Honggang Yu

Phosphatase and Tensin Homolog (PTEN) Represses Colon Cancer Progression through Inhibiting Paxillin Transcription via PI3K/AKT/NF-κB Pathway

The tumor suppressor gene phosphatase and tensin homolog (PTEN) is frequently mutated in colon cancer. However, the potential contribution of loss of PTEN to colon cancer progression remains unclear. In this study, we demonstrated that PTEN overexpression or knockdown in Lovo colon cancer cells decreased or increased paxillin expression, respectively. Moreover, paxillin reversed PTEN-mediated inhibition of Lovo cell invasion and migration. Overexpression of PTEN in an orthotropic colon cancer nude mice model inhibited tumor formation and progression. In addition, PTEN protein level was negatively correlated with that of paxillin in human colon cancer tissues. Mechanistically, we identified three NF-κB binding sites on paxillin promoter and confirmed that paxillin was a direct transcriptional target of NF-κB. Our findings reveal a novel mechanism by which PTEN inhibits the progression of colon cancer by inhibiting paxillin expression downstream of PI3K/AKT/NF-κB pathway. Thereby, PTEN/PI3K/AKT/NF-κB/paxillin signaling cascade is an attractive therapeutic target for colon cancer progression.Background: The tumor suppressor gene PTEN is associated with colon cancer progression. Results: PAXILLIN contributes to the inhibition of progression by PTEN. Conclusion: PTEN represses colon cancer progression through inhibiting paxillin transcription via PI3K/AKT/NF-κB pathway. Significance: PTEN/PI3K/AKT/NF-κB/paxillin signaling cascade is an attractive therapeutic target for colon cancer progression. The tumor suppressor gene phosphatase and tensin homolog (PTEN) is frequently mutated in colon cancer. However, the potential contribution of loss of PTEN to colon cancer progression remains unclear. In this study, we demonstrated that PTEN overexpression or knockdown in Lovo colon cancer cells decreased or increased paxillin expression, respectively. Moreover, paxillin reversed PTEN-mediated inhibition of Lovo cell invasion and migration. Overexpression of PTEN in an orthotropic colon cancer nude mice model inhibited tumor formation and progression. In addition, PTEN protein level was negatively correlated with that of paxillin in human colon cancer tissues. Mechanistically, we identified three NF-κB binding sites on paxillin promoter and confirmed that paxillin was a direct transcriptional target of NF-κB. Our findings reveal a novel mechanism by which PTEN inhibits the progression of colon cancer by inhibiting paxillin expression downstream of PI3K/AKT/NF-κB pathway. Thereby, PTEN/PI3K/AKT/NF-κB/paxillin signaling cascade is an attractive therapeutic target for colon cancer progression.

Lysophosphatidic acid stimulates activation of focal adhesion kinase and paxillin and promotes cell motility, via LPA1-3, in human pancreatic cancer.

BackgroundPancreatic cancer is highly metastatic and with poor prognosis. In previous studies, lysophosphatidic acid (LPA) was shown to be a critical component of ascites which promoted the invasion and metastasis of pancreatic cancer. Two focal adhesion proteins, focal adhesion kinase (FAK) and paxillin, were crucially involved in cell migration, cytoskeleton reorganization, and the dynamics of focal adhesion.ObjectivesThis study examined the involvement of LPA1–3 in LPA-induced activation of FAK and paxillin, and in cell motility, in pancreatic cancer PANC-1 cells.MethodsReverse transcriptase polymerase chain reaction analysis was used to examine mRNA expression of LPA receptors in PANC-1. Cellular protein expression of FAK and paxillin was analyzed by western blotting. The subcellular location of FAK and paxillin was visualized by immunofluorescence. Cell migration was measured by use of a transwell migration chamber.ResultsThree LPA receptors (LPA1, LPA2, and LPA3) were significantly expressed in PANC-1 cells. Treatment with LPA induced both time and dose-dependent tyrosine phosphorylation of FAK and paxillin. LPA also affected translocation of FAK and paxillin from cytoplasm to focal adhesions at the cell periphery and enhanced cell motility of PANC-1. Pretreatment with 3-(4-(4-((1-(2-chlorophenyl)ethoxy)carbonyl amino)-3-methyl-5-isoxazolyl)benzylsulfanyl)propanoic acid (Ki16425), an antagonist of LPA1 and LPA3, before LPA attenuated the LPA-induced tyrosine phosphorylation and redistribution of FAK and paxillin and abrogated LPA-induced cellular migration activity.ConclusionsThese results suggest LPA induces activation of FAK and paxillin via LPA1–3, which may contribute to the increased cell motility in human pancreatic cancer PANC-1 cells. Thus, an understanding of the regulation by LPA of cell motility in pancreatic cancer could identify novel targets for therapy.

Thymoquinone Pretreatment Overcomes the Insensitivity and Potentiates the Antitumor Effect of Gemcitabine Through Abrogation of Notch1, PI3K/Akt/mTOR Regulated Signaling Pathways in Pancreatic Cancer.

BackgroundThe gemcitabine-insensitivity remains the main challenge for pancreatic cancer treatment. Thymoquinone, the predominant bioactive ingredient of Nigella sativa, has been shown to possess promising anti-cancer and chemo-sensitizing effects on pancreatic cancer, however, its meticulous mechanism is still indistinct.AimThe objective of the present study was to investigate the potency of thymoquinone in combination with gemcitabine in inducing apoptosis and preventing the development of gemcitabine-insensitivity in pancreatic cancer cells.MethodsThe anti-tumor effects of thymoquinone and gemcitabine were analyzed via evaluation of alterations of cell viability, tumor weight, apoptosis-related proteins, caspase-3, -9 activities and NF-κB DNA binding activity in pancreatic cancer cells in vitro and PANC-1 cells orthotopic xenograft in vivo.ResultsThymoquinone pretreatment following gemcitabine treatment synergistically caused an increase in pancreatic cancer cells apoptosis and tumor growth inhibition both in vitro and in vivo. The novel combinational regimen also contributes to alterations of multiple molecular signaling targets, such as the suppression of Notch1, NICD accompanying with up-regulation of PTEN, the inactivation of Akt/mTOR/S6 signaling pathways, and the suppression of phosphorylation and nuclear translocation of p65 induced by TNF-α. Thymoquinone pretreatment and gemcitabine also induced down-regulation of anti-apoptotic Bcl-2, Bcl-xL, XIAP and up-regulation and activation of pro-apoptotic molecules including Caspase-3, Caspase-9, Bax and increased release of cytochrome c.ConclusionsThis novel modality of thymoquinone pretreatment can enhance the anti-cancer activity of gemcitabine and may be a promising option in the treatment of pancreatic cancer.

Overexpression of AKT decreases the chemosensitivity of gastric cancer cells to cisplatin in vitro and in vivo

Cisplatin (CDDP) is one of the most efficacious and widely used cytotoxic anticancer drugs used for the treatment of numerous types of cancer. However, its efficacy is limited as a result of acquired drug resistance. AKT overexpression may provide a potential mechanism leading to the resistance of human gastric cancer cells; however, the precise mechanism of the development of CDDP drug resistance remains uncertain. In the present study, we demonstrate that CDDP resistance is associated with AKT overexpression at the cellular and molecular level. We also observed that increased expression levels of AKT were sufficient to inhibit the resistance of gastric cancer cells to CDDP and that overexpressed AKT interacted with reactive oxygen species which were generated by CDDP. These results indicate that AKT activity is essential for the regulation of CDDP resistance in gastric cancer cells. Our results further demonstrate that AKT induces gastric cancer cells to become resistant to CDDP through the Janus kinasexa02 (JAK2)/signal transducer and activator of transcriptionxa03 (STAT3) signaling pathway. Taken together, these data support a potential role for AKT overexpression and the JAK2/STAT3 pathway in the development of CDDP drug resistance in human gastric cancer cells. We hypothesize that AKT may represent a future pharmacological target for the inhibition of CDDP resistance in human cancer.

Mutations in the S gene and in the overlapping reverse transcriptase region in chronic hepatitis B Chinese patients with coexistence of HBsAg and anti-HBs

BACKGROUNDnThe mechanism underlying the coexistence of hepatitis B surface antigen and antibodies to HBsAg in chronic hepatitis B patients remains unknown.nnnAIMSnThis research aimed to determine the clinical and virological features of the rare pattern.nnnMETHODSnA total of 32 chronic hepatitis B patients infected by HBV genotype C were included: 15 carrying both HBsAg and anti-HBs (group I) and 17 solely positive for HBsAg (group II). S gene and reverse transcriptase region sequences were amplified, sequenced and compared with the reference sequences.nnnRESULTSnThe amino acid variability within major hydrophilic region, especially the a determinant region, and within reverse transcriptase for regions overlapping the major hydrophilic region in group I is significantly higher than those in group II. Mutation sI126S/T within the a determinant was the most frequent change, and only patients from group I had the sQ129R, sG130N, sF134I, sG145R amino acid changes, which are known to alter immunogenicity.nnnCONCLUSIONSnIn chronic patients, the concurrent HBsAg/anti-HBs serological profile is associated with an increased aa variability in several key areas of HBV genome. Additional research on these genetic mutants are needed to clarify their biological significance for viral persistence.

Downregulation of TRIM21 contributes to hepatocellular carcinoma carcinogenesis and indicates poor prognosis of cancers

The aim of our work is to clarify the clinical implication and functional role of tripartite motif 21 (TRIM21) in hepatocellular carcinoma (HCC). We validated that TRIM21 was downregulated in liver cancer samples by immunohistochemical (IHC) staining. We also demonstrated that its downregulation was associated with several clinicopathologic features such as tumor numbers, T stage, Barcelona Clinic Liver Cancer (BCLC) stage, and Cancer of the Liver Italian Program (CLIP) stage of HCC patients. Importantly, the expression of TRIM21 in tumor samples is significantly correlated with the prognosis of the patients. We further silenced TRIM21 in HCC cell HepG2 and LM3 and confirmed that TRIM21 silencing will promote cancer cell proliferation (CCK-8 assay), colony forming (plate colony-forming assay), migration (transwell assay), and the ability of antiapoptosis (annexin V-FITC/PI staining) in vitro. Then, we predicted gene sets influenced by TRIM21 by using bioinformatic tools. For the first time, we prove that TRIM21 is a potential tumor suppressor in HCC and its low expression indicates poor prognosis. Our findings provide useful insight into the mechanism of HCC origin and progression and offer clues to novel HCC therapies.

SSRP1 Contributes to the Malignancy of Hepatocellular Carcinoma and Is Negatively Regulated by miR-497

The aim of this study is to clarify the clinical implication and functional role of structure specific recognition protein 1 (SSRP1) in hepatocellular carcinoma (HCC) and explore the underlying mechanism of aberrant high expression of SSRP1 in cancers. In the present investigation, we validated that SSRP1 was upregulated in HCC samples. We also demonstrated that its upregulation was associated with several clinicopathologic features such as higher serum AFP level, larger tumor size, and higher T stage of HCC patients; and its high expression indicated shorter overall survival and faster recurrence. To investigate the role of SSRP1 in HCC progression, both loss- and gain-function models were established. We demonstrated that SSPR1 modulated both proliferation and metastasis of HCC cells in vitro and vivo. Furthermore, we demonstrated that SSRP1-modulated apoptosis process and its knockdown increased the sensitivity of HCC cells to doxorubicin, 5-Fluorouracil, and cisplatin. We also identified microRNA-497 (miR-497) as a posttranscriptional regulator of SSRP1. Ectopic expression of miR-497 inhibited 3-untranslated-region–coupled luciferase activity and suppressed endogenous SSRP1 expression at both messenger RNA and protein levels. For the first time, we proved that SSRP1 upregulation contributed to HCC development and the tumor-suppressive miR-497 served as its negative regulator.

Aberrant high expression level of MORC2 is a common character in multiple cancers

Microrchidia 2 (MORC2) plays important roles in DNA damage repair and lipogenesis, but the clinical and functional role of MORC2 in cancer remains largely unexplored. In this study, we showed that MORC2 was widely expressed in human tissues while significantly up-regulated in most cancer types using immunohistochemical staining and analysis of messenger RNA expression profile of more than 2000 human tissue samples from 15 different organs (lung, prostate, liver, breast, brain, stomach, colon/rectum, pancreas, ovary, endometrium, skin, nasopharynx, kidney, esophagus, and bladder). We also found that the MORC2 expression level in high-grade cancer tissues was much more elevated and associated with unfavorable pathological characteristics, poor overall survival, and disease-free survival in several kinds of cancers such as non-small cell lung cancer and breast cancer. Gene set enrichment analysis was used to predict the genes modulated by MORC2, and the results showed that dysregulation of MORC2 in tumor may take part in the cell cycle regulation and genomic instability. We observed that MORC2 knockdown would arrest the cell cycle progress, and the genome of tumors with high MORC2 expression contained more point mutations and gene copy number variation, which validates our gene set enrichment analysis results. The results also showed that MORC2 knockdown would significantly inhibit the proliferation, colony forming, migration, and invasion in multiple cancer cell lines. Taken together, these results highlight the importance of MORC2 in tumorigenesis and cancer progression, and it may act as a potential diagnostic marker and therapeutic target for these diseases.

AGXT2L1 is down-regulated in heptocellular carcinoma and associated with abnormal lipogenesis

Aims To clarify the clinical implications and functional role of the alanine-glyoxylate aminotransferase 2-like 1 (AGXT2L1) gene in hepatocellular carcinoma (HCC). Methods and results We confirmed that AGXT2L1 was down-regulated in liver cancer samples by immunohistochemical (IHC) staining. We also demonstrated that this down-regulation was associated with several clinicopathological features such as alpha fetoprotein (AFP) serum level and T stage. Furthermore, we showed with Kaplan-Meier analysis that expression of AGXT2L1 in tumour samples was significantly correlated with patient prognosis. The bioinformatic tool indicated that AGXT2L1 plays a role in the lipid metabolic process of HCC tissue, while siRNA silenced the expression of AGXT2L1 in HCC 97H and LM3 cells, confirming that down-regulation of AGXT2L1 promotes the lipogenesis of cancer cells. Conclusions For the first time, we have shown that AGXT2L1 is down-regulated in HCC and its low expression indicates a poor prognosis. Our findings also demonstrated that AGXT2L1 is a crucial gene in the abnormal lipogenesis of HCC tissue.

Function of the macrophage‑capping protein in colorectal carcinoma

To investigate the role of macrophage-capping protein (CapG) in the development and progression of colorectal carcinoma (CRC), immunohistochemistry (IHC), Kaplan-Meier survival analysis, wound healing and Transwell migration assays were performed. The IHC results demonstrated that CapG was relatively highly expressed in CRC tissue compared with non-tumor tissue (P<0.001), and that the expression of CapG was significantly associated with the tumor site, differentiation, lymph node metastasis and clinical stage (P=0.021, P=0.036, P=0.012 and P=0.009, respectively). Wound healing and Transwell migration assays demonstrated that the reduction of CapG expression in a CRC cell line by RNA interference was associated with significantly impaired motility (P<0.001). Kaplan-Meier survival analysis revealed that the expression of CapG in tumor samples was not significantly associated with disease-free survival time. In conclusion, CapG was overexpressed in CRC and was associated with tumor progression; therefore, it may be a useful prognostic biomarker and therapeutic target in CRC.