Paclitaxel side chain production

In an alternate process for the preparation of the C-13 paclitaxel side chain, the enantioselective enzymatic hydrolysis of racemic acetate ci5 -3-(acetyloxy)-4-phenyl-2-azetidinone 38 (Eignre 16.10B), to the corresponding (S)-alcohol 39 and the nnreacted desired (l )-acetate 38 was demonstrated [63] nsing lipase PS-30 from Pseudomonas cepacia (Amano International Enzyme Company) and BMS lipase (extracellnlar lipase derived from the fermentation of Pseudomonas sp. SC 13856). Reaction yields of more than 48% (theoretical maximnm yield 50%) with EEs greater than 99.5% were obtained for the (R)-38. BMS lipase and lipase PS-30 were immobilized on Accnrel polypropylene (PP), and the immobilized lipases were reused (10 cycles) without loss of enzyme activity, productivity, or the EE of the product (R)-38. The enzymatic process was scaled up to 250 L (2.5 kg substrate input) using immobilized BMS lipase and lipase PS-30. Prom each reaction batch, R-acetate 38 was isolated in 45 mol% yield (theoretical maximum yield 50%) and 99.5% EE. The (R)-acetate was chemically converted to (R)-alcohol 39. The C-13 paclitaxel side-chain synthon (2R,3S-37 or R-39) produced by either the reductive or resolution process could be coupled to bacattin III 34 after protection and deprotection to prepare paclitaxel by a semisynthetic process [64]. 

The above in situ strategy was also applied to Schiff base substrates (in Ueu of aldehydes or ketones), affording azuidination products in fair yield, moderate dr, and outstanding enantioselectivity [44, 45]. The specific examples of benzyUdene transfer illustrated in Table 4 confirm that electron-poor (entry 1), electron-rich (entry 4), aliphatic (entry 2), and a,p-unsaturated groups are tolerated in the A-SES imine electrophiles. As shown in Scheme 17, the utility of the method was showcased in an unconventional preparation of paclitaxel side-chain 49 that ultimately begins with 3-furfural. The optically pure heterocyclic aziridine 48 was obtained as an inconsequential diastereomeric mixture by the trio of coordinated phase transfer, achiral rhodium, and chiral sulfide catalyses, albeit in the antipodal series (see ent-47) subsequent conversicm to the target amido ester was by way of standard manipulations [130]. 

Taxol (Paclitaxel) 137, a natural product derived from the bark of the Pacific yew, Taxus brevifolia [213-215], and the hemisynthetic analogue Docetaxel (Taxotere) 138, two recent and promising antitumour agents, have been the matter of extensive in vivo and in vitro animal metabolic studies. The major metabolites of taxol excreted in rat bile [216] were identified as a C-4 hydroxylated derivative on the phenyl group of the acyl side chain at C-13 (139), another aromatic hydroxylation product at the mefa-position on the benzoate group at C-2 (140) and a C-13 deacylated metabolite (baccatin III, 142) the structure of six minor metabolites could not be determined. The major human liver microsomal metabolite, apparently different from those formed in rat [217], has been identified as the 6a-hydroxytaxol (141) [218, 219]. A very similar metabolic pattern was... 

Fig. 2. Schematic representation of paclitaxel biosynthesis. Dimethylallyl-diphosphate and isopentenyl-diphosphate are condensed through geranylgeranyl diphosphate synthase activity to render geranylgeranyl-diphosphate (GGPP). GGPP is converted into taxa-4(5), 11 (12)-diene in a reaction catalyzed by the taxane synthase (TS). A series of reactions catalyzed by cytochrome P450 monoxygenases lead to the production of a taxane intermediate that is further converted to baccatin III through enzymes-driven oxidation and oxetane ring formation. The side chain moiety of paclitaxel is derived from L-phenylalanine. Three consecutive arrows mean multiple steps. Ac, acetyl Bz, benzoyl.

Genisson et al. also prepared two diastereomers of 2 -hydroxymethyl analogs through an asymmetric Baylis-Hillman reaction-like sequence to prepare a A-substimted 2 -methylene C-13 side chain (Scheme 3-3). After incorporation of the side chain to C-13 of 7,10-ii/-Troc-10-DAB, the product was then subjected to osmylation to yield 2 (R)- and 2 (5)-hydroxymethyl docetaxel 7a-b analogs stereoselectively. Both taxoids displayed less tubulin polymerization abilities than did paclitaxel, but the major product 2 / -isomer 7a is more active than the minor one 2 5-isomer 7b. ... 

Nearly all of the impurities contained the characteristic paclitaxel core substructure as indicated by the characteristic product ion at w/z 509 with variations due to modifications. Many of these taxanes contained a side-chain similar to paclitaxel, with variations occurring on the terminal amide of the side chain. The product ions that differed from the characteristic side-chain ions of paclitaxel (m/z 286) by values indicative of specific substructures were used to identify these terminal amide variations. A comparison with the paclitaxel substructural template indicated structural differences beyond the position of the amide group in the side-chain substructure. 

Initially, researchers focused on the separation of paclitaxel from cephalomannine, a paclitaxel analogue that shares the C-13 ester side-chain and, therefore, exhibits cytotoxic activity. Cephalomannine was found to elute close to paclitaxel and caused interferences for determination and purification purposes. Lately, the main taxane targets are baccatin III and 10-deacetyl-baccatin III (10-DAB III). Both compounds lack the C-13 side-chain (Fig. 1) thus, they do not show antitumour activity. Both baccatins, especially 10-DAB III, serve as synthons for the synthesis of paclitaxel or analogues. 7-Epi-paclitaxel differs from paclitaxel only in the stereochemistry of the hydroxy group in the C-7 position. 7-Epi-paclitaxel, a product of paclitaxel epimerization, shows cytotoxic activity and difficulty in separation from paclitaxel. The deacetyl derivatives of paclitaxel, 7-epi-paclitaxel, baccatin III, and cephalomannine, are also quite often the... 

A chief problem in the early development of taxane pharmaceuticals was the establishment of a steady and dependable supply. Taxanes are present in low quantities in plants. In contrast, taxines are relatively abundant in plants (especially in Taxus baccata and Taxus cuspidate), they can serve as an alternative starting material for semisynthetic production of paclitaxel derivatives. The major taxines are taxine A and taxine There is a significant structural resemblance between taxine B and taxanes (Fig. 1). Both groups share the main taxane ring moreover, the C-5 side chain of the taxines has a close spatial position to the C-13 side chain of the taxanes (the latter is essential for antitumor activity). A biosynthetic hypothesis involved the intermediacy of a C-5 to a C-13 ester transfer it was also demonstrated that taxine B can be converted into a baccatin V derivative. Taxines do not show antitumor activity, whereas the cardiotoxicity of taxol is lower compared to taxines but is undesirable for... 

Another Bristol-Meyers Squibb process represents an enzymatic route for the production of side-chain precursors of Paclitaxel (33, Scheme 10) [69]. Racemic czs-azetidinone acetate (rac-31) is subjected to the hydrolytic treatment of Pseudomonas cepacia lipase (PCL), which is used in its immobilized form on polypropylene beads. Thus, (3R,4S)-acetate 31 can be obtained in high ee as well as the remaining alcohol 32. The process takes place in 150 1 reactors where 1.2 kg mc-31/batch can be resolved with a hydrolysis rate of 0.12 g/lh. Lowering the reaction temperature to 5 °C after full conversion causes (3R,4S)-31 subsequently to crystallize. Due to the immobilization, the enzyme can be reused for at least ten cycles without any loss of activity, productivity, or optical purity of the product. Paclitaxel is finally accessible by further chemical steps. 

Prize in Chemistry. (The other half of the 2001 prize was awarded to W. Knowles and R. Noyori for their development of catalytic asymmetric reduction reactions see Section 7.14A.) The following reaction, involved in an enantioselective synthesis of the side chain of the anticancer drug paclitaxel (Taxol), serves to illustrate Sharpless s catalytic asymmetric dihydroxylation. The example utilizes a catalytic amount of K20s02(0H)4, an OSO4 equivalent, a chiral amine ligand to induce enan-tioselectivity, and NMO as the stoichiometric co-oxidant. The product is obtained in 99% enantiomeric excess (ee) ... 

Paditaxel, a complex terpene molecule with antimitotic activity (commercialized as Taxol ), is used for various cancer treatments, including ovarian and breast cancer treatments. The isolation of this naturally occurring compound requires the harvesting of a large amount of trees and involves difficult purification. A semisynthetic route has been developed based on the coupling of baccatin III (a diterpenoid that can be isolated from yew leaves) with the chiral side-chain (2R,3S)-N-benzoyl-3-phenylisoserine ethyl ester. The latter could be obtained from the diastereoselective microbial reduction of racemic 2-keto-3-(N-benzoylamino)-3-phenylpropionic acid ethyl ester using whole cells of Hansemda sp. (Figure 13.12). Despite concomitant production of the undesired anti diastereomers (2R,3R)-N-benzoyl-3-phenylisoser-ine ethyl ester and (2S,3S)-N-benzoyl-3-phenylisoserine ethyl ester (up to 20%) due to nonperfect selectivity, the C-13 side-chain synthon required for paclitaxel... 

Paclitaxel (Taxol) 57, a natural terpenoid product derived from the bark of the Pacific yew Taxus brevifolia [83-85], and its hemisynthetic analog docetaxel (Taxotere) 58, are two recently discovered promising antitumor agents. At this moment, very few data have been published about the microbial metabolism of taxoid compounds only site-specific hydrolyses of acyl side chains at C-13 or C-10 by extracellular and intracellular esterases of Nocardioides albus SC 13911 and N. lutea SC 13912, respectively, have been reported [86]. On the other hand, Hu et al. [87-89] recently described some fungal biotransformations of related more abundant natural taxane diterpenes extracted from Chinese yews or their cell cultures, in order to obtain new active substances or precursors for hemisyn-thesis of paclitaxel analogs. 

---