Inflammation is par for the course in ocular gene therapy; preventing it should be the goal because once it develops, treatment can be difficult.
Reviewed by Dr Christine N. Kay.
Low-grade inflammation seems to be associated with baseline disease in inherited retinal dystrophies (IRDs), according to Dr Christine N. Kay, a surgeon at Vitreoretinal Associates, in Gainesville, Florida, United States. It can be argued that patients with IRDs may be predisposed to inflammation. Adeno-associated virus (AAV)-based gene therapy in these patients may trigger severe inflammatory responses that can lead to permanent visual loss in some.
Examples of diseases include zonular instability in cataract surgeries resulting in IOL dislocations in retinitis pigmentosa (RP), presence of cells in the anterior vitreous, cystoid macular oedema in RP and retinal antibodies in serum samples from patients with RP.
A murine study found that inflammation preceded retinal degeneration in CNGB1 mice.1 Dr Kay also cited Phase 1/2 trials of gene therapy for RPE65-associated Leber congenital amaurosis, choroideremia and RPGR-associated X-linked RP, in which ocular inflammation was dose dependent, with patients who received higher doses at higher risk for vitritis, retinitis and choroiditis.
“At … best … gene therapies treat disease and restore/rescue function,” Dr Kay explained. “At worst … [they introduce] a new antigen and create a new problem.”
Among the numerous factors that stimulate inflammation are delivery mode; vector/capsid manufacturing; promoters and transgene expression; surgical considerations; host characteristics; and dose (the most clinically relevant inflammation occurs with doses of 1 x 1011 vg/eyeor higher).5
The immune response varies according to the mode of administration (subretinal versus intravitreal). Most trials today evaluate vectors injected subretinally, whereas suprachoroidal injections are used less frequently.
“Intravitreal delivery … is [generally] more inflammatory than subretinal delivery and can lead to [increases in] aqueous vector levels, systemic biodistribution to spleen/lymph nodes, [and] systemic dissemination from … anterior chamber [outflow],” Dr Kay said. This delivery, in turn, can trigger more “neutralising antibody responses, which can interfere with effective transgene expression and lead to greater humoral responses and … reduced therapeutic efficacy.”
Regarding the vector capsid serotype, baseline immunity can be species-dependent in animal testing. Dose-dependent increases in binding and neutralising antibodies after AAV injection and in increased serum levels of binding antibodies appear to be correlated with inflammation. However, injection dose is the most important factor affecting increased serum antibody level.
In terms of the roles that vector design and manufacturing play in inflammation, we need to know more about the effects of the production platform (triple transfection in HEK293 cells and herpes- and baculovirus-based systems), of the empty capsid ratio (which correlates with increased transduction efficiency) and of process residuals and contaminants. (Manufacturing of AAV vectors produces three types of capsid—empty, partial and full; the ratio pinpoints the number of empty capsids.) Nevertheless, as previously noted, it is vector dose that correlates most closely with the inflammatory response.6
As another study reports, strong correlation has been observed between AAV toxicity and cis-regulatory sequences.7 Ubiquitous promoters (retinal pigment epithelial promoters) were more toxic than photoreceptor-specific promoters; the reason may be that ubiquitous promoters drive higher levels of transgene expression in more cell types.
Surgical delivery of AAV also plays a role in inflammation. Pro-inflammatory factors of surgery include variability in bleb size, location and dilution with intraocular irrigation (BSS); surgery-related adverse events (particularly haemorrhage); operative duration; and vitreous egress of the vector.
Finally, host (recipient) characteristics include patient age, pigmentation (of the iris), immune system hyperactivity, human leukocyte antigen type and neutralising antibodies. AAV can activate innate pattern recognition receptors that lead to the release of inflammatory cytokines and type I interferons.
A delayed (adaptive) immune response occurs days or weeks after innate immunity. A cellular response can result in increased T cells and a humoral response in increased B cells, plasma cells and antibodies.
In the battle between AAV and the immune system, these are the important points to remember:
Because “our primary responsibility as physicians is to do no harm … we need to acknowledge inflammation in AAV-based gene therapy … discuss it, and learn … to control it,” Dr Kay concluded.
Christine N. Kay, MD
This article was adapted from Dr Christine Kay’s presentation at the 25th annual meeting of the American Society of Gene & Cell Therapy in May 2022. Dr Kay is an investigator for and consultant to numerous companies involved in retinal gene therapy research.