Therapeutic potential of urinary extracellular vesicles in delivering functional proteins and modulating gene...

Chronic kidney disease (CKD) is a widespread health concern, impacting approximately 600 million individuals worldwide and marked by a progressive decline in kidney function. A common form of CKD is autosomal dominant polycystic kidney disease (ADPKD), which is the most inherited genetic kidney disease and affects greater than 12.5 million individuals globally. Given that there are over 400 pathogenic PKD1/PKD2 mutations in patients with ADPKD, relying solely on small molecule drugs targeting a single signaling pathway has not been effective in treating ADPKD. Urinary extracellular vesicles (uEVs) are naturally released by cells from the kidneys and the urinary tract, and uEVs isolated from non-disease sources have been reported to carry functional polycystin-1 (PC1) and polycystin-2 (PC2), the respective products of PKD1 and PKD2 genes that are mutated in ADPKD. uEVs from non-disease sources, as a result, have the potential to provide a direct solution to the root of the disease by delivering functional proteins that are mutated in ADPKD. To test our hypothesis, we first isolated uEVs from healthy mice urine and conducted a comprehensive characterization of uEVs. Then, PC1 levels and EV markers CD63 and TSG101 of uEVs were confirmed via ELISA and Western blot. Following characterization of uEVs, the in vitro cellular uptake, inhibition of cyst growth, and gene rescue ability of uEVs were demonstrated in kidney cells. Next, upon administration of uEVs in vivo, uEVs showed bioavailability and accumulation in the kidneys. Lastly, uEV treatment in ADPKD mice (Pkd1^fl/fl;Pax8-rtTA;Tet-O-Cre) showed smaller kidney size, lower cyst index, and enhanced PC1 levels without affecting safety despite repeated treatment. In summary, we demonstrate the potential of uEVs as natural nanoparticles to deliver protein and gene therapies for the treatment of chronic and genetic kidney diseases such as ADPKD.