Elsevier

Toxicology

Volume 130, Issue 1, 1 September 1998, Pages 29-41
Toxicology

Evaluation of omeprazole genotoxicity in a battery of in vitro and in vivo assays

https://doi.org/10.1016/S0300-483X(98)00101-2Get rights and content

Abstract

Omeprazole, a proton pump inhibitor of wide use in the treatment of gastric acid-related disorders, was evaluated for its genotoxic effects in both rat and human cultured cells and in the intact rat. DNA repair synthesis, as revealed by autoradiography, was detected in primary cultures of metabollically competent rat hepatocytes exposed to concentrations ranging from 10 to 100 mg/l, but the responses cannot be considered as clearly positive. Under the same experimental conditions any significant evidence of DNA repair was absent in primary hepatocytes from two human donors. At the same concentrations a modest but dose-related increase of micronucleated cells, that reached the level of statistical significance at 33 mg/l, was present in primary rat hepatocytes and in one of two human donors. In human lymphocytes exposed to subtoxic concentrations ranging from 0.78 to 12.5 mg/l a reproducible concentration dependent clastogenic effect was absent. In partially hepatectomized female rats treated with a single p.o. dose of 1000 mg/kg, the frequency of micronucleated cells was 5.2-fold higher than in controls in the liver, but only 2.0-fold higher in polychromatic erythrocytes of the bone marrow. In rats of the same sex given azoxymethane as initiator of colon carcinogenesis the oral administration for 8 successive weeks of 10 mg/kg omeprazole on alternate days increased the response to azoxymethane, as indicated by the occurrence in colon mucosa of a modest but statistically significant increase in both the average number and size of aberrant crypt foci. Taken as a whole, our results suggest that omeprazole behaves as a weak genotoxic agent for the rat liver. Reliable information about the potential genotoxic risk to humans requires further studies on primary cells from a wide number of donors.

Introduction

Omeprazole is a drug of worldwide use in the treatment of peptic ulcers and other gastric acid-related disorders. In the acidic canaliculi of parietal cells it is converted to a sulphenamide derivative which binds covalently to the H+, K+-ATPase and thereby produces a long-lasting inhibition of gastric acid secretion (Larsson et al., 1988). In long-term experiments carried out in rats omeprazole was found to induce, with an incidence higher in females than in males, gastric enterochromaffin-like cell hyperplasia and carcinoids in the fundus as well as an increased frequency of neoplastic hyperplastic nodules in the liver (Ekman et al., 1985, Havu, 1986), but it should be noted that the enterochromaffin-like cell hyperplasia was subsequently shown to be mediated by hypergastrinemia (Ryberg et al., 1990). The results of short-term genotoxicity tests performed to evaluate the safety of omeprazole have been previously reported and discussed (Mereto et al., 1993). In brief, omeprazole was not mutagenic to bacteria and mammalian cells, did not induce chromosomal aberrations in human lymphocytes, was negative in mouse bone marrow chromosome aberrations and micronucleus tests, and did not induce DNA fragmentation in the rat liver. In contrast, definitive conclusions cannot be drawn from the results of other studies or from their interpretation (Mereto et al., 1993). These studies concern: the computer based analysis of the relationship between chemical structure and putative genotoxic activity; the ability of omeprazole to form DNA adducts and to elicit DNA repair synthesis in the rat gastric mucosa; and the increase in the frequency of total nuclear anomalies (micronuclei, pyknosis and karyorrhexis) in the rat forestomach and descending colon mucosa. Moreover, omeprazole has been found not to induce DNA fragmentation in the rat gastric mucosa by Mereto et al. (1993), whereas Furihata et al. (1996) judged equivocal the response obtained in the same assay.

Taking into account that the available data are not sufficient to definitely exclude a genotoxic activity of omeprazole or its sulphenamide derivative, and that this drug may be administered at high doses for long periods of time, e.g. to prevent ulcerations in the Zollinger–Ellison syndrome, new information provided by short- and medium-term assays so far not performed should be considered useful for a more reliable assessment of its potential carcinogenic risk to humans. The present study was carried out to examine the capability of omeprazole of inducing: (1) DNA repair synthesis in metabolically competent primary cultures of rat and human hepatocytes; (2) micronuclei formation in human peripheral blood lymphocytes and in replicating primary rat and human hepatocytes; (3) increase of micronucleated liver cells and of micronucleated polychromatic erythrocytes (MnPCE) in the bone marrow in partially hepatectomized rats; and (4) aberrant crypt foci (ACF) in the rat colon mucosa. The rationale for the choice of the above mentioned assays was based, not only on their absence among those performed on omeprazole, but also on the following strategy. The first two assays were performed to verify whether omeprazole produced genotoxic effects in human lymphocytes, which are essentially deficient of the enzyme systems catalyzing the biotransformation of xenobiotics, and in primary hepatocytes, which possess a comprehensive biotransformation capability; the use of both rat and human hepatocytes was considered necessary to verify a possible interspecies difference. The micronucleus test in bone marrow and liver cells and the ACF test in colon mucosa were performed to assess in vivo the possible influence of pharmakokinetic events not reproducible in in vitro systems.

Section snippets

Chemicals

Omeprazole (pharmaceutical grade) was kindly provided by Astra Hässle AB (Mölndal, Sweeden); a commercial preparation of omeprazole (Mepral) was used in the ACF assay. Azoxymethane (AOM), collagenase type IV, lectin from Phaseulus vulgaris (PHA-M), cytochalasin B, William’s medium E (WME), Eagle’s minimal essential medium (MEM), RPMI-1640 medium, and epidermal growth factor (EGF) were purchased from Sigma Chimica (Milan, Italy); N-nitrosodimethylamine (NDMA) from Merck (Darmstadt, Germany);

Induction of DNA repair

Data of Table 1 indicate that a 20-h exposure to omeprazole concentrations ranging from 10 to 100 mg/l elicited DNA repair synthesis in primary hepatocytes from female rats. An increase in both the number of NNG and the percentage of repairing cells was detected in three independent experiments performed with cells from three different donors, and in each experiment one or two doses produced an increase of NNG exceeding our laboratory-specific threshold (5 NNG) for a positive response. However,

Discussion

This study was undertaken to evaluate the possible genotoxic activity of omeprazole in a battery of in vitro and in vivo assays to our knowledge not yet employed in the safety assessment of this worldwide used drug.

In primary rat hepatocytes both positive and negative responses were obtained: DNA repair was detected in three independent experiments but in two of them the response should be classified as equivocal due to the decrease of cytoplasmic counts and the lack of dose-dependence; an

Acknowledgements

This research was supported by a grant of MURST (Italy) targeted project ‘New Assessment Approaches in Toxicology’ (1995/1996).

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