title: Acefylline activates filaggrin deimination by peptidylarginine deiminases in the upper epidermis.
abstract: BACKGROUND
  Peptidylarginine deiminases (PADs) catalyze deimination (or citrullination), a calcium-dependent post-translational modification involved in several physiological processes and human diseases, such as rheumatoid arthritis and cancer. Deimination of filaggrin (FLG) by PAD1 and PAD3 during the last steps of keratinocyte differentiation is a crucial event for the epidermis function and homeostasis. This allows the complete degradation of FLG, leading to the production of free amino acids and their derivatives that are essential for epidermal photoprotection and moisturizing of the stratum corneum.
  
  
  OBJECTIVE
  To increase the flux of this catabolic pathway, we searched for activators of PADs.
  
  
  METHODS
  A large chemical library was screened first in silico and then by using an automated assay based on an indirect colorimetric measurement of recombinant human PAD activity. Potential activators were then confirmed using a recombinant human FLG as a substrate, and secondly after topical application at the surface of three-dimensional reconstructed human epidermis.
  
  
  RESULTS
  The data obtained after the library screening pointed to xanthine derivatives as potential PAD activators. Among seven xanthine derivatives tested at 50-300μM, caffeine, theobromine and acefylline proved to be the most potent enhancers of in vitro deimination of FLG by PAD1 and PAD3. After topical application of a gel formulation containing 3% acefylline at the surface of reconstructed epidermis, immunoblotting analysis showed an increase in the total amount of deiminated proteins, and confocal microscopy showed an enhanced deimination in the stratum corneum. This demonstrated the activation of PADs in living cells.
  
  
  CONCLUSIONS
  As a PAD activator, acefylline will be useful to study the role of deimination and could be proposed to increase or correct the hydration of the cornified layers of the epidermis.
authors.affiliation: Paul Sabatier University (UPS), Toulouse, France; Centre National de la Recherche Scientifique (CNRS), UMR5165, Toulouse, France; Institut National de la Santé Et de la Recherche Médicale (INSERM), U1056, Toulouse, France.
authors.forename: Marie-Claire
authors.initials: MC
authors.lastname: Méchin
authors.affiliation: Paul Sabatier University (UPS), Toulouse, France; Centre National de la Recherche Scientifique (CNRS), UMR5165, Toulouse, France; Institut National de la Santé Et de la Recherche Médicale (INSERM), U1056, Toulouse, France.
authors.forename: Laura
authors.initials: L
authors.lastname: Cau
authors.affiliation: Centre de Recherche & Développement Pierre Fabre Dermo-Cosmétique et Médicament, Toulouse, France.
authors.forename: Marie-Florence
authors.initials: MF
authors.lastname: Galliano
authors.affiliation: Centre de Recherche & Développement Pierre Fabre Dermo-Cosmétique et Médicament, Toulouse, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Daunes-Marion
authors.affiliation: Centre de Recherche & Développement Pierre Fabre Dermo-Cosmétique et Médicament, Toulouse, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Poigny
authors.affiliation: Centre de Recherche & Développement Pierre Fabre Dermo-Cosmétique et Médicament, Toulouse, France.
authors.forename: Jean-Louis
authors.initials: JL
authors.lastname: Vidaluc
authors.affiliation: Centre de Recherche & Développement Pierre Fabre Dermo-Cosmétique et Médicament, Toulouse, France.
authors.forename: Sandrine
authors.initials: S
authors.lastname: Bessou-Touya
authors.affiliation: Ibaraki University, Ibaraki, Japan.
authors.forename: Hidenari
authors.initials: H
authors.lastname: Takahara
authors.affiliation: Paul Sabatier University (UPS), Toulouse, France; Centre National de la Recherche Scientifique (CNRS), UMR5165, Toulouse, France; Institut National de la Santé Et de la Recherche Médicale (INSERM), U1056, Toulouse, France.
authors.forename: Guy
authors.initials: G
authors.lastname: Serre
authors.affiliation: Centre de Recherche & Développement Pierre Fabre Dermo-Cosmétique et Médicament, Toulouse, France.
authors.forename: Hélène
authors.initials: H
authors.lastname: Duplan
authors.affiliation: Paul Sabatier University (UPS), Toulouse, France; Centre National de la Recherche Scientifique (CNRS), UMR5165, Toulouse, France; Institut National de la Santé Et de la Recherche Médicale (INSERM), U1056, Toulouse, France. Electronic address: michel.simon@udear.cnrs.fr.
authors.forename: Michel
authors.initials: M
authors.lastname: Simon
mesh_terms: D000279:Administration, Cutaneous; D002478:Cells, Cultured; D003198:Computer Simulation; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology*; D000078404:Epidermal Cells; D004817:Epidermis / Q000187:drug effects* / Q000201:enzymology; D000091344:Filaggrin Proteins; D006801:Humans; D006867:Hydrolases / Q000378:metabolism*; D007381:Intermediate Filament Proteins / Q000737:chemistry / Q000378:metabolism*; D015603:Keratinocytes / Q000187:drug effects* / Q000201:enzymology; D008958:Models, Molecular; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D000076342:Protein-Arginine Deiminases; D054852:Small Molecule Libraries; D013329:Structure-Activity Relationship; D013806:Theophylline / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology
chemical_list: D020536:Enzyme Activators; C000717249:FLG protein, human; D000091344:Filaggrin Proteins; D007381:Intermediate Filament Proteins; D054852:Small Molecule Libraries; D013806:Theophylline; D006867:Hydrolases; D000076342:Protein-Arginine Deiminases; C019140:acefylline
keywords: Epidermal barrier; Filaggrin; Post-translational modification; Skin; Xanthine