Nanobodies are emerging as critical tools for drug design. Several have been recently created to serve as inhibitors of severe acute respiratory syndrome coronavirus s (SARS-CoV-2) entry in the host cell by targeting surface-exposed spike protein. Here we have established a pipeline that instead targets highly conserved viral proteins made only after viral entry into the host cell when the SARS-CoV-2 RNA-based genome is translated. As proof of principle, we designed nanobodies against the SARS-CoV-2 non-structural protein (Nsp)9, which is required for viral genome replication. One of these anti-Nsp9 nanobodies, 2NSP23, previously characterized using immunoassays and nuclear magnetic resonance spectroscopy for epitope mapping, was expressed and found to block SARS-CoV-2 replication specifically. We next encapsulated 2NSP23 nanobody into lipid nanoparticles (LNPs) as mRNA. We show that this nanobody, hereby referred to as LNP-mRNA-2NSP23, is internalized and translated in cells and suppresses multiple SARS-CoV-2 variants, as seen by qPCR and RNA deep sequencing. These results are corroborated in three-dimensional reconstituted human epithelium kept at air-liquid interface to mimic the outer surface of lung tissue. These observations indicate that LNP-mRNA-2NSP23 is internalized and, after translation, it inhibits viral replication by targeting Nsp9 in living cells. We speculate that LNP-mRNA-2NSP23 may be translated into an innovative strategy to generate novel antiviral drugs highly efficient across coronaviruses.