The Development of N-α-(2-Carboxyl)benzoyl-N5-(2-fluoro-1-iminoethyl)-L-ornithine Amide (o-F-amidine) and N-α-(2-Carboxyl)benzoyl-N5-(2-chloro-1-iminoethyl)-L-ornithine Amide (o-Cl-amidine) As Second Generation Protein Arginine Deiminase (PAD) Inhibitors

Protein arginine deiminase (PAD) activity is upregulated in a number of human diseases, including rheumatoid arthritis, ulcerative colitis, and cancer. These enzymes, there are five in humans (PADs 1-4 and 6), regulate gene transcription, cellular differentiation, and the innate immune response. Building on our successful generation of F-and Cl-amidine, which irreversibly inhibit all of the PADs, a structure activity relationship was performed to develop second generation compounds with improved potency and selectivity. Incorporation of a carboxylate ortho to the backbone amide resulted in the identification of N-alpha-(2-carboxyl)benzoyl-N-5-(2-fluoro-1-iminoethyl)-L-ornithine amide (o-F-amidine) and N-alpha-(2-carboxyl)benzoyl-N-5-(2-chloro-1-iminoethyl)-L-ornithine amide (o-Cl-amidine), as PAD inactivators with improved potency (up to 65-fold) and selectivity (up to 25-fold). Relative to F- and Cl-amidine, the compounds also show enhanced potency in cellulo. As such, these compounds will be versatile chemical probes of PAD function.