The Use of In Vitro Data in Understanding Human Health Effects and Risk

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The Use of In Vitro Data in Understanding Human
Health Effects and Risk

• James Bus, PhD, DABT, ATS
• The Dow Chemical Company
• TestSmart DNT2
• Reston, VA
• November 13, 2008

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NAS/NRC Reports

• Toxicity Testing in the 21st Century A Vision
  and Strategy (2007)
• High-throughput technologies directed at
  toxicity pathways
• Future paradigm of rapid and inexpensive toxicity
  information on broad spectrum of chemicals
• Application of Toxicogenomic Technologies to
  Predictive Toxicology and Risk Assessment (2007)
• Identify challenges and opportunities of new
  technologies for improving risk assessment

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Technical Requirements for High-Throughput
Testing (HTP) Drugs vs Environmental Chemicals

• Pharmaceutical development
• Efficacy and toxicity screening - focused on
  known targets
• Doses near projected therapeutic ranges
• Tolerates low sensitivity (high false negatives
  and positives) comprehensive animal testing
  back-up
• Environmental chemical assessment
• Breadth of testing needed concern for wide
  range of potential toxicities
• Exposures range over many orders of magnitude
• Exposures ongoing in real world

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HTP Challenge Biological Complexity and
Inter-Organ Phenomena
Bus and Popp, Fd.Chem.Tox., 1987
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HTP Challenge Biological Complexity and
Intra-Organ Phenomena

• Example Paraquat vs Diquat
• Both redox cycling oxidants
• Paraquat, but not diquat, actively and
  selectively transported into alveolar cells by
  amine transporter
• HTP screening might not identify paraquat as the
  greater hazard and risk
• Paraquat toxicity controlled by lung cell
  specific transport

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HTP Challenge Dose-Dependent Transitions
Complexities in Conventional In Vivo Animal Tests
Slikker et.al., TAAP 2004
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HTP Challenge (Opportunity!) Dosimetry Context
for Interpretation of Toxicity Responses

• HTP strength allows evaluation over wide
  dose-response range
• but a word of caution
• HTP current focus on prediction of high-dose
  animal toxicity
• Society of Toxicology Risk Assessment Task Force
  
• The relevance of using doses that are many
  multiples of conceivable human exposuresis, at
  most, quite dubious. the predicted risks may
  have little or no relationship to risk in the
  real world. Connolly, Beck Goodman, Tox. Sci.
  49 1-4 (1999)

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HTP Opportunity Characterize Shape of Dose
Response at Low Doses

Naciff et.al., Tox.Sci. 2005
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HTP Challenge (Opportunity!) Providing
Dosimetry Context to in vivo Toxicity Tests

• Animal toxicity studies reported in units of
  external dose (mg/kg/day or ppm)
• Animal toxicity tests often serve as primary
  basis for estimation of human health risks
• HTP data described in units of concentration,
  i.e., reflecting internal dose at target
• Dosimetry of current toxicity tests has no direct
  context to HTP tests
• e.g., NOEL and LOEL values not linked to internal
  blood and/or tissue concentrations

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Determining toxicokinetic parameters for
environmental chemicals in in vivo animal
toxicity studies
Saghir et.al.TAAP, 2006
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Comparing Dosimetry in Rodent Toxicity Tests to
Human Exposures Improving Risk Evaluation
From Hays et.al., Reg.Toxicol.Pharmacol 47
96-109, 2007
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• The objective of HTP testing is to provide more
  toxicity information, more rapidly, on much
  larger numbers of environmental chemicals
• but dont overlook opportunity to refine the
  underlying assumptions and principles of hazard
  and risk assessment
• explore low end of dose-response thresholds?
• rapid insights into key events of mode of
  action
• cross-species extrapolation human relevance?
• susceptible subpopulations when are they
  vulnerable?
• mixtures how to add?
• characterizing compound toxicity by category
  approach

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Conclusions

• Opportunities for new technologies to toxicology
• Improve toxicity screening
• Advance weight-of-evidence analyses
• Increase pace of data generation
• Reduce and refine use of animals
• but, some caveats

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Caveats

• Need tier-based framework for structuring data
  generation
• some considerations
• Built on data-mining of existing DNTs
• Dosimetry information
• Exposure information Margin of Exposure
  triggers
• Mode of Action insights

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Caveats
Build on and integrate with, but dont
overlook experiences and principles developed
from decades of whole animal testing and,
most importantly, dont forget Its the
Dose that Makes the Poison

(Paracelsus)

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HTP Challenge Dose Response Context

• Dose response assessment
• Context to in vivo animal tests
• External dose (ppm mg/kg etc.)
• Internal dose (blood, tissue concentration)
• Context to real world human exposures
• Measured or modeled exposure
• Internal dose assessment (biomonitoring)

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Susceptible subpopulations When are they more
vulnerable?

• Enhanced susceptibility to organophosphate
  insecticide chlorpyrifos due to altered
  metabolism
• PON1 gene encodes protein important to
  detoxification of chlorpyrifos
• Variations in PON1 expression identified in
  humans
• Greater than 60-folod inter-individual difference
  in detoxification
• Opportunity to create PON1 variants in animal
  models
• PON1 knock-outs
• Humanized mice knock-out mouse PON1 and
  knock-in human PON1 variants

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HPT Opportunity Susceptibility

• Impact of PON-1 on Dose Response

PON-1 KO?
Wild-type
Response
Dose