This is a multiple-PI R21 project to support mechanistic studies of chaperone-mediate autophagy (CMA) in corneal epithelial cystinotic cells, and to implement translational studies to repair CMA in corneal cystinosis. In lysosomal storage disorders (LSDs), characterized by genetic defects leading to anomalous accumulation of metabolites in lysosomes, cells are affected by lysosomal malfunction frequently leading to cell death. Cystinosis is a lysosomal storage disorder resulting from defects in the cystine transporter cystinosin (CTNS). Increased levels of intralysosomal cystine lead to cell malfunction which is especially manifested in kidneys and eyes. Although treatment with cysteamine retards corneal deterioration, cell malfunction and progressive corneal injury occur despite cystine depletion therapy, suggesting that cystine accumulation is not the only cause of all the defects in cystinosis. We revealed a defective mechanism of chaperone-mediated autophagy in cystinosis. Defective CMA is directly linked to human disease, including eye disease. Our data highlight that CMA impairment is an important contributor to the pathogenesis of cystinosis and underline the need for complementary treatments to cystine-depletion therapies in cystinosis. Here, we aim to study the molecular mechanisms of defective CMA in corneal cystinosis. Furthermore, we propose translational studies that involve the use of already identified small-molecule activators of CMA to protect the cornea, in a mouse model of cystinosis.
In Specific Aim1, we will elucidate mechanisms that mediate LAMP2A destabilization at the lysosomal membrane in corneal epithelial cystinotic cells. We will characterize the LAMP2A trafficking defects in cystinotic epithelial cells and define the importance of CTNS in these processes using vesicle-tracking and super-resolution microscopy.
In Specific Aim 2, we will utilize CMA activators, in vivo, to decrease corneal disease in cystinosis. To this end, we will test CMA activator efficacy in decreasing cystine content, crystal development, endoplasmic reticulum stress and CMA substrate accumulation, and in inducing lysosomal preservation in the cystinotic mice cornea. We will also establish the compound efficacy in preventing and/or decreasing corneal inflammation, corneal degeneration and cell death, and we will combine selected CMA activators with cysteamine to determine the potential additive protective effects of these compounds in corneal cystinosis. It is expected that these basic and translational studies will increase our understanding of CMA in cystinosis and will help develop new therapies for the treatment of corneal cystinosis that will be beneficial for both children and young adults.

Public Health Relevance

Lysosomal storage diseases are caused by genetic defects leading to anomalous accumulation of metabolites in lysosomes, the degradative and recycling compartment of the cell. These toxic products, when accumulated in large amounts, can be harmful to humans. The identification of novel mechanisms that regulate autophagy and compounds that improve cellular and organ function could be useful for the control of several human diseases including lysosomal storage disorders.

National Institute of Health (NIH)
National Eye Institute (NEI)
Exploratory/Developmental Grants (R21)
Project #
Application #
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Mckie, George Ann
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Scripps Research Institute
La Jolla
United States
Zip Code