Exposure of Infants to Isoniazid via Breast Milk After Maternal Drug Intake of Recommended Doses Is Clinically Insignificant Irrespective of Metaboliser Status. A Physiologically-Based Pharmacokinetic (PBPK) Modelling Approach to Estimate Drug Exposure of Infants via Breast-Feeding

Isoniazid is a first-line anti-tuberculosis drug recommended for treatment of drug-susceptible Mycobacterium tuberculosis infections. Breast-feeding is not contra-indicated while undergoing isoniazid therapy, even though isoniazid was found to migrate into breast milk, leading to infant drug exposure. Exposure assessment of isoniazid in infants exposed to the drug via breast milk has so far not accounted for the polymorphic expression of the isoniazid metabolising enzyme N-acetyltransferase 2. The aim of this study was to re-visit the safety assessment of maternal isoniazid therapy for infants exposed to the drug via breast milk, while accounting for fast and slow metabolisers in the adult and infant population, as well as for slower metabolism in small infants than in adults. We applied a physiologically-based pharmacokinetic (PBPK) modelling approach to estimate mother and infant external and internal drug exposure non-invasively. Validity of our PBPK models was confirmed through comparison of simulated results with experimental data. Highest recommended oral doses for mothers are daily 300 mg or 900 mg every 3 days. Simulation of maternal intake of 300 mg resulted in oral exposures of 0.58 (95%CI: 0.42–0.69) mg/day and 1.49 (1.22–1.50) mg/day for infants of fast and slow metabolising mothers, respectively. Oral exposures of infants within the first 24 h after maternal intake of 900 mg were 1.75 (1.25–2.06) mg/day and 4.46 (4.00–4.50) mg/day. Maximal drug concentrations in infant plasma ranged between 0.04 and 0.78 mg/L for the two dosing regimens. We therefore conclude that infant exposure to isoniazid via breast milk after maternal drug intake of highest recommended doses is very low. We expect that such low exposure levels most likely do not cause any clinically significant adverse effects in nursed infants

Physiologically Based Toxicokinetic Modelling as a Tool to Support Risk Assessment: Three Case Studies

In this contribution we present three case studies of physiologically based toxicokinetic (PBTK) modelling in regulatory risk assessment. (1) Age-dependent lower enzyme expression in the newborn leads to bisphenol A (BPA) blood levels which are near the levels of the tolerated daily intake (TDI) at the oral exposure as calculated by EFSA. (2) Dermal exposure of BPA by receipts, car park tickets, and so forth, contribute to the exposure towards BPA. However, at the present levels of dermal exposure there is no risk for the adult. (3) Dermal exposure towards coumarin via cosmetic products leads to external exposures of two-fold the TDI. PBTK modeling helped to identify liver peak concentration as the metric for liver toxicity. After dermal exposure of twice the TDI, the liver peak concentration was lower than that present after oral exposure with the TDI dose. In the presented cases, PBTK modeling was useful to reach scientifically sound regulatory decisions

In Vitro–In Vivo Extrapolation by Physiologically Based Kinetic Modeling: Experience With Three Case Studies and Lessons Learned

Physiologically based kinetic (PBK) modeling has been increasingly used since the beginning of the 21st century to support dose selection to be used in preclinical and clinical safety studies in the pharmaceutical sector. For chemical safety assessment, the use of PBK has also found interest, however, to a smaller extent, although an internationally agreed document was published already in 2010 (IPCS/WHO), but at that time, PBK modeling was based mostly on in vivo data as the example in the IPCS/WHO document indicates. Recently, the OECD has published a guidance document which set standards on how to characterize, validate, and report PBK models for regulatory purposes. In the past few years, we gained experience on using in vitro data for performing quantitative in vitro–in vivo extrapolation (QIVIVE), in which biokinetic data play a crucial role to obtain a realistic estimation of human exposure. In addition, pharmaco-/toxicodynamic aspects have been introduced into the approach. Here, three examples with different drugs/chemicals are described, in which different approaches have been applied. The lessons we learned from the exercise are as follows: 1) in vitro conditions should be considered and compared to the in vivo situation, particularly for protein binding; 2) in vitro inhibition of metabolizing enzymes by the formed metabolites should be taken into consideration; and 3) it is important to extrapolate from the in vitro measured intracellular concentration and not from the nominal concentration to the tissue/organ concentration to come up with an appropriate QIVIVE for the relevant adverse effects

Prediction of the dose range for adverse neurological effects of amiodarone in patients from an in vitro toxicity test by in vitro–in vivo extrapolation

Amiodarone is an antiarrhythmic agent inducing adverse effects on the nervous system, among others. We applied physiologically based pharmacokinetic (PBPK) modeling combined with benchmark dose modeling to predict, based on published in vitro data, the in vivo dose of amiodarone which may lead to adverse neurological effects in patients. We performed in vitro-in vivo extrapolation (IVIVE) from concentrations measured in the cell lysate of a rat brain 3D cell model using a validated human PBPK model. Among the observed in vitro effects, inhibition of choline acetyl transferase (ChAT) was selected as a marker for neurotoxicity. By reverse dosimetry, we transformed the in vitro concentration-effect relationship into in vivo effective human doses, using the calculated in vitro area under the curve (AUC) of amiodarone as the pharmacokinetic metric. The upper benchmark dose (BMDU) was calculated and compared with clinical doses eliciting neurological adverse effects in patients. The AUCs in the in vitro brain cell culture after 14-day repeated dosing of nominal concentration equal to 1.25 and 2.5 mu M amiodarone were 1.00 and 1.99 mu g*h/mL, respectively. The BMDU was 385.4 mg for intravenous converted to 593 mg for oral application using the bioavailability factor of 0.65 as reported in the literature. The predicted dose compares well with neurotoxic doses in patients supporting the hypothesis that impaired ChAT activity may be related to the molecular/cellular mechanisms of amiodarone neurotoxicity. Our study shows that predicting effects from in vitro data together with IVIVE can be used at the initial stage for the evaluation of potential adverse drug reactions and safety assessment in humans

Testing the thresholds of toxicological concern values using a new database for food-related substances.

Abstract The Threshold of Toxicological Concern (TTC) concept integrates data on exposure, chemical structure, toxicity and metabolism to identify a safe exposure threshold value for chemicals with insufficient toxicity data for risk assessment. The TTC values were originally derived from a non-cancer dataset of 613 compounds with a potentially small domain of applicability. There is interest to test whether the TTC values are applicable to a broader range of substances, particularly relevant to food safety using EFSA's new OpenFoodTox database. After exclusion of genotoxic compounds, organophosphates or carbamates or those belonging to the TTC exclusion categories, the remaining 329 substances in the EFSA OpenFoodTox database were categorized under the Cramer decision tree, into low (Class I), moderate (II), or high (III) toxicity profile. For Cramer Classes I and III the threshold values were 1000 μg/person per day (90% confidence interval: 187–2190) and 87 μg/person per day (90% confidence interval: 60–153), respectively, compared to the corresponding original threshold values of 1800 and 90 μg/person per day. This confirms the applicability of the TTC values to substances relevant to food safety. Cramer Class II was excluded from our analysis because of containing too few compounds. Comparison with the Globally Harmonized System of classification confirmed that the Cramer classification scheme in the TTC approach is conservative for substances relevant to food safety

Safety evaluation of steviol glycoside preparations, including rebaudioside AM, obtained by enzymatic bioconversion of highly purified stevioside and/or rebaudioside A stevia leaf extracts

[EN] The EFSA Panel on Food Additives and Flavourings (FAF) provides a scientific opinion on the safety of steviol glycoside preparations, including rebaudioside AM, obtained by enzymatic bioconversion of highly purified stevioside and/or rebaudioside A stevia leaf extracts. These steviol glycoside preparations are produced via enzymatic bioconversion of highly purified stevioside and/or rebaudioside A extracts obtained from stevia plant using two UDP-glucosyltransferases and one sucrose synthase enzymes produced by the genetically modified strains of E. coli K-12 that facilitate the transfer of glucose to purified stevia leaf extracts via glycosidic bonds. The Panel considered that the parental strain is a derivative of E. coli K-12 which is well characterised and its safety has been documented; therefore, it is considered to be safe for production purposes. The Panel concluded that there is no safety concern for steviol glycoside preparations, including rebaudioside AM, obtained by enzymatic bioconversion of highly purified stevioside and/or rebaudioside A stevia leaf extracts using UDP-glucosyltransferases and sucrose synthase enzymes produced by the genetically modified strains of E. coli K-12, to be used as a food additive. The Panel recommends the European Commission to consider the proposal of establishing separate specifications for steviol glycoside preparations, including rebaudioside AM, obtained by enzymatic bioconversion of highly purified stevioside and/or rebaudioside A stevia leaf extracts in Commission Regulation (EU) No 231/2012. (C) 2021 European Food Safety Authority. EFSA Journal published by John Wiley and Sons Ltd on behalf of European Food Safety Authority.Younes, M.; Aquilina, G.; Castle, L.; Engel, K.; Fowler, P.; Frutos Fernandez, MJ.; Furst, P.... (2021). Safety evaluation of steviol glycoside preparations, including rebaudioside AM, obtained by enzymatic bioconversion of highly purified stevioside and/or rebaudioside A stevia leaf extracts. EFSA Journal. 19(8):1-22. https://doi.org/10.2903/j.efsa.2021.669112219

Safety evaluation of crosslinked polyacrylic acid polymers (carbomer) as a new food additive

[EN] The EFSA Panel on Food Additives and Flavourings (FAF) provides a scientific opinion on the safety of crosslinked polyacrylic acid polymers (carbomer) proposed for use as food additive in solid and liquid food supplements. Carbomer is formed from the monomer, acrylic acid, which is polymerised and crosslinked with allyl pentaerythritol (APE). The polymers are synthesised in ethyl acetate using as free-radical polymerisation initiator. In vivo data showed no evidence for systemic availability or biotransformation of carbomer. Carbomer does not raise a concern regarding genotoxicity. Considering the available data set, the Panel derived an acceptable daily intake (ADI) of 190 mg/kg body weight (bw) per day based on a no observed adverse effect level (NOAEL) of 1,500 mg/kg bw per day from a sub-chronic 13-week study in rat, applying a compound specific uncertainty factor (UF) of 8. At the proposed maximum use levels, the exposure estimates ranged at the mean from 1.1 to 90.2 mg/kg bw per day and at the p95 from 12.5 to 237.4 mg/kg bw per day. At the proposed typical use level, the exposure estimates ranged at the mean from 0.7 to 60.2 mg/kg bw per day and at the p95 from 10.3 to 159.5 mg/kg bw per day. The Panel noted that the maximum proposed use levels would result in exposure estimates close to or above the ADI. The Panel also noted that level of exposure to carbomer from its proposed use is likely to be an overestimation. Taking a conservative approach, the Panel considered that exposure to carbomer would not give rise to a safety concern if the proposed maximum use level for solid food supplements is lowered to the typical use level reported by the applicant. (C) 2021 European Food Safety Authority. EFSA Journal published by John Wiley and Sons Ltd on behalf of European Food Safety Authority.Younes, M.; Aquilina, G.; Engel, K.; Fowler, P.; Frutos Fernandez, MJ.; Furst, P.; Gürtler, R.... (2021). Safety evaluation of crosslinked polyacrylic acid polymers (carbomer) as a new food additive. EFSA Journal. 19(8):1-26. https://doi.org/10.2903/j.efsa.2021.669312619

Safety evaluation of long-chain glycolipids from Dacryopinax spathularia

[EN] The EFSA Panel on Food Additives and Flavourings (FAF) provides a scientific opinion on the safety of long-chain glycolipids from Dacryopinax spathularia (also called AM-1) as a food additive. AM-1 is a purified mixture of long-chain glycolipid congeners obtained by fermentation of the edible non-genetically modified fungus Dacryopinax spathularia. AM-1 glycolipids have very low oral bioavailability and overall available toxicology data do not demonstrate any adverse effects of the proposed food additive. Considering the available data set the Panel established an ADI of 10 mg/kg bw per day based on a range of NOAELs between 1,000 and 1,423 mg/kg bw per day (the highest doses tested), from the reproductive and a prenatal developmental toxicity studies in rats and 90-day studies in rat and dog. At the proposed maximum use levels, the exposure estimates ranged at the mean from 0.01 to 1.07 mg/kg bw per day and at the p95 from 0 to 3.1 mg/kg mg/kg bw per day. At the proposed typical use levels, the exposure estimates ranged at the mean from < 0.01 mg/kg bw per day to 0.23 mg/kg bw per day and at the p95 from 0 to 0.64 mg/kg bw per day. The Panel noted that the highest estimate of exposure of 3.1 mg/kg bw per day (in toddlers) is within the established ADI of 10 mg/kg bw per day and concluded that the exposure to long-chain glycolipids from Dacryopinax spathularia does not raise a safety concern at the uses and use levels proposed by the applicant.Younes, M.; Aquilina, G.; Engel, K.; Fowler, P.; Frutos Fernandez, MJ.; Furst, P.; Gurtler, R.... (2021). Safety evaluation of long-chain glycolipids from Dacryopinax spathularia. EFSA Journal. 19(6):1-28. https://doi.org/10.2903/j.efsa.2021.660912819

Safety evaluation of buffered vinegar as a food additive

[EN] The EFSA Panel on Food Additives and Flavourings (FAF) provides a scientific opinion on the safety of buffered vinegar as a new food additive. Buffered vinegar is a liquid or dried product prepared by adding sodium/potassium hydroxides (E 524 to E 525) and sodium/potassium carbonates (E 500 to E 501) to vinegar, compliant with European Standard EN 13188:2000 and exclusively obtained from an agricultural source origin (except wood/cellulose). The primary constituents of buffered vinegar are acetic acid and its salts. No biological or toxicological data obtained with the proposed food additive were submitted by the applicant as part of the dossier as, following oral ingestion, buffered vinegar dissociates into the acetic anion and acetate a natural constituent of the diet, and of the human body for which extensive data on their biological effects exist and for which EFSA in 2013 has previously concluded that the establishment of an acceptable daily intake (ADI) is not considered necessary. At the proposed maximum/typical use levels, the mean exposure to buffered vinegar from its use as a food additive expressed as acetic acid equivalents ranged from 8.9 mg/kg body weight (bw) per day in infants to 280.3 mg/kg bw per day in children. The 95th percentile of exposure to buffered vinegar ranged from 27.9 mg/kg bw per day in infants to 1,078 mg/kg bw per day in toddlers. The Panel concluded that there is no safety concern for the use of buffered vinegar as a food additive at the proposed maximum/typical use levels. The Panel could not conclude on the safety for the proposed uses at quantum satis as Group I food additive since the resulting exposure could not be estimated.The Panel wishes to thank the following for the support provided to this scientific output: Alkiviadis Stagkos-Georgiadis.Younes, M.; Aquilina, G.; Degen, G.; Engel, K.; Fowler, P.; Frutos Fernandez, MJ.; Fürst, P.... (2022). Safety evaluation of buffered vinegar as a food additive. EFSA Journal. 20(7):1-21. https://doi.org/10.2903/j.efsa.2022.735112120

Safety of the proposed amendment of the specifications for enzymatically produced steviol glycosides (E 960c): Rebaudioside D produced via enzymatic bioconversion of purified stevia leaf extract

[EN] The EFSA Panel on Food Additives and Flavourings (FAF Panel) provides a scientific opinion on the safety of a proposed amendment of the specifications of enzymatically produced steviol glycosides (E 960c) with respect to the inclusion of rebaudioside D produced via enzyme-catalysed bioconversion of purified stevia leaf extract. Rebaudioside D (95% on dry basis) is produced via enzymatic bioconversion of purified stevia leaf extract using uridine diphosphate (UDP)-glucosyltransferase (UGT) and sucrose synthase enzymes produced by the genetically modified yeast K. phaffii UGT-A, that facilitates the transfer of glucose to purified stevia leaf extract via glycosidic bonds. The same enzymes from K. phaffii UGT-A may be used in the manufacturing process of the food additive, rebaudioside M produced via enzyme modification of steviol glycosides from stevia (E 960c(i)). The Panel considered that separate specifications would be needed for this food additive produced via the manufacturing process described in the current application, aligned with those already established for E 960c(i). The Panel concluded that there is no toxicological concern for Rebaudioside D produced via enzymatic bioconversion of purified stevia leaf extract using UDP-glucosyltransferase and sucrose synthase produced by a genetically modified strain of the yeast K. phaffii. However, based on the available data, the Panel could not exclude the possibility that some residual amount of DNA coding for the kanamycin resistance gene could remain in the final product. Should this gene propagate in microbiota due to the presence of recombinant DNA in the final product, this would be of concern. Therefore, the Panel concluded that the safety of Rebaudioside D produced via this enzymatic bioconversion was not sufficiently demonstrated with the available data given that the absence of recombinant DNA was not shown.Younes, M.; Aquilina, G.; Engel, K.; Fowler, P.; Frutos Fernandez, MJ.; Fürst, P.; Gürtler, R.... (2022). Safety of the proposed amendment of the specifications for enzymatically produced steviol glycosides (E 960c): Rebaudioside D produced via enzymatic bioconversion of purified stevia leaf extract. EFSA Journal. 20(5):1-23. https://doi.org/10.2903/j.efsa.2022.729112320