Gene correction of SCID-related genes in hematopoietic stem and progenitor cells

• US 9,833,479 B2
• Filed: 03/14/2017
• Issued: 12/05/2017
• Est. Priority Date: 07/30/2014
• Status: Active Grant

• Current Assignee: Sangamo Therapeutics, Inc.
• Original Assignee: Sangamo Therapeutics, Inc.
• Inventors: Conway, Anthony, Cost, Gregory J., Holmes, Michael C., Urnov, Fyodor
• Primary Examiner(s): BURKHART, MICHAEL D
• Application Number:
  US15/458,258
• Publication Number:
  US 20170189450A1
• Time in Patent Office: 266 Days
• Field of Search: None
• CPC Class Codes: A61K 2035/124 the cells being hematopoietic, bone marrow derived or blood cells
• 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: A61K 35/545 Embryonic stem cells; Pluripotent stem cells; Induced pluripotent stem cells; Uncharacterised stem cells
• CPC Class Codes: A61K 38/1709 from mammals
• CPC Class Codes: A61K 38/1793 for cytokines; for lymphokines; for interferons
• CPC Class Codes: A61K 48/005 characterised by an aspect of the 'active' part of the composition delivered, i.e. the nucleic acid delivered
• CPC Class Codes: C07K 14/4705 stimulating, promoting or activating activity
• CPC Class Codes: C07K 14/7155 for interleukins [IL]
• CPC Class Codes: C12N 15/907 in mammalian cells
• CPC Class Codes: C12N 9/22 Ribonucleases RNAses, DNAses catalytic nucleic acids C12N15/113

Abstract

The present disclosure is in the field of genome engineering, particularly targeted integration of a functional SCID-related genes (e.g., IL2RG, RAG1 and/or RAG2 gene) into the genome of a cell for provision of proteins lacking or deficient in SCID.

Claims

1. A host cell comprising an exogenous sequence integrated into intron 1 of an endogenous IL2RG gene or intron 1 or 2 of an endogenous RAG gene using a nuclease comprising a zinc finger protein, a TAL-effector domain or a single guide RNA (sgRNA) DNA-binding domain that binds to a sequence comprising a target site as shown in any of SEQ ID NOs:

47-60 or 81-83 or a target site bound by any SEQ ID NO;

85-95.

2. The host cell of claim 1, wherein the nuclease comprises the zinc finger protein that comprises recognition helix regions as shown in a single row of Table 1.

3. The host cell of claim 1, wherein the sgRNA comprises a DNA-binding guide RNA as shown in Table 5.

4. The host cell of claim 1, wherein the TALE-effector domain comprises hypervariable diresidues (RVDs) as shown in in a single row of Table 3.

5. The host cell of claim 1, wherein the exogenous sequence is selected from the group consisting of:

a sequence encoding an IL2RG polypeptide integrated into intron 1 of an endogenous IL2RG gene;

a sequence encoding a RAG polypeptide integrated into intron 1 or 2 of an endogenous RAG gene; and

combinations thereof.

6. The host cell of claim 5, wherein the exogenous sequence comprises a cDNA selected from the group consisting of a sequence comprising exons 2 through 8 of a wild type IL2RG gene;

a sequence comprising a full-length IL2RG gene;

a sequence comprising exon 3 of a wild type RAG gene and a sequence comprising a full-length RAG gene.

7. The host cell of claim 1, wherein the cell is a hematopoietic stem cell or an induced pluripotent stem cell (iPSC).

8. A method of treating or preventing SCID or Omenn Syndrome in a subject, the method comprising administering a host cell according to claim 1 to the subject.

9. The method of claim 8, wherein the cell is a hematopoietic stem cell.