Amy Chang

PURPOSE: Polysaccharide krestin (PSK) is a mushroom extract that has been long used in Asia and recently in Western countries as a treatment for cancer due to its presumed immune potentiating effects. Although there have been reports of clinical responses after patients have ingested PSK, the mechanism of action of the agent remains undefined. The current study was undertaken to investigate the mechanism of the antitumor actions of PSK. EXPERIMENTAL DESIGN: The immunostimulatory effect of PSK was first evaluated in vitro using splenocytes from neu transgenic mice and Toll-like receptor (TLR) 2 knockout (TLR2(-/-)) mice. Then the immunostimualtory and antitumor effect of PSK was determined using tumor-bearing neu transgenic mice, TLR2(-/-), and wild-type C57BL/6 mice. RESULTS: We demonstrate that PSK is a selective TLR2 agonist, and the activation of dendritic cells (DC) and T cells by PSK is dependent on TLR2. Oral administration of PSK in neu transgenic mice significantly inhibits breast cancer growth. Selective depletion of specific cell populations suggests that the antitumor effect of PSK is dependent on both CD8(+) T cell and NK cells, but not CD4(+) T cells. PSK does not inhibit tumor growth in TLR2(-/-) mice suggesting that the antitumor effect is mediated by TLR2. CONCLUSION: These results demonstrate that PSK, a natural product commonly used for the treatment of cancer, is a specific TLR2 agonist and has potent antitumor effects via stimulation of both innate and adaptive immune pathways.

Pancreatic cancer is characterized by extensive stromal desmoplasia, which decreases blood perfusion and impedes chemotherapy delivery. Breaking the stromal barrier could both increase perfusion and permeabilize the tumor, enhancing chemotherapy penetration. Mechanical disruption of the stroma can be achieved using ultrasound-induced bubble activity-cavitation. Cavitation is also known to result in microstreaming and could have the added benefit of actively enhancing diffusion into the tumors. Here, we report the ability to enhance chemotherapeutic drug doxorubicin penetration using ultrasound-induced cavitation in a genetically engineered mouse model (KPC mouse) of pancreatic ductal adenocarcinoma. To induce localized inertial cavitation in pancreatic tumors, pulsed high-intensity focused ultrasound (pHIFU) was used either during or before doxorubicin administration to elucidate the mechanisms of enhanced drug delivery (active vs. passive drug diffusion). For both types, the pHIFU exposures that were associated with high cavitation activity resulted in disruption of the highly fibrotic stromal matrix and enhanced the normalized doxorubicin concentration by up to 4.5-fold compared with controls. Furthermore, normalized doxorubicin concentration was associated with the cavitation metrics (P < 0.01), indicating that high and sustained cavitation results in increased chemotherapy penetration. No significant difference between the outcomes of the two types, that is, doxorubicin infusion during or after pHIFU treatment, was observed, suggesting that passive diffusion into previously permeabilized tissue is the major mechanism for the increase in drug concentration. Together, the data indicate that pHIFU treatment of pancreatic tumors when resulting in high and sustained cavitation can efficiently enhance chemotherapy delivery to pancreatic tumors. .

Multiple TLR agonists have been shown to have antitumor effects in animal models. However, the therapeutic efficacy of TLR agonist monotherapy in cancer treatment has been limited, and the mechanisms of failure remain unknown. We demonstrate that topical treatment with a TLR-7 agonist, imiquimod, can elicit significant regression of spontaneous breast cancers in neu transgenic mice, a model of human HER-2/neu(+) breast cancer. However, tumor growth progressed once imiquimod therapy was ended. Gene expression analysis using tumor-derived RNA demonstrated that imiquimod induced high levels of IL-10 in addition to TNF-alpha and IFN-gamma. Elevated levels of circulating IL-10 were also detected in sera from imiquimod-treated mice. Elevated serum IL-10 appeared to be derived from IL-10 and dual cytokine secreting (IFN-gamma(+) and IL-10(+)) CD4(+) T cells rather than CD4(+)CD25(+)Foxp3(+) T regulatory cells, which were also induced by imiquimod treatment. Blockade of IL-10, but not TGF-beta, enhanced the antitumor effect of imiquimod by significantly prolonging survival in treated mice. These data suggest that the excessive inflammation induced by TLR agonists may result in a self-regulatory immunosuppression via IL-10 induction and that blocking IL-10 could enhance the therapeutic efficacy of these agents.