Abstract

Cells deficient in poly(ADP-ribose) polymerase (PARP-1) or DNA-dependent protein kinase (DNA-PK) are highly radiosensitive. Potent, specific inhibitors of PARP-1 (AG14361) and DNA-PK (NU7026) developed in collaboration with KuDOS Pharmaceuticals (UK) and Agouron/Pfizer GRD (CA), respectively, are radiosensitizers when used alone and in combination [1]. To fully exploit the potential of PARP-1 and DNA-PK inhibitors for cancer therapy, a mechanistic understanding of their interactions is essential. Reciprocal regulation by the two enzymes, and the effects of AG14361 and NU7026, were investigated using both purified enzyme assays and a permeabilised cell system. A 30 bp double stranded oligonucleotide was used to stimulate the activity of the enzymes. PARP-1 activity was not affected by the presence of DNA-PK, nor was DNA-PK activity affected by PARP-1 provided the substrates ATP and NAD were present. However, in the absence of ATP or the presence of NU7026, DNA-PK inhibited PARP-1 activity by 25% and 50% respectively. In the absence of NAD or the presence of AG14361, PARP-1 inhibited DNA-PK activity by 75%.Very similar results were obtained using the inhibitors in permeabilised cells either proficient or deficient in PARP-1 or DNA-PK. Ionizing radiation-induced DNA double strand break (DSB) repair in these cell lines was inhibited by both inhibitors and reduced DSB repair was also seen in the deficient cell lines. These data suggest that both PARP-1 and DNA-PK bind to DNA ends and co-operate in DSB repair. References: 1.Veuger SJ, Curtin NJ, Golding BT et al: Radiosensitisation and modulation of DNA repair by novel inhibitors of PARP and DNA-PK in vitro. Proc Am Assoc Cancer Res, 2002; 43: 4142

Keywords: PARP-1, DNA-PK, DNA Repair