Shikonin-mediated PD-L1 degradation suppresses immune evasion in pancreatic cancer by inhibiting NF-κB/STAT3 and NF-κB/CSN5 signaling pathways.

Pancreatic cancer (PC) is a highly fatal malignancy with few effective therapies currently available. Recent studies have shown that PD-L1 inhibitors could be potential therapeutic targets for the treatment of PC. The present study aims to investigate the effect of Shikonin on immune evasion in PC with the involvement of the PD-L1 degradation. Initially, the expression patterns of PD-L1 and NF-κB in PC were predicted in-silico using the GEPIA database, and were subsequently validated using PC tissues. Thereafter, the correlation of NF-κB with STAT3, CSN5 and PD-L1 was examined. PC cells were treated with Shikonin, NF-κB inhibitor, STAT3 activator, and CSN5 overexpression plasmid to investigate effects on PD-L1 glycosylation and immune evasion in PC. Finally, in vivo tumor formation was induced in C57BL/6J mice, in order to verify the in vitro results. PD-L1, NF-κB, NF-κB p65, STAT3, and CSN5 were highly expressed in PC samples, and NF-κB was positively correlated with STAT3/CSN5/PD-L1. Inhibition of NF-κB decreased PD-L1 glycosylation and increased PD-L1 degradation, whereas activated STAT3 and overexpressed CSN5 reversed these trends. Shikonin blocked immune evasion in PC, and lowered the expression of PD-L1, NF-κB, NF-κB p65, STAT3 and CSN5 in vivo and in vitro. The findings indicated Shikonin inhibited immune evasion in PC by inhibiting PD-L1 glycosylation and activating the NF-κB/STAT3 and NF-κB/CSN5 signaling pathways. These effects of Shikonin on PC cells may bear important potential therapeutic implications for the treatment of PC.

Ruan Z ，Liang M ，Shang L ，Lai M ，Deng X ，Su X ... -  《-》

Overexpression of DAPK1-mediated inhibition of IKKβ/CSN5/PD-L1 axis enhances natural killer cell killing ability and inhibits tumor immune evasion in gastric cancer.

Gastric cancer (GC) originates from the stomach and is a prevalent human malignancy. Dysfunction of death associated protein kinase 1 (DAPK1) has been identified as a major regulator involved in the development and progression of GC. However, there's limited data regarding the regulatory mechanism of GC. Herein, we investigated role of DAPK1 in natural killer (NK) cell killing ability and immune evasion of GC cells and mediated pathway. Samples from GC-related gene expression profile and clinical samples from 67 patients with GC were collected to determine the expression of DAPK1, IκB kinase β (IKKβ), programmed death receptor-ligand 1 (PD-L1), and photomorphogenesis 9 (COP9) signalosome 5 (CSN5). The binding affinity among DAPK1, IKKβ, CSN5, and PD-L1 was characterized to verify the underlying mechanism. GC lines were transfected with overexpressed plasmid or siRNA to determine the effect of DAPK1/IKKβ/CSN5/PD-L1 axis on NK cell killing ability and immune evasion of GC cells. GC cells and tissues presented low expression of DAPK1 and high expression of IKKβ, CSN5 and PD-L1. IKKβ, negatively regulated by DAPK1, was capable of activating CSN5 and upregulating PD-L1 expression. Overexpression of DAPK1 promoted NK cell killing ability and reduced immune evasion, coupled with reduction of NK cell apoptosis and increases in levels of TNF-α, IFN-γ, CD107a, and Granzyme B cytokines. The tumor-suppressing properties of DAPK1 through downregulation of IKKβ/CSN5/PD-L1 axis in GC were further confirmed in vivo. In summary, overexpression of DAPK1 promoted the NK cell killing ability and restrained immune evasion of GC cells, providing a potential therapeutic strategy for GC treatment by modulating immune evasion.

Guo Z ，Zhou C ，Zhou L ，Wang Z ，Zhu X ，Mu X ... -  《-》

Deubiquitination and Stabilization of PD-L1 by CSN5.

Pro-inflammatory cytokines produced in the tumor microenvironment lead to eradication of anti-tumor immunity and enhanced tumor cell survival. In the current study, we identified tumor necrosis factor alpha (TNF-α) as a major factor triggering cancer cell immunosuppression against T cell surveillance via stabilization of programmed cell death-ligand 1 (PD-L1). We demonstrated that COP9 signalosome 5 (CSN5), induced by NF-κB p65, is required for TNF-α-mediated PD-L1 stabilization in cancer cells. CSN5 inhibits the ubiquitination and degradation of PD-L1. Inhibition of CSN5 by curcumin diminished cancer cell PD-L1 expression and sensitized cancer cells to anti-CTLA4 therapy.

Lim SO ，Li CW ，Xia W ，Cha JH ，Chan LC ，Wu Y ，Chang SS ，Lin WC ，Hsu JM ，Hsu YH ，Kim T ，Chang WC ，Hsu JL ，Yamaguchi H ，Ding Q ，Wang Y ，Yang Y ，Chen CH ，Sahin AA ，Yu D ，Hortobagyi GN ，Hung MC ... -  《-》

PDIA6 promotes pancreatic cancer progression and immune escape through CSN5-mediated deubiquitination of β-catenin and PD-L1.

Protein Disulfide Isomerase Family A Member 6 (PDIA6) is an endoplasmic reticulum protein that is capable of catalyzing protein folding and disulfide bond formation. Abnormally elevated expression of PDIA6 has been reported to predict poor outcomes in various cancers. Herein, gain-of- and loss-of-function experiments were performed to investigate how PDIA6 participated in the carcinogenesis of pancreatic cancer (PC). By analyzing the protein expression of PDIA6 in 28 paired PC and para carcinoma specimens, we first found that PDIA6 expression was higher in PC samples. Both the overall survival and disease-free survival rates of PC patients with higher PDIA6 expression were poorer than those with lower PDIA6 (n = 178). Furthermore, knockdown of PDIA6 impaired the malignancies of PC cells - suppressed cell proliferation, invasion, migration, cisplatin resistance, and xenografted tumor growth. PDIA6-silenced PC cells were more sensitive to cytotoxic natural killer (NK) cells. Overexpression of PDIA6 had opposite effects on PC cells. Interestingly, COP9 signalosome subunit 5 (CSN5), a regulator of E3 ubiquitin ligases known to promote deubiquitination of its downstream targets, was demonstrated to interact with PDIA6, and its expression was increased in PC cells overexpressing PDIA6. Additionally, PDIA6 overexpression promoted deubiquitination of β-catenin and PD-L1 and subsequently upregulated their expression in PC cells. These alterations were partly reversed by CSN5 shRNA. Collectively, the above results demonstrate that PDIA6 contributes to PC progression, which may be associated with CSN5-regulated deubiquitination of β-catenin and PD-L1. Our findings suggest PDIA6 as a potential target for the treatment of PC.

Ma Y ，Xia P ，Wang Z ，Xu J ，Zhang L ，Jiang Y ... -  《NEOPLASIA》

The deubiquitinase USP22 regulates PD-L1 degradation in human cancer cells.

Wang Y ，Sun Q ，Mu N ，Sun X ，Wang Y ，Fan S ，Su L ，Liu X ... -  《Cell Communication and Signaling》