Light-Activated COS/H2S Donation from Photocaged Thiocarbamates

Article abstract of DOI:10.1021/acs.orglett.7b00808

Hydrogen sulfide (H₂S) is an important biomolecule, and responsive chemical tools for its delivery are needed. Here, we utilize the photocleavable o-nitrobenzyl group to unmask caged thiocarbamates and to access photoactivated H₂S releasing molecules. These donors function by the initial release of carbonyl sulfide (COS), which is quickly hydrolyzed to H₂S by carbonic anhydrase (CA). Our investigations demonstrate that o-nitrobenzyl-caged thiocarbamates can serve as a donor platform for the bio-orthogonal stimulated release of COS/H₂S.

Full text of DOI:10.1021/acs.orglett.7b00808

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Letter
pubs.acs.org/OrgLett
Light-Activated COS/H S Donation from Photocaged Thiocarbamates
2
Yu Zhao, Sarah G. Bolton, and Michael D. Pluth*
Department of Chemistry and Biochemistry, Material Science Institute, Institute of Molecular Biology, University of Oregon, Eugene,
Oregon 97403, United States
*
S Supporting Information
ABSTRACT: Hydrogen sulfide (H S) is an important
2
biomolecule, and responsive chemical tools for its delivery
are needed. Here, we utilize the photocleavable o-nitrobenzyl
group to unmask caged thiocarbamates and to access
photoactivated H S releasing molecules. These donors function
2
by the initial release of carbonyl sulfide (COS), which is quickly
hydrolyzed to H S by carbonic anhydrase (CA). Our
2
investigations demonstrate that o-nitrobenzyl-caged thiocarbamates can serve as a donor platform for the bio-orthogonal
stimulated release of COS/H S.
2
ydrogen sulfide (H S) is an important biological gaseous
molecule that exhibits potent protective activities in
mammals and which is recognized as the third member of the
2
H
gasotransmitter family, along with nitric oxide (NO) and
1
carbon monoxide (CO). Endogenous H S is synthesized
2
primarily from cysteine and homocysteine by enzymes such as
cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE),
and cysteine aminotransferase (CAT)/3-mercaptopyruvate
2
sulfur transferase (3-MST). Both endogenous H S synthesis
2
and exogenous H S administration exhibit promising protec-
2
3
tions in different stages of diseases. For example, H S can relax
blood vessels by activating KATP channels and can rescue
2
4
cardiomyocytes from oxidative stress damage by scavenging
5
cellular reactive oxygen species (ROS). Furthermore, H S-
2
releasing nonsteroidal anti-inflammation drugs (H S−
2
NSAIDs), which couple NSAIDs with H S-releasing molecules,
2
Figure 1. Examples of selected synthetic H S donors that release H S
directly.
2
2
have been shown to significantly diminish NSAID-induced G6I
damage while maintaining potent anti-inflammation activities.
All of these findings illustrate the positive roles of H S in
human health and also highlight the therapeutic potential of
2
1
0
oxygen species (ROS) as well as bioorthogonal triggers such
11
synthetic H S-releasing molecules against different types of
as the cyclooctene/tetrazine click reaction. Further demon-
strating the utility of intermediate COS release to access H S
2
2
damage and diseases.
To modulate biological levels of H S, researchers often use
donors, similar approaches have also been used by other
researchers to access donor motifs triggered by biological
nucleophiles and esterases.
To expand the basic chemistry of triggered COS/H S
2
release, we envisioned that caged-thiocarbamate compounds
with photoactivatable triggering groups could serve as a new
2
H S-releasing molecules (“H S donors”) rather than inorganic
2
2
12
13
sulfide salts, such as sodium sulfide (Na S) and sodium
2
hydrogen sulfide (NaSH), due to the rapid and uncontrollable
7
release of H S from these inorganic precursors. Because of
2
these limitations, synthetic H S donors have attracted
2
7e,8
tunable H S release platform that responds to specific bio-
significant attention in the past several years (Figure 1),
2
orthogonal stimuli. Photoactivatable triggering groups have
been widely used in chemistry and biology and offer advantages
of high modularity and rapid triggering by an external
although numerous limitations remain. Aligned with the need
to develop new methods of H S release, our laboratory recently
2
reported a new class of H S donors by engineering the release
2
14
stimulus. For example, photoactivated donors that release
of carbonyl sulfide (COS) from caged-thiocarbamate mole-
15
16
17
18
and H2S
9
CO, nitroxyl (HNO), NO,
have all been
cules. The released COS is quickly hydrolyzed to H S by the
2
ubiquitous enzyme carbonic anhydrase (CA) (Scheme 1a). We
have used this general COS release strategy to develop
platforms activated by endogenous triggers including reactive
Received: March 17, 2017
©
XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.7b00808
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Organic Letters
Letter
Scheme 1. (a) Caged Thiocarbamates as Tunable H2S
Donors; (b) ONB-Caged Thiocarbamates as Photolabile
7.4, 10 mM) containing CA (25 μg/mL). Consistent with our
hypothesis, we observed a time-dependent H S release from
2
H S Donors
2
both PhotoTCM-1 and PhotoTCM-2 with a peaking time for
H S release of ∼10 min (Figure 2).
2
reported and provide powerful tools for delivering these small
molecule payloads in diverse applications. Herein, we report the
first class of photolabile COS releasing molecules by employing
18a,19
the photoactivated o-nitrobenzyl (ONB) protecting group
Scheme 1) and demonstrate the compatibility of caged
(
thiocarbamates with ONB photocleavage mechanisms.
To test our hypothesis that ONB-functionalized thiocarba-
Figure 2. H S release from PhotoTCM-1 and PhotoTCM-2 in
aqueous buffer at pH 7.4.
2
mates can function as photolabile COS/H S donors, we
2
prepared two donors (PhotoThioCarbaMate: PhotoTCM-1
and PhotoTCM-2), the parent carbamate molecule (Photo-
CarbaMate: PhotoCM-1), which releases CO /H O instead of
To confirm that the observed COS/H S release was due to
2
2
2
10
photocleavage of the ONB protecting group, a series of control
experiments were performed. As shown in Figure 3a, UV
irradiation of a PBS solution containing CA does not generate
H S. Similarly, PhotoTCM-1 does not generate H S in the
COS/H S, and a thiocarbamate (TCM-1) lacking the ONB-
2
photoactivated trigger (Scheme 2). PhotoTCM-1 and
Scheme 2. Synthesis of Photolabile H S Donors and
2
2
2
absence of UV irradiation. Upon UV irradiation of PhotoTCM-
in the absence of CA (bar 4 in Figure 3a), we observed low
Photolabile Carbamate
1
levels of H S, which is consistent with the slow background
2
hydrolysis of COS to H S in the absence of CA. As expected,
2
TCM-1, which lacks the ONB group, does not release H2S
upon the treatment of UV light, showing that the
thiocarbamate linkage is not photodegraded to generate H S.
2
Similarly, no H S release was observed from the carbamate
2
control compound PhotoCM-1. As expected, incubation of
PhotoTCM-1 with CA under UV exposure results in a
significant enhancement of H S concentration, indicating UV
2
irradiation is essential to trigger H S release. Moreover, the
2
measured H S is significantly reduced in the presence of
2
acetazolamide (AAA), a well-known CA inhibitor. In addition,
incubation of PhotoTCM-1 with cellular nucleophiles, such as
cysteine, homocysteine, reduced glutathione, oxidized gluta-
thione, serine, and lysine, in the presence of CA does not
PhotoTCM-2 were prepared by treatment of o-nitrobenzyl
alcohols with p-fluorophenyl isothiocyanate in the presence of
sodium hydride in good yield. PhotoCM-1 was synthesized
from the reaction between o-nitrobenzyl alcohol and p-
fluorophenyl isocyanate in the presence of triethylamine.
With these compounds in hand, we next tested their H2S
generate H S, demonstrating PhotoTCM-1 is stable toward
2
cellular nucleophiles (Figure 3b). We note that PhotoTCM-1
was incubated with the nucleophiles for 10 min to match the
peaking time for the COS/H S release experiments. We did not
2
releasing profiles in the presence of CA. Common H S-
investigate longer incubation times in this preliminary report,
although it is possible that some side reactivity could occur at
extended times or with more potent biological nucleophiles.
Taken together, these studies demonstrate that ONB-function-
alized thiocarbamates provide a functional platform to access
photolabile H S donors and that H S production can be
2
responsive electrodes contain photosensitive components and
produce a photocurrent under UV light, making their use
impractical for these investigations. Therefore, we chose to use
the methylene blue assay to monitor H S release from
2
PhotoTCM-1 (50 μM) and PhotoTCM-2 (50 μM) upon
irradiation with an LED UV light (365 nm) in PBS buffer (pH
2
2
triggered and tuned by UV irradiation.
B
DOI: 10.1021/acs.orglett.7b00808
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Organic Letters
Letter
group is compatible with thiocarbamate linkages, thus providing
a broad platform for further expansion. The time-dependent
H S release after UV activation suggests that these compounds
2
can be used as new efficacious photolabile COS/H S donors.
2
Further applications of these as well as other COS-releasing
constructs triggered by different mechanisms are ongoing in
our laboratory.
ASSOCIATED CONTENT
Supporting Information
■
*
S
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.or-
glett.7b00808.
Experimental procedures and characterization data
(PDF)
AUTHOR INFORMATION
Corresponding Author
E-mail: pluth@uoregon.edu.
■
*
ORCID
Michael D. Pluth: 0000-0003-3604-653X
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
Figure 3. (a) H S release from PhotoTCM-1, TCM-1, and
PhotoCM-1 under various conditions. (b) Effects of amino acids
■
2
Research reported in this publication was supported by the
Sloan Foundation and the Dreyfus Foundation. NMR spec-
troscopy capabilities in the UO CAMCOR facility are
supported by the NSF (CHE-1427987). We thank Prof.
Ramesh Jasti (UO) and his group for assistance with the LED
photolysis experiments.
(
500 μM, 5 mM for GSH) on H S release from PhotoTCM-1.
2
Absorbance measurements were acquired after PhotoTCM-1 was
incubated with amino acids in PBS for 10 min.
The proposed COS/H S release mechanism from the
2
PhotoTCM donors is shown in Scheme 3. UV-mediated
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