Analysis of micro and molecular mechanism of lymphatic metastasis, and development of new strategy targeting for tumor lymphatic vessel

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We investigated the interaction between CD44 and its ligand, hyaluronic acid, on lymph node metastasis of colorectal cancers. In 90% of nude mice, lymph node metastasis was established by alloic transplantation of LS174T and LS174T-NEO, which expressed high levels of CD44. In contrast, Colo320 and LS174T-AS2, which were transfected with antisense CD44 gene, had no expressions of CD44 and never metastasized to lymph nodes by alloic transplantation in nude mice. In immunohistochemistry of lymphatic vessels, LS174T-NEO showed marked lymphatic invasion, in contrast to minimal lymphatic invasion in LS174T-AS2 and Colo320. LS174T and LS174T-NEO widely attached to normal lymph nodes, and it was inhibited by anti-hyaluronic acid processing. On the other hand, the attachment to lymph nodes was significantly reduced in LS174T-AS2.We evaluated the ability of colorectal cancer cell lines on metastasis to lymph nodes, and metastatic (or anti-metastatic) genes were screened by cDNA microarray. Twenty three of human colorectal cancer cell lines were investigated on the lymph node metastasis in alloic transplantation model and the incidence of metastasis was varied widely as follows : KM12c 100%(7/7), CloneA 100%(9/9), HT29 94.4%(17/18),..., SW1116 25%(2/8), DLD1 0%(0/13). Cell lines were divided into three groups (high, middle and low) according to the incidence of lymph node metastasis. Based on cDNA microarray analysis together with in vivo experiments, 107 candidates of metastasis-related genes were extracted. The expression levels of some of those genes were confirmed elevated or depressed with real-time RT-PCR as well. Further, there was a relationship between expression levels of candidate genes and clinical lymph node metastasis in colorectal cancers.