[An Innovative Combination in the Combat Against Trypanosoma cruzi: The Synergistic Effect of Benznidazole and...

Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, is one of the most significant neglected tropical diseases. Approximately 6-7 million people worldwide are estimated to be infected with T.cruzi, with about 12000 deaths annually attributed to the disease. In Türkiye, natural transmission does not occur due to the absence of vectors carrying the parasite. However, touristic or commercial travels to endemic areas and migration from these regions increase the risk of Chagas disease emergence in the country. Currently, only two drugs, benznidazole (BENZ) and nifurtimox, are available for the treatment of Chagas disease. These drugs have limitations, including effectiveness only during the acute or early stages of infection, adverse side effects and the potential for the parasite to develop resistance. Consequently, there is an urgent need for new, safe and effective therapeutic alternatives for treating Chagas disease. This study aimed to investigate the efficacy and cytotoxicity of essential oil components (EOCs) such as α-pinene, isoborneol, carvacrol, coumarin and eugenol against T.cruzi and their synergy with BENZ. In this study, the T.cruzi ATCC 50825 reference strain was used. BENZ was selected as the reference drug and α-pinene, isoborneol, carvacrol, coumarin, and eugenol were chosen as EOCs. Cytotoxic activity was evaluated using the L929 mouse fibroblast cell line. Antitrypanosomal and cytotoxic activities were determined using the broth microdilution method and interactions between BENZ and EOCs were assessed using the checkerboard method. The IC50 values indicating the cytotoxic effects of EOCs and BENZ on fibroblast cells at 24, 48, and 72 hours were as follows: α-pinene (26.68-20.79 µg/mL), isoborneol (128.7-73.63 µg/mL), carvacrol (4.56-2.49 µg/mL), coumarin (169.3-108.6 µg/mL), eugenol (6.04-1.65 µg/mL), and BENZ (1011-806.9 µg/mL). The IC50 values for antitrypanosomal activity at 48 hours were 11.8 µg/mL, 114.6 µg/mL, 3.9 µg/mL, 268.6 µg/mL, 19.5 µg/mL, and 11.7 µg/mL for the same compounds, respectively. Carvacrol, α-pinene and eugenol demonstrated the most potent activity and especially carvacrol and α-pinene being comparable to the reference drug BENZ. Selectivity index (SI) calculations revealed high selective activity (SI> 1) for BENZ and α-pinene, while other EOCs exhibited lower selectivity (SI< 1). Nevertheless, synergistic interactions were identified in all combinations of EOCs with BENZ. The mechanisms underlying drug resistance in T.cruzi remain poorly understood. Natural compounds with multiple biological effects, new drug formulations and innovative combinations can contribute to overcoming drug resistance. Among these strategies, essential oil components, known for their biological activities against various microorganisms, offer promising alternatives. In addition to the strong antitrypanosomal activities of EOCs, their synergistic interaction with BENZ may fill important gaps in the literature and guide future studies in designing next-generation drug combinations, reducing side effects and preventing the development of resistance.