[go: up one dir, main page]

WO2025145028A1 - Compositions de cellules endothéliales cornéennes et procédés d'administration associés - Google Patents

Compositions de cellules endothéliales cornéennes et procédés d'administration associés Download PDF

Info

Publication number
WO2025145028A1
WO2025145028A1 PCT/US2024/062087 US2024062087W WO2025145028A1 WO 2025145028 A1 WO2025145028 A1 WO 2025145028A1 US 2024062087 W US2024062087 W US 2024062087W WO 2025145028 A1 WO2025145028 A1 WO 2025145028A1
Authority
WO
WIPO (PCT)
Prior art keywords
aggregates
cec
cells
composition
cell
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/US2024/062087
Other languages
English (en)
Inventor
Arnaud Lacoste
Jay YASIN
Frada BERENSHTEYN
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Aurion Biotech Inc
Original Assignee
Aurion Biotech Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Aurion Biotech Inc filed Critical Aurion Biotech Inc
Publication of WO2025145028A1 publication Critical patent/WO2025145028A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • 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
    • A61K35/30—Nerves; Brain; Eyes; Corneal cells; Cerebrospinal fluid; Neuronal stem cells; Neuronal precursor cells; Glial cells; Oligodendrocytes; Schwann cells; Astroglia; Astrocytes; Choroid plexus; Spinal cord tissue
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/12—Carboxylic acids; Salts or anhydrides thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/42—Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0048—Eye, e.g. artificial tears
    • 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
    • C12N5/0621—Eye cells, e.g. cornea, iris pigmented cells
    • 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
    • C12N2501/00—Active agents used in cell culture processes, e.g. differentation
    • C12N2501/70—Enzymes
    • C12N2501/72—Transferases [EC 2.]
    • C12N2501/727—Kinases (EC 2.7.)
    • 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
    • C12N2513/00—3D culture
    • 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
    • C12N2533/00—Supports or coatings for cell culture, characterised by material
    • C12N2533/50—Proteins
    • C12N2533/54—Collagen; Gelatin

Definitions

  • Corneal endothelial disease is a sight-threatening and debilitating condition affecting millions of people throughout the world. When corneal endothelial cells die or degrade, they do not regenerate. If left untreated, corneal endothelial cell loss can cause corneal edema and loss of vision. Although topical therapy can relieve symptoms of early-stage disease, the only treatments for more severe corneal endothelial disease are full- or partial-thickness corneal transplantation, referred to as penetrating (PK) or endothelial (DSAEK, DMEK) keratoplasty, respectively. Although corneal transplants are effective, there are disadvantages with these procedures, including limited donor organ supply and the risk of donor rejection. In addition, post-operative recovery for corneal transplant patients requires that the patients lie flat on their backs for up to three days for the transplant to adhere to the corneal stroma.
  • CECs corneal endothelial cells
  • AURN001 is a combination cell therapy including corneal endothelial cells (i.e., neltependocel (allogeneic human corneal endothelial cells) and a Rho kinase (ROCK) inhibitor compound (i.e., Y-27632).
  • ROCK Rho kinase
  • Adhesion of the CECs is promoted by ROCK inhibition through the suppression of the Rho/ROCK/MLC (myosin light chain)-signaling cascade.
  • ROCK inhibitor the dissociated cells contract due to actin cytoskeleton activation, which leads to a significant decrease in the cell surface area and a decrease in cell adhesion (Okumura N, Sakamoto Y, Fujii K, et al. Sci Rep. 2016;6:1-11 .)
  • CEC therapy presents an exciting treatment for those having corneal endothelial diseases.
  • CEC corneal endothelial cell
  • CEC aggregate compositions and methods enable faster CEC delivery relative to methods that rely on CEC suspensions, thereby promoting cell adhesion and viability and reducing the amount of time that patients spend in a prone position following injection.
  • the present methods and compositions also have the advantage of potentially eliminating a requirement for a combination therapy involving a Rho kinase inhibitor.
  • the methods described herein relate to a corneal endothelial cell (CEC) composition including a plurality of aggregates of human corneal endothelial cells (CEC), wherein an aggregate includes at least two cells.
  • CEC corneal endothelial cell
  • At least 1% e.g., 1 %, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or more
  • at least 50% of the CECs are in aggregate form relative to single-cell form.
  • at least 80% of the CECs are in aggregate form relative to single-cell form.
  • at least 90% of the CECs are in aggregate form relative to single-cell form.
  • At least 50% of the CECs are in aggregate form in aggregates comprising at least 50 cells relative to single-cell form. In some embodiments, at least 80% of the CECs are in aggregate form in aggregates comprising at least 50 cells relative to single-cell form. In some embodiments, at least 90% of the CECs are in aggregate form in aggregates comprising at least 50 cells relative to single-cell form.
  • the plurality of aggregates have a diameter of at least about 20 microns (e.g. 20 microns, 30 microns, 40 microns, 50 microns, 60 microns, 70 microns, 80 microns, 90 microns, 100 microns, 150 microns, 200 microns, 250 microns, 300 microns, 350 microns, 400 microns, 450 microns, 500 microns, 550 microns, 600 microns, 650 microns, 700 microns, 750 microns, 800 microns, 850 microns, 900 microns, 950 microns, 1 mm, 1.1 mm, 1.2 mm, 1 .3 mm, 1 .4 mm, 1 .5 mm, 1 .6 mm, 1 .7 mm, 1 .8 mm, 1 .9 mm, or more than about 2 mm).
  • 20 microns e.g. 20 microns, 30 microns, 40 micro
  • the plurality of aggregates have a diameter of at least about 50 microns. In some embodiment, the plurality of aggregates have a diameter of at least about 100 microns. In some embodiment, the plurality of aggregates, e.g., at least 60%, 70%, or 80% of aggregates, have a diameter of about 50-300 microns. In some embodiment, the plurality of aggregates, e.g., at least 60%, 70%, or 80% of aggregates, have a diameter of about 50-250 microns. In some embodiment, the plurality of aggregates, e.g., at least 60%, 70%, or 80% of aggregates, have a diameter of about 50-200 microns.
  • the plurality of aggregates e.g., at least 60%, 70%, or 80% of aggregates, have a diameter of about 100-300 microns. In some embodiment, the plurality of aggregates, e.g., at least 60%, 70%, or 80% of aggregates, have a diameter of about 100-250 microns. In some embodiment, the plurality of aggregates, e.g., at least 60%, 70%, or 80% of aggregates, have a diameter of about 100-200 microns.
  • the plurality of aggregates is at a density of at least about one aggregate per 300 microliters. In some embodiments, the plurality of aggregates is at a density of at least about one aggregate to one million aggregates per 300 microliters. In some embodiments, the plurality of aggregates is at a density of about 1 x 10 4 to about 1 x 10 5 aggregates per 300 microliters. In some embodiments, the plurality of aggregates is at a density of about 1 x 10 4 to about 1 x 10 s aggregates per 300 microliters.
  • the CEC composition further includes a cell substrate.
  • the cell substrate is selected from the group consisting of collagen, gelatin, cellulose, polystyrene, polyester, polycarbonate, poly(N-isopropylacrylamide), polylactic acid, polyglycolic acid, hydroxyapatite, and amniotic membrane.
  • the composition does not include a rho kinase inhibitor.
  • the present disclosure provides a pharmaceutical composition including the CEC composition described herein and a pharmaceutically acceptable carrier.
  • the pharmaceutical composition includes an effective dose of corneal endothelial cells (CECs) for the treatment of a corneal disorder.
  • CECs corneal endothelial cells
  • the present disclosure provides a method of manufacturing corneal endothelial cell (CEC) aggregates, said method including seeding CECs into a three- dimensional culture; and incubating the CECs in the three-dimensional culture to form CEC aggregates.
  • CEC corneal endothelial cell
  • the CECs are seeded onto a two-dimensional culture prior to seeding the CECs into the three-dimensional culture.
  • the three- dimensional culture includes a scaffold.
  • the three-dimensional culture includes a scaffold-free suspension.
  • the method further includes subjecting CEC to fluid dynamics that result in aggregate formation.
  • the three-dimensional culture is maintained for less than one minute. In some embodiments, the three-dimensional culture is maintained for at least about 1 hour. In some embodiments, the three-dimensional culture is maintained for at least about 10 hours. In some embodiments, the three-dimensional culture is maintained for at least about 18 hours.
  • the method further includes plating the CEC aggregates onto a cell substrate.
  • the cell substrate is selected from the group consisting of collagen, gelatin, cellulose, polystyrene, polyester, polycarbonate, poly(N-isopropylacrylamide), polylactic acid, polyglycolic acid, hydroxyapatite, and amniotic membrane.
  • the method further includes harvesting the CEC aggregates.
  • the method includes incorporating the CEC aggregates into a pharmaceutical composition.
  • the pharmaceutical composition lacks a Rho kinase inhibitor.
  • the CECs are human CECs (hCECs).
  • a method for treating or preventing a corneal endothelial disease in a subject in need thereof including administering an effective amount of any CEC aggregate composition provided herein.
  • the method includes administering an effective amount a pharmaceutical composition provided herein.
  • a method for treating or preventing a corneal endothelial disease in a human subject in need thereof including administering an effective amount of a composition including corneal endothelial cell (CEC) aggregates to an eye of a subject.
  • CEC corneal endothelial cell
  • the subject lies face down in a prone position for less than three hours after administration of the composition including the CEC aggregates.
  • the method involves administering the composition including the CEC aggregates to the subject in the absence of a rho kinase inhibitor.
  • the method involves administering the composition including the CEC aggregates to an anterior chamber of the eye of the subject.
  • the effective amount of the CEC aggregates includes at least about 1 x 10 3 CECs. In some embodiments, the effective amount of the CEC aggregates includes about 1 x 10 3 to about 2 x 10 6 CECs. In some embodiments, the effective amount of the CEC aggregates includes about 1 x 10 5 to about 2 x 10 6 CECs.
  • the corneal endothelial disease is a bullous keratopathy, a corneal edema, a corneal leukoma, a corneal endothelial inflammation, or a corneal dystrophy.
  • Figs. 1A-1 E depict the results of an assay evaluating the kinetics of corneal endothelial cell (CEC) aggregate adhesion.
  • Fig. 1 A depicts images of plates (AGGREWELLTM) shortly after seeding a CEC suspension into each well (left panel) and 24 hours after incubation to permit the formation of aggregates (right panel).
  • Figs. 1 B and 1C depicts micrographs at 5x magnification (Fig. 1 B) and 10x magnification (Fig. 1 C) of CEC aggregates plated on collagen-coated plate after incubation at each of the indicated time points (i.e., 15 min, 60 min, and 180 min).
  • Fig. 1 A depict images of plates (AGGREWELLTM) shortly after seeding a CEC suspension into each well (left panel) and 24 hours after incubation to permit the formation of aggregates (right panel).
  • Figs. 1 B and 1C depicts micrographs at 5x magnification (Fig. 1 B)
  • FIG. 1 D depicts micrographs at 10x magnification of a cell monolayer formed by CEC aggregates plated on a collagen-coated plate after 14 days of incubation.
  • FIG. 1 E depicts micrographs at 10x magnification of a cell monolayer formed by CEC aggregates (left panel) and a CEC suspension (right panel) plated on a collagen-coated plate after 25 days of incubations.
  • Fig. 2 graphically depicts the results of an assay evaluating the adhesion of CEC aggregates to a collagen-coated plate in the presence (+Y) and absence (-Y) of Y-27632 (an inhibitor of Rho-associated, coiled-coil containing protein kinase [ROCK]).
  • ROCK protein kinase
  • corneal endothelial disease refers to a disease affecting corneal endothelial cells.
  • corneal endothelial diseases include bullous keratopathy, corneal endothelial dystrophies (e.g., cornea guttata, Fuchs endothelial corneal dystrophy, posterior polymorphous corneal dystrophy, and congenital hereditary corneal endothelial dystrophy), iridocorneal endothelial syndrome, viral diseases (e.g., cytomegalovirus endotheliitis and herpetic endotheliitis), exfoliation syndrome, and corneal endothelial graft rejection; as well as inflammation or physical damage associated with external factors, such as keratouveitis, interstitial keratitis, corneal endotheliitis, corneal endothelial cell loss after corneal transplantation, corneal injury after intraocular surgery (e.g., cataract surgery, vitre
  • corneal endothelial dystrophies
  • corneal endothelial cell refers to a cell derived from a corneal endothelium layer or a cell that otherwise has functional and biochemical characteristics of cells in the corneal endothelium layer, including but not limited to primary culture cells, cultured or subcultured cells, and cells induced to differentiate from undifferentiated cells such as stem cells (e.g., embryonic stem cells or induced pluripotent stem cells (iPSCs)) .
  • stem cells e.g., embryonic stem cells or induced pluripotent stem cells (iPSCs)
  • iPSCs induced pluripotent stem cells
  • the cornea is composed of five layers, corneal epithelium, Bowman's membrane (external boundary), Lamina intestinal, Descemet's membrane (internal boundary), and corneal endothelium, in order from the outside (body surface).
  • the corneal endothelium is a single layer of cells that covers the posterior cornea. Markers for characterizing CECs and methods of identifying CECs are known in the art (See e.g., Hamuro J, et al. Invest Ophthalmol Vis Sci. 2016 Aug 1 ;57(10):4385-92. doi: 10.1167/iovs.16-19771. PMID: 27564520; Wongvisavavit, R., etal (2021 ). Regenerative medicine, 16(09), 871 -891 ).
  • cell aggregate or “aggregate form” refers to a plurality of corneal endothelial cells (e.g., at least two CECs) clustered together into a three-dimensional structure.
  • the term “treating” in the context of a corneal endothelial disease refers to therapeutic treatment in order to alleviate one or more symptoms of a corneal endothelial disease.
  • a subject who is treated according to the methods described herein has a corneal endothelial disease, such that treatment alleviates or slows progression of one or more symptoms of the disease.
  • Exemplary symptoms of the corneal endothelial disease include, but are not limited to, loss of vision, blindness, mechanical disruption of the visual axis, opacification and decreased vision, or an otherwise impairment of visual function.
  • the present disclosure provides a method of reducing or ameliorating these symptoms. That is, in some embodiments, the present disclosure provides a method of increasing vision by administering CEC aggregates to a subject having a corneal endothelial disease.
  • preventing refers to a prophylactic measure whereby symptoms of a corneal endothelial disease are inhibited in a subject who may be at risk of developing a corneal endothelial disease or who has been diagnosed with a corneal endothelial disease but is not yet showing certain symptoms, e.g., is not yet showing vision loss.
  • the disclosure provides a method of preventing vision loss in a patient who is at risk of a corneal endothelial disease or who has been diagnosed with a corneal endothelial disease who is at risk of vision loss.
  • the terms “patient” and ‘subject” are used interchangeably herein.
  • the patient is a human patient.
  • the corneal endothelial cell (CEC) aggregates of the provided compositions have properties that improve delivery times and help mediate cell viability and adhesion for proliferation in vivo (e.g., following transplantation into a cornea of a subject).
  • Cell aggregates have several advantages over cells in suspension, including the ability to sink and attach to the posterior side of the cornea faster than current CEC cell suspensions.
  • CEC aggregates also have improved viability relative to CECs in suspension, which are more prone to apoptosis and cell death.
  • a CEC composition comprising a plurality of aggregates of corneal endothelial cells (CEC), such as human CECs (hCECs).
  • CEC corneal endothelial cells
  • each cell aggregate comprises a plurality of viable CECs.
  • the exact number of cells per aggregate may vary, one skilled in the art will recognize that the size of each aggregate and the number of cells per aggregate is limited by the capacity of nutrients to diffuse to the central cells, and that this number may vary within the composition.
  • Cell aggregates may comprise a minimal number of cells (e.g., two or three cells) per aggregate, or may comprise hundreds of cells per aggregate. Accordingly, an individual composition may have aggregates with a range of sizes and cell numbers.
  • At least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or more than 90% of the cells in the composition are in aggregate form.
  • the cells in the present compositions are primarily in aggregate form in contrast to cell suspensions, which primarily include single cells in non-aggregate form.
  • at least about 50%, 60%, 70%, 80%, 90%, or more than 90% of the cells in the composition are in aggregates.
  • at least about 50% of the cells are in aggregate form in the composition.
  • the plurality of aggregates is at a density of at least about 10,000 aggregates per 300 microliters (e.g., 10,000 aggregates, 25,000 aggregates, 50,000 aggregates, 75,000 aggregates, 100,000 aggregates or more per 300 microliters). In some embodiments, the plurality of aggregates is at a density of at least about 100,000 aggregates per 300 microliters (e.g., 100,000 aggregates, 250,000 aggregates, 500,000 aggregates, 750,000 aggregates, 1 ,000,000 aggregates or more per 300 microliters).
  • the plurality of aggregates are at a density of about 1 to about 1 x 10 6 aggregates per 300 microliters (e.g., about 1 to about 1 x 10 2 aggregates, about 1 to about 1 x 10 3 aggregates, about 1 to about 1 x
  • the plurality of cell aggregates includes from about two to 1000 or more cells per aggregate. In some embodiments, the plurality of aggregates have an average of at least about 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, or more than 1000 cells per aggregate. In certain embodiments, the plurality of aggregates has an average of at least about 50 cells per aggregate.
  • a subset of at least about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, and 95% of the aggregates in the population have a cell number above a given threshold.
  • at least about 50% of the aggregates have at least 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, or more than 100 cells.
  • at least about 50% of the aggregates have at least about 50 cells.
  • the cell aggregates are from about 20 microns (i.e., about two cells) to about 1 mm or more in size (e.g., diameter).
  • the plurality of cell aggregates have an average diameter of at least about 20 microns, 30 microns, 40 microns, 50 microns, 60 microns, 70 microns, 80 microns, 90 microns, 100 microns, 150 microns, 200 microns, 250 microns, 300 microns, 350 microns, 400 microns, 450 microns, 500 microns, 550 microns, 600 microns, 650 microns, 700 microns, 750 microns, 800 microns, 850 microns, 900 microns, 950 microns, 1 mm, 1 .1 mm, 1 .2 mm, 1 .3 mm, 1 .4 mm, 1 .5 mm, 1 .6 mm, 1 .7 mm, 1 .8 mm, 1 .9 mm, or more than about 2 mm.
  • the plurality of aggregates have a diameter of at least about 50 microns or 100 microns. In some embodiment, the plurality of aggregates, e.g., at least 60%, 70%, or 80% of aggregates, have a diameter of about 50-300 microns, 50-250 microns, 50-200 microns, 100-300 microns, 100-250 microns, or 100-200 microns.
  • the size of the cell aggregates may vary within a composition (e.g., non-uniform in size). However, in some embodiments, the size of the aggregates present may be substantially uniform in size. By “substantially uniform in size” it is meant that the aggregates' size distribution has a spread not larger than about 10%. In one embodiment, the aggregates' size distribution has a spread not larger than about 5%.
  • the cell aggregates used herein can be of various shapes, such as, for example, a sphere, a cylinder, rod-like, or cuboidal (i.e., cubes), among others.
  • the aggregates are spheroidal in shape.
  • spheroidal cell aggregates it is meant that while the aggregate is generally shaped like a sphere or ellipsoid, the radii of curvature of the aggregate may not be substantially equal for all points on the surface of the aggregate (i.e., vary by substantially more than 10% over all points on the surface of the aggregate).
  • the plurality of aggregates is non-uniform in shape. In some embodiments, the plurality of aggregates is substantially uniform in shape. By “substantially uniform in shape” it is meant that the spread in uniformity of the aggregates is not more than about 10%. In another embodiment, the spread in uniformity of the aggregates is not more than about 5%.
  • the CEC aggregates of the composition are in a monolayer structure. In further embodiments, the CEC aggregates are layered onto a cell substrate.
  • the substrate may act as a scaffold for cultivating the corneal endothelial cells or may only carry the corneal endothelial cell layer after culture. In certain embodiments, the substrate is used for culturing the corneal endothelial cells and also acts as a scaffold that can be transplanted after completion of the culture.
  • the substrate examples include polymer materials derived from naturally-occurring substances such as collagen, gelatin, cellulose and the like, synthesized polymer materials such as polystyrene, polyester, polycarbonate, poly(N-isopropylacrylamide) and the like, biodegradable polymer materials such as polylactic acid, polyglycolic acid and the like, hydroxyapatite, amniotic membrane and the like.
  • the aforementioned substrate is collagen.
  • compositions herein include a substrate (e.g., a collagen substrate) and a cultured CEC aggregate layer.
  • CECs are (1 ) cultured in a two-dimensional culture vessel (e.g., culture dish, culture tube, culture tank etc.),
  • the cells are passaged via further sub-cultures (e.g., 1 -100 passages), (3) the cells are cultured in three-dimensional culture to form cell aggregates; and/or (4) the aggregates are further plated on a cell substrate (e.g., collagen substrate).
  • a cell substrate e.g., collagen substrate
  • the cultured corneal endothelial cells may have at least one, at least two, or all of the following characteristics (e.g., characteristics similar to CECs found in vivo).
  • the cells may have a monolayer structure.
  • the cell density of the cells may be about 10 to -about 10,000 cells/mm2.
  • the visual flat plane shape of the cells may be approximately hexagonal.
  • cells may be regularly aligned.
  • the cells may express markers characteristic of CECs found in vivo.
  • the CECs have a cell surface expression of a marker selected from the group consisting of CD166 positive, CD44 negative to CD44 weakly positive, CD24 negative to weakly positive, CD44 negative to weakly positive, CD105 negative to weakly positive, CD26 negative to weakly positive, CD200 negative to weakly positive, and CD90 negative to weakly positive phenotypes.
  • the CECs have a cell surface expression at the end of P4 selected from the group consisting of sodium-potassium ATPase, ZO-1 , VDAC3, SLC4A4, CLCN3, COL4A2, COL8A1 , COL8A2, CDH2, CD98, CD166, CD340, Integrin a3
  • the composition herein may have functions similar to those of the corneal endothelial cells in living organisms.
  • the present composition may use other agents in combination such as a steroid agent, antibiotic agent, or a ROCK inhibitor.
  • the additional agent e.g., antibiotic or ROCK inhibitor
  • salts formed with a free carboxyl group derived from hydrochloric acid, phosphoric acid, acetic acid, oxalic acid, tartaric acid or the like, salts formed with a free amine group, derived from isopropylamine, triethylamine, 2-ethylaminoethanol, histidine, procaine or the like, and salts derived from sodium, potassium, ammonium, calcium, ferric hydroxide or the like.
  • the CEC aggregate compositions provided herein may be formulated with or without a rho kinase inhibitor.
  • ROCK inhibition was considered necessary to promote adhesion of corneal endothelial cell in cell culture (see, e.g. U.S. Patent No. US1 1633404B2).
  • CEC aggregates may achieve adhesion in the presence or absence of a Rho kinase inhibitor.
  • Rho kinase or “ROCK” (Rho-associated coiled-coil forming kinase: Rho-bound kinase) refers to serine/threonine kinase which is activated with activation of Rho. Examples thereof include ROKalpha (ROCK-II: Leung, T. et al., J. Biol. Chem., 270, 29051 -29054, 1995), p160ROCK (ROKbeta, ROCK-I: Ishizaki, T. et al., The EMBO J., 15(8), 1885-1893, 1996) and other proteins having serine/threonine kinase activity.
  • ROKalpha ROK-II: Leung, T. et al., J. Biol. Chem., 270, 29051 -29054, 1995
  • p160ROCK ROKbeta, ROCK-I: Ishizaki, T. et al.
  • ROCK inhibitors used as a combined agent include compounds disclosed in US Patent No. 4678783 , Japanese Patent No. 3421217 , WO 95/28387 , WO 99/20620 , WO 99/61403 , WO 02/076976 , WO 02/076977 , WO 2002/083175 , WO 02/100833 , WO 03/059913 , WO 03/062227 , WO 2004/009555 , WO 2004/022541 , WO 2004/108724 , WO 2005/003101 , WO 2005/039564 , WO 2005/034866 , WO 2005/037197 , WO 2005/037198 , WO 2005/035501 , WO 2005/035503 , WO 2005/035506 , WO 2005/080394 , WO 2005/103050 , WO 2006/057270 , WO 2007/026664 , and the like.
  • Such compounds can be manufactured by the method described in each disclosed document.
  • Examples thereof include 1 -(5- isoquinolinesulfonyl)homopiperazine or a salt thereof (e.g., fasudil or fasudil hydrochloride), (+)- trans-4-(1 -aminoethyl)-1 -(4-pyridylcarbamoyl)cyclohexanecarboxamide or a salt thereof (e.g., Y- 27632 ((R)-(+)-trans-(4-pyridyl)-4-(1 -aminoethyl)-cyclohexanecarboxamide dihydrochloride monohydrate), and the like), and preferably comprising Y-27632.
  • 1 -(5- isoquinolinesulfonyl)homopiperazine or a salt thereof e.g., fasudil or fasudil hydrochloride
  • the CEC aggregate composition includes a Rho kinase inhibitor.
  • a Rho kinase inhibitor may be added when culturing, proliferating, differentiating or maturing aggregates of corneal endothelial cells.
  • Such an agent may be included in a cell composition for administration to a subject or provided in a separately administered form.
  • the additional agent may be provided as a kit or combined agent. When used as a kit or combined agent, a package insert that describes the usage method thereof may also be combined.
  • the compositions lack a Rho kinase inhibitor.
  • a Rho kinase inhibitor may be omitted when culturing, proliferating, differentiating, or maturing aggregates of corneal endothelial cells.
  • compositions including the CEC aggregate compositions described herein and a pharmaceutically acceptable carrier or excipient.
  • pharmaceutically acceptable refers to a material, such as a carrier or diluent, which does not abrogate the biological activity or properties of the compound, and is relatively nontoxic, i.e., the material may be administered to an individual without causing undesirable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained.
  • carrier refers to a culture, infusion vehicle, irrigating solution, diluent, adjuvant, excipient, or vehicle administered in conjunction with a medicament, such as a cellular composition provided herein.
  • Such a composition contains a therapeutically effective amount of cellular agent together with a suitable amount of carrier, such that the composition is provided in a form suitable for administration to a patient.
  • the pharmaceutically acceptable carrier is a cell infusion vehicle.
  • the cell infusion vehicle can be any solution in which a cell can be maintained.
  • Cell infusion vehicles include those which can be used as an intraocular irrigating solution or the like. Examples of solutions used as a cell infusion vehicle include Opti-MEM (e.g., with or without additional supplements), Opeguard-MA, Opeguard-F, and the like.
  • the cell infusion vehicle may further comprise additional components, such as at least one of albumin, ascorbic acid (or ascorbate), and lactic acid (or lactate). Addition of these components may facilitate cell maintenance.
  • albumin, ascorbic acid (or ascorbate), and lactic acid (or lactate) are added to a cell infusion vehicle.
  • a solution using Opeguard-MA® or Opti-MEM and at least one, two or all three of albumin, ascorbic acid, and lactic acid is used.
  • the composition can be prepared as a pharmaceutical composition adapted to administration to humans in accordance with a known method. Such a composition can be administered by injection or infusion. When a composition is to be administered by injection, the composition can be distributed by using an injection bottle containing cell infusion solution, aseptic agent-grade water or saline.
  • Corneal endothelial cells may be collected by any conventional methods known in the art from the cornea of a suitable corneal donor.
  • the CECs may be isolated by stripping Descemet's membrane, followed by enzyme treatment to remove the collagen matrix. These cells may undergo further analysis to confirm their biological characteristics and to verify criteria for therapeutic use. Markers for characterizing CECs and methods of identifying CECs are known in the art (See, e.g., Hamuro J, et al. Invest Ophthalmol Vis Sci. 2016 Aug 1 ;57(10):4385-92. doi: 10.1167/iovs.16-19771 . PMID: 27564520; and Wongvisavavit, R., et al (2021 ). Regenerative medicine, 16(09), 871 -891 , which are each hereby incorporated by reference).
  • the CECs are from a human CEC primary cell line.
  • Homogeneous corneal endothelial cells may be prepared using methods known in the art. For example, the Descemet's membrane and the endothelial cell layer of a corneal tissue may be detached from the corneal stroma, transferred into a culture vessel (e.g., a culture dish), and treated with an enzyme, such as collagenase A.
  • the CECs with Descemet's membrane and the endothelial cell layer are digested in a basal growth medium (e.g., OPTI-MEM® I Reduced Serum Media (Thermo Fisher Scientific, Inc., e.g., free of ammonium meta vanadate combined with fetal bovine serum (e.g., 8%)), which may be supplemented with additional components such as calcium chloride (e.g., 200 mg/L), chondroitin sulfate (e.g., 0.08%), and/or an antibiotic (e.g., gentamicin).
  • the CECs may be digested at 37°C for two to 24 hours.
  • the corneal endothelial cells are detached from the Descemet's membrane.
  • the corneal endothelial cells remaining in the Descemet's membrane can be further detached by mechanical methods, such as pipetting.
  • This step may additionally include one or more washing steps (e.g., using the basal growth medium without an enzyme).
  • the corneal endothelial cells may then be cultivated in a suitable culture medium that permits growth of CECs (e.g., in an initial culture at passage 0).
  • a suitable culture medium that permits growth of CECs (e.g., in an initial culture at passage 0).
  • the CECs are cultured in a basal growth medium (e.g., OPTI-MEM® I Reduced Serum Media (Thermo Fisher Scientific, Inc., e.g., free of ammonium meta vanadate and combined with fetal bovine serum (e.g., 8%)).
  • the basal growth medium is further supplemented with an epidermal growth factor (EGF) and/or ascorbic acid (e.g., 20 pg/mL).
  • EGF epidermal growth factor
  • ascorbic acid e.g., 20 pg/mL
  • the basal growth medium further comprises a Rho-associated protein kinase (ROCK)-inhibitor, such as Y-27632.
  • a Rho-associated protein kinase (ROCK)-inhibitor such as Y-27632.
  • DMEM Denssion Eagle's Medium
  • FBS fetal bovine serum
  • b-FGF basic-fibroblast growth factor
  • antibiotics such as penicillin, streptomycin and the like can be used.
  • the CECs are initially cultured in two-dimensional cell culture before generation of cell aggregates.
  • adherent cells are grown in a monolayer system on a flat surface, e.g., in a culture dish, plate, or flask.
  • the culture vessel has a surface coated with Type I collagen, Type IV collagen, fibronectin, laminin or an extracellular matrix of bovine corneal endothelial cells and the like.
  • a conventional culture vessel treated with a commercially available coating agent, such as a FNC coating mix may be used.
  • the temperature for cultivating corneal endothelial cells is not limited as long as the cells proliferate.
  • the cells are cultured at a temperature of about 25°C to about 45°C or about 30°C to about 40°C.
  • the cells are cultured at a temperature of about 37° C.
  • the cells may be cultured in a conventional incubator for cell culture under humidification in an environment of about 5-10% CO 2 . Subculture
  • the cultured corneal endothelial cells prior to generating the cell aggregates, can be subjected to a subculture via the passaging of cells into fresh growth medium.
  • sub-confluent or confluent cells are subjected to the subculture.
  • the subculture includes one or more of the following steps. First, the cells may be detached from the surface of the culture vessel (e.g., by a treatment with trypsin- EDTA, TrypLE enzyme, etc.) and recovered. A culture medium may then be added to the recovered cells to generate a cell suspension.
  • centrifugation may be performed during or after recovery of the cells (e.g., 500 rpm (30 G)-1000 rpm (70 G), 1 to 10 minutes). Such centrifugal treatment generates a cell suspension with a high cell density.
  • the cell suspension may then be seeded and cultured in a culture vessel in the same manner as in the above-mentioned initial culture.
  • the dilution ratio during passage may vary depending on the condition of the cells. In some embodiments, the dilution ratio during passage is about 1 :2-1 :4. In some embodiments, the dilution ratio is about 1 :3.
  • the culture time may vary depending on the condition of the cells to be used. In some embodiments, the culture time is 7- 30 days. This subculture can be performed multiple times where necessary. In some embodiments, the growth medium is exchanged every three to four days.
  • the cultured CEOs are passaged at least one time prior to generation of CEO aggregates. In certain embodiments, the cultured CECs are passaged at least two times prior to generation of CEC aggregates. In some embodiments, the CECs are collected after passage two. In some embodiments, the CECs are collected after passage three.
  • the three-dimensional culture comprises a scaffold, such as a hydrogel, bioceramic, metallic, or polymer scaffold.
  • the scaffold is a solid surface (e.g., a plate or well) having a plurality of microwells or cavities (see, e.g., International Publication Nos. W02008106771 A1 , which is hereby incorporated by reference).
  • the solid scaffold comprising microwells is an AGGREWELLTM plate.
  • the microwells may be of a variety of sizes, depending on the particular embodiment and the intended use of plate. For example, wells may have a dimension of about 100 microns, 200 microns, 400 microns, or about 800 microns. In one embodiment, the microwell is about 400 microns in diameter.
  • the surface may include a plurality of microwells (e.g., at least 1000 microwells).
  • an anti-adherent coating e.g., pluronic acid or commercially available anti-adherent solutions, e.g., StemCell Technologies, cat#421254
  • an anti-adherent coating may be applied to sidewalls of the scaffold surface.
  • sidewalls may be Matrigel coated.
  • the surface is washed with the anti-adherence solution prior to plating the CEC cells on the surface.
  • the surface is further washed with a cell culture medium.
  • cells can be seeded onto the surface to generate cell aggregates.
  • the number of cells in a suspension of corneal endothelial cells can be counted and a desired number of cells can be added to the surface in culture medium.
  • a desired number of cells per microwell can be added (e.g., 10, 20, 30, 40, 50, or more cells per microwell).
  • the cells may be centrifuged (e.g., at 100xg for 3 minutes) to ensure the cells are sedimented to the bottom of the scaffold surface.
  • the three-dimensional culture comprises a scaffold-free culture that promotes formation of CEC aggregates, e.g., forced-floating method, hanging drop method, or agitation-based method.
  • forced-floating method a cell suspension is loaded into the wells of a low adhesion polymer-coated microplate. The microplate is then centrifuged to force the cells to form aggregates.
  • hanging drop method a cell suspension is loaded into the wells of a hanging drop plate. The suspension will hang from the plate in droplets. The cells aggregate in the tips of these drops and form aggregates.
  • agitation-based method a cell suspension is placed in a rotating bioreactor. The cells do not adhere to the walls due to the continuous stirring, resulting in the formation of aggregates.
  • the cells may be incubated for a period of time to permit aggregate formation.
  • the cells are incubated for at least about 1 hour (e.g., at least about 1 hour, 2 hours, 5 hours, 10 hours ,15 hours, 18 hours, 20 hours, 24 hours, or 48 hours or any value therebetween) during which the cells form aggregates ("first incubation").
  • the cells are incubated for about 1 hour to 48 hours, 1 to 2 hours, 2 to 5 hours, 5 to 10 hours, 10 to 15 hours, 16 to 24 hours, or 18 to 20 hours.
  • the cells may be incubated for shorter periods of time (e.g., in instances where additional methodologies, such as centrifugation, are used to facilitate aggregate formation). Accordingly, in some embodiments, the cells are incubated for less than one hour (e.g., less than 1 hour, 45 minutes, 30 minutes, 15 minutes, 10 minutes, 5 minutes, 1 minute, 30 seconds, 20 seconds, 10 seconds, 10 minutes to less than an hour, 10-50 minutes, 10-40 minutes, 10-30 minutes, 10-20 minutes), during which the cells form aggregates.
  • additional methodologies such as centrifugation
  • the aggregates may be recovered (e.g., by either pipetting or by spinning out the aggregates).
  • the aggregates may be maintained in suspension for a period ranging from 1 to 6 days ("second incubation").
  • the aggregates may then be harvested for analysis or further processing.
  • aggregates may self-organize over time. This may occur in the original well plate during the first incubation or after recovery during the second incubation.
  • the CEC aggregates may be plated onto a cell substrate to form a cell layer.
  • the substrate is used for culturing the corneal endothelial cells and also acts as a scaffold that can be transplanted after completion of the culture.
  • the aforementioned substrate include polymer materials derived from naturally-occurring substances such as collagen, gelatin, cellulose and the like, synthesized polymer materials such as polystyrene, polyester, polycarbonate, poly(N- isopropylacrylamide) and the like, biodegradable polymer materials such as polylactic acid, polyglycolic acid and the like, hydroxyapatite, amniotic membrane and the like.
  • the cell substrate is collagen (e.g., a collagen-coated plate).
  • the number of cells to be seeded may be adjusted to form a cell layer having a desired cell density in the finally-produced corneal endothelial preparation.
  • the cells are seeded to form a cell layer having a cell density of about 1 ,000-about 5,000 cells/mm 2 .
  • the aggregates may be incubated at any suitable temperature and time to permit adhesion to the cell substrate.
  • the plate is incubated at a temperature of about 25°C to about 45°C or about 30°C to about 40°C.
  • the cells are cultured at a temperature of about 37° C.
  • the culture time may vary depending on the condition of the cells to be used. For example, in some embodiments, the culture time is 3-30 days.
  • the CEC aggregates are incubated on the cell substate for e.g., at least 3 days, at least one week, at least two weeks, at least three weeks or more than three weeks.
  • the CEC aggregates may then be collected for incorporation into a composition, e.g., a pharmaceutical composition for therapeutic uses.
  • the aggregates may be mixed with a suitable a carrier to maintain viability of the corneal endothelial cells before transplantation.
  • suitable a carrier include an OPTIM-MEM I medium, corneoscleral graft presentation solution OPTISOL-GSTM, an eye preservation solution for corneal transplantation EPIITM, saline, phosphate buffered saline (PBS) and the like.
  • the eye of the recipient is prepared by mechanically removing abnormal extracellular material and degenerated CECs on Descemet’s membrane (e.g., in the central 8 mm diameter area) of the cornea. This may be performed with a silicone cannula through a 1.6 mm incision at the corneal limbus under local anesthesia. A desired number of cells (e.g., a total of at least 1 x10 5 , at least 1 x10 6 CECs, etc.) can then be injected with a syringe into the anterior chamber of the eye. Postoperative care may include topical steroids and/or prophylactic antibiotics.
  • the patient is not required to lay face-down for three hours, as the CEC aggregates rapidly settle on the inner cornea and allow for cell adhesion.
  • the subject lies face down for less than three hours (e.g., less than 180 min, 170 min, less than 160 min, less than 150 min, less than 140 min, less than 130 min, less than 120 min, less than 110 min, less than 100 min, less than 90 min, less than 80 min, less than 70 min, less than 60 min, less than 50 min, less than 40 min, less than 30 min, less than 20 min, less than 10 min) after administration of the composition comprising the CEC aggregates.
  • the subject lies face down for less than three hours, e.g., less than 120 minutes, less than 60 minutes, less than 30 minutes, about 30-60 minutes.
  • the dosage of the composition that is effective in therapy of a specific disorder or condition may vary depending on the properties of the disorder or condition. However, such an amount can be determined by those skilled in the art by a standard clinical technique based on the descriptions herein. Furthermore, an in vitro assay can be used in some cases to assist the identification of the optimal dosing range. The precise dose to be used in a preparation may also vary depending on the administration pathway or the severity of the disease or disorder.
  • Example 1 Materials and methods - Spheroid generation for faster cell human Corneal Endothelial Cell (HCEC) attachment to the substrate for rapid cell delivery
  • HCEC Corneal Endothelial Cell
  • the CEC HCEC spheroids were generated using AGGREWELL 400TM plates (Stemcell Technologies, Cat. No. 3441 1/34415). 24-well AGGREWELLTM plates were used, where each well contained -1 ,200 microwells with each microwell measuring 400 urn in diameter.
  • Single cells were generated from passage 2 and passage 3 and plated at a concentration of 637,000 cells per well of a collagen-coated clear flat bottom tissue culture plates (Corning, cat#354407) and allowed to attach for the period of 3hrs.
  • Example 2 CEC aggregate generation and study of adhesion kinetics
  • CEC aggregates i.e. , spheroids
  • a cell suspension including approximately 1 .80x10 6 viable cells was utilized for spheroid creation in this Example.
  • a 1 x6 well plate (AGGREWELLTM) was seeded for 24 hours, with 50 cells per microwell ('-6000 spheres for each well of a six well plate). The number of cells in the aggregates was assessed 24 hours after seeding. As shown in Fig. 1A (right panel), within 24 hours, CEC aggregates of 50+ cells were formed.
  • Spheroids were removed from each well were seeded into 6x wells of a collagen-coated 24-well plate to measure and image the kinetics of aggregate attachment. After incubation for 15, 60 or 180 minutes, medium and unattached aggregates in each well were removed and the remaining attached aggregates were subject to analyses. For the analyses, images and timelapse videos were acquired, the number of cells in the attached aggregates were counted, and the number of viable cells in the attached aggregates were counted. Images of the aggregates at each time point are shown at 5x and 10x magnification in Figs. 1 B and 1C, respectively. The kinetics of CEC aggregate sedimentation was additionally assessed by video imaging (data not shown).
  • Example 3 CEC aggregate adhesion kinetics in the presence and absence of a Rho kinase inhibitor (Y-27632)
  • This Example evaluated the impact of Y-27632 (an inhibitor of Rho-associated, coiled- coil containing protein kinase [ROCK]) on adhesion of CEC aggregates.
  • Y-27632 an inhibitor of Rho-associated, coiled- coil containing protein kinase [ROCK]
  • Rho kinase inhibitors were considered necessary to promote adhesion of corneal endothelial cell in cell culture (see, e.g, U.S. Patent No. US11633404B2).
  • CEC aggregates (i.e., spheroids) were generated using the method described in Example 1 . 18-20 hours after aggregate formation, the aggregates were harvested and plated into collagen-coated clear flat bottom tissue culture plates (Corning, cat#354407). After plating, spheroids were placed into a 37°C incubator and allowed to attach for 20 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, and 3 hours. Spheroids were plated in culture media in either presence or absence of 100uM Y-27632 (Eurofins CDMO Alphore, Y-27632 Dichloride, Item code: 70F089P1). The total viable cells attached, the percent of viable cells and the percent adhesion are summarized in Table 1 .

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Animal Behavior & Ethology (AREA)
  • Biomedical Technology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Medicinal Chemistry (AREA)
  • Epidemiology (AREA)
  • Ophthalmology & Optometry (AREA)
  • Biotechnology (AREA)
  • Cell Biology (AREA)
  • Organic Chemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Developmental Biology & Embryology (AREA)
  • Neurology (AREA)
  • Wood Science & Technology (AREA)
  • Genetics & Genomics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Neurosurgery (AREA)
  • Immunology (AREA)
  • Dermatology (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Inorganic Chemistry (AREA)
  • Microbiology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • Virology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)

Abstract

L'invention concerne des compositions comprenant des agrégats de cellules endothéliales cornéennes (CEC) et des méthodes d'administration desdits agrégats à des patients pour le traitement d'une maladie endothéliale cornéenne.
PCT/US2024/062087 2023-12-29 2024-12-27 Compositions de cellules endothéliales cornéennes et procédés d'administration associés Pending WO2025145028A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US202363616104P 2023-12-29 2023-12-29
US63/616,104 2023-12-29

Publications (1)

Publication Number Publication Date
WO2025145028A1 true WO2025145028A1 (fr) 2025-07-03

Family

ID=96219797

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2024/062087 Pending WO2025145028A1 (fr) 2023-12-29 2024-12-27 Compositions de cellules endothéliales cornéennes et procédés d'administration associés

Country Status (1)

Country Link
WO (1) WO2025145028A1 (fr)

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090232772A1 (en) * 2004-12-09 2009-09-17 Shiro Amano Human corneal endothelial cell-derived precursor cells, cellular aggregates, methods for manufacturing the same, and methods for transplanting precursor cells and cellular aggregates

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090232772A1 (en) * 2004-12-09 2009-09-17 Shiro Amano Human corneal endothelial cell-derived precursor cells, cellular aggregates, methods for manufacturing the same, and methods for transplanting precursor cells and cellular aggregates

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
LI ET AL.: "A Novel Method of Isolation, Preservation, and Expansion of Human Corneal Endothelial Cells.", IOVS., vol. 48, no. 2, February 2007 (2007-02-01), pages 614 - 620, XP008104571, Retrieved from the Internet <URL:https://iovs.arvojournals.org/article.aspx?articleid=2125977> [retrieved on 20251202], DOI: 10.1167/iovs.06-1126 *
OKUMURA NAOKI, KOIZUMI NORIKO, UENO MORIO, SAKAMOTO YUJI, TAKAHASHI HIROAKI, TSUCHIYA HIDEAKI, HAMURO JUNJI, KINOSHITA SHIGERU: "ROCK Inhibitor Converts Corneal Endothelial Cells into a Phenotype Capable of Regenerating In Vivo Endothelial Tissue", THE AMERICAN JOURNAL OF PATHOLOGY, vol. 181, no. 1, 1 July 2012 (2012-07-01), US , pages 268 - 277, XP093134411, ISSN: 0002-9440, DOI: 10.1016/j.ajpath.2012.03.033 *
ONG HON SHING, ANG MARCUS, MEHTA JODHBIR: "Evolution of therapies for the corneal endothelium: past, present and future approaches", BRITISH JOURNAL OF OPHTHALMOLOGY, vol. 105, no. 4, 1 April 2021 (2021-04-01), GB , pages 454 - 467, XP093333571, ISSN: 0007-1161, DOI: 10.1136/bjophthalmol-2020-316149 *

Similar Documents

Publication Publication Date Title
Xia et al. Magnetic human corneal endothelial cell transplant: delivery, retention, and short-term efficacy
Peh et al. Human corneal endothelial cell expansion for corneal endothelium transplantation: an overview
KR102073730B1 (ko) 인간 ｒｐｅ 세포의 생산 방법 및 인간 ｒｐｅ 세포의 제약 제제
EP1888123B1 (fr) Thérapie cellulaire pour la dégénérescence oculaire
US20220169982A1 (en) Rpe cell populations and methods of generating same
EA026193B1 (ru) Композиции и способы лечения заболеваний сетчатки
KR20150004899A (ko) 각막 내피 세포 접착 촉진제
JP2018501281A (ja) 網膜疾患を治療する方法
ES2880346T3 (es) Evaluación de poblaciones de células epiteliales de pigmento retiniano
WO2020045642A1 (fr) Composition et procédé pour la conservation ou la culture de cellules oculaires
JP2025020312A (ja) 多能性幹細胞由来の臨床グレードの角膜内皮細胞の作製および凍結保存
TW202130806A (zh) 用於產生視網膜色素上皮細胞之方法
US20250268837A1 (en) Composition for promoting angiogenesis comprising extracellular vesicles derived from three-dimensional spheroid-type cell aggregate
US20090047738A1 (en) Feeder cell derived from tissue stem cell
WO2025145028A1 (fr) Compositions de cellules endothéliales cornéennes et procédés d&#39;administration associés
WO2025145016A1 (fr) Procédés et compositions de culture de cellules endothéliales cornéennes (cec)
KR102334886B1 (ko) 성장판 재생용 조성물
RU2809003C2 (ru) Способы выделения и культивирования человеческих ретинальных клеток-предшественников
EP3739040B1 (fr) Procédé de mise en feuille pour des cellules dérivées de cellules souches pluripotentes
TW202417614A (zh) 用於治療眼部病症的感光救援細胞（prc）組成物及方法

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 24914364

Country of ref document: EP

Kind code of ref document: A1