title: PA401, a novel CXCL8-based biologic therapeutic with increased glycosaminoglycan binding, reduces bronchoalveolar lavage neutrophils and systemic inflammatory markers in a murine model of LPS-induced lung inflammation.
abstract: BACKGROUND
  Neutrophils play a fundamental role in a number of chronic lung diseases. Among the mediators of their recruitment to the lung, CXCL8 (IL-8) is considered to be one of the major players. CXCL8 exerts its chemotactic activity by binding to its GPCR receptors (CXCR1/R2) located on neutrophils, as well as through interactions with glycosaminoglycans (GAGs) on cell surfaces including those of the microvascular endothelium. Binding to GAG co-receptors is required to generate a solid-phase haptotactic gradient and to present IL-8/CXCL8 in a proper conformation to its receptors on circulating neutrophils.
  
  
  METHODS
  We have engineered increased GAG-binding affinity into human CXCL8, thereby obtaining a competitive inhibitor that displaces wild-type IL-8/CXCL8 from GAGs. By additionally knocking-out the GPCR binding domain of the chemokine, we generated a dominant negative protein (dnCXCL8; PA401) with potent anti-inflammatory characteristics proven in vivo in a murine model of LPS-induced lung inflammation (Adage et al., 2015). Here we have further investigated PA401 activity in this pulmonary model by evaluating plasma changes induced by LPS on white blood cells (WBC) and a broad range of inflammatory markers, especially chemokines, by addressing immediate effects of PA401 on these parameters in healthy and LPS exposed mice.
  
  
  RESULTS
  Aerosolized LPS induced a significant increase in bronchoalveolar lavage (BAL) neutrophils after 3 and 7h, as well as an increase in total WBC and changes in 21 of the 59 measured plasma markers, mostly belonging to the chemokine family. PA401 treatment in saline exposed mice didn't induce major changes in any of the measured parameters. When administered to LPS aerosolized mice, PA401 caused a significant normalization of KC/mCXCL1 and other inflammatory markers, as well as of blood WBC count. In addition, BAL neutrophils were significantly reduced, confirming the previously observed lung anti-inflammatory activity of PA401 in this experiment.
  
  
  CONCLUSIONS
  PA401 is a new promising biologic therapeutic with a novel and unique mechanism of action for interfering with neutrophilic lung inflammation, that also normalizes plasma inflammatory markers.
authors: ProtAffin Biotechnologie AG, Graz, Austria.
authors: Tiziana
authors: T
authors: Adage
authors: Accelera S.r.l., Nerviano (MI), Italy; Janssen Pharmaceutica NV, Beerse, Belgium(1).
authors: Francesca
authors: F
authors: Del Bene
authors: Accelera S.r.l., Nerviano (MI), Italy.
authors: Francesco
authors: F
authors: Fiorentini
authors: ProtAffin Biotechnologie AG, Graz, Austria; Merus B.V., Utrecht, The Netherlands(1).
authors: Robert P
authors: RP
authors: Doornbos
authors: ProtAffin Biotechnologie AG, Graz, Austria; Institute of Pharmaceutical Sciences, University of Graz, Graz, Austria.
authors: Christina
authors: C
authors: Zankl
authors: ProtAffin Biotechnologie AG, Graz, Austria.
authors: Michael R
authors: MR
authors: Bartley
authors: ProtAffin Biotechnologie AG, Graz, Austria; Institute of Pharmaceutical Sciences, University of Graz, Graz, Austria. Electronic address: andreas.kungl@uni-graz.at.
authors: Andreas J
authors: AJ
authors: Kungl
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism*; D001992:Bronchoalveolar Lavage Fluid; D004195:Disease Models, Animal; D006025:Glycosaminoglycans / Q000378:metabolism*; D016209:Interleukin-8 / Q000378:metabolism* / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D011014:Pneumonia / Q000139:chemically induced*; D011994:Recombinant Proteins / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D015415:Biomarkers; D006025:Glycosaminoglycans; D016209:Interleukin-8; D008070:Lipopolysaccharides; C000610402:PA401 protein; D011994:Recombinant Proteins
keywords: Blood inflammatory markers; Engineered CXCL8; Glycosaminoglycans; Lung inflammation