In order to reduce the inherent risk of serum-supplemented media in cancer patients treated with LAK cells and induce high cytotoxic activity, we have developed a serum-free medium designated as RD5F. RD5F medium consists of transferrin, 2-aminoethanol, 2-mercaptoethanol, sodium selenite and interleukin-2 in addition to nutrient mixture. The cytotoxic activity of LAK cells generated in RD5F is higher than that in medium containing 10% human AB serum against Raji, K562 cells, and oral cancer cells. We have also found that insulin and insulin-like growth factor-1 (IGF-1), which is considered to be the most important hormone of serum-free media for animal cells, inhibited the induction of cytotoxic activity of LAK cells derived from PBL and RLNL, and that transferrin was an essential component of serum-free media for induction of LAK cells. The results obtained by two-color FACS analysis suggested that insulin stimulated differentiation of PBL to CD4+ cells in serum-free culture. Tumor infiltrating lymphocytes (TIL) generated in RD5F medium exhibited CD3 + HLA-DR + phenotype and much higher cytotoxicity against autologous tumor cells than that against allogenic cells. These results strongly suggest the possibility that insulin and IGF-1 act as immunosuppresive factors. In addition, RD5F medium was suggested to be useful in the adoptive immunotherapy of cancer.
