Aquaporin 1 (AQP1) is a plasma membrane water‐transporting protein expressed strongly in tumor microvascular endothelia. We previously reported impaired angiogenesis in implanted tumors in AQP1‐deficient mice and reduced migration of AQP1‐deficient endothelial cells in vitro. Here, we investigated the consequences of AQP1 deficiency in mice that spontaneously develop well‐differentiated, luminal‐type breast adenomas with lung metastases mouse mammary tumor virus‐driven polyoma virus middle T oncogene (MMTV‐PyVT). AQP1+/+ MMTV‐PyVT mice developed large breast tumors with total tumor mass 3.5 ± 0.5 g and volume 265 ± 36 mm3 (se, 11 mice) at age 98 d. Tumor mass (1.6±0.2 g) and volume (131±15 mm3, 12 mice) were greatly reduced in AQP1–/– MMTV‐PyVT mice (P<0.005). CD31 immunofluorescence showed abnormal microvascular anatomy in tumors of AQP1–/– MMTV‐PyVT mice, with reduced vessel density. HIF‐1α expression was increased in tumors in AQP1–/– MMTV‐PyVT mice. The number of lung metastases (5± 1/mouse) was much lower than in AQP1+/+ MMTV‐PyVT mice (31±8/mouse, P<0.005). These results implicate AQP1 as an important determinant of tumor angiogenesis and, hence, as a potential drug target for adjuvant therapy of solid tumors.—Esteva‐Font, C., Jin, B.‐J., Verkman, A. S. Aquaporin‐1 gene deletion reduces breast tumor growth and lung metastasis in tumor‐producing MMTV‐PyVT mice. FASEB J. 28, 1446–1453 (2014). www.fasebj.org