NADPH oxidase isoform 2 is an enzyme complex, which generates reactive oxygen species (ROS) and contributes to oxidative stress. The p47phox-p22phox interaction is critical for activation of the catalytical NOX2 domain, and p47phox is a potential target for therapeutic intervention. By screening 2,500 fragments using fluorescence polarization (FP) and a thermal shift assay (TSA) and validation by surface plasmon resonance (SPR) we found eight hits towards the tandem SH3 domain of p47phox (p47phoxSH3A-B) with KD values of 400-600 μM. Structural studies revealed that fragments 1 and 2 bound two separate binding sites in the elongated conformation of p47phoxSH3A-B and these competed with p22phox for binding to p47phoxSH3A-B. Chemical optimization led to a dimeric compound with the ability to potently inhibit the p47phoxSH3A-B-p22phox interaction (Ki of 20 μM). Thereby, we reveal a new way of targeting p47phox and present the first report of druglike molecules able to bind p47phox and inhibit its interaction with p22phox.