Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of New Neocryptolepine Derivatives
Identifieur interne : 000797 ( Istex/Corpus ); précédent : 000796; suivant : 000798Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of New Neocryptolepine Derivatives
Auteurs : Tim H. M. Jonckers ; Sabine Van Miert ; Kanyanga Cimanga ; Christian Bailly ; Pierre Colson ; Marie-Claire De Pauw-Gillet ; Hilde Van Den Heuvel ; Magda Claeys ; Filip Lemière ; Eddy L. Esmans ; Jef Rozenski ; Ludo Quirijnen ; Louis Maes ; Roger Dommisse ; Guy L. F. Lemière ; Arnold Vlietinck ; Luc PietersSource :
- Journal of Medicinal Chemistry [ 0022-2623 ] ; 2002.
Abstract
On the basis of the original lead neocryptolepine or 5-methyl-5H-indolo[2,3-b]quinoline, an alkaloid from Cryptolepis sanguinolenta, derivatives were prepared using a biradical cyclization methodology. Starting from easily accessible educts, this approach allowed the synthesis of hitherto unknown compounds with a varied substitution pattern. As a result of steric hindrance, preferential formation of the 3-substituted isomers over the 1-substituted isomers was observed when cyclizing N-(3-substituted-phenyl)-N‘-[2-(2-trimethylsilylethynyl)phenyl]carbodiimides. All compounds were evaluated for their activity against chloroquine-sensitive as well as chloroquine-resistant Plasmodium falciparum strains, for their activity against Trypanosoma brucei and T. cruzi, and for their cytotoxicity on human MRC-5 cells. Mechanisms of action were investigated by testing heme complexation using ESI-MS, inhibition of β-hematin formation, DNA interactions (DNA−methyl green assay and linear dichroism), and inhibition of human topoisomerase II. Neocryptolepine derivatives with a higher antiplasmodial activity and a lower cytotoxicity than the original lead have been obtained. This selective antiplasmodial activity was associated with inhibition of β-hematin formation. 2-Bromoneocryptolepine was the most selective compound with an IC50 value against chloroquine-resistant P. falciparum of 4.0 μM in the absence of cytotoxicity (IC50 > 32 μM). Although cryptolepine, a known lead for antimalarials also originally isolated from Cryptolepis sanguinolenta, was more active (IC50 = 2.0 μM), 2-bromoneocryptolepine showed a low affinity for DNA and no inhibition of human topoisomerase II, in contrast to cryptolepine. Although some neocryptolepine derivatives showed a higher antiplasmodial activity than 2-bromocryptolepine, these compounds also showed a higher affinity for DNA and/or a more pronounced cytotoxicity. Therefore, 2-bromoneocryptolepine is considered as the most promising lead from the present work for new antimalarial agents. In addition, 2-bromo-, 2-nitro-, and 2-methoxy-9-cyanoneocryptolepine exhibited antitrypanosomal activity in the micromolar range in the absence of obvious cytotoxicity.
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DOI: 10.1021/jm011102i
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<record><TEI wicri:istexFullTextTei="biblStruct"><teiHeader><fileDesc><titleStmt><title>Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of New Neocryptolepine Derivatives</title><author><name sortKey="Jonckers, Tim H M" sort="Jonckers, Tim H M" uniqKey="Jonckers T" first="Tim H. M." last="Jonckers">Tim H. M. Jonckers</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation><affiliation><mods:affiliation> These authors contributed equally to this work.</mods:affiliation></affiliation></author><author><name sortKey="Van Miert, Sabine" sort="Van Miert, Sabine" uniqKey="Van Miert S" first="Sabine" last="Van Miert">Sabine Van Miert</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> These authors contributed equally to this work.</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Cimanga, Kanyanga" sort="Cimanga, Kanyanga" uniqKey="Cimanga K" first="Kanyanga" last="Cimanga">Kanyanga Cimanga</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Bailly, Christian" sort="Bailly, Christian" uniqKey="Bailly C" first="Christian" last="Bailly">Christian Bailly</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> INSERM U-524, Lille.</mods:affiliation></affiliation></author><author><name sortKey="Colson, Pierre" sort="Colson, Pierre" uniqKey="Colson P" first="Pierre" last="Colson">Pierre Colson</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Institute of Chemistry, University of Liège.</mods:affiliation></affiliation></author><author><name sortKey="De Pauw Gillet, Marie Claire" sort="De Pauw Gillet, Marie Claire" uniqKey="De Pauw Gillet M" first="Marie-Claire" last="De Pauw-Gillet">Marie-Claire De Pauw-Gillet</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Institute of Anatomy, University of Liège.</mods:affiliation></affiliation></author><author><name sortKey="Van Den Heuvel, Hilde" sort="Van Den Heuvel, Hilde" uniqKey="Van Den Heuvel H" first="Hilde" last="Van Den Heuvel">Hilde Van Den Heuvel</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Claeys, Magda" sort="Claeys, Magda" uniqKey="Claeys M" first="Magda" last="Claeys">Magda Claeys</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Lemiere, Filip" sort="Lemiere, Filip" uniqKey="Lemiere F" first="Filip" last="Lemière">Filip Lemière</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Esmans, Eddy L" sort="Esmans, Eddy L" uniqKey="Esmans E" first="Eddy L." last="Esmans">Eddy L. Esmans</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Rozenski, Jef" sort="Rozenski, Jef" uniqKey="Rozenski J" first="Jef" last="Rozenski">Jef Rozenski</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Rega Institute for Medical Research.</mods:affiliation></affiliation></author><author><name sortKey="Quirijnen, Ludo" sort="Quirijnen, Ludo" uniqKey="Quirijnen L" first="Ludo" last="Quirijnen">Ludo Quirijnen</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Tibotec-Virco NV.</mods:affiliation></affiliation></author><author><name sortKey="Maes, Louis" sort="Maes, Louis" uniqKey="Maes L" first="Louis" last="Maes">Louis Maes</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Tibotec-Virco NV.</mods:affiliation></affiliation></author><author><name sortKey="Dommisse, Roger" sort="Dommisse, Roger" uniqKey="Dommisse R" first="Roger" last="Dommisse">Roger Dommisse</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Lemiere, Guy L F" sort="Lemiere, Guy L F" uniqKey="Lemiere G" first="Guy L. F." last="Lemière">Guy L. F. Lemière</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Vlietinck, Arnold" sort="Vlietinck, Arnold" uniqKey="Vlietinck A" first="Arnold" last="Vlietinck">Arnold Vlietinck</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Pieters, Luc" sort="Pieters, Luc" uniqKey="Pieters L" first="Luc" last="Pieters">Luc Pieters</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation><affiliation><mods:affiliation> To whom correspondence should be addressed. Phone and fax: (32)3 820 27 09. E-mail pieters@uia.ua.ac.be.</mods:affiliation></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">ISTEX</idno><idno type="RBID">ISTEX:7040C93C2FEF52E1D59CF67C9FAF41EB0DF5CA74</idno><date when="2002" year="2002">2002</date><idno type="doi">10.1021/jm011102i</idno><idno type="url">https://api.istex.fr/ark:/67375/TPS-05PG9VCQ-8/fulltext.pdf</idno><idno type="wicri:Area/Istex/Corpus">000797</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Corpus" wicri:corpus="ISTEX">000797</idno></publicationStmt><sourceDesc><biblStruct><analytic><title level="a" type="main" xml:lang="en">Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of
New Neocryptolepine Derivatives</title><author><name sortKey="Jonckers, Tim H M" sort="Jonckers, Tim H M" uniqKey="Jonckers T" first="Tim H. M." last="Jonckers">Tim H. M. Jonckers</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation><affiliation><mods:affiliation> These authors contributed equally to this work.</mods:affiliation></affiliation></author><author><name sortKey="Van Miert, Sabine" sort="Van Miert, Sabine" uniqKey="Van Miert S" first="Sabine" last="Van Miert">Sabine Van Miert</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> These authors contributed equally to this work.</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Cimanga, Kanyanga" sort="Cimanga, Kanyanga" uniqKey="Cimanga K" first="Kanyanga" last="Cimanga">Kanyanga Cimanga</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Bailly, Christian" sort="Bailly, Christian" uniqKey="Bailly C" first="Christian" last="Bailly">Christian Bailly</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> INSERM U-524, Lille.</mods:affiliation></affiliation></author><author><name sortKey="Colson, Pierre" sort="Colson, Pierre" uniqKey="Colson P" first="Pierre" last="Colson">Pierre Colson</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Institute of Chemistry, University of Liège.</mods:affiliation></affiliation></author><author><name sortKey="De Pauw Gillet, Marie Claire" sort="De Pauw Gillet, Marie Claire" uniqKey="De Pauw Gillet M" first="Marie-Claire" last="De Pauw-Gillet">Marie-Claire De Pauw-Gillet</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Institute of Anatomy, University of Liège.</mods:affiliation></affiliation></author><author><name sortKey="Van Den Heuvel, Hilde" sort="Van Den Heuvel, Hilde" uniqKey="Van Den Heuvel H" first="Hilde" last="Van Den Heuvel">Hilde Van Den Heuvel</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Claeys, Magda" sort="Claeys, Magda" uniqKey="Claeys M" first="Magda" last="Claeys">Magda Claeys</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Lemiere, Filip" sort="Lemiere, Filip" uniqKey="Lemiere F" first="Filip" last="Lemière">Filip Lemière</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Esmans, Eddy L" sort="Esmans, Eddy L" uniqKey="Esmans E" first="Eddy L." last="Esmans">Eddy L. Esmans</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Rozenski, Jef" sort="Rozenski, Jef" uniqKey="Rozenski J" first="Jef" last="Rozenski">Jef Rozenski</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Rega Institute for Medical Research.</mods:affiliation></affiliation></author><author><name sortKey="Quirijnen, Ludo" sort="Quirijnen, Ludo" uniqKey="Quirijnen L" first="Ludo" last="Quirijnen">Ludo Quirijnen</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Tibotec-Virco NV.</mods:affiliation></affiliation></author><author><name sortKey="Maes, Louis" sort="Maes, Louis" uniqKey="Maes L" first="Louis" last="Maes">Louis Maes</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Tibotec-Virco NV.</mods:affiliation></affiliation></author><author><name sortKey="Dommisse, Roger" sort="Dommisse, Roger" uniqKey="Dommisse R" first="Roger" last="Dommisse">Roger Dommisse</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Lemiere, Guy L F" sort="Lemiere, Guy L F" uniqKey="Lemiere G" first="Guy L. F." last="Lemière">Guy L. F. Lemière</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Chemistry, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Vlietinck, Arnold" sort="Vlietinck, Arnold" uniqKey="Vlietinck A" first="Arnold" last="Vlietinck">Arnold Vlietinck</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation></author><author><name sortKey="Pieters, Luc" sort="Pieters, Luc" uniqKey="Pieters L" first="Luc" last="Pieters">Luc Pieters</name><affiliation><mods:affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</mods:affiliation></affiliation><affiliation><mods:affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</mods:affiliation></affiliation><affiliation><mods:affiliation> To whom correspondence should be addressed. Phone and fax: (32)3 820 27 09. E-mail pieters@uia.ua.ac.be.</mods:affiliation></affiliation></author></analytic><monogr></monogr><series><title level="j" type="main">Journal of Medicinal Chemistry</title><title level="j" type="abbrev">J. Med. Chem.</title><idno type="ISSN">0022-2623</idno><idno type="eISSN">1520-4804</idno><imprint><publisher>American Chemical Society</publisher><date type="e-published" when="2002-06-29">2002</date><date when="2002-08-01">2002</date><biblScope unit="vol">45</biblScope><biblScope unit="issue">16</biblScope><biblScope unit="page" from="3497">3497</biblScope><biblScope unit="page" to="3508">3508</biblScope></imprint><idno type="ISSN">0022-2623</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0022-2623</idno></seriesStmt></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract">On the basis of the original lead neocryptolepine or 5-methyl-5H-indolo[2,3-b]quinoline, an alkaloid from Cryptolepis sanguinolenta, derivatives were prepared using a biradical cyclization methodology. Starting from easily accessible educts, this approach allowed the synthesis of hitherto unknown compounds with a varied substitution pattern. As a result of steric hindrance, preferential formation of the 3-substituted isomers over the 1-substituted isomers was observed when cyclizing N-(3-substituted-phenyl)-N‘-[2-(2-trimethylsilylethynyl)phenyl]carbodiimides. All compounds were evaluated for their activity against chloroquine-sensitive as well as chloroquine-resistant Plasmodium falciparum strains, for their activity against Trypanosoma brucei and T. cruzi, and for their cytotoxicity on human MRC-5 cells. Mechanisms of action were investigated by testing heme complexation using ESI-MS, inhibition of β-hematin formation, DNA interactions (DNA−methyl green assay and linear dichroism), and inhibition of human topoisomerase II. Neocryptolepine derivatives with a higher antiplasmodial activity and a lower cytotoxicity than the original lead have been obtained. This selective antiplasmodial activity was associated with inhibition of β-hematin formation. 2-Bromoneocryptolepine was the most selective compound with an IC50 value against chloroquine-resistant P. falciparum of 4.0 μM in the absence of cytotoxicity (IC50 > 32 μM). Although cryptolepine, a known lead for antimalarials also originally isolated from Cryptolepis sanguinolenta, was more active (IC50 = 2.0 μM), 2-bromoneocryptolepine showed a low affinity for DNA and no inhibition of human topoisomerase II, in contrast to cryptolepine. Although some neocryptolepine derivatives showed a higher antiplasmodial activity than 2-bromocryptolepine, these compounds also showed a higher affinity for DNA and/or a more pronounced cytotoxicity. Therefore, 2-bromoneocryptolepine is considered as the most promising lead from the present work for new antimalarial agents. In addition, 2-bromo-, 2-nitro-, and 2-methoxy-9-cyanoneocryptolepine exhibited antitrypanosomal activity in the micromolar range in the absence of obvious cytotoxicity.</div></front></TEI><istex><corpusName>acs</corpusName><keywords><teeft><json:string>neocryptolepine</json:string><json:string>cryptolepine</json:string><json:string>hematin</json:string><json:string>antiplasmodial</json:string><json:string>cytotoxicity</json:string><json:string>heme</json:string><json:string>topoisomerase</json:string><json:string>mmol</json:string><json:string>antimalarial</json:string><json:string>derivative</json:string><json:string>falciparum</json:string><json:string>cytotoxic</json:string><json:string>neocryptolepine derivative</json:string><json:string>antiplasmodial activity</json:string><json:string>carbodiimides</json:string><json:string>plasmodium</json:string><json:string>hematin formation</json:string><json:string>alkaloid</json:string><json:string>chem</json:string><json:string>column chromatography</json:string><json:string>chloroquine</json:string><json:string>antitrypanosomal</json:string><json:string>test compound</json:string><json:string>medicinal chemistry</json:string><json:string>eluent</json:string><json:string>room temperature</json:string><json:string>brucei</json:string><json:string>antitrypanosomal activity</json:string><json:string>parasite</json:string><json:string>cruzi</json:string><json:string>dichroism</json:string><json:string>cryptolepis</json:string><json:string>cyclization</json:string><json:string>jonckers</json:string><json:string>biradical</json:string><json:string>assay</json:string><json:string>antwerp</json:string><json:string>general procedure</json:string><json:string>sanguinolenta</json:string><json:string>cryptolepis sanguinolenta</json:string><json:string>antimalarial agent</json:string><json:string>medicinal</json:string><json:string>positive control</json:string><json:string>human topoisomerase</json:string><json:string>higher antiplasmodial activity</json:string><json:string>heme monomer</json:string><json:string>heme detoxification process</json:string><json:string>datum</json:string><json:string>hydrochloride salt</json:string><json:string>more antiplasmodially active</json:string><json:string>more cytotoxic</json:string><json:string>concentration range</json:string><json:string>present work</json:string><json:string>flow rate</json:string><json:string>green assay</json:string><json:string>methyl green</json:string><json:string>mass spectrum</json:string><json:string>electrospray ionization</json:string><json:string>neocryptolepine derivative journal</json:string><json:string>electric linear dichroism</json:string><json:string>compound</json:string><json:string>plasmodium falciparum</json:string><json:string>more active</json:string><json:string>higher affinity</json:string><json:string>heme dimer</json:string><json:string>spectral datum</json:string><json:string>drug complex</json:string><json:string>selective compound</json:string><json:string>plasmodium parasite</json:string><json:string>obvious cytotoxicity</json:string><json:string>steric hindrance</json:string><json:string>biradical cyclization</json:string><json:string>malaria pigment</json:string></teeft></keywords><author><json:item><name>JONCKERS Tim H. M.</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Chemistry, University of Antwerp.</json:string><json:string>These authors contributed equally to this work.</json:string></affiliations></json:item><json:item><name>VAN MIERT Sabine</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>These authors contributed equally to this work.</json:string><json:string>Department of Pharmaceutical Sciences, University of Antwerp.</json:string></affiliations></json:item><json:item><name>CIMANGA Kanyanga</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Pharmaceutical Sciences, University of Antwerp.</json:string></affiliations></json:item><json:item><name>BAILLY Christian</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>INSERM U-524, Lille.</json:string></affiliations></json:item><json:item><name>COLSON Pierre</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Institute of Chemistry, University of Liège.</json:string></affiliations></json:item><json:item><name>DE PAUW-GILLET Marie-Claire</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Institute of Anatomy, University of Liège.</json:string></affiliations></json:item><json:item><name>VAN DEN HEUVEL Hilde</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Pharmaceutical Sciences, University of Antwerp.</json:string></affiliations></json:item><json:item><name>CLAEYS Magda</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Pharmaceutical Sciences, University of Antwerp.</json:string></affiliations></json:item><json:item><name>LEMIèRE Filip</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Chemistry, University of Antwerp.</json:string></affiliations></json:item><json:item><name>ESMANS Eddy L.</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Chemistry, University of Antwerp.</json:string></affiliations></json:item><json:item><name>ROZENSKI Jef</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Rega Institute for Medical Research.</json:string></affiliations></json:item><json:item><name>QUIRIJNEN Ludo</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Tibotec-Virco NV.</json:string></affiliations></json:item><json:item><name>MAES Louis</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Tibotec-Virco NV.</json:string></affiliations></json:item><json:item><name>DOMMISSE Roger</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Chemistry, University of Antwerp.</json:string></affiliations></json:item><json:item><name>LEMIèRE Guy L. F.</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Chemistry, University of Antwerp.</json:string></affiliations></json:item><json:item><name>VLIETINCK Arnold</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Pharmaceutical Sciences, University of Antwerp.</json:string></affiliations></json:item><json:item><name>PIETERS Luc</name><affiliations><json:string>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</json:string><json:string>Department of Pharmaceutical Sciences, University of Antwerp.</json:string><json:string>To whom correspondence should be addressed. Phone and fax: (32)3 820 27 09. E-mail pieters@uia.ua.ac.be.</json:string></affiliations></json:item></author><arkIstex>ark:/67375/TPS-05PG9VCQ-8</arkIstex><language><json:string>unknown</json:string></language><originalGenre><json:string>research-article</json:string></originalGenre><abstract>On the basis of the original lead neocryptolepine or 5-methyl-5H-indolo[2,3-b]quinoline, an alkaloid from Cryptolepis sanguinolenta, derivatives were prepared using a biradical cyclization methodology. Starting from easily accessible educts, this approach allowed the synthesis of hitherto unknown compounds with a varied substitution pattern. As a result of steric hindrance, preferential formation of the 3-substituted isomers over the 1-substituted isomers was observed when cyclizing N-(3-substituted-phenyl)-N‘-[2-(2-trimethylsilylethynyl)phenyl]carbodiimides. All compounds were evaluated for their activity against chloroquine-sensitive as well as chloroquine-resistant Plasmodium falciparum strains, for their activity against Trypanosoma brucei and T. cruzi, and for their cytotoxicity on human MRC-5 cells. Mechanisms of action were investigated by testing heme complexation using ESI-MS, inhibition of β-hematin formation, DNA interactions (DNA−methyl green assay and linear dichroism), and inhibition of human topoisomerase II. Neocryptolepine derivatives with a higher antiplasmodial activity and a lower cytotoxicity than the original lead have been obtained. This selective antiplasmodial activity was associated with inhibition of β-hematin formation. 2-Bromoneocryptolepine was the most selective compound with an IC50 value against chloroquine-resistant P. falciparum of 4.0 μM in the absence of cytotoxicity (IC50 > 32 μM). Although cryptolepine, a known lead for antimalarials also originally isolated from Cryptolepis sanguinolenta, was more active (IC50 = 2.0 μM), 2-bromoneocryptolepine showed a low affinity for DNA and no inhibition of human topoisomerase II, in contrast to cryptolepine. Although some neocryptolepine derivatives showed a higher antiplasmodial activity than 2-bromocryptolepine, these compounds also showed a higher affinity for DNA and/or a more pronounced cytotoxicity. Therefore, 2-bromoneocryptolepine is considered as the most promising lead from the present work for new antimalarial agents. In addition, 2-bromo-, 2-nitro-, and 2-methoxy-9-cyanoneocryptolepine exhibited antitrypanosomal activity in the micromolar range in the absence of obvious cytotoxicity.</abstract><qualityIndicators><score>10</score><pdfWordCount>8973</pdfWordCount><pdfCharCount>63791</pdfCharCount><pdfVersion>1.2</pdfVersion><pdfPageCount>12</pdfPageCount><pdfPageSize>612 x 792 pts (letter)</pdfPageSize><pdfWordsPerPage>748</pdfWordsPerPage><pdfText>true</pdfText><refBibsNative>true</refBibsNative><abstractWordCount>279</abstractWordCount><abstractCharCount>2210</abstractCharCount><keywordCount>0</keywordCount></qualityIndicators><title>Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of New Neocryptolepine Derivatives</title><genre><json:string>research-article</json:string></genre><host><title>Journal of Medicinal Chemistry</title><language><json:string>unknown</json:string></language><issn><json:string>0022-2623</json:string></issn><eissn><json:string>1520-4804</json:string></eissn><volume>45</volume><issue>16</issue><pages><first>3497</first><last>3508</last></pages><genre><json:string>journal</json:string></genre></host><ark><json:string>ark:/67375/TPS-05PG9VCQ-8</json:string></ark><categories><wos><json:string>1 - science</json:string><json:string>2 - chemistry, medicinal</json:string></wos><scienceMetrix><json:string>1 - natural sciences</json:string><json:string>2 - chemistry</json:string><json:string>3 - medicinal & biomolecular chemistry</json:string></scienceMetrix><scopus><json:string>1 - Life Sciences</json:string><json:string>2 - Pharmacology, Toxicology and Pharmaceutics</json:string><json:string>3 - Drug Discovery</json:string><json:string>1 - Life Sciences</json:string><json:string>2 - Biochemistry, Genetics and Molecular Biology</json:string><json:string>3 - Molecular Medicine</json:string></scopus><inist><json:string>1 - sciences appliquees, technologies et medecines</json:string><json:string>2 - sciences biologiques et medicales</json:string><json:string>3 - sciences biologiques fondamentales et appliquees. psychologie</json:string><json:string>4 - physiologie vegetale et developpement</json:string></inist></categories><publicationDate>2002</publicationDate><copyrightDate>2002</copyrightDate><doi><json:string>10.1021/jm011102i</json:string></doi><id>7040C93C2FEF52E1D59CF67C9FAF41EB0DF5CA74</id><score>1</score><fulltext><json:item><extension>pdf</extension><original>true</original><mimetype>application/pdf</mimetype><uri>https://api.istex.fr/ark:/67375/TPS-05PG9VCQ-8/fulltext.pdf</uri></json:item><json:item><extension>zip</extension><original>false</original><mimetype>application/zip</mimetype><uri>https://api.istex.fr/ark:/67375/TPS-05PG9VCQ-8/bundle.zip</uri></json:item><json:item><extension>txt</extension><original>false</original><mimetype>text/plain</mimetype><uri>https://api.istex.fr/ark:/67375/TPS-05PG9VCQ-8/fulltext.txt</uri></json:item><istex:fulltextTEI uri="https://api.istex.fr/ark:/67375/TPS-05PG9VCQ-8/fulltext.tei"><teiHeader><fileDesc><titleStmt><title level="a" type="main">Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of
New Neocryptolepine Derivatives</title></titleStmt><publicationStmt><authority>ISTEX</authority><publisher>American Chemical Society</publisher><availability><licence>Copyright © 2002 American Chemical Society</licence><p>American Chemical Society</p></availability><date type="e-published" when="2002-06-29">2002</date><date when="2002-08-01">2002</date><date type="Copyright" when="2002">2002</date></publicationStmt><notesStmt><note type="content-type" source="research-article" scheme="https://content-type.data.istex.fr/ark:/67375/XTP-1JC4F85T-7">research-article</note><note type="publication-type" scheme="https://publication-type.data.istex.fr/ark:/67375/JMC-0GLKJH51-B">journal</note></notesStmt><sourceDesc><biblStruct type="article"><analytic><title level="a" type="main" xml:lang="en">Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of
New Neocryptolepine Derivatives</title><author xml:id="author-0000"><persName><surname>Jonckers</surname><forename type="first">Tim H. M.</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>†</ref><p>
Department of Chemistry, University of Antwerp.</p></note><note place="foot"><ref>‡</ref><p>
These authors contributed equally to this work.</p></note></author><author xml:id="author-0001"><persName><surname>van Miert</surname><forename type="first">Sabine</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>‡</ref><p>
These authors contributed equally to this work.</p></note><note place="foot"><ref>§</ref><p>
Department of Pharmaceutical Sciences, University of Antwerp.</p></note></author><author xml:id="author-0002"><persName><surname>Cimanga</surname><forename type="first">Kanyanga</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>§</ref><p>
Department of Pharmaceutical Sciences, University of Antwerp.</p></note></author><author xml:id="author-0003"><persName><surname>Bailly</surname><forename type="first">Christian</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>‖</ref><p>
INSERM U-524, Lille.</p></note></author><author xml:id="author-0004"><persName><surname>Colson</surname><forename type="first">Pierre</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>⊥</ref><p>
Institute of Chemistry, University of Liège.</p></note></author><author xml:id="author-0005"><persName><surname>De Pauw-Gillet</surname><forename type="first">Marie-Claire</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>#</ref><p>
Institute of Anatomy, University of Liège.</p></note></author><author xml:id="author-0006"><persName><surname>van den Heuvel</surname><forename type="first">Hilde</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>§</ref><p>
Department of Pharmaceutical Sciences, University of Antwerp.</p></note></author><author xml:id="author-0007"><persName><surname>Claeys</surname><forename type="first">Magda</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>§</ref><p>
Department of Pharmaceutical Sciences, University of Antwerp.</p></note></author><author xml:id="author-0008"><persName><surname>Lemière</surname><forename type="first">Filip</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>†</ref><p>
Department of Chemistry, University of Antwerp.</p></note></author><author xml:id="author-0009"><persName><surname>Esmans</surname><forename type="first">Eddy L.</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>†</ref><p>
Department of Chemistry, University of Antwerp.</p></note></author><author xml:id="author-0010"><persName><surname>Rozenski</surname><forename type="first">Jef</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>∇</ref><p>
Rega Institute for Medical Research.</p></note></author><author xml:id="author-0011"><persName><surname>Quirijnen</surname><forename type="first">Ludo</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>◇</ref><p>
Tibotec-Virco NV.</p></note></author><author xml:id="author-0012"><persName><surname>Maes</surname><forename type="first">Louis</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>◇</ref><p>
Tibotec-Virco NV.</p></note></author><author xml:id="author-0013"><persName><surname>Dommisse</surname><forename type="first">Roger</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>†</ref><p>
Department of Chemistry, University of Antwerp.</p></note></author><author xml:id="author-0014"><persName><surname>Lemière</surname><forename type="first">Guy L. F.</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>†</ref><p>
Department of Chemistry, University of Antwerp.</p></note></author><author xml:id="author-0015"><persName><surname>Vlietinck</surname><forename type="first">Arnold</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>§</ref><p>
Department of Pharmaceutical Sciences, University of Antwerp.</p></note></author><author xml:id="author-0016" role="corresp"><persName><surname>Pieters</surname><forename type="first">Luc</forename></persName><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</affiliation><note place="foot"><ref>§</ref><p>
Department of Pharmaceutical Sciences, University of Antwerp.</p></note><affiliation role="corresp"> To whom correspondence should be addressed. Phone and fax: (32) 3 820 27 09. E-mail pieters@uia.ua.ac.be.</affiliation></author><idno type="istex">7040C93C2FEF52E1D59CF67C9FAF41EB0DF5CA74</idno><idno type="ark">ark:/67375/TPS-05PG9VCQ-8</idno><idno type="DOI">10.1021/jm011102i</idno></analytic><monogr><title level="j" type="main">Journal of Medicinal Chemistry</title><title level="j" type="abbrev">J. Med. Chem.</title><idno type="acspubs">jm</idno><idno type="coden">jmcmar</idno><idno type="pISSN">0022-2623</idno><idno type="eISSN">1520-4804</idno><imprint><publisher>American Chemical Society</publisher><date type="e-published" when="2002-06-29">2002</date><date when="2002-08-01">2002</date><biblScope unit="vol">45</biblScope><biblScope unit="issue">16</biblScope><biblScope unit="page" from="3497">3497</biblScope><biblScope unit="page" to="3508">3508</biblScope></imprint></monogr></biblStruct></sourceDesc></fileDesc><profileDesc><abstract><graphic url="jm011102in00001.tif"></graphic><p>On the basis of the original lead neocryptolepine or 5-methyl-5<hi rend="italic">H</hi>-indolo[2,3-<hi rend="italic">b</hi>]quinoline, an
alkaloid from <hi rend="italic">Cryptolepis </hi><hi rend="italic">sanguinolenta</hi>, derivatives were prepared using a biradical cyclization
methodology. Starting from easily accessible educts, this approach allowed the synthesis of
hitherto unknown compounds with a varied substitution pattern. As a result of steric hindrance,
preferential formation of the 3-substituted isomers over the 1-substituted isomers was observed
when cyclizing <hi rend="italic">N</hi>-(3-substituted-phenyl)-<hi rend="italic">N</hi><hi rend="italic">‘</hi><hi rend="italic">-</hi>[2-(2-trimethylsilylethynyl)phenyl]carbodiimides.
All compounds were evaluated for their activity against chloroquine-sensitive as well as
chloroquine-resistant <hi rend="italic">Plasmodium </hi><hi rend="italic">falciparum</hi> strains, for their activity against <hi rend="italic">Trypanosoma
brucei</hi> and <hi rend="italic">T. </hi><hi rend="italic">cruzi</hi>, and for their cytotoxicity on human MRC-5 cells. Mechanisms of action
were investigated by testing heme complexation using ESI-MS, inhibition of β-hematin
formation, DNA interactions (DNA−methyl green assay and linear dichroism), and inhibition
of human topoisomerase II. Neocryptolepine derivatives with a higher antiplasmodial activity
and a lower cytotoxicity than the original lead have been obtained. This selective antiplasmodial
activity was associated with inhibition of β-hematin formation. 2-Bromoneocryptolepine was
the most selective compound with an IC<hi rend="subscript">50</hi> value against chloroquine-resistant <hi rend="italic">P. falciparum</hi>
of 4.0 μM in the absence of cytotoxicity (IC<hi rend="subscript">50</hi> > 32 μM). Although cryptolepine, a known lead
for antimalarials also originally isolated from <hi rend="italic">Cryptolepis </hi><hi rend="italic">sanguinolenta</hi>, was more active (IC<hi rend="subscript">50</hi>
= 2.0 μM), 2-bromoneocryptolepine showed a low affinity for DNA and no inhibition of human
topoisomerase II, in contrast to cryptolepine. Although some neocryptolepine derivatives showed
a higher antiplasmodial activity than 2-bromocryptolepine, these compounds also showed a
higher affinity for DNA and/or a more pronounced cytotoxicity. Therefore, 2-bromoneocryptolepine is considered as the most promising lead from the present work for new antimalarial
agents. In addition, 2-bromo-, 2-nitro-, and 2-methoxy-9-cyanoneocryptolepine exhibited
antitrypanosomal activity in the micromolar range in the absence of obvious cytotoxicity.
</p></abstract><textClass ana="subject"><keywords scheme="document-type-name"><term>Article</term></keywords></textClass><langUsage><language ident="zxx"></language></langUsage></profileDesc></teiHeader></istex:fulltextTEI></fulltext><metadata><istex:metadataXml wicri:clean="corpus acs not found" wicri:toSee="no header"><istex:xmlDeclaration>version="1.0" encoding="UTF-8"</istex:xmlDeclaration><istex:document><article article-type="research-article" specific-use="acs2jats-1.1.23" dtd-version="1.1d1"><front><journal-meta><journal-id journal-id-type="acspubs">jm</journal-id><journal-id journal-id-type="coden">jmcmar</journal-id><journal-title-group><journal-title>Journal of Medicinal Chemistry</journal-title><abbrev-journal-title>J. Med. Chem.</abbrev-journal-title></journal-title-group><issn pub-type="ppub">0022-2623</issn><issn pub-type="epub">1520-4804</issn><publisher><publisher-name>American Chemical Society</publisher-name></publisher><self-uri>pubs.acs.org/jmc</self-uri></journal-meta><article-meta><article-id pub-id-type="doi">10.1021/jm011102i</article-id><article-categories><subj-group subj-group-type="document-type-name"><subject>Article</subject></subj-group></article-categories><title-group><article-title>Synthesis, Cytotoxicity, and Antiplasmodial and Antitrypanosomal Activity of
New Neocryptolepine Derivatives</article-title></title-group><contrib-group><contrib contrib-type="author"><name name-style="western"><surname>Jonckers</surname><given-names>Tim H. M.</given-names></name><xref rid="jm011102iAF2"><sup>†</sup></xref><xref rid="jm011102iAF3"><sup>‡</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>van Miert</surname><given-names>Sabine</given-names></name><xref rid="jm011102iAF4"><sup>§</sup></xref><xref rid="jm011102iAF3"><sup>‡</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Cimanga</surname><given-names>Kanyanga</given-names></name><xref rid="jm011102iAF4"><sup>§</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Bailly</surname><given-names>Christian</given-names></name><xref rid="jm011102iAF5"><sup>‖</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Colson</surname><given-names>Pierre</given-names></name><xref rid="jm011102iAF6"><sup>⊥</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>De Pauw-Gillet</surname><given-names>Marie-Claire</given-names></name><xref rid="jm011102iAF10"><sup>#</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>van den Heuvel</surname><given-names>Hilde</given-names></name><xref rid="jm011102iAF4"><sup>§</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Claeys</surname><given-names>Magda</given-names></name><xref rid="jm011102iAF4"><sup>§</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Lemière</surname><given-names>Filip</given-names></name><xref rid="jm011102iAF2"><sup>†</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Esmans</surname><given-names>Eddy L.</given-names></name><xref rid="jm011102iAF2"><sup>†</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Rozenski</surname><given-names>Jef</given-names></name><xref rid="jm011102iAF11"><sup>∇</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Quirijnen</surname><given-names>Ludo</given-names></name><xref rid="jm011102iAF15"><sup>◇</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Maes</surname><given-names>Louis</given-names></name><xref rid="jm011102iAF15"><sup>◇</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Dommisse</surname><given-names>Roger</given-names></name><xref rid="jm011102iAF2"><sup>†</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Lemière</surname><given-names>Guy L. F.</given-names></name><xref rid="jm011102iAF2"><sup>†</sup></xref></contrib><contrib contrib-type="author"><name name-style="western"><surname>Vlietinck</surname><given-names>Arnold</given-names></name><xref rid="jm011102iAF4"><sup>§</sup></xref></contrib><contrib contrib-type="author" corresp="yes"><name name-style="western"><surname>Pieters</surname><given-names>Luc</given-names></name><xref rid="jm011102iAF1">*</xref><xref rid="jm011102iAF4"><sup>§</sup></xref></contrib><aff>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department of
Chemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,
F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,
Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for Medical
Research, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium
</aff></contrib-group><author-notes><fn id="jm011102iAF2"><label>†</label><p>
Department of Chemistry, University of Antwerp.</p></fn><fn id="jm011102iAF3"><label>‡</label><p>
These authors contributed equally to this work.</p></fn><fn id="jm011102iAF4"><label>§</label><p>
Department of Pharmaceutical Sciences, University of Antwerp.</p></fn><fn id="jm011102iAF5"><label>‖</label><p>
INSERM U-524, Lille.</p></fn><fn id="jm011102iAF6"><label>⊥</label><p>
Institute of Chemistry, University of Liège.</p></fn><fn id="jm011102iAF10"><label>#</label><p>
Institute of Anatomy, University of Liège.</p></fn><fn id="jm011102iAF11"><label>∇</label><p>
Rega Institute for Medical Research.</p></fn><fn id="jm011102iAF15"><label>◇</label><p>
Tibotec-Virco NV.</p></fn><corresp id="jm011102iAF1">
To whom correspondence should be addressed. Phone and fax: (32)
3 820 27 09. E-mail pieters@uia.ua.ac.be.</corresp></author-notes><pub-date pub-type="epub"><day>29</day><month>06</month><year>2002</year></pub-date><pub-date pub-type="ppub"><day>01</day><month>08</month><year>2002</year></pub-date><volume>45</volume><issue>16</issue><fpage>3497</fpage><lpage>3508</lpage><supplementary-material xlink:href="jm011102i_s.pdf" orientation="portrait" position="float"></supplementary-material><history><date date-type="received"><day>13</day><month>11</month><year>2001</year></date><date date-type="asap"><day>29</day><month>06</month><year>2002</year></date><date date-type="issue-pub"><day>01</day><month>08</month><year>2002</year></date></history><permissions><copyright-statement>Copyright © 2002 American Chemical Society</copyright-statement><copyright-year>2002</copyright-year><copyright-holder>American Chemical Society</copyright-holder></permissions><abstract><graphic content-type="abstract-graphic" xlink:href="jm011102in00001.tif" orientation="portrait" position="float"></graphic><p>On the basis of the original lead neocryptolepine or 5-methyl-5<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline, an
alkaloid from <italic toggle="yes">Cryptolepis </italic><italic toggle="yes">sanguinolenta</italic>, derivatives were prepared using a biradical cyclization
methodology. Starting from easily accessible educts, this approach allowed the synthesis of
hitherto unknown compounds with a varied substitution pattern. As a result of steric hindrance,
preferential formation of the 3-substituted isomers over the 1-substituted isomers was observed
when cyclizing <italic toggle="yes">N</italic>-(3-substituted-phenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]carbodiimides.
All compounds were evaluated for their activity against chloroquine-sensitive as well as
chloroquine-resistant <italic toggle="yes">Plasmodium </italic><italic toggle="yes">falciparum</italic> strains, for their activity against <italic toggle="yes">Trypanosoma
brucei</italic> and <italic toggle="yes">T. </italic><italic toggle="yes">cruzi</italic>, and for their cytotoxicity on human MRC-5 cells. Mechanisms of action
were investigated by testing heme complexation using ESI-MS, inhibition of β-hematin
formation, DNA interactions (DNA−methyl green assay and linear dichroism), and inhibition
of human topoisomerase II. Neocryptolepine derivatives with a higher antiplasmodial activity
and a lower cytotoxicity than the original lead have been obtained. This selective antiplasmodial
activity was associated with inhibition of β-hematin formation. 2-Bromoneocryptolepine was
the most selective compound with an IC<sub>50</sub> value against chloroquine-resistant <italic toggle="yes">P. falciparum</italic>
of 4.0 μM in the absence of cytotoxicity (IC<sub>50</sub> > 32 μM). Although cryptolepine, a known lead
for antimalarials also originally isolated from <italic toggle="yes">Cryptolepis </italic><italic toggle="yes">sanguinolenta</italic>, was more active (IC<sub>50</sub>
= 2.0 μM), 2-bromoneocryptolepine showed a low affinity for DNA and no inhibition of human
topoisomerase II, in contrast to cryptolepine. Although some neocryptolepine derivatives showed
a higher antiplasmodial activity than 2-bromocryptolepine, these compounds also showed a
higher affinity for DNA and/or a more pronounced cytotoxicity. Therefore, 2-bromoneocryptolepine is considered as the most promising lead from the present work for new antimalarial
agents. In addition, 2-bromo-, 2-nitro-, and 2-methoxy-9-cyanoneocryptolepine exhibited
antitrypanosomal activity in the micromolar range in the absence of obvious cytotoxicity.
</p></abstract><custom-meta-group><custom-meta><meta-name>document-id-old-9</meta-name><meta-value>jm011102i</meta-value></custom-meta></custom-meta-group></article-meta></front><body><sec id="d7e385"><title>Introduction</title><p>Malaria is one of the most serious parasitic diseases
confronting both developing and industrialized nations,
causing more than 2.5 million deaths annually and a
staggering amount of chronic ill health. The main
species of human malaria parasites are <italic toggle="yes">Plasmodium
falciparum</italic>, <italic toggle="yes">P. vivax</italic>, <italic toggle="yes">P. ovale</italic>, and <italic toggle="yes">P. malariae</italic>. Malaria
caused by <italic toggle="yes">P. falciparum</italic> is the most severe form. A
major problem at the moment in the prevention and
treatment of malaria is the growing resistance of the
malarial parasite <italic toggle="yes">P. falciparum</italic> to currently available
drugs such as chloroquine. The incidence of malaria is
increasing, especially in tropical and subtropical areas;
in addition, morbidity is rising in the industrialized
world. Therefore, the development of new chemotherapeutic treatments for this disease is urgently needed.<named-content content-type="bibref-group"><xref rid="jm011102ib00001" ref-type="bibr"></xref>,<xref rid="jm011102ib00002" ref-type="bibr"></xref></named-content></p><p>The first antimalarial compound to be discovered,
which also served as the lead compound for synthetic
antimalarials of the chloroquine/mefloquine type, was
the alkaloid quinine. A lot of other leads for potential
new antimalarial agents have been characterized since
then, and many of them have been isolated from
medicinal plants.<xref rid="jm011102ib00003" ref-type="bibr"></xref> One of these is the plant alkaloid
cryptolepine (Figure <xref rid="jm011102if00001"></xref>) (5-methyl-5<italic toggle="yes">H-</italic>indolo[3,2-<italic toggle="yes">b</italic>]quinoline), the major alkaloid of the African plant <italic toggle="yes">Cryptolepis
sanguinolenta</italic>. Infusions of <italic toggle="yes">Cryptolepis </italic>root have a long-standing reputation in the treatment of malaria in
Central and West Africa (Ghana, Congo). Indeed, cryptolepine showed potent in vitro antiplasmodial activity
and no cross-resistance with chloroquine, and it was also
active in vivo (in mice infected with <italic toggle="yes">P. berghei</italic>).<named-content content-type="bibref-group"><xref rid="jm011102ib00004" ref-type="bibr"></xref>,<xref rid="jm011102ib00005" ref-type="bibr"></xref></named-content> In
addition to its antiplasmodial activity cryptolepine also
has antibacterial properties<xref rid="jm011102ib00006" ref-type="bibr"></xref> and is highly cytotoxic. Its
cytotoxicity is due to interaction with DNA and inhibition of topoisomerase II.<xref rid="jm011102ib00007" ref-type="bibr"></xref> Also, the minor alkaloid
neocryptolepine (<bold>1a</bold>) (Figure <xref rid="jm011102if00001"></xref>) (5-methyl-5<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline), from the same plant, showed in vitro
antiplasmodial activity against chloroquine-resistant
strains of <italic toggle="yes">P. falciparum</italic>.<xref rid="jm011102ib00005" ref-type="bibr"></xref> However, a direct comparison
of the cytotoxicity of cryptolepine and neocryptolepine demonstrated that the latter was much less cytotoxic.<xref rid="jm011102ib00008" ref-type="bibr"></xref> Synthetic δ-carbolines, benzo-δ-carbolines, and
cryptolepine derivatives were evaluated for their cytotoxic, antiplasmodial, and antitrypanosomal activities.
Most benzo-δ-carbolines and cryptolepine derivatives
remained relatively cytotoxic; 1-methyl-δ-carboline was
selected as the most promising antimalarial compound.<xref rid="jm011102ib00009" ref-type="bibr"></xref>
Synthetic derivatives of cryptolepine have also been
evaluated for their antihyperglycaemic properties.<xref rid="jm011102ib00010" ref-type="bibr"></xref>
Cryptolepine also displays a series of other pharmacological effects, such as antimuscarinic, noradrenergic
receptor antagonistic, antihypertensive, vasodilative,
antithrombotic, antipyretic, and antiinflammatory properties,<xref rid="jm011102ib00010" ref-type="bibr"></xref> some of which may contribute to the relatively
high systemic toxicity observed during preliminary in
vivo experiments in mice.<xref rid="jm011102ib00011" ref-type="bibr"></xref><fig id="jm011102if00001" position="float" orientation="portrait"><label>1</label><caption><p>Two constituents of<italic toggle="yes">Cryptolepis sanguinolenta</italic>.</p></caption><graphic xlink:href="jm011102if00001.tif" position="float" orientation="portrait"></graphic></fig></p><p>Therefore, and because of its reduced cytotoxicity
compared to cryptolepine, neocryptolepine was selected
in the present work as a lead for the development of
new antimalarial agents. The antiplasmodial properties
of a series of synthetic neocryptolepine derivatives are
discussed, as well as their activity in a series of
functional assays relating to possible mechanisms of
action, to establish structure−activity relationships. In
addition, the antitrypanosomal activity against <italic toggle="yes">T. </italic><italic toggle="yes">cruzi</italic>
and <italic toggle="yes">T. brucei</italic> was also evaluated.
</p></sec><sec id="d7e505"><title>Chemistry</title><p>During the past 8 years, several research groups
developed synthetic pathways toward <bold>1a</bold>. In 1994,
Peczynska-Czoch et al<italic toggle="yes">.</italic><xref rid="jm011102ib00012" ref-type="bibr"></xref> reported the coupling between
substituted 2-chloroquinolines and benzotriazole. The
molecules thus obtained were subjected to a Graebe−Ullmann reaction with polyphosphoric acid (PPA) to
give the corresponding 6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinolines in
moderate yield. The subsequent transformation of these
molecules into the corresponding neocryptolepine derivatives is straightforward. Molina et al<italic toggle="yes">.</italic><named-content content-type="bibref-group"><xref rid="jm011102ib00013" ref-type="bibr"></xref>,<xref rid="jm011102ib00014" ref-type="bibr"></xref></named-content> reported
the synthesis of <bold>1a</bold> via an iminophosphorane methodology, while Timári et al<italic toggle="yes">.</italic><xref rid="jm011102ib00015" ref-type="bibr"></xref> synthesized neocryptolepine
via a Pd-catalyzed cross-coupling strategy. Although the
above-mentioned procedures are straightforward for the
synthesis of <bold>1a</bold>, using these methods for the synthesis
of derivatives of neocryptolepine with a diverse substitution pattern is not that obvious. A much more promising procedure has been developed by Schmittel et al<italic toggle="yes">.</italic><xref rid="jm011102ib00016" ref-type="bibr"></xref>
who reported a biradical cyclization reaction on <italic toggle="yes">N</italic>-[4-methyl-2-(2-trimethylsilylethynyl)phenyl]-<italic toggle="yes">N</italic>‘-phenylcarbodiimide yielding 9-methyl-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline, which is an obvious precursor of
9-methylneocryptolepine. Similar to this is Shi's thermolysis of <italic toggle="yes">N</italic>-[2-(1-alkynyl)phenyl]-<italic toggle="yes">N</italic>‘-phenylcarbodiimides in γ-terpinene or <italic toggle="yes">p</italic>-xylene giving 6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinolines.<xref rid="jm011102ib00017" ref-type="bibr"></xref> It is worth noting that in 1992 Saito and
co-workers<xref rid="jm011102ib00018" ref-type="bibr"></xref> published the Lewis acid induced intramolecular Diels−Alder reaction of conjugated carbodiimides leading to indolo[2,3-<italic toggle="yes">b</italic>] quinolines if the reaction
conditions are carefully chosen. From our point of view,
the methods of Schmittel and Shi seemed to be the most
convenient, since they allow an easy alteration of the
substitution pattern of the A-ring of <bold>1a</bold> by starting from
appropriately substituted and commercially available
phenyl isothiocyanates and phenyl isocyanates. Variation of the D-ring can arise by starting from the
appropriately substituted <italic toggle="yes">o</italic>-iodoanilines. Moreover, the
possibility of introducing different substituents at position 11 of the C-ring arising from the presence of a
suitably substituted alkynyl group<xref rid="jm011102ib00019" ref-type="bibr"></xref> in the carbodiimide
made this method the most attractive one (Scheme <xref rid="jm011102ih00001"></xref>).
<fig id="jm011102ih00001" position="float" fig-type="scheme" orientation="portrait"><label>1</label><caption><p>General Synthetic Sequence</p></caption><graphic xlink:href="jm011102ih00001.tif" position="float" orientation="portrait"></graphic></fig></p><p>The reaction sequence starts with a Sonogashira
reaction on <italic toggle="yes">o</italic>-iodoanilines <bold>2 </bold>and <bold>2</bold>‘ giving the 2-trimethylsilylethynylanilines <bold>3 </bold>and<bold> 3</bold>‘, respectively, in good
yields. Compound <bold>2</bold>‘ was synthesized starting from
4-aminobenzonitrile using a combination of ICl and
CaCO<sub>3</sub> in MeOH as an iodine source.<xref rid="jm011102ib00020" ref-type="bibr"></xref> First, we tried
to obtain all the carbodiimides <bold>7a</bold><bold>−</bold><bold>o </bold>via the corresponding thioureas, but unfortunately compound <bold>3</bold>‘ did
not react with the phenyl isothiocyanates <bold>4a</bold><bold>−</bold><bold>j</bold>. In these
cases, we synthesized the carbodiimides <bold>7k</bold><bold>−</bold><bold>o</bold> via an
aza-Wittig reaction of the iminophoshorane <bold>5</bold>‘ with
phenyl isocyanates<sup>14</sup> <bold>6k</bold><bold>−</bold><bold>o</bold>. Another problem was encountered in synthesizing the thioureas. When the
conditions described in the Schmittel paper were used,
the formation of compounds <bold>5a</bold><bold>−</bold><bold>j</bold> was rather troublesome. These intermediates could only be obtained in
40−50% yield after 5 days of reaction time and a
problematic chromatographic purification. We therefore
tried to develop a more practical method to overcome
this problem. Replacing the solvent acetone by ethanol
had a dramatic effect on the reaction rate as well as on
the yield. After a solution of 2-(2-trimethylsilylethynyl)aniline <bold>3</bold> and the appropriate isothiocyanate in ethanol
was stirred in the presence of a catalytic amount of
(dimethylamino)pyridine (DMAP) for 1−2 h at 40 °C, a
white precipitate is formed, which after filtration and
drying gives the corresponding thioureas <bold>5a</bold><bold>−</bold><bold>j</bold> in good
yields (60−70%). Transformation of the thioureas into
the carbodiimides <bold>7a</bold><bold>−</bold><bold>j </bold>was achieved using the procedure reported by Fell and Coppola.<xref rid="jm011102ib00021" ref-type="bibr"></xref> Most of these
carbodiimides are quite susceptible to degradation.
Therefore, after their purification, these products were
immediately used in the cyclization reaction. Such
unstable behavior of carbodiimides has been reported
before.<xref rid="jm011102ib00022" ref-type="bibr"></xref> After biradical cyclization in the presence of
1,4-cyclohexadiene, substituted 11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinolines <bold>8a</bold><bold>−</bold><bold>o</bold> were obtained in good
yields (Table <xref rid="jm011102it00001"></xref>). These compounds were subsequently
transformed into the new neocryptolepine derivatives
<bold>1a</bold><bold>−</bold><bold>o</bold> by methylation of the 5-nitrogen atom followed
by desilylation.<xref rid="jm011102ib00017" ref-type="bibr"></xref> Initially ethanol was used instead of
dimethylformamide (DMF) as the solvent for the methylation; the use of the latter, however, greatly enhanced the yield. The desilylation can be performed
prior to the methylation, but this in general reduces the
overall yield. The outcome of the biradical cyclization
does not seem to be influenced by the nature of the R
and R‘ substituents. This is in contrast to the effect of
the substituent at the alkyne terminus, which determines the reaction product of the biradical cyclization.<xref rid="jm011102ib00016" ref-type="bibr"></xref>
Besides the products listed in Table <xref rid="jm011102it00001"></xref>, we also prepared
7-trimethylsilylnaphtho[1,2-<italic toggle="yes">b</italic>]-α-carboline <bold>8p</bold> and 2,4-dichloro-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline <bold>8q
</bold>(via route A), but these compounds could not be methylated even after prolonged reaction times (Figure <xref rid="jm011102if00002"></xref>). A
low electron density in the A-ring caused by an electron-withdrawing substituent may deactivate the 5-nitrogen
atom toward nucleophilic attack. However, this possibility can be ruled out by the successful synthesis of
2-cyanoneocryptolepine (<bold>1j</bold>) and 2-nitroneocryptolepine
(<bold>1h</bold>). It therefore seems that the presence of a substituent at the 4-position prohibits the introduction of a
methyl group at the 5-nitrogen atom because of steric
hindrance.
<fig id="jm011102if00002" position="float" orientation="portrait"><label>2</label><caption><p>Cyclization products resistant to methylation.</p></caption><graphic xlink:href="jm011102if00002.tif" position="float" orientation="portrait"></graphic></fig><table-wrap id="jm011102it00001" position="float" orientation="portrait"><label>1</label><caption><p>Biradical Cyclization of Carbodiimides<bold>7</bold><bold>a</bold>−<bold>o</bold> to
11-Trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinolines <bold>8</bold><bold>a</bold>−<bold>o</bold></p></caption><oasis:table colsep="2" rowsep="2"><oasis:tgroup cols="8"><oasis:colspec colnum="1" colname="1"></oasis:colspec><oasis:colspec colnum="2" colname="2"></oasis:colspec><oasis:colspec colnum="3" colname="3"></oasis:colspec><oasis:colspec colnum="4" colname="4"></oasis:colspec><oasis:colspec colnum="5" colname="5"></oasis:colspec><oasis:colspec colnum="6" colname="6"></oasis:colspec><oasis:colspec colnum="7" colname="7"></oasis:colspec><oasis:colspec colnum="8" colname="8"></oasis:colspec><oasis:tbody><oasis:row><oasis:entry namest="1" nameend="1">compd
type</oasis:entry><oasis:entry namest="2" nameend="2">R‘</oasis:entry><oasis:entry namest="3" nameend="3">R</oasis:entry><oasis:entry namest="4" nameend="4">yield,<italic toggle="yes"><sup>a</sup></italic><sup></sup> %</oasis:entry><oasis:entry namest="5" nameend="5">compd
type</oasis:entry><oasis:entry namest="6" nameend="6">R‘</oasis:entry><oasis:entry namest="7" nameend="7">R</oasis:entry><oasis:entry namest="8" nameend="8">yield,<italic toggle="yes"><sup>a</sup></italic><sup></sup> %
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>a</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">H
</oasis:entry><oasis:entry colname="4">60
</oasis:entry><oasis:entry colname="5"><bold>i</bold></oasis:entry><oasis:entry colname="6">H
</oasis:entry><oasis:entry colname="7">SCH<sub>3</sub></oasis:entry><oasis:entry colname="8">89
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>b</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">OCH3
</oasis:entry><oasis:entry colname="4">92
</oasis:entry><oasis:entry colname="5"><bold>j</bold></oasis:entry><oasis:entry colname="6">H
</oasis:entry><oasis:entry colname="7">CN
</oasis:entry><oasis:entry colname="8">81
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>c</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">Br
</oasis:entry><oasis:entry colname="4">89
</oasis:entry><oasis:entry colname="5"><bold>k</bold></oasis:entry><oasis:entry colname="6">CN
</oasis:entry><oasis:entry colname="7">H
</oasis:entry><oasis:entry colname="8">70
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>d</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">Cl
</oasis:entry><oasis:entry colname="4">85
</oasis:entry><oasis:entry colname="5"><bold>l</bold></oasis:entry><oasis:entry colname="6">CN
</oasis:entry><oasis:entry colname="7">Cl
</oasis:entry><oasis:entry colname="8">71
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>e</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">F
</oasis:entry><oasis:entry colname="4">82
</oasis:entry><oasis:entry colname="5"><bold>m</bold></oasis:entry><oasis:entry colname="6">CN
</oasis:entry><oasis:entry colname="7">OCH<sub>3</sub></oasis:entry><oasis:entry colname="8">91
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>f</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">I
</oasis:entry><oasis:entry colname="4">74
</oasis:entry><oasis:entry colname="5"><bold>n</bold></oasis:entry><oasis:entry colname="6">CN
</oasis:entry><oasis:entry colname="7">CF<sub>3</sub></oasis:entry><oasis:entry colname="8">74
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>g</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">CH<sub>3</sub></oasis:entry><oasis:entry colname="4">76
</oasis:entry><oasis:entry colname="5"><bold>o</bold></oasis:entry><oasis:entry colname="6">CN
</oasis:entry><oasis:entry colname="7">F
</oasis:entry><oasis:entry colname="8">72
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>h</bold></oasis:entry><oasis:entry colname="2">H
</oasis:entry><oasis:entry colname="3">NO<sub>2</sub></oasis:entry><oasis:entry colname="4">62
</oasis:entry><oasis:entry colname="5"></oasis:entry><oasis:entry colname="6"></oasis:entry><oasis:entry colname="7"></oasis:entry><oasis:entry colname="8"></oasis:entry></oasis:row></oasis:tbody></oasis:tgroup></oasis:table><table-wrap-foot><p><italic toggle="yes"><sup>a</sup></italic><sup></sup> Yields given refer to analytically pure compounds.</p></table-wrap-foot></table-wrap></p><p>We were also interested in some non-methylated
derivatives of <bold>1a</bold>, and therefore, we prepared 2-methoxy-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10b</bold>), 2-chloro-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10d</bold>), 2-fluor-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10e</bold>),
and 2,4-dichloro-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10q</bold>) by desilylation of the corresponding silylated compounds (<bold>8b,
8d, 8e, 8q</bold>) (Scheme <xref rid="jm011102ih00002"></xref>).
<fig id="jm011102ih00002" position="float" fig-type="scheme" orientation="portrait"><label>2</label><caption><p>Synthesis of Some Desilylated Compounds<italic toggle="yes"><sup>a</sup></italic><sup></sup></p><p><fn id="d7e1080"><p><italic toggle="yes"><sup>a</sup></italic><sup></sup> Compound <bold>b</bold>: R<sub>1</sub> = OCH<sub>3</sub>, R<sub>2</sub> = H. Compound <bold>d</bold>: R<sub>1</sub> = Cl, R<sub>2</sub>
= H. Compound <bold>e</bold>: R<sub>1</sub> = F, R<sub>2</sub> = H. Compound <bold>q</bold>: R<sub>1</sub> = Cl, R<sub>2</sub> =
Cl.</p></fn></p></caption><graphic xlink:href="jm011102ih00002.tif" position="float" orientation="portrait"></graphic></fig></p><p>Besides 2- and 9-substituted neocryptolepine derivatives, we wanted to prepare other isomers of the
products synthesized so far for structure−activity relationship (SAR) studies. As a starting point, a further
modification of the A-ring was investigated. However,
some important remarks have to be made in this
context. First of all, from a chemical point of view, the
iminophosphorane route (route B, Scheme <xref rid="jm011102ih00001"></xref>) proved to
be more practical than the thiourea pathway (route A,
Scheme <xref rid="jm011102ih00001"></xref>). Therefore, we decided to use this approach
for the synthesis of the extra set of neocryptolepine
derivatives using the iminophosphorane (<bold>5</bold>‘ ‘).<xref rid="jm011102ib00017" ref-type="bibr"></xref> Second,
if one uses the biradical cyclization reaction to construct
the tetracyclic core starting from <italic toggle="yes">N</italic>-(3-substituted-phenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimides (<bold>7r</bold><bold>−</bold><bold>u</bold>), one can expect a mixture of the 1- and
3-substituted isomers because two different ring-closure
reactions can occur (Scheme <xref rid="jm011102ih00003"></xref>, Table <xref rid="jm011102it00002"></xref>).
<fig id="jm011102ih00003" position="float" fig-type="scheme" orientation="portrait"><label>3</label><caption><p>Neocryptolepine Derivatives<bold>1r</bold>−<bold>1</bold><bold>y</bold></p></caption><graphic xlink:href="jm011102ih00003.tif" position="float" orientation="portrait"></graphic></fig><table-wrap id="jm011102it00002" position="float" orientation="portrait"><label>2</label><caption><p>Synthesis of Isomeric Neocryptolepine Precursors</p></caption><oasis:table colsep="2" rowsep="2"><oasis:tgroup cols="5"><oasis:colspec colnum="1" colname="1"></oasis:colspec><oasis:colspec colnum="2" colname="2"></oasis:colspec><oasis:colspec colnum="3" colname="3"></oasis:colspec><oasis:colspec colnum="4" colname="4"></oasis:colspec><oasis:colspec colnum="5" colname="5"></oasis:colspec><oasis:tbody><oasis:row><oasis:entry namest="1" nameend="1">compd
type</oasis:entry><oasis:entry namest="2" nameend="2">carbodiimide <bold>7</bold></oasis:entry><oasis:entry namest="3" nameend="3">reaction
product <bold>8</bold></oasis:entry><oasis:entry namest="4" nameend="4">substitution</oasis:entry><oasis:entry namest="5" nameend="5">yield,<italic toggle="yes"><sup>a</sup></italic><sup></sup> %
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>r</bold></oasis:entry><oasis:entry colname="2"><bold>7r</bold></oasis:entry><oasis:entry colname="3"><bold>8r</bold></oasis:entry><oasis:entry colname="4">1-OCH<sub>3</sub></oasis:entry><oasis:entry colname="5">8
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"></oasis:entry><oasis:entry colname="2"></oasis:entry><oasis:entry colname="3"><bold>8v</bold></oasis:entry><oasis:entry colname="4">3-OCH<sub>3</sub></oasis:entry><oasis:entry colname="5">41
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>s</bold></oasis:entry><oasis:entry colname="2"><bold>7s</bold></oasis:entry><oasis:entry colname="3"><bold>8v</bold></oasis:entry><oasis:entry colname="4">1-Br
</oasis:entry><oasis:entry colname="5">13
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"></oasis:entry><oasis:entry colname="2"></oasis:entry><oasis:entry colname="3"><bold>8w</bold></oasis:entry><oasis:entry colname="4">3-Br
</oasis:entry><oasis:entry colname="5">55
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>t</bold></oasis:entry><oasis:entry colname="2"><bold>7t</bold></oasis:entry><oasis:entry colname="3"><bold>8t</bold></oasis:entry><oasis:entry colname="4">1-Cl
</oasis:entry><oasis:entry colname="5"><1<italic toggle="yes"><sup>b</sup></italic><sup></sup></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"></oasis:entry><oasis:entry colname="2"></oasis:entry><oasis:entry colname="3"><bold>8x</bold></oasis:entry><oasis:entry colname="4">3-Cl
</oasis:entry><oasis:entry colname="5">52
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>u</bold></oasis:entry><oasis:entry colname="2"><bold>7u</bold></oasis:entry><oasis:entry colname="3"><bold>8u</bold></oasis:entry><oasis:entry colname="4">1-CF<sub>3</sub></oasis:entry><oasis:entry colname="5">4
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"></oasis:entry><oasis:entry colname="2"></oasis:entry><oasis:entry colname="3"><bold>8y</bold></oasis:entry><oasis:entry colname="4">3-CF<sub>3</sub></oasis:entry><oasis:entry colname="5">39</oasis:entry></oasis:row></oasis:tbody></oasis:tgroup></oasis:table><table-wrap-foot><p><italic toggle="yes"><sup>a</sup></italic><sup></sup> Yields given refer to analytically pure compounds.<italic toggle="yes"><sup>b</sup></italic><sup></sup> Product
not characterized.</p></table-wrap-foot></table-wrap></p><p>Both isomers were formed with the 3-isomer predominating in each case. In the case of the chloro-substituted
carbodiimide (<bold>7t</bold>), only a very low amount of the
1-isomer (<bold>8t</bold>) was formed (less than 1%). The preferential formation of the 3-isomers is undoubtedly the result
of the absence of steric hindrance between the
substituent and the bulky trimethylsilyl group when the ring-closure reaction takes place.
</p></sec><sec id="d7e1356"><title>Results and Discussion</title><p><bold>(a) </bold><bold>In Vitro</bold><bold> Antiplasmodial Activity and Cytotoxicity.</bold> All synthesized compounds and cryptolepine
from natural origin<sup>5</sup> were evaluated as their hydrochloride salts for in vitro antiplasmodial activity against a
chloroquine-sensitive and a chloroquine-resistant <italic toggle="yes">P.
falciparum </italic>strain and for cytotoxicity on a human cell
line (MRC-5 cells) (Table <xref rid="jm011102it00003"></xref>). All compounds were
marginally more active against the chloroquine-resistant strain than against the chloroquine-sensitive one,
so the discussion will be focused on activity against the
first one. It is confirmed that neocryptolepine (<bold>1a</bold>), the
original lead in this work, is less cytotoxic than cryptolepine; however, the antiplasmodial activity is also
reduced. Many of the 2-substituted neocryptolepine
derivatives are more antiplasmodially active than neocryptolepine itself but are also more cytotoxic. The most
notable compound in this regard is 2-methylneocryptolepine (<bold>1g</bold>), with a cytotoxicity and antiplasmodial
activity comparable to those of cryptolepine. This is in
agreement with earlier findings for a series of methyl-substituted indolo[2,3-<italic toggle="yes">b</italic>]quinoline derivatives.<xref rid="jm011102ib00012" ref-type="bibr"></xref> Although 2-methylneocryptolepine was not included in
this study, it was found that the cytotoxicity (against
KB cells) was strongly influenced by the position and
the number of methyl substituents, the most cytotoxic
one being 2,5,9,11-tetramethyl-5<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline. The 2-halo-substituted neocryptolepines, on the
other hand (2-bromo-, 2-chloro-, 2-fluoro-, and 2-iodoneocryptolepine) (<bold>1c</bold>, <bold>1d</bold>, <bold>1e</bold>, and <bold>1f</bold>, respectively), are
more active against <italic toggle="yes">P. </italic><italic toggle="yes">falciparum</italic> than neocryptolepine
(<bold>1a</bold>) and less cytotoxic, the most active and selective
compound being 2-bromoneocryptolepine (<bold>1c</bold>) with an
IC<sub>50</sub> against cloroquine-resistant <italic toggle="yes">P. falciparum</italic> of 4.0
μM while the IC<sub>50</sub> on the MRC-5 cells is >32 μM.
Removal of the <italic toggle="yes">N</italic>-methyl group, as in compounds <bold>10d</bold>
(compared to 2-chloroneocryptolepine <bold>1d</bold>) and <bold>10e</bold> (compared to 2-fluoroneocryptolepine <bold>1e</bold>), leads to a complete
loss of biological activity in the concentration range
tested. When comparing 1-, 2-, and 3-substituted neocryptolepine isomers, e.g., in the Br-substituted series,
1-bromo substitution (<bold>1s</bold>) leads to a loss of antiplasmodial activity; 3-bromoneocryptolepine (<bold>1w</bold>) has about the
same antiplasmodial activity against the chloroquine-resistant strain, but it is more cytotoxic. In the methoxy-substituted series, 2-methoxyneocryptolepine (<bold>1b</bold>) and
3-methoxyneocryptolepine (<bold>1v</bold>) are especially more antiplasmodially active than neocryptolepine (<bold>1a</bold>) but also
more cytotoxic. Interestingly, introduction of a 9-cyano
substituent, as in compound <bold>1k</bold> (compared to neocryptolepine <bold>1a</bold>), compound <bold>1l</bold> (compared to 2-chloroneocryptolepine <bold>1d</bold>), and compound <bold>1m </bold>(compared to
2-methoxyneocryptolepine <bold>1b</bold>), leads to a loss of cytotoxicity but also to a reduction of the antiplasmodial
activity.
<table-wrap id="jm011102it00003" position="float" orientation="portrait"><label>3</label><caption><p>In Vitro Cytotoxic, Antiplasmodial, and Antitrypanosomal Activity, DNA Interaction (DNA Methyl Green Assay), and Inhibition of β-Hematin Formation of Synthetic Neocryptolepine Derivatives</p></caption><oasis:table colsep="2" rowsep="2"><oasis:tgroup cols="8"><oasis:colspec colnum="1" colname="1"></oasis:colspec><oasis:colspec colnum="2" colname="2"></oasis:colspec><oasis:colspec colnum="3" colname="3"></oasis:colspec><oasis:colspec colnum="4" colname="4"></oasis:colspec><oasis:colspec colnum="5" colname="5"></oasis:colspec><oasis:colspec colnum="6" colname="6"></oasis:colspec><oasis:colspec colnum="7" colname="7"></oasis:colspec><oasis:colspec colnum="8" colname="8"></oasis:colspec><oasis:tbody><oasis:row><oasis:entry namest="1" nameend="1">compd</oasis:entry><oasis:entry namest="2" nameend="2">cytotoxicity
(MRC-5 cells)
IC<sub>50</sub> (μM)</oasis:entry><oasis:entry namest="3" nameend="3"><italic toggle="yes">Plasmodium falciparum</italic>
(chloroquine sensitive)
(Ghana strain)
IC<sub>50</sub> (μM)</oasis:entry><oasis:entry namest="4" nameend="4"><italic toggle="yes">Plasmodium falciparum</italic>
(chloroquine resistant)
(W2 strain)
IC<sub>50</sub> (μM)</oasis:entry><oasis:entry namest="5" nameend="5"><italic toggle="yes">Trypanosoma</italic><italic toggle="yes">brucei</italic>
IC<sub>50</sub> (μM)</oasis:entry><oasis:entry namest="6" nameend="6"><italic toggle="yes">Trypanosoma</italic><italic toggle="yes">cruzi</italic>
IC<sub>50</sub> (μM)</oasis:entry><oasis:entry namest="7" nameend="7">DNA
interaction
IC<sub>50</sub> (μM)</oasis:entry><oasis:entry namest="8" nameend="8">inhibition of
β-hematin
formation<italic toggle="yes"><sup>a</sup></italic><sup></sup></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1a</bold></oasis:entry><oasis:entry colname="2">11.0 ± 1.4
</oasis:entry><oasis:entry colname="3">27.3 ± 5.7
</oasis:entry><oasis:entry colname="4">14.0 ± 1.7
</oasis:entry><oasis:entry colname="5">4.0 ± 0.1
</oasis:entry><oasis:entry colname="6">4.0 ± 0.1
</oasis:entry><oasis:entry colname="7">92.8 ± 9.7
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1b</bold></oasis:entry><oasis:entry colname="2">4.0 ± 0.1
</oasis:entry><oasis:entry colname="3">4.3 ± 0.6
</oasis:entry><oasis:entry colname="4">4.7 ± 0.6
</oasis:entry><oasis:entry colname="5">0.3 ± 0.1
</oasis:entry><oasis:entry colname="6">1.7 ± 0.6
</oasis:entry><oasis:entry colname="7">77.9 ± 4.4
</oasis:entry><oasis:entry colname="8">−
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>10b</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="7">>1000
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1c</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">6.0 ± 6.1
</oasis:entry><oasis:entry colname="4">4.0 ± 0.1
</oasis:entry><oasis:entry colname="5">1.0 ± 0.1
</oasis:entry><oasis:entry colname="6">>32
</oasis:entry><oasis:entry colname="7">>400
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1d</bold></oasis:entry><oasis:entry colname="2">16.5 ± 0.7
</oasis:entry><oasis:entry colname="3">21.0 ± 8.9
</oasis:entry><oasis:entry colname="4">5.0 ± 0.1
</oasis:entry><oasis:entry colname="5">1.3 ± 0.6
</oasis:entry><oasis:entry colname="6">12.0 ± 1.7
</oasis:entry><oasis:entry colname="7">>500
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>10d</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6">>32
</oasis:entry><oasis:entry colname="7">>1000
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1e</bold></oasis:entry><oasis:entry colname="2">15.0 ± 0.1
</oasis:entry><oasis:entry colname="3">19.3 ± 3.8
</oasis:entry><oasis:entry colname="4">4.7 ± 0.6
</oasis:entry><oasis:entry colname="5">14.3 ± 0.6
</oasis:entry><oasis:entry colname="6">6.0 ± 1.0
</oasis:entry><oasis:entry colname="7">>500
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>10e</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6">>32
</oasis:entry><oasis:entry colname="7">>1000
</oasis:entry><oasis:entry colname="8">−
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1f</bold></oasis:entry><oasis:entry colname="2">16.0 ± 0.1
</oasis:entry><oasis:entry colname="3">17.7 ± 5.1
</oasis:entry><oasis:entry colname="4">6.3 ± 0.6
</oasis:entry><oasis:entry colname="5">1.0 ± 0.1
</oasis:entry><oasis:entry colname="6">18.3 ± 0.6
</oasis:entry><oasis:entry colname="7">>400
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1g</bold></oasis:entry><oasis:entry colname="2">0.95 ± 0.07
</oasis:entry><oasis:entry colname="3">2.7 ± 2.1
</oasis:entry><oasis:entry colname="4">2.3 ± 0.6
</oasis:entry><oasis:entry colname="5">0.1 ± 0.1
</oasis:entry><oasis:entry colname="6"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="7"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="8"><italic toggle="yes">b</italic></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1h</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">29.0 ± 1.7
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">0.7 ± 0.1
</oasis:entry><oasis:entry colname="6"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="7">>1000
</oasis:entry><oasis:entry colname="8"><italic toggle="yes">b</italic></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1i</bold></oasis:entry><oasis:entry colname="2">5.0 ± 0.1
</oasis:entry><oasis:entry colname="3">4.0 ± 1.0
</oasis:entry><oasis:entry colname="4">3.7 ± 0.6
</oasis:entry><oasis:entry colname="5">0.8 ± 0.1
</oasis:entry><oasis:entry colname="6">4.0 ± 0.1
</oasis:entry><oasis:entry colname="7">>600
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1j</bold></oasis:entry><oasis:entry colname="2">16.0 ± 0.1
</oasis:entry><oasis:entry colname="3">17.0 ± 1.0
</oasis:entry><oasis:entry colname="4">15.3 ± 0.6
</oasis:entry><oasis:entry colname="5">4.0 ± 1.0
</oasis:entry><oasis:entry colname="6">16.3 ± 0.6
</oasis:entry><oasis:entry colname="7">>600
</oasis:entry><oasis:entry colname="8"><italic toggle="yes">b</italic></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1k</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="7">>1000
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1l</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="7"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="8"><italic toggle="yes">b</italic></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1m</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">28.3 ± 3.5
</oasis:entry><oasis:entry colname="4">17.0 ± 6.2
</oasis:entry><oasis:entry colname="5">1.0 ± 0.1
</oasis:entry><oasis:entry colname="6">1.0 ± 0.1
</oasis:entry><oasis:entry colname="7">>500
</oasis:entry><oasis:entry colname="8">−
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1n</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">14.0 ± 2.6
</oasis:entry><oasis:entry colname="4">6.7 ± 1.1
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6">>32
</oasis:entry><oasis:entry colname="7">>200
</oasis:entry><oasis:entry colname="8"><italic toggle="yes">b</italic></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1o</bold></oasis:entry><oasis:entry colname="2">11.0 ± 5.7
</oasis:entry><oasis:entry colname="3">16.3 ± 1.2
</oasis:entry><oasis:entry colname="4">14.7 ± 4.9
</oasis:entry><oasis:entry colname="5">4.0 ± 0.1
</oasis:entry><oasis:entry colname="6">>32
</oasis:entry><oasis:entry colname="7">>600
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>10q</bold></oasis:entry><oasis:entry colname="2">>32
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="7">>1000
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1r</bold></oasis:entry><oasis:entry colname="2">5.0 ± 0.1
</oasis:entry><oasis:entry colname="3">17.7 ± 0.6
</oasis:entry><oasis:entry colname="4">12.3 ± 4.2
</oasis:entry><oasis:entry colname="5">1.3 ± 0.6
</oasis:entry><oasis:entry colname="6">4.0 ± 0.1
</oasis:entry><oasis:entry colname="7">>1000
</oasis:entry><oasis:entry colname="8">−
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1s</bold></oasis:entry><oasis:entry colname="2">16.0 ± 0.1
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">16.7 ± 0.6
</oasis:entry><oasis:entry colname="6">15.7 ± 0.6
</oasis:entry><oasis:entry colname="7">>600
</oasis:entry><oasis:entry colname="8">−
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1u</bold></oasis:entry><oasis:entry colname="2">16.0 ± 0.1
</oasis:entry><oasis:entry colname="3">>32
</oasis:entry><oasis:entry colname="4">>32
</oasis:entry><oasis:entry colname="5">>32
</oasis:entry><oasis:entry colname="6">16.3 ± 0.6
</oasis:entry><oasis:entry colname="7">>400
</oasis:entry><oasis:entry colname="8"><italic toggle="yes">b</italic></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1v</bold></oasis:entry><oasis:entry colname="2">3.5 ± 0.7
</oasis:entry><oasis:entry colname="3">3.3 ± 0.6
</oasis:entry><oasis:entry colname="4">1.7 ± 0.6
</oasis:entry><oasis:entry colname="5">0.4 ± 0.1
</oasis:entry><oasis:entry colname="6">1.0 ± 0.1
</oasis:entry><oasis:entry colname="7">150.3 ± 4.0
(37% inhibition)
</oasis:entry><oasis:entry colname="8">−
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1w</bold></oasis:entry><oasis:entry colname="2">18.5 ± 0.7
</oasis:entry><oasis:entry colname="3">30.0 ± 3.5
</oasis:entry><oasis:entry colname="4">4.7 ± 0.6
</oasis:entry><oasis:entry colname="5">27.3 ± 4.5
</oasis:entry><oasis:entry colname="6">>32
</oasis:entry><oasis:entry colname="7">>600
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1x</bold></oasis:entry><oasis:entry colname="2">15.5 ± 0.7
</oasis:entry><oasis:entry colname="3">21.7 ± 4.0
</oasis:entry><oasis:entry colname="4">4.7 ± 0.6
</oasis:entry><oasis:entry colname="5">9.3 ± 1.5
</oasis:entry><oasis:entry colname="6">17.0 ± 2.0
</oasis:entry><oasis:entry colname="7">>700
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1y</bold></oasis:entry><oasis:entry colname="2">18.0 ± 0.1
</oasis:entry><oasis:entry colname="3">16.3 ± 0.6
</oasis:entry><oasis:entry colname="4">15.7 ± 1.5
</oasis:entry><oasis:entry colname="5">16.7 ± 1.5
</oasis:entry><oasis:entry colname="6">>32
</oasis:entry><oasis:entry colname="7">>500
</oasis:entry><oasis:entry colname="8">+
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1">cryptolepine
</oasis:entry><oasis:entry colname="2">1.5 ± 0.7
</oasis:entry><oasis:entry colname="3">2.3 ± 0.6
</oasis:entry><oasis:entry colname="4">2.0 ± 0.1
</oasis:entry><oasis:entry colname="5">3.0 ± 0.1
</oasis:entry><oasis:entry colname="6"><italic toggle="yes">b</italic></oasis:entry><oasis:entry colname="7">65.7 ± 3.0
</oasis:entry><oasis:entry colname="8">+</oasis:entry></oasis:row></oasis:tbody></oasis:tgroup></oasis:table><table-wrap-foot><p><italic toggle="yes"><sup>a</sup></italic><sup></sup> +, inhibition of β-hematin formation; −, no inhibition of β-hematin formation.<italic toggle="yes"><sup>b</sup></italic><sup></sup> Not tested.</p></table-wrap-foot></table-wrap></p><p><bold>(b) DNA Interactions and Inhibition of </bold><bold>β</bold><bold>-Hematin Formation.</bold> All compounds were also evaluated in
functional assays related to possible mechanisms of
action. The first mechanism of action to be proposed for
cryptolepine as an antimalarial agent was intercalation
into DNA, like 9-aminoacridine,<xref rid="jm011102ib00004" ref-type="bibr"></xref> but conflicting results
have been published.<xref rid="jm011102ib00023" ref-type="bibr"></xref> For chloroquine, an aminoquinoline, the hypothesis that interaction with DNA is
responsible for the antiplasmodial effect has been
abandoned. Recent hypotheses propose that quinoline
antimalarials, such as quinine and chloroquine, active
against the intraerythrocytic stages of the <italic toggle="yes">Plasmodium</italic>
parasite act by interfering with the digestion of hemoglobin in the acid food vacuole of the <italic toggle="yes">Plasmodium</italic>
parasite. While the globin part is used as a source of
amino acids, the heme part is toxic to the parasite and
is therefore converted to an insoluble crystalline substance known as hemozoin or malaria pigment. The
latter process is thought to be inhibited by the quinoline
antimalarials.<xref rid="jm011102ib00001" ref-type="bibr"></xref> It has been shown that the corresponding in vitro process, the spontaneous polymerization of
hematin to β-hematin in cell-free systems, is also
inhibited by quinoline antimalarials such as chloroquine
and quinine.<xref rid="jm011102ib00024" ref-type="bibr"></xref> For chloroquine, it has also been claimed
that depolymerization of malarial hemozoin, and/or
inhibition of the glutathione-dependent degradation of
heme in the cytosol of the parasite, may be responsible
in part for its activity.<named-content content-type="bibref-group"><xref rid="jm011102ib00025" ref-type="bibr"></xref>,<xref rid="jm011102ib00026" ref-type="bibr"></xref></named-content> Cryptolepine, similarly to
chloroquine, was able to inhibit the formation of β-hematin in vitro, and it was proposed that this mechanism
is likely to be responsible, at least in part, for its
antiplasmodial activity.<named-content content-type="bibref-group"><xref rid="jm011102ib00011" ref-type="bibr"></xref>,<xref rid="jm011102ib00027" ref-type="bibr"></xref></named-content></p><p>Taking this into account, all compounds synthesized
were evaluated not only for their inhibiting effect on
the polymerization of hematin to β-hematin in cell-free
systems but also in the DNA−methyl green assay, a
colorimetric microassay for the detection of agents that
interact with DNA.<xref rid="jm011102ib00028" ref-type="bibr"></xref> Compounds interacting with DNA
are able to displace methyl green from a methyl green−DNA complex, leading to a loss of color and a decrease
in absorbance. IC<sub>50</sub> values obtained in this assay (i.e.,
the concentration of test compound leading to a 50%
decrease of absorbance, corresponding to 50% displacement of the dye from the DNA complex), together with
the effect on the formation of polymerized β-hematin,
are displayed in Table <xref rid="jm011102it00003"></xref>. Cryptolepine, neocryptolepine
(<bold>1a</bold>), and many of the substituted neocryptolepine
derivatives are able to inhibit the formation of β-hematin in cell-free systems. For some compounds, most
notably those lacking the <italic toggle="yes">N</italic>-methyl group (compounds
<bold>10b</bold>, <bold>10d</bold>, <bold>10e</bold>, and <bold>10q</bold>), this is not accompanied by an
antiplasmodial activity, so obviously the ability to
inhibit β-hematin formation is not enough to show in
vitro antiplasmodial activity. This may be related to the
presence of the compound at the site of action, i.e., the
heme detoxification process in the plasmodial food
vacuole. Only four compounds show affinity for DNA
in the concentration range tested: cryptolepine, being
the most potent one (IC<sub>50</sub> 65.7 ± 3.0 μM), followed by
2-methoxyneocryptolepine (<bold>1b</bold>) (77.9 ± 4.4 μM) and
neocryptolepine (<bold>1a</bold>) (92.8 ± 9.7 μM); for 3-methoxyneocryptolepine (<bold>1v</bold>) 37% displacement was observed at
a concentration of 150 μM. Cryptolepine and neocryptolepine (<bold>1a</bold>), but not 2-methoxy- (<bold>1b</bold>) or 3-methoxyneocryptolepine (<bold>1v</bold>), are capable of inhibiting the
formation of β-hematin. Nevertheless, the last two
compounds also show a higher antiplasmodial activity
than neocryptolepine (<bold>1a</bold>) itself.
</p><p>From these data, it is obvious that there are at least
two different targets for the antiplasmodial activity of
cryptolepine, neocryptolepine, and the neocryptolepine
derivatives synthesized, and some typical compounds
can be selected. Neocryptolepine derivatives, such as
2-bromoneocryptolepine (<bold>1c</bold>), are more antiplasmodially
active than neocryptolepine (<bold>1a</bold>) itself, are able to inhibit
the formation of β-hematin, and show a reduced or no
cytotoxicity and no DNA interactions in the concentration range tested. Their antiplasmodial activity is likely
to be due to a specific mechanism of action (inhibition
of the heme detoxification process in the parasite). On
the other hand, a compound such as 2-methoxyneocryptolepine (<bold>1b</bold>) is also more antiplasmodially active
than neocryptolepine (<bold>1a</bold>) but is not able to inhibit
formation of β-hematin. However, it shows affinity for
DNA, and hence, it is also cytotoxic. The antiplasmodial
activity is not due to a selective mechanism of action.
Other compounds, and most notably cryptolepine itself,
show inhibition of β-hematin formation as well as DNA
interactions and most probably have at least two targets
in the <italic toggle="yes">Plasmodium</italic> parasite: the heme detoxification
process and DNA-containing structures. To support this
hypothesis, more detailed experiments were performed
for some selected compounds, i.e., cryptolepine, neocryptolepine (<bold>1a</bold>), 2-methoxy- (<bold>1b</bold>), 2-bromo- (<bold>1c</bold>), and
2-chloroneocryptolepine (<bold>1d</bold>) (see below).
</p><p>Earlier work on synthetic indoloquinolines has already led to compounds with a higher antiplasmodial activity than cryptolepine, such as 11-methylcryptolepine<sup>9</sup> and 2,7-dibromocryptolepine.<xref rid="jm011102ib00011" ref-type="bibr"></xref> However, 11-methylcryptolepine was also more cytotoxic than cryptolepine. Although the antiplasmodial activity of 2,7-dibromocryptolepine was confirmed in in vivo experiments in mice infected with <italic toggle="yes">Plasmodium berghei</italic> without apparent toxicity to the host, this compound also
showed slightly higher in vitro cytotoxicity than cryptolepine, which may limit its clinical usefulness. δ-Carbolines such as 1-methyl-δ-carboline showed a selective
antiplasmodial activity and no cytotoxicity on L6 cells.<xref rid="jm011102ib00009" ref-type="bibr"></xref>
This compound was found to be specifically localized in
a parasite structure that could correspond to the
parasite nucleus. Inhibition of β-hematin formation was
not evaluated. Although in the present work we have
not been able to design neocryptolepine derivatives with
a higher antiplasmodial activity than cryptolepine, it
has been possible to improve the antiplasmodial activity
of neocryptolepine while at the same time reducing its
cytotoxicity, i.e., to obtain more selective antiplasmodial
compounds.
</p><p><bold>(c) DNA Binding. Electric Linear Dichroism
(ELD) Measurements.</bold> It is now well established that
cryptolepine and related indoloquinolines (in particular
ellipticines) interact with DNA as intercalators.<named-content content-type="bibref-group"><xref rid="jm011102ib00007" ref-type="bibr"></xref>,<xref rid="jm011102ib00029" ref-type="bibr"></xref></named-content>
Neocryptolepine (<bold>1a</bold>) also intercalates into DNA but
exhibits a reduced affinity for DNA compared to cryptolepine.<xref rid="jm011102ib00008" ref-type="bibr"></xref> Electric linear dichroism (ELD) experiments
provide firm evidence that the newly designed neocryptolepine analogues also behave as typical DNA intercalating agents. Figure <xref rid="jm011102if00003"></xref>a shows the ELD spectra for
2-methoxy- (<bold>1b</bold>), 2-bromo- (<bold>1c</bold>), and 2-chloroneocryptolepine (<bold>1d</bold>) bound to calf thymus DNA. Their mode of
binding to DNA was analyzed on the basis of the highest
ELD values obtained when the drug molecules are fully
bound to DNA, i.e., for P/D ratios of 20. In each case,
the reduced dichroism is negative in sign in the 300−400 nm region corresponding to the absorption band of
the indoloquinoline unit. The reduced dichroism measured for the drug−DNA complexes at 320−360 nm is
almost identical to that obtained with DNA alone at 260
nm (Figure <xref rid="jm011102if00003"></xref>b). This implies that the drug is oriented
parallel to the DNA base pairs, as expected for an
intercalation binding mode. ELD defines the orientation
of drug bound to DNA, whereas the DNA−methyl green
assay defines the relative affinity for DNA. 2-Chloro-
(<bold>1d</bold>) and especially 2-bromoneocryptolepine (<bold>1c</bold>) have
a low affinity for DNA, which is in agreement with their
reduced cytotoxicity, whereas 2-methoxyneocryptolepine
(<bold>1b</bold>) has a higher affinity for DNA.
<fig id="jm011102if00003" position="float" orientation="portrait"><label>3</label><caption><p>(a) ELD spectra for (○) 2-methoxy-, (□) 2-bromo-, (▪) 2-chloro-, and (▴) neocryptolepine bound to calf thymus DNA. (b) Dependence of the reduced dichroism Δ<italic toggle="yes">A</italic>/<italic toggle="yes">A</italic> on the
electric field strength for the different drug−DNA complexes
at 350 nm and for DNA alone at 260 nm (·). ELD measurements were performed at a P/D ratio of 20 (200 μM DNA, 10
μM drug) in 1 mM sodium cacodylate buffer, pH 7.0.</p></caption><graphic xlink:href="jm011102if00003.tif" position="float" orientation="portrait"></graphic></fig></p><p><bold>(d) Inhibition of Topoisomerase II.</bold> In contrast to
cryptolepine, which is a DNA intercalator and a potent
poison for topoisomerase II, neocryptolepine poorly
inhibits the DNA relaxing enzyme. We have shown
recently that cryptolepine is about 5 times more potent
than neocryptolepine at inhibiting the unwinding of
closed circular duplex DNA by human topoisomerase II.<xref rid="jm011102ib00008" ref-type="bibr"></xref>
Both DNA relaxation and cleavage assays with plasmid
DNA and <sup>32</sup>P-labeled DNA restriction fragments, respectively, were performed to evaluate the effects of the
2-substituted neocryptolepine analogues on topoisomerase
II activity (human p170 isoform), but none of them
showed an effect superior to that of neocryptolepine. In
the presence of 2-methoxy (<bold>1b</bold>), 2-bromo- (<bold>1c</bold>), or 2-chloroneocryptolepine (<bold>1d</bold>) (20 μM each), topoisomerase II
mediated DNA cleavage was found to be slightly stimulated, but in all cases, the extent of DNA cleavage never
exceeded 15% of the DNA products (data not shown).
As discussed previously, we consider that topoisomerase
II may not represent the essential target for the cytotoxicity of neocryptolepine.<xref rid="jm011102ib00029" ref-type="bibr"></xref> However, it is important
to bear in mind that these experiments were performed
with commercially available human topoisomerase II
and it remains possible that these compounds selectively
interfere with the activity of <italic toggle="yes">Plasmodium</italic> topoisomerase
II. For some compounds known as inhibitors of topoisomerase II, antiplasmodial activity has been reported,
e.g., the fluoroquinolone antibiotics, which are gyrase
(bacterial, prokaryotic topoisomerase II) inhibitors.<xref rid="jm011102ib00030" ref-type="bibr"></xref> For
<italic toggle="yes">P. falciparum</italic>, two distinct topoisomerase II activities
have been demonstrated: a nuclear eukaryotic type II
and a bacterial (prokaryotic) type II topoisomerase
activity associated with plastid DNA replication.<xref rid="jm011102ib00031" ref-type="bibr"></xref> This
plastid is an atypical organelle essential for parasite
survival and represents an effective target for parasitic
drug design.<xref rid="jm011102ib00032" ref-type="bibr"></xref> Therefore, it would be quite useful to
compare the effects of the test drugs on mammalian
versus plasmodial topoisomerase II. This hypothesis will
be explored in our laboratories.
</p><p><bold>(e) Characterization of Heme</bold><bold>:</bold><bold> Gas-Phase Ion
Drug Complexes by </bold><bold>Electrospray Ionization (ESI)
Mass Spectrometry.</bold> Recently, Wright et al.<xref rid="jm011102ib00033" ref-type="bibr"></xref> have
described a mass spectrometric method to detect complexes formed between a test compound and heme,
which was adapted in our laboratory and applied to
cryptolepine, neocryptolepine (<bold>1a</bold>), and its 2-methoxy-
(<bold>1b</bold>), 2-bromo- (<bold>1c</bold>), and 2-chloro- (<bold>1d</bold>) derivatives.
Figure <xref rid="jm011102if00004"></xref>a illustrates the ESI mass spectrum obtained
for a mixture containing 40 μM heme and 100 μM
cryptolepine in MeOH/H<sub>2</sub>O (1:1; v/v; pH 7). Under these
conditions, stable gas-phase cryptolepine complexes
were detected at <italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 848, <italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 1463, and <italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 1481,
corresponding to the complex of the drug with the heme
monomer [FP:C]<sup>+</sup>, the heme dimer [(2FP − H):C]<sup>+</sup>, and
the dehydrated dimer of hematin [(FP−O−FP):C]<sup>+</sup>
respectively. For neocryptolepine and its derivatives,
only the complex with the heme monomer [FP:C]<sup>+</sup> and
the dimer [(2PF − H):C]<sup>+</sup> could be detected (data not
shown). The relative strength of binding between the
drugs and the monomeric and dimeric heme forms was
assessed using low-energy collision-induced dissociation.
Figure <xref rid="jm011102if00004"></xref>b illustrates the collision-induced dissociation
(CID) spectra obtained for the complexes between the
heme monomer and cryptolepine, neocryptolepine (<bold>1a</bold>),
and 2-bromoneocryptolepine (<bold>1c</bold>). Ions corresponding to
the drug complexes with the heme monomer [FP:C]<sup>+</sup>
were selected and collided with helium. In each case,
an FP<sup>+</sup> peak (<italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 616) was produced because of loss of
the drug that, relative to the precursor [FP:C]<sup>+</sup> peak,
was about 12-fold weaker with cryptolepine than with
neocryptolepine, indicating a higher binding capacity of
cryptolepine. On the basis of the CID spectral data, the
order of stability for [FP:C]<sup>+</sup> complexes of neocryptolepine and its derivatives was 2-methoxy > 2-Cl >
2-Br > neocryptolepine (the ion intensity ratios FP<sup>+</sup>/[FP:C]<sup>+</sup> were 0.81, 0.88, 0.98, and 1.22, respectively;
relative standard deviation (RSD) = 0.02). The order of
stability for the complexes with the heme dimer [(2FP
− H):C]<sup>+</sup> of neocryptolepine and its derivatives inferred
from the CID spectral data was 2-methoxy > 2-Br >
neocryptolepine > 2-Cl (the ion intensity ratios [2FP −
H]<sup>+</sup>/[(2FP − H):C]<sup>+</sup> were 0.12, 0.24, 0.25, and 0.27,
respectively; RSD = 0.02). These data demonstrate that
neocryptolepine and its derivatives show a stronger
binding to the dimeric heme form than to the monomeric
one. Apparently all selected compounds were able to
form complexes with the monomeric and dimeric forms
of heme, but 2-methoxyneocryptolepine (<bold>1b</bold>) was not
able to inhibit the polymerization of hematin to β-hematin in cell-free systems as described above. Whereas
complex formation as detected by MS may be a prerequisite for inhibition of β-hematin formation, it does not
necessarily indicate that the test compound is actually
capable of inhibiting this polymerization reaction.
<fig id="jm011102if00004" position="float" orientation="portrait"><label>4</label><caption><p>(a) ESI mass spectrum of a mixture of 40 μM heme and 100 μM cryptolepine in MeOH/H<sub>2</sub>O (1:1). The ion at <italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 1253
corresponds to the complex consisting of the heme dimer and methanol [(2FP−H):MeOH]<sup>+</sup>. (b) CID spectra obtained for the complex
consisting of the heme monomer with cryptolepine, neocryptolepine (<bold>1a</bold>), and 2-bromoneocryptolepine (<bold>1c</bold>).
</p></caption><graphic xlink:href="jm011102if00004.tif" position="float" orientation="portrait"></graphic></fig></p><p><bold>(f) Antitrypanosomal Activity.</bold> Because of the
antitrypanosomal activity of cryptolepine,<xref rid="jm011102ib00009" ref-type="bibr"></xref> all compounds synthesized were also tested against <italic toggle="yes">T. brucei</italic>
and <italic toggle="yes">T. </italic><italic toggle="yes">cruzi</italic> (Table <xref rid="jm011102it00003"></xref>). Cryptolepine showed an IC<sub>50</sub>
against <italic toggle="yes">T. brucei</italic> of 3.0 μM, but this is obviously not
due to a selective mechanism of action, in view of its
antiplasmodial and cytotoxic activity. The same is true
for a series of other compounds, most notably 2-methylneocryptolepine (<bold>1g</bold>), with an IC<sub>50</sub> value against <italic toggle="yes">T.
brucei</italic> of 0.1 μM and also a significant cytotoxicity (IC<sub>50</sub>
on MRC-5 cells of 0.95 μM). More specific antitrypanosomal activity against at least one of the two <italic toggle="yes">Trypanosoma</italic> species tested, combined with absence of obvious
cytotoxicity in the concentration range tested, is observed for 2-bromo- (<bold>1c</bold>), 2-nitro- (<bold>1h</bold>), and 2-methoxy-9-cyanoneocryptolepine (<bold>1m</bold>). The last compound showed
an IC<sub>50</sub> of 1.0 μM against <italic toggle="yes">T. brucei</italic> as well as <italic toggle="yes">T. cruzi</italic>,
whereas the IC<sub>50</sub> on the MRC-5 cells was >32 μM.
Therefore, this neocryptolepine derivative could be
considered as a new lead structure with antitrypanosomal potential. Additional exploration in this area is
warranted.
</p></sec><sec id="d7e2524"><title>Conclusion</title><p>In conclusion, some new neocryptolepine (indolo[2,3-<italic toggle="yes">b</italic>]quinoline) derivatives were synthesized with a more
selective antiplasmodial activity than the original lead.
This activity is most probably due to inhibition of the
heme detoxification process (formation of hemozoin), as
demonstrated by the investigation of heme complexation and the polymerization of hematin to β-hematin
in cell-free systems. 2-Bromoneocryptolepine (<bold>1c</bold>) was
the most selective compound with an IC<sub>50</sub> value against
chloroquine-resistant <italic toggle="yes">P. falciparum</italic> of 4.0 μM, in the
absence of obvious cytotoxicity (IC<sub>50</sub> > 32 μM). Although
this was higher than the IC<sub>50</sub> value of cryptolepine (2.0
μM), 2-bromoneocryptolepine showed a low affinity for
DNA, and no inhibition of human topoisomerase II.
Although some neocryptolepine derivatives with an in
vitro antiplasmodial activity higher than that of 2-bromoneocryptolepine were obtained, these compounds also
showed a higher affinity for DNA and/or a more
pronounced cytotoxicity on a human cell line, as observed for cryptolepine as well. From the present work,
2-bromoneocryptolepine is therefore considered as the
most promising lead for potentially new antimalarial
agents. In addition, 2-bromo-, 2-nitro-, and 2-methoxy-9-cyanoneocryptolepine derivatives exhibited some antitrypanosomal activity and could be considered as new
lead structures for antitrypanosomal agents.
</p></sec><sec id="d7e2548"><title>Experimental Section</title><p><bold>General Experimental Procedures.</bold> NMR spectra were
recorded on Varian Unity and Bruker DRX-400 spectrometers,
operating at 400 MHz for <sup>1</sup>H and 100 MHz for <sup>13</sup>C and with
CDCl<sub>3</sub> as the solvent unless otherwise stated. In every case
tetramethylsilane was used as the internal standard. Chemical
shifts are given in ppm and <italic toggle="yes">J</italic> values in Hz. Multiplicity is
indicated using the following abbreviations: d for a doublet, t
for a triplet, m for a multiplet, etc. In those cases where
standard <sup>1</sup>H and <sup>13</sup>C measurements were insufficient, additional 2D measurements (i.e., COSY, HETCOR or HSQC,
LR-HETCOR or HMBC) were performed for complete structure elucidation. IR spectra were recorded on a Bruker Vector
22 spectrometer. Low-resolution mass spectra were recorded
on a triple quadrupole mass spectrometer (Quattro II, Micromass, Manchester, U.K.) using electrospray ionization (ESI).
Samples were dissolved in CH<sub>3</sub>CN and infused at 5 μL/min
into the mass spectrometer. Fragmentation was induced by
low-energy collisional activation using an Ar gas pressure of
approximately 10<sup>-3</sup> mbar and a collision energy between 25
and 35 eV depending on the compound. Exact mass measurements were performed on a quadrupole/orthogonal-acceleration
time-of-flight (Q/oaTOF) tandem mass spectrometer (qTof 2,
Micromass, Manchester, U.K.) equipped with a standard
electrospray ionization (ESI) interface. Samples were diluted
in an appropriate solvent (MeOH/0.1% formic acid 90/10), and
1 μL aliquots were injected at a flow rate of 5 μL/min using
the CapLC system (Waters, Brussels, Belgium). The mass
scale was corrected by an internal lock-mass provided by the
protonated molecule of 2‘-deoxyadenosine (<italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 252.1096),
which was added into the LC stream at a flow rate of 2.5 μL/min (concentrated 10<sup>-5</sup> M using the same solvent composition)
using a low dead volume tee union. Combined spectra of the
compound were processed using the Masslynx software suite
(Micromass, Manchester, U.K.). Melting points were determined on a Büchi B-545 apparatus and are uncorrected. All
reagents were purchased from commercial sources (Acros,
Aldrich, Lancaster) and were used as such. THF was distilled
from sodium benzophenone. Column chromatography was
performed on Kieselgel 60 (Merck), 0.040−0.063 mm. Thin-layer chromatography was performed on precoated ALUGRAM
SIL G/UV<sub>254</sub> Kieselgel 60 0.25 mm TLC plates. For the sake
of brevity, only the data for complete characterization of the
final products (i.e., the new neocryptolepine derivatives) are
given. Characterization data for the intermediate compounds
are supplied in the Supporting Information. Analytical HPLC
using two independent systems was performed to check the
purity of the products: (column A) Merck, Lichrospher Si 600
(4 mm × 250 mm); (column B) Merck, Lichrocart-Lichrospher
100 RP 18 (4 mm × 250 mm). Also, two different solvent
systems were used: (system A) 1-propanol−hexane 5%/water
95:5 or 1-propanol−hexane 5%/water 90:10; (system B) gradient from 80% to 100% acetonitrile (ACN) in 20 min. Experimental details of the HPLC analysis are reported for all test
compounds in the following form: column, mobile phase,
retention time. The flow rate in system A was 0.65 mL/min,
and the flow rate was 1.0 mL/min in system B.
</p><p><bold>4-Amino-3-iodobenzenecarbonitrile (2</bold><bold>‘</bold><bold>). </bold>4-Aminobenzonitrile (1.0 g, 8.47 mmol) is dissolved in 20 mL of dry
methanol while stirring. The flask is cooled to 0 °C, after which
CaCO<sub>3</sub> (2.5 g, 25 mmol) is added. Then a solution of ICl in
HCl (4 mL, 5 M ICl in concentrated HCl) is added dropwise
in 10 min while maintaining the temperature at 0−5 °C. After
complete addition, the cooling bath is removed and the mixture
is stirred for 30 min at room temperature and further for 2
days at 35 °C. After completion of the reaction, 60 mL of a 0.5
M Na<sub>2</sub>S<sub>2</sub>O<sub>3</sub> solution is added followed by 100 mL of water. The
mixture is extracted with CH<sub>2</sub>Cl<sub>2</sub> (2 × 80 mL), and after
removal of the solvent the crude product was purified by
column chromatography with CH<sub>2</sub>Cl<sub>2</sub> as the eluent.
</p><p><bold>2-(2-Trimethylsilylethynyl)aniline (3). </bold>Pd(PPh<sub>3</sub>)<sub>2</sub>Cl<sub>2</sub> (280
mg, 0.40 mmol) and CuI (64 mg, 0.33 mmol) are added to a
stirred solution of 2-iodoaniline (8.76 g, 0.04 mol) in Et<sub>3</sub>N (100
mL). To this suspension, trimethylsilylacetylene (6 mL, 0.43
mol) is added, and the resulting mixture is stirred for 18 h at
room temperature. Then 100 mL of water is added and the
mixture is extracted with CH<sub>2</sub>Cl<sub>2</sub> (3 × 50 mL). The solvent is
removed in vacuo, and the product is purified by column
chromatography using CH<sub>2</sub>Cl<sub>2</sub> as the eluent.
</p><p><bold>4-Amino-3-(2-trimethylsilylethynyl)benzenecarbonitrile (3</bold><bold>‘</bold><bold>). </bold>To a stirred solution of 4-amino-3-iodobenzenecarbonitrile (0.60 g, 2.5 mmol) (<bold>2</bold>‘) in THF (10 mL), Et<sub>3</sub>N (5
mL), Pd(PPh<sub>3</sub>)<sub>2</sub>Cl<sub>2</sub> (17.2 mg, 0.024 mmol), and CuI (4.2 mg,
0.022 mmol) are added. To this solution trimethylsilylacetylene
(0.36 mL, 2.6 mmol) is added, and the resulting mixture is
stirred for 24 h at room temperature. Then 20 mL of water is
added and the mixture is extracted with CH<sub>2</sub>Cl<sub>2</sub> (3 × 30 mL).
The solvent is removed in vacuo, and the product is purified
by column chromatography using heptane/ether (1:3) as the
eluent.
</p><p><bold>3-(2-Trimethylsilylethynyl)-4-triphenylphosphoranylideneaminobenzenecarbonitrile </bold><bold>(5</bold><bold>‘</bold><bold>).</bold> A flask is charged
with 4-amino-3-(2-trimethylsilylethynyl)benzenecarbonitrile
(<bold>3</bold>‘) (1.30 g, 6.07 mmol), triphenylphosphine (3.28 g, 12.5
mmol), Et<sub>3</sub>N (12 mL), CCl<sub>4</sub> (7 mL), and CH<sub>3</sub>CN (15 mL). The
resulting mixture is stirred at room temperature for 24 h.
Then, 20 mL of MeOH is added and stirring is continued for
30 min. After the addition of 100 mL of water, the mixture is
extracted with CH<sub>2</sub>Cl<sub>2</sub> (3 × 50 mL), after which the solvent is
removed. The product is then purified by column chromatography using EtOAc/MeOH (98:2) as the eluent.
</p><p><bold>General Procedure for the Preparation of Thioureas
5a</bold><bold>−</bold><bold>j, 5p, and 5q. </bold>2-(2-Trimethylsilylethynyl)aniline (<bold>3</bold>) (1.2
g, 6.35 mmol) and 6.5 mmol of the appropriate 4-substituted
phenyl isothiocyanate (<bold>4a</bold><bold>−</bold><bold>j</bold>) are dissolved in EtOH (25 mL).
A few crystals of DMAP (<italic toggle="yes">N,N</italic>-(dimethylamino)pyridine) are
added, and the resulting solution is stirred at 40 °C in an oil
bath. After 1−2 h, a white precipitate is formed. The solid is
filtered off and washed with 20 mL of cold EtOH and
subsequently dried in vacuo to yield the corresponding thiourea. The filtrate is stirred further at 40 °C, giving another
amount of precipitate after a few hours. The following products
were prepared in this manner:
</p><p><italic toggle="yes">N</italic>-phenyl-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5a</bold>)
</p><p><italic toggle="yes">N</italic>-(4-methoxyphenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5b</bold>)
</p><p><italic toggle="yes">N</italic>-(4-bromophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5c</bold>)
</p><p><italic toggle="yes">N</italic>-(4-chlorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5d</bold>)
</p><p><italic toggle="yes">N</italic>-(4-Fluorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5e</bold>)
</p><p><italic toggle="yes">N</italic>-(4-iodophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5f</bold>)
</p><p><italic toggle="yes">N</italic>-(4-methylphenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5g</bold>)
</p><p><italic toggle="yes">N</italic>-(4-isothiocyanatophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5h</bold>)
</p><p><italic toggle="yes">N</italic>-(4-methylthiophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5i</bold>)
</p><p><italic toggle="yes">N</italic>-(4-cyanophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5j</bold>)
</p><p><italic toggle="yes">N</italic>-(1-naphthyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5p</bold>)
</p><p><italic toggle="yes">N</italic>-(2,4-dichlorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]thiourea (<bold>5q</bold>)
</p><p><bold>General Procedure for the Synthesis of Carbodiimides 7a</bold><bold>−</bold><bold>j, 7p, and 7q from Thioureas </bold><bold>5a</bold><bold>−</bold><bold>j, 5p, and 5q.</bold> A
two-necked flask equipped with a drying tube is charged with
the appropriate thiourea (2.13 mmol). Dry CH<sub>2</sub>Cl<sub>2</sub> (25 mL) was
added, followed by Et<sub>3</sub>N (0.59 mL, 4.25 mmol) and a catalytic
amount of DMAP. The solution is stirred, and CH<sub>3</sub>SO<sub>2</sub>Cl (0.5
mL, 6.40 mmol) is added dropwise. It is important that the
rate of addition is kept low because of the very exothermic
nature of the reaction. After complete addition, the solution
is stirred for 15 min at room temperature, after which 50 mL
of water is added. The mixture is extracted with CH<sub>2</sub>Cl<sub>2</sub> (3 ×
30 mL), and the solvent is removed on a rotavapor. Purification
of the product using column chromatography (CH<sub>2</sub>Cl<sub>2</sub> as
eluent) yields the carbodiimide as a viscous oil. The product
is used immediately in the next step to prevent degradation.
The following products were prepared in this manner:
</p><p><italic toggle="yes">N</italic>-phenyl-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7a</bold>)
</p><p><italic toggle="yes">N</italic>-(4-methoxyphenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7b</bold>)
</p><p><italic toggle="yes">N</italic>-(4-bromophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7c</bold>)
</p><p><italic toggle="yes">N</italic>-(4-chlorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7d</bold>)
</p><p><italic toggle="yes">N</italic>-(4-fluorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7e</bold>)
</p><p><italic toggle="yes">N</italic>-(4-iodophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7f</bold>)
</p><p><italic toggle="yes">N</italic>-(4-methylphenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7g</bold>)
</p><p><italic toggle="yes">N</italic>-(4-isothiocyanatophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7h</bold>)
</p><p><italic toggle="yes">N</italic>-(4-methylthiophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7i</bold>)
</p><p><italic toggle="yes">N</italic>-(4-cyanophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7j</bold>)
</p><p><italic toggle="yes">N</italic>-(1-naphthyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7p</bold>)
</p><p><italic toggle="yes">N</italic>-(2,4-dichlorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7q</bold>)
</p><p><bold>General Procedure for the Synthesis of Carbodiimides 7k</bold><bold>−</bold><bold>o from Iminophosphorane </bold><bold>5</bold><bold>‘</bold><bold> and of 7r</bold><bold>−</bold><bold>u from
Iminophosphorane 5</bold><bold>‘</bold><bold>‘</bold><bold>.</bold> Iminophosphorane <bold>5</bold>‘ or <bold>5</bold>‘ ‘ (2.1
mmol) is dissolved in 20 mL of dry toluene. To this solution
the appropriate substituted phenyl isocyanate (2.2 mmol) is
added, and the resulting solution is stirred at 70 °C for 1 h.
The solvent is removed in vacuo, and the product is purified
by column chromatography using CH<sub>2</sub>Cl<sub>2</sub> as the eluent. The
product is used immediately in the next step. The following
products were prepared in this manner:
</p><p><italic toggle="yes">N</italic>-[4-cyano-2-(2-trimethylsilylethynyl)phenyl]-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-phenylcarbodiimide (<bold>7k</bold>)
</p><p><italic toggle="yes">N</italic>-(4-chlorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic><italic toggle="yes">-</italic>[4-cyano-2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7l</bold>)
</p><p><italic toggle="yes">N</italic>-[4-cyano-2-(2-trimethylsilylethynyl)phenyl]-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-(4-methoxyphenyl)carbodiimide (<bold>7m</bold>)
</p><p><italic toggle="yes">N</italic>-[4-cyano-2-(2-trimethylsilylethynyl)phenyl]-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-(4-trifluoromethylphenyl)carbodiimide (<bold>7n</bold>)
</p><p><italic toggle="yes">N</italic>-[4-cyano-2-(2-trimethylsilylethynyl)phenyl]-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-(4-fluorophenyl)carbodiimide (<bold>7o</bold>)
</p><p><italic toggle="yes">N</italic>-(3-methoxyphenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7r</bold>)
</p><p><italic toggle="yes">N</italic>-(3-bromophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7s</bold>)
</p><p><italic toggle="yes">N</italic>-(3-chlorophenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7t</bold>)
</p><p><italic toggle="yes">N</italic>-(3-trifluoromethylphenyl)-<italic toggle="yes">N</italic><italic toggle="yes">‘</italic>-[2-(2-trimethylsilylethynyl)phenyl]carbodiimide (<bold>7u</bold>)
</p><p><bold>General Procedure for the Conversion of Carbodiimides 7a</bold><bold>−</bold><bold>u into the Substituted </bold><bold>11-Trimethylsilyl-6</bold><bold><italic toggle="yes">H</italic></bold><bold>-indolo[2,3-</bold><bold><italic toggle="yes">b</italic></bold><bold>]quinolines (8a</bold><bold>−</bold><bold>y).</bold> This reaction has to be
performed using freshly prepared carbodiimides. The carbodiimide (approximately 2.0 mmol) is dissolved in 30 mL of
mesitylene. Helium gas is bubbled through the solution for
10 min. After the solution was degassed, 1,4-cyclohexadiene
(3 mL, 31.7 mmol) is added and the solution is refluxed for 18
h under a nitrogen atmosphere. The solution is cooled to room
temperature, and the solvent is removed under high vacuum.
Purification of the product is performed by column chromatography using CH<sub>2</sub>Cl<sub>2</sub>/EtOAc (7:3) as the solvent. The following products were prepared in this manner:
</p><p>11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8a</bold>)
</p><p>2-methoxy-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8b</bold>)
</p><p>2-bromo-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8c</bold>)
</p><p>2-chloro-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8d</bold>)
</p><p>2-fluoro-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8e</bold>)
</p><p>2-iodo-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8f</bold>)
</p><p>2-methyl-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8g</bold>)
</p><p>2-isothiocyanato-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8h</bold>)
</p><p>2-methylthio-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8i</bold>)
</p><p>2-cyano-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8j</bold>)
</p><p>9-cyano-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8k</bold>)
</p><p>2-chloro-9-cyano-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8l</bold>)
</p><p>9-cyano-2-methoxy-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8m</bold>)
</p><p>9-cyano-2-trifluoromethyl-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8n</bold>)
</p><p>9-cyano-2-fluoro-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8o</bold>)
</p><p>7-trimethylsilylnaphtho[1,2-<italic toggle="yes">b</italic>]-α-carboline (<bold>8p</bold>)
</p><p>2,4-dichloro-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8q</bold>)
</p><p>1-methoxy-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8r</bold>)
</p><p>1-bromo-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8s</bold>)
</p><p>1-trifluoromethyl-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8u</bold>)
</p><p>3-methoxy-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8v</bold>)
</p><p>3-bromo-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8w</bold>)
</p><p>3-chloro-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8x</bold>)
</p><p>3-trifluoromethyl-11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>8y</bold>)
</p><p><bold>General Procedure for the Conversion of the Substituted </bold><bold>11-Trimethylsilyl-6</bold><bold><italic toggle="yes">H</italic></bold><bold>-indolo[2,3-</bold><bold><italic toggle="yes">b</italic></bold><bold>]quinolines</bold> <bold>8b,
8d, 8e, 8q</bold> <bold>into the Desilylated </bold><bold>6</bold><bold><italic toggle="yes">H</italic></bold><bold>-Indolo[2,3-</bold><bold><italic toggle="yes">b</italic></bold><bold>]quinolines
10b, 10d, 10e, 10q.</bold> The appropriate 11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (0.35 mmol) is dissolved in ethanol (30
mL), after which NaOH (10 mL, 6 N) is added. The mixture is
stirred for 3 h at 70 °C and cooled to room temperature. Water
(50 mL) is added, and the mixture is extracted with CH<sub>2</sub>Cl<sub>2</sub>
(3 × 50 mL). Purification of the product is performed by column
chromatography using CH<sub>2</sub>Cl<sub>2</sub>/EtOAc (7:3) as the eluent. The
following products were prepared in this manner:
</p><p>2-methoxy-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10b</bold>)
</p><p>2-chloro-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10d</bold>)
</p><p>2-fluoro-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10e</bold>)
</p><p>2,4-dichloro-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (<bold>10q</bold>)
</p><p><bold>General Procedure for the Conversion of the Substituted </bold><bold>11-Trimethylsilyl-6</bold><bold><italic toggle="yes">H</italic></bold><bold>-indolo[2,3-</bold><bold><italic toggle="yes">b</italic></bold><bold>]quinolines 8a</bold><bold>−</bold><bold>y
into the Substituted </bold><bold>11-Trimethylsilylneocryptolepines</bold><bold>9a</bold><bold>−</bold><bold>y.</bold> The appropriate 11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinoline (0.625 mmol) is dissolved in DMF (30 mL). To this
solution, CH<sub>3</sub>I (3 mL, 48 mmol) is added, and the solution is
refluxed overnight. The mixture is then cooled to room
temperature, after which it is poured into 50 mL of a 1 M Na<sub>2</sub>CO<sub>3</sub> solution followed by extraction with CH<sub>2</sub>Cl<sub>2</sub> (3 × 50 mL).
After removal of the solvent, the product is purified by column
chromatography using CH<sub>2</sub>Cl<sub>2</sub>/EtOAc (7:3) as the solvent. In
two cases, namely, the synthesis of 9-cyano-2-trifluoromethyl-11-trimethylsilylneocryptolepine (<bold>9n</bold>) and 9-cyano-2-fluoro-11-
trimethylsilylneocryptolepine (<bold>9o</bold>), formation of the compounds
was accompanied by the presence of a large amount of
desilylated product. Unfortunately, these products could not
be separated from each other. Therefore, the mixture of the
two was used in the desilylation reaction (see below). As a
consequence, no spectral data were recorded for these two
compounds. After the reaction in which substituted 11-trimethylsilyl-6<italic toggle="yes">H</italic>-indolo[2,3-<italic toggle="yes">b</italic>]quinolines were formed, the
1-substituted compound is separated from the 3-substituted
one and other impurities by using a silica column and CH<sub>2</sub>Cl<sub>2</sub>/EtOAc (7:3) as the eluent. The following products were
prepared in this manner:
</p><p>11-trimethylsilylneocryptolepine (<bold>9a</bold>)
</p><p>2-methoxy-11-trimethylsilylneocryptolepine (<bold>9b</bold>)
</p><p>2-bromo-11-trimethylsilylneocryptolepine (<bold>9c</bold>)
</p><p>2-chloro-11-trimethylsilylneocryptolepine (<bold>9d</bold>)
</p><p>2-fluoro-11-trimethylsilylneocryptolepine (<bold>9e</bold>)
</p><p>2-iodo-11-trimethylsilylneocryptolepine (<bold>9f</bold>)
</p><p>2-methyl-11-trimethylsilylneocryptolepine (<bold>9g</bold>)
</p><p>2-nitro-11-trimethylsilylneocryptolepine (<bold>9h</bold>)
</p><p>2-methylthio-11-trimethylsilylneocryptolepine (<bold>9i</bold>)
</p><p>2-cyano-11-trimethylsilylneocryptolepine (<bold>9j</bold>)
</p><p>9-cyano-11-trimethylsilylneocryptolepine (<bold>9k</bold>)
</p><p>2-chloro-9-cyano-11-trimethylsilylneocryptolepine (<bold>9l</bold>)
</p><p>9-cyano-2-methoxy-11-trimethylsilylneocryptolepine (<bold>9m</bold>)
</p><p>9-cyano-2-trifluoromethyl-11-trimethylsilylneocryptolepine (<bold>9n</bold>)
</p><p>9-cyano-2-fluoro-11-trimethylsilylneocryptolepine (<bold>9o</bold>)
</p><p>1-methoxy-11-trimethylsilylneocryptolepine (<bold>9r</bold>)
</p><p>1-bromo-11-trimethylsilylneocryptolepine (<bold>9s</bold>)
</p><p>1-trifluoromethyl-11-trimethylsilylneocryptolepine (<bold>9u</bold>)
</p><p>3-methoxy-11-trimethylsilylneocryptolepine (<bold>9v</bold>)
</p><p>3-bromo-11-trimethylsilylneocryptolepine (<bold>9w</bold>)
</p><p>3-chloro-11-trimethylsilylneocryptolepine (<bold>9x</bold>)
</p><p>3-trifluoromethyl-11-trimethylsilylneocryptolepine (<bold>9y</bold>)
</p><p><bold>General Procedure for the Conversion of the Substituted 11-Trimethylsilylneocryptolepines </bold><bold>9a</bold><bold>−</bold><bold>y</bold> <bold>into the
Neocryptolepine Derivatives</bold> <bold>1a</bold><bold>−</bold><bold>y. </bold>The appropriate 11-trimethylsilylneocryptolepine (0.35 mmol) is dissolved in 30
mL of ethanol, after which NaOH (10 mL, 6 N) is added. The
mixture is stirred for 3 h at 70 °C and cooled to room
temperature. Water (50 mL) is added, and the mixture is
extracted with CH<sub>2</sub>Cl<sub>2</sub> (3 × 50 mL). Purification of the product
is performed by column chromatography using acetone as the
eluent. 3-Methoxyneocryptolepine (<bold>1v</bold>) is purified by column
chromatography using toluene/diethylamine (9.5:0.5) as the
eluent. The following products were prepared in this manner:
</p><p>neocryptolepine (<bold>1a</bold>)
</p><p>2-methoxyneocryptolepine (<bold>1b</bold>)
</p><p>2-bromoneocryptolepine (<bold>1c</bold>)
</p><p>2-chloroneocryptolepine (<bold>1d</bold>)
</p><p>2-fluoroneocryptolepine (<bold>1e</bold>)
</p><p>2-iodoneocryptolepine (<bold>1f</bold>)
</p><p>2-methylneocryptolepine (<bold>1g</bold>)
</p><p>2-nitroneocryptolepine (<bold>1h</bold>)
</p><p>2-methylthioneocryptolepine (<bold>1i</bold>)
</p><p>2-cyanoneocryptolepine (<bold>1j</bold>)
</p><p>9-cyanoneocryptolepine (<bold>1k</bold>)
</p><p>2-chloro-9-cyanoneocryptolepine (<bold>1l</bold>)
</p><p>9-cyano-2-methoxyneocryptolepine (<bold>1m</bold>)
</p><p>9-cyano-2-trifluoromethylneocryptolepine (<bold>1n</bold>)
</p><p>9-cyano-2-fluoroneocryptolepine (<bold>1o</bold>)
</p><p>1-methoxyneocryptolepine (<bold>1r</bold>)
</p><p>1-bromoneocryptolepine (<bold>1s</bold>)
</p><p>1-trifluoromethylneocryptolepine (<bold>1u</bold>)
</p><p>3-methoxyneocryptolepine (<bold>1v</bold>)
</p><p>3-bromoneocryptolepine (<bold>1w</bold>)
</p><p>3-chloroneocryptolepine (<bold>1x</bold>)
</p><p>3-trifluoromethylneocryptolepine (<bold>1y</bold>)
</p><p><bold>Conversion to Hydrochloride Salts. </bold>All compounds were
biologically evaluated as their hydrochloride salts. Compounds
were dissolved in 0.1 N HCl in MeOH, and the solutions were
evaporated to dryness after 30 min.
</p><p><bold>In</bold> <bold>Vitro Activity against </bold><bold><italic toggle="yes">P. falciparum</italic></bold><bold>.</bold> For the determination of the antiplasmodial activity, the parasite lactate
dehydrogenase assay was used, with slight modifications, as
previously described by Makler et al.<xref rid="jm011102ib00034" ref-type="bibr"></xref> Briefly, the assay is
based on the observation that the lactate dehydrogenase (LDH)
enzyme of <italic toggle="yes">Plasmodium falciparum</italic> has the ability to rapidly
use 3-acetylpyridine NAD (APAD) as a coenzyme in the
reaction leading to the formation of pyruvate from lactate. Test
compounds were added in 2-fold serial dilutions to <italic toggle="yes">Plasmodium falciparum</italic> (chloroquine-sensitive Ghana strain or chloroquine-resistant W2 strain) cultures (1% parasitaemia, 2%
HCT) in 96-well plates. After 48 h at 37 °C and a gas mixture
of 93% N<sub>2</sub>, 4% CO<sub>2</sub>, and 3% O<sub>2</sub>, the parasite cultures were
frozen at −20 °C to lyse the erythrocytes and to store the plates
until further processing. After thawing, 20 μL of the lysed
culture was added to 100 μL of Malstat reagent (Flow Inc.),
and the formation of APADH was determined. Adding 40 μg
of nitroblue tetrazolium (NBT) and 2 μg of phenazine ethosulfate (PES) to the Malstat reagent promoted the spectrophotometric assessment of LDH activity. As APADH is formed,
the NBT is reduced and forms a blue formazan product that
can be detected visually and measured spectrophotometrically
at 650 nm. Determination of the IC<sub>50</sub> values was performed
in triplicate (mean ± SD). Cryptolepine<sup>9,11</sup> (Table <xref rid="jm011102it00003"></xref>) and
chloroquine were used as positive controls. IC<sub>50</sub> values of
chloroquine were 0.01 μM (Ghana) and 0.09 μM (W2).
</p><p><bold>In</bold> <bold>Vitro Acitivity against </bold><bold><italic toggle="yes">T. brucei</italic></bold><bold> Trypomastigotes.
</bold>IC<sub>50</sub> values against <italic toggle="yes">Trypanosoma brucei</italic> were determined in
triplicate (mean ± SD) as described before.<xref rid="jm011102ib00035" ref-type="bibr"></xref> Briefly, bloodstream forms of <italic toggle="yes">T. </italic><italic toggle="yes">brucei</italic> were cultivated in HMI-9 medium.
In a 96-well microplate, 10 000 hemoflagellates were incubated
at different concentrations of the test compound for 4 days.
Parasite multiplication was measured colorimetrically (490
nm) following addition of MTT, which converts to an water-soluble formazan product. Suramin was used as positive
control (IC<sub>50</sub> = 0.3 ± 0.1 μM).
</p><p><bold>In</bold> <bold>Vitro Acitivity against Intracellular </bold><bold><italic toggle="yes">T. cruzi</italic></bold><bold>
Amastigotes.</bold> IC<sub>50</sub> values against <italic toggle="yes">Trypanosoma </italic><italic toggle="yes">cruzi</italic> were
determined in triplicate (mean ± SD) as described before.<xref rid="jm011102ib00035" ref-type="bibr"></xref>
Briefly, primary mouse peritoneal macrophages were seeded
in 96-well microplates at 30 000 cells per well. After 24 h,
about 10 000 trypomastigotes of <italic toggle="yes">T. cruzi</italic> were added per well
together with 2-fold dilutions of the drug. The cultures were
incubated at 37 °C in5% CO<sub>2</sub>/95% air for 4 days. Following
fixation in MeOH and Giemsa staining, the drug activity was
semiquantitatively scored as a percent reduction of the total
parasite load (free trypomastigotes and intracellular amastigotes) compared with untreated control cultures. Scoring was
performed microscopically. Nifurtimox was used as a positive
control (IC<sub>50</sub> = 0.4 ± 0.1 μM).
</p><p><bold>Cytotoxicty on MRC-5 Cells.</bold> A human diploid embryonic
lung cell line (MRC-5) was used to assess the cytotoxicity of
the test compounds as described before.<xref rid="jm011102ib00035" ref-type="bibr"></xref> Briefly, MRC-5 cells
were seeded at 5000 cells per well in 96-well microtiter plates.
After 24 h, the cells were washed and 2-fold dilutions of the
drug in 200 μL of standard culture medium (RPMI + 5% FCS)
were added. The final DMSO concentration of the culture
remained below 0.5%. The cultures were incubated with
different concentrations of test compounds at 37 °C in 5% CO<sub>2</sub>/95% air for 7 days. Untreated cultures were included as
controls. Cytotoxicity was determined using the colorimetric
MTT assay and scored as a percent reduction of absorption at
540 nm of treated cultures versus untreated control cultures.
Determination of the IC<sub>50</sub> values was performed in triplicate
(mean ± SD). Vinblastine was used as a positive control (IC<sub>50</sub>
< 10 nM).
</p><p><bold>Inhibition of </bold><bold>β</bold><bold>-Hematin Formation. </bold>An in vitro method
was used to measure the inhibition of β-hematin formation,
based on the original method described by Egan et al.<xref rid="jm011102ib00024" ref-type="bibr"></xref> and
modified by Wright et al.<xref rid="jm011102ib00011" ref-type="bibr"></xref> Heme refers to Fe(II)protoporphyrin
IX, hemin is Fe(II)protoporphyrin IX chloride, and hematin is
Fe(II)protoporphyrin IX hydroxide. Briefly, 0.1 M NaOH was
added to Fe(II)hemin (Fluka; HPLC purity, >98%) (typically
7.5 mg), which forms hematin. Polymerization to β-hematin,
which has been characterized as [Fe(III)protoporphyrin IX]<sub>2</sub>,<xref rid="jm011102ib00036" ref-type="bibr"></xref>
proceeds after addition of a 9.9 M acetate buffer, pH 5. Test
compounds (3 equiv with respect to hemin) were added to the
hematin solutions prior to acidification. After incubation for
40 min at 60 °C, cooling and filtration, the precipitate was
washed with water and dried, and FT-IR (Perkin-Elmer FT-IR 1760) was used to check the presence of β-hematin, which
shows sharp bands at 1660 and 1207 cm<sup>-1</sup>. The spectra are
examined on the basis of the presence or absence of the two
typical peaks. When they are absent the compound has
inhibited the formation of β-hematin. A sample containing
hemin but without the test compound, which was not incubated, was used as a negative control (no formation of
β-hematin). A sample containing hemin without the test
compound, which was incubated for 40 min at 60 °C, was used
as a positive control (formation of β-hematin).
</p><p><bold>Characterization of Heme</bold><bold>:</bold><bold> Gas-Phase Ion Drug Complexes by Electrospray Ionization </bold><bold>(ESI) Mass Spectrometry. </bold>Abbreviations are the following: C, cationic form of the
drug; FP, heme = ferriprotoporphyrin (depending on the
conditions, FP may be present in an unliganded form [FP]<sup>+</sup>,
in a dimeric form [2FP − H]<sup>+</sup>, as a dehydrated dimer of
hematin [FP−O−FP]<sup>+</sup>, or as the CH<sub>3</sub>OH complex of the latter
dimeric form [FP−O−FP + MeOH]<sup>+</sup>). Mixtures of the drugs
(hydrochloride salts) with FP were prepared in MeOH/H<sub>2</sub>O (1:1; v/v; pH 7). Positive-ion mass spectra were recorded on an
Autospec-oa TOF mass spectrometer (Micromass, Manchester,
U.K.) employing electrospray ionization. Low-energy collision-induced dissociation (CID) spectra were acquired at a collision
energy (<italic toggle="yes">E</italic><sub>lab</sub>) of 400 eV using helium as the collision gas.
Helium was introduced into the collision cell until the signal
of the very weak FP<sup>+</sup>:CH<sub>3</sub>OH complex (<italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 648) reached 60%
of its original value. Under these conditions, the CID spectrum
of the FP<sup>+</sup>:MeOH complex revealed the FP<sup>+</sup> ion (<italic toggle="yes">m</italic>/<italic toggle="yes">z</italic> 616) as
the base peak, and the residual FP<sup>+</sup>:MeOH ion had a relative
abundance (RA) of 3%. To allow comparisons between binding
strengths of the different [FP:C]<sup>+</sup> complexes, the CID experiments were performed on the same day using the same
collision gas pressure.
</p><p><bold>DNA</bold><bold>−</bold><bold>Methyl Green Assay. </bold>The DNA−methyl green
assay is a simple microtiter assay for the detection of compounds that bind DNA. Agents that displace methyl green
from a DNA−methyl green complex (deoxyribonucleic acid
methyl green; Sigma) are detected spectrophotometrically
(Labsystems Multiscan MCC/340) by a decrease in absorbance
at 620 nm.<xref rid="jm011102ib00028" ref-type="bibr"></xref> DNA−methyl green was suspended in 100 mL of
0.05 M Tris-HCl buffer, pH 7.5, containing 7.5 mM MgSO<sub>4</sub> and
was stirred at 37 °C for 24 h. The dissolved samples were
dispensed into wells of a 96-well microtiter plate. Solvent was
removed under vacuum, and 200 μL of the DNA−methyl green
solution was added. The initial absorbance was compared with
the final absorbance (after 24 h) in order to calculate the IC<sub>50</sub>
value (50% displacement of methyl green from DNA). Determination of the IC<sub>50</sub> values was performed in triplicate (mean
± SD). The decrease in absorbance observed represents the
initial rapid displacement of methyl green from DNA by the
drug, followed by the slower reaction with water that yields
the colorless carbinol.
</p><p><bold>Electric Linear Dichroism (ELD) Measurements.</bold> ELD
measurements were performed with a computerized optical
measurement system using the procedures previously outlined.<xref rid="jm011102ib00037" ref-type="bibr"></xref> All experiments were conducted with a 10 mm path
length Kerr cell having a 1.5 mm electrode separation. The
samples were oriented under an electric field strength varying
from 1 to 13 kV/cm. The concentration of the test compound
was 10 μM in the presence of DNA at 200 μM unless stated
otherwise. This electrooptical method has proved to be most
useful for determining the orientation of the drugs bound to
DNA. It has the additional advantage that it senses only the
orientation of the polymer-bound ligand; free ligand is isotropic
and does not contribute to the signal.
</p><p><bold>Topoisomerase II Mediated DNA Cleavage Assay.</bold>
Supercoiled pKMp27 DNA (0.5 μg) was incubated with 4 units
of human topoisomerase II (TopoGen Inc.) at 37 °C for 30 min
in relaxation buffer (50 mM Tris, pH 7.8, 50 mM KCl, 10 mM
MgCl<sub>2</sub>, 1 mM dithiothreitol, 1 mM EDTA, and 0.5 mM ATP)
in the presence of varying concentrations of the test compound.
Reactions were terminated by adding SDS to 0.25% and
proteinase K to 250 μg/mL. DNA samples were then added to
the electrophoresis dye mixture (3 μL) and electrophoresed in
a 1% agarose gel containing ethidium bromide (1 μg/mL) at
room temperature for 2 h at 120 V. Gels were washed and
photographed under UV light.
</p></sec></body><back><ack><title>Acknowledgments</title><p>We thank J. Verreydt, J.
Schrooten, N. Verhaert, V. Van Heurck, W. Van Lierde,
and W. Van Dongen for their technical assistance. T.
Jonckers thanks the “Vlaams Instituut voor de Bevordering van het Wetenschappelijk Technologisch Onderzoek in de Industrie (IWT)” for a scholarship. He is also
indebted to the foundation “Rosa Blanckaert” for a
research grant. Financial support from the Fund for
Scientific Research (FWO-Vlaanderen) (Project Nos.
G.0119.96, G.0082.98, G.0334.00, and 1.5.139.00) and
from the Special Research Fund of the University of
Antwerp (Concerted Research Project No. 99/3/34, fellowship for S. van Miert) is gratefully acknowledged.
Dr. Colin Wright (University of Bradford, U.K.) is kindly
acknowledged for assistance with the assay on inhibition of β-hematin formation.</p></ack><app-group><app id="d7e4223"><title>Appendix</title><p><table-wrap id="jm011102it00004" position="float" orientation="portrait"><label>4</label><caption><p>Degree of Purity for All Compounds</p></caption><oasis:table colsep="2" rowsep="2"><oasis:tgroup cols="6"><oasis:colspec colnum="1" colname="1"></oasis:colspec><oasis:colspec colnum="2" colname="2"></oasis:colspec><oasis:colspec colnum="3" colname="3"></oasis:colspec><oasis:colspec colnum="4" colname="4"></oasis:colspec><oasis:colspec colnum="5" colname="5"></oasis:colspec><oasis:colspec colnum="6" colname="6"></oasis:colspec><oasis:tbody><oasis:row><oasis:entry namest="1" nameend="1">compd</oasis:entry><oasis:entry namest="2" nameend="2">% purity,
system A<italic toggle="yes"><sup>a</sup></italic><sup></sup></oasis:entry><oasis:entry namest="3" nameend="3">% purity,
system B<italic toggle="yes"><sup>b</sup></italic><sup></sup></oasis:entry><oasis:entry namest="4" nameend="4">compd</oasis:entry><oasis:entry namest="5" nameend="5">% purity,
system A<italic toggle="yes"><sup>a</sup></italic><sup></sup></oasis:entry><oasis:entry namest="6" nameend="6">% purity,
system B<italic toggle="yes"><sup>b</sup></italic><sup></sup></oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1a</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">100.0
</oasis:entry><oasis:entry colname="4"><bold>1k</bold></oasis:entry><oasis:entry colname="5">100.0
</oasis:entry><oasis:entry colname="6">95.7
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1b</bold></oasis:entry><oasis:entry colname="2">97.1
</oasis:entry><oasis:entry colname="3">99.2
</oasis:entry><oasis:entry colname="4"><bold>1l</bold></oasis:entry><oasis:entry colname="5">99.0
</oasis:entry><oasis:entry colname="6">96.4
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>10b</bold></oasis:entry><oasis:entry colname="2">98.3
</oasis:entry><oasis:entry colname="3">95.3
</oasis:entry><oasis:entry colname="4"><bold>1m</bold></oasis:entry><oasis:entry colname="5">100.0
</oasis:entry><oasis:entry colname="6">96.8
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1c</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">95.6
</oasis:entry><oasis:entry colname="4"><bold>1n</bold></oasis:entry><oasis:entry colname="5">96.2
</oasis:entry><oasis:entry colname="6">94.5
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1d</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">100.0
</oasis:entry><oasis:entry colname="4"><bold>1o</bold></oasis:entry><oasis:entry colname="5">97.6
</oasis:entry><oasis:entry colname="6">95.0
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>10d</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">95.7
</oasis:entry><oasis:entry colname="4"><bold>10q</bold></oasis:entry><oasis:entry colname="5">100.0
</oasis:entry><oasis:entry colname="6">93.2
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1e</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">100.0
</oasis:entry><oasis:entry colname="4"><bold>1r</bold></oasis:entry><oasis:entry colname="5">100.0
</oasis:entry><oasis:entry colname="6">97.1
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>10e</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">95.9
</oasis:entry><oasis:entry colname="4"><bold>1s</bold></oasis:entry><oasis:entry colname="5">100.0
</oasis:entry><oasis:entry colname="6">97.1
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1f</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">95.5
</oasis:entry><oasis:entry colname="4"><bold>1u</bold></oasis:entry><oasis:entry colname="5">98.4
</oasis:entry><oasis:entry colname="6">95.2
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1g</bold></oasis:entry><oasis:entry colname="2">98.0
</oasis:entry><oasis:entry colname="3">96.1
</oasis:entry><oasis:entry colname="4"><bold>1v</bold></oasis:entry><oasis:entry colname="5">92.8
</oasis:entry><oasis:entry colname="6">99.5
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1h</bold></oasis:entry><oasis:entry colname="2">95.4
</oasis:entry><oasis:entry colname="3">95.1
</oasis:entry><oasis:entry colname="4"><bold>1w</bold></oasis:entry><oasis:entry colname="5">99.3
</oasis:entry><oasis:entry colname="6">96.8
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1i</bold></oasis:entry><oasis:entry colname="2">99.4
</oasis:entry><oasis:entry colname="3">95.5
</oasis:entry><oasis:entry colname="4"><bold>1x</bold></oasis:entry><oasis:entry colname="5">100.0
</oasis:entry><oasis:entry colname="6">95.8
</oasis:entry></oasis:row><oasis:row><oasis:entry colname="1"><bold>1j</bold></oasis:entry><oasis:entry colname="2">100.0
</oasis:entry><oasis:entry colname="3">95.0
</oasis:entry><oasis:entry colname="4"><bold>1y</bold></oasis:entry><oasis:entry colname="5">99.5
</oasis:entry><oasis:entry colname="6">95.2</oasis:entry></oasis:row></oasis:tbody></oasis:tgroup></oasis:table><table-wrap-foot><p><italic toggle="yes"><sup>a</sup></italic><sup></sup> System A: column, MERCK, Lichrospher Si 600 (4 mm × 250
mm); solvent system, 1-propanol−hexane 5%/water 95:5 or 1-propanol−hexane 5%/water 90:10.<italic toggle="yes"><sup>b</sup></italic><sup></sup> System B: column, MERCK,
Lichrocart-Lichrospher 100 RP 18 (4 mm × 250 mm); solvent
system, gradient from 80% to 100% acetonitrile in 20 min.</p></table-wrap-foot></table-wrap></p></app></app-group><notes notes-type="si"><sec id="d7e4509"><title><ext-link xlink:href="/doi/suppl/10.1021%2Fjm011102i">Supporting Information Available</ext-link></title><p>Characterization of
intermediate compounds and target compounds. This material is available free of charge via the Internet at <uri xlink:href="http://pubs.acs.org">http://pubs.acs.org</uri>.
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M.</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Chemistry, University of Antwerp.</affiliation><affiliation> These authors contributed equally to this work.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">VAN MIERT</namePart><namePart type="given">Sabine</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> These authors contributed equally to this work.</affiliation><affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">CIMANGA</namePart><namePart type="given">Kanyanga</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">BAILLY</namePart><namePart type="given">Christian</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> INSERM U-524, Lille.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">COLSON</namePart><namePart type="given">Pierre</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Institute of Chemistry, University of Liège.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">DE PAUW-GILLET</namePart><namePart type="given">Marie-Claire</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Institute of Anatomy, University of Liège.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">VAN DEN HEUVEL</namePart><namePart type="given">Hilde</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">CLAEYS</namePart><namePart type="given">Magda</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">LEMIèRE</namePart><namePart type="given">Filip</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Chemistry, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">ESMANS</namePart><namePart type="given">Eddy L.</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Chemistry, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">ROZENSKI</namePart><namePart type="given">Jef</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Rega Institute for Medical Research.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">QUIRIJNEN</namePart><namePart type="given">Ludo</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Tibotec-Virco NV.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">MAES</namePart><namePart type="given">Louis</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Tibotec-Virco NV.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">DOMMISSE</namePart><namePart type="given">Roger</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Chemistry, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">LEMIèRE</namePart><namePart type="given">Guy L. F.</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Chemistry, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="family">VLIETINCK</namePart><namePart type="given">Arnold</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal" displayLabel="corresp"><namePart type="family">PIETERS</namePart><namePart type="given">Luc</namePart><affiliation>Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, Department ofChemistry, University of Antwerp, B-2020 Antwerp, Belgium, INSERM U-524, Institut de Recherches sur le Cancer de Lille,F-59045 Lille, France, Institute of Chemistry, University of Liège, Sart-Tilman (B6), B-4000 Liège, Belgium, CRCE,Histology−Cytology, Institute of Anatomy (L3), University of Liège, B-4020 Liège, Belgium, Rega Institute for MedicalResearch, B-3000 Leuven, Belgium, and Tibotec-Virco NV, B-2800 Mechelen, Belgium</affiliation><affiliation> Department of Pharmaceutical Sciences, University of Antwerp.</affiliation><affiliation> To whom correspondence should be addressed. Phone and fax: (32)3 820 27 09. E-mail pieters@uia.ua.ac.be.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><typeOfResource>text</typeOfResource><genre type="research-article" displayLabel="research-article" authority="ISTEX" authorityURI="https://content-type.data.istex.fr" valueURI="https://content-type.data.istex.fr/ark:/67375/XTP-1JC4F85T-7">research-article</genre><originInfo><publisher>American Chemical Society</publisher><dateCreated encoding="w3cdtf">2002-06-29</dateCreated><dateIssued encoding="w3cdtf">2002-08-01</dateIssued><copyrightDate encoding="w3cdtf">2002</copyrightDate></originInfo><abstract>On the basis of the original lead neocryptolepine or 5-methyl-5H-indolo[2,3-b]quinoline, an alkaloid from Cryptolepis sanguinolenta, derivatives were prepared using a biradical cyclization methodology. Starting from easily accessible educts, this approach allowed the synthesis of hitherto unknown compounds with a varied substitution pattern. As a result of steric hindrance, preferential formation of the 3-substituted isomers over the 1-substituted isomers was observed when cyclizing N-(3-substituted-phenyl)-N‘-[2-(2-trimethylsilylethynyl)phenyl]carbodiimides. All compounds were evaluated for their activity against chloroquine-sensitive as well as chloroquine-resistant Plasmodium falciparum strains, for their activity against Trypanosoma brucei and T. cruzi, and for their cytotoxicity on human MRC-5 cells. Mechanisms of action were investigated by testing heme complexation using ESI-MS, inhibition of β-hematin formation, DNA interactions (DNA−methyl green assay and linear dichroism), and inhibition of human topoisomerase II. Neocryptolepine derivatives with a higher antiplasmodial activity and a lower cytotoxicity than the original lead have been obtained. This selective antiplasmodial activity was associated with inhibition of β-hematin formation. 2-Bromoneocryptolepine was the most selective compound with an IC50 value against chloroquine-resistant P. falciparum of 4.0 μM in the absence of cytotoxicity (IC50 > 32 μM). Although cryptolepine, a known lead for antimalarials also originally isolated from Cryptolepis sanguinolenta, was more active (IC50 = 2.0 μM), 2-bromoneocryptolepine showed a low affinity for DNA and no inhibition of human topoisomerase II, in contrast to cryptolepine. Although some neocryptolepine derivatives showed a higher antiplasmodial activity than 2-bromocryptolepine, these compounds also showed a higher affinity for DNA and/or a more pronounced cytotoxicity. Therefore, 2-bromoneocryptolepine is considered as the most promising lead from the present work for new antimalarial agents. In addition, 2-bromo-, 2-nitro-, and 2-methoxy-9-cyanoneocryptolepine exhibited antitrypanosomal activity in the micromolar range in the absence of obvious cytotoxicity.</abstract><relatedItem type="host"><titleInfo><title>Journal of Medicinal Chemistry</title></titleInfo><titleInfo type="abbreviated"><title>J. Med. Chem.</title></titleInfo><genre type="journal" authority="ISTEX" authorityURI="https://publication-type.data.istex.fr" valueURI="https://publication-type.data.istex.fr/ark:/67375/JMC-0GLKJH51-B">journal</genre><identifier type="ISSN">0022-2623</identifier><identifier type="eISSN">1520-4804</identifier><identifier type="acspubs">jm</identifier><identifier type="coden">JMCMAR</identifier><identifier type="uri">pubs.acs.org/jmc</identifier><part><date>2002</date><detail type="volume"><caption>vol.</caption><number>45</number></detail><detail type="issue"><caption>no.</caption><number>16</number></detail><extent unit="pages"><start>3497</start><end>3508</end></extent></part></relatedItem><identifier type="istex">7040C93C2FEF52E1D59CF67C9FAF41EB0DF5CA74</identifier><identifier type="ark">ark:/67375/TPS-05PG9VCQ-8</identifier><identifier type="DOI">10.1021/jm011102i</identifier><accessCondition type="use and reproduction" contentType="restricted">Copyright © 2002 American Chemical Society</accessCondition><recordInfo><recordContentSource authority="ISTEX" authorityURI="https://loaded-corpus.data.istex.fr" valueURI="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-X5HBJWF8-J">ACS</recordContentSource><recordOrigin>Copyright © 2002 American Chemical Society</recordOrigin></recordInfo></mods><json:item><extension>json</extension><original>false</original><mimetype>application/json</mimetype><uri>https://api.istex.fr/ark:/67375/TPS-05PG9VCQ-8/record.json</uri></json:item></metadata><annexes><json:item><extension>tiff</extension><original>true</original><mimetype>image/tiff</mimetype><uri>https://api.istex.fr/document/7040C93C2FEF52E1D59CF67C9FAF41EB0DF5CA74/annexes/tiff</uri></json:item></annexes><serie></serie></istex></record>Pour manipuler ce document sous Unix (Dilib)
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