4. Discussion
It is widely known that, besides the well-known limitations of current treatments, other drawbacks are the natural resistance of the T. cruzi genotype to the drugs used and the cross-resistance of current drugs (Wilkinson et al. , 2008; Mejia et al. , 2012). Here, the in vitro resistance of the three morphological forms ofT. cruzi Arequipa to the reference drug BNZ was characterized.T. cruzi Arequipa shows moderate resistance to BNZ compared to other strains used for CD drug discovery. It is well known that resistance to BNZ is strain-dependent, with IC50 values ranging from 1 µM to over 200 µM (Vela et al. , 2021). Therefore, ignoring the parasite’s genetic variability during CD drug discovery is not advisable.
Regarding T. cruzi infection, it should be noted that it is dependent on the genetic composition of the infecting T. cruzistrain (Toledo et al. , 2004; Santos et al. , 2010; Rodriguez et al. , 2014) and the genetic background of the animal model used as a host (Caldas et al. , 2008), that is, the host-parasite interactions (Campbell et al. , 2004). Hence, it is really important to know the infective capacity of the strains used for CD drug discovery, that is, virulence (quantitative) and tropism (qualitative) in BALB/c mice, before in vivo assays (Rodriguezet al. , 2014). T. cruzi is able to parasitize a large variety of cells (de Souza et al. , 2010; Rodriguez et al. , 2014; Lewis and Kelly, 2016; Lewis et al. , 2018) and its tissue homing ability has been reported to be strain-specific (Tibayrenc and Telleria, 2010). Here, the acute-phase parasitaemia and the chronic-phase tropism of T. cruzi Arequipa were evaluated in BALB/c mice in order to use this strain as a suitable tool for CD drug discovery.
It should be noted that most in vivo chemotherapy has focused on acute-phase infections, partially because it is simpler to monitor the course of parasitaemia (Canavaci et al. , 2010; Romanha et al. , 2010; Buckner, 2011; Chatelain and Konar, 2015). However, the ability to cure chronic-phase infections is the main need from a clinical viewpoint. Effectiveness of current drugs is especially limited during the chronic-phase of CD (Wilkinson et al. , 2008; Chatelain, 2014; Scarim et al. , 2018), so chronic-phase infections should be the main research focus in animal models (DNDi. Drugs for Neglected Diseases Initiative, 2018). Notably, this strain does not cause mortality, which is ultimately useful for comparing thein vivo trypanocidal efficacy of potential compounds and the reference drug BNZ in late-chronic phase.
T. cruzi Arequipa shows an attractive tropism for the evaluation of potential new compounds for the treatment of CD. T. cruziArequipa shows tropism towards: 1) brain, which allows us to evaluate the ability of potential compounds to cross the blood-brain barrier; 2) adipose tissue, which allows us to evaluate the solubility of compounds in lipid rich environments; and 3) heart, which allows us to use this strain as a model for cardiac CD. It has to be highlighted that these organs/tissues are the ones that have maintained the infection most after treatment using current potential candidates for the treatment of CD (including BNZ), showing nested parasites in them (Ferreira et al. , 2011; Tanowitz et al. , 2016); lesser drug accessibility or parasite susceptibility in these environments could be the reasons for the lower efficacy (Nagajyothi et al. , 2013; Tanowitz et al. , 2016). It has been postulated that inappropriate pharmacokinetics/pharmacodynamics between current drugs and tissue location of parasites is linked to the inability to reliably cure chronic infections (Urbina, 2002; Perin et al. , 2017).
Currently, drugs against CD present variable activity in the acute- and chronic-phases of the disease, and the effectiveness of treatments, especially during the chronic-phase, is not as effective as they should be (Martín-Escolano et al. , 2020a). For these reason, thein vivo resistance of T. cruzi Arequipa in both acute- and chronic-phase was evaluated. The treatment strategy using BALB/c mice was as follows: 1) Drugs were administered orally because it is the preferred route for the treatment of parasitic diseases in developing countries, it leads to better patient compliance, and it has a low cost (Espuelas et al. , 2012; DNDi. Drugs for Neglected Diseases Initiative, 2018); 2) Given that a compound showing a reasonable parasitaemia reduction following 5 days of treatment can be defined as a lead compound (Chatelain, 2014), the treatment guideline was for 5 consecutive days in order to evaluate the resistance of T. cruziArequipa to oral BNZ treatment. Moreover, to evaluate if potential compounds (for further research) show higher trypanocidal activity than BNZ in the first in vivo screening phase (Romanha et al. , 2010), the treatment was at subcurative doses of BNZ (20 mg·kg-1 per day). As expected, mice treated in the chronic-phase showed lower infection levels than those treated in the acute-phase. It is widely known that BNZ is more effective in chronic phase, probably because the parasite burden is significantly low and limited to far few locations (Francisco et al. , 2016).