Increased, Retinoic Acid Receptor Activation

This KE refers to increased activation of the retinoic acid receptor (RAR) occurring in complex biological systems in vivo, such as tissues and organs. Thus, increased activation of RAR is here distinguished from direct agonism of the RAR and from increased (or ectopic) availability or synthesis of its endogenous ligand, all‑trans retinoic acid (atRA). Instead, it may occur downstream of these events. This KE occurs in tissues and organs in vivo, and it is not a tissue‑specific event.

RARs are ligand-activated receptors regulated by retinoid ligands, particularly atRA. They mediate several crucial biological processes (e.g., morphogenesis, cell growth regulation, apoptosis, homeostasis etc.). Perturbations of RAR signaling are associated with adverse developmental and physiological outcomes. RARs are members of the nuclear hormone receptor superfamily. There are three major subtypes: RARα, RARβ, and RARγ. RARα may be broadly expressed, while RARβ is related to neurogenesis and development of epithelial tissues, and RARγ in bone and cartilage development. There are different isoforms for each subtypes arising from alternative splicing (RARα1-2, RARβ1–4, and RARγ1-2) (Germain et al, 2006, Petkovich et al, 2022).

The canonical signaling mechanism of RARs involves heterodimerization with retinoid X receptors (RXRs), DNA binding to retinoic acid response elements (RAREs), and ligand‑dependent regulation of transcription. RARs heterodimerize with RXRs, with the RAR-RXR complex binding to RAREs in promoters of their target genes. In the absence of ligand, RAR–RXR complexes are often associated with co‑repressor proteins (e.g., NCoR and SMRT), maintaining transcriptional repression. Upon ligand binding with atRA or other retinoids, the RAR–RXR heterodimers undergo conformational changes that promote dissociation of co‑repressors and recruitment of co‑activator proteins (e.g., SRC1-3), leading to chromatin remodeling and transcriptional activation of several target genes (Germain et al, 2006, Petkovich et al, 2022, Rhinn et al, 2012, Gutierrez et al, 2014).

Increased RAR activation can result from direct agonism of RAR or from increased availability of atRA, its main endogenous ligand; for instance by inhibiting CYP26 enzyme activity required for degradation of atRA (Ross et al, 2013). RAR agonism can arise from several retinoids that possess higher potency compared to the endogenous ligand. Increased RAR activation can also be caused by increased atRA levels (due to increased synthesis, e.g., CYP26B inhibition) that has been associated with homeostasis imbalances. Both mechanisms increase RAR‑dependent gene expression and disrupt the tightly controlled retinoid‑regulated biological processes (Germain et al, 2006, Altucci et al, 2007, Stevison et al, 2017, Kedishvili et al, 2013).

Non‑canonical RAR signaling mechanisms, independent of direct transcriptional regulation, have been described in some experimental systems. However, their conservation and quantitative contribution to adverse outcomes in vivo remain unclear (Al Tanoury et al 2013). Accordingly, this KE addresses decreased RAR activation via the canonical, transcription‑dependent pathway.

This KE describes increased RAR activation in vivo; however, presently there are no methods available to directly measure this endpoint. Currently, indirect assessment of RAR activation is limited to in vitro RAR and RAR-RXR transactivation assays (OECD Detailed Paper Review on Retinoids, Validation report on a RAR transactivation method). AOP developers are encouraged to add new methods to measure RAR activation levels in vivo whenever they become available.

RARs are well-conserved throughout vertebrate evolution, therefore this KE should be considered broadly applicable across vertebrate species. Regarding invertebrates, evidence for the presence of RARs has been reported within chordate phyla, including non-vertebrate chordates. However, multiple invertebrate lineages appear to have lost RAR genes. Although components of RA signaling have been described in some invertebrate species, it remains debated whether they possess a functional RA signaling pathway (Stevison et al, 2017). AOP developers are encouraged to add additional relevant knowledge to expand on the applicability to also include other vertebrates.

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Altucci L, Leibowitz MD, Ogilvie KM, de Lera AR, Gronemeyer H. RAR and RXR modulation in cancer and metabolic disease. Nat Rev Drug Discov. 2007;6(10):793-810. doi:10.1038/nrd2397

Kedishvili NY. Enzymology of retinoic acid biosynthesis and degradation. J Lipid Res. 2013;54(7):1744-1760. doi:10.1194/jlr.R037028

Germain P, Chambon P, Eichele G, et al. International Union of Pharmacology. LX. Retinoic acid receptors. Pharmacol Rev. 2006;58(4):712-725. doi:10.1124/pr.58.4.4

Gutierrez-Mazariegos J, Schubert M, Laudet V. Evolution of retinoic acid receptors and retinoic acid signaling. Subcell Biochem. 2014;70:55-73. doi:10.1007/978-94-017-9050-5_4

Organisation for Economic Co-operation and Development (OECD). Detailed Review Paper on the Retinoid System. OECD Environment Directorate, Chemicals and Biotechnology Committee; 2021. Accessed April 23, 2026. https://www.oecd.org/en/publications/detailed-review-paper-on-the-retinoid-system_4fbb70a9-en.html

Organisation for Economic Co‑operation and Development (OECD). Peer Review of the Retinoic Acid Receptor Transactivation Method Validation: Validation Report. OECD; 2025. Accessed April 23, 2026. https://www.oecd.org/content/dam/oecd/en/events/2025/09/peer-review-of-the-retinoic-acid-receptor-transactivation-method-validation/validation-report-retinoic-acid-receptor-transactivation-method.pdf

Petkovich M, Chambon P. Retinoic acid receptors at 35 years. J Mol Endocrinol. 2022;69(4):T13-T24. Published 2022 Oct 11. doi:10.1530/JME-22-0097

Rhinn M, Dollé P. Retinoic acid signalling during development. Development. 2012;139(5):843-858. doi:10.1242/dev.065938

Ross AC, Zolfaghari R. Cytochrome P450s in the regulation of cellular retinoic acid metabolism. Annu Rev Nutr. 2011;31:65-87. doi:10.1146/annurev-nutr-072610-145127

Stevison F, Hogarth C, Tripathy S, Kent T, Isoherranen N. Inhibition of the all-trans Retinoic Acid (atRA) Hydroxylases CYP26A1 and CYP26B1 Results in Dynamic, Tissue-Specific Changes in Endogenous atRA Signaling. Drug Metab Dispos. 2017;45(7):846-854. doi:10.1124/dmd.117.075341