fish replacement - plymouth.ac.uk
TRANSCRIPT
Fish replacement: Current status minimising the use of fish in tests
Dr Stewart Owen
Current trends in fish in vitro toxicology: Applications of 3Rs principles SETAC Brussels.
10th May, 2017
Peddling an alternative path
Standing on giant shoulders…
Prof Awadhesh Jha
Dr Matt Baron
Dr Richard Maunder
Dr Laura Langan
Prof John Sumpter
Dr Luigi Margiotta-Casaluci
Prof Mariann Rand-Weaver
Dr Nic Bury, Prof Christer Hogstrand
Dr Leon Barron, Dr Tom Miller
Dr Lucy Stott, Elisabeth Chang
Prof Charles Tyler
Dr Lina Gunnarsson
Dr Jon Green
Prof Kevin Chipman
Dr Chibuzor Uchea
And many more……
Dr Tim Martin
Dr Russell Davenport
Dr Andrew Goodhead
Prof Alistair Boxall
Dr Philipp Antczak
How could you use fewer fish?
A few approaches that we have been trying:
Build a virtual fish
In vitro alternatives – Gills, Guts and Livers
Build a fish that isn’t really a fish
In vivo early life stages
Build a prediction
Get it right first time with qAOP predictions
What about: Could we save fish in standard designs?
What is the point of a freshwater and solvent control?
Get the biodegradation assays right
Prof Jason Snape (AZ)
Newcastle University
Dr Tim Martin
Dr Russell Davenport
Dr Andrew Goodhead
80% of chemicals and 95% of APIs
currently fail the key biodegradation
screening tests (OECD 301 series tests)
used to assign chemical persistence
Find a route to the top…
Pharmacology is equivalent:
fish plasma concentration within
pharmacological range
Validation of PECs and in vivo
plasma concentrations and effects in fish
No Risk
Risk prioritisation
In vitro fish
metabolism models
Bio-
informatic
read-across
Fish plasma
concentration
Target
conservation in fish
Human and
mammalian data
Pharmaceutical
s enter fish
In silico
uptake models
In vitro
gill cell uptake
models
Below human Cmax No Risk
Dilution &
%
connectivit
y
Average
amount
of API per
capita per day
PECExcreta
PECInfluent
PECEffluent
PECSurface-water
% metabolism &
metabolite
identification
Dilution & %
connectivity
Dilution, mixing and
degradation (½ life)
Partitioning (Kd)
and degradation (½
life)
PECSediment
PECSludge PECSoil
PECGroundwater
Partitioning (Kd)
and degradation (½
life)
Partitioning (Kd),
leaching &
degradation (½ life)
Applicatio
n rate and
mixing
depth
MECSediment MECGroundwater
MECSoil
MECSurfacewater
Pharmacology is equivalent:fish plasma concentration within
pharmacological range
Validation of PECs and in vivoplasma concentrations and effects in fish
Low Risk
Risk prioritisation
In vitro fish metabolism models
Bio-informaticread-across
Fish plasmaconcentration
Targetconservation in fish
Human andmammalian data
Pharmaceuticals enter fish
In silicouptake models
In vitrogill cell uptake
models
Below human CmaxLow Risk
Dilution & % connectivity
Average amount
of API per
capita per day
PECExcreta
PECInfluent
PECEffluent
PECSurface-water
% metabolism & metabolite identification
Dilution & % connectivity
Dilution, mixing anddegradation (½ life)
Partitioning (Kd)and degradation (½ life)
PECSediment
PECSludgePECSoil
PECGroundwater
Partitioning (Kd)and degradation (½ life)
Partitioning (Kd), leaching &
degradation (½ life)
Application rate and
mixing
depth
MECSedimentMECGroundwater
MECSoil
MECSurfacewater
TR128: Guidance on assessment and application of Adverse
Outcome Pathways (AOPs) relevant to the Endocrine System
ISSN-2079-1526-128 (online)
ECETOC 2016
Can we use AOPs?
Can an AOP be predictive?
Can an AOP be quantitative?
Can a Path be a Network?
Quantitative AOP might be predictive
Margiotta-Casaluci L., et al. (2016). Internal exposure dynamics drive the
Adverse Outcome Pathways of synthetic glucocorticoids in fish. Nature
Scientific Reports 6: 21978.
How do you use fewer fish?
Could you try:
Build a virtual fish
In vitro alternatives – Gills, Guts and Livers
Build a fish that isn’t really a fish
In vivo early life stages
Build a prediction
Get it right first time with qAOP predictions
What about:
Could we save fish in standard designs?
What is the point of a freshwater and solvent control?
Confidentiality Notice
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24
Thank you:
To all our collaborators, colleagues and funding partners.
BBSRC/AstraZeneca IPA (BB/I00646X/1) ‘Mode-Of-Action Ecotoxicology of
Pharmaceuticals’. PIs Prof John Sumpter and Dr Mariann Rand-Weaver. Dr Luigi
Margiotta-Casaluci. July 2011-June 2014. Brunel University.
BBSRC/AstraZeneca IPA (BB/L01016X/1) ‘Moving up a dimension: 3D in vitro models
as effective alternatives to live fish studies’. PIs Prof Awadhesh Jha and Prof Simon
Jackson. Dr Matthew Baron and Dr Richard Maunder March 2014-Feb 2017.
Plymouth University.
Innovative Medicines Initiative (IMI) under grant agreement no.115735—iPiE: Intelligent
led assessment of Pharmaceuticals in the Environment; resources of which are
composed of financial contribution from the European Union's Seventh Framework
Programme (FP7/2015-2018) and European Federation of Pharmaceutical Industries
and Associations (EFPIA) companies' in kind contribution
Refs
• Baron M.G., Mintram K.S., Owen S.F., Hetheridge M.J., Moody J.A., Purcell
W.M., Jackson S.K., Jha J.N. (2017). Pharmaceutical metabolism in fish: using
a 3-D hepatic in vitro model to assess clearance. PLoS one 12(1), e0168837.
doi:10.1371/journal.pone.0168837
• Patel A., Panter G.H., Trollope H.T., Glennon Y.C., Owen S.F., Sumpter J.P.,
Rand-Weaver M. (2016). Testing the “read-across hypothesis” by investigating
the effects of ibuprofen on fish. Chemosphere 163, 592-600
• Miller T.H., Baz-Lomba J.A., Harman C., Reid M.J., Owen S.F., Bury N.R.,
Thomas K.V., Barron L.P. (2016). The first attempt at non-linear in silico
prediction of sampling rates for polar organic chemical integrative samplers
(POCIS). Environmental Science & Technology 50 (15), 7973-7981. [Editors
choice award and cover of issue]
• Margiotta-Casaluci L., Owen S.F., Huerta B., Rodríguez-Mozaz S., Kugathas
S., Barceló D., Rand-Weaver M., Sumpter J.P. (2016). Internal exposure
dynamics drive the Adverse Outcome Pathways of synthetic glucocorticoids in
fish. Nature Scientific Reports 6: 21978.
• McCarthy I.D., Owen S.F., Watt P.W., Houlihan D.F. (2016). Individuals
maintain similar rates of protein synthesis over time on the same plane of
nutrition under controlled environmental conditions. PloS one 11 (3),
e0152239.
• Langan L.M., Dodd N.J.F., Owen S.F., Purcell W.M., Jackson S.K., Jha A.N.
(2016). Direct measurements of oxygen in the middle of 3D tissue cultures
using electron paramagnetic resonance spectroscopy. PloS one 11 (2),
e0149492.
• Green J.M., Metz J., Lee O., Trznadel M., Takesono A., Brown A.R., Owen
S.F., Kudoh T., Tyler C.R. (2016). High-Content and Semi-Automated
Quantification of Responses to Estrogenic Chemicals Using a Novel
Translucent Transgenic Zebrafish. Environmental Science & Technology 50
(12), 6536–6545.
• Schnell, S., Stott, S.C., Hogstrand, C., Wood, C.M., Kelly, S.P., Pärt, P., Owen,
S.F., & Bury N.R. (2016). Procedures for the reconstruction, primary culture
and experimental use of rainbow trout gill epithelia. Nature Protocols 11 (3),
490-498.
• Miller T.H., McEneff G.L., Stott L.C., Owen S.F., Bury N.R., & Barron L.P.
(2016). Assessing the reliability of uptake and elimination kinetics modelling
approaches for estimating bioconcentration factors in the freshwater
invertebrate, Gammarus pulex. Science of The Total Environment 547, 396-
404.
• Brown A.R., Owen S.F., Peters J., Zhang Y., Soffker M., Paull G.C., Hosken
D.J., Wahab M.A., Tyler C.R. (2015). Climate change and pollution speed
declines in zebrafish populations. Proceedings of the National Academy of
Sciences 112 (11), E1237-E1246.
• Uchea C., Owen S.F., Chipman K. (2015). Functional xenobiotic metabolism
and efflux transporters in trout hepatocyte spheroid cultures. Toxicology
Research 4 (2), 494-507.
• Miller T.H., McEneff G.L., Brown R.J., Owen S.F., Bury N.R., Barron L.P.
(2015). Pharmaceuticals in the freshwater invertebrate, Gammarus pulex,
determined using pulverised liquid extraction, solid phase extraction and liquid
chromatography–tandem mass spectrometry. Science of The Total
Environment 511, 153-160.
• Stott L.C., Schnell S., Hogstrand C., Owen S.F., Bury N.R. (2015). A primary
fish gill cell culture model to assess pharmaceutical uptake and efflux:
Evidence for passive and facilitated transport. Aquatic Toxicology 159, 127–
137.
• Corcoran J., Winter M.J., Lange A., Cumming R., Owen S.F, Tyler C.R. (2015).
Effects of the lipid regulating drug clofibric acid on PPARα-regulated gene
transcript levels in common carp (Cyprinus carpio) at pharmacological and
environmental exposure levels. Aquatic Toxicology 161, 127-137.
• Margiotta-Casaluci L., Owen S.F., Cumming R.I., de Polo A., Winter M.J.,
Panter G.H., Rand-Weaver M., Sumpter J.P. (2014). Quantitative cross-species
extrapolation between humans and fish: The case of the anti-depressant
fluoxetine. PloS one 9 (10), e110467.
• Parker T., Libourel A-P., Hetheridge M.J., Cumming R.I., Sutcliffe T.P.,
Goonesinghe A.C., Ball J.S., Owen S.F., Chomis Y., Winter M.J., (2014). A
multi-endpoint in vivo larval zebrafish (Danio rerio) model for integrated
cardiovascular functional assessment. Journal of Pharmacological and
Toxicological Methods 69(1):30-38.
• Corcoran J., Lange A., Cumming R., Owen S.F., Ball J.S., Tyler C.R., Winter
M.J. (2014). Bioavailability of the imidazole antifungal agent clotrimazole and
its effects on key biotransformation genes in the common carp (Cyprinus
carpio). Aquatic Toxicology 152, 57-65.
• Rand-Weaver M., Margiotta-Casaluci L., Patel A., Panter G.H., Owen S.F.,
Sumpter J. (2013). The Read-Across Hypothesis and Environmental Risk
Assessment of Pharmaceuticals. Environmental Science and Technology 47
(20), 11384–11395.
• Readman G.D., Owen S.F., Murrell J.C., Knowles T.G. (2013). Do fish perceive
anaesthetics as aversive? PLoS ONE 8(9): e73773.
• Uchea C., Sarda S., Schulz-Utermoehl T., Owen S., Chipman, K. (2013). In
vitro models of xenobiotic metabolism in trout for use in environmental
bioaccumulation studies. Xenobiotica. 43(5), 421-431.
• Bickley L.K., Brown A.R., Hosken D.J., Hamilton P.B., Le Page G., Paull G.C.,
Owen S.F., and Tyler C.R. (2012). Interactive effects of inbreeding and
endocrine disruption on reproduction in a model laboratory fish. Evolutionary
Applications. 6(2): 279-289.
• Baron M.G., Purcell W.M., Jackson S.K., Owen S.F., Jha A.N. (2012). Towards
a more representative in vitro method for fish ecotoxicology: morphological and
biochemical characterisation of three-dimensional spheroidal hepatocytes.
Ecotoxicology 21(8):2419-2429.
• Wilkes, L., Owen, S.F., Readman, G.D., Sloman, K.A. and Wilson, R.W. (2012).
Does structural enrichment for toxicology studies improve zebrafish welfare?
Applied Animal Behaviour Science 139(1): 143-150.
• Brown A.R., Bickley L.K., Ryan T.A., Paull G.C., Hamilton P.B., Owen S.F.,
Sharpe A.D. and Tyler C.R. (2012). Differences in sexual development in
inbred and outbred zebrafish (Danio rerio) and implications for chemical
testing. Aquatic Toxicology. 112–113: 27–38.
• Bartram, A.E., Huggett, D.B., Hutchinson, T.H., Hetheridge, M.J., Kinter, L.B.,
Ericson, J.F., Constantine, L.B., Sumpter, J.P. Owen, S.F. (2012). Liver and gill
EROD activity in rainbow trout (Oncorhynchus mykiss) exposed to the beta-
blocker Propranolol in vivo and in vitro. Environmental Toxicology 27(10): 573–
582.
• Brown, A.R., Bickley, L.K., Le Page, G., Hosken, D.J., Paull, G.C., Hamilton,
P.B., Owen, S.F., Robinson, J., Sharpe, A.D., Tyler, C.R. (2011). Are
toxicological responses in laboratory (inbred) Zebrafish representative of those
in outbred (wild) populations? A case study with an endocrine disrupting
chemical. Environmental Science and Technology 45: 4166–4172.
• Owen, S.F., Huggett, D.B., Hutchinson, T.H., Hetheridge, M.J., Kinter, L.B.,
Ericson, J.F., Constantine, L.B. and Sumpter, J.P. (2010). The value of
repeating studies and multiple controls: replicated 28‐day growth studies of
rainbow trout exposed to clofibric acid. Environmental Toxicology and
Chemistry 29(12): 2831–2839.
• Winter M.J., Owen S.F., Murray-Smith R., Panter G.H., Hetheridge M.J, Kinter,
L.B. (2010). Using preclinical data to aid the aquatic Environmental Risk
Assessment of human pharmaceuticals: concepts, considerations and
challenges. Integrated Environmental Assessment and Management 6(1): 38–
51.
• Brown, A.R., Hosken, D.J., Balloux, F., Bickley, L.K., LePage, G., Owen, S.F.,
Hetheridge, M.J., Tyler, C.R. (2009) Genetic variation, inbreeding and chemical
exposure - combined effects in wildlife and critical considerations for
ecotoxicology. Philosophical Transactions of the Royal Society B 364: 3377-
3390.
• Owen S.F., Huggett D.B., Hutchinson T.H., Hetheridge M.J., Kinter L.B.,
Ericson J.F., Sumpter J.P. (2009) Uptake of propranolol, a cardiovascular
pharmaceutical, from water into fish plasma and its effects on growth and
organ biometry. Aquatic Toxicology 93(4): 217-224.
• Williams T.D., Readman G.D., Owen S.F. (2009). Key issues concerning
environmental enrichment for laboratory-held fish species. Laboratory Animals
43:1-14.
• Winter M.J., Redfern W.S., Owen S.F., Valentin J-P., Hutchinson T.H. (2008).
Validation of a larval zebrafish locomotor assay for assessing the seizure
liability of early-stage development drugs. Journal of Pharmacological and
Toxicological Methods 57(3): 176 –187 [Most cited article in journal]
• Owen, S.F., Giltrow, E., Huggett, D.B., Hutchinson, T.H., Saye, J.-A, Winter,
M.J., and Sumpter, J.P. (2007). Comparative physiology, pharmacology and
toxicology of β-blockers: mammals versus fish. Aquatic Toxicology 82: 145–
162.