Extrahepatic gene therapy for kidney diseases via polymeric mesoscale nanoparticles

Introduction: The last 5-10 years have seen a rapid expansion of the clinical utility of gene therapies. Many of these are focused on liver diseases, with some exceptions including vaccines and ocular therapies. This is in large part due to the relative abundance of gene therapy carriers that target the liver and other accessible sites, which primarily include lipid nanoparticles, viral delivery mechanisms, and N-acetylgalactosamine conjugates. More recently, substantial pre-clinical efforts have gone into the discovery of novel systemically-administered extrahepatic gene delivery tools, with some notable successes. However, there has been very limited success in the development of kidney-targeted gene therapy carriers to treat renal diseases. Here, we focused on the development of a non-viral gene delivery tool composed of FDA-approved polymeric materials to treat chronic kidney diseases.

Materials and

Methods: We optimized the formulation of kidney-targeted polymeric mesoscale nanoparticles (MNPs) to maximize the encapsulation of siRNA and mRNA gene therapies for kidney diseases. Specifically, we designed a two-step mixture-nanoprecipitation formulation to design MNPs composed of poly-lactic-co-glycolic polyethylene glycol di-block (PLGA-PEG) polymer. This process was separately optimized for siRNA and mRNA loading. Several MNP formulations loading a luciferase reporter mRNA were evaluated for their ability to target the kidneys and express the target protein in hairless immunocompetent mice upon systemic administration using in vivo whole-animal bioluminescence imaging. Similarly, several MNP formulations loaded with siRNAs that target inflammatory cytokines (IL-6, IL-1B, TNFa) were evaluated in both mouse and rat models of chronic kidney disease. Specifically, we investigated the therapeutic efficacy on kidney function and inflammation in a mouse model of unilateral uretral obstruction (UUO) and a rat model of Dahl Salt-Sensitive (DSS) salt-induced hypertension. We evaluated blood and urine markers of renal function in both models, as well as performed a histological assessment of renal inflammation.

Results, Conclusions, and Discussions: We optimized mesoscale nanoparticles for maximal siRNA and mRNA therapy loading while maintaining a 300 – 400 nm polymeric MNP diameter. In healthy immunocompetent hairless mice, we found that MNPs specifically localize to the kidneys and deliver functional luciferase reporter mRNA via in vivo bioluminescence imaging. In both mouse and rat chronic kidney disease models (UUO and DSS, respectively), we administered inflammatory cytokine-silencing siRNA-loaded MNPs. These studies demonstrated substantial therapeutic efficacy and reduction of inflammation from kidney-targeted siRNA delivery as measured by renal fibrosis and injury markers. MNPs localize to the kidneys with up to 26-fold specificity compared to other organs, therefore there is little to no off-target delivery or toxicity. We anticipate further pre-clinical development of both mRNA and siRNA-targeted delivery to the kidneys for chronic kidney diseases will result in a highly specific novel therapy with minimal off-target effects, a first-in-class indication for extrahepatic gene delivery.

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