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Mode of Action (MOA) Evaluation and Derivation of a Cancer and Non-Cancer Reference Value for 4-Vinylcyclohexene

Authors: C. Bevan, M. Gargas, C. Kirman, and J. Vergnes

Session: Risk Assessment Applications
Date and Time: March 17, 2009 9 AM to 12: 30 PM
Location: Exhibit Hall

Abstract No. 839

4-Vinylcyclohexene (VCH) induces follicular loss and tumors in the ovaries of mice, but not rats. Alternative MOAs for the mouse ovarian tumors were evaluated using the modified Hill criteria for causality in the IPCS Human Relevance Framework. There is a high degree of confidence that VCH acts through a non-genotoxic, non-linear (threshold) MOA in producing mouse ovarian tumors. The critical event is the destruction of ovarian primordial and primary follicles by VCH diepoxide through an apoptotic mechanism. Complete oocyte loss results in ovarian failure, which increases plasma FSH levels from the loss of negative feedback from 17β-estradiol and inhibin on the hypothalamus/ pituitary. Ovarian tumor development occurs from the elevated plasma FSH levels, perhaps through alteration in signaling pathways affecting cell growth. The MOA is relevant to humans, but due to metabolic differences between mice and humans, humans are expected to be less susceptible than mice to the ovarian effects from comparable VCH doses. Although there are limited in vitro data indicating genotoxic potential from the VCH epoxide metabolites, particularly VCH diepoxide, the weight of evidence is not compelling that genotoxicity is important in the MOA for mouse ovarian tumors. A 13-week mouse inhalation study was considered to be the key study, and a critical adverse effect (ovarian toxicity) with a clear NOAEL of 250 ppm (47 ppm when adjusted for continuous exposure) was identified. A benchmark dose evaluation also resulted in a BMDL value of 49 ppm. A total uncertainty factor of 100 was used: 10 (intraspecies variability), 10 (subchronic to chronic), and 1 (interspecies) because of mouse/human differences in VCH diepoxide production and similarity between VCH and 1,3-butadiene metabolism. A non-cancer reference value of 0.47 ppm (470 ppb) was derived. Based on the proposed MOA, this value is expected to be adequately protective against ovarian tumors.

Liver Effects in Mice Given Trichloroacetic acid (TCA) in a 14-Day Drinking Water Study are PPARαa-dependent

Authors: P. Dugard, and C. Bevan

Session: Hepatotoxicity: In Vivo Studies
Date and Time: March 17, 2009 1:00 PM to 4:30 PM
Location: Exhibit Hall

Abstract No. 1136

Exposure of mice to perchloroethylene (perc) is known to increase peroxisomes and cell proliferation in the liver, hepatocellular hypertrophy, and liver tumors. TCA is the major metabolite of perc and is known to induce the same responses, including tumors, in the livers of mice when given in drinking water. This study was conducted to determine whether the liver effects seen in mice exposed to TCA in drinking water are PPARαa-dependent. Male and female B6C3F1 mice, male 129/sv wild-type (WT) and male PPARαa-null mice were given 0, 1 or 2.5 g/L TCA in drinking water for 5 (male B6C3F1 mice only), or 14 days. Liver tissue was examined by light and electron microscopy, cell proliferation was quantified by BrDU labeling, and apoptosis was measured by using Tdt-mediated dUTP nick end labelling (TUNEL) staining. Effects in B6C3F1 mice were: increased relative liver weights (1 and 2.5 g/L); increased palmitoyl CoA oxidase activity, with males > females (1 and 2.5 g/L); increased number and volume density of peroxisomes (1 and 2.5 g/L); non-statistically significant increase in cell proliferation in males at 5 days (316% and 267%, respectively); dose-dependent decrease in apoptosis in males at 5 days (12% and 31%, respectively); and eosinophilia and/or hepatocyte hypertrophy (1 and 2.5 g/L). Similar effects were seen in the WT male mice at 14 days, except that there was no change in cell proliferation. Palmitoyl CoA oxidase activity was increased to a greater extent in male WT mice compared to B6C3F1 mice. No liver effects were seen in the PPARαa-null mice except for evidence of fatty vacuolation in all PPARαa-null mice, which appeared more widespread in the treated versus non-treated mice. Thus, peroxisome proliferation occurring in the livers of TCA-treated mice is dependent on PPARαa. The results also suggest that perc-induced liver effects in mice are mediated by PPARαa.

Contribution of Trichloroacetic Acid to Liver Tumors Observed in Perchloroethylene (Perc)-exposed Mice.

Authors: L.M. Sweeney, C.R. Kirman, M.L. Gargas, and P.H. Dugard

Session Title: Risk Assessment Research
Date & Time: March 18, 2009 from 9:00 AM to 12:30 PM
Location: Exhibit Hall

Abstract Number: 1491

Perchloroethylene (Perc) is a solvent used in dry cleaning operations and industrial applications such as metal degreasing. Perc has been found to produce increases in hepatocellular carcinomas and/or adenomas in male and female mice in chronic inhalation bioassays. Perc is metabolized primarily to trichloroacetic acid (TCA), which is also a mouse hepatocarcinogen. The fractional conversion of perchloroethylene to TCA by mice was determined from physiologically based pharmacokinetic (PBPK) modeling of TCA in mouse blood at the conclusion of inhalation exposure of male and female B6C3F1 mice to 10, 50, 100, or 200 ppm perc for 6 hrs/day for 5 days. The dose-dependent bioavailability of TCA in male and female B6C3F1 mice exposed to TCA in drinking water was estimated by optimizing the fit of time course blood, plasma, and liver TCA concentrations for a TCA doses ranging from 12 to 800 mg/kg/day to predictions of a previously published TCA PBPK model. Using the PBPK models, the area under the liver TCA concentration vs. time curve (liver TCA AUC) was calculated for TCA and perc bioassays. Benchmark dose analyses were conducted to determine the dose-response relationship between liver TCA AUC and the additional risk of hepatocellular adenomas or carcinomas (combined) in mice ingesting TCA. Using the dose-response relationships derived for the TCA-exposed mice, the contribution of TCA produced by metabolism to the additional risk of liver adenomas and carcinomas in mice exposed to perchloroethylene by inhalation was computed. The analysis indicated that the levels of TCA observed in perchloroethylene-exposed mice are sufficient to explain the incidence of liver adenomas and carcinomas.

Intravenous- and Inhalation-route Pharmacokinetics of Propanol and Its Metabolite, Propionic Acid.

Authors: J. Soelberg, T.S. Poet, A.L. Busby, L.M. Sweeney, and W. Faber.

Session: Risk Assessment Research
Date and Time: March 18, 2009 9 AM to 12:30 PM
Location: Risk Assessment Research, Exhibit Hall

Abstract No. 1483

n-Propanol is widely used in many industrial applications, potentially leading to inhalation exposures in the workplace . Extrapolation between observed effects in animals and potential human effects requires detailed route-specific pharmacokinetic analyses. Inhalation and intravenous (iv) exposure studies were conducted in rats to determine the blood pharmacokinetics of propanol and its major metabolite, propionic acid. Inhalation-route studies included constant exposures to 500 or 3500 ppm 13C-propanol with concurrent measurement of respiratory rates using plethysmography and measurement of 13C-propanol and 13C-propionic acid in blood. Radiolabeled material was used to isolate the metabolite from endogenous propionic acid. Respiratory rates were steady for the 2 hr inhalation exposures. The average minute volume for 4 rats exposed to 3500 ppm propanol over 2 hr was 116 ± 22 ml/hr. Blood propanol and propionic acid peaked within the 2 hr exposure and dropped immediately post exposure. For 3500 ppm exposures, peak propanol was 360 ± 67 nmol/ml, several fold higher than observed after a 25 mg/kg iv dose. Propionic acid however was proportionally less than after the iv dose, most likely indicating saturable metabolism of propanol. The iv exposures included a 30 minute infusion to achieve steady state with analysis for 13C-propanol and 13C-propionic acid in blood and urine. Rats were also dosed iv with 13C-propionic acid to determine the acid kinetics independent of propanol metabolism. The Cmax and AUC of 13C- propionic acid were ~6x higher following a propionic acid dose than following an equimolar dose of propanol. The rate of elimination of propionic acid in blood post iv dosing was the same following a propanol dose as for a propionic acid dose. Propanol in urine was close to detection limits. These studies show that propanol is readily absorbed via inhalation and metabolized to propionic acid. (ACC Oxo Panel Sponsored)