Delineating the role of prolactin-inducible protein (Pip) in breast cancer lung metastasis

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Evangelista de Lima Terceiro, Lucas
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In Canada, approximately 28,600 women were diagnosed with breast cancer (BC) in 2022, and about 40% of women currently fail primary management strategies and ultimately succumb to this disease. Despite many advances in the treatment of BC, metastatic relapse remains a major challenge for patients. The prolactin-inducible protein (PIP) is a highly breast specific gene product of BC. Because of such relevant attributes, it is considered a valuable BC biomarker frequently used in the clinic to determine the origin of an unknown metastatic carcinoma. However, the role of PIP in BC progression is still poorly understood. To address this role, our laboratory developed a novel transplantable mouse model of BC utilizing the 4T1-mouse metastatic BC cell line and determined that while over-expression of mouse homologous (Pip) suppressed tumor development in the primary tumor in the breast, it enhanced metastasis in lungs. We hypothesized that in the lung, Pip promotes a more aggressive phenotype in the BC cells. To further address this hypothesis Pip-expressing (4T1/Pip) and control BC cells (4T1/EV) were isolated from primary tumors and metastatic lungs of these mouse models and grown as ex vivo cell cultures. Several indicators of cellular transformation and metastatic progression were evaluated. Gene expression analysis of 4T1/Pip cells isolated from the metastatic lungs identified an up-regulation of genes involved in extracellular matrix disassembly, angiogenesis, positive regulation of cell migration and cell proliferation. Functional analysis studies showed that in the lungs, 4T1/Pip cells displayed increased proliferation, enhanced migration and colony formation compared to control. As well, morphological analysis of ex vivo cell cultures revealed that the 4T1/Pip BC cells developed a mesenchymal-like morphology compared to controls. Key EMT genes were also found up-regulated, whereas some epithelial markers were down-regulated in the 4T1/Pip cells from lungs. Furthermore, 4T1/Pip cells showed higher ERK1/2 phosphorylation ratio and cell proliferation when cultured on a fibronectin-coated surface, suggesting that Pip may activate tumor progression pathways through fibronectin fragments interacting with membrane proteins. Collectively, these studies show that in the lung, Pip-expressing breast cancer cells acquire more aggressive traits consistent with the metastatic phenotype, suggesting that Pip may function differently in different tumor microenvironments. This work reveals a novel function of Pip in regulating breast cancer progression and metastasis.
Prolactin Inducible Protein, Triple Negative Breast Cancer, Transplantable Mouse Model, Lung Metastasis