Dolastatin 11, a Marine Depsipeptide, Arrests Cells at Cytokinesis and Induces Hyperpolymerization of Purified Actin

Abstract

The successful synthesis of dolastatin 11, a depsipeptide originally isolated from the mollusk Dolabella auricularia, permitted us to study its effects on cells. The compound arrested cells at cytokinesis by causing a rapid and massive rearrangement of the cellular actin filament network. In a dose-and time-dependent manner, F-actin was rearranged into aggregates, and subsequently the cells displayed dramatic cytoplasmic retraction. The effects of dolastatin 11 were most similar to those of the sponge-derived depsipeptide jasplakinolide, but dolastatin 11 was about 3-fold more cytotoxic than jasplakinolide in the cells studied. Like jasplakinolide, dolastatin 11 induced the hyperassembly of purified actin into filaments of apparently normal morphology. Dolastatin 11 was qualitatively more active than jasplakinolide and, in a quantitative assay we developed, dolastatin 11 was twice as active as jasplakinolide and 4-fold more active than phalloidin. However, in contrast to jasplakinolide and phalloidin, dolastatin 11 did not inhibit the binding of a fluorescent phalloidin derivative to actin polymer nor was it able to displace the phalloidin derivative from polymer. Thus, despite its structural similarity to other agents that induce actin assembly (all are peptides or depsipeptides), dolastatin 11 may interact with actin polymers at a distinct drug binding site.

(−)-Doliculide, a New Macrocyclic Depsipeptide Enhancer of Actin Assembly

“…6). As previously shown, dolastatin 11 (16) was unable to prevent the binding of FITC-phalloidin to actin polymer, whereas jasplakinolide (16,17) displaced the phalloidin analog. Quantitatively, doliculide was almost identical to jasplakinolide as an inhibitor of FITC-phalloidin binding to actin polymer.…”

“…Polymerization was examined both by the standard fluorescence assay (copolymerization of actin and pyrenyl-actin) (23) and by a centrifugal assay described previously (16).…”

“…16 for further details), actin was diluted to 25 M in the above solution and left at 0°C for 1 h. Various concentrations of drug in 5 l of dimethyl sulfoxide were added to 95-l aliquots of the actin solution. Reaction mixtures were incubated for 2 h at 22°C and centrifuged for 30 min at 45,000 rpm in a Beckman TLA 55 rotor in an Optima TLX micro-ultracentrifuge at 22°C.…”

Developmental Biology of Neoplastic Growth

“…were reported to enhance actin polymerization (Bai et al 2001;. Although precise kinetic parameters are not available, (−)-doliculide has similar EC 50 values to those of jasplakinolide on actin polymerization and on inhibition of phalloidin-binding to F-actin, whereas dolastatin 11 is twice as active as jasplakinolide but has no effect on the binding of phalloidin to actin (Bai et al 2001;.…”

“…were reported to enhance actin polymerization (Bai et al 2001;. Although precise kinetic parameters are not available, (−)-doliculide has similar EC 50 values to those of jasplakinolide on actin polymerization and on inhibition of phalloidin-binding to F-actin, whereas dolastatin 11 is twice as active as jasplakinolide but has no effect on the binding of phalloidin to actin (Bai et al 2001;. It was also demonstrated that dolastatin 11 binds to F-actin at a site different from the phalloidin-binding site, though their modes of binding to and stabilization of actin filaments are similar (Oda et al 2003).…”