Polysaccharide krestin (PSK) is a mushroom extract that has long been used in Asia as an anti-cancer drug, although the mechanism of action is unclear. Using neu-transgenic

(neu-tg) mice, a model of human HER2 positive breast cancer, we investigated the anti-tumor effect of PSK and its potential mechanisms. We found that oral administration of PSK significantly inhibited the growth of both implanted and spontaneous breast tumors in these mice. In vitro PSK treatment stimulated splenocytes proliferation and secretion of TNFalpha. PSK activated dendritic cells (DC) from neu-tg mice as shown by increased expression of CD86, CD80, and MHCII. The expression of toll-like receptor 2 (TLR2) on DC was also increased. To study the potential activating effect of PSK on TLRs, we used HEK293 cells transfected with different TLRs and incubated them with PSK (0.5 to 1500ug/ml, 16 hour). PSK activated NFkB in HEK cells transfected with TLR2 in a dose-dependent manner, but had no effect on HEK transfected with other TLRs (4, 7, or 8). To further investigate the role of TLR2 in mediating the immunostimulatory effect of PSK, we used splenocytes from TLR2 knockout (k/o) mice and WT mice and treated them with PSK (25 to 400ug/ml, 72 hour). PSK stimulated the secretion of TNFalpha in splenocytes from WT mice dose-dependently but not in splenocytes from k/o mice. These results suggest that TLR2 is critical in mediating the immunostimulatory effect of PSK.