Sulfo-click chemistry with 18F-labeled thio acids

Article abstract of DOI:10.1039/c8cc08717b

The first application of sulfo-click chemistry with ¹⁸F-labeled thio acids is described. The simple one-pot/three-component reaction proceeded rapidly within 30 min using mild reaction conditions to give various ¹⁸F-labeled small molecule N-acylsulfonamides in radiochemical conversions of 38-99%, and radiolabeled peptides in 20-25% isolated and decay-corrected radiochemical yields.

Full text of DOI:10.1039/c8cc08717b

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Sulfo-click chemistry with F-labeled thio acids†
Jenna Urkow, Cody Bergman and Frank Wuest
*
Cite this:DOI: 10.1039/c8cc08717b
Received 31st October 2018,
Accepted 8th January 2019
DOI: 10.1039/c8cc08717b
rsc.li/chemcomm
The first application of sulfo-click chemistry with 18F-labeled thio considered as electron-deficient azides, which react much faster
acids is described. The simple one-pot/three-component reaction with thio acids than electron-rich azides like benzyl azide.11
proceeded rapidly within 30 min using mild reaction conditions to
give various 18F-labeled small molecule N-acylsulfonamides in ‘sulfo-click’ chemistry, is a rather new metal-free click chemistry
The thio acid/sulfonyl azide reaction, also referred to as
radiochemical conversions of 38–99%, and radiolabeled peptides variation for chemoselective ligation and bioconjugation reac-
in 20–25% isolated and decay-corrected radiochemical yields.
tions to yield N-acylsulfonamides as interesting bioisosteres of
carboxylic acid compounds.14–17 N-Acyl sulfonamides are a highly
Click chemistry has evolved into an important and highly versatile versatile class of compounds known for many decades, and N-acyl
mainstream method for efficient and selective bioconjugation sulfonamides have recently regained new interest in medicinal
reactions in chemical sciences, including radiopharmaceutical chemistry, including solid support linker synthesis strategies,
chemistry for targeted molecular imaging and radiotherapy.1–3 bioconjugation and ligation chemistry, and kinetic target-guided
Prominent examples include copper(I)-catalyzed alkyne–azide syntheses for drug discovery and chemical biology.17
cycloadditions (CuAAC),4,5 various strain-promoted cycloadditions
The rapid reaction kinetics and the chemical orthogonality
for copper-free click chemistry reactions,6,7 Staudinger ligations,8 of the reaction between thio acids and sulfonylazides as
and oxime/hydrazone formations.9,10 The biorthogonal nature of electron-deficient azides make sulfo-click chemistry a particularly
these bio-conjugation methods has been particularly useful for valuable and reliable bioconjugation method for the chemoselec-
the radiolabeling of various peptides, peptidomimetics and tive modification of peptides of proteins for biomedical research
proteins with a high degree of structural and functional diversity as recently demonstrated for the synthesis of cyclic peptides and
with respect to the used targeting vectors and radionuclides.4,5
site-specific conjugation of peptides with fluorophores and metal
The search for innovative click chemistry and bioconjuga- chelators.14,16 Moreover, sulfo-click chemistry can be performed
tion methods for the efficient and chemoselective introduction in aqueous solvents with a high degree of functional group
of radionuclides into targeting vectors like peptides and proteins compatibility. The proposed mechanism of the sulfo-click reac-
under mild conditions is a very active research area in current tion follows a stepwise addition–cyclization–elimination sequence
radiopharmaceutical chemistry. Amidations are among the via a distinct thiatriazoline intermediate (Scheme 1).11
most prevalent bioconjugation methods in radiopharmaceutical
However, despite the favorable chemoselectivity and versatility
chemistry using a wide range of different reaction conditions of sulfo-click chemistry for bioconjugation reactions, sulfo-click
compatible with even complex in vitro and in vivo studies. In this chemistry has not yet been used for the synthesis of radio-
line, the reaction between azides and thio acids is a rather new pharmaceuticals for molecular imaging and therapy.
bioconjugation method to form respective amide bonds, and
Among the arsenal of commonly used radionuclides for the
the reaction proved to be a very useful method for the rapid design and synthesis of molecular imaging agents, short-lived
conversion of organic azides into corresponding amides.11–13 The positron emitter fluorine-18 (18F) offers several important
scope of thio acid/azide amidation reactions was recently further advantages such as high production yields on small biomedical
expanded through the use of sulfonyl azides. Sulfonyl azides are cyclotrons, high spatial resolution through low positron energy
(0.635 MeV).
Moreover, the convenient 109.8 min half-life which allows
multi-step radiosynthesis procedures and extended positron
Department of Oncology, University of Alberta, Edmonton, Canada.
E-mail: wuest@ualberta.ca; Fax: +1-780-432-8483; Tel: +1-780-989-8150
emission tomography (PET) imaging studies in humans and
small animals.18 Consequently, 18F is among the most frequently
† Electronic supplementary information (ESI) available: Experimental section
and spectral data of compounds. See DOI: 10.1039/c8cc08717b
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Scheme 1 Addition/cyclization/elimination reaction sequence between
thio acids and sulfonylazides as electron-deficient azides to form
N-acylsulfonamides.
Scheme 2 Synthesis of 18F-labeled thio acids [18F]1 and [18F]2.
used radionuclides for the synthesis and application of PET
imaging agents. Radiochemistry with 18F typically exploits either
direct radiofluorination chemistry starting from nucleophilic
[18F]fluoride, electrophilic 18F labeling precursors, or bioconju-
gation reaction concepts using 18F-labeled prosthetic group
chemistry. Many 18F-labeled prosthetic groups are directed for
bioconjugation chemistry with amine or sulfhydryl groups as
typical functional groups prevalent in many peptides and
proteins. Bioconjugation reactions with 18F-labeled prosthetic
groups mainly exploit acylation chemistry of lysine residues or
the N-terminal end with 18F-labeled active esters such as
N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB) or Michael addition
Scheme 3 One-pot/three-component sulfo-click coupling reaction for
the synthesis of 18F-labeled N-acyl sulfonamides.
chemistry of cysteine residues with 18F-labeled maleimides such corresponding 18F-labeled N-acyl sulfonamides in the presence
N-[2-(4-[18F]fluorobenzamido)ethyl]-maleimide ([18F]FBEM).19
of various sulfonylazides (R-SO2N3, 1–2 mg) utilizing an one-
Proposed sulfo-click chemistry with 18F requires 18F-labeled pot/three-component coupling reaction procedure in a 2 mL
thio acids or, likewise, 18F-labeled sulfonyl azides as building Eppendorf vial (A: [18F]SFB/NaSH/R-SO2N3; B: [18F]FBA/succinic
blocks for the formation of respective 18F-labeled N-acyl sulfon- thioanhydride/R-SO2N3) (Scheme 3).
amides. Herein, we describe the first synthesis and use of two
In the case of in situ synthesis of [18F]1 starting from [18F]SFB
18F-labeled thio acids (4-[18F]fluorobenzothioic S-acid [18F]1 and and NaSH, we did not notice release of [18F]fluoride as an
4-((4-([18F]fluoro)benzyl)amino)-4-oxobutane-thioic S-acid [18F]2) indication for a possible competitive nucleophilic aromatic
as novel 18F-labeled sulfo-click chemistry building blocks for substitution reaction between [18F]SFB and NaSH leading to
bioconjugation reactions with various sulfonylazides.
the corresponding thiophenol with [18F]fluorine as a leaving
The synthesis and handling of isolated thio acids can be group. Radiolabeling reactions involving model compounds
quite challenging since thio acids possess nucleophilic proper- and drug-like compounds were performed on an analytical scale
ties and they can oxidize readily to form corresponding disulfide (20–30 MBq of [18F]SFB or [18F]FBA). Radiosyntheses with peptides
dimers. Therefore, we considered in situ formation of 18F-labeled were performed at radioactivity amounts of 100–150 MBq for
thio acids and immediate subsequent reaction with sulfonyl- [18F]SFB or [18F]FBA, respectively. All radiolabeling reactions
azides. Feasibility of sulfo-click chemistry with in situ formed proceeded under mild conditions at 30–50 1C within 30 min.
18F-labeled thio acids [18F]1 and [18F]2 was tested in one-pot/
The first set of reactions explored the feasibility of sulfo-click
three-component coupling reactions with various aryl sulfonyl- chemistry with 18F-labeled thio acids [18F]1 and [18F]2 in the
azides representing small molecule model compounds, a presence of several commercially available aryl sulfonylazides
sulfonylazide-containing celecoxib derivative as an example for (2,4,6-triisopropylbenzenesulfonyl azide, 4-(azidosulfonyl)-benzoic
a drug-like compound, and sulfonyl azide-modified tetra-peptide acid, 4-acetamidobenzenesulfonyl azide and 4-methyl-benzene-
Gly-Leu-Ser-Phe as a model peptide for proposed bioconjugation sulfonyl azide) as model compounds. Formation of respective
of peptides with 18F using sulfo-click chemistry. Synthesis of 18F-labeled N-acyl aryl sulfonamides was confirmed through radio-
18F-labeled thio acids [18F]1 and [18F]2 was accomplished in situ TLC analysis of the reaction mixtures in comparison with non-
through rapid reaction of readily available 18F-labeled building radioactive und fully characterized reference compounds 3–10 and
blocks [18F]SFB20 and 4-[18F]fluorobenzyl amine ([18F]FBA)21 with reported radiochemical conversions (RCCs) were determined by
NaSH and succinic thioanhydride, respectively (Scheme 2).
radio-TLC representing percentage of radiolabeled products
[
18F]3–[18F]10 present in the reaction mixture (for full details,
Formation of thio acids [18F]1 and [18F]2 was monitored by
radio-TLC analysis (see ESI†). Radio-TLC-analysis confirmed including synthesis and characterization of reference com-
formation of [18F]1 and [18F]2 at a radiochemical conversion pounds, see ESI†).
(RCC) of 78% and 73%, respectively. In situ-formed thio acids
[
The used aryl sulfonylazides as reaction partners for the
18F]1 and [18F]2 were not isolated and directly converted into sulfo-click reaction with thio acids [18F]1 and [18F]2 differ in
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Scheme 5 Radiosynthesis of N-acyl sulfonamides [18F]12 and [18F]13
based on anti-inflammatory drug celecoxib.
Scheme 4 Structure and radiochemical conversions (RCCs) of aryl
N-acyl sulfonamides [18F]3–[18F]10.
The feasibility of sulfo-click chemistry with thio acids [18F]1
and [18F]2 for 18F-labeling of peptides was further tested in a
third set of reactions with a sulfonylazide-decorated model
peptide Gly-Leu-Ser-Phe. Peptide synthesis was accomplished
using standard Fmoc-based automated solid-phase peptide
synthesis with 4-(azidosulfonyl)benzoic acid as building block
to attach the sulfonylazide motif onto the N-terminal end of
tetrapeptide Gly-Leu-Ser-Phe (for full details see ESI†).
their electronic and steric nature. The structures and obtained
RCCs of 18F-labeled aryl N-acylsulfonamides [18F]3–[18F]10 are
summarized in Scheme 4.
18F-Labeled thio acid [18F]1 afforded comparable high RCCs
(70–99%) of respective N-acyl sulfonamide model compounds
[
18F]3–[18F]6 as also determined by radio-TLC analysis repre-
senting the percentage of product present in the reaction
mixture. Thio acid [18F]2 gave comparable RCCs for compounds
[
18F]8 and [18F]9, whereas compounds [18F]7 and [18F]10 were
Radiolabeling of sulfonylazide-decorated model tetra-
peptide 14 with thio acids [18F]1 and [18F]2 was accomplished
using established one-pot/three component sulfo-click synthesis
procedure to give 18F-labeled peptides [18F]15 and [18F]16 within
a 30 min reaction time in moderate decay-corrected radio-
chemical yields of 25 and 20%, respectively, after HPLC purifica-
tion. The obtained radiochemical yields of 20–25% are somewhat
lower compared to the typically observed 30–50% radiochemical
yield using other acylation reactions with prominent 18F-labeled
prosthetic groups like [18F]SFB for peptide labeling with 18F.19
It is known from the literature that the best results for sulfo-
click chemistry are achieved when the reaction was performed
in DMF as the solvent in the presence of 2,6-lutidine as an
auxiliary base.14 However, the scope of the present work was to
demonstrate the principle feasibility of sulfo-click chemistry
with thio acids [18F]1 and [18F]2 as novel bioconjugation method
for peptide labeling with 18F under mild reaction conditions. No
further optimization of the reaction conditions was performed.
The synthesis of 18F-labeled peptides [18F]15 and [18F]16
using sulfo-click chemistry with in situ-formed thio acids [18F]1
and [18F]2 is shown in Scheme 6.
only obtained in 38% and 30%, respectively, according to their
radio-TLC-based RCC. A possible explanation could be the less
acidic character of [18F]2 compared to [18F]1 in combination
with the steric demand and/or more electron-rich character of
2,4,6-triisopropylbenzenesulfonyl azide and 4-methylbenzene–
sulfonyl as coupling partners in the sulfo-click reaction.
A second set of reactions tested sulfo-click chemistry with
18F-labeled thio acids [18F]1 and [18F]2 for the synthesis of drug-
like compounds. Replacement of the typical cyclooxygenase-2
(COX-2) sulfonamide pharmacophore by a sulfonylazide bio-
isostere provided a series of interesting compounds as analo-
gues of prominent COX-2 inhibitor and anti-inflammatory drug
celecoxib for sulfo-click chemistry. Interestingly, replacement
of sulfonamide group in celecoxib with a sulfonylazide motif
resulted in a switch of the COX-2/COX-1 selectivity profile with
the formation of a more pronounced COX-1 selective inhibitor.22
Synthesis of celecoxib-based sulfonylazide 11 as labeling pre-
cursor for sulfo-click chemistry with thio acids [18F]1 and [18F]2
was accomplished based on reported literature procedures.23
Reference compounds 12 and 13 were prepared by the
reaction of sulfonyl azide 11 with isolated non-radioactive thio
acids 1 and 2 (for full experimental details see ESI†). Reference
compounds were also tested in a COX enzyme inhibitory
assay24 demonstrating that celecoxib-based N-acyl sulfon-
amides 12 and 13 are potent and selective COX-2 inhibitors
(12: COX-2 IC50 = 0.3 mM, COX-1 4 100 mM; 13: COX-2 IC50 = 0.6 mM,
COX-1 4 100 mM).
In conclusion, we have demonstrated the first example of sulfo-
click chemistry with 18F-labeled thio acids and corresponding
Sulfo-click chemistry with thio acids [18F]1 and [18F]2
afforded 18F-labeled N-acyl sulfonamides [18F]12 and [18F]13
in comparable radiochemical conversions of 67 and 55%,
respectively, as determined by radio-TLC analysis representing
the percentage of desired products present in the reaction
mixture (Scheme 5).
Scheme 6 Synthesis of 18F-labeled peptides [18F]15 and [18F]16 using
sulfo-click chemistry as bioconjugation method.
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Conflicts of interest
There are no conflicts to declare.
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