Fundação de Amparo à Pesquisa do Estado de São Paulo [grant number 2018/09948-0 to NSM and 2015/22031-0 to SS]; Fundação de Amparo à Pesquisa do Estado da Bahia [grant number PNX0002/2014]; and Conselho Nacional de Desenvolvimento Científico Tecnológico [grant number 424729/2018-0 to NSM and 477143/2011-3, 445655/2014-3, and INCTV, to SS], PRIN 2016 (prot. 20152TE5PK) (to AM), AIRC 2016 (n. 19162) (to AM), and PE-2013-02355271 (to AM), PRIN 2017 (prot. 2017JL8SRX) (to RR).Chagas disease is an illness caused by the protozoan parasite Trypanosoma cruzi, a ecting more than 7 million people in the world. Benznidazole and nifurtimox are the only drugs available for treatment and in addition to causing several side e ects, are only satisfactory in the acute phase of the disease. Sirtuins are NAD+-dependent deacetylases involved in several biological processes, which have become drug target candidates in various disease settings. T. cruzi presents two sirtuins, one cytosolic (TcSir2rp1) and the latter mitochondrial (TcSir2rp3). Here, we characterized the e ects of human sirtuin inhibitors against T. cruzi sirtuins as an initial approach to develop specific parasite inhibitors. We found that, of 33 compounds tested, two inhibited TcSir2rp1 (15 and 17), while other five inhibited TcSir2rp3 (8, 12, 13, 30, and 32), indicating that specific inhibitors can be devised for each one of the enzymes. Furthermore, all inhibiting compounds prevented parasite proliferation in cultured mammalian cells. When combining the most e ective inhibitors with benznidazole at least two compounds, 17 and 32, demonstrated synergistic e ects. Altogether, these results support the importance of exploring T. cruzi sirtuins as drug targets and provide key elements to develop specific inhibitors for these enzymes as potential targets for Chagas disease treatment