ABSTRACT Down Syndrome (DS), or Trisomy 21 (T21), is a highly common cause of cognitive disability in children, and also impacts individuals throughout life, by increased risks of congenital heart disease, viral susceptibility, leukemia, and conditions such as metabolic, autoimmune and autism spectrum disorders. Furthermore, as individuals with DS live longer, we now know that most develop early-onset Alzheimer Disease (AD). Hence, understanding how three copies of tiny chromosome 21 elevates risks of conditions that afflict the non-DS population is broadly important, especially for the pressing medical challenge of AD. Identification of how T21 impacts specific certain cell-types to cause various aspects of DS has been hampered by variation between all people, and the genetic and phenotypic complexity of DS. Thus, a critical need is for improved ways to identify core effects of T21 on specific cell types and tissues, and the corresponding gene pathways involved. We have developed an inducible chromosome silencing system in DS iPS cells that allows a “reductionist” approach, keeping other variables constant while manipulating just chromosome 21 over-expression. In the first funding period, we validated the utility of this system to correct the known hematopoietic cell pathogenesis in DS and now have extended this to study neurogenesis and angiogenesis, which we will further investigate here. Our progress includes a major advance, as we have identified XIST “minigenes” which repress clustered genes in just a small chromosomal region, which we show for the “Down syndrome critical region”. This innovation has broad translational implications and will be used to in multiple ways, including in relation to APP and AD. One sub-aim will also test proof-of-principle of this approach for smaller chromosome duplication disorders, as in autism. Our main focus is to “dissect” the impact of T21 on genome-wide pathways and cell phenotypes, and distinguish ongoing functional effects in a given cell-type versus formation of different cell types. Our approach can define the immediate and direct effects of T21 over-expression, rather than marginal differences in tissues that have diverged in differentiation and pathology, and thus are far downstream of the root effects in cells. We will investigate what cells T21 impacts, when it impacts, and, critically, when effects are preventable or reversible. Such knowledge is key to inform targets for drug development. This work also may advance the longer-term prospects that “chromosome therapy” may eventually provide a therapeutic strategy. Finally, this work also has broad basic impact for understanding genome balance, the coordinated levels of expression for genes throughout the genome. .

PROJECT NARRATIVE The intent of this proposal is to improve our understanding of how an extra chromosome 21 affects cellular processes that result in cognitive deficits and increased risks for other conditions in Down syndrome, particularly early-onset Alzheimer’s disease (AD). Using a novel method of “trisomy silencing” (correction) to inactivate one chromosome 21 in DS stem cells, we will test central concepts, question some existing assumptions, and identify differences in cell state or development to inform therapeutic strategies. Our studies will improve understanding of how and when trisomy impacts brain and vascular systems, and when effects may be reversible. Additionally, a breakthrough approach will be tested in relation to AD and autism.