Human topoisomerase I has been suggested to be implicated in the maintenance of genomic stability via its ability to regulate genome topology during transcription and replication. In the present study, we demonstrate by whole-genome array comparative genomic hybridization (aCGH) and fluorescence in situ hybridisation (FISH) analysis that topoisomerase I deficiency results in chromosome 5p gain in the cervical cancer cell line, HeLa-CCL2. In contrast, chromosome 5p copy number remained unaffected by topoisomerase I down-regulation in the non-cancer cell line HEK293T, as demonstrated by FISH analysis. Chromosome 5p gain is the most frequent genetic alteration in invasive cervical cancer, which leads to overexpression of genes involved in proliferation and occurs primarily at late stages in cancer development. The amplification of this region upon topoisomerase I down-regulation specifically in HeLa-CCL2 cells may indicate an important role of topoisomerase I in preventing malignant progression of precancerous lesions in the cervix.