Treatment or prevention of medical conditions associated with herpes virus infection

Abstract

The present invention provides a method of treating or preventing a medical condition associated with herpes virus infection comprising administering a therapeutically effective amount of an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface and an HDAC inhibitor. Kits and pharmaceutical products comprising an HDAC inhibitor and CAR-containing immune cells are also provided. In addition, an antibody that binds to gp350 of Epstein-Barr virus present on the cell surface of a target cell is provided.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit of priority to European Patent Application No. 22180615.1 filed on June 23, 2022, the contents of which are hereby incorporated by reference in their entirety for all purposes. Technical Field

[0003] The present invention provides a method for treating or preventing a medical condition associated with herpes virus infection, the method comprising administering a therapeutically effective amount of immune cells and HDAC inhibitors, the immune cells comprising a chimeric antigen receptor (CAR) on their cell surface. A kit and a pharmaceutical product comprising an HDAC inhibitor and immune cells containing CAR are also provided. In addition, an antibody is provided that binds to the gp350 of Epstein-Barr virus present on the cell surface of a target cell. Background Art

[0004] The characteristic of herpes virus infection is usually latent infection, which can eventually lead to the development of medical conditions such as cancer. Epstein-Barr virus (EBV) is a human herpes virus related to the development of various human tumors. These tumors include B-, T- and NK-cell lymphomas, nasopharyngeal carcinomas and gastric cancers. Although the genome of EBV encodes more than 80 viral proteins, among these proteins, only a few are usually expressed in tumor cells infected by EBV. Therefore, using immunotherapy means, such as EBV-specific CAR-T cells are difficult to target and eliminate such tumor cells.

[0005] Glycoprotein 350 / 220 (gp350) is a viral protein expressed mainly during the proliferative lytic cycle. However, independently of this, gp350 has also been found on some latent EBV-infected tumor cells, i.e., cells that do not produce detectable amounts of virus and only produce a small amount of viral protein. However, gp350 is usually present at low expression levels on these cells. Gp350 can be targeted, for example, by gp350-specific CAR-T cell targeting, and for example in WO 2019 / 201995 A1 and Slabik et al., (2020), Molecular Therapy: Oncolytics, 18: 504-524 are described.

[0006] The expression of gp350 on EBV infected tumor cells can be increased, for example, by inducing the lytic cycle of EBV, which can be achieved in several ways. For example, transfection / transduction of a vector encoding the EBV protein BZLF1 (Zta) or treatment with a histone deacetylase (HDAC) inhibitor such as valproic acid, vorinostat or TPA has been described (Ghosh et al., (2012), Blood, 119: 1008-1017).

[0007] However, the CAR-T cells with specificity for proteins expressed on cells with latent herpes simplex virus infection and the expression of herpes simplex virus-specific proteins in the cells cannot achieve effective treatment individually. Therefore, there is still a need to further improve the therapy of the medical conditions caused by latent herpes simplex virus infection. The fundamental technical problem of the present application is to meet this demand. Summary of the invention

[0008] The above technical problems are solved by the subject matter defined in the claims. The inventors surprisingly found that treating cells with HDAC inhibitors increases the surface expression of herpes virus antigens, which can then be targeted by immune cells such as CAR-T cells, thereby acting synergistically in the treatment of medical conditions associated with herpes virus infection.

[0009] Therefore, the present invention provides an HDAC inhibitor for treating or preventing a medical condition associated with herpes virus infection, the method comprising further administering an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface, wherein the CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or to a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell infected by the herpes virus.

[0010] In a related embodiment, the present invention relates to an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface, which is used to treat or prevent a medical condition associated with a herpes virus infection, the method comprising further administering an HDAC inhibitor, wherein the CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or to a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell infected by the herpes virus.

[0011] Human cells may be latently infected by the herpes virus. Human cells may be acutely infected by the herpes virus. The infection of the herpes virus can be continuous. Human cells can be persistently infected by the herpes virus.

[0012] HDAC inhibitors can induce the lytic cycle of herpesviruses. HDAC inhibitors can induce the expression of herpesvirus antigens.

[0013] The HDAC inhibitor can be selected from butyric acid, valproic acid, MS275 (Entinostat), apicidin, largazole, oxamflatin, scriptaid, suberoylanilide hydroxamic acid (SAHA, Vorinostat), LBH589 (Panobinostat), 12-O-tetradecanoylphorbol-13-acetate (TPA) and PXD101 (Belinostat), preferably SAHA, TPA or valproic acid, more preferably SAHA.

[0014] The medical condition associated with herpes virus infection can be a herpes virus-associated cancer, a chronic active herpes virus infection, a reactivated herpes virus infection, or a primary herpes virus infection.

[0015] A medical condition associated with a herpes virus infection may be a herpes virus-associated cancer, wherein herpes virus antigens are present on the surface of cancer cells.

[0016] The herpes virus may be EBV.

[0017] The medical condition may be an EBV-associated cancer selected from a lymphoproliferative disorder (LPD), such as a B-cell lymphoma, including Burkitt's lymphoma (BL), Hodgkin's lymphoma (HL), diffuse large B-cell lymphoma (DLBCL), T-cell lymphoma, or a post-transplantation lymphoproliferative disorder (PTLD), or an epithelial cell carcinoma (nasopharyngeal carcinoma, lung carcinoma, breast carcinoma), a lymphoepithelioma, a lymphoid stromal carcinoma (GCLS, such as gastric cancer) or a glioma.

[0018] The medical condition associated with EBV infection may be chronic active EBV infection (CAEBV) or primary EBV infection (eg infectious mononucleosis).

[0019] The herpes virus antigen may be involved in the binding of the virus to a receptor on a human target cell. The herpes virus antigen may be an Epstein-Barr virus antigen (EBV antigen).

[0020] EBV antigen can be present in the cell surface infected by EBV, preferably present in the cancer cell infected by EBV, the B cell infected by EBV or the epithelial cell surface infected by EBV. EBV antigen can be the protein (such as gB, gL or gH) of EBV virion envelope protein or EBV envelope complex. Preferably, herpes virus antigen is EBV glycoprotein 350 / 220 (gp350 / 220).

[0021] The immune cells may be selected from T lymphocytes, NK cells, macrophages and dendritic cells, wherein the T lymphocytes are preferably cytotoxic T lymphocytes or T helper cells, more preferably cytotoxic T lymphocytes.

[0022] The CAR may comprise or consist of any one of SEQ ID NO: 2, 12, 22 or 32.

[0023] HDAC inhibitors can be administered to patients before or simultaneously with immune cell administration. HDAC inhibitors can be administered to patients after immune cell administration. HDAC inhibitors can be administered to patients simultaneously with immune cell administration.

[0024] The present invention also relates to a method for treating or preventing a medical condition associated with a herpes virus infection, the method comprising administering a therapeutically effective amount of an immune cell as defined herein and an HDAC inhibitor as defined herein, the immune cell comprising a chimeric antigen receptor (CAR) as defined herein on its cell surface.

[0025] The present invention further relates to a kit of parts comprising an immune cell as defined herein, comprising a chimeric antigen receptor (CAR) as defined herein on its cell surface, and an HDAC inhibitor as defined herein.

[0026] The present invention also relates to a pharmaceutical product comprising an immune cell as defined herein, comprising a chimeric antigen receptor (CAR) as defined herein on its cell surface, and an HDAC inhibitor as defined herein.

[0027] The present invention also relates to the kit or the pharmaceutical product, which is used for combined use in a method for treating or preventing a medical condition associated with herpes simplex virus infection. The combined use may include administering an HDAC inhibitor to a patient before the immune cell is administered, administering an HDAC inhibitor and immune cells to the patient simultaneously, or administering immune cells to the patient after the HDAC inhibitor is administered.

[0028] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0029] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:3, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:4; and / or

[0030] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0031] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0032] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having the amino acid sequence shown in SEQ ID NO:5, a heavy chain CDR2 having the amino acid sequence shown in SEQ ID NO:6, and a heavy chain CDR3 having the amino acid sequence shown in SEQ ID NO:7, and

[0033] A light chain variable region comprising a light chain CDR1 having the amino acid sequence shown in SEQ ID NO:8, a light chain CDR2 having the amino acid sequence shown in SEQ ID NO:9, and a light chain CDR3 having the amino acid sequence shown in SEQ ID NO:10; and / or

[0034] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0035] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0036] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 13, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 14; and / or

[0037] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0038] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0039] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having the amino acid sequence shown in SEQ ID NO: 15, a heavy chain CDR2 having the amino acid sequence shown in SEQ ID NO: 16, and a heavy chain CDR3 having the amino acid sequence shown in SEQ ID NO: 17, and

[0040] A light chain variable region comprising a light chain CDR1 having the amino acid sequence shown in SEQ ID NO: 18, a light chain CDR2 having the amino acid sequence shown in SEQ ID NO: 19, and a light chain CDR3 having the amino acid sequence shown in SEQ ID NO: 20; and / or

[0041] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0042] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0043] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 23, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 24; and / or

[0044] (b) binds to the same epitope in gp350 of the EBV protein as the epitope to which the antibody of (a) binds. The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0045] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having the amino acid sequence shown in SEQ ID NO: 25, a heavy chain CDR2 having the amino acid sequence shown in SEQ ID NO: 26, and a heavy chain CDR3 having the amino acid sequence shown in SEQ ID NO: 27, and

[0046] A light chain variable region comprising a light chain CDR1 having the amino acid sequence shown in SEQ ID NO: 28, a light chain CDR2 having the amino acid sequence shown in SEQ ID NO: 29, and a light chain CDR3 having the amino acid sequence shown in SEQ ID NO: 30; and / or

[0047] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0048] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0049] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 33, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 34; and / or

[0050] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0051] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0052] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having the amino acid sequence shown in SEQ ID NO:35, a heavy chain CDR2 having the amino acid sequence shown in SEQ ID NO:36, and a heavy chain CDR3 having the amino acid sequence shown in SEQ ID NO:37, and

[0053] A light chain variable region comprising a light chain CDR1 having the amino acid sequence shown in SEQ ID NO: 38, a light chain CDR2 having the amino acid sequence shown in SEQ ID NO: 39, and a light chain CDR3 having the amino acid sequence shown in SEQ ID NO: 40; and / or

[0054] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0055] The present invention also relates to a CAR comprising any heavy chain variable region and light chain variable region as defined herein or comprising any heavy chain CDR1, CDR2 and CDR3; and light chain CDR1, CDR2, CDR3 as defined herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0056] Figure 1 Shown are the structures of various HDAC inhibitors suitable for use in the present invention.

[0057] Figure 2 SAHA induces the viral lytic cycle in BL41 cells. Cells were treated with SAHA at the indicated concentrations for 3 days, and RNA levels of the EBV immediate early gene BZLF1 and the cellular housekeeping gene GAPDH were determined by RT-PCR, and PCR products were quantified using Image-J.

[0058] Figure 3Induction of gp350 surface expression on BL41 cells is shown, with treatment with vorinostat (suberoylanilide hydroxamic acid; SAHA) (15 μm), valproic acid (VPA) (15 mM) and 12-O-tetradecanoylphorbol-13-acetate TPA (20 ng / ml) as positive controls. Cells were treated for 3 days and gp350 surface expression was measured by flow cytometry with Alexa488-labeled gp350 antibodies. Results are shown as mean fluorescence.

[0059] Figure 4 It is shown that the treatment of EBV-positive BL41 cells increases the IFN-γ released by gp350-specific CAR-T cells.NPC43 cells were treated with SAHA at the indicated concentrations for 3 days. Then, two different types of gp350-specific CAR-T cells (12C10 LH CAR; 7A1 CAR) or mock T cells were added for another 24 hours. Finally, the concentration of IFN-γ in the supernatant was determined by ELISA.

[0060] Figure 5 Treatment of BL41 with SAHA at the indicated concentrations increases the number of bound gp350-specific antibodies over time. Calculations were performed using the QuiFi kit (Dianova) according to the manufacturer's instructions.

[0061] Figure 6 The nucleotide (SEQ ID NO: 1) and amino acid sequences (SEQ ID NO: 2) of the gp350-specific CAR construct 7D11 of the present invention are shown. DETAILED DESCRIPTION

[0062] Herpesviruses are a large family of DNA viruses that cause infections and some diseases (medical conditions associated with herpesvirus infection) in animals, including humans. Herpesviruses can cause both latent and lytic infections. There are nine types of herpesviruses known to primarily infect humans, of which at least five – herpes simplex viruses 1 and 2 (HSV-1 and HSV-2, also called HHV-1 and HHV-2; both of which can cause oral and genital herpes), varicella-zoster virus (or HHV-3; the cause of chickenpox and shingles), Epstein-Barr virus (EBV or HHV-4; which is involved in several diseases, including infectious mononucleosis and some cancers), and human cytomegalovirus (HCMV or HHV-5) – are widespread in humans. More than 90% of adults have been infected with at least one of these, and latent forms of the virus remain in nearly all people who have been infected. Other human herpesviruses are human herpesvirus 6A and 6B (HHV-6A and HHV-6B), human herpesvirus 7 (HHV-7), and Kaposi's sarcoma-associated herpesvirus (KSHV, also known as HHV-8).

[0063] All herpes viruses are nuclear replication, that is, viral DNA is transcribed into mRNA, and viral DNA is replicated in the nucleus of infected cells. When viral particles contact cells with specific types of receptor molecules on the cell surface, infection begins. After the viral envelope glycoprotein binds to the cell membrane receptor, the virion is internalized and disintegrated, so that the viral DNA migrates and enters the nucleus. In the nucleus, replication of viral DNA and transcription of viral genes occur. During acute infection, the infected cell transcribes and cleaves viral genes. In some host cells, a small amount of viral genes called latent associated transcripts (LAT) are accumulated instead. In this way, the virus can exist indefinitely in cells (and therefore in the host) for a long time. Although primary infection is usually accompanied by acute infection and self-limiting clinical disease periods, long-term latency is usually asymptomatic.

[0064] Chromatin dynamics regulates the transcriptional capacity of the entire herpesvirus genome. When the virus enters the cell, the cellular immune response is to protect the cell. The cell achieves protection by wrapping the viral DNA around histones and compressing it into repressive chromatin, which causes the viral DNA to limit the transcription of viral genes and prevent the virus from synthesizing from scratch to eventually reach a state of dormancy or latent infection. If the cell fails to do so and the viral DNA is only partially converted to repressive chromatin, the viral DNA is still available, supporting the transcription of viral genes, the replication of viral DNA, and the morphogenesis and export of newly produced viruses. Herpes viruses can also reactivate from latent infection, enter the lytic phase and begin to produce new viruses. Lytic phase reactivation is usually initiated by external triggers, resulting in the expression of key transcription factors of viral origin in latently infected cells. Key viral transcription factors can act as molecular switches that direct cellular machinery to activate repressive viral chromatin. Reactivation can be incomplete to allow transient expression of viral antigens, but reactivation can also lead to a full outbreak of lytic infection, including the expression of large amounts of viral antigens and de novo synthesis of viruses and the export of infectious virions. Reactivation of latent virus is associated with many diseases (e.g., herpes zoster, pityriasis rosea, cold sores, or genital herpes). After activation, viral genes are transcribed from LAT to multiple lytic genes; these lead to enhanced replication and virus production. Clinically, lytic activation is often accompanied by the onset of nonspecific symptoms (such as low-grade fever, headache, sore throat, malaise, and rash), as well as clinical signs (e.g., swollen or tender lymph nodes) and immunological (e.g., decreased levels of natural killer cells) changes.

[0065] The cell surface expression of herpes virus antigens is also increased by, for example, HDAC inhibitors to activate the lytic cycle in cells infected by herpes virus (latent or long-term), and the herpes virus antigens can be targeted by immune cells comprising CAR. The inventors were surprised to find that treating cells with HDAC inhibitors increased the surface expression of herpes virus antigens, and the herpes virus antigens can then be targeted by immune cells (such as CAR-T cells) that specifically target herpes virus antigens, thereby playing a role in the treatment of medical conditions related to herpes virus infection in a synergistic manner.

[0066] Therefore, the present invention provides an HDAC inhibitor for treating or preventing a medical condition associated with herpes virus infection, the method comprising further administering an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface, wherein the CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or to a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell infected by the herpes virus.

[0067] In a related embodiment, the present invention relates to an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface, which is used to treat or prevent a medical condition associated with a herpes virus infection, the method comprising further administering an HDAC inhibitor, wherein the CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or to a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell infected by the herpes virus.

[0068] The present invention also relates to a method for treating or preventing a medical condition associated with a herpes virus infection, the method comprising administering a therapeutically effective amount of an immune cell as defined herein and an HDAC inhibitor as defined herein, the immune cell comprising a chimeric antigen receptor (CAR) as defined herein on its cell surface.

[0069] The present invention further relates to the use of an HDAC inhibitor in the preparation of a medicament for treating or preventing a medical condition associated with a herpes virus infection, the method comprising further administering an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface, wherein the CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or to a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell (latently) infected with the herpes virus.

[0070] In a related embodiment, the present invention relates to the use of an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface in the preparation of a medicament for treating or preventing a medical condition associated with a herpes virus infection, the method comprising further administering an HDAC inhibitor, wherein the CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell (latently) infected with the herpes virus.

[0071] "Cells infected with herpesviruses" refers to cells containing herpesvirus genomic DNA. Human cells may be latently infected with herpesviruses. "Cells latently infected with herpesviruses" refers to cells that (i) contain herpesvirus genomic DNA viruses and (ii) do not release herpesviruses / virions.

[0072] "Treatment" or "relief" refers to both therapeutic treatment and preventive or prophylactic measures, wherein the purpose is to prevent, slow down (mitigate) or at least partially alleviate or eliminate pathological abnormal conditions in an organism, such as the medical conditions associated with herpes simplex virus infection. Those that need treatment include those that have suffered from the condition and those that tend to have the condition or those in which the disease is to be prevented (prevented). The term "prevention" refers to reducing the likelihood of an organism infecting or developing an abnormal condition (such as the medical condition associated with herpes simplex virus infection).

[0073] The term "therapeutically effective amount" refers to the amount (or number) of a drug for a medical condition associated with herpes virus infection, an HDAC inhibitor according to the present invention, an immune cell comprising CAR on its cell surface, or a pharmaceutical product or a complete kit, or an effective "treatment" of a drug. Specifically, in the case of herpes virus-related cancers, a therapeutically effective amount of HDAC inhibitors and / or immune cells can reduce the number of cancer cells; reduce tumor size; inhibit or stop cancer cells from infiltrating peripheral organs; inhibit and stop tumor metastasis; inhibit and stop tumor growth; alleviate one or more symptoms associated with cancer to a certain extent, or a combination of these effects on cancer cells.

[0074] The present invention is partly based on the induction of the lytic cycle in cells infected by herpesvirus (latency). Without wishing to be bound by theory, the induction of the lytic cycle can increase the cell surface expression of herpesvirus specific proteins (herpesvirus antigens). This can be achieved by, for example, using HDAC inhibitors.

[0075] Chromatin structure and gene transcription (comprising the gene that herpesvirus is relevant) are all subject to the strict regulation and control of the acetylation state of histone molecules in nucleosome.Histone acetyltransferase (HAT) and HDAC play an important role in the epigenetic regulation of this cell gene transcription.The conservative lysine residue in the HAT acetylation histone tail, and is combined with transcription coactivator and other HAT to promote gene transcription, and HDAC is usually combined with the co-repressor protein (such as SMRT, N-Cor, NURD etc.) of different groups to remove acetyl group from the acetylated lysine of histone tail, makes chromatin compaction, and induces transcription inhibition.Therefore, the use of HDAC inhibitor can increase the quantity of herpesvirus antigens and / or their expression level at cell surface.

[0076] As used herein, "HDAC inhibitor" refers to a compound, such as a small molecule, peptide or protein, that inhibits histone deacetylase. One skilled in the art is able to determine whether a compound inhibits a (specific) histone deacetylase. HDAC activity assays are commercially available, including the Amplite TMFluorescence HDAC activity assay kit, Merck (Sigma-Aldrich) HDAC activity assay kit, abcam's histone deacetylase (HDAC) activity assay kit (fluorescence) (ab156064) or Epigentek's Epigenase HDAC activity / inhibition direct assay kit (colorimetric). In the context of the present invention, HDAC inhibitors are preferably capable of inducing the lytic cycle of herpesviruses such as EBV. HDAC inhibitors are preferably also capable of inducing the expression of herpesvirus antigens. HDAC inhibitors are also capable of inducing the surface expression of higher levels of viral antigens.

[0077] The HDAC inhibitor can be selected from butyric acid, valproic acid, MS275 (entinostat), apicidin, largazole, oxamflatin, scriptaid, suberoylanilide hydroxamic acid (SAHA, vorinostat), LBH589 (panobinostat), 12-O-tetradecanoylphorbol-13-acetate (TPA) and PXD101 (belinostat), preferably SAHA, TPA or valproic acid, more preferably SAHA. Figure 1 The structures of these HDAC inhibitors are shown. Table 1 shows HDAC inhibitors suitable for use in the present invention and their HDAC specificity.

[0078] Table 1: Summary of HDAC inhibitors suitable for use in the present invention.

[0079]

[0080]

[0081] The HDAC inhibitor may be butyric acid. The HDAC inhibitor may be valproic acid. The HDAC inhibitor may be MS275 (entinostat). The HDAC inhibitor may be apicidin. The HDAC inhibitor may be largozole, largozale-A or largozole-B. The HDAC inhibitor may be oxamflatin. The HDAC inhibitor may be scriptaid. The HDAC inhibitor may be SAHA (vorinostat). The HDAC inhibitor may be LBH589 (panobinostat). The HDAC inhibitor may be PXD101 (belinostat). The HDAC inhibitor may be TPA.

[0082] There are many types of herpes viruses known to infect humans, and the main herpes viruses of interest within the scope of the present invention are herpes simplex viruses 1 and 2 (HSV-1 and HSV-2, also known as HHV1 and HHV2), varicella zoster virus (VZV, also known as HHV-3), Epstein-Barr virus (EBV or HHV-4), human cytomegalovirus (HCMV or HHV-5), human herpesvirus 6A and 6B (HHV-6A and HHV-6B), human herpesvirus 7 (HHV-7) and Kaposi's sarcoma-associated herpesvirus (KSHV, also known as HHV-8). Herpes viruses have a common structure consisting of a relatively large double-stranded linear DNA genome encoding 70-250 genes enclosed in an icosahedral protein cage called a capsid, which itself is enclosed in a protein layer called a tegument, which contains viral proteins and viral mRNA, and a lipid bilayer membrane called an envelope. When the virus particles contact cells with a specific type of receptor molecule (e.g., CD21) on the cell surface, infection is initiated. Therefore, the herpes virus can be selected from herpes simplex virus 1 and 2 (HSV-1 and HSV-2, also referred to as HHV1 and HHV2), varicella zoster virus (VZV, also referred to as HHV-3), Epstein-Barr virus (EBV or HHV-4), human cytomegalovirus (HCMV or HHV-5), human herpes virus 6A and 6B (HHV-6A and HHV-6B), human herpes virus 7 (HHV-7) and Kaposi's sarcoma-associated herpes virus (KSHV, also referred to as HHV-8), preferably EBV. The herpes virus can be HHV-1. The herpes virus can be HHV-2. The herpes virus can be HHV-3. The herpes virus can be HHV-4 (EBV). The herpes virus can be HHV-5. The herpes virus can be HHV-6A. The herpes virus may be HHV-6B. The herpes virus may be HHV-7. The herpes virus may be HHV-8.

[0083] EBV is a human herpes virus carried by almost all adult populations in the world. Many primary infections occur in childhood and are asymptomatic. Those infections occurring after about 12 or 13 years old are more likely to be accompanied by infectious mononucleosis, which is a self-limiting but temporarily debilitating immunopathology. Whether the infection is accompanied by disease or not, the virus persists. It sets up the incubation period in long-lived memory B cells, and reactivates sporadically, falls off in saliva for oral transmission to new hosts or returns to B cells to supplement B cell storage depots. The vast majority of people will not suffer adverse effects due to the persistence of EBV, but the virus is still relevant to many B cells and epithelial cell malignancies, and may be cause-effect related, and the B cells and epithelial cell malignancies include Burkitt and Hodgkin lymphoma, immunosuppressive post-transplant lymphoproliferative disorders and immunoblastic lymphoma, undifferentiated nasopharyngeal carcinoma and a portion of gastric cancer. It is also associated with autoimmune diseases such as multiple sclerosis (Hutt-Fletcher et al., (2015), Future Virol., 10(10): 1155-1162, Bjornevik et al., (2022), Science, 375(6578): 296-301, Levin et al., (2005), JAMA, 293(20): 2496-500).

[0084] Therefore, the medical condition associated with herpes virus infection can be a cancer associated with herpes virus, wherein herpes virus antigens are present on the surface of cancer cells. For example, this medical condition includes cancer associated with herpes virus or chronic active herpes virus infection or primary herpes virus infection. Particularly relevant is the use of CAR and immune cells as described herein to treat lymphoproliferative disorders (LPD), such as B cell lymphoma and T cell lymphoma, the B cell lymphoma includes Burkitt lymphoma (BL), Hodgkin lymphoma (HL), diffuse large B cell lymphoma (DLBCL). B cell lymphoma is a type of lymphoma that affects B cells. Lymphoma is often referred to as "blood cancer" in lymph nodes. They are more common in the elderly and immunocompromised individuals. B cell lymphoma includes Hodgkin lymphoma and most non-Hodgkin lymphomas.

[0085] DLBCL is a cancer of a B cell. Usually, DLBCL is produced by normal B cells, but it can also represent the malignant transformation of other types of lymphoma or leukemia. Potential immunodeficiency is an important risk factor, and it is also found that the infection of Epstein-Barr virus promotes the development of DLBCL. In one embodiment, the cancer associated with herpes virus is DLBCL.

[0086] Burkitt's lymphoma is a cancer of the B lymphocytes of the lymphatic system, particularly found in the germinal center. Burkitt's lymphoma can be divided into three main clinical variants: endemic, sporadic and immunodeficiency-related variants. EBV infection is present in almost all endemic variants. In one embodiment, the herpes virus-related cancer is Burkitt's lymphoma.

[0087] Also relevant in the context of the present invention is the treatment or prevention of post-transplant lymphoproliferative disorder (PTLD), epithelial cell carcinoma (nasopharyngeal carcinoma, lung cancer, breast cancer), lymphoepithelioma, lymphostromal carcinoma (GCLS, such as gastric cancer) or glioma. PTLD is the name for the proliferation of B cells caused by therapeutic immunosuppression after organ transplantation. These patients may develop infectious mononucleosis-like lesions or polyclonal polymorphic B cell hyperplasia. Therefore, the present invention also relates to the treatment of immunodeficient or immunocompromised patients after chemotherapy, radiotherapy, immunosuppression or transplantation. In one embodiment, the herpes virus-related cancer is PTLD.

[0088] Nasopharynx cancer or nasopharyngeal carcinoma (NPC) is the most common cancer originating from the nasopharynx, most commonly in the posterolateral nasopharynx or pharyngeal recess, accounting for 50% of cases. NPC occurs in children and adults. The association between Epstein-Barr virus and nasopharyngeal carcinoma is clear in World Health Organization (WHO) type II and type III tumors. In one embodiment, the herpes virus-related cancer is NPC.

[0089] Lymphoepithelioma is a poorly differentiated nasopharyngeal carcinoma characterized by significant infiltration of lymphocytes in the tumor-involved area. In the WHO classification system, lymphoepithelioma is also referred to as "category III nasopharyngeal carcinoma." In one embodiment, the herpes virus-related cancer is lymphoepithelioma.

[0090] Lymphoid stromal gastric cancer (GCLS) is a unique histological subtype of gastric cancer characterized by undifferentiated carcinoma mixed with significant lymphoid infiltration. More than 80% of GCLS cases are associated with EBV infection, but it is unclear whether the virus affects disease progression. In one embodiment, the herpes virus-related cancer is GCLS.

[0091] Glioma is a type of tumor that begins in glial cells of the brain or spine. Glioma accounts for about 30% of all brain tumors and central nervous system tumors and 80% of all malignant brain tumors. Studies have revealed that EBV is present in glioma patients at an elevated frequency, suggesting that EBV-related gliomas can be potentially targeted using the present invention. In one embodiment, the herpes virus-related cancer is a glioma.

[0092] The present invention can be used not only for treating the cancer associated with herpes simplex virus, but also for treating actual herpes simplex virus infection itself. Therefore, the medical condition to be treated or prevented and associated with EBV infection can be chronic active EBV infection (CAEBV) or primary EBV infection (e.g. mononucleosis). CAEBV is a rare and usually fatal EBV infection complication, which most often occurs in children or adolescents of Asian or South American pedigrees, although there are also case reports in Spaniards, Europeans and Africans. At present, the only known cure for CAEBV is allogeneic hematopoietic stem cell transplantation (HSCT), and all other treatment options (rituximab, cytotoxic chemotherapy and immunosuppressive therapy) ultimately show as invalid. In one embodiment, the medical condition is CAEBV.

[0093] Infectious mononucleosis, also known as glandular fever, is an infection usually caused by EBV. Most people are infected with the virus as children, when the disease produces few or no symptoms. In adolescents and young adults, the disease usually causes fever, sore throat, swollen lymph nodes in the neck, and fatigue. In one embodiment, the medical condition is infectious mononucleosis.

[0094] Glycoprotein plays an important role in the formation of herpes virus particles and infection of host cells. The most abundant EBV virion glycoprotein is gp350, which is identified as the protein responsible for the attachment of the virus to B lymphocytes. Gp350 is a type 1 membrane protein of 907 residues, which exists as two splice variants in virions, and they have a mass of about 350 and 220 kDa when fully glycosylated. The splicing maintains the reading frame, and the two forms of protein retain the N-terminal attachment site, i.e., a surface without polysaccharides that tethers the virus to CR2 / CD21. It is unclear whether maintaining splice variants has any functional significance. However, since both CR2 and gp350 are modeled as stretched proteins, they initially locate the virus at a distance from the cell surface, and perhaps the sequential combination with gp350 and gp220 is related to making the virus closer to the cell membrane. Therefore, any reference to gp350 herein relates to gp350 or gp220, or vice versa. Thus, the herpes virus antigen may be gp350.

[0095] After attaching to the B cell surface, EBV as an enveloped virus enters the cell through the fusion of its envelope and cell membrane. The fusion occurring from the endocytic vesicle requires the action of four other glycoproteins gB, gH, gL and gp42 (Hutt-Fletcher et al., (2015), Future Virol., 10 (10): 1155-1162). Glycoprotein B is a type I membrane protein expressed in the form of homotrimers. In all herpes viruses, it is generally recognized that gB is the final executor of fusion. Glycoproteins gH, gL and gp42 are regarded as regulators of the process. Glycoprotein gp42 can be present in cells as a type 2 membrane protein or as a soluble protein cut off as a signal sequence. It interacts with gH through the flexible segment in its N-terminal domain and interacts with class II HLA through its C-terminal C-type lectin domain.

[0096] In the context of the present invention, the immune cells comprising CAR on its surface utilize herpes virus antigens to bind and eliminate (latent) cells infected with herpes virus, so as to treat or prevent the medical conditions related to herpes virus infection. Such herpes virus antigens are proteins encoded by herpes virus or protein complexes comprising the protein. Table 2 below shows EBV glycoproteins. These EBV glycoproteins can be used as markers for the lytic phase, or as herpes virus antigens.

[0097] Table 2: Overview of EBV glycoproteins.

[0098]

[0099]

[0100] In a preferred embodiment, herpesvirus antigen is an EBV antigen. In a preferred embodiment, the present invention focuses on the medical condition that targeting EBV-antigen is also treated EBV. In a preferred embodiment, herpesvirus antigen is EBV glycoprotein 350 / 220 (gp350 / gp220). Exemplary EBV gp350 protein is shown in UniProt database, accession number P03200-1, the 1st edition on July 21, 1986. Exemplary EBV gp220 is shown in identical database entry, but position 502 to 750 is missing.

[0101] In a preferred embodiment, the EBV antigen is present on the surface of an EBV infected cell.In a preferred embodiment, the EBV antigen is present on the surface of an EBV infected cancer cell, an EBV infected B cell, or an EBV infected epithelial cell.

[0102] Using the methods described herein and reference methods established in the art, one of skill in the art can identify viral antigens to be targeted (other than those specific antigens described herein) without undue effort.

[0103] The herpes virus antigen subgroup present on the cell surface infected by EBV and possibly present on (latent) infected malignant cells includes gp350 / gp220, LMP1, LMP2, BILF1, BALF4 (gB), BKRF2 (gL), BXLF2 (gH), BBRF3 (gM), BLRF1 (gN). Among them, gp350 / gp220, BILF2, BALF4 (gB), BKRF2 (gL), BXLF2 (gH), BBRF3 (gM), BLRF1 (gN) are particularly preferred as herpes virus antigens.

[0104] The EBV antigen (herpesvirus antigen when the herpesvirus is EBV) may also be in an EBV virion envelope protein or a protein of the EBV envelope complex, such as gB, gL or gH (see also Table 2).

[0105] Although HDAC inhibitors increase the susceptibility of (latently) infected cells to immune cell-mediated killing that has already been induced by the innate immune response, in the context of the present invention, such killing is synergistically supported by administering immune cells (e.g., CAR-T cells) that contain a CAR on their cell surface.

[0106] Immune cells can be selected from T lymphocytes, NK cells, macrophages and dendritic cells, wherein T lymphocytes are preferably cytotoxic T lymphocytes or T helper cells, more preferably cytotoxic T lymphocytes. Immune cells can be T lymphocytes. T lymphocytes can be cytotoxic T lymphocytes. T lymphocytes can be T helper cells. Immune cells can be NK cells. Immune cells can be macrophages. Immune cells can be dendritic cells. It is known in the art that these immune cells respond to unwanted reagents, cells or pathogens (such as cells infected by herpes virus) and show cytotoxicity and / or other beneficial activities. By directing the activity of these cells to specific immunogenic targets, i.e. herpes virus antigens as described herein, the pathogenic cells infected can be eliminated by the corresponding activity of immune cells as described herein.

[0107] In some embodiments, the immune cells of CAR engineering can be engineered to co-express cytokines (such as IL-15, IL-12, IFN-γ, IFN-α, GM-CSF, FLT3L, IL-21, IL-23) or co-stimulatory ligands (CD80, CD86, CD40L) to improve immunotherapy effects. In some embodiments, the immune cells of CAR-engineering can be engineered to co-express additional CAR targeting specific cell lineages.

[0108] According to the present invention, CAR (polypeptide) comprises an extracellular antigen binding domain, which comprises an antibody or antibody fragment binding to a target antigen, a transmembrane domain, and an intracellular domain. CAR is generally described as comprising an extracellular domain (antigen binding domain) derived from an antibody and an intracellular domain comprising a signal transduction module derived from a T cell signaling protein. Exemplary CARs that can be used for the present invention are described in WO2019 / 201995A1, which is incorporated herein by reference.

[0109] In a preferred embodiment, the extracellular domain preferably comprises a variable region of a heavy chain and a light chain from an immunoglobulin, which is constructed as a single chain variable fragment (scFv). ScFv is preferably connected to a hinge region, which provides flexibility and transduces signals to an intracellular signaling domain by anchoring a transmembrane portion. The transmembrane domain is preferably derived from CD8a or CD28. In the first generation of CAR, the signaling domain is composed of the ζ chain of the TCR complex. The term "generation" refers to the structure of the intracellular signaling domain. The second generation CAR is equipped with a single costimulatory domain derived from CD28 or 4-1BB. The third generation CAR has included two costimulatory domains, such as CD28, 4-1BB, ICOS or OX40, CD3ζ. The present invention preferably relates to a second or third generation CAR, although the antigen binding fragments described herein can be used in any given CAR form.

[0110] The CAR used in the context of the present invention includes an extracellular domain (also referred to as a binding domain or antigen binding domain) combined with herpes virus antigens (preferably EBV gp350 / gp220), a transmembrane domain and an intracellular domain or an intracellular signaling domain. The engagement of the antigen binding domain of CAR with the target on the target cell surface causes the clustering of CAR, and the activation stimulus is delivered to the cell containing CAR. The main feature of CAR is their ability to redirect immune effector cell specificity, thereby inducing the proliferation of antigen-specific effector T cells, cytokine production (such as IFN-γ) and the generation of molecules that can mediate the death of target cells expressing surface antigens in a manner independent of major histocompatibility complex (MHC).

[0111] In various embodiments, CAR comprises an extracellular binding domain comprising a (optionally humanized) gp350 / gp220 specific binding domain; a transmembrane domain; one or more intracellular signaling domains. In a specific embodiment, CAR comprises an extracellular binding domain comprising an (optionally humanized) anti-gp350 / gp220 antigen binding fragment; one or more spacer domains; a transmembrane domain; one or more intracellular signaling domains.

[0112] "Extracellular antigen binding domain" or "extracellular binding domain" are used interchangeably and provide a CAR having the ability to specifically bind to herpes virus antigens. The binding domain can be derived from natural, synthetic, semisynthetic or recombinant sources. Preferably, it is a scFv domain.

[0113] "Single-chain Fv" or "scFv" antibody fragments include the VH and VL domains of an antibody, wherein these domains are present in a single polypeptide chain and adopt either orientation (e.g., VL-VH or VH-VL), such as from antibody 7D11 described herein. Typically, the scFv polypeptide also includes a polypeptide linker between the VH and VL domains, which enables the scFv to form a structure required for antigen binding. In a preferred embodiment, the CAR contemplated herein includes an antigen-specific binding domain, which is an scFv and can be a mouse, human or humanized scFv. Single-chain antibodies can be cloned from the V region genes of a hybridoma specific for the desired target. In a specific embodiment, the antigen-specific binding domain is a humanized scFv that binds to EBV gp350 and / or gp220 polypeptides.

[0114] In one embodiment, CAR comprises a scFv derived from the 7D11 antibody described herein. Thus, CAR may comprise the amino acid sequence shown in SEQ ID NO:2. CAR may also consist of the amino acid sequence shown in SEQ ID NO:2. CAR may have an amino acid sequence having at least 85% identity to the amino acid sequence shown in SEQ ID NO:2. CAR may have an amino acid sequence having at least 90% identity to the amino acid sequence shown in SEQ ID NO:2. CAR may have an amino acid sequence having at least 95% identity to the amino acid sequence shown in SEQ ID NO:2. CAR may have an amino acid sequence having at least 98% identity to the amino acid sequence shown in SEQ ID NO:2. CAR may have an amino acid sequence having at least 99% identity to the amino acid sequence shown in SEQ ID NO:2. In another embodiment, CAR comprises a scFv derived from the 6H3 antibody described herein. Thus, CAR may comprise the amino acid sequence shown in SEQ ID NO:12. CAR may also consist of the amino acid sequence shown in SEQ ID NO:12. CAR may have an amino acid sequence having at least 85% identity to the amino acid sequence shown in SEQ ID NO:12. CAR may have an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO: 12. CAR may have an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO: 12. CAR may have an amino acid sequence that is at least 98% identical to the amino acid sequence shown in SEQ ID NO: 12. CAR may have an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 12. In another embodiment, CAR comprises an scFv derived from the 7A4 antibody described herein. Thus, CAR may comprise the amino acid sequence shown in SEQ ID NO: 22. CAR may also consist of the amino acid sequence shown in SEQ ID NO: 22. CAR may have an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 22. CAR may have an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO: 22. CAR may have an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO: 22. CAR may have an amino acid sequence that is at least 98% identical to the amino acid sequence shown in SEQ ID NO: 22. The CAR may have an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 22. In yet another embodiment, the CAR comprises a scFv derived from the 7A8 antibody described herein.Thus, the CAR may comprise the amino acid sequence shown in SEQ ID NO: 32. The CAR may also consist of the amino acid sequence shown in SEQ ID NO: 32. The CAR may have an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 32. The CAR may have an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO: 32. The CAR may have an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO: 32. The CAR may have an amino acid sequence that is at least 98% identical to the amino acid sequence shown in SEQ ID NO: 32. The CAR may have an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 32.

[0115] In one embodiment, CAR comprises VL and VH domains derived from the 7D11 antibody disclosed herein. Thus, CAR may comprise a VH domain as shown in SEQ ID NO:3 and a VL domain as shown in SEQ ID NO:4. CAR may comprise a VH domain having an amino acid sequence with at least 85% sequence identity to the amino acid sequence as shown in SEQ ID NO:3 and a VL domain having an amino acid sequence with at least 85% sequence identity to the amino acid sequence as shown in SEQ ID NO:4. CAR may comprise a VH domain having an amino acid sequence with at least 90% sequence identity to the amino acid sequence as shown in SEQ ID NO:3 and a VL domain having an amino acid sequence with at least 90% sequence identity to the amino acid sequence as shown in SEQ ID NO:4. CAR may comprise a VH domain having an amino acid sequence with at least 95% sequence identity to the amino acid sequence as shown in SEQ ID NO:3 and a VL domain having an amino acid sequence with at least 95% sequence identity to the amino acid sequence as shown in SEQ ID NO:4. CAR may include a VH domain having an amino acid sequence having at least 98% sequence identity with the amino acid sequence as shown in SEQ ID NO: 3 and a VL domain having an amino acid sequence having at least 98% sequence identity with the amino acid sequence as shown in SEQ ID NO: 4. CAR may include a VH domain having an amino acid sequence having at least 99% sequence identity with the amino acid sequence as shown in SEQ ID NO: 3 and a VL domain having an amino acid sequence having at least 99% sequence identity with the amino acid sequence as shown in SEQ ID NO: 4. In another embodiment, CAR comprises VL and VH domains derived from the 6H3 antibody disclosed herein. Thus, CAR may include a VH domain as shown in SEQ ID NO: 13 and a VL domain as shown in SEQ ID NO: 14. CAR may include a VH domain having an amino acid sequence having at least 85% sequence identity with the amino acid sequence as shown in SEQ ID NO: 13 and a VL domain having an amino acid sequence having at least 85% sequence identity with the amino acid sequence as shown in SEQ ID NO: 14. The CAR may comprise a VH domain having an amino acid sequence having at least 90% sequence identity to the amino acid sequence shown in SEQ ID NO: 13 and a VL domain having an amino acid sequence having at least 90% sequence identity to the amino acid sequence shown in SEQ ID NO: 14.CAR may comprise a VH domain having an amino acid sequence having at least 95% sequence identity with the amino acid sequence as shown in SEQ ID NO: 13 and a VL domain having an amino acid sequence having at least 95% sequence identity with the amino acid sequence as shown in SEQ ID NO: 14. CAR may comprise a VH domain having an amino acid sequence having at least 98% sequence identity with the amino acid sequence as shown in SEQ ID NO: 13 and a VL domain having an amino acid sequence having at least 98% sequence identity with the amino acid sequence as shown in SEQ ID NO: 14. CAR may comprise a VH domain having an amino acid sequence having at least 99% sequence identity with the amino acid sequence as shown in SEQ ID NO: 13 and a VL domain having an amino acid sequence having at least 99% sequence identity with the amino acid sequence as shown in SEQ ID NO: 14. In another embodiment, CAR comprises VL and VH domains derived from the 7A4 antibody disclosed herein. Thus, CAR may comprise a VH domain as shown in SEQ ID NO: 23 and a VL domain as shown in SEQ ID NO: 24. The CAR may comprise a VH domain having an amino acid sequence having at least 85% sequence identity with the amino acid sequence as shown in SEQ ID NO: 23 and a VL domain having an amino acid sequence having at least 85% sequence identity with the amino acid sequence as shown in SEQ ID NO: 24. The CAR may comprise a VH domain having an amino acid sequence having at least 90% sequence identity with the amino acid sequence as shown in SEQ ID NO: 23 and a VL domain having an amino acid sequence having at least 90% sequence identity with the amino acid sequence as shown in SEQ ID NO: 24. The CAR may comprise a VH domain having an amino acid sequence having at least 95% sequence identity with the amino acid sequence as shown in SEQ ID NO: 23 and a VL domain having an amino acid sequence having at least 95% sequence identity with the amino acid sequence as shown in SEQ ID NO: 24. The CAR may comprise a VH domain having an amino acid sequence having at least 98% sequence identity with the amino acid sequence as shown in SEQ ID NO: 23 and a VL domain having at least 98% sequence identity with the amino acid sequence as shown in SEQ ID NO: 24. The CAR may comprise a VH domain having an amino acid sequence having at least 99% sequence identity to the amino acid sequence shown in SEQ ID NO: 23 and a VL domain having an amino acid sequence having at least 99% sequence identity to the amino acid sequence shown in SEQ ID NO: 24. In another embodiment, the CAR comprises VL and VH domains derived from the 7A8 antibody disclosed herein.Thus, the CAR may comprise a VH domain as shown in SEQ ID NO: 33 and a VL domain as shown in SEQ ID NO: 34. The CAR may comprise a VH domain having an amino acid sequence having at least 85% sequence identity with the amino acid sequence as shown in SEQ ID NO: 33 and a VL domain having an amino acid sequence having at least 85% sequence identity with the amino acid sequence as shown in SEQ ID NO: 34. The CAR may comprise a VH domain having an amino acid sequence having at least 90% sequence identity with the amino acid sequence as shown in SEQ ID NO: 33 and a VL domain having an amino acid sequence having at least 90% sequence identity with the amino acid sequence as shown in SEQ ID NO: 34. The CAR may comprise a VH domain having an amino acid sequence having at least 95% sequence identity with the amino acid sequence as shown in SEQ ID NO: 33 and a VL domain having an amino acid sequence having at least 95% sequence identity with the amino acid sequence as shown in SEQ ID NO: 34. The CAR may comprise a VH domain having an amino acid sequence having at least 98% sequence identity with the amino acid sequence shown in SEQ ID NO: 33 and a VL domain having an amino acid sequence having at least 98% sequence identity with the amino acid sequence shown in SEQ ID NO: 34. The CAR may comprise a VH domain having an amino acid sequence having at least 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 33 and a VL domain having an amino acid sequence having at least 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 34.

[0116] In the context of the present invention, it is not important whether the HDAC inhibitor or the immune cells containing CAR on its cell surface are first administered to the patient. Both can also be administered simultaneously, for example within 1 hour or less. Therefore, the HDAC inhibitor can be administered to the patient before the immune cells containing CAR on its cell surface are administered. The HDAC inhibitor can be administered to the patient after the immune cells containing CAR on its cell surface. The HDAC inhibitor and the immune cells containing CAR on its surface can also be administered to the patient simultaneously.

[0117] The term "administer" refers to a method of introducing a compound into a cell or tissue (including a blood vessel) of an organism. Exemplary modes of administration include, but are not limited to, oral, rectal, ocular (including intravitreal or intracameral), nasal, topical (including buccal and sublingual), intrauterine, vaginal or parenteral (including subcutaneous, intraperitoneal, intramuscular, intravenous, intradermal, intracranial, intratracheal and epidural), transdermal, intraperitoneal, intracranial, intraventricular, intracerebral, intravaginal, intrauterine or parenteral (e.g., intravenous, intraspinal, subcutaneous or intramuscular) routes. Oral or intravenous administration is preferred.

[0118] HDAC inhibitors and immune cells comprising CAR on their surface can be administered to subjects. In the context of the present invention, "subject" is preferably a person. For example, the subject can be a patient suspected of having a medical condition associated with herpes virus infection or a patient diagnosed with such a medical condition.

[0119] HDAC inhibitors and immune cells containing CAR on their cell surface do not necessarily need to be administered using the same administration method. For example, immune cells containing CAR on their cell surface can be administered intravenously, while HDAC inhibitors are administered orally. However, both can also be administered intravenously.

[0120] The present invention also relates to a kit of parts, comprising an immune cell as defined herein and an HDAC inhibitor as defined herein, wherein the immune cell comprises a chimeric antigen receptor (CAR) on its cell surface. The term "kit of parts", "combined preparations" or also referred to as "juxtapositions" means herein that the components of the combined preparations do not need to exist as a joint (the components in the combination do not directly interact with each other), for example in a composition, so that they can be used separately or sequentially. Therefore, the term "juxtaposition" means that there is no need for a real combination in terms of the physical separation of the components.

[0121] The present invention also relates to a drug product comprising an immune cell as defined herein and an HDAC inhibitor as defined herein, wherein the immune cell comprises a chimeric antigen receptor (CAR) on its cell surface. The drug product can be a composition, wherein both the immune cell and the HDAC inhibitor comprising CAR on its cell surface are combined, for example, in the form of a "ready-to-use" solution for (simultaneous) injection.

[0122] Treatment in the context of the present invention is based on the combination and coordinated administration of HDAC inhibitors and immune cells comprising CAR on their cell surfaces. Therefore, the present invention relates to a kit as defined herein or a pharmaceutical product as defined herein, which is used in combination in a method for treating or preventing a medical condition associated with herpes virus infection. The combined use may include administering an HDAC inhibitor to a patient before the administration of immune cells, administering an HDAC inhibitor and immune cells to the patient at the same time, or administering immune cells to the patient after the HDAC inhibitor is administered.

[0123] Also provided herein are antibodies that bind to the gp350 of EBV. These antibodies can be used to diagnose whether EBV infection or the lytic cycle of EBV has been activated. Here, the presence of gp350 on the surface of target cells in samples obtained from patients indicates that the lytic cycle of EBV has been induced. Antibodies comprising SEQ ID NO:3 and 4 are derived from the 7D11 antibody described herein. Other antibodies comprising SEQ ID NO:13 and 14 are derived from the 6H3 antibody described herein. Other antibodies comprising SEQ ID NO:23 and 24 are derived from the 7A4 antibody described herein. Other antibodies comprising SEQ ID NO:33 and 34 are derived from the 7A8 antibody described herein. As shown in the examples, these antibodies show enhanced efficacy in targeting and killing EBV-infected cells, and show reduced toxicity. The same applies to effector cells (e.g., CAR-T cells) expressing CAR on their cell surfaces, wherein CAR comprises variable domains of antibodies of the present invention, such as SEQ ID NO:3 and 4; SEQ ID NO:13 and 14; SEQ ID NO:23 and 24; or SEQ ID NO:33 and 34.

[0124] Therefore, the present invention relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0125] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:3, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:4; and / or

[0126] (b) Binds to the same epitope in the EBV gp350 protein as the epitope to which the antibody of (a) binds.

[0127] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:2, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:2, or the antibody is an antibody that binds to the same epitope.

[0128] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:2, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:2, or the antibody is an antibody that binds to the same epitope.

[0129] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:2, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:2, or the antibody is an antibody that binds to the same epitope.

[0130] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:2, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:2, or the antibody is an antibody that binds to the same epitope.

[0131] The present invention further relates to an antibody to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:3, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:4, or the antibody is an antibody that binds to the same epitope.

[0132] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:3, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:4, or the antibody is an antibody that binds to the same epitope.

[0133] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:3, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:4, or the antibody is an antibody that binds to the same epitope.

[0134] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:3, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:4, or the antibody is an antibody that binds to the same epitope.

[0135] The present invention also relates to an antibody, wherein the antibody is an antibody comprising a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 3 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 4.

[0136] The present invention also relates to an antibody, wherein the antibody is an antibody composed of a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 3 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 4.

[0137] In this context, the present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0138] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having the amino acid sequence shown in SEQ ID NO:5, a heavy chain CDR2 having the amino acid sequence shown in SEQ ID NO:6, and a heavy chain CDR3 having the amino acid sequence shown in SEQ ID NO:7, and

[0139] A light chain variable region, the light chain variable region comprising a light chain CDR1 having the amino acid sequence shown in SEQ ID NO: 8, a light chain CDR2 having the amino acid sequence shown in SEQ ID NO: 9, and a light chain CDR3 having the amino acid sequence shown in SEQ ID NO: 10; and / or

[0140] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0141] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0142] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 13, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 14; and / or

[0143] (b) Binds to the same epitope in the EBV gp350 protein as the epitope to which the antibody of (a) binds.

[0144] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 12, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 12, or the antibody is an antibody that binds to the same epitope.

[0145] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO: 12, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO: 12, or the antibody is an antibody that binds to the same epitope.

[0146] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO: 12, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO: 12, or the antibody is an antibody that binds to the same epitope.

[0147] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 12, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 12, or the antibody is an antibody that binds to the same epitope.

[0148] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:13, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:14, or the antibody is an antibody that binds to the same epitope.

[0149] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO: 13, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO: 14, or the antibody is an antibody that binds to the same epitope.

[0150] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:13, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:14, or the antibody is an antibody that binds to the same epitope.

[0151] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 13, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 14, or the antibody is an antibody that binds to the same epitope.

[0152] The present invention also relates to an antibody, wherein the antibody is an antibody comprising a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 13 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 14.

[0153] The present invention also relates to an antibody, wherein the antibody is an antibody composed of a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 13 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 14.

[0154] In this context, the present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0155] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having the amino acid sequence shown in SEQ ID NO: 15, a heavy chain CDR2 having the amino acid sequence shown in SEQ ID NO: 16, and a heavy chain CDR3 having the amino acid sequence shown in SEQ ID NO: 17, and

[0156] A light chain variable region comprising a light chain CDR1 having an amino acid sequence as shown in SEQ ID NO: 18, a light chain CDR2 having an amino acid sequence as shown in SEQ ID NO: 19, and a light chain CDR3 having an amino acid sequence as shown in SEQ ID NO: 20; and / or

[0157] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0158] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0159] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 23, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 24; and / or

[0160] (b) Binds to the same epitope in the EBV gp350 protein as the epitope to which the antibody of (a) binds.

[0161] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:22, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:22, or the antibody is an antibody that binds to the same epitope.

[0162] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:22, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:22, or the antibody is an antibody that binds to the same epitope.

[0163] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:22, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:22, or the antibody is an antibody that binds to the same epitope.

[0164] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 22, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO: 22, or the antibody is an antibody that binds to the same epitope.

[0165] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:23, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:24, or the antibody is an antibody that binds to the same epitope.

[0166] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:23, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:24, or the antibody is an antibody that binds to the same epitope.

[0167] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:23, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:24, or the antibody is an antibody that binds to the same epitope.

[0168] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:23, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:24, or the antibody is an antibody that binds to the same epitope.

[0169] The present invention also relates to an antibody, wherein the antibody is an antibody comprising a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 23 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 24.

[0170] The present invention also relates to an antibody, wherein the antibody is an antibody composed of a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 23 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 24.

[0171] In this context, the present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0172] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having the amino acid sequence shown in SEQ ID NO: 25, a heavy chain CDR2 having the amino acid sequence shown in SEQ ID NO: 26, and a heavy chain CDR3 having the amino acid sequence shown in SEQ ID NO: 27, and

[0173] A light chain variable region comprising a light chain CDR1 having an amino acid sequence as shown in SEQ ID NO: 28, a light chain CDR2 having an amino acid sequence as shown in SEQ ID NO: 29, and a light chain CDR3 having an amino acid sequence as shown in SEQ ID NO: 30; and / or

[0174] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0175] The present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0176] (a) comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 33, and the light chain variable region having an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO: 34; and / or

[0177] (b) Binds to the same epitope in the EBV gp350 protein as the epitope to which the antibody of (a) binds.

[0178] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:32, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:32, or the antibody is an antibody that binds to the same epitope.

[0179] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:32, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:32, or the antibody is an antibody that binds to the same epitope.

[0180] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:32, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:32, or the antibody is an antibody that binds to the same epitope.

[0181] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:32, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:32, or the antibody is an antibody that binds to the same epitope.

[0182] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:33, the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in SEQ ID NO:34, or the antibody is an antibody that binds to the same epitope.

[0183] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:33, the light chain variable region has an amino acid sequence that is at least 90% identical to the amino acid sequence shown in SEQ ID NO:34, or the antibody is an antibody that binds to the same epitope.

[0184] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:33, the light chain variable region has an amino acid sequence that is at least 95% identical to the amino acid sequence shown in SEQ ID NO:34, or the antibody is an antibody that binds to the same epitope.

[0185] The present invention further relates to an antibody that binds to the gp350 protein of EBV, wherein the antibody is an antibody comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:33, the light chain variable region has an amino acid sequence that is at least 99% identical to the amino acid sequence shown in SEQ ID NO:34, or the antibody is an antibody that binds to the same epitope.

[0186] The present invention also relates to an antibody, wherein the antibody is an antibody comprising a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 33 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 34.

[0187] The present invention also relates to an antibody, wherein the antibody is an antibody composed of a heavy chain variable region having an amino acid sequence as shown in SEQ ID NO: 33 and a light chain variable region having an amino acid sequence as shown in SEQ ID NO: 34.

[0188] In the context of the present invention, the present invention further relates to an antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody

[0189] (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having an amino acid sequence as shown in SEQ ID NO: 35, a heavy chain CDR2 having an amino acid sequence as shown in SEQ ID NO: 36, and a heavy chain CDR3 having an amino acid sequence as shown in SEQ ID NO: 37, and

[0190] A light chain variable region comprising a light chain CDR1 having an amino acid sequence as shown in SEQ ID NO: 38, a light chain CDR2 having an amino acid sequence as shown in SEQ ID NO: 39, and a light chain CDR3 having an amino acid sequence as shown in SEQ ID NO: 40; and / or

[0191] (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

[0192] The term "antibody" as used herein and in the context of the present invention may include a chimeric antibody, a humanized antibody, a monovalent antibody, a multivalent antibody, a low molecular weight antibody, a diabody or a scFv.

[0193] Chimeric antibodies refer to antibodies comprising variable regions and constant regions of different sources connected to each other. For example, mouse-human heterologous chimeric antibodies are antibodies comprising the heavy and light chain variable regions of mouse antibodies and the heavy and light chain constant regions of human antibodies. The DNA encoding the mouse antibody variable regions is connected to the DNA encoding the human antibody constant regions, and the connection product is integrated into the expression vector to prepare a recombinant vector expressing the chimeric antibody. Cells (recombinant cells) transformed with these vectors can be cultured to express the DNA insert, thereby obtaining the chimeric antibodies produced during the culture process.

[0194] Generally, chimeric antibodies include antibody variable regions of non-human animal origin and constant regions of human antibody origin. In contrast, humanized antibodies include antibody complementary determining regions (CDRs) of non-human animal origin, framework regions (FRs) of human antibody origin, and constant regions of human antibody origin. Humanized antibodies are also referred to as reconstructed human antibodies. Specifically, for example, humanized antibodies comprising non-human animal (e.g., mouse) antibody CDRs transplanted in human antibodies are known in the art. Due to the reduced antigenicity of humanized antibodies in the human body, they can be used as active ingredients of therapeutic agents of the present invention.

[0195] The present invention also provides use of the antibody of the present invention that binds to EBV gp350 in the preparation of a medicament for treating a medical condition associated with herpes virus infection.

[0196] The present invention further provides an antibody of the present invention that binds to gp350 of EBV for use in a method of treating a medical condition associated with a herpes virus infection.

[0197] The present invention also relates to a CAR comprising the heavy chain and light chain variable regions of the antibody of the present invention or the heavy chain CDR1, CDR2 and CDR3 of the antibody of the present invention; and light chain CDR1, CDR2, CDR3. CAR may comprise an amino acid sequence shown in any one of SEQ ID NO: 2, 12, 22 or 32 or consist of an amino acid sequence shown in any one of them.

[0198] In some embodiments, the antibodies of the present invention or described herein that bind to EBV gp350 may comprise a human Fc region modified to enhance effector functions, such as antigen-dependent cell-mediated cytotoxicity (ADCC) and / or complement-dependent cytotoxicity (CDC). This can be achieved by introducing one or more amino acid substitutions in the Fc region of the antibody. For example, cysteine ​​residues may be introduced in the Fc region to allow the formation of interchain disulfide bonds in the region, thereby improving complement-mediated cell killing and antibody-dependent cellular cytotoxicity (ADCC). Homodimeric antibodies with enhanced anti-tumor activity may also be prepared using heterobifunctional cross-linkers known to those skilled in the art. Alternatively, antibodies with dual Fc regions may be engineered.

[0199] In addition, the antibodies described herein may have one or more amino acid substitutions, deletions, additions and / or insertions in their CDR sequences, as long as the resulting antibodies are functionally equivalent to antibodies (a) to (c). The term "functionally equivalent" means that the affinity for gp350 of EBV is equivalent. The term "equivalent" means that compared to antibodies (a) to (c), it has at least 50%, preferably at least 60%, more preferably at least 70%, more preferably at least 80%, even more preferably at least 90%, even more preferably at least 95%, even more preferably at least 99% activity. There is no particular restriction on the upper limit of the activity, which may be higher than the upper limit of antibodies (a) to (c). Affinity can be determined by methods generally known to those skilled in the art.

[0200] The term "homology" is used herein with its common meaning, and is included in the identical amino acids at equivalent positions in the linear amino acid sequences of two proteins compared to each other, and is considered to be a conservative substitution (e.g., by exchanging glutamic acid residues with aspartic acid residues). "Identity" or "sequence identity" refers to the property of the sequence that measures the similarity of a sequence or a relationship. As used in the present invention, the term "sequence identity" or "identity" refers to the percentage of paired identical residues relative to the number of residues in the longer sequence of the two sequences after the polypeptide sequence of the present invention is compared (homology). The identity is measured by dividing the number of identical residues by the total number of residues and multiplying the result by 100.

[0201] For example, the BLASTP program, blastp 2.2.5 version (November 16, 2002; see Altschul, SF et al. (1997) Nucl. Acids Res. 25, 3389-3402) can be used herein to determine the percentage of sequence homology or sequence identity. In this embodiment, the percentage of homology is based on the alignment of the entire polypeptide sequence (optionally including the polypeptide sequence, using the amino acid sequence listed herein as a reference in the pairwise comparison) (matrix: BLOSUM 62; gap penalty: 11.1; cutoff value set to 10 -3 ). This is calculated as the number of "positives" (homologous amino acids) displayed as a percentage of the BLASTP program output divided by the total number of amino acids selected by the program for alignment.

[0202] In one embodiment of the present invention, the antibody binding to gp350 of EBV is conjugated to a cytotoxic substance.

[0203] In a preferred embodiment, the antibody that binds to gp350 of EBV can be conjugated to a cytotoxic substance such as a chemotherapeutic agent, a toxic peptide or a radioactive chemical. Such modified antibodies (hereinafter referred to as antibody conjugates) can be obtained by chemically modifying the obtained antibodies. Methods for antibody modification have been established in the art.

[0204] Examples of chemotherapeutic agents that exert their cytotoxic activity by conjugating to antibodies that bind to gp350 of EBV may include the following chemotherapeutic agents: azaribine, anastrozole, azacytidine, bleomycin, bortezomib, bryostatin-1, busulfan, camptothecin, 10-hydroxycamptothecin, carmustine, celecoxib, chlorambucil, cisplatin, irinotecan, carboplatin, cladribine, cyclophosphamide, cytarabine, dacarbazine, docetaxel, actinomycin, daunorubicin glucuronide, daunorubicin, dexamethasone, diethylstilbestrol, doxorubicin, doxorubicin glucuronide, epirubicin, ethinyl estradiol, estramustine, etoposide, etoposide glucuronide, floxuridine , fludarabine, flutamide, fluorouracil, fluoxymesterone, gemcitabine, hydroxyprogesterone caproate, hydroxyurea, idarubicin, ifosfamide, leucovorin, lomustine, nitrogen mustard, medroxyprogesterone acetate, megestrol acetate, melphalan, mercaptopurine, methotrexate, mitoxantrone, mithramycin, mitomycin, mitotane, phenylbutyric acid, prednisolone, procarbazine, paclitaxel, pentostatin, semustine, streptozotocin, tamoxifen, taxane, taxol, testosterone propionate, thalidomide, thioguanine, thiotepa, teniposide, topotecan, uracil nitrogen mustard, vinblastine, vinorelbine, vincristine.

[0205] Preferably, the chemotherapeutic agent is a low molecular weight chemotherapeutic agent. Such low molecular weight chemotherapeutic agents are unlikely to interfere with the antibody even after being conjugated with the antibody. Low molecular weight chemotherapeutic agents generally have a molecular weight of 100 to 2000, preferably 200 to 1000. All chemotherapeutic agents given above are low molecular weight chemotherapeutic agents. These chemotherapeutic agents include prodrugs that are converted into active chemotherapeutic agents in vivo. The activation of the prodrug can be an enzymatic conversion or a non-enzymatic conversion.

[0206] The antibodies described herein may contain a constant region. The constant region used is not particularly limited, and any constant region can be used. Preferred examples of the constant region used in the present invention may include a constant region of human origin. For example, a constant region derived from human IgG1, human IgG2, human IgG3, or human IgG4 may be used as a heavy chain constant region. In addition, for example, a constant region derived from a human κ chain or a human λ chain may be used as a light chain constant region. The constant region used in the present invention may be a constant region having a native sequence, or may be a modified constant region having a sequence derived from a native sequence by modifying one or more amino acids.

[0207] The antibodies described herein may also contain a framework region (FR). The FR used is not particularly limited, and any FR can be used, as long as the resulting antibody maintains its binding activity to the gp350 of EBV present on the cell surface of the target cell. Preferred examples of the FR used in the present invention may include FRs derived from human antibodies. Since FR replacement techniques that maintain the antigen binding activity of antibodies are known in the art, those skilled in the art can appropriately select FRs. The FR used in the present invention may be a FR having a native sequence, or may be a FR having a sequence derived from a native sequence by modifying one or more amino acids.

[0208] An overview of the SEQ ID NOs and detailed sequences as used in the context of the present invention is given in the following Table 1 (in case of conflict between the sequences shown in Table 1 and the sequences of the sequence listing which has to be submitted for formal reasons, the sequences of Table 1 replace the sequences of the sequence listing):

[0209]

[0210]

[0211]

[0212]

[0213]

[0214]

[0215]

[0216]

[0217]

[0218]

[0219] In the context of the present invention the following abbreviations are used: VH = variable heavy chain; VH = variable heavy chain; VL = variable light chain; CDR = complementarity determining region; VH-CDR = CDR of the variable heavy region; VL-CDR = CDR of the variable light region; CDRs can be determined by using the Kabat algorithm, for example http: / / abysis.org / abysis / .

[0220] *****

[0221] Note that as used herein, the singular forms "a", "an", and "the" include plural references unless the context clearly dictates otherwise. Thus, for example, reference to "an agent" includes one or more of such different agents, and reference to "the method" includes reference to equivalent steps and methods known to those of ordinary skill in the art, which may be modified or substituted for the methods described herein.

[0222] Throughout the specification and claims herein, the singular encompasses the plural, unless the context requires otherwise. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.

[0223] Unless otherwise indicated, the term "at least" preceding a series of elements is understood to refer to each element in the series. Those skilled in the art will recognize or be able to ascertain, using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. These equivalents are also intended to be encompassed by the present invention.

[0224] Wherever used, the term "and / or" includes the meanings of "and," "or," and "all or any other combination of elements connected by the term."

[0225] The term "less than" or conversely "greater than" does not include specific numbers.

[0226] The term "about" means ±10%, preferably ±5%, more preferably ±2%, most preferably ±1%.

[0227] For example, "less than 20" means less than the indicated number. Similarly, "greater than" or "higher than" means greater than or higher than the indicated number, such as "greater than 80%" means greater than or higher than 80% of the specified number.

[0228] Throughout the specification and in the claims that follow, unless the context requires otherwise, the word "comprise" and variations such as "comprises" and "comprising" are to be understood as meaning the inclusion of stated integers or steps or groups of integers or steps, but not the exclusion of any other integers or steps or groups of integers or steps. When used herein, the term "comprises" may be replaced with the terms "contains" or "includes", or sometimes, when used herein, with the term "having". When used herein, "consisting of excludes any element, step or ingredient not specified.

[0229] The term "including" means "including but not limited to." "Including" and "including but not limited to" are used interchangeably.

[0230] It should be understood that the present invention is not limited to the specific methodology, protocols, materials, reagents and substances, etc. described herein, and thus may vary. The terminology used herein is for the purpose of describing specific embodiments only and is not intended to limit the scope of the present invention, which is limited only by the claims.

[0231] All publications cited throughout this specification (including all patents, patent applications, scientific publications, specifications, etc.), whether above or below, are incorporated by reference in their entirety. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate these disclosures by virtue of prior invention. To the extent that material incorporated by reference is inconsistent or inconsistent with this specification, this specification will supersede any such material.

[0232] The contents of all literature and patent documents cited herein are incorporated by reference in their entirety.

[0233] The present invention and its advantages will be better understood from the following examples, which are provided for illustrative purposes only. These examples are not intended to limit the scope of the present invention in any way.

[0234] Example

[0235] Example 1

[0236] CAR-T cells are often used in cancer treatment attempts. The difficulty in turning CAR-T cells into effective anticancer drugs lies in the nature of tumor-specific surface antigens present on cancer cells. Ideally, these surface antigens are expressed only on cancer cells and not on the somatic cells of the organism. But spontaneous cancers originating from host cells rarely meet this condition. In cancer cells infected and driven by oncogenic viruses, the situation is completely different. EBV is such an oncogenic virus, but EBV-infected tumor cells carry the virus mainly in latent form, so that few viral antigens are expressed on the surface of virus-infected cancer cells. HDAC inhibitors can induce lytic viral programs in these cells, so that some viral glycoproteins are expressed on the cell surface and act as so-called new antigens. These new antigens are different from all host cell antigens and can specifically mark virus-infected cells. The virally encoded glycoprotein gp350 on the surface of EBV-infected cancer cells is a favorable condition for targeting and treating cancers associated with EBV or driven by EBV. Gp350-specific antibodies can target gp350. In the case of the present invention, antibodies are applied in the form of CAR-T cells, resulting in CAR-T cells binding to cancer cells. This binding activates CAR-T cells, which induces cancer cell elimination mediated via the effector function of CAR-T cells and the release of cytokines such as interferon gamma (IFN-γ).

[0237] CAR-T cells based on gp350 antibodies 12C10 and 7A1 induced IFN-γ release when they encountered their corresponding gp350 antigens on EBV-infected cells ( Figure 4 We have generated a new gp350 antibody, 7D11 ( Figure 6 ), which is intended to achieve targeted purposes.

[0238] Example 2

[0239] Treatment of cells with HDAC (histone deacetylase) inhibitors results in activation of the lytic cycle of EBV-infected cells, leading to the expression of lytic viral antigens (such as gp350) on the surface of such cells. The expression of the EBV transcription factor BZLF1 is a hallmark of lytic phase induction, as it functions as a molecular switch gene that can initiate the lytic phase in cells latently infected with EBV ( Figure 2 ). During latency, the viral BZLF1 gene is epigenetically repressed, but repression can be reversed by several drugs that act on chromatin components and their post-translational modifications. HDAC inhibitors are such drugs, and some HDAC inhibitors have been used clinically or preclinically ( Figure 1 ). Valproic acid (VPA) and suberoylanilide hydroxamic acid (SAHA) are examples of HDAC inhibitors. The latter is preferred because it induces the lytic cycle of EBV ( Figure 2), and thus increase gp350 surface expression on EBV-infected cells ( Figure 3 ).

[0240] Example 3

[0241] The use of HDAC SAHA did increase the fraction of cells supporting the EBV lytic phase ( Figure 2 ) and gp350 surface expression. Therefore, SAHA drug treatment will lead to increased targeting of gp350-CAR-T cells ( Figure 5 ), and led to more effective elimination of EBV-positive cancer cells. Therefore, the combined application of gp350-CAR-T cells and HDAC / SAHA is the most promising approach for the treatment of EBV-related cancers.

Claims

1. An HDAC inhibitor for use in the treatment or prevention of a medical condition associated with herpes virus infection, The method comprises further administering an immune cell comprising a chimeric antigen receptor (CAR) on its cell surface, The CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or to a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell infected by the herpes virus.

2. The HDAC inhibitor for use according to claim 1, wherein the HDAC inhibitor induces the expression of herpes virus antigens or wherein the HDAC inhibitor induces the lytic cycle of herpes virus.

3. The HDAC inhibitor for use according to claim 1 or 2, wherein the HDAC inhibitor is selected from butyric acid, valproic acid, MS275 (entinostat), apicidin, largazole, oxamflatin, scriptaid, suberoylanilide hydroxamic acid (SAHA, vorinostat), LBH589 (panobinostat), 12-O-tetradecanoylphorbol-13-acetate (TPA) and PXD101 (belinostat), preferably SAHA, TPA or valproic acid, more preferably SAHA.

4. The HDAC inhibitor for use according to any one of the preceding claims, wherein the medical condition associated with herpes simplex virus infection is (i) herpes virus-associated cancer, chronic active herpes virus infection, reactivated herpes virus infection or primary herpes virus infection; (ii) an EBV-associated cancer selected from a lymphoproliferative disorder (LPD), such as a B-cell lymphoma, including Burkitt's lymphoma (BL), Hodgkin's lymphoma (HL), diffuse large B-cell lymphoma (DLBCL), a T-cell lymphoma, or a post-transplantation lymphoproliferative disorder (PTLD), or an epithelial cell carcinoma (nasopharyngeal carcinoma, lung carcinoma, breast carcinoma), a lymphoepithelioma, a lymphoid stromal carcinoma (GCLS, such as gastric carcinoma) or a glioma; or (iii) chronic active EBV infection (CAEBV) or primary EBV infection (e.g. infectious mononucleosis).

5. The HDAC inhibitor for use according to claim 4(i), wherein the medical condition associated with herpes virus infection is a herpes virus-associated cancer, in which herpes virus antigens are present on the surface of cancer cells.

6. An HDAC inhibitor for use according to any one of the preceding claims, wherein the herpes virus is Epstein-Barr virus (EBV).

7. An HDAC inhibitor for use according to any one of the preceding claims, (i) wherein the herpes virus antigen is involved in the binding of the virus to a receptor on a target human cell; (ii) wherein the herpes virus antigen is an Epstein-Barr virus antigen (EBV antigen).

8. The HDAC inhibitor for use according to claim 7(ii), wherein the EBV antigen is present on the surface of EBV-infected cells, preferably on the surface of EBV-infected cancer cells, EBV-infected B cells or EBV-infected epithelial cells, and / or wherein the EBV antigen is an EBV virion envelope protein or a protein of the EBV envelope complex.

9. An HDAC inhibitor for use according to any one of the preceding claims, wherein the herpes virus antigen is EBV glycoprotein 350 / 220 (gp350 / 220).

10. An HDAC inhibitor for use according to any one of the preceding claims, wherein The immune cells are selected from the group consisting of T lymphocytes, NK cells, macrophages and dendritic cells, wherein the T lymphocytes are preferably cytotoxic T lymphocytes or T helper cells, more preferably cytotoxic T lymphocytes.

11. The HDAC inhibitor for use according to any one of the preceding claims, wherein the CAR comprises or consists of any one of SEQ ID NO: 2, 12, 22 or 32.

12. An immune cell comprising a chimeric antigen receptor (CAR) on its cell surface for use in treating or preventing a medical condition associated with herpes virus infection, The method comprises further administering an HDAC inhibitor, The CAR comprises an antibody or antibody fragment that binds to a protein encoded by a herpes virus or to a protein complex comprising the protein (herpes virus antigen), wherein the herpes virus antigen is present on the surface of a human cell infected by the herpes virus.

13. The HDAC inhibitor or immune cell for use according to any one of the preceding claims, wherein the HDAC inhibitor is administered to the patient before, after or simultaneously with the administration of the immune cells.

14. A kit of parts comprising an immune cell as defined in any one of claims 1 to 12 and an HDAC inhibitor as defined in any one of claims 1 to 12, wherein the immune cell comprises a chimeric antigen receptor (CAR) on its cell surface.

15. A pharmaceutical product comprising an immune cell as defined in any one of claims 1 to 12 and an HDAC inhibitor as defined in any one of claims 1 to 12, wherein the immune cell comprises a chimeric antigen receptor (CAR) on its cell surface.

16. A kit of parts according to claim 14 or a pharmaceutical product according to claim 15 for use in combination in a method for treating or preventing a medical condition associated with a herpes virus infection, wherein the combined use comprises administering the HDAC inhibitor to a patient before the administration of the immune cells, administering the HDAC inhibitor and the immune cells to a patient simultaneously, or administering the immune cells to a patient after the administration of the HDAC inhibitor.

17. An antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody (a) comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in any one of SEQ ID NOs: 3, 13, 23 or 33, and the light chain variable region has an amino acid sequence that is at least 85% identical to the amino acid sequence shown in any one of SEQ ID NOs: 4, 14, 24 or 34; and / or (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

18. An antibody that binds to gp350 of EBV present on the cell surface of a target cell, wherein the antibody (a) comprises a heavy chain variable region comprising a heavy chain CDR1 having an amino acid sequence as shown in any one of SEQ ID NOs: 5, 15, 25 or 35, a heavy chain CDR2 having an amino acid sequence as shown in any one of SEQ ID NOs: 6, 16, 26 or 36, and a heavy chain CDR3 having an amino acid sequence as shown in any one of SEQ ID NOs: 7, 17, 27 or 37, and A light chain variable region comprising a light chain CDR1 having an amino acid sequence as shown in any one of SEQ ID NOs: 8, 18, 28 or 38, a light chain CDR2 having an amino acid sequence as shown in any one of SEQ ID NOs: 9, 19, 29 or 39, and a light chain CDR3 having an amino acid sequence as shown in any one of SEQ ID NOs: 10, 20, 30 or 40; and / or (b) Binds to the same epitope in the EBV protein gp350 as the epitope to which the antibody of (a) binds.

19. A CAR comprising heavy chain and light chain variable regions as defined in claim 17 or heavy chain CDR1, CDR2 and CDR3 as defined in claim 18; and light chain CDR1, CDR2, CDR3.

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