The importance of protein kinase G (PKG) for cGMP-dependent cell death and neuroprotection in inherited retinal neurodegeneration

• Applicant: Professor Dr. Francois Paquet-Durand
• Subject Area: Ophthalmology
• Term: from 2012 to 2015
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 212312876

• Final Report Year: 2015

Final Report Abstract

Retinitis Pigmentosa (RP) is a group of inherited retinal degenerations which constitutes one of the leading causes of blindness in the developed world. These are at present untreatable and the underlying neurodegenerative mechanisms are unknown even though the genetic causes are often established. Since elevated levels of cyclic guanosine monophosphate (cGMP) are responsible for photoreceptor cell death in different animal models for human RP, differential regulation of factors involved in cGMP signalling may represent a novel approach for the treatment of RP. The project investigated cGMP-signalling and its role in photoreceptor degeneration using in vivo, ex vivo, and in vitro techniques. A focus was on the activity of cGMP-dependent protein kinase G (PKG), its potential targets during retinal degeneration, and the identification of neuroprotective strategies to halt or delay processes leading to blindness. This was accomplished by in vitro simulation of hereditary retinal degeneration using the PDE6 inhibitor zaprinast on retinal explant cultures, as well as by in vivo phenotyping of PKG1 knock-out (KO) and double-mutant PKG1 KO*rd1 mice. The follow-up of degenerative events in the same individual animals was performed using state-of-the-art, non-invasive technology for both functional and morphological characterization. Unexpectedly, the analysis of both in vitro and in vivo studies showed that PKG1 had only a minor role in hereditary photoreceptor degeneration, and that, furthermore, PKG1 loss-offunction had no effect on the functionality of photoreceptors and the retina as a whole. Since other independent studies have confirmed a role for cGMP-dependent activity in retinal degeneration, the focus of the project shifted towards PKG2. This in turn has provided a basis for the a rational drug design approach for novel PKG2 inhibitors performed together with an industry partner, in the framework of the DRUGSFORD project. In addition, the establishment of a battery of test systems for PKG activity and PKG inhibitors using in vitro organotypic retinal explant cultures and later on in vivo animal models provided a solid foundation and experience for a more extensive test programme. In this way, the project work led to the formation of the larger European DRUGSFORD consortium and further funding from the European Union (HEALTH-F2-2012; www.drugsford.eu).

Publications

• Functional phenotyping of mouse models with ERG. In: Methods in Molecular Biology, Vol. 935: Retinal Degeneration: Methods and Protocols (Weber BH and Langmann T, eds), pp 69-78, New York: Humana Press, 2013
  Tanimoto N, Sothilingam V, Seeliger MW
  
• Inhibition of mitochondrial pyruvate transport by zaprinast causes massive accumulation of aspartate at the expense of glutamate in the retina. J Biol Chem. 288:36129-40, 2013
  Du, Jianhai; Cleghorn, Whitney M.; Contreras, Laura; Lindsay, Ken; Rountree, Austin M.; Chertov, Andrei O.; Turner, Sally J.; Sahaboglu, Ayse; Linton, Jonathan; Sadilek, Martin; Satrústegui, Jorgina; Sweet, Ian R.; Paquet-Durand, François & Hurley, James B.
  
• Retinitis Pigmentosa: Rapid neurodegeneration is governed by slow cell death mechanisms. Cell Death Dis, 4, e488, 2013
  Sahaboglu, A.; Paquet-Durand, O.; Dietter, J.; Dengler, K.; Bernhard-Kurz, S.; Ekström, P. Ar; Hitzmann, B.; Ueffing, M. & Paquet-Durand, F.
  
• How long does a photoreceptor cell take to die? Implications for the causative cell death mechanisms. Adv Exp Med Biol.; 801:575-81, 2014
  Paquet-Durand, F.; Sahaboglu, A.; Dietter, J.; Paquet-Durand, O.; Hitzmann, B.; Ueffing, M. & Ekström, P. A. R.
  
• Identification of a common non-apoptotic cell death mechanism in hereditary retinal degeneration. PLoS One, 9:e112142, 2014
  Arango-Gonzalez, Blanca; Trifunović, Dragana; Sahaboglu, Ayse; Kranz, Katharina; Michalakis, Stylianos; Farinelli, Pietro; Koch, Susanne; Koch, Fred; Cottet, Sandra; Janssen-Bienhold, Ulrike; Dedek, Karin; Biel, Martin; Zrenner, Eberhart; Euler, Thomas; Ekström, Per; Ueffing, Marius & Paquet-Durand, François
  
• Knockout of PARG110 confers resistance to cGMP- induced toxicity in mammalian photoreceptors. Cell Death Dis, 5:e1234, 2014
  Sahaboglu, A.; Tanimoto, N.; Bolz, S.; Garrido, M. G.; Ueffing, M.; Seeliger, M. W.; Löwenheim, H.; Ekström, P. & Paquet-Durand, F.
  
• Novel in situ activity assays for the quantitative molecular analysis of neurodegenerative processes in the retina. Curr Med Chem, 21:3478-93, 2014
  Ekstrom, P.A.R.; Ueffing, M.; Zrenner, E. & Paquet-Durand, F.
  
• Retinitis pigmentosa: impact of different Pde6a point mutations on the disease phenotype. Hum Mol Genet. 24:5486-99, 2015
  Sothilingam, Vithiyanjali; Garcia Garrido, Marina; Jiao, Kangwei; Buena-Atienza, Elena; Sahaboglu, Ayse; Trifunović, Dragana; Balendran, Sukirthini; Koepfli, Tanja; Mühlfriedel, Regine; Schön, Christian; Biel, Martin; Heckmann, Angelique; Beck, Susanne C.; Michalakis, Stylianos; Wissinger, Bernd; Seeliger, Mathias W. & Paquet-Durand, François