Background

Tumour progression is strongly influenced by environment and extracellular factors derived from stroma. Cancer-associated fibroblast (CAF) present predominantly in tumour stroma, actively supporting tumour growth in association with altered genetics. GCH-1, a gene encoding GTP cyclohydrolase (GTPCH) for tetrahydrobiopterin (BH4) synthesis, is one of the genes significantly up-regulated in patients with colon cancer associated-stromal fibroblasts in liver metastasis. However, the functional consequences of GTPCH expression and BH4 synthesis in CAF for tumour angiogenesis and progression are still not known.

Method

We evaluated effects of GTPCH expression and BH4 synthesis on tumour progression using models of cell cultures and mouse xenografts.

Results

Initially immunohistochemistry staining of tissue arrays from 71 breast cancer patients for GTPCH revealed that 60% of them are strongly positive in cancer cells and 50% positive in both stroma and vessels. We co-cultured MDA231-GFP breast cancer cells with genetically modified murine fibroblasts (GCHtet-off), which express GTPCH under doxycycline (Dox) control, and found that the fibroblast-derived BH4 promoted MDA231-GFP clonogenesis, cell proliferation and migration in cultures. These augmentations were prevented by Dox when GTPCH was switched off, or by the GTPCH inhibitor-DAHP in vitro. Furthermore, cotransplantation of MDA231-GFP and GCHtet-off in SCID mice demonstrated BH4 synthesis in the fibroblasts significantly increased MDA231-GFP tumorigenesis, the tumour cell proliferation, invasiveness to smooth muscles and angiogenesis in comparison to MDA231-GFP alone or with cotransplantation of the control, Tet-off-EV. Conversely, both switching off GTPCH expression by Dox and inhibiting its enzymatic activity by DAHP significantly decreased tumour volume and the tumour vascularisation in vivo.

Conclusion

Our findings suggest that up-regulation of GTPCH for BH4 synthesis in murine fibroblasts may provide favourable environment for tumour angiogenesis and progression. Evaluation of genetically altered GTPCH expression in cancers will potentially provide personalised healthcare for patients.