Investigating Adeno-Associated Virus as a Vector for Gene Therapy for Steroid-Resistant Nephrotic Syndrome

Student thesis: Doctoral ThesisDoctor of Philosophy (PhD)


Nephrotic syndrome (NS) is a clinical triad of proteinuria, hypoalbuminaemia and oedema. About 15% of children with NS do not respond to high dose steroids and have steroid-resistant nephrotic syndrome (SRNS). A third of SRNS has an underlying genetic cause, and the most prevalent genetic mutation in Caucasian children is in the NPHS2 gene encoding podocin. Genetic forms of SRNS are mostly unresponsive to immunosuppressants, and these patients progress more quickly to end-stage renal disease. Central to the pathogenesis of nephrotic syndrome is a highly specialised cell called the podocyte, which is an important component of the glomerular filtration barrier.

Gene therapy is an option for treating monogenic diseases that has shown success in other organs e.g. targeting the liver for haemophilia B. Adeno-associated virus (AAV) has been particularly successful when used as a systemic injection in vivo to target Haemophilia B, and is a particularly attractive viral vector due to its relatively favourable safety profile. AAV is non-pathogenic, predominantly episomal and has relatively low immunogenicity compared to other viral vectors. Here, AAV was tested as a vector for gene therapy targeting the podocyte specifically to treat genetic SRNS models in vitro and in vivo.

AAV serotypes 2/8, 2/9 and LK03 were first tested in vitro using GFP and LacZ as reporters. Two podocyte specific promoters (from podocin and nephrin) were also tested. This identified AAV LK03 as being a highly efficient transducer of human podocytes, while AAV 2/8 and 2/9 were both reasonable transducers of mouse podocytes in vitro. The minimal nephrin promoter demonstrated podocyte-specific expression in vitro. Next, AAV serotype LK03 was tested in a human model of genetic nephrotic syndrome in vitro, while AAV 2/9 was used to test a genetic model of nephrotic syndrome in vivo.

AAV LK03 expressing wild type podocin showed partial functional rescue of conditionally immortalised podocytes with the R138Q podocin mutation in vitro. In vivo, AAV 2/9 expressing mouse podocin under a minimal nephrin promoter was injected via the tail vein into conditional podocin knockout mice. This demonstrated partial improvements in urinary albumin: creatinine ratio, in survival, in plasma markers of renal function as well as on histology.

Overall, the AAV serotypes tested above are able to transduce the podocyte, and have demonstrated partial rescue in genetic mouse and human models of nephrotic syndrome.
Date of Award28 Nov 2019
Original languageEnglish
Awarding Institution
  • The University of Bristol
SupervisorMoin Saleem (Supervisor) & Gavin I Welsh (Supervisor)

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