Impact of differential sex response to the vector when developing IL-33 gene therapy for AMD

Abstract

Purpose : Interleukin-33 promotes metabolic homeostasis, resolution of inflammation and can protect against RPE cell death in an in vivo model of outer retinal degeneration. To demonstrate proof of concept for treating atrophic age-related macular degeneration, we characterise efficacy, expression and safety when delivering secreted IL-33 by adeno-associated virus (AAV).

Methods : Male C57BL6/J mice (n = 3-4) were injected intravitreally with AAV vector expressing secreted murine IL-33 or vector expressing no transgene (2E8 gc/eye). At Four weeks mice were exposed to 50K LUX light for 10 minutes unilaterally. As light injury leads to microglia proliferation and monocyte recruitment, retinas were analysed by flow cytometry at day 28. Cx3cr1^CreER:R26-tdtomato (C57BL/6) heterozygous mice (male/female; n = 3-6) were injected intravitreally with a low, mid or high (2E7, 2E8 or 2E9 gc/eye) dose of AAV.IL-33 and AAV.null in the contralateral eye. At four weeks, retinas were processed for characterisation by flow and supernatant IL-33 expression determined by ELISA.

Results : After light injury, male mice treated with 2E8 gc/mL AAV.IL-33 resolved CD45+CD11b+ cells in the eye to naïve levels, whilst these were significantly increased in AAV.null treated eyes (p = 0.0184). After AAV injection only, CD45hiCD3+ percentages were increased with dose, for both vectors. At the high dose males and females had a significant increase (p < 0.0001), whilst no significance was observed at low dose for vectors or sex. At the mid dose, CD45hiCD3+ cells were significantly increased compared to naïve eyes in females only (AAV.IL-33 p = 0.0198, AAV.null p = 0.0124). There were no differences between vectors. Secreted IL-33 increased with vector dose in males and was significantly increased compared to the null vector at mid (p = 0.0251) and high doses (p = 0.0078). In female mice IL-33 secretion did not increase with vector dose or compared to the null vector.

Conclusions : In male mice, IL-33 gene therapy protects against disease processes after light injury. However, female mice had a greater immune response to AAV vectors compared to males, coinciding with a lack of increased transgene expression levels with dose. Importantly, these sex differences could impact translation for female patients, warranting further investigation to appropriately develop IL-33 and other targets into AAV gene therapies suitable for all patient populations.

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