Poly(ADP-ribose) polymerase (
PARP) inhibitors enhance
DNA topoisomerase I (
topo I)
poison-induced cytotoxicity and antitumor activity in vitro and in vivo, but the mechanism has not been defined. We investigated the role of PARP-1 in the response to
topo I
poisons using PARP-1-/- and PARP-1+/+ mouse embryonic fibroblasts and the potent PARP-1 inhibitor,
AG14361 (Ki < 5 nmol/L). PARP-1-/- mouse embryonic fibroblasts were 3-fold more sensitive to
topotecan than PARP-1+/+ mouse embryonic fibroblasts (GI50, 21 and 65 nmol/L, respectively).
AG14361 caused a >3-fold sensitization of PARP-1+/+ cells to
topotecan compared with a <1.4-fold sensitization in PARP-1-/- cells. In human
leukemia K562 cells,
AG14361 caused a 2-fold sensitization to
camptothecin-induced cytotoxicity.
AG14361 did not affect the cellular activity of
topo I as determined by measurement of cleavable complexes and
topo I relaxation activity, showing that sensitization was not due to
topo I activation. In contrast, repair of
DNA following
camptothecin removal, normally very rapid, was significantly retarded by
AG14361, resulting in a 62% inhibition of repair 10 minutes after
camptothecin removal. This led to a 20% increase in the net accumulation of
camptothecin-induced
DNA strand break levels in cells coexposed to
AG14361 for 16 hours. We investigated the DNA repair mechanism involved using a panel of
DNA repair-deficient Chinese hamster ovary cells.
AG14361 significantly potentiated
camptothecin-mediated cytotoxicity in all cells, except the base excision repair-deficient EM9 cells. Therefore, the most likely mechanism for the potentiation of
topo I
poison-mediated cytotoxicity by
AG14361 is via PARP-1-dependent base excision repair.