Analysis of angiogenesis genes from paraffin-embedded breast tumor and lymph nodes

Background. Angiogenesis is important in tumor growth and metastasis. Germ-line polymorphisms critical to the angiogenesis pathway have been shown to confer prognostic information in multiple tumor types. These genes include vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS). Experimental design. We extracted DNA from 53 specimens obtained from 21 patients including a primary breast tumor, and/or a histologically involved lymph node, and/or a histologically normal lymph node. We subsequently genotyped all specimens to evaluate two polymorphisms in the eNOS gene and one polymorphism in the VEGF gene. Results. Chromatographs were generated in 145/159 (91%) samples. When assessing all polymorphisms by site, chromatographs were generated in 42/51 (83%) samples obtained from the primary tumor and 103/108 (95%) from lymph nodes. Chromatographs were generated in 46/53 (87%) samples from the T-786C polymorphism in the 5'-flanking region in the eNOS gene, 49/53 (92%) when assessing the Glu(298)Asp polymorphism in exon 7 in the eNOS gene and 50153 samples (94%) for the (CT)-T-936 polymorphism in the VEGF gene. There wash 100% concordance between analyses from the primary tumor, uninvolved lymph node, and involved lymph node from the same case. Conclusion. We successfully extracted DNA and genotyped several polymorphisms in two genes important in angiogenesis. These genotypes were determined in breast tumors, but also in involved and uninvolved lymph nodes. There was concordance between the genotypes of germline DNA obtained from uninvolved lymph nodes and those determined in tumor samples, implying that the host angiogenic genotype imprints the tumor genotype.