Low-Dose Fluvoxamine Modulates Endocytic Trafficking of SARS-CoV-2 Spike Protein: A Potential Mechanism for Anti-COVID-19 Protection by Antidepressants

Abstract

Commonly prescribed antidepressants may be associated with protection against severe COVID-19. The mechanism of their action in this context, however, remains unknown. Here, I investigated the effect of an antidepressant drug fluvoxamine on membrane trafficking of the SARS-CoV-2 spike protein and its cell host receptor ACE2 in HEK293T cells. A sub-therapeutic concentration (80 nM) of fluvoxamine rapidly upregulated fluid-phase endocytosis, resulting in enhanced accumulation of the spike-ACE2 complex in enlarged early endosomes. Diversion of endosomal trafficking provides a simple cell biological mechanism consistent with the protective effect of antidepressants against COVID-19, highlighting their therapeutic and prophylactic potential.

Keywords: COVID-19; SARS-CoV-2; antidepressants; drug repurposing; endocytosis.

FIGURE 1

Fluvoxamine modulates endocytosis, including that of SARS-CoV-2 spike protein. (A), HEK293T cells were allowed to internalise FM dye for 1 h. Representative images of cells incubated with vehicle or 80 nM fluvoxamine. Arrows denote puncta of internalised label. (B), Quantification of FM dye uptake experiments for various concentrations of fluvoxamine. ***p = 0.0001, Kruskal-Wallis test. ****p < 0.0001, *p < 0.05, Dunn’s multiple comparisons test. (C), HEK293T cells were allowed to internalise FluoroRuby for 1 h. Representative images of cells incubated with vehicle or 80 nM fluvoxamine. Arrows denote puncta of internalised label. (D), Quantification of FluoroRuby uptake experiments for various concentrations of fluvoxamine. **p < 0.01, 1-way ANOVA; ***p < 0.001, *p < 0.05, Holm-Sidak’s multiple comparisons test. (E), HEK293T cells were transfected with a plasmid expressing human ACE2, incubated with the recombinant SARS-CoV-2 spike protein for 1 h and sequentially immunostained for surface (cyan) and internalised (magenta) spike protein. Representative images of cells incubated with vehicle or 80 nM Fluvoxamine. Arrows denote apparent puncta of internalised spike protein. (F), Quantification of the spike protein uptake assay. ****p < 0.0001, unpaired two-tailed t test. (G), Quantification of surface ACE2 (live-labelled by spike protein) vs. total ACE2 labelling following 1 h treatment with 80 nM fluvoxamine. p > 0.05, Mann Whitney test. (H), Accumulation of Spike in EEA1-positive early endosomes following 1 h treatment with vehicle or 80 nM fluvoxamine. Arrows denote apparent puncta of internalised spike protein colocalising with EEA1 puncta. (I), Proposed model for modulation of SARS-CoV-2 endocytic trafficking by fluvoxamine. Fluvoxamine promotes accumulation of SARS-CoV-2 into early/recycling endosomes (EE/RE), thereby diverting it away from cell entry through the plasma membrane or late endosomes/lysosomes (LE/Lyso). Scale bar, 10 μm.