Engineered transferrin receptor binding polypeptides

Patent metadata

Field	Value
Publication number	US-11795232-B2
Application number	US-201916543332-A
Country	US
Kind code	B2
Filing date	Aug 16, 2019
Priority date	Feb 17, 2017
Publication date	Oct 24, 2023
Grant date	Oct 24, 2023

Abstract

Provided herein are polypeptides that bind to a transferrin receptor, methods of generating such polypeptides, and methods of using the polypeptides to target a composition to a transferrin receptor-expressing cell.

First claim

What is claimed is:

1. A polypeptide comprising a CH3 domain that specifically binds to a transferrin receptor, wherein the CH3 domain comprises five, six, seven, or eight substitutions in a set of amino acid positions comprising 118, 119, 120, 122, 210, 211, 212, and 213; and (i) wherein the substitutions and the positions are determined with reference to amino acids 114-220 of SEQ ID NO:1 and (ii) the CH3 domain has at least 80% identity to amino acids 114-220 of any one of SEQ ID NOS:30-46.

2. The polypeptide of claim 1 , wherein the CH3 domain comprises Gly at position 210; Phe at position 211; and/or Asp at position 213.

3. The polypeptide of claim 1 , wherein the CH3 domain comprises at least one position selected from the following: position 118 is Phe or Ile; position 119 is Asp, Glu, Gly, Ala, or Lys; position 120 is Tyr, Met, Leu, Ile, or Asp; position 122 is Thr or Ala; position 210 is Gly; position 211 is Phe; position 212 is His, Tyr, Ser, or Phe; and position 213 is Asp.

4. The polypeptide of claim 1 , wherein the CH3 domain comprises Gly at position 210 and Phe at position 211.

5. The polypeptide of claim 1 , wherein the CH3 domain comprises Gly at position 210 and Asp at position 213.

6. The polypeptide of claim 1 , wherein the CH3 domain comprises Phe at position 211 and Asp at position 213.

7. The polypeptide of claim 1 , wherein the CH3 domain comprises Gly at position 210; Phe at position 211; and Asp at position 213.

8. The polypeptide of claim 1 , wherein the CH3 domain comprises two, three, four, five, six, seven, or eight positions selected from the following: position 118 is Phe or Ile; position 119 is Asp, Glu, Gly, Ala, or Lys; position 120 is Tyr, Met, Leu, Ile, or Asp; position 122 is Thr or Ala; position 210 is Gly; position 211 is Phe; position 212 is His, Tyr, Ser, or Phe; and position 213 is Asp.

9. The polypeptide of claim 1 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of any one of SEQ ID NOS:30-46.

10. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:30 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:30.

11. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:31 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:31.

12. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:32 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:32.

13. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:33 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:33.

14. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:34 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:34.

15. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:35 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:35.

16. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:36 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:36.

17. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:37 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:37.

18. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:38 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:38.

19. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:39 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:39.

20. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:40 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:40.

21. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:41 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:41.

22. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:42 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:42.

23. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:43 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:43.

24. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:44 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:44.

25. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:45 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:45.

26. The polypeptide of claim 9 , wherein the CH3 domain comprises amino acids 118-122 and amino acids 210-213 of SEQ ID NO:46 and has at least 90% identity to amino acids 114-220 of SEQ ID NO:46.

27. The polypeptide of claim 1 , wherein the CH3 domain comprises amino acids 118-213 of any one of SEQ ID NOS:30-46.

28. The polypeptide of claim 1 , wherein the polypeptide comprises the sequence of any one of SEQ ID NOS:30-46 without the first three amino acids “PCP” at the amino-terminal end.

29. The polypeptide of claim 28 , wherein the polypeptide comprises the sequence of any one of SEQ ID NOS:30-46.

30. The polypeptide of claim 1 , wherein the CH3 domain further comprises (i) a Trp at position 139 or (ii) a Ser at position 139, an Ala at position 141, and a Val at position 180, wherein the amino acid positions are determined with reference to SEQ ID NO: 1.

31. The polypeptide of claim 1 , wherein the polypeptide is joined to a CH2 domain.

32. The polypeptide of claim 31 , wherein the CH2 domain contains one or both of the following sets of modifications with reference to the amino acid sequence of SEQ ID NO:1:
(a) Ala at position 7 and at position 8; and
(b) Tyr at position 25, Thr at position 27, and Glu at position 29.

33. The polypeptide of claim 32 , wherein set (a) further comprises Gly at position 102.

34. The polypeptide of claim 31 , wherein the polypeptide is further joined to a Fab.

35. A method for engineering a CH3 domain to specifically bind to a transferrin receptor, the method comprising:
(a) engineering a polynucleotide that encodes the CH3 domain to have at least five amino acid substitutions at a set of amino acid positions comprising 118, 119, 120, 122, 210, 211, 212, and 213,
wherein the substitutions and positions are determined with reference to amino acids 114-220 of SEQ ID NO:1;
(b) expressing a polypeptide comprising the engineered CH3 domain; and
(c) determining whether the engineered CH3 domain binds to the transferrin receptor.

36. The method of claim 35 , wherein the polypeptide comprising the engineered CH3 domain is expressed as a so

Assignees

• Denali Therapeutics Inc

Inventors

• Chen Xiaocheng
• Dennis Mark S
• Kariolis Mihalis
• Silverman Adam P
• Srivastava Ankita
• Watts Ryan J
• Wells Robert C
• Zuchero Joy Yu

Classifications

• C07K16/2881 Primary

  against CD71 · CPC title

• C07K16/005

  constructed by phage libraries · CPC title

• C07K2317/35

  Valency · CPC title

• C07K2317/524

  CH2 domain · CPC title

• C07K2317/526

  CH3 domain · CPC title