Rhoa, Rac1 And Pak1 Evaluation In Paired Stromal Fibroblasts Of Breast Cancer Primary And Of Lymph Node Metastasis: Importance Of These Biomarkers In Lymph Node Invasion

The importance of tumor-stromal cells interactions in breast tumor progression and invasion has been recognized; therefore modifications in the stromal fibroblasts can play a significant role in overall cancer development and progression, the presence of fibroblasts in the breast cancer microenvironment metastatic lymph nodes was described, suggesting its role in invasion process. Here after having evaluated the differential genomic profile of carcinoma associated fibroblasts (CAFs) when compared to fibroblasts derived from tissues adjacent to fibroadenomas (NAFs), we depicted focal adhesion as an altered prominent pathway in microarray studies; we validated these data through confocal assays. In order to verify possible role of fibroblasts in lymph node invasion we constructed a tissue microarray consisting of primary BC samples and corresponding lymph node metastasis and compared the expression of some adhesion markers in fibroblasts of the two locations. For the in vitro part of this work stromal fibroblasts were isolated from four invasive breast carcinoma samples and from four women with benign breast disorders. RNA was extracted from low-passage cultures and analyzed with the CodeLink Human Whole Genome cDNA Microarray platform to assess the genomic differential expression. For evaluation of Rho-GTPases: RhoA and Rac1 and a Rho-GTPase effector PAK1 (P21-activated kinase 1), two distinct TMAs were constructed from stromal component of 43 primary tumors and from matched lymph node samples, respectively, fibroblasts were characterized by the expression of α-SMA and vimentin. Our immunohistochemistry revealed 83,7% RhoA positivity in fibroblasts of primary tumors and 71,4% of respective metastatic lymph nodes (p=0,271); 71,8% of fibroblasts of primaries presented Rac1 positive staining, whereas the frequency of positivity in same cells of lymph nodes was 55,3% (p=0,16). With respect to Pak1 50% of patients were positive in the primary tumor against 73,7% in lymph nodal fibroblasts (p=0,039). Our data point to the importance of focal adhesion factors in stromal cells. Since we did not found significant changes between primary and metastatic lymph nodes, we highlight fibroblasts as an active participant in the invasion to lymph nodes, supporting the idea that metastatic tumor cells continue to be dependent of their microenvironment. Supported by Fapesp Citation Format: Patricia Bortman Rozenchan, Fiorita GL Mundim, Rosimeire A. Roela, Maria LH Katayama, Fatima S. Pasini, Helena Brentani, Eduardo C. Lyra, Maria AAK Folgueira, Maria M. Brentani. RHOA, RAC1 and PAK1 evaluation in paired stromal fibroblasts of breast cancer primary and of lymph node metastasis: Importance of these biomarkers in lymph node invasion. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 186. doi:10.1158/1538-7445.AM2014-186