© 2004 by Oxford University Press
© 2004 Oxford University Press
REVIEW |
Apoptosis as a Novel Target for Cancer Chemoprevention
Affiliations of the authors: Department of Thoracic/Head and Neck Medical Oncology, The University of Texas M.D. Anderson Cancer Center, Houston, TX
Correspondence to: Reuben Lotan, PhD, Department of Thoracic/Head and Neck Medical Oncology, Box 432, The University of Texas M.D. Anderson Cancer Center, 1515 Holcombe Blvd., Houston, TX 77030-4095 (e-mail: rlotan{at}mdanderson.org)
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ABSTRACT |
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 TopNotes Abstract IntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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Cancer chemopreventive agents are typically natural products or their synthetic analogs that inhibit the transformation of normal cells to premalignant cells or the progression of premalignant cells to malignant cells. These agents are believed to function by modulating processes associated with xenobiotic biotransformation, with the protection of cellular elements from oxidative damage, or with the promotion of a more differentiated phenotype in target cells. However, an increasing number of chemopreventive agents (e.g., certain retinoids, nonsteroidal anti-inflammatory drugs, polyphenols, and vanilloids) have been shown to stimulate apoptosis in premalignant and malignant cells in vitro or in vivo. Apoptosis is arguably the most potent defense against cancer because it is the mechanism used by metazoans to eliminate deleterious cells. Many chemopreventive agents appear to target signaling intermediates in apoptosis-inducing pathways. Inherently, the process of carcinogenesis selects against apoptosis to initiate, promote, and perpetuate the malignant phenotype. Thus, targeting apoptosis pathways in premalignant cells—in which these pathways are still relatively intact—may be an effective method of cancer prevention. In this review, we construct a paradigm supporting apoptosis as a novel target for cancer chemoprevention by highlighting recent studies of several chemopreventive agents that engage apoptosis pathways.
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INTRODUCTION |
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TopNotesAbstractIntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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Cancer prevention has become an integral part of cancer control. Common prevention approaches include avoiding exposure to known cancer-causing agents, enhancing host defense mechanisms against cancer, modifying life styles, and chemoprevention. The National Cancer Institute has made cancer prevention research a priority in such diverse areas as early detection and screening, diet and nutrition, cessation of tobacco use, and chemoprevention. Cancer prevention programs are increasingly being added to the multidisciplinary, collaborative efforts at mainstream research institutions, and efforts have intensified to attract researchers to this field.
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CARCINOGENESIS, CHEMOPREVENTION, and APOPTOSIS |
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TopNotesAbstractIntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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Even with improvements in the early detection and treatment of cancer, overall mortality rates for most cancers of epithelial origin have not declined in the last 30 years (1). Carcinomas account for more than 80% of human cancers, with skin, lung, colon, breast, prostate, and uterus being the most frequent sites. Carcinogenesis can be viewed as a process that involves accelerated, and abnormal, cellular changes in which the genes controlling proliferation, differentiation, and apoptosis are transformed under selective environmental pressures (2). Tumor development follows three distinct phases: initiation, promotion, and progression (3,4). The initiation phase is a rapid (within hours or days), irreversible event that occurs when a normal cell is exposed to a carcinogen that causes unrepairable or misrepaired DNA damage. DNA damage itself is not mutagenic unless the resulting somatic mutation is recapitulated via mitosis to yield a clone of the mutated cell. This promotion phase, a protracted process that may require several years or decades to establish, consists of the expansion of mutated cells to form an actively proliferating, multicellular premalignant lesion. During the progression phase, another irreversible event occurs over a relatively short period, perhaps less than 1 year, in which new clones with increased proliferative capacity, invasiveness, and metastatic potential are produced (5). Because the initiation and progression phases are irreversible and relatively transient events, the promotion phase of carcinogenesis may provide the best targets for cancer prevention (6,7).
The term "chemoprevention" was coined by Michael Sporn in 1976, when he referred to the prevention of the development of malignancy by vitamin A and its synthetic analogs, known collectively as retinoids. Sporn (6) contended that the process of carcinogenesis had the potential to be controlled physiologically or pharmacologically during its preneoplastic stages, in which he suggested that the promotion phase could be stabilized, arrested, or reversed (6). Cancer chemoprevention has emerged as an important means of modulating the process of carcinogenesis (1,8). Almost three decades of research indicate that this strategy is promising with respect to reducing the incidence of cancer in well-defined high-risk groups and in the general population (8,9). Chemoprevention, by definition, is the use of agents to slow the progression of, reverse, or inhibit carcinogenesis, thereby lowering the risk of developing invasive or clinically significant disease (1,6,8). Consequently, an effective chemopreventive agent should intervene early in the process of carcinogenesis to eliminate premalignant cells before they become malignant (1,10–12).
Several thousand agents have been reported to have chemopreventive activity, and more than 40 promising agents and agent combinations are currently being evaluated clinically for cancer chemoprevention (12). For example, natural and synthetic retinoids have been effective in arresting or reversing premalignant lesions, such as bronchial metaplasia, oral leukoplakia, uterine cervical dysplasia, and actinic keratoses (1). In randomized trials of patients with familial adenomatous polyposis, the cyclooxygenase-2 inhibitors sulindac and celecoxib inhibited the growth of adenomatous polyps and promoted polyp regression (13). Furthermore, chemoprevention trials have shown that the antiestrogen tamoxifen can reduce the incidence of breast cancer (14) and that the antioxidant vitamin E can reduce the incidence of prostate cancer (8).
Many chemopreventive agents (e.g., retinoids and antiestrogens) are believed to block or delay the progression of transformed cells by modulating cell proliferation or differentiation (1,10,12). Because these agents are thought to promote cytostatic effects, it has been suggested that they should be administered long-term to healthy individuals who have an increased cancer risk. Of course, in this modality, even minor adverse side effects would be unacceptable (10). Long-term toxicity and the possibility of developing resistance to chemopreventive agents are formidable obstacles that could limit the feasibility and success of conventional chemoprevention for many cancers. An alternate chemopreventive approach entails the use of agents that quickly eliminate premalignant cells by inducing them to undergo apoptosis rather than merely slowing their proliferation and/or promoting some degree of differentiation. For example, premalignant lesions could be eradicated and secondary primary tumors prevented with chemopreventive agents that have the capacity to trigger apoptosis in transformed cells. By shifting the outcome of chemoprevention from cytostasis or differentiation to apoptosis, chronic exposure to a particular chemopreventive agent would not be necessary, thereby limiting the risk of long-term toxicity and/or the development of chemoresistance. Novel approaches to drug delivery could also facilitate chemopreventive agent-induced apoptosis in target cells and reduce possible short-term adverse side effects.
Apoptosis is the mechanism used by metazoans to regulate tissue homeostasis through the elimination of redundant or potentially deleterious cells. Apoptosis induction is arguably the most potent defense against cancer. For example, immune system cells destroy cancerous cells (15,16), and most chemotherapeutic agents inhibit tumor cell proliferation (17,18), by inducing apoptosis. The cellular machinery associated with apoptosis is highly conserved, with many similarities existing between phylogenetically divergent species. This similarity may explain why mutations in genes that regulate apoptosis pathways (e.g., p53, Bcl-2 family members, and PTEN) are common in most human cancers, and it underscores the importance of apoptosis resistance in the process of carcinogenesis (19–21). Furthermore, growing evidence suggests that certain chemopreventive agents can trigger apoptosis in transformed cells in vivo and in vitro, which appears to be associated with their effectiveness in modulating the process of carcinogenesis.
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IN VIVO OBSERVATIONS SUPPORTING APOPTOSIS AS A TARGET FOR CHEMOPREVENTIVE AGENTS |
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TopNotesAbstractIntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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Animal studies have demonstrated that certain chemopreventive agents (Table 1) can induce apoptosis in tumor cells in vivo. In the TRAMP mouse model for prostate cancer (22), treatment with oral infusions of a polyphenolic extract isolated from green tea, at doses equivalent to the human consumption of six cups of green tea per day, promoted a statistically significant inhibition of prostate cancer development and increased overall survival. The chemopreventive intervention in prostate carcinogenesis with the green tea extract also caused a statistically significant induction of apoptosis in prostate cancer cells. This activity reportedly reduced the dissemination of these cells, thereby inhibiting their progression and possible metastasis to distant organ sites (22). Oral pretreatment of SKH-1 mice with lyophilized green tea solids for 2 weeks enhanced the ultraviolet (UV)-induced increases in the number of p53-positive cells, p21-positive cells, and apoptotic sunburn cells in the epidermis. Thus, green tea treatment stimulated early adaptive responses to UV irradiation in mouse epidermis by the induction of tumor suppressor genes (i.e., p53 and p21) and the enhancement of apoptosis in vivo (i.e., inducing apoptotic sunburn cells) (23). Furthermore, oral administration of a polyphenolic black tea extract to mice with chemically induced skin tumors inhibited proliferation and enhanced apoptosis in transformed skin cells (24).
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In a mouse model for chemically induced liver carcinogenesis, dietary supplementation with epigallocatechin gallate (a polyphenol isolated from tea leaves) or caloric restriction diminished the frequency and expansion of hepatic lesions by stimulating the induction of apoptosis in transformed hepatocytes (25). Topical applications of caffeine or epigallocatechin gallate inhibited carcinogenesis and selectively increased apoptosis in UVB-induced skin tumors in mice (26). In a rat model for liver carcinogenesis (27), S-adenosyl-L-methionine reduced the incidence of 1,2-dimethylhydrazine/orotic acid-induced hepatic tumors. The decrease in tumor incidence was accompanied by an increase in apoptotic cells in the residual hepatic lesions.
In the Min/+ mouse, an animal with a germline mutation in the adenomatous polyposis coli gene that is essential for normal intestinal cell growth and differentiation, aspirin decreased the spontaneous rate of colon tumor formation by 44% (28). Aspirin also normalized enterocyte growth by increasing apoptosis and reducing the proliferation of preneoplastic intestinal mucosa cells (28). During chemically induced colon carcinogenesis, male F344 rats fed a diet supplemented with perillyl alcohol (a monoterpene isolated from lavender) exhibited a statistically significant reduction in the incidence and multiplicity of invasive adenocarcinomas of the colon compared with animals fed the control (no intervention) diet. Histopathologic evaluation of the treated colons indicated that the chemopreventive activity of perillyl alcohol was mediated through the induction of apoptosis in tumor cells (29). In a related study, the chemopreventive properties of sulindac, curcumin, and phenylethyl-3-methylcaffeate were associated with apoptosis induction in tumor cells from chemically induced colon tumors in male F344 rats. Dietary administration of sulindac, curcumin, or phenylethyl-3-methylcaffeate was associated with a statistically significant increase in the number of apoptotic tumor cells relative to that in animals fed the control diet (30). The flavonoids quercetin and rutin were also examined (31) in the male F344 rat model for chemically induced colon tumors. Either agent given as a dietary supplement increased the frequency of apoptotic cells and caused a redistribution of these cells along the colon crypt axis in the focal area of dysplasia relative to that in animals fed the control diet.
-Difluoromethylornithine (an ornithine decarboxylase inhibitor) was examined as a chemopreventive agent in N-nitrosomethylbenzylamine-induced esophageal cancer in Zn2+-deficient rats (32). Chronic exposure to -difluoromethylornithine in the drinking water substantially reduced the incidence of esophageal tumors from 89% to 10%. Esophageal cells from -difluoromethylornithine-treated animals, compared with esophageal cells from animals given drinking water alone, had increased induction of apoptosis, enhanced expression of the proapoptotic Bcl-2 family member Bax, and reduced expression of proliferating-cell nuclear antigen (32). Dietary administration of the mitochondrial poisons rotenone or capsaicin during 4-nitroquinoline 1-oxide-induced tongue carcinogenesis in rats reduced tumor formation by triggering apoptosis in the transformed cells (33), and dietary administration of rotenone inhibited diethylnitosamine-induced hepatic lesions in mice, again apparently by triggering apoptosis in the transformed hepatocytes (34).
Human chemoprevention trials have also demonstrated an association between a clinical response and the induction of apoptosis in tumor cells (35–37). In a phase I trial of the nonsteroidal anti-inflammatory drug exisulind as a chemopreventive agent in patients with familial adenomatous polyposis (37), increased apoptosis in colon polyps was noted at the maximum tolerated dose. However, this dose appeared to have no statistically significant effect on cell proliferation in the colon polyps or in the frequency at which these lesions were detected. In a prospective pilot study examining the chemopreventive effects of the nonsteroidal anti-inflammatory drug mesalazine in patients with colorectal cancer (35), induction of apoptosis was increased statistically significantly in tumor samples but was unchanged in normal intestinal mucosa. Cell proliferation in the malignant and normal tissue samples was hardly affected by mesalazine, indicating that this agent selectively induced apoptosis in tumor cells without affecting normal cell proliferation. The apparent selectivity in apoptosis induction (35,37) without the inhibition of cell proliferation in transformed intestinal mucosa cells may be dependent on the cell type and/or on the particular chemopreventive agent that was examined in these clinical studies.
Another study (36) examining the effects of mesalazine on apoptosis and proliferation in the uninvolved intestinal mucosa of patients with sporadic polyps of the large bowel reported a statistically significant increase in the frequency of apoptotic cells shortly (i.e., 1–3 days) after the initiation of treatment. The increase in apoptosis remained largely unchanged after longer exposures to mesalazine (up to 14 days). Cell proliferation in the intestinal mucosa was decreased more in the mesalazine treatment group than in the untreated groups.
Thus, these studies illustrate that certain chemopreventive agents can induce apoptosis in tumor cells in various animal models for carcinogenesis and in human chemoprevention trials. Moreover, the induction of apoptosis in tumor cells was apparently associated with the antitumor activity of the respective chemopreventive agents.
The design of clinical chemoprevention trials is evolving, but a few generalities can be defined for each phase (38). A phase I study determines the dose-related safety of a prospective chemopreventive agent and assesses the pharmacokinetics and the toxicity of the agent. Initial doses and schedules in phase I trials are based on toxicity and efficacy data obtained from preclinical studies. Phase II trials use a randomized, blinded, placebo-controlled design to evaluate the dose-response relationship and common toxic side effects that are likely to be associated with long-term (i.e., 3 months or longer) administration of the chemopreventive agent. Several dose levels are evaluated and compared with the modulation of previously validated surrogate end point biomarkers (SEBs). SEBs are defined as measurable biologic processes or molecules that are closely linked to the progression pathway to invasive cancer and that undergo modulation in concert with neoplastic regression (39). Phase II trials can be conducted with individuals who have premalignant lesions or with former cancer patients at risk of developing a second primary tumor. If safety and efficacy are judged satisfactory in these trials, the agent proceeds to evaluation in a randomized, prospective phase III clinical trial. The phase III trial is the ultimate test of a chemopreventive agent efficacy and is designed to measure the incidence of primary tumors and changes in SEBs in relation to dose and toxicity. The longer phase III trials require strict adherence to the chemoprevention protocol by patients and reproducibility in the chemopreventive agent’s formulation.
Although large-scale randomized trials are the gold standard for testing the efficacy of cancer chemopreventive agents and, indeed, of anticancer agents that act at other stages in the cancer process, these trials typically require large study populations, extensive resources, and many years for completion (40). The use of valid SEBs for the intermediate effects of cancer chemoprevention (e.g., apoptosis induction or the loss of clonogenicity) would make it possible to design smaller, short-term prevention trials. SEBs can be assessed by molecular biology techniques, such as immunohistochemistry, applied to tissue specimens obtained by excision, punch, or core-needle biopsy examination. It is likely that a panel of SEBs, rather than a single SEB, will be more advantageous to monitor chemopreventive agent-induced apoptosis in patient samples. The most commonly used SEB for apoptosis induction in the chemoprevention clinical trials discussed thus far is DNA fragmentation. In situ staining for DNA fragmentation can be coupled with an indicator of cellular proliferation (e.g., proliferating-cell nuclear antigen, DNA ploidy, or Ki-67 antigen expression) to determine whether tissue homeostasis has been restored as a result of the chemopreventive intervention (12,32). Other SEBs that can be used to detect chemopreventive agent-induced apoptosis in patient tissue samples include activation of caspases [i.e., cysteine proteases involved in apoptosis, such as the cleavage and activation of caspase-3 (41)], the cleavage of cytokeratin 18 by caspases (42), and enhanced expression of Fas and/or Fas ligand (43). Of course some of the major challenges associated with the use of SEBs in chemoprevention trials will be the standardization of the methods for measuring these intermediate biologic changes as well as the terminology used to describe them (39).
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POTENTIAL MECHANISMS ASSOCIATED WITH CHEMOPREVENTIVE AGENT-INDUCED APOPTOSIS |
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TopNotesAbstractIntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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Apoptosis is triggered by an initiation phase that is highly dependent on cell type and apoptotic stimuli (e.g., oxidative stress, DNA damage, ion fluctuations, and cytokines). In the subsequent effector phase, the cell undergoes distinct biochemical changes that result in the systematic activation of catabolic hydrolases (i.e., proteases and nucleases). These enzymes participate in the degradation phase of apoptosis through the cleavage of proteins and DNA (15,44). Most of the recent advances in the elucidation of apoptosis pathways have come about through the characterization of the effector mechanisms. Effector mechanisms of apoptosis have several components, and two effector mechanisms associated with caspase activation have been characterized extensively—the extrinsic, or death receptor-mediated, effector mechanism and the intrinsic, or mitochondrial-mediated, effector mechanism (45). In addition to mitochondria, other organelles, including the endoplasmic reticulum, Golgi apparatus, and lysosomes, may also have a role in damage sensing, proapoptotic signaling, and caspase activation (46).
The extrinsic pathway of apoptosis is activated at the cell surface when a specific ligand binds to its corresponding cell surface death receptor (Fig 1, A). Death receptors (e.g., tumor necrosis factor receptor [TNFR], TNF-related apoptosis-inducing ligand [TRAIL] receptor, and Fas) belong to the TNFR superfamily (47). After ligand binding (e.g., TNF, TRAIL, and Fas ligand, respectively), death receptors cluster in the plasma membrane and promote the recruitment of adapter proteins (47). Caspase 8 is an apical caspase in the death receptor pathway. The zymogen of caspase 8 can interact with the adapter proteins (e.g., FADD and RIP1) to generate the active form of caspase 8 (47). After activation, caspase 8 can trigger the activation of downstream effector caspases such as caspase 3. In certain cell types, the activation of caspase 8 is sufficient to initiate the proteolytic cascade that results in apoptotic cellular degradation (15,47).
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In addition to activating effector caspases, caspase 8 also targets Bid, a proapoptotic member of the Bcl-2 family. In response to the binding of Fas ligand or TNF to its receptor, caspase 8 induces the cleavage of Bid to yield a truncated C-terminal fragment that translocates from the cytosol to the outer mitochondrial membrane. Oligomers of the C-terminal Bid fragment can trigger pore formation in the outer mitochondrial membrane, allowing apoptogenic mitochondrial proteins, such as cytochrome c (48,49) and endonuclease G (50,51), to be released from the mitochondria (Fig. 1, A). Truncated Bid may also trigger conformational changes in Bax (another proapoptotic member of the Bcl-2 family) that allow it to localize in the outer mitochondrial membrane (52), where Bax can associate with the voltage-dependent anion channel that is also located in the outer mitochondrial membrane. Complexes of Bax and the voltage-dependent anion channel promote pore formation in the outer mitochondrial membrane, allowing the release of apoptogenic mitochondrial proteins (52). Regardless of the mechanism of caspase-8-induced permeabilization of the outer mitochondrial membrane, the presence of apoptogenic mitochondrial proteins in the cytosol can serve to amplify intracellular signals activated by death receptors to trigger apoptosis.
The intrinsic pathway of apoptosis relies solely on the permeabilization of mitochondrial membranes to release the apoptogenic mitochondrial proteins [e.g., cytochrome c (53), endonuclease G (50,51), Smac/DIABLO (54), Omi/HtrA2 (55), apoptosis-inducing factor (AIF) (56), and its homolog AIF-homologous mitochondrion-associated inducer of death (AMID) (57)] required for caspase activation and apoptosis. As they do in the extrinsic pathway, Bcl-2 family members appear to play an important role in the regulation of the intrinsic pathway (Fig. 1, B). During conditions of cell stress, antiapoptotic Bcl-2 family members (e.g., Bcl-2 and Bcl-XL) residing in the outer mitochondrial membrane can be destabilized by the induction of proapoptotic Bcl-2 family members (e.g., Bax, Bad, and Bak). In this scenario, the ratio of proapoptotic Bcl-2 family members to antiapoptotic Bcl-2 family members increases and, by mechanisms that are not completely understood, pores form in the outer mitochondrial membrane, liberating apoptogenic mitochondrial proteins to activate caspases and induce apoptosis (58,59).
Another mechanism implicated in the permeabilization of mitochondrial membranes is the mitochondrial permeability transition. The mitochondrial permeability transition is a rate-limiting and self-amplifying process that is regulated at various levels by several mitochondrial proteins. Many of these proteins are believed to constitute the permeability transition pore complex (17,60) (Fig. 1, B). Normally, proteins in the outer and inner mitochondrial membranes that constitute the permeability transition pore complex are predictably in close proximity to each other and are in a closed or low-conductance conformation (60). Numerous pathologic stimuli (e.g., reactive oxygen species and calcium) and various chemical agents can cause the permeability transition pore complex to adopt an open conformation (17,60), which allows water and solutes can infiltrate the mitochondrial matrix (61). This results in colloidal osmotic swelling of the mitochondrial matrix and permeabilization of the outer mitochondrial membrane, presumably resulting from physical rupture of the outer membrane (62). After the outer membrane fragments, apoptogenic mitochondrial proteins are released to the cytoplasm, where they participate in the degradation phase of apoptosis.
A search of PubMed for studies published since 1994 revealed that several chemopreventive agents have the ability to induce apoptosis in a variety of premalignant and malignant cell types in vitro (Table 2). For some agents, modulation of the cellular target or pathway that was originally envisioned for the agent is independent of its apoptogenic effects, which may account for the prevalent cytotoxicity of some of these agents. For example, although apoptosis triggered by the natural retinoid all-trans-retinoic acid appears to be mediated by the activation of retinoid receptors in breast cancer cells (63), induction of apoptosis by the synthetic retinoid N-(4-hydroxyphenyl)-retinamide appears to be independent of retinoid receptors (64–68). Some nonsteroidal anti-inflammatory drugs are selective inhibitors of cyclooxygenase 2. However, two such agents, sulindac (69–72) and celecoxib (73), can induce apoptosis through cyclooxygenase-2-independent mechanisms. In addition, tamoxifen has been reported to induce apoptosis in a manner that is independent of its antiestrogenic activity (74–76).
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We identified representatives from various classes of chemopreventive agents from recent in vitro studies with sufficient evidence to provide a detailed account of their apoptotic mechanisms (Table 3). Most of these compounds can activate caspases through intrinsic effector mechanisms that are regulated by Bcl-2 family members (e.g., inhibition of Bcl-2 expression or induction of Bax expression) or the mitochondrial permeability transition (e.g., dissipation of mitochondrial inner transmembrane potential). Other agents [e.g., all-trans-retinoic acid (77), sulindac (69), and epigallocatechin gallate (78)] may use extrinsic effectors instead.
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Several of the classes of chemopreventive compounds contain agents that can promote reactive oxygen species generation or trigger oxidative stress [e.g., N-(4-hydroxyphenyl)retinamide (64–68,79), celecoxib (80), indomethacin (81), epigallocatechin gallate (82), curcumin (83), tamoxifen (75,84), capsaicin (85–88), resiniferatoxin (86,89), rotenone (90,91), and deguelin (92)], which appears to be associated with apoptosis induction in various cell types (86–91). Reactive oxygen species can promote divergent cellular effects depending on the extent of their production and the enzymatic or nonenzymatic mechanisms available for their dismutation in a given cell type. Thus, reactive oxygen species can serve as mitogenic stimuli, senescence promoters, or cell death mediators (93). Mitochondria are the primary cellular site of reactive oxygen species production (93–96), and, under certain conditions, elevated mitochondrial reactive oxygen species generation can serve as an apoptotic signal (93,94,97). Many chemopreventive agents with prooxidant activity also appear to have a direct [e.g., inhibition of mitochondrial respiration (86,92,98)] and/or indirect [e.g., promotion of the dissipation of mitochondrial inner transmembrane potential (74,83,85,86,89,92,98,99)] ability to disrupt mitochondrial function to trigger the intrinsic pathway of cell death. Furthermore, enhanced ceramide production promoted by N-(4-hydroxyphenyl)-retinamide (67) and celecoxib (80) could also promote the production of reactive oxygen species, impair mitochondrial function, and trigger intrinsic effector mechanisms for cell degradation, considering that ceramide has been implicated in these processes (94). Therefore, such chemopreventive agents may induce apoptosis, at least in part, by their mitochondrial toxicity. This hypothesis does not seem to be a broad conceptual leap because several novel chemotherapeutic agents are also considered mitochondriotoxic (17).
We have observed that the apoptogenic effects of N-(4-hydroxyphenyl)retinamide (98), capsaicin (86), resiniferatoxin (86), and deguelin (92) are conspicuously diminished in skin cancer cells that have been depleted of mitochondrial DNA (i.e., 
0 cells) and that are therefore functionally deficient in mitochondrial respiration. Other studies have demonstrated that 0 cells are more resistant than their parental counterparts to the apoptogenic effects of cancer chemotherapeutic agents (100,101), ceramide (102), and extrinsic mediators of apoptosis, such as TNF (103) and TRAIL (104), implying that the disruption of mitochondrial respiration by these agents is associated with cell death. The inability of the 0 mitochondria to conduct bioenergetic processes appears to correspond to the mitochondrial alterations observed in various malignant tumor cells (105–109), implying that certain aspects of mitochondrial function [e.g., electron transport, which is inhibited by N-(4-hydroxyphenyl)retinamide, capsaicin, resiniferatoxin, and deguelin] are realistic targets for chemoprevention. However, the window of opportunity for exploiting these targets to trigger apoptosis probably closes as tumor cells progress to a more malignant phenotype.
Many chemopreventive agents can also modulate genes or proteins that respond to conditions of oxidative stress to trigger apoptosis (Table 3). For example, the induction of Fas and Fas ligand is responsive to conditions of oxidative stress (110–112), and all-trans-retinoic acid reportedly induces apoptosis by enhancing Fas ligand expression (77). Oxidative stress appears to be associated with the modulation of Bcl-2 family members in several cell systems (113–118). The studies summarized in Table 3 indicate that the induction of Bax or Bak or the inhibition of Bcl-2 or Bcl-XL are commonly associated with apoptosis induction after exposure to certain chemopreventive agents [e.g., sulindac (72), indomethacin (119), tamoxifen (120), capsaicin (121), deguelin (122), and genistein (123)]. Likewise, many of these agents can modulate nuclear factor NF-
B [e.g., tamoxifen (75), epigallocatechin gallete (124), curcumin (125), resvertrol (126), and genistein (127)], which has also been implicated in regulatory mechanisms associated with oxidative stress (114,116,128,129). Thus, some chemopreventive agents may trigger an oxidative stress response by direct or indirect effects on the mitochondria because these organelles are important regulators of cellular redox homeostasis (130).
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CONCLUSIONS AND PERSPECTIVES |
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TopNotesAbstractIntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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Apoptosis is subverted during tumorigenesis, presumably through the systematic loss of regulatory control mechanisms, ultimately resulting in the generation of a malignant phenotype and resistance to chemotherapy and radiation therapy. Investigations to further elucidate the mechanisms associated with chemopreventive agent-induced apoptosis should provide increased opportunities to develop novel, selectively targeted agents or drugs for chemoprevention.
It is anticipated that the life span of human beings will continue to increase, which will inevitably be associated with an escalation of age-related diseases such as cancer. Numerous epidemiologic studies have provided a basis for primary cancer prevention through the elimination of carcinogens from our diet and the environment. However, given that the carcinogens responsible for many forms of cancer are unknown, it seems highly unlikely that all individuals will be able to eliminate the risk of developing cancer during their lifetimes. Our knowledge of the etiology of certain familial forms of breast, colon, and skin cancer, for example, also indicates that certain individuals are predisposed to cancer irrespective of environmental factors. Advances in the last two decades have provided very sensitive methods, such as endoscopy, blood screening tests, and mutational analysis, for the early detection of cancer that allow an opportunity for intervention to prevent the development of invasive or clinically significant disease. Likewise, the potential benefit of cancer chemoprevention appears promising given the results obtained from clinical trials, animal carcinogenesis models, and in vitro studies. Collectively, these considerations support a chemopreventive approach to manage the magnitude and severity of cancer at present and in the future.
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NOTES |
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TopNotesAbstractIntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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This work was supported in part by a Cancer Prevention Fellowship (N.H.), sponsored by NCI grant R25 CA57780; and the Irving and Nadine Mansfield and Robert David Levitt Cancer Research Chair and USPHS Project Grant PO1 CA68233 from the NCI (R.L.).
The first and second authors have contributed equally to this work and should be considered co–first authors.
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TopNotesAbstractIntroductionCarcinogenesis, Chemoprevention,...In Vivo Observations Supporting...Potential Mechanisms Associated...Conclusions and PerspectivesReferences |
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