GP73 Is Indispensable For T Cell Anti-tumor Activity

Background and objectiveThe homeostasis of T cells maintains the balance of the body’s internal environment,and metabolism plays an important role in it.Especially in the tumor microenvironment,the metabolism of T cells is converted from oxidative phosphorylation to glycolysis,and the energy to maintain the activity of T cells decreases.In addition,the inhibitory receptors on the surface of T cells are also activated by signals from tumor cells and regulatory immune cells,which inhibit the secretion of cytokines by T cells,resulting in immune escape of tumor cells.Therefore,tumor immunotherapy with the purpose of reactivating T cells has been fruitful in clinical tumor treatment,bringing hope to patients.However,tumor immunotherapy is only effective for some patients,and the treatment effect for most patients is poor,and it is often accompanied by side effects.With the increasing incidence of tumors,safer and more efficient treatment methods are urgently needed.GP73 is a transmembrane glycoprotein located on the Golgi apparatus and is involved in the body’s metabolic pathways.Studies have found that in tumor tissues such as hepatocellular carcinoma,colon cancer,and prostate cancer,the expression level of GP73 tends to increase exponentially,and GP73 also continuously activates the EGFR/RTK signaling pathway and the downstream AKT/S6K signaling pathway,eventually leading to tumor growth and metastasis.T cells play an important role in anti-tumor immunity,and whether GP73 is involved in affecting the function of T cells remains unclear.This study focused on the function of GP73 in T cells,aiming to provide data support for clinical tumor immunotherapy.MethodThe expression levels of GP73 before and after activation of human and mouse T cells by CD3/CD28 antibody were detected by flow cytometry.The Cre-Loxp system was used to construct T cell GP73-specific knockout mice,and through subcutaneous tumor implantation and T cell killing experiments in vitro,it was investigated whether the function of T cells would be affected by the knockout of GP73.Single-cell sequencing analysis of CD45⁺cells isolated from tumors was performed to study the differences in KEGG pathway and gene expression in T cells of knockout mice and control mice.The correlation of GP73 with TOX and HIF-1αin various tumor data was analyzed by using TCGA database.The protein expression levels of TOX and HIF-1αwere detected by Jurket T GP73^kocell line.Co-immunoprecipitation was used to study whether GP73interacts with TOX and HIF-1α.ResultThe expression level of GP73 is higher in human and mouse T cells,and after activation by CD3/CD28 antibody,the expression level of GP73 is further increased.In the tumor-bearing mouse model,the tumor of the knockout mice showed a faster growth ability.Correspondingly,in the in vitro killing experiments of mouse T cells,the knockout mouse T cells also showed relatively weak tumor-killing ability.Single-cell sequencing analysis showed that the expression of exhausted immune checkpoints TOX and CD244in the T cell genome of knockout mice increased,and the levels of glycolysis-related genes HIF-1αwere significantly decreased.TCGA database analysis found that the expression level of GP73 was positively correlated with HIF-1αin hepatocellular carcinoma,colon cancer and melanoma.Consistent with the single-cell sequencing results,the expression level of HIF-1αwas decreased and TOX was increased in Jurket T GP73^kocell line.ConclusionKnockdown of GP73 in T cells attenuates the killing function of T cells against tumor cells,and this effect seems to be achieved by accelerating T cell exhaustion and reducing the metabolic capacity of T cells,and the specific effect mechanism requires further investigation.