Inflammatory breast cancer (IBC) is considered the most lethal form of breast cancer due to its ability to progress quickly and the frequent presence of metastasis at diagnosis. African Americas are disproportionally diagnosed with IBC and often have worse outcomes than Caucasians. By investigating IBC in both African American and Caucasian cell lines we seek to understand the differences in IBC progression and help address disparities by providing new anti-IBC strategies. RhoC GTPase is overexpressed in 90% of IBC tumors and is known to increase cell motility. Sites of inflammation, as seen in IBC, attract tumor associated macrophages (TAMs), which have been found to facilitate the movement and invasion of many breast cancers. We hypothesize that TAMs play a role in increasing RhoC expression in IBC cell lines, consequently leading to IBC's severe migratory and metastatic potential.
A novel microfluidic device created by our team was used to measure the migratory phenotype of IBC cell lines in response to macrophage conditioned media (CM) and cytokine stimulation. IBC cell lines were treated with CM, cytokines, or pathway inhibitors then Western blotting was used to determine protein expression and phosphorylation to identify important signaling pathways.
We found the expression of RhoC significantly increased in two different IBC cell lines, SUM149 (African American) and SUM190 (Caucasian), after culturing with conditioned media from the macrophage-differentiated U937 monocytic cell line. This increase was not detected in either the normal-like MCF-10A breast epithelial cell line or the non-IBC MDA-MB-231 triple negative breast cancer cell line. CM caused a significant increase in the migration distance and frequency of both SUM149 and SUM190 cell lines. Analysis of the CM determined CCL2, CCL5, and IL-8 to be the key mediators in the macrophage CM. Western blotting proposes that CCL2, CCL5, and IL-8 stimulation causes twice as much RhoC expression compared to the control. Further analysis suggests a role for the MAPK pathway in controlling RhoC expression and migration.
Macrophage conditioned media causes an increase in RhoC expression in IBC cell lines and stimulates migration. Individual cytokines can lead to an increase in RhoC possibly through the MAPK pathway. Studies involving RhoC inhibitors are ongoing and could yield promising therapies for the prevention of metastasis in IBC. By understanding the specific mechanism of TAMs’ effects on IBC, we hope to learn how to control the lethal metastatic nature of IBC and improve outcomes for patients of all ethnicities.
Citation Format: Julie Madden, Steve Allen, Yu-Chi Shen, Chelsea Fournier, ZhiFen Wu, Liwei Bao, Sofia Merajver. Macrophages increase the expression of RhoC in inflammatory breast cancer leading to increased migration. [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 3236.