Figure 1.
Antigen sweeping by pH-dependent antigen binding antibody with increased FcRn binding at neutral pH.
In vivo study of NPH-IgG1, PH-IgG1 and PH-v1 in normal mice. Effect of antibodies on the total hsIL-6R plasma concentration was evaluated in a co-injection model and a steady-state model. In the co-injection model, hsIL-6R, hsIL-6R+NPH-IgG1, hsIL-6R+PH-IgG1 and hsIL-6R+PH-v1 were intravenously administered as single doses of 50 µg/kg for hsIL-6R and 1 mg/kg for antibody and a time profile of total hsIL-6R plasma concentration (A) is shown. Each data point represents the mean ± s.d. (n = 3 each). In the steady-state model, steady-state plasma concentration of approximately 20 ng/mL hsIL-6R was maintained using an infusion pump filled with hsIL-6R solution, and NPH-IgG1, PH-IgG1 and PH-v1 were intravenously administered as single doses of 1 mg/kg and a time profile of total hsIL-6R plasma concentration (B) is shown. Each data point represents the mean ± s.d. (n = 3 each).
Table 1.
Mutations and FcRn binding affinity of hIgG1 Fc variants.
Figure 2.
In vivo study of sweeping antibodies in a normal mice hsIL-6R trans-signaling model.
Effect of antibodies on the total hsIL-6R plasma concentration and SAA plasma concentration (as a marker for hsIL-6R antagonism) were evaluated. hsIL-6R was intravenously administered as a single dose of 250 µg/kg. At 2 h, non-neutralizing antibodies PHX-IgG1 and PHX-v1 were intravenously administered as single doses of 30 mg/kg (A, B), and neutralizing antibodies NPH-IgG1, PH-IgG1 and PH-v1 were intravenously administered as single doses of 0.03 mg/kg (C, D). At 24 h, hIL-6 was intravenously administered as a single dose of 8 µg/kg. Total hsIL-6R plasma concentration (A, C) and SAA plasma concentration (B, D) at 30 h is shown. Each data represents the mean ± s.d. for total hsIL-6R plasma concentration and the mean ± s.e. for SAA plasma concentration (n = 3–7 each). ND, not detected (below 0.195 ng/mL). Statistical significance was determined by t-test (*) or Tukey’s multiple comparison test (#) for total hsIL-6R and SAA plasma concentration.
Figure 3.
Characterization of sweeping antibody in hFcRn-Tgm.
(A) In vivo study of NPH-IgG1, PH-IgG1, PH-YTE and PH-v2 in hFcRn-Tgm. Effect of antibodies on the total hsIL-6R plasma concentration was evaluated in a co-injection model. hsIL-6R, hsIL-6R+NPH-IgG1, hsIL-6R+PH-IgG1, hsIL-6R+PH-YTE and hsIL-6R+PH-v2 were intravenously administered as single doses of 50 µg/kg for hsIL-6R and 1 mg/kg for antibody and a time profile of total hsIL-6R plasma concentration is shown. Each data point represents the mean ± s.d. (n = 3 each). (B) Effect of pH-dependent antigen binding and increased binding affinity to FcRn at neutral pH on antigen sweeping in hFcRn-Tgm steady-state model with hsIL-6R plasma concentration of approximately 20 ng/mL. NPH-IgG1, NPH-v2, PH-IgG1, PH-v2 and PH-v0 were intravenously administered as single doses of 1 mg/kg. Time profile of total hsIL-6R plasma concentration is shown. Each data point represents the mean ± s.d. (n = 3 each).
Figure 4.
Effect of hFcRn binding affinity at neutral pH on antigen sweeping profile in hFcRn-Tgm.
(A, B) Effect of FcRn binding affinity at pH 7.0 on antigen sweeping and antibody pharmacokinetics in hFcRn-Tgm steady-state model with hsIL-6R concentration of approximately 20 ng/mL in the presence of human IgG. NPH-IgG1, PH-IgG1, PH-v3, v4, v5 and v6 were intravenously administered as single doses of 1 mg/kg with 1 g/kg of hIgG. Time profiles of total hsIL-6R plasma concentration (A) and antibody plasma concentration (B) are shown. Each data point represents the mean ± s.d. (n = 3–6 each).
Figure 5.
Effect of sweeping antibody on high plasma concentration antigen.
Effect of NPH-IgG1, PH-IgG1 and PH-v6 on a hFcRn-Tgm steady-state model with high hsIL-6R concentration of approximately 250 ng/mL in the presence of human IgG. NPH-IgG1, PH-IgG1 and PH-v6 were intravenously administered as multiple doses of 0.01 mg/kg every other day. Molar baseline hsIL-6R concentration (6.6 nM) is 5-fold higher than antibody concentration at 15 min (1.3 nM). Time profiles of total hsIL-6R plasma concentration (A) and free hsIL-6R percentage over control (B) are shown. Each data point represents the mean ± s.d. for total hsIL-6R concentration (n = 3–5 each). Free hsIL-6R percentage over control is determined from the pooled plasma sample of n = 3–5 each.