Thio-ether functionalized glycolipid amphiphilic compounds reveal a potent activator of SK3 channel with vasorelaxation effect

Article abstract of DOI:10.1039/d1ob00021g

The modulation of SK3 ion channels can be efficiently and selectively achieved by using the amphiphilic compound Ohmline (a glyco-glycero-ether-lipid). We report herein a series of Ohmline analogues featuring the replacement of one ether function by a thi

Full text of DOI:10.1039/d1ob00021g

Organic &
Biomolecular Chemistry
View Article Online
View Journal | View Issue
PAPER
Thio-ether functionalized glycolipid amphiphilic
compounds reveal a potent activator of SK3
channel with vasorelaxation effect†
Cite this: Org. Biomol. Chem., 2021,
19, 2753
Charlotte M. Sevrain,a Delphine Fontaine,b Alicia Bauduin,a Maxime Guéguinou,b
Bei Li Zhang,c Aurélie Chantôme,b Karine Mahéo,b Côme Pasqualin,c
Véronique Maupoil,c Hélène Couthon,a Christophe Vandierb and
a
Paul-Alain Jaffrès
*
The modulation of SK3 ion channels can be efficiently and selectively achieved by using the amphiphilic
compound Ohmline (a glyco-glycero-ether-lipid). We report herein a series of Ohmline analogues featur-
ing the replacement of one ether function by a thioether function located at the same position or shifted
close to its initial position. The variation of the lipid chain length and the preparation of two analogues
featuring either one sulfoxide or one sulfone moiety complete this series. Patch clamp measurements
indicate that the presence of the thioether function (compounds 7 and 17a) produces strong activators of
SK3 channels, whereas the introduction of a sulfoxide or a sulfone function at the same place produces
amphiphiles devoid of an effect on SK3 channels. Compounds 7 and 17a are the first amphiphilic com-
pounds featuring strong activation of SK3 channels (close to 200% activation). The cytosolic calcium con-
centration determined from fluorescence at 3 different times for compound 7b (13 min, 1 h, 24 h)
revealed that the effect is different suggesting that the compound could be metabolized over time. This
compound could be used as a strong SK3 activator for a short time. The capacity of 7b to activate SK3
was then used to induce vasorelaxation via an endothelium-derived hyperpolarization (EDH) pathway. For
the first time, we report that an amphiphilic compound can affect the endothelium dependent
vasorelaxation.
Received 5th January 2021,
Accepted 26th February 2021
DOI: 10.1039/d1ob00021g
rsc.li/obc
bition) were developed to assess the role of SKCa channels and
for potent medical applications.
Introduction
Among the wide diversity of potassium channels,1 SKCa chan-
Briefly, activation of SKCa channels was used concomitantly
nels are activated by and sensitive to cytosolic calcium concen- with N-methyl-D-aspartate receptor antagonists to produce anti-
tration. This family of ion channels is composed of 3 channel nociceptive effects.7 Other works have shown that the acti-
isoforms (SK1, SK2, and SK3 also identified as KCa2.1, KCa2.2, vation of SK2/SK3 channels improved the mobility of trans-
and KCa2.3 with the genes KCNN1, KCNN2, and KCNN3). genic SCA2 mice,8 and the activation of SK3 reduced the
These channels are involved in different physiological roles inflammatory response activated by microglia.9 In addition,
(e.g., regulation of neuronal excitability and sinoatrial node2 activators of SK3 channels may be used in pathologies in
and implication in blood pressure regulation3) and pathologi- which SK3 induced hyperpolarization control excitation–con-
cal situations (e.g. neuronal disease,4 cancer,5 and neuro- traction coupling such as to prevent preterm labor,10–12 to
pathy6). Modulators of SKCa channels (activation or inhi- prevent overactive bladder13 or to treat systemic hypertension
by activating the endothelium-derived hyperpolarizing factor
(EDHF) and nitric oxide-mediated vasodilation.14 On the other
aUniv. Brest, CNRS, CEMCA UMR 6521, 6 Avenue Victor Le Gorgeu, Brest, F-29238
Brest, France. E-mail: pjaffres@univ-brest.fr
bUniv. Tours, INSERM, UMR1069, 37000 Tours, France
cUniv. Tours, Laboratoire STIM EA 7349, and Groupe Physiologie des Cellules
Cardiaques et Vasculaires, Faculté de Pharmacie, 31 avenue Monge, 37200 Tours,
France
hand, to mention a few examples of the effect of the inhibition
of the SK3 channel, it is worth mentioning the possibility to
reduce migration/invasion of microglia;15 an SK3 blocker
could reduce taxane-induced peripheral neuropathy.6 It is
noteworthy that in some cancer cells the expression of the SK3
channel is modified or abnormally expressed (not expressed in
some epithelial cells compared to cancer epithelial cells). For
†Electronic supplementary information (ESI) available. See DOI: 10.1039/
d1ob00021g
This journal is © The Royal Society of Chemistry 2021
Org. Biomol. Chem., 2021, 19, 2753–2766 | 2753

View Article Online
Paper
Organic & Biomolecular Chemistry
instance, in cancer cells (e.g. MDA-MB-435s), the SK3 channel tor of SK3 channels. This compound inhibited SK3 currents by
is expressed, whereas in non-tumor cells, it is not. 73% (effect observed 2 minutes after adding Ohmline at 10 µM
Interestingly, biopsies of cancer cells have shown that SK3 and measured by the patch clamp technique) and inhibited
channels are also expressed in melanoma,16 breast, prostate,17 SK3-dependent cancer cell migration (corresponding to 50% of
and colon cancer.18 When the SK3 channel is expressed in MDA-MB-435s cell migration) at a lower concentration (300
cancer cells, it promotes at least cell migration.19 These results nM) for 24 hours.17 Interestingly this compound is one of the
suggested developing inhibitors of SK3 channels with the aim most selective inhibitors of SK3 since it has a weak effect on
to reduce cancer cell migration. As detailed in a recent review,5 SK1 (−20%) and no effect on both SK2 and IKCa (intermediate
the first strategy to modulate SK3 currents involved the use of conductance calcium activated potassium channel). This com-
peptides (e.g. apamin20,21) or heterocyclic compounds featur- pound is neither a pore blocker (cork effect; observed for tetra-
ing cationic moieties (e.g. UCL168422 and dequalinium23). The ethylammonium31) nor a modulator of the sensitivity of cyto-
inhibition can be also achieved by other classes of heterocyclic solic calcium concentration.24 The amphiphilic nature of
compounds that modified the sensitivity of cytosolic calcium Ohmline that belongs to the class of ether lipids32 suggests a
concentration [Ca]cyt. (e.g. NS8593 24). This action likely occurs different mechanism of action when compared to the other
by interfering with the gating mechanism.25–27 On the other modulators of SK3 channels. The alkyl chain acts as a mem-
hand, other heterocyclic compounds were identified as activa- brane anchor and the glycerol moiety and the lactose unit
tors of SK3 channels (e.g. CyPPA 28 and NS13001 8).
interact with the polar groups of the constituents of the
More recently, we found that some amphiphilic compounds plasma membranes (e.g. phospholipid, cholesterol) as recently
inhibit SK3 channel activity with an interesting selectivity.29,30 shown by 2H NMR and molecular dynamic simulation.33 On
Ohmline (Fig. 1) appeared as an efficient and selective inhibi- this basis, Ohmline could modulate SK3 channel function by
generating a modification of the biophysical properties of the
membrane and/or by a direct interaction with the SK3 protein
within the lipid bilayer. Its action could be viewed as an illus-
tration of the concept of membrane lipid therapy.34 Ohmline
is also efficient in vivo to prevent the occurrence of SK3+ bone
metastases and to reduce lung metastasis in a murine model
of metastatic breast cancer.17 In this case, Ohmline induced
the dissociation of a protein–protein complex constituted by
two ion channels (Orai 1 and SK3) that are located in the
plasma membrane. This compound is also the first amphiphi-
lic compound that reduces SOCE-dependent cell migration
and modifies anti-EGFR monoclonal antibody effects.18
With the aim to identify new Ohmline analogues, we have
previously modulated the structure of the polar head
group,35,36 integrated a phosphate moiety37 and reported a
fluorescent analogue of Ohmline.38 In the continuity of these
studies, we report herein the synthesis and properties of new
Ohmline analogues that feature
a sulfur-based function
(thioether, sulfoxide, and sulfone) in replacement of one ether
function. We also report the replacement of the methoxy
group by an ethoxy group (compounds 17a and 17b,
Scheme 1). All these structural changes aim to modulate steric
Fig. 1 Representative SK3 whole-cell currents recorded in HEK293T
expressing SK3 channels before and after application of Ohmline ana-
logues. Whole-cell currents were generated using the ramp protocol
from −90 to 90 mV in 500 ms from a constant holding of 0 mV with a
pCa 6. (a) Recording of currents under control conditions (vehicle appli-
cation) and after exposition of compounds 7b (top), 17a (middle) and
17b (bottom) at 10 µM. An addition of 100 nM apamin after exposition to
17b is used to inhibit residual SK currents. (b) Kinetics of modulation of
SK3 currents measured at 0 mV with 7b (top), 17a (middle) and 17b
(bottom). Cells were perfused with the vehicle and the moment when
the vehicle is substituted by the analogue is indicated by a black round
figure.
Scheme 1 (a) Chemical structure of Ohmline. (b) Chemical structure of
the new Ohmline analogues reported herein.
2754 | Org. Biomol. Chem., 2021, 19, 2753–2766
This journal is © The Royal Society of Chemistry 2021

View Article Online
Organic & Biomolecular Chemistry
Paper
hindrance and electronic demand. As reported below, some of emphasize that the precursors 4a and 4b are racemic because
the new compounds are strong activators or strong inhibitors we have previously shown that the configuration at the sn-2
of SK3 channels. One of the activators (7b) is the first amphi- position of the glycerol unit of Ohmline has no effect on the
philic compound that affects the endothelium dependent modulation of SK3 function.29
vasorelaxation.
Then, we prepared analogues of compound 7b in which the
thioether function was oxidized in sulfoxide or sulfone
(Scheme 3). The synthesis of the sulfoxide starts with 4b that
was oxidized with mCPBA (1 eq. at room temperature) as
reported by Ayers et al.51 The sulfoxide 8 was glycosylated with
Results and discussion
The replacement of the glycerol moiety present in Ohmline by lactose-acetate trichloroacetimidate 5. This reaction, which is
catalyzed by BF3, produced a mixture of compounds that was
a thioglycerol unit was the starting point of this study. We
hypothesized that the replacement of the ether group by a impossible to separate. The presence of the sulfoxide that can
thioether would induce geometrical modifications similar to also interact with BF3 is likely the reason for the side reaction
(Scheme 3a). To overcome these difficulties, we investigated a
those observed when we compare diethylether and diethyl-
thioether (diethyl ether features C–O–C angles of 112° and C– second approach for achieving the oxidation of the thioether
O bonds are close to 1.43 Å (ref. 39), whereas diethylthioether after the glycosylation step. Accordingly, the addition of one
equivalent of mCPBA to the thioether 6b yields the expected
features a C–S–C bond angle of 100° and a C–S bond length
close to 1.81 Å (ref. 40)). As a consequence, these modifications sulfoxide 9 as a mixture of diastereoisomers. The deprotection
could influence the interaction of Ohmline analogues with the of the alcohol functions present on the lactose moiety pro-
duces the expected compound 10 (Ohmline-4-SO) in 93% yield
other amphiphiles present in the plasma membranes and,
then, impact SK3 channel function. It is noteworthy that it was (Scheme 3b). The preparation of the sulfone was achieved by
previously evidenced that the incorporation of the thioether using the strategy initially attempted for the sulfoxide
function within lipid chains of ionic liquids,41 bolaamphi-
(Scheme 3c). The oxidation of compound 4b with an excess of
philes42 or cationic amphiphiles, influenced their physico- mCPBA (5 equivalents) for a longer reaction time (4 h) pro-
chemical properties.43,44 The other differences induced by the duced almost quantitatively the sulfone 11. Its glycosylation
with the activated lactose 5 produced compound 12 in 54%
replacement of one ether moiety by a thioether function are:
(i) the difference of polarity due to the respective electro- yield. Finally, the deprotection of alcohol functions yielded
negativity of the oxygen and sulfur atoms; (ii) the size of the compound 13 (Ohmline 4-SO2) in quantitative yield. A fourth
sulfur atom, which is larger than the oxygen atom and more analogue of Ohmline was prepared with the aim to shift the
polarizable; and (iii) the possibility of the thioether derivatives thioether moiety away from the polar head group by one
to be oxidized in sulfoxide45 and subsequently in sulfone.46,47
methylene unit (Scheme 4). The synthesis scheme is comple-
The synthesis of the thioether analogues of Ohmline uses tely different and starts with the synthesis of the vinyl alcohol
thioglycerol as a starting material (see the ESI† for full details
of the synthesis), which is converted into the thioether 4a or
4b in four synthesis steps (overall yield: 40%).48–50 Compound
4a or 4b that possesses either a C12 or C16 lipid chain
(Scheme 2) was then engaged in a glycosylation reaction with
the lactose acetate 5 activated at its anomeric position with a
trichloroacetimidate moiety to produce compound 6a or 6b.
The deprotection of the seven alcohol functions, achieved by
transesterification with potassium carbonate in methanol (for
7a) or with sodium methanolate (for 7b), produced 7a and 7b
in 37% and 100% yields, respectively. It is important to
Scheme 3 (a) Attempts to synthesize compound 10 by a glycosylation
reaction involving the sulfoxide 8. (b) Synthesis of the sulfoxide analogue
of Ohmline 10 (Ohmline-4-SO) by oxidation of the thioether 6b. (c)
Synthesis of the sulfone 13 from 4b. Synthesis of thio-ether analogues
Scheme 2 Synthesis of thio-ether analogues of Ohmline 7a and 7b.
of Ohmline 7a and 7b.
This journal is © The Royal Society of Chemistry 2021
Org. Biomol. Chem., 2021, 19, 2753–2766 | 2755

View Article Online
Paper
Organic & Biomolecular Chemistry
currents using the whole cell configuration of the patch clamp
technique. For these acute tests, we used a concentration of
10 µM for each of the new amphiphilic compounds as already
performed with Ohmline (Ohmline reduces SK3 current by
−73%).29 The results of the patch clamp measurements are
summarized in Table 1. First, we can observe that the replace-
ment of the ether link by a thioether function produces a
strong activator of the SK3 channel (7b) but this effect
depends on the length of the lipid chain. Indeed, the compari-
son of 7a and 7b indicates that the reduction of the length of
the lipid chain from 16 carbon atoms (same as Ohmline) to 12
carbon atoms abolished the effect on SK3. We hypothesize
that a short lipid chain reduces the membrane anchor pro-
perties of the amphiphilic compounds, thus abolishing the
inhibition of SK3.
Scheme 4 Synthesis of the thioether analogue of Ohmline 17a-b.
Then, when the position of the sulfur atom was changed,
the compound retained its activation capacity (comparison of
compounds 7b and 17a). Fig. 1a shows typical SK3 currents
recorded at membrane potentials varying from −90 to +90 mV
for 500 ms under control conditions (vehicle) and after the
application of compounds 7b and 17a. We observed that the
amplitude of outward currents strongly increased after the
application of compounds 7b and 17a. SK3 currents were ana-
lysed at 0 mV to minimize chloride currents and they showed
an increase close to 200% (Table 1). The entire time course of
these experiments is depicted in Fig. 1b and we can observe a
relatively slow effect of these compounds. Indeed, we found
that the maximum of the activation was reached 5 minutes
after the application of either compound 7b or 17a (Fig. 1a
and b), whereas the maximum inhibitory effect of Ohmline is
observed only 2 minutes after application.29 It can be specu-
lated that compounds 7b and 17a are incorporated more
slowly than Ohmline within the plasma membrane. Then, we
investigated the consequence of the presence of a sulfoxide or
sulfone function. Interestingly, the introduction of a sulfoxide
or sulfone function at the same place where the thioether func-
tion was present in compound 7b produced amphiphiles with
reduced or no action on SK3 currents (comparison of com-
pound 7b with 10 and 13). The oxidation of thioether pro-
duced more polar functions (sulfoxide or sulfone) that likely
influenced the position of the amphiphilic compound within
the plasma membrane. Moreover, steric hindrance constraints
and supramolecular interactions are completely different from
a thioether to a sulfoxide or a sulfone.56 Nevertheless, these
results show for the first time that the incorporation of
thioether in place of an ether function produced the first
amphiphilic activators of SK3, whereas the oxidation of this
thioether function produced almost non-active compounds.
We next tested compound 17b that differed from com-
pound 17a in the presence of an ethoxy group at the sn-2 posi-
tion in place of a methoxy group present in 17a. We have pre-
viously shown that the methoxy group at the sn-2 position has
an important effect on the modulation capacity of SK3.
Indeed, LysoPAF which is an ether lipid possessing a phospho-
choline polar head group and an unsubstituted hydroxy group
at the sn-2 position has no effect on SK3.29 The result was
by a regioselective opening of the vinyl epoxide in acidic media
by adopting a reported procedure.52 Then, we applied a photo-
click thiol–ene reaction53 that engaged compounds 14a and
14b with hexadecanethiol. For this reaction, the conditions
were inspired from a procedure we have previously optimized
to prepare other types of amphiphilic compounds.54,55 Then,
the intermediate 15a or 15b was engaged in a glycosylation
reaction with 5 in the presence of BF3–Et2O to produce 16a or
16b in 35% and 41% yield, respectively. The deprotection of
the alcohol function with sodium methanolate (17a) or potass-
ium carbonate (17b) produced compound 17a or 17b in 87%
and 99% yield, respectively.
Then, we tested the capacity of the molecules 7a, 7b, 10, 13,
17a and 17b (Table 1 and Fig. 1) to change the activity of SK3
Table 1 Effect of Ohmline’s analogues on SK3 currents
Structure
SK3 currenta
7a
−9.7% 5.2
(N = 4)
7b
+177% 67
(N = 4)
10
+29% 8.6 (N
= 5)
13
−19.1% 1.8
(N = 5)
17a
17b
+203% 59
(N = 5)
−91.9% 1.8
(N = 4)
a Acute effect of 10 µM Ohmline analogues in SK3 current recorded at
0 mV in HEK293 cells expressing the recombinant SK3 channel.
Results are normalized to control conditions (vehicle) and expressed as
mean S.E.M. N corresponds to the number of cells tested (indepen-
dent experiments).
2756 | Org. Biomol. Chem., 2021, 19, 2753–2766
This journal is © The Royal Society of Chemistry 2021

View Article Online
Organic & Biomolecular Chemistry
Paper
unexpected because the replacement of the methoxy 18% and 15%, respectively, on HCT116). Under similar con-
group by an ethoxy group converts the activator 17a in a new ditions, Ohmline reduces cytosolic calcium by 26% (ref. 18)) as
and original inhibitor of the SK3 channel 17b that reduced expected for an inhibitor of the SK3 channel (Fig. 2).59 On
SK3 currents by −91.9 1.8% (Fig. 1). The kinetics of the inhi- PC3, 7b and 17a increased the calcium concentration by +24%
bition is even quicker than that of Ohmline because the and +70%, respectively. Then, we focused our investigation on
maximum inhibitory effect is reached after 30 seconds (Fig. 1a, 7b, which is a direct thio-analogue of Ohmline. The same
right).
experiment was repeated for 7b after 24 h because this time
We have previously reported that the modulation of SK3 corresponds to the migration assay that was negative for 7b.
channel activity influences the migration of various cancer Surprisingly, we found that after 24 h, SOCE were significantly
cells. The underlying mechanism implies a variation of cytoso- reduced as shown in Fig. 2E and F. This result likely explains
lic calcium concentration through calcium entries from the the absence of the increase of PC3 cell migration after 24 h
extracellular space by calcium channels. More precisely, when (Fig. ESI3-1†). Altogether, these results suggest that 7b should
the activity of SK3 is increased, it leads to plasma membrane
hyperpolarization which increases the driving force for
calcium entries through calcium channels. In contrast, a
reduction of SK3 activity induces a depolarization of the
plasma membrane that subsequently reduces the driving force
for calcium entries. In this respect, SK3 channels control
calcium entries that occur following two mechanisms: 1 – con-
stitutive entries: the SK3 channel forms a complex with the
calcium channel Orai1 that is constitutively open leading to a
constitutive calcium influx.59 2 – Store Operated Calcium
Entries (SOCE): the SK3 channel forms a complex with the
calcium channel Orai1 and TRPC1 that is induced by the reti-
cular STIM1 protein leading to SOCE (a variation of SOCE
reduces or promotes colon and prostate cancer cell migration
in the case of Ohmline and CYPPA, respectively18,57,58). It is
noteworthy that these two types of calcium entries have
different relative importance in cell migration depending on
the cell line. For instance, the MDA-MB-435s cell line features
mostly SK3-dependent constitutive entries, whereas PC3 and
HCT116 feature mostly SK3-dependent calcium entries via
SOCE.
Then, we assess the effect of the strong SK3 activators (7b
and 17a) and inhibitor (17b) on the migration of
MDA-MB-435s. Before this, it must be noted that these com-
pounds were not toxic according to MTT assays (Fig. ESI6-1†).
For the migration assays, we applied the same protocol we
used previously that involved observing the effect of these
compounds (1 µM) on cell migration after 24 h. As shown in
Fig. ESI3-1,† none of these compounds have an effect on the
migration of MDA-MB-435s (under similar conditions,
Ohmline reduces MDA-MB-435s cell migration by 50%).29
Then, we focused on the SK3 activators (7b and 17a) and
tested their effect on the migration of PC3 at 1 µM and 10 µM.
Under these conditions, neither 7b nor 17a had an impact on F340/F380 fluorescence ratio after addition of 2 mM Ca2+ and normal-
the migration of PC3 (Fig. ESI3-1†). The discrepancy between
the strong effect of 7b, 17a, and 17b on SK3 activity and the
absence of an effect on cell migration raises questions on the
Fig. 2 Effect of compounds 7b and 17a at 10 µM on cytosolic calcium
concentration of the HCT116 colon cancer cell line and PC3 prostate
cancer cell line. Mean traces representing Ca2+ signals (indicated by the
ized to control conditions) triggered by adding 2 μM thapsigargin (Tg) to
HCT116 (A) and PC3 cells (C) with or without pre-treatment of the
chosen drugs (shown in various coloured lines, each at 10 μM dose with
different times). (B–D) Scatter plots showing the peak SOCE levels trig-
effect of these compounds on calcium entries.
gered by Tg in control cells and cells treated with a 10 μM dose of each
Accordingly, the cytosolic calcium concentration induced drug after a short period of exposure (13 minutes). Scatter plots showing
the peak SOCE levels triggered by Tg in control cells and cells pre-
treated with each drug at a 10 μM dose over 24 h (E and F) or 1 h (G and
H). Each value (mean SD) was derived from 3–5 individual experiments
and a total of 20–52 wells. The statistical comparison among the values
by SOCE was measured on two SK3+ cell lines (colon HCT116
and prostate PC3 cancer cell lines) at a short time of 13 min
(recording the data at a shorter time was difficult due to experi-
mental constraints). It is observed that 7b and 17a (10 µM)
for all conditions was performed using one-way ANOVA followed by
induced an increase of cytosolic calcium concentration (by Dunnett’s test or Mann–Whitney test. NS, **P < 0.01, ***P < 0.001.
This journal is © The Royal Society of Chemistry 2021
Org. Biomol. Chem., 2021, 19, 2753–2766 | 2757

View Article Online
Paper
Organic & Biomolecular Chemistry
activate cell migration after 13 min application and then
should weakly inhibit cell migration after 24 h application. To
evaluate the effect of 7b on SOCE at an intermediate time, we
recorded calcium entries in PC3 and HCT116 1 h after the
addition of 7b (10 µM). Interestingly, after 1 h application,
compound 7b has no effect on SOCE (Fig. 2G and H). Thus,
this compound time dependently switches from an activator
(13 min) to a non-active compound (1 h) and to a weak inhibi-
tor of SOCE (24 h) suggesting a time-dependent metaboliza-
tion. It must be noted that Ohmline in a similar experiment
and after 24 h reduces SOCE18 suggesting a limited metaboli-
zation over 24 h. The only difference between Ohmline and 7b
is the replacement of an ether function by a thioether func-
tion. It is likely that the metabolization of 7b comes from this
difference. The more likely possibility would be the oxidation
of the thioether function to a sulfoxide (10) and then to a
sulfone (13). 10 and 13 are a weak activator and a weak inhibi-
tor, respectively, of SK3 (Table 1). In conclusion, the effects of
Fig. 3 Relaxant effect of 7b (1 µM) on isolated rat mesenteric artery
rings pre-contracted with U46619 (3 µM) in endothelium-intact (E+) and
endothelium denuded (E−) rings. * p < 0.05 (Student t-test).
Conclusions
10 and 13 on SK3 activity support the hypothesis that 7b could We report herein the synthesis of Ohmline analogues in which
be oxidized over time leading to its conversion to a weak acti- the ether function linked to the lipid chain is replaced by a
vator and then to a weak inhibitor. This variation of the effect thioether. We also prepared one sulfoxide and one sulfone
over time could explain the absence of an effect on cell analogues. The evaluation of these compounds by patch clamp
migration after 24 h. Nevertheless, a dedicated study, which is measurements indicated that only the thioether containing
beyond the scope of this work, would be necessary to confirm compound 7b exhibited a strong activation effect on SK3
this hypothesis.
(+177% 67). The presence of a shorter lipid chain (12 carbon
Then, we studied 17b, which is a strong inhibitor of SK3 atoms instead of 16 atoms in 7b) produced a compound
(Table 1). 17b has no effect on cell migration (1 µM; 24 h) as devoid of any action on SK3, thus emphasizing the importance
shown on Fig. ESI3-1.† It has also no effect on SOCE after a of a long lipid chain. Then, we report the synthesis of one ana-
short time (13 min) as shown in Fig. ESI4-1.† 13 min is likely logue of compound 7b featuring a thioether function shifted
too short for its metabolization. Therefore, this result suggests by one methylene unit away from the polar head group. We
that its effect on SK3 activity and subsequently on calcium found that this compound 17a also features a strong and rapid
entries is compensated by a concomitant action on other part- activation effect (+203% 59) on SK3 channels at 10 µM. If 7b
ners (e.g. ion channels and receptors) that have not been and 17a are strong activators of SK3, unlike Ohmline, they
identified so far. Our conclusion is that 17b has a strong inhi- have no effect on cancer cell migration after 24 h. This behav-
bition effect on the SK3 channel but this effect is likely not iour raised questions about their possible metabolization over
selective leading to the absence of a variation of calcium time. Then, we recorded the SK3-dependent calcium entries at
entries and subsequently to the absence of an effect on cell different times (13 min, 1 h and 24 h) triggered by 7b by using
migration.
fluorescence techniques. We report for the first time that 7b is
Then, we selected compound 7b, because it is the direct a strong and rapid SK3 activator, which is correlated with the
thio-analogue of Ohmline, to assess its effect on vasorelaxa- calcium entry SOCE as shown by cytosolic calcium measure-
tion. Indeed, the activation of the SK3 channel, found to be ments. However, after 24 h a strong decrease of SOCE was
expressed in endothelial cells of small mesenteric arteries, is measured, indicating that 7b has a time dependent effect:
supposed to induce vasorelaxation via the EDHF pathway.3 first, it increases SOCE, then has no effect (1 h) and after 24 h
Moreover, a vasorelaxation effect is usually observed after a it reduces SOCE. Its biochemical transformation, possibly by
short exposure time to a potent drug (maximal effect usually oxidation, is one hypothesis but the non-selective effect via its
observed before 30 min). This feature was compatible with the action on other ion channels (e.g. inhibition of the calcium
properties of 7b, which is a strong activator for a short time. channel) is also hypothesized and will require further investi-
Thus, we tested the capacity of the SK3 activator 7b to gations. The last modulation dealt with the replacement of the
induce vasorelaxation by modulating excitation–contraction methoxy group present in compound 17a by an ethoxy group
coupling of mesenteric smooth muscle. The activator 7b to produce compound 17b. Surprisingly, 17b has a strong
was tested on isolated rat mesenteric arteries precontracted inhibitory effect on SK3 at 10 µM (−91.9% 1.8) pointing out
with the thromboxane analog U46619. As expected, acute the great influence of the substituent present on the central
exposure to the compound 7b at 1 µM induced an endo- oxygen atom. However, 17b has no effect on cancer cell
thelium-dependent vasorelaxant effect (−34%+/−9%, n = 8) migration after 24 h, suggesting non-specific action. Finally,
which reached a maximum value after about 20 min of incu- 7b, which is a strong activator for a short time, is the first
bation (Fig. 3).
amphiphilic compound that induces vasorelaxation via an
2758 | Org. Biomol. Chem., 2021, 19, 2753–2766
This journal is © The Royal Society of Chemistry 2021

View Article Online
Organic & Biomolecular Chemistry
Paper
endothelium-derived hyperpolarization, thus opening a new (s, CH2 fatty chain); 28.9 (s, CH2 fatty chain); 22.7 (s, CH2 fatty
field of investigation for synthetic amphiphilic compounds.
chain); 20.8 (s, OCH3); 14.1 (s, CH3 fatty chain).
1-S-Hexadecyl-2-O-methyl-rac-thioglycer-3-yl-2,3,4,6-tetra-O-
acetyl-β-D-galactopyranosyl-(1–4)-2,3,6-tri-O-acetyl-β-D-gluco-
pyranoside (6b). A solution of lactose trichloroacetimidate 5
(442 mg, 0.57 mmol, 1.0 eq.) and 4b 49 (200 mg, 0.58 mmol,
1.02 eq.) in dry CH2Cl2 (15 mL) is stirred with molecular sieves
4 Å for one hour under N2. At 0 °C, BF3·Et2O (28 µL,
0.23 mmol, 0.4 eq.) is added dropwise and the mixture is
stirred for 24 hours at room temperature. The mixture is
quenched by the addition of water (10 mL). The organic layer
is washed twice with aqueous saturated NaHCO3 solution (2 ×
10 mL) and an aqueous saturated NaCl solution (10 mL). The
organic layer is dried with MgSO4, filtered and concentrated to
give the crude compound 6b. The product is purified by
chromatography on silica gel (eluent: petroleum spirit/ethyl
acetate: 8/2 to 7/3) to give 6b (284 mg) in 51% yield (estimated
Experimental
General information
Commercially available chemicals were used without
additional purification. The products were characterized by H
1
and 13C NMR spectroscopy (Bruker AC 300, Avance DRX 400
and Avance DRX 500 spectrometers) and mass spectrometry
(BrukerAutoflex MALDI TOF-TOF III, LRF200CID and Waters
Synapt XS). For NMR, scalar coupling constants J are given in
hertz. The following abbreviations are used: s for singlet, d for
doublet, t for triplet, q for quadruplet, qt for quintuplet and m
for multiplet. The purity of all final compounds was ≥95% as
determined by TLC (one spot after staining with I2 and/or
H2SO4). The description of the synthesis of compounds 1–4 is
reported in ESI-1.†
1
purity: 95%). Rf (petroleum spirit/ethyl acetate (7 : 3)): 0.28; H
NMR (CDCl3, 399.972): 5.33 (d, 1 H, JHH = 3.2 Hz, H4′); 5.18 (t,
1 H, JHH = 10.0 Hz, H3); 5.09 (dd, 1 H, JHH = 10.0 Hz, JHH = 7.9
Hz, H2′); 4.95–4.88 (m, 2 H, H2 + H3′); 4.53–4.45 (m, 3 H, H1 +
H6a + H1′); 4.11–4.06 (m, 3 H, H6b + H6′a + H6′b); 3.98–3.85 (m,
Synthesis
1-S-Dodecyl-2-O-methyl-rac-thioglycer-3-yl-2,3,4,6-tetra-O-
acetyl-β-D-galactopyranosyl-(1–4)-2,3,6-tri-O-acetyl-β-D-gluco- 2 H, H5′ + Ha CH2 sn-3); 3.78 (t, 1 H, JHH = 10.0 Hz, H4);
pyranoside (6a). A solution of lactose trichloroacetimidate 5 60 3.67–3.54 (m, 2 H, H5 + Hb CH2 sn-3); 3.48–3.42 (m, 1 H, CH
(850 mg, 1.08 mmol, 1.0 eq.) and 4a (300 mg, 1.03 mmol, 0.95 sn-2); 3.39–3.38 (2 s, 3 H, OCH3 two diastereoisomers);
eq.) in dry CH2Cl2 (4 mL) is stirred with molecular sieves 4 Å 2.63–2.59 (m, 2 H, CH2 sn-1); 2.49 (t, 2 H, JHH = JHH = 7.4 Hz,
for one hour under an inert atmosphere. At 0 °C, BF3·Et2O in CH2 α fatty chain); 2.13 (s, 3 H, CH3 acetyl); 2.10 (s, 3 H, CH3
dry CH2Cl2 (2 mL) (196 µL, 1.08 mmol, 1 eq.) is added drop- acetyl); 2.04 (s, 3 H, CH3 acetyl); 2.03 (s, 9 H, CH3 acetyl); 1.95
wise and the mixture is stirred for 4 hours at 0 °C under an (s, 3 H, CH3 acetyl); 1.54 (qt, 2 H, JHH = 7.6 Hz, CH2 β fatty
inert atmosphere. Triethylamine is added at 0 °C and the chain); 1.34–1.22 (m, 26 H, CH2 fatty chain); 0.86 (t, 3 H, JHH
=
mixture is stirred for 15 minutes. The mixture is concentrated 6.8 Hz, CH3 fatty chain); 13C NMR (CDCl3, 125.804): 170.4 (s,
and the compound is diluted in ethyl acetate (20 mL), filtered CvO); 170.2 (s, CvO); 170.1 (s, CvO); 169.8 (s, CvO); 169.7
on Celite and concentrated to give the crude compound 6a, (s, CvO); 169.1 (s, CvO); 101.1 (s, C1′); 101.0–100.9 (C1 two
which is purified on a column of silica gel (eluent: hexane/ diastereoisomers); 79.9–79.7 (CH sn-2, two diastereoisomers);
ethyl acetate: 9/1 to 8/2) to give 6a (270 mg) in 29% yield. Rf 76.2 (s, C4); 72.7 (s, C3); 72.6 (s, C5); 71.6 (s, C2); 71.0 (C3′); 70.6
1
(hexane/acetone (6/4)): 0.8; H NMR (CDCl3, 500.253): 5.33 (d, (C5′); 69.5 (CH2 sn-3); 69.0 (s, C2′); 66.6 (s, C4′); 62.0 (s, C6); 60.8
1 H, H4′); 5.2 (t, 1H, H2′); 5.1 (m, 1 H, H3′); 5.0–4.85 (m, 2H, H1 (s, C6′); 58.1–57.7 (OCH3, two diastereoisomers); 33.2–32.8(CH2
+ H1′); 4.6–4.4 (m, 3H, H6b + H2 + H1′); 4.2–4.0 (m, 3 H, H6b
+
sn-1, two diastereoisomers); 32.6 (CH2 α fatty chain); 31.9 (s,
H6a + H5′); 3.7–3.4 (m, 1 H, H5′ + H6a); 3.7–3.55 (m, 2 H, CH2 CH2 fatty chain); 29.7 (s, CH2 fatty chain); 29.6 (s, CH2 fatty
sn-3); 3.5–3.4 (m, 1 H, CH sn-2); 3.4 (2s, 3 H, OCH3 two dia- chain); 29.5 (s, CH2 fatty chain); 29.4 (s, CH2 fatty chain); 29.3
stereoisomers); 2.7–2.6 (m, 2 H, CH2 sn-1); 2.52 (t, 2H, JHH = 7 (s, CH2 fatty chain); 28.9 (s, CH2 fatty chain); 22.7 (s, CH2 fatty
Hz, CH2 α fatty chain); 2.0–2.2 (m, 21H, CH3 acetyl); 1.55 (qt, 2 chain); 20.9 (s, OCH3); 20.8 (s, OCH3); 20.7 (s, OCH3); 20.5 (s,
H, JHH = 7 Hz, CH2 β fatty chain); 1.35 (m, 2H, CH2 fatty OCH3); 14.1 (s, CH3 fatty chain); mass (MALDI-TOF; matrix
chain); 1.3–1.2 (m, 26 H, CH2 fatty chain); 0.88 (t, 3 H, JHH = 7 dithranol: 100 mg mL−1): m/z calcd for C46H76O19SNa [M +
Hz, CH3 fatty chain); 13C NMR (CDCl3, 125.803): 170.4 (s, Na]+: 987.4599, found 987.4651.
CvO); 170.2 (s, CvO); 170.1 (s, CvO); 169.8 (s, CvO); 169.7
(s, CvO); 169.1 (s, CvO); 101.1 (s, C1′); 101.0–100.9 (C1 two syl-(1–4)-β-D-glucopyranoside (7a
1-S-Dodecyl-2-O-methyl-rac-thioglycer-3-yl-β-D-galactopyrano-
Ohmline-4S). K2CO3
–
diastereoisomers); 79.8 (CH sn-2, two diastereoisomers); 76.2 (182 mg, 1.32 mmol, 6 eq.) is added to a solution of 6a
(s, C4); 72.6 (s, C5); 71.6 (s, C5′); 71.0 (CH2 fatty chain); 70.9 (200 mg, 0.22 mmol, 1.0 eq.) in MeOH (5 mL). The mixture is
(CH2 sn-1); 70.7 (C2); 69.5 (CH2 sn-3); 69.1 (s, C2′); 66.6 (s, C4′); stirred at room temperature for 2 hours. After filtration on
62.0 (s, C6); 60.8 (s, C6′); 58.0–57.7 (OCH3, two diastereo- resin DOWEX, the filtrate is concentrated to give the crude
isomers); 33.2 (CH2 fatty chain); 33.1 (CH2 α fatty chain); 32.8 compound 7a. The crude compound is purified using a
(s, CH2 fatty chain); 32.7 (s, CH2 fatty chain); 31.9 (s, CH2 fatty column of silica gel (CHCl3/MeOH: 80/20) to give 7a (50 mg) in
chain); 32.7 (s, CH2 fatty chain); 31.9 (s, CH2 fatty chain); 29.8 37% yield (estimated purity: 95%). 1H NMR (DMSO-d6,
(s, CH2 fatty chain); 29.7 (s, CH2 fatty chain); 29.6 (s, CH2 fatty 500.252): 5.10 (brs, 2 H, 2 OH); 4.8 (brs, 1 H, OH); 4.66 (brs, 2
chain); 29.5 (s, CH2 fatty chain); 29.4 (s, CH2 fatty chain); 29.2 H, 2 OH); 4.51 (brs, 2 H, 2 OH); 4.21–4.18 (m, 2 H, H1 + H1′);
This journal is © The Royal Society of Chemistry 2021
Org. Biomol. Chem., 2021, 19, 2753–2766 | 2759

View Article Online
Paper
Organic & Biomolecular Chemistry
3.9–3.72 (m, 3 H, H6a + CH2 sn-3); 3.58–3.54 (m, 3 H, H5 + H6b bined organic layers are washed with a saturated aqueous solu-
+ H4′); 3.53–3.42 (m, 5 H, H5′ + H6′a + H6′b + H6b + CH sn-2); tion of NaHCO3 (30 mL) and brine (30 mL). The organic layer
3.42–3.27 (m, 7 H, H3 + H4 + H2′ + H3′ + OCH3); 2.99 (m, 1 H, is dried on MgSO4, filtered and concentrated to give 8 (193 mg;
1
H2); 2.70–2.57 (m, 2 H, CH2 sn-1); 2.60–2.50 (m, 4 H, CH2
α
93% yield) as a white solid. Rf (CH2Cl2/MeOH: 95/5): 0.39; H
fatty chain + DMSO); 1.51–1.48 (m, 1 H, CH2 β fatty chain); NMR (CDCl3, 500.133): 3.80–3.70 (m, 2 H, CH sn-2 + Ha CH2
1.32–1.28 (m, 2 H, CH2 fatty chain); 1.23 (s, 24 H, CH2 fatty sn-3); 3.40–3.35 (m, 1 H, Hb CH2 sn-3); 3.40–3.35 (2 s, 3 H,
chain); 0.87–0.83 (t, 3 H, JHH = 7 Hz, CH3 fatty chain); 13C NMR OCH3 two diastereoisomers); 2.98–2.61 (m, 4 H, CH2 sn-1 +
(DMSO-d6, 125.803): 103.9 (s, C1′); 103.0–102.9 (C1 diastereo- CH2 α fatty chain); 1.68 (qt, 2 H, JHH = 7.0 Hz, CH2 β fatty
isomers); 80.8 (C4) 79.6–79.5 (CH sn-2, diastereoisomers); 75.5 chain); 1.38–1.17 (m, 26 H, CH2 fatty chain); 0.80 (t, 3 H, JHH
=
(s, C5′); 75.0 (s, C3); 74.9 (s, C5); 73.3 (s, C2); 73.1 (s, C3′); 70.6 7.0 Hz, CH3 fatty chain); 13C NMR (CDCl3, 75.475): 76.1–75.8
(C2′); 69.5–69.3 (CH2 sn-3, diastereoisomers); 68.1 (C4′); (CH sn-2, two diastereoisomers); 62.3–62.0 (CH2 sn-3, two dia-
60.5–60.4 (2s, C6 + C6′); 57.0–56.9 (OCH3, diastereoisomers); stereoisomers); 57.9–57.1 (OCH3, two diastereoisomers);
32.4–32.2 (CH2 sn-1, diastereoisomers); 32.1 (CH2 α fatty 55.7–52.6 (CH2 sn-1, two diastereoisomers); 53.2 (CH2 α fatty
chain); 31.4 (s, CH2 fatty chain); 29.3 (s, CH2 fatty chain); 29.1 chain); 31.9 (CH2 fatty chain); 29.7 (CH2 fatty chain); 29.5 (CH2
(s, CH2 fatty chain); 28.8 (s, CH2 fatty chain); 28.7 (s, CH2 fatty fatty chain); 29.3 (CH2 fatty chain); 29.2 (CH2 fatty chain); 28.8
chain); 28.3 (s, CH2 fatty chain); 22.2 (s, CH2 fatty chain); 14.0 (CH2 fatty chain); 22.7 (CH2 fatty chain); 14.1 (CH3 fatty chain);
(s, CH3 fatty chain); mass (MALDI-TOF; matrix dithranol: mass (ESI QqTOF): m/z calcd for C20H42O3SNa [M + Na]+:
100 mg mL−1): m/z calcd for C28H54O12SK [M + K]+: 653.297, 385.2752, found 385.2759.
found: 653.412.
1-S-Hexadecylsulfinyl-2-O-methyl-rac-thioglycer-3-yl-2,3,4,6-
1-S-Hexadecyl-2-O-methyl-rac-thioglycer-3-yl-β-D-galactopyr-
tetra-O-acetyl-β-D-galactopyranosyl-(1–4)-2,3,6-tri-O-acetyl-β-D-
anosyl-(1–4)-β-D-glucopyranoside (7b – Ohmline-4S). MeONa glucopyranoside (9). To a stirred solution of 6b (150 mg,
in methanol (2.8 mL, 0.02 mmol, 0.5 eq.) is added to a solu- 0.16 mmol, 1.0 eq.) in dry CH2Cl2 (5 mL) is added very slowly
tion of 6b (40 mg, 4.14 × 10−5 mol, 1.0 eq.) in MeOH (5 mL). mCPBA (77%) (34 mg, 0.16 mmol, 1.0 eq.) in dry CH2Cl2
The mixture is stirred at room temperature for 15 hours. (5 mL) at 0 °C. The mixture is stirred at 0 °C and the reaction
Amberlyst IR-120 (H+) is added and the mixture is stirred for is followed by TLC (petroleum ether/ethyl acetate: 3/7). The
30 minutes at room temperature. The reaction mixture is mixture is quenched with a saturated aqueous solution of
warmed (reflux), quickly filtered and concentrated to give the NaHSO3 (10 mL). The aqueous layer is extracted three times
crude compound 7b (28 mg) in a quantitative yield (estimated with CH2Cl2 (3 × 20 mL) and the combined organic layers are
purity: >95%). 1H NMR (DMSO-d6, 500.133): 5.12–5.10 (m, 2 H, washed with a saturated aqueous solution of NaHCO3 (30 mL)
2 OH); 4.80 (brs, 1 H, OH); 4.68–65 (m, 2 H, 2 OH); 4.52–4.50 and brine (30 mL). The organic layer is dried upon MgSO4, fil-
(m, 2 H, 2 OH); 4.22–4.20 (m, 2 H, H1 + H1′); 3.83–3.72 (m, 3 H, tered and concentrated to give 9 (148 mg; 97% yield; estimated
1
H6a + CH2 sn-3); 3.62–3.60 (m, 3 H, H5 + H6b + H4′); 3.53–3.43 purity: >95%). Rf (petroleum spirit/ethyl acetate: 7/3): 0.10; H
(m, 4 H, H5′ + H6′a + H6′b + CH sn-2); 3.36–3.27 (m, 7 H, H3 + NMR (CDCl3, 399.972): 5.34 (d, 1 H, JHH = 3.2 Hz, H4′); 5.19 (td,
H4 + H2′ + H3′ + OCH3); 3.00–2.98 (m, 1 H, H2); 2.70–2.64 (m, 1 1 H, JHH = 9.8 Hz, JHH = 2.8 Hz, H3); 5.10 (dd, 1 H, JHH = 10.4
H, Ha CH2 sn-1); 2.60–2.50 (m, 3 H, CH2 α fatty chain + Hb CH2 Hz, JHH = 8.0 Hz, H2′); 4.96–4.90 (m, 2 H, H2 + H3′); 4.53–4.46
sn-1); 1.52–1.47 (m, 2 H, β CH2 fatty chain); 1.32–1.23 (m, 26 (m, 3 H, H1 + H6a + H1′); 4.12–4.06 (m, 3 H, H6b + H6′a + H6′b);
H, CH2 fatty chain); 0.85 (t, 3 H, JHH = 6.8 Hz, CH3 fatty chain); 3.98–3.61 (m, 6 H, H4 + H5 + H5′ + CH2 sn-3 + CH sn-2);
13C NMR (DMSO-d6, 125.803): 103.9 (s, C1′); 103.0–102.9 (C1 3.46–3.38 (4 s, 3 H, OCH3 diastereoisomers); 2.98–2.61 (m, 4
diastereoisomers); 80.8 (C4) 79.6–79.5 (CH sn-2, diastereo- H, CH2 sn-1 + CH2 α fatty chain); 2.14 (s, 3 H, CH3 acetyl); 2.11
isomers); 75.5 (s, C5′); 75.0 (s, C3); 74.9 (s, C5); 73.3 (s, C2); 73.1 (s, 3 H, CH3 acetyl); 2.05 (s, 3 H, CH3 acetyl); 2.03 (s, 3 H, CH3
(s, C3′); 70.6 (C2′); 69.5–69.3 (CH2 sn-3, diastereoisomers); 68.1 acetyl); 2.02 (s, 6 H, 2 CH3 acetyl); 1.95 (s, 3 H, CH3 acetyl);
(C4′); 60.5–60.4 (2s, C6 + C6′); 57.0–56.9 (OCH3, diastereo- 1.75–1.71 (m, 2 H, CH2 fatty chain); 1.32–1.23 (m, 26 H, CH2
isomers); 32.4–32.2 (CH2 sn-1, diastereoisomers); 32.1 (CH2 α fatty chain); 0.87 (t, 3 H, JHH = 6.8 Hz, CH3 fatty chain); 13C
fatty chain); 31.4 (s, CH2 fatty chain); 29.3 (s, CH2 fatty chain); NMR (CDCl3, 75.474): 170.3 (s, CvO); 170.1 (s, CvO); 170.0 (s,
29.1 (s, CH2 fatty chain); 28.8 (s, CH2 fatty chain); 28.7 (s, CH2 CvO); 169.7 (s, CvO); 169.0 (s, CvO); 101.1 (s, C1′);
fatty chain); 28.3 (s, CH2 fatty chain); 22.2 (s, CH2 fatty chain); 101.0–100.6 (C1 diastereoisomers); 76.1 (s, C4); 74.0 (m, CH sn-
14.0 (s, CH3 fatty chain); mass (ESI QqTOF) m/z calcd for 2); 72.8 (s, C3); 72.6 (s, C5); 71.6(s, C2); 71.0 (C3′); 70.7 (C5′);
C32H62O12SNa [M + Na]+: 693.3860, found 693.3857.
70.3 (m, CH2 sn-3); 69.1 (s, C2′); 66.6 (s, C4′); 61.8 (s, C6); 60.8
3-(Hexadecylsulfinyl)-2-methoxypropan-1-ol (8). To a stirred (s, C6′); 58.5 (m, OCH3, diastereoisomers); 55.9 (m, CH2 sn-1,
solution of 4b 49 (198 mg, 0.571 mmol, 1.0 eq.) in dry CH2Cl2 diastereoisomers); 52.8 (m, CH2 α fatty chain); 31.9 (s, CH2
(5 mL) is added very slowly mCPBA (77%) (128 mg, fatty chain); 29.7 (s, CH2 fatty chain); 29.5 (s, CH2 fatty chain);
0.571 mmol, 1.0 eq.) in dry CH2Cl2 (5 mL) at 0 °C. The mixture 29.4 (s, CH2 fatty chain); 29.2 (s, CH2 fatty chain); 28.8 (s, CH2
is stirred at 0 °C and the reaction is followed by TLC (CH2Cl2/ fatty chain); 22.7 (s, CH2 fatty chain); 20.9 (s, OCH3); 20.8 (s,
MeOH: 95/5). The mixture is quenched with a saturated OCH3); 20.6 (s, OCH3); 20.5 (s, OCH3); 14.1 (s, CH3 fatty chain);
aqueous solution of NaHSO3 (10 mL). The aqueous layer is mass (ESI QqTOF): m/z calcd for C46H77O20S [M + H]+:
extracted three times with CH2Cl2 (3 × 20 mL) and the com- 981.4729, found 981.4731.
2760 | Org. Biomol. Chem., 2021, 19, 2753–2766
This journal is © The Royal Society of Chemistry 2021

View Article Online
Organic & Biomolecular Chemistry
Paper
1-S-Hexadecylsulfinyl-2-O-methyl-rac-thioglycer-3-yl-β-D-galac-
date 5 60 (202 mg, 0.26 mmol, 1.0 eq.) and 11 (100 mg,
topyranosyl-(1–4)-β-D-glucopyranoside (10
–
Ohmline-4-SO). 0.26 mmol, 1.02 eq.) in dry CH2Cl2 (5 mL) is stirred with mole-
K2CO3 (2.8 mg, 0.02 mmol, 0.5 eq.) is added to a solution of 9 cular sieves 4 Å for one hour under an inert atmosphere. At
(40 mg, 0.04 mmol, 1.0 eq.) in MeOH (10 mL). The mixture is 0 °C, BF3·Et2O (13 µL, 0.10 mmol, 0.4 eq.) is added dropwise
stirred at room temperature overnight. Amberlyst IR-120 (H+) and the mixture is stirred for 17 hours at room temperature
is added and the mixture is stirred for 30 minutes at room under an inert atmosphere. The mixture is quenched by the
temperature. The reaction mixture is warmed to reflux, quickly addition of water (3 mL). The organic layer is washed twice
filtered and concentrated to give 10 in a quantitative yield with aqueous saturated NaHCO3 solution (2 × 3 mL) and an
1
(28 mg); H NMR (DMSO-d6, 500.133): 5.20–5.13 (m, 1 H, OH); aqueous saturated NaCl solution (3 mL). The organic layer is
5.07 (brs, 1 H, OH); 4.76 (brs, 1 H, OH); 4.66–4.62 (m, 2 H, 2 dried with MgSO4, filtered and concentrated. The crude com-
OH); 4.57–4.49 (m, 2 H, 2 OH); 4.25–4.18 (m, 2 H, H1 + H1′); pound is purified by chromatography on silica gel (eluent: pet-
3.94–3.25 (m, 17 H, H3 + H4 + H5 + H6a + H6b + H2′ + H3′ + H4′
+
roleum spirit/ethyl acetate: 1/1) to give 12 (140 mg; 54% yield)
H5′ + H6′a + H6′b + CH sn-2 + CH2 sn-3 + OCH3); 3.09–3.02 (m, 1 as a colorless oil. Rf (petroleum spirit/ethyl acetate: 1/1): 0.35;
H, H2); 3.09–2.80 (m, 2 H, CH2 sn-1); 2.77–2.49 (m, 2 H, CH2 α 1H NMR (CDCl3, 399.972): 5.34 (d, 1 H, JHH = 3.2 Hz, H4′); 5.19
fatty chain); 1.65–1.60 (m, 2 H, CH2 β fatty chain); 1.39–1.32 (t, 1 H, JHH = 9.8 Hz, H3); 5.12 (dd, 1 H, JHH = 9.8 Hz, JHH = 8.0
(m, 2 H, CH2 fatty chain); 1.32–1.20 (m, 26 H, CH2 fatty chain); Hz, H2′); 4.96–4.87 (m, 2 H, H2 + H3′); 4.52–4.47 (m, 3 H, H1 +
0.86 (t, 3 H, JHH = 6.8 Hz, CH3 fatty chain); 13C NMR (DMSO- H6a + H1′); 4.12–4.06 (m, 3 H, H6b + H6′a + H6′b); 3.93–3.87 (m,
d6, 125.771): 103.8 (s, C1′); 103.0–102.8 (C1 diastereoisomers); 3 H, H5′ + CH sn-2 + Ha CH2 sn-3); 3.78 (t, 1 H, JHH = 9.8 Hz,
80.7–80.6 (C4 diastereoisomers); 75.5 (s, C5′); 74.9 (brs, C3
+
H4); 3.70–3.59 (m, 2 H, H5 + Hb CH2 sn-3); 3.41–3.40 (2 s, 3 H,
C5); 74.7–73.7 (m, CH sn-2 diastereoisomers); 73.2 (s, C2); 73.1 OCH3 two diastereoisomers); 3.28–3.12 (m, 1 H, Ha CH2 sn-1);
(s, C3′); 70.5 (C2′); 69.6–69.2 (CH2 sn-3, diastereoisomers); 68.1 3.10–2.90 (m, 3 H, Hb CH2 sn-1 + CH2 α fatty chain); 2.16 (s, 3
(C4′); 60.5–60.4 (2s, C6 + C6′); 57.3–56.6 (m, OCH3 diastereo- H, CH3 acetyl); 2.14 (s, 3 H, CH3 acetyl); 2.12 (s, 3 H, CH3
isomers); 54.5–51.8 (m, CH2 sn-1 diastereoisomers); 52.5–51.3 acetyl); 2.05 (s, 3 H, CH3 acetyl); 2.03 (s, 6 H, 2 CH3 acetyl);
(m, CH2 α fatty chain diastereoisomers); 31.3 (s, CH2 fatty 1.95 (s, 3 H, CH3 acetyl); 1.83–1.80 (m, 2 H, CH2 fatty chain);
chain); 29.0 (s, CH2 fatty chain); 28.8 (s, CH2 fatty chain); 28.7 1.42–1.24 (m, 26 H, CH2 fatty chain); 0.87 (t, 3 H, JHH = 6.6 Hz,
(s, CH2 fatty chain); 28.1 (s, CH2 fatty chain); 22.1 (s, CH2 fatty CH3 fatty chain); 13C NMR (CDCl3, 75.474): 170.3 (s, CvO);
chain); 13.9 (s, CH3 fatty chain); mass (ESI QqTOF): m/z calcd 170.1 (s, CvO); 170.0 (s, CvO); 169.7 (s, CvO); 169.6 (s,
for C32H63O13S [M + H]+: 687.3989, found 687.3982.
CvO); 169.1 (s, CvO); 101.1 (s, C1′); 100.8–100.4 (C1 two dia-
stereoisomers); 76.1 (s, C4); 75.2–74.9 (CH sn-2, two diastereo-
3-(Hexadecylsulfonyl)-2-methoxypropan-1-ol (11).49 To
a
stirred solution of 4b 49 (200 mg, 0.577 mmol, 1.0 eq.) in dry isomers); 72.8 (s, C3); 72.6 (s, C5); 71.5(s, C2); 71.0 (C3′); 70.7
CH2Cl2 (10 mL) is added mCPBA (500 mg, 2.885 mmol, 5.0 (C5′); 69.1 (s, C2′); 68.5 (CH2 sn-3); 66.6 (s, C4′); 61.8 (s, C6); 60.8
eq.) dissolved in a few mL of dry CH2Cl2 at 0 °C. The mixture (s, C6′); 58.0–57.7 (OCH3, two diastereoisomers); 55.0 (CH2 sn-
is stirred at room temperature for 4 hours. The mixture is 1, two diastereoisomers); 54.8 (CH2 α fatty chain); 31.9 (s, CH2
quenched with a saturated aqueous solution of NaHSO3 fatty chain); 29.7 (s, CH2 fatty chain); 29.1 (s, CH2 fatty chain);
(10 mL). The aqueous layer is extracted three times with 28.5 (s, CH2 fatty chain); 22.7 (s, CH2 fatty chain); 21.8 (s, CH2
CH2Cl2 (3 × 20 mL) and the combined organic layers are fatty chain); 20.8 (s, OCH3); 20.6 (s, OCH3); 20.5 (s, OCH3); 14.1
washed with a saturated aqueous solution of NaHCO3 (20 mL) (s, CH3 fatty chain); mass (MALDI-TOF; matrix dithranol:
and brine (20 mL). The organic layer is dried with MgSO4, fil- 100 mg mL−1): m/z calcd for C46H76O21SNa [M + Na]+:
tered and concentrated to give 11 (209 mg; 98% yield). Rf 1019.4497, found 1019.4607. Mass (ESI QqTOF): m/z calcd for
(CH2Cl2/MeOH: 95/5): 0.66; 1H NMR (CDCl3, 399.992): C46H76O21SNa [M + Na]+: 1019.4497, found 1019.4493.
3.85–3.82 (m, 2 H, CH sn-2 + Ha CH2 sn-3); 3.63–3.59 (m, 1 H,
Hb CH2 sn-3); 3.43 (s, 3 H, OCH3); 3.37 (dd, 1 H, JHH = 14.8 Hz, topyranosyl-(1–4)-β-D-glucopyranoside (13
1-S-Hexadecylsulfonyl-2-O-methyl-rac-thioglycer-3-yl-β-D-galac-
Ohmline-4-SO2).
–
JHH = 8.0 Hz, Ha CH2 sn-1); 3.10–3.00 (m, 3 H, Hb CH2 sn-1 + NaOMe (0.5 M solution in MeOH, 48 µL, 1.3 mg, 0.03 mmol,
CH2 α fatty chain); 1.82 (qt, 2 H, JHH = 6.6 Hz, CH2 β fatty 0.3 eq.) is added to a solution of 12 (80 mg, 0.08 mmol, 1.0
chain); 1.41 (qt, 2 H, JHH = 6.8 Hz, CH2 fatty chain); 1.36–1.23 eq.) in MeOH (10 mL). The mixture is stirred at room tempera-
(m, 24 H, CH2 fatty chain); 0.87 (t, 3 H, JHH = 6.6 Hz, CH3 fatty ture for 5 hours. Amberlyst IR-120 (H+) is added and the
chain); 13C NMR (CDCl3, 75.475): 76.4 (CH sn-2); 62.0 (CH2 sn- mixture is stirred for 30 minutes at room temperature. The
3); 57.6 (OCH3); 55.0 (CH2 sn-1); 54.6 (CH2 α fatty chain); 32.0 reaction mixture is warmed to reflux, quickly filtered and con-
(CH2 fatty chain); 29.7 (CH2 fatty chain); 29.6 (CH2 fatty chain); centrated to give the compound 13 (53 mg) in a quantitative
1
29.4 (CH2 fatty chain); 29.3 (CH2 fatty chain); 29.1 (CH2 fatty yield. H NMR (DMSO-d6, 399.972): 5.28–5.12 (m, 2 H, 2 OH);
chain); 28.5 (CH2 fatty chain); 22.7 (CH2 fatty chain); 21.9 (CH2 4.85 (brs, 1 H, OH); 4.71–65 (m, 2 H, 2 OH); 4.60–4.52 (m, 2 H,
fatty chain); 14.2 (CH3 fatty chain); mass (ESI QqTOF): m/z 2 OH); 4.25–4.18 (m, 2 H, H1 + H1′); 3.89–3.74 (m, 3 H, H6a
+
calcd for C20H42O4SNa [M + Na]+: 401.2702, found 401.2701.
CH sn-2 + Ha CH2 sn-3); 3.61–3.30 (m, 16 H, H3 + H4 + H5 +
H6b + H2′ + H3′ + H4′ + H5′ + H6′a + H6′b + Hb CH2 sn-3 + OCH3 +
1-S-Hexadecylsulfonyl-2-O-methyl-rac-thioglycer-3-yl-2,3,4,6-
tetra-O-acetyl-β-D-galactopyranosyl-(1–4)-2,3,6-tri-O-acetyl-β-D- CH2 sn-1); 3.08–3.02 (m, 3 H, H2 + CH2 α fatty chain); 1.68 (qt,
glucopyranoside (12). A solution of lactose trichloroacetimi- 2 H, JHH = 7.6 Hz, CH2 β fatty chain); 1.36–1.23 (m, 26 H, CH2
This journal is © The Royal Society of Chemistry 2021
Org. Biomol. Chem., 2021, 19, 2753–2766 | 2761

View Article Online
Paper
Organic & Biomolecular Chemistry
fatty chain); 0.85 (t, 3 H, JHH = 6.8 Hz, CH3 fatty chain); 13C fatty chain); 22.7 (CH2 fatty chain); 14.1 (CH3 fatty chain);
NMR (DMSO-d6, 125.803): 103.9 (s, C1′); 103.0–102.8 (C1 two mass (ESI QqTOF) m/z calcd for C21H44O2SNa [M + Na]+:
diastereoisomers); 80.7 (C4) 75.5 (CH sn-2); 75.0 (brs, C5′ + C3 + 383.2960, found 383.2959.
C5); 73.2 (s, C2); 73.1 (s, C3′); 70.6 (C2′); 68.9–68.4 (CH2 sn-3,
4-(Hexadecylthio)-2-ethoxybutan-1-ol (15b). Compound 14b
two diastereoisomers); 68.1 (C4′); 60.5–60.4 (2s, C6 + C6′); 57.1 (400 mg, 3.44 mmol, 1.0 eq.) and 1-hexadecanethiol (3.2 mL,
(OCH3); 54.1 (CH2 sn-1); 53.7 (CH2 α fatty chain); 31.3 (s, CH2 10.3 mmol, 3.0 eq.) are mixed under nitrogen. DMPA (20 mg,
fatty chain); 29.0 (s, CH2 fatty chain); 28.7 (s, CH2 fatty chain); 10% wt) is added and the solution is placed under UV for
28.5 (s, CH2 fatty chain); 27.7 (s, CH2 fatty chain); 22.1 (s, CH2 24 hours at room temperature. The product is purified by
fatty chain); 21.1 (s, CH2 fatty chain); 14.0 (s, CH3 fatty chain); chromatography on silica gel (eluent: pentane/ethyl acetate: 8/
mass (ESI QqTOF): m/z calcd for C32H62O14SNa [M + Na]+: 2) to give 15b (460 mg; 36% yield) as a colorless liquid. Rf
725.3758, found 725.3755.
(pentane/ethyl acetate: 8/2): 0.41; 1H NMR (CDCl3, 399.922):
2-Methoxy-but-3-en-1-ol (14a). To a stirred solution of buta- 3.67–3.62 (m, 1H, CHa sn3); 3.57–3.52 (m, 2H, CH2 –OEt);
diene oxide (575 µL, 7.13 mmol, 1.0 eq.) in methanol (7 mL) is 3.48–3.43 (m, 2 H, CHb sn3 + CH sn2); 2.55 (t, 2 H, JHH = 7.2
added Al(OTf)3 (3.4 mg, 0.007 mmol, 0.001 eq.) in methanol Hz, CH2 α S); 2.48 (t, 2 H, JHH = 7.2 Hz, CH2 α S fatty chain);
(3 mL) dropwise. The mixture is stirred at 100 °C for 1 hour 2.13 (s, 1H, OH); 1.83–1.78 (m, 1H, CHa sn1); 1.73–1.69 (m,
(oil bath temperature). The solvent is removed carefully under 1H, CHb sn1); 1.55–1.49 (m, 2H, CH2 βS fatty chain); 1.34–1.31
vacuum (T = 25 °C, P = 160 mbar) to obtain quantitatively the (m, 2H; CH2 γS fatty chain), 1.21 (s, 23H, CH2 fatty chain); 1.17
compound 14a as a colorless liquid (728 mg). Rf (petroleum (t, 3H, CH3 –OEt); 0.84 (t, 3H, CH3 fatty chain); 13C NMR
spirit/ethyl acetate: 1/1): 0.65; 1H NMR (CDCl3, 399.972): (CDCl3, 75.474): 78.5 (CH sn2); 65.3 (CH2 –OEt); 63.97 (CH2
5.65–5.57 (m, 1 H, H3); 5.28–5.22 (m, 2 H, CH2 H4); 3.67–3.63 sn3); 32.3; 32.0; 31.2; 29.8; 29.6; 29.5; 29.4; 29.0; 28.1 (CH2 αS +
(m, 1 H, H2); 4.51–3.45 (m, 2 H, CH2 H1); 3.28 (s, 3 H, OCH3); CH2 fatty chain); 22.8 (CH2 sn1) 15.7 (CH3 –OEt); 14.2 (CH3
13C NMR (CDCl3, 75.5): 134.9 (CH alkene); 119.4 (CH2 alkene); fatty chain); mass (Maldi TOF, Matrix HCCA): m/z calcd for
83.5 (CH); 65.3 (CH2); 56.6 (OCH3); mass (MALDI-TOF; matrix C22H46O2SNa [M + Na]+: 397.311, found 397.620.
DCTB 10 mg mL−1): m/z calcd for C5H10O2 [M]+: 102.0681,
found 102.0659.
1-O-[-4-(Hexadecylthio)-2-methoxybutyl]-2,3,4,6-tetra-O-acetyl-
β-D-galactopyranosyl-(1–4)-2,3,6-tri-O-acetyl-β-D-glucopyranoside
2-Ethoxy-but-3-en-1-ol (14b). To a stirred solution of buta- (16a). A solution of lactose trichloroacetimidate 5 60 (425 mg,
diene oxide (575 µL, 7.13 mmol, 1.0 eq.) in ethanol (7 mL) is 0.54 mmol, 1.0 eq.) and 15a (200 mg, 0.55 mmol, 1.02 eq.) in
added Al(OTf)3 (3.4 mg, 0.007 mmol, 0.001 eq.) in ethanol dry CH2Cl2 (10 mL) is stirred with molecular sieves 4 Å for one
(3 mL) dropwise. The mixture is stirred at 100 °C for 1 hour hour under an inert atmosphere. At 0 °C, BF3·Et2O (27 µL,
(heater block insert). The solvent is removed carefully under 0.216 mmol, 0.4 eq.) is added dropwise and the mixture is
vacuum (T = 25 °C, P = 6 mbar) to obtain the compound 14b stirred for 15 hours at room temperature under an inert atmo-
(1.44 g; 87% yield) as a colorless liquid. Rf (petroleum spirit/ sphere. The mixture is quenched by the addition of water
ethyl acetate: 1/1): 0.67; 1H NMR (CDCl3, 399.920): 5.73–5.65 (3 mL). The organic layer is washed twice with aqueous satu-
(m, 1H, H3); 5.34–5.25 (m, 2H, CH2 H4); 3.85–3.81 (m, 1 H, H2); rated NaHCO3 solution (2 × 3 mL) and an aqueous saturated
3.69–3.49 (m, 3 H, CH2 H1 + CH2OEt); 3.43–3.38 (m, 1 H, CH2 NaCl solution (3 mL). The organic layer was dried on MgSO4,
H1′); 1.22–1.19 (t, 3H, JHH = 7.2 Hz CH3OEt); 13C NMR (CDCl3, filtered and concentrated to give 16a which was purified by
75.5): 135.3 (CH alkene); 117.8 (CH2 alkene); 81.4 (CH); 64.9 chromatography on silica gel (eluent: petroleum spirit/ethyl
(CH2OEt); 63.8 (CH2); 14.8 (CH3OEt).
acetate: 8/2 to 6/4) to give 16a (180 mg) in 35% yield. Rf (pet-
4-(Hexadecylthio)-2-methoxybutan-1-ol (15a). Compound roleum spirit/ethyl acetate: 6/4): 0.65; 1H NMR (CDCl3,
14a (200 mg, 1.96 mmol, 1.0 eq.) and 1-hexadecanethiol 399.992): 5.34 (d, 1 H, JHH = 2.8 Hz, H4′); 5.19 (t, 1 H, JHH = 9.4
(1.81 mL, 5.87 mmol, 3.0 eq.) are mixed under argon. DMPA Hz, H3); 5.11 (dd, 1 H, JHH = 10.2 Hz, JHH = 7.8 Hz, H2′);
(20 mg, 10% wt) is added and the solution is placed under UV 4.95–4.87 (m, 2 H, H2 + H3′); 4.52–4.45 (m, 3 H, H1 + H6a
+
for 24 hours at room temperature. The product is purified by H1′); 4.11–4.03 (m, 3 H, H6b + H6′a + H6′b); 3.87–3.80 (m, 2 H,
chromatography on silica gel (eluent: petroleum spirit/ethyl H5′ + Ha CH2 HA); 3.80 (t, 1 H, JHH = 9.4 Hz, H4); 3.59–3.56 (m,
acetate: 8/2) to give pure 15a with 32% yield (218 mg). Rf (pet- 1 H, H5); 3.53–3.39 (m, 2 H, CH HC + Hb CH2 HA); 3.36–3.35 (2
roleum spirit/ethyl acetate: 8/2): 0.50; 1H NMR (CDCl3, s, 3 H, OCH3 diastereoisomers); 2.55–2.52 (m, 2 H, CH2 HD);
399.992): 3.73 (ABX, part A, dd, 1 H, JHH = 11.6 Hz, JHH = 3.6 2.48 (t, 2 H, JHH = 7.4 Hz, CH2 α fatty chain); 2.13 (s, 3 H, CH3
Hz, Ha CH2 HA); 3.51 (ABX, part B, dd, 1 H, JHH = 11.6 Hz, JHH acetyl); 2.10 (s, 3 H, CH3 acetyl); 2.04 (s, 3 H, CH3 acetyl); 2.02
= 5.2 Hz, Hb CH2 HA); 3.40 (s, 4 H, HB + OCH3); 2.57 (t, 2 H, (s, 9 H, 3 CH3 acetyl); 1.94 (s, 3 H, CH3 acetyl); 1.72–1.70 (m, 2
JHH = 7.0 Hz, CH2 α fatty chain); 2.51 (t, 2 H, JHH = 7.2 Hz, CH2 H, CH2 HC); 1.56 (qt, 2 H, JHH = 7.4 Hz, CH2 β fatty chain);
HD); 1.88 (h, 1 H, JHH = 6.8 Hz, Ha CH2 HC); 1.77 (h, 1 H, JHH
=
1.34–1.19 (m, 26 H, CH2 fatty chain); 0.87 (t, 3 H, JHH = 6.8 Hz,
6.8 Hz, Hb CH2 HC); 1.59 (qt, 2 H, JHH = 7.4 Hz, CH2 β fatty CH3 fatty chain); 13C NMR (CDCl3, 75.474): 170.4 (s, CvO);
chain); 1.35–1.119 (m, 26 H, CH2 fatty chain); 0.88 (t, 3 H, JHH 170.2 (s, CvO); 170.1 (s, CvO); 169.9 (s, CvO); 169.7 (s,
= 6.6 Hz, CH3 fatty chain); 13C NMR (CDCl3, 75.474): 80.2 (CB); CvO); 169.2 (s, CvO); 101.2 (s, C1′); 100.8 (s, C1); 78.6–78.4
63.4 (CA); 57.3 (OCH3); 32.2 (CH2 α fatty chain); 31.9 (CH2 fatty (CB); 76.4 (s, C4); 72.8 (s, C3); 72.7 (s, C5); 71.7(s, C2); 71.1 (C3′);
chain); 30.6 (CC); 29.7; 29.5; 29.3; 29.2; 28.9; 27.9 (CD + CH2 70.8 (C5′); 72.2–70.3 (CA); 69.2 (s, C2′); 66.7 (s, C4′); 62.1 (s, C6);
2762 | Org. Biomol. Chem., 2021, 19, 2753–2766
This journal is © The Royal Society of Chemistry 2021

View Article Online
Organic & Biomolecular Chemistry
Paper
60.9 (s, C6′); 58.3–57.7 (m, OCH3, diastereoisomers); 32.3 (m, 3.78–3.75 (m, 2 H, H6a + Ha CH2 HA); 3.61–3.29 (m, 15 H, H3 +
CH2 α fatty chain); 32.0 (s, CH2 fatty chain); 31.6 (s, CC); 29.8; H4 + H5 + H6b + H2′ + H3′ + H4′ + H5′ + H6′a + H6′b + Hb CH2 HA
29.5; 29.4; 29.0; 27.9(CD + CH2 fatty chain); 22.8 (s, CH2 fatty + CH HB + OCH3); 3.02–3.00 (m, 1 H, H2); 2.54–2.44 (m, 4 H,
chain); 21.0 (s, OCH3); 20.9 (s, OCH3); 20.7 (s, OCH3); 20.6 (s, CH2 HD + CH2 α fatty chain); 1.72–1.62 (m, 2 H, CH2 HC); 1.52
OCH3); 14.2 (s, CH3 fatty chain); mass (MALDI TOF; matrix (qt, 2 H, JHH = 6.6 Hz, CH2 β fatty chain); 1.36–1.23 (m, 26 H,
dithranol: 100 mg mL−1): m/z calcd for C47H78O19S [M + Na]+: CH2 fatty chain); 0.87 (t, 3 H, JHH = 6.8 Hz, CH3 fatty chain);
1001.4756, found 1001.4678.
1-O-[-4-(Hexadecylthio)-2-ethoxybutyl]-2,3,4,6-tetra-O-acetyl-
β-D-galactopyranosyl-(1–4)-2,3,6-tri-O-acetyl-β-D-glucopyrano-
13C NMR (DMSO-d6, 75.474): 103.9 (s, C1′); 102.9–102.7 (C1 dia-
stereoisomers); 80.8 (C4); 78.0 (CB); 75.5 (s, C5′); 75.0 (s, C3);
74.9 (s, C5); 73.2 (s, C2); 73.1 (s, C3′); 70.5 (C2′); 70.2–69.8 (CA,
side (16b). A solution of lactose trichloroacetimidate 5 60 diastereoisomers); 68.1 (C4′); 60.6–60.4 (2s, C6 + C6′); 57.0–56.8
(1.44 g, 1.84 mmol, 1.5 eq.) and 15b (460 mg, 1.23 mmol, 1.0 (OCH3, diastereoisomers); 31.3 (CH2 α fatty chain); 32.1 (CC);
eq.) in dry CH2Cl2 (15 mL) is stirred with molecular sieves 4 Å 29.0; 28.7; 28.6; 28.2; 27.1 (CD + CH2 fatty chain); 22.1 (s, CH2
for one hour under an inert atmosphere. At 0 °C, BF3·Et2O fatty chain); 13.9 (s, CH3 fatty chain). Mass (ESI, 100 mg
(160 µL, 0.98 mmol, 0.8 eq.) is added dropwise and the mL−1): m/z calcd for C33H64O12SNa [M + Na]+: 707.40, found
mixture is stirred for 15 hours at room temperature under an 707.40 [M + Na]+.
inert atmosphere. The mixture is quenched by addition of
water (5 mL). The organic layer is washed twice with aqueous (1–4)-β-D-glucopyranoside
1-O-[-4-(Hexadecylthio)-2-ethoxybutyl]-β-D-galactopyranosyl-
(17b Ohmline-2-OEt-5S).
–
saturated NaHCO3 solution (2 × 10 mL) and an aqueous satu- Potassium carbonate (0.021 mg, 0.15 mmol, 0.5 eq.) is added
rated NaCl solution (10 mL). The organic layer is dried on to a solution of 16b (300 mg, 0.31 mmol, 1.0 eq.) in methanol
MgSO4, filtered and concentrated to give 16b which was puri- (20 mL). The mixture is stirred at room temperature for
fied by chromatography on silica gel (eluent: pentane/ethyl 6 hours. Amberlyst IR-120 (H+) is added and the mixture is
acetate: 7/3) to give 16b (500 mg; 41% yield). Rf (pentane/ethyl stirred for 30 minutes at room temperature. The reaction
1
3
acetate: 7/3): 0.4; H NMR (CDCl3, 500.133): 5.2 (d, 1 H, JHH
=
mixture is warmed to reflux, quickly filtered and concentrated
3.0, H4′); 5.07 (t, 1 H, JHH = 9.5 Hz, H3); 4.97–4.94 (dd, 1 H, JHH to give the compound 17b (211 mg; 99% yield; estimated
= 10.5 Hz, JHH = 8.0 Hz, H2′); 4.85–4.82 (m, 1 H, H3′); 4.78–4.73 purity). 1H NMR (DMSO-d6, 500.251): 5.2–8 (m, 2 H, 2 OH);
(q, 1H, JHH = 9.50 Hz, JHH = 18 Hz, H2); 4.43–4.34 (m, 3 H, H1 + 4.8–4.3 (m, 4 H, OH); 4.67–64 (m, 2 H, 2 OH); 4.22–4.19 (m, 2
H6a + H1′); 4.01–3.94 (m, 3 H, H6a′ + H6b′ + H6b); 3.80 (t, 1 H, H, H1 + H1′); 3.76–3.71 (m, 2 H, H6a + CH2 HA); 3.61–3.41 (m,
JHH = 7.0 Hz, H5′); 3.68 (t, 2H, JHH = 9.0 Hz, H4 + HA1); 15H, H3 + H4 + H5 + H6b + H2′ + H3′ + H4′ + H5′ + H6′a + H6′b
+
3.50–3.30 (m, 5H, H5 + HB + HA2 + CH2 –OEt diastereoisomers); Hb CH2 HA + CH HB + CH2 –OEt); 3.0–2.98 (m, 1 H, H2);
2.45–2.33 (m, 4 H, HD + CH2 fatty chain); 2.03–1.81 (m, 25 H, 2.51–2.44 (m, 4 H, CH2 HD + CH2 α fatty chain); 1.60 (m, 1H,
CH3 acetyl); 1.61–1.54 (m, 2 H, CH2 HC); 1.42 (qt, 2 H, JHH = 7.4 CHC1); 1.49 (m, 1H, CHC2); 1.52 (m, 1 H, CH2 β fatty chain);
Hz, CH2 β fatty chain); 1.23 (m, 2H, CH2 γ fatty chain); 1.11 (s, 1.33–1.22 (m, 25 H, CH2 fatty chain); 1.1–1.06 (m, 2H, CH2
26 H, CH2 fatty chain); 1.0 (t, 3H, JHH = 7.0 Hz, CH3 –OEt); 0.74 –OEt); 0.87 (t, 3 H, JHH = 7.0 Hz, CH3 fatty chain); 13C NMR
(t, 3 H, JHH = 7.0 Hz, CH3 fatty chain); 13C NMR (CDCl3, (DMSO-d6, 125.804): 103.8 (s, C1′); 102.8 + 102.7 (C1 diastereo-
125.771): 169.94(s, CvO); 169.92 (s, CvO); 169.8 (s, CvO); isomers); 80.8 (C4); 76.2 (CB); 75.5 (s, C5′); 75.0 (s, C3); 74.8 (s,
169.6 (s, CvO); 169.4 (s, CvO); 169.2 (s, CvO); 169.0 (s, C5); 73.2 (s, C2); 73.1 (s, C3′); 70.7 (CA1 diastereoisomers); 70.5
CvO); 168.7 (s, CvO); 100.7 (s, C1′); 100.33 + 100.28 (s, C1α + (C2′); 70.3 (CA2, diastereoisomers); 68.1 (C4′); 64.4 + 64.3 (2s,
β); 76.4–76.1 (CB); 75.9 (s, C4); 72.6 (s, C3); 72.3 (s, C5); 72.1 CH2 –OEt diastereoisomers); 60.5–60.3 (2s, C6 + C6′); 31.7 (CH2
(CA1); 71.3 (s, C2); 70.7 (C3′); 70.4 (CA2) 70.3 (C5′); 68.8 (s, C2′); α fatty chain); 32.1 (CC); 40.0; 28.9; 28.8; 28.7; 28.6; 28.1; 27.1
66.4 (s, C4′); 65.6–65.0 (OCH2 diastereoisomers); 61.7 (s, C6′); (CD + CH2 fatty chain); 22.1 (s, CH2 fatty chain); 15.5 (s, CH3
61.0 (s, C6); 31.7–31.5 (m, CH2 α fatty chain + Cc); 30.0 (s, CH2 –OEt diastereoisomers); 13.9 (s, CH3 fatty chain). Mass (ESI
fatty chain); 29.3, 29.2, 29.0, 28.6, 27.5, 26.5 (CD + CH2 fatty QqTOF): m/z calcd for C34H66O12SNa [M + Na]+: 721.4173,
chain); 22.3 (s, CH2 fatty chain); 20.6–20.1 (m, OCH3); 15.3 (s, found 721.4171.
OEt); 13.8 (CH3 fatty chain). Mass (Maldi TOF, Matrix HCCA):
Cell culture
m/z calcd for C48H80O19SK [M + K]+: 1031.465, found 1031.567.
1-O-[-4-(Hexadecylthio)-2-methoxybutyl]-β-D-galactopyrano-
HEK293T was obtained from the American Type Culture
syl-(1–4)-β-D-glucopyranoside (17a – Ohmline-5S). NaOMe (0.5 Collection (ATCC) and cultured in Dulbecco’s modified Eagle’s
M solution in MeOH, 33 µL, 0.90 mg, 0.017 mmol, 0.1 eq.) is medium, supplemented with 10% (v : v) fetal bovine serum
added to a solution of 16a (163 mg, 0.17 mmol, 1.0 eq.) in (Lonza, France). The cells were grown under a humidified
methanol (10 mL). The mixture is stirred at room temperature atmosphere at 37 °C with 95% air and 5% CO2. HEK293T cells
for one hour. Amberlyst IR-120 (H+) is added and the mixture were transduced as previously described to achieve stable
is stirred for 30 minutes at room temperature. The reaction expression of homotetramer SK3 channels.16,29 Human cancer
mixture is warmed to reflux, quickly filtered and concentrated cell lines MDA-MB-435s, HCT116 and PC3 were purchased at
1
to give 17a (99 mg; 87% yield). H NMR (DMSO-d6, 300.131): the American Type Culture Collection and grown respectively
5.12–5.09 (m, 2 H, 2 OH); 4.77 (brs, 1 H, OH); 4.67–64 (m, 2 H, in Alpha Dulbecco’s modified Eagle’s medium (USA,
2 OH); 4.54–4.50 (m, 2 H, 2 OH); 4.22–4.19 (m, 2 H, H1 + H1′); Manassas, Virginia), McCoy’s 5A medium and RPMI medium
This journal is © The Royal Society of Chemistry 2021
Org. Biomol. Chem., 2021, 19, 2753–2766 | 2763

View Article Online
Paper
Organic & Biomolecular Chemistry
containing 10% foetal bovine serum (Hyclone, Lonza, 10 minutes. The maximum of fluorescence (peak of calcium
Levallois-Perret, France) at 37 °C, 5% CO2 and at moisture sat- influx (F340/F380)) after injection of 2 mM Ca2+ is measured
uration as already described.17,18,58 We have tested compounds and normalized to control conditions. Data acquisition was
on these cell lines because we already demonstrated that the performed using SoftMax pro software. Fura-2-loaded cells
SK3 channel is expressed in these cells and promotes their were alternately illuminated with 340 and 380 nm.
calcium entries (including SOCE, see below) and their Fluorescence emission was collected at 510 nm. Figures
migration.17–19
showing Ca2+ traces display averages from several wells per
condition and are representative of several independent
recordings.
Electrophysiological recordings
Experiments were performed in the whole-cell configuration of
the patch-clamp technique. The patch pipettes were filled with
solution containing 145 mM KCl, 1 mM MgCl2, 1 mM Mg-ATP,
Vasorelaxation experiments
Animals. Male Wistar rats (350–450 g) were purchased from
0.7 mM CaCl2, 1 mM EGTA and 10 mM HEPES (PCa 6). The CER Janvier (Le Genest, St Isle, France). All experiments invol-
pH was adjusted to 7.2 with KOH. The bath solution contained ving animals were approved by the institutional ethical com-
140 mM NaCl, 4 mM KCl, 2 mM CaCl2, 1 mM MgCl2, 10 mM mittee (Comité d’éthique en expérimentation animal Val de
HEPES, 0.33 mM NaH2PO4, and 11.5 mM D-glucose. The pH Loire, Tours, France, Permit number: 2016090711251954) and
was adjusted to 7.4 with NaOH. Patch-clamp experiments and carried out in accordance with European guidelines on animal
data analyses were conducted as previously described.26 The experimentation and the French “Ministère de l’Enseignement
effects of compounds on HEK293T cells expressing SK3 were supérieur, de la Recherche et de l’Innovation”
measured using a ramp protocol from +100 mV to −100 mV
with a holding potential of 0 mV (500 ms duration; 4 s inter- thetised with a bolus injection of sodium pentobarbital (60 mg
vals) to inactive endogenous potassium currents. The ampli- kg−1, i.p.). After intravenous injection of heparin (500 IU kg−1
Preparation of isolated mesenteric arteries. Rats were anaes-
)
tude of the apamin-sensitive current was obtained by subtrac- the mesenteric bed irrigating the ileum was removed and
tion of the amplitude of the current before and after the appli- immediately placed in a Krebs–Henseleit solution containing
cation of 100 nmol L−1 apamin (a specific SKCa blocker).
119 mM NaCl, 4.7 mM KCl, 1.18 mM KH2PO4, 1.17 mM
MgSO4, 1.36 mM CaCl2, 25 mM NaHCO3 and 5.5 mM glucose.
After dissection of surrounding tissue, the second-order
In vitro cell viability and cell migration assay
Cell viability was determined using the tetrazolium salt mesenteric resistance arteries were cut into 1.5–2 mm ring seg-
reduction method (MTT) and cell migration assay was per- ments and were individually mounted in a Mulvany–Halpern
formed in 24-well plate cell culture inserts with a pore size of myograph (Multi Myograph system-610, DMT, Aarhus,
8
μm as previously described.61 HCT116 and PC3 cell Denmark). The 9 ml organ chamber was filled with Krebs’
migration was performed as already described.18,54 Briefly, solution continuously bubbled with 95% O2/5% CO2 and
after 24 h, stationary cells were removed from the top side, maintained at 37 °C.
whereas migrated cells in the bottom side of the inserts were
Experiment protocols. Experiments were started after an
fixed with DAPI (Sigma-Aldrich) and then counted. Three inde- equilibration period of at least 30 minutes. The mesenteric
pendent experiments were performed and these were done in arteries were stretched to a tension of 3 mN mm−1 which
triplicate.
corresponds to the physiological conditions. The measure-
ments of isometric force (mN mm−1) were performed accord-
ing to the methods described by Mulvany and Halpern.62
Ca2+ entry measurements
Time-lapse [Ca2+] cytosolic measurements were performed Isometric wall tensions of mesenteric arteries were measured
using
a FlexStation III fluorometer (Molecular Devices). with force transducers connected to a Powerlab system and
HCT116 and PC3 cells were seeded on 96-well flat clear-bottom recorded using Chart 5 software. To evaluate the vasorelaxant
black microplates (#3603, Corning) at a density of 3.104 cells effect of the compounds, the mesenteric artery rings were pre-
per well. HCT116 and PC3 cells were grown on fibronectin- contracted with U46619 (3 µM), an agonist of thromboxane A2
coated microplates. One day after seeding, HCT116 and PC3 receptors. The role of the endothelium in the relaxation effect
cells were incubated with 4 mmol L−1 Fura-2/AM in culture induced by 7b (1 µM) has been tested in mesenteric artery
medium for 35 minutes at 37 °C protected from light. rings with or without the endothelium. The endothelium was
Afterward, the cells were gently washed three times with removed by gently rubbing the mesenteric artery luminal
HEPES buffered saline (140 mmol L−1 NaCl, 1.13 mmol L−1 surface with a human hair. The integrity or functional removal
MgCl2, 4.7 mmol L−1 KCl, 2 mmol L−1 CaCl2, 10 mmol L−1 of the endothelium was examined from the endothelium-
D-glucose, and 10 mmol L−1 HEPES, pH 7.4) and left for dependent relaxation response to acetylcholine (10 µM).
5 minutes at room temperature protected from light. For store
Statistics
operated Ca2+ entry (SOCE) experiments, cells were put in
HEPES buffered saline Ca2+-free solution and treated with Statistical analyses have been performed using SigmaStat
thapsigargin (4 µM) (T7458, Life-Technologies): to deplete the Software (version 3.0.1a, Systat Software, Inc). Unless otherwise
intracellular store before injection of 2 mM CaCl2 after indicated, data were expressed as mean standard error of the
2764 | Org. Biomol. Chem., 2021, 19, 2753–2766
This journal is © The Royal Society of Chemistry 2021

View Article Online
Organic & Biomolecular Chemistry
Paper
mean (N, number of experiments and n, number of cells from
independent experiments). For comparison between more
than two means we used Kruskal–Wallis one way analysis of
variance followed by Dunn’s or Dunnet’s post hoc tests as
appropriate. Comparisons between two means were made
using the Mann–Whitney test and using Student t-text for
vasorelaxation experiments. Differences were considered sig-
nificant when p < 0.05.
6 C. Rua, M. Guéguinou, I. Soubai, E. Viel, M. Potier-
Cartereau, A. Chantome, C. Barbe, P. Bougnoux, C. Barin-
Le Guellec and C. Vandier, Clin. Cancer Res., 2018, 24,
5313–5320.
7 L. Hipólito, A. K. Fakira, D. Cabañero, R. Blandón,
S. M. Carlton, J. A. Morón and Z. Melyan, Pain, 2015, 156,
849–858.
8 A. W. Kasumu, C. Hougaard, F. Rode, T. A. Jacobsen,
J. M. Sabatier, B. L. Eriksen, D. Strøbæk, X. Liang,
P. Egorova, D. Vorontsova, P. Christophersen, L. C. B. Rønn
and I. Bezprozvanny, Chem. Biol., 2012, 19, 1340–1353.
9 A. M. Dolga, T. Letsche, M. Gold, N. Doti, M. Bacher,
N. Chiamvimonvat, R. Dodel and C. G. Culmsee, Glia,
2012, 60, 2050–2064.
Author contributions
HC, CV, and PAJ conceived the study and the new analogues of
Ohmline and CMS, AB and HC performed the organic syn-
thesis and the characterization of the new amphiphiles. KM, 10 C. T. Bond, R. Sprengel, J. M. Bissonnette, W. A. Kaufmann,
CP, VM and CV conceived the biological studies. DF performed
in vitro cell viability and cell migration assays. AC performed
the electrophysiological recordings. MG performed Ca2+ entry
D. Pribnow, T. Neelands, T. Storck, M. Baetscher, J. Jerecic,
J. Maylie, H. G. Knaus, P. H. Seeburg and J. P. Adelman,
Science, 2000, 289, 1942–1946.
measurements. BLZ performed the mesenteric artery tone 11 A. Brown, T. Cornwell, I. Korniyenko, V. Solodushko,
measurements. PAJ and CV wrote the paper with input from
all authors.
C. T. Bond, J. P. Adelman and M. S. Taylor, Am. J. Physiol.:
Cell Physiol., 2007, 292, C832–C840.
12 S. L. Pierce, J. D. K. Kresowik, K. G. Lamping and
S. K. England, Biol. Reprod., 2008, 78, 1058–1063.
13 H. Lee, B. H. Koh, L. E. Peri, R. D. Corrigan, H. T. Lee,
N. E. George, B. P. Bhetwal, Y. Xie, B. A. Perrino, T. C. Chai,
K. M. Sanders and S. D. Koh, Sci. Rep., 2017, 7, 12245.
14 S. Brähler, A. Kaistha, V. J. Schmidt, S. E. Wölfle, C. Busch,
B. P. Kaistha, M. Kacik, A. L. Hasenau, I. Grgic, H. Si,
C. T. Bond, J. P. Adelman, H. Wulff, C. de Wit, J. Hoyer and
R. Köhler, Circulation, 2009, 119, 2323–2332.
Conflicts of interest
There are no conflicts to declare.
Acknowledgements
We acknowledge CNRS, INSERM, la ligue contre le cancer, 15 T. Siddiqui, S. Lively, R. Ferreira, R. Wong and
CANCEN, la région Bretagne, Université de Brest, and L. C. Schlichter, PLoS One, 2014, 9, e106087.
Université de Tours, Région Centre-Val de Loire for their finan- 16 A. Chantome, A. Girault, M. Potier, C. Collin, P. Vaudin,
cial support. We also thank Isabelle Domingo for her technical
assistance, the Cancéropôle Grand Ouest and the platforms
J. C. Pagès, C. Vandier and V. Joulin, Exp. Cell Res., 2009,
315, 3620–3630.
(RMN-RPE and spectrométrie de masse) from the University of 17 A. Chantôme, M. Potier-Cartereau, L. Clarysse, G. Fromont,
Brest. This study was funded by Institut National du Cancer
(INCa No 2018-151),
S. Marionneau-Lambot, M. Guéguinou, J. C. Pagès,
C. Collin, T. Oullier, A. Girault, F. Arbion, J. P. Haelters,
P. A. Jaffrès, M. Pinault, P. Besson, V. Joulin, P. Bougnoux
and C. Vandier, Cancer Res., 2013, 73, 4852–4861.
18 M. Guéguinou, T. Harnois, D. Crottes, N. Deliot,
A. Gambade, A. Chantôme, J. P. Haelters, P. A. Jaffrès,
M. L. Jourdan, G. Weber, P. Bougnoux, O. Mignen,
N. Bourmeyster, B. Constantin, T. Lecomte, C. Vandier and
M. Potier-Cartereau, Oncotarget, 2016, 7, 36168–36184.
Notes and references
1 Q. Kuang, P. Purhonen and H. Hebert, Cell. Mol. Life Sci.,
2015, 72, 3677–3693.
2 A. G. Torrente, R. Zhang, H. Wang, A. Zaini, B. Kim, X. Yue,
K. D. Philipson and J. I. Goldhaber, J. Physiol., 2017, 595, 19 M. Potier, V. Joulin, S. Roger, P. Besson, M. L. Jourdan,
3847–3865.
J. Y. LeGuennec, P. Bougnoux and C. Vandier, Mol. Cancer
Ther., 2006, 5, 2946–2953.
3 F. C. Yap, D. S. Weber, M. S. Taylor, M. I. Townsley,
B. S. Comer, J. Maylie, J. P. Adelman and M. T. Lin, 20 C. Hanselmann and S. Grissmer, J. Physiol., 1996, 496, 627–
Am. J. Physiol.: Heart Circ. Physiol., 2016, 310, H1151–
H1163.
4 B. Honrath, I. E. Krabbendam, C. Culmsee and
A. M. Dolga, Neurochem. Int., 2017, 109, 13–23.
637.
21 K. L. Weatherall, V. Seutin, J. F. Liégeois and N. V. Marrion,
Proc. Natl. Acad. Sci. U. S. A., 2011, 108, 18494–18499.
22 P. M. Dunn, Eur. J. Pharmacol., 1999, 368, 119–123.
5 A. Girault, J. P. Haelters, M. Potier-Cartereau, A. Chantome, 23 D. Strøbaek, T. D. Jørgensen, P. Christophersen,
P. A. Jaffrès, P. Bougnoux, V. Joulin and C. Vandier, Curr.
Med. Chem., 2012, 19, 697–713.
P. K. Ahring and S. P. Olesen, Br. J. Pharmacol., 2000, 129,
991–999.
This journal is © The Royal Society of Chemistry 2021
Org. Biomol. Chem., 2021, 19, 2753–2766 | 2765

View Article Online
Paper
Organic & Biomolecular Chemistry
24 D. Strobaek, C. Hougaard, T. H. Johansen, U. S. Sorensen, 42 S. Drescher, A. Meister, G. Graf, G. Hause, A. Blume and
E. O. Nielsen, K. S. Nielsen, R. D. Taylor, P. Pedarzani and
P. Christophersen, Mol. Pharmacol., 2006, 70, 1771–1782.
25 C. H. Lee and R. MacKinnon, Science, 2018, 360, 508–513.
26 R. Vitello, F. Kerff and J. F. Liégeois, Acta Physiol., 2021,
231, e13574.
27 Y. W. Nam, M. Cui, R. Orfali, A. Viegas, N. Nguyen,
E. H. M. Mohammed, K. A. Zoghebi, S. Rahighi, K. Parang
and M. Zhang, Acta Physiol., 2021, 231, e13552.
B. Dobner, Chem. – Eur. J., 2008, 14, 6796–6804.
43 A. Bouraoui, M. Berchel, R. Ghanem, V. Vié, G. Paboeuf,
L. Deschamps, O. Lozach, T. Le Gall, T. Montier and
P. A. Jaffrès, Org. Biomol. Chem., 2019, 17, 3609–3616.
44 A. Bouraoui, R. Ghanem, M. Berchel, V. Vié, Y. Le Guen,
G. Paboeuf, L. Deschamps, T. Le Gall, T. Montier and
P. A. Jaffrès, ChemPhysChem, 2019, 20, 2187–2194.
45 W. Vogt, Free Radicals Biol. Med., 1995, 18, 93–105.
28 C. Hougaard, B. L. Eriksen, S. Jørgensen, T. H. Johansen, 46 J. Tang, K. A. Usmani, E. Hodgson and R. L. Rose, Chem.-
T. Dyhring, L. S. Madsen, D. Strøbæk and
P. Christophersen, Br. J. Pharmacol., 2007, 151, 655–665.
29 A. Girault, J. P. Haelters, M. Potier, A. Chantôme,
Biol. Interact., 2004, 147, 319–329.
47 S. C. Mitchell and R. M. Nickson, Sulfur Rep., 1993, 13,
161–195.
M. Pineau, S. Marionneau-Lambot, T. Oullier, G. Simon, 48 D. D. Lawson, H. R. Getz and D. A. Miller, J. Org. Chem.,
H. Couthon-Gourvès, P. A. Jaffrès, V. Joulin, B. Corbel, 1961, 26, 615–616.
P. Bougnoux and C. Vandier, Curr. Cancer Drug Targets, 49 S. Morris-Natschke, J. R. Surles, L. W. Danie, M. E. Berens,
2011, 11, 1111–1125.
30 C. Vandier, P. Bougnoux, A. Chantome, B. Corbel,
E. J. Modest and C. Piantadosi, J. Med. Chem., 1986, 29,
2114–2117.
A. Girault, J. P. Haelters, V. Joulin, M. Potier-Cartereau and 50 R. L. Alexander, S. L. Morris-Natschke, K. S. Ishaq,
G. Simon, Method for preventing cancer metastasis, PCT
Int. Appl., WO2011101408A120110825, 2011.
R. A. Fleming and G. L. Kucera, J. Med. Chem., 2003, 46,
4205–4208.
31 L. Heginbotham and R. MacKinnon, Neuron, 1992, 8, 483– 51 B. Ayers, H. Long, E. Sim, I. A. Smellie, B. L. Wilkinson and
491. A. J. Fairbanks, Carbohydr. Res., 2009, 344, 739–746.
32 P. A. Jaffrès, C. Gajate, A. M. Bouchet, H. Couthon-Gourvès, 52 D. B. G. Williams and M. Lawton, Org. Biomol. Chem., 2005,
A. Chantôme, M. Potier-Cartereau, P. Besson, P. Bougnoux, 3, 3269–3272.
F. Mollinedo and C. Vandier, Pharmacol. Ther., 2016, 165, 53 A. Wimmer and B. König, Beilstein J. Org. Chem., 2018, 14,
114–131. 54–83.
33 F. Herrera, C. M. Sevrain, P. A. Jaffrès, H. Couthon-Gourvès, 54 D. Afonso, T. Le Gall, H. Couthon-Gourvès, A. Grélard,
A. Grélard, E. J. Dufourc, A. Chantôme, M. Potier-
Cartereau, C. Vandier and A. Bouchet, ACS Omega, 2017, 2,
6361–6370.
S. Prakash, M. Berchel, N. Kervarec, E. J. Dufourc,
T. Montier and P. A. Jaffrès, Soft Matter, 2016, 12, 4516–
4520.
34 P. V. Escriba, Biochim. Biophys. Acta, 2017, 1859, 1493–1506. 55 M. Berchel, T. Le Gall, O. Lozach, J. P. Haelters, T. Montier
35 C. M. Sevrain, J. P. Haelters, A. Chantôme, H. Couthon- and P. A. Jaffrès, Org. Biomol. Chem., 2016, 14, 2846–2853.
Gourvès, M. Gueguinou, M. Potier-Cartereau, C. Vandier 56 T. J. Deming, Bioconjugate Chem., 2017, 28, 691–700.
and P. A. Jaffrès, Org. Biomol. Chem., 2013, 11, 4479–4487.
36 W. Berthe, C. M. Sevrain, A. Chantôme, A. M. Bouchet,
M. Gueguinou, Y. Fourbon, M. Potier-Cartereau,
57 F. Bery, S. Figiel, S. Kouba, D. Fontaine, M. Guéguinou,
M. Potier-Cartereau, C. Vandier, R. Guibon, F. Bruyère,
G. Fromont and K. Mahéo, Int. J. Mol. Sci., 2020, 21, 4786.
J. P. Haelters, H. Couthon-Gourvès, C. Vandier and 58 S. Figiel, F. Bery, A. Chantôme, D. Fontaine, C. Pasqualin,
P. A. Jaffrès, ChemMedChem, 2016, 11, 1531–1539.
37 C. M. Sevrain, J. P. Haelters, A. Chantôme, H. Couthon-
Gourvès, A. Girault, C. Vandier and P. A. Jaffrès,
MedChemComm, 2012, 3, 1471–1478.
38 A. Bauduin, M. Papin, A. Chantôme, H. Couthon,
L. Deschamps, J. Requejo-Isidro, C. Vandier and
P. A. Jaffrès, Eur. J. Med. Chem., 2021, 209, 112894.
39 D. Andri, R. Fourme and K. Zechmeister, Acta Crystallogr.,
1972, B28, 2389.
V. Maupoil, I. Domingo, R. Guibon, F. Bruyère, M. Potier-
Cartereau, C. Vandier, G. Fromont and K. Mahéo, Cancers,
2019, 11, 1814.
59 M. Potier-Cartereau, W. Raoul, G. Weber, K. Mahéo,
R. Rapetti-Mauss, M. Gueguinou, P. Buscaglia, C. Goupille,
N. Le Goux, S. Abdoul-Azize, T. Lecomte, G. Fromont,
A. Chantome, O. Mignen, O. Soriani and C. Vandier, Rev.
Physiol., Biochem. Pharmacol., 2020, DOI: 10.1007/
112_2020_24.
40 O. Gereben and L. Pusztai, J. Phys. Chem. B, 2012, 116, 60 M. Yamabe, K. Kaihatsu and Y. Ebara, Bioconjugate Chem.,
9114–9121. 2018, 29, 1490–1494.
41 R. A. O’Brien, A. Mirjafari, K. M. Mattson, S. M. Murray, 61 M. Potier, A. Chantome, V. Joulin, A. Girault, S. Roger,
N. Mobarrez, E. A. Salter, A. Wierzbicki, J. H. Davis and
K. N. West, J. Phys. Chem. B, 2014, 118, 10232–
10239.
P. Besson, M.-L. Jourdan, J.-Y. Le Guennec, P. Bougnoux
and C. Vandier, Br. J. Pharmacol., 2011, 162, 464–479.
62 M. J. Mulvany and W. Halpern, Circ. Res., 1977, 41, 19–26.
2766 | Org. Biomol. Chem., 2021, 19, 2753–2766
This journal is © The Royal Society of Chemistry 2021