PDE4DIP contributes to colorectal cancer growth and chemoresistance through modulation of the NF1/RAS signaling axis.

Phosphodiesterase 4D interacting protein (PDE4DIP) is a centrosome/Golgi protein associated with cyclic nucleotide phosphodiesterases. PDE4DIP is commonly mutated in human cancers, and its alteration in mice leads to a predisposition to intestinal cancer. However, the biological function of PDE4DIP in human cancer remains obscure. Here, we report for the first time the oncogenic role of PDE4DIP in colorectal cancer (CRC) growth and adaptive MEK inhibitor (MEKi) resistance. We show that the expression of PDE4DIP is upregulated in CRC tissues and associated with the clinical characteristics and poor prognosis of CRC patients. Knockdown of PDE4DIP impairs the growth of KRAS-mutant CRC cells by inhibiting the core RAS signaling pathway. PDE4DIP plays an essential role in the full activation of oncogenic RAS/ERK signaling by suppressing the expression of the RAS GTPase-activating protein (RasGAP) neurofibromin (NF1). Mechanistically, PDE4DIP promotes the recruitment of PLCγ/PKCε to the Golgi apparatus, leading to constitutive activation of PKCε, which triggers the degradation of NF1. Upregulation of PDE4DIP results in adaptive MEKi resistance in KRAS-mutant CRC by reactivating the RAS/ERK pathway. Our work reveals a novel functional link between PDE4DIP and NF1/RAS signal transduction and suggests that targeting PDE4DIP is a promising therapeutic strategy for KRAS-mutant CRC.

RSPO2 promotes progression of ovarian cancer through dual receptor-mediated FAK/Src signaling activation.

R-spondin 2 (RSPO2) drives the potentiation of Wnt signaling and is implicated in tumorigenesis in multiple cancers, but its role in ovarian cancer has not been investigated. Here, we reported that RSPO2 promoted the growth and metastasis of ovarian cancer through the activation of FAK/Src signaling cascades. RSPO2 enhanced the autophosphorylation of FAK and Src through a unique dual receptors mechanism. First, RSPO2-LGR4 interaction prevented the endocytic degradation of LGR4 and promoted LGR4-mediated translocation of Src to the plasma membrane. Second, RSPO2 directly bound to integrin ß3 as a ligand and enhanced the stability of integrins, and both actions potentiated autoactivation of FAK and/or Src in ovarian cancer cells. RSPO2 expression was increased in ovarian tumors and was associated with poor prognosis in patients. Our study highlights the importance of RSPO2 in ovarian tumor progression and suggests that targeting RSPO2/FAK/Src cascades may constitute potential approaches to inhibit the progression of aggressive ovarian cancer.

NLRC3 deficiency promotes cutaneous wound healing due to the inhibition of p53 signaling.

Cutaneous wound healing is a complicated process that is characterized by an initial inflammatory phase followed by a proliferative phase. NLRC3 plays important roles in innate immunity, inflammatory regulation and tumor cell growth. However, the function of NLRC3 in wound healing remains unclear. Here, we investigated the function of NLRC3 in acute cutaneous wound healing using Nlrc3 gene knockout (Nlrc3-/-) mice. Our results demonstrated that skin wound repair in Nlrc3-/- mice was significantly accelerated compared with that in wild-type (WT) mice. NLRC3 deficiency promoted the inflammatory and proliferative phases in wounds enhanced the inflammatory response and increased re-epithelialization and granulation tissue formation, and these phenotypes were primarily ascribed to regulatory effects on p53 signaling. Mechanistically, we uncovered novel crosstalk between NLRC3 and p53 signaling and revealed that NLRC3 could mediate the ubiquitination and degradation of p53 in an Hsp90-dependent manner. In conclusion, our study suggests that NLRC3 is a critical negative regulator of the inflammatory response and cell proliferation during wound healing and that blocking NLRC3 may represent a potential approach for accelerating wound healing.

Long noncoding RNA LINC00930 promotes PFKFB3-mediated tumor glycolysis and cell proliferation in nasopharyngeal carcinoma.

BACKGROUND: Metabolic reprogramming is a hallmark of cancer. However, the roles of long noncoding RNAs (lncRNAs) in cancer metabolism, especially glucose metabolism remain largely unknown. RESULTS: In this study, we identified and functionally characterized a novel metabolism-related lncRNA, LINC00930, which was upregulated and associated with tumorigenesis, lymphatic invasion, metastasis, and poor prognosis in nasopharyngeal carcinoma (NPC). Functionally, LINC00930 was required for increased glycolysis activity and cell proliferation in multiple NPC models in vitro and in vivo. Mechanistically, LINC00930 served as a scaffold to recruit the RBBP5 and GCN5 complex to the PFKFB3 promoter and increased H3K4 trimethylation and H3K9 acetylation levels in the PFKFB3 promoter region, which epigenetically transactivating PFKFB3, and thus promoting glycolytic flux and cell cycle progression. Clinically, targeting LINC00930 and PFKFB3 in combination with radiotherapy induced tumor regression. CONCLUSIONS: Collectively, LINC00930 is mechanistically, functionally and clinically oncogenic in NPC. Targeting LINC00930 and its pathway may be meaningful for treating patients with NPC.

Hsa_circ_001659 serves as a novel diagnostic and prognostic biomarker for colorectal cancer.

Colorectal cancer (CRC) is prevalent worldwide and novel diagnostic and prognostic biomarkers are needed to improve precision medicine. Circular RNAs (circRNAs) are currently being considered as emerging tumor biomarkers. Herein, we aimed to explore the possible clinical application of circRNAs in the early diagnosis and prognostic prediction of CRC. First, candidate circRNA was selected by integrating analysis of Gene Expression Omnibus (GEO) database using GEO2R program. ROC curve analysis demonstrated the predictive values and likelihood ratios of circ_001659 were satisfactory for the diagnosis of CRC, including patients in early-stage disease or patients with carcinoembryonic antigen (CEA)-negative status. Moreover, serum circ_001659 may be a novel biomarker in the assessment of successful treatment and remission of cancer tracking. We further investigated the oncogenic role of circ_001659. In vivo and in vitro experiments indicated that circ_001659 could promote CRC cell invasion and migration. Mechanistically, circ_001659 was localized in the nucleus, recruited the RBBP5 to Vimentin promoter and increased H3K4 trimethylation level on the Vimentin promoter region, which epigenetically activated Vimentin transcription. Our findings demonstrate that circ_001659 could be a useful serum biomarker for CRC diagnosis and prognosis. Targeting circ_001659 and its pathway may be meaningful for treating patients with CRC.

Anti-TGF-ß attenuates tumor growth via polarization of tumor associated neutrophils towards an anti-tumor phenotype in colorectal cancer.

Tumor associated neutrophils (TANs) play important roles in the progress of CRC. Since tumor microenvironments could influence the phenotypes of TANs, altering the tumor microenvironment to polarize the phenotype of TANs may be a new strategy for tumor treatment. This study aims to investigate the effect of anti-TGF-ß on the polarization of TANs from a pro-tumor phenotype towards an anti-tumor phenotype in CRC. In this work, CRC patients had more infiltration of TANs and higher expression of TGF-ß in CRC tissue when compared with the controls. In vitro, SW480 cells were co-cultured with primed neutrophils, which simulated the TANs in the tumor microenvironment, and TGF-ß was blocked by anti-TGF-ß (1D11) in order to polarize TANs. Anti-TGF-ß treatment increased the cytotoxicity of TANs and decreased the metastatic chemoattractants secreted by TANs, and ultimately increased the apoptosis of CRC cells significantly while remarkably suppressing the migration of tumor cells. The changes of signaling pathways in the TANs and tumor cells were explored. The results showed that anti-TGF-ß attenuated CRC may be partly mediated by suppression of PI3K/AKT signaling pathways in TANs and partly mediated by suppression of TGF-ß/Smad signaling pathways in tumor cells. Furthermore, the tumor in the mice treated with 1D11 was obviously smaller and had reverse tumorigenesis compared with the controls, while neutrophil depletion reduced the anti-tumor effect of 1D11. Our data suggest that anti-TGF-ß attenuates tumor growth via the polarization of TANs to an anti-tumor phenotype in CRC, which provides new strategies for CRC treatment.

Resveratrol Promoted Interferon-α-Induced Growth Inhibition and Apoptosis of SMMC7721 Cells by Activating the SIRT/STAT1.

Interferon-α (IFN-α) resistance is a major hurdle in the treatment of hepatocellular carcinoma (HCC). Signal transducers and activators of transcription 1 (STAT1) play a key role in exerting the antiproliferative and proapoptotic effects of IFN-α on tumors. In this study, we aimed to investigate whether resveratrol can promote IFN-α-induced growth inhibition and the apoptosis on HCC cells through the SIRT/STAT1 pathway. We found that IFN-α induced growth inhibition and apoptosis of SMMC7721 cells, and the effects could be significantly enhanced and blocked by resveratrol and EX527, respectively. Resveratrol not only activated SIRT1 but also induced phosphorylation of STAT1. Further study revealed that ablation of STAT1 reduced the combined antitumor effects of IFN-α and resveratrol, lowered the rate of apoptosis, and improved the viability of SMMC7721 cells. Whereas STAT1 overexpression strengthened the combined antitumor effects of resveratrol and IFN-α. Our findings suggest a novel strategy of using resveratrol to enhance the response of HCC to IFN-α treatment through the SIRT/STAT1 pathway.

Poly(ADP-Ribose) Polymerase Inhibitor PJ34 Attenuated Hepatic Triglyceride Accumulation in Alcoholic Fatty Liver Disease in Mice.

Poly(ADP-ribose) polymerase (PARP) is an NAD-consuming enzyme and its specific role in the pathogenesis of alcoholic fatty liver disease (AFLD) remains elusive. In this study, we applied PJ34 [N-(5,6-dihydro-6-oxo-2-phenanthridinyl)-2-acetamide hydrochloride] to inhibit hepatic PARP activity to examine the corresponding pathologic alteration in AFLD in mice and the underlying molecular mechanism. We found that PJ34 decreased the intracellular triglyceride (TG) content in hepatocytes. Moreover, PJ34 suppressed the gene expression of diglyceride acyltransferases DGAT1 and DGAT2 and elevated intracellular NAD+ levels in hepatocytes. These mechanistic observations were validated in alcohol-fed mice injected with PJ34 intraperitoneally. Our results indicate that the PJ34 injection attenuated hepatic TG accumulation in alcohol-fed mice. Furthermore, PJ34 injection lowered the gene expression of hepatic sterol regulatory element binding protein 1c, DGAT1, and DGAT2, whereas PJ34 injection augmented hepatic NAD+ levels in alcohol-fed mice. Finally, nicotinamide riboside supplementation alleviated hepatic TG accumulation in alcohol-fed mice. These data indicate that applying PARP-specific inhibitor PJ34 by intraperitoneal injection attenuated hepatic NAD+ depletion and TG accumulation in alcohol-fed mice and may be a potential candidate for use in AFLD therapy.

Nrf2 mediates the protective effects of homocysteine by increasing the levels of GSH content in HepG2 cells.

Homocysteine (Hcy) and glutathione (GSH) are crucial reduction­oxidation mediators. The underlying mechanisms governing the effects of Hcy on GSH generation in the progression of alcoholic liver disease has so far received little attention. The present study hypothesized that the antioxidant transcriptional factor nuclear factor (erythroid­derived 2)­like 2 (Nrf2) may participate in Hcy­mediated regulation of GSH production in HepG2 human liver cancer cells. MTT assay was used to study the cytotoxicity of homocysteine, western blot analysis and immunofluorescence staining were used to determine the effect of Hcy on Nrf2 expression. Our data demonstrated that HepG2 cells exposed to exogenous levels of Hcy (0­100 µM) exhibited elevated GSH levels in a concentration­dependent manner. Furthermore, 4­hydroxynonenal (4­HNE)­induced cell injury was attenuated by Hcy; however, this protective effect was blocked by the GSH­production inhibitor buthionine sulfoximine. Hcy treatment was able to induce Nrf2 protein expression in HepG2 cells. Treatment with the Nrf2 activator tert­butylhydroquinone (0­100 µM) increased GSH expression in a concentration­dependent manner; however, Nrf2­siRNA abolished the Hcy­induced increase in GSH expression and cellular protection in 4­HNE­stressed HepG2 cells. In conclusion, the antioxidant transcriptional factor Nrf2 was demonstrated to mediate the Hcy­induced increase in GSH expression levels and cellular protection in HepG2 cells.

STAT1 inhibits human hepatocellular carcinoma cell growth through induction of p53 and Fbxw7.

BACKGROUND: Aberrant STAT1 signaling is observed in human hepatocellular carcinoma (HCC) and has been associated with the modulation of cell proliferation and survival. However, the role of STAT1 signaling in HCC and its underlying mechanism remain elusive. METHODS: We transiently transfected pcDNA3.1-STAT1 and STAT1 siRNA into SMMC7721 and HepG2 cells. Western blot and qRT-PCR examined the expression of protein and RNA of target genes. Cell viability was assessed using MTT assay, and cell cycle and apoptosis were analyzed by flow cytometry. RESULTS: We found that STAT1 overexpression increased protein expression of p53 and Fbxw7, and downregulated the expression of cyclin A, cyclin D1, cyclin E, CDK2, Hes-1 and NF-κB p65. These changes led to growth inhibition and induced G0/G1 cell cycle arrest and apoptosis in SMMC7721 and HepG2 cells. Conversely, ablation of STAT1 had the opposite effect on p53, Fbxw7, Hes-1, NF-κB p65, cyclin A, cyclin D1, cyclin E and CDK2, and improved the viability of SMMC7721 and HepG2 cells. CONCLUSIONS: Our data indicate that STAT1 exerts tumor-suppressive effects in hepatocarcinogenesis through induction of G0/G1 cell cycle arrest and apoptosis, and may provide a basis for the design of new therapies for the intervention of HCC in the clinic.

RBP-J-interacting and tubulin-associated protein induces apoptosis and cell cycle arrest in human hepatocellular carcinoma by activating the p53-Fbxw7 pathway.

Aberrant Notch signaling is observed in human hepatocellular carcinoma (HCC) and has been associated with the modulation of cell growth. However, the role of Notch signaling in HCC and its underlying mechanism remain elusive. RBP-J-interacting and tubulin-associated (RITA) mediates the nuclear export of RBP-J to tubulin fibers and downregulates Notch-mediated transcription. In this study, we found that RITA overexpression increased protein expression of p53 and Fbxw7 and downregulated the expression of cyclin D1, cyclin E, CDK2, Hes-1 and NF-κB p65. These changes led to growth inhibition and induced G0/G1 cell cycle arrest and apoptosis in SMMC7721 and HepG2 cells. Our findings indicate that RITA exerts tumor-suppressive effects in hepatocarcinogenesis through induction of G0/G1 cell cycle arrest and apoptosis and suggest a therapeutic application of RITA in HCC.

Inhibitory effect of baicalin on collagen-induced arthritis in rats through the nuclear factor-κB pathway.

This study focused on the potential therapeutic effect of baicalin on collagen-induced arthritis (CIA) in rats and the underlying mechanisms. The CIA rats were injected with baicalin (50, 100, or 200 mg/kg) once daily for 30 days. The rats were monitored for clinical severity of arthritis, and joint tissues were used for radiographic assessment and histologic examination. We quantified tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) in experimental animals and used Western blots to assess levels of protein abundance, phosphorylation, and acetylation of nuclear factor (NF)-κB p65 and sirtuin 1 (sirt1) protein expression in joint tissues. Human fibroblast-like synoviocytes from rheumatoid arthritis (HFLS-RA) were adopted in further mechanistic investigations. Baicalin intraperitoneal injection for 30 days dose-dependently blocked clinical manifestations of CIA, such as functional impairment and swollen red paws. Meanwhile, it alleviated collagen-induced joint inflammation injury and inhibited the secretion of TNF-α and IL-1ß in both rat synovium and HFLS-RA. Further mechanistic investigations revealed that baicalin suppresses NF-κB p65 protein expression and phosphorylation in synovial tissue and human-derived synoviocytes. Moreover, the acetylation of NF-κB p65 was downregulated by baicalin, which negatively correlates with the baicalin-induced upregulation of sirt1 expression in the same conditions. The data indicate that CIA in rats can be alleviated by baicalin treatment via relieving joint inflammation, which is related to the suppression of synovial NF-κB p65 protein expression and the elevation of its deacetylation by sirt1.