The efficacy of anti-TNF-α therapies highlights the role of TNF-α in the pathogenesis of rheumatoid arthritis (RA). However, the mechanism of action of these agents is poorly understood at the molecular level. The aim of this study was to characterize the effects of anti-TNF-α treatment on the global gene expression profile in peripheral blood mononuclear cells (PBMCs) of responder RA patients. Changes in gene expression were determined using oligonucleotide microarrays (25,341 genes) in PBMCs obtained before and after 12 wk of treatment with either etanercept or adalimumab from responder RA patients. Two hundred fifty-one genes displayed significant changes (false discovery rate < 0.1%) in expression level (178 upregulations with mean fold change = 1.5 and 73 downregulations with mean fold change = -1.50) after 12 wk of treatment. Importantly, the expression of several genes, including those coding for the calcium binding proteins S100A12 and A8, CD14 antigen, Selectin P, or ribosomal protein L39, reported to be upregulated in RA patients, were found to be decreased after anti-TNF-α treatment. Globally, inflammation, immune response, apoptosis, protein synthesis, and mitochondrial oxido-reduction were the most affected pathways in response to anti-TNF-α treatment. The obtained gene expression signature in PBMCs provides new information to better understand the mechanisms of action of anti-TNF-α treatment in RA patients.