Bringing in vitro analysis closer to in vivo: Studying doxorubicin toxicity and associated mechanisms in 3D human microtissues with PBPK-based dose modelling

Marcha Verheijen*, Yannick Schrooders, Hans Gmuender, Ramona Nudischer, Olivia Clayton, James Hynes, Steven Niederer, Henrik Cordes, Lars Kuepfer, Jos Kleinjans, Florian Caiment

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review


Doxorubicin (DOX) is a chemotherapeutic agent of which the medical use is limited due to cardiotoxicity. While acute cardiotoxicity is reversible, chronic cardiotoxicity is persistent or progressive, dose-dependent and irreversible. While DOX mechanisms of action are not fully understood yet, 3 toxicity processes are known to occur in vivo: cardiomyocyte dysfunction, mitochondrial dysfunction and cell death. We present an in vitro experimental design aimed at detecting DOX-induced cardiotoxicity by obtaining a global view of the induced molecular mechanisms through RNA-sequencing. To better reflect the in vivo situation, human 3D cardiac micro tissues were exposed to physiologically-based pharmacokinetic (PBPK) relevant doses of DOX for 2 weeks. We analysed a therapeutic and a toxic dosing profile. Transcriptomics analysis revealed significant gene expression changes in pathways related to "striated muscle contraction" and "respiratory electron transport", thus suggesting mitochondrial dysfunction as an underlying mechanism for cardiotoxicity. Furthermore, expression changes in mitochondrial processes differed significantly between the doses. Therapeutic dose reflects processes resembling the phenotype of delayed chronic cardiotoxicity, while toxic doses resembled acute cardiotoxicity. Overall, these results demonstrate the capability of our innovative in vitro approach to detect the three known mechanisms of DOX leading to toxicity, thus suggesting its potential relevance for reflecting the patient situation. Our study also demonstrated the importance of applying physiologically relevant doses during toxicological research, since mechanisms of acute and chronic toxicity differ.
Original languageEnglish
Pages (from-to)184-192
Number of pages9
JournalToxicology Letters
Publication statusPublished - 15 Sept 2018


  • Doxorubicin
  • Cardiotoxicity
  • Transcriptomics
  • Physiologically-based pharmacokinetic modeling
  • 3D microtissues
  • Mitochondrial dysfunction

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