The role of HNF4A in GATA4-deficiency phenotypes of hepatocellular carcinoma.

2018 ◽  
Vol 36 (4_suppl) ◽  
pp. 284-284
Author(s):  
Yu Bin Tan ◽  
Timothy Shuen ◽  
Han Chong Toh
Keyword(s):  
Gene Expression ◽  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Correlation Analysis ◽  
Cell Cycle Arrest ◽  
Cell Line ◽  
Transgenic Mouse ◽  
Cell Lines ◽  
Downstream Target ◽  
Cycle Arrest

284 Background: Hepatocellular carcinoma (HCC) is the 2nd leading global cause of cancer death. Recently, we have discovered previously undescribed deletion and germline mutation of GATA4 and showed that GATA4 is a key differentiation driver and metabolic regulator in HCC. However, as GATA4 is mostly deleted in HCC, targeting GATA4-downstream molecules is ideal. In this study, proof-of-concept experiments were conducted to show that introduction of HNF4A, which is a GATA4-regulated downstream target, could partially rescue the impaired phenotypes in GATA4-deficient HCC cell line. Methods: Correlation analysis using gene expression microarray of human HCC samples was conducted to identify the genes that are positively correlated with GATA4. A transgenic mouse model with a liver-specific conditional GATA4 knockout was designed to identify liver morphology and gene expression changes which are correlated with the loss of Gata4 in the mouse liver. CRISPR-mediated knockout of GATA4 and lentiviral HNF4A overexpression was carried out in a GATA4-deficient HCC cell lines, PLC/PRF/5 and Hep3B, followed by proliferation, apoptosis, cell cycle and senescence functional assays. Results: Pearson correlation analysis from human HCC samples showed that expression of HNF4A is positively correlated with that of GATA4. Livers from conditional Gata4 knockout mice had lower Hnf4a gene expression when compared to age-matched control mice. Loss of function analysis by CRISPR-mediated GATA4 knockout further showed downregulation of HNF4A in GATA4-deficient PLC/PRF/5 cell line. Lentiviral HNF4A overexpression in PLC/PRF/5 and Hep3B demonstrated reduced proliferation and increased apoptosis while PLC/PRF/5 also showed cell cycle arrest at G2/M phase when compared to control. However, no senescence induction was detected in HNF4A-overexpressing cells. Conclusions: Transgenic mouse data, CRISPR-mediated knockout and analysis of HCC samples showed that HNF4A is a key GATA4-downstream target. HNF4A overexpression decreases proliferation, increases apoptosis and cell cycle arrest in GATA4-deficient HCC cell lines, thus representing a possible therapeutic target for HCC.

scholarly journals Coptidis Rhizome inhibits cell growth in HONE-1 cell line by inducing cell cycle arrest and apoptosis

2020 ◽  
Author(s):  
Kim Fey Leu ◽  
Menaga Subramaniam ◽  
Xinghua Wang ◽  
Zao Yang ◽  
Lee Fah Yap ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Line ◽  
Cell Lines ◽  
Cancer Cell ◽  
Cytotoxic Effect ◽  
Global Gene Expression ◽  
Chinese Herbal ◽  
Cycle Arrest

Abstract Background Nasopharyngeal carcinoma (NPC) is among the most common head and neck malignancies seen among adults in Malaysia. Therefore, discovery of novel anti-cancer herbal drugs is of importance. In this study, the cytotoxic effect was conducted on a traditional Chinese herbal prescription (Xiao Xian Xiong Decoction (XXXD) that is made up of 3 Chinese herbal medicines, namely Huanglian (Coptidis Rhizome), Banxia (Pinellia Rhizome), Gualuo (Fructus Trichosanthis).Methods The cytotoxic effect of the individual herb and in combination of two and three herbs was studied on 8 nasopharyngeal cancer cell lines. Global gene expression analysis was carried on extracted RNA using nCounter XT Gene Expression Assay.Results TWO-1, TWO-4, HONE-1, SUNE-1, CNE-2, HK-1, CNE-1 and C666-1 treated with Huanglian, the IC50 values obtained were 24.48, 11.77, 4.48, 10.72 6.32, 11.10, 6.77 and 27.30 µg/ml, respectively. For combination of Huanglian and Banxia, the IC50 values obtained were 74.09 µg/ml (TWO-1), 25.80 µg/ml (TWO-4), 38.10 µg/ml (HONE-1), 29.46 µg/ml (SUNE-1), 19.0 µg/ml (CNE-2) and 20.12 µg/ml (HK-1) but did not exert 50% cell killing in CNE-1 and C666-1 cell lines. The IC50 value attained for the combination of Huanglian and Gualuo was 40.70 µg/ml in HONE-1 cell line. The IC50 values obtained for XXXD (triple combination of Huanglian, Banxia and Gualuo)-treated in HONE-1 and CNE-2 cell lines were 88.55 and 92.42 µg/ml, respectively. Out of all these 7 groups of herbal samples, Huanglian showed the highest cytotoxicity against 8 NPC cell lines with the lowest IC50 value of 4.48 µg/ml recorded in HONE-1. Global gene expression showed Huanglian significantly downregulated genes associated with cell cycle arrest and apoptosis, and thus inhibit HONE-1 cell growth.Conclusions This study suggest that Huanglian could be a potent anticancer herb targeting HONE-1 cancer cell line.

Anticancer Effect of Amygdalin (Vitamin B-17) on Hepatocellular Carcinoma Cell Line (HepG2) in the Presence and Absence of Zinc

2020 ◽  
Vol 20 (4) ◽  
pp. 486-494
Author(s):  
Mohamed A. El-Desouky ◽  
Abdelgawad A. Fahmi ◽  
Ibrahim Y. Abdelkader ◽  
Karima M. Nasraldin
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Cytochrome C ◽  
Cell Cycle Arrest ◽  
Cell Line ◽  
Hepg2 Cell ◽  
Caspase 3 ◽  
Vitamin B ◽  
Cycle Arrest ◽  
Hepg2 Cell Lines

Background: Amygdalin (Vitamin B-17) is a naturally occurring vitamin found in the seeds of the fruits of Prunus Rosacea family including apricot, bitter almond, cherry, and peach. Objective: The purpose of this study was to examine the effect of amygdalin with and without zinc on hepatocellular carcinoma (HepG2) cell line. Methods: MTT assay was used to evaluate the cytotoxicity of amygdalin without zinc, amygdalin + 20μmol zinc, and amygdalin + 800μmol zinc on HepG2 cell lines. The cell cycle distribution assay was determined by flow cytometry. Apoptosis was confirmed by Annexin V-FITC/PI staining assay. Moreover, the pathway of apoptosis was determined by the percentage of change in the mean levels of P53, Bcl2, Bax, cytochrome c, and caspase-3. Results: Amygdalin without zinc showed strong anti-HepG2 activity. Furthermore, HepG2 cell lines treatment with amygdalin + 20μmol zinc and amygdalin + 800μmol zinc showed a highly significant apoptotic effect than the effect of amygdalin without zinc. Amygdalin treatment induced cell cycle arrest at G2/M and increased the levels of P53, Bax, cytochrome c, and caspase-3 significantly, while it decreased the level of anti-apoptotic Bcl2. Conclusion: Amygdalin is a natural anti-cancer agent, which can be used for the treatment of hepatocellular carcinoma. It promotes apoptosis via the intrinsic cell death pathway (the mitochondria-initiated pathway) and cell cycle arrest at G/M. The potency of amygdalin in HepG2 treatment increased significantly by the addition of zinc.

Wogonoside Exerts Anti-Leukemia of AML Cells Via Restricting Phospholipid Scramblase 1 Gene Expression and Promoting Its Translocation Into Nuclear

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4282-4282
Author(s):  
Yan Chen ◽  
Bao-An Chen ◽  
Qing-long Guo
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Lines ◽  
Underlying Mechanism ◽  
Phase Cell ◽  
Western Blots ◽  
Phospholipid Scramblase ◽  
Cycle Arrest ◽  
Phospholipid Scramblase 1

Abstract Abstract 4282 Objective: To evaluate the antileukemic effect of wogonoside and reveal the underlying mechanism. Method: In this study trypan blue dye exclusion assay, MTT assay, and soft agar colony formation assay were used to analysis growth inhibition of wogonoside the on AML (acute human promyelocytic) cell lines. Propidium iodide (PI)-staining and cell cycle-regulatory proteins detecting by western blots were applied to exam whether wogonoside could induce cell cycle arrest. Then a series of experiment were used to assess the ability of wogonoside to overcome the AML associated differentiation block, by using Giemsa staining, Nitroblue tetrazolium (NBT) reduction assay, and cell-surface differentiation antigens expression analysis. Real time PCR, western blots, cycloheximide inhibition test and RNA interference, nuclear and cytoplasmic fractionation, immunofluorescent staining were used to investigate the underlying mechanism. In this point we mainly focus that wogonoside exerts antileukemic by modulating of PLSCR1 gene expression, as well as influence its subcellular localization to play a role in regulating gene transcription. Result: It was demonstrated that wogonoside have the capacity to decrease the growth of myeloid cell lines by induction of G0/1 phase cell cycle arrest and differentiation. This effect is mediated by the increasing in mRNA and up-regulating protein expression of phospholipids scramblase 1 (PLSCR1). Meanwhile wogonoside promoted PLSCR1 traffic into the nucleus, which let PLSCR1 to play a role in regulating cell cycle and differentiation-related genes transcription including p21, p27, c-myc and IP3R1. Conclusion: Wogonoside induced AML cell lines to undergo differentiation and G1 phase arrest by restricting phospholipid scramblase 1 gene expression and promoting its translocation into nuclear. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Melatonin enhances the anti-tumor effect of sorafenib via AKT/p27-mediated cell cycle arrest in hepatocarcinoma cell lines

RSC Advances ◽  
2017 ◽  
Vol 7 (34) ◽  
pp. 21342-21351 ◽  
Author(s):  
Fei Long ◽  
Chengyong Dong ◽  
Keqiu Jiang ◽  
Yakun Xu ◽  
Xinming Chi ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Lines ◽  
Cycle Arrest ◽  
Hepatocarcinoma Cell ◽  
Proposed Model ◽  
Hcc Cells

Proposed model elucidating the role of MT in regulating the proliferation of hepatocellular carcinoma (HCC) cells treated with sorafenib.

scholarly journals Bortezomib Is Effective in Treating T-ALL, Inducting G2/M Cell Cycle Arrest and WEE1 Downregulation

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4360-4360
Author(s):  
SIN Chun-fung ◽  
Timothy Ming-hun Wan ◽  
Aarmann Anil Mohinani Mohan ◽  
Yinxia Qiu ◽  
Anan Jiao
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Line ◽  
Cell Lines ◽  
Mechanism Of Action ◽  
Therapeutic Efficacy ◽  
Cyclin B1 ◽  
M Cell ◽  
Cycle Arrest ◽  
Cell Cycle Modulation

Abstract T lymphoblastic leukaemia (T-ALL) is an aggressive haematological malignancy with poor outcome, especially for relapse/refractory disease. Early T- cell precursor acute lymphoblastic leukaemia (ETP-ALL) is a recently identified subtype of T-ALL with worse treatment outcome compared with other subtypes of T-ALL and treatment options are limited. T-ALL frequently harbors genetic aberrations leading to cell cycle dysregulation and it is one of the major molecular pathogenesis of T-ALL. WEE1 is a protein kinase that is responsible for inhibiting mitosis with unrepaired damaged DNA via inactivating CDK1. WEE1 is highly express in adult T-ALL and its overexpression is associated with adverse prognosis in various cancers. Inhibiting WEE1 expression is a novel approach of therapy. Bortezomib is a 26S proteosome inhibitor and it is FDA approved for treating plasma cell myeloma and mantle cell lymphoma. Bortezomib had been demonstrated therapeutic efficacy in clinical setting for relapse/refractory paediatric T-ALL and B-ALL when combined with chemotherapy. Despite its therapeutic efficacy in clinical studies, the mechanism of action of Bortezomib in T-ALL remain uncertain. The role of Bortezomib in cell cycle modulation had not been established in T-ALL. Moreover, it had not been demonstrated that the effect of Bortezomib in WEE1 expression in T-ALL. Here, we present our study that demonstrated the therapeutic efficacy of Bortezomib in treating T-ALL via cell cycle modulation and downregulation of WEE1 by Bortezomib. T-ALL cell lines including MOLT16, MOLT4, LOUCY and CEM were used in the study. Cell viability was measured by trypan blue. Apoptosis and cell cycle analysis were measured by flow cytometry. Western blot of WEE1, p53, cyclin B1, p21 and p27 were performed. Our result showed that Bortezomib reduce the cell viability of T-ALL cell lines in dose and time-dependent manner. Bortezomib was also sensitive towards LOUCY, a T-ALL cell line with ETP-ALL phenotype. It implied that Bortezomib could be a promising therapy for ETP-ALL. Bortezomib also triggered apoptosis in various T-ALL and the effect of apoptosis was more pronounced after 72 hours of treatment when compared with 24-hour. Again, Bortezomib was able to induce apoptosis in LOUCY cell line. G2/M cell cycle arrest was observed in various T-ALL upon treatment of Bortezomib. The effect on cell cycle modulation was also observed in LOUCY cell line. The protein expression of p21 and p27 were increased after the treatment of Bortezomib. The level of cyclin B1 was increased also. There was upregulation of p53 after Bortezomib treatment. Strikingly, the protein expression level of WEE1 was reduced. The findings of WEE1 downregulation by Bortezomib is a novel findings. We also showed that Bortezomib downregulate WEE1 mRNA expression by quantitative PCR. Our study showed that Bortezomib is active against T-ALL cell lines, including ETP-ALL cell line, LOUCY and modulates cell cycle with G2/M arrest. Bortezomib had been shown to increase the level of p21, p27 and cyclin B1 and induced G2/M cell cycle arrest in glioblastoma cells. However, studies on cell cycle modulation by Bortezomib in T-ALL are scarce. Here, we demonstrated Bortezomib stabilized p21, p27 and upregulation of cyclin B1 in T-ALL as well, which could account for the G2/M cell cycle arrest. We first showed that downregulation of WEE1 after treatment with Bortezomib, in protein level as well as in mRNA level. Recent study showed that inhibition of WEE1 is a novel target of therapy in T-ALL. WEE1 is upregulated in T-ALL to prevent entry of mitosis with unrepaired damaged DNA. The downregulation of WEE1 by Bortezomib as showed by our study could reverse its effect and leads to apoptosis of leukaemic cells. In summary, our study provides the insight on mechanism of action of Bortezomib in modulating cell cycle in T-ALL. Moreover, it is the first study to demonstrate WEE1 downregulation by Bortezomib in T-ALL. These findings not only enhance our understanding of mechanism of action of Bortezomib in T-ALL, but also rationalized the use of certain synergistics combination therapy with Bortezomib in treating T-ALL, e.g., chemotherapeutic agents, PARP inhibitors which could damage DNA of leukaemic cells. Further research is needed to explore those combination therapy in T-ALL and molecular mechanism of downregulation of WEE1 by Bortezomib in T-ALL. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

PI3KCA and PIM Inhibitors in Peripheral T-Cell Lymphomas

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4917-4917
Author(s):  
Esperanza Martin-Sanchez ◽  
Socorro M. Rodriguez-Pinilla ◽  
Luis Lombardia ◽  
Margarita Sanchez-Beato ◽  
Beatriz Dominguez-Gonzalez ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
T Cell ◽  
Cell Cycle Arrest ◽  
Cell Lines ◽  
Gene Expression Signature ◽  
Dependent Manner ◽  
Expression Signature ◽  
Cycle Arrest ◽  
T Cell Lymphomas

Abstract Abstract 4917 T-cell lymphomas (TCL) are a heterogeneous group of aggressive malignancies lacking specific and efficient therapy. Unfortunately, there are neither animal models nor representative cell lines for most TCL types, making functional and pharmacogenomics studies even more difficult. PI3K and PIM are kinases involved in cell proliferation, frequently altered in human cancer that seems to play a critical role in T-cell development and activation. Genomic studies have identified PIK3CD subunit to be significantly associated with in activation of CD40, NF-kB and TCR-pathways. The aim of this project is to determine the efficiency of PI3K inhibitors (PI3Ki) and PIM inhibitors (PIMi) in TCL, looking for biomarkers of their mechanism of action and to identify markers that could identify responders from non-responders. Twenty PTCL and seven reactive lymph nodes were studied using gene expression microarrays. We performed an in silico analysis using the Connectivity Map program to identify drugs that could potentially reverse PTCL gene expression signature. Among them, several PI3K/mTOR inhibitors were found. A panel of 6 TCL cell lines belonging to different TCL subgroups were treated with 3 PI3Ki (LY294002, ETP-45658, GDC-0941) and one PIMi (ETP-39010). Functional studies were also done to establish the role of each of the targeted genes. In vitro studies showed that PI3Ki induced G1 cell cycle arrest in all cell lines, and apoptosis in a portion of them, in a time/dose-dependent manner. We also observed a decrease in the levels of pAKT(S473), pGSK3B(S9) and p-p70S6K(T389) after treatment. In addition, both the analysis of the PTCL gene expression signature as well as western blot studies on TCL cell lines has shown overexpression of PIM family genes, A decrease in cell viability, and a strong induction of apoptosis in all cell lines was seen after PIM inhibition, without cell cycle arrest. Several diagnostic and pharmacodynamic biomarkers of PIMi have been identified at the mRNA and protein level in both cell lines In conclusion, our results indicate that PI3Ki and PIMi are effective therapeutic approaches for TCLs, identifying potential markers for patient's stratification and pharmacodynamic assessment. Disclosures: No relevant conflicts of interest to declare.

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