In recent years, immunotherapy, especially programmed cell death protein 1 (PD-1) / programmed cell death ligand 1 (PD-L1) axis targeted therapy becomes the first line strategy in several types of tumors. However, its low immune response remains to be solved. Tumor microenvironment (TME) regulation is potential to improve immune response of anti-PD-1/PD-L1 therapy. Here, several screening platforms based on important markers, including adenosine triphosphate (ATP), calreticulin (CRT), major histocompatibility complex class II molecules (MHC II), cluster of differentiation 80 (CD80), cluster of differentiation 86 (CD86), and C-X-C motif chemokine 10 (CXCL10), were established, which were aimed at discovering TCM derived products in regulating TME. ATP and CRT take part in dendritic cells (DC) recruitment and phagocytosis, respectively. MHC II, CD80, and CD86 are markers on mature DC'S, and CXCL10 recruits T cells. Fangchinoline, alisol B 23-acetate, platycodin D, 18 B-glycyrrhetintic acid, and wogonoside increased the concentration of released ATP from tumor cells and were screened out via ATP secretion based screening platform. Celastrol and psoralidin were screened out as potential products to induce CRT relocation via CRT-RFP U2OS cell-based screening platforms. The maturation model of DC was developed by detecting the expression of MHC II, CD80, and CD86 on the DC's membrane. Moreover, cycleanine, psoralidin, tetrandrine, baicalein, cepharanthine and icaritin were found to up regulating the mRNA level of CXCL10. Psoralidin isolated from the seed of Psoraled corylifolia L. enhanced the secretion of CXCL10 in MC38 cells and the recruitment of T cells in co-culture model. Meanwhile, it down regulated the secretion of CCL2 in MC38 cells and the recruitment of myeloid-derived suppressor cells (MDSCs). Additionally, it reduced the mRNA level of arginase 1 in bone marrow derived MDSCs. In all, screen systems based on several markers in TME were established and potential products were obtained. Moreover, psoralidin regulated recruitment of immune cells and modulated chemokines' secretion, which gave potential method to improve immune response of PD-1/PD-LI targeted therapy.

近年來，以細胞程式性死亡受體 1 (Programmed cell death protein 1, PD-1)抗體及細胞程式死亡-配體1(Programmedcelldeathligand1,PD-I1）抗體治療為代表的免疫治療成為多類癌種的一線治療策略，但其較低的免疫響應率仍有待解 決。腫瘤微環境（Tumor microenvironment, TME）中的免疫細胞浸潤和細胞因 子表達等條件在免疫治療中發揮著很大作用，因此調節腫瘤免疫微環境可作為提高 PD-L1 抗體響應率的一種潛在方法。我們基於腫瘤免疫中不同環節的幾個重要標誌物建立了篩選平臺，旨在發現調節腫瘤免疫微環境的中藥來源產物。 鈣網本白（Calreticulin, CRT）、三磷酸腺苷（Adenosine triphosphate, ATP)、第 二類主要組織相容性複合體(MaiorhistocompatibilitycomplexclassIImolecules,MHCI)、分化群 80 (Cluster of Differentiation 80, CD80）、分化群 86 (Cluster of Difterentiation 86, CD86） 和干擾素伽瑪誘導的10千道關頓蛩白（C-※-C motif chemokine 10, CXCL10） 被選為篩選標誌物。ATP 可招募樹突細胞 (Dendritic cell. Dc) : CRT 可以促進 DC 吞噬作用：MHIC 11, CD80和 CD86為 DC成熟指標；CXCL10 可招募 T細胞。 通過檢測腫瘤細胞分泌的 ATP的濃度，發現防己諾林城 (Fangchinolin)、23-乙 醯濹瀉醇B（Alisol B 23-acetate)、桔梗皂苷D(Platycodin D)、18 B-甘草次酸 (18 B-Glycyrrhetintic Acid)和漢黃芩苷（Wogonoside）可增加從腫瘤細胞中釋 放的 ATP。通過建立 CRT-RFP U2OS 細胞模型進行篩選，得到調控 CRT遷移的 化合物，例如雷公藤紅素 (Celastrol)和補骨（Psoralidin, PSo)。建立了以 MHCI、CD80及CD86為檢測標誌的DC成熟檢測模型。此外，通過基於 CXCL10轉錄水準測定的節選平臺，篩選發現輪環藤城（Cycleanine）、補骨脂 定(Psoralidin）、粉防己域（Tetrandrine）、黃芩素（Baicalein）、千金藤素 (Cepharanthine）和脫水淫羊藿素(Icaritin）可以在mRNA水準上調CXCI10。 PSO作為從 Psoralea corylifoliaL.種子中分離出的一種小分子，可上調 MC38細 胞中 CXCL10 蛋白的分泌並在共培養模型中增加工細胞的募集，同時下調 CCL2的分泌並在共培養模型中減少 MDSC的募集。此外，PSO 可降低 MDSC 中精胺酸晦1的mRNA水準。 綜上所述，我們建立了基於腫瘤微環境中的幾個標誌物的篩選平臺，並篩選獲得了多個中藥來源的潛在活性產物。PSO 參興調飾免疫細胞的募集並調籪趨化 因子的分泌，為改善 PD-1/PD-I1抗體治療的響應率提供了選擇。