Lentinus edodes (Berk.) Singer fruiting bodies were cultivated on substrates composed of beech sawdust, wheat bran, and calcium sulfate hemihydrate (gypsum), containing different proportions of olive oil press cakes (OOPC). We determined the influence of OOPC on fruiting bodies production and proliferation of CCRF-CEM leukemia cells. A negative influence of OOPC on mycelia growth and maturation was noticed. When growth medium contained 80% OOPC, fruiting bodies ceased forming. To investigate the cytotoxicity on CCRF-CEM cells in vitro, cells were treated with crude polysaccharides extracted from L. edodes fruiting bodies. Also in this case a negative correlation between OOPC content and cytotoxicity was found.