Cell-Based Phenotypic Drug Screening Identifies Luteolin as Candidate Therapeutic for Nephropathic Cystinosis

J Am Soc Nephrol. 2020 Jul;31(7):1522-1537. doi: 10.1681/ASN.2019090956. Epub 2020 Jun 5.

Abstract

Background: Mutations in the gene that encodes the lysosomal cystine transporter cystinosin cause the lysosomal storage disease cystinosis. Defective cystine transport leads to intralysosomal accumulation and crystallization of cystine. The most severe phenotype, nephropathic cystinosis, manifests during the first months of life, as renal Fanconi syndrome. The cystine-depleting agent cysteamine significantly delays symptoms, but it cannot prevent progression to ESKD and does not treat Fanconi syndrome. This suggests the involvement of pathways in nephropathic cystinosis that are unrelated to lysosomal cystine accumulation. Recent data indicate that one such potential pathway, lysosome-mediated degradation of autophagy cargoes, is compromised in cystinosis.

Methods: To identify drugs that reduce levels of the autophagy-related protein p62/SQSTM1 in cystinotic proximal tubular epithelial cells, we performed a high-throughput screening on the basis of an in-cell ELISA assay. We then tested a promising candidate in cells derived from patients with, and mouse models of, cystinosis, and in preclinical studies in cystinotic zebrafish.

Results: Of 46 compounds identified as reducing p62/SQSTM1 levels in cystinotic cells, we selected luteolin on the basis of its efficacy, safety profile, and similarity to genistein, which we previously showed to ameliorate other lysosomal abnormalities of cystinotic cells. Our data show that luteolin improves the autophagy-lysosome degradative pathway, is a powerful antioxidant, and has antiapoptotic properties. Moreover, luteolin stimulates endocytosis and improves the expression of the endocytic receptor megalin.

Conclusions: Our data show that luteolin improves defective pathways of cystinosis and has a good safety profile, and thus has potential as a treatment for nephropathic cystinosis and other renal lysosomal storage diseases.

Keywords: apoptosis; autophagy; endocytosis; lysosomal storage disease; reactive.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Transport Systems, Neutral / genetics
  • Animals
  • Antioxidants / adverse effects
  • Antioxidants / pharmacology*
  • Apoptosis / drug effects
  • Autophagy / drug effects
  • Cells, Cultured
  • Cystinosis / drug therapy*
  • Cystinosis / metabolism
  • Disease Models, Animal
  • Drug Evaluation, Preclinical / methods*
  • Endocytosis / drug effects
  • Humans
  • Kidney Tubules, Proximal / pathology
  • Low Density Lipoprotein Receptor-Related Protein-2 / metabolism
  • Luteolin / adverse effects
  • Luteolin / pharmacology*
  • Lysosomes / drug effects
  • Mice
  • Oxidative Stress / drug effects
  • Phenotype
  • RNA, Messenger / metabolism*
  • Sequestosome-1 Protein / genetics
  • Sequestosome-1 Protein / metabolism
  • Zebrafish

Substances

  • Amino Acid Transport Systems, Neutral
  • Antioxidants
  • CTNS protein, human
  • Low Density Lipoprotein Receptor-Related Protein-2
  • RNA, Messenger
  • SQSTM1 protein, human
  • Sequestosome-1 Protein
  • Sqstm1 protein, mouse
  • Luteolin