Anti-ILT2 / 4 monoclonal antibody preparation, kit containing same and use thereof

Abstract

An anti-ILT2 / 4 monoclonal antibody preparation, a kit containing same and the use thereof. The anti-ILT2 / 4 monoclonal antibody contains HCDR1, HCDR2 and HCDR3 having amino acid sequences as shown in SEQ ID NO: 1, SEQ ID NO: 29 and SEQ ID NO: 3, respectively, and LCDR1, LCDR2 and LCDR3 having amino acid sequences as shown in SEQ ID NOs: 4-6, respectively. The preparation can fully inhibit the aggregation, degradation and precipitation of the anti-ILT2 / 4 monoclonal antibody, thereby maintaining the biological activity of the effective components thereof, facilitating long-term storage and clinical use, and reducing adverse events in clinical use caused by changes in terms of drug quality.

Description

A preparation of anti-ILT2 / 4 monoclonal antibody, a drug kit containing the same, and its application

[0001] This application claims priority to Chinese patent application No. 2024101775792, filed on February 8, 2024. This application incorporates the entirety of the aforementioned Chinese patent application. Technical Field

[0002] The present invention belongs to the field of biomedicine, and specifically relates to an anti-ILT2 / 4 monoclonal antibody preparation, a drug kit containing the same, and applications thereof. Background Art

[0003] The immune system is a critical barrier to the development and progression of malignant tumors. Tumor immunotherapy, exemplified by immune checkpoint blockade (ICB), has achieved remarkable success in recent years. However, clinical application of tumor immunotherapy still faces significant challenges, such as low response rates, primary and secondary drug resistance. The development of new immune checkpoint molecules and combination therapy strategies will be key to remodeling T cell anti-tumor activity and overcoming clinical immune resistance.

[0004] ILT2 stands for Immunoglobulin-like transcript 2, immunoglobulin-like transcription factor 2, also known as lymphocyte immunoglobulin-like receptor B1 (LILRB1). ILT4 stands for Immunoglobulin-like transcript 4, immunoglobulin-like transcription factor 4, also known as lymphocyte immunoglobulin-like receptor B2 (LILRB2). They are two inhibitory receptors in the immunoglobulin-like transcript (ILTs) family. They have similar structures. Their extracellular segments contain four Ig-like domains and four and three immunoreceptor tyrosine-based inhibitory motif (ITIM) domains, respectively, within the cell. They transmit inhibitory signals to the cell through these ITIM domains, thereby regulating the function of immune cells. It is precisely this structural similarity that makes it possible to use the same antibody to simultaneously inhibit the functions of ILT2 and ILT4.

[0005] ILT2 and ILT4 share common ligands including class I HLA and HLA-G, with HLA-G having a higher affinity than other class I HLA members. When ILT2 / 4 bind to HLA, the intracellular ITIM recruits SH-2 domain-containing SHP-1 or SHP-2 phosphatases, inhibiting activation signaling and immune function in these cells.

[0006] Under physiological conditions, ILT4 is primarily expressed on the surface of myeloid cells, including monocytes, macrophages, dendritic cells, and neutrophils. In addition to being expressed on these cells, ILT2 is also expressed on some NK, T, and B cells. Within the tumor microenvironment, ILT2 or ILT4 is highly expressed in tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs), and DCs, driving the transformation of the tumor microenvironment toward an immunosuppressive state, thereby promoting tumor growth.

[0007] A recombinant, fully human anti-ILT2 / 4 S228P IgG4κ monoclonal antibody, designated anti-ILT2 / 4, binds to cell surface ILT2 / ILT4 molecules with similar potency, thereby reprogramming tumor-associated myeloid cells into a proinflammatory state. The resulting proinflammatory response may better stimulate anti-tumor T cell responses (especially when combined with PD-1 / PD-L1 antibodies), thereby activating the tumor immune microenvironment and ultimately treating the tumor.

[0008] As biological macromolecules, they are easily affected by physical (such as freeze-thaw, oscillation, light, temperature, etc.) or chemical (such as pH, organic solvents, microbial contamination, etc.) factors, which may lead to aggregation, degradation, reduced purity, reduced activity, etc., resulting in reduced clinical efficacy of the molecule and may even endanger patient health during clinical use. Summary of the Invention

[0009] In order to solve the problem that anti-ILT2 / 4 monoclonal antibodies are easily affected by physical (such as freeze-thaw, oscillation, light, temperature, etc.) or chemical factors (such as pH, organic solvents, microbial contamination, etc.), resulting in aggregation, degradation, reduced purity, and reduced activity, the present invention provides a preparation of anti-ILT2 / 4 monoclonal antibodies, a drug kit containing the same, and its use. The inventors of the present invention screened and studied the formulation of anti-ILT2 / 4 preparations and ultimately obtained a solution preparation that is most suitable for anti-ILT2 / 4 monoclonal antibodies and can stably preserve the monoclonal antibodies. The preparation can fully inhibit the aggregation, degradation, precipitation, etc. of anti-ILT2 / 4 proteins, thereby maintaining the biological activity of its effective components, facilitating long-term storage and clinical use, and greatly reducing adverse events in clinical use caused by changes in drug quality.

[0010] In order to solve the above technical problems, the first aspect of the present invention provides a preparation of an anti-ILT2 / 4 monoclonal antibody, wherein the anti-ILT2 / 4 monoclonal antibody comprises HCDR1, HCDR2 and HCDR3 as shown in SEQ ID NO: 1, SEQ ID NO: 29 and SEQ ID NO: 3, respectively, wherein X1 is N, A or T, X2 is G, A or S, and LCDR1, LCDR2 and LCDR3 as shown in SEQ ID NOs: 4 to 6, respectively.

[0011] In some embodiments of the present invention, the anti-ILT2 / 4 monoclonal antibody comprises HCDR1, HCDR2 and HCDR3 with amino acid sequences as shown in SEQ ID NO: 1, SEQ ID NO: 7 and SEQ ID NO: 3, respectively, and LCDR1, LCDR2 and LCDR3 with amino acid sequences as shown in SEQ ID NOs: 4 to 6, respectively.

[0012] In some embodiments of the present invention, the anti-ILT2 / 4 monoclonal antibody comprises HCDR1, HCDR2 and HCDR3 whose amino acid sequences are shown in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3, respectively, and LCDR1, LCDR2 and LCDR3 whose amino acid sequences are shown in SEQ ID NOs: 4 to 6, respectively.

[0013] In some embodiments of the present invention, the anti-ILT2 / 4 monoclonal antibody comprises HCDR1, HCDR2 and HCDR3 whose amino acid sequences are shown in SEQ ID NO: 1, SEQ ID NO: 8 and SEQ ID NO: 3, respectively, and LCDR1, LCDR2 and LCDR3 whose amino acid sequences are shown in SEQ ID NOs: 4 to 6, respectively.

[0014] In some embodiments of the present invention, the anti-ILT2 / 4 monoclonal antibody comprises HCDR1, HCDR2 and HCDR3 whose amino acid sequences are shown in SEQ ID NO: 1, SEQ ID NO: 9 and SEQ ID NO: 3, respectively, and LCDR1, LCDR2 and LCDR3 whose amino acid sequences are shown in SEQ ID NOs: 4 to 6, respectively.

[0015] In some embodiments of the present invention, the anti-ILT2 / 4 monoclonal antibody comprises HCDR1, HCDR2 and HCDR3 with amino acid sequences as shown in SEQ ID NO: 1, SEQ ID NO: 10 and SEQ ID NO: 3, respectively, and LCDR1, LCDR2 and LCDR3 with amino acid sequences as shown in SEQ ID NOs: 4 to 6, respectively.

[0016] In some embodiments of the present invention, the VH of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as shown in SEQ ID NO: 30, or at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 30, wherein X is N, A or T, and X2 is G, A or S; the VL of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as shown in SEQ ID NO: 12, or at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 12.

[0017] In some embodiments of the present invention, the VH of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 13; and the VL of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 12.

[0018] In some embodiments of the present invention, the VH of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 11; and the VL of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 12.

[0019] In some embodiments of the present invention, the VH of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 14; and the VL of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 12.

[0020] In some embodiments of the present invention, the VH of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 15; and the VL of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 12.

[0021] In some embodiments of the present invention, the VH of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 16; and the VL of the anti-ILT2 / 4 monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO: 12.

[0022] In some preferred embodiments of the present invention, the HC of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as shown in SEQ ID NO: 19, SEQ ID NO: 17, SEQ ID NO: 20, SEQ ID NO: 21 or SEQ ID NO: 22, or an amino acid sequence that is at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 19, SEQ ID NO: 17 or SEQ ID NO: 20; the LC of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as shown in SEQ ID NO: 18, or an amino acid sequence that is at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 18.

[0023] In some more preferred embodiments of the present invention, the formulation further comprises a buffer, a stabilizer and / or a surfactant.

[0024] In some embodiments of the present invention, the buffer is selected from acetic acid-sodium acetate buffer and PB buffer (sodium dihydrogen phosphate-disodium hydrogen phosphate buffer).

[0025] In some preferred embodiments of the present invention, the buffer is acetic acid-sodium acetate buffer.

[0026] In some preferred embodiments of the present invention, the concentration of the buffer is not less than 10 mM, for example, 10 mM, 20 mM, 30 mM, 40 mM or 50 mM.

[0027] In some preferred embodiments of the present invention, the concentration of the buffer is 10 mM.

[0028] In some embodiments of the present invention, the pH range of the formulation is pH 4-6.

[0029] In some preferred embodiments of the present invention, the pH range of the preparation is pH 4.5 to 5.5.

[0030] In some embodiments of the present invention, the pH range of the formulation is pH 4.0, 4.5, 5.0 or 5.5.

[0031] In some embodiments of the present invention, the stabilizer is sucrose and / or trehalose.

[0032] In some preferred embodiments of the present invention, the stabilizer is trehalose.

[0033] In some preferred embodiments of the present invention, the concentration of the stabilizer ranges from 1% to 20%.

[0034] In some preferred embodiments of the present invention, the concentration of the stabilizer ranges from 7% to 9%.

[0035] In some embodiments of the present invention, the concentration of the stabilizer is 7%, 8% or 9%.

[0036] In some embodiments of the present invention, the surfactant is selected from one or more of polysorbate 20, polysorbate 80 and poloxamer 188.

[0037] In some preferred embodiments of the present invention, the surfactant is polysorbate 20.

[0038] In some preferred embodiments of the present invention, the concentration of the surfactant is in the range of 0.001% to 0.1%.

[0039] In some preferred embodiments of the present invention, the concentration of the surfactant is in the range of 0.01 to 0.1%.

[0040] In some more preferred embodiments of the present invention, the concentration of the surfactant is in the range of 0.02% to 0.06%.

[0041] In some specific embodiments of the present invention, the concentration of the surfactant is 0.02%, 0.04% or 0.06%.

[0042] In some embodiments of the present invention, the concentration of the anti-ILT2 / 4 monoclonal antibody is 1 to 30 mg / mL.

[0043] In some preferred embodiments of the present invention, the concentration of the anti-ILT2 / 4 monoclonal antibody is 10-30 mg / mL.

[0044] In some more preferred embodiments of the present invention, the concentration of the anti-ILT2 / 4 monoclonal antibody is 15-25 mg / mL.

[0045] In some specific embodiments of the present invention, the concentration of the anti-ILT2 / 4 monoclonal antibody is 15 mg / mL, 20 mg / mL or 25 mg / mL.

[0046] In some embodiments of the present invention, the formulation is a liquid formulation.

[0047] In some specific embodiments of the present invention, the pH of the anti-ILT2 / 4 monoclonal antibody formulation is 5.0, the concentration of the anti-ILT2 / 4 monoclonal antibody is 20 mg / mL, the polysorbate 20 content is 0.04%, and the trehalose content is 8%; or

[0048] The pH is 4.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 15 mg / mL, the polysorbate 20 content is 0.02%, and the trehalose content is 7%; or,

[0049] The pH is 5.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 15 mg / mL, the polysorbate 20 content is 0.06%, and the trehalose content is 7%; or,

[0050] The pH is 5.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 25 mg / mL, the polysorbate 20 content is 0.02%, and the trehalose content is 7%; or,

[0051] The pH is 4.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 15 mg / mL, the polysorbate 20 content is 0.06%, and the trehalose content is 9%; or,

[0052] The pH is 5.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 25 mg / mL, the polysorbate 20 content is 0.06%, and the trehalose content is 9%; or,

[0053] The pH is 4.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 25 mg / mL, the polysorbate 20 content is 0.02%, and the trehalose content is 9%; or,

[0054] The pH is 5.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 15 mg / mL, the polysorbate 20 content is 0.02%, and the trehalose content is 9%; or,

[0055] The pH is 4.5, the concentration of the anti-ILT2 / 4 monoclonal antibody is 25 mg / mL, the polysorbate 20 content is 0.06%, and the trehalose content is 7%, where % represents the mass-to-volume ratio.

[0056] In order to solve the above technical problems, the second aspect of the present invention provides a medicine box containing the preparation as described in the first aspect of the invention, characterized in that the medicine box also includes a container.

[0057] In some preferred embodiments of the present invention, the container is a polycarbonate bottle, a polypropylene bottle (PP bottle), an ultra-low density polyethylene bag (ULDPE bag) or an ethylene-vinyl acetate copolymer bag (EVA bag).

[0058] In order to solve the above technical problems, the third aspect of the present invention provides the use of the preparation according to the first aspect of the invention in the preparation of products for diagnosing, treating and / or preventing diseases or conditions mediated by ILT4 and / or ILT2.

[0059] In some preferred embodiments of the present invention, the disease or condition is cancer.

[0060] In some more preferred embodiments of the present invention, the cancer is ILT4-positive and / or ILT2-positive cancer.

[0061] In some further preferred embodiments of the present invention, the cancer is a solid cancer or a blood cancer;

[0062] The blood cancer is, for example, B-cell leukemia, lymphoma, acute myeloid leukemia, Burkitt's lymphoma, mantle cell lymphoma, acute lymphoblastic leukemia, chronic lymphocytic leukemia, diffuse large B-cell lymphoma, follicular lymphoma, or marginal zone lymphoma;

[0063] The solid cancer is, for example, head and neck squamous cell carcinoma, renal cell carcinoma, ovarian cancer, urothelial carcinoma, diffuse large B-cell carcinoma, non-small cell lung cancer, melanoma, metastatic renal cell carcinoma, metastatic colorectal cancer, small cell lung cancer, metastatic non-small cell lung cancer, breast cancer, lung cancer, head and neck cancer, colorectal cancer, prostate cancer, skin cancer, stomach cancer, intestinal cancer, cervical cancer, uterine cancer, endometrial cancer, bladder cancer, brain cancer, esophageal cancer, liver cancer, kidney cancer, testicular cancer, mesothelioma, glioblastoma, bile duct cancer, or pancreatic cancer.

[0064] In order to solve the above technical problems, the fourth aspect of the present invention provides a method for immunodetection or determination of ILT4 and / or ILT2, which comprises contacting the preparation described in the first aspect of the present invention with a sample, and determining whether ILT4 and / or ILT2 are present in the sample based on the combined situation.

[0065] In some preferred embodiments of the present invention, the detection is for non-diagnostic purposes, and the applicable scenarios of the non-diagnostic purposes can be laboratory research and development or immune detection or determination in the environment, etc.

[0066] In order to solve the above technical problems, the fifth aspect of the present invention provides a method for treating and / or preventing diseases or conditions mediated by ILT4 and / or ILT2, which comprises administering a therapeutically effective amount of the preparation as described in the first aspect of the invention to a patient in need.

[0067] In some preferred embodiments of the present invention, the disease or condition is cancer.

[0068] In some more preferred embodiments of the present invention, the cancer is ILT4-positive and / or ILT2-positive cancer.

[0069] In some further preferred embodiments of the present invention, the cancer is a solid cancer or a blood cancer;

[0070] The blood cancer is, for example, B-cell leukemia, lymphoma, acute myeloid leukemia, Burkitt's lymphoma, mantle cell lymphoma, acute lymphoblastic leukemia, chronic lymphocytic leukemia, diffuse large B-cell lymphoma, follicular lymphoma, or marginal zone lymphoma;

[0071] The solid cancer is, for example, head and neck squamous cell carcinoma, renal cell carcinoma, ovarian cancer, urothelial carcinoma, diffuse large B-cell carcinoma, non-small cell lung cancer, melanoma, metastatic renal cell carcinoma, metastatic colorectal cancer, small cell lung cancer, metastatic non-small cell lung cancer, breast cancer, lung cancer, head and neck cancer, colorectal cancer, prostate cancer, skin cancer, stomach cancer, intestinal cancer, cervical cancer, uterine cancer, endometrial cancer, bladder cancer, brain cancer, esophageal cancer, liver cancer, kidney cancer, testicular cancer, mesothelioma, glioblastoma, bile duct cancer, or pancreatic cancer.

[0072] In order to solve the above technical problems, the sixth aspect of the present invention provides a combination therapy, which comprises administering the preparation as described in the first aspect of the invention to patients in need; and a second therapeutic agent; the second therapeutic agent preferably comprises other anti-tumor antibodies or a pharmaceutical composition comprising the other anti-tumor antibodies, and / or one or more of a group consisting of hormone preparations, targeted small molecule preparations, proteasome inhibitors, imaging agents, diagnostic agents, chemotherapeutic agents, oncolytic drugs, cytotoxic agents, cytokines, activators of co-stimulatory molecules, inhibitors of inhibitory molecules and vaccines.

[0073] In order to solve the above technical problems, the seventh aspect of the present invention provides a preparation as described in the first aspect of the present invention, which is used to treat and / or prevent diseases or conditions mediated by ILT4 and / or ILT2.

[0074] In some preferred embodiments of the present invention, the disease or condition is cancer.

[0075] In some more preferred embodiments of the present invention, the cancer is ILT4-positive and / or ILT2-positive cancer.

[0076] In some further preferred embodiments of the present invention, the cancer is a solid cancer or a blood cancer;

[0077] The blood cancer is, for example, B-cell leukemia, lymphoma, acute myeloid leukemia, Burkitt's lymphoma, mantle cell lymphoma, acute lymphoblastic leukemia, chronic lymphocytic leukemia, diffuse large B-cell lymphoma, follicular lymphoma, or marginal zone lymphoma;

[0078] The solid cancer is, for example, head and neck squamous cell carcinoma, renal cell carcinoma, ovarian cancer, urothelial carcinoma, diffuse large B-cell carcinoma, non-small cell lung cancer, melanoma, metastatic renal cell carcinoma, metastatic colorectal cancer, small cell lung cancer, metastatic non-small cell lung cancer, breast cancer, lung cancer, head and neck cancer, colorectal cancer, prostate cancer, skin cancer, stomach cancer, intestinal cancer, cervical cancer, uterine cancer, endometrial cancer, bladder cancer, brain cancer, esophageal cancer, liver cancer, kidney cancer, testicular cancer, mesothelioma, glioblastoma, bile duct cancer, or pancreatic cancer.

[0079] On the basis of conforming to the common sense in this field, the above-mentioned preferred conditions can be arbitrarily combined to obtain the preferred embodiments of the present invention.

[0080] The reagents and raw materials used in the present invention are commercially available.

[0081] The positive progress of the present invention is that the preparation provided by the present invention can improve the storage stability of anti-ILT2 / 4 antibodies, fully inhibit the aggregation, degradation, precipitation, etc. of anti-ILT2 / 4 proteins, and can maintain the biological activity of its effective components during long-term storage without significant changes in various quality indicators, thereby facilitating clinical use and greatly reducing adverse events caused by changes in drug quality during clinical use. BRIEF DESCRIPTION OF THE DRAWINGS

[0082] FIG1 is a graph showing the binding ability of ILT4 antibodies to human / monkey ILT4-ECD proteins.

[0083] FIG2 is an ELISA assay to detect the binding ability of ILT-4 antibodies to ILT2-ECD protein.

[0084] FIG3 shows the activation effect of ILT4 antibodies on macrophages detected by ELISA.

[0085] FIG4 shows the activation effect of ILT4 antibodies on peripheral blood mononuclear cells detected by ELISA.

[0086] FIG5 is an ELISA-based assay for the activation of ILT4 antibodies in mixed lymphocyte reaction.

[0087] FIG6 is a pharmacodynamics experiment of ILT4 antibody in SU8686 mouse tumor model.

[0088] Figure 7 is a trend diagram of the effects of protein concentration and pH on aggregates of samples accelerated at 40°C for 2 weeks obtained by DOE software analysis.

[0089] Figure 8 shows the influence of protein concentration and pH on the content of small molecule components in CE-SDS(NR) samples accelerated at 40°C for 2 weeks, as analyzed by DOE software. W stands for week, and 40°C 2W stands for 40°C for 2 weeks. DETAILED DESCRIPTION

[0090] The present invention is further illustrated by way of examples below, but the present invention is not limited to the scope of the examples. Experimental methods in the following examples where specific conditions are not specified were performed according to conventional methods and conditions, or selected according to the product specifications.

[0091] Example 1

[0092] Example 1 Antibody affinity detection experiment

[0093] Harbour H2L2 transgenic mice (Harbour Antibodies BV, a transgenic mouse carrying a human immunoglobulin immune repertoire, producing antibodies with complete human variable domains and rat constant domains) were immunized with the huILT4-ECD-Fc protein (UniprotKB NO. Q8N423 extracellular region positions 1-460). Antibodies with the sequences shown in Table 1 were obtained through IgG subtype conversion and PTM optimization screening. The antibodies listed in Table 1 were tested for their binding kinetics to human ILT4-ECD protein using biolayer interferometry (BLI) (Fortebio octet RED 96e).

[0094] The test antibody was diluted to a final concentration of 6 μg / mL and directly immobilized onto the AHC biosensor for kinetic measurements. The antigen protein (HuILT4, UniprotKB No. Q8N423) was diluted in 0.02% PBST20 to three concentrations: 100 nM, 50 nM, and 25 nM. The sample was injected for 70 s, with an association time of 300 s and a dissociation time of 300 s. The sample was then regenerated in 10 mM glycine-HCl (pH 1.5) for 15 s. The association rate (k₁₀) and dissociation rate (k₁₀) were calculated using a simple one-to-one Languir binding model (Octet Red 96 data analysis software). The equilibrium dissociation constant (k₁₀) was calculated as the ratio k₁₀ / k₁₀. The antibody affinity test results are shown in Table 2 below.

[0095] Table 1 Antibody sequences

[0096] Table 2 Antibody affinity test results

[0097] As can be seen from the results in Table 2, the affinity of PR302264, PR302264-2 and PR302264-3 antibodies is better than that of the control antibody PR001795 (Merck: US20180298096).

[0098] Example 2 Detection of the binding ability of antibodies to ILT4-ECD at the protein level

[0099] The binding ability of the antibodies listed in Table 1 to the ILT4-ECD protein was determined by ELISA. Binding ability was determined by comparing the binding curves of different antibodies to the ILT4-ECD protein. The specific experimental procedure was as follows: human ILT4-ECD and cynomolgus macaque ILT4-ECD proteins were diluted sequentially to a concentration of 1 μg / mL and added to a 96-well plate, 100 μL per well, and incubated at 4°C overnight. The 96-well plate was washed three times with PBST and then incubated at 37°C for 1 hour with 2% BSA in PBS. The test antibody was serially diluted (100 nM, 10-fold dilution) and added to the 96-well plate. The plate was incubated at 37°C for 1 hour. After washing three times with PBST, an anti-human IgG Fc-HRP secondary antibody (5000x dilution) was added and incubated at 37°C for 30-60 minutes. After washing three times with PBST, TMB color development solution was added for 5-15 minutes, and then color development was terminated with stop solution.

[0100] Table 3 Results of antibody binding ability test with ILT4-ECD at protein level

[0101] The test results are shown in Table 3 and Figure 1. The PR302264 antibody has the ability to bind to the human ILT4-ECD protein, and is comparable to the control antibody PR301366 (Five Prime: WO2020014132); and the PR302264 antibody also has the ability to bind to the monkey ILT4-ECD protein.

[0102] Example 3 Detection of Antibody Binding Ability to ILT2-ECD at the Protein Level

[0103] The antibodies listed in Table 1 were tested for their binding ability to the ILT2-ECD (UniprotKB No. Q8NHL6) protein using an ELISA assay. Binding ability was determined by comparing the binding curves of different antibodies to the ILT2-ECD protein. The specific experimental procedure was as follows: Human ILT2-ECD protein (UniprotKB No. Q8NHL6) was serially diluted to a concentration of 1 μg / mL and added to a 96-well plate (100 μL per well) and incubated at 4°C overnight. The 96-well plate was washed three times with PBST and then incubated at 37°C for 1 hour with 2% BSA in PBS. The test antibody was serially diluted (100 nM, 10-fold dilution) and added to the 96-well plate. The plate was incubated at 37°C for 1 hour. After washing three times with PBST, an anti-human IgG Fc-HRP secondary antibody (5000x dilution) was added and incubated at 37°C for 30-60 minutes. After washing three times with PBST solution, TMB colorimetric solution was added for 5-15 minutes, and then the color development was terminated by adding stop solution. The test results are shown in Figure 2. PR302264-2 and PR302264-3 antibodies have strong binding ability to human ILT2.

[0104] Example 4 Cross-reaction of antibodies with other ILT4 family proteins

[0105] Take 100 μL of human LILRA1 (R&D System, Cat: 9226-T4-050), LILRA2 (R&D System, Cat: 9040-T4-050), LILRA3 (R&D System, Cat: 9517-T4-100), LILRA4 (R&D System, Cat: 9517-T4-100) with a concentration of 1 μg / mL. System, Cat: 8914-T4-050), LILRA5 (R&D System, Cat: 8956-T4-100), LILRA6 (R&D System, Cat: 9088-T4-050), LILRB3 (R&D System, Cat: 9159-T5-050), LILRB4 (R&D System, Cat: 8488-T4-025), LILRB5 (R&D ELISA coating was performed using the protein from the ELISA system (Cat. No. 8478-T4-025) at 4°C overnight. The cells were washed four times with PBST and blocked with 200 μL of a solution of PBST (PBS + 0.05% Tween 20) + 2% BSA at 37°C for 1 hour. The cells were then washed four times with PBST. Add 100 μL of 100 nM ILT4 antibody and positive control antibodies of each protein (LILRA1 Antibody: R&D System, Cat: MAB30851; LILRA2 Antibody: R&D System, Cat: MAB6364; LILRA3 Antibody: R&D System, Cat: MAB2574; LILRA4 Antibody: R&D System, Cat: MAB6287; LILRA5 Antibody: R&D System, Cat: MAB6754; LILRA6 Antibody: R&D System, Cat: MAB8656; LILRA3 Antibody: R&D System, Cat: MAB1806-100; LILRA4 Antibody: R&D System, Cat: MAB24251; LILRA5 Antibody: R&D System, Cat: MAB3065) to the ELISA plate wells respectively. Incubate at 37°C for 1 hour. Wash four times with PBST. Add 100 μl of Goat Anti-Human IgG, (Fab)-HRP (1:5000, Jackson, Cat: 109-035-098) and incubate at 37°C for 1 hour. Wash four times with PBST.100 μL of TMB substrate (Biopanda, Cat: TMB-S-003) was added and incubated at room temperature for 5 minutes. The reaction was then terminated by adding 50 μL of stop buffer (Solarbio, Cat#: C1058). OD 450 nm readings were recorded using a microplate reader (Molecular Devices, Spectramax 384plus). The control antibody PR304683 (NGM: WO2021127200) was used.

[0106] Table 4 Cross-reaction results of antibodies with other ILT4 family proteins

[0107] The results are shown in Table 4. In addition to binding to ILT4, PR302264-2 has cross-binding ability with LIR6 and ILT2.

[0108] Example 5 Detection of Antibody Activation Effect on Macrophages - Cytokine TNF-α Secretion

[0109] CD14 positive cells were isolated from human PBMC (Shanghai Saili Biotechnology Co., Ltd., XFB-HP050A) using a human CD14 cell isolation kit (Miltenyi, 130-050-021). The isolated CD14 positive cells were resuspended and diluted to 8 × 10 5 / mL, 100 μL of cell suspension was added to each well of a 96-well cell culture plate. 50 μL of serially diluted (5-fold dilutions, starting at a final concentration of 50 nM) antibody was then added to the 96-well plate. MCSF (R&D System, 216-MC-025 / CF) was diluted to 400 ng / mL and 50 μL was added to each well. The cells were incubated at 37°C in a CO2 incubator. On the third day, the medium was changed, maintaining the same antibody and MCSF concentrations. On the sixth day, the medium was changed again, maintaining the same antibody and MCSF concentrations, and LPS was added to a final concentration of 50 ng / mL. After 24 hours, the cell supernatant was collected and TNF-α levels were measured using an ELISA kit (Ebioscience, 88-7346-88). As shown in Figure 3, the PR302264 antibody had a strong activating effect on macrophages.

[0110] Example 6 Detection of the Effect of Antibodies on Peripheral Blood Mononuclear Cells - Cytokine TNF-α Secretion

[0111] Use healthy human PBMC and resuspend the cells in 1640 complete medium and dilute them to 2*10 6 / mL, 100μL of cell suspension was added to each well of a 96-well cell culture plate. 50μL of serially diluted antibodies (maximum final concentration: 30nM, diluted to 1nM, and then diluted five-fold) were added to the 96-well plate. LPS was diluted to 400ng / mL and 50μL was added to each well. After three days of incubation at 37°C in a CO2 incubator, the cell supernatant was collected and assayed for TNF-α levels using an ELISA kit. The results, as shown in Figure 4, show that antibodies PR302264-2 and PR302264-3 have a strong activating effect on PBMCs.

[0112] Example 7: Detection of the Effect of Antibodies in Mixed Lymphocyte Reaction - Cytokine IFN-γ Secretion

[0113] CD14 positive cells were isolated from human PBMC using a human CD14 cell isolation kit (Miltenyi, 130-050-021). The isolated CD14 positive cells were resuspended and diluted to 1.5*10 5 / mL, take a 96-well cell culture plate and add 100μL of cell suspension to each well; take 50μL of serially diluted (5-fold dilution, starting and ending at 50nM) antibody to the 96-well cell plate; dilute MSCF (R&D System, 216-MC-025 / CF) to 400ng / mL and add 50μL to each well. Incubate at 37℃ in a CO2 incubator. Change the medium on the third day, keeping the antibody and MCSF concentrations unchanged. On the sixth day, change the medium again, keeping the antibody and MCSF concentrations unchanged, and add LPS to a final concentration of 50ng / mL. On the seventh day, take another human PBMC and sort out CD3-positive cells using the human CD3 sorting kit (Miltenyi, 130-045-501). The isolated CD3-positive cells are resuspended and diluted to 1.5*10 in 1640 complete medium. 6 / mL. The supernatant from the 96-well cell culture plate was removed, and 100 μL of CD3-positive cell suspension and 50 μL of diluted antibody were added, and the volume was finally refilled to 200 μL. After 72 hours of co-incubation, the cell supernatant was collected and the IFN-γ content was measured using an ELISA kit. The results, as shown in Figure 5, show that the antibody PR302264-2 exhibited a strong activating effect in the mixed lymphocyte reaction assay.

[0114] Example 8 In vivo pharmacodynamics experiments of antibodies

[0115] This study investigated the antitumor activity of an anti-human ILT4 antibody using the SU8686 mouse tumor model. The pharmacodynamics of PR302264-2 antibody administered alone (20 mg / kg) in the SU8686 mouse model were investigated. Human pancreatic cancer SU8686 cells (ATCC) were subcutaneously inoculated into female hHSC-NOG-EXL humanized mice (Vitamin B). The antibody group was treated as follows: After tumor cell inoculation, the tumors were allowed to grow to 100 mm in size. 3 The first administration was started at 3:10 p.m. on the 3rd, 7th, 10th, 14th, and 17th days, and the corresponding antibodies were intraperitoneally injected into each group of mice. The tumor volume was measured at the same time, and the mice were euthanized. As shown in Figure 6, the anti-human ILT4 antibody PR302264-2 of the present application has a clear inhibitory effect on tumor growth compared with the negative control, among which the inhibitory effect of PR302264-2 on tumors is better than that of the control antibodies PR001795 (Merck: US20180298096) and PR304683 (NGM: WO2021127200).

[0116] Example 9 pH screening

[0117] To identify the optimal pH range for the anti-ILT2 / 4 antibody (PR302264-2), the inventors conducted a pH gradient screening of the molecule. This screening examined the antibody's purity across the pH range of 4 to 8, including high-temperature accelerated, freeze-thaw, and light-induced precipitation, as well as changes in size exclusion chromatography (SEC), purity of the acidic and basic peaks, and the main peak measured by capillary electrophoresis with sodium dodecyl sulfate (CE-SDS(NR)), and whole-column imaging capillary isoelectric focusing (icIEF), as well as changes in binding activity. The results were summarized in Tables 5-9.

[0118] Size Exclusion Chromatography (SEC):

[0119] Experimental conditions were set up as follows: Waters 2695 liquid chromatograph (Waters, USA); TSKgelG3000SWXL tandem chromatographic columns (purchased from Waters, USA); the mobile phase was 0.2 M phosphate buffer, 0.25 M potassium chloride, pH 6.2, 0.5 mL / min, the injection volume was 100 μg, and the detection wavelength was 280 nm.

[0120] Capillary electrophoresis of sodium dodecyl sulfate (CE-SDS(NR)):

[0121] Capillary electrophoresis was performed using a SCIEX capillary electrophoresis instrument (PA800PLUS, purchased from SCIEX), using a SCIEX capillary cartridge (Cat. No. U738), SCIEX capillaries (Cat. No. 338451), and a SCIEX low-pH SDS (Cat. No. C57805). 25 μL of the 4.0 mg / mL sample solution was mixed, followed by the addition of 70 μL of low-pH SDS and 5 μL of 200 μM nem. After centrifugation, the sample was incubated in a 70°C water bath for 10 minutes and then cooled. Injection was performed at 5 kV for 20 seconds, and separation was performed at 15 kV for 40 minutes. A UV detector was used with a wavelength set at 214 nm.

[0122] Whole-column imaging capillary isoelectric focusing (icIEF):

[0123] Experimental conditions: ProteinSimple Maurice capillary isoelectric focusing instrument (Protein Simple, USA); amphoteric electrolyte solution: 3-10 μL Pharmalyte 3-10 (purchased from GE, USA, catalog number: 17045501), 8-10.5 μL Pharmalyte 3-10 (purchased from GE, USA, catalog number: 17045601), 0.5 μL pI Marker 7.05 (purchased from Protein Simple, USA, catalog number: 102226), 0.5 μL pI Marker 9.46 (purchased from Protein Simple, USA, catalog number: 102349), appropriate amount of 1% methylcellulose (purchased from Protein Simple, catalog number: 101876), appropriate amount of 0.5% methylcellulose (purchased from Protein Simple, catalog number: 101876), appropriate amount of 0.5% methylcellulose (purchased from Protein Simple, catalog number: Simple, Cat. No. 102505), an appropriate amount of sulfonated dimethylethylamine (NDSB) (Merck, D01955G), and an appropriate amount of taurine (T) (Sigma, T0625-100G). The injection solution consisted of 80 μL of the above-mentioned ampholyte solution and 20 μL of a 1.0 mg / mL protein solution. The sample system was pre-focused at 1500 V for 1 minute and focused at 3000 V for 10 minutes before detection.

[0124] Binding activity assay:

[0125] The binding activity of ILT2 and ILT4 was detected by ELISA.

[0126] Table 5 Zero point sample results

[0127] Table 6 Results of samples at 40℃ in the dark for 7 days

[0128] Compared with Table 6, there were relatively fewer aggregates and relatively less aggregation increase at pH 4-6, indicating that pH 4-6 is a relatively suitable buffer pH for anti-ILT2 / 4 antibodies.

[0129] Table 7 Results of samples irradiated at 40℃ for 7 days

[0130] As shown in Table 7, compared with Table 4, under strong light irradiation conditions, the aggregates further increased compared with the light-proof experimental group, indicating that strong light irradiation increased antibody aggregation and precipitation.

[0131] Table 8 Freeze-thaw experiment results

[0132] The freeze-thaw experiment found that, as shown in Table 8, a large amount of precipitation appeared in the samples in freeze-thaw result 1. In a subsequent supplementary experiment, the surfactant polysorbate 20 was added to the freeze-thaw result 1. It was found that the precipitation of the freeze-thaw samples was significantly reduced after the addition of polysorbate 20. However, the precipitation of the samples with pH 7 to pH 8 was not significantly reduced after the addition of polysorbate 20. Therefore, the freeze-thaw experiment was supplemented and freeze-thaw was performed after adding polysorbate 20.

[0133] Table 9 shows the results of freeze-thaw supplementation experiments.

[0134] As shown in Table 9, no precipitation was observed in the samples of the supplementary freeze-thaw experiment, which proves that the addition of surfactant excipients such as polysorbate 20, polysorbate 80, and poloxamer 188 to the formulation can effectively inhibit protein precipitation.

[0135] Therefore, a pH 4-6 formulation, the addition of polysorbate 20, and the addition of sugar alcohol stabilizers such as sucrose and trehalose can effectively improve the stability of anti-ILT2 / 4 antibodies.

[0136] Example 10 Excipient Screening

[0137] Excipient screening was performed using Design of Experiments (DOE), and the effects of pH, antibody concentration, polysorbate 20 content, and trehalose content on the stability of the anti-ILT2 / 4 antibody were simultaneously studied using statistical methods. The specific research conditions are shown in Table 10.

[0138] Table 10 Effects of pH, antibody concentration, polysorbate 20 content, and trehalose content on the stability of anti-ILT2 / 4 antibodies

[0139] The results of the DOE experimental samples are shown in Table 11.

[0140] Table 11 DOE experimental sample results Note: Binding activities are for ILT2 and ILT4 targets respectively.

[0141] The freeze-thaw and oscillation results showed that the anti-ILT2 / 4 antibody had good stability within the above experimental research parameter range, and there was no significant change in various quality parameters.

[0142] The 40°C accelerated results showed that the aggregates, CE-SDS (NR) small molecule components, and icIEF acidic peaks of different formulations increased to varying degrees. Data statistical analysis using DOE software showed that the aggregates increased with increasing protein concentration and pH (see Figure 7); the CE-SDS (NR) small molecule components increased with increasing protein concentration and decreasing pH (see Figure 8). The two were affected in opposite ways by pH, but overall, within the research range, the degree of accelerated change was within an acceptable range. The icIEF acidic peak mainly increased with increasing storage temperature and time. The accelerated test results showed that the degree of change was also within an acceptable range.

[0143] That is, among the 11 prescriptions in the above table, within the range of the prescriptions studied, the anti-ILT2 / 4 antibodies can maintain a relatively high stability.

[0144] Example 11 Prescription Confirmation

[0145] The central point prescription of Example 10, specifically No. 2 / 7 / 11 in Table 10 (three replicate groups), was confirmed by performing freezing, freeze-thaw, 2-8°C, 25°C, and 40°C stability tests.

[0146] The results are shown in Table 12.

[0147] Table 12 Stability test Note: “ / ” means not tested.

[0148] The results in the above table show that anti-ILT2 / 4 has good stability in this preparation and can maintain stability for a long time.

[0149] The antibody sequences used in the present invention are shown in Table 13.

[0150] Table 13 Antibody sequence list

[0151] Although the above describes specific embodiments of the present invention, it should be understood by those skilled in the art that these are merely illustrative and that various changes or modifications may be made to these embodiments without departing from the principles and essence of the present invention. Therefore, the scope of protection of the present invention is defined by the appended claims.

Claims

1. A preparation of an anti-ILT2 / 4 monoclonal antibody, characterized in that: The anti-ILT2 / 4 monoclonal antibody comprises HCDR1, HCDR2 and HCDR3 with amino acid sequences shown in SEQ ID NO: 1, SEQ ID NO: 29 and SEQ ID NO: 3, respectively, and LCDR1, LCDR2 and LCDR3 with amino acid sequences shown in SEQ ID NOs: 4 to 6, respectively.

2. The preparation according to claim 1, wherein The VH of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as set forth in SEQ ID NO: 30, or at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 30; the VL of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as set forth in SEQ ID NO: 12, or at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 12; Preferably, the HC of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as shown in SEQ ID NO: 19, SEQ ID NO: 17, SEQ ID NO: 20, SEQ ID NO: 21 or SEQ ID NO: 22, or an amino acid sequence that is at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 19, SEQ ID NO: 17, SEQ ID NO: 20, SEQ ID NO: 21 or SEQ ID NO: 22; the LC of the anti-ILT2 / 4 monoclonal antibody comprises an amino acid sequence as shown in SEQ ID NO: 18, or an amino acid sequence that is at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 18; More preferably, the formulation further comprises a buffer, a stabilizer and / or a surfactant.

3. The preparation according to claim 1 or 2, wherein The buffer is selected from acetic acid-sodium acetate buffer and PB buffer, preferably acetic acid-sodium acetate buffer; Preferably, the concentration of the buffer solution is not less than 10 mM, selected from 10 mM, 20 mM, 30 mM, 40 mM or 50 mM, preferably 10 mM.

4. The preparation according to any one of claims 1 to 3, wherein The pH range of the preparation is pH 4 to 6, preferably pH 4.5 to 5.

5.

5. The preparation according to any one of claims 1 to 4, wherein The stabilizer is sucrose and / or trehalose, preferably trehalose; Preferably, the concentration of the stabilizer is in the range of 1% to 20%, more preferably 7% to 9%.

6. The preparation according to any one of claims 1 to 5, wherein The surfactant is selected from one or more of polysorbate 20, polysorbate 80 and poloxamer 188, preferably polysorbate 20; Preferably, the concentration of the surfactant is in the range of 0.001% to 0.1%, preferably 0.01% to 0.1%, and more preferably 0.02% to 0.06%.

7. The preparation according to any one of claims 1 to 6, wherein The concentration of the anti-ILT2 / 4 monoclonal antibody is 1-30 mg / mL, preferably 10-30 mg / mL, and more preferably 15-25 mg / mL.

8. The preparation according to any one of claims 1 to 7, wherein The preparation is a liquid preparation.

9. A kit comprising the preparation according to any one of claims 1 to 8, characterized in that The kit further comprises a container; Preferably, the container is a polycarbonate bottle, a polypropylene bottle, an ultra-low density polyethylene bag or an ethylene-vinyl acetate copolymer bag.

10. Use of the preparation according to any one of claims 1 to 8 in the preparation of a product for diagnosing, treating and / or preventing diseases or conditions mediated by ILT4 and / or ILT2; Preferably, the disease or condition is cancer; More preferably, the cancer is ILT4-positive and / or ILT2-positive cancer; More preferably, the cancer is a solid cancer or a blood cancer; The blood cancer is, for example, B-cell leukemia, lymphoma, acute myeloid leukemia, Burkitt's lymphoma, mantle cell lymphoma, acute lymphoblastic leukemia, chronic lymphocytic leukemia, diffuse large B-cell lymphoma, follicular lymphoma or marginal zone lymphoma; The solid cancer is, for example, head and neck squamous cell carcinoma, renal cell carcinoma, ovarian cancer, urothelial carcinoma, diffuse large B-cell carcinoma, non-small cell lung cancer, melanoma, metastatic renal cell carcinoma, metastatic colorectal cancer, small cell lung cancer, metastatic non-small cell lung cancer, breast cancer, lung cancer, head and neck cancer, colorectal cancer, prostate cancer, skin cancer, stomach cancer, intestinal cancer, cervical cancer, uterine cancer, endometrial cancer, bladder cancer, brain cancer, esophageal cancer, liver cancer, kidney cancer, testicular cancer, mesothelioma, malignant glioma, bile duct cancer or pancreatic cancer.

11. A method for immunodetection or determination of ILT4 and / or ILT2, the method comprising contacting the preparation according to any one of claims 1 to 8 with a sample, and determining whether ILT4 and / or ILT2 are present in the sample based on the binding conditions; Preferably, the detection is for non-diagnostic purposes.

12. A method for treating and / or preventing a disease or condition mediated by ILT4 and / or ILT2, the method comprising administering a therapeutically effective amount of the formulation according to any one of claims 1 to 8 to a patient in need thereof.

13. The method according to claim 12, wherein: The disease or condition is cancer; Preferably, the cancer is ILT4-positive and / or ILT2-positive cancer; More preferably, the cancer is a solid cancer or a blood cancer; The blood cancer is, for example, B-cell leukemia, lymphoma, acute myeloid leukemia, Burkitt's lymphoma, mantle cell lymphoma, acute lymphoblastic leukemia, chronic lymphocytic leukemia, diffuse large B-cell lymphoma, follicular lymphoma or marginal zone lymphoma; The solid cancer is, for example, head and neck squamous cell carcinoma, renal cell carcinoma, ovarian cancer, urothelial carcinoma, diffuse large B-cell carcinoma, non-small cell lung cancer, melanoma, metastatic renal cell carcinoma, metastatic colorectal cancer, small cell lung cancer, metastatic non-small cell lung cancer, breast cancer, lung cancer, head and neck cancer, colorectal cancer, prostate cancer, skin cancer, stomach cancer, intestinal cancer, cervical cancer, uterine cancer, endometrial cancer, bladder cancer, brain cancer, esophageal cancer, liver cancer, kidney cancer, testicular cancer, mesothelioma, malignant glioma, bile duct cancer or pancreatic cancer.

14. A combination therapy comprising administering the preparation according to any one of claims 1 to 8 to a patient in need thereof; and a second therapeutic agent; the second therapeutic agent preferably comprises other anti-tumor antibodies or pharmaceutical compositions comprising the other anti-tumor antibodies, and / or one or more of the group consisting of hormone preparations, targeted small molecule preparations, proteasome inhibitors, imaging agents, diagnostic agents, chemotherapeutic agents, oncolytic drugs, cytotoxic agents, cytokines, activators of co-stimulatory molecules, inhibitors of inhibitory molecules and vaccines.

15. The preparation according to any one of claims 1 to 8, for use in treating and / or preventing diseases or conditions mediated by ILT4 and / or ILT2; Preferably, the disease or condition is cancer; More preferably, the cancer is ILT4-positive and / or ILT2-positive cancer; More preferably, the cancer is a solid cancer or a blood cancer; The blood cancer is, for example, B-cell leukemia, lymphoma, acute myeloid leukemia, Burkitt's lymphoma, mantle cell lymphoma, acute lymphoblastic leukemia, chronic lymphocytic leukemia, diffuse large B-cell lymphoma, follicular lymphoma or marginal zone lymphoma; The solid cancer is, for example, head and neck squamous cell carcinoma, renal cell carcinoma, ovarian cancer, urothelial carcinoma, diffuse large B-cell carcinoma, non-small cell lung cancer, melanoma, metastatic renal cell carcinoma, metastatic colorectal cancer, small cell lung cancer, metastatic non-small cell lung cancer, breast cancer, lung cancer, head and neck cancer, colorectal cancer, prostate cancer, skin cancer, stomach cancer, intestinal cancer, cervical cancer, uterine cancer, endometrial cancer, bladder cancer, brain cancer, esophageal cancer, liver cancer, kidney cancer, testicular cancer, mesothelioma, malignant glioma, bile duct cancer or pancreatic cancer.

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