COMPOSITIONS AND METHODS FOR THE EXPANSION OF HEMATOPOIETIC STEM AND PROGENITOR CELLS

• US 20190314407A1
• Filed: 10/31/2018
• Published: 10/17/2019
• Est. Priority Date: 10/31/2017
• Status: Abandoned Application

• Current Assignee: Edigene Biotechnology Incorporated
• Original Assignee: Magenta Therapeutics, Inc.
• Inventors: BOITANO, Anthony, COOKE, Michael, GONCALVES, Kevin A.
• Application Number:
  US16/177,252
• Publication Number:
  US 20190314407A1
• Time in Patent Office: Days
• CPC Class Codes: A61K 2035/124 the cells being hematopoietic, bone marrow derived or blood cells
• CPC Class Codes: A61K 31/444 containing a six-membered ring with nitrogen as a ring heteroatom, e.g. amrinone
• CPC Class Codes: A61K 31/4985 Pyrazines or piperazines ortho- or peri-condensed with heterocyclic ring systems
• CPC Class Codes: 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
• CPC Class Codes: A61K 35/28 Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
• CPC Class Codes: C12N 15/102 Mutagenizing nucleic acids
• CPC Class Codes: C12N 15/1058 Directional evolution of libraries, e.g. evolution of libraries is achieved by mutagenesis and screening or selection of mixed population of organisms
• CPC Class Codes: C12N 15/1079 Screening libraries by altering the phenotype or phenotypic trait of the host reporter assays C12N15/1086
• CPC Class Codes: C12N 15/63 Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
• CPC Class Codes: C12N 2501/065 Modulators of histone acetylation
• CPC Class Codes: C12N 2501/19 Growth and differentiation factors [GDF]
• CPC Class Codes: C12N 2501/999 Small molecules not provided for elsewhere
• CPC Class Codes: C12N 2510/00 Genetically modified cells
• CPC Class Codes: C12N 2740/16043 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12N 5/0647 Haematopoietic stem cells; Uncommitted or multipotent progenitors
• CPC Class Codes: C12P 21/00 Preparation of peptides or proteins single cell protein C12N1/00
• CPC Class Codes: Y02A 50/30 Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

Provided herein are compositions and methods useful for the expansion of hematopoietic stem and progenitor cells, such as those that have been genetically modified, for instance, to express a heterologous transgene. In accordance with the composition and methods described herein, hematopoietic stem and progenitor cells may be genetically modified, for example, to express transgenes encoding therapeutic proteins. Genetically modified hematopoietic stem and progenitor cells may be expanded, for instance, by treatment ex vivo with an aryl hydrocarbon receptor antagonist, and may be infused into a patient, such as a patient in need of hematopoietic stem cell transplant therapy. Thus, provided herein are methods for the treatment of various stem cell disorders, including hematopoietic diseases, metabolic disorders, cancers, and autoimmune diseases, among others.

Claims

1. A method of producing an expanded population comprising genetically modified hematopoietic stem or progenitor cells ex vivo, the method comprising:

culturing a population comprising genetically modified hematopoietic stem or progenitor cells ex vivo in a medium comprising an amount of an aryl hydrocarbon receptor antagonist effective to substantially inhibit differentiation of the genetically modified hematopoietic stem or progenitor cells, thereby producing an expanded population comprising genetically modified hematopoietic stem or progenitor cells, wherein;

the genetically modified hematopoietic stem or progenitor cells were produced by (i) disrupting an endogenous gene in a plurality of hematopoietic stem or progenitor cells or (ii) modifying a plurality of hematopoietic stem or progenitor cells to express a polynucleotide introduced into the plurality of hematopoietic stem or progenitor cells.

2. The method of claim 1, wherein prior to (i) disrupting an endogenous gene in the plurality of hematopoietic stem or progenitor cells or (ii) modifying the plurality of hematopoietic stem or progenitor cells to express a polynucleotide introduced into the plurality of hematopoietic stem or progenitor cells, the plurality of hematopoietic stem or progenitor cells is contacted with an aryl hydrocarbon receptor antagonist.

5. The method of claim 1, wherein the disrupting an endogenous gene in a plurality of hematopoietic stem or progenitor cells comprises contacting the hematopoietic stem or progenitor cells with a nuclease that catalyzes cleavage of an endogenous nucleic acid in the hematopoietic stem or progenitor cell or the modifying a plurality of hematopoietic stem or progenitor cells to express a polynucleotide introduced into the plurality of hematopoietic stem or progenitor cells comprises contacting the hematopoietic stem or progenitor cells with a vector containing the polynucleotide.

6. The method of claim 5, wherein the nuclease is selected from the group consisting of a CRISPR-associated protein, a transcription activator-like effector nuclease, a meganuclease and a zinc finger nuclease or the vector is selected from the group consisting of a viral vector and a transposable element.

7. The method of claim 6, wherein the nuclease is caspase 9, the viral vector is selected from the group consisting of adenovirus (Ad), retrovirus, poxvirus, adeno-associated virus, baculovirus, herpes simplex virus, and a vaccinia virus, or the transposable element is selected from the group consisting of a piggy bac transposon and a sleeping beauty transposon.

9. The method of claim 1, wherein upon transplantation of the expanded population comprising genetically modified hematopoietic stem or progenitor cells to a patient, the genetically modified hematopoietic stem or progenitor cells, or progeny thereof, maintain disruption of the endogenous gene or continue to express the polynucleotide for at least 2 days.

24. The method of claim 7, wherein the retrovirus is selected from the group consisting of a lentivirus and a γ

-retrovirus.

35. The method of claim 1, wherein the expanded population and/or the population comprising genetically modified hematopoietic stem or progenitor cells further comprises hematopoietic stem or progenitor cells that are not genetically modified.

36. The method of claim 35, wherein (1) the genetically modified hematopoietic stem or progenitor cells are expanded at a rate that is proportional to the relative quantity of genetically modified hematopoietic stem or progenitor cells present in the population prior to culturing in the medium comprising the aryl hydrocarbon receptor antagonist, (2) the hematopoietic stem or progenitor cells that are not genetically modified do not out-compete the genetically modified hematopoietic stem or progenitor cells for expansion during the culturing in the medium comprising the aryl hydrocarbon receptor antagonist, (3) the hematopoietic stem cells that are genetically modified are expanded more rapidly than the hematopoietic stem or progenitor cells that are not genetically modified and/or (4) upon transplantation of the expanded population comprising genetically modified hematopoietic stem or progenitor cells into a patient, the ratio of the hematopoietic stem or progenitor cells, or progeny thereof, that are genetically modified to the total quantity of hematopoietic stem cells in a sample bone marrow or peripheral blood sample isolate from the patient is at least 75% of the ratio of genetically modified hematopoietic stem or progenitor cells to the total quantity of the hematopoietic stem or progenitor cells present in the expanded population at the time of administration of the cells to the patient.

45. The method of claim 1, wherein the expanded population comprising genetically modified hematopoietic stem or progenitor cells exhibits a higher engraftment potential relative to a population of hematopoietic stem or progenitor cells not cultured in the medium comprising the aryl hydrocarbon receptor antagonist.

46. The method of claim 1, wherein prior to the disrupting an endogenous gene in the plurality of hematopoietic stem or progenitor cells or the modifying a plurality of hematopoietic stem or progenitor cells to express a polynucleotide introduced into the plurality of hematopoietic stem or progenitor cells, the plurality of hematopoietic stem or progenitor cells are mobilized and isolated from a donor.

48. The method of claim 46, wherein the plurality of hematopoietic stem or progenitor cells are mobilized by contacting the plurality of hematopoietic stem or progenitor cells with a mobilizing amount of a CXCR4 antagonist and/or a CXCR2 agonist.

52. The method of claim 1, wherein prior to disrupting an endogenous gene in a plurality of hematopoietic stem or progenitor cells, thereby producing a population comprising genetically modified hematopoietic stem or progenitor cells or prior to modifying a plurality of hematopoietic stem or progenitor cells to express a polynucleotide introduced into the plurality of hematopoietic stem or progenitor cells, thereby producing a population comprising genetically modified hematopoietic stem or progenitor cells, the plurality of hematopoietic stem or progenitor cells is contacted with an aryl hydrocarbon receptor antagonist during a period of time sufficient to induce cell cycling.

56. The method of claim 52, wherein the plurality of hematopoietic stem or progenitor cells are contacted with an agent in an amount sufficient to induce cell cycling during the period of time.

60. A human blood cell preparation comprising the expanded population comprising genetically modified hematopoietic stem or progenitor cells, or progeny thereof, prepared according to the method of claim 1.

61. A method of treating a disorder in a patient, the method comprising producing an expanded population comprising genetically modified hematopoietic stem or progenitor cells in accordance with the method of claim 1 and infusing the resulting hematopoietic stem or progenitor cells into the patient.

86. The method of claim 61, wherein (1) the disorder is selected from the group consisting of a hemoglobinopathy disorder, a myelodysplastic disorder, an immunodeficiency disorder, a metabolic disorder, cancer, an autoimmune disorder, and a neurological disorder (2) the hematopoietic stem or progenitor cells are autologous with respect to the patient, (3) the hematopoietic stem or progenitor cells are allogeneic with respect to the patient, (4) the hematopoietic stem or progenitor cells are HLA-matched with respect to the patient, (5) the hematopoietic stem or progenitor cells, or progeny thereof, maintain hematopoietic stem cell functional potential after two or more days following infusion of the hematopoietic stem or progenitor cells into the patient, (6) the hematopoietic stem or progenitor cells, or progeny thereof, localize to hematopoietic tissue and/or reestablish hematopoiesis following infusion of the hematopoietic stem or progenitor cells into the patient, and/or (7) upon infusion into the patient the hematopoietic stem or progenitor cells give rise to recovery of a population of cells selected from the group consisting of megakaryocytes, thrombocytes, platelets, erythrocytes, mast cells, myeoblasts, basophils, neutrophils, eosinophils, microglia, ganulocytes, monocytes, osteoclasts, antigen-presenting cells, macrophages, dendritic cells, natural killer cells, T-lymphocytes, and B-lymphocytes.

93. A kit comprising a plurality of hematopoietic stem or progenitor cells and a package insert, wherein the package insert instructs a user to perform the method of claim 1.

95. The method of claim 1, wherein the aryl hydrocarbon receptor antagonist is SR-1, Compound 2, or a compound represented by formula (IV) or formula (V)

96. The method of claim 1, wherein the aryl hydrocarbon receptor antagonist is selected from the group consisting of

3-4. -4. (canceled)

8. (canceled)

10-23. -23. (canceled)

25-34. -34. (canceled)

37-44. -44. (canceled)

47. (canceled)

49-51. -51. (canceled)

53-55. -55. (canceled)

57-59. -59. (canceled)

62-85. -85. (canceled)

87-92. -92. (canceled)

94. (canceled)

97-150. -150. (canceled)