Design and screening of novel molecular compounds targeting lactate dehydrogenase of Babesia microti

Abstract

Abstract Background Human babesiosis is caused by several species within the Babesia genus, primarily Babesia microti, Babesia duncani, and Babesia divergens, all of which infect human red blood cells (RBCs). Clinically, the disease manifests with symptoms such as fever, anemia, jaundice, and hemoglobinuria, with B. microti being the most prevalent of these species. Our previous research has shown that B. microti primarily relies on lactate dehydrogenase (LDH)-mediated anaerobic glycolysis, rather than the tricarboxylic acid cycle (TCA cycle), to generate ATP for its intracellular survival. Because LDH is a promising drug target, it can be inhibited by compounds such as gossypol and 3,5-dihydroxy-2-naphthoxylic acid (DHNA). In this study, we conduct a structure-based optimization of DHNA, leading to the development of a novel library of compounds derived from its structure. Methods Two compounds were identified and synthesized through molecular docking, on the basis of the crystal structure of Babesia microti lactate dehydrogenase (BmLDH). The effects of these compounds were evaluated using several methods, including surface plasmon resonance (SPR) assays, enzyme activity inhibition tests, in vitro growth inhibition assays against B. microti, and mammalian cytotoxicity tests. Results Compounds target A (TA) (−36.0) and B (TB) (−43.8), both exhibiting low CDOCKER energy values, achieved final purities of 96.6% and 97.5%, respectively. Surface plasmon resonance (SPR) experiments showed that TA and TB had comparable dissociation constant (K D) values of 11.3 × 10−6 M and 13.2 × 10−6 M, respectively. However, enzyme activity inhibition assays indicated that TB was more potent, with an half-maximal inhibitory concentration (IC50) value of 23.8 μM, compared with TA’s IC50 of 71.6 μM. Additionally, TB demonstrated a strong ability to inhibit the in vitro growth of B. microti, with an IC50 value of 111.7 μM. Conclusions In this study, two compounds capable of inhibiting the growth of B. microti were obtained. Although both compounds showed moderate inhibitory activity against recombinant BmLDH (rBmLDH) and the growth of B. microti, there is potential to enhance their efficacy through further structural modifications, particularly of compound TB. Graphical Abstract

Keywords

• Babesia microti
• Lactate dehydrogenase
• Compound
• Computer-aided drug design
• Surface plasmon resonance