Cancer cells experience higher oxidative stress from reactive oxygen species (ROS) than non-malignant cells due to genetic alterations and abnormal growth and as a result, maintenance of the anti-oxidant glutathione (GSH) is essential for their survival and proliferation1–3. Under elevated ROS conditions endogenous l-Cysteine (l-Cys) production is insufficient for GSH synthesis, necessitating l-Cys uptake, predominantly in its disulfide form l-Cystine (CSSC) via the xCT(−) transporter. Here we show that administration of an engineered, pharmacologically optimized, human Cyst(e)inase enzyme mediates sustained depletion of the extracellular l-Cys and CSSC pool in mice and non-human primates, selectively causes cell cycle arrest and death (PI and Annexin-V staining) in cancer cells due to depletion of intracellular GSH and ensuing elevated ROS, yet results in no apparent toxicities in mice even after months of continuous treatment. Cyst(e)inase suppressed the growth of prostate carcinoma allografts, reduced tumor growth in prostate and breast cancer xenografts and doubled the median survival time of TCL1-Tg:p53−/− mice that develop disease resembling human chronic lymphocytic leukemia. The observation that enzyme-mediated depletion of the serum l-Cys and CSSC pool suppresses the growth of multiple tumors, yet is very well tolerated for prolonged periods suggests that Cyst(e)inase represents a safe and effective therapeutic modality for inactivating anti-oxidant cellular responses in a wide range of malignancies4,5.

Green tea is now recognized as the most effective cancer preventive beverage. In one study, 10 Japanese-size cups of green tea daily supplemented with tablets of green tea extract limited the recurrence of colorectal polyps in humans to 50%. Thus, cancer patients who consume green tea and take anticancer drugs will have double prevention. We studied the effects of combining ())-epigallocatechin gallate (EGCG) and anticancer drugs, focusing on inhibition of cell growth and induction of apoptosis. Numerous anticancer drugs, such as tamoxifen, COX-2 inhibitors, and retinoids were used for the experiments, and the combination of EGCG and COX-2 inhibitors consistently induced the enhancement of apoptosis. To study the mechanism of the enhancement, we paid special attention to the enhanced expressions of DDIT3 (growth arrest and DNA damage-inducible 153, GADD153), GADD45A, and CDKN1A (p21 ⁄ WAF1 ⁄ CIP1) genes, based on our previous evidence that a combination of EGCG and sulindac specifically induced upregulated expression of GADD153 and p21 genes in PC-9 lung cancer cells. The synergistic enhancements of apoptosis and GADD153 gene expression in human non-small cell lung cancer cells by the combination of EGCG and celecoxib were mediated through the activation of the MAPK signaling pathway. This article reviews the synergistic enhancement of apoptosis, gene expression, and anticancer effects using various combinations of EGCG and anticancer drugs, including the combination of ())-epicatechin (EC) and curcumin. Based on the evidence, we present a new concept: green tea catechins as synergists with anticancer drugs. (Cancer Sci 2011; 102: 317-323) ''C ancer chemoprevention'' was defined in 1976,(1) and Michael Sporn (2) introduced the term ''combination cancer chemoprevention'' in the journal Nature in 1980. He defined the term as the combined use of several drugs with different mechanisms of action exerting marked synergistic preventive effects. In 1983, we began to study the cancer preventive activity of green tea catechins.(3,4) Green tea extract chemically contains at least four tea catechins: EGCG, ECG, EGC, and EC. The first three catechins induce cancer preventive activities, whereas EC is inactive.(5) We first reported that combinations with the active catechins and inactive EC induced synergistic effects on induction of apoptosis and inhibition of cell growth of human lung cancer cell line PC-9, and inhibition of TNF-a release from BALB ⁄ 3T3 cells treated with okadaic acid, a tumor promoter.(6) This suggests that whole green tea, which is a mixture of green tea catechins, is a more effective and practical cancer preventive than green tea catechins alone.(6) Using tritium ( 3 H)-EGCG we showed that EC induced the enhanced incorporation of 3 H-EGCG and other active green tea catechins into cells.(6) In addition, we recently showed that drinking 10 cups of green tea supplemented with green tea tablets significantly (50%) prevented recurrence of colorectal adenomas in patients who had received polypectomy 1 yea...

Male Hi-Myc mice were placed on three dietary regimens [30% calorie restriction (CR), overweight control (modified AIN76A with 10kcal% fat), and a diet-induced obesity regimen (DIO) 60kcal% fat]. All diet groups had approximately similar incidence of hyperplasia and low grade PIN in the ventral prostate at 3 and 6 months of age. However, 30% CR significantly reduced the incidence of in situ adenocarcinomas at 3 months compared to the DIO group and at 6 months compared to both the overweight control and DIO groups. Furthermore, the DIO regimen significantly increased the incidence of adenocarcinoma with aggressive stromal invasion, as compared to the overweight control group (96% vs. 65% respectively, p=0.02) at the 6 month time point. Additionally, at both 3 and 6 months, only in situ carcinomas were observed in mice maintained on the 30% CR diet. Relative to overweight control, DIO increased, while 30% CR reduced activation of Akt, mTORC1, Stat3 and NFκB (p65) in ventral prostate. DIO also significantly increased (and 30% CR decreased) numbers of T-lymphocytes and macrophages in the ventral prostate compared to overweight control. The mRNA levels for IL1α, IL1β, IL6, IL7, IL23, IL27, NFκB1 (p50), TNFα and VEGF family members were significantly increased in the ventral prostate of the DIO group compared to both the overweight control and 30% CR diet groups. Collectively, these findings suggest that enhanced growth factor (Akt/mTORC1 and Stat3) and inflammatory (NFκB, cytokines) signaling may play a role in dietary energy balance effects on prostate cancer progression in Hi-Myc mice.

Despite much recent progress, prostate cancer continues to represent a major cause of cancer-related mortality and morbidity in men. Prostate cancer is the most common nonskin neoplasm and second leading cause of death in men. 6-Shogaol (6-SHO), a potent bioactive compound in ginger (Zingiber officinale Roscoe), has been shown to possess anti-inflammatory and anticancer activity. In the present study, the effect of 6-SHO on the growth of prostate cancer cells was investigated. 6-SHO effectively reduced survival and induced apoptosis of cultured human (LNCaP, DU145, and PC3) and mouse (HMVP2) prostate cancer cells. Mechanistic studies revealed that 6-SHO reduced constitutive and interleukin (IL)-6-induced STAT3 activation and inhibited both constitutive and TNF-a-induced NF-kB activity in these cells. In addition, 6-SHO decreased the level of several STAT3 and NF-kB-regulated target genes at the protein level, including cyclin D1, survivin, and cMyc and modulated mRNA levels of chemokine, cytokine, cell cycle, and apoptosis regulatory genes (IL-7, CCL5, BAX, BCL2, p21, and p27). 6-SHO was more effective than two other compounds found in ginger, 6-gingerol, and 6-paradol at reducing survival of prostate cancer cells and reducing STAT3 and NF-kB signaling. 6-SHO also showed significant tumor growth inhibitory activity in an allograft model using HMVP2 cells. Overall, the current results suggest that 6-SHO may have potential as a chemopreventive and/or therapeutic agent for prostate cancer and that further study of this compound is warranted. Cancer Prev Res; 7(6); 627-38. Ó2014 AACR.

High-throughput screening of a natural compound library was performed to identify the most efficacious combinatorial treatment on prostate cancer. Ursolic acid, curcumin and resveratrol were selected for further analyses and administered in vivo via the diet, either alone or in combination, in a mouse allograft model of prostate cancer. All possible combinations of these natural compounds produced synergistic effects on tumor size and weight, as predicted in the screens. A subsequent untargeted metabolomics and metabolic flux analysis using isotopically labeled glutamine indicated that the compound combinations modulated glutamine metabolism. In addition, ASCT2 levels and STAT3, mTORC1 and AMPK activity were modulated to a greater extent by the combinations compared to the individual compounds. Overall, this approach can be useful for identifying synergistic combinations of natural compounds for chemopreventive and therapeutic interventions.

Obesity is a prognostic risk factor in the progression of prostate cancer (PCa), however, the molecular mechanisms involved are unclear. In this study, we provide preclinical proof of concept for the role of a pro-inflammatory CXCL12-CXCR4/CXCR7 signaling axis in an obesity-driven mouse model of HiMyc prostate cancer. Analysis of the stromal vascular fraction (SVF) from periprostatic white adipose tissue (ppWAT) from obese HiMyc mice at 6 months of age revealed a dramatic increase in mRNAs encoding various chemokines, cytokines, growth factors and angiogenesis mediators, with CXCL12 among the most significantly upregulated genes. Immunofluorescence staining of ventral prostate tissue from obese HiMyc mice revealed high levels of CXCL12 in the stromal compartment as well as high staining for CXCR4 and CXCR7 in the epithelial compartment of tumors. PCa cell lines derived from HiMyc tumors (HMVP2 and derivative cell lines) displayed increased protein expression of both CXCR4 and CXCR7 compared to protein lysates from a non-tumorigenic prostate epithelial cell line (NMVP cells). CXCL12 treatment stimulated migration and invasion of HMVP2 cells but not NMVP cells. These effects of CXCL12 on HMVP2 cells were inhibited by the CXCR4 antagonist AMD3100 as well as knockdown of either CXCR4 or CXCR7. CXCL12 treatment also produced rapid activation of STAT3, NFkB, and MAPK signaling in HMVP2 cells which was again attenuated by either AMD3100 or knockdown of CXCR4 or CXCR7. Collectively, these data suggest that CXCL12 secreted by stromal cells activates invasiveness of PCa cells and may play a role in driving tumor progression in obesity. Targeting the CXCL12-CXCR4/CXCR7 axis could lead to novel approaches for offsetting the effects of obesity on PCa progression.

Fat tissue, overgrowing in obesity, promotes the progression of various carcinomas. Clinical and animal model studies indicate that adipose stromal cells (ASC), the progenitors of adipocytes, are recruited by tumors and promote tumor growth as tumor stromal cells. Here, we investigated the role of ASC in cancer chemoresistance and invasiveness, the attributes of tumor aggressiveness. By using human cell co-culture models, we demonstrate that ASC induce epithelial-mesenchymal transition (EMT) in prostate cancer cells. Our results for the first time demonstrate that ASC interaction renders cancer cells more migratory and resistant to docetaxel, cabazitaxel, and cisplatin chemotherapy. To confirm these findings in vivo, we compared cancer aggressiveness in lean and obese mice grafted with prostate tumors. We show that obesity promotes EMT in cancer cells and tumor invasion into the surrounding fat tissue. A hunter-killer peptide D-CAN, previously developed for targeted ASC ablation, suppressed the obesity-associated EMT and cancer progression. Importantly, cisplatin combined with D-CAN was more effective than cisplatin alone in suppressing growth of mouse prostate cancer allografts and xenografts even in non-obese mice. Our data demonstrate that ASC promote tumor aggressiveness and identify them as a target of combination cancer therapy.

Perturbing redox homeostasis potentially constitutes a selective cancer-killing strategy. An engineered human enzyme, cyst(e)inase that degrades extracellular cysteine ( l -Cys) and cystine (CSSC) leading to depletion of intracellular l -Cys and glutathione (GSH) was evaluated for its effects on pancreatic cancer cell lines. Cyst(e)inase caused oxidative stress and apoptosis in only Panc1 cells, whereas MIA-PaCa2 and BxPC3 cells demonstrated survival under conditions of cyst(e)inase-mediated l -Cys depletion through maintenance of mitochondrial metabolism and lower levels of reactive oxygen species (ROS). A correlation was also observed between thioredoxin 1 protein levels and resistance to cyst(e)inase treatment. Notably, cyst(e)inase in combination with auranofin, a thioredoxin reductase inhibitor, caused a synergistic increase in mitochondrial ROS and apoptosis and inhibition of mitophagy in the more resistant cells. In addition, auranofin treatment sensitized the more resistant pancreatic cancer xenografts to cyst(e)inase without systemic toxicity. These data provide strong rationale to further investigate therapeutic strategies that target multiple antioxidant pathways for treatment of pancreatic ductal adenocarcinoma.