Biology subjects

Jamalpoor, A.

Publications and source records attributed to Jamalpoor, A..

3 recordsLinked to original sources

bioRxiv2023

Gene surgery as a potential treatment option for Nephropathic Cystinosis in vitro

Nephropathic cystinosis is a rare monogenetic kidney disease caused by mutations in the lysosomal transporter cystinosin (encoded by CTNS) that, to date, has no cure. The hallmark of this disease is lysosomal accumulation of cystine and decline in proximal tubular function leading to kidney failure early in life. In this project, we developed a novel gene repair strategy using CRISPR/Cas9 Homology-Independent Targeted Integration (HITI) to restore CTNS. A novel, non-viral peptide-mediated approach was used to deliver the Cas9-guideRNA ribonucleoprotein (RNP) complex and repair templates to conditionally immortalized proximal tubule epithelial cell (ciPTEC) lines. The repair constructs contained either mCherry (1.7 kb), the CTNS Superexon (1.7 Kb) or both (3.2 Kb). The results demonstrated that the smaller mCherry construct achieved a higher repair efficiency (63%) compared to the CTNS-mCherry construct (16%). Clonal expansion of repaired cells showed restoration of lysosomal cystine levels in 70-75% of the clones, which was accompanied by improved mitochondrial bioenergetics. In conclusion, CRISPR/Cas9 HITI can be used to precisely insert repair templates into the genome, resulting in a functional cystinosin restoration, and a reversal of the cystinotic disease phenotype.

bioengineering↗

bioRxiv2020

The lysosomal V-ATPase B1 subunit in proximal tubule is linked to nephropathic cystinosis

Variants in ATP6V1B1, the gene encoding the B1 subunit of the vacuolar H+-ATPase lead to distal renal tubular acidosis and hearing loss of variable degree. Apart from metabolic acidosis, a 23-month-old girl with pathogenic ATP6V1B1 variants presented with increased granulocyte cystine levels, and symptoms of renal Fanconi syndrome, indicating an unknown link to proximal tubulopathy and cystinosis, a disorder of the proximal tubule caused by variants in the CTNS gene. Here, we demonstrated that ATP6V1B1 is expressed in proximal tubules of human kidney tissue, though to a lesser extent than in distal tubules. Further, we used CRISPR/Cas9 technology to selectively knockout ATP6V1B1 or CTNS in human renal proximal tubule cells and performed a full metabolomic and proteomic analysis to compare their phenotype to isogenic control cells. In line with the clinical data, the loss of ATP6V1B1 disrupted the chemiosmotic coupling between the lysosomal V ATPase B1 subunit and cystinosin and resulted in intralysosomal accumulation of cystine and autophagy activation in renal proximal tubule cells. In conclusion, we identified ATP6V1B1 as a central player in renal proximal tubule cells, regulating lysosomal cystine transport and autophagy, and its absence can lead to proximal tubule dysfunction.

pharmacology and toxicology↗

bioRxiv2020

Cysteamine-bicalutamide combination treatment restores alpha-ketoglutarate and corrects proximal tubule phenotype in cystinosis

Nephropathic cystinosis is a severe monogenetic kidney disorder caused by mutations in CTNS, encoding the lysosomal transporter cystinosin, resulting in lysosomal cystine accumulation. The sole treatment, cysteamine, slows down the disease progression, but does not correct the established proximal tubulopathy. Here, we developed a new therapeutic strategy by applying an omics-based strategy to expand our knowledge on the complexity of the disease and prioritize drug targets in cystinosis. We identified alpha-ketoglutarate as a key metabolite linking cystinosin loss, lysosomal autophagy defect and proximal tubular impairment in cystinosis. This insight offered a bicalutamide-cysteamine combination treatment as a novel dual target pharmacological approach for the phenotypical correction of cystinotic proximal tubule cells, patient-derived kidney tubuloids and cystinotic zebrafish.

pharmacology and toxicology↗