Recombinant polyclonal proteins targeting COVID-19 and methods of use thereof

A pharmaceutical composition comprising:

• a mixture of at least 100 unique human recombinant antibodies specifically binding to SARS-CoV-2 receptor binding domain (RBD) or SARS-CoV-2 spike S1, wherein the mixture has an EC50 lower than 0.001 mg/ml,
• a pharmaceutically acceptable excipient, and
• a compound selected from benzalkonium chloride, benzyl alcohol, benzoic acid, salicylic acid, thimerosal, phenethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid and hydrogen peroxide in an amount effective to preserve the pharmaceutical composition,
• wherein the mixture of at least 100 unique human recombinant antibodies has been generated by the process of
• a. isolating single cells from a blood sample from a human donor exposed to an antigen of SARS CoV-2;
• b. amplifying polynucleotides, wherein each of the polynucleotides encodes a cognate pair of heavy chain and light chain variable regions from one of the single cells by overlap extension reverse transcriptase polymerase chain reaction (OE-RT-PCR);
• c. cloning the polynucleotides obtained from the amplification into expression vectors, thereby obtaining constructs encoding antibody fragments;
• d. expressing the antibody fragments from the constructs;
• e. enriching a subset of the constructs based on the binding activities of the antibody fragments against SAR-CoV-2 RBD or SAR-CoV-2 spike S1;
• f. generating antibody expression constructs using the subset of the constructs of step e, wherein each of the antibody expression constructs encodes a light chain variable region, a kappa or lambda-type light chain constant region, a heavy chain variable region, and a heavy chain IgG constant region,
• g. introducing the antibody expression constructs of step f into a cell line, and
• h. expressing antibodies from the antibody expression constructs in the cell line, thereby obtaining the at least 100 unique human recombinant antibodies;
• wherein each of the at least 100 unique human recombinant antibodies is an IgG antibody comprising a cognate pair of heavy chain and light chain variable regions;
• and
• wherein each of the cognate pair of heavy chain and light chain variable regions comprises a heavy chain variable region and a light chain variable region from a single cell out of the blood sample.