CN118047874A - Bispecific antibody and application of bispecific antibody and T cells in preparation of tumor treatment drugs - Google Patents

Abstract

The invention provides a bispecific antibody and application of the bispecific antibody and T cells thereof in preparation of tumor treatment medicines, wherein the bispecific antibody comprises a first functional region targeting CD3 and a second functional region targeting PD-L1, and the first functional region is of an scFv structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 1 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 2; the second functional region is in a Fab structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 3 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 4. The bispecific antibody adopts an IgG type structure, so that the half life in vivo can be prolonged, and the rejection reaction can be reduced. T activation patterns are optimized, and the activation state of T cells in vivo can be maintained by using a combined stimulation strategy of TGF-beta, IL-25 and IL-33. The bispecific antibody provided by the invention can inhibit the growth of tumor cells, regulate the expression level of immune factors and prolong the production period of animals by being combined with activated T cells.

Description

Bispecific antibody and application of bispecific antibody and T cells in preparation of tumor treatment drugs

Technical field:

The invention belongs to the field of biotechnology research and development, and particularly provides a bispecific antibody and application of the bispecific antibody and T cells in preparation of a tumor treatment drug.

The background technology is as follows:

Tumor immunotherapy is an emerging tumor treatment means, has the advantages of strong anti-tumor capability, lasting curative effect, small side effect and the like, and is widely applied to tumor treatment, especially difficult-to-cure malignant tumors. Early tumor immunotherapy was based on monoclonal antibodies, from the original murine antibody, to humanized antibodies, and then to fully humanized antibodies, and several monoclonal antibody drugs have been approved for the market, such as rituximab, trastuzumab, bevacizumab, cetuximab, and the like, without losing weight-scale drugs, but clinical performance is still poor due to off-target effects, short half-life, difficulty in adapting to factors such as tumor microenvironment, and the like.

For this reason, researchers have developed bispecific antibodies (bispecific antibody, bsAb) that target two target antigens simultaneously, and that reduce the risk of off-target effects well, and are therefore receiving general attention in the industry. By 2023, 14 BsAb drugs were approved worldwide, 9 of which were used to treat hematological malignancies, including blinatumomab, mosunetuzumab, teclistamab, epcoritamab and glofitamab, etc. (see Antonio Tapia Galisteo,et al.Bi-and trispecific immune cell engagers for immunotherapy ofhematological malignancies.JHematol Oncol.2023;16:83).

Among the numerous BsAbs, CD3 bispecific antibodies (CD 3-BsAbs) have become increasingly important as an emerging therapeutic modality in the field of cancer immunotherapy, CD3-BsAbs act by simultaneously binding to tumor-associated antigens (TAAs) expressed on tumor cells and CD3 on T cells, CD3-BsAbs cross-link these two cell types to form immune synapses, similar to the natural T Cell Receptor (TCR)/peptide-Major Histocompatibility Complex (MHC) complex (see Xu H,et al.Retargeting T cells to GD2 pentasaccharide on human tumors using Bispecific humanized antibody.Cancer Immunol.Res.2015;3:266–277), this synapse results in T cell activation, secretion of inflammatory cytokines and cytolytic molecules, thereby killing tumor cells in the process, the advantage of CD3-BsAbs is that any T cell can act as an effector cell, no matter what the TCR specificity is, TCR signaling does not require the participation of the antigen binding domain of the TCR for these BsAbs, but rather by activating CD3 signaling (see Wu Z,et al.T cell engaging bispecific antibody(T-BsAb):From technology to therapeutics.Pharmacol.Ther.2018;182:161–175), so CD3-BsAbs can utilize all available T cells, not limited to tumor-specific T cells).

Currently, CD3-BsAb has shown great potential in the treatment of hematological cancers, FDA approved blinatumomab (CD 3xCD 19) has been successfully used in clinical treatment of some B cell malignancies, blinatumomab is a CD3xCD19 BsAb without Fc tail, more than 40% of adult patients treated with blinatumomab show complete or partial remission compared to standard chemotherapy, the median overall survival has been prolonged for several months (see Franquiz M.J,et al.Blinatumomab for the Treatment of Adult B-Cell Acute Lymphoblastic Leukemia:Toward a New Era of Targeted Immunotherapy.Biologics.2020;14:23–34). in addition to blinatumomab, many other CD 3-BsAbs are currently undergoing clinical trials against mature B cell markers such as CD19, CD20, CD38 and B Cell Maturation Antigen (BCMA) as well as bone marrow markers such as CD33 and CD 123. However, in contrast to the success of CD3-BsAb in hematological malignancies, the effect of these antibodies in solid tumors is still quite limited due to the following obstacles in the treatment of solid tumors:

The first is off-target toxicity, in the case of hematological cancers, temporary depletion of B cells or bone marrow subpopulations is reversible so long as they are not targeted to hematopoietic stem cells, thereby replenishing blood. However, solid tumor targets are often expressed on tissues of healthy organs, which can lead to immunopathological phenomena, even organ failure and can be fatal, as demonstrated by preclinical mouse studies using EGFR-targeted CD3-BsAb (see Lutterbuese R,et al.T cell-engaging BiTE antibodies specific for EGFR potently eliminate KRAS-and BRAF-mutated colorectal cancer cells.Proc.Natl.Acad.Sci.USA.2010;107:12605–12610).

Second is insufficient availability of effector cells in the immune tumor microenvironment (immunosuppressive tumor microenvironment, TME). For hematological malignancies, cancer cells in the blood are surrounded by T cells, so that CD3-BsAb is extracted from the endless effector cell pool, whereas solid tumors require T cell infiltration to exert therapeutic efficacy. In this case, "inflamed" tumors may appear, which are infiltrated by immune cells and often respond to immune checkpoint therapy (see Chen D.S, et al elements ofcancer immunity AND THE CANCER-immune set point. Nature.2017;541: 321-330); or "immune desert" tumors, immunocytopenia or lack of immunogenicity, resulting in less priming of tumor-specific T cells homing to the tumor (see anitis E,et al.Mechanisms regulating T-cell infiltration and activity in solidtumors.Ann.Oncol.2017;28:xii18–xii32).

Thirdly, the quality of infiltrating T cells is not high. TME inhibits immune cells, including cancer-associated fibroblasts (CAF), bone Marrow Derived Suppressor Cells (MDSC), and regulatory T cells (T regs), producing TGF-beta, IL-10, indoleamine 2, 3-dioxygenase and arginase and like regulatory factors that block T cell metabolism and activation (see Tormoen G.W,et al.Role of the immunosuppressive microenvironment in immunotherapy.Adv.Radiat.Oncol.2018;3:520–526).. Furthermore, effector T cells exhibit "depletion" characteristics due to chronic antigen stimulation, which is demonstrated in the expression of inhibitory immune checkpoints (see Grywalska E,et al.Immune-checkpoint inhibitors for combating T-cell dysfunction in cancer.Onco Targets Ther.2018;11:6505–6524).

Based on the research, the invention designs and obtains the bispecific antibody targeting CD3 and PD-L1, and uses the bispecific antibody and activated T cells for treating solid tumors, thereby improving the anti-tumor effect and providing a new way for developing novel tumor immunotherapy.

Disclosure of Invention

The first aspect of the invention provides a bispecific antibody, which is characterized by comprising a first functional region targeting CD3 and a second functional region targeting PD-L1, wherein the first functional region is of an scFv structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO.1 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 2; the second functional region is in a Fab structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 3 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 4.

The target selection in CD3 BsAb is very important, directly related to tumor specificity and treatment safety, and Her2, EGFR and EpCAM have been reported to be qualified candidate targets. Immune checkpoint antibodies have been a major breakthrough in the field of tumor immunotherapy in recent years, and have achieved a surprising clinical effect by identifying a number of inhibitory checkpoints in addition to PD-1/PD-L1 and performing therapeutic validation, including CTLA-4, LAG-3, TIM-3 and TIGIT 9. There is evidence that LAG-3 and PD-L1 are promising therapeutic targets. (see Reader CS,et al.Abstract 2874:The tetravalent structure of FS118,a bispecific antibody targeting LAG-3and PD-L1,is required for its novel mechanism of LAG-3shedding.Cancer Res.2022;82:2874). thus, selection of CD3 and PD-L1 in the present application constructs bispecific antibodies.

Further, the first functional region is connected to the N-terminal end of the second functional region.

Further, the heavy chain of the bispecific antibody comprises a domain as shown in scFv-VH-CH-hinge-Fc in sequence from the N-terminal to the C-terminal, wherein scFv targets CD3, VH is a heavy chain variable region targeting PD-L1, and CH is a heavy chain constant region; the light chain of the bispecific antibody comprises a domain from the N-terminus to the C-terminus as shown in VL-CL, which is the heavy chain variable region targeting PD-L1, and CL is the light chain constant region.

Further, the Fc region is a human IgG Fc region.

Further, the Fc region is a human IgG4 Fc region.

The structure of BsAb varies widely and can be broadly divided into IgG-like BsAb and non-IgG-like BsAb. IgG-like bsabs are Y-shaped, with Fc fragments having a variety of advantages in production and clinical treatment. The complete antibody structure not only facilitates the purification process, but also increases the stability of the product, and IgG-like bsabs have a longer half-life in vivo than non-IgG-like bsabs. More importantly, the Fc fragment mediates innate and adaptive immune responses, playing a vital role in antitumor activity; however, because of the linkage association, a more stringent quality control scheme is required (see Ma J,et al.Bispecific antibodies:from research to clinical application.Front Immunol.2021;12:626616). non-IgG-like BsAb, which does not contain an Fc fragment, which has unique advantages, such BsAb is simple in structure and can be easily produced in eukaryotic and prokaryotic expression systems, in addition, the number of antigen binding sites is flexible, the valences of the two targets can be tailored (see Johnson S,et al.Effector cell recruitment with novel Fv-based dual-affinity re-targeting protein leads to potent tumor cytolysis and in vivo B-cell depletion.J Mol Biol.2010;399:436–49). in the present invention, igG-like BsAb structure has been selected in order to reduce graft-versus-host reactions of antibodies during use, improve safety and efficacy of treatment).

In a second aspect, the invention provides an anti-tumour composition comprising said bispecific antibody and T cells.

Further, the preparation method of the T cell comprises the following steps: extracting mononuclear cells from peripheral blood, and separating by CD3/CD28 antibody magnetic beads to obtain T cells; the T cells were subjected to TGF-beta, IL-25 and IL-33 incubation.

Further, the preparation method of the T cell comprises the following steps: the incubation was performed at 37℃for 24h using TGF-. Beta.with a final concentration of 1.5ng/mL, IL-25 at 50ng/mL and IL-33 at 80 ng/mL.

As previously mentioned, a major obstacle in the treatment of solid tumors is the lack of sufficient, fully activated T cells in the tumor microenvironment, and thus the present invention contemplates the supplementation of T cells by in vitro injection, thereby acting synergistically with bispecific antibodies targeting CD3 and PD-L1, and in order to increase T cell activity, it is also activated in vitro using cytokines. Activation and differentiation of T cells is associated with a variety of cytokines including interleukin family, interferon family, tumor necrosis factor family, transforming growth factor family, and the like. The mechanism of action of TGF-beta is complex, which is generally considered to be unfavorable for T cell activation and anti-tumor treatment, but the TGF-beta can generate synergistic action with certain cytokines, so that the T cell proliferation and differentiation are promoted, for example, the TGF-beta can be synergistic with IL-6, th17 cell differentiation which expresses retinoic acid related orphan receptor gamma (RORgamma) and generates high level IL-17 is favored (see Park H,et al.Adistinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17.Nat.Immunol.2005,6:1133–41); to be synergistic with IL-4, th9 cell differentiation (Veldhoen M,et al.TGFβin the context of an inflammatory cytokine milieu supports de novo differentiation of IL-17-producing T cells.Immunity.2006,24:179–89). which induces IL-9 secretion is creatively combined with IL-25 and IL-33 in the invention, and the dosage of the TGF-beta is regulated, so that the T cell activation can be well promoted, and stronger anti-tumor effect is generated.

In a third aspect, the present invention provides an application of the bispecific antibody and/or the antitumor composition in preparing antitumor drugs.

Further, the tumors are lung cancer and liver cancer.

Advantageous effects

The invention provides a bispecific antibody and application of the bispecific antibody and T cells thereof in preparation of a tumor treatment drug, and the bispecific antibody and the T cells have the following specific beneficial effects:

(1) Screening to obtain antigen binding domains targeting CD3 and PD-L1 capable of binding highly specifically to the antigen of interest;

(2) The bispecific antibody targeting CD3 and PD-L1 simultaneously is constructed, and the antibody adopts an IgG type structure, so that the half life in vivo can be prolonged, and the rejection reaction can be reduced;

(3) Optimizing the T cell activation mode, and using a combined stimulation strategy of TGF-beta, IL-25 and IL-33 to maintain the activation state of the T cells in vivo;

(4) The bispecific antibody provided by the invention can inhibit the growth of tumor cells, regulate the expression level of immune factors and prolong the production period of animals by being combined with activated T cells.

Drawings

Fig. 1: schematic diagram of bispecific antibody structure;

Fig. 2: in vitro tumor inhibition experiments;

Fig. 3: animal survival curves;

fig. 4: levels of TNF- α expression in serum;

fig. 5: IL-2 expression level in serum;

Fig. 6: IFN-gamma expression levels in serum.

Detailed Description

The experimental methods described in the following examples, unless otherwise specified, are all conventional; the reagent biological material and the detection kit can be obtained from commercial sources unless otherwise specified.

Example 1 design and preparation of bispecific antibodies

The bispecific antibody with the Fc fragment is designed in the invention, so that the in vivo half-life of the bispecific antibody is obviously prolonged due to the addition of the Fc fragment, and the immune rejection reaction can be reduced by using the human IgG4 Fc region, so that serious toxic and side effects are avoided.

The monoclonal antibody targeting PD-L has been obtained in early use, and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 3 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 4, and the affinity equilibrium constant KD of the monoclonal antibody targeting PD-L is 5.51E-10M. The scFv structure of the target human CD3 is then screened and obtained by phage display technology, wherein the scFv structure comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 1 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 2, and the affinity equilibrium constant KD of the scFv structure with the target antigen is identified to be 3.07E-10M. As shown in fig. 1, based on the above, the present invention uses the monoclonal antibody of PD-Ll as the second functional region, which has a complete antibody structure, and the Fc region is a human IgG4 Fc region, which can prolong the half-life in vivo and reduce the immune rejection reaction caused by heterologous proteins; the N-terminus of the antibody is linked to scFv targeting human CD3 to form a first functional region, thereby forming a bispecific antibody. The method comprises the following specific steps:

The novel peptide comprises a first functional region targeting CD3 and a second functional region targeting PD-L1, wherein the first functional region is of an scFv structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 1 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 2; the second functional region is in a Fab structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 3 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 4. The nucleotide sequence encoding the anti-CD 3 scFv was ligated to the N-terminus of the heavy chain nucleotide of the anti-PD-L1 antibody, while the light chain portion still employed the light chain nucleotide of the anti-PD-L1 antibody. The nucleotide is transferred into pGEX-4T-1 expression vector, the recombinant plasmid is named pGEX-CD3-PD-L1, and the plasmid is transfected into E.coli DH5 alpha infected cell. Coating the plasmid into LB solid culture medium containing ampicillin resistance, screening positive clones, extracting plasmids for preliminary enzyme digestion identification, and sequencing the positive clones to identify that the nucleotide sequence is correct.

1 Mu L of recombinant plasmid pGEX-CD3-PD-L1 is transformed into competent cells of escherichia coli BL21, positive clones are selected and inoculated into 100mL of LB liquid medium, the culture is carried out at 37 ℃ under shaking at 200r/min until OD ₆₀₀ is about 0.8, and IPTG with the final concentration of 0.2mmol/L is added for induction culture, and the culture is carried out at 20 ℃ under shaking at 200r/min for 24 hours. After the induction expression is finished, the cells are collected by centrifugation at 12000r/min for 5min, and after ultrasonic crushing, the antibodies are separated and purified by utilizing a GST chromatographic column. SDS-PAGE and Coomassie brilliant blue staining are used for identifying the purity of the antibody, and the detection purity reaches more than 90 percent, thereby meeting the experimental requirements.

Example 2 extraction and activation of T cells

Taking 12mL of peripheral venous blood of healthy volunteers, putting the peripheral venous blood into a heparin anticoagulation tube, adding 6mL of lymphocyte separation liquid into a 15mL centrifuge tube, then adding 6mL of blood by adherence, and centrifuging at 1000rpm for 30min at 4 ℃; sucking the white membrane layer (PERIPHERAL BLOOD MONONUCLEAR CELLS, PBMCs) into a 50mL centrifuge tube, adding X-VIVO culture medium for cleaning, centrifuging at 500rpm for 10min, and discarding supernatant; 15mL of erythrocyte lysis buffer is added, the mixture is incubated for 10min at room temperature, centrifuged for 10min at 500rpm, and the supernatant is discarded; adding the X-VIVO culture medium for washing once, and activating and screening by using the X-VIVO complete culture medium containing anti-human CD3/CD28 antibody magnetic beads to obtain cells for subculture, and carrying out 3-5 passages altogether, and marking the cells as T cells.

Taking T cells in logarithmic growth phase, discarding the original culture medium, washing 3 times by using sterile PBS, adding TGF-beta with the final concentration of 1.5ng/mL, IL-25 with the final concentration of 50ng/mL and IL-33 with the final concentration of 80ng/mL, and incubating at 37 ℃ for 24 hours; the medium was discarded, the cells were digested with 0.5% pancreatin, washed 3 times with sterile PBS, and centrifuged AT 500rpm for 10min to collect the cells, which were designated AT cells.

Example 3 bispecific antibodies and T cell in vitro anti-tumor experiments

In the invention, human liver cancer cells HepG2 and human non-small cell lung cancer cells A549 are taken as objects, and the tumor inhibition capacity of T cells is examined. Resuscitating and culturing tumor cells, culturing to logarithmic growth phase, inoculating T cells and tumor cells into 96-well plates according to an effective target ratio of 5:1, and grouping the cells, wherein each group comprises 3 compound wells, namely: bsAb+T cells, T cells extracted by a traditional method and 100ng/mL of BsAb targeting CD3 and PD-L1 provided by the invention are added; bsAb+AT cells, activated T cells and 100ng/mL of the BsAb targeting CD3 and PD-L1 provided in the invention were added, tumor cells without T cells and antibodies were used as a negative control group, and medium-only wells were used as a blank control group. After 24h of co-cultivation, 10mL of MTT solution was added to each well, incubated at 37℃for 4h, and absorbance at 450nm (OD value) was measured with a microplate reader, and each set of experiments was repeated 3 times. The inhibition ratio was calculated as follows, inhibition ratio= (negative control OD value-experimental OD value)/(negative control OD value-blank OD value) ×100%.

As shown in figure 2, the bispecific antibody provided by the invention can effectively activate T cells to play an anti-tumor role, and the activated T cells can generate a stronger tumor killing effect, so that the tumor killing capability of the T cells is greatly improved after the dual activation of the bispecific antibody and the combined cytokine.

Example 4 bispecific antibodies and T cell in vivo anti-tumor experiments

4.1 Animal model preparation and treatment

Resuscitates and cultures A549 cells, cultures to logarithmic phase, uses 0.25% pancreatin digestion, adjusts the cell concentration to about 2X 10 ⁶/mL, uses C57BL/6 mice as study object, and each mouse right armpit subcutaneous inoculates cell suspension 0.2mL, observes the tumor growth condition. Selecting 30 mice with successful modeling, randomly dividing into 3 groups, namely BsAb+AT groups, injecting 50mg/kg targeted CD3 and PD-L1 bispecific antibody every 3 days, and injecting 1X 10 ⁶ activated T cells every week for 4 weeks; for BsAb+T group, 50mg/kg of targeted CD3 and PD-L1 bispecific antibody was injected every 3 days, and 1X 10 ⁶ T cells extracted by the conventional method were injected every week for 4 weeks; control group was injected with an equal amount of physiological saline and administered for 4 weeks. Another 10 normal mice were taken as normal control group.

4.2 Animal survival observations

Animals were observed daily for survival and recorded and survival curves were plotted. As shown in fig. 3, the BsAb and T cell therapy provided by the present invention can significantly prolong the survival time of experimental animals, and activated T cells seem to obtain better therapeutic effects, which indicates that the T cells in vivo are hardly satisfied with the need of anti-tumor effect due to the influence of tumor microenvironment in experimental animals, and the anti-tumor effect is significantly enhanced by external T cell supplementation, especially activated T cells.

4.3 Detection of inflammatory factors

The orbit is taken to blood, placed in an anticoagulation tube containing heparin, and the supernatant is collected by centrifugation to collect plasma. The serum cytokines TNF-alpha, IL-2, IFN-gamma content of tumor-bearing mice were detected using an ELISA kit (available from Calvin Biotechnology Co., ltd.) according to the kit instructions.

The results are shown in figures 4-6, the expression levels of TNF-alpha, IL-2 and IFN-gamma in the tumor model group are greatly reduced compared with the normal group, and after BsAb and T cell treatment, the expression levels of various immune factors are recovered, which shows that the method can effectively activate the immune system in vivo, stimulate the secretion of immune factors, and mobilize the autoimmune mechanism of the organism against tumor cells. Considering that the autoimmune system of the organism is fully mobilized by BsAb, the advantage of convenient secretion of immune factors is not obvious by using activated T cells, and the advantages of TNF-alpha and IL-2 are not obviously different from those of BsAb+T groups, but are obviously increased in IFN-gamma expression, thus showing stronger immunoregulation effect.

Claims (10)

1. A bispecific antibody, which is characterized by comprising a first functional region targeting CD3 and a second functional region targeting PD-L1, wherein the first functional region is of an scFv structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 1 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 2; the second functional region is in a Fab structure and comprises a heavy chain variable region with an amino acid sequence shown as SEQ ID NO. 3 and a light chain variable region with an amino acid sequence shown as SEQ ID NO. 4.

2. The bispecific antibody of claim 1, wherein the first functional region is linked to the N-terminus of the second functional region.

3. The bispecific antibody of claim 2, wherein the heavy chain of the bispecific antibody comprises in order from N-terminus to C-terminus a domain as shown in scFv-VH-CH-hinge-Fc, wherein scFv targets CD3, VH is the heavy chain variable region targeting PD-L1, CH is the heavy chain constant region; the light chain of the bispecific antibody comprises a domain from the N-terminus to the C-terminus as shown in VL-CL, which is the heavy chain variable region targeting PD-L1, and CL is the light chain constant region.

4. The bispecific antibody of claim 3, wherein the Fc region is a human IgG Fc region.

5. The bispecific antibody of claim 4, wherein the Fc region is a human IgG4 Fc region.

6. An anti-tumor composition comprising the bispecific antibody of any one of claims 1-5 and a T cell.

7. The anti-tumor composition according to claim 6, wherein the T cell preparation method comprises: extracting mononuclear cells from peripheral blood, and separating by CD3/CD28 antibody magnetic beads to obtain T cells; the T cells were subjected to TGF-beta, IL-25 and IL-33 incubation.

8. The anti-tumor composition according to claim 7, wherein the T cell preparation method comprises: the incubation was performed at 37℃for 24h using TGF-. Beta.with a final concentration of 1.5ng/mL, IL-25 at 50ng/mL and IL-33 at 80 ng/mL.

9. Use of a bispecific antibody according to any one of claims 1-5 and/or an anti-tumor composition according to any one of claims 1-5 for the preparation of an anti-tumor medicament.

10. The use according to claim 9, wherein the tumors are lung cancer and liver cancer.