While searching for novel anti-echinococcosis drugs, we have been focusing on glycolysis which is relied on byEchinococcusfor energy production and intermediates for other metabolic processes. The aim of this study was to investigate the potential therapeutic implication of glycolytic inhibitors onEchinococcus. Our results demonstrate that at an initial concentration of 40 μM, all inhibitors of glycolysis used in the current experiment [3-bromopyruvate (3-BrPA), ornidazole, clorsulon (CLS), sodium oxamate and 2,6-dihydroxynaphthalene (NA-P₂)] show considerable in vitro effects againstEchinococcus granulosusprotoscoleces andEchinococcus multilocularismetacestodes. Among them, 3-BrPA exhibited the highest activity which was similar to that of nitazoxanide (NTZ) and more efficacious than albendazole (ABZ). The activity of 3-BrPA was dose dependent and resulted in severe ultrastructural destructions, as visualized by electron microscopy. An additional in vivo study in mice infected withE. multilocularismetacestodes indicates a reduction in parasite weight after the twice-weekly treatment of 25 mg/kg 3-BrPA for 6 weeks, compared to that of the untreated control. In particular, in contrast to ABZ, the administration of 25 mg/kg 3-BrPA did not cause toxicity to the liver and kidney in mice. Similarly, at the effective dose againstEchinococcuslarvae, 3-BrPA showed no significant toxicity to human hepatocytes. Taken together, the results suggest that interfering with the glycolysis of the parasite may be a novel chemotherapeutical option and 3-BrPA, which exhibited a remarkable activity againstEchinococcus, may be a promising potential drug against cystic echinococcosis (CE) and alveolar echinococcosis (AE).

2-Deoxy-D-glucose (2-DG) is a glucose analog used as a promising anticancer agent. It exerts its effects by inhibiting the glycolytic energy metabolism to deplete cells of energy. The larval stage of Echinococcus relies on glycolysis for energy production. Therefore, in this study, we investigated the in vitro and in vivo efficacy of 2-DG against the larval stage of Echinococcus granulosus and E. multilocularis. 2-DG exhibited significant time- and dose-dependent effects against in vitro cultured E. granulosus protoscoleces and E. multilocularis metacestodes. A daily oral administration of 500 mg/kg 2-DG in E. multilocularis-infected mice effectively reduced the weight of metacestodes. Notably, the combination treatment, either 2-DG (500 mg/kg/day) + albendazole (ABZ) (200 mg/kg/day) or 2-DG (500 mg/kg/day) + half-dose of ABZ (100 mg/kg/day), exhibited a potent therapeutic effect against E. multilocularis, significantly promoting the reduction of metacestodes weight compared with the administration of 2-DG or ABZ alone. Furthermore, the combination significantly promoted apoptosis of the cells of metacestodes and inhibited glycolysis in metacestodes, compared with the administration of 2-DG or ABZ alone. In conclusion, 2-DG exerts an effective activity against the larval stage of Echinococcus. Thus, it may be a promising anti-Echinococcus drug, and its combination with ABZ may provide a new strategy for the treatment of echinococcosis in humans.

Background: Cystic echinococcosis (CE), caused by the larval stage of Echinococcus granulosus (sensu stricto), is a life-threatening but neglected zoonosis. Glycolytic enzymes are crucial molecules for the survival and development of E. granulosus. The aim of this study was to investigate the molecular characterization, immunogenicity, tissue distribution and serodiagnostic potential of E. granulosus hexokinase (EgHK), the first key enzyme in the glycolytic pathway. Methods: EgHK was cloned and expressed in Escherichia coli. Specific serum antibodies were evaluated in mice immunized with recombinant EgHK (rEgHK). The location of EgHK in the larval stage of E. granulosus was determined using fluorescence immunohistochemistry, and the potential of rEgHK as a diagnostic antigen was investigated in patients with CE using indirect enzyme-linked immunosorbent assay (ELISA). Results: Recombinant EgHK could be identified in the sera of patients with CE and in mouse anti-rEgHK sera. High titers of specific immunoglobulin G were induced in mice after immunization with rEgHK. EgHK was mainly located in the tegument, suckers and hooklets of protoscoleces and in the germinal layer and laminated layer of the cyst wall. The sensitivity and specificity of the rEgHK-ELISA reached 91.3% (42/46) and 87.8% (43/49), respectively. Conclusions: We have characterized the sequence, structure and location of EgHK and investigated the immunoreactivity, immunogenicity and serodiagnostic potential of rEgHK. Our results suggest that EgHK may be a promising candidate for the development of vaccines against E. granulosus and an effective antigen for the diagnosis of human CE. Graphical

Echinococcosis is a zoonotic disease caused by cestode species of the genus Echinococcus, which demonstrates considerable medical and veterinary concerns. The development of novel drugs for echinococcosis treatment is urgently needed. In this study, we demonstrated that lonidamine (LND) and 6-aminonicotinamide (6-AN) exhibited considerable in vitro effects against both larval- and adult-stage of E. granulosussensu stricto (s. s.) and E. multilocularis. The combination of LND and 6-AN exhibited a significantly higher activity than the single drug treatment. These results highlight the therapeutic potential of LND, 6-AN and the combination of LND and 6-AN for the treatment of echinococcosis.

The changes of biotransformation enzymes will substantially affect the host's ability to metabolize drugs and other xenobiotic compounds. In order to further elucidate this process and promote the development in treatment of echinococcosis, we investigated the effects of Echinococcus multilocularis infection and drug treatment on biotransformation enzymes in mouse liver. In microsomal and cytosolic fractions, from the six activities assayed, significant decrease of glutathione S-transferases (GST) activity and significant increase of 7-pentoxyresorufin (PROD) and NADPH-cytochrome P450 reductase (CPR) activity were observed in the mice infected with E. multilocularis metacestodes. In addition, after six weeks treatment of albendazole in E. multilocularis infected mice, significant decreased GST activity and significant increase of 7- ethoxyresorufin (EROD), PROD, and particularly 3-fold higher 7-methoxyresorufin (MROD) activity were observed. The 3-bromopyruvate treated mice only exhibited significantly lower GST activity. Our results demonstrate that E. multilocularis metacestodes infection can affect the activities of main hepatic biotransformation enzymes and such alterations of activity may further affect the hepatic biotransformation of xenobiotics. Moreover, albendazole and 3-bromopyruvate, the promising potential drug against Echinococcus, affected different hepatic biotransformation enzymes and may affect their metabolism. The findings will help to develop rational treatments with less side effects and promote the development of more efficient treatments against E. multilocularis.

Alveolar echinococcosis (AE) is a fatal zoonosis caused by the larval stage of Echinococcus multilocularis , but there are no fully effective treatments for this disease to date. The aim of this study was to investigate the potential therapeutic implication of photodynamic therapy (PDT) for E. multilocularis . We synthesized a novel nanophotosensitizer, TPEDCC-PEG, with aggregation-induced emission (AIE) properties and investigated its in vitro and in vivo PDT effects against the larval stage of E. multilocularis , as well as the anti- Echinococcus mechanism. TPEDCC-PEG exhibited excellent physicochemical properties and biocompatibility as a nanophotosensitizer. TPEDCC-PEG PDT, the treatment of TPEDCC-PEG with 410 nm laser irradiation, showed significant dose- and time- dependent protoscolicidal effect on in vitro cultured protoscoleces. Meanwhile, the infection experiments in vivo showed that the growth of protoscoleces survived after TPEDCC-PEG PDT was significantly suppressed in vitro , and the weight of metacestodes resulting from such protoscoleces in TPEDCC-PEG PDT mice was significant less than that in controls. Once every 3 days PDT administration (410 nm, 10 min) of TPEDCC-PEG (10 mg/kg) for 9 days in E. multilocularis -infected mice was highly effective in reducing the volume of metacestode cysts, resulting in a significant inhibition in metacestodes growth and severe structural destructions of metacestodes. Furthermore, TPEDCC-PEG PDT significantly increased the production of reactive oxygen species (ROS), inhibited the expression of glutathione in protoscoleces, and was accompanied by the depolarization of mitochondrial membrane potential. Western blot analysis revealed significantly reduced expression of the Bcl-2 protein alongside increased expression of caspase-3, Bax, and cytochrome c (Cyt C) proteins in protoscoleces following TPEDCC-PEG PDT, which hints the induction of apoptosis would associate with oxidative stress and involve the mitochondrial pathway. Taken together, the results suggest that PDT with TPEDCC-PEG, which exerted potent parasiticidal effects against E. multilocularis both in vitro and in vivo , could thus serve as a promising therapeutic option for echinococcosis.