Dr. Lee’s group is exploring the use of a novel gene transfer vehicle, exosomes, to deliver missing portions of the PWS genes to the hypothalamus. In this pilot study, they seek to develop PWS-specific exosomes and test how well these can deliver genes to neurons and other cells.
Theresa Strong, Director of Research Programs, shares details on this project in this short video clip.
Watch the full webinar describing the 11 research projects funded in this grant cycle here.
Prader-Willi syndrome (PWS) is caused by the lack of expression of genes located on chromosome 15 in region q11.2-q13.1. Often the first clue apparent at birth is low muscle tone and a poor interest in feeding with ineffective suckling. In early childhood, there is a transition through several nutritional phases to excessive appetite and an inability to feel full, leading to a condition called hyperphagia. Despite much interest in this area, the mechanism by which this transition occurs remains poorly understood. One clue to understanding this transition is clearly defining the affected portion of chromosome 15 that is causing PWS in an individual. Such data from several individuals with small missing pieces of the chromosome implicate a gene cluster called SNORD116 composed of small non-coding RNAs. SNORD116 expression is specific to the paternal chromosome and notably is expressed primarily in the brain. In particular, a part of the brain called the hypothalamus has high expression of SNORD116. There is a specific nucleus or collection of densely packed neuronal cells within the hypothalamus called the arcuate nucleus that seems to play a role in feeding behavior. In fact, it has been established that deletion of the paternal SNORD116 alone is necessary and sufficient to cause all of the hallmark features of PWS.
For the 2018 FPWR application, we propose to engineer exosomes that can specifically target the hypothalamus in PWS. Exosomes are lipid-based extracellular vesicles that are secreted by cells and used to communicate with other cells via their vesicle-bound cargo, which can include RNAs, DNAs, proteins, or lipids. Exosome targeting and cargo delivery are governed by the “like attracts like” principle, where they are most likely to interact with cells that express the same surface proteins as their own. Exosomes have attracted much attention over the past several years as an effective pharmacological delivery system, due to their relatively small size ideally-suited for the transport of non-coding RNAs, low levels of immune-mediated response commonly associated with virus-based gene therapy, their ability to readily cross the blood-brain and placental barriers, and the exciting possibility of targeting them to specific tissues. Most importantly, because exosomes are not viral particles and do not contain synthetic vector DNA, they have high translational potential in humans.
We will engineer PWS-specific exosomes by identifying proteins that are found on the surface of hypothalamic tissues from PWS mice and humans. We will then use neuronal cell lines to express these PWS-specific proteins and to generate exosomes that carry these proteins on their vesicle surface. We will use these PWS-specific exosomes to deliver SNORD116 to cell lines derived from different tissue types to test maximal delivery of SNORD116 to cells of hypothalamic origin. After confirming the successful delivery of SNORD116, we will document reversal of PWS-associated gene expression patterns obtained from a mouse study. Successful development of PWS-specific exosomes will allow us to move forward in testing SNORD116 delivery in rodents and primates. Use of exosomes are innovative and to our knowledge have not been utilized as a vessel for the delivery of SNORD116 to the brain, and we believe that successful implementation of this highly-translational research can take us one step closer to a cure for PWS.
Richard Lee, PhD.
Richard Lee, PhD.