Method for inducing cardiac differentiation of pluripotent stem cell

1. A method for inducing cardiac differentiation of a pluripotent stem cell, which comprises the steps of

(1) culturing the pluripotent stem cell in a medium containing one or more WNT signaling activators, and

(2) replacing the medium from step (1) with a medium containing one or more WNT signaling inhibitors wherein a plurality of cardiac differentiated cells are produced,

wherein said one or more WNT signaling inhibitors comprise the compound of Formula (I):

wherein

R 1 to R 5 are each independently a hydrogen atom; a halogen atom; a hydroxyl group; a linear or branched alkoxy group having 1 to 5 carbon atoms; a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; or a group —NR 12 R 13 , wherein R 12 and R 13 are each independently a hydrogen atom, an oxygen atom, or a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; wherein two adjacent groups among R 1 to R 5 may join together to form —O—CH 2 —O—or —O—(CH 2 ) 2 —O—,

R 6 to R 9 are each independently a hydrogen atom; a halogen atom; a hydroxyl group; a linear or branched alkoxy group having 1 to 5 carbon atoms; a linear or branched alkoxy group having 1 to 5 carbon atoms which is substituted with a group —C(O)A, wherein A is a saturated or unsaturated 5- or 6-membered ring which is unsubstituted or substituted with a linear or branched alkyl group having 1 to 5 carbon atoms and the ring may contain 1 or 2 atoms independently selected from a nitrogen atom, an oxygen atom and a sulfur atom; a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; or a group —NR 12 R 13 , wherein R 12 and R 13 are each independently a hydrogen atom, an oxygen atom, or a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; wherein two adjacent groups among R 6 to R 9 may join together to form —O—CH 2 —O—or —O—(CH 2 ) 2 —O—,

R 10 to R 11 are each independently a hydrogen atom; or a linear or branched alkyl group having 1 to 5 carbon atoms,

X is a sulfur atom, and

n is an integer of 0 to 6,

or a salt thereof.

2. The method of claim 1 , wherein said compound of Formula (I) is selected from the group consisting of

or a salt thereof.

3. The method of claim 2 , wherein the one or more WNT signaling inhibitors comprise

or a salt thereof.

4. The method of claim 1 , wherein the one or more WNT signaling inhibitors further comprise a WNT signaling inhibitor selected from XAV939 and IWP-2.

5. The method of claim 1 , wherein the one or more WNT signaling activators comprise a GSK3β inhibitor.

6. The method of claim 5 , wherein the GSK3β inhibitor is selected from BIO and CHIR99021.

7. The method of claim 1 , wherein the medium in the step (1) and the medium in the step (2) do not comprise serum.

8. The method of claim 1 , wherein the medium in the step (1) and the medium in the step (2) do not comprise a cytokine.

9. The method of claim 1 , wherein the medium in the step (1) and the medium in the step (2) comprise albumin.

10. The method of claim 1 , wherein the medium in the step (1) and the medium in the step (2) do not comprise a protein other than albumin.

11. The method of claim 1 , wherein the two culturing steps are conducted in the absence of a xenogeneic component.

12. The method of claim 1 , wherein the pluripotent stem cell is a monkey or human pluripotent stem cell.

13. The method of claim 1 , wherein the differentiated cells include a cardiomyocyte.

14. A kit for promoting cardiac differentiation comprising one or more WNT signaling activators and/or one or more WNT signaling inhibitors, wherein the kit is used for the method of claim 1 ,

wherein said one or more WNT signaling inhibitors comprise the compound of Formula (I):

wherein

R 1 to R 5 are each independently a hydrogen atom; a halogen atom; a hydroxyl group; a linear or branched alkoxy group having 1 to 5 carbon atoms; a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; or a group —NR 12 R 13 , wherein R 12 , and R 13 are each independently a hydrogen atom, an oxygen atom, or a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; wherein two adjacent groups among R 1 to R 5 may join together to form —O—CH 2 —O—or —O—(CH 2 ) 2 —O—,

R 6 to R 9 are each independently a hydrogen atom; a halogen atom; a hydroxyl group; a linear or branched alkoxy group having 1 to 5 carbon atoms; a linear or branched alkoxy group having 1 to 5 carbon atoms which is substituted with a group —C(O)A, wherein A is a saturated or unsaturated 5- or 6-membered ring which is unsubstituted or substituted with a linear or branched alkyl group having 1 to 5 carbon atoms and the ring may contain 1 or 2 atoms independently selected from a nitrogen atom, an oxygen atom and a sulfur atom; a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; or a group —NR 12 R 13 , wherein R 12 , and R 13 are each independently a hydrogen atom, an oxygen atom, or a linear or branched alkyl group having 1 to 5 carbon atoms which is unsubstituted or substituted with a halogen atom; wherein two adjacent groups among R 6 to R 9 may join together to form —O—CH 2 —O—or —O—(CH 2 ) 2 —O—,

R 10 to R 11 are each independently a hydrogen atom; or a linear or branched alkyl group having 1 to 5 carbon atoms,

X is a sulfur atom, and

n is an integer of 0 to 6,

or a salt thereof.

15. The method of claim 1 , wherein

R 1 , R 4 , R 5 , R 6 and R 9 each are a hydrogen atom,

R 2 and R 3 are a methoxy group, an ethoxy group or a propoxy group,

R 10 and R 11 are a hydrogen atom, and

n is an integer of 0 to 4.

16. The method of claim 15 , wherein

R 7 is a halogen atom, and

R 8 is a hydrogen atom.

17. The method of claim 16 , wherein n is 2 or 3.

18. The method of claim 17 , wherein R 2 and R 3 are a methoxy group.