Generation of deoxyadenosine triphosphate donor cells and uses thereof

Patent metadata

Field	Value
Publication number	US-2025145957-A1
Application number	US-202418807485-A
Country	US
Kind code	A1
Filing date	Aug 16, 2024
Priority date	Feb 18, 2022
Publication date	May 8, 2025
Grant date	—

Abstract

Provided herein are compositions and methods directed toward the discovery of improved methods for generating deoxyATP (dATP) in cells that can be delivered to a graft site in the heart to enhance cardiac function.

First claim

1 . A dATP donor cell comprising a nucleic acid sequence encoding a ribonucleotide reductase expression cassette operably linked to a muscle-specific promoter.

2 . The dATP donor cell of claim 1 , wherein the cell is an induced pluripotent stem cell or an embryonic stem cell; and/or wherein the cell is a human cell.

3 . (canceled)

4 . The dATP donor cell of claim 2 , wherein the cell is capable of forming gap junctions with a cardiomyocyte, and the cell comprises a cardiomyocyte, a cardiac progenitor cell, a fibroblast, a mesenchymal cell, a smooth muscle cell, an endothelial cell, or a hematopoietic cell.

5 . (canceled)

6 . The dATP donor cell of claim 4 , wherein the cell is a cardiac progenitor cell or a cardiomyocyte, and wherein the cardiac progenitor cell or cardiomyocyte are in vitro-differentiated.

7 . (canceled)

8 . The dATP donor cell of claim 1 , wherein the ribonucleotide reductase expression cassette comprises a sequence encoding a ribonucleotide reductase catalytic subunit (RRM1) and a ribonucleotide reductase regulatory subunit (RRM2).

9 . The dATP donor cell of claim 8 , wherein the RRM2 subunit comprises a double mutation and has increased stability compared to the wild-type RRM2 subunit.

10 . The dATP donor cell of claim 9 , wherein the double mutation comprises 30AAA/49ARA mutations in SEQ ID NO. 26.

11 . The dATP donor cell of claim 8 , wherein the sequence of the RRM1 subunit is SEQ ID NO: 1, and the sequence of the RRM2 subunit comprises SEQ ID NO: 3.

12 . The dATP donor cell of claim 1 , wherein the muscle-specific promoter is CK8m or CK8e.

13 . (canceled)

14 . The dATP donor cell of claim 12 , wherein the muscle-specific promoter is a cardiac muscle-specific promoter selected from the group consisting of: MSEC-320, MSEC-455c, MSEC-455a, MSEC-571, MSEC-725a, MSEC-875, and CK8m.

15 . (canceled)

16 . A dATP donor cell comprising a nucleic acid sequence encoding a ribonucleotide reductase expression cassette operably linked to a constitutively active promoter.

17 . The dATP donor cell of claim 16 , wherein the constitutively active promoter is selected from CMV, Thymidine Kinase, SV40, EF1A, and CAG promoters.

18 . The dATP donor cell of claim 16 , wherein the cell is an induced pluripotent stem cell or an embryonic stem cell.

19 . (canceled)

20 . The dATP donor cell of claim 18 wherein the cell is capable of forming gap junctions with a cardiomyocyte, and the cell comprises a cardiomyocyte, a cardiac progenitor cell, a fibroblast, a mesenchymal cell, a smooth muscle cell, an endothelial cell, or a hematopoietic cell.

21 . The dATP donor cell of claim 20 , wherein the cell is a cardiac progenitor cell or a cardiomyocyte, and wherein the cardiac progenitor cell or cardiomyocyte are in vitro-differentiated.

22 . (canceled)

23 . The dATP donor cell of claim 16 , wherein the cell is a human cell.

24 . The dATP donor cell of claim 16 , wherein the ribonucleotide reductase expression cassette comprises sequence encoding a ribonucleotide reductase catalytic subunit (RRM1) and a ribonucleotide reductase regulatory subunit (RRM2).

25 . The dATP donor cell of claim 24 , wherein the RRM2 subunit comprises a double mutation and has increased stability compared to the wild-type RRM2 subunit.

26 . The dATP donor cell of claim 25 , wherein the double mutation comprises 30AAA/49ARA mutations in SEQ ID NO. 26.

27 . The dATP donor cell of claim 24 , wherein the sequence of the RRM1 subunit is SEQ ID NO: 1, and the sequence of the RRM2 subunit comprises SEQ ID NO: 3.

28 - 74 . (canceled)

Assignees

• Univ Washington

Inventors

• Regnier Michael
• Murry Charles E
• Hauschka Stephen D

Classifications

• C12Y117/04001

  Ribonucleoside-diphosphate reductase (1.17.4.1) · CPC title

• C12N2830/15

  chimeric enhancer/promoter combination · CPC title

• C12N2506/45

  from artificially induced pluripotent stem cells · CPC title

• C12N15/85

  for animal cells · CPC title

• C12N9/0093 Primary

  acting on CH or CH2 groups (1.17) · CPC title