RESUMEN
INTRODUCTION: The first total synthesis of ω-phenyl Δ6 fatty acids (FA) and their cytotoxicity (A549) and leishmanicidal (L. infantum) activities are described. The novel 16-phenyl-6-hexadecynoic acid (1) and the known 16-phenylhexadecanoic acid (2) were synthesized in 7-8 steps with overall yields of 46 % and 41 %, respectively. The syntheses of the unprecedented 10-phenyl-6-decynoic acid (3), 10-cyclohexyl-6-decynoic acid (4) and 10-(4-methoxyphenyl)-6-decynoic acid (5) was also performed in 3 steps with 73-76 % overall yields. The use of lithium acetylide coupling enabled the 4-step synthesis of 10-phenyl-6Z-decenoic acid (6) with a 100 % cis-stereochemistry. The cytotoxicity of these novel FA was determined against A549 cells and L. infantum promastigotes and amastigotes. Among the ω-phenylated FA, the best cytotoxicity towards A549 was displayed by 1, with an IC50 of 18 ± 1 µM. On the other hand, among the C10 acids, the ω-cyclohexyl acid 4 presented the best cytotoxicity (IC50 = 40 ± 2 µM) towards A549. RESULTS: Based on caspase-3/7 studies neither of the FA induced apoptosis in A549, thus implying other mechanisms of cell death. CONCLUSION: The antileishmanial studies were performed with the top Leishmania donovani topoisomerase IB (LdTopIB) inhibitors, namely 1 and 2 (EC50 between 14 and 36 µM, respectively), acids that did not stabilize the cleavage complexes between LdTopIB and DNA. Acids 1 and 2 displayed cytotoxicity towards L. infantum amastigotes (IC50 = 3-6 µM) and L. infantum promastigotes (IC50 = 60- 70 µM), but low toxicity towards murine splenocytes. Our results identified 1 as the optimum ω- phenylated acid of the series.
Asunto(s)
Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Antiprotozoarios/síntesis química , Antiprotozoarios/farmacología , Ácidos Grasos Omega-6/síntesis química , Ácidos Grasos Omega-6/farmacología , Leishmania infantum/efectos de los fármacos , Inhibidores de Topoisomerasa I/farmacología , Antineoplásicos/química , Antiprotozoarios/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , ADN-Topoisomerasas de Tipo I/metabolismo , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Ácidos Grasos Omega-6/química , Humanos , Estructura Molecular , Pruebas de Sensibilidad Parasitaria , Proteínas Recombinantes/metabolismo , Relación Estructura-Actividad , Inhibidores de Topoisomerasa I/síntesis química , Inhibidores de Topoisomerasa I/químicaRESUMEN
Sponges biosynthesize α-methoxylated fatty acids with unusual biophysical and biological properties and in some cases they display enhanced anticancer activities. However, the antiprotozoal properties of the α-methoxylated fatty acids have been less studied. In this work, we describe the total synthesis of (5Z,9Z)-(±)-2-methoxy-5, 9-eicosadienoic acid (1) and its acetylenic analog (±)-2-methoxy-5,9-eicosadiynoic acid (2), and report that they inhibit (EC50 values between 31 and 22 µM) the Leishmania donovani DNA topoisomerase IB enzyme (LdTopIB). The inhibition of LdTopIB (EC50 = 53 µM) by the acid (±)-2-methoxy-6-icosynoic acid (12) was studied as well. The potency of LdTopIB inhibition followed the trend 2 > 1 > 12, indicating that the effectiveness of inhibition depends on the degree of unsaturation. All of the studied α-methoxylated fatty acids failed to inhibit the human topoisomerase IB enzyme (hTopIB) at 100 µM. However, the α-methoxylated fatty acids were capable of inhibiting an active but truncated LdTopIB with which camptothecin (CPT) cannot interact suggesting that the methoxylated fatty acids inhibit LdTopIB with a mechanism different from that of CPT. The diunsaturated fatty acids displayed low cytotoxicity towards Leishmania infantum promastigotes (EC50 values between 260 and 240 µM), but 12 displayed a better cytotoxicity towards Leishmania donovani promastigotes (EC50 = 100 µM) and a better therapeutic index.
Asunto(s)
Camptotecina/farmacología , Ácidos Grasos Insaturados/química , Ácidos Grasos Insaturados/farmacología , Leishmania donovani/efectos de los fármacos , Inhibidores de Topoisomerasa I/química , Inhibidores de Topoisomerasa I/farmacología , Antiprotozoarios/química , Antiprotozoarios/farmacología , ADN-Topoisomerasas de Tipo I/metabolismo , HumanosRESUMEN
The fatty acids (±)-2-methoxy-6Z-heptadecenoic acid (1), (±)-2-methoxy-6-heptadecynoic acid (2) and (±)-2-methoxyheptadecanoic acid (3) were synthesized and their inhibitory activity against the Leishmania DNA topoisomerase IB enzyme (LdTopIB) determined. Acids 1 and 2 were synthesized from 4-bromo-1-pentanol, the former in ten steps and in 7% overall yield, while the latter in seven steps and in 14% overall yield. Acid 3 was prepared in six steps and in 42% yield from 1-hexadecanol. Acids 1-3 inhibited the LdTopIB enzyme following the order 2 > 1 ⪢ 3, with 2 displaying an EC(50) = 16.6 ± 1.1 µM and 3 not inhibiting the enzyme. Acid 1 preferentially inhibited the LdTopIB enzyme over the human TopIB enzyme. Unsaturation seems to be a prerequisite for effective inhibition, rationalized in terms of weak intermolecular interactions between the active site of LdTopIB and either the double or triple bonds of the fatty acids. Toxicity towards Leishmania donovani promastigotes was also investigated resulting in the same order 2 > 1 > 3, with 2 displaying an EC(50) = 74.0 ± 17.1 µM. Our results indicate that α-methoxylation decreases the toxicity of C(17:1) fatty acids towards L. donovani promastigotes, but improves their selectivity index.
RESUMEN
2-Alkynoic fatty acids display antimycobacterial, antifungal, and pesticidal activities but their antiprotozoal activity has received little attention. In this work we synthesized the 2-octadecynoic acid (2-ODA), 2-hexadecynoic acid (2-HDA), and 2-tetradecynoic acid (2-TDA) and show that 2-ODA is the best inhibitor of the Leishmania donovani DNA topoisomerase IB enzyme (LdTopIB) with an EC(50)=5.3±0.7µM. The potency of LdTopIB inhibition follows the trend 2-ODA>2-HDA>2-TDA, indicating that the effectiveness of inhibition depends on the fatty acid carbon chain length. All of the studied 2-alkynoic fatty acids were less potent inhibitors of the human topoisomerase IB enzyme (hTopIB) as compared to LdTopIB. 2-ODA also displayed in vitro activity against Leishmania donovani (IC(50)=11.0µM), but it was less effective against other protozoa, Trypanosoma cruzi (IC(50)=48.1µM) and Trypanosoma brucei rhodesiense (IC(50)=64.5µM). The antiprotozoal activity of the 2-alkynoic fatty acids, in general, followed the trend 2-ODA>2-HDA>2-TDA. The experimental information gathered so far indicates that 2-ODA is a promising antileishmanial compound.
Asunto(s)
Alquinos/farmacología , Antiprotozoarios/farmacología , ADN-Topoisomerasas de Tipo I/metabolismo , Ácidos Grasos Insaturados/farmacología , Leishmania donovani/enzimología , Inhibidores de Topoisomerasa I/farmacología , Alquinos/síntesis química , Alquinos/química , Animales , Antiprotozoarios/síntesis química , Antiprotozoarios/química , Línea Celular , Relación Dosis-Respuesta a Droga , Ácidos Grasos Insaturados/síntesis química , Ácidos Grasos Insaturados/química , Humanos , Leishmania donovani/efectos de los fármacos , Macrófagos/efectos de los fármacos , Ratones , Ratones Endogámicos BALB C , Estructura Molecular , Pruebas de Sensibilidad Parasitaria , Ratas , Relación Estructura-Actividad , Inhibidores de Topoisomerasa I/síntesis química , Inhibidores de Topoisomerasa I/química , Trypanosoma brucei rhodesiense/efectos de los fármacos , Trypanosoma cruzi/efectos de los fármacosRESUMEN
La utilización intensiva de fármacos antiparasitarios es la causa principal de la aparición de microorganismos parásitos multirresistentes en las regiones del planeta donde son precisamente endémicos. Los agentes etiológicos de las denominadas enfermedades tropicales -malaria, criptosporiodiosis, enfermedad del sueño, enfermedad de Chagas o los distintos tipos de leishmaniosis- son protozoos unicelulares sobre los que no se ha desarrollado en la actualidad ninguna vacuna eficaz y cuyo tratamiento se basa en medidas sanitarias preventivas y en el uso de medicamentos. La quimioterapia antiparasitaria actual es cara, no está ausente de efectos adversos y no supone beneficios a las empresas que la comercializan, por lo que la inversión en I & D es marginal comparada con la llevada a cabo para otros procesos patológicos de menor relevancia médica. La identificación de las ADN topoisomerasas como dianas farmacológicas se basa en los excelentes resultados obtenidos en los ensayos clínicos llevados a cabo con los derivados de la camptotecina en la terapia antitumoral. Las importantes diferencias estructurales entre las ADN topoisomerasas de tipo I de tripanosomas y leishmanias con respecto a sus homólogas de mamífero ha abierto un nuevo campo de investigación que combina las técnicas de biología molecular con la cristalización de proteínas para poder diseñar nuevos fármacos dirigidos específicamente a su inhibición. Revisamos aquí las características de estas nuevas dianas farmacológicas, así como los compuestos que en el momento están siendo utilizados para su inhibición en los agentes parasitarios que causan las principales enfermedades tropicales.
The intensive use of antiparasitic drugs is the main cause of the emergence of multiresistant parasite strains on those regions where these parasites are endemic. The aetiological agents of the so-called tropical diseases viz. malaria, cryptosporidiosis, sleeping sickness, Chagas disease or leishmaniasis, among others, are unicellular protozoan parasites with no immune-prophylactic treatment and where the chemotherapeutical treatment is still under controversy. At present, the chemotherapeutic approach to these diseases is expensive, has side or toxic effects and it does not provide economic profits to the Pharmaceuticals which then have no or scarce enthusiasm in R & D investments in this field. The identification of type I DNAtopoisomerases as promising drug targets is based on the excellent results obtained with camptothecin derivatives in anticancer therapy. The recent finding of significant structural differences between human type I DNAtopoisomerase and their counterparts in trypanosomatids has open a new field in drug discovery, the aim is to find structural insights to be targeted by new drugs. This review is an update of DNA-topoisomerases as potential chemotherapeutic targets against the most important protozoan agents of medical interest.
Asunto(s)
Animales , Humanos , Antineoplásicos/farmacología , Eucariontes/enzimología , Inhibidores de Topoisomerasa I , Antineoplásicos/química , Reparación del ADN , ADN-Topoisomerasas de Tipo I/genética , ADN-Topoisomerasas de Tipo I/metabolismo , Diseño de Fármacos , Eucariontes/genética , Leishmania/enzimología , Leishmania/genética , Neoplasias/tratamiento farmacológico , Infecciones por Protozoos/parasitología , Relación Estructura-Actividad , Trypanosoma/enzimología , Trypanosoma/genéticaRESUMEN
The intensive use of antiparasitic drugs is the main cause of the emergence of multiresistant parasite strains on those regions where these parasites are endemic. The aetiological agents of the so-called tropical diseases viz. malaria, cryptosporidiosis, sleeping sickness, Chagas disease or leishmaniasis, among others, are unicellular protozoan parasites with no immune-prophylactic treatment and where the chemotherapeutical treatment is still under controversy. At present, the chemotherapeutic approach to these diseases is expensive, has side or toxic effects and it does not provide economic profits to the Pharmaceuticals which then have no or scarce enthusiasm in R & D investments in this field. The identification of type I DNA-topoisomerases as promising drug targets is based on the excellent results obtained with camptothecin derivatives in anticancer therapy. The recent finding of significant structural differences between human type I DNA-topoisomerase and their counterparts in trypanosomatids has open a new field in drug discovery, the aim is to find structural insights to be targeted by new drugs. This review is an update of DNA-topoisomerases as potential chemotherapeutic targets against the most important protozoan agents of medical interest.