Direct, efficient and selective tritiations of paclitaxel and photoaffinity taxoids

Article abstract of DOI:10.1016/S0040-4039(00)01652-X

Radiolabeled paclitaxel and three photophore-bearing derivatives were prepared by organoiridium-mediated direct tritium exchange under both catalytic and stoichiometric conditions. The resulting tritiated taxoids had specific activities ranging from 53 to 195 Ci/mmol. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4039(00)01652-X

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 41 (2000) 9015–9019
Direct, efficient and selective tritiations of paclitaxel and
photoaffinity taxoids
Arthur Y. L. Shu* and J. Richard Heys
Radiochemistry Section, Department of Synthetic Chemistry, SmithKline Beecham Pharmaceuticals, PO Box 1539,
King of Prussia, PA 19406, USA
Received 12 September 2000; revised 20 September 2000; accepted 21 September 2000
Abstract
Radiolabeled paclitaxel and three photophore-bearing derivatives were prepared by organoiridium-
mediated direct tritium exchange under both catalytic and stoichiometric conditions. The resulting
tritiated taxoids had specific activities ranging from 53 to 195 Ci/mmol. © 2000 Elsevier Science Ltd. All
rights reserved.
Keywords: organoiridum-mediated tritium exchange; paclitaxel; photoaffinity taxoids; Crabtree’s catalyst.
Azidophenyl, benzoylphenyl and diazirenyl groups are among the most common examples of
photophores used in photoaffinity labeling; used in tagged form, usually with a radiolabel, they
are powerful tools for the identification of cognate macromolecules.1 Although the incorpora-
tion of photophores into ligands of interest can be achieved by straightforward chemical means,2
preparations of their radiolabeled counterparts, especially in high specific activities, can be
challenging.
Our laboratory has been engaged in labeling aromatic molecules using organoiridium-medi-
ated exchange in the presence of isotopic hydrogen gas.3 The advantage of this method is that
it allows the direct utilization of substrates already available, without resorting to the separate
and sometimes elaborate synthesis of precursors, such as unsaturated or multi-halogenated
derivatives. In addition, high levels of isotopic content can often be realized even when only a
limited amount of deuterium or tritium gas is utilized. This methodology requires the aromatic
substrate to possess a functionality capable of coordinating with the iridium center, which
directs the substrate’s ortho hydrogens to undergo exchange through a metallacycle intermedi-
ate. Our aim was to extend this approach to labeling photophore substructures of complex
ligands. We selected paclitaxel and several photoreactive derivatives as test platforms for testing
the organoiridium-mediated direct tritium exchange.
* Corresponding author. Tel: (610)270-4373; fax: (610)270-4110; e-mail: art shu-1@sbphrd.com
–
0040-4039/00/$ - see front matter © 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(00)01652-X

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The mechanism of action of paclitaxel (1)4 was in part elucidated with the use of radiolabeled
photoaffinity derivatives 2 and 3, with azido and benzoyl groups attached to the para position
of the sidechain C-3% benzamide, respectively;5 other paclitaxel derived photoaffinity analogs in
tritiated forms have since emerged.6 These were prepared either by multistep sequences involving
handling of radioactive intermediates or separate syntheses of unsaturated precursors prior to
tritiation. The iridium-mediated isotopic exchange could lend itself to tritiating these analogs
directly, providing the stabilities and directing abilities of various photoactivating groups could
be calibrated, and the complex core structure of paclitaxel could remain intact under the
exchange conditions.
Our previous experience with benzophenones indicated that the benzoylphenyl moiety is
stable to the exchange process and is a good directing group.7 In a model study, treatment of
a solution of N-boc-4-benzoyl- -phenylalanine (5) in CH2Cl2 with [(COD)Ir(PPh3)2]BF4 (6, 0.3
L
equiv.) and 1.8 Ci of carrier-free T2 (specific activity 29 Ci/milliatom T) for 18 h resulted in [3H]5
with a specific activity of 46 Ci/mmol and tritium distribution ratio at Ta:Tb of 1:1.5.8
The azido group was unstable to the exchange conditions and devoid of directing ability.
However, in the presence of a catalytic amount of iridium complex and a strong directing group
such as an amide, exchange adjacent to the latter function can be successful. For example,
although the activated ester 7b was not labeled under a variety of conditions, exposure of
pyrrolidine 4-azidobenzamide (7a) to 0.2 equiv. of 6 and an excess of D2 for 18 h afforded 63%
of 7a with the positions ortho to the amide fully deuterated.
Likewise, the diazirenyl group was compatible with the exchange conditions, but had no
directing ability. When pyrrolidine 4-diazirenylbenzamide (8a) was exposed to either stoichio-
metric or catalytic amounts (0.2 equiv.) of complex 6 and an excess of D2, the exchanged
product was recovered in high yields. Only the positions ortho to the aminocarbonyl function
were fully deuterated. Similar to 7b, succinyl 4-diazirenylbenzoate (8b) failed to incorporate any
deuterium.
With some experimental parameters defined for these three photoactivating groups, we turned
our attention to the feasibility of performing direct isotopic exchange on paclitaxel. When 1 (2
mg) was treated with complex 6 (0.3 equiv.) in the presence of 4 Ci of carrier-free T2 for 18 h,
the purified [3H]1 had specific activity of 53 Ci/mmol with tritiation occurring exclusively at the
ortho positions of the sidechain C-3% benzamide.9 Various ways of making [3H]1 have previously

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been reported,10 but none provided such a high level of label incorporation in a single step. In
agreement with the results from our model studies described above and our previous findings,11
the sidechain C-3% benzamide, being a stronger Lewis base than the C-2 benzoate, was the sole
directing group under these catalytic conditions. Increasing the iridium complex loading to 1.5
equiv. and using Crabtree’s catalyst12 resulted in further tritiation of paclitaxel [3H]1a at the
ortho positions (Tc) in the C-2 benzoate ring as well as sidechain C-3% benzylic site (Tb) in the
ratio of 78:7:15 for Ta:Tb:Tc, respectively. The specific activity was raised to 61 Ci/mmol.
To access photophore-bearing derivatives 2–4, we chose Ojima’s chiral b-lactam semisynthetic
approach.13 (−)-(3R,4S)-b-lactam 9 was acylated with the corresponding para substituted
benzoyl chlorides to 1-acyl-b-lactams 10–12. These b-lactams, having the C-3 hydroxyl protected
as the TIPS ether, were coupled directly with 7-TES-baccatin-III using LiHMDS in THF to
provide doubly silylated taxoids 13–15.14 Desilylation of 13–15 in the presence of HF/pyridine
in a mixture of pyridine and acetonitrile gave the desired substrates 2–4 (Scheme 1).
Scheme 1.
Each of the photoaffinity analogs 2–4 was subjected to the catalytic organoiridium-mediated
tritium exchange conditions described above for parent paclitaxel, except for replacement of
complex 6 with Crabtree’s catalyst. The resulting products all had tritium incorporated at the
ortho positions of the sidechain C-3% benzamide. Additional tritium was observed in [3H]3 (23%
of total) at the combined sites Td and Te that are ortho to the benzophenone carbonyl, reflecting
its intermediate directing ability between the C-3% benzamide and the C-2 benzoate. The specific
activities of [3H]2–4 ranged from 56 to 71 Ci/mmol.
Taxoids 3 and 4, having photophores that are compatible with the exchange conditions, were
additionally exposed to high-loading iridium complex tritiation (3 and 1.5 equiv., respectively).

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The resulting exchanged products exhibited similar labeling patterns as [3H]1a. Diazirene-bear-
ing analog [3H]4a was labeled at the sites Ta:Tb:Tc in the ratio of 67.8:3.5:28.7 with a specific
activity of 82 Ci/mmol. The isotopic distribution for [3H]3a was more complex. Substantial
tritium buildup occurred at the benzophenone–carbonyl directed sites Td and Te, resulting in
the ratio of 28:3:13:30:26 for Ta:Tb:Tc:Td:Te. When the amount of iridium complex was
elevated further to 5 equiv., a noticeable increase of tritium content took place proportionally
at sites Tb and Tc,15 with a ratio of 26(Ta):4(Tb):18(Tc):27(Td):25(Te). The specific activity was
raised from 179 to 195 Ci/mmol.
In summary, organoiridium-mediated direct isotope exchange is a powerful methodology for
labeling compounds with complex structures regioselectively and to high specific activity, as
demonstrated in the one-step tritiations of paclitaxel and related photoaffinity analogs.
Acknowledgements
3
We thank S. H. Levinson, A. L. Freyer and F. G. Vogt for H NMR measurements, L. B.
Killmer for MS measurements, and C. R. Newsome for chiral HPLC assays on b-lactam
intermediates.
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3
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