Sorafenib induces ferroptosis by promoting TRIM54-mediated FSP1 ubiquitination and degradation in hepatocellular carcinoma.

BACKGROUND: Ferroptosis is a unique form of regulated cell death that provided a new opportunity for cancer therapy. Ferroptosis suppressor protein 1 (FSP1) is a key regulator in the NAD(P)H/FSP1/CoQ10 antioxidant system, which sever as an oxide redox enzyme to scavenge harmful lipid hydroperoxides and escape from ferroptosis in cells. This study aimed to investigate the role of FSP1 on sorafenib-induced ferroptosis and disclosed the underlying mechanisms. METHODS: Cell viability, malondialdehyde (MDA), glutathione (GSH), and lipid reactive oxygen species levels were assessed using indicated assay kits. The levels of FSP1 and glutathione peroxidase 4 (GPX4) in the patients with HCC were analyzed based on the database. Western blot and quantitative real-time PCR were performed to detect the protein and mRNA expression. Co-immunoprecipitation was applied to detect the interaction between proteins. Tumor xenograft experiments were used to evaluate whether overexpression of FSP1-inhibited sorafenib-induced ferroptosis in vivo. RESULTS: We verified that sorafenib-induced ferroptosis in HCC. Furthermore, we found that sorafenib decreased the protein level of FSP1, and knockdown FSP1 rendered HCC cells susceptible to sorafenib-induced ferroptosis. Co-immunoprecipitation and ubiquitination assays showed that sorafenib accelerated the TRIM54-mediated FSP1 ubiquitination and degradation. Sorafenib-induced ferroptosis was abrogated by TRIM54 suppression. Mechanically, sorafenib-promoted TRIM54 ubiquitinated and degraded FSP1 by means of the ERK pathway. Moreover, FSP1 enhanced tumor development and decreased HCC cellular susceptibility to sorafenib in vivo. CONCLUSIONS: Sorafenib facilitated the TRIM54-mediated FSP1 ubiquitination through the ERK pathway, thereby inducing ferroptosis in HCC cells.

HPV E7-drived ALKBH5 promotes cervical cancer progression by modulating m6A modification of PAK5.

Human papillomavirus (HPV) infection is a causative agent of cervical cancer (CC). N6-methyladenosine (m6A) modification is implicated in carcinogenesis and tumor progression. However, the involvement of m6A modification in HPV-involved CC remains unclear. Here we showed that HPV E6/7 oncoproteins affected the global m6A modification and E7 specifically promoted the expression of ALKBH5. We found that ALKBH5 was significantly upregulated in CC and might serve as a valuable prognostic marker. Forced expression of ALKBH5 enhanced the malignant phenotypes of CC cells. Mechanistically, we discovered that E7 increased ALKBH5 expression through E2F1-mediated activation of the H3K27Ac and H3K4Me3 histone modifications, as well as post-translational modification mediated by DDX3. ALKBH5-mediated m6A demethylation enhanced the expression of PAK5. The m6A reader YTHDF2 bound to PAK5 mRNA and regulated its stability in an m6A-dependent manner. Moreover, ALKBH5 promoted tumorigenesis and metastasis of CC by regulating PAK5. Overall, our findings herein demonstrate a significant role of ALKBH5 in CC progression in HPV-positive cells. Thus, we propose that ALKBH5 may serve as a prognostic biomarker and therapeutic target for CC patients.

PAK5 potentiates slug transactivation of N-cadherin to facilitate metastasis of renal cell carcinoma.

Renal cell carcinoma (RCC) is an aggravating cancer with a poor prognosis and a high rate of metastasis. PAK5, a p21-activated kinases, has shown to be overexpressed in a variety of cancers, including RCC. In previous studies, we discovered that PAK5 regulates cell migration and invasion in RCC cell lines. However, the underlying mechanisms remain obscure. In this study, we consolidated that PAK5 confers a pro-metastatic phenotype RCC cells in vitro and exacerbates metastasis in vivo. High PAK5 expression was associated with an advanced TNM stage and a lower overall survival. Furthermore, PAK5 increases the expression level of N-cadherin. In terms of mechanism, PAK5 bound to Slug and phosphorylated it at serine 87. As a result, phosphorylated Slug transactivated N-cadherin, accelerating the epithelial-mesenchymal transition. Collectively, Slug is a novel PAK5 substrate, and PAK5-mediated phosphorylation of Slug-S87 increases N-cadherin and the pro-metastatic phenotype of RCC, implying that phosphorylated Slug-S87 could be a therapeutic target in progressive RCC.

COPS6 promotes tumor progression and reduces CD8+ T cell infiltration by repressing IL-6 production to facilitate tumor immune evasion in breast cancer.

Due to poor T cell infiltration, tumors evade immune surveillance. Increased CD8+ T cell infiltration in breast cancer suggests a satisfactory response to immunotherapy. COPS6 has been identified as an oncogene, but its role in regulating antitumor immune responses has not been defined. In this study, we investigated the impact of COPS6 on tumor immune evasion in vivo. Tumor transplantation models were established in C57BL/6 J mice and BALB/c nude mice. Flow cytometry was conducted to identify the role of COPS6 on tumor-infiltrating CD8+ T cells. By analyzing the TCGA and GTEx cohort, we found that COPS6 expression was significantly up-regulated in a variety of cancers. In human osteosarcoma cell line U2OS and non-small cell lung cancer cell line H1299, we showed that p53 negatively regulated COPS6 promoter activity. In human breast cancer MCF-7 cells, COPS6 overexpression stimulated p-AKT expression as well as the proliferation and malignant transformation of tumor cells, whereas knockdown of COPS6 caused opposite effects. Knockdown of COPS6 also significantly suppressed the growth of mouse mammary cancer EMT6 xenografts in BALB/c nude mice. Bioinformatics analysis suggested that COPS6 was a mediator of IL-6 production in the tumor microenvironment and a negative regulator of CD8+ T cell tumor infiltration in breast cancer. In C57BL6 mice bearing EMT6 xenografts, COPS6 knockdown in the EMT6 cells increased the number of tumor-infiltrating CD8+ T cells, while knockdown of IL-6 in COPS6KD EMT6 cells diminished tumor infiltrating CD8+ T cells. We conclude that COPS6 promotes breast cancer progression by reducing CD8+ T cell infiltration and function via the regulation of IL-6 secretion. This study clarifies the role of p53/COPS6/IL-6/CD8+ tumor infiltrating lymphocytes signaling in breast cancer progression and immune evasion, opening a new path for development of COPS6-targeting therapies to enhance tumor immunogenicity and treat immunologically "cold" breast cancer.

Alternative splicing of HSPA12A pre-RNA by SRSF11 contributes to metastasis potential of colorectal cancer.

BACKGROUND: Dysregulation of alternative splicing (AS) induced by serine/arginine-rich proteins has recently been linked to cancer metastasis. Nonetheless, as a member of the serine/arginine-rich protein family, the involvement of SRSF11 in colorectal cancer (CRC) is unknown. METHODS: The TCGA dataset and clinical samples were used to assess SRSF11 expression levels in CRC. For SRSF11, functional experiments were conducted both in vitro and in vivo. RNA-seq technology was used to analyze and screen SRSF11-triggered AS events, which were then confirmed by in vivo UV crosslinking and immunoprecipitation (CLIP) and mini-gene reporter assays. Jalview software was used to determine the preferential binding motif with relation to exon skipping (ES) events. Furthermore, coimmunoprecipitation (Co-IP) and Phospho-tag SDS-PAGE experiments were used to investigate PAK5-mediated phosphorylation regulation on SRSF11, and in vitro kinase experiments validated the interaction. RESULTS: In CRC, SRSF11 was discovered to be overexpressed and associated with a poor prognosis. And SRSF11 played a pro-metastatic role in vitro and in vivo. By screening SRSF11-regulated AS events, we identified the binding motif of SRSF11-triggered splicing-switching of HSPA12A AS, which specifically regulated HSPA12A AS by directly binding to a motif in exon 2. Mechanistically, the HSPA12A transcript with exon 2 retention increased N-cadherin expression by promoting RNA stability. Furthermore, the oncogenic kinase PAK5 phosphorylated SRSF11 at serine 287, protecting it from ubiquitination degradation. CONCLUSIONS: SRSF11 exerts pro-metastatic effects in CRC by inhibiting the AS of HSPA12A pre-RNA. Our findings point to SRSF11-regulated HSPA12A splicing as a novel relationship between SRSF11-regulated splicing and CRC metastasis and suggest a PAK5/SRSF11/HSPA12A axis as a potential therapeutic target and prognostic biomarker in CRC.

Correlation between crowdedness in emergency departments and anxiety in Chinese patients.

BACKGROUND: Emergency department (ED) overcrowding is a severe health care concern, while anxiety and depression rates among ED patients have been reported to be substantially higher compared to the general population. We hypothesized that anxiety due to over crowdedness may lead to adverse events in EDs. AIM: To investigate correlations between crowdedness in EDs and anxiety of patients and nurses, and to identify factors affecting their anxiety. METHODS: In this prospective observational study, a total 43 nurses and 389 emergency patients from two tier III hospitals located in Beijing were included from January 2016 to August 2017. Patients were grouped into inpatients when they were hospitalized after diagnoses, or into outpatients when they were discharged after treatments. The State Trait Anxiety Inventory (STAI Form Y) questionnaire was used to investigate patient and nurse anxieties, while crowdedness of EDs was evaluated with the National Emergency Department Over Crowding Score. RESULTS: The present results revealed that state anxiety scores (49.50 ± 6.00 vs 50.80 ± 2.80, P = 0.005) and trait anxiety scores (45.40 ± 5.70 vs 46.80 ± 2.70, P = 0.002) between inpatients (n = 173) and outpatients (n = 216) were significantly different, while the state anxiety of nurses (44.70 ± 5.80) was different from those of both patient groups. Generalized linear regression analysis demonstrated that multiple factors, including crowdedness in the ED, were associated with state and trait anxieties for both inpatients and outpatients. In addition, there was an interaction between state anxiety and trait anxieties. However, multivariable regression analysis showed that while overcrowding in the ED did not directly correlate with patients' and nurses' anxiety levels, the factors that did correlate with state and trait anxieties of inpatients were related to crowdedness. These factors included waiting time in the ED, the number of patients treated, and the number of nurses in the ED, whereas for nurses, only state and trait anxieties correlated significantly with each other. CONCLUSION: Waiting time, the number of patients treated, and the number of nurses present in the ED correlate with patient anxiety in EDs, but crowdedness has no effect on nurse or patient anxiety.