In this study, we aimed to investigate the potential relationship between gossypol-induced cytotoxicity of human promyelocytic leukemia cell line (HL-60) leukemic cells and intracellular serine/threonine protein phosphatase (PP) dynamics and human telomerase reverse transcriptase (hTERT) activity. Gossypol was found to be cytotoxic in HL-60 cells with the IC50 dose of 4.5 mu M. The combination of gossypol and okadaic acid in IC50 doses revealed the increased cytotoxicity in a time-dependent manner. Treatment of cells with gossypol has shown significant decrease in PP2A activity. The expression of the PP2A catalytic subunit was downregulated in gossypol-treated cells with 24 hours' intervals. hTERT mRNA levels were gradually decreased. In conclusion, during gossypol-induced cytotoxicity, intracellular activity and expression of PP2A was decreased as well as the activity of hTERT. The variation of hTERT activity in gossypol-treated HL-60 cells may be the potential reason for the phosphatase interaction during the gossypol treatment of leukemic cells resulting in cellular cytotoxicity.