PD-L1 and TROP-2 targeted conjugates containing effector molecules and their use

Abstract

This application provides a multispecific targeted conjugate comprising a first targeting moiety that specifically recognizes PD-L1, a second targeting moiety that specifically recognizes Trop-2, and an effector molecule, wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site. In some embodiments, the multispecific targeted conjugate further includes a cleavage site between the first targeting moiety and the conjugation site, wherein the second targeting moiety conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site. This application also provides novel anti-PD-L1 antibody constructs and novel anti-Trop-2 antibody constructs. Furthermore, pharmaceutical compositions and treatment methods using the multispecific targeted conjugate are also provided.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims priority to International Patent Application PCT / CN2023 / 096036, filed on 24 May 2023, and International Patent Application PCT / CN2023 / 095850, filed on 23 May 2023, the contents of each of these applications incorporated herein by reference in their entirety.

[0002] Reference to electronic sequence listings The contents of the electronic sequence listing (199872000542SEQLIST.xml; size: 246,381 bytes; and creation date: April 26, 2024) are incorporated herein by reference in their entirety.

[0003] Field of Invention This application relates to a targeted conjugate containing an effector molecule and its use. [Background technology]

[0004] Antibody-drug conjugates (ADCs) are targeted therapies designed to preferentially deliver a drug ("payload") to diseased tissue expressing a surface antigen recognized by the antibody. ADCs often consist of an antibody conjugated to a therapeutic agent (e.g., a cytotoxic agent) via a chemical linker that enables the release of the therapeutic agent to treat the disease. Currently, 10 ADCs are approved by the FDA for therapeutic use, and over 100 are in clinical trials. The majority of currently approved or clinically evaluated ADCs are small-molecule drug-antibody conjugates. Current ADCs incorporate standard chemotherapeutic agents, including, for example, the auristatin monomethyl auristatin E (MMAE), calicheamicin, SN-38, Dxd (exatecan derivative), and meitansine derivative 1 (DM1). See, for example, Polakis, PharmaRev, 2016, 68(1)3-19.

[0005] While ADCs possess conceptual merit and promising clinical outcomes, developing effective ADC therapies remains remarkably challenging. The overall design of the ADC, the selection of target tissues, the antibodies, chemical linkers, drug binding sites, and the properties of the drug payload all influence the efficacy and risks of the ADC (see, e.g., Chau et al., Lancet 2019;394:793-804). For example, early ADCs in clinical trials suffer from the drawback of immunogenicity of the mouse antibodies used, and recent advances in ADC development rely on humanized antibodies. Other unresolved challenges in ADCs include a) the ineffectiveness of the drug payload, b) premature release of the drug leading to loss of efficacy and enhanced toxicity, c) the level of target expression, d) suboptimal target selectivity, e) insufficient penetration into solid tumor tissue, and f) side effects such as thrombocytopenia and neuropathy, as seen with KADCYLA® and ADCETRIS®. ADC platforms, as highly specific payload delivery systems in vivo, are useful for applications beyond small molecule drug delivery. For example, siRNA can be fused to antibodies, potentially leading to efficient methods for in vivo mRNA knockdown (see Baumer et al., Nature Protocols, 2016, 11:22-36). Such antibody-mediated siRNA delivery has shown promising results in the treatment of colorectal cancer (see Baumer et al., Clin Cancer Res 2015 15;21(6):1383-94). A wider range of designs, antibody types, and therapeutic ranges are still needed for ADC platforms. [Prior art documents] [Non-patent literature]

[0006] [Non-Patent Document 1] Polakis,PharmaRev,2016,68(1)3-19 [Non-Patent Document 2] Chau et al.,Lancet 2019;394:793-804 [Non-Patent Document 3] Baumer et al.,Nature Protocols,2016,11:22-36 [Non-Patent Document 4] Baumer et al., Clin Cancer Res 2015 15;21(6):1383-94 [Overview of the project] [Means for solving the problem]

[0007] In one embodiment, the present application provides a multispecific targeting conjugate comprising a first targeting portion that specifically recognizes PD-L1, a second targeting portion that specifically recognizes Trop-2, and an effector molecule, wherein the effector molecule is conjugated to the second targeting portion via a conjugation site.

[0008] In some embodiments of the multispecific targeted conjugate described above, the multispecific targeted conjugate further includes a cleavage site between the first targeting portion and the conjugation site, where the second targeting portion, conjugated with the effector molecule, can be released from the multispecific targeted conjugate by cleavage at the cleavage site.

[0009] In some embodiments of the multispecific targeting conjugate described above, the multispecific targeting conjugate has the structure of formula 1: [ka] (In the formula, A1 is the first targeting region, A2 is the second targeting region, P is the cleavage site, C is the conjugation site, L is the linker, D is the effector molecule, x is 0 or 1, a is 1 to 20, and b is 1 to 20).

[0010] In some embodiments of any of the multispecific targeting conjugates described above, the first and / or second targeting portion comprises one or more targeted peptides or antibodies or their antigen-binding fragments. In some embodiments, the antibody or its antigen-binding fragment is selected from the group consisting of full-length antibodies, diabodies, scFv, scFab, Fab, Fab', F(ab')2, single-domain antibodies (sdAb), dsFv, and combinations thereof.

[0011] In some embodiments of any of the multispecific targeted conjugates described above, cleavage is induced by conditions at the target site. In some embodiments, the conditions at the target site are selected from the group consisting of proteases, pH changes, redox changes, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof. In some embodiments, the target site is a disease site such as a tumor, for example, a solid tumor. In some embodiments, the conditions are the tumor microenvironment.

[0012] In some embodiments of the above multispecific targeted conjugates, cleavage is performed by a protease. In some embodiments, the protease is urokinase plasminogen activator (uPA), regmine, plasmin, TMPRSS3, TMPRSS4, TMPRSS6, MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-10, MMP-12, MMP-13, MMP-14, MT1-MMP, cathepsin D, cathepsin K, cathepsin S, ADAM10, ADAM12, ADA The protease is selected from the group consisting of MTS, caspase-1, caspase-2, caspase-3, caspase-4, caspase-5, caspase-6, caspase-7, caspase-8, caspase-9, caspase-10, caspase-11, caspase-12, caspase-13, caspase-14, TACE, human neutrophil elastase, β-secretase, fibroblast-associated protein, matryptase, PSMA, and PSA. In some embodiments, the protease is MMP. In some embodiments, the cleavage site that can be cleaved by MMP includes the amino acid sequence of SEQ ID NOs. 71 and 137-143. In some embodiments, the MMP is MMP-9. In some embodiments, the cleavage site that can be cleaved by MMP-9 includes the amino acid sequence of SEQ ID NOs. 71 or 142. In some embodiments, the protease is uPA. In some embodiments, the cleavage site that can be cleaved by uPA includes the amino acid sequence of SEQ ID NO: 214.

[0013] In some embodiments of any of the multispecific targeted conjugates described above, the effector molecule is a therapeutic agent or oligonucleotide. In some embodiments, the effector molecule is a therapeutic agent selected from the group consisting of, for example, protein-based drugs, small molecule drugs, cytotoxic agents, toxins, immunomodulators, anti-inflammatory agents, anti-infective agents, and epigenetic modulators. In some embodiments, the therapeutic agent is exatecan.

[0014] In some embodiments of the multispecific targeting conjugates described above, the multispecific targeting conjugate comprises two or more effector molecules.

[0015] In some embodiments of the multispecific targeted conjugate described above, x is 0. In some embodiments, x is 1. In some embodiments, a is 1 to 10, for example, 1. In some embodiments, b is 2 to 10, for example, 4 to 6.

[0016] In some embodiments of the multispecific targeting conjugate described above, the first targeting portion is located at the N-terminus of the second targeting portion.

[0017] In some embodiments of any of the multispecific targeting conjugates described above, the conjugation site includes a transglutaminase conjugation site. In some embodiments, the transglutaminase conjugation site includes the amino acid sequence of any of SEQ ID NOs. 145-196, for example, SEQ ID NOs. 147 or 188.

[0018] In some embodiments of any of the multispecific targeting conjugates described above, the conjugation site (e.g., a transglutaminase conjugation site) comprises two or more glutamine-containing tags fused in series with each other.

[0019] In some embodiments of any of the above multispecific targeting conjugates (for example, the multispecific targeting conjugate of formula 1), L is given by formula:(Gly) n -(PEG) m -VC-PAB-(DMAE) k (wherein n, m, and k are integers, n≧1, m≧2, and k is 0 or 1) is expressed by the formula: (Gly) n -(PEG) m -VA-PAB-(DMAE) k(wherein n, m, and k are integers, n≧1, m≧2, and k is 0 or 1) is expressed as follows: In some embodiments, L is (Gly)6-amide-PEG8-VA-PAB. In some embodiments, L is (Gly)6-amide-PEG8-VC-PAB.

[0020] In some embodiment of the multispecific targeting conjugate described above, L-(D) a Equation A: [ka] The formula includes the structure shown by the wavy line (wherein the formula, the wavy line indicates the site of covalent bonding to the conjugation site C).

[0021] In some embodiments of the multispecific targeted conjugates described above, the second targeted portion includes one or more sdAbs (e.g., VHH) that specifically recognize Trop-2 (anti-Trop-2 sdAb or anti-Trop-2 VHH). In some embodiments, one or more anti-Trop-2 Each sdAb independently contains: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 42 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or a variant thereof containing up to three amino acid mutations; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 35 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 40 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 45 or a variant thereof containing up to three amino acid mutations; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 36 or a variant thereof containing up to three amino acid mutations, and the amino acid sequence of SEQ ID NO: 41 iv) CDR2 or its variant containing up to three amino acid mutations, and CDR3 or its variant containing up to three amino acid mutations containing the amino acid sequence of SEQ ID NO: 46; iv) CDR1 or its variant containing up to three amino acid mutations containing the amino acid sequence of SEQ ID NO: 38, CDR2 or its variant containing up to three amino acid mutations containing the amino acid sequence of SEQ ID NO: 43, and CDR3 or its variant containing up to three amino acid mutations containing the amino acid sequence of SEQ ID NO: 48; v) CDR1 or its variant containing up to three amino acid mutations containing the amino acid sequence of SEQ ID NO: 39, CDR2 or its variant containing up to three amino acid mutations containing the amino acid sequence of SEQ ID NO: 44, and CDR3 or its variant containing up to three amino acid mutations containing the amino acid sequence of SEQ ID NO: 49;vi) CDR1 containing the amino acid sequence of SEQ ID NO: 53 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 58 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 63 or its variant containing up to three amino acid mutations; vii) CDR1 containing the amino acid sequence of SEQ ID NO: 51 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 56 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 61 or its variant containing up to three amino acid mutations; viii) CDR1 containing the amino acid sequence of SEQ ID NO: 52 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 57 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 62 or its variant containing up to three amino acid mutations (i) CDR1 containing the amino acid sequence of SEQ ID NO: 54 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 59 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 64 or a variant thereof containing up to three amino acid mutations; (x) CDR1 containing the amino acid sequence of SEQ ID NO: 55 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 60 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 65 or a variant thereof containing up to three amino acid mutations; (xi) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 205 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or a variant thereof containing up to three amino acid mutations;xii) CDR1 containing the amino acid sequence of SEQ ID NO: 53 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 58 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 206 or a variant thereof containing up to three amino acid mutations; xiii) CDR1 containing the amino acid sequence of SEQ ID NO: 55 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 60 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 243 or a variant thereof containing up to three amino acid mutations;Alternatively, an anti-Trop-2 VHH comprises CDR1 containing the amino acid sequence of SEQ ID NO: 216 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 217 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 218 or a variant thereof containing up to three amino acid mutations. In some embodiments, one or more anti-Trop-2 VHHs independently contain any of the amino acid sequences of SEQ ID NOs: 50, 66, 123-136, and 203. In some embodiments, a second targeting moiety comprises a first anti-Trop-2 sdAb (e.g., VHH) and a second anti-Trop-2 sdAb (e.g., VHH) fused in series with each other. In some embodiments, the first anti-Trop-2 sdAb (e.g., VHH) and the second anti-Trop-2 sdAb (e.g., VHH) bind to different Trop-2 epitopes. In some embodiments, the first or second anti-Trop-2 sdAb is an anti-Trop-2 VHH comprising CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47. In some embodiments, the first or second anti-Trop-2 VHH comprises any of the amino acid sequences of SEQ ID NOs: 50, 124, and 126. In some embodiments, the first or second anti-Trop-2 sdAb is an anti-Trop-2 VHH comprising CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63. In some embodiments, the first or second anti-Trop-2 VHH comprises any of the amino acid sequences of SEQ ID NOs: 66, 123, and 131. In some embodiments, the first or second anti-Trop-2 sdAb is an anti-Trop-2 VHH comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, the first or second anti-Trop-2 VHH comprises the amino acid sequence of SEQ ID NO: 203.

[0022] In some embodiments of the multispecific targeting conjugate described above, the first targeting portion comprises one or more antibodies or their antigen-binding fragments (anti-PD-L1 antibodies or their antigen-binding fragments) that specifically recognize PD-L1. In some embodiments, one or more anti-PD-L1 antibodies or their antigen-binding fragments are independently: i) a heavy chain CDR1 containing the amino acid sequence of SEQ ID NO: 3 ("H-CDR1") or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8 or a variant thereof containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13 or a variant thereof containing up to three amino acid mutations, a light chain CDR1 containing the amino acid sequence of SEQ ID NO: 20 ("L-CDR1") or a variant thereof containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25 or a variant thereof containing up to three amino acid mutations, and L containing the amino acid sequence of SEQ ID NO: 30 -CDR3 or its variants containing up to three amino acid mutations; ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 1 or its variants containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 6 or its variants containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 11 or its variants containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 18 or its variants containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 23 or its variants containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 28 or its variants containing up to three amino acid mutations;iii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 2 or its variant containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 7 or its variant containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 12 or its variant containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 19 or its variant containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 24 or its variant containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 29 or its variant containing up to three amino acid mutations; iv) H-CDR1 containing the amino acid sequence of SEQ ID NO: 4 or its variant containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 9 or its variant containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 14 or its variant containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 21 or its variant containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 26 or its variant containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 31 or its variant containing up to three amino acid mutations; v) H-CDR1 containing the amino acid sequence of SEQ ID NO: 5 or its variant containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 10 or its variant containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 15 or its variant containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 22 or its variant containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 27 or its variant containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 32 or its variant containing up to three amino acid mutations;Or vi) an H-CDR1 containing the amino acid sequence of SEQ ID NO: 244 or a variant thereof containing up to three amino acid mutations, an H-CDR2 containing the amino acid sequence of SEQ ID NO: 245 or a variant thereof containing up to three amino acid mutations, an H-CDR3 containing the amino acid sequence of SEQ ID NO: 246 or a variant thereof containing up to three amino acid mutations, an L-CDR1 containing the amino acid sequence of SEQ ID NO: 247 or a variant thereof containing up to three amino acid mutations, an L-CDR2 containing the amino acid sequence of SEQ ID NO: 248 or a variant thereof containing up to three amino acid mutations, and an L-CDR3 containing the amino acid sequence of SEQ ID NO: 249 or a variant thereof containing up to three amino acid mutations. In some embodiments, one or more anti-PD-L1 antibodies or antigen-binding fragments independently include: i) a variant of the heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO: 16 or having at least about 85% sequence identity to SEQ ID NO: 16, and a variant of the light chain variable region (VL) containing the amino acid sequence of SEQ ID NO: 33 or having at least about 85% sequence identity to SEQ ID NO: 33; ii) a variant of the VH containing the amino acid sequence of SEQ ID NO: 219 or having at least about 85% sequence identity to SEQ ID NO: 219, and a variant of the VL containing the amino acid sequence of SEQ ID NO: 220 or having at least about 85% sequence identity to SEQ ID NO: 220; iii) a variant of the VH containing the amino acid sequence of SEQ ID NO: 212 or having at least about 85% sequence identity to SEQ ID NO: 212. iv) A variant containing at least approximately 85% sequence identity to the original sequence, and a VL containing the amino acid sequence of SEQ ID NO: 213 or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 213; iv) A VH containing the amino acid sequence of SEQ ID NO: 212 or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 212, and a VL containing the amino acid sequence of SEQ ID NO: 33 or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 33; v) A VH containing the amino acid sequence of SEQ ID NO: 212 or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 212, and a VL containing the amino acid sequence of SEQ ID NO: 221 or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 221;vi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 213 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 222 viiii) A VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and a VL containing the amino acid sequence of SEQ ID NO: 221 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 221;ix) A VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and a VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 222 ;x) A VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and a VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 222;xi) A VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and a VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 224; xii) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 225; xiii) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226;xiv) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 224; xvi) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xvii) VH containing the amino acid sequence of SEQ ID NO: 227 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 227 , and a variant containing the amino acid sequence of SEQ ID NO: 226, or a VL containing at least approximately 85% sequence identity to SEQ ID NO: 226; xviii) A variant containing the amino acid sequence of SEQ ID NO: 228, or a VH containing at least approximately 85% sequence identity to SEQ ID NO: 228, and a variant containing the amino acid sequence of SEQ ID NO: 226, or a VL containing at least approximately 85% sequence identity to SEQ ID NO: 226; xix) A variant containing the amino acid sequence of SEQ ID NO: 229, or a VH containing at least approximately 85% sequence identity to SEQ ID NO: 229, and a VL containing the amino acid sequence of SEQ ID NO: 226, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 226; xx) A variant containing the amino acid sequence of SEQ ID NO: 230, or a VH containing at least approximately 85% sequence identity to SEQ ID NO: 230, and a VL containing the amino acid sequence of SEQ ID NO: 226, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 226;xxi) VH containing the amino acid sequence of SEQ ID NO: 230 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 230, and VL containing the amino acid sequence of SEQ ID NO: 220 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 220; xxii) VH containing the amino acid sequence of SEQ ID NO: 219 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 219, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 226; or xxiii) VH containing the amino acid sequence of SEQ ID NO: 250 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 250, and VL containing the amino acid sequence of SEQ ID NO: 251 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 251. In some embodiments, the first targeting moiety comprises a full-length anti-PD-L1 antibody. In some embodiments, the full-length anti-PD-L1 antibody comprises: i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) a heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) a heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and an amino acid sequence of SEQ ID NO: 86; Light chain containing the amino acid sequence; vi) Heavy chain containing the amino acid sequence of SEQ ID NO 87 and light chain containing the amino acid sequence of SEQ ID NO 88; vii) Heavy chain containing the amino acid sequence of SEQ ID NO 91 and light chain containing the amino acid sequence of SEQ ID NO 92; viiii) Heavy chain containing the amino acid sequence of SEQ ID NO 93 and light chain containing the amino acid sequence of SEQ ID NO 94; ix) Heavy chain containing the amino acid sequence of SEQ ID NO 95 and light chain containing the amino acid sequence of SEQ ID NO 96; x) Heavy chain containing the amino acid sequence of SEQ ID NO 97 and light chain containing the amino acid sequence of SEQ ID NO 98; xi) Heavy chain containing the amino acid sequence of SEQ ID NO 99 and light chain containing the amino acid sequence of SEQ ID NO 100; xii) Heavy chain containing the amino acid sequence of SEQ ID NO 101 and light chain containing the amino acid sequence of SEQ ID NO 102; xiii) Heavy chain containing the amino acid sequence of SEQ ID NO 103 and light chain containing the amino acid sequence of SEQ ID NO 104; xiv) Heavy chain containing the amino acid sequence of SEQ ID NO 105 and light chain containing the amino acid sequence of SEQ ID NO 106; xv) Heavy chain containing the amino acid sequence of SEQ ID NO 107 and SEQ ID NO 1 A light chain containing the amino acid sequence of 08; xvi) a heavy chain containing the amino acid sequence of SEQ ID NO: 109 and a light chain containing the amino acid sequence of SEQ ID NO: 110; xvii) a heavy chain containing the amino acid sequence of SEQ ID NO: 111 and a light chain containing the amino acid sequence of SEQ ID NO: 112; xviii) a heavy chain containing the amino acid sequence of SEQ ID NO: 113 and a light chain containing the amino acid sequence of SEQ ID NO: 114; xix) a heavy chain containing the amino acid sequence of SEQ ID NO: 115 and a light chain containing the amino acid sequence of SEQ ID NO: 116; xx) a heavy chain containing the amino acid sequence of SEQ ID NO: 117 and a light chain containing the amino acid sequence of SEQ ID NO: 118; xxi) a heavy chain containing the amino acid sequence of SEQ ID NO: 119 and a light chain containing the amino acid sequence of SEQ ID NO: 120; xxii) a heavy chain containing the amino acid sequence of SEQ ID NO: 121 and a light chain containing the amino acid sequence of SEQ ID NO: 122; xxiii) a heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO: 34; or xxiv) a heavy chain containing the amino acid sequence of SEQ ID NO: 241 and a light chain containing the amino acid sequence of SEQ ID NO: 242. In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment comprises i) a VH containing the amino acid sequence of SEQ ID NO: 16 and a VL containing the amino acid sequence of SEQ ID NO: 33; or ii) a VH containing the amino acid sequence of SEQ ID NO: 250 and a VL containing the amino acid sequence of SEQ ID NO: 251.In some embodiments, the full-length anti-PD-L1 antibody comprises i) a heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO: 34; or ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 241 and a light chain containing the amino acid sequence of SEQ ID NO: 242.

[0023] In some embodiments of the multispecific targeting conjugates described above, the multispecific targeting conjugate has, from the N-terminus to the C-terminus, i) an anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - conjugation site - any linker 2 - anti-Trop-2 sdAb (e.g., VHH); ii) an anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - cleavage site - any linker 2 - conjugation site - any linker 3 - anti-Trop-2 sdAb (e.g., VHH); iii) an anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - conjugation site - any linker 2 - first anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - second anti-Trop-2 sdAb (e.g., VHH); or iv) comprising an anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - cleavage site - any linker 2 - conjugation site - any linker 3 - first anti-Trop-2 sdAb (e.g., VHH) - any linker 4 - second anti-Trop-2 sdAb (e.g., VHH).

[0024] In some embodiments of the multispecific targeted conjugates described above, the multispecific targeted conjugate comprises a full-length anti-PD-L1 antibody, wherein the multispecific targeted conjugate comprises, from N-terminus to C-terminus, a fusion polypeptide: heavy chain of the full-length anti-PD-L1 antibody - any linker 1 - any cleavage site - any linker 2 - conjugation site - any linker 3 - one or more anti-Trop-2 sdAbs (e.g., VHH), i) the heavy chain comprises the amino acid sequence of SEQ ID NO: 17, the light chain comprises the amino acid sequence of SEQ ID NO: 34, and the fusion polypeptide comprises any amino acid sequence of SEQ ID NOs: 67, 69, 215, and 238-240, or ii) the heavy chain comprises the amino acid sequence of SEQ ID NO: 241, the light chain comprises the amino acid sequence of SEQ ID NO: 242, and the fusion polypeptide comprises any amino acid sequence of SEQ ID NOs: 231-237. In some embodiments, the fusion polypeptide comprises any amino acid sequence of SEQ ID NOs: 67, 69, and 215.

[0025] Furthermore, isolated antibody constructs (anti-PD-L1 antibody constructs) containing a targeting moiety that specifically recognizes PD-L1 (anti-PD-L1 targeting moiety) are also provided, where the anti-PD-L1 targeting moiety is i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3 or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8 or a variant thereof containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13 or a variant thereof containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20 or a variant thereof containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25 or a variant thereof containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30 or a variant thereof containing up to three amino acid mutations; ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 1 or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 6 or a variant thereof containing up to three amino acid mutations, and the amino acid sequence of SEQ ID NO: 11 iii) H-CDR3 containing an amino acid sequence or its variant containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO. 18 or its variant containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO. 23 or its variant containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO. 28 or its variant containing up to three amino acid mutations; iii) H-CDR1 containing the amino acid sequence of SEQ ID NO. 2 or its variant containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO. 7 or its variant containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO. 12 or its variant containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO. 19 or its variant containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO. 24 or its variant containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO. 29 or its variant containing up to three amino acid mutations;iv) H-CDR1 containing the amino acid sequence of SEQ ID NO: 4 or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 9 or a variant thereof containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 14 or a variant thereof containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 21 or a variant thereof containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 26 or a variant thereof containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 31 or a variant thereof containing up to three amino acid mutations; or v) Includes H-CDR1 containing the amino acid sequence of SEQ ID NO. 5 or its variant containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO. 10 or its variant containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO. 15 or its variant containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO. 22 or its variant containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO. 27 or its variant containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO. 32 or its variant containing up to three amino acid mutations.

[0026] In some embodiments of the above-described isolated anti-PD-L1 antibody construct, the anti-PD-L1 targeting moiety is: i) a VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and a VL containing the amino acid sequence of SEQ ID NO: 33 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 33; ii) a VH containing the amino acid sequence of SEQ ID NO: 219 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 219, and a VL containing the amino acid sequence of SEQ ID NO: 220 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 220; iii) a VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and a VL containing the amino acid sequence of SEQ ID NO: 213 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 213; iv) a VH containing the amino acid sequence of SEQ ID NO: 212 or at least a) variant containing approximately 85% sequence identity, and a VL containing the amino acid sequence of SEQ ID NO: 33, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 33; v) a VH containing the amino acid sequence of SEQ ID NO: 212, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 212, and a VL containing the amino acid sequence of SEQ ID NO: 221, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 221; vi) a VH containing the amino acid sequence of SEQ ID NO: 16, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 16, and a VL containing the amino acid sequence of SEQ ID NO: 213, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 213; vii) a VH containing the amino acid sequence of SEQ ID NO: 212, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 212, and a VL containing the amino acid sequence of SEQ ID NO: 222, or a variant containing at least approximately 85% sequence identity to SEQ ID NO: 222;viii) A VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and a VL containing the amino acid sequence of SEQ ID NO: 221 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 221; ix) A VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and a VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 222; x) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 224; xi i) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 225; xiii) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xi v) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 224;xvi) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xvii) VH containing the amino acid sequence of SEQ ID NO: 227 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 227, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xviii) VH containing the amino acid sequence of SEQ ID NO: 228 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 228, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xix) VH containing the amino acid sequence of SEQ ID NO: 229 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 229, and a VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226;xx) A VH containing the amino acid sequence of SEQ ID NO: 230 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 230, and a VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226;xxi) A VH containing the amino acid sequence of SEQ ID NO: 230 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 230, and a VL containing the amino acid sequence of SEQ ID NO: 220 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 220; or xxii) A VH containing the amino acid sequence of SEQ ID NO: 219 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 219, and a VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226.

[0027] In some embodiments of the isolated anti-PD-L1 antibody constructs described above, the anti-PD-L1 targeting moiety is selected from the group consisting of full-length antibody, diabody, scFv, scFab, Fab, Fab', F(ab')2, sdAb, dsFv, and combinations thereof.

[0028] In some embodiments of the isolated anti-PD-L1 antibody constructs described above, the anti-PD-L1 targeting moiety is a full-length antibody (a full-length anti-PD-L1 antibody). In some embodiments, the full-length anti-PD-L1 antibody is composed of: i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) a heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) a heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) a heavy chain containing the amino acid sequence of SEQ ID NO: 87 and a light chain containing the amino acid sequence of SEQ ID NO: 88; vii) a heavy chain containing the amino acid sequence of SEQ ID NO: 91 and a light chain containing the amino acid sequence of SEQ ID NO: 92; viii) a heavy chain containing the amino acid sequence of SEQ ID NO: 93 and a light chain containing the amino acid sequence of SEQ ID NO: 94; ix) a heavy chain containing the amino acid sequence of SEQ ID NO: 95 and a light chain containing the amino acid sequence of SEQ ID NO: 96; x) a heavy chain containing the amino acid sequence of SEQ ID NO: 97 and a light chain containing the amino acid sequence of SEQ ID NO: 98; xi) xii) A heavy chain containing the amino acid sequence of SEQ ID NO: 99 and a light chain containing the amino acid sequence of SEQ ID NO: 100; xiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 103 and a light chain containing the amino acid sequence of SEQ ID NO: 104; xiv) A heavy chain containing the amino acid sequence of SEQ ID NO: 105 and a light chain containing the amino acid sequence of SEQ ID NO: 106; xv) A heavy chain containing the amino acid sequence of SEQ ID NO: 107 and a light chain containing the amino acid sequence of SEQ ID NO: 108; xvi ) Heavy chain containing the amino acid sequence of SEQ ID NO: 109 and light chain containing the amino acid sequence of SEQ ID NO: 110; xvii) Heavy chain containing the amino acid sequence of SEQ ID NO: 111 and light chain containing the amino acid sequence of SEQ ID NO: 112; xviii) Heavy chain containing the amino acid sequence of SEQ ID NO: 113 and light chain containing the amino acid sequence of SEQ ID NO: 114; xix) Heavy chain containing the amino acid sequence of SEQ ID NO: 115 and light chain containing the amino acid sequence of SEQ ID NO: 116; xx) Heavy chain containing the amino acid sequence of SEQ ID NO: 117 and light chain containing the amino acid sequence of SEQ ID NO: 118;xxi) A heavy chain containing the amino acid sequence of SEQ ID NO: 119 and a light chain containing the amino acid sequence of SEQ ID NO: 120; xxii) A heavy chain containing the amino acid sequence of SEQ ID NO: 121 and a light chain containing the amino acid sequence of SEQ ID NO: 122; or xxiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO: 34.

[0029] In some embodiments of the isolated anti-PD-L1 antibody constructs described above, the isolated anti-PD-L1 antibody construct is multispecific.

[0030] In some embodiments of the isolated anti-PD-L1 antibody constructs described above, the isolated anti-PD-L1 antibody construct is selected from the group consisting of bispecific T cell engagers (BiTEs), bispecific killer cell engagers (BiKEs), triplicate killer cell engagers (TriKEs), trifunctional hybrid antibodies (triomabs), biaffinity retargeting constructs (DARTs), chimeric antigen receptors (CARs), genetically modified T cell receptors (TCRs), antibody-drug conjugates (ADCs), and combinations thereof.

[0031] In some embodiments of the isolated anti-PD-L1 antibody constructs described above, the isolated anti-PD-L1 antibody construct is a multispecific targeted conjugate comprising a first targeting moiety that specifically recognizes PD-L1, a second targeting moiety that specifically recognizes Trop-2, and an effector molecule, wherein the first targeting moiety is an anti-PD-L1 targeting moiety, and the effector molecule is conjugated to the second targeting moiety via a conjugation site. In some embodiments, the multispecific targeted conjugate further includes a cleavage site between the first targeting moiety and the conjugation site, wherein the second targeting moiety conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site.

[0032] Furthermore, an isolated antibody construct (anti-Trop-2 antibody construct) containing a targeting moiety that specifically recognizes Trop-2 (anti-Trop-2 targeting moiety) is also provided, where the anti-Trop-2 targeting moiety is: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 42 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or a variant thereof containing up to three amino acid mutations; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 35 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 40 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 45 or a variant thereof containing up to three amino acid mutations; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 36 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 41 or a variant thereof containing up to three amino acid mutations, and sequence number iv) CDR3 containing the amino acid sequence of sequence number 46 or its variant containing up to three amino acid mutations; iv) CDR1 containing the amino acid sequence of sequence number 38 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of sequence number 43 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of sequence number 48 or its variant containing up to three amino acid mutations; v) CDR1 containing the amino acid sequence of sequence number 39 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of sequence number 44 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of sequence number 49 or its variant containing up to three amino acid mutations; vi) CDR1 containing the amino acid sequence of sequence number 53 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of sequence number 58 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of sequence number 63 or its variant containing up to three amino acid mutations;vii) CDR1 containing the amino acid sequence of SEQ ID NO: 51 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 56 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 61 or a variant thereof containing up to three amino acid mutations; viii) CDR1 containing the amino acid sequence of SEQ ID NO: 52 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 57 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 62 or a variant thereof containing up to three amino acid mutations; ix) CDR1 containing the amino acid sequence of SEQ ID NO: 54 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 59 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 64 or a variant thereof containing up to three amino acid mutations ;x) CDR1 containing the amino acid sequence of SEQ ID NO: 55 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 60 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 65 or its variant containing up to three amino acid mutations;xi) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 205 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or its variant containing up to three amino acid mutations;xii) CDR1 containing the amino acid sequence of SEQ ID NO: 53 or its variant containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 58 or its variant containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 206 or its variant containing up to three amino acid mutations;xiii) A VHH (anti-Trop-2 VHH) comprising a CDR1 containing the amino acid sequence of SEQ ID NO: 55 or a variant thereof containing up to three amino acid mutations, a CDR2 containing the amino acid sequence of SEQ ID NO: 60 or a variant thereof containing up to three amino acid mutations, and a CDR3 containing the amino acid sequence of SEQ ID NO: 243 or a variant thereof containing up to three amino acid mutations; or xiiiv) a VHH (anti-Trop-2 VHH) comprising a CDR1 containing the amino acid sequence of SEQ ID NO: 216 or a variant thereof containing up to three amino acid mutations, a CDR2 containing the amino acid sequence of SEQ ID NO: 217 or a variant thereof containing up to three amino acid mutations, and a CDR3 containing the amino acid sequence of SEQ ID NO: 218 or a variant thereof containing up to three amino acid mutations. In some embodiments, the anti-Trop-2 VHH comprises any of the amino acid sequences of SEQ ID NOs: 50, 66, 123-136, and 203.

[0033] In some embodiments of the isolated anti-Trop-2 antibody construct described above, the isolated anti-Trop-2 antibody construct comprises two or more anti-Trop-2 VHH molecules fused in series with one another.

[0034] In some embodiments of the isolated anti-Trop-2 antibody constructs described above, the isolated anti-Trop-2 antibody construct is multispecific.

[0035] In some embodiments of the isolated anti-Trop-2 antibody constructs described above, the isolated anti-Trop-2 antibody construct comprises a first anti-Trop-2 VHH and a second anti-Trop-2 VHH. In some embodiments, the first anti-Trop-2 VHH and the second anti-Trop-2 VHH bind to different Trop-2 epitopes. In some embodiments, the first or second anti-Trop-2 VHH comprises CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47. In some embodiments, the first or second anti-Trop-2 VHH comprises the amino acid sequence of SEQ ID NO: 50, 124, and 126. In some embodiments, the first or second anti-Trop-2 VHH comprises CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63. In some embodiments, the first or second anti-Trop-2 VHH comprises one of the amino acid sequences of SEQ ID NOs. 66, 123, and 131. In some embodiments, the first or second anti-Trop-2 VHH comprises CDR1 containing the amino acid sequence of SEQ ID NOs. 216, CDR2 containing the amino acid sequence of SEQ ID NOs. 217, and CDR3 containing the amino acid sequence of SEQ ID NOs. 218. In some embodiments, the first or second anti-Trop-2 VHH comprises the amino acid sequence of SEQ ID NOs. 203.

[0036] In some embodiments of the isolated anti-Trop-2 antibody constructs described above, the isolated anti-Trop-2 antibody construct is selected from the group consisting of BiTE, BiKE, TriKE, DART, CAR, genetically modified TCR, ADC, and combinations thereof.

[0037] In some embodiments of the isolated anti-Trop-2 antibody constructs described above, the isolated anti-Trop-2 antibody construct is a multispecific targeted conjugate comprising a first targeting moiety that specifically recognizes PD-L1, a second targeting moiety that specifically recognizes Trop-2, and an effector molecule, wherein the second targeting moiety is an anti-Trop-2 targeted moiety, and the effector molecule is conjugated to the second targeting moiety via a conjugation site. In some embodiments, the multispecific targeted conjugate further includes a cleavage site between the first targeting moiety and the conjugation site, wherein the second targeting moiety conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site.

[0038] Furthermore, pharmaceutical compositions comprising a pharmaceutically acceptable carrier, either or any of the above-mentioned isolated anti-PD-L1 antibody constructs, are also provided.

[0039] Furthermore, pharmaceutical compositions comprising any and any of the above-mentioned isolated anti-Trop-2 antibody constructs, and a pharmaceutically acceptable carrier, are also provided.

[0040] Furthermore, pharmaceutical compositions are also provided that include a pharmaceutically acceptable carrier, which may or may not include any of the above-described multispecific targeting conjugates. In some embodiments, at least two of the multispecific targeting conjugates in the pharmaceutical composition contain different numbers of effector molecules. In some embodiments, the average molar ratio of effector molecules to multispecific targeting moieties in the pharmaceutical composition is at least about 1:1.

[0041] Also provided is a method for treating an individual's Trop-2 positive cancer (e.g., recurrent Trop-2 positive cancer), comprising administering an effective amount of any of the above multispecific targeted conjugates or any of the above pharmaceutical compositions to the individual. In some embodiments, the Trop-2 positive cancer is selected from the group consisting of breast cancer, cervical cancer, colorectal cancer, esophageal cancer, gastric cancer, lung cancer, oral squamous cell carcinoma, ovarian cancer, pancreatic cancer, kidney cancer, prostate cancer, gastric cancer, thyroid cancer, head and neck cancer, bladder cancer, urothelial carcinoma, uterine cancer, leukemia, extranodal nasal lymphoma, and non-Hodgkin lymphoma (NHL), gallbladder cancer, biliary tract cancer, endometrial cancer, and glioblastoma. In some embodiments, the Trop-2 positive cancer is prostate cancer, lung cancer, cervical cancer, or colorectal cancer.

[0042] Also provided are isolated nucleic acids encoding one or more polypeptide chains of the multispecific targeted conjugate (e.g., multispecific targeted portion), isolated anti-PD-L1 antibody construct, or isolated anti-Trop-2 antibody construct described herein, vectors containing such isolated nucleic acids, and host cells containing such isolated nucleic acids or vectors. Also provided is a method for producing any of the multispecific targeted conjugate, isolated anti-PD-L1 antibody construct, or isolated anti-Trop-2 antibody construct, comprising: (i) culturing a host cell containing any of the isolated nucleic acids or vectors described herein, or any of the host cells described herein, under conditions suitable for the expression of any polypeptide portion of the multispecific targeted conjugate (e.g., multispecific targeted portion), isolated anti-PD-L1 antibody construct, or isolated anti-Trop-2 antibody construct described herein; and (ii) obtaining any polypeptide portion of the multispecific targeted conjugate, isolated anti-PD-L1 antibody construct, or isolated anti-Trop-2 antibody construct described herein from the host cell.

[0043] Also provided herein is a method for producing any of the above-mentioned multispecific targeting conjugates, comprising conjugating an effector molecule to a second targeting portion via a conjugation site. In some embodiments, the method comprises the steps of (a) reacting a multispecific targeting portion, including a first targeting portion, an arbitrary cleavage site, a second targeting portion, and a conjugation site, with a linker reagent to form a multispecific targeting portion-linker intermediate, and (b) reacting the multispecific targeting portion-linker intermediate with a nucleophile of an effector molecule portion to form a multispecific targeting conjugate, or the method comprises the steps of (c) reacting an effector molecule portion with a linker reagent to form a linker-effector molecule intermediate, and ( d) The method includes the step of reacting the linker-effector molecular intermediate with a conjugation site of a multispecific targeting moiety comprising a first targeting moiety, an optional cleavage site, a second targeting moiety, and a conjugation site, thereby forming a multispecific targeting conjugate, or the method includes the step of reacting the multispecific targeting moiety comprising a first targeting moiety, an optional cleavage site, a second targeting moiety, and a conjugation site with a linker-effector molecular conjugate, thereby forming a multispecific targeting conjugate. In some embodiments, the conjugation is mediated by transglutaminase. In some embodiments, the method further includes expressing the multispecific targeting moiety before step (a), step (d), or step (e). In some embodiments, expressing a multispecific targeting moiety includes (i) culturing a host cell containing an isolated nucleic acid or vector encoding one of the multispecific targeting moieties described herein, and (ii) obtaining the expressed multispecific targeting moiety from the host cell. [Brief explanation of the drawing]

[0044] [Figure 1] A schematic structure of an exemplary multispecific targeting conjugate is shown, comprising: a first targeting moiety that specifically recognizes PD-L1, e.g., a full-length anti-PD-L1 antibody; a second targeting moiety that specifically recognizes Trop-2, e.g., one or more anti-Trop-2 sdAbs (e.g., VHH) fused in series with each other; an effector molecule, e.g., a small molecule drug; an arbitrary protease cleavage site; a conjugation site; and an effector molecule linker.

[0045] [Figure 2] The serum titers of four BALB / c mice immunized with PD-L1 against human PD-L1(F19~R238)-Fc fusion protein, as determined by ELISA, are shown.

[0046] [Figure 3] The results of binding of exemplary anti-PD-L1 antibodies to Flp-In-CHO cells expressing full-length human PD-L1 (fluorescence-activated cell sorting; FACS) are shown. PD-L1-BMK1 (atezolizumab biosimilar) was a positive control. Human IgG1 isotype antibody was a negative control. [Figure 4] The results of binding of exemplary anti-PD-L1 antibodies to Flp-In-CHO cells expressing full-length human PD-L1 (fluorescence-activated cell sorting; FACS) are shown. PD-L1-BMK1 (atezolizumab biosimilar) was a positive control. Human IgG1 isotype antibody was a negative control.

[0047] [Figure 5] The results of binding (FACS) of exemplary anti-PD-L1 antibodies to Flp-In-CHO cells expressing full-length cynomolgus monkey PD-L1 are shown. PD-L1-BMK1 was a positive control. Human IgG1 isotype antibody was a negative control. [Figure 6] The results of binding (FACS) of exemplary anti-PD-L1 antibodies to Flp-In-CHO cells expressing full-length cynomolgus monkey PD-L1 are shown. PD-L1-BMK1 was a positive control. Human IgG1 isotype antibody was a negative control.

[0048] [Figure 7] This shows FACS results of a competitive assay of exemplary anti-PD-L1 antibodies that block the binding of hPD-1(ECD)-Avitag-His-tagged biotin to Flp-In-CHO cells expressing full-length human PD-L1. PD-L1-BMK1 was a positive control. Human IgG1 isotype antibody was a negative control. [Figure 8] This shows FACS results of a competitive assay of exemplary anti-PD-L1 antibodies that block the binding of hPD-1(ECD)-Avitag-His-tagged biotin to Flp-In-CHO cells expressing full-length human PD-L1. PD-L1-BMK1 was a positive control. Human IgG1 isotype antibody was a negative control.

[0049] [Figure 9] This shows FACS results of a competitive assay of exemplary anti-PD-L1 antibodies that block the binding of hCD80(ECD)-Fc-Avitag-biotin to Flp-In-CHO cells expressing full-length human PD-L1. PD-L1-BMK1 was a positive control. Human IgG1 isotype antibody was a negative control. [Figure 10] This shows FACS results of a competitive assay of exemplary anti-PD-L1 antibodies that block the binding of hCD80(ECD)-Fc-Avitag-biotin to Flp-In-CHO cells expressing full-length human PD-L1. PD-L1-BMK1 was a positive control. Human IgG1 isotype antibody was a negative control.

[0050] [Figure 11] The dose-response curves of an immunological blockade reporter assay with a humanized anti-PD-L1 antibody in Jurkat-NFAT-Luc2-PD-1 effector cells and 293T-OS8-PD-L1 cells are shown. Atezolizumab served as a positive control. Human IgG1 isotype served as a negative control.

[0051] [Figure 12]This image shows exemplary binding of anti-Trop-2 VHH-hIgG1 Fc protein to human Trop-2 by ELISA. Sacituzumab (ALGA106A) and datopotamab (ALGA106B) served as positive controls. Human IgG1 isotypes served as negative controls.

[0052] [Figure 13] This shows the binding of exemplary anti-Trop-2 VHH-hIgG1 Fc protein to MDA-MB-468 (a human triple-negative breast cancer cell line expressing Trop-2) by FACS. Sacituzumab (ALGA106A) and datopotamab (ALGA106B) served as positive controls. Human IgG1 isotypes served as negative controls.

[0053] [Figure 14] The curves show the time course of internal translocation of exemplary anti-Trop-2 VHH-hIgG1 Fc protein into MDA-MB-468 cells. Sacituzumab served as a positive control. [Figure 15] The curves show the time course of internal translocation of exemplary anti-Trop-2 VHH-hIgG1 Fc protein into MDA-MB-468 cells. Sacituzumab served as a positive control.

[0054] [Figure 16] FACS analysis shows the binding affinity of llama parental Trop-2-B-YC-82 and humanized anti-Trop-2 VHH-hIgG1 Fc proteins to MDA-MB-468. Sacituzumab (ALGA106A) served as a positive control. Human IgG1 isotypes served as negative controls.

[0055] [Figure 17] The curves show the time course of internal translocation of llama parental Trop-2-B-YC-82 and humanized anti-Trop-2 VHH-hIgG1 Fc protein into MDA-MB-468 cells. Sacituzumab (ALGA106A) served as a positive control.

[0056] [Figure 18] FACS analysis shows the binding affinity of llama parental Trop-2-A-258 and humanized anti-Trop-2 VHH-hIgG1 Fc proteins to MDA-MB-468. Sacituzumab (ALGA106A) and datopotamab (ALGA106B) served as positive controls. Human IgG1 isotypes served as negative controls.

[0057] [Figure 19] The curves show the time course of internal translocation of llama parental Trop-2-A-258 and humanized anti-Trop-2 VHH-hIgG1 Fc protein into MDA-MB-468 cells. Sacituzumab (ALGA106A) and datopotamab (ALGA106B) served as positive controls.

[0058] [Figure 20] This shows the time-dependent stability of the multispecific Ab ALGA105S and ALGA105X in human plasma as measured by ELISA.

[0059] [Figure 21] This shows the internal migration efficiency of pHAb dye-conjugated multispecific antibodies in the A431 cell line. Sacituzumab (ALGA106A) and datopotamab (ALGA106B) served as anti-Trop-2 positive controls. [Figure 22] This shows the internal migration efficiency of pHAb dye-conjugated multispecific antibodies in the HCC827 cell line. Sacituzumab (ALGA106A) and datopotamab (ALGA106B) served as anti-Trop-2 positive controls. [Figure 23] This shows the internal migration efficiency of pHAb dye-conjugated multispecific antibodies in the HCT15-hPD-L1-hTrop-2 cell line. Sacituzumab (ALGA106A) and datopotamab (ALGA106B) served as anti-Trop-2 positive controls.

[0060] [Figure 24]The cytotoxic dose-response curves in the PC3-hPD-L1-hTrop-2 cell line are shown. Non-conjugate sacituzumab (ALGA106A), datopotamab (ALGA106B), ALGA105S, and ALGA105X served as controls. [Figure 25] The cytotoxic dose-response curves in the HCT15-hPD-L1-hTrop-2 cell line are shown. Non-conjugate sacituzumab (ALGA106A), datopotamab (ALGA106B), ALGA105S, and ALGA105X served as controls.

[0061] [Figure 26] The mean body weight curves over time in MC38-hPD-L1-hTrop-2 syngeneic tumor mouse models after a single intravenous dose of the test ADC, non-conjugate multispecific Ab, or durvalumab (anti-PD-L1 mAb) in combination therapy.

[0062] [Figure 27] The images show the time-course tumor growth curves of the MC38-hPD-L1-hTrop-2 syngeneic tumor mouse model after a single intravenous dose of the test ADC, non-conjugate multispecific Ab, or durvalumab (anti-PD-L1 mAb) in combination therapy. [Figure 28] The images show the time-course tumor growth curves of the MC38-hPD-L1-hTrop-2 syngeneic tumor mouse model after a single intravenous dose of the test ADC, non-conjugate multispecific Ab, or durvalumab (anti-PD-L1 mAb) in combination therapy. [Figure 29] The images show the time-course tumor growth curves of the MC38-hPD-L1-hTrop-2 syngeneic tumor mouse model after a single intravenous dose of the test ADC, non-conjugate multispecific Ab, or durvalumab (anti-PD-L1 mAb) in combination therapy.

[0063] [Figure 30] This shows the stability of ADCs in human plasma as measured by ELISA. ALGA106B (datopotamab)-ADC served as a control.

[0064] [Figure 31] Exemplary cytotoxic dose-response curves of ADCs in the Trop-2-positive A431 cell line are shown. ALGA106A (sacituzumab)-ADC and ALGA106B (datopotamab)-ADC served as positive controls. [Figure 32] The cytotoxic dose-response curves of exemplary ADCs in the Trop-2-positive HCC827 cell line are shown. ALGA106A (sacituzumab)-ADC and ALGA106B (datopotamab)-ADC served as positive controls.

[0065] [Figure 33] This shows the time-course tumor growth curves of an MC38-hPD-L1-hTrop-2 syngeneic tumor mouse model after a single intravenous dose of ALGA105X-ADC and after re-transplantation by a second subcutaneous injection of MC38-hPD-L1-hTrop-2 cells without subsequent ALGA105X-ADC administration.

[0066] [Figure 34] Figure 33 shows the average body weight of mice administered ALGA105X-ADC in the re-implantation experiment. [Modes for carrying out the invention]

[0067] This application provides compositions and treatment methods using a targeted conjugate comprising a targeted moiety and an effector molecule, wherein the targeted moiety specifically recognizes PD-L1 and / or Trop-2, and the effector molecule is conjugated to the targeted moiety via a conjugation site, and the targeted moiety linked to the effector molecule can be released from the targeted conjugate by cleavage. The targeted moiety that recognizes PD-L1 and / or Trop-2 binds to cell surface PD-L1 and / or Trop-2 expressed on diseased tissue or diseased cells (e.g., tumor cells). When the targeted conjugate enters a target site (e.g., tumor microenvironment), cleavage of the cleavage site in the targeted conjugate may be induced, which leads to the release of the targeted moiety linked to the effector molecule at the target site. In some embodiments, the targeted conjugate is multispecific. In some embodiments, the targeted conjugate comprises a first targeted portion and a second targeted portion, which are fused to each other via an arbitrary cleavage site and a conjugation site to which one or more effector molecules are conjugated. For example, the targeted portion may be designed by fusioning one or more scFv, scFab, or single-domain antibodies (e.g., anti-Trop-2 VHH) with a clinically efficacy and safety proven monoclonal antibody (e.g., anti-PD-L1 mAb) to provide multispecificity of target recognition and to result in enhanced clinical efficacy. The targeted conjugates described herein may have one or more of the following properties: i) enhanced selective target recognition ability and efficacy compared to conventional ADCs; ii) enhanced safety by targeting the drug to the tumor site and optionally releasing the drug conjugated to the anti-Trop-2 targeted portion (similarly localized to the tumor site); and iii) the anti-Trop-2 sdAb(plural)(e.g., VHH(plural)) portion conjugated to the drug, upon release, exhibits superior tissue penetration compared to conventional IgG-ADCs.

[0068] The present invention addresses one or more unmet needs, including efficient and / or highly specific drug payload release, high permeability, payload delivery to tumor tissue, enhanced antibody target selectivity, and reduced toxicity (e.g., toxicity to non-target cells) / enhanced safety profile. Furthermore, the multispecific targeted conjugate of the present invention employs an immunomodulator (anti-PD-L1 Ab) together with an ADC in a single construct, thereby achieving the following two antitumor mechanisms: i) the ADC portion exhibits target tumor cell killing, and ii) the immunomodulator portion improves the inhibition of immune checkpoints against effector T cells by tumor cells, thereby reducing effector T cell depletion and evasion of the tumor cell immune response. The PD-L1 / Trop-2 multispecific targeting conjugates described herein may: i) effectively treat Trop-2-positive cancers in vivo, exhibiting significantly superior therapeutic efficacy compared to either the sacituzumab-ADC + durvalumab control or the datopotamab-ADC + durvalumab control combination; ii) demonstrate enhanced therapeutic efficacy when a cleavage site is used between the anti-PD-L1 targeting moiety and the anti-Trop-2 targeting moiety conjugated to the effector molecule; iii) demonstrate a superior safety profile comparable to either the sacituzumab-ADC + durvalumab control or the datopotamab-ADC + durvalumab control combination; iv) possess a superior PK profile, reflected in stability in human plasma, for example, compared to the datopotamab-ADC control; and v) demonstrate a remarkably long-lasting immunological memory and safety profile against tumor cells in vivo, potentially offering promising therapeutic efficacy against recurrent tumors or preventing cancer recurrence.

[0069] Accordingly, one aspect of the present application provides a multispecific targeted conjugate comprising a first targeting portion that specifically recognizes PD-L1, a second targeting portion that specifically recognizes Trop-2, and an effector molecule, wherein the effector molecule is conjugated to the second targeting portion via a conjugation site. In some embodiments, the multispecific targeted conjugate further includes a cleavage site between the first targeting portion and the conjugation site, wherein the second targeting portion conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site.

[0070] Another aspect of this application provides a novel anti-PD-L1 antibody construct comprising a targeting moiety that specifically recognizes PD-L1. Another aspect of this application provides a novel anti-Trop-2 antibody construct comprising a targeting moiety that specifically recognizes Trop-2.

[0071] Also provided are pharmaceutical compositions, kits, and manufactured articles comprising any of the multispecific targeted conjugates, anti-PD-L1 antibody constructs, or anti-Trop-2 antibody constructs described herein. Furthermore, methods for treating Trop-2-positive cancer in an individual using any of the multispecific targeted conjugates or anti-Trop-2 antibody constructs described herein, or any of these pharmaceutical compositions, are also provided. Methods for constructing any of the multispecific targeted conjugates described herein are also provided.

[0072] I. Definition Unless otherwise specifically defined below, terms are used herein in the manner commonly used in the art.

[0073] The term "targeting portion," as used herein, refers to a polypeptide-based binding molecule that specifically binds to a target molecule, or a portion thereof that contributes to specific binding. Both antibody-based and non-antibody-based binding molecules, or portions thereof, are intended herein.

[0074] As used herein, the term "therapeutic agent" refers to a molecule that has a therapeutic effect. A therapeutic agent may be any suitable molecular entity other than oligonucleotides.

[0075] The term "effector molecule," as used herein, refers to a molecule that may be used for therapeutic and / or diagnostic purposes. Examples of effector molecules include, but are not limited to, therapeutic agents, diagnostic labels, and oligonucleotides.

[0076] The term "conjugation" refers to the chemical linking of two chemical groups or parts to each other by one or more covalent or noncovalent bonds. Conjugation can be direct between the two chemical groups or parts, or indirectly through a third chemical group or part (e.g., a linker) that bridges the two chemical groups or parts.

[0077] The term "conjugation site" refers to a site that directly links two chemical parts.

[0078] The term "antibody," used in its broadest sense, encompasses a variety of antibody structures. An "antibody" can refer to an immunoglobulin molecule or fragment thereof (including the basic four-stranded antibody unit) that can specifically bind to a specific epitope of an antigen. Antibodies can be intact immunoglobulins derived from natural or recombinant sources, or they can be the immunoreactive portion of an intact immunoglobulin. The antibodies of the present invention may exist in various forms, including, for example, polyclonal antibodies, monoclonal antibodies, intracellular antibodies ("intrabodies"), antigen-binding fragments (e.g., Fv, Fab, Fab', F(ab)2, and F(ab')2), as well as single-chain antibodies (scFv), heavy-chain antibodies, such as camelid antibodies, and humanized antibodies (Harlow et al., 1999, Using Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, NY; Harlow et al., 1989, Antibodies: A Laboratory Manual, Cold Spring Harbor, New York; Houston et al., 1988, Proc. Natl. Acad. Sci. USA 85:5879-5883; Bird et al., 1988, Science 242:423-426).

[0079] A full-length antibody comprises two heavy chains and two light chains. The variable regions of the light and heavy chains are responsible for antigen binding. The variable domains of the heavy and light chains may be referred to as "VH" and "VL," respectively. The variable regions in both chains typically contain three highly variable loops referred to as complementarity-determining regions (CDRs) (light chain (LC)CDRs containing LC-CDR1, LC-CDR2, and LC-CDR3; heavy chain (HC)CDRs containing HC-CDR1, HC-CDR2, and HC-CDR3). The CDR boundaries of the antibodies and antigen-binding fragments disclosed herein may be defined or identified by the rules of Kabat, Chothia, or Al-Lazikani (Al-Lazikani 1997, Chothia 1985, Chothia 1987, Chothia 1989, Kabat 1987, Kabat 1991). The three CDRs of the heavy or light chain are more highly conserved than the CDRs and interpose between adjacent stretches as framework regions (FRs) known to form a scaffold for supporting the hypervariable loop. The constant regions of the heavy and light chains do not participate in antigen binding but exhibit various effector functions. Antibodies are assigned to classes based on the amino acid sequence of the constant region of their heavy chains. The five major classes or isotypes of antibodies are IgA, IgD, IgE, IgG, and IgM, which are characterized by the presence of α, δ, ε, γ, and μ heavy chains, respectively. Some of the major antibody classes are classified into subclasses, e.g., IgG1 (γ1 heavy chain), IgG2 (γ2 heavy chain), IgG3 (γ3 heavy chain), IgG4 (γ4 heavy chain), IgA1 (α1 heavy chain), or IgA2 (α2 heavy chain).

[0080] The term “antigen-binding fragment” as used herein refers to an antibody fragment that includes, for example, a diabody, Fab, Fab', F(ab')2, Fv fragment, disulfide-stabilized Fv fragment (dsFv), (dsFv)2, bispecific dsFv (dsFv-dsFv'), disulfide-stabilized diabody (dsdiabody), single-stranded Fv (scFv), scFv dimer (bivalent diabody), a multispecific antibody formed from a portion of an antibody containing one or more CDRs, a camelized single-domain antibody, a nanobody, a domain antibody, a bivalent domain antibody, or any other antibody fragment that binds to an antigen but does not contain a complete antibody structure. An antigen-binding fragment can bind to the same antigen to which a parent antibody or parent antibody fragment (e.g., parent scFv) binds. In some embodiments, an antigen-binding fragment may contain one or more CDRs derived from a particular human antibody that have been transplanted into a framework region derived from one or more different human antibodies.

[0081] "Fv" is the smallest antibody fragment containing a complete antigen recognition and binding site. This fragment consists of a dimer in which one heavy chain variable domain and one light chain variable domain are tightly and noncovalently associated. The folding of these two domains creates six hypervariable loops (three from the heavy chain and three from the light chain) that provide amino acid residues for antigen binding and confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of Fv containing only three antigen-specific CDRs) has the ability to recognize and bind to the antigen, albeit with lower affinity than the entire binding site.

[0082] "Single-chain Fv", also abbreviated as "sFv" or "scFv", is an antibody fragment that contains VH and VL antibody domains linked to a single polypeptide chain. In some embodiments, the scFv polypeptide further includes a polypeptide linker between the VH and VL domains that enables the scFv to form a structure desirable for antigen binding. For a review of scFv, see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenberg and Moore eds., Springer-Verlag, New York, pp. 269-315 (1994).

[0083] "Nanobody" (Nanobody® is a registered trademark of Ablynx), also known as single-domain antibody (sdAb) or single variable domain antibody, is an antibody fragment consisting of a single monomeric antibody variable domain. Like full-length antibodies, nanobodies can selectively bind to specific antigens. Nanobodies have a molecular weight of only 12-15 kDa and are much smaller than typical full-length antibodies (150-160 kDa). A nanobody is a peptide chain approximately 110 amino acids in length that contains one variable domain (VH) of a heavy-chain antibody or a typical IgG. Unlike full-length antibodies, nanobodies such as VHH or V NAR do not exhibit cytotoxicity induced by the complement system because they lack the Fc region. Nanobodies derived from camelids (e.g., VHH) and fish (e.g., V NAR ) can bind to hidden antigens that are inaccessible to full-length antibodies, such as the active site of an enzyme. Nanobodies can be obtained by immunization with a desired antigen in camelids or sharks and subsequent isolation of the mRNA encoding the heavy-chain antibody. Camelids are members of the biological family Camelidae, the only existing family of the suborder Tylopoda. Camels, dromedaries, llamas, alpacas, vicuñas, and guanacos belong to this group. Alternatively, nanobodies can be generated by screening synthetic libraries.

[0084] The "VHH domain," also known as VHH, VHH antibody fragments, and VHH antibodies are a type of nanobody. VHH was first described as the immunoglobulin antigen-binding (variable) domain of "heavy-chain antibodies" (i.e., "antibodies lacking a light chain") (Hamers-Casterman et al (1993) Nature 363:446-448). The term "VHH domain" has been chosen to distinguish these variable domains from the heavy-chain variable domains (referred to herein as "VH domains") and light-chain variable domains (referred to herein as "VL domains") present in conventional quadruple-chain antibodies. For further explanation of VHH and nanobodies, refer to Muyldermans' review article (J. Biotechnol. 74:277-302, 2001) and the following patent applications listed as general background art: WO94 / 04678, WO95 / 04079, WO96 / 34103; WO94 / 25591, WO99 / 37681, WO00 / 40968, WO00 / 43507, WO00 / 65057, WO01 / 40310, WO01 / 44301, EP1134231, and WO02 / 4 8193;WO97 / 49805, WO01 / 21817, WO03 / 035694, WO03 / 054016, and WO03 / 055527;WO03 / 050531;WO01 / 90190;WO03 / 025020;WO04 / 041867, WO04 / 041862, WO04 / 041865, WO04 / 041863, WO04 / 062551, WO05 / 044858, WO06 / 40153, WO06 / 079372, WO06 / 122786, WO06 / 122787, and WO06 / 122825. As described in these references, nanobodies (particularly VHH sequences and partially humanized nanobodies) may be characterized in particular by the presence of one or more "characteristic residues" in one or more framework sequences.Further descriptions of nanobodies, including humanization and / or camelization of nanobodies, as well as other modifications, parts or fragments, derivatives or “nanobody fusions,” multivalent constructs (including some non-limiting examples of linker sequences), and different modifications to increase the half-life of nanobodies, and their preparations, can be found, for example, in WO08 / 101985 and WO08 / 142164.

[0085] An “isolated” antibody is one identified, separated, and / or recovered from components of its production environment (e.g., natural or recombinant). Preferably, the isolated polypeptide is not associated with all other components from its production environment. Contaminations of the production environment, such as those from recombinant transcellular cells, are typically substances that interfere with the study, diagnostic, or therapeutic use of the antibody and may include enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. In preferred embodiments, the polypeptide is purified to (1) more than 95% by weight of the antibody, and in some embodiments to more than 99% by weight, as determined, for example by the Lowry method; (2) to a degree sufficient to obtain at least 15 residues of the N-terminal or internal amino acid sequence using a spinning cup sequencer; or (3) to homogeneity by SDS-PAGE under non-reducing or reducing conditions using Coomassie blue, or preferably silver staining. The isolated antibody contains the antibody in its in vivo in-situ within recombinant cells, because at least one component of the antibody’s natural environment is absent. However, typically, isolated polypeptides or antibodies are prepared by at least one purification step.

[0086] The "variable region" or "variable domain" of an antibody refers to the amino-terminal domain of the antibody's heavy or light chain. The variable domains of the heavy and light chains may be referred to as "VH" and "VL," respectively. These domains are typically the most variable parts of the antibody (compared to other antibodies of the same class) and contain the antigen-binding site. Antibodies derived solely from camelid species have a single heavy-chain variable region, which is referred to as "VHH." Therefore, VHH is a special type of VH.

[0087] The term "variable" refers to the fact that the sequences of specific segments of the variable domain vary widely among antibodies. The V domain mediates antigen binding and determines the specificity of a particular antibody to that particular antigen. However, variability is not uniformly distributed across the entire range of the variable domain. Rather, it is concentrated in three segments called hypervariable regions (HVRs) in both the light and heavy chain variable domains. The more highly conserved portion of the variable domain is called the framework region (FR). The natural heavy and light chain variable domains each contain four FR regions, primarily in a β-sheet structure, linked by three HVRs, which link the β-sheet structure and, in some cases, form loops that constitute part of it. The HVRs of each chain are held in close proximity to each other by the FR regions and, together with the HVRs from the other chain, contribute to the formation of the antibody's antigen-binding site (see Kabat et al., Sequences of Immunological Interest, Fifth Edition, National Institute of Health, Bethesda, MD (1991)). The constant domain does not directly participate in antibody binding to antigens, but it exhibits various effector functions, such as antibody-dependent cell-mediated cytotoxicity.

[0088] As used herein, the terms “CDR” or “complementarity-determining region” are intended to mean discontinuous antigen-binding sites found within the variable regions of both heavy and light chain polypeptides. These specific regions have been identified by Kabat et al., J.Biol.Chem.252:6609-6616(1977), Kabat et al., USDept.of Health and Human Services, “Sequences of proteins of immunological interest”(1991), Chothia et al., J.Mol.Biol.196:901-917(1987), Al-Lazikani B.et al. al., J.Mol.Biol.,273:927-948(1997), MacCallum et al.,J.Mol.Biol.262:732-745 (1996), Abhinandan and Martin,Mol.Immunol.,45:3832-3839(2008), Lefranc MPet As described by al., Dev. Comp. Immunol., 27:55-77 (2003), and Honegger and Pluckthun, J. Mol. Biol., 309:657-670 (2001), the definitions therein include duplication or subsets of amino acid residues when compared to one another. Nevertheless, the application of any definition to refer to an antibody or its transplanted antibody or variant CDR is intended to be within the scope of the definitions and terms used herein. For example, CDR prediction algorithms and interfaces are known in the art, including Abhinandan and Martin, Mol.Immunol., 45:3832-3839 (2008), Ehrenmann F. et al., Nucleic Acids Res., 38:D301-D307 (2010), and Adolf-Bryfogle J. et al., Nucleic Acids Res., 43:D432-D438 (2015). The contents of the references cited in this paragraph are incorporated herein by reference in their entirety for use in this application and for the possibility of inclusion in one or more claims herein.In some embodiments, the CDR sequences provided herein are based on IMGT definitions. For example, the CDR sequence may be determined by the VBASE2 tool (see also Retter I, Althaus HH, Munch R, Muller W: VBASE2, an integrative V gene database. Nucleic Acids Res. 2005 Jan 1;33 (Database issue): D671-4 (which is incorporated herein by reference in its entirety)).

[0089] The terms “Fc region,” “Fc domain,” or “Fc” refer to the non-antigen-binding region at the C-terminus of an immunoglobulin heavy chain, which includes at least a portion of the constant region. This term encompasses both native and variant Fc regions. In some embodiments, the Fc domain is selected from the group consisting of Fc fragments of IgG, IgA, IgD, IgE, IgM, and combinations and hybrids thereof. In some embodiments, the Fc domain is derived from human IgG. In some embodiments, the Fc domain includes the Fc domain of human IgG1, IgG2, IgG3, IgG4, or combinations or hybrid IgG. In some embodiments, the Fc domain has reduced effector function compared to the corresponding wild-type Fc domain (e.g., reduced effector function by at least about 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, or 95%, as measured by the level of antibody-dependent cell-mediated cytotoxicity (ADCC)). In some embodiments, the human IgG heavy chain Fc region extends from Cys226 to the carboxyl terminus of the heavy chain. However, the C-terminal lysine (Lys447) of the Fc region may or may not be present without affecting the structure or stability of the Fc region. In some embodiments, the targeting region includes a variant Fc region having at least one amino acid substitution compared to the Fc region of wild-type IgG or wild-type antibody.

[0090] As used herein, the term “monoclonal antibody” refers to an antibody obtained from a substantially homogeneous population of antibodies; that is, the individual antibodies constituting the population are identical except for possible naturally occurring mutations and / or post-translational modifications (e.g., isomerization, amidation) that may be present in small amounts. Monoclonal antibodies are highly specific and directed to a single antigenic site. In contrast to polyclonal antibody preparations, which typically contain different antibodies directed to different determinants (epitopes), each monoclonal antibody is directed to a single determinant on the antigen. In addition to their specificity, monoclonal antibodies have the advantage that they are synthesized by hybridoma culture and are not contaminated with other immunoglobulins. The modifier “monoclonal” describes the characteristics of an antibody when obtained from a substantially homogeneous population of antibodies and should not be interpreted as requiring the production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with this application are, for example, those produced by the hybridoma method (e.g., Kohler and Milstein, Nature, 256:495-97 (1975), Hongo et al., Hybridoma, 14(3):253-260 (1995), Harlow et al., Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2 nd(ed. 1988), Hammerling et al., in: Monoclonal Antibodies and T-Cell Hybridomas 563-681 (Elsevier, NY, 1981)), Recombinant DNA method (see U.S. Patent Nos. 4,816,567), Phage display technology (e.g., Clackson et al., Nature, 352:624-628 (1991), Marks et al., J. Mol. Biol. 222:581-597 (1992), Sidhu et al., J. Mol. Biol. 338(2):299-310 (2004), Lee et al., J. Mol. Biol. 340(5):1073-1093 (2004), Fellouse, Proc. Natl. Acad. Sci. USA See 101(34):12467-12472(2004) and Lee et al., J.Immunol.Methods 284(1-2):119-132(2004), and techniques for producing human or human-like antibodies in animals having a human immunoglobulin locus or a part or all of a gene encoding a human immunoglobulin sequence (e.g., WO1998 / 24893, WO1996 / 34096; WO1996 / 33735, WO1991 / 10741, Jakobovits et al., Proc.Natl.Acad.Sci.USA 90:2551(1993), Jakobovits et al., Nature 362:255-258(1993), Bruggemann et al., Year in Immunol. 7:33 (1993), U.S. Patent Nos. 5,545,807, 5,545,806, 5,569,825, 5,625,126, 5,633,425, and 5,661,016, Marks et al., Bio / Technology 10:779-783 (1992), Lonberg et al., Nature 368:856-859 (1994), Morrison, Nature 368:812-813 (1994), Fishwild et al., Nature Biotechnol. 14:845-851 (1996), Neuberger, Nature Biotechnol.It can be produced by various techniques, including (see 14:826 (1996) and Lonberg and Huszar, Intern. Rev. Immunol. 13:65-93 (1995)).

[0091] In this specification, monoclonal antibodies specifically include "chimeric" antibodies (immunoglobulins) and fragments of such antibodies (to the extent that they exhibit the desired biological activity), in which a portion of the heavy chain and / or light chain is identical or homologous to a corresponding sequence of an antibody derived from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain(s) is identical or homologous to a corresponding sequence of an antibody derived from another species or belonging to another antibody class or subclass (insofar as they exhibit the desired biological activity) (U.S. Patent No. 4,816,567; Morrison et al., Proc. Natl Acad. Sci. USA, 81:6851-6855 (1984)). Chimeric antibodies of interest in this specification include PRIMATTZFD® antibodies, in which the antigen-binding region of the antibody is derived, for example, from an antibody produced by immunizing a macaque monkey with the antigen of interest. As used herein, "humanized antibodies" are used as a subset of "chimeric antibodies".

[0092] The “humanized” form of a non-human (e.g., mouse) antibody is a chimeric antibody containing a minimal number of sequences derived from non-human immunoglobulin. In some embodiments, the humanized antibody is a human immunoglobulin (recipient antibody) in which residues derived from the recipient’s HVR (as defined below) are replaced with residues (donor antibody) derived from a non-human species, e.g., mouse, rat, rabbit, or non-human primate, possessing the desired specificity, affinity, and / or capabilities. In some examples, framework ("FR") residues of the human immunoglobulin are replaced with corresponding non-human residues. Furthermore, the humanized antibody may contain residues not found in the recipient antibody or donor antibody. These modifications may be made to further improve antibody performance, such as binding affinity. Generally, humanized antibodies contain substantially all of at least one, usually two, variable domains, where all or substantially all of the hypervariable loops in these variable domains correspond to non-human immunoglobulin sequences, and all or substantially all of the FR regions correspond to human immunoglobulin sequences, although the FR regions may contain one or more substitutions of individual FR residues that improve antibody performance, such as binding affinity, isomerization, and immunogenicity. The number of these amino acid substitutions in the FR is usually 6 or less in the H chain and 3 or less in the L chain. Humanized antibodies also optionally contain at least a portion of the immunoglobulin constant region (Fc), usually at least a portion of human immunoglobulin. Suitable human acceptor antibodies may be selected from conventional databases, such as the KABAT database, Los Alamos database, AbM, and Swiss Protein database, based on homology to the nucleotide and amino acid sequences of the donor antibody. Human antibodies characterized by homology (based on amino acids) to the framework region of the donor antibody may be suitable for providing a heavy chain constant region and / or a heavy chain variable framework region for insertion of the donor CDR. Suitable acceptor antibodies capable of providing a light chain constant region or a variable framework region can be selected in a similar manner. It should be noted that the heavy and light chains of the acceptor antibody do not need to originate from the same acceptor antibody.Prior art describes several methods for producing such humanized antibodies (see, for example, EP-A-0239400 and EP-A-054951). For further details, see, for example, Jones et al., Nature 321:522-525 (1986); Riechmann et al., Nature 332:323-329 (1988); and Presta, Curr. Op. Struct. Biol. 2:593-596 (1992). See, for example, Vaswani and Hamilton, Ann. Allergy, Asthma & Immunol. 1:105-115 (1998), Harris, Biochem. Soc. Transactions 23:1035-1038 (1995), Hurle and Gross, Curr. Op. Biotech. 5:428-433 (1994), and U.S. Patents No. 6,982,321 and 7,087,409.

[0093] A “human antibody” is an antibody having an amino acid sequence corresponding to the amino acid sequence of an antibody produced by a human, and / or an antibody produced using any of the human antibody production techniques disclosed herein. This definition of a human antibody specifically excludes humanized antibodies containing non-human antigen-binding residues. Human antibodies can be produced using various techniques known in the art, including phage display libraries. Hoogenboom and Winter, J.Mol.Biol., 227:381 (1991); Marks et al., J.Mol.Biol., 222:581 (1991). The method described in Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, p.77 (1985); Boerner et al., J.Immunol., 147(1):86-95 (1991) is also available for the preparation of human monoclonal antibodies. See also van Dijk and van de Winkel, Curr. Opin. Pharmacol. 5:368-74 (2001). Human antibodies can be prepared by administering antigens to transgenic animals, such as immunized xenomouses, in which the endogenous locus has been deactivated while the transgenic animals have been modified to produce such antibodies in response to antigen exposure (see, for example, U.S. Patents 6,075,181 and 6,150,584 relating to XENOMOUSE® technology). See also, for example, Li et al., Proc. Natl Acad. Sci. USA, 103:3557-3562 (2006), relating to human antibodies produced by human B-cell hybridoma technology.

[0094] As used herein, “treatment” or “to treat” means an effort to achieve a beneficial or desired outcome, including clinical outcomes. In line with the spirit of this application, beneficial or desired clinical outcomes include, but are not limited to, one or more of the following: reducing one or more symptoms caused by the disease; reducing the severity of the disease; stabilizing the disease (e.g., preventing or delaying disease exacerbation); preventing or delaying the spread of the disease; delaying the onset or recurrence of the disease; delaying or slowing the progression of the disease; improving the state of the disease; achieving remission (whether partial or complete) of the disease; reducing the dosage of one or more other drugs required to treat the disease; delaying disease progression; improving quality of life; and / or extending survival. Reduction of the pathological outcomes of the disease is also included in “treatment.” The methods of this application are intended to address any one or more of these aspects of treatment.

[0095] The terms “individual,” “subject,” and “patient” are used interchangeably herein to describe mammals, including humans. In some embodiments, the individual is a human. In some embodiments, the individual is suffering from a disease or condition (e.g., cancer). In some embodiments, the individual requires treatment.

[0096] As understood in the art, “effective dose” refers to the amount of drug (e.g., a targeted conjugate) sufficient to produce a desired therapeutic outcome (e.g., reducing the severity or duration of cancer, stabilizing the severity of cancer, or eliminating one or more symptoms of cancer) or a desired diagnostic outcome. In therapeutic use, beneficial or desired outcomes include, for example, reducing one or more symptoms of a disease (biochemical, histological, and / or behavioral symptoms) (including its complications and intermediate pathological phenotypes seen in the course of disease development), improving the quality of life of a person with a disease, reducing the dose of other drugs required to treat the disease, enhancing the effect of another drug, delaying disease progression, and / or extending the patient’s survival time. In some embodiments, an effective dose of a drug may extend survival time (including overall survival and progression-free survival), produce an objective response (including complete or partial response), reduce one or more signs or symptoms of a disease or condition to some extent, and / or improve the quality of life of the subject.

[0097] With respect to polypeptide sequences and antibody sequences identified herein, “amino acid sequence identity percentage (%)” is defined as the percentage of amino acid residues in a candidate sequence that are identical to the amino acid residues of the polypeptide sequence being compared, after sequence alignment, and any conservative substitutions are considered as part of sequence identity. Alignment for the purpose of determining the amino acid sequence identity percentage can be achieved in various ways within the scope of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, Megalign (DNASTAR), or MUSCLE software. Those skilled in the art can determine appropriate parameters for evaluating the alignment, including any algorithm necessary to obtain the maximum alignment over the entire length of the sequences being compared. However, for the purposes of this specification, the % amino acid sequence identity value is generated using the sequence comparison computer program MUSCLE (Edgar, RC, Nucleic Acids Research 32(5):1792-1797, 2004; Edgar, RC, BMC Bioinformatics 5(1):113, 2004 (each of these is incorporated herein by reference in whole for all purposes)).

[0098] Amino acid substitutions include, but are not limited to, the substitution of one amino acid with another in a polypeptide. Exemplary substitutions are shown in Table 1. Amino acid substitutions can be introduced into antibodies of interest, and the product can be screened for desired activity, such as retention / improvement of antigen binding, decreased immunogenicity, or improvement of ADCC or CDC. [Table 1] Amino acids can be classified according to the following common side-chain characteristics. (1) Hydrophobic: norleucine, met, ala, val, leu, ile; (2) Neutral hydrophilic: Cys, Ser, Thr, Asn, Gln; (3) Acidic: asp, glu; (4) Basicity: His, Lys, Arg; (5) Residues that affect the direction of the chain: gly, pro; (6) Aromatic: Trp, Tyr, Phe. Non-conservative substitution involves replacing one member of one of these classes with one of another.

[0099] The terms “polypeptide” or “peptide” are used herein to encompass all types of naturally occurring and synthetic proteins, including protein fragments of all lengths, fusion proteins, and modified proteins, such as, but not limited to, glycoproteins and all other types of modified proteins (e.g., proteins obtained by phosphorylation, acetylation, myristoylation, palmitoylation, glycosylation, oxidation, formylation, amidation, polyglutamylation, ADP-ribosylation, PEGylation, biotinylation, etc.).

[0100] The term "fusion" refers to the genetic linking of two polypeptide fragments to provide a single, continuous polypeptide ("fusion polypeptide"). The two polypeptide fragments may be linked directly to each other or via another polypeptide placed between them. Standard recombinant DNA techniques or chemical gene synthesis may be used to provide the nucleic acid that genetically encodes the fusion polypeptide.

[0101] The term “epitope,” as used herein, refers to a specific group of atoms or amino acids on an antigen to which an antibody binds. Two antibodies or antigen-binding fragments may bind to the same epitope within an antigen if they exhibit competitive binding to the antigen.

[0102] As used herein, the terms “specifically bind,” “specifically recognize,” and “specific” refer to a measurable and reproducible interaction, e.g., binding, between a target and a targeting moiety (e.g., a targeted peptide or antibody or its antigen-binding fragment). In certain embodiments, specific binding determines the presence of a target in the presence of a heterogeneous population of molecules, including biomolecules (e.g., cell surface receptors). For example, a targeting moiety that specifically recognizes a target (which may be an epitope) is a targeting moiety (e.g., an antibody) that binds to this target with higher affinity, avidity, more readily, and / or for a longer duration compared to its binding to other molecules. In some embodiments, the degree of binding of a targeting moiety to unrelated molecules is less than 10% of its binding to the target, as measured, for example, by radioimmunoassay (RIA). In some embodiments, a targeting moiety that specifically binds to a target is 10 -5 M or less, 10 -6 M or less, 10 -7 M or less, 10 -8 M or less, 10 -9 M or less, 10 -10 M or less, 10 -11 M or less, or 10 -12 It has a dissociation constant (KD) of M or less. In some embodiments, the targeting moiety specifically binds to an epitope on a protein that is conserved between proteins of different species. In some embodiments, specific binding may include, but is not required, exclusive binding. The binding specificity of the targeting moiety can be determined experimentally by methods known in the art. Such methods include, but are not limited to, Western blotting, ELISA, RIA, ECL, IRMA, EIA, BIACORE®, and peptide scanning.

[0103] As used herein, “to link,” “to conjugate,” “to ligate,” and “to fuse” are used interchangeably to refer to the linking of two chemical moieties by either covalent or non-covalent bonds. The linking may be direct or indirect, for example, via a linker.

[0104] The term "pharmaceutical composition" refers to a preparation that enables the biological activity of its active ingredients to be effective, and that does not contain additional ingredients that are unacceptably toxic to the subject to which the preparation is administered.

[0105] A "pharmaceutically acceptable carrier" refers to one or more components other than the active ingredient in a pharmaceutical formulation that are non-toxic to the target. Examples of pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers, cryoprotective agents, tonic agents, preservatives, and combinations thereof. Pharmaceutically acceptable carriers or excipients preferably meet the necessary criteria for toxicity and manufacturing testing and / or are included in an Inactive Ingredient Guide prepared by the U.S. Food and Drug Administration or another state / federal government, or are listed in the United States Pharmacopeia or other generally recognized pharmacopoeias for use in mammals, more specifically in humans.

[0106] The term “package insert” is used to refer to the instructions for use that are customarily included on the market packaging of a therapeutic product, which include information on indications, usage, dosage, administration, combination therapy, contraindications, and / or warnings regarding the use of such therapeutic product.

[0107] "Manufactured Article" is any manufactured product (e.g., package or container) or kit comprising at least one reagent, e.g., a drug for the treatment of a disease or condition (e.g., cancer) or a probe for the specific detection of a biomarker described herein. In certain embodiments, the manufactured product or kit may be advertised, distributed or sold as a unit for carrying out the method described herein.

[0108] It will be understood that the embodiments of the present invention described herein include embodiments consisting of and / or essentially consisting of.

[0109] In this specification, references to values ​​or parameters "about" include (and describe) variations relating to the value or parameter itself. For example, a statement referring to "about X" includes a statement of "X".

[0110] As used herein, any reference to a value or parameter that is not a certain value or parameter generally means and describes something that is "other than" a certain value or parameter. For example, "This method is not used to treat disease of type X" means that this method is used to treat disease of a type other than X.

[0111] As used herein, the term "approximately X to Y" has the same meaning as "approximately X to approximately Y".

[0112] As used herein and in the appended claims, the singular forms "a," "an," or "the" include plural nouns unless explicitly indicated by the context.

[0113] As used herein, terms such as “and / or” and “A and / or B” are intended to encompass both A and B; A or B; A (alone); and B (alone). Similarly, as used herein, terms such as “and / or” and “A, B, and / or C” are intended to encompass each of the following specific expressions: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0114] II. Multiple Specificity Targeted Conjugates One aspect of this application provides a multispecific targeted conjugate comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2 (e.g., an sdAb, e.g., VHH), and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188) (hereinafter also referred to as the "PD-L1 / Trop-2 multispecific targeted conjugate"). In some embodiments, the multispecific targeted conjugate further includes a cleavage site between the first targeting moiety and the conjugation site, wherein the second targeting moiety conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site. Accordingly, in some embodiments, a multispecific targeted conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2 (e.g., sdAb, e.g., VHH), and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), wherein the multispecific targeted conjugate further comprises a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeting moiety and the conjugation site, and the second targeting moiety conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site.In some embodiments, a multispecific targeting conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2 (e.g., an sdAb, e.g., VHH), and an effector molecule (e.g., exatecan), wherein the effector molecule conjugates to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188). The multispecific targeted conjugate includes an arbitrary cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeted portion and the conjugation site, and the second targeted portion conjugated with the effector molecule may be released from the multispecific targeted conjugate by cleavage at the arbitrary cleavage site, and the multispecific targeted conjugate has the structure of formula 1: [ka] (wherein A1 is a first targeting region, A2 is a second targeting region, P is an arbitrary cleavage site, C is a conjugation site, L is a linker, D is an effector molecule, x=0 or 1, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)). In some embodiments, cleavage is induced by conditions at the target site. In some embodiments, conditions at the target site are selected from the group consisting of proteases, pH changes, redox changes, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof. In some embodiments, the target site is a disease site. In some embodiments, the disease is a tumor (e.g., a solid tumor), and the conditions are the tumor microenvironment. In some embodiments, cleavage occurs extracellularly, for example, outside tumor cells within the tumor microenvironment. In some embodiments, the second targeting moiety comprises one or more (e.g., 1, 2, 3, 4, or 5, e.g., 2) anti-Trop-2 sdAbs (e.g., VHH). In some embodiments, the first and / or second targeting moiety comprises one or more targeted peptides or antibodies or their antigen-binding fragments. In some embodiments, the antibody or its antigen-binding fragment is selected from the group consisting of full-length antibodies, diabodies, scFv, scFab, Fab, Fab', F(ab')2, single-domain antibodies (sdAbs), dsFv, and combinations thereof. Any of the anti-PD-L1 targeting moieties described herein (see, for example, sections II and III) may be used as the first targeting moiety in a PD-L1 / Trop-2 multispecific targeting conjugate. Any of the anti-Trop-2 targeting moieties described herein (e.g., anti-Trop-2 VHH) (see, for example, sections II and IV) may be used as a second targeting moiety in a PD-L1 / Trop-2 multispecific targeting conjugate. In some embodiments, cleavage is performed by a protease.In some embodiments, the protease is urokinase plasminogen activator (uPA), regmine, plasmin, TMPRSS3, TMPRSS4, TMPRSS6, MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-10, MMP-12, MMP-13, MMP-14, MT1-MMP, cathepsin D, cathepsin K, cathepsin S, ADAM10, ADAM12, ADA The protease is selected from the group consisting of MTS, caspase-1, caspase-2, caspase-3, caspase-4, caspase-5, caspase-6, caspase-7, caspase-8, caspase-9, caspase-10, caspase-11, caspase-12, caspase-13, caspase-14, TACE, human neutrophil elastase, β-secretase, fibroblast-associated protein, matryptase, PSMA, and PSA. In some embodiments, the protease is MMP, for example, MMP-9. In some embodiments, the cleavage site that can be cleaved by MMP includes any of the amino acid sequences of SEQ ID NOs. 71 and 137-143. In some embodiments, the cleavage site that can be cleaved by MMP-9 includes the amino acid sequence of SEQ ID NOs. 71 or 142. In some embodiments, the protease is uPA. In some embodiments, the cleavage site that can be cleaved by uPA includes the amino acid sequence of SEQ ID NOs. 214. In some embodiments, the effector molecule is a therapeutic agent or oligonucleotide, for example, a therapeutic agent. In some embodiments, the therapeutic agent is selected from the group consisting of protein-based drugs, small molecule drugs, cytotoxic agents, toxins, immunomodulators, anti-inflammatory agents, anti-infective agents, and epigenetic modulators. In some embodiments, the therapeutic agent is exatecan. In some embodiments, the multispecific targeted conjugate comprises two or more effector molecules. In some embodiments, x is 0. In some embodiments, x is 1. In some embodiments, the first targeting portion is located at the N-terminus of the second targeting portion. In some embodiments, the conjugation site includes a transglutaminase conjugation site.In some embodiments, the transglutaminase conjugation site includes the amino acid sequence of any of SEQ ID NOs: 145-196, for example, SEQ ID NO: 147 or 188. In some embodiments, the transglutaminase conjugation site includes two or more glutamine-containing tags fused in series with each other. In some embodiments, L is the formula:(Gly). n -(PEG) m -VC-PAB-(DMAE) k (wherein n, m, and k are integers, n≧1, m≧2, and k is 0 or 1) is expressed by the formula: (Gly) n -(PEG) m -VA-PAB-(DMAE) k (wherein n, m, and k are integers, n≧1, m≧2, and k is 0 or 1) is expressed as follows: In some embodiments, L is (Gly)6-amide-PEG8-VA-PAB. In some embodiments, L is (Gly)6-amide-PEG8-VC-PAB. In some embodiments, L-(D) a The structure of equation A is: [ka] (In the formula, the wavy line indicates the site of covalent binding to conjugation site C) is included. In some embodiments, the second targeting moiety includes one or more (e.g., 1 or 2) sdAbs (anti-Trop-2 sdAbs, e.g., anti-Trop-2 VHH) that specifically recognize Trop-2. In some embodiments, the second targeting moiety includes a first anti-Trop-2 sdAb (e.g., VHH) and a second anti-Trop-2 sdAb (e.g., VHH) fused in series with each other. In some embodiments, the first anti-Trop-2 sdAb (e.g., VHH) and the second anti-Trop-2 sdAb (e.g., VHH) bind to different Trop-2 epitopes. In some embodiments, the first targeting moiety includes one or more antibodies or antigen-binding fragments (anti-PD-L1 antibodies or antigen-binding fragments) that specifically recognize PD-L1.

[0115] A portion of a multispecific targeted conjugate described herein that does not contain an effector molecule (or additionally does not contain an effector molecule linker) is also referred to herein as the “targeting portion” or “multispecific targeted portion,” which includes a first targeted portion, a second targeted portion, and optionally, a conjugation site and / or cleavage site. For example, A1-(P) of Formula 1 x -The C-A2 portion is referred to as the targeted portion in this specification.

[0116] In some embodiments, a multispecific targeting conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2 (e.g., an sdAb, e.g., VHH), and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), and the first targeting moiety independently comprises i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, and H-CDR1 containing the amino acid sequence of SEQ ID NO: 8. - CDR2, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) comprising one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments thereof, including H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249.In some embodiments, a multispecific targeting conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2 (e.g., sdAb, e.g., VHH), and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), wherein the multispecific targeting conjugate further comprises a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeting moiety and the conjugation site, and the second targeting moiety conjugated with the effector molecule The first targeted portion may be released from the multispecifically targeted conjugate by cleavage at the cleavage site, and independently comprises one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments thereof, i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249.In some embodiments, a multispecific targeting conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2 (e.g., an sdAb, e.g., VHH), and an effector molecule (e.g., exatecan), wherein the effector molecule conjugates to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188). The multispecific targeted conjugate includes an arbitrary cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeted portion and the conjugation site, and the second targeted portion conjugated with the effector molecule may be released from the multispecific targeted conjugate by cleavage at the arbitrary cleavage site, and the multispecific targeted conjugate has the structure of formula 1: [ka] (wherein A1 is a first targeting portion, A2 is a second targeting portion, P is an arbitrary cleavage site, C is a conjugation site, L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule, x=0 or 1, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)), and the first targeting portion independently includes i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, and H-CDR2 containing the amino acid sequence of SEQ ID NO: 13 ii) comprising one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments thereof, comprising H-CDR3, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249. In some embodiments, one or more anti-PD-L1 antibodies or their antigen-binding fragments are independently: i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 222; viii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 222;x) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 224; xii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 225; xiii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 226; xiv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 224; xvi) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 226; xvii) Sequence number VH containing the amino acid sequence of sequence number 227 and VL containing the amino acid sequence of sequence number 226; xviii) VH containing the amino acid sequence of sequence number 228 and VL containing the amino acid sequence of sequence number 226; xix) VH containing the amino acid sequence of sequence number 229 and VL containing the amino acid sequence of sequence number 226; xx) VH containing the amino acid sequence of sequence number 230 and VL containing the amino acid sequence of sequence number 226; xxi) VH containing the amino acid sequence of sequence number 230 and VL containing the amino acid sequence of sequence number 220; xxii) VH containing the amino acid sequence of sequence number 219 and VL containing the amino acid sequence of sequence number 226; or xxiii) VH containing the amino acid sequence of sequence number 250 and VL containing the amino acid sequence of sequence number 251. In some embodiments, the first targeting portion comprises a full-length anti-PD-L1 antibody. In some embodiments, the full-length anti-PD-L1 antibody comprises: i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) a heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) a heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) a heavy chain containing the amino acid sequence of SEQ ID NO: 87 and a light chain containing the amino acid sequence of SEQ ID NO: 88; vii) a heavy chain containing the amino acid sequence of SEQ ID NO: 91 and a light chain containing the amino acid sequence of SEQ ID NO: 92;viii) Heavy chain containing the amino acid sequence of SEQ ID NO: 93 and light chain containing the amino acid sequence of SEQ ID NO: 94; ix) Heavy chain containing the amino acid sequence of SEQ ID NO: 95 and light chain containing the amino acid sequence of SEQ ID NO: 96; x) Heavy chain containing the amino acid sequence of SEQ ID NO: 97 and light chain containing the amino acid sequence of SEQ ID NO: 98; xi) Heavy chain containing the amino acid sequence of SEQ ID NO: 99 and light chain containing the amino acid sequence of SEQ ID NO: 100; xii) Heavy chain containing the amino acid sequence of SEQ ID NO: 101 and light chain containing the amino acid sequence of SEQ ID NO: 102; xiii) Heavy chain containing the amino acid sequence of SEQ ID NO: 103 and light chain containing the amino acid sequence of SEQ ID NO: 104; xiv) Heavy chain containing the amino acid sequence of SEQ ID NO: 105 and light chain containing the amino acid sequence of SEQ ID NO: 106; xv) Heavy chain containing the amino acid sequence of SEQ ID NO: 107 and light chain containing the amino acid sequence of SEQ ID NO: 108; xvi) Sequence xvii) Heavy chain containing amino acid sequence number 109 and light chain containing amino acid sequence number 110; xviii) Heavy chain containing amino acid sequence number 111 and light chain containing amino acid sequence number 112; xix) Heavy chain containing amino acid sequence number 115 and light chain containing amino acid sequence number 116; xx) Heavy chain containing amino acid sequence number 117 and light chain containing amino acid sequence number 118; xxi) Heavy chain containing amino acid sequence number 119 and light chain containing amino acid sequence number 120; xxii) Heavy chain containing amino acid sequence number 121 and light chain containing amino acid sequence number 122; xxiii) Heavy chain containing amino acid sequence number 17 and light chain containing amino acid sequence number 34;Alternatively, (xxiv) the molecule comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 241 and a light chain containing the amino acid sequence of SEQ ID NO: 242. In some embodiments, cleavage is induced by conditions at a target site. In some embodiments, conditions at the target site are selected from the group consisting of proteases, pH changes, redox changes, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof. In some embodiments, the target site is a disease site such as a tumor (e.g., a solid tumor). In some embodiments, the conditions are the tumor microenvironment. In some embodiments, cleavage is induced by a protease, e.g., MMP (e.g., MMP-9) or uPA. In some embodiments, cleavage sites that can be cleaved by MMP include any of the amino acid sequences of SEQ ID NO: 71 and 137-143. In some embodiments, cleavage sites that can be cleaved by MMP-9 include the amino acid sequence of SEQ ID NO: 71. In some embodiments, cleavage sites that can be cleaved by uPA include the amino acid sequence of SEQ ID NO: 214. In some embodiments, the effector molecule is a therapeutic agent or oligonucleotide, e.g., a therapeutic agent. In some embodiments, the conjugation site includes a transglutaminase conjugation site comprising, for example, any amino acid sequence of SEQ ID NOs. 145-196, e.g., SEQ ID NOs. 147 or 188. In some embodiments, the first targeting moiety is located at the N-terminus of the second targeting moiety. In some embodiments, the second targeting moiety comprises one or more (e.g., 1 or 2) anti-Trop-2 sdAbs (e.g., VHH). In some embodiments, the second targeting moiety comprises a first anti-Trop-2 sdAb and a second anti-Trop-2 sdAb fused in series with each other. In some embodiments, the first anti-Trop-2 sdAb and the second anti-Trop-2 sdAb bind to different Trop-2 epitopes. In some embodiments, one or more anti-Trop-2 sdAbs (e.g., VHH) are fused to the C-terminus of one or both heavy chains of a full-length anti-PD-L1 antibody (e.g., interconnected by a conjugation site). In some embodiments, a linker-effector molecular conjugate (L-(D); a) contains any of formulas A to J, for example, the structure of formula A.

[0117] In some embodiments, a multispecific targeting conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2, and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), wherein the second targeting moiety independently comprises i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; and ii) the amino acid sequence of SEQ ID NO: 53 iii) comprising one or more (e.g., one or two) VHH (anti-Trop-2 VHH) that specifically recognize Trop-2, comprising CDR1 containing the amino acid sequence, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iv) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; or v) comprising one or more (e.g., one or two) VHH (anti-Trop-2 VHH) that specifically recognize Trop-2, including CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218.In some embodiments, a multispecific targeted conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2, and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), wherein the multispecific targeted conjugate further comprises a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeting moiety and the conjugation site, wherein the second targeting moiety conjugated with the effector molecule may be released from the multispecific targeted conjugate by cleavage at the cleavage site. The second targeted portion independently comprises one or more (e.g., one or two) VHHs (anti-Trop-2 VHHs) that specifically recognize Trop-2, including i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218.In some embodiments, a multispecific targeted conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2, and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), and the multispecific targeted conjugate includes an arbitrary cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeting moiety and the conjugation site, and the second targeting moiety conjugated with the effector molecule may be released from the multispecific targeted conjugate by cleavage at the arbitrary cleavage site, and the multispecific targeted conjugate has the structure of Formula 1: [ka] (wherein A1 is the first targeting portion, A2 is the second targeting portion, P is an arbitrary cleavage site, C is a conjugation site, L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule, x=0 or 1, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)), and the second targeting portion independently comprises i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, sequence The following are examples of the following embodiments, comprising one or more (e.g., 1 or 2) VHHs (anti-Trop-2 VHHs) that specifically recognize Trop-2, comprising: CDR2 containing the amino acid sequence of sequence number 58, and CDR3 containing the amino acid sequence of sequence number 63; iii) CDR1 containing the amino acid sequence of sequence number 37, CDR2 containing the amino acid sequence of sequence number 205, and CDR3 containing the amino acid sequence of sequence number 47; iv) CDR1 containing the amino acid sequence of sequence number 53, CDR2 containing the amino acid sequence of sequence number 58, and CDR3 containing the amino acid sequence of sequence number 206; or v) CDR1 containing the amino acid sequence of sequence number 216, CDR2 containing the amino acid sequence of sequence number 217, and CDR3 containing the amino acid sequence of sequence number 218. In some embodiments, one or more anti-Trop-2 VHHs independently comprise one of the amino acid sequences of sequence numbers 50, 66, 123-136, and 203. In some embodiments, the second targeting moiety comprises a first anti-Trop-2 VHH and a second anti-Trop-2 VHH fused in series with each other. In some embodiments, the first anti-Trop-2 VHH and the second anti-Trop-2 VHH bind to different Trop-2 epitopes. In some embodiments, the first targeting moiety is located at the N-terminus of the second targeting moiety. In some embodiments, the first targeting moiety comprises a full-length anti-PD-L1 antibody. In some embodiments, one or more anti-Trop-2 VHH molecules are fused to the C-terminus of one or both heavy chains of the full-length anti-PD-L1 antibody (e.g., interconnected by conjugation sites).In some embodiments, cleavage is induced by conditions at the target site. In some embodiments, the conditions at the target site are selected from the group consisting of proteases, pH changes, redox changes, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof. In some embodiments, the target site is a disease site such as a tumor (e.g., a solid tumor). In some embodiments, the conditions are the tumor microenvironment. In some embodiments, cleavage is induced by a protease, e.g., MMP (e.g., MMP-9) or uPA. In some embodiments, the cleavage site that can be cleaved by MMP includes any of the amino acid sequences of SEQ ID NOs. 71 and 137-143. In some embodiments, the cleavage site that can be cleaved by MMP-9 includes the amino acid sequence of SEQ ID NOs. 71. In some embodiments, the cleavage site that can be cleaved by uPA includes the amino acid sequence of SEQ ID NOs. 214. In some embodiments, the effector molecule is a therapeutic agent or oligonucleotide, e.g., a therapeutic agent. In some embodiments, the conjugation site includes a transglutaminase conjugation site comprising, for example, any amino acid sequence of SEQ ID NOs. 145-196, for example, SEQ ID NOs. 147 or 188. In some embodiments, a linker-effector molecular conjugate (L-(D)) is used. a ) contains any of formulas A to J, for example, the structure of formula A.

[0118] In some embodiments, a multispecific targeting conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2, and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), and the first targeting moiety independently comprises i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, The second targeted portion comprises one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments, each containing H-CDR3 containing the amino acid sequence of sequence number 246, L-CDR1 containing the amino acid sequence of sequence number 247, L-CDR2 containing the amino acid sequence of sequence number 248, and L-CDR3 containing the amino acid sequence of sequence number 249, wherein the second targeted portion independently comprises: i) CDR1 containing the amino acid sequence of sequence number 37, CDR2 containing the amino acid sequence of sequence number 42, and CDR3 containing the amino acid sequence of sequence number 47; ii) CDR1 containing the amino acid sequence of sequence number 53, CDR2 containing the amino acid sequence of sequence number 58, and CDR3 containing the amino acid sequence of sequence number 63; iii) CDR1 containing the amino acid sequence of sequence number 37, CDR2 containing the amino acid sequence of sequence number 205, and CDR3 containing the amino acid sequence of sequence number 47; iv) CDR1 containing the amino acid sequence of sequence number 53, CDR2 containing the amino acid sequence of sequence number 58, and CDR3 containing the amino acid sequence of sequence number 206;Or (v) comprising one or more (e.g., one or two) VHHs (anti-Trop-2 VHHs) that specifically recognize Trop-2, including CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, a multispecific targeted conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2, and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), and the multispecific targeted conjugate further comprises a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeting moiety and the conjugation site, and the second targeting moiety conjugated with the effector molecule may be released from the multispecific targeted conjugate by cleavage at the cleavage site, and the first targeting moiety is independently i) comprising one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments, each comprising: i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249, wherein the second targeted portion independently comprises: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47;ii) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) comprising one or more (e.g., one or two) VHHs (anti-Trop-2 VHHs) that specifically recognize Trop-2, including CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, a multispecific targeted conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), a second targeting moiety that specifically recognizes Trop-2, and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), and the multispecific targeted conjugate includes an arbitrary cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214) between the first targeting moiety and the conjugation site, and the second targeting moiety conjugated with the effector molecule may be released from the multispecific targeted conjugate by cleavage at the arbitrary cleavage site, and the multispecific targeted conjugate has the structure of Formula 1: [ka] (wherein A1 is the first targeting portion, A2 is the second targeting portion, P is an arbitrary cleavage site, C is a conjugation site, L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule, x=0 or 1, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)), and the first targeting portion independently includes i ) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, and the amino acid sequence of SEQ ID NO: 246 The second targeted portion comprises one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments, each containing H-CDR3 containing the amino acid sequence of SEQ ID NO: 247, L-CDR1 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249, wherein the second targeted portion independently comprises: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206;Or (v) comprising one or more (e.g., one or two) VHHs (anti-Trop-2 VHHs) that specifically recognize Trop-2, including CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, one or more anti-PD-L1 antibodies or their antigen-binding fragments are independently: i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 222; viii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 and SEQ ID NO: VL containing the amino acid sequence of 222;x) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 222;xi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 224;xii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 225;xiii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 226;xiv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 225;xv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 224;xvi) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 226;xvii) VH containing the amino acid sequence of SEQ ID NO: 227 and VL containing the amino acid sequence of SEQ ID NO: 226;xviii) VH containing the amino acid sequence of SEQ ID NO: 228 and VL containing the amino acid sequence of SEQ ID NO: 226;xix) VH containing the amino acid sequence of SEQ ID NO: 229 and VL containing the amino acid sequence of SEQ ID NO: 226; xx) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 226; xxi) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 220; xxii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 226; or xxiii) VH containing the amino acid sequence of SEQ ID NO: 250 and VL containing the amino acid sequence of SEQ ID NO: 251. In some embodiments, the first targeting moiety comprises a full-length anti-PD-L1 antibody. In some embodiments, the full-length anti-PD-L1 antibody comprises: i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) a heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) a heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) a heavy chain containing the amino acid sequence of SEQ ID NO: 87 and a light chain containing the amino acid sequence of SEQ ID NO: 88; vii) a heavy chain containing the amino acid sequence of SEQ ID NO: 91 and a light chain containing the amino acid sequence of SEQ ID NO: 92; viiii) a heavy chain containing the amino acid sequence of SEQ ID NO: 93 and a light chain containing the amino acid sequence of SEQ ID NO: 94 Including a light chain; ix) A heavy chain containing the amino acid sequence of SEQ ID NO: 95 and a light chain containing the amino acid sequence of SEQ ID NO: 96; x) A heavy chain containing the amino acid sequence of SEQ ID NO: 97 and a light chain containing the amino acid sequence of SEQ ID NO: 98; xi) A heavy chain containing the amino acid sequence of SEQ ID NO: 99 and a light chain containing the amino acid sequence of SEQ ID NO: 100; xii) A heavy chain containing the amino acid sequence of SEQ ID NO: 101 and a light chain containing the amino acid sequence of SEQ ID NO: 102; xiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 103 and a light chain containing the amino acid sequence of SEQ ID NO: 104; xiv) A heavy chain containing the amino acid sequence of SEQ ID NO: 105 and a light chain containing the amino acid sequence of SEQ ID NO: 106; xv) A heavy chain containing the amino acid sequence of SEQ ID NO: 107 and a light chain containing the amino acid sequence of SEQ ID NO: 108; xvi) A heavy chain containing the amino acid sequence of SEQ ID NO: 109 and a light chain containing the amino acid sequence of SEQ ID NO: 110;xvii) Heavy chain containing the amino acid sequence of SEQ ID NO: 111 and light chain containing the amino acid sequence of SEQ ID NO: 112; xviii) Heavy chain containing the amino acid sequence of SEQ ID NO: 113 and light chain containing the amino acid sequence of SEQ ID NO: 114; xix) Heavy chain containing the amino acid sequence of SEQ ID NO: 115 and light chain containing the amino acid sequence of SEQ ID NO: 116; xx) Heavy chain containing the amino acid sequence of SEQ ID NO: 117 and light chain containing the amino acid sequence of SEQ ID NO: 118; xxi) Heavy chain containing the amino acid sequence of SEQ ID NO: 119 and light chain containing the amino acid sequence of SEQ ID NO: 120; xxii) Heavy chain containing the amino acid sequence of SEQ ID NO: 121 and light chain containing the amino acid sequence of SEQ ID NO: 122; xxiii) Heavy chain containing the amino acid sequence of SEQ ID NO: 17 and light chain containing the amino acid sequence of SEQ ID NO: 34;Alternatively, it comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 241 and a light chain containing the amino acid sequence of SEQ ID NO: 242. In some embodiments, one or more anti-Trop-2 VHHs independently contain any of the amino acid sequences of SEQ ID NOs: 50, 66, 123-136, and 203. In some embodiments, the second targeting moiety comprises a first anti-Trop-2 VHH and a second anti-Trop-2 VHH fused in series with each other. In some embodiments, the first anti-Trop-2 VHH and the second anti-Trop-2 VHH bind to different Trop-2 epitopes. In some embodiments, the first targeting moiety is located at the N-terminus of the second targeting moiety. In some embodiments, one or more anti-Trop-2 VHHs are fused to the C-terminus of one or both heavy chains of the full-length anti-PD-L1 antibody (e.g., interconnected by a conjugation site). In some embodiments, cleavage is induced by conditions at the target site. In some embodiments, the conditions at the target site are selected from the group consisting of proteases, pH changes, redox changes, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof. In some embodiments, the target site is a disease site such as a tumor (e.g., a solid tumor). In some embodiments, the conditions are the tumor microenvironment. In some embodiments, cleavage is performed by a protease, e.g., MMP (e.g., MMP-9) or uPA. In some embodiments, the cleavage site that can be cleaved by MMP includes any of the amino acid sequences of SEQ ID NOs. 71 and 137-143. In some embodiments, the cleavage site that can be cleaved by MMP-9 includes the amino acid sequence of SEQ ID NOs. 71. In some embodiments, the cleavage site that can be cleaved by uPA includes the amino acid sequence of SEQ ID NOs. 214. In some embodiments, the effector molecule is a therapeutic agent or oligonucleotide, e.g., a therapeutic agent. In some embodiments, the conjugation site includes a transglutaminase conjugation site comprising, for example, any amino acid sequence of SEQ ID NOs. 145-196, for example, SEQ ID NOs. 147 or 188. In some embodiments, a linker-effector molecular conjugate (L-(D); a ) contains any of formulas A to J, for example, the structure of formula A.

[0119] In some embodiments, the first targeting portion (e.g., any anti-PD-L1 targeting portion described herein) is located at the N-terminus of the second targeting portion (e.g., any anti-Trop-2 targeting portion described herein). In some embodiments, the multispecific targeting conjugate includes, from the N-terminus to the C-terminus, i) the first targeting portion - any linker 1 - conjugation site - any linker 2 - second targeting portion; ii) the first targeting portion - any linker 1 - cleavage site - any linker 2 - conjugation site - any linker 3 - second targeting portion; iii) the first targeting portion - any linker 1 - second targeting portion - any linker 2 - conjugation site; or iv) the first targeting portion - any linker 1 - cleavage site - any linker 2 - second targeting portion - any linker 3 - conjugation site. In some embodiments, the first targeting moiety comprises one or more anti-PD-L1 antibodies or their antigen-binding fragments (e.g., full-length antibodies). In some embodiments, the second targeting moiety comprises one or more anti-Trop-2 antibodies or their antigen-binding fragments, e.g., anti-Trop-2 sdAb (e.g., VHH) or scFv. In some embodiments, the multispecific targeting conjugate has the following configurations from the N-terminus to the C-terminus: i) anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - conjugation site - any linker 2 - anti-Trop-2 sdAb (e.g., VHH); ii) anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - cleavage site - any linker 2 - conjugation site - any linker 3 - anti-Trop-2 sdAb (e.g., VHH); iii) anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - conjugation site - any linker 2 - first anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - second anti-Trop-2 sdAb (e.g., VHH); iv) anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - cleavage site - any linker 2 - conjugation site - any linker 3 - first anti-Trop-2 sdAb(e.g., VHH)-any linker 4-second anti-Trop-2 sdAb(e.g., VHH);v) Anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - conjugation site; vi) Anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - cleavage site - any linker 2 - anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - conjugation site; vii) Anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - first anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - second anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - conjugation site; viiii) Anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - first anti-Trop-2 sdAb(e.g., VHH)-any linker 2-conjugation site-any linker 3-second anti-Trop-2 sdAb(e.g., VHH); ix) anti-PD-L1 antibody or its antigen-binding fragment-any linker 1-cleavage site-any linker 2-first anti-Trop-2 sdAb(e.g., VHH)-any linker 3-second anti-Trop-2 sdAb(e.g., VHH)-any linker 4-conjugation site; or x) anti-PD-L1 antibody or its antigen-binding fragment-any linker 1-cleavage site-any linker 2-first anti-Trop-2 sdAb(e.g., VHH)-any linker 3-conjugation site-any linker 4-second anti-Trop-2 sdAb(e.g., VHH). Any peptide linker described herein, for example, any of SEQ ID NOs. 207-209, may be used herein. In some embodiments, the cleavage site is an MMP (e.g., MMP-9) or a uPA cleavage site. In some embodiments, the cleavage site includes any of SEQ ID NOs. 71, 137-143, and 214, for example, the amino acid sequence of SEQ ID NOs. 71 or 214. In some embodiments, the conjugation site includes any of SEQ ID NOs. 145-196, for example, the amino acid sequence of SEQ ID NOs. 147 or 188.

[0120] In some embodiments, the first targeting portion (e.g., any anti-PD-L1 targeting portion described herein) is located at the C-terminus of the second targeting portion (e.g., any anti-Trop-2 targeting portion described herein). In some embodiments, the multispecific targeting conjugate includes, from the N-terminus to the C-terminus, i) the second targeting portion - any linker 1 - conjugation site - any linker 2 - the first targeting portion; ii) the second targeting portion - any linker 1 - conjugation site - any linker 2 - cleavage site - any linker 3 - the first targeting portion; iii) the conjugation site - any linker 1 - the second targeting portion - any linker 2 - the first targeting portion; or iv) the conjugation site - any linker 1 - the second targeting portion - any linker 2 - cleavage site - any linker 3 - the first targeting portion. In some embodiments, the first targeting moiety comprises one or more anti-PD-L1 antibodies or their antigen-binding fragments (e.g., full-length antibodies). In some embodiments, the second targeting moiety comprises one or more anti-Trop-2 antibodies or their antigen-binding fragments, e.g., anti-Trop-2 sdAb (e.g., VHH) or scFv. In some embodiments, the multispecific targeting conjugate has, from the N-terminus to the C-terminus, i) an anti-Trop-2 sdAb (e.g., VHH) - any linker 1 - conjugation site - any linker 2 - an anti-PD-L1 antibody or its antigen-binding fragment; ii) an anti-Trop-2 sdAb (e.g., VHH) - any linker 1 - conjugation site - any linker 2 - cleavage site - any linker 3 - an anti-PD-L1 antibody or its antigen-binding fragment; iii) a first anti-Trop-2 sdAb (e.g., VHH) - any linker 1 - a second anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - conjugation site - any linker 3 - an anti-PD-L1 antibody or its antigen-binding fragment; iv) a first anti-Trop-2 sdAb (e.g., VHH) - any linker 1 - a second anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - conjugation site - any linker 3 - cleavage site - any linker 4 - anti-PD-L1 antibody or its antigen-binding fragment;v) Conjugation site - any linker 1 - anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - anti-PD-L1 antibody or its antigen-binding fragment; vi) Conjugation site - any linker 1 - anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - cleavage site - any linker 3 - anti-PD-L1 antibody or its antigen-binding fragment; vii) Conjugation site - any linker 1 - first anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - second anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - anti-PD-L1 antibody or its antigen-binding fragment; viiii) first anti-Trop-2 sdAb (e.g., VHH) - any linker 1 - conjugation site - any linker 2 - second anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - anti-PD-L1 antibody or its antigen-binding fragment; ix) conjugation site - any linker 1 - first anti-Trop-2 sdAb (e.g., VHH) - any linker 2 - second anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - cleavage site - any linker 4 - anti-PD-L1 antibody or its antigen-binding fragment; or x) first anti-Trop-2 sdAb (e.g., VHH) - any linker 1 - conjugation site - any linker 2 - second anti-Trop-2 sdAb (e.g., VHH) - any linker 3 - cleavage site - any linker 4 - anti-PD-L1 antibody or its antigen-binding fragment. Any peptide linker described herein, e.g., any of SEQ ID NOs. 207-209 may be used herein. In some embodiments, the cleavage site is an MMP (e.g., MMP-9) or a uPA cleavage site. In some embodiments, the cleavage site includes the amino acid sequence of SEQ ID NOs. 71, 137-143, and 214, for example, SEQ ID NOs. 71 or 214. In some embodiments, the conjugation site includes the amino acid sequence of SEQ ID NOs. 145-196, for example, SEQ ID NOs. 147 or 188.

[0121] In some embodiments, the multispecific targeting conjugate comprises one or more anti-Trop-2 sdAbs (e.g., VHH) fused to a full-length anti-PD-L1 antibody. In some embodiments, one or more anti-Trop-2 sdAbs (e.g., VHH) are fused to the C-terminus of one or both heavy chains, the C-terminus of one or both light chains, the N-terminus of one or both heavy chains, and / or the N-terminus of one or both light chains of the full-length anti-PD-L1 antibody. In some embodiments, two or more anti-Trop-2 sdAbs (e.g., VHH) are fused in series with each other and further fused to the C-terminus of one or both heavy chains, the C-terminus of one or both light chains, the N-terminus of one or both heavy chains, and / or the N-terminus of one or both light chains of the full-length anti-PD-L1 antibody. In some embodiments, one or more (e.g., one or two) anti-Trop-2 sdAbs (e.g., VHH) are fused to the C-terminus of each heavy chain of the full-length anti-PD-L1 antibody. In some embodiments, the C-terminus of the heavy chain of the anti-PD-L1 antibody is fused to the N-terminus of the anti-Trop-2 sdAb (e.g., VHH). In some embodiments, the C-terminus of the light chain of the anti-PD-L1 antibody is fused to the N-terminus of the anti-Trop-2 sdAb (e.g., VHH). In some embodiments, the targeting moiety comprises any of the anti-PD-L1 antibodies or their antigen-binding fragments described herein and / or any of the anti-Trop-2 antibodies or their antigen-binding fragments. In some embodiments, the multispecific targeting conjugate comprises, from the N-terminus to the C-terminus, a fusion polypeptide: heavy chain of a full-length anti-PD-L1 antibody - any linker 1 - any cleavage site (e.g., SEQ ID NO: 71 or 214) - any linker 2 - conjugation site (e.g., SEQ ID NO: 147 or 188) - any linker 3 - one or more anti-Trop-2 sdAbs (e.g., VHH).

[0122] In some embodiments, a multispecific targeting conjugate is provided comprising a first targeting moiety that specifically recognizes PD-L1, a second targeting moiety that specifically recognizes Trop-2, and an effector molecule (e.g., exatecan), wherein the effector molecule is conjugated to the second targeting moiety via a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188) by an effector molecule linker (L), and the first targeting moiety is anti-PD-L 1. A full-length antibody, the multispecific targeting conjugate has a fusion polypeptide from the N-terminus to the C-terminus: heavy chain of a full-length anti-PD-L1 antibody - any linker 1 - any cleavage site (e.g., MMP cleavage site, e.g., MMP-9 cleavage site, e.g., SEQ ID NO: 71; or uPA cleavage site, e.g., SEQ ID NO: 214) - any linker 2 - conjugation site (e.g., transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188) - any linker 3 - one or more (e.g., 1 or 2) anti-Trop-2 The fusion polypeptide comprises an sdAb (e.g., VHH), i) the heavy chain comprises the amino acid sequence of SEQ ID NO: 17, the light chain comprises the amino acid sequence of SEQ ID NO: 34, and the fusion polypeptide comprises any of the amino acid sequences of SEQ ID NOs: 67, 69, 215, and 238-240, or ii) the heavy chain comprises the amino acid sequence of SEQ ID NO: 241, the light chain comprises the amino acid sequence of SEQ ID NO: 242, and the fusion polypeptide comprises any of the amino acid sequences of SEQ ID NOs: 231-237. In some embodiments, the fusion polypeptide comprises any of the amino acid sequences of SEQ ID NOs: 67, 69, and 215. In some embodiments, if the multispecific targeting conjugate comprises two anti-Trop-2 sdAbs (e.g., VHH), they may be the same or different and may bind to the same or different Trop-2 epitopes. In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0123] In some embodiments, the structure of Equation 1: [ka] A multispecific targeted conjugate is provided, comprising (wherein A1 is a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting moiety that specifically recognizes Trop-2 (e.g., sdAb, e.g., VHH), P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule (e.g., exatecan), x=0, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)). In some embodiments, the first targeting moiety comprises one or more (e.g., one) anti-PD-L1 antibodies or their antigen-binding fragments, e.g., a full-length anti-PD-L1 antibody. In some embodiments, the structure of Formula 1: [ka] (wherein A1 is a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting moiety that specifically recognizes Trop-2 (e.g., sdAb, e.g., VHH), P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule (e.g., exatecan), x=0, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)) multispecific targeting A conjugate is provided, wherein the first targeted portion independently comprises one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments thereof, comprising: i) H-CDR1 comprising the amino acid sequence of SEQ ID NO: 3, H-CDR2 comprising the amino acid sequence of SEQ ID NO: 8, H-CDR3 comprising the amino acid sequence of SEQ ID NO: 13, L-CDR1 comprising the amino acid sequence of SEQ ID NO: 20, L-CDR2 comprising the amino acid sequence of SEQ ID NO: 25, and L-CDR3 comprising the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 comprising the amino acid sequence of SEQ ID NO: 244, H-CDR2 comprising the amino acid sequence of SEQ ID NO: 245, H-CDR3 comprising the amino acid sequence of SEQ ID NO: 246, L-CDR1 comprising the amino acid sequence of SEQ ID NO: 247, L-CDR2 comprising the amino acid sequence of SEQ ID NO: 248, and L-CDR3 comprising the amino acid sequence of SEQ ID NO: 249. In some embodiments, the second targeting moiety comprises one or more (e.g., one or two) anti-Trop-2 antibodies or their antigen-binding fragments, for example, a first anti-Trop-2 sdAb and a second anti-Trop-2 sdAb fused in series with each other. In some embodiments, the structure of Formula 1: [ka] A multispecific targeting conjugate is provided, comprising (wherein A1 is a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting moiety that specifically recognizes Trop-2, P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule (e.g., exatecan), x=0, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)), wherein the second targeting moiety is independently i) SEQ ID NO: 37 ii) CDR1 containing the amino acid sequence of SEQ ID NO: 42, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218, comprising one or more (e.g., 1 or 2) VHH (anti-Trop-2 VHH) that specifically recognize Trop-2. In some embodiments, the structure of Formula 1: [ka] (In the formula, A1 is a first targeting region that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting region that specifically recognizes Trop-2, P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), and L is a linker (e.g., (Gly)6-amide) A multispecific targeting conjugate is provided, comprising i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, and L containing the amino acid sequence of SEQ ID NO: 20. -CDR1, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) One or more (e.g., one) anti-PD-L1 antibodies comprising H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249. The antigen-binding fragment comprises the second targeting portion, independently comprising: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47;iv) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) comprising one or more (e.g., one or two) VHHs (anti-Trop-2 VHHs) that specifically recognize Trop-2, comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, the structure of Formula 1: [ka] A multispecific targeted conjugate is provided, comprising (wherein A1 is a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting moiety that specifically recognizes Trop-2 (e.g., sdAb, e.g., VHH), P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule (e.g., exatecan), x=1, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)). In some embodiments, the first targeting moiety comprises one or more (e.g., one) anti-PD-L1 antibodies or their antigen-binding fragments, e.g., a full-length anti-PD-L1 antibody. In some embodiments, the structure of Formula 1: [ka] (wherein A1 is a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting moiety that specifically recognizes Trop-2 (e.g., sdAb, e.g., VHH), P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule (e.g., exatecan), x=1, a=1~20 (e.g., 1~10, e.g., 1), b=1-20 (e.g., 2~10 or 4~6)) multispecific targeting A conjugate is provided, wherein the first targeted portion independently comprises one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments thereof, comprising: i) H-CDR1 comprising the amino acid sequence of SEQ ID NO: 3, H-CDR2 comprising the amino acid sequence of SEQ ID NO: 8, H-CDR3 comprising the amino acid sequence of SEQ ID NO: 13, L-CDR1 comprising the amino acid sequence of SEQ ID NO: 20, L-CDR2 comprising the amino acid sequence of SEQ ID NO: 25, and L-CDR3 comprising the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 comprising the amino acid sequence of SEQ ID NO: 244, H-CDR2 comprising the amino acid sequence of SEQ ID NO: 245, H-CDR3 comprising the amino acid sequence of SEQ ID NO: 246, L-CDR1 comprising the amino acid sequence of SEQ ID NO: 247, L-CDR2 comprising the amino acid sequence of SEQ ID NO: 248, and L-CDR3 comprising the amino acid sequence of SEQ ID NO: 249. In some embodiments, the second targeting moiety comprises one or more (e.g., one or two) anti-Trop-2 antibodies or their antigen-binding fragments, for example, a first anti-Trop-2 sdAb and a second anti-Trop-2 sdAb fused in series with each other. In some embodiments, the structure of Formula 1: [ka] A multispecific targeting conjugate is provided, comprising (wherein A1 is a first targeting moiety that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting moiety that specifically recognizes Trop-2, P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), L is a linker (e.g., (Gly)6-amide-PEG8-VA-PAB), D is an effector molecule (e.g., exatecan), x=1, a=1~20 (e.g., 1~10, e.g., 1), and b=1-20 (e.g., 2~10 or 4~6)), where the second targeting moiety is independently i) SEQ ID NO ii) CDR1 containing the amino acid sequence of 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218, comprising one or more (e.g., 1 or 2) VHH (anti-Trop-2 VHH) that specifically recognize Trop-2. In some embodiments, the structure of Formula 1: [ka] (In the formula, A1 is a first targeting region that specifically recognizes PD-L1 (e.g., a full-length antibody), A2 is a second targeting region that specifically recognizes Trop-2, P is a cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), C is a conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), and L is a linker (e.g., (Gly)6-amide) A multispecific targeting conjugate is provided, comprising i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, and L containing the amino acid sequence of SEQ ID NO: 20. -CDR1, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) One or more (e.g., one) anti-PD-L1 antibodies comprising H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249. The antigen-binding fragment comprises the second targeting portion, independently comprising: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47;iv) comprising one or more (e.g., one or two) VHHs (anti-Trop-2 VHHs) that specifically recognize Trop-2, comprising CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, two or more anti-Trop-2 antibodies or their antigen-binding fragments (e.g., sdAb, e.g., VHHs) bind to different Trop-2 epitopes. In some embodiments, one or more anti-PD-L1 antibodies or their antigen-binding fragments are independently: i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 222; vii i) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 222; x) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 224; xii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 225; xiii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 226; xiv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 224xvi) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 226; xvii) VH containing the amino acid sequence of SEQ ID NO: 227 and VL containing the amino acid sequence of SEQ ID NO: 226; xviii) VH containing the amino acid sequence of SEQ ID NO: 228 and VL containing the amino acid sequence of SEQ ID NO: 226; xix) VH containing the amino acid sequence of SEQ ID NO: 229 and VL containing the amino acid sequence of SEQ ID NO: 226; xx) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 226; xxi) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 220; xxii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 226; or xxiii) VH containing the amino acid sequence of SEQ ID NO: 250 and VL containing the amino acid sequence of SEQ ID NO: 251. In some embodiments, the first targeting portion comprises a full-length anti-PD-L1 antibody. In some embodiments, the full-length anti-PD-L1 antibody comprises: i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) a heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) a heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) a heavy chain containing the amino acid sequence of SEQ ID NO: 87 and a light chain containing the amino acid sequence of SEQ ID NO: 88; vii) the amino acid sequence of SEQ ID NO: 91 A heavy chain containing the sequence and a light chain containing the amino acid sequence of SEQ ID NO: 92; viiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 93 and a light chain containing the amino acid sequence of SEQ ID NO: 94; ix) A heavy chain containing the amino acid sequence of SEQ ID NO: 95 and a light chain containing the amino acid sequence of SEQ ID NO: 96; x) A heavy chain containing the amino acid sequence of SEQ ID NO: 97 and a light chain containing the amino acid sequence of SEQ ID NO: 98; xi) A heavy chain containing the amino acid sequence of SEQ ID NO: 99 and a light chain containing the amino acid sequence of SEQ ID NO: 100; xii) A heavy chain containing the amino acid sequence of SEQ ID NO: 101 and a light chain containing the amino acid sequence of SEQ ID NO: 102; xiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 103 and a light chain containing the amino acid sequence of SEQ ID NO: 104;xiv) Heavy chain containing the amino acid sequence of SEQ ID NO: 105 and light chain containing the amino acid sequence of SEQ ID NO: 106; xv) Heavy chain containing the amino acid sequence of SEQ ID NO: 107 and light chain containing the amino acid sequence of SEQ ID NO: 108; xvi) Heavy chain containing the amino acid sequence of SEQ ID NO: 109 and light chain containing the amino acid sequence of SEQ ID NO: 110; xvii) Heavy chain containing the amino acid sequence of SEQ ID NO: 111 and light chain containing the amino acid sequence of SEQ ID NO: 112; xviii) Heavy chain containing the amino acid sequence of SEQ ID NO: 113 and light chain containing the amino acid sequence of SEQ ID NO: 114 ;xix) Heavy chain containing the amino acid sequence of SEQ ID NO: 115 and light chain containing the amino acid sequence of SEQ ID NO: 116;xx) Heavy chain containing the amino acid sequence of SEQ ID NO: 117 and light chain containing the amino acid sequence of SEQ ID NO: 118;xxi) Heavy chain containing the amino acid sequence of SEQ ID NO: 119 and light chain containing the amino acid sequence of SEQ ID NO: 120;xxii) Heavy chain containing the amino acid sequence of SEQ ID NO: 121 and light chain containing the amino acid sequence of SEQ ID NO: 122;xxiii) Heavy chain containing the amino acid sequence of SEQ ID NO: 17 and light chain containing the amino acid sequence of SEQ ID NO: 34;Alternatively, (xxiv) comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 241 and a light chain containing the amino acid sequence of SEQ ID NO: 242. In some embodiments, one or more anti-Trop-2 VHH molecules independently contain any of the amino acid sequences of SEQ ID NOs: 50, 66, 123-136, and 203. In some embodiments, the second targeting moiety comprises a first anti-Trop-2 VHH and a second anti-Trop-2 VHH fused in series with each other. In some embodiments, the first targeting moiety is located at the N-terminus of the second targeting moiety. In some embodiments, cleavage is induced by conditions at the target site. In some embodiments, the conditions at the target site are selected from the group consisting of proteases, pH changes, redox changes, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof. In some embodiments, the target site is a disease site such as a tumor (e.g., a solid tumor). In some embodiments, the conditions are the tumor microenvironment. In some embodiments, cleavage is performed by a protease, e.g., MMP (e.g., MMP-9) or uPA. In some embodiments, the cleavage site that can be cleaved by MMP includes any of the amino acid sequences of SEQ ID NOs. 71 and 137-143. In some embodiments, the cleavage site that can be cleaved by MMP-9 includes the amino acid sequence of SEQ ID NOs. 71. In some embodiments, the cleavage site that can be cleaved by uPA includes the amino acid sequence of SEQ ID NOs. 214. In some embodiments, the effector molecule is a therapeutic agent or oligonucleotide, e.g., a therapeutic agent. In some embodiments, the conjugation site includes a transglutaminase conjugation site, e.g., any of the amino acid sequences of SEQ ID NOs. 145-196, e.g., SEQ ID NOs. 147 or 188. In some embodiments, a linker-effector molecule conjugate (L-(D); a ) contains any of formulas A to J, for example, the structure of formula A.

[0124] In some embodiments, (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, and a first anti-Trop-2 sdAb (e.g., VHH); (b) a second heavy chain of the same full-length anti-PD-L1 antibody, from the N-terminus to the C-terminus, an optional linker 3, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 4, and a second anti-Trop-2 A multispecific targeting conjugate is provided, comprising: (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing a first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing a second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the following polypeptide chains are provided: (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of the full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, and a first anti-Trop-2 sdAb (e.g., VHH); (b) a second polypeptide chain comprising, from the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 3, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 4, and a second anti-Trop-2 sdAb (e.g., VHH); (c) a third polypeptide chain comprising, from the N-terminus to the C-terminus, a first light chain of the full-length anti-PD-L1 antibody; and (d) a fourth polypeptide chain comprising, from the second light chain of the full-length anti-PD-L1 antibody.A multispecific targeted conjugate is provided comprising (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the full-length anti-PD-L1 antibody is i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3 , H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) comprising H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249. In some embodiments, the following polypeptide chains are provided: (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of the full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, and a first anti-Trop-2 sdAb (e.g., VHH); (b) a second polypeptide chain comprising, from the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 3, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 4, and a second anti-Trop-2 sdAb (e.g., VHH); (c) a third polypeptide chain comprising, from the N-terminus to the C-terminus, a first light chain of the full-length anti-PD-L1 antibody; and (d) a fourth polypeptide chain comprising, from the second light chain of the full-length anti-PD-L1 antibody.A multispecific targeted conjugate is provided comprising (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the first and second anti-Trop-2 sdAb (e.g., VHH) independently comprises: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, the following polypeptide chains are provided: (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of the full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, and a first anti-Trop-2 sdAb (e.g., VHH); (b) a second polypeptide chain comprising, from the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 3, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 4, and a second anti-Trop-2 sdAb (e.g., VHH); and (c) a third polypeptide chain comprising, from the N-terminus to the C-terminus, a first light chain of the full-length anti-PD-L1 antibody;A multispecific targeting conjugate is provided comprising: (d) a fourth polypeptide chain comprising a second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the full-length anti-PD-L1 antibody is provided comprising i) the amino acid sequence of Sequence ID No. 3 H-CDR1 containing the amino acid sequence of SEQ ID NO: 8, H-CDR2 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249, and the first and second anti-Trop-2 sdAb (e.g., VHH) independently includes: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206;or v) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, the full-length anti-PD-L1 antibody comprises i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and SEQ ID NO: 221 VL containing the amino acid sequence of ;vi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 213;vii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 222;viii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 221;ix) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 222;x) VH containing the amino acid sequence of SEQ ID NO: 223 and the amino acid sequence of SEQ ID NO: 222 VL containing the amino acid sequence of SEQ ID NO: 16; xii) VH containing the amino acid sequence of SEQ ID NO: 224; xiii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 225; xiv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 224; xvi) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 226; xvii) VH containing the amino acid sequence of SEQ ID NO: 227 and VL containing the amino acid sequence of SEQ ID NO: 226; xviii) VH containing the amino acid sequence of SEQ ID NO: 228 and VL containing the amino acid sequence of SEQ ID NO: 226; xix) VH containing the amino acid sequence of SEQ ID NO: 229 and VL containing the amino acid sequence of SEQ ID NO: 226; xx) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 226;xxi) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 220; xxii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 226; or xxiii) VH containing the amino acid sequence of SEQ ID NO: 250 and VL containing the amino acid sequence of SEQ ID NO: 251. In some embodiments, the full-length anti-PD-L1 antibody comprises i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) SEQ ID NO: 81; iv) Heavy chain containing the amino acid sequence of SEQ ID NO: 82 and light chain containing the amino acid sequence of SEQ ID NO: 83 and light chain containing the amino acid sequence of SEQ ID NO: 84; v) Heavy chain containing the amino acid sequence of SEQ ID NO: 85 and light chain containing the amino acid sequence of SEQ ID NO: 86; vi) Heavy chain containing the amino acid sequence of SEQ ID NO: 87 and light chain containing the amino acid sequence of SEQ ID NO: 88; vii) Heavy chain containing the amino acid sequence of SEQ ID NO: 91 and light chain containing the amino acid sequence of SEQ ID NO: 92; viiii) Heavy chain containing the amino acid sequence of SEQ ID NO: 93 and light chain containing the amino acid sequence of SEQ ID NO: 94 Light chain;ix) Heavy chain containing the amino acid sequence of SEQ ID NO: 95 and light chain containing the amino acid sequence of SEQ ID NO: 96;x) Heavy chain containing the amino acid sequence of SEQ ID NO: 97 and light chain containing the amino acid sequence of SEQ ID NO: 98;xi) Heavy chain containing the amino acid sequence of SEQ ID NO: 99 and light chain containing the amino acid sequence of SEQ ID NO: 100;xii) Heavy chain containing the amino acid sequence of SEQ ID NO: 101 and light chain containing the amino acid sequence of SEQ ID NO: 102;xiii) Heavy chain containing the amino acid sequence of SEQ ID NO: 103 and light chain containing the amino acid sequence of SEQ ID NO: 104;xiv) Including the amino acid sequence of SEQ ID NO: 105 xv) Heavy chain and light chain containing the amino acid sequence of SEQ ID NO: 106; xvi) Heavy chain containing the amino acid sequence of SEQ ID NO: 107 and light chain containing the amino acid sequence of SEQ ID NO: 108; xvii) Heavy chain containing the amino acid sequence of SEQ ID NO: 109 and light chain containing the amino acid sequence of SEQ ID NO: 110; xviii) Heavy chain containing the amino acid sequence of SEQ ID NO: 113 and light chain containing the amino acid sequence of SEQ ID NO: 114; xix) Heavy chain containing the amino acid sequence of SEQ ID NO: 115 and light chain containing the amino acid sequence of SEQ ID NO: 116 A light chain containing an amino acid sequence;xx) A heavy chain containing the amino acid sequence of SEQ ID NO. 117 and a light chain containing the amino acid sequence of SEQ ID NO. 118;xxi) A heavy chain containing the amino acid sequence of SEQ ID NO. 119 and a light chain containing the amino acid sequence of SEQ ID NO. 120;xxii) A heavy chain containing the amino acid sequence of SEQ ID NO. 121 and a light chain containing the amino acid sequence of SEQ ID NO. 122;xxiii) A heavy chain containing the amino acid sequence of SEQ ID NO. 17 and a light chain containing the amino acid sequence of SEQ ID NO. 34; or xxiv) A heavy chain containing the amino acid sequence of SEQ ID NO. 241 and a light chain containing the amino acid sequence of SEQ ID NO. 242.In some embodiments, the first and second anti-Trop-2 sdAb (e.g., VHH) independently comprise any of the amino acid sequences of SEQ ID NOs. 50, 66, 123-136, and 203. In some embodiments, the first and second anti-Trop-2 sdAb bind to different Trop-2 epitopes. One or more arbitrary linkers may be the same or different (or absent), and may be, for example, any of SEQ ID NOs. 144, 204, 207-211, and 252, or any of SEQ ID NOs. 207-209. The first and second anti-Trop-2 sdAb (e.g., VHH) may be the same or different and may bind to the same or different Trop-2 epitopes. The first and second conjugation sites may be the same or different, for example, any of SEQ ID NOs: 145-196, for example, SEQ ID NOs: 147 or 188. The first and second effector molecules may be the same or different, for example, a therapeutic agent or an oligonucleotide. The first and second effector molecule linkers may be the same or different. In some embodiments, the multispecific targeting conjugate contains a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the linker-effector molecule conjugate contains any of formulas A-J, for example, the structure of formula A.

[0125] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 231; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 231; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises structures of formulas A to J, for example, formula A.

[0126] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 232; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 232; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0127] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 238; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 238; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 90; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 90; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0128] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 239; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 239; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 90; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 90; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0129] In some embodiments, (a) from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, and a first anti-Trop-2 (b) A first polypeptide chain containing sdAb (e.g., VHH); (b) From the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 4, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 5, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 6, and a second anti-Trop-2 A multispecific targeting conjugate is provided, comprising: (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing a first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing a second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, (a) from the N-terminus to the C-terminus, a first heavy chain of the full-length anti-PD-L1 antibody, an optional linker 1, and a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71;(b) A first polypeptide chain comprising a uPA cleavage site (e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, and a first anti-Trop-2 sdAb (e.g., VHH); (b) From the N-terminus to the C-terminus, a second heavy chain of the anti-PD-L1 full-length antibody, an optional linker 4, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 5, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 6, and a second anti-Trop-2 (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule linker (e.g., (Gly)6-amide-PEG8-V A multispecific targeted conjugate is provided, comprising a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via A-PAB), wherein the full-length anti-PD-L1 antibody comprises i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30;or ii) comprising H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249. In some embodiments, (a) from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, and a first anti-Trop-2 (b) A first polypeptide chain containing sdAb (e.g., VHH); (b) From the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 4, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 5, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 6, and a second anti-Trop-2 (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB);(f) a multispecific targeted conjugate is provided comprising a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the first and second anti-Trop-2 sdAb (e.g., VHH) independently comprises: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, (a) from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, and a first anti-Trop-2 (b) A first polypeptide chain comprising an sdAb (e.g., VHH); (b) A second polypeptide chain comprising, from the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 4, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 5, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 6, and a second anti-Trop-2 sdAb (e.g., VHH);A multispecific targeted conjugate is provided, comprising: (c) a third polypeptide chain comprising the first light chain of the full-length anti-PD-L1 antibody; (d) a fourth polypeptide chain comprising the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the full-length anti-PD-L1 antibody is also provided. The antibodies include i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249, and the first and second anti-Trop-2 sdAb (e.g., VHH) independently includes: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206;or v) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, the full-length anti-PD-L1 antibody comprises i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 213; v ii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 222; viii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 222; x) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 224; xii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 225; xiii) containing the amino acid sequence of SEQ ID NO: 16; VL;xiv) containing the amino acid sequence of VH and SEQ ID NO: 226; VL;xv) containing the amino acid sequence of VH and SEQ ID NO: 225; VL;xvi) containing the amino acid sequence of VH and SEQ ID NO: 224; VL;xvii) containing the amino acid sequence of VH and SEQ ID NO: 226; VL;xviii) containing the amino acid sequence of VH and SEQ ID NO: 228 VL containing the amino acid sequence; xix) VH containing the amino acid sequence of SEQ ID NO. 229 and VL containing the amino acid sequence of SEQ ID NO. 226; xx) VH containing the amino acid sequence of SEQ ID NO. 230 and VL containing the amino acid sequence of SEQ ID NO. 226; xxi) VH containing the amino acid sequence of SEQ ID NO. 230 and VL containing the amino acid sequence of SEQ ID NO. 220; xxii) VH containing the amino acid sequence of SEQ ID NO. 219 and VL containing the amino acid sequence of SEQ ID NO. 226; or xxiii) VH containing the amino acid sequence of SEQ ID NO. 250 and VL containing the amino acid sequence of SEQ ID NO. 251. In some embodiments, the full-length anti-PD-L1 antibody is: i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) a heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) a heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) an amino acid sequence of SEQ ID NO: 87 vii) Heavy chain containing amino acid sequence and light chain containing amino acid sequence of SEQ ID NO: 88; vii) Heavy chain containing amino acid sequence of SEQ ID NO: 91 and light chain containing amino acid sequence of SEQ ID NO: 92; viii) Heavy chain containing amino acid sequence of SEQ ID NO: 93 and light chain containing amino acid sequence of SEQ ID NO: 94; ix) Heavy chain containing amino acid sequence of SEQ ID NO: 95 and light chain containing amino acid sequence of SEQ ID NO: 96; x) Heavy chain containing amino acid sequence of SEQ ID NO: 97 and light chain containing amino acid sequence of SEQ ID NO: 98; xi) Heavy chain containing amino acid sequence of SEQ ID NO: 99 and light chain containing amino acid sequence of SEQ ID NO: 100;xii) A heavy chain containing the amino acid sequence of SEQ ID NO: 101 and a light chain containing the amino acid sequence of SEQ ID NO: 102; xiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 103 and a light chain containing the amino acid sequence of SEQ ID NO: 104; xiv) A heavy chain containing the amino acid sequence of SEQ ID NO: 105 and a light chain containing the amino acid sequence of SEQ ID NO: 106; xv) A heavy chain containing the amino acid sequence of SEQ ID NO: 107 and a light chain containing the amino acid sequence of SEQ ID NO: 108; xvi) A heavy chain containing the amino acid sequence of SEQ ID NO: 109 and a light chain containing the amino acid sequence of SEQ ID NO: 110; xvii) A heavy chain containing the amino acid sequence of SEQ ID NO: 111 and a light chain containing the amino acid sequence of SEQ ID NO: 112; xviii) Heavy chain containing the amino acid sequence of SEQ ID NO: 113 and light chain containing the amino acid sequence of SEQ ID NO: 114; xix) Heavy chain containing the amino acid sequence of SEQ ID NO: 115 and light chain containing the amino acid sequence of SEQ ID NO: 116; xx) Heavy chain containing the amino acid sequence of SEQ ID NO: 117 and light chain containing the amino acid sequence of SEQ ID NO: 118; xxi) Heavy chain containing the amino acid sequence of SEQ ID NO: 119 and light chain containing the amino acid sequence of SEQ ID NO: 120; xxii) Heavy chain containing the amino acid sequence of SEQ ID NO: 121 and light chain containing the amino acid sequence of SEQ ID NO: 122; xxiii) Heavy chain containing the amino acid sequence of SEQ ID NO: 17 and light chain containing the amino acid sequence of SEQ ID NO: 34;Or xxiv) comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 241 and a light chain containing the amino acid sequence of SEQ ID NO: 242. In some embodiments, the first and second anti-Trop-2 sdAb (e.g., VHH) independently contain any of the amino acid sequences of SEQ ID NOs: 50, 66, 123-136, and 203. In some embodiments, the first and second anti-Trop-2 sdAb bind to different Trop-2 epitopes. One or more arbitrary linkers may be the same or different (or absent), for example, any of SEQ ID NOs: 144, 204, 207-211, and 252, or any of SEQ ID NOs: 207-209. The first and second anti-Trop-2 sdAb (e.g., VHH) may be the same or different and may bind to the same or different Trop-2 epitopes. In some embodiments, cleavage is induced by conditions at the target site. In some embodiments, the conditions at the target site are selected from the group consisting of proteases, pH changes, redox changes, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof. In some embodiments, the target site is a disease site such as a tumor (e.g., a solid tumor). In some embodiments, the conditions are the tumor microenvironment. The first and second cleavage sites may be the same or different, for example, any of SEQ ID NOs: 71, 137-143, and 214. The first and second conjugation sites may be the same or different, for example, any of SEQ ID NOs: 145-196, for example, SEQ ID NOs: 147 or 188. The first and second effector molecules may be the same or different, for example, a therapeutic agent or an oligonucleotide. The first and second effector molecule linkers may be the same or different. In some embodiments, the multispecific targeting conjugate contains a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the linker-effector molecule conjugate (L-(D); a) contains any of formulas A to J, for example, the structure of formula A.

[0130] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 233; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 233; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0131] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 234; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 234; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0132] In some embodiments, (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 3, and a second anti-Trop-2 sdAb (e.g., VHH); (b) a second heavy chain of the same full-length anti-PD-L1 antibody, an optional linker 4, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 5, a third anti-Trop-2 sdAb (e.g., VHH), an optional linker 6, and a fourth anti-Trop-2 A multispecific targeting conjugate is provided, comprising: (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing a first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing a second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 3, and a second anti-Trop-2 sdAb (e.g., VHH);(b) From the N-terminus to the C-terminus, the second heavy chain of the anti-PD-L1 full-length antibody, an optional linker 4, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 5, a third anti-Trop-2 sdAb (e.g., VHH), an optional linker 6, and a fourth anti-Trop-2 (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a multispecific targeted conjugate containing a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). A total-length anti-PD-L1 antibody is provided, wherein the antibody comprises i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249. In some embodiments, (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 3, and a second anti-Trop-2 sdAb (e.g., VHH);(b) From the N-terminus to the C-terminus, the second heavy chain of the anti-PD-L1 full-length antibody, an optional linker 4, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 5, a third anti-Trop-2 sdAb (e.g., VHH), an optional linker 6, and a fourth anti-Trop-2 A multispecific targeted conjugate is provided, comprising: (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the first, second, third, and fourth anti-Trop-2 sdAb (e.g., VHH) independently includes: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206;Alternatively, v) comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 2, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 3, and a second anti-Trop-2 sdAb (e.g., VHH); (b) a second heavy chain of the same full-length anti-PD-L1 antibody, an optional linker 4, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 5, a third anti-Trop-2 sdAb (e.g., VHH), an optional linker 6, and a fourth anti-Trop-2 (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule linker (e.g., (Gly)6-amide-PEG8-V A multispecific targeted conjugate is provided, comprising a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via A-PAB), wherein the full-length anti-PD-L1 antibody comprises i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30;or ii) comprising H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249, wherein the first, second, third, and fourth anti-Trop-2 sdAb (e.g., VHH) independently comprises: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, the full-length anti-PD-L1 antibody is: i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 222; viii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 222;x) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 224; xii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 225; xiii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 226; xiv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 225; xv) Amino acid sequence of SEQ ID NO: 223 VH containing the amino acid sequence and VL containing the amino acid sequence of SEQ ID NO 224; xvi) VH containing the amino acid sequence of SEQ ID NO 223 and VL containing the amino acid sequence of SEQ ID NO 226; xvii) VH containing the amino acid sequence of SEQ ID NO 227 and VL containing the amino acid sequence of SEQ ID NO 226; xviii) VH containing the amino acid sequence of SEQ ID NO 228 and VL containing the amino acid sequence of SEQ ID NO 226; xix) VH containing the amino acid sequence of SEQ ID NO 229 and VL containing the amino acid sequence of SEQ ID NO 226; xx) containing the amino acid sequence of SEQ ID NO 230;

[0133] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 235; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 235; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0134] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 236; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 236; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0135] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 237; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 237; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 242; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0136] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 67; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 67; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 68; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 68; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0137] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 215; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 215; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 68; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 68; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0138] In some embodiments, (a) from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 4, and a second anti-Trop-2 (b) A first polypeptide chain containing an sdAb (e.g., VHH); (b) From the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 5, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 6, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 7, a third anti-Trop-2 sdAb (e.g., VHH), an optional linker 8, and a fourth anti-Trop-2 A multispecific targeting conjugate is provided, comprising: (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing a first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing a second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, (a) from the N-terminus to the C-terminus, a first heavy chain of the full-length anti-PD-L1 antibody, an optional linker 1, and a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71;(b) A first polypeptide chain comprising a uPA cleavage site (e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 4, and a second anti-Trop-2 sdAb (e.g., VHH); (b) From the N-terminus to the C-terminus, a second heavy chain of the anti-PD-L1 full-length antibody, an optional linker 5, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 6, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 7, and a third anti-Trop-2 (c) a second polypeptide chain comprising an sdAb (e.g., VHH), an optional linker 8, and a fourth anti-Trop-2 sdAb (e.g., VHH); (d) a third polypeptide chain comprising the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain comprising the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA A multispecific targeted conjugate is provided, comprising a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via -PAB), wherein the full-length anti-PD-L1 antibody comprises i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30;or ii) comprising H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249. In some embodiments, (a) from the N-terminus to the C-terminus, a first heavy chain of a full-length anti-PD-L1 antibody, an optional linker 1, a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 4, and a second anti-Trop-2 (b) A first polypeptide chain containing an sdAb (e.g., VHH); (b) From the N-terminus to the C-terminus, a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 5, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 6, a second conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 7, a third anti-Trop-2 sdAb (e.g., VHH), an optional linker 8, and a fourth anti-Trop-2 (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing the second light chain of the full-length anti-PD-L1 antibody; (c) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB);(f) a multispecific targeted conjugate is provided comprising a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the first, second, third, and fourth anti-Trop-2 sdAb (e.g., VHH) independently comprises: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, (a) a first polypeptide chain comprising, from the N-terminus to the C-terminus, a first heavy chain of the full-length anti-PD-L1 antibody, an optional linker 1, a first cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71; or a uPA cleavage site, e.g., SEQ ID NO: 214), an optional linker 2, a first conjugation site (e.g., a transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), an optional linker 3, a first anti-Trop-2 sdAb (e.g., VHH), an optional linker 4, and a second anti-Trop-2 sdAb (e.g., VHH); (b) a second heavy chain of the full-length anti-PD-L1 antibody, an optional linker 5, a second cleavage site (e.g., an MMP cleavage site, e.g., an MMP-9 cleavage site, e.g., SEQ ID NO: 71);or uPA cleavage site (e.g., SEQ ID NO: 214), any linker 6, second conjugation site (e.g., transglutaminase conjugation site, e.g., SEQ ID NO: 147 or 188), any linker 7, third anti-Trop-2 sdAb (e.g., VHH), any linker 8, and fourth anti-Trop-2 A multispecific targeting conjugate is provided, comprising: (c) a second polypeptide chain containing sdAb (e.g., VHH); (d) a third polypeptide chain containing the first light chain of the full-length anti-PD-L1 antibody; (e) a fourth polypeptide chain containing the second light chain of the full-length anti-PD-L1 antibody; (e) a first effector molecule (e.g., exatecan) conjugated to the first conjugation site via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to the second conjugation site via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB), wherein the The anti-PD-L1 full-length antibody comprises i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3, H-CDR2 containing the amino acid sequence of SEQ ID NO: 8, H-CDR3 containing the amino acid sequence of SEQ ID NO: 13, L-CDR1 containing the amino acid sequence of SEQ ID NO: 20, L-CDR2 containing the amino acid sequence of SEQ ID NO: 25, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30; or ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 244, H-CDR2 containing the amino acid sequence of SEQ ID NO: 245, H-CDR3 containing the amino acid sequence of SEQ ID NO: 246, L-CDR1 containing the amino acid sequence of SEQ ID NO: 247, L-CDR2 containing the amino acid sequence of SEQ ID NO: 248, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 249, and the first, second, third, and fourth anti-Trop-2 sdAb (e.g., VHH) independently comprises: i) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 42, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 63;iii) CDR1 containing the amino acid sequence of SEQ ID NO: 37, CDR2 containing the amino acid sequence of SEQ ID NO: 205, and CDR3 containing the amino acid sequence of SEQ ID NO: 47; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 53, CDR2 containing the amino acid sequence of SEQ ID NO: 58, and CDR3 containing the amino acid sequence of SEQ ID NO: 206; or v) CDR1 containing the amino acid sequence of SEQ ID NO: 216, CDR2 containing the amino acid sequence of SEQ ID NO: 217, and CDR3 containing the amino acid sequence of SEQ ID NO: 218. In some embodiments, the full-length anti-PD-L1 antibody is: i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16;

[0139] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 240; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 240; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 90; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 90; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0140] In some embodiments, a multispecific targeted conjugate is provided comprising: (a) a first polypeptide chain comprising the amino acid sequence of SEQ ID NO: 69; (b) a second polypeptide chain comprising the amino acid sequence of SEQ ID NO: 69; (c) a third polypeptide chain comprising the amino acid sequence of SEQ ID NO: 70; (d) a fourth polypeptide chain comprising the amino acid sequence of SEQ ID NO: 70; (e) a first effector molecule (e.g., exatecan) conjugated to a first conjugation site in the first polypeptide chain via a first effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB); and (f) a second effector molecule (e.g., exatecan) conjugated to a second conjugation site in the second polypeptide chain via a second effector molecule linker (e.g., (Gly)6-amide-PEG8-VA-PAB). In some embodiments, the multispecific targeting conjugate comprises a total of about 1 to about 20 effector molecules (e.g., exatecan), for example, about 2 to about 20 or about 4 to about 6 effector molecules. In some embodiments, the effector molecules are therapeutic agents or oligonucleotides. In some embodiments, the linker-effector molecule conjugate comprises any of formulas A to J, for example, the structure of formula A.

[0141] In some embodiments, a targeted conjugate is provided that includes a targeting moiety that specifically recognizes PD-L1, where the PD-L1 targeting moiety is a full-length anti-PD-L1 antibody comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO: 34. In some embodiments, the targeted conjugate further comprises a linker-effector molecule conjugate of formula A conjugated to the PD-L1 targeting moiety via a transglutaminase conjugation site (e.g., any of SEQ ID NOs: 145-196, e.g., SEQ ID NOs: 147 or 188). In some embodiments, the targeted conjugate further comprises one or more protease cleavage sites, e.g., any of SEQ ID NOs: 71, 137-143, and 214. In some embodiments, the protease cleavage site is located between the PD-L1 targeting moiety and the transglutaminase conjugation site. In some embodiments, the protease cleavage site is not located between the PD-L1 targeting site and the transglutaminase conjugation site.

[0142] In some embodiments, a targeted conjugate is provided that includes a targeting moiety that specifically recognizes Trop-2, wherein the Trop-2 targeted moiety independently includes one or more anti-Trop-2 VHH molecules containing any of the amino acid sequences of SEQ ID NOs. 50, 66, and 203. In some embodiments, the targeted conjugate further includes a linker-effector molecule conjugate of formula A conjugated to the Trop-2 targeted moiety via a transglutaminase conjugation site (e.g., any of SEQ ID NOs. 145-196, e.g., SEQ ID NOs. 147 or 188). In some embodiments, the targeted conjugate further includes one or more protease cleavage sites, e.g., any of SEQ ID NOs. 71, 137-143, and 214. In some embodiments, the protease cleavage site is located between the Trop-2 targeted moiety and the transglutaminase conjugation site. In some embodiments, the protease cleavage site is not located between the Trop-2 targeting site and the transglutaminase conjugation site.

[0143] In some embodiments, the conjugation site (C) of the multispecific targeting conjugate may include any preferred number of glutamine-containing tags fused in series with each other. In some embodiments, the number of glutamine-containing tags fused in series with each other may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. In some embodiments, the multispecific targeting conjugate may include any preferred number of effector molecules (D) and effector molecule linkers (L). In some embodiments, the number of effector molecules (D) may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20. In some embodiments, the number of linkers L may be 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.

[0144] The multispecific targeted conjugates described herein may include any of the targeted peptides, antibodies or their antigen-binding fragments, isolated antibody constructs, effector molecules, conjugation sites, cleavage sites (e.g., protease cleavage sites), peptide linkers, and effector molecule linkers described in Parts II-IV and Sections A-G below.

[0145] In some embodiments, the multispecific targeting conjugate contains a single effector molecule. In some embodiments, the multispecific targeting conjugate contains multiple effector molecules. In some embodiments, the multispecific targeting conjugate contains a single effector molecule. In some embodiments, the multispecific targeting conjugate contains two or more identical effector molecules. In some embodiments, the multispecific targeting conjugate contains two or more different effector molecules. In some embodiments, the multispecific targeting conjugate contains a single copy of each effector molecule. In some embodiments, the multispecific targeting conjugate contains two or more copies of each effector molecule.

[0146] In some embodiments, multispecific targeted conjugates exhibit a high drug load. The term "drug load" refers to the ratio between the number of effector molecules and the number of targeted portions (e.g., antibodies or their antigen-binding fragments) in a multispecific targeted conjugate. For example, an antibody conjugated to a total of eight effector molecules has a drug load of 8. Each molecule in a multispecific targeted conjugate has an integer drug load. However, in a composition, multispecific targeted conjugates of different molecules may have different drug load values. Therefore, a composition may have an average drug load of an integer or non-integer value.

[0147] In some embodiments, the multispecific targeting conjugate has an effector molecule to targeting portion (e.g., a first targeting portion and / or a second targeting portion) ratio of at least about 1:1, 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 11:1, 12:1, 13:1, 14:1, 15:1, 16:1, 17:1, 17:1, 18:1, 19:1, or 20:1. In some embodiments, the multispecific targeting conjugate has an effector molecule to targeting portion (e.g., a first targeting portion and / or a second targeting portion) ratio of approximately 20:1 or less, 19:1 or less, 18:1 or less, 17:1 or less, 16:1 or less, 15:1 or less, 14:1 or less, 13:1 or less, 12:1 or less, 11:1 or less, 10:1 or less, 9:1 or less, 8:1 or less, 7:1 or less, 6:1 or less, 5:1 or less, 4:1 or less, 3:1 or less, 2:1 or less, or 1:1 or less. In some embodiments, the multispecific targeting conjugate has an effector molecule to targeting moiety (e.g., a first targeting moiety and / or a second targeting moiety) ratio of approximately 1:1-2:1, 2:1-4:1, 4:1-6:1, 4:1-8:1, 1:1-10:1, 2:1-10:1, 1:1-16:1, 4:1-20:1, 10:1-20:1, 1:1-20:1, or 2:1-20:1.

[0148] In some embodiments, the drug load of the multispecifically targeted conjugate is at least about 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 11:1, 12:1, 13:1, 14:1, 15:1, 16:1, 17:1, 17:1, 18:1, 19:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1, 60:1, 70:1, 80:1, 90:1, 100:1, or more. In some embodiments, the drug load of the multispecifically targeted conjugate is one of the following: approximately 100:1 or less, 90:1 or less, 80:1 or less, 70:1 or less, 60:1 or less, 50:1 or less, 40:1 or less, 30:1 or less, 20:1 or less, 19:1 or less, 18:1 or less, 17:1 or less, 16:1 or less, 15:1 or less, 14:1 or less, 13:1 or less, 12:1 or less, 11:1 or less, 10:1 or less, 9:1 or less, 8:1 or less, 7:1 or less, 6:1 or less, 5:1 or less, 4:1 or less, 3:1 or less, or 2:1 or less. In some embodiments, the drug load of the multispecifically targeted conjugate is one of the following: approximately 2:1–4:1, 2:1–8:1, 2:1–10:1, 2:1–16:1, 4:1–20:1, 10:1–20:1, 20:1–40:1, 40:1–100:1, 2:1–20:1, 2:1–40:1, or 10:1–40:1.

[0149] Furthermore, the multispecific targeting portion of any of the multispecific targeting conjugates described herein (for example, a multispecific targeting conjugate that does not contain one or more effector molecules) is also provided. In some embodiments, i) first and second polypeptide chains each containing the amino acid sequence of SEQ ID NO: 231, and third and fourth polypeptide chains each containing the amino acid sequence of SEQ ID NO: 242; ii) first and second polypeptide chains each containing the amino acid sequence of SEQ ID NO: 232, and third and fourth polypeptide chains each containing the amino acid sequence of SEQ ID NO: 242; iii) first and second polypeptide chains each containing the amino acid sequence of SEQ ID NO: 233, and third and fourth polypeptide chains each containing the amino acid sequence of SEQ ID NO: 242; iv) first and second polypeptide chains each containing the amino acid sequence of SEQ ID NO: 234, and third and fourth polypeptide chains each containing the amino acid sequence of SEQ ID NO: 242; v) first and second polypeptide chains each containing the amino acid sequence of SEQ ID NO: 235, and third and fourth polypeptide chains each containing the amino acid sequence of SEQ ID NO: 242; vi) first and second polypeptide chains each containing the amino acid sequence of SEQ ID NO: 236 vii) Peptide chains, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 242; vii) First and second polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 237, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 242; viii) First and second polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 238, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 90; ix) First and second polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 239, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 90; x) First and second polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 67, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 68; xi) First and second polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 215, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 68;xii) A multispecific targeting moiety is provided comprising first and second polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 240, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 90; or xiii) a first and second polypeptide chain, each containing the amino acid sequence of SEQ ID NO: 69, and third and fourth polypeptide chains, each containing the amino acid sequence of SEQ ID NO: 90.

[0150] Also provided are isolated nucleic acids encoding one or more polypeptide chains of any of the multispecific targeting conjugates described herein, for example, isolated nucleic acids encoding any of the multispecific targeting moieties described herein. Also provided are vectors containing such isolated nucleic acids, and host cells containing such isolated nucleic acids or vectors.

[0151] A. First targeting region that specifically recognizes PD-L1 The PD-L1 / Trop-2 multispecific targeting conjugates described herein include a first targeting moiety that specifically recognizes PD-L1. In some embodiments, the first targeting moiety includes (or consists of, or is essentially composed of) one or more PD-L1 targeting peptides, or one or more (e.g., 1, 2, 3, 4, or 5, e.g., 1) anti-PD-L1 antibodies or their antigen-binding fragments. In some embodiments, the first targeting moiety includes one or more (e.g., 1, 2, 3, 4, or 5, e.g., 1) anti-PD-L1 antibodies or their antigen-binding fragments. In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment is selected from the group consisting of full-length antibodies, diabodies, scFv, scFab, Fab, Fab', F(ab')2, sdAb, dsFv, and combinations thereof. The anti-PD-L1 antibody or its antigen-binding fragment may be human, humanized, mouse, camelid, or chimeric. In some embodiments, the first targeting moiety is monospecific. In some embodiments, the first targeting moiety is multispecific (e.g., recognizes two or more different PD-L1 epitopes). In some embodiments, the first targeting moiety is monovalent. In some embodiments, the first targeting moiety is polyvalent (e.g., contains two or more anti-PD-L1 antigen-binding fragments, or a bivalent full-length anti-PD-L1 antibody or F(ab')2). Either the anti-PD-L1 targeting moiety or isolated anti-PD-L1 antibody construct described herein (see, for example, below and Section III) may be used as the first targeting moiety in the PD-L1 / Trop-2 multispecific targeting conjugate described herein. In some embodiments, the first targeting moiety is a full-length anti-PD-L1 antibody. Targeted peptides

[0152] In some embodiments, the first targeting moiety or anti-PD-L1 targeting moiety comprises (or consists of, or is essentially composed of) one or more targeted peptides that specifically bind to PD-L1.

[0153] The term "targeted peptide" refers to a non-antibody-based polypeptide that specifically binds to a target molecule, such as a cell surface molecule at a target site. In some embodiments, the targeting moiety (e.g., an anti-PD-L1 targeting moiety or an anti-Trop-2 targeting moiety) is a fusion protein comprising an antibody or its antigen-binding fragment fused to a targeted peptide that binds to a target molecule (e.g., PD-L1 or Trop-2).

[0154] In some embodiments, the targeted polypeptide includes a non-antibody scaffold. A non-antibody scaffold is a genetically engineered protein scaffold that produces specificity for different types of targets. Compared to antibodies, genetically engineered protein scaffolds have a much smaller size and simpler structure, which facilitates recombinant gene expression, construction of bifunctional fusion proteins, and tissue penetration. See, for example, Skerra, Curr. Opin. Biotech. 2007, 18:298-304. More than 50 different non-antibody scaffolds have been reported. See, for example, Vazquez-Lombardi, Rodrigo, et al. Drug discovery today 20.10(2015):1271-1283. Examples of non-antibody scaffolds include, but are not limited to, lipocalin, anticalin (an artificial antibody-mimetic protein derived from human lipocalin), "T-bodies," peptides (e.g., BICYCLE® peptides), affibodies (antibody mimetics composed of α-helices, e.g., 3-helix bundles), peptidebodies (peptide-Fc fusions), DARPin (designed ankyrin repeat protein (a genetically engineered antibody-mimetic protein consisting of repeating motifs)), affimers, avimers, notchin (a protein structural motif containing three disulfide crosslinks), monobodies, affinity clamps, external domains, receptor external domains, receptors, cytokines, ligands, immune cytokines, and centirin. See, for example, WO2019084060 incorporated herein by reference.

[0155] In some embodiments, the targeting moiety includes antikalin. Antikalin is one of the more ambitiously developed non-antibody scaffolds, and numerous lead compounds targeted against CTLA-4, hepcidin, hepatocyte growth factor receptor (HGFR; MET), IL-4Ra, and IL-23 / IL-17 are in preclinical trials. PRS-050 (ANGIOCAL®; Pieris) is a VEGF-A-targeted anti-angiogenic antikalin currently in Phase I clinical trials. MP0112 (Molecular Partners / Allergan), a VEGF-A-targeted DARPin targeting retinal neovascularization, is also currently being evaluated in clinical trials. FDA-approved non-antibody monoscaffolds include the plasma kallikrein inhibitors Kunitz domain KALBITOR® (ecalantide; Dyax), BERINERT® (CSL-Behring), and CINRYZE® (ViroPharma / Shire), used in the treatment of hereditary angioedema, as well as the bradykinin receptor antagonist FIRAZYR® (Shire). See, for example, Vazquez-Lombardi, Rodrigo, et al. Drug discovery today 20.10 (2015):1271-1283.

[0156] In some embodiments, the targeted peptide comprises a polypeptide derived from a ligand of a receptor or target molecule (e.g., PD-L1 or Trop-2). In some embodiments, the targeted polypeptide is an inhibitory polypeptide that completely or partially (e.g., at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90%, or 95%) blocks the binding of the target molecule and its ligand or receptor. In some embodiments, the targeted peptide is at least about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 100, 150, 200, 250, 300, 350, 400, 450, 500 amino acids, or any of these lengths.

[0157] In some embodiments, the targeted peptide includes a stabilizing domain. The stabilizing domain may be any suitable domain that stabilizes the targeted peptide. In some embodiments, the stabilizing domain extends the half-life of the targeted peptide in vivo. In some embodiments, the stabilizing domain is an Fc domain, e.g., one of the Fc domains described in the “Antibody or its Antigen-Binding Fragment” section. In some embodiments, the stabilizing domain is an albumin domain. In some embodiments, the targeted peptide and the stabilizing domain are fused to each other via a linker, e.g., a peptide linker.

[0158] Peptide linkers may have naturally occurring or non-natural sequences. For example, a sequence derived from the hinge region of a heavy-chain-only antibody may be used as a linker. Peptide linkers may be of any suitable length. In some embodiments, peptide linkers do not take on a rigid three-dimensional structure and tend to provide flexibility to the polypeptide. In some embodiments, the peptide linker is a mobile linker. Exemplary mobile linkers include glycine polymers, glycine-serine polymers, glycine-alanine polymers, alanine-serine polymers, and other mobile linkers known in the art. In some embodiments, the peptide linker includes a substrate sequence for an enzymatic reaction. In some embodiments, the peptide linker includes a substrate sequence for an enzyme that links the targeted peptide and the stabilizing domain.

[0159] Targeted peptides can be obtained using methods known in the art, for example, by screening a library of polypeptides. Polypeptides can be prepared using chemical synthesis or generated using recombinant DNA technology. Nucleic acid constructs, vectors, and host cells for preparation that encode any of the targeted peptides described herein are also provided.

[0160] Antibodies or their antigen-binding fragments In some embodiments, the first targeting moiety or anti-PD-L1 targeting moiety includes a variant of an anti-PD-L1 antibody or its antigen-binding fragment known in the art, or a variant of any of the anti-PD-L1 antibodies or their antigen-binding fragments described herein. For example, a variant of the antibody or its antigen-binding fragment may include one or more modifications or mutations of the amino acid sequence of the exemplary antibody ("parent antibody") while retaining the overall molecular structure of the parent antibody amino acid sequence. Any region of the parent antibody or its antigen-binding fragment, e.g., one or more framework regions, one or more CDR regions, one or more constant regions, or any combination thereof, may have its amino acid sequence modified. Types of modifications or mutations include substitutions, insertions, deletions, or combinations thereof of one or more amino acids in the parent antibody. In some embodiments, a variant of the anti-PD-L1 antibody or its antigen-binding fragment comprises a CDR (e.g., H-CDR1, H-CDR2, H-CDR3, L-CDR1, L-CDR2, and / or L-CDR3 of sdAb; or CDR1, CDR2, and / or CDR3) having at least about 80% (e.g., at least about 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99%) sequence identity to the corresponding CDR of the parent antibody. In some embodiments, a variant of the anti-PD-L1 antibody or its antigen-binding fragment comprises one or more CDRs having one, two, or three amino acid mutations (e.g., substitutions, e.g., conserved substitutions) compared to the corresponding CDR(s) of the parent antibody. In some embodiments, the variant of the anti-PD-L1 antibody or its antigen-binding fragment includes a VH having an amino acid sequence identical to that of the VH of the parent antibody by at least about 80% (e.g., at least about 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99%). In some embodiments, the variant of the anti-PD-L1 antibody or its antigen-binding fragment includes a VL having an amino acid sequence identical to that of the VL of the parent antibody by at least about 80% (e.g., at least about 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99%).In some embodiments, a variant of an anti-PD-L1 antibody or an antigen-binding fragment thereof comprises an sdAb (e.g., VHH) having an amino acid sequence that is at least about 80% (e.g., any one of at least about 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99%) identical to the sdAb (e.g., VHH) of the parent antibody. In some particular embodiments, a variant of an anti-PD-L1 antibody or an antigen-binding fragment thereof comprises one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, or fifteen conservative or non-conservative substitutions, and / or one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, or fifteen additions and / or deletions to the amino acid sequence shown in any of the VH, VL, heavy chain, and / or light chain (or sdAb) of the parent antibody.

[0161] The anti-PD-L1 antibody or its antigen-binding fragment may be (or may be derived from) any anti-PD-L1 antibody or its antigen-binding fragment known in the art. Exemplary anti-PD-L1 antibodies include, but are not limited to, atezolizumab (e.g., Tecentriq®), avelumab (e.g., Bavencio®), durvalumab (e.g., Imfinzi®), BGB-A333, SHR-1316 (HTI-1088), CK-301, BMS-936559, and emvafolimab (KN035, ASC). 22), CS1001, MDX-1105(BMS-936559), LY3300054, STI-A1014, FAZ053, CX-072, INCB086550, GNS-1480, CA -170, CK-301, M-7824, HTI-1088 (HTI-131, SHR-1316), MSB-2311, AK-106, AVA-004, BBI-801, CA-327, CBA- 0710, CBT-502, FPT-155, IKT-201, IKT-703, 10-103, JS-003, KD-033, KY-1003, MCLA-145, MT-5050, SNA-02, BCD-135, APL-502 (CBT-402 or TQB2450), IMC-001, KD-045, INBRX-105, KN-046, IMC-2102, IMC-2101, KD Examples include -005, IMM-2502, 89Zr-CX-072, 89Zr-DFO-6E11, KY-1055, MEDI-1109, MT-5594, SL-279252, DSP-106, Gensci-047, REMD-290, N-809, PRS-344, FS-222, GEN-1046, BH-29xx, FS-118, their biosimilars, and their derivatives. In some embodiments, an anti-PD-L1 antibody or its antigen-binding fragment that competes for binding to PD-L1 with any of these art-recognized anti-PD-L1 antibodies or their antigen-binding fragments, or any of the novel anti-PD-L1 antibodies or their antigen-binding fragments described herein, may also be used. In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment is a derivative of any of the anti-PD-L1 antibodies or their antigen-binding fragments described herein.In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment is derived from an anti-PD-L1 antibody selected from the group consisting of durvalumab, atezolizumab, and avelumab.

[0162] In some embodiments, the first targeting moiety or anti-PD-L1 targeting moiety comprises (or consists of or is essentially composed of) one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments thereof, where the one or more anti-PD-L1 antibodies or antigen-binding fragments independently include H-CDR1 comprising the amino acid sequence of SEQ ID NO: 3, H-CDR2 comprising the amino acid sequence of SEQ ID NO: 8, H-CDR3 comprising the amino acid sequence of SEQ ID NO: 13, L-CDR1 comprising the amino acid sequence of SEQ ID NO: 20, L-CDR2 comprising the amino acid sequence of SEQ ID NO: 25, and L-CDR3 comprising the amino acid sequence of SEQ ID NO: 30. In some embodiments, the first targeting moiety or anti-PD-L1 targeting moiety comprises (or consists of or essentially comprises) one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments, wherein the one or more anti-PD-L1 antibodies or antigen-binding fragments are independently: i) VH containing the amino acid sequence of SEQ ID NO: 17 and VL containing the amino acid sequence of SEQ ID NO: 34; ii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 221; vi ) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 and VL containing the amino acid sequence of SEQ ID NO: 222; viiii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 222; x) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 224; xii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 225; xiii) VH containing the amino acid sequence of SEQ ID NO: 16 and VL containing the amino acid sequence of SEQ ID NO: 226;xiv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 224; xvi) VH containing the amino acid sequence of SEQ ID NO: 223 and VL containing the amino acid sequence of SEQ ID NO: 226; xvii) VH containing the amino acid sequence of SEQ ID NO: 227 and VL containing the amino acid sequence of SEQ ID NO: 226; xviii) VH containing the amino acid sequence of SEQ ID NO: 228 and VL containing the amino acid sequence of SEQ ID NO: 226; xix) VH containing the amino acid sequence of SEQ ID NO: 229 and VL containing the amino acid sequence of SEQ ID NO: 226; xx) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 226; xxi) VH containing the amino acid sequence of SEQ ID NO: 230 and VL containing the amino acid sequence of SEQ ID NO: 220; or xxii) VH containing the amino acid sequence of SEQ ID NO: 219 and VL containing the amino acid sequence of SEQ ID NO: 226. In some embodiments, the first targeting moiety or anti-PD-L1 targeting moiety is a full-length anti-PD-L1 antibody, for example, i) a heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) a heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) a heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) a heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) a heavy chain containing the amino acid sequence of SEQ ID NO: 87 vii) Heavy chain containing the amino acid sequence of SEQ ID NO: 88 and light chain containing the amino acid sequence of SEQ ID NO: 91 and light chain containing the amino acid sequence of SEQ ID NO: 92; viii) Heavy chain containing the amino acid sequence of SEQ ID NO: 93 and light chain containing the amino acid sequence of SEQ ID NO: 94; ix) Heavy chain containing the amino acid sequence of SEQ ID NO: 95 and light chain containing the amino acid sequence of SEQ ID NO: 96; x) Heavy chain containing the amino acid sequence of SEQ ID NO: 97 and light chain containing the amino acid sequence of SEQ ID NO: 98; xi) Heavy chain containing the amino acid sequence of SEQ ID NO: 99 and light chain containing the amino acid sequence of SEQ ID NO: 100; xii) Heavy chain containing the amino acid sequence of SEQ ID NO: 101 and light chain containing the amino acid sequence of SEQ ID NO: 102;xiii) Heavy chain containing the amino acid sequence of SEQ ID NO: 103 and light chain containing the amino acid sequence of SEQ ID NO: 104; xiv) Heavy chain containing the amino acid sequence of SEQ ID NO: 105 and light chain containing the amino acid sequence of SEQ ID NO: 106; xv) Heavy chain containing the amino acid sequence of SEQ ID NO: 107 and light chain containing the amino acid sequence of SEQ ID NO: 108; xvi) Heavy chain containing the amino acid sequence of SEQ ID NO: 109 and light chain containing the amino acid sequence of SEQ ID NO: 110; xvii) Heavy chain containing the amino acid sequence of SEQ ID NO: 111 and light chain containing the amino acid sequence of SEQ ID NO: 112; xviii) Heavy chain containing the amino acid sequence of SEQ ID NO: 113 and light chain containing the amino acid sequence of SEQ ID NO: 114 ;xix) A heavy chain containing the amino acid sequence of SEQ ID NO: 115 and a light chain containing the amino acid sequence of SEQ ID NO: 116;xx) A heavy chain containing the amino acid sequence of SEQ ID NO: 117 and a light chain containing the amino acid sequence of SEQ ID NO: 118;xxi) A heavy chain containing the amino acid sequence of SEQ ID NO: 119 and a light chain containing the amino acid sequence of SEQ ID NO: 120;xxii) A heavy chain containing the amino acid sequence of SEQ ID NO: 121 and a light chain containing the amino acid sequence of SEQ ID NO: 122; or xxiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO: 34, comprising (or consisting of, or essentially being) a full-length anti-PD-L1 antibody comprising any of these. In some embodiments, the first targeting moiety or anti-PD-L1 targeting moiety comprises (or consists of or is essentially composed of) one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments, wherein the one or more anti-PD-L1 antibodies or antigen-binding fragments independently comprise: i) a VH containing the amino acid sequence of SEQ ID NO: 16 and a VL containing the amino acid sequence of SEQ ID NO: 33; and / or (ii) a heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO: 34.

[0163] In some embodiments, the first targeting moiety or anti-PD-L1 targeting moiety comprises (or consists of or is essentially composed of) one or more (e.g., one) anti-PD-L1 antibodies or antigen-binding fragments, wherein the one or more anti-PD-L1 antibodies or antigen-binding fragments are independently: H-CDR1 containing the amino acid sequence of SEQ ID NO: 244 or a variant thereof containing up to three (e.g., three, two, or one) amino acid mutations (e.g., substitutions, e.g., conserved substitutions); H-CDR2 containing the amino acid sequence of SEQ ID NO: 245 or a variant thereof containing up to three (e.g., three, two, or one) amino acid mutations (e.g., substitutions, e.g., conserved substitutions); H-CDR containing the amino acid sequence of SEQ ID NO: 246 The variants include: L-CDR1 containing up to three (e.g., three, two,...

Claims

1. A multispecific targeted conjugate comprising a first targeting portion that specifically recognizes PD-L1, a second targeting portion that specifically recognizes Trop-2, and an effector molecule, wherein the effector molecule is conjugated to the second targeting portion via a conjugation site.

2. The multispecific targeted conjugate according to claim 1, wherein the multispecific targeted conjugate further includes a cleavage site between the first targeted portion and the conjugation site, and the second targeted portion conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site.

3. The multispecific targeting conjugate has the structure of formula 1: 【Chemistry 25】 (In the formula, A1 is the first targeting portion, A2 is the second targeting portion, P is the cutting site, C is the conjugation site, L is a linker, D is an effector molecule, x = 0 or 1, a = 1 to 20, A multispecific targeting conjugate according to claim 1 or 2, comprising b = 1 to 20.

4. (i) The first and / or second targeting portion comprises one or more targeted peptides or antibodies or antigen-binding fragments thereof, and / or (ii) The multispecific targeted conjugate according to any one of claims 1 to 3, wherein the effector molecule is a therapeutic agent or an oligonucleotide.

5. The multispecific targeted conjugate according to any one of claims 2 to 4, wherein the aforementioned cleavage is induced by conditions at the target site.

6. (i) The conditions at the target site are selected from the group consisting of protease, pH change, redox change, hypoxia, oxidative stress, high heat, extracellular ATP concentration, and combinations thereof, and / or (ii) The multispecific targeted conjugate according to claim 5, wherein the target site is the site of a disease.

7. The multispecific targeted conjugate according to claim 6, wherein the disease is a tumor and the condition is the tumor microenvironment.

8. The multispecific targeted conjugate according to any one of claims 2 to 7, wherein the cleavage is performed by a protease.

9. The multispecific targeted conjugate according to claim 8, wherein the protease is MMP or uPA.

10. (i) The cleavage site that can be cleaved by the MMP includes any of the amino acid sequences of SEQ ID NOs: 71 and 137-143, and / or (ii) The multispecific targeted conjugate according to claim 9, wherein the cleavage site that can be cleaved by the uPA includes the amino acid sequence of SEQ ID NO:

214.

11. The multispecific targeted conjugate according to any one of claims 1 to 10, wherein the effector molecule is a therapeutic agent.

12. The multispecific targeted conjugate according to claim 11, wherein the therapeutic agent is exatecan.

13. A multispecific targeted conjugate according to any one of claims 3, 4, 11, and 12, wherein x is 0.

14. A multispecific targeted conjugate according to any one of claims 3, 4, 11, and 12, wherein x is 1.

15. (i) a is 1 to 10 and / or (ii) The multispecific targeted conjugate according to any one of claims 3 to 14, wherein b is 2 to 10.

16. The multispecific targeted conjugate according to any one of claims 1 to 15, wherein the conjugation site includes a transglutaminase conjugation site.

17. (i) The transglutaminase conjugation site comprises any of the amino acid sequences of SEQ ID NOs: 145 to 196, and / or (ii) The multispecific targeted conjugate according to claim 16, wherein the transglutaminase conjugation site comprises two or more glutamine-containing tags fused in series with respect to each other.

18. (i) L is the formula: (Gly) ｎ - (PEG) ｍ -VC-PAB-(DMAE) ｋ (wherein n, m, and k are integers, n ≥ 1, m ≥ 2, and k is 0 or 1) or it can be expressed as follows: (ii) L is the formula: (Gly) ｎ - (PEG) ｍ -VA-PAB-(DMAE) ｋ (wherein n, m, and k are integers, n ≥ 1, m ≥ 2, and k is 0 or 1) or it can be expressed as follows: (iii) L is (Gly) ６ - Amide - PEG ８ -VA-PAB or (iv) L is (Gly) ６ ６ -amide-PEG ８ ８ -VC-PAB, the multispecific targeting conjugate according to any one of claims 3 to 17.

19. L-(D) ａ However, the structure of equation A: 【Chemistry 26】 A multispecific targeted conjugate according to any one of claims 3 to 18, comprising (wherein the formula, the wavy line indicates the site of covalent bonding to the conjugation site C).

20. The multispecific targeted conjugate according to any one of claims 1 to 19, wherein the second targeted portion includes one or more sdAbs (anti-Trop-2 sdAbs) that specifically recognize Trop-2.

21. The one or more anti-Trop-2 sdAb compounds described above, each independently, i) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 42 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or a variant thereof containing up to three amino acid mutations; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 35 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 40 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 45 or a variant thereof containing up to three amino acid mutations; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 36 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 41 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 46 or a variant thereof containing up to three amino acid mutations; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 38 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 43 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 48 or a variant thereof containing up to three amino acid mutations; v) CDR1 containing the amino acid sequence of SEQ ID NO: 39 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 44 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 49 or a variant thereof containing up to three amino acid mutations; vi) CDR1 containing the amino acid sequence of SEQ ID NO: 53 or a variant thereof containing up to three amino acid mutations; CDR2 containing the amino acid sequence of SEQ ID NO: 58 or a variant thereof containing up to three amino acid mutations; and CDR3 containing the amino acid sequence of SEQ ID NO: 63 or a variant thereof containing up to three amino acid mutations; vii) CDR1 containing the amino acid sequence of SEQ ID NO: 51 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 56 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 61 or a variant thereof containing up to three amino acid mutations; viiii) CDR1 containing the amino acid sequence of SEQ ID NO: 52 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 57 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 62 or a variant thereof containing up to three amino acid mutations; ix) CDR1 containing the amino acid sequence of SEQ ID NO: 54 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 59 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 64 or a variant thereof containing up to three amino acid mutations; x) CDR1 containing the amino acid sequence of SEQ ID NO: 55 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 60 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 65 or a variant thereof containing up to three amino acid mutations; xi) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 205 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or a variant thereof containing up to three amino acid mutations; xi) CDR1 containing the amino acid sequence of SEQ ID NO: 53 or a variant thereof containing up to three amino acid mutations; CDR2 containing the amino acid sequence of SEQ ID NO: 58 or a variant thereof containing up to three amino acid mutations; and CDR3 containing the amino acid sequence of SEQ ID NO: 206 or a variant thereof containing up to three amino acid mutations; xiiii) CDR1 containing the amino acid sequence of SEQ ID NO: 55 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 60 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 243 or a variant thereof containing up to three amino acid mutations; or The multispecific targeted conjugate according to claim 20, comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 217 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 218 or a variant thereof containing up to three amino acid mutations, which is an anti-Trop-2 VHH.

22. The multispecific targeted conjugate according to claim 20 or 21, wherein each of the one or more anti-Trop-2 sdAb is an anti-Trop-2 VHH, each independently containing one of the amino acid sequences of SEQ ID NOs. 50, 66, 123-136, and 203.

23. The multispecific targeted conjugate according to any one of claims 20 to 22, wherein the second targeted portion comprises a first anti-Trop-2 sdAb and a second anti-Trop-2 sdAb fused in series with respect to each other.

24. The multispecific targeted conjugate according to claim 23, wherein the first anti-Trop-2 sdAb and the second anti-Trop-2 sdAb bind to different Trop-2 epitopes.

25. The multispecific targeted conjugate according to any one of claims 1 to 24, wherein the first targeting portion comprises one or more antibodies or antigen-binding fragments (anti-PD-L1 antibodies or antigen-binding fragments) that specifically recognize PD-L1.

26. The one or more anti-PD-L1 antibodies or their antigen-binding fragments independently i) Heavy chain CDR1 containing the amino acid sequence of SEQ ID NO: 3 ("H-CDR1") or a variant thereof containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO: 8 or a variant thereof containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO: 13 or a variant thereof containing up to three amino acid mutations; Light chain CDR1 containing the amino acid sequence of SEQ ID NO: 20 ("L-CDR1") or a variant thereof containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO: 25 or a variant thereof containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30 or a variant thereof containing up to three amino acid mutations; ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 1 or a variant thereof containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO: 6 or a variant thereof containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO: 11 or a variant thereof containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO: 18 or a variant thereof containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO: 23 or a variant thereof containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO: 28 or a variant thereof containing up to three amino acid mutations; iii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 2 or a variant thereof containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO: 7 or a variant thereof containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO: 12 or a variant thereof containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO: 19 or a variant thereof containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO: 24 or a variant thereof containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO: 29 or a variant thereof containing up to three amino acid mutations; iv) H-CDR1 containing the amino acid sequence of SEQ ID NO: 4 or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 9 or a variant thereof containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 14 or a variant thereof containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 21 or a variant thereof containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 26 or a variant thereof containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 31 or a variant thereof containing up to three amino acid mutations; or v) The multispecific targeted conjugate according to claim 25, comprising H-CDR1 containing the amino acid sequence of SEQ ID NO: 5 or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 10 or a variant thereof containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 15 or a variant thereof containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 22 or a variant thereof containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 27 or a variant thereof containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 32 or a variant thereof containing up to three amino acid mutations.

27. The one or more anti-PD-L1 antibodies or their antigen-binding fragments independently i) A variant thereof containing the heavy chain variable region (VH) with the amino acid sequence of SEQ ID NO: 16 or having at least about 85% sequence identity to SEQ ID NO: 16, and a variant thereof containing the light chain variable region (VL) with the amino acid sequence of SEQ ID NO: 33 or having at least about 85% sequence identity to SEQ ID NO: 33; ii) VH containing the amino acid sequence of SEQ ID NO: 219 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 219, and VL containing the amino acid sequence of SEQ ID NO: 220 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 213 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 33 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 221 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 213 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 222; viiii) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 221 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 222; x) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 224; xi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 225; xiiii) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xiv) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 224; xvi) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 226; xvii) VH containing the amino acid sequence of SEQ ID NO: 227 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 227, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 226; xviiii) VH containing the amino acid sequence of SEQ ID NO: 228 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 228, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xix) VH containing the amino acid sequence of SEQ ID NO: 229 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 229, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xx) VH containing the amino acid sequence of SEQ ID NO: 230 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 230, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xxi) VH containing the amino acid sequence of SEQ ID NO: 230 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 230, and VL containing the amino acid sequence of SEQ ID NO: 220 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 220; or xxii) A multispecific targeted conjugate according to claim 25 or 26, comprising a VH containing the amino acid sequence of SEQ ID NO: 219 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 219, and a VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO:

226.

28. The multispecific targeted conjugate according to any one of claims 25 to 27, wherein the first targeted portion comprises a full-length anti-PD-L1 antibody.

29. The aforementioned full-length anti-PD-L1 antibody, i) A heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) A heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) A heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) A heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) A heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) A heavy chain containing the amino acid sequence of SEQ ID NO: 87 and a light chain containing the amino acid sequence of SEQ ID NO: 88; vii) A heavy chain containing the amino acid sequence of SEQ ID NO: 91 and a light chain containing the amino acid sequence of SEQ ID NO: 92; viiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 93 and a light chain containing the amino acid sequence of SEQ ID NO: 94; ix) A heavy chain containing the amino acid sequence of SEQ ID NO: 95 and a light chain containing the amino acid sequence of SEQ ID NO: 96; x) A heavy chain containing the amino acid sequence of SEQ ID NO: 97 and a light chain containing the amino acid sequence of SEQ ID NO: 98; xi) A heavy chain containing the amino acid sequence of SEQ ID NO: 99 and a light chain containing the amino acid sequence of SEQ ID NO: 100; xi) A heavy chain containing the amino acid sequence of SEQ ID NO: 101 and a light chain containing the amino acid sequence of SEQ ID NO: 102; xiiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 103 and a light chain containing the amino acid sequence of SEQ ID NO: 104; xiv) A heavy chain containing the amino acid sequence of SEQ ID NO: 105 and a light chain containing the amino acid sequence of SEQ ID NO: 106; xv) A heavy chain containing the amino acid sequence of SEQ ID NO: 107 and a light chain containing the amino acid sequence of SEQ ID NO: 108; xvi) A heavy chain containing the amino acid sequence of SEQ ID NO: 109 and a light chain containing the amino acid sequence of SEQ ID NO: 110; xvii) A heavy chain containing the amino acid sequence of SEQ ID NO: 111 and a light chain containing the amino acid sequence of SEQ ID NO: 112; xviiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 113 and a light chain containing the amino acid sequence of SEQ ID NO: 114; xix) A heavy chain containing the amino acid sequence of SEQ ID NO: 115 and a light chain containing the amino acid sequence of SEQ ID NO: 116; xx) A heavy chain containing the amino acid sequence of SEQ ID NO: 117 and a light chain containing the amino acid sequence of SEQ ID NO: 118; xxi) A heavy chain containing the amino acid sequence of SEQ ID NO: 119 and a light chain containing the amino acid sequence of SEQ ID NO: 120; xxii) A heavy chain containing the amino acid sequence of SEQ ID NO: 121 and a light chain containing the amino acid sequence of SEQ ID NO: 122; or The multispecific targeted conjugate according to claim 28, comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO:

34.

30. The multispecific targeting conjugate extends from the N-terminus to the C-terminus. i) Anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - conjugation site - any linker 2 - anti-Trop-2 sdAb; ii) Anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - cleavage site - any linker 2 - conjugation site - any linker 3 - anti-Trop-2 sdAb; iii) Anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - conjugation site - any linker 2 - first anti-Trop-2 sdAb - any linker 3 - second anti-Trop-2 sdAb; or iv) A multispecific targeted conjugate according to any one of claims 1 to 29, comprising an anti-PD-L1 antibody or its antigen-binding fragment - any linker 1 - cleavage site - any linker 2 - conjugation site - any linker 3 - first anti-Trop-2 sdAb - any linker 4 - second anti-Trop-2 sdAb.

31. The multispecific targeted conjugate according to any one of claims 1 to 30, wherein the multispecific targeted conjugate comprises a full-length anti-PD-L1 antibody, and the multispecific targeted conjugate comprises, from the N-terminus to the C-terminus, a fusion polypeptide: heavy chain of the full-length anti-PD-L1 antibody - any linker 1 - any cleavage site - any linker 2 - conjugation site - any linker 3 - one or more anti-Trop-2 sdAb, the heavy chain comprises the amino acid sequence of SEQ ID NO: 17, the light chain comprises the amino acid sequence of SEQ ID NO: 34, and the fusion polypeptide comprises the amino acid sequence of any of SEQ ID NOs: 67, 69, 215, and 238-240.

32. An isolated antibody construct (anti-PD-L1 antibody construct) comprising a targeting portion that specifically recognizes PD-L1 (anti-PD-L1 targeting portion), wherein the anti-PD-L1 targeting portion is i) H-CDR1 containing the amino acid sequence of SEQ ID NO: 3 or a variant thereof containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO: 8 or a variant thereof containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO: 13 or a variant thereof containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO: 20 or a variant thereof containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO: 25 or a variant thereof containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO: 30 or a variant thereof containing up to three amino acid mutations; ii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 1 or a variant thereof containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO: 6 or a variant thereof containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO: 11 or a variant thereof containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO: 18 or a variant thereof containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO: 23 or a variant thereof containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO: 28 or a variant thereof containing up to three amino acid mutations; iii) H-CDR1 containing the amino acid sequence of SEQ ID NO: 2 or a variant thereof containing up to three amino acid mutations; H-CDR2 containing the amino acid sequence of SEQ ID NO: 7 or a variant thereof containing up to three amino acid mutations; H-CDR3 containing the amino acid sequence of SEQ ID NO: 12 or a variant thereof containing up to three amino acid mutations; L-CDR1 containing the amino acid sequence of SEQ ID NO: 19 or a variant thereof containing up to three amino acid mutations; L-CDR2 containing the amino acid sequence of SEQ ID NO: 24 or a variant thereof containing up to three amino acid mutations; and L-CDR3 containing the amino acid sequence of SEQ ID NO: 29 or a variant thereof containing up to three amino acid mutations; iv) H-CDR1 containing the amino acid sequence of SEQ ID NO: 4 or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 9 or a variant thereof containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 14 or a variant thereof containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 21 or a variant thereof containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 26 or a variant thereof containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 31 or a variant thereof containing up to three amino acid mutations; or v) Including H-CDR1 containing the amino acid sequence of SEQ ID NO: 5 or a variant thereof containing up to three amino acid mutations, H-CDR2 containing the amino acid sequence of SEQ ID NO: 10 or a variant thereof containing up to three amino acid mutations, H-CDR3 containing the amino acid sequence of SEQ ID NO: 15 or a variant thereof containing up to three amino acid mutations, L-CDR1 containing the amino acid sequence of SEQ ID NO: 22 or a variant thereof containing up to three amino acid mutations, L-CDR2 containing the amino acid sequence of SEQ ID NO: 27 or a variant thereof containing up to three amino acid mutations, and L-CDR3 containing the amino acid sequence of SEQ ID NO: 32 or a variant thereof containing up to three amino acid mutations.

33. The anti-PD-L1 targeting portion is i) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 33 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 33; ii) VH containing the amino acid sequence of SEQ ID NO: 219 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 219, and VL containing the amino acid sequence of SEQ ID NO: 220 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 220; iii) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 213 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 213; iv) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 33 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 33; v) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 221 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 221; vi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 213 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 213; vii) VH containing the amino acid sequence of SEQ ID NO: 212 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 212, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 222; viiii) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 221 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 221; ix) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 222; x) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 222 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 222; xi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 224; xi) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 225; xiiii) VH containing the amino acid sequence of SEQ ID NO: 16 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 16, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xiv) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 225 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 225; xv) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 224 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 224; xvi) VH containing the amino acid sequence of SEQ ID NO: 223 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 223, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 226; xvii) VH containing the amino acid sequence of SEQ ID NO: 227 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 227, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 226; xviiii) VH containing the amino acid sequence of SEQ ID NO: 228 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 228, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xix) VH containing the amino acid sequence of SEQ ID NO: 229 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 229, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xx) VH containing the amino acid sequence of SEQ ID NO: 230 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 230, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least approximately 85% sequence identity to SEQ ID NO: 226; xxi) VH containing the amino acid sequence of SEQ ID NO: 230 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 230, and VL containing the amino acid sequence of SEQ ID NO: 220 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 220; or xxii) An isolated anti-PD-L1 antibody construct according to claim 32, comprising VH containing the amino acid sequence of SEQ ID NO: 219 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO: 219, and VL containing the amino acid sequence of SEQ ID NO: 226 or a variant thereof containing at least about 85% sequence identity to SEQ ID NO:

226.

34. The isolated anti-PD-L1 antibody construct according to claim 32 or 33, wherein the anti-PD-L1 targeting portion is a full-length antibody (a full-length anti-PD-L1 antibody).

35. The aforementioned full-length anti-PD-L1 antibody, i) A heavy chain containing the amino acid sequence of SEQ ID NO: 89 and a light chain containing the amino acid sequence of SEQ ID NO: 90; ii) A heavy chain containing the amino acid sequence of SEQ ID NO: 79 and a light chain containing the amino acid sequence of SEQ ID NO: 80; iii) A heavy chain containing the amino acid sequence of SEQ ID NO: 81 and a light chain containing the amino acid sequence of SEQ ID NO: 82; iv) A heavy chain containing the amino acid sequence of SEQ ID NO: 83 and a light chain containing the amino acid sequence of SEQ ID NO: 84; v) A heavy chain containing the amino acid sequence of SEQ ID NO: 85 and a light chain containing the amino acid sequence of SEQ ID NO: 86; vi) A heavy chain containing the amino acid sequence of SEQ ID NO: 87 and a light chain containing the amino acid sequence of SEQ ID NO: 88; vii) A heavy chain containing the amino acid sequence of SEQ ID NO: 91 and a light chain containing the amino acid sequence of SEQ ID NO: 92; viiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 93 and a light chain containing the amino acid sequence of SEQ ID NO: 94; ix) A heavy chain containing the amino acid sequence of SEQ ID NO: 95 and a light chain containing the amino acid sequence of SEQ ID NO: 96; x) A heavy chain containing the amino acid sequence of SEQ ID NO: 97 and a light chain containing the amino acid sequence of SEQ ID NO: 98; xi) A heavy chain containing the amino acid sequence of SEQ ID NO: 99 and a light chain containing the amino acid sequence of SEQ ID NO: 100; xi) A heavy chain containing the amino acid sequence of SEQ ID NO: 101 and a light chain containing the amino acid sequence of SEQ ID NO: 102; xiiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 103 and a light chain containing the amino acid sequence of SEQ ID NO: 104; xiv) A heavy chain containing the amino acid sequence of SEQ ID NO: 105 and a light chain containing the amino acid sequence of SEQ ID NO: 106; xv) A heavy chain containing the amino acid sequence of SEQ ID NO: 107 and a light chain containing the amino acid sequence of SEQ ID NO: 108; xvi) A heavy chain containing the amino acid sequence of SEQ ID NO: 109 and a light chain containing the amino acid sequence of SEQ ID NO: 110; xvii) A heavy chain containing the amino acid sequence of SEQ ID NO: 111 and a light chain containing the amino acid sequence of SEQ ID NO: 112; xviiii) A heavy chain containing the amino acid sequence of SEQ ID NO: 113 and a light chain containing the amino acid sequence of SEQ ID NO: 114; xix) A heavy chain containing the amino acid sequence of SEQ ID NO: 115 and a light chain containing the amino acid sequence of SEQ ID NO: 116; xx) A heavy chain containing the amino acid sequence of SEQ ID NO: 117 and a light chain containing the amino acid sequence of SEQ ID NO: 118; xxi) A heavy chain containing the amino acid sequence of SEQ ID NO: 119 and a light chain containing the amino acid sequence of SEQ ID NO: 120; xxii) A heavy chain containing the amino acid sequence of SEQ ID NO: 121 and a light chain containing the amino acid sequence of SEQ ID NO: 122; or The isolated anti-PD-L1 antibody construct according to claim 34, comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 17 and a light chain containing the amino acid sequence of SEQ ID NO:

34.

36. The isolated anti-PD-L1 antibody construct according to any one of claims 32 to 35, wherein the isolated anti-PD-L1 antibody construct is multispecific.

37. The isolated anti-PD-L1 antibody construct according to any one of claims 32 to 36, wherein the isolated anti-PD-L1 antibody construct is a multispecific targeted conjugate comprising a first targeting portion that specifically recognizes PD-L1, a second targeting portion that specifically recognizes Top-2, and an effector molecule, the first targeting portion being an anti-PD-L1 targeted portion, and the effector molecule being conjugated to the second targeting portion via a conjugation site.

38. The isolated anti-PD-L1 antibody construct according to claim 3, wherein the multispecific targeted conjugate further includes a cleavage site between the first targeted portion and the conjugation site, and the second targeted portion conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site.

39. An isolated antibody construct (anti-Trop-2 antibody construct) comprising a targeting portion that specifically recognizes Trop-2 (anti-Trop-2 targeting portion), wherein the anti-Trop-2 targeting portion is i) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 42 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or a variant thereof containing up to three amino acid mutations; ii) CDR1 containing the amino acid sequence of SEQ ID NO: 35 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 40 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 45 or a variant thereof containing up to three amino acid mutations; iii) CDR1 containing the amino acid sequence of SEQ ID NO: 36 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 41 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 46 or a variant thereof containing up to three amino acid mutations; iv) CDR1 containing the amino acid sequence of SEQ ID NO: 38 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 43 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 48 or a variant thereof containing up to three amino acid mutations; v) CDR1 containing the amino acid sequence of SEQ ID NO: 39 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 44 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 49 or a variant thereof containing up to three amino acid mutations; vi) CDR1 containing the amino acid sequence of SEQ ID NO: 53 or a variant thereof containing up to three amino acid mutations; CDR2 containing the amino acid sequence of SEQ ID NO: 58 or a variant thereof containing up to three amino acid mutations; and CDR3 containing the amino acid sequence of SEQ ID NO: 63 or a variant thereof containing up to three amino acid mutations; vii) CDR1 containing the amino acid sequence of SEQ ID NO: 51 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 56 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 61 or a variant thereof containing up to three amino acid mutations; viiii) CDR1 containing the amino acid sequence of SEQ ID NO: 52 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 57 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 62 or a variant thereof containing up to three amino acid mutations; ix) CDR1 containing the amino acid sequence of SEQ ID NO: 54 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 59 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 64 or a variant thereof containing up to three amino acid mutations; x) CDR1 containing the amino acid sequence of SEQ ID NO: 55 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 60 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 65 or a variant thereof containing up to three amino acid mutations; xi) CDR1 containing the amino acid sequence of SEQ ID NO: 37 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 205 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 47 or a variant thereof containing up to three amino acid mutations; xi) CDR1 containing the amino acid sequence of SEQ ID NO: 53 or a variant thereof containing up to three amino acid mutations; CDR2 containing the amino acid sequence of SEQ ID NO: 58 or a variant thereof containing up to three amino acid mutations; and CDR3 containing the amino acid sequence of SEQ ID NO: 206 or a variant thereof containing up to three amino acid mutations; xiiii) CDR1 containing the amino acid sequence of SEQ ID NO: 55 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 60 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 243 or a variant thereof containing up to three amino acid mutations; or The isolated antibody construct is VHH (anti-Trop-2 VHH), comprising CDR1 containing the amino acid sequence of SEQ ID NO: 216 or a variant thereof containing up to three amino acid mutations, CDR2 containing the amino acid sequence of SEQ ID NO: 217 or a variant thereof containing up to three amino acid mutations, and CDR3 containing the amino acid sequence of SEQ ID NO: 218 or a variant thereof containing up to three amino acid mutations.

40. The isolated anti-Trop-2 antibody construct according to claim 39, wherein the anti-Trop-2 VHH comprises any of the amino acid sequences of SEQ ID NOs. 50, 66, 123-136, and 203.

41. The isolated anti-Trop-2 antibody construct according to claim 39 or 40, wherein the isolated anti-Trop-2 antibody construct comprises two or more anti-Trop-2 VHH molecules fused in series with each other.

42. The isolated anti-Trop-2 antibody construct according to any one of claims 39 to 41, wherein the isolated anti-Trop-2 antibody construct is multispecific.

43. The isolated anti-Trop-2 antibody construct according to any one of claims 39 to 42, wherein the isolated anti-Trop-2 antibody construct comprises a first anti-Trop-2 VHH and a second anti-Trop-2 VHH.

44. The isolated anti-Trop-2 antibody construct according to any one of claims 39 to 43, wherein the isolated anti-Trop-2 antibody construct is a multispecific targeted conjugate comprising a first targeting portion that specifically recognizes PD-L1, a second targeting portion that specifically recognizes Top-2, and an effector molecule, the second targeting portion being the anti-Trop-2 targeted portion, and the effector molecule being conjugated to the second targeting portion via a conjugation site.

45. The isolated anti-Trop-2 antibody construct according to claim 44, wherein the multispecific targeted conjugate further includes a cleavage site between the first targeted portion and the conjugation site, and the second targeted portion conjugated with the effector molecule can be released from the multispecific targeted conjugate by cleavage at the cleavage site.

46. i) an isolated anti-PD-L1 antibody construct according to any one of claims 32 to 38, or an isolated anti-Trop-2 antibody construct according to any one of claims 39 to 45, and ii) optionally a pharmaceutical composition comprising a pharmaceutically acceptable carrier.

47. A pharmaceutical composition comprising one or more multispecific targeted conjugates according to any one of claims 1 to 31, 37, 38, 44, and 45, and optionally a pharmaceutically acceptable carrier.

48. A method for treating a Trop-2 positive cancer in an individual, comprising administering to the individual an effective amount of a multispecific targeted conjugate according to any one of claims 1 to 31, 37, 38, 44, and 45, or a pharmaceutical composition according to claim 47.

49. A method for producing a multispecific targeted conjugate according to any one of claims 1 to 31, 37, 38, 44, and 45, comprising conjugating the effector molecule to the second targeted portion via the conjugation site.

50. The method described above is (a) The step of reacting the first targeting portion, the second targeting portion, and the multispecific targeting portion including the conjugation site with a linker reagent to form a multispecific targeting portion-linker intermediate, (b) The step of reacting the multispecific targeting moiety-linker intermediate with a nucleophile of the effector molecular moiety, thereby forming the multispecific targeting conjugate, or the step of Or the method described above, (c) A step of reacting the effector molecule with a linker reagent to form a linker-effector molecule intermediate, (d) A step of reacting the linker-effector molecular intermediate with the conjugation site of the multispecific targeting portion, which includes the first targeting portion, the second targeting portion, and the conjugation site, thereby forming the multispecific targeting conjugate, or the step of Or the method described above, (e) The method according to claim 49, comprising the step of reacting the multispecific targeting portion, which includes the first targeting portion, the second targeting portion, and the conjugation portion, with a linker-effector molecule conjugate, thereby forming the multispecific targeting conjugate.

51. The method according to claim 49 or 50, wherein the conjugation is mediated by transglutaminase.

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