时也普遍提示：在采用炎症抑制剂治疗胃癌细胞时，需关注雌激素、甲状腺激素及催产素通路的综合调控效应。