Abstract 1469: The inflammatory pre-metastatic niche in reliably metastatic models of triple negative breast cancer
Authors: Balestrieri Kassondra , Pittman H Keith , Britt Matthew , Ramez Mohamed , Vohra Nasreen , Verbanac Kathryn 
Published: July 2016

The purpose of this study was to evaluate Triple Negative Breast Cancer (TNBC) murine models to use for investigations of the pre-metastatic niche. We selected two murine TNBC based on published gene expression studies which demonstrated molecular profiles that mirrored human breast tumor subtypes. The 2225L murine tumor has similar gene expression patterns to human tumors of the basal-like phenotype. When 2225L was implanted in the mammary fat pad or sc flank of syngeneic naïve Balb/c female mice, no metastases were observed. Tumors were subsequently resected at approximately 600-800 mm3 to promote the growth of seeded metastases. To select for a consistently metastatic 2225L variant, primary tumors or lung metastases from the most consistently metastatic 2225L tumors were serially passaged sc. The resulting variant (2225LM) is highly metastatic, with lung metastases in 90% (43/48) of mice from the most recent 4 cohorts compared to 27% (47/176) of mice in 66 previous passages. Metastases were not observed at any other site. We also studied the T11 murine tumor, which is representative of the claudin-low phenotype. However, when tested for mouse pathogens prior to implantation, T11 was positive for lactate dehydrogenase elevating virus (LDEV). To eliminate LDEV, T11 tumors were serially passaged into athymic nude rats. The resulting LDEV-free T11 tumor was used for in vivo studies. After sc implantation of T11 in Balb/c mice, consistent lung metastases were observed (89%; 24/27 mice). Tumor-infiltrating immune cell subsets were assessed using flow cytometry. CD11b+ Ly6G- and CD11b+ Ly6G+ cells comprised the majority of tumor-infiltrating immune cells and both subsets were more prevalent in 2225LM than T11. Infiltrating lymphoid (CD3+) cells were a minor component of both tumors. In preliminary studies to quantify lung-infiltrating immune cells, levels of CD11b+ Ly6G+ cells were 1.5 fold higher in 2225LM and T11 tumor-bearing mice compared to controls. Prior to the development of frank metastases (12-30 days post-2225L implant), lung analysis by ELISA showed a steady increase in the neutrophil chemoattractant KC and the pro-tumor M2/N2 chemokine marker MCP-1. Analysis of lungs with visible 2225LM metastases detected increased levels of KC (6-27 fold) and MCP-1 (11-23 fold), when compared to naïve mice. These chemokines were not elevated at non-metastatic sites (i.e. kidney). To conclude, our lab has selected, developed and characterized two TNBC mouse models. Both tumor cell lines are reliable metastatic models of TNBC and will be used to further characterize the inflammatory microenvironment of the pre-metastatic niche, including the continuum of gene expression patterns and changing cellular components.

Citation Format: Kassondra Balestrieri, H Keith Pittman, Matthew Britt, Mohamed Ramez, Nasreen Vohra, Kathryn Verbanac. The inflammatory pre-metastatic niche in reliably metastatic models of triple negative breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1469.