ABSTRACT
The proteasomal 19S regulatory particle (RP) associated deubiquitinases (DUBs) have attracted much attention owing to their potential as a therapeutic target for cancer therapy. Identification of new entities against 19S RP associated DUBs and illustration of the underlying mechanisms is crucial for discovery of novel proteasome blockers. In this study, a series of 4-arylidene curcumin analogues were identified as potent proteasome inhibitor by preferentially blocking deubiquitinase function of proteasomal 19S RP with moderate 20S CP inhibition. The most active compound 33 exhibited a major inhibitory effect on 19S RP-associated ubiquitin-specific proteases 14, along with a minor effect on ubiquitin C-terminal hydrolase 5, which resulted in dysfunction of proteasome, and subsequently accumulated ubiquitinated proteins (such as IκB) in several cancer cells. Remarkably, though both 19S RP and 20S CP inhibition induced significantly endoplasmic reticulum stress and triggered caspase-12/9 pathway activation to promote cancer cell apoptosis, the 19S RP inhibition by 33 avoided slow onset time, Bcl-2 overexpression, and PERK-phosphorylation, which contribute to the deficiencies of clinical drug Bortezomib. These systematic studies provided insights in the development of novel proteasome inhibitors for cancer treatment.
Subject(s)
Antineoplastic Agents/pharmacology , Curcumin/analogs & derivatives , Curcumin/pharmacology , Deubiquitinating Enzymes/antagonists & inhibitors , Proteasome Endopeptidase Complex , Proteasome Inhibitors/pharmacology , A549 Cells , Animals , Antineoplastic Agents/chemistry , CHO Cells , Cell Proliferation/drug effects , Cell Proliferation/physiology , Cricetinae , Cricetulus , Deubiquitinating Enzymes/metabolism , Dose-Response Relationship, Drug , Drug Delivery Systems/methods , Humans , Proteasome Endopeptidase Complex/metabolism , Proteasome Inhibitors/chemistryABSTRACT
Natural killer (NK) cells play an important role in preventing cancer development. NK group 2 member D (NKG2D) is an activating receptor expressed in the membrane of NK cells. Tumour cells expressing NKG2DL become susceptible to an immune-dependent rejection mainly mediated by NK cells. The paradoxical roles of transforming growth factor beta (TGF-ß) in regulation of NKG2DL are presented in many studies, but the mechanism is unclear. In this study, we showed that TGF-ß up-regulated the expression of NKG2DLs in both PC3 and HepG2 cells. The up-regulation of NKG2DLs was characterized by increasing the expression of UL16-binding proteins (ULBPs) 1 and 2. TGF-ß treatment also increased the expression of transcription factor SP1. Knockdown of SP1 significantly attenuated TGF-ß-induced up-regulation of NKG2DLs in PC3 and HepG2 cells, suggesting that SP1 plays a key role in TGF-ß-induced up-regulation of NKG2DLs. TGF-ß treatment rapidly increased SP1 protein expression while not mRNA level. It might be due to that TGF-ß can elevate SP1 stability by activating PI3K/AKT signalling pathway, subsequently inhibiting GSK-3ß activity and decreasing the association between SP1 and GSK-3ß. Knockdown of GSK-3ß further verified our findings. Taken together, these results revealed that AKT/GSK-3ß-mediated stabilization of SP1 is required for TGF-ß induced up-regulation of NKG2DLs. Our study provided valuable evidence for exploring the tumour immune modulation function of TGF-ß.
Subject(s)
Glycogen Synthase Kinase 3 beta/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Receptors, NK Cell Lectin-Like/metabolism , Sp1 Transcription Factor/metabolism , Transforming Growth Factor beta/pharmacology , Up-Regulation/drug effects , Hep G2 Cells , Humans , Models, Biological , Phosphatidylinositol 3-Kinases/metabolism , Protein Stability/drug effects , Protein Transport/drug effects , Signal Transduction/drug effectsABSTRACT
Human leukocyte antigen class I antigens (HLA-I) is essential in immune response by presenting antigenic peptides to cytotoxic T lymphocytes. Downregulation of HLA-I is observed in primary and metastatic prostate cancers, which facilitates them escape from immune surveillance, thereby promotes prostate cancer progression. In addition, elevated level of growth factors like TGF-ß or EGF in microenvironment is related to the prostate cancer deterioration. Thus, we wondered whether TGF-ß or EGF was involved in the regulation of HLA-I during the development of prostate cancer cells. In this study, we demonstrated that TGF-ß and EGF both downregulated the expression of HLA-I, thereby attenuated the cytotoxic T cell mediated lysis of prostate cancer cells. Next, we revealed that TGF-ß and EGF induced downregulation of HLA-I is associated with classical epithelial-mesenchymal transition (EMT) morphological changes and expression profiles. We further illustrated that overexpression of Snail is crucial for HLA-I downregulation and its association with EMT. At last, we discussed that NF-κB/p65 is the plausible target for Snail to induce HLA-I downregulation. Taken together, this is the first evidence to reveal that both TGF-ß and EGF can induce HLA-I downregulation which is then proven to be associated with EMT in prostate cancer cells. These discoveries provide a deeper understanding of growth factors induced immune escape and introduce potential therapeutic targets for prostate cancers.
