Understanding the effect of ivermectin in non-severe COVID-19 patients through transcriptomics analyses

Abstract

Abstract Motivation: Ivermectin inhibits SARS-CoV-2 replication in vitro at concentrations 35 times higher than approved clinical dosages for humans. This finding prompted numerous randomized clinical trials to assess its effectiveness. In a clinical trial conducted on non-severe COVID-19 patients to assess the efficacy of ivermectin reducing the transmission of SARS-CoV-2 trial, the ivermectin-treated group exhibited 30% less cough and 50% less anosmia/hyposmia, as well as trends indicating lower viral loads and IgG titers compared to the placebo group. The present study was conducted within the framework of this clinical trial to identify the immunomodulatory mechanisms of early ivermectin treatment during COVID-19, and to elucidate the molecular pathways activated during initial SARS-CoV-2 infection as well as their implications for the immune response to COVID-19. Methods: In the clinical trial, 24 COVID-19 patients were randomly assigned in a 1:1 ratio to ivermectin (400 μg/kg) single oral dose or placebo. Peripheral blood was collected from these patients on days 1 (< 72 hours after symptoms onset and first day of treatment), 4 and 7 in PAXGene tubes and RNA extracted for RNA-Seq analyses. Differential Gene Expression (DGE) analysis was performed using DESeq followed by functional analysis using Gene Ontology (GO) framework. In addition, camera method was applied as it adjusts for inter-gene correlation, preventing the overestimation of variability and enabling accurate control of False Discovery Rate (FDR). In this approach, DGE was not performed, and Blood Transcriptional Modules (BTMs) were used for the direct assessment of differences in immune-related gene sets. Results: On study day 1, all patients showed a notable upregulation of genes associated with viral response, some of them previously reported in COVID-19 studies, as well as genes associated with inflammation, innate response, and induction of apoptosis. As the disease progressed, the response became more adaptive. Ivermectin reduced inflammation on day 4. The ivermectin-treated group showed a rather adaptive response, mainly characterized by an upregulation of B and plasma cells-related BTMs on day 4, and an upregulation of many cell cycle-related BTMs, probably of T-cells, on day 7. The placebo group showed a rather innate response, mainly characterized by an upregulation of monocytes, neutrophiles and dendritic cell-related BTMs on day 4, and an upregulation of erythrocyte and leukocyte-related BTMs on day 7. Conclusions: Ivermectin may reduce inflammation and produce a switch to an adaptive immune response, but no clear effect on COVID-19 responses was observed. Current evidence does not strongly support the use of ivermectin for COVID-19 treatment or prevention, but an immunomodulatory effect of ivermectin cannot be discarded. Availability: Code is available at https://github.com/celiatorresvilanova/TFM Contact: celia.torres@estudiants.uvic.cat Supplementary information: Supplementary data encompass the analysis of the duplicates, which is available at the end of this thesis in the ‘Supplementary material’ and a table of differentially expressed genes (DEGs) related to the immune response in the disease progression (Supplementary file 1) available at https://github.com/celiatorresvilanova/TFM