Ebola virus (EboV) is the etiologic agent of an haemorrhagic fever characterised by a high mortality rate. No therapeutics are available for prevention or treatment of this disease. Recently, it has been demonstrated that viral entry into target cells requires the activity of the acid sphingomyelinase (aSMase), cathepsins and the presence of the intracellular receptor Niemann-Pick C1 (NPC1) protein. The antiarrhythmic drug Amiodarone (Amio) and its main metabolite MDEA are able to perturb the cellular endocytic pathway, with the generation of large vacuoles and mislocalization of cellular proteins, such as NPC1, and lipids. The aim of this work is to evaluate if Amio/MDEA are able to interfere with EboV infection. We evaluated the effect of the compounds on viral entry by confocal microscopy and citofluorimetric analysis of Vero cells incubated with: i) Ebola virus-like particles (EboVLPs) characterised by the VP40 matrix protein fused in frame to the GFP, and ii) Ebola-glycoprotein (GP)-pseudotyped VSV∆G (EboVSV) expressing the GFP in place of its native envelope glycoprotein. In addition, we analysed the Amio effect on key enzymatic activities required for EboV entry. Finally, the effect of the compounds were evaluated on wild-type EboV entry into target cells and viral progeny release under BSL4 condition. Our results demonstrate that Amio is able to inhibit EboVSV replication in a dose-dependent manner. By using the EboVLPs we observed that Amio reduces the binding of VLPs to the cell surface in pre-treated cells, due to the inhibition of the aSMase activity, and their trafficking through the endocytic pathway. Moreover our observations with EboVSV suggest an inhibition of the viral envelope fusion with the endosomal membranes. No effect on cellular cathepsins activity was observed in treated cells. Significantly, the combination of 2 µM Amio/MDEA (average serum concentration found in patients treated for arrhythmia), or 10µM Amio, significantly inhibit wild-type EboV replication in vitro by affecting both the number of infected cells and viral progeny release. Our data suggest that Amio and MDEA inhibit Ebola virus infection by interfering with the entry step. Noteworthy, Amio already exerts its effect at concentrations reached in treated patients.