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WO2019113522A1 - Cellules de crête neurale pour revitaliser des allogreffes crâniennes - Google Patents

Cellules de crête neurale pour revitaliser des allogreffes crâniennes Download PDF

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WO2019113522A1
WO2019113522A1 PCT/US2018/064583 US2018064583W WO2019113522A1 WO 2019113522 A1 WO2019113522 A1 WO 2019113522A1 US 2018064583 W US2018064583 W US 2018064583W WO 2019113522 A1 WO2019113522 A1 WO 2019113522A1
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mscs
cells
allografts
graft
derived
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PCT/US2018/064583
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English (en)
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Dmitriy SHEYN
Juliane Glaeser
Zulma Gazit
Wafa TAWACKOLI
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Cedars-Sinai Medical Center
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/28—Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/29—Parathyroid hormone, i.e. parathormone; Parathyroid hormone-related peptides
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0618—Cells of the nervous system
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
    • C12N5/0662—Stem cells
    • C12N5/0668—Mesenchymal stem cells from other natural sources
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2506/00—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells
    • C12N2506/45—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells from artificially induced pluripotent stem cells

Definitions

  • Described herein are methods and compositions for use with cranial allografts, including grafts coated with mesenchymal stem cells (MSCs) from neural crest cells (NCCs) produced from induced pluripotent stem cell (iPSCs).
  • MSCs mesenchymal stem cells
  • NCCs neural crest cells
  • iPSCs induced pluripotent stem cell
  • Bone grafts are typically used to repair such defects. While the use of autografts is associated with donor site morbidity, allografts consist of nonviable tissue and relies on the invasion of host cells and tissues. Revitalization of cranial allografts is challenging due to the limited reservoir of resident stem cells in the membranous bones of the craniofacial complex. Intermittent Parathyroid hormone (PTH) therapy enhances revitalization of structural allografts in vivo, although host cell engraftment and integration of the allograft is found to be partial.
  • PTH Parathyroid hormone
  • BM-MSCs bone marrow-derived mesenchymal stem cells
  • MSCs mesenchymal stem cells
  • iPSCs Induced pluripotent stem cells
  • iNCCs induced neural crest cells
  • MSCs induced neural crest cells
  • the Inventors aimed to demonstrate a successful differentiation of iPSCs into iNCC-MSCs and to evaluate the impact of iNCC-MSC seeding onto allografts in combination with intermittent PTH therapy on the graft integration and revitalization compared to a BM-MSC/allograft and allograft only treatment in a mouse calvarial defect model.
  • cranium-specific iPSC-derived stem cells that are demonstrated as capable of revitalizing structural allografts. These cells are seeded on the allografts and coating cranial allografts with mesenchymal stem cells derived from induced neural crest cells, provides a new, effective therapeutic avenue for cranial defects. This approach improves allograft function by combination with a reproducible and inexhaustible source of cranium-specific MSCs.
  • iNCC induced neural crest
  • BM bone marrow
  • PTH intermittent parathyroid hormone
  • a method for treating a cranial bone defect including transplanting in a subject, a graft including a quantity of mesenchymal stem cells (MSCs), wherein the graft treats a cranial bone defect in the subject.
  • the MSCs are derived from neural crest cells (NCCs).
  • the NCCs are derived from induced pluripotent stem cells (iPSCs).
  • the method includes administration of PTH.
  • the quantity of PTH includes 0.1 to 1, 1-10, 10-20, 20-30, 30-40, or at least 40ug/kg.
  • the quantity of MSCs includes at least 0.2xl0 6 , 0.5xl0 6 , lxlO 6 , 2xl0 6 , 3xl0 6 , 4xl0 6 or 5xl0 6 cells.
  • treating a cranial bone defect includes one or more of osteogenesis, osteoconduction, osteoinduction, bone volume increase, and bone graft incorporation.
  • the graft is from cranium.
  • the graft is from a long bone.
  • the grafts are allografts.
  • the grafts are autografts
  • a composition including a graft including a quantity of mesenchymal stem cells (MSCs).
  • the MSCs are derived from neural crest cells (NCCs).
  • the NCCs are derived from induced pluripotent stem cells (iPSCs).
  • the graft is an allograft.
  • the graft is from cranium.
  • the graft is from a long bone.
  • the MSCs are coated on the surface of the graft.
  • Figure 1 The potential of allografts to regenerate calvarial defects coated with iNCC- MSC+PTH is higher than with BM-MSC+PTH.
  • A A higher bone volume in the iNCC- MSC+PTH group was detected by pCT, DBU was calculated as BY (week 3)-BV(day 1) in each mouse;
  • B H&E staining shows an improved integration of iNCC-MSC+PTH allograft compared to controls (Top).
  • Immunostaining of the allograft-host junction shows an increased expression of osteocalcin (OC) and bone sialoprotein (BSP) of Dil labeled cells in the iNCC- MSC+PTH group (Bottom). Yellow arrows: allograft-host junction site indicate significance: p ⁇ 0.05.
  • iNCC-MSCs present MSC phenotype and show similar cell viability in vivo.
  • A The expression of MSC surface consensus markers in iNCC-MSC was tested using flow cytometry and found similar to BM-MSCs.
  • B BLI imaging of cell-seeded allografts post- surgery showed that both BM-MSCs and iNCC-MSCs proliferated during the first two weeks and survived on the allograft for at least 6 weeks.
  • bone grafts are typically used to repair cranial loss defects due to trauma or tumor resection.
  • Autografts are associated with donor site morbidity and allografts consist of nonviable tissue with limited function as an osteoconductive scaffold not capable of stimulate new bone formation.
  • allograft healing relies on the invasion of host cells and tissue and occurs at an extremely slow rate. Revitalization is difficult due to the limited reservoir of resident stem cells in the membranous bone of the craniofacial complex.
  • BM-MSCs bone marrow derived mesenchymal stem cells
  • MSCs and neural crest cells are both used in various approaches in craniofacial biology because of their developmental similarities.
  • NCCs neural crest cells
  • the rarity and difficulty of isolating NCCs has prevented feasibility of direct use NCCs in patients.
  • Easily accessible MSCs are readily available in adult tissues such as bone marrow and fat tissues. As a result, they have been a leading choice for regenerative medicine application.
  • NCCs can be derived from induced pluripotent stem cells (iPSCs).
  • iPSCs induced pluripotent stem cells
  • the source material of iPSCs provide a potentially inexhaustible source of patient-specific cells that can be reprogrammed to iPSC-derived NCCs and subsequently to MSCs and might therefore resolve the unmet need for cranium-specific MSCs.
  • iNCCs can be expanded for long term under conditions of bFGF supplementation and TGFP inhibition.
  • iPSC-derived NCCs can possess osteogenic and chondrogenic potential in vitro after mesenchymal induction, there are little or no description of use of iNCC derived MSCs for bone or cartilage repair in vivo.
  • MSC-like cells from iNCCs attempted to investigate cartilage and bone repair in vivo using an athymic nude rat osteochondral defect model.
  • the MSC-like cells were reported as not affecting regenerative repair of osteochondral defects in vivo.
  • iPS- derived MSCs would be an attractive source of cells for regenerative therapy applications for osteochondral repair, it is clear that development of an effective local delivery system, would be required to fulfill their potential for future therapeutic use in vivo.
  • a method for treating a cranial bone defect including transplanting in a subject, a graft including a quantity of mesenchymal stem cells (MSCs), wherein the graft treats a cranial bone defect in the subject.
  • the MSCs are derived from neural crest cells (NCCs) (i.e., neural crest cell-derived MSCs (NCC-MSCs), and induced pluripotent stem cell neural crest cell-derived mesenchymal stem cells (iNCC-MSCs).
  • NCCs neural crest cells
  • iNCC-MSCs induced pluripotent stem cell neural crest cell-derived mesenchymal stem cells
  • the NCCs are derived from induced pluripotent stem cells (iPSCs) (i.e, induced pluripotent stem cell derived neural crest cells (iNCCs).
  • iPSCs induced pluripotent stem cells
  • iNCC-MSCs can be by a variety of techniques known in the art.
  • NCCs can be generated from iPSCs using a modified stem cell maintenance medium by including Fgf2 (8ng/mL), optionally including Igf-l (200ng/mL) and small molecules such as GSK3 inhibitor IX (BIO) (2-4 mM) and SB431542 (20mM).
  • NCCs can also be generated using sonic hedgehog (200ng/mL,) FGF8 (lOOng/mL), brain-derived neurotrophic factor (BDNF) (20ng/mL).
  • FGF8 lOOng/mL
  • BDNF brain-derived neurotrophic factor
  • MSCs can be generated from iNCCs by culturing iNCCs in an exemplary MSC medium, such as alpha- MEM (aMEM), 10% fetal bovine serum and 5ng/mL bFGF.
  • the method includes administration of PTH.
  • the quantity of PTH includes 0.01 to 0.1, 0.1 to 1, 1-10, 10-20, 20-30, 30-40, or at least 40ug/kg.
  • the quantity of MSCs includes at least 0.2xl0 6 , 0.5xl0 6 , lxl0 6 , 2xl0 6 , 3xl0 6 , 4xl0 6 or 5xl0 6 cells.
  • the MSCs express one or more of CD73+, CD44+, Cdl3+, PDGFRcr+ Sca-l+ and Gli-l+.
  • the MSCs do not express one or more of CD45- Terl 19-
  • NCCs express one or more of p75+, Hnkl+, AP2+ and FoxD3+.
  • NCCs do not express one or more of Sox2-, Oct4-, Nanog- and Pax6-.
  • the NCCs express one or more of: CD29, CD57, CD73, CD271, TFAP2A, SoxlO, Pax3, and nestin.
  • MSCs, including iNCC-MSCs express one or more of: CD29, CD 105, CD90 and CD44.
  • treating a cranial bone defect includes one or more of osteogenesis, osteoconduction, osteoinduction, bone volume increase, and bone graft incorporation.
  • the graft is from a flat bone.
  • the graft is from cranium.
  • the graft is from a long bone. Examples of these sources include calavarial, iliac crest, chin, tibial, rib, and resected bone section.
  • the grafts are allografts. In other embodiments, the grafts are autografts
  • the iPSCs are generated from ceils reprogrammed from a blood draw from a subject, including for example, blood cell derived iPSCs (BC-iPSCs).
  • the transplant subject is the same as the donor subject for a blood draw from which BC-iPSCs are derived.
  • a composition including a graft including a quantity of mesenchymal stem cells (MSCs).
  • the MSCs are derived from neural crest cells (NCCs).
  • the NCCs are derived from induced pluripotent stem cells (iPSCs), such cells described herein as induced pluripotent stem cell neural crest cells (iNCCs), and if differentiated to mesenchymal stem cells are described herein as iNCC-MSCs.
  • the graft is an allograft.
  • the graft is from a flat bone.
  • the graft is from cranium.
  • the graft is from a long bone.
  • the MSCs are coated on the surface of the graft.
  • the composition includes contacting a graft with MSCs, including iNCC-MSCs, optionally including culturing of the MSCs and graft.
  • contacting a graft with MSCs and/or culturing of the MSCs includes use of low or non-adherent culture substrates.
  • the MSCs express one or more of CD73+, CD44+, Cdl3+, PDGFRaN Sca-l+ and GH-1+.
  • the MSCs do not express one or more of CD45- Terl 19-
  • NCCs express one or more of p75+, Hnkl+, AP2+ and FoxD3+.
  • NCCs do not express one or more of Sox2-, Oct4-, Nanog- and Pax6-.
  • the NCCs express one or more of: CD29, CD57, CD73, CD271, TFAP2A, SoxlO, Pax3, and nestin.
  • MSCs, including iNCC- MSCs express one or more of: CD29, CD105, CD90 and CD44.
  • the iPSCs are obtained from a subject including cells reprogrammed from a blood draw.
  • cells reprogrammed from a blood draw are made by a method including contacting a quantity of blood cells with one or more vectors encoding a reprogramming factor, and delivering a quantity of reprogramming factors into the blood cells, culturing the blood cells in a reprogramming media, and further wherein delivering the reprogramming factors, and culturing in a reprogramming media generates blood cell derived induced piuripotent stem cells (iPSCs). Further information on iPSC reprogramming is found in U.S. App. No. 15/184,241 and PCX App. No.
  • iPSCs obtained from a subject include cells reprogrammed lymphoblastoid cells or lymphoblast cell lines (LCLs). Further information on iPSC reprogramming is found in Barrett, R. et al. Reliable Generation of Induced Pluripotent Stem Cells from Human Lymphoblastoid Cell Lines. Stem Cells Transl Med. 2014 Dec;3(l2): 1429-34, which is fully incorporated by reference herein.
  • generating iPSCs includes providing a quantity of cells, delivering a quantity of reprogramming factors into the cells, culturing the cells in a reprogramming media for at least 4 days, wherein delivering the reprogramming factors, and culturing generates induced pluripotent stem cells.
  • the cells are primary culture cells.
  • the cells are blood cells (BCs).
  • the blood cells are T-cells.
  • the blood cells are non-T- cells.
  • the cells are mononuclear cells (MNCs), including for example peripheral blood mononuclear cells (PBMCs).
  • the cells are primary granulocytes, monocytes and B-lymphocytes.
  • the reprogramming factors are Oct-4, Sox-2, Klf-4, c-Myc, Lin-28, SV40 Large T Antigen (“SV40LT”), and short hairpin RNAs targeting p53 (“shRNA- p53”).
  • these reprogramming factors are encoded in a combination of vectors including pEP4 E02S ET2K, pCXLE-hOCT3/4-shp53-F, pCXLE-hSK, pCXLE-hUL and pCXWB-EBNAl .
  • the reprogramming media is embryonic stem cell (ESC) media.
  • the reprogramming media includes bFGF. In various embodiments, the reprogramming media is E7 media. In various embodiments, the reprogramming E7 media includes L-Ascorbic Acid, Transferrin, Sodium Bicarbonate, Insulin, Sodium Selenite and/or bFGF. In different embodiments, the reprogramming media comprises at least one small chemical induction molecule. In certain other embodiments, the reprogramming media includes PD0325901, CHIR99021, HA-100, and A-83-01. In other embodiments, the culturing the blood cells in a reprogramming media is for 4-30 days.
  • the iPSCs are capable of serial passaging as a cell line.
  • the iPSCs possess genomic stability.
  • Genomic stability can be ascertained by various techniques known in the art. For example, G-band karyotyping can identify abnormal cells lacking genomic stability, wherein abnormal cells possess about 10% or more mosaicism, or one or more balanced translocations of greater than about 5, 6, 7, 8, 9, 10 or more Mb.
  • genomic stability can be measured using comparative genomic hybridization (aCGH) microarray, comparing for example, iPSCs against iPSCs from a non blood cell source such as fibroblasts.
  • aCGH comparative genomic hybridization
  • Genomic stability can include copy number variants (CNVs), duplications/deletions, and unbalanced translocations.
  • CNVs copy number variants
  • iPSCs exhibit no more than about 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 15, 16, 17, 18, 19, or 20 Mb average size of amplification and deletion.
  • BC-iPSCs exhibit no more than about 20-30 Mb average size of amplification and deletion.
  • iPSCs exhibit no more than about 30-40 Mb average size of amplification and deletion.
  • iPSCs exhibit no more than about 40-50 Mb average size of amplification and deletion.
  • the average number of acquired de novo amplification and deletions in iPSCs is less than about 5, 4, 3, 2, or 1.
  • de novo amplification and deletions in fib-iPSCs are at least two-fold greater than in PBMC -iPSCs.
  • the methods produce iPSC cell lines collectively exhibiting about 20%, 15%, 10%, 5% or less abnormal karyotypes over 4-8, 9-13, 13-17, 17-21, 21-25, or 29 or more passages when serially passaged as a cell line.
  • the reprogramming factors are delivered by techniques known in the art, such as nucleofection, transfection, transduction, electrofusion, electroporation, microinjection, cell fusion, among others.
  • the reprogramming factors are provided as RNA, linear DNA, peptides or proteins, or a cellular extract of a pluripotent stem cell.
  • the cells are treated with sodium butyrate prior to delivery of the reprogramming factors.
  • the cells are incubated or 1, 2, 3, 4, or more days on a tissue culture surface before further culturing. This can include, for example, incubation on a Matrigel coated tissue culture surface.
  • the reprogramming conditions include application of norm-oxygen conditions, such as 5% O2, which is less than atmospheric 21% O2.
  • the reprogramming media is embryonic stem cell (ESC) media.
  • the reprogramming media includes bFGF.
  • the reprogramming media is E7 media.
  • the reprogramming E7 media includes L- Ascorbic Acid, Transferrin, Sodium Bicarbonate, Insulin, Sodium Selenite and/or bFGF.
  • the reprogramming media comprises at least one small chemical induction molecule.
  • the at least one small chemical induction molecule comprises PD0325901, CHIR99021, HA-100, A-83-01, valproic acid (VP A), SB431542, Y-27632 or thiazovivin (“Tzv”).
  • culturing the BCs in a reprogramming media is for at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 days.
  • compositions of mesenchymal stem cells (MSCs), inducing MSCs derived from neural crest cells (NCCs), further including NCCs derived from induced pluripotent stem cells (iNCCs), further including induced pluripotent stem cells derived from blood cells (BCs-iPSCs). Further described here in a graft including one or more the aforementioned cell types. Further described herein is a method of using a composition including a graft including the quantity of mesenchymal stem cells (MSCs). In various embodiments, this includes a graft including induced pluripotent stem cell-derived neural crest cell derived mesenchymal stem cells (iNCC-MSCs).
  • the iPSCs are generated from cells reprogrammed from a blood draw from a subject, including for example, blood cell derived iPSCs (BC-iPSCs).
  • the method promotes one or more of osteogenesis, osteoconduction, osteoinduction, bone volume increase, and bone graft incorporation.
  • the method includes transplanting the graft into a subject with a cranial bone defect.
  • the transplant subject is the same as the donor subject for a blood draw from which BC-iPSCs are derived.
  • iNCC-MSCs human iPSCs were reprogramed by the Cedars-Sinai iPSC Core Facility from healthy human fibroblasts, which were nucleofected with episomal plasmid vectors. The iPSC lines were expanded and differentiated to iNCCs. NCC phenotype was verified using immunofluorescent staining and flow cytometry for NC markers. Differentiation of iNCCs to MSCs was performed by culturing the cells in standard media and passing. As reference, BM-MSCs were isolated from whole bone marrow aspirates using standard plastic adherence.
  • Osteogenic differentiation of iNCC-MSCs and BM-MSCs was shown in terms of a quantitative alkaline phosphatase (ALP) assay.
  • the iNCC-MSCs’ adipogenic differentiation potential was analyzed using Oil Red O staining.
  • the tumorigenic potential of the iNCC-MSCs was determined using the soft agar assay in vitro and teratoma formation assay in vivo. To exclude teratoma formation, both iNCCs and iNCC-MSCs were injected intramuscularly into NOD/SCID mice.
  • iNCC-MSCs structural calvarial allografts were harvested from FVB/N mice and decellularized chemically and enzymatically to exclude cell remnants. Both BM-MSCs and iNCC-MSCs were transduced with a lentiviral vector encoding for Luciferase reporter gene under constitutive ubiquitin promoter. Per allograft, 10 5 transduced cells were seeded using non-attachment culture plates. Unattached cells were washed out and counted. A calvarial defect (5mm in diameter) was created in NOD/SCID mice and implanted with allografts, with or without cell coating.
  • NGFR-P75 and HNK1 neural crest marker expression via immunofluorescent staining and flow cytometry indicated the successful differentiation of iPSCs into iNCCs. Further differentiation of iNCCs into MSCs demonstrated by the expression of all five consensus MSC markers, tested by flow cytometry. Differentiation of iNCC-MSCs into the osteogenic and adipogenic lineages were shown via ALP activity after 14 days of exposure to osteogenic media, which was comparable between iNCC-MSCs and BM-MSCs. Quantification of fat vacuoles via Oil Red O staining revealed a similar uptake of the stain by both BM-MSCs and iNCC-MSCs. No higher tumorigenic potential of iNCC-MSCs was detected compared to BM- MSCs, tested in week 1, 2 and 4. No teratoma formation was detected after 8 weeks (10 6 cells per injection, n 5).
  • iNCC-MSCs can be generated to present MSC phenotype, including markers such as CD29, CD 105, CD90, and CD44 (Fig. 2A). Imaging of cell-seeded allografts post-surgery showed that both BM-MSCs and iNCC-MSCs proliferated during the first two weeks and survived on the allograft for at least 6 weeks (Fig. 2B).
  • Fig. 2A Imaging of cell-seeded allografts post-surgery showed that both BM-MSCs and iNCC-MSCs proliferated during the first two weeks and survived on the allograft for at least 6 weeks (Fig. 2B).
  • iPSCs can be obtained from a subject including cells reprogrammed from a blood draw, such cells are described as possess reduced mutational load and genomic stability when compared to other cell sources such as fibroblasts. Thereafter, blood cell derived iPSCs (BC- iPSCs) are differentiated into neural crest cells in accordance with methods descried herein. Graft recipient subjects can be the same as the blood cell donor subject, thereby providing patient-specific immunocompatability.
  • iPSC induced pluripotent stem cell
  • iNCC neural crest cell
  • MSCs mesenchymal stem cell
  • iNCC-MSCs mesenchymal stem cell
  • transplant techniques including use with autografts and allografts, and the particular use of the products created through the teachings of the invention.
  • Various embodiments of the invention can specifically include or exclude any of these variations or elements.
  • the numbers expressing quantities of ingredients, properties such as concentration, reaction conditions, and so forth, used to describe and claim certain embodiments of the invention are to be understood as being modified in some instances by the term“about.” Accordingly, in some embodiments, the numerical parameters set forth in the written description and attached claims are approximations that can vary depending upon the desired properties sought to be obtained by a particular embodiment. In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodiments of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as practicable. The numerical values presented in some embodiments of the invention may contain certain errors necessarily resulting from the standard deviation found in their respective testing measurements.
  • the terms“a” and“an” and“the” and similar references used in the context of describing a particular embodiment of the invention can be construed to cover both the singular and the plural.
  • the recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context.

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Abstract

L'invention concerne des procédés et des compositions associés à l'utilisation de cellules dérivées de cellules souches pluripotentes induites (CSPi) spécifiques du crâne dont on a démontré qu'elles étaient capables de revitaliser des allogreffes structurales. En particulier, les cellules de crête neurale dérivées de CSPi ((iNCC) peuvent être différenciées en cellules souches mésenchymateuses ((iNCC-MSC), cellules qui sont ensemencées sur des allogreffes. Le revêtement d'allogreffes crâniennes avec des cellules souches mésenchymateuses dérivées de cellules de crête neurale induites offre une nouvelle voie thérapeutique efficace pour les défauts crâniens. Cette approche améliore la fonction de l'allogreffe par combinaison avec une source reproductible et inépuisable de MSC spécifiques du crâne. Les résultats présentés dans la présente invention démontrent une intégration et une revitalisation améliorées des allogreffes revêtues de crête neurale induite (iNCC-MSC) par rapport aux allogreffes revêtues de MSE de moelle osseuse (BM), les deux appliquées en combinaison avec une thérapie intermittente par l'hormone parathyroïdienne (PTH) dans un modèle de défaut calvarial.
PCT/US2018/064583 2017-12-08 2018-12-07 Cellules de crête neurale pour revitaliser des allogreffes crâniennes WO2019113522A1 (fr)

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US201762596545P 2017-12-08 2017-12-08
US62/596,545 2017-12-08

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