PT - JOURNAL ARTICLE AU - Olga Minenkova AU - Daniela Santapaola AU - Ferdinando Maria Milazzo AU - Anna Maria Anastasi AU - Gianfranco Battistuzzi AU - Caterina Chiapparino AU - Antonio Rosi AU - Giuseppe Gritti AU - Gianmaria Borleri AU - Alessandro Rambaldi AU - Clélia Dental AU - Cécile Viollet AU - Bruno Pagano AU - Laura Salvini AU - Emanuele Marra AU - Laura Luberto AU - Antonio Rossi AU - Anna Riccio AU - Emilio Merlo Pich AU - Maria Gabriella Santoro AU - Rita De Santis TI - Human single-chain antibodies neutralize SARS-CoV-2 variants by engaging an essential epitope of the spike: a new weapon against COVID-19 AID - 10.1101/2021.06.04.447066 DP - 2021 Jan 01 TA - bioRxiv PG - 2021.06.04.447066 4099 - http://biorxiv.org/content/early/2021/06/04/2021.06.04.447066.short 4100 - http://biorxiv.org/content/early/2021/06/04/2021.06.04.447066.full AB - As of June 2021, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains a global emergency and effective therapeutic interventions for the treatment and prevention of coronavirus disease 2019 (COVID-19) are urgently needed. SARS-CoV-2-neutralizing monoclonal antibodies (mAbs) represent a promising approach to COVID-19 therapy. However, the recently described accumulating mutations in the SARS-CoV-2 spike protein are challenging the efficacy of approved and investigational mAbs, whose widespread use is also hampered by their significant costs and possible side effects, including Antibody-Dependent Enhancement (ADE). Here we describe a cluster of SARS-CoV-2 neutralizing human single chain variable fragment antibodies, identified by phage display, sharing a common VH CDR3 sequence. Phage libraries were built by amplifying variable domains of immunoglobulin genes from cDNA derived from lymphocytes of COVID-19 convalescent subjects living in Bergamo, Italy. The scFv76-cluster antibodies (scFv76-cl Abs) exhibit high affinity for the spike receptor binding domain (RBD) of Wuhan strain and emerging variants, leading to inhibition of RBD/human ACE2 interaction. The antigenic epitope recognized by scFv76 was mapped in the receptor binding motif (RBM) of RBD at residues L455, F456, Y473, N487 and Y489. None of these residues has been to date listed among the RBD mutations of SARS-CoV-2 variants of concern (VOCs), suggesting an important role of such epitope in viral infectivity. Treatment with scFv76-cl Abs is effective against SARS-CoV-2, as determined by in vitro experiments of viral infection, replication, cytopathogenicity and spike-mediated syncytia formation. Moreover, their intranasal administration is shown to counteract infection in two independent animal models. Overall, the biochemical and biological characteristics of scFv76-cl Abs are compatible with their clinical use for COVID-19 therapy by intranasal or aerosol administration. To our knowledge, this is the first example of promising human anti-SARS-CoV-2 scFv antibodies as drug candidates for COVID-19 therapy.Competing Interest StatementOM, EMP and RDS are employees of Alfasigma SpA and are named as inventors on a patent application on the name of the same Company. Patent application is related to scFv76-cluster antibody sequences and to the use of scFv antibodies encoded by such sequences as drugs for prevention and treatment of COVID-19.