Small-molecule anticancer agents kill cancer cells by harnessing reactive oxygen species in an iron-dependent manner

Article abstract of DOI:10.1039/c7ob03086j

In the course of generating a library of open-chain epothilones, we discovered a new class of small molecule anticancer agents that has no effect on tubulin but instead kills selected cancer cell lines by harnessing reactive oxygen species in an iron-dependent manner. Results of the preliminary studies are consistent with the recently described cell death mechanism ferroptosis. Studies are in progress to confirm ferroptosis as the cell death mechanism and to identify the specific molecular targets of these small molecule anticancer agents.

Full text of DOI:10.1039/c7ob03086j

View Article Online
View Journal
Organic &
Biomolecular
Chemistry
Accepted Manuscript
This article can be cited before page numbers have been issued, to do this please use: S. Fedorka, L. M.
V. Tillekeratne, W. Taylor, A. Al-Hamashi, K. So, Ibtissam. Gad, R. Shah, Veronika. Kholodovych and Hanan.
.Alqahtani, Org. Biomol. Chem., 2018, DOI: 10.1039/C7OB03086J.
This is an Accepted Manuscript, which has been through the
Volume 14 Number
1 7 January 2016 Pages 1–372
Royal Society of Chemistry peer review process and has been
accepted for publication.
Organic &
Biomolecular
Chemistry
www.rsc.org/obc
Accepted Manuscripts are published online shortly after
acceptance, before technical editing, formatting and proof reading.
Using this free service, authors can make their results available
to the community, in citable form, before we publish the edited
article. We will replace this Accepted Manuscript with the edited
and formatted Advance Article as soon as it is available.
You can find more information about Accepted Manuscripts in the
author guidelines.
Please note that technical editing may introduce minor changes
to the text and/or graphics, which may alter content. The journal’s
standard Terms & Conditions and the ethical guidelines, outlined
in our author and reviewer resource centre, still apply. In no
event shall the Royal Society of Chemistry be held responsible
for any errors or omissions in this Accepted Manuscript or any
consequences arising from the use of any information it contains.
ISSN 1477-0520
COMMUNICATION
Takeharu Haino et al.
Solvent-induced emission of organogels based on tris(phenylisoxazolyl)
benzene
rsc.li/obc

Page 1 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
Small-molecule anticancer agents kill cancer cells by
harnessing reactive oxygen species in an iron-dependent
manner
Sara R. Fedorkaa, Kevin Soa, Ayad AlꢀHamashia, Ibtissam Gadb, Ronit Shahb,
Veronika Kholodovychb, Hanan D. Alqahtanib, William R. Taylorb* and L. M.
Viranga Tillekeratnea*
a
Department of Medicinal and Biological Chemistry, College of Pharmacy and Pharmaceutical Sciences,
The University of Toledo, 2801, W. Bancroft Street, Toledo, Ohio 43606, United States
b
Department of Biological Sciences, College of Natural Sciences and Mathematics, The University of
Toledo, 2801, W. Bancroft Street, Toledo, Ohio 43606, United States
*These authors contributing equally to this work and are coꢀcorresponding authors.
Abstract:
In the course of generating a library of openꢀchain epothilones, we discovered a new
class of small molecule anticancer agents that has no effect on tubulin but instead kills selected
cancer cell lines by harnessing reactive oxygen species in an ironꢀdependent manner. Results of
the preliminary studies are consistent with the recently described cell death mechanism
ferroptosis. Studies are in progress to confirm ferroptosis as the cell death mechanism and to
identify the specific molecular targets of these small molecule anticancer agents.

Organic & Biomolecular Chemistry
Page 2 of 46
View Article Online
DOI: 10.1039/C7OB03086J
Introduction
Epothilones 1,2 (Fig. 1), first isolated from the myxobacterium Sorangium cellulosum in
the 1980s,1 are potent cytotoxic agents2 that induce apoptosis through microtubule stabilization
like the blockbuster drug paclitaxel.3,4 Since their isolation, they received the attention of the
scientific community due to their potent cytotoxicity against a variety of cancer cell lines
including breast, lung, and prostate cancers.2 Both paclitaxel and epothilone molecules bind to
the α/β tubulin heterodimer subunits increasing tubulin aggregation to microtubules, while
decreasing α/β heterodimer disassociation from microtubules.3
We previously reported the synthesis and biological activity of a new class of structurally
simplified openꢀchain epothilones 3 and 4 (Fig. 1)5 in which the C9ꢀC15 sector of the epothilone
molecules (1 and 2 , Fig. 1)2–4 has been completely deleted, while retaining the two biologically
relevant sectors C1ꢀC8 and C16ꢀC20 connected by a rigid cyclopentene molecular scaffold for
conformational stability.
Fig. 1. Natural epothilones 1,2 and synthetic open-chain epothilones 3,4.
The diastereomer 3, with stereochemical assignments as in natural epothilones, had weak
but selective activity against SNBꢀ75 (GI50 = 21.9 ꢀM) CNS cancer and OVCARꢀ4 (GI50 = 45.1
ꢀM) ovarian cancer cell lines. A computational study performed by RusinskaꢀRoszak et al6
suggested a high degree of conformational similarity between these openꢀchain molecules and
natural epothilones and confirmed that conformational requirements for tubulin binding are

Page 3 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
being met with the openꢀchain analogues. In carrying out a structureꢀactivity relationship study
of these molecules, we focused on the substituent at the 2ꢀpositon of the aromatic ring as the rest
of the structure embodied most of the important structural features important for activity.
Thiazole was chosen as the aromatic moiety to be consistent with the thiazole of natural
epothilones. Thus, we designed a library of openꢀchain epothilone analogues 5a-f (Scheme 1)
with a variety of 2ꢀsubstituted thiazole rings to investigate their effects on biological activity.
One of the compounds showed strong cytotoxic activity on several cell lines in the NCI 60
human tumor cell line screen tested at 10ꢀ5 M. Follow up studies in our laboratory traced the
observed activity to a part of the molecule which kills cells by a nonapoptotic mechanism of
action. Preliminary studies show that these compounds do not target tubulin, but kills cells by a
mechanism that shows the hallmarks of the recently described cell death mechanism
“ferroptosis”,7 characterized by dependence on iron and ROS.
Results and Discussion
Chemistry
Compounds 5a-f were derived by the esterification of the carboxylic acid 7 with 2ꢀsubstituted
thiazole cyclopentenols 6a-f as shown in the retrosynthetic analysis in Scheme 1.

Organic & Biomolecular Chemistry
Page 4 of 46
View Article Online
DOI: 10.1039/C7OB03086J
Scheme 1. Retrosynthetic analysis of openꢀchain epothilones
We synthesized the carboxylic acid 7 as reported previously (scheme 2),5 with the
exception that the two diastereomers were separated efficiently by flash chromatography at the
aldehyde stage. This was found to be easier and more efficient than separating the two carboxylic
acids as reported earlier. The two aldehydes were then separately converted to the corresponding
carboxylic acids 7 and 7a by Pinnick oxidation.8
Scheme 2. Synthesis of carboxylic acids 7 and 7a
The synthesis of cyclopentenols bearing aromatic moieties 6a-f began with
dibromothiazole 12, which by nucleophilic substitution, metalꢀhalogen exchange followed by

Page 5 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
alkylation or Sonogashira coupling, produced 2ꢀsubstituted 4ꢀbromothiazole derivatives 13-17
in good yields (scheme 3).9–11 They were treated with nꢀbutyllithium followed by trimethyltin
chloride at ꢀ78oC to produce the corresponding tin derivatives (Scheme 4a). tꢀButyllithium
proved more effective for the stannylation of the TMSꢀacetylene derivative 17. The tin
derivatives were partially purified by passing through a plug of silica gel deactivated with 5%
triethylamine in hexanes and were immediately subjected to Stille coupling with
iodocyclopentenone 19 in the presence of palladium catalyst to obtain the arylꢀsubstituted
cyclopentenones 20a-e in good yields (schemes 4).11 The TMS group of 20e was removed with
K2CO3 in methanol to obtain 21 prior to reduction (Scheme 4b). Cyclopentenones 20a-d and 21
were selectively reduced using (R)-CBS catalyst to obtain cyclopentenols 6a-e.5 The
stereochemistry of 6a-f was confirmed using the Mosher ester model.12,13 Enantioselective
reduction of the piperonyl ketone 20c to alcohol 6c was not successful and the product was
obtained as a racemic mixture, which was used in the next reaction without separation.
Compound 21 was subjected to click chemistry with azidotrimethylsilane and copper iodide to
obtain the triazole ketone 22 (scheme 5).14

Organic & Biomolecular Chemistry
Page 6 of 46
View Article Online
DOI: 10.1039/C7OB03086J
S
CH3
N
Br
13
i) n-BuLi, -78 oC, 4 h
ii) Dimethyl Sulfate,
-78 oC, 16 h,
52%
, PPh
TMS
3
-
Na+SCH3
Pd(Ph3)4, CuI, Toluene,
Et3N, 140 oC, 48 h
81%
EtOH , 25oC 2 h
94%
S
Br
S
S
S
TMS
N
N
N
Br
Br
12
Br
17
14
i) Pd(Ph3)4, CuI,
DIPA, THF
ii) Phenyl acetylene,
THF, 86%
Piperidine, 50oC
22 h, 86%
S
S
N
N
Br
N
16
Br
15
Scheme 3. Synthesis of 2ꢀsubstitutedꢀbromothiazoles

Page 7 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
Scheme 4. Synthesis of 2ꢀsubstituted cyclopentenols 6a-e
The triazole product 22 was insoluble in most organic solvents except for large quantities
of methanol that precluded CBS reduction. It was methylated15 with MeI, and subjected to CBS
reduction to obtain 6f.

Organic & Biomolecular Chemistry
Page 8 of 46
View Article Online
DOI: 10.1039/C7OB03086J
NH
N
S
S
TMSꢀN3, CuI, DMF
MeOH, w 140 oC, 2 h
N
i) NaOMe, MeOH,
N
N
70%
w, 65 oC 7 min
ii) MeI, w, 90 oC,
30 min, 36%
O
O
22
21
N
N
S
S
N
N
N
N
Rꢀ2ꢀmethylꢀCBS
boraneꢀMe2S
N
N
THF, 0 oC, 40 min
54%, 70% ee
OH
O
20f
6f
Scheme 5. Synthesis of cyclopentenol 6f
We previously reported that desilylation of the TBS groups after esterification of 7
resulted in cleavage of the ester moiety.5 Therefore, as before the silyl groups were first removed
with TFA and the desilylated carboxylic acid was esterified with cyclopentenols 6a-f using DCC
and DMAP to yield openꢀchain epothilones 5a-f (Scheme 6). The purification of the final
products was carried out on preparative TLC plates, which were first deactivated with 5% Et3Nꢀ
hexanes to yield the final products 5 a-f. Other purification methods including column
chromatography on silica gel and reversed phase HPLC resulted in decomposition of the product
and in most cases, the corresponding alcohol was isolated as a breakdown product.

Page 9 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
Scheme 6. Synthesis of target openꢀchain epothilone molecules
Biological studies
The compounds were tested in the NCI 60 cell line oneꢀdose screen at 10ꢀ5 M. The compounds
with methyl (5a), thiomethoxy (5b), piperidinyl (5c) and substituted triazole (5f)ꢀ thiazole rings
showed no activity on any of the cell lines. Interestingly, the two alkyne analogues with
acetylene and phenyl acetylene substituted thiazole rings showed activity against some cell lines.
The phenylacetylene derivative (5d) inhibited the growth of nonꢀsmall cell lung cancer HOPꢀ62
(35%), nonꢀsmall cell lung cancer NCIꢀH226 (45%), melanoma UACCꢀ62 (40%) and renal
cancer A498 (78%) cell lines. The acetylene derivative (5e) showed 75% growth inhibition
against nonꢀsmall cell lung cancer cell line HOPꢀ62 and 25% growth inhibitory activity against
CNS cancer U251 cell line. What was more striking was its lethal effect on nonꢀsmall cell lung
cancer NCIꢀH522 (ꢀ62%), melanoma LOX IMVI (ꢀ23%), ovarian cancer IGROV1 (ꢀ32%) and
renal cancer UOꢀ31 (ꢀ36%). Intrigued by the remarkable lethality of the analogue 5e to NCIꢀ
H522 lung cancer cell line, we repeated this analysis. NCIꢀH522 cells were highly sensitive to

Organic & Biomolecular Chemistry
Page 10 of 46
View Article Online
DOI: 10.1039/C7OB03086J
analogue 5e (LC50 of ~0.6 ꢀM) whereas HOPꢀ62 cells were less sensitive (LC50 of ~30 ꢀM)
(Supplementary Fig. S1). These results show that analogue 5e with a linear terminal alkyne
substituent group at the 2ꢀposition of the thiazole ring has a profound and selective lethal effect
on NCIꢀH522 cells.
During their synthesis, esters 5a-f were found to be rather unstable and underwent ready
hydrolysis in the presence of nucleophilic solvents like water and methanol. We therefore
speculated that the observed cytotoxicity of compound 5e may in fact, be due to compound 6e
formed by hydrolysis of the ester 5e in the assay media, possibly aided by cellular esterases. As
speculated, compound 6e showed similar potency as 5e in the cellular assay and also had a
similar selectivity profile (see below). Therefore, our subsequent studies focused on 6e.
Comparison of several cancer cell lines exposed to 6e for three days indicated an LC50 of 1.5 ꢀM
for NCIꢀH522, and an LC50 of 3.6 ꢀM for the fibrosarcoma cell line HT1080 (Fig. 2). Colon
cancer cell line HCT116, lung cancer HOPꢀ62 and breast cancer cell line MCF7 were relatively
unaffected by compound 6e (Fig. 2). The large difference in sensitivities of these various cell
lines suggests that 6e is not a general toxin, but relies on metabolic and/or genetic factors in
certain tumor cells to allow sensitivity.
We then synthesized the (R)ꢀenantiomer 23 and the racemic alcohol 24 (Fig. 3) using (S)ꢀ
CBS catalyst and direct NaBH4 reduction, respectively. We also synthesized the corresponding
oxime 25 by reaction of the ketone 21 with hydroxylamine hydrochloride and NaOAc in
methanol (Fig. 3). The three compounds killed NCIꢀH522 cells with equal efficacy compared to
6e and had LC50 values of 2.6 ꢀM, 2.5 ꢀM and 2.3 ꢀM, respectively. The ketone 21 was less
lethal with an LC50 value of 8.0 ꢀM (Supplementary Fig. S1). These results show that cytotoxic

Page 11 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
effect is independent of the stereochemistry of the alcohol function and that a Hꢀbond donor is
probably preferred over a Hꢀbond acceptor at this position.
Fig. 2. Survey of cytotoxicity. Cells were exposed to the indicated compounds at 10 ꢀM and
analyzed by methylene blue staining 3 days later. LC50 values of various cell lines determined 3
days post treatment. Cells were treated one day after plating. Cell viability was determined using
methylene blue staining.

Organic & Biomolecular Chemistry
Page 12 of 46
View Article Online
DOI: 10.1039/C7OB03086J
S
S
S
N
N
N
OH
OH
N
HO
24
25
23
Figure 3. Other analogues synthesized
Rapid, non-apoptotic cell death in response to compounds 5e and 6e
Initial timeꢀlapse microscopy indicated that 50% of cells were killed by 5e in ~10 h and
100% by 18 hours (Fig. 4A). The rapid mode of killing of NCIꢀH522 cells by 5e suggests that
the mechanism of cell death is unlikely to be cell cycleꢀdependent. Similar rapid death was
observed with the smaller 6e compound. Immunofluorescence analysis showed that compound
6e had no overt effects on tubulin in interphase or mitosis, suggesting that the compound does
not mimic epothilone in mechanism of action (Fig. 4B). Furthermore, NCIꢀH522 cells were able
to progress through mitosis in the presence of 6e (Supplementary Fig. S2). The rapid mode of
cell death suggested that 6e might directly disrupt mitochondrial function; however, membrane
potentials were intact until late in the death process (Supplementary Fig. S3). Additional timeꢀ
lapse microscopy indicated that cell death was not associated with membrane blebbing (Fig. 4C).
Cells lost contact with the substratum shortly before losing membrane integrity and pyknotic
nuclei were not evident (Fig. 4D). Inhibiting caspases with ZVAD blocked killing by taxol but
not compound 6e (Fig. 4E). These observations suggest that 6e does not induce apoptosis.

Page 13 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
Fig. 4. Rapid cell death induced by compound 5e and 6e. (A) NCIꢀH522 cells were analyzed by
timeꢀlapse microscopy to determine time of death. Imaging was started at the time of drug
addition. (B) 6e does not affect tubulin. NCIꢀH522 were exposed to 10 ꢀM 6e for 6 hours and
then analyzed by confocal immunofluorescence with antibodies against alphaꢀtubulin and
borealin (to detect centromeres). DNA was stained with Hoechst 33342. For comparison, cells
were exposed to either epothilone B or taxol for 6 hours before analysis. (C) Morphological
features of 6eꢀinduced death. NCIꢀH522 cells exposed to compound 6e at 10 ꢀM were analyzed
by timeꢀlapse microscopy. Cell death is characterized by loss of contact with the substratum

Organic & Biomolecular Chemistry
Page 14 of 46
View Article Online
DOI: 10.1039/C7OB03086J
followed by loss of membrane integrity. (D) Example of cytoplasmic shrinkage during cell death
in the presence of 10 ꢀM 6e. Immunofluorescence was performed as described in “B”, except
cells were stained with phalloidin to detect actin. A wideꢀfield image is shown. (E) Lack of
protective effect of ZVAD on 6eꢀinduced death. Cells were coꢀtreated with ZVAD and either
10 ꢀM compound 6e, or 10 ꢀM taxol. Survival was assessed using methylene blue staining of
fixed adherent cells.
Ferroptotic cell death in cells exposed to compound 6e
Next, we investigated potential nonꢀapoptotic mechanisms of death in response to
compound 6e. Additional studies suggest that these compounds kill cells by inducing ferroptosis,
a novel ironꢀdependent, nonꢀapoptotic form of cell death.7 Ferroptosis is a recently described
mechanism of death characterized by a dependence on iron and ROS. Similarly to erastin, a
prototypical inducer of ferroptosis, iron chelators ciclopirox olamine and hydroxyurea abolished
killing of NCIꢀH522 cells by 6e (Fig. 5A). Sensitivity was enhanced by adding ferric citrate,
suggesting that iron is the relevant cation (Fig. 5B). Death of NCIꢀH522 cells induced by 6e was
also blocked by the free radical scavengers trolox and butylated hydroxyanisole (Fig. 5A). Thus,
compound 6e relies on ROS for cytotoxic activity. ROS are under multiple levels of control in
cells.16 Elevation of ROS, to the extent that they overwhelm antioxidant capacity, can induce
damage to proteins, lipids, and nucleic acids causing cell death. Cellular sources of ROS include
incomplete reduction of O2 during electron transport to form superoxide and direct generation of
superoxide by the membrane bound NADPH oxidases.16 Inhibiting NADPH oxidases using
diphenyleneiodonium chloride blocked killing by 6e, suggesting that these enzymes are

Page 15 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
important providers of ROS needed for killing (Fig. 5A). As predicted, exposure of NCIꢀH522
for 6 hours to compound 6e increased the level of intracellular ROS (Fig. 5C, D). ROS elevation
by 6e was dependent on iron and NADPH oxidase and was blocked by trolox (Fig. 5D).
Fig. 5. Compound 6e induces ferroptosis. (A) Effect of ferroptosis modulators on 6e toxicity.
NCIꢀH522 were exposed to 10 ꢀM 6e with or without the compounds indicated. Viability was
analyzed 24 hours later using methylene blue. CPO: Ciclopirox olamine; HU: Hydroxyurea;
BHA: Butylated hydroxyanisole; DIC: diphenyleneiodonium chloride. All modulators eliminated
6eꢀinduced killing (only 6e alone [+] versus DMSO [ꢀ] was significantly different using a
student’s tꢀtest). (B) Elevating iron enhances 6e toxicity. NCIꢀH522 cells were exposed to 6e
with or without 25 ꢀM ferric citrate. Viability was determined by methylene blue staining 2 days

Organic & Biomolecular Chemistry
Page 16 of 46
View Article Online
DOI: 10.1039/C7OB03086J
later. (C) Elevation of ROS in NCIꢀH522 cells exposed to 10 ꢀM compound 6e for 4 hours.
Representative microscopic fields of H2DCFDAꢀstained cells are shown. Dye was extracted and
quantified in (D).
Conclusions
During the course of our studies to develop structurally simplified epothilone analogues,
we discovered a new class of small molecules that kill selected cancer cell lines by a novel
mechanism of action. The cytotoxic activity is cell cycleꢀindependent and does not involve
apoptosis. The compounds had no effect on tubulin or mitochondrial function. Preliminary
studies show that they harness reactive oxygen species to kill cancer cells in an ironꢀdependent
manner showing the hallmarks of ferroptosis, a recently reported mechanism of action. Studies
are in progress to further confirm the ferroptotic cell death mechanism and to determine the
specific molecular targets of these promising “drugꢀlike” small molecule anticancer agents.
Experimental Section
General Information
All reactions were carried out under nitrogen using anhydrous solvents unless otherwise noted.
Tetrahydrofuran (THF) was distilled from Na and benzophenone under nitrogen prior to use.
NMR spectra were recorded on Varian VXRS 400 MHz, Varian INOVA 600 MHz and Bruker
Avance 600 MHz spectrometers and calibrated using residual undeuterated solvent as internal
reference. Optical rotations were recorded on Rudolph AUTOPOL III and Rudolph AUTOPOL
IV 589/586 polarimeters. Highꢀresolution mass spectra (HRMS) were recorded on a LCT
Electrospray mass spectrometer at the Central Instrument Facility, Mass Spectrometry

Page 17 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
Laboratory, the Wayne State University, Detroit, Michigan) and on a Micromass QꢀTof II
Electrospray mass spectrometer at the Mass Spectrometry and Proteomics Facility, the Ohio
State University, Columbus, Ohio. Ozone was generated on a Welsbach Model Tꢀ408
commercial ozone generator. A Biotage Initiator was used in microwave synthesis. Crude
products were purified by flash column chromatography on silica gel (standard grade, 40ꢀ63 µm,
230 x 400 mesh) purchased from Sorbent Technologies and using RediSep prepacked cartridges
from Teledyne ISCO Inc. on a Combiflash Companion. 1000 ꢀ Uniplates purchased from
Analtech Inc. were used in preparative thin layer chromatography using commercial solvents as
specified. HPLC analyses were performed on a Waters 1525 Binary Pump system with Waters
2487 Dual Wavelength Absorbance Detector on a Supleco C18 reverse phase column (5 ꢀm, 15
cm x 4.6 mm) and on a Symmetry C18 reverse phase column (5 ꢀ, 15 cm x 4.6 mm) using a
linear gradient of 60ꢀ100% acetonitrile in water over 10ꢀ20 min; flow rate of 1 mL/min and UV
detection at 254 nm. Structural integrity and purity of the test compounds were determined by
the composite of 1H and 13C NMR, HRMS and HPLC and were found to be > 95% pure.
A. Synthesis of (3S,6R,7S)-3,7-dihydroxy-4,4,6,8-tetramethyl-5-oxononoic acid (7) and the
(3S,6S,7R) diastereomer (7a).
(3S,6R,7S)-3,7-bis(tert-butyldimethylsilyloxy)-4,4,6,8-tetramethyl-5-oxononanal (11) and
(3S,6S,7R)-3,7-bis(tert-butyldimethylsilyloxy)-4,4,6,8-tetramethyl-5-oxononanal (11a)
Ozone was passed through a solution of diastereomers (7S)ꢀ3,7ꢀdihydroxyꢀ2,4,6,6ꢀ
o
tetramethyldecꢀ9ꢀenꢀ5ꢀone (10)8 (1.00 g, 2.12 mmol, 1 equiv) in DCM (12 mL) at ꢀ78 C until

Organic & Biomolecular Chemistry
Page 18 of 46
View Article Online
DOI: 10.1039/C7OB03086J
the reaction mixture reached a dark blue color. Nitrogen was bubbled through the solution for 15
min and triphenylphosphine (612 mg, 2.33 mmol, 1.1 equiv) was added. The reaction mixture
was stirred at room temperature overnight and concentrated in vacuo. The mixture of aldehydes
was separated by flash column chromatography on silica gel in 10% DCMꢀhexanes, monitored
by NMR, to obtain the (3S,6R,7S) diastereomer 11 (150 mg), the mixture of the two aldehydes
(416 mg), followed by (3S,6S,7R) diastereomer 11a (245 mg) as colorless oils with an overall
yield of 69%.
Data for the (3S,6R,7S) diastereomer 11: 1H NMR (400 MHz, CDCl3): δ 9.76 (s, 1H), 4.57 (t, J =
5.2 Hz, 1H), 3.70 (d, J = 7.6 Hz, 1H), 3.07ꢀ3.00 (m, 1H), 2.51ꢀ2.37 (m, 3H), 1.43ꢀ1.33 (m, 1H),
13
1.17 (m, 3H), 1.03 (m, 3H), 0.66 (m, 9H), 0.01 (m, 6H); C NMR (CDCl3, 100 MHz): δ 218.18,
201.55, 77.85, 70.42, 53.92, 49.24, 45.44, 33.25, 26.43, 26.20, 26.12, 22.71, 21.31, 19.44, 18.76,
18.31, 16. 16.035, ꢀ3.24, ꢀ3.51, ꢀ3.10, ꢀ4.22.
Data for the (3S,6S,7R) diastereomer 11a: 1H NMR (400 MHz, CDCl3): δ 9.71 (s, 1H), 4.45 (t, J
= 4.8 Hz, 1H), 3.70 (d, J = 6.8 Hz, 1H), 3.05ꢀ3.01 (m, 1H), 2.48ꢀ2.32 (m, 3H), 1.44ꢀ1.38 (m,
13
1H), 1.14 (s, 3H), 0.98 (d, J = 8.0 Hz, 3H), 0.81 (m, 9H), 0.01 (m, 6H); C NMR (CDCl3, 100
MHz): δ 218.98, 201.47, 77.87, 71.25, 53.80, 49.79, 46.09, 46.05, 39.92, 32.89, 29.92, 26.46,
26.17, 26.10, 26.06, 24.12, 21.51, 18.791, 18.74, 18.32, 16.09, 15.86, ꢀ3.15, ꢀ3.46, ꢀ3.86, ꢀ4.28.
(3S,6R,7S)-3,7-bis(tert-butyldimethylsilyloxy)-4,4,6,8-tetramethyl-5-oxononanoic acid (7).
To a solution of aldehyde 11 (710 mg, 1.50 mmol, 1 equiv) and 2ꢀmethylꢀ2ꢀbutene (7.6 mL) in
tꢀbutanol (32 mL) was added a solution of NaClO2 (1.244 g, 13.76 mmol, 9.1 equiv) and

Page 19 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
NaH2PO4 (1.248 g, 10.38 mmol, 6.9 equiv) in water (9 mL) dropwise. The reaction mixture was
stirred for 1 h at RT and the reaction was quenched with saturated aqueous ammonium chloride
(40 mL) and water (40 mL). The reaction mixture was extracted with EtOAc (3 x 15 mL). The
combined organic extract was dried over anhydrous sodium sulfate and concentrated in vacuo.
1
The crude product was taken directly to the next step without further purification. H NMR (400
MHz, CDCl3): δ 4.52ꢀ4.49 (m, 1H), 3.73 (dd, J = 8.0 Hz, 2.0 Hz, 1H), 3.09ꢀ3.01 (m, 1H), 2.47
(dd, J = 16.4 Hz, 3.6 Hz, 1H), 2.33 (dd, J = 16.4 Hz, 6.8 Hz, 1H), 1.46ꢀ1.38 (m, 1H), 1.16 (m,
3H), 1.01 (m, 3H), 0.98ꢀ0.82 (m, 9H), 0.03 (m, 6H).
(3S,6R,7S)-3,7-dihydroxy-4,4,6,8-tetramethyl-5-oxononanoic acid.
o
A solution of the carboxylic acid 7 (350 mg, 0.72 mmol) in DCM (15 mL) was cooled to 0 C.
o
TFA (3.0 mL, 20% in DCM) was added and the reaction mixture was stirred at 4 C for 25 h.
Water was added (5 mL) and the reaction was evaporated to dryness under reduced pressure.
The residue was dried azeotropically with toluene to obtain the desilylated (3S,6R,7S) carboxylic
acid (168 mg, 90%) as a colorless oil: 1H NMR (400 MHz, CDCl3): δ 4.23 (dd, J = 10.0 Hz, 2.4
Hz, 1H), 3.33 (dd, J = 8.8 Hz, 1.6 Hz, 1H), 3.26ꢀ3.21 (m, 1H), 2.51ꢀ2.37 (m, 3H), 1.69ꢀ1.63 (m,
1H), 1.15 (d, J = 1.6 Hz, 6H), 1.03 (d, J = 6.8 Hz, 3H), 0.98 (d, J = 6.8 Hz, 3H), 0.85 (d, J = 6.8
Hz, 3H); 13C NMR (CDCl3, 100 MHz): δ 222.47, 177.25, 76.79, 72.47, 52.33, 41.079, 36.71,
30.66, 21.61, 19.54, 19.30, 19.17, 10.61.
(3S,6S,7R)-3,7-bis(tert-butyldimethylsilyloxy)-4,4,6,8-tetramethyl-5-oxononanoic acid (7a).

Organic & Biomolecular Chemistry
Page 20 of 46
View Article Online
DOI: 10.1039/C7OB03086J
To a solution (3S,6S,7R)ꢀ3,7ꢀbis(tertꢀbutyldimethylsilyloxy)ꢀ4,4,6,8ꢀtetramethylꢀ5ꢀoxononanal
11 a (282 mg, 0.569 mmol, 1 equiv) and 2ꢀmethylꢀ2ꢀbutene (3.0 mL) in tꢀbutanol (12 mL) was
added a solution of NaClO2 (497 mg, 5.50 mmol, 9.1 equiv.) and NaH2PO4 (497 mg, 4.15 mmol,
6.9 equiv.) in water (4 mL) dropwise. The reaction was stirred at RT for 1 h and the reaction
was quenched with saturated aqueous ammonium chloride (15 mL) and water (15 mL). The
crude reaction mixture was extracted with EtOAc (3 x 10 mL). The combined organic layers
were dried over anhydrous sodium sulfate, filtered, and concentrated in vacuo. The crude
product was passed through a silica gel plug to obtain the (3S,6S,7R) carboxylic acid 7a (241 mg,
1
87%) as a colorless oil: H NMR (400 MHz, CDCl3): δ 4.39 (m, 1H), 3.72 (dd, J = 8.0 Hz, 1.2
Hz, 1H), 3.08 (m, 1H), 2.46 (dd, J = 16.4 Hz, 2.8 Hz, 1H), 2.29 (dd, J = 16.4 Hz, 6.8 Hz, 1H),
1.45ꢀ1.42 (m, 1H), 1.17 (m, 3H), 1.08 (m, 3H), 0.88ꢀ0.73 (m, 9H), 0.03 (m, 6H); 13C (CDCl3,
100 MHz): δ 218.51, 178.65, 77.87, 73.40, 53.81, 46.07, 40.43, 32.94, 26.48, 26.22, 23.73,
21.53, 19.04, 18.80, 18.41, 16.18, 15.90, ꢀ3.18, ꢀ3.46, ꢀ4.05.
(3S,6S,7R)-3,7-dihydroxy-4,4,6,8-tetramethyl-5-oxononanoic acid
A solution the carboxylic acid 7a (882 mg, 1.81 mmol) in DCM (58 mL) was cooled to 0 oC and
o
TFA (11.6 mL, 20% in DCM) was added. The reaction mixture was stirred at 4 C for 25 h.
Water (15 mL) was added and the reaction mixture was evaporated to dryness under reduced
pressure. The residue was dried by azeotropically with toluene to obtain the desilylated
(3S,6S,7R) carboxylic acid (450 mg, 95%) as a colorless oil: 1H NMR (400 MHz, CDCl3): δ 4.26
(dd, J = 10.0 Hz, 2.4 Hz, 1H), 3.30ꢀ3.22 (m, 2H), 2.53ꢀ2.40 (m, 2H), 1.70ꢀ1.63 (m, 1H), 1.20 (s,
3H), 1.15 (s, 3H), 1.06 (d, J = 7.2 Hz, 3H), 1.00 (d, J = 6.4 Hz, 3H), 0.86 (d, J = 7.2 Hz, 3H); 13C

Page 21 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
NMR (CDCl3, 100 MHz): δ 222.61, 176.67, 76.58, 72.44, 52.16, 41.33, 36.38, 30.82, 21.60,
19.66, 19.21, 19.05, 10.54.
B. Synthesis of 2-substituted thiazole cyclopentenols (6a- f).
2-Methyl-4-bromothiazole (13).
Synthesized using conditions described by Hofle et al.10 (0.765, 52%) reddish oil: TLC Rf = 0.68
(20% EtOAcꢀhexanes); 1H NMR (400 MHz, CDCl3): δ 7.04 (s, 1H), 2.71 (s, 3H); 13C NMR
(CDCl3, 100 MHz): δ 167.20, 123.94, 116.30, 19.41.
2-(Methylthio)-4-bromothiazole (14).
Synthesized using conditions described by Nicolaou et al.11 (1.415 g, 94%) yellow solid: TLC Rf
= 0.48 (2% EtOAcꢀhexanes); 1H NMR (600 MHz, CDCl3): δ 7.05 (s, 1H), 2.68 (s, 3H); 13C
NMR (CDCl3, 100 MHz): δ 124.44, 115.68, 16.85.
2-(Piperidin-1-yl)-4-bromothiazole (15).
Synthesized using conditions described by Nicolaou et al.11 (1.30, 86%) yellow solid: TLC Rf =
0.43 (5% EtOAcꢀhexanes); 1H NMR (400 MHz, CDCl3): δ 6.35 (s, 1H), 3.42 (d, J = 5.6 Hz, 4H),
1.63 (s, 6H); 13C NMR (CDCl3, 100 MHz): δ 171.03, 121.70, 102.97, 49.26, 25.14, 24.08.

Organic & Biomolecular Chemistry
Page 22 of 46
View Article Online
DOI: 10.1039/C7OB03086J
2-(Phenylethynyl)-4-bromothiazole (16).
A mixture of 2,4ꢀdibromothiazole 12 (2.92 g, 12.0 mmol, 1 equiv), Pd(Ph3)4 (0.685 g, 0.60
mmol, 5 mol%), and CuI (0.228 g, 0.12 mmol, 10 mol %) was placed in a threeꢀnecked round
bottom flask under nitrogen. Anhydrous THF (30 mL) was added, followed by N,Nꢀ
diisopropylamine (2.55 mL, 18.02 mmol, 1.5 equiv). A solution of phenylacetylene (2.0 g, 17.98
mmol, 1.5 equiv) in anhydrous THF (6 mL) was slowly added to the reaction mixture via a
syringe pump over 7 h. The reaction mixture was stirred for an additional 9 h. Water (50 mL)
was added and the crude reaction mixture was extracted with ethyl acetate (3 x 20 mL). The
combined organic extract was washed with brine, dried over anhydrous sodium sulfate, and
concentrated in vacuo. The crude product was purified by flash column chromatography on
silica gel in 5% EtOAc in hexanes and recrystallized from EtOAcꢀhexanes to obtain product 16
o
(3.174 g, 86%) as brown crystals: TLC Rf = 0.59 (10 % EtOAcꢀhexanes); mp 79ꢀ80 C; IR vmax
:
1
3300, 3000, 2900, 2350 cmꢀ1; H NMR (600 MHz, acetoneꢀd6): δ 7.79 (s, 1H), 7.66 (m, 2H),
7.51 (m, 3H); 13C NMR (CDCl3, 100 MHz): δ 149.93, 132.249, 130.12, 128.78, 126.11, 121.10,
118.82, 95.69, 81.60; HRMS (m/z): [M + Na]+ calcd for C11H6NSBr, 285.9302; found,
285.9314.
2−((Trimethylsilyl)ethynyl)-4-bromothiazole (17).
A mixture of 2,4ꢀdibromothiazole 12 (3.000 g, 12.34 mmol, 1 equiv), triphenylphosphine (486
mg, 1.851 mmol, 5 mol%), CuI (120 mg, 0.617 mmol, 5 mol%), and Pd(Ph3)2Cl2 (120 mg, 0.173
mmol, 1.4 mol %) was placed in a threeꢀneck round bottom flask under nitrogen. Anhydrous
toluene (42 mL) was added followed by anhydrous Et3N (2.2 mL, 15.04 mmol, 1.3 equiv) and

Page 23 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
trimethylsilylacetylene (2.6 mL, 18.51 mmol, 1.5 equiv). The reaction mixture was refluxed at
o
140 C for 2 d. It was poured into water (50 mL) and extracted with EtOAc (3x 20 mL). The
combined organic extract was dried over anhydrous sodium sulfate and concentrated in vacuo.
The crude product was purified by flash chromatography on silica gel in DCMꢀhexanes to obtain
o
17 (2.63 g, 81%) as a brownish/red solid: TLC Rf = 0.72 (10 % EtOAcꢀhexanes); mp 37 C; IR
vmax: 3300, 2900, 2250 cmꢀ1; 1H NMR (400 MHz, CDCl3): δ 7.18 (s, 1H), 0.24 (s, 9H); 13C
NMR (CDCl3, 100 MHz): δ 149.54, 125.87, 118.85, 103.03, 95.52, ꢀ0.41; HRMS (m/z): [M +
H]+ calcd for C8H11NSSiBr, 259.9565; found, 259.9565.
2-Methyl-3-(2-methylthiazol-4-yl)cyclopent-2-enone (20a).
A solution of 2ꢀmethylꢀ4ꢀbromothiazole 15 (351 mg, 1.97 mmol, 1 equiv) in anhydrous ether (3
mL) was cooled to ꢀ78 oC and nꢀBuLi (1.30 mL of 2.5 M solution in THF, 3.35 mmol, 1.7 equiv)
o
was added dropwise. The reaction mixture was stirred at ꢀ78 C for 2 h and a solution of
trimethytin chloride (785 mg, 3.94 mmol, 2 equiv) in anhydrous ether (2 mL) was added
dropwise. The reaction mixture was stirred for an additional 1 h and allowed to warm to room
temperature slowly. It was diluted with hexanes, passed through a silica gel plug deactivated
with 5% Et3Nꢀhexanes, and eluted with EtOAc. The partially purified product was concentrated
in vacuo to obtain a reddish oil which was immediately taken to the next step. 1H NMR (600
MHz, CDCl3): δ 7.19 (s, 1H), 2.75 (s, 3H), 0.33 (t, J = 3.6 Hz, 9H). A mixture of 3ꢀiodoꢀ2ꢀ
methylcyclopentꢀ2ꢀenone 10 (350 mg, 1.57 mmol, 0.8 equiv) and Pd(Ph3)4 (455 mg, 0.394
mmol, 20 mol%) was placed in a ꢀw vial under nitrogen. A solution of the tin derivative
prepared above in anhydrous DMF (4.5 mL) was added via a syringe, and the vial was heated for

Organic & Biomolecular Chemistry
Page 24 of 46
View Article Online
DOI: 10.1039/C7OB03086J
2 h 15 min at 145 oC in the microwave synthesizer. The reaction mixture was poured into water
(15 mL) and extracted with EtOAc (3 x 5 mL). The combined organic extract was dried over
anhydrous sodium sulfate and concentrated in vacuo. The crude product was purified by flash
chromatography on silica gel in EtOAcꢀhexanes and recrystallized from EtOAcꢀhexanes to
obtain 20 a (336 mg, 88%) as tan crystals: TLC Rf = 0.39 (40% EtOAcꢀhexanes); mp 89ꢀ90 oC;
1H NMR (400 MHz, CDCl3): δ 7.45 (s, 1H), 2.95ꢀ2.92 (m, 2H), 2.73 (s, 3H), 2.49ꢀ2.47 (m, 2H),
2.09 (t, J = 2.0 Hz, 3H); 13C NMR (CDCl3, 100 MHz): δ 210.24, 165.80, 158.67, 152.28,
136.60, 120.32, 33.89, 28.24, 19.56, 10.33; HRMS (m/z): [M + Na]+ calcd for C10H11NOS,
216.0459; found, 216.0461.
2-Methyl-3-(2-(methylthio)thiazol-4-yl)cyclopent-2-enone (20b).
A solution of 2ꢀthiomethylꢀ4ꢀbromothiazole 14 (300 mg, 1.43 mmol, 1 equiv) in anhydrous ether
o
(10 mL) was cooled to ꢀ78 C. nꢀBuLi (1.80 mL, 1.6 M in hexanes, 2.86 mmol, 2 equiv) was
o
added dropwise and the reaction mixture was stirred at ꢀ78 C for 2 h. A solution of trimethyltin
chloride (708 mg, 3.57 mmol, 2.5 equiv) in anhydrous ether (3 mL) was added dropwise. The
o
reaction mixture was stirred at ꢀ78 C for an additional 1 h and was allowed to warm slowly to
room temperature. The reaction mixture was diluted with hexanes, passed through a silica gel
plug deactivated with 5% Et3Nꢀhexanes, and eluted with EtOAc. The partially purified product
was concentrated in vacuo to obtain a reddish oil which was taken immediately to the next step.
1H NMR (400 MHz, CDCl3): δ 7.22 (s, 1H), 2.69 (s, 3H), 0.32 (t, J = 28.0 Hz, 9H). 3ꢀIodoꢀ2ꢀ
methylcyclopentꢀ2ꢀenone 10 (318 mg, 1.43 mmol, 1 equiv) and Pd(Ph3)4 (330 mg, 0.286 mmol,
20 mol%) were placed in a ꢀw vial under nitrogen. A solution of the tin derivative prepared

Page 25 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
above in anhydrous DMF (15 mL) was added via syringe, and the vial was heated for 2 h in the
o
microwave synthesizer at 145 C. The reaction mixture was poured into water (30 mL) and
extracted with EtOAc (3 x 10 mL). The combined organic extract was dried over anhydrous
sodium sulfate and concentrated in vacuo. The product was purified by flash chromatography on
silica gel in EtOAcꢀhexanes, and recrystallized from EtOAcꢀhexanes to obtain the product 20b
o
1
(148 mg, 46%) as a pale yellow solid: TLC Rf = 0.60 (40% EtOAcꢀhexanes); mp 102 C; H
NMR (600 MHz, CDCl3): δ 7.46 (s, 1H), 2.92ꢀ2.90 (m, 2H), 2.73 (s, 3H), 2.52ꢀ2.50 (m, 2H),
2.14 (t, J = 2.0 Hz, 3H); 13C NMR (100 MHz, CDCl3): δ 210.27, 166.73, 157.83, 152.64, 137.03,
119.64, 33.93, 27.92, 16.71, 10.40; HRMS (m/z): [M + Na]+ calcd for C10H11NOS, 248.0180;
found, 248.0189.
2-Methyl-3-(2-(piperidin-1-yl)thiazol-4-yl)cyclopent-2-enone (20c).
A solution of 2ꢀpiperdinꢀ1ꢀylꢀ4ꢀbromothiazole 15 (550 mg, 2.22 mmol, 1 equiv) in anhydrous
o
ether (19 mL) under nitrogen was cooled to ꢀ78 C. nꢀBuLi (1.60 mL, 1.6 M in hexanes, 2.66
o
mmol, 1.2 equiv) was added dropwise and the reaction mixture was stirred at ꢀ78 C for 2 h. A
solution of trimethyltin chloride (590 mg, 3.33 mmol, 1.5 equiv) in anhydrous ether (7 mL) was
o
added dropwise. The reaction mixture was stirred at ꢀ78 C for an additional 1 h and was
allowed to warm slowly to room temperature. The reaction mixture was diluted with hexanes,
passed through a silica gel plug deactivated with 5% Et3Nꢀhexanes, and eluted with EtOAc. The
partially purified product was concentrated in vacuo to obtain a yellowish oil which was taken
1
immediately to the next step. H NMR (400 MHz, CDCl3): δ 6.56 (s, 1H), 3.47 (m, 4H), 1.65
(m, 6H), 0.27 (t, J = 20 Hz, 9H). 3ꢀIodoꢀ2ꢀmethylcyclopentꢀ2ꢀenone 10 (338 mg, 1.52 mmol, 1

Organic & Biomolecular Chemistry
Page 26 of 46
View Article Online
DOI: 10.1039/C7OB03086J
equiv) and Pd(Ph3)4 (352 mg, 0.304 mmol, 20 mol%) were placed in a ꢀw vial under nitrogen.
A solution of the tin derivative prepared above in anhydrous DMF (10 mL) was added via
o
syringe. The vial was heated for 2 h in the microwave synthesizer at 140 C. The reaction
mixture was poured into water (20 mL) and extracted with EtOAc (3 x 8 mL). The combined
organic extract was dried over anhydrous sodium sulfate and concentrated in vacuo. The crude
product was purified by flash chromatography on silica gel in EtOAcꢀhexanes and recrystallized
from EtOAcꢀhexanes to obtain 20 c (341 mg, 58%) as yellow crystals: TLC Rf = 0.57 (40%
o
1
EtOAcꢀhexanes); mp 108 C; H NMR (600 MHz, CDCl3): δ 6.88 (s, 1H), 3.49 (m, 4H), 2.85
(m, 2H), 2.46 (m, 2H), 2.13 (t, J = 2.0 Hz, 3H), 1.66 (m, 6H); 13C NMR (CDCl3, 100 MHz):
δ 221.25, 210.71, 170.84, 159.11, 149.68, 136.08, 109.85, 49.76, 33.93, 27.60, 25.33, 24.37,
10.39. HRMS (m/z): [M + Na]+ calcd for C14H18N2O, 285.1038; found, 285.1046.
2-Methyl-3-(2-(phenylethynyl)thiazol-4-yl)cyclopent-2-enone (20d).
A solution of 2ꢀ(phenylethynyl)ꢀ4ꢀbromothiazole 16 (1.000 g, 3.78 mmol, 1 equiv) in anhydrous
o
ether (15 mL) was cooled to ꢀ78 C and nꢀBuLi (2.3 mL, 2.5 M in hexanes, 5.67 mmol, 1.5
o
equiv) was added dropwise. The reaction mixture was stirred at ꢀ78 C for 2 h and a solution of
trimethyltin chloride (1.503 g, 7.56 mmol, 2 equiv) in anhydrous ether (5 mL) was added
o
dropwise. The reaction mixture was stirred for an additional 1 h at ꢀ78 C before it was allowed
to warm to room temperature slowly. The reaction mixture was diluted with hexanes, passed
through a silica gel plug deactivated with 5% Et3Nꢀhexanes, and eluted with EtOAc. The
partially purified product was concentrated in vacuo to obtain a reddish oil which was used
1
immediately in the next step. H NMR (400 MHz, CDCl3): δ 7.59 (m, 2H), 7.38 (m, 3H), 0.37

Page 27 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
(t, J = 27.6 Hz, 9H). 3ꢀIodoꢀ2ꢀmethylcyclopentꢀ2ꢀenone 10 (678 mg, 3.02 mmol, 0.8 equiv) and
Pd(Ph3)4 (873 mg, 0.756 mmol, 20 mol%) were placed in a ꢀw vial under nitrogen. A solution of
the tin derivative prepared above in anhydrous DMF (15 mL) was added via syringe. The vial
o
was heated at for 2 h 15 min in the microwave synthesizer at 145 C. The reaction mixture was
poured into water (50 mL) and extracted with EtOAc (3 x 20 mL). The combined organic
extract was dried over anhydrous sodium sulfate and concentrated in vacuo. The crude product
was purified by flash chromatography on silica gel in EtOAcꢀhexanes and recrystallized from
EtOAcꢀhexanes to obtain the product 20d (816 mg, 77%) as a brown solid: TLC Rf = 0.45 (40%
o
1
EtOAcꢀhexanes); mp 117 C; IR vmax: 3320, 3200, 2900, 2250, 1800, 1750 cmꢀ1; H NMR (400
MHz, CDCl3): δ 7.64 (s, 1H), 7.61 (m, 2H), 7.38 (m, 3H), 3.03 (m, 2H), 2.54 (m, 2H), 2.13 (t, J
= 2.0 Hz, 3H); 13C NMR (CDCl3, 100 MHz): δ 210.09, 157.99, 153.08, 148.76, 137.61, 132.27,
130.03, 128.76, 121.87, 121.24, 95.08, 82.16, 33.96, 28.40, 10.44; HRMS (m/z): [M + Na]+
calcd for C17H13NOS, 302.0616; found, 302.0620.
2-Methyl-3-(2-((trimethylsilyl)ethynyl)thiazol-4-yl)cyclopent-2-enone (20e).
A solution of 2−((trimethylsilyl)ethynyl)ꢀ4ꢀbromothiazole 17 (100 mg, 0.384 mmol, 1 equiv) in
o
anhydrous ether (3 mL) under nitrogen was cooled to ꢀ78 C and tꢀBuLi (240 ꢀL, 1.6 M, 0.786
mmol, 1 equiv) was added dropwise. The reaction was stirred at ꢀ78 oC for 1 h and a solution of
trimethyltin chloride (150 mg, 0.768 mmol, 2 equiv) in anhydrous ether (2 mL) was added via
o
syringe. The reaction mixture was stirred at ꢀ78 C for an additional 1 h and was allowed to
warm to room temperature slowly. The reaction mixture was diluted with hexanes, passed
through a silica gel plug deactivated with 5% Et3Nꢀhexanes, and eluted with EtOAc. It was

Organic & Biomolecular Chemistry
Page 28 of 46
View Article Online
DOI: 10.1039/C7OB03086J
1
concentrated in vacuo to obtain a yellow oil, which was immediately taken to the next step. H
NMR (600 MHz, CDCl3): δ 7.33 (s, 1H), 0.34 (t, J = 14.8 Hz, 9H), 0.25 (m, 3H). 3ꢀIodoꢀ2ꢀ
methylcyclopentꢀ2ꢀenone 10 (83 mg, 3.84 mmol, 1 equiv) and Pd(Ph3)4 (88 mg, 0.078 mmol, 20
mol%) were placed in a ꢀw vial under nitrogen. A solution of tin derivative prepared above in
anhydrous DMF (2 mL) was added via syringe. The ꢀw vial was heated for 2 h 15 min in the
o
microwave synthesizer at 145 C. The reaction mixture was poured into water (30 mL) and
extracted with EtOAc (3 x 10 mL). The combined organic extract was dried over anhydrous
sodium sulfate and concentrated in vacuo. The crude product was purified by flash
chromatography on silica gel in EtOAcꢀhexanes and recrystallized from DCMꢀhexanes to obtain
20e as tan star shaped crystals (50 mg, 47%): TLC Rf = 0.63 (40% EtOAcꢀhexanes); mp 108ꢀ
110 oC; IR vmax: 3300, 2900, 2200, 1750 cmꢀ1; 1H NMR (600 MHz, CDCl3): δ 7.58 (s, 1H), 2.99
(t, J = 2.0 Hz, 2H), 2.52 (m, 2H), 2.10 (d, J = 2.0 Hz, 3H), 0.28 (s, 9H); 13C NMR (CDCl3, 100
MHz): δ 209.71, 157.66, 152.65, 148.22, 137.37, 121.79, 102.05, 96.15, 33.74, 28.18, 10.23, ꢀ
0.46; HRMS (m/z): [M + Na]+ calcd for C14H17NOSSi, 298.0698; found, 298.0680.
3-(2-Ethynylthiazol-4-yl)-2-methylcyclopent-2-enone (21).
A solution of 2ꢀmethylꢀ3ꢀ(2ꢀ((trimethylsilyl)ethynyl)thiazolꢀ4ꢀyl)cyclopentꢀ2ꢀenone 20e (50 mg,
0.182 mmol, 1 equiv) and K2CO3 (3 mg, 0.022 mmol, 12 mol%) in methanol (1 mL) was stirred
for 5 min. The crude product was poured into water (8 mL) and extracted with EtOAc (3 x 3
mL). The combined organic extract was dried over anhydrous sodium sulfate and concentrated
in vacuo. The product was recrystallized from DCMꢀhexanes to obtain 21 as tan needles: TLC
o
1
Rf = 0.33 (40% EtOAcꢀhexanes); mp 138ꢀ139 C; IR: 3000, 2900, 2250, 1750 cmꢀ1; H NMR

Page 29 of 46
Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C7OB03086J
(400 MHz, CDCl3): δ 7.62 (s, 1H), 3.52 (s, 3H), 2.99ꢀ2.96 (m, 2H), 2.53ꢀ2.51 (m, 2H), 2.11 (t, J
13
= 2.0 Hz, 3H); C NMR (CDCl3, 100 MHz): δ 209.98, 157.56, 153.08, 147.50, 137.84, 121.90,
83.24, 76.31, 33.92, 28.31, 10.38; HRMS (m/z): [M + Na]+ calcd for C11H9NOS, 226.0303;
found, 226.0298.
3-(2-(1H-1,2,3-Triazol-4-yl)-2-methylcyclopent-2-enone (22).
3ꢀ(2ꢀEthynylthiazolꢀ4ꢀyl)ꢀ2ꢀmethylcyclopentꢀ2ꢀenone 21 (60 mg, 0.295 mmol, 1 equiv) and CuI
(3 mg, 0.015 mmol, 5 mol%) were placed in a ꢀw vial under nitrogen. Anhydrous DMF (3.6
mL) and anhydrous MeOH (440 ꢀL, 9:1 ratio DMF/MeOH) were added, followed by
azidotrimethylsilane (58 ꢀL, 0.443 mmol, 1.5 equiv). The vial was heated for 2 h in the
o
microwave synthesizer at 140 C. The reaction mixture was poured into water (10 mL) and
extracted with EtOAc (3 x 4 mL). The combined organic extract was dried over anhydrous
sodium sulfate and concentrated in vacuo. The crude compound was purified by flash
chromatography on silica gel in EtOAcꢀhexanes and recrystallized from methanol to obtain
product 22 (51 mg, 70%) as a white solid: TLC Rf = 0.34 (50% EtOAcꢀhexanes); mp 241ꢀ243
1
oC; H NMR (400 MHz, CDCl3): δ 8.25 (S, 1H), 7.67 (s, 1H), 3.04ꢀ2.98 (m, 2H), 2.57ꢀ2.55 (m,
2H), 2.20 (d, J = 4.0 Hz, 3H); 13C NMR (DMSOꢀd6, 100 MHz): δ 208.87, 158.46, 157.43,
152.32, 141.26, 135.32, 141.26, 135.47, 122.25, 33.25, 27.46, 9.99; HRMS (m/z): [M + Na]+
calcd for C11H10N4OS, 269.0473; found, 269.0489.
2-Methyl-3-(2-(1-methyl-1H-triazol -4-yl)thiazol-4-yl)cyclopent-2-enone (20f).