Endometrial carcinoma is one of the most common cancers in women. A limited number of endometrial carcinoma cell lines are available for studies of signal transduction pathways and experimental therapeutics in vitro. However, these cell lines have not been comprehensively characterized. In this study, we used genome-wide microarray-based comparative genomic hybridization (aCGH) technology to characterize five of the more commonly used endometrial cancer cell lines. We detected DNA copy–number gains in chromosomal regions 2q, 3p, 3q, 5q, 7p, 17q, and19q in all five cell lines. Other common sites of copy–number gains, which were detected in four of five cell lines, included segments of chromosomes 1, 6, 8, 9, 11, 12, and 16. In all five cell lines, we found DNA copy–number losses in regions 3p, 10p, 10q, 11q, 11p, 14q, 15q, 18p, and 21q. Other common sites of genetic aberrations included segments of chromosomes 1, 2, 4, 5, 6, 16, 20, and 22. The genes involved in the copy-number alterations included the oncogenes PIK3CA (3q26.3), K-ras (12p12.1), R-ras (19q13.3-qter), Raf-1 (3p25), EGFR (7p12), Akt1 (14q32.32), and Akt2 (19q13.1-q13.2). A pathway analysis showed that genes in the PI3K and Wnt pathways are commonly affected. Our characterization of genomic alterations in these five commonly used endometrial cancer cell lines provides valuable genomic information for research that focuses on these key oncogenic pathways in endometrial cancer.