Mem Inst Oswaldo Cruz, Rio de Janeiro, Vol. 121 (Suppl. 2) 2026
Review
Advances and challenges in the search for new treatments for Chagas disease
1Fundação Oswaldo Cruz-Fiocruz, Instituto Carlos Chagas, Curitiba, PR, Brasil
2Fundação Oswaldo Cruz-Fiocruz, Instituto René Rachou, Belo Horizonte, MG, Brasil
DOI: 10.1590/0074-02760250299
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ABSTRACT
The treatment of Chagas disease (CD) has relied for more than five decades on two drugs, benznidazole (BZ) and nifurtimox (NTX), both with significant limitations and severe adverse effects. Their limited efficacy during the chronic phase underscores the urgent need for new chemotherapeutic strategies. The pursuit of new therapeutic agents for CD is focused on identifying molecular targets essential for Trypanosoma cruzi survival that are either absent or highly divergent in human, thereby enhancing selectivity and minimising off-target toxicity. These targets exploit the parasite’s unique biology at multiple levels. Key metabolic vulnerabilities include: ergosterol biosynthesis, a sterol pathway distinct from human cholesterol metabolism; glycosomal metabolism, reflecting parasite´s unique compartmentalisation of glycolysis; and redox homeostasis, which depends on trypanothione rather than glutathione. Additional promising avenues involve the parasite’s genetic and epigenetic regulation, mRNA processing and translational control. Furthermore, virulence-associated factors, and specific enzymes such as type I nitroreductase (NTR-1) can be exploited for selective prodrug activation. The complex genomic organisation and pronounced plasticity of T. cruzi complicate the identification of novel therapeutic targets. The abundance of proteins annotated as hypothetical or of unknown function further obscure critical metabolic pathways that could serve as druggable targets. In this context, the discovery of new drugs for CD strategically integrates phenotypic, target-based, and computational approaches, all of which require rigorous validation through preclinical in vitro and in vivo studies. Although modern approaches have yielded several promising lead compounds, successfully controlling CD will also depend on overcoming socioeconomic and access-related barriers to ensure that new therapies reach the populations most affected by this neglected tropical disease.
REFERENCES
01. WHO - World Health Organization. Chagas disease (also known as American trypanosomiasis). 2025. Available from: https:// www.who.int/news-room/fact-sheets/detail/chagas-disease- (american-trypanosomiasis) (Accessed 28th September 2025].
02. Manne-Goehler J, Reich MR, Wirtz VJ. Access to care for Chagas disease in the United States: a health systems analysis. Am Soc Trop Med Hyg. 2015; 93(1): 108-13. https://doi.org/10.4269/ ajtmh.14-0826.
03. Henao-Martínez AF, Colborn K, Parra-Henao G. Overcoming research barriers in Chagas disease — designing effective implementation science. Parasitol Res. 2017; 116(1): 35-44. https://doi. org/10.1007/s00436-016-5291-z.
04. Chaves GC, Arrieche MA-S, Rode J, Mechali D, Reis PO, Alves RV, et al. Estimación de la demanda de medicamentos antichagásicos: una contribución para el acceso en América Latina. Rev Panam Salud Publica. 2017; 41: 1. https://doi.org/10.26633/RPSP.2017.45.
05. Echeverría LE, Marcus R, Novick G, Sosa-Estani S, Ralston K, Zaidel EJ, et al. WHF IASC roadmap on Chagas disease. Glob Heart. 2020; 15(1): 26. https://doi.org/10.5334/gh.484.
06. PAHO - Pan American Health Organization. Less than 10% of people with Chagas receive a diagnosis. PAHO/WHO. 2023. Available from: https://www.paho.org/en/news/13-4-2023-less-10-peoplechagas- receive-diagnosis (Accessed 15th December 2025).
07. Santos FLN, Costa VM, Silva RA. Chagas disease in Brazil: new challenges and perspectives for old problems. Mem Inst Oswaldo Cruz. 2025; 120: e240279. https://doi.org/10.1590/0074- 02760240279.
08. Lima-Costa MF, Peixoto SV, Ribeiro ALP. Chagas disease and mortality in old age as an emerging issue: 10 year follow-up of the Bambuí population-based cohort study (Brazil). Int J Cardiol. 2010; 145(2): 362-3. https://doi.org/10.1016/j.ijcard.2010.02.036.
09. Almeida ILGID, Oliveira LFLD, Figueiredo PHS, Oliveira RDDB, Damasceno TR, Silva WT, et al. The health-related quality of life in patients with Chagas disease: the state of the art. Rev Soc Bras Med Trop. 2022; 55: e0657-2021. https://doi.org/10.1590/0037- 8682-0657-2021.
10. Altcheh J, Moscatelli G, Caruso M, Moroni S, Bisio M, Miranda MR, et al. Population pharmacokinetics of benznidazole in neonates, infants and children using a new pediatric formulation. PLoS Negl Trop Dis. 2023; 17(5): e0010850. https://doi. org/10.1371/journal.pntd.0010850.
11. Cardoso CS, Ribeiro ALP, Oliveira CDL, Oliveira LC, Ferreira AM, Bierrenbach AL, et al. Beneficial effects of benznidazole in Chagas disease: NIH SaMi-Trop cohort study. PLoS Negl Trop Dis. 2018; 12(11): e0006814. https://doi.org/10.1371/journal.pntd.0006814.
12. Hasslocher-Moreno AM, Saraiva RM, Sangenis LHC, Xavier SS, de Sousa AS, Costa AR, et al. Benznidazole decreases the risk of chronic Chagas disease progression and cardiovascular events: a long-term follow up study. EClinicalMedicine. 2021; 31: 100694. https://doi.org/10.1016/j.eclinm.2020.100694.
13. Viotti R, Vigliano C, Lococo B, Bertocchi G, Petti M, Alvarez MG, et al. Long-term cardiac outcomes of treating chronic Chagas disease with benznidazole versus no treatment: a nonrandomized trial. Ann Intern Med. 2006; 144(10): 724-34. https://doi. org/10.7326/0003-4819-144-10-200605160-00006.
14. de Lana M, Suave LT, Assis JCSO, de Assis GFM, Milagre MM, Alessio GD, et al. Parasitological cure and clinical benefits of benznidazole treatment in patients from the Jequitinhonha Valley, MG, Brazil, with recent chronic infection by Trypanosoma cruzi II. Mem Inst Oswaldo Cruz. 2025; 120: e240229. https://doi. org/10.1590/0074-02760240229.
15. Camara EJN, Mendonca VRR, Souza LCL, Carvalho JS, Lessa RA, Gatto R, et al. Elevated IL-17 levels and echocardiographic signs of preserved myocardial function in benznidazole-treated individuals with chronic Chagas’ disease. Int J Infect Dis. 2019; 79: 123-30. https://doi.org/10.1016/j.ijid.2018.11.369.
16. Cruz CV, Rabinovich A, Moscatelli G, Moroni S, González N, Garcia‐ Bournissen F, et al. Adverse events associated with benznidazole treatment for Chagas disease in children and adults. Br J Clin Pharmacol. 2024; 90(12): 3334-47. https://doi.org/10.1111/bcp.16214.
17. Swett MC, Rayes DL, Campos SV, Kumar RN. Chagas disease: epidemiology, diagnosis, and treatment. Curr Cardiol Rep. 2024; 26(10): 1105-12. https://doi.org/10.1007/s11886-024-02113-7.
18. Alonso-Padilla J, Cotillo I, Presa JL, Cantizani J, Peña I, Bardera AI, et al. Automated high-content assay for compounds selectively toxic to Trypanosoma cruzi in a myoblastic cell line. PLoS Negl Trop Dis. 2015; 9(1): e0003493. https://doi.org/10.1371/ journal.pntd.0003493.
19. Alonso-Padilla J, Rodríguez A. High throughput screening for anti- Trypanosoma cruzi drug discovery. PLoS Negl Trop Dis. 2014; 8(12): e3259. https://doi.org/10.1371/journal.pntd.0003259.
20. Gabaldón-Figueira JC, Martinez-Peinado N, Escabia E, Ros- Lucas A, Chatelain E, Scandale I, et al. State-of-the-art in the drug discovery pathway for Chagas disease: a framework for drug development and target validation. Res Rep Trop Med. 2023; 14: 1-19. https://doi.org/10.2147/RRTM.S415273.
21. Canavaci AMC, Bustamante JM, Padilla AM, Brandan CMP, Simpson LJ, Xu D, et al. In vitro and in vivo high-throughput assays for the testing of anti-Trypanosoma cruzi compounds. PLoS Negl Trop Dis. 2010; 4(7): e740. https://doi.org/10.1371/journal. pntd.0000740.
22. Murta SMF, Santana PAL, Dit Lapierre TJWJ, Penteado AB, El Hajje M, Vinha TCN, et al. New drug discovery strategies for the treatment of benznidazole-resistance in Trypanosoma cruzi, the causative agent of Chagas disease. Expert Opin Drug Discov. 2024; 19(6): 741-53. https://doi.org/10.1080/17460441.2024.2349155.
23. Oliveira DSDS, Da Silva ED, Galvão Jr FF, Moura DMN, Sales- Junior PA, Brelaz-de-Castro MCA, et al. Reporter genes and transgenic Trypanosoma cruzi (Kinetoplastida, Trypanosomatidae): applications for screening new drugs against Chagas disease. Front Med. 2025; 12: 1591148. https://doi.org/10.3389/ fmed.2025.1591148.
24. Soeiro MNC, Sales-Junior PA, Pereira VRA, Vannier-Santos MA, Murta SMF, Sousa AS, et al. Drug screening and development cascade for Chagas disease: an update of in vitro and in vivo experimental models. Mem Inst Oswaldo Cruz. 2024; 119: e240057. https://doi.org/10.1590/0074-02760240057.
25. Freitas-Junior LH, Chatelain E, Kim HA, Siqueira-Neto JL. Visceral leishmaniasis treatment: What do we have, what do we need and how to deliver it? Int J Parasitol Drugs Drug Resist. 2012; 2: 11-9. https://doi.org/10.1016/j.ijpddr.2012.01.003.
26. Kratz JM, Gonçalves KR, Romera LMD, Moraes CB, Bittencourt- Cunha P, Schenkman S, et al. The translational challenge in Chagas disease drug development. Mem Inst Oswaldo Cruz. 2022; 117: e200501. https://doi.org/10.1590/0074-02760200501.
27. De Rycker M, Wyllie S, Horn D, Read KD, Gilbert IH. Anti-trypanosomatid drug discovery: progress and challenges. Nat Rev Microbiol. 2023; 21(1): 35-50. https://doi.org/10.1038/s41579- 022-00777-y.
28. Chatelain E, Ioset JR. Phenotypic screening approaches for Chagas disease drug discovery. Expert Opin Drug Discov. 2018; 13(2): 141-53. https://doi.org/10.1080/17460441.2018.1417380.
29. Khare S, Nagle AS, Biggart A, Lai YH, Liang F, Davis LC, et al. Proteasome inhibition for treatment of leishmaniasis, Chagas disease and sleeping sickness. Nature. 2016; 537(7619): 229-33. https://doi.org/10.1038/nature19339.
30. De Souza ML, Lapierre TJWJD, Marques GVL, Ferraz WR, Penteado AB, Trossini GHG, et al. Molecular targets for Chagas disease: validation, challenges and lead compounds for widely exploited targets. Expert Opin Ther Targets. 2023; 27(10): 911-25. https://doi.org/10.1080/14728222.2023.2264512.
31. Bisio MMC, Medina LSJ, García-Bournissen F, Gulin JEN. Listen to what the animals say: a systematic review and meta-analysis of sterol 14-demethylase inhibitor efficacy for in vivo models of Trypanosoma cruzi infection. Parasitol Res. 2024; 123(6): 248. https://doi.org/10.1007/s00436-024-08257-3.
32. Adasme MF, Bolz SN, Adelmann L, Salentin S, Haupt VJ, Moreno- Rodríguez A, et al. Repositioned drugs for Chagas disease unveiled via structure-based drug repositioning. Int J Mol Sci. 2020; 21(22): 8809. https://doi.org/10.3390/ijms21228809.
33. Jesus GCD, Ribeiro TS, Andricopulo AD, Ferreira LLG. Molecular docking and quantitative structure-activity relationships for a series of Trypanosoma cruzi dihydroorotate dehydrogenase inhibitors. Braz J Pharm Sci. 2025; 61: e24064. https://doi.org/10.1590/ s2175-97902025e24064.
34. Domingues KZA, De Fátima A, Fachi MM, Lazo REL, Ferreira L, Pontarolo R. Drug repurposing for trypanosomiasis: using machine learning models and polypharmacology to identify multitarget candidates. J Braz Chem Soc. 2025; https://doi. org/10.21577/0103-5053.20250028.
35. Mansoldo FRP, Carta F, Angeli A, Cardoso VDS, Supuran CT, Vermelho AB. Chagas disease: perspectives on the past and present and challenges in drug discovery. Molecules. 2020; 25(22): 5483. https://doi.org/10.3390/molecules25225483.
36. Andrade HM, Murta SMF, Chapeaurouge A, Perales J, Nirdé P, Romanha AJ. Proteomic analysis of Trypanosoma cruzi resistance to benznidazole. J Proteome Res. 2008; 7(6): 2357-67. https://doi. org/10.1021/pr700659m.
37. Hennig K, Abi-Ghanem J, Bunescu A, Meniche X, Biliaut E, Ouattara AD, et al. Metabolomics, lipidomics and proteomics profiling of myoblasts infected with Trypanosoma cruzi after treatment with different drugs against Chagas disease. Metabolomics. 2019; 15(9): 117. https://doi.org/10.1007/s11306-019-1583-5.
38. Trochine A, Creek DJ, Faral-Tello P, Barrett MP, Robello C. Benznidazole Biotransformation and multiple targets in Trypanosoma cruzi revealed by metabolomics. PLoS Negl Trop Dis. 2014; 8(5): e2844. https://doi.org/10.1371/journal.pntd.0002844.
39. Mejía-Jaramillo AM, Ospina-Zapata H, Fernandez GJ, Triana- Chávez O. Transcriptomic analysis of benznidazole-resistant Trypanosoma cruzi clone reveals nitroreductase I-independent resistance mechanisms. PLoS One. 2025; 20(2): e0314189. doi: 10.1371/ journal.pone.0314189.
40. García‐Huertas P, Mejía‐Jaramillo AM, González L, Triana‐ Chávez O. Transcriptome and functional genomics reveal the participation of adenine phosphoribosyltransferase in Trypanosoma cruzi resistance to benznidazole. J Cell Biochem. 2017; 118(7): 1936-45. https://doi.org/10.1002/jcb.25978.
41. Resende BC, Oliveira ACS, Guañabens ACP, Repolês BM, Santana V, Hiraiwa PM, et al. The influence of recombinational processes to induce dormancy in Trypanosoma cruzi. Front Cell Infect Microbiol. 2020; 10: 5. https://doi.org/10.3389/fcimb.2020.00005.
42. Lima DA, Gonçalves LO, Reis-Cunha JL, Guimarães PAS, Ruiz JC, Liarte DB, et al. Transcriptomic analysis of benznidazole-resistant and susceptible Trypanosoma cruzi populations. Parasit Vectors. 2023; 16(1): 167. https://doi.org/10.1186/s13071-023-05775-4.
43. Costa FC, Francisco AF, Jayawardhana S, Calderano SG, Lewis MD, Olmo F, et al. Expanding the toolbox for Trypanosoma cruzi: a parasite line incorporating a bioluminescence-fluorescence dual reporter and streamlined CRISPR/Cas9 functionality for rapid in vivo localisation and phenotyping. PLoS Negl Trop Dis. 2018; 12(4): e0006388. https://doi.org/10.1371/journal.pntd.0006388.
44. Lander N, Chiurillo MA, Storey M, Vercesi AE, Docampo R. CRISPR/Cas9-mediated endogenous C-terminal Tagging of Trypanosoma cruzi genes reveals the acidocalcisome localization of the inositol 1,4,5-trisphosphate receptor. J Biol Chem. 2016; 291(49): 25505-15. https://doi.org/10.1074/jbc.M116.749655.
45. Santi AMM, Murta SMF. Antioxidant defence system as a rational target for Chagas disease and Leishmaniasis chemotherapy. Mem Inst Oswaldo Cruz. 2022; 117: e210401. https://doi. org/10.1590/0074-02760210401.
46. Romanha AJ, Castro SL, Soeiro MNC, Lannes-Vieira J, Ribeiro I, Talvani A, et al. In vitro and in vivo experimental models for drug screening and development for Chagas disease. Mem Inst Oswaldo Cruz. 2010; 105(2): 233-8. https://doi.org/10.1590/S0074- 02762010000200022.
47. Francisco AF, Jayawardhana S, Olmo F, Lewis MD, Wilkinson SR, Taylor MC, et al. Challenges in Chagas disease drug development. Molecules. 2020; 25(12): 2799. https://doi.org/10.3390/molecules25122799.
48. Hall BS, Meredith EL, Wilkinson SR. Targeting the substrate preference of a type I nitroreductase to develop antitrypanosomal quinone-based prodrugs. Antimicrob Agents Chemother. 2012; 56(11): 5821-30. https://doi.org/10.1128/AAC.01227-12.
49. Hall BS, Wilkinson SR. Activation of benznidazole by trypanosomal type I nitroreductases results in glyoxal formation. Antimicrob Agents Chemother. 2012; 56(1): 115-23. https://doi. org/10.1128/AAC.05135-11.
50. Wilkinson SR, Kelly JM. Trypanocidal drugs: mechanisms, resistance and new targets. Expert Rev Mol Med. 2009; 11: e31. https:// doi.org/10.1017/S1462399409001252.
51. Frade VP, Simões NS, Couto NRB, Sanches C, Oliveira CDL. Ideal benznidazole dose regimen in chronic chagasic patients: a systematic review. Rev Inst Med Trop São Paulo. 2020; 62: e52. https://doi.org/10.1590/s1678-9946202062052.
52. Hernández Y, Marson ME, Fernández ML, Sued O, Frola C, Lloret SP, et al. Pharmacokinetics of two pharmaceutical presentations of benznidazole in adult Trypanosoma cruzi infected population. Mem Inst Oswaldo Cruz. 2025; 120: e240177. https://doi. org/10.1590/0074-02760240177.
53. Molina I, Salvador F, Sánchez-Montalvá A, Artaza MA, Moreno R, Perin L, et al. Pharmacokinetics of benznidazole in healthy volunteers and implications in future clinical trials. Antimicrob Agents Chemother. 2017; 61(4): e01912-16. https://doi.org/10.1128/ AAC.01912-16.
54. Perin L, Fonseca KDS, De Carvalho TV, Carvalho LM, Madeira JV, Medeiros LDF, et al. Low-dose of benznidazole promotes therapeutic cure in experimental chronic Chagas’ disease with absence of parasitism in blood, heart and colon. Exp Parasitol. 2020; 210: 107834. https://doi.org/10.1016/j.exppara.2020.107834.
55. Ramos LG, De Souza KR, Sales Júnior PA, Câmara CC, Castelo- Branco FS, Boechat N, et al. Tackling the challenges of human Chagas disease: a comprehensive review of treatment strategies in the chronic phase and emerging therapeutic approaches. Acta Trop. 2024; 256: 107264. https://doi.org/10.1016/j.actatropica. 2024.107264.
56. Mazzeti AL, Capelari-Oliveira P, Bahia MT, Mosqueira VCF. Review on experimental treatment strategies against Trypanosoma cruzi. J Exp Pharmacol. 2021; 13: 409-32. https://doi. org/10.2147/JEP.S267378.
57. Murta SMF, Romanha AJ. Characterization of Trypanosoma cruzi. Mem Inst Oswaldo Cruz. 1999; 94(Suppl. 1): 177-80. https:// doi.org/10.1590/S0074-02761999000700024.
58. Bunkofske ME, Sanchez-Valdez FJ, Tarleton RL. The importance of persistence and dormancy in Trypanosoma cruzi infection and Chagas disease. Curr Opin Microbiol. 2025; 86: 102615. https:// doi.org/10.1016/j.mib.2025.102615.
59. Espinoza‐Chávez RM, Rezende Júnior CO, Souza ML, Chelucci RC, Michelan‐Duarte S, Krogh R, et al. Structure‐metabolism relationships of benzimidazole derivatives with anti‐ Trypanosoma cruzi activity for Chagas disease. ChemMedChem. 2024; 19(20): e202400293. https://doi.org/10.1002/cmdc.202400293.
60. De Souza CC, De Azevedo-França JA, Barrias E, Cavalcante SCF, Vieira EG, Ferreira AMC, et al. Silver and copper-benznidazole derivatives as potential antiparasitic metallodrugs: synthesis, characterization, and biological evaluation. J Inorg Biochem. 2023; 239: 112047. https://doi.org/10.1016/j.jinorgbio.2022.112047.
61. De Melo IM, Camargo TP, Da Silva VA, Dos Santos EG, Caldas IS, Pontes CLM, et al. Discovery of a new eugenol-benznidazole hybrid active against different evolutive stages of Trypanosoma cruzi. Bioorganic Chem. 2025; 154: 107993. https://doi. org/10.1016/j.bioorg.2024.107993.
62. Pitombeira MCSR, Sales Júnior PA, Murta SMF, Romanha A, Luccas PH, Nonato MC, et al. New 2‐nitroimidazole‐ N ‐acylhydrazones, analogs of benznidazole, as anti‐ Trypanosoma cruzi agents. Arch Pharm (Weinheim). 2024; 357(7): 2400059. https:// doi.org/10.1002/ardp.202400059.
63. Carvalho DB, Costa PAN, Portapilla GB, Das Neves AR, Shiguemoto CYK, Pelizaro BI, et al. Design, synthesis and antitrypanosomatid activity of 2-nitroimidazole-3,5-disubstituted isoxazole compounds based on benznidazole. Eur J Med Chem. 2023; 260: 115451. https://doi.org/10.1016/j.ejmech.2023.115451.
64. Freire ES, Da Silva LP, Silva ADC, Vaz De Castro PAS, De Araújo GR, Otta DA, et al. New drugs and promising drug combinations in the treatment of Chagas disease in Brazil: a systematic review and meta-analysis. Arch Med Res. 2025; 56(1): 103084. https://doi. org/10.1016/j.arcmed.2024.103084.
65. Mazzeti AL, Gonçalves KR, Mota SLA, Pereira DE, Diniz LDF, Bahia MT. Combination therapy using nitro compounds improves the efficacy of experimental Chagas disease treatment. Parasitology. 2021; 148(11): 1320-7. https://doi.org/10.1017/ S0031182021001001.
66. Araujo-Lima CF, Christoni LSA, Justo G, Soeiro MNC, Aiub CAF, Felzenszwalb I. Atorvastatin downregulates in vitro methyl methanesulfonate and cyclophosphamide alkylation-mediated cellular and DNA injuries. Oxid Med Cell Longev. 2018; 2018: 7820890. https://doi.org/10.1155/2018/7820890.
67. Almeida-Silva J, Menezes DS, Fernandes JMP, Almeida MC, Vasco-dos-Santos DR, Saraiva RM, et al. The repositioned drugs disulfiram/diethyldithiocarbamate combined to benznidazole: searching for Chagas disease selective therapy, preventing toxicity and drug resistance. Front Cell Infect Microbiol. 2022; 12: 926699. https://doi.org/10.3389/fcimb.2022.926699.
68. Saraiva RM, Portela LF, Silveira GPED, Gomes NLDS, Pinto DP, Silva ACAD, et al. Disulfiram repurposing in the combined chemotherapy of Chagas disease: a protocol for phase I/II clinical trial. Med Case Rep Study Protoc. 2021; 2(7): e0110. https://doi. org/10.1097/MD9.0000000000000110.
69. Gulin JEN, Bisio MMC, Rocco D, Altcheh J, Solana ME, García- Bournissen F. Miltefosine and benznidazole combination improve anti-Trypanosoma cruzi in vitro and in vivo efficacy. Front Cell Infect Microbiol. 2022; 12: 855119. https://doi.org/10.3389/ fcimb.2022.855119.
70. Pandey RP, Nascimento MS, Franco CH, Bortoluci K, Silva MN, Zingales B, et al. Drug repurposing in Chagas disease: chloroquine potentiates benznidazole activity against Trypanosoma cruzi in vitro and in vivo. Antimicrob Agents Chemother. 2022; 66(11): e00284-22. https://doi.org/10.1128/aac.00284-22.
71. De Melo GD, Coatnoan N, Gouault N, Cupif JF, Renault J, Cosson A, et al. Prodrugs as new therapies against Chagas disease: in vivo synergy between Trypanosoma cruzi proline racemase inhibitors and benznidazole. J Glob Antimicrob Resist. 2022; 28: 84-9. https://doi.org/10.1016/j.jgar.2021.10.030.
72. Vilas-Boas DF, Oliveira RRG, Gonçalves-Santos E, Silva LS, Diniz LF, Mazzeti AL, et al. 4-nitrobenzoylcoumarin potentiates the antiparasitic, anti-inflammatory and cardioprotective effects of benznidazole in a murine model of acute Trypanosoma cruzi infection. Acta Trop. 2022; 228: 106314. https://doi.org/10.1016/j. actatropica.2022.106314.
73. González S, Wall RJ, Thomas J, Braillard S, Brunori G, Díaz IC, et al. Short-course combination treatment for experimental chronic Chagas disease. Sci Transl Med. 2023; 15(726): eadg8105. https://doi.org/10.1126/scitranslmed.adg8105.
74. Ortiz JE, Piñeiro M, Martinez-Peinado N, Barrera P, Sosa M, Bastida J, et al. Candimine from Hippeastrum escoipense (Amaryllidaceae): anti-Trypanosoma cruzi activity and synergistic effect with benznidazole. Phytomedicine. 2023; 114: 154788. https://doi. org/10.1016/j.phymed.2023.154788.
75. Barbosa JMC, Pedra-Rezende Y, Mata-Santos HA, Vilar-Pereira G, Melo TGD, Ramos IP, et al. Preclinical evaluation of combined therapy with amiodarone and low-dose benznidazole in a mouse model of chronic Trypanosoma cruzi infection. Biomed Pharmacother. 2024; 175: 116742. https://doi.org/10.1016/j.biopha. 2024.116742.
76. Jones K, Versteeg L, Damania A, Keegan B, Kendricks A, Pollet J, et al. Vaccine-linked chemotherapy improves benznidazole efficacy for acute Chagas disease. Infect Immun. 2018; 86(4): e00876- 17. https://doi.org/10.1128/IAI.00876-17.
77. Prochetto E, Bontempi I, Rodeles L, Cabrera G, Vicco M, Cacik P, et al. Assessment of a combined treatment with a therapeutic vaccine and benznidazole for the Trypanosoma cruzi chronic infection. Acta Trop. 2022; 229: 106334. https://doi.org/10.1016/j. actatropica.2022.106334.
78. Joshi G, Quadir SS, Yadav KS. Road map to the treatment of neglected tropical diseases: Nanocarriers interventions. J Controlled Release. 2021; 339: 51-74. https://doi.org/10.1016/j.jconrel. 2021.09.020.
79. Gomes DC, Medeiros TS, Pereira ELA, Da Silva JFO, Oliveira JWF, Fernandes-Pedrosa MDF, et al. From benznidazole to new drugs: nanotechnology contribution in Chagas disease. Int J Mol Sci. 2023; 24(18): 13778. https://doi.org/10.3390/ijms241813778.
80. Alvarez VE, Iribarren PA, Niemirowicz GT, Cazzulo JJ. Update on relevant trypanosome peptidases: validated targets and future challenges. Biochim Biophys Acta Proteins Proteomics. 2021; 1869(2): 140577. https://doi.org/10.1016/j.bbapap.2020.140577.
81. Barros-Alvarez X, Gualdrón-López M, Acosta H, Cáceres AJ, Graminha MS, Michels PAM, et al. Glycosomal targets for antitrypanosomatid drug discovery. Curr Med Chem. 2014; 21(15): 1679-706. https://doi.org/10.2174/09298673113209990139.
82. García-Huertas P, Cardona-Castro N. Advances in the treatment of Chagas disease: promising new drugs, plants and targets. Biomed Pharmacother. 2021; 142: 112020. https://doi. org/10.1016/j.biopha.2021.112020.
83. Lepesheva GI, Villalta F, Waterman MR. Targeting Trypanosoma cruzi sterol 14α-demethylase (CYP51). Adv Parasitol. 2011; 75: 65-87. https://doi.org/10.1016/B978-0-12-385863-4.00004-6.
84. Soeiro MNC, De Souza EM, Da Silva CF, Batista DDGJ, Batista MM, Pavão BP, et al. In vitro and in vivo studies of the antiparasitic activity of sterol 14α-Demethylase (CYP51) inhibitor VNI against drug-resistant strains of Trypanosoma cruzi. Antimicrob Agents Chemother. 2013; 57(9): 4151-63. https:// doi.org/10.1128/AAC.00070-13.
85. Guedes-da-Silva FH, Batista DDGJ, Da Silva CF, Pavão BP, Batista MM, Moreira OC, et al. Successful aspects of the coadministration of sterol 14α-Demethylase inhibitor VFV and benznidazole in experimental mouse models of Chagas disease caused by the drug-resistant strain of Trypanosoma cruzi. ACS Infect Dis. 2019; 5(3): 365-71. https://doi.org/10.1021/acsinfecdis.8b00253.
86. Molina I, Gómez I Prat J, Salvador F, Treviño B, Sulleiro E, Serre N, et al. Randomized trial of posaconazole and benznidazole for chronic Chagas’ disease. N Engl J Med. 2014; 370(20): 1899-908. https://doi.org/10.1056/NEJMoa1313122.
87. Torrico F, Gascon J, Ortiz L, Alonso-Vega C, Pinazo MJ, Schijman A, et al. Treatment of adult chronic indeterminate Chagas disease with benznidazole and three E1224 dosing regimens: a proof-ofconcept, randomised, placebo-controlled trial. Lancet Infect Dis. 2018; 18(4): 419-30. https://doi.org/10.1016/S1473-3099(17)30538-8.
88. Francisco AF, Lewis MD, Jayawardhana S, Taylor MC, Chatelain E, Kelly JM. Limited ability of posaconazole to cure both acute and chronic Trypanosoma cruzi infections revealed by highly sensitive in vivo imaging. Antimicrob Agents Chemother. 2015; 59(8): 4653-61. https://doi.org/10.1128/AAC.00520-15.
89. MacLean LM, Thomas J, Lewis MD, Cotillo I, Gray DW, De Rycker M. Development of Trypanosoma cruzi in vitro assays to identify compounds suitable for progression in Chagas’ disease drug discovery. PLoS Negl Trop Dis. 2018; 12(7): e0006612. https://doi.org/10.1371/journal.pntd.0006612.
90. Cherkesova TS, Hargrove TY, Vanrell MC, Ges I, Usanov SA, Romano PS, et al. Sequence variation in CYP51A from the Y strain of Trypanosoma cruzi alters its sensitivity to inhibition. FEBS Lett. 2014; 588(21): 3878-85. https://doi.org/10.1016/j.febslet. 2014.08.030.
91. Rodriguez Gini AL, Da Cunha PST, João EE, Man Chin C, Dos Santos JL, Serra EC, et al. TrypPROTACs unlocking new therapeutic strategies for Chagas disease. Pharmaceuticals. 2025; 18(6): 919. https://doi.org/10.3390/ph18060919.
92. Meier A, Erler H, Beitz E. Targeting channels and transporters in protozoan parasite infections. Front Chem. 2018; 6: 88. https://doi. org/10.3389/fchem.2018.00088.
93. Da Costa KM, Salustiano EJ, Valente RC, Freire-de-Lima L, Mendonça- Previato L, Previato JO. Thiol efflux mediated by an ABCClike transporter participates for Trypanosoma cruzi adaptation to environmental and chemotherapeutic stresses. bioRxiv [Preprint]. 2020; 03.26.009753. https://doi.org/10.1101/2020.03.26.009753.
94. Petravicius PO, Costa-Martins AG, Silva MN, Reis-Cunha JL, Bartholomeu DC, Teixeira MMG, et al. Mapping benznidazole resistance in trypanosomatids and exploring evolutionary histories of nitroreductases and ABCG transporter protein sequences. Acta Trop. 2019; 200: 105161. https://doi.org/10.1016/j.actatropica.2019.105161.
95. Lima DA, Costa-Silva HM, Albergaria KSS, Ribeiro JM, Resende DDM, Santarossa BA, et al. Glycosomal ABC transporter 3 (GAT3) deletion enhances the oxidative stress responses and reduces the infectivity of Trypanosoma cruzi. PLoS Negl Trop Dis. 2025; 19(9): e0013479. https://doi.org/10.1371/journal.pntd.0013479.
96. Cruz-Bustos T, Potapenko E, Storey M, Docampo R. An intracellular ammonium transporter is necessary for replication, differentiation, and resistance to starvation and osmotic stress in Trypanosoma cruzi. mSphere. 2018; 3(1): e00377-17. https://doi. org/10.1128/mSphere.00377-17.
97. Veira C, Benítez D, Pérez-Díaz L, Álvarez G, Cerecetto H, Aguilera E. Looking for approved-medicines to be repositioned as anti-Trypanosoma cruzi agents. Identification of new chemotypes with good individual- or in combination-biological behaviours. Mem Inst Oswaldo Cruz. 2025; 120: e240183. https://doi. org/10.1590/0074-02760240183.
98. Pandey AM, Malwal SR, Valladares-Delgado M, Labrador- Fagúndez L, Stella BG, Díaz-Pérez LJ, et al. Anti-parasitics with a triple threat: targeting parasite enzymes, the proton motive force, and host cell-mediated killing. ACS Infect Dis. 2025; 11(6): 1539- 51. https://doi.org/10.1021/acsinfecdis.5c00094.
99. Cabello-Donayre M, Cabello-Donayre I, Guerra D, Orrego LM, Morales JC, Cautain B, et al. A yeast-based high-throughput screen identifies inhibitors of trypanosomatid HRG heme transporters with potent leishmanicidal and trypanocidal activity. Int J Antimicrob Agents. 2024; 63(3): 107092. https://doi.org/10.1016/j. ijantimicag.2024.107092.
100. Tevere E, Mediavilla MG, Di Capua CB, Merli ML, Robello C, Berná L, et al. Transcriptomic response to different heme sources in Trypanosoma cruzi. BioRxiv Prepr Serv Biol [Preprint]. 2025; 2025.07.14.664770. https://doi.org/10.1101/2025.07.14.664770.
Financial support: CNPq (309994/2023-3 to SMFM; 446909/2024-6 to AMMS), FIOCRUZ (Chamada de Redes Fiocruz Minas / Rede FarVac; Programa INOVA FIOCRUZ), FAPEMIG (RED-00110-23; APD-01401-25). SMFM is research fellow supported by CNPq.
+ Corresponding author: silvane.murta@fiocruz.br | 
http://orcid.org/0000-0002-8523-2155
Received 23 October 2025
Accepted 22 January 2026
HOW TO CITE
Santi AMM, Lima DA, Murta SMF. Advances and challenges in the search for new treatments for Chagas disease. Mem Inst Oswaldo Cruz. 2026; 121(Suppl. 2): e250299.
HANDLING EDITOR
Rubem Figueiredo Sadok Menna-Barreto | https://orcid.org/0000-0002-1352-0641
