The lignan glycosides lyoniside and saracoside poison the unusual type IB topoisomerase of Leishmania donovani and kill the parasite both in vitro and in vivo

Abstract

Lignans are diphenyl propanoids with vast range of biological activities. The present study provides an important insight into the anti-leishmanial activities of two lignan glycosides, viz. lyoniside and saracoside. These compounds inhibit catalytic activities of topoisomerase IB (LdTopIB) of Leishmania donovani in non-competitive manner and stabilize the LdTopIB mediated cleavage complex formation both in vitro and in Leishmania promastigotes and subsequently inhibit the religation of cleaved strand. These two compounds not only poison LdTopIB but also can interact with the free enzyme LdTopIB. We have also shown that lyoniside and saracoside are cytotoxic to promastigotes and intracellular amastigotes. The protein-DNA complex formation leads to double strand breaks in DNA which ultimately triggers apoptosis-like cell death in the parasite. Along with their cytotoxicity towards sodium antimony gluconate (SAG) sensitive AG83 strain, their ability to kill SAG resistant GE1 strain makes these two compounds potential anti-leishmanial candidates. Not only they effectively kill L. donovani amastigotes inside macrophages in vitro, lyoniside and saracoside demonstrated strong anti-leishmanial efficacies in BALB/c mice model of leishmaniasis. Treatment with these lignan glycosides produce nitric oxide and reactive oxygen species which result in almost complete clearance of the liver and splenic parasite burden. These compounds do not inhibit human topoisomerase IB upto 200μM concentrations and had poor cytotoxic effect on uninfected cultured murine peritoneal macrophages upto 100μM concentrations. Taken together it can be concluded that these compounds can be developed into excellent therapeutic agent against deadly disease leishmaniasis.

Keywords: Antileishmanial agent; Apoptosis; DMSO; DNA topoisomerase IB poison; DTT; EtBr; FBS; IPTG; K(D); L. donovani topoisomerase I large subunit; L. donovani topoisomerase I small subunit; L. donovani topoisomerase IB; LdTOP1L; LdTOP1S; LdTopIB; Lignan glycosides; M199; Ni(2+)-nitriloacetate-agarose; Ni-NTA; P11 cellulose; SAG; V(max); dimethyl sulphoxide; dissociation constant; dithiothreitol; ethidium bromide; fetal bovine serum; hTopIB; human topoisomerase IB; isopropyl β-D-thiogalactopyranoside; maximal velocity; medium199; phosphocellulose; sodium antimony gluconate.