Characterization of Gene Therapy Associated Uveitis Following Intravitreal Adeno-Associated Virus Injection in Mice

Abstract

Purpose To characterize the intraocular immune cell infiltrate induced by intravitreal adeno-associated virus (AAV) gene therapy. Methods AAV vectors carrying plasmids expressing green fluorescent protein under the control of PR2.1 were injected intravitreally into AAV naive and AAV primed C57Bl/6 mice. Clinical inflammation was assessed using optical coherence tomography. Intraocular immune cell populations were identified and quantified by flow cytometry on days 1, 7, and 29 after intravitreal injection and compared with sham and fellow eye controls. Results Optical coherence tomography inflammation score and total CD45+ cell number were significantly higher in AAV injected eyes compared to uninjected fellow eye and sham injected controls. Clinically apparent inflammation (vitritis on optical coherence tomography) and cellular inflammation (CD45+ cell number) was significantly increased in AAV injected eyes and peaked around day 7. Vitritis resolved by day 29, but cellular inflammation persisted through day 29. On day 1, neutrophils and activated monocytes were the dominant cell populations in all AAV injected eyes. On day 7, eyes of AAV exposed animals had significantly more dendritic cells and T cells than eyes of AAV naive animals. By day 29, CD8– T cells were the dominant CD45+ cell population in AAV injected eyes. Conclusions Intravitreal AAV injection in mice generates clinically evident inflammation that is mild and seems to resolve spontaneously. However, the total number of intraocular CD45+ cells, particularly T cells, remain elevated. Both innate and adaptive immune cells respond to intravitreal AAV regardless of prior immune status, but the adaptive response is delayed in AAV naive eyes.

Subretinal Therapy: Technological Solutions to Surgical and Immunological Challenges

“…Following an intravitreal injection, a dose dependent immune response is observed presenting as cells in either the anterior chamber and/or the vitreous. Both cellular and humoral factors have been implicated ( 45 – 47 ). In mice, an AAV injection into the vitreous leads to a transient mild spontaneously resolving inflammation but the total number of CD45+ T cells remains elevated, even weeks after the injection.…”

“…In mice, an AAV injection into the vitreous leads to a transient mild spontaneously resolving inflammation but the total number of CD45+ T cells remains elevated, even weeks after the injection. Both innate and adaptive immunity play a role regardless of prior immune status ( 45 ). A patient treated for Lebers with a single intravitreal dose of 9 × 10 10 vg and high pre-treatment titers of IgG neutralizing antibodies to AAV2, developed a significant post-injection ocular inflammation ( 46 ).…”

“…While improved pre-injection screening may reduce the incidence of such events, they limit the applicability of this approach. The presence of inflammation also will limit the efficiency of transfection or the duration of the effect if it leads to the elimination of transfected cells ( 45 , 49 , 50 ).…”

Advances in Ophthalmic Optogenetics: Approaches and Applications

“…One general concern for AAV vectors is that pre-existing immunity and subsequent induced adaptive immunity following vector administration can significantly reduce retinal gene expression, as shown in pre-clinical studies [ 82 ]. Furthermore, there is growing appreciation for the risk of gene therapy associated uveitis following AAV administration, which appears to be related to the vector dose and route of administration [ 83 , 84 ]. However, optogenetic clinical trials thus far have demonstrated that intravitreal delivery of AAV-based vectors is mostly safe and well-tolerated [ 8 ].…”

Therapeutic Applications of Adeno-Associated Virus (AAV) Gene Transfer of HLA-G in the Eye

“…Although there are several reports linking AAV ocular gene therapy to an immune/inflammatory response, especially at high doses, no adverse effects were reported following AAV-HLA-G subconjunctival, intracorneal, or intravitreal dosing [ 14 , 15 , 16 , 42 , 55 , 56 , 57 , 58 ]. Another concern for ocular gene therapy is the durability of the transgene expression following ocular delivery.…”