Sunitinib inhibited angiogenesis, lymphangiogenesis, and lymph node metastasis in the m.f.p. model. (a) Immunohistochemical analysis of the primary tumor treated with sunitinib, DC101, or vehicle. Tumors were stained with anti-CD31 for endothelial cells (upper) and anti-LYVE-1 for lymphatic endothelial cells (lower). Photographs were taken under a microscope at a magnification of ×8. Photographs of representative areas are shown. The black line represents 50 μm. (b) Effects of sunitinib and DC101 on tumor angiogenesis and lymphangiogenesis. The microvessel density (MVD, top) was determined by staining for CD31, and the lymphatic microvessel density (LVD, bottom) by staining for LYVE-1. *P < 0.05 as compared with control. (c) Lymph node metastasis was detected by bioluminescent imaging in vivo. The primary tumor (left) was identified on day 1 of treatment, and tumor metastasis around the axillary lymph node was detected after treatment with vehicle control (right, upper) or sunitinib (right, lower). Ex vivo data of the lymph nodes confirmed metastasis from the MDA-MB-231LN primary tumor. Three representative photographs from each treatment group are shown. LVD, lymphatic microvessel density; m.f.p., mammary fat pad; MVD, microvessel density.