Mutations in bone stem cells can impair bone metabolism and regeneration. Osteogenesis imperfect (OI) is the most common form of genetic bone disease and characterized by bone fragility with clinical manifestations varying from a mild increase in fractures to severe bone deformities and death. Individuals with OI have mutations that alter procollagen structure or production and lead to abnormal collagen fibril formation disrupting bone development and renewal. Treatment options for OI are limited and do not alleviate the complications seen in OI. Our recent work has demonstrated that AAV gene targeting vectors can disrupt the mutant collagen alleles that cause OI and that targeted cells produced normal collagen and formed bone. While these results indicate the effectiveness of genomic editing with targeting frequencies approaching 1%, an enhancement of AAV gene targeting is required to provide more robust in vivo genetic manipulations. Here we will expand upon our successes and propose three lines of investigations to develop a therapeutic approach for patients with inherited bone disease. First we will modify collagen genes using a CRISPR-AAV system that will allow for the disruption or correction of genes that cause OI in vivo. Second, we will investigate whether genetically corrected bone stem cells have a growth advantage in vivo and how TGF-? signaling affects the extra-skeletal manifestations of bone disease. Finally we will conduct a series of experiments in a preclinical rabbit model to establish the optimal gene targeting delivery protocol. Successful completion of this proposal will lead to an improved understand of TGF-? signaling and stem cell kinetics in bone and advance AAV gene targeting toward a clinical trial.

Public Health Relevance

This project addresses several key issues central to the success of gene therapy and the treatment of inherited bone diseases. These include the correction of genetic mutations, improve our understanding of the interactions between cells in the bone, and improve our understanding of cell signaling on bone and other tissues. The strategies developed during the course of this research could be applied to a variety of other stem cells types and used to treat many other disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
3R01AR067707-03S1
Application #
9708157
Study Section
Program Officer
Chen, Faye H
Project Start
2018-07-15
Project End
2020-04-30
Budget Start
2018-09-20
Budget End
2019-04-30
Support Year
3
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Mayo Clinic, Rochester
Department
Type
DUNS #
006471700
City
Rochester
State
MN
Country
United States
Zip Code
55905
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Dudakovic, Amel; Gluscevic, Martina; Paradise, Christopher R et al. (2017) Profiling of human epigenetic regulators using a semi-automated real-time qPCR platform validated by next generation sequencing. Gene 609:28-37
Dudakovic, Amel; Camilleri, Emily T; Xu, Fuhua et al. (2015) Epigenetic Control of Skeletal Development by the Histone Methyltransferase Ezh2. J Biol Chem 290:27604-17