Two efficient protocols for the synthesis of tert-butyl (5S,6R,2E, 7E)-5-[(tert-butyldimethylsilyl)oxy]-6-methyl-8-phenyl-2, 7-octadienoate, a major component of the cryptophycins, are reported. The first utilized the Noyori reduction and Frater alkylation of methyl 5-benzyloxy-3-oxopentanoate to set two stereogenic centers, which became the C16 hydroxyl and C1' methyl of the cryptophycins. The second approach started from 3-p-methoxybenzyloxypropanal and a crotyl borane reagent derived from (-)-alpha-pinene to set both stereocenters in a single step and provided the dephenyl analogue, tert-butyl (5S,6R,2E)-5-[(tert-butyldimethylsilyl)oxy]-6-methyl-2, 7-octadienoate, in five steps. This compound was readily converted to the 8-phenyl compound via Heck coupling. The silanyloxy esters were efficiently deprotected and coupled to the C2-C10 amino acid fragment to provide desepoxyarenastatin A and its dephenyl analogue. The terminal olefin of the latter was further elaborated via Heck coupling. Epoxidation provided cryptophycin-24 (arenastatin A).

A library with 63 paclitaxel analogues modified at the C10 position of paclitaxel has been prepared using parallel solution phase synthesis. Most of the C10 analogues were slightly less active than paclitaxel in the tubulin assembly assay and had reduced potency in the B16 melanoma and MCF-7 cell line cytotoxicity assays. These modifications at C10, however, did not lead to the total loss of activity, indicating that the C10 moiety of paclitaxel may not be directly involved in the drug-microtubule interactions, but could influence its binding affinity to P-glycoprotein. Approximately 50% of the analogues demonstrated better activity against the drug resistant cell line MCF7-ADR. However, the increase in activity was 10-fold at most. This result demonstrates that the cytotoxicity against this drug resistant cancer cell line is sensitive to structural changes at the C10 position of paclitaxel. It was also found that the presence of a nitrogen atom in the C10 substituent might play a role in the interaction of analogues with microtubules.

To study the mechanism of binding of the antitumor agent paclitaxel to microtubules and tubulin, we have synthesized a fluorescent analogue of the drug. A dimethylamino group was introduced onto the 3'-N-benzoyl group of paclitaxel. This compound was synthesized from N-debenzoylpaclitaxel and 3-(dimethylamino)benzoyl chloride in 67% yield. N-Debenzoyl-N-[3-(dimethylamino)benzoyl]-paclitaxel has activity similar to paclitaxel in inducing microtubule assembly and binds to tubulin at the paclitaxel-binding site. Under assembly conditions, binding of this paclitaxel analogue to tubulin occurs in a time-dependent manner and is accompanied by a large increase in fluorescence intensity, as well as a large blue shift in the emission maximum. In addition, evidence is presented to show that this compound also binds to tubulin in the dimeric state, but the binding affinity is much lower (Kd = 49 +/- 8 microM at 25 degrees C) than that reported for polymeric tubulin. The fluorescent paclitaxel analogue, with a high quantum yield, will be a useful tool in studying the mechanism of paclitaxel binding to tubulin and the environment of the paclitaxel-binding site on tubulin.

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Chemical, conformational and structure-activity studies of taxol and related taxanes are detailed. Semisynthetic methodology for the preparation of taxol and related analogues with modified C-l3 phenylisoserine side chains was developed and analogues, modified at the C-3' phenyl group and the N-benzoyl group, were prepared. 3'-Cyclohexyl and 2-cyclohexylcarbonyl taxol analogues and C-13 side chain homologated derivatives were synthesized. Methods for the selective hydrolysis of all ester groups in baccatin ΙΠ and the conversion of 4-deacetylbaccatin III to 4-deacetyltaxol are reported. Reduction of taxanes with samarium diiodide provided 10-deacetyl derivatives as well as 9-dihydrotaxanes. Conformational analysis of taxol and other bioactive derivatives demonstrated the formation of hydrophobically clustered conformations in aqueous solvents.The discovery by the Potier group that 10-deacetylbaccatin III (4) can be isolated in significant quantities from a regenerable source, the needles of the European yew tree Taxus baccata L., was the most significant finding in the attempt to secure the long term supply of the anticancer agent taxol (1) through semisynthesis (Fig. 1) (7). Extraction of the fresh needles yields 4 in amounts of up to lg/kg, which is about ten times the amount of taxol isolated from the bark (0. lg/kg). It is of importance to note that the needles are a fully regenerable source and that their harvest does not threaten the survival of the yew species. The availability of 4 also facilitated semisynthetic studies directed at the elucidation of the taxol pharmacophore (2,3). Synthesis and Biological Evaluation of C-13 Chain Modified Taxol AnaloguesSince the C-13 Af-benzoyl-3-phenylisoserine side chain of taxol is of crucial importance for taxol's cytotoxicity (4), efficient methodology for the asymmetric synthesis of the C-13 side chain 2 and its attachment to baccatin III required development (5). NOTE: Paclitaxel is the generic name for Taxol, which is now a registered trademark. 0097-6156/95/0583-0217$08.00/0