Supplementary Figure S9: Human TNBC cell lines MDA-MB-231 and MDA-MB-468 were injected in the T4 mammary fat pad of 12 week old nude mice. When tumors reached size of 100mm3, mice were randomized in Vehicle and AZD9668 100mg/kg groups. Mice were then treated daily B.I.D until the endpoint of Vehicle tumors reaching 2000mm3. (A, B) Tumor growth curves of mice treated with either Vehicle or AZD9668 100mg/kg in the MDA-MB-231 and MDA- MB-468 models respectively. Endpoint tumors harvested were subject to BrdU staining. (C, D) Representative images BrdU staining and (E, F) their corresponding quantitation in the MDA- MB-231 and MDA-MB-468 models respectively. (G) Spontaneous tumors isolated from PyMT NE+/+ were implanted in FVB NE+/+ mice. When tumors reached a measurable size of 30- 50mm3, mice were randomized into Vehicle and AZD9668 100mg/kg group and treated B.I.D until 400-500mm3. Tumor growth curves, as a function of time are shown.
ARTICLE ABSTRACT
Metastatic disease remains the leading cause of death due to cancer, yet the mechanism(s) of metastasis and its timely detection remain to be elucidated. Neutrophil elastase (NE), a serine protease secreted by neutrophils, is a crucial mediator of chronic inflammation and tumor progression. In this study, we used the PyMT model (NE+/+ and NE−/−) of breast cancer to interrogate the tumor-intrinsic and -extrinsic mechanisms by which NE can promote metastasis. Our results showed that genetic ablation of NE significantly reduced lung metastasis and improved metastasis-free survival. RNA-sequencing analysis of primary tumors indicated differential regulation of tumor-intrinsic actin cytoskeleton signaling pathways by NE. These NE-regulated pathways are critical for cell-to-cell contact and motility and consistent with the delay in metastasis in NE−/− mice. To evaluate whether pharmacologic inhibition of NE inhibited pulmonary metastasis and phenotypically mimicked PyMT NE−/− mice, we utilized AZD9668, a clinically available and specific NE inhibitor. We found AZD9668 treated PyMT-NE+/+ mice showed significantly reduced lung metastases, improved recurrence-free, metastasis-free and overall survival, and their tumors showed similar molecular alterations as those observed in PyMT-NE−/− tumors. Finally, we identified a NE-specific signature that predicts recurrence and metastasis in patients with breast cancer. Collectively, our studies suggest that genetic ablation and pharmacologic inhibition of NE reduces metastasis and extends survival of mouse models of breast cancer, providing rationale to examine NE inhibitors as a treatment strategy for the clinical management of patients with metastatic breast cancer.
