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MP90-02 CARBOHYDRATE RESTRICTION OPPOSES TUMOR-PROMOTING EFFECTS OF OBESITY IN THE HI-MYC TRANSGENIC MOUSE MODEL OF PROSTATE CANCER.
THE JOURNAL OF UROLOGYâ
Vol. 195, No. 4S, Supplement, Tuesday, May 10, 2016
synthesis, in prostate cancer progression. We determined the impact of modifying serum cholesterol levels on tumor progression in PTENdeficient transgenic mouse model of prostate cancer. METHODS: PTENloxP/loxP-Cre+ mice consuming ad libitum high fat, high cholesterol diets (40% fat, 1.25% cholesterol) were randomized after weaning to receive the cholesterol uptake inhibitor, ezetimibe (Zetia; 30 mg/kg diet), or no intervention, and sacrificed at 2, 3, or 4 months of age. At sacrifice, prostates were removed, weighed, and fixed in formalin. Serum cholesterol and testosterone levels were measured by ELISA and prostate androgens measured by mass spectrometry. Proliferation and apoptosis in tumor epithelium and stroma was assessed by Ki67 and TUNEL assay, respectively. RESULTS: Mice randomized to Zetia had lower serum cholesterol levels (p¼0.031). Cholesterol levels were positively correlated with prostate weight (p¼0.033) and tumor epithelial proliferation (p¼0.069), and negatively correlated with tumor epithelial apoptosis (p¼0.004). Tumor stromal cell proliferation was reduced in the Zetia group (p¼0.010). Increased cholesterol levels were associated with elevated intraprostatic DHEA, testosterone and androstendione (p¼0.043, p¼0.074, p¼0.031, respectively) but was unrelated to serum androgens (p¼0.08). Mice consuming Zetia diet had lower rates of intracystic adenocarcinoma at 2 and 3 months of age (0% vs. 14% and 30% vs. 44%, respectively). CONCLUSIONS: Using a PTEN-deficient transgenic mouse model of prostate cancer, we found that serum cholesterol reduction lowered intraprostatic androgen concentrations and slowed prostate cancer progression, supporting a potentially important role for cholesterol in prostate cancer. This study may provide new insight into the use of cholesterol-lowering interventions for prostate cancer prevention and treatment. Source of Funding: 1-R01-CA131235-01A1, 1K24CA160653
MP90-02 CARBOHYDRATE RESTRICTION OPPOSES TUMOR-PROMOTING EFFECTS OF OBESITY IN THE HI-MYC TRANSGENIC MOUSE MODEL OF PROSTATE CANCER. Emma Allott, Chapel Hill, NC; Everardo Macias, Los Angeles, CA; George Thomas, Portland, OR; Stephen Hursting, Chapel Hill, NC; Stephen Freedland*, Los Angeles, CA INTRODUCTION AND OBJECTIVES: Previously, we showed carbohydrate restriction slowed tumor growth in xenograft mouse prostate cancer models. Herein, we examined the impact of carbohydrate restriction on tumor progression within the context of dietinduced obesity in the Hi-Myc transgenic mouse model of prostate cancer. METHODS: At weaning, Hi-myc male transgenic mice were randomized to Western diet (WD; 40% fat, 42% carbohydrate; n¼39) or no carbohydrate ketogenic diet (NCKD; 82% fat, 1% carbohydrate; n¼44) for 3 or 6 months. At sacrifice, prostates were weighed and prostate histology, proliferation and apoptosis were evaluated by H&E, Ki67 and TUNEL, respectively. Body composition and bone mineral content were assessed by DEXA, serum cytokines measured using multiplex, and Akt/mTOR signaling assessed by Western blotting. RESULTS: Caloric intake was higher in the NCKD group, resulting in elevated body weights at 6 months of age (45.3g vs. 38.9g; p¼0.008). While mice randomized to NCKD had higher rates of adenocarcinoma at 6 months (100% vs. 80%, p¼0.04), they had significantly smaller prostates at both 3 and 6 months, following adjustment for body weight (p¼0.004 and p¼0.002, respectively). Despite elevated body weights, serum MCP-1 and IL-1a levels were lower in the NCKD group (p¼0.046 and p¼0.118, respectively) and relative Akt phosphorylation and phospho-S6 ribosomal protein levels were reduced in prostates of NCKD mice.
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CONCLUSIONS: Despite higher caloric intake and elevated body weights, carbohydrate restriction reduced prostate weight, lowered MCP-1 levels, and downstream markers of mTOR/Akt activity. Together, these data suggest that carbohydrate restriction may reduce systemic inflammation and oppose the tumor-promoting effects of obesity, resulting in slower prostate cancer growth, possibility through reduced mTOR/Akt activity. Source of Funding: 1-R01-CA131235-01A1, 1K24CA160653
MP90-03 REACTIVE OXYGEN SPECIES INDUCTION BY CABAZITAXEL VIA INHIBITION OF THE SESTRIN FAMILY IN CASTRATIONRESISTANT PROSTATE CANCER Takeo Kosaka*, Hiroshi Hongo, Yasumasa Miyazaki, Eiji Kikuchi, Akira Miyajima, Mototsugu Oya, Tokyo, Japan INTRODUCTION AND OBJECTIVES: Cabazitaxel (CBZ) has become the most effective cytotoxic agent to demonstrate an improvement in survival in men with docetaxel-refractory CRPC. Accumulating evidence has suggested reactive oxygen species (ROS) production induced by taxanes in cancer cells may influence taxaneinduced cell death or drug resistance. However, ROS production by CBZ or its mechanistic action remains to be characterized in CRPC. In this study, we investigated PCa cell sensitivity to docetaxel (DTX) or CBZ by measurement of ROS production. METHODS: Three cell lines were used: LNCaP, a human PCa cell line that exhibits androgen-dependent proliferation; C4-2, a human PCa cell line that exhibits androgen-independent proliferation; and C42AT6, a CRPC cell line derived from C4-2. These cells were treated with increasing concentrations of docetaxel or CBZ in vitro. Gene expression profiles of CBZ- or DTX-treated cells were analyzed using a whole human genome microarray. FACS analysis was performed using a fluorogenic marker of ROS. Furthermore, we investigated the changes in expression of antioxidant enzymes induced by DTX and CBZ. RESULTS: Cabazitaxel showed significantly higher cytotoxicity than docetaxel in human CRPC cells. FACS analysis revealed that treatment with cabazitaxel significantly increased ROS generation than docetaxel in C4-2AT6 cells, accompanied by higher expression of phospho-p38. Microarray analysis revealed significant changes in ROS regulation-related genes by CBZ. qPCR analysis revealed CBZ induced C4-2AT6 cell death, accompanied by significantly decreased expression of Sestrin-3 (SESN3). To investigate whether CBZ-mediated cell death was caused by induction of ROS generation, C4-2AT6 cells were incubated with CBZ in the presence of the antioxidant N-acetylcysteine (NAC), which was observed to reduce cabazitaxel-induced cytotoxicity. Microarray analysis revealed dynamic changes in ROS regulation-related genes by CBZ, and qPCR analysis demonstrated CBZ reduced mRNA expression of SESN3. We observed that ROS reduction by SESN3 was significantly inhibited by cabazitaxel in a dose-dependent manner, but not by docetaxel. When C4-2AT6 cells were treated with cabazitaxel in the presence of siRNAs against SESN3, significant cytotoxicity was observed, accompanied by enhanced ROS production. CONCLUSIONS: Our results indicated that the higher sensitivity of human CRPC to CBZ, compared with DTX, involves ROS production via inhibition of SESN3 expression, an antioxidant enzyme. Source of Funding: This work was supported in part by a Grant-in-Aid for Young Scientists from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.
Vol. 195, No. 4S, Supplement, Tuesday, May 10, 2016
synthesis, in prostate cancer progression. We determined the impact of modifying serum cholesterol levels on tumor progression in PTENdeficient transgenic mouse model of prostate cancer. METHODS: PTENloxP/loxP-Cre+ mice consuming ad libitum high fat, high cholesterol diets (40% fat, 1.25% cholesterol) were randomized after weaning to receive the cholesterol uptake inhibitor, ezetimibe (Zetia; 30 mg/kg diet), or no intervention, and sacrificed at 2, 3, or 4 months of age. At sacrifice, prostates were removed, weighed, and fixed in formalin. Serum cholesterol and testosterone levels were measured by ELISA and prostate androgens measured by mass spectrometry. Proliferation and apoptosis in tumor epithelium and stroma was assessed by Ki67 and TUNEL assay, respectively. RESULTS: Mice randomized to Zetia had lower serum cholesterol levels (p¼0.031). Cholesterol levels were positively correlated with prostate weight (p¼0.033) and tumor epithelial proliferation (p¼0.069), and negatively correlated with tumor epithelial apoptosis (p¼0.004). Tumor stromal cell proliferation was reduced in the Zetia group (p¼0.010). Increased cholesterol levels were associated with elevated intraprostatic DHEA, testosterone and androstendione (p¼0.043, p¼0.074, p¼0.031, respectively) but was unrelated to serum androgens (p¼0.08). Mice consuming Zetia diet had lower rates of intracystic adenocarcinoma at 2 and 3 months of age (0% vs. 14% and 30% vs. 44%, respectively). CONCLUSIONS: Using a PTEN-deficient transgenic mouse model of prostate cancer, we found that serum cholesterol reduction lowered intraprostatic androgen concentrations and slowed prostate cancer progression, supporting a potentially important role for cholesterol in prostate cancer. This study may provide new insight into the use of cholesterol-lowering interventions for prostate cancer prevention and treatment. Source of Funding: 1-R01-CA131235-01A1, 1K24CA160653
MP90-02 CARBOHYDRATE RESTRICTION OPPOSES TUMOR-PROMOTING EFFECTS OF OBESITY IN THE HI-MYC TRANSGENIC MOUSE MODEL OF PROSTATE CANCER. Emma Allott, Chapel Hill, NC; Everardo Macias, Los Angeles, CA; George Thomas, Portland, OR; Stephen Hursting, Chapel Hill, NC; Stephen Freedland*, Los Angeles, CA INTRODUCTION AND OBJECTIVES: Previously, we showed carbohydrate restriction slowed tumor growth in xenograft mouse prostate cancer models. Herein, we examined the impact of carbohydrate restriction on tumor progression within the context of dietinduced obesity in the Hi-Myc transgenic mouse model of prostate cancer. METHODS: At weaning, Hi-myc male transgenic mice were randomized to Western diet (WD; 40% fat, 42% carbohydrate; n¼39) or no carbohydrate ketogenic diet (NCKD; 82% fat, 1% carbohydrate; n¼44) for 3 or 6 months. At sacrifice, prostates were weighed and prostate histology, proliferation and apoptosis were evaluated by H&E, Ki67 and TUNEL, respectively. Body composition and bone mineral content were assessed by DEXA, serum cytokines measured using multiplex, and Akt/mTOR signaling assessed by Western blotting. RESULTS: Caloric intake was higher in the NCKD group, resulting in elevated body weights at 6 months of age (45.3g vs. 38.9g; p¼0.008). While mice randomized to NCKD had higher rates of adenocarcinoma at 6 months (100% vs. 80%, p¼0.04), they had significantly smaller prostates at both 3 and 6 months, following adjustment for body weight (p¼0.004 and p¼0.002, respectively). Despite elevated body weights, serum MCP-1 and IL-1a levels were lower in the NCKD group (p¼0.046 and p¼0.118, respectively) and relative Akt phosphorylation and phospho-S6 ribosomal protein levels were reduced in prostates of NCKD mice.
e1145
CONCLUSIONS: Despite higher caloric intake and elevated body weights, carbohydrate restriction reduced prostate weight, lowered MCP-1 levels, and downstream markers of mTOR/Akt activity. Together, these data suggest that carbohydrate restriction may reduce systemic inflammation and oppose the tumor-promoting effects of obesity, resulting in slower prostate cancer growth, possibility through reduced mTOR/Akt activity. Source of Funding: 1-R01-CA131235-01A1, 1K24CA160653
MP90-03 REACTIVE OXYGEN SPECIES INDUCTION BY CABAZITAXEL VIA INHIBITION OF THE SESTRIN FAMILY IN CASTRATIONRESISTANT PROSTATE CANCER Takeo Kosaka*, Hiroshi Hongo, Yasumasa Miyazaki, Eiji Kikuchi, Akira Miyajima, Mototsugu Oya, Tokyo, Japan INTRODUCTION AND OBJECTIVES: Cabazitaxel (CBZ) has become the most effective cytotoxic agent to demonstrate an improvement in survival in men with docetaxel-refractory CRPC. Accumulating evidence has suggested reactive oxygen species (ROS) production induced by taxanes in cancer cells may influence taxaneinduced cell death or drug resistance. However, ROS production by CBZ or its mechanistic action remains to be characterized in CRPC. In this study, we investigated PCa cell sensitivity to docetaxel (DTX) or CBZ by measurement of ROS production. METHODS: Three cell lines were used: LNCaP, a human PCa cell line that exhibits androgen-dependent proliferation; C4-2, a human PCa cell line that exhibits androgen-independent proliferation; and C42AT6, a CRPC cell line derived from C4-2. These cells were treated with increasing concentrations of docetaxel or CBZ in vitro. Gene expression profiles of CBZ- or DTX-treated cells were analyzed using a whole human genome microarray. FACS analysis was performed using a fluorogenic marker of ROS. Furthermore, we investigated the changes in expression of antioxidant enzymes induced by DTX and CBZ. RESULTS: Cabazitaxel showed significantly higher cytotoxicity than docetaxel in human CRPC cells. FACS analysis revealed that treatment with cabazitaxel significantly increased ROS generation than docetaxel in C4-2AT6 cells, accompanied by higher expression of phospho-p38. Microarray analysis revealed significant changes in ROS regulation-related genes by CBZ. qPCR analysis revealed CBZ induced C4-2AT6 cell death, accompanied by significantly decreased expression of Sestrin-3 (SESN3). To investigate whether CBZ-mediated cell death was caused by induction of ROS generation, C4-2AT6 cells were incubated with CBZ in the presence of the antioxidant N-acetylcysteine (NAC), which was observed to reduce cabazitaxel-induced cytotoxicity. Microarray analysis revealed dynamic changes in ROS regulation-related genes by CBZ, and qPCR analysis demonstrated CBZ reduced mRNA expression of SESN3. We observed that ROS reduction by SESN3 was significantly inhibited by cabazitaxel in a dose-dependent manner, but not by docetaxel. When C4-2AT6 cells were treated with cabazitaxel in the presence of siRNAs against SESN3, significant cytotoxicity was observed, accompanied by enhanced ROS production. CONCLUSIONS: Our results indicated that the higher sensitivity of human CRPC to CBZ, compared with DTX, involves ROS production via inhibition of SESN3 expression, an antioxidant enzyme. Source of Funding: This work was supported in part by a Grant-in-Aid for Young Scientists from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.