In vitro activity: PF 1069 (Compound 55) is a PARP inhibitor, with IC50s of 8 and 0.3 μM for PARP-1 and PARP-2, respectively. UPF 1069 (1 μM) reduces the residual PARP activity by approximately 80% of PARP-1-deficient fibroblasts, but only slightly inhibits the enzymic activity in wild-type fibroblasts. UPF 1069 (0.1-1 μM) markedly enhances CA1 hippocampal damage. UPF 1069 (10 μM) also exacerbates oxygen-glucose deprivation (OGD) damage in organotypic hippocampal slices. However, UPF 1069 alleviates the damage cuased by OGD in mixed cortical cell cultures, shows a potent neuroprotective activity both at a concentration (1 μM) selectively acting on PARP-2 and at a concentration (10 μM) inhibiting both PARP-1 and PARP-2 activities.

Kinase Assay: PARP activity is evaluated by utilizing commercially available recombinant bovine PARP-1 and mouse PARP-2. Briefly, the enzymatic reaction is carried out in 100 μL of 50 mM Tris-HCl (pH 8.0) containing 5 mM MgCl2, 2 mM dithiothreitol, 10 μg sonicated calf thymus DNA, 0.2 μCi [adenine-2,8-3H]NAD and recombinant enzyme PARP-1 or PARP-2 (0.03 U per sample). Different concentrations of the putative inhibitors are added, and the mixture is incubated for 1 h at 37°C. The reaction is terminated by adding 1 mL of 10% trichloroacetic acid (w/v) and centrifuged. Pellets are then washed twice with 1 mL of H2O and resuspended in 1 mL of 0.1 M NaOH. The radioactivity incorporated from [adenine-2,8-3H]NAD into proteins is evaluated by liquid scintillation spectrometry.

Br J Pharmacol. 2009 Jul;157(5):854-62.