Should metabolic saturation of 1,4-DX be considered as a necessary precursor event for the following tumor endpoints that could be used as a point of departure (i.e., if you are protective of metabolic saturation, you are then also protective of tumor response)?

Answer Explanations

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  Expert 4

  Endpoint	Yes	No	I cannot answer
  Rodent liver tumors	1	0	0
  Rat nasal cavity squamous cell carcinoma	0	1	0
  Rat peritoneal mesothelioma	0	1	0

  Normally, integrative analysis for the dose-response information for each/all of the key events in the hypothesized MOA is preferred rather than reliance on a single precursor event. The choice of yes/no relates also not only to relative confidence in the data supporting metabolic saturation being a key event but also to the required degree of certainty (i.e., science policy).  There are no data on plasma levels of 1,4-DX in the critical carcinogenesis bioassays in which tumours were observed.  Longest term data are restricted to 90 day studies and the range reported for metabolic saturation is broad (e.g., 30 - 100 ug/mL blood in Lacanfroni et al., 2023).
  Confidence that cyp 2E1 induction is a critical key event in the mode of action for 1, 4-DX based on the available data is greatest for rodent liver tumors followed by rat nasal squamous cell carcinomas. For peritoneal  mesotheliomas, relevant data and analyses are lacking. Evidence that is most convincing for liver tumours in rodents include unpublished data addressing essentiality (i.e., that in cyp 2E1 knockout mice referenced in the commentary of Wang et al. (2022) and empirical support (with few exceptions dose-reponse and temporal concordance support tumours being observed at high doses likely to induce cyp 2E1).  Additional data on toxicokinetics in mechanistic studies investigating dose-response for hypothesized key events and cyp 2E1 knock out models (to consider essentiality) are likely to more robustly support metabolic saturation as a necessary precursor event.  
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  Expert 2

  Endpoint	Yes	No	I cannot answer
  Rodent liver tumors	0	1	0
  Rat nasal cavity squamous cell carcinoma	0	1	0
  Rat peritoneal mesothelioma	0	1	0

  The consequences of secondary metabolites on potential events leading to mutagenesis in susceptible populations have not been excluded.  
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  Expert 3

  Endpoint	Yes	No	I cannot answer
  Rodent liver tumors	1	0	0
  Rat nasal cavity squamous cell carcinoma	0	1	0
  Rat peritoneal mesothelioma	1	0	0

  The overall MoA data indicate that, absent an exposure/dose sufficient to stimulate a metabolic cyp2e1 induction response (directly associated with doses sufficient to initially as well as post-induction metabolically saturate cyp2e1), follow-on systemic key events do not occur.  Although likely shared for nasal tumors, the nasal response and the systemic associated MoA is confounded by direct contact with high-dose DX drinking water.  Although the metabolic saturation MoA may not completely be the driver for the nasal MoA, the observation that DX is not genotoxic and nasal tumors are consistently restricted the highest test doses supports a conclusion this response is also a threshold high-dose specific response in rodents that lacks quantitative MoA relevance to humans (high-dose systemic dosing and/or high-dose direct contact MoA).
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  Expert 6

  Endpoint	Yes	No	I cannot answer
  Rodent liver tumors	1	0	0
  Rat nasal cavity squamous cell carcinoma	1	0	0
  Rat peritoneal mesothelioma	1	0	0

  The saturation hypothesis is highly plausible also since saturation with 'overflow' of the parent DX and tumorigenesis are both high dose events and seem to run roughly in parallel when it comes to dose-response considerations. A similar mode of action in the nasal mucosa is also plausible. For mesothelioma a similar mechanism appears plausible. 
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  Expert 5

  Endpoint	Yes	No	I cannot answer
  Rodent liver tumors	1	0	0
  Rat nasal cavity squamous cell carcinoma	0	0	1
  Rat peritoneal mesothelioma	0	0	1

   Based onthe available data, metabolic saturation certainly should be considered as a MOA for 1,4 DX induce liver tumors but some additional experimentl studies seem warranted to fill the data gaps for this MOA.  The nasal tumors and mesothelioma require further study and further experimental data to resolve the MOA of these two tumor sites.