WNT substitute and method for lacrimal gland regeneration

Abstract

Methods and compositions for regenerating lacrimal gland cells for the treatment of dry eye disorders are provided. In particular, the disclosure provides methods and compositions for modulating WNT signaling in a subject by administering an engineered WNT signaling agonist to modulate the regeneration of tear-producing acinar cells for the treatment of tear-deficient dry eye disease. The Specified provides an engineered multivalent bispecific WNT signaling modulator comprising a) at least one first antigen-binding domain that binds to at least one of Lrp6 and / or Lrp5; and b) at least one second antigen-binding domain that binds to at least one FZD receptor.

Claims

1. a. At least one first antigen-binding domain that binds to at least one of Lrp6 and / or Lrp5; and b. At least one second antigen-binding domain that binds to at least one FZD receptor. A modified multivalent bispecific WNT signaling modulator, including a modified multivalent bispecific WNT signaling modulator.

2. The modified WNT signaling modulator according to claim 1, which is a tetravalent bispecific molecule.

3. a. Two light chains, each containing a polypeptide sequence having at least 95% identity with SEQ ID NO: 3; and b. Two heavy chains, each containing a polypeptide sequence having at least 95% identity with SEQ ID NO: 4, SEQ ID NO: 5, or SEQ ID NO:

6. The manipulated WNT signaling modulator according to claim 2, comprising:

4. The manipulated WNT signaling modulator according to claim 3, wherein each of the two light chains comprises a VHH domain containing one or more or more of the amino acid residues N29Q and / or DDD substituted with VHH3-H4 DED.

5. The modified WNT signaling modulator according to claim 3, wherein each heavy chain comprises the amino acid residues CDRH1 of SEQ ID NO: 20, CDRH2 of SEQ ID NO: 21, and CDRH3 of SEQ ID NO:

22.

6. The modified WNT signaling modulator according to any one of claims 2 to 5, wherein the WNT signaling modulator molecule further comprises amino acid modifications including L234A amino acid substitution, L235A amino acid substitution, and P329G amino acid substitution, numbered according to the EU index.

7. The manipulated WNT signaling modulator according to claims 2 to 6, wherein the heavy chain of Sequence ID No. 5 comprises an Fc region mutation having at least three amino acid modifications selected from the group consisting of I253A, H310A, and H435A numbered according to the EU index.

8. The manipulated WNT signaling modulator according to claims 2 to 6, wherein the heavy chain of Sequence ID No. 6 comprises an Fc region mutation having at least three amino acid modifications selected from the group consisting of I253A, H310A, and H435Q numbered according to the EU index.

9. The manipulated WNT signaling modulator according to claim 1, which is a trivalent bispecific molecule.

10. a. A first antigen-binding domain that binds to one of Lrp6 and / or Lrp5; and b. Two second antigen-binding domains that bind to at least two FZD receptors. The manipulated WNT signaling modulator according to claim 9, comprising:

11. The modified WNT signaling modulator according to claim 10, wherein the anti-LRP5 / 6 is an N-terminal "hole" adduct comprising a polypeptide sequence having 95% identity with respect to the combination of SEQ ID NO: 8, SEQ ID NO: 7, and SEQ ID NO:

9.

12. The modified WNT signaling modulator according to claim 10, wherein the anti-LRP5 / 6 is a C-terminal "hole" adduct comprising a polypeptide sequence having 95% identity with respect to the combination of SEQ ID NO: 13, SEQ ID NO: 7, and SEQ ID NO:

9.

13. The manipulated WNT signaling modulator according to claim 10, wherein the anti-LRP5 / 6 is an N-terminal "knob" adduct comprising a polypeptide sequence having 95% identity with respect to the combination of SEQ ID NO: 11, SEQ ID NO: 7, and SEQ ID NO:

12.

14. The manipulated WNT signaling modulator according to claim 10, wherein the anti-LRP5 / 6 is a C-terminal "knob" adduct comprising a polypeptide sequence having 95% identity with respect to the combination of SEQ ID NO: 10, SEQ ID NO: 7, and SEQ ID NO:

12.

15. A nucleic acid comprising a sequence encoding an engineered multivalent bispecific WNT signaling modulator according to any one of claims 1 to 14, wherein the nucleic acid is optionally DNA, cDNA, RNA, mRNA, modified mRNA, or a DNA / RNA hybrid.

16. A vector comprising the nucleic acid described in claim 15.

17. The vector according to claim 16, which is an expression vector comprising a promoter operatively linked to the nucleic acid according to claim 15.

18. A pharmaceutically acceptable diluent, additive, or carrier, a. An operated WNT signaling modulator according to any one of claims 1 to 14; b. The nucleic acid according to any one of claims 15; and / or c. The vector according to any one of claims 16 to 17 A pharmaceutical composition containing the following.

19. A method for regenerating lacrimal gland cells in a subject, The process of administering a WNT signaling modulator to the subject. Methods that include...

20. The method according to claim 19, wherein the subject is a mammal or a human patient.

21. The method according to claim 19, wherein the lacrimal gland cells are acinar cells, progenitor cells, ductal cells, myoepithelial cells, or immune cells.

22. The method according to claim 19, wherein the manipulated WNT signaling modulator is a WNT agonist.

23. The aforementioned modified WNT signaling agonist i. Administered to one or more of the target eyes, either intralacrimally or by eye drops, as necessary; ii. Administered topically, as necessary, into the eyes, ears, nose (into the nasal cavity as necessary), skin (transcutaneously or onto the skin as necessary), mucous membranes, skin, or vagina, or by inhalation; or iii. Administered parenterally, if necessary, by injection (intravenous, intramuscular, subcutaneous, intradermal, intrathecal, intra-arterial, intra-articular, or intraperitoneal administration, if necessary) or by inhalation; or iv. Administered enterally, or, if necessary, orally, sublingually, buccally, or rectally. The method according to claims 19 to 22.

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