Engineered immunoglobulin Fc polypeptides displaying improved complement activation

• US 10,457,737 B2
• Filed: 02/09/2016
• Issued: 10/29/2019
• Est. Priority Date: 02/09/2015
• Status: Active Grant

• Current Assignee: Research & Development Foundation
• Original Assignee: Research & Development Foundation
• Inventors: Georgiou, George, Lee, Chang-Han
• Primary Examiner(s): Dahle, Chun W
• Application Number:
  US15/019,395
• Publication Number:
  US 20160297886A1
• Time in Patent Office: 1,358 Days
• Field of Search: None
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: C07K 16/00 Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies antibodies with enzymatic activity, e.g. abzymes C12N9/0002
• CPC Class Codes: C07K 16/2887 against CD20
• CPC Class Codes: C07K 2317/41 Glycosylation, sialylation, or fucosylation
• CPC Class Codes: C07K 2317/52 Constant or Fc region; Isotype
• CPC Class Codes: C07K 2317/524 CH2 domain
• CPC Class Codes: C07K 2317/72 Increased effector function due to an Fc-modification
• CPC Class Codes: C07K 2317/73 Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
• CPC Class Codes: C07K 2317/732 Antibody-dependent cellular cytotoxicity [ADCC]
• CPC Class Codes: C07K 2317/734 Complement-dependent cytotoxicity [CDC]
• CPC Class Codes: C07K 2317/77 Internalization into the cell
• CPC Class Codes: C07K 2317/90 characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
• CPC Class Codes: C07K 2317/92 Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
• CPC Class Codes: C07K 2317/94 Stability, e.g. half-life, pH, temperature or enzyme-resistance

Abstract

Methods and compositions involving polypeptides having an aglycosylated antibody Fc domain are provided. In certain embodiments, polypeptides have an aglycosylated Fc domain that contains one or more substitutions compared to a native Fc domain. Additionally, some embodiments involve an Fc domain that is binds some Fc receptors but not others. For example, polypeptides are provided with an aglycosylated Fc domain that selectively binds C1q, and optionally activating Fc receptors, but that is significantly reduced for binding to the inhibitory FcγRIIb receptor. Furthermore, methods and compositions are provided for promoting complement dependent cytotoxicity (CDC) using a polypeptide having a modified aglycosylated Fc domain and a second non-Fc binding domain, which can be an antigen binding region of an antibody or a non-antigen binding region. Some embodiments concern antibodies with such polypeptides, which may have the same or a different non-Fc binding domain.

Claims

1. A polypeptide comprising an aglycosylated variant human IgG Fc domain capable of binding human C1q, wherein the Fc domain comprises amino acid substitutions selected from the group consisting of:

(a) K320E and Q386R,

(b) L235K, G236M, G237R, and L351Q,

(c) V308A, S337P, K338Q, K340R, Q342P, R344G, E345Y, and F372L,

(d) M252V, G341A, and L351Q,

(e) M252V, K246N, K322E, R344G, E345Y, and F372L,

(f) M252V, F242L, K338Q, G341A, and E345Y, and

(g) F242L, M252V, K338Q, G341A, and E345Y;

wherein the numbering of the residues in the Fc domain is that of the EU index as in Kabat.

2. The polypeptide of claim 1, wherein the polypeptide does not detectably bind Fcγ

receptors.

3. The polypeptide of claim 1, wherein the polypeptide has specific affinity for binding human C1q and activating Fc receptors.

4. The polypeptide of claim 3, wherein the polypeptide does not detectably bind Fcγ

RIIb.

5. The polypeptide of claim 1, wherein the aglycosylated variant human IgG Fc domain comprises substitutions K320E and Q386R.

6. The polypeptide of claim 5, wherein the aglycosylated variant human IgG Fc domain is Fc801 (SEQ ID NO:

7).

7. The polypeptide of claim 1, wherein the aglycosylated variant human IgG Fc domain comprises substitutions L235K, G236M, G237R, and L351Q.

8. The polypeptide of claim 7, wherein the aglycosylated variant human IgG Fc domain is Fc802 (SEQ ID NO:

10).

9. The polypeptide of claim 3, wherein the aglycosylated variant human IgG Fc domain comprises substitutions V308A, S337P, K338Q, K340R, Q342P, R344G, E345Y, and F372L.

10. The polypeptide of claim 9, wherein the aglycosylated variant human IgG Fc domain is Fc805 (SEQ ID NO:

22).

11. The polypeptide of claim 1, wherein the aglycosylated variant human IgG Fc domain further comprises the substitutions M252V, G341A, and L351Q.

12. The polypeptide of claim 11, wherein the aglycosylated variant human IgG Fc domain is Fc-V18 (SEQ ID NO:

41).

13. The polypeptide of claim 1, wherein the aglycosylated variant human IgG Fc domain further comprises the substitutions K246N, K322E, R344G, E345Y, F372L, and M252V.

14. The polypeptide of claim 13, wherein the aglycosylated variant human IgG Fc domain is Fc-V23 (SEQ ID NO:

43).

15. The polypeptide of claim 1, wherein the aglycosylated variant human IgG Fc domain comprises the substitutions F242L, M252V, K338Q, G341A, and E345Y.

16. The polypeptide of claim 15, wherein the aglycosylated variant human IgG Fc domain is Fc-V24 (SEQ ID NO:

44).