designer ds2  (GenScript corporation)

Journal: NPJ Vaccines

Article Title: DS2 designer pre-fusion F vaccine induces strong and protective antibody response against RSV infection

doi: 10.1038/s41541-024-01059-9

Figure Lengend Snippet: A The pre-fusion and post-fusion conformational states of the RSV F glycoprotein are depicted, with colored antigenic sites (Ø, V, IV, III, II, and I) recognized by distinct groups of monoclonal antibodies (mAbs). B Specific mutations introduced to maintain the prefusion conformation in the three designer pre-F glycoproteins DS2, DS-Cav1, and SC-TM. C A schematic diagram illustrates the recombinant AdC68 vector expressing the three designer pre-F glycoproteins of RSV. Each glycoprotein was presented with signal peptide (SP), transmembrane (TM), and cytoplasmic tail (CT) from RSV A2 strain. The coding sequences of each designer pre-F were inserted into the E1 region of the AdC68 vector, controlled of the cytomegalovirus (CMV) promoter, and terminated by a bovine growth hormone (BGH) polyadenylation signal sequence. D A comparison of the expression and epitope display of various designer pre-F on the surface of infected HEK 293T cells. Following infection with recombinant AdC68, HEK 293T cells were analyzed by flow cytometry using a panel of mAbs targeting various epitopes in the pre- and post-F glycoproteins of RSV. The names and sites specificity of each mAb in the F glycoprotein of RSV are indicated. Motavizumab and Palivizumab, two clinically evaluated antibody drugs, recognize the identical epitope within site II. The X-axis indicates the PE fluorescence intensity while Y-axis denotes the number (counts) of the events. The percentage in each of the flow cytometry graph represents the percentage of PE-positive cells.

Article Snippet: Codon-optimized genes encoding the wild-type (WT) fusion glycoprotein (F) of RSV A2 strain (GenBank accession no: ACO83301.1), along with structure-based designer DS2, DS-Cav1, and SC-TM prefusion F glycoproteins – , were synthesized by GenScript Biotech Corporation, China.

Techniques: Recombinant, Plasmid Preparation, Expressing, Sequencing, Comparison, Infection, Flow Cytometry, Fluorescence

Journal: NPJ Vaccines

Article Title: DS2 designer pre-fusion F vaccine induces strong and protective antibody response against RSV infection

doi: 10.1038/s41541-024-01059-9

Figure Lengend Snippet: A Timeline and grouping for vaccination and monitoring of antibody response among five groups of vaccinated mice. Fifty 6-week-old BALB/c mice, ten in each group, were immunized with indicated dose and route of designer vaccines or controls at week 0 and again at week 3. Blood samples were collected every two weeks to monitor the binding and neutralizing titers to RSV pre-F and live RSV, respectively. B , C Comparison of serum neutralizing activity to live RSV and binding activity to each of the designer pre-F and WT F recombinant glycoproteins among the five groups of vaccinated mice (n = 10). Bar graphs depict the week 10 neutralizing titers (ID 50 ) and binding activities (ED 50 ) to recombinant DS2, DS-Cav1, SC-TM, and WT. Each dot represents one animal and color-coded according to the vaccinated group as shown in Fig. . Dotted lines represent assay limit of detection. D Experimental design for evaluating the protective activity of AdC68-DS2 in mice. Ten weeks after immunization with either AdC68-DS2 or AdC68 empty vector, mice were intranasally challenged with 3 × 10 5 PFU of live RSV Long strain. E , F All animals were carefully monitored for body weight changes, for viral loads in the lungs by qPCR and plaque assays, and for pathological analysis of lung tissue by H&E staining (n = 5). The scale bar is 200 μm for 5× and 50 μm for 20×. VL vascular lumen, BL bronchiolar lumen. All data are presented as median ±interquartile range. The Mann–Whitney U test is used for comparisons between two independent groups and Kruskal–Wallis H test for comparisons among multiple independent groups, as the data sets are not uniformly normally distributed. The results shown are representatives of two independent experiments. The p -values are marked in the graphs.

Article Snippet: Codon-optimized genes encoding the wild-type (WT) fusion glycoprotein (F) of RSV A2 strain (GenBank accession no: ACO83301.1), along with structure-based designer DS2, DS-Cav1, and SC-TM prefusion F glycoproteins – , were synthesized by GenScript Biotech Corporation, China.

Techniques: Vaccines, Binding Assay, Comparison, Activity Assay, Recombinant, Plasmid Preparation, Staining, MANN-WHITNEY

Journal: NPJ Vaccines

Article Title: DS2 designer pre-fusion F vaccine induces strong and protective antibody response against RSV infection

doi: 10.1038/s41541-024-01059-9

Figure Lengend Snippet: A Overall experimental procedure for isolating and characterizing DS2-specific mAbs from bone marrow antibody-secreting cells (ASCs). The image was created with BioRender.com, with permission. B Representative bone marrow ASCs secreting DS2-specific antibodies were captured by the surrounding DS2-coated polystyrene particles, detected using a secondary anti-mouse IgG antibody conjugated with Alexa Fluor™ 488, and visualized under a fluorescence microscope. C Phylogenetic analysis of the heavy and light chains of 29 DS2-specific mAbs, and their relative genetic relatedness to control mAbs (D25, CR9501, 101F, MPE8, Motavizumab, and 4D7) targeting six major antigenic sites (Ø, V, IV, III, II and I) in the F glycoprotein of RSV. Nirsevimab, a commercially available antibody drug derived from D25, specifically recognizes the site Ø antigenic site. The epitope specificity for each of the 29 DS2-specific mAbs, defined by the competitive SPR with the six control mAbs, is indicated along with their binding (ED 50 ) and neutralizing (ID 50 ) abilities. The results shown are representatives of two independent experiments. The branch length is drawn to scale to assess genetic relatedness among the mAbs.

Article Snippet: Codon-optimized genes encoding the wild-type (WT) fusion glycoprotein (F) of RSV A2 strain (GenBank accession no: ACO83301.1), along with structure-based designer DS2, DS-Cav1, and SC-TM prefusion F glycoproteins – , were synthesized by GenScript Biotech Corporation, China.

Techniques: Fluorescence, Microscopy, Control, Derivative Assay, Binding Assay

Journal: NPJ Vaccines

Article Title: DS2 designer pre-fusion F vaccine induces strong and protective antibody response against RSV infection

doi: 10.1038/s41541-024-01059-9

Figure Lengend Snippet: Phenotypic and genotypic characterization of the DS2-specific neutralizing mAbs

Article Snippet: Codon-optimized genes encoding the wild-type (WT) fusion glycoprotein (F) of RSV A2 strain (GenBank accession no: ACO83301.1), along with structure-based designer DS2, DS-Cav1, and SC-TM prefusion F glycoproteins – , were synthesized by GenScript Biotech Corporation, China.

Techniques: Sequencing, Control

Journal: NPJ Vaccines

Article Title: DS2 designer pre-fusion F vaccine induces strong and protective antibody response against RSV infection

doi: 10.1038/s41541-024-01059-9

Figure Lengend Snippet: A Lateral and top view of mAb60 Fab in complex with DS2 pre-fusion F trimer. The heavy chain of mAb60 Fab is shown in red, and the light chain in magenta. The DS2 pre-fusion F trimer is depicted in dark gray as a molecular surface, with one of the protomers depicted in green as a ribbon diagram. B Modeling analysis of mAb60 Fab binding to both pre- and post-fusion F monomers. mAb60 binds to the antigenic site II, similar to Motavizumab, but distinct from antibodies to antigenic site Ø, which are only present in the pre-fusion F and are bound by D25 (in teal) and Nirsevimab (in coral). C The epitope and the paratope residues involved in mAb60 Fab binding to the DS2 pre-fusion F. Epitope residues are shown in green and depicted as a molecular surface, while paratope residues are highlighted with yellow circles and represented as ribbon diagrams. The epitope of Motavizumab is outlined by a dark orange dashed line. D Interactions between mAb60 Fab and DS2 pre-fusion F at the binding interface. Hydrogen bonds are shown as pure-blue dotted lines, and salt bridges as light-yellow lines. E Superimpose of mAb60 Fab onto the post-fusion F (PDB: 5J3D).

Article Snippet: Codon-optimized genes encoding the wild-type (WT) fusion glycoprotein (F) of RSV A2 strain (GenBank accession no: ACO83301.1), along with structure-based designer DS2, DS-Cav1, and SC-TM prefusion F glycoproteins – , were synthesized by GenScript Biotech Corporation, China.

Techniques: Binding Assay