Hepatocellular Carcinoma and Nuclear Paraspeckles: Induction in Chemoresistance and Prediction for Poor Survival

Background/Aims: Hepatocellular carcinoma (HCC) represents the second most common cause of cancer-related deaths worldwide, not least due to its high chemoresistance. The long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1), localised in nuclear paraspeckles, has been shown to enhance chemoresistance in several cancer types. Since data on NEAT1 in HCC chemosensitivity are completely lacking and chemoresistance is linked to poor prognosis, we aimed to study NEAT1 expression in HCC chemoresistance and its link to HCC prognosis. Methods: NEAT1 expression was determined in either sensitive, or sorafenib, or doxorubicin resistant HepG2, PLC/PRF/5, and Huh7 cells by qPCR. Paraspeckles were detected by immunostaining of paraspeckle component 1 (PSPC1) in cell culture and in a cohort of HCC patients. PSPC1 expression was correlated with clinical data. The expression of transcript variants of NEAT1 and transcripts encoding the paraspeckle-associated proteins was analysed in the TCGA liver cancer data set. Results: NEAT1 was overexpressed in all three sorafenib and doxorubicin resistant cell lines. Paraspeckles were present in all chemoresistant cells, whereas no signal was detected in the sensitive cells. Expression of NEAT1 transcripts as well as transcripts encoding PSPC1, NONO, and RBM14 was increased in tumour tissue. Expression of PSPC1, NONO, and RBM14 transcripts was significantly associated with poor survival, whereas NEAT1 expression was not. Immunohistochemical analysis revealed that nuclear and cytoplasmic PSPC1-positivity was significantly associated with shorter overall survival of HCC patients. Conclusion: Our data show an induction of NEAT1 in HCC chemoresistance and a high correlation of transcripts encoding paraspeckle-associated proteins with poor survival in HCC. Therefore, NEAT1, PSPC1, NONO, and RBM14 might be promising targets for novel HCC therapies, and the paraspeckle-associated proteins might be clinical markers and predictors for poor survival in HCC

The mRNA-binding Protein TTP/ZFP36 in Hepatocarcinogenesis and Hepatocellular Carcinoma

Hepatic lipid deposition and inflammation represent risk factors for hepatocellular carcinoma (HCC). The mRNA-binding protein tristetraprolin (TTP, gene name ZFP36) has been suggested as a tumor suppressor in several malignancies, but it increases insulin resistance. The aim of this study was to elucidate the role of TTP in hepatocarcinogenesis and HCC progression. Employing liver-specific TTP-knockout (lsTtp-KO) mice in the diethylnitrosamine (DEN) hepatocarcinogenesis model, we observed a significantly reduced tumor burden compared to wild-type animals. Upon short-term DEN treatment, modelling early inflammatory processes in hepatocarcinogenesis, lsTtp-KO mice exhibited a reduced monocyte/macrophage ratio as compared to wild-type mice. While short-term DEN strongly induced an abundance of saturated and poly-unsaturated hepatic fatty acids, lsTtp-KO mice did not show these changes. These findings suggested anti-carcinogenic actions of TTP deletion due to effects on inflammation and metabolism. Interestingly, though, investigating effects of TTP on different hallmarks of cancer suggested tumor-suppressing actions: TTP inhibited proliferation, attenuated migration, and slightly increased chemosensitivity. In line with a tumor-suppressing activity, we observed a reduced expression of several oncogenes in TTP-overexpressing cells. Accordingly, ZFP36 expression was downregulated in tumor tissues in three large human data sets. Taken together, this study suggests that hepatocytic TTP promotes hepatocarcinogenesis, while it shows tumor-suppressive actions during hepatic tumor progression

Hepatische Steatose und Krebsentwicklung : die Rolle des insulin-like growth factor 2 mRNA bindenden Proteins p62 / IGF2BP2-2 / IMP2-2 und der Fettsäureelongase ELOVL6

The IGF2 mRNA binding protein p62/IGF2B2-2/IMP2-2 induces hepatic steatosis in mice, is overexpressed in HCC patients, and is associated with the overexpression of the tumorigenic growth factor IGF2. The underlying mechanisms involved in p62’s actions during liver disease are poorly characterized. Therefore, the aim of this study was to elucidate the mechanisms of p62 on lipogenesis and tumorigenesis. Since alterations in hepatic lipid and fatty acid composition are linked to liver pathogenesis, we analyzed liver lipids and fatty acid composition of p62 transgenic animals, and found that almost all lipid classes and fatty acids were elevated. The elevated ratio of C18 to C16 fatty acids was attributed to IGF2-mediated SREBF1 and ELOVL6 activation, and was shown to be responsible for steatosis development. Elevated levels of inflammatory markers provided evidence that p62 transgenic animals are susceptible to hepatic inflammation. Therefore, these results demonstrate an important role of p62 and ELOVL6 in the development of steatosis and steatohepatitis. Interestingly, though, ELOVL6 was downregulated in HCC patients and in a murine HCC model. It therefore seems unlikely that ELOVL6 has a pathophysiological role in HCC. Still, p62 amplifies HCC development, which might be attributed to DLK1/RAC1 mediated activation of the antiapoptotic ERK pathway. Taken together, our data underline the role of p62 in liver pathologies and provide evidence for p62 as a prognostic marker or therapeutic target.Das IGF2 mRNA bindende Protein p62/IGF2B2-2/IMP2-2 induziert im Mausmodell eine Fettleber, ist in HCC Patienten überexprimiert und erhöht die Expression des Wachstumsfaktors IGF2. Die Mechanismen, über die p62 die Leber-Pathophysiologie beeinflusst, sind weitgehend unbekannt. Ziel dieser Studie war die Aufklärung der Rolle von p62 in der Lipogenese und Karzinogenese. Veränderte Lipidstoffwechsel können zur Leberpathogenese beitragen. Die Untersuchung der hepatischen Fette aus den Lebern von p62 transgenen Tieren ergab, dass fast alle Lipide und Fettsäuren akkumuliert vorlagen. Das erhöhte Verhältnis von C18 zu C16 Fettsäuren, das durch eine IGF2-vermittelte Aktivierung von SREBF1 und ELOVL6 verursacht wurde, führte zur Steatose in diesen Tieren. Erhöhte inflammatorische Marker in den transgenen Lebern ließen auf eine verstärkte Entzündungsneigung schließen. Somit scheinen p62 und ELOVL6 eine wichtige Rolle in der Steatose- und der Steatohepatitis-Entstehung zu spielen. Interessanterweise war ELOVL6 aber im humanen und murinen HCC herrunterreguliert. Somit ist eine pathophysiologische Rolle von ELOVL6 in der Hepatokarzinogenese unwahrscheinlich. Die kanzerogenen Effekte von p62 scheinen durch die verstärkte Expression von DLK1 und RAC1, die den antiapoptotischen ERK Signalweg aktivieren, vermittelt zu werden. Unsere Daten belegen eine wichtige Rolle von p62 in der Leberpathophysiologie. Somit könnte p62 als Biomarker oder therapeutisches Target von Interesse sein

The IGF2 mRNA binding protein p62/IGF2BP2-2 induces fatty acid elongation as a critical feature of steatosis

Liver-specifi c overexpression of the insulin-like growth factor 2 ( IGF2 ) mRNA binding protein p62/ IGF2BP2-2 induces a fatty liver, which highly expresses IGF2 . Because IGF2 expression is elevated in patients with steatohepatitis, the aim of our study was to elucidate the role and interconnection of p62 and IGF2 in lipid metabolism. Expression of p62 and IGF2 highly correlated in human liver disease. p62 induced an elevated ratio of C18:C16 and increased fatty acid elongase 6 (ELOVL6) protein, the enzyme catalyzing the elongation of C16 to C18 fatty acids and promoting nonalcoholic steatohepatitis in mice and humans. The p62 overexpression induced the activation of the ELOVL6 transcriptional activator sterol regulatory element binding transcription factor 1 (SREBF1). Recombinant IGF2 induced the nuclear translocation of SREBF1 and a neutralizing IGF2 antibody reduced ELOVL6 and mature SREBF1 protein levels. Concordantly, p62 and IGF2 correlated with ELOVL6 in human livers. Decreased palmitoyl-CoA levels, as found in p62 transgenic livers, can explain the lipogenic action of ELOVL6. Accordingly, p62 represents an inducer of hepatic C18 fatty acid production via a SREBF1-dependent induction of ELOVL6. These fi ndings underline the detrimental role of p62 in liver disease. © 2014 by the American Society for Biochemistry and Molecular Biology, Inc

Hepatocellular carcinoma and nuclear paraspeckles : induction in chemoresistance and prediction for poor survival

Background/Aims: Hepatocellular carcinoma (HCC) represents the second most common cause of cancer-related deaths worldwide, not least due to its high chemoresistance. The long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1), localised in nuclear paraspeckles, has been shown to enhance chemoresistance in several cancer types. Since data on NEAT1 in HCC chemosensitivity are completely lacking and chemoresistance is linked to poor prognosis, we aimed to study NEAT1 expression in HCC chemoresistance and its link to HCC prognosis. Methods: NEAT1 expression was determined in either sensitive, or sorafenib, or doxorubicin resistant HepG2, PLC/PRF/5, and Huh7 cells by qPCR. Paraspeckles were detected by immunostaining of paraspeckle component 1 (PSPC1) in cell culture and in a cohort of HCC patients. PSPC1 expression was correlated with clinical data. The expression of transcript variants of NEAT1 and transcripts encoding the paraspeckle-associated proteins was analysed in the TCGA liver cancer data set. Results: NEAT1 was overexpressed in all three sorafenib and doxorubicin resistant cell lines. Paraspeckles were present in all chemoresistant cells, whereas no signal was detected in the sensitive cells. Expression of NEAT1 transcripts as well as transcripts encoding PSPC1, NONO, and RBM14 was increased in tumour tissue. Expression of PSPC1, NONO, and RBM14 transcripts was significantly associated with poor survival, whereas NEAT1 expression was not. Immunohistochemical analysis revealed that nuclear and cytoplasmic PSPC1-positivity was significantly associated with shorter overall survival of HCC patients. Conclusion: Our data show an induction of NEAT1 in HCC chemoresistance and a high correlation of transcripts encoding paraspeckle-associated proteins with poor survival in HCC. Therefore, NEAT1, PSPC1, NONO, and RBM14 might be promising targets for novel HCC therapies, and the paraspeckle-associated proteins might be clinical markers and predictors for poor survival in HCC