Adult granulosa cell tumors (AGCTs) arise from the estrogen-producing granulosa cells. Treatment of recurrence remains a clinical challenge, as systemic anti-hormonal treatment or chemotherapy is only effective in selected patients. We established a method to rapidly screen for drug responses in vitro using direct patient-derived cell lines in order to optimize treatment selection. The response to 11 monotherapies and 12 combination therapies, including chemotherapeutic, anti-hormonal, and targeted agents, were tested in 12 AGCT-patient-derived cell lines and an AGCT cell line (KGN). Drug screens were performed within 3 weeks after tissue collection by measurement of cell viability 72 h after drug application. The potential synergy of drug combinations was assessed. The human maximum drug plasma concentration (Cmax) and steady state (Css) thresholds obtained from available phase I/II clinical trials were used to predict potential toxicity in patients. Patient-derived AGCT cell lines demonstrated resistance to all monotherapies. All cell lines showed synergistic growth inhibition by combination treatment with carboplatin, paclitaxel, and alpelisib at a concentration needed to obtain 50% cell death (IC50) that are below the maximum achievable concentration in patients (IC50 < Cmax). We show that AGCT cell lines can be rapidly established and used for patient-specific in vitro drug testing, which may guide treatment decisions. Combination treatment with carboplatin, paclitaxel, and alpelisib was consistently effective in AGCT cell lines and should be further studied as a potential effective combination for AGCT treatment in patients.

Nephropathic cystinosis is a rare and severe disease caused by disruptions in the CTNS gene. Cystinosis is characterized by lysosomal cystine accumulation, vesicle trafficking impairment, oxidative stress, and apoptosis. Additionally, cystinotic patients exhibit weakening and leakage of the proximal tubular segment of the nephrons, leading to renal Fanconi syndrome and kidney failure early in life. Current in vitro cystinotic models cannot recapitulate all clinical features of the disease which limits their translational value. Therefore, the development of novel, complex in vitro models that better mimic the disease and exhibit characteristics not compatible with 2-dimensional cell culture is of crucial importance for novel therapies development. In this study, we developed a 3-dimensional bioengineered model of nephropathic cystinosis by culturing conditionally immortalized proximal tubule epithelial cells (ciPTECs) on hollow fiber membranes (HFM). Cystinotic kidney tubules showed lysosomal cystine accumulation, increased autophagy and vesicle trafficking deterioration, the impairment of several metabolic pathways, and the disruption of the epithelial monolayer tightness as compared to control kidney tubules. In particular, the loss of monolayer organization and leakage could be mimicked with the use of the cystinotic kidney tubules, which has not been possible before, using the standard 2-dimensional cell culture. Overall, bioengineered cystinotic kidney tubules recapitulate better the nephropathic phenotype at a molecular, structural, and functional proximal tubule level compared to 2-dimensional cell cultures.

Chronic kidney disease (CKD) is increasingly linked to intestinal barrier dysfunction, which contributes to systemic inflammation and disease progression. We investigated whether 2′-fucosyllactose (2′-FL), a human milk oligosaccharide, could modulate intestinal epithelial barrier function under uremic toxin exposure and examined the role of the aryl hydrocarbon receptor (AhR). Using wild-type (WT) and AhR knockout (AhR KO) Caco-2 cell lines, we assessed barrier function and cellular responses through transepithelial electrical resistance (TEER), paracellular permeability (Papp), tight junction (TJ) gene expression, AhR target gene activation, and metabolic assays. AhR deficiency impaired epithelial maturation, confirming its role in barrier development. Short-term exposure to uremic toxins (UTs) did not significantly affect TEER or paracellular permeability under the conditions tested. However, 2′-FL reduced paracellular permeability in both WT and AhR KO cells in the presence of FCS. These effects occurred without changes in TJ gene expression, indicating mechanisms beyond transcriptional regulation. AhR target genes were not induced by UTs, suggesting limited receptor activation, and metabolic activity was unaffected. Overall, 2′-FL reduced paracellular permeability under selected experimental conditions, while the impaired barrier development observed in AhR-deficient cells supports a role for AhR in intestinal epithelial barrier development.