1,5-Disubstituted 1,2,3-Triazoles as Amide Bond Isosteres Yield Novel Tumor-Targeting Minigastrin Analogs

Article abstract of DOI:10.1021/acsmedchemlett.0c00636

1,5-Disubstituted 1,2,3-triazoles (1,5-Tz) are considered bioisosteres of cis-amide bonds. However, their use for enhancing the pharmacological properties of peptides or proteins is not yet well established. Aiming to illustrate their utility, we chose the peptide conjugate [Nle15]MG11 (DOTA-dGlu-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2) as a model compound since it is known that the cholecystokinin-2 receptor (CCK2R) is able to accommodate turn conformations. Analogs of [Nle15]MG11 incorporating 1,5-Tz in the backbone were synthesized and radiolabeled with lutetium-177, and their pharmacological properties (cell internalization, receptor binding affinity and specificity, plasma stability, and biodistribution) were evaluated and compared with [Nle15]MG11 as well as their previously reported analogs bearing 1,4-disubstituted 1,2,3-triazoles. Our investigations led to the discovery of novel triazole-modified analogs of [Nle15]MG11 with nanomolar CCK2R-binding affinity and 2-fold increased tumor uptake. This study illustrates that substitution of amides by 1,5-disubstituted 1,2,3-triazoles is an effective strategy to enhance the pharmacological properties of biologically active peptides.

Full text of DOI:10.1021/acsmedchemlett.0c00636

pubs.acs.org/acsmedchemlett
Letter
1,5-Disubstituted 1,2,3-Triazoles as Amide Bond Isosteres Yield
Novel Tumor-Targeting Minigastrin Analogs
Nathalie M. Grob, Roger Schibli, Martin Béhé, Ibai E. Valverde,* and Thomas L. Mindt*
Cite This: ACS Med. Chem. Lett. 2021, 12, 585−592
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ABSTRACT: 1,5-Disubstituted 1,2,3-triazoles (1,5-Tz) are considered bioisosteres of cis-amide bonds. However, their use for
enhancing the pharmacological properties of peptides or proteins is not yet well established. Aiming to illustrate their utility, we
chose the peptide conjugate [Nle15]MG11 (DOTA-DGlu-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2) as a model compound since it is
known that the cholecystokinin-2 receptor (CCK2R) is able to accommodate turn conformations. Analogs of [Nle15]MG11
incorporating 1,5-Tz in the backbone were synthesized and radiolabeled with lutetium-177, and their pharmacological properties
(cell internalization, receptor binding affinity and specificity, plasma stability, and biodistribution) were evaluated and compared
with [Nle15]MG11 as well as their previously reported analogs bearing 1,4-disubstituted 1,2,3-triazoles. Our investigations led to the
discovery of novel triazole-modified analogs of [Nle15]MG11 with nanomolar CCK2R-binding affinity and 2-fold increased tumor
uptake. This study illustrates that substitution of amides by 1,5-disubstituted 1,2,3-triazoles is an effective strategy to enhance the
pharmacological properties of biologically active peptides.
KEYWORDS: 1,2,3-Triazoles, peptidomimetics, structure−activity relationships, radiopharmaceuticals, tumor targeting, cancer
ive-membered N-heterocycles have attracted attention as
amide bond mimics for the past 20 years. Due to the
dipolar moment, are capable of mimicking both trans- and cis-
amide bonds, depending on their 1,4- or 1,5-substitution
pattern, respectively, while being synthetically accessible
(Figure 2).13
The practicality of 1,2,3-triazoles as peptidomimetics has
been greatly enhanced by the discovery of the copper(I)-
catalyzed azide−alkyne cycloaddition (CuAAC), which
selectively gives access to 1,4-disubstituted 1,2,3-triazoles
(1,4-Tz).14,15 Since then, 1,4-Tz have been quickly adopted
as bioisosteres of amide bonds in peptidomimetics to modify
the pharmacological properties of peptides or proteins.16−20
The systematic replacement of amide bonds by 1,4-Tz (termed
F
similar size, polarity, planarity, and their ability to create
hydrogen bonds, imidazoles, oxa(dia)zoles, tetrazoles, and
triazoles, among others, have been used to replace amide
bonds in order to improve the pharmacological characteristics
of biologically active compounds.1 These replacements are
performed to overcome the inherently low proteolytic stability
of amides or to probe binding conformations since hetero-
cycles can mimic conformationally locked amide bonds.2−6
Despite the large number of examples illustrating the potential
of replacing amide bonds with small heterocycles in drug-like
compounds with low molecular weight,5,6 the use of this
approach in peptides is not very common yet. Disubstituted
tetrazoles as well as 1,2,4- and 1,2,3-triazoles are among the
heterocycles that allow a precise replacement of an amide bond
(Figure 1).7−9 Triazole synthesis can be performed in solution
or on solid support, the latter making their use particularly
appealing for peptide chemists.10−12 In this context, 1,2,3-
triazoles (Tz) are heterocycles that, while having a similar
Received: November 30, 2020
Accepted: March 8, 2021
Published: March 16, 2021
https://doi.org/10.1021/acsmedchemlett.0c00636
© 2021 American Chemical Society
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Figure 1. Mimetics of amide bonds based on N-heterocycles.
in nuclear medicine for targeting a variety of tumors such as
medullary thyroid carcinoma and small cell lung cancer. We
have previously shown that single or multiple 1,4-Tz can be
incorporated between residues DGlu10 to Trp14 of compound
1. In the majority of the examples, the nanomolar affinity of the
resulting peptidomimetics toward the target receptor is
preserved and an increased proteolytic stability was observed.
In addition, several linear and cyclic peptides and peptidomi-
metics mimicking β-turns have shown high affinities toward the
CCK2R.36−38 On the basis of these reports, we set out to study
the influence of amide-to-1,5-Tz switches on the CCK2R-
binding properties of compound 1 in positions likely to be
involved in β-turn conformation. Triazole-containing analogs
of compound 1 were synthesized, radiolabeled with [177Lu]-
Lu3+, and evaluated in vitro (cell internalization, receptor
affinity and specificity, metabolic stability) and in vivo
(biodistribution in mice bearing CCK2R-positive xenografts).
With the aim of identifying 1,5-Tz-modified derivatives of
compound 1 with retained tumor-targeting properties, bonds
between Gly13-Trp14 (5), Tyr12-Gly13 (6), Ala11-Tyr12 (7) were
substituted with a 1,5-Tz moiety. Modifications between
Trp14-Nle15 and DGlu10-Ala11 were discarded since these
bonds are distant from the suspected location of the
turn.29,30 The heterocycles were inserted in the peptide
sequence following the strategy exemplified for conjugate 7
in Scheme 1. Azido benzyl esters of amino acids were
synthesized via diazo transfer using 1H-imidazole-1-sulfonyl
azide.39 Amino alkynes were synthesized from commercially
available L-amino acids as previously described.24,27 Despite
our efforts, we were not able to obtain the excellent optical
purities that we observed in the past.24,27 As a result, amino
alkyne mimics of Fmoc-Ala-OH and Fmoc-Tyr(tBu)-OH were
isolated in a L:D ratio of 66:34 and 90:10, respectively (see
Supporting Information).24,27 Partial racemization during the
Figure 2. Comparison between trans-amide and cis-amide bonds and
1,4- (A) and 1,5-disubstituted 1,2,3-triazoles (B).
a triazole scan) has been used as a tool to probe the
contribution of the backbone to the stability and the activity of
biologically relevant peptides similar to an N-methylamide
scan.21 So far, this approach has been applied to enkephalin,22
angiotensin,23 bombesin,24−26 neurotensin,27,28 and minigas-
trin.29,30 Examples of peptidomimetics based on 1,5-
disubstituted 1,2,3-triazoles (1,5-Tz) are scarce. 1,5-Tz,
obtained by the ruthenium-catalyzed azide−alkyne cyclo-
addition (RuAAC), are useful to probe the bioactive
conformation of peptides, since they can mimic constrained
structures such as β-turns or hairpins,17,31,32 and their use as
valuable amide bioisosteres in peptide backbones has been
demonstrated both theoretically33,34 and experimentally.11,17,35
We have recently reported the triazole scan of [Nle15]MG11
(compound 1, Figure 3) labeled with lutetium-177
(
177Lu).29,30 This conjugate consists of a peptide with high
affinity for the CCK2R coupled to the macrocyclic chelator
DOTA to enable labeling with radiometals and shows potential
Figure 3. Structure of [Nle15]MG11 with numbering of the amino acid residues.
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Letter
Scheme 1. Synthesis of 1,5-Tz-containing peptidomimetic DOTA conjugate 7.
Seyferth−Gilbert homologation has been observed in the
past,29 and alternative procedures have since been proposed for
the synthesis of optically pure amino alkynes.34 Despite the
inconvenience, amino alkynes were used in the peptidomimetic
syntheses as a mixture of enantiomers. Cp*RuCl(PPh3)2 and
reaction temperatures of 60 °C 40,41 afforded the cycloaddition
products in solution within 2−8 h (see Supporting
Information). RuAAC was followed by deprotection of the
carboxylic acid by hydrogenation using Pd/C as catalyst. The
obtained pseudodipeptide building blocks were coupled to the
growing peptide sequence on solid support using HATU as a
coupling reagent. The remaining amino acids were coupled
using standard Fmoc/tBu solid-phase peptide synthesis
(SPPS) until completion of the DOTA-peptide conjugates
(Scheme 1). After cleavage from solid support, deprotection by
TFA, and purification by RP-HPLC, the three DOTA-
substituted peptidomimetics 5−7 were obtained in satisfactory
yields (30−50%) and characterized by analytical RP-HPLC
and HR-MS (see Supporting Information). In the case of the
syntheses of 6 and 7, as was expected based on the
enantiomeric purity of the α-substituted amino alkynes, two
diastereoisomers were found in the same ratios as the L:D ratios
of the aminoalkynes (see Supporting Information). The two
diastereoisomers were separated by RP-HPLC, and only the
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Table 1. Summary of in Vitro Results of Compounds [177Lu]Lu-1−7
internalization at 4 h
(% a. d.)
IC50 (mean),
(95% confidence interval) (nM)
half-life
(h)
b
compd
sequence
a
a
a
a
[
[
[
[
[
[
[
177Lu]Lu-1
177Lu]Lu-2
177Lu]Lu-3
177Lu]Lu-4
177Lu]Lu-5
177Lu]Lu-6
177Lu]Lu-7
[
[
[
[
[
[
[
177Lu]Lu-DOTA-DGlu-Ala-Tyr- Gly-Trp-Nle-Asp-Phe-NH2
32.2 3.2
41.7 3.9
54.3 5.1
29.6 2.7
1.5 0.9
15.4 (11.0−21.1)
15.6 (12.3−19.7)
1.7 (1.3−2.3)
20.9 (17.0−25.7)
447.7 (352.8−568.1)
10.5 (8.1−13.7)
3.9
3.8
2.6
51.4
91.1
2.2
177Lu]Lu-DOTA-DGlu-Ala-Tyr-Glyψ[1,4-Tz]Trp-Nle-Asp-Phe-NH2
177Lu]Lu-DOTA-DGlu-Ala-Tyrψ[1,4-Tz]Gly-Trp-Nle-Asp-Phe-NH2
177Lu]Lu-DOTA-DGlu-Alaψ[1,4-Tz]Tyr-Gly-Trp-Nle-Asp-Phe-NH2
177Lu]Lu-DOTA-DGlu-Ala-Tyr-Glyψ[1,5-Tz]Trp-Nle-Asp-Phe-NH2
177Lu]Lu-DOTA-DGlu-Ala-Tyrψ[1,5-Tz]Gly-Trp-Nle-Asp-Phe-NH2
177Lu]Lu-DOTA-DGlu-Alaψ[1,5-Tz]Tyr-Gly-Trp-Nle-Asp-Phe-NH2
50.9 4.1
30.3 1.4
64.8 (51.7−81.3)
b
6.7
Data of reference compound [177Lu]Lu-1 and 1,4-Tz analogs [177Lu]Lu-2−4 are reproduced for comparison.29 Competition experiments were
a
performed with nonradioactive analogs of the compounds 1−7 labeled with 175Lu.
major compounds, which were assumed to have the desired
stereochemistry, were later examined in in vitro and in vivo
assays (Scheme 1).
The affinity (IC50, Table 1) of the novel triazolopeptides
toward the CCK2R was determined by a competition-binding
assay. Briefly, the established CCK2R-ligand [177Lu]Lu-PP-
F11N (DOTA-(DGlu)6-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2)43
was added to A431-CCK2R cells and displaced by increasing
concentrations of nonradioactive 175Lu-labeled peptidomimet-
ics 5−7 for 1 h. The cells were washed and lysed to determine
the bound fraction of [177Lu]Lu-PP-F11N as a function of the
concentration of added competitors (see Table 1 and the
Supporting Information). As anticipated from the internal-
ization assays, compound [175Lu]Lu-5 with the lowest cell
internalization had the lowest affinity for its target (IC50 ∼ 450
nM). Compounds [175Lu]Lu-6 and [175Lu]Lu-7 showed IC50
values of 11 nM and 65 nM, respectively, with only conjugate
Compounds 5−7 were radiolabeled with [177Lu]LuCl3 to
obtain radioconjugates [177Lu]Lu-5−7 (Table 1) in high
radiochemical yields and purities of ≥95%, with specific molar
activities between 25 and 50 MBq/nmol (not optimized).29,30
Representative chromatograms of quality controls by γ-HPLC
can be found in the Supporting Information.
Receptor-mediated internalization of the radioconjugates
was evaluated using A431 cells stably transfected with the
CCK2R (A431-CCK2R cells)42 and is expressed as the percent
of added radioactivity per 0.85 million cells (Table 1 and
Figure 4). Blocking experiments (performed by addition of a
[
175Lu]Lu-6 approaching the affinity of the reference
compound [175Lu]Lu-1 (15 nM). Unlike compound
177Lu]Lu-2, with a 1,4-Tz between Gly13-Trp14 mimicking a
[
trans-amide, compound [175Lu]Lu-5, with a 1,5-Tz that mimics
a cis-amide bond, did not show high nanomolar affinity toward
the receptor. This result could indicate that the backbone of
the peptide requires a linear (all-trans) conformation from the
C-terminus until Gly13. This observation confirms our previous
hypothesis that Gly13 separates the C-terminal part of
[Nle15]MG11, which is buried in the receptor cavity, from
the N-terminal part that is pointing toward the extracellular
domain of the receptor.29 In the same previous study, we
observed an increased interaction between the CCK2R and
1,4-Tz compound [175Lu]Lu-3 likely as the result of an
additional cation−π interaction between the triazole ring and
Arg356 of the receptor.29 Similarly, the 1,5-Tz isomer
[
175Lu]Lu-6 showed an improved affinity toward the CCK2R
in comparison to the reference compound [175Lu]Lu-1 (10 nM
vs 15 nM). Even though the improvement of affinity of
Figure 4. Specific internalization kinetics of conjugates [177Lu]Lu-1 to
7 in A431-CCK2R cells at 37 °C. Data of previously reported
compounds [177Lu]Lu-1−429 are included for comparison (n = 3 in
triplicates).
[
175Lu]Lu-6 was not as pronounced as in the case of the 1,4-Tz
analog [175Lu]Lu-3, the presence of an aromatic ring between
Tyr12 and Gly13 indeed seems beneficial for the receptor
interaction. Despite showing good cell internalization,
[
175Lu]Lu-7 displayed a decreased affinity toward the
5000-fold excess of minigastrin) decreased the internalization
of radiolabeled compounds to less than 1% in all cases, hence
demonstrating specific receptor-mediated interaction (see
Supporting Information). With the exception of compound
CCK2R in comparison with its all-amide counterpart and its
1,4-Tz isomer (65 nM vs 15 nM and 21 nM, respectively). We
conclude that the position between Ala11-Tyr12 does not play a
significant role in receptor binding due to its tolerance toward
modifications.
[
177Lu]Lu-5, the other 1,5-Tz analogs of compound 1 were
able to bind and internalize into receptor-positive A431-
CCK2R cells. Conjugates [177Lu]Lu-6 and [177Lu]Lu-7
reached similar or higher cell internalizations than reference
compound 1 (51−30% vs 32%, respectively), and the
internalization rates and kinetics resembled closely the ones
of their 1,4-substituted counterparts [177Lu]Lu-3 and -4,
respectively (54% and 30%, respectively, Table 1, Figure 4).
Next, we studied the proteolytic degradation of compounds
[
177Lu]Lu-5-7 in vitro to determine if the use of 1,5-Tz benefits
the stability of this regulatory peptide. The metabolic half-lives
(t1/2) of the conjugates were determined by incubation of the
radiolabeled peptides in human blood plasma followed by
quantification of their metabolites by γ-HPLC over 24 h
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(Table 1). The position of the amide-to-triazole switch
utilizing 1,5-Tz had distinct effects on the metabolic stability
of the resulting peptidomimetic conjugates in comparison to
the previously studied 1,4-Tz-substituted minigastrins.24,26,27
Peptidomimetics [177Lu]Lu-5 and [177Lu]Lu-7 showed an
increased stability in comparison to reference compound
us to the conclusion that the contribution of differently
substituted triazoles to the stability of a peptide may vary and
does not follow a general pattern.
In vivo studies with mice bearing CCK2R-positive tumor
xenografts were performed to investigate whether peptidomi-
metics [177Lu]Lu-6 and 7 retained the biodistribution profile of
reference compound [177Lu]Lu-1 (see Supporting Information
for details). Compound 5 was discarded for in vivo studies due
to its reduced cell internalization and low affinity toward the
CCK2R. Uptake of radioactivity in organs is expressed as % of
injected dose per gram of organ or tissue (% ID/g). Female
CD1 nu/nu mice were xenografted with A431-CCK2R cells
and randomly assigned to groups of four mice. The results of
the experiment are summarized in Figure 6 (see Supporting
Information for complete data sets). At 4 h postinjection, all
compounds showed typical biodistribution profiles of radio-
labeled peptides with a fast clearance from the blood and
unspecific uptake in the kidneys caused by renal excretion
(Figure 6). Specific receptor-mediated uptake was demon-
strated by co-injection with a 6000-fold excess of minigastrin
(blocking), which led to a significant decrease of the uptake in
the receptor-positive tumor and organs, e.g., the stomach (see
Supporting Information). Compound [177Lu]Lu-6 showed the
highest tumor uptake of the investigated 1,5-Tz peptidomi-
metics (3.9% ID/g). Similarly, a more than 2-fold increased
uptake in the tumor was also previously observed for the
analogous 1,4-Tz isomer [177Lu]Lu-3. This is consistent with
their improved CCK2R affinity and cell internalization in
comparison with reference compound [177Lu]Lu-1 in vitro
(Table 1). Tumor uptake of compound [177Lu]Lu-7 was
inferior to its 1,4-Tz counterpart [177Lu]Lu-4 (1.7% ID/g vs
0.8% ID/g,). The difference in the tumor uptake of
compounds 7 and 4 could be due to a decreased receptor
affinity of compound 7 in comparison with compound 1 or 4
as well as to the superior proteolytic resistance of compound
[
[
177Lu]Lu-1, whereas the t1/2 was decreased for compound
177Lu]Lu-6 (2.2 h vs 3.9 h). It appears that while the insertion
of a Tz heterocycle can improve the affinity of the resulting
peptidomimetic toward its receptor, it can also increase its
susceptibility toward degradation by proteases. The degrada-
tion kinetics of 1,4- vs 1,5-Tz peptidomimetics of MG11
differed not only depending on the position of the heterocycle
in the sequence but also on the substitution pattern of the
triazole (Figure 5). Degradation of compound [177Lu]Lu-5 was
Figure 5. Degradation kinetics of 177Lu-labeled peptide conjugates
after incubation in human blood plasma for 24 h at 37 °C. Data points
show the mean SD, n = 2−3, for values of t1/2; see Table 1.
[
177Lu]Lu-4.
We herein report the use of 1,5-Tz heterocycles as
metabolically stable mimics of cis-amide bonds in biologically
active linear peptides. 1,5-Tz were placed in the backbone of
the radiolabeled peptide [Nle15]MG11 in the vicinity of a
proposed β-turn. The obtained peptidomimetics were
compared side-by-side with the all-amide bond reference
compound [Nle15]MG11 as well as with previously studied
1,4-Tz analogs, which adopt trans-conformations for the
probed amide bonds.
significantly slower than its 1,4-Tz counterpart [177Lu]Lu-2
(t1/2 = 3.8 h vs 91.1 h). The behavior was reversed in the case
of conjugate [177Lu]Lu-7, which was metabolized faster than
1,4-Tz compound [177Lu]Lu-4 (t1/2 = 51.4 h vs 6.7 h). Finally,
degradation kinetics of compounds [177Lu]Lu-3 and
[
177Lu]Lu-6 were comparable (Figure 5). These results lead
Figure 6. Comparison of uptakes of compounds [177Lu]Lu-1, -3, -4, -6, and -7 in selected organs of mice xenografted with A431-CCK2R positive
cells at 4 h p.i. (n = 4). Data of compounds [177Lu]Lu-1, -3, and -4 are reproduced for comparison. Superscripted pos indicates receptor-positive
tissue. Data points show the mean SD, n = 4, for values of all collected tissues and tumor-to-tissue ratios; see the Supporting Information.
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In comparison to the reference compound, some of the
novel 1,5-Tz peptidomimetics showed improved biological
properties such as enhanced plasma stability and affinity
toward the CCKR2 in vitro or increased tumor uptake in vivo,
thus providing another example for the successful application
of 1,5-Tz as bioisosteres of amide bonds in bioactive peptides.
Furthermore, the direct comparison of 1,4-Tz versus 1,5-Tz
containing peptidomimetics revealed that a general prediction
about the effect of the substitution pattern of the heterocycle
on the biological properties of a peptide requires further
investigations.
We encourage medicinal and peptide chemists who study
1,4-Tz-based peptidomimetics to consider also the use of 1,5-
Tz as both heterocycles show high potential to enhance the
pharmacological properties of bioactive peptides and thus
expand the peptidomimetic toolbox for drug discovery.
in vitro and in vivo data. M.B. and N.M.G. assisted with the in
vivo experiments. R.S. contributed to interpretations of the
results. I.E.V., N.M.G., and T.L.M. wrote and revised the
manuscript. All authors have given approval to the final version
of the manuscript.
Funding
This work was supported by the Swiss National Science
Foundation (Grant 200021-157076 to T.L.M.) and the
́
́
Conseil Regional Bourgogne Franche-Comte (Grant 2018Y-
07062 to I.E.V.).
Notes
The authors declare the following competing financial
interest(s): R.S. and M.B. are inventors of patent
WO201567473. T.L.M., M.B., R.S., I.E.V., and N.M.G. have
submitted patent application WO 2019/057445 A1 as
inventors.
ASSOCIATED CONTENT
* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acsmedchemlett.0c00636.
■
ACKNOWLEDGMENTS
sı
■
This work is part of the project “Pharmacoimagerie et Agents
Théranostiques” supported by the Université de Bourgogne
and Conseil Régional de Bourgogne through the Plan d’Action
Régional pour l’Innovation (PARI), the Région Bourgogne
Franche-Comté through the ANER program, and the Euro-
pean Union through the PO FEDER-FSE Bourgogne 2014/
2020 programs. GDR CNRS “Agents d’Imagerie Moléculaire”
2037 is thanked for its interest in this research. We thank the
Synthesis and full characterization of peptides 5−7 and
[
177Lu]5−7 (UV- and γ-HPLC, mass spectrometric
data) and detailed results of in vitro (internalization,
competition binding, metabolic stability) and in vivo
experiments (PDF)
̀
“Plateforme d’Analyse Chimique et de Synthese Moléculaire
AUTHOR INFORMATION
Corresponding Authors
■
de l’Université de Bourgogne” (PACSMUB, http://www.
wpcm.fr) for access to spectroscopy instrumentation. I.E.V.
thanks Prof. Anthony Romieu and Dr. Adrien Normand for
Ibai E. Valverde − Institut de Chimie Moléculaire de
l’Université de Bourgogne, UMR CNRS 6302, Université de
Bourgogne Franche-Comté, 21000 Dijon, France;
orcid.org/0000-0002-5802-4131; Phone: +33 380 39 90
48; Email: ibai.valverde@u-bourgogne.fr
Thomas L. Mindt − Ludwig Boltzmann Institute Applied
Diagnostics, General Hospital of Vienna, 1090 Vienna,
Austria; Department of Inorganic Chemistry, Faculty of
Chemistry, University of Vienna, 1090 Vienna, Austria;
Department of Biomedical Imaging and Image Guided
Therapy, Medical University of Vienna, 1090 Vienna,
Austria; Phone: +43 14040025350;
̂
scientific discussions, Dr. Jerome Bayardon for scientific
discussions and technical support with chiral HPLC, and Dr.
Quentin Bonnin and Marie-José Penouilh for HR-MS
(Université de Bourgogne, Dijon, France). DOTA-tris(tBu)
ester was a generous gift from Chematech (Dijon, France).
A431-CCK2R cells were a kind gift of Dr. Luigi Aloj
(University of Cambridge, U.K.). We thank Stefan Imobersteg
(PSI, Villigen, Switzerland) for animal care.
ABBREVIATIONS
■
ψ[Tz], a 1,2,3-triazole substituting an amide bond; BSA,
bovine serum albumin; CCK2R, cholecystokinin-2 receptor;
DOTA, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic
acid; HATU, O-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetrame-
thyluronium hexafluorophosphate; RP-HPLC, reverse-phase
high-performance liquid chromatography; HR-MS, high-
resolution mass spectrometry; ID, injected dose; MG,
minigastrin; Nle, norleucine; SPPS, solid-phase peptide
synthesis; TFA, trifluoroacetic acid
Email: thomas.mindt@lbiad.lbg.ac.at
Authors
Nathalie M. Grob − Department of Chemistry and Applied
Biosciences, ETH Zurich, 8093 Zurich, Switzerland
̈
Roger Schibli − Department of Chemistry and Applied
Biosciences, ETH Zurich, 8093 Zurich, Switzerland; Center
̈
for Radiopharmaceutical Sciences, Division of Biology and
Chemistry, Paul Scherrer Institute, 5232 Villigen, Switzerland
Martin Béhé − Center for Radiopharmaceutical Sciences,
Division of Biology and Chemistry, Paul Scherrer Institute,
5232 Villigen, Switzerland; orcid.org/0000-0002-1110-
2665
REFERENCES
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Complete contact information is available at:
https://pubs.acs.org/10.1021/acsmedchemlett.0c00636
Author Contributions
I.E.V., T.L.M., and N.M.G. designed the compounds and
planed the in vitro and in vivo experiments together with M.B.,
and I.E.V. performed the chemical synthesis of the compounds.
N.M.G conducted radiolabeling, in vitro assays, and evaluated
590
https://doi.org/10.1021/acsmedchemlett.0c00636
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Letter
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