Chemoprevention of colorectal cancer: a systematic review in rodents.
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References to Min mice studies
that are not in PubMed (e.g., meetings abstracts)
- Ansari, S. H., DiBaise, J., Gulizia, J., Karolski, W. J., Ratashak, A., Wali, R. K., and Roy, H. K. Polyethylene glycol (PEG) 3350 suppresses intestinal tumorigenesis in the Min mouse. Gastroenterology, 122: A-215 #S1367, 2002.
Numerous studies have demonstrated that PEG is extraordinarily potent in the chemoprevention of experimental colon carcinogenesis (CC), suppressing >90% of azoxymethane-induced tumors in rats. However, concerns have been raised by the demonstration that PEG 8000 promoted colon adenomas in a transgenic mouse model (Naigamwalla et al., Cancer Res 2000). PEG 3350 (Miralax) is clinically used for the treatment of chronic constipation. We, therefore, examined the effects of PEG 3350 using the MIN mouse, a genetic model of CC. In order to investigate the mechanism of action of PEG, we evaluated cell proliferation by proliferating cellular nuclear antigen (PCNA). Furthermore, we assessed ß-catenin expression, a crucial regulator of cell proliferation in CC.
METHODS: Twenty male MIN mice 5-6 weeks old were randomized to PEG 3350 (10 % in drinking water) or control. Intestinal transit time was assessed by indigo carmine technique. Mice were sacrificed after 10 weeks, the small bowel (SB) and colon removed. Tumors were scored under magnification by 3 independent observers blinded to treatment group. PCNA and ß-catenin was detected in the uninvolved mucosa by immunohistochemistry using monoclonal antibodies and a Vectastain elite kit.
RESULTS: PEG 3350 was well-tolerated with no overt evidence of toxicity. The stool number was 95?4 in the control group, and this was significantly increased by PEG to 159?9. There was a concomitant decrease in transit time. Animal body weight gain was 88% greater in the PEG (p=0.01 versus control), presumably reflecting a lower tumor burden. Tumor multiplicity results are summarized in the table (* p<0.05, ** p<0.01) PEG treatment decreased total tumor number by 48%. Moreover, large tumors (>5 mm) were inhibited by 78%. PCNA expression was unchanged in the proximal small bowel was PEG (10.5?4 versus 11?3.8). ß-catenin expression was unaltered by PEG supplementation.
CONCLUSIONS: Our data demonstrates, for the first time, that PEG suppressed tumors in the MIN mouse. Moreover, this is the first study with PEG formulation that is in clinical practice. We demonstrate that inhibition of proliferation did not occur with PEG, in agreement with our previous in vitro report (Roy et al., FEBS Letters 2001). Further studies are ongoing to elucidate the mechanisms involved in chemoprevention.
TUMOR MULTIPLICITY (mean and S.D., in CONtrols and PEG-fed mice)
Total tumors, CON 42.0 SD16.6, PEG 22.2 SD5.5**
Prox Sml Bw, CON 08.2 SD02.3, PEG 05.5 SD2.8*
Dist Sml Bwl, CON 30.5 SD14.6, PEG 14.1 SD4.6**
Colon Rectu., CON 03.2 SD01.9, PEG 02.6 SD1.4
Cooke D., Schwarz M., Gescher A.J., Marczylo T.H.
Effect of cyanidin-3-glucoside and an anthocyanin mixture from bilberry on adenoma development in the Apc min mouse model of intestinal carcinogenesis - relationsship with tissue anthocyanin levels.
2006, accepted for publication Int.J.Cancer
- Itano, O., Naoto Kurihara, Kunhua Fan, Sadanori Abe, Levy Kopelovich, Leonard Augenlicht, Winfried Edelmann, Raju Kucherlapati, Kan Yang, Martin Lipkin.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. # 3159
Sulindac increases colonic tumors in mice with Apc, Mlh1 and Apc/Mlh1 mutations.
Strang Cancer Prevention Center, New York, NY, DCP, National Cancer Institute, Bethesda, MD, Albert Einstein College of Medicine, Bronx, NY, Brigham and Women's Hospital, Boston, MA.
We previously reported that sulindac had a dual effect on tumor development in Apc min/+ mice, inhibiting tumors in the small intestine and increasing tumors in the large intestine (Carcinogenesis, 22:1871-1875, 2001).
In this study we investigated this dual response in mice with a single Apc1638+/- or Mlh1+/- targeted mutation, and in compound mice with Mlh1+/-Apc1638+/- double mutations. All mice were fed control AIN-76A diet with or without 0.02% sulindac for 6 months.
The results were as follows:
(1) in Apc1638+/- mice tumor incidence in the small intestine decreased after feeding the sulindac diet (0% after sulindac vs 63% before, P<0.001). However, in the colon there was a significant increase in tumor incidence after feeding sulindac (92% vs 0%), with the number of tumors/mouse (2 vs 0; P<0.001). These findings in Apc1638+/- mice are similar to our previous observations in Apcmin/+ mice;
(2) In contrast, the tumor incidence in Mlh1+/- mice fed sulindac increased both in the small intestine (41% vs 13%; P<0.08) and in the colon (91% vs 6%; P<0.001). Similarly, tumor multiplicity and size also increased in the Mlh1+/- mice fed sulindac both in small intestine and colon;
(3) Mlh1+/-Apc1638+/- double mutant mice fed sulindac showed an increase in tumor incidence (100% vs 9%), multiplicity (1 vs 0.1) and size (5 vs .04 mm3 ) in the colon, all at P<0.001. Large intestinal tumors were located on the right side of the colon near the junction of cecum and proximal colon, and histologically malignant tumors were more frequently seen in the colon after sulindac treatment. In the small intestine, sulindac feeding of double mutant mice caused a decrease in tumor volume (P <.08) with no change in tumor incidence and multiplicity. Apoptosis and Bax/Bcl-xL expression were not substantially affected in tumors of small intestine and colon of Mlh1+/- mice fed sulindac.
In summary, sulindac increased tumor development in the colon of mice with Apc1638+/-, Mlh1+/-, and Mlh1+/-Apc1638+/- mutations. In the small intestine mice carrying the Apc mutation responded to sulindac with decreased tumor growth, and mice with the mismatch repair mutation responded to sulindac with increased tumor growth.
Jacoby, R. F., Cole, C. E., Hawk, E. T., and Lubet, R. A. Urodeoxycholate plus low dose sulindac is an effective chemopreventive agent combination that is well tolerated and decreases adenoma multiplicity in the Apc mutant Min mouse. Proc. AACR. 43, #3322, 2002.
Although sulindac and other NSAIDs are very effective colon tumor chemopreventive agents, we tested whether their therapeutic profile might be improved by use at lower more tolerable doses in combination with drugs acting via other mechanisms such as the bile salt ursodeoxycholate. Ursodeoxycholate caused a dose-dependent decrease in intestinal tumor multiplicity in the Min mouse. Combined treatment with low dose sulindac was indeed less toxic with normal weight gain and fewer gastrointestinal ulcerations compared to high dose sulindac, furthermore, combined treatment was more effective than either agent alone (see Table, below). These experiments suggest that cyclooxygenase inhibition when combined with other drugs is a very promising approach for colon cancer prevention.
Treatment, number of mice, Tumors/mouse +/- std.error, %/control, p-value
Control group, 8 mice, 17.9 T/m +/- 2.8, 100%
Ursodeoxycholate 500 ppm, 7 mice, 14.0 T/m +/- 1.7, 78%
Ursodeoxycholate 1500 ppm, 6 mice, 12.3 T/m +/- 1.7, 69% *
Urso. 500 + Sulindac 75, 7 mice, 5.3 T/m +/- 0.7, 30% **
Urso. 1500 + Sulindac 75, 7 mice, 3.1 T/m +/- 0.9, 17% ** +
Urso. 4500 + Sulindac 75, 8 mice, 2.1 T/m +/- 0.6, 12% ** ++
Sulindac 75 ppm, 8 mice, 6.4 T/m +/- 1.3, 36% **
Sulindac 150 ppm, 8 mice, 1.6 T/m +/- 0.5, 9% **
*p< 0.05, **p < 0.01 compared to control group;
+p < 0.05, ++p < 0.01 compared to Sulindac 75 ppm
- Ju, J., Mousumi Bose, Zhe Hou, Jie Liao, Adelaide M. Carothers, Chung S. Yang.
Inhibition of intestinal tumorigenesis in ApcMin mice by (-)-epigallocatechin-3-gallate (EGCG): Possible mechanisms involved.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 3137
We studied the inhibitory activity of (-)-epigallocatechin-3-gallate (EGCG), the most abundant catechin in green tea, against intestinal tumorigenesis in ApcMin mice, which have been recognized as a genetically relevant animal model to human intestinal carcinogenesis and utilized extensively for various chemoprevention studies. Male ApcMin mice on a C57BL/6J background were given either 0.32% EGCG as drinking fluid or water starting at 5 weeks of age. The experiment was terminated at 11 weeks of age. ApcMin mice that received 0.32% EGCG had significantly less small intestinal tumors than the control group that did not receive any treatment (16.5±12.5, N=18 vs. 29.9±13.4 N=19; p<0.005). To investigate whether 0.32% EGCG as drinking fluid modulates ß-catenin signaling and arachidonic acid metabolism, we measured small intestinal tumoric levels of E-cadherin and cPLA2 proteins by Western Blot analysis. Increased E-cadherin and decreased cPLA2 protein levels were found in small intestinal tumor lysates from 0.32% EGCG-treated ApcMin mice, when compared to those respective levels in small intestinal tumor lysates from control-treated ApcMin mice. These results suggest that EGCG effectively suppresses ß-catenin signaling and aberrant arachidonic acid metabolism in ApcMin mice, and these actions may contribute to the inhibition of tumorigenesis in ApcMin mice (supported by grant CA 88961).
- MacGregor, D. J., Kim, Y. S., Siddiki, B. B., Kwan, J., Sleisenger, M. H., and Johnson, L. K. Induction of colon cancer cell apoptosis in vitro and inhibition of intestinal tumor formation in Min mice by balsalazide and metabolites. Gastrointestinal Oncol., A553, 1996.
Merkel C.E., Wen-Chi L. Chang, Renata A. Coudry, Tianyu Li, Harry S. Cooper, Ronald A. Lubet, Insu P. Lee, Vernon E. Steele, Margie L. Clapper
Effect of bexarotene and mushroom extract lsy16 on intestinal neoplasia in the Apc Min/+ mouse model
Proc Amer Assoc Cancer Res 2006;47:[Abstract #3943].
Previous studies by others have demonstrated the ability of bexarotene (Targretin), a selective retinoid X receptor (RXR) agonist, to inhibit both mammary and lung cancers in a variety of animal models. An extract (lsy16) isolated from the mushroom Agaricus blazei Murill Kyowa has also been shown to decrease the formation of chemically induced pulmonary adenomas and colonic aberrant crypt foci (ACF) significantly. The goal of the present study was to assess the chempreventive activity of bexarotene and mushroom extract lsy16 in the ApcMin/+ mouse model of spontaneous intestinal tumorigenesis. Male ApcMin/+ mice (65 days of age) were maintained on a modified AIN 76A diet and randomized to receive either bexarotene, lsy16 or control diet (n = 10 per group). Bexarotene (400, 600 or 800 ppm in the diet) was administered for the duration of the study (45 days). lsy16 extract (100, 200 or 400 mg/kg body weight) was administered in the drinking water on Days 1-14, with animals receiving untreated drinking water on Days 15-45. Body weights were obtained weekly. At the time of sacrifice (Day 45), small intestines were collected for gross lesion count and histopathological evaluation. Bexarotene was well tolerated by all mice except those receiving the highest dose (800 ppm), for which a 20% reduction in survival was observed. The body weights of the remaining bexarotene-treated animals were decreased significantly on Days 35 and 42 (p = 0.01; Wilcoxon test). However, after Bonferroni multi-comparison adjustment, only the body weights of animals receiving 600 ppm bexarotene remained significantly different from those of controls. All animals survived exposure to lsy16, and treatment had no effect on body weight (p = 0.55; Wilcoxon test). No significant difference was observed in the number of gross small intestinal lesions present in bexarotene-treated vs. control animals (mean ± SEM - control 15.6 ± 5.2; 400 ppm 21.8 ± 7.7; 600 ppm 26.2 ± 9.3; 800 ppm 19.6 ± 7.4; p = 0.48 by Wilcoxon test). Likewise, administration of lsy16 had no effect on the number of gross small intestinal lesions per animal (mean ± SEM - control 16.8 ± 6.3; 100 mg/kg 20.9 ± 7.9; 200 mg/kg 21.0 ± 8.5; 400 mg/kg 14.5 ± 5.5; p = 0.82 by Wilcoxon test). Histopathological analyses have confirmed the lack of effect of bexarotene on spontaneous intestinal tumorigenesis. Similar histopathological reviews of tissues from animals exposed to lsy16 remain in progress. The bexarotene data from the present study are consistent with recent findings from others, which indicate the inability of this agent to inhibit tumorigenesis in mouse models of urinary bladder, prostate and ovarian cancer. The contrasting activity of the lsy16 extract against chemically induced ACF and spontaneous ApcMin/+ intestinal adenomas suggests that this agent may not exhibit chemopreventive activity against intestinal cells with an existing Apc mutation. (Supported by NIH N01 CN43309.)
Päivärinta, Essi M., Anne-Maria Pajari, Riitta Törrönen, Marja Mutanen.
Finnish wild berries decrease the number and size of intestinal adenomas in the Min mice.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 3896
University of Helsinki, Helsinki, Finland and University of Kuopio, Kuopio, Finland.
Epidemiological studies indicate that high consumption of vegetables, fruits and berries may prevent colon cancer. These foods contain a number of different phenolic compounds which have been shown to inhibit several stages of carcinogenesis in vitro. However, only a few studies have investigated the effects of berries on colon carcinogenesis in vivo. Therefore, we studied the effects of bilberry (Vaccinium myrtillus), lingonberry (Vaccinium vitis-idaea) and cloudberry (Rubus chamaemorus) on adenoma formation in the multiple intestinal neoplasia (Min) mice, a homologue of human familial adenomatous polyposis. At the age of 5 weeks the male and female Min mice were assigned to either a high-fat AIN93-G-based control diet or a diet containing 10% (w/w) freeze-dried bilberry, lingonberry or cloudberry. The mice were fed the experimental diets for 10 weeks. The concentrations of antocyanins and flavonols in the bilberry diet were 5532 mg and 104 mg/kg, and in the lingonberry diet 471 mg and 97 mg/kg, respectively. The cloudberry diet contained only 2 mg flavonols but 1564 mg ellagic acid/kg diet. After the feeding period, the mice were sacrified and the number, diameter and location of adenomas were determined by an inverse light microscope. All berries significantly decreased the number of adenomas in the small intestine, the average reduction being 40%. The mean number (±SD) of adenomas in the small intestine were 39 ± 17 (P <0.05) in the bilberry, 37 ± 13 (P<0.01) in the lingonberry and 43 ± 15 (P <0.05) in the cloudberry group compared to 65 ± 55 in the control group. The cloudberry and lingonberry also significantly decreased the mean size of adenomas which was most pronounced in the distal small intestine: the mean diameter of adenomas were 0.77 ± 0.10 mm (P <0.01) in the lingonberry, 0.69 ± 0.11 mm (P <0.01) in the cloudberry, and 0.97 ± 0.14 mm in the control groups. On the contrary, the mice fed bilberry had larger adenomas in the distal small intestine than the mice fed the control diet (1.06 ± 0.23 vs. 0.97 ± 0.14 mm; P <0.05). These results suggest that all three berries studied prevent the formation of new adenomas but only lingonberry and cloudberry prevent the growth of existing adenomas in the Min mice. Because the phenolic profiles of berries are different, their effects on the intestinal mucosa may be mediated through different pathways. It is also possible that the effects of berries on adenoma formation are independent of their phenolic profiles. Further studies are required to investigate the mechanims behind the chemopreventive effects of berries
- Pajari, A. M., Rajakangas, J., Paivarinta, E., Kosam, V. M., Rafter, J., and Mutanen, M. Inulin modulates intestinal tumor formation partly through an accumulation of cytosolic B-catenin in Min mice. AACR special conference in Cancer Research. Colon cancer: genetics to prevention. Philadelphia, Pennsylvania, March 7-10, 2002, A-16, 2002.
- Pajari, Anne-Maria, Seija Oikarinen, Marja Mutanen.
Both flaxseed mix and flaxseed oil suppress intestinal tumorigenesis in Min mice.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 3916
University of Helsinki, Helsinki, Finland.
Flaxseed is a rich source of plant lignans as well as a-linolenic acid, both of which may have cancer-protective effects. In this study, the effects of flaxseed mix and flaxseed oil on intestinal tumorigenesis were investigated in Min mice. At the age of 5 weeks, male and female mice were randomly assigned to the following AIN-93-based dietary groups: high-fat non-fiber control diet, the high-fat diet enriched with flaxseed mix (defatted flaxseed, flaxseed oil and wheat fiber) or with flaxseed oil at the same level as found in the flaxseed mix diet. The mice (n = 12/dietary group) were fed the experimental diets for ten weeks. The mean number and size of adenomas were decreased in the small intestine in the mice fed flaxseed mix compared to mice fed control diet (41 ± 17 vs. 59 ± 20; mean ± SD, P = 0.023 and 0.9 mm ± 0.1 vs. 1.2 mm ± 0.1, P = 0.002). Flaxseed oil resulted in a significant reduction in adenoma size (1.0 mm ± 0.2 vs. 1.2 mm ± 0.1, P = 0.012) and a tendency towards lower adenoma number (45 ± 23 vs. 59 ± 20, P = 0.095) when compared to the control mice. The flaxseed mix mice had higher levels of ß-catenin (1.5 ± 1.0 vs. 0.5 ± 0.6, relative intensity by western blotting, P = 0.025) and protein kinase C ? (0.54 ± 0.32 vs. 0.27 ± 0.23, P = 0.043) in the membrane fraction of the normal appearing mucosa than mice fed the control diet. Flaxseed oil feeding also resulted higher levels of ß-catenin and PKC ? in the mucosal membrane relative to the control group but the differences were statistically significant (P = 0.028) only in male mice. In the adenoma tissue, no differences in levels of ß-catenin or COX-2 were found between the dietary groups. These data suggest that the flaxseed mix and, to a lesser extent, flaxseed oil may prevent intestinal tumor formation in part by affecting adhesion and migration in the normal appearing mucosa in Min mice. The mechanisms by which the flaxseed mix and oil contribute to adenoma size and thus adenoma growth need to be studied further
- Rao, C. V., Malisetty, S. V., Cooma, I., and Reddy, B. S. Chemoprevention of FAP polyps and carcionmas by iNOS and COX2 selective inhibitors administered individually and in combination in the APC Min mice model. Proc. AACR. 43, #3323, 2002.
Inducible nitric oxide synthase (iNOS)/nitric oxide plays a pivotal role in colon tumor growth and provides an important molecular target for colon cancer prevention. We and others have consistently shown in preclinical models that cyclooxygenase-1 (COX-1) and COX-2 inhibitors are potent chemopreventive agents against the development of familial as well as chemically-induced colon cancers. Chemopreventive efficacy of iNOS selective inhibitor administered alone or in combination with COX-2 selective inhibitor was investigated in an animal model of familial adenomatous polyposis (FAP) containing germline mutation of the APC gene. In addition, we studied the specific involvement of iNOS in different stages of colon tumor development in APCmin mice. Six-week old male C57BL/6J-APCmin (heterozygous) or wild-type mice were fed high-fat diets containing 0, 100 ppm 1,4-phenylenebis(1,2-ethanediyl)-diisothiourea (PBIT), an iNOS-inhibitor, or 600-ppm nimesulide (NS), a COX-2 inhibitor, or combination of 50-ppm PBIT and 300-ppm NS for 80 days. At 19 weeks of age, all mice were sacrificed, and their intestines were evaluated for tumors. Multiple samples of normal appearing mucosa and tumors harvested from small intestines and colons were assayed for iNOS and COX-2 protein and RNA expressions using Western blot and RT-PCR. We found that the COX-2 inhibitor, NS, decreased the rate of formation of intestinal adenomas >80% (P<0.0001). However, the iNOS-inhibitor, PBIT slightly but significantly inhibited intestinal tumors (P<0.05). Interestingly, PBIT but not NS significantly inhibited (P<0.0001; ~90% inhibition) the colon adenocarcinomas. Combination of low-doses of PBIT and NS significantly suppressed intestinal adenomas and adenocarcinomas (P<0.0001). In addition, COX-2 was highly expressed in intestinal adenomas and adenocarcinomas whereas iNOS expression was high only in colon adenocarcinomas. These observations demonstrate for the first time that the iNOS-selective inhibitor, PBIT, administered individually or in combination with a COX-2 inhibitor suppresses the colon adenocarcinomas suggesting the chemopreventive potential of these agents against genetically predisposed neoplastic lesions, such as FAP. (Supported by NCI Grant CA-80003).
Rao, Chinthalapally V., Malisetty V. Swamy, Jayadev Raju, Joel Reinhardt, Levy Kopelovich.
The p53 rescue agent CP-313198 inhibits intestinal adenomas in APCmin-mice and aberrant crypt foci in the colon of carcinogen-treated F344 rats.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004.
Institute for Cancer Prevention, Valhalla, NY and National Cancer Institute - Division of Cancer Prevention, Bethesda, MD.
CP-31398, a styrylquinazoline, is shown to rescue destabilized mutant p53 expression and to promote the activity of wild-type p53. Cells exposed to CP-31398 undergo cell cycle arrest and/or apoptosis. The present study explored the chemopreventive effects of CP-31398 as a p53 modulator of 1) intestinal adenomas in an animal model of familial adenomatous polyposis (FAP) containing a germline mutation in the APC gene, and 2) aberrant crypt foci (ACF) in an azoxymethane (AOM)-treated rat model. Eight week-old female C57B/6J-APCmin (heterozygous) and wild mice were fed AIN-76A diets containing 0, 100 or 200 ppm of CP-31398 for 70 days. At ~120 days of age, all mice were sacrificed, and their intestines were screened for adenomas. In the rat model, seven-week old male F344 rats were fed AIN-76A diets containing 0, 100 or 200, or 400 ppm of CP-31398. Colonic ACF were induced by s.c. administration of AOM (15 mg/kg bw., once weekly for 2 weeks, at the age of 8 and 9 weeks). Eight weeks after AOM treatment, rats were sacrificed and colonic ACF were evaluated. Multiple samples of intestinal and colonic lesions were harvested from both APCmin mice and carcinogen treated rats and they were assayed for expression and activity of p53 and apoptosis. We found that dietary CP-31398 suppressed the development of intestinal (P<0.0001) tumor formation in APCmin mice in a dose-dependent manner (100 ppm, 36% and 200 ppm, 75%). Dietary CP-31398 also produced significant inhibition of AOM-induced colonic aberrant crypt foci with =4 crypts (p<0.05-0.005) and reduced total number of ACF (P<0.01) in F344 rats. Administration of CP-31398 effected a significant increase of p53 accumulation and translocation to nucleoli in the adenomas of APCmin mice. In addition, significant levels of apoptotic cells were observed in the adenomas of mice fed 200 ppm of CP-31398. These observations demonstrate, for the first time, that the p53-modulating agent CP-31398 possesses chemopreventive activity against intestinal neoplastic lesions in genetically-predisposed APCmin mice as well as carcinogen-induced preneoplastic colonic lesions in rats. [Supported by NIH grants CA-25114 and CA-94962]
- Sale, S.L. Richard G. Tunstall, Gerry A. Potter, William P. Steward, Andreas J. Gescher.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004.
Comparison of the effects of trans-3,4',5-trihydroxystilbene (resveratrol) and 3, 4, 4',5-tetramethoxystilbene (DMU212) in the ApcMin/+ mouse and on cyclooxygenase-2 (COX-2) expression in cells in vitro.
University of Leicester, Leicester, United Kingdom and De Montfort University, Leicester, United Kingdom.
Resveratrol is a naturally occurring polyphenol with cancer chemopreventive properties in preclinical models of carcinogenesis, including those of colorectal cancer. Recently DMU212, an analogue of resveratrol, was found to be superior to resveratrol in terms of growth-inhibitory and pro-apoptotic properties in transformed cells, when compared with their untransformed counterparts (Lu J. et al., Carcinogenesis, 22: 321-328, 2001). We wished to compare the ability of resveratrol and DMU212 to prevent malignancies associated with Apc mutations.
To that end ApcMin/+ mice received resveratrol or DMU212 admixed in the diet at 0.05, 0.2 and 0.5% from weaning to 18 weeks of age, after which adenoma numbers were scored.
Resveratrol at 0.05, 0.2 and 0.5% in the diet reduced adenoma number to 97 ± 31, 73 ± 10, and 70 ± 13 % of numbers in untreated controls, respectively; the decrease of adenoma number at the two higher doses compared to control was significant (p<0.05). DMU212 at the same doses reduced adenoma number to 85 ± 16, 76 ± 8 and 88 ± 22 % of control, respectively; here the decrease was significant only in the case of the 0.2 % dose (p<0.05). Expression of COX-2, which is thought to contribute to cancer progression and is a target of resveratrol, was measured by Western blotting in the human-derived colon cell lines HCEC and HCA-7. Resveratrol inhibited COX-2 expression in both cell lines at concentrations of 5µM and above, whilst DMU212 had a marginal effect, which was significant only at 50µM in HCEC cells. In order to compare the pharmacokinetic properties of these stilbenes, mice received them at 240 mg/kg via the intragastric route and drug concentrations were measured by HPLC in plasma and small intestinal and colonic mucosa. The ratios of area of plasma or target tissue concentration versus time curves of resveratrol over DMU 212 were 3.5, 0.1 and 0.15 for the plasma, small intestinal and colonic mucosa, respectively.
The results suggest that whilst DMU212 is more available than resveratrol in the small intestine and colon, both agents seem to exert adenoma retarding activity in the ApcMin/+ mouse to a comparable extent.
Swamy, M.V. Jayadev Raju, Jagan M. R. Patlolla, Bandaru S. Reddy, Chinthalapally V. Rao.
Chemoprevention of familial adenomatous polyposis (FAP) by lipitor and celebrex administered individually and in combination to male APCmin-mice.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 2363
Institute for Cancer Prevention, Valhalla, NY.
Preclinical and clinical studies have established cyclooxygenase (COX)-2 inhibitors as effective agents for the prevention and treatment of colorectal cancer. The COX-2 inhibitor, celebrex, suppressed colonic polyps up to 30% in FAP patients; however its chemopreventive efficacy in clinical studies is moderate suggesting a need for improvement. Previously, we have shown that a combination of HMG Co-A reductase (HMG-R) inhibitor, with COX-2 inhibitor synergistically increased the efficacy against colon carcinogenesis by enhancing apoptosis in vitro. Thus, experiments were designed to assess the chemopreventive efficacy of lipitor (HMG-R inhibitor), and celebrex, individually or in combination in an animal model of FAP based on a germline mutation of the APC gene. We also determined the levels of apoptosis, COX-2 and HMG-R expression and activities in intestinal tumors of APCmin mice. Six-week old male C57BL/6J-APCmin (heterozygous) or wild-type mice were fed high-fat diets containing 0 or 100 ppm lipitor or 300 ppm celebrex, or a combination of both for 80 days. At 18 weeks of age, all mice were sacrificed, and their intestines were screened for tumors. Multiple samples of normal appearing mucosa and of tumors harvested from small intestines and colons were assayed for apoptosis (TUNEL), and for HMG-R and COX-2 protein expression (Western blot) and activities (ex-vivo enzyme activity-substrate conversion). We observed that 100 ppm lipitor significantly (p<0.002) suppressed intestinal polyp formation. As anticipated, 300 ppm celebrex decreased the rate of formation of intestinal polyps by ~76% (p<0.0001). Importantly, the combination of lipitor and celebrex in the diet suppressed the intestinal polyps by >86% (p<0.0001) as compared to those fed the control diet group. The inhibition of intestinal adenoma formation by lipitor plus celebrex was highly significant (p<0.005) when compared to tumor inhibition achieved by celebrex alone. In addition, increased rates of apoptosis in intestinal tumors (p<0.01) were observed when animals were fed lipitor, celebrex, and their combinations. Similarly, significant decreases in HMGR-R and COX-2 activities were observed in animals fed lipitor plus celebrex. These observations demonstrate for the first time that the HMG-R inhibitor, lipitor suppresses intestinal tumorigenesis; importantly, when administered together with celebrex, it significantly increases the chemopreventive efficacy of COX-2 inhibitor against genetically predisposed intestinal tumor model. (Supported by NCI Grant CA-94962).
- Wilkinson, R.W., Anderson J. Ryan, Stephen R. Wedge, Ian T. Pyrah, Nikki Mandir, Robert A. Goodlad.
The VEGF receptor tyrosine kinase inhibitor ZD6474 significantly reduces intestinal tumor burden in the Min mouse.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 4657
AstraZeneca, Macclesfield, United Kingdom and Cancer Research UK, London, United Kingdom.
Angiogenesis is required to support tumor growth beyond 1-2 mm in diameter. The angiogenic 'switch' appears to be a distinct phenotypic phase of tumorigenesis, which may occur even during the preneoplastic stages of tumor development. ZD6474 is an orally active inhibitor of vascular endothelial growth factor receptor-2 (VEGFR-2, KDR) tyrosine kinase activity, with additional activity against epidermal growth factor receptor (EGFR) tyrosine kinase. In order to examine the role of VEGFR-2-dependent angiogenesis in early colorectal tumorigenesis, the effect of ZD6474 was investigated in multiple intestinal neoplasia (Min) mice, which spontaneously develop numerous benign polyps due to a mutation of the adenomatous polyposis coli (APC) gene, as in familial APC in humans.
ZD6474 (12.5, 25 or 50 mg/kg/day p.o.) or vehicle (1% polysorbate 80 in deionized water p.o.) was administered to 6-week old Min mice (n=12 per group, mixed male/female) for a period of 28 days. At necropsy, the small intestines and colons were isolated, rinsed and weighed. The small bowel was divided into three equal sections (proximal, middle and distal), dissected longitudinally and spread onto filter paper, as was the entire colon. The gut preparations were then fixed in Carnoy's fixative for 3 hours and then stored in 70% ethanol. The intestines were later assessed under a stereomicroscope (x20 magnification) for polyp number and diameter. Polyp volume was derived from polyp diameter, assuming a hemispherical shape in the small intestine and a spherical shape in the colon. The tumor burden was calculated as the product of polyp number and polyp volume.
In the small bowel, ZD6474 (50 mg/kg/day) was associated with a significant reduction both in mean polyp number per mouse (90.9 ± 20.7 vs 49.5 ± 12.9 in control vs treated, respectively, P=0.033) and in mean polyp diameter (1.18 ± 0.04 mm vs 0.95 ± 0.05 mm, P=0.005), resulting in a 75% decrease in polyp burden (76.5 ± 11.7 mm3 vs 18.3 ± 5.6 mm3, P=0.005). In addition, ZD6474 (50 mg/kg/day) treatment also resulted in fewer polyps in the colon (mean polyp number 3.46 ± 0.98 vs 0.83 ± 0.34 in control and treated, respectively, P=0.019).
These results support the hypothesis that the angiogenic switch may occur at an early, premalignant stage of tumor development and that VEGF/VEGFR-2 signaling plays a key role in the process. The present study has shown that ZD6474 significantly reduces polyp growth and number in the Min mouse model of intestinal adenomas, and provides a scientific rationale for studying the effects of VEGFR-2 signaling inhibitors in earlier disease in the clinic. ZD6474 is currently in Phase II clinical development
- Yang K., Osamu Itano, Kunhua Fan, Sadanori Abe, Zhiqian Dong, Ronald Lubet, Levy Kopelovich, Winfried Edelmann, Raju Kucherlapati, Martin Lipkin.
Tumors in small intestine and colon of Mlh1/Apc mutant mice after administration of the cyclooxygenase-2 inhibitor nimesulide.
Proc AACR, 45, AACR 95th annual meeting, Orlando, March 2004. 3161
Cyclooxygenase-2 (COX-2) is up-regulated in various experimental models of colon cancer. Nimesulide, a selective inhibitor of COX-2, has been shown previously to decrease tumors in the small and large intestine of Apc mutant mice, and in the colon of azoxymethane (AOM)-treated mice. In this study we evaluated the effect of nimesulide on tumor development and apoptosis in a compound mutant mouse model carrying both an Apc and an Mlh1 mismatch repair mutation. Apc1638+/-Mlh1+/- mice were fed AIN-76A diet with and without 0.04% nimesulide for 6 months. The findings are shown in Table1. Results of this study indicate that (1) Nimesulide, a selective inhibitor of COX-2, significantly decreased tumor growth in the small intestine of Apc1638+/-Mlh1+/- mice carrying both Apc and mismatch repair mutations. Concomitantly, apoptosis was significantly increased in tumors, crypt epithelial cells, and villous epithelial cells in the small intestine of these mice. (2) Nimesulide increased tumor incidence and volume in the colon of Apc1638+/-Mlh1+/- mice, and nimesulide-induced apoptosis was only marginally seen in the colon. In conclusion, the dual effect on tumor development (decrease in small intestine vs. increase in colon) seen with a COX-1 inhibitor sulindac, appears to be shared with a COX-2 inhibitor in Apc1638+/-Mlh1+/- mice.
Supported by NCI Awards N01-CN85164 and N01-CN15116
Corpet DE & Taché S, 2002, Nutrition & Cancer - & - DE Corpet & F Pierre, 2003, Cancer Epidemiol. Biomarkers Prevention
Polyp prevention in Min mice, Database & Mirror site