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Influence of Study Design on Developmental and Reproductive Toxicology Study Outcomes

Paul M. D. Foster
Toxicologic Pathology (2017) DOI: https://doi.org/10.1177/0192623316671608 PMID: 27708197

Publication

Abstract

Regulatory studies of developmental and reproductive toxicity (DART) studies have remained largely unchanged for decades, with exposures occurring at various phases of the reproductive cycle and toxicity evaluations at different ages/times depending on the study purpose. The National Toxicology Program has conducted studies examining the power to detect adverse effects where there is a prenatal exposure, but evaluations occur postnatally. In these studies, examination is required of only 1 male and female pup from each litter beyond weaning. This provides poor resolving power to detect rare events (e.g., reproductive tract malformations). If an adverse effect is detected, there is little confidence in the shape of the dose-response curve (and the Benchmark Dose or No Observed Adverse Effect Level [NOAEL]). We have developed a new protocol to evaluate DART, the modified one generation study, with exposure commencing with pregnant animals and retention of 4 males and females from each litter beyond weaning to improve statistical power. These animals can be allocated to specific cohorts that examine subchronic toxicity, teratology, littering, and neurobehavioral toxicity in the same study. This approach also results in a reduction in animal numbers used, compared with individual stand-alone studies, and offers increased numbers of end points evaluated compared with recent Organization for Economic Cooperation and Development proposals.

Figures

Figure 1. Diagrammatic representation of the mammalian reproductive cycle.

Source: Adapted from Foster and Gray (2013).

Figure 2. Diagrammatic representation of a typical multigeneration study.

Diagrammatic representation of a typical multigeneration study with reference to the mammalian reproductive cycle. Dosing is continuous throughout the cycle with assessments made at multiple life stages. Source: Adapted from Foster and Gray (2013).

Figure 3. Diagrammatic representation of the current EPA test guideline.

Diagrammatic representation of the current Environmental Protection Agency (EPA) test guideline for fertility and reproductive effects (Office of Prevention, Pesticides and Toxic Substances [OPPTS] 870.3800). Source: Adapted from Foster and Gray (2013). Q = quarantine; PBE = prebreed exposure; ECE = estrous cycle evaluation; M = mating; G = gestation; L = lactation; W = weaning; N = necropsy; C = cull; VO = vaginal opening; PPS = preputial separation; PND = postnatal day; AGD = anogenital distance.

Figure 4. Power curves for the detection of rare adverse reproductive malformations.

Power curves for the detection of rare adverse reproductive malformations from a developmental and reproductive toxicity study. The study is assumed to have 20 litters/group and a 0% background incidence. The vertical dotted line indicates a 10% incidence of an adverse outcome, which would be detected 4.7% with 1 pup/litter 66.4% with 3 pups per litter and 86.5% with 4 pups/litter. Source: Adapted from Blystone et al. (2010).

Figure 5. Diagrammatic representation of the NTP modified one-generation reproduction study.

Diagrammatic representation of the National Toxicology Program modified one-generation reproduction study. Only 10 pups per sex (on reaching adulthood) are required for the subchronic cohort, and thus sufficient numbers of animals would be available for evaluations of other developmental toxicity that may include effects on the developing immune or nervous systems (see Figure 6). G = gestation; L = lactation; PND = postnatal day; GD = gestation day; M = mating; VO = vaginal opening; PPS = belanopreputial separation; EC = estrous cyclicity evaluation.

Figure 6. Diagrammatic representation: one-generation reproduction study with a developmental neurotoxicity cohort.

Diagrammatic representation of the National Toxicology Program modified one-generation reproduction study with a developmental neurotoxicity cohort.

Figure 7. Diagrammatic representation of the Organization for Economic Cooperation.

Diagrammatic representation of the Organization for Economic Cooperation and Development 443 extended one-generation reproduction study (2012).
P = parental generation; M = males; F = females; F1 = first filial generation.

Tables

Table 1. Estimation of Animals Employed on Standard, Stand-alone, NTP Toxicity Studies.

Estimation of Animals Employed on Standard, Stand-alone, National Toxicology Program Toxicity Studies in Comparison to the Modified One-Generation (MOG) Study.