Granted Patent
B2
US 12,391,734 · App. 18/584,363 · Granted Aug 19, 2025
PD-1 homing endonuclease variants, compositions, and methods of use
Inventors:
Jasdeep Mann (Seattle, WA); Joel Gay (Seattle, WA); Jordan Jarjour (Seattle, WA); Joy Zhang (Seattle, WA)
Assignee:
REGENERON PHARMACEUTICALS, INC.
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Abstract
The present disclosure provides improved genome editing compositions and methods for editing a PD-1 gene. The disclosure further provides genome edited cells for the prevention, treatment, or amelioration of at least one symptom of, a cancer, an infectious disease, an autoimmune disease, an inflammatory disease, or an immunodeficiency.
Claims (30)
1. A polypeptide comprising an I-OnuI homing endonuclease (HE) variant that cleaves a target site in the human program cell death 1 (PD-1) gene, wherein the I-OnuI HE variant comprises the amino acid sequence set forth in any one of SEQ ID NOs: 11, 12, 60, 62, and 63.
2. The polypeptide of claim 1 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 11.
3. The polypeptide of claim 1 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 12.
4. The polypeptide of claim 1 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 60.
5. The polypeptide of claim 1 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 62.
6. The polypeptide of claim 1 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 63.
7. The polypeptide of claim 1 , wherein the polypeptide binds and cleaves the polynucleotide sequence set forth in SEQ ID NO: 30.
8. The polypeptide of claim 1 , further comprising a TALE DNA binding domain comprising about 9.5 TALE repeat units to about 15.5 TALE repeat units.
9. The polypeptide of claim 8 , wherein the TALE DNA biding domain binds a polynucleotide sequence set forth in SEQ ID NO: 31.
10. A polynucleotide encoding the polypeptide of claim 1 .
11. An mRNA encoding the polypeptide of claim 1 .
12. A vector comprising a polynucleotide encoding the polypeptide of claim 1 .
13. A method of editing a human PD-1 gene in a cell comprising: introducing a polynucleotide encoding a polypeptide into the cell,
wherein the polypeptide comprises an I-OnuI HE variant comprising the amino acid sequence set forth in any one of SEQ ID NOs: 11, 12, 60, 62, and 63,
wherein expression of the polypeptide creates a double strand break at a target site in a human PD-1 gene; and
wherein the cell is a hematopoietic cell.
14. The method of claim 13 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 11.
15. The method of claim 13 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 12.
16. The method of claim 13 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 60.
17. The method of claim 13 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 62.
18. The method of claim 13 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 63.
19. The method of claim 13 , wherein the polypeptide binds and cleaves the polynucleotide sequence set forth in SEQ ID NO: 30.
20. The method of claim 13 , wherein the polypeptide further comprises a TALE DNA binding domain comprising about 9.5 TALE repeat units to about 15.5 TALE repeat units.
21. The method of claim 20 , wherein the TALE DNA biding domain binds a polynucleotide sequence set forth in SEQ ID NO: 31.
22. The method of claim 13 , wherein the double strand break is repaired by non-homologous end joining (NHEJ).
23. The method of claim 13 , comprising introducing a donor repair template into the cell, wherein the donor repair template is incorporated into the human PD-1 gene by homology directed repair (HDR) at the site of the double-strand break.
24. The method of claim 13 , wherein the hematopoietic cell is a T cell; a CD3 + , CD4 + , and/or CD8 + cell; an immune effector cell; a cytotoxic T lymphocytes (CTLs), a tumor infiltrating lymphocytes (TILs) or a helper T cells; and/or a natural killer (NK) cell or natural killer T (NKT) cell.
25. The method of claim 13 , wherein the source of the cell is peripheral blood mononuclear cells, bone marrow, lymph nodes tissue, cord blood, thymus issue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, or tumors.
26. The method of claim 13 , wherein the polynucleotide encoding the polypeptide is an mRNA.
27. The method of claim 23 , wherein the donor repair template encodes a PD-1 gene or portion thereof comprising one or more mutations compared to the wild type PD-1 gene; or wherein the donor repair template encodes one or more of an immunopotency enhancer, an immunosuppressive signal damper, or an engineered antigen receptor.
Assignments (3)
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Jun 25, 2024
From: 2SEVENTY BIO, INC.
To: REGENERON PHARMACEUTICALS, INC.
Reel/Frame 067822/0847 →
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Jun 20, 2024
From: MANN, JASDEEP; GAY, JOEL; JARJOUR, JORDAN; ZHANG, JOY
To: BLUEBIRD BIO, INC.
Reel/Frame 067783/0688 →
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Jun 20, 2024
From: BLUEBIRD BIO, INC.
To: 2SEVENTY BIO, INC.
Reel/Frame 067796/0552 →
Continuity (5)
Continuation
16330039
Provisional Application
62414279
· Oct 28, 2016
Provisional Application
62385079
· Sep 8, 2016
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