Glaucoma is the leading cause of irreversible blindness worldwide. In glaucoma, retinal ganglion cells (RGCs), the projection neurons that transmit vision from the retina to the brain, are injured and die, partially as a result of increased intraocular pressure (IOP). Current therapies (laser, surgery and eye drops) all act by lowering IOP. Unfortunately, lowering IOP can produce undesirable side effects and/or be difficult to achieve. Moreover, in some patients, RGC loss continues despite significant IOP reduction. Lacking are "neuroprotective" agents that directly interfere with the cell death process in RGCs. The development of safe and efficacious neuroprotective agents would improve glaucoma therapy by complementing the currently available IOP treatment options. In order to identify such compounds, the Zack group at Johns Hopkins developed a high- content, high-throughput small molecule screen using RGCs. This takes advantage of the ability to isolate and culture primary murine RGCs and then uses automated fluorescent imaging and biochemical assays to screen through small molecule libraries in order to identify compounds that promote RGC survival and neurite outgrowth. Through this screen broad-spectrum protein kinase inhibitors such as Sutent and VX-680 were identified as neuroprotective. Unfortunately the identified compounds are weakly active and extremely toxic. To parse out the individual kinases responsible for cell death and survival, an RNA interference-based approach was utilized. Our collaborators at Johns Hopkins adapted their primary RGC platform for high- throughput siRNA-based screening and used it to screen an arrayed library of siRNAs targeting the mouse kinome. The top two hits identified (i.e. genes whose knockdown promoted survival) were a little explored kinase, DLK and its substrate, MKK7. Independently, Califia Bio has designed and synthesized drug-like inhibitors of this protein kinase target for potential treatment of neuropathy. Potent Califia Bio DLK inhibitors are very active in the Johns Hopkins'RGC protection assay. We have identified a potential lead series from several screening hits from a Califia Bio proprietary scaffold. We present a medicinal chemistry plan and screening cascade to optimize these screening hits into advanced lead compounds with appropriate in vitro PK profiles for direct dosing to the eye to minimize systemic toxicity risk and limited off-target activity. Compounds will be optimized to provide sufficient drug like molecules for preliminary efficacy tests in animals. In phase two, we will optimize compounds for ocular delivery formulations, pharmacokinetic properties, in vivo activity and characterize them in preliminary toxicology and geneotoxicity assays.
Glaucoma is the leading cause of blindness worldwide, and is caused by the death of retinal ganglion cells (RGCs), the nerve cells in the retina that carry vision from the eye to the brain. We are have identified DLK a s a key protein kinase involved in degeneration of RGCs and have identified screening hits which target DLK signaling pathways. We will optimize these molecules into treatments for glaucoma.