H-2-168

Background: Harmine (HM) has several pharmacological effects; however, severe neurotoxicity limits its clinical application and development. HM neurotoxicity is associated with abnormal energy metabolism. This study aimed to explore the roles and underlying mechanisms of mitochondrial fusion and division in HM derivative H-2-168-induced neurotoxicity. Methods: PC12 cells were treated with H-2-168, Mdivi-1 (an inhibitor of mitochondrial division), or a combination of both. Cell viability, levels of reactive oxygen species (ROS), adenosine triphosphate (ATP), lactic dehydrogenase (LDH), mitochondrial morphology, and membrane potential were measured. Immunofluorescence (IF) and western blotting were used to determine the expression of apoptosis-, mitochondrial fusion-, and division-related proteins. Additionally, PC12 cells with Drp1 knockdown or Mfn2 overexpression were generated to explore their effects. Results: H-2-168 alone or in combination with Mdivi-1 significantly reduced PC12 cell viability, induced apoptosis, and impaired mitochondrial function. These effects were accompanied by increased levels of ROS and LDH, reduced ATP levels, upregulation of caspase-3, cytochrome c (Cyt-c), Drp1, and Fis1, and downregulation of Mfn2 and OPA1. Additionally, Drp1 knockdown or Mfn2 overexpression further enhanced the H-2-168-induced reduction in cell viability. Conclusions: These data implied that H-2-168 may initiate apoptosis in PC12 cells by influencing the balance between mitochondrial fusion and division, accompanied by changes in energy metabolism, which may induce neurotoxicity.

Cystic echinococcosis (CE) is a chronic zoonotic parasitic disease caused by the larvae of the Echinococcus granulosus sensu lato (s.l.) cluster. The current existing drugs have limited therapeutic efficacy against cystic echinococcosis, and thus, there is an urgent need to develop new drugs.

In this study, 7 harmine (HM) derivatives were screened and the effects of HM derivatives on E. granulosus sensu stricto (s.s.) were evaluated by in vitro and mouse experiments. The safety of the HM derivatives was assessed by cytotoxicity assays, acute toxicity study in animals and subacute toxicity study.

These results show that the HM derivatives H-2-168 and DH-004 exhibited more significant antiparasitic effects at an initial concentration of 40 μM. The results of further studies showed that H-2-168 and DH-004 had dose-dependent effects against protoscoleces and had satisfactory therapeutic outcomes in vivo. Electron microscopy observations demonstrated that H-2-168 and DH-004 caused severe disruption of the parasite ultrastructure. Notably, the results of the acute toxicity and subchronic toxicity studies showed that H-2-168 and DH-004 had significantly improved safety. In addition, we found that H-2-168 and DH-004 induced DNA damage in E. granulosus s.s., which may be the mechanism by which these drugs produce their therapeutic effects.

Overall, the data from this work demonstrate that H-2-168 and DH-004 are highly effective candidate compounds with low toxicity for the treatment of CE and will provide a new therapeutic strategy for CE pharmacological treatment.