Kaempferol (KO) and kaempferol 7-O-rhamnoside (KR) are natural products from various oriental herbs such as Geranii Herba. Previous studies have reported some biological activities of KO and KR; however, their effects on PD-1/PD-L1 interaction have not been reported yet. To elucidate their inhibitory activities on PD-1/PD-L1 protein-protein interaction (PPI), biochemical assays including competitive ELISA and biolayer interferometry (BLI) systems were performed. Cellular PD-1/PD-L1 blocking activity was measured in a co-culture system with PD-1 Jurkat and PD-L1/aAPC CHO-K1 cells by T-cell receptor (TCR) activation-induced nuclear factor of activated T cells (NFAT)-luciferase reporter assay. The detailed binding mode of action was simulated by an in silico docking study and pharmacophore analysis. Competitive ELISA revealed that KO and its glycoside KR significantly inhibited PD-1/PD-L1 interaction. Cellular PD-1/PD-L1 blocking activity was monitored by KO and KR at non-cytotoxic concentration. Surface plasmon resonance (SPR) and biolayer interferometry (BLI) analysis suggested the binding affinity and direct inhibition of KR against PD-1/PD-L1. An in silico docking simulation determined the detailed mode of binding of KR to PD-1/PD-L1. Collectively, these results suggest that KR could be developed as a potent small molecule inhibitor for PD-1/PD-L1 blockade.

中文翻译：

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山emp酚及其糖苷，山emp酚7-O-鼠李糖苷在体外抑制PD-1 / PD-L1相互作用。

山emp酚（KO）和山emp酚7-O-鼠李糖苷（KR）是来自各种东方草药（如Geranii Herba）的天然产物。先前的研究已经报道了KO和KR的某些生物活性。然而，它们对PD-1 / PD-L1相互作用的影响尚未见报道。为了阐明其对PD-1 / PD-L1蛋白-蛋白相互作用（PPI）的抑制活性，进行了包括竞争性ELISA和生物层干涉术（BLI）系统在内的生化测定。在PD-1 Jurkat和PD-L1 / aAPC CHO-K1细胞共培养系统中，通过T细胞受体（TCR）激活诱导的激活T细胞核因子测量了细胞PD-1 / PD-L1阻断活性（NFAT）荧光素酶报告基因检测。通过计算机对接研究和药效团分析来模拟详细的结合作用模式。竞争性ELISA显示，KO及其糖苷KR显着抑制了PD-1 / PD-L1的相互作用。通过非细胞毒性浓度的KO和KR监测细胞对PD-1 / PD-L1的阻断活性。表面等离振子共振（SPR）和生物层干涉法（BLI）分析表明KR对PD-1 / PD-L1具有结合亲和力和直接抑制作用。计算机对接模拟确定了KR与PD-1 / PD-L1结合的详细模式。总的来说，这些结果表明KR可以作为PD-1 / PD-L1阻断的有效小分子抑制剂而开发。表面等离振子共振（SPR）和生物层干涉法（BLI）分析表明KR对PD-1 / PD-L1具有结合亲和力和直接抑制作用。计算机对接模拟确定了KR与PD-1 / PD-L1结合的详细模式。总的来说，这些结果表明KR可以作为PD-1 / PD-L1阻断的有效小分子抑制剂而开发。表面等离振子共振（SPR）和生物层干涉法（BLI）分析表明KR对PD-1 / PD-L1具有结合亲和力和直接抑制作用。计算机对接模拟确定了KR与PD-1 / PD-L1结合的详细模式。总的来说，这些结果表明KR可以作为PD-1 / PD-L1阻断的有效小分子抑制剂而开发。