Methods of Manufacture of Immunocompatible Amniotic Membrane Products

Patent metadata

Field	Value
Publication number	US-2020281981-A1
Application number	US-202016736066-A
Country	US
Kind code	A1
Filing date	Jan 7, 2020
Priority date	Feb 18, 2010
Publication date	Sep 10, 2020
Grant date	—

Abstract

Provided herein is a placental product comprising an immunocompatible amniotic membrane. Such placental products can be cryopreserved and contain viable therapeutic cells after thawing. The placental product of the present invention is useful in treating a patient with a tissue injury (e.g. wound or burn) by applying the placental product to the injury. Similar application is useful with ligament and tendon repair and for engraftment procedures such as bone engraftment.

First claim

1 .- 19 . (canceled)

20 . A method of generating an immunocompatible amniotic membrane, the method comprising:
(a) isolating an amniotic membrane from a placenta; (b) depleting the amniotic membrane of a substantial portion of one or more types of immunogenic maternal cells; and (c) cryopreserving the amniotic membrane in a cryopreservation medium at a freezing temperature, thereby generating the immunocompatible amniotic membrane, wherein the immunocompatible amniotic membrane comprises at least 70% viable therapeutic cells, wherein the viable therapeutic cells are native to the amniotic membrane, wherein the viable therapeutic cells comprise two or more cell types selected from mesenchymal stem cells (MSCs), fibroblasts, and epithelial cells, and wherein the immunocompatible amniotic membrane produces a non-immunogenic response in a mixed lymphocyte reaction assay.

21 . The method of claim 20 , wherein the immunogenic maternal cells are maternal leukocytes, maternal dendritic cells, maternal decidual cells, or a combination thereof.

22 . The method of claim 20 , wherein (b) comprises refrigerating the amniotic membrane in the cryopreservation medium for at least 10 minutes.

23 . The method of claim 22 , wherein the refrigerating comprises incubating the amniotic membrane in the cryopreservation medium at a temperature of between about −10° C. and about 15° C. for a time of at least 10 minutes.

24 . The method of claim 23 , wherein the temperature is between 2° C. and 8° C.

25 . The method of claim 23 , wherein the time is at least 30 minutes.

26 . The method of claim 20 , wherein the freezing temperature is between −85° C. and −75° C.

27 . The method of claim 20 , wherein the cryopreservation medium comprises a cell-permeating cryopreservative.

28 . The method of claim 27 , wherein the cell-permeating cryopreservative comprises dimethylsulfoxide (DMSO).

29 . The method of claim 20 , wherein (b) comprises removing trophoblasts from the amniotic membrane.

30 . The method of claim 20 , wherein (b) comprises selectively killing or inactivating functional CD14+ macrophages.

31 . The method of claim 20 , wherein (b) comprises treating the amniotic membrane with an anti-TNF-α antibody.

32 . The method of claim 20 , wherein (b) comprises treating the amniotic membrane with IL-10.

33 . The method of claim 20 , wherein the immunocompatible amniotic membrane generates less than about 50 pg/ml of IL-2αR in the mixed lymphocyte reaction assay.

34 . The method of claim 20 , wherein the immunocompatible amniotic membrane generates less than about 20 pg/ml of IL-2αR in the mixed lymphocyte reaction assay.

35 . The method of claim 20 , wherein the non-immunogenic response is determined by measuring a level of TNF-α release after lipopolysaccharide (LPS) stimulation.

36 . The method of claim 35 , wherein the immunocompatible amniotic membrane produces a level of TNF-α release after LPS stimulation that is less than 420 pg/ml.

37 . The method of claim 35 , wherein the immunocompatible amniotic membrane produces a level of TNF-α release after LPS stimulation that is less than 200 pg/ml.

38 . The method of claim 20 , wherein the viable therapeutic cells further comprise stromal cells present at about 2,000 to about 15,000 cells per cm 2 of the immunocompatible amniotic membrane.

39 . The method of claim 20 , wherein the viable therapeutic cells comprise MSCs, wherein at least 40% of the MSCs remain are viable after a freeze-thaw cycle.

Assignees

• Osiris Therapeutics Inc

Inventors

• Tom Samson
• Danilkovitch Alla
• Yoo Dana
• Jansen Timothy
• Kuang Jin-Qiang
• Marconi Jennifer Michelle

Classifications

• A01N1/125 Primary

  Freeze protecting agents, e.g. cryoprotectants or osmolarity regulators · CPC title

• A61K38/1841

  Transforming growth factor [TGF] · CPC title

• A61K38/1825

  Fibroblast growth factor [FGF] · CPC title

• A61K38/57

  from animals; from humans {(A61K38/553, A61K38/556 take precedence)} · CPC title

• A61K35/50

  Placenta; Placental stem cells; Amniotic fluid; Amnion; Amniotic stem cells · CPC title