Tox21 Enricher: Web‐based chemical and functional enrichment analysis tool for Tox21 toxicity screening platform
Authors: Hur Junguk , Danes Larson , Krout Dakota , Hsieh Jui‐Hua , Auerbach Scott 
Published: November 2023

Humans are exposed to tens of thousands of chemicals that are used in our daily life, some at levels that may pose a health risk. For many of these chemicals, there are limited toxicological information which makes risk assessment impossible. The United States Toxicology Testing in the 21st Century (Tox21) program was established to develop more efficient and human relevant toxicity assessment methods. The Tox21 program is currently screening over 10,000 chemicals, the Tox21 10K library, using quantitative high‐throughput screening (qHTS) of assays that measure effects on toxicity pathways. To date, more than 70 assays have yielded >12 million concentration‐response curves by Tox21 researchers. One approach to using this data is to perform chemical‐relational analysis based on the patterns of activity across Tox21 assays and then to use nearest neighbor‐based prediction to infer the toxicological properties of untested chemicals via their association with tested chemicals. One approach to inferring the specific properties is to perform chemical annotation enrichment of chemical neighborhoods. Here, we present Tox21 Enricher, a web‐based chemical annotation enrichment tool for Tox21 assay data. Tox21 Enricher identifies significantly over‐represented chemical and biological annotations among lists of chemicals (neighborhoods) facilitating the identification of the toxicological properties or mechanisms in the chemical set. Currently, Tox21 Enricher supports 34 chemical annotation categories, including KEGG pathway ChemDisease, and Gene Ontology from Comparative Toxicogenomics Database; known target and Anatomical Therapeutic Chemical codes from DrugBank; predicted in vivo and in vitro toxicity; Chemical substructures from ToxPrint; a number of clinical and toxicological classifications from DrugMatrix; PubChem PharmAction and MeSH terms. Tox21 Enricher uses a modified Fisher's exact test, a widely used statistical test in gene set enrichment analysis, to assess the significance of enrichment. Tox21 Enricher allows users to submit lists of chemicals using their CAS Registry Numbers as input. Enriched chemical and biological annotations are presented in multiple formats: matrices in Excel file, heatmap images visualizing the level of similarity and dissimilarity of enriched annotations across the input chemical sets, and annotation network views illustrating the associations among the significant annotations. Tox21 Enricher also allows users to search using chemical structures (SMILES strings) that are not contained in the Tox21 10K library. This feature allows researchers to identify list of chemicals that structurally similar to chemicals in the library and then perform chemical annotation enrichment to infer the potential toxicological properties on chemicals not in the Tox21 library.

Tox21 Enricher is freely available at http://hurlab.med.und.edu/Tox21Enricher. Powered with advanced features like concept‐network visualization and structure‐based query, Tox21 Enricher is expected to enhance the toxicity research.