GJB1 gene mutations affecting the gap junction protein Cx32 cause CMT1X, one of the commonest forms of inherited demyelinating peripheral neuropathy. We previously showed that targeted expression of virally delivered Cx32 in Schwann cells following a single intrathecal injection of lentiviral vectors can improve the outcome of the progressive neuropathy that develops after 3 months of age in Cx32 KO mouse model, indicating that gene replacement could offer a therapeutic benefit for this disorder.

The aim of this study was to examine whether late gene replacement therapy after the onset of peripheral neuropathy can provide a therapeutic benefit in the connexin32 (Cx32) knockout (KO) model of X-linked Charcot-Marie-Tooth disease (CMT1X).  

We delivered by intrathecal injection a lentiviral vector LV-Mpz.GJB1 carrying the GJB1/Cx32 gene into 6-month old Cx32 KO mice in order to assess the expression of Cx32 in peripheral tissues. We then proceeded to treatment trial where groups of randomized mice received either the LV-Mpz.GJB1 or the LV-Mpz.Egfp (mock vector). The outcome was examined at the age of 8 and 10 months of age by behavioral, electrophysiological and morphological analysis. We further measured the emerging clinically relevant biomarker neurofilament light (Nfl) levels in blood as an additional outcome measure in treatment groups.

Intrathecal delivery of the therapeutic vector in 6-mo old Cx32 KO mice resulted in expression of Cx32 in lumbar roots and sciatic nerves correctly localized at the paranodal myelin areas. Treatment trial showed better motor performance and increased nerve conduction velocities in treated mice, accompanied by morphological improvement of myelination and reduced inflammation compared to controls. Elevation of blood Nfl levels was also ameliorated in fully treated animals.

Intrathecal gene delivery after the onset of peripheral neuropathy still offers a significant therapeutic benefit in this disease model providing a proof of principle for treating patients with CMT1X.