Division of Medical Genetics

Basic Research

Gene Therapy in Animal Models of Glycogen Storage Disease and Phenylketonuria

Principle Investigator: Dwight Koeberl, MD, PhD

The focus of research in my laboratory has been gene therapy with viral vectors, including adeno-associated virus (AAV) vectors and adenovirus vectors. AAV vectors have been pseudotyped with alternative AAV serotypes to enhance tropism for target tissues, including the liver, heart, and skeletal muscle. Pseudotyped AAV vectors enhanced gene transfer in the knockout mouse models for glycogen storage disease (GSD). GSD results from the deficiency of specific enzymes involved in the storage and retrieval of glucose in the body: glucose-6-phosphatase (G6Pase) in GSD-Ia (von Gierke disease) and acid -glucosidase (GAA) in GSD-II (Pompe disease). G6Pase deficiency in GSD-Ia affects primarily liver and kidney, while GAA deficiency in GSD-II causes primarily muscle disease. AAV vectors transduced liver and striated muscle to replace the deficient enzyme in GSD-Ia and GSD-II mice, subsequently demonstrating efficacy through reduction in glycogen storage and correction of associated biomarkers. Summarizing highlights of our research over the past several years:

GSD-Ia: G6Pase-knockout (G6Pase-KO) mice provide a model for the biochemical abnormalities of GSD-Ia, although early mortality complicates research with both the murine and canine models of GSD-Ia. We prolonged survival and reversed the biochemical abnormalities in GSD-Ia mice. Recently we effectively treated GSD-Ia dogs and mice, demonstrating efficacy during a one year follow-up period.

GSD-II/Pompe disease: Successful gene therapy in GSD-II mice required the evasion of immune responses to introduced GAA; importantly, immune responses complicated enzyme replacement therapy for GSD-II and emphasized the need for gene therapy. We evaded immune responses to introduced GAA by liver-restricted expression with an AAV vector, demonstrating the ability to achieve efficacy by inducing tolerance to human GAA.

Phenylketonuria/PKU: We demonstrated long-term biochemical correction of PKU in mice with an AAV2/8 vector, which is a very significant disorder detected by newborn screening and currently inadequately treated by dietary therapy. Phenylalanine levels in mice were corrected in the blood, and elevated phenylalanine causes mental retardation and birth defects in children born to affected women.

Identification of Gene Regulatory Elements

Principle Investigator: Gregory Crawford, PhD

Now that the human genome has been sequenced, the next major hurdle genomics researchers face is understanding what the genome is telling us. My research involves identifying gene regulatory elements (switches that turn genes "on" and "off") across the genome to help us understand how chromatin structure dictates cell function and fate. For the last 25 years, mapping DNase I hypersensitive sites has been the gold standard method to identify the location of active regulatory elements, including promoters, enhancers, silencers, and locus control regions. I have developed two high-thoughput technologies that can identify most DNase I hypersensitive sites from potentially any cell type from any species with a sequenced genome. We are combining this data with other wet-lab and computational data types to better understand how these regulatory regions control global gene expression in different cell types, different stages of development, and disease.

Genetic and epigenetic basis of neurodevelopmental disorders

Principle Investigator: Yong-hui Jiang, MD, PhD

We are interested in understanding the genetic and epigenetic basis of neurodevelopmental disorders with emphasis on genomic imprinting disorders of Angelman syndrome and Prader-Willi syndrome as well as autism spectrum disorder.
Angelman syndrome is caused by deficiency of brain-specific maternally expressed ubiquitin protein ligase 3A (UBE3A) genes or maternal origin of chromosomal deletion of 15q11-q13. Using a mouse model of Ube3a deficiency we created by gene targeting, we are dissecting the pathogenesis of Angelman syndrome. We are interested in identifying protein substrates of Ube3a and exploring the treatment of Angelman syndrome by epigenetic modifications.
Prader-Willi syndrome is caused by deficiency of the paternal chromosome 15q11-q13 region. There is evidence supporting that deficiency of HBII-85 SnoRNAs in the 15q11-q13 region is responsible for major features of the Prader-Willi syndrome. The function of HBII-85 is unknown. We are interested in identifying the targets of modified by HBII-85 SnoRNAs. Using the mouse model with a deletion of HBII-85 SnoRNA cluster, we will dissect the function of HBII-85 and explore the treatment of Prader-Willi syndrome by using DNA methylation inhibitors.
Autism spectrum disorder is a neurobehavioral disorder with a strong genetic basis. However, the genetic basis for majority of individuals with autism spectrum disorder is unknown. We hypothesize that both genetic and epigenetic defects contribute to the etiology of autism spectrum disorders. We will use medical re-sequencing strategy to identify the genetic basis and use genome wide DNA methylation scan to screen the epigenetic candidates contributing to autism spectrum disorder. One of the most common chromosomal abnormalities found in autism spectrum disorder is the maternal duplication of the 15q11-q13 region. Mutations in synaptic scaffolding protein SHANK3 were also reported in a small but significant set of individuals with autism spectrum disorder. We are modeling autism spectrum disorder in mice by generating and characterizing mice with Shank3 deficiency and the maternal duplication of human15q11-q13 homologous region. Using these mouse models, we will test a hypothesis that autism spectrum disorder is a disorder of synaptic dysfunction.