Susceptibility of the HT-29 human colon adenocarcinoma cell line and human myeloid leukemia cell line U937 to hypericin-mediated photocytotoxicity was investigated and compared in this study. Cellular parameters as viability, cell number, metabolic activity and total protein amount were monitored in screening experiments with subsequent cell-cycle analysis and apoptosis detection to determine the cellular response of the different tumor types to various concentrations of photoactivated hypericin. The results show concentration dependence of the photosensitizer's cytotoxicity on the studied cell lines, with higher sensitivity of U937 cells. Whereas the two extreme hypericin concentrations (1 × 10⁻⁹ M and 1 × 10⁻⁶ M) resulted in similar changes in all tested cellular parameters on the two studied cell lines, 1 × 10⁻⁸ M and 1 × 10⁻⁷ M hypericin treatment resulted in different responses of the cell lines in all monitored parameters except for viability. Although leukemic cells proved sensitive to both 1 × 10⁻⁸ M and 1 × 10⁻⁷ M hypericin, significant changes on HT-29 cells were detected only after the 1 × 10⁻⁷ M hypericin concentration. Cell-cycle arrest was related to simultaneously occurring apoptosis in colon cancer. Remarkable is the difference in cell-cycle profile where G2/M arrest in colon cancer cells versus accumulation of leukemic cells in the S phase appears. This suggests that hypericin treatment affecting the cell-cycle machinery of different cancer cells is not universal in effect.