Anti-hiv antibody 10-1074 variants
An antibody and antigen technology, applied in the direction of antibodies, antibody medical components, antiviral agents, etc., can solve the problem of not fully revealing the detailed composition of serum responses.
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Embodiment 1
[0178] 1 0-1074 Identification and Characterization of Variants of Broadly Neutralizing Antibodies - Round 1
[0179] The first round of variants was generated using transient expression in HEK293 cells, as shown in Table 9, including MS-203, MS-204, MS-205, MS-206, MS-207, MS-208, MS-209, MS -210, MS-211, MS-212, MS-213, MS-214, MS-215, MS-216, MS-217, MS-218, MS-219, MS-220 and MS-224, and by protein A chromatography for purification. The characterization methods used to analyze the variants are listed in Table 8, including Size Exclusion Chromatography (SEC), Differential Scanning Fluorescence (DSF), Low pH Stability, and Relative Solubility Assay (RSA). The antibody buffer was exchanged into phosphate buffered saline and used for analysis. Assays used to analyze the first round of variants included: SEC, to quantify purified monomer and high molecular weight species; DSF, to characterize the stability of CH2 and Fab domains during thermal ramping, and retention of neu...
Embodiment 2
[0184] 10-1074 Identification and Characterization of Variants of Broadly Neutralizing Antibodies—Round 2
[0185]A second round of combinatorial variants was designed based on the first round of variants described in the previous section. The combination variants tested in the second round of optimization are shown in Table 12 and consisted of ten double combinations, ten triple combinations, five quadruple combinations, and one Amino acid modification composition) composition. These variants include MS-200, MS-201, MS-202, MS-225, MS-226, MS-227, MS-228, MS-229, MS-230, MS-231, MS-232, MS- 233, MS-234, MS-235, MS-236, MS-237, MS-238, MS-239, MS-240, MS-241, MS-242, MS-243, MS-244, and MS-245. Combinatorial variants were generated using transient expression in HEK293 cells and purified by protein A chromatography. Antibodies were buffer exchanged into phosphate buffered saline prior to use in the assay. Assays used to analyze the second round of variants included: SEC t...
Embodiment 3
[0212] Characterization of oligomeric species and HMW formation of 10-1074 variants during virus inactivation and purification steps
[0213] Figure 1 shows the anti-HIV antibody 10-1074 variant MS-194 ( Figure 1A ) and MS-203 ( Figure 1B ) for characterization. Arrows indicate peaks in the HP-SEC spectrum corresponding to oligomeric species formed during virus inactivation. figure 2 Comparison of the degree of aggregation represented by high molecular weight ("HMW") and oligomeric species levels after each purification step of the 10-1074 antibody variants MS-194, MS-200, MS-201 and MS-203 is shown. Quantitative.
[0214] The ExpiCHO-S was used essentially as described by ThermoFisher (Cat. No. A29133, Document Part No. A29518) TM The "maximum titer" approach was used to produce molecules MS-194, MS-200, MS-201 and MS-203. Using expifectamine, the pcDNA3.4 expression vector containing the light chain or heavy chain coding region was co-transfected into CHO-S cells wi...
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