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Aspirin, taken daily over at least one year, may exert chemopreventive effects against the early stages of colorectal carcinogenesis Preclinical, observational, and clinical data consistently show that non-steroidal anti-inflammatory drugs (NSAIDs)—particularly aspirin—reduce colorectal carcinogenesis.1 Scores of animal studies show that NSAIDs inhibit the development of colorectal neoplasia across the spectrum of disease, ranging from aberrant crypt foci (ACF) to cancer.2 Human data confirm these findings with dozens of observational studies reporting 40–50% reductions in colorectal adenoma incidence, cancer incidence, and cancer associated mortality among aspirin users. The most compelling data were published earlier this year. Two randomised placebo controlled trials conducted in patients at moderate risk for colorectal cancer reported that aspirin administered at doses as low as 81–325 mg/day reduced the development of adenomas by up to 35% after a few years of use.3,4 In one trial, greater effects were seen against more advanced lesions.3 Colorectal adenomas are established as common non-obligate precursors of colorectal cancer.5 Within the last decade, ACF have been identified in rodent models of carcinogenesis, and have been proposed as precursors of colorectal adenomas and cancers.6 With the use of recently developed high resolution/magnifying endoscopes, researchers are now making quantitative (for example, number, size, crypt multiplicity) and qualitative (for example, morphology) real time in vivo assessments of ACF in humans.7 ACF—or at least a subset of them—may represent important risk markers for adenoma-carcinoma development in humans. They …

Relevant and feasible surrogate end-points are needed for the evaluation of intervention strategies against cancer and other chronic, life-threatening diseases. Carcinogenesis can be viewed as a process of progressive disorganization. This process is characterized by the accumulation of genotypic lesions and corresponding tissue and cellular abnormalities, including loss of proliferation and apoptosis controls. Potential surrogate end-points for cancer incidence include both phenotypic and genotypic biomarkers of this progression. In the US National Cancer Institute chemoprevention programme, histological modulation of a precancer (intraepithelial neoplasia) has so far been the primary phenotypic surrogate end-point in chemoprevention trials. Additionally, high priority has been given to biomarkers measuring specific and general genotypic changes correlated with the carcinogenesis progression model for the targeted cancer (e.g., progressive genomic instability as measured by loss of heterozygosity or amplification at specific microsatellite loci). Other potential surrogate end-points include proliferation and differentiation indices, specific gene and general chromosome damage, cell growth regulatory molecules, and biochemical activities (e.g., enzyme inhibition). Serum biomarkers thought to be associated with cancer progression (e.g., prostate-specific antigen) are particularly appealing surrogate end-points because of accessibility. Potentially chemopreventive effects of the test agent may also be measured (e.g., tissue and serum estrogen levels in studies of steroid aromatase inhibitors). To establish chemopreventive efficacy, prevention of virtually all biomarker lesions, or of those lesions with particular propensity for progression, may be required. Ideally, the phenotype and genotype of any new or remaining precancers in the target tissue of chemopreventive agent-treated subjects would show less, and certainly no greater, potential for progression than those of placebo-treated subjects.

Esophageal cancer, with an estimated number of 12,300 new cases in the year 2000, is relatively uncommon in the United States but produces a high number of annual deaths, estimated at 12,100. Moreover, the incidence of the adenocarcinoma histologic type of esophageal cancer has been rising over the past two decades. Identification of risk factors could lead to primary prevention, as well as earlier diagnosis, treatment, and increased survival. Multiple risk factors are associated with the development of esophageal adenocarcinoma. These include Barrett's esophagus, acid peptic disorders, motor disorders of the esophagus, other malignancies, medications, environmental exposures, diet, and nutrition. However, no one particular risk factor is responsible for the rising incidence of esophageal cancer. Several preventive strategies are under investigation using such agents as nonsteroidal anti-inflammatory drugs (NSAIDs), selenium, alpha-difluoromethylornithine (DFMO), and retinoids. As we gain more insight into the biology of this disease, other risk factors will hopefully be identified that will enable us to develop effective prevention strategies and, thus, reverse the current rising incidence of esophageal carcinoma.

Genetic knockout or pharmacological inhibition of cyclooxygenase-2 decreases the number and size of adenomas in mouse models of familial adenomatous polyposis. Epidemiological and clinical studies in humans indicate that the entire class of nonsteroidal anti-inflammatory drugs (NSAIDs) that inhibit both COX-1 and COX-2 enzymes are promising colon cancer chemopreventive agents. We used the Apc mutant Min mouse model to test combinations of agents that might maximize preventive benefit with minimal toxicity because they act via different mechanisms. Min mice (n = 144) were exposed to low doses of the nonselective COX inhibitor piroxicam and the ornithine decarboxylase (ODC) inhibitor difluoromethylornithine (DFMO), beginning at the time they were weaned and continuing throughout the duration of the experiment. Piroxicam at 12, 25, and 50 ppm in the diet caused dose-dependent decreases in the number of tumors in the middle and distal portions of the small intestine. This decrease in tumor multiplicity was associated with a striking decrease in the size of those tumors that did grow out. In contrast, none of the doses of piroxicam alone decreased tumor multiplicity in the proximal portion of the intestine (duodenum). Exposure to DFMO (0.5 or 1.0% in water) caused a dose-dependent decrease in tumor multiplicity in the middle and distal portions of the small intestine. However, this decreased multiplicity was not associated with a striking decrease in the size of the tumors. Combined treatment of mice with piroxicam plus DFMO was much more effective than either agent alone and resulted in a significant number of mice totally free of any intestinal adenomas (P0.001), in contrast to the 100% incidence and high multiplicity in control Min mice. In addition to this profound effectiveness in reducing tumor number, the few residual tumors in mice treated with the combined drugs were markedly smaller in size than tumors that arose from control Min mice. These experiments suggest that selective COX-2 inhibition combined with ODC inhibition is a very promising approach for colon cancer prevention. These COX-2 and ODC inhibitor drugs were not overtly toxic at the doses used when administered to mice after weaning. However, when treatment was begun in utero, the Mendelian expected progeny ratio of 1:1 that we routinely obtained in untreated control litters was no longer observed. Apc(min)/+ progeny of pregnant dams treated with piroxicam and/or DFMO were reduced in number and their ratio to Apc+/+ progeny was decreased to approximately 0.28:1. Thus, these agents are effective against adenomas that have homozygous mutation of the APC gene and also select against fetuses bearing a heterozygous mutation in the APC gene.

This paper proposes a scientific basis and possible strategy for applying surrogate end points in chemopreventive drug development. The potential surrogate end points for cancer incidence described are both phenotypic (at the tissue, cellular, and molecular levels) and genotypic biomarkers. To establish chemopreventive efficacy in randomized, placebo-controlled clinical trials, it is expected that in most cases it will be critical to ensure that virtually all of the biomarker lesions are prevented or that the lesions prevented are those with the potential to progress. This would require that both the phenotype and genotype of the target tissue in agent-treated subjects, especially in any new or remaining precancers, are equivalent to or show less progression than those of placebo-treated subjects. In the National Cancer Institute chemoprevention program, histological modulation of a precancer (intraepithelial neoplasia) has thus far been the primary phenotypic surrogate end point in chemoprevention trials. Additionally, we give high priority to biomarkers measuring specific and general genotypic changes correlating to the carcinogenesis progression model for the targeted cancer (e.g., progressive genomic instability as measured by loss of heterozygosity or amplification at a specific microsatellite loci). Other potential surrogate end points that may occur earlier in carcinogenesis are being analyzed in these precancers and in nearby normal appearing tissues. These biomarkers include proliferation and differentiation indices, specific gene and general chromosome damage, cell growth regulatory molecules, and biochemical activities (e.g., enzyme inhibition). Serum biomarkers also may be monitored (e.g., prostate-specific antigen) because of their accessibility. Potentially chemopreventive drug effects of the test agent also may be measured (e.g., tissue and serum estrogen levels in studies of steroid aromatase inhibitors). These initial studies are expected to expand the list of validated surrogate end points for future use. Continued discussion and research among the National Cancer Institute, the Food and Drug Administration, industry, and academia are needed to ensure that surrogate end point-based chemoprevention indications are feasible.

Mounting evidence suggests that lipoxygenase (LO)-catalyzed products have a profound influence on the development and progression of human cancers. Compared with normal tissues, significantly elevated levels of LO metabolites have been found in lung, prostate, breast, colon, and skin cancer cells, as well as in cells from patients with both acute and chronic leukemias. LO-mediated products elicit diverse biological activities needed for neoplastic cell growth, influencing growth factor and transcription factor activation, oncogene induction, stimulation of tumor cell adhesion, and regulation of apoptotic cell death. Agents that block LO-catalyzed activity may be effective in preventing cancer by interfering with signaling events needed for tumor growth. In fact, in a few studies, LO inhibitors have prevented carcinogen-induced lung adenomas and rat mammary gland cancers. During the past 10 years, pharmacological agents that specifically inhibit the LO-mediated signaling pathways are now commercially available to treat inflammatory diseases such as asthma, arthritis, and psoriasis. These well-characterized agents, representing two general drug effect mechanisms, are considered good candidates for clinical chemoprevention studies. One mechanism is inhibition of LO activity (5-LO and associated enzymes, or 12-LO); the second is leukotriene receptor antagonism. Although the receptor antagonists have high potential in treating asthma and other diseases where drug effects are clearly mediated by the leukotriene receptors, enzyme activity inhibitors may be better candidates for chemopreventive intervention, because inhibition of these enzymes directly reduces fatty acid metabolite production, with concomitant damping of the associated inflammatory, proliferative, and metastatic activities that contribute to carcinogenesis. However, because receptor antagonists have aerosol formulations and possible antiproliferative activity, they may also have potential, particularly in the lung, where topical application of such formulations is feasible.

Recently, several promising strategies have been advanced for improving the efficiency of new agent development. These include pharmacokinetic/pharmacodynamic (PK/PD) and intermediate endpoint biomarker (IEB) monitoring. Here, we review their essential role as practical tools for guiding the evaluation of agents for cancer chemoprevention (CP) and provide examples of CP agents that utilize these approaches. Several important categories of IEBs are delineated, including histologically based (intraepithelial neoplasias and nuclear morphometry). The use of select IEBs combined with a Bayesian method for clinical trial monitoring for rapid identification of ineffective or promising agents is discussed. The similarities between IEB and TDM are described. Finally, we present future tools for enhanced monitoring of CP agents that will impact on laboratory medicine and are also applicable to many other drug classes, e.g., laser capture microdissection and cDNA chip microarrays that assess gene expression patterns of precancerous and cancerous lesions.

Chemoprevention has four goals: (1) inhibition of carcinogens, (2) logical intervention for persons at genetic risk for cancer, (3) treatment of precancerous lesions, and (4) confirmation and translation of leads from dietary epidemiology into intervention strategies. The National Cancer Institute has described a multidisciplinary, cancer science-based program for chemopreventive drug development that addresses these objectives, and has collaborated with the US Food and Drug Administration to provide consensus guidance for applying this approach. A critical component is the identification and characterization of intermediate biomarkers of cancer and their validation as surrogate end points for cancer incidence in clinical chemoprevention trials. More than 40 agents in the program are currently on the clinical development path (preclinical toxicology and phase I clinical safety studies or phase II/III efficacy trials), with the major effort in phase II studies to identify and characterize intermediate biomarkers. The continually advancing knowledge of molecular and tissue-based carcinogenesis mechanisms will provide leads to new chemopreventive agents with increased specificity for carcinogenesis-related activities and, hence, reduced toxicity by virtue of minimal effects on normal cell and tissue functions. Results from the Human Genome Project will help identify and evaluate the potential for chemopreventive intervention in cohorts at genetic risk and will provide specific target lesions for intervention strategies.

Strategies for chemopreventative drug development are based on the use of well-characterized agents, intermediate biomarkers correlating to cancer incidence, and suitable cohorts for efficacy studies. Since chemoprevention is applied over the long-term, chemopreventive drugs must have low toxicity. Strategies for enhancing chemopreventive drug efficacy and minimizing toxicity include combinations of drugs with complementary mechanisms and/or synergistic activity; coadministration of drugs to counter the toxicity of the chemopreventive agents; and pursuit of related compounds that retain efficacy with reduced side effects. Because of its slow development, cancer is not a feasible end point for clinical evaluation of chemoprevention, and so intermediate biomarkers that can serve as surrogate end points are crucial. Particularly important biomarkers are the morphometric and cytometric changes defining intraepithelial neoplasia (IEN). Cohorts for chemoprevention trials should have high incidences of the cancer or intermediate biomarker(s) under study within the trial duration.

In vivo studies were conducted to compare the activity of three non-steroidal anti-inflammatory drugs as inhibitors of urinary bladder carcinogenesis induced in B6D2F1 (BDF) mice by N-butyl-N-(4-hydroxybutyl)nitrosamine (OH-BBN). Mice received continuous dietary exposure to non-toxic doses of aspirin, sulindac or ketoprofen beginning 1 week prior to the first of eight weekly doses of 7.5 mg OH-BBN; studies were terminated at 24 weeks after the first carcinogen dose. Both dose levels of sulindac (200 and 400 mg/kg diet) and both dose levels of ketoprofen (40 and 80 mg/kg diet) reduced the incidence of transitional cell carcinoma of the urinary bladder by70% from that seen in dietary controls. The high dose of sulindac conferred the greatest protection against bladder cancer induction. In contrast, when administered at 400 and 800 mg/kg diet aspirin was inactive as a chemopreventive agent in the OH-BBN/BDF bladder cancer model. The significant potency of sulindac and ketoprofen as inhibitors of urinary bladder carcinogenesis, when considered with their history of safe human use, suggests that these agents merit further study as drugs for cancer chemoprevention in this target tissue.

Possible chemopreventive mechanisms include carcinogen-blocking activities, antioxidant/anti-inflammatory activities and antiproliferation/antiprogression activities. Carcinogen-blocking activities encompass inhibition of carcinogen uptake, inhibition of carcinogen formation or activation, deactivation or detoxification of carcinogens, prevention of carcinogen binding to DNA, and enhancement of the level or fidelity of DNA repair. Antioxidant/anti-inflammatory activities include scavenging of reactive electrophiles and oxygen radicals, and inhibition of arachidonic acid metabolism. Antiproliferation/antiprogression activities comprise modulation of signal transduction, modulation of hormonal and growth factor activity, inhibition of aberrant oncogene activity, inhibition of polyamine metabolism, induction of terminal differentiation, restoration of immune responses, enhancement of intercellular communication, restoration of tumour suppressor function, induction of apoptosis, telomerase inhibition, correction of DNA methylation imbalances, inhibition of angiogenesis, inhibition of basement membrane degradation, and activation of antimetastasis genes. In evaluating the potential efficacy of chemopreventive agents several mechanistic parameters are weighed: (1) the number of chemoprevention-related pharmacological activities, (2) the impact of the agent on likely carcinogenesis pathways to the targeted cancer, (3) pharmacodynamics, and (4) specificity for chemopreventive activity compared with interference with normal cellular function. Mechanistic data are important throughout the development process for chemopreventive drugs, and they are particularly important in the earlier phases of identifying promising candidate agents and characterizing efficacy. In vitro mechanistic assays are a first step in evaluating chemopreventive potential. Mechanistic considerations are also useful in defining animal efficacy models and in interpreting the results of assays in these models. Mechanistic data are also applied in designing short-term Phase II clinical chemoprevention trials that use reductions in intermediate biomarkers of cancer rather than cancer incidence as end points. The basis for identifying and evaluating these biomarkers is in understanding carcinogenesis and chemopreventive mechanisms.