Dissecting Paclitaxel–Microtubule Association: Quantitative Assessment of the 2′-OH Group (original) (raw)

Paclitaxel (PTX) is a microtubule-stabilizing agent that is widely used in cancer chemotherapy. This structurally complex natural product acts by binding to β-tubulin in assembled microtubules. The 2′-hydroxyl group in the flexible side chain of PTX is an absolute requirement for activity, but its precise role in the drug-receptor interaction has not been specifically investigated. The contribution of the 2′-OH group to the affinity and tubulin-assembly efficacy of PTX has been evaluated through quantitative analysis of PTX derivatives possessing side chain deletions: 2′deoxy-PTX, N-debenzoyl-2′-deoxy-PTX and baccatin III. The affinity of 2′-deoxy-PTX for stabilized microtubules was more than 100-fold less than that of PTX and only about 3-fold greater than the microtubule affinity of baccatin III. No microtubule binding activity was detected for the analog N-debenzoyl-2′-deoxy-PTX. The tubulin-assembly efficacy of each ligand was in concordance with the microtubule binding affinity, as was the trend in cytotoxicities. Molecular dynamics simulations revealed that the 2′-OH group of PTX can form a persistent hydrogen bond with D26 within the microtubule binding site. The absence of this interaction between 2′-deoxy-PTX and the receptor can account for the difference in binding free energy. Computational analyses also provide a possible explanation for why N-debenzoyl-2′-deoxy-PTX is inactive, in spite of the fact that it is essentially a substituted baccatin III. We propose that the hydrogen bonding interaction between the 2′-OH group and D26 is the most important stabilizing interaction that PTX forms with tubulin in the region of the C-13 side chain. We further hypothesize that the substituents at the 3′-position function to orient the 2′-OH group for a productive hydrogen bonding interaction with the protein. The diterpenoids paclitaxel (PTX, Taxol ®) and docetaxel (TXT, Taxotere ®) are highly effective drugs for the treatment of a variety of cancers. PTX, TXT and related taxanes act by interfering with cellular microtubule dynamics through binding to β-tubulin.(1) PTX was the first substance found to bind to microtubules and stabilize them against disassembly.(2) The clinical success of PTX and TXT prompted many studies of structure-activity relationships in this class and searches for other molecular entities that display PTX-like † Financial support was provided by the National Institutes Health (Grant No. CA-69571).