Clinical significance of quantitative analysis of carcinoembryonic antigen assessed by flow cytometry in fresh human gastric cancer cells

Background Gastric cancer (GC) is a multifactorial disease with high mortality. Anti-HER2 therapy is a promising strategy in GC treatment and trastuzumab was approved by FDA (Food and Drug Administration) as the first and the second line of treatment of the disease. Purpose The aim of the study was to examine the effectiveness of a combination of etoposide with trastuzumab or pertuzumab in AGS gastric cancer cells and breast cancer cells such as MCF-7, MDA-MB-231 and HCC1954. Methods and findings The cytotoxic effects of the tested compounds against gastric and breast cancer cells were checked by MTT (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide) assay. The anti-proliferative potential was analyzed by the incorporation of [3H]-thymidine into DNA. Fluorescent microscopy and flow cytometry was used to demonstrate the effect of the compounds on apoptosis. The mitochondrial membrane potential, and the activity of caspase-8 and caspase-9 were assessed. Autophagosomes and autolysosomes formation was checked by flow cytometry. The concentrations of Beclin-1, LC3A and LC3B were performed using ELISA. The expression of LC3A/B was also determined. The results from our study proved that the combination of etoposide with anti-HER2 antibodies was not cytotoxic against breast cancer cells, whereas the combination of etoposide with anti-HER2 antibodies decreased viability and DNA biosynthesis in gastric cancer cells. The interaction of etoposide with pertuzumab or trastuzumab induced programmed cell death via extrinsic and intrinsic apoptotic pathways in AGS gastric cancer cells, but did not affect autophagy, where a decrease of Beclin-1, LC3A and LC3B was observed in comparison with the untreated control. Conclusions The study demonstrated that etoposide (12.5 μM) with pertuzumab represent a promising strategy in gastric cancer treatment, but further in vivo examinations are also required.

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Chemokine receptor CXCR4 expression and its regulation in gastric cancer cells

Journal of Clinical Oncology ◽

10.1200/jco.2009.27.15_suppl.e22173 ◽

2009 ◽

Vol 27 (15_suppl) ◽

pp. e22173-e22173

Author(s):

H. Lee ◽

H. Kim ◽

S. Kim ◽

H. Yun ◽

S. Kim ◽

...

Keyword(s):

Gastric Cancer ◽

Flow Cytometry ◽

Steady State ◽

Cancer Cells ◽

Chemokine Receptor ◽

Cxcr4 Expression ◽

Human Gastric Cancer ◽

Rt Pcr ◽

Gastric Cancer Cells ◽

Membrane Expression

e22173 Background: The chemokine receptor CXCR4 is associated with the biological behavior in several kinds of cancer, but few studies have addressed the expression and regulation of CXCR4 in gastric cancer. Methods: Five gastric cancer cell lines were studied. The expression of CXCR4 was investigated using RT-PCR, Westerning blotting, flow cytometry, and immunofluorescence staining. The regulation of CXCR4 expression by hypoxia, dexamethasone, and the proinflammatory cytokines was evaluated. Results: CXCR4 mRNA and proteins were detectable by RT-PCR and Western blot analysis, respectively, in all five cell lines. However, MKN-28, MKN-45, MKN-74, and SNU 16 cells did not express membrane CXCR4, but had abundant CXCR4 in their cytoplasm, as determined by flow cytometry and immunofluorescence staining. In contrast, a small population of KATO III cells expressed membrane CXCR4. Hypoxia up-regulated CXCR4 proteins and enhanced membrane expression of CXCR4 in human gastric cancer KATO III cells, which constitutively expressed membrane CXCR4 in a steady state, as revealed by Western blotting and flow cytometry, respectively. This hypoxia-induced expression of CXCR4 was mediated via hypoxia-inducible factor (HIF)-1α. In addition, KATO III cells exposed to hypoxia demonstrated enhanced migration in response to stromal cell-derived factor (SDF)-1α, a specific ligand for CXCR4. However, MKN-28, MKN-45, MKN-74, and SNU-16 cells, which lack membrane CXCR4 in a steady state, showed no change in CXCR4 expression in hypoxic condition. Treatment with IFN-γ, TGF-β, TNF-α, and dexamethasone did not induce any change in CXCR4 expression in all five gastric cancer cells. Conclusions: This study suggests that hypoxia up-regulates the membrane expression of functional CXCR4 via HIF-1α in human gastric cancer cells that basally express membrane CXCR4 in a steady state in vitro. No significant financial relationships to disclose.

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