Kinesin family member 18B activates mTORC1 signaling via actin gamma 1 to promote the recurrence of human hepatocellular carcinoma.

The mechanistic target of rapamycin complex 1 (mTORC1) signaling pathway is frequently reported to be hyperactivated in hepatocellular carcinoma (HCC) and contributes to HCC recurrence. However, the underlying regulatory mechanisms of mTORC1 signaling in HCC are not fully understood. In the present study, we found that the expression of kinesin family member 18B (KIF18B) was positively correlated with mTORC1 signaling in HCC, and the upregulation of KIF18B and p-mTOR was associated with a poor prognosis and HCC recurrence. Utilizing in vitro and in vivo assays, we showed that KIF18B promoted HCC cell proliferation and migration through activating mTORC1 signaling. Mechanistically, we identified Actin gamma 1 (γ-Actin) as a binding partner of KIF18B. KIF18B and γ-Actin synergistically modulated lysosome positioning, promoted mTORC1 translocation to lysosome membrane, and prohibited p70 S6K from entering lysosomes for degradation, which finally led to the enhancement of mTORC1 signaling transduction. Moreover, we found that KIF18B was a direct target of Forkhead box M1, which explains the potential mechanism of KIF18B overexpression in HCC. Our study highlights the potential of KIF18B as a therapeutic target for the treatment of HCC.

The aberrant upregulation of exon 10-inclusive SREK1 through SRSF10 acts as an oncogenic driver in human hepatocellular carcinoma.

Deregulation of alternative splicing is implicated as a relevant source of molecular heterogeneity in cancer. However, the targets and intrinsic mechanisms of splicing in hepatocarcinogenesis are largely unknown. Here, we report a functional impact of a Splicing Regulatory Glutamine/Lysine-Rich Protein 1 (SREK1) variant and its regulator, Serine/arginine-rich splicing factor 10 (SRSF10). HCC patients with poor prognosis express higher levels of exon 10-inclusive SREK1 (SREK1L). SREK1L can sustain BLOC1S5-TXNDC5 (B-T) expression, a targeted gene of nonsense-mediated mRNA decay through inhibiting exon-exon junction complex binding with B-T to exert its oncogenic role. B-T plays its competing endogenous RNA role by inhibiting miR-30c-5p and miR-30e-5p, and further promoting the expression of downstream oncogenic targets SRSF10 and TXNDC5. Interestingly, SRSF10 can act as a splicing regulator for SREK1L to promote hepatocarcinogenesis via the formation of a SRSF10-associated complex. In summary, we demonstrate a SRSF10/SREK1L/B-T signalling loop to accelerate the hepatocarcinogenesis.

USP5 facilitates non-small cell lung cancer progression through stabilization of PD-L1.

PD-L1(CD274) is a well-known immunosuppressive molecule, which confers immunoescape features to cancer cells and has become one of the major targets in cancer immunotherapies. Understanding the regulatory mechanisms that control PD-L1 protein expression is important for guiding immune checkpoint blockade therapy. Here, we showed that ubiquitin specific peptidase 5 (USP5) was a novel PD-L1 deubiquitinase in non-small cell lung cancer (NSCLC) cells. USP5 directly interacted with PD-L1 and deubiquitinated PD-L1, therefore enhances PD-L1 protein stability. Meanwhile, USP5 protein levels were highly elevated and positively correlated to PD-L1 levels in NSCLC tissues, and were closely correlated with poor prognosis of these patients. In addition, knockdown of USP5 retarded tumor growth in the Lewis lung carcinoma mouse model. Thus, we identified that USP5 was a new regulator of PD-L1 and targeting USP5 is a promising strategy for cancer therapy.

Salmonella flagella confer anti-tumor immunological effect via activating Flagellin/TLR5 signalling within tumor microenvironment.

mediated cancer therapy has achieved remarkable anti-tumor effects in experimental animal models, but the detailed mechanism remains unsolved. In this report, the active involvement of the host immune response in this process was confirmed by comparing the tumor-suppressive effects of Salmonella in immunocompetent and immunodeficient mice bearing melanoma allografts. Since flagella are key inducers of the host immune response during bacterial infection, flagella were genetically disrupted to analyse their involvement in Salmonella-mediated cancer therapy. The results showed that flagellum-deficient strains failed to induce significant anti-tumor effects, even when more bacteria were administered to offset the difference in invasion efficiency. Flagella mainly activate immune cells via Flagellin/Toll-like receptor 5 (TLR5) signalling pathway. Indeed, we showed that exogenous activation of TLR5 signalling by recombinant Flagellin and exogenous expression of TLR5 both enhanced the therapeutic efficacy of flagellum-deficient Salmonella against melanoma. Our study highlighted the therapeutic value of the interaction between Salmonella and the host immune response through Flagellin/TLR5 signalling pathway during Salmonella-mediated cancer therapy, thereby suggesting the potential application of TLR5 agonists in the cancer immune therapy.

Dysregulation of Wnt/ß-catenin signaling by protein kinases in hepatocellular carcinoma and its therapeutic application.

Wnt/ß-catenin signaling is indispensable for many biological processes, including embryonic development, cell cycle, inflammation, and carcinogenesis. Aberrant activation of the Wnt/ß-catenin signaling can promote tumorigenicity and enhance metastatic potential in hepatocellular carcinoma (HCC). Targeting this pathway is a new opportunity for precise medicine for HCC. However, inhibiting Wnt/ß-catenin signaling alone is unlikely to significantly improve HCC patient outcome due to the lack of specific inhibitors and the complexity of this pathway. Combination with other therapies will be an important next step in improving the efficacy of Wnt/ß-catenin signaling inhibitors. Protein kinases play a key and evolutionarily conserved role in the Wnt/ß-catenin signaling and have become one of the most important drug targets in cancer. Targeting Wnt/ß-catenin signaling and its regulatory kinase together will be a promising HCC management strategy. In this review, we summarize the kinases that modulate the Wnt/ß-catenin signaling in HCC and briefly discuss their molecular mechanisms. Furthermore, we list some small molecules that target the kinases and may inhibit Wnt/ß-catenin signaling, to offer new perspectives for preclinical and clinical HCC studies.

Erianin inhibits the oncogenic properties of hepatocellular carcinoma via inducing DNA damage and aberrant mitosis.

Natural compounds have been confirmed as one of the most feasible solutions for hard-to-treat cancers such as hepatocellular carcinoma (HCC). Erianin, a natural bibenzyl compound from Dendrobium chrysotoxum, has been recently discovered with anticancer property in cancer cells. However, the roles and the molecular mechanisms of erianin in HCC remain unknown. The present study evaluates the effect of erianin on human HCC cells by inhibiting cell proliferation, inducing apoptotic-related cell death and hampering tumorigenicity. Furthermore, it was found that erianin could cause irreparable DNA damage, induce G2/M arrest and deregulate mitotic regulators. It was also observed that many cells with damaged DNA induced by erianin could overcome G2/M arrest and enter mitosis, leading to abnormal mitosis, and subsequently mitotic catastrophe and apoptotic-related cell death. The present study confirmed that erianin could be a potential antitumor agent for HCC clinical treatment.

Honokiol inhibits bladder tumor growth by suppressing EZH2/miR-143 axis.

The oncoprotein EZH2, as a histone H3K27 methyltransferase, is frequently overexpressed in various cancer types. However, the mechanisms underlying its role in urinary bladder cancer (UBC) cells have not yet fully understood. Herein, we reported that honokiol, a biologically active biphenolic compound isolated from the Magnolia officinalis inhibited human UBC cell proliferation, survival, cancer stemness, migration, and invasion, through downregulation of EZH2 expression level, along with the reductions of MMP9, CD44, Sox2 and the induction of tumor suppressor miR-143. Either EZH2 overexpression or miR-143 inhibition could partially reverse honokiol-induced cell growth arrest and impaired clonogenicity. Importantly, it was first revealed that EZH2 could directly bind to the transcriptional regulatory region of miR-143 and repress its expression. Furthermore, honokiol treatment on T24 tumor xenografts confirmed its anticancer effects in vivo, including suppression tumor growth and tumor stemness, accompanied by the dysregulation of EZH2 and miR-143 expressions. Our data suggest a promising therapeutic option to develop drugs targeting EZH2/miR-143 axis, such as honokiol, for bladder cancer treatment.

GOLPH3 is a potential therapeutic target and a prognostic indicator of poor survival in bladder cancer treated by cystectomy.

Golgi phosphoprotein 3 (GOLPH3) has been reported to be involved in the development of several human cancers. However, its clinical significance and biological role in bladder cancer remains unclear. In this study, we sought to analyze the GOLPH3 expression in bladder cancer samples and cells, and explore its clinical significance and biological role. We found that GOLPH3 was significantly increased in bladder cancer tissues and cells. Overexpression of GOLPH3 had significant correlation with poorer survival for bladder cancer patients treated by cystectomy. Knockdown of GOLPH3 inhibited the proliferation, migration and invasion of cancer cells, and tumor growth in a xenograft mouse model. GOLPH3 silencing inhibited AKT/m-TOR signaling, increased the cyclin-dependent kinase (CDK) inhibitor p27 and decreased the CDK regulator cyclin D1 and matrix metallopeptidase 9 (MMP9). Thus, GOLPH3 is likely to play important roles in bladder cancer progression via modulating AKT/mTOR signaling, and it is a novel prognostic biomarker and promising therapeutic target for bladder cancer.

Truncated ERG Oncoproteins from TMPRSS2-ERG Fusions Are Resistant to SPOP-Mediated Proteasome Degradation.

SPOP mutations and TMPRSS2-ERG rearrangements occur collectively in up to 65% of human prostate cancers. Although the two events are mutually exclusive, it is unclear whether they are functionally interrelated. Here, we demonstrate that SPOP, functioning as an E3 ubiquitin ligase substrate-binding protein, promotes ubiquitination and proteasome degradation of wild-type ERG by recognizing a degron motif at the N terminus of ERG. Prostate cancer-associated SPOP mutations abrogate the SPOP-mediated degradation function on the ERG oncoprotein. Conversely, the majority of TMPRSS2-ERG fusions encode N-terminal-truncated ERG proteins that are resistant to the SPOP-mediated degradation because of degron impairment. Our findings reveal degradation resistance as a previously uncharacterized mechanism that contributes to elevation of truncated ERG proteins in prostate cancer. They also suggest that overcoming ERG resistance to SPOP-mediated degradation represents a viable strategy for treatment of prostate cancers expressing either mutated SPOP or truncated ERG.

Hypoxia-inducible miR-182 enhances HIF1α signaling via targeting PHD2 and FIH1 in prostate cancer.

Activation of hypoxia-inducible factor 1α (HIF1α) controls the transcription of genes governing angiogenesis under hypoxic condition during tumorigenesis. Here we show that hypoxia-responsive miR-182 is regulated by HIF1α at transcriptional level. Prolyl hydroxylase domain enzymes (PHD) and factor inhibiting HIF-1 (FIH1), negative regulators of HIF1 signaling, are direct targets of miR-182. Overexpression of miR-182 in prostate cancer cells led to a reduction of PHD2 and FIH1 expression and an increase in HIF1α level either under normoxic or hypoxic condition. Consistently, inhibition of miR-182 could increase PHD2 and FIH1 levels, thereby reducing the hypoxia-induced HIF1α expression. Matrigel plug assay showed that angiogenesis was increased by miR-182 overexpression, and vice versa. miR-182 overexpression in PC-3 prostate cancer xenografts decreased PHD2 and FIH1 expression, elevated HIF1α protein levels, and increased tumor size. Lastly, we revealed that the levels of both miR-182 and HIF1α were elevated, while the expression PHD2 and FIH1 was downregulated in a mouse model of prostate cancer. Together, our results suggest that the interplay between miR-182 and HIF1α could result in a sustained activation of HIF1α pathway, which might facilitate tumor cell adaption to hypoxic stress during prostate tumor progression.

δ-tocotrienol induces human bladder cancer cell growth arrest, apoptosis and chemosensitization through inhibition of STAT3 pathway.

Vitamin E intake has been implicated in reduction of bladder cancer risk. However, the mechanisms remain elusive. Here we reported that δ-tocotrienol (δ-T3), one of vitamin E isomers, possessed the most potent cytotoxic capacity against human bladder cancer cells, compared with other Vitamin E isomers. δ-T3 inhibited cancer cell proliferation and colonogenicity through induction of G1 phase arrest and apoptosis. Western blotting assay revealed that δ-T3 increased the expression levels of cell cycle inhibitors (p21, p27), pro-apoptotic protein (Bax) and suppressed expression levels of cell cycle protein (Cyclin D1), anti-apoptotic proteins (Bcl-2, Bcl-xL and Mcl-1), resulting in the Caspase-3 activation and cleavage of PARP. Moreover, the δ-T3 treatment inhibited ETK phosphorylation level and induced SHP-1 expression, which was correlated with downregulation of STAT3 activation. In line with this, δ-T3 reduced the STAT3 protein level in nuclear fraction, as well as its transcription activity. Knockdown of SHP-1 partially reversed δ-T3-induced cell growth arrest. Importantly, low dose of δ-T3 sensitized Gemcitabine-induced cytotoxic effects on human bladder cancer cells. Overall, our findings demonstrated, for the first time, the cytotoxic effects of δ-T3 on bladder cancer cells and suggest that δ-T3 might be a promising chemosensitization reagent for Gemcitabine in bladder cancer treatment.

Hepatic miR-378 targets p110α and controls glucose and lipid homeostasis by modulating hepatic insulin signalling.

Understanding the regulation of insulin signalling in tissues provides insights into carbohydrate and lipid metabolism in physiology and disease. Here we show that hepatic miR-378/378* expression changes in response to fasting and refeeding in mice. Mice overexpressing hepatic miR-378/378* exhibit pure hepatic insulin resistance. miR-378 inhibits hepatic insulin signalling through targeting p110α, a subunit of PI3K and hence a critical component of insulin signalling. Knockdown of hepatic p110α mimics the effect of miR-378, while restoration of p110α expression abolishes the action of miR-378 on insulin signalling as well as its systemic effects on glucose and lipid homeostasis. miR-378/378* knockout mice display hypoglycemia and increased hepatic triglyceride level with enhanced insulin sensitivity. Inhibition of hepatic p110α in miR-378/378* knockout mice corrects the abnormal glucose tolerance. Finally, we show that overexpression of hepatic miR-378/378* ameliorates hepatic steatosis in ob/ob mice without exacerbating hyperglycemia. Our findings establish fasting-responsive miR-378 as a critical regulator of hepatic insulin signalling.

Overexpression of HP1γ is associated with poor prognosis in non-small cell lung cancer cell through promoting cell survival.

Heterochromatin protein 1γ (HP1γ), which binds to di- or trimethylated lysine 9 on histone H3 (H3K9), plays an important role in chromatin packaging and gene transcriptional regulation. Recently, HP1γ has been implicated in cancer development. However, its clinical relevance and functional role in non-small cell lung cancer (NSCLC) remain elusive. In this study, we found that HP1γ expression was elevated in NSCLC samples at the messenger RNA (mRNA) level compared to adjacent normal lung tissues. In a cohort of 108 NSCLC patients, HP1γ overexpression is significantly associated with N stage (P = 0.003), pathological tumor-node-metastasis (TNM) stage (P = 0.013), smoking status (P = 0.009), and gender (P = 0.042). Patients with a high level of HP1γ expression showed a poorer overall survival rate than those with low HP1γ expression (P = 0.017). Multivariate analysis revealed that HP1γ expression is an independent prognostic marker. We also found knockdown of HP1γ in A549 and NCI-H1975 cells induced apoptosis accompanied with suppressed cell proliferation and colony formation. Consistently, pro-apoptotic proteins, Bax and GADD45α, were upregulated in response to HP1γ depletion. Altogether, our data suggested that HP1γ plays an important role in promoting NSCLC and may represent a novel prognostic biomarker and therapeutic target for the disease.

A long noncoding RNA Sox2ot regulates lung cancer cell proliferation and is a prognostic indicator of poor survival.

Sox2 overlapping transcript (Sox2ot) is a long noncoding RNA (lncRNA), localized on human chromosome 3q26.33, which is frequently amplified in lung squamous cell carcinomas (SCCs). However, its roles in lung cancer remain under investigation. In this study, we found that Sox2ot was up-regulated over two folds in 53.01% of human primary lung cancers (44/83). The expression level of Sox2ot is significantly higher in SCCs than that in adenocarcinomas (ADCs) of the lung. Further study found high Sox2ot expression predicted poor survival in lung cancer patients (P=0.0053), implying Sox2ot is a novel prognostic factor. In two human lung cancer cell lines, HCC827 and SK-MES-1, knocking down Sox2ot inhibited cell proliferation by inducing G2/M arrest, with a concomitant decrease of cells in S phase. Reduced protein levels of Cyclin B1 and Cdc2 were also observed. Importantly, knocking down Sox2ot decreased EZH2 expression and reintroduction of EZH2 allowed Sox2ot knockdown cells progressed through G2/M phase, which correlates with the restoration of Cyclin B1 and Cdc2 expressions. Altogether, our data suggested that Sox2ot plays an important role in regulating lung cancer cell proliferation, and may represent a novel prognostic indicator for the disease.

Hedgehog signaling downregulates suppressor of fused through the HIB/SPOP-Crn axis in Drosophila.

Hedgehog (Hh) signaling plays vital roles in animal development and tissue homeostasis, and its misregulation causes congenital diseases and several types of cancer. Suppressor of Fused (Su(fu)) is a conserved inhibitory component of the Hh signaling pathway, but how it is regulated remains poorly understood. Here we demonstrate that in Drosophila Hh signaling promotes downregulation of Su(fu) through its target protein HIB (Hh-induced BTB protein). Interestingly, although HIB-mediated downregulation of Su(fu) depends on the E3 ubiquitin ligase Cul3, HIB does not directly regulate Su(fu) protein stability. Through an RNAi-based candidate gene screen, we identify the spliceosome factor Crooked neck (Crn) as a regulator of Su(fu) level. Epistasis analysis indicates that HIB downregulates Su(fu) through Crn. Furthermore, we provide evidence that HIB retains Crn in the nucleus, leading to reduced Su(fu) protein level. Finally, we show that SPOP, the mammalian homologue of HIB, can substitute HIB to downregulate Su(fu) level in Drosophila. Our study suggests that Hh regulates both Ci and Su(fu) levels through its target HIB, thus uncovering a novel feedback mechanism that regulates Hh signal transduction. The dual function of HIB may provide a buffering mechanism to fine-tune Hh pathway activity.

Steroid receptor coactivator-3 regulates glucose metabolism in bladder cancer cells through coactivation of hypoxia inducible factor 1α.

Cancer cell proliferation is a metabolically demanding process, requiring high glycolysis, which is known as "Warburg effect," to support anabolic growth. Steroid receptor coactivator-3 (SRC-3), a steroid receptor coactivator, is overexpressed and/or amplified in multiple cancer types, including non-steroid targeted cancers, such as urinary bladder cancer (UBC). However, whether SRC-3 regulates the metabolic reprogramming for cancer cell growth is unknown. Here, we reported that overexpression of SRC-3 accelerated UBC cell growth, accompanied by the increased expression of genes involved in glycolysis. Knockdown of SRC-3 reduced the UBC cell glycolytic rate under hypoxia, decreased tumor growth in nude mice, with reduction of proliferating cell nuclear antigen and lactate dehydrogenase expression levels. We further revealed that SRC-3 could interact with hypoxia inducible factor 1α (HIF1α), which is a key transcription factor required for glycolysis, and coactivate its transcriptional activity. SRC-3 was recruited to the promoters of HIF1α-target genes, such as glut1 and pgk1. The positive correlation of expression levels between SRC-3 and Glut1 proteins was demonstrated in human UBC patient samples. Inhibition of glycolysis through targeting HK2 or LDHA decelerated SRC-3 overexpression-induced cell growth. In summary, overexpression of SRC-3 promoted glycolysis in bladder cancer cells through HIF1α to facilitate tumorigenesis, which may be an intriguing drug target for bladder cancer therapy.

Pao Pereira Extract Suppresses Castration-Resistant Prostate Cancer Cell Growth, Survival, and Invasion Through Inhibition of NFκB Signaling.

Pao extract, derived from bark of Amazonian tree Pao Pereira, is commonly used in South American medicine. A recent study showed that Pao extract repressed androgen-dependent LNCaP prostate cancer cell growth. We hypothesize that Pao extract asserts its anticancer effects on metastatic castration-resistant prostate cancer (CRPC) cells. Pao extract suppressed CRPC PC3 cell growth in a dose- and time-dependent manner, through induction of apoptosis and cell cycle arrest. Pao extract treatment induced cell cycle inhibitors, p21 and p27, and repressed PCNA, Cyclin A and Cyclin D1. Furthermore, Pao extract also induced the upregulation of pro-apoptotic Bax, reduction of anti-apoptotic Bcl-2, Bcl-xL, and XIAP expression, which were associated with the cleavage of PARP protein. Moreover, Pao extract treatment blocked PC3 cell migration and invasion. Mechanistically, Pao extract suppressed phosphorylation levels of AKT and NFκB/p65, NFκB DNA binding activity, and luciferase reporter activity. Pao inhibited TNFα-induced relocation of NFκB/p65 to the nucleus, NFκB/p65 transcription activity, and MMP9 activity as shown by zymography. Consistently, NFκB/p65 downstream targets involved in proliferation (Cyclin D1), survival (Bcl-2, Bcl-xL, and XIAP), and metastasis (VEGFa, MMP9, and GROα/CXCL1) were also downregulated by Pao extract. Finally, forced expression of NFκB/p65 reversed the growth inhibitory effect of Pao extract. Overall, Pao extract induced cell growth arrest, apoptosis, partially through inhibiting NFκB activation in prostate cancer cells. These data suggest that Pao extract may be beneficial for protection against CRPC.

EZH2 regulates cancer cell migration through repressing TIMP-3 in non-small cell lung cancer.

Histone methylations play important roles in human cancer metastasis. Enhancer of zeste homolog 2 (EZH2) is a key component of the polycomb repressor complex 2, which is responsible for histone H3K27 methylation. EZH2 is overexpressed in lung cancer and epigenetically silences tumor suppressor genes. Here, we showed that EZH2 was up-regulated in lung cancer and had a positive correlation with pathologic stage, nodal involvement in lung cancer patients. Moreover, overexpression of EZH2 was correlated with reduced tissue inhibitor of metalloproteinase-3 (TIMP-3) expression, which was shown to be negatively associated with tumor metastasis. Of note, overall survival time of patients with high EZH2/low TIMP-3 expression was significantly shorter than that of patients with low EZH2/high TIMP-3 (P = 0.031). RNA interfering and pharmacologic inhibition of EZH2 reduced histone H3 lysine 27 tri-methylation level and increased TIMP-3 expression level. Knockdown of EZH2 by siRNA significantly reduced A549 cancer cell migration. In contrast, reduction of TIMP-3 in A549 cells partially rescued EZH2 deficiency-induced loss of cell migration capacity. Taken together, our findings indicate that EZH2 accelerates cancer cell migration, in part, via the repression of TIMP-3 expression, suggesting a potential mechanism by which EZH2 promotes lung cancer progression and metastasis.