Síntese, atividade anti-Trypanosoma cruzi e antitumoral de aciltiossemicarbazidas e 1,3,4-tiadiazóis do S-(-) e R-(+)-limoneno

Abstract

RESUMO: A sintese de substancias quimicas a partir de substratos opticamente ativos isolados de fontes naturais e uma forma eficiente de obtencao de compostos quirais mais complexos e biologicamente ativos. Este trabalho implementou novas metodologias de sintese de 1,3,4-tiadiazois e aciltiossemicarbazidas a partir do limoneno. O limoneno e um monoterpeno abundante encontrado nas formas enantiomericas R e S e apresenta diversas atividades biologicas, incluindo antiprotozoaria e antitumoral. Tambem apresentam estas atividades os 1,3,4-tiadiazois, classe de compostos heteroaromaticos, e as aciltiosemicarbazidas, importante classe de intermediarios de sintese. Derivados terpenicos nitrogenados, como isotiocianoterpenos e tiossemicarbazidas contendo a unidade terpenica em posicao pouco usual, servem como precursores sinteticos para ambas as classes bioativas supracitadas e sua sintese ja e consolidada no grupo de pesquisa ao qual pertence este estudo. A partir das tiossemicarbazidas do limoneno e aldeidos (benzaldeido, derivados de benzaldeido, cinamaldeido e aldeidos heteroaromaticos), 31 derivados 2-amino-1,3,4-tiadiazol do limoneno 5-substituidos - 21 do S-(-)-limoneno e 10 do R-(+)-limoneno - foram sintetizados em um procedimento one-pot envolvendo ciclizacao oxidativa por Fe(III), com rendimentos de 63-91%. Quatro 1-aciltiossemicarbazidas 4-substituidas de cada enantiomero do limoneno foram obtidas pela adicao de hidrazidas aos isotiocianoterpenos, com rendimentos de 52-68%. Os precursores hidrazida tambem foram sintetizados por novo procedimento, com rendimentos de 82-98%. As aciltiossemicarbazidas e os 1,3,4-tiadiazois foram avaliados in vitro como agentes antitumorais, sobre varias linhagens de celulas, e agentes anti-Trypanosoma cruzi. Nos ensaios antiproliferativos para células tumorais, os derivados do S-(-)-limoneno apresentaram os resultados de atividade mais expressivos dentre a serie 1,3,4-tiadiazois, enquanto que não houve correlacao especifica das atividades das aciltiossemicarbazidas com a quiralidade da porcao terpenica. As aciltiossemicarbazidas apresentaram atividade inibitoria mais pronunciada para celulas de ovario resistente a multiplos farmacos (NCI-ADR/RES), com o menor valor de concentração inibitoria (GI50) de 2,4 mol.L-1 para a 1-benzoil-tiossemicarbazida do S-(-)-limoneno. Os menores valores de GI50 sobre uma linhagem celular especifica ocorreram para o 5-nitrotiofenil-1,3,4-tiadiazol do S-(-)-limoneno (0,9 mol.L-1, celulas de mama MCF-7) e para o 5-nitrofuranil-1,3,4-tiadiazol do S-(-)-limoneno (0,8 mol.L-1, celulas de colon HT29), que apresentou efeito citostatico (TGI) e citocida (LC50) mais pronunciado que o controle positivo doxorrubicina em algumas linhagens. Na avaliacao de atividade anti-Trypanosoma cruzi, a serie do S-(-)-limoneno de 1,3,4-tiadiazois novamente apresentou os resultados mais significativos em formas tripomastigotas do protozoario. Tres derivados (5-nitrofuranil-1,3,4-tiadiazois do R-(+) e do S-(-)-limoneno e 5-nitrotiofenil-1,3,4-tiadiazol do S-(-)-limoneno) foram mais ativos que o controle positivo benzinidazol, com o menor valor de concentração inibitoria de 5,5 mol.L-1. A 1-(2-fenilacetil)-tiossemicarbazida do R-(+)-limoneno exibiu atividade expressiva para as formas tripomastigotas e epimastigotas do protozoario e um dos menores valores de toxicidade em celulas LLC-MK2. Os resultados de atividades biologicas significativos para os derivados quirais 1,3,4-tiadiazos e aciltiossemicarbazidas, sintetizados a partir do limoneno, com materiais de partida simples e procedimentos comuns, corroboraram o proposito deste trabalho

ABSTRACT: The synthesis of chemical substances employing chiral substrates isolated from natural sources is an efficient way of obtaining more complex and biologically active compounds. This work developed new methodologies to synthesize 1,3,4-thiadiazoles and acylthiosemicarbazides from limonene. Limonene is an abundant monoterpene existing in R and S enantiomeric forms and possesses a wide range of biological activities, such as antiprotozoal and antitumor. 1,3,4-thiadiazoles, a class of heteroaromatic compounds, and acylthiosemicarbazides, an important class of synthetic intermediates, also show those biological activities. Terpene-based nitrogen-containing derivatives, like isothiocyanoterpenes and thiosemicarbazides bearing a terpene moiety at an unusual position are useful synthetic precursors to both bioactive classes of chemicals previously described, as well as their synthesis protocol is already established in the research group to which this study belongs. Starting from limonene thiosemicarbazides and aldehydes (benzaldehyde, benzaldehyde derivatives, cinnamaldehyde and heteroaromatic aldehydes), thirty-one 5-substituted 2-amino-1,3,4-thiadiazoles from limonene - 21 from S-(-)-limonene e 10 from R-(+)-limonene - were synthesized by oxidative cyclization with Fe(III) via one-pot procedure in 63-91% yields. Addition of hydrazides to isothyocianoterpenes yielded four 4-substituted 1-acylthiosemicarbazides from each enantiomer of limonene in 52-68%. Hydrazides precursors were also synthesized by new procedure in 82-98% yields. All acylthiosemicarbazides and 1,3,4-thiadiazoles were evaluated in vitro as antitumor agents for several cell lines and as anti-Trypanosoma cruzi agents. The antiproliferative essays for tumor cells afforded the most expressive results for the S-(-)-limonene derivatives among the 1,3,4-thiadiazoles series, whereas no specific correlation was observed between the acylthiosemicarbazides activities and the chirality of the terpene moiety. Acylthiosemicarbazides presented remarkable activity for breast resistant NCI/ADR cancer cell line furnishing the lowest value of cellular growth inhibition (GI50) as 2,4 mol.L-1 for 1-benzoyl-thiosemicarbazide from S-(-)-limonene. The lowest values of GI50 for a single cell line were found for 5-nitrothiophenyl-1,3,4-tiadiazole from S-(-)-limonene (0,9 mol.L-1, breast MCF-7 cells) and for 5-nitrofuranyl-1,3,4-thiadiazole from S-(-)-limonene (0,8 mol.L-1, colon HT29 cells). The latter also presented better cytostatic (TGI) and cytocidal (LC50) effects than the positive control doxorubicin for some of the cell lines. The anti-Trypanosoma cruzi evaluation also afforded the most meaningful results for the S-(-)-limonene series among the 1,3,4-thiadiazoles, against the protozoan trypomastigote forms. Three derivatives (5-nitrofuranyl-1,3,4-thiadiazole from R-(+) and S-(-)-limonene and 5-nitrothiophenyl-1,3,4-tiadiazole from S-(-)-limonene) were more active than the positive control benznidazole, being found 5,5 mol.L-1 as the lowest value of inhibitory concentration. 1-(2-phenylacetyl)- thiosemicarbazide from R-(+)-limonene furnished pronounced inhibitory activities for both trypomastigote and epimastigote T. cruzi forms, besides one of the smallest cytotoxicity values upon LLC-MK2 cells. The achievement of remarkable results of biological activities for chiral 1,3,4-thiadiazoles and acylthiosemicarbazides synthesized from limonene, employing simple starting materials and standard procedures, supported the aim of this work