PROJECT SUMMARY Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the most common, monoallelic nephropathy worldwide, characterized by continuous renal cysts growth leading to end stage kidney disease. A single FDA approved therapy is available, but it merely slows cyst growth and impacts quality of life, highlighting the urgent need for new treatment options. ADPKD presents with high phenotypic variability, suggesting that mechanisms beyond the genetic mutation to either PKD1 or PKD2 influence disease severity. Recent data have implicated immune cells as modulators of disease severity. Multiple publications highlight that M2-like renal macrophages promote cystic progression. Conversely, data from my K01, using the slowly progressive, orthologous Pkd1 p.R3277C (Pkd1RC/RC) ADPKD model, show that CD8+ T cells can inhibit renal cyst growth. We further found that the renal cystic microenvironment displays multiple features of immunosuppression such as increased numbers of CD4+ regulatory T cells, metabolic reprogramming, and engagement of immune checkpoints. Indeed, immune checkpoint inhibitors, which lead to reactivation of CD8+ T cells, alleviate cystic disease in the Pkd1RC/RC model. What remains unknown are the mechanisms that drive immunosuppression, which may present important/novel therapeutic targets. In cancer, a disease that parallels PKD at the cellular and molecular level, activation of the complement pathway has been shown to promote an immunosuppressive microenvironment. The complement cascade is a central part of the innate immune system that regulates adaptive immunity. Complement has been shown to be upregulated in PKD cells, patient kidneys/cyst fluid, and murine models, including the Pkd1RC/RC mouse as shown by our preliminary data. Further, genetic loss or non-selective inhibition of the complement protein C3 slows cyst growth in murine PKD models. We hypothesize that complement signaling drives PKD, in part, by creating an immunosuppressive microenvironment which impairs the adaptive immune system to halt cyst growth. This project has two aims. In Aim 1, we will utilize fluorescent-activated cell sorting, flow cytometry, and multispectral immunofluorescent imaging to determine which cells are key producers and responders of complement signaling in the Pkd1RC/RC kidney. We will further correlate these findings to disease severity. In Aim 2, we will test if targeted complement inhibition alleviates cystic disease. This will be the first of such studies using a clinically relevant, specific complement inhibitor in a model orthologous to ADPKD, the Pkd1RC/RC mouse. We will further utilize sophisticated 64-fluorophore capable flow cytometry methods to investigate how complement inhibition alters the cystic immune microenvironment with specific focus on immunosuppressive features. These investigations directly expand upon my NIDDK K01 work and will provide key preliminary data for a competitive R01 application. Obtaining this R03 will further facilitate my transition into an independent investigator with an innovative, preclinical, and translational research program that focuses on the role of the cystic microenvironment as modulator of and therapeutic target for PKD.

PROJECT NARRATIVE While recent data implicate innate and adaptive immune cells as key modulators of Autosomal Dominant Polycystic Kidney Disease (ADPKD) severity the mechanisms that shape the immune milieu to either support or halt cyst progression are not well-defined. This proposal studies complement, which is known to be activated in PKD and drives immunosuppression as one such possible mechanism. The proposed research will identify the cells that are key producers of and responders to complement signaling and investigate the efficacy of complement inhibition on halting PKD progression and altering the cystic microenvironment using an orthologous model of ADPKD.