Solid-phase synthesis of smac peptidomimetics incorporating triazoloprolines and biarylalanines

Article abstract of DOI:10.1021/co200078u

Apoptotic induction mechanisms are of crucial importance for the general homeostasis of multicellular organisms. In cancer the apoptotic pathways are downregulated, which, at least partly, is due to an abundance of inhibitors of apoptosis proteins (IAPs) that block the apoptotic cascade by deactivating proteolytic caspases. The Smac protein has an antagonistic effect on IAPs, thus providing structural clues for the synthesis of new pro-apoptotic compounds. Herein, we report a solid-phase approach for the synthesis of Smac-derived tetrapeptide libraries. On the basis of a common (N-Me)AVPF sequence, peptides incorporating triazoloprolines and biarylalanines were synthesized by means of Cu(I)-catalyzed azide-alkyne cycloaddition and Pd-catalyzed Suzuki cross-coupling reactions. Solid-phase procedures were optimized to high efficiency, thus accessing all products in excellent crude purities and yields (both typically above 90%). The peptides were subjected to biological evaluation in a live/dead cellular assay which revealed that structural decorations on the AVPF sequence indeed are highly important for cytotoxicity toward HeLa cells.

Full text of DOI:10.1021/co200078u

RESEARCH ARTICLE
pubs.acs.org/acscombsci
Solid-Phase Synthesis of Smac Peptidomimetics Incorporating
Triazoloprolines and Biarylalanines
Sebastian T. Le Quement,† Mette Ishoey,† Mette T. Petersen,† Jacob Thastrup,‡ Grith Hagel,‡ and
Thomas E. Nielsen*,†
†Department of Chemistry, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark
‡2CureX, Department of Surgery, K, Bispebjerg Hospital, Bispebjerg Bakke 23, DK-2400 Copenhagen NV, Denmark
S Supporting Information
b
ABSTRACT: Apoptotic induction mechanisms are of crucial
importance for the general homeostasis of multicellular organ-
isms. In cancer the apoptotic pathways are downregulated,
which, at least partly, is due to an abundance of inhibitors of
apoptosis proteins (IAPs) that block the apoptotic cascade by
deactivating proteolytic caspases. The Smac protein has an
antagonistic effect on IAPs, thus providing structural clues for the synthesis of new pro-apoptotic compounds. Herein, we report a
solid-phase approach for the synthesis of Smac-derived tetrapeptide libraries. On the basis of a common (N-Me)AVPF sequence,
peptides incorporating triazoloprolines and biarylalanines were synthesized by means of Cu(I)-catalyzed azideÀalkyne cycloaddi-
tion and Pd-catalyzed Suzuki cross-coupling reactions. Solid-phase procedures were optimized to high efficiency, thus accessing all
products in excellent crude purities and yields (both typically above 90%). The peptides were subjected to biological evaluation in a
live/dead cellular assay which revealed that structural decorations on the AVPF sequence indeed are highly important for
cytotoxicity toward HeLa cells.
KEYWORDS: solid-phase synthesis, IAP inhibition, Smac, peptidomimetics, Cu(I)-catalyzed azideÀalkyne cycloaddition, Suzuki
cross-coupling
’ INTRODUCTION
The apoptotic circuitry ensures the general homeostasis of
multicellular organisms via control of cellular proliferation.1,2
This circuitry is subject to tight regulation through numerous
cellular signaling cascades and feed-back mechanisms, where
cysteine-aspartic acid proteases (caspases) play a predominant
role. In short, two main pathways for controlled cell destruction
are generally accepted: an intrinsic (mitochondria-mediated)
which typically acts as response to intracellular damage; and an
extrinsic which involves the membrane-bound death-receptor
ligands and responds to extracellular stimuli such as chemother-
apeutic agents.1 Both pathways converge at the level of activation
of caspases 3 and 7 and ultimately lead to cell death.3 A variety of
caspases are implicated in these cellular cascades where they
function as degraders of the cellular proteome. One of the
hallmarks of cancer is the evasion of natural apoptotic signals,
which enable malignant cells to proliferate, even when subjected
to radio/chemotherapies, that heavily rely on the cells to engage
in their own cellular destruction.4 This resistance to chemother-
apeutic agents is partly mediated by the inhibitors of apoptosis
proteins (IAPs) which suppress the activity of caspases through
various mechanisms.2,4À8 IAPs are a group of proteins currently
Figure 1. Smac-derived AVPI peptide and selected examples of Smac
comprising 8 members (XIAP, cIAP-1 and -2, ML-IAP, NAIP,
ILP-2, survivin, and Apollon) which are characterized by one or
more baculoviral IAP repeats (BIR) but can also incorporate
other important domains such as the CARD or RING domains.9
The mechanism of action of IAPs is still subject to investigation,
mimetics.
Received: April 29, 2011
Revised:
Published: September 12, 2011
September 5, 2011
r
2011 American Chemical Society
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Figure 2. Synthetic strategy for the construction of triazoloproline- and biarylalanine-containing libraries 8À11.
Scheme 1. Synthesis of Fmoc-Protected Azidoproline
Building Blocks 16 and 21a
but it is generally accepted that XIAP plays a predominant role in
the intrinsic pathway by inhibition of caspase-9 and the down-
stream executioner caspases 3 and 7.6 cIAP-1 and -2, on the other
hand, have been shown to modulate a range of mechanisms, such
asE3 ubiquitin ligaseactivity.4,10,11 cIAPs are thus implicated in the
death-receptor pathway, but might also function through binding
and sequestering of the second mitochondria-derived activator
of caspase (Smac) protein,12,13 which is an endogenous inhibitor
of XIAP, and thereby prevent Smac’s pro-apoptotic activity.14
In the past decade, small molecule antagonistic modulation of
IAPs aiming at the re-establishment of normal apoptotic levels
has emerged as a promising anticancer strategy.9,14À16 The
discovery of the mode of action of Smac,17À19 has provided an
entry point for synthetic endeavors in the hunt for such mol-
ecules.20,21 Smac is released in the cytosol upon permeabilization
of the mitochondrial membrane and antagonizes IAP function by
virtue of its N-4 terminal (Smac-derived AVPI peptide 1, Figure 1),
both as a monomer and dimer.9 A multitude of highly potent Smac
mimetics, mimicking the action of the N-4 terminal residues of
the Smac protein have been reported, and at least five IAP-
antagonists are currently undergoing clinical trials.14 The molec-
ular scaffolds typically rely on structural modifications of the
AVPI Smacpeptidebutnovelscaffolds have also been reported.22À28
SAR studies and investigations of the binding groove of XIAP
have led to the proposal of a general pharmacophore model of
the binding interaction of Smac with XIAP.15 This model notably
suggests the introduction of specific proline substituents and
replacement of isoleucine with larger hydrophobic groups to be
important structural elements for fine-tuning the biological activity.
Not surprisingly, a number of reported Smac peptidomimetics
incorporate various types of aromatic functionalities at these
positions (2 and 3, Figure 1).14,29 Other investigations, aiming at
improved bioavailability through enhanced cell permeability, have
focused on the design of constrained peptidomimetics (4, Figure 1),
where turn-mimicking bicyclic motifs have been incorporated in
replacement of the proline residue.30À35 Interestingly, a number
of highly potent polymeric structures mimicking dimeric Smac,
a Reagents and conditions: (a) Cs2CO3, MeI, DMF; (b) DPPA, DEAD,
PPh3, THF; (c) 1 M NaOH (aq), MeOH; (d) 50% TFA (CH2Cl2),
0 °C; (e) FmocCl, 10% Na2CO3 (aq), MeCN, À20 °C; (f) DIAD, PPh3,
THF, 0 °C; (g) NaN3, MeOH, 40 °C; (h) MsCl, Et3N, CH2Cl2, 0 °C; (i)
NaN3, DMSO, 80 °C; (j) LiOH (aq), THF.
thought to simultaneously bind several IAP BIR-domains, have
also been reported.14,29,36 A number of such bi- or trivalent
compounds are connected through aliphatic or aromatic bridging
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Scheme 2. Solid-Phase Synthesis of Solid-Supported Azidopeptides 26 and 27a
a Reagents and conditions: (a) Fmoc-Rink-OH, TBTU, NEM, DMF; (b) 20% Piperidine (DMF), (c) Fmoc-Phe-OH, TBTU, NEM, DMF; (d) 16 or 21,
TBTU, NEM, DMF; (e) Fmoc-Val-OH, TBTU, NEM, DMF; (f) Boc-(N-Me)Ala-OH, TBTU, NEM, DMF; (g) 95% TFA (aq).
Table 1. Optimization of Reaction Parameters for the CuAAC Reaction
entry
1-octyne (equiv)
CuI (equiv)
DIPEA (equiv)
NaAsc (equiv)
time (h)
purity of 8{1}a (%)
1
2
3
4
5
6
40
40
40
20
2
2
2
2
5
5
2
50
50
0
2
0
0
0
0
0
20
20
20
20
48
48
0
89
52
50
50
100
71
0
10
>95
a As indicated by RP-HPLC of the crude product (215 nm).
moieties installed on the pyrrolidine ring of the proline residue
(3, Figure 1).14
We have previously transferred the synthesis of a potent Smac
peptidomimetic incorporating an alkyl-substitution at the 3-posi-
tion of the proline residue (5, Figure 1) to the solid phase.37
Along these lines, we herein wish to present a further extension of
the strategy to the construction of solid-supported libraries of
Smac mimetics, based on a common (N-Me)AVPF motif, display-
ing diverse substitution patterns at the proline and phenylalanine
positions. Specifically, we have taken advantage of the Cu(I)-
catalyzed azideÀalkyne cycloaddition (CuAAC) and the Suzuki
cross-coupling reactions to introduce aromatic substituents in
the parent scaffold in the form of 3-substituted triazoloprolines
and biarylalanines, respectively (Figure 2).
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Table 2. Synthesis of Triazoloproline Libraries 8 and 9
entry
R
product, purity (%)a
1
hexyl-
propyl-
butyl-
8{1}, >95; 9{1}, >95
8{2}, >95; 9{2}, >95
8{3}, >95; 9{3}, >95
8{4}, >95; 9{4}, >95
8{5}, >95; 9{5}, >95
8{6}, >95; 9{6}, >95
8{7}, >95; 9{7}, >95
8{8}, 93; 9{8}, 93
2
3
4
pentyl-
5
cyclopropyl-
6
cyclopentyl-
7
cyclohexyl-
8
cyclohexylmethyl-
phenylthiomethyl-
phenoxymethyl-
benzoyloxymethyl-
phenyl-
9
8{9}, 94; 9{9}, 94
10
11
12
13
14
15
16
17
18
19
20
8{10}, 93; 9{10}, 93
8{11}, 94; 9{11}, 94
8{12}, 93; 9{12}, 93
8{13}, 94; 9{13}, >95
8{14}, >95; 9{14}, >95
8{15}, >95; 9{15}, >95
8{16}, >95; 9{16}, >95
8{17}, >95; 9{17}, >95
8{18}, >95; 9{18}, >95
8{19}, >95; 9{19}, >95
8{20}, >95b; 9{20}, 71c
4-phenoxyphenyl-
3,5-dimethoxyphenyl-
3,5-difluorophenyl-
3,4-dichlorophenyl-
6-methoxynaphtyl-
2-pyridyl-
3-thienyl-
2-carboxyethyl-
a As indicated by RP-HPLC of the crude product (215 nm). b Repetitive CuAAC reactions needed for full conversion. c Repetitive CuAAC reactions
failed to give full conversion.
’ RESULTS AND DISCUSSION
Scheme 3. Solid-Phase Synthesis of Solid-Supported Iodo-
peptides 30 and 31a
Synthesis of Triazoloproline Libraries 8 and 9. Originally
developed on solid support,38 CuAAC has now become a widely
used reaction in organic synthesis.39À41 The scope of this robust
and regioselective reaction keeps expanding, and 1,4-disubstitut-
ed-1,2,3-triazoles now find applications in many areas of organic
chemistry, including the synthesis of peptidomimetics and oligo-
nucleotides for biological investigation.40 Bearing in mind the
reported effect of introducing hydrophobic appendages in the
vicinity of the pyrrolidine ring, we speculated if we could take
advantage of the CuAAC reaction to access Smac mimetic
compounds of types 8 and 9 (Figure 2).42À45 Furthermore, we
were interested in correlating the biological activity with the
stereochemical orientation of the triazole substituent on the
pyrrolidine ring. Along these lines, we envisioned the synthesis of
two triazoloproline-containing libraries incorporating cis- and
trans-triazoloproline residues, respectively.
a Reagents and conditions: (a) Fmoc-Rink-OH, TBTU, NEM, DMF; (b)
20% Piperidine (DMF), (c) Fmoc-(3/4-I)Phe-OH, TBTU, NEM, DMF;
(d) Fmoc-Pro-OH, TBTU, NEM, DMF; (e) Fmoc-Val-OH, TBTU, NEM,
DMF; (f) Boc-(N-Me)Ala-OH, TBTU, NEM, DMF; (g) 95% TFA (aq).
The required building blocks for solid-phase synthesis, Fmoc-cis-
azidoproline 16 and Fmoc-trans-azidoproline 21, were synthesized
(Scheme 1) according to modified literature procedures.46À50
Starting with Boc-protected trans-hydroxyproline 12, the methyl
ester of cis-azidoproline 14 was accessed via reaction with methyl
iodide followed by direct azidation with DPPA under Mitsunobu
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Table 3. Synthesis of Biarylalanine Libraries 10 and 11
entry
Ar
product, purity(%)a
1
phenyl-
10{1}, >95; 11{1}, >95
10{2}, >95; 11{2}, 94
10{3}, 91; 11{3}, >95
10{4}, >95; 11{4}, >95
10{5}, >95; 11{5}, >95
10{6}, >95; 11{6}, >95
10{7}, >95; 11{7}, 90
10{8}, 94; 11{8}, >95
10{9}, 91; 11{9}, >95
10{10}, 82b; 11{10}, 92b
10{11}, >95; 11{11}, >95
10{12}, 88; 11{12}, 83
10{13}, 90; 11{13}, 90
10{14}, 0c; 11{14}, 0c
10{15}, 0c; 11{15}, 0c
10{16}, 0c; 11{16}, 0c
10{17}, 0d; 11{17}, 0d
10{18}, 0d; 11{18}, 0d
2
4-hydroxyphenyl-
4-methoxyphenyl-
4-fluorophenyl-
3-methoxyphenyl-
3-methylphenyl-
3-nitrophenyl-
3
4
5
6
7
8
2-methylphenyl-
2,4-dimethoxyphenyl-
2,6-dimethylphenyl-
3,4-dichlorophenyl-
3-thienyl-
9
10
11
12
13
14
15
16
17
18
2-benzothienyl-
4-pyridyl-
6-indolyl-
2-thienyl-
4-mercaptophenyl-
pentafluorophenyl-
a As indicated by RP-HPLC of crude product (215 nm). b The Suzuki cross-coupling was repeated once to achieve full conversion. c Complex reaction
mixture. d No conversion of starting material.
conditions. The ester was then subjected to alkaline hydrolysis to
access Boc-protected azidoproline 15. Acid-mediated Boc de-
protection, immediately followed by reaction with FmocCl,
yielded the desired Fmoc-cis-azidoproline 16 in 20% overall
yield. Fmoc-trans-azidoproline was also synthesized from trans-
hydroxyproline 12. First, trans-hydroxyproline 12 was lactonized
via intramolecular Mitsunobu esterification to yield bicyclic
compound 17. Then, the bicyclic lactone was ring-opened via
treatment with sodium azide in MeOH, affording esterified cis-
hydroxyproline 18. This cis-hydroxyproline was then converted
to Boc-protected trans-azidoproline 20 in a 3-step sequence,
involving mesylation, SN2-inversion with sodium azide in DMSO,
and hydrolysis with aqueous LiOH. Final conversion of the Boc
protecting group to the Fmoc protecting group afforded the
desired Fmoc-trans-azidoproline 21 in 43% overall yield.
Buildings blocks 16 and 21 were hereafter utilized in the
synthesis of support-bound azidoproline-containing tetrapep-
tides 26 and 27 (Scheme 2). Starting with the amino-function-
alized PEGA800 resin (0.4 mmol/g),51 the Rink linker was
attached using a TBTU-mediated amide coupling procedure.52
Consecutive deprotections of the Fmoc protecting group with
piperidine, and coupling of amino acids using the TBTU proto-
col gave resin-bound Boc-protected azides 26 and 27. For
product characterization purposes, both tetrapeptides were
released from the solid phase while simultaneously being Boc-
deprotected, by acidic treatment with 95% TFA (aq), and were
isolated in excellent crude purities (>95%) and yields (>90%).
With solid-supported azides 26 and 27 available, we set out to
construct the desired triazoloproline libraries. Initial experiments
with standard protocols for solid-phase CuAAC reactions, how-
ever, did not yield the desired triazoloproline-containing tetra-
peptides in satisfying purities.38,45,53À55 Consequently, a rapid
screen of reaction parameters was therefore performed on solid-
supported cis-azidopeptide 26. Reactants, reagent stoichiome-
tries and reaction times were investigated (Table 1). In contrast
to our previous work,54 the use of stoichiometric quantities of
sodium ascorbate (NaAsc) proved detrimental for the reaction
(Table 1, entry 1). Long reaction times and the use of DIPEA as
cosolvent, on the other hand, proved to be crucial for efficient
conversion of solid-supported substrate 26 and allowed isolation of
the desired triazoloproline 8{1} in excellent purity (Table 1, entry 6).
These optimized reaction conditions were then applied to the
synthesis of triazoloproline-containing libraries 8 and 9, by
reaction of azidopeptides 26 and 27 with a range of commercially
available and structurally diverse alkynes (Table 2). In general,
excellent purities were achieved after cleavage from the solid
phase. The reaction with 4-pentynoic acid, however, proved
problematic (Table 2, entry 20). The resin had to be subjected to
the CuAAC reaction conditions three times before full conver-
sion into cis-triazoloproline 8{20} was achieved. Surprisingly, in
the case of the related trans-azide 27, the reaction seemed to stop
at about 70% conversion and 3À4 repetitive CuAAC reactions
failed to provide the desired triazoloproline 9{20} in higher
purity.
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Chart 1. Screening Data from Live/Dead Cellular STS-Potentiation Assay on Triazoloproline Libraries 8 and 9
RESEARCH ARTICLE
With cis- and trans-triazoloproline libraries 8 and 9, both
containing 20 members, at hand, we set out to investigate the
synthesis of the projected biarylalanine libraries 10 and 11.
Synthesis of Biarylalanine Libraries 10 and 11. The palla-
dium-catalyzed Suzuki cross-coupling reaction has become a
powerful method in medicinal chemistry for the construction
of arylÀaryl bonds.56 Although scarcely abundant in natural
products, the biaryl motif now constitutes an important structur-
al element for small molecule drug discovery efforts. The Suzuki
cross-coupling is an attractive reaction for combinatorial and
parallel synthesis. Reaction conditions are generally mild and
compatible with most functional groups, and boronic acids are
readily available and stable to heat, air, and moisture. Inspired by
the potent and selective mimetics made by introducing hydro-
phobic moieties around the Ile residue of the Smac-derived AVPI
sequence, we decided to investigate the Suzuki cross-coupling for
the introduction of more lipophilic moieties in the IAP binding
groove. Specifically, we desired to examine if we could introduce
3- and 4-iodophenylalanines in our solid-phase synthesis scheme
and carry out subsequent Pd-catalyzed cross-coupling with a set
of arylboronic acids, thus creating biarylalanine libraries. First,
solid-supported aromatic iodides 30 and 31 were easily accessed
using commercially available N-protected amino acids and
standard solid-phase peptide synthesis protocols (Scheme 3).
Several approaches have been reported for Suzuki cross-
coupling reactions on solid support.57À61 After extensive opti-
mization, our preferred method for Suzuki cross-coupling reac-
tions on the PEGA resin involves the use of Pd(dppf)Cl2/K3PO4
in an aqueous mixture of t-BuOH and toluene at room tempera-
ture. Thus, solid-supported aromatic iodides 30 and 31 were
subjected to Suzuki cross-coupling with a range of aromatic
boronic acids incorporating various electron donating and with-
drawing substituents (Table 3). Notably, some boronic acids,
such as pentafluorophenylboronic acid and mercaptophenyl-
boronic, did not undergo cross-coupling reactions and only aryl
iodides 32 and 33 could be recovered after cleavage from the
solid phase (Table 3, entries 17 and 18). Other coupling partners
resulted in very complex reaction mixtures (Table 3, entries
14À16). Despite these unsuccessful experiments, all other selected
arylboronic acids smoothly underwent the desired cross-coupling
(Table 3, entries 1À13) in good to excellent purities (88 to >95%),
thus generating the projected 3- and 4-biarylalanine libraries 10 and 11.
Biological Evaluation of Smac Mimetics. The resulting four
Smac mimetic libraries were subjected to biological evaluation.
These were first tested for potentiation of the known pro-
apoptotic agent staurosporine (STS) in a live/dead fluorometric
assay. The assay was conducted on HeLa cells and the results are
depicted as a ratio of dead to live cells normalized to treatment
with STS alone (100 nM). All peptides were administered at a
30/60 μM concentration along with STS (100 nM).62 For
matters of comparison the peptides were tested along with pre-
viously described 5 and (N-Me)AVPF peptides (Charts 1 and 2).
It must be emphasized that the presented cellular assay is a
potentiation assay where cells are concomitantly treated with
STS (100 nM, Smin). The STS addition affects the cells by
bringing them to the brink of apoptosis, which enables the
measurement of a cellular apoptosis response not possible with
the Smac mimetics alone. The effect of the STS addition (100
nM, Smin) wasexamined, andnosignificantdeviationwasobserved
when compared to the media control without STS added. In this
way the observed differences in cellular responses are caused by
the presence of a given Smac mimetic.
On the basis of the present data set, it was difficult to deduce
structureÀactivity relationships. However, specific trends within
the libraries were quite evident. For instance, cis-triazolopro-
lines were generally more potent than their trans counterparts.
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Chart 2. Screening Data from Live/Dead Cellular STS-Potentiation Assay on Biarylalanine Libraries 10 and 11
RESEARCH ARTICLE
present investigation, we have taken advantage of this powerful
Table 4. Fitted Dose-Response Values for Selected Com-
pounds with and without STS
synthetic tool to generate libraries of structurally modified
Smac peptidomimetics. We have developed efficient metho-
dology for the 3-dimensional display of hydrophobic pharma-
cophore elements in the form of triazoloprolines and biaryl-
alanines, respectively, at the proline and isoleucine positions of
the parent Smac AVPI peptide, relying on robust and finely
optimized CuAAC and Suzuki cross-coupling reactions. The
protocols were optimized to very high efficiency (typical
purities >90%) and utilized for the generation of four distinct
libraries incorporating cis- or trans-triazoloproline and 3- or
4-biarylalanine residues. Subsequent biological screening of all
compounds revealed promising bioactives from each generic
library, thus confirming the relevance of the synthetic approach.
The presented biological data must be considered as the results
of a preliminary investigation. The specific mode of action of
the synthesized peptides cannot be deduced from the per-
formed cell survival assay. However, on the basis of prior art, it is
likely that the compounds partly function via IAP inhibition.
On-going efforts aim at the synthesis of new AVPI analogues
and for their detailed biological evaluation in various assays,
including protein binding studies and pull-down experiments
with supported AVPI analogues. The present strategy and findings
provide an entry for the generation of new Smac peptidomimetics
and justify the solid-phase synthesis and screening of larger com-
binatorial libraries.
Ca with STSb
(μM) ((SD)
Ca without STS
(μM) ((SD)
entry
compound
1
2
3
4
5
6
8{2}
10{6}
11{6}
11{13}
5
12.2 (2.0)
31.8 (2.3)
21.6 (2.7)
2.7 (0.4)
27.4 (2.5)
72.1 (10.7)
95.9 (13.7)
13.5 (0.4)
106.6 (14.7)
35.2 (3.9)
2.0 (0.003)
STS
a Concentration needed to reach a dead/live ratio 4-fold higher than STS
alone (100 nM). b STS concentration: 100 nM.
In general, within triazoloproline libraries 8 and 9, triazoles
derived from aliphatic alkynes appeared more active. In addition,
biarylalanine libraries 10 and 11 seemed significantly more
biologically active than the triazoloproline libraries. The results
of concentration-gradient experiments with and without STS are
shown for 4 selected compounds in Table 4. To our delight,
when tested along with STS, all peptides proved more active
potentiators than our starting hit 5, with benzothienyl-containing
compound 11{13} (Table 4, entry 4) showing very satisfying
single-digit potentiating activity (2.7 μM). But more interest-
ingly, when tested without STS, compound 11{13} was shown to
possess low micromolar activity (13.5 μM) on its own, indicating
a biological mode of action different from sole Smac mimicry.
’ CONCLUSION
’ EXPERIMENTAL PROCEDURES
There are only few reports on solid-phase organic synthesis
strategies for the formation of Smac peptidomimetics. In the
All experimental procedures are reported in the Supporting
Information.
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Supporting Information. Experimental procedures in-
b
cluding general methods, solution-phase and solid-phase synthetic
procedures and biological procedures. Analytical data for all building
blocks and compounds cleaved from the solid support. Fitted dose-
response curves. This material is available free of charge via the
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Corresponding Author
*E-mail: ten@kemi.dtu.dk.
Funding Sources
The research leading to these results has received funding from
the European Union’s Seventh Framework Programme managed
by REA-Research Executive Agency http://ec.europa.eu/
research/rea ([FP7/2007-2013] [FP7/2007-2011]) under grant
agreement no. [232447]. The Technical University of Denmark,
Danish Council for Independent Research, Natural Sciences
(TEN), Carlsberg Foundation (STLQ), and Torkil Holm Foun-
dation are gratefully acknowledged for financial support.
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