C-terminal gating modulator for Cav1.4 L-type channels

In this project we will elaborate on therapeutic options to treat a retinal disease, named congenital stationary night blindness type 2 (CSNB2) which is caused by mutations in so-called Cav1.4 L-type calcium channels. These calcium channels are integral proteins at the synaptic side of retinal photoreceptors where they couple light signals to neurotransmitter release. After further processing in the retina the visual information is finally transferred to the brain. Cav1.4 L-type calcium channels are mostly open and hardly close and thereby allow continuous calcium influx important to maintain sustained neurotransmitter release at retinal synapses. Actually, closing of these channels which usually occurs after calcium entry is actively suppressed by an inhibitory domain in the C-terminus, which forms the end of the protein. Patients with mutations in CANA1F, the gene encoding Cav1.4 L-type calcium channel, show typical CSNB2 features, as such photophobia, reduced visual acuity, strabismus, nystagmus and variable levels of night blindness. Currently there is no drug available to cure or at least alleviate the disease. CSNB2 variants which result in a truncation of the C-terminus of Cav1.4 L-type calcium channels are of particular interest for our research. These mutations introduce closing of the channels, a process called inactivation. This inactivation of Cav1.4 L-type calcium channel is likely to reduce the calcium influx at the synaptic side in the retina. Changes in calcium influx alter neurotransmitter release followed by impaired processing of the visual input explaining the visual symptoms particularly the name giving night blindness. When we expressed truncated Cav1.4 L-type calcium channels previously identified in CSNB2 patients in host cells we have shown that we can rescue the channel function when we deliver the missing part of the C-terminus as a separate protein. This also means that the opening and closing process called gating - of a calcium channel can be modulated. Whether or not this concept also works in-vivo has first to be proven in a mouse model that mimics a human truncation mutation. In this project we will therefore apply both viral and non- viral vector-based methods to directly deliver the coding DNA sequence of the distal Cav1.4 C-terminus to the retina of mutant mice. We will (sub-)retinally inject adeno-associated viral vectors as these are considered the current gold standard tool for retinal gene transfer. In the latter approach, we will establish specialized lipid-based nanocarriers which are administrated into the eye for the delivery of DNA. In case we succeed with our manipulation of channel gating after successful expression of the Cav1.4 C-terminus in the mouse retina this would provide a novel therapeutic strategy not only in the retina but also beyond for clinical indications in cardiovascular disease and/or neuropathic pain.