Abstract 985: Stromal PDGFR-α activation stalls mammary ductal development and increases tumorigenic potential of mammary epithelia

Abstract

Abstract One of the major confounding factors in the treatment of breast cancer is the cellular heterogeneity of the breast. An important cellular component of the normal breast as well as tumor stroma is the fibroblast. Fibroblasts are cells of mesenchymal origin that express platelet derived growth factor receptor alpha (PDGFR-α), a pharmaceutically targetable receptor tyrosine kinase. While activation of PDGFR-α was shown to be a poor prognostic factor in glioblastoma and gastro-intestinal stromal tumors, the role of stromal PDGFR-α signaling in mammary development and carcinomas is yet to be explored. To elucidate the role of stromal PDGFR-α signaling in this context, we utilized mice harboring the activating Pdgfra-D842V mutation knocked into the endogenous Pdgfra locus under a Lox-STOP-Lox cassette. To obtain selective activation of PDGFR-α in the fibroblastic stroma, these mice were crossed with fibroblast specific Fsp-cre mice. Whole mount carmine alum staining revealed that the D842V mutants have severely impaired mammary ductal development. Further analyses at the histological level revealed that the D842V glands have increased extra-cellular matrix (ECM) deposition (Masson's trichrome) likely due to the observable increase in fibroblasts (vimentin positivity). Immuno-fluorescent dual staining for keratin 8 and Ki67 demonstrated that stromal PDGFR-α activation increases the proliferation of neighboring mammary ductal cells, potentially making them more tumorigenic. To gain insight into the molecular mechanisms underlying this stromal-epithelial cross-talk, primary mammary epithelial cells and fibroblasts were isolated for downstream assays. Western blot analyses showed that mutant mammary fibroblasts have increased levels of activated PLC-γ, AKT and JNK which are known oncogenic pathways in the stroma. Gene expression arrays followed by global pathway analyses showed that pro-tumorigenic processes such as proliferation and metastasis are enriched in the epithelium from the mutant mice, while pro-tumor pathways such as KRAS and JNK were found to be enriched in the mutant fibroblasts. Interestingly, Pdgfra-D842V mutant mice have significantly increased stiffness as measured by atomic force microscopy of naïve tissue. This result implies that the observed increase in fibroblast number and resulting ECM deposition is sufficient to alter the entire mammary tissue. Since breast density correlates with higher breast cancer risk and tumor invasiveness, this phenotype reveals a novel function for stromal PDGFR-α signaling in breast cancer susceptibility. Studies are underway to evaluate whether these mice are predisposed to mammary tumor formation. Given our present data, we conclude that activation of stromal PDGFR-α signaling not only abrogates mammary development, but also makes the mammary ductal epithelium more tumorigenic. Citation Format: Anisha Mathur, Gina M. Sizemore, Subhasree Balakrishnan, Vasudha C. Shukla, Maria Cuitino, Anthony J. Trimboli, Samir Ghadiali, Gustavo W. Leone, Michael C. Ostrowski. Stromal PDGFR-α activation stalls mammary ductal development and increases tumorigenic potential of mammary epithelia. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 985. doi:10.1158/1538-7445.AM2015-985