Pharmacokinetics of Monoclonal Antibody and Antibody Fragments in The Mouse Eye Following Intravitreal Administration

Abstract

Mice are rarely used in pharmacokinetic (PK) studies of ocular therapeutics due to the small size of their eyes and challenges in drug administration, tissue collection, and analysis of drug concentrations. Therefore, ocular PK of protein therapeutics in mouse eye following intravitreal (IVT) administration is not known. Here, we have presented the first of its kind investigation, to study the PK of 4 different size non-binding protein therapeutics in mouse plasma, cornea/ICB, vitreous humor, retina, and posterior cup (including choroid) following IVT administration. Administered proteins include trastuzumab (150 kDa) and F(ab)₂ (100 kDa), Fab, and scFv (27 kDa) fragments of trastuzumab. An imaging and injection apparatus suitable for performing small (50 nL) IVT injections in mice was developed, and techniques for enucleation of the eye and dissection of ocular tissues were developed. Furthermore, a sensitive enzyme-linked immunosorbent assay (ELISA) for detection of proteins in very small amounts of ocular tissues were developed. It was observed that elimination from the vitreous chamber was the primary driver of PK in the cornea/ICB, retina, posterior cup, and plasma. Trastuzumab displays first-order kinetics in the vitreous humor with a half-life of 18.8 h. F(ab)₂, Fab, and ScFv show biphasic PK profiles with distribution phases becoming more rapid as molecular weight decreases, and terminal elimination becoming longer as molecular weight decreases, with terminal half-lives of 16.3, 20.6, and 48.9 h, respectively. The mean residence times of trastuzumab, F(ab)₂, Fab, and scFv in the vitreous humor were 26.0, 12.2, 10.7, and 8.16 h, respectively. It was found that the mean residence time in vitreous humor doubles with an increase in molecular weight of ∼69 kDa. Interestingly, the PK of proteins measured in the un-injected eye suggest the presence of a pathway for drug transfer between the eyes, which needs to be further validated. Overall, the findings presented here pave the way for drug discovery and development studies of protein therapeutics for ophthalmic indications in mice.

Keywords: Antibody drug(s); Antibody(s); Macromolecular drug delivery; Ophthalmic drug delivery; Pharmacokinetics; Preclinical pharmacokinetics; Protein delivery; Protein(s); Residence time(s); Site-specific delivery.