# US 12,454,565 B2 · PD-L1 analog fusion proteins for antigen specific immunotherapy and methods of use

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- Number: US 12,454,565 B2
- Document ID: US12454565B2
- Status: Granted
- Kind: B2
- Application number: 19/071,424
- Filed: 2025-03-05
- Published: 2025-09-25
- Granted: 2025-10-28
- Earliest claimed: 2024-03-21
- Assignees: Akston Biosciences Corporation
- Applicants: Akston Biosciences Corporation
- Inventors: Todd C. Zion; Thomas M. Lancaster
- Web page: https://patentgrep.com/patent/US12454565B2

## CPC

- C07K 14/70532: {B7 molecules, e.g. CD80, CD86}
- A61K 39/00115: {Apoptosis related proteins, e.g. survivin or livin}
- A61K 39/00: Medicinal preparations containing antigens or antibodies (materials for immunoassay G01N33/53)
- A61K 2039/6056: {Antibodies}
- C07K 2319/30: Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
- 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)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)

## Abstract

The present disclosure provides recombinantly manufactured fusion proteins comprising a Programmed Death-Ligand 1 (PD-L1) protein fragment or an analog thereof linked to a canine Fc fragment. Embodiments include the administration of the fusion proteins to patients as a treatment for cancers, tumors or other diseases associated with expression of the PD-L1 protein in dogs. Exemplary Fc fusion proteins and pharmaceutical formulations of exemplary Fc fusion proteins are provided, in addition to methods of use and preparation.

## Claims

### Claim 1

A fusion protein comprising a PD-L1 analog and an Fc fragment, wherein the PD-L1 analog and the Fc fragment are connected by a peptide linker, wherein the PD-L1 analog consists of the sequence:
(SEQ ID NO: 23)
LIGYGGADYKRITLKVHAPYRNISQRISVDPVTSEHELMCQAEGYPEAE
VIWTSSDHRVLSGKTTITNSNREEKLFNVTSTLNINATANEIFYCTFQR
SGPEENNTAELVIPERLPVPASERTHFMILGPF,
and
wherein the Fc fragment comprises the sequence:
(SEQ ID NO: 1)
DCPKCPAPEMLGGPSVFIFPPKPKDTLLIARTPEVTCVVVDLDPEDPEV
QISWFVDGKQMQTAKTQPREEQFNGTYRVVSVLPIGHQDWLKGKQFTCK
VNNKALPSPIERTISKARGQAHQPSVYVLPPSREELSKNTVSLTCLIKD
FFPPDIDVEWQSNGQQEPESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQ
RGDTFICAVMHEALHNHYTQESLSHSPG.

### Claim 2 (depends on 1)

The fusion protein of claim 1, wherein the PD-L1 analog and the Fc fragment are connected by the peptide linker comprising the sequence:
(SEQ ID NO: 6)
GGGGQGGGSGGQGGGGG.

### Claim 3

A fusion protein comprising a PD-L1 analog consisting of SEQ ID NO: 23 and an Fc fragment, wherein the PD-L1 analog and the Fc fragment are connected by a peptide linker, wherein the fusion protein comprises the sequence:
(SEQ ID NO: 24)
LIGYGGADYKRITLKVHAPYRNISQRISVDPVTSEHELMCQAEGYPEAE
VIWTSSDHRVLSGKTTITNSNREEKLFNVTSTLNINATANEIFYCTFQR
SGPEENNTAELVIPERLPVPASERTHFMILGPFGGGGQGGGSGGQGGGG
GDCPKCPAPEMLGGPSVFIFPPKPKDTLLIARTPEVTCVVVDLDPEDPE
VQISWFVDGKQMQTAKTQPREEQFNGTYRVVSVLPIGHQDWLKGKQFTC
KVNNKALPSPIERTISKARGQAHQPSVYVLPPSREELSKNTVSLTCLIK
DFFPPDIDVEWQSNGQQEPESKYRTTPPQLDEDGSYFLYSKLSVDKSRW
QRGDTFICAVMHEALHNHYTQESLSHSPG.

### Claim 4 (depends on 1)

The fusion protein of claim 1, wherein the fusion protein is a homodimer comprising two identical monomers bound together via one or more disulfide bonds.

### Claim 5 (depends on 1)

The fusion protein of claim 1, wherein the Fc fragment is glycosylated.

### Claim 6 (depends on 1)

An immunogenic composition comprising a fusion protein according to claim 1 and a pharmaceutically acceptable carrier.

### Claim 7 (depends on 6)

The immunogenic composition of claim 6, further comprising an adjuvant.

### Claim 8

A fusion protein comprising the sequence of SEQ ID NO: 24 or pharmaceutical composition thereof for use in treatment for cancers or tumors in dogs.

### Claim 9 (depends on 8)

A treatment method comprising administering via injection the fusion protein or pharmaceutical composition of claim 8 to a dog in need thereof, wherein duration of response to dose levels varying from 1 μg to 100 μg of the fusion protein demonstrates increased anti-PD-L1 protein antibody titers at all dose levels after 1, 2, and 3 doses up to at least 56 days post injection.

### Claim 10 (depends on 9)

The method of claim 9, wherein said fusion protein or pharmaceutical composition thereof is administered subcutaneously or intramuscularly.

### Claim 11 (depends on 1)

A method of treating cancers or tumors in a dog, the method comprising administering a fusion protein according to claim 1 or pharmaceutical composition thereof to a dog in need thereof.

### Claim 12 (depends on 11)

The method of claim 11, wherein said fusion protein or pharmaceutical composition thereof is administered via injection.

### Claim 13 (depends on 12)

The method of claim 12, wherein said fusion protein or pharmaceutical composition thereof is administered subcutaneously or intramuscularly.

### Claim 14 (depends on 11)

The method of claim 11, wherein said fusion protein or pharmaceutical composition thereof is administered as a priming vaccine, the method further comprising administering a booster of said fusion protein or pharmaceutical composition thereof 7 to 14 days after administering said priming vaccine.

### Claim 15 (depends on 11)

The method of claim 11, wherein said dog is antibody-positive to PD-L1 before said administering step, wherein said fusion protein or pharmaceutical composition thereof is administered as a booster vaccine to said dog.

### Claim 16 (depends on 3)

A method of treating cancers or tumors in a dog, the method comprising administering a fusion protein according to claim 3 or pharmaceutical composition thereof to a dog in need thereof.

### Claim 17 (depends on 16)

The method of claim 16, wherein said fusion protein or pharmaceutical composition thereof is administered via injection.

### Claim 18 (depends on 17)

The method of claim 17, wherein said fusion protein or pharmaceutical composition thereof is administered subcutaneously or intramuscularly.

### Claim 19 (depends on 16)

The method of claim 16, wherein said fusion protein or pharmaceutical composition thereof is administered as a priming vaccine, the method further comprising administering a booster of said fusion protein or pharmaceutical composition thereof 7 to 14 days after administering said priming vaccine.

### Claim 20 (depends on 16)

The method of claim 16, wherein said dog is antibody-positive to PD-L1 before said administering step, wherein said fusion protein or pharmaceutical composition thereof is administered as a booster vaccine to said dog.