Intracrine PTHrP Induces Secretory Differentiation and Tumor Development in a Breast Cancer Model

Abstract

Parathyroid hormone-related protein (PTHrP) is required for embryonic mammary gland development and has important functions during lactation, when its production is greatly upregulated by alveolar epithelial cells and secreted into the maternal circulation to mobilize skeletal calcium stores for milk production. PTHrP is also produced by breast cancers and GWAS studies suggest that it influences breast cancer risk. However, the exact functions of PTHrP in breast cancer biology remain unsettled. We used a tetracycline-reg-ulated, mouse mammary tumor virus long terminal repeat (MMTV) system to control the timing of PTHrP overexpression in mammary epithelial cells (Tet-PTHrP mice) and bred these mice with the MMTV-PyMT (Tet-PTHrP;PyMT mice), a standard model of breast cancer, to analyze the impact of PTHrP overexpression on normal mammary gland biology and in breast cancer progression. Virgin Tet-PTHrP mice treated with Dox developed epi-thelial hyperplasia with increased epithelial cell proliferation, expression of Elf5, activation of Stat5 and premature secretory differentiation of mammary epithelial cells with milk pro-duction. We also found that PTHrP overexpression does not impair normal lactation, but it did delay post-lactation involution. In Tet-PTHrP;PyMT mice, overexpression of PTHrP shortened tumor latency and accelerated tumor growth, ultimately reducing overall survival. Tumors overproducing PTHrP histologically resembled secretory carcinomas of the breast and also exhibited increased expression of nuclear pSTAT5 and Elf5, several markers of secretory differentiation, and multiple milk proteins. Treating isolated tumor cells from Tet-PTHrP;PyMT mice with Dox stimulated cell proliferation, activated STAT5 and increased milk protein gene expression, but adding exogenous PTHrP did not. Also, neither ablating the (PTH1R) in epithelial cells nor treating Tet-PTHrP;PyMT mice with an anti-PTH1R antibody prevented secretory differentiation or altered tumor latency, suggesting that PTHrP acts in a cell-autonomous, intracrine manner. Finally, we analyzed single-cell sequencing data derived from 27 different human breast tumors and found that PTHrP expression is associated with upregulation of genes involved in milk production and STAT5 signaling in human breast cancers. Our study suggests that PTHrP promotes pathways leading to secre-tory differentiation and proliferation in both normal mammary epithelial cells and in breast tumor cells.