The prognostic significance and impact of the CXCR4-CXCR7-CXCL12 axis in primary cutaneous melanoma.

BACKGROUND: Expression of the chemokine receptor CXCR4 is known to regulate melanoma metastasis to distant sites with high expression of the CXCL12 ligand. However, the prognostic impact of CXCR4 expression and potential for autocrine-mediated activation of prosurvival mitogen-activated protein kinase signalling remains enigmatic. Furthermore, expression of the decoy receptor CXCR7 within the local cutaneous melanoma microenvironment remains undefined. OBJECTIVES: To define the contribution and prognostic impact of CXCR4-CXCR7-CXCL12 signalling in primary cutaneous melanomas and the immediate tumour microenvironment. METHODS: Immunohistochemical/immunofluorescent expression of CXCR4, CXCR7 or CXC12 was analysed in human metastatic melanoma cell lines, primary cutaneous cell types and a retrospective cohort of primary melanomas/benign naevi. CXCL12 secretion by melanoma/cutaneous cells was evaluated by enzyme-linked immunosorbent assay, and autocrine CXCR4-CXCL12 signalling was investigated by addition of a CXCL12-neutralizing antibody. RESULTS: CXCR4 expression was significantly higher in primary melanomas that subsequently metastasized after 7 years (P = 0·037). Stratification for American Joint Committee on Cancer (AJCC) stage II disease revealed significantly decreased disease-free survival in patients with > 50% CXCR4 expression (P = 0·036), while comparative analysis of CXCL12 expression in the adjacent epidermis of all AJCC stage melanomas revealed increased CXCL12 correlated with prolonged time to metastasis (P = 0·014). CXCR7 was expressed within the primary melanoma microenvironment but was absent on primary tumours. Addition of anti-CXCL12 to BRAF-mutant melanoma cells resulted in downregulation of phospho-CXCR4 and phospho-extracellular signal-related kinase, indicating autocrine CXCR4-CXCL12 signalling. CONCLUSIONS: CXCR4 expression defines a potential prognostic biomarker for AJCC stage II melanoma. Moreover, targeting the CXCR4-CXCR7-CXCL12 axis may represent a novel therapeutic strategy to prevent early melanoma progression.

Inhibition of CXCR4-CXCL12 chemotaxis in melanoma by AMD11070.

BACKGROUND: Despite intensive research and novel adjuvant therapies, there is currently no cure for metastatic melanoma. The chemokine receptor CXCR4 controls metastasis to sites such as the liver; however, the therapeutic blockade with the existing agents has proven difficult. METHODS: AMD11070, a novel orally bioavailable inhibitor of CXCR4, was tested for its ability to inhibit the migration of melanoma cells compared with the commonly described antagonist AMD3100. RESULTS: AMD11070 abrogated melanoma cell migration and was significantly more effective than AMD3100. Importantly for the clinical context, the expression of B-RAF-V600E did not the affect the sensitivity of AMD11070. CONCLUSION: Liver-resident myofibroblasts excrete CXCL12, which is able to promote the migration of CXCR4-expressing tumour cells from the blood into the liver. Blockade of this axis by AMD11070 thus represents a novel therapeutic strategy for both B-RAF wild-type and mutated melanomas.

Therapy with nonglycosaminoglycan-binding mutant CCL7: a novel strategy to limit allograft inflammation.

Chemokines are immobilized by binding to glycosaminoglycans (GAGs). A non-GAG-binding mutant CCL7 (mtCCL7) was developed that retained its affinity for chemokine receptors. This mtCCL7 induced leukocyte chemotaxis in diffusion gradients but did not stimulate trans-endothelial migration (p<0.01). Unlike wild-type CCL7, mtCCL7 persisted in the circulation of BALB/c mice for more than 6 h and prevented leukocyte infiltration of skin isografts (p<0.05). Treatment with mtCCL7 marginally increased the survival of C57BL/6 to BALB/c skin allografts and reduced graft infiltration by CD3+ cells (p<0.05). Importantly, mtCCL7 promoted long-term (>40 day) graft survival following minor histocompatibility (HY) antigen mismatched C57BL/6 skin transplantation; control grafts were rejected by day 24. Treatment with mtCCL7 produced a significant decrease in the frequency of IFN-gamma producing donor-reactive splenic T cells, reduced CCR2 expression by circulating leukocytes for 6 h (p<0.01) and blocked the normal increase in affinity of alpha4beta1 integrins for VCAM-1 following transient chemokine stimulation. These data suggest that mtCCL7 persists in the circulation and reduces both specific T-cell priming and the capacity of circulating immune cells to respond to GAG-bound chemokine at sites of developing inflammation.

Modulatory effects of heparin and short-length oligosaccharides of heparin on the metastasis and growth of LMD MDA-MB 231 breast cancer cells in vivo.

Expression of the chemokine receptor CXCR4 allows breast cancer cells to migrate towards specific metastatic target sites which constitutively express CXCL12. In this study, we determined whether this interaction could be disrupted using short-chain length heparin oligosaccharides. Radioligand competition binding assays were performed using a range of heparin oligosaccharides to compete with polymeric heparin or heparan sulphate binding to I(125) CXCL12. Heparin dodecasaccharides were found to be the minimal chain length required to efficiently bind CXCL12 (71% inhibition; P<0.001). These oligosaccharides also significantly inhibited CXCL12-induced migration of CXCR4-expressing LMD MDA-MB 231 breast cancer cells. In addition, heparin dodecasaccharides were found to have less anticoagulant activity than either a smaller quantity of polymeric heparin or a similar amount of the low molecular weight heparin pharmaceutical product, Tinzaparin. When given subcutaneously in a SCID mouse model of human breast cancer, heparin dodecasaccharides had no effect on the number of lung metastases, but did however inhibit (P<0.05) tumour growth (lesion area) compared to control groups. In contrast, polymeric heparin significantly inhibited both the number (P<0.001) and area of metastases, suggesting a differing mechanism for the action of polymeric and heparin-derived oligosaccharides in the inhibition of tumour growth and metastases.