Bispecific antibodies against cd3 and cd20 for treatment of leirscht's syndrome

Abstract

Methods for the clinical treatment of Richt's Syndrome in a human subject using bispecific antibodies that bind to CD3 and CD20 are provided.

Description

Field of the Invention

[0001] The present invention relates to the use of bispecific antibodies that target both CD3 and CD20 in the treatment of Richter's syndrome (RS). Advantageous treatment regimens are also provided. Background of the Invention

[0003] Chronic lymphocytic leukemia (CLL) is a B-cell malignancy that arises from the uncontrolled proliferation of immature lymphocytes in the bone marrow and involves circulating tumor cells in the blood. CLL is characterized by the accumulation of clonal CD5+CD19+CD20+CD23+ B cells in the bone marrow, blood, and lymphoid organs such as lymph nodes and spleen (Zenz et al., Nat Rev Cancer 2010;10:37-50). CLL is generally a slow-growing cancer. CLL is primarily a disease of older adults, with a median age at diagnosis of 70 years. CLL is the most common leukemia in adults in Western countries, accounting for approximately 25% to 30% of all leukemias in the US, with an estimated 20,720 new cases and 3,930 deaths (Siegel et al., CA Cancer J Clin 2019;69:7-34). Worldwide, there are approximately 105,000 cases per year, with 35,000 deaths (Global Burden of Disease Cancer, Fitzmaurice et al., JAMA Oncol 2018;4:1553-68).

[0004] In contrast, lymphoma arises from the uncontrolled proliferation of lymphocytes in organs outside the bone marrow. Although in some lymphomas, the bone marrow may also have tumor cell infiltration. Lymphoma cells generally will not appear in the peripheral blood.

[0005] Richter’s syndrome (RS), also known as Richter’s transformation (RT), is the development of an aggressive lymphoma that occurs in the context of CLL or SLL (small lymphocytic lymphoma) (Swerdlow et al., 2017; WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. International Agency for Research on Cancer, Lyon, France). It has been estimated that the annual incidence of RS in patients with CLL is approximately 0.5% to 1%, with an overall incidence of approximately 5% to 16% in all patients with CLL (Rossi et al., Br J Haematol, 2008; 142, 202-215). In a recent study, it was found that approximately 2%-10% of all CLL / SLL patients develop Richter’s syndrome during their disease course (Wang Y, et al. Haematologica. 2020;105:765-73). Approximately 90% of the time, RS presents as diffuse large B-cell lymphoma (RS-DLBCL) (Parikh et al., Br J Haematol, 2013; 162, 774-782; Rossi et al., Br J Haematol, 2008; 142,202-215). Approximately 80% of RS-DLBCL is associated with the original CLL clone and has historically been chemo-refractory, with a median survival of 12 months (Eyre et al., Br J Haematol, 2016; 175, 43-54.; Langerbeins et al., Am J Hematol 89, 2014; E239-243; Rogers et al., Br J Haematol, 2018; 180,259-266; Tsimberidou et al., Clin Lymphoma Myeloma Leuk, 2013; 13, 568-574). Monotherapy with novel agents such as BTK inhibitors and BCL2 inhibitors has little changed the outcome of RS. In 29 patients with RS-DLBCL treated with acalabrutinib, the median ORR, duration of response, and median PFS were 38%, 5 months, and 3 months, respectively (Hillmen et al., Blood, 2016; 128, 60-60). In 7 patients with RS-DLBCL, venetoclax achieved an ORR of 43%, with an unknown duration.Checkpoint inhibitors such as pembrolizumab and nivolumab, as monotherapy or in combination with a BTK inhibitor, have demonstrated an ORR ranging from 40% to 60%, with a relatively short PFS of 4 months (Ding et al., Blood 2017;129, 3419-3427; Jain et al., Blood 2016; 128, 59-59; Younes et al., Blood 2017;130, 833-833). Due to the short duration of response to chemoimmunotherapy, autologous and allogeneic stem cell transplantation has been used as post-induction therapy to extend survival in eligible / young RS patients. However, approximately 80% to 90% of RS patients are not eligible for transplantation due to: 1) inability to achieve remission through induction therapy; 2) comorbidities or age. In summary, in the era of novel agents, the incidence of RS has not decreased, and the prognosis of subjects with RS remains poor.

[0006] Accordingly, there remains an unmet need for treatment options for patients with Richter's syndrome. SUMMARY OF THE INVENTION

[0008] Provided herein are methods of treating human subjects exhibiting Richter's syndrome by administering a bispecific antibody that binds to CD3 and CD20, and particularly advantageous clinical treatment regimens.

[0009] In one aspect, provided herein is a method of treating Richter's syndrome in a human subject, the method comprising administering (e.g., subcutaneously) to the subject an effective amount of a bispecific antibody (e.g., epcoritamab), the bispecific antibody comprising:

[0010] (i) a first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a variable heavy chain (VH) region and a variable light chain (VL) region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences of the VH region sequence of SEQ ID NO: 6, and the VL region comprises the CDR1, CDR2, and CDR3 sequences of the VL region sequence of SEQ ID NO: 7; and

[0011] (ii) a second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region and a VL region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences of the VH region sequence of SEQ ID NO: 13, and the VL region comprises the CDR1, CDR2, and CDR3 sequences of the VL region sequence of SEQ ID NO: 14;

[0012] The bispecific antibody is administered at a dose in the range of 12 - 60 mg over a 28-day cycle. In some embodiments, the bispecific antibody is administered at a dose of 12 mg, 24 mg, 48 mg, or 60 mg (or a dose of about 12 mg, 24 mg, 48 mg, or 60 mg). In some embodiments, the bispecific antibody is administered at a dose of 12 mg (or a dose of about 12 mg). In some embodiments, the bispecific antibody is administered at a dose of 24 mg (or a dose of about 24 mg). In some embodiments, the bispecific antibody is administered at a dose of 48 mg (or a dose of about 48 mg). In some embodiments, the bispecific antibody is administered at a dose of 60 mg (or a dose of about 60 mg).

[0013] In some embodiments, the bispecific antibody is administered once a week, for example, for 2.5 28-day cycles (i.e., on days 15 and 22 of cycle 1, and on days 1, 8, 15, and 22 of cycles 2 - 3). In some embodiments, the bispecific antibody is administered once every two weeks after a once-a-week administration, for example, for 6 28-day cycles. In some embodiments, the bispecific antibody is administered once every four weeks after a once-every-two-weeks administration. In another embodiment, a sensitizing dose of the bispecific antibody (e.g., 0.05 - 0.35 mg, e.g., 0.16 mg or about 0.16 mg) is administered two weeks before a once-a-week dose of 24 mg or 48 mg. In a further embodiment, the sensitizing dose is administered one week before an intermediate dose, and the intermediate dose is administered one week before the first once-a-week dose of 24 mg or 48 mg.

[0014] In some embodiments, the bispecific antibody is administered over a 28-day cycle, wherein:

[0015] a) In cycle 1, a sensitizing dose (e.g., 0.05 - 0.35 mg, e.g., 0.16 mg or about 0.16 mg) is administered on day 1, an intermediate dose (e.g., 0.6 - 1.2 mg, e.g., 0.8 mg or about 0.8 mg) is administered on day 8, and a full dose of 12 - 60 mg (e.g., 12 mg, 24 mg, 48 mg, or 60 mg) is administered on days 15 and 22;

[0016] b) In cycles 2 - 3, a full dose of 12 - 60 mg (e.g., 12 mg, 24 mg, 48 mg, or 60 mg) is administered on days 1, 8, 15, and 22;

[0017] c) During cycles 4 - 9, administer a full dose of 12 - 60 mg (such as 12 mg, 24 mg, 48 mg, or 60 mg) on days 1 and 15; and

[0018] d) During cycle 10 and subsequent cycles, administer a full dose of 12 - 60 mg (such as 12 mg, 24 mg, 48 mg, or 60 mg) on day 1.

[0019] In a further embodiment, the subject has refractory and / or relapsed Richter's syndrome following two prior anti - neoplastic therapies.

[0020] In some embodiments, the subject is treated with prophylaxis against cytokine release syndrome (CRS). In some embodiments, the prophylaxis includes administering a corticosteroid (such as prednisolone, at a dose of, for example, 100 mg or its equivalent, including an oral dose) on, for example, the same day as the bispecific antibody. In some embodiments, corticosteroids are further administered on the second, third, and fourth days after administration of the bispecific antibody.

[0021] In some embodiments, the subject is pre - medicated with, such as an antihistamine (such as diphenhydramine, at a dose of, for example, 50 mg or its equivalent, intravenously or orally) and / or an antipyretic (such as acetaminophen, at a dose of, for example, 560 - 1000 mg) to reduce the reaction to the injection. In some embodiments, the pre - medication is administered on the same day as the bispecific antibody.

[0022] In some embodiments, the prophylaxis and pre - medication are administered during cycle 1. In some embodiments, when the subject experiences CRS greater than grade 1 after the last administration of the bispecific antibody in cycle 1, the prophylaxis is administered during cycle 2. In some embodiments, when the subject experiences CRS greater than grade 1 during the last administration of the bispecific antibody in a previous cycle, the prophylaxis is continued in subsequent cycles. In another embodiment, the pre - medication is administered during cycle 2. In another embodiment, the pre - medication is administered during subsequent cycles.

[0023] In some embodiments, if a subject develops grade 1 CRS, the subject is treated with antipyretics and hydration. In some embodiments, if a subject develops grade 2 CRS, the subject is treated with tocilizumab and / or dexamethasone or its methylprednisolone equivalent. In some embodiments, if a subject develops grade 3 CRS, the subject is treated with tocilizumab and dexamethasone (e.g., at a dose of 10 - 20 mg or its methylprednisolone equivalent, e.g., administered every 6 hours). In another embodiment, if a subject develops grade 4 CRS, the subject is treated with tocilizumab and methylprednisolone (e.g., at a dose of 1000 mg / day). In another embodiment, if a subject does not respond to tocilizumab, tocilizumab is switched to siltuximab.

[0024] In some embodiments, prophylaxis against tumor lysis syndrome (TLS) is administered to a subject. In some embodiments, prophylaxis against TLS includes administering one or more urate-lowering agents prior to administration of the bispecific antibody. In some embodiments, allopurinol and rasburicase are administered as urate-lowering agents. In a further embodiment, allopurinol is administered at least 72 hours prior to administration of the bispecific antibody. In a further embodiment, rasburicase is administered after administration of allopurinol and prior to administration of the bispecific antibody. In some embodiments, when a subject exhibits signs of TLS, supportive therapies such as rasburicase and / or allopurinol may be used.

[0025] In some embodiments, a subject treated by the methods described herein achieves a complete response, partial response, or disease stabilization, e.g., as defined by the Lugano criteria (Cheson et al., 2014).

[0026] In some embodiments, the first antigen-binding region of the bispecific antibody comprises VHCDR1, VHCDR2, and VHCDR3 comprising the amino acid sequences set forth in SEQ ID NO: 1, 2, and 3, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprising the amino acid sequences set forth in SEQ ID NO: 4, the sequence GTN, and SEQ ID NO: 5, respectively; and the second antigen-binding region comprises VHCDR1, VHCDR2, and VHCDR3 comprising the amino acid sequences set forth in SEQ ID NO: 8, 9, and 10, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprising the amino acid sequences set forth in SEQ ID NO: 11, the sequence DAS, and SEQ ID NO: 12, respectively.

[0027] In some embodiments, the first antigen-binding region of the bispecific antibody comprises a VH region and a VL region, the VH region comprising the amino acid sequence of SEQ ID NO: 6 and the VL region comprising the amino acid sequence of SEQ ID NO: 7; and the second antigen-binding region comprises a VH region and a VL region, the VH region comprising the amino acid sequence of SEQ ID NO: 13 and the VL region comprising the amino acid sequence of SEQ ID NO: 14.

[0028] In some embodiments, the first binding arm of the bispecific antibody is derived from a humanized antibody, preferably from a full-length IgG1, λ (lambda) antibody. In some embodiments, the second binding arm of the bispecific antibody is derived from a human antibody, preferably from a full-length IgG1, κ (kappa) antibody. In some embodiments, the bispecific antibody is a full-length antibody with a human IgG1 constant region.

[0029] In some embodiments, the bispecific antibody comprises an inert Fc region, e.g., an Fc region in which the amino acids at positions corresponding to L234, L235, and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E, and A, respectively. In some embodiments, the bispecific antibody comprises substitutions that promote bispecific antibody formation, e.g., in which the amino acid at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 in the first heavy chain is L, and in the second heavy chain, the amino acid at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa. In some embodiments, the bispecific antibody has both an inert Fc region (e.g., substitutions at L234, L235, and D265 such as L234F, L235E, and D265A) and substitutions that promote bispecific antibody formation (e.g., F405L and K409R). In a further embodiment, the bispecific antibody comprises a heavy chain constant region comprising the amino acid sequences of SEQ ID NO: 19 and 20.

[0030] In some embodiments, the bispecific antibody comprises a first heavy chain and a first light chain comprising (or consisting of) the amino acid sequences listed in SEQ ID NO: 24 and 25, respectively, and a second heavy chain and a second light chain comprising (or consisting of) the amino acid sequences listed in SEQ ID NO: 26 and 27, respectively. In some embodiments, the bispecific antibody is acrivastat or a biosimilar thereof. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1: Results from a Phase 1b / 2 Trial: Treatment Emergent Adverse Events; Data Cutoff: September 8, 2022.

[0032] Figure 2 : Results from a Phase 1b / 2 Trial: CRS Events by Administration Period; Data Cutoff: September 8, 2022.

[0033] Figure 3 : Results from a Phase 1b / 2 Trial: Depth and Duration of Response; Data Cutoff September 8, 2022.

[0034] Figure 4 : Results from a Phase 1b / 2 Trial; Tumor Reduction Relative to Baseline; Data Cutoff: September 8, 2022.

[0035] Figure 5 : Clinical Case Study (RS-DLBCL). A: Baseline PET / CT scan, mesenteric mass: 11.6 x 7.2 cm, maximum standardized uptake value (SUV) of abdominal / pelvic mass 6.3; B: 12-week PET / CT scan, mesenteric mass: 0.5 x 0.5 cm, maximum SUV in abdominal / pelvic mass 2.0; C: 76-week PET / CT scan, mesenteric mass 0.5 x 0.5 cm, only background uptake. Detailed Description of the Invention

[0037] Definitions

[0038] As used herein, the term "immunoglobulin" refers to a class of structurally related glycoproteins that consists of two pairs of polypeptide chains, one pair of light (L) low molecular weight chains and one pair of heavy (H) chains, all four of which are interconnected by disulfide bonds. The structure of immunoglobulins has been well characterized (see, e.g., Fundamental Immunology Ch. 7 (Paul, W. ed., 2nd ed. Raven Press, N.Y. (1989))). Briefly, each heavy chain typically contains a heavy chain variable region (abbreviated herein as VH or V H ) and a heavy chain constant region (abbreviated herein as CH or C H ). The heavy chain constant region typically contains three domains CH1, CH2, and CH3. The hinge region is the region between the CH1 and CH2 domains of the heavy chain and is highly flexible. The disulfide bonds in the hinge region are part of the interaction between the two heavy chains in an IgG molecule. Each light chain typically contains a light chain variable region (abbreviated herein as VL or V L ) and a light chain constant region (abbreviated herein as CL or C L)。The light chain constant region typically contains one domain CL. The VH and VL regions can be further subdivided into hypervariable regions (or HV regions, which can be hypervariable in the form of sequence and / or structure-defined loops), also known as complementarity-determining regions (CDRs), interspersed with more conserved regions, called framework regions (FRs). Each VH and VL is typically composed of three CDRs and four FRs, arranged in the following order from the amino terminus to the carboxy terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4 (see also Chothia and Lesk J MolBiol 1987;196:901-17). Unless otherwise stated or inconsistent with the context, the CDR sequences herein are identified according to the IMGT rules (Brochet X., Nucl Acids Res 2008;36:W503-508; Lefranc MP., Nucl Acids Res 1999;27:209-12; www.imgt.org / ). Unless otherwise stated or inconsistent with the context, references to amino acid positions in the constant region are according to EU numbering (Edelman et al., PNAS. 1969; 63:78-85; Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed 1991 NIH Publication No. 91-3242). For example, according to EU numbering, SEQ ID NO: 15 shows the amino acid positions 118-447 of the IgG1 heavy chain constant region.

[0039] As used herein, the term "amino acid corresponding to position..." refers to the amino acid position number in the human IgG1 heavy chain. The corresponding amino acid positions in other immunoglobulins can be found by alignment with human IgG1. Thus, an amino acid or segment in one sequence that "corresponds to" an amino acid or segment in another sequence is the amino acid or segment that is typically aligned with the other amino acid or segment using a standard sequence alignment program (such as ALIGN, ClustalW or a similar program) with default settings and has at least 50%, at least 80%, at least 90% or at least 95% identity to the human IgG1 heavy chain. Aligning sequences or segments of sequences and thereby determining the positions in a sequence that correspond to the amino acid positions according to the present invention is within the ability of one of ordinary skill in the art.

[0040] As used herein, in the context of the present invention, the term "antibody" (Ab) refers to an immunoglobulin molecule that has the ability to specifically bind an antigen under typical physiological conditions and has a half-life over a significant period of time, such as at least about 30 minutes, at least about 45 minutes, at least about 1 hour, at least about 2 hours, at least about 4 hours, at least about 8 hours, at least about 12 hours, about 24 hours or longer, about 48 hours or longer, about 3, 4, 5, 6, 7 days or more, or any other relevant functional defined period (such as a time sufficient to induce, promote, enhance, and / or regulate a physiological response associated with antibody binding to an antigen and / or a time sufficient to enable the antibody to recruit effector activity). The variable regions of the heavy and light chains of the immunoglobulin molecule contain binding domains that interact with the antigen. Unless otherwise specified, the term antibody also encompasses polyclonal antibodies, monoclonal antibodies (mAbs), antibody-like polypeptides, chimeric antibodies, and humanized antibodies. The antibodies generated can be of any isotype.

[0041] As used herein, the term "antibody fragment" or "antigen-binding fragment" refers to a fragment of an immunoglobulin molecule that retains the ability to specifically bind an antigen and can be generated by any known technique, such as enzymatic cleavage, peptide synthesis, and recombinant techniques. Examples of antibody fragments include (i) Fab' or Fab fragments, monovalent fragments consisting of the VL, VH, CL, and CH1 domains, or monovalent antibodies as described in WO2007059782 (Genmab); (ii) F(ab') 2 fragments, bivalent fragments containing two Fab fragments linked by a disulfide bond in the hinge region; (iii) Fd fragments, consisting essentially of the VH and CH1 domains; (iv) Fv fragments, consisting essentially of the VL and VH domains of a single arm of an antibody; (v) dAb fragments (Ward et al., Nature 1989; 341: 54446), which consist essentially of the VH domain and are also referred to as domain antibodies (Holt et al.; Trends Biotechnol 2003; 21: 484-90); (vi) camelid or nanobodies (Revets et al.; Expert Opin Biol Ther 2005; 5: 111-24) and (vii) isolated complementarity-determining regions (CDRs). In addition, although the two domains VL and VH of the Fv fragment are encoded by separate genes, they can be joined using recombinant methods by a synthetic linker to enable them to be made as a single protein chain, in which the VL and VH regions pair to form a monovalent molecule (referred to as a single-chain antibody or single-chain Fv (scFv), see, for example, Bird et al., Science 1988; 242:42326, and Huston et al., PNAS 1988;85:587983). Such single-chain antibodies are encompassed by the term antibody fragment, unless otherwise specified or the context clearly dictates otherwise.

[0042] As used herein, the term "antibody binding region" or "antigen binding region" refers to the region that interacts with an antigen and contains both the VH region and the VL region. When used herein, the term antibody refers not only to monospecific antibodies but also to multispecific antibodies that contain multiple, such as two or more, for example three or more, different antigen binding regions. Unless otherwise specified or clearly inconsistent with the context, the term antigen binding region includes antibody fragments that are antigen binding fragments, i.e., retain the ability to specifically bind an antigen.

[0043] As used herein, the term "isotype" refers to the immunoglobulin class encoded by the heavy chain constant region gene (e.g., IgG1, IgG2, IgG3, IgG4, IgD, IgA, IgE, or IgM). When referring to a specific isotype such as IgG1, the term is not limited to a specific isotype sequence such as a specific IgG1 sequence, but is used to indicate that the antibody is more closely related in sequence to that isotype such as IgG1 than to other isotypes. Thus, for example, an IgG1 antibody can be a sequence variant of a naturally occurring IgG1 antibody, which may include variations in the constant region.

[0044] As used herein, the term "bispecific antibody" or "bs" or "bsAb" refers to an antibody that has two different antigen binding regions defined by different antibody sequences. Bispecific antibodies can be in any form.

[0045] As used herein, the terms "half molecule", "Fab-arm", and "arm" refer to a heavy chain-light chain pair.

[0046] When a bispecific antibody is described as comprising a half molecule antibody "derived from" a first parental antibody and a half molecule antibody "derived from" a second parental antibody, the term "derived from" indicates that the bispecific antibody is generated by recombining the half molecules from each of the first and second parental antibodies into the resulting bispecific antibody by any known method. In this context, "recombination" is not intended to be limited by any particular recombination method and thus includes all methods for generating the bispecific antibodies described herein, including, for example, recombination by half molecule exchange (also known as "controlled Fab-arm exchange"), and recombination at the nucleic acid level and / or by co-expression of the two half molecules in the same cell.

[0047] As used herein, in the context of an antibody, the term "full-length" indicates that the antibody is not a fragment, but rather contains all of the domains of that isotype typically found in nature for that particular isotype. For example, a VH, CH1, CH2, CH3, hinge, VL, and CL domains of an IgG1 antibody. A full-length antibody can be engineered. An example of a "full-length" antibody is acalabrutinib.

[0048] As used herein, the term "Fc region" refers to the antibody region consisting of the Fc sequences of the two heavy chains of an immunoglobulin, wherein the Fc sequence comprises at least a hinge region, a CH2 domain, and a CH3 domain.

[0049] As used herein, the term "heterodimeric interaction between the first and second CH3 regions" refers to the interaction between the first CH3 region and the second CH3 region in a first CH3 / second CH3 heterodimeric protein.

[0050] As used herein, the term "homodimeric interaction of the first and second CH3 regions" refers to the interaction between the first CH3 region and another first CH3 region in a first CH3 / first CH3 homodimeric protein and the interaction between the second CH3 region and another second CH3 region in a second CH3 / second CH3 homodimeric protein.

[0051] As used herein, in the context of an antibody binding to a predetermined antigen, the term "binding" generally refers to binding having an affinity corresponding to a K D of about 10 -6 M or less, such as 10 -7 M or less, such as about 10 -8 M or less, such as about 10 -9 M or less, about 10 -10 M or less or about 10 -11 M or less, as determined by, for example, biolayer interferometry (BLI) technology in an Octet HTX instrument using the antibody as a ligand and the antigen as an analyte, and wherein the antibody binds to the predetermined antigen with an affinity corresponding to a K D that is at least ten-fold lower, such as at least 100-fold lower, for example at least 1,000-fold lower, such as at least 10,000-fold lower, for example at least 100,000-fold lower, than its K D for binding to a non-specific antigen other than the predetermined antigen or a closely related antigen (e.g., BSA, casein). The amount with a lower binding K D depends on the K D of the antibody, such that when the K D of the antibody is very low, then the K D for binding to the antigen is lower than the KD A low amount may be at least 10,000-fold (i.e., the antibody is highly specific).

[0052] As used herein, the term "K" D "(M) refers to the dissociation equilibrium constant of a particular antibody-antigen interaction. As used herein, affinity and K D are negatively correlated, i.e., a higher affinity is intended to mean a lower K D and a lower affinity is intended to mean a higher K D .

[0053] As used herein, the term "isolated antibody" refers to an antibody that is substantially free of other antibodies having different antigen specificities. In a preferred embodiment, an isolated bispecific antibody that specifically binds to CD20 and CD3 is additionally substantially free of monospecific antibodies that specifically bind to CD20 or CD3.

[0054] As used herein, the term "CD3" refers to the human cluster of differentiation 3 protein, which is part of the T cell coreceptor protein complex and is composed of four different chains. CD3 also exists in other species, and thus, the term "CD3" is not limited to human CD3 unless inconsistent with the context. In mammals, the complex contains the CD3γ (gamma) chain (human CD3γ chain UniProtKB / Swiss-Prot No P09693, or cynomolgus monkey CD3γ UniProtKB / Swiss-Prot No Q95LI7), the CD3δ (delta) chain (human CD3δ UniProtKB / Swiss-Prot No P04234, or cynomolgus monkey CD3δ UniProtKB / Swiss-Prot No Q95LI8), two CD3ε (epsilon) chains (human CD3ε UniProtKB / Swiss-Prot No P07766, SEQ ID NO: 28; cynomolgus monkey CD3ε UniProtKB / Swiss-Prot No Q95LI5; or rhesus monkey CD3ε UniProtKB / Swiss-Prot No G7NCB9) and the CD3ζ chain (zeta) chain (human CD3ζ UniProtKB / Swiss-Prot No P20963, cynomolgus monkey CD3ζ UniProtKB / Swiss-Prot No Q09TK0). These chains associate with a molecule called the T cell receptor (TCR) and generate an activation signal in T lymphocytes. The TCR and CD3 molecules together constitute the TCR complex.

[0055] As used herein, the term "CD3 antibody" or "anti-CD3 antibody" refers to an antibody that specifically binds to the antigen CD3, particularly human CD3ε (epsilon).

[0056] The term "human CD20" or "CD20" refers to human CD20 (UniProtKB / Swiss-Prot No P11836, SEQ ID NO:29) and includes any variants, isotypes, and species homologs of CD20 that are naturally expressed by cells (including tumor cells) or expressed on cells transfected with the CD20 gene or cDNA. Species homologs include rhesus CD20 (Macaca mulatta; UniProtKB / Swiss-Prot No H9YXP1) and cynomolgus CD20 (Macaca fascicularis; UniProtKB No G7PQ03).

[0057] As used herein, the term "CD20 antibody" or "anti-CD20 antibody" refers to an antibody that specifically binds to the antigen CD20, particularly human CD20.

[0058] As used herein, the term "CD3xCD20 antibody", "anti-CD3xCD20 antibody", "CD20xCD3 antibody" or "anti-CD20xCD3 antibody" refers to a bispecific antibody that comprises two different antigen-binding regions, one of which specifically binds to the antigen CD20 and one of which specifically binds to CD3.

[0059] As used herein, the term "DuoBody-CD3xCD20" refers to an IgG1 bispecific CD3xCD20 antibody that comprises a first heavy and light chain pair as defined in SEQ ID NO: 24 and SEQ ID NO: 25, respectively, and a second heavy and light chain pair as defined in SEQ ID NO: 26 and SEQ ID NO: 27, respectively. The first heavy and light chain pair comprises a region that binds to human CD3ε (epsilon), and the second heavy and light chain pair comprises a region that binds to human CD20. The first binding region comprises VH and VL sequences as defined in SEQ ID NO: 6 and 7, and the second binding region comprises VH and VL sequences as defined in SEQ ID NO: 13 and 14. Such bispecific antibodies can be prepared as described in WO 2016 / 110576.

[0060] The present invention also provides antibodies that comprise functional variants of the heavy chain, light chain, VL region, VH region, or one or more CDRs of the antibodies of the examples. Functional variants of heavy chain, light chain, VL, VH, or CDR used in the context of an antibody still allow the antibody to retain at least a substantial proportion (at least about 90%, 95% or more) of the functional characteristics of the "reference" and / or "parent" antibody, including affinity and / or specificity / selectivity for specific epitopes of CD20 and / or CD3, Fc inertness, and PK parameters such as half-life, Tmax, Cmax. Such functional variants generally retain significant sequence identity with the parent antibody and / or have heavy and light chains of substantially similar length. The percentage identity between two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps that need to be introduced to achieve an optimal alignment of the two sequences and the length of each gap (i.e., % homology = # of identical positions / total # of positions x 100). The percentage identity between two nucleotide or amino acid sequences can be determined, for example, using the algorithm of E. Meyers and W. Miller, Comput. Appl. Biosci 4, 11-17 (1988), incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. Additionally, the percentage identity between two amino acid sequences can be determined using the algorithm of Needleman and Wunsch, J. Mol. Biol. 48, 444-453 (1970). Exemplary variants include variants that differ from the heavy and / or light chain, VH and / or VL, and / or CDR regions of the parent antibody sequence primarily by conservative substitutions; for example, 10, such as 9, 8, 7, 6, 5, 4, 3, 2, or 1 substitution(s) in the variant can be conservative amino acid residue substitutions.

[0061] Conservative substitutions can be defined by substitutions within the amino acid classes reflected in the following table:

[0062] Table 1: Amino Acid Residue Classes for Conservative Substitutions

[0063]

[0064] Unless otherwise indicated, the following nomenclature is used to describe mutations: i) an amino acid substitution at a given position is written, for example, as K409R, which means that the lysine at position 409 is replaced with arginine; and ii) for a particular variant, a specific three-letter or one-letter code, including the codes Xaa and X, is used to indicate any amino acid residue. Thus, the replacement of lysine with arginine at position 409 is designated: K409R, and the replacement of lysine with any amino acid residue at position 409 is designated K409X. If the lysine at position 409 is deleted, it is designated K409*.

[0065] As used herein, the term "humanized antibody" refers to a genetically engineered non-human antibody that contains a human antibody constant domain and a non-human variable domain that has been modified to contain a high degree of sequence homology with a human variable domain. This can be achieved by transplanting the six non-human antibody CDRs that together form the antigen-binding site onto a homologous human acceptor framework region (FR) (see WO92 / 22653 and EP0629240). To fully reconstruct the binding affinity and specificity of the parental antibody, it may be necessary to replace framework residues of the parental antibody (i.e., the non-human antibody) with human framework regions (backmutations). Structural homology modeling may assist in identifying amino acid residues in the framework region that are important for antibody binding properties. Thus, a humanized antibody can contain non-human CDR sequences, primarily a human framework region that optionally contains one or more amino acid backmutations relative to the non-human amino acid sequence, and a fully human constant domain. The VH and VL of the CD3 arm used herein in DuoBody-CD3xCD20 represent humanized antigen-binding regions. Optionally, additional amino acid modifications that are not necessarily backmutations can be applied to obtain a humanized antibody with preferred characteristics such as affinity and biochemical properties.

[0066] As used herein, the term "human antibody" refers to an antibody having variable and constant regions derived from human germline immunoglobulin sequences. A human antibody may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by in vitro random or site-specific mutagenesis or by in vivo somatic mutation). However, as used herein, the term "human antibody" is not intended to include antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences. The VH and VL of the CD20 arm used in DuoBody-CD3xCD20 represent the human antigen-binding regions. The human monoclonal antibodies of the present invention can be produced by a variety of techniques, including conventional monoclonal antibody methods such as the standard somatic hybridization technique of Kohler and Milstein, Nature 256: 495 (1975). While somatic hybridization procedures are preferred, in principle, other techniques for producing monoclonal antibodies can be employed, such as viral or oncogenic transformation of B lymphocytes or phage display techniques using human antibody gene libraries. A suitable animal system for preparing hybridomas that secrete human monoclonal antibodies is the murine system. The production of hybridomas in mice is a well-established procedure. Immunization protocols and techniques for isolating immunized spleen cells for fusion are known in the art. The fusion partners (e.g., murine myeloma cells) and fusion procedures are also known. Thus, transgenic or transchromosomal mice or rats carrying a portion of the human immune system rather than a murine or rat system can be used to generate human monoclonal antibodies. Thus, in one embodiment, the human antibodies are obtained from transgenic animals, such as mice or rats, that carry human germline immunoglobulin sequences rather than animal immunoglobulin sequences. In such embodiments, the antibodies are derived from the human germline immunoglobulin sequences introduced into the animal, but the final antibody sequences are the result of further modification of the human germline immunoglobulin sequences by somatic hypermutation and affinity maturation by the endogenous animal antibody mechanisms (see, e.g., Mendez et al. Nat Genet 1997;15:146-56). The VH region and VL region of the CD20 arm used in DuoBody-CD3xCD20 represent the human antigen-binding regions.

[0067] As used herein, the term “biosimilar” (e.g., of an approved reference product / biologic) means a biological product that is similar to a reference product based on data from: (a) analytical studies that show the biological product is highly similar to the reference product, notwithstanding minor differences in clinically inactive components; (b) animal studies (including toxicity assessments); and / or (c) one or more clinical studies (including assessments of immunogenicity and pharmacokinetics or pharmacodynamics) sufficient to demonstrate safety, purity, and potency under one or more appropriate conditions of use for which the reference product is approved and is to be used and for which approval is sought (e.g., there are no clinically meaningful differences between the biological product and the reference product with respect to the safety, purity, and potency of the product). In some embodiments, the biosimilar biological product and the reference product use the same one or more mechanisms of action for one or more conditions of use specified, recommended, or suggested in the proposed label, but only to the extent of one or more mechanisms of action known for the reference product. In some embodiments, one or more conditions of use specified, recommended, or suggested in the proposed label for the biological product have been previously approved for the reference product. In some embodiments, the route of administration, dosage form, and / or strength of the biological product is the same as that of the reference product. A biosimilar can be, for example, an antibody currently known to have the same primary amino acid sequence as a marketed antibody, but prepared in a different cell type or by a different method of production, purification, or formulation.

[0068] As used herein, the term “reducing conditions” or “reducing environment” means conditions or an environment in which a substrate (here, a cysteine residue in the antibody hinge region) is more likely to be reduced rather than oxidized.

[0069] As used herein, the term “recombinant host cell” (or simply “host cell”) is intended to mean a cell into which an expression vector, such as an expression vector encoding an antibody described herein, has been introduced. Recombinant host cells include, for example, transfected cell lines such as CHO, CHO-S, HEK, HEK293, HEK-293F, Expi293F, PER.C6, or NSO cells, as well as lymphocytes.

[0070] As used herein, "Richter's syndrome" or "Richter's transformation" are used interchangeably to refer to the transformation of chronic lymphocytic leukemia (CLL) into an aggressive lymphoma. Richter's syndrome occurs in the context of CLL or SLL (Swerdlow et al., 2017; WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. International Agency for Research on Cancer, Lyon, France) and occurs in approximately 10% to 15% of patients with CLL. In most cases, CLL evolves into diffuse large B-cell lymphoma (DLBCL), which maintains a clonal relationship with the original leukemia phase, while the remaining patients develop a Hodgkin lymphoma variant. The survival of patients with RS is generally poor, and subjects carrying selective chromosomal aberrations or those clonally related to CLL experience the worst prognosis and outcomes (Allan and Furman, 2019; Int J Hematol Oncol, 7 (4), p. IJH09, Falchi, et al., 2014; Blood, 123 (18), pp. 2783-27903).

[0071] Several genetic and immune factors may contribute to transformation. In recent years, additional risk factors have been identified, such as TP53 disruption, NOTCH1 mutation, CDKN2A loss, and MYC activation ((Rossi, et al., 2018; Blood, 131 (25), pp. 2761-2772, Chigrinova, et al., 2013; Blood, 122 (15), pp. 2673-2682, Fabbri, et al., 2013; J Exp Med, 210 (11) 13), pp. 2273-2288, Parikh et al., 2014; Blood, 123 (11), pp. 1647-1657). Additionally, the biased use of subgroup 8 V4-39 stereotyped immunoglobulin genes increases the risk of RS development 24-fold (Parikh et al., 2013; Br J Haematol, 162(6), pp. 774-782, Rossi et al., 2009, Clin Cancer Res, 15(13), pp. 4415-4422), indicating a driving role of B cell receptor (BCR) signaling in transformation. Overall, the molecular profile of RS is heterogeneous, lacks a unifying lesion, and does not overlap genetically with de novo DLBCL (Fabbri, et al., 2013; J Exp Med, 210 (11) 13), pp. 2273-2288). Dysregulation of underlying transcriptional programs and signaling pathways may account for the aggressive clinical phenotype of RS (Allan and Furman, 2019; Int J Hematol Oncol, 7 (4), p. IJH09).

[0072] The term "treatment" refers to the administration of an effective amount of a therapeutically active antibody described herein for the purpose of alleviating, ameliorating, arresting, or eradicating (curing) a symptom or disease state such as CLL. Treatment can result in a complete response (CR), partial response (PR), or stable disease (SD), e.g., as defined by the Lugano criteria (Cheson et al., 2014), as shown in Table 2.

[0073] Treatment can be continued, e.g., until disease progression (PD) or unacceptable toxicity.

[0074] As used herein, the term "administering" refers to the physical introduction of a composition (or formulation) comprising a therapeutic agent into a subject using any of a variety of methods and delivery systems known to those of skill in the art. Preferred routes of administration of the antibodies described herein include intravenous, intraperitoneal, intramuscular, subcutaneous, spinal or other parenteral routes of administration, such as by injection or infusion. As used herein, the phrase "parenteral administration" refers to a mode of administration other than enteral and topical administration, usually by injection, and includes, but is not limited to, intravenous, intraperitoneal, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intratracheal, subcutaneous, subepidermal, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion, as well as in vivo electroporation. Alternatively, the therapeutic agents described herein may be administered via non-parenteral routes, such as topical, epidermal or mucosal routes of administration, such as intranasal, oral, vaginal, rectal, sublingual or topical administration. Administration may also be effected, for example, once, multiple times and / or over one or more extended periods. In the methods described herein, a bispecific antibody (e.g., acorecrtamab) is administered subcutaneously. Other agents used in combination with the bispecific antibody (such as for cytokine release syndrome prophylaxis or tumor lysis syndrome (TLS) prophylaxis) may be administered via other routes, such as intravenously or orally.

[0075] The term "effective amount" or "therapeutically effective amount" refers to an amount that is effective to achieve the desired therapeutic result at the required dosage and for the required period of time. For example, a dosage of a bispecific antibody (e.g., acorecrtamab) for subcutaneous administration in the range of 12 - 60 mg, as defined herein, may be defined as such an "effective amount" or "therapeutically effective amount". The therapeutically effective amount of an antibody can vary depending on factors such as the disease state, age, sex and weight of the individual, as well as the ability of the antibody to elicit a desired response in the individual. A therapeutically effective amount is also an amount in which any toxic or detrimental effects of the antibody or antibody moiety are offset by the therapeutic beneficial effects. In some embodiments, a patient treated by the methods described herein will exhibit an improvement in ECOG performance status. The therapeutically effective amount or dose of a drug includes a "prophylactically effective amount" or "prophylactically effective dose", which is any amount of the drug that inhibits the development or recurrence of a disease when the drug is administered alone or in combination with another therapeutic agent to a subject at risk of developing a disease or condition (e.g., cytokine release syndrome) or at risk of having a recurrence of a disease.

[0076] As used herein, the term "inhibiting tumor growth" includes any measurable reduction in tumor growth, such as inhibiting tumor growth by at least about 10%, such as at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 99% or 100%.

[0077] As used herein, the term "subject" refers to a human patient, e.g., a human patient suffering from Richter's syndrome. The terms "subject" and "patient" are used interchangeably herein.

[0078] As used herein, the term "buffer" refers to a pharmaceutically acceptable buffer. The term "buffer" encompasses those reagents that maintain the pH of a solution within an acceptable range, for example, and includes, but is not limited to, acetate, histidine, TRIS® (tris(hydroxymethyl)aminomethane), citrate, succinate, glycolate, and the like. Generally, as used herein, a "buffer" has a pKa and buffering capacity suitable for a pH range of about 5 to about 6, preferably about 5.5.

[0079] As used herein, "disease progression" or "PD" refers to a situation in which one or more indices of lymphoma indicate that the disease is progressing despite treatment. In some embodiments, disease progression is defined according to the Lugano criteria (Cheson et al., 2014), as shown in Table 2.

[0080] As used herein, a "surfactant" is a compound commonly used in pharmaceutical formulations to prevent the adsorption of a drug to a surface and / or aggregation. In addition, a surfactant reduces the surface tension (or interfacial tension) between two liquids or between a liquid and a solid. For example, when present at very low concentrations (e.g., 5% w / v or less, such as 3% w / v or less, such as 1% w / v or less, such as 0.4% w / v or less, such as less than 0.1% w / v or less, such as 0.04% w / v), exemplary surfactants can significantly reduce the surface tension. Surfactants are amphiphilic, meaning they typically consist of hydrophilic and hydrophobic or lipophilic groups and are thus able to form micelles or similar self-assembled structures in aqueous solutions. Known pharmaceutical surfactants include glyceryl monooleate, benzethonium chloride, sodium docusate, phospholipids, polyethylene alkylethers, sodium lauryl sulfate, and trioctanoin (anionic surfactants); benzalkonium chloride, citrimonium bromide, cetylpyridinium chloride, and phospholipids (cationic surfactants); and α-tocopherol, glyceryl monooleate, myristyl alcohol, phospholipids, poloxamer, polyoxyethylene alkyl ethers, polyoxyethylene castor oil derivatives, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene stearate, polyoxylhydroxystearate, polyoxylglyceride, polysorbates such as polysorbate 20 or polysorbate 80, propylene glycol dilaurate, propylene glycol monolaurate, sorbitan esters sucrose palmitate, sucrose stearate, trioctanoin, and TPGS (nonionic and zwitterionic surfactants).

[0081] As used herein, a "diluent" is a pharmaceutically acceptable (safe and non-toxic for administration to humans) diluent that can be used to prepare a pharmaceutical composition or formulation (the terms "composition" and "formulation" are used interchangeably herein). Preferably, such a diluent of the composition only dilutes the antibody concentration and not the buffer and stabilizer. Thus, in one embodiment, the diluent contains the same buffer and stabilizer as present in the pharmaceutical composition of the invention. Further exemplary diluents include sterile water, bacteriostatic water for injection (BWFI), a pH buffer solution (preferably an acetate buffer), a sterile saline solution, Ringer's solution, or a dextrose solution. In one embodiment, the diluent comprises an acetate buffer and sorbitol or consists essentially of an acetate buffer and sorbitol.

[0082] As used herein, the term "about" means ±10% of a specified value.

[0083] Richter's syndrome treatment regimen

[0084] Richter's syndrome (RS), also known as Richter's transformation, is a rare complication of chronic lymphocytic leukemia (CLL) and / or small lymphocytic lymphoma (SLL). It is characterized by the sudden transformation of CLL / SLL into a significantly more aggressive large cell lymphoma. In most cases, CLL, which usually grows slowly or indolently, transforms into a common type of non-Hodgkin lymphoma (NHL) known as diffuse large B cell lymphoma (DLBCL). Cases of transformation into Hodgkin lymphoma (HL) / Hodgkin disease (HD) and some types of T cell lymphoma have been reported more rarely.

[0085] Although the exact cause of Richter's syndrome remains unclear, certain factors are thought to increase the risk of developing RS in patients already diagnosed with CLL / SLL. These risk factors include certain inherited genetic characteristics (e.g., BCL-2, CD38, LRP4 genotypes) and specific genetic mutations. For example, patients carrying deletions of chromosomes 11q and 17p, an unmutated IGVH gene, a NOTCH-1 mutation, shortened telomere length, elevated levels of ζ-associated protein (ZAP-70), β2-microglobulin (B2M), and CD38, and / or having advanced disease (Rai stage III-IV with >3 cm lymph nodes) at the time of initial CLL diagnosis are all considered to be at greater risk of developing RS.

[0086] Richter's syndrome is characterized by sudden clinical deterioration. Currently available therapies have shown limited responses with unsatisfactory safety profiles. The median overall survival ranges from a few months to approximately 1 year. Since there is no established standard of care, there is a need to provide therapies with novel modes of action that provide durable responses with tolerable safety profiles. One such therapy is the treatment of RS patients with a bispecific antibody that binds to CD3 and CD20 ("anti-CD3xCD20 antibody").

[0087] Accordingly, in one aspect, the present disclosure provides a method of treating Richter's syndrome in a human subject, the method comprising administering (e.g., subcutaneously) to the subject an effective amount of a bispecific antibody comprising:

[0088] (i) a first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a variable heavy chain (VH) region and a variable light chain (VL) region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences of the VH region sequence of SEQ ID NO: 6, and the VL region comprises the CDR1, CDR2, and CDR3 sequences of the VL region sequence of SEQ ID NO: 7; and

[0089] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region and a VL region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences in the VH region sequence of SEQ ID NO: 13, and the VL region comprises the CDR1, CDR2, and CDR3 sequences in the VL region sequence of SEQ ID NO: 14.

[0090] Preferably, the bispecific antibody is administered at a dose in the range of 12 - 60 mg over a 28-day period.

[0091] In some embodiments, the bispecific antibody is a full-length antibody. In some embodiments, the bispecific antibody is an antibody with an inert Fc region. In some embodiments, the bispecific antibody is a full-length antibody with an inert Fc region.

[0092] In some embodiments, the bispecific antibody is administered at a dose of 12 mg (or a dose of about 12 mg). In some embodiments, the bispecific antibody is administered at a dose of 24 mg (or a dose of about 24 mg). In some embodiments, the bispecific antibody is administered at a dose of 48 mg (or a dose of about 48 mg). In some embodiments, the bispecific antibody is administered at a dose of 60 mg (or a dose of about 60 mg).

[0093] Regarding the 12 - 60 mg dose of the bispecific antibody to be administered, or any other specified dose, it should be understood that this amount refers to the amount of the bispecific antibody representing the full-length antibody, such as acoramutab as defined in the Examples section. Thus, administering a dose of 24 mg of the bispecific antibody can be referred to as administering the dose of the bispecific antibody described herein, where this dose corresponds to a 24 mg dose of acoramutab. When, for example, the molecular weight of the antibody used is significantly different from the molecular weight of a full-length antibody such as acoramutab, a person of ordinary skill in the art can readily determine the amount of antibody to be administered. For example, the amount of the antibody can be calculated by dividing the molecular weight of the antibody by the weight of a full-length antibody such as acoramutab and multiplying the result by the specified dose as described herein. As long as the bispecific antibody (e.g., a functional variant of DuoBody-CD3xCD20) has highly similar characteristics to DuoBody-CD3xCD20 in terms of plasma half-life, Fc inertness, and / or binding characteristics to CD3 and CD20 (i.e., in terms of CDR and epitope binding characteristics), such an antibody is suitable for use in the methods provided herein at the doses described for full-length antibodies such as acoramutab.

[0094] In one embodiment, the bispecific anti-CD3xCD20 antibody is administered at a dose in the range between 12 mg and 60 mg. In some embodiments, the bispecific antibody is administered at a dose of 12 mg or about 12 mg. In some embodiments, the bispecific antibody is administered at a dose of 24 mg or about 24 mg. In some embodiments, the bispecific antibody is administered at a dose of 48 mg or about 48 mg. In some embodiments, the bispecific antibody is administered at a dose of 60 mg or about 60 mg.

[0095] In some embodiments, the dose of the bispecific antibody is administered once a week (weekly administration) in a 28-day cycle. In some embodiments, the weekly administration is carried out for 2.5 28-day cycles (i.e., 10 times). In one embodiment, the dose is administered for 2.5 28-day cycles (i.e., 10 times; on days 15 and 22 of cycle 1, and on days 1, 8, 15, and 22 of cycles 2 and 3). In some embodiments, after the weekly administration, the interval between administrations of the bispecific antibody can be reduced to once every two weeks (bi-weekly administration). In some embodiments, such bi-weekly administration can be carried out for 6 28-day cycles (i.e., 12 times). In some embodiments, after the bi-weekly administration, the interval between administrations of the bispecific antibody can be further reduced to once every four weeks. In one embodiment, the once-every-four-week administration can be carried out for an extended period of time, e.g., for at least 1 cycle, at least 2 cycles, at least 3 cycles, at least 4 cycles, at least 5 cycles, at least 6 cycles, at least 7 cycles, at least 8 cycles, at least 9 cycles, at least 10 cycles, at least 11 cycles, at least 12 cycles, at least 13 cycles, at least 14 cycles, at least 15 cycles, at least 16 cycles, at least 17 cycles, or between 1 - 20 cycles, between 1 - 19 cycles, between 1 - 18 cycles, between 1 - 17 cycles, between 1 - 16 cycles, between 1 - 15 cycles, between 1 - 14 cycles, between 1 - 13 cycles, between 1 - 12 cycles, between 1 - 10 cycles, between 1 - 5 cycles, between 5 - 20 cycles, between 5 - 15 cycles, or between 5 - 10 cycles of a 28-day cycle. In some embodiments, acalabrutinib is administered once every four weeks until disease progression (e.g., as defined by the Lugano criteria (Cheson et al., 2014), as shown in Table 2) or unacceptable toxicity. In one embodiment, the once-weekly dose is administered in cycles 1 - 3 (and may include a sensitizing dose and an intermediate dose, as described below), the once-every-two-week dose is administered in cycles 4 - 9, and the once-every-four-week dose is administered from cycle 10 onwards.

[0096] It should be understood that the doses referred to herein may also be referred to as full doses or flat doses in the above circumstances, where, for example, once-weekly doses, once-every-two-week doses, and / or once-every-four-week doses are administered at the same level. Thus, when preferably selecting a dose of 48 mg for each once-weekly administration, each once-every-two-week administration, and each once-every-four-week administration, the same dose of 48 mg should be administered. Before administering this dose, a sensitizing dose or a sensitizing dose and one or more subsequent intermediate (second sensitizing) doses may be administered. This may be advantageous as it may help reduce the risk and severity of cytokine release syndrome (CRS), a side effect that can occur during treatment with the bispecific anti-CD3xCD20 antibody described herein. Such sensitizing doses or sensitizing and intermediate doses are at lower doses compared to the fixed or full doses.

[0097] Thus, in some embodiments, a sensitizing dose of the bispecific antibody may be administered before administering a once-weekly dose of 12 - 60 mg. In one embodiment, the sensitizing dose is administered two weeks before administering the first once-weekly dose of 12 - 60 mg in cycle 1. The sensitizing dose may be in the range of 20 - 2000 µg (0.02 mg - 2 mg), such as in the range of 50 - 1000 µg (0.05 mg to 1 mg) or 70 - 350 µg (0.07 mg to 0.35 mg). The sensitizing dose may be, for example, 80, 100, 120, 140, 160, 180, 200, 220, 240, 260, 280, 300, 320, 350, 400, 450, 500, 600, 700, 800, 900, or 1000 µg, or about 80, about 100, about 120, about 140, about 160, about 180, about 200, about 220, about 240, about 260, about 280, about 300, about 320, about 350, about 400, about 450, about 500, about 600, about 700, about 800, about 900, or about 1000 µg. In a preferred embodiment, the sensitizing dose is between 50 and 350 µg (0.05 and 0.35 mg, respectively). In a more preferred embodiment, the sensitizing dose is 160 µg (0.16 mg) or about 160 µg (about 0.16 mg). In the most preferred embodiment, the sensitizing dose is 160 µg (0.16 mg) or about 160 µg (about 0.16 mg) of the full-length bispecific antibody.

[0098] In some embodiments, one or more intermediate doses of the bispecific antibody are administered after a sensitizing dose and before a first once-weekly dose of 12 - 60 mg. In one embodiment, the sensitizing dose is administered on day 1 of cycle 1 and the intermediate dose is administered on day 8, followed by the first once-weekly dose of 12 - 60 mg on days 15 and 22, i.e., the sensitizing dose is administered one week before the intermediate dose (i.e., day 1 of cycle 1), and the intermediate dose is administered one week before the first once-weekly dose of 12 - 60 mg (day 8 of cycle 1). The one or more intermediate doses are selected from the range between the sensitizing dose and the fixed or full dose. For example, the one or more intermediate doses can be in the range of 200 - 8000 μg (0.2 - 8 mg), e.g., in the range of 400 - 4000 μg (0.4 - 4 mg) or 600 - 2000 μg (0.6 - 2 mg). The intermediate dose can be, for example, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500 or 1600 μg, or about 200, about 300, about 400, about 500, about 600, about 700, about 800, about 900, about 1000, about 1100, about 1200, about 1300, about 1400, about 1500 or about 1600 μg. In a preferred embodiment, the intermediate dose is in the range of 600 and 1200 μg (0.6 and 1.2 mg, respectively). The presently preferred embodiment uses an intermediate dose which is 800 μg (0.8 mg) or about 800 μg (0.8 mg). The most preferred embodiment uses an intermediate dose which is 800 μg or about 800 μg (0.8 mg) of the full-length bispecific antibody.

[0099] In some embodiments, the bispecific antibody is administered (e.g., subcutaneously) in 28-day cycles, wherein

[0100] a) in cycle 1, a sensitizing dose is administered on day 1, an intermediate dose is administered on day 8, and a full dose of 12 - 60 mg is administered on days 15 and 22;

[0101] b) in cycles 2 - 3, a full dose of 12 - 60 mg is administered on days 1, 8, 15 and 22;

[0102] c) in cycles 4 - 9, a full dose of 12 - 60 mg is administered on days 1 and 15; and

[0103] d) in cycle 10 and subsequent cycles, a full dose of 12 - 60 mg is administered on day 1.

[0104] In some embodiments, the bispecific antibody is administered in a 28-day cycle (e.g., subcutaneously), wherein

[0105] a) in cycle 1, a sensitizing dose in the range of 0.05 - 0.35 is administered on day 1, an intermediate dose in the range of 0.6 - 1.2 mg is administered on day 8, and a full dose of 12 - 60 mg is administered on days 15 and 22;

[0106] b) in cycles 2 - 3, a full dose of 12 - 60 mg is administered on days 1, 8, 15, and 22;

[0107] c) in cycles 4 - 9, a full dose of 12 - 60 mg is administered on days 1 and 15; and

[0108] d) in cycle 10 and subsequent cycles, a full dose of 12 - 60 mg is administered on day 1.

[0109] In some embodiments, the bispecific antibody is administered in a 28-day cycle (e.g., subcutaneously), wherein

[0110] a) in cycle 1, a sensitizing dose of 160 μg is administered on day 1, an intermediate dose of 800 μg is administered on day 8, and a full dose of 12 - 60 mg is administered on days 15 and 22;

[0111] b) in cycles 2 - 3, a full dose of 12 - 60 mg is administered on days 1, 8, 15, and 22;

[0112] c) in cycles 4 - 9, a full dose of 12 - 60 mg is administered on days 1 and 15; and

[0113] d) in cycle 10 and subsequent cycles, a full dose of 12 - 60 mg is administered on day 1.

[0114] In some embodiments, the bispecific antibody is administered in a 28-day cycle (e.g., subcutaneously), wherein

[0115] a) in cycle 1, a sensitizing dose is administered on day 1, an intermediate dose is administered on day 8, and a full dose of 12 mg or about 12 mg is administered on days 15 and 22;

[0116] b) in cycles 2 - 3, a full dose of 12 mg or about 12 mg is administered on days 1, 8, 15, and 22;

[0117] c) in cycles 4 - 9, a full dose of 12 mg or about 12 mg is administered on days 1 and 15; and

[0118] d) In cycle 10 and subsequent cycles, administer a full dose of 12 mg or approximately 12 mg on day 1.

[0119] In some embodiments, the bispecific antibody is administered in 28-day cycles (e.g., subcutaneously), wherein

[0120] a) In cycle 1, administer a sensitizing dose in the range of 0.05 - 0.35 on day 1, an intermediate dose in the range of 0.6 - 1.2 mg on day 8, and a full dose of 12 mg on days 15 and 22;

[0121] b) In cycles 2 - 3, administer a full dose of 12 mg on days 1, 8, 15, and 22;

[0122] c) In cycles 4 - 9, administer a full dose of 12 mg on days 1 and 15; and

[0123] d) In cycle 10 and subsequent cycles, administer a full dose of 12 mg on day 1.

[0124] In some embodiments, the bispecific antibody is administered in 28-day cycles (e.g., subcutaneously), wherein

[0125] a) In cycle 1, administer a sensitizing dose of 160 μg on day 1, an intermediate dose of 800 μg on day 8, and a full dose of 12 mg or approximately 12 mg on days 15 and 22;

[0126] b) In cycles 2 - 3, administer a full dose of 12 mg or approximately 12 mg on days 1, 8, 15, and 22;

[0127] c) In cycles 4 - 9, administer a full dose of 12 mg or approximately 12 mg on days 1 and 15; and

[0128] d) In cycle 10 and subsequent cycles, administer a full dose of 12 mg or approximately 12 mg on day 1.

[0129] In some embodiments, the bispecific antibody is administered in 28-day cycles (e.g., subcutaneously), wherein

[0130] a) In cycle 1, administer a sensitizing dose on day 1, an intermediate dose on day 8, and a full dose of 24 mg or approximately 24 mg on days 15 and 22;

[0131] b) In cycles 2 - 3, administer a full dose of 24 mg or approximately 24 mg on days 1, 8, 15, and 22;

[0132] c) In cycles 4 - 9, administer a full dose of 24 mg or approximately 24 mg on days 1 and 15; and

[0133] d) In cycle 10 and subsequent cycles, administer a full dose of 24 mg or approximately 24 mg on day 1.

[0134] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0135] a) In cycle 1, administer a sensitizing dose within the range of 0.05 and 0.35 on day 1, an intermediate dose within the range of 0.6 and 1.2 mg on day 8, and a full dose of 24 mg on days 15 and 22;

[0136] b) In cycles 2 - 3, administer a full dose of 24 mg on days 1, 8, 15, and 22;

[0137] c) In cycles 4 - 9, administer a full dose of 24 mg on days 1 and 15; and

[0138] d) In cycle 10 and subsequent cycles, administer a full dose of 24 mg on day 1.

[0139] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0140] a) In cycle 1, administer a sensitizing dose of 160 μg on day 1, an intermediate dose of 800 μg on day 8, and a full dose of 24 mg or approximately 24 mg on days 15 and 22;

[0141] b) In cycles 2 - 3, administer a full dose of 24 mg or approximately 24 mg on days 1, 8, 15, and 22;

[0142] c) In cycles 4 - 9, administer a full dose of 24 mg or approximately 24 mg on days 1 and 15; and

[0143] d) In cycle 10 and subsequent cycles, administer a full dose of 24 mg or approximately 24 mg on day 1.

[0144] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0145] a) In cycle 1, administer a sensitizing dose on day 1, an intermediate dose on day 8, and a full dose of 48 mg or approximately 48 mg on days 15 and 22;

[0146] b) In cycle 2 - 3, administer a full dose of 48 mg or approximately 48 mg on days 1, 8, 15, and 22;

[0147] c) In cycles 4 - 9, administer a full dose of 48 mg or approximately 48 mg on days 1 and 15; and

[0148] d) In cycle 10 and subsequent cycles, administer a full dose of 48 mg or approximately 48 mg on day 1.

[0149] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0150] a) In cycle 1, administer a sensitizing dose within the range of 0.05 - 0.35 mg on day 1, an intermediate dose within the range of 0.6 - 1.2 mg on day 8, and a full dose of 48 mg on days 15 and 22;

[0151] b) In cycles 2 - 3, administer a full dose of 48 mg on days 1, 8, 15, and 22;

[0152] c) In cycles 4 - 9, administer a full dose of 48 mg on days 1 and 15; and

[0153] d) In cycle 10 and subsequent cycles, administer a full dose of 48 mg on day 1.

[0154] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0155] a) In cycle 1, administer a sensitizing dose of 160 μg on day 1, an intermediate dose of 800 μg on day 8, and a full dose of 48 mg or approximately 48 mg on days 15 and 22;

[0156] b) In cycles 2 - 3, administer a full dose of 48 mg or approximately 48 mg on days 1, 8, 15, and 22;

[0157] c) In cycles 4 - 9, administer a full dose of 48 mg or approximately 48 mg on days 1 and 15; and

[0158] d) In cycle 10 and subsequent cycles, administer a full dose of 48 mg or approximately 48 mg on day 1.

[0159] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0160] a) In Cycle 1, a sensitizing dose is administered on Day 1, an intermediate dose on Day 8, and a full dose of 60 mg or approximately 60 mg on Days 15 and 22;

[0161] b) In Cycles 2 - 3, a full dose of 60 mg or approximately 60 mg is administered on Days 1, 8, 15, and 22;

[0162] c) In Cycles 4 - 9, a full dose of 60 mg or approximately 60 mg is administered on Days 1 and 15; and

[0163] d) In Cycle 10 and subsequent cycles, a full dose of 60 mg or approximately 60 mg is administered on Day 1.

[0164] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0165] a) In Cycle 1, a sensitizing dose in the range of 0.05 - 0.35 is administered on Day 1, an intermediate dose in the range of 0.6 - 1.2 mg on Day 8, and a full dose of 60 mg on Days 15 and 22;

[0166] b) In Cycles 2 - 3, a full dose of 60 mg is administered on Days 1, 8, 15, and 22;

[0167] c) In Cycles 4 - 9, a full dose of 60 mg is administered on Days 1 and 15; and

[0168] d) In Cycle 10 and subsequent cycles, a full dose of 60 mg is administered on Day 1.

[0169] In some embodiments, the bispecific antibody is administered in 28 - day cycles (e.g., subcutaneously), wherein

[0170] a) In Cycle 1, a sensitizing dose of 160 μg is administered on Day 1, an intermediate dose of 800 μg on Day 8, and a full dose of 60 mg or approximately 60 mg on Days 15 and 22;

[0171] b) In Cycles 2 - 3, a full dose of 60 mg or approximately 60 mg is administered on Days 1, 8, 15, and 22;

[0172] c) In Cycles 4 - 9, a full dose of 60 mg or approximately 60 mg is administered on Days 1 and 15; and

[0173] d) In Cycle 10 and subsequent cycles, a full dose of 60 mg or approximately 60 mg is administered on Day 1.

[0174] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28-day cycles, wherein

[0175] a) in Cycle 1, a sensitizing dose is administered on Day 1, an intermediate dose is administered on Day 8, and full doses of 12 - 60 mg are administered on Days 15 and 22;

[0176] b) in Cycles 2 - 3, full doses of 12 - 60 mg are administered on Days 1, 8, 15, and 22;

[0177] c) in Cycles 4 - 9, full doses of 12 - 60 mg are administered on Days 1 and 15; and

[0178] d) in Cycle 10 and subsequent cycles, full doses of 12 - 60 mg are administered on Day 1.

[0179] In some embodiments, the bispecific antibody is acrivastamab, which is administered (e.g., subcutaneously) in 28-day cycles, wherein

[0180] a) in Cycle 1, a sensitizing dose in the range of 0.05 - 0.35 is administered on Day 1, an intermediate dose in the range of 0.6 - 1.2 mg is administered on Day 8, and full doses of 12 - 60 mg are administered on Days 15 and 22;

[0181] b) in Cycles 2 - 3, full doses of 12 - 60 mg are administered on Days 1, 8, 15, and 22;

[0182] c) in Cycles 4 - 9, full doses of 12 - 60 mg are administered on Days 1 and 15; and

[0183] d) in Cycle 10 and subsequent cycles, full doses of 12 - 60 mg are administered on Day 1.

[0184] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28-day cycles, wherein

[0185] a) in Cycle 1, a sensitizing dose of 160 μg is administered on Day 1, an intermediate dose of 800 μg is administered on Day 8, and full doses of 12 - 60 mg are administered on Days 15 and 22;

[0186] b) in Cycles 2 - 3, full doses of 12 - 60 mg are administered on Days 1, 8, 15, and 22;

[0187] c) in Cycles 4 - 9, full doses of 12 - 60 mg are administered on Days 1 and 15; and

[0188] d) in Cycle 10 and subsequent cycles, full doses of 12 - 60 mg are administered on Day 1.

[0189] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28-day cycles, wherein

[0190] a) In cycle 1, a sensitizing dose is administered on day 1, an intermediate dose on day 8, and a full dose of 12 mg or about 12 mg on days 15 and 22;

[0191] b) In cycles 2-3, a full dose of 12 mg or about 12 mg is administered on days 1, 8, 15, and 22;

[0192] c) In cycles 4-9, a full dose of 12 mg or about 12 mg is administered on days 1 and 15; and

[0193] d) In cycle 10 and subsequent cycles, a full dose of 12 mg or about 12 mg is administered on day 1.

[0194] In some embodiments, the bispecific antibody is acrivastamab, which is administered (e.g., subcutaneously) in 28-day cycles, wherein

[0195] a) In cycle 1, a sensitizing dose in the range of 0.05 - 0.35 is administered on day 1, an intermediate dose in the range of 0.6 - 1.2 mg on day 8, and a full dose of 12 mg on days 15 and 22;

[0196] b) In cycles 2-3, a full dose of 12 mg is administered on days 1, 8, 15, and 22;

[0197] c) In cycles 4-9, a full dose of 12 mg is administered on days 1 and 15; and

[0198] d) In cycle 10 and subsequent cycles, a full dose of 12 mg is administered on day 1.

[0199] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28-day cycles, wherein

[0200] a) In cycle 1, a sensitizing dose of 160 μg is administered on day 1, an intermediate dose of 800 μg on day 8, and a full dose of 12 mg or about 12 mg on days 15 and 22;

[0201] b) In cycles 2-3, a full dose of 12 mg or about 12 mg is administered on days 1, 8, 15, and 22;

[0202] c) In cycles 4-9, a full dose of 12 mg or about 12 mg is administered on days 1 and 15; and

[0203] d) In cycle 10 and subsequent cycles, administer a full dose of 12 mg or about 12 mg on day 1.

[0204] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28-day cycles, wherein

[0205] a) In cycle 1, administer a sensitizing dose on day 1, an intermediate dose on day 8, and a full dose of 24 mg or about 24 mg on days 15 and 22;

[0206] b) In cycles 2-3, administer a full dose of 24 mg or about 24 mg on days 1, 8, 15, and 22;

[0207] c) In cycles 4-9, administer a full dose of 24 mg or about 24 mg on days 1 and 15; and

[0208] d) In cycle 10 and subsequent cycles, administer a full dose of 24 mg or about 24 mg on day 1.

[0209] In some embodiments, the bispecific antibody is acrivastamab, which is administered (e.g., subcutaneously) in 28-day cycles, wherein

[0210] a) In cycle 1, administer a sensitizing dose within the range of 0.05 - 0.35 on day 1, an intermediate dose within the range of 0.6 - 1.2 mg on day 8, and a full dose of 24 mg on days 15 and 22;

[0211] b) In cycles 2-3, administer a full dose of 24 mg on days 1, 8, 15, and 22;

[0212] c) In cycles 4-9, administer a full dose of 24 mg on days 1 and 15; and

[0213] d) In cycle 10 and subsequent cycles, administer a full dose of 24 mg on day 1.

[0214] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28-day cycles, wherein

[0215] a) In cycle 1, administer a sensitizing dose of 160 μg on day 1, an intermediate dose of 800 μg on day 8, and a full dose of 24 mg or about 24 mg on days 15 and 22;

[0216] b) In cycles 2-3, administer a full dose of 24 mg or about 24 mg on days 1, 8, 15, and 22;

[0217] c) In cycles 4 - 9, administer a full dose of 24 mg or approximately 24 mg on days 1 and 15; and

[0218] d) In cycle 10 and subsequent cycles, administer a full dose of 24 mg or approximately 24 mg on day 1.

[0219] In some embodiments, the bispecific antibody is acoretozumab, which is administered subcutaneously in a 28 - day cycle, wherein

[0220] a) In cycle 1, administer a sensitizing dose on day 1, an intermediate dose on day 8, and a full dose of 48 mg or approximately 48 mg on days 15 and 22;

[0221] b) In cycles 2 - 3, administer a full dose of 48 mg or approximately 48 mg on days 1, 8, 15, and 22;

[0222] c) In cycles 4 - 9, administer a full dose of 48 mg or approximately 48 mg on days 1 and 15; and

[0223] d) In cycle 10 and subsequent cycles, administer a full dose of 48 mg or approximately 48 mg on day 1.

[0224] In some embodiments, the bispecific antibody is acoretozumab, which is administered (e.g., subcutaneously) in a 28 - day cycle, wherein

[0225] a) In cycle 1, administer a sensitizing dose within the range of 0.05 - 0.35 on day 1, an intermediate dose within the range of 0.6 - 1.2 mg on day 8, and a full dose of 48 mg on days 15 and 22;

[0226] b) In cycles 2 - 3, administer a full dose of 48 mg on days 1, 8, 15, and 22;

[0227] c) In cycles 4 - 9, administer a full dose of 48 mg on days 1 and 15; and

[0228] d) In cycle 10 and subsequent cycles, administer a full dose of 48 mg on day 1.

[0229] In some embodiments, the bispecific antibody is acoretozumab, which is administered subcutaneously in a 28 - day cycle, wherein

[0230] a) In cycle 1, administer a sensitizing dose of 160 μg on day 1, an intermediate dose of 800 μg on day 8, and a full dose of 48 mg or approximately 48 mg on days 15 and 22;

[0231] b) In cycle 2 - 3, administer a full dose of 48 mg or about 48 mg on days 1, 8, 15, and 22;

[0232] c) In cycles 4 - 9, administer a full dose of 48 mg or about 48 mg on days 1 and 15; and

[0233] d) In cycle 10 and subsequent cycles, administer a full dose of 48 mg or about 48 mg on day 1.

[0234] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28 - day cycles, wherein

[0235] a) In cycle 1, administer a sensitizing dose on day 1, an intermediate dose on day 8, and a full dose of 60 mg or about 60 mg on days 15 and 22;

[0236] b) In cycles 2 - 3, administer a full dose of 60 mg or about 60 mg on days 1, 8, 15, and 22;

[0237] c) In cycles 4 - 9, administer a full dose of 60 mg or about 60 mg on days 1 and 15; and

[0238] d) In cycle 10 and subsequent cycles, administer a full dose of 60 mg or about 60 mg on day 1.

[0239] In some embodiments, the bispecific antibody is acrivastamab, which is administered (e.g., subcutaneously) in 28 - day cycles, wherein

[0240] a) In cycle 1, administer a sensitizing dose within the range of 0.05 - 0.35 on day 1, an intermediate dose within the range of 0.6 - 1.2 mg on day 8, and a full dose of 60 mg on days 15 and 22;

[0241] b) In cycles 2 - 3, administer a full dose of 60 mg on days 1, 8, 15, and 22;

[0242] c) In cycles 4 - 9, administer a full dose of 60 mg on days 1 and 15; and

[0243] d) In cycle 10 and subsequent cycles, administer a full dose of 60 mg on day 1.

[0244] In some embodiments, the bispecific antibody is acrivastamab, which is administered subcutaneously in 28 - day cycles, wherein

[0245] a) In cycle 1, a sensitizing dose of 160 μg is administered on day 1, an intermediate dose of 800 μg is administered on day 8, and a full dose of 60 mg or approximately 60 mg is administered on days 15 and 22;

[0246] b) In cycles 2 - 3, a full dose of 60 mg or approximately 60 mg is administered on days 1, 8, 15, and 22;

[0247] c) In cycles 4 - 9, a full dose of 60 mg or approximately 60 mg is administered on days 1 and 15; and

[0248] d) In cycle 10 and subsequent cycles, a full dose of 60 mg or approximately 60 mg is administered on day 1.

[0249] In one embodiment, a sensitizing dose of 80 μg and an intermediate dose of 800 μg are selected on days 1 and 8, respectively, of the first cycle. In some embodiments, a sensitizing dose of 80 μg and an intermediate dose of 1200 μg are selected on days 1 and 8, respectively, of the first cycle. In some embodiments, a sensitizing dose of 80 μg and an intermediate dose of 1600 μg are selected on days 1 and 8, respectively, of the first cycle. In some embodiments, a sensitizing dose of 160 μg and an intermediate dose of 1200 μg are selected on days 1 and 8, respectively, of the first cycle. In some embodiments, a sensitizing dose of 160 μg and an intermediate dose of 1600 μg are selected on days 1 and 8, respectively, of the first cycle.

[0250] In one embodiment, the subject has a clinical history of CLL / SLL that has transformed to aggressive lymphoma; for example, the DLBCL subtype. In a further embodiment, Richter's syndrome has the DLBCL subtype.

[0251] In one embodiment, a human subject has a measurable disease as determined by both: a) a fluorodeoxyglucose (FDG)-positron emission tomography (PET) CT scan that shows a positive lesion compatible with the anatomic tumor site determined by CT (or MRI); and b) a CT scan (or MRI) with ≥2 well-defined lesions / nodules with a long axis >1.5 cm and a short axis >1.0 cm or 1 well-defined lesion / nodule with a long axis >2.0 cm and a short axis ≥1.0 cm of involvement.

[0252] In some embodiments, the human subject has received at least one line of therapy prior to treatment with the methods described herein. For example, in one embodiment, the subject has received one prior line of therapy. In some embodiments, the subject has refractory and / or relapsed Richter's syndrome following a prior anti-neoplastic therapy. Relapse can be defined as evidence of disease progression in a subject who has previously achieved CR or PR for at least 6 months. Refractory disease can be defined as treatment failure (failure to achieve CR or PR) or progression within 6 months of the last dose of therapy. In some embodiments, the subject has received three prior lines of therapy. In some embodiments, the subject has received more than three prior lines of therapy. In some embodiments, the subject has received one, two, three, or more prior lines of therapy. In some embodiments, the subject has received at least two prior lines of therapy. In one embodiment, the prior line of therapy comprises a systemic anti-neoplastic therapy.

[0253] In some embodiments, the subject has received one or more (e.g., at least two) prior lines of therapy for chronic lymphocytic leukemia (CLL) and / or for small lymphocytic lymphoma (SLL).

[0254] Prior lines of therapy for CLL and / or SLL can particularly include chemoimmunotherapy.

[0255] In other embodiments, prior lines of therapy for CLL and / or SLL include therapy with a targeting agent such as a BCL2 inhibitor or a BTK inhibitor.

[0256] In further embodiments, prior lines of therapy for CLL and / or SLL include CAR T cell therapy.

[0257] In some embodiments, the subject has received prior therapy for Richter's syndrome, such as prior therapy selected from:

[0258] i) rituximab combined with cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP),

[0259] ii) rituximab combined with dexamethasone, cytarabine, and cisplatin (R-DHAP),

[0260] iii) venetoclax combined with rituximab, etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin (VR-EPOCH).

[0261] In some embodiments, the subject treated according to the invention achieves a complete metabolic response or a partial metabolic response.

[0262] In other embodiments, subjects treated according to the invention achieve a complete response, partial response, or disease stabilization.

[0263] In other embodiments, a subject receives acoramab according to the invention as a first-line therapy for Richter's syndrome. In further embodiments, the method according to the invention is a first-line therapy for Richter's syndrome.

[0264] According to these embodiments, subjects treated according to the invention as a first-line therapy for Richter's syndrome achieve a complete metabolic response or a partial metabolic response.

[0265] In other embodiments, wherein a subject is treated according to the invention as a first-line therapy for Richter's syndrome, the subject achieves a complete response, partial response, or disease stabilization.

[0266] In some embodiments, a human subject must have a clinical history of CLL / SLL with biopsy-proven transformation to aggressive lymphoma (i.e., DLBCL subtype). In some embodiments, a human subject is considered ineligible for chemoimmunotherapy or refuses intensive chemotherapy, at the discretion of the investigator.

[0267] In some embodiments, a human subject must have measurable disease as determined by both: a) fluorodeoxyglucose (FDG)-positron emission tomography (PET) CT scan, which shows positive lesions compatible with the anatomic tumor sites determined by CT (or MRI); and b) CT scan (or MRI) with ≥2 well-defined lesions / nodules with a long axis >1.5 cm and short axis >1.0 cm or 1 well-defined lesion / nodule with a long axis >2.0 cm and short axis ≥1.0 cm.

[0268] In some embodiments, the ECOG performance status score of a human subject is 0 or 2. Information regarding the ECOG performance status score can be found, for example, in Oken et al., Am J Clin Oncol 1982 Dec;5(6):649-55).

[0269] In some embodiments, the human subject has acceptable laboratory parameters: (1) creatinine clearance or serum creatinine (using the Cockcroft-Gault formula > 45 mL / min or serum creatinine ≤ 1.5 times the upper limit of normal (x ULN)), (2) serum alanine transaminase (≤ 2.5 × ULN), (3) serum aspartate transaminase (≤ 2.5 × ULN), (4) bilirubin (≤ 1.5 × ULN, unless due to Gilbert syndrome), (5) hemoglobin (≥ 9.0 g / dL, unless anemia is due to bone marrow involvement in CLL), (6) absolute neutrophil count (≥ 1.0 × 10 9 / L (1000 / μL), unless neutropenia is due to bone marrow involvement in CLL), platelet count (≥ 30 × 10 9 / L (30000 / μL)), and coagulation status (PT / INR / aPTT ≤ 1.5 × ULN).

[0270] The human subject receiving the treatment described herein can be a patient with one or more inclusion criteria listed in Example 2, or a patient without one or more exclusion criteria listed in Example 2.

[0271] Human subjects with Richter's syndrome are classified as having a CD20-positive cancer. Thus, prior cancer treatments that such human subjects can receive include anti-CD20 monoclonal antibodies (e.g., rituximab). During such treatment or any other treatment, RS may be refractory to the treatment or have relapsed. Thus, in one embodiment, the subject has received treatment with an anti-CD20 monoclonal antibody such as rituximab or obinutuzumab prior to treatment with the bispecific antibody. In some embodiments, during the prior treatment with an anti-CD20 antibody or a combination of an anti-CD20 monoclonal antibody with a therapeutic agent (e.g., cyclophosphamide, doxyrubicin hydrochloride, vincristine sulfate, prednisone (R-CHOP)), RS relapsed or was refractory to the treatment.

[0272] The methods described herein are advantageous for treating Richter's syndrome. However, treatment can be terminated when disease progression occurs or unacceptable toxicity appears.

[0273] The response of RS patients to the methods described herein can be evaluated according to the Lugano criteria (Cheson et al., 2014), as shown in Table 2.

[0274] Table 2: Lugano Criteria (Cheson et al., 2014)

[0275]

[0276]

[0277]

[0278]

[0279] Abbreviations: 5PS = 5-point scale; CT = computed tomography; FDG = fluorodeoxyglucose; IHC = immunohistochemistry; LDi = longest diameter of the lesion; MRI = magnetic resonance imaging; PET = positron emission tomography; PPD = cross product of LDi and the perpendicular diameter; SDi = shortest axis perpendicular to LDi; SPD = sum of the products of the perpendicular diameters of multiple lesions.

[0280] 1. A score of 3 for many subjects (especially if at intermediate scans) indicates a good prognosis with standard treatment. However, in trials where PET is involved in the investigation of downstaging, a score of 3 may preferably be considered an inadequate response (to avoid undertreatment).

[0281] • Measured dominant (target) lesions: Select up to six of the largest dominant nodules, nodular masses, and extranodal lesions that can be clearly measured in two diameters.

[0282] o Nodules should preferably be from different regions of the body and should include the mediastinal and retroperitoneal regions where applicable.

[0283] o Non-nodular lesions include lesions in solid organs (e.g., liver, spleen, kidney, lung), gastrointestinal involvement, skin lesions, or lesions noted on palpation.

[0284] • Non-measured lesions: Any disease, dominant disease, and truly evaluable disease that is not selected for measurement should be considered non-measured.

[0285] o These sites include any nodules, nodular masses, and extranodal sites that are not selected as dominant or measurable or do not meet the requirements for measurability but are still considered abnormal, as well as truly evaluable disease, which is any site of suspected disease that will be difficult to quantitatively follow up by measurement, including pleural effusion, ascites, bone lesions, leptomeningeal disease, abdominal masses, and other lesions that cannot be confirmed and followed up by imaging.

[0286] • In the Waldeyer's ring or extranodal sites (such as the GI tract, liver, bone marrow), FDG uptake can be greater than that in the mediastinum with a complete metabolic response, but should not be higher than the surrounding normal physiological uptake (such as bone marrow activation caused by chemotherapy or myeloid growth factors).

[0287] 2. PET 5PS: 1 = no uptake above background; 2 = uptake ≤ mediastinum; 3 = uptake > mediastinum but ≤ liver; 4 = uptake moderately > liver; 5 = uptake significantly higher than liver and / or new lesions; X = new uptake areas unlikely to be related to lymphoma. Source: (Cheson et al., 2014, J Clin Oncol 32, 3059 - 3068).

[0288] Subjects treated according to the methods described herein preferably experience an improvement in at least one sign of lymphoma.

[0289] In one embodiment, the treated subjects exhibit a complete metabolic response or a partial metabolic response as measured by PET (see Table 2, PET - CT - based response), and Figure 4 it is shown that 8 out of 9 RS patients achieved at least a 50% reduction in tumor compared to baseline.

[0290] In one embodiment, the treated subjects exhibit a complete response (CR), partial response (PR), or stable disease (SD) as defined by the Lugano criteria (Cheson et al., 2014) (see, for example, Table 2). As Figure 4 shown, 8 out of 9 RS patients achieved at least a 50% reduction in tumor size compared to baseline.

[0291] In some embodiments, the methods described herein produce at least one therapeutic effect selected from extended survival (such as progression - free survival or overall survival), optionally compared to another therapy or placebo. In some embodiments, subjects are treated with the methods described herein until disease progression (PD) or unacceptable toxicity.

[0292] When using methods in human subjects, cytokine release syndrome (CRS) may occur, which utilize immune cell- and bispecific antibody-based approaches that act by activating immune effector cells, such as by engaging CD3 (Lee et al., Biol Blood Marrow Transplant 2019; 25:625-38, which is incorporated herein by reference). Thus, in some embodiments, CRS mitigation is conducted in conjunction with the methods described herein. As part of CRS mitigation, as described herein, a sensitizing dose and / or intermediate dose is selected prior to administration of the full dose (e.g., 12-60 mg). CRS can be classified according to standard practice (e.g., as outlined in Lee et al., Biol Blood Marrow Transplant. 2019 Apr;25(4):625-638, which is incorporated herein by reference). CRS can include the over-release of cytokines, such as pro-inflammatory cytokines, such as IL-6, TNF-α, or IL-8, which can lead to adverse effects, such as fever, nausea, vomiting, and chills. Thus, although bispecific antibodies such as acalabrutinib have unique anti-tumor activity, their immunological mode of action may trigger unwanted "side" effects, namely, induction of an unwanted inflammatory response. Accordingly, patients may further undergo concomitant therapy, prophylaxis, and / or premedication with, for example, painkillers, antipyretics, and / or anti-inflammatory drugs to mitigate possible CRS symptoms.

[0293] Thus, in one embodiment, a human subject in the methods described herein is treated for prevention of CRS. In a preferred embodiment, the prevention comprises administering a corticosteroid to the subject. In one embodiment, the prevention (e.g., corticosteroid) is administered on the same day as the bispecific antibody. The prevention (e.g., corticosteroid) can also be administered on a subsequent day. In some embodiments, the prevention (e.g., corticosteroid) is further administered on subsequent days 2, 3, and 4. It should be understood that when referring to further medication (such as prevention), days 2, 3, and 4 are relative to the administration of the bispecific antibody administered on day 1. For example, when the antibody is administered on day 15 of a cycle and prevention is also administered, the prevention corresponding to days 2, 3, and 4 is days 16, 17, and 18 of that cycle. In some embodiments, the prevention is administered on the day of administration of the bispecific antibody and on subsequent days 2 - 4. When the prevention is administered on the same day as the bispecific antibody, it is preferably administered 30 - 120 minutes before the administration of the bispecific antibody. An exemplary corticosteroid suitable for the methods and uses described herein is prednisolone. In some embodiments, the corticosteroid is prednisolone. In some embodiments, prednisolone is administered at an intravenous dose of 100 mg or its equivalent (including oral dose). Exemplary corticosteroid equivalents of prednisolone that can be used for CRS prevention, as well as dose equivalents, are shown in Table 6.

[0294] In addition, in some embodiments, a human subject in the methods described herein is treated with premedication to reduce the reaction to the injection. In one embodiment, the premedication comprises administration of an antihistamine. In some embodiments, the premedication comprises administration of an antipyretic. In a further embodiment, the premedication comprises systemic administration of an antihistamine and an antipyretic.

[0295] An exemplary antihistamine suitable for premedication is diphenhydramine. In some embodiments, the antihistamine is diphenhydramine. In one embodiment, diphenhydramine is administered at an intravenous or oral dose of 50 mg or its equivalent. An exemplary antipyretic suitable for premedication is acetaminophen. In some embodiments, the antipyretic is acetaminophen. In one embodiment, acetaminophen is administered at an oral dose of 560 - 1000 mg (such as 650 - 1000 mg) or its equivalent. In some embodiments, the premedication is administered on the same day as the bispecific antibody. In some embodiments, the premedication is administered on the same day as the bispecific antibody, for example, 30 - 120 minutes before the administration of the bispecific antibody.

[0296] Preliminary medication and / or prophylaxis can be administered at least during the initial stage of treatment. In some embodiments, the preliminary medication and / or prophylaxis are administered during the first four administrations of the bispecific antibody. For example, the preliminary medication and / or prophylaxis can be administered as described herein during the first 28-day cycle of bispecific antibody administration. In some embodiments, the preliminary medication is administered during Cycle 1. In some embodiments, the prophylaxis is administered during Cycle 1.

[0297] Generally, the risk of reaction during the initial treatment subsides after several administrations, e.g., after the first four administrations (the first cycle). Thus, and when a human subject does not experience CRS, prophylaxis against CRS can be discontinued. However, when a human subject experiences CRS greater than Grade 1, CRS prophylaxis can be continued. Similarly, the preliminary medication can also be optionally continued. CRS grading can be performed as described in Tables 7 and 8.

[0298] In a further embodiment, in the method described herein, when a human subject experiences CRS greater than Grade 1 after the fourth (i.e., last) administration of the bispecific antibody in Cycle 1, prophylaxis is administered during the second 28-day cycle (i.e., Cycle 2). In addition, when a human subject experiences CRS greater than Grade 1 at the last administration of the bispecific antibody in the previous cycle, prophylaxis can be continued during subsequent cycles. Any preliminary medication can be optionally administered during the second cycle. In some embodiments, prophylaxis is administered during Cycle 2. Further preliminary medication can also be optionally administered in subsequent cycles. In some embodiments, the preliminary medication is administered during subsequent cycles (after Cycle 2).

[0299] In one embodiment, a preliminary medication and prophylaxis against CRS are administered, wherein the preliminary medication comprises an antihistamine such as diphenhydramine (e.g., at an intravenous or oral dose of 50 mg or its equivalent), and the prophylaxis comprises an antipyretic such as acetaminophen (e.g., at an oral dose of 650 - 1000 mg or its equivalent), and a corticosteroid such as prednisolone (e.g., at an intravenous dose of 100 mg or its equivalent). In some embodiments, the preliminary medication and prophylaxis are administered 30 - 120 minutes before the administration of the bispecific antibody. On Days 2 and 3 and optionally Day 4 thereafter, further prophylaxis is administered, including systemic administration of a corticosteroid such as prednisolone (e.g., at an intravenous dose of 100 mg or its equivalent). In some embodiments, the preliminary medication and prophylaxis schedule are preferably administered during the first four administrations of the bispecific antibody, e.g., during the first 28-day cycle of bispecific antibody administration as described herein. In addition, in the case where CRS greater than Grade 1 occurs, for example, at the last administration of the previous cycle, subsequent cycles can include the same administration schedule, wherein the preliminary medication as part of the administration schedule is optional.

[0300] During treatment of human subjects with RS using the dosages and treatment regimens described herein, CRS can be well managed while effectively controlling and / or treating RS. As described in the examples, subjects treated with the methods described herein may experience manageable CRS. In some cases, subjects receiving the treatment described herein may develop grade 1 CRS as defined by standard practice. In other cases, subjects may develop manageable grade 2 CRS as defined by standard practice. Thus, subjects receiving the treatment described herein may have manageable grade 1 or 2 CRS as defined by standard practice. According to the standard classification of CRS, grade 1 CRS includes fever to at least 38°C, no hypotension, and no hypoxia, and grade 2 CRS includes fever to at least 38°C plus hypotension not requiring vasopressors and / or hypoxia requiring oxygen by low-flow nasal cannula or blow-by. Such manageable CRS may occur in cycle 1. Human subjects receiving the treatment described herein may also have CRS greater than grade 2 as defined by standard practice during treatment. Thus, human subjects receiving the treatment described herein may also have grade 3 CRS as defined by standard practice during said treatment. Such manageable CRS may further occur during cycle 1 and subsequent cycles.

[0301] Human subjects treated according to the methods described herein may also experience fever, fatigue, and injection site reactions. They may also experience neurotoxicity, partial seizures, agraphia associated with CRS, or confusional states associated with CRS.

[0302] As described above, although CRS prophylaxis has been received, subjects may develop CRS during treatment with the methods described herein. CRS grading criteria are described in Tables 7 and 8.

[0303] In one embodiment, if a subject develops grade 1 CRS, an antibiotic is administered to the subject, i.e., a subject who develops grade 1 CRS is treated with an antibiotic if an infection occurs. In some embodiments, the antibiotic is continued until neutropenia (if present) resolves. In some embodiments, subjects with grade 1 CRS presenting with systemic symptoms are treated with NSAIDs.

[0304] In one embodiment, subjects who develop grade 2 CRS are treated with an intravenous fluid bolus and / or oxygen supplementation. In some embodiments, subjects who develop grade 2 CRS are treated with vasopressors. In some embodiments, subjects with grade 2 CRS with comorbidities are treated with tocilizumab (a humanized antibody against the IL-6 receptor, such as ACTEMRA ®Treatment with commercially available) and / or steroids (e.g., dexamethasone or its methylprednisolone equivalent). In a further embodiment, dexamethasone is administered to a subject presenting with concurrent ICANS. In yet another embodiment, if a subject does not show improvement in CRS symptoms within, for example, 6 hours, or if a subject begins to deteriorate after initial improvement, the second dose of tocilizumab is administered together with a dose of corticosteroid. In some embodiments, if a subject is refractory to tocilizumab after three administrations, additional cytokine therapy, such as an anti-IL-6 antibody (e.g., siltuximab) or an IL-1R antagonist (e.g., anakinra), is administered to the subject.

[0305] In one embodiment, a subject developing grade 3 CRS is supported with vasopressors (e.g., norepinephrine) and / or treated with oxygen supplementation. In some embodiments, a subject with grade 3 CRS is treated with tocilizumab, or tocilizumab in combination with a steroid (e.g., dexamethasone or its methylprednisolone equivalent). In some embodiments, dexamethasone is administered to a subject presenting with concurrent ICANS. In a further embodiment, if a subject is refractory to tocilizumab after three administrations, additional cytokine therapy, such as an anti-IL-6 antibody (e.g., siltuximab) or an IL-1R antagonist (e.g., anakinra), is administered to the subject.

[0306] In one embodiment, a subject developing grade 4 CRS is supported with vasopressors and / or treated with oxygen supplementation (e.g., via positive pressure ventilation, such as CPAP, BiPAP, intubation, or mechanical ventilation). In some embodiments, if a subject develops grade 4 CRS, at least two vasopressors are administered to the subject. In some embodiments, a steroid is further administered to the subject, i.e., tocilizumab and a steroid are administered to the subject. In some embodiments, the steroid is dexamethasone. In some embodiments, the steroid is methylprednisolone. In a further embodiment, dexamethasone is administered to a subject presenting with concurrent ICANS. In yet a further embodiment, if a subject is refractory to tocilizumab after three administrations, additional cytokine therapy, such as an anti-IL-6 antibody (e.g., siltuximab) or an IL-1R antagonist (e.g., anakinra), is administered to the subject. In some embodiments, if a subject is refractory to tocilizumab, the administration of tocilizumab is switched to the administration of an anti-IL-6 antibody (e.g., siltuximab). In some embodiments, if a subject is refractory to tocilizumab, tocilizumab is switched to an IL-1R antagonist (e.g., anakinra).

[0307] In some embodiments, a human subject receives prophylactic treatment for tumor lysis syndrome (TLS), i.e., the subject is treated with prophylaxis against tumor lysis syndrome (TLS). Classification and grading of tumor lysis syndrome can be performed using methods known in the art, such as those described in Howard et al., N Engl J Med 2011;364:1844-54 and Coiffier et al., J Clin Oncol 2008;26:2767-78. In some embodiments, prophylactic treatment for TLS includes administering one or more urate-lowering agents prior to administration of the bispecific antibody, i.e., prophylaxis against TLS includes administering one or more urate-lowering agents prior to administration of the bispecific antibody. Exemplary urate-lowering agents include allopurinol and rasburicase. Thus, in one embodiment, prophylactic treatment for TLS includes administering allopurinol and / or rasburicase. In some embodiments, prophylactic treatment for TLS includes administering allopurinol and / or rasburicase prior to administration of the bispecific antibody. In one embodiment, allopurinol is administered 72 hours prior to the bispecific antibody. In some embodiments, rasburicase is initiated after administration of allopurinol but prior to administration of the bispecific antibody. Prior to subsequent doses of the bispecific antibody, a re-evaluation of the subject's TLS risk category can be performed. If the maximum diameter of all measurable lymph nodes < 5 cm and ALC < 25×10 9 / L, the subject is considered to be at low risk for TLS. If the maximum diameter of any measurable lymph node ≥ 5 cm but < 10 cm or ALC ≥ 25×10 9 / L, the subject is considered to be at moderate risk for TLS. If (a) the maximum diameter of any measurable lymph node ≥ 10 cm, or (b) ALC ≥ 25×10 9 / L and the maximum diameter of any measurable lymph node ≥ 5 cm but < 10 cm, the subject is considered to be at high risk for TLS. Subjects with lymphocyte count > 100×10 9 / L are considered high risk. In some embodiments, when a subject shows signs of TLS, supportive therapies such as rasburicase and / or allopurinol can be used.

[0308] In one embodiment, the bispecific antibody used in the methods described herein is administered subcutaneously and is thus formulated in a pharmaceutical composition to be compatible with subcutaneous (s.c.) administration, i.e., having a formulation and / or concentration that permits pharmaceutically acceptable s.c. administration at the doses described herein. In some embodiments, subcutaneous administration is by injection. For example, a formulation of DuoBody-CD3xCD20 that is compatible with a subcutaneous formulation and can be used in the methods described herein has been previously described (see, e.g., WO2019155008, which is incorporated herein by reference). In some embodiments, the bispecific antibody can be formulated using sodium acetate trihydrate, acetic acid, sodium hydroxide, sorbitol, polysorbate 80, and water for injection and has a pH of 5.5 or about 5.5. In some embodiments, the bispecific antibody is provided as a 5 mg / mL or 60 mg / mL concentrate. In other embodiments, the desired dose of the bispecific antibody is reconstituted to a volume of about 1 mL for subcutaneous injection.

[0309] In one embodiment, a suitable pharmaceutical composition for a bispecific antibody can comprise the bispecific antibody, 20 - 40 mM acetate, 140 - 160 mM sorbitol, and a surfactant such as polysorbate 80, and has a pH of 5.3 - 5.6. In some embodiments, the pharmaceutical formulation can comprise an antibody concentration in the range of 5 - 100 mg / mL, e.g., 48 or 60 mg / mL of the bispecific antibody, 30 mM acetate, 150 mM sorbitol, 0.04% w / v polysorbate 80, and has a pH of 5.5. Such formulations can be diluted with, e.g., a formulation buffer to allow appropriate dosing and subcutaneous administration.

[0310] The volume of the pharmaceutical composition is appropriately selected to permit subcutaneous administration of the antibody. For example, the volume to be administered ranges from about 0.3 mL to about 3 mL, such as 0.3 mL to 3 mL. The volume to be administered can be 0.5 mL, 0.8 mL, 1 mL, 1.2 mL, 1.5 mL, 1.7 mL, 2 mL, or 2.5 mL, or about 0.5 mL, about 0.8 mL, about 1 mL, about 1.2 mL, about 1.5 mL, about 1.7 mL, about 2 mL, or about 2.5 mL. Thus, in some embodiments, the volume to be administered is 0.5 mL or about 0.5 mL. In some embodiments, the volume to be administered is 0.8 mL or about 0.8 mL. In some embodiments, the volume to be administered is 1 mL or about 1 mL. In some embodiments, the volume to be administered is 1.2 mL or about 1.2 mL. In some embodiments, the volume to be administered is 1.5 mL or about 1.5 mL. In some embodiments, the volume to be administered is 1.7 mL or about 1.7 mL. In some embodiments, the volume to be administered is 2 mL or about 2 mL. In some embodiments, the volume to be administered is 2.5 mL or about 2.5 mL.

[0311] The methods (or uses of the CD3xCD20 antibody) described herein are for treating human patients suffering from CLL. It is understood that the methods described herein can be the first or a first partial treatment provided to such patients. However, the patient may have undergone prior treatment for CLL and / or Richter's syndrome. Prior treatment can include, but is not limited to, one or more of chemotherapy, immunotherapy, and targeted therapy, or combinations thereof. Most commonly, the standard of care for RS involves treatment with a combination of cytotoxic chemotherapy and anti-CD20 monoclonal antibodies. It is understood that the methods described herein can also be used in combination with other treatments.

[0312] In one embodiment, the bispecific antibody for use in the methods described herein comprises:

[0313] (i) a first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a variable heavy chain (VH) region and a variable light chain (VL) region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences within the amino acid sequence of SEQ ID NO: 6, and the VL region comprises the CDR1, CDR2, and CDR3 sequences within the amino acid sequence of SEQ ID NO: 7; and

[0314] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region and a VL region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences within the amino acid sequence of SEQ ID NO: 13, and the VL region comprises the CDR1, CDR2, and CDR3 sequences within the amino acid sequence of SEQ ID NO: 14.

[0315] The CDR1, CDR2, and CDR3 regions can be identified from the variable heavy chain region and the variable light chain region using methods known in the art. The CDR regions from the variable heavy chain region and the variable light chain region can be annotated according to IMGT (see Lefranc et al., Nucleic Acids Research 1999;27:209-12 and Brochet. Nucl Acids Res 2008;36:W503-8).

[0316] In some embodiments, the bispecific antibody comprises:

[0317] (i) A first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises VHCDR1, VHCDR2, and VHCDR3 comprising the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprising the amino acid sequences set forth in SEQ ID NOs: 4, the GTN sequence, and SEQ ID NO: 5, respectively; and

[0318] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises VHCDR1, VHCDR2, and VHCDR3 comprising the amino acid sequences set forth in SEQ ID NOs: 8, 9, and 10, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprising the amino acid sequences set forth in SEQ ID NOs: 11, the sequence DAS, and SEQ ID NO: 12, respectively.

[0319] In some embodiments, the bispecific antibody comprises:

[0320] (i) A first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a VH region having the amino acid sequence of SEQ ID NO: 6 and a VL region having the amino acid sequence of SEQ ID NO: 7; and

[0321] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region having the amino acid sequence of SEQ ID NO: 13 and a VL region having the amino acid sequence of SEQ ID NO: 14.

[0322] In some embodiments, the bispecific antibody is a full-length antibody. In some embodiments, the bispecific antibody comprises an inert Fc region. In one embodiment, the bispecific antibody is a full-length antibody and may have an inert Fc region. In some embodiments, the first binding arm targeting CD3 is derived from a humanized antibody, such as derived from a full-length IgG1, λ (lambda) antibody, such as H1L1 as described in WO2015001085 (which is incorporated herein by reference), and / or the second binding arm targeting CD20 is derived from a human antibody, such as derived from a full-length IgG1, κ (kappa) antibody, such as clone 7D8 as described in WO2004035607 (which is incorporated herein by reference). The bispecific antibody can be produced from two half-antibodies, each of the two half-antibodies comprising the respective first and second binding arms listed, for example, in SEQ ID NOs: 24 and 25 and SEQ ID NOs: 26 and 27. The half-antibodies can be produced in CHO cells, and the bispecific antibody can be generated, for example, by Fab-arm exchange. In one embodiment, the bispecific antibody is a functional variant of DuoBody-CD3xCD20.

[0323] Thus, in some embodiments, the bispecific antibody comprises (i) a first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a VH region having an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 6 or a VH region having the amino acid sequence of SEQ ID NO: 6 but having 1, 2 or 3 mutations (e.g., amino acid substitutions), and a VL region having an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to SEQ ID NO: 7 or a VL region having the amino acid sequence of SEQ ID NO: 7 but having 1, 2 or 3 mutations (e.g., amino acid substitutions); and

[0324] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region having an amino acid sequence that is at least 85%, 90%, 95%, 98% or 99% identical to SEQ ID NO: 13 or a VH region having the amino acid sequence of SEQ ID NO: 13 but having 1, 2 or 3 mutations (e.g., amino acid substitutions), and a VL region having an amino acid sequence that is at least 85%, 90%, 95%, 98% or 99% identical to SEQ ID NO: 14 or a VL region having the amino acid sequence of SEQ ID NO: 14 but having 1, 2 or 3 mutations (e.g., amino acid substitutions).

[0325] In one embodiment, the bispecific antibody comprises:

[0326] (i) A first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a heavy chain having the amino acid sequence of SEQ ID NO: 24 and a light chain having the amino acid sequence of SEQ ID NO: 25; and

[0327] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region having the amino acid sequence of SEQ ID NO: 26 and a VL region having the amino acid sequence of SEQ ID NO: 27.

[0328] In some embodiments, the bispecific antibody comprises (i) a first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a heavy chain having an amino acid sequence that is at least 85%, 90%, 95%, 98% or 99% identical to SEQ ID NO: 24 or a heavy chain having the amino acid sequence of SEQ ID NO: 24 but having 1, 2 or 3 mutations (e.g., amino acid substitutions), and a light chain having an amino acid sequence that is at least 85%, 90%, 95%, 98% or 99% identical to SEQ ID NO: 25 or a light chain having the amino acid sequence of SEQ ID NO: 25 but having 1, 2 or 3 mutations (e.g., amino acid substitutions); and

[0329] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a heavy chain having an amino acid sequence that is at least 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 26 or a heavy chain having the amino acid sequence of SEQ ID NO: 26 but having 1, 2, or 3 mutations (e.g., amino acid substitutions), and a light chain having an amino acid sequence that is at least 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 27 or a light chain having the amino acid sequence of SEQ ID NO: 27 but having 1, 2, or 3 mutations (e.g., amino acid substitutions).

[0330] A variety of constant regions or variants thereof can be used in bispecific antibodies. In one embodiment, the antibody comprises an IgG constant region, such as a human IgG1 constant region, e.g., the human IgG1 constant region defined in SEQ ID NO: 15, or any other suitable IgG1 allotype. In some embodiments, the bispecific antibody is a full-length antibody having a human IgG1 constant region. In some embodiments, the first binding arm of the bispecific antibody is derived from a humanized antibody, preferably derived from a full-length IgG1, λ (lambda) antibody. In one embodiment, the first binding arm of the bispecific antibody is derived from a humanized antibody, e.g., derived from a full-length IgG1, λ (lambda) antibody, and thus comprises a λ light chain constant region. In some embodiments, the first binding arm comprises the λ light chain constant region defined in SEQ ID NO: 22. In some embodiments, the second binding arm of the bispecific antibody is derived from a human antibody, preferably from a full-length IgG1, κ (kappa) antibody. In some embodiments, the second binding arm of the bispecific antibody is derived from a human antibody, preferably derived from a full-length IgG1, κ (kappa) antibody, and thus may comprise a κ light chain constant region. In some embodiments, the second binding arm comprises the κ light chain constant region defined in SEQ ID NO: 23. In a preferred embodiment, the first binding arm comprises the λ light chain constant region defined in SEQ ID NO: 22 and the second binding arm comprises the κ light chain constant region defined in SEQ ID NO: 23.

[0331] It should be understood that the constant region portion of the bispecific antibody may comprise modifications that allow for efficient formation / production of the bispecific antibody and / or provide an inert Fc region. Such modifications are well known in the art.

[0332] Bispecific antibodies in different formats are known in the art (reviewed by Kontermann, Drug Discov Today 2015;20:838-47; MAbs, 2012;4:182-97). Thus, the bispecific antibodies used in the methods and uses described herein are not limited to any particular bispecific format or method of producing them. For example, bispecific antibodies can include, but are not limited to, bispecific antibodies having complementary CH3 domains to enforce heterodimerization, Knob-into-Hole molecules (Genentech, WO9850431), CrossMAb (Roche, WO2011117329), or electrostatically paired molecules (Amgen, EP1870459 and WO2009089004; Chugai, US201000155133; Oncomed, WO2010129304).

[0333] Preferably, the bispecific antibody comprises an Fc region containing a first heavy chain and a second heavy chain, the first heavy chain having a first Fc sequence containing a first CH3 region and the second heavy chain having a second Fc sequence containing a second CH3 region, wherein the sequences of the first and second CH3 regions are different and such that the heterodimeric interaction between the first and second CH3 regions is stronger than each of the homodimeric interactions of the first and second CH3 regions. More details regarding these interactions and how to achieve them are provided in, for example, WO2011131746 and WO2013060867 (Genmab), which are incorporated herein by reference. In one embodiment, the bispecific antibody comprises (i) an amino acid L at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 in the first heavy chain and an amino acid R at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 in the second heavy chain, or vice versa.

[0334] Bispecific antibodies can comprise modifications in the Fc region to render the Fc region inert or non - activated. Thus, in the bispecific antibodies disclosed herein, one or both of the heavy chains can be modified such that, relative to an unmodified bispecific antibody, the antibody induces a lesser degree of Fc - mediated effector function. Fc - mediated effector function can be measured by determining Fc - mediated CD69 expression on T cells (i.e., CD3 antibody - mediated, Fcγ receptor - dependent CD3 cross - linking - induced CD69 expression), by binding to Fcγ receptors, by binding to Clq, or by inducing Fc - mediated FcγR cross - linking. In particular, the heavy chain constant region sequence can be modified such that, when compared to a wild - type (unmodified) antibody, Fc - mediated CD69 expression is reduced by at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 99% or 100%, wherein the Fc - mediated CD69 expression is determined in a PBMC - based functional assay, such as described in Example 3 of WO2015001085. Modifications of the heavy and light chain constant region sequences can also result in reduced binding of Clq to the antibody. The reduction can be at least 70%, at least 80%, at least 90%, at least 95%, at least 97% or 100% compared to an unmodified antibody, and Clq binding can be determined, for example, by ELISA. In addition, the Fc region can be modified such that the antibody mediates at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 99% or 100% less Fc - mediated T - cell proliferation compared to an unmodified antibody, wherein the T - cell proliferation is measured in a PBMC - based functional assay. Examples of amino acid positions that can be modified, for example in an IgG1 isotype antibody, include positions L234 and L235. Thus, in one embodiment, a bispecific antibody can comprise a first heavy chain and a second heavy chain, and wherein in both the first heavy chain and the second heavy chain, the amino acid residues at the positions corresponding to positions L234 and L235 in the human IgG1 heavy chain according to Eu numbering are F and E, respectively. In addition, the D265A amino acid substitution can reduce binding to all Fcγ receptors and prevent ADCC (Shields et al., JBC 2001;276:6591 - 604). Thus, a bispecific antibody can comprise a first heavy chain and a second heavy chain, wherein in both the first heavy chain and the second heavy chain, the amino acid residue at the position corresponding to position D265 in the human IgG1 heavy chain according to Eu numbering is A.

[0335] In one embodiment, in the first and second heavy chains of a bispecific antibody, the amino acids at the positions corresponding to positions L234, L235 and D265 in the human IgG1 heavy chain are F, E and A, respectively. Antibodies having these amino acids at these positions are examples of antibodies with an inert or non - activated Fc region.

[0336] In some embodiments, the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein in both the first and second heavy chains, the amino acids at positions corresponding to positions L234, L235, and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E, and A, respectively. In some embodiments, the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein in the first heavy chain, the amino acid at the position corresponding to position F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and wherein in the second heavy chain, the amino acid at the position corresponding to position K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa. In a preferred embodiment, the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein (i) in both the first and second heavy chains, the amino acids at positions corresponding to positions L234, L235, and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E, and A, respectively, and (ii) in the first heavy chain, the amino acid at the position corresponding to position F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and wherein in the second heavy chain, the amino acid at the position corresponding to position K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa.

[0337] Regarding the bispecific antibodies described herein, those bispecific antibodies having the combination of three amino acid substitutions L234F, L235E, and D265A and additionally the K409R or F405L mutation, as described above, can be designated with the suffix "FEAR" or "FEAL", respectively.

[0338] The amino acid sequence of the wild-type IgG1 heavy chain constant region can be identified herein as SEQ ID NO: 15. Consistent with the embodiments disclosed above, the bispecific antibody can comprise an IgG1 heavy chain constant region carrying an F405L substitution and can have the amino acid sequence listed in SEQ ID NO: 17 and / or an IgG1 heavy chain constant region carrying a K409R substitution and can have the amino acid sequence listed in SEQ ID NO: 18, and having further substitutions that render the Fc region inert or non-activated. Thus, in one embodiment, the bispecific antibody comprises a combination of IgG1 heavy chain constant regions, wherein the amino acid sequence of one of the IgG1 heavy chain constant regions carries the L234F, L235E, D265A, and F405L substitutions (e.g., as listed in SEQ ID NO: 19) and the amino acid sequence of the other IgG1 heavy chain constant region carries the L234F, L235E, D265A, and K409R substitutions (e.g., as listed in SEQ ID NO: 20). Thus, in some embodiments, the bispecific antibody comprises a heavy chain constant region comprising the amino acid sequences of SEQ ID NOs: 19 and 20.

[0339] In a preferred embodiment, the bispecific antibody for use in the methods and uses described herein comprises a first binding arm comprising a heavy chain and a light chain as defined in SEQ ID NOs: 24 and 25, respectively, and a second binding arm comprising a heavy chain and a light chain as defined in SEQ ID NOs: 26 and 27, respectively. Such antibodies may also be referred to herein as DuoBody-CD3xCD20. In addition, variants of such antibodies are contemplated for use in the methods and uses described herein. In some embodiments, the bispecific antibody comprises a heavy chain and a light chain consisting of the amino acid sequences listed in SEQ ID NOs: 24 and 25, respectively, and a heavy chain and a light chain consisting of the amino acid sequences listed in SEQ ID NOs: 26 and 27, respectively. In some embodiments, the bispecific antibody is acalabrutinib (CAS 2134641-34-0) or a biosimilar thereof.

[0340] Kit

[0341] Also provided herein are kits comprising a pharmaceutical composition comprising a bispecific antibody that binds to CD3 and CD20 according to the invention, such as DuoBody-CD3xCD20 or acalabrutinib, in a therapeutically effective amount suitable for use in the methods described herein, and a pharmaceutically acceptable carrier. The kit may optionally further comprise, for example, instructions including a dosing schedule to allow a practitioner (e.g., a physician, nurse, or patient) to administer the composition contained therein to a patient suffering from CLL. The kit may also include a syringe.

[0342] Optionally, the kit includes a plurality of packaged single doses (e.g., doses between 12 - 60 mg, such as doses of 12 mg, 24 mg, 36 mg, 48 mg, or 60 mg) of a pharmaceutical composition, each containing an effective amount of a bispecific antibody for single administration according to the methods described herein. Instruments or devices necessary for administering the pharmaceutical composition may also be included in the kit. For example, the kit may provide one or more prefilled syringes containing a certain amount of the bispecific antibody.

[0343] Further embodiments

[0344] 1. A method of treating Richter's syndrome (RS) in a human subject, the method comprising administering to the subject a bispecific antibody comprising:

[0345] (i) a first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a variable heavy chain (VH) region and a variable light chain (VL) region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences of the VH region sequence of SEQ ID NO: 6, and the VL region comprises the CDR1, CDR2, and CDR3 sequences of the VL region sequence of SEQ ID NO: 7; and

[0346] (ii) a second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region and a VL region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences of the VH region sequence of SEQ ID NO: 13, and the VL region comprises the CDR1, CDR2, and CDR3 sequences of the VL region sequence of SEQ ID NO: 14;

[0347] wherein the bispecific antibody is administered in a dose ranging from 12 - 60 mg in a 28-day cycle.

[0348] 2. The method of embodiment 1, wherein the bispecific antibody is administered in a dose of 24 mg.

[0349] 3. The method of embodiment 1, wherein the bispecific antibody is administered in a dose of 48 mg.

[0350] 4. The method according to any one of embodiments 1 - 3, wherein the bispecific antibody is administered once a week (once-weekly administration).

[0351] 5. The method of embodiment 4, wherein the once-weekly administration is carried out for 2.5 28-day cycles.

[0352] 6. The method according to embodiment 4 or 5, wherein after said once-weekly administration, the bispecific antibody is administered once every two weeks (bi-weekly administration).

[0353] 7. The method according to embodiment 6, wherein said bi-weekly administration is carried out for six 28-day cycles.

[0354] 8. The method according to embodiment 6 or 7, wherein after said bi-weekly administration, the bispecific antibody is administered once every four weeks.

[0355] 9. The method according to any one of embodiments 4-8, wherein a sensitizing dose of the bispecific antibody is administered in the 1st cycle of said 28-day cycle before administering a first once-weekly dose of 12-60 mg.

[0356] 10. The method according to embodiment 9, wherein said sensitizing dose is administered two weeks before administering a first once-weekly dose of 12-60 mg.

[0357] 11. The method according to embodiment 9 or 10, wherein said sensitizing dose is in the range of 0.05 - 0.35 mg.

[0358] 12. The method according to any one of embodiments 9-11, wherein said sensitizing dose is 0.16 mg or about 0.16 mg.

[0359] 13. The method according to any one of embodiments 9-12, wherein an intermediate dose of the bispecific antibody is administered after administering said sensitizing dose and before administering said first once-weekly dose of 12 - 60 mg.

[0360] 14. The method according to embodiment 13, wherein said sensitizing dose is administered on day 1 and said intermediate dose is administered on day 8 before a first once-weekly dose of 12-60 mg on days 15 and 22 of the 1st cycle.

[0361] 15. The method according to embodiment 13 or 14, wherein said intermediate dose is in the range of 0.6-1.2 mg.

[0362] 16. The method according to any one of embodiments 13-15, wherein said intermediate dose is 0.8 mg or about 0.8 mg.

[0363] 17. The method according to any one of embodiments 13-16, wherein the bispecific antibody is administered in 28-day cycles, wherein:

[0364] a) In the first cycle, a sensitizing dose is administered on day 1, an intermediate dose on day 8, and a full dose of 12 - 60 mg on days 15 and 22;

[0365] b) In the second - third cycles, a full dose of 12 - 60 mg is administered on days 1, 8, 15, and 22;

[0366] c) In the fourth - ninth cycles, a full dose of 12 - 60 mg is administered on days 1 and 15; and

[0367] d) In the tenth cycle and subsequent cycles, a full dose of 12 - 60 mg is administered on day 1.

[0368] 18. The method according to embodiment 17, wherein the full dose is 24 mg or about 24 mg.

[0369] 19. The method according to embodiment 17, wherein the full dose is 48 mg or about 48 mg.

[0370] 20. The method according to any one of embodiments 1 - 19, wherein the bispecific antibody is administered subcutaneously.

[0371] 21. The method according to any one of embodiments 1 - 20, wherein the subject has a clinical history of CLL / SLL that has transformed to aggressive lymphoma; for example, having the DLBCL subtype.

[0372] 22. The method according to any one of embodiments 1 - 21, wherein the Richter's syndrome has the DLBCL subtype.

[0373] 23. The method according to any one of embodiments 1 - 22, wherein the subject has received one or more (such as at least two) previous lines of therapy for chronic lymphocytic leukemia (CLL) and / or for small lymphocytic lymphoma (SLL).

[0374] 24. The method according to any one of the embodiments, wherein the previous line of therapy for CLL and / or SLL includes chemoimmunotherapy.

[0375] 25. The method according to any one of the embodiments, wherein the previous line of therapy for CLL and / or SLL includes therapy using targeting agents such as BCL2 inhibitors or BTK inhibitors.

[0376] 26. The method according to any one of the embodiments, wherein the previous line of therapy for CLL and / or SLL includes CAR T - cell therapy.

[0377] 27. The method according to any one of embodiments 1-23, wherein the subject has received a previous therapy for Richter's syndrome, such as a previous therapy selected from the following:

[0378] i) Rituximab combined with cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP),

[0379] ii) Rituximab combined with dexamethasone, cytarabine, and cisplatin (R-DHAP), and

[0380] iii) Venetoclax combined with rituximab, etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin (VR-EPOCH).

[0381] 28. The method according to any one of embodiments 1-27, wherein the subject achieves a complete metabolic response or a partial metabolic response.

[0382] 29. The method according to any one of embodiments 1-27, wherein the subject achieves a complete response, a partial response, or disease stabilization.

[0383] 30. The method according to any one of embodiments 1-26, wherein the subject receives acelumab as a first-line therapy for Richter's syndrome.

[0384] 31. The method according to any one of embodiments 1-30, wherein the subject achieves a complete metabolic response or a partial metabolic response.

[0385] 32. The method according to any one of embodiments 1-30, wherein the subject achieves a complete response, a partial response, or disease stabilization.

[0386] 33. The method according to any one of embodiments 1-26 and 30-32, which is a first-line therapy for Richter's syndrome.

[0387] 34. The method according to any one of embodiments 21-29, wherein the subject has refractory and / or relapsed Richter's syndrome after receiving the previous therapy.

[0388] 35. The method according to any one of embodiments 1-34, wherein the subject is treated with prophylaxis against cytokine release syndrome (CRS).

[0389] 36. The method according to embodiment 35, wherein the prophylaxis comprises administering a corticosteroid to the subject.

[0390] 37. The method according to embodiment 35 or 36, wherein the corticosteroid is administered on the same day as the bispecific antibody.

[0391] The method according to embodiment 37, wherein the corticosteroid is further administered on the second, third, and fourth days after administration of the bispecific antibody.

[0392] 39. The method according to any one of embodiments 36 - 38, wherein the corticosteroid is prednisolone.

[0393] 40. The method according to embodiment 39, wherein the prednisolone is administered at an intravenous dose of 100 mg or its equivalent including an oral dose.

[0394] 41. The method according to any one of embodiments 1 - 40, wherein a premedication is administered to the subject to reduce the reaction to the injection.

[0395] 42. The method according to embodiment 41, wherein the premedication includes an antihistamine.

[0396] 43. The method according to embodiment 42, wherein the antihistamine is diphenhydramine.

[0397] 44. The method according to embodiment 43, wherein the diphenhydramine is administered at an intravenous or oral dose of 50 mg or its equivalent.

[0398] 45. The method according to any one of embodiments 41 - 44, wherein the premedication includes an antipyretic.

[0399] 46. The method according to embodiment 45, wherein the antipyretic is acetaminophen.

[0400] 47. The method according to embodiment 46, wherein the acetaminophen is administered at an oral dose of 560 to 1000 mg or its equivalent.

[0401] 48. The method according to any one of embodiments 41 - 47, wherein the premedication is administered on the same day as the bispecific antibody.

[0402] 49. The method according to any one of embodiments 35 - 48, wherein the prophylaxis is administered during cycle 1.

[0403] 50. The method according to any one of embodiments 41 - 49, wherein the premedication is administered during cycle 1.

[0404] 51. The method according to embodiment 49 or 50, wherein when the subject experiences CRS greater than grade 1 after the last administration of the bispecific antibody in cycle 1, the prophylaxis is administered during cycle 2.

[0405] 52. The method according to embodiment 51, wherein when the subject experiences CRS greater than grade 1 in the last administration of the bispecific antibody in the previous cycle, the prophylaxis is continued in subsequent cycles.

[0406] 53. The method according to any one of embodiments 41 - 52, wherein the premedication is administered during cycle 2.

[0407] 54. The method according to embodiment 53, wherein the premedication is administered during subsequent cycles.

[0408] 55. The method according to any one of embodiments 1 - 54, wherein if the subject develops grade 1 CRS, antibiotics are administered to the subject.

[0409] 56. The method according to any one of embodiments 1 - 54, wherein if the subject develops grade 2 or 3 CRS, vasopressors are administered to the subject.

[0410] 57. The method according to any one of embodiments 1 - 54, wherein if the subject develops grade 4 CRS, at least two vasopressors are administered to the subject.

[0411] 58. The method according to any one of embodiments 1 - 57, wherein if the subject develops grade 2, 3, or 4 CRS, tocilizumab is administered to the subject.

[0412] 59. The method according to embodiment 58, wherein steroids are further administered to the subject.

[0413] 60. The method according to embodiment 59, wherein the steroid is dexamethasone.

[0414] 61. The method according to embodiment 59, wherein the steroid is methylprednisolone.

[0415] 62. The method according to any one of embodiments 58 - 61, wherein if the subject is refractory to tocilizumab, tocilizumab is switched to an anti - IL - 6 antibody (e.g., siltuximab).

[0416] 63. The method according to any one of embodiments 58 - 61, wherein if the subject is refractory to tocilizumab, tocilizumab is switched to an IL - 1R antagonist (e.g., anakinra).

[0417] 64. The method according to any one of embodiments 1 - 63, wherein the subject is treated with prophylaxis against tumor lysis syndrome (TLS).

[0418] The method according to embodiment 64, wherein the prophylaxis against TLS comprises administering one or more urate-lowering agents before administering the bispecific antibody.

[0419] The method according to embodiment 65, wherein the one or more urate-lowering agents comprise rasburicase and / or allopurinol.

[0420] The method according to any one of embodiments 1-66, wherein the subject achieves a complete response, a partial response, or disease stabilization.

[0421] The method according to any one of embodiments 1-67, wherein:

[0422] (i) The first antigen-binding region comprises VHCDR1, VHCDR2, and VHCDR3, and VLCDR1, VLCDR2, and VLCDR3, wherein VHCDR1, VHCDR2, and VHCDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 4, the sequence GTN, and SEQ ID NO: 5, respectively; and

[0423] (ii) The second antigen-binding region comprises VHCDR1, VHCDR2, and VHCDR3, and VLCDR1, VLCDR2, and VLCDR3, wherein VHCDR1, VHCDR2, and VHCDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 8, 9, and 10, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 11, the sequence DAS, and SEQ ID NO: 12, respectively.

[0424] The method according to any one of embodiments 1-68, wherein:

[0425] (i) The first antigen-binding region comprises a VH region comprising the amino acid sequence of SEQ ID NO: 6 and a VL region comprising the amino acid sequence of SEQ ID NO: 7; and

[0426] (ii) The second antigen-binding region comprises a VH region comprising the amino acid sequence of SEQ ID NO: 13 and a VL region comprising the amino acid sequence of SEQ ID NO: 14.

[0427] The method according to any one of embodiments 1-68, wherein the first binding arm of the bispecific antibody is derived from a humanized antibody, preferably derived from a full-length IgG1, λ (lambda) antibody.

[0428] 71. The method according to embodiment 70, wherein the first binding arm of the bispecific antibody comprises a lambda light chain constant region, and the lambda light chain constant region comprises the amino acid sequence set forth in SEQ ID NO: 22.

[0429] 72. The method according to any one of embodiments 1-71, wherein the second binding arm of the bispecific antibody is derived from a human antibody, preferably derived from a full-length IgG1,κ (kappa) antibody.

[0430] 73. The method according to embodiment 72, wherein the second binding arm comprises a kappa light chain constant region, and the kappa light chain constant region comprises the amino acid sequence set forth in SEQ ID NO: 23.

[0431] 74. The method according to any one of embodiments 1-73, wherein the bispecific antibody is a full-length antibody having a human IgG1 constant region.

[0432] 75. The method according to any one of embodiments 1-74, wherein the bispecific antibody comprises an inert Fc region.

[0433] 76. The method according to any one of embodiments 1-75, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein in both the first heavy chain and the second heavy chain, the amino acids at the positions corresponding to positions L234, L235 and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E and A, respectively.

[0434] 77. The method according to any one of embodiments 1-76, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein in the first heavy chain, the amino acid at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and wherein in the second heavy chain, the amino acid at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa.

[0435] 78. The method according to any one of embodiments 1-77, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein

[0436] (i) in both the first heavy chain and the second heavy chain, the amino acids at the positions corresponding to positions L234, L235 and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E and A, respectively, and

[0437] (ii) In the first heavy chain, the amino acid at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and in the second heavy chain, the amino acid at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa.

[0438] 79. The method according to embodiment 78, wherein the bispecific antibody comprises a heavy chain constant region comprising the amino acid sequences of SEQ ID NOs: 19 and 20.

[0439] 80. The method according to any one of embodiments 1-79, wherein the bispecific antibody comprises heavy and light chains comprising the amino acid sequences listed in SEQ ID NOs: 24 and 25, respectively, and heavy and light chains comprising the amino acid sequences listed in SEQ ID NOs: 26 and 27, respectively.

[0440] 81. The method according to any one of embodiments 1-80, wherein the bispecific antibody comprises heavy and light chains consisting of the amino acid sequences of SEQ ID NOs: 24 and 25, respectively, and heavy and light chains consisting of the amino acid sequences of SEQ ID NOs: 26 and 27, respectively.

[0441] 82. The method according to any one of embodiments 1-81, wherein the bispecific antibody is acalabrutinib or a biosimilar thereof.

[0442] 1a. A bispecific antibody, comprising:

[0443] (i) A first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a variable heavy chain (VH) region and a variable light chain (VL) region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences in the VH region sequence of SEQ ID NO: 6, and the VL region comprises the CDR1, CDR2, and CDR3 sequences in the VL region sequence of SEQ ID NO: 7; and

[0444] (ii) A second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region and a VL region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences in the VH region sequence of SEQ ID NO: 13, and the VL region comprises the CDR1, CDR2, and CDR3 sequences in the VL region sequence of SEQ ID NO: 14;

[0445] It is used for the treatment of Richter's syndrome (RS) in human subjects, and the treatment includes administering the bispecific antibody to the subject at a dose in the range of 12 - 60 mg in a 28-day cycle.

[0446] 2a. The bispecific antibody used in Embodiment 1a, wherein the bispecific antibody is administered at a dose of 24 mg.

[0447] 3a. The bispecific antibody used in Embodiment 1a, wherein the bispecific antibody is administered at a dose of 48 mg.

[0448] 4a. The bispecific antibody used in any one of Embodiments 1a - 3a, wherein the bispecific antibody is administered once a week (administered once a week).

[0449] 5a. The bispecific antibody used in Embodiment 4a, wherein the once-a-week administration is carried out for 2.5 28-day cycles.

[0450] 6a. The bispecific antibody used in Embodiment 4a or 5a, wherein after the once-a-week administration, the bispecific antibody is administered once every two weeks (administered once every two weeks).

[0451] 7a. The bispecific antibody used in Embodiment 6a, wherein the once-every-two-weeks administration is carried out for six 28-day cycles.

[0452] 8a. The bispecific antibody used in Embodiment 6a or 7a, wherein after the once-every-two-weeks administration, the bispecific antibody is administered once every four weeks.

[0453] 9a. The bispecific antibody used in any one of Embodiments 4a - 8a, wherein a sensitizing dose of the bispecific antibody is administered in the 1st cycle of the 28-day cycle before administering the first once-a-week dose of 12 - 60 mg.

[0454] 10a. The bispecific antibody used in Embodiment 9a, wherein the sensitizing dose is administered two weeks before administering the first once-a-week dose of 12 - 60 mg.

[0455] 11a. The bispecific antibody used in Embodiment 9a or 10a, wherein the sensitizing dose is in the range of 0.05 - 0.35 mg.

[0456] 12a. The bispecific antibody used in any one of Embodiments 9a - 11a, wherein the sensitizing dose is 0.16 mg or about 0.16 mg.

[0457] 13a. A bispecific antibody used in any one of embodiments 9a - 12a, wherein an intermediate dose of the bispecific antibody is administered after the administration of the sensitizing dose and before the administration of the first once - weekly dose of 12 - 60 mg.

[0458] 14a. A bispecific antibody for the use as in embodiment 13a, wherein the sensitizing dose is administered on day 1, and the intermediate dose is administered on day 8, before the first once - weekly dose of 12 - 60 mg on days 15 and 22 of cycle 1.

[0459] 15a. A bispecific antibody used in embodiment 13a or 14a, wherein the intermediate dose is in the range of 0.6 - 1.2 mg.

[0460] 16a. A bispecific antibody used in any one of embodiments 13a - 15a, wherein the intermediate dose is 0.8 mg or about 0.8 mg.

[0461] 17a. A bispecific antibody used in any one of embodiments 13a - 16a, wherein the bispecific antibody is administered in a 28 - day cycle, wherein:

[0462] a) In cycle 1, the sensitizing dose is administered on day 1, the intermediate dose is administered on day 8, and the full dose of 12 - 60 mg is administered on days 15 and 22;

[0463] b) In cycles 2 - 3, the full dose of 12 - 60 mg is administered on days 1, 8, 15, and 22;

[0464] c) In cycles 4 - 9, the full dose of 12 - 60 mg is administered on days 1 and 15; and

[0465] d) In cycle 10 and subsequent cycles, the full dose of 12 - 60 mg is administered on day 1.

[0466] 18a. A bispecific antibody used in embodiment 17a, wherein the full dose is 24 mg or about 24 mg.

[0467] 19a. A bispecific antibody used in embodiment 17a, wherein the full dose is 48 mg or about 48 mg.

[0468] 20a. A bispecific antibody used in any one of embodiments 1a - 19a, wherein the bispecific antibody is administered subcutaneously.

[0469] The bispecific antibody used in any one of embodiments 1a-20a, wherein the subject has a clinical history of CLL / SLL that has transformed into aggressive lymphoma; for example, having a DLBCL subtype.

[0470] 22a. The bispecific antibody used in any one of embodiments 1a-21a, wherein the Richter's syndrome has a DLBCL subtype.

[0471] 23a. The bispecific antibody used in any one of embodiments 1a-21a, wherein the subject has received one or more (such as at least two) previous lines of therapy for chronic lymphocytic leukemia (CLL) and / or for small lymphocytic lymphoma (SLL).

[0472] 24a. The bispecific antibody used in any one of the embodiments, wherein the previous line of therapy for CLL and / or SLL includes chemoimmunotherapy.

[0473] 25a. The bispecific antibody used in any one of the embodiments, wherein the previous line of therapy for CLL and / or SLL includes therapy using targeting agents such as BCL2 inhibitors or BTK inhibitors.

[0474] 26a. The bispecific antibody used in any one of the embodiments, wherein the previous line of therapy for CLL and / or SLL includes CAR T cell therapy.

[0475] 27a. The bispecific antibody used in any one of embodiments 1a-23a, wherein the subject has received previous therapy for Richter's syndrome, such as previous therapy selected from:

[0476] i) Rituximab combined with cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP),

[0477] ii) Rituximab combined with dexamethasone, cytarabine, and cisplatin (R-DHAP), and

[0478] iii) Venetoclax combined with rituximab, etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin (VR-EPOCH).

[0479] 28a. The bispecific antibody used in any one of embodiments 1a-27a, wherein the subject achieves a complete metabolic response or a partial metabolic response.

[0480] 29a. The bispecific antibody used in any one of embodiments 1a-27a, wherein the subject achieves a complete response, a partial response, or disease stabilization.

[0481] 30a. The bispecific antibody used in any one of embodiments 1a - 26a, wherein the subject receives acalabrutinib as a first - line therapy for Richter's syndrome.

[0482] 31a. The bispecific antibody used in any one of embodiments 1a - 30a, wherein the subject achieves a complete metabolic response or a partial metabolic response.

[0483] 32a. The bispecific antibody used in any one of embodiments 1a - 30a, wherein the subject achieves a complete response, a partial response, or disease stabilization.

[0484] 33a. The bispecific antibody used in any one of embodiments 1a - 26a and 30a - 32a, the method being a first - line therapy for Richter's syndrome.

[0485] 34a. The bispecific antibody used in any one of embodiments 21a - 29a, wherein the subject has refractory and / or relapsed Richter's syndrome after receiving the previous therapy.

[0486] 35a. The bispecific antibody used in any one of embodiments 1a - 34a, wherein the subject is treated with prophylaxis against cytokine release syndrome (CRS).

[0487] 36a. The bispecific antibody used in embodiment 35a, wherein the prophylaxis includes administering corticosteroids to the subject.

[0488] 37a. The bispecific antibody used in embodiment 35a or 36a, wherein the corticosteroid is administered on the same day as the bispecific antibody.

[0489] 38a. The bispecific antibody used in embodiment 37a, wherein the corticosteroid is further administered on the second, third, and fourth days after administering the bispecific antibody.

[0490] 39a. The bispecific antibody used in any one of embodiments 36a - 38a, wherein the corticosteroid is prednisolone.

[0491] 40a. The bispecific antibody used in embodiment 39a, wherein the prednisolone is administered at an intravenous dose of 100 mg or its equivalent including an oral dose.

[0492] 41a. The bispecific antibody used in any one of embodiments 1a - 40a, wherein a pre - medication is administered to the subject to reduce the reaction to the injection.

[0493] 42a. The bispecific antibody used in embodiment 41a, wherein the pre - medication includes an antihistamine.

[0494] 43a. The bispecific antibody used in embodiment 42a, wherein the antihistamine is diphenhydramine.

[0495] 44a. The bispecific antibody used in embodiment 43a, wherein the diphenhydramine is administered at an intravenous or oral dose of 50 mg or its equivalent.

[0496] 45a. The bispecific antibody used in any one of embodiments 41a - 44a, wherein the premedication includes an antipyretic.

[0497] 46a. The bispecific antibody used in embodiment 45a, wherein the antipyretic is acetaminophen.

[0498] 47a. The bispecific antibody used in embodiment 46a, wherein the acetaminophen is administered at an oral dose of 560 to 1000 mg or its equivalent.

[0499] 48a. The bispecific antibody used in any one of embodiments 41a - 47a, wherein the premedication is administered on the same day as the bispecific antibody.

[0500] 49a. The bispecific antibody used in any one of embodiments 35a - 48a, wherein the prophylaxis is administered during cycle 1.

[0501] 50a. The bispecific antibody used in any one of embodiments 41a - 49a, wherein the premedication is administered during cycle 1.

[0502] 51a. The bispecific antibody used in embodiment 49a or 50a, wherein when the subject experiences CRS greater than grade 1 after the last administration of the bispecific antibody in cycle 1, the prophylaxis is administered during cycle 2.

[0503] 52a. The bispecific antibody used in embodiment 51a, wherein when the subject experiences CRS greater than grade 1 in the last administration of the bispecific antibody in a previous cycle, the prophylaxis is continued in subsequent cycles.

[0504] 53a. The bispecific antibody used in any one of embodiments 41a - 52a, wherein the premedication is administered during cycle 2.

[0505] 54a. The bispecific antibody used in embodiment 53a, wherein the premedication is administered during subsequent cycles.

[0506] The bispecific antibody used in any one of embodiments 1a - 54a, wherein if the subject develops grade 1 CRS, an antibiotic is administered to the subject.

[0507] 56a. The bispecific antibody used in any one of embodiments 1a - 54a, wherein if the subject develops grade 2 or 3 CRS, a vasopressor is administered to the subject.

[0508] 57a. The bispecific antibody used in any one of embodiments 1a - 54a, wherein if the subject develops grade 4 CRS, at least two vasopressors are administered to the subject.

[0509] 58a. The bispecific antibody used in any one of embodiments 1a - 57a, wherein if the subject develops grade 2, 3, or 4 CRS, tocilizumab is administered to the subject.

[0510] 59a. The bispecific antibody used in embodiment 58a, wherein a steroid is further administered to the subject.

[0511] 60a. The bispecific antibody used in embodiment 59a, wherein the steroid is dexamethasone.

[0512] 61a. The bispecific antibody used in embodiment 59a, wherein the steroid is methylprednisolone.

[0513] 62a. The bispecific antibody used in any one of embodiments 58a - 61a, wherein if the subject is refractory to tocilizumab, tocilizumab is switched to an anti - IL - 6 antibody (e.g., siltuximab).

[0514] 63a. The bispecific antibody used in any one of embodiments 58a - 61a, wherein if the subject is refractory to tocilizumab, tocilizumab is switched to an IL - 1R antagonist (e.g., anakinra).

[0515] 64a. The bispecific antibody used in any one of embodiments 1a - 63a, wherein the subject is treated with prophylaxis against tumor lysis syndrome (TLS).

[0516] 65a. The bispecific antibody used in embodiment 64a, wherein the prophylaxis against TLS includes administering one or more urate - lowering agents before administering the bispecific antibody.

[0517] 66a. The bispecific antibody used in embodiment 65a, wherein the one or more urate - lowering agents include rasburicase and / or allopurinol.

[0518] The bispecific antibody used in any one of embodiments 1a - 66a, wherein the subject achieves a complete response, a partial response, or disease stabilization.

[0519] 68a. The bispecific antibody used in any one of embodiments 1a - 67a, wherein:

[0520] (i) The first antigen - binding region comprises VHCDR1, VHCDR2, and VHCDR3, and VLCDR1, VLCDR2, and VLCDR3, wherein VHCDR1, VHCDR2, and VHCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 1, 2, and 3, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 4, the sequence GTN, and SEQ ID NO: 5, respectively; and

[0521] (ii) The second antigen - binding region comprises VHCDR1, VHCDR2, and VHCDR3, and VLCDR1, VLCDR2, and VLCDR3, wherein VHCDR1, VHCDR2, and VHCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 8, 9, and 10, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 11, the sequence DAS, and SEQ ID NO: 12, respectively.

[0522] 69a. The bispecific antibody used in any one of embodiments 1a - 68a, wherein:

[0523] (i) The first antigen - binding region comprises a VH region comprising the amino acid sequence of SEQ ID NO: 6 and a VL region comprising the amino acid sequence of SEQ ID NO: 7; and

[0524] (ii) The second antigen - binding region comprises a VH region comprising the amino acid sequence of SEQ ID NO: 13 and a VL region comprising the amino acid sequence of SEQ ID NO: 14.

[0525] 70a. The bispecific antibody used in any one of embodiments 1a - 68a, wherein the first binding arm of the bispecific antibody is derived from a humanized antibody, preferably derived from a full - length IgG1, λ (lambda) antibody.

[0526] 71a. The bispecific antibody used in embodiment 70a, wherein the first binding arm of the bispecific antibody comprises a λ light - chain constant region, and the λ light - chain constant region comprises the amino acid sequence set forth in SEQ ID NO: 22.

[0527] 72a. A bispecific antibody used in any one of embodiments 1a - 71a, wherein the second binding arm of the bispecific antibody is derived from a human antibody, preferably derived from a full - length IgG1,κ (kappa) antibody.

[0528] 73a. The bispecific antibody used in embodiment 72a, wherein the second binding arm comprises a κ light chain constant region, and the κ light chain constant region comprises the amino acid sequence set forth in SEQ ID NO: 23.

[0529] 74a. The bispecific antibody used in any one of embodiments 1a - 73a, wherein the bispecific antibody is a full - length antibody having a human IgG1 constant region.

[0530] 75a. The bispecific antibody used in any one of embodiments 1a - 74a, wherein the bispecific antibody comprises an inert Fc region.

[0531] 76a. The bispecific antibody used in any one of embodiments 1a - 75a, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, and in both the first heavy chain and the second heavy chain, the amino acids at positions corresponding to positions L234, L235, and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E, and A, respectively.

[0532] 77a. The bispecific antibody used in any one of embodiments 1a - 76a, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein in the first heavy chain, the amino acid at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and in the second heavy chain, the amino acid at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa.

[0533] 78a. The bispecific antibody used in any one of embodiments 1a - 77a, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein

[0534] (i) in both the first heavy chain and the second heavy chain, the amino acids at positions corresponding to positions L234, L235, and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E, and A, respectively, and

[0535] (ii) In the first heavy chain, the amino acid at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and in the second heavy chain, the amino acid at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa.

[0536] 79a. The bispecific antibody used in embodiment 78a, wherein the bispecific antibody comprises a heavy chain constant region comprising the amino acid sequences of SEQ ID NOs: 19 and 20.

[0537] 80a. The bispecific antibody used in any one of embodiments 1a - 79a, wherein the bispecific antibody comprises heavy and light chains comprising the amino acid sequences listed in SEQ ID NOs: 24 and 25, respectively, and heavy and light chains comprising the amino acid sequences listed in SEQ ID NOs: 26 and 27, respectively.

[0538] 81a. The bispecific antibody used in any one of embodiments 1a - 80a, wherein the bispecific antibody comprises heavy and light chains consisting of the amino acid sequences of SEQ ID NOs: 24 and 25, respectively, and heavy and light chains consisting of the amino acid sequences of SEQ ID NOs: 26 and 27, respectively.

[0539] 82a. The bispecific antibody used in any one of embodiments 1a - 81a, wherein the bispecific antibody is acelretumab or a biosimilar thereof.

[0540] The present disclosure is further illustrated by the following examples, which should not be construed as further limitations. The content of all the figures and all references, Genbank sequences, patents, and published patent applications cited throughout this application are hereby expressly incorporated by reference. Examples

[0541] DuoBody - CD3xCD20

[0542] DuoBody - CD3xCD20 is a bsAb that recognizes the T - cell antigen CD3 and the B - cell antigen CD20. DuoBody - CD3xCD20 elicits potent T - cell - mediated killing of CD20 - expressing cells. DuoBody - CD3xCD20 has a regular IgG1 structure.

[0543] Two parental antibodies, IgG1-CD3-FEAL (humanized IgG1λ, CD3ε-specific antibody with heavy and light chain sequences as set forth in SEQ ID NO: 24 and 25, respectively) and IgG1-CD20-FEAR (derived from the human IgG1κ CD20-specific antibody 7D8 with heavy and light chain sequences as set forth in SEQ ID NO: 26 and 27, respectively) were prepared as separate biological intermediates. Each parental antibody contains one of the complementary mutations in the CH3 domain required to generate the DuoBody molecule (F405L and K409R, respectively). The parental antibodies contain three additional mutations in the Fc region (L234F, L235E, and D265A; FEA). The parental antibodies were produced in the mammalian Chinese hamster ovary (CHO) cell line using standard suspension cell culture and purification techniques. Subsequently, DuoBody-CD3xCD20 was prepared by the controlled Fab-arm exchange (cFAE) process (Labrijn et al. 2013, Labrijn et al. 2014, Gramer et al. 2013). The parental antibodies were mixed and subjected to controlled reducing conditions. This resulted in the separation of the parental antibodies that re-assembled upon re-oxidation. In this way, a highly pure DuoBody-CD3xCD20 preparation (approx. 93% - 95%) was obtained. After further polishing / purification, a final product approaching 100% purity was obtained. The concentration of DuoBody-CD3xCD20 was measured by absorbance at 280 nm using the theoretical extinction coefficient Ɛ = 1.597 mL∙mg -1 cm -1 , and the DuoBody-CD3xCD20 concentration was measured by absorbance at 280 nm. This product has received the international proprietary name of acriviroc.

[0544] Acriviroc is prepared as a sterile, clear, colorless to pale yellow solution (5 mg / mL or 60 mg / mL) and is provided as a concentrate for subcutaneous (SC) injection solution. Acriviroc contains buffers and tonicifying agents. All excipients and their amounts in the formulated product are pharmaceutically acceptable for a subcutaneous injection product. A suitable dose is reconstituted to a volume of approximately 1 mL for subcutaneous injection.

[0545] Example 1: A Phase 1b / 2, open-label, safety, and efficacy study of acriviroc in relapsed / refractory chronic lymphocytic leukemia and Richter's syndrome (RS)

[0546] The objective of this Phase 1b / 2 study was to evaluate the safety and preliminary efficacy of single-agent acralizumab in subjects with Richter's syndrome (RS). The study was an open-label, 2-part (dose escalation and expansion), multicenter study designed to assess the safety, tolerability, PK, pharmacodynamics, immunogenicity, and preliminary efficacy of single-agent acralizumab in subjects 18 years of age or older with relapsed and / or refractory (R / R) chronic lymphocytic leukemia (CLL) or Richter's syndrome.

[0547] This trial consisted of 2 parts: dose escalation (Part 1) and expansion (Part 2). The overall study design is further disclosed in WO 2021 / 224499. This disclosure provides the objectives of the dose escalation portion, including identification of the recommended Phase 2 dose (RP2D) and the maximum tolerated dose (MTD). Acralizumab was studied at 2 full dose levels: 24 mg and 48 mg. An escalating dosing regimen was applied: 0.16 mg / 0.8 mg / 24 mg and 0.16 mg / 0.8 mg / 48 mg (sensitization / intermediate / full dose).

[0548] An expansion cohort (Expansion Cohort 1) investigating the treatment of relapsed or refractory chronic lymphocytic leukemia (R / R CLL) is also disclosed in WO 2021 / 224499.

[0549] Finally, the disclosure in WO 2021 / 224499 includes preliminary results from the dose escalation phase data, which indicated that acralizumab was well tolerated in patients with R / R CLL at dose levels up to 48 mg and had encouraging clinical activity in patients with high-risk disease.

[0550] Target

[0551] Expansion Cohort 2, Richter's syndrome

[0552] The primary objective of this arm of Expansion Cohort 2 was to evaluate the preliminary efficacy of acralizumab in subjects with Richter's syndrome (endpoint: ORR).

[0553] The secondary objectives of Expansion Cohort 2 include evaluating the preliminary efficacy of acralizumab (endpoints: overall response rate (ORR), duration of response (DOR), complete response (CR), time to response (TTR), progression-free survival (PFS), overall survival (OS), and time to next anti-cancer therapy (TTNT)), assessing the MRD status in peripheral blood and bone marrow (endpoint: incidence of undetectable MRD), evaluating the safety and tolerability of acralizumab (endpoints: incidence and severity of adverse events (AE), serious adverse events (SAE), cytokine release syndrome (CRS), immune effector cell-associated neurotoxicity syndrome (ICAN), and tumor lysis syndrome (TLS), as well as the incidence of dose interruption, dose delay, and dose intensity), establishing the pharmacokinetic (PK) and pharmacodynamic characteristics of acralizumab (endpoints: PK parameters and pharmacodynamic parameters), and evaluating the immunogenicity of acralizumab (endpoint: incidence of anti-drug antibodies (ADA) to acralizumab).

[0554] The exploratory objectives of the expansion part include evaluating biomarkers predictive of clinical response to acralizumab (endpoints: expression of CD20 and evaluation of immune populations, phenotypes, functions, and blood).

[0555] Overview of study design

[0556] Expansion Cohort 2, Richter's syndrome

[0557] Approximately 70 subjects with Richter's syndrome (RS) were enrolled in the expansion part.

[0558] As shown below, acralizumab is administered as a subcutaneous (SC) injection in 4-week cycles (i.e., 28 days) until one or more discontinuation criteria are met:

[0559] · Cycles 1 - 3: Days 1, 8, 15, and 22 (QW)

[0560] · Cycles 4 - 9: Days 1 and 15 (Q2W)

[0561] · Cycles 10 and later: Day 1 (Q4W)

[0562] An escalating dosing method is used to mitigate the likelihood of CRS: a priming dose of 0.16 mg on Day 1 of Cycle 1, then an intermediate dose of 0.8 mg on Day 8 of Cycle 1, then full doses of 48 mg on Days 15 and 22 of Cycle 1, and full doses of 48 mg in subsequent cycles.

[0563] The primary efficacy endpoint of the expansion cohort was the ORR as evaluated using the iwCLL 2018 criteria (Table 2). Secondary efficacy endpoints included DOR, CR, TTR, PFS, OS, and TTNT. The incidence of MRD negativity and undetectable MRD were also evaluated as secondary efficacy endpoints. MRD assessment indicated how many cancer cells remained in subjects who were in remission during or after treatment had been administered. Safety endpoints in the expansion cohort included the incidence and severity of AE / SAE, the incidence and severity of tumor lysis syndrome (TLS), immune effector cell-associated neurotoxicity syndrome (ICANS), and CRS, and the incidence of treatment interruptions and delays.

[0564] Inclusion criteria

[0565] 1. Prior to any screening procedures, subjects must sign the ICF indicating that he or she understands the purpose of the trial and the procedures required and is willing to participate in the trial prior to any other trial-related assessments or procedures. In cases where local or national specific regulations require it, if each subject consents to provide samples for genomic biomarker analysis (DNA), he or she must sign a separate ICF. If a subject refuses consent for DNA research in these specific areas, the subject remains eligible to participate in the trial.

[0566] 2. Subjects must be at least 18 years of age.

[0567] 3. Must have a clinical history of CLL / SLL with biopsy-proven transformation to aggressive lymphoma (i.e., DLBCL subtype).

[0568] 4. Considered ineligible for chemoimmunotherapy or refused intensive chemotherapy according to the investigator's judgment.

[0569] 5. Must have measurable disease as determined by both

[0570] a. Fluorodeoxyglucose (FDG)-positron emission tomography (PET) CT scan that shows positive lesions compatible with the anatomic tumor sites determined by CT (or MRI); and

[0571] b) CT scan (or MRI) with ≥2 well-defined lesions / nodules with long axis > 1.5 cm and short axis > 1.0 cm or 1 well-defined lesion / nodule with long axis > 2.0 cm and short axis ≥ 1.0 cm.

[0572] 6. ECOG performance status score of 0, 1, or 2

[0573] 7. Evidence of CD20 positivity at screening

[0574] 8. Have the following acceptable laboratory parameters:

[0575] Table 4

[0576]

[0577] 8. During the 2-week period prior to the first dose of acrivirocizumab, the cumulative dose of corticosteroids received was less than 250 mg of prednisone equivalent.

[0578] 9. Subjects must have available fresh bone marrow material at screening.

[0579] 10. Women of reproductive potential must agree to use adequate contraception during the trial and for 12 months after the last administration of acrivirocizumab. Adequate contraception is defined as highly effective contraceptive methods.

[0580] 11. Women of childbearing potential must have a negative serum (β-HCG) pregnancy test at screening and a negative serum or urine pregnancy test before treatment administration on day 1 of each cycle.

[0581] 12. Women must agree not to donate eggs (oocytes, ova) for assisted reproductive purposes throughout the trial until 12 months after the last treatment.

[0582] 13. Men who have sexual intercourse with women of reproductive potential and have not had a vasectomy must agree to use barrier contraceptive methods, such as condoms with spermicidal foam / gel / membrane / cream / suppository and a diaphragm or cervical / vault cap with spermicidal foam / gel / membrane / cream / suppository, and all men must also not donate sperm during the trial and for 12 months after receiving the last dose of acrivirocizumab.

[0583] 14. Subjects must be willing and able to comply with the prohibitions and restrictions specified in this protocol.

[0584] Exclusion criteria

[0585] 1. Diagnosed with Richter's syndrome without the DLBCL subtype, such as Hodgkin's lymphoma, prolymphocytic leukemia.

[0586] 2. The subject received autologous HSCT within 3 months prior to the first dose of acrivirocizumab.

[0587] 3. The subject has received more than 1 prior line of therapy for RS.

[0588] 4. The subject has received prior treatment with a CD3 × CD20 bispecific antibody

[0589] 5. The subject has received any prior allogeneic HSCT or solid organ transplant

[0590] 6. The subject has received treatment with an anti-cancer agent, such as

[0591] a. Small molecules, such as a BTK inhibitor, BCL2 inhibitor, or PI3K inhibitor within 5 half-lives prior to the first dose of acralizumab; or

[0592] b. An anti-CD20 mAb or chemotherapy within 2 weeks prior to the first dose of acralizumab; or

[0593] c. A radio-conjugated or toxin-conjugated antibody or CAR-T cell therapy within 4 weeks or 5 half-lives (whichever is shorter) prior to the first dose of acralizumab

[0594] d. The subject has received treatment with an investigational drug within 4 weeks or 5 half-lives (whichever is shorter) prior to the first dose of acralizumab.

[0595] 7. The subject has an autoimmune disease or other disease requiring permanent or high-dose immunosuppressive therapy

[0596] 8. The subject has clinically significant heart disease, including but not limited to:

[0597] a. An unstable or uncontrolled disease / condition related to or affecting cardiac function, such as unstable angina, congestive heart failure of New York Heart Association class III or IV (see Appendix 3), arrhythmia (CTCAE v5.0 grade 2 or higher), or clinically significant electrocardiogram (ECG) abnormalities

[0598] b. Myocardial infarction, intracranial hemorrhage, or stroke within the past 6 months

[0599] c. Screening 12-lead ECG shows a corrected baseline QT interval (QTcF) >480 by the Fridericia formula

[0600] 9. The subject has received a live vaccine within 28 days prior to the first dose of acralizumab

[0601] 10. The subject has toxicity from a prior anti-cancer therapy that has not resolved to baseline levels or is grade 1 or lower, excluding alopecia and peripheral neuropathy

[0602] 11. The subject has known CNS involvement at screening

[0603] 12. The subject has a past or current known malignancy different from the inclusion diagnosis, except for the following:

[0604] a. Cervical cancer at stage 1B or lower

[0605] b. Non-invasive basal cell or squamous cell skin cancer

[0606] c. Non-invasive superficial bladder cancer

[0607] d. Prostate cancer with current PSA level < 0.1 ng / mL

[0608] e. Any curable cancer with a CR duration > 2 years

[0609] 13. The subject has a suspected allergic reaction, hypersensitivity or intolerance to acrivastamab or its excipients

[0610] 14. The subject has undergone major surgery within 4 weeks prior to enrollment

[0611] 15. The subject has a known history of hepatitis B / seropositive (unless due to vaccination or immunity from resolved natural infection, or unless due to passive immunity from immunoglobulin therapy):

[0612] a. Positive test for antibody to hepatitis B core antigen (anti-HBc), and

[0613] b. Negative test for antibody to hepatitis B surface antigen (anti-HBs).

[0614] 16. Known history or ongoing hepatitis C infection that has not been cured

[0615] 17. Known history of seropositive HIV infection. Note: HIV testing at screening is required only when mandated by local health authorities or institutional standards

[0616] 18. The subject is pregnant or breastfeeding, or is a female planning to become pregnant during the period of enrollment in this trial or within 12 months after the last dose of acrivastamab

[0617] 19. The subject is a male planning to become a father during the period of enrollment in this trial or within 12 months after the last dose of acrivastamab

[0618] 20. The subject has any condition that is not in the best interest of the subject (e.g., compromising well-being) or that may prevent, limit or confound the protocol-specified assessments

[0619] 21. The subject has uncontrolled concurrent illnesses at the time of enrollment or within 2 weeks prior to the first dose of acrivastamab, such as a persistent or active infection requiring intravenous antibiotic treatment

[0620] Pre-medication and CRS prophylaxis

[0621] Premedication with corticosteroids, antihistamines, and antipyretics is mandatory as described in Table 5. For each dose of acralizumab in Cycle 1, corticosteroids for 4 consecutive days are mandatory to prevent / reduce the severity from potential CRS symptoms as described in Table 5. For the administration of acralizumab in Cycle 2 and later, CRS prophylaxis with corticosteroids is optional. Corticosteroids can be administered by the intravenous or oral route using the recommended dose or equivalent.

[0622] Table 5. Prophylactic corticosteroid administration before or after acralizumab administration

[0623]

[0624]

[0625]

[0626] Table 6: Corticosteroid dose equivalent - conversion table

[0627]

[0628] Supportive care for cytokine release syndrome

[0629] CRS is graded according to the ASTCT grading of CRS (Tables 7 and 8), and for the treatment of CRS, subjects should receive supportive care. Supportive care can include, but is not limited to,

[0630] • Saline infusion

[0631] • Systemic glucocorticoids, antihistamines, antipyretics

[0632] • Blood pressure support (vasopressin, vasopressor)

[0633] • Support for low - flow and high - flow oxygen and positive pressure ventilation

[0634] • IV administration of monoclonal antibodies against IL - 6R, such as tocilizumab

[0635] • If there is no response to repeated tocilizumab, monoclonal antibodies against IL - 6, such as IV siltuximab.

[0636] • Administration of anakinra

[0637] • Blood product support, analgesics, skin and oral care, etc. should be based on local guidelines and the decision of the investigator.

[0638] Table 7: Grading and Management of Cytokine Release Syndrome

[0639] According to the American Society for Transplantation and Cellular Therapy (ASTCT), formerly the American Society for Blood and Marrow Transplantation (ASBMT), the unified definition and grading criteria for CRS are presented as follows.

[0640] Grading of Cytokine Release Syndrome

[0641]

[0642] Table 8: Grading and Management of Cytokine Release Syndrome

[0643]

[0644]

[0645] Prevention and management of tumor lysis syndrome

[0646] For prophylactic treatment of tumor lysis syndrome, subjects receive urate-lowering agents before administration of acralizumab, allopurinol is given at least 72 hours before the first dose of acralizumab, and rasburicase is initiated before starting acralizumab. Increased oral hydration should be received before the first dose and maintained during dosing. The TLS risk category of the subject is re-evaluated before subsequent doses.

[0647] Study assessments

[0648] Bone marrow assessment

[0649] Fresh bone marrow aspirates are obtained at screening (i.e., within 21 days before Cycle 1 Day 1) and at complete response (CR) or when there are clinical indications. Fresh bone marrow biopsies are obtained at screening and at CR or nodular partial response (PR) (nPR) or when there are clinical indications. Bone marrow evaluation includes morphological examination as well as flow cytometry or immunohistochemistry.

[0650] Radiological assessment

[0651] For RS, it must be performed at screening (i.e., within 3 weeks before the first dose of GEN3013) 18F-FDG-PET CT (or CT / MRI and FDG-PET when PET CT is not available). For subjects with FDG-avid tumors at screening, all subsequent disease assessments will be performed using FDG-PET CT with a 5-point scale (Barrington et al., 2014). For subjects with non-avid or variably FDG-avid tumors, CT scans with IV contrast will be performed on the neck / thorax / abdomen / pelvis / other known lesions. The CT component of PET CT can be used to replace an independent CT / MRI only if the CT component of PET CT has similar diagnostic quality to contrast-enhanced CT performed without PET. If contrast-enhanced PET CT is not available, an independent diagnostic CT / MRI and standard FDG-PET should be performed. If separate CT and PET scanners are used and the subject undergoes both scans on the same day, PET must be performed before CT and IV contrast to avoid compromising the PET results. For any given subject, the PET CT acquisition method (e.g., administration of intravenous contrast) should be consistent between screening and subsequent evaluations.

[0652] The imaging assessment schedule for both dose escalation and expansion is performed as described in the visit assessment schedule (section 1). CT scans with contrast are the recommended imaging modality. MRI can be used only if contrast CT is medically contraindicated or the CT scan frequency exceeds local standards.

[0653] MRI can be used to evaluate disease sites that cannot be adequately imaged with CT (MRI must be obtained at screening and all subsequent response evaluations when MRI is desired). For all other disease sites, MRI studies cannot replace the required neck, thorax, abdomen, and pelvis CT scans.

[0654] If necessary, additional imaging assessments can be performed at any time during the trial, at the investigator's discretion, to support the efficacy evaluation of the subject. Any clinical suspicion of disease progression at any time requires immediate physical examination and imaging assessment, rather than waiting for the next pre-scheduled imaging assessment.

[0655] Minimal residual disease (MRD) assessment

[0656] MRD in the blood is evaluated by flow cytometry and next-generation sequencing. Blood samples are requested at fixed time points and at CR after treatment initiation. As an exploratory analysis, when the subject reaches CR, a portion of the aspirate collected to confirm CR is used to evaluate MRD.

[0657] Disease response and disease progression assessment

[0658] Perform tumor response according to imaging assessment to inform decisions on continuous therapy. Response assessment is completed according to Table 2 of the Lugano criteria (Cheson et al., 2014, J Clin Oncol 32, 3059-3068). Since local palliative radiotherapy is permitted for non-target lesions, if given during the trial, these lesions should no longer be included in the response assessment.

[0659] Lugano criteria (Cheson et al., 2014) Target and non-target lesions

[0660] Target lesions should consist of up to six of the largest dominant nodules, nodular masses, or other lymphoma lesions that can be measured in two dimensions and should preferably be from different body regions representing the overall disease burden of the subject, including mediastinal disease and retroperitoneal disease (if applicable). At baseline, the longest diameter (LDi) of measurable nodules must be greater than 15 mm. Measurable extranodal disease may be included among the six representative target lesions. At baseline, measurable extranodal lesions should be greater than 10 mm in LDi. All other lesions (including nodular disease, extranodal disease, and evaluable disease) should be followed up as non-target lesions (e.g., skin, GI, bone, spleen, liver, kidney, pleural effusion or pericardial effusion, ascites, bone, bone marrow).

[0661] Dissociated and confluent lesions

[0662] Over time, lesions may dissociate or may become confluent. In the case of dissociated lesions, the individual products of the perpendicular diameters (PPDs) of these nodules should be added together to represent the PPD of the dissociated lesions; this PPD is added to the sum of the PPDs of the remaining lesions to measure response. If there is subsequent growth of any or all of these discrete nodules, the nadir of each individual nodule is used to determine progression. In the case of confluent lesions, the PPD of the confluent mass is compared to the sum of the PPDs of the individual nodules, where an increase in the PPD of the confluent mass of more than 50% compared to the sum of the individual nodules is required to indicate progressive disease (PD). The LDi and shortest diameter (SDi) are no longer required to determine progression.

[0663] Endpoints are defined as follows:

[0664] Overall response rate (ORR), defined as the proportion of subjects who achieve a PR or CR response before the start of subsequent therapy.

[0665] Time to response (TTR), defined in responders as the time between the first dose of acoramidis and the initial documentation of PR or CR.

[0666] Duration of response (DOR), defined in responders as the time between the initial record of PR or CR and the date of disease progression or death (whichever occurs first).

[0667] Progression-free survival (PFS), defined as the time between the date of the first administration of acrilumab and the date of disease progression or death (whichever occurs first).

[0668] Overall survival (OS), defined as the time between the date of the first administration of acrilumab and the date of death.

[0669] MRD negativity rate, defined as the proportion of subjects with at least one undetectable MRD result according to a specific threshold before the start of subsequent therapy.

[0670] Clinical safety assessment

[0671] Safety will be evaluated by measuring adverse events, laboratory test results, ECG, vital signs measurements, physical examination findings, and ECOG performance status. Immunoeffector cell-related neurotoxicity syndromes (e.g., as described by Lee et al., Biol Blood Marrow Transplant 2019;25:625-638), constitutional symptoms (B symptoms), tumor flare reactions, and survival will also be evaluated.

[0672] Immunophenotyping

[0673] Absolute B-cell and T-cell counts in fresh whole blood will be determined by flow cytometry to monitor changes associated with acrilumab treatment. T-cell activation and exhaustion phenotypes will be evaluated using flow cytometry and markers to assess the association of such markers with drug target engagement, treatment efficacy, and / or the safety of acrilumab. Other immune phenotypes of circulating immune cells in whole blood (e.g., the level of regulatory T cells that can suppress T-cell function) will be determined by flow cytometry to evaluate the association of such markers with T-cell activation / exhaustion phenotypes, subject response, and the MOA of acrilumab.

[0674] Analysis of cytokines and endothelial cell activation markers

[0675] Since T-cell activation after the initial administration of acrilumab can lead to cytokine release causing CRS, cytokine levels will be closely monitored. Levels of cytokines (such as IL-2, IL15, IL-6, IL-8, IL-10, IFNγ, and / or TNFα) will be measured in plasma samples using an array-based ligand-binding assay. Other cytokines may also be assayed to evaluate the association of such markers with AEs occurring after treatment and with the outcome of acrilumab.

[0676] Preliminary results:

[0677] The first patient was enrolled on November 17, 2021.

[0678] Data cutoff was July 15, 2022:

[0679] Ten Richter's syndrome (RS) patients (median age 69.5 years; range 53 - 79 years) had received acralizumab 48 mg, with a follow-up of ≥12 weeks. The median time from RS diagnosis to the first dose of acralizumab was 0.03 years (range: 0.0 - 0.1). Prior therapies for RS included rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP), rituximab, dexamethasone, cytarabine, and cisplatin (R-DHAP), and venetoclax plus dose-adjusted rituximab, etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin (VR-EPOCH), and 50% of patients received acralizumab as first-line treatment for RS. The median treatment duration was 2.5 months (range: 0.5 - 6.5), and 5 (50%) patients received ongoing treatment. The most common treatment-emergent adverse events (TEAEs) of any grade were cytokine release syndrome (CRS) (90%; 30% grade 1, 60% grade 2), anemia (30%), diarrhea (40%), hypophosphatemia (10%), injection site reaction (30%), and thrombocytopenia (30%). Notable grade 3 - 4 TEAEs included neutropenia (n = 4; 2 patients grade 3, 2 patients grade 4), anemia (n = 2), and COVID-19 (n = 2). Most CRS events were associated with the first full dose of acralizumab. All CRS events resolved (median resolution time, 3 days), no patients discontinued treatment due to CRS, and 7 patients received tocilizumab. No cases of immune effector cell-associated neurotoxicity (ICANS) were observed. One patient developed clinical tumor lysis syndrome (grade 2), which resolved within 3 days. No patients discontinued treatment due to AEs. Two patients died due to disease progression. Antitumor activity was observed early (most responses were seen at the first [week 6] assessment), with an overall response rate of 60% and a complete response rate of 50%.

[0680] Data cutoff was September 8, 2022 (efficacy) / September 16, 2022 (safety):

[0681] A total of 10 patients in the Richter's cohort had been dosed with 48 mg acralizumab and were evaluable for response. The median treatment duration was 3.5 months (range: 0.5 - 9.3). The median number of 28-day cycles of acralizumab was 4 (range 1 - 11). The primary patient characteristics are provided in Tables 9 and 10 below, and the treatment history is provided in Tables 11 and 12.

[0682] Table 9: Patient characteristics

[0683]

[0684] Data on CLL characteristics were obtained from the local laboratory. a The IGHV mutation status was unknown in 2 patients. b The TP53 mutation status was not mutated in 4 patients and unknown in 1 patient. c The NOTCH1 mutation status was not mutated in 4 patients and unknown in 4 patients. d The trisomy 12 status was negative in 8 patients and unknown in 1 patient. e The Del17p status was negative in 7 patients. f The Del11q status was negative in 7 patients. g The Del13q status was negative in 4 patients and unknown in 2 patients.

[0685] Table 10: Patient characteristics

[0686]

[0687] Data cutoff: September 16, 2022. a The cell of origin was unknown in 3 patients.

[0688] Table 11: Treatment history

[0689]

[0690] Data cutoff: September 16, 2022.

[0691] Table 12: Treatment history

[0692]

[0693] a The response to VR-EPOCH was unknown.

[0694] R-CHOP, rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone; R-DHAP, rituximab, dexamethasone, cytarabine, and cisplatin; VR-EPOCH, venetoclax plus dose-adjusted rituximab, etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin.

[0695] The most common treatment-emergent adverse events such as Figure 1As shown. Adverse events were mainly low-grade, including CRS; no ICANS events were observed. One case of clinical tumor lysis syndrome (grade 2) was observed; it resolved within 3 days. One grade 5 (fatal) treatment-emergent adverse event (TEAE) was observed: overall physical health deterioration in the context of progressive disease; not related to acrivastat. Six patients experienced dose delays due to TEAEs. There were no TEAEs leading to discontinuation.

[0696] CRS events were recorded as shown in Table 13. CRS events by dosing period were as Figure 2 shown. The occurrence of CRS was predictable, with most cases occurring after the first full dose of acrivastat. No grade 3 or higher CRS events were observed. All CRS events resolved, and none led to treatment discontinuation.

[0697] Table 13: CSR Events

[0698]

[0699] Data cutoff: September 16, 2022. a Grading was performed according to the Lee et al. 2019 criteria. b The median was based on the Kaplan-Meier estimate of the longest CRS duration among patients with CRS.

[0700] The depth and duration of response were as Figure 3 shown. The median time to response was 1.3 months (range: 1.1 - 2.4 months). The median time to complete response was 1.4 months (range: 1.1 - 2.8 months). The best overall response data are provided in Table 14 below.

[0701] Table 14: Best Overall Response

[0702]

[0703] Data cutoff: September 8, 2022. Median follow-up: 4.9 months (range, 0.6 - 9.3).

[0704] a Based on the modified response-evaluable population, defined as patients with ≥1 target lesion at baseline and ≥1 response evaluation after baseline and / or patients who died within 60 days after the first dose. b Response was evaluated according to the Lugano 2014 criteria. c The patient stopped treatment at C1D15 due to progression and did not receive any scans.

[0705] Tumor reduction relative to baseline (best overall response) was as Figure 4 shown.

[0706] Figure 5 Shows a clinical case study of RS-DLBCL.

[0707] Patient history:

[0708] • 76-year-old male

[0709] • Diagnosed with SLL in July 2019, started ibrutinib

[0710] • Transformed to RS-DLBCL in October 2020

[0711] • Treated with 3 cycles of R-CHOP for RS-DLBCL, mixed response

[0712] Axicabtagene ciloleucel treatment:

[0713] • First dose: SPD = 105 cm 2

[0714] • CR at weeks 6, 12, 17, 23, 36, 48, 62, 76; DS = 1, sum of products of diameters (SPD) = 2.8 cm 2

[0715] • Patient in continuous CR for over 70 weeks and still on treatment (last dose C22D1)

[0716] As of August 15, 2023, 30 subjects had been treated with at least one dose of axicabtagene ciloleucel. The overall response rate and complete metabolic response rate observed were similar to those as of September 2022, with a consistent, manageable, and tolerable safety profile.

[0717] Conclusion:

[0718] Based on data cut-off data from September 8, 2022 (efficacy) / September 16, 2022 (safety), the following conclusions were drawn: In patients with RS, SC axicabtagene ciloleucel showed a manageable safety profile with low-grade CRS events. Most CSR events occurred in cycle 1 after the first full dose of axicabtagene ciloleucel. All CSR events resolved and none led to treatment discontinuation. This safety profile was consistent with previous reports of axicabtagene ciloleucel monotherapy and no new safety signals were reported. Preliminary efficacy findings showed that SC axicabtagene ciloleucel had encouraging single-agent activity in RS-DLBCL, with high overall and complete response rates observed, and most responses observed at the first (week 6) assessment.

[0719] Based on data cutoff from September 2022, the conclusions remain favorable: acralizumab shows promising activity, with high overall response and CMR rates and a tolerable safety profile.

[0720] - Overall response rate (ORR): 60%; Complete metabolic response rate (CMR): 50%

[0721] - Only low-grade cytokine release syndrome (CRS); all resolved

[0722] - No ICANS events

[0723] - 1 case of grade 2 CTL, which resolved in 3 days

[0724] - No discontinuation due to treatment-emergent adverse events (TEAEs).

[0725] As of August 15, 2023, acralizumab continues to show high preliminary ORR and CR rates, with a manageable and tolerable safety profile. This suggests that acralizumab has the potential to be an effective, convenient, and tolerable treatment option with a favorable benefit-risk profile for patients with RS.

[0726] Table 13: Sequence Listing

[0727]

[0728]

[0729]

[0730]

[0731]

[0732]

[0733] The bold and underlined are FE; A; L and R, corresponding to positions 234 and 235; 265; 405 and 409 respectively, and the positions are according to EU numbering. In the variable region, the CDR regions annotated according to the definition of IMGT are underlined.

Claims

1. A method for treating Richter's syndrome (RS) in a human subject, the method comprising administering to the subject a bispecific antibody, the bispecific antibody comprising: (i) a first binding arm comprising a first antigen-binding region that binds to human CD3ε (epsilon) and comprises a variable heavy chain (VH) region and a variable light chain (VL) region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences in the VH region sequence of SEQ ID NO: 6, and the VL region comprises the CDR1, CDR2, and CDR3 sequences in the VL region sequence of SEQ ID NO: 7; and (ii) a second binding arm comprising a second antigen-binding region that binds to human CD20 and comprises a VH region and a VL region, wherein the VH region comprises the CDR1, CDR2, and CDR3 sequences in the VH region sequence of SEQ ID NO: 13, and the VL region comprises the CDR1, CDR2, and CDR3 sequences in the VL region sequence of SEQ ID NO: 14; wherein the bispecific antibody is administered in a dose ranging from 12 - 60 mg in a 28-day cycle.

2. The method of claim 1, wherein the bispecific antibody is administered in a dose of 24 mg.

3. The method of claim 1, wherein the bispecific antibody is administered in a dose of 48 mg.

4. The method according to any one of claims 1 - 3, wherein the bispecific antibody is administered once a week (administered weekly).

5. The method of claim 4, wherein the once-a-week administration is carried out for 2.5 28-day cycles.

6. The method according to claim 4 or 5, wherein after the once-a-week administration, the bispecific antibody is administered once every two weeks (administered every two weeks).

7. The method of claim 6, wherein the once-every-two-weeks administration is carried out for six 28-day cycles.

8. The method according to claim 6 or 7, wherein after the once-every-two-weeks administration, the bispecific antibody is administered once every four weeks.

9. The method according to any one of claims 4 - 8, wherein a priming dose of the bispecific antibody is administered in the first cycle of the 28-day cycle before administering the first once-a-week dose of 12 - 60 mg.

10. The method of claim 9, wherein the priming dose is administered two weeks before administering the first once-a-week dose of 12 - 60 mg.

11. The method according to claim 9 or 10, wherein the priming dose is in the range of 0.05 - 0.35 mg.

12. The method according to any one of claims 9 - 11, wherein the priming dose is 0.16 mg or about 0.16 mg.

13. The method according to any one of claims 9 - 12, wherein an intermediate dose of the bispecific antibody is administered after administering the priming dose and before administering the first once-a-week dose of 12 - 60 mg.

14. The method according to claim 13, wherein the sensitizing dose is administered on day 1 and the intermediate dose is administered on day 8, prior to the first once-weekly dose of 12 - 60 mg on days 15 and 22 of the first cycle.

15. The method according to claim 13 or 14, wherein the intermediate dose is in the range of 0.6 - 1.2 mg.

16. The method according to any one of claims 13 - 15, wherein the intermediate dose is 0.8 mg or about 0.8 mg.

17. The method according to any one of claims 13 - 16, wherein the bispecific antibody is administered in 28 - day cycles, wherein: e) in the first cycle, the sensitizing dose is administered on day 1, the intermediate dose is administered on day 8, and the full dose of 12 - 60 mg is administered on days 15 and 22; f) in the second to third cycles, the full dose of 12 - 60 mg is administered on days 1, 8, 15, and 22; g) in the fourth to ninth cycles, the full dose of 12 - 60 mg is administered on days 1 and 15; and h) in the tenth cycle and subsequent cycles, the full dose of 12 - 60 mg is administered on day 1.

18. The method according to claim 17, wherein the full dose is 24 mg or about 24 mg.

19. The method according to claim 17, wherein the full dose is 48 mg or about 48 mg.

20. The method according to any one of claims 1 - 19, wherein the bispecific antibody is administered subcutaneously.

21. The method according to any one of claims 1 - 20, wherein the subject has a clinical history of CLL / SLL that has transformed to aggressive lymphoma; for example, having the DLBCL subtype.

22. The method according to any one of claims 1 - 21, wherein Richter's syndrome has the DLBCL subtype.

23. The method according to any one of claims 1 - 22, wherein the subject has received one or more (such as at least two) prior lines of therapy for chronic lymphocytic leukemia (CLL) and / or for small lymphocytic lymphoma (SLL).

24. The method according to any one of the claims, wherein the prior line of therapy for CLL and / or SLL includes chemoimmunotherapy.

25. The method according to any one of the claims, wherein the prior line of therapy for CLL and / or SLL includes therapy using targeting agents such as BCL2 inhibitors or BTK inhibitors.

26. The method according to any one of the claims, wherein the prior line of therapy for CLL and / or SLL includes CAR T cell therapy.

27. The method according to any one of claims 1 - 23, wherein the subject has received prior therapy for Richter's syndrome, such as prior therapy selected from: i) rituximab combined with cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP), ii) rituximab combined with dexamethasone, cytarabine, and cisplatin (R-DHAP), and iii) Venetoclax in combination with rituximab, etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin (VR-EPOCH).

28. The method according to any one of claims 1-27, wherein the subject achieves a complete metabolic response or a partial metabolic response.

29. The method according to any one of claims 1-27, wherein the subject achieves a complete response, a partial response, or disease stabilization.

30. The method according to any one of claims 1-26, wherein the subject receives acalabrutinib as a first-line therapy for Richter's syndrome.

31. The method according to any one of claims 1-30, wherein the subject achieves a complete metabolic response or a partial metabolic response.

32. The method according to any one of claims 1-30, wherein the subject achieves a complete response, a partial response, or disease stabilization.

33. The method according to any one of claims 1-26 and 30-32, which is a first-line therapy for Richter's syndrome.

34. The method according to any one of claims 21-29, wherein the subject has refractory and / or relapsed Richter's syndrome after receiving the previous therapy.

35. The method according to any one of claims 1-34, wherein the subject is treated with prophylaxis against cytokine release syndrome (CRS).

36. The method according to claim 35, wherein the prophylaxis comprises administering a corticosteroid to the subject.

37. The method according to claim 35 or 36, wherein the corticosteroid is administered on the same day as the bispecific antibody.

38. The method according to claim 37, wherein the corticosteroid is further administered on the second, third, and fourth days after administration of the bispecific antibody.

39. The method according to any one of claims 36-38, wherein the corticosteroid is prednisolone.

40. The method according to claim 39, wherein the prednisolone is administered at an intravenous dose of 100 mg or its equivalent including an oral dose.

41. The method according to any one of claims 1-40, wherein a premedication is administered to the subject to reduce the reaction to the injection.

42. The method according to claim 41, wherein the premedication comprises an antihistamine.

43. The method according to claim 42, wherein the antihistamine is diphenhydramine.

44. The method according to claim 43, wherein the diphenhydramine is administered at an intravenous or oral dose of 50 mg or its equivalent.

45. The method according to any one of claims 41-44, wherein the premedication comprises an antipyretic.

46. The method according to claim 45, wherein the antipyretic is acetaminophen.

47. The method according to claim 46, wherein the acetaminophen is administered at an oral dose of 560 to 1000 mg or its equivalent.

48. The method according to any one of claims 41-47, wherein the premedication is administered on the same day as the bispecific antibody.

49. The method according to any one of claims 35 - 48, wherein the prophylaxis is administered during Cycle 1.

50. The method according to any one of claims 41 - 49, wherein the premedication is administered during Cycle 1.

51. The method according to claim 49 or 50, wherein when the subject experiences CRS greater than Grade 1 after the last administration of the bispecific antibody in Cycle 1, the prophylaxis is administered during Cycle 2.

52. The method according to claim 51, wherein when the subject experiences CRS greater than Grade 1 in the last administration of the bispecific antibody in a previous cycle, the prophylaxis is continued in subsequent cycles.

53. The method according to any one of claims 41 - 52, wherein the premedication is administered during Cycle 2.

54. The method according to claim 53, wherein the premedication is administered during subsequent cycles.

55. The method according to any one of claims 1 - 54, wherein if the subject develops Grade 1 CRS, an antibiotic is administered to the subject.

56. The method according to any one of claims 1 - 54, wherein if the subject develops Grade 2 or 3 CRS, a vasopressor is administered to the subject.

57. The method according to any one of claims 1 - 54, wherein if the subject develops Grade 4 CRS, at least two vasopressors are administered to the subject.

58. The method according to any one of claims 1 - 57, wherein if the subject develops Grade 2, 3, or 4 CRS, tocilizumab is administered to the subject.

59. The method according to claim 58, wherein a steroid is further administered to the subject.

60. The method according to claim 59, wherein the steroid is dexamethasone.

61. The method according to claim 59, wherein the steroid is methylprednisolone.

62. The method according to any one of claims 58 - 61, wherein if the subject is refractory to tocilizumab, tocilizumab is switched to an anti - IL - 6 antibody (e.g., siltuximab).

63. The method according to any one of claims 58 - 61, wherein if the subject is refractory to tocilizumab, tocilizumab is switched to an IL - 1R antagonist (e.g., anakinra).

64. The method according to any one of claims 1 - 63, wherein the subject is treated with prophylaxis against tumor lysis syndrome (TLS).

65. The method according to claim 64, wherein the prophylaxis against TLS comprises administering one or more urate - lowering agents before administering the bispecific antibody.

66. The method according to claim 65, wherein the one or more urate - lowering agents comprise rasburicase and / or allopurinol.

67. The method according to any one of claims 1 - 66, wherein the subject achieves a complete response, partial response, or disease stability.

68. The method according to any one of claims 1 - 67, wherein: (i) The first antigen-binding region comprises VHCDR1, VHCDR2, and VHCDR3, and VLCDR1, VLCDR2, and VLCDR3, wherein VHCDR1, VHCDR2, and VHCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 1, 2, and 3, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 4, the sequence GTN, and SEQ ID NO: 5, respectively; and (ii) The second antigen-binding region comprises VHCDR1, VHCDR2, and VHCDR3, and VLCDR1, VLCDR2, and VLCDR3, wherein VHCDR1, VHCDR2, and VHCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 8, 9, and 10, respectively, and VLCDR1, VLCDR2, and VLCDR3 comprise the amino acid sequences set forth in SEQ ID NO: 11, the sequence DAS, and SEQ ID NO: 12, respectively.

69. The method according to any one of claims 1-68, wherein: (i) The first antigen-binding region comprises a VH region comprising the amino acid sequence of SEQ ID NO: 6 and a VL region comprising the amino acid sequence of SEQ ID NO: 7; and (ii) The second antigen-binding region comprises a VH region comprising the amino acid sequence of SEQ ID NO: 13 and a VL region comprising the amino acid sequence of SEQ ID NO:

14.

70. The method according to any one of claims 1-68, wherein the first binding arm of the bispecific antibody is derived from a humanized antibody, preferably derived from a full-length IgG1, λ (lambda) antibody.

71. The method according to claim 70, wherein the first binding arm of the bispecific antibody comprises a λ light chain constant region, and the λ light chain constant region comprises the amino acid sequence set forth in SEQ ID NO:

22.

72. The method according to any one of claims 1-71, wherein the second binding arm of the bispecific antibody is derived from a human antibody, preferably derived from a full-length IgG1, κ (kappa) antibody.

73. The method according to claim 72, wherein the second binding arm comprises a κ light chain constant region, and the κ light chain constant region comprises the amino acid sequence set forth in SEQ ID NO:

23.

74. The method according to any one of claims 1-73, wherein the bispecific antibody is a full-length antibody having a human IgG1 constant region.

75. The method according to any one of claims 1-74, wherein the bispecific antibody comprises an inert Fc region.

76. The method according to any one of claims 1-75, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein in both the first heavy chain and the second heavy chain, the amino acids at positions corresponding to positions L234, L235 and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E and A, respectively.

77. The method according to any one of claims 1-76, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein in the first heavy chain, the amino acid at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and wherein in the second heavy chain, the amino acid at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa.

78. The method according to any one of claims 1-77, wherein the bispecific antibody comprises a first heavy chain and a second heavy chain, wherein (i) in both the first heavy chain and the second heavy chain, the amino acids at positions corresponding to positions L234, L235 and D265 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 are F, E and A, respectively, and (ii) in the first heavy chain, the amino acid at the position corresponding to F405 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is L, and wherein in the second heavy chain, the amino acid at the position corresponding to K409 in the human IgG1 heavy chain constant region of SEQ ID NO: 15 is R, or vice versa.

79. The method according to claim 78, wherein the bispecific antibody comprises a heavy chain constant region comprising the amino acid sequences of SEQ ID NOs: 19 and 20.

80. The method according to any one of claims 1-79, wherein the bispecific antibody comprises heavy and light chains comprising the amino acid sequences listed in SEQ ID NOs: 24 and 25, respectively, and heavy and light chains comprising the amino acid sequences listed in SEQ ID NOs: 26 and 27, respectively.

81. The method according to any one of claims 1-80, wherein the bispecific antibody comprises heavy and light chains consisting of the amino acid sequences of SEQ ID NOs: 24 and 25, respectively, and heavy and light chains consisting of the amino acid sequences of SEQ ID NOs: 26 and 27, respectively.

82. The method according to any one of claims 1-81, wherein the bispecific antibody is acrivastat or a biosimilar thereof.

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