2026
Research Square 2026 Jul 6. doi:10.21203/rs.3.rs-10146411/v1 preprint
Sustained optogenetic expression in skeletal muscle is limited by host adaptive immunity, but preserved with targeted immunosuppression
UNSW Sydney, Macquarie University, Children's Medical Research Institute, The University of Sydney.
Service type: Stock strains
Abstract
Obstructive sleep apnoea (OSA) is characterised by recurrent upper airway collapse during sleep due to inadequate neuromuscular compensation of the upper airway. Despite its high prevalence, affecting upto one billion people worldwide, existing therapies are often poorly tolerated or variably effective, leaving persistent residual disease and a major unmet clinical need. Optogenetic approaches to muscle stimulation have emerged as a novel strategy to address this need by enabling precise, non-invasive, direct targeting of upper airway neuromuscular dysfunction. Recombinant adeno associated viral (rAAV) vector–mediated expression of light sensitive proteins (opsins) in upper airway muscles enables targeted muscle activation and airway dilation in response to light in preclinical rodent models of upper airway hypotonia and atonia. However, in rat models, opsin expression following rAAV delivery was transient—robust at 3 weeks but significantly diminished by 12 weeks post administration—consistent with known immune related barriers to achieving durable transgene expression and clinical translation of muscle directed rAAV gene therapies. To assess whether immunosuppression mitigated opsin loss, rats received intramuscular rAAV delivery to the tongue and were treated with either systemic prednisolone, a combination of rapamycin and cyclosporin, or vehicle control three times a week from week -1 to week +12. Only combined rapamycin/cyclosporin significantly reduced immune cell infiltration (~ 20% reduction; p≤0.02). Correspondingly, opsin expression was significantly higher in this group (one way ANOVA; p<0.0001), and light stimulation increased muscle activation by approximately two-fold (linear mixed model; p<0.001). In contrast, prednisolone did not differ from the control group, with both cohorts showing little to no response to stimulation (LMM, p>0.05). While the immunological mechanisms underlying opsin loss are not yet fully defined, these findings demonstrate that sustained functional transgene expression in upper airway muscle can be achieved with immunosuppression, supporting optogenetic muscle stimulation as a viable gene therapy–based approach for OSA. More broadly, they underscore the importance of immune management for achieving durable expression in rAAV‑mediated therapies, particularly those involving long‑term expression of transgene products that are non-endogenous or incompletely immune-tolerized.
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