Genotoxicity and cytotoxicity of antineoplastic drugs at environmentally relevant concentrations after long-term exposure

Abstract

Introduction: 5-fluorouracil (5-FU) and methotrexate (MTX) are the antineoplastic drugs most commonly used worldwide. Considered cytotoxic, these pharmaceuticals exhibit low specificity, causing damage not only to cancer cells but also to healthy cells in organisms. After being consumed and metabolized, these drugs are excreted through urine and feces, followed by wastewater treatment. However, conventional treatments do not have the capacity to completely remove these substances, risking their introduction into freshwater systems. This could pose a risk to human health even at low concentrations.

Aims: Thus, the present study aimed to investigate the genotoxicity, cytotoxicity, and mutagenicity of 5-FU and MTX at environmentally relevant concentrations after a long-term exposure, using adult male rats as an experimental model.

Methods: Male Wistar rats (70 days old) were distributed into 4 groups (n = 10/group): control, received only vehicle; MTX, received methotrexate at 10ngL^-1; 5-FU received 5-fluorouracil at 10ngL^-1; and MTX + 5-FU, received a combination of MTX and 5-FU at 10ngL^-1 each. The period of exposure was from postnatal day (PND) 70 to PND 160, through drinking water. After that, the animals were euthanized and the samples (liver, testis, femoral bone marrow, and peripheral blood) were obtained.

Results: Increased DNA fragmentation was observed in the peripheral blood, liver, and testis, altering the parameters of the tail moment and tail intensity in the Comet assay. Besides, the change in the ratio between PCE and NCE indicates bone marrow suppression.

Conclusion: These findings warn the adverse effects for the general population worldwide chronically exposed to these drugs at trace concentration unintentionally.

Keywords: 5-fluorouracil; DNA damage; Emerging contaminants; Genetic toxicology; Methotrexate; Rats.

Fig. 1

Comet assay on whole blood samples from experimental animals at PND 160. A: Tail moment (a.u). B: Tail intensity (%). Values expressed as mean ± SD, Kruskal-Wallis with Dunn’s a posteriori test. For all groups with the same letter superscript, the difference between the means is not statistically significant. Tail moment: MTX vs control, p = 0.0143. Tail intensity: MTX vs control, p = 0.0099.

Fig. 2

Comet assay in liver of experimental animals at DPN 160. A: Tail moment (a.u). B: Tail intensity (%). Values expressed as mean ± SD, Kruskal-Wallis with Dunn’s a posteriori test. For all groups with the same letter superscript, the difference between the means is not statistically significant. Tail moment: 5-FU vs control (p = 0.0052), MTX (p = 0.0043), 5-FU + MTX (<0.0001), 5-FU + MTX vs control (p = 0.0101), MTX (P = <0.0001).

Fig. 3

Comet assay in testis of experimental animals at PND 160. A: Tail moment (a.u). B: Tail intensity (%). Values expressed as mean ± SD, Kruskal-Wallis with Dunn’s a posteriori test. For all groups with the same letter superscript, the difference between the means is not statistically significant. Tail moment: 5-FU vs control (P < 0.0001), MTX vs control (p = 0.035), 5-FU + MTX vs control (p = 0.0027). Tail intensity: 5-FU vs control (P < 0.0001), MTX (p = 0.0289), and 5-FU + MTX vs control (p = 0.0053).