# US 2025/0145679 A1 · Heterodimeric polypeptide complexes comprising il-15 variants and uses thereof

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- Number: US 2025/0145679 A1
- Document ID: US20250145679A1
- Status: Published application
- Kind: A1
- Application number: 18/835,695
- Filed: 2023-02-28
- Published: 2025-05-08
- Earliest claimed: 2022-02-28
- Applicants: Wuxi Biologics Ireland Limited
- Inventors: Yu LIANG; Mengmeng SUN; Lei WU; Ruipu XIN; Jianqing XU; Jijie GU
- Web page: https://patentgrep.com/patent/US20250145679A1

## CPC

- C07K 14/5443: {IL-15}
- A61K 39/39: characterised by the immunostimulating additives, e.g. chemical adjuvants
- A61K 45/06: Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
- A61K 47/64: Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent (peptidic linkers A61K47/65)
- A61P 35/00: Antineoplastic agents
- C07K 14/7155: {for interleukins [IL]}
- C07K 16/2818: {against CD28 or CD152}
- C07K 19/00: Hybrid peptides {, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes}
- C12N 5/0636: {T lymphocytes}
- C12N 5/0646: {Natural killers cells [NK], NKT cells}
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- A61K 2039/5158: {Antigen-pulsed cells, e.g. T-cells}
- A61K 2039/585: {wherein the target is cancer}
- C07K 2317/33: Crossreactivity, e.g. for species or epitope, or lack of said crossreactivity
- C07K 2317/569: Single domain, e.g. dAb, sdAb, VHH, VNAR or nanobody®
- C07K 2317/74: Inducing cell proliferation
- C07K 2317/92: Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
- C07K 2319/00: Fusion polypeptide
- C07K 2319/30: Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
- C07K 2319/33: fusions for targeting to specific cell types, e.g. tissue specific targeting, targeting of a bacterial subspecies
- C12N 2501/2315: Interleukin-15 (IL-15)
- C12N 2510/00: Genetically modified cells
- 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)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Provided in the present disclosure are IL-15 variants, IL-15/IL-15Rα-Fc fusion proteins comprising the IL-15 variants and IL-15/IL-15Rα fusion protein comprising the IL-15 variants the methods of producing the same and the uses thereof. The IL-15 variants, fusion proteins, and polypeptide complexes of the disclosure can be used as a potent agent for the treatment of cancers and infectious diseases.

## Claims

### Claim at position 1

1-44. (canceled)

### Claim 5 (depends on 51)

The polypeptide complex of claim 51, comprising a first chain and a second chain, wherein:
(a) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 19, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 20;
(b) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 70, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 71;
(c) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 72, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 73;
(d) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 74, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 75;
(e) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 21, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 22;
(f) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 23, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 24;
(g) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 25, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 26;
(h) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 76, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 77;
(i) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 13, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 14;
(j) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 15, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 16;
(k) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 17, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 18;
(l) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 39, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 40;
(m) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 41, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 42;
(n) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 43, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 44;
(o) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 45, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 46;
(p) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 47, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 48; or
(q) the first chain comprises an amino acid sequence as set forth in SEQ ID No: 49, and the second chain comprises an amino acid sequence as set forth in SEQ ID No: 50.

### Claim 45

A polypeptide complex comprising an Interleukin-15 (IL-15) variant domain, an IL-15Rα domain, a first dimerization domain and a second dimerization domain, wherein:
the IL-15 variant domain comprises an IL-15 variant peptide of the amino acid sequence of any of SEQ ID Nos: 51-69; and
the first dimerization domain and the second dimerization domain associates together to form a dimer.

### Claim 46 (depends on 45)

The polypeptide complex of claim 45, wherein the IL-15Rα domain comprises or consists of a wild-type IL-15Rα protein, an IL-15Rα variant or any fragment thereof that retains IL-15 binding activity, such as a fragment as set forth in SEQ ID No: 2.

### Claim 47 (depends on 46)

The polypeptide complex of claim 46, further comprising one or more antigen-binding moieties, optionally in VHH, Fab, scFv, VH or VL format.

### Claim 48 (depends on 47)

The polypeptide complex of claim 47, wherein the antigen-binding moiety specifically binds to a target antigen selected from PD-1, PD-L1, PD-L2, CTLA-4, LAG3, TIM-3, TIM4, 4-1BB, OX-40, OX-40L, GITR, A2aR, TIGIT, CD96, PVRIG, CD226, 5T4, VISTA, VSIG3, VSIG4, ICOS, CD28, CD3, CD4, CD8, CD45, CD44v6, CD27, CD47, SIRPAα, SLAMF7, CD24, Siglec10, Siglec15, Siglec8, VSIR, VSIG4, PSGL-1, C5AR1, BTN1A1, BTN3A1, CD70, RANKL, CSF1R, CSF2RB, TNFRSF1/1a/1b, BDCA2, BTLA, C5aR, NKG2A, NKG2D, NKp30, NKp46, CD16a, CD56, CD166, FCGR3, CD2, Neurophilin-1, CCR8, CCR2, CCR4, CCR5, CCR6, CCR7, CCR8, GCGR, CXCR2, CXCR4, CXCR5, CALCRL, ETAR, GLP1R, CX3CR1, GPR1, GPR17, GPR20, GPR30, GPR34, GPR-65, GPCR78, GPRC5D, GPR84, LGR4, LGR5, VEGF, VEGFR, HER2, HER3, Trop2, pCAD, ERα, EGFR, de2-7 EGFR, EGFRvIII, PSMA, PSCA, PSA, TAG-72, SEZ6, SEZ6L, SEZ6L2, SEMA4D, DLL3, GD2, GPC3, KLB, KLRB1, KLRG1, GPC1, PCSK9, EpCAM, p-Cadherin, Caludin 6, Caludin 18.2, FGFR2b, FGFR3, FGFR4, MUC1, MUC13, MUC16, MUC17, MUCL3, FolRa, TfR, TF, TFR, TFPI, c-Met, NY-ESO-1, GUCY2C, LIV-1, Integrin αvβ6, Integrin α10β1, Integrin α3, Integrin α5β4, Integrin αvβ, Integrin αvβ8, ROR1, ROR2, PRLR, PTK7, B7-H3, Nectin-4, NetG1, Ax1, CD147, LRRC15, Napi2b, STEAP1, LY6G6D, LYPD1, MACRO, MerTK, MICA, MICB, MSLN, Mkars, G12D, CDH3, CDH6, CDH17, APLA2, CAIX, CD46, CD47, CLDN6, EphA3, Fucosyl-GM1, ITGA3, Kallikrein, MISRII, Podocalyxin, RON, ROBO1, PAUF, PLA2, Podocalyxin, PRLR, PTK7, TM4SF1, TMEFF2, TREAKR, TREM-1, TREM-2, uPARAP, TYRP1, KAAG1, RU2AS, CD146, CD63, Endoglin, Globo H, IGF-1R, TEM1, TEM8, TAX1BP3, ADAM-9, ENPP3, EphA2, EphA3, FcRH5, NaPi3b, TWEAK, DLK1, SORT1, SSTR2, STEAP1, CD25, CD39, GARP, LRRC33, LAIR1, LAMP3, LAP, LEPR, LILRB1, LILRB2, LILRB4, RAGE, FGL1, TPBG, PDGFRB, TGFBR2, CEACAM1, CEACAM5, CEACAM6, Carcinoembryonic antigen (CEA), ICAM1, A33, CAMPATH-1 (CDw52), Carboanhydrase IX (MN/CA IX), CD248, PDPN, ITGB1, ITGAV, CD20, CD19, CD21, CD22, CLL, BCMA, DCLK1, DDR1, DLK1, DPEP3, DKK1, CD5, CD13, CD30, CD33, CD34, CD36, CD37, CD38, CD43, CD52, CD55, CD94, CD99, CD7, CD71, CD73, CD74, CD79a, CD79b, CD229, CD132, CD133, G250, CSF1R (CD115), HLA-DR, HLA-G, HTRA1, TRA-1-60, IGFR, IL-2 receptor, MCSP (Melanoma-associated cell surface chondroitin sulphate proteoglycan), ART1, ASGR1, B7H3, B7-H4, B7H6, CD124, c-Kit (CD117), CD7, Clex12A, Clever-1, IL-13RA2, IL-11RA, IL-31RA, IL-4RA, IFNAR, ActRIIb, IL-7R, SLAMF7, Fms-like tyrosine kinase 3 (FLT-3, CD135), GFRA1, BTLA, GloboH, CSF2RB, chondroitin sulfate proteoglycan 4 (CSPG4), ITGA4, Clec5a, Clec7a, Clec9a, Clec12a, CLEC14, CD205, CD206, CD200R1, CD228, CD229, CD40, CD40L, FcRn, TLR8, TLR9, TNFR2, LTBR, CD44, CD93, PDGF, PDGFR-alpha (CD140a), PDGFR-beta (CD140b), CD146, CD147, CRTH2, TNF-α, TGF-β, IL1RAcP, TSLP, DR5, ST2, fibroblast activating protein (FAP), CDCP1, Derlin1, Tenascin, frizzled 1-10, the vascular antigens VEGFR2 (KDR/FLK1), VEGFR3 (FLT4, CD309), Endoglin, Tie2 and other TAAs, I/O checkpoints, tumor microenvironment targets, or autoimmune and inflammatory diseases associated targets.

### Claim 49 (depends on 47)

The polypeptide complex of claim 47, wherein the first and second antigen-binding moieties are in VHH format and specifically bind to PD-1, optionally the first and second antigen-binding moieties have same amino acid sequences as set forth in SEQ ID No: 80.

### Claim 50 (depends on 45)

The polypeptide complex of claim 45, wherein the first dimerization domain is one chain of an immunoglobulin Fc region, and the second dimerization domain is the other chain of the immunoglobulin Fc region, wherein the Fc region comprises a human IgG Fc such as a human IgG1 Fc, IgG2 Fc, IgG3 Fc, IgG4 Fc, or a variant thereof.

### Claim 51 (depends on 50)

The polypeptide complex of claim 50, wherein the Fc variant comprises:
(a) a human IgG4 Fc engineered to comprise one or more of the following: S228P mutation, F234A/L235A mutations, M252Y/S254T/T256E mutations and “knob into hole” structure; or
(b) a human IgG1 Fc engineered to comprise one or more of the following: M252Y/S254T/T256E mutations, G236R/L328R mutations and “knob into hole” structure.

### Claim 53 (depends on 45)

A fusion protein, comprising the IL-15 variant peptide as defined in claim 45 operably linked to a non-IL-15 moiety, wherein the non-IL-15 moiety is a moiety that can enhance the half-life of the fusion protein in vivo or an IL-15Rα domain, optionally the IL-15Rα domain comprises or consists of a wild-type IL-15Rα protein, an IL-15Rα variant or any fragment thereof that retains IL-15 binding activity.

### Claim 54

An IL-15 variant peptide consisting of the amino acid sequence of any of SEQ ID Nos: 51-69.

### Claim 55 (depends on 45)

A nucleic acid molecule, comprising a nucleic acid sequence(s) encoding the first and second chain of the polypeptide complex of claim 45.

### Claim 56 (depends on 55)

A host cell or a vector comprising the nucleic acid molecule of claim 55.

### Claim 57 (depends on 45)

A pharmaceutical composition comprising the polypeptide complex of claim 45, as well as a pharmaceutically acceptable carrier.

### Claim 58 (depends on 45)

An immunoconjugate comprising the polypeptide complex of claim 45 conjugated to a moiety selected from a chemical compound (including a cytotoxic agent, a cytostatic agent), polypeptide, carbohydrate, lipid, nucleic acid or a combination thereof.

### Claim 59 (depends on 45)

A method for producing the polypeptide complex of claim 45, comprising the steps of:
culturing a host cell comprising a vector(s) encoding one or both chains of the polypeptide complex; and
isolating the polypeptide complex from the host cell culture.

### Claim 60 (depends on 45)

A method for treating or preventing cancer, an infectious disease, an immunodeficiency or lymphopenia in a subject, comprising administering an effective amount of the polypeptide complex of claim 45 to the subject.

### Claim 61 (depends on 60)

The method of claim 60, which further comprises administering an additional anti-tumor therapy, such as cell therapy, targeted therapy, chemotherapy and gene therapy.

### Claim 62 (depends on 60)

The method of claim 60, wherein the cancer is selected from breast cancer, lung cancer, colon cancer, ovarian cancer, melanoma, bladder cancer, renal cell carcinoma, liver cancer, prostate cancer, stomach cancer, pancreatic cancer, lymphoma, leukemia, and multiple myeloma.

### Claim 63 (depends on 55)

A method for treating or preventing cancer, an infectious disease, an immunodeficiency or lymphopenia in a subject, comprising administering a cell therapy (including tumor-infiltrating lymphocyte (TIL) therapy, T cell receptor (TCR) therapy, chimeric antigen receptor (CAR) T cell therapy, and NK cell therapy) or a gene therapy (including using lentivirus, AAV, poxvirus, herpes zoster virus, oncolytic virus, or other RNA/DNA vectors for gene delivery), wherein the cell therapy and the gene therapy comprises delivering the nucleic acid molecule of claim 55 to the subject.

### Claim 64 (depends on 63)

The method of claim 63, wherein the cancer is selected from breast cancer, lung cancer, colon cancer, ovarian cancer, melanoma, bladder cancer, renal cell carcinoma, liver cancer, prostate cancer, stomach cancer, pancreatic cancer, lymphoma, leukemia, and multiple myeloma.