Intrathecal CLN6 gene therapy has shown limited success in human clinical trials. Increasing evidence suggests that multiple forms of NCL share a common molecular pathway, with CLN5 emerging as a key node in this network. Notably, CLN5 expression is reduced in the brains of CLN6-affected sheep, suggesting a functional CLN5 deficiency in this model. CLN5 is known to interact with several other NCL proteins – including CLN1, CLN2, CLN3, CLN6, CLN7 and CLN8 – raising the possibility that co-delivery of CLN5 might improve the efficacy of CLN6 gene therapy.
To test this, we treated pre-symptomatic CLN6-affected sheep via intracerebroventricular injection with self-complementary adeno-associated virus serotype 9 (AAV9) vectors expressing ovine CLN6 alone, ovine CLN5 alone, or a dual therapy combining both vectors, at a range of doses. The higher dose CLN6 monotherapy and dual therapy markedly improved clinical outcomes, doubling survival to 36 months and significantly delaying disease progression. Neuroimaging showed reduced intracranial volume loss, whilst post-mortem histopathology revealed widespread CNS transduction, and trends toward preserved cortical thickness, and reduced lysosomal storage and neuroinflammation. In contrast, CLN5 monotherapy and lower doses of CLN6 or dual therapy did not confer substantial clinical or neuropathological benefit.
These results highlight the importance of dose, delivery route, and therapeutic synergy in treating NCLs. They support the pursuit of higher-dose CLN6 gene therapy, improved CNS targeting, and exploration of dual-gene strategies as a path forward for CLN6 Batten disease. These approaches may overcome the limitations of previous clinical trials and also more effectively treat other NCL subtypes with shared molecular pathology.