Treatment methods for patients with reduced sensitivity to BCL-2 inhibitors

Abstract

This provides improved treatments for cancers, including CD70-expressing cancers such as myeloid malignancies. [Solution] A CD70-binding antibody or its antigen-binding fragment for use in the treatment of a human subject with myeloid malignancy resistant to BCL-2 inhibitors, and a method for treating the subject's myeloid malignancy, comprising the steps of (a) selecting a human subject with myeloid malignancy having reduced sensitivity to BCL-2 inhibitors or being refractory, and (b) administering the CD70-binding antibody or its antigen-binding fragment to the subject. In one embodiment, the myeloid malignancy is acute myeloid leukemia (AML). In one embodiment, the CD70-binding antibody is xatuzumab. In one embodiment, a BCL-2 inhibitor is co-administered with the CD70-binding antibody or its antigen-binding fragment.

Description

[Technical Field]

[0001] (Sequence Listing) This application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety.

[0002] (Field of the Invention) The present invention relates to therapies including combination therapies for the treatment of cancer, particularly the treatment of relapsed or refractory myeloid malignancies. The therapy is particularly useful for the treatment of acute myeloid leukemia (AML), including monocytic AML. The combination therapy comprises an antibody or antigen-binding fragment thereof that binds CD70, and a BCL-2 inhibitor, such as venetoclax or a pharmaceutically acceptable salt thereof. [Background Art]

[0003] (Background of the Invention) In recent years, the development of new cancer treatments has come to focus on molecular targets involved in cancer progression, particularly proteins. The list of molecular targets involved in tumor growth, invasion and metastasis continues to expand, and the list includes proteins overexpressed by tumor cells, as well as targets associated with systems that support tumor growth such as the vascular system and immune system. The number of therapeutic agents or anticancer agents designed to interact with these molecular targets also continues to increase. Currently, numerous targeted cancer therapeutics have been approved for clinical use, and many more are under development.

[0004] CD70 has been identified as a particularly interesting molecular target due to its constitutive expression in many types of hematological malignancies and solid carcinomas (Junker et al., (2005) J Urol. 173:2150-3; Sloan et al., (2004) Am J Pathol. 164:315-23; Held-Feindt and Mentlein, (2002) Int J Cancer, 98:352-6; Hishima et al., (2000) Am J Surg Pathol. 24:742-6; Lens et al., (1999) Br J Haematol. 106:491-503; Boursalian et al., (2009) Adv Exp Med Biol. 647:108-119; Wajant H., (2016) Expert Opin Ther Targets, 20(8):959-973). CD70 is a type II transmembrane glycoprotein belonging to the tumor necrosis factor (TNF) superfamily, and it mediates its action through binding to its congener cell surface receptor, CD27. Both CD70 and CD27 are expressed by multiple cell types of the immune system, and the CD70-CD27 signaling pathway is involved in regulating several different aspects of the immune response. This reflects the fact that CD70 overexpression occurs in various autoimmune diseases, including rheumatoid and psoriatic arthritis and lupus (Boursalian et al., (2009) Adv Exp Med Biol. 647:108-119; Han et al., (2005) Lupus, 14(8):598-606; Lee et al., (2007) J Immunol. 179(4):2609-2615; Oelke et al., (2004) Arthritis Rheum. 50(6):1850-1860).

[0005] CD70 expression has been linked to poor prognosis in several cancers, including B-cell lymphoma, renal cell carcinoma, and breast cancer (Bertrand et al., (2013) Genes Chromosomes Cancer, 52(8):764-774; Jilaveanu et al., (2012) Hum Pathol. 43(9):1394-1399; Petrau et al., (2014) J Cancer, 5(9):761-764). CD70 expression is also found in metastatic tissue in a high percentage of cases, indicating an important role of this molecule in cancer progression (Jacobs et al., (2015) Oncotarget, 6(15):13462-13475). The constitutive expression of CD70 and its receptor CD27 on hematopoietic tumor cells is thought to be related to the role of the CD70-CD27 signaling center, which directly regulates tumor cell proliferation and survival (Goto et al., (2012) Leuk Lymphoma, 53(8):1494-1500; Lens et al., (1999) Br J Haematol. 106(2):491-503; Nilsson et al., (2005) Exp Hematol. 33(12):1500-1507; van Doorn et al., (2004) Cancer Res. 64(16):5578-5586).

[0006] Upregulated CD70 expression on tumors, particularly solid tumors that do not co-express CD27, also contributes to immunosuppression in the tumor microenvironment in various ways. For example, CD70 binding to CD27 on regulatory T cells (Tregs) has been shown in mice to increase the frequency of Tregs, decrease tumor-specific T cell responses, and promote tumor growth (Claus et al., (2012) Cancer Res. 72(14):3664-3676). As demonstrated in cells of renal cell carcinoma, glioma, and glioblastoma, CD70-CD27 signaling can also weaken the immune response by inducing apoptosis of tumor-induced T lymphocytes (Chahlavi et al., (2005) Cancer Res. 65(12):5428-5438; Diegmann et al., (2006) Neoplasia, 8(11):933-938; Wischusen et al., (2002) Cancer Res, 62(9):2592-2599). Finally, CD70 expression is also associated with T cell depletion, which causes lymphocytes to adopt a more differentiated phenotype and be unable to kill tumor cells (Wang et al., (2012) Cancer Res, 72(23):6119-6129; Yang et al., (2014) Leukemia, 28(9):1872-1884).

[0007] BCL-2 (B-cell lymphoma 2) is an essential mitochondrial outer membrane protein that inhibits apoptotic death in several cells, including lymphocytes. Overexpression of BCL-2 in cancer cells confers resistance to apoptosis, and therefore inhibition of this protein can promote tumor cell death. Recent clinical trials have reported that adding the highly specific BCL-2 inhibitor venetoclax to the current standard of care for acute myeloid leukemia (AML), such as hypomethylating agents (HMA), can significantly improve response rates and overall survival in newly diagnosed AML patients (75 years of age or older or with comorbidities) who are unable to receive intensive induction chemotherapy (Dinardo, (2020), New England Journal of Medicine). These findings have led to recent FDA approval of this regimen for this population, and it is now considered standard care.

[0008] The combination of venetoclax and HMA azacitidine results in a remission rate of approximately 70% for AML. However, a small but significant number of patients do not achieve remission and remain refractory. Furthermore, the majority of patients who achieve remission eventually relapse.

[0009] Therefore, improving therapies is still necessary for the treatment of cancers, including CD70-expressing cancers such as myeloid malignancies. [Overview of the project]

[0010] (Summary of the invention) In a first aspect, the present invention provides an anti-CD70 antibody or a CD70-conjugated fragment thereof for use in the treatment of myeloid malignancies in human subjects resistant to treatment with BCL-2 inhibitors.

[0011] A further aspect of the present invention is a method for treating myeloid malignancies in humans. This method is (a) Selecting human subjects with myeloid malignancies that have reduced sensitivity to BCL-2 inhibitors or are refractory, (b) The step of administering to the human subject an antibody that binds to CD70 or an antigen-binding fragment thereof.

[0012] In one embodiment, the BCL-2 inhibitor is venetoclax or a pharmaceutically acceptable salt thereof.

[0013] In one embodiment, the myeloid malignancy is selected from the group consisting of acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), myeloproliferative neoplasm (MPN), chronic myeloid leukemia (CML), and myelomonocytic leukemia (CMML).

[0014] In one embodiment, the myeloid malignancy is AML.

[0015] In one embodiment, the AML is a monocyte-type AML.

[0016] In one embodiment, the myeloid malignancy is MDS.

[0017] In one embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject.

[0018] In one embodiment, at least one of BCL-2 and CD117 is downregulated, and at least one of CD11b, CD68, CD64, CD70, BCL2A1, and MCL1 is upregulated.

[0019] In one embodiment, step (a) includes determining the CD70 expression level of malignant myeloid cells of the human subject.

[0020] In one embodiment, CD70 is upregulated compared to CD70 expression levels measured before or during treatment with a BCL-2 inhibitor.

[0021] In one embodiment, said human subject has a clinical history comprising (a) treatment with a BCL-2 inhibitor, and (b) absence of remission in response to treatment with said BCL-2 inhibitor.

[0022] In one embodiment, the treatment with a BCL-2 inhibitor in said clinical history further comprises treatment with a hypomethylating agent (HMA).

[0023] In one embodiment, said human subject has a clinical history comprising (a) treatment with a BCL-2 inhibitor, (b) partial or complete remission, and (c) partial or complete relapse.

[0024] In one embodiment, the treatment with a BCL-2 inhibitor in said clinical history further comprises treatment with a hypomethylating agent (HMA).

[0025] In one embodiment, the hypomethylating agent (HMA) is co-administered with the antibody that binds to CD70 or an antigen-binding fragment thereof.

[0026] In one embodiment, the BCL-2 inhibitor is co-administered with the antibody that binds to CD70 or an antigen-binding fragment thereof.

[0027] In one embodiment, the antibody that binds to CD70 or the antigen-binding fragment thereof comprises HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, the amino acid sequence of said HCDR1 consists of SEQ ID NO: 1; the amino acid sequence of said HCDR2 consists of SEQ ID NO: 2; the amino acid sequence of said HCDR3 consists of SEQ ID NO: 3; the amino acid sequence of said LCDR1 consists of SEQ ID NO: 4; the amino acid sequence of said LCDR2 consists of SEQ ID NO: 5; and the amino acid sequence of said LCDR3 consists of SEQ ID NO: 6.

[0028] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 90% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 90% the same amino acid sequence as SEQ ID NO: 8.

[0029] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having the same amino acid sequence as SEQ ID NO: 7 and a variable light chain domain (VL) having the same amino acid sequence as SEQ ID NO: 8.

[0030] In one embodiment, the amino acid sequence identical to VH consisting of sequence number 7 by at least 90% includes HCDR1, HCDR2, and HCDR3. The amino acid sequence of HCDR1 is sequence number 1; The amino acid sequence of HCDR2 consists of sequence number 2; and, The amino acid sequence of HCDR3 consists of sequence number 3; and, The amino acid sequence that is at least 90% identical to the VL consisting of SEQ ID NO: 8 comprises LCDR1, LCDR2, and LCDR3, wherein the amino acid sequence of LCDR1 consists of SEQ ID NO: 4; The amino acid sequence of LCDR2 consists of sequence number 5; and the amino acid sequence of LCDR3 consists of sequence number 6.

[0031] In one embodiment, the HMA is selected from the group consisting of azacitidine, decitabine, and guadecitabine.

[0032] In one embodiment, the BCL-2 inhibitor is venetoclax or a pharmaceutically acceptable salt thereof.

[0033] In one embodiment, the antibody that binds to CD70 is xatuzumab.

[0034] One aspect of the present invention is a method for identifying and treating a patient who should be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy, and the method is: (i) To measure the differentiation status of the patient's myeloid cells, (ii) Determine whether the patient has differentiated monocytic AML, and identify patients with differentiated monocytic AML as patients who should be treated with the anti-CD70 antibody or its CD70-binding fragment, and (iii) The method comprising the step of administering the anti-CD70 antibody or its CD70 conjugate fragment to a patient identified as a patient to be treated with the anti-CD70 antibody or its CD70 conjugate fragment.

[0035] One aspect of the present invention is an anti-CD70 antibody or a CD70-conjugated fragment thereof for use in the treatment of myeloid malignancies in patients resistant to treatment with BCL-2 inhibitors.

[0036] A further aspect of the present invention is a CD70-binding antibody or its antigen-binding fragment for use in the treatment of myeloid malignancies in patients resistant to treatment with BCL-2 inhibitors.

[0037] In one embodiment, the patient has previously received treatment with a BCL-2 inhibitor or treatment with a BCL-2 inhibitor and a hypomethylating agent (HMA).

[0038] In one embodiment, the myeloid malignancy is selected from acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), myeloproliferative neoplasm (MPN), chronic myeloid leukemia (CML), and myelomonocytic leukemia (CMML).

[0039] In one embodiment, the myeloid malignancy is AML or MDS.

[0040] In one embodiment, the patient is identified as having differentiated monocyte-type AML based on different expression levels.

[0041] In one embodiment, before the treatment, (i) Measuring the differentiation state of the patient’s bone marrow cells, and, (ii) A selection is made which includes the step of determining whether the patient has differentiated monocytic AML, A therapeutically effective dose of the anti-CD70 antibody or its anti-CD70 conjugate fragment is administered to a patient having differentiated monocytic AML.

[0042] In one embodiment, the patient is identified as having differentiated monocytic AML based on differential expression levels of at least one monocyte marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1.

[0043] In one embodiment, the patient exhibits downregulated expression of at least one of the BCL-2 and CD117, and upregulated expression of at least one of the CD11b, CD68, CD64, BCL2A1, and MCL1.

[0044] In one embodiment, the human subject is (a) Treatment with BCL-2 inhibitors, and, (b) Absence of remission in response to treatment with the BCL-2 inhibitor, He has a clinical history that includes [this].

[0045] In one embodiment, the treatment with the BCL-2 inhibitor in the clinical history further includes treatment with a hypomethylating agent (HMA).

[0046] In one embodiment, the human subject is (a) Treatment with BCL-2 inhibitors, (b) Partial or complete remission, (c) Partial or complete relapse, He has a clinical history that includes [this].

[0047] In one embodiment, the treatment with the BCL-2 inhibitor in the clinical history further includes treatment with a hypomethylating agent (HMA).

[0048] In one embodiment, a hypomethylating agent (HMA) is co-administered with an antibody or its antigen-binding fragment that binds to CD70.

[0049] In one embodiment, a BCL-2 inhibitor is co-administered with an antibody or its antigen-binding fragment that binds to CD70.

[0050] In one embodiment, a BCL-2 inhibitor and a hypomethylating agent (HMA) are co-administered with an antibody or its antigen-binding fragment that binds to CD70.

[0051] In one embodiment, the CD70 expression level in the patient is measured.

[0052] In one embodiment, CD70 is upregulated compared to CD70 expression levels measured before or during treatment with a BCL-2 inhibitor.

[0053] In one embodiment, the BCL-2 inhibitor-resistant patient is a patient who is relapsed or refractory to BCL-2 inhibitors.

[0054] In one embodiment, the BCL-2 inhibitor is venetoclax or a pharmaceutically acceptable salt thereof.

[0055] In one embodiment, the low-methylating agent (HMA) is azacitidine, decitabine, or guadecitabine.

[0056] In one embodiment, the patient is resistant to combination therapy with a BCL-2 inhibitor and HMA.

[0057] In one embodiment, the patient is resistant to the combination therapy of venetoclax and azacitidine.

[0058] In one embodiment, the anti-CD70 antibody or its CD70-binding fragment comprises a variable heavy chain domain (VH) and a variable light chain domain (VL), wherein the VH domain and VL domain are represented by the following CDR sequence: HCDR1 consisting of sequence number: 1; HCDR2 consisting of sequence number: 2; HCDR3 consisting of sequence number: 3; LCDR1 consisting of sequence number 4; LCDR2 consisting of sequence number: 5; Includes LCDR3, which has sequence number 6.

[0059] In one embodiment, the anti-CD70 antibody or its anti-CD70 conjugate fragment comprises a variable heavy chain domain (VH) consisting of or at least 90% identical to SEQ ID NO: 7, and a variable light chain domain (VL) consisting of or at least 90% identical to SEQ ID NO: 8.

[0060] In one embodiment, the amino acid differences in the amino acid sequence that is at least 90% identical to that of VH consisting of sequence number 7 do not exist in the CDR sequence of VH, and the amino acid differences in the amino acid sequence that is at least 90% identical to that of VL consisting of sequence number 8 do not exist in the CDR sequence of VL.

[0061] In one embodiment, the anti-CD70 antibody is xatuzumab.

[0062] In one embodiment, the hypomethylating agent (HMA) is co-administered with the anti-CD70 antibody or its anti-CD70 conjugated fragment.

[0063] In one embodiment, the BCL-2 inhibitor is co-administered with the anti-CD70 antibody or its CD70-binding fragment.

[0064] One aspect of the present invention is a method for identifying a patient to be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy, and the method is: (i) to measure the differentiation status of the patient's myeloid cells, and, (ii) Determining whether the patient has differentiated monocytic AML, The method involves identifying patients with differentiated monocytic AML as patients who should be treated with the anti-CD70 antibody or its CD70-binding fragment.

[0065] In one embodiment, steps (i) and (ii) are performed with a sample obtained from a patient with the myeloid malignancy.

[0066] In one embodiment, the patient's bone marrow sample contains CD45bright / SSChigh / CD38+ / CD34- / CD33+ / CD11b+ / CD70+ phenotype cells or CD45bright / SSChigh / CD34- / CD117- / CD11b+ / CD68+ / CD14+ / CD64+ phenotype cells. [Brief explanation of the drawing]

[0067] (Brief explanation of the drawing) [Figure 1]Figure 1A shows the treatment history and flow analysis of bone marrow (BM) specimens at the time of diagnosis for patient Pt-51. In the CD45 / SSC plot, the Mono, Prim, and Lym gates represent subpopulations of monocytes, primitives, and lymphocytes, respectively. The CD34 / CD117 and CD68 / CD11b plots show the immunophenotypes of the gated primitive subpopulations. Arrows highlight the target population. Figure 1 is adapted from the literature by Pei et al. (2020). Figure 1B shows the treatment history and flow analysis of bone marrow (BM) specimens at the time of diagnosis for patient Pt-72. In the CD45 / SSC plot, the Mono, Prim, and Lym gates represent subpopulations of monocytes, primitives, and lymphocytes, respectively. The CD34 / CD117 and CD68 / CD11b plots show the immunophenotypes of the gated monocyte subpopulations. Arrows highlight the target population. Figure 1 is adapted from the literature by Pei et al. (2020). Figure 1C is a violin plot showing the median fluorescence intensity (MFI) of CD117, CD11b, CD68, and CD64 in mono-AML (N=5) and prim-AML (N=7) quantified by flow cytometry analysis. Each dot represents a unique AML. The Mann-Whitney test was used for significance determination. Figure 1 is adapted from the literature by Pei et al. (2020). Figure 1D shows the survival rates of low-ROS LSCs sorted from mono-AML (N=5) and prim-AML (N=7) after 24-hour in vitro treatment with VEN alone or in combination with a fixed dose of 1.5 μmol / L AZA. Mean ± SD of technical triplicates. All survival data were normalized to untreated (UNT) controls. VEN: Venetoclax. AZA: Azacitidine. Figure 1 is adapted from the literature by Pei et al. (2020).

[0068] [Figure 2]Figure 2A is a bar graph showing BCL2 expression in low-ROS prim-AML (N=7) and low-ROS mono-AML (N=5). Each dot represents a unique AML. Mean ± SD. Figure 2 is adapted from Pei et al. (2020). Figure 2B is a bar graph showing BCL2 expression in the FAB-M0 (N=16), M1 (N=44), M2 (N=40), M0 / 1 / 2 (N=100), and M5 (N=21) subclasses of AML from the TCGA (The Cancer Genome Atlas) dataset. Each dot represents a unique AML. Figure 2 is adapted from Pei et al. (2020). Figure 2C is an image of Western blot results showing BCL-2 protein-level expression in prim-AML (N=5) and mono-AML (N=4). Actin is used as a loading control. Figure 2 is adapted from the literature by Pei et al. (2020).

[0069] [Figure 3] Figure 3A shows the treatment history of patient Pt-12 and a flow analysis of its diagnostic (Dx) and relapse (Rl) specimens. In the CD45 / SSC plot, the Mono, Prim, and Lym gates identify the monocyte, early-stage, and lymphocyte populations, respectively. The CD34 / CD117 and CD68 / CD11b plots show the immunophenotypes of the gated early-stage subpopulation (P-AML) and monocyte subpopulation (M-AML). Arrows highlight the target population in the CD45 / SSC plot. Figure 3 is adapted from the literature by Pei et al. (2020). Figure 3B shows the treatment history of patient Pt-65 and a flow analysis of its diagnostic (Dx) and relapse (Rl) specimens. In the CD45 / SSC plot, the Mono, Prim, and Lym gates identify the monocyte, early-stage, and lymphocyte populations, respectively. The CD34 / CD117 and CD68 / CD11b plots show the immunophenotypes of the gated early-stage subpopulation (P-AML) and the monocyte subpopulation (M-AML). The arrows highlight the target population, particularly the monocyte subpopulation, in the CD45 / SSC plot. Figure 3 is adapted from Pei et al., (2020).

[0070] [Figure 4] Figure 4 is a bar graph showing CD70 mRNA expression levels in the FAB-M0 (N=13), M1 (N=39), M2 (N=37), M3 (N=14), M4 (N=32), and M5 (N=18) subclasses of AML. Patients with the highest CD70 expression belong to the M5 subtype, which contains more than 80% monocytic AML cells in the bone marrow.

[0071] [Figure 5] Figure 5 shows flow cytometry analysis of bone marrow samples from VEN+AZA refractory monocytic disease (A) and VEN+AZA refractory disease (B) including a mixed phenotype of monocytic and early-type AML cells. Gating of monocytic cells with CD34, CD11b, CD14, and CD64 results in higher levels of CD70 expression in monocytic AML cells, in contrast to early-type cells (A and B). Early-type cells also show CD70 expression (B).

[0072] [Figure 6] Figure 6A shows a bar graph (left) comparing the median fluorescence intensity (MFI) of CD70 in early-type AML cells and monocyte-type AML cells, and a paired expression analysis for each sample (right) showing that monocyte-type AML cells within the same patient sample exhibit higher CD70 expression levels than early-type AML cells. Figure 6B shows a bar graph (left) comparing the percentage of CD70-positive early-type AML cells and monocyte-type AML cells, and a paired analysis for each sample (right) showing that CD70-positive malignant cells exhibit a higher percentage of CD70 expression in the monocyte-type AML cell population. CD70 expression levels are higher in monocyte-type malignant AML cells than in early-type AML cells.

[0073] [Figure 7]Figure 7A shows flow cytometry analysis of mixed phenotype and monocytic AML samples used to evaluate NK-dependent killing by xatuzumab. In both samples, the monocytic CD38+ / CD33+ / CD11b+ subpopulation expressed high levels of CD70 on the plasma membrane. Figure 7B is a bar graph showing the effects on monocytic and early-stage AML cells after administration of xatuzumab, 41D12 FcDead antibody, and vehicle control. Xatuzumab can significantly mediate NK-dependent cell killing in monocytic and early-stage AML cells of VEN+AZA-sensitive mixed phenotype AML. A one-way ANOVA test was used to determine significance. *p<0.05. Figure 7C is a bar graph showing the effects on monocytic AML cells after administration of xatuzumab, 41D12 FcDead antibody, and vehicle control. Xatuzumab can significantly mediate NK-dependent cell killing in VEN+AZA-resistant monocytic AML cells. A one-way ANOVA test was used to determine significance. ***p<0.001.

[0074] [Figure 8] Figure 8 is a bar graph showing median CD70 expression from transcriptome analyses of gene expression performed in early-type and monocyte-type low-ROS LSCs from bone marrow-derived AML samples. Asymmetric Wilcoxon tests were used to compare both LSC subpopulations. *p<0.05.

[0075] [Figure 9]Figure 9 is a bar graph showing the effect of antibody therapy on leukemia stem cells from CD70-positive, VEN+AZA-resistant monocytic AML bone marrow samples. Data were normalized to isotype control, blocking anti-CD70 antibody 41D12 FcDead, and antibody-free control colony-forming units (CFUs) for xatuzumab. VEN+AZA-resistant monocytic AML bone marrow samples were incubated with NK cells (1:5 T:E ratio) in the presence of antibody (10 μg / ml), and then cultured in CFU medium to confirm whether LSCs were also efficiently targeted by xatuzumab-mediated NK-dependent ADCC. Only xatuzumab, and not the control or blocking antibody, was able to significantly reduce the number of LSCs that colonized in the medium. The plot shows data from triplicate independent studies in three different AML bone marrow samples of VEN+AZA-resistant monocytic AML. One-way ANOVA was used to determine significance. ****p<0.0001.

[0076] [Figure 10] Figure 10 is a bar graph showing the efficacy of anti-CD70 antibody therapy in the presence of NK cells in a patient-derived xenograft mouse model. NSGS mice were transplanted with VEN+AZA-resistant monocytic AML bone marrow samples. When the graft survival level in the bone marrow reached approximately 25% of the leukemia cells, the animals were treated with xatuzumab or VEN+AZA, with or without a single infusion of 1.5 × 10⁶ NK cells, every three days for three doses. After nine days, the animals were sacrificed, bone marrow was isolated from the femur, and the number of malignant monocytic cells in the bone marrow was measured. Unlike xatuzumab alone, VEN+AZA, and VEN+AZA combined with NK cells, animals treated with xatuzumab combined with NK cells showed a significant reduction in the level of monocytic AML cells in the mouse bone marrow. The Mann-Whitney test was used to determine significance. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. [Modes for carrying out the invention]

[0077] (Detailed explanation) (A.Definition) Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art of the present invention.

[0078] Acute Myeloid Leukemia – As used herein, “acute myeloid leukemia” or “AML” refers to a hematopoietic neoplasm involving myeloid cells. AML is characterized by clonal proliferation of bone marrow progenitor cells with reduced differentiation potential. Patients with AML present with an accumulation of blast cells in the bone marrow. As used herein, “blast cells” or simply “blasts” refers to clonal bone marrow progenitor cells exhibiting disrupted differentiation potential. Blast cells also typically accumulate in the peripheral blood of AML patients. AML is usually diagnosed when a patient shows 20% or more blast cells in the bone marrow or peripheral blood. As used herein, the terms “patient” and “human subject” are used interchangeably.

[0079] According to the World Health Organization (WHO) classification scheme, AML generally encompasses the following subtypes: AML with recurrent genetic abnormalities; AML with myelodysplastic-associated changes; treatment-associated myeloid neoplasms; myelosarcoma; Down syndrome-associated myeloproliferative disorders; blastic plasmacytoid dendritic cell neoplasms; and AML unclassified (e.g., acute megakaryoblastic leukemia, acute basophilic leukemia).

[0080] AML can also be categorized according to the French-American-British (FAB) classification system, and includes the subtypes M0 (acute myeloblastic leukemia, most undifferentiated); M1 (acute myeloblastic leukemia, undifferentiated); M2 (acute myeloblastic leukemia, granulocyte mature); M3 (promyelocytic or acute promyelocytic leukemia (APL)); M4 (acute myelomonocytic leukemia); M4eo (myelomonocytic leukemia with myeloeosinophilia); M5 (acute monoblastic leukemia (M5a) or acute monocytic leukemia (M5b)); M6 (acute erythroleukemia, including erythroleukemia (M6a) and the very rare pure erythroleukemia (M6b)); or M7 (acute megakaryoblastic leukemia).

[0081] "Antibody" - As used herein, the term "antibody" is intended to encompass full-length antibodies and their variants, and includes, but is not limited to, modified antibodies, humanized antibodies, and germline antibodies. The term "antibody," as commonly used herein, refers to an immunoglobulin polypeptide having a combination of two heavy chains and two light chains, the polypeptide having significant specific immune response activity against the antigen of interest (CD70 in this specification). For IgG class antibodies, this antibody contains two identical polypeptide light chains with a molecular weight of approximately 23,000 daltons, and two identical heavy chains with a molecular weight of 53,000 to 70,000. These four chains are linked by disulfide bonds in a "Y" configuration, with the light chains bracketing the heavy chains from the opening of the "Y" over a variable region. The light chains of antibodies are classified as either kappa or lambda (κ, λ). Each heavy chain class may be conjugated to either a kappa light chain or a lambda light chain. Generally, light and heavy chains are covalently linked to each other, and the "tails" of these two heavy chains are linked to each other by disulfide covalent bonds, or by non-covalent bonds if the immunoglobulin is produced by a hybridoma, B cell, or genetically modified host cell. In the heavy chain, the amino acid sequence continues from the N-terminus at the end of the Y-shaped branch to the C-terminus at the bottom of each chain.

[0082] Those skilled in the art will understand that heavy chains are classified as gamma, mu, alpha, delta, or epsilon (γ, μ, α, δ, ε), with several subclasses (e.g., γ1-γ4). The properties of this chain determine the “class” of an antibody, respectively, as IgG, IgM, IgA, IgD, or IgE. Immunoglobulin subclasses (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1, etc., are well-characterized and known to provide functional specialization. As used herein, the term “antibody” encompasses antibodies derived from any class or subclass of an antibody.

[0083] "Antigen-binding fragment" - As used herein, the term "antigen-binding fragment" refers to a portion or part of a full-length antibody, or an antibody chain containing fewer amino acid residues than an intact or complete antibody, while maintaining antigen-binding activity. Antigen-binding fragments of antibodies include peptide fragments that exhibit specific immunoreactive activity against the same antigen (e.g., CD70). As used herein, the term "antigen-binding fragment" is intended to encompass antibody fragments selected from antibody light chain variable domains (VL); antibody heavy chain variable domains (VH); single-chain antibodies (scFv); F(ab')2 fragments; Fab fragments; Fd fragments; Fv fragments; single-arm (monovalent) antibodies; diabodies, triabodies, tetrabodies, or any antigen-binding molecules formed by combinations, assemblies, or complexes of such antigen-binding fragments. As used herein, the term "antigen-binding fragment" may also encompass antibody fragments selected from the group consisting of unibodies, domain antibodies, and nanobodies. The fragments can be obtained, for example, by chemical or enzymatic treatment of intact or complete antibodies or antibody chains, or by recombinant means.

[0084] "BCL-2" - As used herein, "BCL-2" or "BCL-2 protein" or "BCL2" refers to the first member of the BCL-2 protein family identified in humans, namely B-cell lymphoma 2. The cDNA encoding human BCL-2 was cloned in 1986, and the important role of this protein in inhibiting apoptosis was revealed in 1988. BCL-2 has been confirmed to be upregulated in several different types of cancer. For example, BCL-2 is activated by t(14;18) chromosomal translocation in follicular lymphoma. Amplification of the BCL-2 gene has also been reported in different cancers, including leukemia (e.g., CLL), lymphoma (e.g., B-cell lymphoma), and several solid tumors (e.g., small cell lung cancer). Human BCL-2 is encoded by the BCL2 gene (UniProtKB-P10415) and has amino acid sequences shown as NCBI reference sequences NP_000624.2 and NP_000648.2.

[0085] "BCL-2 Family" - As used herein, the term "BCL-2 Family" or "BCL-2 Protein Family" refers to a collection of pro-apoptosis and anti-apoptosis proteins associated with BCL-2; see Delbridge et al., (2016) Nat Rev Cancer. 16(2):99-109. At least 16 members of this family exist, classified into three functional groups: (i) BCL-2-like proteins (e.g., BCL-2, BCL-XL / BCL2L1, BCLW BCL2L2, MCL2, BFL1 / BCL2A1); (ii) BAX and BAK; and (iii) BH3 monoproteins (e.g., BIM, PUMA, BAD, BMF, BID, NOXA, HRK, BIK). The BCL-2 protein family, along with its apoptosis-inhibiting members (e.g., BCL-2, BCL-XL), plays a crucial role in regulating essential apoptotic pathways, typically antagonizing pro-apoptotic members (e.g., BAX and BIM). De-regulation of BCL-2 family members, for example, through gene translocation, amplification, overexpression, and mutation, has been observed in many cancers. The downstream effect of this de-regulation is often apoptosis resistance, thereby promoting cancer growth.

[0086] "BCL2A1"—as used herein, "BCL2A1" or B-cell lymphoma 2-associated protein A1 refers to the apoptosis-inhibiting BCL2 protein, which exerts important survival-promoting functions. BCL2A1 has been reported to be upregulated in AML and associated with venetoclax resistance. Zhang et al. (2020) Nat Cancer 1:826-839.

[0087] "BCL-2 inhibitor" - As used herein, a BCL-2 inhibitor means any agent, compound, or molecule capable of specifically inhibiting the activity of BCL-2, in particular an agent, compound, or molecule capable of inhibiting the apoptosis inhibitory activity of BCL-2. Examples of BCL-2 inhibitors suitably used in the combinations described herein include B-cell lymphoma homology 3 (BH3) mimetic compounds (Merino et al., (2018) Cancer Cell. 34(6):879-891). Specific BCL-2 inhibitors are not limited to those listed below, but include venetoclax, ABT-737 (Oltersdorf, T et al., (2005) Nature 435:677-681), navitoclax / ABT-263 (Tse, C et al., (2008) Cancer Res. 68:3421-3428), BM-1197 (Bai, L et al., (2014) PLoS ONE 9:e99404), S44563 (Nemati, F et al., (2014) PLoS ONE 9:e80836), BCL2-32 (Adam, A et al., (2014) Blood 124:5304), and AZD4320 (Hennessy, EJ et al., (2015) ACS Medicinal This includes the Chemistry annual meeting (https: / / www.acsmedchem.org / ama / orig / abstracts / mediabstractf2015.pdf abstr. 24), and S55746 (International Standard Randomized Controlled Trial Registry Number (ISRCTN) http: / / www.isrctn.com / ISRCTN04804337 (2016)). Further examples of BCL-2 inhibitors are described in the literature by Ashkenazi, A et al., (2017) Nature Reviews Drug Discovery 16:273-284, which is incorporated herein by reference.

[0088] "CD70" - As used herein, the terms "CD70," "CD70 protein," or "CD70 antigen" are interchangeable and refer to a member of the TNF ligand family that is a ligand for TNFRSF7 / CD27. CD70 is also known as CD27L or TNFSF7. The terms "human CD70," "human CD70 protein," or "human CD70 antigen" are interchangeable and refer specifically to the native human CD70 protein naturally expressed in the human body and / or on the surface of cultured human cell lines, as well as human homologs including recombinant forms and fragments thereof. Specific examples of human CD70 include the amino acid sequence shown in NCBI reference sequence deposit number NP_001243, or polypeptides having its extracellular domain.

[0089] "Xatuzumab" - as used herein, "xatuzumab" is a monoclonal IgG1 anti-CD70 antibody, also known as ARGX-110. ARGX-110 has been shown to inhibit the interaction of CD70 with its receptor CD27 (Silence et al., (2014) MAbs. Mar-Apr;6(2):523-32, incorporated herein by reference). In particular, ARGX-110 has been shown to inhibit CD70-induced CD27 signaling. The level of CD27 signaling can be determined, for example, by measuring serum soluble CD27 as described in Riether et al. (J. Exp. Med. (2017) 214(2);359-380) or by measuring IL-8 expression as described in Silence et al. (MAbs (2014) 6(2):523-32). Although not constrained by theory, inhibition of CD27 signaling is thought to reduce Treg cell activation and / or proliferation, thereby decreasing the inhibition of antitumor effector T cells. AGRX-110 has also been demonstrated to deplete CD70-expressing tumor cells. In particular, ARGX-110 has been shown to lyse CD70-expressing tumor cells and increase antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) of CD70-expressing cells (Silence et al., cited above). ARGX-110 is defucosylated to enhance ADCC.

[0090] Table 1 shows the amino acid sequences of the six CDR, VH, and VL molecules of ARGX-110 or xatuzumab. (Table 1) [Table 1]

[0091] "Staging of AML" - Most cancers are staging based on tumor size and extent. The staging of AML is often characterized by the number and accumulation of leukemia cells in other organs, such as the liver or spleen. The staging, or progression, of AML is an important factor in evaluating treatment options. The response to BCL-2 inhibitors (or BCL-2 inhibitors and hypomethylating agents) in AML patients correlates closely with the stage of progression; early-stage AML is susceptible, while monocytic AML or "differentiated monocytic AML" (this term is used interchangeably in this text) is more resistant to treatment with BCL-2 inhibitors. Primary AML cells have different properties from more differentiated monocytic AML cells and therefore show different responses to therapy. The expression of monocytic markers can help distinguish primary AML cells from monocytic AML cells. Such monocyte-type markers include BCL-2, CD117, CD11b, CD68, CD64, CD70, BCL2A1, MCL1, and other markers. Non-limiting examples of other monocyte-type markers include CD38, CD34, CD33, and CD14. Monocyte-type AML cells can also be characterized as CD45 bright cells and SSC high cells. The gene expression levels of these monocyte-type markers are more upregulated or more downregulated in monocyte-type tumor cells depending on the stage of AML progression. Myeloid differentiation status correlates with decreased BCL2 expression in AML patients. Therefore, more differentiated monocyte-type AML is very likely to be more refractory to BCL-2 inhibitor-based therapy.

[0092] "Downregulated expression level" - as used herein, "downregulated expression level" refers to a reduced expression level. This means that the expression level of the monocyte marker is trending downward. A downregulated expression level of a monocyte marker is an expression level that is lower than the previous expression level. The previous expression level may be the expression level measured in a patient before or during treatment with a BCL-2 inhibitor (or before or during treatment with a BCL-2 inhibitor and a hypomethylating agent). The previous expression level may also be the baseline expression level of the monocyte marker on monocyte tumor cells.

[0093] "Clinical history treatment" - as used herein, "clinical history treatment" means a prior treatment, for example, a prior treatment with an antibody that binds to CD70 or an antigen-binding fragment thereof.

[0094] "Leukemia stem cells"—as used herein, "leukemia stem cells" or "LSCs" are a subset of blast cells associated with AML. LSCs are blast cells with stem cell characteristics that, when transplanted into immune-deficient recipients, may initiate leukemic disease. LSCs are self-renewing by inducing leukemia, but can also partially differentiate into non-LSC type conventional blast cells that resemble the original disease but are not self-renewing. LSCs occur at a frequency ranging from 1 in 10,000 to 1 in 1,000,000 as a proportion of primary AML blast cells (Pollyea and Jordan, (2017), Blood, 129:1627-1635, incorporated herein by reference). LSCs may also be characterized as cells that are CD34+, CD38-, and optionally CD45- and / or CD123+. LSCs may also be characterized as CD45dim, SSClow, CD90+CD34+ cells.

[0095] "Myeloid malignancies" - as used herein, the term "myeloid malignancies" refers to any clonal disease of hematopoietic stem cells or progenitor cells. Myeloid malignancies or myeloid diseases include chronic and acute forms. Chronic forms include myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPNs), and chronic myelomonocytic leukemia (CMML), while acute forms include acute myeloid leukemia (AML).

[0096] "NK-dependent ADCC" - As used herein, "NK-dependent antibody-dependent cell-mediated cytotoxicity (ADCC)" is an adaptive immune response mediated by natural killer (NK) cells. NK-dependent ADCC is initiated by the activation of NK cells by an antibody. NK-dependent ADCC may also be initiated by the activation of NK cells by an anti-CD70 antibody.

[0097] "Resistance" - As used herein, the terms "resistance" or "resistance" refer to a decrease in sensitivity to treatment in human subjects, such as resistance to a therapy, e.g., BCL-2 inhibitor therapy. The term "resistance" includes initial resistance to therapy or relapse after an initial response to therapy. When a patient relapses, it means that the patient initially responded to therapy but ultimately relapsed, resulting in the patient no longer showing a positive response to treatment. The term "resistance" also includes refractory patients, as well as relapsed patients. A refractory response means that the patient shows no response at all to the treatment given. Patients who do not achieve remission are refractory.

[0098] "Standard intensification chemotherapy"—as used herein, the term "standard intensification chemotherapy" (also referred herein as "intensification induction therapy" or "induction therapy") refers to the so-called "7+3" induction chemotherapy, characterized by seven days of high-dose cytarabine followed by three days of anthracycline (e.g., daunorubicin or idarubicin). Standard intensification chemotherapy may be administered to eligible, newly diagnosed AML patients with the aim of inducing complete remission of AML, typically after successful chemotherapy, with the intention of performing stem cell transplantation on the patient. Not all newly diagnosed AML patients are eligible for standard intensification chemotherapy as described herein.

[0099] "Upregulated expression level" - As used herein, the term "upregulated expression level" refers to an increased or higher expression level. This means an upward trend in the expression level of a monocyte marker. An upregulated expression level of a monocyte marker is a higher expression level compared to a prior expression level. The prior expression level may be the expression level measured in patients before or during treatment with a BCL-2 inhibitor (or before or during treatment with a BCL-2 inhibitor and a hypomethylating agent). The prior expression level may also be the baseline expression level of the monocyte marker on monocyte tumor cells.

[0100] "Venetoclax" - As used herein, the term "venetoclax" refers to a compound having the following chemical structure: [ka]

[0101] Venetoclax is a potent, selective, and orally bioavailable inhibitor of the BCL-2 protein. Its empirical formula is C45H50C1N7O7S and its molecular weight is 868.44. Venetoclax has very low water solubility. Chemically, venetoclax can be expressed as 4-(4-{[2-(4-chlorophenyl)-4,4dimethylcyclohexa-1-en-1-yl]methyl}piperazine-1-yl)-N-({3-nitro-4-[(tetrahydro-2H-pyran-4ylmethyl)amino]phenyl}sulfonyl)-2-(1H-pyrrolo[2,3-b]pyridine-5-yloxy)benzamide). Other names for venetoclax include ABT-199; chemical name 1257044-40-8; and GDC-0199.

[0102] Venetoclax was approved in 2015 by the U.S. Food and Drug Administration (FDA) for the treatment of adult patients with chronic lymphocytic leukemia (CLL) or small lymphocytic leukemia (SLL) who had previously received at least one course of therapy. Venetoclax is distributed and marketed by AbbVie Inc. under the trade name Veneclexta®. Venetoclax is also approved in the United States for use in combination with azacitidine, decitabine, or low-dose cytarabine for the treatment of newly diagnosed acute myeloid leukemia (AML) in adults aged 75 years or older, or in adults with comorbidities that prevent the use of intensive induction chemotherapy.

[0103] (B. Methods) The BCL-2 protein is an apoptosis inhibitory member of the BCL-2 family and is upregulated in many different types of cancer. Overexpression of BCL-2 allows tumor cells to evade apoptosis by capturing pro-apoptotic proteins. BCL-2 is highly expressed in many hematological malignancies and is a major pro-survival protein in diseases such as chronic lymphocytic leukemia (CLL), follicular lymphoma, and mantle cell lymphoma. Inhibiting BCL-2 involves inhibiting the apoptosis-inhibiting or pro-survival activity of this protein.

[0104] Apoptosis inhibitory members of the BCL-2 family, including BCL-2, have been reported to be overexpressed in AML primary samples (Bogenberger et al., (2014) Leukemia 28(2); 1657-65). Overexpression of BCL-2 has also been reported in leukemia stem cells (LSCs) obtained from AML patients (Lagadinou et al., (2013) Cell Stem Cell 12(3); 329-341). Inhibition of BCL-2 in an ex vivo LSC population led to the selective eradication of quiescent LSCs (Lagadinou et al., (2013) Cell Stem Cell 12(3); 329-341).

[0105] While not limiting its scope by theory, the method of the present invention is considered particularly effective for the treatment of AML due to the therapeutic effect of combining a CD70 antibody or its antigen-binding fragment with a BCL-2 inhibitor, especially the combined effect at the LSC level. The self-renewal ability of LSCs means that the persistence of these cells is a major factor contributing to disease relapse.

[0106] Furthermore, the inventors surprisingly found an increased proportion of monocytic AML cells in patients with myeloid malignancies refractory to treatment with the BCL-2 inhibitor venetoclax and the hypermethylating agent (HMA) azacitidine. The presence of monocytic AML cells was found to increase the risk of disease recurrence. The inventors also discovered that monocytic AML cells express significantly higher CD70 levels compared to less differentiated, early-stage AML cells. Venetoclax and azacitidine-resistant monocytic AML cells were sensitive to treatment with anti-CD70 antibodies both in vitro and in vivo in mouse disease models. While not theoretically limiting, it was hypothesized that monocytic AML cells are targeted by anti-CD70 antibody-mediated NK cell-dependent ADCC. Therefore, the anti-CD70 antibodies described herein are considered particularly effective for treating myeloid malignancies resistant to BCL-2 inhibitors such as venetoclax. In one aspect, the present invention provides a method for treating myeloid malignancies in human subjects. The method is particularly useful in treating human subjects with myeloid malignancies that have reduced sensitivity to or are refractory to BCL-2 inhibitors such as venetoclax. The method comprises the steps of (a) selecting a human subject with a myeloid malignancy that has reduced sensitivity to or is refractory to BCL-2 inhibitors, and (b) administering to the human subject an antibody that binds to CD70 or an antigen-binding fragment thereof.

[0107] In one embodiment, human subjects have failed to respond to treatment of myeloid malignancies with BCL-2 inhibitors. In another embodiment, human subjects have shown a clinical response to treatment of myeloid malignancies with BCL-2 inhibitors, but subsequently experienced relapse of myeloid malignancies. The clinical response may be any clinical response, including complete response, partial response, or minimal response. In one embodiment, human subjects have shown a clinical response to treatment of myeloid malignancies with BCL-2 inhibitors, but subsequently experienced a decline in their clinical response to said BCL-2 inhibitors. In another embodiment, human subjects have shown a clinical response to treatment of myeloid malignancies with BCL-2 inhibitors, but subsequently became refractory to treatment with said BCL-2 inhibitors. In yet another embodiment, human subjects have not shown a significant clinical response to treatment of myeloid malignancies with BCL-2 inhibitors.

[0108] In some embodiments, human subjects have failed to treat myeloid malignancies with venetoclax or a pharmaceutically acceptable salt thereof. In some embodiments, human subjects have shown a clinical response to treatment of myeloid malignancies with venetoclax or a pharmaceutically acceptable salt thereof, but have subsequently experienced relapse of myeloid malignancies. The clinical response may be any clinical response, including complete response, partial response, or minimal response. In some embodiments, human subjects have shown a clinical response to treatment of myeloid malignancies with venetoclax or a pharmaceutically acceptable salt thereof, but have subsequently experienced a decline in their clinical response to venetoclax or a pharmaceutically acceptable salt thereof. In some embodiments, human subjects have shown a clinical response to treatment of myeloid malignancies with venetoclax or a pharmaceutically acceptable salt thereof, but have subsequently become refractory to treatment with venetoclax or a pharmaceutically acceptable salt thereof. In other embodiments, human subjects have not demonstrated a significant clinical response to the treatment of myeloid malignancies with venetoclax or its pharmaceutically acceptable salts.

[0109] Venetoclax for use in the methods described herein can be provided in any suitable form that effectively inhibits the BCL-2 protein. Such forms include, but are not limited to, any suitable polymorph, amorphous or crystalline form, or any isomer or tautomer form. In some embodiments, the combination therapy described herein includes venetoclax synthesized according to the process described in US2010 / 0305122 (incorporated herein by reference). In another embodiment, the method described herein includes venetoclax formed or synthesized according to any of the processes described in CN107089981(A), CN107648185(A), EP3333167, WO2017 / 156398, WO2017 / 212431, WO2018 / 009444, WO2018 / 029711, WO2018 / 069941, WO2018 / 157803, and WO2018 / 167652 (all incorporated herein by reference). In a certain embodiment, the method described herein includes venetoclax in any crystalline or salt form described in WO2012 / 071336 (incorporated herein by reference).

[0110] Pharmacovigilant salts for use in accordance with the present invention include salts of acidic or basic groups.Pharmacovigilant acid addition salts include, but are not limited to, hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate, acidic phosphate, isonicotinate, acetate, lactate, salicylate, citrate, tartrate, pantothenate, hydrogen tartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucaronate, sugarate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, and pamoate (i.e., 1,1'-methylene-bis-(2-hydroxy-3-naphthoate)).Pharmacovigilant salts can be formed with various amino acids. Suitable base salts include, but are not limited to, aluminum salts, calcium salts, lithium salts, magnesium salts, potassium salts, sodium salts, zinc salts, and diethanolamine salts.

[0111] In one embodiment, human subjects have failed to treat myeloid malignancies with venetoclax. In another embodiment, human subjects have shown a clinical response to treatment of myeloid malignancies with venetoclax, but subsequently experienced relapse of myeloid malignancies. The clinical response may be any clinical response, including complete response, partial response, or minimal response. In one embodiment, human subjects have shown a clinical response to treatment of myeloid malignancies with venetoclax, but subsequently experienced a decline in their clinical response to venetoclax. In another embodiment, human subjects have shown a clinical response to treatment of myeloid malignancies with venetoclax, but subsequently became refractory to treatment with venetoclax. In yet another embodiment, human subjects have not shown a significant clinical response to treatment of myeloid malignancies with venetoclax.

[0112] This method involves administering a CD70-binding antibody or its antigen-binding fragment to a human subject. Administration may be carried out using any suitable route of administration, including, but is not limited to, oral, intravenous, intraperitoneal, pulmonary, and subcutaneous parenteral administration. For parenteral administration (e.g., intravenous, intraperitoneal, pulmonary, and subcutaneous), the CD70-binding antibody or its antigen-binding fragment may be administered to the human subject by injection or infusion.

[0113] As described herein, CD70 has already been characterized as a noteworthy target for anti-cancer therapy. CD70 is constitutively expressed in many types of hematological malignancies and solid tumors, and its expression is associated with poor prognosis in some cancers. Antibodies targeting CD70 have been developed, and some are moving toward clinical development.

[0114] Antibodies targeting CD70 have proven particularly effective in treating myeloid malignancies, especially in patients with acute myeloid leukemia (AML). Results from a Phase I / II clinical trial testing the CD70 antibody, ARGX-110 (xatuzumab), in AML patients demonstrated remarkable efficacy, particularly in newly diagnosed patients classified as ineligible for standard intensive chemotherapy (see WO2018 / 229303). Of particular note in this trial was the efficient reduction of leukemia stem cells (LSCs) in AML patients when used in combination with azacitidine. Examination of LSCs isolated from patients during the trial revealed evidence of increased asymmetric division of LSCs, indicating differentiation into myeloid cells. In summary, these results suggest that CD70 antibodies deplete the LSC pool in AML patients, thereby increasing the likelihood of remission and reducing the risk of relapse.

[0115] In one embodiment, the antibody that binds to CD70 is xatuzumab.

[0116] Additional CD70 antibodies or their antigen-binding fragments that may be used in the methods described herein include antibody-drug conjugates (ADCs). An ADC is an antibody conjugated to an active substance, such as auristatin and mytansin or other cytotoxic agents. Certain ADCs maintain antibody blocking and / or effector functions (e.g., ADCC, CDC, ADCP) while also delivering the conjugated active substance to cells expressing a target (e.g., CD70). Examples of anti-CD70 ADCs include borsetuzumab mahodotin (also known as SGN-75, Seattle Genetics), SGN-70A (Seattle Genetics), and MDX-1203 / BMS936561 (Bristol-Myers Squibb), each of which can be used in accordance with the present invention. Suitable anti-CD70 ADCs are also described in WO2008074004 and WO2004073656, which are also incorporated herein by reference, respectively.

[0117] In one embodiment, the antigen-binding fragment of the antibody bound to CD70 is independently selected from the group consisting of antibody light chain variable domain (VL); antibody heavy chain variable domain (VH); single-chain antibody (scFv); F(ab')2 fragment; Fab fragment; Fd fragment; Fv fragment; single-arm (monovalent) antibody; diabody, triabody, tetrabody, or any antigen-binding molecule formed by a combination, assembly, or complexation of these antigen-binding fragments.

[0118] In some embodiments, the myeloid malignancy is selected from the group consisting of acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), myeloproliferative neoplasm (MPN), chronic myeloid leukemia (CML), and chronic myelomonocytic leukemia (CMML). In some embodiments, the myeloid malignancy is AML. In some embodiments, the myeloid malignancy is MDS. In some embodiments, the myeloid malignancy is MPN. In some embodiments, the myeloid malignancy is CML. In some embodiments, the myeloid malignancy is CMML.

[0119] As described above, in a certain embodiment, the myeloid malignancy is AML. Acute myeloid leukemia is also known as acute myeloid leukemia, acute myeloid leukemia, acute granulocytic leukemia, and acute nonlymphocytic leukemia. According to the American Cancer Society's 2020 estimates of leukemia in the United States, there were approximately 19,940 new cases of AML, the majority of which were adults, and approximately 11,180 deaths from AML, the majority of which were adults. AML is the most common type of adult leukemia. However, AML accounts for only about 1% of all cancers and is remarkably rare overall. AML is generally a disease of the elderly and is rarely seen in those under 45 years of age. The average age of first diagnosis of AML is about 68 years, but AML can also occur in children.

[0120] In some embodiments, myeloid malignancies are monocytic AML. Acute monocytic leukemia (AMoL, or AML-M5), also known as monoblastic AML, is considered a type of acute myeloid leukemia (AML). According to World Health Organization (WHO) criteria, patients with AML-M5 must have more than 20% blasts in their bone marrow, and more than 80% of those blasts must be of the monocytic lineage.

[0121] As described above, in a certain embodiment, the myeloid malignancy is MDS. Myelodysplastic syndrome (MDS) is a group of various bone marrow disorders (cancers) in which the bone marrow does not produce enough healthy blood cells. MDS is often referred to as "bone marrow insufficiency disorder." In patients with myelodysplastic syndrome, hematopoietic stem cells (immature cells) in the bone marrow do not mature into red blood cells, white blood cells, or platelets. These immature blood cells are called blasts and do not perform their intended function, dying in the bone marrow or dying immediately after entering the bloodstream. As a result, there is less room for healthy white blood cells, red blood cells, and platelets to form in the bone marrow. When there are fewer healthy blood cells, patients are more susceptible to infections, anemia, or easy bleeding. Various types of MDS include, but are not limited to, refractory anemia, refractory anemia with ring sideroblasts, refractory anemia with blast cell proliferation, refractory cytopenia with polycytic lineage dysplasia, refractory cytopenia with monocytic lineage dysplasia, myelodysplastic syndromes associated with isolated deletion (5q) chromosome abnormalities, chronic myelomonocytic leukemia (CMML), and unclassified myelodysplastic syndromes.

[0122] In one embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject. The relevant expression levels can be determined using any suitable method, including, but not limited to, fluorescence-activated cell sorting (FACS), fluorescence microscopy using detectable (e.g., fluorescently labeled) antibodies specific to the relevant cell surface molecules(s), and mRNA expression analysis. In one embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of BCL-2 in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of CD117 in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of CD11b in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of CD68 in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of CD64 in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of BCL2A1 in the malignant myeloid cells of the human subject. In one embodiment, step (a) includes determining the expression level of MCL1 in the malignant myeloid cells of the human subject.

[0123] In some embodiments, at least one of BCL-2 and CD117 is downregulated, and at least one of CD11b, CD68, CD64, CD70, BCL2A1, and MCL1 is upregulated. In some embodiments, BCL-2 is downregulated and CD11b is upregulated. In some embodiments, BCL-2 is downregulated and CD68 is upregulated. In some embodiments, BCL-2 is downregulated and CD64 is upregulated. In some embodiments, BCL-2 is downregulated and CD70 is upregulated. In some embodiments, BCL-2 is downregulated and BCL2A1 is upregulated. In some embodiments, BCL-2 is downregulated and MCL1 is upregulated. In one embodiment, CD117 is downregulated and CD11b is upregulated. In one embodiment, CD117 is downregulated and CD68 is upregulated. In one embodiment, CD117 is downregulated and CD64 is upregulated. In one embodiment, CD117 is downregulated and CD70 is upregulated. In one embodiment, CD117 is downregulated and BCL2A1 is upregulated. In one embodiment, CD117 is downregulated and MCL1 is upregulated.

[0124] In a further embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of CD117, CD11b, and CD68. In a certain embodiment, step (a) includes determining the expression level of CD117 in malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD11b in malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD68 in malignant myeloid cells of the human subject. In a certain embodiment, CD117 is downregulated. In a certain embodiment, CD11b is upregulated. In a certain embodiment, CD68 is upregulated. In a certain embodiment, CD11b is upregulated and CD68 is upregulated. In a certain embodiment, CD117 is downregulated, CD11b is upregulated and CD68 is upregulated.

[0125] In a further embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of CD64, CD34, CD117, CD11b, CD68, and CD14 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD64 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD34 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD117 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD11b in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD68 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD14 in the malignant myeloid cells of the human subject. In one embodiment, CD64 is upregulated. In one embodiment, CD34 is downregulated. In one embodiment, CD117 is downregulated. In one embodiment, CD11b is upregulated. In one embodiment, CD68 is upregulated. In one embodiment, CD14 is upregulated. In one embodiment, CD64 is upregulated and CD34 is downregulated. In one embodiment, CD64 is upregulated and CD117 is downregulated. In one embodiment, CD64 is upregulated and CD11b is upregulated. In one embodiment, CD64 is upregulated and CD68 is upregulated. In one embodiment, CD64 is upregulated and CD14 is upregulated. In one embodiment, CD34 is downregulated and CD117 is downregulated. In another embodiment, CD34 is downregulated and CD11b is upregulated. In yet another embodiment, CD34 is downregulated and CD68 is upregulated.In one embodiment, CD34 is downregulated and CD14 is upregulated. In one embodiment, CD117 is downregulated and CD14 is upregulated. In one embodiment, CD68 is upregulated and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated and CD117 is downregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated and CD11b is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated and CD68 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD117 is downregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD34 is downregulated, CD117 is downregulated, and CD11b is upregulated. In one embodiment, CD34 is downregulated, CD117 is downregulated, and CD68 is upregulated. In one embodiment, CD34 is downregulated, CD11b is upregulated, and CD68 is upregulated. In one embodiment, CD34 is downregulated, CD117 is downregulated, and CD14 is upregulated. In one embodiment, CD34 is downregulated, CD11b is upregulated, and CD14 is upregulated. In another embodiment, CD117 is downregulated, CD11b is upregulated, and CD14 is upregulated. In yet another embodiment, CD34 is downregulated, CD68 is upregulated, and CD14 is upregulated.In one embodiment, CD117 is downregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD117 is downregulated, and CD11b is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD117 is downregulated, and CD68 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD11b is upregulated, and CD68 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD117 is downregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD11b is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD117 is downregulated, CD11b is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD117 is downregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated. In another embodiment, CD34 is downregulated, CD117 is downregulated, CD11b is upregulated, and CD68 is upregulated.In one embodiment, CD34 is downregulated, CD117 is downregulated, CD11b is upregulated, and CD14 is upregulated. In one embodiment, CD34 is downregulated, CD117 is downregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD34 is downregulated, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD117 is downregulated, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD117 is downregulated, CD11b is upregulated, and CD68 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD117 is downregulated, CD11b is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD117 is downregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD34 is downregulated, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated. In one embodiment, CD64 is upregulated, CD117 is downregulated, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated. In another embodiment, CD34 is downregulated, CD117 is downregulated, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated.In one embodiment, CD64 is upregulated, CD34 is downregulated, CD117 is downregulated, CD11b is upregulated, CD68 is upregulated, and CD14 is upregulated.

[0126] In a further embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of CD34, CD38, CD11b, CD33, and CD70 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD34 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD38 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD11b in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD33 in the malignant myeloid cells of the human subject. In a certain embodiment, step (a) includes determining the expression level of CD70 in the malignant myeloid cells of the human subject. In a certain embodiment, CD34 is downregulated. In a certain embodiment, CD38 is upregulated. In one embodiment, CD11b is upregulated. In one embodiment, CD33 is upregulated. In one embodiment, CD70 is upregulated. In one embodiment, CD34 is downregulated and CD38 is upregulated. In one embodiment, CD34 is downregulated and CD33 is upregulated. In one embodiment, CD34 is downregulated and CD70 is upregulated. In one embodiment, CD38 is upregulated and CD33 is upregulated. In one embodiment, CD38 is upregulated and CD11b is upregulated. In one embodiment, CD38 is upregulated and CD70 is upregulated. In one embodiment, CD33 is upregulated and CD11b is upregulated. In one embodiment, CD33 is upregulated and CD70 is upregulated. In another embodiment, CD38 is upregulated, CD33 is upregulated and CD34 is downregulated.In one embodiment, CD38 is upregulated, CD33 is upregulated, and CD11b is upregulated. In one embodiment, CD38 is upregulated, CD34 is downregulated, and CD11b is upregulated. In one embodiment, CD33 is upregulated, CD34 is downregulated, and CD11b is upregulated. In one embodiment, CD38 is upregulated, CD33 is upregulated, and CD70 is upregulated. In one embodiment, CD38 is upregulated, CD34 is downregulated, and CD70 is upregulated. In one embodiment, CD33 is upregulated, CD34 is downregulated, and CD70 is upregulated. In one embodiment, CD38 is upregulated, CD11b is upregulated, and CD70 is upregulated. In one embodiment, CD33 is upregulated, CD11b is upregulated, and CD70 is upregulated. In one embodiment, CD34 is downregulated, CD11b is upregulated, and CD70 is upregulated. In one embodiment, CD38 is upregulated, CD33 is upregulated, CD34 is downregulated, and CD11b is upregulated. In one embodiment, CD38 is upregulated, CD33 is upregulated, CD34 is downregulated, and CD70 is upregulated. In one embodiment, CD38 is upregulated, CD33 is upregulated, CD11b is upregulated, and CD70 is upregulated. In one embodiment, CD38 is upregulated, CD34 is downregulated, CD11b is upregulated, and CD70 is upregulated. In one embodiment, CD33 is upregulated, CD34 is downregulated, CD11b is upregulated, and CD70 is upregulated.In one embodiment, CD38 is upregulated, CD33 is upregulated, CD34 is downregulated, CD11b is upregulated, and CD70 is upregulated.

[0127] In a further embodiment, step (a) includes determining the expression level of at least one marker selected from the group consisting of CD38, CD11b, and CD33 in the malignant myeloid cells of the human subject. In one embodiment, CD38 is upregulated, CD33 is upregulated, and CD11b is upregulated.

[0128] In further embodiments, step (a) includes determining the expression level of at least one marker selected from the group consisting of CD45, CD11b, and CD117 in the malignant myeloid cells of the human subject. In some embodiments, CD45 is upregulated. In some embodiments, CD45 is upregulated and CD11b is upregulated. In some embodiments, CD45 is upregulated and CD117 is downregulated. In some embodiments, CD45 is upregulated, CD11b is upregulated and CD117 is downregulated.

[0129] In further embodiments, step (a) includes determining the expression level of CD45 and determining the SSC value. In one embodiment, cells are characterized as CD45 bright and SSC high.

[0130] In a specific embodiment, treatment with a clinical history of BCL-2 inhibitors (e.g., venetoclax) upregulated CD70 expression in myeloid cells. Patients with myeloid malignancies who have failed treatment with BCL-2 can then be treated with an antibody that binds to CD70 or its antigen-binding fragment (e.g., xatuzumab). Treatment with an antibody that binds to CD70 or its antigen-binding fragment then upregulates BCL-2 expression in myeloid cells. Therefore, treatment with a BCL-2 inhibitor (e.g., venetoclax) and treatment with an antibody that binds to CD70 or its antigen-binding fragment (e.g., xatuzumab) have a synergistic effect in patients with myeloid malignancies and improve the treatment response in these patients. In a specific embodiment, an anti-CD70 antibody or its CD70-binding fragment is combined with a BCL-2 inhibitor (co-administered) for use in the treatment of myeloid malignancies in patients who are resistant to treatment with a BCL-2 inhibitor. In one embodiment, step (a) includes determining the CD70 expression level of malignant myeloid cells of the human subject. The relevant expression level can be determined using any suitable method, including, but not limited to, fluorescence-activated cell sorting (FACS) and fluorescence microscopy using a detectable (e.g., fluorescently labeled) antibody specific to CD70.

[0131] In one embodiment, CD70 is upregulated compared to CD70 expression levels measured before or during treatment with a BCL-2 inhibitor. In one embodiment, the treatment with the BCL-2 inhibitor includes treatment with venetoclax. In one embodiment, the treatment with the BCL-2 inhibitor includes treatment with a BCL-2 inhibitor other than venetoclax.

[0132] In one embodiment, the human subject is (a) Treatment with BCL-2 inhibitors, and, (b) Absence of remission in response to treatment with the BCL-2 inhibitor, The patient has a clinical history that includes the following. In one embodiment, the absence of remission means the absence of complete remission. In another embodiment, the absence of remission means the absence of at least partial remission. In one embodiment, the treatment with a BCL-2 inhibitor in the clinical history is treatment with venetoclax. In another embodiment, the treatment with a BCL-2 inhibitor in the clinical history is treatment with a BCL-2 inhibitor other than venetoclax.

[0133] In one embodiment, treatment with a BCL-2 inhibitor as described in the clinical history further includes treatment with a hypomethylating agent (HMA). Hypomethylating agents inhibit the normal methylation of DNA and / or RNA. Examples of hypomethylating agents, though not limited to them, include azacitidine, decitabine, and guadecitabine.

[0134] Azacitidine is an analog of cytidine, and decitabine is its deoxy derivative. Guadecitabine is a cytidine deaminase-resistant prodrug of decitabine. Azacitidine and decitabine are DNA methyltransferase (DNMT) inhibitors and are known to upregulate gene expression by promoter hypomethylation. Such hypomethylation disrupts cellular function and consequently produces cytotoxic effects.

[0135] In one embodiment, the human subject is (a) Treatment with BCL-2 inhibitors, (b) Partial or complete remission, (c) Partial or complete relapse, He has a clinical history that includes [this].

[0136] In one embodiment, the human subject has a clinical history including treatment with a BCL-2 inhibitor, partial remission, and partial relapse.

[0137] In one embodiment, the human subject has a clinical history including treatment with BCL-2 inhibitors, partial remission, and complete relapse.

[0138] In one embodiment, the human subject has a clinical history including treatment with BCL-2 inhibitors, complete remission, and partial relapse.

[0139] In one embodiment, the human subject has a clinical history including treatment with a BCL-2 inhibitor, complete remission, and complete relapse.

[0140] Further according to these embodiments, in some embodiments, the treatment with the BCL-2 inhibitor in the clinical history further includes treatment with a hypomethylating agent (HMA). In some embodiments, the treatment with the BCL-2 inhibitor in the clinical history is treatment with venetoclax or a pharmaceutically acceptable salt thereof. In other embodiments, the treatment with the BCL-2 inhibitor in the clinical history is treatment with a BCL-2 inhibitor other than venetoclax or a pharmaceutically acceptable salt thereof. As stated above, examples of hypomethylating agents that are not limited to these include azacitidine, decitabine, and guadecitabine.

[0141] Further according to these embodiments, in some embodiments, a hypomethylating agent (HMA) is co-administered with an antibody or antigen-binding fragment thereof that binds to the CD70. In some embodiments, the HMA is selected from the group consisting of azacitidine, decitabine, guadecitabine, and any combination thereof. In some embodiments, the HMA is azacitidine. In some embodiments, the HMA is decitabine. In some embodiments, the HMA is guadecitabine.

[0142] HMA can be administered on the same or substantially the same schedule as the antibody or antigen-binding fragment that binds to D70, or on a different schedule than that of the antibody or antigen-binding fragment that binds to D70. Furthermore, the route of administration of HMA may be the same as that of the antibody or antigen-binding fragment that binds to D70, or it may be different from that of the antibody or antigen-binding fragment that binds to D70.

[0143] Further according to these embodiments, in one embodiment, a BCL-2 inhibitor is co-administered with an antibody or antigen-binding fragment thereof that binds to CD70. The BCL-2 inhibitor may be administered on the same or substantially the same schedule as the antibody or antigen-binding fragment that binds to D70, or on a different schedule than that of the antibody or antigen-binding fragment that binds to D70. Furthermore, the route of administration of the BCL-2 inhibitor may be the same as the route of administration of the antibody or antigen-binding fragment that binds to D70, or it may be different from the route of administration of the antibody or antigen-binding fragment that binds to D70.

[0144] In one embodiment, an antibody or antigen-binding fragment thereof that binds to CD70 is co-administered with a BCL-2 inhibitor and a hypomethylating agent (HMA). In one embodiment, the antibody that binds to CD70 is xatuzumab. In one embodiment, the BCL-2 inhibitor is venetoclax or a pharmaceutically acceptable salt thereof. In one embodiment, the HMA is azacitidine. In one embodiment, xatuzumab is co-administered with venetoclax and azacitidine.

[0145] Further according to these embodiments, in some embodiments, venetoclax or a pharmaceutically acceptable salt thereof is co-administered with an antibody or antigen-binding fragment thereof that binds to CD70. The venetoclax or a pharmaceutically acceptable salt thereof may be administered on the same or substantially the same schedule as the antibody or antigen-binding fragment thereof that binds to D70, or on a different schedule than that of the antibody or antigen-binding fragment thereof that binds to D70. Furthermore, the route of administration of the venetoclax or a pharmaceutically acceptable salt thereof may be the same as the route of administration of the antibody or antigen-binding fragment thereof that binds to D70, or it may be different from the route of administration of the antibody or antigen-binding fragment thereof that binds to D70.

[0146] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, The amino acid sequence of HCDR1 is sequence number 1; The amino acid sequence of HCDR2 is sequence number 2; The amino acid sequence of HCDR3 is sequence number 3; The amino acid sequence of LCDR1 is sequence number 4; The amino acid sequence of LCDR2 consists of sequence number 5; and, The amino acid sequence of LCDR3 is sequence number 6.

[0147] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 90% the same amino acid sequence as SEQ ID NO: 7.

[0148] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 91% the same amino acid sequence as SEQ ID NO: 7.

[0149] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 92% the same amino acid sequence as SEQ ID NO: 7.

[0150] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 93% the same amino acid sequence as SEQ ID NO: 7.

[0151] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 94% the same amino acid sequence as SEQ ID NO: 7.

[0152] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 95% the same amino acid sequence as SEQ ID NO: 7.

[0153] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 96% the same amino acid sequence as SEQ ID NO: 7.

[0154] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 97% the same amino acid sequence as SEQ ID NO: 7.

[0155] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 98% the same amino acid sequence as SEQ ID NO: 7.

[0156] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 99% the same amino acid sequence as SEQ ID NO: 7.

[0157] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 100% the same amino acid sequence as SEQ ID NO: 7.

[0158] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 90% the same amino acid sequence as SEQ ID NO: 8.

[0159] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 91% the same amino acid sequence as SEQ ID NO: 8.

[0160] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 92% the same amino acid sequence as SEQ ID NO: 8.

[0161] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 93% the same amino acid sequence as SEQ ID NO: 8.

[0162] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 94% the same amino acid sequence as SEQ ID NO: 8.

[0163] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 95% the same amino acid sequence as SEQ ID NO: 8.

[0164] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 96% the same amino acid sequence as SEQ ID NO: 8.

[0165] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 97% the same amino acid sequence as SEQ ID NO: 8.

[0166] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 98% the same amino acid sequence as SEQ ID NO: 8.

[0167] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 99% the same amino acid sequence as SEQ ID NO: 8.

[0168] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable light chain domain (VL) having at least 100% the same amino acid sequence as SEQ ID NO: 8.

[0169] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 90% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 90% the same amino acid sequence as SEQ ID NO: 8.

[0170] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 91% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 91% the same amino acid sequence as SEQ ID NO: 8.

[0171] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 92% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 92% the same amino acid sequence as SEQ ID NO: 8.

[0172] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 93% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 93% the same amino acid sequence as SEQ ID NO: 8.

[0173] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 94% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 94% the same amino acid sequence as SEQ ID NO: 8.

[0174] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 95% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 95% the same amino acid sequence as SEQ ID NO: 8.

[0175] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 96% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 96% the same amino acid sequence as SEQ ID NO: 8.

[0176] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 97% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 97% the same amino acid sequence as SEQ ID NO: 8.

[0177] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having an amino acid sequence at least 98% identical to that of SEQ ID NO: 7, and a variable light chain domain (VL) having an amino acid sequence at least 98% identical to that of SEQ ID NO: 8.

[0178] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 99% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 99% the same amino acid sequence as SEQ ID NO: 8.

[0179] In one embodiment, the antibody or antibody-binding fragment that binds to CD70 includes a variable heavy chain domain (VH) having at least 100% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 100% the same amino acid sequence as SEQ ID NO: 8.

[0180] In one embodiment, the amino acid sequence identical to VH consisting of sequence number 7 by at least 90% includes HCDR1, HCDR2, and HCDR3. The amino acid sequence of HCDR1 is sequence number 1; The amino acid sequence of HCDR2 consists of sequence number 2; and, The amino acid sequence of HCDR3 consists of sequence number 3; and, The amino acid sequence that is at least 90% identical to the VL consisting of Sequence ID No. 8 includes LCDR1, LCDR2, and LCDR3. The amino acid sequence of LCDR1 is sequence number 4; The amino acid sequence of LCDR2 consists of sequence number 5; and, The amino acid sequence of LCDR3 is sequence number 6.

[0181] In some embodiments, the method may include one or more additional therapeutic agents, for example, at least one additional anticancer agent, preferably a therapeutic agent for myeloid malignancies. In some embodiments, the additional anticancer agent is a therapeutic agent for acute myeloid leukemia (AML).

[0182] In one embodiment, the CD70 antibody or its antigen-binding fragment is administered in a dose range of 0.1 to 25 mg / kg per dose, preferably 10 mg / kg. Alternatively, the BCL-2 inhibitor, preferably venetoclax or a pharmaceutically acceptable salt thereof, may be administered in a dose range of 100 to 600 mg per dose. In a preferred embodiment, the method described herein includes administration in combination with an additional azacitidine, in which case the azacitidine is administered in a dose of 75 mg / m2. In a more preferred embodiment, the method described herein includes administration in combination with an additional decitabine, in which case the decitabine is administered in a dose of 20 mg / m2.

[0183] In some embodiments, the method further includes monitoring the patient's blast count. The number in the patient's peripheral blood and / or bone marrow may decrease, for example to less than 25%, for example to 5%, for example to less than 5%, for example to minimal residual disease levels, for example to undetectable levels. In some embodiments, the bone marrow blast count decreases by 5% to 25%, and the bone marrow blast percentage decreases by more than 50% compared to before treatment.

[0184] In some embodiments, the method induces partial remission. In some embodiments, the method induces complete remission, optionally accompanied by platelet and / or neutrophil recovery. The method can induce transfusion withdrawal of red blood cells, platelets, or both within 8 weeks, 10 weeks, or 12 weeks or longer. In some embodiments, the method reduces mortality at 30 or 60 days.

[0185] In certain embodiments, the method extends survival. For example, the method may extend survival compared to the standard care agents (including multiple agents) employed to treat certain myeloid malignancies treated with this combination. The method may induce a minimal residual disease (RAM) state.

[0186] In one embodiment, the method further includes the step of administering a bone marrow transplant to the subject. Alternatively or additionally, the method may further include the step of administering one or more additional anticancer agents. The one or more additional anticancer agents may be selected from any agents suitable for the treatment of myeloid malignancies, preferably AML. Preferred agents may be selected from: selectin inhibitors (e.g., GMI-1271); FMS-like tyrosine kinase receptor 3 (FLT3) inhibitors (e.g., midostaurin); cyclin-dependent kinase inhibitors; aminopeptidase inhibitors; JAK / STAT inhibitors; cytarabine; anthracycline compounds (e.g., daunorubicin, idarubicin); doxorubicin; hydroxyurea; Vyxeos; IDH1 or IDH2 inhibitors, such as Idhifa (or enasidenib) or tybusovo (or ivosidenib); smoothed inhibitors, such as glassedigib; BET bromodomain inhibitors; agents targeting CD123 or CD33; HDAC inhibitors; agents targeting LSCs; AML bone marrow niche targeting agents; and NEDD8 activating enzyme inhibitors, such as pevonedistat.

[0187] The CD70 antibody or antigen-binding fragment of the method described herein may be formulated using any suitable pharmaceutical carrier, adjuvant, and / or excipient. The formulation techniques for antibodies for human therapeutic use are well known in the art and have been validated, for example, in the literature by Wang et al., (2007) Journal of Pharmaceutical Sciences, 96:1-26, the contents of which are incorporated herein by reference in their entirety. Pharmaceutically acceptable excipients that can be used in the formulation of this antibody composition include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffering substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based substances (e.g., sodium carboxymethylcellulose), polyethylene glycol, polyacrylate, wax, polyethylene-polyoxypropylene-block polymer, polyethylene glycol, wool fat, and hyaluronidase (e.g., PH20 enzyme).

[0188] BCL-2 inhibitors (preferably venetoclax or a pharmaceutically acceptable salt thereof) may be formulated using any suitable pharmaceutical carrier, adjuvant, and / or excipient. Suitable agents include, for example, encapsulating materials or additives such as absorption enhancers, antioxidants, binders, buffers, coatings, colorants, diluents, disintegrants, emulsifiers, bulking agents, fillers, flavoring agents, humectants, lubricants, fragrances, preservatives, propellants, release agents, sterilizers, sweeteners, solubilizers, wetting agents, and mixtures thereof.

[0189] CD70 antibodies, particularly ARGX-110, have been found to be effective even at relatively low doses in the treatment of myeloid malignancies, especially AML. Therefore, in certain embodiments of all the methods of the present invention, the CD70 antibody or its antigen-binding fragment is administered in doses ranging from 0.1 mg / kg to 30 mg / kg per dose. In certain embodiments of all the methods of the present invention, the CD70 antibody or its antigen-binding fragment is administered in doses ranging from 0.1 mg / kg to 25 mg / kg per dose, for example, in the range of 0.1 mg / kg to 20 mg / kg. In certain embodiments, the CD70 antibody or its antigen-binding fragment is administered in doses ranging from 1 mg / kg to 20 mg / kg per dose. Unless otherwise specified, the ranges described herein include the endpoints of the range; for example, the administration of doses in the range of 0.1 to 25 mg / kg includes administration at a dose of 0.1 mg / kg and administration at a dose of 25 mg / kg, and also includes administration at all doses between the two endpoints.

[0190] In one embodiment of the method of the present invention, the CD70 antibody or its antigen-binding fragment is administered in a dose ranging from 0.1 to 15 mg / kg. In another embodiment, the CD70 antibody or its antigen-binding fragment is administered in a dose ranging from 0.5 to 2 mg / kg. In yet another embodiment, the CD70 antibody or its antigen-binding fragment is administered in doses of 1 mg / kg, 3 mg / kg, 10 mg / kg, or 20 mg / kg. In one preferred embodiment, the CD70 antibody or its antigen-binding fragment is administered in a dose of 1 mg / kg. In one preferred embodiment, the CD70 antibody or its antigen-binding fragment is administered in a dose of 10 mg / kg.

[0191] In one embodiment, multiple doses of CD70 antibody or antigen-binding fragment are administered. In one such embodiment, each dose of CD70 antibody or its antigen-binding fragment is spaced 10 to 20 days apart, optionally 12 to 18 days apart. In one embodiment, each dose of anti-CD70 antibody is spaced 14 to 17 days apart.

[0192] BCL-2 inhibitors, preferably venetoclax or pharmaceutically acceptable salts thereof, may be administered according to any regimen determined to be effective for the compound. FDA prescribing information for the use of venetoclax in the treatment of AML suggests a dosing schedule with an escalation phase followed by a maintenance phase. When venetoclax is prescribed in combination with azacitidine or decitabine, the recommended dosing schedule consists of 100 mg of venetoclax on day 1; 200 mg of venetoclax on day 2; 400 mg of venetoclax on day 3; and thereafter, 400 mg of venetoclax daily in combination with 75 mg / m2 of azacitidine or 20 mg / m2 of decitabine until disease progression or unacceptable toxicity is observed. When vencrecsta® is prescribed in combination with low-dose cytarabine, the recommended dosing schedule is 100 mg of vencrecsta® on day 1; 200 mg of vencrecsta® on day 2; 400 mg of vencrecsta® on day 3; and thereafter, 600 mg of vencrecsta® daily in combination with 20 mg / m2 until disease progression or unacceptable toxicity is observed.

[0193] In one embodiment, each dose of venetoclax or a pharmaceutically acceptable salt thereof, for example, an oral dose, is in the range of 100 mg to 600 mg. In one embodiment, venetoclax or a pharmaceutically acceptable salt thereof is administered at a dose of 400 mg per day. In another embodiment, venetoclax or a pharmaceutically acceptable salt thereof is administered at a dose of 600 mg per day. As described above, a fixed daily dose of venetoclax may be preceded by a dose escalation period of, for example, three days, during which the patient is administered a dose of venetoclax that is increased until the maintenance daily dose is reached.

[0194] In a certain embodiment, the method described herein includes monitoring the number of blast cells in a patient. As used herein, “blast cells” or “blast” refers to myeloblasts or myeloid blasts, which are myeloid progenitor cells in the bone marrow. In healthy individuals, blast cells are not found in peripheral blood circulation, and blast cells should constitute less than 5% of the bone marrow. In patients with myeloid malignancies, particularly AML and MDS, the production of abnormal blasts with disrupted differentiation potential is increased, and this overproduction of abnormal blasts can be detected by monitoring the number of blast cells in the patient’s peripheral blood circulation, bone marrow, or both.

[0195] The proportion of blast cells in bone marrow or peripheral blood can be assessed by methods known in the art, such as flow cytometry analysis or morphological evaluation of cells obtained from a bone marrow biopsy or peripheral blood smear. The proportion of blast cells is determined relative to the total number of cells in the sample. For example, flow cytometry can be used to determine the proportion of blast cells using the number of CD45dim, SSClow cells relative to the total number of cells. As a further example, morphological evaluation can be used to determine the number of morphologically identified blast cells for all cells within the examination field.

[0196] In one embodiment, a method is provided for reducing the proportion of blast cells in the bone marrow to less than 25%, less than 20%, or for example, less than 10%. In another embodiment, a method is provided for reducing the proportion of blast cells in the bone marrow to less than 5%. In yet another embodiment, a method is provided for reducing the proportion of blast cells in the bone marrow to approximately 5% to approximately 25%, wherein the proportion of bone marrow blast cells compared to the proportion of bone marrow blast cells before the implementation of the method (or before treatment) is reduced by more than 50%.

[0197] In one embodiment, a method is provided for reducing the proportion of blast cells in peripheral blood to less than 25%, less than 20%, or, for example, less than 10%. In another embodiment, a method is provided for reducing the proportion of blast cells in peripheral blood to less than 5%. In yet another embodiment, a method is provided for reducing the proportion of blast cells in peripheral blood to about 5% to about 25%, wherein the reduction in the proportion of blast cells in peripheral blood compared to the proportion of peripheral blast cells before the implementation of the method (or before treatment) is more than 50%.

[0198] For the clinical determination of the blast cell percentage, an assessment of cell morphology (also known as cytomorphology) is usually preferred.

[0199] In specific embodiments, the methods described herein induce complete response. In the context of AML treatment, complete response or "complete remission" is defined as: myeloblasts < 5%; absence of circulating blasts and blasts with Auer bodies; absence of extramedullary disease; ANC ≥ 1.0 × 10⁹ / L (1000 μL); platelet count ≥ 100 × 10⁹ / L (100,000 μL), for which refer to Dohner et al., (2017) Blood, 129(4):424-447.

[0200] This method can achieve a complete response with platelet recovery, i.e., a response with a platelet count of >100 × 10⁹ / L (100,000 / μL). This method can achieve a complete response with neutrophil recovery, i.e., a response with a neutrophil count of >1.0 × 10⁹ / L (1000 / μL). Alternatively, or in addition to the above, this method can induce transfusion-free erythrocytes, platelets, or both within 8 weeks, 10 weeks, or 12 weeks or longer.

[0201] In specific embodiments, the methods described herein induce a negative minimal residual disease (MRD) or measurable residual disease (MRD) stage; see Schuurhuis et al., (2018) Blood, 131(12):1275-1291 for details.

[0202] In one embodiment, the method described herein induces complete response without minimal residual disease (CRMRD-), for which see Dohner et al., (2017) Blood, 129(4):424-447.

[0203] This method may achieve a partial response or induce partial remission. In the context of AML treatment, a partial response or partial remission includes a 5% to 25% reduction in the myeloblast percentage and at least a 50% reduction in the pre-treatment myeloblast percentage; please refer to the aforementioned Dohner et al.

[0204] The methods described herein may extend survival. As used herein, the term “survival” refers to overall survival, 1-year survival, 2-year survival, 5-year survival, event-free survival, and progression-free survival. The methods described herein may extend survival compared to gold-standard treatments for the specific disease or condition being treated. Gold-standard treatments may also be identified as best practice, standard care, standard medical care, or standard therapy. For any given disease, there may be one or more gold-standard treatments, for example, depending on different clinical practices in different countries. These treatments already available for myeloid malignancies are diverse and include chemotherapy, radiotherapy, stem cell transplantation, and specific targeted therapies. Furthermore, clinical guidelines in both the United States and Europe govern the standard treatment of myeloid malignancies such as AML. For more information, see O'Donnell et al., (2017) Journal of the National Comprehensive Cancer Network, 15(7):926-957, and Dohner et al., (2017) Blood, 129(4):424-447 (both incorporated by reference).

[0205] The method of the present invention may extend or improve survival compared to patients receiving any standard treatment for myeloid malignancies.

[0206] The methods described herein may include a further step of administering a bone marrow transplant to a patient or subject. The methods described herein may also be used to prepare a patient or subject with a myeloid malignancy for a bone marrow transplant. As stated above, the methods of the present invention may be performed to reduce the absolute or relative number of blast cells in the bone marrow or peripheral blood. In a certain embodiment, the method is performed to reduce the number of blast cells in the bone marrow and / or peripheral blood prior to transplantation. The method may be used to reduce the number of blast cells to less than 5% in preparation for a bone marrow transplant in a patient or subject.

[0207] One aspect of the present invention is a method for identifying and treating a patient who should be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy, and the method is: (i) To measure the differentiation status of the patient's myeloid cells, (ii) Determine whether the patient has differentiated monocytic AML, and identify patients with differentiated monocytic AML as patients who should be treated with the anti-CD70 antibody or its CD70-binding fragment, and (iii) Administer the anti-CD70 antibody or its CD70 conjugate fragment to a patient identified as a patient who should be treated with the anti-CD70 antibody or its CD70 conjugate fragment. The method includes the step of [the specified step].

[0208] In one embodiment, steps (i) and (ii) are performed on a sample obtained from a patient with the myeloid malignancy.

[0209] In one embodiment, the patient's bone marrow sample contains CD45bright / SSChigh / CD38+ / CD34- / CD33+ / CD11b+ / CD70+ phenotype cells or CD45bright / SSChigh / CD34- / CD117- / CD11b+ / CD68+ / CD14+ / CD64+ phenotype cells.

[0210] (C. Pharmaceutical Uses) In a further embodiment, the present invention provides a CD70-binding antibody or an antigen-binding fragment thereof for therapeutic use. In particular, the CD70-binding antibody or the antigen-binding fragment thereof is for use in the treatment of myeloid malignancies in human subjects. In particular, the CD70-binding antibody or the antigen-binding fragment thereof is for use in the treatment of myeloid malignancies in human subjects that are resistant to treatment with BCL-2 inhibitors.

[0211] In a particular embodiment, the human subject is identified as having differentiated monocyte-type AML based on differential expression levels of one or more markers.

[0212] In a special embodiment, before the treatment, (i) Measuring the differentiation state of bone marrow cells in the human subject, (ii) To determine whether the human subject has differentiated monocytic AML, A selection process is carried out, including the step of administering a therapeutically effective dose of the anti-CD70 antibody or its anti-CD70 conjugate fragment to a human subject having differentiated monocytic AML.

[0213] In a further embodiment, the present invention is (a) Selecting human subjects with myeloid malignancies that have reduced sensitivity to BCL-2 inhibitors or are refractory, (b) Administering to the human subject an antibody that binds to CD70 or an antigen-binding fragment thereof, The present invention provides an antibody or antigen-binding fragment thereof that binds to CD70 and is used in a method for treating myeloid malignancies in human subjects, which includes the step of [the specified step].

[0214] In one embodiment, an antibody or antigen-binding fragment thereof that binds to CD70 is provided for use in a method for treating myeloid malignancies in human subjects as described herein, and which is administered in combination with a BCL-2 inhibitor.

[0215] In one embodiment, an antibody or antigen-binding fragment thereof that binds to CD70 is provided for use in a method for treating myeloid malignancies in human subjects as described herein, and which is administered in combination with a hypomethylating agent (HMA).

[0216] In one embodiment, an antibody or antigen-binding fragment thereof that binds to CD70 is provided for use in a method for treating myeloid malignancies in human subjects as described herein, and which is administered in combination with a BCL-2 inhibitor and a hypomethylating agent (HMA).

[0217] In one embodiment, a combination of a CD70-binding antibody or its antigen-binding fragment and a BCL-2 inhibitor is provided for use in a method for treating myeloid malignancies in human subjects as described herein.

[0218] In one embodiment, a combination of a CD70-binding antibody or its antigen-binding fragment and a hypomethylating agent (HMA) is provided for use in a method for treating myeloid malignancies in human subjects as described herein.

[0219] In one embodiment, a combination of a CD70-binding antibody or its antigen-binding fragment, a BCL-2 inhibitor, and a hypomethylating agent (HMA) is provided for use in a method for treating myeloid malignancies in human subjects as described herein.

[0220] In one embodiment, the antibody that binds to CD70 is xatuzumab. In one embodiment, the BCL-2 inhibitor is venetoclax or a pharmaceutically acceptable salt thereof. In one embodiment, the HMA is azacitidine. In one embodiment, the combination is xatuzumab, venetoclax, and azacitidine.

[0221] In one embodiment, the myeloid malignancy is AML. In one embodiment, the myeloid malignancy is monocyte-type AML. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on differential expression levels of one or more markers.

[0222] In one embodiment, the human subject is identified as having differentiated monocytic AML based on differential expression levels of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject. The relevant expression levels can be determined using any suitable method, including, but not limited to, fluorescence-activated cell sorting (FACS), fluorescence microscopy using detectable (e.g., fluorescently labeled) antibodies specific to the relevant cell surface molecules, and mRNA expression analysis. In one embodiment, the human subject is identified as having differentiated monocytic AML based on differential expression levels of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the differential expression level of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of BCL-2 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD117 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD11b in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD68 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the CD64 expression level of the malignant myeloid cells of the human subject. In another embodiment, the human subject is identified as having differentiated monocytic AML based on the BCL2A1 expression level of the malignant myeloid cells of the human subject.In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of MCL1 in the malignant myeloid cells of the human subject.

[0223] In some embodiments, the expression levels of at least one of BCL-2 and CD117 are downregulated, and at least one of CD11b, CD68, CD64, CD70, BCL2A1, and MCL1 is upregulated. In some embodiments, the expression level of BCL-2 is downregulated, and the expression level of CD11b is upregulated. In some embodiments, the expression level of BCL-2 is downregulated, and the expression level of CD68 is upregulated. In some embodiments, the expression level of BCL-2 is downregulated, and the expression level of CD64 is upregulated. In some embodiments, the expression level of BCL-2 is downregulated, and the expression level of CD70 is upregulated. In some embodiments, the expression level of BCL-2 is downregulated, and the expression level of BCL2A1 is upregulated. In one embodiment, the expression level of BCL-2 is downregulated and the expression level of MCL1 is upregulated. In one embodiment, the expression level of CD117 is downregulated and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD117 is downregulated and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD117 is downregulated and the expression level of CD64 is upregulated. In one embodiment, the expression level of CD117 is downregulated and the expression level of CD70 is upregulated. In one embodiment, the expression level of CD117 is downregulated and the expression level of BCL2A1 is upregulated. In one embodiment, the expression level of CD117 is downregulated and the expression level of MCL1 is upregulated.

[0224] In a further embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of at least one marker selected from the group consisting of CD117, CD11b, and CD68. In a certain embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD117 in the malignant myeloid cells of the human subject. In a certain embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD11b in the malignant myeloid cells of the human subject. In a certain embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD68 in the malignant myeloid cells of the human subject. In a certain embodiment, the expression level of CD117 is downregulated. In a certain embodiment, the expression level of CD11b is upregulated. In a certain embodiment, the expression level of CD68 is upregulated. In one embodiment, the expression level of CD11b is upregulated, and the expression level of CD68 is upregulated. In another embodiment, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD68 is upregulated.

[0225] In further embodiments, the human subject is identified as having differentiated monocyte-type AML based on the expression level of at least one marker selected from the group consisting of CD64, CD34, CD117, CD11b, CD68, and CD14 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD64 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD34 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD117 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD11b in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD68 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocytic AML based on the expression level of CD14 in the malignant myeloid cells of the human subject. In one embodiment, the expression level of CD64 is upregulated. In one embodiment, the expression level of CD34 is downregulated. In one embodiment, the expression level of CD117 is downregulated. In one embodiment, the expression level of CD11b is upregulated. In one embodiment, the expression level of CD68 is upregulated. In one embodiment, the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated and the expression level of CD34 is downregulated. In one embodiment, the expression level of CD64 is upregulated and the expression level of CD117 is downregulated. In another embodiment, the expression level of CD64 is upregulated and the expression level of CD11b is upregulated. In yet another embodiment, the expression level of CD64 is upregulated and the expression level of CD68 is upregulated.In one embodiment, the expression level of CD64 is upregulated and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD34 is downregulated and the expression level of CD117 is downregulated. In one embodiment, the expression level of CD34 is downregulated and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD34 is downregulated and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD34 is downregulated and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD117 is downregulated and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD68 is upregulated and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD117 is downregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD117 is downregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression levels of CD64, CD68, and CD14 are upregulated. In another embodiment, the expression levels of CD34, CD117, and CD11b are downregulated.In one embodiment, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD117 is downregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD68 is upregulated.In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD117 is downregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated.In one embodiment, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD68 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In one embodiment, the expression level of CD64 is upregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated. In another embodiment, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated.In one embodiment, the expression level of CD64 is upregulated, the expression level of CD34 is downregulated, the expression level of CD117 is downregulated, the expression level of CD11b is upregulated, the expression level of CD68 is upregulated, and the expression level of CD14 is upregulated.

[0226] In further embodiments, the human subject is identified as having differentiated monocyte-type AML based on the expression level of at least one marker selected from the group consisting of CD34, CD38, CD11b, CD33, and CD70 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD34 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD38 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD11b in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the expression level of CD33 in the malignant myeloid cells of the human subject. In one embodiment, the human subject is identified as having differentiated monocyte-type AML based on the CD70 expression level of the malignant myeloid cells of the human subject. In one embodiment, the CD34 expression level is downregulated. In one embodiment, the CD38 expression level is upregulated. In one embodiment, the CD11b expression level is upregulated. In one embodiment, the CD33 expression level is upregulated. In one embodiment, the CD70 expression level is upregulated. In one embodiment, the CD34 expression level is downregulated and the CD38 expression level is upregulated. In one embodiment, the CD34 expression level is downregulated and the CD33 expression level is upregulated. In one embodiment, the CD34 expression level is downregulated and the CD70 expression level is upregulated. In one embodiment, the expression level of CD38 is upregulated, and the expression level of CD33 is upregulated. In another embodiment, the expression level of CD38 is upregulated, and the expression level of CD11b is upregulated. In yet another embodiment, the expression level of CD38 is upregulated, and the expression level of CD70 is upregulated.In one embodiment, the expression level of CD33 is upregulated, and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD33 is upregulated, and the expression level of CD70 is upregulated. In one embodiment, the expression level of CD38 is upregulated, the expression level of CD33 is upregulated, and the expression level of CD34 is downregulated. In one embodiment, the expression level of CD38 is upregulated, the expression level of CD33 is upregulated, and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD38 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD33 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD11b is upregulated. In one embodiment, the expression levels of CD38 are upregulated, the expression levels of CD33 are upregulated, and the expression levels of CD70 are upregulated. In one embodiment, the expression levels of CD38 are upregulated, the expression levels of CD34 are downregulated, and the expression levels of CD70 are upregulated. In one embodiment, the expression levels of CD33 are upregulated, the expression levels of CD34 are downregulated, and the expression levels of CD70 are upregulated. In one embodiment, the expression levels of CD38 are upregulated, the expression levels of CD11b are upregulated, and the expression levels of CD70 are upregulated. In one embodiment, the expression levels of CD33 are upregulated, the expression levels of CD11b are upregulated, and the expression levels of CD70 are upregulated. In one embodiment, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD70 is upregulated.In one embodiment, the expression level of CD38 is upregulated, the expression level of CD33 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD11b is upregulated. In one embodiment, the expression level of CD38 is upregulated, the expression level of CD33 is upregulated, the expression level of CD34 is downregulated, and the expression level of CD70 is upregulated. In one embodiment, the expression level of CD38 is upregulated, the expression level of CD33 is upregulated, the expression level of CD11b is upregulated, and the expression level of CD70 is upregulated. In one embodiment, the expression level of CD38 is upregulated, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD70 is upregulated. In one embodiment, the expression level of CD33 is upregulated, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD70 is upregulated. In another embodiment, the expression level of CD38 is upregulated, the expression level of CD33 is upregulated, the expression level of CD34 is downregulated, the expression level of CD11b is upregulated, and the expression level of CD70 is upregulated.

[0227] In further embodiments, the human subject is identified as having differentiated monocyte-type AML based on the expression level of at least one marker selected from the group consisting of CD38, CD11b, and CD33 in the malignant myeloid cells of the human subject. In one embodiment, the expression level of CD38 is upregulated, the expression level of CD33 is upregulated, and the expression level of CD11b is upregulated.

[0228] In further embodiments, the human subject is identified as having differentiated monocyte-type AML based on the expression level of at least one marker selected from the group consisting of CD45, CD11b, and CD117 in the malignant myeloid cells of the human subject. In some embodiments, the expression level of CD45 is upregulated. In some embodiments, the expression level of CD45 is upregulated and the expression level of CD11b is upregulated. In some embodiments, the expression level of CD45 is upregulated and the expression level of CD117 is downregulated. In some embodiments, the expression level of CD45 is upregulated, the expression level of CD11b is upregulated and the expression level of CD117 is downregulated.

[0229] In further embodiments, the human subject is identified as having differentiated monocyte-type AML based on the CD45 expression level and the determining SSC value. In one embodiment, the cells are characterized as CD45 bright and SSC high.

[0230] In a specific embodiment, treatment with a BCL-2 inhibitor (e.g., venetoclax) in a clinical history upregulated CD70 expression in myeloid cells. Patients with myeloid malignancies who have failed treatment with BCL-2 can then be treated with an antibody that binds to CD70 or its antigen-binding fragment (e.g., xatuzumab). Treatment with an antibody that binds to CD70 or its antigen-binding fragment then upregulates BCL-2 expression in myeloid cells. Therefore, treatment with a BCL-2 inhibitor (e.g., venetoclax) and an antibody that binds to CD70 or its antigen-binding fragment (e.g., xatuzumab) has a synergistic effect in patients with myeloid malignancies, improving the treatment response in these patients. In a specific embodiment, an anti-CD70 antibody or its CD70-binding fragment is combined with a BCL-2 inhibitor (co-administered) for use in the treatment of myeloid malignancies in patients who are resistant to treatment with the BCL-2 inhibitor. In one embodiment, the CD70 expression level of malignant myeloid cells of the human subject is determined. The relevant expression level can be determined using any suitable method, including, but not limited to, fluorescence-activated cell sorting (FACS) and fluorescence microscopy using a detectable (e.g., fluorescently labeled) antibody specific to CD70.

[0231] In one embodiment, the patient's bone marrow sample contains CD45bright / SSChigh / CD38+ / CD34- / CD33+ / CD11b+ / CD70+ phenotype cells or CD45bright / SSChigh / CD34- / CD117- / CD11b+ / CD68+ / CD14+ / CD64+ phenotype cells.

[0232] All embodiments described herein in relation to therapeutic methods according to the above aspects of the present invention (see in particular Section B) are also applicable to these further aspects and embodiments of the present invention.

[0233] (D. Use for the manufacture of pharmaceuticals) In a further embodiment, the present invention provides the use of an antibody or antigen-binding fragment thereof that binds to CD70 for the manufacture of a pharmaceutical product. In particular, the pharmaceutical product is for the treatment of myeloid malignancies in human subjects, the subject being used specifically for the treatment specified by the method described herein.

[0234] In one embodiment, the use of a combination of a CD70-binding antibody or its antigen-binding fragment and a BCL-2 inhibitor is provided for the manufacture of a therapeutic drug for human myeloid malignancies as described herein.

[0235] In one embodiment, the use of a combination of a CD70-binding antibody or its antigen-binding fragment and a hypomethylating agent (HMA) is provided for the manufacture of a therapeutic drug for human myeloid malignancies as described herein.

[0236] In one embodiment, the use of a combination of a CD70-binding antibody or its antigen-binding fragment, a BCL-2 inhibitor, and a hypomethylating agent (HMA) is provided for the manufacture of a therapeutic drug for human myeloid malignancies as described herein.

[0237] All embodiments described herein in relation to therapeutic methods according to the above aspects of the present invention (see in particular sections B and C) are also applicable to these further aspects and embodiments of the present invention.

[0238] (E. Diagnostic Methods) Further aspects of the present invention relate to diagnostic methods. Thus, in one aspect, the present invention relates to a method for identifying a patient to be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy, and the method is: (i) to measure the differentiation status of the patient's myeloid cells, and, (ii) To determine whether the patient has differentiated monocytic AML, The present invention provides a method in which patients having differentiated monocytic AML, selected by a method including the steps, are identified as patients to be treated with the anti-CD70 antibody or its CD70-binding fragment.

[0239] In one embodiment, steps (i) and (ii) of the method are performed on a sample obtained from the patient and determined.

[0240] As used herein, a sample includes any tissue or fluid sample available from a patient with myeloid malignancy. The sample may be used to determine the differentiation status of the patient's myeloid cells. The sample may contain a detectable amount of a marker, preferably a monocyte-type cell marker. The term “sample” includes tissues, cells, and bodily fluids isolated from a subject, as well as tissues, cells, and fluids present within the subject. In some of these embodiments, the method is a method for determining the differentiation status of a patient's myeloid cells or detecting a marker in vitro. As used herein, “fluid” includes, for example, saliva, mucus, urine, blood, lymph, etc. In some embodiments, the sample includes serum, plasma, or other blood fractions or blood components or lymph obtained from the patient with myeloid malignancy. In other embodiments, the sample includes bone marrow obtained from the patient with myeloid malignancy.

[0241] Therefore, in a further embodiment, a method for identifying a patient to be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy and, (i) Measuring the differentiation status of myeloid cells in samples obtained from patients with the myeloid malignancy, (ii) To determine whether the sample has differentiated monocytic AML, The method is provided, which involves selecting a patient by the steps of the above, wherein the presence of differentiated monocytic AML in the sample identifies the patient from whom the sample was obtained as a patient who should be treated with the anti-CD70 antibody or its CD70-binding fragment.

[0242] In one embodiment, the myeloid cell differentiation status is determined according to the FAB classification system. The FAB system is a well-described and clinically relevant means for classifying AML patients by their differentiation status. This system classifies AML according to the type of cells from which leukemia originates and the maturity of those cells. In one embodiment, the myeloid cell differentiation status is AML-M5. In another embodiment, the myeloid cell differentiation status is AML-M4. In yet another embodiment, the myeloid cell differentiation status is determined according to the WHO classification system.

[0243] In a further embodiment, the level of differentiated monocytic AML in a sample obtained from a patient with myeloid malignancy can be compared to a predetermined cutoff value for the level of differentiated monocytic AML. This allows for evaluation of whether the level of differentiated monocytic AML in the patient is higher, lower, or not higher than the predetermined cutoff value. Such a comparison can be used to determine whether the patient should be selected for treatment with an anti-CD70 antibody or its antigen-binding fragment.

[0244] In a further embodiment, a method for identifying a patient to be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy, (i) Determining the level of differentiated monocytic AML in a sample obtained from a patient with the myeloid malignancy, (ii) Compare the level of differentiated monocytic AML in (i) with a predetermined cutoff value for differentiated monocytic AML. A method is provided in which a patient is selected for treatment with the anti-CD70 antibody or its CD70-binding fragment if the level of differentiated monocytic AML determined in the patient sample is higher than a predetermined cutoff value, and the patient is selected by a method comprising the steps of [step].

[0245] In another embodiment, a method for identifying a patient to be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy, (i) Determining the level of differentiated monocytic AML in a sample obtained from a patient with the myeloid malignancy, (ii) Compare the level of differentiated monocytic AML in (i) with a predetermined cutoff value for differentiated monocytic AML. A method is provided in which a patient is selected for treatment with the anti-CD70 antibody or its CD70-binding fragment if the patient is selected by a method including the steps of, and the level of differentiated monocytic AML determined in the patient sample is not higher than a predetermined cutoff value.

[0246] In another embodiment, a method is provided for identifying a patient to be treated with an anti-CD70 antibody or its antigen-binding fragment, the patient having a myeloid malignancy, and being selected by a method comprising the steps of (i) determining the level of differentiated monocytic AML in a sample obtained from the patient having the myeloid malignancy, and (ii) comparing the level of differentiated monocytic AML in (i) to a predetermined cutoff value for differentiated monocytic AML, wherein the patient is selected for treatment with the anti-CD70 antibody or its CD70-binding fragment if the level of differentiated monocytic AML determined in the patient sample is lower than the predetermined cutoff value.

[0247] In a further embodiment, a method for identifying a patient to be treated with an anti-CD70 antibody or its antigen-binding fragment, wherein the patient has a myeloid malignancy, (i) Determining the level of differentiated monocytic AML in a sample obtained from a patient with the myeloid malignancy, (ii) Compare the level of differentiated monocytic AML in (i) with a predetermined cutoff value for differentiated monocytic AML. A method is provided in which a patient is selected for treatment with the anti-CD70 antibody or its CD70-binding fragment if the patient is selected by a method including the steps of, and the level of differentiated monocytic AML determined in the patient sample is not lower than a predetermined cutoff value.

[0248] In some embodiments, a predetermined cutoff value for differentiated monocytic AML is the mean level of differentiated monocytic AML in a control cohort of AML patients. All embodiments described herein in relation to therapeutic methods according to the above embodiments of the present invention (see in particular sections B and C) are also applicable to these further embodiments and forms of the present invention.

[0249] (Inclusion of references) Various publications have been referenced in the preceding description and through the following examples, each of which is incorporated herein by reference in its entirety. [Examples]

[0250] (Examples) (Example 1) (Patients with monocytic AML disease are more likely to be refractory to venetoclax and azacitidine) To investigate whether differentiation status can predict responsiveness to venetoclax + azacitidine (VEN+AZA) in a clinical setting, we retrospectively re-examined 100 untreated patients who had previously received VEN+AZA and were subsequently diagnosed with AML. Several baseline factors were analyzed to determine their ability to predict disease as treatment-refractory (lack of complete remission (CR), CRi with incomplete peripheral blood cell count recovery, partial remission (PR), or morphologically leukemia-free state (MLFS) as defined by the European Leukemia Network (ELN); Dohner H et al., Blood 2017;129:424-47). The median age of the cohort was 72 years, 20 patients (20%) had a history of preceding hematological impairment, and 64 patients (64%) had disease with a poor prognosis according to ELN criteria.

[0251] To specifically examine features related to myeloid differentiation, we first employed the FAB (France, USA, UK) classification system. Although this system is no longer used clinically, it provides a well-described and clinically relevant means for classifying AML patients by myeloid differentiation status. In the VEN+AZA-treated patient cohort, 13 patients (13%) were identified as the FAB-M5 subtype, defined as the more differentiated monocytic AML phenotype, 8 patients (8%) were FAB-M4, and 77 patients (77%) were FAB-M0 or M1, indicating a less differentiated phenotype. Univariate analysis revealed that sex (P=0.0495), presence of RAS pathway mutations (P=0.0039), and FAB-M5 maturation stage (P<0.0001) were associated with refractory disease to VEN+AZA (Table 2). Multivariate analysis revealed that only the FAB-M5 maturation stage (P=0.0066) could predict refractory responses (Table 2). Specifically, 62% of FAB-M5 patients were refractory to VEN+AZA, compared to 0% of FAB-M4 patients and only 8% of non-FAB-M5 patients. Furthermore, the median overall survival was 89 days for FAB-M5 patients compared to 518 days for non-FAB-M5 patients (P=0.0039). These findings indicate a strong correlation between myeloid differentiation status and resistance to venetoclax-based therapy.

[0252] It is noteworthy that Kuusanmaki et al., (2020) Haematologica 105(3):708-720, reported, based on ex vivo testing, that the highest BCL2 / MCL1 gene expression ratio was observed in M0 / 1 and the lowest in M4 / 5 AML among all mononuclear cell fractions. This group further reported, based on ex vivo characterization and drug susceptibility testing, that gene expression data from mononuclear cell-enriched AML samples showed that M4 / 5 AML expressed low BCL2 and high MCL1 and BCL2A1, which is consistent with the decreased venetoclax sensitivity observed in all mononuclear cell fractions in M4 / 5 samples.

[0253] (Table 2. Baseline characteristics and univariate and multivariate logistic regression analyses of 100 untreated patients who had previously received VEN+AZA and were newly and consecutively diagnosed with AML)

Table 2

[0254] (Example 2)(Monocytic AML has intrinsic VEN+AZA resistance) To understand whether the lack of response to VEN+AZA in monocyte-type AML is due to an endogenous mechanism, we directly evaluated the in vitro sensitivity of VEN+AZA when protection from exogenous factors such as the microenvironment was minimal. Since the FAB system is no longer used clinically, we adopted phenotypic markers that would likely act as surrogates for the FAB-M5 subtype. Previous studies have shown that FAB-M5 patients lack expression of the early-type marker CD117 and upregulate the expression of monocyte-type markers CD11b, CD68, and CD64. References: Xu Y et al., (2006) Leukemia 20:1321-4; Garcia C et al., (2008) Appl Immunohistochem Mol Morphol 16:417-21; Cascavilla N et al., (1998) Haematologica 83:392-7; Di Noto R et al., (1996) Br J Haematol 92:562-4; Naeim F., "Atlas of Hematopathology: Morphology, Immunophenotype, Cytogenetics, and Molecular Approaches," 1st edition, London: Academic Press; 2013. pxi, 743 p. Therefore, we designed a multicolor flow cytometry panel including CD117, CD11b, CD68, and CD64 to distinguish monocytic AML (FAB-M5) patients from early-stage AML (FAB-M0 / M1 / M2) patients. As shown in Figure 1, this approach readily identified two major cell populations within AML patients. For example, patient 51 (Pt-51, typical FAB-M0 / M1 / M2) presented with a single dominant disease population phenotypically primitive, as demonstrated by CD45- medium / SSC-low / CD117+ / CD11b- / CD68- (Figure 1A). This patient achieved complete remission (CR) with VEN+AZA treatment.In contrast, Pt-72 (typical FAB-M5) was refractory to VEN+AZA, and presented a predominant monocytic disease with CD45-bright / SSC-high / CD117- / CD11b+ / CD68+ (Figure 1B). Further analysis of 12 primary AML specimens confirmed the phenotype profile of the primitive specimens versus monocytic specimens (Figure 1C). Hereinafter, these AMLs are referred to as "prim-AML" and "mono-AML", respectively.

[0255] Multiple studies have suggested that leukemic stem cells (LSCs) are key targets for AML therapy. Pollyea DA et al., (2017) Blood 129:1627-35. Previous studies have shown that the reactive oxygen species-low (ROS-low) phenotype enriches for functionally defined LSCs. Lagadinou ED et al., (2013) Cell Stem Cell 12:329-41; Pei S et al., (2018) Cell Stem Cell 23:86-100.e6. Therefore, to more directly evaluate drug response in the LSC subpopulation, we isolated low-ROS cells from prim-AML and mono-AML specimens. Since mono-AML has never been directly characterized by ROS levels, we confirmed in a colony-forming unit (CFU) assay that the low-ROS phenotype enriches potential stem / progenitor cells in mono-AML. These data indicate that the low-ROS phenotype strongly enriches potential stem / progenitor cells in mono-AML, consistent with what has been reported in prim-AML. Next, we treated the low-ROS subpopulation of prim-AML or mono-AML with VEN+AZA in vitro. The results showed that low-ROS LSCs from mono-AML specimens have significantly higher drug resistance than those from prim-AML specimens (Figure 1D), suggesting that at least part of the refractory response observed in FAB-M5 patients can be attributed to intrinsic molecular mechanisms inherent to monocytic AML cells. Figure 1 was adapted from Pei et al., (2020).

[0256] (Example 3) (Monocyte-type AML loses expression of venetoclax target BCL-2) Venetoclax is a BCL-2 specific inhibitor, and several studies have shown that BCL-2 expression strongly correlates with venetoclax sensitivity in vitro. (See Souers AJ et al., 2013; Nat Med 19:202-8; Pan R et al., 2014; Cancer Discov 4:362-75). Intergenetic analysis related to apoptosis regulation revealed a significant and consistent loss of BCL2 in mono-AML (N=5) compared to prim-AML (N=7) (Figure 2A). Analysis of the TCGA AML dataset also showed that loss of BCL2 gene expression progresses through the disease stages of morphological maturation of AML (from FAB-M0 to FAB-M5). As a result, significantly lower BCL2 expression was observed in FAB-M5 compared to FAB-M0 / M1 / M2 in the TCGA AML dataset (Figure 2B). Furthermore, decreased BCL-2 expression in mono-AML was also confirmed at the protein level (Figure 2C). Interestingly, BCL-2 loss also occurs during normal monocyte development. (References: Novershtern N et al., (2011) Cell 144:296-309; Lara-Astiaso D et al., (2014) Science 345:943-9). Consistent BCL-2 loss was observed at the monocyte stage in both human and mouse strains.

[0257] In summary, these analyses indicate that BCL-2 loss is a conserved biological feature during the development and growth of both normal and malignant monocytes. Furthermore, these data suggest that BCL-2 loss in monocyte-type AML may lead to resistance to venetoclax-based therapy. Figure 2 is adapted from Pei et al., (2020).

[0258] (Example 4) (Venetoclax and azacitidine (VEN+AZA) are selected for relapsed monocytic disease) Based on the above findings, we investigated the extent of apparent monocytic disease in patients who initially responded to VEN+AZA therapy but subsequently relapsed. Analysis of AML patients prior to VEN+AZA treatment revealed that the majority of patients showed a mixture of monocytic and primitive phenotypes, and were found to actually have tumors named "MMP-AML" (Mixed monocytic / primitive AML). The characteristics of two MMP-AML patients (Pt-12, Pt-65) were analyzed during treatment (Figures 3A and 3B). At relapse after initial complete remission, both patients showed near-complete disappearance of the primitive subpopulation and the emergence of a dominant monocytic phenotype (CD45-bright / SSC-high / CD117- / CD11b+ / CD68+). Therefore, VEN+AZA treatment was shown to induce a significant in vivo selection of the monocyte subpopulation in each patient (Figures 3A and 3B).

[0259] Notably, this monocyte-selective phenotype appears to be a clinical feature specific to VEN+AZA therapy. Indeed, previous analyses of patients treated with conventional chemotherapy have shown a consistent enrichment of an earlier LSC phenotype. Ho TC et al., (2016) Blood 128:1671-8.

[0260] To further confirm this finding, we analyzed RNA-seq data from 11 sets of diagnostic and relapsed specimens after conventional chemotherapy, from an isolation study by Shlush et al. (Shlush LI et al., 2017, Nature 547:104-8). In this setting, acquisition of the LSC gene expression signature, disappearance of monocyte-type markers (CD11b and CD68), and monocyte-type gene expression signature were observed at the time of relapse, suggesting that the myeloid phenotype is suppressed after chemotherapy.

[0261] Finally, we compared diagnostic and relapsed specimens from six AML patients treated with conventional chemotherapy. None of the cases showed a clear monocyte phenotype at the time of relapse. In fact, two patients who showed monocyte features at diagnosis showed a shift to an earlier phenotype at the time of relapse.

[0262] In summary, these data indicate that early-stage phenotypes are strongly promoted in relapses following conventional chemotherapy, and that the selection of monocyte phenotypes at the time of relapse is a notable feature of VEN+AZA therapy. Figure 3 is adapted from the literature by Pei et al. (2020).

[0263] (Example 5) (Treatment of patients with reduced sensitivity to venetoclax - Xatuzumab monotherapy) At least two adult human patients with AML who have reduced sensitivity to venetoclax or whose condition is refractory will be selected for the study. These patients will receive xatuzumab intravenously (iv) at a dose of approximately 10 mg / kg every 12–14 days. The number of blast cells in each patient will be measured before the start of xatuzumab administration (pre-treatment baseline) and then monitored approximately weekly for at least two weeks after the last or most recent dose of xatuzumab. Flow cytometry will be used to determine the ratio of blast cells to total cell number using the number of CD45dim, SSClow cells. A reduction of at least 5% in the number of blast cells from pre-treatment baseline will indicate the success of the therapeutic intervention.

[0264] (Example 6) (Treatment of patients with reduced sensitivity to venetoclax - Xatuzumab in combination with venetoclax) Two or more adult human patients with AML who have reduced sensitivity to or are refractory to venetoclax will be selected for the study. These patients will be administered xatuzumab intravenously (iv) at a dose of approximately 10 mg / kg every 12–14 days. From the second dose of xatuzumab onward, patients will be administered venetoclax orally (po) at a dose of 400–600 mg daily, starting with a first dose of 100 mg and increasing by 100 mg / day until the target daily dose of 400–600 mg is reached. The number of blast cells in the patients will be measured before the initiation of xatuzumab administration (pre-treatment baseline) and then monitored approximately weekly for at least two weeks after the last or most recent dose of xatuzumab. The ratio of blast cells to total cell number will be determined using flow cytometry, with CD45dim and SSClow cell counts. A reduction of at least 5% in blast count from pre-treatment baseline indicates success of the therapeutic intervention.

[0265] (Example 7) (Treatment of patients with reduced sensitivity to venetoclax - Xatuzumab in combination with venetoclax and azacitidine) Two or more adult human patients with AML who have reduced sensitivity to or are refractory to venetoclax will be selected for the study. These patients will be administered xatuzumab intravenously (iv) at a dose of approximately 10 mg / kg every 12–14 days. From the second dose of xatuzumab onward, patients will be administered venetoclax orally (po) at a dose of 400–600 mg daily, starting with a first dose of 100 mg and increasing by 100 mg / day until the target daily dose of 400–600 mg is reached, using an escalation schedule. Similarly, from the second dose of xatuzumab onward, patients will be administered azacitidine subcutaneously (sc) or ivally (IV) at a dose of 75 mg / m2 daily for 7 days, once in a 4-week cycle. Patient blast counts are measured before the initiation of xatuzumab administration (pre-treatment baseline), and then monitored approximately weekly for at least two weeks after the last or most recent xatuzumab dose. Flow cytometry is used to determine the ratio of blast cells to total cell number using CD45dim and SSClow cell counts. A reduction of at least 5% in blast count from pre-treatment baseline indicates success of the therapeutic intervention.

[0266] (Example 8) (Monocyte-type AML cells express significantly higher CD70 levels compared to less differentiated, early-type AML cells.) Analysis of CD70 mRNA expression revealed that CD70 expression at the transcriptional level was at least 6-fold higher on average in bone marrow samples from AML patients with the FAB M5 subtype (Figure 4), and these cells contained at least 80% of cells differentiated toward monocytic cells (monocytic AML). These monocytic AML cells differed phenotypicly from less differentiated AML cells (early-stage AML and mature AML, FAB M0-M2) and were classified as CD45bright / SSChigh / CD117- / CD11b+ / CD68+. This contrasts with early-stage AML cells showing the CD45medium / SSClow / CD117+ / CD11b- / CD68- phenotype in flow cytometry analysis (Pei et al., 2020). Analysis of the response to VEN+AZA combinations in FAB AML subtypes showed that monocytic blasts of the FAB M5 subtype were associated with refractory disease in response to VEN+AZA combinations. Specifically, 62% of FAB M5 patients and only 8% of non-FAB M5 patients were refractory to the VEN+AZA combination (Pei et al., 2020). Interestingly, the myelomonocytic FAB M4 subtype also shows elevated CD70 levels compared to less differentiated subtypes. Other authors have also noted that this subtype is associated with resistance to VEN+AZA (Zhang et al., 2020; Kuusanmaki et al., 2017). The FAB M4 subtype is a mixed phenotypic leukemia because it consists of a combination of clones of myeloid cell differentiation at different disease stages and at least 20% monocytic blasts. While the VEN+AZA combination therapy shows better efficacy in this subgroup, the monocytic AML cells present in this subgroup may increase the risk of early relapse (Zhang et al., 2020). Furthermore, both the M4 and M5 subtypes exhibit the lowest BCL2 / MCL1 gene expression ratio, which is associated with resistance to Bcl-2 inhibition (Kuusanmaki et al., 2020).

[0267] CD70 expression was tested for in bone marrow samples from patients with monocytic AML and mixed phenotype AML, and the phenotype of CD70-positive cells was confirmed by flow cytometry (Figure 5). Cytometric analysis confirmed high CD70 expression on the plasma membrane of malignant cells in VEN+AZA-resistant monocytic AML cells with the CD45bright / SSChigh / CD34- / CD117- / CD11b+ / CD68+ / CD14+ / CD64+ phenotype (Figure 5). Generally, monocytic disease samples showed the highest CD70 expression (Figure 5A), while early-type blast cells showed very limited CD70 expression. Occasionally, mixed phenotype samples consisting of monocytic leukemia cells and early-type leukemia cells showed relatively high CD70 expression in both monocytic AML cells and early-type AML cells, but early-type cells generally showed low CD70 expression on the cell surface. Figure 5B shows an example of a mixed phenotype sample with high CD70 expression on early-type AML cells and monocytic AML cells obtained from a VEN+AZA refractory patient.

[0268] Because AML samples often exhibit a mixed phenotype with different ratios of early-stage malignant cells and monocyte-type malignant cells, we compared CD70 expression in subpopulations of monocyte-type cells and early-stage cells. Flow cytometry analysis showed that the median fluorescence intensity of CD70 in monocyte-type AML cells was approximately six times higher than in early-stage AML cells. This confirmed high CD70 expression on monocyte-type AML cells (Figure 6A left), which was comparable to data obtained from CD70 mRNA analysis (Figure 4). Sample-by-sample pair analysis also showed higher CD70 expression levels on monocyte-type AML cells than on early-stage AML cells present in the same patient sample (Figure 6A right). Only a limited number of samples showed similarly high CD70 expression on both early-stage and monocyte-type cells. Data from protein expression levels were also confirmed by calculating the percentage of CD70-positive cells. On average, over 50% of monocytic AML cells in the samples showed high CD70 expression, while less than 10% of early-stage AML cells also showed CD70 expression (Figure 6B left). Furthermore, in 95% of the samples, the proportion of CD70-positive monocytic AML cells was higher than the average proportion of early-stage AML cells. Paired analysis also showed a higher proportion of CD70-positive monocytic malignant cells than early-stage AML cells in the same samples (Figure 6B right).

[0269] (Example 9) (CD70-positive, VEN+AZA-resistant monocytic AML cells are efficiently killed by xatuzumab-mediated ADCC) Xatuzumab is a defucosylated anti-human CD70 antibody with enhanced properties for mediating NK-dependent antibody-dependent cell-mediated cytotoxicity (ADCC) (Silence et al., 2014). VEN+AZA-resistant CD70-positive monocytic AML (CD45bright / SSChigh / CD38+ / CD34- / CD33+ / CD11b+ / CD70+), as well as mixed phenotype samples (Figure 7A) containing CD70-positive monocytic cells (CD45bright / SSChigh / CD38+ / CD34- / CD33+ / CD11b+ / CD70+) and CD70-negative VEN+AZA-sensitive early-stage cells (CD45medium / SSClow / CD34+ / CD33- / CD11b- / CD70-, including both CD38+ and CD38- populations), were tested for susceptibility to xatuzumab-mediated ADCC. Both types of primary bone marrow samples were treated with xatuzumab at a concentration of 10 μg / ml and co-incubated with human NK cells isolated by negative selection from healthy donor PBMCs. NK cells were added to monocytic AML bone marrow samples and mixed phenotypic bone marrow samples at target / effector cell (T:E) ratios of 1:5 and 1:15, respectively. The cells were co-cultured in a cell culture incubator at 37°C for 24 hours. Flow cytometry analysis was performed to measure the number of early-stage AML cells and monocytic AML cells and to estimate the ADCC level for specific samples. Monocytic cells in both samples were significantly targeted by xatuzumab-mediated NK cell-dependent ADCC (Figures 7B and 7C, respectively). When a blocking anti-CD70 41D12 FcDead antibody, which impairs effector function, was used as a negative control, no significant antibody-specific effect was detected with the blocking antibody in targeting CD70-positive monocytic cells (Figures 7B and 7C). This supports the specificity of the xatuzumab-mediated effect in targeting CD70-positive VEN+AZA-resistant monocytic AML cells.

[0270] (Example 10) (Xatuzumab effectively targets CD70-positive LSCs derived from VEN+AZA-resistant monocytic AML samples.) Low-ROS enriched leukemia stem cells (LSCs) derived from early-stage AML and monocytic AML exhibit significantly different characteristics. This is because low-ROS LSCs derived from monocytic AML are less dependent on the BCL2 protein for survival and have higher resistance to venetoclax (Pei et al., 2020). Transcriptome analysis of early-stage AML and monocytic AML samples, particularly a comparison of CD70 expression in a subpopulation enriched with low-ROS LSCs, showed that CD70 expression levels in low-ROS LSCs from monocytic disease were significantly higher than those in low-ROS LSCs derived from early-stage AML samples (Figure 8).

[0271] To investigate whether CD70-positive LSCs from VEN+AZA-resistant monocytic AML bone marrow samples could be targeted by xatuzumab, bone marrow samples from VEN+AZA-resistant samples were first incubated with xatuzumab (10 μg / ml) and NK cells isolated from healthy donor PBMCs (1:5, T:E ratio). After 24 hours of incubation, the samples were transferred to Mesocult medium and further cultured to confirm whether LSCs were targeted by xatuzumab-mediated ADCC. Untreated, human IgG1 isotype control, and blocking 41D12 FcDead antibody were used as negative controls. Cells were incubated in a 37°C cell culture incubator for 14 days, and colony-forming units (CFUs) were estimated by counting the number of grown colonies. Under the xatuzumab-treated conditions, a significant decrease in the number of grown colonies was observed compared to the untreated control (Figure 9). No significant effect was detected with either isotype controls or blocking anti-CD70 antibodies.

[0272] (Example 11) (Xatuzumab significantly reduces CD70-positive, VEN+AZA-resistant monocyte-type AML cells in patient-derived xenograft mouse models via an NK-dependent mechanism.) Patient samples injected into NSGS mice engraft in the mouse bone marrow. The efficacy of anti-leukemia compounds can be rigorously measured using a therapeutic approach by determining the reduction of malignant cells in the mouse bone marrow after full engraftment of the patient-derived samples. Cells from bone marrow derived from VEN+AZA-resistant monocytic AML were engrafted at 2×10⁶ cells per NSGS mouse over 42 days. One cohort of animals was treated with vehicle, the combination of 100 mg / kg venetoclax (Ven) and 3 mg / kg azacitidine (Aza), or 10 mg / kg cusatuzumab. A second cohort was first injected with 1.5×10⁶ NK cells isolated from PBMCs of healthy donors, then treated with the same drug combination as the first cohort. Animals were treated every 3 days with vehicle, the VEN+AZA combination, or cusatuzumab. One day after the third administration, animals were sacrificed, bone marrow was isolated from femurs, and samples were analyzed by flow cytometry to determine the number of monocytic AML cells. Flow cytometry analysis showed a significant reduction of malignant human CD45+CD11b+CD117- cells in animals treated with cusatuzumab in the presence of human NK cells. However, in the absence of human NK cells, no significant effect was observed for either VEN+AZA or cusatuzumab, and no significant effect was observed for VEN+AZA in the presence of NK cells (Figure 10). Therefore, cusatuzumab is effective at depleting VEN+AZA-resistant CD70-positive monocytic AML cells via NK-dependent ADCC in vivo in NSGS mice.

[0273] (References) [Table 3]

Claims

1. A pharmaceutical composition for treating monocytic acute myeloid leukemia (AML) in human subjects resistant to treatment with BCL-2 inhibitors, comprising an antibody that binds to CD70 or an antigen-binding fragment thereof.

2. The pharmaceutical composition according to claim 1, wherein the human subject has differentiated monocyte-type AML.

3. Before the treatment, (i) A step of measuring the differentiation state of bone marrow cells of the human subject, and (ii) A step of determining whether or not the human subject has differentiated monocytic AML, A selection is made, and a therapeutically effective dose of the anti-CD70 antibody or its anti-CD70 conjugate fragment is administered to a human subject having differentiated monocytic AML, according to claim 1 or 2.

4. The pharmaceutical composition according to claim 2 or 3, wherein the human subject is identified as having differentiated monocyte-type AML based on the differential expression level (or more) of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, CD70, BCL2A1, and MCL1 in the malignant myeloid cells of the human subject.

5. The pharmaceutical composition according to claim 4, wherein the human subject is identified as having differentiated monocytic AML when at least one expression level(s) of BCL-2 and CD117 is downregulated and / or at least one expression level(s) of CD11b, CD68, CD64, CD70, BCL2A1, and MCL1 is upregulated.

6. The pharmaceutical composition according to claim 3, wherein in step (i), the CD70 expression level of malignant myeloid cells of the human subject is measured.

7. The pharmaceutical composition according to claim 6, wherein CD70 is upregulated in malignant myeloid cells of a human subject compared to the CD70 expression level measured before or during treatment with a BCL-2 inhibitor.

8. The pharmaceutical composition according to any one of claims 1 to 7, wherein a hypomethylating agent (HMA) is co-administered with an antibody or an antigen-binding fragment thereof that binds to CD70.

9. The pharmaceutical composition according to claim 8, wherein the HMA is selected from the group consisting of azacitidine, decitabine, and guadecitabine.

10. The pharmaceutical composition according to any one of claims 1 to 9, wherein the BCL-2 inhibitor is co-administered with an antibody or antigen-binding fragment thereof that binds to CD70.

11. The antibody or antibody-binding fragment that binds to CD70 includes HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, The amino acid sequence of HCDR1 is sequence number 1; The amino acid sequence of HCDR2 is sequence number 2; The amino acid sequence of HCDR3 is sequence number 3; The amino acid sequence of LCDR1 is sequence number 4; The amino acid sequence of LCDR2 consists of sequence number 5; and, The amino acid sequence of LCDR3 is as follows: A pharmaceutical composition according to any one of claims 1 to 10.

12. The pharmaceutical composition according to any one of claims 1 to 11, wherein the antibody or antibody-binding fragment that binds to CD70 comprises a variable heavy chain domain (VH) having at least 90% the same amino acid sequence as SEQ ID NO: 7, and a variable light chain domain (VL) having at least 90% the same amino acid sequence as SEQ ID NO:

8.

13. The pharmaceutical composition according to claim 12, wherein the antibody or antibody-binding fragment that binds to CD70 comprises a variable heavy chain domain (VH) having the same amino acid sequence as SEQ ID NO: 7 and a variable light chain domain (VL) having the same amino acid sequence as SEQ ID NO:

8.

14. The amino acid sequence that is at least 90% identical to VH consisting of Sequence ID No. 7 comprises HCDR1, HCDR2, and HCDR3, The amino acid sequence of HCDR1 is sequence number 1; The amino acid sequence of HCDR2 consists of sequence number 2; and, The amino acid sequence of HCDR3 consists of sequence number 3; and, The amino acid sequence that is at least 90% identical to the VL consisting of Sequence ID No. 8 includes LCDR1, LCDR2, and LCDR3. The amino acid sequence of LCDR1 is sequence number 4; The amino acid sequence of LCDR2 consists of sequence number 5; and, The amino acid sequence of LCDR3 is sequence number 6. The pharmaceutical composition according to claim 13.

15. The pharmaceutical composition according to any one of claims 1 to 14, wherein the BCL-2 inhibitor is venetoclax or a pharmaceutically acceptable salt thereof.

16. The pharmaceutical composition according to any one of claims 1 to 15, wherein the antibody that binds to CD70 is xatuzumab.

17. A method for determining susceptibility to the treatment of monocytic acute myeloid leukemia (AML) with an antibody that binds to CD70 or an antigen-binding fragment thereof, comprising measuring the expression level of at least one marker selected from the group consisting of BCL-2, CD117, CD11b, CD68, CD64, BCL2A1, and MCL1 in malignant myeloid cells isolated from a human subject.

18. The method according to claim 17, wherein the human subject is identified as sensitive to treatment with an antibody or antigen-binding fragment thereof that binds to CD70 when at least one expression level(s) of BCL-2 and CD117 is downregulated and / or at least one expression level(s) of CD11b, CD68, CD64, CD70, BCL2A1, and MCL1 is upregulated.

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