Studies of aberrant crypt foci (ACF).
References not found in Medline

• 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.
  
  
• Do, K.; Barnard, G. 2003
  Effect of concomitant PEG 3350 and celecoxib on development of aberrant crypt foci in rats
  Gastroenterology, Vol: 13X, Issue: suppl. X, Pages: Publ ID= W989 Abstract (Digestive Disease Week) ID 103984
  
  
• Femia Angelo Pietro, Maddalena Salvadori, Piero Dolara, Consuelo Bottini, Luciana Tessitore, Giovanna Caderni
  Proc Amer Assoc Cancer Res 2005; 46: [Abstract #3931]. AACR 96th Annual Meeting
  Characterization of mucin-depleted foci (MDF) as biomarkers of colon carcinogenesis in 1,2-dimethylhydrazine(DMH)-induced rats treated with polyethylene-glycol (PEG).
  Mucin-depleted foci (MDF) characterized by absent or scant mucous production were recently described by our group in the colon of carcinogen-treated rats (1). MDF can be easily identified along the entire mucosal surface of unembedded colon and are histologically dysplastic; moreover, MDF increase in rats treated with known promoters of colon carcinogenesis while decrease with chemopreventive agents such as piroxicam. On this basis, we suggested that MDF represent preneoplastic lesions that could be used as biomarkers for carcinogenesis. To further confirm this hypothesis, we studied MDF in 1,2-dimethylhydrazine (DMH)-initiated rats treated with polyethylene glycol (PEG) which has been demonstrated to inhibit colon cancer and preneoplastic lesions in carcinogen-treated rats (2). F344 male rats were treated weekly with DMH (150 mg/kg x 2, s.c) and then randomized into: control group, given AIN76 diet and water ad libitum and PEG group, fed AIN-76 diet and drinking water supplemented with 5% PEG. Sixteen weeks after the first dose of DMH, the total number of MDF was drastically reduced (P<0.01) in rats treated with PEG (MDF/ colon were 5.2±0.8 in controls and 0.1 ±0.1 in PEG-treated rats, means ±SE). Since PEG reduces colon cancer, MDF are correlated with carcinogenesis and can be proposed as endpoints to study the modulation of colon carcinogenesis in short-term experiments. To further characterize these lesions we studied the expression of p27, an inhibitor of cyclin-dependent kinases, which inhibits progression of the cell cycle (3). Immunohystochemical identification of p27 was determined in paraffin sections of MDF (15 samples) and in their paired normal mucosa (NM), all challenged with p27 antibody (monoclonal mouse anti-human p27Kip1). Compared with NM, the nuclear expression of p27 was markedly reduced in MDF (P<0.001) (cells with nuclear localization of p27 as % of total cells scored were 9.7%±1.4, and 25.4±1.7 % in MDF and NM, respectively, means ±SE). Cytoplasmic expression of p27 was not varied in MDF. In conclusion, MDF are correlated with colon carcinogenesis and have a marked reduction of nuclear p27, since such alteration has been reported in many human tumours including colon cancer, these results confirm the precancerous nature of MDF. 1) Caderni et al., Cancer Res 2003; 63: 2388-92. 2) Parnaud et al., Cancer Res 1999; 59: 5143-47. 3) Weinstein, Carcinogenesis 2000, 21: 857-864.
  
  
• Guruswamy S., Malisetty V. Swamy, Vernon E. Steele, Chinthalapally V. Rao.
  Chemoprevention of colon carcinogenesis by S-Adenosyl-L-methionine in male F344 rats
  Proc Amer Assoc Cancer Res 2006;47:[Abstract #3895].
  A number of studies has shown an increased risk of colon cancer occurs due to the dietary deficiency of methyl group and/or folate levels. S-Adenosyl-L-methionine (SAM) is required for several transmethylation reactions involving polyamines, proteins, and phospholipids and plays an important role in cell proliferation and regulation. In this current study, we investigated the chemopreventive effect of SAM on azoxymethane (AOM) induced colon carcinogenesis using the frequency of aberrant crypt foci (ACF) as an endpoint efficacy marker. Prior to the initiation of chemopreventive efficacy, we assessed the maximum tolerated dose (MTD) of SAM in the AIN-76A diet. For the MTD assay, seven week old male F344 rats were fed either the control AIN-76A or experimental diets containing five different dose (250, 500, 750, 1,000 and 1,500 ppm) levels of SAM for a period of six weeks. MTD was evaluated based on bodyweight gain, symptoms of toxicity and gross pathology. The MTD of SAM was found to be 1,000 ppm when it was administered in the AIN-76A diet. To evaluate the chemopreventive efficacy of SAM, seven week-old rats (18 rats/group) were fed either the control diet or an experimental diet containing either 400 or 800 ppm of SAM which corresponds to 40% and 80% MTD, respectively. At about 8 weeks of age, rats intended for carcinogen treatment were administered AOM by s.c. injection once a week for two weeks at a dose of 15 mg/Kg body weight. The rats were sacrificed 6 weeks after the last AOM injection and the colons were evaluated for ACF by methylene blue staining. Our results suggest that dietary administration of 800 ppm SAM significantly suppressed colonic total ACF formation by 36% (p<0.01), while a 400ppm SAM in the diet showed a 26% suppression. Interestingly, the 800 ppm SAM diet significantly suppressed the number of 4 or more crypts by more than 43% (p<0.005). These results indicate that in rats, SAM can reduce both the number and the frequency of ACF without any change in total body weight gain. SAM is a natural compound and is widely used at the clinical level as an anti-depressant and an anti-hepatotoxic agent and may therefore be an attractive candidate agent for chemoprevention of colon cancer. [Supported by N01-CN-65108, R01-CA-109247 and R01-CA-80003].
  Department of Medicine, Section of Hem/Onc, University of Oklahoma Health Science Center, Oklahoma City, OK, University of Oklahoma Health Science Center, Oklahoma City, OK, Division of Cancer Prevention, National Cancer Institute, Bethesda, MD
  
  
• Jain CK, Agarwal S, and Rao AV: The effect of dietary lycopene on bioavailability, tissue distribution, in vivo antioxidant properties and colonic preneoplasia in rats. Nutr. Res, 19, 1383-1391, 1999.
  Recent studies have suggested a protective role for lycopene, an antioxidant carotenoid, in the prevention of chronic diseases including cancer and coronary heart disease. Tomatoes and tomato products are the major dietary source of lycopene. The aim of this study was to investigate the bioavailability, tissue distribution, and the in vivo antioxidant properties of lycopene and its role in colon carcinogenesis. Male Fischer 344 rats were used in this study. Lycopene in the form of 6% oleoresin was incorporated into an AIN93M diet at a concentration of 10 ppm lycopene. Azoxymethane (AOM) was used as the colon specific chemical carcinogen. Serum and tissue lycopene levels were measured as estimates of lycopene uptake and tissue distribution. Serum TBARS and thiols were measured as indicators of lipid and protein oxidation. Incidence and size of aberrant crypt foci (ACF) were measured as preneoplastic markers. Dietary lycopene was absorbed and distributed to various tissues. An increase in serum thiols and a decrease in serum TBARS was observed in rats fed the lycopene diet. Incidence of ACF in lycopene fed rats showed a trend towards reduced numbers and size. Effects were more pronounced when lycopene was fed during the promotion stage than during the initiation stage. Based on these results it is concluded that dietary lycopene is absorbed by rats and distributed to various tissues. It acts as an antioxidant in reducing oxidation and thereby may protect against AOM induced ACF incidence. Dietary lycopene may play an important role in protecting against oxidative stress and colon carcinogenesis.
  
  
• Kwon, Y. et al., Aging alters the inhibition of colonic ACF by curcumin. Journal of Nutrition. 2002, 132, 11 Suppl. S, 3541S. Abstract from the International research conference on Food, Nutrition & Cancer, Washington DC, July 11-12, 2002.
  
  
• Murillo Genoveva, and Wendy H. Hirschelman, Jerome W. Kosmeder II, Robert M. Moriarty, John M. Pezzuto, Rajendra G. Mehta,
  Zapotin, a novel chemopreventive agent inhibits Azoxymethane-induced aberrant crypt formation in CF-1 mice
  Proc. Am. Assoc. Cancer Res, 43, #631. AACR 2002 Annual Meeting, San Francisco, Abstract
  Zapotin (5,6,2',6'-tetramethoxyflavone) is a naturally occurring flavone in the seeds of White Zapote (Casimiroa edulis Llave), a tropical fruit common in many parts of the world. In the present study, zapotin was chemically synthesized and evaluated for its potential chemopreventive effects on colon carcinogenesis. The formation of aberrant crypt foci (ACF) in response to carcinogen exposure is considered one of the earliest recognizable precancerous lesions. The ACF model has been used extensively to identify modulators of colon carcinogenesis. We investigated the effects of zapotin on the incidence and multiplicity of AOM-induced ACF in mice. CF-1 mice were treated s.c. with 10 mg/kg AOM weekly for 2 weeks. Zapotin (5.0 or 10.0 mg/kg/body weight) was administered intragastrically for 6 weeks, beginning 7 days before the carcinogen treatment and continued until termination of the study. Colons were stained with methylene blue and scored for the presence of ACF. Subsequently, the colons were stained for hexosaminidase (Hex) activity. Loss of Hex activity has been reported as a potential biomarker for colon cancer in rats. Zapotin significantly (p= 0.028) reduced the number of ACF/colon in a dose-dependent manner (14 +/- 2.3 control vs. 6.2 +/- 1.7, and 4.6 +/- 1.4, in 5.0 and 10 mg zapotin groups respectively). Moreover, colons isolated from the zapotin treated mice had significantly (p = 0.0018) fewer crypts with decreased hexosaminidase activity. Loss of enzyme activity was present in both the ACF, as well as, in the morphologically normal crypts of the mice treated with carcinogen. The mechanism of action for zapotin was investigated in HT-29 colon cancer cells. Incubation of HT-29 cells with zapotin (100nM to 10 ?M) for 6 days inhibited cell growth in a dose responsive manner. The cell cycle analysis indicated the cells treated with zapotin were arrested in G2 phase by 18 hours and remained arrested for up to 72 hours. In addition, zapotin induced programmed cell death in HT-29 cells as confirmed by ethidium bromide/acridine orange staining and DNA laddering. These results suggest a protective role of zapotin in colon carcinogenesis. (Supported by P01-CA-48112)
  
  
• Liu Y., Jihyeung Ju, Mou-Tuan Huang, Harold Newmark, Chung S. Yang
  Effects of a combination of calcium and aspirin on azoxymethane-induced aberrant crypt foci formation in the colons of CF-1 mice
  Proc Amer Assoc Cancer Res 2006;47:[Abstract #3172].
  Calcium supplementation and Aspirin have been shown to suppress colon carcinogenesis. The aim of the present study was to investigate whether the combination of calcium and Aspirin will generate a synergistic effect on aberrant crypt foci (ACF) formation in an azoxymethane (AOM)-induced colon tumorigenesis mouse model. Forty female CF-1 mice were randomly divided into 4 groups (10 mice in each group) and injected subcutaneously with AOM 10 mg/kg body weight once a week for two consecutive weeks at ages of 5 and 6 weeks. The basic control diet, a high-fat (20% mixed lipids by weight), low-calcium (1.4 mg /g diet) diet, was formulated to mimic the average Western diet. Three days after the last injection, mice were given a calcium-enriched diet (5.2 mg calcium/g diet), Aspirin-enriched diet (0.2 mg Aspirin/g diet), combination diet (5.2 mg calcium and 0.2 mg Aspirin/g diet) or control diet for 8 weeks until the experiment was terminated. Fixed colon tissues were stained with methylene blue, and the number of ACF containing single or multi-AC was scored under the microscope. Both calcium and Aspirin significantly decreased the total number of ACF/colon by 43% (P<0.05) and 40% (P<0.05), respectively. Combination of calcium and Aspirin significantly decreased number of ACF/colon by 50% (P<0.05). However, statistically significant differences among the extents of inhibition in different groups were not observed. We found a more prominent decrease in the number of large foci (³3 crypts/foci) than the number of small foci (< 3crypts/foci) by all of the treatments: calcium, Aspirin, and their combination decreased the number of large foci by 58%, 66%, and 79%, respectively (P<0.05). These results suggest that calcium and Aspirin have an additive effect in inhibiting colon carcinogenesis. Further studies are required to better characterize the effect (Supported by NIH grant CA 88961).
  Department of Chemical Biology, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ
  
  
• Morioka T., Masumi Suzui, Tatsuya Kinjo, Tatsuya Kaneshiro, Morihiko Inamine, Hideki Mori, Naoki Yoshimi.
  The relationship between mucin-depleted foci stained with Alcian bleu and beta-catenin accumulated crypts in rat colon carcinogenesis induced by 1,2-dimethylhydrazine dihydrochloride
  Proc Amer Assoc Cancer Res 2005;46:[Abstract #].
  The usefulness of mucin-depleted foci (MDF), which has been recently proposed as a new preneoplastic biomarker in rat colon carcinogenesis, was histologically investigated on rat colonic tissues treated with 1,2-Dimethylhydrazine dihydrochloride (DMH). The relationship among aberrant crypt foci (ACF), MDF and beta-catenin accumulated crypts (BCAC) was examined by comparing the corresponding computer-captured images. Twelve male F344 rats were received DMH s.c. at a dose of 40 mg/kg body weight, once a week for 2 weeks, and randomly divided into two groups. And then, while rats in group 1 were given a drinking water, those in group 2 were supplied indomethacin (IND) at 16 ppm in the drinking water for 6 weeks. All animals were sacrificed at the 8 weeks after the first DMH treatment. The resected colons were fixed in 10% formalin, and observed both ACF and MDF with Alcian blue staining (pH 2.5). Then they were histologically and immunohistochemically examined by using en face preparation. In group 1, The numbers of ACF, MDF and the overlapped lesions were counted 150±23, 19.5±5.2, and 0.67±0.5, respectively, as average. The numbers of ACF, MDF and the overlapped lesions (106± 23, 7.5± 1.9, and 0.17± 0.4, respectively) in group 2 treated with IND were significantly decreased, compared with those in group 1. The number of BCAC in group 2 (11.0± 2.7) was also significantly lower than that in group 1 (3.7± 0.8). The reduction (61.5%) of MDF by IND was much stronger than that (29.3%) of ACF. On the analyses with the corresponding computer-captured images, MDF had more frequent histological dysplastic changes (nuclear abnormality, Paneth cell metaplasia and so on) and the overexpression of beta-catenin than ACF. Especially, MDF having over 4 crypts or MDF with ACF appearance were much corresponded to BCAC. These results suggest that MDF may be useful as an early biomarker in colon carcinogenesis.
  Univ. of the Ryukyus, Okinawa, Japan, Gifu University Sch Med, Gifu, Japan
  
  
• 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
  Proc. Am. Assoc. Cancer Res, 43, #377. AACR 2002 Annual Meeting, San Francisco, Abstract
  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.
  
  
• Patlolla J.M.R., Venkat Swamy Malisetty, Jayadev Raju, Barbara Simi, Chinthalapally V Rao.
  Colon cancer chemopreventive properties of b-Escin, a triterpene saponin constituent of Aesculus hippocastanum
  Proc Amer Assoc Cancer Res 2005;46:[Abstract #2474].
  Extracts of Aesculus hippocastanum (horse chestnut) seed have been used in the treatment of chronic venous insufficiency, edema and hemorrhoids. Most of the medicinal beneficial effects of horse chestnut are attributed to its principal component b-escin or aescin. Recent studies suggest that b-escin may possess anti-inflammatory, anti-hyaluronidase, and anti-histamine properties. We have evaluated the preventive efficacy of dietary b-escin on azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) and analyzed the tumor cell growth inhibitory effects in the HT-29 human colon cancer cell line. To evaluate inhibitory properties of b-escin on colonic ACF, 7-week-old male F344 rats were fed experimental diets containing 0, 0.025% or 0.05% b-escin. 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). Rats were continued on experimental diets for 10 weeks and sacrificed. Colons were evaluated histopathologically for ACF and expression levels of markers associated with proliferation and apoptosis. Administration of dietary b-escin significantly suppressed total colonic ACF formation (~43%, p<0.001) when compared control diet group. Importantly, rats fed b-escin showed significantly lower number (~45%, p<0.0005) of multi-crypt foci containing ³ 4 or more. However, b-escin does not produce any significant dose-response effects on prevention of colonic ACF. In colonic crypts of rats fed 0.05% b-escin there was a significant increase in BrdU labeling (p<0.01) and decrease in PCNA labeling (p<0.005) when compared to animals fed the control diet. b-Escin also inhibited proliferation and induced apoptosis in the HT-29 human colon cancer cells in a dose-dependant manner. The results of this study for the first time provide evidence that b-escin, a naturally-occurring triterpene saponin, possess chemopreventive activity against colon carcinogenesis. (Supported by R01 CA-80003 from the National Cancer Institute).
  Department of Medicine, Hem-Onc Section, Oklahoma University Health Science Center, Oklahoma, OK, Department of medicine, Heam-Onc Section,Oklahoma University Health Science Center, Oklahoma, OK, Univ. of Waterloo, Department of Biology, Waterloo, ON, Canada
  
  
• Patlolla J.M.R., Rashid Hamid, Vernon E. Steele, Chinthalapally V. Rao
  Chemopreventive properties of probucol, purpurogallin and ethoxyquin against azoxymethane-induced colonic aberrant crypt foci formation in rats
  Proc Amer Assoc Cancer Res 2006;47:[Abstract #3918].
  Anti-oxidants play an important role in protecting cellular macromolecules from oxidative damage. Several naturally occurring anti-oxidants have shown an inverse relationship for risk of various cancers, including colon cancer. Similarly, recent studies support the protective role cholesterol lowering agents in various cancers. In the present study, a cholesterol lowering agent, probucol, and anti-oxidants purpurogallin, (a naturally-occurring) and ethoxyquin (a synthetic) were evaluated for colon cancer chemopreventive properties using azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) in male F344 rats as a efficacy endpoint. Prior to efficacy studies we evaluated the maximum tolerated dose (MTD) of each agent. For MTD assay, seven week-old rats were fed either control AIN-76 diet, or five different dose levels diets containing probucol, purpurogallin (each 500, 1,000, 2,000, 3,000 and 4,000 ppm) or ethoxyquin (500, 1,000, 1,500, 2,000 and 2,500 ppm) for six weeks. Based on body weight gain and toxicity symptoms, the MTD of probucol, purpurogallin and ethoxyquin is found to be 3,000 ppm, 4,000 ppm and 2,000 ppm, respectively, when administered in AIN-76A diet. To evaluate inhibitory properties of probucol, purpurogallin and ethoxyquin on colonic ACF, seven-week-old male F344 rats were fed experimental diets containing 0, 40, and 80% MTD of each agent. One week later rats received s.c. injections of AOM (15 mg/kg body wt., once weekly for two weeks) or equal volume of normal saline (vehicle). Rats continued on experimental diets for seven weeks before being sacrificed. Colons were evaluated histopathologically for ACF. Administration of probucol at 80% MTD level significantly suppressed total colonic ACF formation (~31%, p<0.001) when compared to the control diet group. Administration of 40 and 80% MTD of purpurogallin and ethoxyquin significantly suppressed total colon ACF formation (~35-48%, p<0.001-0.0001) compared to control diet fed rats. Importantly, rats fed 80% MTD of probucol, and 40 and 80% MTD levels of purpurogallin and ethoxyquin showed significantly lower numbers (~35-55%, p<0.005-0.0001) of multi-crypt foci (³ 4 or more). All three agents produced significant dose-response effects on prevention of colonic ACF. The results of this study for the first time provide evidence that probucol, a cholesterol lowering agent, and purpurogallin, a naturally-occurring agent, possess chemopreventive activity against colon carcinogenesis. (Supported by NO1-CN-65108; R01 CA-80003; and R01 CA-94962 from the National Cancer Institute).
  
  
• Raju, Jayadev, Jagan M. R. Patlolla, Malisetty V. Swamy, Chinthalapally V. Rao.
  Colon cancer preventive effects of Trigonella foenum graecum (fenugreek) seed and its constituent diosgenin using in vitro and in vivo models.
  Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 3155
  Institute for Cancer Prevention, Valhalla, NY.
  Trigonella foenum graecum (fenugreek) is traditionally used to treat various health disorders including diabetes, high cholesterol, wounds, inflammation, and gastrointestinal ailments. Recent studies suggest that fenugreek and its active constituents may possess anticarcinogenic potential. We evaluated the preventive efficacy of dietary fenugreek seed and its major steroid saponin constituent diosgenin on azoxymethane (AOM) -induced rat colon carcinogenesis during initiation and promotional stages. Colonic preneoplastic lesions or aberrant crypt foci (ACF) were chosen as end-points. In addition, we assessed the preventive mechanisms of diosgenin in the HT-29 human colon cancer cell line. To evaluate the effect during the initiation and postinitiation stages, 7-week-old male F344 rats were fed experimental diets containing 0, 1% fenugreek seed powder (FSP), or 0.05% or 0.1% diosgenin continuously and were then injected with AOM a week later; effects during the promotional stage was studied by feeding 1% FSP and 0.1% diosgenin 4-weeks after AOM-injections. Rats were sacrificed 8-weeks after AOM-injection and their colons were either evaluated for aberrant crypt foci (ACF) or for molecular markers. Dietary 1% FSP, 0.05% and 0.1% diosgenin administered continuously suppressed total colonic ACF up to 32%, 24% and 42% (p = 0.001-0.0001), respectively. Dietary 1% FSP and 0.1% diosgenin fed only during the promotional stage also inhibited total ACF up to 33% (p = 0.001) and 39% (p = 0.0001), respectively. Importantly, upon continuous feeding of 1% FSP, 0.05% or 0.1% diosgenin the numbers of colonic ACF with multiplicity = 4 were 38%, 20% and 36% lower than in controls (p = 0.001). Feeding 1% FSP or 0.1% diosgenin resulted in 25% and 32% lower (p = 0.001) incidence of multi-crypt ACF than in controls. Dietary diosgenin inhibited total colonic ACF and multi-crypt ACF formation in a dose-dependent manner. Diosgenin also inhibited proliferation and induced apoptosis in the HT-29 human colon cancer cell lines in a dose-dependent manner. Our results suggest that diosgenin induced apoptosis in HT-29 cells at least in part through modification of bcl-2 and caspase-3 levels. These findings add new insights into the development of fenugreek seeds and/or diosgenin as possible colon cancer-preventive agents. {Supported by NCI Grant CA-80003}
  
  
• Rao CV, Cooma I, Swamy MV, Simi B, and Reddy BS: Modulation of inductible nitric oxide synthase ans cyclooxygenase activities by curcumin during different stages of experimental colon carcinogenesis. Proc. Am. Assoc. Cancer Res, 42, #03084 (abstr), 2001.
  Curcumin, a naturally-occurring compound, is an effective chemopreventive agent against the several chemically induced cancers, including colonic tumors. The exact mechanism(s) of action of curcumin in the inhibition of colon tumorigenesis is not fully understood. To gain an understanding of the mode of action of curcumin we examined its effects on inducible forms of nitric oxide synthase (iNOS) and cyclooxygenase (COX-2) activities during the preneoplastic and neoplastic stages of azoxymethane (AOM)-induced colon carcinogenesis. Groups of 5 week-old male F344 rats were fed AIN-76A semipurified diet. At 7 weeks of age, rats in each dietary group except the vehicle-control groups received s.c. injections of AOM (15 mg/kg body wt) once weekly for 2 weeks. Three days after the AOM treatment groups of rats were transferred from control diet to experimental diets containing 0 or 2,000 ppm of curcumin and continued until termination of the experiment at week 8 or 48. Expression and activities of isoforms NOS and COX were analyzed in normal appearing mucosa, aberrant crypt foci (ACF) and tumors. Administration of curcumin significantly suppressed AOM-induced colonic ACF and tumor multiplicity. AOM treatment significantly increased colonic mucosal iNOS and COX-2 activities (P<0.001) during the preneoplastic stage, but had minimal effect on Ca+dependent NOS (eNOS) and COX-1 activities compared to those in vehicle-treated rats. Compared to adjacent mucosa, colonic tumors showed up to >50-fold higher activities of iNOS and COX-2; and up to >3-fold higher activities of eNOS and COX-1 correlating the protein expression levels of these enzymes. Dietary curcumin inhibited AOM-induced colonic iNOS and COX-2 (>50%) activities during the preneoplastic stage (P<0.01) and in tumors (P<0.0001). Also, curcumin significantly inhibited COX-1 activity (P<0.00) but it had no effect on the eNOS activity in colonic tumors. Immunohistochemical analysis of colonic ACF, showed that only ~8% of ACF express iNOS and <4% ACF express COX-2 protein; in contrast to almost all colon tumors were positive for iNOS and COX-2 protein expression. On the basis of enzyme activities and expression levels, the curcumin-induced inhibitory effect on iNOS occurred predominately at the transcription level, and that on COX-2 was mainly at the post-translational levels. Thus, dietary curcumin modulates AOM-induced iNOS and COX-2 activities during the preneoplastic and malignant stage of colon carcinogenesis by influencing these enzymes at transcription and post-translational levels.(Supported by NCI Grant CA-80003).
  
  
• 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
  
  
• Schmelz E.M., Hu Xu, Arun K. Rishi, Fazlul H. Sarkar, Adhip P. Majumdar.
  Reversal of early stages of colorectal cancer with EGF-receptor related protein (ERRP)
  Proc Amer Assoc Cancer Res 2006;47:[Abstract #1239].
  Inactivation of EGFR family members (EGFRs) represents a promising strategy for the development of novel and selective anticancer therapies. Current anti-EGFR therapies target EGFR or HER-2 but not both. Since most solid tumors express several members of EGFRs, identification of inhibitor(s), targeting multiple members of the EGFR family may provide a therapeutic benefit to a broader patient population. EGF-Receptor Related Protein (ERRP), a 53-55 kDa secretory protein, which we have isolated and characterized as a pan-erbB inhibitor that targets multiple members of EGFRs, is a potential therapeutic agent for several epithelial cancers, including colorectal cancer. We have repeatedly observed that ERRP is highly effective in inhibiting growth of SCID mice xenografts of colon cancer cells. However, it remains to be determined whether ERRP can inhibit/regress growth of chemically-induced colonic neoplasia/tumors in immunocompetent mice. As a first step in achieving this goal, we studied the effect of recombinant ERRP on inhibition/regression of aberrant crypt foci (ACF) formed in the colon of mice after dimethylhydrazine (DMH) treatment. ACF are considered to be the precursor of colorectal adenomas and subsequently carcinoma. CF1 mice were injected (i.p.) once a wk for 6 wks with DMH (30 mg/kg bodyweight) to induce ACF formation, subsequently divided into two groups and injected (i.p.) once a day for 10 days with recombinant ERRP (50 µg/mouse) or vehicle. Administration of ERRP significantly decreased (40-50%) the number of ACF per animal (P<0.05). The number of ACF per focus was also reduced by 30-40% after ERRP treatment, compared to the controls (P<0.01). This was associated with a marked inhibition (50-60%) of proliferation (PCNA immunoreactivity) and stimulation (60-70%) of apoptosis (TUNEL assay) in the colonic mucosa. These changes were also accompanied by marked reduction in activation (tyrosine phopshorylation) of EGFR, HER-2 and HER-3 in the colonic mucosa leading to attenuation of NF-kB activity. In contrast, the levels of alkaline phosphatase, a marker of intestinal differentiation, was greatly elevated in the colonic mucosa following ERRP treatment. Taken together, the results suggest that ERRP is effective in regressing the number and the size of existing ACF, which are precursors of colorectal adenomas and colorectal carcinoma. This could partly be attributed to inhibition of EGFRs signaling and induction of differentiation of the colonic mucosa.
  Karmanos Cancer Inst/Wayne State University, Detroit, MI, Wayne State University, Detroit, MI, VA Med Ctr/Wayne State University, Detroit, MI
  
  
• 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
  
  
• Steele, V.E., Wargovich, W.J., McKee, K., Sharma, S., Wilkinson, B.P., Wyatt, G.P., Gao, P., Kelloff, G.J. Cancer chemoprevention drug development strategies for resveratrol Pharmaceutical Biology, 1998, 36: suppl, 62-68
  No abstract available online, sorry!
  
  
• Suzui M., Takamitsu Morioka, Morihiko Inamine, Hisataka Moriwaki, Naoki Yoshimi.
  Acyclic retinoid inhibits 1,2-dimethylhydrazine-induced aberrant crypt foci formation and colon tumor development in F344 rats and induces apoptosis in HCT116 and SW480 human colon carcinoma cell lines
  Proc Amer Assoc Cancer Res 2005;46:[Abstract #5212].
  Acyclic retinoid (ACR), a novel synthetic retinoid, has recently been demonstrated by us to inhibit the growth of human hepatoma cells and this effect was associated with induction of p21CIP1 and inhibition of expression of cyclin D1. These results suggest that this agent may be useful in the chemoprenevtion and therapy of hepatoma and possibly other human malignancies (Suzui, M. Cancer Res 62: 3997, 2002 & Mol Cancer Ther 3: 309, 2004). In the present studies, we examined the possible chemopreventive effect of ACR on rat colon carcinogenesis during initiation and postinitiation stages. We chose aberrant crypt foci (ACF) as end points in short-term colon carcinogenesis bioassays because ACF are preneoplastic lesions in rodents and humans. Colon tumors were induced to evaluate the preventive effect of ACR in medium-term colon carcinogenesis. In addition, we assessed the mechanism of growth inhibitory effect of ACR in HCT116 and SW480 human colon carcinoma cell lines. Exp. 1: Five-week-old male F344 rats were randomly assigned to the experimental groups. Rats in groups 1-3 were given subcutaneous injections of 1,2-dimethylhydrazine (DMH, 40 mg/kg body weight) once a week for 2 weeks to induce ACF. One week before the first injection of DMH, rats in groups 2 and 3 were fed a diet containing 50 and 150 ppm ACR, respectively, for 5 weeks. Rats in group 4 were fed a diet containing 150 ppm ACR. Rats in group 5 were given the basal diet alone and served as untreated controls. All animals were sacrificed at 5 weeks after the start of the experiment. Oral administration of 50 and 150 ppm ACR significantly decreased the number of ACF (58% and 52% decrease, respectively, P<0.001), when compared with the group treated with DMH alone. Exp. 2: Rats in groups 1-3 were given subcutaneous injections of DMH once a week for 2 weeks. Starting 1 week after the second injection, the rats received 1% dextran sodium sulfate (DSS) in drinking water for 1 week. One week after the DSS treatment, rats in groups 2 and 3 were fed a diet containing 50 and 150 ppm ACR, respectively, for 16 weeks. Rats in groups 4 and 5 were treated in the same protocol as described in Exp. 1. All animals were sacrificed at 16 weeks after the start of the experiment. Continuous oral administration of 50 and 150 ppm ACR caused a significant decrease of tumor multiplicity (P<0.005) and tumor size (P<0.05) compared with the control group. No toxic effects caused by ACR were observed in all organs examined. Furthermore, ACR caused a marked and dose-dependent inhibition of growth of the HCT116 and SW480 human colon carcinoma cell lines. These findings suggest that ACR may be a promising chemopreventive agent for human colon carcinogenesis. These novel effects of ACR provide further evidence that ACR may be a valuable compound in the chemoprevention and therapy of various malignancies.
  Department of Patholgy, University of the Ryukyus Faculty of Medicine, Okinawa, Japan, Department of Internal Medicine, Gifu University School of Medicine, Gifu, Japan
  
  
• Tanaka T., Rikako Suzuki, Hiroyuki Kohno, Kazuya Hata, Shigeyuki Sugie, Shingo Miyamoto, Kuniaki Sugawara, Takashi Sumida, Yoshinobu Hirose.
  Diet supplemented with citrus unshiu segment membrane suppresses azoxymethane-induced colonic aberrant crypt foci and b-catenin accumulated crypts in male db/db mice
  Proc Amer Assoc Cancer Res 2006;47:[Abstract #661].
  Background and aims: Obesity/diabetes is known to be one of the risk factors for colon cancer development, although the mechanism is not fully understood. We previously reported the cancer chemopreventive effects of several compounds in citrus fruits on chemically induced colon tumorigenesis. The current study was designed to know whether male C57BL/KsJ-db/db (db/db) mice possessing obese and diabetic genotypes due to their disruption of the leptin receptor are sensitive to azoxymethane (AOM)-induced colon tumorigenesis using two biomarkers, aberrant crypt foci (ACF) and b-catenin accumulated crypts (BCAC). In addition, the modulatory effects of dietary citrus unshiu segment membrane (CUSM) containing contains fiber and hesperidin on the occurrence of the lesions were determined in male db/db, db/+, and +/+ mice.
  Methods: Male db/db, db/+, and +/+ mice were given 5 weekly subcutaneous injections of AOM (15 mg/kg bw,). Starting one week after the last injection of AOM, they were fed the diet containing 0.02%, 0.1% or 0.5% CUSM for 7 weeks. The frequencies of ACF and BCAC were counted at the end of the study (wk 12).
  Results: At wk 12, a significant increase in the numbers of ACF and BCAC was noted in db/db mice in comparison with those of db/+ and +/+ mice. Dietary administration with CUSM resulted in a reduction in the frequency of ACF in all the genotypes of mice, and the potency was high in order of the db/db mice (53-59%, P<0.001), the db/+ mice (31-48%, P<0.05), and the +/+ mice (31-63%, P<0.05). The number of BCAC was also reduced by feeding with CUSM: 28-61% reduction (P<0.05) in the db/db mice; 20-49% in the db/+ mice; and 21-53% reduction in the +/+ mice. Clinical chemistry revealed that CUSM feeding lowered serum triglycerides, but did not serum leptin and insulin levels in the mice. Other interesting findings included that CUSM feeding reduced the area of islets of Langerhans, liver fatty metamorphosis, and liver fibrosis in the db/db mice that received AOM.
  Conclusion: Our results may indicate that the db/db mice are sensitive to AOM-induced colon carcinogenesis in comparison with the db/+ and +/+ mice, suggesting the positive association between colon cancer development and obesity/diabetes. The findings also suggest that CUSM in diet has chemopreventive ability against the early phase of AOM-induced colon carcinogenesis in the db/db as well as the db/+ and +/+ mice. (Supported financially by the Ministry of Agriculture, Forestry and Fisheries of Japan)
  Kanazawa Medical University, Ishikawa, Japan, BMR Laboratories, Sunplanet Co., Ltd., Gifu, Japan, Kyoto University, Kyoto, Japan, Ehime Beverage Inc., Matsuyama, Japan, Gifu University, Gifu, Japan
  
  
• Verghese Martha, Lloyd T. Walker, Louis Shackelford, Chandramohan B. Chawan
  Inhibitory effects of nondigestible carbohydrates of different chain lengths on azoxymethane-induced aberrant crypt foci in Fisher 344 rats
  Nutrition Research 25 (2005) 859-868
  Colon cancer is one of the leading causes of cancer morbidity and mortality in Western countries. The objective of this study was to elucidate the effect of prebiotic carbohydrates of different chain lengths on azoxymethane-induced aberrant crypt foci in Fisher 344 male rats. After an acclimatization period of 1 week, 70 male weanling rats were divided into 7 groups and fed AIN-93G (Control) and 6 experimental diets that contained control + (maltodextrin; Raftiline HP, Raftiline ST, Raftilose P95, Raftilose Synergy1, and Mix; ORAFTI, Tienen, Belgium). All the rats received 16 mg/kg body weight of azoxymethane dissolved in saline subcutaneous at 7 and 8 weeks of age. The rats continued to receive the assigned diets until killed by carbon dioxide asphyxiation at 17 weeks of age. There was a significant (P b .05) increase in cecal weight and a decrease in cecal pH in rats fed prebiotic carbohydrates. The highest reduction of colonic aberrant crypt foci, both in total number as well as crypt, multiplicity was seen in the group fed Mix (63.9%). Consumption of diets containing Raftilose Synergy1, Raftiline ST, and Raftiline HP showed a reduction of total colonic crypts by 52.2%, 29.6%, and 46.3%, respectively, as compared with the control diet.
  
  
• Volate, Suresh R., Destiny M. Davenport, Ala Y. Issa, Michael J. Wargovich.
  Modulation of aberrant crypt foci and apoptosis by dietary herbal supplements.
  Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 3160
  University of South Carolina, Columbia, SC.
  It is estimated that one third of Americans use dietary herbal supplements on a regular basis. Diets rich in bioactive phytochemicals are associated with reduced risk of certain cancers, notably, colon cancer. Herbal supplements have not been directly tested as sources of bioactive cancer preventives. Hence, this study compares the ability of four herbal flavonoids (quercetin, curcumin, rutin, and silymarin) and one whole herb mixture (ginseng powder) to suppress aberrant crypt foci (ACF) in an azoxymethane (AOM)-induced rat colon cancer model. Second, this study examined the effect of these herbal compounds on apoptosis and the mechanisms by which these compounds evoke apoptosis. The results of this study show that diets containing quercetin, curcumin, silymarin, ginseng and rutin led a 4, 2, 1.8, 1.5, and 1.2 fold decrease in the number of ACFs compared to control, respectively. Histological analysis of the colon mucosa shows that all of the herbal supplements tested induced apoptosis, with quercetin being the most potent (3X increase compared to control). Thus quercetin's chemopreventive activity toward ACF correlates strongly with its function as an inducer of apoptosis. Western blot analysis of Bax (proapoptotic) and bcl-2 (anti-apoptotic) proteins from the colon scraping revealed that quercetin and ginseng induces apoptosis via the mitochondrial pathway. Taken together, the results of this study suggests that out of the five commonly used herbal supplements, quercetin is the most effective at reducing the number of ACFs and it does so by inducing apoptosis via the mitochondrial pathway. The mechanism by with the other compounds evoke apoptosis are being investigated. Supported by AICR grant 00B041-REV
  
  
• 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)
  
  
• Wargovich MJ, Woods C, Liao J, Hollis D, Marquart L, et al.: Calcium fortified whole grain diet inhibits aberrant crypt foci in the rat colon. Proc. Am. Assoc. Cancer Res, 42, #949 (abstr), 2001.
  Whole grains are rich sources of phytochemicals, many of which have anticarcinogenic properties. Whole grain fiber can also act as a carrier for trace minerals, such as calcium (Ca++) delivering it into the colon where it could have chemoprotective effects. To test this hypothesis groups of 10 F344 rats were fed AIN-93G for 4 wks and injected with 15 mg/kg azoxymethane at weeks 1 and 2 to induce ACF. The rats were then randomized to the same diet, supplemented with either whole grain cereal (WGC), or WGC fortified with Ca++ to provide either 75 mg Ca++/d, or with 300 mg Ca++/d; these were fed during a second 4 wk period. Colons were removed at wk 8, fixed and stained to assess ACF number and multiplicity. ACF incidence/colon for the AIN93G group was (144±14) while ACF formation was significantly inhibited by calcium supplementation at the 300 mg Ca++/d level (86±17 compared to AIN93G alone (P=0.019). Ca++ supplementation at the 75 mg/d mg level did not significantly reduce ACF formation. Analysis of crypt multiplicity data revealed that the higher Ca++ diet inhibited ACF regardless of the number of crypts/ACF focus. In conclusion, calcium supplementation in whole grain cereals results in reduced expression of preneoplastic foci in the rat colon.
  
  
• Woods Cynthia J., and Destiny M. Hollis, David Klurfeld, Donald Yowell, Michael J. Wargovich,
  The AIN-93G semi-synthetic diet suppresses azoxymethane-induced aberrant crypt foci in the F344 rat
  Proc. Am. Assoc. Cancer Res, 43, #861. AACR 2002 Annual Meeting, San Francisco, Abstract The semi-purified AIN-76A diet has been widely used as a basal rodent diet in chemoprevention studies for many years. In 1993, this diet was re-formulated to better meet the nutritional requirements of the animals during both growth (AIN-93G) and maintenance (AIN-93M) stages of life. However, retrospective analysis of several studies in our laboratory using the azoxymethane (AOM) model for colonic aberrant crypt foci (ACF) indicates that there may be a reproducible, differential effect of AIN-93G and AIN-76A on AOM-induced ACF formation. Therefore, we sought to directly compare the effects of these basal diets in the AOM ACF model. Male F344 rats were fed either AIN-76A or AIN-93G diet, injected with 15 mg/kg AOM (two times, one week apart), and maintained on the diets for three additional weeks. Neither diet induced ACF formation without AOM. However, treatment with AOM resulted in 22% more ACFs in the rats fed the AIN-76A diet than the rats fed the AIN-93G diet (p<0.02). These data suggest that AOM-induced ACF formation is suppressed by the specific formulation of the AIN-93G diet. Elements that differ between these diets, widely used in tumorigenesis studies, include the source of carbohydrate and the substitution of an omega-3 fatty acid rich oil in the AIN-93G relative to the AIN-76A diet. Additional studies are aimed at identifying potential mechanisms of protection by the AIN-93G diet such as apoptosis, proliferation, and carcinogen activation and/or detoxification. The effects of these diets will also be evaluated in the APCmin mouse model, another common model of intestinal tumorigenesis. These data have wide-ranging implications for animal chemoprevention studies using semi-synthetic diets.
  
  
• 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.