# US 10,113,146 B2 · Method of collecting placental stem cells

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- Number: US 10,113,146 B2
- Document ID: US10113146B2
- Status: Granted
- Kind: B2
- Application number: 15/177,779
- Filed: 2016-06-09
- Published: 2016-10-06
- Granted: 2018-10-30
- Earliest claimed: 2000-12-06
- Assignees: Celularity Inc.
- Applicants: Celularity, Inc.
- Inventors: Robert J. Hariri
- Web page: https://patentgrep.com/patent/US10113146B2

## CPC

- C12N 5/0605: {Cells from extra-embryonic tissues, e.g. placenta, amnion, yolk sac, Wharton's jelly}
- A01N 1/122: Preservation or perfusion media
- A61K 35/50: Placenta; Placental stem cells; Amniotic fluid; Amnion; Amniotic stem cells
- A61M 1/02: Blood transfusion apparatus (blood infusion by syringes A61M5/14)
- A61P 3/00: Drugs for disorders of the metabolism (of the blood or the extracellular fluid A61P7/00)
- A61P 5/14: of the thyroid hormones, e.g. T3, T4
- A61P 7/06: Antianaemics
- A61P 9/00: Drugs for disorders of the cardiovascular system
- A61P 9/10: for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
- A61P 11/00: Drugs for disorders of the respiratory system
- A61P 13/00: Drugs for disorders of the urinary system
- A61P 13/02: of urine or of the urinary tract, e.g. urine acidifiers
- A61P 13/12: of the kidneys
- A61P 17/00: Drugs for dermatological disorders
- A61P 17/02: for treating wounds, ulcers, burns, scars, keloids, or the like
- A61P 21/00: Drugs for disorders of the muscular or neuromuscular system
- A61P 25/00: Drugs for disorders of the nervous system
- A61P 25/16: Anti-Parkinson drugs
- A61P 25/28: for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
- A61P 27/02: Ophthalmic agents
- A61P 27/06: Antiglaucoma agents or miotics
- A61P 29/00: Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C12M 29/10: {Perfusion}
- C12M 47/04: {Cell isolation or sorting (purging biological preparations of unwanted cells C12N5/0081, determining the presence or kind of microorganism C12Q1/04)}
- C12N 5/0603: {Embryonic cells (production of embryos, nuclear transfer A01K67/027); Embryoid bodies}
- C12N 5/0647: {Haematopoietic stem cells; Uncommitted or multipotent progenitors}
- C12N 5/0668: {Mesenchymal stem cells from other natural sources}
- A61K 35/12: Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells (vaccines or medicinal preparations containing antigens or antibodies A61K39/00)
- A61K 48/00: Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61K 2035/124: {the cells being hematopoietic, bone marrow derived or blood cells}
- A61M 2202/0462: Placental blood, umbilical cord blood
- A61M 2210/1458: Placenta
- A61M 2210/1466: Umbilical cord
- C12N 2509/00: Methods for the dissociation of cells, e.g. specific use of enzymes
- C12N 2510/02: Cells for production
- C12N 2533/90: Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- A01N: PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF (preservation of food or foodstuff A23); BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES (preparations for medical, dental or toiletry purposes which kill or prevent the growth or proliferation of unwanted organisms A61K); PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61M: DEVICES FOR INTRODUCING MEDIA INTO, OR ONTO, THE BODY (introducing media into or onto the bodies of animals A61D7/00; means for inserting tampons A61F13/26; devices for administering food or medicines orally A61J; containers for collecting, storing or administering blood or medical fluids A61J1/05); DEVICES FOR TRANSDUCING BODY MEDIA OR FOR TAKING MEDIA FROM THE BODY (surgery A61B; chemical aspects of surgical articles A61L); DEVICES FOR PRODUCING OR ENDING SLEEP OR STUPOR {(Electrotherapy, e.g. producing anaesthesia by the use of alternating or intermittent currents A61N1/36021)}
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C12M: APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; {APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS}

## Abstract

A method of collecting embryonic-like stem cells from a placenta which has been treated to remove residual cord blood by perfusing the drained placenta with an anticoagulant solution to flush out residual cells, collecting the residual cells and perfusion liquid from the drained placenta, and separating the embryonic-like cells from the residual cells and perfusion liquid. Exogenous cells can be propagated in the placental bioreactor and bioactive molecules collected therefrom.

## Claims

### Claim 1

A method of collecting embryonic-like stem cells from a placenta which has been treated to remove residual cord blood said method comprising:
perfusing a placenta which has been drained of cord blood with an anticoagulant perfusion solution to flush out residual cells and embryonic-like stem cells from said drained placenta, and
collecting said residual cells and embryonic-like stem cells and perfusion solution from the drained placenta.

### Claim 2 (depends on 1)

The method of claim 1 further comprising separating said embryonic-like stem cells from said residual cells and anticoagulant perfusion solution.

### Claim 3 (depends on 1)

The method of claim 1 wherein the placenta is perfused with the anticoagulant solution by passing the anticoagulant solution into one or both of the umbilical artery and umbilical vein of said placenta.

### Claim 4 (depends on 1)

The method of claim 1 including collecting said residual cells and said anticoagulant perfusion solution from the part of the placenta that was attached to the wall of the uterus of the mother.

### Claim 5 (depends on 2)

The method of claim 2 including said embryonic-like stem cells are separated from said residual cells and anticoagulant perfusion solution by centrifugation.

### Claim 6 (depends on 1)

The method of claim 1 including
continuing perfusion of said placenta for maintaining placental viability,
flushing out additional residual cells and additional embryonic-like stem cells,
collecting said additional residual cells, said additional embryonic-like stem cells and said additional anticoagulant perfusion solution, and
separating said additional embryonic-like stem cells from said additional residual cells.

### Claim 7 (depends on 6)

The method of claim 6 including maintaining said perfusion of said placenta for stimulating the production of new cells and bioactive molecules, and collecting said newly produced cells and said bioactive molecules from said placenta.

### Claim 8 (depends on 6)

The method of claim 6 wherein the residual cells and embryonic-like stem cells are collected over a period of 24 to 48 hours after birth.

### Claim 9 (depends on 6)

The method of claim 6 wherein the placenta or cells in the placenta are stimulated to produce bioactive molecules.

### Claim 10 (depends on 7)

The method of claim 7 wherein said newly produced cells and bioactive molecules are collected over a period of 24 to 48 hours after birth.

### Claim 11 (depends on 6)

The method of claim 6 wherein the cells in the placenta are genetically engineered with exogenous DNA.

### Claim 12 (depends on 7)

The method of claim 7 wherein the newly produced cells are genetically engineered with exogenous DNA.

### Claim 13

A method of propagating exogenous cells in a placental bioreactor which has been treated to remove residual umbilical cord blood, comprising:
perfusing a placenta which has been drained of cord blood with an anticoagulant perfusion solution to flush out residual cells and embryonic-like stem cells,
collecting said residual cells, said embryonic-like stem cells, and said anticoagulant perfusion solution from the drained placenta,
removing all remaining viable endogenous cells from the drained placenta,
perfusing the placenta with nutrient perfusion solution,
introducing exogenous cells into the perfused placenta,
incubating the perfused placenta with exogenous cells, and
collecting propagated exogenous cells from the perfused placenta.

### Claim 14 (depends on 13)

The method of claim 13 wherein the remaining viable endogenous cells are removed by irradiation of the perfused placenta.

### Claim 15 (depends on 14)

The method of claim 14 wherein the irradiation is selected from electromagnetic, ultra-violet, X-ray, gamma-, and beta-radiation.

### Claim 16 (depends on 13)

The method of claim 13 wherein the exogenous cells are stem cells.

### Claim 17 (depends on 16)

The method of claim 16 wherein the stems cells are hemopoietic pluripotent stem cells.

### Claim 18 (depends on 13)

The method of claim 13 wherein the exogenous cells are stimulated to produce bioactive molecules.

### Claim 19 (depends on 18)

The method of claim 18 wherein the bioactive molecules are collected prior to collecting the propagated exogenous cells.

### Claim 20 (depends on 13)

The method of claim 13 wherein the exogenous cells are stimulated to propagate by introducing growth factors selected from cytokines, hormones, promoters, repressors, proteins, nucleic acids, viruses, and immunogens.

### Claim 21 (depends on 13)

The method of claim 13 wherein the exogenous cells are selected from animal cells, insect cells, or human cells.

### Claim 22 (depends on 13)

The method of claim 13 wherein the exogenous cells in the placenta are genetically engineered with exogenous DNA prior to collecting or during propagation in the perfused placental bioreactor.

### Claim 23 (depends on 22)

The method of claim 22 wherein the exogenous DNA is introduced into the exogenous cells propagated in the perfused placenta via a vector selected from viral, adenoviral, and retroviral vectors.

### Claim 24 (depends on 22)

The method of claim 22 wherein the exogenous DNA is introduced into the exogenous cells by a method selected from mechanical introduction of the exogenous DNA, chemical mediated uptake of the exogenous DNA, and liposomal mediated uptake of the exogenous DNA.