Enantioselective Synthesis of a Cyclopropane Derivative of Spliceostatin A and Evaluation of Bioactivity

Article abstract of DOI:10.1021/acs.orglett.8b03228

Spliceostatin A is a potent inhibitor of spliceosomes and exhibits excellent anticancer activity against multiple human cancer cell lines. We describe here the design and synthesis of a stable cyclopropane derivative of spliceostatin A. The synthesis invo

Full text of DOI:10.1021/acs.orglett.8b03228

Letter
pubs.acs.org/OrgLett
Cite This: Org. Lett. XXXX, XXX, XXX−XXX
Enantioselective Synthesis of a Cyclopropane Derivative of
Spliceostatin A and Evaluation of Bioactivity
Arun K. Ghosh,*,† Guddeti Chandrashekar Reddy,† Satish Kovela,† Nicola Relitti,† Veronica K. Urabe,‡
Beth E. Prichard,‡ and Melissa S. Jurica‡
†Department of Chemistry and Department of Medicinal Chemistry, Purdue University, 560 Oval Drive, West Lafayette, Indiana
47907, United States
‡Department of Molecular Cell and Developmental Biology and Center for Molecular Biology of RNA, University of California,
Santa Cruz, California 95064, United States
S
* Supporting Information
ABSTRACT: Spliceostatin A is a potent inhibitor of spliceosomes and
exhibits excellent anticancer activity against multiple human cancer cell lines.
We describe here the design and synthesis of a stable cyclopropane derivative
of spliceostatin A. The synthesis involved a cross-metathesis or a Suzuki
cross-coupling reaction as the key step. The functionalized epoxy alcohol
ring was constructed from commercially available optically active tri-O-
acetyl-D-glucal. The biological properties of the cyclopropyl derivative
revealed that it is active in human cells and inhibits splicing in vitro comparable to spliceostatin A.
he majority of eukaryotic genes are expressed as
Tprecursor mRNAs (pre-mRNAs) which are then
converted to mRNAs by splicing.1,2 This event is carried out
by a multimegadalton ribonucleoprotein complex, spliceosome.
Upon recognition of a splicing signal, the spliceosome catalyzes
the removal of noncoding sequences (introns) and joins
protein coding sequences (exons) to form mRNAs.3−5 These
are exported to the cytoplasm for translation into proteins.
While these transcription and translation processes are
complex, recent studies have demonstrated that splicing is
pathologically altered to promote the initiation and main-
tenance of cancer.6,7 Splicing involves numerous protein−
protein and protein−RNA interactions which offer oppor-
tunities to manipulate or inhibit the splicing cascade for
therapeutic purposes, particularly in the area of anticancer drug
development.8,9 Presently, a number of natural products and
their derivatives are known to potently inhibit spliceosome
function by binding to the SF3B subunit of U2 SnRNP.10−12
These include FR901464 (1, Figure 1), spliceostatin A (2),
pladienolide B (3), and a semisynthetic derivative E7107,
4.13−15 The precise molecular interactions of these molecules
with SF3B are being investigated using cryo-electron
microscopy.16−18 While these natural products display potent
splicing activity, their clinical use is limited due to chemical
instability and inadequate physiochemical properties. To date,
a semisynthetic derivative of pladienolide, E7107 (4) was
developed with improved pharmacological properties for
clinical development.19 A number of spliceostatin derivatives
have been specifically synthesized as payloads for antibody−
drug conjugates.20,21 Both FR901464 (1) and spliceostatin A
(2) show very potent antitumor properties. FR901464
displayed IC50 values ranging from 0.6 to 3.4 nM against
multiple human cancer cell lines. It also showed effectiveness
Figure 1. Structures of spliceosome inhibitors 1−5.
against solid tumors implanted in mice at a dose range of 0.05
to 1 mg/kg.14,22 Spliceostatin A showed similar activity. As a
result, these compounds attracted much attention for synthesis
and medicinal chemistry development. We recently reported
the synthesis and structure−activity studies of both these
compounds.23,24 Nicolaou and co-workers also reported
Received: October 10, 2018
© XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.8b03228
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Letter
structural modifications of spliceostatin derivatives.25 In our
continued interest in developing molecular probes for splicing
studies, we devised a cyclopropyl derivative of FR901464 and
spliceostatin A, where a cyclopropane is incorporated at the
anomeric site of FR901464 and spliceostatin A to improve
stability and potency. We have devised an enantioselective
synthesis of the epoxide subunit using readily available tri-O-
acetyl-D-glucal as the key starting material and investigated
coupling using cross-metathesis and Suzuki reactions. The
synthesis will provide ready access to stable spliceostatin
derivatives for biological studies.
Scheme 2. Synthesis of Dihydropyranone 18
Our synthetic strategy for the construction of the cyclo-
propyl derivative of spliceostatin A is shown in Scheme 1. We
Scheme 1. Retrosynthesis of Spliceostatin Derivative
presence of a catalytic amount of PPTS at 23 °C for 2 h
afforded benzylidene acetal 14. It was protected as the TBS-
ether using TBSCl and imidazole in the presence of a catalytic
amount of DMAP in DMF at 23 °C for 12 h. The resulting
TBS-ether was treated with NIS in a mixture (95:5) of CH3CN
and water at 23 °C for 15 min, and the mixture was
concentrated. The residue was dissolved in DMF, saturated
NaHCO3 and Na2S2O4 were added, and the mixture was
stirred at 23 °C for 5 h to provide lactol 15 in 81% yield over
two steps.29,30 Oxidation of lactol 15 with Dess-Martin
periodinane (DMP) in the presence of NaHCO3 in CH2Cl2
at 0 to 23 °C for 2 h furnished lactone derivative 16 in 81%
yield. To install the cyclopropane ring, lactone 16 was treated
with Petasis reagent31 in toluene at 23 °C and the resulting
mixture was heated at 60 °C for 48 h to provide the
corresponding enol-ether in 90% yield. Simmons−Smith
cyclopropanation32 of the resulting enol ether with methylene
diiodide and diethyl zinc in CH2Cl2 at 0 to 23 °C for 2 h
afforded 4-oxaspiro[2,5]octane derivative 17 in 42% yield.
Cyclopropane derivative 17 was converted to olefin 11 as
follows. Treatment of TBS-ether 17 with TBAF in THF at 0 to
23 °C for 3 h provided the corresponding alcohol. Oxidation of
the secondary alcohol to the corresponding ketone was
achieved by treatment with DMP at 0 to 23 °C for 2 h.
Wittig-olefination of the resulting ketone with methylene-
triphenylphosphorane in THF at 0 to 23 °C for 3 h provided
initially planned cross metathesis of diene 6 and epoxy olefin 8
using Grubbs’ second generation catalyst. This cross meta-
thesis reaction typically yields a mixture of olefin dimer, diene
dimer, and the desired cross-coupling product. We also
planned to investigate a Suzuki cross-coupling reaction
between the known26,27 pinacol borononate 7 and vinyl iodide
derivative 9 to provide the desired cross-coupling product 5.
The synthesis of pinacol boronate 7 would be achieved in
optically active form using our recently reported procedure.27
The synthesis of cyclopropyl epoxide segment 8 or 9 would
be carried out from alcohol 10 which would be derived from 4-
methoxybenzylidene acetal 11 by reduction with DIBAL-H.
Acetal 11 can be synthesized from δ-lactone 12 by synthetic
manipulation from standard commercially available optically
active tri-O-acetyl-D-glucal 13.
Enantioselective synthesis of 7-methylene-4-oxaspiro[2.5]-
octane derivative 10 is shown in Scheme 2. The preparation of
benzylidene acetal 14 was carried out in multigram scale using
commercially available tri-O-acetyl-D-glucal 13 with minor
modifications.28 Treatment of 13 with K2CO3 in MeOH at 23
°C for 5 h provided the corresponding triol. Reaction of the
resulting triol with 1-(dimethoxy)-4-methoxybenzene in the
B
DOI: 10.1021/acs.orglett.8b03228
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Letter
11 in 64% yield over three steps. DIBAL-H reduction of
benzylidene acetal 11 at −78 to 0 °C for 4 h furnished alcohol
10 in 93% yield.
Scheme 4. Synthesis of Spliceostatin Derivative 5
Conversion of alcohol 10 to epoxy alcohol derivatives 8 and
9 is shown in Scheme 3. Oxidation of alcohol 10 with DMP in
Scheme 3. Synthesis of Epoxy Alcohol Derivatives
the presence of NaHCO3 provided the corresponding
aldehyde. Wittig-olefination of the resulting aldehyde with
methylenetriphenylphosphorane in THF at 0 °C for 1 h
afforded diene 18 in 53% yield over two steps. Removal of the
PMB-group was carried out by exposure of 18 to DDQ in a
mixture (10:1) of CH2Cl2 and phosphate buffer (pH 7.2) at 0
°C for 2 h to furnish homoallylic alcohol 19. Selective
epoxidation of 19 with a catalytic amount (18 mol %) of
VO(acac)2 in the presence of tert-butyl hydroperoxide in
CH2Cl2 at 0 to 23 °C for 1.5 h provided epoxy alcohol
derivative 8 as a single diastereomer (by 1H NMR analysis) in
57% yield (73% brsm). For the synthesis of vinyl iodide
derivative 9, alcohol 10 was oxidized with DMP in the
presence of NaHCO3 to provide the aldehyde in 48% yield.
Takai olefination33 was carried out by reaction of the resulting
aldehyde with a mixture of CrCl2 and CHI3 in THF at 23 °C
for 3 h to furnish vinyl iodide 20 in 85% yield. Exposure of
PMB-derivative 20 to DDQ in a mixture of CH2Cl2 and
phosphate buffer resulted in deprotection of the PMB-group.
Directed epoxidation of the resulting allyic alcohol with tert-
butyl hydroperoxide in the presence of a catalytic amount (18
mol %) of VO(acac)2 provided epoxy alcohol 9 in 64% yield
over two steps.
For the synthesis of the cyclopropyl derivative of
spliceostatin A, we first carried out a cross metathesis34
reaction of diene 6 and epoxy olefin 8 as shown in Scheme 4. A
mixture of epoxy olefin 8 and Grubbs’ II catalyst (15 mol %) in
CH2Cl2 was prepared under an argon atmostphere. To a
stirred solution of diene 6 in CH2Cl2, a one-third portion of
epoxy olefin 8 and Grubbs’ catalyst was added and the
resulting mixture was heated at reflux for 1.5 h. The other two
portions of catalyst and olefin mixture were added successively
in 1.5 h intervals. The resulting mixture was stirred at reflux for
an additional 1.5 h (total 8 h). The reaction was cooled to 23
°C, and the mixture was concentrated. The residue was
purified by silica gel chromatography to furnish cyclopropane
derivative 5 in 27% yield. The cross metathesis reaction also
provided an inseparable mixture of epoxide dimer and diene
dimer as the byproducts (21 and 22, about 25−30%). We then
investigated a Suzuki coupling35 of known boronate 7 and
vinyl iodide derivative 9 using Pd(dppf)2Cl2·DCM (10 mol %)
catalyst in the presence of aqueous Cs2CO3 in THF at 23 °C
for 30 min to provide the coupling product 5 in 29% yield. The
yield for coupling product 5 via cross-metathesis or Suzuki
coupling was comparable.
We evaluated the biological activity of this cyclopropane
spliceostatin A (5) in an in vitro splicing system as previously
described.36 The compound inhibits splicing, but at a slightly
reduced (∼2-fold lower) potency relative to spliceostatin A
(Figure 2). The compound is also active in HeLa cells. It
induces the coalescence of nuclear speckles as observed by
immuno-staining of the splicing factor SFRS2 at similar levels
as spliceostatin A (Figure 3).
In summary, we reported the design, synthesis, and
biological evaluation of a cyclopropane derivative of
spliceostatin A and FR901464. The synthesis of the cyclo-
propane derivatives 8 and 9 for coupling reactions was carried
out enantioselectively from commercially available, optically
C
DOI: 10.1021/acs.orglett.8b03228
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ASSOCIATED CONTENT
■
S
* Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.or-
glett.8b03228.
Experimental procedures in addition to 1H and 13C
NMR spectra are available for all new compounds
(PDF)
AUTHOR INFORMATION
■
Corresponding Author
*E-mail: akghosh@purdue.edu.
ORCID
Figure 2. Impact of compound 5 on in vitro splicing. Average splicing
efficiency relative to inhibitor concentration normalized to no-drug
control. Compound 5; SSA, spliceostatin A.
Arun K. Ghosh: 0000-0003-2472-1841
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
Financial support of this work was provided by the National
Institutes of Health (GM122279) and Purdue University. The
authors thank Ms. Hannah Simpson and Mr. Josh Born (both,
Purdue University) for valuable discussions
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