The dissemination of tumor cells to sites far from the primary tumor (metastasis) is the principal cause of death in cancer patients. Tumor-associated lymphatic vessels are a key conduit for metastatic tumor cells, which typically first colonize the lymph nodes. Although the primary tumor and affected lymph nodes can be removed during surgery, tumor cells inside lymphatic vessels are left behind. Here, we show that in-transit tumor cells inside lymphatic vessels in mice bearing mouse melanomas or human lung tumors give rise to metastases. Using photodynamic therapy with the benzoporphyrin derivative verteporfin, we selectively destroyed lymphatic vessels in mice and pigs. Destruction of tumor-associated lymphatic vessels also eradicated intralymphatic tumor cells and prevented metastasis of mouse melanoma cells and subsequent relapse. Photodynamic therapy, when combined with anti-lymphangiogenic therapy, prevented further tumor invasion of lymphatic vessels. These findings highlight the potential of targeting in-transit tumor cells in patients.