Anti-pd-l1 antibodies and antibody-drug conjugates and their use in the treatment of cancer

Abstract

The present invention provides methods and compositions for treating cancers in subjects, such as non-small cell lung cancer, head and neck cancer, esophageal cancer, ovarian cancer, melanoma, breast cancer, and gastric cancer, such as by administering an antibody and antibody-drug conjugate that bind to programmed death ligand 1 (PD-L1).

Description

[0001] For all purposes, the contents of U.S. Provisional Patent Application No. 63 / 626,868 (filed January 30, 2024), U.S. Provisional Patent Application No. 63 / 557,770 (filed February 26, 2024), U.S. Provisional Patent Application No. 63 / 691,610 (filed September 6, 2024), and U.S. Provisional Patent Application No. 63 / 736,264 (filed December 19, 2024) are incorporated herein by reference. Technical Field

[0002] This invention relates to methods and compositions for treating cancers in subjects, such as non-small cell lung cancer, head and neck cancer, esophageal cancer, ovarian cancer, melanoma, breast cancer, and gastric cancer, such as by administering an antibody and antibody-drug conjugate that bind to programmed death ligand 1 (PD-L1). Background Technology

[0003] PD-L1 (also known as programmed death-ligand 1, B7-H1, or CD274) is a protein that has been shown to be expressed in a variety of cancer cells. PD-L1 is a transmembrane protein that interacts with PD-1 and acts as an "off" switch to inactivate T cells. PD-L1 is typically overexpressed on tumor cells and binds to PD-1, allowing tumors to evade T cell immune responses.

[0004] Several cancers express PD-L1, including melanoma. Melanoma is the most dangerous type of skin cancer. In 2015, 3.1 million people had active melanoma, and it caused 59,800 deaths. The five-year survival rate for stage IV disease is less than 10%, and the median survival is only 6 to 12 months. Therefore, there is a need to improve treatments for melanoma and other cancers that express PD-L1. One type of treatment for PD-L1-expressing cancers involves administering anti-PD-L1 antibodies as immunotherapy. Immuno-oncology is a promising field in cancer treatment, but there is still room for improvement in current therapies.

[0005] This invention addresses the need for improved treatment of cancers such as non-small cell lung cancer, head and neck cancer, esophageal cancer, ovarian cancer, melanoma, breast cancer, and gastric cancer by providing highly specific and effective anti-PD-L1 antibody-drug conjugates.

[0006] All references cited in this article (including patent applications, patent publications, and scientific articles) are incorporated herein by full reference, as if each individual reference were specifically and separately indicated to be incorporated herein by reference. Summary of the Invention

[0007] This article provides a method for treating cancer in human subjects. In some embodiments, the method includes administering to a subject an effective amount of an antibody-drug conjugate (ADC) comprising an anti-PD-L1 antibody or an antigen-binding fragment thereof, and an effective amount of an anti-PD-1 antibody; wherein the anti-PD-L1 antibody or the antigen-binding fragment thereof binds to PD-L1 and is conjugated to one or more units of auristatin E (MMAE); wherein the anti-PD-L1 antibody or the antigen-binding fragment thereof comprises: (i) a light chain variable region comprising light chain CDR1, CDR2, and CDR3 of SEQ ID NO: 8, 9, and 10, respectively, and (ii) a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 of SEQ ID NO: 5, 6, and 7, respectively; and wherein the anti-PD-1 antibody comprises: (i) a light chain variable region comprising light chain CDR1, CDR2, and CDR3 of SEQ ID NO: 20, 21, and 22, respectively, and (ii) a heavy chain variable region comprising heavy ... Heavy chains CDR1, CDR2 and CDR3 of 17, 18 and 19.

[0008] In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment comprises: a heavy chain variable region comprising an amino acid sequence that is at least 85% identical to the amino acid sequence of SEQ ID NO:3 and a light chain variable region comprising an amino acid sequence that is at least 85% identical to the amino acid sequence of SEQ ID NO:4. In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment comprises: a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:3 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:4. In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:1 and a light chain comprising the amino acid sequence of SEQ ID NO:2.

[0009] In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment is conjugated to the MMAE of each unit via a linker. In some embodiments, the linker is an enzyme-cleavable linker, and wherein the linker forms a bond with a sulfur atom of the antibody or its antigen-binding fragment. In some embodiments, the linker has the following formula: -Aa-Ww-Yy-; where -A- is a stretcher unit, a is 0 or 1; -W- is an amino acid unit, w is an integer in the range of 0 to 12; and -Y- is a spacer unit, y is 0, 1, or 2. In some embodiments, the stretcher unit has the structure of formula (1); the amino acid unit is valine-citrulline; and the spacer unit is a PAB group comprising the structure of formula (2):

[0010]

[0011] .

[0012] In some embodiments, the ADC comprises 1 to 20 units of MMAE per antibody or antigen-binding fragment thereof. In some embodiments, the ADC comprises 1 to 10 units of MMAE per antibody or antigen-binding fragment thereof. In some embodiments, the ADC comprises 2 to 8 units of MMAE per antibody or antigen-binding fragment thereof. In some embodiments, the ADC comprises 3 to 5 units of MMAE per antibody or antigen-binding fragment thereof. In some embodiments, the ADC has the following structure:

[0013]

[0014] Where L- represents an anti-PD-L1 antibody or its antigen-binding fragment, and p is 1 to 10. In some embodiments, p is 2 to 8. In some embodiments, p is 3 to 5. In some embodiments, p is 4. In some embodiments, p is 8.

[0015] In some implementation schemes, the cancer is melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), head and neck squamous cell carcinoma (HNSCC), triple-negative breast cancer (TNBC), esophageal squamous cell carcinoma (esophageal SCC), ovarian cancer, urothelial carcinoma, hepatocellular carcinoma (HCC), gastric cancer, or cervical cancer.

[0016] In some embodiments, the ADC is administered to the subject at a dose of about 0.25 to about 5 mg / kg of subject body weight, about 0.5 to about 2.5 mg / kg of subject body weight, or about 1 to about 2 mg / kg of subject body weight. In some embodiments, the subject body weight is the subject's ideal body weight (IBW). In some embodiments, the subject body weight is the subject's adjusted ideal body weight (AIBW). In some embodiments, the anti-PD-1 antibody is administered at a dose of 200 mg. In some embodiments, the ADC is administered about once a week, about once every two weeks, about once every three weeks, or about twice every three weeks. In some embodiments, the ADC is administered on days 1 and 8 of each 21-day cycle. In some embodiments, the anti-PD-1 antibody is administered about once a week, about once every two weeks, about once every three weeks, or about twice every three weeks. In some embodiments, the anti-PD-1 antibody is administered on day 1 of each 21-day cycle.

[0017] In some implementations, the subject has relapsed or refractory metastatic or unresectable solid malignant tumors that are intolerant to standard care. In some implementations, the ADC is administered intravenously. In some implementations, the anti-PD-1 antibody is administered intravenously. In some implementations, one or more treatment effects improve relative to baseline in the subject after administration of the antibody-drug conjugate. In some implementations, one or more treatment effects are selected from the group consisting of: tumor size derived from the cancer, objective response rate, duration of response, time to response, progression-free survival, and overall survival. Attached Figure Description

[0018] Figure 1 Showing the overall study design of a phase I study evaluating SGN-PDL1V. HNSCC = head and neck squamous cell carcinoma; NSCLC = non-small cell lung cancer; PD-L1 = programmed death-ligand 1; SCC = squamous cell carcinoma; TNBC = triple-negative breast cancer. (a) If necessary, alternative doses and schedules may be examined; (b) melanoma, ovarian cancer, TNBC, gastric cancer, or esophageal SCC; (c) PD-L1 negative NSCLC and HNSCC, melanoma, ovarian cancer, TNBC, gastric cancer, or esophageal SCC.

[0019] Figure 2 Preliminary patient data for all subjects during dose escalation and dose optimization are shown.

[0020] Figure 3 Preliminary patient data showing all HNSCC subjects in dose escalation ≥1.25 mg / kg.

[0021] Figure 4 This shows the incidence and grade of treatment-related adverse events (AEs) in patients treated with SGN-PDL1V.

[0022] Figure 5 The pharmacokinetics of SGN-PDL1V are shown compared to unconjugated MMAE.

[0023] Figure 6 The antitumor activity of SGN-PDL1V against PDL1+ tumors is shown by the change in the total tumor diameter.

[0024] Figure 7 The durability of the clinical response of SGN-PDL1V to PDL1+ tumors is shown, as indicated by changes in tumor size over time.

[0025] Figure 8 The antitumor activity of SGN-PDL1V against PDL1+ NSCLC is shown, as indicated by the change in total tumor diameter.

[0026] Figure 9 The durability of the clinical response to SGN-PDL1V in PDL1+ NSCLC is demonstrated by changes in tumor size over time.

[0027] Figure 10 This displays the viable cell counts of PD-L1 negative cells (single culture), PD-L1 negative cells (co-culture), or PD-L1 positive cells (co-culture) when treated with indicated concentrations of PDL1V, isovir alafenamide, or MMAE. Raw fluorescence data were analyzed using Flowjo software and processed in Graphpad Prism 8 using the formula [(PDL1neg or PDL1pos absolute viable cell count) / (mean PDL1neg or PDL1pos absolute viable cell count in the untreated group)].

[100] The percentage of live cells out of untreated cells was derived and plotted.

[0028] Figure 11A This displays the amount of extracellular ATP in each sample, where the raw luminescence data were analyzed in Graphpad Prism 8 and plotted as a fold change relative to the untreated group using the following formula: [Value in experimental group / Mean of untreated group]. The dashed line represents the mean value of the untreated group. Figure 11B This displays the amount of HMGB1 in the supernatant of each sample, where the raw luminescence values ​​were extrapolated from the HMGB1 standard curve of the positive control and plotted in Graphpad Prism 8. The dashed line represents the mean value of the untreated group. Figure 11C This displays the amount of calreticulin on the cell surface of each sample. The raw fluorescence data were analyzed in Flowjo and plotted using Graphpad Prism 8 to show the percentage of calreticulin-positive cells. The dashed line represents the mean value of the untreated group.

[0029] Figure 12A and Figure 12B This shows the amount of CD4+ cell death in donor 1 and donor 2 when treated with PDL1v, isovitoline, or a positive control, respectively. Figure 12C and Figure 12D This shows the amount of CD8+ cell death in donor 1 and donor 2 when treated with PDL1v, isoformicine, or a positive control, respectively. The raw data were analyzed in Flowjo v10 using GraphPad Prism 8, with non-linear, four-parameter curve fitting, the curve formula being [Y = bottom + ((top - bottom) / (1 + 10^((LogEC50 - X))]. HillSlope). Data were calculated using the formula [values ​​in the experimental group / mean of the untreated group]. 100 is calculated and displayed as the percentage of unprocessed data.

[0030] (2Q3W: Days 1 and 8 of a 21-day cycle; AiBW: Adjusted ideal body weight; CI: Confidence interval; cORR: Confirmed objective response rate; CR: Complete response; DCR: Disease control rate; ESCC: Esophageal squamous cell carcinoma; HNSCC: Head and neck squamous cell carcinoma; mDOR: Median duration of confirmed response; mPFS: Median progression-free survival; NE / NA: Not evaluable / unavailable; NSCLC: Non-small cell lung cancer; ORR: Objective response rate; PD: Disease progression; PD-L1: Programmed cell death ligand 1; PR: Partial response; RECIST: Response assessment criteria for solid tumors; SD: Stable disease; TNBC: Triple-negative breast cancer.) Invention Details

[0032] I. Definition

[0033] To facilitate understanding of this disclosure, certain terms are first defined. As used in this application, each of the following terms shall have the meaning set forth below unless expressly provided otherwise herein. Additional definitions will be set forth throughout the application.

[0034] The term “and / or” as used herein should be considered as each of two specified features or components having or not having a particular disclosure of the other. Thus, the term “and / or” as used in phrases such as “A and / or B” herein is intended to include “A and B”, “A or B”, “A alone”, and “B alone”. Similarly, the term “and / or” as used in phrases such as “A, B, and / or C” is intended to cover each of the following: 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.

[0035] It should be understood that the aspects and embodiments of the present invention described herein include “comprising aspects and embodiments,” “consisting of aspects and embodiments,” and “substantially consisting of aspects and embodiments.”

[0036] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. For example, *Concise Dictionary of Biomedicine and Molecular Biology*, Juo, Pei-Show, 2nd ed., 2002, CRC Press; *The Dictionary of Cell and Molecular Biology*, 3rd ed., 1999, Academic Press; and *The Oxford Dictionary of Biochemistry and Molecular Biology*, Revised, 2000, Oxford University Press, provide a general dictionary for those skilled in the art of the many terms used in this disclosure.

[0037] Units, prefixes, and symbols are represented in a form accepted by the International System of Units (SI). Numerical ranges include the numbers that define the range. The headings provided herein are not intended to limit the various aspects of this disclosure, which can be found throughout the specification. Therefore, the terms defined below are defined more fully with reference to the entire specification.

[0038] The terms “PD-L1”, “CD274”, “B7-H1” and “programmed cell death ligand 1” are used interchangeably herein and, unless otherwise specified, include any variant, isoform, and species homolog of human PD-L1 that is normally expressed by cells or expressed on cells transfected with the PD-L1 gene.

[0039] The term "immunoglobulin" refers to a structurally related class of glycoproteins consisting of two pairs of polypeptide chains (a pair of light (L) low molecular weight chains and a pair of heavy (H) chains), all four chains being linked by disulfide bonds. The structure of immunoglobulins has been well characterized. See, for example, Fundamental Immunology Ch. 7 (Paul, W., ed., 2nd ed. Raven Press, N.Y. (1989)). In short, each heavy chain generally contains a heavy chain variable region (abbreviated as V in this paper). H or VH) and heavy chain constant region (C H Or CH). The heavy chain constant region generally contains three structural domains, C H1. C H 2 and C H 3. Heavy chains are typically interconnected via disulfide bonds in so-called "hinge regions." Each light chain generally contains a light chain variable region (abbreviated as V in this paper). L or VL) and light chain constant region (C L Or CL). The constant region of a light chain generally contains a structural domain, C L CL can be a κ (kappa) or λ (lambda) isotype. The terms "constant domain" and "constant region" are used interchangeably herein. Immunoglobulins can be derived from any of the commonly known isotypes, including but not limited to IgA, secreted IgA, IgG, and IgM. IgG subclasses are also well known to those skilled in the art and include, but are not limited to, human IgG1, IgG2, IgG3, and IgG4. "Isotype" refers to an antibody class or subclass (e.g., IgM or IgG1) encoded by a heavy chain constant region gene.

[0040] The terms "variable region" or "variable domain" refer to the structural domains of the antibody heavy or light chain that enable the antibody to bind to the antigen. The variable regions of the heavy and light chains of natural antibodies (V1 and V2, respectively) are... H and V L The variable region (HVR) can be further subdivided into highly variable regions (or highly variable areas, which may be highly variable in the sequence and / or form of structurally defined loops), also known as complementarity determining regions (CDRs). These regions are interspersed with more conserved regions called frame regions (FRs). In the art, the terms "complementarity determining region" and "CDR," synonymous with "hypervariable region" or "HVR," refer to discontinuous amino acid sequences within the antibody variable region that confer antigen specificity and / or binding affinity. Generally, each heavy chain variable region contains three CDRs (CDR-H1, CDR-H2, CDR-H3), and each light chain variable region contains three CDRs (CDR-L1, CDR-L2, CDR-L3). In the art, "frame region" and "FR" refer to the non-CDR portions of the heavy and light chain variable regions. Generally, each full-length heavy chain variable region contains four FRs (FR-H1, FR-H2, FR-H3, and FR-H4), and each full-length light chain variable region contains four FRs (FR-L1, FR-L2, FR-L3, and FR-L4). In each V... H and V LInternally, the three CDRs and four FRs are generally arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4 (see also Chothia and Lesk). J. Mot.Biol ., 195, 901-917 (1987)).

[0041] In the context of this invention, the term "antibody" (Ab) refers to an immunoglobulin molecule, a fragment of an immunoglobulin molecule, or a derivative thereof, which has the ability to specifically bind to an antigen under normal physiological conditions and has a long half-life, such as at least about 30 minutes, at least about 45 minutes, at least about one hour (h), at least about two hours, at least about four hours, at least about eight hours, at least about 12 hours (h), about 24 hours or longer, about 48 hours or longer, about three, four, five, six, seven or more days, or any other relevant functionally defined time period (such as sufficient time to induce, promote, enhance, and / or regulate physiological responses associated with antibody-antigen binding and / or sufficient time for antibody recruitment effector function). The variable regions of the heavy and light chains of an immunoglobulin molecule contain binding domains that interact with the antigen. The constant regions of an antibody (Ab) can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (such as effector cells) and components of the complement system (such as C1q, the first component in the typical pathway of complement activation). Antibodies can also be bispecific antibodies, bifunctional antibodies, multispecific antibodies, or similar molecules.

[0042] As used herein, the term "monoclonal antibody" refers to a formulation of an antibody molecule produced recombinantly from a single primary amino acid sequence. Monoclonal antibody compositions exhibit single binding specificity and affinity for a specific epitope. Therefore, the term "human monoclonal antibody" refers to an antibody exhibiting single binding specificity, having variable and constant regions derived from human germline immunoglobulin sequences. Human monoclonal antibodies can be generated from hybridomas fused to immortalized cells, including B cells obtained from transgenic or transchromosomal nonhuman animals (such as transgenic mice) with genomes containing both human heavy-chain and light-chain transgenes.

[0043] As used herein, the term "isolated antibody" refers to an antibody that substantially does not contain other antibodies with different antigen specificities (e.g., an isolated antibody that specifically binds to PD-L1 substantially does not contain antibodies that specifically bind to antigens other than PD-L1). However, isolated antibodies that specifically bind to PD-L1 may be cross-reactive with other antigens, such as PD-L1 molecules from different species. Furthermore, isolated antibodies may be substantially free of other cellular material and / or chemicals. In one embodiment, the isolated antibody comprises an antibody conjugate attached to another reagent (e.g., a small molecule drug). In some embodiments, the isolated anti-PD-L1 antibody comprises a conjugate of an anti-PD-L1 antibody containing a small molecule drug (e.g., MMAE or MMAF).

[0044] "Human antibody" (HuMAb) refers to an antibody having variable regions, both the FR and CDR in these variable regions being derived from human germline immunoglobulin sequences. Furthermore, if the antibody contains constant regions, those constant regions are also derived from human germline immunoglobulin sequences. Human antibodies disclosed herein may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced through random or site-specific mutagenesis in vitro or through somatic mutations in vivo). However, the term "human antibody" as used herein is not intended to include antibodies in which a CDR sequence derived from the germline of another mammalian species (such as a mouse) has been grafted onto a human frame sequence. The terms "human antibody" and "fully human antibody" are used synonymously.

[0045] As used herein, “humanized antibody” refers to a genetically engineered non-human antibody containing a human antibody constant domain and a non-human variable domain modified to have a high level of sequence homology with the human variable domain. This can be achieved by transplanting the six non-human antibody complementarity-determining regions (CDRs) that together form the antigen-binding site into the homologous human receptor frame region (FR) (see WO92 / 22653 and EP0629240). To fully reconstruct the binding affinity and specificity of the parent antibody, it may be necessary to substitute frame residues from the parent antibody (i.e., the non-human antibody) into the human frame region (back-mutation). Structural homology modeling can help identify amino acid residues in the frame region that are important for the binding properties of the antibody. Therefore, a humanized antibody may contain a non-human CDR sequence (primarily a human frame region, which optionally contains one or more amino acid back mutations of a non-human amino acid sequence) and a fully human constant region. Optionally, additional amino acid modifications (which are not necessarily reversion mutations) can be applied to obtain humanized antibodies with better properties (such as affinity and biochemical properties).

[0046] As used herein, the term "chimeric antibody" refers to an antibody in which the variable region is derived from a non-human species (e.g., rodents) and the constant region is derived from a different species (such as humans). Chimeric antibodies can be generated through antibody engineering. "Antibody engineering" is a general term used for different types of modifications to antibodies, and it is a method well known to those skilled in the art. Specifically, chimeric antibodies can be generated using methods such as those described in Sambrook. et al. Chimeric antibodies are generated using standard DNA techniques described in *Molecular Cloning: A Laboratory Manual*, 1989, New York: Cold Spring Harbor Laboratory Press, Ch. 15. Therefore, chimeric antibodies can be recombinant antibodies engineered genetically or enzymatically. The generation of chimeric antibodies is within the knowledge of those skilled in the art, and therefore, chimeric antibodies generated according to this invention can be produced by methods other than those described herein. Chimeric monoclonal antibodies are developed for therapeutic applications to reduce antibody immunogenicity. They generally contain a non-human (e.g., murine) variable region specific to the antigen of interest and human constant antibody heavy and light chain domains. As used in the context of chimeric antibodies, the terms "variable region" or "variable domain" refer to the region containing both the CDR and framework regions of the immunoglobulin heavy and light chains.

[0047] "Antigen antibody" refers to an antibody that binds to an antigen. For example, anti-PD-L1 antibody is an antibody that binds to the antigen PD-L1.

[0048] An antibody's "antigen-binding portion" or "antigen-binding fragment" refers to one or more fragments of an antibody that retain the ability to specifically bind to antigens bound by the whole antibody. Examples of antibody fragments (e.g., antigen-binding fragments) include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab')2; bifunctional antibodies; linear antibodies; single-chain antibody molecules (e.g., scFv); and multispecific antibodies formed from antibody fragments. Papain digestion of an antibody produces two identical antigen-binding fragments, called "Fab" fragments, each with a single antigen-binding site; and a residual "Fc" fragment, whose name reflects its tendency to crystallize. Pepsin treatment yields the F(ab')2 fragment, which has two antigen-binding sites and is still capable of cross-linking antigens.

[0049] The "percentage (%) sequence identity" relative to a reference polypeptide sequence is defined as the percentage of amino acid residues in the candidate sequence that are identical to those in the reference polypeptide sequence after alignment, and if necessary, introducing vacancies to achieve the maximum percentage sequence identity, and without considering any conservative substitutions as part of the sequence identity. Alignments performed for the purpose of determining percentage amino acid sequence identity can be achieved in various ways within the skill of the art, such as using publicly available computer software, such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine suitable parameters for the alignment, including any algorithms required to achieve the maximum alignment across the full length of the compared sequences. For example, the % sequence identity of a given pair of amino acid sequences A, and / or for a given amino acid sequence B (which can be alternatively expressed as a given pair of amino acid sequences A, and / or for a given amino acid sequence B having or containing a certain % sequence identity) is calculated as follows:

[0050] 100 × fraction X / Y

[0051] Where X is the number of amino acid residues that are scored as a consistent match in the sequence comparison between A and B, and Y is the total number of amino acid residues in B. It should be understood that if the lengths of amino acid sequences A and B are not equal, the % sequence similarity of A to B will not be equal to the % sequence similarity of B to A.

[0052] As used herein, in the context of antibody-antigen binding, the terms "binding" or "specific binding" generally refer to the binding of antibodies to predetermined antigens, as measured by, for example, biofilm interferometry (BLI) techniques in the Octet HTX instrument using antibodies as ligands and antigens as analytes, corresponding to approximately 10 -6 M or smaller K D The combination of affinity, such as 10 -7 M or smaller, such as about 10 -8 M or smaller, such as about 10 -9 M or smaller, approximately 10 -10 M or smaller, or about 10 -11 M or even smaller, and in which the antibody corresponds to K D The K-molecule binds to the predetermined antigen with affinity. D K+ binds to non-specific antigens other than the intended antigen or closely related antigens (e.g., BSA, casein). D At least ten times lower, such as at least 100 times lower, for example at least 1,000 times lower, such as at least 10,000 times lower, for example at least 100,000 times lower. Combined K D The amount of reduction depends on the antibody's K. D Therefore, when the antibody's K D When the level is very low, the K+ that binds to the antigen... D Below the K level that binds to non-specific antigens D The amount can be at least 10,000 times (i.e., the antibody is highly specific).

[0053] As used in this article, the term "K" D "(M)" refers to the dissociation equilibrium constant of a specific antibody-antigen interaction. As used in this paper, affinity and K D It is an inverse correlation, meaning that higher affinity implies lower K. D Furthermore, lower affinity implies higher K. D .

[0054] The term "ADC" refers to an antibody-drug conjugate, which in the context of this invention refers to an anti-PD-L1 antibody conjugated to a drug moiety (e.g., MMAE or MMAF) as described in this application.

[0055] The abbreviations "vc" and "val-cit" refer to the dipeptide linker valine-citrulline.

[0056] The abbreviation VKG refers to the tripeptide linker valine-lysine-glycine.

[0057] The abbreviation "MC" refers to the extended cis-butene diimide hexanoyl group:

[0058]

[0059] The abbreviation "MP" refers to the extended cis-butene diimide propionyl group:

[0060]

[0061] As used herein, a “PEG unit” is an organic moiety comprising repeating ethylene-oxygen subunits (PEG or PEG subunits) and can be polydisperse, monodisperse, or discrete (i.e., having a discrete number of ethylene-oxygen subunits). Polydisperse PEG is a heterogeneous mixture of various sizes and molecular weights, while monodisperse PEG is generally purified from a heterogeneous mixture and thus provides a single chain length and molecular weight. Preferred PEG units comprise discrete PEG, i.e., compounds synthesized stepwise rather than via polymerization. Discrete PEG provides a single molecule with a defined and specified chain length.

[0062] The PEG unit provided herein comprises one or more polyethylene glycol chains, each comprising one or more ethylene oxide subunits covalently attached to each other. The polyethylene glycol chains may be linked together, for example, in a straight-chain, branched-chain, or star configuration. Generally, prior to incorporation into the camptothecin conjugate, at least one of the polyethylene glycol chains has an alkyl portion (i.e., an example representing R) at one end having an electrophilic group substituted for the carbamate nitrogen of the methylene carbamate unit. Generally, the terminal ethylene oxide subunits in each polyethylene glycol chain that do not participate in the covalent attachment to the remainder of the connector unit are modified with a PEG-terminated unit (generally optionally modified with a substituted alkyl group such as -CH3, CH2CH3, or CH2CH2CO2H). Preferably, the PEG unit has a single polyethylene glycol chain having 2 to 24 -CH2CH2O- subunits covalently attached in series and terminated at one end with a PEG-terminated unit.

[0063] "Cancer" refers to a broad group of diseases characterized by the uncontrolled growth of abnormal cells in the body. "Cancer" or "cancer tissue" can include tumors. Unregulated cell division and growth lead to the formation of malignant tumors that can invade adjacent tissues and metastasize to distant parts of the body via the lymphatic system or bloodstream. After metastasis, the distant tumor may be referred to as "derived from" the pre-metastatic tumor.

[0064] The term "antibody-dependent cellular cytotoxicity" (ADCC) is a mechanism that induces cell death, dependent on the interaction between antibody-coated target cells and lysogenic immune cells (also known as effector cells). These effector cells include natural killer cells, monocytes / macrophages, and neutrophils. Effector cells attach to the Fc effector domain(s) of Ig bound to the target cell via their antigen-binding sites. The antibody-coated target cell then dies due to the effector cell activity.

[0065] The term “antibody-dependent cellular phagocytosis” or ADCP refers to the process by which antibody-coated cells are internalized (whether completely or partially) by phagocytizing immune cells (such as macrophages, neutrophils, and dendritic cells) that bind to the (multiple) Fc effector domains of Ig.

[0066] The term "complement-dependent cytotoxicity" or CDC refers to a mechanism that induces cell death in which the Fc effector domains of a target-bound antibody activate a series of enzymatic reactions, ultimately forming pores in the target cell membrane. Generally, antigen-antibody complexes (such as those on antibody-coated target cells) bind to and activate complement component C1q, which in turn activates the complement cascade leading to target cell death. Complement activation can also lead to the deposition of complement components on the target cell surface, which promotes ADCC by binding to complement receptors (e.g., CR3) on leukocytes.

[0067] "Cytostatic effect" refers to the inhibition of cell proliferation. "Cytostatic inhibitor" refers to an agent that has a cytostatic effect on cells, thereby inhibiting the growth and / or expansion of specific subgroups of cells. Cytostatic inhibitors can be conjugated with antibodies or administered in combination with antibodies.

[0068] "Treatment" or "therapy" for a subject refers to any type of intervention or procedure performed on the subject, or the administration of an active agent to the subject, with the aim of reversing, alleviating, improving, inhibiting, slowing, or preventing the onset, worsening, development, severity, or recurrence of disease-related symptoms, complications, conditions, or biochemical indicators. In some implementations, the disease is cancer.

[0069] "Subject" includes any human or non-human animal. The term "non-human animal" includes, but is not limited to, vertebrates such as non-human primates, sheep, dogs, and rodents such as mice, rats, and guinea pigs. In some embodiments, the subject is a human. The terms "subject," "patient," and "individual" are used interchangeably herein.

[0070] An "effective amount," "therapeutic effective amount," or "therapeutic dose" of a drug or therapeutic agent is any amount of the drug, when used alone or in combination with another therapeutic agent, that protects a subject from the onset of disease or promotes the resolution of disease, as demonstrated by a decrease in the severity of disease symptoms, an increase in the frequency and duration of a disease symptom-free period, or prevention of injury or disability caused by the disease. The ability of a therapeutic agent to promote disease resolution can be assessed using a variety of methods known to those skilled in the art, such as assessment in human subjects during clinical trials, assessment in animal model systems predicting efficacy in humans, or assessment by analyzing the activity of the agent in in vitro assays.

[0071] For example, in the treatment of tumors, a therapeutically effective amount of an anticancer agent, relative to one or more untreated subjects, inhibits cell or tumor growth by at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, or at least about 80%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%. In some embodiments, a therapeutically effective amount of an anticancer agent inhibits cell or tumor growth by 100% in one or more treated subjects, relative to one or more untreated subjects.

[0072] In other embodiments of this disclosure, tumor regression may be observed and continue for a period of at least about 20 days, at least about 30 days, at least about 40 days, at least about 50 days, or at least about 60 days.

[0073] Therapeutic effective amounts of a drug (e.g., anti-PD-L1 antibody-drug conjugates) include “preventative effective amounts,” which are any amounts of a drug that, when administered alone or in combination with an anticancer agent to a subject at risk of developing cancer (e.g., a subject with a pre-existing condition) or experiencing recurrence of cancer, inhibit the development or recurrence of cancer. In some embodiments, preventative effective amounts completely prevent the development or recurrence of cancer. “Inhibiting” the development or recurrence of cancer means reducing the likelihood of the development or recurrence of cancer or completely preventing the development or recurrence of cancer.

[0074] As used herein, “subtherapeutic dose” means a dose of a therapeutic compound (e.g., an anti-PD-L1 antibody-drug conjugate) that is lower than the usual or typical dose of the therapeutic compound when administered alone for the treatment of hyperproliferative diseases (e.g., cancer).

[0075] "Immune-associated response pattern" refers to the clinical response pattern commonly observed in cancer patients treated with immunotherapies that induce cancer-specific immune responses or produce antitumor effects by modifying innate immune processes. This response pattern is characterized by a beneficial therapeutic effect following an initial increase in tumor burden or the appearance of new lesions (which would be classified as disease progression and synonymous with drug failure in the evaluation of conventional chemotherapy). Therefore, appropriate evaluation of immunotherapies may require long-term monitoring of the effects of these agents on the target disease.

[0076] For example, an "anticancer agent" promotes cancer regression in a subject. In some implementations, a therapeutically effective amount of the drug promotes cancer regression to the point of elimination. "Promoting cancer regression" means that the administration of an effective amount of the drug (alone or in combination with an anticancer agent) results in a reduction in tumor growth or size, tumor necrosis, a decrease in the severity of at least one disease symptom, an increase in the frequency and duration of symptom-free periods, or prevention of damage or disability due to disease. Furthermore, the terms "effective" and "effectiveness" in relation to treatment include both pharmacological effectiveness and physiological safety. Pharmacological effectiveness refers to the ability of a drug to promote cancer regression in a patient. Physiological safety refers to the level of toxicity produced by the administration of the drug or other adverse physiological effects (adverse effects) at the cellular, organ, and / or organismal level.

[0077] "Sustained response" refers to a sustained effect on reducing tumor growth after treatment has been discontinued. For example, the tumor size may remain the same or smaller compared to its size at the start of the treatment phase. In some embodiments, the sustained response has a duration at least the same as or at least 1.5, 2.0, 2.5, or 3 times longer than the duration of treatment.

[0078] As used in this article, “complete response” or “CR” means the disappearance of all target lesions; “partial response” or “PR” means that the sum of the longest diameter (SLD) of the target lesions is reduced by at least 30% relative to the baseline SLD; and “stable disease” or “SD” means that, relative to the minimum SLD since the start of treatment, the target lesions have neither shrunk sufficiently to meet the PR nor increased sufficiently to meet the PD.

[0079] As used in this article, "progression-free survival" or "PFS" refers to the length of time during and after treatment when the treated disease (e.g., cancer) does not worsen. Progression-free survival can include the amount of time a patient has experienced a complete or partial response, as well as the amount of time a patient has experienced stable disease.

[0080] As used in this article, “overall response rate” or “ORR” refers to the sum of the complete response (CR) rate and the partial response (PR) rate.

[0081] As used in this article, “overall survival” or “OS” refers to the percentage of individuals in a group who are likely to survive after a specific duration.

[0082] The phrase “pharmaceutically acceptable” indicates that a substance or composition must be chemically and / or toxicologically compatible with other ingredients contained in the formulation and / or with the mammals it treats.

[0083] As used herein, the phrase “pharmaceutically acceptable salt” refers to an organic or inorganic salt of a compound of the present invention that is pharmaceutically acceptable. Exemplary salts include, but are not limited to, sulfates, citrates, acetates, oxalates, chlorides, bromides, iodides, nitrates, bisulfates, phosphates, acid phosphates, isonicotinates, lactates, salicylates, acid citrates, tartrates, oleates, tannins, pantothenates, tartrates, ascorbic acid salts, succinates, maleate, gentianate, fumarate, gluconate, glucuronide, gluconate dihydrate, formate, benzoate, glutamate, methanesulfonate (“methanesulfonate”), ethanesulfonate, benzenesulfonate, p-toluenesulfonate, dihydroxynaphthyl acid (i.e., 4,4'-methylene-bis-(2-hydroxy-3-naphthyl acid)) salts, alkali metal (e.g., sodium and potassium) salts, alkaline earth metal (e.g., magnesium) salts, and ammonium salts. Pharmaceutically acceptable salts may involve another molecule, such as an acetate ion, a succinate ion, or other relative ions. The relative ion can be any organic or inorganic part that stabilizes the charge on the parent compound. Furthermore, pharmaceutically acceptable salts may have more than one charged atom in their structure. The case where multiple charged atoms are part of a pharmaceutically acceptable salt means that it may have multiple relative ions. Therefore, pharmaceutically acceptable salts may have one or more charged atoms and / or one or more relative ions.

[0084] "Administration" refers to the physical introduction of a therapeutic agent into a subject using any of a variety of methods and delivery systems known to a person skilled in the art. Exemplary routes of administration for anti-PD-L1 antibody-drug conjugates include intravenous, intramuscular, subcutaneous, intraperitoneal, spinal, or other parenteral administration routes, such as by injection or infusion (e.g., intravenous infusion). As used herein, the phrase "parenteral administration" means a mode of administration other than enteral and local administration, typically by injection, and including but not limited to intravenous, intramuscular, intra-arterial, intrathecal, intralymphatic, intralesional, intracystic, intraorbital, intracardiac, intradermal, intraperitoneal, tracheal, subcutaneous, subepidermal, intra-articular, subcystic, subarachnoid, spinal, epidural, and intrasternal injections and infusions, as well as intracorporeal electroporation. Therapeutic agents may be administered via non-parenteral routes or orally. Other non-parenteral routes include local, epidermal, or mucosal application, such as intranasal, vaginal, rectal, sublingual, or topical. Application may also be performed, for example, once, multiple times, and / or over one or more extended time periods.

[0085] The terms “baseline” or “baseline value” used interchangeably in this document refer to a measurement or characterization of symptoms before or at the start of treatment (e.g., an anti-PD-L1 antibody-drug conjugate as described herein). Baseline values ​​may be compared to reference values ​​to determine the reduction or improvement of symptoms in PD-L1-related diseases (e.g., cancer) covered herein. The terms “reference” or “reference value” used interchangeably in this document refer to a measurement or characterization of symptoms after treatment (e.g., an anti-PD-L1 antibody-drug conjugate as described herein). Reference values ​​may be measured once or multiple times during a dosing regimen or treatment cycle or upon completion of a dosing regimen or treatment cycle. A “reference value” may be an absolute value; a relative value; a value with an upper and / or lower limit; a series of values; an average; a median; or a value compared to a baseline value.

[0086] Similarly, a “baseline value” can be an absolute value; a relative value; a value with an upper and / or lower limit; a series of values; an average; a median; a mean; or a value compared to a reference value. Reference values ​​and / or baseline values ​​can be obtained from a single individual, from two different individuals, or from a group of individuals (e.g., a group of two, three, four, five, or more individuals).

[0087] As used herein, the term "monotherapy" means that the anti-PD-L1 antibody-drug conjugate is the only anticancer agent administered to the subject during a treatment cycle. However, other therapeutic agents may also be administered to the subject. For example, anti-inflammatory agents or other medications administered to a subject with cancer to treat cancer-related symptoms that are not the underlying cancer itself (including, for example, inflammation, pain, weight loss, and general malaise) may be administered during the period of monotherapy.

[0088] As used herein, an “adverse event” (AE) is any unfavorable and usually unintended or unwanted sign (including abnormal laboratory study results), symptom, or illness associated with the use of a medical treatment. A medical treatment may have one or more associated AEs, and these AEs may be of the same or different severity levels. The reference to methods that can “modify adverse events” means treatment regimens that reduce the incidence and / or severity of one or more AEs associated with the use of different treatment regimens.

[0089] As used herein, a “serious adverse event” or “SAE” is an adverse event that meets one of the following criteria:

[0090] For events that are fatal or life-threatening (as used in the definition of serious adverse events), “life-threatening” means an event in which the patient is at risk of death at the time of the event; it does not mean an event that, if it were more serious, could cause death.

[0091] Causes persistent or significant disability / incapacity

[0092] Constitutes congenital abnormalities / birth defects

[0093] Medically significant events are defined as events that endanger the patient or may require medical or surgical intervention to prevent one of the outcomes listed above. Medical and scientific judgment must be exercised in determining whether an adverse event (AE) is "medically significant."

[0094] Hospitalization or extension of existing hospitalization is required, excluding the following: 1) routine treatment or monitoring of underlying diseases unrelated to any deterioration of the condition; 2) selective or pre-planned treatment for a pre-existing condition unrelated to the indication in the study and which has not deteriorated since the informed consent was signed; and 3) social causes and temporary care in the absence of any deterioration in the patient’s overall condition.

[0095] It should be understood that the use of alternatives (e.g., “or”) means any one, both, or any combination of the alternatives. It should be understood that, as used herein, the indefinite article “a / an” means “one or more” of any of the stated or enumerated components.

[0096] The terms “about” or “comprising essentially of” refer to a value or composition within an acceptable margin of error for a particular value or composition, which, as determined by a person of ordinary knowledge in the art, will depend in part on how the value or composition is measured or determined, i.e., the limitations of the measurement system. For example, in accordance with practice in the art, “about” or “comprising essentially of” may mean within 1 or more standard deviations. Alternatively, “about” or “comprising essentially of” may mean a range of up to 20%. Furthermore, particularly in relation to biological systems or processes, the term may mean a value of up to an order of magnitude or up to five times. When a particular value or composition is provided in this application and the claims, unless otherwise stated, the meaning of “about” or “comprising essentially of” should be assumed to be within an acceptable margin of error for that particular value or composition.

[0097] This document's references to "(about)" a value or parameter include (and describe) implementations for that value or parameter itself. For example, a reference to "about X" encompasses and describes "X".

[0098] As used herein, the term “Combined Positive Score” or “CPS” refers to an immunohistochemical method for measuring PD-L1 expression in cancer (such as tumor samples from cancer). CPS is the number of PD-L1-stained cells (tumor cells, lymphocytes, macrophages) divided by the total number of viable tumor cells, multiplied by 100. For some therapeutic applications, a CPS ≥ 1 indicates that the tumor sample expresses PD-L1. For example, subjects with conditions such as gastric cancer, cervical cancer, and head and neck squamous cell carcinoma require a CPS ≥ 1 to be eligible for certain PD-1 or PD-L1 inhibitor therapies. In some cases, a CPS ≥ 10 is required to be eligible for certain PD-1 or PD-L1 inhibitor therapies, such as subjects with epithelial carcinoma of the urethra (bladder cancer), esophageal squamous cell carcinoma (ESCC), or triple-negative breast cancer who are being treated with pembrolizumab.

[0099] As used herein, the term “Tumor Proportion Score” or “TPS” refers to an immunohistochemical method that measures PD-L1 expression in cancer (such as tumor samples from cancer). TPS is the percentage of live tumor cells that show partial or complete membrane staining at any intensity. For some therapeutic treatments, a tumor sample is considered to have PD-L1 expression if TPS ≥ 1%, and a tumor sample is considered to have high PD-L1 expression if TPS ≥ 50%. For example, subjects (such as those with non-small cell lung cancer) need a TPS ≥ 1% to be eligible for certain PD-1 or PD-L1 inhibitor therapies (such as pembrolizumab). In some cases, subjects need a TPS ≥ 50% to be eligible for certain PD-1 or PD-L1 inhibitor therapies (such as cimiprilmab).

[0100] As used herein, the term "ideal body weight" or "IBW" refers to a dimensional description independent of total weight. IBW is an estimate of weight adjusted for sex, height, and optionally, bone structure. IBW can be calculated, for example, using the formulas IBW = 0.9H - 88 (for men) and IBW = 0.9H - 92 (for women), where H = height (in cm). Alternatively, IBW can also be calculated, for example, using the following formulas: IBW (men) = 50 kg + 2.3 kg × (height (inches) - 60); IBW (women) = 45.5 kg + 2.3 kg × (height (inches) - 60).

[0101] As used in this article, the term "adjusted ideal body weight" or "AIBW" refers to a size description that takes into account sex, total weight, and height. For example, AIBW can be calculated using the formula AIBW = IBW + 0.4 (weight (in kg) - IBW).

[0102] As stated herein, unless otherwise indicated, any concentration range, percentage range, ratio range, or integer range shall be understood to include any integer value within the range and (where appropriate) its fraction (such as one-tenth and one-hundredth of an integer).

[0103] The various aspects of this disclosure are described in further detail in the following subsections.

[0104] II. Overview

[0105] This document provides methods for treating various cancers in a subject using an antibody-drug conjugate (ADC) that binds to PD-L1. In one aspect, this document provides methods for treating a subject's cancer using an ADC that binds to PD-L1 and an anti-PD-1 antibody (e.g., pembrolizumab). In some embodiments, the cancer is a solid tumor. In some embodiments, the cancer is head and neck cancer. In a further embodiment, the cancer is head and neck squamous cell carcinoma (HNSCC). In a further embodiment, the cancer is metastatic or unresectable HNSCC. In some embodiments, the cancer is breast cancer. In a further embodiment, the cancer is triple-negative breast cancer (TNBC). In some embodiments, the cancer is lung cancer. In a further embodiment, the cancer is non-small cell lung cancer (NSCLC). In some embodiments, the cancer is esophageal cancer. In a further embodiment, the cancer is esophageal squamous cell carcinoma (SCC). In some embodiments, the cancer is melanoma. In some embodiments, the cancer is ovarian cancer. In some embodiments, the cancer is gastric cancer.

[0106] Pembrolizumab targets the immune checkpoint receptor PD-1 and blocks PD-L1 interaction. In the tumor microenvironment, T cells may lose activity through the binding of PD-1 (expressed on T cells) to PD-L1 (expressed on tumor cells). However, in the presence of unconjugated anti-PD-L1 antibodies, the interaction between PD-1 and PD-L1 is disrupted, leading to anti-tumor T cell activity.

[0107] The combination of an ADC that binds to PD-L1 on the cell surface with a PD-1 checkpoint inhibitor (such as pembrolizumab) offers unique advantages. In one embodiment, the ADC of the present invention comprises an anti-PD-L1 antibody conjugated to one or more units of monomethylolpropionate E (MMAE). In a further embodiment, this ADC induces direct cytotoxicity against PD-L1-positive cells, which reduces overall immunosuppressive PD-1 / PD-L1 signaling in the tumor microenvironment and thus drives increased activity of cytotoxic T cells. In another embodiment, this ADC can also initiate cancer immune processes through MMAE-induced immunogenic cell death.

[0108] III. Target molecule

[0109] Unless otherwise indicated, PD-L1 refers to human PD-L1. An exemplary human protein sequence is assigned UniProtID NO. Q9NZQ7.

[0110] IV. The antibody of the present invention

[0111] Previously, selected antibodies for cancer treatment have been conjugated with cytotoxic agents without sequence modification to produce antibody-drug conjugates (ADCs). These ADCs have generally proven to be as effective as or more effective than unconjugated antibodies in killing tumor cells. Previously, if antibody modification was considered during ADC preparation, some possible modifications could increase antibody binding affinity or enhance antibody activity, such as ADCC. However, it has been found that, at least in some cases, modifying or tuning the ADC antibody by, for example, reducing its binding affinity or ADCC activity, leads to improved ADC efficacy compared to ADCs with unmodified antibodies. Some examples of this include ADCs with anti-PD-L1 antibodies (such as Ab1), which were unexpectedly optimized by modifying the antibody (e.g., reducing its binding affinity). For example, in some cases, anti-PD-L1 ADCs were more effective at killing tumor cells in vitro when the binding affinity of the antibody conjugated to the cytotoxic agent was reduced. In another embodiment, in some cases, anti-PD-L1 ADCs are more effective at killing tumor cells in vitro and in vivo when the binding affinity of the antibody conjugated to the cytotoxic agent is reduced.

[0112] This invention provides antibodies that bind to PD-L1, such as humanized antibodies. In some embodiments, the anti-PD-L1 antibody provided herein may have the heavy chain CDR1 of SEQ ID NO: 5, the heavy chain CDR2 of SEQ ID NO: 6, the heavy chain CDR3 of SEQ ID NO: 7, the light chain CDR1 of SEQ ID NO: 8, the light chain CDR2 of SEQ ID NO: 9, and the light chain CDR3 of SEQ ID NO: 10.

[0113] The anti-PD-L1 antibody of the present invention may also be described or specified in terms of its binding affinity to PD-L1 (e.g., human PD-L1). In some embodiments, the preferred binding affinity includes having a dissociation constant or K0. D : Greater than 2.7nM, 5 nM, 6 nM, 7 nM, 8 nM, 9 nM, 10 nM, 15 nM, 20 nM, 25 nM, 30 nM, 40 nM, 50 nM, 60 nM, 70 nM, 80 nM, 90 nM, 100 nM, 110 nM, 120 nM, 130 nM, 140 nM, 150 nM, 200 nM, 250 nM, 300nM, 400 nM, or 500 nM. In some embodiments, the preferred PD-L1 antibody has binding affinity between 3 nM and 300 nM, 3 nM and 200 nM, 3 nM and 100 nM, 3 nM and 50 nM, 3 nM and 40 nM, 3 nM and 20 nM, 3 nM and 15 nM, 5 nM and 300 nM, and 5 nM and 15 nM. In some embodiments, the preferred PD-L1 antibody has binding affinity at least 2, 3, 3.7, 4, or 5 times greater than that of Ab1. In some embodiments, the binding affinity is a monovalent binding affinity.

[0114] In some embodiments, the binding of the anti-PD-L1 antibody of the present invention is pH-dependent, such that the antibody exhibits differential binding across a pH gradient. In some embodiments, the anti-PD-L1 antibody exhibits maximum binding between a pH of about 4 and a pH of about 10. In some embodiments, maximum binding is between a pH of about 6 and a pH of about 9. In some embodiments, maximum binding is between a pH of about 6.5 and a pH of about 8.

[0115] In animal models or clinical trials, the preferred antibodies of the present invention inhibit cancer (e.g., cell growth, metastasis to the organism, and / or lethality to the organism), as demonstrated for cancer cells proliferating in cultures. Animal models can be established by implanting PD-L1-expressing human tumor cell lines into appropriate immunodeficient rodent strains (e.g., athymic nude mice or SCID mice). These tumor cell lines can be established in immunodeficient rodent hosts as solid tumors via subcutaneous injection or as diffuse tumors via intravenous injection.

[0116] Once established in the host, these tumor models can be used to evaluate the therapeutic efficacy of anti-PD-L1 antibodies or their conjugate forms, as described in the examples.

[0117] The anti-PD-L1 antibody disclosed herein is preferably a monoclonal antibody and may be a multispecific antibody, human antibody, humanized antibody or chimeric antibody, single-chain antibody, Fab fragment, F(ab') fragment, fragment generated from a Fab expression library, and any of the above-described PD-L1 binding fragments. In some embodiments, the anti-PD-L1 antibody of this disclosure specifically binds to PD-L1. The immunoglobulin molecules of this disclosure may belong to any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass of immunoglobulin molecules. In some embodiments, the anti-PD-L1 antibody of this disclosure belongs to the IgG1 type.

[0118] In some embodiments of this disclosure, the anti-PD-L1 antibody is an antigen-binding fragment as described herein (e.g., a human antigen-binding fragment), and includes, but is not limited to, Fab, Fab' and F(ab')2, Fd, single-chain Fvs (scFv), single-chain antibodies, disulfide-linked Fvs (sdFv), and those containing V L or V H Fragments of structural domains. Antigen-binding fragments (including single-chain antibodies) may comprise individual (multiple) variable regions or combinations thereof with all or some of the following: hinge region, CH1, CH2, CH3, and CL domains. This disclosure also includes antigen-binding fragments comprising any combination of variable regions and hinge regions, CH1, CH2, CH3, and CL domains. In some embodiments, the anti-PD-L1 antibody or its antigen-binding fragment is human, rodent (e.g., mouse and rat), donkey, sheep, rabbit, goat, guinea pig, camel, horse, or chicken.

[0119] The anti-PD-L1 antibody disclosed herein may be monospecific, bispecific, trispecific, or multispecific. Multispecific antibodies may be specific to different epitopes of PD-L1, or specific to both PD-L1 and heterologous proteins. See, for example, PCT disclosures WO 93 / 17715; WO 92 / 08802; WO 91 / 00360; WO 92 / 05793; Tutt. et al. , 1991, J. Immunol. 147:60 69; U.S. Patent Nos. 4,474,893; 4,714,681; 4,925,648; 5,573,920; 5,601,819; Kostelny et al. , 1992, J. Immunol. 148:1547 1553.

[0120] The anti-PD-L1 antibody disclosed herein can be described or specified in terms of its specific CDRs. The precise amino acid sequence boundaries of a given CDR or FR can be readily determined using any of a variety of well-known schemes, including the Kabat method described below. et al. (1991), “Sequences of Proteins of Immunological Interest,” 5thEd. Public Health Service, National Institutes of Health, Bethesda, MD (“Kabat” numbering scheme); Al-Lazikani et al. (1997) JMB 273,927-948 (“Chothia” numbering scheme); MacCallum et al. , J. Mol. Biol. 262:732-745 (1996), "Antibody-antigeninteractions: Contact analysis and binding site topography," J. Mol. Biol. 262, 732-745." ("Contact" numbering scheme); Lefranc MP et al., "IMGT unique numbering for immunoglobulin and T cell receptor variable domains and Ig superfamily V-like domains," Dev Comp Immunol, 2003 Jan;27(1):55-77 ("IMGT" numbering scheme); Honegger A and Plückthun A, "Yet another numbering scheme for immunoglobulin variable domains: an automatic modeling and analysis tool," J Mol Biol, 2001Jun 8;309(3):657-70, ("Aho" numbering plan); and Martin et al. , “Modeling antibody hypervariable loops: a combined algorithm,” PNAS, 1989, 86(23):9268-9272 (“AbM” numbering scheme). The boundaries of a given CDR may vary depending on the scheme used for identification. In some implementations, the “CDR” or “complementarity determination region” or individually designated CDRs (e.g., CDR-H1, CDR-H2, CDR-H3) of a given antibody or its region (e.g., its variable region) should be understood to encompass CDRs as defined (or designated) by any of the foregoing schemes. For example, stating that a particular CDR (e.g., CDR-H3) contains a given V H or V L When specifying the amino acid sequence of the corresponding CDR in the region's amino acid sequence, it should be understood that such CDR has a sequence like the corresponding CDR (e.g., CDR-H3) within the variable region as defined by any of the aforementioned schemes. Schemes for identifying a particular CDR can be specified, such as CDRs defined by the Kabat, Chothia, AbM, or IMGT methods.

[0121] The CDR sequences of the anti-PD-L1 antibody and anti-PD-L1 antibody-drug conjugate described in this article are based on, for example, Kabat. et al. The Kabat numbering scheme described in (1991), “Sequences of Proteins of Immunological Interest,” 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD.

[0122] In one aspect, this document provides an anti-PD-L1 antibody and / or an anti-PD-L1 antibody-drug conjugate comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 5, (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO: 6, and (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO: 7; and / or wherein the light chain variable region comprises (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 8, (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO: 9, and (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO: 10, wherein the CDR of the anti-PD-L1 antibody is defined by the Kabat numbering scheme.

[0123] In one aspect, this document provides an anti-PD-L1 antibody and / or an anti-PD-L1 antibody-drug conjugate comprising a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises the amino acid sequence of SEQ ID NO: 3 and the light chain variable domain comprises the amino acid sequence of SEQ ID NO: 4. In another aspect, this document provides an anti-PD-L1 antibody and / or an anti-PD-L1 antibody-drug conjugate comprising a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 1 and the light chain comprises the amino acid sequence of SEQ ID NO: 2.

[0124] In some embodiments, this document provides anti-PD-L1 antibodies and / or anti-PD-L1 antibody-drug conjugates comprising a heavy chain variable domain, the heavy chain variable domain comprising an amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence of SEQ ID NO: 3. In some embodiments, the heavy chain variable domain comprising an amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence of SEQ ID NO: 3 contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to a reference sequence and retains the ability to bind to PD-L1 (e.g., human PD-L1). In some embodiments, a total of 1 to 10 amino acids in SEQ ID NO: 3 have been substituted, inserted, and / or deleted. In some embodiments, substitutions, insertions, or deletions (e.g., 1, 2, 3, 4, or 5 amino acids) occur in regions outside the CDR (i.e., in the FR). In some embodiments, the anti-PD-L1 antibody comprises: a post-translational modified heavy chain variable domain sequence including SEQ ID NO:3.

[0125] In some embodiments, this document provides anti-PD-L1 antibodies and / or anti-PD-L1 antibody-drug conjugates comprising a light chain variable domain, the light chain variable domain comprising an amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence of SEQ ID NO: 4. In some embodiments, the light chain variable domain comprising an amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence of SEQ ID NO: 4 contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to a reference sequence and retains the ability to bind to PD-L1 (e.g., human PD-L1). In some embodiments, a total of 1 to 10 amino acids in SEQ ID NO: 4 have been substituted, inserted, and / or deleted. In some embodiments, substitutions, insertions, or deletions (e.g., 1, 2, 3, 4, or 5 amino acids) occur in regions outside the CDR (i.e., in the FR). In some embodiments, the anti-PD-L1 antibody comprises: a post-translational modified light chain variable domain sequence including SEQ ID NO:4.

[0126] In some implementations, the anti-PD-L1 antibody or anti-PD-L1 antibody-drug conjugate is a monoclonal antibody.

[0127] There are five classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, each with a heavy chain designated α, δ, ε, γ, and μ, respectively. The γ and α classes are further subdivided into subclasses; for example, humans express the following subclasses: IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. IgG1 antibodies can exist in various polymorphic variants (called allotypes) (reviewed in Jefferis and Lefranc 2009). mAbs Vol 1, Issue 4, pp. 1-7), any of which apply to some embodiments herein. Common allotypes in the human population are designated by the letters a, f, n, z, or combinations thereof. In any embodiment herein, the antibody may comprise a heavy chain Fc region containing a human IgG Fc region. In a further embodiment, the human IgG Fc region contains human IgG1.

[0128] Antibodies also include modified derivatives, which are antibodies covalently attached to any type of molecule such that covalent attachment does not impede the antibody's binding to PD-L1 or its ability to exert cell growth inhibition or cytotoxic effects. For example (but not in a limiting manner), antibody derivatives include antibodies that have been modified, for example, by: glycosylation, acetylation, polyethylene glycolation, phosphorylation, amidation, derivatization by known protecting / capping groups, proteolytic cleavage, or binding to cellular ligands or other proteins. Any of these chemical modifications can be performed using known techniques, including but not limited to specific chemical cleavage, acetylation, formylation, and the metabolic synthesis of tunicamycin. Additionally, derivatives may contain one or more atypical amino acids.

[0129] Humanized antibodies

[0130] Humanized antibodies are genetically engineered antibodies in which a CDR (Cellular Receptor Derivative) from a non-human "donor" antibody is grafted into a human "recipient" antibody sequence (see, for example, Queen, US 5,530,101 and 5,585,089; Winter, US 5,225,539; Carter, US 6,407,213; Adair, US 5,859,205; and Foote, US 6,881,557). The recipient antibody sequence can be, for example, a mature human antibody sequence, a complex of such sequences, a common sequence of human antibody sequences, or a germline region sequence. For heavy chains, a preferred recipient sequence is germline V. H Exon V Hl-2 (also referred to as HV1-2 in the literature) (Shin et al, 1991, EMBO J. 10:3641-3645), while for the hinge region (J H Then it is exon J. H -6 (Mattila et al, 1995, Eur. J. Immunol. 25:2578-2582). For the light chain, the preferred acceptor sequence is exon VK2-30 (also referred to as KV2-30 in the literature), while for the hinge region it is exon JK-4 (Hieter et al, 1982, J. Biol. Chem. 257:1516-1522). Therefore, a humanized antibody is an antibody having some or all of the CDRs that are wholly or substantially derived from the donor antibody, and a variable region framework sequence and a constant region (if present) that are wholly or substantially derived from the human antibody sequence. Similarly, a humanized heavy chain has at least one, two, and usually all three CDRs that are wholly or substantially derived from the donor antibody heavy chain, and a heavy chain variable region framework sequence and a heavy chain constant region (if present) that are substantially derived from the human heavy chain variable region framework and constant region sequences. Similarly, the humanized light chain has at least one, two, and usually all three CDRs that are wholly or substantially derived from the donor antibody light chain, and a light chain variable region framework sequence and light chain constant region (if present) that are substantially derived from the human light chain variable region framework and constant region sequences. Except for nanobodies and dAbs, humanized antibodies comprise humanized heavy chains and humanized light chains. When at least 60%, 85%, 90%, 95%, or 100% of the corresponding residues (as defined by Kabat) are identical among the corresponding CDRs, the CDRs in the humanized antibody are substantially derived from the corresponding CDRs in the non-human antibody. When at least 85%, 90%, 95%, or 100% of the corresponding residues as defined by Kabat are identical, the variable region framework sequence of the antibody chain or the constant region of the antibody chain is substantially derived from the human variable region framework sequence or the human constant region, respectively. In some embodiments, the PD-L1 antibody of the present invention is a humanized antibody.

[0131] Although humanized antibodies are typically incorporated into all six CDRs of mouse antibodies (preferably as defined by Kabat), they can also be prepared with fewer CDRs than all CDRs of mouse antibodies (e.g., at least 3, 4, or 5) (e.g., Pascalis et al., J. Immunol. 169:3076, 2002; Vajdos et al., Journal of Molecular Biology, 320: 415-428, 2002; Iwahashi et al., Mol. Immunol. 36:1079-1091, 1999; Tamura et al., Journal of Immunology, 164:1432-1441, 2000).

[0132] Selection of constant region

[0133] The variable regions of the heavy and light chains of humanized antibodies may be linked to at least a portion of the human constant regions. The choice of the constant regions may depend in part on whether antibody-dependent cell-mediated cytotoxicity, antibody-dependent phagocytosis, and / or complement-dependent cytotoxicity are required. For example, human isotypes IgG1 and IgG3 have strong complement-dependent cytotoxicity, human isotype IgG2 has weak complement-dependent cytotoxicity, and human IgG4 lacks complement-dependent cytotoxicity. Human IgG1 and IgG3 also induce stronger cell-mediated effector functions than human IgG2 and IgG4. The light chain constant region may be λ or κ. Antibodies may be expressed as tetramers containing two light chains and two heavy chains, as single heavy chains or light chains, as Fab, Fab', F(ab')2, and Fv, or as single-chain antibodies, wherein the variable domains of the heavy and light chains are linked by spacers.

[0134] Human constant regions exhibit allotropic and isoallotypic variations among different individuals, meaning that the location of a constant region at one or more polymorphic sites can differ among individuals. The difference between isoallotypic and isoallotypic is that serum identifying isoallotypic will bind to one or more non-polymorphic regions of other isotypes.

[0135] One or more amino acids at the amino or carboxyl terminus of the light and / or heavy chains (such as the C-terminal lysine of the heavy chain) may be omitted or derivatized in a certain proportion or in all molecules. Substitutions may be made in the constant region to reduce or increase effector functions such as complement-mediated cytotoxicity or ADCC (see, for example, Winter et al., U.S. Patent No. 5,624,821; Tso et al., U.S. Patent No. 5,834,597; and Lazar et al., Proc. Natl. Acad. Sci. USA 103:4005, 2006), or to prolong the half-life in humans (see, for example, Hinton et al., J. Biol. Chem. 279:6213, 2004).

[0136] Exemplary substitutions include amino acid substitutions of natural amino acids to introduce cysteine ​​residues at amino acid positions 234, 235, 237, 239, 267, 298, 299, 326, 330, or 332, preferably the S239C mutation in the human IgG1 isotype (US 20100158909). The presence of additional cysteine ​​residues allows for interchain disulfide bond formation. Such interchain disulfide bond formation can cause steric hindrance, thereby reducing the affinity of the Fc region-FcyR binding interaction. Multiple cysteine ​​residues introduced in or near the Fc region of the IgG constant region can also serve as sites for conjugation with therapeutic agents (i.e., using thiol-specific agents of the drug (such as maleic anhydride derivatives) to couple cytotoxic drugs). The presence of the therapeutic agent causes steric hindrance, thereby further reducing the affinity of the Fc region-FcyR binding interaction. Other substitutions at any of positions 234, 235, 236 and / or 237 reduce affinity for Fey receptors (particularly FcyRI receptors) (see, for example, US 6,624,821, US 5,624,821).

[0137] The in vivo half-life of an antibody can also affect its effector function. Increasing or decreasing the half-life of an antibody can alter its therapeutic activity. FcRn is a receptor structurally similar to a MHC class I antigen that is non-covalently associated with β2-microglobulin protein. FcRn regulates the catabolism of IgG and its trans-tissue endocytosis (Ghetie and Ward, 2000, Annu. Rev. Immunol. 18:739-766; Ghetie and Ward, 2002, Immunol. Res. 25:97-113). IgG-FcRn interaction occurs at pH 6.0 (the pH of intracellular vesicles) rather than at pH 7.4 (the pH of blood); this interaction allows IgG to be recycled back into circulation (Ghetie and Ward, 2000, Ann. Rev. Immunol. 18:739-766; Ghetie and Ward, 2002, Immunol. Res. 25:97-113). Regions on human IgG1 involved in FcRn binding have been localized (Shields et al, 2001, J. Biol. Chem. 276:6591-604). Alanine substitutions at positions Pro238, Thr256, Thr307, Gln311, Asp312, Glu380, Glu382, or Asn434 in human IgG1 enhance FcRn binding (Shields et al, 2001, J. Biol. Chem. 276:6591-604). IgG1 molecules with these substitutions have a longer serum half-life. Therefore, these modified IgG1 molecules may be able to perform their effector functions for a longer period of time compared to unmodified IgG1, and thus exert their therapeutic efficacy. Other exemplary substitutions for increasing FcRn binding include Gin at position 250 and / or Leu at position 428. All positions in the constant region are numbered using EU.

[0138] Oligosaccharides covalently attached to conserved Asn297 are involved in the ability of IgG to bind FcyR to the Fc region (Lund et al, 1996, J. Immunol. 157:4963-69; Wright and Morrison, 199', Trends Biotechnol. 15:26-31). Engineering of this glycoform of IgG can significantly improve IgG-mediated ADCC. Adding bipartite N-acetylglucosamine modification (Umana et al, 1999, Nat. Biotechnol. 17:176-180; Davies et al, 2001, Biotech. Bioeng. 74:288-94) to this glycoform or removing fucose from this glycoform (Shields et al, 2002, J. Biol. Chem. 277:26733-40; Shinkawa et al, 2003, J. Biol. Chem. 278:6591-604; Niwa et al., 2004, Cancer Res. 64:2127-33) are two examples of IgG Fc engineering that improves the binding between IgG Fc and FcyR, thereby enhancing Ig-mediated ADCC activity.

[0139] Systematic substitution of solvent-exposed amino acids in the Fc region of human IgG1 has generated IgG variants with altered FcyR binding affinity (Shields et al, 2001, J. Biol. Chem. 276:6591-604). Subsets of these variants involving substitutions to Ala at Thr256 / Ser298, Ser298 / Glu333, Ser298 / Lys334, or Ser298 / Glu333Lys334, when compared to parental IgG1, exhibit increased FcyR binding affinity and ADCC activity (Shields et al, 2001, J. Biol. Chem. 276:6591-604; Okazaki et al, 2004, J. Mol. Biol. 336:1239-49).

[0140] The complement-binding activity of antibodies (both C1q binding and CDC activity) can be improved by substitutions at Lys326 and Glu333 (Idusogie et al., 2001, J. Immunol. 166:2571-2575). The same substitutions on the human IgG2 backbone can transform antibody isotypes that are poorly bound to C1q and severely lack complement activation activity into antibody isotypes that can bind to both C1q and mediate CDC (Idusogie et al., 2001, J. Immunol. 166:2571-75). Several other methods have also been applied to improve antibody complement-binding activity. For example, transplanting an 18-amino acid C-terminal fragment of IgM to the C-terminus of IgG greatly enhances its CDC activity. This has been observed even in the case of IgG4, which typically lacks detectable CDC activity (Smith et al., 1995, J. Immunol. 154:2226-36). Furthermore, replacing Ser444, located near the C-terminus of the IgG1 heavy chain, with Cys induces tail-to-tail dimerization of IgG1, resulting in a 200-fold increase in CDC activity compared to monomeric IgG1 (Shopes et al, 1992, J. Immunol. 148:2918-22). Additionally, bispecific bifunctional antibody constructs specific to C1q also confer CDC activity (Kontermann et al., 1997, Nat. Biotech. 15:629-31).

[0141] Complement activity can be reduced by mutating at least one of the amino acid residues 318, 320, and 322 of the heavy chain to a residue with a different side chain (such as Ala). Substituting any one of the three residues with other alkyl-substituted nonionic residues (such as Gly, Leu, or Val) or aromatic nonpolar residues (such as Phe, Tyr, Trp, and Pro) also reduces or eliminates C1q binding. Ser, Thr, Cys, and Met can be used at residues 320 and 322 (but not 318) to reduce or eliminate C1q binding activity.

[0142] C1q binding activity can be modulated but not eliminated by replacing the 318 (Glu) residue with a polar residue. Replacing the 297 (Asn) residue with Ala results in the removal of dissolution activity but only a slight (approximately three-fold) reduction in affinity for C1q. This alteration disrupts the presence of the carbohydrate required for glycosylation site and complement activation. Any other substitution at this site also disrupts the glycosylation site. The following mutations and any combination thereof also reduce C1q binding: D270A, K322A, P329A, and P31IS (see WO 06 / 036291). The L234A / L235A mutation (or the LALA mutation) also reduces C1q binding and FcyR binding. In embodiments, the anti-PD-L1 antibody of the present invention comprises the L234A / L235A mutation.

[0143] The term "human constant region" refers to any arrangement of residues having any natural allotype or occupying a polymorphic position in a natural allotype. Furthermore, up to 1, 2, 5, or 10 mutations (such as those indicated above) relative to the natural human constant region may exist to reduce Fcγ receptor binding or increase binding to FcRN.

[0144] V. Expression of recombinant antibodies

[0145] Humanized antibodies are typically produced through recombinant expression. Recombinant polynucleotide constructs generally include an expression control sequence operatively linked to the coding sequence of the antibody chain, including a naturally related or heterologous promoter region. Preferably, the expression control sequence is a eukaryotic promoter system within a vector capable of transforming or transfecting eukaryotic host cells. Once the vector has been incorporated into a suitable host, the host is maintained at a high level of expression suitable for that nucleotide sequence, and then the cross-reactive antibody is collected and purified.

[0146] Mammalian cells are preferred hosts for expressing nucleotide chains encoding immunoglobulins or fragments thereof. See Winnacker, From Genes to Clones, (VCH Publishers, NY, 1987). A variety of suitable host cell lines capable of secreting intact heterologous proteins have been developed in this field, including CHO cell lines (e.g., DG44), various COS cell lines, HeLa cells, HEK293 cells, L cells, and non-antibody-producing myelomas (including Sp2 / 0 and NSO). Preferably, the cells are non-human. Expression vectors for these cells may include expression control sequences such as origin of replication, promoters, enhancers (Queen et al., Immunol. Rev. 89:49(1986)); and essential processing information sites such as ribosome binding sites, RNA splicing sites, polyadenylation sites, and transcription terminator sequences. Preferred expression control sequences are promoters derived from endogenous genes, cytomegaloviruses, SV40, adenoviruses, bovine papillomaviruses, etc. See Co et al., J. Immunol. 148:1149 (1992).

[0147] Once expressed, the antibody can be purified according to standard procedures in the relevant technical field, including HPLC purification, column chromatography, gel electrophoresis, etc. (see Scopes, Protein Purification (Springer-Verlag, NY, 1982) for general information).

[0148] VI. Nucleic Acids

[0149] This invention further provides nucleic acids encoding any humanized heavy and light chains as described above. Generally, the nucleic acid also encodes a signal peptide fused to the mature heavy and light chains. The coding sequence on the nucleic acid can be operatively linked to regulatory sequences (such as promoters, enhancers, ribosome binding sites, transcription termination signals, etc.) to ensure expression of the coding sequence. The nucleic acid encoding the heavy and light chains can be present in isolated form or can be cloned into one or more vectors. The nucleic acid can be synthesized, for example, by solid-state synthesis or by PCR of overlapping oligonucleotides. The nucleic acids encoding the heavy and light chains can be, for example, linked as a single continuous nucleic acid within an expression vector, or can be separate, for example, each cloned into its own expression vector.

[0150] In some respects, this document also provides nucleic acids encoding anti-PD-L1 antibodies or antigen-binding fragments thereof as described herein. This document further provides vectors comprising nucleic acids encoding anti-PD-L1 antibodies or antigen-binding fragments thereof as described herein. This document further provides host cells expressing nucleic acids encoding anti-PD-L1 antibodies or antigen-binding fragments thereof as described herein. This document further provides host cells comprising vectors containing nucleic acids encoding anti-PD-L1 antibodies or antigen-binding fragments thereof as described herein.

[0151] The anti-PD-L1 antibody described herein can be prepared using well-known recombinant technologies, well-known expression vector systems, and host cells. In one embodiment, the antibody is prepared in CHO cells using the following disclosed GS expression vector system: De la Cruz Edmunds et al. , 2006, Molecular Biotechnology 34; 179-190, EP216846, U.S. Patent No. 5,981,216, WO 87 / 04462, EP323997, U.S. Patent No. 5,591,639, U.S. Patent No. 5,658,759, EP338841, U.S. Patent No. 5,879,936 and U.S. Patent No. 5,891,693.

[0152] The monoclonal anti-PD-L1 antibody described herein can, for example, be produced by Kohler. et al. , Nature The monoclonal antibody can be produced by the hybridoma method described in, 256, 495 (1975), or by a recombinant DNA method. Monoclonal antibodies can also be used, for example, those described in Clackson. et al. , Nature , 352, 624-628 (1991) and Marks et al., JMol, Biol The technique described in ., 222(3):581-597 (1991) is isolated from a phage antibody library. Monoclonal antibodies can be obtained from any suitable source. Thus, for example, monoclonal antibodies can be obtained from hybridomas prepared from mouse spleen B cells obtained from mice immunized with an antigen of interest (e.g., cells expressing the antigen on their surface or in the form of nucleic acids encoding the antigen of interest). Monoclonal antibodies can also be obtained from hybridomas derived from antibody-expressing cells of immunized human or non-human mammals (such as rats, dogs, primates, etc.).

[0153] Antibody-drug conjugates

[0154] Anti-PD-L1 antibodies can be conjugated with cytotoxic or cell growth-inhibiting moieties (including their pharmaceutically compatible salts) to form antibody-drug conjugates (ADCs). Particularly suitable moieties for conjugation with antibodies are cytotoxic agents (e.g., chemotherapeutic agents), prodrug-converting enzymes, radioisotopes or compounds, or toxins (these moieties are collectively referred to as therapeutic agents). For example, anti-PD-L1 antibodies can be conjugated with cytotoxic agents (such as chemotherapeutic agents) or toxins (e.g., cell growth inhibitors or cytotoxic agents, such as abrin, ricin A, Pseudomonas exotoxin, or diphtheria toxin).

[0155] Anti-PD-L1 antibodies can be conjugated to prodrug-converting enzymes. These enzymes can be fused to the antibody recombinantly or chemically conjugated using known methods. Exemplary prodrug-converting enzymes include carboxypeptidase G2, β-glucuronidase, penicillin-V-amidase, penicillin-G-amidase, β-lactamase, β-glucosidase, nitroreductase, and carboxypeptidase A.

[0156] The techniques for conjugating therapeutic agents with proteins (and specifically, with antibodies) are well known. (See, e.g., Arnon et al, "Monoclonal Antibodies For Immunotargeting Of Drugs In CancerTherapy," in Monoclonal Antibodies And Cancer Therapy (Reisfeld et al. eds., Alan R. Liss, Inc., 1985); Hellstrom et al, "Antibodies For Drug Delivery," in Controlled Drug Delivery (Robinson et al. eds., Marcel Dekker, Inc., 2nd ed.1987); Thorpe, "Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: AReview," in Monoclonal Antibodies '84: Biological And Clinical Applications (Pinchera et al. eds., 1985); "Analysis, Results, and Future Prospective of the Therapeutic Use of Radiolabeled Antibody In Cancer Therapy," in Monoclonal Antibodies For Cancer Detection And Therapy (Baldwin et al. See also, for example, PCT Publication WO 89 / 12624).

[0157] Therapeutic agents can be conjugated in a manner that reduces their activity unless they are cleaved from the antibody (e.g., by hydrolysis, antibody degradation, or by a cleaving agent). Such therapeutic agents are attached to antibodies with cleavable adapters that are sensitive to cleavage in the intracellular environment of PD-L1-expressing cancer cells but substantially insensitive to the extracellular environment, such that the conjugate is self-cleaved by the antibody when it is internalized by PD-L1-expressing cancer cells (e.g., in endosomes or, for example, in a lysosomal or caveolear environment by means of pH sensitivity or protease sensitivity).

[0158] A typical ADC contains a linker region between the therapeutic agent and the anti-PD-L1 antibody. As described above, the linker is generally cleavable under intracellular conditions, such that cleavage of the linker in an intracellular environment (e.g., within lysosomes, endosomes, or pits) releases the therapeutic agent from the antibody. The linker may be a peptide linker that is cleaved, for example, by intracellular peptidases or proteases (including lysosomal or endosomal proteases). Generally, the peptide linker is at least two amino acids long or at least three amino acids long. Cleavage agents may include cathepsins B and D and plasminogen lysins (see, for example, Dubowchik and Walker, 1999, Pharm. Therapeutics 83:67-123). The most typical are peptide linkers that are cleavable by enzymes present in PD-L1-expressing cells. For example, a peptide linker that is cleavable by cathepsin-B, a thiol-dependent protease highly expressed in cancerous tissue (e.g., a linker containing Phe-Leu or Gly-Phe-Leu-Gly peptides) may be used. Other such linkers are described, for example, in U.S. Patent No. 6,214,345. In certain embodiments, the peptide linker cleavable by intracellular proteases comprises a Val-Cit linker or a Phe-Lys dipeptide (see, for example, U.S. Patent No. 6,214,345, which describes the synthesis of doxorubicin with a Val-Cit linker). One advantage of using intracellular proteolytically hydrolyzed radioactive agents is that the agent is generally attenuated upon conjugation and the serum stability of the conjugate is generally high.

[0159] Cleavable adapters can be pH-sensitive, meaning they are sensitive to hydrolysis at certain pH values. Generally, pH-sensitive adapters are hydrolyzable under acidic conditions. For example, acid-labile adapters that are hydrolyzable in lysosomes (e.g., hydrazones, ureas, thioureas, cis-aconitic amides, orthoesters, acetals, ketals, etc.) can be used. (See, for example, U.S. Patents 5,122,368; 5,824,805; 5,622,929; Dubowchik and Walker, 1999, Pharm. Therapeutics 83:67-123; Neville et al, 1989, Biol. Chem. 264: 14653-14661). Such adapters are relatively stable under neutral pH conditions (such as the pH in blood) but unstable below pH 5.5 or 5.0 (approximate to the pH of lysosomes). In some embodiments, the hydrolyzable connector is a thioether connector (such as a thioether attached to the therapeutic agent via an acylhydrazone bond, for example, see, for example, U.S. Patent No. 5,622,929).

[0160] Other connectors are cuttable under reducing conditions (e.g., disulfide connectors). Disulfide connectors include those formed using SATA (N-succinimide-S-acetylthioacetate), SPDP (N-succinimide-3-(2-pyridyldithio)propionate), SPDB (N-succinimide-3-(2-pyridyldithio)butyrate), and SMPT (N-succinimide-oxycarbonyl-α-methyl-α-(2-pyridyl-dithio)toluene), SPDB, and SMPT. (See, for example, Thorpe et al, 1987, Cancer Res. 47:5924-5931; Wawrzynczak et al, In Immunoconjugates: Antibody Conjugates in Radioimagery and Therapy of Cancer (CW Vogel ed., Oxford U. Press, 1987. See also U.S. Patent No. 4,880,935)).

[0161] The connector can also be a malonic acid ester connector (Johnson et al, 1995, Anticancer Res. 15:1387-93), a maleic animide benzoyl connector (Lau et al, 1995, Bioorg-Med-Chem. 3(10):1299-1304), or a 3'-N-amide analog (Lau et al, 1995, Bioorg-Med-Chem. 3(10):1305-12). The connector can also be a malonic acid ester connector (Johnson et al, 1995, Anticancer Res. 15:1387-93), a maleic animide benzoyl connector (Lau et al, 1995, Bioorg-Med-Chem. 3(10):1299-1304), or a 3'-N-amide analog (Lau et al, 1995, Bioorg-Med-Chem. 3(10):1305-12).

[0162] The linker can also be an uncuttable linker, such as a maleic anhydride-alkylene or maleic anhydride-aryl linker, which attaches directly to the therapeutic agent (e.g., a drug). The active drug-linker is released through antibody degradation.

[0163] Generally speaking, the adapter is essentially insensitive to the extracellular environment, meaning that when the ADC is present in the extracellular environment (e.g., in plasma), no more than about 20%, generally no more than about 15%, more generally no more than about 10%, and even more generally no more than about 5%, no more than about 3%, or no more than about 1% of the adapter in the ADC sample is cleaved.

[0164] Whether the adapter is substantially insensitive to the extracellular environment can be determined, for example, by independently culturing (a) an ADC (“ADC sample”) and (b) an equimolar amount of unconjugated antibody or therapeutic agent (“control sample”) in plasma for a predetermined period of time (e.g., 2, 4, 8, 16, or 24 hours), and then comparing the amount of unconjugated antibody or therapeutic agent present in the ADC sample with the amount of unconjugated antibody or therapeutic agent present in the control sample, as measured, for example, by high performance liquid chromatography.

[0165] Linkers can also promote cell internalization. Linkers can promote cell internalization when conjugated to a therapeutic agent (i.e., in the case of the linker-therapeutic agent portion of an ADC or ADC derivative as described herein). Alternatively, linkers can promote cell internalization when conjugated to both a therapeutic agent and an anti-PD-L1 antibody (i.e., in the case of an ADC as described herein).

[0166] Anti-PD-L1 antibodies can be conjugated to linkers via heteroatoms of the antibody. These heteroatoms may be natively present on the antibody or may be introduced into the antibody. In some aspects, anti-PD-L1 antibodies are conjugated to linkers via the nitrogen atom of a lysine residue. In other aspects, anti-PD-L1 antibodies are conjugated to linkers via the sulfur atom of a cysteine ​​residue. Cysteine ​​residues may be naturally occurring or engineered into the antibody. Methods for conjugating linkers and drug-linkers to antibodies via lysine and cysteine ​​residues are known in the art.

[0167] Exemplary antibody-drug conjugates include olistatin-based antibody-drug conjugates (i.e., the drug component is an olistatin drug). Olistatin binds to tubulin and has been shown to interfere with microtubule dynamics and nuclear and cell division, and possesses anticancer activity. Generally, olistatin-based antibody-drug conjugates contain a linker between the olistatin drug and the anti-PD-L1 antibody. The linker may be, for example, a cleavable linker (e.g., a peptide linker, a carbohydrate linker) or a non-cleavable linker (e.g., a linker released through antibody degradation). Olistatin includes olistatin T, MMAF, and MMAE. Exemplary synthesis and structures of olistatin are disclosed in U.S. Publications Nos. 7,659,241, 7,498,298, 2009-0111756, 2009-0018086, and 7,968,687, each of which is incorporated herein by reference in its entirety and for all purposes.

[0168] Exemplary antibody-drug conjugates also include camptothecin-based antibody-drug conjugates (i.e., the drug component is a camptothecin drug). Camptothecin is a topoisomerase inhibitor that has been shown to have anticancer activity. Generally, camptothecin-based antibody-drug conjugates contain a linker between the camptothecin drug and the anti-PD-L1 antibody. The linker can be, for example, a cleavable linker (e.g., a peptide linker, a carbohydrate linker) or a non-cleavable linker (e.g., a linker released through antibody degradation). The synthesis and structure of exemplary camptothecin drug-linker are described in PCT / US19 / 025968 (filed April 5, 2019), which is incorporated herein by reference in its entirety and for all purposes.

[0169] Other exemplary antibody-drug conjugates include maytansinoid antibody-drug conjugates (i.e., the drug component is maytansinoid drug) and benzodiazepine antibody-drug conjugates (i.e., the drug component is benzodiazepine (e.g., pyrrolo[l,4]benzodiazepine dimer (PBD dimer), indolinobenzodiazepine dimer, and oxazolidinoneobenzodiazepine dimer)).

[0170] Exemplary antibody-drug conjugates include the following vcMMAE and mcMMAF antibody-drug conjugates, where p represents the drug load and Ab represents the anti-PD-L1 antibody:

[0171]

[0172]

[0173] Or its pharmaceutically acceptable salt.

[0174] Exemplary anti-PD-L1 antibody drug conjugates include camptothecin antibody drug conjugates as follows, where p represents the drug loading and Ab represents the anti-PD-L1 antibody:

[0175] In some implementations, camptothecin ADC has the formula (IC):

[0176]

[0177] Or its pharmaceutically acceptable salt;

[0178] in

[0179] Ab is an anti-PD-L1 antibody;

[0180] y is 1, 2, 3, or 4, or 1 or 4; and

[0181] z is an integer from 2 to 12, or 2, 4, 8, or 12;

[0182] And p is 1 to 16.

[0183] In some aspects of these implementation schemes, p is 2, 3, 4, 5, 6, 7, 8, 9, or 10. In other aspects, p is 2, 4, or 8.

[0184] In some implementations, camptothecin ADC has the following formula:

[0185]

[0186] Or its pharmaceutically acceptable salt;

[0187] Where p is 2, 4, or 8, preferably p is 8.

[0188] In some implementations, camptothecin ADC has the following formula:

[0189]

[0190] Or its pharmaceutically acceptable salt;

[0191] Where p is 2, 4, or 8, preferably p is 8.

[0192] In some implementations, the camptothecin drug-connector has the following formula:

[0193]

[0194] Or its pharmaceutically acceptable salt;

[0195] in

[0196] y is 1, 2, 3, or 4, or 1 or 4; and

[0197] Z is an integer from 2 to 12, or 2, 4, 8, or 12.

[0198] In some implementations, the camptothecin drug-connector has the following formula:

[0199]

[0200] MP-PEG8-VKG-camptothecin

[0201] In some implementations, the camptothecin drug-connector has the following formula:

[0202]

[0203] MP-PEG4-VKG-camptothecin

[0204] In some implementations, the camptothecin drug-connector has the following formula:

[0205]

[0206] MP-PEG12-VKG-camptothecin

[0207] When referring to PD-L1-targeting antibody-drug conjugates, the subscript 'p' indicates drug loading. Depending on the context, this can represent the number of drug-adaptor molecules attached to an individual antibody molecule and thus be an integer value, or it can represent the average drug loading and thus be an integer or non-integer value, but is generally non-integer. Average drug loading represents the average number of drug-adaptor molecules per antibody in the population. Typically, but not always, when we refer to antibodies (e.g., monoclonal antibodies), we are referring to the population of antibody molecules. In compositions containing a population of antibody-drug conjugate molecules, average drug loading is an important quality attribute because it determines the amount of drug that can be delivered to the target cells. The percentage of unconjugated antibody molecules in the composition is included in the average drug loading value.

[0208] In a preferred aspect of the invention, when referring to a composition comprising a group of antibody-drug conjugate compounds, the average drug loading is from 1 to about 16, preferably from about 2 to about 14, and more preferably from about 2 to about 10.

[0209] For MMAE and camptothecin ADCs (such as those illustrated herein), the preferred average drug loading is about 2, 4, or 8, with a particularly preferred average drug loading of about 8. In embodiments, the preferred average drug loading for the MMAE ADC is 2 or 4. In embodiments, the preferred average drug loading for the camptothecin ADC is 4 or 8. In exemplary embodiments, the drug-linker is conjugated to a cysteine ​​residue of a reduced interchain disulfide. In some aspects, the actual drug loading of an individual antibody molecule in a group of antibody-drug conjugate compounds is 1 to 10 (or 6 to 10 or 6 to 8), with a predominantly high drug loading of 8. For example, higher drug loading can be achieved by conjugating the drug-linker to an introduced cysteine ​​residue (such as a cysteine ​​residue introduced at position 239 according to the EU index) in addition to the interchain disulfide.

[0210] The PEG (polyethylene glycol) portion of the drug connector can range from 2 to 36. In all the embodiments described above, the subscript z is preferably 2 to 12, 4 to 12, 8 to 14, 8 to 12, 10 to 12, or 10 to 14, more preferably 2, 4, 8, or 12, and most preferably 8.

[0211] Polydisperse PEG, monodisperse PEG, and discrete PEG can be used to manufacture the polyethylene glycol-modified antibody-drug conjugates of the present invention. Polydisperse PEG is a heterogeneous mixture of (multiple) sizes and molecular weights, while monodisperse PEG is generally purified from a heterogeneous mixture and thus provides a single chain length and molecular weight. The preferred PEG unit is discrete PEG, i.e., a compound synthesized stepwise and not via polymerization. Discrete PEG provides a single molecule with a defined and specified chain length. As with the subscript "p", when referring to a population of antibody-drug conjugates, the value of the subscript "n" can be an average number and can be an integer or a non-integer.

[0212] Classes of cytotoxic agents useful for conjugating anti-PD-L1 antibodies include, for example, anti-microtubule agents, DNA minor groove binding agents, DNA replication inhibitors, and chemotherapy sensitizers. Other exemplary classes of cytotoxic agents include anthracycline, oliquistatin, camptothecin, duocarmycin, etoposide, maytansine-like compounds, and vinca alkaloids. Some exemplary cytotoxic agents include olistatins (e.g., olistatin T, olistatin E, AFP, monomethylolistatin F (MMAF), lipophilic monomethylolistatin F, monomethylolistatin E (MMAE)), DNA minor groove binding agents (e.g., enediyne and lexitropsin), pyroxine, taxanes (e.g., paclitaxel and docetaxel), vinca alkaloids, nicotinamide phosphoribosyltransferase inhibitor (NAMPTi), tubulysin M, doxorubicin, morpholino doxorubicin, and cyanomorpholino doxorubicin.

[0213] Cytotoxic agents can be chemotherapeutic agents, such as doxorubicin, paclitaxel, melphalan, vinca alkaloids, methotrexate, mitomycin C, or etoposide. These agents can also be analogues of CC-1065, calicheamicin, maytansine, dolastatin 10, rhizoxin, or pallytoxin.

[0214] Cytotoxic agents can also include olistatin. Olistatin can be an olistatin E derivative, which is, for example, an ester formed between olistatin E and a keto acid. For example, olistatin E can be reacted with p-acetylbenzoic acid or benzoylvaleric acid to produce AEB and AEB, respectively. Other typical olistatins include olistatin T, AFP, MMAF, and MMAE. The synthesis and structure of various olistatins are described, for example, in US 2005-0238649 and US 2006-0074008.

[0215] Cytotoxic agents can be DNA minor groove binding agents (see, for example, U.S. Patent No. 6,130,237). For example, minor groove binding agents can be CBI compounds or enediynes (e.g., kazimidox).

[0216] Cytotoxic agents or cell growth inhibitors can be anti-tubulin agents. Examples of anti-tubulin agents include taxanes (e.g., Taxol®, Taxotere®), T67 (Tularik), vinca alkaloids (e.g., vincristine, vinblastine, vindesine, and vinorelbine), and oliganthrines (e.g., oliganthrine E, AFP, MMAF, MMAE, AEB, and AEB). Exemplary oliganthrines are shown below in Formulas III through XIII. Other suitable anti-microtubule agents include, for example, baccatin derivatives, taxane analogs (e.g., epothilone A and B), nocodazole, colchicine and colcimid, estramustine, cryptophysin, cemadotin, maytansine, cobretastatin, discodermolide, and eleuthrobin.

[0217] Cytotoxic agents can be maytansine-like agents, which are another group of anti-microtubule agents (e.g., DM1, DM2, DM3, DM4). For example, maytansine-like agents can be maytansine or maytansine containing drug linkers such as DM-1 or DM-4 (ImmunoGen, Inc.; see also Chari et al., 1992, Cancer Res.).

[0218] VIII. Therapeutic Use

[0219] This article provides a method for treating subjects with various cancers using a combination of PD-L1-binding antibody-drug conjugates (ADCs) and anti-PD-1 antibodies (such as pembrolizumab).

[0220] In one aspect, this document provides a method for treating a subject's cancer using a PD-L1-binding ADC and an anti-PD-1 antibody (e.g., pembrolizumab). In some embodiments, the human subject treated with the methods provided herein has previously received cancer treatment. In some embodiments, the human subject treated with the methods provided herein has previously received treatment including or composed of a checkpoint inhibitor (CPI). In some embodiments, the CPI is an anti-PD-1 antibody (e.g., pembrolizumab). In some embodiments, the CPI is a PD-1 inhibitor, a PD-L1 inhibitor, or a PD-L2 inhibitor (including, but not limited to, atezolizumab, pembrolizumab, nivolumab, durvalumab, or aveluma). In a particular embodiment, the CPI is atezolizumab, pembrolizumab, nivolumab, durvalumab, or aveluma. In some embodiments, human subjects treated with the methods described herein have not previously received an agent targeting another stimulating or co-inhibitory T-cell receptor (including, but not limited to, CD137 agonists, CTLA 4 inhibitors, or OX-40 agonists). In specific embodiments, the agent targeting another stimulating or co-inhibitory T-cell receptor is a CD137 agonist, a CTLA 4 inhibitor, or an OX-40 agonist. In some embodiments, human subjects treated with the methods described herein are not eligible for cisplatin treatment. In other embodiments, human subjects treated with the methods described herein are not eligible for cisplatin treatment and have not previously received treatment including or composed of CPIs. In some embodiments, human subjects treated with the methods described herein are not eligible for cisplatin treatment, have not previously received treatment including or composed of CPIs, and have not received adjuvant / platin-based neoadjuvant therapy within the 12 months prior to randomization. In a further embodiment, human subjects treated with the methods provided herein are not eligible for cisplatin treatment, have not previously received treatment including or composed of CPIs, have not received adjuvant / platin-based neoadjuvant therapy within 12 months prior to randomization, and have not previously received systemic therapy for locally advanced or metastatic disease.

[0221] Examples of cancers associated with and suitable for treatment include melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), head and neck cancer, triple-negative breast cancer (TNBC), ovarian cancer, urothelial carcinoma, hepatocellular carcinoma (HCC), gastric cancer, and cervical cancer. In some embodiments, the antibody or antibody-drug conjugate of the present invention is used in a method of treating melanoma. In some embodiments, the antibody or antibody-drug conjugate of the present invention is used in a method of treating NSCLC. In some embodiments, the antibody or antibody-drug conjugate of the present invention is used in a method of treating SCLC. In some embodiments, the antibody or antibody-drug conjugate of the present invention is used in a method of treating head and neck cancer. In some embodiments, the antibody or antibody-drug conjugate of the present invention is used in a method of treating TNBC. Triple-negative breast cancer is a technical term for cancers lacking detectable estrogen and progesterone receptors and lacking HER2 / neu overexpression. In some embodiments, the antibody or antibody-drug conjugate of the present invention is used in a method of treating ovarian cancer. In some embodiments, the antibody or antibody-drug conjugate of the present invention is used in a method of treating urothelial carcinoma. In some embodiments, the antibodies or antibody-drug conjugates of the present invention are used in methods for treating HCC. In some embodiments, the antibodies or antibody-drug conjugates of the present invention are used in methods for treating gastric cancer. In some embodiments, the antibodies or antibody-drug conjugates of the present invention are used in methods for treating cervical cancer. This treatment can be applied to patients with such primary or metastatic tumors. This treatment can also be applied to patients who are refractory to conventional treatment or who have relapsed after responding to such treatment.

[0222] Administering the antibodies of this invention (such as humanized antibodies, alone or in conjugate form) in an effective regimen means, at a dose, route, and frequency, delaying the onset of cancer, reducing the severity of cancer, inhibiting further progression of cancer, and / or improving at least one sign or symptom of cancer. If the patient already has cancer, this regimen may be considered therapeutically effective. If the patient is at an elevated risk of cancer relative to the general population but has not yet experienced symptoms, this regimen may be considered preventatively effective. In some cases, therapeutic or preventative efficacy may be observed in individual patients relative to historical controls or past experiences in the same patients. In other cases, therapeutic or preventative efficacy may be demonstrated in preclinical or clinical trials in treated patient populations relative to a control group of untreated patients.

[0223] Exemplary doses of monoclonal antibodies are 0.1 mg / kg to 50 mg / kg of patient body weight, more generally 0.5 mg / kg to 30 mg / kg, 1 mg / kg to 30 mg / kg, 1 mg / kg to 20 mg / kg, 1 mg / kg to 15 mg / kg, 1 mg / kg to 12 mg / kg, or 1 mg / kg to 10 mg / kg, or 2 mg / kg to 30 mg / kg, 2 mg / kg to 20 mg / kg, 2 mg / kg to 15 mg / kg, 2 mg / kg to 12 mg / kg, or 2 mg / kg to 10 mg / kg, or 3 mg / kg to 30 mg / kg, 3 mg / kg to 20 mg / kg, 3 mg / kg to 15 mg / kg, 3 mg / kg to 12 mg / kg, or 3 mg / kg to 10 mg / kg. Exemplary doses of monoclonal antibodies or antibody-drug conjugates thereof are 0.5 mg / kg to 7.5 mg / kg, 1 mg / kg to 7.5 mg / kg, or 2 mg / kg to 7.5 mg / kg or 3 mg / kg to 7.5 mg / kg of the subject's body weight, or 0.1 to 20, or 0.5 to 5 mg / kg of body weight (e.g., 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mg / kg) or as a fixed dose of 10 to 1500 or 200 to 1500 mg. In some methods, patients are administered doses of at least 0.5 mg / kg, 1.0 mg / kg, 1.5 mg / kg, at least 2 mg / kg, or at least 3 mg / kg every three weeks or longer. In some methods, patients are administered doses of at least 0.5 mg / kg, 1.0 mg / kg, 1.5 mg / kg, 2 mg / kg, or 3 mg / kg every three weeks or more frequently. In some methods, patients are administered a dose of approximately 0.5 mg / kg, 0.75 mg / kg, 1.0 mg / kg, 1.25 mg / kg, 1.5 mg / kg, 1.75 mg / kg, 2 mg / kg, or 3 mg / kg, once every three weeks or more frequently. Among other factors, the dosage depends on the frequency of administration, the patient's condition, and the response to prior treatment (if any), whether prophylactic or therapeutic, and whether the condition is acute or chronic.

[0224] Administration can be parenteral, intravenous, oral, subcutaneous, intra-arterial, intracranial, intrathecal, intraperitoneal, local, intranasal, or intramuscular. It can also be directly targeted to the tumor. Systemic circulation via intravenous or subcutaneous administration is preferred. Intravenous administration can be achieved, for example, through infusions over a period of 30 to 90 minutes or by a single bolus injection.

[0225] Among other factors, the frequency of administration depends on the half-life of the circulating antibody or conjugate, the patient's condition, and the route of administration. In response to changes in the patient's condition or worsening of the treated cancer, the frequency of administration of the anti-PD-L1 antibody or antibody-drug conjugate described herein may be daily, weekly, every two weeks, twice every three weeks, once every three weeks, monthly, quarterly, or at irregular intervals. In some embodiments, administration is approximately weekly. In some embodiments, weekly administration occurs approximately on days 1, 8, and 15 of a 21-day cycle. In some embodiments, administration is approximately once every two weeks. In some embodiments, administration is approximately twice every three weeks. In some embodiments, administration occurs approximately on days 1 and 8 of a 21-day cycle. In some embodiments, administration is approximately once every three weeks. In some embodiments, administration is weekly. In some embodiments, weekly administration occurs on days 1, 8, and 15 of a 21-day cycle. In some embodiments, administration is once every two weeks. In some embodiments, administration is performed on days 1 and 8 of a 21-day cycle. In some embodiments, administration is performed twice every three weeks. In some embodiments, administration is performed once every three weeks. For subcutaneous administration, exemplary dosing frequencies range from daily to monthly, although more frequent or less frequent dosing is also possible. Exemplary frequencies for intravenous administration over a continuous course of treatment are between twice a week or per quarter, although more frequent or less frequent dosing is also possible. Other exemplary frequencies for intravenous administration over a continuous course of treatment are between three times a week or every four weeks, although more frequent or less frequent dosing is also possible. Other exemplary frequencies for intravenous administration over a continuous course of treatment are once every two weeks or once every three weeks, although more frequent or less frequent dosing is also possible. In some embodiments, the dosage is based on the subject's weight. In some embodiments, the subject's weight is the subject's ideal body weight (IBW). In some embodiments, the subject's weight is the subject's adjusted ideal body weight (AIBW). In some embodiments, the dose is 0.5 mg / kg and is administered approximately once a week. In some embodiments, the dosage is 0.5 mg / kg and is administered approximately every 2 weeks. In some embodiments, the dosage is 0.5 mg / kg and is administered approximately twice every 3 weeks. In some embodiments, the dosage is 0.5 mg / kg and is administered approximately every 3 weeks. In some embodiments, the dosage is 0.8 mg / kg and is administered approximately every week. In some embodiments, the dosage is 0.8 mg / kg and is administered approximately every 2 weeks. In some embodiments, the dosage is 0.8 mg / kg and is administered approximately twice every 3 weeks. In some embodiments, the dosage is 0.8 mg / kg and is administered approximately every 3 weeks. In some embodiments, the dosage is 0.9 mg / kg and is administered approximately every week.In some embodiments, the dosage is 0.9 mg / kg and is administered approximately every 2 weeks. In some embodiments, the dosage is 0.9 mg / kg and is administered approximately twice every 3 weeks. In some embodiments, the dosage is 0.9 mg / kg and is administered approximately every 3 weeks. In some embodiments, the dosage is 1.0 mg / kg and is administered approximately every 1 week. In some embodiments, the dosage is 1.0 mg / kg and is administered approximately every 2 weeks. In some embodiments, the dosage is 1.0 mg / kg and is administered approximately twice every 3 weeks. In some embodiments, the dosage is 1.0 mg / kg and is administered approximately every 3 weeks. In some embodiments, the dosage is 1.1 mg / kg and is administered approximately every 1 week. In some embodiments, the dosage is 1.1 mg / kg and is administered approximately every 2 weeks. In some embodiments, the dosage is 1.1 mg / kg and is administered approximately twice every 3 weeks. In some embodiments, the dosage is 1.1 mg / kg and is administered approximately every 3 weeks. In some embodiments, the dosage is 1.2 mg / kg and administered approximately once every week. In some embodiments, the dosage is 1.2 mg / kg and administered approximately once every two weeks. In some embodiments, the dosage is 1.2 mg / kg and administered approximately twice every three weeks. In some embodiments, the dosage is 1.2 mg / kg and administered approximately once every three weeks. In some embodiments, the dosage is 1.25 mg / kg and administered approximately once every week. In some embodiments, the dosage is 1.25 mg / kg and administered approximately once every two weeks. In some embodiments, the dosage is 1.25 mg / kg and administered approximately twice every three weeks. In some embodiments, the dosage is 1.25 mg / kg and administered approximately once every three weeks. In some embodiments, the dosage is 1.3 mg / kg and administered approximately once every week. In some embodiments, the dosage is 1.3 mg / kg and administered approximately once every two weeks. In some embodiments, the dosage is 1.3 mg / kg and administered approximately twice every three weeks. In some embodiments, the dosage is 1.3 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 1.4 mg / kg and administered approximately every week. In some embodiments, the dosage is 1.4 mg / kg and administered approximately every 2 weeks. In some embodiments, the dosage is 1.4 mg / kg and administered twice approximately every 3 weeks. In some embodiments, the dosage is 1.4 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 1.5 mg / kg and administered approximately every week. In some embodiments, the dosage is 1.5 mg / kg and administered approximately every 2 weeks. In some embodiments, the dosage is 1.5 mg / kg and administered twice approximately every 3 weeks. In some embodiments, the dosage is 1.5 mg / kg and administered approximately every 3 weeks.In some embodiments, the dosage is 1.6 mg / kg and administered approximately once every week. In some embodiments, the dosage is 1.6 mg / kg and administered approximately once every two weeks. In some embodiments, the dosage is 1.6 mg / kg and administered approximately twice every three weeks. In some embodiments, the dosage is 1.6 mg / kg and administered approximately once every three weeks. In some embodiments, the dosage is 1.7 mg / kg and administered approximately once every week. In some embodiments, the dosage is 1.7 mg / kg and administered approximately once every two weeks. In some embodiments, the dosage is 1.7 mg / kg and administered approximately twice every three weeks. In some embodiments, the dosage is 1.7 mg / kg and administered approximately once every three weeks. In some embodiments, the dosage is 1.75 mg / kg and administered approximately once every week. In some embodiments, the dosage is 1.75 mg / kg and administered approximately once every two weeks. In some embodiments, the dosage is 1.75 mg / kg and administered approximately twice every three weeks. In some embodiments, the dosage is 1.75 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 1.8 mg / kg and administered approximately every 1 week. In some embodiments, the dosage is 1.8 mg / kg and administered approximately every 2 weeks. In some embodiments, the dosage is 1.8 mg / kg and administered twice approximately every 3 weeks. In some embodiments, the dosage is 1.8 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 1.9 mg / kg and administered approximately every 1 week. In some embodiments, the dosage is 1.9 mg / kg and administered approximately every 2 weeks. In some embodiments, the dosage is 1.9 mg / kg and administered twice approximately every 3 weeks. In some embodiments, the dosage is 1.9 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 2.0 mg / kg and administered approximately every 1 week. In some embodiments, the dosage is 2.0 mg / kg and administered approximately every 2 weeks. In some embodiments, the dosage is 2.0 mg / kg and administered approximately twice every 3 weeks. In some embodiments, the dosage is 2.0 mg / kg and administered approximately once every 3 weeks. In some embodiments, the dosage is 2.1 mg / kg and administered approximately once every week. In some embodiments, the dosage is 2.1 mg / kg and administered approximately once every 2 weeks. In some embodiments, the dosage is 2.1 mg / kg and administered approximately twice every 3 weeks. In some embodiments, the dosage is 2.1 mg / kg and administered approximately once every 3 weeks. In some embodiments, the dosage is 2.2 mg / kg and administered approximately once every week. In some embodiments, the dosage is 2.2 mg / kg and administered approximately once every 2 weeks. In some embodiments, the dosage is 2.2 mg / kg and administered approximately twice every 3 weeks.In some embodiments, the dosage is 2.2 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 2.3 mg / kg and administered approximately every 1 week. In some embodiments, the dosage is 2.3 mg / kg and administered approximately every 2 weeks. In some embodiments, the dosage is 2.3 mg / kg and administered twice approximately every 3 weeks. In some embodiments, the dosage is 2.3 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 2.4 mg / kg and administered approximately every 1 week. In some embodiments, the dosage is 2.4 mg / kg and administered approximately every 2 weeks. In some embodiments, the dosage is 2.4 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 2.4 mg / kg and administered approximately every 3 weeks. In some embodiments, the dosage is 2.5 mg / kg and administered approximately every 1 week. In some embodiments, the dosage is 2.5 mg / kg and administered approximately every 2 weeks. In some implementations, the dose is 2.5 mg / kg and administered approximately twice every 3 weeks. In some implementations, the dose is 2.5 mg / kg and administered approximately once every 3 weeks. In some methods, the patient is administered a dose of approximately 0.5 mg / kg, 0.75 mg / kg, 1.0 mg / kg, 1.25 mg / kg, 1.5 mg / kg, 1.75 mg / kg, 2 mg / kg, or 3 mg / kg on days 1, 8, and 15 of a 21-day cycle. In some methods, the patient is administered a dose of 1.25 mg / kg on days 1, 8, and 15 of a 21-day cycle. In some methods, the patient is administered a dose of 1.5 mg / kg on days 1, 8, and 15 of a 21-day cycle. In some methods, patients are administered a dose of approximately 0.5 mg / kg, 0.75 mg / kg, 1.0 mg / kg, 1.25 mg / kg, 1.5 mg / kg, 1.75 mg / kg, 2 mg / kg, or 3 mg / kg on days 1 and 8 of a 21-day cycle. In some methods, patients are administered a dose of 1.25 mg / kg on days 1 and 8 of a 21-day cycle. In some methods, patients are administered a dose of 1.5 mg / kg on days 1 and 8 of a 21-day cycle.

[0226] The frequency of administration depends on the nature of the cancer (e.g., whether it presents as acute or chronic symptoms) and the condition's response to treatment. For acute symptoms or acute exacerbations of chronic symptoms, 1 to 10 doses are usually sufficient. Sometimes, a single bolus dose (optionally in split doses) is sufficient for acute symptoms or acute exacerbations of chronic symptoms. For recurrence of acute symptoms or exacerbations, treatment can be repeated. For chronic symptoms, antibodies can be administered at regular intervals, such as weekly, bi-weekly, monthly, quarterly, or six-monthly, for at least 1, 5, or 10 years or the patient's lifetime.

[0227] Pharmaceutical compositions intended for parenteral administration are preferably sterile, substantially isotonic, and manufactured under GMP conditions. The pharmaceutical composition may be provided in unit dosage forms (i.e., doses for a single administration). The pharmaceutical composition may be formulated using one or more physiologically acceptable carriers, diluents, excipients, or adjuvants. The formulation depends on the chosen route of administration. For injection, antibodies may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hanks' solution, Ringer's solution, or physiological saline or acetate buffer (to reduce discomfort at the injection site). The solution may contain formulation agents such as suspending agents, stabilizers, and / or dispersants. Alternatively, antibodies may be in lyophilized form and formulated with a suitable medium (e.g., sterile, pyrogen-free water) prior to use. The antibody concentration in liquid formulations may be, for example, from 1 to 100 mg / ml, such as 10 mg / ml.

[0228] Treatment with the antibodies of this invention can be combined with chemotherapy, radiation, stem cell therapy, surgery, and other treatments effective for the condition being treated. Other available classes of agents that can be administered with antibodies and antibody-drug conjugates targeting PD-L1 as described herein include, for example, antibodies against other receptors expressed on cancer cells, anti-microtubule agents (e.g., oliquistatin), DNA minor groove binding agents, DNA replication inhibitors, alkylating agents (e.g., platinum complexes, such as cisplatin, mono(platinum), bis(platinum), and trinuclear platinum complexes, and carboplatin), anthracyclines, antibiotics, antifolate agents, antimetabolites, chemotherapy sensitizers, pyromalacin, etoposide, fluorinated pyrimidines, ionophores, lecithin, nitrosourea, platinol, preformed compounds, purine antimetabolites, puromycin, radiosensitizers, steroids, taxanes, topoisomerase inhibitors, vinca alkaloids, etc.

[0229] Treatment with PD-L1 antibodies or antibody-drug conjugates, optionally in combination with any of the other agents or regimens mentioned above, either alone or in the form of an antibody-drug conjugate, can increase median progression-free survival or overall survival by at least 30% or 40%, but preferably 50%, 60% to 70%, or even 100% or longer, in patients with tumors (e.g., melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), head and neck cancer, triple-negative breast cancer (TNBC), ovarian cancer, urothelial carcinoma, hepatocellular carcinoma (HCC), gastric cancer, and cervical cancer) (especially when the cancer is recurrent or refractory) compared to the same treatment (e.g., chemotherapy) without anti-PD-L1 antibodies alone or in the form of a conjugate. Alternatively, treatment including anti-PD-L1 antibodies, either alone or in conjugate form (e.g., standard chemotherapy), compared to the same treatment (e.g., chemotherapy) without anti-PD-L1 antibodies, either alone or in conjugate form, can increase the complete response rate, partial response rate, or objective response rate (complete + partial) in patients with tumors by at least 30% or 40%, but preferably 50%, 60% to 70%, or even 100%.

[0230] Generally, in clinical trials (e.g., phase II, II / III, or III trials), the increase in median progression-free survival and / or response rate in patients treated with the aforementioned standard therapy plus anti-PD-L1 antibodies, either alone or in conjugate form, is statistically significant, for example, at p = 0.05, 0.01, or even 0.001, compared to a control group receiving standard therapy alone (or plus placebo). Complete and partial response rates are determined using objective criteria commonly used in cancer clinical trials, such as those listed or accepted by the National Cancer Institute and / or the Food and Drug Administration.

[0231] In some embodiments, the anti-PD-L1 antibody or antibody-drug conjugate described herein is administered in combination with an anti-PD-1 antibody. In a further embodiment, the anti-PD-1 antibody is pembrolizumab. In some embodiments, the anti-PD-1 antibody is administered simultaneously with the anti-PD-L1 antibody or antibody-drug conjugate described herein. In some embodiments, the anti-PD-1 antibody and the anti-PD-L1 antibody or antibody-drug conjugate described herein are administered sequentially. In some embodiments, "simultaneously" means administering the anti-PD-L1 antibody or antibody-drug conjugate described herein to the subject at an interval of less than one hour with the anti-PD-1 antibody, such as less than about 30 minutes, less than about 15 minutes, less than about 10 minutes, or less than about 5 minutes. In some embodiments, sequential administration means administering the anti-PD-L1 antibody or antibody-drug conjugate described herein with the anti-PD-1 antibody at intervals of at least 1 hour, at least 2 hours, at least 3 hours, at least 4 hours, at least 5 hours, at least 6 hours, at least 7 hours, at least 8 hours, at least 9 hours, at least 10 hours, at least 11 hours, at least 12 hours, at least 13 hours, at least 14 hours, at least 15 hours, or at least 16 hours. Hours, at least 17 hours, at least 18 hours, at least 19 hours, at least 20 hours, at least 21 hours, at least 22 hours, at least 23 hours, at least 24 hours, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 5 days, at least 7 days, at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 6 weeks, at least 2 months, at least 3 months, at least 4 months, at least 5 months, or at least 6 months.

[0232] In some embodiments, the PD-1 is pembrolizumab or a biosimilar thereof. In some embodiments, pembrolizumab or a biosimilar thereof is administered at a dose of about 200 mg. In some embodiments, pembrolizumab or a biosimilar thereof is administered at a dose of 200 mg. In some embodiments, pembrolizumab or a biosimilar thereof is administered at a dose of about 400 mg. In some embodiments, pembrolizumab or a biosimilar thereof is administered at a dose of 400 mg. In some embodiments, pembrolizumab or a biosimilar thereof is administered approximately every 3 weeks. In some embodiments, pembrolizumab or a biosimilar thereof is administered every 3 weeks. In some embodiments, pembrolizumab or a biosimilar thereof is administered approximately every 6 weeks. In some embodiments, pembrolizumab or a biosimilar thereof is administered every 6 weeks. In some embodiments, pembrolizumab or a biosimilar thereof is administered at a dose of about 200 mg approximately every 3 weeks. In some embodiments, pembrolizumab or its biosimilar is administered at a dose of 200 mg every 3 weeks. In some embodiments, pembrolizumab or its biosimilar is administered at a dose of approximately 400 mg every approximately 6 weeks. In some embodiments, pembrolizumab or its biosimilar is administered at a dose of 400 mg every 6 weeks. In some embodiments, pembrolizumab or its biosimilar is administered intravenously. In some embodiments, a first dose of an anti-PD-L1 antibody or antibody-drug conjugate is administered to the subject before administering the first dose of pembrolizumab or its biosimilar. In some embodiments, a first dose of an anti-PD-L1 antibody or antibody-drug conjugate is administered to the subject for at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 2 months, at least 3 months, at least 4 months, at least 5 months, or at least 6 months before administering the first dose of pembrolizumab or its biosimilar.

[0233] Treatment with anti-PD-L1 antibodies or antibody-drug conjugates (optionally in combination with the anti-PD-1 antibodies described above) can increase median progression-free survival or overall survival in patients with cancer (including solid tumors). In some implementations, the cancer is melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), head and neck squamous cell carcinoma (HNSCC), triple-negative breast cancer (TNBC), esophageal squamous cell carcinoma (esophageal SCC), ovarian cancer, urothelial carcinoma, hepatocellular carcinoma (HCC), gastric cancer, or cervical cancer.

[0234] In some embodiments, treatment with an anti-PD-L1 antibody or antibody-drug conjugate (optionally in combination with the anti-PD-1 antibody described above) can drive bystander-effect-based cytotoxicity against PD-L1-negative tumor cells. In some embodiments, treatment with an anti-PD-L1 antibody or antibody-drug conjugate (optionally in combination with the anti-PD-1 antibody described above) can drive bystander-effect-based cytotoxicity against PD-L1-negative tumor cells when PD-L1-negative tumor cells are in a tumor microenvironment that also contains PD-L1-positive tumor cells. In some further embodiments, treatment with an anti-PD-L1 antibody or antibody-drug conjugate induces bystander-effect-based cytotoxicity in PD-L1-negative tumor cells at levels at least about one of the following higher than those induced by antibodies or antibody-drug conjugates that are not specific to PD-L1: 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 2 times, 5 times, 10 times, 100 times, 1000 times, or higher, optionally wherein the PD-L1-negative tumor cells are in a tumor microenvironment that also contains PD-L1-positive tumor cells.

[0235] Immunogenic cell death is a regulated activation that recruits immune cells to the tumor microenvironment. This activation is induced by the release of damage-associated molecular patterns (DAMPs) from tumor cells, such as cell surface chaperone calreticulin (CRT), extracellular adenosine triphosphate (eATP), and high-mobility group box 1 protein (HMGB1). In some embodiments, treatment with anti-PD-L1 antibodies or antibody-drug conjugates (optionally in combination with the aforementioned anti-PD-1 antibodies) can drive an increase in immunogenic cell death. In some embodiments, treatment with anti-PD-L1 antibodies or antibody-drug conjugates (optionally in combination with the aforementioned anti-PD-1 antibodies) can induce activation that recruits immune cells to the tumor microenvironment. In some implementations, treatment with an anti-PD-L1 antibody or antibody-drug conjugate (optionally in combination with the anti-PD-1 antibody described above) can induce activation of immune cells recruited to the tumor microenvironment at levels at least about one of the following higher than those induced by antibodies or antibody-drug conjugates that are not specific to PD-L1: 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 2 times, 5 times, 10 times, 100 times, 1000 times, or higher. In some implementations, treatment with an anti-PD-L1 antibody or antibody-drug conjugate (optionally in combination with the anti-PD-1 antibody described above) can induce the release of CRT, eATP, and / or HMGB1 at levels at least about one of the following higher than those induced by antibodies or antibody-drug conjugates that are not specific to PD-L1: 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 2 times, 5 times, 10 times, 100 times, 1000 times, or higher.

[0236] In some embodiments, treatment with anti-PD-L1 antibodies or antibody-drug conjugates (optionally in combination with the anti-PD-1 antibodies described above) does not result in significant depletion of immune cells. In some embodiments, treatment with anti-PD-L1 antibodies or antibody-drug conjugates (optionally in combination with the anti-PD-1 antibodies described above) induces immune cell death at levels less than any one of about 2%, 5%, 10%, 15%, 20%, or 30% compared to the level induced by antibodies or antibody-drug conjugates that are not specific to PD-L1, at least one of the following: 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 2 times, 5 times, 10 times, 100 times, 1000 times, or less. In some implementations, the immune cells include one or more of CD4+ T cells, CD8+ T cells, macrophages, and / or dendritic cells.

[0237] In some embodiments, treatment with an anti-PD-L1 antibody or antibody-drug conjugate (optionally in combination with the anti-PD-1 antibody described above) results in the targeted depletion of immunosuppressive cells, such as immune cells that suppress natural anti-tumor immunity. In some embodiments, treatment with an anti-PD-L1 antibody or antibody-drug conjugate (optionally in combination with the anti-PD-1 antibody described above) induces at least about one of the following, compared to the death of immunosuppressive cells at the start of treatment: 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70%, or more. In some embodiments, treatment with an anti-PD-L1 antibody or antibody-drug conjugate (optionally in combination with the anti-PD-1 antibody described above) induces immunosuppressive cell death at levels at least one of the following higher than those induced by antibodies or antibody-drug conjugates that are not specific to PD-L1: 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 2-fold, 5-fold, 10-fold, 100-fold, 1000-fold, or higher. In some embodiments, the immunosuppressive cells comprise one or more of regulatory T cells (Tregs), myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), tumor-associated neutrophils (TANs), and / or tumor-associated dendritic cells (tDCs).

[0238] Checkpoint inhibitors and combination therapy with pembrolizumab

[0239] In some implementations, subjects treated with the methods provided herein may possess certain phenotypic or genotypic characteristics. In some implementations, subjects may possess any permutation and combination of the phenotypic or genotypic characteristics described herein.

[0240] In some embodiments, phenotypic or genotypic characteristics are determined histologically, cytologically, or both. In some embodiments of the methods provided herein, the histological and / or cytological determination of phenotypic and / or genotypic characteristics is performed based on recently analyzed tissues as described in the American Society of Clinical Oncology / American Society of Pathologists (ASCO / CAP) guidelines, which are incorporated herein by reference in their entirety. In some embodiments, phenotypic or genotypic characteristics are determined by sequencing including next-generation sequencing (e.g., NGS from Illumina, Inc.), DNA hybridization, and / or RNA hybridization.

[0241] In various aspects or embodiments of the methods provided herein, human subjects suitable for use with the methods provided herein have not received prior treatment with any checkpoint inhibitor (CPI). A CPI is defined as a PD-1 inhibitor, PD-L1 inhibitor, or PD-L2 inhibitor (including, but not limited to, atezolizumab, pembrolizumab, nivolumab, durvalumab, or avelumab). In some embodiments, human subjects suitable for use with the methods provided herein have not received prior treatment with a PD-1 inhibitor, PD-L1 inhibitor, or PD-L2 inhibitor. In some embodiments, human subjects suitable for use with the methods provided herein have not received prior treatment with atezolizumab, pembrolizumab, nivolumab, durvalumab, or avelumab. As used herein, the term "immune checkpoint inhibitor" or "checkpoint inhibitor" (CPI) refers to a molecule that completely or partially reduces, inhibits, interferes with, or modulates one or more checkpoint proteins. Several checkpoint proteins are known, such as CTLA-4 and its ligands CD80 and CD86; and PD-1 and its ligands PD-L1 and PD-L2 (Pardoll, Nature Reviews Cancer , 2012, 12(252-264). Other exemplary checkpoint proteins include LAG-3, B7, TIM3 (HAVCR2), OX40 (CD134), GITR, CD137, CD40, VTCN1, IDO1, CD276, PVRIG, TIGIT, CD25 (IL2RA), IFNAR2, IFNAR1, CSF1R, VSIR (VISTA), or HLA. These proteins appear to be responsible for co-stimulatory or inhibitory interactions in T cell responses. Immune checkpoint proteins appear to regulate and maintain self-tolerance and the duration and magnitude of physiological immune responses. Immune checkpoint inhibitors include antibodies or antibody-derived antibodies.

[0242] In some embodiments, the checkpoint inhibitor used in the methods provided herein may be an inhibitor or activator of a checkpoint protein upregulated in cancer. In some specific embodiments, the checkpoint inhibitor used in the methods provided herein may be an inhibitor or activator of a checkpoint protein, including LAG-3, B7, TIM3 (HAVCR2), OX40 (CD134), GITR, CD137, CD40, VTCN1, IDO1, CD276, PVRIG, TIGIT, CD25 (IL2RA), IFNAR2, IFNAR1, CSF1R, VSIR (VISTA), or HLA. In some embodiments, the checkpoint inhibitor used in the methods provided herein may be selected from inhibitors or activators belonging to the group consisting of: PD-1 inhibitors, PD-L1 inhibitors, PD-L2 inhibitors, CTLA-4 inhibitors, LAG-3 inhibitors, B7 inhibitors, TIM3 (HAVCR2) inhibitors, OX40 (CD134) inhibitors, GITR agonists, CD137 agonists, or CD40 agonists, VTCN1 inhibitors, IDO1 inhibitors, CD276 inhibitors, PVRIG inhibitors, TIGIT inhibitors, CD25 (IL2RA) inhibitors, IFNAR2 inhibitors, IFNAR1 inhibitors, CSF1R inhibitors, VSIR (VISTA) inhibitors, or HLA-targeting therapeutic agents. Such inhibitors, activators, or therapeutic agents are further provided below.

[0243] In some embodiments, the checkpoint inhibitor is a CTLA-4 inhibitor. In one embodiment, the CTLA-4 inhibitor is an anti-CTLA-4 antibody. Examples of anti-CTLA-4 antibodies include, but are not limited to, those described in U.S. Patent Nos. 5,811,097, 5,811,097, 5,855,887, 6,051,227, 6,207,157, 6,682,736, 6,984,720, and 7,605,238, all of which are incorporated herein by reference in their entirety. In one embodiment, the anti-CTLA-4 antibody is tremelimumab (also known as ticilimumab or CP-675,206). In another embodiment, the anti-CTLA-4 antibody is ipilimumab (also known as MDX-010 or MDX-101). Ipilimumab is a fully human monoclonal IgG antibody that binds to CTLA-4. Ipilimumab is marketed under the brand name Yervoy™.

[0244] In some implementations, the checkpoint inhibitor is a PD-1 / PD-L1 inhibitor. Examples of PD-1 / PD-L1 inhibitors include, but are not limited to, those described in U.S. Patent Nos. 7,488,802, 7,943,743, 8,008,449, 8,168,757, and 8,217,149, and PCT Patent Application Publications WO2003042402, WO2008156712, WO2010089411, WO2010036959, WO2011066342, WO2011159877, WO2011082400, and WO2011161699, all of which are incorporated herein by reference in their entirety.

[0245] "PD-1 antagonist" means any compound or biomolecule that blocks the binding of PD-L1 expressed on cancer cells to PD-1 expressed on immune cells (T cells, B cells, or natural killer T cells) and, in a particular embodiment, also blocks the binding of PD-L2 expressed on cancer cells to PD-1 expressed on immune cells. Alternative names or synonyms for PD-1 and its ligands include: PD-1 is PDCD1, PD-1, CD279, and SLEB2; PD-L1 is PDCD1L1, PDL1, B7H1, B7-4, CD274, and B7-H; and PD-L2 is PDCD1L2, PDL2, B7-DC, Btdc, and CD273. In any treatment method, pharmaceutical agent, or use of the present invention in which a human subject is being treated, the PD-1 antagonist blocks the binding of human PD-L1 to human PD-1 and, in a particular embodiment, blocks the binding of human PD-L1 and PD-L2 to human PD-1. The human PD-1 amino acid sequence can be found at NCBI locus number NP_005009. The human PD-L1 and PD-L2 amino acid sequences can be found at NCBI loci number NP_054862 and NP_079515, respectively.

[0246] In some embodiments, the checkpoint inhibitor is a PD-1 inhibitor or antagonist. In one embodiment, the PD-1 inhibitor or antagonist is an anti-PD-1 antibody. In one embodiment, the anti-PD-1 antibody is BGB-A317, nivolumab (also known as ONO-4538, BMS-936558, or MDX1106), or pembrolizumab (also known as MK-3475, SCH 900475, or lambrolizumab). In one embodiment, the anti-PD-1 antibody is nivolumab. Nivolumab is a human IgG4 anti-PD-1 monoclonal antibody and is marketed under the brand name Opdivo™. In another embodiment, the anti-PD-1 antibody is pembrolizumab. Pembrolizumab is a humanized monoclonal IgG4 antibody and is marketed under the brand name Keytruda™. In yet another embodiment, the anti-PD-1 antibody is the humanized antibody CT-011. CT-011 alone showed no response to treatment of relapsed acute myeloid leukemia (AML). In yet another embodiment, the anti-PD-1 antibody is the fusion protein AMP-224. In another embodiment, the anti-PD-1 antibody is BGB-A317. BGB-A317 is a monoclonal antibody in which the ability to bind to Fcγ receptor I is specifically engineered to be removed, and it has a unique binding characteristic to PD-1 with high affinity and superior target specificity. In one embodiment, the anti-PD-1 antibody is cimipril. In another embodiment, the anti-PD-1 antibody is camrelizumab. In a further embodiment, the anti-PD-1 antibody is sintilimab. In some embodiments, the PD-1 antibody is tislelizumab. In some embodiments, the anti-PD-1 antibody is TSR-042. In yet another embodiment, the anti-PD-1 antibody is PDR001. In yet another embodiment, the anti-PD-1 antibody is toripalimab.

[0247] In some embodiments, the checkpoint inhibitor is a PD-L1 inhibitor. In one embodiment, the PD-L1 inhibitor is an anti-PD-L1 antibody. In one embodiment, the anti-PD-L1 antibody is MEDI4736 (dvorumab). In another embodiment, the anti-PD-L1 antibody is BMS-936559 (also known as MDX-1105-01). In yet another embodiment, the PD-L1 inhibitor is atezolizumab (also known as MPDL3280A and Tecentriq®). In a further embodiment, the PD-L1 inhibitor is avelumab.

[0248] In one embodiment, the checkpoint inhibitor is a PD-L2 inhibitor. In one embodiment, the PD-L2 inhibitor is an anti-PD-L2 antibody. In one embodiment, the anti-PD-L2 antibody is rHIgM12B7A.

[0249] In one embodiment, the checkpoint inhibitor is a lymphocyte activation gene-3 (LAG-3) inhibitor. In one embodiment, the LAG-3 inhibitor is a soluble Ig fusion protein IMP321 (Brignone). et al. , J. Immunol. , 2007, 179 (4202-4211). In another embodiment, the LAG-3 inhibitor is BMS-986016.

[0250] In one embodiment, the checkpoint inhibitor is a B7 inhibitor. In one embodiment, the B7 inhibitor is a B7-H3 inhibitor or a B7-H4 inhibitor. In one embodiment, the B7-H3 inhibitor is the anti-B7-H3 antibody MGA271 (Loo et al. , Clin.Cancer Res. , 2012, 3834).

[0251] In one implementation, the checkpoint inhibitor is a TIM3 (T cell immunoglobulin domain and mucin domain 3) inhibitor (Fourcade). et al. , J. Exp. Med. , 2010, 207 , 2175-86; Sakurashi et al. , J. Exp. Med. , 2010, 207 , 2187-94).

[0252] In one embodiment, the checkpoint inhibitor is an OX40 (CD134) agonist. In one embodiment, the checkpoint inhibitor is an anti-OX40 antibody. In one embodiment, the anti-OX40 antibody is anti-OX-40. In another embodiment, the anti-OX40 antibody is MEDI6469.

[0253] In one embodiment, the checkpoint inhibitor is a GITR agonist. In one embodiment, the checkpoint inhibitor is an anti-GITR antibody. In one embodiment, the anti-GITR antibody is TRX518.

[0254] In one embodiment, the checkpoint inhibitor is a CD137 agonist. In another embodiment, the checkpoint inhibitor is an anti-CD137 antibody. In one embodiment, the anti-CD137 antibody is urelumab. In yet another embodiment, the anti-CD137 antibody is PF-05082566.

[0255] In one embodiment, the checkpoint inhibitor is a CD40 agonist. In one embodiment, the checkpoint inhibitor is an anti-CD40 antibody. In one embodiment, the anti-CD40 antibody is CF-870,893.

[0256] In one implementation, the checkpoint inhibitor is recombinant human interleukin IL-15 (rhIL-15).

[0257] In one implementation, the checkpoint inhibitor is a VTCN inhibitor. In another implementation, the VTCN inhibitor is FPA150.

[0258] In one embodiment, the checkpoint inhibitor is an IDO inhibitor. In one embodiment, the IDO inhibitor is INCB024360. In another embodiment, the IDO inhibitor is indoximod. In one embodiment, the IDO inhibitor is epacadostat. In another embodiment, the IDO inhibitor is BMS986205. In yet another embodiment, the IDO inhibitor is Navoximod. In one embodiment, the IDO inhibitor is PF-06840003. In another embodiment, the IDO inhibitor is KHK2455. In yet another embodiment, the IDO inhibitor is RG70099. In one embodiment, the IDO inhibitor is IOM-E. In another embodiment, the IDO inhibitor is IOM-D.

[0259] In some embodiments, the checkpoint inhibitor is a TIGIT inhibitor. In some embodiments, the TIGIT inhibitor is an anti-TIGIT antibody. In one embodiment, the TIGIT inhibitor is MTIG7192A. In another embodiment, the TIGIT inhibitor is BMS-986207. In yet another embodiment, the TIGIT inhibitor is OMP-313M32. In one embodiment, the TIGIT inhibitor is MK-7684. In another embodiment, the TIGIT inhibitor is AB154. In yet another embodiment, the TIGIT inhibitor is CGEN-15137. In one embodiment, the TIGIT inhibitor is SEA-TIGIT. In another embodiment, the TIGIT inhibitor is ASP8374. In yet another embodiment, the TIGIT inhibitor is AJUD008.

[0260] In some embodiments, the checkpoint inhibitor is a VSIR inhibitor. In some embodiments, the VSIR inhibitor is an anti-VSIR antibody. In one embodiment, the VSIR inhibitor is MTIG7192A. In another embodiment, the VSIR inhibitor is CA-170. In yet another embodiment, the VSIR inhibitor is JNJ 61610588. In one embodiment, the VSIR inhibitor is HMBD-002.

[0261] In some embodiments, the checkpoint inhibitor is a TIM3 inhibitor. In some embodiments, the TIM3 inhibitor is an anti-TIM3 antibody. In one embodiment, the TIM3 inhibitor is AJUD009.

[0262] In some embodiments, the checkpoint inhibitor is a CD25 (IL2RA) inhibitor. In some embodiments, the CD25 (IL2RA) inhibitor is an anti-CD25 (IL2RA) antibody. In one embodiment, the CD25 (IL2RA) inhibitor is daclizumab. In another embodiment, the CD25 (IL2RA) inhibitor is basiliximab.

[0263] In some embodiments, the checkpoint inhibitor is an IFNAR1 inhibitor. In some embodiments, the IFNAR1 inhibitor is an anti-IFNAR1 antibody. In one embodiment, the IFNAR1 inhibitor is anifrolumab. In another embodiment, the IFNAR1 inhibitor is sifalimumab.

[0264] In some embodiments, the checkpoint inhibitor is a CSF1R inhibitor. In some embodiments, the CSF1R inhibitor is an anti-CSF1R antibody. In one embodiment, the CSF1R inhibitor is pexidartinib. In another embodiment, the CSF1R inhibitor is emactuzumab. In yet another embodiment, the CSF1R inhibitor is cabilazumab. In one embodiment, the CSF1R inhibitor is ARRY-382. In another embodiment, the CSF1R inhibitor is BLZ945. In yet another embodiment, the CSF1R inhibitor is AJUD010. In one embodiment, the CSF1R inhibitor is AMG820. In another embodiment, the CSF1R inhibitor is IMC-CS4. In yet another embodiment, the CSF1R inhibitor is JNJ-40346527. In one embodiment, the CSF1R inhibitor is PLX5622. In another embodiment, the CSF1R inhibitor is FPA008.

[0265] In some embodiments, the checkpoint inhibitor is a therapeutic agent targeting HLA. In some embodiments, the therapeutic agent targeting HLA is an anti-HLA antibody. In one embodiment, the therapeutic agent targeting HLA is GSK01. In another embodiment, the therapeutic agent targeting HLA is IMC-C103C. In yet another embodiment, the therapeutic agent targeting HLA is IMC-F106C. In one embodiment, the therapeutic agent targeting HLA is IMC-G107C. In yet another embodiment, the therapeutic agent targeting HLA is ABBV-184.

[0266] The methods described herein may be used, when appropriate, in combination with one or more second active agents as described herein to treat diseases as understood herein and in the art to which they pertain.

[0267] PD-1 antagonists and pembrolizumab

[0268] This article provides methods for treating various cancers in subjects using a combination of an antibody-drug conjugate (ADC) binding to PD-L1 and a PD-1 antagonist. In some embodiments, the PD-1 antagonist is an anti-PD-1 antibody. In further embodiments, the anti-PD-1 antibody is pembrolizumab. In some embodiments, the PD-1 antagonist is selected from the group consisting of pembrolizumab, nivolumab, cimiprimab, durvalumab, atezolizumab, dostarlimab, avelumab, or pidilizumab. In some embodiments, the treatment is first-line therapy. In other embodiments, the treatment is second-line therapy.

[0269] “Pembrolizumab” or “pembro” is a humanized IgG4 mAb having the structure described in WHO Drug Information, Vol. 27, No. 2, pages 161-162 (2013). Pembrolizumab has the heavy chain CDR1 of SEQ ID NO: 17, the heavy chain CDR2 of SEQ ID NO: 18, the heavy chain CDR3 of SEQ ID NO: 19, the light chain CDR1 of SEQ ID NO: 20, the light chain CDR2 of SEQ ID NO: 21, and the light chain CDR3 of SEQ ID NO: 22. Pembrolizumab has the heavy chain sequence of SEQ ID NO: 23 and the light chain sequence of SEQ ID NO: 24. Pembrolizumab has the heavy chain variable region sequence of SEQ ID NO: 25 and the light chain variable region sequence of SEQ ID NO: 26.

[0270] In another embodiment, the PD-1 antagonist is an antibody or antigen-binding protein comprising a heavy chain variable region or a light chain variable region having at least 95%, 90%, 85%, 80%, 75%, or 50% sequence identity with SEQ ID NO:25 or SEQ ID NO:26, respectively, and exhibiting specific binding to PD-1. In another embodiment, the PD-1 antagonist is an antibody or antigen-binding protein comprising a heavy chain variable region or a light chain variable region having at most 1, 2, 3, 4, or 5 or more amino acid substitutions for SEQ ID NO:25 or SEQ ID NO:26, respectively, and exhibiting specific binding to PD-1.

[0271] In some embodiments, the PD-1 antagonist is administered after the ADC provided herein. In other embodiments, the PD-1 antagonist is administered concurrently with the ADC provided herein (e.g., during the same dosing period). In still other embodiments, the PD-1 antagonist is administered after the ADC provided herein.

[0272] In some embodiments, the amount of PD-1 antagonist used in the various methods provided herein can be determined using standard clinical techniques. In some embodiments, the amount of PD-1 antagonist (e.g., pembrolizumab) used in the various methods is provided in the examples.

[0273] In some embodiments, the subjects who can be treated with the methods provided herein are mammals. In some embodiments, the subjects who can be treated with the methods provided herein are humans.

[0274] IX. Products and Reagent Kits

[0275] In some embodiments, the subjects who can be treated by the methods provided herein are mammals. In some embodiments, the subjects who can be treated by the methods provided herein are humans. In another aspect, an article or kit comprising the anti-PD-L1 antibody or anti-PD-L1 antibody-drug conjugate described herein is provided. In the methods of the present invention, the article or kit may further comprise instructions for use of the anti-PD-L1 antibody or anti-PD-L1 antibody-drug conjugate described herein. Thus, in some embodiments, the article or kit comprises instructions for use of the anti-PD-L1 antibody or anti-PD-L1 antibody-drug conjugate described herein in treating a subject with cancer (e.g., melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), head and neck cancer, triple-negative breast cancer (TNBC), ovarian cancer, urothelial carcinoma, hepatocellular carcinoma (HCC), gastric cancer, and cervical cancer), comprising administering an effective amount of the anti-PD-L1 antibody or anti-PD-L1 antibody-drug conjugate described herein to the subject. In some embodiments, the subject is a human.

[0276] The product or kit may further include a container. Suitable containers include, for example, bottles, vials (e.g., dual-chamber vials), syringes (such as single-chamber or dual-chamber syringes), and test tubes. In some embodiments, the container is a vial. The container can be formed from a variety of materials (such as glass or plastic). The container holds the formulation.

[0277] The product or kit may further include a label or instruction leaflet located on or attached to the container, which may indicate guidance on formulation reconfiguration and / or use. The label or instruction leaflet may further indicate that the formulation is intended for use subcutaneously, intravenously (e.g., intravenous infusion), or by other modes of administration for the treatment of a subject's cancer (e.g., melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), head and neck cancer, triple-negative breast cancer (TNBC), ovarian cancer, urothelial carcinoma, hepatocellular carcinoma (HCC), gastric cancer, and cervical cancer). The container holding the formulation may be a single-use vial or a reusable vial, which allows for repeated administration of the reconfigured formulation. The product or kit may further include a second container containing a suitable diluent. The product or kit may further include other materials required from a commercial, therapeutic, and user perspective, including additional buffers, diluents, filters, needles, syringes, and an instruction leaflet containing instructions for use.

[0278] The article or kit may optionally further include a container containing a second agent, wherein the anti-PD-L1 antibody or anti-PD-L1 antibody-drug conjugate is the first agent, and the article or kit further includes instructions on a label or package insert for treating a subject with an effective amount of the second agent. In some embodiments, the second agent is used to eliminate one or more adverse events or reduce their severity.

[0279] In some embodiments, the anti-PD-L1 antibody or anti-PD-L1 antibody-drug conjugate is present in the container as a lyophilized powder. In some embodiments, the lyophilized powder is in a tightly sealed container, such as a vial, ampoule, or sachet indicating the amount of active agent. When administering the medicine by injection, ampoules of sterile water or saline for injection may be optionally provided as part of the kit, allowing the components to be mixed prior to administration. If desired, such kits may further include one or more of a variety of conventional pharmaceutical components, such as containers having one or more pharmaceutically acceptable carriers, additional containers, etc., as will be apparent to a person skilled in the art. The kit may also include printed instructions in the form of a package insert or label indicating the amount of the component to be administered, instructions for administration, and / or instructions for mixing the components.

[0280] In some embodiments, the anti-PD-L1 antibody-drug conjugate is a sterile, preservative-free, white to off-white cake or powder supplied in single-dose glass vials for reconstitution for intravenous administration. The formulation may contain pharmacopoeia-compliant excipients. Appropriate packaging and storage conditions will be provided for the drug. In some embodiments, the anti-PD-L1 antibody-drug conjugate can be reconstituted with 4.4 mL of sterile water for injection (compliant with USP, Ph. Eur., or equivalent). In some embodiments, for administration, the reconstituted solution is diluted with 5% dextran injection (USP or equivalent) prior to intravenous infusion.

[0281] X. Other Applications

[0282] The anti-PD-L1 antibodies described herein (such as humanized anti-PD-L1 antibodies) can be used to detect PD-L1 in the context of clinical diagnosis or treatment or in research. PD-L1 expression in cancer provides an indication that the cancer is suitable for treatment with the antibodies of this invention. The antibodies are also available as research reagents for laboratory studies to detect PD-L1-carrying cells and their responses to various stimuli. In such uses, monoclonal antibodies can be labeled with fluorescent molecules, spin-labeled molecules, enzymes, or radioisotopes, and can be provided in the form of kits containing all the reagents necessary for PD-L1 assays. The antibodies described herein can be used to detect PD-L1 protein expression and determine whether cancer is suitable for treatment with PD-L1 ADCs.

[0283] All patent applications, websites, other publications, registration numbers, etc., cited above or below are incorporated by full reference for all purposes, as if each individual item were specifically and individually indicated to be incorporated by reference in this manner. Where different versions of a sequence are associated with registration numbers at different times, it means that the version is associated with the registration number at the effective filing date of this application. The effective filing date means the earlier of the actual filing date that mentions the registration number (if applicable) or the filing date of the priority application. Similarly, where different versions of publications, websites, etc., are published at different times, unless otherwise stated, it refers to the version published closest to the effective filing date of that application. Unless otherwise specifically stated, any feature, step, component, embodiment, or aspect of the invention may be used in combination with any other feature, step, component, embodiment, or aspect. Although the invention has been described somewhat in detail by way of illustration and examples for purposes of clarity and understanding, it is clear that certain changes and modifications may be made within the scope of the appended claims.

[0284] Example

[0285] Example 1: Phase 1 study of SGN-PDL1V in advanced solid tumors

[0286] A phase 1, open-label, multicenter study was designed to evaluate the safety, tolerability, pharmacokinetic (PK), and antitumor activity of SGN-PDL1V as monotherapy or in combination with pembrolizumab in adults with selected advanced solid tumors. SGN-PDL1V is an ADC comprising an anti-PD-L1 antibody conjugated to MMAE. The anti-PD-L1 antibody comprises a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 of SEQ ID NOs: 5, 6, and 7, respectively; and a light chain variable region comprising light chain CDR1, CDR2, and CDR3 of SEQ ID NOs: 8, 9, and 10, respectively. The anti-PD-L1 antibody also comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 3 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 4. The anti-PD-L1 antibody also comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 1 and a light chain comprising the amino acid sequence of SEQ ID NO: 2.

[0287] The combination of SGN-PDL1V, which binds to PD-L1 on the cell surface, with PD-1 checkpoint inhibitors such as pembrolizumab offers unique advantages. SGN-PDL1V induces direct cytotoxicity against PD-L1-positive cells, reducing overall immunosuppressive PD-1 / PD-L1 signaling in the tumor microenvironment and thus driving increased cytotoxic T cell activity. SGN-PDL1V can also initiate cancer immune processes through MMAE-induced immunogenic cell death. Ultimately, SGN-PDL1V has demonstrated promising clinical activity in patients with relapsed / refractory solid tumors to checkpoint inhibitors such as pembrolizumab, suggesting that the combination of PD-L1+ tumor debulking by SGN-PDL1V and PD-(L)1 blockade by pembrolizumab can provide patients with additional therapeutic benefits beyond pembrolizumab alone.

[0288] This study includes dose escalation for multiple tumor types (Part A), a cohort for subsequent dose and schedule optimization (Part B, as needed), a disease-specific cohort for dose-expanded monotherapy (Part C), and a run-in cohort for combination safety with pembrolizumab (Part D).

[0289] This study includes dose escalation (Part A), which will recruit subjects with HNSCC, non-small cell lung cancer (NSCLC), triple-negative breast cancer (TNBC), or esophageal squamous cell carcinoma (SCC) who have programmed death-ligand 1 (PD-L1) expression ≥1 based on a composite positive score (CPS), tumor proportion score (TPS), or immune cell score (% IC). The initial dosing schedule to be tested is days 1 and 8 of a 21-day cycle (2Q3W). In the 2Q3W schedule, the initial dose level to be evaluated is 0.5 mg / kg. Dose escalation will be performed in increments of 0.25 mg / kg, up to a dose level of 2.5 mg / kg (if tolerated). Alternative dosing schedules may be initiated at any time with additional subjects (e.g., days 1, 8, and 15 of a 28-day cycle [3Q4W]; or days 1 and 15 of a 28-day cycle [2Q4W]; or day 1 of a 21-day cycle [Q3W]). Provided the dose intensity is the same or lower, the initial dose of the alternative dosing schedule is the same as the permitted dose in the initial dosing schedule (i.e., 2Q3W). The modified toxicity probability interval (mTPI) dose escalation rule is applied individually to each dosing schedule.

[0290] Part B can be initiated to further evaluate the recommended SGN-PDL1V dosing schedules for different tumor types in Part A. This part will allow for the optimization of the recommended dosing and schedules for expansion. The selection of open-enrollment (multiple) schedules will be determined by the trial commissioner in consultation with the Safety Monitoring Committee (SMC) based on safety, dose-limiting toxicity (DLT), PK, and available data on initial antitumor activity. Part B can recruit subjects with selected tumor types who meet the eligibility criteria described in Part A. Different doses / schedules for different tumor types can be evaluated in a parallel cohort manner. Subjects participating in Part B will be randomly assigned to different doses and schedules with equal probability for their specific tumor type. If sufficient evidence exists after Part A to support continuing a particular SGN-PDL1V dosing schedule, Part B can be omitted, provided that at least 6 subjects are dosed with the recommended dose and schedule.

[0291] Once the recommended amplification dose and schedule are selected by SMC, dose expansion with a specific disease amplification cohort (Part C) and combination safety introduction with SGN-PDL1V and pembrolizumab (Part D) can be initiated to further evaluate the safety, tolerability, and antitumor activity of SGN-PDL1V.

[0292] During dose amplification (Part C), the following cohorts can be evaluated:

[0293] HNSCC; PD-L1 expression is not considered.

[0294] NSCLC; PD-L1 expression not considered.

[0295] Additional signal search cohort in subjects with melanoma, ovarian cancer, TNBC, gastric cancer, or esophageal SCC.

[0296] Biological cohorts can also be initiated, including PD-L1 negative NSCLC and HNSCC, TNBC, esophageal SCC, melanoma, gastric cancer, or ovarian cancer.

[0297] During Part D, the combination of SGN-PDL1V and pembrolizumab will be evaluated in first-line (1L) HNSCC patients with PD-L1 expression ≥1 based on TPS or CPS. Approximately 12 subjects will be recruited in this cohort. To assess the safety of the combination therapy, the initial 6 subjects will be administered SGN-PDL1V plus pembrolizumab. Dosing decisions for subsequent subjects in the cohort will be based on the incidence of DLT. If the SMC deems the combination of SGN-PDL1V and pembrolizumab safe, a future protocol amendment may expand this cohort to approximately 40 subjects. Additional combinations may also be explored in future protocol amendments.

[0298] In Part D of this study, the planned dose of pembrolizumab was 200 mg IV every 3 weeks (Q3W). Based on the overall data generated in the pembrolizumab development program, 200 mg Q3W is the appropriate dose of pembrolizumab for adults across all indications.

[0299] The 200 mg Q3W dose is justified for the following reasons:

[0300] Clinical data from eight randomized studies of melanoma and non-small cell lung cancer (NSCLC) tumor types demonstrated a flat dose- and exposure-efficacy relationship from 2 mg / kg Q3W to 10 mg / kg every 2 weeks (Q2W), representing an exposure range of approximately 5 to 7.5 times (refer to pembrolizumab IB).

[0301] Population pharmacokinetic (PK) analysis showed that both fixed-dose and weight-based dosing provided similar control of PK variability and had considerable overlap in exposure distribution, supporting the applicability of 200 mg Q3W.

[0302] Clinical data showed improved benefit-risk (including overall survival) across multiple tumor types at 200 mg Q3W, and

[0303] Pharmacological data showed that at 200 mg Q3W, full target saturation was achieved in both systemic circulation (inferred from PK data) and tumors (inferred from physiological PK [PBPK] analysis).

[0304] A total of 2262 patients with melanoma and NSCLC were recruited across eight randomized dose-comparison studies, covering different disease scenarios (treatment-naïve, previously treated, PD-L1 enriched, and all patients) and different treatment profiles (monotherapy and combination therapy with chemotherapy). Five studies compared 2 mg / kg Q3W with 10 mg / kg Q2W (KN001 cohort B2, KN001 cohort D, KN002, KN010, and KN021), and three studies compared 10 mg / kg Q3W with 10 mg / kg Q2W (KN001 cohort B3, KN001 cohort F2, and KN006). All of these studies demonstrated a flat dose- and exposure-response relationship across the studied doses, representing a difference of approximately 5 to 7.5 times in exposure. 2 mg / kg (or a fixed dose of 200 mg) Q3W provided a similar response to the highest dose studied. Subsequently, a flat dose-exposure-response relationship was also observed in other tumor types, including head and neck squamous cell carcinoma (HNSCC), bladder cancer, gastric cancer, and classic Hodgkin lymphoma, confirming that 200 mg Q3W is an appropriate dose independent of tumor type. These findings are consistent with the MOA of pembrolizumab, which acts by interacting with immune cells rather than by directly binding to cancer cells.

[0305] Furthermore, pharmacological data showed target saturation at 200 mg Q3W. First, PK data in KN001, evaluating target-mediated drug disposition, ultimately demonstrated PD-1 saturation in systemic circulation at doses significantly lower than 200 mg Q3W. Second, PBPK analysis was performed to predict tumor PD-1 saturation across extensive tumor penetration and expression. This assessment concluded that pembrolizumab at 200 mg Q3W achieved complete PD-1 saturation in both hematology and tumors.

[0306] Finally, population PK analysis of pembrolizumab (characterizing the effect of body weight and other patient covariates on exposure) showed that fixed dosing provided similar control of PK variability as weight-based dosing, and that the exposure distributions from the 200 mg Q3W fixed dose and the 2 mg / kg Q3W dose had considerable overlap. Supported by these PK characteristics, and considering the advantages of fixed dosing in reducing dosing complexity and potential dosing errors, the 200 mg Q3W fixed dose was chosen for evaluation across all pembrolizumab regimens.

[0307] Benefits / Risk Assessment

[0308] ADCs (such as SGN-PDL1V) have potential antitumor activity in relapsed and refractory solid tumors and could benefit a patient population with poor prognosis who have exhausted all other available treatment options. The toxicities observed in a preclinical study of SGN-PDL1V were largely consistent with MMAE toxicities previously reported with other vedotin ADCs, and no target-related toxicities were observed. This preclinical study represents the potential toxicities of SGN-PDL1V and supports evaluation in human clinical studies. This patient population requires novel treatment options with potential benefits and manageable, monitorable risks.

[0309] Experimental objectives and purposes

[0310] This study will evaluate the safety, tolerability, pharmacokinetic (PK), and antitumor activity of SGN-PDL1V (monotherapy) in subjects with advanced solid malignancies and in combination with pembrolizumab in subjects with metastatic or unresectable HNSCC. Table 1 summarizes the specific objectives and corresponding endpoints of this study.

[0311] Table 1: Targets and Corresponding Endpoints

[0312]

[0313]

[0314] ac-MMAE = Antibody-conjugated monomethylolpropionate E; ADA = Anti-drug antibody; AE = Adverse reaction; DLT = Dose-limiting toxicity; DOR = Duration of response; EORTC = European Organisation for Research and Treatment of Cancer; HNSCC = Head and neck squamous cell carcinoma; iRECIST: Modified RECIST 1.1 for immunotherapy agents; MMAE = Monomethylolpropionate E; MTD = Maximum tolerated dose; ORR = Objective response rate; OS = Overall survival; PFS = Progression-free survival; PK / PD = Pharmacokinetics / Pharmacodynamics; PRO = Patient-reported outcome; QLQ = Quality of life questionnaire (core and head and neck); RECIST v1.1 = Standards for assessing response to solid tumors, version 1.1.

[0315] Research Plan

[0316] Overall research design

[0317] This is a phase 1, open-label, dose-escalation and dose-expansion multicenter study evaluating the safety and tolerability of SGN-PDL1V monotherapy and combination therapy in adults with histologically or cytologically confirmed metastatic or unresectable solid tumors that are recurrent or refractory or intolerant to SoC. Once the safety and tolerability of SGN-PDL1V (dose escalation and dose / timetable optimization, Part A and Part B, respectively) are established, and the SMC has recommended expanded dose and timetable, SGN-PDL1V will be evaluated in Part C as monotherapy and in combination with pembrolizumab (Part D). Figure 1 Present the overall research design.

[0318] In Part A, approximately 80 subjects are expected to be evaluated in the dose escalation phase of this study. The dose escalation phase of this study will be conducted using a modified probability toxicity interval (mTPI) design to assess the safety and tolerability of SGN-PDL1V and identify its mean toxicity dose (MTD). Initially, the dosing schedule for testing will be days 1 and 8 of a 21-day cycle (2Q3W). Alternative dosing schedules may be initiated at any time with additional subjects (e.g., days 1, 8, and 15 of a 28-day cycle [3Q4W]; or days 1 and 15 of a 28-day cycle [2Q4W]; or day 1 of a 21-day cycle [Q3W]). The mTPI dose escalation rule will be applied individually to each dosing schedule. Not all alternative dosing schedules will be evaluated.

[0319] After dose escalation is complete, dose and schedule optimization (Part B) can be initiated following consultation with the SMC to further evaluate the different SGN-PDL1V doses and / or schedules recommended in Part A. This part will allow for the optimization of the dose and schedule recommendations used for expansion. Subjects meeting the eligibility criteria described in Part A can be recruited for Part B. Different doses and schedules for different tumor types can be evaluated in a parallel cohort manner. Subjects participating in Part B will be randomly assigned to different doses and schedules with equal probability for their specific tumor type. If sufficient evidence exists after Part A to support continuing a particular SGN-PDL1V dosing schedule, Part B can be omitted, provided that at least six subjects are dosed at the recommended dose and schedule.

[0320] Figure 2 Preliminary patient data are shown for all subjects in both dose escalation (Part A) and dose optimization (Part B). The uORR is 23%. Figure 3 This presents preliminary patient data for HNSCC subjects in dose escalation (≥1.25 mg / kg, Part A), with a cORR of 21% and an uORR of 50%.

[0321] Dose escalation (Part C) will be performed using selected disease-specific cohorts. Following completion of dose escalation (Part A) and dose and schedule optimization (Part B, if necessary), a disease-specific cohort for HNSCC, excluding PD-L1 expression, may be initiated by the trial commissioner in consultation with the SMC. Additional disease-specific cohorts for NSCLC, excluding PD-L1 expression, may also be initiated. Additional signal search cohorts may be initiated for subjects with melanoma, ovarian cancer, triple-negative breast cancer (TNBC), gastric cancer, or esophageal squamous cell carcinoma (SCC), and biological cohorts may also be initiated. The doses to be examined in Part C will be equal to or lower than the MTD and / or recommended dose and schedule determined in Parts A and B.

[0322] The combination safety initiation (Part D) will recruit approximately 12 participants to evaluate the combination of SGN-PDL1V and pembrolizumab. To assess the safety of the combination therapy, the initial 6 participants will receive SGN-PDL1V plus pembrolizumab. Dosing decisions for subsequent participants in the cohort will be based on the incidence of DLT.

[0323] In Part D, SGN-PDL1V (starting at a dose one level lower than the recommended monotherapy dose identified in Parts A and / or B) is combined with the approved dose of standard pembrolizumab in a Q3-week cycle. Part D will be initiated once the SMC determines that the recommended expanded monotherapy dose and schedule are safe and tolerable. If the SMC deems the combination of SGN-PDL1V and pembrolizumab safe, future protocol revisions may expand this cohort to approximately 40 subjects (Part E). Additional combinations may also be explored in future protocol revisions.

[0324] Tumor types included in dose escalation (Part A) and dose and schedule optimization (Part B)

[0325] For dose escalation (Part A) and dose and schedule optimization (Part B), subjects must have NSCLC, HNSCC, TNBC, or esophageal SCC. Additionally, all subjects must have PD-L1 expression ≥1 based on historical testing, using tumor proportion score (TPS), CPS, or immune cell score (%IC).

[0326] Part A: Dose Elevation Cohort

[0327] Approximately 80 subjects are expected to be evaluated in the dose escalation phase of this trial. Dose escalation will be conducted using an mTPI design to assess the safety and tolerability of SGN-PDL1V and to identify its MTD and recommended dose. Safety, pharmacodynamics, biomarker analysis, and preliminary antitumor activity will be used to determine the recommended dose and schedule.

[0328] The mTPI design uses a Bayesian model to calculate the posterior probability of the three-time interval, reflecting the relative distance between the toxicity rate at each dose level and the target dose-limiting toxicity (DLT) rate. The dosing-decision rule is determined with a target DLT rate of 25% and a 5% margin. The three-time intervals will be (0%, 20%), [20%, 30%], and (30%, 100%), and the corresponding dose-decision rules will be:

[0329] 1. If the current dose-dependent leukemia (DLT) rate is likely to be <20%, then increase the dose.

[0330] 2. If the current dose-dependent DLT rate is likely between 20% and 30%, maintain that level.

[0331] 3. If the current dose may result in a DLT rate >30%, then reduce the dose.

[0332] The dose-finding decisions are shown in Table 2. “E” indicates increasing the dose, “S” indicates maintaining the same dose, and “D” indicates decreasing the dose. The decision “DU” means that the current dose level is unacceptable due to high toxicity. If the posterior probability of a DLT rate higher than 25% is greater than 95%, the dose will be defined as having unacceptable toxicity.

[0333] Part A recruitment will proceed on a cohort-by-cohort basis. Decisions regarding dose escalation, dose level modifications, and subsequent cohort size will be made by the trial commissioner in consultation with the SMC. Dose levels will not be skipped. When initially assessing a dose level, subjects in that cohort must be observed throughout the entire DLT period before recruitment for higher doses can proceed. Note: For each new dose-escalation cohort initiated as recommended by the SMC, no more than one subject should receive treatment within a 24-hour period.

[0334] At each dose level, at least two subjects with DLT-evaluable (DE) will be treated until the first DLT is observed. After the first DLT is observed, at least three DE subjects will be required at each dose level before escalation to all higher doses. Subjects deemed DLT-unevaluable during Cycle 1 may be replaced. At least six DE subjects will be observed at the estimated MTD or recommended dose before being determined. The MTD will be estimated based on data from all subjects across all evaluable doses. If the MTD is not achieved, the recommended dose will be determined using safety, pharmacodynamics, and biomarker analyses, as well as preliminary antitumor activity. Decreasing to lower or intermediate dose levels may be performed at any time after consultation between the trial commissioner and the SMC.

[0335] During dose escalation, additional subjects may be recruited at a test dose level that is considered tolerable (hereinafter referred to as "backfilling"). DLT observed at lower dose levels will be considered together with all data; SMC will make recommendations for future escalations based on the current and lower dose levels at which DLT was observed in backfilled subjects, using the mTPI model.

[0336] Subjects with solid tumors may continue treatment until disease progression (PD), clinical deterioration, unacceptable toxicity, withdrawal of consent, or study termination, according to the Solid Tumor Response Assessment Criteria version 1.1 (RECIST v1.1), whichever occurs first.

[0337] Table 2: Trial Calculation Table for Dose Discovery in mTPI Design

[0338]

[0339] D = Decrease to the next lower dose; DLT = Dose-limiting toxicity; DU = Current dose is unacceptably toxic; E = Increase to the next higher dose; mTPI = Modified probability interval for toxicity; S = Maintain at the current dose.

[0340] Initially, SGN-PDL1V will be administered at the dose levels shown in Table 3 in a 2Q3W schedule. An alternative dosing schedule may be initiated at any time (Table 4). As long as the dose intensity is the same or lower, the initial dose of the alternative dosing schedule will be the same as the permitted dose in the initial pattern. The mTPI dose escalation rule will be applied individually to each dosing schedule. Not all alternative dosing schedules require evaluation.

[0341] Part B: Dosage and Schedule Optimization

[0342] Dosage and schedule optimization can be initiated after consultation with the SMC to evaluate the SGN-PDL1V dosing schedule recommended in Part A. This part will allow for the optimization of the recommended dosage and schedule for expansion. Part B can be used to recruit subjects with selected tumor types who meet the eligibility criteria described in Part A. Approximately 80 subjects will be evaluated in Part B. Subjects participating in Part B will be randomly assigned with equal probability to different dosages and schedules for their specific tumor type.

[0343] If, after completing Part A, there is sufficient evidence to support continuing with a particular dosage and schedule, Part B may be omitted, provided that at least six subjects are administered the recommended dosage and schedule. SMC will make recommendations based on the data from Parts A and B, suggesting dosages and schedules to be evaluated in Parts C and D (and Part E). The trial client will make the final decision regarding the dosage and schedule.

[0344] Table 3: Planned Dosage Levels

[0345]

[0346] 2Q3W = Day 1 & 8 q21; 3Q4W = Day 1 & 8 & 15 q28; 2Q4W = Day 1 & 15 q28; Q3W = Day 1 q21.

[0347] Table 4: Schedule of Initial and Alternative Treatments

[0348]

[0349] Collect and analyze the preliminary results of Part A and Part B.

[0350] Table 5 shows the preliminary baseline characteristics of patients recruited in the dose escalation and optimization cohorts, consistent with the study cohort of patients with PD-L1+ tumors who received extensive prior therapy. The median number of prior treatment lines received by patients was 3.

[0351] Table 5: Baseline characteristics of patients

[0352]

[0353] (CPI: Checkpoint Inhibitor; ECOG: East Coast Oncology Collaboration)

[0354] Table 6 presents a preliminary summary of adverse events, showing that PDL1V has a manageable safety profile overall, with a small number of advanced treatment-related AEs. No dose-limiting toxicities or treatment-related deaths were observed across all dose levels (ranging from 0.5 to 1.75 mg / kg). Figure 4 As shown, most treatment-related adverse events (TRAEs) are low-grade (grade 1 / 2), and very few are grade 3 or 4 events.

[0355] Table 6: Occurrence of adverse events at indicated dose levels

[0356]

[0357] On days 1 and 8 of the 21-day cycle, use adjusted ideal body weight. AE: Adverse events; DLT: Dose-limiting toxicity; TEAE: Treatment-induced adverse events.

[0358] Figure 5 The cumulative effect of PDL1V was negligible across clinically active dose levels (1.25 mg / kg to 1.75 mg / kg) and when administered on days 1 and 8 of a 21-day cycle; the terminal elimination half-life was approximately 3.8 days. Figure 5 As shown, antibody-conjugated MMAEs (left panel) and unconjugated MMAEs (right panel) exhibit dose-proportional pharmacokinetics at the active dose level.

[0359] Figure 6Preliminary patient data for all subjects are shown in both dose escalation (Part A) and dose optimization (Part B). Tumor reductions were observed across tumor types, with the largest reductions observed in patients with NSCLC and HNSCC, indicated by green circles and purple triangles. Evidence of activity was observed at active dose levels of 1.25 mg / kg or higher. Table 7 shows what appears to be a dose-response relationship across tumor types, with higher confirmed objective response rates (cORR) observed at higher dose levels.

[0360] Table 7: Response rates at indicated dose levels

[0361]

[0362] Table 8 also shows that cORR was observed across different PD-L1 expression levels in both NSCLC and HNSCC. Figure 7 The clinical response across the active dose was durable, with a median duration of response of 7.9 months. The median follow-up period for patients was 9.4 months.

[0363] Table 8: Response rates in NSCLC or HNSCC patients with PDL-1-indicating expression

[0364]

[0365] As shown in Table 9, with NSCLC on the left and HNSCC on the right, the cORR was generally observed at higher dose levels in both tumor types. For NSCLC, the best overall response (ORR) was observed at 1.5 mg / kg. In HNSCC, the highest response was observed at 1.75 mg / kg (on a 21-day cycle, D1 and D8). However, due to the need for frequent dose modifications at this level, investigations are underway at 1.75 mg / kg (every weekly). Expansion cohorts at 1.5 mg / kg (on 21-day cycles, D1 and D8) have also been initiated in both NSCLC and HNSCC.

[0366] Table 9: Antitumor activity of PDL1V in NSCLC and HNSCC

[0367]

[0368] For 32 patients with PD-L1+ NSCLC Figure 8 Antitumor activity was observed across different histological subtypes, particularly in adenocarcinoma (~69% of samples), adenosquamous (marked with dark green diamonds), and squamous histological morphology (marked with orange diamonds). Furthermore, Figure 9These clinical responses were durable, with a median duration of response of 5.6 months. Among the 32 patients, the best cORR (33.3%) was observed in those receiving 1.5 mg / kg (2Q3W), the recommended dose for advancing to the dose expansion phase of this study.

[0369] Overall, the investigator-assessed objective response rate (ORR) across all doses and tumor types was 27.3% (12.7% confirmed), and the median duration of confirmed response was 7.9 months. Objective responses were observed starting at 1.25 mg / kg and were independent of PDL1 expression. Specifically, clinically meaningful and durable responses were observed in patients with severe, previously treated NSCLC and HNSCC.

[0370] Part C: Dosage Amplification

[0371] After determining the MTD or identifying the recommended dose, two dose amplification cohorts (Part C), a signal search cohort, and a biological cohort can be initiated. Approximately 150 subjects will receive treatment at or below the MTD or recommended dose in specific disease amplification cohorts to further characterize the safety, tolerability, PK, and antitumor activity of SGN-PDL1V.

[0372] The C-part queue may include:

[0373] HNSCC patients excluding those with PD-L1 expression (approximately 40 participants),

[0374] NSCLC patients excluding those with PD-L1 expression (approximately 40 participants),

[0375] Signal search cohort (approximately 40 subjects) with ovarian cancer, TNBC, gastric cancer, or esophageal SCC or melanoma.

[0376] Biological Cohort: Approximately 30 participants in the biological cohort of PD-L1-negative NSCLC and HNSCC, melanoma, ovarian cancer, TNBC, gastric cancer, or esophageal SCC will be gated based on data generated from other cohorts and will be required to undergo additional biopsies to provide additional tissue for the SGN-PDL1V biomarker study, which will compare tumor samples before and after treatment to characterize clinical MOA and the correlation between sensitivity / resistance at MTD or recommended dose. Participants recruited in the biological cohort must have tumors present at appropriate, palpable, and feasible biopsy sites.

[0377] For each expansion cohort with a planned sample size of approximately 40 participants, an interim analysis (IA) will be conducted after approximately 15 participants have achieved an evaluable response, while recruitment continues. Futility assessment will be performed using the predicted probability of success (PPoS) method, where success is defined as having a post-hoc probability greater than 0.80, i.e., a response rate greater than the background response rate. If the estimated PPoS for the given interim data is <10%, the trial sponsor and SMC consultant will subsequently decide to further halt recruitment for that cohort after careful evaluation of all data.

[0378] Part D: Import of Composite Security

[0379] Part D aims to evaluate the safety and tolerability of the combination of SGN-PDL1V and pembrolizumab. Approximately 12 subjects will be recruited in this safety induction cohort.

[0380] SGN-PDL1V will be administered starting at a dose level one level lower than the recommended monotherapy dose and according to a schedule determined by the trial commissioner based on the results of Parts A and B and after consultation with the SMC.

[0381] To assess the safety of this combination, the initial six subjects will be administered SGN-PDL1V plus pembrolizumab. Safety will be assessed by the incidence of DLT. Dosing decisions will be made using the following rules:

[0382] 1. The initial dose for the first 6 subjects will be one dose level lower than the recommended expansion dose identified in Part A and / or Part B plus 200 mg pembrolizumab Q3W.

[0383] i. If none or one of the initial 6 subjects has experienced DLT, the additional 6 subjects will receive the recommended expanded dose of SGN-PDL1V plus 200 mg pembrolizumab Q3W.

[0384] ii. If two or more of the initial six subjects (as described in 1) experience DLT, SMC will conduct a further data review and recommend whether to have approximately six additional subjects receive SGN-PDL1V plus 200 mg pembrolizumab Q3W at a dose level two lower than the recommended monotherapy amplification dose, or whether to consider ending the combination.

[0385] Treatment duration

[0386] Subjects may continue treatment with SGN-PDL1V until a disease progression (PD) is identified according to RECIST v1.1 (parts A, B, and C) or according to iRECIST (part D only), unacceptable toxicity, clinical deterioration, initiation of subsequent therapy, withdrawal of consent, death, or study termination, whichever occurs first. In part D, 200 mg of pembrolizumab will be administered in combination with SGN-PDL1V on a Q3-week dosing schedule for a maximum of 35 doses (approximately 2 years). If a subject in part D discontinues either study drug for any reason, that subject will be considered to have discontinued both study treatments.

[0387] Dose-limiting toxicity

[0388] DLT will be assessed during dose escalation in Part A for monotherapy and in Part D for combination therapy. The DLT assessment period will be the first cycle (21 or 28 days). DLT is defined as any of the following criteria associated with SGN-PDL1V monotherapy or combination therapy with pembrolizumab (not attributable solely to pembrolizumab) during the DLT assessment period, excluding toxicities clearly associated with disease progression or comorbidities. Grading will be based on the National Cancer Institute Common Terminology Guidelines for Adverse Events (NCI-CTCAE), version 5.0:

[0389] Level 5 toxicity

[0390] Grade 3 or 4 non-hematologic toxicity (non-laboratory), except for the following:

[0391] ○ Grade 3 fatigue or constipation relieved within 72 hours, with or without intervention.

[0392] ○ Grade 3 nausea / vomiting or diarrhea lasting ≤72 hours with adequate antiemetics and other supportive care.

[0393] ○ Level 3 IRR, regardless of duration

[0394] Note: If a Grade 3 IRR occurs in ≥20% of subjects (i.e., 2 or more of the first 10 subjects), all subsequent subjects will require premedication and / or adjustments to the infusion method and protocol as advised by the SMC. For subsequent subjects, any Grade 3 IRR will be considered a DLT.

[0395] Grade 3 or higher electrolyte / non-hematological laboratory abnormalities, except for those that last ≤72 hours, are clinically uncomplicated, and resolve spontaneously or respond to routine medical interventions.

[0396] Grade 3 thrombocytopenia accompanied by clinically significant bleeding

[0397] Grade 4 thrombocytopenia lasting >7 days

[0398] Grade 4 anemia

[0399] Grade 4 hematologic toxicity (excluding anemia), lasting >7 days

[0400] Grade 3 febrile neutropenia

[0401] Dosage delay ≥14 days (attributed to toxicity)

[0402] Basic Principles of Research Design

[0403] This first-in-human (FIH), Phase 1 study, comprising dose escalation (Part A), dose and schedule optimization (Part B), dose expansion with a signal search cohort and a biological cohort (Part C), and combination therapy with pembrolizumab (Part D), aims to evaluate the safety and tolerability of SGN-PDL1V initially administered at 2Q3W in subjects with selected advanced solid tumors, and to estimate the mean time to treatment (MTD) and / or determine the recommended dose and schedule. Preliminary clinical development of SGN-PDL1V will involve evaluation in subjects of candidates for SGN-PDL1V administration who do not have appropriate standard treatment options and are administered SGN-PDL1V based on the treating physician's opinion. Dose escalation will be used to estimate the MTD and / or determine the recommended dose or schedule for SGN-PDL1V. Safety assessments may include the type, incidence, severity, and relevance of adverse events (AEs) and laboratory abnormalities, the incidence of drug-induced leukemia (DLT), and detection of anti-drug antibodies (ADAs). Part B may be opened for dose and schedule optimization and may recruit approximately 80 subjects.

[0404] Once the amplification dose and schedule are selected by the SMC, dose-based amplification with up to four specific disease amplification cohorts can be recruited (Part C), and combination safety delivery with SGN-PDL1V and pembrolizumab can be implemented (Part D). Figure 1 ).

[0405] Basic principles of dose selection

[0406] The starting dose of SGN-PDL1V at 0.5 mg / kg is based on data from a GLP nonclinical toxicology study in cynomolgus monkeys and is considered a reasonable starting dose that may result in minimal toxicity associated with MMAE delivery of SGN-PDL1V while maintaining potential pharmacological activity, consistent with ICH guidelines S6 and S9. Subsequent dose levels used in Parts C and D will be based on clinical data from Parts A and / or B, as well as recommendations from the SMC.

[0407] In Part D of this study, the planned dose of pembrolizumab is 200 mg Q3W, which is the approved dose of pembrolizumab for adults across all indications (see Pembrolizumab IB / Product Information).

[0408] Subject selection and withdrawal

[0409] Participants must meet all eligibility criteria for recruitment in this study. Researchers may not waive eligibility criteria and must review them under good clinical practice audits and / or health administration facility inspections.

[0410] Inclusion Criteria

[0411] 1. Participants must provide written informed consent.

[0412] 1. Tumor type:

[0413] i. Dosage escalation (Part A) and dosage and schedule optimization (Part B):

[0414] Subjects must have histologically or cytologically confirmed metastatic or unresectable HNSCC, NSCLC, TNBC, or esophageal SCC.

[0415] Subjects must have relapsed or refractory disease, have worsened to approved therapies, be intolerant to or refuse such therapies, or have contraindications to such therapies and, in the researcher's judgment, there should be no suitable SoC treatment option. If an available SoC therapy has not yet been administered, the reason for unsuitability (e.g., intolerance or refusal) must be documented.

[0416] In France, participants are eligible only if they experience PD after receiving local SoC treatment or if they are intolerant to existing therapies.

[0417] All subjects in dose escalation must have PD-L1 expression ≥1 based on historical test results according to TPS, CPS, or % IC.

[0418] For subjects with locally advanced / metastatic NSCLC, the determination of tumor genomic alterations (including but not limited to sensitized epidermal growth factor receptor (EGFR) mutations, anaplastic lymphoma kinase (ALK) rearrangements, and ROS1 arrangements) that are targeted by approved and available targeted agents must be made before recruitment.

[0419] Unless there are contraindications or the subject has experienced intolerance due to toxicity, subjects with such tumor genomic aberrations must have received an approved mutation-specific targeted therapy and have experienced disease progression (PD).

[0420] Unless the treatment is contraindicated or the subject has experienced intolerance due to toxicity, subjects with tumor genomic alterations not targeted by an approved targeted agent must have received dual regimens (simultaneously or sequentially) of anti-PD-1 or anti-PD-L1 therapy (if available) and platinum-based chemotherapy and have deteriorated against those regimens.

[0421] Unless there are contraindications to treatment or the subject has experienced intolerance due to toxicity, subjects with HNSCC must have received anti-PD-1 therapy (if available) and platinum-based chemotherapy regimens (simultaneously or sequentially) and have experienced exacerbations to those regimens.

[0422] Tumor tissue is required at the time of recruitment. Archived tumor tissue collected within 24 months prior to recruitment should be provided. If no archived tissue is available, a newly obtained biopsy must be submitted.

[0423] ii. Dosage escalation (Part C):

[0424] Unless there are contraindications, subjects must have relapsed or refractory disease or be intolerant to SoC therapy, as follows:

[0425] HNSCC; PD-L1 expression is not considered.

[0426] ○ Subjects must have histologically or cytologically confirmed HNSCC.

[0427] ○ The primary tumor must originate from the oral cavity, oropharynx, hypopharynx, nasopharynx, or larynx.

[0428] Unless there are contraindications to treatment or the subject has experienced intolerance due to toxicity, the subject must have received prior anti-PD-1 therapy (if eligible) and a platinum-based chemotherapy regimen (simultaneously or sequentially) with progression. The subject must not have received >1 prior line of cytotoxic therapy in locally advanced or metastatic cases. Platinum regimens administered as part of multimodal therapy in curative cases will be counted as one regimen if relapse occurs within ≤6 months after completion.

[0429] ○ Subjects must have previously undergone PD-L1 expression testing and, based on historical testing, should have PD-L1 expression ≥1 or <1 according to TPS or CPS.

[0430] ○ Tumor tissue is required at the time of recruitment. The most recently archived tumor tissue collected within 24 months of recruitment should be provided. If no archived tissue is available, a newly obtained biopsy must be submitted.

[0431] NSCLC; PD-L1 expression not considered:

[0432] ○ Subjects must have histologically or cytologically confirmed NSCLC (stage IIIB, IIIC, IV). Subjects with both SCC and non-SCC histology are eligible. Note: Subjects with neuroendocrine components or histological structures are ineligible.

[0433] For subjects with NSCLC, the determination of tumor genomic alterations (including but not limited to sensitizing EGFR mutations, ALK rearrangements, and ROS1 arrangements) that are targeted by approved and available targeted agents must be made before recruitment.

[0434] - Unless there are contraindications or the subject has experienced intolerance due to toxicity, subjects with such tumor genomic aberrations must have received no more than two approved mutation-specific targeted therapies and have experienced disease progression (PD). Subjects may also have received up to one PD-L1 or anti-PD-1 therapy (concurrent or sequential with platinum-based chemotherapy).

[0435] - Unless the treatment is contraindicated or the subject has experienced intolerance due to toxicity, subjects without approved targeted agents targeting tumor genomic alterations must have received prior anti-PD-1 or anti-PDL1 therapy (if available) and a platinum-based chemotherapy regimen (concurrently or sequentially) and experienced progression / relapse. Subjects must not have received >2 prior lines of cytotoxic therapy. For early or locally advanced disease, subjects who experienced progression or relapse within ≤6 months of the last dose of platinum-based therapy in adjuvant, neoadjuvant, or concomitant radiochemotherapy regimens are eligible.

[0436] - Subjects must have previously undergone PD-L1 expression testing, and based on historical testing, their PD-L1 expression should be ≥1 or <1 according to CPS or TPS.

[0437] Tumor tissue is required at the time of recruitment. The most recently archived tumor tissue collected within 24 months of recruitment should be provided. If no archived tissue is available, a newly obtained biopsy must be submitted.

[0438] Signal search queue:

[0439] - Patients with TNBC, ovarian cancer, esophageal SCC, stomach cancer, or melanoma.

[0440] - Subjects must have relapsed or refractory disease, have worsened to approved therapies, be intolerant to or refuse such therapies, or have contraindications to such therapies and, in the researcher's judgment, there should be no suitable SoC treatment option. If an available SoC therapy has not yet been administered, the reason for unsuitability (e.g., intolerance or refusal) must be documented.

[0441] - Subjects in France are eligible only if they experience PD after receiving local SoC treatment or if they are intolerant to existing therapies.

[0442] Tumor tissue is required at the time of recruitment. The most recently archived tumor tissue collected within 24 months of recruitment should be provided. If no archived tissue is available, a newly obtained biopsy must be submitted.

[0443] Biological cohorts:

[0444] - Subjects with PD-L1 negative NSCLC and HNSCC, TNBC, esophageal SCC, melanoma, gastric cancer or ovarian cancer who meet the above dose amplification criteria.

[0445] - There is no specific number of subjects for each tumor type.

[0446] - Subjects must have palpable tumors for multiple (up to 3) fresh biopsies.

[0447] - If feasible, a fresh baseline biopsy is required; if feasible, an in-treatment biopsy from the same lesion as the pre-drug biopsy is required (cycle 1 only); if feasible, a fresh biopsy from any lesion demonstrating deterioration is required.

[0448] iii. Combinations (Part D):

[0449] Security Import:

[0450] ○ Subjects must have histologically or cytologically confirmed HNSCC disease.

[0451] ○ The primary tumor must originate from the oral cavity, oropharynx, nasopharynx, hypopharynx, or larynx.

[0452] ○ Subjects must not have received anti-PD-1 or PD-L1 therapy or other immunotherapy agents.

[0453] ○ Subjects in metastatic cases must not have received a previous line of cytotoxic therapy. If a subject's disease worsens or relapses 6 or more months after the last dose, cytotoxic therapy should only be used in an adjuvant / neoadjuvant setting.

[0454] ○ Subjects must have PD-L1 expression ≥1 based on historical test results according to CPS or TPS.

[0455] ○ Tumor tissue is required at the time of recruitment. The most recently archived tumor tissue collected within 24 months of recruitment should be provided. If no archived tissue is available, a newly obtained biopsy must be submitted.

[0456] 2. Must be at least 18 years old and legally an adult at the time of consent, and be at least the legal age of majority in their local jurisdiction.

[0457] 3. The Eastern Coast Oncology Cooperative (ECOG) daily performance status score is 0 or 1.

[0458] 4. Disease that is measurable at baseline according to RECIST v1.1.

[0459] 5. The following baseline laboratory data are:

[0460] Absolute neutrophil count (ANC) ≥1000 / µL

[0461] Heme (Hgb) ≥ 9 g / dL

[0462] Platelet count ≥100,000 / μL

[0463] Serum bilirubin ≤1.5 × upper limit of normal (ULN) or ≤3 × ULN for patients with Gilbert's disease.

[0464] The estimated glomerular filtration rate (eGFR) using the Modification of Diet in Renal Disease (MDRD) study formula is ≥45 mL / min / 1.73 m 2 Multiply by body surface area (BSA) (if applicable).

[0465] Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) ≤3 × ULN (≤5 × ULN if there is evidence of malignant disease involving the liver)

[0466] 6. For participants who are of fertility, please meet the following conditions:

[0467] i. A serum or urine pregnancy test (minimum sensitivity 25 mIU / mL or equivalent units of β-human chorionic gonadotropin [β-hCG]) must be negative within 7 days prior to the first dose of SGN-PDL1V. Subjects with false positive results and documented confirmation that they are not pregnant are eligible to participate.

[0468] ii. You must agree not to attempt pregnancy during the study period and for at least 2 months after the final dose of SGN-PDL1V and 4 months after the final dose of pembrolizumab (Part D only).

[0469] iii. You must agree that you will not breastfeed or donate eggs from the time of informed consent until 2 months after the final dose of SGN-PDL1V and 4 months after the final dose of pembrolizumab (Part D only).

[0470] iv. If there is a possibility of pregnancy from sexual activity, at least two acceptable methods of contraception must be used from the time of informed consent, throughout the study and for at least two months after the final administration of SGN-PDL1V and for four months after the final administration of pembrolizumab (Part D only), at least one of which must be highly effective.

[0471] 7. For subjects who could impregnate others, please adhere to the following conditions:

[0472] i. Must agree to refrain from donating sperm from the start of the informed consent process and throughout the entire study period, and for at least four months after the final administration of the investigational drug.

[0473] ii. If there is a possibility of pregnancy from sexual activity with a person of childbearing potential, at least two acceptable methods of contraception must be used from the time of informed consent and throughout the study and for at least four months after the final administration of the study drug, at least one of which must be effective.

[0474] iii. If having sexual intercourse with a pregnant or breastfeeding person, condoms must be used from the time informed consent is obtained and throughout the study and for at least 4 months after the final administration of the study drug.

[0475] Exclusion criteria

[0476] 1. A history of another malignancy within 3 years prior to the first dose of SGN-PDL1V, or any evidence of residual disease from a previously diagnosed malignancy. Exceptions are malignancies with negligible risk of metastasis or death (e.g., 5-year overall survival [OS] ≥90%), such as adequately treated cervical carcinoma in situ, non-melanoma skin cancer, localized prostate cancer, ductal carcinoma in situ, or stage I uterine cancer.

[0477] 2. Known active central nervous system metastases. Subjects with previously treated brain metastases may participate, subject to the following conditions: they must be clinically stable for at least 4 weeks after treatment for brain metastases and prior to enrollment in the study; they must not have new or expanded brain metastases; and they must have discontinued prescribed corticosteroids for symptoms associated with brain metastases for at least 7 days prior to the first dose of SGN-PDL1V.

[0478] 3. Leptomeningeal disease.

[0479] 4. Less than 5 half-lives since prior treatment with an anti-PD-L1 agent (as directed by the protocol) (see Table 10).

[0480] Table 10: Clearance period of previous anti-PDL1 therapy

[0481]

[0482] Note: If the subject has been exposed to an anti-PD-L1 agent not listed above, please contact your healthcare professional for clearance information.

[0483] 5. Previously received medication containing MMAE.

[0484] 6. Existing neuropathy grade ≥2 according to NCI CTCAE v5.0.

[0485] 7. Any grade (according to NCI CTCAE, version 5.0) active viral, bacterial, or fungal infection within 2 weeks prior to the first dose of SGN-PDL1V, unless the investigator considers it clinically insignificant (e.g., onychomycosis). Routine antimicrobial prophylaxis is permitted.

[0486] 8. Known or suspected autoimmune disease or significant autoimmune-related toxicity from previous immuno-oncology-based therapies, and currently experiencing an attack, not yet in complete remission, or at risk of relapse.

[0487] 9. Uncontrolled diabetes mellitus is defined as Hgb A1c ≥8% or Hgb A1c between 7% and <8%, accompanied by unspecified related symptoms of diabetes (polyuria or polydipsia).

[0488] 10. Positive for hepatitis B surface antigen expression. Known active hepatitis C infection (positive for polymerase chain reaction [PCR] or received antiviral therapy for hepatitis C within the last 6 months). Subjects who have received treatment for hepatitis C infection and have a recorded 12-week sustained virologic response are eligible to participate.

[0489] 11. Known to be positive for human immunodeficiency virus (HIV).

[0490] 12. A documented medical history of cerebrovascular events (stroke or transient ischemic attack), unstable angina, myocardial infarction, congestive heart failure, or cardiac symptoms meeting New York Heart Association Class III or IV within 6 months prior to their first dose of SGN-PDL1V.

[0491] 13. History of non-infectious ILD or pneumonia requiring steroids, current ILD or pneumonia, grade ≥3 lung disease unrelated to underlying malignancy, prior lung radiotherapy of >30 Gy within 6 months prior to the first dose of study treatment, and known carbon monoxide lung diffusion capacity [DLCO]; heme correction <50% of predicted value. Subjects with chronic obstructive pulmonary disease (COPD) are eligible if they do not require supplemental oxygen or systemic corticosteroids (prednisolone or equivalent) >10 mg daily.

[0492] 14. Receive treatment with a potent inducer or inhibitor of cytochrome P450 3A (CYP3A) within 14 days prior to the first dose of SGN-PDL1V.

[0493] 15. Four weeks prior to the first dose of SGN-PDL1V, or two weeks prior to the first dose of SGN-PDL1V if the underlying disease has worsened at the time of treatment, there has been incomplete chemotherapy, immunotherapy, biologics, and / or other approved or investigational antitumor therapy (as directed by the protocol).

[0494] 16. Two weeks prior to the first dose of SGN-PDL1V, there was no completed focal radiotherapy or major surgery.

[0495] 17. Subjects who are breastfeeding, pregnant, or planning to become pregnant from the time of informed consent until 2 months after the final dose of SGN-PDL1V and 4 months after the final dose of pembrolizumab (Part D only).

[0496] 18. Known hypersensitivity to any excipients contained in the formulation of SGN-PDL1V or pembrolizumab.

[0497] 19. According to researchers' insights, the estimated life expectancy is <12 weeks.

[0498] 20. Subjects who received the live vaccine within 30 days of their first dose of SGN-PDL1V.

[0499] 21. According to the researchers’ insights, other serious underlying medical conditions may impair a subject’s ability to receive or tolerate the planned treatment and follow-up.

[0500] 22. Any toxicity associated with previous treatment that has not returned to baseline or is grade >1, except for hair loss.

[0501] Parts C and D only:

[0502] 23. Diagnosed with immunodeficiency or received chronic systemic steroid therapy (at a dose of more than 10 mg of prednisolone equivalent daily) or any other form of immunosuppressive therapy within 7 days prior to the first dose of the study drug.

[0503] 24. Having an active autoimmune disease requiring systemic treatment (i.e., the use of disease modulators, corticosteroids, or immunosuppressive drugs) within the past 2 years. Replacement therapies (such as thyroxine, insulin, or physiological corticosteroid replacement therapy for adrenal or pituitary insufficiency) are not considered a form of systemic treatment and are permitted.

[0504] Treatment of subjects

[0505] Treatment applied

[0506] All subjects in this study will receive the study agent SGN-PDL1V. Subjects in Part D will also receive pembrolizumab. Administration of the combination therapy must begin with an IV dose of pembrolizumab, followed by SGN-PDL1V. SGN-PDL1V should not be initiated until at least 30 minutes have elapsed after the completion of pembrolizumab administration. All doses will be administered via IV infusion.

[0507] Dosage and administration

[0508] SGN-PDL1V will initially be administered via IV infusion in a 2-week, 3-week cycle. Alternative dosing frequencies, such as 3-week, 2-week, or 3-week, may be evaluated in Part B, as recommended by the SMC. SGN-PDL1V should not be mixed with other medications.

[0509] Dosing will be based on adjusted ideal body weight (AIBW). AIBW reduces pharmacokinetic variability of SGN-PDL1V across weight groups by increasing exposure in low-weight subjects and decreasing exposure in high-weight subjects, thereby mitigating the risk of potential underdosing or overdosing across all weight groups. AIBW provides adjustment to ideal body weight (IBW) when a subject's actual total body weight (TBW) differs from their IBW. Because AIBW is calculated using the subject's sex, height, and TBW, the percentage adjustment to the total dose will be based on the body mass index (BMI) group.

[0510] During the dose escalation phase in Part A, subjects should remain in the clinic for at least 4 hours after completing SGN-PDL1V administration on Day 1 of cycles 1, 2, and 3. Additional monitoring in subsequent cycles will be considered after reviewing safety data with the SMC. During the dose and schedule optimization phase in Part B, an observation period is required only on Day 1 of cycle 1. If the SMC determines that safety is adequate after reviewing the data from the dose escalation cohort, the observation period may be shortened to 2 hours.

[0511] The duration of infusion will vary depending on the method and dosage of administration. Please refer to the Pharmacy Binder for more details. The initial method of SGN-PDL1V administration will be stepwise infusion. In stepwise infusion, the infusion rate increases at set time intervals until the prescribed maximum infusion rate is reached. The first infusion of SGN-PDL1V will begin at a rate of 50 mg / hour. If well tolerated for the first 30 minutes, the rate will be gradually increased every 30 minutes (no more than twice the rate) until the maximum rate (800 mg / hour) is reached. In subsequent infusions, the infusion rate can be increased more rapidly at shorter time intervals: for example, in the first 15 minutes, if tolerated, the rate can be gradually increased every 15 minutes (no more than twice the rate) until the maximum rate is reached.

[0512] As clinical experience with stepwise infusions develops, the maximum infusion rate may be increased or decreased based on accumulated safety data and / or the recommendations of the SMC; the SMC may also recommend a longer or shorter total infusion duration. Alternative methods of SGN-PDL1V administration may also be evaluated. This may include systematically implementing strategies such as changing the planned infusion duration, administering at a fixed infusion rate, administering in divided doses, administering at the weight-capped dose, administering a flat dose, or changing pre-administration. See Pharmacy Binder for more details. Any material changes to SGN-PDL1V dosing or administration may only be implemented after obtaining regulatory approval where appropriate, in accordance with local regulations.

[0513] If an individual subject is intolerant to the infusion, the duration of the infusion for that subject may be increased; the duration of subsequent infusions may also be increased at the discretion of the researchers after consulting with medical supervisors. Conversely, if a subject tolerates continuous infusion without an IRR > 1, the duration of the infusion may be shortened at the discretion of the researchers after consulting with medical supervisors (i.e., administered at a faster rate), and this can be dose-cohort specific.

[0514] During and at any time after administration, the infusion site should be closely monitored for redness, swelling, pain, and infection. Subjects should be advised to report any redness or discomfort immediately during or after administration.

[0515] Upper limit of weight dose administration

[0516] As pharmacodynamic, clinical activity, and pharmacodynamic data continue to develop, upper weight limit (IV) dosing can be implemented following SMC recommendations. Compared to weight-based dosing (where the total dose is calculated without an upper limit on the subject's weight), upper weight limit dosing restricts the weight used for total dose calculations. The upper limit for dose calculations should be determined based on newly emerging data.

[0517] For example, if the weight limit is 100 kg, subjects weighing ≥100 kg will use 100 kg to calculate the total dose to be administered. For subjects weighing <100 kg, the subject's actual weight will be used to calculate the total dose to be administered.

[0518] Weight-based dosage

[0519] Weight-based dosing is based on the subject's actual weight or according to institutional standards. For subjects experiencing a weight change of ≥10% from baseline, the dose must be adjusted. Subject weight must be measured as described in the event schedule during all relevant assessment windows. Additional dose adjustments for weight changes are permitted according to institutional standards. Rounding to the nearest whole number of milligrams is permitted up to 5% of the nominal dose.

[0520] Dosage modification

[0521] For SGN-PDL1V, dose reduction or dosing interval extension for toxicities (including DLT) may be considered on a case-by-case basis after consultation with a medical supervisor. Dosage modifications for toxicities (including DLT) may be permitted on a case-by-case basis after consultation with a medical supervisor (Table 11). For subjects treated at the lowest dose level, the dose may be reduced to 25% of the most recently administered dose, the dosing frequency may be reduced (e.g., if the initial dosing was 2Q3W, dosing may be changed to Q3W), or treatment may be discontinued. Subjects experiencing DLT in cycle 1 should not receive further treatment with SGN-PDL1V unless toxicities are adequately managed and after consultation with a medical supervisor. The type and severity of observed AEs will inform decision-making. For subjects treated at the lowest dose level, the dose may be reduced to 25% of the most recently administered dose, the dosing frequency may be reduced, or treatment may be discontinued.

[0522] If a subject experiences a clinically significant, unresolved treatment-induced adverse event (TEAE) on or after the next scheduled dosing day of cycle 1, the next dose may be delayed for up to 7 days. Delays for other reasons or lasting >7 days must be discussed with a healthcare professional. Subjects requiring a dose delay >7 days due to an unresolved TEAE may be administered at a reduced dose or with a reduced dosing frequency in subsequent cycles after discussion with a healthcare professional.

[0523] During the DLT period (cycle 1), growth factor and fluid support are discouraged unless medically indicated; subjects receiving growth factors (e.g., G-CSF or GM-CSF) or fluid support for non-DLT causes during this period may be deemed unevaluable for DLT in Parts A and D. Growth factor support should be considered for the prevention or treatment of cytopenia in subsequent cycles. For subjects who still experience relapsed grade 4 neutropenia despite growth factor use, discontinuation, reduction of dosing frequency, or dose reduction to one dose level lower than the current dose may be considered. Table 11 describes recommended dose modifications for study-related toxicities. Consultation with a medical supervisor is required after the occurrence of ≥ grade 3 AEs that may be associated with SGN-PDL1V treatment to determine whether to continue study treatment.

[0524] Table 11: Recommended dosage modifications for SGN-PDL1V-related toxicities.

[0525]

[0526]

[0527]

[0528] For subjects at or below dose level 2, treatment should be resumed at 50% of the most recently administered dose. If a subject is dosed twice for the same adverse event (excluding neutropenia) and the same adverse event recurs, treatment must be permanently discontinued.

[0529] AE = Adverse Event; ILD = Interstitial Lung Disease; IV = Intravenous; IRR = Infusion-Related Reaction

[0530] a. After an ≥ grade 3 AE that may be associated with SGN-PDL1V, treatment may not continue until after consultation with a medical supervisor.

[0531] b. Even with a reduced dose, if grade 3 (or grade 4 neutropenia) toxicity occurs again, the frequency of administration may be reduced after consulting with a medical supervisor.

[0532] c. If a subject experiences a grade 4 IRR, an allergic reaction, or an anaphylaxis, SGN-PDL1V must be permanently discontinued.

[0533] d For all grades of febrile neutropenia, please follow the dosage modification guidelines for grade 4 hematologic toxicity.

[0534] Intrasubject dose escalation is permitted only if the subject meets the procedural criteria for receiving the next SGN-PDL1V cycle, has toxicity ≤ grade 2 observed during the previous treatment cycle, and, in the investigator's opinion, the subject is likely to derive clinical benefit from dose escalation. The maximum intrasubject dose escalation level should be less than or equal to one level of the current actively enrolling level. Additional treatment cycles may be administered at the investigator's discretion, in consultation with medical supervisors, at doses and schedules previously deemed safe by the SMC.

[0535] Pembrolizumab

[0536] Instructions for use of pembrolizumab

[0537] To assess the safety of this combination, an initial six subjects will be administered SGN-PDL1V plus pembrolizumab. Safety will be assessed by the incidence of disease-related thrombosis (DLT). Pembrolizumab is a humanized mAb that blocks the interaction between PD-1 and its ligands PD-L1 and PD-L2. Pembrolizumab is an IgG4κIg with a molecular weight of approximately 149 kDa.

[0538] Pembrolizumab will be supplied in single-use vials as a 100 mg / 4 mL (25 mg / mL) solution. Pembrolizumab for injection is a sterile, preservative-free, clear to slightly milky white, colorless to slightly yellow solution that requires dilution for IV infusion. Each vial contains 100 mg of pembrolizumab in a 4 mL solution. Each 1 mL solution contains 25 mg of pembrolizumab reconstituted with L-histidine, polysorbate, sucrose, and WFI USP.

[0539] Dosage and administration

[0540] In Part D, the investigational treatment with pembrolizumab will be administered via 30-minute IV infusion at a dose of 200 mg every 3 weeks. The site should make every effort to ensure the target infusion time is as close to 30 minutes as possible. However, given the variability of infusion pumps between sites, a window between -5 minutes and +10 minutes (i.e., an infusion time of 30 minutes - 5 minutes / +10 minutes) is permitted. Administration of the combination therapy must begin with the IV administration of pembrolizumab, followed by SGN-PDL1V. SGN-PDL1V should not be initiated until at least 30 minutes have elapsed after the completion of pembrolizumab administration. Unless otherwise instructed, the investigational treatment should be administered according to the product information or institutional guidelines.

[0541] Dosage modification

[0542] Please refer to the product information on pembrolizumab dosage modifications and the pembrolizumab toxicity management guidelines. For the combination cohort, AEs may be attributable to a single study treatment or a combination of study treatments. The final decision regarding causation is made at the investigator's discretion. If the event is clearly related to one of the drugs, follow the specific instructions for that drug. If the event is related to more than one drug, follow the instructions for all drugs related to the event. There may be situations where a subject may not tolerate combination therapy but could benefit from treatment with a single drug. These situations must be discussed and approved by the trial commissioner before the subject can continue treatment. In cases where it may be difficult to attribute to an individual study treatment, the management of pausing or discontinuing study treatment should apply to all study treatments administered.

[0543] Management of treatment-induced adverse events

[0544] The IB for SGN-PDL1V and the product information for pembrolizumab describe common adverse events (AEs) and less common serious findings observed in individual study treatments, respectively. Subjects receiving pembrolizumab should monitor for AEs according to the guidelines in the product information.

[0545] SGN-PDL1V

[0546] Management of infusion-related reactions

[0547] IRR is characterized by an adverse reaction to the infusion of a pharmacological or biological substance. IRR occurs within 24 hours of infusion and can present as a combination of various signs or symptoms, including fever, rigidity, flushing, itching, various types of rash, urticaria, dyspnea, nausea, vomiting, back or abdominal pain, and / or hypotension.

[0548] IRR may occur during the infusion of study treatment. Infusion should be administered in a facility with appropriate equipment and personnel to manage any potential anaphylactic reactions. All supportive measures consistent with recommended subject care should be provided throughout the study, according to institutional standards. Supportive measures may include extending the infusion time and / or administering IRR medication. If a Grade 3 IRR occurs in >20% of subjects, subsequent subjects will require pre-administration or modification. Subjects are advised to remain near the healthcare facility for 24 hours after SGN-PDL1V administration to allow for real-time assessment in the event of a delayed IRR.

[0549] During dose escalation, additional mitigation strategies may be explored to manage IRR as advised or required by the SMC. These mitigation strategies may include, but are not limited to, any or all of the following:

[0550] Slowing down, interrupting, or otherwise adjusting the application of SGN-PDL1V

[0551] Possible pre- or post-infusion medications, for example:

[0552] ○ Antihistamines, such as diphenhydramine 50 mg IV or equivalent and famotidine 40 mg IV or equivalent.

[0553] ○ Antipyretics, such as oral (PO) acetaminophen or paracetamol 500 to 1,000 mg or equivalent.

[0554] ○ Antiemetics, such as ondansetron

[0555] IV fluid support, such as normal saline.

[0556] ○ Anti-rigor drugs, such as meperidine

[0557] ○ Vasopressors

[0558] ○ Corticosteroids, such as hydrocortisone 100 mg IV or equivalent, or methylprednisolone 40 mg IV or equivalent.

[0559] Recommendations for IRR management are detailed in Table 12.

[0560] Table 12: Management of SGN-PDL1V-related infusion-related reactions

[0561]

[0562] IRR = Infusion-related reactions; IV = Intravenous; NSAID = Nonsteroidal anti-inflammatory drug

[0563] All Grade 3 or higher IRR events (occurring during or within 24 hours after infusion) or allergic reactions (occurring >24 hours after infusion), regardless of their relevance to SGN-PDL1V, should be reported immediately to the medical supervisor. Grade 4 IRR events should result in immediate and permanent discontinuation of SGN-PDL1V for the subject. Serious adverse events (SAEs) should be reported within the 24-hour SAE reporting timeframe using the standard SAE form.

[0564] Adverse events

[0565] definition

[0566] Adverse events

[0567] According to the ICH E2A guidelines for Expedited Reporting and 21 CFR 312.32, an adverse event (AE) in an Investigational New Drug (IND) safety report is any unfortunate medical event that occurs in a patient or clinical trial subject who is receiving the drug and that is not necessarily causally related to the treatment.

[0568] When determining whether to record test results, medical conditions, or other events on an adverse event CRF, the following information should be considered:

[0569] From the time of informed consent until the day before Day 1 of the study, only protocol-related adverse events (AEs) need to be recorded. Protocol-related AEs are defined as unfortunate medical events that occur as a result of procedures prescribed in the protocol.

[0570] All medical conditions that were present or persisted prior to administration on day 1 of the study and whose NCI CTCAE grade increased should be documented.

[0571] Medical conditions that were present or persisted before administration on day 1 of the study and whose severity worsened, frequency increased, became associated with SGN-PDL1V, or otherwise worsened but did not meet the threshold for an increase in the NCI CTCAE grade should be documented.

[0572] All adverse events (AEs) from Day 1 of the study (before dosing) to the end of the safety reporting period (regardless of whether they are related to SGN-PDL1V) should be documented. Any complications arising from any procedure (e.g., biopsy), whether or not the procedure is prescribed, should be documented as AEs.

[0573] Generally, abnormal laboratory values ​​should not be recorded as adverse events (AEs) unless they are related to clinical signs or symptoms, require intervention, lead to a surgical adverse event (SAE), or result in study termination or interruption / cessation of study treatment. When an AE is recorded due to a laboratory abnormality, the resulting medical condition should be recorded, not just the abnormality itself (e.g., "anemia" should be recorded, not "hypoglobulin").

[0574]

[0575] Serious adverse events

[0576] An AE should be classified as an SAE if it meets one of the following criteria:

[0577]

[0578] Severity of adverse events

[0579] The severity of adverse events (AEs) should be assessed using the NCI CTCAE version 5.0 rating. AE severity and severity level are assessed independently. "Severity level" characterizes the intensity of the AE. "Severity" is a regulatory definition and serves as guidance for trial commissioners in defining their regulatory reporting obligations.

[0580] Relationship between adverse events and research treatment

[0581] The relationship between each adverse event (AE) and one or more agents (SGN-PDL1V and / or pembrolizumab) in the study treatment should be assessed by the investigator using the following guidelines:

[0582]

[0583] Data analysis methods

[0584] Determination of sample size

[0585] Approximately 160 participants can be recruited in Parts A and B of this study. This number is based on the assumption that the dose escalation cohort in Part A is expected to evaluate approximately 80 participants, while the dose and schedule optimization (Part B) is expected to evaluate approximately 80 participants (if needed). To evaluate the combination of SGN-PDL1V and pembrolizumab, approximately 12 participants will be recruited in the safety induction cohort (Part D).

[0586] Dose escalation and MTD identification will be guided by the DE analysis set in the mTPI design. Due to the dynamic nature of the mTPI design, the exact sample size for this design cannot be predetermined in Part A. Approximately 80 subjects are expected to be evaluated in Part A.

[0587] In Part C, the sample size for the specific disease amplification cohort was determined to be 40 bits to ensure reasonable precision in the estimation of the objective response rate (ORR). The following lists the precise two-sided confidence intervals (CIs) given for the observed ORR.

[0588]

[0589] Definition of research endpoint

[0590] Objective response rate

[0591] ORR (excluding confirmatory response) is defined as the proportion of subjects who achieve partial response (PR) or complete response (CR) as assessed by researchers according to RECIST v1.1. Confirmatory ORR is defined as the proportion of subjects who subsequently achieve confirmed CR or PR according to RECIST v1.1. For ORR calculation, subjects whose disease response cannot be assessed according to response criteria will be rated as "non-evaluable." In ORR calculation, subjects without post-baseline response assessment or with a "non-evaluable" response will be counted as non-responders.

[0592] No deterioration survival period

[0593] Progression-free survival (PFS) is defined as the time from the start of study treatment to the first recorded disease progression (PD) (assessed by the investigator according to RECIST v1.1) or death from any cause, whichever comes first. PFS data will be reviewed on the last effective disease assessment date for subjects without PD or the last effective disease assessment date for subjects without PD prior to recording the start of non-routine antitumor therapy. Subjects lacking tumor response assessment after their first dose of study treatment will have their event time reviewed on Day 1. Details of the review rules will be provided in SAP.

[0594] Overall lifespan

[0595] OS is defined as the time from the start of study treatment to death from any cause. For subjects whose death is unknown, OS will be reviewed at the last known date of survival.

[0596] Duration of objective response

[0597] Duration of Response (DOR) is defined as the time from the first recorded objective tumor response (subsequently confirmed CR or PR) to the first recorded disease progression (PD) (as assessed by the investigator according to RECIST v1.1) or to death from any cause, whichever comes first. DOR data will be reviewed on the date of the last adequate disease assessment for subjects who did not have PD at the time of analysis, or on the date of the last effective disease assessment for subjects who did not have PD before the initiation of subsequent anticancer therapy. Details of the review rules will be provided in SAP. DOR will be calculated only for the subgroup of subjects who achieved CR or PR.

[0598] Subgroup inspection

[0599] All analyses will be presented based on dose level / cohort and totals for selected endpoints (if applicable).

[0600] As an exploratory analysis, subgroup analysis can be performed targeting a selected endpoint. Subgroups may include, but are not limited to, the following:

[0601] Previous treatment

[0602] Disease subtypes

[0603] PD-L1 expression level

[0604] Example 2: Modeling of Tumor Cell Killing Mechanism

[0605] Bystander activity

[0606] PDL1V exhibits potent cytotoxicity against various PD-L1-positive cancer cell lines in vitro. Flow cytometry-based viability assays were performed to assess whether PDL1V also drives dose-dependent bystander effect-based cytotoxicity in PD-L1-positive tumor cells when co-cultured with PD-L1-positive tumor cells.

[0607] Methods: SUDHL4 cells engineered to express PD-L1 and wild-type PD-L1-negative cells were thawed from frozen vials stored at -210°C and transferred to complete growth medium (RPMI 1640 cell culture medium + 10% FBS). Cells were counted and seeded in duplicate (40K cells per well for PD-L1 positive and / or 10K cells per well for PD-L1 negative) into 12-well plates. PDL1V, homovitin ADC, and free MMAE (positive control) were prepared in complete medium and added to appropriate wells at the following concentrations (untreated complete medium was added to wells corresponding to untreated conditions): PDL1V: 50 ng / ml, 5 ng / ml, or 0.5 ng / ml; homovitin: 50 ng / ml, or MMAE: 1.5 nM. The plates were incubated at 37°C and 5% CO2 for 96 hours. Cells were collected, and anti-PD-L1 antibody and fluorescent dye were added to all wells of a 96-well assay plate. Cells were incubated on ice for 30 minutes. Unbound antibody was removed, and cells were incubated with counting beads at room temperature for 15 minutes. Fluorescence was measured using an Attune flow cytometer.

[0608] like Figure 10 As shown, when co-cultured with PD-L1-positive tumor cells, PDL1V induces dose-dependent, bystander-effect-based cytotoxicity in PD-L1-negative tumor cells. For example, at the endpoint, at 5 ng / ml PDL1V, ~100% of PD-L1-negative cells were viable in monoculture, compared to only ~50% of PD-L1-negative cells and ~10% of PD-L1-positive cells in co-culture (as compared to the untreated group). At the endpoint, at 50 ng / ml PDL1V, ~100% of PD-L1-negative cells were viable in monoculture, compared to only ~10% of PD-L1-negative cells and 0% of PD-L1-positive cells in co-culture (as compared to the untreated group).

[0609] Immunogenic cell death

[0610] Immunogenic cell death (ICD) (regulated activation that recruits immune cells to the tumor microenvironment) is induced by damage-associated molecular patterns (DAMPs) released by tumor cells, such as cell surface chaperone calreticulin (CRT), extracellular adenosine triphosphate (eATP), and high-mobility group box 1 (HMGB1). As shown in this example, PD-L1 positive cancer cells treated with PDL1V in vitro released all three DAMPs, indicating that targeted delivery of the MMAE payload drove the induction of ICD.

[0611] Method (eATP Release): Karpas 299 cells were thawed from vials stored at -21°C into complete growth medium (RPMI 1640 cell culture medium + 10% FBS). Cells were counted and seeded in triplicate at 20K cells per well into 96-well plates. PDL1V, isotype vitolamine ADC, unconjugated PD-L1 antibody, and free MMAE (positive control) were prepared in complete medium and added to the appropriate wells (untreated complete medium was added to the wells corresponding to the untreated condition). The RealTime Glo eATP reagent matrix was added to each well of the assay plate according to the manufacturer's instructions. The assay plate was mixed on an orbital oscillator at 500 rpm for 30 seconds to produce a homogenized mixture, and the luminescence of each well was measured using Cytation (Agilent / Bio Tek).

[0612] like Figure 11A As shown, PD-L1 positive cancer cells treated with PDL1V in vitro released much higher levels of eATP than untreated and negative controls (isomorphic vitoline, PD-L1 mAb), and the levels were similar to those of the positive control (MMAE).

[0613] Method (HMGB1 Release): Karpas 299 cells were thawed from vials stored at -21°C into complete growth medium (RPMI 1640 cell culture medium + 10% FBS). Cells were counted and seeded in duplicate (200k cells per well) into 12-well plates. PDL1V, isotype vitolamine ADC, unconjugated PD-L1 antibody, and free MMAE (positive control) were prepared in complete medium and added to appropriate wells (untreated complete medium was added to wells corresponding to untreated conditions). The assay plate was incubated at 37°C and 5% CO2 for 42 hours. The supernatant from each well was transferred to appropriate wells in a new plate, and the cell supernatant was incubated with a mixture of cell supernatant and anti-human HMGB1 antibody at room temperature for 90 minutes. After adding luminescence detection buffer to each well and incubating at room temperature for 4 minutes, the luminescence of each well was measured using an EnVision microplate reader (PerkinElmer).

[0614] like Figure 11B As shown, PD-L1 positive cancer cells treated with PDL1V in vitro released much higher levels of HMGB1 than untreated and negative controls (isovitoline, PD-L1 mAb), and the levels were similar to those of the positive control (MMAE).

[0615] Methods (Calcoretin Release): Karpas 299 cells were thawed from frozen vials stored at -21°C and transferred to complete growth medium (RPMI 1640 cell culture medium + 10% FBS). Cells were counted and seeded at 1.5E6 cells per well into 6-well plates. PDL1V, isotype vidolin ADC, unconjugated PD-L1 antibody, and free MMAE (positive control) were prepared in complete medium and added to appropriate wells (untreated complete medium was added to wells corresponding to untreated conditions). The assay plates were incubated at 37°C and 5% CO2 for 42 hours. Cells were harvested, counted, and seeded at 250K cells per well into 96-well U-shaped plates. Anti-human calretin or anti-human isotype antibody was added to appropriate wells of the assay plates and incubated on ice for 30 minutes. Unbound antibody was removed, and cells were incubated with a fluorescently active stain at room temperature for 15 minutes. Fluorescence was measured using an Attune flow cytometer.

[0616] like Figure 11C As shown, PD-L1 positive cancer cells treated with PDL1V in vitro exhibited significantly higher levels of cell surface calreticulin compared to untreated and negative controls (isomorphic vidotine, PD-L1 mAb), and the levels were similar to those of the positive control (MMAE).

[0617] Effects of PDL1V on immune cell viability

[0618] As an immune checkpoint protein, PD-L1 is expressed on antigen-presenting cells and T cells. Depletion of these cells can affect the tolerability of PDL1V as a monotherapy or limit its efficacy when combined with anti-PD-1 therapy. To investigate the potential for immune cell depletion, CD4+ and CD8+ T cells were evaluated in an in vitro fluorescence-based cytotoxicity assay. As shown in this example, PDL1V exhibited minimal cytotoxic activity across all three cell types (despite expressing PD-L1). These data support the view that PDL1V possesses selective cytotoxicity against PD-L1-positive and PD-L1-negative cancer cells in the tumor microenvironment, but lacks cytotoxicity against PD-L1-positive immune cells involved in innate antitumor immunity.

[0619] Methods: CD3+ pan-T cells were thawed from frozen vials stored at -21°C and transferred to complete growth medium (RPMI 1640 cell culture medium + 10% FBS). Pan-T cells were counted, labeled with amine-reactive CFSE dye, and seeded in duplicate (donor 2) or triplicate (donor 1) into 96-well U-bottom microplates. CD3 / CD28 cross-linked beads and 10-dot, 3-fold serially diluted PDL1V, isotype vitolamine ADC, and positive control drugs were prepared in complete medium and added to appropriate wells. The assay plates were incubated at 37°C and 5% CO2 for 96 hours.

[0620] Prepare fluorescent active dyes and antibodies binding to CD3, CD4, CD8, and PD-L1 according to the manufacturer's instructions, and add them to each well of the assay plate. After incubating on ice for 30 minutes, wash the cells with 1X PBS and measure fluorescence using an Attune flow cytometer.

[0621] like Figure 12A and Figure 12B As shown, despite the expression of PD-L1 in T cells, PDL1V exhibits minimal cytotoxic activity in CD4+ T cells for donors 1 and 2.

[0622] like Figure 12C and Figure 12D As shown, despite the expression of PD-L1 in T cells, PDL1V exhibits minimal cytotoxic activity in CD8+ T cells for donors 1 and 2.

[0623] Informal sequence list

[0624] SEQ ID NO:1 – SG-559-01 LALA hIgG1 heavy chain protein

[0625] QVQLVQSGAEVKKPGSSVKVSCKTSGDTFSTAAISWVRQAPGQGLEWMGGIIPIFGKAHYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYFCARKFHFVSGSPFGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0626] SEQ ID NO: 2 – SG-559-01 Kappa Light Chain - Protein

[0627] EIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0628] SEQ ID NO: 3 – SG-559-01 Heavy Chain Variable Region - Protein

[0629] QVQLVQSGAEVKKPGSSVKVSCKTSGDTFSTAAISWVRQAPGQGLEWMGGIIPIFGKAHYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYFCARKFHFVSGSPFGMDVWGQGTTVTVSS

[0630] SEQ ID NO:4 – SG-559-01 Light chain variable region - protein

[0631] EIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPTFGQGTKVEIK

[0632] SEQ ID NO:5 – SG-559-01 Heavy chain CDR1-protein

[0633] TAAIS

[0634] SEQ ID NO:6 – SG-559-01 Heavy chain CDR2-protein

[0635] GIIPIFGKAHYAQKFQG

[0636] SEQ ID NO:7 – SG-559-01 Heavy chain CDR3-protein

[0637] KFHFVSGSPFGMDV

[0638] SEQ ID NO:8 – SG-559-01 Light chain CDR1-protein

[0639] RASQSVSSYLA

[0640] SEQ ID NO:9 – SG-559-01 Light chain CDR2-protein

[0641] DASNRAT

[0642] SEQ ID NO:10 – SG-559-01 Light chain CDR3-protein

[0643] QQRSNWPT

[0644] SEQ ID NO:11 – SG-559-01 hIgG1 heavy chain protein

[0645] QVQLVQSGAEVKKPGSSVKVSCKTSGDTFSTAAISWVRQAPGQGLEWMGGIIPIFGKAHYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYFCARKFHFVSGSPFGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0646] SEQ ID NO: 12 – SG-559-01 variable heavy chain region - nucleic acid

[0647] caggtccagctggtgcagtctggggctgaggtgaagaagcctgggtcctcggtgaaggtctcctgcaagacttctggagacaccttcagcaccgccgctatcagctgggtgcgacaggcccctggacaagggcttgagtggatgggagggatcatccctatatttggtaaagcacactacgcacagaagttccagggcagagtcacgattaccgcggacgaatccacgagcacagcctacatggagctgagcagcctgagatctgaggacacggccgtgtatttttgtgcgagaaagtttcactttgtttcggggagccccttcggtatggacgtctggggccaagggaccacggtcaccgtctcctca

[0648] SEQ ID NO: 13 – SG-559-01 variable light chain region - nucleic acid

[0649] gaaattgtgttgacacagtctccagccaccctgtctttgtctccaggggaaagagccaccctctcctgcagggccagtcagagtgttagcagctacttagcctggtaccaacagaaacctggccaggctcccaggctcctcatctatgatgcatccaacagggccactggcatcccagccaggttcagtggcagtgggtctgggacagacttcactctcaccatcagcagcctagagcctgaagattttgcagtttattactgtcagcagcgtagcaactggccgacgttcggccaagggaccaaggtggaaatcaaa

[0650] SEQ ID NO: 14 - SG-559-01 LALA hIgG1 Heavy Chain - Nucleic Acid

[0651]

[0652] SEQ ID NO: 15 – SG-559-01 Kappa light chain – nucleic acid

[0653] gaaattgtgttgacacagtctccagccaccctgtctttgtctccaggggaaagagccaccctctcctgcagggccagtcagagtgttagcagctacttagcctggtaccaacagaaacctggccaggctcccaggctcctcatctatgatgcatccaacagggccactggcatcccagccaggttcagtggcagtgggtctgggacagacttcactctcaccatcagcagcctagagcctgaagattttgcagtttattactgtcagcagcgtagcaactggccgacgttcggccaagggaccaaggtggaaatcaaacgtacggtggctgcaccatctgtcttcatcttcccgccatctgatgagcagttgaaatctggaactgcctctgttgtgtgcctgctgaataacttctatcccagagaggccaaagtacagtggaaggtggataacgccctccaatcgggtaactcccaggagagtgtcacagagcaggacagcaaggacagcacctacagcctcagcagcaccctgacgctgagcaaagcagactacgagaaacacaaagtctacgcctgcgaagtcacccatcagggcctgagctcgcccgtcacaaagagcttcaacaggggagagtgt

[0654] SEQ ID NO: 16 – SG-559-01 hIgG1 heavy chain – nucleic acid

[0655]

[0656] SEQ ID NO:17 – Pembrolizumab heavy chain CDR1-protein

[0657] NYYMY

[0658] SEQ ID NO:18 – Pembrolizumab heavy chain CDR2-protein

[0659] GINPSNGGTNFNEKFKN

[0660] SEQ ID NO:19 – Pembrolizumab heavy chain CDR3-protein

[0661] RDYRFDMGFDY

[0662] SEQ ID NO:20 – Pembrolizumab light chain CDR1-protein

[0663] RASKGVSTSGYSYLH

[0664] SEQ ID NO:21 – Pembrolizumab light chain CDR2-protein

[0665] LASYLES

[0666] SEQ ID NO:22 – Pembrolizumab light chain CDR3-protein

[0667] QHSRDLPLT

[0668] SEQ ID NO:23 – Pembrolizumab heavy chain protein

[0669] QVQLVQSGVEVKKPGASVKVSCKASGYTFTNYYMYWVRQAPGQGLEWMGGINPSNGGTNFNEKFKNRVTLTTDSSTTTAYMELKSLQFDDTAVYYCARRDYRFDMGFDYWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK

[0670] SEQ ID NO: 24 – Pembrolizumab light chain - protein

[0671] EIVLTQSPATLSLSPGERATLSCRASKGVSTSGYSYLHWYQQKPGQAPRLLIYLASYLESGVPARFSGSGSGTDFTLTISSLEPEDFAVYYCQHSRDLPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0672] SEQ ID NO: 25 – Pembrolizumab heavy chain variable region - protein

[0673] QVQLVQSGVEVKKPGASVKVSCKASGYTFTNYYMYWVRQAPGQGLEWMGGINPSNGGTNFNEKFKNRVTLTTDSSTTTAYMELKSLQFDDTAVYYCARRDYRFDMGFDYWGQGTTVTVSS

[0674] SEQ ID NO: 26 – Pembrolizumab light chain variable region - protein

[0675] EIVLTQSPATLSLSPGERATLSCRASKGVSTSGYSYLHWYQQKPGQAPRLLIYLASYLESGVPARFSGSGSGTDFTLTISSLEPEDFAVYYCQHSRDLPLTFGGGTKVEIK。

Claims

1. A method for treating cancer in a human subject, comprising administering to the subject: (a) An effective amount of an antibody-drug conjugate (ADC) comprising an anti-PD-L1 antibody or its antigen-binding fragment, and (b) An effective amount of anti-PD-1 antibody; The anti-PD-L1 antibody or its antigen-binding fragment binds to PD-L1 and is conjugated to one or more units of monomethyl guanylate E (MMAE); The anti-PD-L1 antibody or its antigen-binding fragment comprises: (i) a light chain variable region comprising light chain CDR1, CDR2, and CDR3 of SEQ ID NO: 8, 9, and 10, respectively; and (ii) a heavy chain variable region comprising heavy chain CDR1, CDR2, and CDR3 of SEQ ID NO: 5, 6, and 7, respectively; and The anti-PD-1 antibody comprises: (i) a light chain variable region comprising light chain CDR1, CDR2 and CDR3 of SEQ ID NO: 20, 21 and 22, respectively, and (ii) a heavy chain variable region comprising heavy chain CDR1, CDR2 and CDR3 of SEQ ID NO: 17, 18 and 19, respectively.

2. The method of claim 1, wherein the anti-PD-L1 antibody or its antigen-binding fragment comprises: a heavy chain variable region comprising an amino acid sequence that is at least 85% identical to the amino acid sequence of SEQ ID NO:3 and a light chain variable region comprising an amino acid sequence that is at least 85% identical to the amino acid sequence of SEQ ID NO:

4.

3. The method of claim 1 or claim 2, wherein the anti-PD-L1 antibody or its antigen-binding fragment comprises: a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:3 and a light chain variable region comprising the amino acid sequence of SEQ ID NO:

4.

4. The method of any one of claims 1-3, wherein the anti-PD-L1 antibody or its antigen-binding fragment comprises: a heavy chain comprising the amino acid sequence of SEQ ID NO:1 and a light chain comprising the amino acid sequence of SEQ ID NO:

2.

5. The method of any one of claims 1-4, wherein the anti-PD-L1 antibody or its antigen-binding fragment is conjugated to the MMAE of each unit via a linker.

6. The method of claim 5, wherein the adapter is an enzyme-cleavable adapter, and wherein the adapter forms a bond with the sulfur atom of the antibody or its antigen-binding fragment.

7. The method of claim 5 or 6, wherein the connector has the formula: -Aa-Ww-Yy-; wherein -A- is a stretcher unit, a is 0 or 1; -W- is an amino acid unit, w is an integer in the range of 0 to 12; and -Y- is a spacer unit, y is 0, 1, or 2.

8. The method of claim 7, wherein the extended subunit has the structure of formula (1); the amino acid unit is valine-citrulline; and the spacer subunit is a PAB group comprising the structure of formula (2): 。 9. The method of claim 7 or 8, wherein the extension subunit forms a bond with the sulfur atom of the antibody or its antigen-binding fragment; and wherein the spacer subunit is linked to MMAE via a carbamate group.

10. The method of any one of claims 1-9, wherein the ADC comprises 1 to 20 units of MMAE for each antibody or its antigen-binding fragment.

11. The method of any one of claims 1-10, wherein the ADC comprises 1 to 10 units of MMAE for each antibody or its antigen-binding fragment.

12. The method of any one of claims 1-11, wherein the ADC comprises 2 to 8 units of MMAE for each antibody or its antigen-binding fragment.

13. The method of any one of claims 1-12, wherein the ADC comprises 3 to 5 units of MMAE for each antibody or its antigen-binding fragment.

14. The method of any one of claims 1-13, wherein the ADC has the following structure: Where L- represents the anti-PD-L1 antibody or its antigen-binding fragment, and p is 1 to 10.

15. The method of claim 14, wherein p is 2 to 8.

16. The method of claim 14, wherein p is 3 to 5.

17. The method of claim 14, wherein p is 4.

18. The method of claim 14, wherein p is 8.

19. The method of any one of claims 1-18, wherein the cancer is melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), head and neck squamous cell carcinoma (HNSCC), triple-negative breast cancer (TNBC), esophageal squamous cell carcinoma (esophageal SCC), ovarian cancer, urothelial carcinoma, hepatocellular carcinoma (HCC), gastric cancer, or cervical cancer.

20. The method of claim 19, wherein the cancer is NSCLC, HNSCC, TNBC, or esophageal SCC.

21. The method of claim 19, wherein the cancer is NSCLC.

22. The method of claim 19, wherein the cancer is HNSCC.

23. The method of claim 19, wherein the cancer is TNBC.

24. The method of claim 19, wherein the cancer is esophageal SCC.

25. The method of any one of claims 1-24, wherein the ADC is administered to the subject at a dose of about 0.25 to about 5 mg / kg of subject body weight, about 0.5 to about 2.5 mg / kg of subject body weight, or about 1 to about 2 mg / kg of subject body weight, optionally wherein the ADC is administered to the subject at a dose of 1.25 mg / kg or 1.5 mg / kg of subject body weight.

26. The method of any one of claims 1-25, wherein the subject's weight is the subject's ideal weight (IBW).

27. The method of any one of claims 1-25, wherein the subject's weight is the subject's adjusted ideal weight (AIBW).

28. The method of any one of claims 1-27, wherein the anti-PD-1 antibody is administered at a dose of 200 mg.

29. The method of any one of claims 1-28, wherein the ADC is administered approximately once every 1 week, approximately once every 2 weeks, approximately once every 3 weeks, or approximately twice every 3 weeks.

30. The method of any one of claims 1-29, wherein the ADC is administered approximately twice every 3 weeks.

31. The method of any one of claims 1-30, wherein the ADC is applied on the 1st and 8th days of each 21-day cycle.

32. The method of any one of claims 1-31, wherein the anti-PD-1 antibody is administered about once a week, about once every two weeks, about once every three weeks, or about twice every three weeks.

33. The method of any one of claims 1-32, wherein the anti-PD-1 antibody is administered approximately once a week.

34. The method of any one of claims 1-32, wherein the anti-PD-1 antibody is administered on day 1 of each 21-day cycle.

35. The method of any one of claims 1-34, wherein the subject has recurrent or refractory metastatic or unresectable solid malignant tumor disease that is intolerant to standard care.

36. The method of any one of claims 1-35, wherein the route of administration of the ADC is intravenous.

37. The method of any one of claims 1-36, wherein the route of administration of the anti-PD-1 antibody is intravenous.

38. The method of any one of claims 1-37, wherein, after administration of the antibody-drug conjugate, one or more therapeutic effects in the subject are improved relative to baseline.

39. The method of claim 38, wherein the one or more therapeutic effects are selected from the group consisting of: tumor size derived from the cancer, objective response rate, response duration, response time, progression-free survival, and overall survival.

Images