FcRn receptor-mediated pharmacokinetics of therapeutic IgG in the eye

Abstract

Purpose: The goal of this study was to determine the role of the neonatal Fc (FcRn) receptor in eliminating intravitreally administered full-length immunoglobulin G (IgG) across the blood-retinal barrier.

Methods: FcRn receptor expression in normal and laser photocoagulated retinas was compared quantitatively by real-time RT-PCR. The distribution of intravitreally administered full-length IgG was investigated and compared in wild-type and FcRn knockout mouse eyes as well as normal and laser-photocoagulated rat eyes at several time points. Additionally, the pharmacokinetics of intravitreally injected full-length IgG and chicken immunoglobulin Y (IgY) was compared in the normal rat retina.

Results: Intravitreally administered full-length IgG overcame the inner limiting membrane and diffused into the deeper retinal structures in both normal and laser-photocoagulated retinas. Interestingly, IgG was eliminated across the blood-retinal barrier into the blood system in the normal retina, whereas IgY was not. In addition, full-length IgGs did not penetrate across the blood-retinal barrier in the FcRn knockout mouse. Intravitreally injected IgGs were eliminated into the blood system more rapidly in laser-photocoagulated eyes when compared to normal control eyes because of FcRn receptor upregulation in the laser-photocoagulated retina.

Conclusions: FcRn plays an important role in eliminating intravitreally administered full-length IgGs across the blood-retina barrier into the systemic blood system.

Figure 1

The distribution in rat eyes of intravitreally administered fluorescent dye labeled full-length IgG (red). A-C show the distribution of the full-length IgG in the normal retina at 30 min, 6 h, and 15 h post intravitreal administration, respectively. D and E show the distribution of the full-length IgG in the laser-photocoagulated retina at 1 h and 3 h post intravitreal administration, respectively. Intravitreally administered full-length antibody overcame the inner limiting membrane barrier and diffused into the deeper retinal structures in both laser photocoagulated and control rat retinas. A–E: Cell nuclei were stained with DAPI (blue). Abbreviations: inner limiting membrane (ILM), ganglion cell layer (GCL), inner plexiform layer (IPL), inner nuclear layer (INL), outer plexiform layer (OPL), outer nuclear layer (ONL), external limiting membrane (ELM), and Bruch’s membrane (BM).

Figure 2

The distribution in rat eyes of intravitreally administered fluorescent dye labeled full-length IgG (red). A and B show the distribution of the full-length IgG in the live and post-mortem retina 6 h post intravitreal administration, respectively. More antibodies were observed in the post-mortem retina, compared to the live retina, suggesting the presence of active elimination mechanisms for IgG in the live retina. A, B: Cell nuclei were stained with DAPI (blue). Abbreviations: inner limiting membrane (ILM), ganglion cell layer (GCL), inner plexiform layer (IPL), inner nuclear layer (INL), outer plexiform layer (OPL), outer nuclear layer (ONL), and external limiting membrane (ELM).

Figure 3

Correlation between the expression of FcRn receptor and the serum concentration of intravitreally administered bevacizumab. A: Real-time RT–PCR study in rats demonstrated a 1.82-fold upregulation of FcRn mRNA expression in laser-photocoagulated retinas, relative to normal retinas (n=5, p=0.001). B: The serum concentration of bevacizumab in the laser-photocoagulated group (832 ng/ml) was higher than that in the control group (279 ng/ml) 5 h post intravitreal administration of 2 µl bevacizumab at 25 mg/ml (n=4; p<0.05). The error bars are standard error of the mean.

Figure 4

Distribution of intravitreally injected bevacizumab in the FcRn knockout and wild-type mice. A and B show the distribution of intravitreally administered bevacizumab (green) around retinal blood vessels in an FcRn knockout mouse and wild-type mouse 5 h post injection, respectively. C shows the normalized fluorescent intensity across the retinal vascular endothelium in a wild-type mouse. Arrows in B indicate the direction over which the fluorescence profiles were obtained. The normalized fluorescence intensity profile in panel C shows a fluorescence intensity gradient across the retinal vascular endothelium from the abluminal to luminal sides, indicating the penetration of bevacizumab into the blood system in the wild-type mouse. A, B: Cell nuclei were stained with DAPI (blue).

Figure 5

Pharmacokinetics of intravitreally injected bevacizumab and chicken IgY in rats. A–C show the distribution in rat eyes of chicken IgY in the whole retina (A), around the retinal blood vessel (B), and in the choroid (C) 5 h post intravitreal injection as determined by Alexa dye labeled antibodies (red). D–F show the distribution in rat eyes of bevacizumab in the whole retina (D), around the retinal blood vessel (E), and in the choroid (F) 5 h post intravitreal injection as determined by Alexa dye labeled antibodies (red).