Table 1:

Utility of genomics for de-risking positive findings in rodent 2-year carcinogenicity studies - pharmaceutical development case studies

Rodent 2-year carcinogenicity study outcomes	Proposed mode of action	Genomic data	Human relevance?	Impact
Histiocytic sarcoma in multiple tissues (mouse; not rat)	Unknown	1) Deep sequencing of mouse histiocytic sarcomas excluded viral-based tumor etiology. 2) Early onset and persistent activation of IFN γ-associated gene expression signature followed by late onset of histiocytic sarcoma-associated gene expression signature in affected mouse tissues (not observed in rats or in PBMCs from patients in a clinical phase IIb study)	Likely human irrelevant (based on species differences in molecular pathways associated with mouse histiocytic sarcomas)	Reassured health authority and supported further clinical development.
Liver/thyroid tumors (mouse and rat)	CAR activator	1) CAR activator-induced gene expression signatures observed in liver (Peffer et al., 2018; Rooney et al., 2018) from rat 4 week repeat dose toxicity study 2) Absence of AhR-, PPARα, activator-induced gene expression signatures in liver from rat 4 week repeat dose toxicity study (Rooney et al., 2018)	Human-irrelevant mode of action based on established species differences in CAR-mediated proliferative responses and human epidemiological data on phenobarbital	Reassured health authority and supported further clinical development; Integration of genomics and histopathology data from subchronic toxicology studies would also have supported a Category 3a carcinogenicity study waiver under the proposed ICH S1 revision.
Liver/ lung tumors (mouse and rat)	Sustained AhR activation	Careful attention to dose-response with regard to thresholds and kinetics of CYP1A1 and CYP1A2 activation as liver tissue transcriptomic biomarkers of AhR activation tone (Qin et al., 2019; Taylor et al., 2015)	Likely human relevant but only when a therapeutic dose level may be associated with sustained activation above a defined threshold	Reassures both sponsor and health authorities when hypertrophy/ hyperplasia seen in a chronic study is not associated with sustained suprathreshold AhR activity