CoMFA, synthesis, and pharmacological evaluation of (E)-3-(2-carboxy-2- arylvinyl)-4,6-dichloro-1H-indole-2-carboxylic acids: 3-[2-(3-Aminophenyl)-2- carboxyvinyl]-4,6-dichloro-1H-indole-2-carboxylic acid, a potent selective glycine-site NMDA receptor antagonist

J. Med. Chem. 2005, 48, 995-1018

995

CoMFA, Synthesis, and Pharmacological Evaluation of

(E)-3-(2-Carboxy-2-arylvinyl)-4,6-dichloro-1H-indole-2-carboxylic Acids:

3-[2-(3-Aminophenyl)-2-carboxyvinyl]-4,6-dichloro-1H-indole-2-carboxylic Acid, a

Potent Selective Glycine-Site NMDA Receptor Antagonist^†

Bruce M. Baron, Robert J. Cregge, Robert A. Farr, Dirk Friedrich, Raymond S. Gross, Boyd L. Harrison,

David A. Janowick, Donald Matthews, Timothy C. McCloskey, Scott Meikrantz, Philip L. Nyce, Roy Vaz, and

William A. Metz*

Department of Medicinal Chemistry, Aventis Pharmaceuticals, Route 202-206, Bridgewater, New Jersey 08807-0800

Received October 12, 2004

(E)-3-(2-Carboxy-2-phenylvinyl)-4,6-dichloro-1H-indole-2-carboxylic acid, 1, is a potent and

selective antagonist of the glycine site of the N-methyl-^D-aspartate (NMDA) receptor. Using

3D comparative molecular field analysis (CoMFA) to guide the synthetic effort, a series of aryl

diacid analogues of 1 were synthesized to optimize in vivo potency, duration of action, and

binding activity. It was found that the incorporation of a substituted aromatic with an electron

withdrawing group or a heterocyclic group at the 2-position of the 3-propenyl moiety of 1 gave

compounds with better affinity and potency in the murine stroke model. Ultimately this led to

the discovery of 3-[2-(3-aminophenyl)-2-carboxyvinyl]-4,6-dichloro-1H-indole-2-carboxylic acid,

19, as a new potent selective glycine-site NMDA receptor antagonist.

Introduction

Overactivation of the N-methyl-D-aspartate (NMDA)

receptor, an ionotropic glutamate receptor in the brain

and central nervous system, by elevated levels of

glutamic acid as a result of a neurodegenerative event

such as cerebral ischemia or neurotrauma, allows excess

calcium to enter the neuronal cell.¹This calcium influx

activates endogenous, calcium-dependent, degradative

enzymes that are believed to destroy the neuron cell

before the damaged cell can mutate. Glutamate antago-

nists have been shown to disrupt this chain of events

at the surface of the cell at the glycine NMDA site.²

Figure 1. Potent NMDA inhibitors.

Glycine, an obligatory coagonist for the NMDA recep-

tor,³activates ion channels at a strychnine-insensitive

binding site⁴distinct from that utilized by glutamate.⁵

It has been demonstrated that glycine antagonists can

mediate NMDA receptor response in vitro and in vivo⁶

and are noncompetitive antagonists of glutamate. In-

tervention at this site could offer a potential mechanistic

advantage, since a glycine antagonist would not have

to compete with the ischemia-induced elevated levels

of glutamate. Therefore, a glycine-site NMDA receptor

antagonist might have an application for neuroprotec-

tion in stroke and head trauma.⁷

initiated on the basis of 3D comparative molecular field

analysis (CoMFA) of 1 to improve the pharmacological

properties.

CoMFA of Inhibitor 1

CoMFA is a well accepted three-dimensional quanti-

tative structure-activity relationship (3D-QSAR) method

for providing insight into the possible conformation of

a molecule close to or in its active form. We have used

CoMFA to establish a 3D structure that could represent

the active conformation of 1 on the basis of a set of

analogues.

CoMFA studies were performed on approximately 300

compounds related to 1 to provide a series of target

molecules for synthesis and biological evaluation. All

structures shown in Table 1 were constructed in SYBYL

6.6.⁹Compound 1 was subjected to conformational

analysis, and the lowest energy conformation, calculated

using the Tripos 5.2 force field as available within

SYBYL, was used as a template to construct the other

structures listed in Table 1. The ionizable groups in each

compound were kept in the neutral state. All structure-

swere then subjected to optimization using the AM1

Compound 1 (Figure 1) is a potent glycine antagonist

with affinity for the glycine-site NMDA receptor (IC₅₀

vs [³H]glycine of 11 nM) and is centrally active as

demonstrated by anticonvulsant activity (ED₅₀vs au-

diogenic seizures of 12 mg/kg intraperitoneally (ip) and

29 mg/kg intraveneously (iv) vs maximal electroshock

in the rat).⁸A chemical optimization program was

^†Dedicated to Prof. John “Jack” Martin on the occasion of his return

to the classroom from retirement.

* To whom correspondence should be addressed. Phone: (908) 231-

3602. Fax: (908) 231-3602. E-mail: william.metz@aventis.com.

Published on Web 02/01/2005

996 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Table 1. Compounds Used in the CoMFA

Baron et al.

Hamiltonian in MOPAC 4.0 ¹⁰(as supplied within SY-

BYL). The structures were then overlaid using the in-

dole group and the adjacent carboxylic group as the

template.

sented in Figure 2. The positive (0.003) steric field coef-

ficients are shown in green, the negative coefficients

(-0.003) in red, and the electrostatic field coefficients

are shown in yellow (0.009) and cyan (-0.009), respec-

tively.

The overlap of the positive steric coefficient contour

with the positive electrostatic contour suggests a posi-

tively charged or less negatively charged substitution

pattern in that region. The negative coefficient contours

clearly delineate a pocket in the NMDA receptor into

which the compounds fit with the phenyl group of

compound 1 sitting in a pocket that conservatively

prefers a relatively less negatively charged substituent.

As a result of the CoMFA studies a chemical optimiza-

tion program was initiated and directed toward modi-

fication or replacement of the phenyl group at the

2-position of the 3-propenyl moiety of 1.

A region was constructed around the parts of the

structure showing variation only as shown in Figure 2.

The dimensions of the CoMFA¹¹region were 23 × 11 ×

20 Å with a 1 Å spacing along the three axes. A 2 kcal

minimum σ value was used, and correspondingly 8401

out of possibly 10121 points were dropped from the

CoMFA. A proton and an sp³carbon were used as probes

in the CoMFA to evaluate the electrostatic and steric

fields, respectively. A cutoff of 30 kcal and greater in

the steric field was used to drop the electrostatic field

computation at the corresponding CoMFA grid points.

Compounds in bold and underlined in Table 1 were not

used in the learning set of the CoMFA and were only

used as a prediction set. The fitted vs actual pIC50 val-

ues are shown in Figure 3 together with the statistics

from the PLS analysis of the 28 compounds in the

learning set.

Chemistry

Compound 1 was first synthesized in seven steps from

3-iodo-2-(ethoxycarbonyl)-4,6-dichloroindole¹²and 2-phen-

yl-3-(tributylstannyl)acrylic acid methyl ester using a

Stille coupling strategy (Scheme 1). For the task of

chemical optimization a more efficient route was neces-

The steric contribution amounted to 48.5%. Contours

of (std dev) × (coeff) (hereafter referred to as coeffi-

cients), for the steric and electrostatic fields are repre-

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 997

Figure 2. Compounds in Table 1 overlaid after optimzation and shown together with the steric and electrostatic coefficients in

the CoMFA region. Also shown are the steric and electrostatic coefficient histograms.

Scheme 1

sary to identify and prepare a common intermediate

that could accommodate the regiospecific introduction

of a variety of substituted aryl groups and heteroaryls

at the 2-position of the 3-propenyl moiety of 1, prefer-

ably at a late stage in the synthetic sequence.

Several different synthetic stategies were investigated

to prepare a number of structurally diverse analogues

of 1. All of the synthetic routes described have advan-

tages and disadvantages depending on the desired

functionality incorporated.

Suzuki-Miyaura Reaction. The in situ palladium-

(0)-catalyzed Suzuki-Miyaura reaction is an efficient

method for the cross-coupling of aryl boranes with vinyl

bromides.¹³The reaction of the (Z)-indole vinyl bromide

8Z with the appropriate arylboronic acid provides easy

access to analogues of 1. An attractive feature of this

approach is that a variety of arylboronic acids are

commercially available, and those of interest which are

not can easily be prepared from the corresponding

aryllithium (halogen metal exchange) species by treat-

ment with a trialkoxyborane.

Vinyl bromide 8Z was prepared on a multigram scale

utilizing the synthetic sequence shown in Scheme 2.

Formylation of 4,6-dichloro-H-indole-2-carboxylic acid

ethyl ester 5¹⁴affords aldehyde 6. Protection of the

indole nitrogen with p-toluenesulfonyl chloride and

treatment with bromophosphonoacetate 4 (prepared

fresh from the corresponding tert-butyl diethoxyphospho-

998 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

Scheme 3^a

^aReactions and conditions: (a) aryl-B(OH)₂, anhyd. K₂CO₃,

(Ph₃P)₄Pd, tol. or Pd₂(dba)₃, (2-furyl)₃Pd, THF; (b) TFA or HCO₂H;

(c) LiOH, THF, H₂O; (d) KHSO₄. Structures: aryl; a ) phenyl; b

) naphthalen-1-yl; c ) 2,4-dichlorophenyl; d ) 2-furyl; e ) 3-furyl;

f ) 4-chlorophenyl; g ) 2-thienyl; h ) 3-thienyl; i ) 4-methoxy-

phenyl.

dipalladium(0) [Pd₂(dba)₃]/tris(2-furyl)phosphine/K₂-

18

CO₃in tetrahydrofuran (THF) at 55-60 °C (Scheme

Figure 3. Statistic describing the PLS method and the fitted

3). Method A was used for the coupling of the phenyl-,

1-naphthyl-, 2,4-dichlorophenyl-, 2-furyl, 3-furyl, and (p-

chlorophenyl)boronic acids, which gave products 9a-f

in 51-71% yield. Method B was employed for the

coupling of the electron-rich 2-thienyl, 3-thienyl, and (p-

methoxyphenyl)boronic acids to afford the coupled

products 9g-i in 77, 49, and 46% yields, respectively.

The coupled products 9a-i possess only the desired

E-stereochemistry, simplifying purification and char-

acterization of the products and their derivatives. The

Suzuki coupling using method A required reaction times

of 4 h to overnight, while couplings using method B

required reaction times of 6-9 days.

and predicted versus actual pIC₅₀values from the CoMFA.

Scheme 2^a

Attempted coupling of (p-methoxyphenyl)boronic acid

to vinyl bromide 8Z, using [1,1′-bis(diphenylphosphino)-

ferrocene]palladium(II) acetate as a catalyst,¹⁹provided

a 5:1 mixture of 9i and unreacted vinyl bromide 8Z after

10 days in THF at 55-60 °C. The use of Pd₂(dba)₃as

the catalyst in the absence of any phosphine ligands, a

procedure reported to greatly increase the rate and

efficiency of coupling,²⁰did not result in any apparent

coupling after 24 h at 55-60 °C.

Products 9a-i were only slightly more polar than the

vinyl bromide 8Z, making the purification of the crude

diester difficult. Deprotection of the tert-butyl esters of

these crude intermediates with either TFA or HCO₂H

afforded the monoacids 10a-i, which were highly

crystalline. As a result, they were easily separated from

the (Z)-bromo acid and other contaminants, such as

dibenzylideneacetone (method B), by fractional recrys-

tallization or by trituration with pentane containing a

small amount of Et₂O. Hydrolysis of 10a-i with LiOH

in THF/H₂O provided the desired diacids 11a-i.

The absence of a significant upfield shift of either the

tert-butyl ester or the vinyl protons in the proton

NMRspectra of 9-11 indicated that isomerization of the

E-acids to the Z-acids did not occur in either the

coupling reaction or the subsequent deprotection steps.

Notably, the proton NMR spectrum of the 1-naphthyl

ester 9b suggested that the naphthyl ring is situated

directly above the 2-carboethoxy group, as the methyl-

ene protons are highly shielded and differentiated,

^aReactions and conditions: (a) Br₂, aq. NaOH, i-PrOH; (b)

SnCl₂, H₂O, t-BuOH; (c) POCl₃, DMF; (d) pTsCl, anhyd. K₂CO₃;

(e) 4, LiHMDS, THF.

noacetate¹⁵via dibromination with NaOBr and mon-

odebromination with SnCl₂) under Horner-Emmons

conditions provided the indole vinyl bromide 8 as an

E/Z mixture. Fractional recrystallization (pentane/

Et₂O) gave the desired vinyl bromide 8Z in 65% yield.

The stereochemistry of the vinyl bromide was as-

signed using proton NMR. The vinyl proton of vinyl

bromide 8Z is deshielded and gives a singlet at 8.2 ppm,

while the proton of 8E is observed as a singlet at 7.5

ppm. The tert-butyl ester protons of vinyl bromide 8Z

give a singlet at 1.6 ppm, while the corresponding

protons of 8E are shielded by the indole ring, shifting

their signal upfield to 1.0 ppm.

Vinyl bromide 8Z was coupled with various com-

mercially available arylboronic acids¹⁶using one of two

methods. Method A involves the use of (Ph₃P)₄Pd/K₂-

17

CO₃as the catalyst in toluene at 90-100 °C, while

method B involves the use of tris(dibenzylideneacetone)

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 999

Scheme 4^a

Scheme 5^a

^aReactions and conditions: (a) HNR₁R₂, EDC, HOBt; (b) LiOH,

THF, H₂O; (c) KHSO₄. Structures: a, R₁) H, R₂) Ph; b, R₁) H,

R₂) Bn; c, R₁) H, R₂) Me; d, R₁, R₂) H; e, R₁, R₂) Me; f, R₁

) H, R₂) CH₂CH₂Ph; g, R₁, R₂) -(CH₂CH₂)₂O; h, R₁) H, R₂)

OBn; i, R₁) H, R₂) OH; j, R₁) tBOC, R₂) OTBDMS.

^aReactions and conditions: (a) NaH/MeOH, THF, HCO₂Et; (b)

MeI, DMF; (c) TMSOTf, CH₂Cl₂; (d) LiOH, THF, H₂O (e) KHSO₄.

Structures: X ) a, p-F; b, p-Cl; c, p-Br; d, p-I; e, p- Me; f, p-CF₃;

g, m-NO₂; h, o-Cl.

absorbing at 3.63 and 2.79 ppm. This conformation was

indeed confirmed in the molecular modeling global

minimization of 9b in preparation for a CoMFA analysis

of diacids 11.

Enol ether 15a was prepared in a two-step, one-pot

reaction starting with methyl p-fluorophenylacetate

(14a), NaH, and HCO₂Me in THF. A drop of MeOH was

added to initiate the reaction and impede the occurrence

of a sudden exotherm. Quenching with MeI in dimeth-

ylformamide (DMF) provided the crude enol ether 15a

in 82% yield, which was subsequently condensed with

the indole ester 5 in the presence of a Lewis acid.

Several acids (BF₃‚OEt₂, Me₃OBF₄, and TiCl₄) were

evaluated for the condensation reaction. TMSOTf pro-

vided the best yield and cleanest conversion to the

desired product. An equivalent amount of Lewis acid

was necessary for the reaction to occur since the

formation of MeOH during the reaction consumes the

catalyst. Typically the enol ethers were used in the

coupling step without further purification due their

instability to silica gel. TMSOTf-promoted coupling

between the indole 5 and enol ether 15a gave the

desired diester 16a in 53% as a 3:2 mixture of E/Z

isomers. Hydrolysis of 16a using aqueous NaOH solu-

tion at 95 °C for 16 h afforded the diacid following acid

workup. Crystallization gave 17a in 71% yield as the

desired E isomer.

The p-chlorophenyl, p-bromophenyl, p-iodophenyl,

p-tolyl, p-trifluoromethylphenyl, m-nitrophenyl, and

o-chlorophenyl analogues, compounds 17b-h, respec-

tively, were prepared in a fashion similar to 17a,

utilizing the corresponding phenylacetic esters 14b-h.

All were obtained as E/Z mixtures. Saponification and

recrystallization of the diacids afforded the desired E

isomers, as identified by the characteristic location of

the vinyl proton at ∼8.1 ppm in the proton NMR

spectra. The application of this procedure appears to be

limited to enol ethers having either electron-withdraw-

ing or weakly electron-donating aryl groups, since

neither the m- and p-methoxyphenyl- nor the 2- or

3-thienyl-substituted enol ethers afforded any of the

desired coupled products.

A key feature in this synthetic route was the intro-

duction of a tert-butyl ester at the vinyl position,

distinguishing that ester from the ethyl ester at the

2-position on the indole ring. The difference in reactivity

of these esters allowed for selective hydrolysis of the tert-

butyl ester 9a with TFA, providing the monoacid 10a,

which upon further functionalization gave a variety of

novel amide derivatives 13a-g (Scheme 4).

Intermediate 10a was reacted with various amines

by activating the carboxylic acid with EDC/HOBT to

afford, after hydrolysis, the amide acid analogues 13a-

g. Synthesis of the hydroxamic acid analogue 13i was

unsuccessful. The initial reaction to couple 10a with

O-benzylhydroxylamine to give the intermediate O-

benzylamide 12h was successful. However, attempted

debenzylation using either HBr/AcOH or PdO/cyclohex-

ene resulted in overreduction to the primary amide 12d.

An alternative route coupling N-t-Boc-O-TBDMS hy-

droxylamine²¹with the acid chloride of 10a (prepared

from the reaction of the carboxylic acid with oxalyl

chloride) gave the N-t-Boc-O-TBDMS amide 12j. Depro-

tection of this compound with TFA/CsF gave the pro-

tected hydroxamic acid 12i. However, removal of the

p-tosyl group and the ethyl ester using either LiOH/

THF/H₂O or K₂CO₃/THF/H₂O resulted in destruction of

the hydroxamic acid moiety as shown by a negative

FeCl₃test. Further attempts were not made to synthe-

size 13i.²²Additional amide analogues of 13 were not

synthesized due to the inactivity of compounds 13a-g

in the DBA/2J audiogenic seizure model. This lack of

activity was attributed to poor CNS penetration.

Nuclear Overhauser effect (NOE) studies on amide

ester 12g confirmed the stereochemical assignments,

since strong NOEs were observed between the vinyl

proton and the proximal morpholine protons, while only

a weak NOE was observed between the vinyl proton and

the phenyl protons.

m-Aminophenyl analogue 19 was synthesized from

m-nitrophenyl diester 16g (Scheme 6). Initial attempts

to reduce the nitro group using Fe(0) gave poor yields

and a mixtue of byproducts. Ultimately SnCl₂was

employed to give the desired m-aminophenyl diester 18

in 92% yield. The caveat in using this method was the

difficult removal of the tin salts formed during the

workup. Conversion of diester 18 to diacid 19 required

rigorous hydrolysis conditions. Following treatment

with LiOH, the aqueous layer was washed with EtOAc

and acidified to pH 4 with 0.5 M NaHSO₄solution. The

Lewis Acid-Catalyzed Enol Ether Condensation.

The utility of the Suzuki-Miyaura coupling reaction

was limited by the substituent on the arylboronic acid

aromatic ring. Arylboronic acids possessing electron-

withdrawing substituents on the aromatic ring either

gave poor yields or no reaction with the starting

material being recovered. The Lewis acid-catalyzed

(trimethylsilyltriflate) condensation of indole ester 5 via

a preformed enol ether 15a-h offered an alternative

method for preparing target compounds 17a-h (Scheme

5).

1000 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

Scheme 6^a

^aReactions and conditions: (a) SnCl₂, H₂O, i-PrOH; (b) LiOH, THF, H₂O; (c) KHSO₄. Structures: a, R₁) H, R₂) COMe; b, R₁) H,

R₂) COPh; c, R₁) H, R₂) CO₂Me; d, R₁) H, R₂) SO₂Me; e, R₁, R₂) Me; f, R₁) H, R₂) Me.

Scheme 7^a

resulting precipitate was extracted with EtOAc. Re-

moval of the solvent under reduced pressure afforded

the crude product that was purified by recrystallization.

Although several solvent systems such as EtOAc, EtOAc/

hexane, acetone, and CH₃CN/H₂O were examined, none

were effective in affording pure product. A two-step

sequence, involving an initial crystallization from EtOAc

followed by precipitation from MeOH with H₂O, gave

pure 19 after drying under a stream of nitrogen, first

at room temperature and then in vacuo at 80 °C.

m-Acylaminophenyl diacids 21a-d were prepared by

treatment of 18 with the corresponding acid chloride

and Et₃N followed by saponification of the crude diesters

20a-d. The resulting diacids were then purified by

^aReactions and conditions: (a) R-CH₂CN, 22a-d; (b) H₂SO₄,

recrystallization. The m-dimethylamino analogue 21e

HOAc; (c) NO₂C₆H₄CH₂CN; (d) 6N NaOH; (e) KHSO₄; ( f) SnCl₂.

was prepared by reductive amination of the m-amino

Structures: 22a, R ) Ph; 22b, R ) 2-pyridyl; 22c, R ) 3-pyridyl;

group.²³Treatment of the m-aminophenyl diester 18

22d, R ) 4-pyridyl.

with paraformaldehyde and NaBH₃CN in HOAc pro-

vided the desired m-(dimethylamino)phenyl diester 20e.

Saponification of the crude diester with LiOH followed

by workup gave the desired diacid 21e. A small amount

of a methylated indole material, observed in the crude

diester product, was removed by recrystallization of the

diacid from EtOAc. The m-(methylamino)phenyl ana-

logue 21f was prepared by formylation of the m-

aminophenyl diester 18 with HCO₂Et followed by

selective reduction with H₃B‚SMe₃. Saponification of the

resulting diester 20f with LiOH, followed by workup,

gave the desired diacid.

Knoevenagel Reaction. Utilization of the enol ether

method was not applicable to the synthesis of analogues

with substituted aromatics containing electron-donating

groups. Therefore, the classical Knoevenagel condensa-

tion reaction²⁴was explored as an alternative method

to prepare analogues of this type. Attempts to condense

aldehyde 6 with prospective arylacetic esters failed;

however, when the appropriate arylacetonitrile 22a-d

was used, the corresponding arylpropenenitriles 23a-d

were generated, providing an easy entry into a limited

series of analogues 24a-d (Scheme 7).

23b-d and the p-aminophenyl analogue 25b were the

corresponding nitriles successfully hydrolyzed to the

desired diacids 24b-d and 26b.

Mukaiyama Reaction. Although the condensation

of N-tosyl aldehyde 7 with phosphonate 4 gave vinyl

bromides 8 in good yield under Horner-Emmons condi-

tions, the condensation of 7 with ethyl (diethylphospho-

no)phenylacetate was unsuccessful. Therefore, to pre-

pare analogues of 1 which incorporate an arylacetic

ester containing a relatively inactivated methylene

group, an alternative coupling method using the Lewis

acid-catalyzed aldol reaction of aldehyde 6 with various

ketene silyl acetals such as 28, also known as the

Mukaiyama reaction²⁶(Scheme 8), was explored. The

success of the Mukaiyama reaction would rely on the

transfer of the silyl group to the oxygen of the aldol to

drive the reaction to completion.

Condensation of 1-methoxy-1-(trimethylsiloxy)-2-phen-

ylethylene (28a)²⁷with aldehyde 6 in the presence of

(Ph₃P-O-Ph₃P)²⁺‚2TfO^{- 28}at -78 to 0 °C gave 54% of

erythro/threo aldols 30a and 12% of the corresponding

TMS ethers 29a.

A detailed account of the synthesis of analogues of 1

via this route has been reported.²⁵A limitation of this

method is that in most cases the hydrolysis of the

arylpropenenitriles proceeds only to the respective

amides, which will not hydrolyze further, even under

more rigorous conditions, to the desired carboxylic acids.

Only in the case of the substituted pyridine analogues

While 1-methoxy-1-(trimethylsiloxy)-2-(2-pyridyl)eth-

ylene (28b) gave only the TMS ether 29b, the ketene

silyl acetal of methyl (m-phenoxyphenyl)acetate (28d)

gave mostly the dehydrated product 31d directly.

The aldol products proved to be versatile intermedi-

ates. Treatment with p-TsOH in refluxing toluene

results in dehydration to the E/Z unsaturated esters in

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1001

Scheme 8^a

^aReactions and conditions: (a) TMSOTf, Et₃N; (b) Ph₃P-O-PPh₃²⁺‚2TfO^-; (c) pTsOH or Tf₂O; (d) LiOH, THF, H₂O; (e) KHSO₄.

Structures: a; R₁) Me, R₂) Ph; b, R₁) Et, R₂) 2-pyridyl; c, R₁) Me, R₂) 3-methoxyphenyl; d, R₁) Me, R₂) 3-phenoxyphenyl; e,

R₁) Me, R₂) Bn; f, R₁) Me, R₂) phenethyl; g, R₁) Me, R₂) cyclohexyl.

high yield. However, treatment of the silylated 2-pyridyl

aldols 29b with TFA at 25-45 °C gave exclusively ethyl

2-pyridylacetate and aldehyde 6 from a retro-aldol

condensation. To avoid the retro-aldol and obtain the

unsaturated ester 31b directly, the silylated 2-pyridyl

aldol 29b was treated with Tf₂O at 0-25 °C in the

absence of added base. The triflate, generated in situ,

is apparently unstable and undergoes base-catalyzed

elimination to the olefin. Running the reaction in the

presence of proton sponge [1,8-bis(dimethylamino)-

naphthalene] gave a complex mixture of products. The

retro-aldol was not completely suppressed, but a ∼4.5:1

ratio of unsaturated esters 31b/retro-aldol products

were obtained. Saponification of ester 31b provided pure

E diacid 32b; however, upon drying at 125 °C, the diacid

isomerized to a 6:1 E/Z mixture. The desired E isomers

of diacids 32c,d were obtained exclusively after crystal-

lization upon saponification of the intermediate E/Z

diesters 31c,d.

The preparation of analogues with the proximal

phenyl ring replaced by benzyl, phenethyl, and cyclo-

hexyl residues was also possible using the Mukaiyama

route. Treatment of intermediates 29e,g with p-TsOH

in refluxing toluene resulted in dehydration to the novel

unsaturated esters 31e and 31g in high yield. When (1-

methoxy-3-phenyl-1-propenyloxy)trimethylsilane (28e)

was condensed with aldehyde 6 under similar conditions

and the mixture allowed to warm to room temperature

with stirring over 16 h, neither the aldol nor the TMS

ether 29e/30e was observed due to in situ dehydration,

possibly induced by excess Lewis acid during the

extended reaction time. As a result, the desired unsat-

urated diester 31e was isolated directly in moderate

yield and was found to consist only of the Z isomer.

Assignment of olefin geometry in 31e is based on NOE

effects and vicinal proton-carbon coupling constants

involving the olefinic proton, specifically a strong NOE

interaction (ca. 20%) with the benzylic methylene

protons, a large anti-coupling (ca. 13 Hz) to the ester

carbonyl carbon, and a smaller syn-coupling (ca. 6.5 Hz)

to the benzylic methylene carbon. Base hydrolysis of the

unsaturated benzyl diester 31e gave benzyl diacid

analogue 32e exclusively as the Z isomer, also deter-

mined by NOE studies. Likewise, in the analogous

preparation of cyclohexyl diester 31g and subsequent

hydrolysis to diacid 32g, only the respective Z isomers

were isolated. In contrast, the adduct obtained from

condensation of (1-methoxy-4-phenyl-1-butenyloxy)tri-

methylsilane (28f) with 6 under the same conditions

gave analytical data consistent with structure 31f (ca.

1:1 diastereomeric mixture), resulting from cyclization

via intramolecular Friedel-Crafts alkylation.

Synthesis of the Saturated Analogue, S-3-(2-Car-

boxy-2-phenylethyl)-4,6-dichloro-1H-indole-2-

carboxylic Acid (36)

The CoMFA analysis showed that a low-energy con-

former of the S-enantiomer of 3-(2-carboxy-2-phenyl-

ethyl)-4,6-dichloro-1H-indole-2-carboxylic acid (36) over-

lapped very well with the lowest energy conformer of

receptor-docked 1 (Figure 4). To test the validity of these

calculations, compound 36 was synthesized, in its ra-

cemic form, along with several related alkyl diacid

analogues and evaluated for NMDA receptor antagonist

activity (Scheme 9).

The most obvious synthetic route to dihydro analogue

36 is the direct hydrogenation of 1. Unfortunately,

hydrogenation of 1 using H₂at 60 psi with 10% Pd/C

resulted in dechlorination, making it necessary to

develop an alternate synthetic route. This was ac-

complished starting with N-tosyl aldehyde 7. Protection

of the indole nitrogen was necessary prior to reduction

of the aldehyde to the alcohol. The aldehyde was

reduced with NaBH₄to give the alcohol 33, which was

then treated with SOCl₂in refluxing toluene to afford

the corresponding chloride 34. Chloride 34 was con-

verted with NaI to the iodide, which was reacted in situ

with the anion of ethyl phenylacetate. The resulting

dihydro diester, 35, was hydrolyzed to afford the dihydro

diacid 36 in moderate to good yield (Scheme 10).

The Mukaiyama aldol route was also explored for the

preparation of compound 36. Treatment of the aldol

1002 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

that have been evaluated through all the tests, including

the stroke model, did not permit correlation of stroke

results with any single test result, nor is it currently

known what combination of in vivo and in vitro tests

best correlate with activity in the stroke model. Thus,

it would appear that good to excellent activity in most

of the critical path tests and models is needed to identify

those compounds which will exhibit significant neuro-

protective results in the stroke model (see, for example,

38 vs 19).

In Vitro Activity. The in vitro pharmacology de-

scribed below reflects the critical discovery path used

for evaluating the glycine site antagonists synthesized.

The two main criteria for compound selection were the

binding of the inhibitor to the NMDA receptor complex

and functional assays to assess in vitro “efficacy”. These

two areas are outlined to demonstrate the primary

activities measured.

The binding assays measured the affinity of the

inhibitor and the benchmark/reference compounds for

the glycine site on the NMDA receptor. The functional

assays demonstrated NMDA antagonism in vitro and

protection of the neuronal cell cultures from anoxia.

Figure 4. Overlay of compound 1, with compound 36 in which

the unsaturation has been removed.

Scheme 9

Compound 19 was found to be a potent antagonist

(K_i) 24 nM) of [³H]glycine binding to its recognition

site on the NMDA receptor complex of rat brain. Activity

resides exclusively with the E isomer of 19. The Z

isomer has been prepared and evaluated in key critical

path assays as well as in in vitro assays predictive of

side effect liability. The K_ivalue for the Z isomer in the

[³H]glycine binding assay was 553 nM. Values for 19

and reference compounds (Figure 1) are shown in Table

2.

Scheme 10^a

Several biochemical methods confirmed 19 as a

functional antagonist at the glycine site. For example,

in a cerebellar slice preparation, NMDA elicits a con-

centration-dependent increase in the content of the

second messenger substance cGMP. Compound 19

completely inhibited this response with an IC₅₀of 364

nM. Values for 19 relative to other NMDA antagonists

are summarized Table 2.

^aReactions and conditions: (a) NaBH₄; (b) SOCl₂; (c) NaI; (d)

PhCH₂CO₂Et, NaH; (e) KOH; (f) HCl.

Scheme 11^a

Antagonism of the NMDA response was noncompeti-

tive in nature. Thus, in the presence of a fully effective

concentration of 19, increasing the concentration of

NMDA could not restore the functional response. Inhi-

bition of the cGMP functional response was due to an

action of 19 at the glycine recognition site in that it was

readily reversible by addition of an excess amount of

the glycine-mimetic D-serine. Compound 19 (1 µM)

completely inhibited the NMDA response. Addition of

D-serine resulted in a concentration-dependent restora-

tion of the NMDA response. With a concentration of

D-serine equal to 14.5 µM 50% restoration was achieved.

These results indicate that 19 is a noncompetitive

inhibitor of NMDA, and therefore the potency of 19

would be unaffected by ischemia-induced elevations of

extracellular glutamate.

In Vivo Activity. Following the characterization of

the in vitro activity, several critical issues were ad-

dressed using a series of in vivo models. In vivo evidence

of NMDA antagonism was obtained in mice by monitor-

ing effects of intravenously administered compound 19

on harmaline-induced elevations of cerebellar cGMP

content. Harmaline is known to activate the climbing

^aReactions and conditions: (a)TFA, Et₃SiH; (b) LiOH, THF,

H₂O; (c) KHSO₄.

products, 29a/30a, with TFA/Et₃SiH at room tempera-

ture gave only dihydro ester 37 without any trace of

unsaturated ester 31a resulting from dehydration

(Scheme 11). SnCl₄/Et₃SiH was also effective, but less

convenient, in reducing the aldol products. Saponifica-

tion of dihydro ester 37, followed by crystallization from

cyclohexane/EtOAc, gave dihydro diacid 36 in good

yield.

Pharmacology

Compounds were advanced from the binding assay

to the in vivo anticonvulsant tests, the in vitro (inhibi-

tion of cGMP accumulation stimulated by NMDA in rat

cerebellar slices) and in vivo (inhibition of harmaline-

induced cGMP elevation in the cerebellum) biochemical

functional assays in parallel. The number of compounds

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1003

Table 2. Summary of Activities for NMDA Antagonists in Tests Used for Compound Selection

in vitro tests, IC₅₀,^anM in vivo functional tests, ED₅₀,^amg/kg

binding

affinity,^b

nM

functional

potency

harmaline

increase in cGMP,

mouse, iv

audiogenic

seizures, DBA/2J,^d

mouse, ip

quinolinic acid

seizures,^d

mouse, iv

maximal

electroshock,

rat, iv

compd

(cGMP)^c

1

10.9

24.3

9.6

462

114

20

282

364

87

2560

840

183

262

31

59

15

12.2

10.8

18.6

16.9

8.5

37.0

14.9

29.5

55.0

>64

14.7

>64

5.2

29.0

30.6

19

11h

21a

21c

38

39

40

35

54.1

>64

3.5

nd

<0.05

>128

46.3

3.7

15.2

13.2

>25

1

>32

3.5

^aNumbers reported are the mean of two or more experiments unless indicated. Standard errors are (10-30% of the mean. ^bIC50

values represent the concentration of the compound required to reduce the binding of the tritiated ligand by 50% for rat cortical and

hippocampal membrane strychnine-insensitive gycine binding sites. ^cInhibition of glutmatate stimulated accumulation of cyclic GMP in

neonatal rat cerebral slices. ^dDBA and QAS models were used to assess CNS penetration. The ED₅₀is defined as that dose protecting

50% of the mice.

fiber afferent pathway to the cerebellum that utilizes

glutamate as its neurotransmitter. This provides a

means for in situ activation of NMDA receptors and thus

is an in vivo counterpart of the cGMP assay performed

with cerebellar slices. Compound 19 inhibited the har-

maline response with an ED₅₀of 15 mg/kg. Comparative

data are provided for compound 11h and reference

NMDA antagonists in Table 2.

To demonstrate brain penetration, the audiogenic

seizure-prone mouse (DBA/2J) was treated intraperi-

toneally with compound 19 followed by presentation of

a loud sound stimulus (110 dB, 11 kHz, 25 s) to initiate

the onset of convulsions. Compound 19 provided protec-

tion in this model with an ED₅₀) 10.8 mg/kg. This

model is known to be sensitive to NMDA antagonists

and has been used as an initial test by a number of

pharmaceutical companies developing these compounds.

Compound 19, when given iv, antagonized the sei-

zures induced by the intracerebroventricular (icv) ad-

ministration of quinolinic acid, an NMDA agonist, with

an ED₅₀) 14.9 mg/kg. This value is consistent with the

dose range for biochemical activity of 19 as an antago-

nist at the NMDA receptor (i.e. harmaline-elevated

cGMP). A time course analysis found that 20.7 mg/kg

of compound 19 protected 60% of the mice at 5 min after

injection, 20% at either 1 or 2 h, and 0% at 4 h. Thus,

compound 19 had a peak effect at 5 min with duration

of approximately 1 h after injection in mice.

In the rat, compound 19 antagonized maximal elec-

troshock-induced seizures with an ED₅₀) 30.6 mg/kg

iv. A time course analysis found that 54.7 mg/kg of 19

protected 70% of the rats at 5 min and 10% at 1 h after

iv injection. Thus, the duration of effect in rats is similar

to that seen in the mouse vide supra. These tests

enabled the selection of compound 19 for evaluation in

the stroke model and provided guidance for the condi-

tions used in the animal stroke experiments

Summary of in Vitro and in Vivo Data on

Compound 19 Used To Make Compound Selection.

Table 2 summarizes the key data used to choose

compound 19 for advancement. Key competitors and

compounds of interest are 38 (CoCensys/Ciba), 39

(Glaxo Wellcome), and 40 (Merck), which are currently

in late-phase clinical trials for the treatment of stroke.

In mouse behavioral tests or in mouse-derived tissues,

38 was as potent or 2-3-fold more potent than 19.

However, in the rat, 19 was more potent in the maximal

electroshock seizure test. Thus, both compounds 19 and

38 show a similar broad pattern of efficacy in seizure

models. In contrast, 39 is a potent receptor antagonist

in in vitro tests; however, this compound exhibited no

activity following iv or ip administration. Activity was

detected in these tests following direct administration

into the brain (icv). These data are consistent with a

limited penetration of 39 into the intact nervous system.

Further evaluation of these competing compounds will

focus on their comparative activity in the stroke models.

The selection of compound 19 for evaluation in a

therapeutic model of stroke was derived by a logical

demonstration of glycine/NMDA receptor antagonism,

from initial binding studies, and through functional

assays, to in vivo activities.

Activity in the Stroke Model. Stroke is defined as

a focal neurological deficit that is sudden in onset and

stable over time. It is caused by an interruption in blood

flow through a cerebral vessel and may be thrombotic,

embolic, or hemorrhagic in origin. The resulting neu-

rological damage is related to the density of ischemia

(i.e., residual blood flow in the ischemic zone) and its

duration. Because of the inherent variability of the

human condition, two different animal models of focal

cerebral ischemia were chosen to represent the disease,

dMCAo (distal middle cerebral artery occlusion) and

pMCAo (proximal middle cerebral artery occlusion).

These models, and their key features are outlined in

Table 3.

The models were chosen to represent the human

disease by providing a range of ischemic insult severity.

Thus, the dMCAo model, using permanent ischemia in

a strain with little collateral circulation, gives a severe

ischemia with only a small fraction of salvageable tissue.

In contrast, the pMCAo, using a 3 h period of ischemia,

followed by reperfusion in a strain with good collateral

circulation, provides a model of moderately severe

ischemia with a high fraction of salvageable tissue.

Consistent with these features, postischemic adminis-

tration of compound 19 reduced infarct volume by 28%

in the dMCAo and 77% in the pMCAo models. These

data indicate that compound 19 shows a clear pattern

of neuroprotection in animal models believed to be

predictive of effectiveness in human stroke.

Compound 19 reduced infarct volume in both perma-

nent and ischemia-reperfusion models (Table 4). In

either model, similar reductions in infarct volume were

1004 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

Table 3. Summary of the Properties of the Stroke Models Used To Evaluate Antagonist 19

feature

dMCAo^a

pMCAo^b

animal/strain

location arterial occlusion

type of “block”

rat/spontaneously hypertensive (SH rat) rat/Wistar

distal segment of MCA

electrocautery

proximal segment of MCA

monofilament

duration of ischemia

density of ischemia

severity of ischemia

typical lesion volume

brain areas affected

permanent

temporary (3 h)

moderate, good collateral flow + reperfusion

high, little collateral flow

high

moderate

160 mm³, 11% of hemisphere volume

cerebral cortex

190 mm³, 21% of hemisphere volume

cortex and striatum

50%

typical infarct reduction by neuroprotectants 20%

^adMCAo ) distal middle cerebral artery occlusion. ^bpMCAo ) proximal middle cerebral artery occlusion.

Table 4. Summary of Results with Compound 19

dosing regimen

duration (h)

neuroprotection (% infarct reduction)

bolus (mg/kg)

infusion (mg/kg/h)

total dose (mg/kg)

initiation (min)

dMCAo (%)

pMCAo (%)

70

35

17.5

8.8

4.5

2.3

35

18

35

70

35

17.5

8.8

4.5

2.3

35

18

25

6

4

490

245

123

62

31.5

16

245

126

135

-15

30

32^a

nd

13

0

73^a

77^a

68^a

49^a

48^a

40^a

12

30

57^a

76^a

71^a

50^a

41^a

9

30

120

180

240

360

480

-15

30

44^a

58^a

59^a

13

nd

28a

nd

^ap < 0.05.

Table 5. Summary of Results with Competitors

dosing regimen

compd bolus (mg/ kg) infusion (mg/kg/h) duration (h) total dose (mg/kg) initiation (min)

neuroprotection (% infarct reduction)

dMCAo (%)

pMCAo (%)

38

39

10

30

7

6

52

30

-15

30

18^a

33^a

24^a

nd

0

50^a

nd

30

nd

180

-15

180

28

-25^b

32

nd

^ap < 0.05. ^bIndicates increased infarct volume with 39 treatment (not significant).

observed whether the drug was given prior to or

following the ischemic insult. Compound 19 was also

compared to benchmark NMDA antagonists in the two

models (Table 5). In every case, the extent of efficacy

observed with compound 19 was greater than that

produced by the reference compounds. While the dose

for the benchmarks was fully efficacious on the basis of

published data, the greater efficacy seen with compound

19 may reflect selection of a suboptimal dose of the

reference compounds. Alternatively, the results could

indicate a greater degree of efficacy for compound 19

in the therapeutic models.

is one of the most potent compounds tested to date in

the binding assay but is not as active overall as 19. All

of the compounds prepared showed good to excellent

binding activity (,10 µM, the minimal acceptable

binding affinity) against [³H]glycine on the NMDA

receptor and several compounds, 21a and 21c, show

potent activity in the DBA/2J model. Compound 19 has

a T_1/2> 1 h following iv administration.

As predicted by the CoMFA analysis, significant

increase in binding affinity and potency were obtained

by the incorporation of an aromatic group with an

electron-withdrawing subtituent or heterocycle at the

2-position of the 3-propenyl moiety of lead structure 1.

Although CoMFA analysis indicated that racemic com-

pound 36 and related dihydro analogues may have

significantly improved binding activity, upon biological

evaluation they were not any more potent or have better

in vivo efficacy than compound 1 to warrant further

investigation of this series.

Conclusions

Analogue 19 is the most fully characterized of the new

compounds, being generally more potent than 1 and

showing efficacy in the stroke model which differenti-

ates this compound from 1. The 3-thienyl analogue 11h

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1005

as milligrams per milliliter. The absorptivity (A) and molar

absorptivity (ꢀ) are calculated at appropriate wavelengths.

Experimental Section

General Methods. Except where noted otherwise, reagents

and starting materials were obtained from common com-

mercial sources and used as received. Anhydrous solvents were

purchased from Aldrich Chemical Co., Inc. in Sure/Seal

bottles. Other reaction solvents and chromatographic, recrys-

tallization, and workup solvents were spectroscopic grade and

used as received. When required, reactions were run under

an atmosphere of dry nitrogen or argon in oven-dried flasks.

The phrase “concentrated in vacuo” indicates rotary evapo-

ration on a Buchi apparatus at 25-50 °C and 15-20 Torr

(water aspirator) or 30 mmHg (KNF Neuberger Model UN

726.12FTP diaphragm pump), unless stated otherwise. Room

temperature is abbreviated as “rt”. Vacuum drying was

performed at <10 Torr at the temperatures noted. Melting

points were determined on a Thomas-Hoover Uni-melt capil-

lary melting point apparatus. Melting points and boiling points

are reported uncorrected.

Abbreviations: Et₃N, triethylamine; p-TsOH, para-toluene-

sulfonic acid; TFA, trifluoroacetic acid; TFAA, trifluoroacetic

anhydride; TMSOTf, trimethylsilyl triflate; TBDMS, tert-

butyldimethylsilyl; EDC, 1-(3-(dimethylamino)propyl)-3-eth-

ylcarbodiimide hydrochloride; HOBT, 1-hydroxybenzotriazole

hydrate; Tf₂O, trifluoromethanesulfonic anhydride; LiHMDS,

lithium hexamethyldisilazide.

Dibromo(diethoxyphosphoryl)acetic Acid tert-Butyl

Ester,¹⁵3. Fresh NaOBr was prepared via dropwise addition

of Br₂(40.0 g, 12.8 mL, 0.25 mol) to a cooled (ice bath), stirred

solution of NaOH (20.0 g, 0.5 mol) in H₂O (60 mL). After 25

min tert-butyldiethylphosphonoacetate (13.5 g, 12.6 mL, 0.053

mol) was added dropwise over 5 min at ice-bath temperature.

When the addition was complete, the mixture was extracted

with CH₂Cl₂(2 × 70 mL) and the combined extracts were

washed with H₂O (1 × 100 mL), dried (MgSO₄), and concen-

trated to give 3 as a colorless oil (20.9 g, 95%), R_f) 0.25, 50%

Et₂O/hexane. ¹H NMR (300 MHz, CDCl₃): δ 4.38 (m, 4H), 1.54

(s, 9H, t-Bu), 1.40 (overlapping t, 6H, J ) 7.1 Hz).

Thin-layer chromatography (TLC) was performed on glass-

backed, silica gel 60F-254 plates (EM) coated to a thickness

of 0.25 mm. The plates were eluted with solvent systems (v/v)

as described and visualized by iodine vapor, UV light, or a

staining reagent such as KMnO₄solution.²⁹Gas chromatog-

raphy (GC) was performed on a Hewlett-Packard 5890 Series

II gas chromatograph equipped with a Hewlett-Packard 3392A

integrator. Separations were carried out on a 15 m × 0.32 mm

i.d. fused silica capillary column (DB-5, 0.25 mm film) from J

& W Scientific. Reversed-phase high-performance liquid chro-

matographic (HPLC) analyses were performed on a Waters

system composed of a Model 600E pump, a Model 486 detector,

and a Model 746 data module. The column (250 mm × 4.6 mm

i.d.), from Eka Nobel (Nobel Industries, Sweden), was packed

with C₈Kromasil (spherical 13 µm particles, KR100-13-C8).

The mobile phase consisted of CH₃CN buffer mixtures, the

buffer being made up of 900 parts distilled water, 100 parts

CH₃CN, and 1 part TFA. Preparative TLC was performed on

glass-backed silica gel 60F-254 plates (EM) coated to a

thickness of 2 mm. Preparative radial TLC was accomplished

using a Chromatotron (Harris Industries) system. Flash

chromatography was carried out using EM Science silica gel

60 (40-63 mm) according to the literature procedure³⁰using

the solvent systems as described.

Infrared (IR) spectra were recorded on a Mattson Galaxy

Series 5020 infrared spectrophotometer with samples prepared

as indicated and are reported in wavenumbers (cm^-1). ¹H NMR

spectra were recorded on Varian Gemini-300, Unity-300,

Unity-400, or UnityPlus-500 spectrometers with chemical

shifts (δ) reported in parts per million (ppm) relative to

internal tetramethylsilane (0.0 ppm) or residual protonated

solvent (chloroform, 7.26 ppm; dimethyl sulfoxide (DMSO),

2.50 ppm) as a reference. Signals were designated as s

(singlet), d (doublet), t (triplet), q (quartet), p (pentuplet), m

(multiplet), and br (broad), etc. Coupling constants (J) are

reported in hertz. First-order analyses of spectra were at-

tempted when possible; consequently, chemical shifts and

coupling constants for multiplets may only be approximate.

¹³C NMR spectra were recorded on the Varian Gemini or Unity

instruments (75 or 100 MHz), with chemical shifts (δ) reported

in ppm relative to CDCl₃(77.0 ppm) unless stated otherwise.

Mass spectra (MS) were obtained on a Finnigan MAT Model

TSQ 700 mass spectrometer system by chemical ionization at

120 eV using methane (CI, 120 eV) unless otherwise indicated

as for example with electron impact, MS (EI, 70 eV). The

relative peak height in percent for the base peak and the

molecular ion designated as M are given in parentheses. High-

resolution mass spectrometric analysis (exact mass spectra)

was performed in the FAB mode at a mass resolution of 10 000

(10% valley definition) using a VG-Fisons Autospec mass

spectrometer. Exact mass values were determined for the

protonated molecular ions (M⁺+ 1). Ultraviolet (UV) spectra

were recorded on a Perkin-Elmer Lambda 4C spectrophoto-

meter and reported in nanometers (nm). The sample was

prepared in the solvent indicated at a concentration expressed

Bromo(diethoxyphosphoryl)acetic Acid tert-Butyl Es-

ter,¹⁵4. To a stirred, cooled (ice bath) solution of the dibro-

mophosphonoacetate 3 (20.8 g, 0.05 mol) in t-BuOH (100 mL)

was added, in portions, a solution of SnCl₂‚2H₂O (11.0 g, 0.049

mol) in H₂O (100 mL). The cloudy white reaction mixture was

stirred at ice-bath temperature for 15 min and then extracted

with CH₂Cl₂(3 × 100 mL). The combined extracts were washed

with H₂O (2 × 100 mL), dried (MgSO₄), and concentrated in

vacuo to give 4 as a pale yellow oil (15.2 g, 91%), R_f) 0.24,

1

50% Et₂O/hexane. H NMR (300 MHz, CDCl₃): δ 4.24-4.31

(m, 5H, 2 CH₂, overlapping d, J ) 15.2 Hz, CHBr), 1.51 (s,

9H, t-Bu), 1.35-1.40 (overlapping t, 6H, J ) 7.0 Hz).

4,6-Dichloro-3-formyl-1H-indole-2-carboxylic Acid Eth-

yl Ester, 6. To a stirred suspension of 4,6-dichloro-1H-indole-

2-carboxylic acid ethyl ester (5)¹⁴(20.0 g, 0.077 mol) in

anhydrous CH₂Cl₂(100 mL) and DMF (9.0 mL, 0.117 mol) at

rt was added POCl₃(18.0 g, 11.0 mL, 0.117 mol). The mixture

was then heated at reflux for 2.5 h, after which time it was

allowed to cool to rt and the precipitate filtered and washed

sparingly with H₂O. The crude solid was then treated with 1

M NaOAc/H₂O (300 mL) and stirred at rt for 1 h, filtered,

washed with H₂O, and placed under high vacuum (0.5 Torr)

for 16 h to afford 6 (14.3 g, 64.7%) as an off-white solid: mp

216-217 °C. ¹H NMR (300 MHz, CDCl₃): δ 10.80 (s, 1H), 9.40

(br s, 1H, NH), 7.39 (s, 1H, Ar), 7.35 (s, 1H, Ar), 4.52 (q, 2H,

J ) 7.2 Hz), 1.47 (t, 3H, J ) 7.1 Hz).

4,6-Dichloro-3-formyl-1-(toluene-4-sulfonyl)-1H-indole-

2-carboxylic Acid Ethyl Ester, 7. To a stirred suspension

of 6 (14.3 g, 0.057 mol), K₂CO₃(32.1 g, 0.23 mol), and DMF

(200 mL) at rt was added p-TsCl (10.9 g, 0.057 moL) in portions

(a slight exotherm was observed). After stirring for 30 min at

rt, the mixture was diluted with H₂O (100 mL) and the solid

precipitate filtered and washed with additional H₂O. This

crude solid was then triturated with Et₂O (100 mL), filtered,

and dried under vacuum (0.5 Torr) to yield 7 (18.5 g, 84%) as

1

a white powder: mp 189-191 °C (dec). H NMR (300 MHz,

CDCl₃): δ 10.71 (s, 1H), 8.00 (m, 3H), 7.36 (m, 3H), 4.61 (q,

2H, J ) 7.2 Hz), 2.41 (s, 3H), 1.49 (t, 3H, J ) 7.1 Hz).

(Z)-3-(2-Bromo-2-(tert-butoxycarbonyl)vinyl)-4,6-dichlo-

ro-1-(toluene-4-sulfonyl)-1H-indole-2-carboxylic Acid Eth-

yl ester, 8. To a stirred solution of 4 (15.2 g, 0.046 mol) in

dry THF (200 mL) at -78 °C under N₂was added LiHMDS (1

M in THF, 45.9 mL, 0.046 mol) dropwise over 10 min. This

was followed by the portionwise addition of aldehyde 7 (13.0

g, 0.029 mol) as a suspension in THF (50 mL). The mixture

was stirred at -78 °C for 30 min and then allowed to warm to

rt and stirred for an additional 1.5 h. A clear dark mixture

resulted that was quenched with H₂O (100 mL) and concen-

trated to remove the THF. The residual oil was then diluted

with CH₂Cl₂(100 mL), washed with H₂O (1 × 50 mL) followed

by brine, dried (MgSO₄), and concentrated in vacuo to give an

amber oil which solidified upon standing. Recrystallization

from Et₂O afforded the desired Z isomer of 8 (9.6 g, 53%) as a

1006 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

1

white solid: mp 131-132 °C. H NMR (300 MHz, CDCl₃): δ

Hz), 7.7-7.8 (m, 3 H), 7.4-7.5 (m, 5 H), 7.29 (d, 1 H, J ) 1.7

Hz), 7.1-7.2 (m, 3 H), 7.00 (dd, 1 H, J ) 7.0, 1.1 Hz), 3.63

(bm, 1 H), 2.79 (bm, 1 H), 2.35 (s, 3 H), 1.40 (s, 9 H), 0.79 (t,

3 H, J ) 7.2 Hz)scyclohexane, aromatic impurity present.

8.21 (s, 1H), 7.95 (m, 3H), 7.30 (m, 3H), 4.42 (q, 2H, J ) 7.2

Hz), 2.41 (s, 3H), 1.56 (s, 9H, tBu), 1.36 (t, 3H, J ) 7.1 Hz).

Anal. C₂₅H₂₄BrCl₂SNO₆: C, H, N. The E isomer was obtained

by slow fractional crystallization of the mother liquor from

EtOAc/cyclohexane to afford vinyl bromide 8E as large color-

less rods: mp 117-119.5 °C. ¹H NMR (CDCl₃): δ 7.99 (d, 1 H,

J ) 1.7 Hz), 7.96 (d, 2 H, J ) 8.7 Hz), 7.50 (s, 1 H), 7.33 (d, 2

H, J ) 8.7 Hz), 7.27 (d, 1 H, J ) 1.7 Hz), 4.42 (q, 2 H, J ) 7.2

Hz), 2.41 (s, 3 H), 1.39 (t, 3 H, J ) 7.2 Hz), 1.00 (s, 9 H)strace

Et₂O present.

General Procedure I for Suzuki-Miyaura Coupling

Reaction.^13cMethod A. To a stirred suspension of the vinyl

bromide 8Z (1.08 g, 1.75 mmol), arylboronic acid,¹⁶(2.63 mmol)

and K₂CO₃(0.483 g, 3.5 mmol) in toluene (20 mL) under N₂

was added Pd(PPh₃)₄(0.20 g, 10 mol %), and the mixture was

heated to 90 °C for 1 h. After 1 h the mixture was concentrated

and then diluted with CH₂Cl₂, washed successively with H₂O,

saturated NaHCO₃, saturated tartaric acid, and then brine,

dried (MgSO₄), and concentrated to give a yellow-orange

semisolid which was immediately flash-chromatographed us-

ing 25% Et₂O/hexane on SiO₂to provide the Suzuki coupled

product.

Method B. Tris(dibenzylideneacetone)dipalladium(0) [Pd₂-

(dba)₃] (412 mg, 0.450 mmol) and tri-2-furylphosphine (837 mg,

3.60 mmol) were added to dry THF (60 mL) with stirring under

N₂. After 5 min, vinyl bromide 8Z (1.856 g, 3.01 mmol), the

electron-rich arylboronic acid (9.20 mmol), and powdered K₂-

CO₃(1.27 g, 9.20 mmol) were added to the yellow-green

suspension. The reaction mixture was stirred at 55-60 °C for

6-9 days, then cooled to rt, and diluted with cyclohexane (120

mL). The mixture was poured onto a 100 mm × 70 mm SiO₂

column (prepacked with 3:1 cyclohexane/EtOAc) and eluted

with 3:1 cyclohexane/EtOAc. The eluant was concentrated in

vacuo and the residue purified by flash chromatography (4:1

cyclohexane/EtOAc) to give the Suzuki coupled product.

(E)-3-(2-(tert-Butoxycarbonyl)-2-phenylvinyl)-4,6-dichlo-

ro-1-(toluene-4-sulfonyl )-1H-indole-2-carboxylic Acid

Ethyl Ester, 9a. Phenylboronic acid (0.32 g, 2.63 mmol) was

coupled to 8 using general procedure I, method A, to afford

9a (388 mg, 36.3%) as a white amorphous solid. ¹H NMR (300

MHz, CDCl₃): δ 7.90 (s, 1H), 7.81 (s, 1H), 7.62 (m, 2H), 7.02-

7.26 (m, 8H), 4.12 (q, 2H, J ) 7.1 Hz), 2.39 (s, 3H), 1.53 (s,

9H, tBu), 1.24 (t, 3H, J ) 7.4 Hz). Anal. C₃₁H₂₉Cl₂SNO₆: C,

H, N.

(E)-3-(2-Carboxy-2-phenylvinyl)-4,6-dichloro-1-(toluene-

4-sulfonyl)-1H-indole-2-carboxylic Acid Ethyl Ester, 10a.

tert-Butyl ester (9a; 388.0 mg, 0.63 mmol) was hydrolyzed with

TFA (1.5 mL) in CH₂Cl₂(20 mL) with stirring at rt for 15 min.

The mixture was diluted with additional CH₂Cl₂, washed with

H₂O, dried (MgSO₄), and concentrated to give 10a (345 mg,

98%) as a white amorphous solid. ¹H NMR (300 MHz,

CDCl₃): δ 9.48 (br s, 1H), 8.09 (s, 1H), 7.91 (s, 1H), 7.62 (d,

2H, J ) 8.4 Hz), 7.07-7.27 (m, 8H), 4.14 (q, 2H, J ) 7.2 Hz),

2.38 (s, 3H), 1.22 (t, 3H, J ) 7.1 Hz).

(E)-3-(2-Carboxy-2-phenylvinyl)-4,6-dichloro-1H-indole-

2-carboxylic Acid, 11a from Diester 9a via Acid 10a. Vinyl

bromide 8Z (928 mg, 1.50 mmol) was coupled with phenylbo-

ronic acid (445 mg, 2.59 mmol) using general procedure I,

method A, to afford diester 9a. Removal of the tert-butyl ester

was accomplished by stirring 9a in TFA (7 mL) for 1 h. Workup

followed by flash chromatography (95:4:1 CH₂Cl₂/MeOH/

HOAc) provided acid 10a, which was hydrolyzed using LiOH‚

H₂O (122 mg, 2.91 mmol, 3.4 equiv) in 9:1 THF/H₂O (20 mL)

at reflux for 7.5 h followed by recrystallization from cyclohex-

ane/EtOAc to provide 11a as an ivory powder. ¹H NMR

(DMSO-d₆): δ 12.95 (bs, 2 H), 12.14 (s, 1 H), 8.10 (s, 1 H),

7.32 (s, 1 H), 7.15 (s, 1H), 7.08 (m, 3 H), 6.97 (m, 2 H).

Removal of the tert-butyl ester was accomplished by stirring

9b in TFA (7 mL) for 1 h. Workup followed by flash chroma-

tography (95:4:1 CH₂Cl₂/MeOH/HOAc) provided acid 10b (527

1

mg) as a pale yellow oil. H NMR (CDCl₃): δ 9.60 (bs, 1 H),

8.35 (s, 1 H), 7.85 (d, 1 H, J ) 1.7 Hz), 7.72-7.8 (m, 3 H), 7.44

(d, 2 H, J ) 8.4 Hz), 7.4 (m, 2 H), 7.29 (d, 1 H, J ) 1.7 Hz),

7.25 (m, 1 H), 7.14 (d, 2 H, J ) 8.0 Hz), 7.07 (dd, 1 H, J ) 7.1,

1.1 Hz), 3.7 (bm, 1 H), 2.95 (bm, 1 H), 2.34 (s, 3 H), 0.83 (t, 3

H, J ) 7.1 Hz)scyclohexane and EtOAc present. MS: m/z 610,

608 (M⁺+ 1), 564, 562, 438, 436, 157 (100).

Hydrolysis of acid 10b (527 mg, 0.866 mmol) using LiOH‚

H₂O (122 mg, 2.91 mmol, 3.4 equiv) in 9:1 THF/H₂O (20 mL)

at reflux for 7.5 h followed by recrystallization from cyclohex-

ane/EtOAc provided 11b as an ivory powder: mp 265 °C (dec).

IR (KBr): ν_max3252, 1696, 1248, 1235, 777 cm^{-1 1}H NMR

.

(DMSO-d₆): δ 12.8 (bs, 2 H), 11.96 (s, 1 H), 8.33 (s, 1 H), 7.83

(d, 1 H, J ) 8.3 Hz), 7.76 (d, 1 H, J ) 8.0 Hz), 7.67 (dd, 1 H,

J ) 7.3, 2.0 Hz), 7.2-7.40 (m, 4 H), 7.21 (d, 1 H, J ) 1.7 Hz),

7.15 (d, 1 H, J ) 1.7 Hz)sMeCN and EtOAc present. MS: m/z

438, 436 (M⁺+ 29 - H₂O), 427, 426 (M⁺+ 1), 425 (M⁺), 410,

408 (100), 382, 380. Anal. C₂₂H₁₃Cl₂NO₄: C, H, N.

3-[2-Carboxy-2-(2,4-dichlorophenyl)vinyl]-4,6-dichloro-

1H-indole-2-carboxylic Acid, 11c from Diester 9c via

Acid 10c. Vinyl bromide 8Z (821 mg, 1.33 mmol) was coupled

with (2,4-dichlorophenyl)boronic acid (382 mg, 2.00 mmol) via

general procedure I, method A. Flash chromatography (6:1

cyclohexane/EtOAc) gave diester 9c (500 mg) as an ivory

powder: mp 160.5-163.5 °C. IR (KBr): ν_max1723, 1370, 1277,

1196, 1181, 1159, 579 cm^-1. ¹H NMR (CDCl₃): δ 7.94 (d, 1 H,

J ) 1.7 Hz), 7.91 (s, 1 H), 7.62 (d, 2 H, J ) 8.4 Hz), 7.35 (d, 1

H, J ) 2.0 Hz), 7.28 (d, 1 H, J ) 1.4 Hz), 7.23 (d, 2 H, J ) 8.4

Hz), 6.86 (dd, 1 H, J ) 8.3, 1.9 Hz), 6.75 (d, 1 H, J ) 8.3 Hz),

4.26 (q, 2 H, J ) 7.1 Hz), 2.40 (s, 3 H), 1.49 (s, 9 H), 1.43 (s,

cyclohexane), 1.28 (t, 3 H, J ) 7.1 Hz). MS: m/z 712, 710 (M⁺

+ 29), 684, 682 (M⁺+ 1), 628, 530, 474, 456, 454, 157 (100).

Anal. C₃₁H₂₇Cl₄NO₆S: C, H, N.

Removal of the tert-butyl ester (vide supra) afforded acid

10c (389 mg, 47%) as fine ivory granules: mp 202-206 °C

(physical change ∼150 °C). IR (KBr): ν_max1730, 1697, 1373,

1273, 1196, 1181, 581 cm^-1. ¹H NMR (CDCl₃): δ 8.17 (s, 1 H),

7.95 (d, 1 H, J ) 1.7 Hz), 7.62 (d, 2 H, J ) 8.4 Hz), 7.36 (d, 1

H, J ) 2.0 Hz), 7.29 (d, 1 H, J ) 1.7 Hz), 7.24 (d, 2 H, J ) 8.3

Hz), 6.93 (dd, 1 H, J ) 8.3, 2.1 Hz), 6.83 (d, 1 H, J ) 8.3 Hz),

4.26 (q, 2 H, J ) 7.2 Hz), 2.40 (s, 3 H), 1.43 (s, cyclohexane),

1.28 (t, 3 H, J ) 7.2 Hz). MS: m/z 628, 626 (M⁺+ 1, with 4

Cl), 582, 456, 454, 157 (100). Anal. C₂₇H₁₉Cl₄NO₆S‚¹/₂C₆H₁₂:

C, H, N.

Hydrolysis of 10c (421 mg, 0.671 mmol) gave diacid 11c as

a fine ivory solid, which was dried under vacuum at 110 °C:

mp 251 °C (dec). IR (KBr): ν_max1694, 1240, 1215 cm^{-1 1}H

.

NMR (DMSO-d₆): δ 12.9 (bs, 2 H), 12.20 (s, 1 H), 8.18 (s, 1

H), 7.42 (d, 1 H, J ) 2.2 Hz), 7.33 (d, 1 H, J ) 1.8 Hz), 7.18 (d,

1 H, J ) 1.8 Hz), 7.15 (dd, 1 H, J ) 8.3, 2.2 Hz), 6.94 (d, 1 H,

J ) 8.3 Hz), 1/6 mol EtOAc present. MS: m/z 446, 444 (M⁺

+

1, with 4 Cl), 430, 428 (100), 426 (M⁺+ 1 - H₂O, with 4 Cl),

190, 99, 85. Anal. C₁₈H₉Cl₄NO₄: C, H, N.

3-(2-Carboxy-2-(furan-2-yl)vinyl)-4,6-dichloro-1H-indole-

2-carboxylic Acid, 11d from Diester 9d via Acid 10d.

Vinyl bromide 8Z (1.00 g, 1.62 mmol) was coupled with furan-

2-boronic acid (270 mg, 2.4 mmol) using general procedure I,

method A, to afford diester 9d (165 mg, 17%). Removal of the

tert-butyl ester gave crude acid 10d (0.1 g, 0.2 mmol), which

was hydrolyzed to afford 11d (59 mg, 59%) as an ivory solid:

1

mp 237-239 °C (dec). H NMR (DMSO-d₆): δ 13.3 (bs, 1 H),

3-(2-Carboxy-2-(naphthalen-1-yl)vinyl)-4,6-dichloro-

1H-indole-2-carboxylic Acid, 11b from Diester 9b via

Acid 10b. Vinyl bromide 8Z (928 mg, 1.50 mmol) was coupled

with 1-naphthaleneboronic acid (445 mg, 2.59 mmol) using

general procedure I, method A, to afford diester 9b (453 mg,

12.95 (bs, 1 H), 12.32 (s, 1 H), 7.90 (s, 1 H), 7.40 (d, 1 H, J )

1.8 Hz), 7.29 (dd, 1 H, J ) 1.7, 0.6 Hz), 7.12 (d, 1 H, J ) 1.8

Hz), 6.43 (dm, 1 H, J ) 3.4 Hz), 6.31 (dd, 1 H, J ) 3.4, 1.8

Hz). MS: m/z 368, 367, 366 (M⁺+ 1), 365 (M⁺), 350, 348 (M⁺

+ 1 - H₂O), 322, 279 (100), 85.

1

45%). H NMR (CDCl₃): δ 8.11 (s, 1 H), 7.87 (d, 1 H, J ) 1.7

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1007

3-(2-Carboxy-2-(furan-3-yl)vinyl)-4,6-dichloro-1H-indole-

2-carboxylic Acid, 11e from Diester 9e via Acid 10e. Vinyl

bromide 8Z (1.00 g, 1.62 mmol) was coupled with furan-3-

boronic acid (270 mg, 2.4 mmol) using general procedure I,

method A, to provide diester 9e (292 mg, 30%) as a tan oil.

Removal of the tert-butyl ester gave crude acid 10e, which was

hydrolyzed (vide supra) to give 11e (170 mg, 96%) as an ivory

solid: mp 223-225 °C (dec). ¹H NMR (DMSO-d₆): δ 13.0 (bs,

2 H), 12.36 (s, 1 H), 7.94 (s, 1 H), 7.48 (m, 1 H), 7.42 (d, 1 H,

J ) 1.7 Hz), 7.34 (m, 1 H), 7.15 (d, 1 H, J ) 1.7 Hz), 5.78 (m,

1 H). MS: m/z 368, 367, 366 (M⁺+ 1), 365 (M⁺), 350, 348 (M⁺

+ 1 - H₂O, 100).

(E)-3-[2-Carboxy-2-(4-chlorophenyl)vinyl]-4,6-dichloro-

1H-indole-2-caboxylic Acid, 11f from Diester 9f via Acid

10f. Freshly hydrolyzed 4-chlorobenzeneboronic acid (0.41 g,

2.63 mmol) was coupled with 8 using general procedure I,

method A, to afford 9f (595 mg, 52%) as a white amorphous

solid. ¹H NMR (300 MHz, CDCl₃): δ 7.93 (s, 1H), 7.84 (s, 1H),

7.60 (d, 2H, J ) 8.6 Hz), 7.23-7.26 (s overlapping d, 3H, J )

9 Hz), 7.07 (q, 4H, J ) 8.4 Hz), 4.16 (q, 2H, J ) 7.2 Hz), 2.39

(s, 3H), 1.52 (s, 9H, tBu), 1.25 (t, 3H, J ) 7.2 Hz). Anal. C₃₁H₂₈-

Cl₃SNO₆: C, H, N.

Hz), 7.08 (d, 1 H, J ) 2.8 Hz), 7.08 (d, 1 H, J ) 3.6 Hz), 6.81

(dd, 1 H, J ) 3.6, 2.8 Hz), 4.22 (q, 2 H, J ) 7.2 Hz), 2.40 (s, 3

H), 1.28 (t, 3 H, J ) 7.2 Hz)sEt₂O present. ¹H NMR (DMSO-

d₆): δ 13.07 (bs, 1 H), 7.88 (d, 1 H, J ) 1.7 Hz), 7.74 (d, 2 H,

J ) 8.4 Hz), 7.66 (s, 1 H), 7.57 (d, 1 H, J ) 1.7 Hz), 7.44 (d, 2

H, J ) 8.4 Hz), 7.27 (dd, 1 H, J ) 5.0, 2.9 Hz), 7.09 (dd, 1 H,

J ) 2.9, 1.2 Hz), 6.63 (dd, 1 H, J ) 5.0, 1.2 Hz), 4.11 (q, 2 H,

J ) 7.1 Hz), 2.36 (s, 3 H), 1.14 (t, 3 H, J ) 7.1 Hz)scyclohexane

and trace EtOAc present. MS: m/z 592 (M⁺+ 29), 566, 564

(M⁺+ 1), 548, 546 (M⁺+ 1 - H₂O), 520, 518, 394, 392, 157

(100), 139. Anal. C₂₅H₁₉Cl₂NO₆S₂: C, H, N.

Hydrolysis (vide supra) of acid 10h (1.147 g, 2.03 mmol) gave

11h as fine yellow crystals: mp 228-232 °C (dec). IR (KBr):

ν_max1690 cm^{-1 1}H NMR (DMSO-d₆): δ 13.3 (bs, 1 H), 12.8

.

(bs, 1 H), 12.24 (s, 1 H), 8.01 (s, 1 H), 7.37 (d, 1 H, J ) 1.7 Hz),

7.20 (dd, 1 H, J ) 5.0, 3.0 Hz), 7.12 (d, 1 H, J ) 1.7 Hz), 7.03

(dd, 1 H, J ) 3.0, 1.2 Hz), 6.66 (dd, 1 H, J ) 5.0, 1.2 Hz). MS:

m/z 410 (M⁺+ 29), 392, 383, 381 (M⁺), 366, 364 (100), 338,

336. Anal. C₁₆H₉Cl₂NO₄S: C, H, N.

3-[2-Carboxy-2-(4-methoxyphenyl)vinyl]-4,6-dichloro-

1H-indole-2-carboxylic Acid, 11i from Diester 9i via Acid

10i. Vinyl bromide 8Z (623 mg, 1.01 mmol) was coupled with

(p-methoxyphenyl)boronic acid (456 mg, 3.00 mmol) using

general procedure I, method B, to give diester 9i (427 mg). ¹H

NMR (CDCl₃): δ 7.91 (d, 1 H, J ) 1.7 Hz), 7.75 (s, 1 H), 7.63

(d, 2 H, J ) 8.5 Hz), 7.24 (d, 1 H, J ) 1.7 Hz), 7.21 (d, 2 H, J

) 8.6 Hz), 6.98 (d, 2 H, J ) 8.9 Hz), 6.64 (d, 2 H, J ) 8.9 Hz),

4.15 (q, 2 H, J ) 7.1 Hz), 3.74 (s, 3 H), 2.37 (s, 3 H), 1.53 (s, 9

H), 1.23 (t, 3 H, J ) 7.1 Hz). MS: m/z 672 (M⁺+ 29), 644 (M⁺

+ 1), 598, 590, 589, 588 (M⁺- C₄H₈, 100), 572, 570.

tert-Butyl ester 9f (213.0 mg, 0.328 mmol) was hydrolyzed

to give 10f (188 mg, 96%, 3:1 E/Z) as a white amorphous solid.

Further hydrolysis (LiOH‚H₂O) gave 114 mg (87%) of the crude

diacid as a yellow amorphous solid. Recrystallization from CH₂-

Cl₂/MeOH afforded diacid 11f (18 mg, 14%) as a pale yellow

1

solid: mp 252-254 °C (dec). H NMR (300 MHz, DMSO-d₆):

δ 13.1 (bs, 1.5 H), 12.2 (s, 1 H), 8.11 (s, 1 H), 7.34 (s, 1 H), 7.17

(m, 3 H), 6.97 (m, 2 H).

3-(2-Carboxy-2-(thiophen-2-yl)vinyl)-4,6-dichloro-1H-

indole-2-carboxylic Acid, 11g from Diester 9g via Acid

10g. Thiophene-2-boronic acid (1.12 g, 9.20 mmol) was coupled

with vinyl bromide 8 via general procedure I, method B, to

give 1.80 g (88%) of diester 9g (1.80 g, 88%) as an amorphous

solid. ¹H NMR (CDCl₃): δ 7.97 (d, 1 H, J ) 1.7 Hz), 7.80 (d, 2

H, J ) 8.5 Hz), 7.64 (s, 1 H), 7.27 (d, 2 H, J ) 8.5 Hz), 7.23 (d,

1 H, J ) 1.7 Hz), 7.16 (dd, 1 H, J ) 5.1, 1.2 Hz), 6.83 (dd, 1 H,

J ) 3.7, 1.2 Hz), 6.75 (dd, 1 H, J ) 5.1, 3.7 Hz), 4.24 (q, 2 H,

J ) 7.1 Hz), 2.39 (s, 3 H), 1.57 (s, 9 H), 1.26 (t, 3 H, J ) 7.1

Hz).

Removal of the tert-butyl ester (vide supra) with 96%

HCO₂H (6 mL), followed by recrystallization from cyclohexane/

EtOAc gave acid 10i (276 mg) as light ivory crystals: mp 175-

178 °C. IR (KBr): ν_max1728, 1692, 1371, 1271, 1250, 1179 cm^-1

.

¹H NMR (CDCl₃): δ 8.01 (s, 1 H), 7.92 (d, 1 H, J ) 1.7 Hz),

7.63 (d, 2 H, J ) 8.5 Hz), 7.26 (d, 1 H, J ) 1.7 Hz), 7.22 (d, 2

H, J ) 8.5 Hz), 7.01 (d, 2 H, J ) 8.8 Hz), 6.67 (d, 2 H, J ) 8.9

Hz), 4.16 (q, 2 H, J ) 7.2 Hz), 3.76 (s, 3 H), 2.38 (s, 3 H), 1.24

(t, 3 H, J ) 7.2 Hz)strace cyclohexane present. MS (CI, 70

eV): m/z 616 (M⁺+ 29), 590, 588 (M⁺+ 1), 572, 570, 157 (100),

139. Anal. C₂₈H₂₃Cl₂NO₇S: C, H, N.

Removal of the tert-butyl ester afforded 10g (1.31 g, 77%)

Hydrolysis of acid 10i (258 mg, 0.438 mmol) gave 11i as

fine yellow needles: mp 252 °C (dec). IR (KBr): ν_max1690,

as fine ivory crystals: mp 184-187 °C. IR (KBr): ν_max1726,

1696, 1373, 1275, 1206, 1196, 1177 cm^-^{1 1}H NMR (DMSO-

.

1

1611, 1248, 1177 cm^-1. H NMR (DMSO-d₆): δ 13.5-12.3 (2

d₆): δ 13.2 (bs, 1 H), 7.92 (d, 1 H, J ) 1.7 Hz), 7.80 (d, 2 H, J

) 8.4 Hz), 7.60 (s, 1 H), 7.57 (d, 1 H, J ) 1.7 Hz), 7.44 (d, 2 H,

J ) 8.4 Hz), 7.40 (dd, 1 H, J ) 4.7, 1.5 Hz), 6.84-6.8 (m, 2 H),

4.14 (q, 2 H, J ) 7.1 Hz), 2.37 (s, 3 H), 1.13 (t, 3 H, J ) 7.1

Hz)strace cyclohexane present. MS: m/z 594, 592 (M⁺+ 29),

566, 564 (M⁺+ 1, 100), 548, 546 (M⁺+ 1 - H₂O), 520, 518,

394, 392. Anal. C₂₅H₁₉Cl₂NO₆S₂: C, H, N.

H), 12.14 (s, 1 H), 7.99 (s, 1 H), 7.33 (d, 1 H, J ) 1.7 Hz), 7.14

(d, 1 H, J ) 1.7 Hz), 6.89 (d, 2 H, J ) 8.7 Hz), 6.65 (d, 2 H, J

) 8.7 Hz) 3.62 (s, 3 H). MS: m/z 407 (M⁺+ 1), 406, 405 (M⁺),

390, 388 (100), 362, 360. Anal. C₁₉H₁₃Cl₂NO₅: C, H, N.

(E)-4,6-Dichloro-3-(2-phenyl-2-(phenylcarbamoyl)vinyl)-

1-(toluene-4-sulfonyl )-1H-indole-2-carboxylic Acid Ethyl

Ester, 12a. To the acid ester 10a (340 mg, 0.61 mmol) in CH₂-

Cl₂(20 mL) at rt was added SOCl₂(80.0 mg, 0.05 mL, 0.671

mmol). After stirring for 15 min, aniline (62.5 mg. 0.061 mL,

0.671 mmol) was added and immediately a white precipitate

formed. The reaction was stirred an additional 2 h, diluted

with CH₂Cl₂, washed with H₂O, then with 1 N HCl, dried

(MgSO₄), and concentrated to a yellow oil. Flash chromatog-

raphy (3:1 hexane/Et₂O) provided 12a (262 mg, 68%) as a white

solid: mp 84-85 °C. ¹H NMR (300 MHz, CDCl₃): δ 7.90 (d,

2H, J ) 7.9 Hz), 7.61 (d, 2H, J ) 8.4 Hz), 7.49 (d, 2H, J ) 8.4

Hz), 7.11-7.33 (m, 12H), 4.21 (q, 2H, J ) 7.1 Hz), 2.39 (s, 3H),

1.28 (t, 3H, J ) 7.1 Hz). Anal. C₃₃H₂₆Cl₂SN₂O₅: C, H, N.

Hydrolysis of acid 10g (1.24 g, 2.20 mmol) gave 11g as a

bright yellow solid: mp 239-244 °C (dec). IR (KBr): ν_max1690

1

cm^-1. H NMR (DMSO-d₆): δ 7.92 (s, 1 H), 7.38 (d, 1 H, J )

1.7 Hz), 7.28 (dd, 1 H, J ) 5.1, 1.2 Hz), 7.12 (d, 1 H, J ) 1.7

Hz), 6.87 (dd, 1 H, J ) 3.7, 1.2 Hz), 6.77 (dd, 1 H, J ) 5.1, 3.7

Hz). MS: m/z 410 (M⁺+ 29), 392, 384, 383, 382 (M⁺+ 1), 381

(M⁺), 366, 364 (100), 338, 336. Anal. C₁₆H₉Cl₂NO₄S: C, H, N.

3-(2-Carboxy-2-(thiophen-3-yl)vinyl)-4,6-dichloro-1H-

indole-2-carboxylic Acid, 11h from Diester 9h via Acid

10h. Vinyl bromide 8Z (1.85 g, 3.00 mmol) was coupled with

thiophene-3-boronic acid (1.16 g, 9.07 mmol) using the general

procedure I, method B, to provide after 9 days diester 9h (1.53

4,6-Dichloro-3-(2-(phenylcarbamoyl)vinyl)-1H-indole-

2-carboxylic Acid, 13a. Hydrolysis (vide supra) of amide

ester 12a (0.68 g, 1.1 mmol) gave 13a (109 mg, 23%) as an

ivory powder: mp 276 °C (dec). IR (KBr): ν_max3397, 3298, 1690,

1

g) as a yellow oil. H NMR (CDCl₃): δ 7.94 (d, 1 H, J ) 1.7

Hz), 7.74 (d, 2 H, J ) 8.4 Hz), 7.72 (s, 1 H), 7.26 (d, 2 H, J )

8.0 Hz), 7.24 (d, 1 H, J ) 1.7 Hz), 7.03 (d, 1 H, J ) 4.4 Hz),

7.02 (d, 1 H, J ) 1.5 Hz), 6.77 (dd, 1 H, J ) 4.7, 1.6 Hz), 4.20

(q, 2 H, J ) 7.15 Hz), 2.40 (s, 3 H), 1.55 (s, 9 H), 1.28 (t, 3 H,

J ) 7.15 Hz).

1616, 1597, 1524, 1441, 1316 cm^-^{1 1}H NMR (DMSO-d₆): δ

.

13.31 (bs, 1 H), 12.18 (s, 1 H), 9.67 (s, 1 H), 7.81 (s, 1 H), 7.7-

7.8 (m, 2 H), 7.3-7.4 (m, 3 H), 7.1 (m, 5 H), 7.0 (m, 2 H)s

ether, MeCN, EtOAc present. ¹³C NMR (DMSO-d₆): δ 166.5,

161.7, 140.6, 139.1, 137.0, 136.2, 129.7, 128.7, 128.5, 127.7,

127.4, 127.3, 127.1, 123.5, 122.2, 120.8, 120.3, 115.9, 111.1.

MS: m/z 451 (M⁺+ 1), 136, 120 (100). Anal. C₂₄H₁₆Cl₂N₂O₃:

C, H, N.

Diester 9h was treated with TFA to give acid 10h (835 mg,

49%) as ivory crystals: mp 197-200 °C (dec). IR (KBr): ν_max

1723, 1694, 1373, 1277, 1206, 1196, 1177 cm^{-1 1}H NMR

.

(CDCl₃): δ 8.00 (s, 1 H), 7.95 (d, 1 H, J ) 1.7 Hz), 7.74 (d, 2

H, J ) 8.5 Hz), 7.27 (d, 2 H, J ) 8.5 Hz), 7.25 (d, 1 H, J ) 1.7

1008 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

4,6-Dichloro-3-(2-(benzylcarbamoyl)-2-phenylvinyl)-

1H-indole-2-carboxylic Acid, 13b from Amide Ester 12b.

Acid ester 10a (443 mg, 0.793 mmol) was reacted with

benzylamine (95 mL, 0.87 mmol), HOBT (118 mg, 0.771 mmol),

and EDC (167 mg, 0.871 mmol) in CH₂Cl₂(5 mL) to afford,

after recrystallization from CH₂Cl₂/Et₂O, 12b (0.34 g, 66%) as

a white powder: mp 150-153 °C. IR (KBr): ν_max3401, 1730,

mL) were heated at reflux for 16 h. Upon allowing the reaction

to cool to rt, the reaction was diluted with Et₂O and the

resulting solid was filtered, washed with Et₂O, and dried under

vacuum to yield a yellow solid. Recrystallization from acetone/

H₂O gave nitrile 23a (0.765 g, 40%) as a pale yellow solid: mp

215-217 °C (dec). IR (KBr): ν_max3350, 3086, 3059, 3034, 2986,

2939, 2904, 2222, 1730, 1685, 1606, 1558, 1531, 1496, 1475,

1448, 1438, 1421, 1388, 1367, 1342, 1321, 1303 cm^-1. ¹H NMR

(DMSO-d₆): δ 12.81 (s, 1 H), 8.33 (s, 1 H), 7.76 (d, 2 H, J )

7.0 Hz), 7.52 (m, 4 H), 7.35 (s, 1 H), 4.34 (q, 2 H, J ) 6.7 Hz),

1.25 (t, 3 H, J ) 6.8 Hz)sthe NMR suggested the presence of

1

1670, 1516, 1371 cm^-1. H NMR (CDCl₃): δ 7.88 (d, 1 H, J )

1.7 Hz), 7.83 (s, 1 H), 7.60 (m, 1 H), 7.58 (m, 1 H), 7.1-7.3 (m,

11 H), 7.1 (m, 2 H), 5.89 (t, 1 H, J ) 5.7 Hz), 4.54 (d, 2 H, J )

5.7 Hz), 4.21 (q, 2 H, J ) 7.1 Hz), 2.38 (s, 3 H), 1.26 (t, 3 H, J

) 7.1 Hz)sEt₂O present. ¹³C NMR (CDCl₃): δ 165.9, 160.7,

145.5, 140.6, 138.0, 137.1, 134.3, 134.0, 132.5, 129.8, 129.6,

128.7, 128.6, 128.3, 128.1, 127.8, 127.5, 127.1, 125.7, 125.6,

121.5, 114.0, 62.6, 44.1, 21.7, 13.8. MS: m/ z 675 (M⁺+ 29),

647 (M⁺+ 1), 493, 157 (100), 139. Anal. C₃₄H₂₈Cl₂N₂O₅S: C,

H, N.

a paramagnetic impurity. MS (EI, eV): m/z 384 (M⁺), (M⁺

-

HCO₂Et, 100). Anal. C₂₀H₁₄Cl₂N₂O₂: C, H, N.

Nitrile 23a (225 mg, 0.584 mmol) was heated in H₂SO₄(3

mL) and HOAc (3 mL) at 70 °C for 16 h. The reaction was

allowed to cool to rt and was poured onto H₂O (50 mL), and

the resulting solid was filtered, washed with additional H₂O,

and dried to give crude amide as a 65:35 E/Z mixture.

Recrystallization from acetone/H₂O provided 212 mg (90%) of

the intermediate E amide ester as a tan solid: mp 267-270

°C (dec). A solution of amide ester (202 mg, 0.50 mmol) and

LiOH‚H₂O (42 mg, 1.0 mmol) in 1:1 THF/H₂O (10 mL) was

heated at 60 °C for 16 h. The THF was then evaporated and

the residue diluted with water and acidified to pH 1 with 6 N

HCl. A precipitate formed that was filtered and washed with

additional H₂O and washed sparingly with Et₂O. Drying under

high vacuum afforded 13d (136 mg, 73%) as a tan solid, which

was an ∼5:1 E/Z isomeric mixture. Further hydrolysis of

amide carboxylic acid 13d to afford 1 failed by standard

hydrolysis methods. For additional information and conditions

see ref 22.

Hydrolysis (vide supra) of amide ester 12b (0.44 g, 0.68

mmol) afforded 13b (153 mg, 48%) as a white powder: mp 262

°C (dec). IR (KBr): ν_max3416, 3322, 1674, 1613, 1557, 1526,

1499, 1454, 1236, 1211 cm^-^{1 1}H NMR (DMSO-d₆): δ 12.07

.

(bs, 1 H), 8.05 (t, 1 H, J ) 6.1 Hz), 7.73 (s, 1 H), 7.2-7.3 (m,

6 H), 7.1 (m, 4 H), 6.9-7.0 (m, 2 H), 4.40 (d, 2 H, J ) 6.1 Hz).

¹³C NMR (DMSO-d₆): δ 167.2, 161.7, 140.1, 139.9, 136.8, 136.2,

129.5, 128.5, 128.2, 127.8, 127.5, 127.2, 127.0, 127.0, 126.6,

122.5, 120.6, 115.8, 111.0, 42.7. MS: m/z 493 (M⁺+ 29), 465

(M⁺+ 1), 429, 197. Anal. C₂₅H₁₈Cl₂N₂O₃: C, H, N.

4,6-Dichloro-3-(2-(methylcarbamoyl)-2-phenylvinyl)-

1H-indole-2-carboxylic Acid, 13c from Amide Ester 12c.

Acid ester 10a (800 mg, 1.43 mmol) was reacted (vide supra)

with methylamine hydrochloride (116 mg, 1.72 mmol), to afford

12c (376 mg, 46%) as a white powder. IR (KBr): ν_max3441,

1722, 1514, 1383, 1370 cm^-1. ¹H NMR (DMSO-d₆): δ 7.84 (m,

1 H), 7.65 (d, 2 H, J ) 8.6 Hz), 7.58 (m, 1 H), 7.53 (m, 1 H),

7.41 (d, 2 H, J ) 8.6 Hz), 7.33 (s, 1 H), 7.12-7.26 (m, 3 H),

6.93 (m, 2 H), 4.10 (q, 2 H, J ) 7.2 Hz), 2.69 (d, 3 H, J ) 4.6

Hz), 2.37 (s, 3 H), 1.13 (t, 3 H, J ) 7.1 Hz). MS: m/z 571 (M⁺

+ 1), 417 (100), 386, 157. Anal. C₂₈H₂₄Cl₂N₂O₅S: C, H, N.

4,6-Dichloro-3-(2-(dimethylcarbamoyl)-2-phenylvinyl)-

1H-indole-2-carboxylic Acid, 13e from Amide Ester 12e.

Acid ester 10a (0.81 g, 1.5 mmol) was reacted (vide supra) with

Me₂NH to give 12e (0.75 g, 88%) as a white powder: mp 73-

1

81 °C. IR (KBr): ν_max1732, 1638, 1389, 1371 cm^-1. H NMR

(CDCl₃): δ 7.95 (d, 1 H, J ) 1.7 Hz), 7.73 (m, 1 H), 7.69 (m, 1

H), 7.1-7.3 (m, 8 H), 6.99 and 6.69 (s, 1 H total, 1:5 ratio),

4.14 (q, 2 H, J ) 7.2 Hz), 3.06 and 3.04 and 3.00 (s, 6 H total),

2.39 (s, 3 H), 1.34 and 1.25 (t, 3 H total, J ) 7.2 Hz, 1:5 ratio).

¹³C NMR (CDCl₃): δ 170.2, 160.8, 145.6, 143.0, 137.1, 134.5,

134.3, 132.4, 129.9, 128.4, 128.1, 128.0, 128.0, 127.6, 127.3,

125.5, 122.3, 121.1, 120.4, 118.8, 114.0, 110.5, 62.7, 61.6, 38.2,

34.8, 26.9 (C₆H₁₂), 21.7, 14.2, 13.7. MS: m/z 585 (M⁺+ 1), 431

(100),157. Anal. C₂₉H₂₆Cl₂N₂O₅S: C, H, N.

Hydrolysis of amide ester 12c (335 mg, 0.586 mmol) using

LiOH‚H₂O (42 mg, 1.0 mmol) in THF (12 mL) and H₂O (5 mL)

provided 13c (58 mg, 25%) as a white powder. IR (KBr): ν_max

3421, 3233, 1665, 1632, 1528, 1298, 1242 cm^{-1 1}H NMR

.

(DMSO-d₆): δ 13.25 (bs, 1 H), 12.06 (s, 1 H), 7.66 (s, 1 H),

7.35 (m, 1 H), 7.31 (m, 1 H), 7.10 (m, 4 H), 6.93 (m, 2 H), 2.70

(d, 3 H, J ) 4.6 Hz). MS: m/z 389 (M⁺+ 1, 100), 371, 358,

353. Anal. C₁₉H₁₄Cl₂N₂O₃: C, H, N.

Hydrolysis of amide ester 12e (0.75 g, 1.3 mmol) gave 13e

(308 mg, 60%) as a white powder. The acid contained 6% of

the Z isomer by HPLC and NMR. For 13e: mp 170-176 °C.

IR (KBr): ν_max3233, 1692, 1613, 1557, 1534, 1497, 1443, 1402,

1209 cm^-1. ¹H NMR (DMSO-d₆): δ 13.27 (bs, 1 H), 12.18 (s, 1

H), 7.37 (d, 1 H, J ) 1.7 Hz), 7.1 (m, 4 H), 7.0 (m, 2 H), 6.85

(s, 1 H), 3.15 and 2.65 (s, 3 H total, 94:6 ratio), 2.96 and 2.62

(s, 3 H total, 94:6 ratio). ¹³C NMR (DMSO-d₆): δ 170.1, 161.8,

139.7, 137.1, 136.1, 128.7, 127.8, 127.4, 127.2, 122.3, 121.2,

120.7, 115.6, 111.1, 37.8, 34.2. MS: m/z 431 (M⁺+ 29), 403

(M⁺+ 1, 100), 157. Anal. C₂₀H₁₆Cl₂N₂O₃: C, H, N.

3-(2-Carbamoyl-2-phenylvinyl)-4,6-dichloro-1H-indole-

2-carboxylic Acid, 13d from Amide Ester 12d. Acid ester

10a (0.77 g, 1.4 mmol) was treated with NH₄Cl and Et₃N (1.1

equiv) to give, after two recrystallizations (CH₂Cl₂/Et₂O), 12d

(233 mg, 30%) as an ivory powder: mp 205-208 °C. IR (KBr):

1

ν_max1728, 1711, 1688, 1593, 1371 cm^-1. H NMR (CDCl₃): δ

7.88 (d, 1 H, J ) 1.7 Hz), 7.82 (s, 1 H), 7.61 (m, 1 H), 7.58 (s,

1 H), 7.1-7.3 (m, 8 H), 5.63 (bs, 1 H), 5.57 (bs, 1 H), 4.21 (q,

2 H, J ) 7.2 Hz), 2.38 (s, 3 H), 1.27 (t, 3 H, J ) 7.2 Hz)sEt₂O,

CH₂Cl₂present. ¹³C NMR (CDCl₃): δ 167.7, 160.6, 145.6, 140.0,

137.2, 134.6, 134.0, 132.6, 129.9, 129.7, 129.5, 128.6, 128.6,

128.3, 128.1, 127.1, 125.7, 125.5, 121.3, 114.0, 62.6, 21.7, 13.8.

MS: m/z 585 (M⁺+ 29), 557 (M⁺+ 1), 540, 511, 403, 386, 157

(100), 139, 129. Anal. C₂₇H₂₂Cl₂N₂O₅S: C, H, N.

4,6-Dichloro-3-(2-(phenethylcarbamoyl)-2-phenylvinyl)-

1H-indole-2-carboxylic Acid, 13f from Amide Ester 12f.

Acid ester 10a (1.08 g, 1.93 mmol) was reacted with phen-

ethylamine to afford 12f (0.87 g, 68%) as white needles: mp

108-112 °C. IR (KBr): ν_max1730, 1514, 1371, 1269, 1194, 1181,

Hydrolysis of amide ester 12d (233 mg, 0.418 mmol)

following recrystallization (CH₂Cl₂/Et₂O) afforded 13d (114 mg,

73%) as a tan powder: mp 169 °C (dec). IR (KBr): ν_max3393,

1

665, 581 cm^-1. H NMR (CDCl₃): δ 7.87 (d, 1 H, J ) 1.7 Hz),

7.75 (s, 1 H), 7.59 (m, 1 H), 7.57 (m, 1 H), 7.1-7.3 (m, 11 H),

6.9-7.0 (m, 2 H), 5.56 (bt, 1 H, J ) 5.6 Hz), 4.19 (q, 2 H, J )

7.2 Hz), 3.58 (q, 2 H, J ) 6.5 Hz), 2.82 (t, 2 H, J ) 6.8 Hz),

2.37 (s, 3 H), 1.26 (t, 3 H, J ) 7.2 Hz). ¹³C NMR (CDCl₃): δ

165.7, 160.6, 145.5, 140.8, 138.6, 137.1, 134.3, 134.0, 132.5,

129.8, 129.5, 128.7, 128.6, 128.4, 128.1, 127.3, 127.1, 126.5,

125.6, 121.5, 114.0, 62.5, 41.1, 35.3, 26.9, 21.6, 13.8. MS: m/z

689 (M⁺+ 29), 661 (M⁺+ 1, 100), 615, 505, 157. Anal. C₃₅H₃₀-

Cl₂N₂O₅S: C, H, N.

3283, 1680, 1615, 1578, 1559, 1534, 1240 cm^-^{1 1}H NMR

.

(DMSO-d₆): δ 13.24 (bs, 1 H), 12.06 (s, 1 H), 7.71 (s, 1 H),

7.31 (d, 1 H, J ) 1.7 Hz), 7.27 (bs, 1 H), 7.1 (m, 4 H), 6.9 (m,

3 H)sEt₂O present. ¹³C NMR (DMSO-d₆): δ 168.7, 161.7,

140.1, 136.8, 136.5, 129.4, 128.6, 127.9, 127.3, 127.3, 126.8,

122.4, 120.6, 116.2, 111.0. MS: m/z 403 (M⁺+ 29), 375 (M⁺

1, 100), 358. Anal. C₁₈H₁₂Cl₂N₂O₃: C, H, N.

+

3-(2-Carbamoyl-2-phenylvinyl)-4,6-dichloro-1H-indole-

2-carboxylic Acid, 13d from 6 via 23a. A mixture of

aldehyde 6 (1.43 g, 5.0 mmol), phenylacetonitrile (0.59 g, 0.60

mL, 5.0 mmol), and 4 drops of piperidine in 95% EtOH (30

Hydrolysis of amide ester 12f (0.87 g, 1.3 mmol) gave 13f

(321 mg, 51%) of as an ivory powder: mp 232 °C (dec). IR

(KBr): ν_max3418, 1611, 1555, 1528, 1447, 1379, 1333, 1281

Potent Glycine-Site NMDA Receptor Antagonist

cm^{-1 1}H NMR (DMSO-d₆): δ 12.43 (bs, 1 H), 7.75 (s, 1 H),

7.64 (bs, 1 H), 7.2-7.4 (m, 6 H), 6.97 (s, 5 H), 6.84 (d, 1 H, J

) 1.6 Hz), 3.43 (q, 2 H, J ) 6.8 Hz), 2.81 (t, 2 H, J ) 7.2 Hz).

¹³C NMR (DMSO-d₆): δ 167.9, 139.5, 138.3, 136.8, 136.0, 135.1,

130.0, 128.7, 128.3, 127.0, 126.5, 126.2, 126.0, 122.2, 119.3,

111.1, 40.8, 35.1. MS: m/z 507 (M⁺+ 29), 479 (M⁺+ 1), 435,

240, 148, 105 (100). Anal. C₂₆H₂₀Cl₂N₂O₃: C, H, N.

4,6-Dichloro-3-(3-(morpholin-4-yl)-3-oxo-2-phenylpro-

penyl)-1H-indole-2-carboxylic Acid, 13g from Amide

Ester 12g. Acid ester 10a (450 mg, 0.806 mmol) was reacted

with morpholine (80 mL, 0.92 mmol) to afford 12g (411 mg,

81%) as a white amorphous solid. ¹H NMR (CDCl₃): δ 7.95

(d, 1 H, J ) 1.7 Hz), 7.71 (d, 2 H, J ) 8.4 Hz), 7.0-7.3 (m, 8

H), 6.73 (s, 1 H), 4.12 (q, 2 H, J ) 7.2 Hz), 3.71 (bs, 4 H), 3.51

(bs, 4 H), 2.40 (s, 3 H), 1.23 (t, 3 H, J ) 7.2 Hz). Anal. C₃₁H₂₈-

Cl₂N₂O₆S: C, H, N.

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1009

.

stirring 16 h, was concentrated, diluted with DMF (50 mL),

cooled to 0 °C, and treated with MeI (6.8 g, 3 mL, 0.476 mol).

After stirring the reaction for 2.5 h at rt, the reaction was

diluted with H₂O, extracted with Et₂O, and washed twice with

H₂O, dried (MgSO₄), and concentrated to give the crude enol

ether, which was used without further purification. TMSOTf

(2.2 mL, 11.0 mmol) was added dropwise over 10 min followed

by the addition of the indole ester 5 (2.82 g, 10.9 mmol)

portionwise to the crude enol ether (2.8 g, 0.012 mol) in ClCH₂-

CH₂Cl (40 mL) at rt. The reaction was heated at 70 °C for 8 h

and then concentrated to a dark residue. This was diluted with

Et₂O, washed with saturated NaHCO₃, dried (MgSO₄), and

concentrated to a light brown solid that was chromatographed

(hexane: Et₂O: 3:1) to give the diester. The diester was then

subjected to base hydrolysis using excess LiOH‚H₂O heating

to 95 °C overnight to provide the desired diacid.

3-[2-(4-Fluorophenyl)-2-carboxyvinyl]-4,6-dichloro-1H-

indole-2-carboxylic Acid, 17a from 15a via 16a. (4-Fluo-

rophenyl)acetic acid methyl ester (14a; 10.9 g, 94%) was

prepared from (p-fluorophenyl)acetic acid (10.6 g, 68.8 mmol)

as a white solid which was used without further purification.

Hydrolysis of amide ester 12g (337 mg, 0.537 mmol) gave

acid 13g as a white crystalline powder: mp > 275 °C. IR

(KBr): ν_max1711, 1605, 1466, 1443 cm^-1. ¹H NMR (CDCl₃): δ

13.40 (bs, 1 H), 12.20 (s, 1 H), 7.37 (d, 1 H, J ) 1.8 Hz), 7.09-

7.15 (m, 4 H), 7.00-7.05 (m, 2 H), 6.90 (s, 1 H), 3.6 (m, 4

H-overlapping H₂O peak), 3.33 (bs, 4 H). MS: m/z 485 (M⁺

+

The enol ether 15a was generated from 14a (10.9 g, 64.8

mmol), as described in general procedure II, to afford 15a (11.2

g, 82%) as a light yellow/green solid: TLC, hexane/EtOAc (2:

1), R_f) 0.52. Condensation of enol ether 15a (1.74 g, 8.28

mmol) with indole 5 (1.94 g, 7.52 mmol), vida supra, afforded

16a (1.74 g, 53%) as a light yellow solid: TLC, cyclohexane/

EtOAc (7:3), R_f) 0.5. IR (KBr): ν_max1721, 1686, 1532, 1350,

41), 475, 473 (M⁺+ 29), 447, 446, 445 (M⁺+ 1, 100), 206. Anal.

C₂₂H₁₈Cl₂N₂O₄: C, H, N.

3-(2-(Benzyloxycarbamoyl)-2-phenylvinyl)-4,6-dichloro-

1-(toluene-4-sulfonyl)-1H-indole-2-carboxylic Acid Ethyl

Ester, 12h. The reaction (vide supra) of acid ester 10a (0.77

g, 1.4 mmol) with O-benzylhydroxylamine hydrochloride and

Et₃N (1.2 equiv) afforded 12h (1.33 g, 96%) as a yellow

powder: ∼7:1 E/Z ratio. IR (KBr): ν_max3422, 1730, 1678, 1371,

1271, 1194, 1177, 1007, 700, 665, 581 cm^-1. ¹H NMR (DMSO-

d₆): δ 11.25 and 11.16 (major) (2s, 1 H), 7.86 (major) (d, 1 H,

J ) 1.7 Hz), 7.7 (m, 2 H), 7.59 (major) (d, 1 H, J ) 1.7 Hz),

7.3-7.5 (m, 7 major and 9 minor H), 7.1-7.2 (m, 4 H), 6.8-

6.9 (m, 2 H), 4.9 (m, 2 H), 4.14 and 4.06 (major) (2q, 2 H, J )

7.2 Hz), 2.38 (major) (s, 3 H), 1.24 and 1.12 (major) (2t, 3 H, J

) 7.2 Hz). MS: m/z 691 (M⁺+ 29), 663 (M⁺+ 1), 509, 403,

386, 157, 107 (100), 91, 79.

1

1323, 1242 cm^-1. H NMR (DMSO-d₆): δ 12.25-12.40 (bs, 1

H), 8.08 (s, 1 H), 7.42-7.55 (m, 1 H), 7.38 (d, 1 H, J ) 1.6 Hz),

7.24 (m, 1 H), 7.20 (d, 1 H, J ) 1.6 Hz), 6.89-6.98 (m, 4 H),

4.27 (q, 2 H, J ) 7.1 Hz), 3.80 (s, 3 H), 1.25 (t, 3 H, J ) 7.1

Hc). ¹⁹F NMR (DMSO-d₆): δ -114.04 to -114.14 (m, 1 F).

MS: m/z 436 (M⁺+ 1), 404 (M⁺+ 1 - MeOH), 376. Anal.

C₂₁H₁₆Cl₂FNO₄: C, H, N.

Hydrolysis of p-fluoro ester 16a (900 mg, 2.06 mmol) in THF

(12 mL) and MeOH (6 mL) with aqueous 1.0 M NaOH (12 mL,

12.0 mmol) at 70 °C for 2.5 h gave, after recrystallization from

EtOAc, 17a (575 mg, 71%) as a light yellow solid. IR (KBr):

ν_max3434, 3281, 1687, 1615, 1510, 1238 cm^-1. ¹H NMR (DMSO-

d₆): δ 12.19 (s, 1 H), 8.08 (s, 1 H), 7.33 (d, 1 H, J ) 1.6 Hz),

7.16 (d, 1 H, J ) 1.6 Hz), 6.89-6.98 (m, 4 H). ¹⁹F NMR (DMSO-

d₆): δ -114.65 to +114.75 (m, 1 F). MS: m/z 394 (M⁺), 376

(M⁺- 18), 348, 174, 130, 85. Anal. C₁₈H₁₀Cl₂FNO₄: C, H, N.

4,6-Dichloro-3-(2-(hydroxycarbamoyl)-2-phenylvinyl)-

1-(toluene-4-sulfonyl)-1H-indole-2-carboxylic Acid Ethyl

Ester, 12i from Amide Ester 12j. To a solution of the acid

ester 10a (1.28 g, 2.29 mmol) in anhydrous CH₂Cl₂(10 mL)

was added DMF (2 drops) followed by oxalyl chloride (0.28 mL,

3.2 mmol). The reaction mixture was stirred at rt for 45 min.

The reaction was concentrated in vacuo, and the residue was

taken up in MeCN (6 mL). To this solution at 0 °C was added

a solution of N-t-Boc-O-TBDMS hydroxylamine (0.79 g, 3.2

mmol), DMAP (29 mg, 0.23 mmol), and Et₃N (0.45 mL, 3.2

mmol) in MeCN (9 mL). The reaction mixture was allowed to

stir at 0 °C for 4 h. The solution was concentrated in vacuo,

and the residue taken up in EtOAc, washed with water (2×),

and dried (MgSO₄). The residue was filtered through a short

column of SiO₂(eluting with CH₂Cl₂) to give 12j (1.44 g, 69%)

as a pale yellow solid. To a solution of 12j (1.44 g, 1.83 mmol)

in CH₂Cl₂(20 mL) at 0 °C was added CsF (0.31 g, 2.0 mmol)

and TFA (3.4 mL, 44 mmol). The reaction mixture was stirred

at 0 °C for 5 h, then diluted with CH₂Cl₂, washed with water,

and dried (MgSO₄). Filtration through SiO₂(eluting with

5-10% MeOH/CHCl₃) afforded 12i (0.34 g, 32%) as an ivory

powder. IR (KBr): ν_max1728, 1597, 1389, 1373, 1269, 1196,

(E)-3-[2-Carboxy-2-(4-chlorophenyl)-vinyl]-4,6-dichloro-

1H-indole-2-carboxylic Acid, 17b. Methyl (p-chlorophenyl)-

acetate (4.4 g, 0.024 mol) was treated as described in general

procedure II to afford the crude enol ether 15b (2.8 g, 52%) as

a yellow oil. ¹H NMR (300 MHz, CDCl₃): δ 7.56 (s, 1H), 7.24-

7.31 (m, 4 H), 3.74 (s, 3H), 3.65 (s, 3H). Compound 15b (2.8 g,

0.012 mol) was reacted with 5 to give methyl ethyl ester 16b

1

(1.2 g, 24%) as a white solid: mp 185-187 °C. H NMR (300

MHz, CDCl₃): δ 9.23 (br s, 1H), 8.23 (s, 1H), 7.23 (s, 1H), 7.12

(s, 1H), 6.99-7.10 (m, 4H), 4.31 (q, 2H, J ) 7.2 Hz), 3.87 (s,

3H), 1.34 (t, 3H, J ) 7.1 Hz). IR (KBr): ν_max3418, 3308, 3099,

3088, 3038, 2984, 2953, 2906, 1701, 1612, 1558, 1531, 1491,

1437, 1394, 1369 cm^-1. CIMS (CH⁴): m/e (rel intens) 482 (M

+ C₂H₅⁺, 20), 452 (M + H⁺, 70), 420 (M + H - MeOH, 100).

Anal. C₂₁H₁₆Cl₃NO₄: C, H, N.

Hydrolysis of 16b (1.1 g, 2.40 mmol) afforded the diacid 17b

(0.90 g, 90%) as a pale yellow solid: mp 252-254 °C (dec). ¹H

NMR (400 MHz, DMSO-d₆): δ 13.1 (br s, 1.5 H), 12.2 (s, 1H),

8.11 (s, 1H), 7.34 (s, 1H), 7.17 (m, 3H), 6.97 (m, 2H). ¹³C NMR

(100 MHz, DMSO-d₆): δ 167.4, 161.5, 136.9, 135.6, 134.7,

134.2, 131.6, 131.5, 128.8, 127.1, 127.0, 122.3, 120.9, 115.0,

111.2. IR (KBr): ν_max3429, 3271, 3161, 3099, 3090, 3049, 2694,

2661, 2660, 2615, 2542, 1695, 1635, 1614, 1558, 1535, 1493,

1456, 1435, 1410, 1394, 1369 cm^-1. CIMS (CH₄): m/e (rel inten)

438 (M + C₂H₅⁺, 3), 410 (M + H⁺, 15). Anal. C₁₈H₁₀Cl₃NO₄‚

1.0C₃H₆O: C, H, N.

1

1182, 1090, 1003, 581 cm^-1. H NMR (DMSO-d₆): δ 10.68 (s,

1 H), 9.04 (m, 1 H), 7.84 (d, 1 H, J ) 1.7 Hz), 7.65 (d, 2 H, J

) 8.3 Hz), 7.56 (d, 1 H, J ) 1.7 Hz), 7.39 (d, J ) 8.3 Hz), 7.1-

7.2 (m, 4 H), 6.9 (m, 2 H), 4.08 (q, 2 H, J ) 7.1 Hz), 2.36 (s, 3

H), 1.12 (s, 3 H). ¹³C NMR (DMSO-d₆): δ 164.6, 159.7, 146.3,

140.8, 136.3, 134.4, 133.0, 131.8, 130.3, 129.4, 128.9, 127.9,

127.8, 127.8, 126.8, 125.5, 122.3, 120.6, 113.4, 62.2, 21.1, 13.5.

MS: m/z 573 (M⁺+ 1), 540, 529 (100), 499, 483, 386, 375, 157.

General Procedure II for Lewis Acid-Catalyzed Enol

Ether Coupling. The respective enol ether was prepared by

reacting the desired substituted methyl phenyl acetate (0.023

mol) with NaH (95%, 1.3 g) in THF (50 mL) at 0 °C followed

by the addition of HCO₂CH₃(3.0 mL) and a few drops of

MeOH. The reaction was allowed to warm to rt and, after

3-[2-(4-Bromophenyl)-2-carboxyvinyl]-4,6-dichloro-1H-

indole-2-carboxylic Acid, 17c from 15c via 16c. (4-Bro-

mophenyl)acetic acid methyl ester (14c; 4.76 g, 90%) was

prepared as an oil from (p-bromophenyl)acetic acid (5.0 g, 23

1010 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

mmol) and used without further purification. The enol ether

15c was prepared (vide supra) from 14c (4.76 g, 20.8 mmol)

to afford 15c (5.13 g, 91%) as a colorless oil, which was used

without further purification. Condensation of 15c (5.13 g 18.92

mmol) with indole 5 (4.44 g, 17.2 mmol) provided 16c (5.54 g,

65%) as a light yellow solid that was used without further

purification.

tan powder. IR (KBr): ν_max3304, 1717, 1705, 1682, 1325, 1298,

1244, 1169, 1126, 1111, 1069 cm^-1. MS: m/z 514 (M⁺+ 29),

486 (M⁺+ 1), 466, 454 (100).

Hydrolysis (vide supra) of diester 16f (5.0 g, 10 mmol),

followed by recrystallization from EtOAc and acetone/cyclo-

hexane, gave 17f (1.46 g, 34%) as an ivory powder. IR (KBr):

ν_max3096, 1692, 1615, 1327, 1244, 1219, 1171, 1130, 1111, 1069

1

cm^-1. H NMR (DMSO-d₆): δ 13.13 (bs, 1 H), 12.22 (s, 1 H),

Hydrolysis of 16c (5.52 g, 11.1 mmol) provided 17c (2.92 g,

58%) as pale yellow crystals. IR (KBr): ν_max3425, 3049, 1691,

8.20 (s, 1 H), 7.5 (m, 2 H), 7.35 (d, 1 H, J ) 1.3 Hz), 7.2 (m, 3

H)sacetone present. ¹³C NMR (DMSO-d₆): δ 167.2, 161.4,

140.2, 136.9, 135.5, 134.9, 130.6, 128.9, 127.4, 127.0, 125.9,

124.03 124.0, 122.4, 122.3, 121.0, 114.7, 111.3. ¹⁹F NMR

(DMSO-d₆): δ -60.6 (s). MS: m/z 443 (M⁺+ 1), 426 (100).

Anal. C₁₉H₁₀F₃Cl₂NO₄: C, H, N.

1614, 1558, 1242 cm^-^{1 1}H NMR (DMSO-d₆): δ 12.96 (bs, 2

.

H), 12.21 (s, 1 H), 8.11 (s, 1 H), 7.35 (d, 1 H, J ) 1.8 Hz), 7.32

(m, 2 H), 7.29 (m, 2 H), 7.17 (d, 2 H, J ) 1.8 Hz). MS: m/z 456

(M⁺+ 1), 438 (M⁺+ H - 18), 410, 286, 206, 123. Anal. C₁₈H₁₀-

Cl₂BrNO₄‚H₂O: C, H, N.

3-[2-(4-Iodophenyl)-2-carboxyvinyl]-4,6-dichloro-1H-

indole-2-carboxylic Acid, 17d from 15d via 16d. (4-

Iodophenyl)acetic acid methyl ester (14d; 5.01 g, 94%) was

prepared (vide supra) and used without further purification.

The enol ether 15d was prepared from 14d, (5.01 g, 18.1 mmol)

to afford 15d (5.37 g, 93%) as a yellow oil, which was used

without further purification. Condensation of enol ether 15d

(3.84 g, 12.1 mmol) with indole 5 (2.83 g, 11.0 mmol) in 1,2-

dichloroethane (30 mL), using TMSOTf (2.30 mL, 12.1 mmol),

provided 16d (4.95 g, 87%, 7:3 ratio of E/Z isomers) as a white

powder: mp 213-215 °C (dec). IR (KBr): ν_max3416, 3306, 1748,

3-[2-Carboxy-2-(3-nitrophenyl)vinyl]-4,6-dichloro-1H-

indole-2-carboxylic Acid, 17g from 15g via 16g. Methyl

ester 14g (21.5 g, 100%) was prepared from m-nitrophenyl-

acetic acid (20.0 g, 110 mmol) as a white solid and was used

without further purification. The NMR agreed with published

1

data. H NMR (CDCl₃): δ 8.17 (d, 1 H, J ) 1.1 Hz), 8.14 (dd,

1 H, J ) 7.7, 1.0 Hz), 7.63 (dd, 1 H, J ) 7.7, 1.1 Hz), 7.52 (t,

1 H, J ) 7.7 Hz), 3.75 (s, 2 H), 3.73 (s, 3 H).

Methyl m-nitrophenylacetate (14g; 15.3 g, 78.3 mmol) was

treated as described in general procedure II to afford enol ether

15g (13.9 g, 75%) as a yellow solid: mp 101-103 °C.

1717, 1614, 1485, 1321, 1294, 1240 cm^-^{1 1}H NMR (DMSO-

.

This was reacted with indole 5 (11.7 g, 45.2 mmol) to afford

m-nitro diester 16g (13.0 g, 62%) as a yellow solid: TLC,

cyclohexane/EtOAc (7:3), R_f) 0.5. IR (KBr): ν_max1721, 1686,

1532, 1350, 1323, 1242 cm^-1. MS: m/z 463 (M⁺+ 1), 462, 433

(M⁺), 433, 431, 417, 371, 238, 210. The isomers can be

separated by fractional recrystallization from EtOAc/cyclo-

hexane. The Z isomer of 16g precipitates as a yellow powder,

which can then be recrystallized from Et₂O/cyclohexane to

obtain pure Z isomer of 16g as yellow needles: mp 178-180

°C. IR (KBr): ν_max3408, 3316, 1715, 1530, 1443, 1350, 1319,

d₆): δ 12.45 (bs, 1 H), 8.14 (s, 1 H), 7.70 and 7.50 (2d, 2 H, J

) 8.5 Hz), 7.45 and 7.39 (2d, 1 H, J ) 1.7 Hz), 7.22 and 7.21

(2d, 1 H, J ) 1.7 Hz), 6.94 and 6.75 (2d, 2 H, J ) 8.4 Hz), 4.19

(q, 2 H, J ) 7.0 Hz), 3.78 and 3.64 (2s, 3 H), 1.25 (t, 3 H, J )

7.0 Hz). MS: m/z 543 (M⁺), 512 (M⁺- MeOH), 498, 445, 417,

386, 326, 291, 213. Anal. C₂₁H₁₆Cl₂INO₄: C, H, N.

Hydrolysis of the p-iodo diester 16d (0.95 g, 1.8 mmol)

afforded 17d (520 mg, 56%) as a white powder. IR (KBr): ν_max

3427, 3312, 3271, 1692, 1613, 1240, 1221 cm^{-1 1}H NMR

.

1238, 1209, 1182 cm^-^{1 1}H NMR (DMSO-d₆): δ 12.46 (bs, 1

.

(DMSO-d₆): δ 13.09 (bs, 2 H), 12.21 (s, 1 H), 8.11 (s, 1 H),

7.45-7.50 (m, 2 H), 7.35 (d, 1 H, J ) 1.8 Hz), 7.17 (d, 1 H, J

) 1.8 Hz), 6.75-6.80 (m, 2 H). MS: m/z 502 (M⁺+ 1), 484,

456, 403, 375, 329. Anal. C₁₈H₁₀Cl₂INO₄: C, H, N.

H), 8.27 (t, 1 H, J ) 1.9 Hz), 8.22 (dm, 1 H, J ) 8.2 Hz), 7.92

(dm, 1 H, J ) 8.0 Hz), 7.71 (t, 1 H, J ) 8.0 Hz), 7.60 (s, 1 H),

7.44 (d, 1 H, J ) 1.7 Hz), 7.17 (d, 1 H, J ) 1.7 Hz), 4.26 (q, 2

H, J ) 7.1 Hz), 3.41 (s, 3 H), 1.23 (t, 3 H, J ) 7.1 Hz). ¹³C

NMR (DMSO-d₆): δ 166.0, 160.6, 147.7, 139.6, 137.1, 134.5,

133.8, 133.8, 129.9, 129.1, 127.3, 125.6, 122.7, 122.4, 122.0,

121.2, 116.4, 111.4, 61.1, 51.6, 13.9. MS: m/z 491 (M⁺+ 29),

462 (M⁺), 431 (100). Anal. C₂₁H₁₆Cl₂N₂O₆: C, H, N. The pure

E isomer of 16g was isolated as an ivory powder: mp 173-

175 °C. IR (KBr): ν_max3399, 3304, 1715, 1556, 1532, 1437,

3-(2-Carboxy-2-p-tolylvinyl)-4,6-dichloro-1H-indole-2-

carboxylic Acid, 17e from 15e via 16e. p-Tolylacetic acid

methyl ester (14e; 3.38 g, 88%) was prepared as an oil (3.50

g, 23.3 mmol) and used without further purification. The enol

ether 15e was prepared (vide supra) to afford 15e (2.36 g, 57%)

as a colorless oil. ¹H NMR (CDCl₃): δ 7.58 (s, 1 H), 7.24 (m, 4

H), 3.57 (s, 3 H), 3.76 (s, 3 H), 2.40 (s, 3 H). Condensation of

enol ether 15e (2.36 g, 11.4 mmol) with indole 5 (1.94 g, 7.52

1350, 1321, 1300, 1242 cm^-^{1 1}H NMR (DMSO-d₆): δ 12.35

.

1

mmol) provided 16e (2.73 g, 61%) as a light yellow solid. H

(bs, 1 H), 8.25 (s, 1 H), 7.96 (dm, 1 H, J ) 7.6 Hz), 7.86 (m, 1

H), 7.39 (t, 1 H, J ) 7.6 Hz), 7.36 (dm, 1 H, J ) 7.6 Hz), 7.33

(d, 1 H, J ) 1.7 Hz), 7.14 (d, 1 H, J ) 1.7 Hz), 4.18 (q, 2 H, J

) 7.1 Hz), 3.81 (s, 3 H), 1.23 (t, 3 H, J ) 7.1 Hz). ¹³C NMR

(DMSO-d₆): δ 165.8, 159.8, 146.8, 137.1, 136.7, 136.3, 136.0,

133.7, 129.4, 129.1, 126.9, 125.7, 124.5, 122.2, 122.1, 121.3,

114.7, 111.4, 61.1, 52.5, 13.2. MS: m/z 491 (M⁺+ 29), 462 (M⁺),

431 (100). Anal. C₂₁H₁₆Cl₂N₂O₆: C, H, N.

NMR (CDCl₃): δ 12.32 and 12.28 (2 s, 1 H), 8.08 (s, 1 H), 7.38

(s, 1 H), 7.2 (s, 1 H), 6.8-7.0 (dd, 4 H), 4.2 (q, 2 H), 3.78 (s, 3

H), 3.76 (s, 3 H), 2.16 (s, 3 H), 1.24 (t, 3 H).

Hydrolysis of 16e (2.73 g, 6.32 mmol) afforded 17e (1.53 g,

62%) as a light yellow solid. IR (KBr): ν_max3371, 1691, 1694,

1612, 1558, 1215 cm^-^{1 1}H NMR (DMSO-d₆): δ 12.76 (bs, 2

.

H), 12.12 (s, 1 H), 8.02 (s, 1 H), 7.32 (d, 1 H, J ) 1.8 Hz), 7.14

(d, 1 H, J ) 1.8 Hz), 6.89 (d, 2 H J ) 8.5 Hz), 6.85 (d, 2 H, J

) 8.5 Hz), 2.15 (s, 3 H). MS: m/z 389 (M^{+ -}1), 372 (M⁺- 18),

344, 115. Anal. C₁₉H₁₃Cl₂NO₄‚H₂O: C, H, N.

Hydrolysis (vide supra) of the m-nitro diester 16g (876 mg,

1.89 mmol) in THF (8 mL) and H₂O (4 mL) with LiOH (272

mg, 11.3 mmol) at 70 °C for 16 h provided, after recrystalli-

zation from EtOAc, diacid 17g (354 mg, 44%) of as a light

yellow powder: mp 272-275 °C. IR (KBr): ν_max3414, 3298,

3-[2-Carboxy-2-(4-(trifluoromethyl)phenyl)vinyl]-4,6-

dichloro-1H-indole-2-carboxylic Acid, 17f from 15f via

16f. (4-(Trifluoromethyl)phenyl)acetic acid methyl ester (14f)

was prepared according to a literature procedure.³¹The spectra

agreed with the published spectra.

Compound 14f (7.87 g, 36.1 mmol) was treated as described

in general procedure II to afford 15f (7.27 g, 77%) as a yellow

oil. IR (KBr): ν_max1740, 1713, 1632, 1620, 1437, 1327, 1279,

1262, 1194, 1165 cm^-1. ¹H NMR (CDCl₃): δ 7.61 (s, 1 H), 7.59

(d, 2 H, J ) 8.2 Hz), 7.47 (d, 2 H, J ) 8.2 Hz), 3.87 (s, 3 H),

3.75 (s, 3 H). ¹³C NMR (CDCl₃): δ 160.4, 130.5, 127.9, 124.6

(q, J ) 3.8 Hz), 62.2, 51.6. ¹⁹F NMR (CDCl₃): δ -63.1 Hz.

MS: m/z 521 (2M⁺+ 1), 289 (M⁺+ 29), 261 (M⁺+ 1), 257,

241, 229 (100), 213, 173, 75.

1691, 1612, 1530, 1350, 1242 cm^-^{1 1}H NMR (DMSO-d₆): δ

.

13.20 (bs, 1 H), 12.28 (s, 1 H), 8.27 (s, 1 H), 7.95-8.00 (m, 1

H), 7.85-7.90 (m, 1 H), 7.30-7.45 (m, 3 H), 7.19 (s, 1 H). MS:

m/z 420 (M⁺), 403 (M⁺- H₂O), 377, 258, 230, 195. Anal. C₁₈H₁₀-

Cl₂N₂O₆‚H₂O: C, H, N.

(E)-3-[2-Carboxy-2-(2-chlorophenyl)vinyl]-4,6-dichloro-

1H-indole-2-carboxylic Acid, 17h from 15h via 16h. (2-

Chlorophenyl)acetic acid methyl ester (14h; 4.1 g, 22 mmol)

was treated as described in general procedure II to afford the

1

crude enol ether 15h (2.85 g, 56%) as a yellow oil. H NMR

(CDCl₃): δ 7.57 (s, 1 H), 7.39-7.43 (m, 1 H), 7.21-7.26 (m, 3

H), 3.83 (s, 3 H), 3.70 (s, 3 H).

Compound 15f (5.62 g, 21.6 mmol) was then reacted with

indole 5 (5.07 g, 19.6 mmol) to afford 16f (5.07 g, 53%) as a

The crude enol ether 15h (2.8 g, 12 mmol) was treated (vide

supra) to afford diester 16h (1.3 g, 27%) as a white solid: mp

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1011

201-206 °C. IR (KBr): ν_max3420, 3410, 3304, 3084, 3024, 2987,

415 (M⁺- 18), 387, 373, 326, 235, 176, 123. Anal. C₂₀H₁₄-

Cl₂N₂O₅‚0.5HOAc‚0.5EtOAc: C, H, N.

2951, 2904, 1724, 1705, 1678, 1641, 1612, 1558, 1533, 1473,

1435, 1390, 1369, 1323, 1292, 1242, 1197 cm^-^{1 1}H NMR

.

3-[2-(3-(Benzoylamino)phenyl)-2-carboxyvinyl]-4,6-

dichloro-1H-indole-2-carboxylic Acid, 21b from 18 via

20b. The benzamide diester 20b was prepared (vide supra)

from m-amino diester 18 (866 mg, 2.0 mmol) and PhCOCl (0.58

mL, 5.0 mmol). The crude product was used without further

purification.

Hydrolysis of the benzamide diester 20b (920 mg, 1.43

mmol) gave, after crystallization from EtOAc/hexane, 21b (350

mg, 56%) as a yellow powder: mp 237-238 °C (dec). IR

(KBr): ν_max3420, 3275, 1686, 1611, 1537, 1234 cm^-1. ¹H NMR

(DMSO-d₆): δ 12.16 (s, 1 H), 10.07 (s, 1 H), 8.06 (s, 1 H), 7.85-

7.95 (m, 2 H), 7.45-7.60 (m, 4 H), 7.31 (d, 1 H, J ) 1.8 Hz),

7.13 (d, 1 H, J ) 1.8 Hz), 6.94 (t, 1 H, J ) 7.7 Hz), 6.65 (dd, 1

H, J ) 1.0, 7.7 Hz). MS: m/z 495 (M⁺), 256, 151. Anal. (C₂₅H₁₆-

Cl₂N₂O₅) C, H, N.

(CDCl₃): δ 8.99 (bs, 1 H), 8.32 (s, 1 H), 6.96-7.32 (m, 6 H),

4.35 (q, 2 H, J ) 7.4 Hz), 3.88 (s, 3 H), 1.37 (t, 3 H, J ) 7.1

Hz). MS: m/z 482 (M⁺+ 29), 452 (M⁺+ 1), 420 (M⁺+ 1 -

MeOH, 100). Anal. C₂₁H₁₆Cl₃NO₄: C, H, N.

Hydrolysis of 16h (1.3 g, 2.9 mmol) gave diacid 17h (0.9 g,

90%) as a pale yellow solid: mp 244-246 °C (dec). IR (KBr):

ν_max3423, 3259, 3161, 3090, 2692, 2681, 2671, 2656, 2613,

2604, 2542, 1695, 1633, 1614, 1558, 1533, 1473, 1438, 1411,

1

1392, 1369, 1338 cm^-1. H NMR (DMSO-d₆): δ 12.7 (bs, 1.2

H), 12.17 (s, 1 H), 8.16 (s, 1 H), 6.99-7.33 (overlapping m, 6

H). ¹³C NMR (DMSO-d₆): δ 167.2, 161.6, 136.7, 135.1, 134.8,

134.6, 133.3, 131.7, 128.9, 128.8, 128.7, 127.2, 127.1, 125.9,

122.6, 120.8, 114.5, 111.2. MS: m/z 438 (M⁺+ 29), 410 (M⁺

1). Anal. C₁₈H₁₀Cl₃NO₄‚0.5C₃H₆O: C, H, N.

+

3-[2-(3-Aminophenyl)-2-carboxyvinyl]-4,6-dichloro-1H-

indole-2-carboxylic Acid, 19 from 18. To a solution of

m-nitro ester 16g (16.2 g, 34.9 mmol) in EtOAc (175 mL) was

added SnCl₂‚2H₂O (47.2 g, 209 mmol). The mixture was heated

to reflux for 4 h and then allowed to cool to rt, neutralized

with saturated NaHCO₃, and diluted with H₂O and EtOAc;

the emulsion was allowed to dissipate, and the aqueous phase

separated. The aqueous phase was extracted with EtOAc (3×),

and the combined organic phases were washed with brine and

dried (MgSO₄). The crude product was purified by flash

chromatography (2:1 hexane/EtOAc) to afford amino diester

18 (13.9 g, 92%) as a light yellow solid: mp 249-251 °C. IR

3-[2-Carboxy-2-(3-(methoxycarbonylamino)phenyl)vi-

nyl]-4,6-dichloro-1H-indole-2-carboxylic Acid, 21c from

18 via 20c. Diester 18 (1.19 g, 2.74 mmol) was treated with

ClCO₂Me (0.25 mL, 3.3 mmol) to afford the pure E isomer of

20c (0.69 g, 51%) as a white powder: mp 192-194 °C. IR

(KBr): ν_max3293, 1742, 1705, 1611, 1553, 1441, 1321, 1300,

1285, 1240 cm^-1. ¹H NMR (CDCl₃): δ 10.61 (bs, 1 H), 8.16 (s,

1 H), 7.25 (d, 1 H, J ) 1.7 Hz), 7.19 (bs, 1 H), 7.04 (bs, 1 H),

7.01 (d, 1 H, J ) 1.7 Hz), 6.93 (t, 1 H, J ) 1.7 Hz), 6.64 (d, 1

H, J ) 7.6 Hz), 4.21 (q, 2 H, J ) 7.1 Hz), 3.79 (s, 3 H), 3.64 (s,

3 H), 2.19 (bs, 1 H), 1.28 (t, 3 H, J ) 7.1 Hz). ¹³C NMR

(CDCl₃): δ 167.3, 160.8, 153.8, 137.3, 137.0, 136.0, 135.9, 134.6,

130.4, 128.0, 127.8, 125.1, 124.6, 123.2, 121.8, 112.0, 117.4,

116.4, 110.9, 61.1, 52.2, 51.9, 14.1. MS: m/z 519 (M⁺+ 29),

490 (M⁺), 459 (100). Anal. C₂₃H₂₀Cl₂N₂O₆: C, H, N. Flash

chromatograhy (2:1 cyclohexane/EtOAc) of the filtrate gave an

∼1:1 mixture of E and Z isomers 20c (0.52 g, 39%) as a pale

yellow powder: mp 87-92 °C. IR (KBr): ν_max3337, 1705, 1611,

(KBr): ν_max3402, 3379, 3327, 1709, 1609, 1321, 1238 cm^-1

.

¹H NMR (DMSO-d₆): δ 12.36 (s, 1 H), 7.45 (d, 1 H, J ) 1.7

Hz), 7.32 (s, 1 H), 7.22 (d, 1 H, J ) 1.7 Hz), 7.05 (t, 1 H, J )

7.8 Hz), 6.67 (d, 1 H, J ) 1.9 Hz), 6.55-6.62 (m, 2 H), 5.16 (s,

1 H), 4.27 (q, 2 H, J ) 7.1 Hz), 3.39 (s, 3 H), 1.25 (t, 3 H, J )

7.1). MS: m/z 433 (M⁺), 432 (M^{+ -}1), 403, 401, 359, 327, 300.

Anal. C₂₁H₁₈Cl₂N₂O₄‚0.5H₂O: C, H, N.

1555, 1545, 1443, 1319, 1300, 1283, 1238 cm^-^{1 1}H NMR

.

To m-amino ester 18 (550 mg, 1.27 mmol) in THF (7 mL)

and H₂O (5 mL) was added LiOH (304 mg, 12.7 mmol). The

mixture was heated to 70 °C for 24 h. The mixture was allowed

to cool, the THF removed in vacuo, and the aqueous phase

washed with EtOAc. The aqueous phase was filtered through

Celite, acidified with aqueous NaHSO₄to pH 4, and extracted

three times with EtOAc. The combined organic phases were

washed with brine and dried (MgSO₄). The crude product was

recrystallized from EtOAc. The recrystallized material was

dissolved in hot MeOH, filtered through Celite, and precipi-

tated by the addition of H₂O. The precipitate was filtered

through a fritted disk, dried first under a stream of N₂, and

then dried in vacuo at 80 °C overnight to afford 19 (310 mg,

62%) as a light yellow powder: mp 211-220 °C. IR (KBr): ν_max

(CDCl₃): δ 9.34 (bs, 1 H), 9.15 (bs, 1 H), 8.20 (s, 1 H), 7.53 (bt,

1 H, J ) 1.7 Hz), 7.44 (m, 1 H), 7.43 (s, 1 H), 7.34 (t, 1 H, J )

8.0 Hz), 7.21 (dt, 1 H, J ) 7.6, 1.4 Hz), 7.18 and 7.16 (2d, 3 H,

J ) 1.7 Hz), 7.10 (d, 2 H, J ) 1.7 Hz), 7.08 (bt, 1 H, J ) 1.7

Hz), 7.00 (t, 1 H, J ) 8.0 Hz), 6.76 (bs, 1 H), 6.73 (dt, 1 H, J

) 7.7, 1.3 Hz), 6.50 (bs, 1 H), 4.33 (q, 2 H, J ) 7.1 Hz), 4.28 (q,

2 H, J ) 7.1 Hz), 3.85 (s, 3 H), 3.79 (s, 3 H), 3.68 (s, 3 H), 3.56

(bs, 3 H), 1.34 (t, 3 H, J ) 7.1 Hz), 1.31 (t, 3 H, J ) 7.1 Hz).

¹³C NMR (CDCl₃): δ 167.5, 167.3, 161.4, 160.7, 159.3, 154.0,

139.3, 137.8, 137.0, 136.8, 136.6, 136.5, 136.5, 136.0, 134.4,

131.9, 131.2, 131.0, 128.9, 128.7, 128.5, 128.1, 124.9, 124.8,

123.5, 123.4, 122.9, 122.4, 122.4, 118.3, 118.1, 117.8, 117.1,

110.6, 110.5, 61.6, 52.4, 52.2, 51.7, 14.2, 14.1. MS: m/z 519

(M⁺+ 29), 490 (M⁺), 459 (100), 234.

Hydrolysis of the diester 20c (1.11 g, 2.26 mmol) provided

21c (279 mg, 28%) as an ivory powder: mp 250 °C (dec). IR

(KBr): ν_max3372, 3081, 1688, 1609, 1589, 1539, 1443, 1294,

1240, 1175 cm^-1. ¹H NMR (DMSO-d₆): δ 12.86 (bs, 2 H), 12.10

(s, 1 H), 9.43 (s, 1 H), 8.05 (s, 1 H), 7.32 (d, 1 H, J ) 1.8 Hz),

7.2 (m, 2 H), 7.12 (d, 1 H, J ) 1.8 Hz), 6.96 (t, 1 H, J ) 7.9

Hz), 6.59 (dt, 1 H, J ) 7.7, 1.3 Hz), 3.59 (s, 3 H). ¹³C NMR

(DMSO-d₆): δ 167.9, 161.7, 153.7, 138.0, 136.8, 136.6, 136.4,

133.7, 128.5, 127.2, 127.1, 123.8, 122.3, 120.6, 119.8, 116.6,

115.2, 111.0, 51.4. MS: m/z 477 (M⁺+ 29), 459, 448 (M⁺), 431

(100), 399, 387. Anal. C₂₀H₁₄Cl₂N₂O₆: C, H, N.

3430, 3246, 1694, 1611, 1240 cm^{-1 1}H NMR (DMSO-d₆): δ

.

12.1 (s, 1 H), 7.93 (s, 1 H), 7.31 (d, 1 H, J ) 1.6 Hz), 7.12 (d,

1 H, J ) 1.1 Hz), 6.67 (t, 1 H, J ) 7.6 Hz), 6.27 (s, 1 H), 6.24

(d, 1 H, J ) 1.1 Hz), 6.10 (d, 1 H, J ) 7.6 Hz). MS: m/z 391

(M⁺), 373 (M⁺- 18), 347, 345, 329, 99. Anal. C₁₈H₁₂Cl₂N₂O₄‚

0.58H₂O: C, H, N.

3-[2-(3-(Acetylamino)phenyl)-2-carboxyvinyl]-4,6-dichlo-

ro-1H-indole-2-carboxylic Acid, 21a from 18 via 20a. To

a solution of the m-amino diester 18 (2.0 g, 4.6 mmol) in CH₂-

Cl₂(45 mL) was added Et₃N (1.9 mL, 14 mmol) and CH₃COCl

(0.82 mL, 12 mmol). The mixture was allowed to stir at rt for

20 h. The reaction was quenched by addition of MeOH and

the mixture diluted with CH₂Cl₂. The organic phase was

washed with brine and dried (MgSO₄). The crude product 20a

was used without further purification.

3-[2-Carboxy-2-(3-(methanesulfonylamino)phenyl)vi-

nyl]-4,6-dichloro-1H-indole-2-carboxylic Acid, 21d from

18 via 20d. Diester 18 (1.12 g, 2.58 mmol) was reacted with

MeSO₂Cl (0.24 mL, 3.1 mmol) to afford 20d (1.19 g, 90%) as a

yellow amorphous solid. IR (KBr): ν_max3410, 3297, 1703, 1609,

Hydrolysis (vide supra) of the N-acetyl ester 20a (2.39 g,

4.62 mmol) in THF (25 mL) and H₂O (20 mL) with LiOH (664

mg, 27.7 mmol) at 70 °C for 16 h gave 21a (1.74 g, 87%) as a

light yellow powder: mp 270-271 °C (dec). IR (KBr): ν_max

1437, 1321, 1240, 1152, 980 cm^-^{1 1}H NMR (CDCl₃); E

.

isomer: δ 9.37 (bs, 1 H), 8.24 (s, 1 H), 7.22 (d, 1 H, J ) 1.7

Hz), 7.07 (d, 1 H, J ) 1.7 Hz), 6.9-7.0 (m, 4 H), 6.65 (bs, 1 H),

4.31 (q, 2 H, J ) 7.2 Hz), 3.87 (s, 3 H), 2.60 (s, 3 H), 1.36 (t, 3

H, J ) 7.2 Hz). ¹H NMR (CDCl₃); Z isomer: 9.32 (bs, 1 H),

7.48 (s, 1 H), 7.3-7.4 (m, 4 H), 7.25 (d, 1 H, J ) 1.7 Hz), 7.12

(d, 1 H, J ) 1.7 Hz), 7.10 (m, 1 H), 4.35 (q, 2 H, J ) 7.2 Hz),

3.53 (s, 3 H), 3.06 (s, 3 H), 1.32 (t, 3 H, J ) 7.2 Hz). ¹³C NMR

3414, 3279, 1688, 1613, 1557, 1242 cm^{-1 1}H NMR (DMSO-

.

d₆): δ 12.14 (s, 1 H), 9.73 (s, 1 H), 8.03 (s, 1 H), 7.39 (dd, 1 H,

J ) 2.0, 7.7 Hz), 7.31 (d, 1 H, J ) 1.5 Hz), 7.24 (d, 1 H, J ) 2.0

Hz), 7.13 (d, 1 H, J ) 1.5 Hz), 6.94 (t, 1 H, J ) 7.7 Hz), 6.57

(dd, 1 H, J ) 1.0, 7.7 Hz), 1.94 (s, 3 H). MS: m/z 432 (M^{+ -}1),

1012 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

(CDCl₃): δ 167.2, 167.0, 161.3, 160.9, 140.0, 136.8, 136.7, 136.7,

136.6, 136.1, 136.0, 135.8, 134.9, 132.4, 131.3, 131.2, 129.6,

128.7, 128.5, 127.2, 125.2, 125.0, 124.9, 123.1, 122.6, 122.5,

122.2, 120.4, 120.1, 119.9, 118.1, 117.0, 110.7, 110.6, 61.8, 61.7,

52.5, 51.8, 39.4, 38.6, 21.0, 14.2. MS: m/z 511 (M⁺+ 1), 510

(M⁺), 479, 203, 186 (100), 160.

H), 6.2 (m, 3 major H), 2.67 and 2.38 (major) (2s, 3 H). ¹³C

NMR (DMSO-d₆): δ (major peaks) 168.1, 161.8, 148.6, 137.8,

136.9, 136.3, 133.0, 128.6, 127.4, 126.6, 122.6, 120.6, 117.1,

116.1, 112.7, 111.1, 110.8, 29.4. MS: m/z 433 (M⁺+ 29), 405

(M⁺+ 1, 100), 387, 361, 123. Anal. C₁₉H₁₄Cl₂N₂O₄‚0.91H₂O:

C, H, N.

Hydrolysis of the diester 20d (1.17 g, 2.29 mmol) provided

21d (0.50 g, 47%) as an ivory powder: mp > 270 °C. IR (KBr):

ν_max3418, 3300, 3200, 3094, 1686, 1615, 1319, 1306, 1240,

1140, 980 cm^-1. ¹H NMR (DMSO-d₆): δ 12.93 (bs, 2 H), 12.18

(s, 1 H), 9.44 (s, 1 H), 8.07 (s, 1 H), 7.32 (d, 1 H, J ) 1.7 Hz),

7.13 (d, 1 H, J ) 1.7 Hz), 7.08 (dt, 1 H, J ) 7.7, 0.7 Hz), 6.9

(m, 2 H), 6.80 (dm, 1 H, J ) 7.7 Hz), 2.48 (s, 3 H). ¹³C NMR

(DMSO-d₆): δ 167.6, 161.5, 137.1, 136.9, 136.8, 136.4, 134.0,

128.7, 127.9, 127.1, 126.9, 125.9, 122.2, 121.3, 120.8, 118.9,

115.4, 111.1, 38.0. MS: m/z 479, 468 (M⁺), 451 (100), 266, 186.

Anal. C₁₉H₁₄Cl₂N₂O₆S: C, H, N.

3-[2-Carboxy-2-(3-(dimethylamino)phenyl)vinyl]-4,6-

dichloro-1H-indole-2-carboxylic Acid, 21e from 18 via

20e. To a solution of the m-amino diester 18 (1.08 g, 2.5 mmol)

in AcOH (15 mL) was added paraformaldehyde (751 mg, 25.0

mmol) followed by NaBH₃CN (786 mg, 12.5 mmol), and the

mixture was stirred at rt for 20 h. The mixture was then

adjusted to pH < 9 with 1 M NaOH solution, extracted with

EtOAc, and the organic phases washed with brine and dried

(MgSO₄), affording after flash chromatography (2:1 hexane/

EtOAc) dimethylamino diester 20e (970 mg, 84%) as a light

yellow powder. ¹H NMR (DMSO-d₆): δ 12.24 (s, 1 H), 7.99 (s,

1 H), 7.34 (d, 1 H, J ) 1.6 Hz), 7.20 (d, 1 H, J ) 1.6 Hz), 6.92

(t, 1 H, J ) 8.5 Hz), 6.43 (dd, 1 H, J ) 8.5, 1.1 Hz), 6.30 (d, 1

H, J ) 8.5 Hz), 6.24 (d, 1 H, J ) 1.1 Hz), 4.20 (q, 2 H, J ) 7.4

Hz), 3.75 (s, 3 H), 2.58 (s, 6 H), 1.25 (t, 3 H, J ) 7.4 Hz).

3-[2-(4-Aminophenyl)-2-cyanovinyl]-4,6-dichloro-1H-

indole-2-carboxylic acid, 26a from 6 via 25a. To the

aldehyde 6 (858 mg, 3.0 mmol) in EtOH (45 mL) was added

(p-nitrophenyl)acetonitrile (584 mg, 3.6 mmol) and piperidine

(30 µL, 0.3 mmol). The mixture was refluxed for 18 h and

cooled to rt. The precipitate was filtered and washed with

EtOH and dried to afford 25a (1.18 g, 91%) as a bright yellow

solid. IR (KBr): ν_max3402, 3283, 2224, 1709, 1684, 1609, 1522,

1

1344, 1238 cm^-1. H NMR (DMSO-d₆): δ 12.92 (s, 1 H), 8.65

(s, 1 H), 8.36 (d, 1 H, J ) 8.9 Hz), 8.03 (d, 1 H, J ) 8.9 Hz),

7.53 (d, 1 H, J ) 1.6 Hz), 7.37 (d, 1 H, J ) 1.6 Hz), 4.34 (q, 2

H, J ) 7.1 Hz), 1.24 (t, 3 H, J ) 7.1). MS: m/z 430 (M⁺+ 1),

-

412, 387, 384 (M⁺EtOH). Anal. C₂₀H₁₃Cl₂N₃O₄: C, H, N.

Reduction of the p-nitro ester 25a (6.93 g, 16.1 mmol) in

EtOH (50 mL) with SnCl₂‚2H₂O (18.2 g, 80.5 mmol) yielded,

after flash chromatography (2:1 hexane/EtOAc), p-amino ester

25b (6.20 g, 96%) as a bright yellow solid. IR (KBr): ν_max3385,

3302, 2222, 1690, 1622, 1609, 1514, 1238 cm^-^{1 1}H NMR

.

(DMSO-d₆): δ 10.15 (s, 1 H), 7.85 (s, 1 H), 7.52 (d, 1 H, J )

7.4 Hz), 7.31 (s, 1 H), 7.14 (d, 1 H, J ) 1.3 Hz), 6.74 (d, 1 H,

J ) 7.4 Hz), 4.36 (q, 2 H, J ) 7.1 Hz), 1.27 (t, 3 H, J ) 7.1).

MS: m/z 400 (M⁺+ H), 399 (M⁺), 354 (M⁺- EtOH), 325, 290.

Anal. C₂₀H₁₅Cl₂N₃O₂: C, H, N.

Hydrolysis of the p-amino ester 25b (970 mg, 2.10 mmol)

afforded, after crystallization from EtOAc/hexane, nitrile acid

26a (494 mg, 56%) as a light yellow powder. IR (KBr): ν_max

3387, 3285, 2224, 1695, 1610, 1558, 1514, 1235 cm^-1. ¹H NMR

(DMSO-d₆): δ 12.54 (s, 1 H), 7.88 (s, 1 H), 7.47 (d, 1 H, J )

1.7 Hz), 7.40 (d, 1 H, J ) 8.5 Hz), 7.27 (d, 1 H, J ) 1.7 Hz),

Hydrolysis of 20e (970 mg, 2.10 mmol) provided, after

crystallization from EtOAc/hexane, 21e (494 mg, 56%) as a

light yellow powder: mp 256-258 °C. IR (KBr): ν_max3425,

1692, 1602, 1557, 1240 cm^-1. ¹H NMR (DMSO-d₆): δ 12.10 (s,

1 H), 7.99 (s, 1 H), 7.32 (d, 1 H, J ) 1.6 Hz), 7.14 (d, 1 H, J )

1.6 Hz), 6.91 (t, 1 H, J ) 7.7 Hz), 6.25-6.45 (m, 3 H), 2.62 (s,

6 H). MS: m/z 419 (M⁺), 401 (M⁺- 18), 383, 329, 271, 178.

Anal. C₂₀H₁₄Cl₂N₂O₅‚0.5H₂O‚0.5EtOAc: C, H, N.

-

6.66 (d, 1 H, J ) 8.5 Hz). MS: m/z 372 (M⁺), 371 (M⁺1),

336, 328, 285. Anal. C₁₈H₁₁Cl₂N₃O₂: C, H, N.

3-[2-(4-Aminophenyl)-2-carboxyvinyl]-4,6-dichloro-1H-

indole-2-carboxylic Acid, 26b. To the p-amino ester 25b

(6.20 g, 15.5 mmol) was added AcOH (20 mL) and H₂SO₄(20

mL). The reaction mixture was heated to 70 °C and stirred

for 3 h. The reaction mixture was cooled in an ice bath and

the resulting precipitate collected by filtration and dried in

vacuo affording the intermediate amide ester (3.0 g, 45%) as

a brown solid. ¹H NMR (DMSO-d₆): δ 12.24,12.20 (s, 1 H),

7.64 (s, 1 H), 7.40 (m, 2 H), 7.2 (s, 2 H), 6.84 (d, 2 H), 6.80 (d,

2 H), 4.20 (q, 2 H), 3.4-4.4 (bs, 2 H), 1.25 (t, 3 H). To the amide

ester (1.90 g, 5.38 mmol) was added 6 N NaOH (20 mL), and

the mixture was heated to 105 °C for 14 h. The mixture was

then cooled to 0 °C, acidified to pH 3 with 6 N HCl, and the

resulting precipitate collected by filtration and dried in vacuo

to afford 26b (1.34 g, 66%) as a brown solid. IR (KBr): ν_max

3-[2-Carboxy-2-(3-(methylamino)phenyl)vinyl]-4,6-

dichloro-1H-indole-2-carboxylic Acid, 21f from 18 via

20f. A mixture of m-amino diester 18 (5.45 g, 12.6 mmol) and

HCO₂Et (800 mL) was allowed to stir at rt overnight. By the

next day all the solids had dissolved. The HCO₂Et was

removed in vacuo, and the residue was taken up in EtOAc,

washed with saturated aqueous NaHCO₃, and dried (MgSO₄).

The crude formamide was dissolved in THF (50 mL), and BH₃‚

SMe₂complex (15 mL of a 2 M solution in THF) was added.

The resulting solution was heated to 60 °C for 15 min, then

cooled to rt, quenched with MeOH, and stirred for 2 h. The

crude material was chromatographed (7:3 cyclohexane/EtOAc)

to afford 20f (2.5 g, 44% of a ∼6:1 mixture of E/Z isomers) as

a yellow powder. IR (KBr): ν_max3410, 3306, 1707, 1680, 1607,

3395, 3271, 1724, 1612, 1176, 1082 cm^{-1 1}H NMR (DMSO-

.

d₆): δ 12.12 (s, 1 H), 7.87 (s, 1 H), 7.33 (d, 1 H, J ) 1.8 Hz),

7.10 (d, 1 H, J ) 1.8 Hz), 6.62 (d, 2 H, J ) 8.6 Hz), 6.23 (d, 2

H, J ) 8.6 Hz). MS: m/z 392 (M⁺+ 1), 390 (M^{+ -}1), 354, 348,

346, 310. Anal. C₁₈H₁₂Cl₂N₂O₄: C, H, N.

1

1559, 1441, 1321, 1296, 1244 cm^-1. H NMR (CDCl₃): δ 9.28

and 9.12 (major) (bs, 1 H), 8.13 (major) and 7.52 (s, 1 H), 7.23

(t, 1 minor H, J ) 7.8 Hz), 7.09 (d, 1 major H, J ) 1.7 Hz),

7.01 (d, 1 major H, J ) 1.7 Hz), 6.9 (m, 1 major + 1 minor H),

6.86 (m, 1 minor H), 6.62 (m, 1 minor H), 6.46 (m, 1 major H),

6.3 (m, 2 major + 1 minor H), 4.34 and 4.28 (major) (q, 2 H, J

) 7.2 Hz), 3.85 (major) and 3.58 (s, 3 H), 2.88 and 2.55 (major)

(s, 3 H), 1.35 (major) and 1.34 (t, 3 H, J ) 7.2 Hz). ¹³C NMR

(CDCl₃): δ (major peaks) 167.7, 161.0, 148.3, 137.5, 136.5,

136.1, 133.8, 131.0, 128.5, 128.2, 124.6, 123.7, 122.2, 118.9,

117.5, 113.8, 111.9, 110.9, 61.4, 52.3, 30.4, 14.1. MS: m/z 475

(M⁺+ 29), 447 (M⁺+ 1, 100), 415, 373, 341.

General Procedure III for the Mukaiyama Condensa-

tion of Ketene Silyl Acetals with 6. The ketene silyl acetals

28a-g of the arylalkylesters 27a-g were generated by treat-

ing the esters (29.3 mmol) at 0 °C, under N₂, with a cold

solution of TMSOTf (99%, 6.33 mL, 33.0 mmol) and Et₃N (4.6

mL, 33.0 mmol) in anhydrous Et₂O (60 mL), dropwise over 10

min. After stirring the reaction for 4 h, the reaction was

allowed to gradually warm to rt, at which time the colorless

reaction acquired a yellow denser layer. This was separated

from the Et₂O layer, and the Et₂O layer was concentrated to

a colorless oil. In a separate reaction vessel Ph₃PO (1.3 g, 4.68

mmol) was cooled to 0 °C under N₂in CH₂Cl₂. A solution of

Tf₂O (0.40 mL, 2.34 mmol) in CH₂Cl₂(30 mL) was added

dropwise over 5 min. After 10 min this solution was added

dropwise over 5 min to a stirred suspension of the freshly

prepared ketene silyl acetal and 6 (3.7 g, 12.85 mmol) in CH₂-

Cl₂at -78 °C. The reaction was stirred at -78 °C for 1 h

Hydrolysis of diester 20f (1.9 g, 4.2 mmol) gave 21f (0.70 g,

41%, a ∼6:1 mixture of E/Z isomers) as an ivory powder: mp

193 °C (dec). IR (KBr): ν_max3426, 1694, 1609, 1557, 1535, 1437,

1414, 1366, 1331, 1236 cm^-^{1 1}H NMR (DMSO-d₆): δ 12.25

.

and 12.07 (major) (2s, 1 H), 7.93 (major) and 7.39 (2s, 1 H),

7.42 and 7.30 (major) (2d, 1 H, J ) 1.7 Hz), 7.18 and 7.12

(major) (2d, 1 H, J ) 1.7 Hz), 7.09 (m, 1 minor H), 6.76 (t, 1

major H, J ) 7.8 Hz), 6.6-6.7 (m, 2 minor H), 6.51 (m, 1 minor

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1013

during which time a white solid started to precipitate. The

reaction was allowed to gradually warm to rt overnight. After

16 h the reaction was poured on H₂O and extracted with

EtOAc, washed with saturated NaHCO₃followed by brine,

dried (MgSO₄), and concentrated to an amber oil which

solidified upon standing. Recrystallization from hot Et₂O/

CHCl₂afforded pure product.

Silyl ether 29b (577 mg, 1.10 mmol) was treated (vide supra)

with Tf₂O (200 µL, 1.19 mmol) to afford diester 31b (195 mg

of >95% pure E) as fine white crystals. Recrystallization/

trituration of the oily yellow mother liquor with Et₂O gave an

additional 254 mg (449 mg total, 49%) of light yellow crystals

containing a small amount of aldehyde from the retro-aldol

reaction. For 31b: mp 152-153 °C (dec). IR (KBr): ν_max1717,

1248 cm^-1. ¹H NMR (CDCl₃): δ 9.05 (bs, 1 H), 8.44 (ddd, 1 H,

J ) 4.8, 1.8, 1.0 Hz), 8.26 (s, 1 H), 7.36 (td, 1 H, J ) 7.8, 1.8

Hz), 7.22 (d, 1 H, J ) 1.7 Hz), 7.10 (d, 1 H, J ) 1.7 Hz), 7.0

(m, 2 H), 4.36 (q, 2 H, J ) 7.1 Hz), 4.22 (q, 2 H, J ) 7.1 Hz),

1.42 (s, 3.6 H, cyclohexane), 1.34 (2t, 6 H, J ) 7.1 Hz). MS:

m/z 473 (M⁺+ 41), 463, 461 (M⁺+ 29), 435, 433 (M⁺+ 1,

100), 361, 359. Anal. C₂₁H₁₈Cl₂N₂O₄‚0.3C₆H₁₂: C, H, N.

4,6-Dichloro-3-(2-(methoxycarbonyl)-2-phenylvinyl)-

1H-indole-2carboxylic Acid Ethyl Ester, 31a from 27a via

29a/30a. Methyl phenylacetate 27a (1.80 mL, 12.5 mmol) was

treated as described in general procedure III and reacted with

aldehyde 6 (1.57 g, 5.49 mmol) to afford 29a (85:15 mixture of

two diastereomers, 333 mg, 12%,) and 30a (253 mg of less polar

diastereomer A, 623 mg of diastereomeric mixture, and 407

mg of more polar diastereomer B; total 1.283 g, 54%), all as

white solids. The TMS ethers 29a were recrystallized from

cyclohexane to give fine white crystals: mp 150-153 °C. IR

(KBr): ν_max3329, 1730, 1688, 1238, 1167, 1080, 843 cm^-1. ¹H

NMR (CDCl₃): δ 9.01 and 8.68 (major) (two s, 1 H), 7.28

(major) and 7.14 (two d, 1 H, J ) 1.8 Hz), 7.2-7.25 (m, 2 H of

minor) and 6.95-7.07 (m, 5 H of major and 3 H of minor), 7.18

(major) and 7.14 (two d, 1 H, J ) 1.8 Hz), 6.76 and 6.62 (major)

(two d, 1 H, J ) 10.1 and 10.5 Hz), 4.93 and 4.72 (major) (two

d, 1 H, J ) 10.1 and 10.5 Hz), 4.54 (q, J ) 7.1 Hz; 2H of minor),

4.39 and 4.30 (two dq, J ) 10.8, 7.1 Hz, 2H of major), 3.75

(major) and 3.74 (two s, 3 H), 1.53 and 1.47 (major) (two t, 3

H, J ) 7.1 Hz), -0.05 (major) and -0.07 (two s, 9 H). MS:

m/z 536 (M⁺+ 29), 510, 508 (M⁺+ 1), 494, 492, 420, 418, 388,

386, 360, 358 (100). Anal. C₂₄H₂₇Cl₃NO₅Si: C, H, N. The

diastereomerically pure fractions of alcohols 30a were indi-

vidually recrystallized from cyclohexane/EtOAc. Alcohol 30a

(diastereomer A) was obtained as fine white needles: mp 206-

208 °C (change at 170 °C). IR (KBr): ν_max1740, 1686, 1246

cm^-1. ¹H NMR (DMSO-d₆): δ 12.24 (bs, 1 H), 7.49 (d, 1 H, J )

1.8 Hz), 7.25-7.48 (m, 5 H), 7.29 (d, 1 H, J ) 1.8 Hz), 6.44

(dd, 1 H, J ) 9.9, 7.1 Hz), 5.24 (d, 1 H, J ) 7.1 Hz), 4.56 (bs,

1 H), 4.43 (q, 2 H, J ) 7.1 Hz), 3.18 (s, 3 H), 1.43 (t, 3 H, J )

7.1 Hz). MS: m/z 438, 436 (M⁺+ 1), 420, 418, 388, 386, 288,

286 (100). Anal. C₂₁H₁₉Cl₂NO₅: C, H, N. Aldol 30a (diastereo-

mer B) was isolated as white granules: mp 177-178 °C. IR

(KBr): ν_max1736, 1723, 1686, 1310, 1244, 1167 cm^-1. ¹H NMR

(CDCl₃): δ 8.99 (bs, 1 H), 7.10-7.15 (m, 3 H), 6.97-7.03 (m,

4 H), 6.64 (dd, 1 H, J ) 11.2, 10.1 Hz), 5.30 (bs, 1 H), 4.50 (q,

2 H, J ) 7.1 Hz), 4.38 (bd, 1 H), 3.78 (s, 3 H), 1.49 (t, 3 H, J

) 7.1 Hz)strace cyclohexane and EtOAc present. MS: m/z 438,

436 (M⁺+ 1), 420, 418, 388, 386, 288, 286 (100). Anal. C₂₁H₁₉-

Cl₂NO₅: C, H, N.

Hydrolysis of diester 31b (429 mg, 0.991 mmol) gave 32b

(as the E isomer) which upon drying in vacuo at 125 °C

isomerized to a 6:1 E/Z mixture of isomers: mp 187-189 °C

1

(dec). IR (KBr): ν_max3426, 1684, 1611, 1240 cm^-1. H NMR

(DMSO-d₆); E isomer only: δ 13.2 (bs, 2 H), 12.23 (s, 1 H),

8.33 (dm, 1 H, J ) 4.8 Hz), 8.16 (s, 1 H), 7.54 (td, 1 H, J ) 7.8,

1.8 Hz), 7.35 (d, 1 H, J ) 1.8 Hz), 7.13 (d, 1 H, J ) 1.8 Hz), 7.1

(m, 1 H), 7.01 (dt, 1 H, J ) 7.9, 1.0 Hz). MS: m/z 405 (M⁺

+

29), 379, 377 (M⁺+ 1), 335, 333 (100), 291, 289, 288, 287, 260,

258, 253. Anal. C₁₇H₁₀Cl₂N₂O₄‚0.8H₂O: C, H, N.

3-[2-Carboxy-2-(3-methoxyphenyl)vinyl]-4,6-dichloro-

1H-indole-2-carboxylic Acid, 32c from 27c via 30c and

31c. Methyl (m-methoxyphenyl)acetate (27c) was prepared

according to a literature procedure.³²The spectra agreed with

the published spectra.

The aldol condensation was carried out as described in

general procedure III using 1-methoxy-1-(trimethylsiloxy)-2-

(m-methoxyphenyl)ethylene (28c). The erythro/threo alcohols

30c (2.42 g, 31%) were obtained as a white powder, including

pure threo alcohol 30c (0.85 g) as a white powder: mp 252 °C

(dec). IR (KBr): ν_max3401, 1736, 1690, 1611, 1559, 1435, 1319,

1

1242, 1165 cm^-1. H NMR (CDCl₃): δ 8.96 (bs, 1 H), 7.13 (d,

1 H, J ) 1.6 Hz), 7.03 (bs, 1 H), 6.89 (t, 1 H, J ) 8.0 Hz), 6.80

(bs, 1 H), 6.67 (dd + m, 2 H, J ) 11.3, 10.1 Hz), 6.54 (ddd, 1

H, J ) 8.2, 2.5, 0.9 Hz), 5.26 (bs, 1 H), 4.49 (q, 2 H, J ) 7.1

Hz), 4.39 (m, 1 H), 3.78 (s, 3 H), 3.65 (s, 3 H), 1.48 (t, 3 H, J

) 7.1 Hz)splus minor impurities. ¹³C NMR (CDCl₃): δ 172.9,

159.1, 136.3, 131.3, 128.9, 123.3, 121.2, 113.9, 110.4, 67.7, 62.4,

59.1, 55.1, 52.3, 14.3. MS: m/z 494 (M⁺+ 29), 466 (M⁺+ 1),

448, 416, 286 (100), 181, 149, 121. Anal. Calcd for C₂₂H₂₁Cl₂-

NO₆.

Dehydration of the m-methoxy aldol products 30c gave 31c

(1.5 g, 63%, ∼3:2 ratio of E/Z isomers) as a ivory powder: mp

147-150 °C. IR (KBr): ν_max3302, 1717, 1678, 1609, 1559, 1435,

To a stirred solution of alcohols 30a (diastereomeric mixture,

232 mg, 0.532 mmol) in toluene (25 mL) at 65 °C under N₂

was added p-TsOH (10 mg). The solution was heated at gentle

reflux for 2.5 h and then allowed to cool to rt before being

washed twice with H₂O. Concentration in vacuo gave diesters

31a (231 mg, 100%) as a white solid, which was a ∼2:1 mixture

of E/Z isomers by NMR.

1

1321, 1289, 1242, 1179 cm^-1. H NMR (CDCl₃): δ 9.34 (bs, 1

H Z isomer), 9.15 (bs, 1 H E isomer), 8.18 (s, 1 H E isomer),

7.45 (s, 1 H Z isomer), 7.33 (t, 1 H Z isomer, J ) 7.9 Hz), 7.16

(d, 1 H E isomer, J ) 1.7 Hz), 7.1 (m, 2 H E isomer + 2 H Z

isomer), 7.01 (m, 1 H E isomer), 6.92 (m, 1 H Z isomer), 6.7-

6.6 (m, 2 H E isomer + 2 H Z isomer), 4.34 (t, 2 H E isomer,

J ) 7.1 Hz), 4.27 (t, 2 H Z isomer, J ) 7.1 Hz), 3.86 (s, 3 H E

isomer), 3.85 (s, 3 H Z isomer), 3.59 (s, 3 H Z isomer), 3.54 (s,

3 H E isomer), 1.35 (t, 3 H E isomer, J ) 7.1 Hz), 1.32 (t, 3 H

Z isomer, J ) 7.1 Hz). ¹³C NMR (CDCl₃): δ 167.7, 167.3, 161.4,

160.8, 159.4, 158.5, 139.7, 137.1, 136.9, 136.7, 136.5, 136.4,

134.2, 131.6, 131.2, 131.0, 129.2, 128.6, 128.5, 128.4, 124.9,

124.6, 123.6, 122.4, 122.3, 122.2, 120.7, 118.1, 117.3, 115.3,

114.0, 113.4, 113.2, 110.5, 110.5, 61.5, 55.3, 54.9, 52.4, 51.8,

14.2, 14.1. MS: m/z 476 (M⁺+ 29), 447 (M⁺), 416 (100). Anal.

C₂₂H₁₉Cl₂NO₅: C, H, N.

3-(2-Carboxy-2-(pyridin-2-yl)vinyl)-4,6-dichloro-1H-in-

dole-2-carboxylic Acid, 32b from 27b via 29b and 31b.

The ketene silyl acetal 28b was prepared from ethyl 2-pyridyl-

acetate as described in general procedure III to afford silyl

ether 29b (1.178 g). The early and late fractions containing

minor impurities were recrystallized from Et₂O/pentane to give

a second and third crop of crystals (overall yield of 1.804 g,

61%, a mixture of erythro/threo isomers) as fine ivory crys-

tals: mp 161-176 °C. IR (KBr): ν_max1732, 1719, 1248, 1211,

1177, 1082, 843 cm^-1. ¹H NMR (CDCl₃): δ 10.82 and 10.71 (s

and bs, 0.2 H), 9.32, 9.30, 9.23 and 8.84 (major) (4 s, 1 H),

8.62 and 8.23 (major) (2 m, 1 H), 7.13-7.72 (m, 5 H), 6.88-

6.97 (m, 1 H), 6.79, 6.75 (major) and 6.72 (3 d, 1 H, J ) 10.3,

10.5, and 10.3 Hz, respectively), 5.19, 5.17 (major), 4.99

(major), and 4.98 (4 d, 1 H, J ) 9.9, 10.2, 10.4 and 10.3 Hz,

respectively), 4.12-4.56 (m, 4 H), 3.73-3.86 (m, 0.6 H), 1.36-

1.53 (m, 4 H), 1.29 (t, 2 H, J ) 7.1 Hz), 0.75-0.85 (m, 1 H),

-0.05, -0.06, -0.31, and -0.33 (4 s, 9 H). MS: m/z 525, 524,

523 (M⁺+ 1), 522 (M⁺), 509, 507, 487, 435, 433, 360, 358 (100).

Anal. C₂₄H₂₈Cl₂N₂O₅Si: C, H, N.

Hydrolysis (vide supra) of the diesters 31c gave, after flash

chromatography (5-7% AcOH/CH₂Cl₂) and recrystallization

from acetone/H₂O, 32c (640 mg, 49%) as a ivory powder: mp

269 °C (dec). IR (KBr): ν_max3414, 3405, 3352, 1690, 1613, 1559,

1289, 1248, 1215 cm^-^{1 1}H NMR (DMSO-d₆): δ 13.03 (bs, 2

.

H), 12.15 (s, 1 H), 8.05 (s, 1 H), 7.33 (d, 1 H, J ) 1.7 Hz), 7.16

(d, 1 H, J ) 1.7 Hz), 7.00 (t, 1 H, J ) 8.0 Hz), 6.64 (ddd, 1 H,

J ) 8.3, 2.6, 1.0 Hz), 6.5-6.6 (m, 2 H), 3.52 (s, 3 H). ¹³C NMR

(DMSO-d₆): δ 167.6, 161.6, 157.9, 137.1, 136.8, 136.8, 133.8,

128.7, 128.1, 127.1, 126.7, 122.5, 121.9, 120.7, 115.6, 115.4,

1014 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

112.2, 111.1, 54.5. MS: m/z 416, 405 (M⁺), 388 (100). Anal.

C₁₉H₁₃Cl₂NO₅: C, H, N.

61.6 t, 61.1 t, 51.5 q, 51.4 q, 47.8 d, 47.1 d, 38.3 d, 38.0 d, 29.2

t, 29.0 t, 27.4 t, 27.2 t, 14.3 q, 13.8 q. CIMS (CH₄): m/e (rel

intens) 474 (M + C₂H₅⁺, 8), 446 (M + H⁺, 24), 385 (M⁺- HCO₂-

Me), 189 (100). Anal. C₂₃H₂₁Cl₂NO₄: C, H, N.

3-[2-Carboxy-2-(3-phenoxyphenyl)vinyl]-4,6-dichloro-

1H-indole-2-carboxylic Acid, 32d from 27d via 29d/30d

and 31d. Methyl (m-phenoxyphenyl)acetate 27d was prepared

according to the literature procedure.^{32 1}H NMR (CDCl₃): δ

7.2-7.4 (m, 4 H), 7.14 (m, 1 H), 7.0 (m, 4 H), 3.73 (s, 3 H),

3.64 (s, 2 H).

Mukaiyama coupling of aldehyde 6 (2.0 g, 7.0 mmol) with

the ketene silyl acetal 28d, prepared from 27d (5.08 g, 21.0

mmol) was carried out as described in general procedure III

to afford TMS ethers 29d (0.92 g, 22%) and alcohols 30d (1.27

g, 34%).

Alcohols 30d (1.27 g, 2.40 mmol) were treated with p-TsOH‚

H₂O (40 mg, 0.21 mmol) to afford crude diester 31d. ¹H NMR

(DMSO-d₆): δ 12.35 (d, 1 H), 8.1 (s, 1 H), 7.0-7.5 (m, 5 H),

6.9 (d, 2 H), 6.8 (2 H), 6.7 (2 H), 4.1-4.4 (m, 2 H), 2.5 (s, 3 H),

1.1-1.3 (3 H).

4,6-Dichloro-3-(2-cyclohexyl-2-(methoxycarbonyl)vinyl)-

1H-indole-2-carboxylic Acid Ethyl Ester, 31g. Mukaiyama

coupling of aldehyde 6 (1.20 g, 4.19 mmol) with the ketene

silyl acetal 28g, prepared from methyl cyclohexylacetate 27g

(1.97 g, 12.6 mmol), as described in the general procedure III

gave alcohols 29g (0.62 g, 33%). Dehydration of alcohols 29g

(629 mg, 1.40 mmol) in toluene (10 mL) with p-TsOH‚H₂O (30

mg, 0.16 mmol) for 12 h gave, after recrystalization from hot

1

CH₂Cl₂, diester Z-31g (382 mg, 64%) as a yellow powder. H

NMR (CDCl₃): δ 9.23 (s, 1 H), 7.29 (s, 1 H), 7.16 (m, 1 H),

7.08 (d, 1 H), 4.35 (q, 2 H), 3.5 (s, 3 H), 1.6-2.05 (m, 5 H),

1.2-1.5 (m, 9 H).

3-(2-Carboxy-2-cyclohexylvinyl)-4,6-dichloro-1H-indole-

2-carboxylic Acid, 32g. Hydrolysis (vide supra) of diester

Z-31g (382 mg, 0.90 mmol) in THF (5 mL) using 1 M NaOH

(5.5 mL) gave Z-ester acid (250 mg, 71%) in which the methyl

ester of the side chain had remained unhydrolyzed. IR (KBr):

Hydrolysis of diester 31d (1.19 g, 2.32 mmol) provided 32d

as a yellow crystalline solid. IR (KBr): ν_max3418, 3283, 1686,

1

1613, 1489, 1240 cm^-1. H NMR (DMSO-d₆): δ 12.20-13.60

1

ν_max3300, 2928, 2855, 1694, 1556, 1535, 1248 cm^-1. H NMR

(bs, 2H), 12.20 (s, 1 H), 8.10 (s, 1 H), 7.35 (m, 4 H), 7.18 (s, 1

H), 7.09 (m, 1 H), 6.96 (d, 2 H, J ) 8.8 Hz), 6.82 (d, 2 H, J )

8.8 Hz), 6.72 (d, 2 H, J ) 8.8 Hz). MS: m/z 468 (M⁺+ 1), 450

(M⁺+ 1 - H₂O), 424. Anal. C₂₄H₁₅Cl₂NO₅‚0.9 C₄H₁₀O: C, H,

N.

(DMSO-d₆): δ 12.5 (br, 1 H), 12.02 (s, 1 H), 7.39 (d, 1 H, J )

1.8 Hz), 7.15 (d, 1 H, J ) 1.8 Hz), 7.06 (s, 1 H), 1.60-1.94 and

1

1.12-1.42 (m, 11 H). H NMR (DMSO-d₆+ D₂O): δ 3.34 (s,

-CO₂Me). MS: m/z 396 (M⁺+ 1), 364 (M⁺- MeOH), 336.

Anal. Calcd for C₁₉H₁₉Cl₂NO₄: C, 57.89; H, 4.86; N, 3.55.

Found; C, 56.50; H, 4.64; N, 3.35. Further hydrolysis of the

Z-ester acid as above for an additional 12 h gave, after

acidification and recrystallization from EtOAc/cyclohexane,

diacid Z-32g (192 mg, 55.8%) as white crystals. IR (KBr): ν_max

3279, 2930, 2857, 1696, 1559, 1451, 1248, 1203 cm^-1. ¹H NMR

(DMSO-d₆): δ 13.3-12.3 (br, 2 H), 12.05 (s, 1 H), 7.37 (d, 1 H,

J ) 1.8 Hz), 7.12 (d, 1 H, J ) 1.8 Hz), 6.93 (s, 1 H), 1.63-1.91

(m, 5 H), 1.10-1.42 (m, 6 H). MS (CI, NH₃): m/z 401, 399 (M⁺

3-(2-Carboxy-3-phenylpropenyl)-4,6-dichloro-1H-indole-

2-carboxylic Acid, 32e. The ketene silyl acetal of methyl

hydrocinnamate (28e) was prepared and coupled with 6 as

previously described to give 2.0 g (36%) of diester Z-31e as a

white solid: mp 150-150.5 °C. IR (KBr): ν_max3304, 3086,

3063, 3028, 2986, 2951, 2910, 2847, 1803, 1709, 1651, 1610,

1

1557 cm^-1. H NMR (CDCl₃): δ 9.36 (bs, 1 H), 7.32-7.41 (m,

4 H), 7.22-7.27 (m, 1 H), 7.14 (bt, 1 H, J ) 1.3 Hz), 7.04 (d, 1

H, J ) 1.7 Hz), 7.00 (d, 1 H, J ) 1.7 Hz), 4.27 (q, 2 H, J ) 7.2

+ NH₄, 100), 381 (M⁺+ NH₄

Cl₂NO₄: C, H, N.

H₂O), 355, 96. Anal. C₁₈H₁₇-

+

-

Hz), 3.89 (bs, 2 H), 3.57 (s, 3 H), 1.31 (t, 3 H, J ) 7.1 Hz). ¹³

C

NMR (CDCl₃): δ 167.5, 161.1, 138.8, 136.7, 134,9, 132.3, 130.5,

129.2, 128.4, 128.3, 126.4, 124.5, 122.3, 122.1, 118.3, 110.6,

77.3, 77.0, 76.7, 61.3, 51.5, 40.5, 14.2. MS: m/z 460 (M⁺+ 29),

432 (M⁺+ 1), 400 (100). Anal. C₂₂H₁₉Cl₂NO₄: C, H, N.

Hydrolysis of diester 31e (575 mg, 1.33 mmol) with 6 N

NaOH (2 mL) and heating at 80 °C for 1 h, followed by acidic

workup and recrystallization (acetone/H₂O), afforded diacid

Z-32e (484 mg, 93%) as a white solid: mp 244-244.5 °C (dec).

IR (KBr): ν_max3387, 3346, 3084, 3065, 3028, 2714, 2596, 1689,

4,6-Dichloro-3-(hydroxymethyl)-1-(toluene-4-sulfonyl)-

1H-indole-2-carboxylic Acid Ethyl Ester, 33. NaBH₄(1.10

g, 29 mmol) was added to a stirred solution of N-tosyl aldehyde

7 (12.3 g, 28 mmol) in THF (125 mL), and the reaction mixture

was stirred for 2 h at 35 °C. The THF was evaporated, and

H₂O (50 mL) was added to the residue. The H₂O layer was

extracted with EtOAc (2 × 80 mL), and the combined organic

layers were dried (Na₂SO₄) and concentrated in vacuo to give

alcohol 33 (10.7 g, 87%) as a white solid. ¹H NMR (DMSO-d₆):

δ 7.99 (s, 1 H), 7.85 (m, 2 H), 7.28 (m, 3 H), 4.95 (s, 2 H), 4.51

(q, 2 H, J ) 7.1 Hz), 2.39 (s, 3 H), 1.42 (t, 3 H, J ) 7.1 Hz).

4,6-Dichloro-3-(chloromethyl)-1-(toluene-4-sulfonyl)-

1H-indole-2-carboxylic Acid Ethyl Ester, 34. A mixture

of alcohol 33 (5.0 g, 11 mmol), SOCl₂(1.5 mL, 20 mmol), and

toluene (50 mL) was heated at reflux for 1 h. The reaction

mixture was concentrated in vacuo to give chloride 34 (5.2 g,

100%) as a dark brown solid, which was used without further

purification. ¹H NMR (DMSO-d₆): δ 7.82-7.98 (m, 3 H), 7.23-

7.37 (m, 3 H), 4.95 (s, 2 H), 4.52 (q, 2 H, J ) 7.2 Hz), 2.40 (s,

3 H), 1.45 (t, 3 H, J ) 7.1 Hz).

4,6-Dichloro-3-(2-(ethoxycarbonyl)-2-phenylethyl)-1H-

indole-2-carboxylic Acid Ethyl Ester, 35. To the crude

chloride 34 (5.2 g, 11 mmol) dissolved in THF (15 mL) was

added NaI (1.7 g, 11 mmol), and the reaction mixture was

stirred at rt under N₂for 4 h. In a separate reaction flask, the

anion of ethyl phenylacetate (6.0 g, 36 mmol) was generated

in THF (75 mL) by the addition of NaH (1.6 g, 36 mmol, 55%

dispersion in oil). After stirring for 3 h at 40 °C, the resulting

yellow suspension was cooled to -40 °C and the crude chloride

34/NaI/THF solution was added. The reaction mixture was

allowed to warm to rt and stirred for 18 h, after which it was

diluted with H₂O (150 mL), extracted with Et₂O (3 × 80 mL),

dried (Na₂SO₄), and concentrated in vacuo to give a crude

brown solid. Recrystallization from 9:1 hexane/EtOAc gave

dihydro diester 35 (1.5 g, 30%) as a white solid. ¹H NMR

(CDCl₃): δ 8.90 (br s, 1 H), 7.25 (d, 1 H, J ) 2 Hz), 7.14-7.20

(m, 5 H), 7.15 (d, 1 H, J ) 2 Hz), 3.98-4.40 (overlapping q, 4

1614, 1558 cm^-^{1 1}H NMR (DMSO-d₆): δ 12.66 (bs, 1.5 H),

.

12.14 (s, 1 H), 7.39 (d, 1 H, J ) 2.1 Hz), 7.24-7.37 (m, 4 H),

7.16-7.23 (m, 1 H), 7.20 (bs, 1 H), 7.14 (d, 1 H, J ) 1.8 Hz),

3.76 (s, 2 H). ¹³C NMR (DMSO-d₆): δ 167.6, 162.3, 139.5, 136.9,

135.1, 130.8, 128.8, 128.3, 128.1, 127.6, 126.3, 125.9, 122.3,

120.6, 117.5, 111.1, 40.2. MS: m/z 400 (M⁺+ 29), 390 (M⁺

+

1), 389 (M⁺), 390 (M⁺- H₂O, 100). Anal. C₁₉H₁₃Cl₂NO₄‚

0.2H₂O: C, H, N.

4,6-Dichloro-3-(2-(methoxycarbonyl)-1,2,3,4-tetrahy-

dronaphthalen-1-yl)-1H -indole-2-carboxylic Acid Ethyl

Ester, 31f. Ketene acetal 28f was generated from methyl

4-phenylbutyrate 27f (4.2 g, 23.6 mmol) and was reacted with

6 (3.7 g, 12.85 mmol) as described in the general procedure

III to afford 31f directly (2.0 g, 35%; ca. 1:1 mixture of

diastereomers) as a white solid: mp 165-166.5 °C. IR (KBr):

ν_max3331, 3099, 3061, 3018, 2992, 2982, 2949, 2865, 2843,

1718, 1697, 1610, 1556, 1523, 1489, 1437, 1369, 1350, 1319,

1

1280, 1236, 1172, 1114 cm^-1. H NMR (300 MHz, CDCl₃): δ

9.28 and 9.11 (two brs, 1 H), 7.25 and 7.22 (two d, 1H, J ) 2

Hz), 7.17 and 7.00 (two d, 1H, J ) 2 Hz), 6.89-7.14 (series of

m, 3H), 6.69 (m, 1H), 5.97 and 5.82 (two d, 1H, J ) 11 Hz),

4.45 and 4.10 (two AB-m, 2H), 3.59 (m, 1H), 3.49 and 3.47 (two

s, 3H), 2.92-3.22 (m, 2H), 2.06-2.36 (m, 2H), 1.42 and 1.02

(two t, 3H, J ) 7 Hz). ¹³C NMR (75 MHz, CDCl₃; multiplicities

derived from APT spectrum): δ 175.6 s, 175.3 s, 161.9 s, 160.3

s, 139.5 s, 139.1 s, 137.9 s, 136.3 s, 135.6 s, 135.5 s, 130.9 s,

130.6 s, 128.8 s, 128.4 d, 128.1 s, 128.0 d, 127.7 d, 127.6 d,

127.4 s, 126.2 s, 126.0 d, 125.9 d, 125.6 d, 125.3 d, 124.9 s,

123.9 s, 123.5 s, 123.2 d, 122.8 d, 122.3 s, 110.6 d, 110.4 d,

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1015

H), 3.98-4.15 (m, 2 H), 3.75-3.92 (m, 1 H), 1.41 (t, 3 H, J )

7 Hz), 1.16 (t, 3H, J ) 7 Hz).

When the assay was to be performed, the membranes were

thawed at ambient temperatue and diluted with 9 volumes of

10 mM HEPES-KOH, pH 7.4, and incubated at 25 °C for 15

min. This was followed by centrifugation at 44000g for 15 min

and then resuspension with 10 mM HEPES-KOH at pH 7.4

using a polytron. The incubation/resuspension process was

repeated an additional 2 times, and the final pellet was

resuspended in 6 volumes of 50 mM HEPES-KOH at pH 7.4.

Incubation vials, in triplicate, received 50 µL of 200 nM [³H]-

glycine, 50 µL of 100 nM strychnine, 50 µL of various

concentrations of test compounds diluted with 50 mM HEPES-

KOH at ph 7.4, and 200 µL of membrane suspension (400 µg

of protein/aliquot) in a final volume of 0.5 mL. Incubations

were carried out at 4 °C for 30 min and were terminated by

centrifugation at 46000g for 10 min. The supernatants were

decanted, and the pellets were rinsed rapidly with 2 mL of

ice-cold 50 mM HEPES-KOH at pH 7.4, then dissolved in 4

mL of Ready Protein (Beckman Instruments), and counted by

liquid scintillation spectrometry.

Specific binding of [³H]glycine is measured as the total

radioactivity bound minus that bound to the receptors in the

presence of 0.1 mM ^D-serine. Total membrane-bound radioac-

tivity is less that of the 2% added to the assay vials. Since

these conditions limit the total binding to less than 10% of

the radioactivity, the concentration of free ligand does not

change appreciably during the assay. The IC₅₀values reported

represent the molar concentration of a compound required to

reduce glycine binding by 50%, and were mean values from

two or more experiments; standard errors in all cases were

within 10-30% of the mean.

3-(2-Carboxy-2-phenylethyl)-4,6-dichloro-1H-indole-2-

carboxylic Acid, 36. The dihydro diester 35 (820 mg, 1.9

mmol) was heated at reflux in a mixture of KOH (700 mg, 10.6

mmol) and EtOH (50 mL). After 16 h, the EtOH was evapo-

rated and the residue acidified to pH 1 with aqueous HCl. The

precipitate formed was washed with H₂O and dried to provide

dihydro diacid 36 (600 mg, 86%) as a white solid. Precipitation

of the crude product from EtOH with 2-butanone gave the pure

dihydro diacid 36 (390 mg, 56%) as a white solid.

3-(2-Carboxy-2-phenylethyl)-4,6-dichloro-1H-indole-2-

carboxylic Acid, 36 from 29a via 37. To a stirred solution

of aldol 29a (395 mg of the less polar diastereomer, 0.905

mmol) in dry CH₂Cl₂(4 mL) under N₂was added Et₃SiH (200

µL, 1.25 mmol) followed by TFA (0.90 mL). After stirring for

16 h at rt, the reaction mixture was diluted with EtOAc/Et₂O,

washed with H₂O (2×), and concentrated in vacuo to give 700

mg of a light yellow solid. Recrystallization from cyclohexane/

trace EtOAc gave diester 37 (320 mg, 84%) as a white solid:

mp 160-164 °C (dec). IR (KBr): ν_max3325, 1734, 1715, 1701,

1676, 1321, 1240 cm^-1. ¹H NMR (CDCl₃): δ 8.88 (bs, 1 H), 7.24

(d, 1 H, J ) 1.7 Hz), 7.15-7.25 (m, 5 H), 7.13 (d, 1 H, J ) 1.7

Hz), 4.35 (dq, 1 H, J ) 11, 7 Hz), 4.23 (dq, 1 H, J ) 11, 7 Hz),

4.07-4.15 (m, 2 H), 3.79-3.88 (m, 1 H), 3.60 (s, 3 H), 1.40 (t,

3 H, J ) 7.1 Hz). MS: m/z 422, 421, 420 (M⁺+ 1), 419, 390,

388, 362, 360 (100), 272, 270. Anal. C₂₁H₁₉Cl₂NO₄: C, H, N.

Quinolinic Acid Seizures. One group of 10 mice (CD-1

mice) were administered 0.01-100 µg of test compound

intracerebroventricularly (icv) in a volume of 5 µL of saline.³⁵

A second control group containg an equal number of mice were

administered an equal volume of saline as a control. Ap-

proximately 5 min later, both groups were administered 7.7

µg of quinolinic acid icv in a volume of 5 µL of saline. This

dose of quinolinic acid was found to cause clonic-tonic seizures

in 90-100% of otherwise untreated mice. Immediately follow-

ing quinolinic acid administration, animals were observed for

the next 15 min for the occurrence of clonic-tonic seizures.

Those mice not convulsing during the observation period were

considered protected. The ED₅₀is defined as that dose that

protects 50% of the mice.

NMDA-Elicited Cyclic cGMP. The most potent com-

pounds, 11h and 19, were confirmed as antagonists by their

ability to inhibit NMDA-stimulated accumulation of cyclic

GMP. Similar assays were run with known NMDA antagonists

38 (CoCensys), 39 (Glaxo), and 40 (Merck) in order to evaluate

the affinity of the compounds for the glutamic acid binding

site of the NMDA receptor complex. NMDA-elicited cyclic GMP

(cGMP) was measured in 8-day old CD rat cerebellar slices.

Cystolic free Ca²⁺concentrations were measured in fur-2 AM-

loaded cultured rat cerebellar granule cells (8 days in vitro)

using microspectrofluorimetry.³⁵Na⁺and Ca²⁺currents were

measured by the whole-cell configuration of the patch-clamp

technique. Rat brain neurons (CD rat, 4-18 days in vitro) were

continually perfused at 0.5 mL/min at 22 °C in a HEPES

buffered extracellular saline solution consisting of the following

(mM): 142 NaCl; 1 CaCl₂; 8 KCl; 10 glucose; and 10 HEPES,

pH 7.4. Patch pipets were fabricated from Fisher brand

microhematocrit capillary tubes and filled with an intracellular

solution consisting of the following (mM): 153 CsCl₂; 10

HEPES; and 5 EGTA, pH 7.38. Test compounds were prepared

as 10 mM stock solutions in DMSO. Immediately prior to use,

appropriate concentrations were prepared by dilution with

external solution (DMSO final concentration ranged from 0.001

to 1.0%). NMDA (100 µM)/glycine (2 µM) and/or test com-

pounds were applied via a U-tube (rapid solution exchanger)

positioned within 200 µm of the neuron of interest. Via this

method a complete solution change in the vicinity of a cultured

neuron was achieved well within 500 ms.³⁶The relative

potencies of test compounds were assessed by comparison of

the response to NMDA (100 µM)/ glycine (2 µM) obtained in

Hydrolysis of diester 37 (320 mg, 0.76 mmol) in 5:1 THF/

H₂O (24 mL) using LiOH‚H₂O (95 mg, 2.3 mol) at reflux for 5

h gave 350 mg of yellow solid. Recrystallization from cyclo-

hexane/EtOAc gave 36 (280 mg, 74.0%) as fine ivory needles:

mp 246-248 °C (dec). IR (KBr): ν_max1699 cm^{-1 1}H NMR

.

(DMSO-d₆): δ 13.18 (br s, 1 H), 12.37 (br s, 1 H), 11.87 (s, 1

H), 7.35 (d, 1 H, J ) 1.8 Hz), 7.1-7.22 (m, 5 H), 7.15 (d, 1 H,

J ) 1.8 Hz), 3.98 (dd, 1 H, J ) 8.3, 7.1 Hz), 3.90 (dd, 1 H, J )

13.4, 7.0 Hz), 3.78 (dd, 1 H, J ) 13.4, 8.7 Hz). MS: m/z 379,

378 (M⁺+ 1), 377, 334, 332 (100), 244, 242. Anal. C₁₈H₁₃Cl₂-

NO₄: C, H, N.

Receptor Binding. Affinity for brain strychnine glycine

binding on the NMDA receptor complex were performed using

rat cortical and hippocampal membranes.^33,34Approximately

50-60 young male Sprague-Dawley rats (C-D strain) were

sacrificed by decapitation and their cerebral cortices and

hippocampi removed. The two brain regions were combined

and homogenized in 15 volumes of ice-cold 0.32 M sucrose

using Teflon glass homogenized (10 passes at 400 rpm). The

homogenates were centrifuged at 1000g for 10 min, and the

supernatants were transferred and recentrifuged at 44,000 x

gravity for 20 min. The upper white part of the pellets were

suspended with a pipet in ice-cold water and homogenized with

a polyron (setting 6 for 10 s) and centrifuged at 44000g for 15

min. Pellets were then suspended in 6 volumes of water and

placed in a dry ice/methanol bath until frozen, followed by

thawing at 37 °C in a shaking water bath. The freeze/thaw

process was repeated, and the final volumes of the suspension

were adjusted to 15 volumes with water and centrifuged at

44000g for 15 min. The resulting pellets were then suspended

in 15 volumes of 10 mM HEPES-KOH (N-2-(hydoxyethyl)-

piperazine-N′-2-ethanesulfonic acid-potassium hydroxide) at

pH 7.4 containing 0.04% Triton X-100 (v/v), incubated at 37

°C for 15 min and centrifuged at 44000g for 15 min. The pellets

were resuspended in 15 volumes of 10 mM HEPES-KOH at

pH 7.4 with a polytron (setting of 6 for 10 s) and centrifuged

at 44000g for 15 min. This resuspension/centrifugation process

was repeated an additional 2 times. The membranes were then

resuspended in 3 volumes of 10 mM HEPES and stored at -80

°C.

1016 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4

Baron et al.

the presence of the antagonist relative to that elicited by

NMDA/glycine alone. Recordings from 3 to 8 cells were

collected for each compound examined. All cells successfully

patched responded to NMDA/glycine.

clusion dosing. The dosing regimen was as in the previous

experiment (bolus + 6 h infusion) but was initiated 30 min

after the onset of ischemia. Compound 19 produced a dose-

dependent reduction in infarct volume. This was significant

at doses of 4.5 mg/kg bolus + 4.5 mg/kg/h × 6 h infusion (total

dose of 31.5 mg/kg) and above but not at the lowest dose (2.25

mg/kg bolus + 2.25 mg/kg/h × 6 h infusion). Therefore, the

minimal effective dose (MED) is 31.5 mg/kg given over 6 h. In

two separate experiments, drug concentrations were measured

in plasma samples from rats receiving the bolus + infusion

regimen corresponding to the MED. Plasma concentrations of

19 ranged from 21 to 25 µg/mL.

Harmaline-Elevated Cyclic GMP. Inhibition of harma-

line-elevated cyclic GMP accumulation was measured in the

cerebellum of CD-1 mice.³⁷Cyclic GMP content was normalized

to the amount of membrane protein. Rat brain (CD rat)

dihydroxyphenylalanine accumulation and extracellur dopam-

ine levels were assayed according to procedures reported.^38-40

Maximal Electroshock. Maximal electroshock-induced

seizure testing was performed using CDF rats.⁴¹Assessment

of anxiolytic potential was performed using the separation-

induced vocalization test in CD rat pups.^42,43Assessment of

potential ataxia was measured by disruption of rotorod

performance in CD-1 mice⁴¹or by the ability of a compound to

decrease the amount of time that an 8-day old CD rat pup

was able to retain its grasp on the lip of an ice bucket.⁴³Effects

of compounds on sensory gating were assessed by their

measured effect on prepulse inhibition of the startle reflex in

Wistar rats.⁴¹

Reduced infarct volume in rats using MCAo was determined

using male Sprague-Dawley rats.⁴⁵The rat is anesthetized

with halothane in a mixture of O₂and NO (1:2 ratio) and a

midline incision is made in the ventral neck region. An

indwelling venous catheter is placed in the jugular vein. Under

a dissecting microscope, the left common carotid artery is

identified at its bifurcation into the external carotid artery and

internal carotid artery. Two ties are placed on the external

carotid artery. The internal carotid artery is exposed distally

to the point of its bifurcation into the intracranial internal

carotid artery and the pterygopalatine artery. A small cut is

made in the distal segment of the external carotid artery and

a 3-0 nylon monofilament is introduced into the lumen of the

external carotid artery. The two previously placed ties are

tightened around the monofilament. The external carotid

artery is cut and reflected caudally so that the monofilament

can be advanced into the internal carotid artery, past the distal

internal carotid artery/pterygopalatine artery bifurcation and

continuing into the intranial segment of the internal carotid

artery to a distance of 20 nm, at which point the origin of the

middle cerebral artery is occluded. The ties are then tightened,

and the wound is closed. Compound or vehicle alone is

administered intravenously at a predetermined time postis-

chemia and dosing can be single, multiple, or continuous

infusion. Animals are given food and water and allowed to

survive for 24 h. Prior to sacrifice, the rat is weighed and given

a battery of four neurological tests to measure muscle strength,

grooming skills, postural reflexes, and sensorimotor integra-

tion.⁴⁶The animal is then decapitated, the brain removed,

sliced into six sections, and incubated in 2% 2,3,5-triphenyltet-

razolium chloride for 30 min.⁴⁷The area of infarction is clearly

visible. The infarct area is determined by computer-assisted

image analysis for each of the six sections and integrated over

the anterior-posterior extent of the brain to yield the infarct

volume.

DBA Audiogenic Seizure.⁴⁴Typically one group of six to

eight male DBA/2J audiogenic susceptible mice was admin-

istered 0.01-10.0 µg of the test compound, as a solution in

methylcellulose, into the lateral ventricle of the brain or from

0.1 mg to 300 mg intraperitoneally. A second group of mice

was administered an equal volume of a saline control by the

same route. A period of 5 min to 4 h later, the mice were placed

individually into a glass jar and exposed to a sound stimulus

of 110 dB (12 kHz) for 30 s. Each mouse was observed during

the sound exposure for signs of seizure activity. Mice not

displaying tonic hind limb extension were considered protected.

Animals treated with probenecid were coadministered pro-

benecid (100 mg/kg, ip) and test compound 1 h prior to the

seizure stimulus.

dMCAo and pMCAo Focal Cerebral Ischemia Mod-

els.^45,46Compound 19 was administered at three doses vs

saline vehicle in the dMCAo (distal middle cerebral artery

occlusion) model. Drug administration was begun with a bolus

dose given 15 min prior to the onset of ischemia and main-

tained by continuous infusion. The highest dose of 19 (70 mg/

kg bolus + 70 mg/kg/h infusion for 6 h) showed significant

neuroprotection reducing infarct volume by 32%. While the

dose of 35 mg/kg + 35 mg/kg/h for 6 h demonstrated a trend

toward neuroprotection (12% reduction of infarct volume), it

did not reach statistical significance. This experiment validates

the concept that a molecule of this structural type can provide

significant infarct volume reduction in a severe ischemia

model. These results were used to establish the upper dose

level for postocclusion testing in the dMCAo and the ischemia-

reperfusion pMCAo (proximal middle cerebral artery occlusion)

models and serve as a starting point for dose optimization.

A second study employed poststroke administration of 19

in the dMCAo model. Drug treatment was initiated 30 min

after the onset of ischemia. A dosing regimen of 35 mg/kg bolus

+ 25 mg/kg/h infusion for 4 h was used and resulted in a 28%

reduction in infarct volume, which is similar to that achieved

with preischemia dosing.

Animals. Adult male mice (Charles River CD-1), alult male

rats (Charles River CD, Wistar, or CDF), or 18 day old, male

audiogenic seizure mice (DBA/2J Jackson Laboratories) were

used for the in vivo studies. Neonatal (Charles River CD) rats

were used for cell culture and brain slice experiments. Animals

were housed in animal rooms on a 14 h light/dark cycle and

allowed free access to food and water. Neonatal rats were

housed with the mother in a breeding box with sawdust

bedding. All experiments were performed during the light

phase of the light/dark cycle. Drugs were dissolved in distilled

water, saline, or 50 mM Tris base and injected at a volume of

1 mL/kg (rats) or 10 mL/kg (mice).

Antagonists 19 and 11h and all competitor compounds were

administered iv, the desired route of administration for acute

stroke treatment. Anticonvulsant experiments in mice and rats

indicated that the half-life of effect for 19 and 11h was

approximately 30 min to 1 h. Therefore, in the stroke experi-

ments, sustained administration would be required to provide

NMDA antagonism during the period of excitotoxic cell death.

Bolus doses of 19 and 11h were chosen to be within the range

necessary for anticonvulsant activity measured with the rat

maximal electroshock test. Preliminary pharmacokinetic in-

formation was obtained to define the appropriate values for

infusion doses. This allowed maintenance of steady-state

plasma drug concentrations, providing a sustained pharma-

cological effect.

Data Analysis. Concentration-response data were fit to a

logistic function using nonlinear regression analysis. Antagonist-

induced responses (e.g., NMDA-stimulated cGMP formation)

used a function of the following form: response ) B + MXⁿ/

(Kⁿ+ Xⁿ), where B ) observed basal response, M ) calculated

maximal response, X ) concentration of agonist, n ) slope

factor, and K ) calculated EC₅₀of the agonist. Antagonist-

induced inhibition of responses was fit using a similar model

of the following form: inhibition ) 100Xⁿ/(Kⁿ+ Xⁿ), where X

) concentration of antagonist, n ) slope factor, and K ) IC₅₀

of the antagonist. Dose-response data were fit using linear

regression of log-transformed values.

Acknowledgment. The authors wish to thank the

Analytical and Structural Sciences department for their

Following this initial demonstration of efficacy in the

pMCAo model, the results were extended to include postoc-

Potent Glycine-Site NMDA Receptor Antagonist

Journal of Medicinal Chemistry, 2005, Vol. 48, No. 4 1017

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assistance in characterization and analysis of the de-

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Supporting Information Available: Results from el-

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1549.

JM0491849

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CoMFA, synthesis, and pharmacological evaluation of (E)-3-(2-carboxy-2- arylvinyl)-4,6-dichloro-1H-indole-2-carboxylic acids: 3-[2-(3-Aminophenyl)-2- carboxyvinyl]-4,6-dichloro-1H-indole-2-carboxylic acid, a potent selective glycine-site NMDA receptor antagonist

DOI: 10.1021/jm0491849

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