Predicting carcinogenic mechanisms of non-genotoxic carcinogens via combined analysis of global dna methylation and in vitro cell transformation

Sung Hee Hwang, Hojin Yeom, Byeal I. Han, Byung Joo Ham, Yong Moon Lee, Mi Ryung Han, Michael Lee

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)


An in vitro cell transformation assay (CTA) is useful for the detection of non-genotoxic carcinogens (NGTXCs); however, it does not provide information on their modes of action. In this study, to pursue a mechanism-based approach in the risk assessment of NGTXCs, we aimed to develop an integrated strategy comprising an in vitro Bhas 42 CTA and global DNA methylation analysis. For this purpose, 10 NGTXCs, which were also predicted to be negative through Derek/Sarah structure–activity relationship analysis, were first tested for transforming activity in Bhas 42 cells. Methylation profiles using reduced representation bisulfite sequencing were generated for seven NGTXCs that were positive in CTAs. In general, the differentially methylated regions (DMRs) within promoter regions showed slightly more bias toward hypermethylation than the DMRs across the whole genome. We also identified 13 genes associated with overlapping DMRs within the promoter regions in four NGTXCs, of which seven were hypermethylated and six were hypomethylated. Using ingenuity pathway analysis, the genes with DMRs at the CpG sites were found to be enriched in cancer-related categories, including “cell-to-cell signaling and interaction” as well as “cell death and survival”. Moreover, the networks related to “cell death and survival”, which were considered to be associated with carcinogenesis, were identified in six NGTXCs. These results suggest that epigenetic changes supporting cell transformation processes occur during non-genotoxic carcinogenesis. Taken together, our combined system can become an attractive component for an integrated approach for the testing and assessment of NGTXCs.

Original languageEnglish
Article number5387
Pages (from-to)1-18
Number of pages18
JournalInternational journal of molecular sciences
Issue number15
Publication statusPublished - 2020 Aug 1

Bibliographical note

Funding Information:
This research was supported by a grant (18182MFDS367) from Korea’s Ministry of Food and Drug Safety in 2019 and by the National Research Foundation of Korea (NRF) grant funded by the Ministry of Science and ICT (MIST) (NRF-2019R1F1A1048733). The content is solely the responsibility of the authors and does not necessarily represent the official views of the Ministry of Food and Drug Safety. This research was also supported by the Collaborative Genome Program for Fostering New Post-Genome Industry of the National Research Foundation (NRF) funded by the Ministry of Science and ICT (MSIT) (NRF-2017M3C9A6047623).

Publisher Copyright:
© 2020 by the authors. Licensee MDPI, Basel, Switzerland. T.


  • In vitro cell transformation assay
  • Methylation profiles
  • Non-genotoxic carcinogen
  • RRBS

ASJC Scopus subject areas

  • Catalysis
  • Molecular Biology
  • Spectroscopy
  • Computer Science Applications
  • Physical and Theoretical Chemistry
  • Organic Chemistry
  • Inorganic Chemistry


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