Development of a Novel Method for Deriving Thresholds of Toxicological Concern (TTCs) for Vaccine Constituents

Abstract Safety assessment relating to the presence of impurities, residual materials and contaminants in vaccines is a focus area of research at the United States Food and Drug Administration (FDA). Sponsors who submit Investigational New Drug (IND) applications for new vaccine products must report the results of safety assessments to the Division of Vaccines and Related Products Applications (DVRPA). Scientifically defining thresholds of toxicological concern (TTCs) as they apply to vaccine constituents will provide a useful aid to the sponsors and public regarding safety assessments of compounds for which there is little or no toxicity data. TTCs are mathematically modeled and extrapolated levels, below which adverse human health effects are not expected to occur (Kroes, 2004). In this project, we accessed DVRPA\u27s submission databases and open source data to yield an initial chemical test set. Using INCHEM, RepDose, RTECS and TOXNET, we gathered LD50 and TDLo data. Using a structure-based decision tree, provided in the ToxTree software package, (3) different algorithms (The Cramer extended, the In vivo rodent micronucleus assay, and the Benigni-Bossa rule base for carcinogenicity by ISS) were applied to assign the initial test set (n= 197) of chemicals into structural families based on structural alerts (SAs). This resulted in six (6) potential methods for elucidating TTCs: In vivo rodent micronucleus assay/ LD50, Benigni-Bossa/ LD50, Cramer extended/ LD50, In vivo rodent micronucleus assay/ TDLo, Benigni-Bossa/ TDLo, and the Cramer extended/ TDLo. After each algorithm designated two structural families each, the distribution of TDLo\u27s and LD50\u27s for each structural family was subjected to a preliminary data analysis using JMP statistical software version 9. Based on an analysis of quantiles, skew, and kurtosis, it was concluded that the TDLo dataset was of poor quality and was dropped from further analysis, and that the In vivo rodent micronucleus assay algorithm failed to partition the initial test set in a meaningful way, so it too was culled from further consideration. This resulted in (2) remaining TTC methods for further consideration: Benigni-Bossa/ LD50 and the Cramer extended/ LD50. The remaining methods were subjected to internal validation based on Gene-Tox, CCRIS, CPDB, IARC, and EPA classaifications for genotoxic mutagenicity and carcinogenicity. Validation parameters were calculated for both methods and it was determined that the Benigni-Bossa/ LD50 method outperformed the Cramer extended/ LD50 method in terms of specificity (87.2 vs. 48.1%#37;), accuracy (65.2 vs. 52.94%#37;), positive predictivity (66.6 vs. 50%#37;), negative predictivity (64.8 vs. 56.5%#37;), ROC+ (2 vs. 1) and ROC- (1.84 vs. 1.3). These results indicated that the Benigni-Bossa/ LD50 was the most appropriate for calculating TTCs for vaccine constituents. For each class, the lower 2.5th percentile LD50 was extrapolated to a TTC value using safety estimates derived using uncertainty factors (UF) and adjusting for adult human weight. Final TTCs were designated as 18.06 μg/ person and 20.616 μg/ person for the Benigni-Bossa positive and negative structural families