Page 587 - The Toxicology of Fishes

Chemical Carcinogenesis in Fishes 567

process; for example, BNF may alter cyp, increasing detoxification and reducing initial DNA damage
by AFB . Finally, I3C may alter AFB pharmacokinetics.
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As has been presented above, DEN is a complete carcinogen in the trout and medaka models. Inhibition
of the DEN-induced hepatocarcinogenesis was demonstrated using I3C (Fong et al., 1988). To test for
possible inhibitory effects on DEN-induced hepatic tumors, trout were prefed a diet containing BNF,
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I3C, or Arochlor 1254 and, following a recovery period, were exposed to a tumorigenic concentration
of DEN in water for 24 hours. Fish were allowed to grow for 42 weeks and then assessed for liver tumor
formation. Liver DNA ethylguanine levels were reduced in I3C-pretreated fish, were increased in BNF-
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pretreated fish, and showed no significant effect with Arochlor 1254. The authors suggested that I3C
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inhibition was mediated by effects on O -ethylguanine formation and, therefore, on the initiation phase
of carcinogenesis.
Chlorophyllin (CHL), a food-grade derivative of the green plant pigment chlorophyll, proved to be a
potent, dose-responsive inhibitor of AFB –DNA adduction and hepatocarcinogenesis in the trout model
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(Breinholt et al., 1995a). These effects occurred when CHL was fed in a diet containing AFB . When
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CHL was fed after exposure to AFB , it neither enhanced nor suppressed AFB liver tumorigenesis.
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These findings suggest that CHL interferes with early events in AFB carcinogenesis. A subsequent in
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vitro study (Breinholt et al., 1995b) revealed that CHL formed a strong noncovalent complex with AFB 1
and inhibited phase I metabolism. A third study by Breinholt et al. (1999) focused on the complex
formation and in situ protective mechanisms within the target organ, liver. Bioavailability differences
rather than in situ organ inhibitory mechanisms are likely the cause of the protection afforded by CHL.
Finally, the cancer prevention properties shown by CHL against AFB were tested using another class
1
of carcinogen, the PAHs (Harttig et al., 1996). Dibenzo(a,l)pyrene (DB(a,l)P) is the most carcinogenic
of the PAHs tested in the trout model to date. Features include the occurrence of tumors of liver, stomach,
and swim bladder. Use of this compound provided the opportunity to determine whether CHL would
be effective in inhibition of multi-organ carcinogenesis. CHL reduced DB(a,l)P toxicity, inhibited
DB(a,l)P–DNA adducts, and strongly inhibited DB(a,l)P mutagenesis in the Salmonella assay.
As we have seen, inhibition of chemical carcinogenesis, or cancer chemoprevention, has been studied
only in the trout model. Each of the examples presented herein suggests that the protection occurs by
inhibition of early events, including xenobiotic uptake or distribution and alteration in phase I metabolism
favoring detoxification. The final result of these changes is a net reduction in carcinogen–DNA adduction.
This does not mean that other mechanisms are not operative in later stages, as between initiation and
promotion or progression; they simply have not been addressed. Work to date has not considered prolif-
eration, differentiation, or cell death. As presented above, only one paper has included serial analysis with
emphasis on factors governing the cell cycle (Orner et al., 1998). The general approach has been to alter
early events reducing DNA adduction and then to determine the effect of such alteration on eventual
tumor incidence. What is now increasingly possible and is a critical challenge is a careful analysis during
pathogenesis of fish neoplasia incorporating self-sufficiency in growth signals, insensitivity to growth-
inhibitory (antigrowth) signals, evasion of programmed cell death (apoptosis), limitless replicative poten-
tial, sustained angiogenesis, and tissue invasion and metastasis (Hanahan and Weinberg, 2000).

Immunologic Factors
The immune system and its role in the carcinogenesis have been studied in mammalian models, although
parallel studies in fish models have yet to be conducted. Regulatory T (T-reg) cells are an important
component of the immune system that were implicated in cancer progression as early as 1974 (Umiel
and Globerson, 1974). Subsequent work during the 1980s focused on the disruption of T-reg cells and
attempted to induce tumor immunity (Orentas et al., 2006). It is the functional inhibition of such cells
that is currently the goal of effective anticancer immune therapies in humans. To date, only a limited
number of CD4 T-cell gene homologs have been isolated from fish, including pufferfish (Fugu rubripes)
(Suetaka et al., 2006), trout (Oncorhynchus mykiss) (Dijkstra et al., 2005), sea lamprey (Petromyzon
marinus) (Pancer et al., 2004), and channel catfish (Ictalurus punctatus) (GenBank Accession Nos.
ABD93355 and ABD93354). It is noteworthy that the CD4 homologs characterized thus far display
many of the structural motifs found in mammalian counterparts. An exception is a particular motif