(A) SK, SKM, P1 and sN1 cells were subcutaneously injected into the flanks of nude mice, respectively, and tumor volumes measured once a week (n = 4 for each group). Tumor growth curves are shown. (B) Six weeks later, tumors were excised and weighed. C–E, Tumors dissected from mice subjected to subcutaneous injection with control and SK-sNEDD4 cells were introduced orthotopically into nude mice and monitored weekly with IVIS. (C) left, Representative IVIS images of mice 5 weeks after tumor implantation (9 mice/group). The metastasis-free survival curves are shown (C, right). Mice were sacrificed 8 weeks after implantation. Livers (the implantation site is highlighted with arrows in panel D) and lungs were excised, fixed and embedded in paraffin. Hematoxylin & Eosin (H&E) staining was performed on liver (E, left) and lung (E, right) tissue sections. N, normal; T, tumor; *P < 0.05; **P < 0.01; ***P < 0.001.