RESUMO
The aim of embryo-fetal developmental toxicity assessments for pharmaceuticals is to inform potential risk of adverse pregnancy outcome, which has traditionally relied on studies in pregnant animals. Recent updates to international safety guidelines (ICH S5R3) have incorporated information on how to use weight of evidence and alternative assays to reduce animal use while still informing risk of fetal harm. Uptake of these alternative approaches has been slow due to limitations in understanding how alternative assays translate to in vivo effects and then relevance to human exposure. To understand the predictivity of new approach methodologies for developmental toxicity (DevTox NAMs), we used two pharmaceutical examples (glasdegib and lorlatinib) to illustrate the value of DevTox NAMs to complement weight of evidence (WoE) assessments while considering the relationship of concentration-effect levels in NAMs to in vivo studies. The in vitro results generated in a battery of assays (mEST, rWEC, zebrafish, and human based stem cells) confirmed the WoE based on literature and further confirmed by preliminary embryo-fetal development data. The data generated for these two compounds supports integrating DevTox NAMs into the developmental toxicity assessment for advanced cancer indications.
RESUMO
Targeted protein degradation is the selective removal of a protein of interest through hijacking intracellular protein cleanup machinery. This rapidly growing field currently relies heavily on the use of the E3 ligase cereblon (CRBN) to target proteins for degradation, including the immunomodulatory drugs (IMiDs) thalidomide, lenalidomide, and pomalidomide which work through a molecular glue mechanism of action with CRBN. While CRBN recruitment can result in degradation of a specific protein of interest (e.g., efficacy), degradation of other proteins (called CRBN neosubstrates) also occurs. Degradation of one or more of these CRBN neosubstrates is believed to play an important role in thalidomide-related developmental toxicity observed in rabbits and primates. We identified a set of 25 proteins of interest associated with CRBN-related protein homeostasis and/or embryo/fetal development. We developed a targeted assay for these proteins combining peptide immunoaffinity enrichment and high-resolution mass spectrometry and successfully applied this assay to rabbit embryo samples from pregnant rabbits dosed with three IMiDs. We confirmed previously reported in vivo decreases in neosubstrates like SALL4, as well as provided evidence of neosubstrate changes for proteins only examined in vitro previously. While there were many proteins that were similarly decreased by all three IMiDs, no compound had the exact same neosubstrate degradation profile as another. We compared our data to previous literature reports of IMiD-induced degradation and known developmental biology associations. Based on our observations, we recommend monitoring at least a major subset of these neosubstrates in a developmental test system to improve CRBN-binding compound-specific risk assessment. A strength of our assay is that it is configurable, and the target list can be readily adapted to focus on only a subset of proteins of interest or expanded to incorporate new findings as additional information about CRBN biology is discovered.
Assuntos
Proteólise , Proteômica , Talidomida , Ubiquitina-Proteína Ligases , Animais , Coelhos , Proteômica/métodos , Ubiquitina-Proteína Ligases/metabolismo , Talidomida/análogos & derivados , Talidomida/farmacologia , Proteólise/efeitos dos fármacos , Feminino , Embrião de Mamíferos/metabolismo , Embrião de Mamíferos/efeitos dos fármacos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Lenalidomida/farmacologia , GravidezRESUMO
BACKGROUND: Abrocitinib is a Janus kinase (JAK) 1 selective inhibitor approved for the treatment of atopic dermatitis. Female reproductive tissues were unaffected in general toxicity studies, but an initial female rat fertility study resulted in adverse effects at all doses evaluated. A second rat fertility study was conducted to evaluate lower doses and potential for recovery. METHODS: This second study had 4 groups of 20 females each administered abrocitinib (0, 3, 10, or 70 mg/kg/day) 2 weeks prior to cohabitation through gestation day (GD) 7. In addition, 2 groups of 20 rats (0 or 70 mg/kg/day) were dosed for 3 weeks followed by a 4-week recovery period before mating. All mated females were evaluated on GD 14. RESULTS: No effects were observed at ≤10 mg/kg/day. At 70 mg/kg/day (29x human exposure), decreased pregnancy rate, implantation sites, and viable embryos were observed. All these effects reversed 4 weeks after the last dose. CONCLUSIONS: Based on these data and literature on the potential role of JAK signaling in implantation, we hypothesize that these effects may be related to JAK1 inhibition and, generally, that peri-implantation effects such as these, in the absence of cycling or microscopic changes in nonpregnant female reproductive tissues, are anticipated to be reversible.
Assuntos
Fertilidade , Janus Quinase 1 , Pirimidinas , Sulfonamidas , Feminino , Animais , Gravidez , Ratos , Fertilidade/efeitos dos fármacos , Janus Quinase 1/antagonistas & inibidores , Janus Quinase 1/metabolismo , Pirimidinas/farmacologia , Sulfonamidas/farmacologia , Ratos Sprague-Dawley , Implantação do Embrião/efeitos dos fármacos , Inibidores de Janus Quinases/farmacologia , Taxa de GravidezRESUMO
Traditionally, understanding potential developmental toxicity from pharmaceutical exposures has been based on the results of ICH guideline studies in two species. However, support is growing for the use of weight of evidence approaches when communicating the risk of developmental toxicity, where the intended pharmacologic mode of action affects fundamental pathways in developmental biology or phenotypic data from genetically modified animals may increasingly be included in the overall assessment. Since some concern surrounds the use of data from knockout (KO) mice to accurately predict the risk for pharmaceutical modulation of a target, a deeper understanding of the relevance and predictivity of adverse developmental effects in KO mice for pharmacological target modulation is needed. To this end, we compared the results of embryo-fetal development (EFD) studies for 86 drugs approved by the FDA from 2017 to 2019 that also had KO mouse data available in the public domain. These comparisons demonstrate that data from KO mouse models are overall highly predictive of malformations or embryo-fetal lethality (MEFL) from EFD studies, but less so of a negative outcome in EFD studies. This information supports the use of embryo-fetal toxicity data in KO models as part of weight of evidence approaches in the communication of developmental toxicity risk of pharmaceutical compounds.