methods for increasing bone growth and/or enhancing wound healing, for example, fracture repair; inducing division and/or inducing differentiation of a hematopoietic stem cell ; Fibroblast Growth Factor-2; bone morphogenetic proteins
Inventors
Lau, Kin-Hing William • Baylink, David J. • Hall, Susan L. • Chen, Shin-Tai • Mohan, Subburaman
Assignees
Loma Linda University • US Department of Veterans Affairs
Publication Number
US-7816140-B2
Publication Date
2010-10-19
Expiration Date
2026-06-13
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Abstract
The present disclosure provides compositions and methods for increasing bone growth and/or enhancing wound healing, for example, fracture repair. The disclosure provides recombinant nucleic acids useful for promoting bone growth. For example, the disclosure provides recombinant nucleic acids that encode a fibroblast growth factor-2 (FGF-2) analog. The disclosure also provides vectors and cells incorporating these nucleic acids, as well as FGF-2 analogs encode by them. The disclosure also provides a mouse system of bone marrow transplantation and methods for producing as well as methods for using the system. Methods for inducing division and/or inducing differentiation of a hematopoietic stem cell are also provided, as are methods for enhancing bone growth and/or wound repair (for example, fracture repair).
Core Innovation
The invention provides compositions and methods for increasing bone growth and enhancing wound healing, particularly fracture repair, by employing recombinant nucleic acids that encode osteogenic growth factors such as fibroblast growth factor-2 (FGF-2) and analogs thereof. These nucleic acids are incorporated into vectors and cells, including hematopoietic stem cells or progenitor cells, which can home to and engraft in bone marrow to promote bone formation in vivo. The methods encompass ex vivo transduction of hematopoietic stem cells with nucleic acids encoding osteogenic growth factors, followed by transplantation into subjects to enhance bone growth and repair.
The problem being addressed arises from the inconsistent and limited success of previous approaches using FGF-2 for bone growth and repair, including attempts at gene transfer methods that failed to produce significant improvements in various models. There is a need for effective compositions and methods that provide improved delivery and expression of osteogenic growth factors to enhance bone formation systemically or locally, overcoming prior limitations in therapeutic efficacy for skeletal disorders such as osteoporosis and fractures.
Claims Coverage
The claims include two independent claims covering ex vivo methods of inducing division and differentiation of hematopoietic stem cells using nucleic acids encoding secretable analogs of FGF-2 with specific mutations and secretion signal sequences, followed by transplantation into subjects to enhance bone growth or repair.
Ex vivo expression of secretable FGF-2 analog with BMP2/4 secretion signal and cysteine mutations in hematopoietic stem cells
The method involves expressing in a hematopoietic stem cell a heterologous nucleic acid encoding a secretable analog of human FGF-2 comprising a BMP2/4 hybrid secretion signal sequence and mutations substituting alanine for cysteine at position 70 and asparagine for cysteine at position 88, promoting stem cell self-renewal, bone growth, and angiogenesis in vivo.
Introduction of transduced hematopoietic stem or progeny cells into subjects for enhancing bone growth or repair
Transplanting hematopoietic stem cells or their progeny, which express the described osteogenic growth factor, into subjects in need to promote systemic or local bone formation and healing, including treatment of conditions such as osteoporosis, fractures, and osteogenesis imperfecta.
Use of specific promoters and viral vector systems for nucleic acid delivery and regulated expression
Nucleic acids can be introduced by electroporation, lipid-mediated transformation, viral transduction with replication defective vectors such as adenovirus, adeno-associated virus, murine leukemia virus or lentivirus, and can be operably linked to constitutive, tissue-specific, or regulatable promoters including tetracycline-inducible promoters to control expression in target cells or tissues.
Selection and enrichment of appropriate hematopoietic stem cells expressing CXCR4 and other markers for delivery
The hematopoietic stem cells can be pluripotent and enriched from bone marrow, cord blood, or peripheral blood, expressing CXCR4 and other markers such as CD34+, lin−, AC133+, CD45−, and can be derived from human or mouse sources for transplantation.
The independent claims focus on ex vivo methods of transducing hematopoietic stem cells with nucleic acids encoding a modified, secretable FGF-2 analog containing BMP2/4 secretion signals and specific cysteine-substituting mutations, deploying these cells into subjects to enhance bone growth and repair, with delivery performed using various viral vectors and with controlled transgene expression via regulatable promoters.
Stated Advantages
Provides increased efficacy of treatment using osteogenic growth factors such as FGF-2 for enhanced bone growth and repair.
Enables large-scale and consistent secretion of stable and biologically active FGF-2 analogs by engineered hematopoietic stem cells.
Allows targeted delivery of osteogenic growth factors to bone marrow, facilitating systemic and local bone formation with reduced side effects.
Supports regulation of therapeutic gene expression with inducible promoters to control bone growth and minimize adverse effects.
Offers an improved bone marrow transplantation model yielding robust, long-term engraftment with reduced host morbidity and mortality for assessment of osteogenic therapies.
Documented Applications
Treatment of skeletal disorders including osteoporosis, osteogenesis imperfecta, and bone fractures.
Enhancement of wound healing, including fracture repair.
Gene therapy approaches employing hematopoietic stem or progenitor cells as vehicles to deliver osteogenic growth factors to bone tissue.
Use of recombinant FGF-2 analogs to promote bone formation and angiogenesis in vivo.
Screening methods for identifying agents that modulate bone growth using a mouse bone marrow transplantation model.
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