Synthesis and antiprotozoal activity of novel bis-benzamidino imidazo[1,2-a]pyridines and 5,6,7,8-tetrahydro-imidazo[1,2-a]pyridines

The key dinitrile intermediates 4a-d were synthesized by reaction of phenacyl bromide 1 and the appropriate 2-amino-5-bromopyridines to yield 3a-d. Suzuki coupling of 3a-d with 4-cyanophenylboronic acid yielded the 2,6-bis(4-cyanophenyl)-imidazo[1,2-a]pyridine derivatives 4a-d. The bis-amidoximes 5a-d, obtained from 4a-d by the action of hydroxylamine, were converted to the bis-O-acetoxyamidoximes which on catalytic hydrogenation in a mixture of ethanol/ethyl acetate gave the acetate salts of 2,6-bis[4-(amidinophenyl)]-imidazo[1,2-a]pyridines 7a-d. In contrast, catalytic hydrogenation of the bis-O-acetoxyamidoxime of 5a in glacial acetic acid gave the saturated analogue 2,6-bis[4-(amidinophenyl)]-5,6,7,8-tetrahydro-imidazo[1,2-a]pyridine 8. O-Methylation of the amidoximes 5a-d gave the N-methoxyamidines 6a-d. The diamidines showed strong DNA binding affinity, were very active in vitro against T. b. r. exhibiting IC(50) values between 7 and 38nM, but were less effective against P. f. with IC(50) values between 23 and 92nM. Two of the diamidines 7c and 7d were slightly more active than furamidine but less active than azafuramidine in the T. b. r. STIB900 mouse model. Only one prodrug 6b showed moderate activity in the same mouse model.     

Evaluation of cell permeation of a potent 5alpha-reductase inhibitor using MALDI-TOF MS

N-(Dicyclohexyl)acetyl-piperidine-4-benzylidene-4-carboxylic acid (1), although a very potent in vitro 5alpha-steroid reductase (5alphaR) type 2 inhibitor, showed only marginal in vivo activity in rats. Since this could be due to hindered cellular uptake of the carboxylic acid, acid (1) and its corresponding methyl ester (1a) were compared with respect to their permeation properties. In the parallel artificial membrane permeation assay (PAMPA), 1a showed a higher %flux of 55 versus 6 for 1. Considering the high potency of 1 and better permeation of 1a, the use of 1a as a prodrug for 1 was explored using the human prostate carcinoma cell line DU145. Esterase activity, a prerequisite for this prodrug concept was detected employing 4-nitrophenyl acetate (4-NPA) as a substrate. After incubation of DU145 cells with 1 and 1a, respectively, permeated 1a and its hydrolysis to 1 were unequivocally observed by MALDI-TOF MS analyses, whereas 1 could not be detected inside the cells above the detection limit. Regarding biological activity, 1a showed a stronger inhibition of 5alphaR in intact DU145 cells than 1 (IC50 values, 4 microM and > 10 microM for 1a and 1, respectively). These results suggest that the in vivo activity of 1 might be increased by the use of its methyl ester prodrug 1a.     

Synthesis and activity of a folate peptide camptothecin prodrug

A folate receptor targeted camptothecin prodrug was synthesized using a hydrophilic peptide spacer linked to folate via a releasable disulfide carbonate linker. The conjugate was found to possess high affinity for folate receptor-expressing cells and inhibited cell proliferation in human KB cells with an IC(50) of 10nM. Activity of the prodrug was completely blocked by excess folic acid, demonstrating receptor-mediated uptake.     

Aromatic esters of progesterone as 5alpha-reductase and prostate growth inhibitors

The aim of this study was to determine the biological activity of 4 steroidal derivatives (9a, 9b and 10a, 10b) prepared from the commercially available 17alpha acetoxyprogesterone, where 9a, 9b, have the Delta(4)-3-oxo structure and 10a and 10b an epoxy group at C-4 and C-5. These steroids were tested as inhibitors of 5alpha-reductase enzyme, which is present in androgen-dependent tissues and converts testosterone to its more active reduced metabolite dihydrotestosterone. The pharmacological effect of these steroids was demonstrated by the significant decrease of the weight of the prostate gland of gonadectomized hamsters treated with testosterone plus finasteride or with steroids 10a and 10b. For the studies in vitro the IC(50) values were determined by measuring the steroid concentration that inhibits 50% of the activity of-5alpha-reductase. In this study we also determined the capacity of these steroids to bind to the androgen receptor present in the rat prostate cytosol. The results from this work indicated that compounds 9a, 9b, 10a, and 10b inhibited the 5alpha reductase activity with IC(50) values of 360, 370, 13 and 4.9 nM respectively. However these steroids did not bind to the androgen receptors since none competed with labeled mibolerone. Steroid 10b, an epoxy steroidal derivative containing bromine atom in the ester moiety, was the most active inhibitor of 5alpha-reductase enzyme, present in human prostate homogenates with an IC(50) value of 4.9 nM and also showed in vivo pharmacological activity since it decreased the weight of the prostate from hamsters treated with testosterone in a similar way as finasteride.     

Some new acyclic nucleotide analogues as antiviral prodrugs: synthesis and bioactivities in vitro

A series of ester analogues of acyclic nucleotide PMPA and PMEA were synthesized as potent antiviral agents. The antiviral evaluation results indicated that bis benzyl ester prodrug of PMPA 5f and bis allyl ester prodrug of PMEA 5g exhibited potent antiviral activities. The IC(50) of 5f for HBV was 2.15 microM, and the IC(50) of 5g for HIV-1 was 1.61 microM.     

Studies of the inhibitory activity of MK-0591 (3-[1-(4-chlorobenzyl)-3-(t-butylthio)-5-(quinolin-2-yl-methoxy)-indol- 2-yl]-2,2-dimethyl propanoic acid) on arachidonic acid metabolism in human phagocytes.

We have investigated the inhibitory activity of compound MK-0591 (3-[1-(4-chlorobenzyl)-3-(t-butylthio)-5-(quinolin-2-yl-methoxy)-i ndol-2- yl]-2,2-dimethyl propanoic acid) on 5-lipoxygenase (5-LO) product synthesis in various human phagocytes stimulated with either the ionophore A23187, opsonized zymosan (OPZ), platelet-activating factor (PAF), or formyl-methionyl-leucyl-phenylalanine (fMLP). The lipoxygenase products were analyzed by reversed-phase HPLC. MK-0591 inhibited the formation of 5-hydroxyeicosatetraenoic acid, leukotriene (LT) B4, its omega-oxidation products, and 6-trans-isomers with IC50 values of 2.8-4.8 nM in A23187-stimulated neutrophils. In these conditions, arachidonic acid at a concentration of 10 microM had no effect on MK-0591 inhibitory activity. In neutrophils stimulated with OPZ, the synthesis of LTB4, its omega-oxidation products, and 6-trans-isomers was inhibited with IC50 values of 9.5-11.0 nM. MK-0591 inhibited 5-LO product synthesis in A23187-stimulated blood monocytes, eosinophils, and alveolar macrophages with IC50 values of 0.3-0.9, 3.7-5.3, and 8.5-17.3 nM, respectively. In neutrophils primed with granulocyte--macrophage colony-stimulating factor and stimulated with PAF, lipoxygenase product synthesis was inhibited with IC50 values of 7.7-8.7 nM. At the concentration of 1 microM, MK-0591 had no inhibitory effect on 15-lipoxygenase activity in human polymorphonuclear leukocytes, nor on human platelet 12-lipoxygenase and cyclooxygenase. In conclusion, MK-0591 is a very potent and specific inhibitor of 5-LO product synthesis in various types of human phagocytes.     

Synthesis and evaluation of novel 8,5-fused bicyclic peptidomimetic compounds as interleukin-1beta converting enzyme (ICE) inhibitors

An 8,5-fused bicyclic peptidomimetic ring system generated by a stereoselective ring metathesis reaction was elaborated into potent inhibitors of interleukin-1beta converting enzyme (ICE, caspase-1). Multiple compounds were found that exhibited ICE IC50 values < 10 nM and were selective over caspase-3 and caspase-8. These active analogs generally possessed good activity (IC50 values < 100 nM) in a whole cell assay measuring IL-1beta production. Pharmacokinetic analysis of the ethyl acetal prodrug form of a selected active lead revealed a compound with a reasonable plasma half-life (1.1 h) and good oral bioavailability (30%).     

Angiotensin I converting enzyme inhibitors containing unnatural alpha-amino acid analogues of phenylalanine

The activity of three angiotensin I converting enzyme (ACE) inhibitors with unique related structures was assessed in vitro and in vivo. The three compounds were (S)(-)-1,2,3,4-tetrahydro-2-(3-mercapto-1-oxopropyl)-3-isoquinoline carboxylic acid (EU-4865), 1,2,3,4-tetrahydro-2-(3-mercapto-1-oxopropyl)-1- isoquinolinecarboxylic acid (EU-4881), and (S)(-)-1,2,3,4-tetrahydro-1-(3-mercapto-1-oxopropyl)-2- quinolinecarboxylic acid (EU-5031). In vitro EU-4881 was a competitive inhibitor that lacked potency (IC50 = 1980 nM) against purified ACE. The other two compounds were equipotent (IC50 = 41 nM) against purified ACE but differed in their inhibition kinetics. EU-4865 (Ki = 38 nM) was a noncompetitive inhibitor, and EU-5031 (Ki = 6.9 nM) was a competitive inhibitor. Against caveolae membrane-bound ACE EU-4881 also lacked potency (IC50 = 2852 nM). In vivo in the conscious acute aortic coarctate (AAC) rat it also lacked potency, having an ED30 (oral dose decreasing blood pressure 30 mmHg) greater than 100 mg/kg. The activity of EU-4865 and EU-5031 in the caveolae membrane-bound ACE and AAC rat reflected their different Ki values rather than their similar IC50 values. In vitro, EU-4865 and EU-5031 had IC50 values of 19 and 6.7 nM, respectively, and in vivo, they had ED30 values of 52 and 1.1 mg/kg, respectively. These results suggest that ACE has a binding requirement for a carboxy-terminus, hydrophobic amino acid that is important for in vivo activity.     

Synthesis of poly(ethylene glycol)-based saquinavir prodrug conjugates and assessment of release and anti-HIV-1 bioactivity using a novel protease inhibition assay

Various poly(ethylene glycol)(PEG)-based prodrug conjugates of the HIV-1 protease inhibitor (PI) saquinavir (SQV) were prepared using several types of chemical groups potentially capable of modifying its pharmacokinetic properties. These prodrug conjugates included SQV-cysteine-PEG3400, SQV-cysteine-PEG3400-biotin, SQV-cysteine(R.I.CK-Tat9) [a cationic retro-inverso-cysteine-lysine-Tat nonapeptide]-PEG3400, and SQV-cysteine(R.I.CK(stearate)-Tat9)-PEG3400. SQV was linked to cysteine to form a releasable SQV-cysteine ester bond in all of the conjugates. The amino group of the cysteine moiety provided an attachment site for a slower-degrading amide bond with N-hydroxysuccinimide-activated forms of PEG- and PEG-biotin. Disulfide bonds were used to attach the cationic peptides, R.I.CK-Tat9 and R.I.CK(stearate)-Tat9 to the cysteine moiety in order to provide cell-specific release. An assay was established and validated for measuring the activity of SQV and other protease inhibitors in biological samples. In this assay, cleavage of an internally quenched fluorescent substrate, Arg-Glu(EDANS)-Ser-Gln-Asn-Tyr-Pro-Ile-Val-Gly-Lys(DABCYL)-Arg by HIV-1 protease was inhibited by SQV in a dose-dependent manner at concentrations of 0.05-0.5 microM. All prodrug conjugates were shown to be inactive in this assay until the ester bond was cleaved and active SQV was released. The prodrug reconversion half-lives in 0.1 N HCl, phosphate-buffered saline (PBS) at pH 7.4 and in spiked plasma at 37 degrees C were 9, 14, and 0.9 h, respectively. The anti-HIV-1 activity (ED(50)) of the PEG-based SQV prodrug conjugates was evaluated in MT-2 cells using an MTT assay. The activity of conjugated SQV was reduced (ED(50) = 900 nM) for the PEG only conjugate, but restored with the addition of biotin (ED(50) = 125 nM), R.I.CK-Tat9 (ED(50) = 15 nM), and R.I.CK(stearate)-Tat9 (ED(50) = 62 nM) as compared to maximum achievable anti-HIV-1 activity (unconjugated SQV, control, ED(50) = 15 nM), suggesting enhanced cellular uptake of conjugates. Cytotoxicity (LD(50)) was assessed for all prodrug conjugates using non-HIV-1 infected cells and was found to be in the micromolar range. The difference between the LD(50) and ED(50) suggests a favorable therapeutic index for the prodrug conjugates. In conclusion, these promising initial results demonstrate that the reconversion of the conjugate prodrugs was complete and that active SQV was released. Since the major delivery advantages of PEG prodrug conjugates can only be observed in vivo, issues of reconversion and elimination half-lives in plasma will have to be further studied in an in vivo model. The current results also demonstrate that the protease inhibition assay is a simple yet effective bioanalytical tool that can be used to assess the release and anti-HIV-1 activity of HIV-1 PIs from their prodrug forms.     

Synthesis and biological activity of potent heterocyclic thiol-based inhibitors of endothelin-converting enzyme-1

Directed screening of metalloprotease inhibitors identified CGS 30084 (1) as a potent inhibitor of endothelin-converting enzyme-1 (ECE-1) in vitro (IC(50)=77 nM). Herein we report the syntheses and biological activities of analogues containing modified biphenyl moieties, bearing heterocyclic proximal rings. Compound 20, the thioacetate ethyl ester prodrug derivative of compound 19a, was found to be an orally active and potent inhibitor of ECE-1 activity in rats.     

Calyculin A and okadaic acid: inhibitors of protein phosphatase activity

Calyculin A and okadaic acid induce contraction in smooth muscle fibers. Okadaic acid is an inhibitor of phosphatase activity and the aims of this study were to determine if calyculin A also inhibits phosphatase and to screen effects of both compounds on various phosphatases. Neither compound inhibited acid or alkaline phosphatases, nor the phosphotyrosine protein phosphatase. Both compounds were potent inhibitors of the catalytic subunit of type-2A phosphatase, with IC50 values of 0.5 to 1 nM. With the catalytic subunit of protein phosphatase type-1, calyculin A was a more effective inhibitor than okadaic acid, IC50 values for calyculin A were about 2 nM and for okadaic acid between 60 and 500 nM. The endogenous phosphatase of smooth muscle myosin B was inhibited by both compounds with IC50 values of 0.3 to 0.7 nM and 15 to 70 nM, for calyculin A and okadaic acid, respectively. The partially purified catalytic subunit from myosin B had IC50 values of 0.7 and 200 nM for calyculin A and okadaic acid, respectively. The pattern of inhibition for the phosphatase in myosin B therefore is similar to that of the type-1 enzyme.     

Prodrugs of 3-(3,4-dichlorobenzyloxy)-2-amino-6-fluorobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (MGS0039): a potent and orally active group II mGluR antagonist with antidepressant-like potential

3-(3,4-Dichlorobenzyloxy)-2-amino-6-fluorobicyclo[3.1.0]hexane-2,6-dicarboxylic acid 5 (MGS0039) is a highly selective and potent group II metabotropic glutamate receptor (mGluR) antagonist (antagonist activities for mGluR2; IC50=20.0 nM, mGluR3; IC50=24.0 nM) and is detected in both plasma (492 ng/mL) and brain (13.2 ng/g) at oral administration of 10 ng/mL [J. Med. Chem.2004, 47, 4750], but the oral bioavailability of 5 was 10.9%. In order to improve the oral bioavailability of 5, prodrugs of 5 were discovered by esterification of carboxyl group on C6-position of bicyclo[3.1.0]hexane ring. Among these compounds, 6-alkyl esters exhibited approximately 10-fold higher concentrations of 5 in the plasma and brain of rats after oral administration (e.g., ethyl ester of 5; plasma, Cmax=20.7+/-1.3 microM) compared to oral administration of 5 (plasma, Cmax=2.46+/-0.62 microM). 3-(3,4-Dichlorobenzyloxy)-2-amino-6-fluorobicyclo[3.1.0]hexane-2,6-dicarboxylic acid 6-heptyl ester (7ao), a prodrug of MGS0039, showed antidepressant-like effects in rat forced swimming test and mouse tail suspension test following oral administration. Moreover, following oral administration of 7ao in mice, high concentrations of MGS0039 were detected in both the brain and plasma, while 7ao was barely detected. In this paper, we report the synthesis, in vitro metabolic stabilities, and pharmacokinetic profiles of the prodrugs of 5, and the antidepressant-like effects of 7ao.     

Oxidative stimuli-responsive nanoprodrug of camptothecin kills glioblastoma cells

The purpose of this study was to prepare and characterize nanometer-sized prodrug (nanoprodrug) of camptothecin. The camptothecin prodrug was synthesized using tetraethylene glycol spacer linked via carbonate bond to camptothecin and via ester bond to α-lipoic acid. The nanoprodrug was prepared through the spontaneous emulsification mechanism using the mixture of camptothecin prodrug and α-tocopherol which served as a structural matrix. The nanoprodrug was activated readily by porcine liver esterase and, with a much slower rate, by hydrolytic degradation. Upon longterm storage, the α-lipoic acid moiety of the camptothecin prodrug gradually oxidized without loss of structural integrity and therapeutic efficacy. Interestingly, the hydrolytic activation was negligible before the oxidation, but was significantly accelerated after the oxidation of the α-lipoic acid moiety, suggesting an oxidative stimuli-responsive activation of the prodrug. The camptothecin nanoprodrug was found to possess significant inhibitory effect on the proliferation of U87-MG glioma cells with an IC₅₀ of 20 nM.     

Synthesis and biological activity of novel potent endothelin-converting enzyme-1 inhibitors

Through directed screening of metalloprotease inhibitors, CGS 30084 (1) has been identified as a potent endothelin-converting enzyme-1 (ECE-1) inhibitor in vitro (IC50 = 77 nM). Herein we report the syntheses and biological activities of analogues derived from this lead, based on modifications of the biphenyl moiety. Compound 10, the thioacetate methyl ester prodrug derivative of compound 6m, was found to be an orally active and potent inhibitor of ECE-1 activity in rats.     

Dicationic DNA-targeted antiprotozoal agents: naphthalene replacement of benzimidazole

A series of naphthalene analogues of highly active benzimidazole diamidines were synthesized using sequential Stille and Suzuki coupling reactions for preparation of the bis-nitrile intermediates. All of the diamidines showed strong DNA affinities as judged by high DeltaTm values with poly(dA-dT). The dicationic compounds were quite active in vitro versus Trypanosoma brucei rhodesiense (T. b. r.) exhibiting IC50 values ranging from 4 to 98 nM. These compounds were also active versus Plasmodium falciparum (P. f.) giving IC50 values ranging from 4 to 33 nM. Two of the compounds showed good activity in vivo in the STIB900 model for acute African trypanosomiasis; one gave 3/4 cures and the other gave 4/4 cures on ip dosage of 20 mg/kg for 4 days. The amidoxime prodrugs of the naphthalene analogues were essentially ineffective.     

Identified a morpholinyl-4-piperidinylacetic acid derivative as a potent oral active VLA-4 antagonist

An investigation into the structure-activity relationship of a lead compound, prolyl-5-aminopentanoic acid 4, led to the identification of a novel series of 4-piperidinylacetic acid, 1-piperazinylacetic acid, and 4-aminobenzoic acid derivatives as potent VLA-4 antagonists with low nanomolar IC(50) values. A representative compound morpholinyl-4-piperidinylacetic acid derivative (13d: IC(50)=4.4 nM) showed efficacy in the Ascaris-antigen sensitized murine airway inflammation model by oral administration.     

Pharmacology of the leukotriene antagonist verlukast: the (R)-enantiomer of MK-571.

Verlukast (MK-679) (3-[(3-(2-(7-chloro-2-quinolinyl)-(E)-ethenyl)phenyl)[3-(dimethylamino)- 3- oxopropyl)thio)methyl)-thio)propionic acid) is a potent and selective inhibitor of [3H]leukotriene D4 binding in guinea-pig (IC50 = 3.1 +/- 0.5 nM) and human (IC50 = 8.0 +/- 3.0 nM) lung homogenates and dimethyl sulfoxide differentiated U937 cell membrane preparations (IC50 = 10.7 +/- 1.6 nM) but is essentially inactive versus [3H]leukotriene C4 binding in guinea-pig lung homogenates (IC50 values of 19 and 33 microM). Functionally, when tested at 60 nM, it antagonized contractions of guinea-pig trachea (GPT) induced by leukotriene C4, leukotriene D4, and leukotriene E4 (respective-log KB values of 8.6, 8.8, and 8.9) and contractions of human trachea (HT) induced by leukotriene D4 (-log KB value 8.3 +/- 0.2). In contrast, verlukast (20-200 nM) failed to antagonize contractions of GPT induced by leukotriene C4 in the presence of 45 mM L-serine borate. Intravenous (i.v.) and aerosol verlukast antagonized bronchoconstriction (BC) induced in anaesthetized guinea pigs by i.v. leukotriene D4 but did not block BC to arachidonic acid or histamine. Intraduodenal verlukast (0.25 mg/kg) antagonized leukotriene D4 (0.2 micrograms/kg) induced BC in guinea pigs. Oral and aerosol administration blocked leukotriene D4-induced BC in conscious squirrel monkeys. Orally administered compound also blocked ovalbumin-induced BC in conscious sensitized rats treated with methysergide (3 micrograms/kg). The pharmacological profile for verlukast is similar to that of the racemic compound, MK-571. Verlukast is currently in clinical development for the treatment of asthma and related diseases.     

Discovery of a novel CCR3 selective antagonist

In searching for a novel CCR3 receptor antagonist, we designed a library that included a variety of carboxamide derivatives based on the structure of our potent antagonists for human CCR1 and CCR3 receptors, and screened the new compounds for inhibitory activity against 125I-Eotaxin binding to human CCR3 receptors expressed in CHO cells. Among them, two 2-(benzothiazolethio)acetamide derivatives (1a and 2a) showed binding affinities with IC50 values of 750 and 1000 nM, respectively, for human CCR3 receptors. These compounds (1a and 2a) also possessed weak binding affinities for human CCR1 receptors. We selected la as a lead compound for derivatization to improve in vitro potency and selectivity for CCR3 over CCRI receptors. Derivatization of la by incorporating substituents into each benzene ring of the benzothiazole and piperidine side chain resulted in the discovery of a compound (1b) exhibiting 820-fold selectivity for CCR3 receptors (IC50 = 2.3 nM) over CCR1 receptors (IC50 = 1900 nM). This compound (1b) also showed potent functional antagonist activity for inhibiting Eotaxin (IC50 = 27 nM)- or RANTES (IC50 = 13 nM)-induced Ca2+ increases in eosinophils.     

Synthesis and inhibitory activity of acyl-peptidyl-pyrrolidine derivatives toward post-proline cleaving enzyme; a study of subsite specificity.

Several pyrrolidine derivatives have been synthesized and examined for their inhibitory activity on post-proline cleaving enzymes from Flavobacterium meningosepticum and bovine brain. Almost all the compounds tested in this study inhibited the activity of both enzymes at low IC50 values (from nM to microM) but a specificity difference was observed with alkylacyl-peptidyl-pyrrolidine derivatives which strongly inhibited only the bacterial enzyme. The most effective inhibitors have a proline residue on their P2 sites and a substituted or unsubstituted phenoxybutyryl moiety on their P3 sites. Thus phenoxybutyryl-prolyl-pyrrolidine is the most effective partial structure of the inhibitors. The best inhibitors found were: 4-(4-benzylphenoxy)butyryl-prolyl-pyrrolidine for bacterial enzyme (IC50 1.4 nM) and 4-phenylbutyryl-thioprolyl-pyrrolidine for bovine brain enzyme (IC50 67 nM). In the passive avoidance test, using amnesic rats experimentally induced with scopolamine, the pyrrolidine derivatives which had potent inhibitory activity toward post-proline cleaving enzymes also showed strong anti-amnesic activities at doses of 1-5 mg/kg, i.p.     

Abietane diterpenoids and triterpenoic acids from Salvia cilicica and their antileishmanial activities

Bioguided-fractionation of an acetone extract of the roots of Salvia cilicica (Lamiaceae) led to isolation of two new diterpenes, 7-hydroxy-12-methoxy-20-nor-abieta-1,5(10),7,9,12-pentaen-6,14-dione and abieta-8,12-dien-11,14-dione (12-deoxy-royleanone), together with oleanolic acid, ursolic acid, ferruginol, inuroyoleanol and cryptanol. Their structures were determined spectroscopically, which included HREIMS and 2D NMR spectroscopic analysis. The new abietane derivatives showed appreciable in vitro antileishmanial activity against intracellular amastigote forms of both Leishmania donovani (IC(50) values of 170 and 120 nM, respectively) and Leishmania major (IC(50) values of 290 and 180 nM, respectively). The triterpenoic acids were found to be potently active against amastigote (IC(50) values of 7-120 nM) and moderately active against promastigote stages (IC(50) values of 51-137 nM) of the two Leishmania species.