Ninjinyoeito (NYT) and Juzentaihoto (JTT) are immunomodulatory Kampo medicines, and clinically used to support cancer therapy. Our have previously shown that these medicines can suppress the differentiation of Myeloid-derived suppressor cells (MDSC), which is known to be accumulated in tumor and potently inhibits T-cell and NK-cell activity. In the tumor-bearing state, MDSC migrates to the tumor microenvironment (TME) and becomes an activated state that produces immunosuppressive factors, to promote tumor growth. In this study, we have investigated the effects of NYT and JTT on migration and activation in TME. To prepare MDSC, bone-marrow cells were isolated from C57BL/6J mice and differentiated into MDSC by the treatment with IL-6 and GM-CSF. The treatment of 4T1 breast cancer cells-conditioned media to MDSC markedly increased expression of immumosuppressive factors, arginase-1 and iNOS, and these increase in mRNA inhibited by NYT and JTT. In Transwell assay, migration of MDSC was stimulated by treatment of 4T1 cells or 4T1-conditioned media, NYT and JTT significantly inhibited this migration. These data indicated that, NYT and JTT suppressed not only differentiation, but also the activation and the migration of MDSC in TME. The multi-steps inhibition may be important in the immunomodulatory effects of these Kampo medicines.