Lin Zheng

Association between Hepatic Steatosis and Entecavir Treatment Failure in Chinese Patients with Chronic Hepatitis B

Background The coexistence of HBV infection and nonalcoholic fatty liver disease (NAFLD) becomes characteristic of liver disease in China, with unknown bilateral influence. We aimed to investigate the effect of hepatic steatosis, a common hepatocyte change in NAFLD, on antiviral therapy in patients with chronic hepatitis B (CHB). Methods and Findings We carried out a prospective nested case control study in CHB patients receiving Entecavir for initial antiviral therapy, by recording demographic, anthropometric and clinical data at baseline, 24wk, 48wk and 96wk. Univariate analysis and multivariate logistic regression were applied to find out independent factors of hepatic steatosis and Entecavir treatment failure. The rates of HBV-DNA clearance, HBeAg seroconversion and ALT normalization were compared between CHB patients with and without steatosis by post hoc analysis. A total of 267 Chinese patients with CHB entered final analysis, with overall percentages of hepatic steatosis and HBeAg positive as 30.5% and 62.4%. Multivariate analysis showed waist circumference, serum TG and uric acid levels were independent factors of hepatic steatosis. The response rates to Entecavir were 54.9%, 63.8%, 74.2% at 24wk, 48wk and 96wk. Hepatic steatosis was revealed as an independent factor of Entecavir treatment failure by multivariate logistic regression at 24wk, 48wk and 96wk. In CHB patients with hepatic steatosis, HBV-DNA clearance and HBeAg seroconversion were both lower throughout the follow-up, but only the former reached statistical significance. Besides, ALT normalization was also significantly lower at 24wk and 48wk. Conclusion Hepatic steatosis is significantly associated with Entecavir treatment failure and metabolic factors are independent factors of hepatic steatosis in CHB patients, which called for a specified antiviral strategy in CHB patients with NAFLD.

The role of Ursodeoxycholic acid in non-alcoholic steatohepatitis: a systematic review

BackgroundNon-alcoholic steatohepatitis (NASH) is a condition that occurs during the progression of non-alcoholic fatty liver disease. Effective therapy for NASH is still lacking. In this study, we investigated the effects of Ursodeoxycholic acid (UDCA) in the treatment of NASH.MethodsWestern and Chinese databases were searched by independent investigators using appropriate MESH headings to identify randomized, controlled Western and Chinese clinical trials, published between January 1990 and October 2012, testing the effects of UDCA in patients with NASH. Patient characteristics and trial endpoints were analyzed, with quality assessment according to widely acknowledged criteria. Pu2009<u20090.05 was defined as statistically significant in all trials.ResultsTwelve qualified randomized clinical trials, including six from China and involving 1160 subjects, were selected. Seven of these trials assessed the effects of UDCA Monotherapy, with the other five testing combinations of UDCA with vitamin E, polyene phosphatidylcholine, silymarin, glycyrrhizin and tiopronin. The duration of therapy ranged from 3 to 24xa0months, with two studies using high doses of UDCA (23–35xa0mg/kg/d). The average quality point was 2.69, and was significantly lower in articles from China than in those from Western countries (2.2u2009±u20090.4 vs. 3.8u2009±u20091.1, respectively, pu2009<u20090.05). UDCA Monotherapy significantly improved liver function in five studies and improved steatosis and fibrosis in two studies. All five studies assessing UDCA combination therapy showed significant improvements liver function, while two studies also improved steatosis and inflammation. One study of high-dose UDCA showed significant improvements in ALT, γGT and liver fibrosis, whereas the other study showed no significant change in ALT and liver pathology.ConclusionsUDCA therapy is effective in NASH, especially when combined with other drugs. However, the low quality of these studies and the heterogeneity of their results precluded further meta-analysis. Additional carefully designed clinical trials are needed, especially in China.

GDC-0941 sensitizes breast cancer to ABT-737 in vitro and in vivo through promoting the degradation of Mcl-1

The present study showed that GDC-0941 potently sensitized breast cancer to ABT-737 in vitro and in vivo. ABT-737 exhibited limited lethality in breast cancer cells; however, when combined with GDC-0941, it displayed strong synergistic cytotoxicity and enhanced caspase-mediated apoptosis. GDC-0941 promoted proteasomal degradation of Mcl-1, of which the overexpression has been validated to confer ABT-737 resistance, thereby enhanced the anticancer efficacy of ABT-737. Furthermore, the combination of GDC-0941 and ABT-737 exerted increased anti-tumor efficacy on MDA-MB-231 xenograft models. Overall, our data described unprecedentedly the promising therapeutic potential and underlying mechanisms of combining GDC-0941 with ABT-737 in treating breast cancer.

Transition from hepatic steatosis to steatohepatitis: Unique microRNA patterns and potential downstream functions and pathways

Background and Aim:u2002 This study aimed to explore the unique miRNA responsible for transition from hepatic steatosis to steatohepatitis and to investigate the functions and pathways of their downstream targets.

Upregulating Noxa by ER Stress, Celastrol Exerts Synergistic Anti-Cancer Activity in Combination with ABT-737 in Human Hepatocellular Carcinoma Cells

The human hepatocellular carcinoma (HCC) represents biologically aggressive and chemo-resistant cancers. Owing to the low affinity with the apoptotic factor Mcl-1, the BH3 mimetic drug ABT-737 failed to exert potent cancer-killing activities in variety of cancer models including HCC. The current study demonstrated that combining ABT-737 and Celastrol synergistically suppressed HCC cell proliferation, and induced apoptosis which was accompanied with the activation of caspase cascade and release of cytochrome c from mitochondria. Further study revealed that the enhanced Noxa caused by Celastrol was the key factor for the synergy, since small interfering RNA-mediated knockdown of Noxa expression in HCC cells resulted in decreased apoptosis and attenuated anti-proliferative effects of the combination. In addition, our study unraveled that, upon Celastrol exposure, the activation of endoplasmic reticulum (ER) stress, specifically, the eIF2α-ATF4 pathway played indispensable roles in the activation of Noxa, which was validated by the observation that depletion of ATF4 significantly abrogated the Noxa elevation by Celastrol. Our findings highlight a novel signaling pathway through which Celastrol increase Noxa expression, and suggest the potential use of ATF4-mediated regulation of Noxa as a promising strategy to improve the anti-cancer activities of ABT-737.

Simultaneous NF-κB inhibition and E-cadherin upregulation mediate mutually synergistic anticancer activity of celastrol and SAHA in vitro and in vivo.

Suberoylanilide hydroxamic acid (SAHA) is a promising histone deacetylase (HDAC) inhibitor approved by the US Food and Drug Administration (FDA) and whose clinical application for solid tumours is partially limited by decreased susceptibility in cancer cells due to nuclear factor (NF)‐κB activation. As an NF‐κB inhibitor, celastrol exhibits potent anticancer effects but has failed to enter clinical trials due to its toxicity. In this report, we demonstrated that the combination of celastrol and SAHA exerted substantial synergistic efficacy against human cancer cells in vitro and in vivo accompanied by enhanced caspase‐mediated apoptosis. This drug combination inhibited the activation of NF‐κB caused by SAHA monotherapy and consequently led to increased apoptosis in cancer cells. Interestingly, E‐cadherin was dramatically downregulated in celastrol‐resistant cancer cells, and E‐cadherin expression was closely related to decreased sensitivity to celastrol. However, our combination treatment significantly augmented the expression of E‐cadherin, suggesting that mutual mechanisms contributed to the synergistic anticancer activity. Furthermore, the enhanced anticancer efficacy of celastrol combined with SAHA was validated in a human lung cancer 95‐D xenograft model without increased toxicity. Taken together, our data demonstrated the synergistic anticancer effects of celastrol and SAHA due to their reciprocal sensitisation, which was simultaneously regulated by NF‐κB and E‐cadherin; thus, the combination of celastrol and SAHA was superior to other combination regimens that rely on a single mechanism. Our findings not only open new opportunities for the clinical development of SAHA but should also motivate the clinical investigation of celastrol, which has been hampered by its toxicity.

Nutlin-3 inhibits epithelial–mesenchymal transition by interfering with canonical transforming growth factor-β1-Smad-Snail/Slug axis

Enormous efforts have been made to explore small molecules that interfere with the TGF-β signaling pathway, so as to inhibit EMT and the cancer metastasis, but few agents have been identified. In this study, we demonstrated that Nutlin-3 could abolish the down-regulation of E-cadherin induced by TGF-β1 in p53-deficient cancer cells. Further studies revealed that Nutlin-3 prohibited EMT by blocking the phosphorylation of Smad2/3, resulting in the decreased transcription of Snail/Slug. In addition, Nutlin-3 suppressed the motility of cancer cells, and potentiated the anti-proliferative activity of gefitinib and lapatinib. Collectively, Nutlin-3 could inhibit EMT and enhance the anti-cancer activity of EGFR inhibitors by interfering with the canonical TGF-β1-Smad-Snail/Slug axis, in a p53-independent manner.

Identification of novel inhibitors of p53–MDM2 interaction facilitated by pharmacophore-based virtual screening combining molecular docking strategy

3D pharmacophore models for inhibitors of p53–MDM2 interaction were developed. The pharmacophore-based virtual screening of an in-house compound database combined with docking studies led to the identification of compound MCL0527 as a novel lead (MDM2 binding IC50 = 4.23 μM). Initial structure optimization yielded some derivatives with improved p53–MDM2 binding inhibitory activities. Furthermore, all target compounds showed potent anti-proliferative effect against human tumor cell lines with a generally selective profile on wild-type p53 cell lines.

MDM2 promotes epithelial|[ndash]|mesenchymal transition and metastasis of ovarian cancer SKOV3 cells

Background:Metastasis accounts for the most lethal reason for the death of ovarian cancer patients, but remains largely untreated. Epithelial–mesenchymal transition (EMT) is critical for the conversion of early-stage ovarian tumours into metastatic malignancies. Thus the exploration of the signalling pathways promoting EMT would open potential opportunities for the treatment of metastatic ovarian cancer. Herein, the putative role of MDM2 in regulating EMT and metastasis of ovarian cancer SKOV3 cells was investigated.Methods:The regulatory effects by MDM2 on cell motility was emulated by wound-healing and transwell assays. The effects on EMT transition and Smad pathway were studied by depicting the expression levels of epithelial marker E-cadherin as well as key components of Smad pathway. To evaluate the clinical relevance of our findings, the correlation of MDM2 expression levels with the stages of 104 ovarian cancer patients was investigated by immunohistochemistry assay.Results:We demonstrate that MDM2 functions as a key factor to drive EMT and motility of ovarian SKOV3 cells, by facilitating the activation of TGF-β-Smad pathway, which results in the increased transcription of snail/slug and the subsequent loss of E-cadherin levels. Such induction of EMT is sustained in either E3 ligase-depleted MDM2 or E3 ligase inhibitor HLI-373-treated cells, while being impaired by the N-terminal deletion of MDM2, which is also reflected by the inhibitory effects against EMT by Nutlin-3a, the N-terminal targeting agent. The expression levels of MDM2 is highly correlated with the stages of the ovarian cancer patients, and the higher expression of MDM2 together with TGFB are closely correlated with poor prognosis and predict a high risk of ovarian cancer patients.Conclusions:This study suggests that MDM2 activates Smad pathway to promote EMT in ovarian cancer metastasis, and targeting the N-terminal of MDM2 can reprogram EMT and impede the mobility of cancer cells.

Abstract 1434: MDM2 promotes tumor cell migration through the induction of epithelial to mesenchymal transition

Objectives: Epithelial to mesenchymal transition (EMT), which occurs in the initiation of metastasis, offers tumor cells increased invasive properties. MDM2 overexpression has been reported to predict distant metastasis, with mechanisms undefined. The present study will investigate the role of MDM2 in regulating EMT, so as to promote the discovery and development of novel targets for anti-metastasis therapy. Methods: (1) MDM2 expression in human ovarian adenocarcinoma tissues was examined by immunohistochemical staining; (2) SKOV3 cells were transfected with MDM2 plasmid, then the alteration of cell motility was detected using wound-healing and transwell assays; the protein and mRNA expression of E-cadherin and Slug/Snail were detected by Western Blot and real-time PCR respectively; (3) Smad2, Smad3 or Smad4 was silenced by siRNA, then E-cadherin protein expression affected by MDM2 overexpression was determined by Western Blot; (4) After transfection with MDM2 siRNA, TGF-β-triggered activation EMT and cell migration were observed; (5) The effect of MDM2 inhibitor Nutlin-3 on EMT and cell motility were observed by Western Blot and transwell assays. Results: MDM2 expression in 123 clinical samples was examined by immunohistochemical staining, which identified a significant correlation between MDM2 expression and clinical staging. The reduction of E-cadherin protein expression, as the indicator of the occurrence of EMT, was observed in SKOV3 cells transfected with MDM2 plasmid, which led to increased cell motility. In addition, evident upregulation of Slug and Snail expression was induced by MDM2 overexpression. Smad complexes directly activate the transcription of Slug and Snail. In our study, after silencing Smad2, Smad3 or Smad4, the inhibitory effect of MDM2 on E-cadherin expression was counteracted, indicating that Smad pathway might be critical for MDM2-promoted EMT. Then we found that TGF-β-Smad-induced EMT and cell migration could be suppressed by silencing MDM2. Further investigation suggested that the MDM2 inhibitor Nutlin-3 could prevent the downregulation of E-cadherin induced by TGF-β or MDM2 overexpression through interrupting Smad activation. Additionally, Nutlin-3 exhibited inhibitory effect on cell migration, which might be owing to its antagonism on the EMT. Conclusion: In summary, this study explored the molecular mechanisms underlying the role of MDM2 in cancer progression. MDM2 expression has been demonstrated to be closely correlated to human ovarian adenocarcinoma clinical staging, indicating the possible utilization of MDM2 in prognosis. Further study clarified MDM2 promoted tumor cell migration through the induction of EMT, which provides new theoretical evidences for the roles that MDM2 plays in tumor metastasis. Our study implicates that MDM2 may be a potential target for inhibiting metastasis and contributes to the translational research of MDM2 inhibitors as anti-metastatic agents. Citation Format: Lin Zheng, Yeping Wu, Tianyi Zhou, Hong Zhu, Qiaojun He, Bo Yang. MDM2 promotes tumor cell migration through the induction of epithelial to mesenchymal transition. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1434. doi:10.1158/1538-7445.AM2015-1434