Development of Conjugated Oligonucleotide Therapeutics for Neuronal Disease

PROJECT SUMMARYDTx technology leverages lipidationthe covalent conjugation of long chain fatty acidsLCFAto oligonucleotide therapeuticsto enable delivery of siRNA into cells and tissues outside of the hepatocyteA lipid motif has been identified that enables the activity of siRNA in vivofollowing intravitreal injection to the eyeand across multiple primary cells including neuronsDTx lipidated siRNA is highly efficacious and at least an order of magnitude more potent at repressing mRNA expression when compared to conjugates of other fatty acidse gDHAthat have been utilized to facilitate siRNA uptake into neurons in vivo or in vitroIn this applicationDTx proposes to evaluate the potential of this novel technology to deliver siRNA and or antisenseASto neurons in vivo to understand broadlyif the technology is useful for neurodegenerative diseases and more specificallyif it can be utilized to prevent neuronal degeneration in a rat model of retinitis pigmentosaRPThe first aimSAwill explore whether DTx technology can enable the delivery of AS molecules into neurons as effectively as it enables siRNAWhile applications of AS and siRNA technology to repress mRNA expression overlap to some degreethere are situationsdue to their distinct modes of actionwhere utilizing one modality over the other can be highly advantageousIn this aiman ASpreviously demonstrated to have in vivo activitywill be conjugated to the DTx lipid motifIts ability to repress mRNA expression both ex vivo in primary neurons and in vivo in retinal neurons will be evaluated through a combination of qPCR and quantitative in situ hybridizationq ISHSAwill evaluate the potential of DTx conjugated siRNA to repress mRNA expression following intracerebroventricularICVinjection to the brainIts ability to repress mRNA expression in neurons will be evaluated by qPCR and qISH relative to DHA conjugated siRNAan approach demonstrated to enable siRNA activity following ICV injection to the mouse brainThe final aimSAwill evaluate whether DTx technology can be applied to siRNA to prevent neuronal degeneration in a rat model of RP driven by a mutationPHin the rhodopsin geneAn siRNA that potently and selectively targets the PH mutant allele will be conjugated to DTx technology and evaluated in vivo in the PH rat RP model for the ability to selectively repress PH expression to prevent photoreceptor cell death and to preserve photoreceptor functionIn this applicationwe lay out a path to better understand our platform technology for neuronal indications in the eye and CNSThe long duration of action of siRNA AS therapeuticsand the inherent safety advantages of keeping a drug confined to a single compartmentmakes local delivery an attractive and commercially viable approach should we succeed in enabling siRNA and improving AS activity in the eye and brainWe expect data from the proposed studies to guide future grant phaseSBIR applications aimed at developing therapeutics for neurodegenerative diseases in the CNS such as Huntington sAlzheimer s and Parkinson s disease and provide the data and a compelling rationale for a phaseSBIR aimed at moving DTx conjugated PH AS siRNA into early clinical development!!NARRATIVEThe biggest hurdle limiting the expansion of oligonucleotides as a therapeutic class is deliveryfinding safe and effective ways to get antisenseASand siRNA therapeutics into cells outside of the liverDTx has developed a technologybased on the conjugation of long chain fatty acidsLCFAsto oligonucleotidesthat enables the delivery of siRNA in vivo and to primary human endothelial cellshuman skeletal muscle cellshuman adipocyteshuman T cellshuman trabecular meshwork cells and primary rat neuronsIn this applicationwe propose to explore whether this technology has the potential to treat neurodegenerative diseases such as retinitis pigmentosaAlzheimer s diseasefrontotemporal dementiaHuntington s disease and or progressive supranuclear palsy!!