ARTHRITIS & RHEUMATISM Vol. 56, No. 9, September 2007, pp 3138–3148 DOI 10.1002/art.22842 © 2007, American College of Rheumatology Pattern of Interleukin-1 Secretion in Response to Lipopolysaccharide and ATP Before and After Interleukin-1 Blockade in Patients With CIAS1 Mutations Marco Gattorno, 1 Sara Tassi, 2 Sonia Carta, 2 Laura Delfino, 2 Francesca Ferlito, 1 Maria Antonietta Pelagatti, 1 Andrea D’Osualdo, 3 Antonella Buoncompagni, 1 Maria Giannina Alpigiani, 4 Maria Alessio, 5 Alberto Martini, 1 and Anna Rubartelli 2 Objective. To examine the synthesis, processing, and secretion of interleukin-1 (IL-1), as well as the clinical and biologic effects of IL-1 blockade, in patients with chronic infantile neurologic, cutaneous, articular (CINCA) syndrome and Muckle-Wells syndrome (MWS), in an effort to understand the molecular mech- anisms linking mutations of the CIAS1 gene and IL-1 hypersecretion, and the underlying response to IL-1 receptor antagonist (IL-1Ra). Methods. Six patients with CINCA syndrome or MWS were treated with IL-1Ra and followed up longi- tudinally. Monocytes obtained from the patients and from 24 healthy donors were activated with lipopolysac- charide (LPS) for 3 hours, and intracellular and se- creted IL-1 levels were determined by Western blotting and enzyme-linked immunosorbent assay before and after exposure to exogenous ATP. Results. LPS-induced IL-1 secretion was mark- edly increased in monocytes from patients with CIAS1 mutations. However, unlike in healthy subjects, secre- tion of IL-1 was not induced by exogenous ATP. Treatment with IL-1Ra resulted in a dramatic clinical improvement, which was paralleled by an early and strong down-regulation of LPS-induced IL-1 secretion by the patients’ cells in vitro. Conclusion. Our results showed that the require- ments of ATP stimulation for IL-1 release observed in healthy individuals are bypassed in patients bearing CIAS1 mutations. This indicates that cryopyrin is the direct target of ATP and that the mutations release the protein from the requirement of ATP for activation. In addition, the dramatic amelioration induced by IL-1Ra treatment is at least partly due to the strong decrease in IL-1 secretion that follows the first injections of the antagonist. These findings may have implications for other chronic inflammatory conditions characterized by increased IL-1. Interleukin-1 (IL-1) plays a pivotal role in the pathogenesis of autoinflammatory diseases and repre- sents a potential target of therapeutic intervention in both monogenic and multifactorial inflammatory dis- eases (1). Unlike most cytokines, IL-1 lacks a secretory signal peptide and is externalized by monocytic cells through a nonclassic pathway, arranged in 2 steps (2,3). First, Toll-like receptor ligands, such as lipopolysaccha- ride (LPS), induce gene expression and synthesis of the inactive IL-1 precursor (proIL-1). Monocytes stimu- lated with LPS alone release only 20% of the IL-1 over 24–48 hours (4). A second stimulus, such as exogenous ATP, strongly enhances the proteolytic mat- uration and secretion of IL-1 (5,6). ATP-triggered Supported in part by grants from the Associazione Italiana per la Ricerca sul Cancro, the Ministero della Salute (Ricerca Cor- rente e Finalizzata), and the Comitato Interministeriale per la Pro- grammazione Economica (grant 02/07/2004, CBA project). 1 Marco Gattorno, MD, Francesca Ferlito, PhD, Maria An- tonietta Pelagatti, MD, Antonella Buoncompagni, MD, Alberto Mar- tini, MD: Second Division of Pediatrics, IRCCS, Istituto G. Gaslini, and University of Genoa, Genoa, Italy; 2 Sara Tassi, PhD, Sonia Carta, PhD, Laura Delfino, Technician, Anna Rubartelli, MD: Istituto Na- zionale per la Ricerca sul Cancro, Genoa, Italy; 3 Andrea D’Osualdo, PhD: Laboratory of Molecular Gentics, IRCCS, Istituto G. Gaslini, Genoa, Italy; 4 Maria Giannina Alpigiani, MD: University of Genoa, Genoa, Italy; 5 Maria Alessio, MD: Federico II University, Naples, Italy. Drs. Gattorno and Tassi contributed equally to this work. Address correspondence and reprint requests to Marco Gat- torno, MD, IRCCS, Istituto G. Gaslini, Pediatria II, Largo G. Gaslini 5, 16147 Genoa, Italy. E-mail: marcogattorno@ospedale-gaslini.ge.it, or to Anna Rubartelli, MD, Istituto Nazionale per la Ricerca sul Cancro, Largo Rosanna Benzi 10, 16132 Genoa, Italy. E-mail: anna.rubartelli@istge.it. Submitted for publication September 15, 2006; accepted in revised form May 18, 2007. 3138