Exposure to peroxisome proliferators: reassessment of the potential carcinogenic hazard.

Melnick (1) recently suggested that because peroxisome proliferation is not established as an obligatory step in the carcinogenicity of peroxisome proliferators (PPs), the proposal that the peroxisome proliferator di(2-ethylhexyl)phthalate (DEHP) poses no carcinogenic risk to humans (2) due to species differences in peroxisome proliferation should be viewed as an unvalidated hypothesis (1). In this context, Melnick (1) raised the recent downgrading by the International Agency for Research on Cancer (IARC) of DEHP (3) to “not classifiable as to its risk to humans (group 3)” based on their conclusion that it produces rodent liver tumors by a mechanism involving peroxisome proliferation, which they judged to be not relevant to humans (3). As illustrated by Melnick (1), there is a large body of data to correlate the phenomenon of rodent liver peroxisome proliferation with rodent liver cancer but “published studies have not established peroxisome proliferation per se as an obligatory pathway on the carcinogenicity of DEHP” (1). This focuses attention on the need, as also suggested by O’Brien et al. (4), for a fundamental review of how PPs induce liver cancer in rodents and the relevance of these rodent tumors for humans. There are two distinct usage patterns for PPs: as drugs such as clofibrate for the treatment of hypolipidemia (5) and in nonclinical applications such as the plasticiser DEHP. Most PPs are carcinogenic to the rodent liver, and the task of assessing their human carcinogenic potential has fallen to different regulatory agencies, depending on the primary usage pattern of the particular PP in question. There seems to be little regulatory concern regarding the safety of the clinical PPs, yet continuing uncertainty regarding the safety of the nonpharmaceutical PPs. This presents an untenable situation that we suggest is unjustified. Hypolipidemic fibrates such as clofibrate and gemfibrozil have been used extensively over the past 20 years to treat cardiovascular disease and are enjoying a revival due to recent reconfirmation of efficacy (6). However, by 1980 several of these agents had become associated with a rodent-specific response known as hepatic peroxisome proliferation (7), a property shared by a number of nonpharmaceutical chemicals (8). Additionally, a link between rodent liver peroxisome proliferation and an increased risk of rodent liver carcinogenesis was emerging (7). Nonetheless, clinical side effects of the fibrate PPs are rare, and analyses of causes of death during treatment show no evidence of an adverse effect and no evidence of an increase in malignant disease compared to the normal population (5). Specifically, the carcinogenic risk to humans of gemfibrozil and clofibrate has been formally evaluated by IARC (9,10), and in the case of clofibrate, for which most clinical data exist, IARC (9) concluded that “the mechanism of liver carcinogenesis in clofibrate-treated rats would not be operative in humans.” This conclusion was based on the observation that clofibrate causes peroxisome proliferation and cell proliferation in rodent but not human hepatocytes and on the results of extensive epidemiologic studies, particularly the World Health Organization trial on clofibrate that included 208,000 man-years of observation (11,12). Further, a meta-analysis (13) of the results from six clinical trials on clofibrate also found no excess cancer mortality (9). It therefore appears that there are no remaining concerns about the human carcinogenic potential of the clinical PPs and that the rodent liver effects have been set aside as probable laboratory curiosities. However, this is not true for the nonpharmaceutical PPs; several regulatory agencies continue to be concerned about their carcinogenic potential to the human liver. The unease of these agencies is due to their belief that in the absence of a definitive mechanism of PP-induced rodent liver carcinogenesis, it is not possible to make a clear statement on the human safety of these chemicals. Nonetheless, there are now several strong lines of evidence that PPinduced rodent liver carcinogenesis is not relevant to humans, which supports the conclusion drawn by IARC for the clinically used PPs and the recent IARC decision to downgrade DEHP from group 2B (possibly carcinogenic to humans) to group 3 (3).These lines of evidence are as follows: • Direct genetic toxicity has been eliminated as a common mechanism of carcinogenic action for PPs in general (14). Thus, rodent hepatocarcinogenicity must occur via a nongenotoxic mechanism that correlates with peroxisome proliferation, although, as pointed out by Melnick (1), the hepatocarcinogenicity is unlikely to be caused by peroxisome proliferation per se, as initially suggested (15). • There are marked species differences in the induction of peroxisomes, with human hepatocytes being resistant (8,16,17). These data provide evidence that the phenomenon of PP-induced peroxisome proliferation is rodent specific. • PPs suppress rodent hepatocyte apoptosis (18–20) and induce rodent hepatocyte replication (8). This duality of effects provides a plausible mode of rodent carcinogenic action based on liver growth perturbation (21,22). As well as being resistant to peroxisome proliferation, human hepatocytes are also resistant to PP-mediated induction of replication and suppression of apoptosis (8,16,17). Whatever the precise mechanism by which PPs induce rodent liver cancer, rodent liver peroxisome proliferation, induction of the peroxisomal gene acyl CoA oxidase (ACO) (23), hypertrophy (24), and carcinogenicity (25) are all mediated through activation of the peroxisome proliferator-activated receptor (PPARα). This is illustrated dramatically by the absence of all of these responses in PPARα knockout mice treated with the PPs DEHP or Wyeth-14,643 (24–26). Although human liver expresses around 10-fold less PPARα mRNA than the rodent liver (27,28), evidence suggests that the hypolipidemic effects of the fibrate drugs in humans are also mediated by activation of PPARα, leading to regulation of the apolipoprotein (Apo) genes such as ApoA1 (29). Thus, PPARα levels in human liver may be sufficient to mediate PPinduced hypolipidemia, but insufficient to activate the gene battery associated with rodent peroxisome proliferation and cancer (30). In addition to these quantitative data, there are species differences in the molecular sequence of the PPARα response elements (PPREs) located upstream of genes associated with rodent peroxisome proliferation such as ACO. In the rat, ACO responds to PPs via a functional PPRE, whereas the equivalent gene in humans does not (31,32). Thus, the human ACO gene does not respond to PPs even in the presence of excess PPARα (31–33). Similarly, recent data have shown that PPARα cannot induce the battery of peroxisome proliferation-associated genes in human hepatoma cells (33,34). Conversely, the human ApoA1 gene is responsive to fibrate PPs, whereas the equivalent rat gene is not (35). Such findings isolate the human hypolipodaemic effects of PPs from the rodent cancer effects. Although the precise mechanism of the carcinogenic action of PPs in the rodent liver remains to be determined, all of the phenomena associated with this response of rodent hepatocytes (peroxisome proliferation, ACO gene expression, induction of cell proliferation, and the suppression of apoptosis) are absent in human hepatocytes. This body of data provides a plausible mode of carcinogenic action for the rodent liver,