Therapeutic combinations of acalabrutinib and a multispecific antibody to treat b-cell malignancies

Abstract

The present disclosure relates, in general, to therapeutic combinations of acalabrutinib and a multispecific antibody that binds to human CD3 and CD19, and to corresponding methods of treatment, pharmaceutical compositions, and kits.

Description

THERAPEUTIC COMBINATIONS OF ACALABRUTINIB AND A MULTISPECIFIC ANTIBODY TO TREAT B-CELL MALIGNANCIESBACKGROUND OF THE DISLCOSURE

[0001] The B-cell antigen receptor (BCR) is implicated in the pathogenesis of several B-cell malignancies, including diffuse large B-cell diffuse lymphoma, mantle cell lymphoma, chronic lymphocytic leukemia, and small lymphocytic lymphoma. Bruton Tyrosine Kinase (BTK) is an essential kinase downstream of the BCR signalling complex. Acalabrutinib (also known as ACP-196 and by the chemical name of 4-{8-amino-3-[(2S)-l-(but-2-ynoyl)pyrrolidin-2- yl]imidazo[l,5-a]pyrazin-l-yl}-N-(pyridin-2-yl)benzamide) is a selective, covalent BTK inhibitor that is the active pharmaceutical ingredient in the drug product CALQUENCE®. CALQUENCE® is FDA approved for the treatment of adults with relapsed or refractory mantle cell lymphoma, chronic lymphocytic leukemia, and small lymphocytic lymphoma, and is also being evaluated in clinical trials for the treatment of diffuse large B-cell lymphoma.

[0002] Non-Hodgkin lymphoma represents a broad spectrum of diseases arising from lymphocytes “frozen” at various stages of development. Although understanding of the biology and genetics of non-Hodgkin lymphoma has increased in recent years, few impactful or curative therapies have emerged for the treatment of relap sed / refractory or aggressive non-Hodgkin lymphoma. There currently remains a high unmet medical need due to the disease-associated morbidity and mortality of non-Hodgkin lymphoma and the few efficacious treatment options.

[0003] Non-Hodgkin lymphomas include diverse neoplasms of the lymphoid compartment; in aggregate, approximately 74,000 new cases are reported each year (National Cancer Institute [NCI] Surveillance, Epidemiology and End Results [SEER] 2020). In the United States (US), roughly 80% of these derive from the B-cell lineage. Among the B-cell non-Hodgkin lymphomas (B-NHL) the subtypes can be loosely grouped into ‘indolent’ lymphomas, including chronic lymphocytic leukemia / small lymphocytic lymphoma, follicular lymphoma (FL), and marginal zone lymphoma (MZL), and the ‘aggressive’ lymphomas including diffuse large B-cell lymphoma (DLBCL), high grade B-cell lymphoma (HGBL), mantle-cell lymphoma (MCL), and transformed indolent lymphomas; additional types and subtypes exist (Swerdlow 2017).

[0004] The present disclosure relates to the unexpected finding that a combination of acalabrutinib and a multispecific antibody can provide an efficacious therapeutic option for treating B-cell malignancies such as mantle cell lymphoma (MCL), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), and / or diffuse large B-cell lymphoma (DLBCL) as described below.SUMMARY OF THE DISCLOSURE

[0005] In one aspect, the present disclosure relates to methods of treating a B-cell malignancy in a human subject in need thereof, comprising administering to the subject a first amount of a compound of Formula I:or a pharmaceutically acceptable salt thereof, and a second amount of a CD3 x CD19 bispecific antibody or antigen-binding molecule, wherein said antibody or antigen-binding molecule is capable of specific binding to an epitope of CD3 and an epitope of CD 19, and wherein said antibody or antigen-binding molecule comprises:(1) a first Heavy Chain Variable Domain that comprises a CDRH1 (SEQ ID NO: 1), CDRH2 (SEQ ID NO: 2), and CDRH3 (SEQ ID NO: 3);(2) a Light Chain Variable Domain that comprises a CDRL1 (SEQ ID NO: 4), CDRL2 (SEQ ID NO: 5), and CDRL3(SEQ ID NO: 6); and(3) a second Heavy Chain Variable Domain that comprises a CDRH1 (SEQ ID NO: 22), CDRH2 (SEQ ID NO: 23), and CDRH3 (SEQ ID NO: 24);wherein the first amount and the second amount together comprise a therapeutically effective amount. In some aspects, the antibody or antigen-binding molecule comprises the first Heavy Chain Variable Domain comprising the amino acid sequence of SEQ ID NO: 7 and the Light Chain Variable Domain comprising the amino acid sequence of SEQ ID NO: 8. In another aspect, the antibody or antigen-binding molecule is a three-chain antibody-like molecule (TCA) comprising a first, a second, and a third polypeptide chain, wherein said first polypeptide chain has the amino acid sequence of SEQ ID NO: 11, said second polypeptide chain has the amino acid sequence of SEQ ID NO: 18, and said third polypeptide chain has the amino acid sequence of SEQ ID NO: 20. In another aspect, the antibody or antigen-binding molecule is AZD0486.

[0006] In another aspect, the B-cell malignancy is selected from the group consisting of mantle cell lymphoma; follicular lymphoma; de novo diffuse large B-cell lymphoma; transformed diffuse large B-cell lymphoma; T-cell / histiocyte-rich large B-cell lymphoma; primary cutaneous diffuse large B-cell lymphoma; leg-type primary cutaneous diffuse large B-cell lymphoma; Epstein-Barr virus-positive diffuse large B-cell lymphoma; diffuse large B-cell lymphoma associated with chronic inflammation; primary mediastinal large B-cell lymphoma; intravascular large B-cell lymphoma; anaplastic lymphoma kinase-positive (ALK+) large B-cell lymphoma; and high-grade B-cell lymphoma with rearrangements of MYC and BCL2 or of BCL6 and MYC.

[0007] In another aspect, the B-cell malignancy is non-Hodgkin lymphoma.

[0008] In another aspect, the B-cell malignancy is diffuse large B-cell lymphoma. In another aspect, the diffuse large B-cell lymphoma is selected from the group consisting of chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), or B-cell acute lymphoblastic leukemia (B-ALL), de novo diffuse large B-cell lymphoma, relapsed / refractory diffuse large B-cell lymphoma, and transformed diffuse large B-cell lymphoma. In another aspect, the diffuse large B-cell lymphoma is selected from the group consisting of germinal center B-cell diffuse large B-cell lymphoma and activated B-cell diffuse large B-cell lymphoma. In another aspect, the diffuse large B-cell lymphoma is activated B-cell diffuse large B-cell lymphoma.

[0009] In some aspects, the B-cell malignancy is mantle cell lymphoma (MCL). In some aspects, the B-cell malignancy is chronic lymphocytic leukemia (CLL). In some aspects, theB-cell malignancy is small lymphocytic lymphoma (SLL). In some aspects, the B-cell malignancy is diffuse large B-cell lymphoma (DLBCL).

[0010] In another aspect, the B-cell hematological malignancy is selected from the group consisting of non-Hodgkin’s lymphoma (NHL), Hodgkin’s lymphoma, mantle cell lymphoma (MCL), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), B-cell acute lymphoblastic leukemia (B-ALL), Burkitt’s lymphoma, multiple myeloma, myelodysplastic syndromes, and myelofibrosis.

[0011] In another aspect, the human subject has previously received at least one prior chemo-immunotherapy for the diffuse large B-cell lymphoma.

[0012] In another aspect, the B-cell malignancy is selected from the group consisting of mantle cell lymphoma, chronic lymphocytic leukemia, and small lymphocytic lymphoma.

[0013] In another aspect, the method comprises orally administering to the human subject the compound of Formula I, or a pharmaceutically acceptable salt thereof, and administering AZD0486 by an intravenous infusion (IV). In another aspect, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is co-administered to the human subject with AZD0486. In another aspect, the compound of Formula I, or a pharmaceutically acceptable salt thereof, is orally administered to the human subject before or after AZD0486.

[0014] In another aspect, the first amount of the compound of Formula I, or pharmaceutically acceptable salt thereof, administered to the human subject is from about 75 mg to about 225 mg daily (free base equivalent weight). In another aspect, the first amount of the compound of Formula I, or pharmaceutically acceptable salt thereof, administered to the human subject is about 100 mg once daily. In another aspect, the first amount of the compound of Formula I, or pharmaceutically acceptable salt thereof, administered to the human subject is about 100 mg twice daily. In another aspect, the compound of Formula I, or pharmaceutically acceptable salt thereof, is orally administered to the human subject under a continuous dosing schedule. In another aspect, the second amount of AZD0486, administered to the human subject is from about 30 pg to about 30000 pg. In some aspects, the AZD0486 is administered according to a 28-day treatment cycle treatment cycle. In some aspects, the treatment cycle is modified to include a priming dose. In some aspects, the priming dose is from about 150 pg to about 1500 pg. In some aspects, the priming dose is from about 270 pg to about 1000 pg. In some aspects,the priming dose is administered at a first timepoint in the first treatment cycle, and a full dose is administered at all subsequent timepoints. In some aspects, the priming dose is administered on day 1 of the first treatment cycle, a full dose is administered on day 15 of the first treatment cycle, and the full dose is administered on days 1 and 15 of all subsequent treatment cycles. In some aspects, the treatment cycle is modified to include at least two priming doses. In some aspects, the first priming dose is from about 150 pg to about 540 pg. In some aspects, the second priming dose is from about 800 pg to about 1200 pg. In some aspects, the first priming dose is about 270 pg and the second priming dose is about 1000 pg. In some aspects, the first priming dose is administered on day 1 of the first treatment cycle, the second priming dose is administered on day 8 of the first treatment cycle, a full dose is administered on day 15 of the first treatment cycle, and the full dose is administered on days 1 and 15 of all subsequent treatment cycles. In some aspects, optionally the patient receives six treatment cycles.

[0015] In another aspect, the disclosure relates to therapeutic combinations for simultaneous, separate, or sequential administration, wherein the combination comprises a compound of Formula I:or a pharmaceutically acceptable salt thereof, and AZD0486.

[0016] In another aspect, the disclosure relates to use of a combination comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, and AZD0486, for the treatment of a B-cell malignancy.

[0017] In another aspect, the disclosure relates to use of a combination comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, and AZD0486, in the manufacture of a medicament for the treatment of a B-cell malignancy.

[0018] In another aspect, the present disclosure relates to pharmaceutical compositions comprising: a compound of Formula I:or a pharmaceutically acceptable salt thereof, AZD0486 and a pharmaceutically acceptable carrier.

[0019] In another aspect, the present disclosure relates to kits comprising: a first pharmaceutical composition comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier; and a second pharmaceutical composition comprising AZD0486 and a pharmaceutically acceptable carrier.BRIEF DESCRIPTION OF THE DRAWINGS

[0020] FIGs. 1 A-1D show plots of tumor volume over time of individual mouse against REC- 1 cells for vehicle (1A), Acalabrutinib (IB), AZD0486 (1C), and Acalabrutinib + AZD0486 (ID) treatment groups as described in Example 1.

[0021] FIG. 2 shows a plot of mean tumor volume over time showing the antitumor activity of vehicle, Acalabrutinib, AZD0486, and Acalabrutinib + AZD0486 treatment groups against REC-1 cells as described in Example 1.

[0022] FIG. 3 shows the corresponding area under the log curve (Log AUC) values for each animal based on the plots shown in FIGs. 1 A-1D as described in Example 1.DETAILED DESCRIPTION OF THE DISCLOSURE

[0023] The practice of the present methods will employ, unless otherwise indicated, conventional techniques of molecular biology (including recombinant techniques), microbiology, cell biology, biochemistry, and immunology, which are within the skill of the art. Such techniques are explained fully in the literature, such as, “Molecular Cloning: A Laboratory Manual”, second edition (Sambrook et al., 1989); “Oligonucleotide Synthesis” (M. J. Gait, ed., 1984); “Animal Cell Culture” (R. I. Freshney, ed., 1987); “Methods in Enzymology” (Academic Press, Inc.); “Current Protocols in Molecular Biology” (F. M. Ausubel et al., eds., 1987, and periodic updates); “PCR: The Polymerase Chain Reaction”, (Mullis et al., ed., 1994); “A Practical Guide to Molecular Cloning” (Perbal Bernard V., 1988); “Phage Display: A Laboratory Manual” (Barbas et al., 2001); Harlow, Lane and Harlow, Using Antibodies: A Laboratory Manual: Portable Protocol No. I, Cold Spring Harbor Laboratory (1998); and Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory; (1988).

[0024] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range is encompassed within the disclosure. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges is also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0025] Unless indicated otherwise, antibody residues herein are numbered according to the Kabat numbering system (e.g., Kabat et al., Sequences of Immunological Interest. 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991))

[0026] In the following description, numerous specific details are set forth to provide a more thorough understanding of the methods disclosed herein. However, it will be apparent to one of skill in the art that the present methods may be practiced without one or more of these specific details. In other instances, well-known features and procedures well known to those skilled in the art have not been described in order to avoid obscuring the methods disclosed herein.

[0027] All references cited throughout the disclosure, including patent applications and publications, are incorporated by reference herein in their entirety.I. Definitions

[0028] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in the art to which this disclosure belongs.

[0029] The term “combination” can refer to simultaneous, separate, or sequential administration of two or more agents. In one aspect, “combination” can refer to simultaneous administration (e.g., administration of both agents in a single dosage form). In another aspect, “combination” refers to separate administration (e.g., administration of both agents in separate dosage forms, but at substantially the same time). In a further aspect of the disclosure, “combination” refers to sequential administration (e.g., where a first agent is administered, followed by a delay, followed by administration of a second or further agent). Where the administration is sequential or separate, the delay in administering the later component should be neither too long nor too short, so as not to lose the benefit of the combination.

[0030] The terms “co-administration,” “in combination with,” “simultaneous,” and “concurrent,” as used herein, encompass administration of two or more active pharmaceutical ingredients to a subject and include simultaneous administration in separate compositions, administration at different times in separate compositions, or administration in a composition in which two or more active pharmaceutical ingredients are present.

[0031] The term “effective amount” or “therapeutically effective amount” refers to that amount of a compound or combination of compounds as described herein that is sufficient to effect the intended application including, but not limited to, disease treatment. A therapeutically effective amount may vary depending upon the intended application (in vitro or in vivo), or the subject and disease condition being treated (e.g., the weight, age and gender of the subject), the severity of the disease condition, the manner of administration, etc. which can readily be determined byone of ordinary skill in the art. The term also applies to a dose that will induce a particular response in target cells (e.g., the reduction of platelet adhesion and / or cell migration). The specific dose will vary depending on the particular compounds chosen, the dosing regimen to be followed, whether the compound is administered in combination with other compounds, timing of administration, the tissue to which it is administered, and the physical delivery system in which the compound is carried.

[0032] A “therapeutic effect” as that term is used herein, encompasses a therapeutic benefit and / or a prophylactic benefit. A prophylactic effect includes delaying or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof.

[0033] The terms “treat,” “treating,” and “treatment” refer to at least partially alleviating, inhibiting, preventing and / or ameliorating a condition, disorder, or disease, such as B-cell malignancies. The terms “treatment of B-cell malignancies” includes both in vitro and in vivo treatments, including in warm-blooded animals such as humans. The effectiveness of treatment of B-cell malignancies can be assessed in a variety of ways, including but not limited to: inhibiting cancer cell proliferation (including the reversal of cancer growth); promoting cancer cell death (e.g., by promoting apoptosis or another cell death mechanism); improvement in symptoms; duration of response to the treatment; delay in progression of disease; and prolonging survival. Treatments can also be assessed with regard to the nature and extent of side effects associated with the treatment. Furthermore, effectiveness can be assessed with regard to biomarkers, such as levels of expression or phosphorylation of proteins known to be associated with particular biological phenomena. Other assessments of effectiveness are known to those of skill in the art.

[0034] The term “QD” means quaque die, once a day, or once daily. The term “BID” means bis in die, twice a day, or twice daily.

[0035] The term “pharmaceutically acceptable salt” refers to salts derived from a variety of organic and inorganic counter ions known in the art. Pharmaceutically acceptable acid addition salts can be formed with inorganic acids and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid and phosphoric acid. Organic acids from which salts can be derived include, for example,acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid and salicylic acid. Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese and aluminum. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins. Examples include isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine.

[0036] “Pharmaceutically acceptable carrier” or “pharmaceutically acceptable excipient” is intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and inert ingredients. The use of such pharmaceutically acceptable carriers or pharmaceutically acceptable excipients for active pharmaceutical ingredients is well known in the art. Except insofar as any conventional pharmaceutically acceptable carrier or pharmaceutically acceptable excipient is incompatible with the active pharmaceutical ingredient, its use in the therapeutic compositions of the disclosure is contemplated. Additional active pharmaceutical ingredients, such as other drugs, can also be incorporated into the described compositions and methods.

[0037] When ranges are used herein to describe, for example, dosage amounts, all combinations and sub-combinations of ranges and specific aspects therein are intended to be included.

[0038] Use of the term “about” when referring to a number or a numerical range means that the number or numerical range referred to is an approximation within experimental variability (or within statistical experimental error), and thus the number or numerical range may vary. The variation is typically from 0% to 15%, from 0% to 10%, or from 0% to 5% of the stated number or numerical range.

[0039] Unless the context requires otherwise, the terms "comprise," "comprises," and “comprising" are used on the basis and clear understanding that they are to be interpreted inclusively, rather than exclusively, and that Applicant intends each of those words to be so interpreted in construing this patent, including the claims below.

[0040] The abbreviations listed in Table 1 below have the meanings indicated in that table.Table 1

[0041] As used in this application, the amount of a compound refers to the amount of thatcompound in its free base form (i.e., the free base equivalent weight).

[0042] By “consisting essentially of’, it is meant a limitation of the scope of composition or method described to the specified materials or steps that do not materially affect the basic and novel characteristic(s) of the subject methods.

[0043] By “consisting of’, it is meant the exclusion from the composition, method, or kit of any element, step, or ingredient not specified in the claim.

[0044] Antibody residues herein are numbered according to the Kabat numbering system and the EU numbering system. The Kabat numbering system is generally used when referring to a residue in the variable domain (approximately residues 1-113 of the heavy chain) (e.g., Kabat et al., Sequences of Immunological Interest. 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991)). The “EU numbering system” or “EU index” is generally used when referring to a residue in an immunoglobulin heavy chain constant region (e.g., the EU index reported in Kabat et al., supra). The “EU index as in Kabat” refers to the residue numbering of the human IgGl EU antibody. Unless stated otherwise herein, references to residue numbers in the variable domain of antibodies mean residue numbering by the Kabat numbering system. Unless stated otherwise herein, references to residue numbers in the constant domain of antibodies mean residue numbering by the EU numbering system.

[0045] Antibodies, also referred to as immunoglobulins, conventionally comprise at least one heavy chain and one light chain, where the amino terminal domain of the heavy and light chains is variable in sequence, hence is commonly referred to as a variable region domain, or a variable heavy (VH) or variable light (VL) domain. The two domains conventionally associate to form a specific binding region, although as will be discussed here, specific binding can also be obtained with heavy chain-only variable sequences, and a variety of non-natural configurations of antibodies are known and used in the art.

[0046] A “functional” or “biologically active” antibody or antigen-binding molecule (including heavy chain-only antibodies and multi-specific (e.g., bispecific) three-chain antibody-like molecules (TCAs), described herein) is one capable of exerting one or more of its natural activities in structural, regulatory, biochemical or biophysical events. For example, a functional antibody or other binding molecule, e.g., a TCA, may have the ability to specifically bind an antigen and the binding may in turn elicit or alter a cellular or molecular event such as signal transduction or enzymatic activity. A functional antibody or other bindingmolecule, e.g., a TCA, may also block ligand activation of a receptor or act as an agonist or antagonist. The capability of an antibody or other binding molecule, e.g., a TCA, to exert one or more of its natural activities depends on several factors, including proper folding and assembly of the polypeptide chains.

[0047] The term “antibody” herein is used in the broadest sense and specifically covers monoclonal antibodies, polyclonal antibodies, monomers, dimers, multimers, multi-specific antibodies (e.g., bispecific antibodies), heavy chain-only antibodies, three chain antibodies, single chain Fv (scFv), nanobodies, etc., and also includes antibody fragments, so long as they exhibit the desired biological activity (Miller et al (2003) Jour, of Immunology 170:4854-4861). Antibodies may be murine, human, humanized, chimeric, or derived from other species.

[0048] The term “antibody” may reference a full-length heavy chain, a full length light chain, an intact immunoglobulin molecule; or an immunologically active portion of any of these polypeptides, i.e., a polypeptide that comprises an antigen binding site that immunospecifically binds an antigen of a target of interest or part thereof, such targets including but not limited to, cancer cell or cells that produce autoimmune antibodies associated with an autoimmune disease. The immunoglobulin disclosed herein can be of any type (e.g., IgG, IgE, IgM, IgD, and IgA), class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl and IgA2) or subclass of immunoglobulin molecule, including engineered subclasses with altered Fc portions that provide for reduced or enhanced effector cell activity. Light chains of the subject antibodies can be kappa light chains (Vkappa) or lambda light chains (Vlambda). The immunoglobulins can be derived from any species. In one aspect, the immunoglobulin is of largely human origin.

[0049] The term “monoclonal antibody” as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical except for possible naturally occurring mutations that may be present in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to conventional (polyclonal) antibody preparations which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. Monoclonal antibodies in accordance with the present methods can be made by the hybridoma method first described by Kohler et al. (1975) Nature 256:495, and can also bemade via recombinant protein production methods (see, e.g., U.S. Patent No. 4,816,567), for example.

[0050] The term “variable”, as used in connection with antibodies, refers to the fact that certain portions of the antibody variable domains differ extensively in sequence among antibodies and are used in the binding and specificity of each particular antibody for its particular antigen. However, the variability is not evenly distributed throughout the variable domains of antibodies. It is concentrated in three segments called hypervariable regions both in the light chain and the heavy chain variable domains. The more highly conserved portions of variable domains are called the framework regions (FRs). The variable domains of native heavy and light chains each comprise four FRs, largely adopting a P-sheet configuration, connected by three hypervariable regions, which form loops connecting, and in some cases forming part of, the P-sheet structure. The hypervariable regions in each chain are held together in close proximity by the FRs and, with the hypervariable regions from the other chain, contribute to the formation of the antigen-binding site of antibodies (see Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)). The constant domains are not involved directly in binding an antibody to an antigen, but exhibit various effector functions, such as participation of the antibody in antibody dependent cellular cytotoxicity (ADCC).

[0051] The term “hypervariable region” when used herein refers to the amino acid residues of an antibody which are responsible for antigen-binding. The hypervariable region generally comprises amino acid residues from a “complementarity determining region” or “CDR” (e.g., residues 31-35 (Hl), 50-65 (H2) and 95-102 (H3) in the heavy chain variable domain; Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)) and / or those residues from a “hypervariable loop” residues 26-32 (Hl), 53-55 (H2) and 96-101 (H3) in the heavy chain variable domain; Chothia and Lesk J. Mol. Biol. 196:901-917 (1987)). In some aspects, “CDR” means a complementary determining region of an antibody as defined in Lefranc, MP et al., IMGT, the international ImMunoGeneTics database, Nucleic Acids Res., 27:209-212 (1999). “Framework Region” or “FR” residues are those variable domain residues other than the hypervariable region / CDR residues as herein defined.

[0052] Exemplary CDR designations are shown herein, however one of skill in the art will understand that a number of definitions of the CDRs are commonly in use, including the Kabatdefinition (see “Zhao et al. A germline knowledge based computational approach for determining antibody complementarity determining regions.” Mol Immunol. 2010;47:694-700), which is based on sequence variability and is the most commonly used. The Chothia definition is based on the location of the structural loop regions (Chothia et al. “Conformations of immunoglobulin hypervariable regions.” Nature. 1989; 342:877-883). Alternative CDR definitions of interest include, without limitation, those disclosed by Honegger, “Yet another numbering scheme for immunoglobulin variable domains: an automatic modeling and analysis tool.” JMolBioL 2001;309:657-670; Ofiran et al. “Automated identification of complementarity determining regions (CDRs) reveals peculiar characteristics of CDRs and B-cell epitopes.” J Immunol. 2008;181 :6230-6235; Almagro “Identification of differences in the specificity-determining residues of antibodies that recognize antigens of different size: implications for the rational design of antibody repertoires.” J Mol Recognit. 2004;17: 132-143; and Padlanet al. “Identification of specificity-determining residues in antibodies.” Faseb J. 1995;9: 133-139., each of which is herein specifically incorporated by reference.

[0053] The terms “heavy chain-only antibody,” and “heavy chain antibody” are used interchangeably herein and refer, in the broadest sense, to antibodies, or more or more portions of an antibody, e.g., one or more arms of an antibody, lacking the light chain of a conventional antibody. The terms specifically include, without limitation, homodimeric antibodies comprising the VH antigen -binding domain and the CH2 and CH3 constant domains, in the absence of the CHI domain; functional (antigen-binding) variants of such antibodies, soluble VH variants, Ig-NAR comprising a homodimer of one variable domain (V-NAR) and five C- like constant domains (C-NAR) and functional fragments thereof; and soluble single domain antibodies (sUniDabs™). In one aspect, a heavy chain-only antibody is composed of a variable region antigen-binding domain composed of framework 1, CDR1, framework 2, CDR2, framework 3, CDR3, and framework 4. In another aspect, a heavy chain-only antibody is composed of an antigen-binding domain, at least part of a hinge region and CH2 and CH3 domains. In another aspect, a heavy chain-only antibody is composed of an antigen-binding domain, at least part of a hinge region and a CH2 domain. In a further aspect, a heavy chain-only antibody is composed of an antigen-binding domain, at least part of a hinge region and a CH3 domain. Heavy chain-only antibodies in which the CH2 and / or CH3 domain is truncated are also included herein. In a further aspect, a heavy chain is composed of an antigen binding domain, and at least one CH (CHI, CH2, CH3, or CH4) domain but no hinge region.The heavy chain-only antibody can be in the form of a dimer, in which two heavy chains are disulfide bonded or otherwise, covalently or non-covalently, attached with each other. The heavy chain-only antibody may belong to the IgG subclass, but antibodies belonging to other subclasses, such as IgM, IgA, IgD and IgE subclass, are also included herein. In a particular aspect, a heavy chain antibody is of the IgGl, IgG2, IgG3, or IgG4 subtype, in particular the IgGl subtype. In one aspect, the heavy chain-only antibodies herein are used as a binding (targeting) domain of a chimeric antigen receptor (CAR). The definition specifically includes human heavy chain-only antibodies produced by human immunoglobulin transgenic rats (UniRat™), called UniAbs™. The variable regions (VH) of UniAbs™ are called UniDabs™, and are versatile building blocks that can be linked to Fc regions or serum albumin for the development of novel therapeutics with multi-specificity, increased potency and extended half-life. Since the homodimeric UniAbs™ lack a light chain and thus a VL domain, the antigen is recognized by one single domain, i.e., the variable domain of the heavy chain of a heavy-chain antibody (VH or VHH).

[0054] An “intact antibody chain” as used herein is one comprising a full length variable region and a full length constant region (Fc). An intact “conventional” antibody comprises an intact light chain and an intact heavy chain, as well as a light chain constant domain (CL) and heavy chain constant domains, CHI, hinge, CH2 and CH3 for secreted IgG. Other isotypes, such as IgM or IgA may have different CH domains. The constant domains may be native sequence constant domains (e.g., human native sequence constant domains) or amino acid sequence variants thereof. The intact antibody may have one or more “effector functions” which refer to those biological activities attributable to the Fc constant region (a native sequence Fc region or amino acid sequence variant Fc region) of an antibody. Examples of antibody effector functions include Clq binding; complement dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; and down regulation of cell surface receptors. Constant region variants include those that alter the effector profile, binding to Fc receptors, and the like.

[0055] Depending on the amino acid sequence of the Fc (constant domain) of their heavy chains, antibodies and various antigen-binding proteins can be provided as different classes. There are five major classes of heavy chain Fc regions: IgA, IgD, IgE, IgG, and IgM, and several of these may be further divided into “subclasses” (isotypes), e.g., IgGl, IgG2, IgG3, IgG4, IgA, and IgA2. The Fc constant domains that correspond to the different classes ofantibodies may be referenced as a, 5, a, y, and p, respectively. The subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known. Ig forms include hinge-modifications or hingeless forms (Roux et al (1998) J. Immunol. 161 :4083-4090; Lund et al (2000) Eur. J. Biochem. 267:7246-7256; US 2005 / 0048572; US 2004 / 0229310). The light chains of antibodies from any vertebrate species can be assigned to one of two types, called K (kappa) and X (lambda), based on the amino acid sequences of their constant domains. Antibodies in accordance with aspects of the methods can comprise kappa light chain sequences or lambda light chain sequences.

[0056] A “functional Fc region” possesses an “effector function” of a native-sequence Fc region. Non-limiting examples of effector functions include Clq binding; CDC; Fc-receptor binding; ADCC; ADCP; down-regulation of cell-surface receptors (e.g., B-cell receptor), etc. Such effector functions generally require the Fc region to interact with a receptor, e.g., the FcyRI; FcyRIIA; FcyRIIBl; FcyRIIB2; FcyRIIIA; FcyRIIIB receptors, and the low affinity FcRn receptor; and can be assessed using various assays known in the art. A “dead” or “silenced” Fc is one that has been mutated to retain activity with respect to, for example, prolonging serum half-life, but which does not activate a high affinity Fc receptor, or which has a reduced affinity to an Fc receptor.

[0057] A “native-sequence Fc region” comprises an amino acid sequence identical to the amino acid sequence of an Fc region found in nature. Native-sequence human Fc regions include, for example, a native-sequence human IgGl Fc region (non -A and A allotypes); native-sequence human IgG2 Fc region; native-sequence human IgG3 Fc region; and nativesequence human IgG4 Fc region, as well as naturally occurring variants thereof.

[0058] A “variant Fc region” comprises an amino acid sequence that differs from that of a native-sequence Fc region by virtue of at least one amino acid modification, such as one or more amino acid substitution(s). In one aspect, the variant Fc region has at least one amino acid substitution compared to a native-sequence Fc region or to the Fc region of a parent polypeptide, e.g., from about one to about ten amino acid substitutions, such as 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, or from about one to about five amino acid substitutions in a native-sequence Fc region or in the Fc region of the parent polypeptide. The variant Fc region herein will possess at least about 80% homology with a native-sequence Fc region and / or with an Fc region of a parent polypeptide, at least about 90% homology therewith, or at least about 95% homology therewith.

[0059] The human IgG4 Fc amino acid sequence (UniProtKB No. P01861) is incorporated herein. Silenced IgGl is described, for example, in Boesch, A.W., et al., “Highly parallel characterization of IgG Fc binding interactions.” MAbs, 2014. 6(4): p. 915-27, the disclosure of which is incorporated herein by reference in its entirety.

[0060] Other Fc variants are possible, including, without limitation, one in which a region capable of forming a disulfide bond is deleted, or in which certain amino acid residues are eliminated at the N-terminal end of a native Fc, or a methionine residue is added thereto. Thus, in some aspects, one or more Fc portions of an antibody can comprise one or more mutations in the hinge region to eliminate disulfide bonding. In yet another aspect, the hinge region of an Fc can be removed entirely. In still another aspect, an antibody can comprise an Fc variant.

[0061] Further, an Fc variant can be constructed to remove or substantially reduce effector functions by substituting (mutating), deleting or adding amino acid residues to effect complement binding or Fc receptor binding. For example, and not limitation, a deletion may occur in a complement-binding site, such as a Clq-binding site. Techniques for preparing such sequence derivatives of the immunoglobulin Fc fragment are disclosed in International Patent Publication Nos. WO 97 / 34631 and WO 96 / 32478. In addition, the Fc domain may be modified by phosphorylation, sulfation, acylation, glycosylation, methylation, farnesylation, acetylation, amidation, and the like.

[0062] In some aspects, an antibody comprises a variant human IgG4 CH3 domain sequence comprising a T366W mutation, which can optionally be referred to herein as an IgG4 CH3 knob sequence. In some aspects, an antibody comprises a variant human IgG4 CH3 domain sequence comprising a T366S mutation, an L368A mutation, and a Y407V mutation, which can optionally be referred to herein as an IgG4 CH3 hole sequence. The IgG4 CH3 mutations described herein can be utilized in any suitable manner so as to place a “knob” on a first heavy chain constant region of a first monomer in an antibody dimer, and a “hole” on a second heavy chain constant region of a second monomer in an antibody dimer, thereby facilitating proper pairing (heterodimerization) of the desired pair of heavy chain polypeptide subunits in the antibody.

[0063] In some aspects, an antibody comprises a heavy chain polypeptide subunit comprising a variant human IgG4 Fc region comprising an S228P mutation, an F234A mutation, an L235A mutation, and a T366W mutation (knob). In some aspects, and antibody comprises a heavychain polypeptide subunit comprising a variant human IgG4 Fc region comprising an S228P mutation, an F234A mutation, an L235A mutation, a T366S mutation, an L368A mutation, and a Y407V mutation (hole).

[0064] The term “Fc-region-comprising antibody” refers to an antibody that comprises an Fc region. The C-terminal lysine (residue 447 according to the EU numbering system) of the Fc region may be removed, for example, during purification of the antibody or by recombinant engineering of the nucleic acid encoding the antibody. Accordingly, an antibody having an Fc region according to this disclosure can comprise an antibody with or without K447.

[0065] Aspects of the disclosure include antibodies comprising a heavy chain-only variable region in a monovalent or bivalent configuration. As used herein, the term “monovalent configuration” as used in reference to a heavy chain-only variable region domain means that only one heavy chain-only variable region domain is present, having a single binding site. In contrast, the term “bivalent configuration” as used in reference to a heavy chain-only variable region domain means that two heavy chain-only variable region domains are present (each having a single binding site), and are connected by a linker sequence. Non-limiting examples of linker sequences are discussed further herein, and include, without limitation, GS linker sequences of various lengths. When a heavy chain-only variable region is in a bivalent configuration, each of the two heavy chain-only variable region domains can have binding affinity to the same antigen, or to different antigens (e.g., to different epitopes on the same protein; to two different proteins, etc.). However, unless specifically noted otherwise, a heavy chain-only variable region denoted as being in a “bivalent configuration” is understood to contain two identical heavy chain-only variable region domains, connected by a linker sequence, wherein each of the two identical heavy chain-only variable region domains have binding affinity to the same target antigen.

[0066] Aspects of the disclosure include antibodies having multi-specific configurations, which include, without limitation, bispecific, trispecific, etc. A large variety of methods and protein configurations are known and used in bispecific monoclonal antibodies (BsMAB), tri-specific antibodies, etc.

[0067] Various methods for the production of multivalent artificial antibodies have been developed by recombinantly fusing variable domains of two or more antibodies. In some aspects, a first and a second antigen-binding domain on a polypeptide are connected by apolypeptide linker. One non-limiting example of such a polypeptide linker is a GS linker, having an amino acid sequence of four glycine residues, followed by one serine residue, and wherein the sequence is repeated n times, where n is an integer ranging from 1 to about 10, such as 2, 3, 4, 5, 6, 7, 8, or 9. Non-limiting examples of such linkers include GGGGS (n=l) and GGGGSGGGGS (n=2). Other suitable linkers can also be used, and are described, for example, in Chen et al., Adv Drug Deliv Rev. 2013 October 15; 65(10): 1357-69, the disclosure of which is incorporated herein by reference in its entirety.

[0068] The term “three-chain antibody like molecule” or “TCA” is used herein to refer to antibody-like molecules comprising, consisting essentially of, or consisting of three polypeptide subunits, two of which comprise, consist essentially of, or consist of one heavy and one light chain of a monoclonal antibody, or functional antigen-binding fragments of such antibody chains, comprising an antigen-binding region and at least one CH domain. This heavy chain / light chain pair has binding specificity for a first antigen. The third polypeptide subunit comprises, consists essentially of, or consists of a heavy-chain only antibody comprising an Fc portion comprising CH2 and / or CH3 and / or CH4 domains, in the absence of a CHI domain, and one or more antigen binding domains (e.g., two antigen binding domains) that binds an epitope of a second antigen or a different epitope of the first antigen, where such binding domain is derived from or has sequence identity with the variable region of an antibody heavy or light chain. Parts of such variable region may be encoded by VH and / or VL gene segments, D and JH gene segments, or JL gene segments. The variable region may be encoded by rearranged VHDJH, VLDJH, VHJL, or VLJL gene segments.

[0069] A TCA binding compound makes use of a “heavy chain only antibody” or “heavy chain antibody” or “heavy chain polypeptide” which, as used herein, mean a single chain antibody comprising heavy chain constant regions CH2 and / or CH3 and / or CH4 but no CHI domain. In one aspect, the heavy chain antibody is composed of an antigen-binding domain, at least part of a hinge region and CH2 and CH3 domains. In another aspect, the heavy chain antibody is composed of an antigen-binding domain, at least part of a hinge region and a CH2 domain. In a further aspect, the heavy chain antibody is composed of an antigen-binding domain, at least part of a hinge region and a CH3 domain. Heavy chain antibodies in which the CH2 and / or CH3 domain is truncated are also included herein. In a further aspect, the heavy chain is composed of an antigen binding domain, and at least one CH (CHI, CH2, CH3, or CH4) domain but no hinge region. The heavy chain only antibody can be in the form of a dimer, inwhich two heavy chains are disulfide bonded other otherwise covalently or non-covalently attached to each other, and can optionally include an asymmetric interface between one or more of the CH domains to facilitate proper pairing between polypeptide chains. The heavy-chain antibody may belong to the IgG subclass, but antibodies belonging to other subclasses, such as IgM, IgA, IgD and IgE subclass, are also included herein. In a particular aspect, the heavy chain antibody is of the IgGl, IgG2, IgG3, or IgG4 subtype, in particular the IgGl subtype or the IgG4 subtype. Non-limiting examples of a TCA binding compound are described in, for example, WO2017 / 223111 and W02018 / 052503, the disclosures of which are incorporated herein by reference in their entirety.

[0070] Heavy-chain antibodies constitute about one fourth of the IgG antibodies produced by the camelids, e.g., camels and llamas (Hamers-Casterman C., et al. Nature. 363, 446-448 (1993)). These antibodies are formed by two heavy chains but are devoid of light chains. As a consequence, the variable antigen binding part is referred to as the VHH domain and it represents the smallest naturally occurring, intact, antigen-binding site, being only around 120 amino acids in length (Desmyter, A., et al. J. Biol. Chem. 276, 26285-26290 (2001)). Heavy chain antibodies with a high specificity and affinity can be generated against a variety of antigens through immunization (van der Linden, R. H., et al. Biochim. Biophys. Acta. 1431, 37-46 (1999)) and the VHH portion can be readily cloned and expressed in yeast (Frenken, L. G. J., et al. J. Biotechnol. 78, 11-21 (2000)). Their levels of expression, solubility and stability are significantly higher than those of classical F(ab) or Fv fragments (Ghahroudi, M. A. et al. FEBS Lett. 414, 521-526 (1997)). Sharks have also been shown to have a single VH-like domain in their antibodies, termed VNAR. (Nuttall et al. Eur. J. Biochem. 270, 3543-3554 (2003); Nuttall et al. Function and Bioinformatics 55, 187-197 (2004); Dooley et al., Molecular Immunology 40, 25-33 (2003)).

[0071] The terms “CD 19” and “cluster of differentiation 19” as used herein refer to a molecule expressed during all phases of B cell development until terminal differentiation into plasma cells. The term “CD 19” includes a CD 19 protein of any human and non-human animal species, and specifically includes human CD 19 as well as CD 19 of non-human mammals.

[0072] The term “human CD 19” as used herein includes any variants, isoforms and species homologs of human CD19 (UniProt P15391), regardless of its source or mode of preparation. Thus, “human CD 19” includes human CD 19 naturally expressed by cells and CD 19 expressed on cells transfected with the human CD 19 gene.

[0073] The term “CD3” refers to the human CD3 protein multi-subunit complex. The CD3 protein multi-subunit complex is composed to 6 distinctive polypeptide chains. These include a CD3y chain (SwissProt P09693), a CD35 chain (SwissProtP04234), two CD3s chains (SwissProt P07766), and one CD3(^ chain homodimer (SwissProt 20963), and which is associated with the T-cell receptor a and P chain. The term “CD3” includes any CD3 variant, isoform and species homolog which is naturally expressed by cells (including T-cells) or can be expressed on cells transfected with genes or cDNA encoding those polypeptides, unless noted.

[0074] A “CD 19 x CD3 antibody” is a multispecific heavy chain-only antibody, such as a bispecific heavy chain-only antibody, which comprises two different antigen-binding regions, one of which binds specifically to CD 19 and one of which binds specifically to CD3.

[0075] The terms “anti-CD19 heavy chain-only antibody,” “CD 19 heavy chain-only antibody,” “anti-CD19 heavy chain antibody” and “CD 19 heavy chain antibody” are used herein interchangeably to refer to a heavy chain-only antibody as hereinabove defined, immunospecifically binding to CD 19, including human CD 19, as hereinabove defined. The definition includes, without limitation, human heavy chain antibodies produced by transgenic animals, such as transgenic rats or transgenic mice expressing human immunoglobulin, including UniRatslM producing human anti -CD 19 UniAblM antibodies, as hereinabove defined.

[0076] “Percent (%) amino acid sequence identity” with respect to a reference polypeptide sequence is defined as the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in the reference polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared. For purposes herein, however, % amino acid sequence identity values are generated using the sequence comparison computer program ALIGN-2.

[0077] An “isolated” antibody is one which has been identified and separated and / or recovered from a component of its natural environment. Contaminant components of its natural environment are materials which would interfere with diagnostic or therapeutic uses for the antibody, and may include enzymes, hormones, and other proteinaceous or nonproteinaceous solutes. In some aspects, the antibody will be purified (1) to greater than 95% by weight of antibody as determined by the Lowry method, or more than 99% by weight, (2) to a degree sufficient to obtain at least 15 residues of N-terminal or internal amino acid sequence by use of a spinning cup sequenator, or (3) to homogeneity by SDS-PAGE under reducing or nonreducing conditions using Coomassie blue or silver stain. Isolated antibody includes the antibody in situ within recombinant cells since at least one component of the antibody's natural environment will not be present. Ordinarily, however, isolated antibody will be prepared by at least one purification step.

[0078] Antibodies of the present disclosure include multi-specific antibodies. Multi-specific antibodies have more than one binding specificity. The term "multi-specific" specifically includes "bispecific" and "trispecific," as well as higher-order independent specific binding affinities, such as higher-order polyepitopic specificity, as well as tetravalent antibodies and antibody fragments. The terms "multi- specific antibody," "multi-specific heavy chain-only antibody," "multi-specific heavy chain antibody," "multi-specific Uni Ab™", and "multi-specific binding compound" are used herein in the broadest sense and cover all antibodies with more than one binding specificity. The multi-specific heavy chain anti-CD19 antibodies of the present disclosure specifically include antibodies immunospecifically binding to one single epitope on a CD 19 protein, such as a human CD 19, and to an epitope on a different protein, such as, for example, a CD3 protein (i.e., bivalent and monoparatopic). The multi- specific heavy chain anti-CD19 antibodies of the present disclosure specifically include antibodies immunospecifically binding to two or more non-overlapping epitopes on a CD 19 protein, such as a human CD19 (i.e., bivalent and biparatopic). The multi-specific heavy chain anti-CD19 antibodies of the present disclosure also specifically include antibodies immunospecifically binding to an epitope on a CD 19 protein, such as human CD 19 and to an epitope on a different protein, such as, for example, a CD3 protein, such as human CD3 (i.e., bivalent and biparatopic). The multi-specific heavy chain anti-CD19 antibodies of the present disclosure also specifically include antibodies immunospecifically binding to two or more non-overlapping or partially overlapping epitopes on a CD 19 protein, such as a human CD 19protein, and to an epitope on a different protein, such as, for example, a CD3 protein, such as human CD3 protein (i.e., trivalent and biparatopic).

[0079] An “epitope” is the site on the surface of an antigen molecule to which a single antibody molecule binds. Generally, an antigen has several or many different epitopes and reacts with many different antibodies. The term specifically includes linear epitopes and conformational epitopes.

[0080] “Epitope mapping” is the process of identifying the binding sites, or epitopes, of antibodies on their target antigens. Antibody epitopes may be linear epitopes or conformational epitopes. Linear epitopes are formed by a continuous sequence of amino acids in a protein. Conformational epitopes are formed of amino acids that are discontinuous in the protein sequence, but which are brought together upon folding of the protein into its three-dimensional structure.

[0081] The term “valent” as used herein refers to a specified number of binding sites in an antibody molecule.

[0082] A “monovalent” antibody has one binding site. Thus, a monovalent antibody is also monospecific.

[0083] A “multi-valent” antibody has two or more binding sites. Thus, the terms “bivalent”, “trivalen ’, and “tetravalent” refer to the presence of two binding sites, three binding sites, and four binding sites, respectively. Thus, a bispecific antibody according to the present disclosure is at least bivalent and may be trivalent, tetravalent, or otherwise multi-valent. A bivalent antibody in accordance with aspects of the disclosure may have two binding sites to the same epitope (i.e., bivalent, monoparatopic), or to two different epitopes (i.e., bivalent, biparatopic).

[0084] A large variety of methods and protein configurations are known and used for the preparation of bispecific monoclonal antibodies (BsMAB), tri-specific antibodies, and the like.

[0085] The term “three-chain antibody like molecule” or “TCA” is used herein to refer to antibody-like molecules comprising, consisting essentially of, or consisting of three polypeptide subunits, two of which comprise, consist essentially of, or consist of one heavy chain and one light chain of a monoclonal antibody, or functional antigen-binding fragments of such antibody chains, comprising an antigen-binding region and at least one CH domain.This heavy chain / light chain pair has binding specificity for a first antigen. The third polypeptide subunit comprises, consists essentially of, or consists of a heavy chain-only antibody comprising an Fc portion comprising CH2 and / or CH3 and / or CH4 domains, in the absence of a CHI domain, and an antigen binding domain that binds an epitope of a second antigen or a different epitope of the first antigen, where such binding domain is derived from or has sequence identity with the variable region of an antibody heavy or light chain. Parts of such variable region may be encoded by VH and / or VL gene segments, D and JH gene segments, or JL gene segments. The variable region may be encoded by rearranged VHDJH, VLDJH, VHJL, or VLJL gene segments. A TCA protein makes use of a heavy chain-only antibody as hereinabove defined.

[0086] The term “human antibody” is used herein to include antibodies having variable and constant regions derived from human germline immunoglobulin sequences. The human antibodies herein may include amino acid residues not encoded by human germline immunoglobulin sequences, e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo. The term “human antibody” specifically includes heavy chain-only antibodies having human heavy chain variable region sequences, produced by transgenic animals, such as transgenic rats or mice, in particular UniAbs™ produced by UniRats™, as defined above.

[0087] By a “chimeric antibody” or a “chimeric immunoglobulin” is meant an immunoglobulin molecule comprising amino acid sequences from at least two different Ig loci, e.g., a transgenic antibody comprising a portion encoded by a human Ig locus and a portion encoded by a rat Ig locus. Chimeric antibodies include transgenic antibodies with non-human Fc-regions or artificial Fc-regions, and human idiotypes. Such immunoglobulins can be isolated from animals of the present disclosure that have been engineered to produce such chimeric antibodies.

[0088] Antibody “effector functions” refer to those biological activities attributable to the Fc region (a native sequence Fc region or amino acid sequence variant Fc region) of an antibody. Examples of antibody effector functions include Clq binding; complement dependent cytotoxicity (CDC); Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; down regulation of cell surface receptors (e.g., B-cell receptor; BCR), etc.

[0089] “Antibody-dependent cell-mediated cytotoxicity” and “ADCC” refer to a cell-mediated reaction in which nonspecific cytotoxic cells that express Fc receptors (FcRs) (e.g., Natural Killer (NK) cells, neutrophils, and macrophages) recognize bound antibody on a target cell and subsequently cause lysis of the target cell. The primary cells for mediating ADCC, NK cells, express FcyRIII only, whereas monocytes express FcyRI, FcyRII and FcyRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol 9:457-92 (1991). To assess ADCC activity of a molecule of interest, an in vitro ADCC assay, such as that described in US Patent No. 5,500,362 or 5,821,337 may be performed. Useful effector cells for such assays include peripheral blood mononuclear cells (PBMC) and Natural Killer (NK) cells. Alternatively, or additionally, ADCC activity of the molecule of interest may be assessed in vivo, e.g., in an animal model such as that disclosed in Clynes etal. PNAS (USA) 95:652-656 (1998).

[0090] “Complement dependent cytotoxicity” or “CDC” refers to the ability of a molecule to lyse a target in the presence of complement. The complement activation pathway is initiated by the binding of the first component of the complement system (Clq) to a molecule (e.g. an antibody) complexed with a cognate antigen. To assess complement activation, a CDC assay, e.g., as described in Gazzano-Santoro et al., J. Immunol. Methods 202: 163 (1996), may be performed.

[0091] “ CD 19-positive” is or refers to an expression of CD 19 on >50% of tumor cells as assessed by a pathologist using immunohistochemistry (H4C) or flow cytometry.

[0092] The term “characterized by expression of CD 19” broadly refers to any disease or disorder in which CD 19 expression is associated with or involved with one or more pathological processes that are characteristic of the disease or disorder. Such disorders include, but are not limited to, B-cell neoplasms.

[0093] The terms “subject,” “individual,” and “patient” are used interchangeably herein to refer to a mammal being assessed for treatment and / or being treated. In an aspect, the mammal is a human. The terms “subject,” “individual,” and “patient” encompass, without limitation, individuals having cancer, individuals with autoimmune diseases, with pathogen infections, and the like. Subjects may be human, but also include other mammals, particularly those mammals useful as laboratory models for human disease, e.g., mouse, rat, etc.

[0094] A “sterile” formulation is aseptic or free or essentially free from all living microorganisms and their spores. A “frozen” formulation is one at a temperature below 0 °C.

[0095] A “stable” formulation is one in which the composition therein essentially retains its physical stability and / or chemical stability and / or biological activity upon storage. In one aspect, the formulation essentially retains its physical and chemical stability, as well as its biological activity upon storage. The storage period is generally selected based on the intended shelf-life of the formulation. Various analytical techniques for measuring protein stability are available in the art and are reviewed in Peptide and Protein Drug Delivery, 247-301. Vincent Lee Ed., Marcel Dekker, Inc., New York, N.Y., Pubs. (1991) and Jones. A. Adv. Drug Delivery Rev. 10: 29-90) (1993), for example. Stability can be measured at a selected temperature for a selected time period. Stability can be evaluated qualitatively and / or quantitatively in a variety of different ways, including evaluation of aggregate formation (for example using size exclusion chromatography, by measuring turbidity, and / or by visual inspection); by assessing charge heterogeneity using cation exchange chromatography, image capillary isoelectric focusing (icIEF) or capillary zone electrophoresis; amino-terminal or carboxy-terminal sequence analysis; mass spectrometric analysis; SDS-PAGE analysis to compare reduced and intact antibody; peptide map (for example tryptic or LYS-C) analysis; evaluating biological activity or antigen binding function of the antibody; etc. Instability may involve any one or more of: aggregation, deamidation (e.g., Asn deamidation), oxidation (e.g., Met oxidation), isomerization (e.g., Asp isomeriation), clipping / hydrolysis / fragmentation (e.g., hinge region fragmentation), succinimide formation, unpaired cysteine(s), N-terminal extension, C-terminal processing, glycosylation differences, etc.II. AZD0486 (anti-CD3 / anti-CD19)

[0096] The present disclosure relates, in part, to therapeutic combinations and corresponding methods for the treatment of B-cell malignancies, including non-Hodgkin lymphomas such as diffuse large B-cell lymphoma. In particular, the present disclosure relates to methods of treatment comprising administration of a therapeutic combination of acalabrutinib, or a pharmaceutically acceptable salt thereof, and a bispecific three chain antibody-like molecule (TCA), to a subject in need thereof, particularly a human subject in need thereof, to treat a B-cell malignancy.

[0097] In some aspects, a TCA is referred to as AZD0486, and comprises an anti-CD3 binding domain that is paired with an anti-CD 19 VH binding domain, wherein the anti-CD3 VH domainand the anti-CD3 VL domain together have binding affinity for CD3, and the TCA further comprises a heavy chain variable domain of a heavy chain-only antibody having binding affinity to CD 19, in a monovalent configuration, and the TCA further comprises a variant human IgG4 Fc domain comprising a first heavy chain constant region sequence comprising an S228P mutation, an F234A mutation, an L235A mutation, and a T366W mutation (knob), and a second heavy chain constant region sequence comprising an S228P mutation, an F234A mutation, an L235A mutation, a T366S mutation, an L368A mutation, and a Y407V mutation (hole).

[0098] In some aspects, a multi-specific antibody comprises a CD3-binding VH domain that is paired with a light chain variable domain. In certain aspects, the light chain is a fixed light chain. In some aspects, the CD3-binding VH domain comprises a CDR1 sequence of SEQ ID NO: 1, a CDR2 sequence of SEQ ID NO: 2, and a CDR3 sequence of SEQ ID NO: 3, in a human VH framework. In some aspects, the fixed light chain comprises a CDR1 sequence of SEQ ID NO: 4, a CDR2 sequence of SEQ ID NO: 5, and a CDR3 sequence of SEQ ID NO: 6, in a human VL framework. Together, the CD3 -binding VH domain and the light chain variable domain have binding affinity for CD3. In some aspects, a CD3 -binding VH domain comprises a heavy chain variable region sequence of SEQ ID NO: 7. In some aspects, a CD3-binding VH domain comprises a sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 99% percent identity to the heavy chain variable region sequence of SEQ ID NO: 7. In some aspects, a fixed light chain comprises a light chain variable region sequence of SEQ ID NO: 8. In some aspects, a fixed light chain comprises a sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 99% percent identity to the heavy chain variable region sequence of SEQ ID NO: 8

[0099] Multi-specific antibodies comprising the above-described CD3-binding VH domain and light chain variable domain have advantageous properties, for example, as described in PCT Publication No. W02018 / 052503, the disclosure of which is incorporated by reference herein in its entirety.Table 2. Anti-CD3 Heavy and Light Chain CDR1, CDR2, CDR3 amino acid sequences.Table 3. Anti-CD3 heavy and light chain variable region amino acid sequences.Table 4: Human IgG4 Fc region sequence with silencing mutations.Table 5: Additional sequences.

[0100] In some aspects, bispecific or multi-specific antibodies are provided, which may have any of the configurations discussed herein, including, without limitation, a bispecific three- chain antibody like molecule. In some aspects, a bispecific antibody can comprise at least one heavy chain variable region having binding specificity for CD 19, and at least one heavy chainvariable region having binding specificity for a different protein, e.g., CD3. In some aspects, a bispecific antibody can comprise a heavy chain / light chain pair that has binding specificity for a first antigen, and a heavy chain from a heavy chain-only antibody, comprising an Fc portion comprising CH2 and / or CH3 and / or CH4 domains, in the absence of a CHI domain, and an antigen binding domain that binds an epitope of a second antigen or a different epitope of the first antigen, in a monovalent or bivalent configuration. In one particular aspect, a bispecific antibody comprises a heavy chain / light chain pair that has binding specificity for an antigen on an effector cell (e.g., a CD3 protein on a T-cell), and a heavy chain from a heavy chain-only antibody comprising an antigen-binding domain that has binding specificity for CD 19, in a monovalent or bivalent configuration.

[0101] In some aspects, where an antibody of the present disclosure is a bispecific antibody, one arm of the antibody (one binding moiety, or one binding unit) is specific for human CD 19, while the other arm may be specific for target cells, tumor-associated antigens, targeting antigens, e.g., integrins, etc., pathogen antigens, checkpoint proteins, and the like. Target cells specifically include cancer cells. In some aspects, one arm of the antibody (one binding moiety, or one binding unit) is specific for human CD 19, while the other arm is specific for CD3.

[0102] In one aspect, an antibody is a bispecific TCA comprising a first polypeptide comprising SEQ ID NO: 11, a second polypeptide comprising SEQ ID NO: 18, and a third polypeptide comprising SEQ ID NO: 20. In some aspects, CD 19 CDR1 comprises a polypeptide comprising SEQ ID NO: 22. In some aspects, CD 19 CDR2 comprises a polypeptide comprising SEQ ID NO: 23. In some aspects, CD 19 CDR3 comprises a polypeptide comprising SEQ ID NO: 24. This antibody is also referred to as AZD0486, and is also described in PCT publication WO2021 / 222578, the disclosure of which is incorporated by reference herein in its entirety. In aspects AZD0486 is a is a bispecific TCA comprising a first polypeptide comprising SEQ ID NO: 11, a second polypeptide comprising SEQ ID NO: 18, and a third polypeptide comprising SEQ ID NO: 20.III. Preparation of Antibodies

[0103] The multispecific antibodies of the present disclosure can be prepared by methods known in the art. In one aspect, the heavy chain antibodies herein are produced by transgenic animals, including transgenic mice and rats in which the endogenous immunoglobulin genes are knocked out or disabled. In another aspect, the heavy chain antibodies herein are produced in UniRat™. UniRat™ have their endogenous immunoglobulin genes silenced and use ahuman immunoglobulin heavy-chain translocus to express a diverse, naturally optimized repertoire of fully human HCAbs. While endogenous immunoglobulin loci in rats can be knocked out or silenced using a variety of technologies, in UniRat™ the zinc-finger (endo)nuclease (ZNF) technology was used to inactivate the endogenous rat heavy chain J- locus, light chain CK locus and light chain Ck locus. ZNF constructs for microinjection into oocytes can produce IgH and IgL knock out (KO) lines. For details see, e.g., Geurts et al., 2009, Science 325:433. Characterization of Ig heavy chain knockout rats has been reported by Menoret et al., 2010, Eur. J. Immunol. 40:2932-2941. Advantages of the ZNF technology are that non-homologous end joining to silence a gene or locus via deletions up to several kb can also provide a target site for homologous integration (Cui et al., 2011, Nat Biotechnol 29:64-67). Human heavy chain antibodies produced in UniRat™ are called UniAbs™ and can bind epitopes that cannot be attacked with conventional antibodies. Their high specificity, affinity, and small size make them ideal for mono- and poly-specific applications.

[0104] In addition to UniAbs™, specifically included herein are heavy chain-only antibodies lacking the camelid VHH framework and mutations, and their functional VH regions. Such heavy chain-only antibodies can, for example, be produced in transgenic rats or mice which comprise fully human heavy chain-only gene loci as described, e.g., in W02006 / 008548, but other transgenic mammals, such as rabbit, guinea pig, rat can also be used. Heavy chain-only antibodies, including their VHH or VH functional fragments, can also be produced by recombinant DNA technology, by expression of the encoding nucleic acid in a suitable eukaryotic or prokaryotic host, including, for example, mammalian cells (e.g., CHO cells), E. coli or yeast.

[0105] Domains of heavy chain-only antibodies combine advantages of antibodies and small molecule drugs: can be mono- or multi-valent; have low toxicity; and are cost-effective to manufacture. Due to their small size, these domains are easy to administer, including oral or topical administration, are characterized by high stability, including gastrointestinal stability; and their half-life can be tailored to the desired use or indication. In addition, VH and VHH domains of HCAbs can be manufactured in a cost-effective manner.

[0106] In a particular aspect, the heavy chain antibodies of the present disclosure, including UniAbs™, have the native amino acid residue at the first position of the FR4 region (amino acid position 101 according to the Kabat numbering system), substituted by another amino acid residue, which is capable of disrupting a surface-exposed hydrophobic patch comprising orassociated with the native amino acid residue at that position. Such hydrophobic patches are normally buried in the interface with the antibody light chain constant region but become surface exposed in HCAbs and are, at least partially, for the unwanted aggregation and light chain association of HCAbs. In one aspect, the substituted amino acid residue is charged. In another aspect, the substituted amino acid residue is positively charged, such as lysine (Lys, K), arginine (Arg, R) or histidine (His, H). In another aspect the heavy chain-only antibodies derived from the transgenic animals contain a Trp to Arg mutation at position 101. In a particular aspect, the resultant HCAbs have high antigen-binding affinity and solubility under physiological conditions in the absence of aggregation.

[0107] As part of the present disclosure, human IgG anti-CD19 heavy chain antibodies with unique sequences from UniRatlM animals (UniAblM) were identified that bind human CD 19 in ELISA (recombinant CD 19 extracellular domain) protein and cell-binding assays. The identified heavy chain variable region (VH) sequences are positive for human CD 19 protein binding and / or for binding to CD 19+ cells, and are all are negative for binding to cells that do not express CD 19.

[0108] The antibodies described herein bind CD 19-positive Burkitt’s lymphoma cell lines Daudi (ATCC® CCL-2131M), Raji (ATCC® CCL-86™), and Ramos (ATCC® CRL-1596™), and some are cross-reactive with the CD 19 protein of Cynomolgus macaque. In addition, they can be engineered to provide cross-reactivity with the CD 19 protein of any animal species, if desired.

[0109] The anti -CD 19 heavy chain antibodies, such as UniAbs™ herein may have an affinity for CD 19 with a Kd of from about 10'6to around about 10’11, including without limitation: from about 10'6to around about 10'10; from about 10'6to around about 10'9; from about 10'6to around about 10'8; from about 10'8to around about 10’11; from about 10'8to around about 10'10; from about 10'8to around about 10'9; from about 10'9to around about 10’11; from about 10'9to around about 10'10; or any value within these ranges. The affinity selection may be confirmed with a biological assessment for modulating, e.g., blocking, a CD 19 biological activity, including in vitro assays, pre-clinical models, and clinical trials, as well as assessment of potential toxicity.

[0110] Heavy chain antibodies binding to non-overlapping epitopes on a CD19 protein, e.g., UniAbs™ can be identified by competition binding assays, such as enzyme-linkedimmunoassays (ELISA assays) or flow cytometric competitive binding assays. For example, one can use competition between known antibodies binding to the target antigen and the antibody of interest. By using this approach, one can divide a set of antibodies into those that compete with the reference antibody and those that do not. The non-competing antibodies are identified as binding to a distinct epitope that does not overlap with the epitope bound by the reference antibody. Often, one antibody is immobilized, the antigen is bound, and a second, labeled (e.g., biotinylated) antibody is tested in an ELISA assay for ability to bind the captured antigen. This can be performed also by using surface plasmon resonance (SPR) platforms, including ProteOn XPR36 (BioRad, Inc), Biacore 2000 and Biacore T200 (GE Healthcare Life Sciences), and MX96 SPR imager (Ibis technologies B.V.), as well as on biolayer interferometry platforms, such as Octet Red384 and Octet HTX (ForteBio, Pall Inc). For further details see the examples herein.

[0111] Typically, an antibody “competes” with a reference antibody if it causes about 15-100% reduction in the binding of the reference antibody to the target antigen, as determined by standard techniques, such as by the competition binding assays described above. In various aspects, the relative inhibition is at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50% at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95% or higher.IV. Acalabrutinib

[0112] The compound of Formula (I) is also known by the International Nonproprietary Name of acalabrutinib. For clarity, Table 6 below presents the corresponding chemical name, compound codes, and chemical structure for acalabrutinib (which are used interchangeably throughout this application with respect to the compound of Formula (I)).TABLE 6

[0113] International Publication No. W02013 / 010868 discloses acalabrutinib (Example 6) and describes the synthesis of acalabrutinib. International Publication No. W02020 / 043787 further describes the synthesis of acalabrutinib. International Publication No. W02017 / 002095 describes crystalline forms of acalabrutinib. International Publication No. WO2021 / 255246 describes acalabrutinib maleate dosage forms. International Publication Nos. W02013 / 010868, W02017 / 002095, W02020 / 043787, and WO2021 / 255246 are each incorporated by reference in their entirety. Unless otherwise stated, a general reference to acalabrutinib in this application includes acalabrutinib free base and pharmaceutically acceptable salts of acalabrutinib.V. Therapeutic Combinations, Methods of Treatment, and Uses

[0114] The present disclosure relates, in part, to therapeutic combinations and corresponding methods for the treatment of B-cell malignancies. In particular, the present disclosure relates to methods of treatment comprising administration of a therapeutic combination of acalabrutinib, or a pharmaceutically acceptable salt thereof, and AZD0486 to a subject in need thereof, particularly a human subject in need thereof, to treat a B-cell malignancy.

[0115] It has been found that therapeutic combinations of acalabrutinib, or a pharmaceutically acceptable salt thereof, and AZD0486 can be more effective than either agent alone in the treatment of B-cell malignancies. In some aspects, the combinations and methods of treatment discussed below exhibit synergistic effects that may result in greater efficacy, decreased side effects, the use of less active pharmaceutical ingredient to achieve a given clinical result, orother synergistic effects. Such combinations can provide enhanced efficacy by, for example, promoting cancer cell death, inhibiting cancer growth (e.g., inhibiting an increase in tumor volume), and / or increasing duration of response.

[0116] As reflected in the study results described later in the Examples, the combination of AZD0486 and acalabrutinib has broader and greater activity in mantle cell lymphoma than either agent alone. Acalabrutinib also has shown single agent activity in several diffuse large B-cell lymphoma cell lines, particularly those classified as activated B-cell (ABC) subtype.

[0117] Accordingly, in one aspect, the present disclosure relates to therapeutic combinations for simultaneous, separate, or sequential administration, wherein the combination comprises a compound of Formula I:or a pharmaceutically acceptable salt thereof, and AZD0486.

[0118] In another aspect, the present disclosure relates to methods of treating a B-cell malignancy in a subject in need thereof, comprising administering to the subject a first amount of a compound of Formula I:or a pharmaceutically acceptable salt thereof, and a second amount of AZD0486, wherein the first amount and the second amount together comprise a therapeutically effective amount. In another aspect, the first amount and the second amount together comprise a synergistic amount for treating the B-cell malignancy.

[0119] In some aspects, the subject is a mammal. In one aspect, the subject is a companion animal. In another aspect, the subject is a canine, feline, or equine. In another aspect, the subject is a human.

[0120] In some aspects, acalabrutinib is administered in its non-salt form (i.e., free base form).

[0121] In some aspects, acalabrutinib is administered as a pharmaceutically acceptable salt form. In one aspect, the pharmaceutically acceptable salt form is acalabrutinib maleate. In another aspect, the pharmaceutically acceptable salt form is acalabrutinib maleate monohydrate. In another aspect, the pharmaceutically acceptable salt form is the crystalline Form A of acalabrutinib maleate monohydrate. See, for example, International Publication Nos. W02017 / 002095 and WO2021 / 255246 which are incorporated by reference in their entirety. Acalabrutinib maleate monohydrate Form A also can be referred to by the alternative nomenclature of acalabrutinib maleate monohydrate Form 1. Unless otherwise stated, any reference in this disclosure to an amount of acalabrutinib, acalabrutinib maleate, or acalabrutinib maleate monohydrate is based on the acalabrutinib free base equivalent weight. For example, 100 mg refers to 100 mg of acalabrutinib free base or an equivalent amount of acalabrutinib maleate or acalabrutinib maleate monohydrate.

[0122] In another aspect, the present disclosure relates to use of a combination comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, and AZD0486 for the treatment of a B-cell malignancy.

[0123] In another aspect, the present disclosure relates to use of a combination comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, and AZD0486, in the manufacture of a medicament for the treatment of a B-cell malignancy.

[0124] In some aspects, the B-cell malignancy is aggressive lymphoma.

[0125] In some aspects, the B-cell malignancy is non-Hodgkin lymphoma.

[0126] In some aspects, the B-cell malignancy is selected from the group consisting of B-cell acute lymphoblastic leukemia, mature B-cell acute lymphoblastic leukemia, and diffuse large B-cell lymphoma.

[0127] In some aspects, the B-cell malignancy is selected from the group consisting of mantle cell lymphoma; follicular lymphoma; de novo diffuse large B-cell lymphoma; transformed diffuse large B-cell lymphoma; T-cell / histiocyte-rich large B-cell lymphoma; primary cutaneous diffuse large B-cell lymphoma; leg-type primary cutaneous diffuse large B-cell lymphoma; Epstein-Barr virus-positive diffuse large B-cell lymphoma; diffuse large B-cell lymphoma associated with chronic inflammation; primary mediastinal large B-cell lymphoma; intravascular large B-cell lymphoma; anaplastic lymphoma kinase-positive (ALK+) large B-cell lymphoma; and high-grade B-cell lymphoma with rearrangements of MYC and BCL2 or of BCL6 and MYC.

[0128] In some aspects, the B-cell malignancy is selected from the group consisting of de novo diffuse large B-cell lymphoma; transformed diffuse large B-cell lymphoma; T-cell / histiocyte-rich large B-cell lymphoma; primary cutaneous diffuse large B-cell lymphoma; leg-type primary cutaneous diffuse large B-cell lymphoma; Epstein-Barr viruspositive diffuse large B-cell lymphoma; diffuse large B-cell lymphoma associated with chronic inflammation; primary mediastinal large B-cell lymphoma; intravascular large B-cell lymphoma; anaplastic lymphoma kinase-positive (ALKY) large B-cell lymphoma; and high-grade B-cell lymphoma with rearrangements of MYC and BCL2 or of BCL6 and MYC.

[0129] In some aspects, the B-cell malignancy is diffuse large B-cell lymphoma. In one aspect, the diffuse large B-cell lymphoma is selected from the group consisting of de novo diffuse large B-cell lymphoma, relap sed / refractory diffuse large B-cell lymphoma, and transformed diffuse large B-cell lymphoma.

[0130] In some aspects, the diffuse large B-cell lymphoma is de novo diffuse large B-cell lymphoma.

[0131] In some aspects, the diffuse large B-cell lymphoma is relapsed / refractory diffuse large B-cell lymphoma.

[0132] In some aspects, the diffuse large B-cell lymphoma is transformed diffuse large B-cell lymphoma. In one aspect, the transformed diffuse large B-cell lymphoma is Richter syndrome.

[0133] In some aspects, the diffuse large B-cell lymphoma is selected from the group consisting of the germinal center B-cell diffuse large B-cell lymphoma and activated B-cell diffuse large B-cell lymphoma subtypes. In one aspect, the diffuse large B-cell lymphoma is selected from the group consisting of relap sed / refractory germinal center B-cell diffuse large B-cell lymphoma and relapsed / refractory activated B-cell diffuse large B-cell lymphoma.

[0134] In some aspects, the diffuse large B-cell lymphoma is activated B-cell diffuse large B-cell lymphoma. In one aspect, the diffuse large B-cell lymphoma is relapsed / refractory activated B-cell diffuse large B-cell lymphoma.

[0135] In some aspects, the B-cell malignancy is selected from the group consisting of mantle cell lymphoma, chronic lymphocytic leukemia, and small lymphocytic lymphoma.

[0136] In some aspects, the B-cell malignancy is mantle cell lymphoma. In one aspect, the mantle cell lymphoma is relapsed or refractory mantle cell lymphoma.

[0137] In some aspects, the B-cell malignancy is chronic lymphocytic leukemia. In one aspect, the chronic lymphocytic leukemia is relapsed or refractory chronic lymphocytic leukemia.

[0138] In some aspects, the B-cell malignancy is small lymphocytic lymphoma. In one aspect, the small lymphocytic lymphoma is relapsed or refractory small lymphocytic lymphoma.

[0139] Diagnosis of the specific B-cell malignancy from which a subject is suffering can be made in accordance with accepted clinical practice. See, for example, the 2016 classification guidelines established by the World Health Organization (WHO) for lymphoid neoplasms, or the National Comprehensive Cancer Network (NCCN) classification guidelines for non-Hodgkin lymphoma.

[0140] In some aspects, the human subject has previously received at least one prior chemo-immunotherapy for the B-cell malignancy. In one aspect, the B-cell malignancy is diffuse large B-cell lymphoma or chronic lymphocytic leukemia. In another aspect, B-cell malignancy is diffuse large B-cell lymphoma and the prior chemo-immunotherapy comprises treatment with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (i.e., R-CHOP). In another aspect, the B-cell malignancy is chronic lymphocytic leukemia and the prior chemo-immunotherapy comprises treatment with an anti-CD20 monoclonal antibody such as obinutuzumab, rituximab, or ofatumumab.VI. Combination Dosages and Dosing Regimens

[0141] The antibodies and pharmaceutical compositions described herein can be used for the treatment of diseases and conditions characterized by the expression of a target protein (e.g., CD3, CD19), including, without limitation, the conditions and diseases described further herein. In some aspects, the antibodies and pharmaceutical compositions described herein can be used for the treatment of diseases and conditions characterized by the expression of CD 19.

[0142] Effective doses of the compositions of the present disclosure for the treatment of disease vary depending upon many different factors, including means of administration, target site, physiological state of the patient, whether the patient is human or an animal, other medications administered, and whether treatment is prophylactic or therapeutic. Usually, the patient is a human, but nonhuman mammals may also be treated, e.g., companion animals such as dogs, cats, horses, etc., laboratory mammals such as rabbits, mice, rats, etc., and the like. Treatment dosages can be titrated to optimize safety and efficacy.

[0143] Typically, antibody compositions are prepared as injectables, either as liquid solutions or suspensions; solid forms suitable for solution in, or suspension in, liquid vehicles prior to injection can also be prepared. The pharmaceutical compositions herein are suitable for intravenous or subcutaneous administration, directly or after reconstitution of solid (e.g., lyophilized) compositions. The preparation also can be emulsified or encapsulated in liposomes or micro particles such as polylactide, polyglycolide, or copolymer for enhanced adjuvant effect, as discussed above. Langer, Science 249: 1527, 1990 and Hanes, Advanced Drug Delivery Reviews 28: 97-119, 1997. The agents of this disclosure can be administered in the form of a depot injection or implant preparation which can be formulated in such a manner as to permit a sustained or pulsatile release of the active ingredient. The pharmaceutical compositions are generally formulated as sterile, substantially isotonic and in full compliance with all Good Manufacturing Practice (GMP) regulations of the U.S. Food and Drug Administration.

[0144] The compositions for administration will commonly comprise an antibody or other agent (e.g., another ablative agent) dissolved in a pharmaceutically acceptable carrier, such as an aqueous carrier. A variety of aqueous carriers can be used, e.g., buffered saline and the like. These solutions are sterile and generally free of undesirable matter. These compositions may be sterilized by conventional, well known sterilization techniques. The compositions may contain pharmaceutically acceptable auxiliary substances as required to approximatephysiological conditions such as pH adjusting and buffering agents, toxicity adjusting agents and the like, e.g., sodium acetate, sodium chloride, potassium chloride, calcium chloride, sodium lactate and the like. The concentration of active agent in these formulations can vary widely, and will be selected primarily based on fluid volumes, viscosities, body weight and the like in accordance with the particular mode of administration selected and the patient's needs (e.g., Remington's Pharmaceutical Science(15th ed., 1980) and Goodman & Gillman, The Pharmacological Basis of Therapeutics (Hardman et al., eds., 1996)).

[0145] The amount of acalabrutinib administered to the subject will be dependent on the subject being treated, the severity of the disorder or condition, the rate of administration, the disposition of the compounds, and the discretion of the prescribing physician. In the various aspects described throughout this specification and unless otherwise specifically stated, all amounts recited for acalabrutinib, or a pharmaceutically acceptable salt thereof, are based on the free base equivalent weight of acalabrutinib, or the pharmaceutically acceptable salt thereof.

[0146] An effective amount of the combination of acalabrutinib and AZD0486 can be administered simultaneously, separately, or sequentially, and in either single or multiple doses, by any of the accepted modes of administration of agents having similar utilities, including rectal, buccal, intranasal and transdermal routes, by intra-arterial injection, intravenously, parenterally, intramuscularly, subcutaneously, orally, topically, or as an inhalant. In some aspects, acalabrutinib is administered orally to a human subject.

[0147] In some embodiments, the pharmaceutical compositions are suitable for oral administration. Pharmaceutical compositions suitable for oral administration can be presented as unit dosage forms, such as tablets, capsules, liquids, or aerosol sprays each containing a predetermined amount of the active ingredients. In one aspect, the pharmaceutical composition is an oral unit dosage form. In another aspect, the pharmaceutical composition is a tablet. In another aspect, the pharmaceutical composition is a capsule. In another aspect, the pharmaceutical composition is a liquid pharmaceutical composition suitable for oral consumption.

[0148] In some aspects, acalabrutinib and AZD0486 are administered simultaneously to the subject. In some aspects, acalabrutinib and AZD0486 are administered separately to the subject. In some aspects, acalabrutinib and AZD0486 are administered sequentially to the subject.

[0149] In some aspects, acalabrutinib is administered before AZD0486 is administered within a dosing cycle. In one aspect, acalabrutinib is administered at least 2 hours, at least 4 hours, at least 6 hours, at least 8 hours, at least 12 hours, at least 16 hours, at least 24 hours, or at least 48 hours before AZD0486 is administered within a dosing cycle. In another aspect, acalabrutinib is administered no more than 2 hours, no more than 4 hours, no more than 6 hours, no more than 8 hours, no more than 12 hours, no more than 16 hours, no more than 24 hours, or no more than 48 hours before AZD0486 is administered within a dosing cycle. In another aspect, acalabrutinib is administered from 2 to 4 hours before; from 4 to 6 hours before; from 6 to 8 hours before; from 8 to 12 hours before; from 12 to 16 hours before; from 16 to 24 hours before; from 20 to 28 hours before; or from 24 to 48 hours before AZD0486 is administered within a dosing cycle.

[0150] In some aspects, AZD0486 is administered before acalabrutinib is administered within a dosing cycle. In one aspect, AZD0486 is administered at least 2 hours, at least 4 hours, at least 6 hours, at least 8 hours, at least 12 hours, at least 16 hours, at least 24 hours, or at least 48 hours before acalabrutinib is administered within a dosing cycle. In another aspect, AZD0486 is administered no more than 2 hours, no more than 4 hours, no more than 6 hours, no more than 8 hours, no more than 12 hours, no more than 16 hours, no more than 24 hours, or no more than 48 hours before acalabrutinib is administered within a dosing cycle. In another aspect, AZD0486 is administered from 2 to 4 hours before; from 4 to 6 hours before; from 6 to 8 hours before; from 8 to 12 hours before; from 12 to 16 hours before; from 16 to 24 hours before; from 20 to 28 hours before; or from 24 to 48 hours before acalabrutinib is administered within a dosing cycle.A. Acalabrutinib Dosages and Dosins Regimens

[0151] In combinations with AZD0486, acalabrutinib generally is administered at a daily dosage from about 50 mg to about 400 mg. In some aspects, acalabrutinib is administered at a daily dosage from about 50 mg to about 350 mg. In one aspect, acalabrutinib is administered at a daily dosage from about 50 mg to about 300 mg. In another aspect, acalabrutinib is administered at a daily dosage from about 50 mg to about 250 mg. In another aspect, acalabrutinib is administered at a daily dosage from about 75 mg to about 225 mg. In another aspect, acalabrutinib is administered at a daily dosage from about 100 mg to about 200 mg. In another aspect, acalabrutinib is administered at a daily dosage from about 75 mg to about 125 mg. In another aspect, acalabrutinib is administered at a daily dosage from about 175 mg toabout 225 mg. In another aspect, acalabrutinib is administered at a daily dosage of about 100 mg. In another aspect, acalabrutinib is administered at a daily dosage of about 200 mg. In another aspect, acalabrutinib is administered at a daily dosage of about 400 mg.

[0152] In some aspects, acalabrutinib is administered to the subject once daily (QD). In one aspect, acalabrutinib is administered at a dosage from about 50 mg to about 400 mg once daily. In another aspect, acalabrutinib is administered at a dosage from about 50 mg to about 350 mg once daily. In another aspect, acalabrutinib is administered at a dosage from about 50 mg to about 300 mg once daily. In another aspect, acalabrutinib is administered at a dosage from about 50 mg to about 250 mg once daily. In another aspect, acalabrutinib is administered at a dosage from about 75 mg to about 225 mg once daily. In another aspect, acalabrutinib is administered at a dosage from about 100 mg to about 200 mg once daily. In another aspect, acalabrutinib is administered at a dosage from about 75 mg to about 125 mg once daily. In another aspect, acalabrutinib is administered at a dosage from about 175 mg to about 225 mg once daily. In another aspect, acalabrutinib is administered at a dosage of about 100 mg once daily. In another aspect, acalabrutinib is administered at a dosage of about 200 mg once daily. In another aspect, acalabrutinib is initially administered at a dosage of about 200 mg once daily and the acalabrutinib dosage is subsequently reduced during the dosing cycle to a dosage of about 100 mg once daily.

[0153] In some aspects, acalabrutinib is administered to the subject twice daily (BID). In one aspect, acalabrutinib is administered at a dosage from about 25 mg to about 200 mg twice daily. In another aspect, acalabrutinib is administered at a dosage from about 25 mg to about 150 mg twice daily. In another aspect, acalabrutinib is administered at a dosage from about 50 mg to about 125 mg twice daily. In another aspect, acalabrutinib is administered at a dosage from about 90 mg to about 110 mg twice daily. In another aspect, acalabrutinib is administered at a dosage from about 95 mg to about 105 mg twice daily. In another aspect, acalabrutinib is administered at a dosage of about 100 mg twice daily. In another aspect, acalabrutinib is initially administered at a dosage of about 100 mg twice daily and the acalabrutinib dosage is subsequently reduced during the dosing cycle to a dosage of about 100 mg once daily. In another aspect, acalabrutinib is administered at a dosage of about 200 mg twice daily.

[0154] In some aspects, acalabrutinib is administered under a continuous dosing schedule. A continuous dosing schedule includes no holidays during a dosing cycle. For example, in a seven-day dosing cycle, continuously dosed acalabrutinib would be given on days one, two,three, four, five, six, and seven. The dosing cycle would then repeat for the desired number of cycles. In one aspect, for example, acalabrutinib is administered for more than 1, 2, 3, 4, 5, 6, 7, 14, 21, 28, 35, 42, 49, or 56 days. In one aspect, the dosing cycle is 28 days. Administration of acalabrutinib and repeat of the dosing cycle can continue as long as tolerable and beneficial for the subject.

[0155] In some aspects, acalabrutinib is administered to the subject under an intermittent dosing schedule. In contrast to a continuous dosing schedule, an intermittent dosage schedule can include dosage holidays. For example, intermittently dosed acalabrutinib in a seven-day dosing cycle might be given on days one and two, but not given on days three, four, five, six, or seven. The dosing cycle would then repeat. This illustration could be referred to as a 2 days on / 5 days off schedule, where acalabrutinib is given for two days followed by a five-day holiday. Similarly, intermittently dosed acalabrutinib in a seven-day dosing cycle might be given on days one, two, three, and four, but not given on days five, six, or seven. The dosing cycle would then repeat. This illustration could be referred to as a 4 days on / 3 days off schedule, where acalabrutinib is given for four days followed by a three-day holiday.B. AZD0486 Dosages and Dosing Regimens

[0156] In some aspects, a patient has received at least 2 lines of systemic therapy approved for B-NHL. In some aspects, a patient has previously been exposed to, intolerant of, ineligible for, or declined therapy to a typical second line therapy such as high dose chemotherapy followed by autologous (Stem Cell Transplant (SCT)).

[0157] In some aspects, the methods involve administering a single dose of AZD0486 on a 28-day cycle for at least one cycle. In some aspects, a AZD0486 dose is selected from the group consisting of: 30 pg, 90 pg, 270 pg, 800 pg, 2400 pg, 7200 pg, 15000 pg, 30000 pg, 40000 pg, 50000 pg, 60000 pg, 70000 pg, 80000 pg, 90000 pg and 100000 pg. In some aspects, the methods involve modifying the frequency of dosing to include additional time between doses. In some aspects, the treatment cycle is repeated two or more times. In some aspects, AZD0486 is administered to the patient as a monotherapy. In some aspects, AZD0486 is administered by an intravenous infusion (IV).

[0158] In some aspects, the patient will receive one or more priming doses prior to the target dose. In some aspects, the patient will receive a single priming dose on day 1 of the first administration cycle. In some aspects, the patient will receive two priming doses. In someaspects, the patient will receive two priming doses, with the first priming dose administered on day 1 of the first administration cycle and the second priming dose administered on day 2 of the first administration cycle.

[0159] In aspects where at least one priming dose is administered, the priming dose is from 150 pg to 1500 pg. In aspects, the priming dose is from 175 pg to 1250 pg. In aspects, the priming dose is from 200 pg to 1000 pg. In aspects, the priming dose is from 200 pg to 500 pg. In aspects, the priming dose is from 250 pg to 1000 pg. In aspects, the priming dose is from 250 pg to 500 pg. In aspects, the priming dose is from 270 pg to 1000 pg.

[0160] In aspects of the methods, two priming doses are administered. In aspects where two priming doses are administered, the first priming dose is from about 150 pg to about 540 pg. In aspects, the first priming dose is from about 175 pg to about 500 pg. In aspects, the first priming dose is from about 200 pg to about 400 pg. In aspects, the first priming dose is from about 200 pg to about 300 pg. In aspects, the first priming dose is from about 250 pg to about 300 pg. In aspects, the first priming dose is about 200 pg, 210 pg, 220 pg, 230 pg, 240 pg, 250 pg, 260 pg, 270 pg, 280 pg, 290 pg, 300 pg, 310 pg, 320 pg, 330 pg, 340 pg, or 350 pg. In aspects, the first priming dose is about 270 pg. In aspects, the first priming dose is 270 pg.

[0161] In aspects where two priming doses are administered, the second priming dose is from about 800 pg to about 1200 pg. In aspects, the second priming dose is from about 850 pg to about 1150 pg. In aspects, the second priming dose is from about 900 pg to about 1100 pg. In aspects, the second priming dose is from about 950 pg to about 1050 pg. In aspects, the second priming dose is about 950 pg, 960 pg, 970 pg, 980 pg, 990 pg, 1000 pg, 1010 pg, 1020 pg, 1030 pg, 1040 pg, or 1050 pg. In aspects, the second priming dose is about 1000 pg. In aspects, the second priming dose is 1000 pg.VII. Pharmaceutical Compositions

[0162] It is another aspect of the present disclosure to provide pharmaceutical compositions comprising one or more multispecific binding compounds of the present disclosure in admixture with a suitable pharmaceutically acceptable carrier and pharmaceutical compositions comprising acalabrutinib, or a pharmaceutically acceptable salt thereof, in admixture with a suitable pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers as used herein are exemplified, but not limited to, adjuvants, solid carriers, water,buffers, or other carriers used in the art to hold therapeutic components, or combinations thereof.

[0163] In one aspect, a pharmaceutical composition comprises a compound of Formula I:or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. In another aspect, the composition comprises acalabrutinib in its free base form. In another aspect, the composition comprises acalabrutinib in its pharmaceutically acceptable salt form. In a further aspect, the composition comprises acalabrutinib maleate. In a further aspect, the composition comprises acalabrutinib maleate monohydrate. In a further aspect, the composition comprises crystalline acalabrutinib maleate monohydrate Form A. In another aspect, the composition is a dosage form disclosed in International Publication No. WO2021 / 255246. In another aspect, the composition is a 100 mg Calquence® capsule. In another aspect, the composition is a 100 mg Calquence® tablet.

[0164] In one aspect, a pharmaceutical composition comprises AZD0486 and a pharmaceutically acceptable carrier. In another aspect, a pharmaceutical composition comprises a heavy chain antibody (e.g., Uni Ab™) that binds to CD 19. In another aspect, a pharmaceutical composition comprises a multi-specific (including bispecific) heavy chain antibody (e.g., Uni Ab™) with binding specificity for two or more non-overlapping epitopes on a CD 19 protein. In one aspect, a pharmaceutical composition comprises a multi-specific (including bispecific) heavy chain antibody (e.g., UniAb™) with binding specificity to CD19 and with binding specificity to a binding target on an effector cell (e.g., a binding target on a T cell, such as, e.g., a CD3 protein on a T cell).

[0165] The pharmaceutical compositions typically are formulated to provide a therapeutically effective amount of the combinations described in this specification. The pharmaceutical compositions also may comprise one or more pharmaceutically acceptable excipients, carriers, diluents, and / or fillers. Each of acalabrutinib and AZD0486 as active ingredients can be combined in an intimate admixture with a pharmaceutical carrier according to conventional pharmaceutical compounding techniques. The carrier can take a wide variety of forms depending on the form of preparation desired for administration.

[0166] Pharmaceutical compositions of the antibodies used in accordance with the present disclosure are prepared for storage by mixing proteins having the desired degree of purity with optional pharmaceutically acceptable carriers, excipients or stabilizers (see, e.g. Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980)), such as in the form of lyophilized formulations or aqueous solutions. Acceptable carriers, excipients, or stabilizers are nontoxic to recipients at the dosages and concentrations employed, and include buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions such as sodium; metal complexes (e.g. Zn-protein complexes); and / or non-ionic surfactants such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG).

[0167] In some aspects, the pharmaceutical compositions are suitable for oral administration. Pharmaceutical compositions suitable for oral administration can be presented as unit dosage forms, such as tablets, capsules, liquids, or aerosol sprays each containing a predetermined amount of the active ingredients. In one aspect, the pharmaceutical composition is an oral unit dosage form. In another aspect, the pharmaceutical composition is a tablet. In another aspect, the pharmaceutical composition is a capsule. In another aspect, the pharmaceutical composition is a liquid pharmaceutical composition suitable for oral consumption.

[0168] In some aspects, the pharmaceutical composition is an oral unit dosage form comprising acalabrutinib, or a pharmaceutically acceptable salt thereof, in an amount from about 75 mg to about 125 mg, and a pharmaceutically acceptable carrier. In another aspect, the pharmaceutical composition is an oral unit dosage form comprising acalabrutinib, or a pharmaceutically acceptable salt thereof, in an amount of about 100 mg, and a pharmaceutically acceptable carrier.

[0169] In some aspects, pharmaceutical compositions for parenteral administration are sterile and substantially isotonic and manufactured under Good Manufacturing Practice (GMP) conditions. Pharmaceutical compositions for parenteral administration can be provided in unit dosage form (i.e., the dosage for a single administration). The formulation depends on the route of administration chosen. The antibodies herein can be administered by intravenous injection or infusion or subcutaneously. For injection administration, the antibodies herein can be formulated in aqueous solutions, such as in physiologically-compatible buffers to reduce discomfort at the site of injection. The solution can contain carriers, excipients, or stabilizers as discussed above. Alternatively, antibodies can be in lyophilized form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.

[0170] In some aspects, the pharmaceutical compositions described above are for use in the treatment of the B-cell malignancies described in this specification. For example, in one aspect, the B-cell malignancy is non-Hodgkin lymphoma. In another aspect, the B-cell malignancy is diffuse large B-cell lymphoma. In another aspect, the B-cell malignancy is activated B-cell (ABC) diffuse large B-cell lymphoma. In another aspect, the B-cell malignancy is mantle cell lymphoma. In another aspect, the B-cell malignancy is chronic lymphocytic leukemia. In another aspect, the B-cell malignancy is small lymphocytic lymphoma.VIII. Kits

[0171] Also within the scope of the present disclosure are kits comprising the active agents and formulations thereof, of the disclosure and instructions for use. The kit can further contain at least one additional reagent, e.g., a chemotherapeutic drug, etc. Kits typically include a label indicating the intended use of the contents of the kit. The term “label” as used herein includes any writing, or recorded material supplied on or with a kit, or which otherwise accompanies a kit.

[0172] The kits include each of acalabrutinib, or a pharmaceutically acceptable salt thereof, and AZD0486 either alone or in combination in suitable packaging. The kits are for coadministration of acalabrutinib, or a pharmaceutically acceptable salt thereof, and AZD0486 simultaneously, separately, or sequentially. The kits optionally comprise written material that can include instructions for use, discussion of clinical studies and listing of side effects. Such kits may also include information, such as scientific literature references, package insert materials, clinical trial results, and / or summaries of these and the like, which indicate or establish the activities and / or advantages of the composition, and / or which describe dosing, administration, side effects, drug interactions, or other information useful to the health care provider. Such information may be based on the results of various studies, for example, studies using experimental animals involving in vivo models and studies based on human clinical trials.

[0173] In some aspects, the kit comprises: a first pharmaceutical composition comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier; and a second pharmaceutical composition comprising AZD0486 and a pharmaceutically acceptable carrier.

[0174] In some aspects, the first pharmaceutical composition and the second pharmaceutical composition are each unit dosage forms. In aspects, the first pharmaceutical composition is in an oral unit dosage form and the second pharmaceutical composition is an injectable unit dosage form. In another aspect, the first pharmaceutical composition and the second pharmaceutical composition are each oral unit dosage forms. In another aspect, the firstpharmaceutical composition and the second pharmaceutical composition are each injectable unit dosage forms. In aspects, the first pharmaceutical composition comprises acalabrutinib, or a pharmaceutically acceptable salt thereof, at a dosage as provided in this disclosure. In aspects, the second pharmaceutical composition comprises AZD0486 at a dosage as provided in this disclosure.

[0175] In some aspects, acalabrutinib, or a pharmaceutically acceptable salt thereof, and AZD0486 are provided as a single composition within a container in the kit. In one aspect, therefore, the kit comprises a pharmaceutical composition comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, AZD0486, and a pharmaceutically acceptable carrier. In aspects, the first pharmaceutical composition is in an oral unit dosage form and the second pharmaceutical composition is an injectable unit dosage form. In another aspect, the first pharmaceutical composition and the second pharmaceutical composition are each oral unit dosage forms. In another aspect, the first pharmaceutical composition and the second pharmaceutical composition are each injectable unit dosage forms. In aspects, the first pharmaceutical composition comprises acalabrutinib, or a pharmaceutically acceptable salt thereof, at a dosage as provided in this disclosure. In aspects, the second pharmaceutical composition comprises AZD0486 at a dosage as provided in this disclosure.

[0176] In some aspects, the kits may further contain another active pharmaceutical ingredient. In one aspect, acalabrutinib, or a pharmaceutically acceptable salt thereof, and AZD0486, and another active pharmaceutical ingredient are provided as separate pharmaceutical compositions in separate containers within the kit. In another aspect, acalabrutinib, or a pharmaceuticallyacceptable salt thereof, and AZD0486, and the agent are provided as a single composition within a container in the kit. Suitable packaging and additional articles for use (e.g., measuring cup for liquid preparations, foil wrapping to minimize exposure to air, and the like) are known in the art and may be included in the kit. The kits can be provided, marketed and / or promoted to health providers, including physicians, nurses, pharmacists, formulary officials, and the like. The kits may also, in selected aspects, be marketed directly to the consumer.

[0177] In some aspects, the kits may further comprise a pharmaceutical composition comprising a therapeutically effective amount of cyclophosphamide, doxorubicin, vincristine, prednisone, or combinations thereof. The kit is for co-administration of acalabrutinib, cyclophosphamide, doxorubicin, vincristine, and / or prednisone, either simultaneously or separately.

[0178] In some aspects, the kits described above are for use in the treatment of the B-cell malignancies described in this specification. For example, in one aspect, the B-cell malignancy is non-Hodgkin lymphoma. In another aspect, the B-cell malignancy is diffuse large B-cell lymphoma. In another aspect, the B-cell malignancy is activated B-cell (ABC) diffuse large B-cell lymphoma. In another aspect, the B-cell malignancy is mantle cell lymphoma. In another aspect, the B-cell malignancy is chronic lymphocytic leukemia. In another aspect, the B-cell malignancy is small lymphocytic lymphoma.EXAMPLESExample 1: In vivo Activity Against REC-1 Lymphoblast Cells

[0179] In order to demonstrate the antitumor activity of AZD0486 in combination with acalabrutinib, a combination of AZD0486 and acalabrutinib was evaluated for in vivo activity in an MCL subcutaneous model using immunodeficient mice humanized with pan-T cells. The results of this study with respect to subcutaneously inoculated REC-1 lymphoblast cells are presented in FIGs. 1-3, and show responsiveness against the REC-1 lymphoblast cells.

[0180] Xenografts were established by subcutaneous (SC) injection of 5 * 106REC-1 cells suspended in 1 : 1 mixture of 50 pL of phosphate buffered saline (PBS) and 50 pL of Matrigel (Cultrex-BME-type 3) into the right flank of 6-to-8 week old animals. The date of implantation was recorded as day 0. The REC-1 cell line used in the study shown in Table 7 below.Table 7

[0181] Animals were randomized into groups based on tumor volume assessed on day 6. There were no animal substitutions. Average tumor volumes and bodyweights in each group are reported in Table 8.Table 8. Average tumor volume and bodyweight in animal groups after randomization

[0182] Human T cells were isolated from frozen human PBMCs (Donor EN01000571) using Pan T Cell Isolation Kit (Miltenyi Biotec 130-096-535) according to the manufacturer’sinstruction. Isolated T cells were activated using CD3 / CD28 / CD2 T cell activator kit (STEMCELL, 10990), 50 lU / mL of interleukin 2 (Roche, 11147528001) and Penicillin (100 IU / mL) / Streptomycin (100 pg / mL) for 3 days in ImmunoCult XF T cell expansion Medium (STEMCELL, 10981). At day 3, cells were expanded and maintained in media supplemented with 50 lU / mL of interleukin 2. The expansion was repeated every 2 days for a total of 9 days. Cells were collected and 10 x io6cells in 200 pL PBS were injected intravenously in NSG mice 6 days after tumor inoculation.

[0183] 40 female mice were used in this study. All mice were inoculated subcutaneously with 5 x 106REC-1 cells. At day 6 post tumor inoculation, each mouse was injected with 10 x 106human T cells expanded from healthy human PBMCs. Mice from each tumor model were assigned into 4 groups based on the randomization methods mentioned above. At day 7, group 1 and 2 received vehicle orally (PO) and group 3 and 4 received 20 mg / kg of acalabrutinib (administered twice a day, with the intervals of 10 and 14 hours (BID, 10 / 14). At day 13, group 1 and 3 received vehicle PBS IP, and group 2 and 4 received 0.65 mg / kg of AZD0486 IP. Group designation and dose levels are presented in Table 9.Table 9. Group Designation and Dose Levels in Study

[0184] Tumor growth was measured twice weekly using a digital calliper. The tumor volume was calculated using the formula: Tumor volume (TV) (mm3) = (d2x D) / 2, where d and D is the shortest and longest diameter in mm, respectively. The bodyweights and tumor volumes were measured, in biological safety cabinet, by using StudyLog™ software (version 4 and above). Tumor volumes of individual mouse are shown for each group in FIGs. 1 A-1D. Solid line indicates the start day of Acalabrutinib BID PO dosing and dashed lines indicate the days of IP dosing with AZD0486.

[0185] During the study, animals with tumors exceeding 2,000 mm3were considered to have reached the study endpoint and censored values were imputed by the last observation carried forward method.

[0186] The mean of tumor volume + / - SEM across the course of the experiment for each treatment group is shown in FIG. 2 and corresponding area under the log curve (Log AUC) values for each animal are shown in FIG. 3. For the analyses of tumor growth, area under the curve (AUC) values were calculated based on tumor volume versus day following the trapezoid rule. Censored values for animals reaching study endpoint were imputed by the last observation carried forward method. Log-transformation was performed for AUC values to reduce skewness and statistical significance was determined using one-way ANOVA (*p<0.0332, **p<0.0021, ***p<0.0002, ****p<0.0001). Data was graphed and analysed using Excel and GraphPad Prism 10.RESULTS

[0187] On day 29 post-tumor inoculation, tumor volume was reduced in AZD0486 + acalabrutinib treated animals (Average= 28.168mm3; SEM= 13.33) compared to the vehicle treated group (Average= 1535.03 lmm3; SEM= 100.88), AZD0486 treated group (Average= 461.333mm3; SEM= 153.69) and acalabrutinib treated group (Average= 1089.986mm3; SEM= 147.43) as shown in FIGs. 1 A-1D.

[0188] As shown in FIGs. 2-3, the Log(AUC) tumor volume analysis confirmed significant tumor growth inhibition of AZD0486 and acalabrutinib monotherapies over vehicle -treated groups (p<0.0001 and p<0.0332, respectively). Additionally, the combination acalabrutinib + AZD0486 induced a significant tumor burden control over monotherapy treatment with acalabrutinib or AZD0486 (pO.OOOl and p<0.0002, respectively).CONCLUSIONS

[0189] AZD0486 combination treatment with acalabrutinib resulted in significant anti -tumor efficacy over monotherapies in a humanized mouse model engrafted subcutaneously with REC-1 tumor B cells (MCL).Example 2: Phase I / II Study of AZD0486 in Patients with Mature B-Cell MalignanciesStudy Design:

[0190] In order to evaluate the safety and preliminary efficacy of AZD0486 administered as monotherapy and / or in combination with other anticancer agents, a Phase I / II study will be conducted on patients with mature B-Cell malignancies. This Phase VII master study will be an open-label, multi -center study that will include 3 substudies.

[0191] Each substudy population is defined as follows: Substudy 1 : Relap sed / refractory (R / R) Chronic lymphocytic leukaemia (CLL) / Small lymphocytic lymphoma (SLL); Substudy 2: R / R Mantle-cell lymphoma (MCL); and Substudy 3: Large B-cell lymphoma (LBCL) or R / R B- cell non-Hodgkin lymphoma (B-NHL).

[0192] The Substudy 1 Group will consist of 2 cohorts (Cohort 1 A and IB). Cohort 1 A will receive AZD0486 as a monotherapy through subcutaneous (SC) injection. Cohort IB will receive AZD0486 and acalabrutinib combination therapy, with AZD0486 administered as SC injection and acalabrutinib administered orally twice daily.

[0193] The Substudy 2 Group will consist of 2 cohorts (Cohort 2A and 2B). Cohort 2A will receive AZD0486 as a monotherapy through SC injection. Cohort 2B will receive AZD0486 and acalabrutinib combination therapy, with AZD0486 administered as SC injection and acalabrutinib administered orally twice daily.

[0194] The Substudy 3 Group will receive AZD0486 as intravenous (IV) infusion in combination with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone) chemotherapy as per standard of care.

[0195] Additional substudies and cohorts may be added to allow further evaluation of AZD0486 monotherapy and in combination with other anticancer agents in defined populations. Table 10 shows indication-specific and study treatment-specific information summary of the planned substudies.Table 10. Listing of SubstudiesOutcome Measures:

[0196] The primary objectives of the study are to assess the safety and tolerability of AZD0486 as monotherapy and in combination with other anticancer agents across mature B-cell malignancies. Said safety and tolerability will be measured by various metrics, including the number of participants with Adverse Events (AE), Serious Adverse Events (SAE), Adverse Events of Special Interest (AESI), and Dose Limiting Toxicity (DLTs).

[0197] The secondary outcome measures of the study include Objective Response Rate (ORR), Complete Response (CR) Rate, Duration of Response (DoR), Maximum Observed Concentration (Cmax), Area Under the Concentration-time Curve (AUC), Minimum Observed Concentration (Cmin), Time to Reach Maximum Concentration (Tmax), Trough Plasma Concentration (Ctrough), Half Life (tl / 2) of AZD0486, Clearance (CL) of AZD0486, and the number of participants with Anti-drug Antibody (ADA) for AZD0486.Definitions:

[0198] Dose Limiting Toxicity (DLT) is defined as a treatment-emergent adverse event (TEAE) that is not due to the participant’s underlying malignancy or other extraneous cause and meets the criteria. A DLT observation period will be 28 days after the first target dose of AZD0486 in monotherapy cohorts and 28 days after starting combination therapy.

[0199] Objective Response Rate (ORR) is defined as the proportion of participants achieving Objective Response (OR), defined by Complete Response (CR) or Partial Response (PR), atany point during therapy based on assessment by investigators. CR rate is proportion of participants achieving CR as best response based on disease-specific response criteria.

[0200] Duration of Response (DoR) is defined as the time from the date of first documented overall response (CR or PR) until the date of documented disease progression, relapse, or death due to any cause in the absence of disease progression or relapse, whichever occurs earlier. The end of response should coincide with date of progression or relapse or death from any cause used for the PFS endpoint.Selection of Study Population:

[0201] Inclusion criteria:1. Must be 18 years of age or over when enrolled in the study.2. Participant must meet the Eastern Cooperative Oncology Group (ECOG) performance status of 0 to 2, using contraception during treatment and at least 6 months after final dose, and has confirmed CD 19 expression if previously received anti-CD19 therapy.3. For the Substudy 1 Group, participants with CLL must require treatment according to the international workshop on Chronic Lymphocytic Leukemia (iwCLL) criteria, participants with SLL must have at least 1 measurable site per Lugano classification for NHL, and have an absolute lymphocyte count of <10,000. For cohort 1A, participants must have received at least 2 prior lines of therapy, including a Bruton tyrosine kinase inhibitor (BTKi) and B-cell lymphoma 2 inhibitor (BCL2i). For cohort IB, participants should have had received at least 1 prior line of therapy and be BTKi-sensitive. BTKi-sensitive is defined as BTKi-naive, discontinued BTKi (other than acalabrutinib) due to toxicity, or disease progressed 2 years or more from end of previous BTKi therapy.4. For the Substudy 2 Group, participants must be diagnosed with MCL per WHO criteria, classified as Clinical Stage II, III, or IV under Ann Arbor Classification, have at least 1 measurable site per Lugano classification for NHL, and have an absolute lymphocyte count of <10,000. For cohort 2A, MCL must have relapsed or progressed after 2 or more lines of therapy, including BTKi. For cohort 2B, MCL must have relapsed or progressed after 1 or more line of therapy, not including a BTKi.5. For the Substudy 3 Group, participants must be diagnosed with Large B-cell lymphoma per WHO 2022 criteria or R / R B-NHL after at least 1 prior line of therapy, have anIntemational Prognostic Index (IP I) score of 2 to 5, have at least 1 measurable site per Lugano classification for NHL, have left ventricular ejection fraction (LVEF) of >50%, and using contraception at least 12 months after the last dose of R-CHOP or 6 months after the last dose of AZD0486.

[0202] Exclusion criteria:1. A participant will not be eligible for treatment if he / she meets any of the following criteria: has central nervous system (CNS) lymphoma, has undergone surgery within 14 days of receiving the study drug, has clinically significant cardiovascular (CV) disease, exhibits unresolved Grade >2 AEs from prior anticancer therapy (except alopecia or fatigue), has received any anticancer therapy within 5 half-lives or 21 days (whichever is shorter) prior to treatment, has received radiation therapy within 28 days, has received a CAR-T or auto-haematopoietic stem cell transplant (HSCT) within 12 weeks or received a T-cell engager (TCE) within 8 weeks of the first dose of study drug treatment, or experienced Grade > 3 cytokine release syndrome (CRS) or immune effector cell-associated neurotoxicity syndrome (ICANS) event.2. For the Substudy 1 Group, participants will not be eligible for treatment if suspected transformation of CLL / SLL into more aggressive form of lymphoma. For cohort IB, participants exhibiting the following will not be eligible for treatment: bleeding diathesis, requires treatment with a strong CYP3A inhibitor or inducer, history of intracranial hemorrhage (ICH) or stroke within 6 months before first dose of study drug, GI malabsorption, receiving a vitamin K antagonist.3. For cohort 2B, participants exhibiting the following will not be eligible for treatment: bleeding diathesis, requires treatment with a strong CYP3 A inhibitor or inducer, history of ICH or stroke within 6 months before first dose of study drug, GI malabsorption, receiving a vitamin K antagonist.4. For Substudy 3 Group, participants with the following will not be eligible for treatment: Mediastinal grey-zone lymphoma, Burkitt lymphoma, Richter's transformation, primary effusion large B-cell lymphoma (LBCL), CNS lymphoma, receiving a cumulative dose of anthracycline >150 mg / m2.

[0203] This written description uses examples to disclose and to enable any person skilled in the art to practice the disclosure, including making and using any of the disclosed salts,substances, or compositions, and performing any of the disclosed methods or processes. The patentable scope of the disclosure is defined by the claims, and may include other examples that occur to those skilled in the art. Such other examples are intended to be within the scope of the claims if they have elements that do not differ from the literal language of the claims, or if they include equivalent elements with insubstantial differences from the literal language of the claims. While aspects of the disclosure are shown and described in this specification, such aspects are provided by way of example only and are not intended to otherwise limit the scope of the disclosure. Various alternatives to the described aspects of the disclosure may be employed in practicing the disclosure. Section headings as used in this section and the entire disclosure are not intended to be limiting.

[0204] All references (patent and non-patent) cited above are incorporated by reference into this patent application. The discussion of those references is intended merely to summarize the assertions made by their authors. No admission is made that any reference (or a portion of any reference) is relevant prior art (or prior art at all). Applicants reserve the right to challenge the accuracy and pertinence of the cited references.

Claims

WHAT IS CLAIMED IS:

1. A method of treating a B-cell malignancy in a human subject in need thereof, comprising administering to the human subject a first amount of a compound of Formula I:or a pharmaceutically acceptable salt thereof, and a second amount of a CD3 x CD 19 bispecific antibody or antigen-binding molecule , wherein said antibody or antigen-binding molecule is capable of specific binding to an epitope of CD3 and an epitope of CD 19, and wherein said antibody or anti gen -binding molecule comprises:(1) a first Heavy Chain Variable Domain that comprises a CDRHI (SEQ ID NO: 1), CDRH2 (SEQ ID NO: 2), and CDRH3 (SEQ ID NO: 3);(2) a Light Chain Variable Domain that comprises a CDRLI (SEQ ID NO: 4), CDRL2 (SEQ ID NO: 5), and CDRL3(SEQ ID NO: 6); and(3) a second Heavy Chain Variable Domain that comprises a CDRHI (SEQ ID NO: 22), CDRH2 (SEQ ID NO: 23), and CDRH3 (SEQ ID NO: 24); wherein the first amount and the second amount together comprise a therapeutically effective amount.

2. The method of claim 1, wherein said antibody or antigen-binding molecule comprises the first Heavy Chain Variable Domain comprising the amino acid sequence of SEQ ID NO: 7 and the Light Chain Variable Domain comprising the amino acid sequence of SEQ ID NO:

83. The method of claim 1, wherein said antibody or antigen-binding molecule is a three- chain antibody-like molecule (TCA) comprising a first, a second, and a third polypeptide chain, wherein said first polypeptide chain has the amino acid sequence of SEQ ID NO: 11, said second polypeptide chain has the amino acid sequence of SEQ ID NO: 18, and said third polypeptide chain has the amino acid sequence of SEQ ID NO:

204. The method of claim 1, wherein said antibody or antigen-binding molecule is AZD0486.

5. The method of claim 1, wherein the B-cell malignancy is non-Hodgkin lymphoma.

6. The method of claim 1, wherein the B-cell malignancy is selected from the group consisting of mantle cell lymphoma; follicular lymphoma; de novo diffuse large B-cell lymphoma; transformed diffuse large B-cell lymphoma; T-cell / histiocyte-rich large B-cell lymphoma; primary cutaneous diffuse large B-cell lymphoma; leg-type primary cutaneous diffuse large B-cell lymphoma; Epstein-Barr virus-positive diffuse large B-cell lymphoma; diffuse large B-cell lymphoma associated with chronic inflammation; primary mediastinal large B-cell lymphoma; intravascular large B-cell lymphoma; anaplastic lymphoma kinase-positive (ALKA) large B-cell lymphoma; and high-grade B-cell lymphoma with rearrangements of MYC and BCL2 or of BCL6 and MYC.

7. The method of claim 1, wherein the B-cell malignancy is selected from the group consisting of chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), or B-cell acute lymphoblastic leukemia (B-ALL), de novo diffuse large B-cell lymphoma; transformed diffuse large B-cell lymphoma; T-cell / histiocyte-rich large B-cell lymphoma; primary cutaneous diffuse large B-cell lymphoma; leg-type primary cutaneous diffuse large B-cell lymphoma; Epstein-Barr virus-positive diffuse large B-cell lymphoma; diffuse large B-cell lymphoma associated with chronic inflammation; primary mediastinal large B-cell lymphoma; intravascular large B-cell lymphoma; anaplastic lymphoma kinase-positive (ALKA) large B-cell lymphoma; and high-grade B-cell lymphoma with rearrangements of MYC and BCL2 or of BCL6 and MYC.

8. The method of claim 1, wherein the B-cell malignancy is selected from the group consisting of diffuse large B-cell lymphoma (DLBCL), mantle cell lymphoma (MCL), chronic lymphocytic leukemia (CLL), and small lymphocytic lymphoma (SLL).

9. The method of claim 8, wherein the B-cell malignancy is mantle cell lymphoma (MCL).

10. The method of claim 8, wherein the B-cell malignancy is chronic lymphocytic leukemia (CLL).

11. The method of claim 8, wherein the B-cell malignancy is small lymphocytic lymphoma (SLL).

12. The method of claim 8, wherein the B-cell malignancy is diffuse large B-cell lymphoma (DLBCL).

13. The method of claim 8, wherein the diffuse large B-cell lymphoma is selected from the group consisting of de novo diffuse large B-cell lymphoma, relap sed / refractory diffuse large B-cell lymphoma, and transformed diffuse large B-cell lymphoma.

14. The method of claim 8, wherein the diffuse large B-cell lymphoma is selected from the group consisting of germinal center B-cell diffuse large B-cell lymphoma and activated B-cell diffuse large B-cell lymphoma.

15. The method of claim 8, wherein the diffuse large B-cell lymphoma is activated B-cell diffuse large B-cell lymphoma.

16. The method of any one of claims 8 to 15, wherein the human subject has previously received at least one prior chemo-immunotherapy for the diffuse large B-cell lymphoma.

17. The method of any one of claims 1 to 16 wherein the method comprises orally administering to the human subject the compound of Formula I, or a pharmaceutically acceptable salt thereof, and AZD0486 is administered by an intravenous infusion (IV).

18. The method of claim 17, wherein the compound of Formula I, or a pharmaceutically acceptable salt thereof, is co-administered to the human subject with AZD0486.

19. The method of claim 18, wherein the compound of Formula I, or a pharmaceutically acceptable salt thereof, is orally administered to the human subject before or after AZD0486.

20. The method of any one of claims 1 to 19, wherein the first amount of the compound of Formula I, or a pharmaceutically acceptable salt thereof, administered to the human subject is from about 75 mg to about 225 mg daily.

21. The method of any one of claims 1 to 19, wherein the first amount of the compound of Formula I, or a pharmaceutically acceptable salt thereof, administered to the human subject is about 100 mg once daily.

22. The method of any one of claims 1 to 19, wherein the first amount of the compound of Formula I, or a pharmaceutically acceptable salt thereof, administered to the human subject is about 100 mg twice daily.

23. The method of any one of claims 1 to 22, wherein the compound of Formula I, or a pharmaceutically acceptable salt thereof, is orally administered to the human subject under a continuous dosing schedule.

24. The method of any one of claims 1 to 23, wherein the second amount of AZD0486, administered to the human subject is from about 30 pg to about 30000 pg.

25. The method according to claim 24, wherein the AZD0486 is administered according to a 28-day treatment cycle treatment cycle.

26. The method of claim 25, wherein the treatment cycle is modified to include a priming dose.

27. The method according to claim 26, wherein the priming dose is from about 150 pg to about 1500 pg.

28. The method according to claim 27, wherein the priming dose is from about 270 pg to about 1000 pg.

29. The method according to any one of claims 26-28, wherein the priming dose is administered at a first timepoint in the first treatment cycle, and a full dose is administered at all subsequent timepoints.

30. The method according to claim 29, wherein the priming dose is administered on day 1 of the first treatment cycle, a full dose is administered on day 15 of the first treatment cycle, and the full dose is administered on days 1 and 15 of all subsequent treatment cycles.

31. The method according to claim 26, wherein the treatment cycle is modified to include at least two priming doses.

32. The method according to claim 31, wherein the first priming dose is from about 150 pg to about 540 pg.

33. The method according to claim 31 or claim 32, wherein the second priming dose is from about 800 pg to about 1200 pg.

34. The method according to any one of claims 31-33, wherein the first priming dose is about 270 pg and the second priming dose is about 1000 pg.

35. The method according to any one of claims 31-34, wherein the first priming dose is administered on day 1 of the first treatment cycle, the second priming dose is administered on day 8 of the first treatment cycle, a full dose is administered on day 15 of the first treatment cycle, and the full dose is administered on days 1 and 15 of all subsequent treatment cycles.

36. A combination for simultaneous, separate, or sequential administration, wherein the combination comprises a compound of Formula I:or a pharmaceutically acceptable salt thereof, and AZD0486.

37. Use of a combination comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, andAZD0486, for the treatment of a B-cell malignancy.

38. Use of a combination comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, andAZD0486, in the manufacture of a medicament for the treatment of a B-cell malignancy.

39. A pharmaceutical composition comprising: a compound of Formula I:or a pharmaceutically acceptable salt thereof,AZD0486, and a pharmaceutically acceptable carrier.

40. A kit comprising: a first pharmaceutical composition comprising a compound of Formula I:or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier; and a second pharmaceutical composition comprising AZD0486 and a pharmaceutically acceptable carrier.

Images