Most monogenic forms of inherited retinopathies are associated with genes expressed in photoreceptors (PR) or retinal pigment epithelium (RPE) where they encode proteins that are critical for PR structure, function and survival. Specific cellular processes and biochemical pathways implicated in retinal dystrophies include: PR development, morphogenesis, phototransduction, visual cycle, cellular metabolism, protein folding, among others. The vision and survival of rod and cone require continuous renewal of the visual chromophore, the 11-cis retinal, which forms together with the opsin proteins, the rhodopsin and cone visual pigments.
11-cis retinal is produced from vitamin A, an all-trans retinol, which is supplied with provitamin carotene from diet. In the outer segment (OS) discs of PR, light cause photoisomerization of 11-cis retinal to all-trans retinal, this is then reduced to all-trans retinol. Renewal of 11-cis retinal is provided by the visual cycle, an enzymatic pathway which involves the adjacent RPE and Müller cells. Stargardt pathophysiology involves rapid accumulation of toxic bisretinoid by-products such as A2E in the RPE.
1- Lipid metabolism is of major importance for retina integrity and its deregulation can lead to pigmentary retinopathies. Lipid disorders have been implicated in macular degeneration such as the age-related macular degeneration (AMD) and Stargardt disease. We have identified FATP1 as a gene that regulates the (visual) cycle of retinoids in the retina. As such it could act as a "modifier" of retinal disease fate.
2- Carbonyl and oxidative stress (COS) play an important role in Stargardt's disease as well as in neurodegenerative diseases of aging such as age-related macular degeneration (AMD). We are developing lipophenolic chemicals that are effective as anti-COS: first, to slow the formation of bisretinoid derivatives, such as A2E, by countering all-trans retinal toxicity in photoreceptor cells; second, to eliminate oxidative stressors by preventing A2E oxidation-related damage in RPE. Based on our expertise in retinal and visual cycle biochemistry, we will work on innovative projects based on obtaining RPE cells or retinal organoids from iPSCs of Stargardt patients with varying degrees of severity for phenotype analysis and pharmacological therapy.
We also hope to better understand the action of these lipophenolic compounds by studying differential transcriptional responses to the retina and/or the compounds in the retina in vitro (cell culture) and in vivo (Staragardt animal model).
Our team has identified several mutations in the visual cycle genes responsible for retinal dystrophies (Marlhens et al., Nat Genet, 1997; Sénéchal et al., Am J Ophthalmol. 2006; Humbert et al., Invest Ophthalmol Vis Sci. 2006; Ksantini et al., Ophthalmic Genet. 2010). We have now developed biochemical analysis and animal models to study molecular mechanisms and pathophysiology of the visual cycle. We’ve started up the strategies by searching for retinoid isomerase (RPE65)-interacting proteins. A two-hybrid screening with the bait RPE65 has revealed several classes of proteins that can interact with. Among them, the metabolic protein FATP1 is an acyl-CoA-synthetase which mobilizes long chain fatty acid. We demonstrated the co-expression of FATP1 and RPE65 in cell cultures results in significant inhibition of 11-cis-retinol formation (Guignard et al., JBC, 2010). This was the first demonstration of a RPE65 interacting protein which inhibits the isomerisation. We then studied the phenotype of Fatp1-deficient mice provided by Andreas Stahl. Interestingly, these mice showed a slight but significant decrease of the visual function and several retinal alterations with age (Chekroud et al., PLoS One. 2012, Cia et al., J Cell Mol Med 2016). Transgenic mice overexpressing the human FATP1 gene in RPE identify a previously unknown role of FATP1 as an important regulator of retinoid metabolism and photoreceptor homeostasis (Cubizolle A., et al., PLoS ONE 2017, Van Der Brink DM. and Cubizolle A., et al., PLoS Genet., 2018)
Using pharmaceutical chemistry, we have been able to synthesize molecules that combine alkylated polyphenols and long-chain unsaturated fatty acids, which we call lipophenols. These compounds have the dual anti-COS function of acting on both all-trans retinal and oxidative stress (A2E). We evaluated the protective capacity against COS in rodent primary cells and a human RPE line (Cubizolle et al., J Cell Mol Med. 2017; Cubizolle et al., J Cell Mol Med. 2020) using dozens of molecules with different polypenolic backbones, different fatty acids in phenolic or carbon positions, and alkylation and deuteration substitutions (Moine et al. Antioxidants (Basel) 2018; Rosell et al., Antioxidants (Basel), 2019). We have identified several LEADs that have been evaluated preclinically in an Abca4-/- mouse model of STGD1 (Taveau and Cubizolle et al., Exp Mol Med 2020).
Biochemistry and molecular biology, yeast two-hydrid screening, recombinant protein interaction, cell cultures, cell death and cell survival measurements, HLPC analysis of retinoids, analytical chemistry to probe the visual cycle, physiology of the visual function in mice.
Taveau N. and Cubizolle A., et al, Exp Mol Med. 2020
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- David Cia (UMR Inserm 1107, Laboratoire de Biophysique Neurosensorielle, Facultés de Médecine et de Pharmacie, Clermont-Ferrand).
- Céline Crauste, Joseph Vercauteren et Thierry Durand, Institute of Biomolecules Max Mousseron (IBMM), UMR5247-CNRS-UM1-UM2-ENSCM, Faculty of Pharmacy, Montpellier.
- Francisco Veas, Institut de Recherche pour le Développement (IRD - France), Molecular Comparative Immuno-Physiopathology Health, Research Professor
- Peter Humphries, Ocular Genetics Unit, Department of Genetics, Trinity College Dublin, Dublin, Ireland.