PD-L1-targeting immunoglobulin monovariable domain

Abstract

This technology provides polypeptides that bind to PD-L1. This technology also provides nucleic acids, vectors, and compositions.

Description

[Technical Field]

[0001] This technology relates to immunoglobulin single variable domains (ISVDs) that target PD-L1, and more particularly to polypeptides and constructs comprising or essentially comprising one or more such ISVDs. The technology also relates to nucleic acid molecules encoding ISVDs and polypeptides, as well as vectors containing nucleic acids, and compositions containing ISVDs, polypeptides, nucleic acids, or vectors. Furthermore, the technology relates to these products used in methods for treating subjects suffering from cancer. The technology also relates to methods for producing these products. [Background technology]

[0002] The PD-1 (programmed cell death-1) receptor (CD279) is expressed on the surface of activated T cells. Its ligands, PD-L1 (B7-H1; CD274) and PD-L2 (B7-DC; CD273), are transmembrane proteins belonging to the B7 family, commonly expressed in dendritic cells and macrophages. PD-1 and PD-L1 / PD-L2 belong to a family of immune checkpoint proteins that act as co-inhibitors, capable of halting or limiting the progression of the T cell response. The interaction between PD-1 / PD-L1 ensures that the immune system is activated only at the appropriate time to minimize the potential for chronic autoimmune inflammation. The role of PD-L1 in the biology, cancer immunology, and immunotherapy is described by Sun et al. 2018 (Immunity 48:434) and Salmaninejad et al. 2019 (J. Cell Physiol. 234:16824).

[0003] Numerous biological compounds aimed at blocking PD-L1 have already been approved for cancer treatment. Atezolizumab (Tecentriq; Roche) is a humanized IgG1 monoclonal antibody with modified Fc and is approved for urothelial carcinoma, non-small cell lung cancer, small cell lung cancer, hepatocellular carcinoma, melanoma, alveolar soft part sarcoma, and triple-negative breast cancer. Durvalumab (Imfinzi; MedImmune / Astrazeneca) is a humanized IgG1κ monoclonal antibody and is approved for certain types of bladder cancer, lung cancer, and biliary tract cancer. BMS-936559 / MDX-1105 (Medarex / BMS) is a humanized IgG4 monoclonal antibody. Avelumab (Bavencio; Merck KGaA / Pfizer) is a fully human monoclonal antibody and is approved for the treatment of Merkel cell carcinoma, urothelial carcinoma, and renal cell carcinoma.

[0004] Shuguang et al. (2018, Protein Cell, 9:135) described a comparative study of the binding properties of these anti-PD-L1 monoclonal antibodies, elucidating the rules governing the binding properties of anti-PD-L1 biopharmaceuticals. Due to their large size and the complexity of the tumor microenvironment, monoclonal antibodies have low permeability, spurring the development of smaller molecule biopharmaceuticals with superior penetration efficiency. Embafolimab (KN035; 3D Medicines / Alphamab Oncology / Simcere Pharmaceutical Group) is a single-domain-Fc fusion protein currently marketed for the treatment of advanced biliary tract cancer and soft tissue sarcoma. Other immunoglobulin monovariable domains targeting PD-L1 are described in International Publication No. 2008 / 071447. [Overview of the project] [Problems that the invention aims to solve]

[0005] Improved small molecule biopharmaceuticals targeting PD-L1 remain in need. Such small molecule biopharmaceuticals are useful not only for their relatively small size, high affinity, potency, and / or efficacy, but also for their ability to be combined with other (cancer) target conjugates within a single chain while maintaining good manufacturing yields. Furthermore, biopharmaceuticals should demonstrate safety, convenience, and patient compliance, thereby improving patients' quality of life. [Means for solving the problem]

[0006] In some embodiments, the technology relates to a monovariable immunoglobulin domain that specifically targets PD-L1. In some embodiments, the technology relates to a high-affinity monovariable immunoglobulin domain that specifically targets PD-L1. In some embodiments, the technology relates to a high-potency monovariable immunoglobulin domain that specifically targets PD-L1. In some embodiments, the technology relates to a highly effective monovariable immunoglobulin domain that specifically targets PD-L1.

[0007] Targeting PD-L1 with small (small-sized) biopharmaceuticals, such as monovalent or bivalent immunoglobulin monovariable domains, while maintaining high affinity, potency, and / or efficacy, may be advantageous in certain applications where very small conjugates are desired. For example, small conjugates may be advantageous in facilitating the manufacture and further handling of therapeutic compounds, either in penetration into the tumor environment or when additional conjugates targeting other (tumor) targets are involved within the same construct.

[0008] This technology provides a monovariable immunoglobulin domain that specifically targets PD-L1 and blocks the interaction between PD1 and PD-L1 (having an IC50 equivalent to or better than the benchmark). This technology provides a monovariable immunoglobulin domain that specifically targets PD-L1 and blocks the interaction between CD80 and PD-L1 (having an IC50 equivalent to or better than the benchmark). This technology provides a monovariable immunoglobulin domain that specifically targets PD-L1 and exhibits efficacy with an IC50 equivalent to or better than the benchmark in bioassays such as competition with PD1 in flow cytometry or functionality in tetanus toxoid autoantigen-specific co-culture assays (tetanus toxoid (TT) assays).

[0009] The immunoglobulin monovariate domain of this technology specifically binds to PD-L1 in humans, cynomolgus monkeys, and mice, with a KD difference of up to 5 times. This technology also includes an immunoglobulin monovariate domain that does not bind to PD-L2, and / or binds to PD-L2. -4 Higher than M / Worse K D This technology provides an immunoglobulin monovariate domain that binds to the B7 family. This technology does not bind to other members of the B7 family, and / or 10 -4 Higher than M / Worse K D It provides a single variable immunoglobulin domain that binds to other members of the B7 family.

[0010] The immunoglobulin monovariate domain of this technology, as well as the polypeptide and constructs containing the immunoglobulin monovariate domain, showed high efficacy in competitive assays with PD-1 and CD80, demonstrated high efficacy and effectiveness in TT assays, and could be efficiently produced (for example, in microbial hosts such as Pichia and P. pastoris).

[0011] In particular, this technology provides the following embodiments:

[0012] 3.1 Embodiment Family 7 In some embodiments, an ISVD that specifically binds to PD-L1 is an ISVD that inhibits the interaction between PD-L1 and PD1. An ISVD that specifically binds to PD-L1 binds to the PD1 binding site on PD-L1. In some embodiments, an ISVD that specifically binds to PD-L1 interacts with a conformational epitope on PD-L1. In some embodiments, an ISVD that specifically binds to PD-L1 is an ISVD that interacts with the following amino acids of the PD-L1 protein: Y56, E58, E60, D62, N63, Q66, H78 and / or R113. By blocking this epitope on the PD-L1 protein with a monovalent immunoglobulin monovariable domain, PD1 binding and subsequent signaling can already be inhibited very efficiently.

[0013] Embodiment 1. An immunoglobulin monovariate domain (ISVD) that specifically binds to human PD-L1 and consists of essentially four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), and interacts with one or more amino acids selected from the amino acids Y56, E58, E60, D62, N63, Q66, H78 and R113 of the PD-L1 protein.

[0014] Embodiment 2. An immunoglobulin single variable domain (ISVD) as described in Embodiment 1, which interacts with the following amino acids of the PD-L1 protein: Y56, E58, D62, N63, Q66, and H78.

[0015] Embodiment 3. An immunoglobulin monovariate domain (ISVD) according to Embodiment 1 or 2, which interacts with the following amino acids of the PD-L1 protein: Y56, E58, E60, D62, N63, Q66, H78, and R113.

[0016] Embodiment 4. An immunoglobulin single variable domain (ISVD) according to any of Embodiments 1 to 3, wherein CDR2 and CDR3 interact with one or more amino acids of the PD-L1 protein selected from Y56, E58, E60, D62, N63, Q66, H78, and R113.

[0017] Embodiment 5. An immunoglobulin single variable domain (ISVD) according to any one of Embodiments 1 to 4, wherein CDR2 and CDR3 interact with the following amino acids of the PD-L1 protein: Y56, E58, D62, N63, Q66, and H78.

[0018] Embodiment 6. An immunoglobulin single variable domain (ISVD) according to any one of Embodiments 1 to 5, wherein CDR2 and CDR3 interact with the following amino acids of the PD-L1 protein: Y56, E58, E60, D62, N63, Q66, H78, and R113.

[0019] Embodiment 7. An immunoglobulin single variable domain (ISVD) according to any one of Embodiments 1 to 6, wherein S51, S53 and N56 (Kabat numbered) in CDR2 form interaction sites with amino acids of the PD-L1 protein.

[0020] Embodiment 8. An immunoglobulin single variable domain (ISVD) according to any one of Embodiments 1 to 7, wherein A98, A99, A100, A100a, I100d and I100g (Kabat numbered) in CDR3 form interaction sites with amino acids of the PD-L1 protein.

[0021] Embodiment 9. An immunoglobulin monovariate domain (ISVD) according to any one of Embodiments 1 to 8, wherein S51, S53 and N56 (Kabat numbering) of CDR2 and A98, A99, A100, A100a, I100d and I100g (Kabat numbering) of CDR3 form interaction sites with epitopes on the PD-L1 protein.

[0022] Embodiment 10. An immunoglobulin single variable domain (ISVD) according to any one of Embodiments 1 to 9, wherein S51, S53 and N56 (Kabat numbering) of CDR2 and A98, A99, A100, A100a, I100d and I100g (Kabat numbering) of CDR3 form interaction sites with one or more amino acids of a PD-L1 protein selected from Y56, E58, E60, D62, N63, Q66, H78 and R113.

[0023] Embodiment 11. An immunoglobulin monovariate domain (ISVD) according to any one of Embodiments 1 to 8, wherein S51, S53 and N56 (Kabat numbered) in CDR2 and A98, A99, A100, A100a, I100d and I100g (Kabat numbered) in CDR3 form interaction sites with the following amino acids of the PD-L1 protein: Y56, E58, D62, N63, Q66 and H78.

[0024] Embodiment 12. An immunoglobulin monovariate domain (ISVD) according to any one of Embodiments 1 to 8, wherein S51, S53 and N56 (Kabat numbered) in CDR2 and A98, A99, A100, A100a, I100d and I100g (Kabat numbered) in CDR3 form interaction sites with the following amino acids of the PD-L1 protein: Y56, E58, E60, D62, N63, Q66, H78 and R113.

[0025] Embodiment 13. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR2 (AbM numbering) consists of the amino acid sequence X1X2SX3SX4X5NX6X7 (Sequence ID 600), where X1, X2, X3, X4, X5, X6, and X7 are each any amino acid independently selected. and -CDR3 (AbM numbering) is the amino acid sequence X1X2X3AAAAX4X5IX6X7IX8X9X10 (Sequence number 603) consists of X1, X2, X3, X4, X5, X6, X7, X8, X9 and X 10 These are any amino acids that are independently selected, Immunoglobulin single variable domain (ISVD).

[0026] Embodiment 14. -CDR2 (AbM numbering) consists of the amino acid sequence X1X2SX3SX4X5NX6X7 (Sequence ID 600), where X1 is selected from A, S, T, P, and G; X2 is selected from M, L, I, V, and C; X3 is selected from A, S, T, P, and G; X4 is selected from A, S, T, P, and G; X5 is selected from R, H, and K; X6 is selected from A, S, T, P, and G; and X7 is selected from N and Q; and -CDR3 (AbM numbering) is the amino acid sequence X1X2X3AAAAX4X5IX6X7IX8X9X 10 (Sequence code 603) consists of, where X1 is selected from A, S, T, P and G, X2 is selected from A, S, T, P and G, X3 is selected from F, Y and W, X4 is selected from A, S, T, P and G, X5 is selected from A, S, T, P and G, X6 is selected from A, S, T, P and G, X7 is selected from N, Q, I, L, M, V and C, X8 is selected from F, Y and W, X9 is selected from D and E, and X 10 This is selected from F, Y, and W; An immunoglobulin monovariate domain (ISVD) as described in Embodiment 13.

[0027] Embodiment 15. An immunoglobulin monovariate domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 13 or 14; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 13 or 14; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 13 or 14; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 17 or 109; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 109, Immunoglobulin single variable domain (ISVD).

[0028] Embodiment 16. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence X1ISSX2GRX3TN (SEQ ID NO: 598) in which amino acid residue X1 is selected from A and G, amino acid residue X2 is selected from S and T, and amino acid residue X3 is selected from N, T, and F; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence X1ISSX2GRX3TN (SEQ ID NO: 598), wherein amino acid residue X1 is selected from A and G, amino acid residue X2 is selected from S and T, and amino acid residue X3 is selected from N, T, and F; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from amino acid sequence X1ISSX2GRX3TN (SEQ ID NO: 598), wherein amino acid residue X1 is selected from A and G, amino acid residue X2 is selected from S and T, and amino acid residue X3 is selected from N, T, and F; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) in which amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, amino acid residue X3 is selected from A, P and S, and amino acid residue X4 is selected from N and I; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), wherein amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, and amino acid residue X3 is selected from N and I; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), where amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L, and T, and amino acid residue X3 is selected from N and I. Immunoglobulin single variable domain (ISVD).

[0029] Embodiment 17. An ISVD according to Embodiments 13-16, wherein the amino acid sequences of CDR2 and CDR3 (AbM numbered) have at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequences of CDR2 and CDR3 of an ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 311-324.

[0030] Embodiment 18. -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 13; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 17; or -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 14; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 109; or -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 103; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 109; or -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 14; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 17; or -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 103; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 110; or -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 13; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 111; or -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 104; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 112; ISVD as described in any one of Embodiments 13 to 17.

[0031] Embodiment 19. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 23 or 24; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; c) Amino acid sequences having a difference of 6, 5, 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 23 or 24; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 17 or 109; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 39; Immunoglobulin single variable domain (ISVD).

[0032] Embodiment 20. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605) where amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from S and T, amino acid residue X3 is selected from T, N and F, amino acid residue X4 is selected from S and P, amino acid residue X5 is selected from E and K, and amino acid residue X6 is selected from G, V and A; b) An amino acid sequence having at least 80% amino acid identity with amino acid sequence X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605), wherein amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from S and T, amino acid residue X3 is selected from T, N and F, amino acid residue X4 is selected from S and P, amino acid residue X5 is selected from E and K, and amino acid residue X6 is selected from G, V and A; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from amino acid sequence X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605), wherein amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from S and T, amino acid residue X3 is selected from T, N, and F, amino acid residue X4 is selected from S and P, amino acid residue X5 is selected from E and K, and amino acid residue X6 is selected from G, V, and A; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) in which amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, amino acid residue X3 is selected from A, P and S, and amino acid residue X4 is selected from I and N; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), wherein amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, amino acid residue X3 is selected from A, P and S, and amino acid residue X4 is selected from I and N; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), in which amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L, and T, amino acid residue X3 is selected from A, P, and S, and amino acid residue X4 is selected from I and N; Immunoglobulin single variable domain (ISVD).

[0033] Embodiment 21. The ISVD according to Embodiment 19 or 20, wherein the amino acid sequences of CDR2 and CDR3 (Kabat numbered) have at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequences of CDR2 and CD3 of an ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 311-324.

[0034] Embodiment 22. -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 23; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 17; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 109; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 109; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 17; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 17; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 122; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 110; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 123; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 111; ISVD as described in any one of Embodiments 19 to 21.

[0035] Embodiment 23. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 11 or 98 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 11 or 98; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 11 or 98; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 13 or 14 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 13 or 14; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 13 or 14; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 17 or 109 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 109, Immunoglobulin single variable domain (ISVD).

[0036] Embodiment 24. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence GRX1FSX2NX3MG (SEQ ID NO: 596) in which amino acid residue X1 is selected from T, K, and G, amino acid residue X2 is selected from G and R, and amino acid residue X3 is selected from T and A; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence GRX1FSX2NX3MG (SEQ ID NO: 596), wherein amino acid residue X1 is selected from T, K, and G, amino acid residue X2 is selected from G and R, and amino acid residue X3 is selected from T and A; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence GRX1FSX2NX3MG (SEQ ID NO: 596), wherein amino acid residue X1 is selected from T, K, and G, amino acid residue X2 is selected from G and R, and amino acid residue X3 is selected from T and A; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence X1ISSX2GRX3TN (SEQ ID NO: 598) in which amino acid residue X1 is selected from A and G, amino acid residue X2 is selected from S and T, and amino acid residue X3 is selected from N, T, and F; e) An amino acid sequence having at least 80% amino acid identity with amino acid sequence X1ISSX2GRX3TN (SEQ ID NO: 598), wherein amino acid residue X1 is selected from A and G, amino acid residue X2 is selected from S and T, and amino acid residue X3 is selected from N, T, and F; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from amino acid sequence X1ISSX2GRX3TN (SEQ ID NO: 598), wherein amino acid residue X1 is selected from A and G, amino acid residue X2 is selected from S and T, and amino acid residue X3 is selected from N, T, and F; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) in which amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, amino acid residue X3 is selected from A, P and S, and amino acid residue X4 is selected from N and I; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), wherein amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, and amino acid residue X3 is selected from N and I; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), where amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L, and T, and amino acid residue X3 is selected from N and I; Immunoglobulin single variable domain (ISVD).

[0037] Embodiment 25. An ISVD according to Embodiment 23 or 24, wherein the amino acid sequence of the CDR (AbM numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with respect to the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NOs: 311-324.

[0038] Embodiment 26. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 11; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 13; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 17; or -CDR1 consists of the amino acid sequence of SEQ ID NO: 11; -CDR2 consists of one of the amino acid sequences of SEQ ID NOs: 13 and 14; -CDR3 consists of the amino acid sequence of SEQ ID NO: 17; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 98; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 14; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 98; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 103; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 99; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 103; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 99; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 14; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 11; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 14; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 17; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 100; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 103; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 110; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 100; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 13; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 111; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 101; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 104; -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 112; ISVD as described in any one of Embodiments 23 to 25.

[0039] Embodiment 27. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 22 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 22; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 22; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 23 or 24 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; f) Amino acid sequences having a difference of 6, 5, 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 23 or 24; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 17 or 109 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 109, Immunoglobulin single variable domain (ISVD).

[0040] Embodiment 28. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence X1NX2MG (SEQ ID NO: 604) in which amino acid residue X1 is selected from G and R, and amino acid residue X2 is selected from T and A; b) An amino acid sequence having at least 80% amino acid identity with amino acid sequence X1NX2MG (SEQ ID NO: 604), wherein amino acid residue X1 is selected from G and R, and amino acid residue X2 is selected from T and A; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from amino acid sequence X1NX2MG (SEQ ID NO: 604), wherein amino acid residue X1 is selected from G and R, and amino acid residue X2 is selected from T and A; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605), where amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from S and T, amino acid residue X3 is selected from T, N and F, amino acid residue X4 is selected from S and P, amino acid residue X5 is selected from E and K, and amino acid residue X6 is selected from G, V and A; e) An amino acid sequence having at least 80% amino acid identity with amino acid sequence X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605), wherein amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from S and T, amino acid residue X3 is selected from T, N and F, amino acid residue X4 is selected from S and P, amino acid residue X5 is selected from E and K, and amino acid residue X6 is selected from G, V and A; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from amino acid sequence X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605), in which amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from S and T, amino acid residue X3 is selected from T, N, and F, amino acid residue X4 is selected from S and P, amino acid residue X5 is selected from E and K, and amino acid residue X6 is selected from G, V, and A; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) in which amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, amino acid residue X3 is selected from A, P and S, and amino acid residue X4 is selected from I and N; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), wherein amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L and T, amino acid residue X3 is selected from A, P and S, and amino acid residue X4 is selected from I and N; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601), where amino acid residue X1 is selected from A and P, amino acid residue X2 is selected from I, L, and T, amino acid residue X3 is selected from A, P, and S, and amino acid residue X4 is selected from I and N; Immunoglobulin single variable domain (ISVD).

[0041] Embodiment 29. An ISVD according to Embodiment 27 or 28, wherein the amino acid sequence of the CDR (Kabat numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NOs: 311-324.

[0042] Embodiment 30. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 22; -CDR2 (Kabat numbering) consists of one of the amino acid sequences of sequence numbers 23 and 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 17; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 23; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 17; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 109; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 109; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 109; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 17; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 118; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 122; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 110; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 118; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 123; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 111; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 118; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 123; -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 111; ISVD as described in any one of Embodiments 27 to 29.

[0043] Embodiment 31. The amino acid sequence is, i) When determining the degree of amino acid identity by ignoring the amino acid residues that form the CDR sequence, it has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NOs. 4, SEQ ID NOs. 5, SEQ ID NOs. 7 and SEQ ID NOs. 311-324; and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 30.

[0044] Embodiment 32. An ISVD according to any one of Embodiments 1 to 31, which is essentially derived from a heavy chain variable domain sequence derived from a conventional quadrichain antibody, or is essentially derived from a heavy chain variable domain sequence derived from a heavy chain antibody.

[0045] Embodiment 33. An ISVD according to any one of Embodiments 1 to 32, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0046] Embodiment 34. An ISVD according to any one of Embodiments 1 to 33, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 5 and SEQ ID NO: 311 to 321, or selected from the group consisting of amino acid sequences having an amino acid sequence identity of more than 80%, preferably more than 90%, more preferably more than 95%, for example 99% or more, with at least one of the amino acid sequences of SEQ ID NO: 4, 5 and SEQ ID NO: 311 to 321.

[0047] Embodiment 35. An ISVD according to any one of Embodiments 1 to 34, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 7 and SEQ ID NO: 311 to 339.

[0048] Embodiment 36. When the ISVD is determined by surface plasmon resonance, 5.10 -9 ~10 -11 mol / liter or less, preferably, 10 -9 ~5.10 -11 mol / liter or 5.10 -10 ~10 -10 mol / liter or more preferably 10 -10 ~10 -11 mol / liter dissociation constant (K D ), and specifically binds to human PD-L1. An ISVD according to any one of Embodiments 1 to 35.

[0049] Embodiment 37. When the ISVD is determined by surface plasmon resonance, 10 5 M -1 s -1 ~about 10 7 M -1 s -1 , preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more preferably 10 6 M -1 s -1 ~10 7 M -1 s -1 , for example 10 6 M -1 s-1 ~5.10 6 M -1 s -1 k on An ISVD according to any one of embodiments 1 to 36, which specifically binds to human PD-L1 at a rapid rate.

[0050] Embodiment 38. When ISVD is determined by surface plasmon resonance, 10 -3 s -1 (t1 / 2=0.69s)~10 -6 s -1 (Provides a nearly irreversible complex over a period of several days t1 / 2), preferably 10 -3 s -1 ~5.10 -6 s -1 , comfortable 5.10 -4 s -1 ~5.10 -6 s -1 For example, 5.10 -4 s -1 ~10 -5 s -1 k off ISVD according to any one of Embodiments 1 to 37, which specifically binds to human PD-L1 at a rapid rate.

[0051] Embodiment 39. An ISVD according to any one of Embodiments 1 to 38, which specifically binds to human PD-L1 and cynomolgus monkey PD-L1, but does not bind to other members of the B7 family.

[0052] Embodiment 40. An ISVD according to any one of Embodiments 1 to 39, which specifically binds to PD-L1 in humans and cynomolgus monkeys, but does not bind to PD-L2.

[0053] Embodiment 41. In a TT assay monitored by IFN-γ production (for example, as described in the Examples), 10 -8 M or less, more preferably 10 -9 M or less, or even 5.10 -10An ISVD according to any one of Embodiments 1 to 40, having a potency (EC50 value) of M or less. For example, in such a TT assay, the immunoglobulin monovariate domain of this technology is 10 -11 M~10 -8 M, for example, 10 -10 M~10 -8 M, 10 -10 M~10 -9 M or 10 -11 M~10 -9 It may have the efficacy of M (EC50 value).

[0054] Embodiment 42. An ISVD according to any one of Embodiments 1 to 41, having an efficacy of at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95% in a TT assay monitored by IFN-γ production (e.g., as described in the Examples).

[0055] Embodiment 43. A polypeptide or construct comprising, or essentially comprising, one or more ISVDs as described in any one of Embodiments 1 to 42, further comprising one or more other groups, residues, parts or binding units which may optionally be linked via one or more linkers.

[0056] Embodiment 44. The polypeptide or construct according to Embodiment 43, wherein the one or more other groups, residues, parts or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 526, 529, 533, 541, 542, 546, 547, 548, 551, 552, 554, 560, 563, 572, 582, 583, 590 and 593.

[0057] Embodiment 45. The polypeptide or construct according to Embodiment 43 or 44, wherein the one or more linkers are one or more amino acid sequences.

[0058] Embodiment 46. A polypeptide or construct according to any one of Embodiments 43 to 45, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0059] Embodiment 47. A polypeptide or construct according to any one of Embodiments 43 to 46, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0060] Embodiment 48. A polypeptide or construct according to any one of Embodiments 43 to 47, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelized VH, domain antibodies, single-domain antibodies and dAbs.

[0061] Embodiment 49. A polypeptide or construct according to any one of Embodiments 43 to 48, which is a polyvalent construct.

[0062] Embodiment 50. A polypeptide or construct according to any one of Embodiments 43 to 49, which is a polyspecific construct.

[0063] Embodiment 51. A polypeptide or construct according to any one of Embodiments 46 to 50, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0064] Embodiment 52. The polypeptide or construct according to Embodiment 51, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and small proteins or peptides that specifically bind to serum proteins.

[0065] Embodiment 53. The polypeptide or construct according to Embodiment 52, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of human serum albumin or fragments thereof.

[0066] Embodiment 54. The polypeptide or construct according to Embodiment 52, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0067] Embodiment 55. The polypeptide or construct according to Embodiment 54, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of ISVD, VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAb, which specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0068] Embodiment 56. The polypeptide or construct according to Embodiment 55, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0069] Embodiment 57. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct according to Embodiment 56.

[0070] Embodiment 58. The polypeptide or construct according to Embodiment 57, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0071] Embodiment 59. The polypeptide or construct according to Embodiment 54, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0072] Embodiment 60. A polypeptide or construct according to any one of Embodiments 43 to 59, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0073] Embodiment 61. A polypeptide or construct according to any one of Embodiments 43 to 60, further comprising C-terminal extension.

[0074] Embodiment 62. The C-terminal extension is C-terminal extension (X) n The polypeptide or construct according to Embodiment 61, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0075] Embodiment 63. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 42, or a polypeptide according to any one of Embodiments 43 to 60.

[0076] Embodiment 64. The nucleic acid according to Embodiment 63, which is a form of a genetic construct.

[0077] Embodiment 65. A non-human host or host cell that expresses an ISVD according to any one of Embodiments 1 to 42 or a polypeptide according to any one of Embodiments 43 to 62, or is capable of expressing them under appropriate conditions, and / or contains a nucleic acid according to any one of Embodiments 63 or 64.

[0078] Embodiment 66. A method for producing an ISVD according to any one of Embodiments 1 to 42 or a polypeptide according to any one of Embodiments 43 to 62, comprising at least the following steps: a) The step of expressing the nucleic acid described in Embodiment 63 or 64 in a suitable host cell or a non-human host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 42 or the polypeptide described in any one of Embodiments 43 to 62. Methods that include...

[0079] Embodiment 67. A method for producing an ISVD according to any one of Embodiments 1 to 42 or a polypeptide according to any one of Embodiments 43 to 62, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 65 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one of Embodiments 1 to 42 or a polypeptide as described in at least one of Embodiments 43 to 62; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 42 or the polypeptide described in any one of Embodiments 43 to 62. Methods that include...

[0080] Embodiment 68. A composition comprising an ISVD according to at least one embodiment 1 to 42, a polypeptide or construct according to at least one embodiment 43 to 62, or a nucleic acid according to at least one embodiment 63 or 64.

[0081] Embodiment 69. The composition according to Embodiment 68, which is a pharmaceutical composition.

[0082] Embodiment 70. The composition according to Embodiment 68 or 69, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0083] Embodiment 71. An ISVD according to any one of Embodiments 1 to 42, a polypeptide or construct according to any one of Embodiments 43 to 62, or a composition according to any one of Embodiments 68 to 70, for use as a pharmaceutical.

[0084] Embodiment 72. An ISVD according to any one of Embodiments 1 to 42, a polypeptide or construct according to any one of Embodiments 43 to 62, or a composition according to any one of Embodiments 68 to 70, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0085] Embodiment 73. An ISVD according to any one of Embodiments 1 to 42, a polypeptide or construct according to any one of Embodiments 43 to 62, or a composition according to any one of Embodiments 68 to 70, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0086] Embodiment 74. An ISVD according to any one of Embodiments 1 to 42, a polypeptide or construct according to any one of Embodiments 43 to 62, or a composition according to any one of Embodiments 68 to 70, for use in the diagnosis, prevention and / or treatment of cancer.

[0087] Embodiment 75. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 42, a polypeptide or construct according to any one of Embodiments 43 to 62, or a composition according to any one of Embodiments 68 to 70.

[0088] Embodiment 76. A method according to Embodiment 75 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease or disorder an ISVD according to at least one Embodiment 1 to 42, a polypeptide or construct according to one of Embodiments 43 to 62, or a composition according to one of Embodiments 68 to 70 in a pharmaceutically active amount.

[0089] Embodiment 77. A method according to any one of Embodiments 75 or 76 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 42, a polypeptide or construct according to any one of Embodiments 43 to 62, or a composition according to any one of Embodiments 68 to 70.

[0090] 3.2 Embodiment Family 42

[0091] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of Sequence ID No. 132; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 132; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 132; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 134 or 136 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 134 or 136; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 134 or 136; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 143 or 145 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 143 or 145; Immunoglobulin single variable domain (ISVD).

[0092] Embodiment 2. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence GGTFRHYVMG (Sequence ID 132); b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence X1X2SWSGSGX3Y (SEQ ID NO: 607) in which amino acid residue X1 is selected from A or G, amino acid residue X2 is selected from I, S, and V, and amino acid residue X3 is selected from T and Q; e) An amino acid sequence having at least 80% amino acid identity with amino acid sequence X1X2SWSGSGX3Y (SEQ ID NO: 607), wherein amino acid residue X1 is selected from A or G, amino acid residue X2 is selected from I, S, and V, and amino acid residue X3 is selected from T and Q; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from amino acid sequence X1X2SWSGSGX3Y (SEQ ID NO: 607), where amino acid residue X1 is selected from A or G, amino acid residue X2 is selected from I, S, and V, and amino acid residue X3 is selected from T and Q; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608) where amino acid residue X1 is selected from A, V, R, K, and E, amino acid residue X2 is selected from Q and G, amino acid residue X3 is selected from V, K, Q, H, and E, and amino acid residue X4 is selected from L and R; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608), wherein amino acid residue X1 is selected from A, V, R, K, and E, amino acid residue X2 is selected from Q and G, amino acid residue X3 is selected from V, K, Q, H, and E, and amino acid residue X4 is selected from L and R; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608), where amino acid residue X1 is selected from A, V, R, K, and E, amino acid residue X2 is selected from Q and G, amino acid residue X3 is selected from V, K, Q, H, and E, and amino acid residue X4 is selected from L and R; Immunoglobulin single variable domain (ISVD).

[0093] Embodiment 3. An ISVD according to Embodiment 1 or 2, wherein the amino acid sequence of the CDR (AbM numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NOs. 340 to 379.

[0094] Embodiment 4. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 134; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 143; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 144; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 145; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 147; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 148; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 149; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 150; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 149; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 145; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 147; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 149; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 150; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 144; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 151; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 132; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 137; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 152; An ISVD as described in any one of Embodiments 1 to 3.

[0095] Embodiment 5. An immunoglobulin monovariate domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 159; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 159; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 159; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 160 or 162 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 160 or 162; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 160 or 162; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 143 or 145 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 143 or 145; Immunoglobulin single variable domain (ISVD).

[0096] Embodiment 6. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of HYVMG (SEQ ID NO: 159) b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence HYVMG (SEQ ID NO: 159); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence HYVMG (SEQ ID NO: 159); and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610), where amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from I, S and V, and amino acid residue X3 is selected from T and Q; e) An amino acid sequence having at least 80% amino acid identity with amino acid sequence X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610), wherein amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from I, S and V, and amino acid residue X3 is selected from T and Q; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610), where amino acid residue X1 is selected from G and A, amino acid residue X2 is selected from I, S and V, and amino acid residue X3 is selected from T and Q; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608) where amino acid residue X1 is selected from A, V, R, K, or E, amino acid residue X2 is selected from Q or G, amino acid residue X3 is selected from V, K, Q, H, or E, and amino acid residue X4 is selected from R or L; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608), wherein amino acid residue X1 is selected from A, V, R, K, or E, amino acid residue X2 is selected from Q or G, amino acid residue X3 is selected from V, K, Q, H, or E, and amino acid residue X4 is selected from R or L; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608), where amino acid residue X1 is selected from A, V, R, K, or E, amino acid residue X2 is selected from Q or G, amino acid residue X3 is selected from V, K, Q, H, or E, and amino acid residue X4 is selected from R or L; Immunoglobulin single variable domain (ISVD).

[0097] Embodiment 7. An ISVD according to Embodiment 5 or 6, wherein the amino acid sequence of the CDR (AbM numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NOs. 340 to 379.

[0098] Embodiment 8. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 160; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 143; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 144; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 145; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 147; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 148; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 149; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 150; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 148; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 144; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 166; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 145; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 147; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 149; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 147; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 150; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 151; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 163; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 152; ISVD as described in any one of Embodiments 5 to 7.

[0099] Embodiment 9. The amino acid sequence is, i) When determining the degree of amino acid identity while ignoring the amino acid residues that form the CDR sequence, it has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NOs. 340-379; and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 8.

[0100] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is essentially composed of a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or is essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0101] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0102] Embodiment 12. An ISVD according to any one of Embodiments 1 to 11, which is a humanized ISVD selected from the group consisting of SEQ ID NOs: 341 to 378, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99% amino acid sequence identity with at least one amino acid sequence of SEQ ID NOs: 341 to 378.

[0103] Embodiment 13. An ISVD according to any one of Embodiments 1 to 12, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 340 to 419.

[0104] Embodiment 14. Dissociation constant (K) determined by surface plasmon resonance method D ) is 5.10 -9 ~10 -11 moles / liter or less, preferably 10 -9 ~5.10 -11 moles / liter or 5.10 -10 ~10 -10 moles / liter, comfortable, 10 -10 ~10 -11 An ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 in moles / liter.

[0105] Embodiment 15. When ISVD is determined by surface plasmon resonance, 10 4 M -1 s -1 ~about 10 7 M -1 s-1 Preferably 10 5 M -1 s -1 ~10 7 M -1 s -1 More preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 For example 10 5 M -1 s -1 ~5.10 6 M -1 s -1 at a k on rate that specifically binds to human PD-L1, the ISVD according to any one of Embodiments 1 to 14.

[0106] In Embodiment 16. When the ISVD is determined by surface plasmon resonance, 10 -3 s -1 (t1 / 2 = 0.69 s)~10 -6 s -1 (providing a complex that is substantially irreversible with a t1 / 2 of several days), preferably 10 -3 s -1 ~5.10 -6 s -1 More preferably 5.10 -4 s -1 ~5.10 -6 s -1 For example 5.10 -4 s -1 ~10 -5 s -1 at a k off rate that specifically binds to human PD-L1, the ISVD according to any one of Embodiments 1 to 15.

[0107] In Embodiment 17. The ISVD according to any one of Embodiments 1 to 16 that specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0108] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, which specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0109] Embodiment 19. In a TT assay monitored by IFN-γ production (for example, as described in the Examples), 10 -8 M or less, more preferably 10 -9 M or less, or even 5.10 -10 An ISVD according to any one of Embodiments 1 to 16, having a potency (EC50 value) of M or less. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10 -11 M~10 -8 M, for example, 10 -10 M~10 -8 M, 10 -10 M~10 -9 M or 10 -11 M~10 -9 It may have the efficacy of M (EC50 value).

[0110] Embodiment 20. An ISVD according to any one of Embodiments 1 to 19, having an efficacy of at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95% in a TT assay monitored via IFN-γ production (e.g., a TT assay as described in the Examples).

[0111] Embodiment 21. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 20, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0112] Embodiment 22. The polypeptide or construct according to Embodiment 21, wherein the one or more other groups, residues, parts or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 559, 561, 568, 575, 578, 580, 585, 587, 589, 591 and 595.

[0113] Embodiment 23. The polypeptide or construct according to Embodiment 21 or 22, wherein the one or more linkers are one or more amino acid sequences.

[0114] Embodiment 24. A polypeptide or construct according to any one of Embodiments 21 to 23, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0115] Embodiment 25. A polypeptide or construct according to any one of Embodiments 21 to 24, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0116] Embodiment 26. A polypeptide or construct according to any one of Embodiments 21 to 25, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0117] Embodiment 27. A polypeptide or construct according to any one of Embodiments 21 to 26, which is a polyvalent construct.

[0118] Embodiment 28. A polypeptide or construct according to any one of Embodiments 21 to 27, which is a polyspecific construct.

[0119] Embodiment 29. A polypeptide or construct according to any one of Embodiments 24 to 28, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0120] Embodiment 30. The polypeptide or construct according to Embodiment 29, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that can bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that can specifically bind to serum proteins.

[0121] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0122] Embodiment 32. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0123] Embodiment 33. The polypeptide or construct according to Embodiment 32, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0124] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0125] Embodiment 35. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct described in Embodiment 34.

[0126] Embodiment 36. The polypeptide or construct according to Embodiment 35, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0127] Embodiment 37. The polypeptide or construct according to Embodiment 34, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0128] Embodiment 38. A polypeptide or construct according to any one of Embodiments 21 to 37, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0129] Embodiment 39. A polypeptide or construct according to any one of Embodiments 21 to 38, further comprising C-terminal extension.

[0130] Embodiment 40. The C-terminal extension is C-terminal extension (X) n The polypeptide or construct according to Embodiment 39, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0131] Embodiment 41. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 20, or a polypeptide according to any one of Embodiments 21 to 40.

[0132] Embodiment 42. The nucleic acid described in Embodiment 41, which is a form of a genetic construct.

[0133] Embodiment 43. A non-human host or host cell that expresses an ISVD described in any one of Embodiments 1 to 20 or a polypeptide described in any one of Embodiments 21 to 40, or is capable of expressing them under appropriate conditions, and / or contains the nucleic acid described in Embodiment 41 or 42.

[0134] Embodiment 44. A method for producing an ISVD according to any one of Embodiments 1 to 20 or a polypeptide according to any one of Embodiments 21 to 40, comprising at least the following steps: a) A step of expressing the nucleic acid described in Embodiment 41 or 42 in a suitable host cell or a non-human host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 20 or the polypeptide described in any one of Embodiments 21 to 40. Methods that include...

[0135] Embodiment 45. A method for producing an ISVD according to any one of Embodiments 1 to 20 or a polypeptide according to any one of Embodiments 21 to 40, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one of Embodiments 1 to 20 or a polypeptide as described in at least one of Embodiments 21 to 40; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 20 or the polypeptide described in any one of Embodiments 21 to 40. Methods that include...

[0136] Embodiment 46. A composition comprising an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to at least one of Embodiments 21 to 40, or a nucleic acid according to at least one Embodiment 41 or 42.

[0137] Embodiment 47. The composition according to Embodiment 46, which is a pharmaceutical composition.

[0138] Embodiment 48. The composition according to Embodiment 46 or 47, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0139] Embodiment 49. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use as a pharmaceutical.

[0140] Embodiment 50. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0141] Embodiment 51. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0142] Embodiment 52. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of cancer.

[0143] Embodiment 53. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47.

[0144] Embodiment 54. A method according to Embodiment 53 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0145] Embodiment 55. A method according to Embodiment 53 or 54 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to one of Embodiments 21 to 40, or a composition according to one of Embodiments 46 to 47.

[0146] 3.3 Embodiment Family 18

[0147] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 169 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 169; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 169; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 171 or 175 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 171 or 175; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 171 or 175; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 192 or 193 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 192 or 193; Immunoglobulin single variable domain (ISVD).

[0148] Embodiment 2. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence GSIFTSAVME (SEQ ID NO: 169); b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence GSIFTSAVME (SEQ ID NO: 169); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence GSIFTSAVME (SEQ ID NO: 169); and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence RIGSGGRX1A (SEQ ID NO: 611) in which amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P, and G; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence RIGSGGRX1A (SEQ ID NO: 611), wherein amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P, and G; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from amino acid sequence RIGSGGRX1A (SEQ ID NO: 611), wherein amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P, and G; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence VX1GGYIY (SEQ ID NO: 612) in which amino acid residue X1 is selected from M and I; h) An amino acid sequence having at least 80% amino acid identity with amino acid sequence VX1GGYIY (SEQ ID NO: 612), wherein amino acid residue X1 is selected from M and I; i) An amino acid sequence having a difference of 3, 2, or 1 amino acid from the amino acid sequence VX1GGYIY (SEQ ID NO: 612), wherein amino acid residue X1 is selected from M and I; Immunoglobulin single variable domain (ISVD).

[0149] Embodiment 3. An ISVD according to Embodiment 1 or 2, wherein the amino acid sequence of the CDR (AbM numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NOs. 420 to 450.

[0150] Embodiment 4. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 171; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 192; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 172; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 173; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 174; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 175; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 176; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 177; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 178; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 171; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 179; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 180; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 169; -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 181; and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 193. An ISVD as described in any one of Embodiments 1 to 3.

[0151] Embodiment 5. An immunoglobulin monovariate domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 197 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 197; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 197; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 198 or 202 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 198 or 202; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 198 or 202; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 192 or 193 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 192 or 193; Immunoglobulin single variable domain (ISVD).

[0152] Embodiment 6. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SAVME (SEQ ID NO: 197) b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SAVME (SEQ ID NO: 197); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SAVME (SEQ ID NO: 197); and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613) in which amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P, and G, and amino acid residue X2 is selected from P and V; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613), wherein amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P, and G, and amino acid residue X2 is selected from P and V; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613), wherein amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P, and G, and amino acid residue X2 is selected from P and V; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence VX1GGYIY (SEQ ID NO: 612) in which amino acid residue X1 is selected from M and I; h) An amino acid sequence having at least 80% amino acid identity with amino acid sequence VX1GGYIY (SEQ ID NO: 612), wherein amino acid residue X1 is selected from M and I; i) An amino acid sequence having a difference of 3, 2, or 1 amino acid from the amino acid sequence VX1GGYIY (SEQ ID NO: 612), wherein amino acid residue X1 is selected from M and I; Immunoglobulin single variable domain (ISVD).

[0153] Embodiment 7. An ISVD according to Embodiment 5 or 6, wherein the amino acid sequence of the CDR (AbM numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NOs. 420 to 450.

[0154] Embodiment 8. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 198; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 192; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 199; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 200; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 201; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 202; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 203; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 204; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 205; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 198; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 206; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 207; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 208; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 209; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 193. ISVD as described in any one of Embodiments 5 to 7.

[0155] Embodiment 9. The amino acid sequence is, i) When determining the degree of amino acid identity by ignoring the amino acid residues that form the CDR sequence, it has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NOs. 420-450; and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 8.

[0156] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is essentially composed of a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or is essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0157] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0158] Embodiment 12. An ISVD according to any one of Embodiments 1 to 11, which is a humanized ISVD selected from the group consisting of SEQ ID NOs: 421 to 449, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example 99% or more amino acid sequence identity with at least one amino acid sequence of SEQ ID NOs: 421 to 449.

[0159] Embodiment 13. An ISVD according to any one of Embodiments 1 to 12, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs. 420 to 481.

[0160] Embodiment 14. Dissociation constant (K) determined by surface plasmon resonance method D ) is 5.10 -9 ~10 -11 moles / liter, preferably 10 -9 ~5.10 -11 moles / liter or 10 -9 ~10 -10 moles / liter or comfortable 10 -9 ~10 -11 An ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 in moles / liter.

[0161] Embodiment 15. When ISVD is determined by surface plasmon resonance, 10 4 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 4 M -1 s -1 ~10 7 M -1 s -1 , more 10 5 M -1 s -1 ~10 7 M -1 s -1 For example, 5.10 5 M -1 s -1 ~5.10 6 M-1 s -1 k on An ISVD according to any one of Embodiments 1 to 14, which specifically binds to human PD-L1 at a rapid rate.

[0162] Embodiment 16. When ISVD is determined by surface plasmon resonance, 10 -3 s -1 (t1 / 2=0.69s)~10 -6 s -1 (Provides a nearly irreversible complex over a period of several days t1 / 2), preferably 10 -3 s -1 ~5.10 -6 s -1 , more 10 -4 s -1 ~10 -6 s -1 For example, 10 -4 s -1 ~10 -5 s -1 k off An ISVD according to any one of Embodiments 1 to 15, which specifically binds to human PD-L1 at a rapid rate.

[0163] Embodiment 17. An ISVD according to any one of Embodiments 1 to 16, which specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0164] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, which specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0165] Embodiment 19. In a TT assay monitored by IFN-γ production (for example, as described in the Examples), 10 -8 M or less, more preferably 10 -9 M or less, or even 5.10 -10 An ISVD according to any one of Embodiments 1 to 16, having a potency (EC50 value) of M or less. For example, in such a TT assay, the immunoglobulin monovariate domain of the present technology has a titer (EC50 value) of 10-11 M~10 -8 For example, during M -10 M~10 -8 During M, 10 -10 M~5.10 -9 Between M, or 10 -11 M~5.10 -9 It can be within the range of M.

[0166] Embodiment 20. An ISVD according to any one of Embodiments 1 to 19, having an efficacy of at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95% in a TT assay monitored via IFN-γ production (e.g., a TT assay as described in the Examples).

[0167] Embodiment 21. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 20, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0168] Embodiment 22. The polypeptide or construct according to Embodiment 21, wherein the one or more other groups, residues, parts or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 535, 536, 538, 570, 576, 581, 586, 588, 592 and 594.

[0169] Embodiment 23. The polypeptide or construct according to Embodiment 21 or 22, wherein the one or more linkers are one or more amino acid sequences.

[0170] Embodiment 24. A polypeptide or construct according to any one of Embodiments 21 to 23, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0171] Embodiment 25. A polypeptide or construct according to any one of Embodiments 21 to 24, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0172] Embodiment 26. A polypeptide or construct according to any one of Embodiments 21 to 25, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0173] Embodiment 27. A polypeptide or construct according to any one of Embodiments 21 to 26, which is a polyvalent construct.

[0174] Embodiment 28. A polypeptide or construct according to any one of Embodiments 21 to 27, which is a polyspecific construct.

[0175] Embodiment 29. A polypeptide or construct according to any one of Embodiments 24 to 28, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0176] Embodiment 30. The polypeptide or construct according to Embodiment 29, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that can bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that can specifically bind to serum proteins.

[0177] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0178] Embodiment 32. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0179] Embodiment 33. The polypeptide or construct according to Embodiment 32, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0180] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0181] Embodiment 35. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct described in Embodiment 34.

[0182] Embodiment 36. The polypeptide or construct according to Embodiment 35, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0183] Embodiment 37. The polypeptide or construct according to Embodiment 34, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0184] Embodiment 38. A polypeptide or construct according to any one of Embodiments 21 to 37, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0185] Embodiment 39. A polypeptide or construct according to any one of Embodiments 21 to 38, further comprising C-terminal extension.

[0186] Embodiment 40. The C-terminal extension is C-terminal extension (X) nThe polypeptide or construct according to Embodiment 39, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0187] Embodiment 41. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 20, or a polypeptide according to any one of Embodiments 21 to 40.

[0188] Embodiment 42. The nucleic acid described in Embodiment 41, which is a form of a genetic construct.

[0189] Embodiment 43. A non-human host or host cell that expresses an ISVD described in any one of Embodiments 1 to 20 or a polypeptide described in any one of Embodiments 21 to 40, or is capable of expressing them under appropriate conditions, and / or contains the nucleic acid described in Embodiment 41 or 42.

[0190] Embodiment 44. A method for producing an ISVD according to any one of Embodiments 1 to 20 or a polypeptide according to any one of Embodiments 21 to 40, comprising at least the following steps: a) A step of expressing the nucleic acid described in Embodiment 41 or 42 in a suitable host cell or a non-human host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 20 or the polypeptide described in any one of Embodiments 21 to 40. Methods that include...

[0191] Embodiment 45. A method for producing an ISVD according to any one of Embodiments 1 to 20 or a polypeptide according to any one of Embodiments 21 to 40, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one of Embodiments 1 to 20 or a polypeptide as described in at least one of Embodiments 21 to 40; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 20 or the polypeptide described in any one of Embodiments 21 to 40. Methods that include...

[0192] Embodiment 46. A composition comprising an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to at least one of Embodiments 21 to 40, or a nucleic acid according to at least one Embodiment 41 or 42.

[0193] Embodiment 47. The composition according to Embodiment 46, which is a pharmaceutical composition.

[0194] Embodiment 48. The composition according to Embodiment 46 or 47, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0195] Embodiment 49. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use as a pharmaceutical.

[0196] Embodiment 50. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0197] Embodiment 51. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0198] Embodiment 52. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of cancer.

[0199] Embodiment 53. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47.

[0200] Embodiment 54. A method according to Embodiment 53 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0201] Embodiment 55. A method according to Embodiment 53 or 54 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to one of Embodiments 21 to 40, or a composition according to one of Embodiments 46 to 47.

[0202] 3.4 Embodiment Family 16

[0203] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 218 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 218; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 218; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 222 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 222; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 222; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 234 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 234; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 234; Immunoglobulin single variable domain (ISVD).

[0204] Embodiment 2. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence ERTFX1SYSMG (SEQ ID NO: 617) in which amino acid residue X1 is selected from S and G; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence ERTFX1SYSMG (SEQ ID NO: 617) in which amino acid residue X1 is selected from S and G; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence ERTFX1SYSMG (SEQ ID NO: 617) in which amino acid residue X1 is selected from S and G; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence X1IX2YRGAGPY (SEQ ID NO: 614) in which amino acid residue X1 is selected from V and P, and amino acid residue X2 is selected from G and F; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence X1IX2YRGAGPY (SEQ ID NO: 614), wherein amino acid residue X1 is selected from V and P, and amino acid residue X2 is selected from G and F; f) Amino acid sequences that differ by 3, 2, or 1 amino acid from the amino acid sequence X1IX2YRGAGPY (SEQ ID NO: 614), in which amino acid residue X1 is selected from V and P, and amino acid residue X2 is selected from G and F; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence RDGAVASIPQAFX1S (SEQ ID NO: 615) in which amino acid residue X1 is selected from A and T; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence RDGAVASIPQAFX1S (SEQ ID NO: 615) in which amino acid residue X1 is selected from A and T; i) Amino acid sequences in which amino acid residue X1 is selected from A and T and has an amino acid difference of 3, 2, or 1 amino acid from the amino acid sequence RDGAVASIPQAFX1S (SEQ ID NO: 615); Immunoglobulin single variable domain (ISVD).

[0205] Embodiment 3. An ISVD according to Embodiment 1 or 2, wherein the amino acid sequence of the CDR (AbM numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NOs. 482 to 497.

[0206] Embodiment 4. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 218, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 222, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 234; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 218, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 223, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 234; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 218, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 224, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 234; or -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 219. -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 222, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 235; An ISVD as described in any one of Embodiments 1 to 3.

[0207] Embodiment 5. An immunoglobulin monovariate domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 241 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 241; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 241; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 242 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 242; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 242; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 234 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 234; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 234; Immunoglobulin single variable domain (ISVD).

[0208] Embodiment 6. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SYSMG (SEQ ID NO: 241) b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SYSMG (SEQ ID NO: 241); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SYSMG (SEQ ID NO: 241); and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616) in which amino acid residue X1 is selected from V and P, and amino acid residue X2 is selected from G and F; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616), wherein amino acid residue X1 is selected from V and P, and amino acid residue X2 is selected from G and F; f) Amino acid sequences that differ by 3, 2, or 1 amino acid from the amino acid sequence X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616), in which amino acid residue X1 is selected from V and P, and amino acid residue X2 is selected from G and F; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence RDGAVASIPQAFX1S (SEQ ID NO: 615) in which amino acid residue X1 is selected from A and T; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence RDGAVASIPQAFX1S (SEQ ID NO: 615) in which amino acid residue X1 is selected from A and T; i) Amino acid sequences in which amino acid residue X1 is selected from A and T and has an amino acid difference of 3, 2, or 1 amino acid from the amino acid sequence RDGAVASIPQAFX1S (SEQ ID NO: 615); Immunoglobulin single variable domain (ISVD).

[0209] Embodiment 7. An ISVD according to Embodiment 5 or 6, wherein the amino acid sequence of the CDR (AbM numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of an ISVD having an amino acid sequence selected from SEQ ID NOs. 482 to 497.

[0210] Embodiment 8. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 242, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 234; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 243, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 234; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 244, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 234; or -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 242, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 235 ISVD as described in any one of Embodiments 5 to 7.

[0211] Embodiment 9. The amino acid sequence is, i) It has 80% amino acid sequence identity with any one of the amino acid sequences of sequence numbers 482-497, and for the purpose of determining the degree of amino acid identity, amino acid residues that form the CDR sequence are not considered. and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 8.

[0212] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is essentially composed of a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or is essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0213] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0214] Embodiment 12. An ISVD according to any one of Embodiments 1 to 11, which is a humanized ISVD selected from the group consisting of SEQ ID NOs: 483 to 496, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99%, amino acid sequence identity with at least one amino acid sequence of SEQ ID NOs: 483 to 496.

[0215] Embodiment 13. An ISVD according to any one of Embodiments 1 to 12, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 482 to 513.

[0216] Embodiment 14. Dissociation constant (K) determined by surface plasmon resonance method D ) is 5.10 -9 ~10 -11 It is less than or equal to moles / liter, preferably 10 -9 ~5.10 -11 moles / liter, or 5.10 -10 ~10 -10 moles / liter, comfortable 10 -9 ~10 -10 ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 in the range of moles / liter.

[0217] Embodiment 15. k determined by surface plasmon resonance method on Speed ​​is 10 5 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more 10 6 M -1 s -1 ~10 7 M -1 s -1 For example, 10 6 M -1 s -1 ~5.10 6 M -1 s -1 ISVD according to any one of Embodiments 1 to 14, which specifically binds to human PD-L1 within the specified range.

[0218] Embodiment 16. k determined by surface plasmon resonance method off Speed ​​is 10 -3 s -1 (t1 / 2=0.69s)~10 -6 s -1 (where t1 / 2 gives a nearly irreversible complex lasting several days), preferably in the range of 10 -3 s -1 ~5.10-5 s -1 , comfortable 5.10 -4 s -1 ~5.10 -5 s -1 For example, 10 -4 s -1 ~10 -5 s -1 ISVD according to any one of Embodiments 1 to 15, which specifically binds to human PD-L1 within the specified range.

[0219] Embodiment 17. An ISVD according to any one of Embodiments 1 to 16, which specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0220] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, which specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0221] Embodiment 19. In a TT assay monitored by IFN-γ production (for example, as described in the Examples), 10 -8 M or less, more preferably 10 -9 M or less, or even 5.10 -10 An ISVD according to any one of Embodiments 1 to 16, having a potency (EC50 value) of M or less. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10 -11 M~10 -8 M, for example, 10 -10 M~10 -8 M, 10 -10 M~10 -9 M or 10 -11 M~10 -9 It may have the efficacy of M (EC50 value).

[0222] Embodiment 20. An ISVD according to any one of Embodiments 1 to 19, having an efficacy of at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95% in a TT assay monitored via IFN-γ production (e.g., a TT assay as described in the Examples).

[0223] Embodiment 21. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 20, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0224] Embodiment 22. The polypeptide or construct according to Embodiment 21, wherein the one or more other groups, residues, parts or binding units are amino acid sequences and are any polypeptide selected from, for example, SEQ ID NOs: 531, 549, 567, 574, 579, 584.

[0225] Embodiment 23. The polypeptide or construct according to Embodiment 21 or 22, wherein the one or more linkers are one or more amino acid sequences.

[0226] Embodiment 24. A polypeptide or construct according to any one of Embodiments 21 to 23, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0227] Embodiment 25. A polypeptide or construct according to any one of Embodiments 21 to 24, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0228] Embodiment 26. A polypeptide or construct according to any one of Embodiments 21 to 25, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0229] Embodiment 27. A polypeptide or construct according to any one of Embodiments 21 to 26, which is a polyvalent construct.

[0230] Embodiment 28. A polypeptide or construct according to any one of Embodiments 21 to 27, which is a polyspecific construct.

[0231] Embodiment 29. A polypeptide or construct according to any one of Embodiments 24 to 28, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0232] Embodiment 30. The polypeptide or construct according to Embodiment 29, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that can bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that can specifically bind to serum proteins.

[0233] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0234] Embodiment 32. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0235] Embodiment 33. The polypeptide or construct according to Embodiment 32, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0236] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0237] Embodiment 35. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct described in Embodiment 34.

[0238] Embodiment 36. The polypeptide or construct according to Embodiment 35, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0239] Embodiment 37. The polypeptide or construct according to Embodiment 34, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0240] Embodiment 38. A polypeptide or construct according to any one of Embodiments 21 to 37, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0241] Embodiment 39. A polypeptide or construct according to any one of Embodiments 21 to 38, further comprising C-terminal extension.

[0242] Embodiment 40. The C-terminal extension is C-terminal extension (X) n The polypeptide or construct according to Embodiment 39, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0243] Embodiment 41. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 20, or a polypeptide according to any one of Embodiments 21 to 40.

[0244] Embodiment 42. The nucleic acid described in Embodiment 41, which is a form of a genetic construct.

[0245] Embodiment 43. A non-human host or host cell that expresses an ISVD described in any one of Embodiments 1 to 20 or a polypeptide described in any one of Embodiments 21 to 40, or is capable of expressing them under appropriate conditions, and / or contains the nucleic acid described in Embodiment 41 or 42.

[0246] Embodiment 44. A method for producing an ISVD according to any one of Embodiments 1 to 20 or a polypeptide according to any one of Embodiments 21 to 40, comprising at least the following steps: a) A step of expressing the nucleic acid described in Embodiment 41 or 42 in a suitable host cell or a non-human host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 20 or the polypeptide described in any one of Embodiments 21 to 40. Methods that include...

[0247] Embodiment 45. A method for producing an ISVD according to any one of Embodiments 1 to 20 or a polypeptide according to any one of Embodiments 21 to 40, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one of Embodiments 1 to 20 or a polypeptide as described in at least one of Embodiments 21 to 40; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 20 or the polypeptide described in any one of Embodiments 21 to 40. Methods that include...

[0248] Embodiment 46. A composition comprising an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to at least one of Embodiments 21 to 40, or a nucleic acid according to at least one Embodiment 41 or 42.

[0249] Embodiment 47. The composition according to Embodiment 46, which is a pharmaceutical composition.

[0250] Embodiment 48. The composition according to Embodiment 46 or 47, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0251] Embodiment 49. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use as a pharmaceutical.

[0252] Embodiment 50. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0253] Embodiment 51. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0254] Embodiment 52. An ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47, for use in the diagnosis, prevention and / or treatment of cancer.

[0255] Embodiment 53. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 20, a polypeptide or construct according to any one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47.

[0256] Embodiment 54. A method according to Embodiment 53 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to one of Embodiments 21 to 40, or a composition according to Embodiment 46 or 47 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0257] Embodiment 55. A method according to Embodiment 53 or 54 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 20, a polypeptide or construct according to one of Embodiments 21 to 40, or a composition according to one of Embodiments 46 to 47.

[0258] 3.5 Embodiment Family 17

[0259] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 256 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 256; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 256; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 267 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 267; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 267; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 279 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 279; i) Amino acid sequences having a difference of 43, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 279; Immunoglobulin single variable domain (ISVD).

[0260] Embodiment 2. The ISVD according to Embodiment 1, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 514.

[0261] Embodiment 3. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 256, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 267, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 279. ISVD as described in Embodiment 1 or 2.

[0262] Embodiment 4. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 293 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 293; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 293; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 299 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 299; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 299; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 279 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 279; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 279; Immunoglobulin single variable domain (ISVD).

[0263] Embodiment 5. The ISVD according to Embodiment 4, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 514.

[0264] Embodiment 6. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 293. -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 299, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 279. ISVD as described in Embodiment 4 or 5.

[0265] Embodiment 7. The amino acid sequence is i) It has 80% amino acid sequence identity with any one of the amino acid sequences of SEQ ID NO: 514, and for the purpose of determining the degree of amino acid identity, amino acid residues that form the CDR sequence are not considered. and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 6.

[0266] Embodiment 8. An ISVD according to any one of Embodiments 1 to 7, which is essentially derived from a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or from a heavy chain variable domain sequence derived from a heavy chain antibody.

[0267] Embodiment 9. An ISVD according to any one of Embodiments 1 to 8, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0268] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 514, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99% amino acid sequence identity with the amino acid sequence of SEQ ID NO: 514.

[0269] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 514 and 520.

[0270] Embodiment 12. Dissociation constant (K) determined by surface plasmon resonance method D ) is 10 -8 ~10 -11 It is less than or equal to moles / liter, preferably 5.10 -9 ~5.10 -11 moles / liter, or 10 -8 ~10 -10 moles / liter, comfortable 5.10 -8 ~10 -10 ISVD according to any one of embodiments 1 to 11, which specifically binds to human PD-L1 in the range of moles / liter.

[0271] Embodiment 13. k determined by surface plasmon resonance method on Speed ​​is 10 5 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more 10 6 M -1 s -1 ~10 7 M -1 s -1 For example, 10 6 M -1 s -1~5.10 6 M -1 s -1 ISVD according to any one of Embodiments 1 to 12, which specifically binds to human PD-L1 within the specified range.

[0272] Embodiment 14. k determined by surface plasmon resonance spectroscopy off Speed ​​is 10 -2 s -1 (t1 / 2=0.69s)~10 -6 s -1 (The range in which t1 / 2 gives a nearly irreversible complex lasting several days is preferably 5.10) -3 s -1 ~5.10 -6 s -1 , comfortable 5.10 -3 s -1 ~10 -5 s -1 For example, 10 -3 s -1 ~10 -5 s -1 ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 within the specified range.

[0273] Embodiment 15. An ISVD according to any one of Embodiments 1 to 14, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to other members of the B7 family.

[0274] Embodiment 16. An ISVD according to any one of Embodiments 1 to 15, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to PD-L2.

[0275] Embodiment 17. The potency (EC50 value) in a TT assay monitored by IFN-γ production (e.g., as described in the example) is 10 -7 M or less, more preferably 5.10 -8 M or lower, and even 10 -8 ISVD according to any one of Embodiments 1 to 16, wherein M is less than or equal to 10. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10-11 M~10 -7 The range of M, for example, 10 -10 M~10 -7 M, 10 -10 M~5.10 -8 It may have an effect within the range of (EC50 value).

[0276] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, wherein the efficacy in a TT assay monitored by IFN-γ production (e.g., as described in the Examples) is at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95%.

[0277] Embodiment 19. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 18, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0278] Embodiment 20. The polypeptide or construct according to Embodiment 19, wherein the one or more other groups, residues, portions or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 543, 545 and 548.

[0279] Embodiment 21. The polypeptide or construct according to Embodiment 19 or 20, wherein the one or more linkers are one or more amino acid sequences.

[0280] Embodiment 22. A polypeptide or construct according to any one of Embodiments 19 to 21, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0281] Embodiment 23. A polypeptide or construct according to any one of Embodiments 19 to 22, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0282] Embodiment 24. The polypeptide or construct according to any one of Embodiments 19 to 23, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0283] Embodiment 25. A polypeptide or construct according to any one of Embodiments 19 to 24, which is a polyvalent construct.

[0284] Embodiment 26. A polypeptide or construct according to any one of Embodiments 19 to 25, which is a polyspecific construct.

[0285] Embodiment 27. A polypeptide or construct according to any one of Embodiments 22 to 26, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0286] Embodiment 28. The polypeptide or construct according to Embodiment 27, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that specifically bind to serum proteins.

[0287] Embodiment 29. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0288] Embodiment 30. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0289] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0290] Embodiment 32. The polypeptide or construct according to Embodiment 31, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0291] Embodiment 33. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct according to Embodiment 32.

[0292] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0293] Embodiment 35. The polypeptide or construct according to Embodiment 32, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0294] Embodiment 36. The polypeptide or construct according to any one of Embodiments 19 to 35, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69. Embodiment 37. A polypeptide or construct according to any one of Embodiments 19 to 36, further comprising C-terminal extension. Embodiment 38. The C-terminal extension is C-terminal extension (X) nThe polypeptide or construct according to Embodiment 37, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0295] Embodiment 39. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 18, or a polypeptide according to any one of Embodiments 19 to 38.

[0296] Embodiment 40. A nucleic acid according to Embodiment 39, which is in the form of a genetic construct.

[0297] Embodiment 41. A non-human host or host cell that expresses an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 39, or is capable of expressing them under appropriate conditions, and / or contains a nucleic acid according to any one of Embodiments 39 or 40.

[0298] Embodiment 42. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) The step of expressing the nucleic acid described in Embodiment 39 or 40 in a suitable host cell or host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0299] Embodiment 43. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one Embodiment 1 to 18 or a polypeptide as described in at least one Embodiment 19 to 38; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0300] Embodiment 44. A composition comprising an ISVD according to at least one embodiment 1 to 18, a polypeptide or construct according to at least one embodiment 19 to 38, or a nucleic acid according to at least one embodiment 39 or 40.

[0301] Embodiment 45. The composition described in Embodiment 44, which is a pharmaceutical composition.

[0302] Embodiment 46. The composition according to Embodiment 44 or 45, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0303] Embodiment 47. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use as a pharmaceutical.

[0304] Embodiment 48. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0305] Embodiment 49. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0306] Embodiment 50. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 for use in the diagnosis, prevention and / or treatment of cancer.

[0307] Embodiment 51. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0308] Embodiment 52. The method of Embodiment 51 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0309] Embodiment 53. The method of Embodiment 51 or 52 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0310] 3.6 Embodiment Family 6

[0311] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 257 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 257; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 257; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 268 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 268; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 268; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 280; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 280; Immunoglobulin single variable domain (ISVD).

[0312] Embodiment 2. The ISVD according to Embodiment 1, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 515.

[0313] Embodiment 3. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 257, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 268, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 280 ISVD as described in Embodiment 1 or 2.

[0314] Embodiment 4. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 294 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 294; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 294; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 300 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 300; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 300; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 280; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 280; Immunoglobulin single variable domain (ISVD).

[0315] Embodiment 5. The ISVD according to Embodiment 4, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 515.

[0316] Embodiment 6. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 294, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 300, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 280 ISVD as described in Embodiment 4 or 5.

[0317] Embodiment 7. The amino acid sequence is i) It has 80% amino acid sequence identity with any one of the amino acid sequences of SEQ ID NO: 515, and for the purpose of determining the degree of amino acid identity, amino acid residues that form the CDR sequence are not considered. and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 6.

[0318] Embodiment 8. An ISVD according to any one of Embodiments 1 to 7, which is essentially derived from a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or from a heavy chain variable domain sequence derived from a heavy chain antibody.

[0319] Embodiment 9. An ISVD according to any one of Embodiments 1 to 8, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0320] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 514, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99% amino acid sequence identity with the amino acid sequence of SEQ ID NO: 514.

[0321] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 515 and 521.

[0322] Embodiment 12. Dissociation constant (K) determined by surface plasmon resonance method D ) is 10 -8 ~10 -11 It is less than or equal to moles / liter, preferably 10 -8 ~10 -10 moles / liter, or 5.10 -9 ~10 -11 moles / liter, comfortable 5.10 -9 ~10 -10 ISVD according to any one of embodiments 1 to 11, which specifically binds to human PD-L1 in the range of moles / liter.

[0323] Embodiment 13. k determined by surface plasmon resonance method on Speed ​​is 10 5 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more 10 6 M -1 s -1 ~10 7 M -1 s -1 For example, 10 6 M -1 s -1 ~5.10 6 M -1 s -1 ISVD according to any one of Embodiments 1 to 12, which specifically binds to human PD-L1 within the specified range.

[0324] Embodiment 14. k determined by surface plasmon resonance spectroscopy off Speed ​​is 10 -2 s -1 (t1 / 2=0.69s)~10 -6 s -1 (where t1 / 2 gives a nearly irreversible complex lasting several days), preferably in the range of 10 -2 s -1 ~5.10 -6 s -1 , comfortable 5.10 -3 s -1 ~5.10 -6 s -1 For example, 5.10 -3 s -1 ~10 -5 s -1 ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 within the specified range.

[0325] Embodiment 15. An ISVD according to any one of Embodiments 1 to 14, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to other members of the B7 family.

[0326] Embodiment 16. An ISVD according to any one of Embodiments 1 to 15, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to PD-L2.

[0327] Embodiment 17. In a TT assay monitored by IFN-γ production (for example, as described in the Examples), 10 -8 M or less, more preferably 10 -9 M or less, or even 5.10 -10 An ISVD according to any one of Embodiments 1 to 16, having a potency (EC50 value) of M or less. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10 -11 M~10 -8 M, for example, 10 -10 M~10 -8 M, 10 -10 M~10 -9 M or 10 -11 M~10 -9 It may have the efficacy of M (EC50 value).

[0328] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, wherein the efficacy in a TT assay monitored by IFN-γ production (e.g., as described in the Examples) is at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95%.

[0329] Embodiment 19. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 18, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0330] Embodiment 20. The polypeptide or construct according to Embodiment 19, wherein the one or more other groups, residues, parts or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 530 and 539.

[0331] Embodiment 21. The polypeptide or construct according to Embodiment 19 or 20, wherein the one or more linkers are one or more amino acid sequences.

[0332] Embodiment 22. A polypeptide or construct according to any one of Embodiments 19 to 21, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0333] Embodiment 23. A polypeptide or construct according to any one of Embodiments 19 to 22, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0334] Embodiment 24. The polypeptide or construct according to any one of Embodiments 19 to 23, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0335] Embodiment 25. A polypeptide or construct according to any one of Embodiments 19 to 24, which is a polyvalent construct.

[0336] Embodiment 26. A polypeptide or construct according to any one of Embodiments 19 to 25, which is a polyspecific construct.

[0337] Embodiment 27. A polypeptide or construct according to any one of Embodiments 22 to 26, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0338] Embodiment 28. The polypeptide or construct according to Embodiment 27, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that specifically bind to serum proteins.

[0339] Embodiment 29. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0340] Embodiment 30. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0341] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0342] Embodiment 32. The polypeptide or construct according to Embodiment 31, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0343] Embodiment 33. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct according to Embodiment 32.

[0344] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0345] Embodiment 35. The polypeptide or construct according to Embodiment 32, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0346] Embodiment 36. The polypeptide or construct according to any one of Embodiments 19 to 35, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0347] Embodiment 37. A polypeptide or construct according to any one of Embodiments 19 to 36, further comprising C-terminal extension.

[0348] Embodiment 38. The C-terminal extension is C-terminal extension (X) n The polypeptide or construct according to Embodiment 37, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0349] Embodiment 39. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 18, or a polypeptide according to any one of Embodiments 19 to 38.

[0350] Embodiment 40. A nucleic acid according to Embodiment 39, which is in the form of a genetic construct.

[0351] Embodiment 41. A non-human host or host cell that expresses an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 39, or is capable of expressing them under appropriate conditions, and / or contains a nucleic acid according to any one of Embodiments 39 or 40.

[0352] Embodiment 42. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) The step of expressing the nucleic acid described in Embodiment 39 or 40 in a suitable host cell or host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0353] Embodiment 43. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one Embodiment 1 to 18 or a polypeptide as described in at least one Embodiment 19 to 38; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0354] Embodiment 44. A composition comprising an ISVD according to at least one embodiment 1 to 18, a polypeptide or construct according to at least one embodiment 19 to 38, or a nucleic acid according to at least one embodiment 39 or 40.

[0355] Embodiment 45. The composition described in Embodiment 44, which is a pharmaceutical composition.

[0356] Embodiment 46. The composition according to Embodiment 44 or 45, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0357] Embodiment 47. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use as a pharmaceutical.

[0358] Embodiment 48. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0359] Embodiment 49. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0360] Embodiment 50. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 for use in the diagnosis, prevention and / or treatment of cancer.

[0361] Embodiment 51. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0362] Embodiment 52. The method of Embodiment 51 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0363] Embodiment 53. The method of Embodiment 51 or 52 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0364] 3.7 Embodiment Family 44

[0365] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 258 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 258; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 258; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 269 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 269; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 269; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 281 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 281; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 281; Immunoglobulin single variable domain (ISVD).

[0366] Embodiment 2. The ISVD according to Embodiment 1, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 516.

[0367] Embodiment 3. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 258, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 269, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 281. ISVD as described in Embodiment 1 or 2.

[0368] Embodiment 4. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 295 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 295; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 295; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 301 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 301; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 301; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 281 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 281; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 281; Immunoglobulin single variable domain (ISVD).

[0369] Embodiment 5. The ISVD according to Embodiment 4, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 516.

[0370] Embodiment 6. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 295. -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 301, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 281. ISVD as described in Embodiment 4 or 5.

[0371] Embodiment 7. The amino acid sequence is i) It has 80% amino acid sequence identity with any one of the amino acid sequences of SEQ ID NO: 516, and for the purpose of determining the degree of amino acid identity, amino acid residues that form the CDR sequence are not considered. and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 6.

[0372] Embodiment 8. An ISVD according to any one of Embodiments 1 to 7, which is essentially derived from a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or from a heavy chain variable domain sequence derived from a heavy chain antibody.

[0373] Embodiment 9. An ISVD according to any one of Embodiments 1 to 8, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0374] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 516, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99% amino acid sequence identity with the amino acid sequence of SEQ ID NO: 516.

[0375] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 516 and 522.

[0376] Embodiment 12. Dissociation constant (K) determined by surface plasmon resonance method D ) is 10 -8 ~10 -11 It is less than or equal to moles / liter, preferably 10 -8 ~10 -10 moles / liter, or 5.10 -9 ~10 -11 moles / liter, comfortable 5.10 -9 ~10 -10 ISVD according to any one of embodiments 1 to 11, which specifically binds to human PD-L1 in the range of moles / liter.

[0377] Embodiment 13. k determined by surface plasmon resonance method on Speed ​​is 10 5 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more 10 6 M -1 s -1 ~10 7 M -1 s -1 For example, 10 6 M -1 s -1 ~5.10 6M -1 s -1 ISVD according to any one of Embodiments 1 to 12, which specifically binds to human PD-L1 within the specified range.

[0378] Embodiment 14. k determined by surface plasmon resonance spectroscopy off Speed ​​is 10 -2 s -1 (t1 / 2=0.69s)~10 -6 s -1 (where t1 / 2 gives a nearly irreversible complex lasting several days), preferably in the range of 10 -2 s -1 ~5.10 -6 s -1 , comfortable 5.10 -3 s -1 ~5.10 -6 s -1 For example, 5.10 -3 s -1 ~10 -5 s -1 ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 within the specified range.

[0379] Embodiment 15. An ISVD according to any one of Embodiments 1 to 14, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to other members of the B7 family.

[0380] Embodiment 16. An ISVD according to any one of Embodiments 1 to 15, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to PD-L2.

[0381] Embodiment 17. The potency (EC50 value) in a TT assay monitored by IFN-γ production (e.g., as described in the example) is 10 -7 M or less, more preferably 5.10 -8 M or lower, and even 10 -8 ISVD according to any one of Embodiments 1 to 16, wherein M is less than or equal to 10. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10 -11 M~5.10-8 The range of M, for example, 10 -10 M~5.10 -8 M, 10 -10 M~10 -8 M or 10 -11 M~10 -8 It may have an effect within the range of M (EC50 value).

[0382] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, wherein the efficacy in a TT assay monitored by IFN-γ production (e.g., as described in the Examples) is at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95%.

[0383] Embodiment 19. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 18, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0384] Embodiment 20. The polypeptide or construct according to Embodiment 19, wherein the one or more other groups, residues, portions or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 556 and 562.

[0385] Embodiment 21. The polypeptide or construct according to Embodiment 19 or 20, wherein the one or more linkers are one or more amino acid sequences.

[0386] Embodiment 22. A polypeptide or construct according to any one of Embodiments 19 to 21, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0387] Embodiment 23. A polypeptide or construct according to any one of Embodiments 19 to 22, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0388] Embodiment 24. The polypeptide or construct according to any one of Embodiments 19 to 23, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0389] Embodiment 25. A polypeptide or construct according to any one of Embodiments 19 to 24, which is a polyvalent construct.

[0390] Embodiment 26. A polypeptide or construct according to any one of Embodiments 19 to 25, which is a polyspecific construct.

[0391] Embodiment 27. A polypeptide or construct according to any one of Embodiments 22 to 26, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0392] Embodiment 28. The polypeptide or construct according to Embodiment 27, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that specifically bind to serum proteins.

[0393] Embodiment 29. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0394] Embodiment 30. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0395] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0396] Embodiment 32. The polypeptide or construct according to Embodiment 31, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0397] Embodiment 33. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct according to Embodiment 32.

[0398] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0399] Embodiment 35. The polypeptide or construct according to Embodiment 32, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0400] Embodiment 36. The polypeptide or construct according to any one of Embodiments 19 to 35, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0401] Embodiment 37. A polypeptide or construct according to any one of Embodiments 19 to 36, further comprising C-terminal extension.

[0402] Embodiment 38. The C-terminal extension is C-terminal extension (X) nThe polypeptide or construct according to Embodiment 37, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0403] Embodiment 39. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 18, or a polypeptide according to any one of Embodiments 19 to 38.

[0404] Embodiment 40. A nucleic acid according to Embodiment 39, which is in the form of a genetic construct.

[0405] Embodiment 41. A non-human host or host cell that expresses an ISVD described in any one of Embodiments 1 to 18 or a polypeptide described in any one of Embodiments 19 to 39, or is capable of expressing them under appropriate conditions, and / or a non-human host or host cell containing the nucleic acid described in any one of Embodiments 39 or 40.

[0406] Embodiment 42. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) The step of expressing the nucleic acid described in Embodiment 39 or 40 in a suitable host cell or host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0407] Embodiment 43. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one Embodiment 1 to 18 or a polypeptide as described in at least one Embodiment 19 to 38; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0408] Embodiment 44. A composition comprising an ISVD according to at least one embodiment 1 to 18, a polypeptide or construct according to at least one embodiment 19 to 38, or a nucleic acid according to at least one embodiment 39 or 40.

[0409] Embodiment 45. The composition described in Embodiment 44, which is a pharmaceutical composition.

[0410] Embodiment 46. The composition according to Embodiment 44 or 45, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0411] Embodiment 47. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use as a pharmaceutical.

[0412] Embodiment 48. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0413] Embodiment 49. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0414] Embodiment 50. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 for use in the diagnosis, prevention and / or treatment of cancer.

[0415] Embodiment 51. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0416] Embodiment 52. The method of Embodiment 51 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0417] Embodiment 53. The method of Embodiment 51 or 52 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0418] 3.8 Embodiment Family 43

[0419] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 259 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 259; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 259; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 270 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 270; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 270; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 282 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 282; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 282; Immunoglobulin single variable domain (ISVD).

[0420] Embodiment 2. The ISVD according to Embodiment 1, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 517.

[0421] Embodiment 3. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 259, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 270, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 282. ISVD as described in Embodiment 1 or 2.

[0422] Embodiment 4. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 296 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 296; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 296; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 302 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 302; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 302; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 282 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 282; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 282; Immunoglobulin single variable domain (ISVD).

[0423] Embodiment 5. The ISVD according to Embodiment 4, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 517.

[0424] Embodiment 6. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 296, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 302, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 282. ISVD as described in Embodiment 4 or 5.

[0425] Embodiment 7. The amino acid sequence is i) It has 80% amino acid sequence identity with any one of the amino acid sequences of Sequence ID No. 517, and for the purpose of determining the degree of amino acid identity, amino acid residues that form the CDR sequence are not considered. and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 6.

[0426] Embodiment 8. An ISVD according to any one of Embodiments 1 to 7, which is essentially derived from a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or from a heavy chain variable domain sequence derived from a heavy chain antibody.

[0427] Embodiment 9. An ISVD according to any one of Embodiments 1 to 8, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0428] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 517, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99% amino acid sequence identity with the amino acid sequence of SEQ ID NO: 517.

[0429] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 517 and 523.

[0430] Embodiment 12. Dissociation constant (K) determined by surface plasmon resonance method D ) is 10 -8 ~10 -11 It is less than or equal to moles / liter, preferably 10 -9 ~5.10 -11 moles / liter, or 10 -8 ~10 -10 moles / liter, comfortable 10 -9 ~10 -10 ISVD according to any one of embodiments 1 to 11, which specifically binds to human PD-L1 in the range of moles / liter.

[0431] Embodiment 13. k determined by surface plasmon resonance method on Speed ​​is 10 5 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more 10 6 M -1 s -1 ~10 7 M -1 s -1 For example, 10 6 M -1 s -1 ~5.10 6 M -1 s -1 ISVD according to any one of Embodiments 1 to 12, which specifically binds to human PD-L1 within the specified range.

[0432] Embodiment 14. k determined by surface plasmon resonance spectroscopy off Speed ​​is 10 -2 s -1 (t1 / 2=0.69s)~10 -6 s -1 (where t1 / 2 gives a nearly irreversible complex lasting several days), preferably in the range of 10 -2 s -1 ~5.10 -6 s -1 , comfortable 5.10 -3 s -1 ~5.10 -6 s -1 For example, 5.10 -3 s -1 ~10 -5 s -1 ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 within the specified range.

[0433] Embodiment 15. An ISVD according to any one of Embodiments 1 to 14, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to other members of the B7 family.

[0434] Embodiment 16. An ISVD according to any one of Embodiments 1 to 15, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to PD-L2.

[0435] Embodiment 17. The potency (EC50 value) in a TT assay monitored by IFN-γ production (e.g., as described in the example) is 10 -7 M or less, more preferably 5.10 -8 M or lower, and even 10 -8 ISVD according to any one of Embodiments 1 to 16, wherein M is less than or equal to 10. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10 -11 M~5.10 -8 The range of M, for example, 10 -10 M~5.10 -8 M, 10 -10 M~10 -8 M or 10 -11 M~10 -8 It may have an effect within the range of M (EC50 value).

[0436] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, wherein the efficacy in a TT assay monitored by IFN-γ production (e.g., as described in the Examples) is at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95%.

[0437] Embodiment 19. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 18, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0438] Embodiment 20. The polypeptide or construct according to Embodiment 19, wherein the one or more other groups, residues, portions or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 557 and 565.

[0439] Embodiment 21. The polypeptide or construct according to Embodiment 19 or 20, wherein the one or more linkers are one or more amino acid sequences.

[0440] Embodiment 22. A polypeptide or construct according to any one of Embodiments 19 to 21, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0441] Embodiment 23. A polypeptide or construct according to any one of Embodiments 19 to 22, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0442] Embodiment 24. The polypeptide or construct according to any one of Embodiments 19 to 23, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0443] Embodiment 25. A polypeptide or construct according to any one of Embodiments 19 to 24, which is a polyvalent construct.

[0444] Embodiment 26. A polypeptide or construct according to any one of Embodiments 19 to 25, which is a polyspecific construct.

[0445] Embodiment 27. A polypeptide or construct according to any one of Embodiments 22 to 26, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0446] Embodiment 28. The polypeptide or construct according to Embodiment 27, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that specifically bind to serum proteins.

[0447] Embodiment 29. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0448] Embodiment 30. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0449] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0450] Embodiment 32. The polypeptide or construct according to Embodiment 31, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0451] Embodiment 33. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct according to Embodiment 32.

[0452] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0453] Embodiment 35. The polypeptide or construct according to Embodiment 32, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0454] Embodiment 36. The polypeptide or construct according to any one of Embodiments 19 to 35, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0455] Embodiment 37. A polypeptide or construct according to any one of Embodiments 19 to 36, further comprising C-terminal extension.

[0456] Embodiment 38. The C-terminal extension is C-terminal extension (X) n The polypeptide or construct according to Embodiment 37, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0457] Embodiment 39. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 18, or a polypeptide according to any one of Embodiments 19 to 38.

[0458] Embodiment 40. A nucleic acid according to Embodiment 39, which is in the form of a genetic construct.

[0459] Embodiment 41. A non-human host or host cell that expresses an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 39, or is capable of expressing them under appropriate conditions, and / or contains a nucleic acid according to any one of Embodiments 39 or 40.

[0460] Embodiment 42. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) The step of expressing the nucleic acid described in Embodiment 39 or 40 in a suitable host cell or host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0461] Embodiment 43. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one Embodiment 1 to 18 or a polypeptide as described in at least one Embodiment 19 to 38; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0462] Embodiment 44. A composition comprising an ISVD according to at least one embodiment 1 to 18, a polypeptide or construct according to at least one embodiment 19 to 38, or a nucleic acid according to at least one embodiment 39 or 40.

[0463] Embodiment 45. The composition described in Embodiment 44, which is a pharmaceutical composition.

[0464] Embodiment 46. The composition according to Embodiment 44 or 45, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0465] Embodiment 47. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use as a pharmaceutical.

[0466] Embodiment 48. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0467] Embodiment 49. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0468] Embodiment 50. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 for use in the diagnosis, prevention and / or treatment of cancer.

[0469] Embodiment 51. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0470] Embodiment 52. The method of Embodiment 51 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0471] Embodiment 53. The method of Embodiment 51 or 52 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0472] 3.9 Embodiment Family 46

[0473] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 260 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 260; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 260; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 271 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 271; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 271; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 283 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 283; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 283; Immunoglobulin single variable domain (ISVD).

[0474] Embodiment 2. The ISVD according to Embodiment 1, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 518.

[0475] Embodiment 3. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 260, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 271, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 283. ISVD as described in Embodiment 1 or 2.

[0476] Embodiment 4. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 297 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 297; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 297; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 303 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 303; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 303; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 283 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 283; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 283; Immunoglobulin single variable domain (ISVD).

[0477] Embodiment 5. The ISVD according to Embodiment 4, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 518.

[0478] Embodiment 6. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 297, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 303, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 283. ISVD as described in Embodiment 4 or 5.

[0479] Embodiment 7. The amino acid sequence is i) It has 80% amino acid sequence identity with any one of the amino acid sequences of SEQ ID NO: 518, and for the purpose of determining the degree of amino acid identity, amino acid residues that form the CDR sequence are not considered. and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 6.

[0480] Embodiment 8. An ISVD according to any one of Embodiments 1 to 7, which is essentially derived from a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or from a heavy chain variable domain sequence derived from a heavy chain antibody.

[0481] Embodiment 9. An ISVD according to any one of Embodiments 1 to 8, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0482] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 518, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99%, amino acid sequence identity with the amino acid sequence of SEQ ID NO: 518.

[0483] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 518 and 524.

[0484] Embodiment 12. Dissociation constant (K) determined by surface plasmon resonance method D ) is 10 -8 ~10 -11 It is less than or equal to moles / liter, preferably 10 -9 ~5.10 -11 moles / liter, or 10 -8 ~10 -10 moles / liter, comfortable 10 -9 ~10 -10 ISVD according to any one of embodiments 1 to 11, which specifically binds to human PD-L1 in the range of moles / liter.

[0485] Embodiment 13. k determined by surface plasmon resonance method on Speed ​​is 10 5 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more 10 6 M -1 s -1 ~10 7 M -1 s -1 For example, 10 6 M -1 s -1 ~5.10 6 M-1 s -1 ISVD according to any one of Embodiments 1 to 12, which specifically binds to human PD-L1 within the specified range.

[0486] Embodiment 14. k determined by surface plasmon resonance spectroscopy off Speed ​​is 10 -2 s -1 (t1 / 2=0.69s)~10 -6 s -1 (where t1 / 2 gives a nearly irreversible complex lasting several days), preferably in the range of 10 -2 s -1 ~5.10 -6 s -1 , comfortable 5.10 -3 s -1 ~5.10 -6 s -1 For example, 5.10 -3 s -1 ~10 -5 s -1 ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 within the specified range.

[0487] Embodiment 15. An ISVD according to any one of Embodiments 1 to 14, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to other members of the B7 family.

[0488] Embodiment 16. An ISVD according to any one of Embodiments 1 to 15, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to PD-L2.

[0489] Embodiment 17. The potency (EC50 value) in a TT assay monitored by IFN-γ production (e.g., as described in the example) is 10 -7 M or less, more preferably 5.10 -8 M or lower, and even 10 -8 ISVD according to any one of Embodiments 1 to 16, wherein M is less than or equal to 10. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10 -11 M~5.10-8 The range of M, for example, 10 -10 M~5.10 -8 M, 10 -10 M~10 -8 M or 10 -11 M~10 -8 It may have an effect within the range of M (EC50 value).

[0490] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, wherein the efficacy in a TT assay monitored by IFN-γ production (e.g., as described in the Examples) is at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95%.

[0491] Embodiment 19. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 18, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0492] Embodiment 20. The polypeptide or construct according to Embodiment 19, wherein the one or more other groups, residues, parts or binding units are amino acid sequences, and for example, the polypeptide of SEQ ID NO: 558.

[0493] Embodiment 21. The polypeptide or construct according to Embodiment 19 or 20, wherein the one or more linkers are one or more amino acid sequences.

[0494] Embodiment 22. A polypeptide or construct according to any one of Embodiments 19 to 21, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0495] Embodiment 23. A polypeptide or construct according to any one of Embodiments 19 to 22, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0496] Embodiment 24. The polypeptide or construct according to any one of Embodiments 19 to 23, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0497] Embodiment 25. A polypeptide or construct according to any one of Embodiments 19 to 24, which is a polyvalent construct.

[0498] Embodiment 26. A polypeptide or construct according to any one of Embodiments 19 to 25, which is a polyspecific construct.

[0499] Embodiment 27. A polypeptide or construct according to any one of Embodiments 22 to 26, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0500] Embodiment 28. The polypeptide or construct according to Embodiment 27, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that specifically bind to serum proteins.

[0501] Embodiment 29. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0502] Embodiment 30. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0503] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0504] Embodiment 32. The polypeptide or construct according to Embodiment 31, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0505] Embodiment 33. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct according to Embodiment 32.

[0506] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0507] Embodiment 35. The polypeptide or construct according to Embodiment 32, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0508] Embodiment 36. The polypeptide or construct according to any one of Embodiments 19 to 35, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0509] Embodiment 37. A polypeptide or construct according to any one of Embodiments 19 to 36, further comprising C-terminal extension.

[0510] Embodiment 38. The C-terminal extension is C-terminal extension (X) nThe polypeptide or construct according to Embodiment 37, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0511] Embodiment 39. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 18, or a polypeptide according to any one of Embodiments 19 to 38.

[0512] Embodiment 40. A nucleic acid according to Embodiment 39, which is in the form of a genetic construct.

[0513] Embodiment 41. A non-human host or host cell that expresses an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 39, or is capable of expressing them under appropriate conditions, and / or contains a nucleic acid according to any one of Embodiments 39 or 40.

[0514] Embodiment 42. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) The step of expressing the nucleic acid described in Embodiment 39 or 40 in a suitable host cell or host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0515] Embodiment 43. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one Embodiment 1 to 18 or a polypeptide as described in at least one Embodiment 19 to 38; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0516] Embodiment 44. A composition comprising an ISVD according to at least one embodiment 1 to 18, a polypeptide or construct according to at least one embodiment 19 to 38, or a nucleic acid according to at least one embodiment 39 or 40.

[0517] Embodiment 45. The composition described in Embodiment 44, which is a pharmaceutical composition.

[0518] Embodiment 46. The composition according to Embodiment 44 or 45, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0519] Embodiment 47. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use as a pharmaceutical.

[0520] Embodiment 48. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0521] Embodiment 49. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0522] Embodiment 50. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 for use in the diagnosis, prevention and / or treatment of cancer.

[0523] Embodiment 51. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0524] Embodiment 52. The method of Embodiment 51 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0525] Embodiment 53. The method of Embodiment 51 or 52 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0526] 3.10 Embodiment Family 45

[0527] Embodiment 1. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 262 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 262; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 262; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 272 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 272; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 272; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 284 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 284; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 284; Immunoglobulin single variable domain (ISVD).

[0528] Embodiment 2. The ISVD according to Embodiment 1, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 519.

[0529] Embodiment 3. -CDR1 (AbM numbering) consists of the amino acid sequence of sequence number 262, -CDR2 (AbM numbering) consists of the amino acid sequence of sequence number 272, and -CDR3 (AbM numbering) consists of the amino acid sequence of sequence number 284. ISVD as described in Embodiment 1 or 2.

[0530] Embodiment 4. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 298 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 298; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO. 298; and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 304 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 304; f) Amino acid sequences having a difference of 6, 5, 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 304; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 284 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 284; i) Amino acid sequences that differ from the amino acid sequence of Sequence ID No. 284 by 4, 3, 2, or 1 amino acid; Immunoglobulin single variable domain (ISVD).

[0531] Embodiment 5. The ISVD according to Embodiment 4, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 519.

[0532] Embodiment 6. -CDR1 (Kabat numbering) consists of the amino acid sequence of sequence number 298, -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 304, and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 284. ISVD as described in Embodiment 4 or 5.

[0533] Embodiment 7. The amino acid sequence is i) It has 80% amino acid sequence identity with any one of the amino acid sequences of SEQ ID NO: 519, and for the purpose of determining the degree of amino acid identity, amino acid residues that form the CDR sequence are not considered. and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the Hallmark residues listed in Table 1. ISVD as described in any one of Embodiments 1 to 6.

[0534] Embodiment 8. An ISVD according to any one of Embodiments 1 to 7, which is essentially derived from a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or from a heavy chain variable domain sequence derived from a heavy chain antibody.

[0535] Embodiment 9. An ISVD according to any one of Embodiments 1 to 8, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or a dAb.

[0536] Embodiment 10. An ISVD according to any one of Embodiments 1 to 9, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 519, or selected from the group consisting of amino acid sequences having more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99% amino acid sequence identity with the amino acid sequence of SEQ ID NO: 519.

[0537] Embodiment 11. An ISVD according to any one of Embodiments 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs. 519 and 525.

[0538] Embodiment 12. Dissociation constant (K) determined by surface plasmon resonance method D ) is 10 -8 ~10 -11 It is less than or equal to moles / liter, preferably 10 -9 ~5.10 -11 moles / liter, or 10 -8 ~10 -10 moles / liter, comfortable 10 -9 ~10 -10 ISVD according to any one of embodiments 1 to 11, which specifically binds to human PD-L1 in the range of moles / liter.

[0539] Embodiment 13. k determined by surface plasmon resonance method on Speed ​​is 10 5 M -1 s -1 ~about 10 7 M -1 s -1 Preferably 5.10 5 M -1 s -1 ~10 7 M -1 s -1 , more 10 6 M -1 s -1 ~10 7 M -1 s -1 For example, 10 6 M -1 s -1 ~5.10 6 M -1 s -1 ISVD according to any one of Embodiments 1 to 12, which specifically binds to human PD-L1 within the specified range.

[0540] Embodiment 14. k determined by surface plasmon resonance spectroscopy off Speed ​​is 10 -2 s -1 (t1 / 2=0.69s)~10 -6 s -1 (where t1 / 2 gives a nearly irreversible complex lasting several days), preferably in the range of 10 -2 s -1 ~5.10 -6 s -1 , comfortable 5.10 -3 s -1 ~5.10 -6 s -1 For example, 5.10 -3 s -1 ~10 -5 s -1 ISVD according to any one of Embodiments 1 to 13, which specifically binds to human PD-L1 within the specified range.

[0541] Embodiment 15. An ISVD according to any one of Embodiments 1 to 14, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to other members of the B7 family.

[0542] Embodiment 16. An ISVD according to any one of Embodiments 1 to 15, which specifically binds to human and cynomolgus monkey (cyno) PD-L1 and does not bind to PD-L2.

[0543] Embodiment 17. The potency (EC50 value) in a TT assay monitored by IFN-γ production (e.g., as described in the example) is 10 -7 M or less, more preferably 5.10 -8 M or lower, and even 10 -8 ISVD according to any one of Embodiments 1 to 16, wherein M is less than or equal to 10. For example, in such a TT assay, the immunoglobulin monovariate domain of the technology is 10 -11 M~5.10 -8 The range of M, for example, 10 -10 M~5.10 -8 M, 10 -10 M~10 -8 M or 10 -11 M~10 -8 It may have an effect within the range of M (EC50 value).

[0544] Embodiment 18. An ISVD according to any one of Embodiments 1 to 17, wherein the efficacy in a TT assay monitored by IFN-γ production (e.g., as described in the Examples) is at least 50%, for example, at least 60%, 70%, 80%, 90%, or 95%.

[0545] Embodiment 19. A polypeptide or construct comprising or essentially comprising one or more ISVDs described in any one of Embodiments 1 to 18, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

[0546] Embodiment 20. The polypeptide or construct according to Embodiment 19, wherein the one or more other groups, residues, portions or binding units are amino acid sequences and are one of the polypeptides selected from, for example, SEQ ID NOs: 566 and 573.

[0547] Embodiment 21. The polypeptide or construct according to Embodiment 19 or 20, wherein the one or more linkers are one or more amino acid sequences.

[0548] Embodiment 22. A polypeptide or construct according to any one of Embodiments 19 to 21, wherein one or more other groups, residues, parts or binding units are immunoglobulin sequences.

[0549] Embodiment 23. A polypeptide or construct according to any one of Embodiments 19 to 22, wherein one or more other groups, residues, parts or binding units are ISVDs.

[0550] Embodiment 24. The polypeptide or construct according to any one of Embodiments 19 to 23, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies and dAbs.

[0551] Embodiment 25. A polypeptide or construct according to any one of Embodiments 19 to 24, which is a polyvalent construct.

[0552] Embodiment 26. A polypeptide or construct according to any one of Embodiments 19 to 25, which is a polyspecific construct.

[0553] Embodiment 27. A polypeptide or construct according to any one of Embodiments 22 to 26, wherein the one or more other groups, residues, parts, or binding units increase the half-life of the polypeptide or construct compared to an ISVD without the one or more other groups, residues, parts, or binding units.

[0554] Embodiment 28. The polypeptide or construct according to Embodiment 27, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and low molecular weight proteins or peptides that specifically bind to serum proteins.

[0555] Embodiment 29. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0556] Embodiment 30. The polypeptide or construct according to Embodiment 28, wherein the one or more other groups, residues, parts or binding units that provide a polypeptide or construct having an extended half-life are selected from the group consisting of binding units that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0557] Embodiment 31. The polypeptide or construct according to Embodiment 30, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelidized VH, domain antibodies, single-domain antibodies or dAbs that specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

[0558] Embodiment 32. The polypeptide or construct according to Embodiment 31, wherein the one or more other groups, residues, parts or binding units that increase the half-life of the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0559] Embodiment 33. The ISVD that specifically binds to human serum albumin essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47 The polypeptide or construct according to Embodiment 32.

[0560] Embodiment 34. The polypeptide or construct according to Embodiment 33, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO: 47.

[0561] Embodiment 35. The polypeptide or construct according to Embodiment 32, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

[0562] Embodiment 36. The polypeptide or construct according to any one of Embodiments 19 to 35, wherein the linker is selected from the group consisting of Sequence IDs 53 to 69.

[0563] Embodiment 37. A polypeptide or construct according to any one of Embodiments 19 to 36, further comprising C-terminal extension.

[0564] Embodiment 38. The C-terminal extension is C-terminal extension (X) n The polypeptide or construct according to Embodiment 37, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4, or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected, preferably independently selected (preferably naturally occurring) from the group consisting of alanine (A), glycine (G), valine (V), leucine (L), or isoleucine (I).

[0565] Embodiment 39. A nucleic acid encoding an ISVD according to any one of Embodiments 1 to 18, or a polypeptide according to any one of Embodiments 19 to 38.

[0566] Embodiment 40. A nucleic acid according to Embodiment 39, which is in the form of a genetic construct.

[0567] Embodiment 41. A non-human host or host cell that expresses an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 39, or is capable of expressing them under appropriate conditions, and / or contains a nucleic acid according to any one of Embodiments 39 or 40.

[0568] Embodiment 42. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) The step of expressing the nucleic acid described in Embodiment 39 or 40 in a suitable host cell or host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0569] Embodiment 43. A method for producing an ISVD according to any one of Embodiments 1 to 18 or a polypeptide according to any one of Embodiments 19 to 38, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell as described in Embodiment 43 under conditions such that the non-human host or host cell expresses and / or produces an ISVD as described in at least one Embodiment 1 to 18 or a polypeptide as described in at least one Embodiment 19 to 38; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD described in any one of Embodiments 1 to 18 or the polypeptide described in any one of Embodiments 19 to 38. Methods that include...

[0570] Embodiment 44. A composition comprising an ISVD according to at least one embodiment 1 to 18, a polypeptide or construct according to at least one embodiment 19 to 38, or a nucleic acid according to at least one embodiment 39 or 40.

[0571] Embodiment 45. The composition described in Embodiment 44, which is a pharmaceutical composition.

[0572] Embodiment 46. The composition according to Embodiment 44 or 45, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

[0573] Embodiment 47. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use as a pharmaceutical.

[0574] Embodiment 48. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

[0575] Embodiment 49. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

[0576] Embodiment 50. An ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 for use in the diagnosis, prevention and / or treatment of cancer.

[0577] Embodiment 51. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of Embodiments 1 to 18, a polypeptide or construct according to any one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45.

[0578] Embodiment 52. The method of Embodiment 51 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45 to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder.

[0579] Embodiment 53. The method of Embodiment 51 or 52 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of an ISVD according to at least one of Embodiments 1 to 18, a polypeptide or construct according to one of Embodiments 19 to 38, or a composition according to Embodiment 44 or 45. [Brief explanation of the drawing]

[0580] [Figure 1A] This figure shows the off-rates of uniquely sequenced anti-PD-L1 clones on human, cynomolgus monkey (cyno), and mouse PD-L1. Off-rates were measured using surface plasmon resonance on Proteon. (A) Dot plot; (B) Dot plot showing cross-reactivity. [Figure 1B]This figure shows the off-rates of uniquely sequenced anti-PD-L1 clones on human, cynomolgus monkey (cyno), and mouse PD-L1. Off-rates were measured using surface plasmon resonance on Proteon. (A) Dot plot; (B) Dot plot showing cross-reactivity. [Figure 2] This figure shows the FACS competition profile of a subset of purified anti-PD-L1 VHH on CHO cells expressing human PD-L1. The competing factor is human PD-1. [Figure 3] This figure shows the FACS competitive profile of a portion of purified anti-PD-L1 VHH on ES-2 cells. The competitive factor is human PD-1. [Figure 4] This figure shows the FACS competition profiles of different lots and benchmark 1 of monovalent ISVD10F11 (ISVD86), bivalent ISVD10F11 (ISVD87), and bivalent ISVD36E11 (ISVD84) on ES-2 cells using human PD-1 as a competitive factor. [Figure 5] This figure shows IFN-γ production in the TT assay compared to benchmark 1 for monovalent and divalent 36E11, 10F11, and 52A01. [Figure 6] This figure shows IFN-γ production in a TT assay for monovalent ISVD10F11 (ISVD40) compared to benchmark 5. [Figure 7] This figure shows IFN-γ production in TT assays for monovalent ISVDs containing parental CDR sequences and ISVDs containing sequence-optimized CDR sequences. (A) A025502469, A025503388 and bivalent benchmarks; (B) A025502469, A025503357, A025503359; (C) A025501511, A025502467, A02552469; (D) A025501511, A025502470, A02552471. [Figure 7-1] Same as above. [Figure 8]This figure shows the structural determination of the ISVD10F11 / PD-L1 interaction site by cryo-electron microscopy (cryo-EM). ISVD10F11 is shown as the upper molecule. The extended loop shown at the bottom is CDR3. The loop on the right is CDR2 of ISVD10F11. The molecule shown at the bottom is PD-L1. [Modes for carrying out the invention]

[0581] 5.1 Definition Unless otherwise indicated or defined, all terms used have their ordinary meanings in the art and will be obvious to those skilled in the art. For example, Sambrook et al. (Molecular Cloning: A Laboratory Manual (2nd.Ed.) Vols. 1-3, Cold Spring Harbor Laboratory Press, 1989), F. Ausubel et al. (Current protocols in molecular biology, Green Publishing and Wiley Interscience, New York, 1987), Lewin (Genes II, John Wiley & Sons, New York, NY, 1985), Old et al. (Principles of Gene Manipulation: An Introduction to Genetic Engineering (2nd edition) University of California Press, Berkeley, CA, 1981); Roitt et al. (Immunology (6th Ed.) Mosby / Elsevier, Edinburgh, 2001), Roitt et al. (Roitt's Essential Immunology (10th Ed.) Blackwell Publishing, UK, 2001), and Janeway et al. al.(Immunobiology(6th See standard handbooks such as (ed.) Garland Science Publishing / Churchill Livingstone, New York, 2005), as well as the general background technologies cited herein.

[0582] Unless otherwise specified, all methods, processes, techniques, and operations not described in detail may be carried out or performed in a manner known to those skilled in the art. For example, in this case as well, please refer to the standard handbook and the general background techniques listed herein and the further references cited therein, as well as the following reviews which describe, for example, protein engineering techniques such as affinity maturation and other techniques for improving the specificity and other desired properties of proteins such as immunoglobulins: Presta (Adv. Drug Deliv. Rev. 58(5-6):640-56, 2006), Levin and Weiss (Mol. Biosyst. 2(1):49-57, 2006), Irving et al. (J. Immunol. Methods 248(1-2):31-45, 2001), Schmitz et al. (Placenta 21 Suppl. A:S106-12, 2000), and Gonzales et al. (Tumour Biol. 26(1):31-43, 2005).

[0583] As used herein, the term “sequence” (for example, “immunoglobulin sequence,” “antibody sequence,” “variable domain sequence,” “VHH sequence,” or “protein sequence”) should generally be understood to include both the relevant amino acid sequence and the nucleic acid or nucleotide sequence encoding it, unless a more restricted interpretation is required in the context.

[0584] Amino acid residues are indicated according to standard three-letter or one-letter amino acid codes. See Table A-2 on page 48 of International Publication No. 08 / 020079.

[0585] Nucleic acids or amino acids are considered "(essentially) isolated" compared to, for example, reaction medium or culture medium if they have been separated from at least one other component normally associated with the source or medium, such as another nucleic acid, another protein / polypeptide, another biological component or macromolecule, or at least one contaminant, impurity or trace component. In particular, nucleic acids or amino acids are considered "(essentially) isolated" if they have been purified by at least 2 times, especially at least 10 times, even more especially at least 100 times, and up to 1000 times or more. Nucleic acids or amino acids in "(essentially) isolated form" are preferably essentially homogeneous when determined using a suitable technique, such as a suitable chromatographic technique such as polyacrylamide gel electrophoresis.

[0586] When a nucleotide sequence or amino acid sequence is said to "contain" or "essentially consist of" another nucleotide sequence or amino acid sequence, this may mean that the latter nucleotide sequence or amino acid sequence is incorporated into the first-referenced nucleotide sequence or amino acid sequence, but more generally, it means that the first-referenced nucleotide sequence or amino acid sequence contains within its sequence stretches of nucleotide or amino acid residues having identical nucleotide or amino acid sequences to the latter sequence, regardless of how the first-referenced sequence was actually generated or obtained (e.g., by any preferred method described herein). Using a non-restrictive example, when a polypeptide is said to contain an immunoglobulin monovariate domain, this may mean that the immunoglobulin monovariate domain sequence is incorporated into the polypeptide sequence, but more usually, it means that the polypeptide contains the immunoglobulin monovariate domain sequence within its sequence, regardless of how the polypeptide was generated or obtained. Furthermore, when a nucleic acid or nucleotide sequence is said to contain another nucleotide sequence, the first-mentioned nucleic acid or nucleotide sequence is preferably such that, when expressed as an expression product (e.g., a polypeptide), the amino acid sequence encoded by the latter nucleotide sequence forms part of the expression product (in other words, the latter nucleotide sequence is within the same reading frame as the larger nucleic acid or nucleotide sequence first mentioned).

[0587] "Consists of (essentially)" means that either the latter nucleic acid sequence or amino acid sequence is identical to a polypeptide (e.g., CDR region; ISVD), or that it corresponds to a polypeptide (e.g., CDR region; ISVD) having a limited number of amino acid residues attached to the amino terminus, carboxyl terminus, or both the amino and carboxyl terminus of an immunoglobulin monovariable domain, for example, 1 to 20 amino acid residues, for example, 1 to 10 amino acid residues, preferably 1 to 6 amino acid residues, for example, 1, 2, 3, 4, 5, or 6 amino acid residues.

[0588] For the purpose of comparing two or more amino acid sequences, the "percentage of sequence identity" between a first amino acid sequence and a second amino acid sequence can be calculated by dividing [the number of amino acid residues in the first amino acid sequence that are identical to the amino acid residue at the corresponding position in the second amino acid sequence] by [the total number of amino acid residues in the first amino acid sequence] and multiplying by [100%], where each deletion, insertion, substitution, or addition of an amino acid residue in the second amino acid sequence is considered a difference at a single amino acid residue (i.e., at a single position) compared to the first amino acid sequence. Typically, for the purpose of determining the percentage of "sequence identity" between two amino acid sequences according to the calculation method outlined above in this specification, the amino acid sequence containing the largest number of amino acid residues is considered the "first" amino acid sequence, and the other amino acid sequences are considered the "second" amino acid sequences.

[0589] As used herein, “amino acid difference” means the deletion, insertion, or substitution, preferably of a single amino acid residue, compared to a reference sequence. In one embodiment, the “amino acid difference” is a substitution. In one embodiment, the amino acid substitution is a conservative substitution. Such a conservative substitution is preferably one in which one amino acid from the following groups (a) to (e) is replaced by another amino acid residue from the same group: (a) small aliphatic, nonpolar, or slightly polar residues: Ala, Ser, Thr, Pro, and Gly; (b) polar, negatively charged residues and their (uncharged) amides: Asp, Asn, Glu, and Gln; (c) polar, positively charged residues: His, Arg, and Lys; (d) large aliphatic, nonpolar residues: Met, Leu, Ile, Val, and Cys; and (e) aromatic residues: Phe, Tyr, and Trp. In one embodiment, the conservative substitution is as follows: Ala to Gly or Ser; Arg to Lys; Asn to Gln or His; Asp to Glu; Cys to Ser; Gln to Asn; Glu to Asp; Gly to Ala or Pro; His to Asn or Gln; Ile to Leu or Val; Leu to Ile or Val; Lys to Arg, Gln or Glu; Met to Leu, Tyr, or Ile; Phe to Met, Leu, or Tyr; Ser to Thr; Thr to Ser; Trp to Tyr; Tyr to Trp; and / or Phe to Val, Ile, or Leu.

[0590] As used herein, the "VHH family" refers to a group of VHH sequences that have the same length (i.e., the same number of amino acids in the sequence) and whose amino acid sequences from position 8 to position 106 (according to Kabat numbering) have an amino acid sequence identity of 89% or more.

[0591] In this specification, the terms “epitope” and “antigenic determinant” are used synonymously to refer to antibody-binding molecules, such as immunoglobulins, conventional antibodies, or immunoglobulin monovariate domains, and in particular to a portion of a macromolecule (e.g., polypeptide or protein (e.g., PD-L1)) recognized by the antigen-binding site of these molecules. An epitope defines the minimal binding site of an immunoglobulin and therefore represents the target of the immunoglobulin's specificity. The portion of an antigen-binding molecule, such as an immunoglobulin, conventional antibody, or immunoglobulin monovariate domain, that recognizes an epitope is called a “paratope.”

[0592] The framework region of a variable domain, such as an immunoglobulin monovariable domain, forms a β-sheet that provides the structural backbone of the domain, while the hypervariable sequence corresponds to three loops on the outer edge of a β-barrel that are opposed to each other within the folded domain. These hypervariable loops are close to each other and form a single hypervariable region at the molecular tip. This is the antigen binding site, or antigen-binding site, or paratope. These three hypervariable loops determine antigen specificity by forming a surface complementary to the antigen and are generally called complementarity-determining regions, or CDRs (CDR1, CDR2, and CDR3). Therefore, the region on the surface of the antibody molecule formed by the opposing arrangement of the CDRs forms the site where the antigen binds.

[0593] Antibodies generally recognize only small regions on the surface of large molecules such as proteins. The structures recognized by antibodies are called antigenic determinants or epitopes. Such sites are likely to be composed of amino acids from different parts of a polypeptide chain that have become close to each other through protein folding. This type of antigenic determinant is known as a conformational epitope or discontinuous epitope because it is a structure that is recognized consisting of protein segments that are close in three-dimensional structure but not continuous in the amino acid sequence of the antigen. In contrast, epitopes composed of a single polypeptide chain segment are called linear epitopes.

[0594] The binding between an antibody-antigen binding site (paratope) and its epitope is a reversible non-covalent interaction. The forces involved in these non-covalent interactions, i.e., the binding itself, are electrostatic forces, hydrogen bonds, van der Waals forces, and hydrophobic forces. Electrostatic interactions occur between charged amino acid side chains, as in salt bridges. Interactions involving electric dipoles, such as hydrogen bonds, or short-range van der Waals forces also occur. Hydrophobic interactions occur when two hydrophobic surfaces are close enough to exclude water. For some antigens, hydrophobic interactions are thought to account for the majority of the binding energy.

[0595] The extent to which each of these forces contributes to the overall interaction depends on the specific antibody and antigen involved. A notable difference between antibody-protein antigen interactions and most other natural protein-protein interactions is that antibodies have many aromatic amino acids at their antigen-binding sites. These amino acids are primarily involved in van der Waals and hydrophobic interactions, and sometimes in hydrogen bonding. Generally, hydrophobic and van der Waals forces act over very short distances and attract two surfaces with complementary shapes; that is, a convex part of one surface must fit into a concave part of the other surface for good bonding to occur. In contrast, electrostatic interactions between charged side chains and hydrogen bonds bridging oxygen and / or nitrogen atoms enhance the interaction as a whole while simultaneously allowing for specific characteristics or reactive groups.

[0596] Amino acids involved in any of these non-covalent interactions between an antibody-antigen binding site (paratope) and its epitope are also said to "form an interaction site" between the antigen-binding site (paratope) and the epitope.

[0597] When a polypeptide (such as an immunoglobulin, antibody, immunoglobulin monovariate domain, or antigen-binding molecule or fragment thereof) is said to be "capable of binding," "capable of specifically binding," "has affinity," and / or "has specificity" to a certain epitope, antigen, or protein (or at least a portion, fragment, or epitope thereof), it is said to be "against" or "directed against" the epitope, antigen, or protein, and is considered a "binding" molecule relating to the epitope, antigen, or protein, or is considered an "anti"epitope, "anti"antigen, or "anti"protein (e.g., "anti"PD-L1).

[0598] The terms “specificity,” “specifically binding,” or “specific binding” refer to the number of different target molecules, such as antigens of the same organism, to which a particular binding unit, such as ISVD, can bind with sufficiently high affinity (see below). “Specificity,” “specifically binding,” or “specific binding” are used herein interchangeably with “selectivity,” “selectively binding,” or “selective binding.” Binding units such as ISVD bind specifically to their designated targets. The specificity / selectivity of a binding unit can be determined based on affinity. Affinity indicates the strength or stability of molecular interactions. Affinity is generally expressed by the KD or dissociation constant, which has units of mol / liter (or M). Affinity can also be expressed as the association constant KA, where KA is equal to 1 / KD, (mol / liter) -1 (or M -1 It has units of ).

[0599] Affinity is a measure of the binding strength between a part of a molecule and a binding site on a target molecule. A smaller KD value indicates a stronger binding strength between the target molecule and the targeting site. Typically, the binding units used in this technology, such as ISVD, have a KD of 10. -5 ~10 -12 moles / liter or less, 10 -7 ~10 -12 moles / liter or less, or 10 -8 ~10 -12With a dissociation constant (KD) of moles / liter (i.e., 10 5 ~10 12 liters / mol or more, 10 7 ~10 12 liters / mol or more, or 10 8 ~10 12 It binds to its target with an association constant (KA) of liters / mol. 10 -4 Any KD value greater than mol / liter (or 10 4 Any KA value less than liters / mol is generally considered to indicate nonspecific binding. The KD of biological interactions considered specific, such as the binding of immunoglobulin sequences to antigens, is typically 10. -5 moles / liter (10000 nM or 10 μM) ~ 10 -12 The concentration is within the range of moles / liter (0.001 nM or 1 pM) or less.

[0600] Therefore, specific / selective binding is determined using the same measurement method, e.g., SPR, when the binding unit (or polypeptide containing it) is 10 -5 ~10 -12 It binds to PD-L1 with a KD value of 10 moles / liter or less. -4 This may mean binding to the relevant B7 family with a KD value greater than mol / liter. An example of a relevant B7 family member is PD-L2. Therefore, in one embodiment of this technology, ISVD binds to (human) PD-L1 at a rate of 10 -5 ~10 -12 It binds with a KD value of mol / liter or less, and for the same type of PD-L2, 10 -4 They bind with a KD value greater than moles / liter.

[0601] Specific binding to a specific target originating from a particular species does not preclude the possibility that the binding unit may also specifically bind to similar targets originating from different species. For example, specific binding to human PD-L1 does not preclude the possibility that the binding unit or the polypeptide containing it may also specifically bind to PD-L1 originating from cynomolgus monkeys ("cyno").

[0602] The specific binding of the binding unit to a specified target can be determined by any suitable method known in itself, including, for example, scatchard analysis and / or competitive binding assays, such as radioimmunoassays (RIAs), enzyme immunoassays (EIAs), and sandwich competitive assays, as well as various variations thereof known in the art; and other techniques further referenced herein.

[0603] The dissociation constant may be the actual dissociation constant or the apparent dissociation constant, as will be obvious to those skilled in the art. Methods for determining the dissociation constant will be obvious to those skilled in the art, and include, for example, the techniques referred to herein. In this regard, 10 -4 moles / liter or 10 -3 moles / liter greater than (for example, 10 -2 It will also be apparent that it may be impossible to measure the dissociation constant (moles / liter). In some cases, as will be apparent to those skilled in the art, the (actual or apparent) dissociation constant can be calculated based on the (actual or apparent) association constant (KA) by the relationship [KD = 1 / KA].

[0604] An amino acid sequence is said to be "cross-reactive" to two different antigens or antigenic determinants (for example, serum albumins from two different mammalian species, e.g., human serum albumin and cynomolgus monkey serum albumin, or PD-L1 from different mammalian species, e.g., human PD-L1, cynomolgus monkey PD-L1, and mouse PD-L1) if the amino acid sequence is specific to these different antigens or antigenic determinants (as defined herein).

[0605] In this specification, the terms “blocking,” “antagonizing,” “competitive,” “competitive,” and “-competitive” are used synonymously to mean the ability of an immunoglobulin, antibody, immunoglobulin monovariate domain, polypeptide, or other conjugate to prevent the binding of another protein, polypeptide, ligand, or conjugate to a particular target. The extent to which an immunoglobulin, antibody, immunoglobulin monovariate domain, polypeptide, or other conjugate can prevent the binding of another ligand to a target, and therefore whether it can be said to “block,” can be determined using a competitive binding assay. Particularly suitable quantitative competitive blocking assays are described in the examples and include, for example, a fluorescence-activated cell sorting (FACS) binding assay using PD-L1 expressed on cells. The degree of blocking can be measured by (reduced) channel fluorescence.

[0606] The following is a general description of a suitable FACS assay for determining whether an immunoglobulin, antibody, immunoglobulin monovariate domain, polypeptide, or other conjugate is blocked or can be blocked. Those skilled in the art will understand that this assay can be used for any immunoglobulin monovariate domain and polypeptide described herein. The FACS instrument (e.g., FACS Canto; Becton Dickinson) should be operated according to the manufacturer's recommendations.

[0607] To evaluate "blocking" or "competition" between two conjugates (e.g., a monovariate immunoglobulin domain and a native ligand or other conjugate) for binding to PD-L1, FACS competition experiments can be performed using cells overexpressing human PD-L1 (e.g., Flp-In(trademark)-293 cells) and parental cells as background cell lines. Various detection reagents can be used, such as monoclonal ANTI-FLAG(registered trademark) M2 antibody (Sigma-Aldrich, catalog number F1804), monoclonal anti-C-myc antibody (Sigma-Aldrich, catalog number WH0004609M2), and monoclonal ANTI-HIS TAG antibody (Sigma-Aldrich, catalog number SAB1305538), each individually labeled. ☆In flow cytometry, a wide range of fluorophores can be used as labels (e.g., PE (R-phycoerythrin), 7-aminoactinomycin D (7-AAD), acridine orange, various AlexaFluor, allophycocyanin (APC), AmCyan, aminocoumarin, APC Cy5, APC Cy7, APC-H7, APC / Alexa Fluor 750, AsRed2, Azami-Green, Azurite, BODIPY FL C5-ceramide, BCECF-AM, Bis-oxonol DiBAC2(3), BODIPY-FL, Calcein, Calcein AM, Caroxy-H2DCFDA, Cascade Blue, Cascade Yellow, Cell Tracker Green, Cerulean, CFSE, Chromomycin A3, CM-H2DCFDA, Cy2, Cy3, Cy3.5, Cy3B, Cy5, Cy5.5, Cy7, CyPet, DAF-FM, DAF-FM Diacetate, DAPI, DCFH (2'7'Diclodihydrofluorescein), DHR, Dihydrocalcein AM, Dihydrorhodamine, Dihydrothidium, DiLC1(5), DiOC6(3), DiOC7(3), dKeima-Red, DRAQ5, Dronpa-Green, various DsRed, dTomato, various DyLight, E.coli BioParticles AF488, E2-Crimson, E2-Orange, EBFP2, ECFP, various eFluor, EGFP, EGFP*, Emerald, eqFP650, eqFP670, ER-Tracker Blue-White DPX, Ethidium Bromide, Express2, EYFP, Fc OxyBurst Green, Fc OxyBurst Green 123, FITC, Fluo-3, Fluo-4, Fluorescein, Fura-2, Fura-Red, GFPuv, H2DCFDA, HcRed1, Hoechst Blue (33258), Hoechst Red (33342), Hydroxycoumarin, HyPer, Indo-1, Indo-1 Blue (Low Ca2+), Indo-1 Violet (high Ca2+), iRFP, J-Red, JC-1, JC-9, Katushka (TurboFP635), Katushka2 Kusabira-Orange, LDS 751, Lisamin Rhodamine B, various Live / Dead products, Lucifer Yellow, Lucifer Yellow CH, Lyso Tracker Blue, Lyso Tracker Green, Lyso Tracker Red, mAmertrine, Marina Blue, mBanana, mCFP, mCherry, mCitrine, Methoxycoumarin, mHoneyDew, Midoriishi-Cyan, Mitramycin, Mito Tracker Deep Red, Mito Tracker Green, Mito Tracker Orange, Mito Tracker Red, MitoFluor Green, mKate (TagFP635), mKate2, mKeima, mKeima-Red, mKO, mKOk, mNeptune, Monocloviman, mOrange, mOrange2, mRaspberry, mPlum, mRFP1, mStrawberry, mTangerine, mTarquoise, mTFP1, mTFP1(Teal), NBD, OxyBurst Green H2DCFDA, OxyBurst Green H2HFFBSA, Pacific Blue, PE (R-phycoerythrin), PE Cy5, PE Cy5.5, PE Cy7, PE Texas Red, PE-Cy5 conjugate, PE-Cy7 conjugate, PerCP (Peridinine Chlorophyll Protein), PerCP Cy5.5, PhiYFP, PhiYFP-m, Propidium Iodide (PI), various Qdots, Red 613, RFP Tomato, Rhod-2, S65A, S65C, S65L, S65T, Singlet Oxygen Sensor Green, Sirius, SNARF, Superfolder GFP, SYTOX Blue, SYTOX Green, SYTOX Orange, T-Sapphire, TagBFP, TagCFP, TagGFP, TagRFP, TagRFP657, TagYFP, tdTomato, Texas Red, Thiazole Orange, TMRE, TMRM, Topaz, TOTO-1, TO-PRO-1, TRITC, TRITC Examples include TruRed, TurboFP602, TurboFP635, TurboGFP, TurboRFP, TurboYFP, Venus, Vybrant CycleDye Violet, Wild Type GFP, X-Rhodamin, Y66F, Y66H, Y66W, YOYO-1, YPet, ZsGreen1, ZsYellow1, and Zymosan A BioParticles AF488 (see http: / / www.thefcn.org / flow-fluorochromes for details). Fluorophores, or simply "fluor," are typically conjugated to antibodies that recognize PD-L1 (e.g., immunoglobulin monovariate domains) or antibodies used as detection reagents. Various conjugated antibodies are commercially available, including (but are not limited to) antibodies conjugated to Alexa Fluor®, DyLight®, Rhodamine, PE, FITC, and Cy3. Each fluorophore has a unique excitation and fluorescence wavelength. The possible label combinations depend on the wavelength of the lamp or laser used to excite the fluorophore and the available detectors.

[0608] To evaluate the competition between two test conjugates (referred to as A and B) for binding to PD-L1, a dilution series of unlabeled conjugate A is added to cells (e.g., 200,000 cells) along with labeled conjugate B*. The concentration of conjugate B* in the test mixture must be high enough to readily saturate the binding site on PD-L1 expressed on the cells. The concentration of conjugate B* that saturates the binding site for the conjugate on PD-L1 expressed on the cells can be determined by titrating the conjugate B* on PD-L1-expressing cells and determining the EC50 value for binding. To act at a saturation concentration, conjugate B* can be used at 100 times its EC50 concentration.

[0609] Cells can be incubated with a mixture of conjugate A and conjugate B*, washed, and then read using FACS. First, a gate is set to complete cells determined from the scattering profile, and the total amount of channel fluorescence is recorded. Another solution of conjugate B* is also prepared. The conjugate B* in this solution must be in the same buffer and at the same concentration as that in the test mixture (containing conjugates A and B*). This other solution is also added to the cells. After incubation and cell washing, a read can be performed using FACS. First, a gate is set to complete cells determined from the scattering profile, and the total amount of channel fluorescence is recorded. If the fluorescence of cells incubated with the mixture of conjugates A and B* is lower than the fluorescence of cells incubated with conjugate B* alone, it indicates that conjugate A is blocking the binding of conjugate B* to PD-L1 expressed on the cells.

[0610] Cross-blocking immunoglobulins, antibodies, immunoglobulin monovariate domains, polypeptides, or other conjugates refer to those that bind to PD-L1 in the above-mentioned competitive FACS and, during the assay and in the presence of a second conjugate, produce fluorescence ranging from 80% to 0.1% (e.g., 80% to 4%) of the maximum fluorescence (the maximum fluorescence measured for the labeled immunoglobulin, antibody, immunoglobulin monovariate domain, polypeptide, or other conjugate alone), specifically from 75% to 0.1% (e.g., 75% to 4%), and further from 70% to 0.1% (e.g., 70% to 4%).

[0611] Competition for binding of two test conjugates (referred to as A* and B*) to PD-L1 can also be evaluated by labeling both conjugates with different fluorophores and adding them to PD-L1-expressing cells. After incubation and cell washing, readings can be performed using FACS. A gate is set for each fluorophore, and the total channel fluorescence is recorded. A decrease and / or disappearance of fluorescence in either fluorophore indicates that the conjugate is blocking binding to PD-L1 expressed on the cells.

[0612] Other methods for determining whether an immunoglobulin, antibody, immunoglobulin monovariate domain, polypeptide, or other conjugate directed at a target blocks, can block, or competitively binds, as defined herein, are described, for example, by Xiao-Chi Jia et al. (Journal of Immunological Methods 288:91-98, 2004) and Miller et al. (Journal of Immunological Methods 365:118-125, 2011).

[0613] As used herein, the term "potency" is a measure of the biological activity of an agonist such as a polypeptide or ISVD. The potency of an agonist can be determined by any preferred method known in the art, such as those described in the Experiments section, for example. Cell culture-based potency assays are often a preferred format for determining biological activity because they measure the physiological response induced by the agonist and can produce results in a relatively short time. Depending on the mechanism of action of the product, various types of cell-based assays can be used, such as a tetanus toxoid autoantigen-specific co-culture assay or an NFAT reporter assay (further described in the Examples section).

[0614] In contrast, the term "efficacy" measures the maximum intensity of the action itself at the saturated concentration of an agent such as an ISVD or polypeptide. Efficacy indicates the maximum response obtained from the agent. It means the ability of the polypeptide to produce the desired (therapeu...

Claims

1. An immunoglobulin monovariate domain (ISVD) that specifically binds to human PD-L1 and consists of essentially four framework regions (FR1-FR4, respectively) and three complementarity-determining regions (CDR1-CDR3, respectively), and interacts with one or more amino acids selected from the PD-L1 protein amino acids Y56, E58, E60, D62, N63, Q66, H78, and R113.

2. The immunoglobulin monovariate domain (ISVD) according to Embodiment 1, wherein one or more of S51, S53, and N56 (Kabat numbering) in CDR2 and / or one or more of A98, A99, A100, A100a, I100d, and I100g (Kabat numbering) in CDR3 form interaction sites with the epitopes on the PD-L1 protein.

3. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR2 (AbM numbering) is the amino acid sequence X １ X ２ SX ３ SX ４ X ５ NX ６ X ７ (SEQ ID NO: 600), where X １ 、X ２ 、X ３ 、X ４ 、X ５ 、X ６ and X ７ are each independently any amino acid selected and -CDR3 (AbM numbering) is the amino acid sequence X １ X ２ X ３ AAAAX ４ X ５ IX ６ X ７ IX ８ X ９ X １０ (Sequence number 603) consists of, where X １ , X ２ , X ３ , X ４ , X ５ , X ６ , X ７ , X ８ , X ９ and X １０ is any amino acid that is independently selected. Immunoglobulin single variable domain (ISVD).

4. -CDR2 (AbM numbering) is the amino acid sequence X １ X ２ SX ３ SX ４ X ５ NX ６ X ７ (Sequence number 600) consists of, where X １ is selected from A, S, T, P, and G, X ２ is selected from M, L, I, V and C, X ３ is selected from A, S, T, P, and G, X ４ is selected from A, S, T, P, and G, X ５ is selected from R, H, and K, X ６ is selected from A, S, T, P, and G, and X ７ This is selected from N and Q; and -CDR3 (AbM numbering) is the amino acid sequence X １ X ２ X ３ AAAAX ４ X ５ IX ６ X ７ IX ８ X ９ X １０ (Sequence number 603) consists of, where X １ is selected from A, S, T, P, and G, X ２ is selected from A, S, T, P, and G, X ３ is selected from F, Y, and W, X ４ is selected from A, S, T, P, and G, X ５ is selected from A, S, T, P, and G, X ６ is selected from A, S, T, P, and G, X ７ is selected from N, Q, I, L, M, V and C, X ８ is selected from F, Y, and W, X ９ is selected from D and E, and X １０ is selected from F, Y, and W. The immunoglobulin monovariate domain (ISVD) according to claim 3.

5. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 13 or 14; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 13 or 14; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 13 or 14; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 17 or 109; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 109 The immunoglobulin monovariate domain (ISVD) according to claim 3 or 4.

6. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid residue X １ A and G are selected, and amino acid residue X ２ is selected from S and T, and amino acid residue X ３ The amino acid sequence X is selected from N, T, and F. １ ISSX ２ GRX ３ TN (Sequence ID 598); b) Amino acid residue X １ A and G are selected, and amino acid residue X ２ is selected from S and T, and amino acid residue X ３ The amino acid sequence X is selected from N, T, and F. １ ISSX ２ GRX ３ An amino acid sequence having at least 80% amino acid identity with TN (SEQ ID NO: 598); c) Amino acid residue X １ A and G are selected, and amino acid residue X ２ is selected from S and T, and amino acid residue X ３ The amino acid sequence X is selected from N, T, and F. １ ISSX ２ GRX ３ Amino acid sequences having a difference of 3, 2, or 1 amino acid from TN (SEQ ID NO: 598); and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ is selected from A and P, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ is selected from N and I, for the amino acid sequence SSWAAAX １ GTX ２ X ３ X ４ IYDY (SEQ ID NO: 601); e) Amino acid residue X １ is selected from A and P, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from N and I, the amino acid sequence SSWAAAX １ GTX ２ X ３ X ４ an amino acid sequence having at least 80% amino acid identity with IYDY (SEQ ID NO: 601); a) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ The amino acid sequence SSWAAAAX is selected from N and I. １ GTX ２ X ３ X ４ Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of IYDY (SEQ ID NO: 601), The immunoglobulin monovariate domain (ISVD) according to any one of claims 3 to 5.

7. The ISVD according to any one of claims 3 to 6, wherein the amino acid sequences of CDR1 and CDR2 (AbM numbered) have at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequences of CDR1 and CDR2 of an ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NOs: 311 to 324.

8. -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 13; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 14; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 103; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 14; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 103; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 110; or -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 13; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 111; or -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 104; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 112; ISVD according to any one of claims 3 to 17.

9. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 23 or 24; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; c) Amino acid sequences having a difference of 6, 5, 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 23 or 24; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 17 or 109; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 39; Immunoglobulin single variable domain (ISVD).

10. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ The amino acid residue X is selected from S and T. ３ The amino acid residue X is selected from T, N, and F. ４ The amino acid residue X is selected from S and P. ５ is selected from E and K, and amino acid residue X ６ The amino acid sequence X is selected from G, V, and A. １ ISSX ２ GRX ３ TNYADX ４ VX ５ X ６ (Sequence ID 605); b) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ The amino acid residue X is selected from S and T. ３ The amino acid residue X is selected from T, N, and F. ４ The amino acid residue X is selected from S and P. ５ is selected from E and K, and amino acid residue X ６ The amino acid sequence X is selected from G, V, and A. １ ISSX ２ GRX ３ TNYADX ４ VX ５ X ６ An amino acid sequence having at least 80% amino acid identity with (SEQ ID NO: 605); c) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ The amino acid residue X is selected from S and T. ３ The amino acid residue X is selected from T, N, and F. ４ The amino acid residue X is selected from S and P. ５ is selected from E and K, and amino acid residue X ６ The amino acid sequence X is selected from G, V, and A. １ ISSX ２ GRX ３ TNYADX ４ VX ５ X ６ Amino acid sequences having a difference of 3, 2, or 1 amino acid from (SEQ ID NO: 605); and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ The amino acid sequence SSWAAAAX is selected from I and N. １ GTX ２ X ３ X ４ IYDY (Sequence ID 601); e) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ The amino acid sequence SSWAAAAX is selected from I and N. １ GTX ２ X ３ X ４ An amino acid sequence having at least 80% amino acid identity with IYDY (SEQ ID NO: 601); f) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ The amino acid sequence SSWAAAAX is selected from I and N. １ GTX ２ X ３ X ４ Amino acid sequences having a difference of 3, 2, or 1 amino acid from IYDY (SEQ ID NO: 601); The immunoglobulin monovariate domain (ISVD) according to claim 9.

11. The ISVD according to claim 9 or 10, wherein the amino acid sequences of CDR2 and CDR3 (Kabat numbered) have at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequences of CDR2 and CD3 of an ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NOs: 311-324.

12. -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 23; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 122; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 110; or -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 123; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 111; ISVD according to any one of claims 9 to 11.

13. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 11 or 98 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 11 or 98; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 11 or 98; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 13 or 14 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 13 or 14; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 13 or 14; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 17 or 109 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 109, Immunoglobulin single variable domain (ISVD).

14. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid residue X １ The amino acid residue X is selected from T, K, and G. ２ A is selected from G and R, and amino acid residue X ３ The amino acid sequence GRX is selected from T and A. １ FSX ２ NX ３ MG (Sequence ID 596); b) Amino acid residue X １ The amino acid residue X is selected from T, K, and G. ２ A is selected from G and R, and amino acid residue X ３ The amino acid sequence GRX is selected from T and A. １ FSX ２ NX ３ An amino acid sequence having at least 80% amino acid identity with MG (SEQ ID NO: 596); c) Amino acid residue X １ The amino acid residue X is selected from T, K, and G. ２ A is selected from G and R, and amino acid residue X ３ The amino acid sequence GRX is selected from T and A. １ FSX ２ NX ３ Amino acid sequences having a difference of 3, 2, or 1 amino acid from MG (SEQ ID NO: 596); and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ A and G are selected, and amino acid residue X ２ is selected from S and T, and amino acid residue X ３ The amino acid sequence X is selected from N, T, and F. １ ISSX ２ GRX ３ TN (Sequence ID 598); e) Amino acid residue X １ A and G are selected, and amino acid residue X ２ is selected from S and T, and amino acid residue X ３ The amino acid sequence X is selected from N, T, and F. １ ISSX ２ GRX ３ An amino acid sequence having at least 80% amino acid identity with TN (SEQ ID NO: 598); f) Amino acid residue X １ A and G are selected, and amino acid residue X ２ is selected from S and T, and amino acid residue X ３ The amino acid sequence X is selected from N, T, and F. １ ISSX ２ GRX ３ Amino acid sequences having a difference of 3, 2, or 1 amino acid from TN (SEQ ID NO: 598); and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ The amino acid sequence SSWAAAAX is selected from N and I. １ GTX ２ X ３ X ４ IYDY (Sequence ID 601); h) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ The amino acid sequence SSWAAAAX is selected from N and I. １ GTX ２ X ３ X ４ An amino acid sequence having at least 80% amino acid identity with IYDY (SEQ ID NO: 601); i) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ The amino acid sequence SSWAAAAX is selected from N and I. １ GTX ２ X ３ X ４ Amino acid sequences having a difference of 3, 2, or 1 amino acid from IYDY (SEQ ID NO: 601); The immunoglobulin monovariate domain (ISVD) according to claim 13.

15. The ISVD according to claim 13 or 14, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NOs: 311-324.

16. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 11; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 13; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR1 consists of the amino acid sequence of SEQ ID NO: 11; -CDR2 consists of one of the amino acid sequences of SEQ ID NOs: 13 and 14; -CDR3 consists of the amino acid sequence of SEQ ID NO: 17; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 98; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 14; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 98; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 103; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 99; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 103; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 99; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 14; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 11; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 14; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 100; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 103; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 110; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 100; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 13; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 111; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 101; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 104; -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 112; ISVD according to any one of claims 13 to 15.

17. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 22 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 22; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 22; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 23 or 24 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; f) Amino acid sequences having a difference of 6, 5, 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 23 or 24; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 17 or 109 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 17 or 109, Immunoglobulin single variable domain (ISVD).

18. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid residue X １ A is selected from G and R, and amino acid residue X ２ The amino acid sequence X is selected from T and A. １ NX ２ MG (Sequence ID 604); b) Amino acid residue X １ A is selected from G and R, and amino acid residue X ２ The amino acid sequence X is selected from T and A. １ NX ２ An amino acid sequence having at least 80% amino acid identity with MG (SEQ ID NO: 604); c) Amino acid residue X １ A is selected from G and R, and amino acid residue X ２ The amino acid sequence X is selected from T and A. １ NX ２ Amino acid sequences having a difference of 3, 2, or 1 amino acid from MG (SEQ ID NO: 604); and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ The amino acid residue X is selected from S and T. ３ The amino acid residue X is selected from T, N, and F. ４ The amino acid residue X is selected from S and P. ５ is selected from E and K, and amino acid residue X ６ The amino acid sequence X is selected from G, V, and A. １ ISSX ２ GRX ３ TNYADX ４ VX ５ X ６ (Sequence ID 605); e) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ The amino acid residue X is selected from S and T. ３ The amino acid residue X is selected from T, N, and F. ４ The amino acid residue X is selected from S and P. ５ is selected from E and K, and amino acid residue X ６ The amino acid sequence X is selected from G, V, and A. １ ISSX ２ GRX ３ TNYADX ４ VX ５ X ６ An amino acid sequence having at least 80% amino acid identity with (SEQ ID NO: 605); f) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ The amino acid residue X is selected from S and T. ３ The amino acid residue X is selected from T, N, and F. ４ The amino acid residue X is selected from S and P. ５ is selected from E and K, and amino acid residue X ６ The amino acid sequence X is selected from G, V, and A. １ ISSX ２ GRX ３ TNYADX ４ VX ５ X ６ Amino acid sequences having a difference of 3, 2, or 1 amino acid from (SEQ ID NO: 605); and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ The amino acid sequence SSWAAAAX is selected from I and N. １ GTX ２ X ３ X ４ IYDY (Sequence ID 601); h) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ The amino acid sequence SSWAAAAX is selected from I and N. １ GTX ２ X ３ X ４ An amino acid sequence having at least 80% amino acid identity with IYDY (SEQ ID NO: 601); i) Amino acid residue X １ A and P are selected, and amino acid residue X ２ is selected from I, L, and T, and amino acid residue X ３ is selected from A, P, and S, and amino acid residue X ４ The amino acid sequence SSWAAAAX is selected from I and N. １ GTX ２ X ３ X ４ Amino acid sequences having a difference of 3, 2, or 1 amino acid from IYDY (SEQ ID NO: 601); The immunoglobulin monovariate domain (ISVD) according to claim 17.

19. The ISVD according to claim 17 or 18, wherein the amino acid sequence of the CDR (Kabat numbered) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NOs: 311-324.

20. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat numbering) consists of one of the amino acid sequences of SEQ ID NOs. 23 and 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 23; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 24; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 109; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat numbering) consists of the amino acid sequence of sequence number 120; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 17; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 118; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 122; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 110; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 118; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 123; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 111; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 118; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 123; -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 111; ISVD according to any one of claims 17 to 19.

21. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of Sequence ID No. 132; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 132; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 132; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 134 or 136 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 134 or 136; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 134 or 136; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 143 or 145 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 143 or 145; Immunoglobulin single variable domain (ISVD).

22. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); b) Amino acid sequences having at least 80% amino acid identity with amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ is selected from A or G, and amino acid residue X ２ is selected from I, S, and V, and amino acid residue X ３ The amino acid sequence X is selected from T and Q. １ X ２ SWSGSGX ３ Y (Sequence ID 607); e) Amino acid residue X １ is selected from A or G, and amino acid residue X ２ is selected from I, S, and V, and amino acid residue X ３ The amino acid sequence X is selected from T and Q. １ X ２ SWSGSGX ３ An amino acid sequence having at least 80% amino acid identity with Y (SEQ ID NO: 607); f) Amino acid residue X １ is selected from A or G, and amino acid residue X ２ is selected from I, S, and V, and amino acid residue X ３ The amino acid sequence X is selected from T and Q. １ X ２ SWSGSGX ３ Amino acid sequences having a difference of 3, 2, or 1 amino acid from Y (SEQ ID NO: 607); and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ is selected from A, V, R, K, and E, and amino acid residue X ２ A is selected from Q and G, and amino acid residue X ３ is selected from V, K, Q, H, and E, and amino acid residue X ４ The amino acid sequence DX is selected from L and R. １ TSX ２ VX ３ X ４ AVEAYDY (Sequence ID 608); h) Amino acid residue X １ is selected from A, V, R, K, and E, and amino acid residue X ２ A is selected from Q and G, and amino acid residue X ３ is selected from V, K, Q, H, and E, and amino acid residue X ４ The amino acid sequence DX is selected from L and R. １ TSX ２ VX ３ X ４ An amino acid sequence having at least 80% amino acid identity with AVEAYDY (SEQ ID NO: 608); i) Amino acid residue X １ is selected from A, V, R, K, and E, and amino acid residue X ２ A is selected from Q and G, and amino acid residue X ３ is selected from V, K, Q, H, and E, and amino acid residue X ４ The amino acid sequence DX is selected from L and R. １ TSX ２ VX ３ X ４ Amino acid sequences having a difference of 3, 2, or 1 amino acid from AVEAYDY (SEQ ID NO: 608); The immunoglobulin single variable domain (ISVD) according to claim 21.

23. The ISVD according to claim 21 or 22, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NOs. 340 to 379.

24. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 134; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 143; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 144; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 145; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 146; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 147; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 148; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 149; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 150; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 149; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 146; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 145; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 147; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 149; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 150; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 144; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 151; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 137; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 152; The ISVD according to any one of claims 21 to 23.

25. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 159; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 159; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 159; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 160 or 162 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 160 or 162; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 160 or 162; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 143 or 145 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 143 or 145; Immunoglobulin single variable domain (ISVD).

26. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of HYVMG (SEQ ID NO: 159) b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence HYVMG (SEQ ID NO: 159); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence HYVMG (SEQ ID NO: 159); and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ is selected from I, S, and V, and amino acid residue X ３ The amino acid sequence X is selected from T and Q. １ X ２ SWSGSGX ３ YYADSVKG (Sequence ID 610); e) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ is selected from I, S, and V, and amino acid residue X ３ The amino acid sequence X is selected from T and Q. １ X ２ SWSGSGX ３ An amino acid sequence having at least 80% amino acid identity with YYADSVKG (SEQ ID NO: 610); f) Amino acid residue X １ A is selected from G and A, and amino acid residue X ２ is selected from I, S, and V, and amino acid residue X ３ The amino acid sequence X is selected from T and Q. １ X ２ SWSGSGX ３ Amino acid sequences having a difference of 3, 2, or 1 amino acid from YYADSVKG (SEQ ID NO: 610); and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ is selected from A, V, R, K, or E, and amino acid residue X ２ is selected from Q or G, and amino acid residue X ３ is selected from V, K, Q, H, or E, and amino acid residue X ４ Amino acid sequence DX, where R or L is selected. １ TSX ２ VX ３ X ４ AVEAYDY (Sequence ID 608); h) Amino acid residue X １ is selected from A, V, R, K, or E, and amino acid residue X ２ is selected from Q or G, and amino acid residue X ３ is selected from V, K, Q, H, or E, and amino acid residue X ４ Amino acid sequence DX, where R or L is selected. １ TSX ２ VX ３ X ４ An amino acid sequence having at least 80% amino acid identity with AVEAYDY (SEQ ID NO: 608); i) Amino acid residue X １ is selected from A, V, R, K, or E, and amino acid residue X ２ is selected from Q or G, and amino acid residue X ３ is selected from V, K, Q, H, or E, and amino acid residue X ４ Amino acid sequence DX, where R or L is selected. １ TSX ２ VX ３ X ４ Amino acid sequences having a difference of 3, 2, or 1 amino acid from AVEAYDY (SEQ ID NO: 608); The immunoglobulin single variable domain (ISVD) according to claim 25.

27. The ISVD according to claim 25 or 26, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NOs. 340 to 379.

28. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 160; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 143; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 144; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 145; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 147; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 148; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 149; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 150; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 148; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 144; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of sequence number 146; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 166; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 145; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 147; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 149; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 147; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 150; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 151; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 163; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 152; ISVD according to any one of claims 25 to 27.

29. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 169 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 169; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 169; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 171 or 175 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 171 or 175; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 171 or 175; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 192 or 193 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 192 or 193; Immunoglobulin single variable domain (ISVD).

30. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence GSIFTSAVME (SEQ ID NO: 169); b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence GSIFTSAVME (SEQ ID NO: 169); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence GSIFTSAVME (SEQ ID NO: 169); and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ The amino acid sequence RIGSGRX is selected from I, R, H, E, K, T, Q, A, S, P, and G. １ A (Sequence ID 611); e) Amino acid residue X １ The amino acid sequence RIGSGRX is selected from I, R, H, E, K, T, Q, A, S, P, and G. １ An amino acid sequence having at least 80% amino acid identity with A (SEQ ID NO: 611); f) Amino acid residue X １ The amino acid sequence RIGSGRX is selected from I, R, H, E, K, T, Q, A, S, P, and G. １ Amino acid sequences having a difference of 3, 2, or 1 amino acid from A (SEQ ID NO: 611); and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ The amino acid sequence VX is selected from M and I. １ GGYIY (Sequence ID 612); h) Amino acid residue X １ The amino acid sequence VX is selected from M and I. １ An amino acid sequence having at least 80% amino acid identity with GGYIY (SEQ ID NO: 612); i) Amino acid residue X １ The amino acid sequence VX is selected from M and I. １ Amino acid sequences having a difference of 3, 2, or 1 amino acid from GGYIY (SEQ ID NO: 612); The immunoglobulin single variable domain (ISVD) according to claim 29.

31. The ISVD according to claim 29 or 30, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NOs. 420 to 450.

32. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 171; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 192; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 172; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 173; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 174; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 175; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 176; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 177; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 178; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 171; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 179; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 180; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 181; and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 193; ISVD according to claim 29 or 30.

33. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 197 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 197; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 197; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 198 or 202 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 198 or 202; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 198 or 202; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 192 or 193 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 192 or 193; Immunoglobulin single variable domain (ISVD).

34. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SAVME (SEQ ID NO: 197) b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SAVME (SEQ ID NO: 197); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SAVME (SEQ ID NO: 197); and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ is selected from I, R, H, E, K, T, Q, A, S, P, and G, and amino acid residue X ２ The amino acid sequence RIGSGRX is selected from P and V. １ AYX ２ DSVKG (Sequence ID 613); e) Amino acid residue X １ is selected from I, R, H, E, K, T, Q, A, S, P, and G, and amino acid residue X ２ The amino acid sequence RIGSGRX is selected from P and V. １ AYX ２ An amino acid sequence having at least 80% amino acid identity with DSVKG (SEQ ID NO: 613); f) Amino acid residue X １ is selected from I, R, H, E, K, T, Q, A, S, P, and G, and amino acid residue X ２ The amino acid sequence RIGSGRX is selected from P and V. １ AYX ２ Amino acid sequences having a difference of 3, 2, or 1 amino acid from DSVKG (SEQ ID NO: 613); and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ The amino acid sequence VX is selected from M and I. １ GGYIY (Sequence ID 612); h) Amino acid residue X １ The amino acid sequence VX is selected from M and I. １ An amino acid sequence having at least 80% amino acid identity with GGYIY (SEQ ID NO: 612); i) Amino acid residue X １ The amino acid sequence VX is selected from M and I. １ Amino acid sequences having a difference of 3, 2, or 1 amino acid from GGYIY (SEQ ID NO: 612); The immunoglobulin monovariate domain (ISVD) according to claim 33.

35. The ISVD according to claim 33 or 34, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NOs. 420 to 450.

36. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 198; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 192; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 199; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 200; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 201; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 202; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 203; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 204; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 205; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 198; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 206; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 207; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 208; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 209; and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 193; ISVD according to any one of claims 33 to 35.

37. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 218 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 218; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 218; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 222 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 222; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 222; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 234 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 234; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 234; Immunoglobulin single variable domain (ISVD).

38. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid residue X １ The amino acid sequence ERTFX selected from S and G １ SYSMG (Sequence ID 617); b) Amino acid residue X １ The amino acid sequence ERTFX selected from S and G １ An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SYSMG (SEQ ID NO: 617); c) Amino acid residue X １ The amino acid sequence ERTFX selected from S and G １ Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SYSMG (SEQ ID NO: 617); and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ is selected from V and P, and amino acid residue X ２ The amino acid sequence X is selected from G and F. １ IX ２ YRGAGPY (SEQ ID NO: 614); e) Amino acid residue X １ is selected from V and P, and amino acid residue X ２ The amino acid sequence X is selected from G and F. １ IX ２ An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of YRGAGPY (SEQ ID NO: 614); f) Amino acid residue X １ is selected from V and P, and amino acid residue X ２ The amino acid sequence X is selected from G and F. １ IX ２ Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of YRGAGPY (SEQ ID NO: 614); and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ The amino acid sequence selected from A and T is RDGAVASIPQAFX １ S (Sequence ID 615); h) Amino acid residue X １ The amino acid sequence selected from A and T is RDGAVASIPQAFX １ An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of S (SEQ ID NO: 615); i) Amino acid residue X １ The amino acid sequence selected from A and T is RDGAVASIPQAFX １ Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of S (SEQ ID NO: 615); The immunoglobulin monovariate domain (ISVD) according to claim 37.

39. The ISVD according to claim 37 or 38, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NOs. 482 to 497.

40. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO:

218. -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 222, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 234; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO:

218. -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 223, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 234; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO:

218. -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 224, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 234; or -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 219, -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 222, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 235; ISVD according to any one of claims 37 to 40.

41. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 241 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 241; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 241; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 242 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 242; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 242; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 234 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 234; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 234; Immunoglobulin single variable domain (ISVD).

42. Essentially, it consists of four framework areas (FR1 to FR4, respectively) and three complementarity determination areas (CDR1 to CDR3, respectively). -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SYSMG (SEQ ID NO: 241) b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence SYSMG (SEQ ID NO: 241); c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence SYSMG (SEQ ID NO: 241); and -CDR2 (Kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid residue X １ is selected from V and P, and amino acid residue X ２ The amino acid sequence X is selected from G and F. １ IX ２ YRGAGPYYEDSVKG (Sequence ID 616); e) Amino acid residue X １ is selected from V and P, and amino acid residue X ２ The amino acid sequence X is selected from G and F. １ IX ２ An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of YRGAGPYYEDSVKG (SEQ ID NO: 616); f) Amino acid residue X １ is selected from V and P, and amino acid residue X ２ The amino acid sequence X is selected from G and F. １ IX ２ Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of YRGAGPYYEDSVKG (SEQ ID NO: 616); and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid residue X １ The amino acid sequence selected from A and T is RDGAVASIPQAFX １ S (Sequence ID 615); h) Amino acid residue X １ The amino acid sequence selected from A and T is RDGAVASIPQAFX １ An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of S (SEQ ID NO: 615); i) Amino acid residue X １ The amino acid sequence selected from A and T is RDGAVASIPQAFX １ Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of S (SEQ ID NO: 615); The immunoglobulin monovariate domain (ISVD) according to claim 41.

43. The ISVD according to claim 41 or 42, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NOs. 482 to 497.

44. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 242, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 234; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 243, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 234; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 244, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 234; or -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 241, -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 242, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 235; ISVD according to any one of claims 41 to 43.

45. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 256 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 256; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 256; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 267 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 267; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 267; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 279 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 279; i) Amino acid sequences having a difference of 43, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 279; Immunoglobulin single variable domain (ISVD).

46. The ISVD according to claim 45, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

514.

47. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO:

256. -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 267, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 279; ISVD according to claim 45 or 46.

48. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 293 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 293; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 293; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 299 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 299; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 299; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 279 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 279; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 279; Immunoglobulin single variable domain (ISVD).

49. The ISVD according to claim 48, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

514.

50. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO:

293. -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 299, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 279; ISVD according to claim 48 or 49.

51. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 257 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 257; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 257; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 268 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 268; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 268; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 280; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 280; Immunoglobulin single variable domain (ISVD).

52. The ISVD according to claim 51, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

515.

53. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 257, -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 268, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 280; ISVD according to claim 51 or 52.

54. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 294 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 294; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 294; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of Sequence ID No. 300 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 300; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 300; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 280; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 280; Immunoglobulin single variable domain (ISVD).

55. The ISVD according to claim 54, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

515.

56. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 294, -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 300, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 280; ISVD according to claim 54 or 55.

57. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 258 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 258; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 258; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 269 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 269; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 269; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 281 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 281; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 281; Immunoglobulin single variable domain (ISVD).

58. The ISVD according to claim 57, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

516.

59. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 258, -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 269, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 281; ISVD according to claim 57 or 58.

60. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 295 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 295; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 295; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 301 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 301; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 301; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 281 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 281; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 281; Immunoglobulin single variable domain (ISVD).

61. The ISVD according to claim 60, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

516.

62. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO:

295. -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 301, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 281; ISVD according to claim 60 or 61.

63. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 259 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 259; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 259; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 270 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 270; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 270; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 282 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 282; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 282; Immunoglobulin single variable domain (ISVD).

64. The ISVD according to claim 63, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

517.

65. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 259, -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 270, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 282; ISVD according to claim 63 or 64.

66. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 296 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 296; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 296; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 302 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 302; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 302; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 282 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 282; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 282; Immunoglobulin single variable domain (ISVD).

67. The ISVD according to claim 66, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

517.

68. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO:

296. -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 302, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 282; ISVD according to claim 66 or 67.

69. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 260 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 260; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 260; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 271 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 271; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 271; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 283 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 283; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 283; Immunoglobulin single variable domain (ISVD).

70. The ISVD according to claim 69, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

518.

71. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 260, -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 271, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 283; ISVD according to claim 69 or 70.

72. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 297 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 297; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 297; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 303 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 303; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 303; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 283 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 283; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 283; Immunoglobulin single variable domain (ISVD).

73. The ISVD according to claim 72, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even more substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

518.

74. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 297, -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 303, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 283; ISVD according to claim 72 or 73.

75. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (AbM numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 262 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 262; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 262; and -CDR2 (AbM numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 272 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 272; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 272; and -CDR3 (AbM numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 284 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 284; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 284; Immunoglobulin single variable domain (ISVD).

76. The ISVD according to claim 75, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

519.

77. -CDR1 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 262, -CDR2 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 272, and -CDR3 (AbM numbering) consists of the amino acid sequence of SEQ ID NO: 284; ISVD according to claim 75 or 76.

78. An immunoglobulin monovariable domain (ISVD) that specifically binds to human PD-L1 and essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), -CDR1 (Kabat numbering) consists of an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 298 b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 298; c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 298; and -CDR2 (kabat numbering) consists of an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 304 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 304; f) Amino acid sequences having a difference of 6, 5, 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 304; and -CDR3 (Kabat numbering) consists of an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 284 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 284; i) Amino acid sequences having a difference of 4, 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 284; Immunoglobulin single variable domain (ISVD).

79. The ISVD according to claim 78, wherein the amino acid sequence of the CDR (AbM numbering) has at least 80% amino acid sequence identity, more preferably at least 90% amino acid sequence identity, for example 95% amino acid sequence identity or 99% or more amino acid sequence identity, or even substantially 100% amino acid sequence identity, with the amino acid sequence of the CDR of the ISVD having an amino acid sequence selected from SEQ ID NO:

519.

80. -CDR1 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO:

298. -CDR2 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 304, and -CDR3 (Kabat numbering) consists of the amino acid sequence of SEQ ID NO: 284; ISVD according to claim 78 or 79.

81. The amino acid sequence is i) Having 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NOs: 4, 5, 7, 311-525, or 527-528, and for the purpose of determining the degree of amino acid identity, the amino acid residues that form the CDR sequence are not considered; and, ii) Preferably, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104, and 108 according to Kabat numbering are selected from the hallmark residues listed in Table 1. ISVD according to any one of claims 1 to 80.

82. ISVD according to any one of claims 1 to 81, which essentially consists of a heavy chain variable domain sequence derived from a conventional quadruple-chain antibody, or which essentially consists of a heavy chain variable domain sequence derived from a heavy chain antibody.

83. An ISVD according to any one of claims 1 to 82, comprising essentially VHH, humanized VHH, camelid VH, a domain antibody, a single-domain antibody, or dAb.

84. An ISVD according to any one of claims 1 to 83, which is a humanized ISVD selected from the group consisting of SEQ ID NOs: 5, 7, 311-321, 326-336, 341-378, 380-418, 421-449, 451-481, 483-496, 498-512 and 527-528, or a humanized ISVD selected from the group consisting of at least one amino acid sequence of SEQ ID NOs: 5, 7, 311-321, 326-336, 341-378, 380-418, 421-449, 451-481, 483-496, 498-512 and 527-528 and an amino acid sequence having amino acid sequence identity of more than 80%, preferably more than 90%, more preferably more than 95%, for example more than 99%.

85. The ISVD according to any one of claims 1 to 84, wherein the amino acid sequence is selected from the group consisting of SEQ ID NOs: 4, 5, 7, 311 to 525 and 527 to 528.

86. A polypeptide or construct comprising or essentially comprising one or more ISVDs as described in any one of claims 1 to 85, and optionally further comprising one or more other groups, residues, parts or binding units, and optionally linked via one or more linkers.

87. The polypeptide or construct according to claim 86, wherein the one or more other groups, residues, parts or binding units are amino acid sequences.

88. The polypeptide or construct according to claim 86 or 87, wherein the one or more linkers are one or more amino acid sequences.

89. The polypeptide or construct according to any one of claims 86 to 88, wherein the one or more other groups, residues, portions or binding units are immunoglobulin sequences.

90. The polypeptide or construct according to any one of claims 86 to 89, wherein the one or more other groups, residues, parts or binding units are ISVDs.

91. The polypeptide or construct according to any one of claims 86 to 90, wherein the one or more other groups, residues, parts or binding units are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibody, single-domain antibody, and dAb.

92. A polypeptide or construct according to any one of claims 86 to 91, which is a polyvalent construct.

93. A polypeptide or construct according to any one of claims 86 to 92, which is a polyspecific construct.

94. The polypeptide or construct according to any one of claims 86 to 93, wherein the one or more other groups, residues, parts, or binding units confer an extension of the half-life to the polypeptide or construct compared to the ISVD without the one or more other groups, residues, parts, or binding units.

95. The polypeptide or construct according to claim 94, wherein the one or more other groups, residues, moieties or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of polyethylene glycol molecules (PEG), serum proteins or fragments thereof, binding units that specifically bind to serum proteins, Fc moieties, and small proteins or peptides that specifically bind to serum proteins.

96. The polypeptide or construct according to claim 95, wherein the one or more other groups, residues, parts or binding units that confer an extension of the half-life to the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

97. The polypeptide or construct according to claim 95, wherein the one or more other groups, residues, portions, or binding units that give the polypeptide or construct an increased half-life are selected from the group consisting of binding units that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulin (such as IgG).

98. The polypeptide or construct according to claim 97, wherein the one or more other groups, residues, parts or binding units that confer an extension of the half-life to the polypeptide or construct are selected from the group consisting of VHH, humanized VHH, camelid VH, domain antibodies, single-domain antibodies or dAb, which specifically bind to serum albumin (e.g., human serum albumin) or serum immunoglobulin (e.g., IgG).

99. The polypeptide or construct according to claim 98, wherein the one or more other groups, residues, parts or binding units that confer an extension of the half-life to the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

100. The ISVD, which specifically binds to human serum albumin, essentially consists of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively). -CDR1 (AbM numbering) has an amino acid sequence selected from the following: a) Amino acid sequence of SEQ ID NO: 43 b) an amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 43; and c) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 43; and -CDR2 (AbM numbering) has an amino acid sequence selected from the following: d) Amino acid sequence of SEQ ID NO: 45 e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 45; f) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of Sequence ID No. 45; and -CDR3 (AbM numbering) has an amino acid sequence selected from the following: g) Amino acid sequence of SEQ ID NO: 47 h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of Sequence ID No. 47; i) Amino acid sequences having a difference of 3, 2, or 1 amino acid from the amino acid sequence of SEQ ID NO: 47; The polypeptide or construct according to claim 99.

101. The polypeptide or construct according to claim 100, wherein CDR1 consists of the amino acid sequence of SEQ ID NO: 43, CDR2 consists of the amino acid sequence of SEQ ID NO: 45, and CDR3 consists of the amino acid sequence of SEQ ID NO:

47.

102. The polypeptide or construct according to claim 101, wherein the ISVD that specifically binds to human serum albumin is selected from the group consisting of ALB8 (SEQ ID NO: 27), ALB23 (SEQ ID NO: 28), ALBX00001 (SEQ ID NO: 41), and ALB23002 (SEQ ID NO: 42).

103. The polypeptide or construct according to any one of claims 86 to 102, wherein the linker is selected from the group consisting of SEQ ID NOs: 53 to 69.

104. A polypeptide or construct according to any one of claims 86 to 103, further comprising C-terminal elongation.

105. The aforementioned C-terminal elongation is C-terminal elongation (X). ｎ The polypeptide or construct according to claim 104, wherein n is 1 to 10, preferably 1 to 5, for example 1, 2, 3, 4 or 5 (preferably 1 or 2, for example 1), and each X is an amino acid residue independently selected from the group consisting of alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I), preferably independently selected (preferably naturally occurring).

106. A nucleic acid encoding an ISVD according to any one of claims 1 to 85, or a polypeptide according to any one of claims 86 to 105.

107. The nucleic acid according to claim 106, which is a form of genetic construct.

108. A non-human host or host cell that expresses an ISVD according to any one of claims 1 to 85 or a polypeptide according to any one of claims 86 to 105, or is capable of expressing them under appropriate conditions, and / or contains the nucleic acid according to claim 106 or 107.

109. A method for producing an ISVD according to any one of claims 1 to 85 or a polypeptide according to any one of claims 86 to 105, comprising at least the following steps: a) A step of expressing the nucleic acid according to claim 106 or 107 in a suitable non-human host cell or host organism, or in another suitable expression system; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD according to any one of claims 1 to 85 or the polypeptide according to any one of claims 86 to 105. Methods that include...

110. A method for producing an ISVD according to any one of claims 1 to 85 or a polypeptide according to any one of claims 86 to 105, comprising at least the following steps: a) A step of culturing and / or maintaining a non-human host or host cell according to claim 108 under conditions such that the non-human host or host cell expresses and / or produces an ISVD according to at least one of claims 1 to 85, or a polypeptide according to at least one of claims 86 to 105; Depending on the situation, the following may follow: b) A step of isolating and / or purifying the ISVD according to any one of claims 1 to 85 or the polypeptide according to any one of claims 86 to 105. Methods that include...

111. A composition comprising at least one ISVD according to any one of claims 1 to 85, at least one polypeptide or construct according to any one of claims 86 to 105, or at least one nucleic acid according to claim 106 or 107.

112. The composition according to claim 111, which is a pharmaceutical composition.

113. The composition according to claim 111 or 112, further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally one or more further pharmaceutically active polypeptides and / or compounds.

114. An ISVD according to any one of claims 1 to 85, a polypeptide or construct according to any one of claims 86 to 105, or a composition according to any one of claims 111 to 113, for use as a pharmaceutical.

115. An ISVD according to any one of claims 1 to 85, a polypeptide or construct according to any one of claims 86 to 105, or a composition according to any one of claims 111 to 113, for use in the diagnosis, prevention and / or treatment of at least one disease and / or disorder.

116. ISVD according to any one of claims 1 to 85, a polypeptide or construct according to any one of claims 86 to 105, or a composition according to any one of claims 111 to 113, for use in the diagnosis, prevention and / or treatment of PD-L1, its biological or pharmacological activity, and / or at least one disease and / or disorder related to a biological pathway or signaling pathway in which PD-L1 is involved.

117. An ISVD according to any one of claims 1 to 85, a polypeptide or construct according to any one of claims 86 to 105, or a composition according to any one of claims 111 to 113, for use in the diagnosis, prevention and / or treatment of cancer.

118. A method for the diagnosis, prevention and / or treatment of at least one disease and / or disorder, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of at least one disease and / or disorder a pharmaceutically active amount of an ISVD according to any one of claims 1 to 85, a polypeptide or construct according to any one of claims 86 to 105, or a composition according to any one of claims 111 to 113.

119. A method according to claim 118 for the diagnosis, prevention and / or treatment of at least one disease or disorder relating to PD-L1, its biological or pharmacological activity, and / or a biological pathway or signaling pathway in which PD-L1 is involved, comprising administering to a subject in need of diagnosis, prevention and / or treatment of at least one disease or disorder a pharmaceutically active amount of at least one ISVD according to any one of claims 1 to 85, a polypeptide or construct according to any one of claims 86 to 105, or a composition according to any one of claims 111 to 113.

120. A method according to claim 118 or 119 for the diagnosis, prevention and / or treatment of cancer, comprising administering to a subject in need of the diagnosis, prevention and / or treatment of cancer a pharmaceutically active amount of at least one ISVD according to any one of claims 1 to 85, a polypeptide or construct according to any one of claims 86 to 105, or a composition according to any one of claims 111 to 113.

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