In a recent issue of the Journal, Yang and Duerksen-Hughes present a new in vitro approach for identifying genotoxic carcinogens. In this study they used p53 induction in vitro as an indicator of genotoxic damage. The authors suggest that this endpoint could be used as an effective tool for identifying environmental carcinogens. In our opinion, p53 induction has clear weaknesses as an endpoint for in vitro genotoxicity testing. It is induced by many types of DNA damage and the most serious problem, which has not been addressed by the authors, are the data on p53 stabilizing mechanisms not related to DNA damage. Thus, p53 can be induced by factors interfering with the degradation of p53, and it was recently shown that inhibition of the ubiquinone pathway by proteasome inhibitors induces p53 and p53-dependent apoptosis (1). In addition, there are mechanisms involving alterations in p53 binding proteins. For example, inhibition of MDM2 gene expression by antisense oligonucleotide treatment (2) or altered MDM2 phosphorylation (3) may influence p53 induction and apoptosis. Also, hypoxia, a known p53 stabilizing factor, induces p53 as a result of hypoxiainducible factor 1α activation (4). Furthermore, ribonucleotidyl depletion induces p53 in the absence of detectable DNA damage (5). Considering this plethora of p53 inducing mechanisms and the wide range of effects that has been ascribed to toxins, the suggested endpoint seems far too unspecific. To illustrate our point, we can use a simple inorganic compound, selenite. The redox cycling metabolites of selenite may have at least two effects of interest: they may produce superoxide which can damage DNA and they may induce hypoxia (6). In addition, selenite may inhibit protein synthesis, which can attenuate any form of induction (7). It has long been known that additional in vitro tests for genotoxicity do not necessarily add predictive power when testing chemicals for carcinogenicity (8). In order to improve in vitro testing, it seems most reasonable to develop specific tests, ideally one test for each type of DNA damage. It may be added that data on in vivo genotoxicity is often lacking in risk assessment. A critical question is whether a compound, shown to be genotoxic in in vitro tests, induces genotoxicity in the target organ or in the target cell type. It is possible that histological data on in vivo p53 expression might prove useful in this situation, especially negative data suggesting lack of induction. Wild-type p53, stabilized by DNA damage, can readily be visualized in many organs, including liver, by sensitive immunohistochemical techniques (9).
[1]
U. Stenius,et al.
Wild-type p53 expression in liver tissue and in enzyme-altered foci: an in vivo investigation on diethylnitrosamine-treated rats.
,
1998,
Carcinogenesis.
[2]
L. Neckers,et al.
Stabilization of wild-type p53 by hypoxia-inducible factor 1α
,
1998,
Nature.
[3]
Jiandong Chen,et al.
Synergistic activation of p53 by inhibition of MDM2 expression and DNA damage.
,
1998,
Proceedings of the National Academy of Sciences of the United States of America.
[4]
J. Turchi,et al.
Mdm-2 phosphorylation by DNA-dependent protein kinase prevents interaction with p53.
,
1997,
Cancer research.
[5]
U. Lopes,et al.
p53-dependent Induction of Apoptosis by Proteasome Inhibitors*
,
1997,
The Journal of Biological Chemistry.
[6]
G. Wahl,et al.
A reversible, p53-dependent G0/G1 cell cycle arrest induced by ribonucleotide depletion in the absence of detectable DNA damage.
,
1996,
Genes & development.
[7]
J. Högberg,et al.
Studies of the role of DNA fragmentation in selenium toxicity.
,
1988,
Biochemical pharmacology.
[8]
B H Margolin,et al.
Prediction of chemical carcinogenicity in rodents from in vitro genetic toxicity assays.
,
1987,
Science.
[9]
P. Emmelot,et al.
Inhibition of in vitro amino acid incorporation by sodium selenite.
,
1974,
Biochemistry.