Design of N-acylprolyltyrosine 'tripeptoid' analogues of neurotensin as potential atypical antipsychotic agents

Article abstract of DOI:10.1021/jm970217c

A series of N-acylprolyltyrosine amides was designed as tripeptoid analogues of neurotensin. The substituted dipeptides were tested in vivo for antidopamine activity by their ability to inhibit the apomorphine-induced climbing in mice and the dopamine-ind

Full text of DOI:10.1021/jm970217c

284
J . Med. Chem. 1998, 41, 284-290
Design of N-Acylp r olyltyr osin e “Tr ip ep toid ” An a logu es of Neu r oten sin a s
P oten tia l Atyp ica l An tip sych otic Agen ts
Tatiana A. Gudasheva,*,† Tatiana A. Voronina,‡ Rita U. Ostrovskaya,‡ Natalya I. Zaitseva,†
Nina A. Bondarenko,‡ Vera K. Briling,† Ludmila S. Asmakova,‡ Grigory G. Rozantsev,† and Sergei B. Seredenin‡
Institute of Pharmacology, Russian Academy of Sciences, Baltijskaya st. 8, Moscow 125315, Russia
Received March 31, 1997X
A series of N-acylprolyltyrosine amides was designed as tripeptoid analogues of neurotensin.
The substituted dipeptides were tested in vivo for antidopamine activity by their ability to
inhibit the apomorphine-induced climbing in mice and the dopamine-induced extrapolatory
behavior impairment in rats. The N-acylprolyltyrosine amides structure-activity relationships
have indicated the size of the N-acyl group and the configuration of amino acids that are
important for the activity. We found that the bioactivity has been increased dramatically when
the n-hydrocarbon chain on the N-acyl group was increased from four to five carbon atoms.
The activity seems to reside exclusively in the L-Tyr diastereomers. All of the compounds tested
were inactive in the cataleptogenic action and did not exhibit the acute toxicity even at doses
500-1000 times higher than ED50 in climbing test. On this basis, the N-acylprolyltyrosine
amides could potentially be a novel class of atypical antipsychotic agents.
Sch em e 1. Stepwise Design of Therapeutically Useful
“Peptoids” from Nonpeptide Drugs and Putative
Endogenous Neuropeptides
In tr od u ction
Neurotensin (NT, pGlu1-Leu2-Tyr3-Glu4-Asn5-Lys6-
Pro7-Arg8-Arg9-Pro10-Tyr11-Ile12-Leu13) is an endogenous
tridecapeptide1 that has a broad range of biological
effects including hypotension, analgesia, and increase
of vascular permeability. In addition, NT has been
shown to possess pharmacological properties similar to
those of dopamine antagonists that are effective anti-
psychotics.2-6 Closer scrutiny of the behavioral data
revealed even closer similarities of centrally adminis-
trated NT and atypical antipsychotic drugs. For ex-
ample, direct injection of NT into the nucleus accum-
bens, a major terminal site of the mesolimbic DA
system, attenuates D-amphetamine-induced hyperactiv-
ity, whereas direct injection of NT into the nucleus
caudatus, a major terminal site of the nigroneostriatal
DA system, does not attenuate D-amphetamine-induced
stereotypic behavior. This finding suggests that NT
might selectively modulate mesolimbicortical DA neu-
rons while exerting few if any effects on the nigroneo-
striatal DA systemsa profile desirable in drug devel-
opment because of the presumed absence of both
extrapyramidal side effect and tardive dyskinesia li-
ability.7 Thus, NT analogues could potentially be a novel
class of neuroleptics (antipsychotics), which could re-
place current anti-dopamine therapies.
Small di-/tripeptides and their derivatives could be
the most perspective drugs among these NT analogues.
These peptides are more available and less polyfunc-
tional in comparison with polypeptides. Besides, small
peptides in some cases are more enzymatically stable
and hence could be orally active.
We now report the design, synthesis, and pharmaco-
logical properties of the substituted prolyltyrosines,
“tripeptoid” analogues of NT. The dipeptide lead was
designed using a drug-based peptide design approach,
which has as a ground the hypothesis that some
psychotropic exogenous drugs are the ligands of specific
neuropeptide receptors. Our approach to the design of
peptide analogues of nonpeptide drugs is outlined in
Scheme 1. Previously we have developed this approach
for the design of dipeptide analogues of the classical
nootropic piracetam, pyroglytamyl-containing dipep-
* To whom correspondence should be addressed. E-mail: uusolari@
solaris.msk.ru. Fax: +7(095)1511261.
† Department of Medicinal Chemistry.
‡ Department of Pharmacology.
X Abstract published in Advance ACS Abstracts, December 15, 1997.
S0022-2623(97)00217-3 CCC: $15.00 © 1998 American Chemical Society
Published on Web 01/07/1998

“Tripeptoid” Analogues of Neurotensin
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 3 285
Ta ble 1. Antagonism of Apomorphine-Induced Mice Climbing
of Prolyltyrosines
Sch em e 2
O
N
R
C
OH
CH CH2
CONH2
NH
R-(X)-Pro-(Y)-Tyr-NH2
a
compd
R
X
Y
ED50, mg/kg ip
10
11
13
(CH3)3COC(O)
(CH3)3COC(O)
C6H5CH2C(O)
L
D
L
L
L
L
2.5 (2.0-3.1)
0.4 (0.3-0.5)
2.2 (1.7-2.8)
3.4 (po)
14
15
16
17
18
19
20
21
C6H5CH2C(O)
C6H5CH2C(O)
CH3(CH2)3C(O)
CH3(CH2)4C(O)
D
L
L
L
L
L
D
L
L
D
L
L
L
L
L
D
4.9 (3.2-75)
>8
3.2 (2.1-5.1)
0.5 (0.4-0.7)
0.5 (0.4-0.7)
3.4 (2.2-5.3)
3.6 (2.7-4.7)
CH3(CH2)7C(O)
H
H
H
>16
17.5 (11.3-27.1)
(-)-sulpiride
a
ED50 (doses required for 50% of maximal response) were
calculated on percent antagonism; 95% confidence limits are
included in parentheses.
treatment of corresponding BOC derivatives with tri-
fluoroacetic acid. Diastereomeric purity of the prepared
1
compounds was not below 98% according to H-NMR
spectroscopy.
P h a r m a cology
Pharmacological results are presented in Tables 1 and
2. As an indication of potential antipsychotic activity,
the present compounds were evaluated in vivo for their
ability to antagonize the apomorphine-induced climbing
response in mice.15 Most of the substances under study
were able to antagonize this climbing. For some of the
active compounds, an additional behavioral test associ-
ated with antipsychotic efficacy, dopamine dependent
extrapolatory behavior paradigm,16 was used. This test
measures the ability of rats placed into a cylinder
plunged in water to solve an “extrapolatory task”. The
rats treated with L-DOPA lost this ability. The typical
and atypical neuroleptics restored the ability of L-DOPA-
treated rats to escape from an acute stress situation.
All of the compounds tested demonstrated antidopamine
activity in this test. The tendency to induce catalepsy
in rats was used as an indication of the propensity to
cause extrapyramidal side effects.17 All of the com-
pounds tested were inactive in this cataleptogenic action
even in doses 500-1000 times higher than the ED50 in
the climbing test. The dipeptides studied did not exhibit
the acute toxicity in doses up to 500-1000 mg/kg ip
(experiments on albino male mice).
tides, bridging the gap between piracetam and the major
metabolite of the memory peptide vasopressin.8-10
The structure of the atypical benzamide neuroleptic,
sulpiride, was used as a starting point for the present
drug-based peptide design. Sulpiride [5-(aminosulfo-
nyl)-N-[(1-ethyl-2-pyrrolidinyl)methyl]-2-methoxybenza-
mide] is a proven antipsychotic agent,11 and its action
is associated with a low incidence of extrapyramidal
disturbances. In this respect sulpiride has a certain
similarity with NT. We assumed that sulpiride acts not
only on D-2 dopamine receptors but also on NT recep-
tors, which are known as able to modulate dopaminergic
system.12
Ch em istr y
The general synthetic route for prolyltyrosines is
presented in Scheme 2. N-Acylprolines were prepared
by acylation of proline with the appropriate acyl chloride
according to Schotten-Baumann; BOC-prolines were
synthesized using di-tert-butyl dicarbonate according to
Pozdnev.13 A coupling of N-acylproline with the corre-
sponding ester of tyrosine was carried out by the mixed
anhydride method using isobutyl chloroformate in the
presence of N-ethylmorpholine according to Anderson.14
Amides of dipeptides were synthesized by ammonolysis
of the corresponding esters. Unsubstituted prolylty-
rosines were obtained as its trifluoroacetic salts by
Resu lts a n d Discu ssion
The structure of sulpiride contains a pyrrolidine ring
and an electronegatively substituted phenyl group
separated by an amide group. From this we have
assumed that sulpiride is a structure analogue of the
-Pro-Tyr- dipeptide fragment of some polypeptides. The
structure of this dipeptide consists of the pyrrolidine-
containing amino acid Pro and the aromatic amino acid

286 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 3
Gudasheva et al.
Ta ble 2. In Vivo Characterization of Some Prolyltyrosines, Sulpiride, and Other Neuroleptics
ED50 (ip), mg/kg
LD50 (ip), mg/kg
acute tox in mice
compound
13
17
19
sulpiride
remoxipride
clozapine
haloperidol
apoma
dopab
catac
2.2 (1.7-2.8)
0.5 (0.4-0.7)
3.4 (2.2-5.3)
17.5 (11.3-27.1)
1.2 (0.5-2.4)
26.2 (9.4-73.2)
0.1 (0.05-0.16)
1.5 (1.2-2.0)
0.4 (0.3-0.6)
>1000
>1000
>500
>500
250
>500
1.6 (1.2-2.1)
>500
16.5 (9.11-24.3)
0.31 (0.29-0.35)
20 (18-27)
g120
8.7d
302d
>32e
0.14 (0.10-0.20)
0.5 (0.4-0.6)f
35g
a
b
Inhibition of apomorphine-induced climbing in mice. Antagonism of extrapolatory behavior impairement induced by DOPA in rats.
c Induction of catalepsy in rats. Value taken from Florvall and Ogren.23 e Value taken from Howard et al.24 f Value taken from Henning
d
et al.25 Value taken from de Paulis et al.26
g
F igu r e 1. Superposition of the (S)-sulpiride (open lines) and
L-Pro-L-Tyr-NH2 (solid lines).
with the OH-substitution Tyr. The simplest model of
such fragment is dipeptide prolyltyrosine amide (Pro-
Tyr-NH2).
O
CH2
SO2NH2
NH
CH3O
N
F igu r e 2. Superposition of the neurotensin (9-13) and its
“tripeptoid” analogue N-caproyl-L-Pro-L-Tyr-NH2 (bold lines).
The type I â-turn conformation is depicted.
(-) Sulpiride
OH
O
CONH2
NH
which could imitate the hydrophobic side chain of Leu13
in the hypothetical â-turn conformation of NT(8-13)
(see Figure 2). In this case the type I â-turn conforma-
tion was used because the Pro-Tyr fragment has better
overlay with sulpiride than that in the type II â-turn
conformation. Thus N-acylprolyltyrosine amides may
be considered as the “tripeptoid” analogues of NT.
In fact, introduction of the N-acyl moiety into dipep-
tide Pro-Tyr-NH2 resulted in increased or at least
conserved activity in the apomorphine climbing test in
mice (cf. 10, 13, 16-18, Table 1). The N-acylprolylty-
rosine amides SAR have indicated the size of N-acyl
group that is critical for the activity increase. The
activity of N-valerylprolyltyrosine amide (cf. 16) is the
same as that of nonsubstituted H-Pro-Tyr-NH2 (cf. 19).
The lengthening of N-valeryl moiety by one CH2 group
(N-caproyl-Pro-Tyr-NH2, cf. 17) results in the increased
activity. Further augmentation of N-caproyl moiety by
three CH2 groups did not change the activity (cf. 18).
N-BOC-prolyltyrosine amide (cf. 10) and N-pheny-
lacetylprolyltyrosine amide (cf. 13) with N-acyl moiety
shorter than N-valeryl have demonstrated activity at
the level of nonsubstituted Pro-Tyr-NH2.
N
H
L-Pro-L-Tyr-NH2
Superimposition of L-Pro-L-Tyr-NH2 and (S)-sulpiride
(using of Dreiding molecular models) revealed the
overlap of pyrrolidine cycles and phenyl rings of both
molecules in unstrained conformations (see Figure 1).
Dipeptide Pro-Tyr-NH2 has been shown to inhibit apo-
morphine-induced climbing in mice without catalepto-
genic activity (Table 2). This result supports our
assumption about the peptidergic mechanism of sulpir-
ide action. The sequence Pro-Tyr corresponds to the
neurotensin fragment NT(10-11), which is present in
the major biologically active metabolite of neurotensin,
NT(8-13).
It is known that the replacement of the Pro10-Tyr11
sequence in NT(8-13) with â-turn mimetics has not
resulted in activity loss that has suggested the presence
of a â-turn-like structure.18-20 NT(8-13) structure-
activity relationships have revealed the residues that
are critical for receptor recognition. For example, an
alanine scan of NT(8-13) suggested21 the relative
importance of the individual amino acid side chains to
These findings are in a good agreement with our
assumption that a sufficiently long N-acyl moiety is able
to imitate the Leu13 side chain which is important to
reveal the activity. The above results are also consistent
the data reported elsewhere20 that NT(8-13) is biologi-
cally active being in the â-turn conformation. The
be as follows: Leu13 > Tyr11 . Ile12 > Arg9 > Pro10
>
Arg8. In order to increase the Pro-Tyr-NH2 dipeptide
activity, we included into its structure an N-acyl group,

“Tripeptoid” Analogues of Neurotensin
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 3 287
-48.2° (c 0.39, CHCl3): Rf 0.75 (CHCl3/C2H5OH, 9:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 1.08 (t,
CH3CH2O, 3H, minor conformer), 1.11 (t, CH3CH2O, 3H, major
conformer), 1.15-1.59 (m, CâH2-CγH2 Pro, 4H), 1.21 (s, (CH3)3C,
9H, major conformer), 1.37 (s, (CH3)3C, 9H, minor conformer),
2.76-3.0 (m, CâH2 Tyr, 2H), 3.15-3.35 (m, CδH2 Pro, 2H), 4.0
(q, CH3CH2O, 2H, minor conformer), 4.03 (q, CH3CH2O, 2H,
major conformer), 4.11 (m, CRH Pro, 1H), 4.31 (m, CRH Tyr,
1H, minor conformer), 4.36 (m, CRH Tyr, 1H, major conformer),
6.63, 7.0 (each m, AA′BB′ system, C6H4 Tyr, 4H), 8.10 (d, NH
Tyr, 1H, minor conformer), 8.15 (d, NH Tyr, 1H, major
conformer), 9.23 (s, OH, 1H, major conformer), 9.25 (s, OH,
1H, minor conformer). Anal. (C21H30N2O6) C, H, N.
results obtained suggest the distance between the
nitrogen atom of Pro10 and the side chain Leu13 to be
not less than six σ-bonds.
To evaluate the influence the amino acid residue
configurations exerts over the activity of compounds
studied, the diastereomers of both Pro-Tyr NH2 and its
N-acyl derivatives were synthesized. The replacement
of L-Tyr by D-Tyr (cf. 15, 21) resulted in the loss of
activity. In contrast, replacement of L-Pro by D-Pro (cf.
11, 14, 20) resulted in activity conservation. This
demonstrates that the effect of the dipeptides studied
is stereoselective in respect to tyrosine, but not proline.
It is concluded that the N-acylprolyltyrosine amides
which carry an N-acyl group longer than N-valeryl and
have the L-configuration of Tyr and the L- or D-config-
uration of Pro constitute a new type of potent antipsy-
chotic agents with the ability to inhibit the apomorphine-
induced climbing without production catalepsy even at
high doses.
N-(ter t-Bu toxyca r bon yl)-D-p r olyl-L-tyr osin e m eth yl es-
ter (2): yield 86% (after column chromatography); oil; [R]20
D
+72.0° (c 0.23, CHCl3); Rf 0.53 (CHCl3/C2H5OH, 11:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 1.29 (s,
(CH3)3C, 9H, major conformer), 1.39 (s, (CH3)3C, 9H, minor
conformer), 1.41-2.09 (m, CâH2-CγH2 Pro, 4H), 2.67-2.99 (m,
CâH2 Tyr, 2H), 3.11-3.35 (m, CδH2 Pro, 2H), 3.59 (s, CH3O,
3H, major conformer), 3.62 (s, CH3O, 3H, minor conformer),
4.05 (dd, CRH Pro, 1H, major conformer), 4.09 (dd, CRH Pro,
1H, minor conformer), 4.46 (m, CRH Tyr, 1H), 6.64, 6.97 (each
m, AA′BB′ system, C6H4 Tyr, 4H); 8.09 (d, J ) 8.77 Hz, NH,
1H, minor conformer), 8.26 (d, J ) 8.05 Hz, NH, 1H, major
conformer), 9.22 (s, OH, 1H). Anal. (C20H28N2O6) C, H, N.
Exp er im en ta l Section
Molecu la r Mod elin g. The structures demonstrating the
overlay of sulpiride with the prolyltyrosine peptide fragments
were obtained by handling with Molecular Dreiding stereo-
models (W. Buchi Scientific Apparatus Flawil, Switzerland).
N-(ter t-Bu toxyca r bon yl)-L-p r olyl-D-tyr osin e m eth yl es-
ter (3): yield 87% (after column chromatography); oil; [R]20
D
-72.0° (c 0.23, CHCl3); Rf 0.60 (CHCl3/C2H5OH, 11:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 1.29 (s,
(CH3)3C, 9H, major conformer), 1.39 (s, (CH3)3C, 9H, minor
conformer), 1.41-2.09 (m, CâH2-CγH2 Pro, 4H), 2.67-2.99 (m,
CâH2 Tyr, 2H), 3.11-3.35 (m, CδH2 Pro, 2H), 3.59 (s, CH3O,
3H, major conformer), 3.62 (s, CH3O, 3H, minor conformer),
4.05 (dd, CRH Pro, 1H, major conformer), 4.09 (dd, CRH Pro,
1H, minor conformer), 4.46 (m, CRH Tyr, 1H), 6.64, 6.97 (each
m, AA′BB′ system, C6H4 Tyr, 4H), 8.09 (d, NH, 1H, minor
conformer), 8.26 (d, NH, 1H, major conformer), 9.22 (s, OH,
1H). Anal. (C20H28N2O6) C, H, N.
The following sequence of operations was used. First, the
hydrophobic regions of the molecules (the phenyl groups and
the pyrrolidine rings) were superimposed. Second, the hydro-
philic parts were fitted: the positive charged nitrogen atoms
of the pyrrolidine rings were fitted precisely; the polar OH-
group of the Tyr side chain and the sulpiride SO2NH2 group
were superimposed; the sulpiride amide group and the peptide
group were placed closely; the corresponding N-H and CdO
bond vectors were oriented parallel. Then, the conformations
have been selected that demonstrated the least strain of the
valent angles and bonds. Finally, the conformations with the
minimum unsuperimposed molecular volumes were chosen.
N-(P h en yla cetyl)-L-p r olyl-L-tyr osin e m eth yl ester (4):
yield 75% (after crystallization from methanol-ether, 1:2, v/v);
mp 146-148 °C; [R]20 -48.0° (c 0.15, DMF); Rf 0.57 (CHCl3/
Ch em istr y. Melting points were determined on a capillary
melting point apparatus in open capillary tubes and are
uncorrected. The structures of the compounds were confirmed
D
CH3OH, 9:1); NMR (DMSO-d6) two conformations evident in
spectrum δ 1.56-2.18 (m, CâH2-CγH2 Pro, 4H), 2.80-2.90 (m,
CâH2 Tyr, 2H, major conformer), 2.82, 3.02 (each dd, CâH2 Tyr,
2H, minor conformer), 3.20, 3.35 (each d, J ) 15.7 Hz, C6H5-
CH2, 2H, major conformer), 3.40-3.50 (m, CδH2 Pro, 2H), 3.55
(s, CH3O, 3H, major conformer), 3.63 (s, CH3O, 3H, minor
conformer), 3.64 (s, C6H5CH2, 2H, minor conformer), 4.35 (m,
CRH Pro, CRH Tyr major conformer, 2H), 4.54 (m, CRH Tyr,
1H, minor conformer), 6.61, 7.01 (each m, AA′BB′ system, C6H4
Tyr, 4H, minor conformer), 6.64, 6.96 (each m, AA′BB′ system,
C6H4 Tyr, 4H, major conformer), 7.0-7.35 (m, C6H5CH2, 5H),
8.13 (d, J ) 7.68 Hz, NH Tyr, 1H, major conformer), 8.54 (d,
J ) 8.20 Hz, NH Tyr, 1H, minor conformer), 9.17 (s, OH, 1H,
minor conformer), 9.21 (s, OH, 1H, major conformer). Anal.
(C23H26N2O5) C, H, N.
1
by elemental analysis and H NMR spectroscopy. Microanaly-
ses agree with calculated values within (0.4%. The NMR
spectra were obtained on a Bruker AC-250 spectrometer using
tetramethylsilane as an internal standard. The NMR peaks
were designated as follows: s, singlet; d, doublet; t, triplet; q,
quartet; m, multiplet. Specific optical rotations were recorded
by automatic polarimeter Perkin-Elmer 241. The TLC was
carried out on Merck silica gel 60 F 254 plates, and spots were
developed in an iodine chamber or under UV light. For column
chromatography, Merck silica gel 60, 230-400 mesh was used.
If required, all solvents used in reaction mixtures were dried
and purified by standard procedures.
Gen er a l Syn th etic Meth od for th e P r ep a r a tion of
N-Acylp r olyltyr osin e Ester s. To a well-stirred solution of
N-acylproline (10.0 mmol)13,22 in 100 mL of chloroform were
added N-ethylmorpholine (10.0 mmol) and isobutyl chlorofor-
mate (10.0 mmol) dropwise at -10 °C. After 2 min a mixture
of the hydrochloride methyl or ethyl ester of tyrosine (10.0
mmol) and N-ethylmorpholine (10.0 mmol) in dimethylforma-
mide (20 mL) was added. Stirring continued for 30 min at
-5 °C, and then the mixture was allowed to stand for 1 h.
The precipitate was separated by filtration; the solvent was
evaporated in vacuo; the residue was dissolved in chloroform,
washed with 5% solution of NaHCO3, water, 1 N HCl, and
water, and dried with Na2SO4; and the solvent was evaporated
to dryness. The residue was purified by column chromatog-
raphy using chloroform as eluent or crystallization from
suitable solvent.
N-(P h en yla cet yl)-D-p r olyl-l-t yr osin e et h yl est er (5):
yield 66% (after trituration with hexane); mp 186-188 °C;
[R]20 +49.5° (c 0.3, CH3OH); Rf 0.56 (CHCl3/CH3OH, 9:1);
D
NMR (DMSO-d6) two conformations evident in spectrum δ 1.09
(t, J ) 7.04 Hz, CH3CH2O, 3H, minor conformer), 1.13 (t, J )
7.04 Hz, CH3CH2O, 3H, major conformer), 1.48-2.2 (m, CâH2-
CγH2 Pro, 4H), 2.7-3.05 (m, CâH2 Tyr, 2H), 3.18-3.4 (m, CδH2
Pro, 2H), 3.65 (s, C6H5CH2, 2H), 4.04 (q, CH3CH2O, 2H), 4.27
(dd, CRH Pro, 1H, minor conformer), 4.33 (dd, CRH Pro, 1H,
major conformer), 4.40 (m, CRH Tyr, 1H, major conformer),
4.44 (m, CRH Tyr, 1H, minor conformer), 6.62, 7.01 (each m,
AA′BB′ system, C6H4 Tyr, 4H, minor conformer), 6.63, 6.95
(each m, AA′BB′ system, C6H4 Tyr, 4H, major conformer), 7.1-
7.34 (m, C6H5CH2, 5H), 8.15 (d, J ) 8.41 Hz, NH, 1H, major
conformer), 8.59 (d, J ) 8.02 Hz, NH, 1H, minor conformer),
9.18 (s, OH, 1H, major conformer), 9.19 (s, OH, 1H, minor
conformer). Anal. (C24H28N2O5) C, H, N.
N-(ter t-Bu toxyca r bon yl)-L-p r olyl-L-tyr osin e eth yl es-
ter (1): yield 82% (after column chromatography); oil; [R]20
D

288 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 3
N-(P h en yla cet yl)-L-p r olyl-D-t yr osin e et h yl est er (6):
Gudasheva et al.
tallization or column chromatography using chloroform-
yield 59% (after trituration with ether); mp 187-188 °C; [R]20
ethanol (9:1, v/v) as eluent.
D
-58.5° (c 0.3, CH3OH); Rf 0.5 (CHCl3/CH3OH, 9:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 1.10 (t,
J ) 7.04 Hz, CH3CH2O, 3H, minor conformer), 1.14 (t, J )
7.04 Hz, CH3CH2O, 3H, major conformer), 1.5-2.2 (m, CâH2-
CγH2 Pro, 4H), 2.7-3.05 (m, CâH2 Tyr, 2H), 3.2-3.4 (m, CγH2
Pro, 2H), 4.04 (q, CH3CH2O, 2H, minor conformer), 4.05 (q,
CH3CH2O, 2H, major conformer), 4.31 (dd, CRH Pro, 1H, minor
conformer), 4.38 (dd, CRH Pro, 1H, major conformer), 4.40 (m,
CRH Tyr, 1H, major conformer), 4.45 (m, CRH Tyr, 1H, minor
conformer), 6.45, 6.62 (each m, AA′BB′ system, C6H4 Tyr, 4H,
major conformer), 6.97, 7.01 (each m, AA′BB′ system, C6H4
Tyr, 4H, minor conformer), 7.09-7.35 (m, C6H5CH2, 5H), 8.15
(d, J ) 8.4 Hz, NH, 1H, major conformer), 8.59 (d, J ) 8.02
Hz, NH, 1H, minor conformer), 9.25 (s, OH, 1H, major
conformer), 9.26 (s, OH, 1H, minor conformer). Anal.
(C24H28N2O5) C, H, N.
N-Va ler yl-L-p r olyl-L-tyr osin e m eth yl ester (7): yield 51%
(after column chromatography); oil; [R]20D -58° (c 0.5, CHCl3);
Rf 0.77 (n-C4H9OH/AcOH/H2O, 5:1:2). NMR (DMSO-d6) two
conformations evident in spectrum δ 0.82 (t, CH3(CH2)3, 3H,
minor conformer), 0.88 (t, CH3(CH2)3, 3H, major conformer),
0.98-2.17 (m, CâH2-CγH2 Pro, 4H), 0.98-2.0 (m, CH3(CH2)2,
4H), 2.23 (t, C3H7CH2C(O), 2H), 2.70-3.04 (dd, CâH2 Tyr, 2H),
3.2-3.5 (m, CδH2 Pro, 2H), 3.56 (s, CH3O, 3H, major con-
former), 3.63 (s, CH3O, 3H, minor conformer), 4.25 (dd, CRH
Pro, 1H, minor conformer), 4.33 (dd, CRH Pro, 1H, major
conformer), 4.33 (m, CRH Tyr, 1H, major conformer), 4.47 (m,
CRH Tyr, 1H, minor conformer), 6.64, 6.99 (each m, AA′BB′
system, C6H4 Tyr, 4H, minor conformer), 6.65, 6.98 (each m,
AA′BB′ system, C6H4 Tyr, 4H, major conformer), 8.07 (d, NH
Tyr, 1H, major conformer), 8.4 (d, NH Tyr, 1H, minor
conformer), 9.18 (s, OH Tyr, 1H, major conformer), 9.2 (s, OH
Tyr, 1H, minor conformer). Anal. (C20H28N2O5) C, H, N.
N-(ter t-Bu toxycar bon yl)-L-pr olyl-L-tyr osin e am ide (10):
yield 60% (after column chromatography); mp 72-75 °C; [R]20
D
-49.7° (c 0.38, CH3OH); Rf 0.33 (CHCl3/C2H5OH, 9:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 1.22 (s,
(CH3)3C, 9H, major conformer), 1.39 (s, (CH3)3C, 9H, minor
conformer), 1.56-2.10 (m, CâH2-CγH2 Pro, 4H), 2.6-3.0 (m,
CâH2 Tyr, 2H), 3.2-3.4 (m, CδH2 Pro, 2H), 4.05 (m, CRH Pro,
1H), 4.28 (m, CRH Tyr, 1H, minor conformer), 4.35 (m, CRH
Tyr, 1H, major conformer), 6.62, 6.98 (each m, AA′BB′ system,
C6H4 Tyr, 4H, major conformer), 6.62, 7.0 (each m, AA′BB′
system, C6H4 Tyr, 4H, minor conformer), 7.0, 7.30 (each s, NH2,
2H, minor conformer), 7.08, 7.15 (each s, NH2, 2H, major
conformer), 7.70 (d, NH, 1H, major conformer), 7.75 (d, NH,
1H, minor conformer), 9.18 (s, OH, 1H, major conformer), 9.20
(s, OH, 1H, minor conformer). Anal. (C19H27N3O5) C, H, N.
N-(ter t-Bu toxycar bon yl)-D-pr olyl-L-tyr osin e am ide (11):
yield 40% (after column chromatography); mp 76-78 °C; [R]20
D
+33.2° (c 0.3, CHCl3), Rf 0.36 (CHCl3/C2H5OH, 9:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 1.28 (s,
(CH3)3C, 9H, major conformer), 1.38 (s, (CH3)3C, 9H, minor
conformer), 1.55-2.08 (m, CâH2-CγH2 Pro, 4H), 2.64, 2.84 (each
dd, CâH2 Tyr, 2H, major conformer), 2.64, 2.98 (each dd, CâH2
Tyr, 2H, minor conformer), 3.10-3.32 (m, CδH2 Pro, 2H), 4.05
(dd, CRH Pro, 1H), 4.26 (m, CRH Tyr, 1H, minor conformer),
4.46 (m, CRH Tyr, 1H, major conformer), 6.62, 6.98 (each m,
AA′BB′ system, C6H4 Tyr, 4H), 7.05, 7.37 (each s, NH2, 2H,
major conformer), 7.16, 7.28 (each s, NH2, 2H, minor con-
former), 7.88 (d, NH Tyr, J ) 8.9 Hz, 1H, major conformer),
8.12 (d, NH Tyr, J ) 8.6 Hz, 1H, minor conformer), 9.15 (c,
OH Tyr, 1H). Anal. (C19H27N3O5) C, H, N.
N-(ter t-Bu toxycar bon yl)-L-pr olyl-D-tyr osin e am ide (12):
yield 46% (after column chromatography); mp 79-80 °C; [R]20
D
-34.0° (c 0.3, CHCl3); Rf 0.36 (CHCl3/C2H5OH, 9:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 1.28 (s,
(CH3)3C, 9H, major conformer), 1.38 (s, (CH3)3C, 9H, minor
conformer), 1.55-2.08 (m, CâH2-CγH2 Pro, 4H), 2.64, 2.84 (each
dd, CâH2 Tyr, 2H, major conformer), 2.64, 2.98 (each dd, CâH2
Tyr, 2H, minor conformer), 3.10-3.32 (m, CδH2 Pro, 2H), 4.05
(dd, CRH Pro, 1H), 4.26 (m, CRH Tyr, 1H, minor conformer),
4.46 (m, CRH Tyr, 1H, major conformer), 6.62, 6.98 (each m,
AA′BB′ system, C6H4 Tyr, 4H), 7.05, 7.37 (each s, NH2, 2H,
major conformer), 7.16, 7.28 (each s, NH2, 2H, minor con-
former), 7.88 (d, NH Tyr, J ) 8.9 Hz, 1H, major conformer),
8.12 (d, NH Tyr, J ) 8.6 Hz, 1H, minor conformer), 9.15 (c,
OH Tyr, 1H). Anal. (C19H27N3O5) C, H, N.
N-Ca p r oyl-L-p r olyl-L-tyr osin e m eth yl ester (8): yield
54% (after crystallization from EtOAc); mp 115-116 °C; [R]20
D
-58° (c 0.4, CHCl3); Rf 0.56 (CHCl3/C2H5OH, 9:1); NMR
(DMSO-d6) two conformations evident in spectrum δ 0.84 (t,
(CH3(CH2)4, 3H, minor conformer), 0.87 (t, CH3(CH2)4, 3H,
major conformer), 1.02-2.20 (m, CâH2-CγH2 Pro, CH3(CH2)4,
12H), 2.7-3.1 (m, CâH2 Tyr, 2H), 3.2-3.45 (m, CδH2 Pro, 2H),
3.56 (s, CH3O, 3H, major conformer), 3.63 (s, CH3O, 3H, minor
conformer), 4.26 (m, CRH Pro, 1H, minor conformer), 4.35 (m,
CRH Pro, 1H, major conformer), 4.35 (m, CRH Tyr, 1H, major
conformer), 4.48 (m, CRH Tyr, 1H, minor conformer), 6.65, 7.00
(each m, AA′BB′ system, C6H4 Tyr, 4H, minor conformer), 6.66,
6.98 (each m, AA′BB′ system, C6H4 Tyr, 4H, major conformer),
8.04 (d, NH Tyr, 1H, major conformer), 8.42 (d, NH Tyr, 1H,
minor conformer), 9.23 (s, OH Tyr, 1H, major conformer), 9.26
(s, OH Tyr, 1H, minor conformer). Anal. (C21H30N2O5) C, H,
N.
N-(P h en yla cetyl)-L-p r olyl-L-tyr osin e a m id e (13): yield
75% (after crystallization from methanol-ether, 1:2, v/v); mp
201-203 °C; [R]20 -89.6° (c 0.3, DMF); Rf 0.75 (CHCl3/C2H5-
D
OH, 9:3); NMR (DMSO-d6) two conformations evident in
spectrum δ 1.54-2.16 (m, CRH2-CγH2 Pro, 4H), 2.69, 2.92 (each
dd, CâH2 Tyr, 2H, minor conformer), 2.74, 2.94 (each dd, CâH2
Tyr, 2H, major conformer), 3.40-3.65 (m, CδH2 Pro, 2H), 3.66,
3.71 (each d, AB system, C6H5CH2, 2H), 4.21 (dd, CRH Pro,
1H, major conformer), 4.38 (dd, CRH Pro, 1H, minor con-
former), 4.28 (m, CRH Tyr, 1H, major conformer), 4.51 (m, CRH
Tyr, 1H, minor conformer), 6.57, 7.05 (each m, AA′BB′ system,
C6H4 Tyr, 4H, minor conformer), 6.62, 6.95 (each m, AA′BB′
system, C6H4 Tyr, 4H, major conformer), 7.0-7.5 (m, C6H5-
CH2, NH2, 7H), 7.71 (d, NH, 1H, major conformer), 8.20 (d,
NH, 1H, minor conformer), 9.09 (s, OH, 1H, minor conformer);
9.14 (s, OH, 1H, major conformer). Anal. (C22H25N3O4) C, H,
N.
N-Non a n oyl-L-p r olyl-L-t yr osin e et h yl est er (9): yield
60% (after trituration with ether); mp 115-117 °C; [R]20
D
-50.4° (c 0.4, CHCl3); Rf 0.9 (CHCl3/CH3OH, 9:1); NMR
(CDCl3) two conformations evident in spectrum δ 0.87 (t, CH3-
(CH2)7, 3H), 1.26 (m, CH3(CH2)5, CH3CH2O, 13H), 1.5-1.7 (m,
C6H13CH2CH2, 2H), 1.75-2.06 (m, CâH2-CγH2 Pro, 4H), 2.28
(t, C7H15CH2C(O), 2H), 2.90, 3.11 (each dd, CâH2 Tyr, 2H,
minor conformer), 2.93, 3.09 (each dd, CâH2 Tyr, 2H, major
conformer), 3.30-3.54 (m, CδH2 Pro, 2H), 4.16 (q, CH3CH2O,
2H), 4.55 (dd, CRH Pro, 1H), 4.77 (m, CRH Tyr, 1H), 6.28 (d,
NH Tyr, 1H, minor conformer), 6.67, 6.97 (each m, AA′BB′
system, C6H4 Tyr, 4H, minor conformer), 6.76, 6.93 (each m,
AA′BB′ system, C6H4 Tyr, 4H, major conformer), 7.27 (s, OH
Tyr, 1H); 7.28 (d, NH Tyr, 1H, major conformer). Anal.
(C25H38N2O5) C, H, N.
N-(P h en yla cetyl)-D-p r olyl-L-tyr osin e a m id e (14): yield
54% (after crystallization from methanol-ether, 1:2, v/v); mp
195-197 °C; [R]20D +61.7° (c 0.4, CH3OH); Rf 0.3 (CHCl3/CH3-
OH, 9:1); NMR (DMSO-d6) two conformations evident in
spectrum in ratio 4:1 δ 1.4-2.1 (m, CâH2-CγH2 Pro, 4H), 2.61
(dd, J ) 14.1 Hz, J ) 10.8 Hz, CâH2 Tyr, 1H, major conformer),
2.68 (dd, J ) 13.9 Hz, J ) 10.8 Hz, CâH2 Tyr, 1H, minor
conformer), 2.94 (dd, J ) 13.9 Hz, J ) 4.0 Hz, CâH2 Tyr, 1H,
minor conformer), 3.03 (dd, J ) 14.1 Hz, J ) 3.7 Hz, CâH2
Tyr, 1H, major conformer), 3.3-3.5 (m, CδH2 Pro, 2H), 3.65
Gen er a l Syn th etic Meth od for th e P r ep a r a tion of
N-Acylp r olyltyr osin e Am id es. A solution of N-acylprolyl-
tyrosine methyl or ethyl ester (5 mmol) in 50 mL of methanol
was cooled to 0 °C. NH3 (dried through NaOH trap) was then
bubbled through the solution for 30 min. The solution was
maintained at room temperature for 48 h. Methanol was
evaporated in vacuo, and the residue was purified by recrys-

“Tripeptoid” Analogues of Neurotensin
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 3 289
(s, C6H5CH2, 2H), 4.2-4.3 (m, CRH Pro, CRH Tyr, 2H, major
conformer), 4.4-4.5 (m, CRH Pro, CRH Tyr, 2H, minor con-
former), 6.6, 7.0, 7.1 (each m, AA′BB′ system, C6H4 Tyr, 4H),
7.1-7.4 (m, C6H5CH2, 5H), 8.3 (d, NH, 1H), 9.23 (s, OH, 1H,
major conformer), 9.25 (s, OH, 1H, minor conformer). Anal.
(C22H25N3O4) C, H, N.
tide amide (1 mmol) in 10 mL of CF3COOH was stirred for 15
min at a room temperature, and the solvent was evaporated
in vacuo. Ether was added to the residue, and the dipeptide
amide crystals were separated as a trifluoroacetic salt.
L-P r olyl-L-tyr osin e a m id e tr iflu or oa ceta te (19): yield
96%; mp 161-163 °C; [R]20 -13° (c 0.37, C2H5OH); Rf 0.28
D
(n-C4H9OH/AcOH/H2O, 5:1:2); NMR (DMSO-d6) δ 1.7-2.4 (m,
CâH2-CγH2 Pro, 4H), 2.68, 2.92 (each dd, J ) 13.9 Hz, J ) 9.96
Hz, J ) 4.14 Hz, J ) 13.71 Hz, CâH2 Tyr, 2H), 3.07-3.25 (m,
CδH2 Pro, 2H), 4.11 (m, CRH Pro, 1H), 4.41 (m, CRH Tyr, 1H),
6.65, 7.05 (each m, AA′BB′ system, C6H4 Tyr, 4H), 7.14, 7.61
(each s, NH2, 2H), 8.48 (s, NH Pro, 1H), 8.68 (d, J ) 8.1 Hz,
NH Tyr, 1H), 9.31 (s, OH Tyr, 1H). Anal. (C14H19N3O3‚CF3-
COOH) N.
N-(P h en yla cetyl)-L-p r olyl-D-tyr osin e a m id e (15): yield
64% (after column chromatography); mp 196-198 °C; [R]20
D
-65.8° (c 0.4, CH3OH); Rf 0.39 (CHCl3/CH3OH, 9:1); NMR
(DMSO-d6) two conformations evident in spectrum in ratio of
3:1 δ 1.4-2.1 (m, CâH2-CγH2 Pro, 4H), 2.61 (dd, J ) 13.89 Hz,
J ) 10.8 Hz, CâH2 Tyr, 1H, major conformer), 2.68 (dd, J )
14.1 Hz, J ) 10.8 Hz, CâH2 Tyr, 1H, minor conformer), 2.94
(dd, J ) 13.9 Hz, J ) 4.0 Hz, CâH2 Tyr, 1H, minor conformer),
3.03 (dd, J ) 14.1 Hz, J ) 3.65 Hz, CâH2 Tyr, 1H, major
conformer), 3.3-3.45 (m, CδH2 Pro, 2H), 3.65 (s, C6H5CH2, 2H),
4.17-4.31 (m, CRH Pro, CRH Tyr, 2H, major conformer), 4.4-
4.54 (m, CRH Pro, CRH Tyr, 2H, minor conformer), 6.63, 6.99
(each m, AA′BB′ system, C6H4 Tyr, 4H, major conformer), 6.63,
7.05 (each m, AA′BB′ system, C6H4 Tyr, 4H, minor conformer),
7.08-7.36 (m, C6H5CH2, 5H), 8.25 (d, NH, 1H), 9.23 (s, OH,
1H, major conformer), 9.25 (s, OH, 1H, minor conformer).
Anal. (C22H25N3O4) C, H, N.
D-P r olyl-L-tyr osin e a m id e tr iflu or oa ceta te (20): yield
93%; mp 93-95 °C (hydroscopic); [R]20 +32° (c 0.16, CHCl3);
D
Rf 0.26 (C4H9OH/CH3COOH/H2O, 5:1:2). NMR (DMSO-d6) δ
1.40-2.13 (m, CâH2-CγH2 Pro, 4H), 2.60, 2.99 (each dd, CâH2
Tyr, 2H), 3.12 (m, CδH2 Pro, 2H), 4.12 (m, CRH Pro, 1H), 4.47
(m, CRH Tyr, 1H), 6.63, 7.01 (each m, AA′BB′ system, C6H4
Tyr, 4H), 7.20, 7.64 (each s, NH2, 2H), 8.39 (s, NH Pro, 1H),
8.69 (d, NH Tyr, 1H), 9.17 (s, OH Tyr, 1H). Anal.
(C14H19N3O3‚CF3COOH) N.
N-Va ler yl-L-p r olyl-L-tyr osin e a m id e (16): yield 91% (af-
L-P r olyl-D-tyr osin e a m id e tr iflu or oa ceta te (21): yield
ter trituration with ether); mp 202-204 °C; [R]20 -60.7° (c
D
94%; mp 94-95 °C (hydroscopic); [R]20 -32° (c 0.2, CHCl3);
D
0.3, CH3OH); Rf 0.48 (CHCl3/C2H5OH, 4:1); NMR (DMSO-d6)
two conformations evident in spectrum δ 0.79 (t, CH3(CH2)3,
3H, minor conformer), 0.89 (t, CH3(CH2)3, 3H, major con-
former), 1.00-2.35 (m, CH3(CH2)3, CâH2-CγH2 Pro, 10H), 2.60-
2.98 (m, CâH2 Tyr, 2H), 3.13-3.59 (m, CδH2 Pro, 2H), 4.16 (m,
CRH Pro, 1H, major conformer), 4.24 (m, CRH Pro, 1H, minor
conformer), 4.24 (m, CRH Tyr, 1H, major conformer), 4.42 (m,
CRH Tyr, 1H, minor conformer), 6.63, 6.97 (each m, AA′BB′
system, C6H4 Tyr, 4H), 7.03, 7.49 (each s, NH2, 2H, minor
conformer), 7.09, 7.12 (each s, NH2, 2H, major conformer), 7.67
(d, NH Tyr, 1H, major conformer), 8.12 (d, NH Tyr, 1H, minor
conformer), 9.17 (s, OH Tyr, 1H, minor conformer), 9.18 (s,
OH Tyr, 1H, major conformer). Anal. (C19H27N3O4) C, H, N.
Rf 0.26 (C4H9OH/CH3COOH/H2O, 5:1:2); NMR (DMSO-d6) δ
1.41-2.13 (m, CâH2-CγH2 Pro, 4H), 2.60, 2.99 (each dd, CâH2
Tyr, 2H), 3.11 (m, CδH2 Pro, 2H), 4.12 (m, CRH Pro, 1H), 4.47
(m, CRH Tyr, 1H), 6.62, 7.00 (each m, AA′BB′ system, C6H4
Tyr, 4H), 7.18, 7.64 (each s, NH2, 2H), 8.39 (s, NH Pro, 1H),
8.69 (d, NH Tyr, 1H), 9.17 (s, OH Tyr, 1H). Anal.
(C14H19N3O3‚CF3COOH) N.
P h a r m a cology. In h ibition of Ap om or p h in e-In d u ced
Clim bin g Beh a vior in Mice. Male mice (C57BL6) weighing
22-25 g were placed in wire mesh cages (12 cm diameter, 14
cm high) and were allowed 1 h for adaptation and exploration
of the new environment. Apomorphine was dissolved ex
tempora in saline containing 0.1% ascorbic acid and injected
5.0 mg/kg sc. This dose caused climbing in all animals for 15
min. The test compound dissolved in saline containing 3%
Tween 80 was injected ip 15 min prior to the apomorphine.
For evaluation of climbing 15 min after apomorphine, 30
readings were taken every 2 min (scoring scale: four paws on
bottom (no climbing), 0; one paw on the wall, 1; two paws on
the wall, 2; three paws on the wall, 3; four paws on the wall
(full climbing), 4). Mice consistently climbing before apomor-
phine injection were discarded. The climbing scores were
individually totaled (maximum score: 120 per mouse) and the
total score of the control (vehicle ip - apomorphine sc) was
set to 100%. ED50 values with 95% confidence limits were
calculated by linear regression analysis (log dose transforma-
tion) using 8-16 mice per dose.
In h ibition of Extr a p ola tor y Beh a vior Im p a ir m en t
In d u ced by DOP A in Ra ts. The experimental animals were
outbread albino male rats (Moscow region, Stolbovaya) weigh-
ing 180-200 g. A rat put into a cylinder plunged in water
with the temperature of 22 °C should solve an “extrapolatory
task”, i.e., to dive under the cylinder’s edge, thus escaping the
stress situation. The animal was taken out of water im-
mediately after the solution of the task. The time during
which the animals were in water did not exceed 2 min. From
80 to 90% of the control rats solved the task. The rats which
did not escape in 2 min were discarded. One day later 0.9%
NaCl solution (intact group) or the suspension in 3% Tween
80 of Madopar (100 mg/kg L-DOPA + 25 mg/kg benzerazide)
(control group) were administrated intraperitonially 60 min
prior the rat being put into a cylinder. The test compounds
dissolved in saline + 3% Tween 80 were injected ip 10 min
prior to Madopar (test group). The animals with successful
escape response were registered. ED50 values with 95%
confidence limits were calculated by linear regression analysis
with eight rats per dose.
N-Ca p r oyl-L-p r olyl-L-tyr osin e a m id e (17): yield 59%
(after trituration from CHCl3); mp 112-113 °C; [R]20 -66° (c
D
0.3, CH3OH); Rf 0.4 (CHCl3/C2H5OH, 9: 1); NMR (DMSO-d6)
two conformations evident in spectrum δ 0.83 (t, CH3(CH2)4,
3H, minor conformer), 0.87 (t, CH3(CH2)4, 3H, major con-
former), 0.99-2.27 (m, CH3(CH2)4, 8H), 1.20-2.36 (m, CâH2-
CγH2 Pro, 4H), 2.66, 2.88 (each m, CâH2 Tyr, 2H, minor
conformer), 2.71, 2.99 (each m, CâH2 Tyr, 2H, major con-
former), 3.26-3.59 (m, CδH2 Pro, 2H), 4.16 (dd, CRH Pro, 1H,
major conformer), 4.25 (m, CRH Pro, 1H, minor conformer),
4.25 (m, CRH Tyr, 1H, major conformer), 4.43 (m, CRH Tyr,
1H, minor conformer), 6.63, 6.98 (each m, AA′BB′ system, C6H4
Tyr, 4H, major conformer), 6.61, 6.99 (each m, AA′BB′ system,
C6H4 Tyr, 4H, minor conformer), 7.09, 7.10 (each s, NH2, 2H,
major conformer), 7.10, 7.47 (each s, NH2, 2H, minor con-
former), 7.66 (d, NH Tyr, 1H, major conformer), 8.10 (d, NH
Tyr, 1H, minor conformer), 9.18 (s, OH Tyr, 1H, minor
conformer), 9.20 (s, OH Tyr, 1H, major conformer). Anal.
(C20H29N3O4) C, H, N.
N-Non a n oyl-L-p r olyl-L-tyr osin e a m id e (18): yield 62%
(after column chromatography); mp 178-180 °C; [R]20D -57.6°
(c 0.3, CH3OH); Rf 0.7 (CHCl3/CH3OH, 9:1); NMR (DMSO-d6)
two conformations evident in spectrum δ 0.86 (t, CH3(CH2)7,
3H), 1.10-2.27 (m, CH3(CH2)7, 14H), 1.50-2.10 (m, CâH2-CγH2
Pro, 4H), 2.6-3.0 (m, CâH2 Tyr, 2H), 3.20-3.50 (m, CδH2 Pro,
2H), 4.16 (m, CRH Pro, 1H, major conformer), 4.26 (m, CRH
Pro, 1H, minor conformer), 4.26 (m, CRH Tyr, 1H, major
conformer), 4.42 (m, CRH Tyr, 1H, minor conformer), 6.62, 6.97
(each m, AA′BB′ system, C6H4 Tyr, 4H, major conformer), 6.62,
7.01 (each m, AA′BB′ system, C6H4 Tyr, 4H, minor conformer),
6.95-7.15 (m, NH2, 2H), 7.67 (d, NH Tyr, 1H, major con-
former), 8.10 (d, NH Tyr, 1H, minor conformer), 9.17 (s, OH
Tyr, 1H, minor conformer), 9.18 (s, OH Tyr, 1H, major
conformer). Anal. (C23H35N3O4) C, H, N.
Gen er a l St r a t egy of t h e N-(ter t-Bu t oxyca r b on yl)-
d ip ep tid e Am id e Dep r otection . A solution of BOC-dipep-
Ca ta lep sy in Ra ts. Induction of catalepsy was tested in
adult outbread male rats (200-250 g) 0.5, 1.0, 1.5, and 2 h

290 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 3
Gudasheva et al.
(12) von Euler, G.; Fuxe, K. Neurotensin reduces the affinity of D-2
dopamine receptors in rat striatal membranes. Acta Physiol.
Scand. 1987, 131, 625-626.
(13) Pozdnev, V. F. The synthesis of tert-butyloxycarbonyl derivatives
of some trifunctional amino acids using di-tert-butylpyrocar-
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after ip administration of the test compound dissolved in saline
containing 3% Tween 80 in the apparatus Morpurgo.
The rats were placed with the forepaws onto a bar situated
7 cm above the floor of a perspex jar (22 × 38 × 16 cm) and
tested for catalepsy according to a rating score (time resting
upon the bar: 10 s, 0; 10-20 s, 1; 20-30 s, 2; >30 s, 3).
Testing was repeated twice for each animal at 2 min intervals.
The cumulated maximal score for one animal during the 2 h
observation period was 15. Scores are expressed as percent
of the maximal score obtainable.
Acu te Toxicity. Male outbred albino mice (Moscow region)
weighing 20-25 g were used. The mice were housed five per
cage and had free access to food and water. The mice were
kept at a temperature of 21-22 °C and humidity of 40-50%
and in a light-controlled room (12/12 h light/dark cycle). The
number of surviving animals was recorded 2, 24, and 48 h after
the injection.
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