A BAL was then performed using 3 individual 0.4?ml volumes of PBS, which were combined. Blood pharmacodynamic bioassay One hundred l of whole blood from each animal was added to 400?l of RPMI media supplemented with penicillin and streptomycin (All Invitrogen). and have been shown to be efficacious in inflammatory arthritis models, did not lead to total blockade of the alveolar macrophages response to GM-CSF. This suggests a significant therapeutic window is possible with GM-CSF axis inhibition. KEYWORDS: Antibody exposure, GM-CSF, GM-CSF Receptor, lung partitioning, pharmacodynamics, pulmonary alveolar proteinosis, rheumatoid arthritis Abbreviations BALbronchoalveolar lavageGM-CSFgranulocyte macrophage colony-stimulating factorGM-CSFRgranulocyte macrophage colony-stimulating factor receptor subuniti.nintranasalLLOQLower Limit of QuantitationNDnot detectablePAPpulmonary Nepafenac alveolar proteinosisPDpharmacodynamicsPKpharmacokineticsRARheumatoid arthritis Introduction Rheumatoid arthritis Nepafenac (RA), a chronic systemic autoimmune disease characterized by inflammation of synovial joints, affects approximately 1% of the population. The debilitating, painful joint swelling and damage can be refractory to, or incompletely modified by, current therapies, including both small molecule and biologic disease modifying anti-rheumatic drugs.1 Consequently, there is still need for novel treatments that are more efficacious. Antibody blockade of granulocyte macrophage colony-stimulating factor (GM-CSF) and knock-out mice have revealed that this cytokine has a pivotal role in inflammation,2 and especially in models of autoimmune inflammation such as collagen-induced arthritis3,4 and experimental autoimmune encephalomyelitis.5 GM-CSF mediates its effects by specifically binding the GM-CSF receptor subunit (GM-CSFR), and then recruiting the signaling common -chain. Antibody blockade of GM-CSFR has a similarly anti-inflammatory effect on arthritis models. 6 GM-CSFR is usually widely expressed on myeloid cells and granulocytes, which are strongly associated with the inflammation in RA.7 GM-CSF is raised in synovial fluid from patients with RA.8,9 These data make the GM-CSF/GM-CSFR axis a stylish target for therapeutic intervention in RA and other autoimmune diseases. Mavrilimumab, an antagonistic antibody targeting GM-CSFR, recently completed a Phase 2b trial in patients with RA who have had an inadequate response to methotrexate.10 Highly significant improvements in the signs and symptoms of arthritis were observed in this study, which replicated the results of an earlier Phase 2 study.11 In addition to its role in inflammation, studies Nepafenac in knock out mice have demonstrated that GM-CSF has a limited role in hematopoiesis, with steady-state effects being limited Nepafenac to some tissue-resident dendritic cell populations.12-15 This was surprising because GM-CSF was originally identified as a molecule that can expand myeloid progenitors.16 Further analysis did demonstrate a requirement for GM-CSF in alveolar macrophage function.12,13 The alveolar macrophages from mice deficient in GM-CSF or the common chain are less able to catabolise surfactant lipids, which leads to the formation of foamy alveolar macrophage and the accumulation of lipoproteinaceous material in the lung.17,18 The lung Cd24a phenotype of these mice and its similarity to a very rare lung condition, known then as idiopathic pulmonary alveolar proteinosis (PAP), led to the discovery that defects in this pathway underlie this disease.19,20 Subsequently PAP has been shown to be strongly associated with anti-GM-CSF autoantibodies,21 or with mutations in either of the GM-CSF receptor subunits.22-25 Additionally, transfer of auto-antibodies from patients with anti-GM-CSF-associated PAP to non-human primates led to the formation of foamy macrophages.26 These results indicate that GM-CSF plays an important role in the terminal differentiation of alveolar macrophages in mice and humans. Dosing of very high levels ( 30?mg/kg/week for 11?weeks) of mavrilimumab in preclinical non-human primate studies was associated with the formation of a very small number of foamy macrophages within the.