Replacing, refining and reducing animal usage in epilepsy research using a non-sentient model

Epilepsy is the commonest serious neurological condition, resulting in considerable morbidity and mortality. Research into the origins and mechanisms of epilepsy has quickly advanced since seizures can be induced in animals to investigate both the mechanism of epilepsy and to identify new treatments. One epilepsy treatment, Valproic acid (VPA), is now the most widely prescribed drug worldwide, but surprisingly, its mechanism of action remains unclear. This project will follow NC3Rs criteria for research by replacing and reducing animals in developing a non-sentient model system for understanding the basis of the anti-epileptic effects of VPA and in identifying novel epilepsy treatments. This is a necessary approach, since the current analysis of one potential new epilepsy drug employs at least two experimental animal models at five drug doses with eight animals per dose - thus around 100 animals are used per compound. Dictyostelium is a non-sentient biomedical model that we have used as a replacement for animals in understanding the cellular effects of VPA. Our recent discovery of a VPA-catalysed inhibition of phosphoinositide (PI) signalling through attenuating de novo inositol biosynthesis in this model is consistent with this effect being the mechanism for seizure control. We have also shown that increased PI inhibition for VPA-related compounds correlates with increased seizure control, and have shown one novel compound strongly increased seizure-control effects in mice compared to VPA. Our data therefore suggests VPA may function in seizure control through inhibition of de novo inositol biosynthesis. This project will define the mechanism of VPA inhibited inositol attenuation. We will then follow a staged process, first identifying novel compounds showing this effect in Dictyostelium, and then analysing a limited number of drugs in animal seizure models to confirm transferability of these effects to animal systems. The project will finally test three compounds on long-term seizure control using new technology in refined experiments to develop better treatments for epilepsy. The project will therefore replace and reduce animal research by employing a simple non-sentient model to better understand the basis of epilepsy and for the development of new epilepsy treatments, and refine animal usage to prove the efficacy of these drugs. We estimate screening 40 compounds will need 100 animals, reducing animal usage in the development of these drugs by 3900. The project is highly likely to help to unravel the complex way in which VPA stops epilepsy in the human population and to develop new treatments for epilepsy.

Chang P, Augustin K, Boddum K, Williams S, Sun M, Terschak JA, Hardege JD, Chen PE, Walker MC, Williams RS (2016) Seizure control by decanoic acid through direct AMPA receptor inhibition. Brain 139(Pt 2): 431-43. doi:10.1093/brain/awv325 Read the paper

Chang P, Zuckermann AM, Williams S, Close AJ, Cano-Jaimez M, McEvoy JP, Spencer J, Walker MC, Williams RS (2015) Seizure control by derivatives of medium chain fatty acids associated with the ketogenic diet show novel branching-point structure for enhanced potency. Journal of Pharmacology and Experimental Therapeutics 352(1): 43-52. doi:10.1124/jpet.114.218768 Read the paper 

Walker MC, Williams RS (2015) New experimental therapies for status epilepticus in preclinical development.  Epilepsy & Behavior 49: 290-3. doi:10.1016/j.yebeh.2015.06.009 Read the paper

Cunliffe VT, Baines RA, Giachello CN, Lin WH, Morgan A, Reuber M, Russell C, Walker MC, Williams RS (2015) Epilepsy research methods update: Understanding the causes of epileptic seizures and identifying new treatments using non-mammalian model organisms.  Seizure 24:44-51. doi:10.1016/j.seizure.2014.09.018 Read the paper

Chang P. et al. (2014) Seizure-induced reduction in PIP3 levels contributes to seizure-activity and is rescued by valproic acid. Neurobiol Dis. 62(100): 296-306. doi:10.1016/j.nbd.2013.10.017 Read the paper

Chang P. et al. (2013) Seizure control by ketogenic diet-associated medium chain fatty acids. Neuropharmacology 69:105-114. doi:10.1016/j.neuropharm.2012.11.004 Read the paper

Pakes N.K., Veltman D.M., Williams R.B. (2013) Zizimin and Dock guanine nucleotide exchange factors in cell function and disease. Small GTPases. 4(1):22-27. doi:10.4161/sgtp.22087 Read the paper 

Pakes N.K. et al. (2012) The Rac GEF ZizB regulates development, cell motility and cytokinesis in DictyosteliumJ Cell Sci. 125(Pt 10):2457-65. doi:10.1242/jcs.100966 Read the paper

Chang P. et al. (2012) The antiepileptic drug valproic acid and other medium-chain fatty acids acutely reduce phosphoinositide levels independently of inositol in Dictyostelium. Dis. Model. Mech 5(1):115-124. doi:10.1242/dmm.008029 Read the paper

Elphick LM, Pawolleck N, Guschina IA, Chaieb L, Eikel D, Nau H, Harwood JL, Plant NJ, Williams RS (2012) Conserved valproic-acid-induced lipid droplet formation in Dictyostelium and human hepatocytes identifies structurally active compounds. Disease Models and Mechanisms 5(2): 231-40.  doi:10.1242/dmm.008391 Read the paper 

Robery S. et al. (2011) Investigating the effect of emetic compounds on chemotaxis in Dictyostelium identifies a non-sentient model for bitter and hot tastant research Plos One, 6(9):e24439. doi:10.1371/journal.pone.0024439 Read the paper

Terbach N. et al. (2011) Identifying an uptake mechanism for the antiepileptic and bipolar disorder treatment valproic acid using the simple biomedical model Dictyostelium. J. Cell. Sci. 1;124(Pt 13):2267-76. doi:10.1242/jcs.084285 Read the paper

Pakes N.K., Jayasinghe S.N., and Williams R.S (2011) Bio-electrospraying and aerodynamically assisted bio-jetting the model eukaryotic Dictyostelium discoideum: assessing stress and developmental competency post-treatment J. Roy. Soc. Interface. 8(61):1185-91. doi: 10.1098/rsif.2010.0696 Read the paper

Ludtmann M.H., Boeckeler K., and Williams R.S (2011) Molecular pharmacology in a simple model system: implicating MAP kinase and phosphoinositide signalling in bipolar disorder Semin. Cell. Dev. Biol. 22(1):105-13. doi:10.1016/j.semcdb.2010.11.002 Read the paper

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Project grant

Status:

Closed

Principal investigator

Professor Robin Williams

Institution

Royal Holloway University of London

Co-Investigator

Professor Matthew C Walker

Grant reference number

G0900775/1

Award date:

Jan 2010 - Apr 2013

Grant amount

£415,373