Optimization of humanized C3 rat light damage model for complement therapeutics

Abstract

Purpose : Complement has been implicated in ocular inflammatory and retinal degenerative diseases. In this study, we optimized humanized C3 rat light damage (LD) as a target engagement model to qualify C3 monoclonal antibodies (mAbs).

Methods : C3 humanized rats received intravitreal injection (25ug in 3ul) of either control or anti-C3 mAb one day before light damage. Rats were then dark adapted overnight. On the day of light damage, pupils were dilated with 1% tropicamide. Animals were light exposed (3000 Lux) for 6hrs using Tek Light T5 blue, fluorescent aquarium light bulb 432W. Animals were euthanized 24, 48 or 72 hours post light damage. Posterior eye cup and serum samples were collected for human C3a ELISA (Quidel) and human C3 ELISA (Abcam), whole rat eyes were collected for C3/C3b immunohistochemistry to evaluate complement activation.

Results : In the posterior eye cup (PEC) of C3 humanized rats, the C3 protein activation peaked at 24 hours and gradually decreased day 2 and 3 post blue light damage as measured by C3a ELISA. However, the serum C3a level remained the same from day 1 to 3 after LD. Different from C3a changes, C3 protein levels in PECs were elevated 24 hours post LD and remained increased until day 3 as measured by C3 ELISA and immunolabeling. C3 mAb delivered either intravitreally or subcutaneously was able to reduce the C3 activation in PEC of C3 humanized rats.

Conclusions : LD induces complement local activation in the eye. This is an effective method to induce complement activation. C3 humanized rat is a useful model for testing human C3 mAb candidates.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.