wrn protects against topo i but not topo ii inhibitors by preventing dna break formation the werner syndrome helicase3exonuclease wrn is a major component of the dna repair and replication machinery to analyze whether wrn is involved in the repair of topoisomeraseinduced dna damage we utilized u2os cells in which wrn is stably downregulated wrnkd and the corresponding wildtype cells wrnwt we show that cells not expressing wrn are hypersensitive to the toxic effect of the topoisomerase i inhibitor topotecan but not to the topoisomerase ii inhibitor etoposide this was shown by mass survival assays colony formation and induction of apoptosis upon topotecan treatment wrn deficient cells showed enhanced dna replication inhibition and sphase arrest whereas after treatment with etoposide they showed the same cell cycle response as the wildtype a considerable difference between wrn and wildtype cells was observed for dna single and doublestrand break formation in response to topotecan topotecan induced dna singlestrand breaks 6h after treatment in both wrnwt and wrnkd cells these breaks were repaired at similar kinetics however in wrnkd but not wrnwt cells they were converted into dna doublestrand breaks dsbs at high frequency as shown by neutral comet assay and phosphorylation of h2ax our data provide evidence that wrn is involved in the repair of topoisomerase i but not topoisomerase iiinduced dna damage most likely via preventing the conversion of dna singlestrand breaks into dsbs during the resolution of stalled replication forks at topo idna complexes we suggest that the wrn status of tumor cells impacts anticancer therapy with topoisomerase i but not topoisomerase ii inhibitors