Glaucoma and other optic neuropathies are characterized by retinal ganglion cell (RGC) non-regenerative
axon damage and eventual cell death. Optic nerve crush (ONC), which transects all RGC axons, is often used
to model this process, and to seek interventions that preserve RGCs and promote regeneration of their axons.
Following ONC in mice, ~80% of the RGCs die within 2 weeks, and virtually none of the survivors regenerate
axons. We and others have used ONC to identify interventions that lead to increased survival, increased
regeneration or both. However, these treatments are only partially effective and many relevant targets are not
well-suited for the clinic. It is therefore important to identify additional and improved promoters of survival and
regeneration. Kinases are key regulators of cellular responses to insults and stresses. Furthermore, because
of their discrete enzymatic activity, kinases are druggable targets. Thus, we performed a preliminary in vivo
CRISPR-assisted kinase screen paired with ONC in adult mice to determine which kinases prevent RGC
survival and axon regeneration. To our excitement, the first round of screening through 750 kinases identified
17 anti-survival and 5 anti-regenerative hits. Many of these hits represent new mechanistic and translational
insights. In the proposed study, we will first examine the mechanistic underpinnings of two novel target kinase
pathways (namely LKB1/SNRK and Mkk4/Mkk7) that we have already verified the efficacy of with larger
experimental cohorts beyond our preliminary screen. These experiments will allow us to more completely
understand their mechanisms of action to formulate ideal therapeutic interventions. Additionally, we will
complete the validation of our remaining target kinases with full experimental cohorts in the ONC model.
Finally, we will choose the most promising gRNAs with robust neuroprotective effects and test their ability to
protect RGCs in our recently developed glaucoma model. At the completion of these studies, we expect to
be well-positioned to leverage the knowledge generated here into druggable targets to treat multiple
conditions causing blindness.