Declaring chemicals "not carcinogenic to humans" requires validation, not speculation. (Correspondence).

Regulatory agencies should provide detailed guidelines on how to use mechanistic and epidemiologic data to dismiss positive cancer evidence obtained from studies in experimental animals. Roberts and Ashby (2002) bemoaned that the U.S. Environmental Protection Agency (EPA) does not provide opportunity to conclude that there is no evidence of carcinogenicity from well-conducted and adequately powered epidemiologic studies. In their letter, Roberts and Ashby provided examples for which they claimed that there is enough information to draw valid conclusions of "no effect." Our examination of their examples and viewpoints reveals several critical issues that they have ignored, but which must be addressed in any future regulatory agency guidelines that may allow the dismissal of carcinogenic effects in animals based on mechanistic and epidemiologic data.

Roberts and Ashby (2002) appealed to the International Agency for Research on Cancer (IARC) guidelines (IARC 2000a) for evaluating epidemiologic studies to rectify the absence of procedures in the U.S. EPA's guidelines (US EPA 1999) that would allow the conclusion that there is no carcinogenic hazard based on well-conducted epidemiologic studies. However, Roberts and Ashby disregarded the IARC distinction between the classifications "inadequate evidence of carcinogenicity" and "evidence suggesting lack of carcinogenicity" in humans. Requirements for the latter are intentionally stringent, including multiple, mutually consistent, adequately powered studies covering the full range of human exposures that exclude with reasonable certainty bias, confounding, and chance and provide individual and pooled estimates of risk near unity with narrow confidence intervals. In particular, IARC (2000a) cautions that "latent periods substantially shorter than 30 years cannot provide evidence for lack of carcinogenicity."

With the IARC criteria in mind, it is surprising that Roberts and Ashby (2002) pointed to epidemiologic studies on clofibrate as an example, where they claim there is sufficient power to detect a cancer increase and that there are enough data to conclude that there is no carcinogenic effect. Although the World Health Organization trial on prevention of ischemic heart disease with clofibrate includes 208,000 man-years of observation in the mortality update (WHO 1984), the 5-year treatment period plus the 8-year mortality follow-up period is insufficient to adequately determine the extent of drug-related cancer incidences or deaths, or to make a valid conclusion of no risk of cancer. Cancer is a group of diseases that most often have latency periods greater than 20 years for clinical manifestation. Further, because there were only 206 total cancer deaths among the 5,331 men in the clofibrate group (WHO 1984), this study has inadequate power to demonstrate lack of a carcinogenic hazard, which should be evaluated for individual sites. This study also provided no information on cancer risk to women. The meta-analysis by Law et al. (1994) did not provide any estimates of cancer risk for clofibrate, though it was cited by Roberts and Ashby (2002) as part of the negative human evidence on the carcinogenicity of this hypolipidemic drug. Certainly, exposure and follow-up durations, gender differences, and study power are critical issues for regulatory agencies to address when formulating guidelines for use of epidemiologic studies to support a claim of no evidence of carcinogenicity in humans despite positive cancer data in experimental animals.

On the use of mechanistic data for evaluating chemical carcinogens, we have repeatedly stressed the need for rigorous testing of mechanistic hypotheses (Huff 1995; Melnick et al. 1997). In this regard it is instructive to examine the basis for IARC's conclusions on the relevance of the carcinogenicity of peroxisome proliferators in experimental animals. As noted by Roberts and Ashby (2002), IARC opined that the mechanism of tumor induction by clofibrate [and di(2-ethylhexyl)phthalate (DEHP)] would not be operative in humans. For DEHP, there is sufficient evidence of carcinogenicity in rats and mice, yet IARC downgraded the classification of this chemical from "possibly carcinogenic in humans" to "not classifiable as to its carcinogenicity to humans" (IARC 2000b). Downgrading DEHP was based on the working group's conclusion that in addition to peroxisome proliferation, cell proliferation induced by DEHP is due to activation of the peroxisome proliferator-activated receptor (PPAR[alpha]) and that peroxisome proliferation had not been documented in human hepatocyte cultures. However, in considering IARC's evaluation, it is important to recognize that humans possess a functional PPAR[alpha] and that the working group noted that effects of DEHP in human liver have not been adequately evaluated. Cell proliferation mediated by PPAR[alpha] was considered by the IARC working group to be the critical step in the carcinogenicity of peroxisome proliferators.

Several deficiencies and critical data gaps in the peroxisome proliferation mode-of-action hypothesis have been noted (Melnick 2001). Moreover, recent studies have shown that hepatocyte proliferation and peroxisome proliferation occur by different mechanisms, and that PPAR[alpha]-independent effects that occur in Kupffer cells are required for the induction of cell proliferation and suppression of apoptosis (Parzefall et al. 2001; Peters et al. 2001; Rose et al. 1999). Although the demonstration of the important role of Kupffer cells in peroxisome proliferator-mediated effects is fairly recent, it seems odd that this subject was not mentioned in the IARC evaluation of DEHP (IARC 2000b; Melnick 2003). Based on current information, it is clear that peroxisome proliferators cause liver tumors in rodents, but the mechanism of action is not understood (Peters et al. 2001).

Tomatis (2002) warned the occupational and environmental health communities that serious public health consequences might follow if decision-making bodies rely on untested mechanistic hypotheses that are later shown experimentally to be incorrect. Thus, when formulating guidelines for acceptance of mechanistic hypotheses, we expect regulatory agencies to require rigorous testing and validation before using incomplete data sets to downgrade the categorization of chemical carcinogens. Such guidelines must also address whether differences in mechanistic events among species are truly qualitative rather than quantitative in nature. For quantitative differences, the guidelines should also require information on the range of parameter variability in exposed humans so that sensitive subpopulations are not ignored in these categorizations. The issue is not singularly the adequacy of mechanistic or epidemiologic evidence, but certainty for protecting public health.

REFERENCES

Huff J. 1995. Mechanisms, chemical carcinogenesis, and risk assessment: cell proliferation and cancer. Am J Ind Med 27:293-300.

IARC. 2000a. Preamble IARC Monogr Eval Carcinog Risks Hum 77:9-31.

IARC. 2000b. Di(2-ethylhexyl)phthalate. IARC Monogr Evaluat Carcinog Risks Hum 77: 41-148.

Law MR, Thompson SG, Wald NJ. 1994. Assessing possible hazards of reducing serum cholesterol. Br Med J 308:373-379.

Melnick RL. 2001. Is peroxisome proliferation an obligatory precursor step in the carcinogenicity of di(2-ethylhexyl)phthalate (DEHP)? Environ Health Perspect 109:437-442.

Melnick RL. 2003. Suppression of crucial information in the IARC evaluation of DEHP [letter]. Int J Occup Environ Health 9:84-85.

Melnick RL, Kohn MC, Huff J. 1997. Weight of evidence versus weight of speculation to evaluate the [alpha]2u-globulin hypothesis. Environ Health Perspect 105:904-906.

Parzefall W, Berger W, Kainzbauer E, Teufelhofer O, Schulte-Hermann R, Thurman RG. 2001. Peroxisome proliferators do not increase DNA synthesis in purified rat hepatocytes. Carcinogenesis 22:519-523.

Peters JM, Rusyn I, Rose ML, Gonzalez FJ, Thurman RG. 2001. Peroxisome proliferator-activated receptor [alpha] is restricted to hepatic parenchymal cells, not Kupffer cells: implications for the mechanism of action of peroxisome proliferators in hepatocarcinogenesis. Carcinogenesis 21:823-826.

Roberts R, Ashby J. 2002. Mechanistic and epidemiologic data: when is enough enough? Environ Health Perspect 110:A502-A503.

Rose MI, Rivera CA, Bradford BU, Graves LM, Cattley RC, Schoonhoven R, et al. 1999. Kupffer cell oxidant production is central to the mechanism of peroxisome proliferators. Carcinogenesis 20:27-33.

Tomatis L. 2002. The IARC Monographs program: changing attitudes towards public health. Int J 0ccup Environ Health 8:144-152.

U.S. EPA. 1999. Guidelines for Carcinogenic Risk Assessment. Draft. Washington, DC: U.S. Environmental Protection Agency.

WHO. 1984. WHO cooperative trial on primary prevention of ischaemic heart disease with clofibrate to lower serum cholesterol: final mortality follow-up. Report of the Committee of Principal Investigators. Lancet 2:600-604.

Ronald L. Melnick
Freya Kamel
James Huff
National Institute of Environmental
Health Sciences
Research Triangle Park, North Carolina
E-mail: [e-mail]