Investigating TRPM3 Modulation and Cyst Reversibility in Ex Vivo and Organoid Models of Autosomal Dominant Polycystic Kidney Disease (ADPKD)

Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary kidney disease, affecting approximately 1 in 1000 live births. It is characterised by multiple fluid-filled cysts in renal tubules. ADPKD is associated with mutations in the PKD1 and PKD2 genes, which encode the proteins polycystin 1 (PC1) and polycystin 2 (PC2). PC1 is a transmembrane protein that acts as a mechano-sensor, while PC2 is a non-selective cation channel that belongs to the transient receptor potential (TRP) superfamily. PC1 and PC2 form a heterodimer complex in the primary cilium, crucial for regulating intracellular calcium levels. Recent research suggests that TRPM3, another TRP family member, is necessary for PC2's ciliary function. Therefore, I analysed the impact of TRPM3 activators (nifedipine and CIM0216) and inhibitors (isosakuranetin, primidone, and diclofenac) on cyst formation in cultured E12.5 mouse kidneys which were exposed to various concentrations of forskolin, a compound known to induce cyst formation in kidney cultures. The results indicated that cyst formation occurred at lower forskolin concentrations in kidney rudiments treated with TRPM3 antagonists. In contrast, TRPM3 agonists significantly reduced cyst formation, especially at high forskolin concentrations, compared to kidneys treated with forskolin alone. Among the TRPM3 agonists tested, nifedipine—an FDA-approved antihypertensive drug—showed potential as a therapeutic for ADPKD.
Isosakuranetin was the TRPM3 antagonist that most significantly increased forskolin sensitivity. I generated a drug-induced PKD model by treating mouse kidney rudiments with isosakuranetin and forskolin to analyse cyst reversibility. Cysts in my PKD model originated from proximal tubules and showed disrupted apical-basal polarity. Upon isosakuranetin and forskolin withdrawal, cysts shrank and eventually disappeared with no detectable cell death.
I showed that siRNA-based downregulation was not a useful technique for designing a reversible cyst-forming organoid model, at least in my experimental conditions.
To test whether similar results in cyst reversibility could be achieved in mutant human systems, I developed a CRISPR/Cas13b-based reversible gene downregulation system for future studies. This tool has the potential for the analysis of cyst regression in human organoid studies by inducing cyst formation through controlled downregulation of PKD1 or PKD2 genes, followed by gene re-expression.
Overall, the study suggests that TRPM3 could be a viable target for ADPKD treatment, and nifedipine may be a promising therapeutic option. The findings also provide detailed findings into cyst regression in an early developmental stage. However, it should be noted that all results obtained in these studies were from ex vivo mouse kidney rudiments. To obtain more reliable results, the data here should be confirmed in human ADPKD models. In this context, nifedipine can be tested in ADPKD patient-derived organoids or PKD1 or PKD2 mutant human organoid models.