Expression profiling of anticancer genes in colorectal cancer patients and their in vitro induction by riproximin, a ribosomal inactivating plant protein.

BACKGROUND: Ectopic expression of anticancer genes (ACGs) imposes antineoplastic effects on transformed cells. Clinically, reduced expression of these genes has been linked with poor prognosis, metastasis and chemo/radiotherapy resistance in cancers. Identifying expression pattern of ACGs is crucial to establish their prognostic and therapeutic relevance in colorectal cancer (CRC). In addition to the clinical perspective, naturally occurring compounds can be explored in parallel for inducing ACGs to achieve cancer cell-specific death. METHODOLOGY: Expression profiles of three ACGs (NOXA, PAR-4, TRAIL) were identified via real-time PCR in CRC clinical isolates. Time lapse-based expression modifications in ACGs were studied in a CRC liver metastasis animal model using microarray methodology. Effects of a purified plant protein (riproximin) on selected ACGs were identified in three primary and metastatic CRC cell lines by real-time PCR. Lastly, importance of the ACGs in a cellular environment was highlighted via bioinformatic analysis. RESULTS: ACGs (except NOXA) were persistently downregulated in clinical isolates when comparing the overall mean expression values with normal mucosa levels. In vivo studies showed a prominent inhibition of NOXA and PAR-4 genes in implanted CRC cells during rat liver colonization. TRAIL showed deviation from this theme while showing marked induction during the early period of liver colonization (days 3 and 6 after CRC cell implantation). Riproximin exhibited substantial potential of inducing ACGs at transcriptome levels in selected CRC cell lines. Bioinformatic analysis showed that vital molecular/functional aspects of a cell are associated with the presence of ACGs. CONCLUSION: ACGs are downregulated in primary and metastatic phase of CRC. Riproximin effectively induces ACGs in CRC cells and can be exploited for clinical investigations over time.

Modulation of the Endothelin System in Colorectal Cancer Liver Metastasis: Influence of Epigenetic Mechanisms?

Targeting of endothelin system genes is a promising strategy in cancer therapy. The modulation of these genes was explored in a model of colorectal cancer (CRC) liver metastasis and in a panel of CRC tumor cell lines that were exposed to the demethylating agent decitabine. The CC531 rat model mimicking CRC liver metastasis was used for tumor cell re-isolation and analysis of the endothelin system genes and DNA methyltransferases (DNMTs) by microarray. To mimic the effects caused by methylation changes, a panel of seven CRC cell lines was treated with the demethylating agent decitabine. Three genes of the endothelin system were potently modulated at messenger RNA (mRNA) level in rat CC531 cells during liver colonization. The concomitant decrease of two DNMTs suggested an influence from altered methylation. Changes in gene expression were also accomplished by exposure of CRC cells to the demethylating agent decitabine, when using daily low concentrations for 3 days, with minimal cytotoxic effects. Sensitive human SW480 cells showed an almost 100fold upregulation of endothelin-1 mRNA compared to untreated cells. This, however, was different in LS174T cells, which showed no significant increase in gene expression although the methylation levels were significantly decreased at a variety of corresponding loci. We suggest that the mechanism induced by methylation on gene expression in metastatic CRC cells can be compromised. The results question the overall success of treating metastatic CRC by methylation inhibitors.

The CCR5 antagonist maraviroc causes remission of pancreatic cancer liver metastasis in nude rats based on cell cycle inhibition and apoptosis induction.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease, and novel therapeutic strategies are urgently needed. Recently, expression of the C-C chemokine receptor 5 (CCR5) and its ligands has been found to play an important role in cancer progression and metastasis. In this study, we blocked the CCR5 receptor by the FDA approved antagonist maraviroc (MVC) in Suit2-007 and MIA-PaCa-2 human PDAC cells. The treatment significantly inhibited their proliferation and induced apoptosis of exposed cells as evidenced by caspases activation and increased Bax levels. Moreover, MVC inhibited the cell cycle by down regulating the proteins of the complexes of cyclin dependent kinase (CDK) 4/6 - Cyclin D and CDK2 - Cyclin E, as well as by increasing the protein levels of CDK inhibitors p18, p21 and p27. In line with this, MVC caused significant retardation of Suit2-007 cells growing in a PDAC liver metastasis xenograft model (p < 0.05). These results suggest that maraviroc could be a promising treatment strategy for PDAC patients with liver metastases.

The insulin-like growth factor binding proteins 3 and 7 are associated with colorectal cancer and liver metastasis.

Over the last few decades, a great deal of attention has been directed to the IGF system for its vital role in regulating cell and tissue survival, growth and differentiation. The insulin-like growth factor binding proteins (IGFBPs), a main constituent of this system, have been implicated in the tumorigenesis of colorectal cancer (CRC). In this study, we intended to shed more light on two essential members; IGFBP3 as representative for the six main IGFBPs and IGFBP7 to represent their related proteins (IGFBP-rps). Our experiments on silencing IGFBP3 or IGFBP7 in the two human CRC cell lines SW480, Caco2, and in the rat CRC cell line CC531 show reduced proliferation, colony formation, and for IGFBP3, also reduced migration. The expression of both genes in 68 human CRC samples was higher in UICC stages II and III than in stages I and IV. Additionally, IGFBP3 was negatively correlated with age (p = 0.05) and positively related to IGFBP7 expression (p = 0.0001). Further, in a liver metastasis experiment, the expression of both genes was drastically increased in response to early metastatic growth in vivo. Since these high levels returned gradually to normal thereafter, it could be assumed that the up-regulation of IGFBPs is vital during the process of homing into the liver and early metastatic dissemination. Our results indicate that IGFBP3 and 7 cannot be simply considered as tumor suppressors but have additional properties, which become evident only during cancer progression and metastasis formation.