PD-L1 targeting immunoglobulin single variable domains

Abstract

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

Description

1. Technical Field This technology relates to immunoglobulin single variable domains (ISVDs) targeting PD-L1, and more particularly to polypeptides and constructs comprising one or more such ISVDs or substantially composed of one or more such ISVDs. It also relates to nucleic acid molecules encoding ISVDs and polypeptides and vectors containing these nucleic acids, and to compositions comprising the ISVD, polypeptide, nucleic acid, or vector. This technology further relates to the use of these products in methods of treating subjects with cancer. This technology also relates to methods of producing these products. 2. Technical Background 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 and are typically expressed on dendritic cells and macrophages. PD-1 and PD-L1 / PD-L2 belong to a family of immune checkpoint proteins that act as co-inhibitors, which can prevent or limit the development of T cell responses. PD-1 / PD-L1 interactions ensure that the immune system is activated only at the appropriate time to minimize the likelihood of chronic autoimmune inflammation. The biology of PD-L1 and its role in cancer immunology and immunotherapy are described in Sun et al. 2018 (Immunity 48: 434) and Salmaninejad et al. 2019 (J. Cell Physiol. 234: 16824).

[0003] Many biological compounds designed to block PD-L1 have been approved for cancer treatment. Atezolizumab (Tecentriq; Roche) is an Fc-modified humanized IgG1 monoclonal antibody approved for urothelial carcinoma, non-small cell lung cancer, small cell lung cancer, hepatocellular carcinoma, melanoma, alveolar soft tissue sarcoma, and triple-negative breast cancer. Devallumab (Imfinzi; Medimus Ltd. / AstraZeneca) is a humanized IgG1κ monoclonal antibody 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 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 biologics. Due to the large size of monoclonal antibodies and the complexity of the tumor microenvironment, their penetration efficiency is low, which has driven the development of small molecule biologics with excellent penetration efficiency. Envafolimab (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 immunoglobulins targeting PD-L1 with single variable domains have been described in WO 2008 / 071447.

[0005] Small molecule biologics targeting PD-L1 still require improvement. These biologics could also be combined with other (cancer) target conjugates in single chains, while maintaining relatively small size, high affinity, potency, and / or efficacy in addition to good manufacturing yields. Furthermore, biologics should demonstrate safety, convenience, patient compliance, and improved quality of life for patients.

[0006] 3. Overview of Existing Technologies In some embodiments, this technology relates to a single variable domain of an immunoglobulin that specifically targets PD-L1. In some embodiments, this technology relates to a high-affinity single variable domain of an immunoglobulin that specifically targets PD-L1. In some embodiments, this technology relates to a high-potency single variable domain of an immunoglobulin that specifically targets PD-L1. In some embodiments, this technology relates to a high-potency single variable domain of an immunoglobulin that specifically targets PD-L1.

[0007] Targeting PD-L1 with small (size-based) biologics (such as monovalent or bivalent immunoglobulins with a single variable domain) while maintaining high affinity, potency, and / or efficacy can be advantageous in certain applications where very small conjugates are desired. Small conjugates can be advantageous, for example, for penetration into the tumor setting, or when additional conjugates targeting other (tumor) targets are involved in the same construct, facilitating the manufacture and further processing of therapeutic compounds.

[0008] This technology provides immunoglobulin monovariable domains that specifically target PD-L1, blocking the interaction between PD1 and PD-L1 (with an IC50 comparable to or better than benchmarks). This technology also provides immunoglobulin monovariable domains that specifically target PD-L1, blocking the interaction between CD80 and PD-L1 (with an IC50 comparable to or better than benchmarks). Furthermore, this technology provides immunoglobulin monovariable domains that specifically target PD-L1, exhibiting potency in bioassays with an IC50 comparable to or better than one or more benchmarks, such as competition with PD1 in flow cytometry or functionality in tetanus toxoid autoantigen-specific co-culture assays (tetanus toxoid (TT) assays).

[0009] This technology specifically binds to human, cynomolgus monkey, and mouse PD-L1 via a single variable domain of immunoglobulin, with a maximum KD difference of up to 5-fold. This technology also provides a method that does not bind to PD-L2 and / or binds at levels higher / lower than 10. -4 M of K D This technology binds to the immunoglobulin monovariable domain of PD-L2. It provides a method that does not bind to other members of the B7 family and / or binds at a level higher / lower than 10. -4 M of K D Combine with the immunoglobulin single variable domain of other members of the B7 family.

[0010] The immunoglobulin monovariable domain of this technology, as well as peptides and constructs containing the immunoglobulin monovariable domain, have shown high potency in competitive assays with PD-1 and CD80 and high potency and efficacy in TT assays, and can be efficiently produced (e.g. in microbial hosts such as Pichia (e.g. Pichia pastoris)).

[0011] Specifically, this technology provides the following embodiments: 3.1 Example Series 7 In some embodiments, the PD-L1-specific ISVD is an ISVD that blocks the interaction between PD-L1 and PD1. The PD-L1-specific ISVD binds to the PD1 binding site on PD-L1. In some embodiments, the PD-L1-specific ISVD interacts with a conformational epitope on PD-L1. In some embodiments, the PD-L1-specific ISVD 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. Blocking this epitope on the PD-L1 protein via a monovalent immunoglobulin monovariable domain has been very effective in preventing PD1 binding and subsequent signal transduction. Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain consisting essentially of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), interacting with one or more amino acids of the PD-L1 protein selected from Y56, E58, E60, D62, N63, Q66, H78 and R113.

[0012] Example 2. The immunoglobulin single variable domain (ISVD) as described in Example 1 interacts with the following amino acids of the PD-L1 protein: Y56, E58, D62, N63, Q66, and H78.

[0013] Example 3. An immunoglobulin single variable domain (ISVD) as described in any one of Examples 1 or 2, which interacts with the following amino acids of the PD-L1 protein: Y56, E58, E60, D62, N63, Q66, H78, and R113.

[0014] Example 4. Immunoglobulin single variable domain (ISVD) as described in any of Examples 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.

[0015] Example 5. Immunoglobulin single variable domain (ISVD) as described in any one of Examples 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.

[0016] Example 6. Immunoglobulin single variable domain (ISVD) as described in any one of Examples 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.

[0017] Example 7. Immunoglobulin single variable domain (ISVD) as described in any of Examples 1 to 6, wherein S51, S53 and N56 (Kabat number) in CDR2 form interaction sites with amino acids of the PD-L1 protein.

[0018] Example 8. Immunoglobulin single variable domain (ISVD) as described in any of Examples 1 to 7, wherein A98, A99, A100, A100a, I100d, and I100g (Kabat number) in CDR3 form interaction sites with amino acids of the PD-L1 protein.

[0019] Example 9. An immunoglobulin single variable domain (ISVD) as described in any of Examples 1 to 8, wherein S51, S53 and N56 (Kabat numbers) in CDR2 and A98, A99, A100, A100a, I100d and I100g (Kabat numbers) in CDR3 form interaction sites with epitopes on the PD-L1 protein.

[0020] Example 10. An immunoglobulin single variable domain (ISVD) as described in any of Examples 1 to 9, wherein S51, S53, and N56 (Kabat number) in CDR2 and A98, A99, A100, A100a, I100d, and I100g (Kabat number) in CDR3 form an interaction site with one or more amino acids selected from Y56, E58, E60, D62, N63, Q66, H78, and R113 of the PD-L1 protein.

[0021] Example 11. An immunoglobulin single variable domain (ISVD) as described in any of Examples 1 to 8, wherein S51, S53 and N56 (Kabat number) in CDR2 and A98, A99, A100, A100a, I100d and I100g (Kabat number) in CDR3 form interaction sites with the following amino acids of the PD-L1 protein: Y56, E58, D62, N63, Q66 and H78.

[0022] Example 12. An immunoglobulin single variable domain (ISVD) as described in any of Examples 1 to 8, wherein S51, S53, and N56 (Kabat number) in CDR2 and A98, A99, A100, A100a, I100d, and I100g (Kabat number) in CDR3 form interaction sites with the following amino acids of the PD-L1 protein: Y56, E58, E60, D62, N63, Q66, H78, and R113.

[0023] Example 13. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR2 (AbM number) consists of the amino acid sequence X1X2SX3SX4X5NX6X7 (SEQ ID NO: 600), where X1, X2, X3, X4, X5, X6 and X7 are any amino acids that are independently selected.

[0024] as well as -CDR3 (AbM number) consists of the amino acid sequence X1X2X3AAAAX4X5IX6X7IX8X9X 10 Composed of (SEQ ID NO: 603), wherein X1, X2, X3, X4, X5, X6, X7, X8, X9 and X 10 Any amino acid selected independently.

[0025] Example 14. Immunoglobulin single variable domain (ISVD) as described in Example 13, wherein -CDR2 (AbM number) consists of the amino acid sequence X1X2SX3SX4X5NX6X7 (SEQ ID NO: 600), wherein 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; as well as -CDR3 (AbM number) consists of the amino acid sequence X1X2X3AAAAX4X5IX6X7IX8X9X 10 (SEQ ID NO: 603) is composed of X1 selected from A, S, T, P and G, X2 selected from A, S, T, P and G, X3 selected from F, Y and W, X4 selected from A, S, T, P and G, X5 selected from A, S, T, P and G, X6 selected from A, S, T, P and G, X7 selected from N, Q, I, L, M, V and C, X8 selected from F, Y and W, X9 selected from D and E, and X 10 Selected from F, Y, and W.

[0026] Example 15. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR2 (AbM number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 13 or 14; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 13 or 14 by 3, 2 or 1 amino acids; as well as -CDR3 (AbM number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 17 or 109 by 4, 3, 2 or 1 amino acid.

[0027] Example 16. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR2 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of 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; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598), wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T, and F; c) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598) by 3, 2 or 1 amino acid, wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T and F; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: d) 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 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) An amino acid sequence that differs from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acids, wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T and the amino acid residue X3 is selected from N and I.

[0028] Example 17. An ISVD as described in Examples 13 to 16, wherein the amino acid sequences of the CDR2 and CDR3 (AbM number) have at least 80% amino acid sequence identity with the amino acid sequences of the CDR2 and CDR3 of an ISVD having amino acid sequences selected from SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 311-324, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0029] Example 18. The ISVD as described in any one of Examples 13 to 17, wherein -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 110. Or one of them -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 111. Or one of them -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 104; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 112.

[0030] Example 19. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR2 (Kabat number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 23 or 24 by 6, 5, 4, 3, 2 or 1 amino acid; as well as -CDR3 (Kabat number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 17 or 39 by 4, 3, 2 or 1 amino acid.

[0031] Example 20. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR2 (Kabat number) consists of an amino acid sequence selected from the following: a) The 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; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of 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) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from S and T, the amino acid residue X3 is selected from T, N and F, the amino acid residue X4 is selected from S and P, the amino acid residue X5 is selected from E and K, and the amino acid residue X6 is selected from G, V and A; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601); wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N; f) An amino acid sequence that differs from the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N.

[0032] Example 21. An ISVD as described in Example 19 or 20, wherein the amino acid sequences of the CDR2 and CDR3 (Kabat numbers) have at least 80% amino acid sequence identity with the amino acid sequences of the CDR2 and CD3 of an ISVD having amino acid sequences selected from SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 311-324, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0033] Example 22. The ISVD as described in any one of Examples 19 to 21, wherein -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 23; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 122; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 110. Or one of them -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 123; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 111.

[0034] Example 23. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 11 or 98; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 11 or 98; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 11 or 98 by 3, 2 or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 13 or 14; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 13 or 14; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 13 or 14 by 3, 2 or 1 amino acids; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 17 or 109; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 17 or 109 by 4, 3, 2 or 1 amino acid.

[0035] Example 24. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) 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; 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) An amino acid sequence that differs from the amino acid sequence GRX1FSX2NX3MG (SEQ ID NO: 596) by 3, 2 or 1 amino acid, wherein the amino acid residue X1 is selected from T, K and G, the amino acid residue X2 is selected from G and R and the amino acid residue X3 is selected from T and A; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598), wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T, and F; f) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T and F; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) 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 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 the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, and the amino acid residue X3 is selected from N and I; i) An amino acid sequence that differs from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acids, wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T and the amino acid residue X3 is selected from N and I.

[0036] Example 25. An ISVD as described in Example 23 or 24, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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 NO: 311-324, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0037] Example 26. The ISVD as described in any one of Examples 23 to 25, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 11; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR1 consists of the amino acid sequence of SEQ ID NO: 11; -CDR2 consists of one of the amino acid sequences of SEQ ID NO: 13 and 14; and -CDR3 consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 98; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 98; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 99; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 99; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 11; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 100; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 110. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 100; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 111. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 101; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 104; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 112.

[0038] Example 27. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 22; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 22; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 22 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 23 or 24; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 23 or 24 by 6, 5, 4, 3, 2 or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 17 or 109; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 17 or 109 by 4, 3, 2 or 1 amino acid.

[0039] Example 28. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of X1NX2MG (SEQ ID NO: 604); wherein the amino acid residue X1 is selected from G and R and wherein the amino acid residue X2 is selected from T and A; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1NX2MG (SEQ ID NO: 604); wherein the amino acid residue X1 is selected from G and R and wherein the amino acid residue X2 is selected from T and A; c) An amino acid sequence that differs from the amino acid sequence X1NX2MG (SEQ ID NO: 604) by 3, 2, or 1 amino acid residue; wherein the amino acid residue X1 is selected from G and R and wherein the amino acid residue X2 is selected from T and A; as well as -CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605), wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from S and T, the amino acid residue X3 is selected from T, N and F, the amino acid residue X4 is selected from S and P, the amino acid residue X5 is selected from E and K, and the amino acid residue X6 is selected from G, V and A; f) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from S and T, the amino acid residue X3 is selected from T, N and F, the amino acid residue X4 is selected from S and P, the amino acid residue X5 is selected from E and K, and the amino acid residue X6 is selected from G, V and A; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of 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; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601); wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N; i) An amino acid sequence that differs from the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N.

[0040] Example 29. An ISVD as described in Example 27 or 28, wherein the amino acid sequence of the CDR (Kabat number) has at least 80% 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 NO: 311-324, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0041] Example 30. The ISVD as described in any one of Examples 27 to 29, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat number) consists of one of the amino acid sequences of SEQ ID NO: 23 and 24; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 23; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 109. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 17. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 118; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 122; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 110. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 118; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 123; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 111. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 118; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 123; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 111.

[0042] Example 31. The ISVD as described in any one of Examples 1 to 30, with the following amino acid sequence i) Having 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 7 and SEQ ID NO: 311-324, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0043] Example 32. The ISVD as described in any one of Examples 1 to 31, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0044] Example 33. The ISVD as described in any one of Examples 1 to 32 is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0045] Example 34. An ISVD as described in any one of Examples 1 to 33, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 5 and SEQ ID NO: 311-321, or selected from the group consisting of an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with at least one of the amino acid sequences of SEQ ID NO: 4, 5 and SEQ ID NO: 311-321.

[0046] Example 35. The ISVD as described in any one of Examples 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: 311-339.

[0047] Example 36. An ISVD as described in any one of Examples 1 to 35, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D The value is 5.10 -9 Up to 10 -11 mol / L or lower, preferably 10 -9 Up to 5.10 -11 mol / L or 5.10 -10 Up to 10 -10 mol / L, or more preferably 10 -10 Up to 10 -11 Moles per liter, as determined by surface plasmon resonance.

[0048] Example 37. An ISVD as described in any one of Examples 1 to 36, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M -1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 6 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0049] Example 38. An ISVD as described in any one of Examples 1 to 37, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -3 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -3 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -4 s -1 and 5.10 -6 s -1 Between, for example, in 5.10 -4 s -1 and 10 -5 s -1 Between, for example, as determined by surface plasmon resonance.

[0050] Example 39. An ISVD as described in any one of Examples 1 to 38, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0051] Example 40. An ISVD as described in any one of Examples 1 to 39, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0052] Example 41. An ISVD as described in any one of Examples 1 to 40, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -8 M or lower, more preferably 10 -9 M or lower, or even 5.10 -10 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 10 -8 Between M, for example, 10 -10 M and 10 -8 Between M, 10 -10 M and 10 -9 Between M or 10 -11 M and 10 -9The effectiveness between M (EC50 value).

[0053] Example 42. An ISVD as described in any one of Examples 1 to 41, wherein the ISVD has at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% efficacy in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0054] Example 43. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 42, or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0055] Example 44. A polypeptide or construct as described in Example 43, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 526, 529, 533, 541, 542, 546, 547, 548, 551, 552, 554, 560, 563, 572, 582, 583, 590 and 593.

[0056] Example 45. A polypeptide or construct as described in any one of Examples 43 to 44, wherein the one or more linkers are one or more amino acid sequences.

[0057] Example 46. A polypeptide or construct as described in any one of Examples 43 to 45, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0058] Example 47. A polypeptide or construct as described in any one of Examples 43 to 46, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0059] Example 48. A polypeptide or construct as described in any one of Examples 43 to 47, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0060] Example 49. A polypeptide or construct as described in any one of Examples 43 to 48, which is a multivalent construct.

[0061] Example 50. A polypeptide or construct as described in any one of Examples 43 to 49, which is a multispecific construct.

[0062] Example 51. A polypeptide or construct as described in any one of Examples 46 to 50, wherein the one or more other groups, residues, portions or binding units provide an increased half-life to the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0063] Example 52. A polypeptide or construct as described in Example 51, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0064] Example 53. A polypeptide or construct as described in Example 52, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0065] Example 54. A polypeptide or construct as described in Example 52, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0066] Example 55. A polypeptide or construct as described in Example 54, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: ISVD, VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody or dAb that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulin (such as IgG).

[0067] Example 56. A polypeptide or construct as described in Example 55, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0068] Example 57. A polypeptide or construct as described in Example 56, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0069] Example 58. A polypeptide or construct as described in Example 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.

[0070] Example 59. The polypeptide or construct as described in Example 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).

[0071] Example 60. A polypeptide or construct as described in any one of Examples 43 to 59, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0072] Example 61. A polypeptide or construct as described in any one of Examples 43 to 60, further comprising a C-terminal extension.

[0073] Example 62. A polypeptide or construct as described in Example 61, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0074] Example 63. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 42 or a polypeptide as described in any one of Examples 43 to 60.

[0075] Example 64. The nucleic acid as described in Example 63, which is in the form of a genetic construct.

[0076] Example 65. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 42, an ISVD or a polypeptide as described in any one of Examples 43 to 62; and / or contains a nucleic acid as described in any one of Examples 63 or 64.

[0077] Example 66. A method for generating an ISVD as described in any one of Examples 1 to 42 or a polypeptide as described in any one of Examples 43 to 62, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 63 or 64 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 42, or the polypeptide as described in any one of Examples 43 to 62.

[0078] Example 67. A method for generating an ISVD as described in any one of Examples 1 to 42 or a polypeptide as described in any one of Examples 43 to 62, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 65 to express and / or produce at least one ISVD as described in any one of Examples 1 to 42 or at least one polypeptide as described in any one of Examples 43 to 62; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 42, or the polypeptide as described in any one of Examples 43 to 62.

[0079] Example 68. A composition comprising at least one ISVD as described in any one of Examples 1 to 42, at least one polypeptide or construct as described in any one of Examples 43 to 62, or at least one nucleic acid as described in any one of Examples 63 or 64.

[0080] Example 69. The composition as described in Example 68 is a pharmaceutical composition.

[0081] Example 70. A composition as described in any one of Examples 68 or 69, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0082] Example 71. An ISVD as described in any one of Examples 1 to 42, a polypeptide or construct as described in any one of Examples 43 to 62, or a composition as described in any one of Examples 68 to 70, used as a pharmaceutical.

[0083] Example 72. An ISVD as described in any one of Examples 1 to 42, a polypeptide or construct as described in any one of Examples 43 to 62, or a composition as described in any one of Examples 68 to 70, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder.

[0084] Example 73. An ISVD as described in any one of Examples 1 to 42, a polypeptide or construct as described in any one of Examples 43 to 62, or a composition as described in any one of Examples 68 to 70, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

[0085] Example 74. An ISVD as described in any one of Examples 1 to 42, a polypeptide or construct as described in any one of Examples 43 to 62, or a composition as described in any one of Examples 68 to 70, for the diagnosis, prevention, and / or treatment of cancer.

[0086] Example 75. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 42, a polypeptide or construct as described in any one of Examples 43 to 62, or a composition as described in any one of Examples 68 to 70.

[0087] Example 76. The method of Example 75, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of the PD-L1, and / or the biological pathways or signaling involving the PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 42, a polypeptide or construct as described in any one of Examples 43 to 62, or a composition as described in any one of Examples 68 to 70.

[0088] Example 77. A method as described in any one of Examples 75 or 76, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 42, a polypeptide or construct as described in any one of Examples 43 to 62, or a composition as described in any one of Examples 68 to 70.

[0089] 3.2 Example Series 42 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 132; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 132; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 132 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 134 or 136; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 134 or 136; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 134 or 136 by 3, 2 or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 143 or 145; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 143 or 145 by 4, 3, 2 or 1 amino acid.

[0090] Example 2. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) Amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); c) An amino acid sequence that differs from the amino acid sequence GGTFRHYVMG (SEQ ID NO: 132) by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1X2SWSGSGX3Y (SEQ ID NO: 607), wherein the amino acid residue X1 is selected from A or G, the amino acid residue X2 is selected from I, S and V, and the amino acid residue X3 is selected from T and Q; f) An amino acid sequence that differs from the amino acid sequence of X1X2SWSGSGX3Y (SEQ ID NO: 607) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A or G, the amino acid residue X2 is selected from I, S and V and the amino acid residue X3 is selected from T and Q. as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of 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; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of 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) An amino acid sequence that differs from the amino acid sequence of DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A, V, R, K and E, the amino acid residue X2 is selected from Q and G, the amino acid residue X3 is selected from V, K, Q, H and E, and the amino acid residue X4 is selected from L and R.

[0091] Example 3. An ISVD as described in Example 1 or 2, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 340-379, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0092] Example 4. The ISVD as described in any one of Examples 1 to 3, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 134; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 143. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 144. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 145. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 146. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 147. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 148. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 149. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 150. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 149. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 146. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 145. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 147. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 149. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 150. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 144. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 151. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 137; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 152.

[0093] Example 5. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 159; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 159; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 159 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 160 or 162; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 160 or 162; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 160 or 162 by 3, 2 or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 143 or 145; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 143 or 145 by 4, 3, 2 or 1 amino acid.

[0094] Example 6. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of HYVMG (SEQ ID NO: 159); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of HYVMG (SEQ ID NO: 159); c) An amino acid sequence that differs from the amino acid sequence of HYVMG (SEQ ID NO: 159) by 3, 2, or 1 amino acid; as well as -CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610); wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from I, S and V, and the amino acid residue X3 is selected from T and Q; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610); wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from I, S and V, and the amino acid residue X3 is selected from T and Q; f) An amino acid sequence that differs from the amino acid sequence of X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from I, S and V, and the amino acid residue X3 is selected from T and Q; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of 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; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608); wherein the amino acid residue X1 is selected from A, V, R, K or E, wherein the amino acid residue X2 is selected from Q or G, wherein the amino acid residue X3 is selected from V, K, Q, H or E, and wherein the amino acid residue X4 is selected from R or L; i) An amino acid sequence that differs from the amino acid sequence of DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A, V, R, K or E, the amino acid residue X2 is selected from Q or G, the amino acid residue X3 is selected from V, K, Q, H or E, and the amino acid residue X4 is selected from R or L.

[0095] Example 7. An ISVD as described in Example 5 or 6, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 340-379, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0096] Example 8. The ISVD as described in any one of Examples 5 to 7, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 160; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 143. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 144. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 145. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 146. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 147. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 148. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 149. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 150.

[0097] Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 148. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 146. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 144. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 146. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 166; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 145. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 147. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 149.

[0098] Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 147. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 150. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 151. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 163; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 152.

[0099] Example 9. The amino acid sequence of the ISVD as described in any one of Examples 1 to 8. i) Having 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 340-379, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0100] Example 10. An ISVD as described in any one of Examples 1 to 9, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0101] Example 11. The ISVD as described in any one of Examples 1 to 10, which is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0102] Example 12. An ISVD as described in any one of Examples 1 to 11, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 341-378, or selected from the group consisting of an amino acid sequence having at least one amino acid sequence of SEQ ID NO: 341-378 having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity.

[0103] Example 13. The ISVD as described in any one of Examples 1 to 12, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 340-419.

[0104] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D The value is 5.10 -9 Up to 10 -11 mol / L or lower, preferably 10 -9 Up to 5.10 -11 mol / L or 5.10-10 Up to 10 -10 mol / L, or more preferably 10 -10 Up to 10 -11 Moles per liter, as determined by surface plasmon resonance.

[0105] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 4 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 10 5 M -1 s -1 and 10 7 M -1 s -1 Between, more preferably in 5.10 5 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 5 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0106] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -3 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -3 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -4 s -1 and 5.10 -6 s -1 Between, for example, in 5.10 -4 s -1 and 10 -5 s -1 Between, for example, as determined by surface plasmon resonance.

[0107] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0108] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0109] Example 19. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -8 M or lower, more preferably 10 -9 M or lower, or even 5.10 -10 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 10 -8 Between M, for example, 10 -10 M and 10 -8 Between M, 10 -10 M and 10 -9 Between M or 10 -11 M and 10 -9 The effectiveness between M (EC50 value).

[0110] Example 20. An ISVD as described in any one of Examples 1 to 19, wherein the ISVD has an efficacy of at least 50%, such as at least 60%, 70%, 80%, 90%, or 95%, in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0111] Example 21. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 20, or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0112] Example 22. A polypeptide or construct as described in Example 21, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 559, 561, 568, 575, 578, 580, 585, 587, 589, 591 and 595.

[0113] Example 23. A polypeptide or construct as described in any one of Examples 21 to 22, wherein the one or more linkers are one or more amino acid sequences.

[0114] Example 24. A polypeptide or construct as described in any one of Examples 21 to 23, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0115] Example 25. A polypeptide or construct as described in any one of Examples 21 to 24, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0116] Example 26. A polypeptide or construct as described in any one of Examples 21 to 25, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0117] Example 27. The polypeptide or construct as described in any one of Examples 21 to 26 is a multivalent construct.

[0118] Example 28. A polypeptide or construct as described in any one of Examples 21 to 27, which is a multispecific construct.

[0119] Example 29. A polypeptide or construct as described in any one of Examples 24 to 28, wherein the one or more other groups, residues, portions or binding units provide an increased half-life for the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0120] Example 30. A polypeptide or construct as described in Example 29, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0121] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0122] Example 32. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0123] Example 33. A polypeptide or construct as described in Example 32, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0124] Example 34. A polypeptide or construct as described in Example 33, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0125] Example 35. A polypeptide or construct as described in Example 34, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0126] Example 36. A polypeptide or construct as described in Example 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] Example 37. The polypeptide or construct as described in Example 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] Example 38. A polypeptide or construct as described in any one of Examples 21 to 37, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0129] Example 39. A polypeptide or construct as described in any one of Examples 21 to 38, further comprising a C-terminal extension.

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

[0131] Example 41. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40.

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

[0133] Example 43. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 20, an ISVD or a polypeptide as described in any one of Examples 21 to 40; and / or contains a nucleic acid as described in any one of Examples 41 or 42.

[0134] Example 44. A method for generating an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 41 or 42 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 20, or the polypeptide as described in any one of Examples 21 to 40.

[0135] Example 45. A method for generating an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 20 or at least one polypeptide as described in any one of Examples 21 to 40; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 20, or the polypeptide as described in any one of Examples 21 to 40.

[0136] Example 46. A composition comprising at least one ISVD as described in any one of Examples 1 to 20, at least one polypeptide or construct as described in any one of Examples 21 to 40, or at least one nucleic acid as described in any one of Examples 41 or 42.

[0137] Example 47. The composition as described in Example 46 is a pharmaceutical composition.

[0138] Example 48. A composition as described in any one of Examples 46 or 47, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0139] Example 49. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, used as a pharmaceutical.

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

[0141] Example 51. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

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

[0143] Example 53. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0144] Example 54. The method of Example 53, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0145] Example 55. A method as described in any one of Examples 53 or 54, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0146] 3.3 Example Series 18 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 169; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 169; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 169 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 171 or 175; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 171 or 175; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 171 or 175 by 3, 2 or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 192 or 193; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 192 or 193 by 3, 2 or 1 amino acid.

[0147] Example 2. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) Amino acid sequence GSIFTSAVME (SEQ ID NO: 169); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence GSIFTSAVME (SEQ ID NO: 169); c) An amino acid sequence that differs from the amino acid sequence GSIFTSAVME (SEQ ID NO: 169) by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of RIGSGGRX1A (SEQ ID NO: 611), wherein the 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 of RIGSGGRX1A (SEQ ID NO: 611), wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P and G; f) An amino acid sequence that differs from the amino acid sequence of RIGSGGRX1A (SEQ ID NO: 611) by 3, 2 or 1 amino acid, wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P and G; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of VX1GGYIY (SEQ ID NO: 612), wherein the amino acid residue X1 is selected from M and I; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of VX1GGYIY (SEQ ID NO: 612), wherein the amino acid residue X1 is selected from M and I; i) An amino acid sequence that differs from the amino acid sequence of VX1GGYIY (SEQ ID NO: 612) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from M and I.

[0148] Example 3. An ISVD as described in Example 1 or 2, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 420-450, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0149] Example 4. The ISVD as described in any one of Examples 1 to 3, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 171; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 192. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 172; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 173; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 174; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 175; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 176; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 177; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 178; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 171; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 179; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 180; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 181; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 193.

[0150] Example 5. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 197; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 197; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 197 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 198 or 202; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 198 or 202; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 198 or 202 by 3, 2 or 1 amino acids; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 192 or 193; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 192 or 193 by 3, 2 or 1 amino acid.

[0151] Example 6. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SAVME (SEQ ID NO: 197); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SAVME (SEQ ID NO: 197); c) An amino acid sequence that differs from the amino acid sequence of SAVME (SEQ ID NO: 197) by 3, 2, or 1 amino acid; as well as -CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613); wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q; A, S, P and G, and wherein the 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 of RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613); wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q; A, S, P and G, and wherein the amino acid residue X2 is selected from P and V; f) An amino acid sequence that differs from the amino acid sequence of RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613) by 3, 2 or 1 amino acid; wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q; A, S, P and G, and wherein the amino acid residue X2 is selected from P and V; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of VX1GGYIY (SEQ ID NO: 612); wherein the amino acid residue X1 is selected from M and I; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of VX1GGYIY (SEQ ID NO: 612); wherein the amino acid residue X1 is selected from M and I; i) An amino acid sequence that differs from the amino acid sequence of VX1GGYIY (SEQ ID NO: 612) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from M and I.

[0152] Example 7. An ISVD as described in Example 5 or 6, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 420-450, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0153] Example 8. The ISVD as described in any one of Examples 5 to 7, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 198; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 192. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 199; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 200; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 201; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 202; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 203; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 204; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 205; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193.

[0154] Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 198; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 206; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 207; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 208; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 209; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 193.

[0155] Example 9. The amino acid sequence of the ISVD as described in any one of Examples 1 to 8. i) Having 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 420-450, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0156] Example 10. An ISVD as described in any one of Examples 1 to 9, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0157] Example 11. The ISVD as described in any one of Examples 1 to 10, which is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0158] Example 12. An ISVD as described in any one of Examples 1 to 11, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 421-449, or selected from the group consisting of an amino acid sequence having an amino acid sequence identity of at least one of the amino acid sequences of SEQ ID NO: 421-449 of greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher.

[0159] Example 13. The ISVD as described in any one of Examples 1 to 12, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 420-481.

[0160] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D The value is 5.10 -9 Up to 10 -11mol / L or lower, preferably 10 -9 Up to 5.10 -11 moles per liter or 10 -9 Up to 10 -10 mol / L, or more preferably 10 -9 Up to 10 -11 Moles per liter, as determined by surface plasmon resonance.

[0161] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 4 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 4 M -1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 5 M -1 s -1 and 10 7 M -1 s -1 Between, for example in 5.10 5 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0162] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -3 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -3 s -1 and 5.10 -6 s -1 Between, more preferably in 10 -4 s -1 and 10 -6 s -1 Between, for example, in 10 -4 s -1 and 10 -5 s -1Between, for example, as determined by surface plasmon resonance.

[0163] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0164] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0165] Example 19. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -8 M or lower, more preferably 10 -9 M or lower, or even 5.10 -10 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 10 -8 Between M, for example, 10 -10 M and 10 -8 Between M, 10 -10 M and 5.10 -9 Between M or 10 -11 M and 5.10 -9 The effectiveness between M (EC50 value).

[0166] Example 20. An ISVD as described in any one of Examples 1 to 19, wherein the ISVD has an efficacy of at least 50%, such as at least 60%, 70%, 80%, 90%, or 95%, in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0167] Example 21. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 20, or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0168] Example 22. A polypeptide or construct as described in Example 21, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 535, 536, 538, 570, 576, 581, 586, 588, 592, and 594.

[0169] Example 23. A polypeptide or construct as described in any one of Examples 21 to 22, wherein the one or more linkers are one or more amino acid sequences.

[0170] Example 24. A polypeptide or construct as described in any one of Examples 21 to 23, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0171] Example 25. A polypeptide or construct as described in any one of Examples 21 to 24, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0172] Example 26. A polypeptide or construct as described in any one of Examples 21 to 25, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0173] Example 27. The polypeptide or construct as described in any one of Examples 21 to 26 is a multivalent construct.

[0174] Example 28. A polypeptide or construct as described in any one of Examples 21 to 27, which is a multispecific construct.

[0175] Example 29. A polypeptide or construct as described in any one of Examples 24 to 28, wherein the one or more other groups, residues, portions or binding units provide an increased half-life for the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0176] Example 30. A polypeptide or construct as described in Example 29, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0177] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0178] Example 32. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0179] Example 33. A polypeptide or construct as described in Example 32, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0180] Example 34. A polypeptide or construct as described in Example 33, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0181] Example 35. A polypeptide or construct as described in Example 34, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0182] Example 36. A polypeptide or construct as described in Example 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] Example 37. The polypeptide or construct as described in Example 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] Example 38. A polypeptide or construct as described in any one of Examples 21 to 37, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0185] Example 39. A polypeptide or construct as described in any one of Examples 21 to 38, further comprising a C-terminal extension.

[0186] Example 40. A polypeptide or construct as described in Example 39, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0187] Example 41. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40.

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

[0189] Example 43. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 20, an ISVD or a polypeptide as described in any one of Examples 21 to 40; and / or contains a nucleic acid as described in any one of Examples 41 or 42.

[0190] Example 44. A method for generating an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 41 or 42 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 20, or the polypeptide as described in any one of Examples 21 to 40.

[0191] Example 45. A method for generating an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 20 or at least one polypeptide as described in any one of Examples 21 to 40; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 20, or the polypeptide as described in any one of Examples 21 to 40.

[0192] Example 46. A composition comprising at least one ISVD as described in any one of Examples 1 to 20, at least one polypeptide or construct as described in any one of Examples 21 to 40, or at least one nucleic acid as described in any one of Examples 41 or 42.

[0193] Example 47. The composition as described in Example 46 is a pharmaceutical composition.

[0194] Example 48. A composition as described in any one of Examples 46 or 47, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0195] Example 49. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, used as a pharmaceutical.

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

[0197] Example 51. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

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

[0199] Example 53. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0200] Example 54. The method of Example 53, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0201] Example 55. A method as described in any one of Examples 53 or 54, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0202] 3.4 Example Series 16 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 218; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 218; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 218 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 222; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 222; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 222 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 234; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 234; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 234 by 3, 2 or 1 amino acids.

[0203] Example 2. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence ERTFX1SYSMG (SEQ ID NO: 617), wherein the 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), wherein the amino acid residue X1 is selected from S and G; c) An amino acid sequence that differs from the amino acid sequence ERTFX1SYSMG (SEQ ID NO: 617) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from S and G; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of X1IX2YRGAGPY (SEQ ID NO: 614), wherein the amino acid residue X1 is selected from V and P, and wherein the 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 of X1IX2YRGAGPY (SEQ ID NO: 614), wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; f) An amino acid sequence that differs from the amino acid sequence of X1IX2YRGAGPY (SEQ ID NO: 614) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615), wherein the 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 of RDGAVASIPQAFX1S (SEQ ID NO: 615), wherein the amino acid residue X1 is selected from A and T; i) An amino acid sequence that differs from the amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A and T.

[0204] Example 3. An ISVD as described in Example 1 or 2, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 482-497, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0205] Example 4. The ISVD as described in any one of Examples 1 to 3, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 218; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 222; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 234. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 218; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 223; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 234. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 218; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 224; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 234. Or one of them -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 219; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 222; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 235.

[0206] Example 5. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 241; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 241; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 241 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 242; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 242; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 242 by 3, 2, or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 234; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 234; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 234 by 3, 2 or 1 amino acids.

[0207] Example 6. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SYSMG (SEQ ID NO: 241); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SYSMG (SEQ ID NO: 241); c) An amino acid sequence that differs from the amino acid sequence of SYSMG (SEQ ID NO: 241) by 3, 2, or 1 amino acid; as well as -CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616); wherein the amino acid residue X1 is selected from V and P, and wherein the 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 of X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616); wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; f) An amino acid sequence that differs from the amino acid sequence of X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616) by 3, 2, or 1 amino acid residues; wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615); wherein the amino acid residue X1 is selected from A and T; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615); wherein the amino acid residue X1 is selected from A and T; i) An amino acid sequence that differs from the amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A and T.

[0208] Example 7. An ISVD as described in Example 5 or 6, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 482-497, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0209] Example 8. The ISVD as described in any one of Examples 5 to 7, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 242; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 234. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 243; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 234. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 244; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 234. Or one of them -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 242; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 235.

[0210] Example 9. The amino acid sequence of the ISVD as described in any one of Examples 1 to 8. i) Having 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 482-497, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0211] Example 10. An ISVD as described in any one of Examples 1 to 9, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0212] Example 11. The ISVD as described in any one of Examples 1 to 10, which is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0213] Example 12. An ISVD as described in any one of Examples 1 to 11, which is a humanized ISVD selected from the group consisting of SEQ ID NO: 483-496, or selected from the group consisting of an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with at least one of the amino acid sequences of SEQ ID NO: 483-496.

[0214] Example 13. The ISVD as described in any one of Examples 1 to 12, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 482-513.

[0215] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D The value is 5.10 -9 Up to 10 -11 mol / L or lower, preferably 10 -9 Up to 5.10 -11 mol / L or 5.10 -10 Up to 10 -10 mol / L, or more preferably 10 -9 Up to 10 -10 Moles per liter, as determined by surface plasmon resonance.

[0216] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M -1 s -1 and 10 7 M -1 s -1 Between, more preferably in 106 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0217] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -3 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -3 s -1 and 5.10 -5 s -1 Between, more preferably at 5.10 -4 s -1 and 5.10 -5 s -1 Between, for example, in 10 -4 s -1 and 10 - 5 s -1 Between, for example, as determined by surface plasmon resonance.

[0218] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0219] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0220] Example 19. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -8 M or lower, more preferably 10 -9 M or lower, or even 5.10 -10 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10-11 M and 10 -8 Between M, for example, 10 -10 M and 10 -8 Between M, 10 -10 M and 10 -9 Between M or 10 -11 M and 10 -9 The effectiveness between M (EC50 value).

[0221] Example 20. An ISVD as described in any one of Examples 1 to 19, wherein the ISVD has an efficacy of at least 50%, such as at least 60%, 70%, 80%, 90%, or 95%, in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0222] Example 21. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 20, or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0223] Example 22. A polypeptide or construct as described in Example 21, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 531, 549, 567, 574, 579, 584.

[0224] Example 23. A polypeptide or construct as described in any one of Examples 21 to 22, wherein the one or more linkers are one or more amino acid sequences.

[0225] Example 24. A polypeptide or construct as described in any one of Examples 21 to 23, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0226] Example 25. A polypeptide or construct as described in any one of Examples 21 to 24, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0227] Example 26. A polypeptide or construct as described in any one of Examples 21 to 25, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0228] Example 27. The polypeptide or construct as described in any one of Examples 21 to 26 is a multivalent construct.

[0229] Example 28. A polypeptide or construct as described in any one of Examples 21 to 27, which is a multispecific construct.

[0230] Example 29. A polypeptide or construct as described in any one of Examples 24 to 28, wherein the one or more other groups, residues, portions or binding units provide an increased half-life for the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0231] Example 30. A polypeptide or construct as described in Example 29, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0232] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0233] Example 32. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0234] Example 33. A polypeptide or construct as described in Example 32, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0235] Example 34. A polypeptide or construct as described in Example 33, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0236] Example 35. A polypeptide or construct as described in Example 34, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0237] Example 36. A polypeptide or construct as described in Example 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.

[0238] Example 37. The polypeptide or construct as described in Example 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).

[0239] Example 38. A polypeptide or construct as described in any one of Examples 21 to 37, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0240] Example 39. A polypeptide or construct as described in any one of Examples 21 to 38, further comprising a C-terminal extension.

[0241] Example 40. A polypeptide or construct as described in Example 39, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0242] Example 41. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40.

[0243] Example 42. The nucleic acid as described in Example 41, which is in the form of a genetic construct.

[0244] Example 43. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 20, an ISVD or a polypeptide as described in any one of Examples 21 to 40; and / or contains a nucleic acid as described in any one of Examples 41 or 42.

[0245] Example 44. A method for generating an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 41 or 42 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 20, or the polypeptide as described in any one of Examples 21 to 40.

[0246] Example 45. A method for generating an ISVD as described in any one of Examples 1 to 20 or a polypeptide as described in any one of Examples 21 to 40, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 20 or at least one polypeptide as described in any one of Examples 21 to 40; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 20, or the polypeptide as described in any one of Examples 21 to 40.

[0247] Example 46. A composition comprising at least one ISVD as described in any one of Examples 1 to 20, at least one polypeptide or construct as described in any one of Examples 21 to 40, or at least one nucleic acid as described in any one of Examples 41 or 42.

[0248] Example 47. The composition as described in Example 46 is a pharmaceutical composition.

[0249] Example 48. A composition as described in any one of Examples 46 or 47, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0250] Example 49. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, used as a pharmaceutical.

[0251] Example 50. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder.

[0252] Example 51. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

[0253] Example 52. An ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47, for the diagnosis, prevention, and / or treatment of cancer.

[0254] Example 53. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0255] Example 54. The method of Example 53, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0256] Example 55. A method as described in any one of Examples 53 or 54, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 20, a polypeptide or construct as described in any one of Examples 21 to 40, or a composition as described in any one of Examples 46 to 47.

[0257] 3.5 Example Series 17 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 256; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 256; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 256 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 267; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 267; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 267 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 279; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 279; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 279 by 43, 2, or 1 amino acid.

[0258] Example 2. An ISVD as described in Example 1, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 514, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0259] Example 3. The ISVD as described in any one of Examples 1 to 2, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 256; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 267; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 279.

[0260] Example 4. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 293; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 293; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 293 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 299; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 299; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 299 by 3, 2, or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 279; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 279; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 279 by 3, 2 or 1 amino acid.

[0261] Example 5. An ISVD as described in Example 4, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 514, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0262] Example 6. The ISVD as described in any one of Examples 4 to 5, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 293; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 299; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 279.

[0263] Example 7. The amino acid sequence of the ISVD as described in any one of Examples 1 to 6. i) It has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 514, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0264] Example 8. An ISVD as described in any one of Examples 1 to 7, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0265] Example 9. The ISVD as described in any one of Examples 1 to 8 is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0266] Example 10. An ISVD as described in any one of Examples 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 an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with the amino acid sequence of SEQ ID NO: 514.

[0267] Example 11. The ISVD as described in any one of Examples 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 514 and 520.

[0268] Example 12. An ISVD as described in any one of Examples 1 to 11, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D ) is 10 -8 Up to 10 -11 Moles per liter or lower, preferably 5.10 -9 Up to 5.10 -11 mol / L or 5.10 -9 Up to 10 -10 mol / L, or more preferably 10 -8 Up to 10 -10 Moles per liter, as determined by surface plasmon resonance.

[0269] Example 13. An ISVD as described in any one of Examples 1 to 12, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M-1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 6 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0270] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -2 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably 5.10 -3 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -3 s -1 and 10 -5 s -1 Between, for example, in 10 -3 s -1 and 10 - 5 s -1 Between, for example, as determined by surface plasmon resonance.

[0271] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0272] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0273] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -7 M or lower, more preferably 5.10-8 M or lower, or even 10 -8 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 10 -7 Between M, for example, 10 -10 M and 10 -7 Between M, 10 -10 M and 5.10 -8 The effectiveness between them (EC50 value).

[0274] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD has at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% efficacy in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0275] Example 19. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 18 or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0276] Example 20. A polypeptide or construct as described in Example 19, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 543, 545, and 548.

[0277] Example 21. A polypeptide or construct as described in any one of Examples 19 to 20, wherein the one or more linkers are one or more amino acid sequences.

[0278] Example 22. A polypeptide or construct as described in any one of Examples 19 to 21, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0279] Example 23. A polypeptide or construct as described in any one of Examples 19 to 22, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0280] Example 24. A polypeptide or construct as described in any one of Examples 19 to 23, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0281] Example 25. A polypeptide or construct as described in any one of Examples 19 to 24, which is a multivalent construct.

[0282] Example 26. A polypeptide or construct as described in any one of Examples 19 to 25, which is a multispecific construct.

[0283] Example 27. A polypeptide or construct as described in any one of Examples 22 to 26, wherein the one or more other groups, residues, portions or binding units provide an increased half-life to the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0284] Example 28. A polypeptide or construct as described in Example 27, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0285] Example 29. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0286] Example 30. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0287] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0288] Example 32. A polypeptide or construct as described in Example 31, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0289] Example 33. A polypeptide or construct as described in Example 32, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0290] Example 34. A polypeptide or construct as described in Example 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.

[0291] Example 35. The polypeptide or construct as described in Example 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).

[0292] Example 36. A polypeptide or construct as described in any one of Examples 19 to 35, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0293] Example 37. A polypeptide or construct as described in any one of Examples 19 to 36, further comprising a C-terminal extension.

[0294] Example 38. A polypeptide or construct as described in Example 37, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0295] Example 39. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38.

[0296] Example 40. The nucleic acid as described in Example 39, in the form of a genetic construct.

[0297] Example 41. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 18, an ISVD or a polypeptide as described in any one of Examples 19 to 39; and / or contains a nucleic acid as described in any one of Examples 39 or 40.

[0298] Example 42. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 39 or 40 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0299] Example 43. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 18 or at least one polypeptide as described in any one of Examples 19 to 38. Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0300] Example 44. A composition comprising at least one ISVD as described in any one of Examples 1 to 18, at least one polypeptide or construct as described in any one of Examples 19 to 38, or at least one nucleic acid as described in any one of Examples 39 or 40.

[0301] Example 45. The composition as described in Example 44 is a pharmaceutical composition.

[0302] Example 46. A composition as described in any one of Examples 44 or 45, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0303] Example 47. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, used as a pharmaceutical.

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

[0305] Example 49. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

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

[0307] Example 51. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0308] Example 52. The method of Example 51, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of the PD-L1, and / or the biological pathways or signaling involving the PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0309] Example 53. A method as described in any one of Examples 51 or 52, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0310] 3.6 Example Series 6 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 257; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 257; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 257 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 268; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 268; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 268 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 280; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 280 by 3, 2 or 1 amino acids.

[0311] Example 2. An ISVD as described in Example 1, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 515, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0312] Example 3. The ISVD as described in any one of Examples 1 to 2, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 257; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 268; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 280.

[0313] Example 4. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 294; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 294; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 294 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 300; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 300; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 300 by 3, 2, or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 280; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 280 by 3, 2 or 1 amino acids.

[0314] Example 5. An ISVD as described in Example 4, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 515, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0315] Example 6. The ISVD as described in any one of Examples 4 to 5, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 294; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 300; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 280.

[0316] Example 7. The amino acid sequence of the ISVD as described in any one of Examples 1 to 6. i) It has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 515, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0317] Example 8. An ISVD as described in any one of Examples 1 to 7, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0318] Example 9. The ISVD as described in any one of Examples 1 to 8 is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0319] Example 10. An ISVD as described in any one of Examples 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 an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with the amino acid sequence of SEQ ID NO: 514.

[0320] Example 11. The ISVD as described in any one of Examples 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 515 and 521.

[0321] Example 12. An ISVD as described in any one of Examples 1 to 11, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D ) is 10 -8 Up to 10 -11 mol / L or lower, preferably 10 -8 Up to 10 -10 mol / L or 5.10 -9 Up to 10 -11 mol / L, or more preferably 5.10 -9 Up to 10 -10 Moles per liter, as determined by surface plasmon resonance.

[0322] Example 13. An ISVD as described in any one of Examples 1 to 12, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M-1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 6 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0323] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -2 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -2 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -3 s -1 and 5.10 -6 s -1 Between, for example, in 5.10 -3 s -1 and 10 -5 s -1 Between, for example, as determined by surface plasmon resonance.

[0324] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0325] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0326] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -8 M or lower, more preferably 10 -9M or lower, or even 5.10 -10 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 10 -8 Between M, for example, 10 -10 M and 10 -8 Between M, 10 -10 M and 10 -9 Between M or 10 -11 M and 10 -9 The effectiveness between M (EC50 value).

[0327] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD has at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% efficacy in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0328] Example 19. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 18 or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0329] Example 20. A polypeptide or construct as described in Example 19, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 530 and 539.

[0330] Example 21. A polypeptide or construct as described in any one of Examples 19 to 20, wherein the one or more linkers are one or more amino acid sequences.

[0331] Example 22. A polypeptide or construct as described in any one of Examples 19 to 21, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0332] Example 23. A polypeptide or construct as described in any one of Examples 19 to 22, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0333] Example 24. A polypeptide or construct as described in any one of Examples 19 to 23, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0334] Example 25. A polypeptide or construct as described in any one of Examples 19 to 24, which is a multivalent construct.

[0335] Example 26. A polypeptide or construct as described in any one of Examples 19 to 25, which is a multispecific construct.

[0336] Example 27. A polypeptide or construct as described in any one of Examples 22 to 26, wherein the one or more other groups, residues, portions or binding units provide an increased half-life to the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0337] Example 28. A polypeptide or construct as described in Example 27, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0338] Example 29. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0339] Example 30. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0340] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0341] Example 32. A polypeptide or construct as described in Example 31, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0342] Example 33. A polypeptide or construct as described in Example 32, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0343] Example 34. A polypeptide or construct as described in Example 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.

[0344] Example 35. The polypeptide or construct as described in Example 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).

[0345] Example 36. A polypeptide or construct as described in any one of Examples 19 to 35, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0346] Example 37. A polypeptide or construct as described in any one of Examples 19 to 36, further comprising a C-terminal extension.

[0347] Example 38. A polypeptide or construct as described in Example 37, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0348] Example 39. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38.

[0349] Example 40. The nucleic acid as described in Example 39, in the form of a genetic construct.

[0350] Example 41. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 18, an ISVD or a polypeptide as described in any one of Examples 19 to 39; and / or contains a nucleic acid as described in any one of Examples 39 or 40.

[0351] Example 42. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 39 or 40 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0352] Example 43. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 18 or at least one polypeptide as described in any one of Examples 19 to 38. Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0353] Example 44. A composition comprising at least one ISVD as described in any one of Examples 1 to 18, at least one polypeptide or construct as described in any one of Examples 19 to 38, or at least one nucleic acid as described in any one of Examples 39 or 40.

[0354] Example 45. The composition as described in Example 44 is a pharmaceutical composition.

[0355] Example 46. A composition as described in any one of Examples 44 or 45, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0356] Example 47. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, used as a pharmaceutical.

[0357] Example 48. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder.

[0358] Example 49. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

[0359] Example 50. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of cancer.

[0360] Example 51. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0361] Example 52. The method of Example 51, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of the PD-L1, and / or the biological pathways or signaling involving the PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0362] Example 53. A method as described in any one of Examples 51 or 52, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0363] 3.7 Example Series 44 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 258; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 258; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 258 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 269; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 269; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 269 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 281; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 281; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 281 by 3, 2 or 1 amino acid.

[0364] Example 2. An ISVD as described in Example 1, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 516, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0365] Example 3. The ISVD as described in any one of Examples 1 to 2, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 258; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 269; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 281.

[0366] Example 4. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 295; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 295; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 295 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 301; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 301; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 301 by 3, 2, or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 281; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 281; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 281 by 3, 2 or 1 amino acid.

[0367] Example 5. An ISVD as described in Example 4, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 516, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0368] Example 6. The ISVD as described in any one of Examples 4 to 5, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 295; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 301; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 281.

[0369] Example 7. The amino acid sequence of the ISVD as described in any one of Examples 1 to 6. i) It has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 516, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0370] Example 8. An ISVD as described in any one of Examples 1 to 7, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0371] Example 9. The ISVD as described in any one of Examples 1 to 8 is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0372] Example 10. An ISVD as described in any one of Examples 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 an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with the amino acid sequence of SEQ ID NO: 516.

[0373] Example 11. The ISVD as described in any one of Examples 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 516 and 522.

[0374] Example 12. An ISVD as described in any one of Examples 1 to 11, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D ) is 10 -8 Up to 10 -11 mol / L or lower, preferably 10 -8 Up to 10 -10 mol / L or 5.10 -9 Up to 10 -11 mol / L, or more preferably 5.10 -9 Up to 10 -10 Moles per liter, as determined by surface plasmon resonance.

[0375] Example 13. An ISVD as described in any one of Examples 1 to 12, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M-1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 6 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0376] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -2 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -2 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -3 s -1 and 5.10 -6 s -1 Between, for example, in 5.10 -3 s -1 and 10 -5 s -1 Between, for example, as determined by surface plasmon resonance.

[0377] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0378] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0379] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -7 M or lower, more preferably 5.10 -8M or lower, or even 10 -8 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 5.10 -8 Between M, for example, 10 -10 M and 5.10 -8 Between M, 10 -10 M and 10 -8 Between M or 10 -11 M and 10 -8 The effectiveness between M (EC50 value).

[0380] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD has at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% efficacy in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0381] Example 19. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 18 or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0382] Example 20. A polypeptide or construct as described in Example 19, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 556 and 562.

[0383] Example 21. A polypeptide or construct as described in any one of Examples 19 to 20, wherein the one or more linkers are one or more amino acid sequences.

[0384] Example 22. A polypeptide or construct as described in any one of Examples 19 to 21, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0385] Example 23. A polypeptide or construct as described in any one of Examples 19 to 22, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0386] Example 24. A polypeptide or construct as described in any one of Examples 19 to 23, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0387] Example 25. A polypeptide or construct as described in any one of Examples 19 to 24, which is a multivalent construct.

[0388] Example 26. A polypeptide or construct as described in any one of Examples 19 to 25, which is a multispecific construct.

[0389] Example 27. A polypeptide or construct as described in any one of Examples 22 to 26, wherein the one or more other groups, residues, portions or binding units provide an increased half-life to the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0390] Example 28. A polypeptide or construct as described in Example 27, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0391] Example 29. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0392] Example 30. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0393] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0394] Example 32. A polypeptide or construct as described in Example 31, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0395] Example 33. A polypeptide or construct as described in Example 32, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0396] Example 34. A polypeptide or construct as described in Example 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.

[0397] Example 35. The polypeptide or construct as described in Example 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).

[0398] Example 36. A polypeptide or construct as described in any one of Examples 19 to 35, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0399] Example 37. A polypeptide or construct as described in any one of Examples 19 to 36, further comprising a C-terminal extension.

[0400] Example 38. A polypeptide or construct as described in Example 37, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0401] Example 39. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38.

[0402] Example 40. The nucleic acid as described in Example 39, in the form of a genetic construct.

[0403] Example 41. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 18, an ISVD or a polypeptide as described in any one of Examples 19 to 39; and / or contains a nucleic acid as described in any one of Examples 39 or 40.

[0404] Example 42. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 39 or 40 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0405] Example 43. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 18 or at least one polypeptide as described in any one of Examples 19 to 38. Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0406] Example 44. A composition comprising at least one ISVD as described in any one of Examples 1 to 18, at least one polypeptide or construct as described in any one of Examples 19 to 38, or at least one nucleic acid as described in any one of Examples 39 or 40.

[0407] Example 45. The composition as described in Example 44 is a pharmaceutical composition.

[0408] Example 46. A composition as described in any one of Examples 44 or 45, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0409] Example 47. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, used as a pharmaceutical.

[0410] Example 48. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder.

[0411] Example 49. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

[0412] Example 50. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of cancer.

[0413] Example 51. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0414] Example 52. The method of Example 51, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of the PD-L1, and / or the biological pathways or signaling involving the PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0415] Example 53. A method as described in any one of Examples 51 or 52, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0416] 3.8 Example Series 43 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 259; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 259; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 259 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 270; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 270; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 270 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 282; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 282; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 282 by 3, 2 or 1 amino acids.

[0417] Example 2. An ISVD as described in Example 1, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 517, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0418] Example 3. The ISVD as described in any one of Examples 1 to 2, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 259; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 270; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 282.

[0419] Example 4. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 296; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 296; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 296 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 302; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 302; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 302 by 3, 2, or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 282; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 282; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 282 by 3, 2 or 1 amino acids.

[0420] Example 5. An ISVD as described in Example 4, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 517, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0421] Example 6. The ISVD as described in any one of Examples 4 to 5, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 296; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 302; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 282.

[0422] Example 7. The amino acid sequence of the ISVD as described in any one of Examples 1 to 6. i) It has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 517, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0423] Example 8. An ISVD as described in any one of Examples 1 to 7, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0424] Example 9. The ISVD as described in any one of Examples 1 to 8 is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0425] Example 10. An ISVD as described in any one of Examples 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 an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with the amino acid sequence of SEQ ID NO: 517.

[0426] Example 11. The ISVD as described in any one of Examples 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 517 and 523.

[0427] Example 12. An ISVD as described in any one of Examples 1 to 11, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D ) is 10 -8 Up to 10 -11 mol / L or lower, preferably 10 -9 Up to 5.10 -11 moles per liter or 10 -8 Up to 10 -10 mol / L, or more preferably 10 -9 Up to 10 -10 Moles per liter, as determined by surface plasmon resonance.

[0428] Example 13. An ISVD as described in any one of Examples 1 to 12, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M-1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 6 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0429] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -2 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -2 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -3 s -1 and 5.10 -6 s -1 Between, for example, in 5.10 -3 s -1 and 10 -5 s -1 Between, for example, as determined by surface plasmon resonance.

[0430] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0431] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0432] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -7 M or lower, more preferably 5.10 -8M or lower, or even 10 -8 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 5.10 -8 Between M, for example, 10 -10 M and 5.10 -8 Between M, 10 -10 M and 10 -8 Between M or 10 -11 M and 10 -8 The effectiveness between M (EC50 value).

[0433] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD has at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% efficacy in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0434] Example 19. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 18 or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0435] Example 20. A polypeptide or construct as described in Example 19, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 557 and 565.

[0436] Example 21. A polypeptide or construct as described in any one of Examples 19 to 20, wherein the one or more linkers are one or more amino acid sequences.

[0437] Example 22. A polypeptide or construct as described in any one of Examples 19 to 21, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0438] Example 23. A polypeptide or construct as described in any one of Examples 19 to 22, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0439] Example 24. A polypeptide or construct as described in any one of Examples 19 to 23, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0440] Example 25. A polypeptide or construct as described in any one of Examples 19 to 24, which is a multivalent construct.

[0441] Example 26. A polypeptide or construct as described in any one of Examples 19 to 25, which is a multispecific construct.

[0442] Example 27. A polypeptide or construct as described in any one of Examples 22 to 26, wherein the one or more other groups, residues, portions or binding units provide an increased half-life to the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0443] Example 28. A polypeptide or construct as described in Example 27, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0444] Example 29. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0445] Example 30. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0446] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0447] Example 32. A polypeptide or construct as described in Example 31, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0448] Example 33. A polypeptide or construct as described in Example 32, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0449] Example 34. A polypeptide or construct as described in Example 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.

[0450] Example 35. The polypeptide or construct as described in Example 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).

[0451] Example 36. A polypeptide or construct as described in any one of Examples 19 to 35, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0452] Example 37. A polypeptide or construct as described in any one of Examples 19 to 36, further comprising a C-terminal extension.

[0453] Example 38. A polypeptide or construct as described in Example 37, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0454] Example 39. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38.

[0455] Example 40. The nucleic acid as described in Example 39, in the form of a genetic construct.

[0456] Example 41. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 18, an ISVD or a polypeptide as described in any one of Examples 19 to 39; and / or contains a nucleic acid as described in any one of Examples 39 or 40.

[0457] Example 42. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 39 or 40 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0458] Example 43. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 18 or at least one polypeptide as described in any one of Examples 19 to 38. Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0459] Example 44. A composition comprising at least one ISVD as described in any one of Examples 1 to 18, at least one polypeptide or construct as described in any one of Examples 19 to 38, or at least one nucleic acid as described in any one of Examples 39 or 40.

[0460] Example 45. The composition as described in Example 44 is a pharmaceutical composition.

[0461] Example 46. A composition as described in any one of Examples 44 or 45, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0462] Example 47. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, used as a pharmaceutical.

[0463] Example 48. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder.

[0464] Example 49. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

[0465] Example 50. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of cancer.

[0466] Example 51. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0467] Example 52. The method of Example 51, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of the PD-L1, and / or the biological pathways or signaling involving the PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0468] Example 53. A method as described in any one of Examples 51 or 52, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0469] 3.9 Example Series 46 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 260; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 260; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 260 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 271; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 271; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 271 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 283; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 283; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 283 by 3, 2 or 1 amino acid.

[0470] Example 2. An ISVD as described in Example 1, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 518, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0471] Example 3. The ISVD as described in any one of Examples 1 to 2, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 260; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 271; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 283.

[0472] Example 4. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 297; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 297; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 297 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 303; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 303; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 303 by 3, 2, or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 283; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 283; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 283 by 3, 2 or 1 amino acid.

[0473] Example 5. An ISVD as described in Example 4, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 518, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0474] Example 6. The ISVD as described in any one of Examples 4 to 5, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 297; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 303; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 283.

[0475] Example 7. The amino acid sequence of the ISVD as described in any one of Examples 1 to 6. i) It has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 518, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0476] Example 8. An ISVD as described in any one of Examples 1 to 7, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0477] Example 9. The ISVD as described in any one of Examples 1 to 8 is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0478] Example 10. An ISVD as described in any one of Examples 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 an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with the amino acid sequence of SEQ ID NO: 518.

[0479] Example 11. The ISVD as described in any one of Examples 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 518 and 524.

[0480] Example 12. An ISVD as described in any one of Examples 1 to 11, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D ) is 10 -8 Up to 10 -11 mol / L or lower, preferably 10 -9 Up to 5.10 -11 moles per liter or 10 -8 Up to 10 -10 mol / L, or more preferably 10 -9 Up to 10 -10 Moles per liter, as determined by surface plasmon resonance.

[0481] Example 13. An ISVD as described in any one of Examples 1 to 12, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M-1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 6 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0482] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -2 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -2 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -3 s -1 and 5.10 -6 s -1 Between, for example, in 5.10 -3 s -1 and 10 -5 s -1 Between, for example, as determined by surface plasmon resonance.

[0483] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0484] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0485] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -7 M or lower, more preferably 5.10 -8M or lower, or even 10 -8 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 5.10 -8 Between M, for example, 10 -10 M and 5.10 -8 Between M, 10 -10 M and 10 -8 Between M or 10 -11 M and 10 -8 The effectiveness between M (EC50 value).

[0486] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD has at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% efficacy in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0487] Example 19. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 18 or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0488] Example 20. A polypeptide or construct as described in Example 19, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as a polypeptide having SEQ ID NO: 558.

[0489] Example 21. A polypeptide or construct as described in any one of Examples 19 to 20, wherein the one or more linkers are one or more amino acid sequences.

[0490] Example 22. A polypeptide or construct as described in any one of Examples 19 to 21, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0491] Example 23. A polypeptide or construct as described in any one of Examples 19 to 22, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0492] Example 24. A polypeptide or construct as described in any one of Examples 19 to 23, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0493] Example 25. A polypeptide or construct as described in any one of Examples 19 to 24, which is a multivalent construct.

[0494] Example 26. A polypeptide or construct as described in any one of Examples 19 to 25, which is a multispecific construct.

[0495] Example 27. A polypeptide or construct as described in any one of Examples 22 to 26, wherein the one or more other groups, residues, portions or binding units provide an increased half-life to the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0496] Example 28. A polypeptide or construct as described in Example 27, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0497] Example 29. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0498] Example 30. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0499] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0500] Example 32. A polypeptide or construct as described in Example 31, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0501] Example 33. A polypeptide or construct as described in Example 32, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0502] Example 34. A polypeptide or construct as described in Example 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.

[0503] Example 35. The polypeptide or construct as described in Example 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).

[0504] Example 36. A polypeptide or construct as described in any one of Examples 19 to 35, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0505] Example 37. A polypeptide or construct as described in any one of Examples 19 to 36, further comprising a C-terminal extension.

[0506] Example 38. A polypeptide or construct as described in Example 37, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0507] Example 39. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38.

[0508] Example 40. The nucleic acid as described in Example 39, in the form of a genetic construct.

[0509] Example 41. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 18, an ISVD or a polypeptide as described in any one of Examples 19 to 39; and / or contains a nucleic acid as described in any one of Examples 39 or 40.

[0510] Example 42. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 39 or 40 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0511] Example 43. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 18 or at least one polypeptide as described in any one of Examples 19 to 38. Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0512] Example 44. A composition comprising at least one ISVD as described in any one of Examples 1 to 18, at least one polypeptide or construct as described in any one of Examples 19 to 38, or at least one nucleic acid as described in any one of Examples 39 or 40.

[0513] Example 45. The composition as described in Example 44 is a pharmaceutical composition.

[0514] Example 46. A composition as described in any one of Examples 44 or 45, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0515] Example 47. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, used as a pharmaceutical.

[0516] Example 48. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder.

[0517] Example 49. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

[0518] Example 50. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of cancer.

[0519] Example 51. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0520] Example 52. The method of Example 51, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of the PD-L1, and / or the biological pathways or signaling involving the PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0521] Example 53. A method as described in any one of Examples 51 or 52, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0522] 3.10 Example Series 45 Example 1. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 262; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 262; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 262 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 272; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 272; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 272 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 284; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 284; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 284 by 3, 2 or 1 amino acid.

[0523] Example 2. An ISVD as described in Example 1, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 519, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0524] Example 3. The ISVD as described in any one of Examples 1 to 2, wherein -CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 262; -CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 272; and -CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO: 284.

[0525] Example 4. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: -CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 298; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 298; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 298 by 3, 2, or 1 amino acid; as well as -CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 304; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 304; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 304 by 6, 5, 4, 3, 2 or 1 amino acid; as well as -CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 284; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 284; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 284 by 4, 3, 2 or 1 amino acid.

[0526] Example 5. An ISVD as described in Example 4, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 519, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

[0527] Example 6. The ISVD as described in any one of Examples 4 to 5, wherein -CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 298; -CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 304; and -CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 284.

[0528] Example 7. The amino acid sequence of the ISVD as described in any one of Examples 1 to 6. i) It has 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 519, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

[0529] Example 8. An ISVD as described in any one of Examples 1 to 7, which is essentially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or essentially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

[0530] Example 9. The ISVD as described in any one of Examples 1 to 8 is basically composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

[0531] Example 10. An ISVD as described in any one of Examples 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 an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with the amino acid sequence of SEQ ID NO: 519.

[0532] Example 11. The ISVD as described in any one of Examples 1 to 10, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 519 and 525.

[0533] Example 12. An ISVD as described in any one of Examples 1 to 11, wherein the ISVD specifically binds to human PD-L1, and the dissociation constant (K) D ) is 10 -8 Up to 10 -11 mol / L or lower, preferably 10 -9 Up to 5.10 -11 moles per liter or 10 -8 Up to 10 -10 mol / L, or more preferably 10 -9 Up to 10 -10 Moles per liter, as determined by surface plasmon resonance.

[0534] Example 13. An ISVD as described in any one of Examples 1 to 12, wherein the ISVD specifically binds to human PD-L1,k on speed at 10 5 M -1 s -1 To about 10 7 M -1 s -1 Between, preferably in 5.10 5 M-1 s -1 and 10 7 M -1 s -1 Between, more preferably in 10 6 M -1 s -1 and 10 7 M -1 s -1 Between, for example, in 10 6 M -1 s -1 and 5.10 6 M -1 s -1 Between, for example, as determined by surface plasmon resonance.

[0535] Example 14. An ISVD as described in any one of Examples 1 to 13, wherein the ISVD specifically binds to human PD-L1,k off speed at 10 -2 s -1 (t1 / 2 = 0.69 s) and 10 -6 s -1 (Providing a near-irreversible complex with t1 / 2 being several days) between, preferably within 10 -2 s -1 and 5.10 -6 s -1 Between, more preferably at 5.10 -3 s -1 and 5.10 -6 s -1 Between, for example, in 5.10 -3 s -1 and 10 -5 s -1 Between, for example, as determined by surface plasmon resonance.

[0536] Example 15. An ISVD as described in any one of Examples 1 to 14, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to other members of the B7 family.

[0537] Example 16. An ISVD as described in any one of Examples 1 to 15, wherein the ISVD specifically binds to human and cynomolgus monkey PD-L1 and does not bind to PD-L2.

[0538] Example 17. An ISVD as described in any one of Examples 1 to 16, wherein the ISVD has a potency (EC50 value) of 10 in TT determinations monitored by IFN-γ generation (e.g., as described in the examples). -7 M or lower, more preferably 5.10 -8M or lower, or even 10 -8 M or lower. For example, in such a TT assay, the immunoglobulin single variable domain of this technique can have 10 -11 M and 5.10 -8 Between M, for example, 10 -10 M and 5.10 -8 Between M, 10 -10 M and 10 -8 Between M or 10 -11 M and 10 -8 The effectiveness between M (EC50 value).

[0539] Example 18. An ISVD as described in any one of Examples 1 to 17, wherein the ISVD has at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% efficacy in a TT measurement monitored via IFN-γ generation (e.g., as described in the examples).

[0540] Example 19. A polypeptide or construct comprising one or more ISVDs as described in any one of Examples 1 to 18 or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

[0541] Example 20. A polypeptide or construct as described in Example 19, wherein one or more other groups, residues, portions or binding units are amino acid sequences, such as one of the polypeptides selected from SEQ ID NO: 566 and 573.

[0542] Example 21. A polypeptide or construct as described in any one of Examples 19 to 20, wherein the one or more linkers are one or more amino acid sequences.

[0543] Example 22. A polypeptide or construct as described in any one of Examples 19 to 21, wherein one or more other groups, residues, portions or binding units are immunoglobulin sequences.

[0544] Example 23. A polypeptide or construct as described in any one of Examples 19 to 22, wherein one or more other groups, residues, portions or binding units are ISVDs.

[0545] Example 24. A polypeptide or construct as described in any one of Examples 19 to 23, wherein one or more other groups, residues, portions or binding units are selected from the group consisting of: VHH, humanized VHH, camelified VH, domain antibody, single-domain antibody and dAb.

[0546] Example 25. A polypeptide or construct as described in any one of Examples 19 to 24, which is a multivalent construct.

[0547] Example 26. A polypeptide or construct as described in any one of Examples 19 to 25, which is a multispecific construct.

[0548] Example 27. A polypeptide or construct as described in any one of Examples 22 to 26, wherein the one or more other groups, residues, portions or binding units provide an increased half-life to the polypeptide or construct compared to an ISVD without the one or more other groups, residues, portions or binding units.

[0549] Example 28. A polypeptide or construct as described in Example 27, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

[0550] Example 29. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

[0551] Example 30. A polypeptide or construct as described in Example 28, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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).

[0552] Example 31. A polypeptide or construct as described in Example 30, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

[0553] Example 32. A polypeptide or construct as described in Example 31, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

[0554] Example 33. A polypeptide or construct as described in Example 32, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: -CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as -CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as -CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

[0555] Example 34. A polypeptide or construct as described in Example 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.

[0556] Example 35. The polypeptide or construct as described in Example 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).

[0557] Example 36. A polypeptide or construct as described in any one of Examples 19 to 35, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

[0558] Example 37. A polypeptide or construct as described in any one of Examples 19 to 36, further comprising a C-terminal extension.

[0559] Example 38. A polypeptide or construct as described in Example 37, wherein the C-terminal extension is a C-terminal extension (X). n , wherein n is 1 to 10, preferably 1 to 5, such as 1, 2, 3, 4 or 5 (and preferably 1 or 2, such as 1); and each X is an independently selected, and preferably independently selected, amino acid residue (preferably naturally occurring) from the group consisting of: alanine (A), glycine (G), valine (V), leucine (L) or isoleucine (I).

[0560] Example 39. A nucleic acid encoding an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38.

[0561] Example 40. The nucleic acid as described in Example 39, in the form of a genetic construct.

[0562] Example 41. A non-human host or host cell that expresses, or is capable of expressing, as described in any one of Examples 1 to 18, an ISVD or a polypeptide as described in any one of Examples 19 to 39; and / or contains a nucleic acid as described in any one of Examples 39 or 40.

[0563] Example 42. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, the method comprising at least the following steps: a) Express the nucleic acid as described in any one of Examples 39 or 40 in a suitable host cell or non-human host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0564] Example 43. A method for generating an ISVD as described in any one of Examples 1 to 18 or a polypeptide as described in any one of Examples 19 to 38, said method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in Example 43 to express and / or produce at least one ISVD as described in any one of Examples 1 to 18 or at least one polypeptide as described in any one of Examples 19 to 38. Optional: b) Isolate and / or purify the ISVD as described in any one of Examples 1 to 18, or the polypeptide as described in any one of Examples 19 to 38.

[0565] Example 44. A composition comprising at least one ISVD as described in any one of Examples 1 to 18, at least one polypeptide or construct as described in any one of Examples 19 to 38, or at least one nucleic acid as described in any one of Examples 39 or 40.

[0566] Example 45. The composition as described in Example 44 is a pharmaceutical composition.

[0567] Example 46. A composition as described in any one of Examples 44 or 45, which is a pharmaceutical composition further comprising at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprising one or more additional pharmaceutically active peptides and / or compounds.

[0568] Example 47. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, used as a pharmaceutical.

[0569] Example 48. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder.

[0570] Example 49. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of PD-L1, and / or the biological pathways or signaling involving PD-L1.

[0571] Example 50. An ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45, for the diagnosis, prevention, and / or treatment of cancer.

[0572] Example 51. A method for diagnosing, preventing, and / or treating at least one disease and / or disorder, the method comprising administering to a subject in need a pharmaceutically active amount of an ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0573] Example 52. The method of Example 51, for the diagnosis, prevention and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of the PD-L1, and / or the biological pathways or signaling involving the PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45.

[0574] Example 53. A method as described in any one of Examples 51 or 52, for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as described in any one of Examples 1 to 18, a polypeptide or construct as described in any one of Examples 19 to 38, or a composition as described in any one of Examples 44 to 45. 4. Attached Figure Descriptions Figure 1: Dissociation rates of anti-PD-L1 clones with unique sequencing against PD-L1 in humans, cynomolgus monkeys, and mice. Dissociation rates were measured using surface plasmon resonance on Proteon. (A) Dot plot; (B) Dot plot showing cross-reactivity.

[0576] Figure 2 FACS competition profiles of some purified anti-PD-L1 VHHs on CHO cells expressing human PD-L1. The competitor is human PD-1.

[0577] Figure 3 FACS competition profiles of some purified anti-PD-L1 VHH on ES-2 cells. The competitor is human PD-1.

[0578] Figure 4 FACS competition profiles of different batches of monovalent ISVD10F11 (ISVD86), bivalent ISVD10F11 (ISVD87), bivalent ISVD36E11 (ISVD84) and baseline 1 on ES-2 cells, with human PD-1 as a competitor.

[0579] Figure 5Compared with baseline 1, IFN-γ production was determined using monovalent and divalent 36E11, 10F11 and 52A01 TT.

[0580] Figure 6 Compared with baseline 5, IFN-γ production was measured in TT using unit price ISVD10F11 (ISVD40).

[0581] Figure 7 : IFN-γ production was determined by TT using monovalent ISVD containing parental CDR sequences and ISVD containing sequence-optimized CDR sequences. (A) A025502469, A025503388 and bivalent baseline; (B) A025502469, A025503357, A025503359; (C) A025501511, A025502467, A02552469; (D) A025501511, A025502470, A02552471.

[0582] Figure 8 The structure of the ISVD10F11 / PD-L1 interaction site was determined using cryo-electron microscopy (cryo-EM). ISVD10F11 was depicted as the top molecule. The extended ring shown below is CDR3. The ring on the right is CDR2 of ISVD10F11. The molecule shown at the bottom is PD-L1. 5. Detailed Implementation 5.1 Definition Unless otherwise stated or defined, all terms used have their ordinary meaning in the art, which is clear to those skilled in the art. For example, refer to standard manuals such as Sambrook et al. (Molecular Cloning: A Laboratory Manual, 2nd Edition, Volumes 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 Edition, Mosby / Elsevier, Edinburgh, 2001), and Roitt et al. (Roitt's Essential Immunology). [Roitt's Foundations of Immunology] (10th edition) Blackwell Publishing, UK, 2001) and Janeway et al. (Immunobiology [Immunobiology] (6th edition) Garland Science Publishing / Churchill Livingstone, New York, 2005) and the general background techniques cited herein.

[0584] As will be clear to those skilled in the art, unless otherwise stated, all methods, steps, techniques and operations not specifically described in detail can be performed in a manner known per se and have been performed in a manner known per se. For example, refer again to the standard manual and general background techniques mentioned in this article and the further references cited therein; and to reviews such as Presta (Adv. Drug Deliv. Rev. . [Advanced Drug Delivery Review] 58 (5-6): 640-56, 2006), Levin and Weiss (Mol. Biosyst. [Molecular Biology Systems] 2(1): 49-57, 2006), Irving et al. (J. Immunol. Methods [Journal of Immunological Methods] 248(1-2): 31-45, 2001), Schmitz et al. (Placenta 21 [Placenta 21] Supplement A: S106-12, 2000), Gonzales et al. (Tumour Biol. [Tumor Biology] 26(1): 31-43, 2005), which describe techniques for protein engineering (such as affinity maturation) and other techniques for improving the specificity and other desired properties of proteins (such as immunoglobulins).

[0585] As used herein, the term “sequence” (e.g., in terms such as “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 that relevant amino acid sequence, unless the context requires a more limited interpretation.

[0586] Amino acid residues will be represented according to standard three-letter or one-letter amino acid codes. Refer to Table A-2 on page 48 of WO 08 / 020079.

[0587] A nucleic acid or amino acid is considered to be "(substantially) isolated" when it has been separated from at least one other component typically associated with it in the source or medium (such as another nucleic acid, another protein / peptide, another biological component or macromolecule, or at least one contaminant, impurity, or trace component), for example, compared to the reaction medium or culture medium from which the nucleic acid or amino acid has been obtained. Specifically, a nucleic acid or amino acid is considered to be "(substantially) isolated" when it has been purified at least 2-fold, particularly at least 10-fold, more particularly at least 100-fold, and up to 1000-fold or more. Nucleic acids or amino acids "in a (substantially) isolated" form are preferably substantially homogeneous, as determined using suitable techniques (such as suitable chromatographic techniques, such as polyacrylamide gel electrophoresis).

[0588] When a nucleotide or amino acid sequence is referred to as "containing" another nucleotide or amino acid sequence, or as "consistently made up of" another nucleotide or amino acid sequence, this may mean that the latter nucleotide or amino acid sequence has been incorporated into the first-mentioned nucleotide or amino acid sequence. More generally, however, this typically means that the first-mentioned nucleotide or amino acid sequence contains within its sequence a segment of nucleotide or amino acid residues having the same nucleotide or amino acid sequence as the latter sequence, regardless of how the first-mentioned sequence has actually been generated or obtained (which may be achieved, for example, by any suitable method described herein). By way of a non-limiting example, when a polypeptide is referred to as containing an immunoglobulin monovariable domain, this may mean that the sequence of said immunoglobulin monovariable domain has been incorporated into the sequence of the polypeptide. More generally, this typically means that the polypeptide contains the sequence of those immunoglobulin monovariable domains within its sequence, regardless of how the polypeptide has been generated or obtained. Furthermore, when a nucleic acid or nucleotide sequence is referred to as containing another nucleotide sequence, the first mentioned nucleic acid or nucleotide sequence is preferably such that when it is 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 in the same reading frame as the first mentioned larger nucleic acid or nucleotide sequence).

[0589] The phrase “(basically) composed of” means that the following nucleic acid sequence or amino acid sequence is identical to or corresponds to a polypeptide (e.g., CDR region; ISVD) having a limited number of amino acid residues added at the N-terminus, C-terminus, or both of the N-terminus and C-terminus of the immunoglobulin monovariable domain, such as 1 to 20 amino acid residues, for example 1 to 10 amino acid residues, and preferably 1 to 6 amino acid residues, such as 1, 2, 3, 4, 5 or 6 amino acid residues.

[0590] 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 corresponding amino acid residues in the second amino acid sequence] by [the total number of amino acid residues in the first amino acid sequence] and multiplying by [100%]. Each deletion, insertion, substitution, or addition of amino acid residues in the second amino acid sequence (compared to the first amino acid sequence) is considered a difference at a single amino acid residue (i.e., a single position). Typically, for the purpose of determining the percentage of "sequence identity" between two amino acid sequences according to the calculation method outlined above, the amino acid sequence with the largest number of amino acid residues is designated as the "first" amino acid sequence, and the other amino acid sequence is designated as the "second" amino acid sequence.

[0591] As used herein, “amino acid difference” refers to the deletion, insertion, or substitution of a single amino acid residue relative to a reference sequence. In one embodiment, “amino acid difference” is substitution. In one embodiment, amino acid substitution is conservative substitution. Such conservative substitution is a substitution in which one amino acid in the following groups (a)-(e) is replaced by another amino acid residue in the same group: (a) small aliphatic, nonpolar, or weakly 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, conservative substitutions are as follows: Ala is substituted with Gly or Ser; Arg is substituted with Lys; Asn is substituted with Gln or His; Asp is substituted with Glu; Cys is substituted with Ser; Gln is substituted with Asn; Glu is substituted with Asp; Gly is substituted with Ala or Pro; His is substituted with Asn or Gln; Ile is substituted with Leu or Val; Leu is substituted with Ile or Val; Lys is substituted with Arg, Gln, or Glu; Met is substituted with Leu, Tyr, or Ile; Phe is substituted with Met, Leu, or Tyr; Ser is substituted with Thr; Thr is substituted with Ser; Trp is substituted with Tyr; Tyr is substituted with Trp; and / or Phe is substituted with Val, Ile, or Leu.

[0592] As used in this specification, “VHH family” refers to a group of VHH sequences that have the same length (i.e., they have the same number of amino acids in their sequences) and in which the amino acid sequence between position 8 and position 106 (according to Kabat numbering) has 89% or more amino acid sequence identity.

[0593] The terms "epitope" and "antigenic determinant" are used interchangeably and refer to a portion of a macromolecule (such as a polypeptide or protein (e.g., PD-L1)) that is recognized by an antigen-binding molecule (such as an immunoglobulin, a conventional antibody, or an immunoglobulin monovariable domain), and more particularly by that antigen-binding site of said molecule. An epitope defines the minimal binding site of an immunoglobulin and thus represents a specific target for the immunoglobulin. The portion of the antigen-binding molecule (e.g., an immunoglobulin, a conventional antibody, or an immunoglobulin monovariable domain) that recognizes the epitope is called the "complementary site."

[0594] These framework regions of variable domains (such as immunoglobulin monovariable domains) form a β-sheet that provides the structural framework for the domain, and these hypervariable sequences correspond to three loops juxtaposed within the folded domain at the outer edge of the β-barrel. These hypervariable loops aggregate to form a single hypervariable site at the molecule's tip. This is the antigen-binding site, antigen-binding site, or complementation site. The three hypervariable loops determine antigen specificity by forming a surface complementary to the antigen and are more commonly referred to as complementarity-determining regions or CDRs (CDR1, CDR2, and CDR3). Thus, the surface of the antibody molecule formed by the juxtaposition of CDRs creates an antigen-binding site.

[0595] Antibodies typically recognize only small regions on the surface of large molecules, such as proteins. The structures recognized by antibodies are called antigenic determinants or epitopes. These sites may consist of amino acids from different parts of a polypeptide chain that are aggregated together by protein folding. Such antigenic determinants are called conformational or discontinuous epitopes because the recognized structures are composed of protein fragments that are discontinuous in the amino acid sequence of the antigen but aggregated together in the three-dimensional structure. In contrast, epitopes composed of a single polypeptide chain segment are called continuous or linear epitopes.

[0596] The binding between an antibody-antigen binding site (complementary site) and its epitope is a reversible, non-covalent interaction. The forces or bonds involved in these non-covalent interactions are electrostatic forces, hydrogen bonds, van der Waals forces, and hydrophobic forces. Electrostatic interactions occur between charged amino acid side chains, such as salt bridges. Interactions also occur between electric dipoles, such as hydrogen bonds, or may involve short-range van der Waals forces. Hydrophobic interactions occur when two hydrophobic surfaces come together to exclude water. For some antigens, hydrophobic interactions may account for the majority of the binding energy.

[0597] Each of these forces contributes to the overall interaction depending on the specific antibody and antigen involved. A significant 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 primarily participate in van der Waals forces and hydrophobic interactions, and sometimes hydrogen bonds. Generally, hydrophobic and van der Waals forces operate over very short ranges and pull two complementary surfaces together: a mound on one surface must fit a valley on the other to achieve good binding. Conversely, electrostatic interactions between charged side chains and hydrogen bonds bridging oxygen and / or nitrogen atoms can adapt to specific features or reactive groups while enhancing the overall interaction.

[0598] The amino acid involved in one of these non-covalent interactions between the antibody-antigen binding site (complementary site) and its epitope is also referred to as the "interaction site" that "forms" the antigen-binding site (complementary site) and the epitope.

[0599] A polypeptide (such as an immunoglobulin, antibody, immunoglobulin monovariable domain, or antigen-binding molecule or fragment thereof) that can "bind" to, or specifically bind to, an epitope, antigen, or protein (or at least a portion, fragment, or epitope thereof), "has affinity for" an epitope, antigen, or protein (or at least a portion, fragment, or epitope thereof), and / or "has specificity for" an epitope, antigen, or protein (or at least a portion, fragment, or epitope thereof) is referred to as "against" or "directed against" the epitope, antigen, or protein, or as a "binding" molecule associated with such epitope, antigen, or protein, or is referred to as "anti" epitope, "anti" antigen, or "anti" protein (e.g., "anti" PD-L1).

[0600] The terms “specificity,” “specific binding,” or “specific binding” refer to the number of different target molecules (such as antigens) from the same organism that a particular binding unit (such as an ISVD) can bind to with a sufficiently high affinity (see below). “Specificity,” “specific binding,” or “specific binding” are used interchangeably herein with “selectivity,” “selective binding,” or “selective binding.” Binding units, such as ISVDs, specifically bind to their designated targets. The specificity / selectivity of a binding unit can be determined based on affinity. Affinity represents the strength or stability of a molecular interaction. Affinity is typically given by KD or the dissociation constant, in units of mol / L (or M). Affinity can also be expressed as the association constant KA, which is equal to 1 / KD, in units of (mol / L).-1 (or M) -1 ).

[0601] Affinity is a measure of the binding strength between a target molecule and its binding sites: the lower the KD value, the stronger the binding strength between the target molecule and the target moiety. Typically, the binding units (e.g., ISVDs) used in this technique will have a KD value of 10. -5 Up to 10 -12 mol / L or lower, 10 -7 Up to 10 -12 moles per liter or lower, or 10 -8 Up to 10 -12 The dissociation constant (KD) of mol / L with its target binding (i.e., association constant (KA)) is 10. 5 Up to 10 12 liters / moles or more, 10 7 Up to 10 12 liters / moles or more, or 10 8 Up to 10 12 (liters / moles). Any value greater than 10 -4 KD value of mol / L (or any value below 10 mol / L) 4 The KA value (in liters per mol) is generally considered to indicate nonspecific binding. KD values ​​considered specific biological interactions (e.g., binding of immunoglobulin sequences to antigens) are typically in the range of 10. -5 mol / L (10000 nM or 10 µM) to 10 -12 In the range of mol / L (0.001 nM or 1 pM) or lower.

[0602] Therefore, specific / selective binding can refer to the use of the same measurement method, such as SPR-binding units (or peptides containing them), at a ratio of 10. -5 Up to 10 -12 KD values ​​of mol / L or lower combined with PD-L1, and with a value greater than 10 -4 The KD value in mol / L is combined with the relevant B7 family member. An example of the relevant B7 family member is PD-L2. Therefore, in embodiments of this technique, ISVD is 10 -5 Up to 10 -12 KD values ​​of mol / L or lower bind to (human) PD-L1 and at a concentration greater than 10 -4 The molar / liter KD value is associated with PD-L2 in the same species.

[0603] Specific binding to a target from a particular species does not preclude the binding unit from specifically binding to a similar target from a different species. For example, specific binding to human PD-L1 does not preclude the binding unit or the polypeptide containing it from specifically binding to PD-L1 from cynomolgus monkeys.

[0604] The specific binding of the binding unit to its designated target can be achieved by any suitable means known per se, including, for example, Scatchard analysis and / or competitive binding assays such as radioimmunoassay (RIA), enzyme immunoassay (EIA) and sandwich competitive assays, and various variants thereof known per se in the art; as well as other techniques further mentioned herein.

[0605] The dissociation constant can be an actual or apparent dissociation constant that is clear to those skilled in the art. Methods for determining the dissociation constant are clear to those skilled in the art and include, for example, the techniques mentioned herein. In this regard, it is also clear that it may be impossible to measure more than 10 -4 moles per liter or 10 -3 moles per liter (e.g., 10) -2 The dissociation constant (in moles per liter). Optionally, 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) using the relationship [KD = 1 / KA].

[0606] An amino acid sequence is described as “cross-reactive” for two different antigens or antigenic determinants (such as, for example, serum albumins from two different mammalian species, such as, for example, human serum albumin and cynomolgus monkey serum albumin, such as, for example, PD-L1 from different mammalian species, such as, for example, human PD-L1, cynomolgus monkey PD-L1 and mouse PD-L1), provided that it is specific for (as defined herein) those different antigens or antigenic determinants.

[0607] The terms “blocking,” “antagonism,” “compete,” “competing,” and “competition” are used interchangeably herein to refer to the ability of an immunoglobulin, antibody, immunoglobulin monovariable domain, peptide, or other binding agent to interfere with the binding of another protein, peptide, ligand, or binding agent to a given target. The extent to which an immunoglobulin, antibody, immunoglobulin monovariable domain, peptide, or other binding agent can interfere with the binding of another ligand to a target, and therefore whether it can be described as “blocking,” can be determined using a competitive binding assay. Particularly suitable quantitative competitive blocking assays are described in the examples and include, for example, fluorescence activated cell sorting (FACS) binding assays using PD-L1 expressed on cells. The degree of blocking can be measured by (reduced) channel fluorescence.

[0608] The following generally describes suitable FACS assays for determining whether or to which immunoglobulins, antibodies, immunoglobulin monovariable domains, peptides, or other binding agents block or are capable of blocking. It should be understood that this assay can be used with any immunoglobulin monovariable domain and peptide described herein. FACS instruments (e.g., FACS Canto; Becton-Dickinson) should be operated according to the manufacturer's recommendations.

[0609] To assess the “blocking” or “competitive” binding of PD-L1 between two binding agents (such as an immunoglobulin monovariable domain and a natural ligand or another binding agent), a FACS competition assay can be performed using cells overexpressing human PD-L1 (such as, for example, Flp-In™-293 cells) and parental cells as a background cell line. Different assays can be used, including, for example, monoclonal ANTI-FLAG® M2 antibody (Sigma-Aldrich, catalog number F1804), monoclonal anti-C-myc antibody (Sigma-Aldrich, catalog number WH0004609M2), and monoclonal anti-HIS-tagged antibody (Sigma-Aldrich, catalog number SAB1305538), each labeled in a different manner. A wide range of fluorophores can be used as markers in flow cytometry (e.g., PE (R-phycoerythrin), 7-aminoactinomycin D (7-AAD), acridine orange, various forms of Alexa Fluor, allophycocyanin (APC), AmCyan, aminocoumarin, APC Cy5, APC Cy7, APC-H7, APC / Alexa Fluor 750, AsRed2, Azami-Green, azurite, BODIPYFL C5-ceramide, BCECF-AM, dioxane DiBAC2(3), BODIPY-FL, calcein, calcein AM, Caroxy-H2DCFDA, strata blue, strata yellow, cell tracer green, sky blue, CFSE, chromomycin A3, CM-H2DCFDA, Cy2, Cy3, Cy3.5, Cy3B, Cy5, Cy5.5, Cy7, CyPet, DAF-FM). DAF-FM diacetate, DAPI, DCFH (2'7' dichlorodihydrofluorescein), DHR, dihydrocalcium chlorophyll AM, dihydrorhodamine, dihydrothiazide, DiLC1(5), DiOC6(3), DiOC7(3), dKeima-red, DRAQ5, Dronpa-green, various forms of DsRed, dTomato, various forms of DyLight, E. coli bioparticles AF488, E2-deep red, E2-orange, EBFP2, ECFP, various forms of eFluor, EGFP, EGFP Emerald Green, eqFP650, eqFP670, ER-Tracer Blue-White DPX, Ethidium Bromide, Express2, EYFP, Fc OxyBurst Green, Fc OxyBurstGreen 123, FITC, Fluorine-3, Fluorine-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, Rhodamine B, various forms of living / dead, Lucifer Yellow, Lucifer Yellow CH, Lyso Tracer Blue, Lyso Tracer Green, Lyso Tracer Red, mAmertrine, Marina Blue, mBanana, mCFP, mCherry, mCitrine, Methoxycoumarin, mHoneyDew, Midoriishi-Cyan, Photomycin, Mito Tracer Deep Red, Mito Tracer Green, Mito Tracer Orange, Mito Tracer Red, MitoFluor Green, mKate (TagFP635) mKate2, mKeima, mKeima-Red, mKO, mKOk, mNeptune, Monochlorobimane, 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 (polydinophytin chlorophyll protein), PerCP Cy5.5. PhiYFP, PhiYFP-m, propidium iodide (PI), various forms of Qdot, 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-sky blue, TagBFP, TagCFP, TagGFP, TagRFP, TagRFP657, TagYFP, tdTomato, Texas Red, Thiazole Orange, TMRE, TMRM, Topaz, TOTO-1, TO-PRO-1, TRITC, TRITC TruRed, TurboFP602, TurboFP635, TurboGFP, TurboRFP, TurboYFP, Venus, Vybrant CycleDye (violet), wild-type GFP, X-Rhodamine, Y66F, Y66H, Y66W, YOYO-1, YPet, ZsGreen1, ZsYellow1, Zymosan A bioparticles AF488 (for more information, please visit: http: / / www.thefcn.org / flow-Fluorochromes). A fluorophore, or simply "fluorophore," is typically attached to an antibody that recognizes PD-L1 (e.g., an immunoglobulin monovariable domain) or an antibody used as a detection reagent. A variety of conjugated antibodies are available, such as (but not limited to) antibodies conjugated to Alexa Fluor®, DyLight®, Rhodamine, PE, FITC, and Cy3. Each fluorophore has a characteristic peak excitation and emission wavelength. The combination of labels that can be used will depend on the wavelength of one or more lamps or lasers used to excite the fluorophore and the available detectors.

[0610] To evaluate the competition between two test binders (referred to as A and B) for binding to PD-L1, a diluted series of cold (unlabeled) binders A and labeled binders B were compared. Add together (e.g., to 200,000 cells). Test the binder B in the mixture. The concentration should be high enough to easily saturate the binding sites on PD-L1 expressed on cells. Bacterial agent B saturates these binding sites on PD-L1 expressed on these cells. The concentration can be obtained by titrating these PD-L1 cells with a series of binding agents B. And the determination of the EC50 value for binding. To operate at saturation concentration, binder B... It can be used at a concentration of 100x EC50.

[0611] Cells were bound with binder A and binder B. After incubating the mixture together and washing the cells, readings can be taken on FACS. First, gates are set for intact cells based on the scattering spectrum, and the total amount of fluorescence in each channel is recorded. Binding agent B was also prepared. A separate solution. The binder B in this solution... It should be used with the test mixture (containing binders A and B) The cells were placed in the same buffer solution and at the same concentration. This separate solution was also added to the cells. After incubation and cell washing, readings could be taken on FACS. First, gates were set for intact cells based on the scattering spectrum, and the total amount of fluorescence in the channels was recorded. This was done with a separate binding agent B. Compared to the fluorescence of cells incubated in solution, the fluorescence of cells incubated with binders A and B was compared. The decrease in fluorescence of cells incubated with the mixture indicates that binder A blocks the flow of the mixture through binder B. It binds to PD-L1 expressed on these cells.

[0612] Cross-blocking immunoglobulins, antibodies, immunoglobulin monovariable domains, peptides, or other binders are such binders that they will bind to PD-L1 in the aforementioned competitive FACS, such that during the assay and in the presence of the second binder, the recorded fluorescence is between 80% and 0.1% (e.g., 80% and 4%) of the maximum fluorescence (measured against individual labeled immunoglobulins, antibodies, immunoglobulin monovariable domains, peptides, or other binders), specifically between 75% and 0.1% (e.g., 75% and 4%) of the maximum fluorescence, and more specifically between 70% and 0.1% (e.g., 70% and 4%) of the maximum fluorescence (as just defined above).

[0613] Two test binders (referred to as A) and B Competition for PD-L1 binding can also be assessed by adding two binding agents (each labeled with a different fluorophore) to these PD-L1-expressing cells. After incubation and cell washing, readings can be taken on FACS. A gating system is set for each fluorophore, and the total amount of fluorescence in the channel is recorded. Decreased and / or absent fluorescence in one of the fluorophores indicates that the binding agent is blocking binding to PD-L1 expressed on the cells.

[0614] Other methods for determining whether immunoglobulins, antibodies, immunoglobulin monovariable domains, peptides, or other binders can block, competitively bind, or are competitive (as defined herein) against a target are described, for example, in 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).

[0615] As used herein, the term "potency" is a measure of the biological activity of a pharmaceutical agent, such as an ISVD or peptide. The potency of a pharmaceutical agent can be determined by any suitable method known in the art, such as those described in the Experimental section. Cell culture-based potency assays are often the preferred form for determining biological activity because they measure the physiological response induced by the agent and produce results within a relatively short timeframe. Based on the product's mechanism of action, various types of cell-based assays can be used, such as, for example, tetanus toxoid autoantigen-specific co-culture assays or NFAT reporter gene assays (as further described in the Examples section).

[0616] Conversely, the "efficacy" of drugs such as ISVDs or peptides measures the maximum intensity of their effect at saturated drug concentrations. Efficacy indicates the maximum response a drug can achieve. It refers to the drug's ability to produce the desired (therapeutic) effect. Drug efficacy can be assessed using in vitro functional assays or in vivo models.

[0617] 5.2 Immunoglobulin Single Variable Domain This technology aims to provide novel drugs for treating PD-L1-related diseases. The inventors unexpectedly discovered that the immunoglobulin single variable domain (ISVD), comprising CDR1 (composed of the amino acid sequence of SEQ ID NO: 11), CDR2 (composed of the amino acid sequence of SEQ ID NO: 13 or 14), and CDR3 (composed of the amino acid sequence of SEQ ID NO: 17), can be used to more forcefully and effectively block PD-L / PD-1 interactions and treat PD-L1-related diseases. The inventors identified a site on PD-L1 involved in the interaction between ISVD and PD-L (blocking of this interaction results in a very effective and powerful inhibition of PD1 / PD-L1 signaling). Blocking this interaction site is sufficient to inhibit PD1 / PD-L1 signaling. Furthermore, this interaction site on PD-L1 or an epitope can be blocked by the single variable domain. Unexpectedly, the inventors determined that only two CDRs in the immunoglobulin single variable domain are involved in this interaction with PD-L1 and the blocking of PD / PD-L1 interactions. Therefore, the inventors observed that only two CDRs (CDR2 and CD3) of the immunoglobulin variable domain were sufficient to block PD / PD-L1 interaction.

[0618] The term "immunoglobulin single variable domain" (ISVD), used interchangeably with "single variable domain," defines an immunoglobulin molecule in which an antigen-binding site is located on and formed by a single immunoglobulin domain. This distinguishes the ISVD from "conventional" immunoglobulins (e.g., monoclonal antibodies) or fragments thereof (e.g., Fab, Fab', F(ab')2, scFv, di-scFv), where two immunoglobulin domains, specifically two variable domains, interact to form an antigen-binding site. Typically, in conventional immunoglobulins, the heavy chain variable domain (VH) and light chain variable domain (VL) interact to form the antigen-binding site. In this case, the complementarity-determining regions (CDRs) of the VH and VL contribute to the formation of the antigen-binding site; a total of six CDRs are involved in the formation of the antigen-binding site.

[0619] Given the above definition, the antigen-binding domains or Fab fragments, F(ab')2 fragments, Fv fragments (such as disulfide-linked Fv or scFv fragments) of conventional 4-chain antibodies (such as IgG, IgM, IgA, IgD, or IgE molecules; known in the art) or the antigen-binding domains of biantibodies derived from such conventional 4-chain antibodies (all known in the art) are generally no...

Claims

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

2. The immunoglobulin single variable domain (ISVD) as described in Example 1, wherein one or more of S51, S53 and N56 (Kabat number) in CDR2 and / or A98, A99, A100, A100a, I100d and I100g (Kabat number) in CDR3 form an interaction site with an epitope on the PD-L1 protein.

3. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the ISVD consisting essentially of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR2 (AbM number) consists of the amino acid sequence X1X2SX3SX4X5NX6X7 (SEQ ID NO: 600), where X1, X2, X3, X4, X5, X6, and X7 are any amino acids chosen independently. as well as - CDR3 (AbM number) consists of the amino acid sequence X1X2X3AAAAX4X5IX6X7IX8X9X 10 Composed of (SEQ ID NO: 603), wherein X1, X2, X3, X4, X5, X6, X7, X8, X9 and X 10 Any amino acid selected independently.

4. The immunoglobulin single variable domain (ISVD) as described in claim 3, wherein... - CDR2 (AbM number) consists of the amino acid sequence X1X2SX3SX4X5NX6X7 (SEQ ID NO: 600), wherein 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; as well as - CDR3 (AbM number) consists of the amino acid sequence X1X2X3AAAAX4X5IX6X7IX8X9X 10 (SEQ ID NO: 603) is composed of X1 selected from A, S, T, P and G, X2 selected from A, S, T, P and G, X3 selected from F, Y and W, X4 selected from A, S, T, P and G, X5 selected from A, S, T, P and G, X6 selected from A, S, T, P and G, X7 selected from N, Q, I, L, M, V and C, X8 selected from F, Y and W, X9 selected from D and E, and X 10 Selected from F, Y, and W.

5. The immunoglobulin single variable domain (ISVD) as described in any one of claims 3 or 4, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR2 (AbM number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 13 or 14; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 13 or 14 by 3, 2 or 1 amino acid; as well as - CDR3 (AbM number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 17 or 109 by 4, 3, 2 or 1 amino acid.

6. The immunoglobulin single variable domain (ISVD) as described in any one of claims 3 to 5, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR2 (AbM number) consists of an amino acid sequence selected from the following: a) 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; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598), wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T, and F; c) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598) by 3, 2 or 1 amino acid, wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T and F; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: d) 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 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; a) An amino acid sequence that differs from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acids, wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T and the amino acid residue X3 is selected from N and I.

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

8. The ISVD as claimed in any one of claims 3 to 17, wherein - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

110. Or one of them - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

111. Or one of them - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 104; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

112.

9. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR2 (Kabat number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 23 or 24 by 6, 5, 4, 3, 2 or 1 amino acid; as well as - CDR3 (Kabat number) 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 that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 17 or 39 by 4, 3, 2 or 1 amino acid.

10. The immunoglobulin single variable domain (ISVD) as described in claim 9, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR2 (Kabat number) consists of an amino acid sequence selected from the following: a) The 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; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of 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) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from S and T, the amino acid residue X3 is selected from T, N and F, the amino acid residue X4 is selected from S and P, the amino acid residue X5 is selected from E and K, and the amino acid residue X6 is selected from G, V and A; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601); wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N; f) An amino acid sequence that differs from the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N.

11. The ISVD of any one of claims 9 or 10, wherein the amino acid sequences of the CDR2 and CDR3 (Kabat numbers) have at least 80% amino acid sequence identity with the amino acid sequences of the CDR2 and CD3 of the ISVD having amino acid sequences selected from SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 311-324, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

12. The ISVD as claimed in any one of claims 9 to 11, wherein - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 23; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 122; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

110. Or one of them - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 123; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

111.

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

14. The immunoglobulin single variable domain (ISVD) as described in claim 13, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) 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; 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) An amino acid sequence that differs from the amino acid sequence GRX1FSX2NX3MG (SEQ ID NO: 596) by 3, 2 or 1 amino acid, wherein the amino acid residue X1 is selected from T, K and G, the amino acid residue X2 is selected from G and R and the amino acid residue X3 is selected from T and A; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598), wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T, and F; f) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TN (SEQ ID NO: 598) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A and G, the amino acid residue X2 is selected from S and T, and the amino acid residue X3 is selected from N, T and F; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) 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 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 the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, and the amino acid residue X3 is selected from N and I; i) An amino acid sequence that differs from the amino acid sequence SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acids, wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T and the amino acid residue X3 is selected from N and I.

15. The ISVD of any one of claims 13 or 14, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having amino acid sequences selected from SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 311-324, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

16. The ISVD as claimed in any one of claims 13 to 15, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 11; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR1 consists of the amino acid sequence of SEQ ID NO: 11; - CDR2 consists of one of the amino acid sequences of SEQ ID NO: 13 and 14; and - CDR3 consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 98; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 98; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 99; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 99; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 11; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 14; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 100; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 103; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

110. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 100; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 13; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

111. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 101; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 104; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

112.

17. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 22; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 22; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 22 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 23 or 24; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 23 or 24; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 23 or 24 by 6, 5, 4, 3, 2 or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 17 or 109; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 17 or 109; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 17 or 109 by 4, 3, 2 or 1 amino acid.

18. The immunoglobulin single variable domain (ISVD) as described in claim 17, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of X1NX2MG (SEQ ID NO: 604); wherein the amino acid residue X1 is selected from G and R and wherein the amino acid residue X2 is selected from T and A; b) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1NX2MG (SEQ ID NO: 604); wherein the amino acid residue X1 is selected from G and R and wherein the amino acid residue X2 is selected from T and A; c) An amino acid sequence that differs from the amino acid sequence X1NX2MG (SEQ ID NO: 604) by 3, 2, or 1 amino acid residue; wherein the amino acid residue X1 is selected from G and R and wherein the amino acid residue X2 is selected from T and A; as well as - CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605), wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from S and T, the amino acid residue X3 is selected from T, N and F, the amino acid residue X4 is selected from S and P, the amino acid residue X5 is selected from E and K, and the amino acid residue X6 is selected from G, V and A; f) An amino acid sequence that differs from the amino acid sequence of X1ISSX2GRX3TNYADX4VX5X6 (SEQ ID NO: 605) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from S and T, the amino acid residue X3 is selected from T, N and F, the amino acid residue X4 is selected from S and P, the amino acid residue X5 is selected from E and K, and the amino acid residue X6 is selected from G, V and A; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of 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; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601); wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N; i) An amino acid sequence that differs from the amino acid sequence of SSWAAAX1GTX2X3X4IYDY (SEQ ID NO: 601) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A and P, the amino acid residue X2 is selected from I, L and T, the amino acid residue X3 is selected from A, P and S, and the amino acid residue X4 is selected from I and N.

19. The ISVD of any one of claims 17 or 18, wherein the amino acid sequence of these CDRs (Kabat numbers) has at least 80% amino acid sequence identity with the amino acid sequence of the CDR of the ISVD having amino acid sequences selected from SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 311-324, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

20. The ISVD as claimed in any one of claims 17 to 19, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; - CDR2 (Kabat number) consists of one of the amino acid sequences of SEQ ID NO: 23 and 24; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 23; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 24; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

109. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 22; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 120; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

17. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 118; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 122; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

110. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 118; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 123; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

111. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 118; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 123; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

111.

21. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 132; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 132; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 132 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 134 or 136; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 134 or 136; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 134 or 136 by 3, 2 or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 143 or 145; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 143 or 145 by 4, 3, 2 or 1 amino acid.

22. The immunoglobulin single variable domain (ISVD) as described in claim 21, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) Amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence GGTFRHYVMG (SEQ ID NO: 132); c) An amino acid sequence that differs from the amino acid sequence GGTFRHYVMG (SEQ ID NO: 132) by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1X2SWSGSGX3Y (SEQ ID NO: 607), wherein the amino acid residue X1 is selected from A or G, the amino acid residue X2 is selected from I, S and V, and the amino acid residue X3 is selected from T and Q; f) An amino acid sequence that differs from the amino acid sequence of X1X2SWSGSGX3Y (SEQ ID NO: 607) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A or G, the amino acid residue X2 is selected from I, S and V and the amino acid residue X3 is selected from T and Q; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of 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; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of 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) An amino acid sequence that differs from the amino acid sequence of DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A, V, R, K and E, the amino acid residue X2 is selected from Q and G, the amino acid residue X3 is selected from V, K, Q, H and E, and the amino acid residue X4 is selected from L and R.

23. The ISVD of any one of claims 21 or 22, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 340-379, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

24. The ISVD as claimed in any one of claims 21 to 23, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 134; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

143. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

144. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

145. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

146. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

147. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

148. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

149. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

150. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

149. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

146. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

145. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

147. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

149. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

150. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 136; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

144. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 135; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

151. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 132; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 137; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

152.

25. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the ISVD consisting essentially of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 159; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 159; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 159 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 160 or 162; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 160 or 162; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 160 or 162 by 3, 2 or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 143 or 145; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 143 or 145; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 143 or 145 by 4, 3, 2 or 1 amino acid.

26. The immunoglobulin single variable domain (ISVD) as described in claim 25, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of HYVMG (SEQ ID NO: 159); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of HYVMG (SEQ ID NO: 159); c) An amino acid sequence that differs from the amino acid sequence of HYVMG (SEQ ID NO: 159) by 3, 2, or 1 amino acid; as well as - CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of 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; e) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610); wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from I, S and V, and the amino acid residue X3 is selected from T and Q; f) An amino acid sequence that differs from the amino acid sequence of X1X2SWSGSGX3YYADSVKG (SEQ ID NO: 610) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from G and A, the amino acid residue X2 is selected from I, S and V, and the amino acid residue X3 is selected from T and Q; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of 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; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608); wherein the amino acid residue X1 is selected from A, V, R, K or E, wherein the amino acid residue X2 is selected from Q or G, wherein the amino acid residue X3 is selected from V, K, Q, H or E, and wherein the amino acid residue X4 is selected from R or L; i) An amino acid sequence that differs from the amino acid sequence of DX1TSX2VX3X4AVEAYDY (SEQ ID NO: 608) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A, V, R, K or E, wherein the amino acid residue X2 is selected from Q or G, wherein the amino acid residue X3 is selected from V, K, Q, H or E, and wherein the amino acid residue X4 is selected from R or L.

27. The ISVD as claimed in claim 25 or 26, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 340-379, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

28. The ISVD as claimed in any one of claims 25 to 27, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 160; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

143. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

144. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

145. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

146. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

147. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

148. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

149. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

150. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

148. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

146. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

144. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

146. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 166; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

145. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

147. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

149. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

147. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 162; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

150. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 161; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

151. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 159; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 163; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

152.

29. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the ISVD consisting essentially of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 169; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 169; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 169 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 171 or 175; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 171 or 175; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 171 or 175 by 3, 2 or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 192 or 193; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 192 or 193 by 3, 2 or 1 amino acid.

30. The immunoglobulin single variable domain (ISVD) as described in claim 29, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) Amino acid sequence GSIFTSAVME (SEQ ID NO: 169); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence GSIFTSAVME (SEQ ID NO: 169); c) An amino acid sequence that differs from the amino acid sequence GSIFTSAVME (SEQ ID NO: 169) by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of RIGSGGRX1A (SEQ ID NO: 611), wherein the 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 of RIGSGGRX1A (SEQ ID NO: 611), wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P and G; f) An amino acid sequence that differs from the amino acid sequence of RIGSGGRX1A (SEQ ID NO: 611) by 3, 2 or 1 amino acid, wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q, A, S, P and G; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of VX1GGYIY (SEQ ID NO: 612), wherein the amino acid residue X1 is selected from M and I; h) An amino acid sequence having at least 80% amino acid identity with the amino acid sequence of VX1GGYIY (SEQ ID NO: 612), wherein the amino acid residue X1 is selected from M and I; i) An amino acid sequence that differs from the amino acid sequence of VX1GGYIY (SEQ ID NO: 612) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from M and I.

31. The ISVD of any one of claims 29 or 30, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 420-450, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

32. The ISVD as claimed in any one of claims 29 to 30, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 171; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

192. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 172; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 173; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 174; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 175; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 176; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 177; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 178; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 171; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 179; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 180; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 169; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 181; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

193.

33. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 197; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 197; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 197 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 198 or 202; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 198 or 202; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 198 or 202 by 3, 2 or 1 amino acids; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 192 or 193; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 192 or 193; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 192 or 193 by 3, 2 or 1 amino acid.

34. The immunoglobulin single variable domain (ISVD) as described in claim 33, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SAVME (SEQ ID NO: 197); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SAVME (SEQ ID NO: 197); c) An amino acid sequence that differs from the amino acid sequence of SAVME (SEQ ID NO: 197) by 3, 2, or 1 amino acid; as well as - CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613); wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q; A, S, P and G, and wherein the 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 of RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613); wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q; A, S, P and G, and wherein the amino acid residue X2 is selected from P and V; f) An amino acid sequence that differs from the amino acid sequence of RIGSGGRX1AYX2DSVKG (SEQ ID NO: 613) by 3, 2 or 1 amino acid; wherein the amino acid residue X1 is selected from I, R, H, E, K, T, Q; A, S, P and G, and wherein the amino acid residue X2 is selected from P and V; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of VX1GGYIY (SEQ ID NO: 612); wherein the amino acid residue X1 is selected from M and I; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of VX1GGYIY (SEQ ID NO: 612); wherein the amino acid residue X1 is selected from M and I; i) An amino acid sequence that differs from the amino acid sequence of VX1GGYIY (SEQ ID NO: 612) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from M and I.

35. The ISVD of any one of claims 33 or 34, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 420-450, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

36. The ISVD as claimed in any one of claims 33 to 35, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 198; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

192. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 199; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 200; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 201; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 202; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 203; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 204; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 205; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 198; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 206; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 207; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 208; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 197; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 209; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

193.

37. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 218; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 218; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 218 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 222; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 222; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 222 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 234; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 234; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 234 by 3, 2 or 1 amino acid.

38. The immunoglobulin single variable domain (ISVD) as described in claim 37, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence ERTFX1SYSMG (SEQ ID NO: 617), wherein the 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), wherein the amino acid residue X1 is selected from S and G; c) An amino acid sequence that differs from the amino acid sequence ERTFX1SYSMG (SEQ ID NO: 617) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from S and G; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of X1IX2YRGAGPY (SEQ ID NO: 614), wherein the amino acid residue X1 is selected from V and P, and wherein the 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 of X1IX2YRGAGPY (SEQ ID NO: 614), wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; f) An amino acid sequence that differs from the amino acid sequence of X1IX2YRGAGPY (SEQ ID NO: 614) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615), wherein the 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 of RDGAVASIPQAFX1S (SEQ ID NO: 615), wherein the amino acid residue X1 is selected from A and T; i) An amino acid sequence that differs from the amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615) by 3, 2 or 1 amino acid residues, wherein the amino acid residue X1 is selected from A and T.

39. The ISVD of any one of claims 37 or 38, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 482-497, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

40. The ISVD as claimed in any one of claims 37 to 40, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 218; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 222; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

234. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 218; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 223; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

234. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 218; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 224; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

234. Or one of them - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 219; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 222; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

235.

41. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 241; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 241; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 241 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 242; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 242; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 242 by 3, 2, or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 234; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 234; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 234 by 3, 2 or 1 amino acid.

42. The immunoglobulin single variable domain (ISVD) as described in claim 41, wherein the immunoglobulin single variable domain is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SYSMG (SEQ ID NO: 241); b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SYSMG (SEQ ID NO: 241); c) An amino acid sequence that differs from the amino acid sequence of SYSMG (SEQ ID NO: 241) by 3, 2, or 1 amino acid; as well as - CDR2 (Kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616); wherein the amino acid residue X1 is selected from V and P, and wherein the 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 of X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616); wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; f) An amino acid sequence that differs from the amino acid sequence of X1IX2YRGAGPYYEDSVKG (SEQ ID NO: 616) by 3, 2 or 1 amino acid; wherein the amino acid residue X1 is selected from V and P, and wherein the amino acid residue X2 is selected from G and F; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615); wherein the amino acid residue X1 is selected from A and T; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615); wherein the amino acid residue X1 is selected from A and T; i) An amino acid sequence that differs from the amino acid sequence of RDGAVASIPQAFX1S (SEQ ID NO: 615) by 3, 2 or 1 amino acid residues; wherein the amino acid residue X1 is selected from A and T.

43. The ISVD of any one of claims 41 or 42, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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: 482-497, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

44. The ISVD as claimed in any one of claims 41 to 43, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 242; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

234. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 243; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

234. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 244; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

234. Or one of them - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 241; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 242; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

235.

45. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 256; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 256; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 256 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 267; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 267; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 267 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 279; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 279; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 279 by 43, 2 or 1 amino acids.

46. ​​The ISVD of claim 45, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

47. The ISVD as claimed in any one of claims 45 or 46, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 256; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 267; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

279.

48. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the ISVD consisting essentially of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 293; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 293; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 293 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 299; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 299; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 299 by 3, 2, or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 279; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 279; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 279 by 3, 2 or 1 amino acid.

49. The ISVD of claim 48, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

50. The ISVD as claimed in any one of claims 48 or 49, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 293; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 299; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

279.

51. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 257; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 257; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 257 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 268; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 268; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 268 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 280; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 280 by 3, 2 or 1 amino acid.

52. The ISVD of claim 51, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

53. The ISVD as claimed in any one of claims 51 or 52, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 257; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 268; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

280.

54. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 294; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 294; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 294 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 300; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 300; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 300 by 3, 2, or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 280; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 280; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 280 by 3, 2 or 1 amino acid.

55. The ISVD of claim 54, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

56. The ISVD as claimed in any one of claims 54 or 55, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 294; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 300; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

280.

57. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 258; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 258; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 258 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 269; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 269; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 269 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 281; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 281; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 281 by 3, 2 or 1 amino acid.

58. The ISVD of claim 57, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

59. The ISVD as claimed in any one of claims 57 to 58, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 258; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 269; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

281.

60. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 295; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 295; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 295 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 301; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 301; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 301 by 3, 2, or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 281; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 281; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 281 by 3, 2 or 1 amino acid.

61. The ISVD of claim 60, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

62. The ISVD as claimed in any one of claims 60 or 61, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 295; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 301; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

281.

63. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 259; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 259; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 259 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 270; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 270; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 270 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 282; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 282; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 282 by 3, 2 or 1 amino acid.

64. The ISVD of claim 63, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

65. The ISVD as claimed in any one of claims 63 or 64, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 259; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 270; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

282.

66. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 296; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 296; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 296 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 302; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 302; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 302 by 3, 2, or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 282; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 282; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 282 by 3, 2 or 1 amino acid.

67. The ISVD of claim 66, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

68. The ISVD as claimed in any one of claims 66 or 67, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 296; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 302; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

282.

69. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 260; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 260; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 260 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 271; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 271; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 271 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 283; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 283; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 283 by 3, 2 or 1 amino acid.

70. The ISVD of claim 69, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

71. The ISVD as claimed in any one of claims 69 or 70, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 260; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 271; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

283.

72. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 297; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 297; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 297 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 303; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 303; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 303 by 3, 2, or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 283; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 283; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 283 by 3, 2 or 1 amino acid.

73. The ISVD of claim 72, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

74. The ISVD as claimed in any one of claims 72 or 73, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 297; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 303; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

283.

75. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (AbM number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 262; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 262; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 262 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 272; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 272; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 272 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 284; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 284; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 284 by 3, 2 or 1 amino acid.

76. The ISVD of claim 75, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

77. The ISVD as claimed in any one of claims 75 or 76, wherein - CDR1 (AbM number) consists of the amino acid sequence of SEQ ID NO: 262; - CDR2 (AbM number) consists of the amino acid sequence of SEQ ID NO: 272; and - CDR3 (AbM number) consists of the amino acid sequence of SEQ ID NO:

284.

78. An immunoglobulin single variable domain (ISVD) that specifically binds to human PD-L1, the immunoglobulin single variable domain being essentially composed of four framework regions (FR1 to FR4) and three complementarity-determining regions (CDR1 to CDR3), wherein: - CDR1 (Kabat number) consists of an amino acid sequence selected from the following: a) The amino acid sequence of SEQ ID NO: 298; b) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 298; c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 298 by 3, 2, or 1 amino acid; as well as - CDR2 (kabat number) consists of an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 304; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 304; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 304 by 6, 5, 4, 3, 2 or 1 amino acid; as well as - CDR3 (Kabat number) consists of an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 284; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 284; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 284 by 4, 3, 2 or 1 amino acid.

79. The ISVD of claim 78, wherein the amino acid sequence of these CDRs (AbM numbers) has at least 80% 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, more preferably at least 90% amino acid sequence identity, such as 95% amino acid sequence identity, or 99% amino acid sequence identity or higher, or even substantially 100% amino acid sequence identity.

80. The ISVD as claimed in any one of claims 78 or 79, wherein - CDR1 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 298; - CDR2 (Kabat number) consists of the amino acid sequence of SEQ ID NO: 304; and - CDR3 (Kabat number) consists of the amino acid sequence of SEQ ID NO:

284.

81. The ISVD according to any one of claims 1 to 80, wherein the amino acid sequence is... i) Having 80% amino acid sequence identity with one of the amino acid sequences of SEQ ID NO: 4, 5, 7, 311-525 or 527-528, wherein the amino acid residues forming these CDR sequences are ignored for the purpose of determining the degree of amino acid identity; And among them: ii) Preferably, according to the Kabat number, one or more amino acid residues at positions 11, 37, 44, 45, 47, 83, 84, 103, 104 and 108 are selected from the marker residues mentioned in Table 1.

82. The ISVD according to any one of claims 1 to 81, which is substantially composed of a heavy chain variable domain sequence derived from a conventional four-chain antibody, or substantially composed of a heavy chain variable domain sequence derived from a heavy chain antibody.

83. The ISVD according to any one of claims 1 to 82, which is substantially composed of VHH, humanized VHH, camelified VH, domain antibody, single domain antibody or dAb.

84. The ISVD according to any one of claims 1 to 83, wherein the humanized ISVD is selected from the group consisting of: SEQ ID NO: 5, 7, 311-321, 326-336, 341-378, 380-418, 421-449, 451-481, 483-496, 498-512 and 527-528, or selected from the group consisting of: an amino acid sequence having greater than 80%, preferably greater than 90%, more preferably greater than 95%, such as 99% or higher amino acid sequence identity with at least one of the amino acid sequences of SEQ ID NO: 5, 7, 311-321, 326-336, 341-378, 380-418, 421-449, 451-481, 483-496, 498-512 and 527-528.

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 NO: 4, 5, 7, 311-525 and 527-528.

86. A polypeptide or construct comprising one or more ISVDs as claimed in any one of claims 1 to 85 or substantially composed of the one or more ISVDs, and optionally further comprising one or more other groups, residues, portions or binding units optionally linked via one or more linkers.

87. The polypeptide or construct of claim 86, wherein one or more other groups, residues, portions or binding units are amino acid sequences.

88. The polypeptide or construct of any one of claims 86 to 87, wherein the one or more linkers are one or more amino acid sequences.

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

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

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

92. The polypeptide or construct according to any one of claims 86 to 91, wherein it is a multivalent construct.

93. The polypeptide or construct according to any one of claims 86 to 92, wherein it is a multispecific construct.

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

95. The polypeptide or construct of claim 94, wherein the one or more other groups, residues, portions or binding units that provide the increased half-life to the polypeptide or construct 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 portions, and small proteins or peptides that specifically bind to serum proteins.

96. The polypeptide or construct of claim 95, wherein the one or more other groups, residues, portions or binding units providing the increased half-life of the polypeptide or construct are selected from the group consisting of human serum albumin or fragments thereof.

97. The polypeptide or construct of claim 95, wherein the one or more other groups, residues, portions or binding units that provide an increased half-life for the polypeptide or construct 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 of claim 97, wherein the one or more other groups, residues, portions or binding units providing the increased half-life of the polypeptide or construct are selected from the group consisting of: VHHs that specifically bind to serum albumin (such as human serum albumin) or serum immunoglobulins (such as IgG), humanized VHHs, camelified VHs, domain antibodies, single-domain antibodies or dAbs.

99. The polypeptide or construct of claim 98, wherein the one or more other groups, residues, portions or binding units that provide the increased half-life to the polypeptide or construct are ISVDs that specifically bind to human serum albumin.

100. The polypeptide or construct of claim 99, wherein the ISVD that specifically binds to human serum albumin is substantially composed of four framework regions (FR1 to FR4, respectively) and three complementarity-determining regions (CDR1 to CDR3, respectively), wherein: - CDR1 (AbM number) has an amino acid sequence selected from the following: a) The 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 SEQ ID NO: 43; and c) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 43 by 3, 2, or 1 amino acid; as well as - CDR2 (AbM number) has an amino acid sequence selected from the following: d) The amino acid sequence of SEQ ID NO: 45; e) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 45; f) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 45 by 3, 2, or 1 amino acid; as well as - CDR3 (AbM number) has an amino acid sequence selected from the following: g) The amino acid sequence of SEQ ID NO: 47; h) An amino acid sequence that has at least 80% amino acid identity with the amino acid sequence of SEQ ID NO: 47; i) An amino acid sequence that differs from the amino acid sequence of SEQ ID NO: 47 by 3, 2 or 1 amino acid.

101. The polypeptide or construct of 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 of 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 of any one of claims 86 to 102, wherein the linker is selected from the group consisting of SEQ ID NO: 53 to 69.

104. The polypeptide or construct of any one of claims 86 to 103, further comprising a C-terminal extension.

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

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

107. The nucleic acid of claim 106, wherein it is in the form of a genetic construct.

108. A non-human host or host cell that expresses, or is capable of expressing, as appropriate, any ISVD as claimed in any one of claims 1 to 85 or a polypeptide as claimed in any one of claims 86 to 105; and / or contains a nucleic acid as claimed in any one of claims 106 or 107.

109. A method for generating an ISVD as claimed in any one of claims 1 to 85 or a polypeptide as claimed in any one of claims 86 to 105, the method comprising at least the following steps: a) Expressing the nucleic acid as described in any one of 106 or 107 in a suitable non-human host cell or host organism or in another suitable expression system; Optional: b) Isolate and / or purify the ISVD as described in any one of claims 1 to 85, or the polypeptide as described in any one of claims 86 to 105.

110. A method for generating an ISVD as claimed in any one of claims 1 to 85 or a polypeptide as claimed in any one of claims 86 to 105, the method comprising at least the following steps: a) The non-human host or host cells are cultured and / or maintained under conditions that cause the non-human host or host cells as described in claim 108 to express and / or produce at least one ISVD as described in any one of claims 1 to 85 or at least one polypeptide as described in any one of claims 86 to 105; Optional: b) Isolate and / or purify the ISVD as described in any one of claims 1 to 85, or the polypeptide as described in any one of claims 86 to 105.

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

112. The composition of claim 111, wherein it is a pharmaceutical composition.

113. The composition of any one of claims 111 or 112, wherein the composition is a pharmaceutical composition further comprises at least one pharmaceutically acceptable carrier, diluent or excipient and / or adjuvant, and optionally comprises one or more additional pharmaceutically active peptides and / or compounds.

114. The ISVD of any one of claims 1 to 85, the polypeptide or construct of any one of claims 86 to 105, or the composition of any one of claims 111 to 113, used as a pharmaceutical.

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

116. The ISVD of any one of claims 1 to 85, the polypeptide or construct of any one of claims 86 to 105, or the composition of any one of claims 111 to 113, for the diagnosis, prevention, and / or treatment of at least one disease and / or disorder associated with PD-L1, the biological or pharmacological activity of said PD-L1, and / or the biological pathways or signaling involving PD-L1 therein.

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

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

119. The method of claim 118, used for the diagnosis, prevention, and / or treatment of at least one disease or disorder associated with PD-L1, the biological or pharmacological activity of said PD-L1, and / or the biological pathways or signaling involving PD-L1, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as claimed in any one of claims 1 to 85, a polypeptide or construct as claimed in any one of claims 86 to 105, or a composition as claimed in any one of claims 111 to 113.

120. The method of any one of claims 118 or 119 for diagnosing, preventing, and / or treating cancer, the method comprising administering to a subject in need a pharmaceutically active amount of at least one ISVD as claimed in any one of claims 1 to 85, a polypeptide or construct as claimed in any one of claims 86 to 105, or a composition as claimed in any one of claims 111 to 113.

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