Rapid Affinity Maturation of Novel Anti-PD-L1 Antibodies by a Fast Drop of the Antigen Concentration and FACS Selection of Yeast Libraries

Table 3

Affinity and rate constants of rhPD-L1-his monovalent binding to anti-PD-L1 IgGs.

mAb	Kinetic analyses^a				Steady-state analyses^b
	ka (1/Ms)^a	kd (1/s)^a	KD (nM)^a	KD fold change	KD1 (nM)^b
Wild type	(5.12 ± 0.72) × 10^5	(1.61 ± 0.30) × 10^−1	314 ± 43.3	1	378 ± 61
3_3	(8.21 ± 0.57) × 10^5	(4.68 ± 0.20) × 10^−2	57.1 ± 4.67	5.49	69.3 ± 3.86
3_7	(7.09 ± 0.69) × 10^5	(6.01 ± 0.06) × 10^−2	84.8 ± 7.71	3.73	101 ± 4.57
3_14	(7.88 ± 0.59) × 10^5	(4.68 ± 0.21) × 10^−2	59.6 ± 5.43	5.26	67.1 ± 5.77
3_17	(7.32 ± 0.33) × 10^5	(3.58 ± 0.20) × 10^−2	49.0 ± 2.21	6.4	61.1 ± 4.98
10_3	(8.35 ± 0.49) × 10^5	(4.37 ± 0.30) × 10^−2	52.3 ± 4.18	6.0	62.9 ± 6.07
10_12	(7.70 ± 0.76) × 10^5	(6.30 ± 0.09) × 10^−2	81.8 ± 7.73	3.83	88.5 ± 7.24

^aKinetic (k_a and k_d) and equilibrium dissociation (K_D) constants for monovalent complexes were calculated by fitting binding curves (Figures 4(a)–4(g)) to the 1 : 1 Langmuir binding model. The equilibrium dissociation constants (K_D) were calculated from the relationship K_D = k_d/k_a. The reported constants are average values obtained from at least four independent analyses using different biosensors, sample preparations, ligand densities and analyte concentration gradients on the flow cell surfaces. Data are reported with standard deviation. K_D fold changes compared to wild type are reported. ^bEquilibrium binding analyses. Equilibrium dissociation constants (K_D1) were calculated by non‐linear curve fitting of 1 : 1 Langmuir binding isotherms by plotting equilibrium response toward analyte concentrations (data not shown). The average values are reported with standard deviation.