Abstract A76: Characterization of CXC-chemokine expression in colorectal cancer tissue and its role in mediating chemoresistance of colorectal cancer cells to conventional and molecularly targeted therapies (2009)

Abstract Aim of study: The aims of this work were (i) to characterize CXC-chemokine and receptor expression in colorectal cancer (CRC) and (ii) study the role of CXC-chemokine signalling in modulating CRC cell sensitivity to Oxaliplatin and anti-EGFR therapy. Methods and Materials: CXC-chemokine and receptor expression was analysed in a retrospective study of colorectal cancer biopsy specimens in stage II and stage III CRC patients (n=254) who took part in a randomized phase III study comparing adjuvant 5FU chemotherapy with surgery alone between 1994 and 1997. For the in vitro work, studies were carried out in a panel of CRC cell lines comprising HCT116 clonal derivatives and LoVo cells. CXCL8 ligand and receptor expression was assessed by flow cytometry, qRT-PCR, western blotting and ELISA. Anti-apoptotic and EGFR signalling was also assessed using qRT-PCR and western blotting. NF-κB activity was assessed by electrophoretic mobility shift assay (EMSA) and Oxaliplatin response was assessed using cell counts. Results: Expression of CXCR1, CXCR2 and CXCL1 was elevated in tumor epithelium relative to the histologically normal adjacent colorectal cells (p&amp;lt;0.001, Wilcoxon signed ranks test). In addition, expression of CXCL1 correlated with the expression of its receptor CXCR2 in the tumor samples. In the CRC cell lines, constitutive expression of the ligands CXCL8 and CXCL1 was detected by immunoblotting and ELISA. Flow cytometry also confirmed the expression of the two receptors, CXCR1 and CXCR2 in these cell lines. The expression of the ligands and receptors was enhanced in response to Oxaliplatin treatment, demonstrated by qRT-PCR and immunoblotting. Furthermore, the administration of a CXCR2 antagonist AZ10397767 sensitized Oxaliplatin-promoted cytotoxicity and apoptosis in the PTEN-deficient HCT116 cells. Therefore, inhibition of CXC-chemokine signaling may modulate Oxaliplatin therapeutic response in PTEN-deficient CRC. Consistent with our published observations in prostate cancer cell lines, CXCL8 signalling increased NF-κB transcriptional activity and potentiated anti-apoptotic protein expression in CRC cells. Further studies demonstrated that CXCL8 signaling affects other survival pathways including activation of the Epidermal Growth Factor Receptor (EGFR). Immunoblotting confirmed that CXCL8 administration promotes post-translational regulation of EGFR through receptor phosphorylation in the CRC cells. Also, qRT-PCR experiments revealed that CXCL8 signaling potentiates the transcription of the EGFR ligands, Amphiregulin, and Transforming Growth factor-α (TGF-α) in CRC cell lines while ELISA detected an increased secretion of Amphiregulin following stimulation with CXCL8; this may play a role in the EGFR activation seen. Conclusions: Our studies suggest CRC malignant epithelium is subject to increased autocrine CXC-chemokine signaling compared to the adjacent normal colorectal tissue and this may reflect a role for the signaling in this disease. In-vitro work suggests that chemotherapy-induced CXC-chemokine signaling may underpin CRC chemoresistance through multiple pathways. These results support a role for CXC-chemokine signaling in the promotion of CRC and modulating sensitivity of this disease to current treatment Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A76.