Additionally, we would like to thank Alejandra Montes-Luna and Ilselena Corts-Paniagua for their assistance with the in vitro assays

Additionally, we would like to thank Alejandra Montes-Luna and Ilselena Corts-Paniagua for their assistance with the in vitro assays. libraries called ALTHEA Gold Plus Vanoxerine Libraries?. Three out of 24 specific clones for the RBD isolated from the libraries, with affinity in the low nanomolar range and sub-optimal in vitro neutralization in PRNT, were affinity optimized via a method called Rapid Ptgs1 Affinity Maturation (RAM). The final molecules reached sub-nanomolar neutralization potency, slightly superior to IgG-A7, while the developability profile over the parental molecules was improved. These results demonstrate that general-purpose libraries are a valuable source of potent neutralizing antibodies. Importantly, since general-purpose libraries are ready-to-use, it could expedite isolation of antibodies for rapidly evolving viruses such as SARS-CoV-2. Keywords: COVID-19, phage display, therapeutic antibodies, affinity maturation, semi-synthetic libraries 1. Introduction The devastating impact of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on human health and the global economy prompted an unprecedented search for diagnostic, prophylactic and therapeutic options to control this viral infection. Due to the success of antibodies in preventing and treating Vanoxerine diverse infectious diseases [1,2], hundreds of academic laboratories as well as small, medium and large biotech companies around the world focused their research efforts on isolating and characterizing anti-SARS-CoV-2 antibodies. Such efforts led to the Emergency Use Authorization (EUA) by the Food and Drug Administration (FDA) and/or the European Agency of Medicines (EMA) of three cocktails of two antibodies each plus three standalone prophylactic and/or therapeutic anti-COVID-19 antibody-based Vanoxerine drugs [3]. Nevertheless, as SARS-CoV-2 continues to evolve into new variants of concern (VOCs) and immune scape variants [4], EUA antibodies have lost efficacy, resulting in a continuous quest for new diagnostic, prophylactic and/or therapeutic antibodies to Vanoxerine manage COVID-19. The variable (V) regions serving as substrate for engineering the EUA antibodies were obtained from immune repertoires of COVID-19 convalescent and/or infected patients, mostly via B-cell selection and V region cloning, with one EUA antibody, Regdanvimab, isolated from an immune phage display library [5]. This has been in part due to the suggestion [6] that immune repertoires commonly generate more potent neutralizing antibodies than those obtained from na?ve, general-purpose, libraries. However, the latter have advantages over immune repertoires that can be exploited to expedite the isolation of antibodies for rapidly evolving viruses. Among others, libraries built with general-purpose antibody repertoires are ready-to-use, avoiding the search for sources of immune repertoires and library construction, thus shortening the discovery phase of V regions to be used as substrate for engineering antibody-based drugs. More importantly, learning from hundreds of therapeutic antibodies that have failed in preclinical and clinical development, the latest generation of general-purpose synthetic or semisynthetic libraries [7] have been designed to maximize the developability profile of the selected V regions includingbut not limited toexpression, aggregation, solubility and chemical and long-term stability, which translates into molecules that could speedily and robustly be developed in therapeutic drugs. We reported in previous works [8,9] the characterization of highly potent antibodies with broad SARS-CoV-2 neutralizing capacity obtained from an immune scFv phage-displayed library. This library was built with the VH repertoire of a convalescent COVID-19 patient infected with the SARS-CoV-2 Delta (B.1.617.2) variant, who was previously vaccinated with a single dose of Convidecia?. Four synthetic VL libraries were used as counterpart of this patient immune VH repertoire. Two of the VL libraries were built with the IGKV4-01 and IGKV3-20 human germline genes, which were designed and tested as part of ALTHEA Gold Libraries? [10]. The other two VL libraries were designed with the IGKV1-39 and IGKV3-11 human germline genes. These additional VLs increased the structural diversity of Vanoxerine the libraries with the potential of generating a more diverse set of antibodies. After panning the library with the SARS-CoV-2 receptor-binding domain of (RBD) wild-type (WT), also known as the Wuhan variant, we obtained a panel of 34 anti-SARS-CoV-2 scFvs encoded by diverse IGHV germline genes, combined with variants of all the.