CELLULAR UPTAKE OF A NOVEL CYTOTOXIC AGENT, CRYPTOPHYCIN-52, BY HUMANTHP-1 LEUKEMIA-CELLS AND H-125 LUNG-TUMOR CELLS

Cryptophycin (CP) is a newly developed anticancer agent isolated from the terrestrial cyanobacteria of the genus Nostoc CP is a mitotic inhi bitor, causing cells to accumulate in mitosis with the disappearance o f intracellular microtubules, In this report, we studied the interacti on and uptake of a new synthetic CP analog, CP-52, with 2 human tumor cell lines, THP-1 and H-125. In vitro colony-forming assay showed that CP-52 has antiproliferative activity against THP-1 and H-125 cell lin es with IC50 of 0.1 ng/ml and 20 mu g/ml, respectively; i.e., THP-1 ce lls are 200,000 times more sensitive to CP-52 than H-125 cells. The up take of CP-52 by the target cells was carried out using tritiated CP-5 2 (H-3-CP-52). The uptake of H-3-CP-52 by both THP-1 and H-125 cells w as rapid, reaching a maximum within 20 min. Dissociation experiments s howed that CP-52 interacts with the target cells irreversibly, presuma bly by binding to specific cellular sites with high affinity. With inc reasing doses of H-3-CP-52, the uptake was found to be saturable, reac hing a steady state as the concentrations of H-3-CP-52 were raised to about 20 mu g/ml. Under this condition, the maximal values of CP-52 up take by THP-1 and H-125 cells was estimated to be 27 and 136 ng/10(5) cells, respectively. The uptake and accumulation of H-3-CP-52 with the target cells was effectively inhibited by prior treatment with unlabe led CP-52 and, to a lesser extent, vinblastine and taxol but not adria mycin, colchicine or mitomycin. In addition, the binding of H-3-CP-52 to purified tubulin was inhibited by vinblastine but not taxol. This f inding suggested that CP-52 and taxol interact and bind to distinct re gions of tubulin molecules. Further, it suggests that, in addition to tubulin, other intracellular and/or membrane components are involved i n mediating the binding of CP-52. (C) 1998 Wiley-Liss, Inc.