Hepatocellular carcinoma (HCC) is a common and deadly cancer with limited treatment options. Through genome‐wide growth depletion screens using clustered regularly interspaced short palindromic repeats and expression profiling of primary HCC tumors, we identified 13 clinically relevant target genes with therapeutic potential. Subsequent functional annotation analysis revealed significant enrichment of these 13 genes in the cell cycle, cell death, and survival pathways. Non–structural maintenance of chromosomes condensin I complex subunit G (NCAPG) was ranked the highest among the depletion screens and multiple HCC expression datasets. Transient inhibition of NCAPG using specific small interfering RNAs resulted in a significant reduction in cell growth, migration, and the down‐regulation of mitochondrial gene expression in vitro. Small homologous RNA–mediated knockdown of NCAPG significantly impaired cell viability, caused aberrant mitotic division, fragmented the mitochondrial network, and increased cell death in vitro. HCC cells with a reduced expression of NCAPG formed significantly smaller xenograft tumors in vivo. Importantly, high NCAPG expression was significantly associated with poorer overall and disease‐free survival in HCC patients. High NCAPG expression is a novel prognostic biomarker to predict HCC early recurrence after surgical resection. In conclusion, NCAPG is an essential gene for HCC tumor cell survival. It represents a promising novel target for treating HCC and a prognostic biomarker for clinical management of HCC.—Wang, Y., Gao, B., Tan, P. Y., Handoko, Y. A., Sekar, K., Deivasigamani, A., Seshachalam, V. P., OuYang, H.‐Y., Shi, M., Xie, C., Goh, B. K. P., Ooi, L. L., Hui, K. M. Genome‐wide CRISPR knockout screens identify NCAPG as an essential oncogene for hepatocellular carcinoma tumor growth. FASEB J. 33, 8759–8770 (2019). http://www.fasebj.org
Early tumor recurrence after curative surgical resection poses a great challenge to the clinical management of hepatocellular carcinoma (HCC). We conducted whole genome expression microarrays on 64 primary HCC tumors with clinically defined recurrence status and cross‐referenced with RNA‐seq data from 18 HCC tumors in the Cancer Genome Atlas project. We identified a 77‐gene signature, which is significantly associated with early recurrent (ER) HCC tumors. This ER‐associated signature shows significant enrichment in genes involved in cell cycle pathway. We performed receiver operating characteristic (ROC) analysis to evaluate the prognostic biomarker potential of these 77 genes and Pearson correlation analysis to identify 11 close clusters. The one gene with the best area under the ROC curve in each of the 11 clusters was selected for validation using reverse‐transcription quantitative PCR in an independent cohort of 24 HCC tumors. NUF2 was identified to be the minimal biomarker sufficient to discriminate ER tumors from LR tumors. NUF2 in combination with liver cirrhosis could significantly improve the detection of ER tumors with an AUROC of 0.82 and 0.85 in the test and validation cohort, respectively. In conclusion, NUF2 in combination with liver cirrhosis is a promising prognostic biomarker for early HCC recurrence.
In our screening projects for anticancer agents from natural resources, artocarpin [6-(3-methyl-1-butenyl)-5,2',4'-trihydroxy-3-isoprenyl-7-methoxyflavone] isolated from wood of jack fruit (Artocarpus heterophyllus) showed potent cytotoxic activity on human T47D breast cancer cells. The mode of action of artocarpin was evaluated by its effect on cell viability, nuclear morphology, cell cycle progression, expression of protein markers for apoptosis, and mitochondrial membrane potential (Delta psi m). These results showed that artocarpin caused a reduction of cell viability in a concentration-dependent manner and an alteration of cell and nuclear morphology. Moreover, the percentage of the sub-G1 phase formation was elevated dose-dependently. Artocarpin induced activation of caspase 8 and 10 as indicated by stronger signal intensity of cleaved-caspase 8 and weaker signal intensity of caspase 10 markers detected after artocarpin treatment. In addition, we also noticed the activation of caspase 3 by artocarpin. There were negligible changes in mitochondrial membrane potential (Delta psi m) due to artocarpin treatment. All together, these data indicated that artocarpin induced apoptosis in T47D cells possibly via an extrinsic pathway.
Purpose: To evaluate the anti-cancer properties of the diethylether extract of Sukun (Artocarpus altilis) wood. Methods: The extract was tested in human T47D breast cancer cells and examined for its effect on cell viability, nuclear morphology and sub-G1 formation. Cell viability was determined by microculture tetrazolium technique (MTT), nuclear morphology investigated using 4'-6-diamidino-2-phenylindole (DAPI) staining technique, and cell cycle progression monitored by sub-G1 apoptosis assay using flow cytometry. Results: The results showed decreasing cell viability in a concentration-dependent manner. Altered cell morphology after treatment with the extract demonstrated that cells experienced apoptosis. Cell cycle analysis indicated that the number of cells in sub-G1 phase rose with increasing concentrations of the extract. Conclusion: The data demonstrate that Sukun wood extract induced apoptosis and sub-G1 phase formation in breast cancer (T47D) cells, and therefore, has a potential as an anti-cancer agent.
Purpose: To investigate the anti-cancer properties of the methanol extract of Piper crocatum Ruiz & Pav. leaves and its mode of action in human breast cancer (T47D) cells. Methods: The anti-cancer property and mechanism of action of the extract were evaluated by its effect on cell viability, nuclear morphology, cell cycle progression and the expression of phosphorylated p44/p42 as a marker for cell proliferation. Results: The results showed that there was a reduction of cell viability by the extract in a concentrationdependent manner and no alteration of nuclear morphology was observed. There were negligible changes in Sub-G1 phase formation after extract treatment. Expression of phosphorylated p44/42 was decreased by the extract only. Inclusion of the extract in the incubation medium decreased insulinstimulated phosphorylation of p44/p42 indicating that the anti-proliferative effect of the extract was via p44/p42 pathway. Conclusion: All together, the data indicate that P. crocatum methanol extract inhibits the growth of human breast cancer (T47D) cells via inhibition of p44/p42 phosphorylation.
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