Mutations in the β-tubulin binding site for peloruside A confer resistance by targeting a cleft significant in side chain binding

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

“…1 The site is situated on the exterior of the MT on β-tubulin near the charged C-terminal tail, which was recently validated with the aid of macrolide-resistant cell lines. 2,3 Laulimalide exhibits microtubule-stabilizing activity and presents new opportunities for the development of antimitotic therapies. 4 It is highly cytotoxic, inhibiting cell proliferation in numerous cancer cell lines at low nM IC 50 values.…”

βI-tubulin mutations in the laulimalide/peloruside binding site mediate drug sensitivity by altering drug–tubulin interactions and microtubule stability

“…3 and 4), these data suggest that the predominant effect of this residue in the absence of a microtubule-targeting agent is one of microtubule destabilization. As with the N337D/R306H double mutation in the study by Begaye et al [34] and V333W in our study, the N337L mutation produced less stable microtubules in the cells and therefore an increase in resistance to MSAs and an increase in sensitivity to VBL, but to a greater degree than V333W. The origin of this reduction in stability of microtubules containing the N337L mutation remains to be fully elucidated, but may lie in the loss of a hydrogen bonding interaction with the carbonyl group of the neighbouring V333 (Fig.…”

“…Begaye et al [34] had previously reported in 1A9 human ovarian cancer cells that a combined N337D mutation heterozygous with R306H induced resistance to PLA. On the basis of our preliminary binding model, it was expected that N337 would hydrogen-bond to PLA; therefore, N337L would exhibit resistance due to loss of this hydrogen bond.…”

“…Supporting the notion that these drugs occupy a non-taxoid site location on β-tubulin, the mutant cells showed normal sensitivity to drugs that bind to taxoid-, vinca-or colchicine-sites on β-tubulin. In addition to these two mutant loci, a later study by Begaye et al [34] demonstrated resistance to PLA and LAU when mutations R306H and A296S at the same amino acid positions as in our study and two additional mutations Y340S and N337D were selected for by culture of 1A9 cells in high concentrations of PLA. Before selecting for actual mutated cells, these residues had been previously identified by computer docking modelling as essential sites for PLA and LAU binding to tubulin [20][21][22].…”

Expression of recombinant alpha and beta tubulins from the yew Taxus cuspidata and analysis of the microtubule assembly in the presence of taxol

“…The recombinant microtubule composed of the alpha and beta tubulins from T. cuspidata was found to be more tolerant to taxol more than the microtubule from HEK293T cells. The treatment of cancer cells with microtubule-associated drugs, including taxol, 5) peloruside A, 33) and laulimalide, 34) facilitates the acquisition of the mutations in the beta tubulin of the resistant cells, although we cloned the alpha and beta tubulin cDNAs from HEK293T cells in the absence of taxol. In addition, the deduced amino acid sequences of alpha and beta human tubulins were identical to those registered in the database (Accession Number: NP_006073.2 and NP_821133.1), suggesting that the recombinant human microtubule is supposedly sensitive to taxol.…”