# US 11,440,971 B2 · Fc-region variants with modified FcRn-binding and methods of use

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- Number: US 11,440,971 B2
- Document ID: US11440971B2
- Status: Granted
- Kind: B2
- Application number: 16/868,972
- Filed: 2020-05-07
- Published: 2020-08-27
- Granted: 2022-09-13
- Earliest claimed: 2014-11-06
- Assignees: Hoffmann-La Roche Inc.
- Applicants: Hoffmann-La Roche Inc.
- Inventors: Ekkehard Moessner; Tilman Schlothauer
- Web page: https://patentgrep.com/patent/US11440971B2

## CPC

- C07K 16/468: {Immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies}
- A61P 9/00: Drugs for disorders of the cardiovascular system
- A61P 27/02: Ophthalmic agents
- C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
- C07K 2317/31: multispecific
- C07K 2317/35: Valency
- C07K 2317/52: Constant or Fc region; Isotype
- C07K 2317/526: CH3 domain
- C07K 2317/528: CH4 domain
- C07K 2317/71: Decreased effector function due to an Fc-modification
- C07K 2317/72: Increased effector function due to an Fc-modification
- C07K 2317/94: Stability, e.g. half-life, pH, temperature or enzyme-resistance
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS

## Abstract

Herein is reported an IgG class Fc-region comprising a first variant Fc-region polypeptide and a second variant Fc-region polypeptide, wherein a) the first variant Fc-region polypeptide is derived from a first parent IgG class Fc-region polypeptide and the second variant Fc-region polypeptide is derived from a second parent IgG class Fc-region polypeptide, whereby the first parent IgG class Fc-region polypeptide is identical to or different from the second parent IgG class Fc-region polypeptide, and b) the first variant Fc-region polypeptide differs from the second variant Fc-region polypeptide in one or more amino acid residues other than those amino acid residues in which the first parent IgG class Fc-region polypeptide differs from the second parent IgG class Fc-region polypeptide, and c) the IgG class Fc-region comprising the first variant Fc-region polypeptide and the second variant Fc-region polypeptide has an affinity to a human Fc-receptor that is different than that of an IgG class Fc-region comprising the first parent IgG class Fc-region polypeptide of a) and the second parent IgG class Fc-region polypeptide of a), wherein either the first Fc-region polypeptide or the second Fc-region polypeptide or both Fc-region polypeptides comprise independently of each other one of the following mutations or combination of mutations: T307H, or Q311H, or E430H, or N434H, or T307H and Q311H, or T307H and E430H, or T307H and N434A, or T307H and N434H, or T307Q and Q311H, or T307Q and E430H, or T307Q and N434H, or T307H and Q311H and E430H and N434A, or T307H and Q311H and E430H and N434H, or T307H and Q311H and E430H and N434Y, or T307Q and Q311H and E430H and N434A, or T307Q and Q311H and E430H and N434H, or T307Q and Q311H and E430H and N434Y, or T307Q and V308P and N434Y and Y436H, or T307H and M252Y and S254T and T256E, or T307Q and M252Y and S254T and T256E, or Q311H and M252Y and S254T and T256E, or E430 H and M252Y and S254T and T256E, or N434H and M252Y and S254T and T256E, or T307H and Q311H and M252Y and S254T and T256E, or T307H and E430H and M252Y and S254T and T256E, or T307H and N434A and M252Y and S254T and T256E, or T307H and N434H and M252Y and S254T and T256E, or T307Q and Q311H and M252Y and S254T and T256E, or T307Q and E430H and M252Y and S254T and T256E, or T307Q and N434H and M252Y and S254T and T256E, or T307H and Q311H and E430H and N434A and M252Y and S254T and T256E, or T307H and Q311H and E430H and N434H and M252Y and S254T and T256E, or T307H and Q311H and E430H and N434Y and M252Y and S254T and T256E, or T307Q and Q311H and E430H and N434A and M252Y and S254T and T256E, or T307Q and Q311H and E430H and N434H and M252Y and S254T and T256E, or T307Q and Q311H and E430H and N434Y and M252Y and S254T and T256E, or T307Q and V308P and N434Y and Y436H and M252Y and S254T and T256E.

## Claims

### Claim 1

An IgG class Fc-region comprising a first variant Fc-region polypeptide and a second variant Fc-region polypeptide,
wherein:
a) the first variant Fc-region polypeptide is derived from a first parent IgG class Fc-region polypeptide and the second variant Fc-region polypeptide is derived from a second parent IgG class Fc-region polypeptide, whereby the first parent IgG class Fc-region polypeptide is identical to or different from the second parent IgG class Fc-region polypeptide, and
b) the first variant Fc-region polypeptide differs from the second variant Fc-region polypeptide in one or more amino acid residues other than those amino acid residues in which the first parent IgG class Fc-region polypeptide differs from the second parent IgG class Fc-region polypeptide, and
c) the IgG class Fc-region comprising the first variant Fc-region polypeptide and the second variant Fc-region polypeptide has an affinity to a human neonatal Fc-receptor that is different than that of an IgG class Fc-region comprising the first parent IgG class Fc-region polypeptide of a) and the second parent IgG class Fc-region polypeptide of a),
wherein either the first variant Fc-region polypeptide or the second variant Fc-region polypeptide or both variant Fc-region polypeptides comprise independently of each other T307Q and V308P and N434Y and Y436H.

### Claim 2 (depends on 1)

The IgG class Fc-region according to claim 1, wherein the amino acid residues in which the first parent IgG class Fc-region polypeptide differs from the second parent IgG class Fc-region polypeptide promote the formation of a heterodimeric IgG class Fc-region.

### Claim 3 (depends on 1)

The IgG class Fc-region according to claim 1, wherein:
i) the first parent IgG class Fc-region polypeptide is selected from the group consisting of: a human IgG1 Fc-region polypeptide, a human IgG2 Fc-region polypeptide, a human IgG3 Fc-region polypeptide, a human IgG4 Fc-region polypeptide, a human IgG1 Fc-region polypeptide comprising the mutations L234A and L235A, a human IgG1 Fc-region polypeptide comprising the mutations Y349C, T366S, L368A, and Y407V, a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, Y349C, T366S, L368A, and Y407V, a human IgG1 Fc-region polypeptide comprising the mutation P329G, a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, and P329G, a human IgG1 Fc-region polypeptide comprising the mutations P329G, Y349C, T366S, L368A, and Y407V, a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, P329G, Y349C, T366S, L368A, and Y407V, a human IgG4 Fc-region polypeptide comprising the mutations S228P and L235E, a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, and P329G, a human IgG4 Fc-region polypeptide comprising the mutations Y349C, T366S, L368A, and Y407V, a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, Y349C, T366S, L368A, and Y407V, a human IgG4 Fc-region polypeptide comprising the mutation P329G, a human IgG4 Fc-region polypeptide comprising the mutations P329G, Y349C, T366S, L368A, and Y407V, a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, P329G, Y349C, T366S, L368A, and Y407V, a human IgG1, IgG2 or IgG4 comprising the mutation K392D, and a human IgG3 comprising the mutation N392D (Kabat EU index numbering system);
and
ii) the second parent IgG class Fc-region polypeptide is selected from the group consisting of: a human IgG1 Fc-region polypeptide, a human IgG2 Fc-region polypeptide, a human IgG3 Fc-region polypeptide, a human IgG4 Fc-region polypeptide, a human IgG1 Fc-region polypeptide comprising the mutations L234A and L235A, a human IgG1 Fc-region polypeptide comprising the mutations S354C and T366W, a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, S354C, and T366W, a human IgG1 Fc-region polypeptide comprising the mutation P329G, a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, and P329G, a human IgG1 Fc-region polypeptide comprising the mutations P329G, S354C, and T366W, a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, P329G, S354C, and T366W, a human IgG4 Fc-region polypeptide comprising the mutations S228P and L235E, a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, and P329G, a human IgG4 Fc-region polypeptide comprising the mutations S354C and T366W, a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, S354C, and T366W, a human IgG4 Fc-region polypeptide comprising the mutation P329G, a human IgG4 Fc-region polypeptide comprising the mutations P329G, S354C, and T366W, a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, P329G, S354C, and T366W, a human IgG1 comprising one or more of the mutations D399K, D356K, and E357K, and a human IgG2, IgG3 or IgG4 comprising one or more of the mutations D399K, E356K, and E357K (Kabat EU index numbering system).

### Claim 4 (depends on 1)

The IgG class Fc-region according to claim 1, wherein:
i) the first parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide and the second parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide; or
ii) the first parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A and L235A, and the second parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A and L235A (Kabat EU index numbering system); or
iii) the first parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, and P329G, and the second parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, and P329G (Kabat EU index numbering system); or
iv) the first parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, S354C, and T366W, and the second parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, Y349C, T366S, L368A, and Y407V (Kabat EU index numbering system); or
v) the first parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, P329G, S354C, and T366W, and the second parent IgG class Fc-region polypeptide is a human IgG1 Fc-region polypeptide comprising the mutations L234A, L235A, P329G, Y349C, T366S, L368A, and Y407V (Kabat EU index numbering system); or
vi) the first parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide and the second parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide; or
vii) the first parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P and L235E, and the second parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P and L235E (Kabat EU index numbering system); or
viii) the first parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, and P329G, and the second parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, and P329G (Kabat EU index numbering system); or
ix) the first parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, S354C, and T366W, and the second parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, Y349C, T366S, L368A, and Y407V (Kabat EU index numbering system); or
x) the first parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, P329G, S354C, and T366W, and the second parent IgG class Fc-region polypeptide is a human IgG4 Fc-region polypeptide comprising the mutations S228P, L235E, P329G, Y349C, T366S, L368A, and Y407V (Kabat EU index numbering system); or
xi) the first parent IgG class Fc-region polypeptide is: (a) a human IgG1, IgG2, or IgG4 Fc-region polypeptide comprising the mutation K392D, or (b) a human IgG3 Fc-region polypeptide comprising the mutation N392D, and the second parent IgG class Fc-region polypeptide is: (c) a human IgG1 Fc-region polypeptide comprising one or more of the mutations D399K, D356K, and E357K or (d) a human IgG2, IgG3 or IgG4 Fc-region polypeptide comprising one or more of the mutations D399K, E356K, and E357K (Kabat EU index numbering system).

### Claim 5 (depends on 1)

An antibody comprising the IgG class Fc-region according to claim 1.

### Claim 6 (depends on 5)

The antibody according to claim 5, wherein the antibody is a monoclonal antibody.

### Claim 7 (depends on 5)

The antibody according to claim 5, wherein the antibody is a human, humanized, or chimeric antibody.

### Claim 8 (depends on 5)

The antibody according to claim 5, wherein the antibody is a bispecific antibody.

### Claim 9 (depends on 5)

The antibody according to claim 5, wherein the antibody is a bivalent antibody.

### Claim 10 (depends on 5)

A pharmaceutical formulation comprising the antibody according to claim 5.

### Claim 11 (depends on 1)

A method for making an IgG class Fc-region, the method comprising:
a) cultivating a mammalian cell comprising one or more nucleic acids encoding the first and second variant Fc-region polypeptides according to claim 1; and
b) recovering the IgG class Fc-region from the cultivation medium.

### Claim 12 (depends on 11)

The method of claim 11, wherein the mammalian cell is a CHO cell.