Tumorigenicity and genotoxicity of 2,7-dinitrofluorene after systemic administration at a low dose level to female rats Free

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2,7-Dinitrofluorene (2,7-diNF) and 9-oxo-2,7-diNF are environmental pollutants formed chiefly from incomplete combustion of fossil fuels. We examined the tumorigenicity in female Sprague-Dawley rats of 2,7-diNF and 9-oxo-2,7-diNF administered by intraperitoneal (i.p.) and oral routes at 10 μmol/kg body weight (b.wt.) or the vehicle (12% DMSO in olive oil) at 2.5 ml/kg b.wt., three times per week for 4 weeks (wk), starting at 4 wk of age (16 or 17 rats/group). The predominant site of tumor induction was the mammary gland. In i.p.-treated rats, Kaplan-Meier estimates of median latency for the combined malignant and benign mammary tumors showed a decrease in 2,7-diNF- and 9-oxo-2,7-diNF-treated groups (42 wk, P=0.003 and 64 wk, P=0.007, respectively) relative to the vehicle-treated (80 wk) group, whereas after oral dosing, there were no significant differences in the median tumor latency between 2,7-diNF- or 9-oxo-2,7-diNF- and the vehicle-treated rats (70 or 82 vs. 77 wk, respectively). At the end point of 90 wk, the malignant mammary tumor incidence in 2,7-diNF-, 9-oxo-2,7-diNF-, and vehicle-treated rats was 44% (P=0.02, compared to vehicle-treated), 25%, and 6%, respectively, after i.p., and 29%, 18%, and 12%, respectively, after oral dosing. Liver and mammary gland DNA was isolated 24 h after the 12^th i.p. or oral dose to 4 rats/group, and analyzed for the presence of N-(deoxyguanosin-8-yl)-2-amino-7-nitrofluorene (dG-2,7-diNF), a deoxyadenosine adduct (dA-2,7-diNF) derived from 2,7-diNF, and a deoxyguanosine adduct (dG-9-oxo-2,7-diNF) derived from 9-oxo-2,7-diNF by HPLC combined with electrospray tandem mass spectrometry. Both dG-2,7-diNF and dA-2,7-diNF were detected in DNA (3.9-59.5 adducts/10⁸ nucleotides) from tissues of 2,7-diNF-treated rats. Only very low levels of dG-9-oxo-2,7-diNF (≤1.2 adducts/10⁸ nucleotides) were detected in the livers and mammary gland of 9-oxo-2,7-diNF-treated rats. None of the adducts was found in DNA from the vehicle-treated rats. The levels of dA-2,7-diNF in individual tissues were unaffected by the mode of treatment, but after i.p. administration, its level was higher (P<0.01) in the mammary gland than liver (13.6 vs. 7.8 adducts/10⁸ nucleotides). In the liver, the level of dG-2,7-diNF was unaffected by the mode of treatment, but in the mammary gland, the level dG-2,7-diNF was higher (P<0.05) after i.p. than oral dosing and also higher (P<0.05) than in the liver (36 vs. 8.6 and vs. 9.1 adducts/10⁸ nucleotides, respectively). The genotoxic effects of 2,7-diNF in vivo are thus consistent with its tumorigenicity for the mammary gland. The data confirm carcinogenic potency of low doses of 2,7-diNF reported by us previously after its intra-mammary application (Carcinogenesis 20: 2017, 1999), and signify its potential relevance for environmental breast cancer.