Localization of erythropoietin receptor mRNA in primary breast tumors by laser capture microdissection

Abstract

e22036 Background: Adverse effects of erythropoiesis stimulating agents on tumor progression and/or survival were observed in recent Phase III clinical trials, however, mechanisms are not understood. Whether tumor erythropoietin receptor (EpoR) expression is associated with erythropoietin-dependent tumor progression remains unclear, owing in part to the lack of specificity of commercial antibodies for protein detection. To overcome issues of protein detection, we previously optimized a quantitative RTPCR approach for reliable measurements of low level EpoR mRNA in primary tumor samples, and observed a 30-fold range of EpoR expression among panels of both breast and head and neck cancer samples. Methods: To test whether EpoR mRNA is expressed in tumor epithelial cells, we performed laser capture microdissection to fractionate 3 breast tumors into tumor epithelial enriched vs. depleted fractions. In each fraction, lineage marker and EpoR mRNA expression was normalized to 3 endogenous control genes. Results: Vascular endothelial markers in tumor epithelial-enriched fractions were reduced by an average of 15.3-fold (vascular endothelial cadherin) and 18.5-fold (platelet cell adhesion molecule 1) while the stromal marker vimentin was reduced by 14.7-fold. EpoR expression was enriched by 4.9-, 7.2-, and 1.1-fold in epithelial enriched fractions. Conclusions: Despite the depletion of endothelial cells, EpoR mRNA was at the same level or enriched in tumor epithelial fractions. These results demonstrate that malignant epithelial cells are a major source of EpoR mRNA in primary tumors. Applying this approach to additional breast cancer specimens as well as other cancer types will improve our understanding of erythropoietin-associated tumor progression. [Table: see text]