Systematic evaluation of Tox21 compounds that target β-adrenergic receptors and their role in cardiotoxicity

Abstract

β-adrenergic receptors play important roles in heart failure and drug-induced cardiotoxicity (DICT). The Tox21 10 K library of drugs and environmental chemicals have been tested for their activity against β-adrenergic receptor subtypes 1 and 2 (ADRB1 and ADRB2), as well as inhibition of the human ether-à-go-go-related gene (hERG) in a quantitative high-throughput screening (qHTS) format. In this study, the Tox21 compound activity profiles in the ADRB1/2 and hERG assays were compared in relation to their DICT potential. The results showed that compounds that acted as ADRB1 agonists, ADRB2 antagonists, or hERG inhibitors were more likely to exhibit DICT. The ADRB1 and ADRB2 assays shared similar compound activity profiles, while the hERG inhibition assay identified a distinct set of active compounds. In addition, we identified structural features that may differentiate the cardiotoxic and non-toxic ADRB1 agonists. Finally, machine learning models were developed for ADRB1 activity prediction based on chemical structure. The models were used to virtually screen a collection of approximately 360 K diverse compounds, with the highest-ranked compounds selected for experimental validation. This work represents the first systematic study of drugs and environmental chemicals against ADRB1/2, providing important insights into β-adrenergic receptor-related cardiotoxicity mechanisms. By clarifying how specific pharmacological interactions contribute to cardiac risk, it provides a framework for early cardiotoxicity prediction and the design of safer therapeutics through integrated profiling and modeling.

Keywords: Cardiotoxicity; High-Throughput Screening; In Vitro Assay; Tox21; hERG; β-Adrenergic Receptors.

Fig. 1.

ADRB1 vs. ADRB2 activity comparison, each grid shows numbers of compounds with activities for ADRB1 and ADRB2 in both agonist (left) and antagonist (right) modes.

Fig. 2.

Distribution of compounds by DICT scores, a measure of cardiotoxicity ranging from 0 to 1, for 1001 compounds in the Tox21 10 K compound library that fall into different ADRB and hERG activity categories, with compound numbers and mean DICT scores of assay activity category in each panel. The x-axis represents the range of DICT scores, while the y-axis indicates the number of compounds.

Fig. 3.

Comparison of active rates of compounds in different assays, i.e., ADRB1/ADRB2 in agonist and antagonist modes and hERG inhibition, stratified by their cardiotoxicity potential measured by DICT score (cardiotoxic: >0.50; non-toxic: <0.20). Active rate is defined as the fraction of active compounds in a specific assay, e.g., ADRB1 agonist, that also fall into a specific cardiotoxicity category, e.g., cardiotoxic (DICT score > 0.50). The enrichment of cardiotoxic compounds in each assay activity category was evaluated by the p-value derived from the Fisher’s exact test comparing the active rates of compounds falling into different DICT score categories.

Fig. 4.

Activity profiles of the 589 cardiotoxic compounds in the Tox21 10 K library (DICT score > 0.20) against different targets including hERG inhibition, ADRB1/2 agonist and antagonist. In the heat map, each column is a compound, and each row is an assay. The heat map is colored by compound activity which can be categorized into: “active agonist”, “inconclusive agonist”, “active antagonist”, “inconclusive antagonist”, “inconclusive”, and “inactive”. A dark red color indicates a strong activator, a dark blue color indicates a strong inhibitor, and inactive compounds are colored in white. Dark gray indicates inconclusive data.

Fig. 5.

2D t-SNE projection of 182 cardiotoxic compounds (DICT score > 0.2) that were also ADRB agonists/antagonists or hERG inhibitors based on ToxPrint.

Fig. 6.

Experimental validation of model-predicted ADRB1 agonists/antagonists. Each panel contains the concentration response curve of a compound, ADRB1 activation (EC₅₀) or inhibition (IC₅₀) potency and compound structure. Compounds from A-C are representative active ADRB1 agonists, compounds from D—F are representative ADRB1 antagonists. Results (EC₅₀ for agonists and IC₅₀ for antagonists) are presented as mean ± standard deviation (SD), and the error bars represent the SD of four independent experiments.