Grant Awarded July 2020 by Cystinosis Ireland the Cystinosis Research Network.
A Cellular Resource for Studying Male Infertility in Cystinosis, Minnie Sarwal, MD, PhD, Professor of Surgery, (Director, Precision Transplant Medicine) University of California San Francisco (UCSF), James Smith, MD, MS (Director, Male Reproductive Health Center Urologist), University of California San Francisco (UCSF), Ann Harris, Professor, Department of Genetics and Genome Sciences, School of Medicine, Case Western Reserve University, Cleveland, Ohio, Elena Levchenko, Professor, Department of Pediatric Nephrology, Leuven, EU, and Swastika Sur, MSc., PhD Postdoctoral Scholar, Sarwal Lab, University of California San Francisco, Department of Surgery.
Total Grant: €10,000
Principal Investigator, Swastika Sur, a Postdoctoral Fellow at the University of California San Francisco, has outlined a research proposal focused on infertility in men with cystinosis.
In addition to various endocrine organs that are affected in cystinosis, hypergonadotropic hypogonadism has been reported as a frequent finding in male cystinosis patients. Although spermatogenesis has shown to be intact at the testicular level in some patients, no male cystinosis patient is known to have naturally fathered a child. The sole treatment for cystinosis is the aminothiol cysteamine, which is highly effective in reducing the intracellular levels of cystine. However, this drug is ineffective in the treatment of male infertility in these patients.
A previous study of the pathophysiology of cystinosis-mediated infertility used a CTNS-/- mouse model. However, the CTNS-/- mouse model generated on C57BL/6 background was found to not be suitable for clarifying the pathogenesis of male infertility in cystinosis. This mouse model partially mimics the renal phenotype of the human disease but was found to have normal testicular morphology and function.
Therefore to this date, the exact pathophysiology of male infertility observed in patients with cystinosis is not yet fully understood, which is a critical unmet need due to the growing population of cystinosis patients, treated with cysteamine reaching young adulthood.
This project proposes to generate isogenic immortalized epididymis and testicular cell models to study infertility associated with cystinosis, by using CRISPR/Cas9 technology to develop a deletion in the CTNS gene of normal human epididymis and testicular cells. The Sarwal group’s ongoing collaborations with Dr Smith at UCSF has enabled access to a rich tissue resource of normal testicular and epidydymal samples that will be used for generating this cystinosis- specific resource. The Sarwal research group also has ethical approvals in place at UCSF and at KU Leuven that will allow Dr Sur to approach cystinotic patients for access to patient biosamples.
Previously, the Sarwal research group successfully generated human immortalized CTNS-/- proximal tubular epithelial cells, and confirmed the phenotype of these newly developed cell line in terms of intracellular cystine levels by HPLC-MS/MS.
In this project, Dr Sur will focus on generating human immortalized CTNS-/- epididymal and testicular cells, followed by phenotype validation so that these cell lines can serve as a resource for the research community to study the pathophysiology of male infertility observed in cystinosis.
The Specific Aims of this project are the following:
- Aim 1: Generate human immortalized CTNS -/- epididymal and testis cell lines by CRISPR/Cas9 and confirm the phenotype to further downstream study of male fertility associated with cystinosis
- Aim 2: Map the molecular perturbations in both cell lines with deletion of CTNS and in tissue samples from male cystinotic patients, by using state of the art genomics that the Sarwal Lab has legacy expertise-in. This will define the clinical utility of the resource generated in Aim 1.
This project will provide a first of its kind human cellular models to study cystinosis-mediated male infertility.