Chemoprevention of azoxymethane-induced rat aberrant crypt foci by dietary zerumbone isolated from Zingiber zerumbet.

The modifying effects of dietary feeding of zerumbone isolated from Zingiber zerumbet on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) were investigated in male F344 rats. Expression of cyclooxygenase (COX)-2 in colonic mucosa exposed to AOM and/or zerumbone was also assayed. In addition, we assessed the effects of zerumbone on cell proliferation activity of crypts by counting silver-stained nucleolar organizer regions protein (AgNORs) in colonic cryptal cell nuclei. To induce ACF rats were given three weekly subcutaneous injections of AOM (15 mg/kg body weight). They were also fed the experimental diet containing 0.01% or 0.05% zerumbone for 5 weeks, starting one week before the first dosing of AOM. AOM exposure produced 84+/-13 ACF/rat at the end of the study (week 5). Dietary administration of zerumbone caused reduction in the frequency of ACF: 72+/-17 (14% reduction) at a dose of 0.01% and 45+/-18 (46% reduction, p<0.001) at a dose of 0.05%. Feeding of zerumbone significantly reduced expression of COX-2 and prostaglandins in colonic mucosa. Zerumbone feeding significantly lowered the number of AgNORs in colonic crypt cell nuclei. These findings might suggest possible chemopreventive ability of zerumbone, through suppression of COX-2 expression, cell proliferating activity of colonic mucosa, and induction of phase II detoxification enzymes in the development of carcinogen-induced ACF.     

In situ quantification of aberrant p53 in colorectal neoplasia

Aberrant p53 protein accumulation was measured immunohistologically in 342 colorectal paraffin-embedded tissue sections from 115 patients (24 with adenocarcinoma, 59 with adenoma and 32 'hospital controls'). Subjective scoring was compared with quantitative cell imaging, including dichotomous (p53⁺/p53^?) status, ng p53^mut mg^?1 enterocyte protein, and tumour burden and patient body 'burden' of aberrant p53. A total of 62.5% cancer patients, 23.7% adenoma patients and 3.1% hospital controls were accorded p53⁺ status on the basis of p53 quantification. Quantitative p53⁺/p53^? assignment had a stronger inverse association with survival (χ²=6.17, p=0.013, Kaplan-Meier test) than subjective 'visual estimation' (χ²=0.57, p=0.449). There was a strong inverse relationship between the p53 'body burden' and the months of post-diagnosis survival (hazard ratio=1.42, p=0.0004, Cox proportional hazards). Absolute quantification for inactivated p53 permits objective and reproducible scoring, adjusts for intra-laboratory immunostaining 'batch effects', corrects for fixation artefacts, and standardizes for inter-laboratory differences in fixation, antibody selection and staining method. Clinically, in situ quantification of p53 will permit more accurate survival prognoses and will inform therapy selection and dose. Ultimately, accurate quantitative tissue/blood p53 correlations may provide a minimally invasive and systemic surrogate measure for these same clinical purposes.     

In situ quantification of aberrant p53 in colorectal neoplasia

Aberrant p53 protein accumulation was measured immunohistologically in 342 colorectal paraffin-embedded tissue sections from 115 patients (24 with adenocarcinoma, 59 with adenoma and 32 'hospital controls'). Subjective scoring was compared with quantitative cell imaging, including dichotomous (p53⁺/p53^-) status, ng p53^mut mg^-1 enterocyte protein, and tumour burden and patient body 'burden' of aberrant p53. A total of 62.5% cancer patients, 23.7% adenoma patients and 3.1% hospital controls were accorded p53⁺ status on the basis of p53 quantification. Quantitative p53⁺/p53^- assignment had a stronger inverse association with survival (χ²=6.17, p=0.013, Kaplan-Meier test) than subjective 'visual estimation' (χ²=0.57, p=0.449). There was a strong inverse relationship between the p53 'body burden' and the months of post-diagnosis survival (hazard ratio=1.42, p=0.0004, Cox proportional hazards). Absolute quantification for inactivated p53 permits objective and reproducible scoring, adjusts for intra-laboratory immunostaining 'batch effects', corrects for fixation artefacts, and standardizes for inter-laboratory differences in fixation, antibody selection and staining method. Clinically, in situ quantification of p53 will permit more accurate survival prognoses and will inform therapy selection and dose. Ultimately, accurate quantitative tissue/blood p53 correlations may provide a minimally invasive and systemic surrogate measure for these same clinical purposes.     

In situ quantification of aberrant p53 in colorectal neoplasia.

Aberrant p53 protein accumulation was measured immunohistologically in 342 colorectal paraffin-embedded tissue sections from 115 patients (24 with adenocarcinoma, 59 with adenoma and 32 'hospital controls'). Subjective scoring was compared with quantitative cell imaging, including dichotomous (p53+/p53-) status, ng p53mut mg(-1) enterocyte protein, and tumour burden and patient body 'burden' of aberrant p53. A total of 62.5% cancer patients, 23.7% adenoma patients and 3.1% hospital controls were accorded p53+ status on the basis of p53 quantification. Quantitative p53+/p53- assignment had a stronger inverse association with survival (chi2=6.17, p=0.013, Kaplan-Meier test) than subjective 'visual estimation' (chi2=0.57, p=0.449). There was a strong inverse relationship between the p53 'body burden' and the months of post-diagnosis survival (hazard ratio=1.42, p=0.0004, Cox proportional hazards). Absolute quantification for inactivated p53 permits objective and reproducible scoring, adjusts for intra-laboratory immunostaining 'batch effects', corrects for fixation artefacts, and standardizes for inter-laboratory differences in fixation, antibody selection and staining method. Clinically, in situ quantification of p53 will permit more accurate survival prognoses and will inform therapy selection and dose. Ultimately, accurate quantitative tissue/blood p53 correlations may provide a minimally invasive and systemic surrogate measure for these same clinical purposes.     

Dietary copper and dimethylhydrazine affect protein kinase C isozyme protein and mRNA expression and the formation of aberrant crypts in colon of rats.

Low dietary copper has been shown to decrease the expression of various protein kinase C (PKC) isozymes and increase the risk of colon cancer development in experimental animals. The purpose of this study was to investigate the relationship between dietary copper and carcinogen administration on PKC isozyme accumulation and aberrant crypt foci (ACF) formation in rats fed 0.9 and 7.7 microg Cu/g diet. After 24 and 31 d on the diets, the rats were injected with either dimethylhydrazine (DMH) (25 mg/kg i.p.) or saline and killed at two time points (2 wk and 8 wk after DMH). Rats fed low dietary copper had significantly lower (p<0.0001) hematocrits, hemoglobin, ceruloplasmin activity and plasma and liver copper concentrations than rats fed adequate dietary copper. Ingestion of low dietary copper significantly (p<0.005) increased the formation of DMH-induced ACF (116.8 vs 59.6). Low dietary copper significantly (p<0.05) decreased the concentration of PKC alpha, delta, and zeta in the colon at 2 wk but not at 8 wk. Thus, changes in PKC isoform protein concentration may be related to increased susceptibility of copper-deficient animals to colon cancer.     

A review of spatial computational models for multi-cellular systems, with regard to intestinal crypts and colorectal cancer development

Colon rectal cancers (CRC) are the result of sequences of mutations which lead the intestinal tissue to develop in a carcinoma following a “progression” of observable phenotypes. The actual modeling and simulation of the key biological structures involved in this process is of interest to biologists and physicians and, at the same time, it poses significant challenges from the mathematics and computer science viewpoints. In this report we give an overview of some mathematical models for cell sorting (a basic phenomenon that underlies several dynamical processes in an organism), intestinal crypt dynamics and related problems and open questions. In particular, major attention is devoted to the survey of so-called in-lattice (or grid) models and off-lattice (off-grid) models. The current work is the groundwork for future research on semi-automated hypotheses formation and testing about the behavior of the various actors taking part in the adenoma–carcinoma progression, from regulatory processes to cell–cell signaling pathways.     

Chemoprevention clinical trials.

As part of a program to develop drugs which will delay or prevent cancer in humans, the Chemoprevention Branch, National Cancer Institute, National Institutes of Health, is sponsoring 12 Phase I and 22 Phase II and III clinical trials. Three agent classes are significantly advanced in the trials. These are the retinoids, including 13-cis-retinoic acid, retinol, and 4-hydroxyphenylretinamide (nine studies), beta-carotene (seven studies), and calcium compounds (three studies). In addition, six promising new compounds are in Phase I or Phase II trials. These are: piroxicam, ibuprofen, oltipraz (a dithiolthione), difluoromethylornithine, glycyrrhetinic acid, and N-acetylcysteine. Key concepts related to the development of cancer chemopreventive agents are (1) the need for long-term administration, (2) the need for oral route of administration, (3) the matching of toxic side effects to degree of cancer risk.     

Optimality in the development of intestinal crypts

Intestinal crypts in mammals are comprised of long-lived stem cells and shorter-lived progenies. These two populations are maintained in specific proportions during adult life. Here, we investigate the design principles governing the dynamics of these proportions during crypt morphogenesis. Using optimal control theory, we show that a proliferation strategy known as a "bang-bang" control minimizes the time to obtain a mature crypt. This strategy consists of a surge of symmetric stem cell divisions, establishing the entire stem cell pool first, followed by a sharp transition to strictly asymmetric stem cell divisions, producing nonstem cells with a delay. We validate these predictions using lineage tracing and single-molecule fluorescence in?situ hybridization of intestinal crypts in infant mice, uncovering small crypts that are entirely composed of Lgr5-labeled stem cells, which become a minority as crypts continue to grow. Our approach can be used to uncover similar design principles in other developmental systems.     

Chemoprevention of cancer with retinoids.

The importance of retinoids for chemoprevention of cancer has received further emphasis with the finding that retinoids can suppress in vitro expression of the malignant phenotype, whether it be caused by chemical carcinogens, radiation, or viral transforming factors. Retinoids have been found to be potent inhibitors of the tumor-promoting effects of phorbol esters. Further advances in the chemistry of synthetic retinoids have led to the development of new agents that show less toxicity and better targeting to specific organ sites than natural retinoids. Synthetic retinoids have been particularly useful for prevention of bladder and breast cancer in experimental animals.     

Chemoprevention with aromatase inhibitors--trial strategies

Estrogen and its catechol metabolites from both the circulation and synthesized within the breast are important in the pathogenesis of breast cancer. Blocking estrogen's effects on the breast with selective estrogen receptor modulators (SERMS) is an ongoing strategy. Thus, tamoxifen and raloxifene reduce risk as monotherapy. Aromatase (estrogen synthetase) inhibitors are a logical alternative to SERMS. To date, SERMS have demonstrated reduction only in estrogen–progesterone receptor positive cancers without reduction in receptor negative tumors. By inhibiting the parent estrogens and their catechol metabolites, true prevention of cancer initiation might occur and reduction not only in the receptor positive but also negative tumors might result. Ongoing adjuvant breast cancer trials are exploring aromatase inhibitors as alternatives to tamoxifen, or in sequence or in combination with tamoxifen. Relative efficacies including reduction in contralateral breast cancer, toxicities and end-organ effects and impact on quality of life, are being explored. Data from these trials will help to guide future chemoprevention strategies. Proof of principal trials in ‘high risk’ cohorts such as premalignant breast lesions, dense screening mammograms, high plasma estradiol levels or increased bone density are already ongoing. Issues such as dose, schedule, therapeutic index and mono versus combination therapy are important to define.     

Chemoprevention of lung cancer by lycopene

An investigation was conducted to assess the chemopreventive potential of lycopene (LP), a naturally occurring hydrocarbon carotenoid found in tomatoes and their products, administered during the post-initiation stage in a multiorgan carcinogenesis model. One hundred eighteen B6C3F1 mice of both sexes were subjected to combined treatment with diethylnitrosamine (DEN), N-methyl-N-nitrosourea (MNU) and 1,2-dimethylhydrazine (DMH) from day 11 after birth to week 9 (DMD treatment) (groups 1 and 2) or given their vehicles (group 3). Then group 1 received LP (25 or 50 ppm in drinking water) for 21 weeks from weeks 11 to 32. Group 2 served as a carcinogen alone control and group 3 was given only LP (25 or 50 ppm). The incidences and multiplicities of lung adenomas plus carcinomas combined in male mice in group 1 receiving LP 50 ppm were significantly decreased as compared to the DMD alone or DMD and LP 25 ppm group values (75.0 vs 18.8%, P < 0.02; 0.94+/-0.17 v.s 0.25+/-0.14, P < 0.001). While hepatocellular carcinomas were lacking in the DMD and LP groups, two cases were found in the DMD alone group (not statistically significant). The values for aberrant crypt foci (ACF) and tumors in the colon and kidney did not show any significant variation among the carcinogen-treated subgroups. The results of this study provide evidence that the tomato carotenoid, lycopene, may have potential as a chemopreventive agent against carcinogenesis in the male lung.     

Chemoprevention of tumors: the role of RAR-beta

Chemoprevention can be defined as the use of specific natural or synthetic chemical agents to reverse, suppress, or prevent carcinogenic progression to invasive cancer. The knowledge of carcinogenic mechanisms provides the scientific rationale for chemoprevention. Epithelial carcinogenesis proceeds through multiple discernible stages of molecular and cellular alterations. Understanding of the multistep nature of carcinogenesis has evolved through highly controlled animal carcinogenesis studies, and these studies have identified three distinct phases: initiation, promotion and progression. Animal model studies have provided evidence that the development of cancer involves many different factors, including alterations in the structures and functions of different genes. Transitions between successive stages can be enhanced or inhibited in the laboratory by different types of agents, such activities providing the fundamental basis for chemoprevention.     

Early commitment and robust differentiation in colonic crypts

Tissue stem cells produce a constant flux of differentiated cells with distinct proportions. Here, we show that stem cells in colonic crypts differentiate early to form precisely 1:3 ratio of secretory to absorptive cells. This precision is surprising, as there are only eight stem cells making irreversible fate decisions, and so large stochastic effects of this small pool should have yielded much larger noise in cell proportions. We use single molecule FISH, lineage‐tracing mice and simulations to identify the homeostatic mechanisms facilitating robust proportions. We find that Delta‐Notch lateral inhibition operates in a restricted spatial zone to reduce initial noise in cell proportions. Increased dwell time and dispersive migration of secretory cells further averages additional variability added during progenitor divisions and breaks up continuous patches of same‐fate cells. These noise‐reducing mechanisms resolve the trade‐off between early commitment and robust differentiation and ensure spatially uniform spread of secretory cells. Our findings may apply to other cases where small progenitor pools expand to give rise to precise tissue cell proportions.