The Microtubule Stabilizing Agent Laulimalide Does Not Bind in the Taxoid Site, Kills Cells Resistant to Paclitaxel and Epothilones, and May Not Require Its Epoxide Moiety for Activity

Abstract

Laulimalide is a cytotoxic natural product that stabilizes microtubules. The compound enhances tubulin assembly, and laulimalide is quantitatively comparable to paclitaxel in its effects on the reaction. Laulimalide is also active in P-glycoprotein overexpressing cells, while isolaulimalide, a congener without the drug's epoxide moiety, was reported to have negligible cytotoxic and biochemical activity [Mooberry et al. (1999) Cancer Res. 59, 653-660]. We report here that laulimalide binds at a site on tubulin polymer that is distinct from the taxoid site. We found that laulimalide, while as active as paclitaxel, epothilone A, and eleutherobin in promoting the assembly of cold-stable microtubules, was unable to inhibit the binding of radiolabeled paclitaxel or of 7-O-[N-(2,7-difluoro-4'-fluoresceincarbonyl)-L-alanyl]paclitaxel, a fluorescent paclitaxel derivative, to tubulin. Confirming this observation, we demonstrated that microtubules formed in the presence of both laulimalide and paclitaxel contained near-molar quantities, relative to tubulin, of both drugs. Laulimalide was active against cell lines resistant to paclitaxel or epothilones A and B on the basis of mutations in the M40 human beta-tubulin gene. We also report that a laulimalide analogue lacking the epoxide moiety, while less active than laulimalide in biochemical and cellular systems, is probably more active than isolaulimalide. Further exploration of the role of the epoxide in the interaction of laulimalide with tubulin is therefore justified.

Macromolecular Accessibility of Fluorescent Taxoids Bound at a Paclitaxel Binding Site in the Microtubule Surface

“…The compounds whose activity has been confirmed can be divided into two different groups, those that bind to the paclitaxel site (paclitaxel and its analogs, epothilones, eleuterobins, sarcodyctins, discodermolide, 3,17-diacetoxy2ethoxy-6oxo-B-homo-estra-1,3,5 (10)-triene) (49, 50, 56 -58), and those that share the laulimalide site (laulimalide and peloruside) (49,59).…”

Low-dose laulimalide represents a novel molecular probe for investigating microtubule organization

“…4 It is highly cytotoxic, inhibiting cell proliferation in numerous cancer cell lines at low nM IC 50 values. 4,5 It is also active in multidrug-resistant cancer cell lines overexpressing P-glycoprotein (P-gp) and is both effective in taxoid-resistant cell lines 5 and synergistic with taxoids. 6 These findings suggest that laulimalide may form the basis for a next-generation antimitotic agent, although we note lingering concerns over toxicity.…”

Peloruside A Does Not Bind to the Taxoid Site on β-Tubulin and Retains Its Activity in Multidrug-Resistant Cell Lines

“…It is therefore important to identify drugs that have similar or improved pharmacological properties to paclitaxel that are also effective in paclitaxel-resistant cancer cells. Most of the microtubule-stabilizing agents have been examined for their susceptibility to MDR, and three look particularly promising because of their lack of interaction with the P-gp pump: epothilone (9 -12); discodermolide (13); and laulimalide (11,14). Recently, it was shown that laulimalide is not only more toxic than paclitaxel in cells with a MDR phenotype, but it also binds to a different site on tubulin to paclitaxel, epothilone, and discodermolide (11).…”

“…Most of the microtubule-stabilizing agents have been examined for their susceptibility to MDR, and three look particularly promising because of their lack of interaction with the P-gp pump: epothilone (9 -12); discodermolide (13); and laulimalide (11,14). Recently, it was shown that laulimalide is not only more toxic than paclitaxel in cells with a MDR phenotype, but it also binds to a different site on tubulin to paclitaxel, epothilone, and discodermolide (11). Because laulimalide does not compete with paclitaxel for binding, it was also found to be effective in paclitaxelresistant cells that have ␤-tubulin gene mutations that modify the taxoid binding site (11), thus providing additional evidence that the laulimalide binding site is unique to that of paclitaxel.…”

“…Recently, it was shown that laulimalide is not only more toxic than paclitaxel in cells with a MDR phenotype, but it also binds to a different site on tubulin to paclitaxel, epothilone, and discodermolide (11). Because laulimalide does not compete with paclitaxel for binding, it was also found to be effective in paclitaxelresistant cells that have ␤-tubulin gene mutations that modify the taxoid binding site (11), thus providing additional evidence that the laulimalide binding site is unique to that of paclitaxel. Interestingly, epothilone and discodermolide, both of which compete with paclitaxel for binding to ␤-tubulin, have different sensitivities to particular mutations of the ␤-tubulin gene (9).…”