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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptNIH Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
J Med Chem. Author manuscript; available in PMC Aug 29, 2009.
Published in final edited form as:
PMCID: PMC2734907
NIHMSID: NIHMS102336

Design, Synthesis and Biological Evaluation of New Generation Taxoids

Abstract

Novel second-generation taxoids with systematic modifications at the C2, C10, and C3’N positions were synthesized and their structure-activity relationships studied. A number of these taxoids exhibited exceptionally high potentency against multidrug drug-resistant cell lines, and several taxoids exhibited virtually no difference in potency against the drug-sensitive and drug-resistant cell lines. These exceptionally potent taxoids were termed “third-generation taxoids”. 19 (SB-T-1214), 14g (SB-T-121303), and 14i (SB-T-1213031), exhibited excellent activity against paclitaxel-resistant ovarian cancer cell lines as well, wherein the drug-resistance is mediated by β-tubulin mutation. These taxoids were found to possess exceptional activity in promoting tubulin assembly, forming numerous very short microtubules similar to those formed by discodermolide. Taxoids 19 and 14g also showed excellent cytotoxicity against 4 pancreatic cancer cell lines, expressing 3–4 multidrug resistant genes. Moreover, taxoid 19 exhibited excellent in vivo efficacy against highly drug-resistant CFPAC-1 pancreatic as well as DLD-1 human colon tumor xenografts in mice.

Introduction

Cancer is one of the most common lethal diseases in the world. In the USA cancer is now the leading cause of death in people under the age of 85.1,2 Among a variety of chemotherapeutic drugs paclitaxel and docetaxel are currently two of the most widely used drugs in the fight against cancer, especially for the treatment of ovarian, breast, and lung cancers as well as kaposi’s sarcoma.3,4 Further clinical applications are ongoing against different types of cancers as well as for combination therapies with other anticancer drugs.3 These “taxane” anticancer drugs bind to the β-tubulin subunit, accelerate the polymerization of tubulin, and stabilize the resultant microtubules, thereby inhibiting their depolymerization.5,6 This results in the arrest of the cell division cycle mainly at the G2/M stage, leading to apoptosis through the cell-signaling cascade. Although both paclitaxel and docetaxel possess potent antitumor activity, it has been shown that treatment with these drugs often encounters undesirable side effects as well as drug resistance.3 Therefore, it is important to develop new taxane anticancer agents with fewer side effects, superior pharmacological properties, and improved activity against various classes of tumors, especially against drug-resistant human cancer.

Accordingly, extensive structure-activity relationship (SAR) studies on paclitaxel and its congeners have been performed in different laboratories, to date, for discovery and development of better taxane anticancer agents.4,716 In the course of our SAR study on taxoids, we found that (i) the C3’-phenyl group was not an essential component for their potent activity and (ii) the modifications of the C10 position with certain acyl groups as well as the replacement of the phenyl group with an alkenyl or alkyl group at the C3’ position made compounds 1–2 orders of magnitude more potent than the parent drugs (paclitaxel and docetaxel) against drug resistant human breast cancer cell lines. These highly potent taxoids were termed “second-generation taxoids”.17 Furthermore, we found that introduction of a substituent (e.g., MeO, N3, Cl, F, etc.) to the meta position of the C2-benzoyl group of the second-generation taxoids, enhanced the activities 2–3 orders of magnitude higher than the parent drugs against drug-resistant human breast cancer cell lines.14,15 We describe here a full account of our work on the synthesis, biological evaluations and SAR of a series of novel new generation taxoids bearing various substituents at the C2, C10, C3’, and C3’N, positions.

Chemical Synthesis

A series of second-generation taxoids bearing different aromatic acyl groups at the C10 position was synthesized by applying the procedure developed by Georg.18 The C7 hydroxy group of 10-deacetylbaccatin III (DAB, 1) was selectively protected as a triethylsilyl ether using triethyl silyl chloride (TESCl) and imidazole in DMF. The resulting 7-TES-DAB was acetylated specifically at the C10 position to give 7-TES-baccatin III (2) in excellent yield (95% for two steps)8,17,19 An isoserine moiety was introduced to the C13 position of 2 through the Ojima-Holton coupling of 2 with N-t-Boc-β-lactam 3a7,20 under the standard conditions in the presence of LiHMDS in THF at −40 °C to afford the corresponding taxoid 4 (Scheme 1).21,22

Scheme 1
Reagents and conditions: (i) (a) TESCl, imidazole, (b) LiHMDS, AcCl, 95% in two steps; (ii) LiHMDS, THF, 95% (iii) N2H4·H2O, EtOH, 85%.

The selective removal of the acetyl group at the C10 position of 4 with hydrazine was possible when the hydroxyl group at the C2’ position was protected as a TIPS ester. This selective deacetylation under mild conditions gave the desired C2’-TIPS-7-TES-10-OH-taxoid 5 in high yield (Scheme 1).

Taxoid 5 served as the key intermediate for the syntheses of a number of second-generation taxoids, bearing different acyl groups at the C10 position. The acylation of the C10 position of 5 was carried out using either an acid chloride with triethylamine (TEA) and DMAP or a carboxylic acid with DIC and DMAP (Scheme 2). For the introduction of a Cbz group, CbzCl and LiHMDS were used (Scheme 2). Yields for the acylation reactions were 75–98% (see Supporting Information for details). The deprotection of C2’-TIPS and 7-TES groups with HF-pyridine in pyridine/acetonitrile (1:1) proceeded smoothly to give desired 10-acyl second-generation taxoids 6a–o in 80–97% yields (Scheme 2). Results are summarized in Table 1.

Scheme 2
Reagents and conditions: (i) (a) RCl, TEA, DMAP, CH2Cl2 or (b) ROH, DIC, DMAP, CH2Cl2 or (c) RCl, LiHMDS, THF, −20 °C (75–98%); (ii) HF/pyridine, pyridine/MeCN (80–97%).
Table 1
Synthesis of Taxoids 6

A series of novel second-generation taxoids, bearing substituents at the meta position of the C-2-benzoyl group, was synthesized (Scheme 5) from the corresponding C2-modified DABs 10 (see Scheme 3 and Scheme 4). The protection of the C7, C10 and C13 hydroxy groups of DAB with TES made it possible to selectively remove the C2-benzoyl moiety with sodium bis(2-methoxyethoxy)aluminumhydride, giving 7,10,13-tris-TES-2-debenzoyl-DAB 8 in excellent yield.23,24 The esterification of the C2-OH of 8 with 3-substituted benzoic acids (R1 = F, Cl, MeO, Me, N3, vinyl) with DIC and DMAP gave various C2-modified tris-TES-DABs 9a–f in 80–90% yields. Removal of all TES groups of 9a–f with HF-pyridine, followed by selective protection of the C7 hydroxyl group of the resulting baccatin gave 2-modified DABs 10a–f. Subsequent selective acylation at the C10 position of 10a–f afforded the corresponding 2,10-modified DABs 11a–l in 71–95% yields (Scheme 3). Results are summarized in Table 2.

Scheme 3
Reagents and conditions: (i) TESCl, imidazole, DMF, RT, 96% (ii) sodium bis(2-methoxyethoxy)aluminumhydride, THF, −10 °C, 97% (iii) DIC, DMAP, CH2Cl2, 85–90% (iv) HF/pyridine, pyridine/MeCN; (v) TESCl, imidazole, DMF, RT, 2h; (vi) ...
Scheme 4
Reagents and conditions: (i) TPAP, NMO, 4Å mol. Sieves, CH2Cl2, 25 °C, 99% (ii) 2-MeO-BzCl, DMAP, TEA, CH2Cl2, 100% (iii) NaBH4, MeOH/THF, 80% at 50% conversion.
Scheme 5
Reagents and conditions: (i) 3a–f (1.2~1.5 equiv), LiHMDS, THF, −40 °C, 30 min; (ii) HF/pyridine, pyridine/MeCN, 0 °C — RT, 18h; (iii) H2/Pd-C, EtOAc/MeOH, RT, 24h.
Table 2
2,10-Modified baccatins 11

The same strategy was applied to the introduction of 2-methoxybenzoyl group at the C10 position, but did not work well. Thus, we employed an alternative method, which is shown in Scheme 4. Selective oxidation of the C-13 position of 10b using TPAP and NMO provided 13-oxobaccatin 12. The C10 position was then acylated with 2-methoxylbenzoyl chloride in the presence of DMAP to afford 10-(2-methoxybenzoyl)-13-oxobaccatin 13. Reduction of the C13 ketone with NaBH4 gave desired 2-(3-methoxybenzoyl)-10-(2-methoxybenzoyl)baccatin 11k in reasonable yield.

The Ojima-Holton coupling reactions of baccatins 11a–l with β-lactams 3a–f in the presence of LiHMDS as a base 17,25 proceeded smoothly to give the corresponding taxoids 14 (see Table 3 for structures). Enantiopure β-lactams with various C4 substituents were readily obtained through efficient chiral ester enolate-imine cyclocondensations15,17,20,22,25,26 or [2+2] ketene-imine cycloaddition, followed by enzymatic optical resolution27. The deprotection of the silyl groups with HF/Pyridine gave desired taxoids 14a–p and 15c–e,g in 65–95% yields (for two steps) (Scheme 5). In addition, taxoids 14g (SB-T-121303) (R1 = MeO, R2 = propanoyl, R3 = 2-methylpropen-1-yl) was subjected to hydrogenation on Pd/C to afford 15g (R3 = 2-methylpropyl) in quantitative yield (Scheme 5). Results are summarized in Table 3.

Table 3
Second- and third-generation taxoids 14 and 15.

We also investigated the effects of the C3’N substituents on the cytotoxicity of second-generation taxoids. Modification of the C3’N position can be made by using enantiopure β-lactams bearing various N-acyl or N-carbalkoxy groups. These β-lactams were readily obtained by reacting NH-free 3-TBSO- or 3-TIPSO-β-lactam with acid chlorides or chloroformates. The resulting β-lactams 16a–h were coupled with 10-deacetyl-10-propanoylbaccatin III or baccatin 11b using 1.5–2 equivalents of β-lactam 16a–h to complete the reactions in excellent yields. Subsequent deprotection of silyl groups with HF/pyridine gave desired taxoids 17a–l in good to excellent yields. Hydrogenation of selected 17 on Pd/C the corresponding 3’-(2-methylpropyl)taxoids 18 in quantitative yields (Scheme 6). Results are summarized in Table 4.

Scheme 6
Reagents and conditions: (i) LiHMDS, THF, −40 −C, 30 min; (ii) HF/pyridine, pyridine/MeCN, 0 °C — RT, 18 h (61–86% for two steps); (iii) H2, Pd/C, EtOAc, RT, 24 h (90–92%).
Table 4
C3’N-Modified second-generation taxoids 17 and 18

Evaluation of Biological Activities

Cytotoxicity of new generation taxoids against human breast and ovarian cancer cell lines

New second-generation taxoids, thus obtained, were evaluated for their cytotoxicity against drug-sensitive (LCC6-WT: P-glycoprotein negative, Pgp−) and drug-resistant (LCC6-MDR: P-glycoprotein positive, Pgp+) human breast cancer cell lines and selected taxoids were also assayed for their potency against human breast cancer cell line MCF7 (Pgp−) and human ovarian cancer cell line NCI/ADR (Pgp+). Cytotoxicity assay results of taxoids 6a–6o, bearing various aromatic groups in the acyl moieties at the C10 position against LCC6-WT and LCC6-MDR cell lines are summarized in Table 5.

Table 5
Cytotoxicity of second-generation taxoids with modifications at C1O (IC50 nM)a

As Table 5 shows, all taxoids assayed exhibit activities similar to or slightly better than that of paclitaxel against LCC6-WT, while their activities against LCC6-MDR are two orders of magnitude better than that of paclitaxel for more than a half of this series of taxoids. The most noteworthy feature revealed in this assay can be seen in the dramatic decrease in the R/S ratio, i.e., the ratio of the IC50 value against the drug-resistant cell line vs. that against the drug-sensitive cell line (paclitaxel’s R/S value is 112 in this assay), which is a excellent indicator to see the level of drug-resistance associated with drugs. A majority of the taxoids in this series exhibit an R/S ratio at or below 3, and taxoid 6d demonstrates almost no difference against drug-resistant and drug-sensitive cell lines with the R/S ratio of 1.2. In general, taxoids with a substituted or unsubstituted benzoyl group (6a–6e) or 2-phenylpropanoyl group (6m–6o) are highly potent against LCC6-MDR, while taxoids with an arylacetyl substituents (6i–6k) show reduced activity against LCC6-MDR although 4 out of 7 taxoids in this group (6h–6k) are more potent than paclitaxel against LCC6-WT. Taxoids 6a and 6d that bear a benzoyl group and a 4-methoxybenzoyl group, respectively, at C10 possess highest potencies against LCC6-MDR (IC50 values of 4.8 and 4.7 nM, respectively). Taxoid 6k, bearing a 4-methoxyphenyl acetyl group at C10 is the least potent (IC50 = 38.4 nM) against LCC6-MDR, but 6k possesses the highest potency against LCC6-WT. Interestingly, elongation of the alkyl chain of 6i–6k just by one carbon restores high potency against LCC6-MDR.

10-Propanoyl-taxoids, bearing different acyl or carboalkoxy groups at the C3’N position were assayed for their cytotoxicity against LCC6-WT (Pgp−), LCC6-MDR (Pgp+), MCF7 (Pgp−), and NCI/ADR (Pgp+) human cancer cell lines and results are summarized in Table 6. As Table 6 shows, most of C3’N-modified taxoids possess better cytotoxicity against LCC6-WT and MCF7 cell lines and one to two orders of magnitude higher potency against drug-resistant LCC6-MDR and NCI/ADR cell lines as compared with paclitaxel. Two taxoids (17d and 17e), bearing cyclopent-2-en-1-yl and cyclohex-2-en-1-yl groups, exhibit high potency against all four cell lines examined and the R/S ratios are 9.2–15. The observed cytotoxicity values for 17d and 17e are comparable to those for the parent second-generation taxoids, 6x (SB-T-1213)17 and 6’x (SB-T-1103)17, shown in Table 2 for comparison. These results indicate that the t-Boc group at the C3’N position, which is the “gold standard”, can be replaced by these cycloalkenoyl groups without losing potency.

Table 6
Cytotoxicity of second-generation taxoids with C3’N modifications (IC50 nM)a

Cytotoxicity assay results of second-generation taxoids bearing a modified C2-benzoyl group at its meta position are summarized in Table 7. The IC50 values of paclitaxel, docetaxel, 6x, 6’x, and 19 (SB-T-1214) are also listed for comparison.17 As Table 7 shows, a majority of these new second-generation taxoids (14 and 15), bearing a t-Boc group at the C3’N position, exhibit remarkable potency against drug-resistant (Pgp+) cancer cell lines, LCC6-MDR and NCI/ADR, i.e., 2–3 orders of magnitude higher potencies than those of paclitaxel and docetaxel. In more than several cases, the R/S ratios are less than 3 and even become less than 1 in three cases (14g for LCC6-WT : LCC6-MDR and MCF7:NCI/ADR as well as 15g for LCC6-WT : LCC6-MDR). In these three cases, it can be said that the Pgp-mediated MDR is completely circumvented by new taxoids 14g and 15g. In addition to 14g and 15g, 14e and 14l also show excellent R/S ratios, i.e., 14e: R/S ratios of 1.3 and 1.2 for LCC6-WT : LCC6-MDR and MCF7:NCI/ADR, respectively; 14l: R/S ratios of 1.0 for LCC6-WT : LCC6-MDR. Accordingly, we have defined these new generation taxoids, which can virtually circumvent the Pgp-mediated MDR as the “third-generation” taxoids. It is interesting to point out that the same C2-benzoyl group modifications to the taxoids, bearing cycloalkanoyl or carbalkoxy groups at the C3’N position, do not have recognizable effects on potency, except for taxoids 18h and 18m, which exhibit sub-nanomolar IC50 values against LCC6-WT and MCF7. Taxoid 18m also shows very high potency against drug-resistant LCC6-MDR (2.6 nM) and NCI/ADR (1.18 nM) cell lines as well.

Table 7
Cytotoxicity of second-generation taxoids with C2-meta modifications (IC50 nM)a

For the effects of the meta substituents of the C2-benzoyl moiety, the potency decreases in the order: F > Cl > N3 > MeO >> CH2=CH- against LCC6-WT (Pgp−), while the order changes to MeO > N3 > Cl > F >> CH2=CH- against LCC6-MDR (Pgp+). Apparently, the former order is more or less reflecting the ability of a taxoid with such a modification to bind microtubules. On the other hand, the latter order reflects the effects of these substituents on the MDR reversal activity of taxoids or simply indicates the extent of interaction of taxoids with Pgp in the reverse order. In both case, the meta-vinyl substitution of the C2-benzoyl moiety (14f) resulted in the least potent taxoid in this comparison. The introduction of a meta-methyl group to the C2-benzoyl moiety (14a) does not seem to have a recognizable positive effect.

For the C3’ position, 2-methylprop-1-enyl and 2-methylpropyl are the best substituents for potency so far. However, the introduction of allyl, (E)-prop-1-enyl, but-3-enyl or (S)-2,2-dimethylcyclopropyl group provided very potent taxoids (14m~14p), especially (S)-2,2-dimethylcyclopropyl group (14p) appears to have a beneficial effect, comparable to those of 2-methylprop-1-enyl and 2-methylpropyl groups.

For the C10 modifications, the potency decreases in the order: 4-MeO(C6H4)CH2CO~2-MeO(C6H4)CO > MeOCO~Ac > cyclo-PrCO ~ EtCO > PhCH2OCO (Cbz) against LCC6-WT, while the order changes to 4-MeO(C6H4)CH2CO > 2-MeO(C6H4)CO~EtCO > MeOCO > Cbz > Ac < cyclo-Pr against LCC6-MDR. However, the decreasing order further changes against MCF7 and NCI/ADR, i.e., Cbz > EtCO > MeOCO > 4-MeO(C6H4)CH2CO > Ac > 2-MeO(C6H4)CO~EtCO against MCF7, while EtCO >> MeOCO > Cbz > 4-MeO(C6H4)CH2CO > Ac > 2-MeO(C6H4)CO~EtCO against NCI/ADR. These results suggest that the cancer cell types including Pgp overexpression are sensitive to the C10 substitution. Nevertheless, the difference in potency for these cases is within the factor of 10. Accordingly, all these highly potent new taxoids are very good candidates for further preclinical studies.

Cytotoxicity of new-generation taxoids against paclitaxel-resistant cancer cells with point mutations in tubulin

Multidrug-resistance to paclitaxel arises from the overexpression of ATP-binding cassette (ABC) transporters,28 but other drug-resistance mechanisms are also involved in paclitaxel-resistance.29 One of the significant mechanism is associated with alterations of its cellular target, tubulin/microtubule.3035 In this regard, two paclitaxel-resistant sublines 1A9PTX10 and 1A9PTX22, derived from 1A9 cell line, have been reported.34 The parental 1A9 is a clone of the human ovarian carcinoma cell line A-2780. Point mutations in class I β-tubulin in both 1A9PTX10 and 1A9PTX22 have been identified by sequence analysis.34 Thus, the cytotoxicity of new generation taxoids against these two paclitaxel-resistant cell lines would provide critical information about their ability to deal with drug-resistance other than MDR. Selected new generation taxoids, 19, 14g and 15g, were assayed against both drug-resistant cell lines and the parental cell line. As Table 8 shows, all three taxoids exhibit extremely potent activity, especially against drug-resistant cell lines 1A9PTX10 and 1A9PTX22, with two orders of magnitude higher potency than paclitaxel. The results clearly demonstrate that these second- and third-generation taxoids possess capability of effectively circumventing the paclitaxel drug-resistance arising from point mutations in tubulins/microtubules besides MDR. This makes the new generation taxoids even more attractive.

Table 8
Cytotoxicity of new generation taxoids against 1A9PTX10 and 1A9PTX22 cell lines (IC50 nM)a
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Cytotoxicity of new generation taxoids against pancreatic cancer cell lines

Currently there are no effective chemotherapeutic treatments for pancreatic cancer and the five year survival rate is less than 5%.36 Pancreatic cancer is refractive to conventional therapy and the expression of various multidrug resistance proteins is believed to be a major factor for the extensive drug resistance.3739 We carried out a RT-PCR analysis of the expression of multidrug resistance genes in lysates of pancreatic cancer cell lines and found that CFPAC-1 and PANC-1 cell lines expressed mdr1, mrp1, mrp2 and lrp genes, responsible for multidrug resistance, while MIA PaCa-2 and BxPC-3 cell lines expressed mrp1, mrp2 and lrp genes.40 Accordingly, it was of particular interest for us to examine the efficacy of new generation taxoids against highly multidrug resistant pancreatic cancer cell lines. Thus, two taxoids, 19 and 14g were evaluated for their cytotoxicity against 4 pancreatic cancer cell lines, MIA PaCa-2, CFPAC-1, BxPC-3 and PANC-1.

As Table 9 shows, new generation taxoids 19 and 14g exhibit excellent cytotoxicity against four pancreatic cancer cell lines with sub-nanomolar to single-digit nanomolar IC50 values except one case for 14g against PANC-1. It is worth mentioning that the third-generation taxoid 14g, which possesses superior potency (>10 times) than the second-generation taxoid 19 against Pgp+ (i.e., mdr1) human ovarian cancer cell line NCI/ADR (see Table 3), exhibits lower potency than taxoid 19 against BxPC-3 and PANC-1 cell lines, especially against the latter (22.6 nM vs. 3.68 nM). The results may suggest that taxoid 14g cannot modulate a combination of multidrug-resistant proteins so efficiently as Pgp alone. Nevertheless, 14g is still highly cytotoxic to PANC-1. On the other hand, taxoid 19 demonstrates remarkably high potency against all four pancreatic cancer cell lines. In addition, taxoid 19 did not show any appreciable cytotoxicity against primary pancreatic ductal cells up to 10 µM concentration. This is a very impressive finding.

Table 9
Cytotoxicity of 19 and 14g against pancreatic cancer cell lines (IC50 nM)a

Preliminary in vivo antitumor activity against pancreatic cancer CFPAC-1 xenograft in nude mice

Encouraged by remarkable in vitro cytotoxicity, a preliminary study on the in vivo antitumor activity of taxoid 19 against CFPAC-1 xenograft in nude mice. Animals were inoculated with 1 million CFPAC-1 pancreatic cancer cells that express high levels of mdr1 in each flank. After the tumor size had become 100 mm3 on day 19, three intravenous injections of 19 (20 mg/kg × 3, 60 mg/kg total dose in Tween 80/EtOH/PBS) were made in three days interval, i.e., days 19, 22 and 25. The treatment was highly efficacious, causing complete reduction in tumor volume. After 8 weeks tumors were recovered from the xenograft and subjected to histopathological analysis. The analysis of the stained tissues revealed that only inflammatory infiltrate and fibrotic issue remained and no trace of cancer cells. Consequently, taxoid 19 can be considered a very promising lead compound for potential treatment of pancreatic cancer.

Microtubule Polymerization Assay

The activities of 19, 14g and 14i (SB-T-1213031) were evaluated in two in vitro tubulin polymerization assays. Paclitaxel was also used as the standard for comparison. Changes in absorbance in this spectrophotometric assay provide a direct measure of turbidity, hence indicate the extent of tubulin polymerization. Taxoids 19, 14g and 14i induced tubulin polymerization in the absence of GTP in a manner similar to paclitaxel (see Figure 1 and Figure 2). The microtubules formed with these new generation taxoids as well as paclitaxel were stable against Ca2+-induced depolymerization. As Figure 1 shows, taxoids 19 and 14g promote rapid polymerization of tubulin at a faster rate than that of paclitaxel. The turbidity of the tubulin solution treated by 19 or 14g reaches plateau quickly and does not change with time. This observation may imply that there is a difference in structure between microtubules formed with the new generation taxoids and those with paclitaxel. Third-generation taxoid 14g causes spontaneous tubulin polymerization, reaching >90% of a plateau within 5 min from onset, while it takes ca. 12 min. for second-generation taxoid 19 to reach the same point.

Figure 1
Tubulin polymerization with 19, 14g and paclitaxel: microtubule protein 1 mg/mL, 37 °C, GTP 1 µM, Drug 10 µM
Figure 2
Tubulin polymerization with 14i and paclitaxel: microtubule protein 1 mg/ml, 37 °C, Drug 10 µM

In a similar manner, the activity of taxoid 14i was compared with that of paclitaxel in a tubulin polymerization assay using a slightly different protocol for tubulin preparation from that used for the experiments presented in Figure 1. As Figure 2 shows, this assay reveals a remarkable difference in the speed of tubulin polymerization between the third-generation taxoid 14i and paclitaxel. Taxoid 14i causes instantaneous polymerization of tubulin, completing the polymerization within 2 min, while paclitaxel promotes the polymerization much more slowly.

Electron Microscopy Analysis

The microtubules formed with new generation taxoids (19, 14g and 14i) were analyzed further by electron microscopy for their morphology and structure in comparison with those formed by using GTP and paclitaxel. The electron micrographs of microtubules formed with three taxoids, paclitaxel and GTP are summarized in Figure 3. As Figure 3A and 3B show, GTP and paclitaxel form long and straight microtubules. The microtubules formed with a second-generation taxoid 19 (Figure 3C) are shorter than those with GTP or paclitaxel. In contrast, the morphology of the microtubules formed by the action of third-generation taxoids 14g and 14i is unique in that those microtubules are very short and numerous (Figure 3D and 3E). The microtubules with taxoid 14g appear to have more curvature that those with 14i. It is worth mentioning that discodermolide (another potent anticancer drug candidate, stabilizing microtubules4144) forms microtubules with similar characteristics with those formed with 14g and 14i, i.e., short and numerous (Figure 3F). It is strongly suggested that the formation of short and numerous microtubules is related to the instantaneous rapid polymerization of tubulin observed with these third-generation taxoids as well as discodermolide.

Figure 3
Electromicrographs of microtubules (20,000×): (A) GTP; (B) paclitaxel; (C) 19; (D) 14g; (E) 14i; (F) discodermolide

Antitumor Activity of taxoid 19 in vivo against Pgp+ DLD-1 human colon tumor xenograft

Taxoid 19 has recently emerged as one of the leading candidates among the second- and third-generation taxoids being studied in the Ojima laboratory, especially in connection with tumor-targeting drug delivery using taxoid conjugates with omega-3 fatty acids.45 Accordingly, this taxoid was evaluated for its efficacy against a drug-resistant human colon tumor xenograft (Pgp+) DLD-1 in severe combined immune deficient mice (SCID). Taxoid was administered intravenously at three doses 3 times using 3 days regimen (q3d×3, on day 5, 8, and 11), starting from day 5 after DLD-1 subcutaneous tumor implantation. Results are summarized in Table 10.

Table 10
Antitumor effect of taxoid 19 delivered i.v. to SCID mice bearing a Pgp+ human colon tumor xenograft, DLD-1

As Table 6 shows, taxoid 19 exhibits remarkable antitumor activity in sharp contrast to paclitaxel. As anticipated, paclitaxel is ineffective against this highly drug-resistant (Pgp+) tumor at its optimal dose (60 mg/kg total dose). The best result for taxoid 19 was obtained at 60 mg/kg total dose (20mg/kg × 3), wherein complete regression of the DLD-1 tumor was achieved in five of five mice (tumor growth delay >150 days). Systemic toxicity profile shows that there was only 3–5% weight loss during the period of day 15 to day 20, and the drug was very well tolerated by animals. At 30 mg/kg total dose (10 mg/kg × 3), tumor growth delay of 37 days was observed, but all animals died before day 60. At 120 mg/kg total dose (40 mg/kg × 3), two animals died due to drug’s toxicity. Nevertheless, three animals survived till the end of the experiment on day 167 (tumor growth delay >150 days). Thus, the total dose of 60 mg/kg (20 mg/kg × 3) appears to be optimal for taxoid 19. The highly promising in vivo antitumor activity of 19 warrants further preclinical evaluation of this second-generation taxoid.

Experimental Section

General Methods

1H and 13C NMR spectra were measured on a Varian 300, 400 or 500 MHz NMR spectrometer or a Bruker AC-250 NMR spectrometer. Melting points were measured on a Thomas Hoover Capillary melting point apparatus and are uncorrected. Specific optical rotations were measured on a Perkin-Elmer Model 241 polarimeter. IR spectra were recorded on a Perkin-Elmer Model 1600 FT-IR spectrophotometer. TLC was performed on Merck DC-alufolien with Kieselgel 60F-254 and column chromatography was carried out on silica gel 60 (Merck; 230–400 mesh ASTM). Purity was determined with a Waters HPLC assembly consisting of dual Waters 515 HPLC pumps, a PC workstation running Millennium 32, and a Waters 996 PDA detector, using a Phenomenex Curosil-B column, employing CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min. High-resolution mass spectra were obtained at the Mass Spectrometry Laboratory, University of Illinois at Urbana-Champaign, Urbana, IL or the Mass Spectrometry Facility, University of California, Riverside, Riverside, CA.

Materials

The chemicals were purchased from Aldrich-Sigma Co. and used as received or purified before use by standard methods. Tetrahydrofuran was freshly distilled from sodium metal and benzophenone. Dichloromethane was also distilled immediately prior to use under nitrogen from calcium hydride. N,N-Dimethylformamide (DMF) was distilled over 4A molecular sieves under reduced pressure. 4-(N,N-Dimethylamino)pyridine (DMAP) was uses as received. 10-Deacetylbaccatin III (DAB, 1) was obtained from Indena, SpA, Italy. 7-Triethylsilylbaccatin III (2),46,47 (3R,4S)-1-tert-butoxycarbonyl-3-triisopropylsiloxy-4-(2-methylprop-1-enyl)azetidin-2-one (3a),7,20 (3R,4S)-1-tert-butoxycarbonyl-3-triisopropylsiloxy-4-(2-methylpropyl)azetidin-2-one (3b),14,48 (3R,4S)-1-(tert-butoxycarbonyl)-3-triisopropylsiloxy-4-(prop-2-enyl)azetidin-2-one (3c)49 (3R,4S)-1-(tert-butoxycarbonyl)-3-triisopropylsiloxy-4-[(E)-prop-1-enyl]azetidin-2-one (3d),7 (3R,4S)-1-(tert-butoxycarbonyl)-3-triisopropylsiloxy-4-(3-butenyl)azetidin-2-one (3e),49 (3R,4S)-1-(tert-butoxycarbonyl)-3-triisopropylsiloxy-4-[(S)-2,2-dimethylcyclopropyl]azetidin-2-one (3f),26 7,10,13-tri(triethylsilyl)-10-deacetylbaccatin III (7),23,49 7,10,13-tri(triethylsilyl)-2-debenzoyl-10-deacetylbaccatin III (8),49,50 7-triethylsilyl-10-deacetyl-10-propanoylbaccatin,17 and 3'-dephenyl-3'-(2-methyl-1-propenyl)-10-(cyclopropanecarbonyl)docetaxel (19)17 were prepared by the literature methods.

Synthesis of C10-modified taxoids 6

C-10 modified second-generation taxoids 6 were synthesized using the route illustrated in Scheme 1 via key intermediates 4 and 5, followed by acylation and deprotection. Typical procedures for the preparation of 4 and 5 as well as that for acylation and deprotection are described below.

7-Triethylsilyl-10-acetyl-2’-triisopropyl-3'-dephenyl-3'-(2-methylprop-1-enyl)docetaxel (4)

To a solution of 7-triethylsilylbaccatin III (2)46,47 (86 mg, 0.12 mmol) and 4-(2-methylprop-1-enyl)-β-lactam 37,20 (82 mg, 0.20 mmol) in 5 mL dry THF was added 1M LiHMDS (0.20 mL, 0.20 mmol) in THF dropwise at −40 °C, and the solution was stirred at the same temperature for 30 min. The reaction was quenched with saturated aqueous NH4Cl, and the aqueous layer was extracted with CH2Cl2. The combined extracts were dried over anhydrous MgSO4 and concentrated in vacuo. The residue was purified by flash chromatography on silica gel (hex:EtOAc = 6:1) to afford 4 (125 mg, 95% yield) as a white solid: 1H (300 MHz, CDCl3) δ 0.54 (m, 6 H), 0.85 (m, 9 H), 1.04 (m, 24 H), 1.15 (s, 3 H), 1.33 (s, 9 H), 1.64 (s, 6 H), 1.74 (m, 4 H), 2.16 (s, 3 H), 2.35 (s, 3 H), 2.49 (m, 1 H), 3.90 (d, J = 7.5 Hz, 1 H), 4.20 (d, J = 8.7 Hz, 1 H), 4.30 (d, 8.7 Hz, 1 H), 4.43 (m, 2 H), 4.80 (m, 2 H), 4.92 (d, J = 8.1, 1 H), 5.33 (d, J = 8.4 Hz, 1 H), 5.63 (d, J = 7.5 Hz, 1 H), 6.08 (t, J = 8.1 Hz, 1 H), 6.35 (s, 1 H), 7.47 (dd, J = 8.1 Hz, 7.2 Hz, 2 H), 7.59 (dd, J = 7.2 Hz, 8.1 Hz, 1 H), 8.09 (d, J = 7.2 Hz, 2 H).

2’-Triisopropyl-3'-dephenyl-3'-(2-methyl-1-propenyl)-7-triethylsilyldocetaxel (5)

To a solution of 4 (31 mg, 0.03 mmol) in 3 mL EtOH was added 1 mL N2H4·H2O, and the mixture was allowed to stir for 1 h. The reaction was quenched with saturated aqueous NH4Cl, and extracted 2 times with EtOAc. The combined organic layers were washed with water and brine, dried over MgSO4, filtered and concentrated in vacuo. The resulting crude solid was purified by flash chromatography on silica gel (hexanes/EtOAc = 3/1) to afford 5 (25 mg, 85% yield) as a white solid: 1H (300 MHz, CDCl3) δ 0.54 (m, 6 H), 0.85 (m, 9 H), 1.04 (m, 24 H), 1.15 (s, 3 H), 1.33 (s, 9 H), 1.64 (s, 6 H), 1.74 (m, 4 H), 2.34 (m, 1 H), 2.37 (s, 3 H), 3.90 (d, J = 7.5 Hz, 1 H), 4.20 (d, J = 8.7 Hz, 1 H), 4.30 (d, 8.7 Hz, 1 H), 4.42 (m, 2 H), 4.80 (m, 2 H), 4.92 (d, J = 8.1, 1 H), 5.11 (d, J = 1.8 Hz, 1 H), 5.33 (d, J = 8.4 Hz, 1 H), 5.63 (d, J = 7.5 Hz, 1 H), 6.15 (t, J = 8.1 Hz, 1 H), 7.47 (dd, J = 8.1 Hz, 7.2 Hz, 2 H), 7.59 (dd, J = 7.2 Hz, 8.1 Hz, 1 H), 8.09 (d, J = 7.2 Hz, 2 H).

Preparation of 2’,7-protected taxoid 5A

Method A. To a solution of 5, 3 equivalents of DMAP, and 3 equivalents of triethylamine in CH2Cl2, were added 3 equivalents of the corresponding acid chloride. The solution was allowed to stir for 12–72 h at 25–40 °C, diluted with EtOAc, and quenched with saturated aqueous NaHCO3. The organic layer was washed with water and brine, dried over MgSO4, filtered and concentrated. The resulting solid was purified by flash chromatography on silica gel to afford the corresponding coupling product, 2’-triisopropyl-3'-dephenyl-3'-(2-methyl-1-propenyl)-7-triethylsilyl-10-acyldocetaxel (5A). Method B. To a solution of 5, 3 equivalents of DMAP, and 3 equivalents of the corresponding acid in CH2Cl2, was added 3 equivalents of DIC. The solution was allowed to stir for 12–72 h at 25 °C, diluted with EtOAc, and quenched with saturated aqueous NaHCO3. The organic layer was washed with water and brine, dried over MgSO4, filtered and concentrated. The resulting solid was purified by flash chromatography on silica gel to afford the desired coupling product 5A.

Preparation of taxoids 6 through deprotection of 5A

A typical procedure is described for the synthesis of 3'-dephenyl-3'-(2-methyl-1-propenyl)-10-(2-methoxybenzoyl)docetaxel (6b): To a solution of 10 mg (0.01 mmol) of 2’-triisopropyl-3'-dephenyl-3'-(2-methyl-1-propenyl)-7-triethylsilyl-10-(2-methoxybenzoyl)docetaxel (5A–b) in 1 mL of pyridine/acetonitrile (1/1) was added dropwise 0.1 mL of HF/pyridine at 0 °C, and the mixture was stirred at room temperature overnight. The reaction was quenched with saturated aqueous NaHCO3. The mixture was then diluted with ethyl acetate, washed with saturated CuSO4 solution and water, dried over anhydrous MgSO4 and concentrated in vacuo. The crude product was purified by column chromatography on silica gel (hexanes/EtOAc = 1/1) to afford 5 mg (90% yield) of taxoid 6b as a white solid: mp 150–158 °C; [α]D20 −75 (c 0.010, CDCl3); 1H NMR (250 MHz, CDCl3) δ 1.20 (m, 6 H), 1.31 (s, 9 H), 1.54 (s, 3 H), 1.63 (s, 3 H), 1.82 (m, 1 H), 1.84 (s, 3 H), 2.01 (s, 3 H), 2.36 (s, 3 H), 2.57 (m, 2 H), 3.39 (bs, 1 H), 3.89 (m, 4 H), 4.20 (m, 2 H), 4.31 (d, J= 8.1 Hz, 1 H), 4.48 (m, 1 H), 4.63 (m, 2 H), 4.98 (d, J = 8.7 Hz, 1 H), 5.25 (m, 1 H), 5.70 (d, J = 6.9 Hz, 1 H), 6.12 (m, 1 H), 6.47 (s, 1 H), 7.00 (m, 2 H), 7.54 (m, 4 H), 8.22 (m, 3 H); 13C (63 MHz, CDCl3) δ 9.6, 15.0, 18.6, 21.9, 22.4, 25.7, 26.6, 28.2, 35.6, 43.3, 45.7, 51.6, 55.9, 58.7, 72.2, 72.4, 73.8, 75.2, 75.6, 78.0, 79.3, 80.0, 81.1, 84.5, 112.1, 120.3, 120.63, 128.6, 129.2, 130.2, 132.7, 133.0, 133.7, 134.6, 160.0, 165.8, 167.0, 170.1, 170.5, 203.8. HRMS (FAB) m/z calcd for C49H61NO16·H+: 920.4069. Found: 920.4068 (Δ = 0.1 ppm). Other taxoids 6a and 6c–o were prepared in the same manner and characterization data are shown below.

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-benzoyldocetaxel (6a)

90% (2 steps), white solid; mp 140–143 °C; 1H NMR (250 MHz, CDCl3) δ 1.12 (s, 3 H), 1.22 (s, 3 H), 1.35 (s, 9 H), 1.58 (s, 3 H), 1.70 (s, 3 H), 1.76 (s, 3 H), 1.85 (m, 1 H), 1.96 (s, 3 H) 2.37 (s, 3 H), 2.60 (m, 2 H), 3.38 (m, 1 H), 3.89 (d, J= 7.0 Hz, 1 H), 4.21 (m, 2 H), 4.32 (d, J = 8.3 Hz, 1 H), 4.52 (m, 1 H), 4.76 (m, 2 H), 4.98 (d, J = 8.0 Hz, 1 H), 5.30 (m, 1 H), 5.71 (d, J = 7.0 Hz, 1 H), 6.21 (m, 1 H), 6.56 (s, 1 H), 7.47 (dd, J = 7.3 Hz, 7.8 Hz, 4 H), 7.58 (m, 2 H), 8.10 (m, 4 H); 13C (63 MHz, CDCl3) δ 9.6, 15.0, 18.6, 22.1, 22.4, 23.5, 25.7, 26.9, 28.2, 35.7, 42.3, 43.3, 45.8, 51.6, 58.7, 72.3, 72.4, 73.8, 75.1, 76.0, 79.2, 80.1, 81.1, 84.5, 120.6, 128.5, 128.7, 129.1, 129.2, 130.0, 130.2, 132.9, 133.7, 142.9, 166.4, 167.0, 170.1, 172.1, 203.6. HRMS (FAB) m/z calcd for C48H59NO15·H+: 890.3963. Found: 890.3966 (Δ = −0.3 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(3-methoxybenzoyl)docetaxel (6c)

87% (2 steps); white solid; mp 154–159 °C; [α]D20 −75 (c 0.010, CDCl3); 1H NMR (250 MHz, CDCl3) δ 1.20 (s, 3 H), 1.37 (s, 15 H), 1.54 (s, 3 H), 1.63 (s, 3 H), 1.82 (m, 1 H), 1.84 (s, 3 H), 2.36 (s, 3 H), 2.57 (m, 2 H), 3.39 (bs, 1 H), 3.89 (m, 4 H), 4.20 (m, 2 H), 4.31 (d, J = 8.1 Hz, 1 H), 4.48 (m, 1 H), 4.63 (m, 2 H), 4.98 (d, J = 8.7 Hz, 1 H), 5.25 (m, 1 H), 5.70 (d, J = 6.9 Hz, 1 H), 6.12 (m, 1 H), 6.54 (s, 1 H), 7.02 (m, 1 H), 7.44 (m, 6 H), 8.02 (m, 2 H); 13C (63 MHz, CDCl3) δ 9.6, 15.0, 18.6, 22.1, 22.4, 25.7, 26.8, 28.2, 33.4, 35.7, 43.3, 45.8, 51.6, 55.4, 58.7, 72.3, 73.8, 75.1, 76.1, 79.2, 80.0, 81.1, 84.5, 105.1, 114.1, 120.1, 120.6, 122.3, 128.7, 129.2, 129.6, 130.2, 130.4, 132.8, 133.7, 159.6, 166.3, 167.0, 169.7, 169.8, 170.1, 170.4, 203.5. HRMS (FAB) m/z calcd for C49H61NO16·H+: 920.4069. Found: 920.4068 (Δ = 0.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(4-methoxybenzoyl)docetaxel (6d)

80%; white solid; mp 148–156 °C; [α]D20 −45 (c 0.010, CDCl3); 1H NMR (250 MHz, CDCl3) δ 1.25 (s, 3 H), 1.35 (s, 9 H), 1.55 (s, 3 H), 1.70 (s, 6 H), 1.77 (s, 3 H), 1.82 (m, 1 H), 1.95 (s, 3 H), 2.37 (s, 3 H), 2.57 (m, 2 H), 3.39 (bs, 1 H), 3.88 (m, 4 H), 4.21 (m, 2 H), 4.31 (d, J= 8.1 Hz, 1 H), 4.55 (m, 1 H), 4.76 (m, 2 H), 4.98 (d, J = 8.7 Hz, 1 H), 5.23 (m, 1 H), 5.75 (d, J = 6.6 Hz, 1 H), 6.21 (m, 1 H), 6.54 (s, 1 H), 6.94 (d, J = 6.5 Hz, 2 H), 7.48 (m, 3 H), 8.03 (d, J = 9 Hz, 2 H), 8.11 (d, J = 6.5 Hz, 2 H); 13C (63 MHz, CDCl3) δ 9.6, 15.0, 18.6, 22.1, 22.4, 25.7, 26.8, 28.2, 33.4, 35.7, 43.3, 45.8, 51.6, 55.4, 58.7, 72.3, 73.8, 75.1, 76.1, 79.2, 80.0, 81.1, 84.5, 105.1, 114.1, 120.1, 120.6, 121.3, 128.7, 129.2, 129.6, 130.2, 131.4, 132.8, 133.7, 159.6, 166.3, 167.4, 169.8, 170.0, 170.1, 170.7, 203.6. HRMS (FAB) m/z calcd for C49H61NO16·H+: 920.4069. Found: 920.4068 (Δ = 0.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(3,4-dimethoxybenzoyl)docetaxel (6e)

92% (for 2 steps); white solid; mp 159–165 °C; 1H NMR (250 MHz, CDCl3) δ 1.25 (s, 6 H), 1.33 (s, 12 H), 1.70 (s, 3 H), 1.76 (s, 3 H), 1.95 (m, 4 H), 2.42 (s, 3 H), 2.66 (m, 2 H), 3.37 (bs, 1 H), 3.88 (m, 1 H), 3.93 (s, 3 H), 3.95 (s, 3 H), 4.18 (m, 2 H), 4.32 (d, J = 8.5 Hz, 1 H), 4.51 (m, 1 H), 4.76 (m, 2 H), 4.98 (d, J = 8.0 Hz, 1 H), 5.31 (m, 1 H), 5.71 (d, J = 6.8 Hz, 1 H), 6.21 (t, J = 8.0 Hz, 1 H), 6.53 (s, 1 H), 6.92 (d, J = 8.5 Hz, 1 H), 7.44 (dd, J = 7.3 Hz, 8.0 Hz, 2 H), 7.57 (m, 2 H), 7.73 (d, J = 8.5 Hz, 1 H), 8.11 (d, J = 7.0 Hz, 2 H); 13C (63 MHz, CDCl3) δ 9.5, 14.2, 15.0, 18.6, 22.1, 22.4, 25.7, 26.9, 28.2, 29.7, 35.6, 43.3, 45.8, 51.6, 56.0, 56.1, 58.7, 72.3, 73.8, 75.1, 75.9, 79.2, 80.0, 81.1, 84.5, 110.4, 112.4, 120.6, 121.5, 124.1, 128.7, 129.2, 130.2, 132.9, 133.7, 138.0, 142.8, 148.8, 153.7, 155.5, 166.2, 167.0, 170.1, 173.1, 203.8. HRMS (FAB) m/z calcd for C50H63NO17·H+: 950.4174. Found: 950.4174 (Δ = 0.0 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(1-naphthoyl)docetaxel (6f)

90% (2 steps); white solid; mp 162–164 °C; 1H NMR (250 MHz, CDCl3) δ 1.21 (s, 3 H), 1.28 (s, 3 H), 1.35 (s, 9 H), 1.73 (s, 3 H), 1.77 (s, 6 H), 1.93 (m, 1 H), 2.00 (s, 3 H), 2.38 (s, 3 H), 2.68 (m, 2 H), 3.37 (bs, 1 H), 3.92 (d, J = 7.0 Hz, 1 H), 4.23 (m, 2 H), 4.33 (d, J = 8.3 Hz, 1 H), 4.58 (m, 1 H), 4.77 (m, 2 H), 5.00 (d, J = 8.0 Hz, 1 H), 5.33 (m, 1 H), 5.73 (d, J = 7.0 Hz, 1 H), 6.23 (t, J = 7.0 Hz, 1 H), 6.69 (s, 1 H), 7.57 (m, 6 H), 7.90 (d, J = 7.5 Hz, 1 H), 8.10 (m, 3 H), 8.32 (d, J = 6.5 Hz, 1 H), 8.89 (d, J = 8.3 Hz, 1 H); 13C (63 MHz, CDCl3) δ 9.6, 15.1, 15.6, 18.6, 22.2, 22.4, 25.7, 26.8, 28.2, 35.7, 43.3, 45.8, 51.6, 58.7, 72.3, 72.4, 73.8, 75.1, 76.0, 79.2, 80.0, 81.1, 84.5, 120.6, 124.5, 125.7, 126.1, 126.4, 128.0, 128.6, 128.7, 129.2, 130.2, 130.8, 131.4, 133.0, 133.7, 134.1 138.0, 143.0, 155.5, 167.0, 167.5, 170.2, 173.0, 203.9. HRMS (FAB) m/z calcd for C52H61NO15·H+: 940.4128. Found: 940.4124 (Δ = 0.4 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(2-napthoyl)docetaxel (6g)

92%; white solid; mp 165–170 °C; 1H NMR (250 MHz, CDCl3) δ 1.32 (s, 3 H), 1.35 (s, 9 H), 1.39 (s, 3 H), 1.57 (s, 3 H), 1.72 (s, 3 H), 1.76 (s, 3 H), 1.95 (m, 1 H), 1.98 (s, 3 H), 2.38 (s, 3 H), 2.50 (m, 2 H), 3.39 (bs, 1 H), 3.92 (d, J = 7.0 Hz, 1 H), 4.23 (m, 2 H), 4.31 (d, J = 8.3 Hz, 1 H), 4.55 (m, 1 H), 4.77 (m, 2 H), 4.99 (d, J = 8.8 Hz, 1 H), 5.25 (m, 1 H), 5.73 (d, J = 6.8 Hz, 1 H), 6.23 (m, 1 H), 6.63 (s, 1 H), 7.51 (m, 5 H), 7.99 (m, 6 H), 8.65 (s, 1 H); 13C (63 MHz, CDCl3) δ 9.6, 14.2, 15.1, 18.6, 22.2, 22.4, 25.7, 26.9, 28.2, 35.7, 43.3, 45.8, 51.6, 58.7, 72.3, 73.4, 73.8, 75.1, 76.2, 79.2, 80.0, 81.1, 84.5, 120.6, 125.2, 126.3, 126.8, 127.8, 128.4, 128.6, 129.2, 129.5, 130.2, 131.8, 132.4, 132.9, 133.7, 135.9, 138.0, 142.9, 155.5, 166.6, 167.0, 170.1, 203.6. HRMS (FAB) m/z calcd for C52H61NO15·H+: 940.4128. Found: 940.4124 (Δ = 0.4 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-benzyloxycarbonyldocetaxel (6h)

97%, white solid; mp 145–150 °C; 1H NMR (300 MHz. CDCl3) δ 1.15 (s, 3 H), 1.23 (s, 3 H), 1.34 (s, 9 H), 1.60 (bs, 1 H), 1.70 (s, 3 H), 1.76 (s, 6 H), 1.85 (m, 1 H), 1.89 (s, 3 H), 2.36 (s, 3 H), 2.54 (m, 2 H), 3.36 (bs, 1 H), 4.19 (m, 2 H), 4.30 (d, J = 8.7 Hz, 1 H), 4.40 (m, 1 H), 4.76 (m, 2 H), 4.96 (d, J = 7.8 Hz, 1 H), 5.23 (s, 2 H), 5.31 (d, J = 7.5 Hz, 1 H), 5.67 (d, J = 6.9 Hz, 1 H), 6.16 (m, 2 H), 7.40 (m, 7 H), 7.61 (dd, J = 7.2 Hz, 7.5 Hz, 1 H), 8.10 (d, J = 7.5 Hz, 2 H); 13C NMR (63 MHz. CDCl3) δ 9.5, 15.1, 18.5, 21.8, 22.4, 25.7, 26.5, 28.2, 35.5, 35.6, 43.1, 45.6, 51.6, 58.2, 58.6, 70.6, 72.1, 72.3, 73.8, 75.0, 78.4, 79.2, 80.0, 81.0, 84.4, 120.6, 128.5, 128.6, 128.7, 129.2, 130.1, 132.5, 133.7, 134.7, 137.9, 143.5, 155.3, 155.5, 166.9, 170.1, 173.0, 203.9. HRMS (FAB) m/z calcd for C49H61NO16·H+: 920.4069. Found: 920.4068 (Δ = 0.1 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(2-methoxyphenylacetyl)docetaxel (6i)

80% (for 2 steps); white solid; mp 148–149 °C; 1H NMR (250 MHz. CDCl3) δ 1.04 (s, 3 H), 1.13 (s, 3 H), 1.35 (s, 9 H), 1.58 (s, 3 H), 1.66 (s, 3 H), 1.76 (s, 3 H), 1.88 (s, 4 H), 2.34 (s, 3 H), 2.39 (m, 2 H), 2.50 (m, 1 H), 3.39 (bs, 1 H), 3.81 (m, 4 H), 4.19 (m, 2 H), 4.32 (m, 2 H), 4.75 (m, 2 H), 4.94 (d, J = 7.5 Hz, 1 H), 5.29 (m, 1 H), 5.64 (d, J = 6.8 Hz, 1 H), 6.14 (t, J = 7.5 Hz, 1 H), 6.29 (s, 1 H), 6.92 (m, 2 H), 7.27 (m, 2 H), 7.46 (m, 2 H), 7.56 (m, 1 H), 8.09 (d, J = 7.0 Hz, 2 H); 13C NMR (63 MHz, CDCl3) δ 9.5, 14.2, 15.1, 18.5, 21.6, 22.4, 25.7, 26.4, 28.2, 30.9, 35.6, 40.1, 43.1, 45.8, 51.5, 55.7, 58.8, 61.2, 72.0, 72.3, 73.7, 74.9, 75.0, 75.8, 79.1, 81.0, 84.4, 87.5, 114.0, 114.6, 120.1, 120.6, 122.5, 128.6, 129.6, 130.4, 130.5, 132.7, 142.5, 159.6, 169.7, 170.5, 172.9, 203.4. HRMS (FAB) m/z calcd for C50H63NO16·H+: 934.4225. Found: 934.4229 (Δ = −0.4 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(3-methoxyphenylacetyl)docetaxel (6j)

80% (for 2 steps); white solid; mp 145–150 °C; 1H NMR (250 MHz. CDCl3) δ 1.07 (s, 3 H), 1.18 (s, 3 H), 1.34 (s, 9 H), 1.59 (s, 3 H), 1.67 (s, 3 H), 1.73 (s, 3 H), 1.76 (s, 3 H) 1.86 (s, 3 H), 1.86 (m, 1 H), 2.17 (m, 4 H), 2.37 (s, 3 H), 2.39 (m, 2 H), 2.55 (m, 1 H), 3.34 (bs, 1 H), 3.79 (m, 4 H), 4.19 (m, 1 H), 4.32 (d, J = 8.4 Hz, 1 H), 4.39 (m, 1 H), 4.73 (m, 2 H), 4.94 (d, J = 8.1 Hz, 1 H), 5.30 (m, 1 H), 5.64 (d, J = 6.8 Hz, 1 H), 6.13 (t, J = 8.1 Hz, 1 H), 6.30 (s, 1 H), 6.88 (m, 2 H), 7.27 (m, 2 H), 7.46 (m, 2 H), 7.60 (m, 1 H), 8.09 (d, J = 7.3 Hz, 2 H); 13C NMR (250 MHz, CDCl3) δ 9.5, 14.5, 15.1, 18.4, 21.6, 22.4, 25.7, 26.4, 28.2, 30.9, 35.6, 40.1, 43.1, 45.6, 51.5, 55.7, 58.8, 61.2, 72.0, 72.3, 73.7, 74.9, 75.0, 75.8, 79.1, 81.0, 84.4, 87.5, 114.0, 114.6, 120.1, 120.6, 122.7, 128.6, 129.6, 130.2, 130.5, 132.5, 142.5, 159.6, 168.9, 170.5, 172.9, 203.6. HRMS (FAB) m/z calcd for C50H63NO16·H+: 934.4225. Found: 934.4229 (Δ = −0.4 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-(4-methoxyphenylacetyl)docetaxel (6k)

60% (for 2 steps); white solid; mp 145–150 °C; 1H NMR (250 MHz. CDCl3) δ 1.07 (s, 3 H), 1.18 (s, 3 H), 1.34 (s, 9 H), 1.59 (s, 3 H), 1.67 (s, 3 H), 1.73 (s, 3 H), 1.76 (s, 3 H) 1.86 (s, 3 H), 1.86 (m, 1 H), 2.17 (m, 4 H), 2.37 (s, 3 H), 2.39 (m, 2 H), 2.55 (m, 1 H), 3.34 (bs, 1 H), 3.79 (m, 4 H), 4.19 (m, 1 H), 4.32 (d, J = 8.4 Hz, 1 H), 4.39 (m, 1 H), 4.73 (m, 2 H), 4.94 (d, J = 8.1 Hz, 1 H), 5.30 (m, 1 H), 5.64 (d, J = 6.9 Hz, 1 H), 6.15 (t, J = 8.1 Hz, 1 H), 6.30 (s, 1 H), 7.03 (d, J = 9.0 Hz, 2 H), 7.45 (m, 2 H), 7.58 (m, 1 H), 8.10 (d, J = 7.3 Hz, 2 H), 8.26 (d, J = 9.0 Hz, 2 H); 13C NMR (63 MHz, CDCl3) δ 9.5, 14.2, 15.1, 18.5, 21.6, 22.4, 25.7, 26.4, 28.2, 30.9, 35.6, 40.1, 43.1, 45.8, 51.5, 55.7, 58.8, 61.2, 72.0, 72.3, 73.7, 74.9, 75.0, 75.8, 79.1, 81.0, 84.4, 87.5, 114.0, 114.6, 120.1, 120.6, 122.5, 128.6, 129.6, 130.4, 130.5, 132.7, 142.5, 159.6, 169.7, 170.5, 172.9, 203.4. HRMS (FAB) m/z calcd for C50H63NO16·H+: 934.4225. Found: 934.4229 (Δ = −0.4 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-hydrocinnamoyldocetaxel(6l)

85% (for 2 steps); white solid; mp 150–155 °C; 1H NMR (250 MHz, CDCl3) δ 1.13 (s, 3 H), 1.21 (s, 3 H), 1.56 (s, 12 H), 1.68 (s, 3 H), 1.72 (s, 3 H), 1.86 (s, 3 H), 1.88 (m, 1 H), 2.16 (s, 3 H), 2.38 (m, 2 H), 2.85 (m, 2 H), 3.20 (m, 2 H), 3.35 (bs, 1 H), 3.7m (m, 1 H), 4.20 (m, 2 H), 4.34 (d, J = 8.3 Hz, 1 H), 4.45 (m, 1 H), 4.77 (m, 2 H), 4.94 (d, J = 7.5 Hz, 1 H), 5.31 (bs, 1 H), 5.66 (d, J = 7.0 Hz, 1 H), 6.27 (m, 2 H), 7.17 (m, 5 H), 7.47 (dd, J = 7.8 Hz, 8.3 Hz, 2 H), 7.60 (m, 1 H), 8.09 (d, J = 7.8 Hz, 2 H); 13C (63 MHz, CDCl3) δ 9.5, 15.0, 18.6, 22.4, 25.7, 26.6, 28.2, 30.0, 35.6, 36.1, 43.2, 45.6, 51.6, 55.3, 58.6, 72.2, 72.3, 73.8, 75.0, 75.5, 79.2, 81.1, 113.9, 117.6, 120.6, 128.6, 129.2, 130.2, 132.2, 133.7, 138.0, 142.6, 158.2, 167.0, 170.1, 173.1, 173.6, 203.8. HRMS (FAB) m/z calcd for C50H63NO15·H+: 918.4276. Found: 918.4276 (Δ = 0.0 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-[3-(2-methoxyphenyl)propanoyl]docetaxel (6m)

97%; white solid; mp 155–159 °C; [α]D20 −45 (c 0.050, CDCl3); 1H NMR (250 MHz, CDCl3) δ 1.13 (s, 3 H), 1.24 (s, 3 H), 1.35 (s, 9 H), 1.67 (s, 3 H), 1.75 (s, 3 H), 1.88 (s, 4 H), 2.01 (s, 3 H), 2.35 (s, 3 H), 2.48 (m, 2 H), 2.79 (m, 2 H), 2.94 (m, 2 H), 3.33 (m, 1 H), 3.82 (m, 4 H), 4.20 (m, 2 H), 4.30 (d, J = 8.5 Hz, 1 H), 4.38 (m, 1 H), 4.75 (m, 2 H), 4.96 (d, J = 8.0 Hz, 1 H), 5.31 (m, 1 H), 5.64 (d, J = 7.0 Hz, 1 H), 6.19 (m, 1 H), 6.29 (s, 1 H), 6.82 (m, 2 H), 7.19 (m, 2 H), 7.45 (m, 2 H), 7.59 (m, 1 H), 8.10 (d, J = 7.3 Hz 2 H); 13C (63 MHz, CDCl3) δ 9.5, 15.0, 18.6, 21.9, 22.4, 25.7, 26.0, 26.6, 28.2, 30.9, 31.6, 33.4, 34.0, 35.6, 43.2, 45.7, 51.6, 55.2, 58.6, 72.2, 72.4, 73.8, 75.1, 75.4, 79.2, 80.0, 81.1, 84.4, 87.3, 105.1, 110.2, 120.4, 120.6, 127.7, 128.5, 128.6, 129.2, 130.0, 130.2, 132.9, 133.7, 137.9, 142.5, 155.4, 157.5, 167.0, 170.1, 173.5, 203.8. HRMS (FAB) m/z calcd for C51H65NO16·H+: 948.4382. Found: 948.4382 (Δ = 0.0 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-[3-(3-methoxyphenyl)propanoyl]docetaxel (6n)

80%; white solid; mp 159–164 °C; 1H NMR (250 MHz, CDCl3) δ 1.12 (s, 3 H), 1.23 (s, 3 H), 1.35 (s, 9 H), 1.55 (s, 3 H), 1.60 (s, 3 H), 1.75 (s, 3 H), 1.87 (s, 3 H), 1.88 (m, 1 H), 2.35 (s, 3 H), 2.49 (m, 2 H), 2.81 (m, 2 H), 2.94 (m, 2 H), 3.34 (m, 1 H), 3.79 (s, 3 H), 3.81 (m, 1 H), 4.20 (m, 2 H), 4.29 (d, J = 8.3 Hz, 1 H), 4.38 (m, 1 H), 4.75 (m, 2 H), 4.92 (d, J = 8.0 Hz, 1 H), 5.30 (bs, 1 H), 5.66 (d, J = 7.0 Hz, 1 H), 6.19 (m, 1 H), 6.29 (s, 1 H), 6.79 (m, 3 H), 7.20 (m, 1 H), 7.47 (m, 2 H), 7.53 (m, 1 H), 8.15 (d, J = 7.5 Hz 2 H); 13C (63 MHz, CDCl3) δ 9.5, 14.9, 18.6, 21.9, 22.4, 25.7, 26.6, 28.2, 30.9, 33.4, 35.6, 35.7, 36.3, 43.2, 45.6, 51.6, 55.2, 58.6, 72.2, 73.7, 75.0, 75.6, 79.2, 81.1, 84.4, 105.2, 111.8, 114.0, 120.6, 128.6, 129.2, 129.5, 130.2, 132.8, 133.7, 141.8, 142.7, 166.9, 169.7, 170.1, 171.3, 173.0, 203.8. HRMS (FAB) m/z calcd for C51H65NO16·H+: 948.4382. Found: 948.4382 (Δ = 0.0 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-10-[3-(4-methoxyphenyl)propanoyl]docetaxel (6o)

80%; white solid; mp 169–172 °C; 1H NMR (250 MHz, CDC13) δ 1.12 (s, 3 H), 1.23 (s, 3 H), 1.35 (s, 9 H), 1.68 (s, 3 H), 1.76 (s, 3 H), 1.86 (s, 6 H), 1.88 (m, 1 H), 2.35 (s, 3 H), 2.48 (m, 2 H), 2.79 (m, 2 H), 2.94 (m, 2 H), 3.33 (d, J = 6.5 Hz, 1 H), 3.78 (s, 3 H), 3.81 (m, 1 H), 4.20 (m, 2 H), 4.30 (d, J = 8.5 Hz, 1 H), 4.42 (m, 1 H), 4.75 (m, 2 H), 4.96 (d, J = 8.0 Hz, 1 H), 5.31 (bs, 1 H), 5.66 (d, J = 7.0 Hz, 1 H), 6.17 (m, 1 H), 6.29 (s, 1 H), 6.82 (d, J = 8.5 Hz, 2 H), 7.13 (d, J = 8.5 Hz, 2 H), 7.47 (dd, J = 7.3 Hz, 7.8 Hz, 2 H), 7.60 (m, 1 H), 8.10 (d, J= 7.3 Hz 2 H); 13C (63 MHz, CDC13) δ 9.5, 14.9, 18.6, 22.4, 25.7, 26.6, 28.2, 30.0, 35.6, 36.1, 43.2, 45.6, 51.6, 55.3, 58.6, 72.2, 72.3, 73.8, 75.0, 75.5, 79.2, 81.1, 84.4, 113.9, 117.6, 120.6, 128.6, 129.2, 130.2, 132.2, 132.8, 133.7, 138.0, 142.6, 158.2, 167.0, 170.1, 173.1, 173.6, 203.8. HRMS (FAB) m/z calcd for C5iH65NO16·H+: 948.4382. Found: 948.4382 (Δ = 0.0 ppm).

Preparation of 7,10,13-tri-TES-2-modified baccatins (9)

A typical procedure is described for the preparation of 7,10,13-tri(triethylsilyl)-2-debenzoyl-2-(3-fluorobenzoyl)-10-deacetylbaccatin III (9c): To a solution of baccatin 8 (200 mg, 0.255 mmol), 3-fluorobenzoic acid (179 mg, 1.28 mmol) and DMAP (31 mg, 0.255 mmol) in 4 mL dichloromethane was added 1,3-diisopropylcarbodiimide (DIC) (193.1 mg, 1.53 mmol) and the mixture was stirred at 40 °C for 40 h. The resulting precipitate was filtered off and washed with ethyl acetate (30 mL). The filtrate was washed with saturated sodium bicarbonate solution (10 mL × 2), dried over anhydrous magnesium sulfate, and concentrated in vacuo. Column chromatography of the residue on silica gel using hexanes/ethyl acetate (7/1) as eluant gave 9c as a white solid (183 mg, 79%): mp 93–95 °C; 1H NMR (CDCl3) δ 0.60 (m, 18 H), 0.98 (m, 27 H), 1.09 (s, 3 H), 1.16 (s, 3 H), 1.57 (bs, 1 H), 1.62 (s, 3 H), 1.85 (m, 1 H), 1.96 (s, 3 H), 2.14 (m, 2 H), 2.26 (s, 3 H), 2.50 (m, 1 H), 3.83 (d, J = 6.9 Hz, 1 H), 4.10 (d, J = 8.1 Hz, 1 H), 4.24 (d, J = 8.1 Hz, 1 H), 4.39 (dd, J = 10.4, 6.9 Hz, 1 H), 4.92 (m, 2 H), 5.17 (s, 1 H), 5.56 (d, J = 7.0 Hz, 1 H), 7.29 (m, 1 H), 7.43 (dd, J = 13.6, 8.0 Hz, 1 H), 7.76 (d, J = 9.2 Hz, 1 H), 7.87 (d, J = 7.7 Hz, 1 H); 13C NMR (CDCl3) δ 4.8, 5.2, 5.9, 6.9, 10.4, 14.6, 20.6, 22.3, 26.3, 37.2, 39.8, 42.9, 46.9, 58.2, 68.2, 72.6, 75.7, 75.9, 76.5, 79.6, 80.7, 84.0, 116.6, 117.0, 120.3, 120.7, 125.6, 130.1, 130.3, 131.7, 131.9, 132.4, 135.6, 139.5, 160.6, 164.5, 165.6, 169.9, 205.6. HRMS (FAB, DCM/NBA) mlz calcd for C47H77O10SiF·H+ 905.4887. Found: 905.4869 (Δ = 2.0 ppm).

Other 7,10,13-tri-TES-2-modified baccatins 9a, 9b and 9d–f were prepared in the same manner and characterization data are summarized in the Supporting Information.

Preparation of 7-TES-2-(3-substituted-benzoyl)-10-acylbaccatins 11

A typical procedure is described for the preparation of 7-triethylsilyl-2-debenzoyl-2-(3-fluorobenzoyl)-10-deacetyl-10-propanoylbaccatin III (11c): (a) To a solution of 9c (181 mg, 0.200 mmol) in 10 mL of pyridine/acetonitrile (1:1) was added dropwise HF/pyridine (70:30, 1.5 mL) at 0 °C, and the mixture was stirred at room temperature for 17 h. To the reaction mixture was added ethyl acetate (100 mL) and saturated aqueous sodium carbonate (15 mL). The organic layer was separated and washed with saturated aqueous copper sulfate (10 mL × 3) and water (10 mL), dried over anhydrous magnesium sulfate, and concentrated in vacuo to afford 2-(3-fluorobenzoyl)-2-debenzoyl-10-deacetylbaccatin III (7-OH-10c) as a white solid (106 mg, 94%).

(b) To a solution of baccatin 7-OH-10c thus obtained (105 mg, 0.187 mmol) and imidazole (54 mg, 0.789 mmol) in dry DMF (3.1 mL) was added chlorotriethylsilane (0.10 mL, 0.592 mmol) dropwise at room temperature. The reaction mixture was stirred at room temperature for 2 h and diluted with ethyl acetate (60 mL). The reaction mixture was then washed with water (5 mL × 3), brine (10 mL), dried over MgSO4 and concentrated in vacuo. Column chromatography of the residue on silica gel (hexanes/ethyl acetate = 2/1 to 1/1) gave 7-TES-2-(3-fluorobenzoyl)-2-debenzoyl-10-deacetylbaccatin III (10c) as a white solid (105 mg, 84% yield): mp 118–120 °C; 1H NMR (CDC13) δ 0.55 (m, 6 H), 0.92 (m, 9 H), 1.06 (s, 6 H), 1.61 (bs, 1 H), 1.71 (s, 3 H), 1.89 (m, 1 H), 2.06 (s, 3 H), 2.21 (s, 1 H), 2.25 (s, 1 H), 2.27 (s, 3 H), 2.47 (m, 1 H), 3.93 (d, J = 7.0 Hz, 1 H), 4.13 (d, J = 7.1 Hz, 1 H), 4.28 (m, 2 H), 4.39 (dd, J = 10.5, 6.6 Hz, 1 H), 4.84 (t, J = 7.8 Hz, 1 H), 4.95 (d, J = 8.3 Hz, 1 H), 5.16 (s, 1 H), 5.55 (d, J = 7.0 Hz, 1 H), 7.29 (m, 1 H), 7.44 (dd, J = 13.6, 7.8 Hz, 1 H), 7.77 (d, J = 8.9 Hz, 1 H), 7.88 (d, J = 7.7 Hz, 1 H); 13C NMR (CDCl3) δ 5.1, 6.7, 9.9, 15.2, 18.5, 22.5, 28.8, 37.2, 38.5, 42.6, 46.9, 57.9, 67.8, 72.9, 74.5, 75.2, 76.4, 78.8, 80.6, 84.2, 116.7, 117.0, 120.5, 120.8, 125.8, 125.9, 130.2, 130.3, 134.9, 160.5, 164.5, 165.8, 170.7,210.2. HRMS (FAB, DCM/NBA) m/z calcd for C35H49O10FSi·H+: 677.3157. Found: 677.3177 (Δ = −2.9 ppm).

(c) To a solution of 10c (104 mg, 0.154 mmol) in dry THF (11 mL) was added LiHMDS (1.0 M in THF, 0.17 mL, 0.17 mmol) dropwise at −40 °C. The mixture was stirred at −40 °C for 5 min. Then freshly distilled chlorotriethylsilane (0.016L, 0.1844 mmol) was added dropwise and the reaction mixture was stirred for 30 min. Then, to the reaction mixture was added aqueous saturated NH4C1 (10 mL) and ethyl acetate (100 mL), and the organic layer was separated and washed with brine, dried over anhydrous magnesium sulfate and concentrated in vacuo. Column chromatography of the residue on silica gel (hexanes/ethyl acetate = 3/1) gave 11c as a white solid (95 mg, 84%): mp 194–196 °C; [α]D20 −74 (c 0.39, CHCl3); 1H NMR (CDCl3) δ 0.56 (q, J = 7.8 Hz, 6 H), 0.89 (t, J = 7.8 Hz, 9 H), 1.00 (s, 3 H), 1.20 (m, 7 H), 1.64 (s, 3 H), 1.83 (m, 1 H), 2.16 (s, 3 H), 2.25 (m, 3 H), 2.40 (m, 4 H), 3.85 (d, J = 6.8 Hz, 1 H), 4.09 (d, J = 8.2 Hz, 1 H), 4.25 (d, J = 8.2 Hz, 1 H), 4.45 (dd, J = 10.2, 6.7 Hz, 1 H), 4.79 (t, J = 8.0 Hz, 1 H), 4.93 (d, J = 9.4 Hz, 1 H), 5.57 (d, J = 7.1 Hz, 1 H), 6.45 (s, 1 H), 7.29 (m, 1 H), 7.44 (dd, J = 13.6, 8.0 Hz, 1 H), 7.74 (d, J = 8.7 Hz, 1 H), 7.86 (d, J = 7.7 Hz, 1 H); 13C NMR (CDCl3) δ 5.3, 6.7, 6.94, 9.2, 9.9, 14.9, 20.1, 22.5, 26.8, 27.7, 37.2, 38.3, 42.7, 47.2, 58.5, 67.8, 72.3, 75.2, 75.5, 76.4, 78.8, 80.7, 84.2, 116.7, 117.0, 120.5, 120.8, 125.9, 130.2, 130.3, 131.6, 131.7, 132.6, 144.1, 165.8, 170.6, 172.8, 202.3. HRMS (FAB)m/z calcd for C38H53O11FSi·H+: 733.3419. Found: 733.3418 (Δ = + 0.2 ppm).

Other 7-TES-2-(3-substituted-benzoyl)-10-acylbaccatins (11a, 11b and 11d–k) were prepared in the same manner and characterization data are summarized in the Supporting Information.

Preparation of 2-debenzoyl-2-(3-methoxybenzoyl)-7-triethylsilyl-10-deacetyl-10-(2-methoxybenzoyl)baccatin III (11k)

(a) To a solution of 104 mg (0.16 mmol) of 2-debenzoyl-2-(3-methoxybenzoyl)-7-triethylsilyl-10-deacetyl baccatin III (10b) in 2 mL of CH2Cl2 was added 21 mg (0.17 mmol) of N-methylmorpholine-N-oxide (NMO) and 20 mg of 4 Å molecular sieves. After the mixture was stirred for 10 min, 37 mg (0.01 mmol) of tetrapropylammonium perruthenate (TPAP) was added and the mixture was allowed to stir for 4 h. The reaction mixture was then filtered and concentrated in vacuo to give 2-debenzoyl-2-(3-methoxybenzoyl)-7-triethylsilyl-10-deacetyl-13-oxo-baccatin III (12) (104 mg, 99% yield) as a white solid: 1H (300 MHz, CDC13) δ 0.49 (m, 6 H), 0.95 (m, 9 H), 1.15 (s, 3 H), 1.22 (s, 3 H), 1.24 (m, 1H), 1.71 (s, 3 H), 1.84 (m, 2 H), 2.10 (s, 3 H), 2.17 (s, 3 H), 2.47 (m, 1 H), 2.60 (d, J = 19.8 Hz, 1 H), 2.89 (d, J = 19.8 Hz, 1H), 3.85 (m, 6 H), 4.11 (d, J = 8.7 Hz, 1 H), 4.35 (m, 4 H), 4.90 (d, J = 8.1Hz, 1H), 5.31 (d, J = 1.8 Hz, 1 H), 5.62 (d, J = 6.6 Hz, 1 H), 7.13 (dd, J = 8.4 Hz, 2.7 Hz, 1 H), 7.38 (t, J = 5.4 Hz, 1 H), 7.58 (s, 3 H), 7.64 (d, J = 7.5 Hz, 1 H). HRMS: m/e calcd for C44H58O13Si·H+: 823.3725. Found: 823.3723 (Δ = −0.2 ppm).

(b) To a solution of 104 mg (0.159 mmol) of 12, DMAP (59 mg, 0.477 mmol) and triethylamine (48 mg, 0.477 mmol) in 2 mL of CH2Cl2 was added 2-methoxylbenzoyl chloride (81 mg, 0.477 mmol). The mixture was allowed to stir overnight and the reaction was quenched with saturated aqueous NaHCO3, and extracted 3 times with EtOAc. The combined organic layers were washed with water and brine, dried over MgSO4, filtered, and concentrated. The resulting solid was purified by column chromatography on silica gel (hexanes/EtOAc = 8/1) to give 2-debenzoyl-2-(3-methoxybenzoyl)-7-triethylsilyl-10-deacetyl-10-(2-methoxybenzoyl)-13-oxo-baccatin III (13) (125 mg, 100%) as a white solid: 1H (300 MHz, CDC13) δ 0.58 (m, 6 H), 0.91 (q, J = 7.8 Hz, 9 H), 1.10 (s, 3 H), 1.12 (s, 3 H), 1.37 (s, 3 H), 1.92 (m, 1 H), 1.97 (bs, 1 H), 2.19 (s, 3 H), 2.30 (s, 3 H), 2.55 (m, 1 H), 2.64 (d, J = 19.8 Hz, 1 H), 2.93 (s, J= 19.8 Hz, 1 H), 3.54 (m, 3 H), 3.85 (m, 8 H), 3.97 (d, J = 6.6 Hz, 1 H), 4.13 (d, J= 8.1 Hz, 1 H), 4.35 (d, J = 8.7 Hz, 1 H), 4.54 (dd, J = 6.6 Hz, 3.9 Hz, 1 H), 4.93 (d, J = 8.1 Hz, 1 H), 5.73 (d, J = 6.6 Hz, 1 H), 6.82 (s, 1 H), 6.66–7.07 (m, 3 H), 7.14–7.19 (m, 2 H), 7.28–7.42 (m, 1 H), 7.48–7.68 (m, 1 H), 8.01 (m, 1 H).

(c) To a solution of 13 (125 mg, 0.159 mmol) in 6 mL MeOH/THF (3/2) at 0 °C was added NaBH4 (100 mg, 6.36 mmol) and the solution was allowed to stir for 5 h. The reation was quenched with saturated NH4C1, and extracted 3 times with CH2Cl2. The organic layer was dried over MgSO4, filtered, and concentrated. The resulting solid was purified by column chromatography on silica gel (hex:EtOAc = 3:1) to give 10k (60 mg, 40% yield; 80% yield based on 50% conversion) as a white solid: 1H (300 MHz, CDCl3) δ 0.58 (m, 6H), 0.91 (q, J = 7.8 Hz, 9 H), 1.10 (s, 3 H), 1.12 (s, 3 H), 1.37 (s, 3 H), 1.92 (m, 2 H), 1.97 (bs, 1 H), 2.19 (s, 3 H), 2.30 (s, 3 H), 2.55 (m, 1 H), 3.54 (m, 1 H), 3.85 (m, 7 H), 3.94 (d, J = 6.9 Hz, 1 H), 4.15 (d, J = 7.8 Hz, 1 H), 4.34 (d, J = 7.8 Hz, 1 H), 4.53 (dd, J = 6.6 Hz, 3.6 Hz, 1 H), 4.84 (m, 1H), 4.97 (d, J = 7.8 Hz, 1 H), 5.66 (d, J = 6.9 Hz, 1 H), 6.69 (s, 1 H), 6.95–7.07 (m, 2 H), 7.14–7.19 (m, 1 H), 7.28–7.42 (m, 2 H), 7.65–7.73 (m, 2 H), 8.01 (m, 1 H).

Synthesis of second- and third-generation taxoids 14 through the Ojima-Holton coupling of baccatins 11 with β-lactams 3a–f

A typical procedure is described for the synthesis of 2-debenzoyl-2-(3-methylbenzoyl)-10-acetyl-3'-dephenyl-3'-(2-methylprop-1-enyl)docetaxel (14a): (a) To a solution of baccatin 11a (50 mg, 0.071 mmol) and (β-lactam 3a (41.1 mg, 0.099 mmol) in 3 mL dry THF was added a 1.0 M solution of LiHMDS in THF (0.099 mL, 0.099 mmol) dropwise at −40 °C, and the solution was stirred at the same temperature for 30 min. The reaction was quenched with aqueous saturated ammonium chloride, and the aqueous layer was extracted with dichloromethane. The combined extracts were dried over anhydrous magnesium sulfate and concentrated in vacuo. The residue was purified on a silica gel column (hexanes/EtOAc = 6/1) to afford of the coupling product, 7-TES-2’-TIPS-14a (67 mg, 84% yield), as a white solid.

(b) To a solution of 7-TES-2’-TIPS-14a (65.0 mg, 0.058 mmol), thus obtained, in 4 mL pyridine/acetonitrile (1/1) was added dropwise HF/pyridine (70/30, 0.5 mL) at 0 °C. The mixture was stirred at room temperature overnight. The reaction was quenched with saturated aqueous NaHCO3. The reaction mixture was diluted with EtOAc, washed with aqueous saturated CuSO4 and water, dried over anhydrous MgSO4 and concentrated in vacuo. The crude product was purified by column chromatography on silica gel (hexanes/EtOAc = 1/1) to afford 14a (47 mg, 96% yield; 81% yield for 2 steps) as a white solid: White solid; mp 145–148 °C; [α]D20 − 85 (c 0.011, CHC13); 1H NMR (300 MHz, CDC13) δ 1.14 (s, 3 H), 1.25 (s, 3 H), 1.34 (s, 9 H), 1.67 (s, 3 H), 1.75 (d, J = 3.6 Hz, 6 H), 1.89 (s, 3 H), 2.23 (s, 3 H), 2.37 (m, 9 H), 2.52 (m, 2 H), 3.80 (d, J = 7.2 Hz, 1 H), 4.18 (m, 2 H), 4.30 (d, J = 8.7 Hz, 1 H), 4.42 (m, 1 H), 4.75 (m, 2 H), 4.96 (d, J = 7.8 Hz, 1 H), 5.29 (m, 2 H), 5.64 (d, J = 7.2 Hz, 1H), 6.16 (t, J = 8.7 Hz, 1 H), 6.30 (s, 1 H), 7.37 (m, 2 H), 7.90 (m, 2 H); 13C (63 MHz, CDC13) δ 9.5, 14.9, 18.5, 20.8, 21.3, 21.8, 22.3, 25.7, 26.6, 28.2, 35.6, 43.2, 25.6, 58.4, 72.1, 72.3, 73.7, 74.9, 75.6, 76.46, 79.1, 81.1, 84.4, 120.7, 127.3, 128.5, 129.1, 130.8, 132.8, 134.4, 138.3, 142.6, 155.4, 167.0, 170.0, 171.3, 203.7. HRMS (FAB) m/z calcd for C44H59NO15·Na+: 864.3782. Found: 864.3803 (Δ = − 2.4 ppm).

Other taxoids 14b–p, 15c–e and 15g were prepared in the same manner and characterization data are shown below.

2-Debenzoyl-2-(3-methoxybenzoyl)-10-acetyl-3'-dephenyl-3'-(2-methylprop-l-enyl)docetaxel (14b)

White solid; 72% (for 2 steps); mp 142–144 °C; [α]D25 – 72 (c 0.70, CHC13); 1H NMR (300 MHz, CDCI3) δ 1.15 (s, 3H), 1.26 (s, 3H), 1.34 (s, 9H), 1.68 (s, 3H), 1.74 (s, 3H), 1.77 (s, 3H), 1.90 (s, 3H), 1.92 (m, 1H), 2.24 (s, 3H), 2.35 (s, 3H), 2.38 (m, 2H), 2.54 (m, 1H), 3.81 (d, J = 6.9 Hz, 1H), 3.87 (s, 3H), 4.18 (d, J = 8.4 Hz, 1H), 4.35 (d, J = 8.4 Hz, 1H), 4.43 (dd, J = 6.6, 10.5 Hz, 1H), 4.75 (br s, 2H), 4.97 (d, J = 8.1 Hz, 1 H), 5.32 (s, 1H), 5.66 (d, J = 7.2 Hz, 1H), 6.18 (t, J = 8.4 Hz, 1H), 6.30 (s, 1H), 7.14 (dd, J = 2.1, 8.1 Hz, 1H), 7.38 (t, J = 7.8 Hz, 1H), 7.64 (s, 1H), 7.70 (d, J = 7.8 Hz, 1H); 13C NMR (75.0 MHz, CDCl3) δ 9.5, 15.0, 18.5, 20.9, 21.8, 22.4, 25.7, 26.7, 28.2, 29.7, 35.6, 43.2, 45.6, 51.5, 55.4, 58.6, 72.2, 72.3, 73.7, 75.1, 75.6, 76.4, 79.1, 80.0, 81.1, 84.4, 114.6, 120.2, 122.6, 122.6, 129.6, 130.4, 132.8, 137.9, 142.7, 143.5, 159.7, 166.8, 170.0, 171.3, 173.1, 203.8. HRMS (DCM/NBA/NACL) calcd. for C44H59NO16Na+: 880.3766, obtained: 880.3732 (Δ = −3.9 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

2-Debenzoyl-2-(3-fluorobenzoyl)-10-deacetyl-10-propanoyl-3'-dephenyl-3'-(2-methylprop-l-enyl)docetaxel (14c)

White solid; 84% (for 2 steps); mp 137–139 °C; [α]20D −78 (c 0.45, CHC13); 1H NMR(CDCl3) δ 1.13(s, 3 H), 1.19 (m, 6 H), 1.33 (m, 9 H), 1.66 (s, 3 H), 1.75 (s, 6 H), 1.89 (s, 3 H), 2.34 (m, 5 H), 2.52 (m, 4 H), 3.37 (m, 1 H), 3.80 (d, J = 7.0 Hz, 1 H), 4.12 (m, 2 H), 4.29 (d, J = 8.5 Hz, 1 H), 4.42 (m, 1 H), 4.76 (m, 1 H), 4.96 (d, J = 8.1 Hz, 1 H), 5.29 (m, 1 H), 5.63 (d, J = 7.2 Hz, 1 H), 6.16 (t, J = 8.4 Hz, 1 H), 6.30 (s, 1 H), 7.29 (m, 1 H), 7.43 (m, 1 H), 7.77 (d, J = 9.1 Hz, 1 H), 7.89 (d, J = 7.8 Hz, 1 H); 13C NMR (CDC13) δ 9.1, 9.5, 14.9, 18.5, 21.8, 22.3, 25.7, 26.6, 27.6, 28.2, 35.5, 43.1, 45.6, 51.6, 58.5, 72.2, 73.7, 75.4, 76.3, 77.6, 79.2, 79.9, 81.0, 84.4, 116.8, 117.1, 120.6, 125.9, 130.3, 130.4, 132.8, 138.0, 142.6, 155.4, 165.7, 170.0, 174.6, 203.7. 19F NMR (CDCl3, using Freon as standard) δ −112.1. HRMS (FAB) m/z calcd for C44H58O15FN·H+: 860.3869. Found: 860.3870 (Δ = − 0.1 ppm).

2-Debenzoyl-2-(3-chlorobenzoyl)-10-deacetyl-10-propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)docetaxel (14d)

White solid; 70% (for 2 steps); mp 142–144 °C; [α]D20 −89 (c 0.090, CHC13); 1H NMR (CDCl3) δ 1.13 (s, 3 H), 1.21 (m, 7 H), 1.33 (s, 9 H), 1.66 (m, 3 H), 1.74 (m, 7 H), 1.82 (s, 3 H), 2.45 (m, 5 H), 2.52 (m, 3 H), 3.80 (d, J = 7.1 Hz, 1 H), 4.12 (m, 2 H), 4.27 (d, J = 8.3 Hz, 1 H), 4.40 (dd, J = 10.6, 6.5 Hz, 1 H), 4.75 (m, 2 H), 4.96 (d, J = 8.2 Hz, 1 H), 5.30 (d, J = 9.2 Hz, 1 H), 5.60 (d, J = 7.1 Hz, 1 H), 6.13 (t, J = 8.6 Hz, 1 H), 6.30 (s, 1 H), 7.40 (t, J = 7.8 Hz, 1 H), 7.56 (d, J = 8.3 Hz, 1 H), 7.97 (d, J = 7.7 Hz, 1 H), 8.10 (s, 1 H); 13C NMR (CDC13) δ 9.0, 9.5, 14.9, 18.6, 21.8, 22.3, 25.6, 26.5, 27.5, 28.1, 35.5, 41.2, 43.1, 45.6, 51.6, 58.4, 72.2, 72.3, 73.7, 75.4, 75.5, 76.3, 79.2, 79.9, 81.0, 84.4, 120.5, 128.3, 130.0, 130.2, 131.0, 132.7, 133.6, 134.7, 137.9, 142.6, 155.4, 165.5, 169.9, 173.1, 174.6, 203.7. HRMS: m/e calcd for C44H58O15NCl·H+: 876.3573. Found: 876.3573 (Δ = 0.0 ppm).

2-Debenzoyl-2-(3-azidobenzoyl)-10-deacetyl-10-propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)docetaxel (14e)

White solid; 71% (for 2 steps); mp 128–130 °C; [α]D20 − 71 (c 0.44, CHC13); 1H NMR (CDCl3) δ 1.14 (s, 3 H), 1.25 (m, 9 H), 1.34 (s, 9 H), 1.66–1.73 (m, 12 H), 1.89 (s, 3 H), 2.37 (m, 5 H), 2.52 (m, 4 H), 3.32 (bs, 1 H), 3.81 (d, J = 6.9 Hz, 1 H), 4.12 (m, 2 H), 4.30 (d, J = 8.1 Hz, 1 H), 4.40 (dd, J = 10.6, 6.8 Hz, 1 H), 4.74 (m, 2 H), 4.96 (d, J = 8.2 Hz, 1 H), 5.29 (m, 1 H), 5.64 (d, J = 7.0 Hz, 1 H), 6.13 (t, J = 9.0 Hz, 1 H), 6.31 (s, 1 H), 7.23 (d, J = 7.5 Hz, 1 H), 7.46 (t, J = 7.9 Hz, 1 H), 7.78 (s, 1 H), 7.86 (d, J = 7.8 Hz, 1 H); 13C NMR (CDC13) δ 9.0, 9.5, 15.0, 18.6, 21.8, 22.5, 25.7, 26.6, 27.6, 28.2, 35.5, 43.2, 45.6, 51.6, 58.6, 72.2, 72.6, 73.7, 75.4, 76.4, 79.2, 81.0, 84.5, 87.3, 120.1, 120.5, 124.3, 126.8, 130.2, 132.7, 140.8, 166.0, 170.1, 174.6, 203.8. HRMS: m/e calcd for C44H58O15N4·H+: 883.3977. Found: 883.3987 (Δ = − 1.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

3'-Dephenyl-3'-(2-methylprop-1-enyl)-2-debenzoyl-2-(3-allylbenzoyl)-10-propanoyldocetaxel (14f)

White solid; 80% (for 2 steps), mp 118–120 °C; 1H NMR (300 MHz, CDC13) δ 1.15 (s, 3 H), 1.21 (t, J = 7.4 Hz, 3 H), 1.23 (s, 3 H), 1.34 (s, 9 H), 1.68 (s, 3 H), 1.75 (s, 3 H), 1.77 (s, 3 H), 1.86 (m, 1 H), 1.90 (s, 3 H), 2.36 (s, 6 H), 2.39 (m, 2 H), 2.37 (s, 3 H), 2.53 (m, 3 H), 3.36 (bs, 1 H), 3.83 (d, J = 6.9 Hz, 1 H), 4.19 (m, 3 H), 4.32 (d, J = 8.4 Hz, 1 H), 4.43 (dd, J = 6.6, 10.5 Hz, 1 H), 4.74 (d, J = 3.0 Hz, 1 H), 4.97 (d, J = 8.4 Hz, 1 H), 5.34 (br d, J = 10.8 Hz, 2 H), 5.67 (d, J = 6.9 Hz, 1 H), 5.85 (d, J = 17.4 Hz, 1 H), 6.18 (t, J = 8.4 Hz, 1 H), 6.32 (s, 1 H), 6.76 (dd, J = 10.5, 17.4 Hz, 1 H), 7.44 (t, J = 7.8 Hz, 1 H), 7.63 (d, J = 7.5 Hz, 1 H), 7.98 (d, J = 7.8 Hz, 1 H), 8.17 (s, 1 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.5, 15.0, 18.5, 20.9, 22.4, 25.7, 26.7, 28.2, 29.7, 35.5, 35.6, 43.2, 45.6, 51.5, 58.6, 72.2, 72.9, 75.1, 75.4, 79.2, 80.0, 81.1, 84.4, 115.4, 120.6, 127.8, 128.9, 129.4, 131.3, 135.9, 138.1, 155.4, 159.7, 165.4, 166.9, 170.0, 171.9, 178.5, 203.8. HRMS (FAB) m/z calcd for C46H61NO15·Na+: 890.3981. Found: 890.3939 (Δ = −4.7 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

2-Debenzoyl-2-(3-methoxybenzoyl)-10-deacetyl-10-propanoyl-3'-dephenyl-3'-(2-methylprop-l-enyl)docetaxel (14g)

White solid; 69% (for 2 steps); mp 130–132 °C; [α]D20 −75 (c 0.080, CHC13); 1H NMR(CDCl3) δ 1.13(s, 3 H), 1.28 (m, 8 H), 1.33 (s, 9 H), 1.66 (m, 3 H), 1.73 (s, 3 H), 1.75 (s, 3 H), 1.89 (m, 5 H), 2.37 (m, 6 H), 2.52 (m, 3 H), 3.80 (d, J = 6.9 Hz, 1 H), 3.86 (s, 3 H), 4.12 (m, 2 H), 4.32 (d, J = 8.5 Hz, 1 H), 4.40 (dd, J = 10.6, 6.8 Hz, 1 H), 4.72 (m, 2 H), 4.96 (d, J = 8.3 Hz, 1 H), 5.30 (d, J = 7.6 Hz, 1 H), 5.64 (d, J = 7.0 Hz, 1 H), 6.16 (t, J = 8.6 Hz, 1 H), 6.30 (s, 1 H), 7.13 (d, J = 7.9 Hz, 1 H), 7.33 (t, J = 8.0 Hz, 1 H), 7.62 (s, 1 H), 7.68 (d, J = 7.6 Hz, 1 H); 13C NMR (CDC13) δ 9.0, 9.5, 14.9, 18.5, 21.8, 22.4, 25.7, 26.6, 27.5, 28.2, 35.5, 43.2, 45.6, 51.5, 55.3, 58.5, 72.2, 72.3, 73.7, 75.1, 75.4, 76.2, 79.1, 79.9, 81.1, 84.4, 114.6, 120.1, 120.6, 122.5, 129.6, 130.4, 132.9, 137.8, 142.5, 155.4, 159.6, 166.8, 170.0, 174.0,174.6,203.8. HRMS: m/e calcd for C45H61O16N·H+: 872.4069. Found: 872.4072 (Δ = −0.4 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

3'-Dephenyl-3'-(2-methylprop-l-enyl)-2-debenzoyl-2-(3-methoxybenzoyl)-10-(cyclopropane-carbonyl)docetaxel (14h)

77% (for 2 steps); mp 144–146 °C; [α]D25–74 (c 0.92, CHC13); 1H NMR (300 MHz, CDC13) δ 1.16 (s, 3H), 1.26 (s, 3H), 1.34 (s, 9H), 1.66 (m, 4H), 1.67 (s, 3H), 1.74 (s, 3H), 1.77 (s, 3H), 1.82 (m, 2H), 1.89 (s, 3H), 2.34 (s, 3H), 2.38 (m, 2H), 2.54 (m, 1H), 3.36 (br s, 1H), 3.81 (d, J = 7.2 Hz, 1H), 3.87 (s, 3H), 4.18 (d, J = 9.0 Hz, 1H), 4.20 (s, 1H), 4.34 (d, J = 8.4 Hz, 1H), 4.41 (dd, J = 6.6, 11.1 Hz, 1H), 4.79 (m, 2H), 4.97 (d, J = 9.0 Hz, 1 H), 5.31 (br s, 1H), 5.66 (d, J = 6.9 Hz, 1H), 6.18 (t, J = 9.0 Hz, 1H), 6.30 (s, 1H), 7.14 (d, J = 8.1 Hz, 1H), 7.37 (t, J = 7.8 Hz, 1H), 7.64 (s, 1H), 7.70 (d, J = 7.5 Hz, 1H); 13C NMR (75.0 MHz, CDC13) δ 9.5, 13.0, 15.0, 18.5, 21.9, 22.4, 25.7, 26.7, 28.2, 29.7, 35.5, 35.6, 43.2, 45.6, 51.5, 55.4, 58.6, 72.2, 72.4, 73.7, 75.1, 75.4, 76.4, 79.2, 80.0, 81.1, 84.4, 114.6, 120.1, 120.6, 122.6, 129.6, 130.4, 132.9, 137.9, 142.7, 155.4, 159.7, 166.8, 170.0, 171.9, 175.1, 203.9. HRMS (DCM/NBA/NACL) calcd. for C46H61NO16Na+: 906.3909, obtained: 906.3888 (Δ = −2.3 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-2-debenzoyl-2-(3-methoxybenzoyl)-10-(methoxycarbonyl)docetaxel (14i)

White solid; 80% (for 2 steps); mp 132–138 °C; 1H NMR (300 MHz, CDCl3) δ 1.15 (s, 3 H), 1.25 (m, 12 H), 1.68 (m, 6 H), 1.89 (m, 1 H), 1.93 (s, 3 H), 2.17 (s, 3 H), 2.35 (s, 3 H), 2.57 (m, 1 H), 3.78 (d, J = 6.9 Hz, 1 H), 3.87 (bs, 7 H), 4.20 (m, 2 H), 4.36 (m, 2 H), 4.73 (m, 2 H), 4.95 (d, J = 7.8 Hz, 1 H), 5.31 (d, J = 7.0, 1 H), 5.65 (d, J = 6.9 Hz, 1 H), 6.17 (m, 2 H), 7.12 (dd, J = 7.8 Hz, 2.4 Hz, 1 H), 7.37 (t, J = 7.8 Hz, 1 H), 7.63 (s, 1 H), 7.68 (d, J = 4.5 Hz, 1 H), 8.01 (s, 1 H); 13C NMR (63 MHz, CDC13) δ 9.4, 15.0, 18.5, 21.7, 22.4, 25.7, 26.6, 28.2, 30.9, 33.3, 35.6, 43.1, 45.6, 51.5, 55.4, 55.6, 58.6, 72.1, 72.3, 73.7, 75.1, 78.3, 79.1, 80.0, 81.1, 84.4, 105.1, 114.6, 120.1, 120.6, 122.5, 129.6, 130.4, 132.5, 137.9, 143.5, 155.8, 159.7, 166.8, 170.1, 176.4, 204.0. HRMS (FAB) m/z calcd for C51H65NO17·H+: 964.4331. Found: 964.4366 (Δ = 3.7 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-2-debenzoyl-2-(3-methoxybenzoyl)-10-(benzyloxy-carbonyl)docetaxel (14j)

White solid; 70% (for 2 steps); mp 145–150 °C; 1H NMR (300 MHz, CDC13) δ 1.15 (s, 3 H), 1.25 (m, 12 H), 1.69 (m, 6 H), 1.89 (m, 1 H), 1.95 (s, 3 H), 2.16 (s, 3 H), 2.35 (s, 3 H), 2.56 (m, 1 H), 3.36 (d, J = 6.9 Hz, 1 H), 3.78 (d, J = 7.2 Hz, 1 H), 3.87 (s, 3 H), 4.19 (m, 2 H), 4.33 (d, J = 8.7 Hz, 1 H), 4.40 (m, 1 H), 4.72 (m, 2 H), 4.95 (d, J = 7.5 Hz, 1 H), 5.23 (s, 2 H), 5.31 (d, J = 7.8 Hz, 1 H), 5.65 (d, J = 7.2 Hz, 1 H), 6.14 (m, 2 H), 7.12 (dd, J = 7.2 Hz, 1.8 Hz, 1 H), 7.34 (m, 6 H), 7.63 (s, 1 H), 7.68 (d, J = 7.8 Hz, 1 H). HRMS (FAB) m/z calcd for C50H63NO17·H+: 950.4174. Found: 950.4164 (Δ = −1.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

3'-Dephenyl-3'-(2-methyl-1-propenyl)-2-debenzoyl-2-(3-methoxybenzoyl)-10-(2-methoxybenzoyl)docetaxel (14k)

White solid; 95% (for 2 steps); mp 143–145 °C; 1H NMR (300 MHz, CDCl3) δ 1.28 (s, 3 H), 1.31 (s, 3 H), 1.34 (s, 9 H), 1.60 (m, 1 H), 1.71 (s, 3 H), 1.75 (s, 3 H), 1.77 (s, 3 H), 1.91 (m, 1 H), 1.96 (s, 3 H), 2.37 (s, 3 H), 2.59 (m, 2 H), 3.35 (bs, 1 H), 3.88 (s, 3 H), 3.91 (s, 3 H), 4.22 (m, 2H), 4.35 (d, J = 8.4 Hz, 1 H), 4.51 (m, 1 H), 4.76 (m, 2 H), 4.98 (d, J = 8.1 Hz,1 H), 5.32 (m, 1 H), 5.71 (d, J = 6.9 Hz, 1 H), 6.21 (m, 1 H), 6.57 (s, 1 H), 6.99 (m, 2 H), 7.13 (dd, J = 8.4 Hz, 1.8 Hz, 1 H), 7.36 (t, J = 7.5 Hz, 1 H), 7.50 (m, 1 H), 7.65 (s, 1 H), 7.70 (d, J = 7.5 Hz, 1 H), 7.99 (dd, J = 7.8 Hz, 2.1 Hz, 1 H); 13CNMR(63 MHz, CDC13) δ 9.56, 15.0, 18.5, 22.4, 25.7, 26.6, 28.2, 35.6, 43.3, 45.7, 55.4, 55.9, 58.7, 72.2, 72.4, 73.7, 75.2, 75.6, 79.2, 81.2, 84.5, 87.6, 105.0, 105.1, 112.1, 120.2, 120.3, 122.6, 129.7, 130.5, 132.7, 134.6, 159.7, 160.0, 165.9, 166.9, 170.1, 203.8. HRMS (FAB) m/z calcd for C50H63NO17·H+: 950.4174. Found: 950.4149 (Δ = −2.7 ppm).

3'-Dephenyl-3'-(2-methyl-1-propenyl)-2-debenzoyl-2-(3-methoxybenzoyl)-10-(4-methoxyphenylacetyl)docetaxel (141)

White solid; 70% (for 2 steps); mp 152–154 °C; 1H NMR (300 MHz, CDC13) δ 1.07 (s, 3 H), 1.18 (s, 3 H), 1.34 (s, 9 H), 1.59 (s, 3 H), 1.67 (s, 3 H), 1.73 (s, 3 H), 1.76 (s, 3 H) 1.86 (s, 3 H), 1.86 (m, 1 H), 2.17 (m, 4 H), 2.37 (s, 3 H), 2.39 (m, 2 H), 2.55 (m, 1 H), 3.34 (bs, 1 H), 3.79 (m, 4 H), 3.68 (s, 3 H), 4.19 (m, 1 H), 4.32 (d, J = 8.4 Hz, 1 H), 4.39 (m, 1 H), 4.73 (m, 2 H), 4.94 (d, J = 8.1 Hz, 1 H), 5.30 (m, 1 H), 5.64 (d, J = 6.9 Hz, 1 H), 6.15 (t, J = 8.1 Hz, 1 H), 6.30 (s, 1 H), 6.86 (d, J = 8.4 Hz, 2 H), 7.12 (dd, J = 7.8 Hz, 2.1 Hz, 1 H), 7.24 (m, 2 H), 7.37 (t, J = 7.5 Hz, 1 H), 7.63 (s, 1 H), 7.68 (d, J = 7.8 Hz, 1 H); 13C NMR (63 MHz, CDC13) δ 9.53, 14.2, 14.9, 18.5, 21.6, 22.4, 25.7, 26.4, 28.2, 30.9, 35.6, 40.1, 43.1, 45.6, 51.5, 55.2, 55.3, 58.5, 72.1, 72.3, 73.7, 75.0, 75.8, 79.0, 79.9, 81.0, 84.4, 87.5, 114.0, 114.6, 120.1, 120.6, 122.5, 129.6, 130.4, 130.5, 132.7, 142.5, 159.6, 166., 170.0, 172.0, 203.4, 206.9. HRMS (FAB) m/z calcd for C51H65NO17·H+: 964.4331. Found: 964.4366 (Δ = 3.7 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(prop-2-enyl)docetaxel (14m)

White solid; 65% (for 2 steps); mp 127–130 °C; [α]D20 −64 (c 0.25, CHC13); 1H NMR (300 MHz, CDCl3) δ 1.13 (s, 3 H), 1.21 (m, 26 H), 1.66 (m, 3 H), 1.88 (s, 3 H), 2.20–2.37 (m, 5 H), 2.52 (m, 3 H), 3.39 (bs, 1 H), 3.80 (d, J = 8.8 Hz, 1 H), 3.89 (s, 3 H), 4.12 (m, 2 H), 4.35 (m, 3 H), 4.61 (m, 1 H), 4.96 (m, 2 H), 5.63 (m, 1.25 H), 5.84 (d, J = 5.8 Hz, 0.5 H), 6.06 (d, J = 5.9 Hz, 0.25 H), 6.23 (t, J = 8.8 Hz, 1 H), 6.30 (s, 1 H), 7.13 (d, J = 5.8 Hz, 1 H), 7.38 (t, J = 8.0 Hz, 1 H), 7.63 (s, 1 H), 7.70 (d, J = 7.6 Hz, 1 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.6, 14.8, 21.9, 22.5, 26.7, 27.6, 28.0, 29.7, 35.5, 43.2, 45.7, 55.3, 58.6, 68.5, 72.2, 73.0, 75.1, 75.3, 79.0, 81.2, 84.5, 87.4, 103.1, 114.1, 120.7, 122.7, 129.8, 130.2, 133.3, 159.7, 167.1, 170.3, 170.7, 174.6, 203.6. HRMS (FAB) m/z calcd for C44H59NO16·H+: 858.3912. Found: 858.3880 (Δ = − 3.7 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-[(E)-prop-1-enyl]docetaxel (14n)

White solid; 80% (for 2 steps); mp 120–122 °C; [α]D20 −100 (c 0.010, CHC13). 1H NMR (300 MHz, CDCl3) δ 1.14 (s, 3 H), 1.32 (s 3 H), 1.66 (s, 3 H), 1.73 (s 3 H), 1.75 (s, 3 H), 1.88 (s, 3 H), 2.36 (m, 6 H), 3.87 (s, 2 H), 4.12 (d, 2 H), 4.27 (m, 2 H), 4.40 (dd, J = 10.6, 6.8 Hz, 1 H), 4.57 (b, 1 H), 4.86 (d, 1 H), 4.96 (d, J = 8.1 Hz, 1 H), 5.60 (m, 2 H), 6.13 (t, J = 8.8 Hz, 1 H), 6.30 (s, 1 H), 7.40 (t, J = 7.8 Hz, 1 H), 7.56 (d, J = 8.3 Hz, 1 H), 7.97 (d, J = 7.7 Hz, 1 H), 8.10 (s, 1 H). 13C NMR (62.9 MHz, CDC13) δ 4.5, 5.0, 10.4, 13.3, 17.3, 18.0, 22.2, 23.0, 23.6, 25.2, 31.0, 31.1, 38.7, 41.1, 50.9, 54.1, 67.7, 68.6, 70.6, 70.9, 73.2, 74.5, 76.6, 79.9, 110.0, 115.8, 118.1, 122.8, 124.3, 125.2, 128.0, 138.0, 149.0, 162.0, 165.6, 170.0, 199.0. HRMS: m/e calcd forC44H59NO16H+: 858.3912 Found: 858.3880 (Δ = −3.7 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(but-3-enyl)docetaxel (14o)

White solid; 71% (for 2 steps); mp 116–117 °C; [α]D20 −37 (c 0.30, CHC13); 1H NMR (300 MHz, CDC13) δ 1.14 (s, 3 H), 1.23 (t, J =7.5 Hz, 3 H), 1.25 (s, 3 H), 1.30 (s, 9 H), 1.39 (m, 2 H), 1.67 (m, 3 H), 1.84 (m, 1 H), 1.88 (s, 3 H), 2.15 (m, 2 H), 2.33 (m, 2 H), 2.37 (s, 3 H), 2.53 (m, 3 H), 3.26 (bs, 1 H), 3.81 (d, J = 6.9 Hz, 1 H), 3.89 (s, 3 H), 4.05 (m, 1 H), 4.18 (d, J = 8.1 Hz, 1 H), 4.23 (s, 1 H), 4.36 (d, J = 8.4 Hz, 1 H), 4.44 (dd, J = 7.2, 10.2 Hz, 1 H), 4.64 (d, J = 9.9 Hz, 1 H), 4.97 (d, J = 8.7 Hz, 1 H), 5.05 (d, J = 8.4 Hz, 1 H), 5.08 (d, J = 15.3 Hz, 1 H), 5.66 (d, J = 7.2 Hz, 1 H), 5.82 (m, 1H), 6.21 (t, J = 8.7 Hz, 1 H), 6.31 (s, 1 H), 7.14 (dd, J = 2.4, 8.1 Hz, 1 H), 7.39 (t, J = 8.1 Hz, 1 H), 7.65 (s, 1 H), 7.72 (d, J = 7.5 Hz, 1 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.5, 14.9, 21.9, 22.6, 26.7, 27.6, 28.1, 29.7, 30.1, 31.4, 35.5, 35.6, 43.2, 45.6, 55.4, 58.6, 72.2, 72.5, 75.1, 75.4, 79.1, 79.8, 81.1, 84.4, 114.4, 115.8, 120.3, 122.7, 129.7, 130.4, 137.2, 155.4, 159.6, 166.9, 170.3, 170.0, 173.9, 174.6, 203.8. HRMS (FAB) m/z calcd for C45H61NO16·Na+: 894.3870. Found: 894.3904 (Δ = −3.1ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-[(S)-2,2-dimethylcyclopropyl]docetaxel (14p)

White solid; 83% (for 2 steps); 1H NMR (CDC13) δ 0.098 (t, J = 4.6 Hz, 1H), 0.62 (dd, J = 8.5, 4.3 H), 1.11 (m, 10 H), 1.24 (s, 6 H), 1.31 (s, 9 H), 1.66 (s, 3 H), 1.83 (s, 1 H), 1.89 (s, 3 H), 2.34 (s, 3 H), 2.37 (m, 2 H), 2. 54 (m, 3 H), 3.34 (d, J = 6.6 Hz, 1 H), 3.51 (t, J = 9.3 Hz, 1 H), 3.80 (d, J = 7.0 Hz, 1 H), 3.85 (s, 3 H), 4.18 (d, J = 8.4 Hz, 1 H), 4.32 (s, 1 H), 4.34 (d, J = 8.4 Hz, 1 H), 4.43 (m, 1 H), 4.80 (d, J = 8.9 Hz, 1 H), 4.97 (d, J = 8.4 Hz, 1 H), 5.65 (d, J = 7.0 Hz, 1 H), 6.15 (t, J = 8.6 Hz, 1 H), 6.30 (s, 1 H), 7.13 (dd, J = 8.0, 2.2 Hz, 1 H), 7.36 (t, J = 8.0 Hz, 1 H), 7.63 (s, 1 H), 7.69 (d, J = 7.5 Hz, 1 H); 13C NMR (CDC13) δ 9.0, 9.5, 14.9, 17.1, 19.3, 20.1, 22.0, 22.6, 26.2, 26.5, 27.1, 27.5, 28.1, 33.2, 35.5, 43.2, 45.5, 55.3, 55.4, 58.5, 72.1, 72.7, 73.0, 75.1, 75.4, 76.4, 79.1, 79.8, 81.1, 84.4, 87.4, 114.4, 120.3, 122.6, 129.6, 130.4, 132.8, 142.6, 155.0, 159.6, 166.7, 169.7, 170.6, 174.6, 203.8. HRMS (FAB) m/z calcd for C46H63NO16·H+: 886.4225. Found: 886.4237 (Δ = −1.3 ppm).

2-Debenzoyl-2-(3-fluorobenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropyl)docetaxel (15c)

White solid; 89% for 2 steps): mp 138–140 °C; [α]D20 −79 (c 0.47, CHC13); 1H NMR (CDCI3) δ 0.95 (s, 3 H), 0.97 (s, 3 H), 1.13 (s, 3 H), 1.28 (m, 18 H), 1.66 (m, 5 H), 2.37 (m, 6 H), 2.52 (m, 4 H), 3.21 (bs, 1 H), 3.80 (d, J = 7.0 Hz, 1 H), 4.12 (m, 2 H), 4.29 (d, J = 8.5 Hz, 1 H), 4.42 (m, 1 H), 4.57 (d, J = 9.7 Hz, 1 H), 4.96 (d, J = 10.1 Hz, 1 H), 5.61 (d, J = 6.8 Hz, 1 H), 6.16 (t, J = 8.4 Hz, 1 H), 6.30 (s, 1 H), 7.29 (m, 1 H), 7.43 (m, 1 H), 7.77 (d, J = 9.1 Hz, 1 H), 7.89 (d, J = 7.8 Hz, 1 H); 13C NMR (CDCl3) δ 9.1, 9.5, 14.9, 21.8, 22.4, 23.2, 24.7, 26.5, 27.6, 28.1, 35.5, 41.2, 43.2, 45.6, 51.3, 57.6, 72.2, 72.6, 73.0, 75.4, 75.5, 76.2, 79.2, 79.6, 81.0, 84.4, 116.8, 117.2, 120.6, 120.9, 126.0, 130.3, 130.4, 142.6, 155.5, 169.9, 174.0, 174.6, 203.7. 19F NMR (CDCl3, using Freon as the standard) δ −112.1. HRMS (FAB) m/z calcd for C44H60O15FN·H+: 862.4025. Found: 862.4022 (Δ = + 0.4 ppm).

2-Debenzoyl-2-(3-chlorobenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropyl)docetaxel (15d)

White solid; 80% (for 2 steps); mp 143–145 °C; [α]D20 −83 (c 0.12, CHC13); 1H NMR (CDC13) δ 0.95 (m, 6 H), 1.12 (s, 3 H), 1.28 (m, 26 H), 1.66 (m, 6 H), 1.88 (s, 3 H), 2.37 (m, 6 H), 2.52 (m, 4 H), 3.21 (bs, 1 H), 3.80 (d, J = 7.1 Hz, 1 H), 4.12 (m, 2 H), 4.27 (d, J = 8.3 Hz, 1 H), 4.40 (dd, J = 10.6, 6.8 Hz, 1 H), 4.57 (d, J = 9.6 Hz, 1 H), 4.96 (d, J = 8.1 Hz, 1 H), 5.60 (d, J = 7.1 Hz, 1 H), 6.13 (t, J = 8.8 Hz, 1 H), 6.30 (s, 1 H), 7.40 (t, J = 7.8 Hz, 1 H), 7.56 (d, J = 8.3 Hz, 1 H), 7.97 (d, J = 7.7 Hz, 1 H), 8.10 (s, 1 H); 13C NMR (CDCl3) δ 9.0, 9.5, 14.9, 21.8, 22.3, 23.2, 24.6, 26.5, 27.5, 28.1, 35.5, 41.2, 43.1, 45.6, 51.3, 58.4, 72.2, 72.6, 73.0, 75.4, 75.5, 76.2, 79.2, 79.6, 81.0, 84.4, 128.3, 130.0, 130.3, 131.0, 132.7, 133.6, 134.7, 142.6, 155.5, 165.5, 169.8, 174.0, 174.6, 203.7. HRMS: m/e calcd for C44H60O15NCl·H+: 878.3730. Found: 878.3728 (Δ = 0.2 ppm).

2-Debenzoyl-2-(3-azidobenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropyl)docetaxel (15e)

White solid; 70% (for 2 steps); mp 132–134 °C; [α]D20 −70 (c 0.47, CHC13); 1H NMR (CDC13) δ 0.95 (m, 6 H), 1.12 (s, 3 H), 1.28 (m, 26 H), 1.66 (m, 6 H), 1.88 (s, 3 H), 2.37 (m, 5 H), 2.52 (m, 4 H), 3.18 (bs, 1 H), 3.82 (d, J = 6.9 Hz, 1 H), 4.12 (m, 4 H), 4.31 (d, J = 8.4 Hz, 1 H), 4.40 (dd, J = 10.2, 6.6 Hz, 1 H), 4.56 (d, J = 9.6 Hz, 1 H), 4.97 (d, J = 8.7 Hz, 1 H), 5.65 (d, J = 6.9 Hz, 1 H), 6.14 (t, J = 8.4 Hz, 1 H), 6.31 (s, 1 H), 7.23 (m, 1 H), 7.46 (t, J = 7.8 Hz, 1 H), 7.88 (d, J = 7.5 Hz, 1 H); 13C NMR (CDCI3) δ 9.0, 9.5, 14.9, 21.8, 22.5, 23.2, 24.7, 26.6, 27.6, 28.1, 35.5, 41.2, 43.2, 45.6, 51.4, 58.5, 72.2, 72.6, 73.0, 75.4, 75.5, 76.3, 78.0, 79.2, 79.7, 81.0, 84.4, 120.2, 124.3, 126.8, 130.1, 132.8, 140.8, 142.6, 155.5, 166.0, 170.1,174.0,174.6,203.8. HRMS: m/e calcd for C44H60N4O15·H+: 885.4133. Found: 885.4134 (Δ = −0.1 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropyl)docetaxel (15g)

White solid; 73% (for 2 steps); mp 132–134 °C; [α]D20 −110 (c 0.070, CHC13); 1HNMR (CDCl3) δ 0.95 (m, 6 H), 1.13 (s, 3 H), 1.28 (m, 8 H), 1.66 (m, 6 H), 1.88 (s, 3 H), 2.37 (m, 6 H), 2.52 (m, 4 H), 3.21 (bs, 1 H), 3.80 (d, J = 6.9 Hz, 1 H), 3.86 (s, 3 H), 4.12 (m, 2 H), 4.30 (d, J = 8.4 Hz, 1 H), 4.40 (dd, J = 10.6, 6.8 Hz, 1 H), 4.57 (d, J = 9.6 Hz, 1 H), 4.96 (d, J = 8.1 Hz, 1 H), 5.63 (d, J = 7.0 Hz, 1 H), 6.16 (t, J = 8.4 Hz, 1 H), 6.30 (s, 1 H), 7.13 (d, J = 7.9 Hz, 1 H), 7.33 (t, J = 8.0 Hz, 1 H), 7.62 (s, 1 H), 7.68 (d, J = 7.6 Hz, 1 H); 13C NMR (CDCl3) δ 9.0, 9.6, 14.9, 21.8, 22.5, 23.3, 24.7, 26.6, 27.6, 28.1, 35.5, 41.2, 43.2, 45.6, 51.2, 55.3, 58.5, 72.2, 72.6, 73.0, 75.4, 75.5, 76.2, 79.1, 79.7, 81.1, 84.4, 114.1, 120.4, 122.7, 129.6, 130.4, 132.9, 142.5, 155.4, 159.6, 166.8, 169.9, 174.0, 174.6, 203.8. HRMS: m/e calcd for C45H63O16N·H+: 874.4225. Found: 874.4224 (Δ = 0.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

Alternative Method for the synthesis of 15g through hydrogenation of 14g

To a reaction flask with activated Pd/C (33 mg, 0.014 mmol) was added a solution of 14g (250 mg, 0.287 mmol) in 10 mL EtOAc with several drops of MeOH under hydrogen atmosphere. The suspension was stirred overnight and the reaction mixture was filtered through Celite. The filtrate was condensed by a rotary evaporator and the residue was purified on a silica gel column using hexanes/EtOAc (1/1) as the eluant to afford 15g (250 mg, 99.8% yield) as a white solid.

Preparation of (3R,4S)-1-acyl-3-trialkylsiloxy-4-(2-methylprop-1-enyl)azetidin-2-ones (16)

A typical procedure is described for the preparation of (3R,4S)-1-cyclohexanecarbonyl-3-triisopropylsiloxy-4-(2-methylprop-1-enyl)azetidin-2-one (16h): To a solution of (3R,4S)-3-triisopropylsiloxy-4-(2-methylprop-1-enyl)azetidin-2-one20 (129 mg, 0.435mmol) and DMAP (53 mg, 0.435mmol) in 1 mL of CH2Cl2 was added triethylamine (0.6 mL, 4.35 mmol) followed by the dropwise addition of cyclohexyl chloroformate (0.14 mL, 0.652 mmol) at 0 °C. The mixture was stirred for 2 h at room temperature, quenched with NH4C1 (5 mL), and extracted with EtOAc (10 mL ×3), washed with brine, dried over MgSO4, and concentrated in vacuo. Column chromatography of the residue on silica gel (hexanes/EtOAc = 20/1) afforded 16h (177 mg, 94% yield) as a colorless oil; 1H NMR (300 MHz, CDC13) δ 1.03 (m, 21 H). 1.14–1.50 (m, 6 H), 1.64–1.93 (m, 4 H), 1.76 (s, 3 H), 1.78 (s, 3 H), 2.91 (m, 1 H), 4.80 (dd, J = 6.3 Hz, J = 9.9 Hz, 1 H), 4.99 (d, J = 6.3 Hz, 1 H), 5.22 (d, J = 9.9 Hz, 1 H).

Other N-modified β-lactams 16a–g were prepared in the same manner. (3R,4S)-3-tert-butyldimethylsiloxy-4-(2-methylprop-1-enyl)azetidin-2-one was prepared in the same manner as that described for the corresponding 3-triisopropylsiloxy-β-lactam.20,22 Characterization data (1H NMR) for 16a–g are summarized in the Supporting Information.

Synthesis of taxoids 17

Taxoids 17a–l were synthesized through the Ojima-Holton coupling of 7-TES-10-deacetyl-10-propanoylbaccatin17 or baccatin 11b with β-lactams 16a–h in the same manner as that described above for the synthesis of 14a–p. For example, 3'-dephenyl-3'-(2-methylprop-1-enyl)-3’N-debenzoyl-3’N-cyclobutanecarbonyl-10-propanoyldocetaxel (17a) was obtained in 86% yield for 2 steps as white solid: mp 149–151 °C [α]D20 −72.6 (c 2.91, CHC13); 1H NMR (300 MHz, CDC13) δ 1.16 (s, 3 H), 1.21–1.26 (m, 15 H), 1.69 (s, 3 H), 1.–2.00 (m, 10 H), 1.83 (s, 3 H), 2.09–2.21 (m, 5 H), 2.44–2.60 (m, 4 H), 2.92 (m, 1 H), 3.82 (d, J = 6.6 Hz, 1 H), 4.32–4.19 (m, 3 H), 4.43 (m, 1 H), 4.97 (d, J = 8.4 Hz, 1 H), 5.04 (m, 1 H), 5.35 (d, J = 8.7 Hz, 1 H), 5.58 (d, J = 8.4 Hz, 1 H), 5.68 (d, J = 7.2 Hz, 1 H), 6.20 (t, 1 H), 6.31 (s, 1 H), 7.47 (t, 2 H), 7.61 (t, 1 H), 8.11 (d, 2 H); 13C NMR (63 MHz, CDC13) δ 0, 9.5, 14.9, 18.6, 21.9, 22.4, 25.2, 25.5, 25.7, 26.7, 27.6, 29.7, 35.7, 39.7, 43.2, 45.6, 50.2, 58.6, 72.1, 72.2, 73.8, 75.0, 75.4, 76.4, 79.1, 81.0, 84.4, 120.0, 128.6, 129.2, 130.1, 133.1, 133.7, 139.0, 142.3, 166.9, 170.2, 172.8, 174.6, 174.9, 203.8. HRMS: m/e calcd for C44H57NO14·H+: 824.3857. Found: 824.3855 (Δ = 0.3 ppm).

Other taxoids 17b–l were synthesized in the same manner and characterization data are shown below.

10-Propanoyl-3'-Dephenyl-3'-(2-methylprop-1-enyl)-3’N-(cyclopentane-carbonyl)docetaxel (17b)

White solid; 78% (for 2 steps); mp 150–152 °C; [α]D20 −75.7 (c 3.75, CHC13); 1H NMR (300 MHz, CDCl3) δ 0.98 (m, 6H), 1.16 (s, 3 H), 1.23 (m, 3 H), 1.26 (s, 9 H), 1.84–1.41 (m, 13 H), 1.88 (s, 3 H), 2.39 (s, 3 H), 2.43–2.59 (m, 3 H), 3.37 (d, J = 5.7 Hz, 1 H), 3.80 (d, J = 7.2 Hz, 1 H), 4.20 (m, 2 H), 4.30 (d, J = 8.1, 1 H), 4.42 (m, 2 H), 4.97 (d, J = 7.5 Hz, 1 H), 5.47 (d, J = 9.4 Hz, 1 H), 5.68 (d, J = 7.2 Hz, 1 H), 6.14 (m, 1 H), 6.30 (s, 1 H), 7.47 (t, 2 H), 7.60 (t, 1 H), 8.12 (d, 2 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.6, 14.9, 21.9, 22.0, 22.5, 23.3, 24.8, 25.8, 25.8, 26.7, 27.6, 29.7, 30.7, 35.6, 35.8, 41.0, 43.2, 45.6, 45.7, 49.6, 58.6, 72.2, 72.6, 72.9, 75.0, 75.4, 76.5, 79.1, 81.1, 84.4, 128.7, 129.2, 130.2, 133.1, 133.6,142.2, 166.9, 170.1, 173.8, 174.6, 176.3, 203.8. HRMS: m/e calcd for C45H59NO14·H+: 838.4014. Found: 838.4011 (Δ = 0.3 ppm).

10-Propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)-3’N-(cyclohexanecarbonyl)docetaxel (17c)

White solid; 85% (for 2 steps); mp 152–155 °C; [α]D20 −78.3 (c 3.78, CHC13); 1HNMR (300 MHz, CDCl3) δ 0.08 (s, 3 H), 1.13–1.18 (m, 6 H), 1.28 (s, 9 H), 1.60 (s, 3 H), 1.69 (br, s, 6 H) 1.72 (m, 1 H), 1.83 (s, 3 H), 2.29 (s, 3 H), 2.31 (s, 2 H), 2.44 (m, 3 H), 3.38 (br s, 1 H), 3.74 (d, J = 6.9 Hz, 1 H), 4.10 (d, J = 8.1Hz, 1H), 4.13(brs 1 H), 4.22 (d, J = 8.1 Hz, 1 H), 4.33 (dd, J = 10.1, 7.5 Hz, 1 H), 4.67 (m, 2 H), 4.88 (d, J = 9.3 Hz, 1 H), 5.23 (d, J = 8.4 Hz, 1 H), 5.59 (d, J = 6.9 Hz, 1 H), 6.06 (m, 1 H), 6.24 (s, 1 H), 7.37 (t, 2 H), 7.51 (t, 1 H), 8.01 (d, 2 H). HRMS: m/e calcd for C46H61NO14·H+: 852.4170. Found: 852.4172 (Δ = −0.2 ppm).

10-Propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)-3’N-debenzoyl-3’N-(cyclopent-1-ene-1-carbonyl)docetaxel (17d)

White solid; 80% (for 2 steps); mp 148–151 °C [α]D20 −70.3 (c 1.45, CHC13); 1HNMR (300 MHz, CDCl3) δ 1.08 (s, 3 H), 1.13–1.18 (m, 6 H), 1.28 (s, 9 H), 1.60 (s, 3 H), 1.69 (br, s, 6 H) 1.72 (m, 1 H), 1.83 (s, 3 H), 2.29 (s, 3 H), 2.31 (s, 2 H), 2.44 (m, 3 H), 3.82 (d, J = 6.6 Hz, 1 H), 4.19 (d, J = 8.7 Hz, 1 H), 4.31 (m, 2 H), 4.42 (m, 1 H), 4.96 (d, J = 4.8 Hz, 1 H), 5.10 (m, 1 H), 5.38 (d, J = 9.0 Hz, 1H), 5.67 (d, J = 7.2 Hz, 1 H), 5.89 (d, J = 8.1 Hz, 1 H), 6.19 (t, 1 H), 6.31 (s, 1 H), 6.51 (dd, 1 H), 7.47 (t, 2 H), 7.61 (t, 1 H), 8.09 (d, 2 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.5, 14.9, 18.6, 21.8, 22.4, 23.2, 25.8, 26.8, 27.6, 31.4, 33.1, 35.6, 35.8, 43.2, 45.6, 50.4, 58.6, 72.0, 72.2, 73.8, 75.0, 75.4, 76.5, 79.0, 81.0, 84.4, 87.5, 120.0, 128.6, 129.2, 130.1, 133.1, 133.7, 138.4, 139.2, 139.5, 142.2, 165.0, 166.8, 170.3, 172.7, 174.6, 203.8. HRMS: m/e calcd for C45H57NO14·H+: 836.3857. Found: 836.3858 (Δ = −0.1 ppm).

10-Propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)-3’N-debenzoyl-3’N-(cyclohex-1-ene-1-carbonyl)docetaxel (17e)

White solid; 75% (for 2 steps); mp 151–154 °C; [α]D20 −67 (c 2.92, CHC13); 1H NMR (300 MHz, CDC13) δ 1.15 (s, 3 H), 1.13–1.18 (m, 6 H), 1.26 (s, 9 H), 1.60 (s, 3 H), 1.69 (br, s, 6 H), 1.72 (m, 1 H), 1.81 (s, 3H), 1.93 (s, 3 H), 2.16 (m, 4 H), 2.38 (s, 3 H), 2.45 – 2.59 (m, 3 H), 3.74 (d, J = 6.9 Hz), 3.82 (d, J = 6.6 Hz, 1 H), 4.20 (d, J = 8.4 Hz, 1 H), 4.30 (m, 2H), 4.41 (m, 1 H), 4.96 (d, J = 7.8 Hz, 1 H), 5.06 (m, 1H), 5.11 (d, J = 3.6 Hz, 1H), 5.37 (d, J = 9.0 Hz, 1 H), 5.67 (d, J = 7.2 Hz, 1 H), 5.93 (d, J = 7.8 Hz, 1H), 6.18 (t, 1 H), 6.31 (s, 1 H), 6.60 (dd, 1H), 7.46 (t, 2 H), 7.60 (t, 1 H), 8.09 (d, 2 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.5, 14.9, 18.6, 21.4, 21.8, 22.0, 22.4, 24.2, 25.4, 25.8, 26.8, 27.6, 29.7, 30.9, 35.6, 35.8, 43.2, 45.6, 50.5, 58.6, 71.9, 72.2, 73.9, 75.0, 75.4, 79.1, 81.0, 84.4, 120.1, 128.6, 129.2, 130.1, 132.5, 133.0, 133.7, 134.7, 139.2, 142.3, 166.9, 168.4, 170.3, 172.8, 174.6, 203.8. HRMS: m/e calcd for C46H59NO14·H+: 850.4014. Found: 850.4018 (Δ = −0.5 ppm).

10-Propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)-3’N-debenzoyl-3’N-(cyclopentyloxycarbonyl)docetaxel (17f)

White solid; 85% (for 2 steps); mp 141–143 °C [α]D20 −74 (c 5.06, CHC13); 1HNMR(300MHz,CDCl3) δ 1.08(s,3H), 1.13–1.18 (m, 6 H), 1.28 (s, 9 H), 1.60 (s, 3 H), 1.69 (br, s, 6 H) 1.72 (m, 1 H), 1.83 (s, 3 H), 2.29 (s, 3 H), 2.31 (s, 2 H), 2.44 (m, 3 H), 3.38 (br s, 1 H), 3.74 (d, J = 6.9 Hz, 1 H), 4.10 (d, J = 8.1 Hz, 1 H), 4.13 (br s 1 H), 4.22 (d, J = 8.1 Hz, 1 H), 4.33 (dd, J = 10.1, 7.5 Hz, 1 H), 4.67 (m, 2 H), 4.88 (d, J = 9.3 Hz, 1 H), 5.23 (d, J = 8.4 Hz, 1 H), 5.59 (d, J = 6.9 Hz, 1 H), 6.06 (m, 1 H), 6.24 (s, 1 H), 7.37 (t, 2 H), 7.51 (t, 1 H), 8.01 (d, 2 H); 13C NMR (63 MHz, CDCl3) δ 9.0, 9.5, 14.9, 18.5, 21.9, 22.4, 23.6, 23.6, 25.7, 26.7, 27.5, 29.7, 32.6, 32.6, 35.6, 35.6, 43.2, 45.7, 51.9, 58.5, 72.1, 72.2, 73.7, 75.0, 75.4, 76.4, 77.9, 79.2, 81.1, 84.4, 120.5, 128.6, 129.2, 130.1, 133.0, 133.7, 138.2, 142.3, 156.1, 166.9, 170.2, 172.8, 174.6, 203.8. HRMS: m/e calcd for C45H59NO15·H+: 854.3963. Found: 854.3960 (Δ = + 0.3 ppm).

10-Propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)-3’N-debenzoyl-3’N-(cyclohexyloxycarbonyl)docetaxel (17g)

White solid; 80% (for 2 steps); mp 142–144 °C [α]D20 −66.5 (c 5.00, CHCl3); 1H NMR (300 MHz, CDC13) δ 1.08 (s, 3 H), 1.13–1.18 (m, 6 H), 1.28 (s, 9 H), 1.60 (s, 3 H), 1.69 (br, s, 6 H) 1.72 (m, 1 H), 1.83 (s, 3 H), 2.29 (s, 3 H), 2.31 (s, 2 H), 2.44 (m, 3 H), 3.38 (br s, 1 H), 3.74 (d, J = 6.9 Hz, 1 H), 4.10 (d, J = 8.1 Hz, 1 H), 4.13 (br s 1 H), 4.22 (d, J = 8.1 Hz, 1 H), 4.33 (dd, J = 10.1, 7.5 Hz, 1 H), 4.67 (m, 2 H), 4.88 (d, J = 9.3 Hz, 1 H), 5.23 (d, J = 8.4 Hz, 1 H), 5.64 (d, J = 7.0 Hz, 1 H), 6.18 (m, 1 H), 6.30 (s, 1 H), 7.46 (t, 2 H), 7.59 (t, 1 H), 8.09 (d, 2 H); 13C NMR (63 MHz, CDCl3) δ 9.0, 9.5, 14.9, 18.5, 21.9, 22.4, 23.6, 23.7, 25.3, 25.7, 26.7, 27.5, 31.8, 35.6, 35.6, 43.2, 45.7, 51.8, 58.5, 72.1, 72.2, 73.7, 75.0, 75.4, 76.4, 79.2, 81.0, 84.4, 120.6, 128.6, 129.2, 130.1, 133.0, 133.7, 138.1, 142.3, 155.8, 166.9, 170.2, 172.8, 174.6, 203.8. HRMS: m/e calcd for C46H61NO15·H+: 868.4120. Found: 868.4120 (Δ = −0.1 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylprop-1-enyl)-3'N-de-tert-butoxycarbony-3'N-(cyclopropanecarbonyl)docetaxel (17h)

White solid; 66% (for 2 steps); mp 141–143 °C; [α]D20 −66.6 (c 11.4, CHC13); 1H NMR (300 MHz, CDC13): δ 1.14 (s, 3 H), 1.28 (m, 9 H), 1.67 (s, 3 H), 1.73 (s, 3 H), 1.77 (s, 3 H), 1.88 (m, 5 H), 2.35 (m, 5 H), 2.52 (m, 3 H), 3.80 (d, J = 6.8 Hz, 1 H), 3.85 (s, 3 H), 4.17 (d, J = 8.7, 1 H), 4.25 (d, J = 3.2 Hz, 1 H), 4.33 (d, J = 8.7 Hz), 4.41 (dd, J = 10.5 Hz, J =6.3 Hz, 1H), 4.96 (d, J =8.1 Hz, 1 H), 5.03 (td, J = 3.2, 8.8 Hz, 1 H), 5.37 (d, J = 8.8 Hz, 1 H), 5.66 (d, J = 6.8 Hz, 1 H), 5.92 (d, J = 8.1 Hz, 1 H), 6.15 (t, J= 8.4 Hz. 1 H), 6.30 (s, 1 H), 7.13 (dd, J = 7.8 Hz, J = 2.4Hz, 1 H), 7.3 (t, J = 7.8m Hz, 2 H) 7.62 (d, 2.4 Hz, 1 H), 7.68 (d, J = 7.8 Hz, 1 H). HRMS m/e calcd for C44H57NO15·H+: 840.3806. Found: 840.3802 (Δ = −0.5 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropen-l-yl)-3'N-dt-tert-butoxycarbony-3'N-(cyclobutanecarbonyl)docetaxel (17i)

White solid; 61% (for 2 steps); mp 143–145 °C; [α]D20 −70.4 (c 13.5, CHC13); 1H NMR (300 MHz, CDC13) δ 1.14 (s, 3 H), 1.28 (m, 8 H), 1.33 (s, 9 H), 1.67 (m, 3 H), 1.73 (s, 3 H), 1.75 (s, 3 H), 1.89 (m, 5 H), 2.37 (m, H), 2.52 (m, 3 H), 2.91 (m, 1 H), 3.80 (d, J = 6.9 Hz, 1 H), 3.85 (s, 3 H), 4.19 (d, J = 8.4, 1 H), 4.25 (s, 1 H), 4.34 (d, J = 8.4 Hz), 4.42 (t, J = 7.2, 1 H), 4.96 (d, J = 8.1 Hz, 1 H), 5.03 (dt, J = 3.3, 8.9 Hz, 1 H), 5.35 (d, J = 8.9 Hz, 1 H), 5.57 (d, J = 8.1 Hz, 1 H), 5.67 (d, J = 7.2 Hz, 1 H), 6.16 (t, J= 8.7 Hz. 1 H), 6.31 (s, 1 H), 7.13 (dd, J = 7.8 Hz, J = 1.8 Hz, 1 H), 7.37 (t, J = 7.8 Hz, 1 H) 7.63 (s, 1 H), 7.70 (d, J = 7.8 Hz, 1 H). HRMS m/e calcd for C45H59O15N·H+: 854.3962. Found: 854.3962 (Δ = 0.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropen-1-yl)-3'N-de-tert-butoxycarbony-3'N-(cyclopentanecarbonyl)docetaxel (17j)

White solid; 77% (for 2 steps); mp 136–138 °C; [α]D20 −74.5 (c 14.1, CHC13); 1H NMR (300 MHz, CDC13) δ 1.14 (s, 3 H), 1.28 (m, 8 H), 1.33 (s, 9 H), 1.67 (m, 3 H), 1.73 (s, 3 H), 1.75 (s, 3 H), 1.89 (m, 5 H), 2.37 (m, H), 2.52 (m, 3 H), 3.80 (d, J = 6.9 Hz, 1H), 3.85 (s, 3 H), 4.19 (d, J = 8.4, 1 H), 4.21 (s, 1 H), 4.32 (d, J = 8.4 Hz), 4.41 (t, J=10.2, 1 H), 4.96 (d, J = 8.4 Hz, 1 H), 5.03 (dt, J = 3.0, 8.7 Hz, 1 H), 5.36 (d, J = 9.0 Hz, 1 H), 5.66 (d, J = 6.9 Hz, 1 H), 5.72 (d, J = 8.1 Hz, 1 H), 6.13 (t, J= 8.7 Hz. 1 H), 6.31 (s, 1 H), 7.13 (dd, J = 7.5 Hz, J = 2.1Hz, 1 H), 7.37 (d, J = 7.5m Hz, 2 H) 7.63 (s, 1 H), 7.69 (d, J = 7.5 Hz, 1 H). HRMS m/e calcd for C46H61O15N·H+: 868.4119. Found: 868.4122 (Δ = −0.3 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropen-1-yl)-3'N-de-tert-butoxycarbony-3'N-(cyclohexanecarbonyl)docetaxel (17k)

White solid; 81% (for 2 steps); mp 151–153 °C; [α]D20 −73.4 (c 14.3, CHC13); 1H NMR (300 MHz, CDC13) δ 1.14 (s, 3 H), 1.28 (m, 8 H), 1.33 (s, 9 H), 1.67 (m, 3 H), 1.73 (s, 3 H), 1.75 (s, 3 H), 1.89 (m, 5 H), 2.37 (m, H), 2.52 (m, 3 H), 3.80 (d, J = 6.9 Hz, 1H), 3.86 (s, 3 H), 4.18–4.23 (m, 2 H), 4.33 (d, J = 8.7 Hz, 1 H), 4.41 (dt, J = 11.1, 6.6 Hz, 1 H), 4.95 (d, J=7.8, 1 H), 5.03 (dt, J = 2.7, 8.4 Hz, 1 H), 5.35 (d, J = 8.4 Hz, 1 H), 5.67 (d, J = 6.9 Hz, 1 H), 5.72 (d, J = 8.1 Hz, 1 H), 6.12 (t, J= 9.0 Hz. 1 H), 6.31 (s, 1 H), 7.13 (dd, J = 7.8 Hz, J = 2.1Hz, 1 H), 7.37 (d, J = 7.8 Hz, 2 H) 7.63 (s, 1 H), 7.69 (d, J = 7.8 Hz, 1 H). HRMS m/e calcd for C47H63O15N·H+: 882.4276. Found: 882.4275 (Δ = 0.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropen-1-yl)-3'N-de-tert-butoxycarbony-3'N-(cyclohexyloxycarbonyl)docetaxel (171)

White solid; 77% (for 2 steps); mp 141–143 °C; [α]D20 −68 (c 10.1, CHC13); 1HNMR (300 MHz, CDC13) δ 1.13 (s, 3 H), 1.28 (m, 8 H), 1.33 (s, 9 H), 1.66 (m, 3 H), 1.73 (s, 3 H), 1.75 (s, 3 H), 1.89 (m, 5 H), 2.37 (m, H), 2.52 (m, 3 H), 3.81 (d, J=7.2 Hz, 1 H), 3.87 (s, 3 H), 4.17 (d, 8.7 Hz, 1 H), 4.23 (d, J = 3.0 Hz, 1 H), 4.34 (d, J = 8.7 Hz), 4.42 (dd, J=10.8, 7.5 Hz, 1 H), 4.49 (m, 1 H), 4.79 (dt, J = 3.0, 8.0 Hz, 1 H), 4.88 (d, J = 8.7 Hz, 1 H), 4.96 (d, J = 7.5 Hz, 2H), 5.32 (d, J = 8.0 Hz, 1 H), 5.66 (d, J = 7.2 Hz, 1 H), 6.19 (t, J = 8.1 Hz. 1 H), 6.31 (s, 1 H), 7.13 (dd, J = 7.8 Hz, J = 2.1Hz, 1 H), 7.37 (d, J = 7.8Hz, 2 H) 7.63 (s, 1 H), 7.69 (d, J = 7.8Hz, 1 H). HRMS m/e calcd for C47H63O16N·H+: 898.4225. Found: 898.4226 (Δ = −0.1 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

Synthesis of Taxoids 18 through hydrogenation of 17

A typical procedure is described for the synthesis of 10-propanoyl-3'-dephenyl-3'-(2-methylpropyl)-3'N-debenzoyl-3'N-(cyclopentanecarbonyl)-docetaxel (18b): To a flask charged with activated 10% Pd/carbon (2 mg, 0.002 mmol) and 17b (13 mg, 0.02 mmol) under hydrogen atmosphere was added EtOAc (12.0 mL) and MeOH (0.02 mL) and the suspension was stirred at room temperature for 24 h. The reaction mixture was filtered over celite and the filtrate was concentrated in vacuo to afford 18b in quantitative yield as white solid: mp 149–152 °C; [α]D20 −114 (c 3.70, CHCl3); 1HNMR (300 MHz, CDC13) δ 0.98 (m, 6H), 1.16 (s, 3 H), 1.23 (m, 3 H), 1.26 (s, 9 H), 1.84–1.41 (m, 13 H), 1.88 (s, 3 H), 2.39 (s, 3 H), 2.43–2.59 (m, 3 H), 3.37 (d, J = 5.7 Hz, 1 H), 3.80 (d, J = 7.2 Hz, 1 H), 4.20 (m, 2 H), 4.30 (d, J = 8.1, 1 H), 4.42 (m, 2 H), 4.97 (d, J = 7.5 Hz, 1 H), 5.47 (d, J = 9.4 Hz, 1 H), 5.68 (d, J = 7.2 Hz, 1 H), 6.14 (m, 1 H), 6.30 (s, 1 H), 7.47 (t, 2 H), 7.60 (t, 1 H), 8.12 (d, 2 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.6, 14.9, 21.9, 22.0, 22.5, 23.3, 24.8, 25.8, 25.8, 26.7, 27.6, 29.7, 30.7, 35.6, 35.8, 41.0, 43.2, 45.6, 45.7, 49.6, 58.6, 72.2, 72.6, 72.9, 75.0, 75.4, 76.5, 79.1, 81.1, 84.4, 128.7, 129.2, 130.2, 133.1, 133.6, 142.2, 166.9, 170.1, 173.8, 174.6, 176.3, 203.8. HRMS: m/e calcd for C45H61NO14·H+: 840.4170. Found: 840.4174 (Δ = −0.4 ppm).

Other taxoids 18c, 18f–h, 18k and 18m were synthesized in the same manner and characterization data are shown below.

10-Propanoyl-3'-dephenyl-3'-(2-methylpropyl)-3'N-debenzoyl-3'N-cyclohexanecarbonyldocetaxel (18c)

White solid; 100%; mp 133–135 °C; [α]D20 −74 (c 4.21, CHC13); 1H NMR (300 MHz, CDC13) δ 1.08 (s, 3 H), 1.13–1.18 (m, 6 H), 1.28 (s, 9 H), 1.60 (s, 3 H), 1.69 (br, s, 6 H) 1.72 (m, 1 H), 1.83 (s, 3 H), 2.29 (s, 3 H), 2.31 (s, 2 H), 2.44 (m, 3 H), 3.38 (br s, 1 H), 3.74 (d, J = 6.9 Hz, 1H), 4.10 (d, J = 8.1 Hz, 1 H), 4.13 (br s 1 H), 4.22 (d, J = 8.1 Hz, 1 H), 4.33 (dd, J = 10.1, 7.5 Hz, 1 H), 4.67 (m, 2 H), 4.88 (d, J = 9.3 Hz, 1 H), 5.23 (d, J = 8.4 Hz, 1 H), 5.59 (d, J = 6.9 Hz, 1 H), 6.06 (m, 1 H), 6.24 (s, 1 H), 7.37 (t, 2 H), 7.51 (t, 1 H), 8.01 (d, 2 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.6, 14.9, 21.8, 22.0, 22.5, 23.3, 24.8, 25.6, 26.7, 27.6, 29.7, 35.6, 35.7, 41.0, 43.2, 45.5, 45.6, 49.2, 58.5, 72.2, 72.6, 72.8, 75.1, 75.4, 79.0, 81.1, 84.4, 128.6, 129.3, 130.2, 133.1, 133.6, 142.2, 166.8, 170.0, 173.8, 174.6, 176.1, 203.8. HRMS: m/e calcd for C45H61NO14·H+: 840.4170. Found: 840.4174 (Δ = −0.4 ppm). HPLC analysis using a Phenomenex Curosil-B column (CH3CN/water (2/3) as the solvent system with a flow rate of 1 mL/min; UV 230 nm and 254 nm) showed ≥95% purity.

10-Propanoyl-3'-dephenyl-3'-(2-methylpropyl)-3'N-debenzoyl-3'N-cyclopentyloxycarbonyldocetaxel (18f)

White solid; 100%; mp 131–133 °C; [α]D20 −67 (c 0.42, CHC13); 1H NMR (300 MHz, CDC13) δ 1.08 (s, 3 H), 1.13–1.18 (m, 6 H), 1.28 (s, 9 H), 1.60 (s, 3 H), 1.69 (br s, 6 H) 1.72 (m, 1 H), 1.83 (s, 3 H), 2.29 (s, 3 H), 2.31 (s, 2 H), 2.44 (m, 3 H), 3.38 (br s, 1 H), 3.74 (d, J = 6.9 Hz, 1H), 4.10 (d, J = 8.1 Hz, 1 H), 4.13 (br s 1 H), 4.22 (d, J = 8.1 Hz, 1 H), 4.33 (dd, J = 10.1, 7.5 Hz, 1 H), 4.67 (m, 2 H), 4.88 (d, J = 9.3 Hz, 1 H), 5.23 (d, J = 8.4 Hz, 1 H), 5.59 (d, J = 6.9 Hz, 1 H), 6.06 (m, 1 H), 6.24 (s, 1 H), 7.37 (t, 2 H), 7.51 (t, 1 H), 8.01 (d, 2 H); 13C NMR (63 MHz, CDC13) δ 9.0, 9.5, 14.9, 18.5, 21.9, 22.4, 23.6, 23.6, 25.7, 26.7, 27.5, 29.7, 32.6, 32.6, 35.6, 35.6, 43.2, 45.7, 51.9, 58.5, 72.1, 72.2, 73.7, 75.0, 75.4, 76.4, 77.9, 79.2, 81.1, 84.4, 120.5, 128.6, 129.2, 130.1, 133.0, 133.7, 138.2, 142.3, 156.1, 166.9, 170.2, 172.8, 174.6, 203.8. HRMS: m/e calcd for C45H61NO15·H+: 856.4119. Found: 854.4121 (Δ = −0.2 ppm).

10-Propanoyl-3'-dephenyl-3'-(2-methylpropyl)-3'N-debenzoyl-3'N-cyclohexyloxycarbonyldocetaxel (18g)

White solid; 100%; mp 142–144 °C; [α]D20 −60 (c 1.28, CHC13); 1HNMR (300 MHz, CDCl3) δ 1.08 (s, 3 H), 1.13–1.18 (m, 6 H), 1.28 (s, 9 H), 1.60 (s, 3 H), 1.69 (br, s, 6 H) 1.72 (m, 1 H), 1.83 (s, 3 H), 2.29 (s, 3 H), 2.31 (s, 2 H), 2.44 (m, 3 H), 3.38 (br s, 1 H), 3.74 (d, J = 6.9 Hz, 1H), 4.10 (d, J = 8.1 Hz, 1 H), 4.13 (br s 1 H), 4.22 (d, J = 8.1 Hz, 1 H), 4.33 (dd, J = 10.1, 7.5 Hz, 1 H), 4.67 (m, 2 H), 4.88 (d, J = 9.3 Hz, 1 H), 5.23 (d, J = 8.4 Hz, 1 H), 5.64 (d, J = 7.0 Hz, 1 H), 6.18 (m, 1 H), 6.30 (s, 1 H), 7.46 (t, 2 H), 7.59 (t, 1 H), 8.09 (d, 2 H); 13C NMR (63 MHz, CDC13) – 9.0, 9.5, 14.9, 18.5, 21.9, 22.4, 23.6, 23.7, 25.3, 25.7, 26.7, 27.5, 31.8, 35.6, 35.6, 43.2, 45.7, 51.8, 58.5, 72.1, 72.2, 73.7, 75.0, 75.4, 76.4, 79.2, 81.0, 84.4, 120.6, 128.6, 129.2, 130.1, 133.0, 133.7, 138.1, 142.3, 155.8, 166.9, 170.2, 172.8, 174.6, 203.8. HRMS: m/e calcd for C46H63NO15·H+: 870.4273. Found: 870.4276 (Δ = + 0.3 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropyl)-3'N-de-tert-butoxycarbony-3'N-(cyclopropanecarbonyl)docetaxel (18h)

White solid; 100%; mp 139–141 °C; [α]D20 −74.5 (c 14.0, CHC13); 1HNMR (400 MHz, CDC13) δ 0.71 (m, 2H), 0.86 (m, 2H), 0.95 (dd, J1 = 6.4 Hz, J2 = 16.4Hz, 6 H), 1.15 (s,3H), 1.21 – 1.41 (m, 8 H), 1.62 – 1.78 (m, 5 H), 1.88 (m, 3H), 2.38 (m, 4H), 2. 54 (m, 4H), 3.66 (bs, 1H), 3.79 (d, J = 7.2 Hz, 1 H), 3.87 (s, 1H), 4.20 (m, 2 H), 4.32 (d, J= 8.4 Hz, 1 H), 4.42 (m, 2 H), 4.96 (dd, J1 = 2 Hz, J2 = 9.6 Hz, 1 H), 5.67 (d, J = 7.2 Hz, 1H), 5.78 (d, J = 9.2 Hz, 1H), 6.14 (m, 1 H), 6.30 (s, 1 H), 7.14 (bdd, 1 H), 7.37 (bt, J = 8 Hz, 1 H), 7.63 (bs, 1H), 7.70 (bd, J = 7.6 Hz, 1 H); 13C NMR (100 MHz, CDC13) δ 7.3, 7.5, 9.0, 9.6, 14.5, 14.8, 21.8, 22.0, 22.5, 23.2, 24.7, 26.7, 27.6, 35.6, 35.7, 40.9, 43.2, 45.5, 50.0, 55.3, 58.5, 72.1, 72.4, 73.0, 75.1, 75.4, 76.4, 78.9, 81.1, 84.4, 114.3, 120.4, 122.6, 129.6, 130.5, 133.0, 142.2, 159.6, 166.7, 170.0, 173.7, 173.8, 174.6, 203.8. HRMS m/e calcd for C44H59O15N·H+: 842.3963. Found: 842.3941 (Δ = −2.6 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropan-1-yl)-3'N-de-tert-butoxycarbony-3'N-(cyclohexanecarbonyl)docetaxel (18k)

White solid; 100%; mp 143–145 °C; [α]D20 −85 (c 14.0, CHC13); 1H NMR (400 MHz, CDC13) δ 0.95 (dd, J1 = 6.4 Hz, J2 = 16.4 Hz, 6 H), 1.15 – 1.37 (m, 16 H), 1.57 – 1.40 (m, 10 H), 1.87 (m, 5H), 2.38 (m, 5H), 2.54 (m, 4H), 3.81 (d, J = 6.8 Hz, 1 H), 3.87 (s, 3H), 4.20 (m, 2 H), 4.33 (d, J = 8.4 Hz, 1 H), 4.43 (m, 2H), 4.97 (bd, J = 9.6 Hz, 1 H), 5.47 (d, J = 9.6 Hz, 1H), 5.68 (d, J = 7.2 Hz, 1H), 6.12 (m, 1 H), 6.30 (s, 1 H), 7.14 (dd, J1 = 2 Hz, J2 = 8.4 Hz, 1 H), 7.37 (bt, J = 8 Hz, 1 H), 7.64 (bs, 1H), 7.70 (bd, J = 7.6 Hz, 1 H); 13C NMR (100 MHz, CDC13) d 9.0, 9.5, 14.8, 21.7, 22.0, 22.5, 23.3, 24.8, 25.5, 25.6, 26.7, 27.6, 29.7, 35.6, 35.7, 41.0, 43.2, 45.4, 45.5, 49.2, 55.3, 58.5, 72.2, 72.6, 72.9, 75.1, 75.4, 76.4, 78.8, 81.2, 84.4, 114.2, 120.4, 122.7, 129.6, 130.5, 133.2, 142.1, 159.6, 166.6, 170.0, 173.7, 174.6, 176.1, 203.8. HRMS m/e calcd for C47H65NO15·H+ 884.4432. Found: 884.4413 (Δ = −2.1 ppm).

2-Debenzoyl-2-(3-methoxybenzoyl)-10-propanoyl-3'-dephenyl-3'-(2-methylpropyl)-3'N-de-tert-butoxycarbony-3'N-(cyclopentyloxycarbonyl)docetaxel (18m)

White solid; 60 % yield (for 3 steps from 11); mp 143–145 °C; [α]D20 −70 (c 13.0, CHC13); 1H NMR (400 MHz, CDC13) δ 0.95 (dd, J1 = 6.4 Hz, J2 = 10 Hz, 6 H), 1.14 (s, 3 H), 1.25 (m, 7 H), 1.35 – 1.72 (m, 15 H), 1.85 (s, 3H), 2.36 (m, 5H), 2.54 (m, 4H), 3.81 (d, J = 8 Hz, 1 H), 3.87 (s, 1H), 4.12–4.20 (m, 3 H), 4.33 (d, J= 8.4 Hz, 1 H), 4.42 (m, 1 H), 4.69 (d, J = 9.6 Hz, 1 H), 4.89 (m, 1 H), 4.98 (d, J1 = 8 Hz, 1 H), 5.66 (d, J = 7.2 Hz, 1H), 6.20 (m, 1 H), 6.30 (s, 1 H), 7.13 (dd, J1 = 2.8 Hz, J2 = 8.4 Hz, 1 H), 7.37 (bt, J = 8 Hz, 1 H), 7.64 (bs, 1H), 7.70 (bd, J = 7.6 Hz, 1 H); 13C NMR (100 MHz, CDC13) δ 9.0, 9.5, 14.8, 21.7, 22.0, 22.5, 23.2, 23.5, 23.6, 24.6, 26.7, 27.5, 32.5, 32.6, 35.5, 41.2, 43.2, 45.6, 51.6, 55.3, 58.5, 72.1, 72.5, 73.0, 75.1, 75.4, 76.4, 77.8, 79.1, 81.1, 84.4, 114.2, 120.5, 122.7, 129.6, 130.4, 133.0, 142.3, 156.1, 159.7, 166.8, 170.0, 173.7, 174.6, 203.8. HRMS m/e calcd for C46H63O16N·H+: 886.4225. Found: 886.4211 (Δ = −1.6 ppm).

In vitro cell growth inhibition assay

(a) Tumor cell growth inhibition was determined according to the method established by Skehan et al.51 Human cancer cells LCC6-WT (Pgp−), MCF-7 (Pgp−), LCC6-MDR (Pgp+) and NCI/ADR (Pgp+), were plated at a density of 400–2,000 cells/well in 96-well plates and allowed to attach overnight. These cell lines were maintained in RPMI-1640 medium (Roswell Park Memorial Institute growth medium) supplemented with 5% fetal bovine serum and 5% Nu serum (Collaborative Biomedical Product, MA). Taxoids were dissolved in DMSO and further diluted with RPMI-1640 medium. Triplicate wells were exposed to various treatments. After 72 h incubation, 100 µL of ice-cold 50% trichloroacetic acid (TCA) was added to each well, and the samples were incubated for 1 h at 4 °C. Plates were then washed five times with water to remove TCA and serum proteins, and 50 µL of 0.4% sulforhodamine B (SRB) was added to each well. Following a 5 min incubation, plates were rinsed five times with 0.1% acetic acid and air-dried. The dye was then solubilized with 10 mM Tris base (pH 10.5) for 5 min on a gyratory shaker. Optical density was measured at 570 nm. The IC50 values were then calculated by fitting the concentration-effect curve data with the sigmoid-Emax model using nonlinear regression, weighted by the reciprocal of the square of the predicted effect.52

(b) Human pancreatic cancer cell lines MIA PaCa-2, CFPAC-1, BxPC-3, and PANC-1 were cultured as specified by ATCC (Manassas, Virginia). For cytotoxicity assays the cells were plated at a density of 2.5 × 104 cells/well in 24-well plates and allowed to adhere overnight. The media was changed the following morning and replaced with media containing taxane derivatives or vehicle control. Taxoids were dissolved in DMSO to 10 mM concentration and were further diluted in appropriate media prior to addition to cells. Each dose of drug or vehicle was tested in triplicate, and the experiment is representative of at least 3 independent trials. After 72 hours of treatment the media was aspirated and the cells were washed in warm PBS. MTT reagent (Sigma) was diluted in RPMI-1640 media without phenol red (Invitrogen), and added to the cells at a concentration of 0.5 mg/mL. After 3 hours of incubation, the reagent was aspirated, the plate was washed with PBS and MTT formazan crystals were dissolved in 500 µL acidified isopropanol (0.1N hydrochloric acid). Subsequently, 100 µL of the solution was transferred to a microtiter plate. Absorbance at 570 nM was measured on a thermomax plate reader (Molecular Devices). The IC50 values were obtained by using the same method as that described for (a).

Tubulin polymerization assay

Assembly and disassembly of calf brain microtubule protein (MTP) was monitored spectrophotometrically (Beckman Coulter DU 640, Fullerton, CA) by recording changes in turbidity at 350 nm at 37 °C.53,54 MTP was diluted to 1mg/mL in MES buffer containing 3 M glycerol. The concentration of tubulin in MTP is 85% and that is taken into consideration when the ratios of tubulin to drug are presented in Figure 1 and Figure 22. Microtubule assembly was carried out with 10 µM taxoid (19, 4g or 14i). Paclitaxel (10 µM) was also used for comparison purpose. Calcium chloride (6 mM) was added to the assembly reaction after 50 min to follow the calcium-induced microtubule depolymerization. For the experiments shown in Figure 3, tubulin stored in liquid nitrogen was centrifuged just before use and protein concentration adjusted to 1 mg/mL, while the experiments in Figure 4 used tubulin stored in liquid nitrogen as it was at 1 mg/mL concentration.

Electron microscopy

Aliquots (50 µL) were taken from in vitro polymerization assays at the end of the reaction and placed onto 300-mesh carbon-coated, formavar-treated copper grids. Samples were then stained with 20 µL of 2% uranyl acetate and viewed with a JEOL model 100CX electron microscope.

Animals and tumor xenografts

Female severe combined immune deficient, (SCID) mice aged six to eight weeks were obtained from the National Cancer Institute (Frederick, Maryland), were housed and monitored at the Medical Research Complex at Roswell Park Cancer Institute. All experimental procedures and protocols were approved by the Institutional Animal Care and Use Committee. The human colon tumor DLD-1 which expresses pgp, were initiated by implantation of approximately 50 mg of non-necrotic tumor fragments on the right flank using a 12-guage trocar needle. Chemotherapy was started when the tumor was established as a palpable mass, (approximately 50–100 mm3 size), 5 days after implantation and continued either every three days or weekly. Each drug treatment group or drug free vehicle consisted of 5 mice per group; untreated controls contained 10 mice per group.

Drug preparation for in vivo experiments

Taxoid 19 was prepared as a 30 mg/mL stock solution in equal parts of Tween 80 (polyoxyethylene-sorbitan monooleate; purchased from Sigma Chemical Company) and absolute ethanol. Each stock solution was further diluted before use in 0.9% NaCl (saline) so that the appropriate concentration of each drug could be injected i.v. via the tail vein, in a volume of approximately 0.4 mL for a 20 g mouse.

In vivo tumor growth inhibition assay against DLD-1 tumor xenograft

For each animal, the tumor length (1) and width (w), each in mm, were measured using electronic calipers and recorded every 3–4 days. Tumor volume (v), in mm3, was calculated using the formula: v = 0.4(1 × w2). The time in days to the pre-determined target tumor volume of 600 mm3 was linearly interpolated from a plot of log (volume) versus time. Statistically significant differences in tumor volumes between control and drug-treated mice were determined by the Cox-Mantel test. For the Cox-Mantel test, the time-to-event data for animals that did not reach the target tumor volume, either because of long-term cure (defined as those animals that were still alive at the conclusion of the experiment whose tumors either completely regressed or did not reach the pre-set target volume) or early death due to drug toxicity, were treated as censored data. All statistical tests were two-sided.

In vivo efficacy assay of taxoid 19 against CFPAC-1 tumor xenograft

A preliminary in vivo efficacy evaluation of taxoid 19 was also performed against human pancreatic cancer xenograft in male Swiss nude mice aged six to eight weeks obtained from Taconic Farms (Hudson, New York). The human pancreatic cancer cell line CFPAC-1, which expresses high levels of mdrl, was used for the experiments. CFPAC-1 cancer cells (1 × 106) were resuspended in 200 µL of PBS and injected bilaterally into the flanks of mice using a 26 gauge needle. Treatment started when the tumor was established as a palpable mass (approximately 100 mm3 in size) 19 days after the injection. Taxoid 19 was prepared as a 30 mg/mL stock solution in Tween 80 and absolute ethanol (1:1). Each stock solution was further diluted by PBS prior to administration so that appropriate concentration of each drug could be injected in a final volume of 0.2 mL. The taxoid was administered i.v. using tail vein injection three times at 3 day intervals on day 19, 22 and 25 (20 mg/kg × 3; total dose 60 mg/kg). The tumor size was measured and the volume calculated in the same manner as that described above.

Supplementary Material

Supporting Information

Supporting Information Available:

1H and 13C NMR spectra of new taxoids and characterization data for synthetic intermediates. This material is available free of charge via the Internet at http://pubs.acs.org.

Acknowledgments

This work was supported by grants from the National Institutes of Health (CA103314 and GM42798 to I.O.; CA083185 and CA077263 to S.B.H.; CA55360 and CA28146 to D.B.-S;CA 73872 to R.J.B.), a Targeted Research Opportunities grant from School of Medicine, State University of New York at Stony Brook, and the Lustgarten Foundation (to D.B.-S.). Generous support from Indena SpA is also gratefully acknowledged.

Abbreviations

DAB
10-deacetylbaccatin III
IC50
half maximal inhibitory concentration
LiHMDS
lithium hexamethyldisilazide
NMO
N-methylmorpholine-N-oxide
PBS
phosphate buffered saline
PDA
photodiode-array
Pgp
P-glycoprotein
R/S
resistance factor (the ratio of the IC50 value against the drug-resistant cell line vs. that against the drug-sensitive cell line)
RT-PCR
reverse transcriptase-polymerase chain reaction
SAR
structure-activity relationship
SCID
severe combined immunodeficiency
TEA
triethylamine
TES
triethylsilyl
TIPS
triisopropylsilyl
TLC
thin layer chromatography
TPAP
tetrapropylammonium perruthenate
WT
wild type

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