Elevated expression of WSB2 degrades p53 and activates the IGFBP3-AKT-mTOR-dependent pathway to drive hepatocellular carcinoma.

Dysregulation of wild-type p53 turnover is a key cause of hepatocellular carcinoma (HCC), yet its mechanism remains poorly understood. Here, we report that WD repeat and SOCS box containing protein 2 (WSB2), an E3 ubiquitin ligase, is an independent adverse prognostic factor in HCC patients. WSB2 drives HCC tumorigenesis and lung metastasis in vitro and in vivo. Mechanistically, WSB2 is a new p53 destabilizer that promotes K48-linked p53 polyubiquitination at the Lys291 and Lys292 sites in HCC cells, leading to p53 proteasomal degradation. Degradation of p53 causes IGFBP3-dependent AKT/mTOR signaling activation. Furthermore, WSB2 was found to bind to the p53 tetramerization domain via its SOCS box domain. Targeting mTOR with everolimus, an oral drug, significantly blocked WSB2-triggered HCC tumorigenesis and metastasis in vivo. In clinical samples, high expression of WSB2 was associated with low wild-type p53 expression and high p-mTOR expression. These findings demonstrate that WSB2 is overexpressed and degrades wild-type p53 and then activates the IGFBP3-AKT/mTOR axis, leading to HCC tumorigenesis and lung metastasis, which indicates that targeting mTOR could be a new therapeutic strategy for HCC patients with high WSB2 expression and wild-type p53.

Elevated FBXL6 activates both wild-type KRAS and mutant KRASG12D and drives HCC tumorigenesis via the ERK/mTOR/PRELID2/ROS axis in mice.

BACKGROUND: Kirsten rat sarcoma (KRAS) and mutant KRASG12D have been implicated in human cancers, but it remains unclear whether their activation requires ubiquitination. This study aimed to investigate whether and how F-box and leucine-rich repeat 6 (FBXL6) regulates KRAS and KRASG12D activity in hepatocellular carcinoma (HCC). METHODS: We constructed transgenic mouse strains LC (LSL-Fbxl6KI/+;Alb-Cre, n = 13), KC (LSL-KrasG12D/+;Alb-Cre, n = 10) and KLC (LSL-KrasG12D/+;LSL-Fbxl6KI/+;Alb-Cre, n = 12) mice, and then monitored HCC for 320 d. Multiomics approaches and pharmacological inhibitors were used to determine oncogenic signaling in the context of elevated FBXL6 and KRAS activation. Co­immunoprecipitation (Co-IP), Western blotting, ubiquitination assay and RAS activity detection assay were employed to investigate the underlying molecular mechanism by which FBXL6 activates KRAS. The pathological relevance of the FBXL6/KRAS/extracellular signal-regulated kinase (ERK)/mammalian target of rapamycin (mTOR)/proteins of relevant evolutionary and lymphoid interest domain 2 (PRELID2) axis was evaluated in 129 paired samples from HCC patients. RESULTS: FBXL6 is highly expressed in HCC as well as other human cancers (P < 0.001). Interestingly, FBXL6 drives HCC in transgenic mice. Mechanistically, elevated FBXL6 promotes the polyubiquitination of both wild-type KRAS and KRASG12D at lysine 128, leading to the activation of both KRAS and KRASG12D and promoting their binding to the serine/threonine-protein kinase RAF, which is followed by the activation of mitogen-activated protein kinase kinase (MEK)/ERK/mTOR signaling. The oncogenic activity of the MEK/ERK/mTOR axis relies on PRELID2, which induces reactive oxygen species (ROS) generation. Furthermore, hepatic FBXL6 upregulation facilitates KRASG12D to induce more severe hepatocarcinogenesis and lung metastasis via the MEK/ERK/mTOR/PRELID2/ROS axis. Dual inhibition of MEK and mTOR effectively suppresses tumor growth and metastasis in this subtype of cancer in vivo. In clinical samples, FBXL6 expression positively correlates with p-ERK (χ2 = 85.067, P < 0.001), p-mTOR (χ2 = 66.919, P < 0.001) and PRELID2 (χ2 = 20.891, P < 0.001). The Kaplan-Meier survival analyses suggested that HCC patients with high FBXL6/p-ERK levels predicted worse overall survival (log­rank P < 0.001). CONCLUSIONS: FBXL6 activates KRAS or KRASG12D via ubiquitination at the site K128, leading to activation of the ERK/mTOR/PRELID2/ROS axis and tumorigenesis. Dual inhibition of MEK and mTOR effectively protects against FBXL6- and KRASG12D-induced tumorigenesis, providing a potential therapeutic strategy to treat this aggressive subtype of liver cancer.

Elevated FBXL6 expression in hepatocytes activates VRK2-transketolase-ROS-mTOR-mediated immune evasion and liver cancer metastasis in mice.

Metastatic hepatocellular carcinoma (HCC) is the most lethal malignancy and lacks effective treatment. FBXL6 is overexpressed in human hepatocellular carcinoma (HCC), but whether this change drives liver tumorigenesis and lung metastasis in vivo remains unknown. In this study, we aimed to identify FBXL6 (F-Box and Leucine Rich Repeat Protein 6) as a key driver of HCC metastasis and to provide a new paradigm for HCC therapy. We found that elevated FBXL6 expression in hepatocytes drove HCC lung metastasis and was a much stronger driver than Kras mutation (KrasG12D/+;Alb-Cre), p53 haploinsufficiency (p53+/-) or Tsc1 loss (Tsc1fl/fl;Alb-Cre). Mechanistically, VRK2 promoted Thr287 phosphorylation of TKT and then recruited FBXL6 to promote TKT ubiquitination and activation. Activated TKT further increased PD-L1 and VRK2 expression via the ROS-mTOR axis, leading to immune evasion and HCC metastasis. Targeting or knockdown of TKT significantly blocked FBXL6-driven immune evasion and HCC metastasis in vitro and in vivo. Notably, the level of active TKT (p-Thr287 TKT) was increased and was positively correlated with the FBXL6 and VRK2 expression levels in HCC patients. Our work provides novel mechanistic insights into FBXL6-driven HCC metastasis and suggests that targeting the TKT-ROS-mTOR-PD-L1/VRK2 axis is a new paradigm for treating patients with metastatic HCC with high FBXL6 expression.

Elevated FBXW10 drives hepatocellular carcinoma tumorigenesis via AR-VRK2 phosphorylation-dependent GAPDH ubiquitination in male transgenic mice.

Hepatocellular carcinoma (HCC), the most common liver cancer, occurs mainly in men, but the underlying mechanism remains to be further explored. Here, we report that ubiquitinated glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is responsible for HCC tumorigenesis in males. Mechanistically, FBXW10 promotes GAPDH polyubiquitination and activation; VRK2-dependent phosphorylation of GAPDH Ser151 residue is critical for GAPDH ubiquitination and activation. Activated GAPDH interacts with TRAF2, leading to upregulation of the canonical and noncanonical NF-κB pathways, and increases PD-L1 and AR-VRK2 expression, followed by induction of immune evasion, HCC tumorigenesis, and metastasis. Notably, the GAPDH inhibitor koningic acid (KA) activates immune response and protects against FBXW10-driven HCC in vivo. In HCC clinical samples, the expression of active GAPDH is positively correlated with that of FBXW10 and VRK2. We propose that the FBXW10/AR/VRK2/GAPDH/NF-κB axis is critical for HCC tumorigenesis in males. Targeting this axis with KA is a potential therapeutic strategy for male HCC patients.

Elevated FBXL18 promotes RPS15A ubiquitination and SMAD3 activation to drive HCC.

BACKGROUND: F-box and leucine-rich repeat protein 18 (FBXL18) is an E3 ubiquitin ligase that is reported to be involved in the tumorigenesis of various types of cancer. However, it remains unknown whether FBXL18 is correlated with hepatocarcinogenesis. METHODS AND RESULTS: In the current study, we found that FBXL18 was highly expressed in HCC tissues and positively associated with poor overall survival of HCC patients. FBXL18 was an independent risk factor for HCC patients. We observed that FBXL18 drove HCC in FBXL18 transgenic mice. Mechanistically, FBXL18 promoted the K63-linked ubiquitination of small-subunit ribosomal protein S15A (RPS15A) and enhanced its stability, increasing SMAD family member 3 (SMAD3) levels and translocation to the nucleus and promoting HCC cell proliferation. Moreover, the knockdown of RPS15A or SMAD3 significantly suppressed FBXL18-mediated HCC proliferation. In clinical samples, elevated FBXL18 expression was positively associated with RPS15A expression. CONCLUSION: FBXL18 promotes RPS15A ubiquitination and upregulates SMAD3 expression, leading to hepatocellular carcinogenesis, and this study provides a novel therapeutic strategy for HCC treatment by targeting the FBXL18/RPS15A/SMAD3 pathway.

FBXW10-S6K1 promotes ANXA2 polyubiquitination and KRAS activation to drive hepatocellular carcinoma development in males.

The incidence rate of human hepatocellular carcinoma (HCC) is approximately three times higher in males than in females. A better understanding of the mechanisms underlying HCC development in males could lead to more effective therapies for HCC. Our previous study found that FBXW10 played a critical role in promoting HCC development in male mice and patients, but the mechanism remains unknown. Here, we found that FBXW10 promoted K63-linked ANXA2 polyubiquitination and activation in HCC tissues from males, and this process was required for S6K1-mediated phosphorylation. Activated ANXA2 further translocated from the cytoplasm to the cell membrane to bind KRAS and then activated the MEK/ERK pathway, leading to HCC proliferation and lung metastasis. Interfering with ANXA2 significantly blocked FBXW10-driven HCC growth and lung metastasis in vitro and in vivo. Notably, membrane ANXA2 was upregulated and positively correlated with FBXW10 expression in male HCC patients. These findings offer new insights into the regulation and function of FBXW10 signaling in HCC tumorigenesis and metastasis and suggest that the FBXW10-S6K1-ANXA2-KRAS-ERK axis may serve as a potential biomarker and therapeutic target in male HCC patients with high FBXW10 expression.

Silencing of FAM111B inhibited proliferation, migration and invasion of hepatoma cells through activating p53 pathway.

BACKGROUND: The function of Family with sequence similarity 111 member B (FAM111B) has been reported in multiple malignancies, but its involvement in occurrence and development of hepatocellular carcinoma (HCC) is still unclear. PURPOSE: To investigate the role of FAM111B in HCC and explore the potential molecular mechanism. METHODS: We examined the mRNA level of FAM111B via qPCR and protein level via immunohistochemistry in human HCC tissues. siRNA was used to construct a FAM111B-knockdown model in HCC cell lines. CCK-8, colony formation, transwell, and wound healing assays were performed to investigate the effect of FAM111B on proliferation, migration and invasion of HCC cell. Gene Set Enrichment Analysis, western blotting, and flow cytometry were carried out to find the related molecular mechanism. RESULTS: Human HCC tumor tissues exhibited higher expression of FAM111B, and high FAM111B expression was associated with poor prognosis. Vitro assays demonstrated that knockdown of FAM111B greatly repressed proliferation, migration and invasion of HCC cells. Furthermore, silencing of FAM111B significantly resulted in cell cycle arrest at G0/G1 and downregulation of epithelial-mesenchymal transition (EMT)-related proteins MMP7 and MMP9 via activation of p53 pathway. CONCLUSION: FAM111B played an essential role in promoting HCC development by regulation of p53 pathway.

FBXO43 increases CCND1 stability to promote hepatocellular carcinoma cell proliferation and migration.

Background and Aims: Abnormal expression of E3 ubiquitin ligase plays an important role in the development and progression of hepatocellular carcinoma (HCC), although the mechanism has remained elusive. This study aimed to investigate the biological function and potential mechanism of FBXO43 in HCC. Methods: FBXO43 expression in tissues and cells were detected by quantitative real-time PCR (qRT-PCR), Western blot, and immunohistochemistry (IHC). The Kaplan-Meier method and Cox regression analysis were used to explore the correlation between the expression level of FBXO43 and the clinical survival. MTT assay, EdU incorporation, colony formation, Transwell, and wound healing assays were performed to evaluate the function of FBXO43 in cell proliferation and migration in vitro. The interaction between FBXO43 and cyclin D1 (CCND1) was assessed by co-immunoprecipitation (Co-IP) assay and in vivo ubiquitination assay. Results: We found that FBXO43 was upregulated in HCC patient tissues and positively associated with poor clinicopathological features. Meanwhile, HCC patients with high expression of FBXO43 had shorter overall survival (OS) and disease-free survival (DFS). Furthermore, knockdown of FBXO43 inhibited HCC cell proliferation, migration and epithelial-mesenchymal transition (EMT) in HCC cells. Mechanistically, FBXO43 interacted with CCND1 and promoted its stability by polyubiquitination, leading to HCC cell proliferation, migration and EMT. Functional rescue experiments demonstrated that knockdown of CCND1 blocks FBXO43-mediated cell proliferation and metastasis. Conclusions: FBXO43, as an independent prognostic biomarker, promotes HCC cell proliferation, metastasis and EMT by stability of CCND1, which provides a new potential strategy for HCC treatment by targeting FBXO43-CCND1 axis.

An optimized protocol to detect protein ubiquitination and activation by ubiquitination assay in vivo and CCK-8 assay.

E3 ubiquitin ligases play a role in protein degradation, cellular localization, and activation, and their dysregulation is associated with human diseases. Here, we present a protocol to detect IGF2BP1 ubiquitination and activation by an E3 ubiquitin ligase FBXO45. We describe steps for preparing cells and transfecting plasmids. We detail the use of western blot to detect IGF2BP1 ubiquitination and a Cell Counting Kit-8 (CCK-8) assay to detect IGF2BP1 activation. This protocol is applicable to other proteins of interest. For complete details on the use and execution of this protocol, please refer to Lin et al. (2021).1.

In vivo analysis of FBXO45-mediated fibrosis and liver tumorigenesis in a chemically induced mouse model of hepatocellular carcinoma.

FBXO45, an E3 ubiquitin ligase highly expressed in liver tumors, is positively correlated with poor survival of hepatocellular carcinogenesis (HCC) patients, but whether FBXO45 drives HCC tumorigenesis remains largely unclear. Here, we describe a protocol that shortens the observation period for HCC tumorigenesis to assess the effects of FBXO45 in a DEN/CCl4-induced HCC mouse model. We describe steps for chemical induction of HCC in FBXO45-overexpressing mice, followed by tissue collection and pathology assessment via quantitative real-time PCR, histology, and immunohistochemistry. For complete details on the use and execution of this protocol, please refer to Lin et al. (2021).1.

Degradation of helicase-like transcription factor (HLTF) by ß-TrCP promotes hepatocarcinogenesis via activation of the p62/mTOR axis.

Helicase-like transcription factor (HLTF) has been found to be involved in the maintenance of genome stability and tumour suppression, but whether its downregulation in cancers is associated with posttranslational regulation remains unclear. Here, we observed that HLTF was significantly downregulated in hepatocellular carcinoma (HCC) tissues and positively associated with the survival of HCC patients. Mechanistically, the decreased expression of HLTF in HCC was attributed to elevated ß-TrCP-mediated ubiquitination and degradation. Knockdown of HLTF enhanced p62 transcriptional activity and mammalian target of rapamycin (mTOR) activation, leading to HCC tumourigenesis. Inhibition of mTOR effectively blocked ß-TrCP overexpression- or HLTF knockdown-mediated HCC tumourigenesis and metastasis. Furthermore, in clinical tissues, decreased HLTF expression was positively correlated with elevated expression of ß-TrCP, p62, or p-mTOR in HCC patients. Overall, our data not only uncover new roles of HLTF in HCC cell proliferation and metastasis, but also reveal a novel posttranslational modification of HLTF by ß-TrCP, indicating that the ß-TrCP/HLTF/p62/mTOR axis may be a new oncogenic driver involved in HCC development. This finding provides a potential therapeutic strategy for HCC patients by targeting the ß-TrCP/HLTF/p62/mTOR axis.

Mutant Kras and mTOR crosstalk drives hepatocellular carcinoma development via PEG3/STAT3/BEX2 signaling.

Background & Aims: Abnormal activation of mTOR through loss of tuberous sclerosis complex (Tsc) frequently occurs in hepatocellular carcinoma (HCC). Mutant Kras could induce aggressive HCCs. Here, we aim to identify the predictive or prognostic biomarkers for HCC patients with Kras mutant and mTOR hyperactivation, and to provide potential therapeutic approaches for this subtype of HCCs. Methods: We generated transgenic mice in which hepatocytic mTOR was hyperactivated through Tsc1 insufficiency with or without oncogenic KrasG12D. Bioinformatics and gain- or loss-of-function studies were used to illustrate the mechanisms underlying oncogenic pathway alterations. Transcriptional profiling was used to identify biomarker for the subtype of HCC. The therapeutic efficacy of targeting mTOR was tested in a liver orthotropic homogeneous murine model. Results: Oncogenic KrasG12D facilitated mTOR activation via the Mek/Erk/ROS axis, leading to HCC tumorigenesis and metastasis. Inhibition of Mek/Erk enhanced the anticancer effect of mTOR inhibitor via reduction of mTOR activity. Paternally expressed 3 (PEG3) was responsible for Kras/Erk- and mTOR-driven HCC. Elevated PEG3 protein interacted with STAT3 and promoted its transcriptional activity, resulting in the upregulation of proliferation- and metastasis-related proteins. Targeting mTOR significantly inhibited these actions in vitro and in vivo. Moreover, in clinical samples, PEG3 was identified as a new poor prognostic marker for HCC patients with Kras/Erk and mTOR hyperactivation. Conclusion: These findings reveal the underlying mechanism of hepatocytic Kras/Erk-driven mTOR activation and its downstream targets (PEG3 and STAT3) in HCC, identify PEG3 as a new prognostic biomarker for HCC with Kras/Erk and mTOR hyperactivation, and provide a potential therapeutic strategy for this subset of HCC patients.

Elevated Sodium Pump α3 Subunit Expression Promotes Colorectal Liver Metastasis via the p53-PTEN/IGFBP3-AKT-mTOR Axis.

The sodium pump α3 subunit is associated with colorectal liver metastasis. However, the underlying mechanism involved in this effect is not yet known. In this study, we found that the expression levels of the sodium pump α3 subunit were positively associated with metastasis in colorectal cancer (CRC). Knockdown of the α3 subunit or inhibition of the sodium pump could significantly inhibit the migration of colorectal cancer cells, whereas overexpression of the α3 subunit promoted colorectal cancer cell migration. Mechanistically, the α3 subunit decreased p53 expression, which subsequently downregulated PTEN/IGFBP3 and activated mTOR, leading to the promotion of colorectal cancer cell metastasis. Reciprocally, knockdown of the α3 subunit or inhibition of the sodium pump dramatically blocked this effect in vitro and in vivo via the downregulation of mTOR activity. Furthermore, a positive correlation between α3 subunit expression and mTOR activity was observed in an aggressive CRC subtype. Conclusions: Elevated expression of the sodium pump α3 subunit promotes CRC liver metastasis via the PTEN/IGFBP3-mediated mTOR pathway, suggesting that sodium pump α3 could represent a critical prognostic marker and/or therapeutic target for this disease.

Elevated FBXO45 promotes liver tumorigenesis through enhancing IGF2BP1 ubiquitination and subsequent PLK1 upregulation.

Dysregulation of tumor-relevant proteins may contribute to human hepatocellular carcinoma (HCC) tumorigenesis. FBXO45 is an E3 ubiquitin ligase that is frequently elevated expression in human HCC. However, it remains unknown whether FBXO45 is associated with hepatocarcinogenesis and how to treat HCC patients with high FBXO45 expression. Here, IHC and qPCR analysis revealed that FBXO45 protein and mRNA were highly expressed in 54.3% (57 of 105) and 52.2% (132 of 253) of the HCC tissue samples, respectively. Highly expressed FBXO45 promoted liver tumorigenesis in transgenic mice. Mechanistically, FBXO45 promoted IGF2BP1 ubiquitination at the Lys190 and Lys450 sites and subsequent activation, leading to the upregulation of PLK1 expression and the induction of cell proliferation and liver tumorigenesis in vitro and in vivo. PLK1 inhibition or IGF2BP1 knockdown significantly blocked FBXO45-driven liver tumorigenesis in FBXO45 transgenic mice, primary cells, and HCCs. Furthermore, IHC analysis on HCC tissue samples revealed a positive association between the hyperexpression of FBXO45 and PLK1/IGF2BP1, and both had positive relationship with poor survival in HCC patients. Thus, FBXO45 plays an important role in promoting liver tumorigenesis through IGF2BP1 ubiquitination and activation, and subsequent PLK1 upregulation, suggesting a new strategy for treating HCC by targeting FBXO45/IGF2BP1/PLK1 axis.

p53 haploinsufficiency and increased mTOR signalling define a subset of aggressive hepatocellular carcinoma.

BACKGROUND & AIMS: p53 mutations occur frequently in human HCC. Activation of the mammalian target of rapamycin (mTOR) pathway is also associated with HCC. However, it is still unknown whether these changes together initiate HCC and can be targeted as a potential therapeutic strategy. METHODS: We generated mouse models in which mTOR was hyperactivated by loss of tuberous sclerosis complex 1 (Tsc1) with or without p53 haplodeficiency. Primary cells were isolated from mouse livers. Oncogenic signalling was assessed in vitro and in vivo, with or without targeted inhibition of a single molecule or multiple molecules. Transcriptional profiling was used to identify biomarkers predictive of HCC. Human HCC materials were used to corroborate the findings from mouse models. RESULTS: p53 haploinsufficiency facilitates mTOR signalling via the PTEN/PI3K/Akt axis, promoting HCC tumorigenesis and lung metastasis. Inhibition of PI3K/Akt reduced mTOR activity, which effectively enhanced the anticancer effort of an mTOR inhibitor. ATP-binding cassette subfamily C member 4 (Abcc4) was found to be responsible for p53 haploinsufficiency- and Tsc1 loss-driven HCC tumorigenesis. Moreover, in clinical HCC samples, Abcc4 was specifically identified an aggressive subtype. The mTOR inhibitor rapamycin significantly reduced hepatocarcinogenesis triggered by Tsc1 loss and p53 haploinsufficiency in vivo, as well as the biomarker Abcc4. CONCLUSIONS: Our data advance the current understanding of the activation of the PTEN/PI3K/Akt/mTOR axis and its downstream target Abcc4 in hepatocarcinogenesis driven by p53 reduction and Tsc1 loss. Targeting mTOR, an unexpected vulnerability in p53 (haplo)deficiency HCC, can be exploited therapeutically to treat Abcc4-positive patients with HCC. LAY SUMMARY: Tsc1 loss facilitates the p53 (haplo)insufficiency-mediated activation of the PTEN/Akt/mTOR axis, leading to the elevated expression of Abcc4 to drive HCC tumorigenesis and metastasis in mice. Inhibition of mTOR protects against p53 haploinsufficiency and Tsc1 loss-triggered tumour-promoting activity, providing a new approach for treating an aggressive subtype of HCC exhibiting high Abcc4 expression.

Reduced IκBα promotes hepatocellular carcinoma cell proliferation and migration via regulation of NF-κB/Erbin axis.

Aberrantly low expression of NF-κB inhibitor α (IκBα) is observed in hepatocellular carcinoma (HCC), yet the underlying mechanism via which IκBα regulates HCC remains largely unknown. Therefore, to determine the potential function of IκBα in hepatocarcinogenesis, the present study used immunohistochemistry (IHC) staining to analyze the associations between IκBα protein expression and clinicopathologic characteristics of 107 patients with HCC. It was found that expression of IκBα was significantly associated with tumor recurrence. Moreover, IκBα protein expression was decreased in 107 HCC tissue samples and was positively associated with overall survival. Mechanistically, it was demonstrated that silencing of IκBα activated NF-κB in both Huh7 and HCCLM3 cells, followed by upregulation of Erbb2 interacting protein (Erbin) at both the mRNA and protein levels, confirmed by reverse transcription-quantitative PCR and western blotting, to promote cell proliferation and migration. Furthermore, knockdown of Erbin significantly attenuated NF-κB-mediated cell proliferation and migration. It was also identified that overexpression of Erbin in HCC tissues promoted both cell proliferation and migration, and was negatively associated with IκBα expression in 107 HCC tissue samples. Thus, these results indicated that downregulation of IκBα promoted HCC tumorigenesis via upregulation of NF-κB-mediated Erbin expression.

Prognostic and predicted significance of Ubqln2 in patients with hepatocellular carcinoma.

PURPOSE: Hepatocellular carcinoma (HCC) is a common malignant cancer and the third leading cause of death worldwide. The molecular mechanism of HCC remains unclear. Recent studies have demonstrated that the ubiquitin-proteasome system (UPS) is associated with HCC. Ubqln2, a member of the UPS, is abnormally expressed in HCC. However, whether Ubqln2 is associated with HCC prognosis remains unknown. PATIENTS AND METHODS: We analyzed the associations between overall survival and various risk factors in 355 HCC tissue samples obtained from the Cancer Genomic Atlas (TCGA) database at the mRNA level and in 166 HCC tissue samples from Southwest Hospital at the protein level. qRCR was used to determinate Ubqln2 expression in cancer and noncancerous tissues. The association between Ubqln2 and Ki-67 was analyzed by immunohistochemistry. The association between Ubqln2 expression and survival was analyzed using Kaplan-Meier curve and Cox proportional hazards models. A nomogram was used to predict the impact of Ubqln2 on prognosis. Mutated genes were analyzed to determine the potential mechanism. RESULTS: Ubqln2 highly expressed in HCC tissues. The Ubqln2 mRNA level had significant relations with UICC tumor stage (P = .022), UICC stage (P = .034) and resection potential (P = .017). Concordantly, the Ubqln2 protein was closely associated with tumor size (P = .005), UICC stage (P = .012), and recurrence (P = .009). Ubqln2 was highly expressed in HCC and positively associated with poor survival. The nomogram precisely predicted the prognosis of HCC patients with high or low Ubqln2 expression. A genomic waterfall plot suggested that Ubqln2 expression was closely associated with mutated CTNNB1. CONCLUSION: Our findings reveal that Ubqln2, an independent risk factor for HCC, is a potential prognostic marker in HCC patients. Ubqln2 expression is positively associated with mutated CTNNB1.

FBXW10 promotes hepatocarcinogenesis in male patients and mice.

Hepatocellular carcinoma (HCC) is reported to associate with abnormal expression of SCF E3 ubiquitin ligases. FBXW10, an F-box protein of the E3 ubiquitin ligases, was abnormally regulated in HCC patients. However, whether FBXW10 is associated with HCC has not yet been evaluated. Here, we analyzed the associations between overall survival and various risk factors in 191 HCC tissues. Univariate and multivariate analyses demonstrated that FBXW10 was an independent risk factor related to HCC prognosis. The results showed that FBXW10, gender and tumor state were strongly associated with overall survival in HCC patients. Furthermore, high expression of FBXW10 was associated with poor survival among male HCC patients but not female HCC patients. FBXW10 was more highly expressed in male HCC tissues and more strongly related to vascular invasion in male HCC patients. Consistent with these findings, the male FBXW10-Tg(+) mice were more susceptible to tumorigenesis, changes in regenerative capacity, and liver injury and inflammation but not changes in liver function than FBXW10-Tg(-) mice. FBXW10 promoted cell proliferation and migration in HCC cell lines. Our findings reveal that FBXW10, an independent risk factor for HCC, promotes hepatocarcinogenesis in male patients, and is also a potential prognostic marker in male patients with HCC.

The FBXW2-MSX2-SOX2 axis regulates stem cell property and drug resistance of cancer cells.

SOX2 is a key transcription factor that plays critical roles in maintaining stem cell property and conferring drug resistance. However, the underlying mechanisms by which SOX2 level is precisely regulated remain elusive. Here we report that MLN4924, also known as pevonedistat, a small-molecule inhibitor of neddylation currently in phase II clinical trials, down-regulates SOX2 expression via causing accumulation of MSX2, a known transcription repressor of SOX2 expression. Mechanistic characterization revealed that MSX2 is a substrate of FBXW2 E3 ligase. FBXW2 binds to MSX2 and promotes MSX2 ubiquitylation and degradation. Likewise, FBXW2 overexpression shortens the protein half-life of MSX2, whereas FBXW2 knockdown extends it. We further identified hypoxia as a stress condition that induces VRK2 kinase to facilitate MSX2-FBXW2 binding and FBXW2-mediated MSX2 ubiquitylation and degradation, leading to SOX2 induction via derepression. Biologically, expression of FBXW2 or SOX2 promotes tumor sphere formation, which is blocked by MSX2 expression. By down-regulating SOX2 through inactivation of FBXW2 E3 ligase, MLN4924 sensitizes breast cancer cells to tamoxifen in both in vitro and in vivo cancer cell models. Thus, a negative cascade of the FBXW2-MSX2-SOX2 axis was established, which regulates stem cell property and drug resistance. Finally, an inverse correlation of expression was found between FBXW2 and MSX2 in lung and breast cancer tissues. Collectively, our study revealed an anticancer mechanism of MLN4924. By inactivating FBXW2, MLN4924 caused MSX2 accumulation to repress SOX2 expression, leading to suppression of stem cell property and sensitization of breast cancer cells to tamoxifen.