Solid-phase synthesis of substituted 2,3-diketopiperazines from reduced polyamides

Article abstract of DOI:10.1016/S0040-4020(00)00253-2

An efficient method for the solid phase synthesis of 1,6-disubstituted 2,3-diketopiperazine and 1,4,5-trisubstituted 2,3-diketopiperazine derivatives is described. The reduction of resin-bound acylated amino acids or resin-bound acylated dipeptides, followed by treatment with oxalyldiimidazole, affords the corresponding diketopiperazines in good yield and high purity. This is an example of a broader approach to the solid phase synthesis of individual heterocyclic compounds using peptides directly or indirectly as starting materials. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4020(00)00253-2

TETRAHEDRON
Pergamon
Tetrahedron 56 (2000) 3319±3326
Solid-Phase Synthesis of Substituted 2,3-Diketopiperazines from
Reduced Polyamides
*
Adel Nefzi, Marc A. Giulianotti and Richard A. Houghten
Torrey Pines Institute for Molecular Studies, 3550 General Atomics Court, San Diego, CA 92121, USA
Received 9 February 2000; accepted 22 March 2000
AbstractÐAn ef®cient method for the solid phase synthesis of 1,6-disubstituted 2,3-diketopiperazine and 1,4,5-trisubstituted 2,3-diketo-
piperazine derivatives is described. The reduction of resin-bound acylated amino acids or resin-bound acylated dipeptides, followed by
treatment with oxalyldiimidazole, affords the corresponding diketopiperazines in good yield and high purity. This is an example of a broader
approach to the solid phase synthesis of individual heterocyclic compounds using peptides directly or indirectly as starting materials. q 2000
Elsevier Science Ltd. All rights reserved.
Introduction
of classic 2,5-diketopiperazines.8 We report here an ef®cient
method for the solid-phase synthesis of 2,3-diketopipera-
zines, a less studied class of compounds. Our approach is
based on the reduction of resin-bound acylated amino acids
or resin-bound acylated dipeptides, followed by the
treatment of the resulting resin-bound polyamines with
oxalyldiimidazole to afford the corresponding 1,6-disubsti-
tuted-2,3-diketopiperazine and 1,4,5-trisubstituted-2,3-diketo-
piperazine derivatives, respectively (Scheme 1).
Combinatorial chemistry enables the rapid generation of
large numbers of diverse compounds, and when combined
with high-throughput screening techniques, offers a power-
ful strategy for the discovery and generation of drug leads.
Heterocyclic compounds having a high degree of structural
diversity have proven to be broadly and economically useful
as therapeutic agents.1 Diketopiperazines are conformation-
ally constrained scaffolds that are common in nature. Many
natural products containing a diketopiperazine structure
have been isolated and have been shown to have a wide
range of biological activities. Included in such compounds
are inhibitors of the mammalian cell cycle,2 inhibitors of
plasminogen activator-1 and topoisomerase.3 Diketopipera-
zines have been reported to be useful as ligands to the
neurokinin-2 receptor, and competitive antagonists to
Substance P at the neurokinin-1 receptor.4 A number of
approaches have been reported for the solid-phase synthesis
of diketopiperazine derivatives: Gordon and Steelle
developed a strategy for the solid-phase synthesis of diketo-
Results and Discussion
Starting from a p-methylbenzhydrylamine (MBHA) resin-
bound acylated amino acid, the amides were reduced in the
presence of borane±THF complex to afford two secondary
amines.9±11 The treatment of the resulting diamine with
oxalyldiimidazole generated the 1,6-disubstituted-2,3-
diketopiperazine 3 in good yield and high purity following
HF cleavage. We initially examined the feasibility of this
synthetic route using ®ve representative l-amino acids (Ser,
Tyr, Val, Ala, Phe), and four carboxylic acids (phenylacetic
acid, acetic acid, cyclohexyl acetic acid, isobutyric acid). A
total of 19 disubstituted diketopiperazines 3 were synthe-
sized as controls (Table 1).
piperazines based on reductive amination on the solid
à Á
5
support; a similar approach has been published by Krchnak
and co-workers for the synthesis of persubstituted 2,5-
diketopiperazine,6 and Scott and co-workers developed an
alternative strategy for the synthesis of a similar diketo-
piperazine library using a-bromocarboxylic acids and a
range of amines.7
In order to increase the amount of diversity around the
diketopiperazine ring, we extended our approach to the
solid-phase synthesis of 1,4,6-trisubstituted-2,3-diketopi-
perazines 6. Starting from the same resin-bound amino
acid 1 and following trityl (Trt) protection of the amino
group, a selective N-alkylation of the amide linked to the
solid support was performed using lithium t-butoxide in
THF, followed by addition of an alkylating agent in
dimethyl sulfoxide (DMSO).12,13 We have successfully
used the following halogenated alkyls for the N-alkylation
All existing reported approaches for the solid-phase
synthesis of diketopiperazines describe only the synthesis
Keywords: combinatorial chemistry; solid-phase synthesis; 2,3-diketo-
piperazines.
* Corresponding author. Tel.: 1858-455-3803; fax: 1858-455-3804;
e-mail: rhoughten@tpims.org
0040±4020/00/$ - see front matter q 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020(00)00253-2

3320
A. Nefzi et al. / Tetrahedron 56 (2000) 3319±3326
Scheme 1. Solid-phase synthesis of 1,6-disubstituted 2,3-diketopiperazines and 1,4,5-trisubstituted 2,3-diketopiperazines from resin-bound polyamines: (a)
Fmoc-Xaa-OH, DIPCDI, HOBt, DMF; (b) 20% piperidine in DMF; (c) Trt-Cl, DIPEA, DCM; (d) R2-X, tBuOLi, DMSO; (e) 2% TFA in DCM, DIPEA/DCM;
(f) Fmoc-Xaa-OH, DIPCDI, DMF; (g) 20% piperidine in DMF; (h) R4COOH, DIPCDI, DMF; (i) BH3±THF, 658C; (j) oxalyldiimidazole, DMF; (k) HF/
anisole.
of amides: methyliodide, ethyliodide, benzylbromide,
2-bromomethylnaphthylene and allylbromide.12 Following
removal of the Trt protecting group with 2% TFA in DCM
and neutralization, the second amino acid was added using
traditional solid-phase peptide chemistry,14 and the resulting
dipeptide 4 was treated with appropriate carboxylic acids to
obtain the acylated dipeptide. The reduction of the amide
groups of the resin-bound N-acylated dipeptide using
diborane in THF at 658C yielded a single tertiary and two
secondary amines 5.11,13 Potential racemization during
tritylation, amide alkylation and reduction was examined
using analytical reverse-phase high performance liquid
chromatography (RP-HPLC). Different dipeptides were
used as controls for the reactions involved, and all possible
diasterioisomers were synthesized. Racemization was
monitored by comparing the respective absorbances at
214 nm of two diastereomeric pairs that do not coelute.
Different pairs for diastereoisomers that do not coelute
were run in different gradients. The lack of diastereomeres
in the RP-HPLC and in the 1H NMR suggest little (,2%) or
no racemization occurred during the N-alkylation.13 The
treatment of the resin-bound polyamine 5 with oxalyldi-
imidazole in anhydrous DMF, followed by HF cleavage,
yielded the desired 1,4,5-trisubstituted-2,3-diketopiperazine
6 (Table 2).
As a ®rst attempt in the synthesis of diketopiperazine
derivatives 6, we initially optimized the reaction conditions
of this synthetic route by the parallel synthesis of 24 indi-
vidual compounds. We chose benzyl bromide as the R2
Table 1. RP-HPLC purity and masses found for the prepared diketopiperazines 3
Entry
R1
R4
HPLC purity (%)a
MW expected
MW found
3a
3b
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
3m
3n
3o
3p
3q
3r
3s
CH2±C6H4±OH
CH2±C6H4±OH
CH2±C6H4±OH
CH2Ph
CH2Ph
CH2Ph
CH2Ph
CH3
CH3
CH3
CH3
CH2OH
CH2OH
CH2OH
CH2OH
CH(CH3)2
CH(CH3)2
CH(CH3)2
CH(CH3)2
CH2Ph
.95
92
89
324.37
316.39
276.33
308.37
232.28
300.40
260.33
232.28
156.18
224.30
184.24
248.28
172.18
240.30
200.24
260.33
184.24
252.35
212.29
325.4 (MH1)
317.3 (MH1)
277.2 (MH1)
309.3 (MH1)
233.2 (MH1)
301.2 (MH1)
261.2 (MH1)
233.2 (MH1)
157.2 (MH1)
225.2 (MH1)
185.1 (MH1)
249.9 (MH1)
173.2 (MH1)
241.2 (MH1)
201.9 (MH1)
261.2 (MH1)
185.2 (MH1)
253.2 (MH1)
213.2 (MH1)
CH2±C6H11
CH(CH3)2
CH2Ph
.95
.95
.95
92
.95
.95
.95
.95
70
83
87
92
90
CH3
CH2±C6H11
CH(CH3)2
CH2Ph
CH3
CH2±C6H11
CH(CH3)2
CH2Ph
CH3
CH2±C6H11
CH(CH3)2
CH2Ph
CH3
CH2±C6H11
CH(CH3)2
.95
89
93
a The products were run on a Vydac C18 column, using a 5±95% gradient of 0.05% TFA in ACN in 7 min. The purity was estimated using analytical traces at
214 nm. The yields obtained in all cases were higher than 75% relative to the initial loading of the resin.

A. Nefzi et al. / Tetrahedron 56 (2000) 3319±3326
Table 2. RP-HPLC purity and masses found for the prepared diketopiperazines 6
3321
Entry
R1
R3
R4
HPLC purity (%)a
MW expected
MW found
6a
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
6m
6n
6o
6p
6q
6r
6s
(CH2)4N(CH3)CH2Ph
CH2Ph
CHCH2CH(CH3)2
(CH2)4N(CH3)CH2Ph
CH2OH
Ph
CH2Ph(m-Cl) (o-Cl)
CH2Ph(m-OMe)
CH2Ph
Cycloheptyl
CH2Ph
CH2Ph
93
92
93
94
76
78
.95
.95
.95
.95
92
.95
.95
.95
.95
95
92
92
84
94
92
.95
93
670.28
485.62
497.30
608.86
471.25
423.25
455.26
598.82
616.38
568.38
554.77
582.82
596.85
497.67
511.70
570.76
612.84
646.86
660.93
584.79
632.83
626.87
616.83
531.29
671.6 (MH1)
486.6 (MH1)
498.3 (MH1)
609.5 (MH1)
472.2 (MH1)
424.2 (MH1)
456.3 (MH1)
599.6 (MH1)
617.6 (MH1)
569.5 (MH1)
555.5 (MH1)
583.7 (MH1)
597.7 (MH1)
498.4 (MH1)
512.6 (MH1)
571.7 (MH1)
613.6 (MH1)
647.9 (MH1)
662.0 (MH1)
585.7 (MH1)
633.6 (MH1)
627.7 (MH1)
617.7 (MH1)
532.2 (MH1)
CH3
CH2Ph
Ph
CH2Ph
CH(CH3)2
CH2Ph
CH(CH3)2
CH2Ph
CH(CH3)2
CH3
CH(CH3)2
CH(CH3)CH2CH3
CH(CH3)2
CH(CH3)CH2CH3
CH3
CH(CH3)2
Ph
Ph
CH2OH
CH3
CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
CH2Ph
CH2Ph(p-OMe)
CH2Ph
CH2Ph
CH2Ph(p-Me)
CH2Ph(p-Me)
CH2Ph(p-Me)
CH2Ph(p-Me)
CH2Ph(p-Me)
CH2Ph(p-OMe)
CH2Ph(p-OEt)
CH2Ph(p-OEt)
CH2-adamantyl
CH2Ph(p-OEt)
CH2Ph(p-Me)
CH2Ph(p-OMe)
CH2Ph(p-Me)
CH2Ph
CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
(CH2)4N(CH3)CH2Ph
CH2Ph
6t
CH3
Ph
6u
6v
6w
6x
CH(CH3)CH2CH3
Ph
CH2Ph
.95
a The products were run on a Vydac C18 column, using a 5±95% gradient of 0.05% TFA in ACN in 7 min. The purity was estimated using analytical traces at
21 nm. The yields obtained in all cases were higher then 75% relative to the initial loading of the resin.
Figure 1. LC-MS of the diketopiperazine 6k obtained from lysine, alanine and 4-methylphenylacetic acid.

3322
A. Nefzi et al. / Tetrahedron 56 (2000) 3319±3326
Figure 2. LC-MS of the diketopiperazine 6o obtained from phenylalanine, isoleucine and 4-methylphenylacetic acid.
alkylating group, ®ve different l-amino acids (Ser, Ala, Phe,
Lys and Leu) for R ®ve amino acids for R3 (Val, Phe, Ile,
Ala and Phenylglycine) and eight carboxylic acids for R4
(phenylacetic acid, p-tolyl acetic acid, p-methoxyphenyl-
acetic acid, p-ethoxyphenylacetic acid, adamantane acetic
acid, m-methoxyphenylacetic acid, cyclohepthyl carboxylic
acid and 3,4-dichlorophenylacetic acid). As shown in Table
2, excellent purities were obtained for all cases. During
N-alkylation with Bzl-Br, the protected Ne-amine of lysine
was also N-benzylated; during the reduction step, the Boc
protecting group was reduced to the N-methyl.12,13 Figs. 1
and 2 show the LC-MS spectra of the diketopiperazines 6k
and l, which are representatives of the purities obtained for
all cases.
cleaved due to the required exposure for six hours to HF
cleavage conditions. These carboxylic acids were therefore
excluded from our libraries as well as the nitro aromatic
carboxylic acids due to the partial reduction of the nitro
group during borane treatment. The majority of the aromatic
carboxylic acids chosen for the library synthesis are deriva-
tives of phenylacetic acid. Using the information from these
control experiments and following the selection of the
appropriate compounds for each of the three positions of
diversity, a trisubstituted 2,3-diketopiperazine library is
being prepared. The synthesis of this library and its use in
screening for the identi®cation of highly active compounds
will be reported elsewhere. We have also successfully used
the strategy described to generate cyclic urea and cyclic
thiourea libraries by treating the generated resin-bound
polyamine 5 with carbonyldiimidazole and thiocarbonyldi-
imidazole, respectively.15
Modi®cations occurring in the functionalized amino acid
side chains during the N-alkylation and reduction steps
have been reported.15 Expanding our optimization and
control compound set, we examined more than 40 amino
acids at the ®rst and second positions of diversity, and 60
carboxylic acids at the third position. These individual
control compounds, in which the individual building blocks
were varied while the other two positions remained ®xed,
were synthesized in order to determine the effect of the
reaction conditions on each of the building blocks. The
amino acids Arg(Pmc), Trp(Boc), and His(Trt) which
yielded aggregation and/or polymerized products were
excluded since they led to the desired product in less than
50%. Excellent yields were obtained for the remaining
amino acids with average HPLC purities higher than 80%.
We also observed that following amide reduction electron
rich substituted benzoic acids (such as p-methoxy-benzoic
acid), which yield N-benzyl derivatives, are completely
Using the `libraries from libraries' concept,16 this laboratory
has reported previously a range of combinatorial libraries
prepared on the solid-phase, then cleaved and utilized in
solution.12,13,17 Such libraries incorporate a wide range of
functionalities, including hydrogen bonding donors and
acceptors, positive and negative charges, functionalities of
differing chiralities, hydrophobic and hydrophilic sub-
stituents, etc.
Conclusion
This work is part of our ongoing efforts in the solid-phase
synthesis of individual small molecule and heterocyclic
compounds and subsequent combinatorial libraries using

A. Nefzi et al. / Tetrahedron 56 (2000) 3319±3326
3323
amino acids and peptides as the starting material.17
N-acylated amino acids and N-acylated dipeptides have
been successfully used for the generation of a variety of
1,6-disubstituted 2,3-diketopiperazines and 1,4,5-trisubsti-
tuted 2,3-diketopiperazines, respectively, in high purity.
The general nature of this approach permits not only large
numbers of individual diketopiperazines to be prepared, but
also mixture-based combinatorial libraries.17
followed by quenching with MeOH. The resin was then
washed with methanol (4£) and the borane disproportion-
ated by treatment with piperidine at 658C overnight. The
resin was then washed with methanol (2£) and DMF (6£)
and dried. The completeness of the reaction was veri®ed by
cleavage and analysis following reduction.
(3) Disubstituted diketopiperazine formation: The cycliza-
tion occurred following treatment of the resin-bound
reduced acylated amino acid overnight with 5-fold
excess of oxalyldiimidazole (0.1 M) in anhydrous DMF.
Following cleavage from the resin with anhydrous HF in
the presence of anisole at 08C for 90 min, the desired
product was extracted with acetonitrile/water (50:50) and
lyophilized.
Experimental
Fmoc-amino acid derivatives and HOBt were purchased
from Calbiochem-Novabiochem Corp. (San Diego, CA),
Bachem Bioscience Inc. (Philadelphia, PA), and Bachem
California (Torrance, CA). MBHA resin (1% divinyl-
benzene, 100±200 mesh, 1 mmol/g substitution) was
purchased from Peninsula Laboratories, Inc. (Belmont,
CA). N,N0-Diisopropylcarbodiimide (DIPCDI) was pur-
chased from Chem Impex International (Wood Dale, IL),
tri¯uoroacetic acid from Halocarbon (River Edge, NJ) and
hydrogen ¯uoride from Air Products (San Marcos, CA). All
other reagents and anhydrous solvents (DMSO and THF)
were purchased from Aldrich Chemical Company
(Milwaukee, WI). Analytical RP-HPLC was performed on
a Beckman System Gold instrument (Fullerton, CA).
Samples were analyzed using a Vydac 218TP54 C18 column
(0.46£25 cm).
1-Phenethyl-6(S)-p-hydroxybenzyltetrahydro-2,3-diketo-
piperazine (3a). 1H NMR (500 MHz, DMSO-d6) d 8.43 (d,
Jˆ4.5 Hz, 1H), 7.32±6.66 (m, 9H), 3.88 (m, 1H), 3.76 (m,
1H), 3.23 (dd, Jˆ13.1, 2.4 Hz) 2.89 (m, 1H), 2.88 (m, 2H),
2.82 (m, 2H), 2.63 (dd, Jˆ13.3, 9.5 Hz, 1H). 13C NMR
(125 MHz, DMSO-d6): 157.4, 157, 156.1, 138.9, 130.1,
128.7, 128.3, 127.3, 126.3, 115.2, 56.9, 54.9, 47.5. ES-MS
calcd for C19H20N2O3: 324.373, found: 325.402 (MH1).
1-Ethyl-6(S)-benzyltetrahydro-2,3-diketopiperazine (3e).
1H NMR (500 MHz, DMSO-d6) d 8.45 (d, Jˆ3.8 Hz, 1H),
7.34±7.23 (m, 5H), 3.72 (m, 1H), 3.64 (dd, Jˆ13.3, 6.9 Hz,
1H), 3.45 (dd, Jˆ13.2, 3.3 Hz), 2.99 (dd, Jˆ13.3, 5.1 Hz,
1H), 2.95 (dd, Jˆ14.4, 5.4 Hz, 1H), 2.88 (dd, Jˆ13.5,
6.92 Hz, 1H), 2.81 (dd, Jˆ13.1, 9.7 Hz, 1H), 1.08 (t,
Jˆ6.9 Hz, 3H). 13C NMR (125 MHz, DMSO-d6): 157.6,
156.8, 137.4, 129.2, 128.5, 126.6, 55.8, 40.5, 40.2, 36.7,
13.1. ES-MS calcd for C13H16N2O2: 232.284, found:
233.207 (MH1).
Typical procedure for the individual synthesis of 1,6-
disubstituted 2,3-diketopiperazine (3)
Solid phase syntheses were carried out using the `teabag'
method, in which the resin is contained within sealed poly-
propylene mesh packets.18 The completeness of amino acid
coupling and N-acylation were veri®ed using the ninhydrin
test.19
1-Isobutyl-6(S)-methyltetrahydro-2,3-diketopiperazine
(3k). 1H NMR (500 MHz, DMSO-d6) d 8.47 (d, Jˆ4.1 Hz,
1H), 3.62 (m, 1H), 3.61 (m, 1H), 3.53 (dd, Jˆ13.4, 7.8 Hz,
1H), 3.06 (m, 1H), 2.70 (dd, Jˆ13.1, 7.2 Hz, 1H), 1.93 (m,
1H), 1.22 (d, Jˆ7.1 Hz, 3H), 0.88 (d, Jˆ6.7 Hz, 3H), 0.82
(d, Jˆ6.7 Hz, 3H). 13C NMR (125 MHz, DMSO-d6): 157.5,
157.4, 51.6, 50.6, 43.1, 26.5, 19.9, 19.9, 16.7. ES-MS calcd
for C9H16N2O2: 184.243, found: 185.105 (MH1).
(1) Amino acid coupling and N-acylation: 100 mg p-
methylbenzydrylamine (MBHA) resin (0.1 mol-equiv./g,
100±200 mesh) was contained within a sealed polypropy-
lene mesh packet. Reactions were carried out in 10 ml poly-
ethylene bottles. Following neutralization with 5%
diisopropylethylamine (DIPEA) in dichloromethane
(DCM), the resin was washed with DCM. The ®rst amino
acid (Fmoc-Xaa-OH, 6 equiv.) was coupled using the
conventional reagents hydroxybenzotriazole (HOBt,
6 equiv.) and diisopropylcarbodiimide (DIPCDI, 6 equiv.)
in anhydrous DMF for 60 min. Following removal of the
Fmoc group with 20% piperidine in DMF (2£10 min) and
washing with DMF (8£), the amino acid was N-acylated
with a carboxylic acid (10 equiv.) in the presence of
DIPCDI (10 equiv.) and HOBt (10 equiv.) overnight in
anhydrous DMF.
1-Ethyl-6(S)-isopropyltetrahydro-2,3-diketopiperazine
1
(3q). H NMR (500 MHz, DMSO-d6) 8.34 (d, Jˆ3.9 Hz,
1H), 3.81 (m, 1H), 3.46 (m, 1H), 3.28 (m, 2H), 2.89 (m,
1H), 1.96 (m, 1H), 1.09 (t, Jˆ6.9 Hz, 1H), 0.96 (d,
Jˆ6.7 Hz, 3H), 0.88 (d, Jˆ6.7 Hz, 3H). 13C NMR
(125 MHz, DMSO-d6): 157.5, 157.4, 51.6, 50.6, 43.1,
26.5, 19.9, 19.9, 16.7. ES-MS calcd for C9H16N2O2:
184.245, found: 185.206 (MH1).
1-Cyclohexylmethyl-6(S)-p-hydroxybenzyltetrahydro-
2,3-diketopiperazine (3b) 1H NMR (500 MHz, DMSO-d6)
8.45 (d, Jˆ5.2 Hz, 1H), 7.01 (d, Jˆ8.1 Hz, 2H), 6.69 (d,
Jˆ8.5 Hz, 2H), 3.57 (dd, Jˆ7.3, 13.3 Hz, 1H), 3.48 (m, 2H),
2.96 (dd, Jˆ5.6, 12.1 Hz), 1H), 2.85 (dd, Jˆ4.8, 13.3 Hz,
1H), 2.65 (dd, Jˆ9.3, 13.2 Hz, 1H), 2.57 (dd, Jˆ6.9,
13.1 Hz, 1H), 1.60 (m, 6H), 1.14 (m, 3H), 0.94 (m, 1H),
0.85 (m, 1H). ES-MS calcd for C18H24N2O3: 316.398,
found: 317.301 (MH1).
(2) Exhaustive reduction of the amide groups: The reduction
was performed in 50 ml kimax tubes under nitrogen. The
resin packet (1 mol-equiv. resin, 100 mg of starting resin,
0.2 mol-equiv. carbonyl) and boric acid (15-fold excess
over each amide bond) were added to each tube. Trimethyl
borate (15-fold excess over each amide bond) was added,
followed by 1 M BH3±THF (40-fold excess over each
amide bond). The tubes were heated at 658C for 72 h,

3324
A. Nefzi et al. / Tetrahedron 56 (2000) 3319±3326
1-Isobutyl-6(S)-p-hydroxybenzyltetrahydro-2,3-diketo-
piperazine (3c). H NMR (500 MHz, DMSO-d6) 8.46 (d,
Jˆ4.35 Hz, 1H), 5.13 (1s, 1H), 3.67 (m, 2H), 3.50 (m,
3H), 3.08 (m, 2H), 1.10 (m, 3H). ES-MS calcd for
C7H12N2O3: 172.186, found: 173.203 (MH1).
1
Jˆ5.2 Hz, 1H), 7.01 (d, Jˆ8.2 Hz, 2H), 6.69 (d, Jˆ8.5 Hz,
2H), 3.53 (m, 3H), 2.96 (dd, Jˆ5.6, 12.7 Hz, 1H), 2.86 (dd,
Jˆ4.9, 13.4 Hz, 1H), 2.66 (dd, Jˆ9.3, 13.4 Hz, 1H), 2.56
(dd, Jˆ7.1, 13.1 Hz, 1H), 1.96 (m, 1H), 0.87 (d, Jˆ6.3 Hz,
3H), 0.81 (d, Jˆ6.8 Hz, 3H). ES-MS calcd for C15H20N2O3:
276.332, found: 277.207 (MH1).
1-Cyclohexylmethyl-6(S)-hydroxymethyltetrahydro-2,3-
diketopiperazine (3n). 1H NMR (500 MHz, DMSO-d6)
8.35 (d, Jˆ4.8 Hz, 1H), 5.12 (s, 1H), 3.63 (dd, Jˆ7.2,
13.4 Hz, 1H), 3.57 (m, 2H), 3.42 (m, 3H), 2.74 (dd,
Jˆ6.9, 13.1 Hz, 1H), 1.63 (m, 6H), 1.19 (m, 3H), 1.16 (m,
1H), 0.88 (m, 1H). ES-MS calcd for C12H20N2O3: 240.301,
found: 241.208 (MH1).
1-Phenethyl-6(S)-benzyltetrahydro-2,3-diketopiperazine
1
(3d). H NMR (500 MHz, DMSO-d6) 8.45 (d, Jˆ5.1 Hz,
2H), 7.30 (m, 5H), 7.21 (m, 5H), 3.87 (m, 1H), 3.50 (m,
1H), 3.26 (dd, Jˆ4.1, 13.17 Hz, 1H), 3.01 (m, 1H), 2.94 (dd,
Jˆ5.5, 13.3 Hz, 1H), 2.81 (m, 4H). ES-MS calcd for
C19H20N2O2: 308.371, found: 309.306 (MH1).
1-Isobutyl-6(S)-hydroxymethyltetrahydro-2,3-diketo-
piperazine (3o). H NMR (500 MHz, DMSO-d6) 8.36 (d,
1
Jˆ4.7 Hz, 1H), 3.61 (m, 4H), 3.54 (m, 2H), 3.31 (dd, Jˆ4.9,
12.9 Hz, 1H), 2.73 (dd, Jˆ7.1, 12.9 Hz, 1H), 1.94 (m, 1H),
0.89 (d, Jˆ6.8, 3H), 0.84 (d, Jˆ6.7 Hz, 3H). ES-MS calcd
for C9H16N2O3: 200.241, found: 201.109 (MH1).
1-Cyclohexylmethyl-6(S)-p-hydroxybenzyltetrahydro-
2,3-diketopiperazine (3f). 1H NMR (500 MHz, DMSO-d6)
8.47 (d, Jˆ5.1 Hz, 1H), 7.32 (m, 2H), 7.24 (m, 3H), 3.56 (m,
3H), 3.01 (dd, Jˆ5.1, 13.3 Hz, 1H), 2.94 (dd, Jˆ5.9,
13.7, Hz, 1H), 2.79 (dd, Jˆ9.5, 13.1 Hz, 1H), 2.57 (dd,
Jˆ7.1, 13.4 Hz, 1H), 1.65 (m, 6H), 0.93 (m, 1H), 0.83 (m,
1H). ES-MS calcd for C18H24N2O2: 300.402, found: 301.208
(MH1).
1-Phenethyl-6(S)-isopropyltetrahydro-2,3-diketopiper-
azine (3p). 1H NMR (500 MHz, DMSO-d6) 8.32 (d,
Jˆ4.36 Hz, 1H), 7.30 (m, 2H), 7.22 (m, 3H), 4.05 (m,
1H), 3.12 (m, 4H), 2.88 (m, 1H), 2.83 (m, 1H), 1.91 (d,
Jˆ6.86 Hz, 3H), 0.84 (d, Jˆ6.80 Hz, 3H). ES-MS calcd
for C15H20N2O2: 260.33, found: 261.20 (MH1).
1-Isobutyl-6(S)-benzyltetrahydro-2,3-diketopiperazine
1
(3g). H NMR (500 MHz, DMSO-d6) 8.49 (d, Jˆ5.18 Hz,
1-Cyclohexylmethyl-6(S)-isopropyltetrahydro-2,3-diketo-
piperazine (3r). H NMR (500 MHz, DMSO-d6) 8.36 (d,
1
1H), 7.33 (m, 3H), 7.24 (m, 2H), 3.63 (m, 1H), 3.54 (m, 2H),
2.99 (dd, Jˆ5.3 Hz, 1H), 2.94 (dd, Jˆ5.9, 13.9 Hz, 1H),
2.79 (dd, Jˆ9.4, 13.1 Hz, 1H), 2.56 (dd, Jˆ7.1, 13.4 Hz,
1H), 1.05 (m, 1H), 0.87 (d, Jˆ6.7 Hz, 3H), 0.81 (d,
Jˆ6.8 Hz, 3H). ES-MS calcd for C15H20N2O2: 260.334,
found: 261.207 (MH1).
Jˆ5.10 Hz, 1H), 3.76 (dd, Jˆ6.76, 13.40 Hz, 1H), 3.57 (dd,
Jˆ4.18, 13.74 Hz, 1H), 3.28 (dd, Jˆ5.89, 13.76 Hz, 1H),
3.17 (dd, Jˆ3.89, 7.53 Hz, 1H), 2.60 (dd, Jˆ7.39, 13.01 Hz,
1H), 1.95 (m, 1H), 1.67 (m, 4H), 1.59 (m, 2H), 1.14 (m, 3H),
0.93 (d, Jˆ6.87 Hz, 3H), 0.87 (d, Jˆ6.77 Hz, 3H). ES-MS
calcd for C14H24N2O2: 252.35, found: 253.20 (MH1).
1-Phenethyl-6(S)-methyltetrahydro-2,3-diketopiperazine
1
(3h). H NMR (500 MHz, DMSO-d6) 8.43 (d, Jˆ3.9 Hz,
1-Isobutyl-6(S)-isopropyltetrahydro-2,3-diketopiperazine
1
1H), 7.29 (m, 2H), 7.24 (m, 3H), 3.86 (ddd, Jˆ5.95, 5.49,
8.6 Hz, 1H), 3.47 (m, 1H), 3.39 (dd, Jˆ4.1, 12.9 Hz, 1H),
3.16 (m, 1H), 2.9 (m, 2H), 1.17 (d, Jˆ6.7 Hz, 3H). ES-MS
calcd for C13H16N2O2: 232.283, found: 233.209 (MH1).
(3s). H NMR (500 MHz, DMSO-d6) 8.39 (d, Jˆ4.88 Hz,
1H), 3.74 (dd, Jˆ6.95, 12.88 Hz, 1H), 3.56 (dd, Jˆ4.41,
13.69 Hz, 1H), 3.29 (dd, Jˆ5.86, 13.73 Hz, 1H), 3.20 (m,
1H), 2.59 (dd, Jˆ7.81, 12.96 Hz, 1H), 1.97 (m, 2H), 0.93 (d,
Jˆ6.87 Hz, 3H), 0.89 (d, Jˆ7.6 Hz, 3H), 0.88 (d,
Jˆ7.14 Hz, 3H), 0.82 (d, Jˆ6.38 Hz, 3H). ES-MS calcd
for C11H20N2O2: 212.29, found: 213.20 (MH1).
1-Ethyl-6(S)-methyltetrahydro-2,3-diketopiperazine (3i).
1H NMR (500 MHz, DMSO-d6) 8.43 (1s, 1H), 3.68 (m, 1H),
3.57 (m, 2H), 3.04 (m, 2H), 1.23 (d, Jˆ6.7 Hz, 3H), 1.09 (t,
Jˆ6.9 Hz, 3H). ES-MS calcd for C7H12N2O2: 156.187,
found: 157.204 (MH1).
Typical procedure for the individual synthesis of 1,4,5-
trisubstituted 2,3-diketopiperazine (6)
1-Cyclohexylmethyl-6(S)-methyltetrahydro-2,3-diketo-
piperazine (3j). H NMR (500 MHz, DMSO-d6) 8.45 (d,
1
(1) Amino acid coupling and selective N-alkylation: The
®rst amino acid was coupled in the same conditions as
described before. Following removal of the protecting
group with 20% piperidine in DMF (1£10 min) and wash
with DMF (8£), the mesh packet was shaken overnight in a
solution of trityl chloride in DCM/DMF (9:1) in the
presence of DIPEA. Completeness of the trityl coupling
was veri®ed using the bromophenol blue color test.20
N-alkylation was performed by treatment of the resin packet
with 0.5 M lithium t-butoxide in THF (20 equiv.) for 10 min
at room temperature. Excess base was removed by cannu-
lation, followed by addition of the individual alkylating
agent (20 equiv.) in anhydrous DMSO. The solution
was vigorously shaken for 2 h at room temperature (this
operation was repeated three times).
Jˆ4.0 Hz, 1H), 3.60 (m, m, 3H), 3.04 (dd, Jˆ5.6, 11.1 Hz,
1H), 2.72 (dd, Jˆ6.8, 13.4 Hz, 1H), 1.63 (m, 6H), 1.21 (d,
Jˆ8.6 Hz, 3H), 1.14 (m, 3H), 0.93 (m, 1H), 0.88 (m, 1H).
ES-MS calcd for C12H20N2O2: 224.301, found: 225.287
(MH1).
1-Phenethyl-6(S)-hydroxymethyltetrahydro-2,3-diketo-
piperazine (31). H NMR (500 MHz, DMSO-d6) 8.33 (d,
1
Jˆ4.7 Hz, 1H), 7.30 (m, 5H), 5.12 (m, 1H), 3.93 (m, 1H),
3.32 (m, 2H), 3.18 (m, 3H), 2.84 (m, 2H). ES-MS calcd for
C13H16N2O3: 248.28, found: 249.20 (MH1).
1-Methyl-6(S)-hydroxymethyltetrahydro-2,3-diketopiper-
azine (3m). 1H NMR (500 MHz, DMSO-d6) 8.34 (d,

A. Nefzi et al. / Tetrahedron 56 (2000) 3319±3326
3325
(2) N-Acylated dipeptide: Upon removal of the trityl from
the a-amino group with 2% TFA in DCM (2£10 min), the
resin packet was washed, neutralized with a solution of 5%
DIEA in DCM, and the second amino acid (Fmoc-Xaa-OH)
d6): 157.58, 155.97, 138.69, 137.53, 131.41, 131.15, 130.21,
129.89, 129.62, 129.17, 129.04, 128.91, 128.73, 128.68,
128.39, 126.79, 126.36, 58.39, 55.32, 54.34, 50.62, 47.94,
47.65, 37.16, 33.00, 28.42, 22.92. ES-MS calcd for
C40H48N4O2: 616.34, found: 617.60 (MH1).
²
coupled in the same conditions as described above. Follow-
ing removal of the Fmoc group, the dipeptide was
N-acylated with a carboxylic acid (10 equiv.) overnight in
the presence of DIPCDI (10 equiv.) and HOBt (10 equiv.) in
anhydrous DMF.
1-{1(S)-[Hydroxymethyl]-1(S)-[benzylaminomethyl)]-
ethyl}-5(S)-benzyl-4-phenethyltetrahydro-2,3-diketopiper-
azine (6e). 1H NMR (500 MHz, DMSO-d6) d 7.24±7.36 (m,
15H), 5.13 (m, 1H), 4.57 (d, Jˆ14.5 Hz, 1H), 4.46 (d,
Jˆ14.5 Hz, 1H), 4.12 (dd, Jˆ8.7, 14.8 Hz, 1H), 3.92 (m,
1H), 3.51 (m, 2H), 3.18±3,26 (m, 5H), 3.08 (dd, Jˆ3.8,
14.8 Hz, 1H), 2.88 (m, 3H). 13C NMR (125 MHz, DMSO-
d6): 158.47, 156.14, 136.92, 136.13, 129.10, 128.72, 128.64,
128.55, 128.23, 127.56, 127.08, 126.88, 60.06, 56.18,
54.57, 49.92, 46.20, 44.96, 44.72, 42.50, 36.22, 34.57,
31.73. ES-MS calcd for C29H33N3O3: 471.25, found: 472.2
(MH1).
(3) Exhaustive reduction of the amide groups: The reduction
was performed in the same conditions as described above
(the characterization and the purities of the intermediate
diethylenetriamines have been reported).13
(4) Trisubstituted diketopiperazine formation: The cycli-
zation occurred following treatment of the reduced acylated
dipeptide overnight with 5-fold excess of oxalyldiimidazole
(0.1 M) in anhydrous DMF. Following cleavage from the
resin with anhydrous HF in the presence of anisole at 08C
for 7 h,21 the desired product was extracted with aceto-
nitrile/water (50:50) and lyophilized.
1-{1(S)-[(Benzylamino)methyl]-5-[benzyl(methyl)amino]-
pentyl}-5(S)-isopropyl-4-phenethyltetrahydro-2,3-diketo-
piperazine (6n). H NMR (500 MHz, DMSO-d6) d 7.08±
1
7.40 (m, 14H), 4.92 (m, 1H), 4.17 (m 1H), 4.05 (m, 1H),
3.98 (m, 2H), 3.69 (d, Jˆ5.38 Hz, 2H), 3.06 (m, 1H), 2.96±
2.88 (m, 5H), 2.77 (m, 2H), 2.26 (1s, 3H), 1.62 (m, 1H),
0.78 (m, 12H). 13C NMR (125 MHz, DMSO-d6): 157.85,
156.26, 136.78, 135.67, 135.31, 132.23, 129.87,
128.99, 128.89, 128.83, 128.74, 128.57, 128.50,
126.73, 59.33, 50.40, 49.95, 35.00, 32.89, 29.74, 20.66,
19.61, 19.18. ES-MS calcd for C32H39N3O3: 497.67,
found: 498.4 (MH1).
For the library synthesis, the selected R1 and R3 groups were
derived from Gly, Phg, Met(O) and the d- and l- forms of
the following amino acids: Ala, Phe, Ile, Leu, Nva,
Ser(tBu), Thr(tBu), Val, Tyr(tBu), Nle, Cha and Nal. In
addition R1 included the d- and l- forms of Lys(Boc). R2
included also b-Ala.
The selected R3 were derived from the following carboxylic
acids: 1-phenyl-1-cyclopropane carboxylic acid, m-tolyl-
acetic acid, 3-¯uorophenylacetic acid, (a,a,a)-tri¯uoro-m-
tolylacetic acid, p-tolylacetic acid, 3-methoxyphenylacetic
acid, 4-methoxyphenylacetic acid, 4-ethoxyphenylacetic
acid, 4-isobutyl-a-methylphenylacetic acid, 3,4-dichloro-
phenylacetic acid, 3,5-bis(tri¯uoromethyl)-phenylacetic acid,
phenylacetic acid, hydrocinnamic acid, 4-phenylbutyric
acid, butyric acid, heptanoic acid, isobutyric acid, isovaleric
acid, 4-methylvaleric acid, trimethylacetic acid, tert-butyl-
acetic acid, cyclohexanecarboxylic acid, cyclohexylacetic
acid, cycloheptanecarboxylic acid, acetic acid, cyclobutane-
carboxylic acid, cyclopentanecarboxylic acid, cyclohexane-
propionic acid, 4-methyl-1-cyclohexanecarboxylic acid,
4-tert-butyl-cyclohexanecarboxylic acid, 1-adamantane-
acetic acid, 3,3-diphenylpropionic acid, dicyclohexylacetic
acid, indole-3-acetic acid, 1-naphthylacetic acid, 3-(3,4,5)-
trimethoxyphenylpropionic acid, 2-norbornaneacetic acid,
cyclopentylacetic acid, 2-ethylbutyric acid.
1-[1(S)-(Benzyl)-2-(benzylamine)]-5(S)-methyl-3-m-meth-
oxyphenethyltetrahydro-2,3-diketopiperazine (6t). 1H
NMR (500 MHz, DMSO-d6) d 7.42±7.19 (m, 11H), 6.85±
6.78 (m, 3H), 5.15 (m, 1H), 4.20 (m, 2H), 3.73 (s, 3H), 3.70
(m, 2H), 3.16 (m, 3H), 2.97 (m, 4H), 2.80 (m, 3H), (m, 3H).
13C NMR (125 MHz, DMSO-d6): 159.35, 157.89, 156.03,
140.38, 136.59, 134.68, 131.79, 129.97, 129.48, 129.05,
128.79, 128.48, 126.71, 120.89, 120.124, 114.239, 113.77,
113.63, 111.87, 54.97, 50.44, 49.47, 46.79, 46.32, 43.13,
42.97, 42.01, 34.70, 33.57, 17.05. ES-MS calcd for
C30H35N3O3: 485.62, found: 486.4 (MH1).
1-{1-[(Benzylamino)methyl]-5(S)-[benzyl(methyl)amino]-
pentyl}-5(S)-methyl-4-p-methoxyphenethyltetrahydro-
2,3-diketopiperazine (6p). 1H NMR (500 MHz, DMSO-d6)
d 7.41±6.86 (m, 14H), 4.72 (m, 1H), 3.72 (s, 3H), 3.70 (m,
2H), 3.17 (m, 3H), 3.08 (m, 1H), 3.00 (m, 4H), 2.86±2.76
(m, 5H), 2.52 (s, 3H), 1.249 (m, 2H), 1.23 (m, 2H), 1.18 (m,
2H), 1.14 (m, 3H). ES-MS calcd for C35H46N3O3: 570.76,
found: 471.7 (MH1).
1-{1(S)-[(Benzylamino)methyl]-5-[benzyl(methyl)amino]-
pentyl}-5(S)-benzyl-4-phenethyltetrahydro-2,3-diketo-
piperazine (6i). 1H NMR (500 MHz, DMSO-d6) d 7.13±7.6
(m, 20H), 4.37 (m, 1H), 4.15±4.23 (m, 2H), 3.67±3.83 (m,
8H), 3.35 (m, 1H), 2.94±3.09 (m, 4H), 2.67±2.84 (m, 4H),
1.58 (m, 2H), 1.16 (m, 1H). 13C NMR (125 MHz, DMSO-
1-{1(S)-[(Benzylamino)methyl]-5-[benzyl(methyl)amino]-
pentyl}-5(S)-isopropyl-4-p-methoxyphenethyltetrahydro-
2,3-diketopiperazine (6q). 1H NMR (500 MHz, DMSO-d6)
d 7.53±6.81 (m, 14H), 4.72 (m, 1H), 3.97 (dd, Jˆ6.56,
13.30 Hz, 2H), 3.69 (d, Jˆ5.25 Hz, 2H), 3.35 (m, 2H),
3.14 (m, 2H), 3.06 (m, 3H), 2.99 (m, 4H), 2.78 (m, 2H),
2.59 (s, 3H), 1.77 (m, 1H), 1.69 (m, 2H), 1.45 (m, 2H), 1.29
(t, Jˆ6.68 Hz, 3H), 1.22 (m, 2H), 0.89 (d, Jˆ5.00 Hz, 3H),
0.84 (d, Jˆ5.00 Hz, 3H). ES-MS calcd for C38H52N4O3:
612.84, found: 613.7 (MH1).
²
It was found that the alkylation of the amide resin linkage dramatically
increases the acid sensitivity of the MBHA resin-bound peptide. The N-
alkylated MBHA resin could be cleaved with 70% TFA in DCM in 30 min.
This cleavage can be avoided through the subsequent use of Fmoc
chemistry.

3326
A. Nefzi et al. / Tetrahedron 56 (2000) 3319±3326
F. Tetrahedron Lett. 1998, 39, 2639±2642. (b) Szardenings, A. K.;
Supporting information
Burkoth, T. S.; Lu, H. H.; Tien, D. W.; Campbell, D. A.
Tetrahedron 1997, 53, 6573±6578. (c) Kowalski, J.; Lipton,
M. A. Tetrahedron Lett. 1996, 37, 5839±5842.
1
Listing of: (a) H NMR spectra of all compounds; (b) the
LC-MS for compounds 6a±x; and (c) HPLCs of diastereo-
isomers that do not coelute.
9. Early reports on the reduction of amides with diborane: (a)
Brown, H. C.; Heim, P. J. Org. Chem. 1973, 38, 912±916. (b)
Oh, I. H.; Yoon, N. M.; Gyoung, Y. S. Bull. Korean Chem. Soc.
1989, 10, 12±15.
Acknowledgements
10. (a) Cuervo, J. H.; Weitl, F.; Ostresh, J. M.; Hamashin, V. T.;
Hannah, A. L.; Houghten, R. A. In Peptides 94, Proceedings of the
23rd European Peptide Symposium; Maia, H. L. S., Ed., ESCOM:
Leiden, 1995; pp 465±466. (b) Kim, J. M.; Wilson, T. E.; Norman,
T. C.; Schultz, P. G. Tetrahedron Lett. 1996, 37, 5309±5312.
11. Ostresh, J. M.; Schoner, C. C.; Hamashin, V. T.; Nefzi, A.;
Meyer, J.-P.; Houghten, R. A. J. Org. Chem. 1998, 63, 8622±8623.
12. DoÈrner, B.; Husar, G. M.; Ostresh, J. M.; Houghten, R. A.
Bioorg. Med. Chem. 1996, 4, 709±714.
This research was supported by National Cancer Institute
Grant no. CA78040 (Houghten).
References
1. (a) Gallop, M. A.; Barret, R. W.; Dower, W. J.; Fodor, S. P. A.;
Gordon, E. M. J. Med. Chem. 1994, 37, 1233±1251. (b) Gordon,
E. M.; Barret, R. W.; Dower, W. J.; Fodor, S. P. A.; Gordon, E. M.
J. Med. Chem. 1994, 37, 1385±1401. (c) Thompson, L. A.; Ellman,
J. A. Chem. Rev. 1996, 96, 555±600. (d) Fruchtel, J. S.; Jung, G.
Angew. Chem., Int. Ed. Engl. 1996, 35, 17±42. (e) Hermkens,
P. H. H.; Ottenheijm, H. C. J.; Rees, D. C. Tetrahedron 1996,
52, 4527±4537. (f) Nefzi, A.; Ostresh, J. M.; Houghten, R. A.
Chem. Rev. 1997, 97, 449±472.
13. Nefzi, A.; Ostresh, J. M.; Houghten, R. A. Tetrahedron 1999,
55, 335±344.
14. Merri®eld, R. B. J. Am. Chem. Soc. 1963, 85, 2149±2150.
15. (a) Nefzi, A.; Ostresh. J. M.; Giulianotti, M.; Houghten, R. A.
J. Comb. Chem. 1999, 1, 195±198. (b) Nefzi, A.; Ostresh, J. M.;
Meyer, J.-P.; Houghten, R. A. Tetrahedron Lett. 1997, 38, 93±96.
2. Cui, C.-B.; Kakeya, H.; Osada, H. J. Antibiot. 1996, 49, 437±
534.
È
16. (a) Ostresh, J. M.; Husar, G. M.; Blondelle, S. E.; Dorner, B.;
Weber, P. A.; Houghten, R. A. Proc. Natl. Acad. Sci., USA 1994,
91, 11138±11141. (b) Houghten, R. A.; Blondelle, S. E.; Dooley,
3. (a) Charlton, P. A.; Faint, R. W.; Bent, F.; Bryans, J.; Chicarelli-
Robinson, I.; Mackie, I.; Machin, S.; Bevan, P. Thromb. Haemost.
1996, 75, 808±812. (b) Funabashi, Y.; Horiguchi, T.; Iinuma, S.;
Tanida, S.; Harada, S. J. Antibiot. 1994, 47, 1202±1208.
4. Barrow, C. J.; Musza, L. L.; Cooper, R. Bioorg. Med. Chem.
Lett. 1995, 5, 377±380.
È
T. C.; Dorner, B.; Eichler, J.; Ostresh, J. M. Mol. Diversity 1996, 2,
41±44.
17. Houghten, R. A.; Pinilla, C.; Appel, J. R.; Blondelle, S. E.;
Dooley, C. T.; Eichler, J.; Nefzi, A.; Ostresh, J. M. J. Med. Chem.
1999, 42, 3743±3778.
5. Gordon, D.; Steele J. BioMed. Chem. Lett. 1995, 5, 47±50.
ÃÁ
6. Krchnak, V.; Weichsel, A. S.; Cabel, D.; Lebl, M. Molecular
18. Houghten, R. A. Proc. Natl. Acad. Sci., USA 1985, 82, 5131±
5134.
Diversity and Combinatorial Chemistry: Libraries and Drug
Discovery; Chaiken, I. M., Janda, K. D., Eds.; American Chemical
Society: Washington, DC, 1996, pp 99±117.
19. Kaiser, E. T.; Colescott, R. L.; Blossinger, C. D.; Cook, P. I.
Anal. Biochem. 1970, 34, 595.
Ï
20. Krchnak, V.; Vagner, J.; Safar, P.; Lebl, M. Coll. Czech.
ÃÁ
Â
ÂÏ
7. Scott, B. O.; Seigmund, A. C.; Marlowe, C. K.; Pei, Y.; Spear,
K. L. Mol. Diversity 1995, 1, 125±128.
Chem. Commun. 1988, 53, 2542±2546.
21. Houghten, R. A.; Bray, M. K.; DeGraw, S. T.; Kirby, C. J. Int.
J. Pept. Protein Res. 1986, 27, 6763±6768.
8. Other reports on the solid-phase synthesis of 2,5-diketopiper-
azines: (a) Fresno, M.; Alsina, J.; Royo, M.; Barany, G.; Albericio,