New pyrazoline derivatives were synthesized and evaluated for their cytotoxic effects on AsPC-1 human pancreatic adenocarcinoma, U87 and U251 human glioblastoma cell lines. 1-[((5-(4-Methylphenyl)-1,3,4-oxadiazol-2-yl)thio)acetyl]-3-(2-thienyl)-5-(4-chlorophenyl)-2-pyrazoline (11) was found to be the most effective anticancer agent against AsPC-1 and U251 cell lines, with IC₅₀ values of 16.8 µM and 11.9 µM, respectively. Tumor selectivity of compound 11 was clearly seen between Jurkat human leukemic T-cell line and human peripheral blood mononuclear cells (PBMC). Due to its promising anticancer activity, compound 11 was chosen for apoptosis/necrosis evaluation and DNA-cleavage analysis in U251 cells. Compound 11-treated U251 cells exhibited apoptotic phenotype at low concentration (1.5 µM). DNA-cleaving efficiency of this ligand was more significant than cisplatin and was clearly enhanced by Fe(II)-H₂O₂-ascorbic acid systems. This result pointed out the relationship between the DNA cleavage and the cell death.