Chemoprevention of colorectal cancer: a systematic review in rats.
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Studies of colonic tumors & cancers in rats,
not yet linked to Medline (e.g., abstracts from congress)
Bi S, Geng BQ, and Yong DG: Effects of menadione on
1,2-dimethylhydrazine-induced mouse colon adenocarcinoma. Acta
Pharmacologica Sinica 14, 17-20, 1993.
- Bull A, Bird RP, Bruce WR, Nigro N, and Medline
A: Effect of calcium on azoxymethane induced intestinal tumors in rats.
Gastroenterology, 92, abstract #1332 (abstr), 1987.
EFFECT OF CALCIUM ON AZOXYMETHANE INDUCED INTESTINAL TUMORS IN RATS.
A.Bull, R.P.Bird, W.R.Bruce, N.Nigro, A.Medline,
Ludwig Inst. for Cancer Res., Toronto Branch, Toronto, Canada and
Wayne State Univ.Sch. of Medicine, Detroit, MI.
Two experimental protocols were used to evaluate the effect of dietary calcium on colon tumorigenesis. In the first (Study A) 120 male Sprague-Dawley rats weighing 100-125 g were divided into 4 dietary groups of 30 rats each. The diets were as follows: Calcium (Ca HP04) 0.1% or 1.0% of the diet was added to either a low fat (2% beef fat + 1% corn oil) diet or a high fat (29% beef fat + 1% corn oil) diet. The Ca:P was 1:1 in all diets. 25 rats in each group were given weekly sc injections of azoxymethane 8 mg/kg for 8 weeks. The other five rats in each group served as non carcinogen controls. Rats were killed and intestinal tumors tabulated 26 weeks after the first carcinogen injection.
In the second study (Study B) 160 F344 female rats received weekly injections of azoxymethane (sc 15 mg/kg for 2 weeks). One week later they were randomized into the 4 dietary groups mentioned above. Each group contained 40 treated and 10 control animals. Nine months after the first injection with carcinogen treatment animals were killed and examined for tumor incidence. Results for the large bowel tumors (average no of tumors/rat) are shown in the table:
-------------- 3% Fat ------------------ ------ 30% Fat ----------
-------- 0.1% Ca -- 1.0% Ca ---- ---- 0.1% Ca -- 1.0% Ca
Study A : 2.9 ------ 3.5 ----------- ------- 3.3 --------- 5.7
Study B : 1.4 ------ 1.7 ----------- ------- 2.0 --------- 2.7
Total tumor yield irrespective of the size and type was enhanced by both high levels of calcium and high levels of fat in the diet. Consequently, the results of this animal experiment do not support the hypothesis that supplemental calcium inhibits intestinal tumor formation.
Corpet D.E. and Parnaud G. Colon Cancer Prevention by Dietary PEG in Rats.
Special Conference and Workshops on Colon Cancer. May 10-13, 2000, McGill Cancer Centre, McGill University, Montreal, Canada.
Background and Methods: Dietary changes could prevent 70-80% of colorectal cancers. Bulking fibers and a high water intake may decrease colon carcinogenesis. We speculated that a non-fermented polymer that increases stools moisture, the osmotic laxative polyethylene-glycol 8000 (PEG), might protect rats against colon carcinogenesis. This hypothesis was tested in 16 independent studies, involving more than 600 rats and mice. The general design was to initiate carcinogenesis in F344 rats by a single injection of azoxymethane (20mg/kg), and to randomize the animals 7 d later to diets or drinking water containing 5% PEG or no PEG (control). Most studies lasted for 30 d with the aberrant crypt foci (ACF) endpoint. Macroscopic tumors were looked for in 2 long-term studies.
ACF Results: All our studies showed a highly significant preventive effect of dietary PEG. In the first study, PEG-fed rats had 100 times fewer large ACF than controls and 20 times fewer total ACF than control (both p<0.0001). Two treated rats had no detectable ACF, a protection never observed before. PEG was much more potent than any of the 80 agents tested before in the ACF model (54 publications). Next studies showed that a 3 d PEG treatment halves the number of ACF in rats, but the inhibition was reversible in part when treatment was discontinued. PEG caused the regression of established ACF. The protection was time- and dose-dependant. PEG was active in both male and female F344 rats, also in SD rats and OF1 mice. ACF initiated by various carcinogens (AOM, MNU and MeIQ) were suppressed by PEG. PEG with MW between 3350 and 12000 (but not PEG 400), and PEG 8000 from five suppliers were active. The prevention was stronger in rats fed a high-fat diet (p < 0.0001) than in rats fed a rodent chow (p=0.02). PEG was active when given via diet or drinking water, but not when injected i.p., or when given only during the initiation phase.
Cancer Results: In an accelerated model of carcinogenesis, rats were fed a high-fat diet, based on cooked casein to promote tumor growth. Dietary PEG decreased the incidence of macroscopic tumors from 70 to 10% (p=0.005), and the multiplicity from 2.1 to 0.1 tumor/rat (p=0.003). No cancer was detected in PEG-fed rats. In the standard rat model also, dietary PEG markedly decreased the incidence of colon cancers from 19/27 to 2/21 (p < 0.0001), and the multiplicity from 3.1 to 0.3 tumor/rat (p<0.0001). Among 60 chemopreventive agents, only celecoxib is as potent as PEG.
Mechanism: The mechanism of PEG effect is not known. We speculate that PEG may act by 3 ways: It bulks, lubricates, and pumps. Bulk: PEG doubled fecal weight and moisture, which markedly diluted promoting bile acids and reduced cytotoxicity of fecal water. Lubricant: PEG, like mucin, can protect epithelia from the fecal stream, and can help the resealing of wounded cells membrane. Pump: HT29 cancer cells were blocked in G0/G1 phase by PEG osmotic pressure. PEG increased the osmotic pressure in the gut of rats, which may block mutated cells, and erase them from the mucosa.
Conclusion: PEG is thus a fast, consistent, and very potent inhibitor of early precursor lesions, and of colon cancer in rats. PEG has no known toxicity in humans. It is on the "generally recognized as safe" list. We think it could be tested in a clinical trial in humans.
- Harris GK, Carlton PS, Liston BW, Gupta AK, and
Stoner GD: Inhibitory effects of freese-dried black raspberries on AOM induced
colon cancer in the F344 rat. Proc. Am. Assoc. Cancer Res, 42, #948 (abstr),
Kim H.S., Jeong-Sang Lee, Tae-Young Oh, Marie Yeo, Ki-Baik Hahm, Young-Joon Surh
Chemopreventive effects of curcumin on azoxymethane-initiated and dextran sulfate sodium-promoted mouse colon carcinogenesis
Proc Amer Assoc Cancer Res 2006;47:[Abstract #2275].
Multiple lines of compelling evidence support the casual relationship between inflammation and cancer. Dextran sulfate sodium (DSS) administration has been reported to induce chronic inflammation in mouse colonic mucosa, which markedly enhances the incidence and the burden of polyps. In an attempt to corroborate the contribution of ulcerative colitis to colon carcinogenesis, we treated male ICR mice with a single i.p dose (10mg/kg body weight) of AOM followed by 2% DSS in drinking water for consecutive 7 days. At the 16th week of AOM plus DSS treatment, 83.33% of mice developed tumors, mostly in rectal and descending colon, which were verified as the polypoid tumors and adenocarcinomas by histopathological examination. In contrast, only 27.27% of AOM-treated and none of DSS-treated mice had tumors. Cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) were markedly up-regulated in the AOM-initiated and DSS-promoted colon tumors. DNA binding activities of nuclear factor-kappaB (NF-kB) and activator protein 1 (AP-1) were also elevated in the colon tumors. In the same model, we examined the chemopreventive effects of curcumin. Oral administration of curcumin (0.1 or 0.25 mmole/kg body wt.) for 14 weeks after initiation with AOM significantly reduced the tumor incidence and the multiplicity (83.33% and average 6.00 tumors/mouse in the AOM plus DSS group; 50.14% and 4.25 tumors/mouse in the low dose curcumin group; 42.86% and 4.00 tumors/mouse in the high dose curcumin group). Curcumin treatment significantly inhibited the expressions of COX-2 and iNOS as well activitation of NF-kB and AP-1 in the colonic tissues. Likewise curcumin reduced activities as well as mRNA expression of several matrix metalloproteinases (MMPs), such as MMP2, MMP9, MMP13, TIMP1, MT1-MMP and MT2-MMP in the colon. Differentially expressed gene analysis revealed the upregulation of several genes including heterogeneous nuclear ribonucleoprotein M (Hnrpm), deleted in malignant brain tumors 1 (DMBT1), interferon induced transmembrane protein 1 (Ifitm 1), in colon tumors compared with the normal tissue. Curcumin treatment inhibited the expressions of these genes. Functional significance of above genes that are upregulated in colitis-associated carcinogenesis is under investigation
Kohno, H., Rikako Suzuki, Yumiko Yasui, Masashi Hosokawa, Kazuo Miyashita, Shigeyuki Sugie, Takuji Tanaka.
Dietary pomegranate seed oil rich in conjugated linolenic acid increases colonic PPAR? expression and decreases malignancy induced by azoxymethane in rats.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 4032
Kanazawa Medical University, Ishikawa, Japan and Hokkaido University, Hakodate, Japan.
Conjugated linolenic acid (CLN) present in some seed oils is a mixture of positional and geometric isomers of the 18:3 fatty acids that have conjugated trienoic double bonds. It is known that seed oil from pomegranate (Punica granatum L.) (PGO) contains more than 80% 9cis(c), 11trans(t), 13c-18:3 as CLN. Our previous short-term experiment demonstrated that seed oil from bitter melon (Momordica charantia) (BMO) rich in 9c, 11t, 13t-CLN, inhibited the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF). In the present study, the possible inhibitory effect of dietary administration of PGO on the development of colonic neoplasms was investigated using an animal colon carcinogenesis model initiated with a colon carcinogen AOM. Male F344 rats were given subcutaneous injections of AOM (20 mg/kg body weight) once a week for 2 weeks to induce colon neoplasms. They also received diets containing 0.01%, 0.1%, 1% PGO or 1% conjugated linoleic acid (CLA) for 32 weeks, starting one week before the first dosing of AOM. At the termination of the study (32 weeks), AOM induced 81% incidence (13/16 rats) of colonic adenocarcinoma. Dietary supplementation with PGO caused significant reduction in the incidence (46% inhibition by 0.01% PGO, P<0.05; 53% inhibition by 0.1% PGO, P<0.01; and 31% inhibition by 1% PGO) and the multiplicity (70% inhibition by 0.01% PGO, P<0.01; 73% inhibition by 0.1% PGO, P<0.005; and 53% inhibition by 1% PGO, P<0.05) of colonic adenocarcinoma, though a clear dose response was not observe. Such inhibition was associated with the increased content of CLA (c9,t11-18:2) in the lipid composition in colonic mucosa and liver. Also, PGO administration in diet enhanced expression of peroxisome proliferator-activated receptor (PPAR?) protein in the non-lesional colonic mucosa. These findings suggest that PGO rich in CLN can suppress AOM-induced colon carcinogenesis and the inhibition might be caused, in part, by modification of lipid composition in the colon and liver and/or increased expression of PPAR? protein level in the colon mucosa. Our findings also suggest that PGO could be applied to preclinical studies of the prevention of colon cancer.
Lee J.S., Hyun Soo Kim, Tae-Young Oh, Marie Yeo, Ki-Baik Hahm, Yung-Jue Bang, Young-Joon Surh.
Chemopreventive Effects of a Specific COX-2 Inhibitor (Celecoxib) and a Novel HDAC Inhibitor (SK-7041) on Azoxymethane-Initiated and Dextran Sulfate Sodium-Promoted Mouse Colon Carcinogenesis
Proc Amer Assoc Cancer Res 2005;46:[Abstract #2472].
Multiple lines of compelling evidence support the causal relationship between inflammation and cancer. In an attempt to clarify the contribution of ulcerative colitis to colon carcinogenesis, we treated male ICR mice with a single i.p dose (10 mg/kg body weight) of azoxymethane (AOM) followed by 2% dextran sulfate sodium salt (DSS) in drinking water for consecutive 7 days. DSS administration caused chronic inflammation, which markedly enhanced the incidence and the burden of AOM-initiated colon tumors. At the 16 week of AOM and DSS treatment, 80% of mice developed colorectal carcinoma, mostly in rectal and descending colon, whereas only 22% of AOM-treated and none of DSS-treated mice developed tumors. These findings suggest ulcerative colitis induced by DSS effectively promotes colon carcinogenesis in mice. Cyclooxygenase-2, inducible nitric oxide synthase, p16, and mutated p53 were markedly up-regulated in the AOM-initiated and DSS-promoted colon tumors. The serum levels of nitric oxide and proinflammatory cytokines, including interleukin (IL)-8, IL-1b, and IL-6, were also elevated in the colon tumors. In addition, the mRNA expression of matrix metalloproteinases (MMPs), such as MMP2, MMP9, MMP13, and MT2-MMP, was increased after AOM plus DSS treatment. On the contrary, the tissue inhibitor of MMP2 (TIMP2) was down-regulated by the same treatment. By utilizing the same mouse model, we evaluated chemopreventive activities of a selective inhibitor of COX-2 (Celecoxib) and a novel histone deacetylase (HDAC) inhibitor (SK-7041). Oral administration of Celecoxib (0.1 or 0.25 mmole/kg) by gavage for 14 weeks significantly lowered the AOM plus DSS-induced tumor incidence and multiplicity (75% and average 3.67 tumors/mouse in the AOM plus DSS group; 44.44% and 1.78/mouse in the low dose Celecoxib group; 40.0% and 1.00/mouse in the high dose Celecoxib group). When the HDAC inhibitor was administrated by i.p. injection 5 times at three-day intervals after AOM and DSS treatment, the tumor incidence and multiplicity were substantially reduced (80% and 3.70/mouse in the AOM plus DSS group; 60% and 2.20/mouse in the 10 mg/kg SK-7041 group; 55.83% and 1.53/mouse in the 30 mg/kg SK-7041). In conclusion, we have established a reliable and effective short-term two-stage mouse colon carcinogenesis model that can be utilized in chemoprevention studies.
National Research Laboratory of Molecular Carcinogenesis and Chemoprevention, College of Pharmacy, Seoul National University, Seoul, Republic of Korea, Genomic Research Center for Gastroenterology, School of Medicine, Ajou University, Suwon, Republic of Korea, National Research Laboratory for Cancer Epigenetics, Cancer Research Institute, College of Medicine, Seoul National University, Seoul, Republic of Korea
- Madar Z, Weiss O, Timar B, Gurevich P, and
Zusman I: The effects of high-fiber diets on chemically induced colon cancer in
rats. The Cancer Journal, 9, 207-211, 1996.
Martinez-Ferrer, Magaly, Martha Verghese, Lloyd Walker, Louis Shackelford.
Lycopene reduces incidence of azoxymethane induced colon tumors in Fisher 344 male rats during initiation and progression stages.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 1524
Vanderbilt University - School of Medicine, Nashville, TN and Alabama A&M University, Normal, AL.
Colorectal cancer is the third leading cause of cancer deaths in the United States. Several epidemiological studies have shown an inverse correlation between dietary intake of lycopene and the incidence of colon cancer. The objectives of this study were: (1) to determine the effect of dietary lycopene (400 ppm 7% fat and 400 ppm 14% fat) on chemically-induced colon tumorigenesis (end point tumor induction) in rats at tumor initiation and promotion stages, and (2) to determine the effect of dietary lycopene on the histopathology of tumors. Fisher 344 male weanling rats were divided into 8 groups of rats and were assigned to: AIN 93G +7% fat and AIN 93G +14% fat (controls), AIN 93G +7% fat+400ppm lycopene and AIN 93G +14% fat + 400ppm lycopene diets. Each lycopene diet was fed during initiation (I), promotion (P) and initiation plus promotion (I+P) stages of carcinogenesis. All the rats except saline controls received 16 mg/kg body weight of AOM at 7 and 8 wk of age. The rats were sacrificed at 46 wk of age. Tumor incidence (%) in colon of rats in the AIN 93G+7% fat and AIN 93G+14% fat were 87 and 100. Tumor incidence (%) in colon of rats in the AIN 93G+7% fat + 400ppm lycopene and AIN 93G+14% fat + 400 ppm lycopene (I, P, I+P) was 76, 60, 45, 86, 55, and 35, respectively. Colon tumors per tumor bearing rats (TBR) were 1.9, 4.4, 1.4, 1.7, 1, 1.8, 2.1, and 1.1 for the AIN 93G (7% and 14% fat), lycopene 400 ppm 7% and lycopene 400 ppm 14% (I, P, I+P), respectively. Histopathology of colon tumors was also compared among groups. Rats fed higher fat showed a higher incidence of developing adenocarcinomas. Dietary lycopene showed an effect in reducing total number of tumors as well as tumor size. This study provides evidence that dietary lycopene suppresses AOM induced colon tumorigenesis in Fisher 344 male rats, especially at the (P) and (I+P) stage, and therefore, colon tumorigenesis may be highly sensitive to dietary intervention
Venkat Swamy Malisetty, Jagan M.R. Patlolla, Jayadev Raju, Lee Ann Marcus, Chang-In Choi, Chinthalapally V Rao
Chemoprevention of colon cancer by diosgenin, a steroidal saponin constituent of fenugreek
Proc Amer Assoc Cancer Res 2005;46:[Abstract #2473].
Trigonella foenum graecum (fenugreek) is traditionally used to treat disorders such as high cholesterol, diabetes, wounds, inflammation, and gastrointestinal ailments. Recently we have shown that diosgenin a major constituent of fenugreek inhibit human colon cancer cell growth and induce apoptosis and suppress the carcinogen-induced colonic aberrant crypt foci in rats (CEBP, 13: 1-7, 2004). However, there are no studies indicating disogenin indeed inhibits carcinoma formation in established animal models of colon cancer. The present double-blind study was designed to assess the potential chemopreventive properties of diosgenin on azoxymethane (AOM)-induced rat colon carcinogenesis. Colonic adenocarcinomas were chosen as chemopreventive efficacy end point. In addition, we assessed the markers of proliferation and apoptosis in colonic tumors and normal appearing crypt to understand the mechanism of tumor growth inhibition of diosgenin. Male F344 rats at 7-weeks of age were fed the control (AIN-76A diet) and one week later, rats received s.c. injections of AOM (15 mg/kg body wt., once weekly for 2 weeks) or equal volume of normal saline (vehicle). One week after the carcinogen treatment groups of rats fed experimental diets containing 0 or 0.1% of diosgenin and continued on the experimental diets for 48 weeks and sacrificed. Colon tumors were evaluated histopathologically and expression levels of markers associated with proliferation and apoptosis were determined in colon tumors and normal appearing colon. Administration of diosgenin significantly suppressed both invasive and non-invasive colon tumor incidence up to 60% (p<0.0004). Similarly, colon tumor multiplicity (adenocarcinomas/rat) significantly suppressed (~68%, p<0.0001) in rats fed diosgenin diet. In colonic crypts and tumors of animals fed diosgenin there was a significant increase in BrdU labeling (p<0.005) and decrease in PCNA labeling (p<0.001) when compared to animals fed the control diet. The results of this study for the first time provide strong evidence that diosgenin, a naturally-occurring steroidal saponin of fenugreek, possess strong chemopreventive activity against colon carcinogenesis. These results pave the way further development of diosgenin for human clinical trials (Supported by R01 CA-80003 and R01 CA-94962 from the National Cancer Institute).
Natarajan, K., Bruce Cornish, Duane P. Matthees, Chandradhar Dwivedi.
Chemopreventive effects of dietary flaxseed oil on colon tumor development.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 3922
South Dakota State University, Brookings, SD.
Fatty acid composition of dietary fat, primarily the levels of ?-3 and ?-6 polyunsaturated fatty acids, has shown profound effect on colon tumor development in animal studies. Fats containing ?-6 fatty acids (e.g., corn oil) enhanced and ?-3 fatty acids (e.g., fish oil) reduced chemically-induced colon tumor in rats. The purpose of the current study was to investigate the effects of dietary flaxseed oil (containing a-linolenic acid, an ?-3 polyunsaturated fatty acid) on azoxymethane-induced colon tumor in rats and to compare with corn oil treated group.
Male Fischer rats, two groups of 30 each were assigned to the AIN-93M diet supplemented with 15% corn oil and 15% flaxseed oil respectively. Carcinogenesis was initiated with subcutaneous injections of azoxymethane (15mg/kg) once a week for three consecutive weeks. After 35 weeks of initiation, the rats were sacrificed under ether anesthesia. Blood was collected by cardiac puncture. The gastrointestinal tract was isolated and flushed with ice-cold normal saline. The site, size and number of tumors were recorded. The fatty acid composition in the serum, colon and tumors was determined using Gas Chromatography-Flame ionization detector (GC/FID).
Colon tumor incidence was found to be 100% and 54%; multiplicity was found to be 3.1 ± 0.8 and 0.7 ± 0.7 tumors/rat in corn and flaxseed oil treated groups respectively. Colon and serum samples of the corn oil group showed an increase in the ?-6 fatty acid levels whereas the flaxseed oil group exhibited an increase in the ?-3 fatty acid levels. The results indicate that dietary flaxseed oil containing high levels of ?-3 fatty acids is effective in preventing colon tumor development when compared to dietary corn oil containing ?-6 fatty acids in rats (Supported by EVERSCO LTD and SDEPSCoR).
- Osawa Emi, and Atsushi Nakajima, Hisahiko Sekihara, Hitoshi Nakagama,
PPAR gamma ligands suppress the formation of ACF and colon tumors induced by azoxymethane in mice
AACR 2002 AACR Annual Meeting, San Francisco Abstract Number: 377
Peroxisome Proliferator-Activated Receptorgamma (PPAR gamma) have been shown to inhibit cell growth for several types of malignancies. Recent studies, however, indicated that PPAR gamma ligands can promote colon carcinogenesis in APCMin mice. Therefore biological functions of PPAR gamma in colon carcinogenesis remain controversial. To clarify the role of PPAR gamma in multistage colon carcinogenesis, effects of PPAR gamma ligands on the induction of aberrant crypt foci (ACF), putative preneoplastic lesions of the colon, and tumor formation were investigated using an azoxymethane (AOM)-induced colon cancer model in BALB/c mice. After the administration of AOM, mice fed the basal diet without PPAR gamma ligand developed 14.8±2.1 ACFs per mice at week 5, and treatment with 200 ppm Pio, 200 ppm Rosi and 1000 ppm Tro significantly reduced the formation of ACF, those being 2.5±1.2 , 2.7±1.9, and 9.3±3.9, respectively (p<0.001). The total numbers of ACs per mouse were also suppressed significantly. Suppressive effect was observed in a dose-dependent manner within the range of 0.7 to 500 ppm of Rosi (separate experiment, data not shown). Histological analysis of a total of 40 ACF (10 ACF from each groups, respectively) revealed all lesions to be hyperplastic, without any B-catenin accumulation. Induction of colon tumors observed 32 weeks after six weekly administration of AOM (10 mg/kg bw) was also markedly suppressed by continuous feeding of 200 ppm Pio. The incidence was reduced to 20% (5/25), in contrast to 47% (14/30) without Pio-treatment (p<0.05). Regarding the sizes of tumors, only 38 % of tumors observed in the Pio-treated group were more than 2 mm in size, in contrast to 79 % of those in mice without Pio-treatment (p<0.05). The majority of lesions developing in both Pio-treated and untreated groups were diagnosed as adenocarcinomas. Colon tumors in both Pio-treated and untreated mice demonstrated no appreciable histological differences on HE staining or B-catenin immunostaining. In conclusion, this study clearly demonstrated the substantial suppressive effect of PPAR gamma on colon carcinogenesis without affecting the histological features of preneoplastic lesions or tumors, and that PPAR gamma ligands could be potential chemopreventive agents for colon carcinogenesis.
Rao C.V., Naveena B. Janakiram, Malisetty V. Swamy, Suresh Guruswamy, Jagan M. r. Patlolla, Ruchita Thungathurthi, Levy Kopelovich, Vernon E. Steele
Chemoprevention of colon cancer by raloxifene, a selective estrogen receptor modulator (SERM)
Proc Amer Assoc Cancer Res 2006;47:[Abstract #5745].
Recent studies suggest that estrogen receptor-b (ER-b) over-expression is positively associated with colon tumor development and evidences from estrogen replacement therapies also indicate a lowered risk of colon cancer. Therefore, in the present study we tested the chemopreventive potential of raloxifene (Evista), a selective estrogen receptor modulator (SERM) in a well-established model of colon cancer. Prior to the efficacy study, we assessed the maximum tolerated dose (MTD) and the dose-response effects of raloxifene on AOM-induced colonic aberrant crypt foci (ACF) in F344 rats. For MTD and ACF evaluation, seven week-old male F344 rats were fed the control diet (modified AIN-76A), or the experimental diets containing five different dose levels (0.31, 0.62, 1.25, 2.5 or 5 ppm) of raloxifene. One week later, all animals were s.c. injected with azoxymethane (AOM) (15 mg/kg body wt., once weekly for 2 weeks). At 15 weeks of age, all rats were killed and evaluated for MTD and colonic ACF. For colon tumor efficacy study, rats were fed diets containing 0 or 1 ppm raloxifene until termination, i.e., 48 weeks after carcinogen treatment. Colonic ACF and tumors were evaluated histopathologically. Expression levels of ER-b and markers associated with proliferation and apoptosis were determined in colon tumors and adjacent mucosa by Western blot and immunohistochemistry. Based on the body weight gain, the MTD of raloxifene in male F344 rats was found to be 5 ppm when administered in the AIN-76A diet. Administration of raloxifene at five different dose levels significantly inhibited AOM-induced total colonic ACF (31-36%, p<0.01) and multi-crypt (4 or more) aberrant foci (23-45%, p<0.05-0.01). Raloxifene at dose ranges of 0.31 -2.5 ppm did not show any dose response effect. Administration of raloxifene at the 1 ppm level significantly suppressed AOM-induced adenocarcinoma incidence by >36% (p<0.02). Similarly, administration of raloxifene suppressed AOM-induced colon adenocarcinoma multiplicity by 63% (p<0.001). AOM-induced colon adenocarcinomas showed significant up-regulation of ER-b and PCNA expression levels when compared to normal appearing colonic mucosa. Administration of raloxifene significantly suppressed AOM-induced colon tumor cell proliferation (PCNA-labeling, p<0.01), and an increase in apoptosis (TUNEL, p<0.05) compared to colon tumors of rats fed the control diet. The results of this study provide clear evidence that ER-b antagonist raloxifene prevents the AOM-induced colon carcinogenesis and its effect is associated with suppression of proliferation and induction of apoptosis in colon tumors. [Supported by NCI-CN-25114, NCI-CN-43300 and 1R01-CA-109247]
University of Oklahoma Cancer Institute, OUHSC, Oklahoma City, OK, Division of Cancer Prevention National Cancer Institute, Bethesda, MD
Rao C.V., Bandaru S. Reddy, Wang C-X, Jagan M.R Patlolla, Xaiaoping Liu, Nengtai Ouyang, Barbara Simi, Levy Kopelovich, Basil Rigas.
Chemoprevention of colon cancer by nitric oxide (NO)-releasing NSAIDs: strategies for developing safe and efficacious agents for colon cancer prevention and treatment
Proc Amer Assoc Cancer Res 2005;46:[Abstract #].
NSAIDs are promising colon cancer chemopreventive agents; however, their prolonged administration causes gastrointestinal toxicity. The rationale for the development of NO-NSAIDs is that they appear free of appreciable adverse effects, while they retain beneficial activities of their parent agents. Recent studies suggest that NO-releasing NSAIDs lack gastrointestinal toxicity and inhibit human colon cancer cell growth, carcinogen-induced colonic aberrant crypt foci in rats and intestinal polyp formation in Min mice. However, there are no studies indicating NO-NSAIDs indeed inhibit carcinoma formation in established animal models of colon cancer. The present double-blind study was designed to assess the potential chemopreventive properties of NO-fluorobiprofen (NCX 1026; NO-FLP), NO-indomethacin (NCX 530; NO-IND) and NO-aspirin (NCX 4016; NO-ASA) against azoxymethane (AOM)-induced colon cancer. Male F344 rats were fed the experimental diets containing 0, 100 and 200 ppm (NO-FLP), 40 and 80 ppm NO-IND, and 1500 and 3000 ppm NO-ASA dose levels representing 40 and 80% MTD. One week later, rats received s.c. injections of AOM (15 mg/kg body wt., once weekly for 2 weeks) and continued on the experimental diets for 46 weeks and sacrificed. Colon tumors were evaluated histopathologically. Catalytic activities of COX including COX-2 and inducible nitric oxide synthase (iNOS), and expression levels of markers associated with proliferation and apoptosis were determined in colon tumors. Administration of NO-FLP at 100 ppm, NO-IND at 40 and 80 ppm and NO-ASA at 3000 ppm significantly suppressed both tumor incidence and multiplicity (p<0.01 and p<0.001). The degree of inhibition was more pronounced with NO-IND at both dose levels (72-76% inhibition) than with the other NO-NSAIDs (43-67%). Administration of NO-IND at 40 and 80 ppm, NO-FLP at 100 and 200 ppm and NO-ASA at 3000 ppm significantly inhibited (p<0.01 to p<0.001) total COX including COX-2 activity (52-75% inhibition) and formation of PGs including PGE2, PGF2a and 6-keto-PGF1a and TxB2 from arachidonic acid (53-77% inhibition) in colon tumors. iNOS activity and b-catenin expression were suppressed in animals given NO-NSAIDs. In colonic crypts and tumors of animals fed NO-NSAIDs there was a significant increase in BrdU labeling and decrease in PCNA labeling when compared to animals fed the control diet. The results of this study provide strong evidence that NO-NSAIDs possess strong chemopreventive activity against colon carcinogenesis with NO-IND and NO-ASA being the most effective; their effect is associated with suppression of COX and iNOS activities and b-catenin levels in colon tumors. These results pave the way for the rational design of human clinical trials (Supported by USPHS contract CN-05112; R01 CA92423; from the National Cancer Institute).
Rao C.V., Malisetty V. Swamy, Suresh Guruswamy, Vernon E. Steele, Levy Kopelovich.
Chemoprevention of colon cancer by CP-31398, a p53 rescue agent administered individually or in combination with low-doses of celecoxib
Proc Amer Assoc Cancer Res 2006;47:[Abstract #3181].
CP-31398, a styrylquinazoline, is shown to rescue destabilized mutant p53 expression and to promote the activity of wild-type p53, i.e., cell cycle arrest and/or apoptosis. Previously, we have shown that COX-2 over-expression would lead to generation of electrophilic PGs that bind and impair the p53 protein in cytosol in colon cancer cells. Also, our studies had shown that CP-31398 (CP) suppresses the intestinal tumors in APCmin mice. The present study explored the chemopreventive effects of CP on 1) azoxymethane (AOM)-induced colon adenocarcinoma formation in male F344 rats and 2) assess whether combined effects of low-doses of CP and celecoxib (CEL) improve the chemopreventive efficacy in rats. Seven-week old, rats (48 /group) were fed control AIN-76A diet and given AOM (15 mg/kg b.w., s.c. once week for 2 weeks). Two weeks after the AOM treatment, rats were fed diets containing 0, 150, 300 ppm of CP-31398, or 300 ppm of celecoxib, or combination of 150 ppm CP plus 300 ppm CEL. Forty-eight weeks after AOM treatment, rats were sacrificed and intestinal tumors were evaluated by histopathologically. Multiple samples of colonic tumors were assayed for expression levels of p53, p21WAF1/CIP and markers of apoptosis (TUNEL) and cell proliferation (BrdU). We found that dietary CP at 150 and 300 ppm levels suppressed the colon adenocarcinoma incidence by 30% (p<0.02) and 44% (P<0.005), respectively. Similarly, dietary CP suppressed colon adenocarcinoma multiplicity by 51% (p<0.005, 150ppm) and 65% (p<0.0001, 300 ppm). CEL at the 300 ppm level significantly suppressed colon adenocarcinoma incidence (60%, p<0.0003) and multiplicity (70%, p<0.0001). Importantly, combinations of 150 ppm CP and 300 ppm CEL suppressed colon adenocarcinoma incidence by 78% and multiplicity by 89%. Combinations of low-doses of CP and CEL produced significantly more inhibitory effect when compared to 300 ppm CP and/or 300 ppm CEL alone. In tumors of rats fed 300 ppm CP and/or combinations of CP plus CEL significantly enhanced p53 and p21WAF1/CIP expression levels were observed when compared to control diet fed rat tumors. Administration of 300 ppm CP-31398, 300 ppm celecoxib and combination of 150 ppm CP-31398 plus 300 ppm celecoxib suppressed colon tumor cell proliferation (BrdU labeling index by 18-32%, p<0.05-0.01) and an increased apoptotic cells were observed in the adenocarcinomas when compared control diet fed colon tumors. These observations demonstrate, for the first time, that the p53-modulating agent CP-31398 possesses dose-dependent chemopreventive activity in well-established models of colon cancer, and the combination of low-doses levels CP-31398 and celecoxib significantly enhance colon cancer chemopreventive efficacy when compared to individual agents. [Supported by NIH grants CA-25114, N01-CN-43300 and CA-94962].
Oklahoma University Health Sciences Center, Oklahoma City, OK, Division of Cancer Prevention, National Cancer Center, Bethesda, MD
Reddy A., Bhanu Arasada, Duane Matthees, Chandradhar Dwivedi
Chemopreventive effects of dietary flaxseed on azoxymethane-induced colon tumor development in rats
Proc Amer Assoc Cancer Res 2005;46:[Abstract #3474].
Studies have shown that fats containing w-6 fatty acids (e.g., corn oil) enhanced and w-3 fatty acids (e.g., fish oil, flaxseed oil) reduced chemically induced colon tumors in rats. Lignans have also been shown to prevent colon cancer development in experimental animals. The objective of the present investigation is to study the effects of dietary flaxseed meal, a source of both w-3 fatty acids and lignans on colon cancer development, and compare with the effects of dietary corn meal. Male Fischer rats, two groups of 24 each were assigned to the AIN-93M diet supplemented with 15% corn meal and 15% flaxseed meal respectively. Carcinogenesis was initiated with subcutaneous injections of azoxymethane (15 mg/kg) once a week for three consecutive weeks. After 35 weeks of initiation, rats were sacrificed under ether anesthesia. Blood was collected by cardiac puncture. The gastrointestinal tract was isolated and flushed with ice-cold normal saline. The site, size and number of tumors were recorded. The fatty acid composition of the serum, colon and tumors was determined using Gas Chromatography-Flame Ionization detector. COX-2 activity was assayed by using Cayman COX activity assay kit in microsomes prepared from colon for both groups. Colon tumor incidence, multiplicity and size were found to be 82.6% and 29.4%; 1.3 and 0.2 tumors/rat; 44.4 and 5.3 mm2 in corn and flaxseed groups respectively. Colon and serum samples of the corn group showed an increase in w-6 fatty acids levels where as the flaxseed group exhibited an increase in w-3 fatty acid levels. COX-2 activity in flaxseed group was significantly lower when compared to corn group. Dietary flaxseed meal containing high levels of w-3 fatty acids and possibly lignans is effective in preventing colon tumor development as compared to dietary corn meal. This investigation is supported from a grant from North Dakota Oil Seed Council.
South Dakota State Univ., Brookings, SD
- Seril DN, Liao J, Ho KLK, Yang CS, and Yang GY:
Inhibition of Chronic Ulcerative Colitis (UC)-Associated Carcinogenesis in Mice
by N-Acetyl-L-Cysteine (NAC): Effects on Oxidative/Nitrosative Damage and
Proliferation - Proc. Am. Assoc. Cancer Res, 42, #4647 (abstr), 2001.
Sivananthan K., Ranjana P. Bird, Andrew W. Maksymiuk, Georgia Lefas, Asher Begleiter.
Effect of post-initiation induction of NQO1 by oltipraz on AOM induced colon tumor formation in Sprague-Dawley rats.
Proc Amer Assoc Cancer Res 2005;46:[Abstract #2476].
Phase II detoxifying enzymes like NADP(H):quinone oxidoreductase 1 (NQO1), glutathione S-transferases (GST) and UDP-glucuronyltransferases (UGT) may prevent carcinogen-induced cancers. Inducers of phase II detoxifying enzymes have been shown to inhibit carcinogen induced colon tumors in rat and mouse models, but the role of NQO1 in these effects was not clearly defined. We showed that oltipraz fed to Sprague-Dawley rats prior to treatment with a colon carcinogen increased NQO1 activity in the colons, without increasing GST or UGT activities, and decreased the number of aberrant crypt foci (ACF) in the colons of these rats. This was the first direct evidence that induction of NQO1 alone can inhibit initiation of colon carcinogenesis. Since oltipraz selectively induced NQO1 in these Sprague-Dawley rats without inducing other phase II detoxifying enzymes, this represents a unique model to study the role of NQO1 in colon carcinogenesis without interference from other phase II enzymes. Previous studies have shown that post-initiation administration of phase II enzyme inducers can also inhibit colon tumor formation in rats. Using the oltipraz fed Sprague-Dawley rat model, we investigated whether selective induction of NQO1 by oltipraz after treatment with the colon carcinogen azoxymethane (AOM) could also inhibit colon tumor formation. Rats were injected with 15 mg/kg AOM weekly for 2 weeks and then were fed control diet or oltipraz diet (control diet containing 200 ppm oltipraz) for 29 weeks. NQO1 activity was increased 2-fold in the colons of rats fed oltipraz diet compared with rats fed control diet, but the oltipraz diet did not increase GST or UGT activities. Rats fed oltipraz diet had 58% fewer ACF than control fed rats at 12 weeks after AOM treatment (p<0.001). However, there was no significant difference in the distribution of crypt multiplicities in the two groups. The frequency of microadenomas in oltipraz fed rats after 29 weeks was 1.07 ± 0.15/rat, while that in control fed rats was 1.87 ± 0.23/rat and this difference was statistically significant (p<0.003). The tumor frequency at 29 weeks was 0.37 ± 0.08/rat in rats fed oltipraz diet and 0.47 ± 0.09/rat in rats fed control diet, while the proportion of animals that developed colon tumors was 0.30 ± 0.05 in oltipraz fed rats and 0.37 ± 0.06 in control fed rats. Although the oltipraz fed rats had a lower tumor frequency and a lower tumor incidence than control fed rats the differences were not statistically significant. The median tumor volume was 432 mm3 in oltipraz fed rats and 219 mm3 in control fed rats. These results suggest that post-initiation induction of NQO1 can inhibit the early stages of colon carcinogenesis, but that this may not inhibit actual tumor formation. In addition, NQO1 induction may promote the growth of tumors once they have developed. (Supported by the Canadian Institutes of Health Research and CancerCare Manitoba Foundation)
Manitoba Institute of Cell Biology, CancerCare Manitoba, Winnipeg, MB, Canada, Department of Biology, University of Waterloo, Waterloo, ON, Canada, Department of Internal Medicine, University of Manitoba, Winnipeg, MB, Canada, Department of Pharmacology & Therapeutics, University of Manitoba, Winnipeg, MB, Canada
Swamy M.V., Suresh Guruswamy, Jagan M R. Patlolla, Ruchita Thungathurthi, Reddy S. Bandaru, Levy Kopelovich, Vernon E. Steele, Chinthalapally V. Rao
Chemoprevention of colon cancer by lovastatin and low-doses of celecoxib administered individually and in combination in F344 rats
Proc Amer Assoc Cancer Res 2006;47:[Abstract #2362].
Different mode of action exerted by combinations of agents represents a practical approach for improving the chemopreventive efficacy without unwanted side effects. Previously, we showed that a combination of HMG-R inhibitors with low-doses levels of COX-2 inhibitor enhance colon tumor cell apoptosis and suppress polyp formation in APCmin mice. Experiments were designed to assess the chemopreventive efficacy of lovastatin (HMG-R inhibitor) individually and/or in combination with low-dose levels of celecoxib (COX-2 inhibitor) on azoxymethane (AOM)-induced rat colon carcinogenesis. Prior to the efficacy study we assessed MTD levels of lovastatin and celecoxib in male F344 rats and found it to be 800 ppm and 5,000 ppm, respectively. At seven weeks of age, rats were fed the control diet (AIN 76A) and colonic tumors were induced by AOM. Two weeks after AOM-treatment rats were fed the diets containing 0, 200, 400 ppm lovastatin, 300 ppm celecoxib, or combination of 200 ppm lovastatin + 300 ppm celecoxib, and continued on the diets for 48 weeks before being sacrificed. Colon tumors were evaluated histopathologically and expression levels of markers for proliferation (BrdU, PCNA, p21waf1/cip1) and apoptosis (TUNEL) were determined by IHC. Dietary lovastatin at 200 ppm suppressed both invasive and non-invasive colon tumor incidence by ~ 35% (p<0.02) and multiplicity by ~52% (p<0.005). Diets with 400 ppm lovastatin suppressed the tumor incidence by 57% (p<0.0003) and multiplicity by 73% (p<0.0001). Lovastatin suppressed colon adenocarcinoma formation in a dose-response manner (p<0.001). Administration of 300 ppm celecoxib significantly suppressed adenocarcinoma incidence ~60% (p<0.0003) and multiplicity by 70% (p<0.0001). Importantly, the combination of low doses of lovastatin (200 ppm=25% MTD) and celecoxib (300 ppm=<6% MTD) inhibited adenocarcinoma incidence by >82% and multiplicity by ~88% as compared to control diet group. Combinations of low-dose levels of lovastatin and celecoxib provided significant (p<0.01) inhibition of colon adenocarcinoma formation when compared to high dose lovastatin or 300 ppm celecoxib alone. Tumors of rats fed lovastatin and/or celecoxib diets showed significantly reduced BrdU (~20-32%, p<0.01-0.001) and PCNA (15-35%, p<0.05-0.01) and increased TUNEL (22-30%, p<0.01-0.001) labeling, and an increased expression of p21waf1/cip1 when compared to control diet fed rat tumors. For the first time, our results suggests that lovastatin suppress AOM-induced colon adenocarcinoma formation in a dose-dependent manner, and the combination of low-dose levels celecoxib further improves the chemopreventive efficacy of lovastatin. These findings further support the development of low-dose combinations of HMGR-inhibitors with celecoxib for the chemoprevention of colon cancer. (Supported by NCI-CA-94962, N01-CN-25114 and N01-CN-43300).
Oklahoma Health Sciences Ce, Okalhoma City, OK, The Rutgers, The State University of New Jersey, New Jersey, NJ, Division of Cancer Prevention, National Cancer Institute, Bethesda, MD
- Tanaka T, Makita H, Ino N, Rahman W, Mori H,
et al.: Modifying effects of the arotinoid mofarotene (ro 40-8757) on
azoxymethane-induced rat colon and liver carcinogenesis. Cancer J 9, 260-268,
- Tanako T, Arai K, Murota I, Hayakawa K, Mizutani T, Mitsuoka T
Effects of feeding sour milk on longevity and tumorigenesis in mice and rats. Bifido Microflora 4, 31-37,
Toth B., Melissa Coles
Chemoprevention of murine large intestinal cancer by celecoxib using serial sacrifice studies
Proc Amer Assoc Cancer Res 2006;47:[Abstract #377].
The aim of this study was to inhibit the development of large intestinal cancer induced 1,2-dimethylhydrazine diHCl (1,2-DMH) by celecoxib (C) in Swiss female mice. In order to achieve our objective, the following experimental procedures were used: Group 1, 1,2-DMH was given as ten weekly subcutaneous injections at 20 mg/g to mice starting at nine weeks of age; Group 2, C was administered at 0.1% dose level in the powdered diet fed to mice which were seven weeks of age followed by 1,2-DMH as described in Group 1; and Group 3, physiological saline (PS) was given as ten weekly subcutaneous injections at 0.01 ml/g to mice which were nine weeks of age at the beginning of the experiment. Each group consisted of 30 mice that were sacrificed 26 weeks after the first injection of 1,2-DMH or PS. In addition, an identical set of experiments was added where mice were sacrificed 35 weeks after the first injection of 1,2-DMH or PS. The number of animals with large intestinal tumors and the total number of intestinal tumors were: Group 1, 29 and 438; Group 2, 18 and 64 (p<0.001); and Group 3, 1 and 1, in the animals sacrificed at 26 weeks. The corresponding tumor incidences in the animals sacrificed at 35 weeks were: Group 1, 30 and 323; Group 2, 23 and 134 (p<0.01); and Group 3, 0 and 0. Using the chi-square analysis, the differences in groups 1 and 2 are statistically significant. The lesions were diagnosed as polypoid adenomas and adenocarcinomas of the cecum, colon and rectum. C is a cyclooxygense-2 enzyme inhibitor that proved to be protective against human colorectal cancer. The current findings corroborate and extend prior results from our earlier investigation where the experiments were not subjected to serial sacrifice conditions. In the current study, the prevention of large intestinal cancer was achieved to a greater degree. Supported by NIH grant 5 RO1 AT001739
Eppley Institute for Research in Cancer, University of Nebraska Medical Center, Omaha, NE
Wali, W.L.; Stoiber, D.M.; Hart, J.A.; Brasitus, T.A.; Bissonnette, M. 2002
Polyethylene glycol suppresses hyperproliferation and increases apoptosis in AOM-induced aberrant crypt foci
Gastroenterology, 122, 5, Pages: A-241, (abstract from Digestive Disease Week)
- Yoshimi N, Qiao Z, Matsunaga K, Mori H, Hanausek M, et al.: The modifying effects of aceglatone, a B-glucuronidase inhibitor, in colon carcinogenesis of F344 rats. Proc. Am. Assoc. Cancer Res, 42, #957 (abstr), 2001.
D-Glucaric acid (GA) is a non-toxic, natural compound found in many fruits and vegetables. Our previous studies have shown that D-glucaro-1,4-lactone, the active metabolite of GA, inhibits
chemically induced tumorigenesis in rodents, in part, by inhibiting the enzyme b-glucuronidase. And GA is related to glucuronidation in a detoxification system. Aceglatone,
2,5-di-O-acetyl-D-1,4-glucaro-6,3-dilactone, is more stable than D-glucaro-1,4-lactone and is known as a postoperative prophylactic agent. We examined the effects on the initiation phase in rat
colon carcinogenesis induced by azoxymethane (AOM), 15mg/kg, s.c., 2 times. Aceglatone(0.5% or 2%) was treated as a diet for 5 weeks. The formation of aberrant crypt foci in the groups treated with
AOM plus 0.5% and 2% aceglatone diet for 5 weeks were significantly reduced compared with that of the group with AOM alone (49.5% and 45.2 %, respectively) at 5 weeks after treatment. In
addition, the tumor incidence in both groups also tends to decrease (69.2% and 68.8%, respectively) at the experimental termination of 36 weeks. It suggests that aceglatone may act as a blocking agent
of carcinogen detoxification system in colon carcinogenesis.
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