Possible Action of Olaparib for Preventing Invasion of Oral Squamous Cell Carcinoma In Vitro and In Vivo.

Despite recent advances in treatment, the prognosis of oral cancer remains poor, and prevention of recurrence and metastasis is critical. Olaparib is a PARP1 inhibitor that blocks polyADP-ribosylation, which is involved in the epithelial-mesenchymal transition (EMT) characteristic of tumor recurrence. We explored the potential of olaparib in inhibiting cancer invasion in oral carcinoma using three oral cancer cell lines, HSC-2, Ca9-22, and SAS. Olaparib treatment markedly reduced their proliferation, migration, invasion, and adhesion. Furthermore, qRT-PCR revealed that olaparib inhibited the mRNA expression of markers associated with tumorigenesis and EMT, notably Ki67, Vimentin, ß-catenin, MMP2, MMP9, p53, and integrin α2 and ß1, while E-Cadherin was upregulated. In vivo analysis of tumor xenografts generated by injection of HSC-2 cells into the masseter muscles of mice demonstrated significant inhibition of tumorigenesis and bone invasion by olaparib compared with the control. This was associated with reduced expression of proteins involved in osteoclastogenesis, RANK and RANKL. Moreover, SNAIL and PARP1 were downregulated, while E-cadherin was increased, indicating the effect of olaparib on proteins associated with EMT in this model. Taken together, these findings confirm the effects of olaparib on EMT and bone invasion in oral carcinoma and suggest a new therapeutic strategy for this disease.

NFE2L1 and NFE2L3 Complementarily Maintain Basal Proteasome Activity in Cancer Cells through CPEB3-Mediated Translational Repression.

Proteasomes are protease complexes essential for cellular homeostasis, and their activity is crucial for cancer cell growth. However, the mechanism of how proteasome activity is maintained in cancer cells has remained unclear. The CNC family transcription factor NFE2L1 induces the expression of almost all proteasome-related genes under proteasome inhibition. Both NFE2L1 and its phylogenetically closest homolog, NFE2L3, are highly expressed in several types of cancer, such as colorectal cancer. Here, we demonstrate that NFE2L1 and NFE2L3 complementarily maintain basal proteasome activity in cancer cells. Double knockdown of NFE2L1 and NFE2L3 impaired basal proteasome activity in cancer cells and cancer cell resistance to a proteasome inhibitor anticancer drug, bortezomib, by significantly reducing the basal expression of seven proteasome-related genes: PSMB3, PSMB7, PSMC2, PSMD3, PSMG2, PSMG3, and POMP Interestingly, the molecular basis behind these cellular consequences was that NFE2L3 repressed NFE2L1 translation by the induction of the gene encoding the translational regulator CPEB3, which binds to the NFE2L1 3' untranslated region and decreases polysome formation on NFE2L1 mRNA. Consistent results were obtained from clinical analysis, wherein patients with cancer having tumors expressing higher levels of CPEB3/NFE2L3 exhibit poor prognosis. These results provide the novel regulatory mechanism of basal proteasome activity in cancer cells through an NFE2L3-CPEB3-NFE2L1 translational repression axis.

NRF3-POMP-20S Proteasome Assembly Axis Promotes Cancer Development via Ubiquitin-Independent Proteolysis of p53 and Retinoblastoma Protein.

Proteasomes are essential protease complexes that maintain cellular homeostasis, and aberrant proteasomal activity supports cancer development. The regulatory mechanisms and biological function of the ubiquitin-26S proteasome have been studied extensively, while those of the ubiquitin-independent 20S proteasome system remain obscure. Here, we show that the cap 'n' collar (CNC) family transcription factor NRF3 specifically enhances 20S proteasome assembly in cancer cells and that 20S proteasomes contribute to colorectal cancer development through ubiquitin-independent proteolysis of the tumor suppressor p53 and retinoblastoma (Rb) proteins. The NRF3 gene is highly expressed in many cancer tissues and cell lines and is important for cancer cell growth. In cancer cells, NRF3 upregulates the assembly of the 20S proteasome by directly inducing the gene expression of the 20S proteasome maturation protein POMP. Interestingly, NRF3 knockdown not only increases p53 and Rb protein levels but also increases p53 activities for tumor suppression, including cell cycle arrest and induction of apoptosis. Furthermore, protein stability and cell viability assays using two distinct proteasome inhibitor anticancer drugs, the 20S proteasome inhibitor bortezomib and the ubiquitin-activating enzyme E1 inhibitor TAK-243, show that the upregulation of the NRF3-POMP axis leads to ubiquitin-independent proteolysis of p53 and Rb and to impaired sensitivity to bortezomib but not TAK-243. More importantly, the NRF3-POMP axis supports tumorigenesis and metastasis, with higher NRF3/POMP expression levels correlating with poor prognoses in patients with colorectal or rectal adenocarcinoma. These results suggest that the NRF3-POMP-20S proteasome assembly axis is significant for cancer development via ubiquitin-independent proteolysis of tumor suppressor proteins.

Multiple regulatory mechanisms of the biological function of NRF3 (NFE2L3) control cancer cell proliferation.

Accumulated evidence suggests a physiological relationship between the transcription factor NRF3 (NFE2L3) and cancers. Under physiological conditions, NRF3 is repressed by its endoplasmic reticulum (ER) sequestration. In response to unidentified signals, NRF3 enters the nucleus and modulates gene expression. However, molecular mechanisms underlying the nuclear translocation of NRF3 and its target gene in cancer cells remain poorly understood. We herein report that multiple regulation of NRF3 activities controls cell proliferation. Our analyses reveal that under physiological conditions, NRF3 is rapidly degraded by the ER-associated degradation (ERAD) ubiquitin ligase HRD1 and valosin-containing protein (VCP) in the cytoplasm. Furthermore, NRF3 is also degraded by ß-TRCP, an adaptor for the Skp1-Cul1-F-box protein (SCF) ubiquitin ligase in the nucleus. The nuclear translocation of NRF3 from the ER requires the aspartic protease DNA-damage inducible 1 homolog 2 (DDI2) but does not require inhibition of its HRD1-VCP-mediated degradation. Finally, NRF3 mediates gene expression of the cell cycle regulator U2AF homology motif kinase 1 (UHMK1) for cell proliferation. Collectively, our study provides us many insights into the molecular regulation and biological function of NRF3 in cancer cells.