Autosomal Dominant Polycystic Kidney Disease (ADPKD) Cell Models for Research

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the most common inherited kidney disorder, affecting approximately 1 in 1000 to 1 in 2500 individuals worldwide. It accounts for about 5-10% of end-stage renal disease (ESRD) cases, with a global prevalence of around 12.5 million cases. The disease is characterized by the progressive development of fluid-filled cysts in the kidneys, leading to renal enlargement and eventual loss of function. The median age of ESRD onset is 54-58 years, and it is the fourth leading cause of ESRD in adults. According to the World Health Organization (WHO), ADPKD contributes significantly to the global burden of chronic kidney disease, with substantial morbidity and mortality. There is no cure, and current treatments focus on managing symptoms and slowing disease progression. The 5-year survival for patients with ESRD due to ADPKD is approximately 50-60%, depending on access to renal replacement therapy. Key risk factors for rapid progression include early diagnosis, hypertension, and a family history of ESRD. The disease also has systemic manifestations, including hepatic cysts, intracranial aneurysms, and cardiac valve abnormalities, which contribute to its clinical complexity.

ADPKD is an ideal model for studying fundamental mechanisms of cystogenesis, epithelial cell proliferation, and fluid secretion. The disease is monogenic in most cases, with mutations in PKD1 or PKD2 accounting for ~85% and ~15% of cases, respectively. This genetic simplicity allows for precise modeling using gene editing. Public datasets, such as those from the National Cancer Institute (NCI) and the Genotype-Tissue Expression (GTEx) project, provide extensive transcriptomic and proteomic data on kidney tissues, enabling researchers to investigate disease pathways. Open questions include the role of ciliary signaling, metabolic reprogramming, and the interplay between genetic and environmental factors. The availability of well-characterized cell lines and organoid models, combined with CRISPR technology, makes ADPKD a tractable system for drug discovery and functional genomics.

Although ADPKD is not a cancer, it shares several proliferative and signaling pathways with neoplasia. The major pathways involved in cystogenesis include:

• • cAMP/PKA Pathway: Elevated cAMP levels stimulate protein kinase A (PKA), leading to increased cell proliferation and fluid secretion. This pathway is a key target for therapeutic intervention.
• • mTOR Pathway: The mammalian target of rapamycin (mTOR) is hyperactivated in ADPKD, promoting cell growth and cyst expansion. Mutations in PKD1 or PKD2 lead to dysregulation of mTOR signaling.
• • Wnt/β-catenin Pathway: Aberrant Wnt signaling contributes to abnormal cell proliferation and differentiation in cystic epithelial cells.
• • MAPK/ERK Pathway: The mitogen-activated protein kinase (MAPK) cascade is activated in ADPKD, driving cell proliferation and survival.

These pathways are interconnected and contribute to the progressive nature of the disease.

Data from ClinVar and COSMIC databases.

The key signaling networks deregulated in ADPKD include:

• • cAMP/PKA: Central to cystogenesis, with elevated cAMP levels leading to increased proliferation and chloride-driven fluid secretion.
• • mTOR: Hyperactivation of mTORC1 promotes cell growth and cyst expansion.
• • Wnt/β-catenin: Aberrant activation leads to increased cell proliferation and dedifferentiation.
• • MAPK/ERK: Activation of this cascade enhances cell proliferation and survival.
• • Calcium Signaling: Loss of polycystin-2 function reduces intracellular calcium, affecting multiple downstream pathways.

Key nodes in these networks include:

• • PKA: Regulates CFTR and other ion channels.
• • mTORC1: Phosphorylates S6K and 4E-BP1.
• • β-catenin: Translocates to the nucleus and activates TCF/LEF transcription factors.
• • ERK1/2: Phosphorylates transcription factors like Elk-1.

These pathways provide multiple targets for therapeutic intervention.

Organoids derived from patient iPSCs or adult stem cells recapitulate cyst formation and are valuable for drug screening. They can be gene-edited to introduce or correct mutations.

Animal models for ADPKD include:

• • Genetically Engineered Mouse Models (GEMMs): Pkd1 and Pkd2 knockout or conditional knockout mice. Examples include Pkd1fl/fl; Cre mice and Pkd2WS25/- mice.
• • Inducible Models: Tamoxifen-inducible Cre recombinase allows temporal control of gene deletion.
• • Rat Models: Pkd1 mutant rats (e.g., Han:SPRD) develop progressive cystic disease.
• • Zebrafish Models: Used for studying ciliary function and early development.
• • Patient-Derived Xenografts (PDX): Not commonly used for ADPKD due to the non-malignant nature, but kidney organoids can be transplanted into mice for in vivo studies.

CRISPR-based gene editing enables the creation of isogenic cell lines with precise mutations in PKD1 or PKD2. These models are essential for studying genotype-phenotype correlations and for drug discovery. Examples include:

• • PKD1 Knockout Cell Lines: Generated by introducing frameshift mutations via CRISPR-Cas9, resulting in loss of polycystin-1. These cells exhibit increased proliferation and altered cAMP signaling.
• • PKD2 Knockout Cell Lines: Similar approach for polycystin-2, leading to disrupted calcium signaling.
• • Knock-in Models: Introduction of specific patient mutations (e.g., R4227X in PKD1) to study their functional impact.

These gene-edited models are commercially available from various sources, ensuring sequence verification and quality control. They accelerate research by providing consistent and reproducible models for high-throughput screening and functional studies.

Gene-edited cell lines are used to validate the function of genes implicated in ADPKD. For example, knocking out PKD1 in renal epithelial cells allows researchers to study downstream effects on signaling pathways and gene expression. Similarly, introducing specific mutations can help determine their pathogenicity. These models are also used in CRISPR screens to identify modifiers of cystogenesis.

Isogenic pairs (wild-type vs. knockout) are ideal for drug screening, as they allow the identification of compounds that specifically target the mutant phenotype. For instance, screening for compounds that inhibit proliferation in PKD1 knockout cells but not in wild-type cells can reveal potential therapeutics. Additionally, gene-edited models can be used to study drug resistance mechanisms, such as the development of resistance to mTOR inhibitors.

CRISPR-based synthetic lethality screens can identify genes that are essential for the survival of ADPKD cells but not normal cells. This approach can uncover novel therapeutic targets and biomarkers. For example, targeting genes involved in metabolic reprogramming may selectively kill cystic cells.