Triple negative breast cancers are defined by their lack of expression of HER, oestrogen receptors and progesterone receptors. They account for around 10-24% of cases. To define a cancer by the biomarkers it does not express is unsatisfactory. Triple negative breast cancer has been linked to aspects of metabolism and metabolic disorders such as diabetes. There are several biomarkers that are of interest and overlap in both their roles in breast cancer and in aspects of metabolism. Fatty acid binding protein 7 (FABP7) is one of 9 FABPs that is involved in the transport, solubilisation and regulation of metabolism of various fatty acids. Expression profiling and immunohistochemistry (IHC) studies have identified FABP7 to be over-expressed in a subtype of breast cancer that can be considered almost synonymous with triple negative breast cancer; basal-like breast cancer, so called because it expresses cytokeratins that are characteristic of basal epithelial cells. The role of FABP7 in breast cancer is not fully understood and studies have given conflicting results in regards to the relation to prognosis. Evidence suggests that FABP7 can be regulated by methylation acetylation and exposure to fatty acids. Insulin like growth factor binding protein-2 (IGFBP-2) is a member of the IGF-axis that is responsible for altering cell growth and metabolism. IGFBP-2 has been found to be over-expressed in many cancers including those of the prostate and breast. Phosphotensin homolog (PTEN) is a tumour suppressor gene that is responsible for dephosphorylating PIP3 to inhibit the Akt pathway and thus inhibit cell growth and promote apoptosis. IGFBP-2 has IGF independent actions; it can down-regulate PTEN through binding of an integrin receptor and therefore have mitogenic and anti-apoptotic effects.
To study the expression of the metabolic biomarkers FABP7, IGFBP-2 and PTEN in clinical cases of Malaysian TN breast cancer. To use appropriate cell lines in order to more fully understand whether epigenetic mechanisms and FAs regulate FABP7 expression. To over-express FABP7 in a breast cancer cell lines and to further understand the role of FABP7 in breast cancer.
IHC was used to assess FABP7, PTEN and IGFBP-2 expression in a cohort of triple negative breast cancer cases. FAs, a demethylation agent-AZA and a histone deacetylase inhibitor-TSA were used to investigate what regulated FABP7 in cell lines. Over-expression experiments were used to understand the effect of FABP7 in breast cancer cell lines.
FABP7 expression in patient samples was associated with lower grade, basal-like phenotype, FAS expression and although not significant, improved patient survival. Treatment of BT-20 and MDA-MB-231 cell lines with AZA and TSA resulted in increases in FABP7 mRNA expression. Fatty acid treatment led to changes in FABP7 mRNA expression. Combinations of AZA and fatty acids gave large increases in FABP7 mRNA expression. Over-expression of FABP7 in BT-20 cells resulted in increased cell viability and although not significant changes in expression of survivin, caspase 9 and their splice variants. IGFBP-2 expression was associated with poor patient survival though this was not significant. PTEN loss was a frequent event in the cohort of triple negative breast cancer cases; 48.3% of cases had PTEN loss. PTEN loss was associated with poor patient survival though this was not significant. PTEN loss was associated with expression of IGFBP-2.
Discussion & Conclusions
FABP7 is likely to play a role in patient survival as demonstrated in the patient samples. FABP7 over-expressing BT-20 cells tended to have increased survivin FL and ?EX3 expression. Both increased mitochondrial activity and survivin expression have been found to be associated with improved prognosis in breast cancer and this may explain some mechanisms by which FABP7 results in better prognosis in the TN breast cancer cases in this study. Since FABP7 mRNA expression was not increased to fold changes comparable to oestrogen receptor re-expression after AZA and TSA treatment, it is unlikely that the FABP7 gene is methylated or regulated by acetylation. It is possible that there are genes upstream of FABP7 such as transcription factors that are regulated my methylation and acetylation and therefore impact on FABP7 expression after treatment with AZA and TSA. This is the first study that demonstrates the significant relationship between IGFBP-2 expression and PTEN loss in patient samples. PTEN loss is a frequent event in TN breast cancer. IGFBP-2 and PTEN loss may be useful markers of prognosis in TN breast cancer.
Dean, S. J. An investigation in to the role of fatty acid binding protein-7, insulin like growth factor binding protein-2 and phosphatase and tensin homolog in triple negative breast cancer; in vitro and in vivo. (Thesis). University of the West of England