Spiro[2.2]pentane as a dissymmetric scaffold for conformationally constrained analogues of glutamic acid: Focus on racemic 1-aminospiro[2.2]pentyl-1,4-dicarboxylic acids

Article abstract of DOI:10.1021/jo020138v

In search for novel conformationally constrained analogues of L-glutamic acid, a diastereodivergent synthesis of the four 1-aminospiro[2.2]pentyl-1,4-dicarboxylic acid racemic pairs is reported along with their stereochemical assignment, conformational analysis, and preliminary biological evaluation as potential glutamate (ionotropic and metabotropic) ligands.

Full text of DOI:10.1021/jo020138v

Sp ir o[2.2]p en ta n e a s a Dissym m etr ic Sca ffold for
Con for m a tion a lly Con str a in ed An a logu es of Glu ta m ic Acid : F ocu s
on Ra cem ic 1-Am in osp ir o[2.2]p en tyl-1,4-d ica r boxylic Acid s
Roberto Pellicciari,*,† Maura Marinozzi,† Emidio Camaioni,† Maria del Carmen Nu`nez,†,|
Gabriele Costantino,† Fabrizio Gasparini,‡ Gianluca Giorgi,§ Antonio Macchiarulo,† and
Natarajan Subramanian‡
Dipartimento di Chimica e Tecnologia del Farmaco, Universita` di Perugia, Via del Liceo, 1,
06123 Perugia Italy; Novartis Pharma, Basel, Switzerland; and Centro Interdipartimentale di Analisi e
Determinazioni Strutturali, Universita` di Siena, Via A. Moro, 53100 Siena, Italy
rp@unipg.it
Received February 27, 2002
In search for novel conformationally constrained analogues of L-glutamic acid, a diastereodivergent
synthesis of the four 1-aminospiro[2.2]pentyl-1,4-dicarboxylic acid racemic pairs is reported along
with their stereochemical assignment, conformational analysis, and preliminary biological evaluation
as potential glutamate (ionotropic and metabotropic) ligands.
CHART 1
In tr od u ction
Several strategies aimed at the reduction or elimina-
tion of the conformational flexibility of biologically active
substances have been developed.1 Among these, rigidifi-
cation through ring insertion,2 induction of hydrophobic
collapse,3 and intramolecular hydrogen bond formation4
are examples of “freezing” techniques that have produc-
tively been employed for the preparation of compounds
endowed with improved properties such as potency,
stability, biovailability, or selectivity. The quest for se-
lectivity, in particular, is of special relevance because
natural hormones, transmitters, and substrates usually
interact in a specific way with heterogeneous populations
of receptors by assuming a cluster of bioactive conforma-
tions. The case of L-glutamate (L-Glu, 1, Chart 1) is a
particularly relevant example. Indeed, L-Glu (1) is the
principal excitatory neurotransmitter in the vertebrate
CNS and is involved in a variety of physiological func-
tions, including neuronal plasticity, long-term potentia-
* Tel: +39 075 585 5120. Fax: +39 075 585 5124.
† Universita` di Perugia.
tion of synaptic activity, and memory, as well as in both
acute or chronic pathological processes such as brain
ischemia, hypoxia, and neurodegenerative diseases. The
variety of pathophysiologal functions exerted by L-Glu
(1) are mediated by two main families of membrane
receptors, the integral membrane spanning ionotropic
glutamate receptors (iGluRs)5 and the G-protein coupled
metabotropic glutamate receptors (mGluRs).6 Each of
these two families, in turn, is further subdivided in a
multiple and heterogeneous variety of receptor subtypes
and isoforms (known as GluR1-GluR6, KA1-KA2, NR1,
NR3 in the case of iGluR’s, and as mGluR1-mGluR8
‡ Novartis Pharma.
§ Universita` di Siena.
| Present address: Departamento de Quimica Organica, Facultad
de Farmacia, Campus de Cartuja s/n, 18071 Granada, Spain.
(1) (a) Wang, P.; Brank, A. S.; Banavali N. K.; Nicklaus, M. C.;
Marquez, V. M.; Christman, J . K.; MacKerell, A. D. J . Am. Chem. Soc.
2000, 122, 12422-12434. (b) Bernardi, A.; Arosio, D.; Manzoni, L.;
Micheli, F.; Pasquarello, A.; Seneci, P. J . Org. Chem. 2001, 66, 6209-
6216. (c) Anderson, A.; Boyd, A. C.; Clark, J . K.; Fielding, L.; Gemmell,
D. K.; Hamilton, M. N.; Maidment, M. S.; May, V.; McGuire, R.;
McPhail, P.; Sansbury, F. H.; Sundaram, H.; Taylor, R. J . Med. Chem.
2000, 43, 4118-4125. (d) Hoepping, A.; J ohnson, K. M.; Clifford, G.;
Flippen-Anderson, J .; Kozikowski, A. P. J . Med. Chem. 2000, 43, 2064-
2071.
(2) Halab, L.; Gosselin, F.; Lubell, W. D. Biopolymers 2000, 55, 101-
122.
(3) Boge, T. C.; Wu, Z. J .; Himes, R. H.; Vander Velde, D. G.; Georg
G. I. Bioorg. Med. Chem. Lett. 1999, 9, 3047-3052.
(4) Meinke, P. T.; Arison, B.; Culberson, J . C.; Fisher, M. H.; Mrozik,
H. J . Org. Chem. 1998, 63, 2591-2596.
(5) The Ionotropic Glutamate Receptors; Monaghan, D. T., Wenthold,
R. J ., Eds.; Humana Press: Totowa, New J ersey, 1997.
(6) Pellicciari, R.; Costantino, G. Curr. Opin. Chem. Biol. 1999, 3,
433-40.
10.1021/jo020138v CCC: $22.00 © 2002 American Chemical Society
Published on Web 07/12/2002
J . Org. Chem. 2002, 67, 5497-5507
5497

Pellicciari et al.
CHART 2
in the case of mGluR’s) with several evidences indicating
a peculiar physio-pathological role for each individual
subtype. The pharmacological characterization of this
vast and complex receptor subtype array has largely been
made possible by the design and synthesis of conforma-
tionally constrained L-Glu (1) analogues.7 We have been
engaged for a long time in this endeavor,8 mainly ad-
dressing our work to the preparation of non-proteino-
genic, acidic amino acids endowed with potency and
selectivity for L-Glu receptor subtypes of the two families.
In this connection, ring insertion has been the strategy
of choice to constrain the glutamate skeleton in search
for its “frozen” subtype selective bioactive conformations;
thus, cyclopentane (2),9 cyclopropane (4, 5),10,7d bicylo-
[3.2.0]heptane (3),7b and bicylo[3.1.0]hexane (6)7c are
relevant examples of ring moieties successfully used to
conformationally rigidify L-Glu (1). More exotic scaffolds
such as bicyclo[1.1.1]pentane (7)8c and the cubyl (8)11
moiety have also been shown to be good surrogates of
the phenyl ring as spacers able to constrain the phar-
macophoric groups of L-Glu (1) into a coplanar disposi-
tion.
More recently, our attention was attracted by the
possibility to employ the unusual spiro[2.2]pentane nucleus
as a new scaffold for fully constrained L-Glu analogues.
Indeed, the 1,4-functionalization of this nucleus gives rise
to the corresponding 1-aminospiro[2.2]pentyl-1,4-dicar-
boxylic acids (22a -d ), conformationally blocked L-Glu
analogues.
Few precedents of biologically active substances incor-
porating the spiropentane nucleus are known, and they
include spiropentylacetic acid and spiropentyl-acetyl-CoA
(9, Chart 2) as inhibitors of acycl-CoA dehydrogenases,12
and spiropentane analogues of deoxyadenosine and de-
oxyguanosine (10, 11, Chart 2) as inhibitors of human
cytomegalovirus and Epstein-Barr virus.13
tions, whose possible biological significance is worthy of
testing; (ii) the 1-4 functionalization of the spiro[2.2]-
pentane nucleus to give the corresponding 1-aminospiro-
[2.2]pentyl-1,4-dicarboxylic acid isomers generates three
new chiral centers, so that four possible diastereoisomeric
pairs are generated; (iii) an appropriate diastereodiver-
gent synthesis would give rise to the availability of a
stereolibrary of compounds in which differently oriented
functional groups will mimic folded or extended confor-
mations of L-Glu (1); (iv) the resulting libraries can be
used to screen a variety of glutamate receptor subtypes
toward a better definition of the conformation-activity
relationships.
We report here the synthesis and the stereochemical
characterization of the four pairs of 1-aminospiro[2.2]-
pentyl-1,4-dicarboxylic acid (22a -d , ASPEDs) isomers.
Resu lts
A number of entries into substituted [2.2]spiropentanes
have been developed, mainly based on strategies involv-
ing the cyclopropanation of allenes or alkylidene cyclo-
propanes via a Simmons-Smith reaction14,15 or diazoal-
kanes12b,17 and diazoesters15-17 addition. Dihalo- and
bromospiro[2.2]pentane derivatives have been also pre-
pared by addition of dihalocarbenes18 or bromocarbenoid
intermediates,21 respectively, to 2-substituted methyl-
enecyclopropanes. Trost et al.20 have described the prepa-
ration of 1,2,4-trisubstituted spiropentanes by conjugate
addition of phenylsulfonium cyclopropylidene to R,â-
unsaturated carbonyl compounds. Recently, the synthesis
of amino acids containing the spiropentyl unit such as
(()-spiropentylglycine (12) and (()-1-amino spiropentane
carboxylic acid (13) has been reported.21 Thus, racemic
13 was prepared by the rhodium-catalyzed decomposition
of dimethyl diazomalonate in the presence of methyl-
enecyclopropane followed by Curtius rearrangement key
The following aspects make the spiro[2.2]pentane
analogues of L-Glu (1) particularly attractive: (i) the
unusual spiro junction between two cyclopropane rings
forces the pseudotorsional angles into unnatural disposi-
(7) (a) For a general review, see: Bra¨uner-Osborne, H.; Egebjerg,
J .; Nielsen, E.; Madsen, U.; Krogsgaard-Larsen, P. J . Med. Chem. 2000,
43, 2609-2645. (b) Kozikowski, A. P.; Steensma, D.; Araldi, G. L.;
Tuckmantel, W.; Wang, S.; Pshenichkin, S.; Surina, E.; Wroblewski,
J . T. J . Med. Chem. 1998, 41, 1641-1650. (c) Monn, J . A.; Valli M. J .;
Massey, S. M.; Wright, R. A.; Salhoff, C. R.; J ohnson, B. G.; Howe, T.;
Alt, C. A.; Rhodes, G. A.; Robey, R. L.; Griffey, K. R.; Tizzano, J . P.;
Kallman, M. J .; Helton, D. R.; Schoepp, D. D. J . Med. Chem. 1997, 40,
528-537. (d) Shimamoto, K.; Ohfune, Y. J . Med. Chem. 1996, 39, 407-
423.
(8) (a) Pellicciari, R.; Luneia, R.; Costantino, G.; Marinozzi, M.;
Natalini, B.; J akobsen, P.; Kanstrup, A.; Lombardi, G.; Moroni, F.;
Thomsen, C. J . Med. Chem. 1995, 38, 3717-3719. (b) Pellicciari, R.;
Marinozzi, M.; Natalini, B.; Costantino, G.; Luneia, R.; Giorgi, G.;
Moroni, F.; Thomsen, C. J . Med. Chem. 1996, 39, 2259-2269. (c)
Pellicciari, R.; Raimondo, M.; Marinozzi, M.; Natalini, B.; Costantino,
G.; Thomsen, C. J . Med. Chem. 1996, 39, 2874-2876. (d) Pellicciari,
R.; Marinozzi, M.; Costantino, G.; Natalini, B.; Moroni, F.; Pellegrini-
Giampietro, D. J . Med. Chem. 1999, 42, 2716-2720.
(14) Ullman, E.; Fanshave, W. J . J . Am. Chem. Soc. 1961, 83, 2379-
2383.
(15) Charette, A. B.; J olicoeur, E.; Bydlinski, G. A. S. Org. Lett. 2001,
3, 3293-3295.
(16) Gajewski, J . J .; Burka, L. T. J . Org. Chem. 1970, 35, 2190-
2196.
(9) Manzoni, O.; Fagni, L.; Pin, J . P.; Rassendren, F.; Poulat, F.;
Sladeczek, F.; Bockaert, J . Mol. Pharmacol. 1990, 38, 1-6.
(10) Pellicciari, R.; Curini, M.; Natalini, B.; Ceccherelli, P. IX
International Symposium on Medicinal Chemistry, Berlin, 1986; p 118.
(11) Pellicciari, R.; Costantino, G.; Giovagnoni, E.; Mattoli, L.;
Brabet, I.; Pin, J .-P. Biorg. Med. Chem. Lett. 1998, 8, 1569-1573.
(12) (a) Tserng, K. Y.; J in, S.-J .; Hoppel, C. L. Biochemistry 1991,
30, 10755. (b) Li, D.; Zhou, H.-Q.; Dakoji, S.; Shin, I.; Oh, E.; Liu, H.-
W. J . Am. Chem. Soc. 1998, 120, 2008-2017.
(17) Konzelman, L. M.; Conley, R. T. J . Org. Chem. 1968, 33, 3828-
3841.
(18) Akhachinskauya, I. V.; Donskaya, N. A.; Kalykina, I. V.;
Oprunenko, Y. F.; Shabarov, Y. S. Zh. Org. Khim. 1989, 25, 1645.
(19) Cheng, C.; Shimo, T.; Somekawa, K.; Kawaminami, M. Tetra-
hedron Lett. 1997, 52, 9005-9008.
(20) Trost, B. M.; Bogdanowicz, M. J . J . Am. Chem. Soc. 1973, 95,
5307-5310.
(21) de Meijere, A.; Ernst, K.; Zuck, B.; Brandl, M.; Kozhushkov, S.
I.; Tamm, M.; Yufit, D. S.; Howard, J . A. K.; Labahn, T. Eur. J . Org.
Chem. 1999, 3105-3115.
(13) Guan, H.-P.; Ksebati, M. B.; Cheng, Y.-C.; Drach, J . C.; Kern,
E. R.; Zemlicka, J . J . Org Chem. 2000, 65, 1280-1290.
5498 J . Org. Chem., Vol. 67, No. 16, 2002

1-Aminospiro[2.2]pentyl-1,4-dicarboxylic Acids
SCHEME 1a
a
(1) tBuO2CCHN2, (Rh(OAc)2)2, CH2Cl2; (2) tBuOK, Et2O; (3) (EtO2C)2CN2, (Rh(OAc)2)2, CH2Cl2; (4) NaOH, MeOH; (5) (a) EtOCOCl,
NaN3, THF, (b) tBuOH; (6) (a) 8 N J ones reagent, (b) 6 N HCl, (c) Dowex, 10% pyr.
sequence.21 Furthermore, the enantioselective synthesis
of 1,1,4-trisubstituted spiropentanes by a dioxaborolane
ligand controlled zinc-mediated cyclopropanation of hy-
droxymethylallenes has also been reported.15
Our initial approach to the synthesis of 1-amino-spiro-
[2,2]pentyl-1,4-dicarboxylic acids (22a -d ) began with the
preparation of the key intermediate tert-butyl (methyl-
enecyclopropyl)carboxylate22 (()-14 from commercially
available 2-bromopropene (15) and tert-butyl diazoac-
etate23 by a dirhodium(II) tetraacetate catalyzed cyclo-
propanation as depicted in Scheme 1.
The bromide (()-16 thus obtained in 56% yield was
submitted to a potassium tert-butoxide induced dehydro-
bromination in DMSO at room temperature to give (()-
14 in 46% yield. A subsequent dirhodium(II) tetraacetate
catalyzed cyclopropanation of (()-14 with a solution of
dimethyl diazomalonate in CH2Cl2 afforded, surprisingly,
only the distal isomer of the 1,1-dimethyl, 5-tert-butyl
spiro[2.2]pentanetricarboxylate ((()-17). No trace of the
other isomer was detected by GC-MS analysis of the
crude reaction mixture. Flash chromatography of the
crude product led to the isolation of pure (()-17 with 33%
yield. The stereochemistry of (()-17 was initially ratio-
nalized with the steric effect exerted by the bulky tert-
butyl moiety of the olefin analogously with our previous
reported result.8d Subsequently, this prediction was
confirmed by comparison of the analytical data (1H NMR,
13C NMR, and HPLC) of the mixture of the final amino
acids (22c + 22d ), obtained by this route with those of
the single isomeric amino acids (22a -d ) resulted from
the alternative route below described.
refluxed overnight in tert-butyl alcohol to give the
mixture of the distal and medial-syn isomeric N-Boc
aminoesters (()-20 and (()-21 (42%). The unseparated
mixture was hydrolyzed with 6 N HCl at 80 °C for 8 h to
give, after purification by ion-exchange resin chromatog-
raphy, the mixture of the 1-amino-spiro[2,2]pentyl-1,4-
dicarboxylic acids ((()-22c and (()-22d , 46%).
To have access to all four possible racemic diastereo-
mers of 1-amino-spiro[2,2]pentyl-1,4-dicarboxylic acids
((()-22a -d ) we decided to modify the above-described
synthetic route in such a way as to decrease the steric
bulk of the tert-butyl moiety on the olefin (Schemes 2 and
3).24 Thus, ethyl methylenecyclopropanecarboxylate ob-
tained from 2-bromopropene according to a known pro-
cedure25 was submitted to LiAlH4 reduction to afford the
corresponding carbinol (()-23,13 which was protected as
tert-butyldimethylsilyl ether using tert-butyldimethylsilyl
chloride and 4-(dimethylamino)pyridine in CH2Cl2 to give
the olefin (()-24 (55%). Subsequent slow addition of a
solution of diethyl diazomalonate in CH2Cl2 to a refluxing
solution of (()-24 and dirhodium(II) tetraacetate in the
same solvent yielded a mixture of the two spiropentyl
diastereoisomers (()-25 and (()-26 in a 1:5 ratio (GC-
MS). Medium-pressure liquid chromatography (MPLC)
of this mixture on silica gel (cyclohexanes-ethyl acetate,
95/5) afforded the less polar proximal isomer (()-25 and
the more polar spiropentyl distal isomer (()-26 in 11%
and 53% yield, respectively. Selective basic monohydroly-
sis of the malonate moiety of 4-(tert-butyldimethylsilyl-
oxymethyl)-spiro[2.2]pentane-1,1-dicarboxylyc acid di-
ethyl ester ((()-25) with 0.5 N NaOH/EtOH at room
temperature gave after careful neutralization (10% citric
acid) a mixture of two proximal and medial-anti diaste-
reoisomeric monoacids (()-27a and (()-27b (90%), since
a new chiral center was introduced. The unresolved
mixture of (()-27a and (()-27b was then transformed
into the corresponding acyl azide by activation with ethyl
chloroformate in the presence of 18-crown-6 and potas-
Selective monohydrolysis of the malonate moiety of (()-
17 with 1 equiv of aqueous NaOH furnished the mixture
of the monoesters (()-18 and (()-19 (72%), which was
converted into the corresponding acyl azides via the
mixed anhydrides (ethyl chloroformate, 18-crown-6, po-
tassium carbonate) followed by treatment with aqueous
sodium azide. The crude acyl azides mixture was then
(22) Brandl, M.; Kozhushkov, S. I.; Yufit, D. S.; Howard, J . A. K.;
de Meijere, A. Eur. J . Org. Chem. 1998, 2785-2795.
(23) Regitz, M.; Hocker, J .; Liedhegener, A. Organic Syntheses;
Wiley: New York, 1973; Collect. Vol. V, p 178.
(24) While this work was in progress, Zemlicka et al.13 reported the
synthesis of all isomers of 1,4-disubstituted spiropentanes by a similar
approach.
(25) Lai, M.-T.; Liu, H.-W. J . Am. Chem. Soc. 1990, 112, 4034-4035.
J . Org. Chem, Vol. 67, No. 16, 2002 5499

Pellicciari et al.
SCHEME 2a
a
(1) TBDSCl, DMAP, CH2Cl2; (2) (a) (EtO2C)2CN2, (Rh(OAc)2)2, CH2Cl2, (b) MPLC.
SCHEME 3a
a(1) NaOH, EtOH; (2) (a) EtOCOCl, NaN3, THF, (b) tBuOH; (3) (a) TBAF, THF, (b) mpic; (4) (a) BN J ones reagent, (b) 6 N HCl, (c)
Dowex, 10% pyr.
sium carbonate and subsequent reaction with an aqueous
solution of sodium azide at 0 °C.26 Refluxing the crude
acyl azide in the presence of tert-butyl alcohol afforded
after flash chromatography (light petroleum-ethyl ac-
etate, 90/10) the corresponding proximal and medial-anti
N-Boc-protected aminospiropentanes (()-28a and (()-
28b (30%), which could not be separated by silica gel
chromatography. Nevertheless, after desilylation by Bu4-
NF in THF to the diastereisomeric proximal and medial-
anti isomeric alcohols (()-29a and (()-29b, these were
isolated individually by MPLC (light petroleum-ethyl
acetate, 3/2) in 21% and 65% yields, respectively. By
following an analogous procedure, selective basic mono-
hydrolysis of the malonate moiety of isomer (()-26
afforded a mixture of the distal and medial-syn monoac-
ids (()-27c and (()-27d (78%), which was converted into
the mixture of the corresponding N-Boc amino distal and
medial-syn derivatives (()-28c and (()-28d (32%). Re-
moval of the silyl protecting group afforded distal and
medial-syn isomers (()-29c and (()-29d , which were
resolved by MPLC in 54% and 35% yields, respectively.
Oxidation of the primary hydroxy group of the separated
isomers (()-29a -d with 8 N J ones’s reagent in acetone
at 0 °C and subsequent final acidic hydrolysis (6 N HCl,
90 °C, 16 h) furnished, after ion-exchange resin chroma-
tography on Dowex 50X2-200 using 10% aqueous pyri-
dine as eluent, the corresponding desired proximal,
medial-anti, distal, and medial-syn spiropentyl amino
acids (()-22a (ASPED-A), (()-22b (ASPED-B), (()-22c
(ASPED-C), and (()-22d (ASPED-D) in 65%, 61%, 71%,
and 68% yield, respectively.
Nom en cla tu r e a n d Ster eoch em ica l Assign m en t.
Functionalization at 1-4 positions of the spiro[2.2]-
(26) Dappen, M. S.; Pellicciari, R.; Natalini, B.; Monahan, J . B.;
Chiorri, C.; Cordi, A. A. J . Med. Chem. 1991, 34, 161-168.
5500 J . Org. Chem., Vol. 67, No. 16, 2002

1-Aminospiro[2.2]pentyl-1,4-dicarboxylic Acids
SCHEME 4
SCHEME 5
pentane moiety generates four pairs of enantiomers. To
distinguish between these enantiomeric pairs it is con-
venient to adopt a trivial stereochemical nomenclature
specifically suited for catching the spatial relationships
between pharmacophoric groups. In 1970, Gayewski and
Burka16 assigned disubstituted spiropentanes with a
stereochemical nomenclature based on the evaluation of
(i) the linear distance between substituents and (ii) the
relative orientation below and above a reference plane.
Thus, as depicted in Scheme 4, the four possible diaste-
reoisomeric disubstituted spiropentanes are classified as
proximal (shortest distance between substituents), distal
(longest distance between substituents), medial-syn (in-
termediate distance and syn disposition referring to the
plane bearing the higher priority substituent), and
medial-anti (intermediate distance and anti disposition
referring to the plane bearing the higher priority sub-
stituent).
Although originally defined for disubstituted spiropen-
tane derivatives, Gayewski and Burka’s rules can be
applied without ambiguity to our trisubstituted spiro-
pentante derivatives 22a -d if relative distances are
considered between substituents with the higher IUPAC
priority. Therefore, having defined this additional crite-
rion, our racemic pair endowed with the largest distance
between the amino group and the distal carboxylate will
be named distal, the one with the shortest distance will
be named proximal. The other two isomers will be
assigned with the medial-syn and medial-anti nomen-
clature according to the orientation above or below the
reference plane containing the highest priority substitu-
ent.
although much weaker, also in derivative 22c and 22d .
As a result of the rigidity of the spiro[2.2]pentane moiety,
this NOE effect must be ascribed to protons lying on the
same face of the spiro[2.2]pentane system. Accordingly,
22a and 22b must be the proximal or the medial-anti
isomers, whereas 22c and 22d must be the distal or
medial-syn ones. No other diagnostic signals were identi-
fied, thus making impossible a further discrimination
between the two diastereoisomeric pairs.
The definitive stereochemical assignment of the new
derivatives was then based upon the single-crystal X-ray
analysis carried out on selected spiropentane amino
acids. Thus, the hydrobromic salts of the racemic spiro-
pentane amino acids 22b, 22c, and 22d were submitted
to X-ray crystallography. Crystal data and relevant
geometrical parameters are reported in Tables 1 and 2.
The solved crystal of the hydrobromic salt of derivative
22d is endowed with a S-chirality at the amino acidic
carbon atom and at the spiro carbon atom, while a
R-chirality is observed at the carbon atom bearing the
distal carboxylate at position 4 (see Figure S9, Supporting
Information). Derivative 22d belongs to a centric space
group, thus confirming its racemic nature. Accordingly,
the crystal structure of the hydrobromic salt of 22d
describes both (1S,3S,4R)-1-aminospiro[2.2]pentyl-1,4-
dicarboxylic acid and its (1R,3R,4S) enantiomer. In an
analogous way, configuration (1S,3R,4R) and (1R,3S,4S)
could be attributed to the racemic mixture 22c, also
belonging to a centric space group. Derivative 22b was
assigned with (1S,3R,4S) configuration, but in this case
the compound crystallizes in a chiral space group (Pna21).
Since the racemic mixture 22a is chemically distinguish-
able from either 22b, 22c, or 22d and possesses NOE
properties similar to those of 22d , it represents the fourth
pair of diastereoisomeric 1-aminospiro[2.2]pentyl-1,4-
dicarboxylic acids and, hence, must be attributed the
remaining (1S,3S,4S) and (1R,3R,4R) configurations.
The next step toward the stereochemical characteriza-
tion of the four pairs 22a -d was the attribution of the
trivial nomenclature above-described. Thus, the spatial
relationships between functional groups in the four pairs
are summarized in Scheme 6 and, according to the chosen
Once having defined these criteria, we faced the
problem of the stereochemical assignment of the four
racemic pairs 22a -d . As a first attempt, we tried to
identify diagnostic protons in bidimensional NMR experi-
ments (Scheme 5).
The chemical shift of all protons was assigned by using
(H,H)-COSY experiments. In particular, the downfield
proton was assigned as H4 (proton in R-position with
respect to the distal carboxylate). Proton H4 shows COSY
effects with two upfield protons that were therefore
assigned as H5 (protons in the â-position with respect to
the distal carboxylate). The remaining two protons are
those in the 2-position of the spiropentane system. From
the examination of NOESY data, NOE correlations were
observed between proximal protons; thus a strong NOE
effect between H4 and both H5 protons was observed in
all isomers 22a -d . This effect was therefore undiagnostic
for the stereochemical assignment. Weak NOE correla-
tions, furthermore, were observed in derivative 22a and
22b between H4 and H2 protons. This effect is present,
J . Org. Chem, Vol. 67, No. 16, 2002 5501

Pellicciari et al.
TABLE 1. Cr ysta l Da ta for Com p ou n d s 22b -d
22d
22c
22b
formula
M
C7H9NO4‚HBr
252.07
C7H9NO4‚HBr
252.07
C7H9NO4‚HBr‚H2O
270.09
crystal size (mm)
temperature (K)
wavelength (Å)
crystal system
space group
a (Å)
0.3 × 0.2 × 0.2
0.3 × 0.3 × 0.2
0.3 × 0.2 × 0.2
293(2)
293(2)
293(2)
0.71073
monoclinic
P21/c (No. 14)
6.922(1)
0.71073
monoclinic
P21/c (No. 14)
8.479(1)
0.71073
orthorhombic
Pna21 (No. 33)
9.051(1)
b (Å)
5.835(1)
10.238(1)
12.242(1)
c (Å)
24.653(3)
11.295(1)
9.123(1)
â (deg)
93.13(1)
994.2(3)
99.11(1)
968.1(2)
U (Å3)
1010.8(2)
Z
4
4
4
F(000)
504
1.684
4.118
omega
504
1.729
4.229
omega
544
1.775
4.064
omega
Dc (g cm-3
)
(Mo K) (mm-1
scan mode
)
scan range/°
scan width/°
2.9e Θe 25.0
1.36
2.4e Θe 25.0
1.04
2.8e Θe 30.0
0.92
scan speed/° min-1
reflections collected
independent reflections
refinement method
no. of parameters refined
R1 (all data)
3
3
3
2641
2320
3094
1738 (Rint ) 0.02)
1705 (Rint ) 0.05)
2924 (Rint ) 0.02)
full-matrix least-squares on F2
full-matrix least-squares on F2
full-matrix least-squares on F2
158
151
176
0.0440
0.0679
1.076
0.079
0.095
1.015
0.0728
0.0953
1.329
wR2 (all data)
goodness-of-fit on F2
TABLE 2. Releva n t Geom etr ica l P a r a m eter s (Å, d eg)
for Non -Hyd r ogen Atom s of Com p ou n d s 22b-d
SCHEME 6
22d
22c
22b
N(1)-C(1)
1.471(4)
1.322(4)
1.202(4)
1.271(4)
1.253(4)
1.538(5)
1.483(4)
1.482(4)
1.468(5)
1.456(5)
1.490(4)
1.532(5)
1.480(5)
118.4(3)
58.1(2)
1.468(6)
1.299(6)
1.215(6)
1.326(6)
1.209(7)
1.540(7)
1.477(7)
1.487(6)
1.464(7)
1.469(7)
1.483(7)
1.535(8)
1.481(8)
118.7(4)
58.0(3)
1.465(7)
1.329(7)
1.202(7)
1.324(8)
1.207(7)
1.550(8)
1.486(7)
1.482(7)
1.481(8)
1.461(9)
1.478(8)
1.524(12)
1.492(11)
118.1(5)
58.4(4)
O(1)-C(6)
O(2)-C(6)
O(3)-C(7)
O(4)-C(7)
C(1)-C(2)
C(1)-C(3)
C(1)-C(6)
C(2)-C(3)
C(3)-C(4)
C(3)-C(5)
C(4)-C(5)
C(5)-C(7)
N(1)-C(1)-C(2)
C(2)-C(1)-C(3)
C(3)-C(1)-C(6)
C(1)-C(2)-C(3)
C(1)-C(3)-C(2)
C(1)-C(3)-C(4)
C(2)-C(3)-C(5)
C(4)-C(3)-C(5)
C(3)-C(4)-C(5)
C(3)-C(5)-C(4)
C(3)-C(5)-C(7)
O(1)-C(6)-O(2)
O(3)-C(7)-O(4)
C(5)-C(7)-O(4)
C(5)-C(7)-O(3)
122.2(3)
59.1(2)
62.8(2)
136.2(3)
135.8(3)
62.6(2)
122.4(4)
58.8(3)
63.1(3)
135.0(4)
138.6(4)
62.6(4)
120.7(4)
58.7(3)
63.0(4)
132.6(5)
138.9(5)
62.4(5)
arrangement of the two spirofused rings, with the sub-
stituents bisecting the corresponding angles of the ring.
As expected, this perpendicular arrangement of the two
cyclopropane rings forces the pseudo-torsional angles
C1-C2-C3-C4 and C2-C3-C4-C5 to values that are
attainable to L-Glu (1) only with a conspicuous energy
expense. As an example, Table 3 reports the energy
values, computed with the Universal Force Field by
considering neutral zwitterionic species and by simulat-
ing a water environment with a dielectric constant of 80,
necessary for L-Glu (1) to achieve the conformation
mimicked by 22a -d .
59.8(2)
57.6(2)
59.1(3)
58.2(3)
59.3(4)
58.2(4)
118.2(3)
125.6(3)
123.7(3)
119.6(3)
116.7(3)
119.4(5)
125.9(5)
123.6(5)
123.5(5)
112.8(5)
118.1(5)
124.8(5)
123.5(6)
123.0(6)
113.5(5)
In alternative to dihedral angles, another parameter
used as an indicator of the geometry of L-Glu analogues
is the distance between functional groups.27 According
to this indicator, L-Glu (1) and L-Glu analogues are
classified as extended or folded on the basis of the
distances d1 and d2 between the distal carboxylate and
the R-amino group or the R-carboxylate, respectively.
criteria, 22a is the proximal, 22b the medial-anti, 22d
the media- syn, and 22c the distal isomeric pair.
1-Am in osp ir o[2.2]p en tyl-1,4-d ica r boxylic Acid s a s
Tools for Con for m a tion a l An a lysis. Although topo-
logically equivalent to L-Glu (1), the 1-aminospiro[2.2]-
pentyl-1,4-dicarboxylic acids (22a -d ) have a three-
dimensional structure that corresponds to high-energy
conformers of the naturally occurring parent derivative.
Indeed, X-ray crystallography confirms a perpendicular
(27) Tellier, F.; Acher, F.; Brabet, I.; Pin, J . P.; Azerad, R. Bioorg.
Med. Chem. 1998, 6, 195-208.
5502 J . Org. Chem., Vol. 67, No. 16, 2002

1-Aminospiro[2.2]pentyl-1,4-dicarboxylic Acids
F IGURE 1. Projection of the four pairs of diastereoisomers of trisubstituted spiropentanes 22a -d and of the nine staggered
conformations of glutamate, into a bidimensional plot according to the d1,d2 distances; d1 is defined as the distance between the
carbons of the two carboxylate groups (C-C), and d2 is defined as the distance between the carbon of the distal carboxylate group
and the nitrogen of the amino acidic amino group.
TABLE 3. En er getic P en a lties Requ ir ed by Glu ta m a te
to Ad op t Con for m a tion s Mim ick ed by 22a -d or Its
Bioa ctive Con for m a tion s a t iGlu R’s or m Glu R’sa
of the nine staggered conformations of glutamate when
projected into the same plot (Figure 1). Thus, the
proximal 1-aminospiro[2.2]pentyl-1,4-dicarboxylic acid
(22a ) is endowed with distances close to the g-g+
conformation of L-Glu (1). The distances d1-d2 in the
medial-syn isomers 22d are closely related to the g-g-
conformation of L-Glu (1), whereas the medial-anti
isomers 22b resemble the anti-anti glutamic acid, and
the distal isomers the g+anti. The bidimensional plot is
also useful to compare the conformational profile of
22a -d with respect to the parent carboxycyclopropyl-
glycines (Figure 2). Thus, the introduction of the spiro-
fused ring reduces the conformational degeneracy of
CCGs from 12 possible conformations to only 4. The
proximal isomers 22a correspond to the eclipsed-anti
conformation of L-CCG-IV (5), the distal pair corresponds
to the pseudoanti-g+ conformation of L-CCG-I (4), the
medial-syn 22d corresponds to the eclipsed-g+ conforma-
tion of L-CCG-IV (5), and, finally the medial-anti pair
22b corresponds to the pseudoanti-anti conformation of
L-CCG-I (4). Of interest also is the possibility of project-
ing into the bidimensional plot the proposed bioactive
conformation of L-Glu (1) experimentally determined
(Figure 2). The proposed bioactive conformation of L-Glu
(1) when acting at mGluR1 was extracted from the X-ray
crystal structure (2.20 Å resolution) of the ligand binding
domain of mGluR128 and shown to match the pseudoanti-
anti conformation of L-CCG-I and the medial-anti isomer
of 1-aminospiro[2.2]pentyl-1,4-dicarboxylic acid (22b).
The putative bioactive conformation of L-Glu (1) acting
at the AMPA receptor was extracted from the X-ray
structure (1.9 Å resolution) S1-S2 construct of the GluR2
receptor29 and shown to match the pseudoanti-g- confor-
mation of L-CCG-II (4) but none of the 1-aminospiro[2.2]-
energetic gap
compound
(kcal/mol)
ASPED A [(()-22a ]
ASPED B [(()-22b]
ASPED C [(()-22d ]
ASPED D [(()-22c]
AMPA-Glu
21.39
13.01
12.38
10.20
4.13
NMDA-Glu
0.80
mGluR1-Glu
1.50
a
Energetic penalties are calculated, by constraining the tor-
sional bonds of glutamate to the values of the putative bioactive
conformations or individual compounds 22a -d and by calculating
the difference in energy from the global minimum conformer. All
of the species are considered in their zwitterionic forms
If the four pairs of ASPED diastereoisomers are plotted
in a bidimensonal plot reporting the d1 versus d2 dis-
tances computed by using the Universal Force Field (see
Experimental Section), a clear picture of the spatial
relationship among functional groups is obtained. In
particular, steroisomers with (d1 + d2) e 7.5 Å can be
classified as folded isomers; those endowed with 7.5 g
(d1 + d2) e 8.0 Å can be classified as semifolded; those
endowed with 8.0 g (d1 + d2) e 8.5 Å can be named
semiextended, and those with the (d1 + d2) g 8.5 Å are
termed extended isomers. Clearly, because of the discrete
nature of the torsional angles for rigid isomers, only
singular points can be represented in the plot. It can be
observed that, quite unexpectedly, the medial-syn 1-amino-
spiro[2.2]pentyl-1,4-dicarboxylic acid (22d ) falls into the
region of folded conformation, whereas the proximal
1-aminospiro[2.2]pentyl-1,4-dicarboxylic acid (22a ) can
rather be classified as a semifolded glutamate analogue.
The other two pairs, the medial-anti 22b and the distal
22c, have a more extended disposition of functional
groups and are projected between the semiextended and
extended region. It is interesting to compare the position
(28) Kunishima, N.; Shimada, Y.; Tsuji, Y.; Sato, T.; Yamamoto, M.;
Kumasaka, T.; Nakanishi, S.; J ingami, H.; Morikawa, K. Nature 2000,
407, 971-977.
(29) Armstrong, N.; Gouaux, E. Neuron 2000, 28, 165-181.
J . Org. Chem, Vol. 67, No. 16, 2002 5503

Pellicciari et al.
F IGURE 2. Comparison between 22a -d and carboxycyclopropylglycine in the d1-d2 space. Again, d1 is defined as the distance
between the carbons of carboxylate groups (C-C), and d2 is defined as the distance between the carbon of the distal carboxylate
group and the nitrogen of the amino acidic amino group. Stars indicate the projection of the proposed bioactive conformations of
glutamate when acting at selected iGluR’s or mGluR’s.
pentyl-1,4-dicarboxylic acid isomers (22a -d ). Finally, the
putative bioactive conformation of L-Glu (1) was inferred
from a homology model of the NMDA-R2 receptor (un-
published result) and shown to correspond to the eclipsed-
g+ conformation of L-CCG-IV (5) and to the media-syn
isomers of 1-aminospiro[2.2]pentyl-1,4-dicarboxylic acid
(22d ). The analysis of this plot clearly confirms that the
extended pseudoanti-anti conformation of L-CCG-I is the
bioactive conformation at mGluR1, whereas the eclipsed-
g+ of L-CCG-IV (5) is the preferred one at the NMDA
receptor. Most interestingly, the plot suggests the medial-
syn isomers of 1-aminospiro[2.2]pentyl-1,4-dicarboxylic
acid (22d ) as a potential NMDA-preferring ligand and
the medial-anti 1-aminospiro[2.2]pentyl-1,4-dicarboxylic
acid (22b) as a potential mimick of the mGluR1-bioactive
conformation of L-Glu (1).
Biologica l Testin g of 1-Am in osp ir o[2.2]p en tyl-1,4-
d ica r boxylic Acid s. The newly synthesized derivatives
22a -d were preliminarily tested in a variety of gluta-
matergic receptor assays, including (i) binding to iono-
tropic glutamate receptors (NMDA, AMPA, and KA), (ii)
F IGURE 3. Superimposition of 22b, 22c, and 22d on the
proposed bioactive conformation of L-Glu (1) when acting at
mGluR1, AMPA, and NMDA, respectively. The superimposi-
tion was accomplished by fitting the amino acidic moiety and
evaluation of their ability to enhance or inhibiting
mGluR1- or mGluR5-induced PI turnover, (iii) evaluation
of their ability to enhance or inhibit the mGluR2-induced
GTP(γ)-35S stimulation. No agonistic or antagonistic
the distal carboxylate oxygens. Only 22c matches the lone pair
activity was displayed up to a concentration of 50 µM at
mGluR1 and mGluR5. On the other hand, two of the
isomeric pairs significantly inhibit the binding of [3H]-
Glu to NMDA receptors: 22c with an IC50 ) 17.5 µM
and 22b with an IC50 ) 39.8 µM.
The preliminary pharmacological characterization of
the new derivatives (22a -d ) did not reveal any particular
activity at glutamate receptors, with exception of weak
affinity of 22b and 22c to the NMDA receptor. It is worth
analyzing these data having in mind the results reported
in Figures 2 and 3. In particular, if the distances between
functional (pharmacophoric) groups are taken as an
indicator of the spatial arrangement of 22a -d , thus using
projection of L-Glu (1).
the concept of folded and extended conformations, then
one would have predicted the new derivatives more active
than they actually are. In particular, the extended
derivative 22b could have been predicted as a mGluR1
ligand. The lack of activity can be attributed either to
the presence of steric bumps or, more likely, to the
peculiar orientation of the functional groups. Indeed, the
visual comparison of the conformations of L-Glu (1) closer
to the individual 22a -d isomers allows verification that,
despite a similarity in d1-d2 distances, the carboxylic
groups have quite a different orientation, which results
5504 J . Org. Chem., Vol. 67, No. 16, 2002

1-Aminospiro[2.2]pentyl-1,4-dicarboxylic Acids
separated. The aqueous phase was extracted with pentane
(3 × 70 mL). The combined organic phases were washed with
water (1 × 70 mL) and dried (Na2SO4). The pentane solution
was removed at atmospheric pressure using a 20-cm Vigreux
column, and the product 14 was distilled: bp 69-70 °C, 19
mbar (6.1 g, 46%). Analytical data were identical with those
previous reported.30
(()-4-(ter t-Bu tyl)-1,1-d im eth yl Sp ir o[2.2]p en ta n e-1,1,4-
tr ica r boxyla te (17). A solution of dimethyl diazomalonate
(0.20 g, 1.25 mmol) in dry dichloromethane (5 mL) was added
via syringe pump over a period of 3 h to a magnetically stirred
solution of 14 (0.20 g, 1.3 mmol) and dirhodium(II) tetraacetate
(0.02 g, 0.045 mmol) in dry dichloromethane (10 mL) kept
under argon at reflux. The reaction mixture was then filtered,
and after evaporation of the solvent, the residue was submitted
to a flash chromatography. Elution with light petroleum-
EtOAc (90:10) afforded 17 (0,121 g, 33%): 1H NMR (CDCl3) δ
1.45-1.70 (m, 11H), 1.90 (d, 1H, J ) 4.8 Hz), 2.00-2.15 (m,
2H), 3.70 (s, 6H); 13C NMR (CDCl3) δ 13.6, 20.0, 21.3, 27.3,
28.0, 52.3, 80.7, 168.5, 170.7.
(()-4-(ter t-Bu tyl)-1-m eth yl Sp ir o[2.2]p en ta n e-1,1,4-tr i-
ca r boxyla tes (18 + 19). A solution of NaOH (0.182 g, 4.55
mmol) in water (1 mL) was added to a magnetically stirred
solution of 17 (1.29 g, 4.55 mmol) in methanol (1 mL). After
72 h of stirring methanol was evaporated in vacuo. The
residual aqueous layer was diluted and extracted with ether
(3 × 4 mL), acidified with a 10% citric acid solution, and
extracted with ethyl acetate (4 × 10 mL). The combined
extracts were dried (Na2SO4) and evaporated to give the
mixture of the two unseparated diastereoisomers 18 + 19 (0.89
g, 72%): 1H NMR (CDCl3) δ 1.30-1. 70 (m, 11H), 1.90-2.30
(m, 3H), 3.70 (s, 3H), 10.00 (brs, 1H).
in an unoptimized interaction of their electronic lone
pairs with the receptorial counterpart (Figure 3).
Thus, our results indicate that the “folded-extended”
concept, based on the distances between functional
groups, although operatively useful and immediate, may
not be appropriate when discussing structure-activity
relationships.
Con clu sion
A diastereoselective route to the synthesis of all four
possible racemic isomers of 1-amino-spiro[2,2]pentyl-1,4-
dicarboxylic acids (22a -d ) has been reported along with
their conformational characterization and preliminary
biological evaluation as glutamate receptor ligands. The
inclusion of the glutamate skeleton into a spiro[2.2]-
pentane scaffold generated a library of molecules with
interesting and quite unusual conformational properties.
When tested on a series of glutamate receptors, only a
moderate affinity to the NMDA receptor could be ob-
served for two pairs (21b and 22c), whereas no activity
at metabotropic receptors was seen. Although these
preliminary biological results are unsatisfactory, the
availability of a synthetic route to this class of confor-
mational analogues of L-Glu (1) can be of importance
since the peculiar conformational properties of the new
derivatives may be exploited in the design of novel
peptidomimetics or in the incorporation of enzyme inhibi-
tors.
(()-4-(ter t-Bu t yl) 1-Met h yl-1-(ter t-b u t oxyca r b on yl)-
a m in o Sp ir o[2.2]p en ta n e-1,1,4-tr ica r boxyla tes (20 + 21).
A magnetically stirred solution of 18 + 19 (0.220 g, 0.81 mmol)
in dry THF (2.5 mL) was treated with potassium carbonate
(0.226 g, 1.63 mmol) and 18-crown-6 (0.033 g, 0.12 mmol) at 0
°C under argon. A solution of ethyl chloroformate (0.098 g, 0.90
mmol) in dry THF (0.5 mL) was then added dropwise, and the
mixture was stirred for 30 min at 0 °C and 90 min at room
temperature. The solution was quickly filtered to remove solids
(washed with 1.5 mL of THF). The solution was recooled to 0
°C, whereupon a solution of sodium azide (0.059 g, 0 0.90
mmol) in water (0.83 mL) was rapidly added. After stirring
for 30 min, the mixture was poured into water (10 mL) and
extracted with ether (3 × 10 mL). The combined ether extracts
were washed with brine (12 mL), dried (Na2SO4), and evapo-
rated in vacuo to give the intermediate acyl azide (0.183 g,
76%). The acyl azide was taken up in tert-butyl alcohol (5.0
mL) and heated to reflux for 17 h. The mixture was cooled
and the solvent was removed in vacuo. The resulting residue
was submitted to a flash chromatography. Elution with light
petroleum-EtOAc (70:30) afforded an unseparated mixture
of 20 + 21 (0.161 g, 58%): 1H NMR (CDCl3) δ 0.80-1.00 (m,
1H), 1.40-1.70 (m, 21H), 1.90-2.10 (m, 1H), 3.73 (s, 3H), 5.20
(brs, 1H).
(()-d ista l- a n d (()-m ed ia l-syn -1-Am in o Sp ir o[2.2]p en -
t a n e-1,4-d ica r boxylic Acid s (22c + 22d ). A mixture of
20 + 21 (0.128 g, 0.37 mmol) and 6 N hydrochloric acid (13
mL) was heated at 80 °C for 8 h. The reaction mixture was
evaporated to dryness, and the residue was submitted to ion
exchange chromatography (Dowex 50WX2-200). Elution with
10% pyridine afforded an unseparated mixture of 22c + 22d
(0.040 g, 63%): 1H NMR (D2O) δ 1.40-11.65 (m, 3H), 1.75-
1.85 (2xd, 1H, J ) 6.4 Hz), 2.00-2.20 (m, 1H); 13C NMR (D2O)
δ 13.7, 14.9, 17.0, 20.0, 20.8, 27.7, 40.4, 172.8, 172.9, 176.8,
177.0. These spectral data are in agreement with those
reported below for single isomers 22c and 22d .
Exp er im en ta l Section
Ch em istr y. Melting points were determined by the capil-
lary method and are uncorrected. 1H and 13C NMR spectra
were obtained with 200 MHz (unless noted) spectrometers. The
abbreviations used are as follows: s, singlet; dd, double
doublet; m, multiplet, p, pseudo. Bidimensional NOESY and
COSY experiments were performed on 400 MHz spectrometer.
TLC were carried out on precoated TLC plates with silica gel
60 F-254. For column chromatography, silica gel 60 (230-400
mesh) was used. GC-MS analyses were carried out with a
gas chromatograph (12.5 m column) equipped with a mass-
selective detector. GC analyses were performed on a gas
chromatograph with a 30 m capillary column.
All final compounds 22a -d displayed an HPLC purity >
95%. HPLC analyses were performed with a workstation
equipped with a UV-vis detector using the following param-
eters: column, 2.5 × 25 mm, 0.5 µm RP-18; flow rate, 0.8 mL/
min; detection at 210 nm; mobile phase, H2O + 0.05%TFA.
(()-ter t-Bu tyl-2-m eth yl, 2-Br om o-cyclop r op a n e-1-ca r -
boxyla te (16). A solution of tert-butyl diazoacetate (58.4 g,
410 mol) in dry dichloromethane (500 mL) was added under
argon via syringe pump to a magnetically stirred solution of
2-bromopropene (15) (38.5 g, 320 mol) and dirhodium(II)
tetraacetate (1.35 g, 3.1 mmol) in dry dichloromethane (400
mL) over a period of 96 h. The reaction mixture was then
filtered, and after evaporation of the solvent, the product 16
was isolated by distillation as a mixture of cis and trans
isomers: bp 55-59 °C, 0.2 mbar (64.3 g, 86%); 1H NMR
(CDCl3) δ 1.40 (dd, 1H, J ) 6.7 and 13.6 Hz), 1.45-1.55 (m,
10 H), 1.87 (s, 3H), 2.25 (dd, 1H, J ) 6.7 and 9.1 Hz).
(()-ter t-Bu t yl-2-m et h ylen e-cyclop r op a n e-1-ca r b oxy-
la te (14). A solution of 16 (20.3 g, 86.3 mmol) in dry DMSO
(11 mL) was added dropwise in 20 min to a magnetically
stirred solution of potassium tert-butoxide (14.4 g, 128.3 mmol)
in dry DMSO (87.5 mL) kept under argon at room tempera-
ture. The reaction mixture was stirred for 30 min at room
temperature and then quenched with cold water (150 mL).
Pentane was added (150 mL) and the organic phase was
(()-(ter t-Bu tyldim eth ylsilyloxym eth yl)-2-m eth ylen ecy-
clop r op a n e (24). A solution of methylenecyclopropylmethanol
(30) de Meijre, A.; Kozhushkov, S. I.; Spaeth, T.; Zefirov, N. S. J .
Org. Chem. 1993, 58, 502-505.
J . Org. Chem, Vol. 67, No. 16, 2002 5505

Pellicciari et al.
(23) (6.61 g, 78.6 mmol) in dry dichloromethane (200 mL) was
treated with 4-(dimethylamino)pyridine (9.6 g, 78.6 mmol).
After stirring for 10 min, this mixture was treated with tert-
butyldimethylsilyl chloride (12.15 g, 80 mmol), and the result-
ing solution was stirred overnight at room temperature under
argon. The reaction was quenched with a saturated solution
of sodium bicarbonate. The aqueous phase was extracted with
dichloromethane (3 × 100 mL). The combined extracts were
washed with brine (1 × 50 mL), dried (Na2SO4), and evapo-
rated, and the mixture was purified by flash chromatography
(n-hexane) to give 24 (8.5 g, 55%) as a colorless oil: 1H NMR
(CDCl3) δ 0.01 (6H, s,), 0.83 (9H, s), 1.20 (2H, m), 1.64 (1H,
m), 3.36 (1H, dd, J ) 11 and 7.5 Hz), 3.59 (1H, dd, J ) 11 and
6.8 Hz), 5.3 (1H, m), 5.35 (1H, m).
(()-(pr oxim a l)-4-(ter t-Bu t yld im et h ylsilyloxym et h yl)-
sp ir o[2.2]p en ta n e-1,1-d ica r boxylic Acid Dieth yl Ester
(25) an d (()-(dista l)-4-(ter t-Bu tyldim eth ylsilyloxym eth yl)-
sp ir o[2.2]p en t a n e-1,1-d ica r b oxylic Acid Diet h yl E st er
(26). A solution of diethyldiazomalonate (13.12 g, 70.4 mmol)
in dry dichloromethane (600 mL) was added via syringe pump
over 90 h to a magnetically stirred solution of the olefin 24
(3.5 g, 17.6 mmol) and dirhodium(II) tetraacetate (0.35 g, 0.78
mmol) in dry dichloromethane (200 mL) at reflux under argon.
The reaction mixture was filtered and evaporated to dryness.
The residue was submitted to flash chromatography; elution
with light petroleum ether-EtOAc (98:2) gave a mixture of
25 and 26 as a colorless oil (25/26 ) 1/5, GC-MS). The mixture
of diastereoisomers 25 and 26 was separated by MPLC.
(cyclohexanes-EtOAc, 95:5) to give 25 (0.7 g, 11%) and 26 (3.3
g, 53%). Da ta for 25: 1H NMR (CDCl3) δ 0.10 (6H, bs), 0.94
(9H, bs), 1.14-1.17 (1H, m), 1.27-1.37 (6H, m), 1.73-1.90 (3H,
m), 3.23 (1H, dd, J ) 8.6 and 10.8 Hz),4.02 (1H, dd, J ) 4.5
and 11.4 Hz), 4.19-4.30 (4H, m); 13CNMR (CDCl3) δ 5.25.2
10.114.0 14.218.4 20.6,21.3, 26.0 28.1 32.9, 61.2, 61.2, 64.4,
169.0, 169.4; GC-MS m/z (relative intensity) 355 (M - H+, 1),
299 (100), 225 (32), 181 (16), 75 (38). Da ta for 26: 1H NMR
(CDCl3) δ 0.01 (6H, bs), 0.85 (9H, bs),1.19-1.26 (7H, m), 1.51-
1.98 (3H, 2m,), 3.43 (1H, dd, J ) 2 6.6 and 10.6 Hz), 3.72 (1H,
dd, J ) 5.3 and 11.4 Hz), 4.06-4.19 (4H, m); 13C NMR (CDCl3)
δ 3.9, 3.8, 11.8, 15.6, 19.7, 20.6, 21.2, 27.1, 29.4, 34.4, 62.5,
61.6, 65.7, 170.4, 170.5; GC-MS m/z (relative intensity) 355
(M - H+, 1), 299 (100), 225 (33), 181 (67), 75 (83).
(()-pr oxima l- an d (()-media l-a n ti-4-(ter t-Bu tyldim eth -
ylsilyloxym eth yl)-spir o[2.2]pen tan e-1,1-dicar boxylic Acid
Mon oeth yl Ester s (27a + 27b). A solution of 0.5 M sodium
hydroxyde (8.4 mL, 4.2 mmol) in water was added to a solution
of 25 (1.5 g, 4.2 mmol) in 95% ethanol, and the resulting
mixture was stirred at room temperature for 36 h. The reaction
mixture was evaporated to dryness, water was added to the
residue, and the mixture was neutralized with 10% citric acid
solution and extracted with ethyl acetate (3 × 100 mL). The
combined extracts were dried (Na2SO4) and evaporated to give
the mixture of products 27a and 27b (1.25 g, 90%) as a
colorless oil: 1H NMR (CDCl3) δ 0.04 (6H, bs), 0.79 (9H, bs),
0.95-1.20 (1H, m), 1.27-1.37 (6H, m), 1.73-2.20 (3H, m), 3.50
(1H, m), 3.80 (1H, m), 4.07-4.30 (2H, m).
(()-d ista l- a n d (()-m ed ia l-syn -4-(ter t-Bu tyld im eth yl-
silyloxym eth yl)-sp ir o[2.2]p en ta n e-1,1-d ica r boxylyc Acid
Mon oeth yl Ester s (27c + 27d ). Monohydrolysis of 26 (4.6
g, 13.0 mmol) was performed according to the procedure
described above for 25. The mixture of the isomers 27c + 27d
(3.3 g, 78%) was obtained as a colorless oil: 1H NMR (CDCl3)
δ 0.04 (6H, s), 0.79 (9H, m), 1.05-1.20 (1H, m), 1.21-1.33 (4H,
m), 1.53-1.75 (1H, m), 3.30-3.52 (1H, m), 3.70 (0.5H, dd, J )
8.6 and 10.8 Hz), 3.90 (0.5H, dd, J ) 4.5 and 11.4 Hz), 4.10-
4.30 (2H, m).
g, 0.38 mmol), and freshly distilled ethylchloroformate (0.49
g, 4.54 mmol). The mixture was kept at 0 °C for 30 min and
at room temperature for 90 min. The reaction mixture was
filtered quickly under argon, and the remaining solid was
washed with tetrahydrofurane (2 × 10 mL). A solution of
sodium azide (0.29 g, 4.54 mmol) in water (5 mL) was added
to the filtrate at 0 °C under argon. After 1 h, the reaction
mixture was poured into cold water (100 mL) and extracted
with ethyl acetate (3 × 100 mL). The combined extracts were
dried (Na2SO4) and evaporated. The residue was dissolved in
tert-butyl alcohol (30 mL), and the solution was refluxed
overnight under argon. The reaction mixture was concentrated
under reduced pressure, and the residue was submitted to
flash chromatography; elution with light petroleum ether-
EtOAc (9:1) gave the product 28a + 28b (0.45 g, 30%) as a
mixture of diastereoisomers: 1H NMR (CDCl3) δ 0.04 (6H, s),
0.78 (9H, s), 0.80-1.00 (1H, m), 1.18-1.28 (3H, m), 1.40 (9H,
s), 1.50-1.70 (1H, m), 1.80-1.98 (1H, m), 3.18-3.32 (1H, m),
3.83-3.91 (1H, m), 3.98-4.26 (4H, m).
(()-d ista l- a n d (()-m ed ia l-syn -4-(ter t-Bu tyld im eth yl-
silyloxym eth yl)-1-ter t-bu toxyca r bon yl Am in osp ir o[2.2]-
p en t a n e-1-ca r b oxylic Acid E t h yl E st er s (28c + 28d ).
Starting from the mixture 18c + 18d (2.4 g, 7.32 mmol) the
isomers 19c + 19d were obtained (0.94 g, 32%) according to
the procedure described above for 18a + 18b: 1H NMR (CDCl3)
δ 0.04 (6H, m), 0.82 (9H, m), 1.05-1.35 (4H, m), 1.42 (9H, bs),
1.45-1.90 (3H, m), 3.38 (1H, dd, J ) 6.6 and 10.6 Hz), 3.85
(1H, dd, J ) 5.3 and 11.4 Hz), 4.08-4.22 (4H, m).
(()-pr oxim a l-1-ter t-Bu toxyca r bon yla m in o-4-h yd r oxy-
m eth ylsp ir o[2.2]p en ta n e-1-ca r boxylic Acid Eth yl Ester
(29a ) a n d (()-m ed ia l-a n ti-1-ter t-Bu toxyca r bon yla m in o-
4-h ydr oxym eth ylspir o[2.2]pen tan e-1-car boxylic Acid Eth -
yl Ester (29b). A solution of 28a + 28b (0.44 g, 1.1 mmol) in
distilled tetrahydrofurane (30 mL) was treated with tetra-n-
butylammonium fluoride (0.44 g, 1.1 mmol). The solution was
stirred at room temperature until no starting material was
detected by TLC. Water (150 mL) was added to the mixture,
and the aqueous phase was extracted with ethyl acetate (4 ×
50 mL). The combined extracts were dried (Na2SO4) and
evaporated to give the mixture of derivatives 29a and 29b,
which was separated by MPLC; elution with light petroleum
ether-EtOAc (3:2) gave 29a (0.065 g, 21%) and 29b (0.202 g,
1
65%) as white solids. Da ta for 29a : mp 77-79 °C; H NMR
(CDCl3) δ 0.72 (1H, pt, J ) 4.9 Hz) 1.05 (1H, dd, J ) 5.0 and
8.2 Hz), 1.17 (3H, t, J ) 7.1 Hz), 1.40 (9H, bs), 1.60-1.70 (1H,
m), 2.01 (1H, d, J ) 4.5 Hz), 3.22 (1H, pt, J ) 10.8 Hz) 3.96-
4.19 (3H, m); 13C NMR (CDCl3) δ 10.2, 14.3, 20.8, 23.3, 28.3,
61.1, 64.0, 79.8, 157.4, 172.0. Da ta for 29b: mp 89-81 °C;
1H NMR (CDCl3) δ 0.84 (2H, m), 1.18 (3H, t, J ) 7.1 Hz), 1.37
(10H, m), 1.66-1.80 (1H, m), 1.85 (1H, d, J ) 4.4 Hz), 3.40
(1H, pt, J ) 8.2 Hz), 3.81 (1H, dd, J ) 3.8 and 11.6 Hz), 4.02-
4.22 (2H, m); 13C NMR (CDCl3) δ 7.7, 14.1, 22.4, 24.0, 28.2,
30.9, 38.7, 62.0, 63.6, 80.0, 155.9, 172.4.
(()-dista l-1-ter t-Bu toxycar bon ylam in o-4-h ydr oxym eth -
ylsp ir o[2.2]p en ta n e-1-ca r boxylic Acid Eth yl Ester (29c)
and (()-media -syn-1-tert-Butoxycarbonylamino-4-hydroxy-
m eth ylsp ir o[2.2]p en ta n e-1-ca r boxylic Acid Eth yl Ester
(29d ). Both isomers were prepared as described above for
spiropentanes 29a and 29b from a mixture of 19c + 19d (0.6
g, 2.33 mmol). MPLC in light petroleum ether-EtOAc (3:2)
gave 29c (360 mg, 54%) and 29d (230 mg, 35%) as white solids.
1
Da ta for 29c: mp 84-86 °C; H NMR (CDCl3) δ 0.84 (1H, t,
J ) 5.2 Hz), 1.20 (3H, t, J ) 7.1 Hz), 1.35-1.50 (10H, m), 1.50-
1.72 (1H, m), 1.93-2.10 (2H, 2m), 3.23 (1H, dd, J ) 9.2 Hz,
10.6 Hz), 3.85 (1H, dd, J ) 4.8 and 10.6 Hz), 4.11 (2H, q, J )
7.1 Hz); 13C,NMR (CDCl3) δ 11.3, 14.3, 19.5, 20.2, 20.8, 28.2,
29.2, 29.8, 38.9, 61.2, 65.3, 80.1, 156.1, 172.0. Da ta for 29d :
mp 82-84 °C; 1HNMR (CDCl3) δ 0.78 (1H, t, J ) 5.1 Hz), 1.10
(3H, t, J ) 7.1 Hz), 1.35-1.50 (10H, m), 1.50-1.72 (1H, m),
1.93-2.10 (2H, 2m), 3.62 (2H, m), 4.17 (2H, m); 13C NMR
(CDCl3) δ 10.4, 14.2, 20.6, 21.4, 28.2, 30.5, 38.6, 61.7, 65.1,
80.0, 155.9, 172.9.
(()-pr oxima l- an d (()-media l-a n ti-4-(ter t-Bu tyldim eth -
ylsilyloxym eth yl)-1-ter t-bu toxyca r bon ylAm in osp ir o[2.2]-
p en ta n e-1-ca r boxylic Acid Eth yl Ester s (28a +28b). A
stirred solution of 27a + 27b (1.24 g, 3.78 mmol) in dry
tetrahydrofurane (30 mL) at 0 °C under argon was treated
with potassium carbonate (1.04 g, 7.55 mmol), 18-crown-6 (0.10
5506 J . Org. Chem., Vol. 67, No. 16, 2002

1-Aminospiro[2.2]pentyl-1,4-dicarboxylic Acids
(()-1-Am in osp ir o[2.2]p en t a n -1,4-d ica r b oxylic Acid s
(22a -d ). A magnetically stirred ice-cold solution of a single
isomer of Boc-aminospiropentanes 29a , 29b, 29c, or 29d (0.06
g, 0.21 mmol) in acetone (10 mL) was treated with 8 N J ones’
reagent (about 0.5 mL) until a persistent orange color was
reached. The reaction mixture was stirred at room tempera-
ture for 1 h. 2-Propanol (1 mL) was added, and after 10 min,
the mixture was filtered through Celite, washing with metha-
nol. The solvent was removed under reduced pressure, and
the residue was used for the next reaction without further
purification. HCl (6 N, 20 mL) was added to a solution of crude
mixture, and the reaction was stirred at 90 °C for 16 h. The
solvent was removed under reduced pressure. The residue was
submitted to ion-exchange resin chromatography (Dowex
50X2-200, 5% pyridine) to give the title compound 22a , 22b,
22c, or 22d .
Da ta for pr oxim a l isom er 22a : 65% yield; mp 239-240
°C; 1H NMR (400 MHz, D2O) δ 1.44 (1H, t, J ) 4.7 Hz, 5-CHa),
1.58-1.61 (2H, m, 2-CHa and 5-CHb); 1.80 (1H, d, J ) 6.3 Hz,
2-CHb); 2.30 (1H, dd, J ) 4.8 and 8.3 Hz, 4-CH); 13C NMR
(D2O) δ 15.5, 16.0, 21.3, 26.2, 40.0, 172.5, 176.7.
Da ta for m ed ia l-a n ti isom er 22b: 61% yield; mp 208-
210 °C; 1H NMR (400 MHz, D2O) δ 1.54-1.61 (2H, m, 5-CH2),
1.65 (1H, d, J ) 6.4 Hz, 2-CHa), 1.88 (1H, d, J ) 6.4 Hz, 2-CHb),
2.22 (1H, dd, J ) 4.9 and 7.8 Hz, 4-CH); 13C NMR (D2O) δ
13.2, 17.7, 21.0, 26.9, 39.3, 172.6, 176.3.
Da ta for d ista l isom er 22c: 71% yield; mp 210-212 °C;
1H NMR (400 MHz, D2O) δ 1.58 (1H, t, J ) 4.8 Hz, 5-CHa),
1.65 (1H, dd, J ) 5.0 and 8.4 Hz, 5-CHb),1.71 (1H, d, J ) 6.5
Hz, 2-CHa), 1.86 (1H, d, J ) 6.6 Hz, 2-CHb), 2.14 (1H, dd, J )
5.0 and 8.2 Hz, 4-CH); 13C NMR (D2O) δ 13.6, 16.8, 20.7, 27.6,
40.1, 171.4, 177.0.
Da ta for m ed ia l-syn isom er 22d : 68% yield; mp 170-
172 °C; 1H NMR (400 MHz, D2O) δ 1.55-1.61 (2H, m, 5-CH2),
1.64 (1H, d, J ) 6.5 Hz, 2-CHa), 1.94 (1H, d, J ) 6.5 Hz, 2-CHb),
2.23 (1H, dd, J ) 4.8 and 8.2 Hz, 4-CH); 13C NMR (D2O) δ
14.9, 16.9, 19.9, 27.6, 40.2, 173.0, 177.1.
given in Table 1. Absorption correction obtained by Ψ scans
was applied to all the data collections. The structures were
solved by direct methods implemented in the SHELX-97
program.31 The refinements were carried out by full-matrix
anisotropic least-squares on F2 for all reflections for non-H
atoms by using the SHELX-97 program.31
Com p u ta tion a l Meth od s. All studied compounds were
built using the Cerius2 libraries using molecular mechanics
and semiempirical calculations. In particular, the structures
of compounds 22a -d were geometry optimized, in their
neutral form, using the MOPAC-AM1 method. The geometries
thus obtained for compounds 22b-d were in good agreement
with the experimentally determined ones. L-Glutamic acid (1)
and the CCGs (4, 5) were submitted to a conformational search
using the grid search routine implemented in Cerius2. Two
torsional angles for L-Glu (1) (t1 defined as C1-C2-C3-C4, t2
defined as C2-C3-C4-C5) and one torsional angle for the CCG
4 and 5 (t1 defined as Ca-Cb-Cc-Cd) were analyzed in a range
of 360° with a step increment of 1°. The energetical profiles
were evaluated by using the Universal force field implemented
in Cerius2 and by correcting the electrostatic contribution by
setting a dielectric constant of 80. All conformers were
minimized using the conjugate gradient until a convergence
gradient of 0.001 was reached. After this protocol, the unique
conformers were stored and used for further analysis. All
calculations were carried out on a SGI O2 R5000 machine
using the Cerius2 software package of Accelrys.
Biology. The preliminary biological screenings were per-
formed as previously reported.32
Ack n ow led gm en t. Financial support of Italian
Minister of University (MURST, COFIN2000) to R.P.
is gratefully acknowledged.
Su p p or tin g In for m a tion Ava ila ble: Characterization
data and spectra for 22a -d . This material is available free of
charge via the Internet at http://pubs.acs.org.
X-r a y An a lysis. Single crystals of 22b-d were obtained
by dissolution in water-48% HBr (10:1) and slow evaporation
at room temperature. Colorless single crystals of each com-
pound were submitted to X-ray data collection on a four-circle
diffractometer with graphite monochromated Mo K radiation
(λ ) 0.71069 Å). Lattice parameters were determined by least-
squares refinement on 40, 49, and 60 randomly selected and
automatically centered reflections, respectively. The ω/2θ scan
technique was used in the data collection. Crystal details are
J O020138V
(31) Sheldrick, G. M. SHELX-97, rel.97-2; University of Goettin-
gen: Goettingen, 1997.
(32) Gasparini, F.; Lingenhohl, K.; Stoehr, N.; Flor, P. J .; Heinrich,
M.; Vranesic, I.; Biollaz, M.; Allgeier, H.; Heckendorn, R.; Urwyler,
S.; Varney, M. A.; J ohnson, E. C.; Hess, S. D.; Rao, S. P.; Sacaan, A.
I.; Santori, E. M.; Velicelebi, G.; Kuhn, R. Neuropharmacology 1999,
38, 1493-1503.
J . Org. Chem, Vol. 67, No. 16, 2002 5507