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HIF-1alpha determines the metastatic potential of gastric cancer cells.

ABSTRACT: Gastric adenocarcinoma is characterised by rapid emergence of systemic metastases, resulting in poor prognosis due to vanished curative treatment options. Better understanding of the molecular basis of gastric cancer spread is needed to design innovative treatments. The transcription factor HIF-1alpha (hypoxia-inducible factor 1alpha) is frequently overexpressed in human gastric cancer, and inhibition of HIF-1alpha has proven antitumour efficacy in rodent models, whereas the relevance of HIF-1alpha for the metastatic phenotype of gastric adenocarcinoma remains elusive. Therefore, we have conducted a comprehensive analysis of the role of HIF-1alpha for pivotal metastasis-associated processes of human gastric cancer. Immunhistochemistry for HIF-1alpha showed specific staining at the invading tumour edge in 90% of human gastric cancer samples, whereas normal gastric tissue was negative and only a minority of early gastric cancers (T1 tumours) showed specific staining. Hypoxia-inducible factor 1alpha-deficient cells showed a significant reduction of migratory, invasive and adhesive properties in vitro. Furthermore, the HIF-1alpha-inhibitor 2-methoxy-estradiol significantly reduced metastatic properties of gastric cancer cells. The accentuated expression at the invading edge together with the in vitro requirement of HIF-1alpha for migration, invasion and adherence argues for a pivotal role of HIF-1alpha in local invasion and, ultimately, systemic tumour spread. These results warrant the exploration of HIF-1alpha-inhibiting substances in clinical treatment studies of advanced gastric cancer.

SUBMITTER: Rohwer N

PROVIDER: S-EPMC2653758 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND: Hypoxia-inducible factor 1 (HIF-1) is a master transcriptional regulator of genes regulating oxygen homeostasis. The HIF-1 protein is composed of two HIF-1alpha and HIF-1beta/aryl hydrocarbon receptor nuclear translocator (ARNT) subunits. The prognostic relevance of HIF-1alpha protein overexpression has been shown in breast cancer. The impact of HIF-1alpha alternative splice variant expression on breast cancer prognosis in terms of metastasis risk is not well known. METHODS: Using real-time quantitative reverse transcription PCR assays, we measured mRNA concentrations of total HIF-1alpha and 4 variants in breast tissue specimens in a series of 29 normal tissues or benign lesions (normal/benign) and 53 primary carcinomas. In breast cancers HIF-1alpha splice variant levels were compared to clinicopathological parameters including tumour microvessel density and metastasis-free survival. RESULTS: HIF-1alpha isoforms containing a three base pairs TAG insertion between exon 1 and exon 2 (designated HIF-1alphaTAG) and HIF-1alpha736 mRNAs were found expressed at higher levels in oestrogen receptor (OR)-negative carcinomas compared to normal/benign tissues (P = 0.009 and P = 0.004 respectively). In breast carcinoma specimens, lymph node status was significantly associated with HIF-1alphaTAG mRNA levels (P = 0.037). Significant statistical association was found between tumour grade and HIF-1alphaTAG (P = 0.048), and total HIF-1alpha (P = 0.048) mRNA levels. HIF-1alphaTAG mRNA levels were also inversely correlated with both oestrogen and progesterone receptor status (P = 0.005 and P = 0.033 respectively). Univariate analysis showed that high HIF-1alphaTAG mRNA levels correlated with shortened metastasis free survival (P = 0.01). CONCLUSIONS: Our results show for the first time that mRNA expression of a HIF-1alphaTAG splice variant reflects a stage of breast cancer progression and is associated with a worse prognosis.See commentary: http://www.biomedcentral.com/1741-7015/8/45.

Project description:BackgroundReduced chemosensitivity of solid cancer cells represents a pivotal obstacle in clinical oncology. Hence, the molecular characterization of pathways regulating chemosensitivity is a central prerequisite to improve cancer therapy. The hypoxia-inducible factor HIF-1alpha has been linked to chemosensitivity while the underlying molecular mechanisms remain largely elusive. Therefore, we comprehensively analysed HIF-1alpha's role in determining chemosensitivity focussing on responsible molecular pathways.Methodology and principal findingsRNA interference was applied to inactivate HIF-1alpha or p53 in the human gastric cancer cell lines AGS and MKN28. The chemotherapeutic agents 5-fluorouracil and cisplatin were used and chemosensitivity was assessed by cell proliferation assays as well as determination of cell cycle distribution and apoptosis. Expression of p53 and p53 target proteins was analyzed by western blot. NF-kappaB activity was characterized by means of electrophoretic mobility shift assay. Inactivation of HIF-1alpha in gastric cancer cells resulted in robust elevation of chemosensitivity. Accordingly, HIF-1alpha-competent cells displayed a significant reduction of chemotherapy-induced senescence and apoptosis. Remarkably, this phenotype was completely absent in p53 mutant cells while inactivation of p53 per se did not affect chemosensitivity. HIF-1alpha markedly suppressed chemotherapy-induced activation of p53 and p21 as well as the retinoblastoma protein, eventually resulting in cell cycle arrest. Reduced formation of reactive oxygen species in HIF-1alpha-competent cells was identified as the molecular mechanism of HIF-1alpha-mediated inhibition of p53. Furthermore, loss of HIF-1alpha abrogated, in a p53-dependent manner, chemotherapy-induced DNA-binding of NF-kappaB and expression of anti-apoptotic NF-kappaB target genes. Accordingly, reconstitution of the NF-kappaB subunit p65 reversed the increased chemosensitivity of HIF-1alpha-deficient cells.Conclusion and significanceIn summary, we identified HIF-1alpha as a potent regulator of p53 and NF-kappaB activity under conditions of genotoxic stress. We conclude that p53 mutations in human tumors hold the potential to confound the efficacy of HIF-1-inhibitors in cancer therapy.

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HIF-1alpha determines the metastatic potential of gastric cancer cells.

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