HGAL expression inhibits breast cancer cell growth, metastasis, and invasion. Mice were injected intravenously with 3 × 10⁶ luciferase-transduced human MDA-MB-231–derived breast cancer subline 4175 cells. D-luciferin was injected intraperitoneally (150 mg/kg body weight) 15 minutes before acquiring images. Bioluminescence images were taken once per week. (A) Representative bioluminescent images of mice with subline 4175 tumors. (B) Tumor growth curve based on photon flux intensity (10 mice in each group) with significant difference between the mouse cohorts with 4175 Mock and 4175 HGAL tumors (P = .002). Mice that were not injected (background; 5 mice) served as a control for background photon flux intensity. Normalized photon flux represents tumor photon flux (luminescence intensity) at the indicated time relative to photon flux measured after cell injection on day 0 that was set to a value of 100. (C) A representative images of lungs with 4175 Mock and 4175 HGAL tumors. (D) Number of lung tumors in mice injected with 4175 Mock and 4175 HGAL cells (P = .001). (E) Representative images of in vitro 3D spheroid cell invasion in 4175 Mock and 4175 HGAL cells. Scale bar represents 200 µm. (F) Quantitative analysis of surface area of 4175 Mock and 4175 HGAL spheroids over a 6-day period (P = .0002). Experiments in panels D, E, and F were performed in triplicate and were repeated twice. ***P = .001.