Tong Ling, Shan Minjie, Zou Wen, Liu XianLing, Felsher Dean W, Wang Jingjing
Division of Oncology, Departments of Medicine and Pathology, Stanford University School of Medicine, Stanford, CA, United States.
Department of Oncology, The Second Xiangya Hospital of Central South University, Changsha, China.
Front Oncol. 2022 Aug 1;12:904969. doi: 10.3389/fonc.2022.904969. eCollection 2022.
The cyclic adenosine monophosphate/phosphodiesterase 4 cAMP/PDE4) pathway is involved in inflammation and immune regulation; however, the effect of cAMP/PDE4 on immune infiltration and immune evasion in lung adenocarcinoma (LUAD) remains unclear.
CBioPortal, which is the The Cancer Genome Atlas (TCGA) online database, and the Kaplan Meier plotter were used to analyze the association between genes and the prognosis of TCGA-LUAD. Tumor Immune Estimation Resource (TIMER) was used to analyze the association between gene expression and immune infiltration. The Genecards database was used to identify the transcription factors of related genes. The lung adenocarcinoma cell line H1299 and A549 were treated with cAMP pathway drugs. Flow cytometry and qRT-PCR were used to detect the PD-L1 protein and gene expression, respectively. A one-way analysis of variance with Tukey's test or a Student's t-test were used.
It was found that PDE4B and CREB1, which are downstream genes of the cAMP/PDE4 axis, were differentially expressed in LUAD and adjacent tissues and are correlated with the prognosis and immune infiltration of LUAD. In the CBioPortal database, cAMP pathway genes are closely related to programmed cell death-ligand 1 (PD-L1) expression in TCGA-LUAD. The protein-protein interaction revealed that there was a direct interaction between CREB1/CREBBP, which are the downstream molecules of the cAMP/PDE4 axis, and MYC; additionally, MYC was predicted to bind to the PD-L1 transcription site and regulate PD-L1 expression. CREB1 was also predicted to transcriptionally bind to both MYC and PD-L1. These results predicted the interaction network of cAMP/PDE4/CREB1/CREBP/MYC/PD-L1, and the core factor may be related to MYC. In the cell experiment, forskolin (an adenylate cyclase activator) and zardaverine (a PDE4 inhibitor) enhance the cAMP pathway and decrease PD-L1 expression, while SQ2253 (an adenylate cyclase inhibitor) inhibits the cAMP pathway and increases PD-L1 expression of the LUAD cell lines H1299 and A549, and MYC regulation by these drugs was positively correlated with PD-L1 regulation, which verified the regulation of the cAMP/PDE4 pathway on MYC and PD-L1.
This study showed that the cAMP/PDE4 pathway may play an important role in PD-L1 regulation and immune infiltration in LUAD.
环磷酸腺苷/磷酸二酯酶4（cAMP/PDE4）信号通路参与炎症和免疫调节；然而，cAMP/PDE4对肺腺癌（LUAD）免疫浸润和免疫逃逸的影响仍不清楚。
利用癌症基因组图谱（TCGA）在线数据库CBioPortal和Kaplan Meier绘图工具分析基因与TCGA-LUAD预后的相关性。使用肿瘤免疫估计资源（TIMER）分析基因表达与免疫浸润的相关性。利用Genecards数据库鉴定相关基因的转录因子。用cAMP信号通路药物处理肺腺癌细胞系H1299和A549。分别采用流式细胞术和qRT-PCR检测PD-L1蛋白和基因表达。采用单因素方差分析及Tukey检验或Student t检验。
发现cAMP/PDE4轴的下游基因PDE4B和CREB1在LUAD组织和癌旁组织中存在差异表达，且与LUAD的预后和免疫浸润相关。在CBioPortal数据库中，cAMP信号通路基因与TCGA-LUAD中程序性细胞死亡配体1（PD-L1）的表达密切相关。蛋白质-蛋白质相互作用分析显示，cAMP/PDE4轴的下游分子CREB1/CREBBP与MYC之间存在直接相互作用；此外，预测MYC可与PD-L1转录位点结合并调节PD-L1表达。还预测CREB1可与MYC和PD-L1发生转录结合。这些结果预测了cAMP/PDE4/CREB1/CREBP/MYC/PD-L1的相互作用网络，核心因子可能与MYC有关。在细胞实验中，福斯可林（一种腺苷酸环化酶激活剂）和扎达维林（一种PDE4抑制剂）可增强cAMP信号通路并降低PD-L1表达，而SQ2253（一种腺苷酸环化酶抑制剂）则抑制cAMP信号通路并增加LUAD细胞系H1299和A549的PD-L1表达，且这些药物对MYC的调节与对PD-L1的调节呈正相关，这证实了cAMP/PDE4信号通路对MYC和PD-L1的调节作用。
本研究表明，cAMP/PDE4信号通路可能在LUAD的PD-L1调节和免疫浸润中发挥重要作用。
