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CRISPR-Cas9 Genome Edited MSCs to Target OA Cartilage Regeneration

CRISPR-Cas9 基因组编辑 MSC 以靶向 OA 软骨再生

基本信息

批准号：
10701421
负责人：
Ming Pei
金额：
$ 36.58万
依托单位：
WEST VIRGINIA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-22 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10701421
关键词：
Address Adipose tissue Affect American Animal Model Animals Anti-Inflammatory Agents Arthroscopy Bone Marrow Bypass COL1A1 gene CRISPR/Cas technology Cartilage Cell Aging Cells Chondrocytes Clinical Collagen Data Defect Degenerative polyarthritis Deterioration Dose Engineering Environment Exclusion Exhibits Feedback Fibrocartilages Future Generations Genetic Engineering Genetic Transcription Human Hydrogen Peroxide Implant In Vitro Inflammation Interleukin-1 Joints Knee Osteoarthritis Knock-in Knowledge Lentivirus Vector Mediating Mesenchymal Modeling Monitor Nude Mice Oryctolagus cuniculus Osteogenesis Patients Physiologic Ossification Proverb Refractory Disease Rejuvenation Reporter Resistance Safety Site Source Synovial Membrane Technology Tissue Engineering Tissue constructs Tissues Translating Translations Vascular blood supply WNT Signaling Pathway aggrecan angiogenesis articular cartilage base cartilage cell cartilage degradation cartilage regeneration cartilage repair clinical translation design disability expectation genome editing immunodeficient mouse model implantation improved in vitro testing in vivo innovation lipid biosynthesis novel therapeutics osteochondral repair osteochondral tissue oxidation preference premature prevent senescence small molecule stem cell proliferation stem cell therapy stem cells subcutaneous tissue regeneration

项目摘要

Project Summary Cartilage defects in patients with knee osteoarthritis are often filled with fibrocartilage. Inflammation in joints renders cartilage defects a refractory disease. Mesenchymal stromal/stem cells (MSCs), a promising cell source for the treatment of cartilage defects, exhibit a site-dependent differentiation preference: bone marrow- derived MSCs (BMSCs) exhibit chondrogenic differentiation but end with endochondral ossification; adipose- derived MSCs (ADSCs) are readily available but have limited chondrogenic potential and tend toward fibrocartilage instead of articular cartilage; and synovium-derived MSCs (SDSCs) are a tissue-specific stem cell for chondrogenic differentiation but have limited accessibility. Moreover, a greater number of MSCs is needed for tissue engineering and regeneration; however, MSCs are prone to become senescent during ex vivo expansion leading to a loss of their proliferation and differentiation potentials. Given that MSCs and fibrocartilage have extensive expression of collagen I (COL1A1), in this proposal, we hypothesize that integration of OStrio (both OCT4 and SOX trio) at the COL1A1 locus allows for MSC rejuvenation and articular differentiation while suppressing fibrocartilage by a negative feedback loop. To achieve this hypothesis, we plan to use cutting-edge CRISPR-Cas9 Genome Editing technology for precise integration of OStrio at the COL1A1 locus of MSCs (Aim 1), followed by fine-tuning of proliferation and chondrogenic differentiation of OStrio-expressing MSCs via WNT signals (Aim 2); lastly, genetically engineered MSCs will be assessed in vivo through subcutaneous implantation (with a sufficient vascular supply) of premature tissue constructs from human OStrio-expressing MSCs in an immunodeficient mouse model for future clinical translation (Aim 3.1) and intra-articular implantation of premature tissue constructs from rabbit OStrio-expressing MSCs in an osteochondral defect rabbit model (Aim 3.2). Our expectation is that OStrio-expressing MSCs have superior proliferation and chondrogenic differentiation capacity in an in vitro study as well as superior ability in cartilage resurfacing and resistance to deterioration from inflammation and oxidation as well as angiogenesis in animal studies. Our objective is to engineer “ideal” MSCs which can be prodigiously expanded and specifically differentiated toward articular cartilage with a robust capacity to resist the harsh osteoarthritic environment by preventing hypertrophic cartilage and fibrocartilage formation. To translate this finding into the clinical setting, we will use cutting-edge CRISPR-Cas9 Genome Editing technology to introduce these factors by targeted integration instead of using lentiviral vectors to improve the safety profile of this new therapy. This project has the potential to bypass a critical bottleneck in articular cartilage regeneration by safely providing large quantities of patient-specific functional chondroprogenitors that can survive a harsh osteoarthritic environment.

项目总结