Inhibiting Corona virus-RNA translation with existing antibiotics and other small molecules (Hämning av Corona virus-mRNA translation med befintliga antibiotika och andra små molekyler)

Recent reports claim that  ribosomal RNA-binding antibiotics e.g. anisomycin, doxycyline, etc. are effective against Coronavirus (CoV), but systematic study and molecular insights are lacking.This project aims at identifying and validating existing antibiotics and other small molecules for specific inhibition of SARS-CoV-2 RNA translation. Since the CoV-2 structural gene translation requires -1 frameshift by formation of a sequence-specific RNA 'pseudoknot', it can be inhibited without affecting the host mRNA translation.Specific binders of CoV-2 RNA will be rapidly screened using an ´antibiotic & small molecule microarray´, using a fluorescence-labelled CoV-2 RNA pseudoknot. Collaboration with Matthew Disney, Scripps Research Inst. Already started. ~1 monthThe hits from the microarray will be tested in a rabbit reticulocyte lysate based translation assay using firefly and renilla luciferase dual reporter system, constructed by replacing ORF-1 and -2 of CoV-2 RNA. The constructs are ready in Sanyal lab and the assay works. 2 monthsThe positive hits from the cell-free assay will be validated in human HEK293(T) or similar cells using the luciferase dual reporter assay. Collaboration with Lars Hellman, Uppsala Univ., the expression vectors are present in Hellman lab. 2 - 3 monthsThe results will be quickly published so that the positive hits can be tested in COVID-19 models. Collaboration with the clinicians will be developed for implementing the hits for COVID-19 therapy. Biokemi och molekylärbiologi; Biofysik; Cellbiologi