Cysteamine–bicalutamide combination therapy corrects proximal tubule phenotype in cystinosis

Abstract

Nephropathic cystinosis is a severe monogenic 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 renal proximal tubulopathy. Here, we developed a new therapeutic strategy by applying omics to expand our knowledge on the complexity of the disease and prioritize drug targets in cystinosis. We identified alpha-ketoglutarate as a potential metabolite to bridge cystinosin loss to autophagy, apoptosis and kidney proximal tubule impairment in cystinosis. This insight combined with a drug screen revealed a bicalutamidecysteamine combination treatment as a novel dual-target pharmacological approach for the phenotypical correction of cystinotic kidney proximal tubule cells, patient-derived kidney tubuloids and cystinotic zebrafish.

Benefits and Toxicity of Disulfiram in Preclinical Models of Nephropathic Cystinosis

“…After treatment with 10 µM DSF, cells were incubated with CellROX Green Reagent (Thermo Fisher Scientific) for 30 min at room temperature and reactive oxygen species (ROS) were measured by live cell flow cytometry. GSH and GSSG levels were analyzed as described by Jamalpoor et al, who have performed extensive metabolomic investigations in the same cell conditions [ 12 ].…”

Bioengineered Cystinotic Kidney Tubules Recapitulate a Nephropathic Phenotype

“…Three ciPTECs lines were used for this study: the healthy control ciPTEC, CTNS WT (also referred to as ciPTEC14.4 in previous literature) [ 24 ], and two cystinotic models ciPTEC CTNS Patient (ciPTEC46.2) [ 34 ] and ciPTEC CTNS −/− . CTNS −/− is an isogenic immortalized cell line derived from ciPTEC14.4 and generated in-house previously [ 9 ], which harbors a biallelic mutation in the exon 4 of the CTNS gene. The ciPTEC46.6 (referred to as “Patient”) is an immortalized cell line derived from a urine sample of a cystinotic patient harboring a 57 kb deletion which includes the first 10 exons of the CTNS gene.…”

“…One of the first manifestations of cystinosis is the clinical presentation of renal Fanconi syndrome, characterized by a severe proximal tubule cell dysfunction at early stages of the disease, which results in a total loss of integrity of the proximal tubule [ 7 ]. Great efforts have been made to elucidate further the underlying pathological mechanisms of nephropathic cystinosis that revealed several hallmarks beyond cystine accumulation, including impaired autophagy, mTOR activation, disrupted vesicle dynamics (lysosomes-autophagosomes interactions), mitochondrial impairment, reactive oxygen species (ROS), and increased cell stress [ 8 , 9 , 10 , 11 , 12 , 13 ]. Despite clinical improvements in prognosis, there is, as of yet, no curative therapy available for cystinosis.…”

“…While valuable, their restricted availability and lack of capacity to stay in culture for more than a few passages urged researchers to immortalize primary cultures following different strategies. An example forms the ciPTEC (conditionally immortalized proximal tubule epithelial cells) [ 24 ], a model that has been shown to be particularly useful for investigating the molecular mechanisms affected in nephropathic cystinosis [ 9 ]. However, individual variability and the lack of a healthy control with the same genetic background as the cystinotic donor are major hurdles in pinpointing the molecular mechanism associated with the disease.…”

“…However, individual variability and the lack of a healthy control with the same genetic background as the cystinotic donor are major hurdles in pinpointing the molecular mechanism associated with the disease. To overcome this issue, recent studies have created isogenic cell lines using CRISPR-Cas by knocking out the CTNS gene [ 9 , 10 ]. In addition, advancements in the culture of primary cells obtained from patients’ urine now allow prolonged in vitro expansion in the form of adult organoids.…”