Toward aplyronine payloads for antibody-drug conjugates: Total synthesis of aplyronines A and D

Article abstract of DOI:10.1039/c7ob03204h

The aplyronines are a family of antimitotic marine macrolides that disrupt cytoskeletal dynamics by dual targeting of both actin and tubulin. Given their picomolar cytotoxicity profile and unprecedented mode of action, the aplyronines represent an excellent candidate as a novel payload for the development of next-generation antibody-drug conjugates (ADCs) for cancer chemotherapy. Enabled by an improved second-generation synthesis of the macrolactone core 5, we have achieved the first total synthesis of the most potent congener aplyronine D together with a highly stereocontrolled synthesis of aplyronine A. To facilitate step economy, an adventurous site-selective esterification of the C7 hydroxyl group was performed to install the N,N,O-trimethylserine pharmacophore to directly afford aplyronines A and D. Toward the assembly of ADCs incorporating an aplyronine warhead, the C29-ester derivative 4 featuring an Fmoc-amino substituted linker attached to the actin-binding tail region was also prepared by adapting this flexible endgame.

Full text of DOI:10.1039/c7ob03204h

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Toward Aplyronine Payloads for Antibody-Drug Conjugates: Total
Synthesis of Aplyronines A and D
Nika Anžiček,† Simon Williams,† Michael P. Housden and Ian Paterson*[a]
Received 00th January 20xx,
Accepted 00th January 20xx
The aplyronines are a family of antimitotic marine macrolides that disrupt cytoskeletal dynamics by dual targeting of both
actin and tubulin. Given their picomolar cytotoxicity profile and unprecedented mode of action, the aplyronines represent
an excellent candidate as a novel payload for the development of next-generation antibody-drug conjugates (ADCs) for
cancer chemotherapy. Enabled by an improved second-generation synthesis of the macrolactone core 5, we have achieved
the first total synthesis of the most potent congener aplyronine D together with a highly stereocontrolled synthesis of
aplyronine A. To facilitate step economy, an adventurous site-selective esterification of the C7 hydroxyl group was
performed to install the N,N,O-trimethylserine pharmacophore to directly afford aplyronines A and D. Toward the
assembly of ADCs incorporating an aplyronine warhead, the C29-ester derivative 4 featuring an Fmoc-amino substituted
linker attached to the actin-binding tail region was also prepared by adapting this flexible endgame.
DOI: 10.1039/x0xx00000x
www.rsc.org/
represented by aplyronines A (1) and D (2), also feature a signature
N,N,O-trimethylserine residue attached to C7, which is generally
present as a mixture of epimers. The configurations of the five
Introduction
alkenes and 17 stereocentres in aplyronine A were determined by
detailed spectroscopic analysis in combination with chemical
degradation and correlation with fragments prepared by
enantioselective synthesis.4 The resulting stereochemical
assignment was validated by the first total synthesis5 and the X-ray
crystallographic analysis of its 1:1 complex with actin.6 Notably, the
eight known members of the aplyronine family differ in the identity
of the amino acid at C29 (N,N-
Natural products offer a rich source of potent bioactive compounds
for the development of new drugs, particularly cytotoxic anticancer
agents.1 Their therapeutic window can be enhanced by targeted
drug delivery to tumour cells, where natural product derivatives
such as the tubulin-binding agents maytansine and monomethyl
auristatin E are clinically validated cytotoxic payloads for antibody-
drug conjugates (ADCs).2 Other highly potent anticancer natural
products with unique mechanisms of action have potential to be
developed as novel ADC payloads, provided they can be obtained in
useful quantities and modified to include a suitable linker for
bioconjugation.
Me
2 N
R
R1
OAc
O
O
OR3
7
OH
O
OH
O
Me
N
1
A promising novel chemotype for targeted delivery by ADCs are the
aplyronines (Figure 1). These complex polyketide metabolites were
isolated by Yamada and co-workers3 from the sea hare Aplysia
kurodai, a gastropod mollusc collected off the coast of the Mie
prefecture in Japan. From a molecular architecture perspective, the
aplyronines are characterised by a highly substituted 24-membered
macrolactone core, with an elaborate side chain terminating in an
N-methyl-N-vinyl formamide moiety and an amino acid residue
attached to C29. The most biologically active congeners, as
29
CHO
23
34
MeO
OMe
1: aplyronine A, R1 = R2 = Me, R3 = N,N,O-trimethylserine
2: aplyronine D, R1 = H, R2 = Me, R3 = N,N,O-trimethylserine
3: aplyronine C, R1 = R2 = Me, R3 = H
4: R1 = H, R2 = (CH2)6NHFmoc, R3 = N,N,O-trimethylserine
Figure 1 Structures of the aplyronine congeners 1–3 and the linker-
modified derivative 4.
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dimethylalanine, N-methylalanine or N,N-dimethylglycine) and derivative
Journal Name
4
for planned antibody conjugation, and the
whether the N,N,O-trimethylserine is attached at C7 or C9, or development of an improved route to the pivotal macrolactone
omitted altogether. These permutations in the position and identity core.
of the amino acids can lead to marked SAR differences, e.g.
aplyronine C (3) lacking the N,N,O-trimethylserine moiety is around
Results and discussion
two orders of magnitude less cytotoxic than aplyronines A and D.
Me
2 N
R
esterification
R1
esterification
The aplyronines represent potential clinical candidates in cancer
chemotherapy, although there are severe supply issues impacting
on their further development. On testing in the US National Cancer
O
1
O
R3
OH
O
7
O
OH
O
OAc
Me
N
27
CHO
29
23
Institute primary screen, aplyronine
A
(NSC687160) displayed
34
aldol
impressive sub-nanomolar antiproliferative activity (mean GI50 = 0.2
nM) in the 60 cell line panel.2a Moreover, the in vivo efficacy of
aplyronine A was demonstrated against a range of five implanted
tumour types in mouse xenograft models, with high survival rates
observed for Lewis lung carcinoma and P388 leukaemia.3a,b
MeO
OMe
Me2N
1: aplyronine A, R1 = R2 = Me, R3
2: aplyronine D, R1 = H, R2 = Me, R3
3: aplyronine C, R1 = R2 = Me, R3 = H
=
OMe
=
1:1 S:R
O
Intriguingly, the minor congener aplyronine D, displaying
a
structural variation in the amino acid residue at C29, was later
reported to possess even greater cytotoxicity than aplyronine A
(e.g. GI50 of 0.071 nM for 2 vs 0.45 nM for 1 in the HeLa-S3 cervical
cancer cell line).3c The aplyronines are known to primarily function
as small molecule mimics of actin-binding proteins,6-8 disrupting
actin dynamics by severing F-actin and sequestering G-actin. While
the details of the actin-binding interaction of the aplyronines have
been well characterised, the exceptional cytotoxicity has recently
been shown to arise from interference with protein-protein
interactions, through the formation of a heterotrimeric complex of
aplyronine A with actin and tubulin, synergistically disrupting
cytoskeletal dynamics.9 Although the precise nature of these
interactions and the associated binding site on tubulin of the
initially formed actin-aplyronine complex are yet to be determined,
this dual protein-targeting mode is unprecedented amongst
antimitotic natural products.10 The combination of exceptional
potency and a unique mechanism of action make the aplyronines
important lead compounds, both as molecular probes to further
explore their interaction with actin and tubulin, and in the context
of providing a new class of cytotoxic payload for next-generation
ADC development.
macrolactonisation
O
TES
O
TES
TES
1
O
O
O
O
OAc
Me
N
27
OH
CHO
28
34
5
6
MeO
TES
OMe
HWE olefination
TES
O
O
1
OTBS
8
PMB TES
O
O
OMe
O
27
O
14
OEt
H
15
23
OAlloc
P
O
OEt
7
Scheme 1 Retrosynthetic analysis highlighting the key fragments
5–8.
As outlined in Scheme 1, our proposed unified synthesis plan for the
aplyronines was focused on the controlled construction and
assembly of the key fragments 5–8, resulting from disconnections
at C27–C28 and C14–C15, and opening of the macrolactone ring.
Importantly, it was considered essential to be able to site-
selectively install the pharmacophoric N,N,O-trimethylserine
residue at C7. To simplify the protecting group strategy for
enhanced step economy, an adventurous esterification onto the C7
Given their extremely scarce natural abundance (e.g. only 2.6 mg of
aplyronine D was isolated from a 300 kg collection of Aplysia
kurodai3c) and structural complexity, the aplyronine macrolides
constitute compelling and challenging synthetic targets.11-13
Notably, the total synthesis of aplyronines A, B and C was first
achieved by the Yamada group,5 with an improved second-
generation route to aplyronine A subsequently reported by Kigoshi
and co-workers.11 Recently, we accomplished a convergent total
synthesis of aplyronine C based on asymmetric aldol chemistry
developed in our group, leading to a significantly shorter route.12a
Moving forward, we aimed to devise a flexible and practical
synthetic strategy to access the most potent aplyronine congeners
and designed analogues for future application as ADC payloads. We
now report an expedient total synthesis of both aplyronines A and
D via a late-stage divergence, together with a linker-modified
alcohol in aplyronine
C (3) was proposed to directly afford
aplyronine A (1). To further streamline the route, we elected to set
out by having all three secondary hydroxyl groups at C7, C9 and C25
in the macrolactone core 5 protected as TES ethers. Building on our
first-generation route,12a the full aplyronine side chain would then
be installed by a late-stage aldol coupling14 to forge the C27–C28
bond between the derived C27 aldehyde and the methyl ketone 6.
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In our previous work, the macrolactonisation of an intermediate The required southern fragment 7 for the planned HWE-type
23,25-dihydroxy acid had generated exclusively the incorrect ring fragment coupling with the phosphonate 8 was assembled from the
size,12b which was followed by Ti(OiPr)4-mediated isomerisation to C21–C27 aldehyde 14 (Scheme 3). Installation of the stereotetrad
give
a 3:1 mixture in favour of the desired 24-membered region of 14 commenced with a titanium-mediated aldol reaction
macrolactone. The resulting requirement for repeated between the chiral ethyl ketone 1516 and aldehyde 16. High
chromatographic separation and re-equilibration of the isomeric diastereoselectivity for the desired syn-adduct 17 (82%, 12:1 dr)
26-membered macrolactone impacted on material throughput. To was achieved by employing Ti(OiPr)Cl3 as the Lewis acid17 to ensure
circumvent this difficulty, we now planned to achieve a site-specific
macrocyclisation step to give solely the desired 24-membered
macrolactone. Hence, the C23 hydroxyl would be orthogonally
protected as its PMB ether in the C15–C27 aldehyde 7. To connect
the southern and northern fragments, we envisaged that this
aldehyde would engage in an (E)-selective HWE-type coupling with
a revised C1–C14 phosphonate 8.
efficient chelation by the benzyl ether of 15 in the proposed bicyclic
aldol transition state. This procedure proved robust on a multi-gram
scale, affording greater diastereoselectivity and operational
simplicity than the previously employed Sn(OTf)2-promoted aldol
reaction.12c,18 Next, an Evans-Tishchenko reduction19 (SmI2, EtCHO)
of the β-hydroxy ketone 17 efficiently configured the C25
stereocentre (93%, >20:1 dr), and the resulting alcohol was
Following this flexible and modular plan, we commenced our
synthesis campaign with the preparation of the northern fragment
8 (Scheme 2). In practice, this was obtained from the 1,3-diol 9,
incorporating the correctly configured C7–C10 stereotetrad, which
was readily prepared on a multigram scale (see the ESI) by our
previously described boron-mediated aldol route12c from the chiral
ketone 10. The diol 9 was first converted into the bis-TES ether
(TESOTf), followed by cleavage of the PMB ether (DDQ) and
transformation of the resulting alcohol (Ph3P, I2, imid.) into the
iodide 11 (55%, 3 steps). Adjustment of the oxidation state at C1
(DIBAL) and silylation of the resulting alcohol (TBSCl) then afforded
n
12, which was alkylated at C11 with the dianion (NaH, BuLi) of 1315
to deliver the desired β-keto phosphonate 8 (54%, 3 steps)
accompanied by a minor regioisomer that was best removed after
fragment coupling.
Scheme 2 Synthesis of the C1–C14 northern fragment 8.
Scheme 3 Synthesis of the C15–C27 southern fragment 7.
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converted into its TES ether (TESOTf), followed by ester cleavage and methylation (MeI, NaH) of the resulting alcohol next installed
(DIBAL) to afford the secondary alcohol 18 (95%, 2 steps). As the C19 methoxy-substituted stereocentre in 23 (88%, 13:1 dr).
dictated by our site-specific macrolactonisation plan, the PMB ether
At this stage, the required cleavage of the C27 benzyl ether in the
presence of a PMB ether, primary TES ether and alkene in 23
proved challenging, but could be accomplished by controlled
treatment with lithium 4,4’-di-tert-butylphenylide (LiDBB) at low
temperature.24 The primary alcohol was then converted into its
Alloc derivative (allyl chloroformate, pyr) to give 24 (84%, 2 steps).
Selective cleavage of the primary TES ether (AcOH) and Dess–
Martin oxidation then afforded the desired C15–C27 southern
fragment 7 (75%, 2 steps). This scalable sequence could be
comfortably carried out to prepare multi-gram quantities of the
aldehyde 7 in a single batch.
at C21 in 18 was at this point transferred over to the C23 position
via reductive opening (DIBAL) of the derived PMP acetal (DDQ),
generating a primary alcohol for Dess–Martin oxidation20 to give
the C21–C27 aldehyde 14 (58%,
3 steps). Moving forward,
elaboration to the C15–C27 fragment 7 was dependent on an HWE
olefination involving the β-keto phosphonate 19 containing the
isolated C17 methyl-bearing stereocentre. This chiral building block
was conveniently accessed through a scalable four-step sequence,
by first employing a pig liver esterase-catalysed hydrolysis of diester
20 to give the acid 21 (96% ee).21 Following selective reduction of
the acid to the primary alcohol (BH3⋅SMe2) and TES ether formation,
the ester was converted (MeP(O)(OMe)2, nBuLi) into the β-keto
phosphonate 19. A Ba(OH)2-mediated HWE coupling22 between 19
The stage was now set to explore our planned fragment coupling
and macrocyclisation strategy (Scheme 4). Firstly,
a similar
and 14 then proceeded smoothly to yield the (E)-enone 22 (85%).
23
sequence of Ba(OH)2-mediated HWE olefination, (R)-Me-CBS
Asymmetric reduction using the (S)-Me-CBS catalyst and BH3⋅SMe2
asymmetric reduction and methylation (Me3O⋅BF4) served to unite
the key fragments 7 and 8, setting in place the required (E)-
trisubstituted alkene and the C13 methoxy-bearing stereocentre in
25 (67%, 3 steps, 15:1 dr). Next, treatment of 25 with DDQ served
to excise both the C1 TBS and C23 PMB ethers with concomitant
oxidation to the dienal.25 Further oxidation to the seco-acid 26
7 + 8
i. Ba(OH)2, aq THF
ii. (R)-Me-CBS, BH3•SMe2,
THF, –10 °C
67%
iii. Me3O•BF4,
proton sponge, CH2Cl2
OTBS
(78%,
2 steps) was then performed using sodium chlorite.
PMB TES
TES
O
TES
O
O
O
OAlloc
Gratifyingly, the critical macrocyclisation of 26 proceeded smoothly
under Yamaguchi conditions26 (TCBC, Et3N, DMAP) to deliver the
desired 24-membered macrolactone core 27 (70%). Finally,
treatment of 27 with Pd(PPh3)4 and dimedone cleanly removed the
13
MeO
OMe
C27 Alloc group to intercept the known macrocyclic intermediate 5
(99%),12a thus demonstrating the viability of this improved second-
generation route.
25
i. DDQ, CH2Cl2,
pH 7 buffer
78% ii. NaClO2, NaH2PO4,
tBuOH, H2O
Moving forward from this key intermediate, we first targeted the
synthesis of aplyronines A (1) and D (2) which differ in the amino
acid residue attached at C29 via a late-stage divergence (Scheme 5).
Following our earlier work,12a,14 the full side chain was first installed
using a boron-mediated aldol reaction (c-Hex2BCl, Et3N) between
TES
O
TES
CO2H
OH
TES
OAlloc
O
O
23
MeO
OMe
the C27 aldehyde 28, obtained by Swern oxidation of 5, and the
methyl ketone 6. The resulting adduct was treated with Burgess
reagent27 to effect the controlled β-elimination of the newly formed
C27 alcohol to give the (E)-enone 29 (65%). This aldol/dehydration
sequence proved remarkably effective, particularly considering the
presence of a potentially labile acetoxy group at the β’ position.
Conjugate reduction of 29 could then be achieved with a catalytic
amount of copper(I) hydride, following an adaptation of a Lipshutz
protocol.28,29 Diastereoselective reduction (Zn(BH4)2) of the
26
TCBC, Et3N, THF;
DMAP, PhMe
70%
O
TES
TES
O
TES
O
O
O
OR
1
27
23
MeO
OMe
resulting ketone then afforded the correctly configured C29 alcohol
30 (56%, 2 steps, 10:1 dr). We rationalise this useful result based on
the participation of an intermediate eight-membered zinc chelate
involving the ketone and acetyl carbonyl groups.30 Strategically, this
advanced intermediate 30 represented the divergence point from
which the majority of the known aplyronine congeners3c and
designed analogues might be accessed.
27: R = Alloc
5: R = H
Pd(PPh3)4,
dimedone,
THF
99%
Scheme 4 Synthesis of the macrolactone core 5.
4 | J. Name., 2012, 00, 1-3
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Initially targeting the synthesis of aplyronine A (1) from 30, the (S)- aplyronine C (3 in Figure 1).32 At this stage, our strategy required
N,N-dimethylalanine moiety at C29 could be introduced under the crucial site-selective esterification of the C7 hydroxyl with
either Keck31 (DCC, DMAP⋅HCl) or Yamaguchi26 (TCBC, DMAP, Et3N)
N,N,O-trimethylserine in the presence of the C9 and C25 secondary
hydroxyls. After some exploratory investigations,33 we found that
5
this transformation could be accomplished under Yamaguchi
conditions with complete site-selectivity at C7, where careful
monitoring by TLC ensured that no bis-esterification product at C9
was formed. The use of racemic N,N,O-trimethylserine gave rise to
(COCl)2, DMSO, Et3N,
CH2Cl2, - 78 °C
99%
O
a
1:1 mixture of (S)- and (R)-diastereomeric esters in good
TES
TES
O
TES
O
O
O
O
approximation to the reported 1.1:1 dr in the natural product. To
our satisfaction, the direct transformation of aplyronine C into
aplyronine A was achieved in 61% yield (82% BRSM), and all the
spectroscopic data agreed with that reported for natural aplyronine
A.3a,34 This pleasing result validated our streamlined protecting
group strategy, which contrasts with the more elaborate differential
protection of the various alcohols employed previously.5,11
H
27
MeO
OMe
28
O
OAc
Me
N
CHO
i. cHex2BCl, Et3N, Et2O, - 78 °C
ii. Burgess reagent, THF
28
6
65%
We now focused on completing the total synthesis of aplyronine D
(2). By substituting the N,N-dimethylalanine ester at C29 for N,N-
dimethylglycine, an analogous sequence was performed from the
pivotal C29 alcohol 30 to afford 2 (43%, 79% BRSM, 3 steps).
Gratifyingly, the spectroscopic and biological data obtained for
synthetic aplyronine D was an excellent match for that reported for
the natural product (copies of NMR spectra were kindly provided by
Prof. Yamada).3c Notably, this constitutes the first total synthesis of
this much rarer, yet apparently more active, aplyronine congener.34
As in the related case of aplyronines A and C, its presumed
biosynthetic precursor with a free hydroxyl at C7 is also likely to be
a natural product. We designate this congener as aplyronine I (see
the ESI, structure 30a), although as far as we are aware this has not
yet been isolated from extracts of Aplysia kurodai.
O
O
TES
O
TES
TES
O
O
O
OAc
Me
N
29
CHO
29
MeO
TES
OMe
i. Cu(OAc)2, PPh3, TMDS, PhMe
ii. ZnBH4, Et2O, 0 °C
56%, 10:1 dr
O
TES
O
TES
O
7
O
O
OH OAc
29
Me
N
CHO
25
9
30
MeO
OMe
For aplyronine A:
For aplyronine D:
Finally, in work towards the synthesis of designed analogues
suitable for ADC applications, we set out to prepare the linker-
modified aplyronine 4 (Scheme 6). Alteration of the actin-binding
tail region of the aplyronines at the C29 position has been shown to
have little effect on the cytotoxicity.35 Furthermore, inspection of
the 1:1 aplyronine A-actin crystal structure6 suggested that
elaboration of the amino acid moiety at C29 would not be expected
to impact on the key ligand-protein binding interactions.
Importantly, our modular synthetic route offers the opportunity to
introduce a range of functional handles at C29 using the alcohol 30
for late-stage diversification. The initially designed aplyronine
i. (S)-N,N-dimethylalanine, DCC, i. N,N-dimethylglycine, TCBC, DMAP
DMAP•HCl, CH2Cl2
ii. HF•pyr, pyr, THF
iii. (rac)-N,N,O-trimethylserine,
TCBC, DMAP, Et3N,
PhH, CH2Cl2
Et3N, THF, PhMe
ii. HF•pyr, pyr, THF
iii. (rac)-N,N,O-trimethylserine,
TCBC, DMAP, Et3N,
PhH, CH2Cl2
49%, 69% BRSM
43%, 79% BRSM
Me2N
R1
O
O
OR2
7
OH
9
O
OH
25
O
OAc
Me
N
CHO
29
analogue
4 contains an Fmoc-protected primary amine for
MeO
OMe
bioconjugation studies, for example through the derived maleimide
or functionalised amide.36
Me2N
=
1: aplyronine A, R1 = Me, R2
2: aplyronine D, R1 = H, R2
OMe
=
1:1 S:R
O
To construct analogue 4, attachment of the modified glycine
derivative 31 (see the ESI for its preparation) again proceeded
smoothly under Yamaguchi esterification conditions (TCBC, DMAP,
Et3N) and the three TES ethers were cleaved on treatment with
Scheme 5 Completion of the total synthesis of aplyronines A and D.
esterification conditions. This afforded product that was
HF·pyr (91%, 2 steps). The trimethylserine residue was then
attached site-selectively at C7 in the resulting triol using a similar
procedure to that developed for aplyronines A and D. Furthermore,
a
configurationally pure with respect to the amino acid stereocentre.
Cleavage of all three TES ethers using HF·pyr then afforded
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it was demonstrated that this transformation could be achieved Pillow (Genentech), Dr Ray Finlay (AstraZeneca) and Dr Andrew
stereospecifically using enantiopure (S)-N,N,O-trimethylserine (see Mortlock (AstraZeneca) for helpful discussions, Prof. K. Yamada
the ESI for its preparation) to afford
4 (58%) as a single (Nagoya) for copies of NMR spectra for aplyronine D, and the EPSRC
diastereomer.37,38 Given the key structural requirement for the UK National Mass Spectrometry Facility at Swansea University for
presence of the N,N,O-trimethylserine moiety for the potent mass spectra.
biological activity of aplyronine A, it is likely that each epimer will
have different binding affinities with tubulin and hence variable
Notes and references
cytotoxicity, thus the preparation of configurationally pure
stereoisomers will allow these SAR effects to be investigated in
future work.39,40
1
a) D. J. Newman and G. M. Cragg, J. Nat. Prod., 2016, 79,
629; b) M. S. Butler, A. A. B. Robertson and M. A. Cooper,
Nat. Prod. Rep., 2014, 31, 1612; c) K. von Schwarzenberg and
A. M. Vollmar, Cancer Lett., 2013, 332, 295; d) S. M. Dalby
30
and I. Paterson, Curr. Opin. Drug Discov. Devel., 2010, 13
777; e) I. Paterson and E. A. Anderson, Science, 2005, 310
,
,
Fmoc
i. 31, TCBC, Et3N, DMAP, THF, PhMe
52% ii. HF•pyr, pyr, THF
451.
ꢀ
NH
Me
N
O
O
2
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iii. (S)-N,N,O-trimethylserine, TCBC
DMAP, Et3N, PhMe, THF
OH
31
Me
Me2N
N
Fmoc
O
OMe
N
H
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,
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O
OH
O
7
O
OH
O
OAc
Me
N
CHO
3
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29
4
MeO
OMe
Scheme 6 Preparation of the C29 linker-modified aplyronine 4.
4
5
Conclusions
In conclusion, we have achieved the total synthesis of the potent
antimitotic macrolides aplyronines A (1.3%) and D (1.1%) in 29 steps
LLS based on an improved second-generation route that has
provided straightforward access to multi-gram quantities of key
intermediates. Additionally, we have prepared the linker-modified
version 4 for bioconjugation studies in the context of exploring the
use of aplyronines as novel cytotoxic payloads for ADCs in targeted
cancer chemotherapy. We envisage that this flexible and modular
synthetic route can provide a sustainable supply of the aplyronines
and their designed analogues for further biological studies, and in
future work will adapt it to generate novel aplyronine-based
payloads for the discovery of improved ADCs.
6
7
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Usui, E. Sumiya, M. Uesugi and H. Kigoshi, J. Am. Chem. Soc.,
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ꢀ
ꢀ
Conflicts of Interest
There are no conflicts of interest to declare.
12 a) I. Paterson, S. J. Fink, L. Y. W. Lee, S. J. Atkinson and S. B.
Blakey, Org. Lett., 2013, 15, 3118; b) I. Paterson, M. D.
Acknowledgements
Woodrow and C. J. Cowden, Tetrahedron Lett., 1998, 39
,
We thank the Public Scholarship, Development, Disability and
Maintenance Fund of the Republic of Slovenia (Studentship to
N.A.), the Todd-Raphael Fund (Scholarship to S.W.), Downing
College, Cambridge (Mays-Wild Fellowship to M.P.H), and Prof.
David Spring for support, Dr. Michael Woodrow for his contribution
to earlier synthetic studies Dr. David Newman (NCI), Dr. Thomas
6041; c) I. Paterson, C. J. Cowden and M. D. Woodrow,
Tetrahedron Lett., 1998, 39, 6037; d) I. Paterson, S. B. Blakey
and C. J. Cowden, Tetrahedron Lett., 2002, 43, 6005.
13 For other synthetic efforts towards the aplyronines, see: a) J.
A. Marshall and B. A. Johns, J. Org. Chem., 2000, 65, 1501; b)
W. P. Hong, M. N. Noshi, A. El-Awa and P. L. Fuchs, Org. Lett.,
2011, 13, 6342; c) M. N. Noshi, A. El-Awa, E. Torres and P. L.
6 | J. Name., 2012, 00, 1-3
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DOI: 10.1039/C7OB03204H
Journal Name
ARTICLE
O
Fuchs, J. Am. Chem. Soc., 2007, 129, 11242; d) M. A. Calter
and X. Guo, Tetrahedron, 2002, 58, 7093; e) M. A. Calter and
J. Zhou, Tetrahedron Lett., 2004, 45, 4847.
OH OH
7
O
OH
OTBS
14 This versatile aldol-based strategy for introducing the
characteristic actin-binding tail of the aplyronines was
employed successfully in the context of our total synthesis of
reidispongiolide A and rhizopodin, as well as simplified
analogues, see: a) I. Paterson, K. Ashton, R. Britton, G.
Cecere, G. Chouraqui, G. J. Florence, H. Knust and J. Stafford,
MeO
OMe
34 Preliminary screening results have confirmed the expected
sub-nanomolar antiproliferative activity of our synthetic
samples of aplyronines A and D in an extensive panel of
human cancer cell lines. Detailed biological evaluation will be
reported in due course.
Chem. Asian J., 2008,
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,
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36 Using an extended aplyronine side chain as a model system,
we have demonstrated the elaboration of the Fmoc-amino
substituted linker into a maleimide-based Michael acceptor
for bioconjugation with cysteine residues, as shown below.
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,
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,
Me
N
R
N
H
O
O
O
OH
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OAc
Me
N
BnO
29
CHO
R = Fmoc
R = H
22 I. Paterson, K.-S. Yeung and J. B. Smaill, Synlett, 1993, 774.
ꢀ
piperidine, MeCN
O
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N
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NaHCO3,
aq THF
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28 a) To enhance the chemoselectivity for reduction of the
enone over the dienoate, we employed triphenylphosphine 37 The biological activity of
as the ligand to form a less active reagent, see: B. A. Baker, Z. context of ADC development.
V. Bosković and B. H. Lipshutz, Org. Lett., 2008, 10, 289; b) 38 Yamada (ref 5) and Kigoshi (ref 11) both reported some
3, 2399.
Me
N
O
N
O
ꢀ
O
O
OH
O
OAc
Me
N
O
ꢀ
BnO
CHO
29
ꢀ
4
will be presented elsewhere in the
W. S. Mahoney, D. M. Brestensky and J. M. Stryker, J. Am.
Chem. Soc., 1988, 110, 291.
racemisation of the activated carboxylic acid under their
reaction conditions. In our case, the retention of
29 A stock solution of copper(I) hydride was prepared from
copper(II) acetate and triphenylphosphine using excess
tetramethyldisiloxane (TMDS) as the reducing agent. D. Lee
and J. Yun, Tetrahedron Lett., 2005, 46, 2037.
configuration to provide
4
temperature employed and shorter time required for a less
hindered substrate.
a single diastereomer in the
esterified product could be attributed to the lower
30 T. Oishi and T. Nakata, Acc. Chem. Res., 1984, 17, 338.
39 It is unclear whether the differing epimeric ratios of amino
acids present in the aplyronines are a natural feature or an
artefact of the isolation procedure. The actual producing
organism for the aplyronines is believed to be cyanobacteria,
which are ingested from algae consumed by the sea hare in
its diet.
31 E. P. Boden and G. E. Keck, J. Org. Chem., 1985, 50, 2394.
32 The overall yield for this second-generation synthesis of
aplyronine C ( ) is 2.7% versus 3.6% for our earlier route (ref
ꢀ
3
12a). However, this new route is streamlined with regard to
protecting group manipulations, more scalable through
avoiding the equilibration of the undesired 26-membered to 40 For related synthetic work from our group, see: I. Paterson
the desired 24-membered macrocycle and can potentially
provide multi-gram quantities of the aplyronines.
and N. Y. S. Lam, J. Antibiot., 2018, doi: 10.1038/ja.2017.111.
…
33 Model studies into the esterification site-selectivity were
carried out on the truncated macrocyclic triol shown below.
These led to the development of suitable reaction conditions
for site-specific attachment of N,N,O-trimethylserine at the
C7 hydroxyl group.
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DOI: 10.1039/C7OB03204H
Toward Aplyronine Payloads for Antibody-Drug Conjugates:
Total Synthesis of Aplyronines A and D
Nika Anžiček, Simon Williams, Michael P. Housden and Ian Paterson*
University Chemical Laboratory Lensfield Road. Cambridge CB2 1EW, UK
We report an expedient total synthesis of aplyronines A and D, together with a
linker-modified analogue for bioconjugation studies.