Structural Determinants of the Dictyostatin Chemotype for Tubulin Binding Affinity and Antitumor Activity Against Taxane- and Epothilone-Resistant Cancer Cells.

A combined biochemical, structural, and cell biology characterization of dictyostatin is described, which enables an improved understanding of the structural determinants responsible for the high-affinity binding of this anticancer agent to the taxane site in microtubules (MTs). The study reveals that this macrolide is highly optimized for MT binding and that only a few of the structural modifications featured in a library of synthetic analogues resulted in small gains in binding affinity. The high efficiency of the dictyostatin chemotype in overcoming various kinds of clinically relevant resistance mechanisms highlights its potential for therapeutic development for the treatment of drug-resistant tumors. A structural explanation is advanced to account for the synergy observed between dictyostatin and taxanes on the basis of their differential effects on the MT lattice. The X-ray crystal structure of a tubulin-dictyostatin complex and additional molecular modeling have allowed the rationalization of the structure-activity relationships for a set of synthetic dictyostatin analogues, including the highly active hybrid 12 with discodermolide. Altogether, the work reported here is anticipated to facilitate the improved design and synthesis of more efficacious dictyostatin analogues and hybrids with other MT-stabilizing agents.

MT-Stabilizer, Dictyostatin, Exhibits Prolonged Brain Retention and Activity: Potential Therapeutic Implications.

Inclusions comprising the microtubule (MT)-stabilizing protein, tau, are found within neurons in the brains of patients with Alzheimer's disease and related neurodegenerative disorders that are broadly referred to as tauopathies. The sequestration of tau into inclusions is believed to cause a loss of tau function, such that MT structure and function are compromised, leading to neuronal damage. Recent data reveal that the brain-penetrant MT-stabilizing agent, epothilone D (EpoD), improves cognitive function and decreases both neuron loss and tau pathology in transgenic mouse models of tauopathy. There is thus a need to identify additional MT-stabilizing compounds with blood-brain barrier (BBB) permeability and slow brain clearance, as observed with EpoD. We report here that the MT-stabilizing natural product, dictyostatin, crosses the BBB in mice and has extended brain retention. Moreover, a single administration of dictyostatin to mice causes prolonged stabilization of MTs in the brain. In contrast, the structurally related MT-stabilizer, discodermolide, shows significantly less brain exposure. Thus, dictyostatin merits further investigation as a potential tauopathy therapeutic.

Total synthesis and biological evaluation of a series of macrocyclic hybrids and analogues of the antimitotic natural products dictyostatin, discodermolide, and taxol.

The design, synthesis, and biological evaluation of a series of hybrids and analogues of the microtubule-stabilizing anticancer agents dictyostatin, discodermolide, and taxol is described. A 22-membered macrolide scaffold was prepared by adapting earlier synthetic routes directed towards dictyostatin and discodermolide, taking advantage of the distinctive structural and stereochemical similarities between these two polyketide-derived marine natural products. Initial endeavors towards accessing novel discodermolide/dictyostatin hybrids led to the adoption of a late-stage diversification strategy and the construction of a small library of methyl-ether derivatives, along with the first triple hybrids bearing the side-chain of taxol or taxotere attached through an ester linkage. Biological assays of the anti-proliferative activity of these compounds in a series of human cancer cell lines, including the taxol-resistant NCI/ADR-Res cell line, allowed the proposal of various structure-activity relationships. This led to the identification of a potent macrocyclic discodermolide/dictyostatin hybrid 12 and its C9 methoxy derivative 38, accessible by an efficient total synthesis and with a similar biological profile to dictyostatin.

Total synthesis and biological evaluation of novel C2-C6 region analogues of dictyostatin.

By exploiting a Still-Gennari HWE coupling with a common C11-C26 aldehyde, a series of C2-C6 modified analogues of the microtubule-stabilising marine natural product dictyostatin were synthesised and evaluated in vitro for growth inhibition against a range of human cancer cell lines, including the (P-glycoprotein efflux-mediated) Taxol-resistant NCI/ADR cell line. Removal of the C6 methyl substituent in dictyostatin was found to be well tolerated and led to the retention of antiproliferative activity in the low nanomolar range (IC(50)=43 nM in the NCI/ADR cell line), while partial and full saturation of the (2Z,4E)-dienoate region led to a progressive reduction in biological potency. The lactone ring size was found to be critical, as C21 to C19 translactonisation to afford 20-membered isodictyostatin analogues led to a significant loss of cytotoxicity. In a series of incubatory experiments performed on the PANC-1 cell line, all three of the 22-membered macrolide analogues acted in an analogous fashion to dictyostatin, through a mechanism of microtubule stabilization, causing both an accumulation of cells at the G2/M phase and formation of characteristic dense intracellular microtubule bundles.

Total synthesis and biological evaluation of potent analogues of dictyostatin: modification of the C2-C6 dienoate region.

By exploiting a Still-Gennari olefination of a common C11-C26 aldehyde with a C4-C10 or C1-C10 beta-ketophosphonate, three modified C2-C6 region analogues of the 22-membered macrolide dictyostatin were synthesised and evaluated in vitro for growth inhibition against a range of human cancer cell lines, including the Taxol-resistant NCI/ADR-Res cell line. 6-Desmethyldictyostatin and 2,3-dihydrodictyostatin displayed potent (low nanomolar) antiproliferative activity, intermediate between dictyostatin and discodermolide, while 2,3,4,5-tetrahydrodictyostatin showed activity comparable to discodermolide. As with dictyostatin, these simplified analogues act through a mechanism of microtubule stabilisation, G2/M arrest and apoptosis.

The bound conformation of microtubule-stabilizing agents: NMR insights into the bioactive 3D structure of discodermolide and dictyostatin.

A protocol based on a combination of NMR experimental data with molecular mechanics calculations and docking procedures has been employed to determine the microtubule-bound conformation of two microtubule-stabilizing agents, discodermolide (DDM) and dictyostatin (DCT). The data indicate that tubulin in assembled microtubules recognizes DDM through a conformational selection process, with minor changes in the molecular skeleton between the major conformer in water solution and that bound to assembled microtubules. For DCT, the deduced bound geometry presents some key conformation differences around certain torsion angles, with respect to the major conformer in solution, and still displays mobility even when bound. The bound conformer of DCT resembles that of DDM and provides very similar contacts with the receptor. Competition experiments indicate that both molecules compete with the taxane-binding site. A model of the binding mode of DDM and DCT to tubulin is proposed.

Development of practical syntheses of the marine anticancer agents discodermolide and dictyostatin.

Initially isolated in trace quantities from deep-sea sponges, the structurally related polyketides discodermolide and dictyostatin share the same microtubule-stabilizing antimitotic mechanism as Taxol. Discodermolide has been the focus of intense research activity in order to develop a practical supply route, and these efforts ultimately allowed its large-scale synthesis and the initiation of clinical trials as a novel anticancer drug. Similarly, the re-isolation and synthesis of dictyostatin continues to stimulate the biological and chemical communities in their quest for the development of new chemotherapeutic agents. This comprehensive review chronicles the synthetic endeavours undertaken over the last 15 years towards the development and realization of practical chemical syntheses of discodermolide and, more recently, dictyostatin, focusing on the methods and strategies employed for achieving overall stereocontrol and key fragment unions, as well as the design and synthesis of novel hybrid structures.

Synthesis and biological evaluation of 10,11-dihydrodictyostatin, a potent analogue of the marine anticancer agent dictyostatin.

By employing a diverted total synthesis strategy with late-stage intermediates, 10,11-dihydrodictyostatin ( 5) was prepared and evaluated in vitro for growth inhibition against a range of human cancer cell lines, including the NCI/ADR Taxol-resistant cell line. This novel dictyostatin analogue was found to retain potent antimitotic activity, with a comparable profile to discodermolide and Taxol, functioning by microtubule stabilization and G2/M arrest. These SAR studies provide further insight into the interaction between dictyostatin ( 1) and its tubulin target.

Synthesis and biological evaluation of novel analogues of dictyostatin.

Novel analogues of the microtubule-stabilising agent dictyostatin were designed using existing SAR information from the structurally related discodermolide, synthesised by a late-stage diversification strategy and evaluated in vitro for growth inhibition against a range of human cancer cell lines, including those known to exhibit Taxol-resistance (AsPC-1, DLD-1, PANC-1, NCI/ADR).

Design, synthesis and biological evaluation of a macrocyclic discodermolide/dictyostatin hybrid.

A 22-membered macrocyclic discodermolide/dictyostatin hybrid has been designed and synthesised; biological evaluation against a range of human cancer cell lines revealed significant levels of growth inhibition.