Role of Netrin-1 in Polycystic Kidney Disease - PROJECT SUMMARY Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in the polycystic kidney disease 1 (PKD1) gene, leading to the development of numerous fluid-filled cysts originating from tubular epithelial cells. These cystic epithelial cells exhibit a hyperproliferative phenotype, ultimately progressing to end-stage renal disease (ESRD). Tolvaptan, a V2-specific vasopressin receptor antagonist, is the first FDA-approved drug for the treatment of ADPKD. However, additional therapeutic targets are needed because tolvaptan is not curative, and its side effects preclude its use in some PKD patients. A critical barrier to developing more effective and safe treatments for PKD is the incomplete understanding of 1) the underlying mechanisms of renal cyst formation, and 2) how cyst formation contributes to the progression to ESRD. Netrin-1 is a laminin-related secreted molecule that is widely expressed in renal tubular epithelial cells and immune cells. It is highly induced after kidney injury and upregulated in many cancers, where it promotes cancer cell proliferation, invasion, and tumor growth. Our preliminary data, along with published studies, demonstrate that netrin-1 levels are significantly upregulated in both human ADPKD kidneys and rodent models of PKD compared to their control counterparts. Additionally, overexpression of netrin-1 in kidney tubules leads to cyst formation and activation of cellular pathways that mimic human polycystic kidney pathology. Neutralizing netrin-1 with a monoclonal antibody suppressed cyst growth in both male and female PKD mice. These data suggest that the upregulation of netrin- 1 contributes to cyst growth. However, the functional role of netrin-1 and its downstream mechanisms in driving cyst growth and PKD progression is still poorly understood and represents a critical knowledge gap. The goal of this proposal is to address the previously unrecognized role of netrin-1 signaling in cyst growth and PKD progression. We hypothesize that aberrant production of netrin-1 in tubular epithelial cells in ADPKD stimulates uncontrolled proliferation of cystic epithelial cells, which is further exacerbated by netrin-1-induced M2 macrophage polarization to promotes cyst expansion through a UNC5b-dependent pathway. To test this hypothesis, we have formulated two specific aims: Aim 1 will test the hypothesis that tubular upregulation of netrin-1 in ADPKD promotes cell proliferation and cyst growth via activation of the UNC5b/ERK/Akt pathway. Aim 2 will test the hypothesis that secreted netrin-1 drives macrophage polarization in cysts by binding to the UNC5b receptor on macrophages, thereby promoting macrophage-mediated cyst growth. To address these aims, we will use cell-specific netrin-1 and UNC5b receptor conditional knockout PKD mice, transgenic mice with netrin-1 overexpression, and pharmacological treatments (netrin-1 and UNC5b neutralizing antibodies), and in vitro cell culture models will be used to rigorously test our hypotheses. Successful completion of this proposal will provide novel insights into the mechanisms by which netrin-1 induces cyst growth and offer the pre-clinical foundation needed to explore netrin-1 as a novel therapeutic target for ADPKD treatment.
