WO2018229706A1

WO2018229706A1 - Combination therapy for the treatment of cancer

Info

Publication number: WO2018229706A1
Application number: PCT/IB2018/054369
Authority: WO
Inventors: Haihui Lu; Matthew John MEYER; Ryan Sullivan
Original assignee: Novartis AG
Current assignee: Novartis AG
Priority date: 2017-06-16
Filing date: 2018-06-14
Publication date: 2018-12-20
Anticipated expiration: 2019-12-16

Abstract

The present invention relates to a pharmaceutical composition comprising an anti-CD32b antibody molecule in combination with an IL-15/IL-15Ra complex. The present combination can be administered independently or separately, in a quantity which is therapeutically effective for the treatment of cancer. The invention also provides the use of such a combination for the manufacture of a medicament; the use of such a combination as a medicine; a kit of part comprising such a combination; and a method of treatment of such a combination.

Description

COMBINATION THERAPY FOR THE TREATMENT OF CANCER

FIELD OF THE INVENTION

The present invention relates to antibody molecules which bind human CD32b in combination with an additional agent that enhances antibody dependent cellular cytotoxicity (ADCC), such as a complex comprising interleukin-15 ("IL-15") bound to IL-15 receptor alpha ("IL-15Ra"). In a specific aspect, the combination is useful in the prevention, treatment, and/or management of disorders in which activating the immune system is beneficial, such as cancer.

BACKGROUND OF THE INVENTION

Fc gamma receptors (FcyR) bind IgG and they are expressed by many immune cells, enabling them to serve as the link between innate and humoral immunity. Activatory FcyRs mediate immune responses including phagocytosis and ADCC (Nimmerjahn & Ravetch (2008) Nature Rev. Immunol. 8(1): 34-47). CD32b (FcyRIIb) is the sole inhibitory FcyR. It is expressed by immune cells including dendritic cells and macrophages and is the only FcyR expressed on B cells (Nimmerjahn & Ravetch, supra;

Amigorena et al., (1989) Eur. J. Immunol. 19(8): 1379-1385). Activation of CD32b results in inhibition of activatory FcyR functions, i.e. inhibition of phagocytosis and ADCC (Smith & Clatworthy, (2010) Nat. Rev. Immunol. 10(5): 328-343), or when cross-linked to the B cell receptor, reduced B cell function (Horton et al., (2011) J. Immunol. 186(7): 4223-4233). As such, an antibody molecule that binds to CD32b can reduce the inhibitory effects of this receptor and when combined with an agent that triggers ADCC, will give rise to an enhanced ADCC response.

The cytokine, interleukin-15 (IL-15), is a member of the four alpha-helix bundle family of lymphokines produced by many cells in the body. IL-15 plays a pivotal role in modulating the activity of both the innate and adaptive immune system, e.g., maintenance of the memory T-cell response to invading pathogens, inhibition of apoptosis, activation of dendritic cells, and induction of Natural Killer (NK) cell proliferation and cytotoxic activity. IL-15 signaling has been shown to occur through the heterodimeric complex of the IL-15 receptor, which consists of three polypeptides, the type-specific IL- 15 receptor alpha ("IL-15Ra"), the IL-2/IL-15 receptor beta (or CD122) ("β"), and the common gamma chain (or CD 132) ("y") that is shared by multiple cytokine receptors. Based on its multifaceted role in the immune system, various therapies designed to modulate IL-15 -mediated function have been explored. Recent reports suggest that IL- 15, when complexed with the sIL- 15Ra, or the sushi domain, maintains its immune enhancing function. Recombinant IL-15 and IL-15/IL-15Ra complexes have been shown to promote to different degrees the expansion of memory CD8 T cells and NK cells and enhance tumor rejection in various preclinical models. Furthermore, tumor targeting of IL-15 or IL-15/IL-15Ra complex containing constructs in mouse models, resulted in improved anti-tumor responses in either

immunocompetent animals transplanted with syngeneic tumors or in T- and B cell-deficient SCID mice (retaining NK cells) injected with human tumor cell lines. Enhanced anti -tumor activity is thought to be dependent on increased half-life of the IL-15 -containing moiety as well as the trans-presentation of IL-15 on the surface of tumor cells leading to enhanced NK and/or CD8 cytotoxic T cell expansion within the tumor. As such, tumor cells engineered to express IL-15 were also reported to promote rejection of established tumors by enhancing T cell and NK cell recruitment, proliferation and function (Zhang et al, (2009) PNAS USA. 106:7513-7518; Munger et al, (1995) Cell Immunol. 165(2):289-293; Evans et al, (1997) Cell Immunol. 179(l):66-73; Klebanoff ei a/., (2004) PNAS USA. 101(7): 1969-74; Sneller et al, (2011) Blood. l l8(26):6845-6848; Zhang et al, (2012) J. Immunol. 188(12):6156-6164).

NK cells are one cellular mediator of ADCC and given the role shown for IL-15 and IL-15/IL- 15Ra complexes in enhancing NK-cell functionality, a role for IL-15 in enhancing ADCC has also been investigated. IL-15 has been shown to enhance NK-cell ADCC in vitro (Carson et al, (1994) J. Exp. Med. 180: 1395-403), including that directed by anti-CD20 mAbs (Moga et al, (2008) Exp. Hematol. 36: 69-77). More recently, a study by Rosario et al, (2015) showed that an IL-15 superagonist augmented the response of NK cells when directed by anti-CD20 mAbs, against lymphoma targets, in vitro and in vivo (Rosario et al, (2015) Clin. Cancer Res. 22(3): 596-608). Since IL-15 has been shown to enhance the CD20 mAb directed ADCC responses by NK cells against lymphoma cells, it is believed that IL-15 could also potentiate the ADCC activity of other therapeutic agents that utilise ADCC as a mechanism of action.

Therefore, therapeutic approaches that enhance anti-tumor immunity could work more effectively when the immune response is optimized by targeting multiple components at one or more stages of an immune response. For example, approaches that enhance cellular and humoral immune responses (e.g., by stimulating (e.g., NK cells) and e.g., disinhibiting ADCC (e.g., by inhibiting CD32b), can result in a more effective and/or prolonged therapeutic response. Therefore combination therapies for cancer immunotherapy are desirable.

SUMMARY OF THE INVENTION

Accordingly, disclosed herein are combination therapies that enhance ADCC and as such can provide a superior beneficial effect, e.g., in the treatment of a disorder, such as an enhanced anti -cancer effect, reduced toxicity and/or reduced side effects, compared to monotherapy administration of the therapeutic agents of the combination. For example, one or more of the therapeutic agents in the combination can be administered at a lower dosage, or for a shorter period of administration or less frequently, than would be required to achieve the same therapeutic effect compared to the monotherapy administration. More specifically, one of the therapeutic agents in the combination can be administered to enhance the effect of the other agent. Thus, compositions and methods for treating cancer and other immune disorders using combination therapies are disclosed.

In one aspect, the combination therapy comprises an agent that modulates the inhibition of activatory FcyR functions such as an antibody to CD32b, in combination with immune enhancing agents such as IL-15 complexed with sIL-15Ra, to enhance the immune system. In an embodiment, the present invention provides a combination comprising an anti-human CD32b antibody molecule in combination with an IL-15/IL-15Ra complex.

In one embodiment, the anti- human CD32b antibody molecule of the combination comprises a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 113, a VHCDR2 amino acid sequence of SEQ ID NO: 114 and a VHCDR3 amino acid sequence of SEQ ID NO: 115, as described in Table 1 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 120, a VLCDR2 amino acid sequence of SEQ ID NO: 121 and a VLCDR3 amino acid sequence of SEQ ID NO: 122 as described in Table 1.

In one embodiment, the anti-CD32b antibody molecule of the combination comprises a heavy chain variable domain (VH) comprising an amino acid sequence at least 90% identical to SEQ ID NO: 116 and a light chain variable domain (VL) comprising an amino acid sequence at least 90% identical to SEQ ID NO: 123, as described in Table 1.

In one embodiment, the anti-CD32b antibody molecule of the combination comprises a VH comprising an amino acid sequence of SEQ ID NO: 1 16 and a VL comprising an amino acid sequence of SEQ ID NO: 123, as described in Table 1.

In one embodiment, the anti-CD32b antibody molecule of the combination comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 118 and a light chain comprising the amino acid sequence of SEQ ID NOs: 125, as described in Table 1.

In another embodiment, the present application discloses an isolated antibody molecule that specifically binds to CD32b within the Fc binding domain of CD32b. In some embodiments, the antibody binds within amino acid residues 107-123 (VLRCHSWKDKPLVKVTF; SEQ ID NO: 250) of CD32b, as described in Table 1.

In an alternative embodiment, the anti-CD32b antibody molecule of the combination comprises the antibody 2B6 as described in PCT Publication WO2004/016750 (Koenig & Maria-Concetta) and Maria-Concetta et al (2007) Immunology, 121 : 392-404. 2B6 comprises a VH having the amino acid sequence of SEQ ID NO: 200 and a VL having the amino acid sequence of SEQ ID NO: 205, as described in Table 1. Alternatively, the anti-CD32b antibody molecule of the combination comprises any one of Antibodies 20, 24, 26 and 28, having a VH and VL amino acid sequence of SEQ ID NO: 210 and 214, 218 and 222, 226 and 230, or 234 and 238, respectively as described in PCT Publication

WO2009/083009 (Van den Brink et al) and as described in Table 1. Alternatively, the anti-CD32b antibody of the combination comprises the antibody 6G11 as described in PCT Publication

WO2012/022985 (Cragg et al) and PCT Publication WO2015/173384 (Frendeus et al), having a VH amino acid sequence of SEQ ID NO: 242 and a VL amino acid sequence of SEQ ID NO: 246, as described in Table 1.

In one embodiment, the IL-15/IL-15Ra complex of the combination may comprise wild-type IL- 15 or an IL-15 derivative covalently or noncovalently bound to wild-type IL-15Ra or an IL-15Ra derivative. In one embodiment, the IL-15/IL-15Ra complex comprises wild-type IL-15 and IL-15Ra. In another embodiment, the IL-15/IL-15Ra complex comprises an IL-15 derivative and wild-type IL-15Ra. In another embodiment, the IL-15/IL-15Ra complex is in the wild-type heterodimeric form. In another embodiment, the IL-15 is human IL-15 and IL-15Ra is human IL-15Ra. In a specific embodiment, the human IL-15 comprises the amino acid sequence of SEQ ID NO: 251 or amino acid residues 49 to 162 of SEQ ID NO: 251 and the human IL-15Ra comprises the amino acid sequence of SEQ ID NO: 256 or a fragment thereof, as described in Table 1. In another embodiment the IL-15 comprises the amino acid sequence of SEQ ID NO: 251 or amino acid residues 49 to 162 of SEQ ID NO: 251 and the IL-15Ra comprises the amino acid sequence of SEQ ID NO: 257 or 260, as described in Table 1. In specific embodiments, the human IL-15 comprises amino acid residues 49 to 162 of the amino acid sequence of SEQ ID NO: 251 and human IL-15Ra comprises the amino acid sequence of SEQ ID NO: 260, as described in Table 1.

In other embodiments, the IL-15Ra is glycosylated such that glycosylation accounts for at least or more than 20%, 30%, 40% or 50% of the mass of the IL-15Ra. In another embodiment, the IL-15/IL- 15Ra complex comprises wild-type IL-15 and an IL-15Ra derivative. In another embodiment, the IL- 15/IL-15Ra complex comprises an IL-15 derivative and an IL-15Ra derivative. In one embodiment, the IL-15Ra derivative is a soluble form of the wild-type IL-15Ra. In another embodiment, the IL-15Ra derivative comprises a mutation that inhibits cleavage by an endogenous protease. In a specific embodiment, the extracellular domain cleavage site of IL-15Ra is replaced with a cleavage site that is specifically recognized by a heterologous protease. In one embodiment, the extracellular domain cleavage site of IL-15Ra is replaced with a heterologous extracellular domain cleavage site (e.g., heterologous transmembrane domain that is recognized and cleaved by another enzyme unrelated to the endogenous processing enzyme that cleaves the IL-15Ra).

In a specific embodiment, the present invention provides a combination comprising an anti- human CD32b antibody molecule in combination with an IL-15/IL-15Ra complex, wherein

i) the anti- human CD32b antibody molecule of the combination comprises a heavy chain variable region (VH) comprising a VHCDR1 amino acid sequence of SEQ ID NO: 113, a VHCDR2 amino acid sequence of SEQ ID NO: 114 and a VHCDR3 amino acid sequence of SEQ ID NO: 115, as described in Table 1 and a light chain variable region (VL) comprising a VLCDR1 amino acid sequence of SEQ ID NO: 120, a VLCDR2 amino acid sequence of SEQ ID NO: 121 and a VLCDR3 amino acid sequence of SEQ ID NO: 122, as described in Table 1; and

ii) the IL-15/IL-15Ra complex is a heterodimeric complex of human IL-15 and human soluble IL-15Ra and wherein the human IL-15 and comprises residues 49 to 162 of the amino acid sequence of SEQ ID NO: 251 and the human soluble IL-15Ra comprises the amino acid sequence of SEQ ID NO: 260, as described in Table 1.

Uses of the Combination Therapies The combinations disclosed herein can result in one or more of: an increase in antigen presentation, an increase in effector cell function (e.g., one or more of T cell proliferation, IFN-a secretion or cytolytic function), inhibition of regulatory T cell function, an effect on the activity of multiple cell types, such as regulatory T cell, effector T cells and NK cells), an increase in tumor infiltrating lymphocytes and an increase in T-cell receptor mediated proliferation. In one embodiment, the use of an anti-CD32b antibody molecule in the combination inhibits, reduces or neutralizes one or more activities of CD32b, resulting in enhanced effector function/ADCC. In one embodiment, the use of an IL-15/IL-15Ra complex in the combination stimulates the immune response and can enhance ADCC resulting from the use of an anti-CD32b antibody molecule. Thus, such combinations can be used to treat or prevent disorders where enhancing an immune response in a subject is desired, e.g. cancer. Such combination therapies can be used, e.g., for cancer immunotherapy and treatment of other conditions, such as chronic infection. In an embodiment, provided herein are methods of treating (e.g., inhibiting, reducing, ameliorating, or preventing) a disorder, e.g., a hyperproliferative condition or disorder (e.g., a cancer) in a subject by administering to the subject an anti-CD32b antibody molecule in combination with an IL-15/IL-15Ra complex. Also provided is an anti-CD32b antibody molecule in combination with an IL-15/IL-15Ra complex for use in the treatment of (e.g., inhibiting, reducing, ameliorating, or preventing) a disorder, e.g., a hyperproliferative condition or disorder (e.g., a cancer) in a subject. Further provided is an anti-CD32b antibody molecule in combination with an IL-15/IL-15Ra complex for use in the preparation of a medicament for the treatment of (e.g., inhibiting, reducing, ameliorating, or preventing) a disorder, e.g., a hyperproliferative condition or disorder (e.g., a cancer) in a subject.

The augmentation of the ADCC activity of an anti-CD32b antibody molecule by an IL-15/IL- 15Ra complex has been demonstrated in Example 1. In a cytotoxicity assay to measure target cell lysis, an anti-CD32b antibody of the present invention showed dose dependent ADCC activity against effector (Daudi) cells. The addition of an IL-15/IL-15Ra complex enhanced the potency of the ADCC activity. In a specific embodiment, the present invention provides a method of enhancing the ADCC activity of an anti-CD32b antibody comprising administering an effective amount of an IL-15/IL-15Ra complex in combination with an anti-CD32b antibody.

In some embodiments, the disorder is selected from B cell malignancies, Hodgkins lymphoma, Non-Hodgkins lymphoma, multiple myeloma, diffuse large B cell lymphoma, acute lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse small cleaved cell lymphoma, MALT lymphoma, mantel cell lymphoma, marginal zone lymphoma, follicular lymphoma, systemic light chain amyloidosis or haematological malignancies. In a specific embodiment, the disorder is selected from B-cell malignancies, non-Hodgkins lymphoma multiple myeloma or chronic lymphocytic leukemia.

In one embodiment, the combination of anti-CD32b antibody molecule and IL-15/IL-15Ra complex are administered to a subject separately or together. In another embodiment, the combination of anti-CD32b antibody molecule and IL-15/IL-15Ra complex are administered simultaneously or sequentially.

The present application also provides nucleic acids encoding the anti-CD32b antibody molecule and/or the IL-15/IL-15Ra complex disclosed herein, as well as a vector comprising the nucleic acid, and a host cell comprising the nucleic acid or the vector. Also provided are methods of producing the anti-CD32b antibody molecule and/or the IL-15/IL-15Ra complex disclosed herein, the method comprising: culturing a host cell expressing a nucleic acid encoding the antibody molecule or complex; and collecting the antibody molecule or complex from the culture.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 shows that ADCC mediated by the anti-CD32b antibody NOV2108 against Daudi cells is enhanced by the IL-15/IL-15Ra complex, hetIL-15. ADCC assays were performed using primary NK cells as effector cells in the absence (grey lines) or presence (black lines) of hetIL-15. Luciferized Daudi cells as target cells were incubated with primary NK cells and antibody NOV2108 in different formats ((A) wild type, (B) N297A Fc silent, (C) afucosylated or (D) an IgG control), in serial dilution for 4 hr. Effector: target ratio (E:T) = 1. Luciferase signal was then measured to determine % lytic activity. The experiment was repeated with two different donor NK cells with the representative data from one experiment is shown.

DETAILED DESCRIPTION OF THE INVENTION

ADCC is a mechanism of immune defense whereby an effector cell (NK cells, macrophages, neutrophils) actively lyse a target cell whose cell surface antigens are bound by specific antibodies engaged with Fc Receptors on the immune cell. Many anti-tumor antibodies have been developed into therapeutics for cancer and an important mechanism of their anti-tumor activity is ADCC. CD32b is expressed on various B cell and plasma cell malignancies, and anti-CD32b antibodies can be used to target the malignant cells. A complex of IL-15 with IL-15Ra (IL-15/IL-15Ra), potently activates NK cells, resulting in proliferation of NK cells as well as enhanced target cell lysis. The present invention provides a combination comprising an antibody molecule that specifically binds to human CD32b protein with an IL-15/IL-15Ra complex, and pharmaceutical compositions, production methods, and methods of use of such a combination.

Definitions

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which this invention pertains.

By "in combination with" or " a combination of, it is not intended to imply that the agents of the combination must be administered at the same time and/or formulated for delivery together, although these methods of delivery are within the scope described herein. The anti-CD32b antibody molecules can be administered prior to, concurrently with, or subsequent to, the IL-15/IL-15Ra complex and vice versa, i.e. the anti-CD32b antibody molecule and the IL-15/IL-15Ra complex can be administered in any order. In general, each agent will be administered at a dose and/or on a time schedule determined for that agent. It will further be appreciated that each therapeutic agent utilized in this combination may be administered together in a single composition or administered separately in different compositions. In general, it is expected that the therapeutic agents of the combination be utilized at levels that do not exceed the levels at which they are utilized individually. In some embodiments, the levels of the agents utilized in combination will be lower than those utilized individually. In some embodiments, the agents of the combination may also be used as entirely separate pharmaceutical dosage forms or pharmaceutical formulations that are also sold independently of each other and where instructions of the possibility of their combined use is or are provided in the package equipment, e.g. leaflet or the like, or in other information e.g. provided to physicians and medical staff.

"CD32A" or "CD32a", as used herein, means human CD32a protein, also referred to as human FCy Receptor 2A or FCyR2A or FCGR2a or FCGR2A. There are two variants known as H131 and R131 (when referenced without the signal sequence) or HI 67 and R167 (when referenced with the signal sequence). The amino acid sequence of the H167 variant is deposited under accession number UniProtKB P12318 and is detailed below:

MTMETQMSQNVCPRNLWLLQPLTVLLLLASADSQAAAPPKAVLKLEPPWINVLQEDSVTLTCQGARSPES DSIQWFHNGNLIPTHTQPSYRFKANNNDSGEYTCQTGQTSLSDPVHLTVLSEWLVLQTPHLEFQEGETIM LRCHSWKDKPLVKVTFFQNGKSQKFSHLDPTFSIPQA HSHSGDYHCTGNIGYTLFSSKPVTITVQVPSM GSSSPMGIIVAVVIATAVAAIVAAVVALIYCRKKRISANSTDPVKAAQFEPPGRQMIAIRKRQLEETNND YETADGGYMTLNPRAPTDDDKNIYLTLPPNDHVNSNN (SEQ ID NO: 247 in Table 1).

"CD32B" or "CD32b", as used herein, means human CD32b protein, also referred to as human FCy Receptor 2B or FCyR2B or FCGR2b or FCGR2B. The amino acid sequence for CD32b variant 1 is deposited under accession number UniProtKB P31994-1 and is detailed below:

MGILSFLPVLATESDWADCKSPQPWGHMLLWTAVLFLAPVAGTPAAPPKAVLKLEPQWINVLQEDSVTLT CRGTHSPESDSIQWFHNGNLIPTHTQPSYRFKA NNDSGEYTCQTGQTSLSDPVHLTVLSEWLVLQTPHL EFQEGETIVLRCHSWKDKPLVKVTFFQNGKSKKFSRSDPNFSIPQA HSHSGDYHCTGNIGYTLYSSKPV TITVQAPSSSPMGIIVAVVTGIAVAAIVAAVVALIYCRKKRISALPGYPECREMGETLPEKPANPTNPDE ADKVGAEN ITYSLLMHPDALEEPDDQNRI (SEQ ID NO: 248 in Table 1).

The amino acid sequence for CD32b variant 2 is deposited under accession number UniProtKB P31994-2 and is detailed below:

MGILSFLPVLATESDWADCKSPQPWGHMLLWTAVLFLAPVAGTPAAPPKAVLKLEPQWINVLQEDSVTLT CRGTHSPESDSIQWFHNGNLIPTHTQPSYRFKA NNDSGEYTCQTGQTSLSDPVHLTVLSEWLVLQTPHL EFQEGETIVLRCHSWKDKPLVKVTFFQNGKSKKFSRSDPNFSIPQA HSHSGDYHCTGNIGYTLYSSKPV TITVQAPSSSPMGIIVAVVTGIAVAAIVAAVVALIYCRKKRISANPTNPDEADKVGAENTITYSLLMHPD ALEEPDDQNRI (SEQ ID NO: 249 in Table 1).

As described herein, an antibody molecule which binds to CD32b binds to human CD32b protein or a fragment thereof. As used herein "huCD32b" refers to human CD32b protein or a fragment thereof. The term "antibody molecule" and the like, as used herein, include whole antibodies and any antigen-binding fragment (i.e., "antigen-binding portion") or single chains thereof. A naturally occurring "antibody molecule" is a glycoprotein comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds. Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region is comprised of three domains, CHI, CH2 and CH3. Each light chain is comprised of a light chain variable region (abbreviated herein as VL) and a light chain constant region. The light chain constant region is comprised of one domain, CL. The VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each VH and VL is composed of three CDRs and four FRs arranged from amino-terminus to carboxy-terminus in the following order: FRl, CDRl, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen. The constant regions of the antibodies may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system. The precise amino acid sequence boundaries of a given CDR can be determined using any of a number of well-known schemes, including those described by Kabat et al., (1991) "Sequences of Proteins of Immunological Interest," 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD ("Kabat" numbering scheme), Al- Lazikani et al, (1997) JMB 273, 927-948 ("Chothia" numbering scheme) and ImMunoGenTics (IMGT) numbering (Lefranc, M.-P., (1999) The Immunologist, 7: 132-136; Lefranc, M.-P. et al, (2003) Dev. Comp. Immunol. 27: 55-77 ("IMGT" numbering scheme). For example, for classic formats, under Kabat, the CDR amino acid residues in the heavy chain variable domain (VH) are numbered 31-35 (HCDRl), 50-65 (HCDR2), and 95-102 (HCDR3); and the CDR amino acid residues in the light chain variable domain (VL) are numbered 24-34 (LCDR1), 50-56 (LCDR2), and 89-97 (LCDR3). Under Chothia the CDR amino acids in the VH are numbered 26-32 (HCDRl), 52-56 (HCDR2), and 95-102 (HCDR3); and the amino acid residues in VL are numbered 26-32 (LCDR1), 50-52 (LCDR2), and 91-96 (LCDR3). By combining the CDR definitions of both Kabat and Chothia, the CDRs consist of amino acid residues 26- 35 (HCDRl), 50-65 (HCDR2), and 95-102 (HCDR3) in human VH and amino acid residues 24-34 (LCDR1), 50-56 (LCDR2), and 89-97 (LCDR3) in human VL. Under IMGT the CDR amino acid residues in the VH are numbered approximately 26-35 (CDRl), 51-57 (CDR2) and 93-102 (CDR3), and the CDR amino acid residues in the VL are numbered approximately 27-32 (CDRl), 50-52 (CDR2), and 89-97 (CDR3) (numbering according to "Kabat"). Under IMGT, the CDR regions of an antibody can be determined using the program IMGT/DomainGap Align.

The terms "antigen-binding fragment", "antigen-binding fragment thereof," "antigen binding portion" of an antibody, and the like, as used herein, refer to one or more fragments of an intact antibody that retain the ability to specifically bind to a given antigen (e.g., CD32b). Antigen binding functions of an antibody can be performed by antibody fragments. Examples of antigen binding fragments encompassed within the term "antigen binding portion" of an antibody include a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CHI domains; a F (ab)2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; an Fd fragment consisting of the VH and CHI domains; an Fv fragment consisting of the VL and VH domains of a single arm of an antibody; a single domain antibody (dAb) fragment (Ward et al., (1989) Nature 341 :544-546), which consists of a VH domain; and an isolated CDR.

Furthermore, although the two domains of the Fv fragment, VL and VH, are coded for by separate genes, they can be joined, using recombinant methods, by an artificial peptide linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv (scFv); see, e.g., Bird et al., (1988) Science 242:423-426; and Huston et al., (1988) PNAS USA. 85:5879-5883). Such single chain antibodies include one or more "antigen binding fragments" of an antibody. These antibody fragments are obtained using conventional techniques known to those of skill in the art, and the fragments are screened for utility in the same manner as are intact antibodies.

Antigen binding fragments can also be incorporated into single domain antibodies, maxibodies, minibodies, intrabodies, diabodies, triabodies, tetrabodies, v-NAR and bis-scFv (see, e.g., Hollinger & Hudson (2005), Nature Biotechnology, 23 (9): 1126-1136). Antigen binding fragments of antibodies can be grafted into scaffolds based on polypeptides such as Fibronectin type III (Fn3) (see U.S. Pat. No. 6,703, 199, which describes fibronectin polypeptide monobodies).

Antigen binding fragments can be incorporated into single chain molecules comprising a pair of tandem Fv segments (VH-CH1-VH-CH1) which, together with complementary light chain polypeptides, form a pair of antigen binding regions (Zapata et al, (1995) Protein Eng. 8 (10): 1057-1062; and U.S. Pat. No. 5,641,870).

As used herein, the term "Affinity" refers to the strength of interaction between antibody and antigen at single antigenic sites. Within each antigenic site, the variable region of the antibody "arm" interacts through weak non-covalent forces with antigen at numerous sites; the more interactions, the stronger the affinity.

The term "amino acid" refers to wild-type and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the wild-type amino acids. Wild- type amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, e.g., hydroxyproline, gamma-carboxyglutamate, and O-phosphoserine. Amino acid analogs refer to compounds that have the same basic chemical structure as a wild-type amino acid, i.e., an alpha carbon that is bound to a hydrogen, a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as a wild- type amino acid. Amino acid mimetics refers to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a wild-type amino acid.

The term "binding specificity" as used herein refers to the ability of an individual antibody combining site to react with one antigenic determinant and not with a different antigenic determinant. The combining site of the antibody is located in the Fab portion of the molecule and is constructed from the hypervariable regions of the heavy and light chains. Binding affinity of an antibody is the strength of the reaction between a single antigenic determinant and a single combining site on the antibody. It is the sum of the attractive and repulsive forces operating between the antigenic determinant and the combining site of the antibody.

Specific binding between two entities means a binding with an equilibrium constant (KA or K_A) of at least 1 x 10⁷ M^"1, 10⁸ M^"1, 10⁹ M^"1, 10¹⁰ M^"1, 10¹¹ M^"1, 10¹² M^"1, 10¹³ M^"1, or 10¹⁴ M^"1. The phrase "specifically (or selectively) binds" to an antigen (e.g., CD32b-binding antibody) refers to a binding reaction that is determinative of the presence of a cognate antigen (e.g., a human CD32b protein) in a heterogeneous population of proteins and other biologies. A CD32b-binding antibody of the invention binds to CD32b with a greater affinity than it does to a non-specific antigen (e.g., CD32a). The phrases "an antibody recognizing an antigen" and "an antibody specific for an antigen" are used interchangeably herein with the term "an antibody which binds specifically to an antigen".

The term "conservatively modified variant" applies to both amino acid and nucleic acid sequences. With respect to particular nucleic acid sequences, conservatively modified variants refers to those nucleic acids which encode identical or essentially identical amino acid sequences, or where the nucleic acid does not encode an amino acid sequence, to essentially identical sequences. Because of the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For instance, the codons GCA, GCC, GCG and GCU all encode the amino acid alanine. Thus, at every position where an alanine is specified by a codon, the codon can be altered to any of the corresponding codons described without altering the encoded polypeptide. Such nucleic acid variations are "silent variations," which are one species of conservatively modified variations. Every nucleic acid sequence herein which encodes a polypeptide also describes every possible silent variation of the nucleic acid. One of skill will recognize that each codon in a nucleic acid (except AUG, which is ordinarily the only codon for methionine, and TGG, which is ordinarily the only codon for tryptophan) can be modified to yield a functionally identical molecule. Accordingly, each silent variation of a nucleic acid that encodes a polypeptide is implicit in each described sequence.

For polypeptide sequences, "conservatively modified variants" include individual substitutions, deletions or additions to a polypeptide sequence which result in the substitution of an amino acid with a chemically similar amino acid. Conservative substitution tables providing functionally similar amino acids are well known in the art. Such conservatively modified variants are in addition to and do not exclude polymorphic variants, interspecies homologs, and alleles of the invention. The following eight groups contain amino acids that are conservative substitutions for one another: 1) Alanine (A), Glycine

(G); 2) Aspartic acid (D), Glutamic acid (E); 3) Asparagine (N), Glutamine (Q); 4) Arginine (R), Lysine (K); 5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V); 6) Phenylalanine (F), Tyrosine (Y), Tryptophan (W); 7) Serine (S), Threonine (T); and 8) Cysteine (C), Methionine (M) (see, e.g., Creighton, Proteins (1984)). In one embodiment, the term "conservative sequence modifications" are used to refer to amino acid modifications that do not significantly affect or alter the binding characteristics of the antibody containing the amino acid sequence.

The term "recognize" as used herein refers to an antibody antigen-binding fragment thereof that finds and interacts (e.g., binds) with its conformational epitope.

The term "blocks" as used herein refers to reducing an interaction or a process, e.g., stopping ligand-dependent or ligand-independent signaling. For example, the interaction or process can be reduced by up to 50%, up to 60%, up to 70%, up to 80%, up to 90% or up to 100%. The terms "cross-block", "cross-blocked", "cross-blocking", "compete", "cross compete" and related terms are used

interchangeably herein to mean the ability of an antibody or other binding agent to interfere with the binding of other antibodies or binding agents to CD32b in a standard competitive binding assay. The ability or extent to which an antibody or other binding agent is able to interfere with the binding of another antibody or binding molecule to CD32b, and therefore whether it can be said to cross-block according to the invention, can be determined using standard competition binding assays. One suitable assay involves the use of the BIAcore^® technology (e.g. by using the BIAcore 3000 instrument (Biacore, Uppsala, Sweden)), which can measure the extent of interactions using surface plasmon resonance technology. Another assay for measuring cross-blocking uses an ELISA-based approach. Although the techniques are expected to produce substantially similar results, measurement by the Biacore technique is considered definitive.

The term "neutralizes" means that an antibody, upon binding to its target, reduces the activity, level or stability of the target; e.g., a CD32b antibody, upon binding to CD32b neutralizes CD32b by at least partially reducing an activity, level or stability of CD32b, such as its role in engaging Fc portions of antibodies.

The term "epitope" means a protein determinant capable of specific binding to an antibody. Epitopes usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and usually have specific three dimensional structural characteristics, as well as specific charge characteristics. Conformational and nonconformational epitopes are distinguished in that the binding to the former but not the latter is lost in the presence of denaturing solvents. The term "epitope" includes any protein determinant capable of specific binding to an immunoglobulin or otherwise interacting with a molecule. Epitopic determinants generally consist of chemically active surface groupings of molecules such as amino acids or carbohydrate or sugar side chains and can have specific three-dimensional structural characteristics, as well as specific charge characteristics. An epitope may be "linear" or "conformational". The term "linear epitope" refers to an epitope with all of the points of interaction between the protein and the interacting molecule (such as an antibody) occurring linearly and/or continuously along the primary amino acid sequence of the protein.

As used herein, the term "high affinity" for an IgG antibody refers to an antibody having a KD of 10^"8 M or less, 10^"9 M or less, or 10^"10 M, or 10^"11 M or less for a target antigen, e.g., CD32b. However, "high affinity" binding can vary for other antibody isotypes. For example, "high affinity" binding for an IgM isotype refers to an antibody having a KD of 10^"7 M or less, or 10^"8 M or less.

The term "human antibody" (or antigen-binding fragment thereof), as used herein, is intended to include antibodies (and antigen-binding fragments thereof) having variable regions in which both the framework and CDR regions are derived from sequences of human origin. Furthermore, if the antibody contains a constant region, the constant region also is derived from such human sequences, e.g., human germline sequences, or mutated versions of human germline sequences. The human antibodies and antigen-binding fragments thereof of the invention may include amino acid residues not encoded by human sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo).

The phrases "monoclonal antibody" or "monoclonal antibody composition" (or antigen-binding fragment thereof) as used herein refers to polypeptides, including antibodies, antibody fragments, bispecific antibodies, etc. that have substantially identical to amino acid sequence or are derived from the same genetic source. This term also includes preparations of antibody molecules of single molecular composition. A monoclonal antibody composition displays a single binding specificity and affinity for a particular epitope.

The term "human monoclonal antibody" (or antigen-binding fragment thereof) refers to antibodies (and antigen-binding fragments thereof) displaying a single binding specificity which have variable regions in which both the framework and CDR regions are derived from human sequences. In one embodiment, the human monoclonal antibodies are produced by a hybridoma which includes a B cell obtained from a transgenic nonhuman animal, e.g., a transgenic mouse, having a genome comprising a human heavy chain transgene and a light chain transgene fused to an immortalized cell.

The phrase "recombinant human antibody" (or antigen-binding fragment thereof), as used herein, includes all human antibodies (and antigen-binding fragments thereof) that are prepared, expressed, created or isolated by recombinant means, such as antibodies isolated from an animal (e.g., a mouse) that is transgenic or transchromosomal for human immunoglobulin genes or a hybridoma prepared therefrom, antibodies isolated from a host cell transformed to express the human antibody, e.g., from a transfectoma, antibodies isolated from a recombinant, combinatorial human antibody library, and antibodies prepared, expressed, created or isolated by any other means that involve splicing of all or a portion of a human immunoglobulin gene, sequences to other DNA sequences. Such recombinant human antibodies have variable regions in which the framework and CDR regions are derived from human germline

immunoglobulin sequences. In one embodiment, such recombinant human antibodies can be subjected to in vitro mutagenesis (or, when an animal transgenic for human Ig sequences is used, in vivo somatic mutagenesis) and thus the amino acid sequences of the VH and VL regions of the recombinant antibodies are sequences that, while derived from and related to human germline VH and VL sequences, may not naturally exist within the human antibody germline repertoire in vivo.

A "humanized antibody" (or antigen-binding fragment thereof), as used herein, is an antibody molecule that retains the reactivity of a non-human antibody while being less immunogenic in humans. This can be achieved, for instance, by retaining the non-human CDR regions and replacing the remaining parts of the antibody with their human counterparts (i.e., the constant region as well as the framework portions of the variable region). See, e.g., Morrison et al, (1984) PNAS. USA, 81 :6851-6855; Morrison & Oi, (1988) Adv. Immunol., 44:65-92; Verhoeyen et al, (1988) Science, 239: 1534-1536; Padlan, (1991) Molec. Immun, 28:489-498 and Padlan, (1994) Molec. Immun., 31 : 169-217. Other examples of human engineering technology include, but is not limited to, Xoma technology disclosed in U.S. Pat. No.

5,766,886.

The term "chimeric antibody" (or antigen-binding fragment thereof) is an antibody molecule in which (a) the constant region, or a portion thereof, is altered, replaced or exchanged so that the antigen binding site (variable region) is linked to a constant region of a different or altered class, effector function and/or species, or an entirely different molecule which confers new properties to the chimeric antibody, e.g., an enzyme, toxin, hormone, growth factor, drug, etc.; or (b) the variable region, or a portion thereof, is altered, replaced or exchanged with a variable region having a different or altered antigen specificity. For example, a mouse antibody can be modified by replacing its constant region with the constant region from a human immunoglobulin. Due to the replacement with a human constant region, the chimeric antibody can retain its specificity in recognizing the antigen while having reduced antigenicity in human as compared to the original mouse antibody.

The terms "identical" or percent "identity," in the context of two or more nucleic acids or polypeptide sequences, refer to two or more sequences or subsequences that are the same. Two sequences are "substantially identical" if two sequences have a specified percentage of amino acid residues or nucleotides that are the same (i.e., 60% identity, optionally 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identity over a specified region, or, when not specified, over the entire sequence), when compared and aligned for maximum correspondence over a comparison window, or designated region as measured using one of the following sequence comparison algorithms or by manual alignment and visual inspection. Optionally, the identity exists over a region that is at least about 50 nucleotides (or 10 amino acids) in length, or more preferably over a region that is 100 to 500 or 1000 or more nucleotides (or 20, 50, 200 or more amino acids) in length. Optionally, the identity exists over a region that is at least 50 nucleotides (or 10 amino acids) in length, or more preferably over a region that is 100 to 500 or 1000 or more nucleotides (or 20, 50, 200 or more amino acids) in length.

For sequence comparison, typically one sequence acts as a reference sequence, to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are entered into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. Default program parameters can be used, or alternative parameters can be designated. The sequence comparison algorithm then calculates the percent sequence identities for the test sequences relative to the reference sequence, based on the program parameters.

A "comparison window", as used herein, includes reference to a segment of any one of the number of contiguous positions selected from the group consisting of from 20 to 600, usually about 50 to about 200, more usually about 100 to about 150 in which a sequence may be compared to a reference sequence of the same number of contiguous positions after the two sequences are optimally aligned. Methods of alignment of sequences for comparison are well known in the art. Optimal alignment of sequences for comparison can be conducted, e.g., by the local homology algorithm of Smith & Waterman (1970) Adv. Appl. Math. 2:482c, by the homology alignment algorithm of Needleman & Wunsch, (1970) J. Mol. Biol. 48:443, by the search for similarity method of Pearson & Lipman, (1988) PNAS USA 85:2444, by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, Wis.), or by manual alignment and visual inspection (see, e.g., Brent et al., Current Protocols in

Molecular Biology, John Wiley & Sons, Inc. (ringbou ed., 2003)).

Two examples of algorithms that are suitable for determining percent sequence identity and sequence similarity are the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al., (1977) Nuc. Acids Res. 25:3389-3402; and Altschul et al, (1990) J. Mol. Biol. 215:403-410, respectively. Software for performing BLAST analyses is publicly available through the National Center for

Biotechnology Information. This algorithm involves first identifying high scoring sequence pairs (HSPs) by identifying short words of length W in the query sequence, which either match or satisfy some positive-valued threshold score T when aligned with a word of the same length in a database sequence. T is referred to as the neighborhood word score threshold (Altschul et al., supra). These initial

neighborhood word hits act as seeds for initiating searches to find longer HSPs containing them. The word hits are extended in both directions along each sequence for as far as the cumulative alignment score can be increased. Cumulative scores are calculated using, for nucleotide sequences, the parameters M (reward score for a pair of matching residues; always >0) and N (penalty score for mismatching residues; always <0). For amino acid sequences, a scoring matrix is used to calculate the cumulative score.

Extension of the word hits in each direction are halted when: the cumulative alignment score falls off by the quantity X from its maximum achieved value; the cumulative score goes to zero or below, due to the accumulation of one or more negative -scoring residue alignments; or the end of either sequence is reached. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses as defaults a word length (N) of 11, an expectation (E) or 10, M=5, N=-4 and a comparison of both strands. For amino acid sequences, the BLASTP program uses as defaults a word length of 3, and expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff & Henikoff, (1989) PNAS USA 89: 10915) alignments (B) of 50, expectation (E) of 10, M=5, N=-4, and a comparison of both strands.

The BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin & Altschul, (1993) PNAS USA 90:5873-5787). One measure of similarity provided by the BLAST algorithm is the smallest sum probability (P (N)), which provides an indication of the probability by which a match between two nucleotide or amino acid sequences would occur by chance. For example, a nucleic acid is considered similar to a reference sequence if the smallest sum probability in a comparison of the test nucleic acid to the reference nucleic acid is less than about 0.2, more preferably less than about 0.01, and most preferably less than about 0.001.

The percent identity between two amino acid sequences can also be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., (1988) 4: 11-17) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. In addition, the percent identity between two amino acid sequences can be determined using the Needleman & Wunsch (J. Mol, Biol. (1970) 48:444-453) algorithm which has been incorporated into the GAP program in the GCG software package (available at www.gcg.com), using either a Blossom 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6.

Other than percentage of sequence identity noted above, another indication that two nucleic acid sequences or polypeptides are substantially identical is that the polypeptide encoded by the first nucleic acid is immunologically cross-reactive with the antibodies raised against the polypeptide encoded by the second nucleic acid, as described below. Thus, a polypeptide is typically substantially identical to a second polypeptide, for example, where the two peptides differ only by conservative substitutions.

Another indication that two nucleic acid sequences are substantially identical is that the two molecules or their complements hybridize to each other under stringent conditions, as described below. Yet another indication that two nucleic acid sequences are substantially identical is that the same primers can be used to amplify the sequence.

As used herein, the term "hybridizes under low stringency, medium stringency, high stringency, or very high stringency conditions" describes conditions for hybridization and washing. Guidance for performing hybridization reactions can be found in Current Protocols in Molecular Biology, John Wiley & Sons, N.Y. (1989), 6.3.1-6.3.6. Aqueous and nonaqueous methods are described in that reference and either can be used. Specific hybridization conditions referred to herein are as follows: 1) low stringency hybridization conditions in 6X sodium chloride/sodium citrate (SSC) at about 45°C, followed by two washes in 0.2X SSC, 0.1% SDS at least at 50°C (the temperature of the washes can be increased to 55°C for low stringency conditions); 2) medium stringency hybridization conditions in 6X SSC at about 45°C, followed by one or more washes in 0.2X SSC, 0.1% SDS at 60°C; 3) high stringency hybridization conditions in 6X SSC at about 45°C, followed by one or more washes in 0.2X SSC, 0.1% SDS at 65°C; and preferably 4) very high stringency hybridization conditions are 0.5M sodium phosphate, 7% SDS at 65°C, followed by one or more washes at 0.2X SSC, 1% SDS at 65°C. Very high stringency conditions (4) are the preferred conditions and the ones that should be used unless otherwise specified.

The term "isolated antibody" (or antigen-binding fragment thereof), as used herein, refers to an antibody molecule that is substantially free of other antibodies having different antigenic specificities (e.g., an isolated antibody that specifically binds CD32b is substantially free of antibodies that specifically bind antigens other than CD32b). Moreover, an isolated antibody may be substantially free of other cellular material and/or chemicals.

The term "isotype" refers to the antibody class (e.g., IgM, IgE, IgG such as IgGl or IgG4) that is provided by the heavy chain constant region genes. Isotype also includes modified versions of one of these classes, where modifications have been made to after the Fc function, for example, to enhance or reduce effector functions or binding to Fc receptors.

The term "Kassoc", "Ka" or "K_on", as used herein, is intended to refer to the association rate of a particular antibody-antigen interaction, whereas the term "Kdis", "Kd," or "K_off", as used herein, is intended to refer to the dissociation rate of a particular antibody-antigen interaction. In one embodiment, the term "KD", as used herein, is intended to refer to the dissociation constant, which is obtained from the ratio of Kd to Ka (i.e. Kd/Ka) and is expressed as a molar concentration (M). KD values for antibodies can be determined using methods well established in the art. A method for determining the KD of an antibody is by using surface plasmon resonance, or using a biosensor system such as a Biacore^® system. Where the dissociation constant is less than about 10^"9 M, solution equilibrium kinetic exclusion KD measurement (MSD-SET) is a preferred method for determining the KD of an antibody (see, e.g. Friquet et al., (1985) J Immnunol. Meth. 77, 305-319; herein incorporated by reference).

The term "IC₅₀" as used herein, refers to the concentration of an antibody or an antigen-binding fragment thereof, which induces an inhibitory response, either in an in vitro or an in vivo assay, which is 50% of the maximal response, i.e., halfway between the maximal response and the baseline.

The term "effector function" refers to an activity of an antibody molecule that is mediated by binding through a domain of the antibody other than the antigen-binding domain, usually mediated by binding of effector molecules. Effector function includes complement-mediated effector function, which is mediated by, for example, binding of the CI component of the complement to the antibody. Activation of complement is important in the opsonisation and lysis of cell pathogens. The activation of complement also stimulates the inflammatory response and may also be involved in autoimmune hypersensitivity. Effector function also includes Fc receptor (FcR)-mediated effector function, which may be triggered upon binding of the constant domain of an antibody to an Fc receptor (FcR). Binding of antibody to Fc receptors on cell surfaces triggers a number of important and diverse biological responses including engulfment and destruction of antibody-coated particles, clearance of immune complexes, release of inflammatory mediators, placental transfer, control of immunoglobulin production and lysis of antibody- coated target cells by killer cells. The latter response, whereby an effector cell of the immune system (e.g.

NK cells, neutrophils and macrophages) actively lyses a target cell, whose membrane -surface antigens have been bound by specific antibodies, is referred to as antibody-dependent cell-mediated cytotoxicity or "ADCC". This is a mechanism of cell-mediated immune defense and is one of the mechanisms through which antibodies, as part of the humoral immune response, can act to limit and contain infection. ADCC is independent of the immune complement system that also lyses target cells but does not require any other cell. The primary cells for mediating ADCC, NK cells, express FcyRIII only, whereas monocytes express FcyRI, FcyRII and FcyRIII. Macrophages, neutrophils and eosinophils can also mediate ADCC (see Ravetch & Kinet (1991) Ann. Rev. Immunol., 9: 457- 492). ADCC is part of the adaptive immune response due to its dependence on a prior antibody response.

An effector function of an antibody may be altered by altering, e.g., enhancing or reducing, the affinity of the antibody for an effector molecule such as an Fc receptor or a complement component. Binding affinity will generally be varied by modifying the effector molecule binding site, and in this case it is appropriate to locate the site of interest and modify at least part of the site in a suitable way. It is also envisaged that an alteration in the binding site on the antibody for the effector molecule need not alter significantly the overall binding affinity but may alter the geometry of the interaction rendering the effector mechanism ineffective as in non-productive binding. It is further envisaged that an effector function may also be altered by modifying a site not directly involved in effector molecule binding, but otherwise involved in performance of the effector function.

The term "nucleic acid" is used herein interchangeably with the term "polynucleotide" and refers to deoxyribonucleotides or ribonucleotides and polymers thereof in either single- or double-stranded form. The term encompasses nucleic acids containing known nucleotide analogs or modified backbone residues or linkages, which are synthetic, wild-type, and non-wild-type, which have similar binding properties as the reference nucleic acid, and which are metabolized in a manner similar to the reference nucleotides. Examples of such analogs include, without limitation, phosphorothioates, phosphoramidates, methyl phosphonates, chiral-methyl phosphonates, 2-O-methyl ribonucleotides, peptide-nucleic acids (PNAs).

Unless otherwise indicated, a particular nucleic acid sequence also implicitly encompasses conservatively modified variants thereof (e.g., degenerate codon substitutions) and complementary sequences, as well as the sequence explicitly indicated. Specifically, as detailed below, degenerate codon substitutions may be achieved by generating sequences in which the third position of one or more selected (or all) codons is substituted with mixed-base and/or deoxyinosine residues (Batzer et al, (1991) Nucleic Acid Res. 19:5081; Ohtsuka ei /., (1985) J. Biol. Chem. 260:2605-2608; and Rossolini et al, (1994) Mol. Cell. Probes 8:91-98).

The term "operably linked" refers to a functional relationship between two or more

polynucleotide (e.g., DNA) segments. Typically, it refers to the functional relationship of a transcriptional regulatory sequence to a transcribed sequence. For example, a promoter or enhancer sequence is operably linked to a coding sequence if it stimulates or modulates the transcription of the coding sequence in an appropriate host cell or other expression system. Generally, promoter transcriptional regulatory sequences that are operably linked to a transcribed sequence are physically contiguous to the transcribed sequence, i.e., they are cis-acting. However, some transcriptional regulatory sequences, such as enhancers, need not be physically contiguous or located in close proximity to the coding sequences whose transcription they enhance.

As used herein, the term, "optimized" means that a nucleotide sequence has been altered to encode an amino acid sequence using codons that are preferred in the production cell or organism, generally a eukaryotic cell, for example, a cell of Pichia, a Chinese Hamster Ovary cell (CHO) or a human cell. The optimized nucleotide sequence is engineered to retain completely or as much as possible the amino acid sequence originally encoded by the starting nucleotide sequence, which is also known as the "parental" sequence. The optimized sequences herein have been engineered to have codons that are preferred in mammalian cells. However, optimized expression of these sequences in other eukaryotic cells or prokaryotic cells is also envisioned herein. The amino acid sequences encoded by optimized nucleotide sequences are also referred to as optimized.

The terms "polypeptide" and "protein" are used interchangeably herein to refer to a polymer of amino acid residues. The terms apply to amino acid polymers in which one or more amino acid residue is an artificial chemical mimetic of a corresponding wild-type amino acid, as well as to wild-type amino acid polymers and non-wild-type amino acid polymer. Unless otherwise indicated, a particular polypeptide sequence also implicitly encompasses conservatively modified variants thereof.

The term "recombinant host cell" (or simply "host cell") refers to a cell into which a recombinant expression vector has been introduced. It should be understood that such terms are intended to refer not only to the particular subject cell but to the progeny of such a cell. Because certain modifications may occur in succeeding generations due to either mutation or environmental influences, such progeny may not, in fact, be identical to the parent cell, but are still included within the scope of the term "host cell" as used herein.

The term "subject" includes human and non-human animals. Non-human animals include all vertebrates, e.g., mammals and non-mammals, such as non-human primates, sheep, dog, cow, chickens, amphibians, and reptiles. Except when noted, the terms "patient" or "subject" are used herein

interchangeably.

The terms "treat," "treated," "treating," and "treatment," include the administration of compositions or antibodies to alleviate or delay the onset of the symptoms, complications, or biochemical indicia of a disease, preventing the development of further symptoms or arresting or inhibiting further development of the disease, condition, or disorder. Treatment can be measured by the therapeutic measures described herein. The methods of "treatment" of the present invention include administration of a CD32b antibody molecule to a subject in order to cure, reduce the severity of, or ameliorate one or more symptoms of cancer or condition associated with cancer, in order to prolong the health or survival of a subject beyond that expected in the absence of such treatment. For example, "treatment" includes the alleviation of a disease symptom in a subject by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or more.

The term "prevent" includes administration of compositions or antibodies to prevent or delay the onset of the disease, or to prevent the manifestation of clinical or subclinical symptoms thereof i.e.

prophylactic administration, or therapeutic suppression or alleviation of symptoms after the manifestation of the disease.

The term "vector" is intended to refer to a polynucleotide molecule capable of transporting another polynucleotide to which it has been linked. One type of vector is a "plasmid", which refers to a circular double stranded DNA loop into which additional DNA segments may be ligated. Another type of vector is a viral vector, wherein additional DNA segments may be ligated into the viral genome. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) can be integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. Moreover, certain vectors are capable of directing the expression of genes to which they are operatively linked. Such vectors are referred to herein as "recombinant expression vectors" (or simply, "expression vectors"). In general, expression vectors of utility in recombinant DNA techniques are often in the form of plasmids. In the present specification, "plasmid" and "vector" may be used interchangeably as the plasmid is the most commonly used form of vector. However, the invention is intended to include such other forms of expression vectors, such as viral vectors (e.g., replication defective retroviruses, adenoviruses and adeno- associated viruses), which serve equivalent functions.

CD32b antibody molecules

Anti-human CD32b antibody molecules for use in a combination of the present invention include isolated antibodies or antigen-binding fragments thereof that bind with a higher affinity for human CD32b protein, than to human CD32a protein. Selectivity for CD32b over CD32a is desired to ensure selective binding to CD32b positive B-cell malignancies and B-cells while lacking binding to CD32a positive immune cells, including monocytes and neutrophils.

Examples of such anti-human CD32b antibody molecules for use in a combination of the present invention include antibodies NOV0281, NOV0308, NOV0563, NOV1216, NOV1218, NOV1219, NOV2106, NOV2107, NOV2108, NOV2109, NOV2110, NOV2111, NOV2112, and NOV2113

(including antibodies with wild type Fc regions or that are afucosylated) whose sequences are listed in Table 1. Details regarding the generation and characterization of the NOV antibodies described herein can be found in PCT/IB2016/057745. Additional examples of anti-CD32b antibody molecules for use in a combination of the present invention include antibody 2B6 (PCT Publication WO2004/016750), antibodies 20, 24, 26 or 28 (PCT Publication WO2009/083009) and antibody 6G11 (PCT Publications WO2012/022985 and WO2015/173384), whose sequences are listed in Table 1.

Additional antibody molecules that specifically bind to CD32b include those comprising (or alternatively, consisting of) a VH amino acid sequence listed in Table 1, wherein no more than about 10 amino acids in a framework sequence (for example, a sequence which is not a CDR) have been mutated (wherein a mutation is, as various non-limiting examples, an addition, substitution or deletion) or wherein no more than about 20 amino acids in a framework sequence (for example, a sequence which is not a CDR) have been mutated (wherein a mutation is, as various non-limiting examples, an addition, substitution or deletion).

Furthermore, antibody molecules that specifically bind to CD32b, include those comprising (or alternatively, consisting of) a VL amino acid sequence listed in Table 1, wherein no more than about 10 amino acids in a framework sequence (for example, a sequence which is not a CDR) have been mutated (wherein a mutation is, as various non-limiting examples, an addition, substitution or deletion) or wherein no more than about 20 amino acids in a framework sequence (for example, a sequence which is not a CDR) have been mutated (wherein a mutation is, as various non-limiting examples, an addition, substitution or deletion).

Other antibody molecules for use in a combination of the present invention include amino acids that have been mutated, yet have at least 60, 70, 80, 90, 91, 92, 93, 94, 95, 96, 97, 98 or 99 percent identity in the CDR regions with the CDR regions depicted in the sequences described in Table 1. In one aspect, other antibody molecules of the invention includes mutant amino acid sequences wherein no more than 1, 2, 3, 4 or 5 amino acids have been mutated in the CDR regions when compared with the CDR regions depicted in the sequence described in Table 1.

Other antibody molecules for use in a combination of the present invention include those wherein the amino acids or nucleic acids encoding the amino acids have been mutated, yet have at least 60, 70, 80, 90 or 95 percent identity to the sequences described in Table 1. In one embodiment, it includes mutant amino acid sequences wherein no more than 1, 2, 3, 4 or 5 amino acids have been mutated in the variable regions when compared with the variable regions depicted in the sequence described in Table 1, while retaining substantially the same therapeutic activity.

In a specific embodiment, antibody molecules, which bind human CD32b and which can be used in a combination of the present invention comprise heavy and light chain variable domain CDRs comprising:

the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 1, 2, and 3, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 8, 9, and 10, respectively;

the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 15, 16, and 17, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 22, 23, and 24, respectively;

the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 29, 30, and 31 respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 36, 37, 38, respectively;

the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 43, 44, and 45, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 50, 51, 52, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 57, 58, and 59, respectively, and the

LCDR1, LCDR2. and LCDR3 sequences of SEQ ID NOs: 64, 65. and 66, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 71, 72, and 73, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 78, 79, and 80, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 85, 86, and 87, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 92, 93. and 94, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 99, 100, and 101, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 106, 107, and 108, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 1 13, 1 14, and 1 15. respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 120, 121, and 122, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 127, 128, and 129, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 134, 135, and 136, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 141, 142, and 143. respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 148, 149, and 150, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 155, 156, and 157, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 162, 163, and 164, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 169, 170, and 171. respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 176, 177, and 178, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 183, 184, and 185, respectively, and the

LCDR1, LCDR2. and LCDR3 sequences of SEQ ID NOs: 190. 191, and 192, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 197, 198, and 199. respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 202, 203, and 204, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 207, 208, and 209, respectively, and the

LCDR1, LCDR2. and LCDR3 sequences of SEQ ID NOs: 211, 212, and 213, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 215, 216, and 217, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 219, 220, and 221, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 223, 224, and 225, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 227, 228, and 229, respectively;

the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 231, 232, and 233, respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 235, 236, and 237, respectively; and the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 239, 240, and 241. respectively, and the

LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 243, 244, and 245, respectively;

wherein the CDRs are numbered according to Kabat numbering (Kabat et al, supra)

In another specific embodiment, antibody molecules, which bind human CD32b and which can be used in a combination of the present invention comprise the following VH and VL amino acid sequences: the VH amino acid sequence of SEQ ID NO: 4 and the VL amino acid sequence of SEQ ID NO: 11; the VH amino acid sequence of SEQ ID NO: 18 and the VL amino acid sequence of SEQ ID NO: 25; the VH amino acid sequence of SEQ ID NO: 32 and the VL amino acid sequence of SEQ ID NO: 39; the VH amino acid sequence of SEQ ID NO: 46 and the VL amino acid sequence of SEQ ID NO: 53; the VH amino acid sequence of SEQ ID NO: 60 and the VL amino acid sequence of SEQ ID NO: 67; the VH amino acid sequence of SEQ ID NO: 74 and the VL amino acid sequence of SEQ ID NO: 81; the VH amino acid sequence of SEQ ID NO: 88 and the VL amino acid sequence of SEQ ID NO: 95; the VH amino acid sequence of SEQ ID NO: 102 and the VL amino acid sequence of SEQ ID NO: 109; the VH amino acid sequence of SEQ ID NO: 116 and the VL amino acid sequence of SEQ ID NO: 123; the VH amino acid sequence of SEQ ID NO: 130 and the VL amino acid sequence of SEQ ID NO: 137; the VH amino acid sequence of SEQ ID NO: 144 and the VL amino acid sequence of SEQ ID NO: 151; the VH amino acid sequence of SEQ ID NO: 158 and the VL amino acid sequence of SEQ ID NO: 165; the VH amino acid sequence of SEQ ID NO: 172 and the VL amino acid sequence of SEQ ID NO: 179; the VH amino acid sequence of SEQ ID NO: 186 and the VL amino acid sequence of SEQ ID NO: 193; the VH amino acid sequence of SEQ ID NO: 200 and the VL amino acid sequence of SEQ ID NO: 205; the VH amino acid sequence of SEQ ID NO: 210 and the VL amino acid sequence of SEQ ID NO: 214; the VH amino acid sequence of SEQ ID NO: 218 and the VL amino acid sequence of SEQ ID NO: 222; the VH amino acid sequence of SEQ ID NO: 226 and the VL amino acid sequence of SEQ ID NO: 230; the VH amino acid sequence of SEQ ID NO: 234 and the VL amino acid sequence of SEQ ID NO: 238; or the VH amino acid sequence of SEQ ID NO: 242 and the VL amino acid sequence of SEQ ID NO: 246.

In another specific embodiment, antibody molecules, which bind human CD32b and which can be used in a combination of the present invention comprise the following heavy chain and light chain amino acid sequences:

the heavy chain amino acid sequence of SEQ ID NO: 6 and the light chain amino acid sequence of SEQ ID NO: 13;

the heavy chain amino acid sequence of SEQ ID NO: 20 and the light chain amino acid sequence of SEQ ID NO: 27;

the heavy chain amino acid sequence of SEQ ID NO: 34 and the light chain amino acid sequence of SEQ ID NO: 41;

the heavy chain amino acid sequence of SEQ ID NO: 48 and the light chain amino acid sequence of SEQ ID NO: 55;

the heavy chain amino acid sequence of SEQ ID NO: 62 and the light chain amino acid sequence of SEQ ID NO: 69;

the heavy chain amino acid sequence of SEQ ID NO: 76 and the light chain amino acid sequence of SEQ ID NO: 83;

the heavy chain amino acid sequence of SEQ ID NO: 90 and the light chain amino acid sequence of SEQ ID NO: 97;

the heavy chain amino acid sequence of SEQ ID NO: 104 and the light chain amino acid sequence of SEQ ID NO: 111;

the heavy chain amino acid sequence of SEQ ID NO: 118 and the light chain amino acid sequence of SEQ ID NO: 125;

the heavy chain amino acid sequence of SEQ ID NO: 132 and the light chain amino acid sequence of SEQ ID NO: 139;

the heavy chain amino acid sequence of SEQ ID NO: 174 and the light chain amino acid sequence of SEQ ID NO: 181; or

the heavy chain amino acid sequence of SEQ ID NO: 188 and the light chain amino acid sequence of SEQ ID NO: 195.

Since each of these antibody molecules can bind to CD32b, the VH, VL, full length light chain, and full length heavy chain sequences (amino acid sequences and the nucleotide sequences encoding the amino acid sequences) can be "mixed and matched" to create other CD32b-binding antibody molecules of the invention. Such "mixed and matched" CD32b-binding antibodies can be tested using the binding assays known in the art (e.g., ELISAs, and other assays described in the Example section). When these chains are mixed and matched, a VH sequence from a particular VH/VL pairing should be replaced with a structurally similar VH sequence. Likewise a full length heavy chain sequence from a particular full length heavy chain/full length light chain pairing should be replaced with a structurally similar full length heavy chain sequence. Likewise, a VL sequence from a particular VH/VL pairing should be replaced with a structurally similar VL sequence. Likewise a full length light chain sequence from a particular full length heavy chain/full length light chain pairing should be replaced with a structurally similar full length light chain sequence.

Accordingly, antibody molecules, for use in a combination of the present invention may comprise: a heavy chain variable region VHCDR1 comprising an amino acid sequence selected from any of SEQ ID NOs: 1, 15, 29, 43, 57, 71, 85, 99, 113, 127, 141, 155, 169, and 183, 197, 207, 215, 223, 231 and 239; a heavy chain variable region VHCDR2 comprising an amino acid sequence selected from any of SEQ ID NOs: 2, 16, 30, 44, 58, 72, 86. 100, 114, 128, 142, 156, 170, 184, 198, 208, 216, 224, 232 and 240 ; a heavy chain variable region VHCDR3 comprising an amino acid sequence selected from any of SEQ ID NOs: 3, 17, 31, 45, 59, 73, 87, 101, 115, 129, 143, 157, 171, 185, 199, 209, 217, 225, 233 and 241; a light chain variable region VLCDR1 comprising an amino acid sequence selected from any of SEQ ID NOs: 8, 22, 36, 50, 64, 78, 92, 106, 120, 134, 148, 162, 176, 190, 202, 211, 219, 227, 235 and 243; a light chain variable region VLCDR2 comprising an amino acid sequence selected from any of SEQ ID NOs: 9, 23, 37, 51, 65, 79, 93, 107, 121, 135, 149, 163, 177, 191, 203, 212, 220, 228, 236 and 244; and a light chain variable region VLCDR3 comprising an amino acid sequence selected from any of SEQ ID NOs: 10, 24, 38, 52, 66, 80, 94, 108, 122, 136, 150, 164, 178, 192, 204, 213, 221, 229, 237 and 245; as described in Table 1, wherein the antibody molecule specifically binds CD32b. Furthermore, antibody molecules, for use in a combination of the present invention may comprise: a heavy chain variable region (VH) comprising an amino acid sequence selected from any of SEQ ID NOs: 4, 18, 32, 46, 60, 74, 88, 102, 116, 130, 144, 158, 172, 186, 200, 210, 218, 226, 234 and 242; and a light chain variable region (VL) comprising an amino acid sequence selected from any of SEQ ID NOs: 11, 25, 39, 53, 67, 81, 95, 109, 123, 137, 151, 165, 179, 193, 205, 214, 222, 230, 238 and 246, as described in Table 1.

In addition, antibody molecules, for use in a combination of the present invention may comprise: a heavy chain comprising an amino acid sequence selected from any of SEQ ID NOs: 6, 20, 34, 48, 62, 76, 90, 104, 118, 132, 174 and 188; and a light chain comprising an amino acid sequence selected from any of SEQ ID NOs: 13, 27, 41, 55, 69, 83, 97, 111, 125, 139, 181, 195, as described in Table 1.

In one embodiment, an antibody molecule, for use in a combination of the present invention and that specifically binds to CD32b is an antibody that is described in Table 1. In one embodiment, an antibody that specifically binds to CD32b is NOV0281. In one embodiment, an antibody that specifically binds to CD32b is NOV0308. In one embodiment, an antibody that specifically binds to CD32b is NOV0563. In one embodiment, an antibody that specifically binds to CD32b is NOV1216. In one embodiment, an antibody that specifically binds to CD32b is NOV1218. In one embodiment, an antibody that specifically binds to CD32b is NOV1219. In one embodiment, an antibody that specifically binds to CD32b is NOV2106. In one embodiment, an antibody that specifically binds to CD32b is NOV2107. In one embodiment, an antibody that specifically binds to CD32b is NOV2108. In one embodiment, an antibody that specifically binds to CD32b is NOV2109. In one embodiment, an antibody that specifically binds to CD32b is NOV2112. In one embodiment, an antibody that specifically binds to CD32b is NOV2113. In one embodiment, an antibody that specifically binds to CD32b is 2B6 (WO2004/016750). In one embodiment, an antibody that specifically binds to CD32b is Ab 20, 24, 26 or 28

(WO2009/083009). In one embodiment, an antibody that specifically binds to CD32b is 6G11

(WO2012/022985 and WO2015/173384).

In one embodiment, an antibody of the invention optimized for expression in a mammalian cell has a full length heavy chain sequence and a full length light chain sequence, wherein one or more of these sequences have specified amino acid sequences based on the antibody molecules described herein or conservative modifications thereof, and wherein the antibodies retain the desired functional properties of the CD32b-binding antibodies and antigen-binding fragments thereof of the invention. Accordingly, the invention provides an isolated monoclonal antibody optimized for expression in a mammalian cell comprising a full length heavy chain and a full length light chain wherein: the full length heavy chain comprises an amino acid sequence selected from any of SEQ ID NOs: 6, 20, 34, 48, 62, 76, 90, 104, 118, 132, 174 and 188, as described in Table 1, and conservative modifications thereof; and the full length light chain comprises an amino acid sequence selected from any of SEQ ID NOs: 13, 27, 41, 55, 69, 83, 97, 111, 125, 139, 181 and 195, as described in Table 1, and conservative modifications thereof; wherein the antibody specifically binds to CD32b and mediates both macrophage and NK cell killing of antibody bound, CD32b positive target cells.

In some embodiments of the CD32b-binding antibody molecules disclosed herein, the antibodies comprise a wild type (WT) Fc sequence. Alternatively, antibody molecules of the invention may be engineered to include modifications within the Fc region, typically to alter one or more functional properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and/or antigen-dependent cellular cytotoxicity. Furthermore, an antibody of the invention may be chemically modified (e.g., one or more chemical moieties can be attached to the antibody) or be modified to alter its glycosylation, again to alter one or more functional properties of the antibody. Each of these

embodiments is described in further detail below. The numbering of residues in the Fc region is that of the EU index of Kabat.

In yet another embodiment, the Fc region is modified to increase the ability of the antibody to mediate antibody dependent cellular cytotoxicity (ADCC) and/or to increase the affinity of the antibody for an Fc-gamma receptor by modifying one or more amino acids. This approach is described further, for example, in PCT Publication WO 00/42072 by Presta and by Lazar et al, (2006) PNAS 103(110): 4005- 4010. Moreover, the binding sites on human IgGl for Fc-gamma RI, Fc-gamma RII, Fc-gamma RIII and FcRn have been mapped and variants with improved binding have been described (see Shields, R. L. et al, (2001) J. Biol. Chem. 276:6591-6604).

In still another embodiment, the glycosylation of an antibody is modified. For example, an aglycoslated antibody can be made (i.e., the antibody lacks glycosylation). Glycosylation can be altered to, for example, increase the affinity of the antibody for "antigen^' . Such carbohydrate modifications can be accomplished by, for example, altering one or more sites of glycosylation within the antibody sequence. For example, one or more amino acid substitutions can be made that result in elimination of one or more variable region framework glycosylation sites to thereby eliminate glycosylation at that site. Such aglycosylation may increase the affinity of the antibody for antigen. Such an approach is described in further detail in U.S. Pat. Nos. 5,714,350 and 6,350,861 by Co et al.

Additionally or alternatively, an antibody can be made that has an altered type of glycosylation, such as a hypofucosylated or afucosylated antibody having reduced amounts of fucosyl residues, or an antibody having increased bisecting GlcNac structures. Such altered glycosylation patterns have been demonstrated to increase the ADCC ability of antibodies. Such carbohydrate modifications can be accomplished by, for example, expressing the antibody in a host cell with altered glycosylation machinery. Cells with altered glycosylation machinery have been described in the art and can be used as host cells in which to express recombinant antibodies of the invention to thereby produce an antibody with altered glycosylation. For example, EP 1,176,195 by Hang et al, describes a cell line with a functionally disrupted FUT8 gene, which encodes a fucosyl transferase, such that antibodies expressed in such a cell line exhibit hypofucosylation. PCT Publication WO 03/035835 by Presta describes a variant

CHO cell line, LecI3 cells, with reduced ability to attach fucose to Asn (297)-linked carbohydrates, also resulting in hypofucosylation of antibodies expressed in that host cell (see also Shields, R. L. et al.,( 2002) J. Biol. Chem. 277:26733-26740). PCT Publication WO 99/54342 by Umana et al. describes cell lines engineered to express glycoprotein-modifying glycosyl transferases (e.g., beta (1,4)— N

acetylglucosaminyl-transferase III (GnTIII)) such that antibodies expressed in the engineered cell lines exhibit increased bisecting GlcNac structures which results in increased ADCC activity of the antibodies (see also Umana et al., (1999 )Nat. Biotech. 17: 176-180). Von Horsten et al., in Glycobiology

20(12): 1607-18 (2010) also describe a method of producing non-fucosylated antibodies by co-expression of antibodies with a heterologous GDP-6-deoxy-D-lyxo-4-hexulose reductase in CHO cells.

In a specific embodiment, the CD32b-binding antibody is afucosylated NOV2108, comprising a WT Fc. In a specific embodiment, the CD32b-binding antibody comprises an HCDR1, HCDR2, and HCDR3 comprising the amino acid sequences of SEQ ID NOs: 113, 114, and 115, respectively, and a LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NOs: 120, 121, and 122 respectively, and wherein the antibody is afucosylated. In another specific embodiment, the CD32b- binding antibody comprises a VH comprising the amino acid sequence of SEQ ID NO: 116 and a VL comprising the amino acid sequence of SEQ ID NO: 123, and wherein the antibody is afucosylated. In yet another embodiment, the CD32b-binding antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 118 and a light chain comprising the amino acid sequence of SEQ ID NO: 125, wherein the antibody is afucosylated.

In a further embodiment, the hinge region of CHI is modified such that the number of cysteine residues in the hinge region is altered, e.g., increased or decreased. This approach is described further in U.S. Pat. No. 5,677,425 by Bodmer et al. The number of cysteine residues in the hinge region of CHI is altered, for example, to facilitate assembly of the light and heavy chains or to increase or decrease the stability of the antibody.

In another embodiment, the Fc hinge region of an antibody is mutated to decrease the biological half-life of the antibody. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc-hinge fragment such that the antibody has impaired

Staphylococcyl protein A (SpA) binding relative to native Fc-hinge domain SpA binding. This approach is described in further detail in U.S. Pat. No. 6,165,745 by Ward et al.

In another embodiment, the antibody is modified to increase its biological half-life. Various approaches are possible. For example, one or more of the following mutations can be introduced: T252L, T254S, T256F, as described in U.S. Pat. No. 6,277,375 to Ward. Alternatively, to increase the biological half-life, the antibody can be altered within the CHI or CL region to contain a salvage receptor binding epitope taken from two loops of a CH2 domain of an Fc region of an IgG, as described in U.S. Pat. Nos. 5,869,046 and 6, 121,022 by Presta et al.

In one embodiment, the Fc region is altered by replacing at least one amino acid residue with a different amino acid residue to alter the effector functions of the antibody. For example, one or more amino acids can be replaced with a different amino acid residue such that the antibody has an altered affinity for an effector ligand but retains the antigen-binding ability of the parent antibody. The effector ligand to which affinity is altered can be, for example, an Fc receptor or the C 1 component of complement. This approach is described in further detail in U.S. Pat. Nos. 5,624,821 and 5,648,260, both by Winter et al.

In another embodiment, one or more amino acids selected from amino acid residues can be replaced with a different amino acid residue such that the antibody has altered Clq binding and/or reduced or abolished complement dependent cytotoxicity (CDC). This approach is described in further detail in U.S. Pat. No. 6, 194,551 by Idusogie et al. In another embodiment, one or more amino acid residues are altered to thereby alter the ability of the antibody to fix complement. This approach is described further in PCT Publication WO 94/29351 by Bodmer et al.

The invention also provides substantially purified nucleic acid molecules which encode polypeptides comprising segments or domains of the CD32b-binding antibody chains described above. Some of the nucleic acids of the invention comprise the nucleotide sequence encoding the heavy chain variable region shown in any of SEQ ID NOs: 4, 18, 32, 46, 60, 74, 88, 102, 116, 130, 144, 158, 172, 186, 200, 210, 218, 226, 234 or 242, and/or the nucleotide sequence encoding the light chain variable region shown in any of SEQ ID NOs: 11, 25, 39, 53, 67, 81, 95, 109, 123, 137, 151, 165, 179, 193, 205, 214, 222, 230, 238 or 246. In a specific embodiment, the nucleic acid molecules are those identified in Table 1 and comprise any of SEQ ID Nos: 5, 19, 33, 47, 61, 75, 89, 103, 117, 131, 145, 159, 173, 187 or 201, encoding a heavy chain variable region, and/or the nucleic acid molecules identified in Table 1 that comprise any of SEQ ID Nos: 12, 26, 40, 54, 68, 82, 96, 110, 124, 138, 152, 166, 180, 194 or 206, encoding a light chain variable region. Some other nucleic acid molecules of the invention comprise nucleotide sequences that are substantially identical (e.g., at least 65, 80%, 95%, or 99%) to the nucleotide sequences of those identified in Table 1. When expressed from appropriate expression vectors, polypeptides encoded by these polynucleotides are capable of exhibiting CD32b antigen binding capacity.

Also provided in the invention are polynucleotides which encode at least one CDR region and usually all three CDR regions from the heavy or light chain of the CD32b-binding antibody set forth in Table 1. Some other polynucleotides encode all or substantially all of the variable region sequence of the heavy chain and/or the light chain of the CD32b-binding antibody set forth in Table 1. Because of the degeneracy of the code, a variety of nucleic acid sequences will encode each of the immunoglobulin amino acid sequences.

The nucleic acid molecules of the invention can encode both a variable region and a constant region of the antibody. Some of the nucleic acid sequences of the invention comprise nucleotides encoding a mature heavy chain sequence that is identical or substantially identical (e.g., at least 80%, 90%, or 99%) to the mature heavy chain sequence set forth in any of SEQ ID NOs: 6, 20, 34, 48, 62, 76, 90, 104, 118, 132, 174 or 188. Some of the nucleic acid sequences of the invention comprise nucleotide encoding a mature light chain sequence that is identical or substantially identical (e.g., at least 80%, 90%, or 99%) to the mature light chain sequence set forth in any of SEQ ID NOs: 13, 27, 41, 55, 69, 83, 97, 111, 125, 139, 181 or 195. In a specific embodiment, the nucleic acid molecules are those identified in Table 1 and comprise any of SEQ ID Nos: 7, 21, 35, 49, 63, 77, 91, 105, 119, 133, 175 or 189, encoding a mature heavy chain, and/or the nucleic acid molecules identified in Table 1 that comprise any of SEQ ID Nos: 14, 28, 42, 56, 70, 84, 98, 112, 126, 140, 182 or 196, encoding a mature light chain.

The polynucleotide sequences can be produced by de novo solid-phase DNA synthesis or by PCR mutagenesis of an existing sequence (e.g., sequences as described in Table 1) encoding a CD32b-binding antibody or its binding fragment. Direct chemical synthesis of nucleic acids can be accomplished by methods known in the art, such as the phosphotriester method of Narang et al, (1979), Meth. Enzymol. 68:90; the phosphodiester method of Brown et al, (1979) Meth. Enzymol. 68: 109; the

diethylphosphoramidite method of Beaucage et al., (1981) Tetra. Lett., 22: 1859; and the solid support method of U.S. Pat. No. 4,458,066. Introducing mutations to a polynucleotide sequence by PCR can be performed as described in, e.g., PCR Technology: Principles and Applications for DNA Amplification, H. A. Erlich (Ed.), Freeman Press, NY, N.Y., 1992; PCR Protocols: A Guide to Methods and Applications, Innis et al. (Ed.), Academic Press, San Diego, Calif, (1990); Mattila et al, (1991) Nucleic Acids Res. 19:967; and Eckert et al, (1991) PCR Methods and Applications 1 : 17.

IL-15

As used herein, the terms "IL-15" and "interleukin-15" refer to a wild-type IL-15 or an IL-15 derivative. In specific embodiments, the IL-15 is isolated and recombinantly produced. As used herein, the terms "wild-type IL-15" and "wild-type interleukin-15" in the context of proteins or polypeptides refer to any mammalian interleukin-15 amino acid sequences, including immature or precursor and mature forms. Non-limiting examples of GeneBank Accession Nos. for the amino acid sequence of various species of wild-type mammalian interleukin-15 include NP_000576 (human, immature form), CAA62616 (human, immature form), NP_001009207 {Felis catus, immature form), AAB94536 (Rattus norvegicus, immature form), AAB41697 {Rattus norvegicus, immature form), NP_032383 {Mus musculus, immature form), AAR19080 (canine), AAB60398 (Macaca mulatta, immature form),

AAI00964 (human, immature form), AAH23698 {mus musculus, immature form), and AAH18149 (human). The amino acid sequence of the immature/precursor form of human IL-15, which comprises the long signal peptide (underlined) and the mature human IL-15 (italicized), as provided in SEQ ID NO: 251 in Table 1. In some embodiments, IL-15 is the immature or precursor form of a mammalian IL-15. In other embodiments, IL-15 is the mature form of a mammalian IL-15. In a specific embodiment, IL-15 is the precursor form of human IL-15. In another embodiment, IL-15 is the mature form of human IL-15. In one embodiment, the IL-15 protein/polypeptide is isolated or purified.

As used herein, the terms "IL-15" and "interleukin-15" in the context of nucleic acids refer to any nucleic acid sequences encoding mammalian interleukin-15, including the immature or precursor and mature forms. Non-limiting examples of GeneBank Accession Nos. for the nucleotide sequence of various species of wild-type mammalian IL-15 include NM_000585 (human), NM_008357 {Mus musculus), and RNU69272 {Rattus norvegicus). The nucleotide sequence encoding the

immature/precursor form of human IL-15, which comprises the nucleotide sequence encoding the long signal peptide (underlined) and the nucleotide sequence encoding the mature human IL-15 (italicized), as provided in SEQ ID NO: 252 in Table 1. In a specific embodiment, the nucleic acid is an isolated or purified nucleic acid. In some embodiments, nucleic acids encode the immature or precursor form of a mammalian IL-15. In other embodiments, nucleic acids encode the mature form of a mammalian IL-15. In a specific embodiment, nucleic acids encoding IL-15 encode the precursor form of human IL-15. In another embodiment, nucleic acids encoding IL-15 encode the mature form of human IL-15.

As used herein, the terms "IL-15 derivative" and "interleukin-15 derivative" in the context of proteins or polypeptides refer to: (a) a polypeptide that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to a wild-type mammalian IL-15 polypeptide; (b) a polypeptide encoded by a nucleic acid sequence that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical a nucleic acid sequence encoding a wild-type mammalian IL-15 polypeptide; (c) a polypeptide that contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more amino acid mutations (i.e., additions, deletions and/or substitutions) relative to a wild-type mammalian IL-15 polypeptide; (d) a polypeptide encoded by nucleic acids can hybridize under high or medium stringency hybridization conditions to nucleic acids encoding a wild-type mammalian IL-15 polypeptide; (e) a polypeptide encoded by a nucleic acid sequence that can hybridize under high or medium stringency hybridization conditions to a nucleic acid sequence encoding a fragment of a wild-type mammalian IL-15 polypeptide of at least 20 contiguous amino acids, at least 30 contiguous amino acids, at least 40 contiguous amino acids, at least 50 contiguous amino acids, at least 100 contiguous amino acids, or at least 150 contiguous amino acids; and/or (f) a fragment of a wild-type mammalian IL-15 polypeptide. IL-15 derivatives also include a polypeptide that comprises the amino acid sequence of a mature form of a mammalian IL-15 polypeptide and a heterologous signal peptide amino acid sequence. In a specific embodiment, an IL-15 derivative is a derivative of a wild-type human IL-15 polypeptide. In another embodiment, an IL-15 derivative is a derivative of an immature or precursor form of human IL-15 polypeptide. In another embodiment, an IL-15 derivative is a derivative of a mature form of human IL-15 polypeptide. In another embodiment, an IL-15 derivative is the IL- 15N72D described in, e.g., Zhu et al., (2009), J. Immunol. 183: 3598 or U.S. Patent No. 8,163,879. In another embodiment, an IL-15 derivative is one of the IL-15 variants described in U.S. Patent No.

8, 163,879. In one embodiment, an IL-15 derivative is isolated or purified.

In a preferred embodiment, IL-15 derivatives retain at least 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of wild-type mammalian IL-15 polypeptide to bind IL-15Ra polypeptide, as measured by assays well known in the art, e.g., ELISA, BIAcore^®, co-immunoprecipitation. In another preferred embodiment, IL-15 derivatives retain at least 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of wild-type mammalian IL-15 polypeptide to induce IL-15 -mediated signal transduction, as measured by assays well-known in the art, e.g., electromobility shift assays, ELISAs and other immunoassays. In a specific embodiment, IL-15 derivatives bind to IL-15Ra and/or Ιί-15Ρνβγ as assessed by, e.g., ligand/receptor binding assays well-known in the art. Percent identity can be determined using any method known to one of skill in the art and as described supra. As used herein, the terms "IL-15 derivative" and "interleukin-15 derivative" in the context of nucleic acids refer to: (a) a nucleic acid sequence that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to the nucleic acid sequence encoding a mammalian IL-15 polypeptide; (b) a nucleic acid sequence encoding a polypeptide that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical the amino acid sequence of a wild-type mammalian IL-15 polypeptide; (c) a nucleic acid sequence that contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more nucleic acid base mutations (i.e., additions, deletions and/or substitutions) relative to the nucleic acid sequence encoding a mammalian IL-15 polypeptide; (d) a nucleic acid sequence that hybridizes under high or medium stringency hybridization conditions to a nucleic acid sequence encoding a mammalian IL-15 polypeptide; (e) a nucleic acid sequence that hybridizes under high or medium stringency hybridization conditions to a fragment of a nucleic acid sequence encoding a mammalian IL-15 polypeptide; and/or (f) a nucleic acid sequence encoding a fragment of a nucleic acid sequence encoding a mammalian IL-15 polypeptide. In a specific embodiment, an IL-15 derivative in the context of nucleic acids is a derivative of a nucleic acid sequence encoding a human IL-15 polypeptide. In another embodiment, an IL-15 derivative in the context of nucleic acids is a derivative of a nucleic acid sequence encoding an immature or precursor form of a human IL-15 polypeptide. In another embodiment, an IL-15 derivative in the context of nucleic acids is a derivative of a nucleic acid sequence encoding a mature form of a human IL-15 polypeptide. In another embodiment, an IL-15 derivative in the context of nucleic acids is the nucleic acid sequence encoding the IL-15N72D described in, e.g., Zhu et al, (2009; supra), or U.S. Patent No. 8,163,879. In another embodiment, an IL-15 derivative in the context of nucleic acids is the nucleic acid sequence encoding one of the IL-15 variants described in U.S. Patent No. 8, 163,879.

IL-15 derivative nucleic acid sequences include codon-optimized nucleic acid sequences that encode mammalian IL-15 polypeptide, including mature and immature forms of IL-15 polypeptide. In other embodiments, IL-15 derivative nucleic acids include nucleic acids that encode mammalian IL-15 RNA transcripts containing mutations that eliminate potential splice sites and instability elements (e.g., A/T or A/U rich elements) without affecting the amino acid sequence to increase the stability of the mammalian IL-15 RNA transcripts. In one embodiment, the IL-15 derivative nucleic acid sequences include the codon-optimized nucleic acid sequences described in PCT Publication WO2007/084342. In certain embodiments, the IL-15 derivative nucleic acid sequence is the codon-optimized sequence in SEQ ID NO: 254 in Table 1 (the amino acid sequence encoded by such a nucleic acid sequence is provided in SEQ ID NO: 255 in Table 1).

In one embodiment, IL-15 derivative nucleic acid sequences encode proteins or polypeptides that retain at least 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of a wild-type mammalian IL-15 polypeptide to bind IL-15Ra, as measured by assays well known in the art, e.g., ELISA, BIAcore^®, co- immunoprecipitation. In another preferred embodiment, IL-15 derivative nucleic acid sequences encode proteins or polypeptides that retain at least 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of a wild-type mammalian IL-15 polypeptide to induce IL-15 -mediated signal transduction, as measured by assays well-known in the art, e.g., electromobility shift assays, ELISAs and other immunoassays. In a specific embodiment, IL-15 derivative nucleic acid sequences encode proteins or polypeptides that bind to IL-15Ra and/or Ιί-15Ρνβγ as assessed by, e.g., ligand/receptor assays well-known in the art.

IL-15Ra

As used herein, the terms "IL-15Ra" and "interleukin-15 receptor alpha" refer to a wild-type IL- 15Ra, an IL-15Ra derivative, or a wild-type IL-15Ra and an IL-15Ra derivative. In specific

embodiments, the IL-15Ra is isolated and recombinantly produced. As used herein, the terms "wild-type IL-15Ra" and "wild-type interleukin-15 receptor alpha" in the context of proteins or polypeptides refer to mammalian interleukin-15 receptor alpha ("IL-15Ra") amino acid sequence, including immature or precursor and mature forms and isoforms. Non-limiting examples of GeneBank Accession Nos. for the amino acid sequence of various wild-type mammalian IL-15Ra include NP 002180 (human), ABK41438 (Macaca mulatto), NP_032384 (Mus musculus), Q60819 (Mus musculus), CAI41082 (human). The amino acid sequence of the immature form of the full length human IL-15Ra, which comprises the signal peptide (underlined) and the mature human IL-15Ra (italicized), as provided in SEQ ID NO: 256 in Table 1. The amino acid sequence of the immature form of the soluble human IL-15Ra, which comprises the signal peptide (underlined) and the mature human soluble IL-15Ra (italicized), as provided in SEQ ID NO: 257 in Table 1. In some embodiments, IL-15Ra is the immature form of a mammalian IL-15Ra polypeptide. In other embodiments, IL-15Ra is the mature form of a mammalian IL-15Ra polypeptide. In certain embodiments, IL-15Ra is the soluble form of mammalian IL-15Ra polypeptide. In other embodiments, IL-15Ra is the full-length form of a mammalian IL-15Ra polypeptide. In a specific embodiment, IL-15Ra is the immature form of a human IL-15Ra polypeptide. In another embodiment, IL-15Ra is the mature form of a human IL-15Ra polypeptide. In certain embodiments, IL-15Ra is the soluble form of human IL-15Ra polypeptide. In other embodiments, IL-15Ra is the full-length form of a human IL-15Ra polypeptide. In one embodiment, an IL-15Ra protein or polypeptide is isolated or purified.

As used herein, the terms "IL-15Ra" and "interleukin-15 receptor alpha" in the context of nucleic acids refer to any nucleic acid sequences encoding mammalian interleukin-15 receptor alpha, including the immature or precursor and mature forms. Non-limiting examples of GeneBank Accession Nos. for the nucleotide sequence of various species of wild-type mammalian IL-15Ra include NM_002189 (human), EF033114 {Macaca mulatto), and NM_008358 (Mus musculus). The nucleotide sequence encoding the immature form of wild-type human IL-15Ra, which comprises the nucleotide sequence encoding the signal peptide (underlined) and the nucleotide sequence encoding the mature human IL- 15Ra (italicized), as provided in SEQ ID NO: 258 in Table 1. The nucleotide sequence encoding the immature form of soluble human IL-15Ra protein or polypeptide, which comprises the nucleotide sequence encoding the signal peptide (underlined) and the nucleotide sequence encoding the mature human soluble IL-15Ra (italicized), as provided in SEQ ID NO in 259 in Table 1). In a specific embodiment, the nucleic acid is an isolated or purified nucleic acid. In some embodiments, nucleic acids encode the immature form of a mammalian IL-15Ra polypeptide. In other embodiments, nucleic acids encode the mature form of a mammalian IL-15Ra polypeptide. In certain embodiments, nucleic acids encode the soluble form of a mammalian IL-15Ra polypeptide. In other embodiments, nucleic acids encode the full-length form of a mammalian IL-15Ra polypeptide. In a specific embodiment, nucleic acids encode the precursor form of human IL-15 polypeptide. In another embodiment, nucleic acids encode the mature of human IL-15 polypeptide. In certain embodiments, nucleic acids encode the soluble form of a human IL-15Ra polypeptide. In other embodiments, nucleic acids encode the full-length form of a human IL-15Ra polypeptide.

As used herein, the terms "IL-15Ra derivative" and "interleukin-15 receptor alpha derivative" in the context of a protein or polypeptide refer to: (a) a polypeptide that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to a wild-type mammalian IL-15 polypeptide; (b) a polypeptide encoded by a nucleic acid sequence that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical a nucleic acid sequence encoding a wild-type mammalian IL-15Ra polypeptide; (c) a polypeptide that contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more amino acid mutations (i.e., additions, deletions and/or substitutions) relative to a wild-type mammalian IL-15Ra polypeptide; (d) a polypeptide encoded by a nucleic acid sequence that can hybridize under high or medium stringency hybridization conditions to a nucleic acid sequence encoding a wild-type mammalian IL-15Ra polypeptide; (e) a polypeptide encoded by a nucleic acid sequence that can hybridize under high or medium stringency hybridization conditions to nucleic acid sequences encoding a fragment of a wild-type mammalian IL-15 polypeptide of at least 20 contiguous amino acids, at least 30 contiguous amino acids, at least 40 contiguous amino acids, at least 50 contiguous amino acids, at least 100 contiguous amino acids, or at least 150 contiguous amino acids; (f) a fragment of a wild-type mammalian IL-15Ra polypeptide; and/or (g) a specific IL-15Ra derivative described herein. IL-15Ra derivatives also include a polypeptide that comprises the amino acid sequence of a mature form of mammalian IL-15Ra polypeptide and a heterologous signal peptide amino acid sequence. In a specific embodiment, an IL-15Ra derivative is a derivative of a wild-type human IL-15Ra polypeptide. In another embodiment, an IL-15Ra derivative is a derivative of an immature form of human IL-15 polypeptide. In another embodiment, an IL-15Ra derivative is a derivative of a mature form of human IL-15 polypeptide. In one embodiment, an IL-15Ra derivative is a soluble form of a mammalian IL-15Ra polypeptide. In other words, in certain

embodiments, an IL-15Ra derivative includes soluble forms of mammalian IL-15Ra, wherein those soluble forms are not naturally occurring. Other examples of IL-15Ra derivatives include the truncated, soluble forms of human IL-15Ra described herein. In a specific embodiment, an IL-15Ra derivative is purified or isolated.

In a preferred embodiment, IL-15Ra derivatives retain at least 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of a wild-type mammalian IL-15Ra polypeptide to bind an IL-15 polypeptide, as measured by assays well known in the art, e.g., ELISA, BIAcore^®, co-immunoprecipitation. In another preferred embodiment, IL-15Ra derivatives retain at least 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of a wild-type mammalian IL-15Ra polypeptide to induce IL- 15 -mediated signal transduction, as measured by assays well-known in the art, e.g., electromobility shift assays, ELISAs and other immunoassays. In a specific embodiment, IL-15Ra derivatives bind to IL-15 as assessed by methods well-known in the art, such as, e.g., ELISAs.

As used herein, the terms "IL-15Ra derivative" and "interleukin-15 receptor alpha derivative" in the context of nucleic acids refer to: (a) a nucleic acid sequence that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to the nucleic acid sequence encoding a mammalian IL-15Ra polypeptide; (b) a nucleic acid sequence encoding a polypeptide that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical the amino acid sequence of a wild-type mammalian IL-15Ra polypeptide; (c) a nucleic acid sequence that contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more nucleic acid mutations (i.e., additions, deletions and/or substitutions) relative to the nucleic acid sequence encoding a mammalian IL-15Ra polypeptide; (d) a nucleic acid sequence that hybridizes under high or medium stringency hybridization conditions to a nucleic acid sequence encoding a mammalian IL-15Ra polypeptide; (e) a nucleic acid sequence that hybridizes under high or medium stringency hybridization conditions to a fragment of a nucleic acid sequence encoding a mammalian IL-15Ra polypeptide; (f) a nucleic acid sequence encoding a fragment of a nucleic acid sequence encoding a mammalian IL-15Ra polypeptide; and/or (g) a nucleic acid sequence encoding a specific IL-15Ra derivative described herein. In a specific embodiment, an IL-15Ra derivative in the context of nucleic acids is a derivative of a nucleic acid sequence encoding a human IL-15Ra polypeptide. In another embodiment, an IL-15Ra derivative in the context of nucleic acids is a derivative of a nucleic acid sequence encoding an immature form of a human IL-15Ra polypeptide. In another embodiment, an IL-15Ra derivative in the context of nucleic acids is a derivative of a nucleic acid sequence encoding a mature form of a human IL-15Ra polypeptide. In one embodiment, an IL-15Ra derivative in the context of nucleic acids refers to a nucleic acid sequence encoding a derivative of mammalian IL-15Ra polypeptide that is soluble. In certain embodiments, an IL-15Ra derivative in context of nucleic acids refers to a nucleic acid sequence encoding a soluble form of mammalian IL-15Ra, wherein the soluble form is not naturally occurring. In some embodiments, an IL-15Ra derivative in the context of nucleic acids refers to a nucleic acid sequence encoding a derivative of human IL-15Ra, wherein the derivative of the human IL-15Ra is a soluble form of IL-15Ra that is not naturally occurring. In specific embodiments, an IL- 15 Ra derivative nucleic acid sequence is isolated or purified.

IL-15Ra derivative nucleic acid sequences include codon-optimized nucleic acid sequences that encode an IL-15Ra polypeptide, including mature and immature forms of IL-15Ra polypeptide. In other embodiments, IL-15Ra derivative nucleic acids include nucleic acids that encode IL-15Ra RNA transcripts containing mutations that eliminate potential splice sites and instability elements (e.g., A/T or A/U rich elements) without affecting the amino acid sequence to increase the stability of the IL-15Ra RNA transcripts. In certain embodiments, the IL-15Ra derivative nucleic acid sequence is the codon- optimized sequence in SEQ ID NO: 261 or 263 in Table 1 (the amino acid sequences encoded by such a nucleic acid sequences are provided in SEQ ID NO: 262 and 264 in Table 1, respectively).

In specific embodiments, IL-15Ra derivative nucleic acid sequences encode proteins or polypeptides that retain at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of a wild-type mammalian IL-15Ra polypeptide to bind IL-15, as measured by assays well known in the art, e.g., ELISA, BIAcore^®, co-immunoprecipitation. In another preferred embodiment, IL- 15Ra derivative nucleic acid sequences encode proteins or polypeptides that retain at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98% or 99% of the function of a wild-type mammalian IL- 15Ra to induce IL-15 -mediated signal transduction, as measured by assays well-known in the art, e.g., electromobility shift assays, ELISAs and other immunoassays. In a specific embodiment, IL-15Ra derivative nucleic acid sequences encode proteins or polypeptides that bind to IL-15 as assessed by methods well-known in the art, such as, e.g., ELISAs.

Described herein is the wild type soluble form of human IL-15Ra. Also described herein are specific IL-15Ra derivatives that are truncated, soluble forms of human IL-15Ra. These specific IL-15Ra derivatives and the soluble form of human IL-15Ra are based, in part, on the identification of the proteolytic cleavage site of human IL-15Ra. Further described herein are soluble forms of IL-15Ra that are characterized based upon glycosylation of the IL-15Ra.

The proteolytic cleavage of human IL-15Ra takes place between the residues (i.e., Glyl70 and His 171) which are in shown in bold and underlined in the provided amino acid sequence of the immature form of the wild-type full length human IL-15Ra:

MAPRRARGCRTLGLPALLLLLLLRPPATRGITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKR KAGTSSLTECVLNKATOVAHWTTPSLKCIRDPALVHQRPAPPSTVTTAGVTPQPESLSPSGKEPA ASSPSSNNTAATTAAIVPGSQLMPSKSPSTGTTEISSHESSHGTPSQTTAKNWELTASASHQPPGV YPOGHSDTTVAISTSTVLLCGLSAVSLLACYLKSROTPPLASVEMEAMEALPVTWGTSSRDEDLE NCSHHL (SEQ ID NO: 256 in Table 1).

Accordingly, in one aspect, provided herein is a soluble form of human IL-15Ra (e.g., a purified soluble form of human IL-15Ra), wherein the amino acid sequence of the soluble form of human IL-15Ra terminates at the site of the proteolytic cleavage of the wild-type membrane -bound human IL-15Ra. In particular, provided herein is a soluble form of human IL-15Ra (e.g., a purified soluble form of human IL-15Ra), wherein the amino acid sequence of the soluble form of human IL-15Ra terminates with PQG (SEQ ID NO: 270 in Table 1), wherein G is Glyl70. In a particular embodiment, provided herein is a soluble form of human IL-15Ra (e.g., a purified soluble form of human IL-15Ra) which has the amino acid sequence shown in SEQ ID NO: 257 in Table 1. In some embodiments, provided herein is an IL- 15Ra derivative (e.g., a purified and/or soluble form of IL-15Ra derivative), which is a polypeptide that: (i) is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to SEQ ID NO: 257 in Table 1; and (ii) terminates with the amino acid sequence PQG (SEQ ID NO: 270 in Table 1). In other particular embodiments, provided herein is a soluble form of human IL-15Ra (e.g., a purified soluble form of human IL-15Ra) which has the amino acid sequence of SEQ ID NO: 260 in Table 1). In some embodiments, provided herein is an IL-15Ra derivative (e.g., a purified and/or soluble form of an IL- 15Ra derivative), which is a polypeptide that is at least 75%, 80%, 85%, 90%, 95%, 98% or 99% identical to SEQ ID NO: 260 in Table 1, and, optionally, wherein the amino acid sequence of the soluble form of the IL-15Ra derivative terminates with PQG (SEQ ID NO: 270 in Table 1).

In some embodiments, provided herein is an IL-15Ra derivative of human IL-15Ra, wherein the IL-15Ra derivative is soluble and: (a) the last amino acids at the C-terminal end of the IL-15Ra derivative consist of amino acid residues PQGHSDTT (SEQ ID NO: 265 in Table 1); (b) the last amino acids at the C-terminal end of the IL-15Ra derivative consist of amino acid residues PQGHSDT (SEQ ID NO: 266 in Table 1); (c) the last amino acids at the C-terminal end of the IL-15Ra derivative consist of amino acid residues PQGHSD (SEQ ID NO: 267 in Table 1); (d) the last amino acids at the C-terminal end of the IL- 15Ra derivative consist of amino acid residues PQGHS (SEQ ID NO: 268 in Table 1); or (e) the last amino acids at the C-terminal end of the IL-15Ra derivative consist of amino acid residues PQGH (SEQ ID NO: 269 in Table 1). In certain embodiments, the amino acid sequences of these IL-15Ra derivatives are at least 75%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 271 in Table 1. In some embodiments, these IL-15Ra derivatives are purified.

In another aspect, provided herein are glycosylated forms of IL-15Ra (e.g., purified glycosylated forms of IL-15Ra), wherein the glycosylation of the IL-15Ra accounts for at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, or 20% to 25%, 20% to 30%, 25% to 30%, 25% to 35%, 30% to 35%, 30% to 40%, 35% to 40%, 35% to 45%, 40% to 50%, 45% to 50%, 20% to 40%, or 25% to 50% of the mass (molecular weight) of the IL-15Ra as assessed by techniques known to one of skill in the art. The percentage of the mass (molecular weight) of IL-15Ra (e.g., purified IL- 15Ra) that glycosylation of IL-15Ra accounts for can be determined using, for example and without limitation, gel electrophoresis and quantitative densitometry of the gels, and comparison of the average mass (molecular weight) of a glycosylated form of IL-15Ra (e.g., a purified glycosylated form of IL- 15Ra) to the non-glycosylated form of IL-15Ra (e.g., a purified non-glycosylated form of IL-15Ra). In one embodiment, the average mass (molecular weight) of IL-15Ra (e.g., purified IL-15Ra) can be determined using MALDI-TOF MS spectrum on Voyager De-Pro equipped with CovalX HM-1 high mass detector using sinapic acid as matrix, and the mass of a glycosylated form of IL-15Ra (e.g., purified glycosylated form of IL-15Ra) can be compared to the mass of the non-glycosylated form of IL-15Ra (e.g., purified non-glycosylated form of IL-15Ra) to determine the percentage of the mass that glycosylation accounts for.

In another aspect, provided herein are glycosylated forms of IL-15Ra, wherein the IL-15Ra is glycosylated (N- or O-glycosylated) at certain amino acid residues. In certain embodiments, provided herein is a human IL-15Ra which is glycosylated at one, two, three, four, five, six, seven, or all, of the following glycosylation sites: (i) O-glycosylation on threonine at position 5 of the amino acid sequence NWELTASASHQPPGVYPQG (SEQ ID NO: 272 in Table 1) in the IL-15Ra;

(ii) O-glycosylation on serine at position 7 of the amino acid sequence NWELTASASHQPPGVYPQG (SEQ ID NO: 272 in Table 1) in the IL-15Ra;

(iii) N-glycosylation on serine at position 8 of the amino acid sequence ITCPPPMSVEHADIWVK (SEQ ID NO: 273 in Table 1) in the IL-15Ra, or serine at position 8 of the amino acid sequence

ITCPPPMSVEHADIWVKSYSLYSRERYICNS (SEQ ID NO: 274 in Table 1) in the IL-15Ra;

(iv) N-glycosylation on Ser 18 of amino acid sequence ITCPPPMSVEHADIWVKSYSLYSRERYICNS (SEQ ID NO: 274 in Table 1) in the IL-15Ra;

(v) N-glycosylation on serine at position 20 of the amino acid sequence

ITCPPPMSVEHADIWVKSYSLYSRERYICNS (SEQ ID NO: 274 in Table 1) in the IL-15Ra;

(vi) N-glycosylation on serine at position 23 of the amino acid sequence

ITCPPPMSVEHADIWVKSYSLYSRERYICNS (SEQ ID NO: 274 in Table 1) in the IL-15Ra; and/or

(vii) N-glycosylated on serine at position 31 of the amino acid sequence

ITCPPPMSVEHADIWVKSYSLYSRERYICNS (SEQ ID NO: 274 in Table 1) in the IL-15Ra.

In specific embodiments, the glycosylated IL-15Ra is a wild-type human IL-15Ra. In other specific embodiments, the glycosylated IL-15Ra is an IL-15Ra derivative of human IL-15Ra. In some embodiments, the glycosylated IL-15Ra is a wild-type soluble human IL-15Ra, such as SEQ ID NO:257 or 260 in Table 1. In other embodiments, the glycosylated IL-15Ra is an IL-15Ra derivative that is a soluble form of human IL-15Ra. In certain embodiments, the glycosylated IL-15Ra is purified or isolated.

IL-15/IL-15Ra complex

As used herein, the term "IL-15/IL-15Ra complex" refers to a complex comprising IL-15 and IL- 15Ra covalently or noncovalently bound to each other. In a preferred embodiment, the IL-15Ra has a relatively high affinity for IL-15, e.g., KD of 10 to 50 pM as measured by a technique known in the art, e.g., KinEx A assay, plasma surface resonance (e.g., BIAcore^® assay). In another preferred embodiment, the IL-15/IL-15Ra complex induces IL-15 -mediated signal transduction, as measured by assays well- known in the art, e.g., electromobility shift assays, ELISAs and other immunoassays. In some embodiments, the IL-15/IL-15Ra complex retains the ability to specifically bind to the βγ chain. In a specific embodiment, the IL-15/IL-15Ra complex is isolated from a cell.

Provided herein are complexes that bind to the βγ subunits of the IL-15 receptor, induce IL-15 signal transduction (e.g., Jak/Stat signal transduction) and enhance IL-15 -mediated immune function, wherein the complexes comprise IL-15 covalently or noncovalently bound to interleukin-15 receptor alpha ("IL-15Ra") (a "IL-15/IL-15Ra complex"). The IL-15/IL-15Ra complex is able to bind to the βγ receptor complex. The IL-15/IL-15Ra complexes may be composed of wild-type IL-15 or an IL-15 derivative and wild-type IL-15Ra or an IL-15Ra derivative. In certain embodiments, an IL-15/IL-15Ra complex comprises IL-15 or an IL-15 derivative and an IL-15Ra described above. In a specific embodiment, an IL-15/IL-15Ra complex comprises IL-15 or an IL-15 derivative and IL-15Ra with the amino acid sequence of SEQ ID NO: 260 in Table 1. In another embodiment, an IL-15/IL-15Ra complex comprises IL-15 or an IL-15 derivative and a glycosylated form of IL-15Ra described supra.

In a specific embodiment, an IL-15/IL-15Ra complex comprises wild-type IL-15 or an IL-15Ra derivative and soluble IL-15Ra (e.g., wild-type soluble human IL-15Ra). In another specific embodiment, an IL-15/IL-15Ra complex is composed of an IL-15 derivative and an IL-15Ra derivative. In another embodiment, an IL-15/IL-15Ra complex is composed of wild-type IL-15 and an IL-15Ra derivative. In one embodiment, the IL-15Ra derivative is a soluble form of IL-15Ra. Specific examples of soluble forms of IL-15Ra are described above. In a specific embodiment, the soluble form of IL-15Ra lacks the transmembrane domain of wild-type IL-15Ra, and optionally, the intracellular domain of wild- type IL-15Ra. In another embodiment, the IL-15Ra derivative is the extracellular domain of wild-type IL-15Ra or a fragment thereof. In certain embodiments, the IL-15Ra derivative is a fragment of the extracellular domain comprising the sushi domain or exon 2 of wild-type IL-15Ra. In some

embodiments, the IL-15Ra derivative comprises a fragment of the extracellular domain comprising the sushi domain or exon 2 of wild-type IL-15Ra and at least one amino acid that is encoded by exon 3. In certain embodiments, the IL-15Ra derivative comprises a fragment of the extracellular domain comprising the sushi domain or exon 2 of wild-type IL-15Ra and an IL-15Ra hinge region or a fragment thereof. In certain embodiments, the IL-15Ra comprises the amino acid sequence of SEQ ID NO: 260 in Table 1.

In another embodiment, the IL-15Ra derivative comprises a mutation in the extracellular domain cleavage site that inhibits cleavage by an endogenous protease that cleaves wild-type IL-15Ra. In a specific embodiment, the extracellular domain cleavage site of IL-15Ra is replaced with a cleavage site that is recognized and cleaved by a heterologous known protease. Non-limiting examples of such heterologous protease cleavage sites include Arg-X-X-Arg (SEQ ID NO: 275 in Table 1), which is recognized and cleaved by furin protease; and A-B-Pro-Arg-X-Y (SEQ ID NO: 276 in Table 1) (A and B are hydrophobic amino acids and X and Y are non-acidic amino acids) and Gly-Arg-Gly, which are recognized and cleaved by thrombin protease.

In another embodiment, the IL-15 is encoded by a nucleic acid sequence optimized to enhance expression of IL-15, e.g., using methods as described in PCT Publications WO 2007/084342 and WO 2010/020047; and U.S. Patent Nos. 5,965,726; 6, 174,666; 6,291,664; 6,414,132; and 6,794,498.

In certain embodiments, provided herein is an IL-15/IL-15Ra complex comprising human IL- 15Ra which is glycosylated at one, two, three, four, five, six, seven, or all, of the glycosylation sites as described supra and with reference to SEQ ID NOs: 272, 273 and 274 in Table 1. In specific embodiments, the glycosylated IL-15Ra is a wild-type human IL-15Ra. In other specific embodiments, the glycosylated IL-15Ra is an IL-15Ra derivative of human IL-15Ra. In some embodiments, the glycosylated IL-15Ra is a wild-type soluble human IL-15Ra, such as SEQ ID NO: 257 or 260 in Table 1. In other embodiments, the glycosylated IL-15Ra is an IL-15Ra derivative that is a soluble form of human IL-15Ra. In certain embodiments, the IL-15/IL-15Ra complex is purified or isolated.

In addition to IL-15 and IL-15Ra, the IL-15/IL-15Ra complexes may comprise a heterologous molecule. In some embodiments, the heterologous molecule increases protein stability. Non-limiting examples of such molecules include polyethylene glycol (PEG), Fc domain of an IgG immunoglobulin or a fragment thereof, or albumin that increase the half-life of IL-15 or IL-15Ra in vivo. In certain embodiments, IL-15Ra is conjugated/fused to the Fc domain of an immunoglobulin (e.g., an IgGl) or a fragment thereof. In a specific embodiment, the IL-15RaFc fusion protein comprises the amino acid sequence of SEQ ID NO: 277 or 278 in Table 1. In another embodiment, the IL-15RaFc fusion protein is the IL-15Ra/Fc fusion protein described in Han et al, (2011), Cytokine 56: 804-810, U.S. Patent No. 8,507,222 or U.S. Patent No. 8, 124,084. In those IL-15/IL-15Ra complexes comprising a heterologous molecule, the heterologous molecule may be conjugated to IL-15 and/or IL-15Ra. In one embodiment, the heterologous molecule is conjugated to IL-15Ra. In another embodiment, the heterologous molecule is conjugated to IL-15.

The components of an IL-15/IL-15Ra complex may be directly fused, using either non-covalent bonds or covalent bonds (e.g., by combining amino acid sequences via peptide bonds), and/or may be combined using one or more linkers. Linkers suitable for preparing the IL-15/IL-15Ra complexes comprise peptides, alkyl groups, chemically substituted alkyl groups, polymers, or any other covalently- bonded or non-covalently bonded chemical substance capable of binding together two or more components. Polymer linkers comprise any polymers known in the art, including polyethylene glycol (PEG). In some embodiments, the linker is a peptide that is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more amino acids long. In a specific embodiment, the linker is long enough to preserve the ability of IL-15 to bind to the IL-15Ra. In other embodiments, the linker is long enough to preserve the ability of the IL-15/IL-15Ra complex to bind to the βγ receptor complex and to act as an agonist to mediate IL-15 signal transduction.

In particular embodiments, IL-15/IL-15Ra complexes are pre-coupled prior to use in the methods described herein (e.g., prior to contacting cells with the IL-15/IL-15Ra complexes or prior to

administering the IL-15/IL-15Ra complexes to a subject). In other embodiments, the IL-15/IL-15Ra complexes are not pre-coupled prior to use in the methods described herein.

In a specific embodiment, an IL-15/IL-15Ra complex enhances or induces immune function in a subject by at least 99%, at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 60%, at least 50%, at least 45%, at least 40%, at least 45%, at least 35%, at least 30%, at least 25%, at least 20%, or at least 10% relative to the immune function in a subject not administered the IL- 15/IL-15Ra complex using assays well known in the art, e.g., ELISPOT, ELISA, and cell proliferation assays. In a specific embodiment, the immune function is cytokine release (e.g., interferon-gamma, IL-2, IL-5, IL-10, IL-12, or transforming growth factor (TGF) -beta). In one embodiment, the IL-15 mediated immune function is NK cell proliferation, which can be assayed, e.g., by flow cytometry to detect the number of cells expressing markers of NK cells (e.g., CD56). In another embodiment, the IL-15 mediated immune function is antibody production, which can be assayed, e.g., by ELISA. In some embodiments, the IL-15 mediated immune function is effector function, which can be assayed, e.g., by a cytotoxicity assay or other assays well known in the art.

Methods of producing antibodies and polypeptides of the combination

Also provided in the invention are expression vectors and host cells for producing the CD32b- binding antibodies and IL-15/IL-15Ra complexes of the combination, as described above. Various expression vectors can be employed to express the polynucleotides encoding the antibody, IL-15 and IL- 15Ra polypeptides. Both viral -based and nonviral expression vectors can be used to produce the polypeptides in a mammalian host cell. Non-viral vectors and systems include plasmids, episomal vectors, typically with an expression cassette for expressing a protein or RNA, and human artificial chromosomes (see, e.g., Harrington et al., (1997) Nat Genet. 15:345). For example, nonviral vectors useful for expression of polypeptides in mammalian (e.g., human) cells include pThioHis A, B & C, pcDNA3.1/His, pEBVHis A, B & C, (Invitrogen, San Diego, Calif), MPSV vectors, and numerous other vectors known in the art for expressing other proteins. Useful viral vectors include vectors based on retroviruses, adenoviruses, adenoassociated viruses, herpes viruses, vectors based on SV40, papilloma virus, HBP Epstein Barr virus, vaccinia virus vectors and Semliki Forest virus (SFV). See, Brent et al, supra; Smith, (1995) Annu. Rev. Microbiol. 49:807; and Rosenfeld et al., (1992) Cell 68: 143.

The choice of expression vector depends on the intended host cells in which the vector is to be expressed. Typically, the expression vectors contain a promoter and other regulatory sequences (e.g., enhancers) that are operably linked to the polynucleotides. In one embodiment, an inducible promoter is employed to prevent expression of inserted sequences except under inducing conditions. Inducible promoters include, e.g., arabinose, lacZ, metallothionein promoter or a heat shock promoter. Cultures of transformed organisms can be expanded under noninducing conditions without biasing the population for coding sequences whose expression products are better tolerated by the host cells. In addition to promoters, other regulatory elements may also be required or desired for efficient expression. These elements typically include an ATG initiation codon and adjacent ribosome binding site or other sequences. In addition, the efficiency of expression may be enhanced by the inclusion of enhancers appropriate to the cell system in use (see, e.g., Scharf et al., (1994) Results Probl. Cell Differ. 20: 125; and Bittner et al., (1987) Meth. Enzymol., 153:516). For example, the SV40 enhancer or CMV enhancer may be used to increase expression in mammalian host cells.

The expression vectors may also provide a secretion signal sequence position to form a fusion protein with polypeptides encoded by inserted CD32b-binding antibody sequences or IL-15 or IL-15Ra sequences. More often, the inserted sequences are linked to a signal sequences before inclusion in the vector. Vectors to be used to receive sequences encoding CD32b-binding antibody light and heavy chain variable domains sometimes also encode constant regions or parts thereof. Such vectors allow expression of the variable regions as fusion proteins with the constant regions thereby leading to production of intact antibodies and antigen-binding fragments thereof. Typically, such constant regions are human.

The host cells for harboring and expressing the CD32b-binding antibody chains and the IL-15 and IL-15Ra proteins can be either prokaryotic or eukaryotic. E. coli is one prokaryotic host useful for cloning and expressing the polynucleotides of the present invention. Other microbial hosts suitable for use include bacilli, such as Bacillus subtilis, and other enterobacteriaceae, such as Salmonella, Serratia, and various Pseudomonas species. In these prokaryotic hosts, one can also make expression vectors, which typically contain expression control sequences compatible with the host cell (e.g., an origin of replication). In addition, any number of a variety of well-known promoters will be present, such as the lactose promoter system, a tryptophan (trp) promoter system, a beta-lactamase promoter system, or a promoter system from phage lambda. The promoters typically control expression, optionally with an operator sequence, and have ribosome binding site sequences and the like, for initiating and completing transcription and translation. Other microbes, such as yeast, can also be employed to express polypeptides of the invention. Insect cells in combination with baculovirus vectors can also be used.

In one embodiment, mammalian host cells are used to express and produce the polypeptides of the present invention. For example, they can be either a hybridoma cell line expressing endogenous immunoglobulin genes or a mammalian cell line harboring an exogenous expression vector. These include any normal mortal or normal or abnormal immortal animal or human cell. For example, a number of suitable host cell lines capable of secreting intact immunoglobulins have been developed including the CHO cell lines, various Cos cell lines, HeLa cells, myeloma cell lines, transformed B-cells and hybridomas. Examples of mammalian cell lines include, but are not limited to, COS, CHO, HeLa, NIH3T3, HepG2, MCF7, HEK 293, HEK 293T, RD, PC 12, hybridomas, pre-B cells, 293, 293H, K562, SkBr3, BT474, A204, M07Sb, ΤΤβΙ, Raji, Jurkat, MOLT-4, CTLL-2, MC-IXC, SK-N-MC, SK-N-MC, SK-N-DZ, SH-SY5Y, C127, NO, and BE(2)-C cells. Other mammalian cell lines available as hosts for expression are known in the art and include many immortalized cell lines available from the American Type Culture Collection (ATCC). The use of mammalian tissue cell culture to express polypeptides is discussed generally in, e.g., Winnacker, FROM GENES TO CLONES, VCH Publishers, N.Y., N.Y., 1987.

Expression vectors for mammalian host cells can include expression control sequences, such as an origin of replication, a promoter, and an enhancer (see, e.g., Queen, et al, (1986) Immunol. Rev. 89:49-68,), and necessary processing information sites, such as ribosome binding sites, RNA splice sites, polyadenylation sites, and transcriptional terminator sequences. These expression vectors usually contain promoters derived from mammalian genes or from mammalian viruses. Suitable promoters may be constitutive, cell type -specific, stage -specific, and/or modulatable or regulatable. Useful promoters include, but are not limited to, the metallothionein promoter, the constitutive adenovirus major late promoter, the dexamethasone-inducible MMTV promoter, the SV40 promoter, the MRP poIIII promoter, the constitutive MPSV promoter, the tetracycline-inducible CMV promoter (such as the human immediate -early CMV promoter), the constitutive CMV promoter, and promoter-enhancer combinations known in the art.

Methods for introducing expression vectors containing the polynucleotide sequences of interest vary depending on the type of cellular host. For example, calcium chloride transfection is commonly utilized for prokaryotic cells, whereas calcium phosphate treatment or electroporation may be used for other cellular hosts. (See generally Sambrook et al., supra). Other methods include, e.g., electroporation, calcium phosphate treatment, liposome-mediated transformation, injection and microinjection, ballistic methods, virosomes, immunoliposomes, polycation:nucleic acid conjugates, naked DNA, artificial virions, fusion to the herpes virus structural protein VP22 (Elliot & O'Hare, (1997) Cell 88:223), agent- enhanced uptake of DNA, and ex vivo transduction.

For long-term, high-yield production of recombinant IL-15 and IL-15Ra polypeptides and/or an anti-CD32b antibody molecule, stable cell lines can be generated. For example, cell lines can be transformed using the nucleic acid constructs described herein which may contain a selectable marker gene on the same or on a separate nucleic acid construct. The selectable marker gene can be introduced into the same cell by co-transfection. Following the introduction of the vector, cells are allowed to grow for 1-2 days in an enriched media before they are switched to selective media to allow growth and recovery of cells that successfully express the introduced nucleic acids. Resistant clones of stably transformed cells may be proliferated using tissue culture techniques well known in the art that are appropriate to the cell type. In a particular embodiment, the cell line has been adapted to grow in serum- free medium. In one embodiment, the cell line has been adapted to grow in serum-free medium in shaker flasks. In one embodiment, the cell line has been adapted to grow in stir or rotating flasks. In certain embodiments, the cell line is cultured in suspension. In particular embodiments, the cell line is not adherent or has been adapted to grow as nonadherent cells. In certain embodiments, the cell line has been adapted to grow in low calcium conditions. In some embodiments, the cell line is cultured or adapted to grow in low serum medium.

In a specific embodiment, a particularly preferred method of high-yield production of a recombinant polypeptide of the present invention is through the use of dihydro folate reductase (DHFR) amplification in DHFR-deficient CHO cells, by the use of successively increasing levels of methotrexate as described in U.S. Patent No. 4,889,803. The polypeptide obtained from such cells may be in a glycosylated form.

In one embodiment, cell lines are engineered to express the stable heterodimer of wild-type human IL-15 and wild-type soluble human IL-15Ra, which can then be purified, and administered to a human. In one embodiment, the stability of the IL-15/IL-15Ra heterodimer is increased when produced from cell lines recombinantly expressing both IL-15 and IL-15Ra.

In a specific embodiment, the host cell recombinantly expresses IL-15 and the full length IL- 15Ra. In another specific embodiment, the host cell recombinantly expresses IL-15 and the soluble form of IL-15Ra. In another specific embodiment, the host cell recombinantly expresses IL-15 and a membrane -bound form of IL-15Ra which is not cleaved from the surface of the cell and remains cell associated. In some embodiments, the host cell recombinantly expressing IL-15 and/or IL-15Ra (full- length or soluble form) also recombinantly expresses another polypeptide (e.g., a cytokine or fragment thereof).

In certain embodiments, such a host cell recombinantly expresses an IL-15 polypeptide in addition to an IL-15Ra polypeptide. The nucleic acids encoding IL-15 and/or IL-15Ra can be used to generate mammalian cells that recombinantly express IL-15 and IL-15Ra in high amounts for the isolation and purification of IL-15 and IL-15Ra, preferably the IL-15 and the IL-15Ra are associated as complexes. In one embodiment, high amounts of IL-15/IL-15Ra complexes refer to amounts of IL-15/IL- 15Ra complexes expressed by cells that are at least 1 fold, 2 fold, 3 fold, 4 fold, 5 fold, 6 fold, 7 fold, 8 fold, 9 fold, 10 fold, 20 fold, or more than 20 fold higher than amounts of IL-15/IL-15Ra complexes expressed endogenously by control cells (e.g., cells that have not been genetically engineered to recombinantly express IL-15, IL-15Ra, or both IL-15 and IL-15Ra, or cells comprising an empty vector). In some embodiments, a host cell described herein expresses approximately 0.1 pg to 25 pg, 0.1 pg to 20 pg, 0.1 pg to 15 pg, 0.1 pg to 10 pg, 0.1 pg to 5 pg, 0.1 pg to 2 pg, 2 pg to 10 pg, or 5 to 20 pg of IL-15 as measured by a technique known to one of skill in the art (e.g., an ELISA). In certain embodiments, a host cell described herein expresses approximately 0.1 to 0.25 pg per day, 0.25 to 0.5 pg per day, 0.5 to 1 pg per day, 1 to 2 pg per day, 2 to 5 pg per day, or 5 to 10 pg per day of IL-15 as measured by a technique known to one of skill in the art (e.g., an ELISA). In a specific embodiment, the IL-15Ra is the soluble form of IL-15Ra. In a specific embodiment, the IL-15Ra is the soluble form of IL-15Ra associated with IL-15 in a stable heterodimer, which increases yields and simplifies production and purification of bioactive heterodimer IL-15/soluble IL-15Ra cytokine.

Recombinant IL-15 and IL-15Ra and an anti-CD32b antibody molecule can be purified using methods of recombinant protein production and purification are well known in the art, e.g., see PCT Publication WO 2007/070488. Briefly, the polypeptide can be produced intracellularly, in the periplasmic space, or directly secreted into the medium. Cell lysate or supernatant comprising the polypeptide can be purified using, for example, hydroxyapatite chromatography, gel electrophoresis, dialysis, and affinity chromatography. Other techniques for protein purification such as fractionation on an ion-exchange column, ethanol precipitation, Reverse Phase HPLC, chromatography on silica, chromatography on heparin SEPHAROSE™ (gel filtration substance; Pharmacia Inc., Piscataway, New Jersey) chromatography on an anion or cation exchange resin (such as a polyaspartic acid column), chromatofocusing, SDS-PAGE, and ammonium sulfate precipitation are also available.

In some embodiments, IL-15 and IL-15Ra are synthesized or recombinantly expressed by different cells and subsequently isolated and combined to form an IL-15/IL-15Ra complex, in vitro, prior to administration to a subject. In other embodiments, IL-15 and IL-15Ra are synthesized or

recombinantly expressed by different cells and subsequently isolated and simultaneously administered to a subject an IL-15/IL-15Ra complex in situ or in vivo. In yet other embodiments, IL-15 and IL-15Ra are synthesized or expressed together by the same cell, and the IL-15/IL-15Ra complex formed is isolated.

Prophylactic and Therapeutic uses

The present invention provides methods of treating a disease or disorder associated with increased CD32b activity or expression by administering to a subject in need thereof an effective amount of an anti-CD32b antibody molecule in combination with an IL-15/IL-15Ra complex. In a specific embodiment, the present invention provides a method of treating indications including, but not limited to, B cell malignancies including Hodgkins lymphoma, Non-Hodgkins lymphoma, multiple myeloma, diffuse large B cell lymphoma, acute lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse small cleaved cell lymphoma, MALT lymphoma, mantel cell lymphoma, marginal zone lymphoma and follicular lymphoma as well as other diseases and conditions including systemic light chain amyloidosis, haematological malignancies and other solid tumors.

In one embodiment, the present invention provides methods of treating a CD32b-related disease or disorder by administering to a subject in need thereof an effective amount of an anti-CD32b antibody molecule in combination with an IL-15/IL-15Ra complex. Examples of known CD32b related diseases or disorders for which the disclosed combination may be useful include but are not limited to: B cell malignancies including Hodgkins lymphoma, Non-Hodgkins lymphoma, multiple myeloma, diffuse large B cell lymphoma, acute lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse small cleaved cell lymphoma, MALT lymphoma, mantel cell lymphoma, marginal zone lymphoma and follicular lymphoma as well as other diseases and conditions including systemic light chain amyloidosis, haematological malignancies and other solid tumors.

Furthermore, the combination of an anti-CD32b antibody molecule with an IL-15/IL-15Ra complex can be used, inter alia, to treat, prevent, delay or reverse disease progression of patients who have become resistant or refractory to antibody treatment. By administering the CD32b-binding antibody molecules, disclosed herein, in combination with an IL-15/IL-15Ra complex, the efficacy of the ADCC effect of the antibody may be enhanced, in full or in part.

In a further embodiment, the combination of an anti-CD32b antibody molecule with an IL-15/IL- 15Ra complex described herein, can be administered to a patient in need thereof in conjunction with another therapeutic agent as discussed below. For example, the combination of the present invention can be co-formulated with, and/or co-administered with, one or more additional therapeutic agents, e.g., one or more anti-cancer agents, cytotoxic or cytostatic agents, hormone treatment, vaccines, and/or other immunotherapies. In other embodiments, the combination can be administered with other therapeutic treatment modalities, including surgery, radiation, cryosurgery, and/or thermotherapy. Such combination therapies may advantageously utilize lower dosages of the administered therapeutic agents, thus avoiding possible toxicities or complications associated with the various monotherapies. As will be appreciated by the skilled artisan, therapies utilizing the combination of the present invention may be administered in conjunction with multiple classes of the agents described above. When the combination of the present invention is administered together with another agent or agents, the two (or more) can be administered sequentially in any order, or simultaneously. In some aspects, the combination of the present invention is administered to a subject who is also receiving therapy with one or more other agents or methods. In other aspects, the combination is administered in conjunction with surgical treatments. The therapy regimen may be additive, or it may produce synergistic results.

Pharmaceutical Compositions

The invention provides pharmaceutical compositions comprising the combination of an anti- CD32b antibody molecule with an IL-15/IL-15Ra complex formulated together or separately with a pharmaceutically acceptable carrier. The anti-CD32b antibody molecule and the IL-15/IL-15Ra complex can be administered to a patient as a "non-fixed combination" meaning that the anti-CD32b antibody molecule and IL-15/IL-15Ra complex are administered as separate entities either simultaneously, concurrently or sequentially with no specific time limits, wherein such administration provides therapeutically effective levels of the two agents in the body of the patient. The term "non-fixed combination" thus defines especially a "kit of parts" in the sense that the combination agents (i) an anti- CD32b antibody molecule and (ii) an IL-15/IL-15Ra complex as defined herein can be dosed

independently of each other or by use of different fixed combinations with distinguished amounts of the combination agents, i.e. simultaneously or at different time points, where the combination agents may also be used as entirely separate pharmaceutical dosage forms or pharmaceutical formulations that are also sold independently of each other and where instructions for the possibility of their combined use is or are provided in the package equipment, e.g. leaflet or the like, or in other information e.g. provided to physicians and medical staff. The independent formulations or the parts of the kit of parts can then, e.g. be administered simultaneously or chronologically staggered, that is at different time points and with equal or different time intervals for any part of the kit of parts. In a specific embodiment, the time intervals are chosen such that the effect on the treated disease in the combined use of the parts is larger than the effect which would be obtained by use of only any one of the combination agents (i) and (ii), thus being jointly active. The ratio of the total amounts of the combination agent (i) to the combination agent (ii) to be administered in the combined preparation can be varied, e.g. in order to cope with the needs of a patient sub-population to be treated or the needs of the single patient which different needs can be due to age, sex, body weight, etc. of the patients.

A pharmaceutical composition of the present invention can additionally contain one or more other therapeutic agents that are suitable for treating or preventing a CD32b-associated disease (e.g., B cell malignancies including Hodgkins lymphoma, Non-Hodgkins lymphoma, multiple myeloma, diffuse large

B cell lymphoma, acute lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse small cleaved cell lymphoma, MALT lymphoma, mantel cell lymphoma, marginal zone lymphoma and follicular lymphoma as well as other diseases and conditions including systemic light chain amyloidosis, haematological malignancies and other solid tumors).

A pharmaceutical composition of the present invention can be administered with a

pharmaceutically acceptable carrier to enhance or stabilize the composition, or facilitate preparation of the composition. Pharmaceutically acceptable carriers include solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible.

A pharmaceutical composition of the present invention can be administered by a variety of methods known in the art. The route and/or mode of administration may vary depending upon the desired results. Administration can be intravenous, intramuscular, intraperitoneal, or subcutaneous, or administered proximal to the site of the target. The pharmaceutically acceptable carrier should be suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal or epidermal administration (e.g., by injection or infusion). Depending on the route of administration, the active compound, i.e., anti-CD32b antibody molecule and/or IL-15/IL-15Ra complex, may be coated in a material to protect the compound from the action of acids and other natural conditions that may inactivate the compound.

The composition should be sterile and fluid. Fluidity can be maintained, for example, by use of coating such as lecithin, by maintenance of required particle size in the case of dispersion and by use of surfactants. In many cases, it is preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol or sorbitol, and sodium chloride in the composition. Long-term absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate or gelatin.

Pharmaceutical compositions of the invention can be prepared in accordance with methods well known and routinely practiced in the art. See, e.g., Remington: The Science and Practice of Pharmacy, Mack Publishing Co., 20th ed., (2000); and Sustained and Controlled Release Drug Delivery Systems, J. R. Robinson, ed., Marcel Dekker, Inc., New York, (1978). Pharmaceutical compositions are preferably manufactured under GMP conditions. Typically, a therapeutically effective dose or efficacious dose of the CD32b-binding antibody molecule and the IL-15/IL-15Ra complex, of the combination, is employed in pharmaceutical compositions of the invention. The anti-CD32b antibody molecules and IL-15/IL-15Ra complex are formulated into pharmaceutically acceptable dosage forms by conventional methods known to those of skill in the art. Dosage regimens are adjusted to provide the optimum desired response (e.g., a therapeutic response). For example, for each component of the combination, a single bolus may be administered, several divided doses may be administered over time or the dose may be proportionally reduced or increased as indicated by the requirements of the therapeutic situation. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the subjects to be treated; each unit contains a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. Actual dosage levels of the active ingredients in the pharmaceutical compositions of the present invention can be varied so as to obtain an amount of the active ingredient which is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient. The selected dosage level depends upon a variety of pharmacokinetic factors including the activity of the particular components of the combination of the present invention employed, the route of administration, the time of administration, the rate of excretion of the particular compound being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compositions employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors.

A physician can start doses of the combination of the invention employed in the pharmaceutical composition at levels lower than that required to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved. In general, effective doses of the compositions of the present invention, for the treatment of an disorder described herein vary depending upon many different factors, including means of administration, target site, physiological state of the patient, other medications administered, and whether treatment is prophylactic or therapeutic. Treatment dosages need to be titrated to optimize safety and efficacy. For systemic administration with an anti-CD32b antibody molecule, the dosage ranges from about 0.0001 to 100 mg/kg, and more usually 0.01 to 15 mg/kg, of the host body weight. An exemplary treatment regime entails systemic administration once every two weeks or once a month or once every 3 to 6 months. For subcutaneous administration of the IL-15/IL-15Ra complex, the dose ranges from about 0.25 to 8 μg/kg/day. An exemplary treatment regime entails subcutaneous administration in a treatment cycle of three times a week for two weeks, followed by a two week break before a repeat of the treatment cycle.

For a combination of the present invention comprising an anti-CD32b antibody molecule with an IL-15/IL-15Ra complex, the antibody and/or IL-15/IL-15Ra complex may be administered on multiple occasions. Intervals between single dosages can be weekly, monthly or yearly. Intervals can also be irregular as indicated by measuring blood levels of CD32b-binding antibody and/or IL-15 in the patient. In some methods of systemic administration of the anti-CD32b antibody, dosage is adjusted to achieve a plasma antibody concentration of 1-1000 μg/ml and in some methods 25-500 μg/ml. In some methods of subcutaneous administration of the IL-15/IL-15Ra complex, dosage is escalated until an y adverse events or dose limiting toxicity are observed. Alternatively, the components of the combination can be administered as a sustained release formulation, in which case less frequent administration is required. Dosage and frequency vary depending on the half-life of the antibody and complex in the patient. The dosage and frequency of administration can vary depending on whether the treatment is prophylactic or therapeutic. In prophylactic applications, a relatively low dosage is administered at relatively infrequent intervals over a long period of time. Some patients continue to receive treatment for the rest of their lives. In therapeutic applications, a relatively high dosage at relatively short intervals is sometimes required until progression of the disease is reduced or terminated, and preferably until the patient shows partial or complete amelioration of symptoms of disease. Thereafter, the patient can be administered a prophylactic regime.

Specific Embodiments, Citation and references

The present invention is not to be limited in scope by the specific embodiments described herein. Indeed, various modifications of the invention in addition to those described herein will become apparent to those skilled in the art from the foregoing description and accompanying figures. Such modifications are intended to fall within the scope of the appended claims.

Various references, including patent applications, patents, and scientific publications, are cited herein; the disclosure of each such reference is hereby incorporated herein by reference in its entirety.

EXAMPLES

The following examples are provided to further illustrate the invention but not to limit its scope. Other variants of the invention will be readily apparent to one of ordinary skill in the art and are encompassed by the appended claims.

EXAMPLE 1: Target cell lysis mediated by an anti-CD32b antibody in combination with an IL- 15/IL-15Ra complex

NK cells are among the most potent effector cells for the ADCC activity of anti-CD32b antibodies. It is believed that the IL-15/IL-15Ra complex "hetIL-15" can augment their ADCC activity by: 1) increasing effectortarget cell ratio as a result of NK cell proliferation; and 2) enhancing effector cell function, such as release of perforin and granzyme, as well as secretion of IFNy. To demonstrate the enhanced effector cell function as in 2) an ADCC assay to measure target cell lysis mediated by the anti- CD32b human IgGl (NOV2108) in combination with hetIL-15 was performed as detailed below.

PBMCs were isolated from a Leukopak (HemaCare catalog# PB001F-3) via a ficoll gradient. Primary NK cells were then negatively selected from PBMCs using Miltenyi beads (catalog# 130-050- 101). Isolated NK cells were incubated with lOOpg/ml of rhIL-2 overnight (Peprotech, #200-02) and then used in the ADCC assay at an appropriate E:T ratio. Luciferized Daudi cells were used as target cells and primary NK cells were used as effector cells; and cells were co-cultured for 4 hr either in the absence or presence of hetIL-15. Following the co-incubation, Bright Glo (Promega, catalog# E2620; 60 μΐ) was added to all wells, with the exception of the appropriate control wells and the luminescence signal was subsequently measured on an Envision (Perkin Elmer).

NOV2108 demonstrated Fc- and dose- dependent ADCC activity against Daudi cells. Addition of hetIL-15 enhanced the potency of both the Fc wild type and Fc -enhanced NOV2108 (afucosylated) in the ADCC assay (Figure 1). The ADCC enhancement by hetIL-15 was evident at concentrations as low as 0.01 and 0.1 ng/ml, and at lng/ml similar ADCC activity as with 0.1 ng/ml hetIL15 was observed. No effect on ADCC activity against Daudi cells was observed with an Fc silent version of the NOV2108 antibody having the modification N297A, with or without the addition of hetIL-15. The EC50 of each antibody in two experiments with different donor NK cells is shown in Table 2, where the increase in potency was reflected by the lower EC50 values.

Table 2: EC50 values of anti-CD32b antibody NOV2108-mediated ADCC activity against Daudi in combination with hetIL-15

EXAMPLE 2: Assay for IFNy expression mediated by an anti-CD32b antibody in combination with an IL-15/IL-15Ra complex

To further demonstrate an enhanced effector cell function of NK cells mediated by anti-CD32b antibodies, an IL-15/IL-15Ra complex (hetIL-15) can be included in an NK cell killing assay. In this NK cell killing assay, NK cells can be pre-treated with hetIL-15, and cultured with Daudi cells and anti- CD32b antibody, and the resulting level of IFNy measured. In a second setting, NK cells can be cultured with Daudi cells, hetIL15 and anti-CD32b, and the resulting level of IFNy measured.

Microplates containing the capture antibody can be first prepared according to the following method. The Capture Antibody (R&D Systems, Cat. no: 840101) is diluted to a working concentration in PBS without carrier protein and used immediately to coat a 96-well microplate (R&D Systems, Cat. No: DY990) with ΙΟΟμΙ per well of the diluted Capture Antibody. The plate is then sealed and incubated overnight at room temperature. After incubation, each well is aspirated and washed with wash buffer twice (0.05% Tween^® 20 in PBS, pH 7.2-7.4), for a total of three washes. The plates are then blocked by adding 300 μΐ of block buffer (1% BSA in PBS, pH 7.2-7.4, 0.2 um filtered) to each well and incubated at room temperature for a minimum of 1 hour. The aspiration/wash step as described above should be repeated.

To perform the cell killing assay, ΙΟΟμΙ of sample or standard (R&D Systems, Cat. No: 840103) in Reagent Diluent (0.1% BSA, 0.05% Tween 20 in Tris-buffered Saline ) or another appropriate diluent, is added to each well, incubated for 2 hours and then aspirated/washed as described above. ΙΟΟμΙ of

Detection Antibody (R&D Systems, Cat. No: 840102), diluted in Reagent Diluent with NGS, is then added to each well, incubated for 2 hours and then aspirated/washed as described above. ΙΟΟμΙ of a working dilution of Streptavidin-HRP (R&D Systems, Cat. No: 893975) is then added to each well, incubated for 20 min at room temperature and then aspirated/washed as described above. lOOul of

Substrate Solution (1 : 1 mixture of Color Reagent A (H202) and Color Reagent B (Tetramethylbenzidine) (R&D Systems, Cat. No: DY999)) is then added to each well and the plate incubated for 20 min at room temperature. 50ul of Stop Solution (2 N H₂SO₄) is then added to each well, tapping the plate to mix thoroughly.

To measure IFNy production, the optical density of each well should be determined immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate as readings made directly at 450 nm without correction may be higher and less accurate.

TABLE 1. Sequence Table

9 LCDR2 QDSKRPS

(Kabat)

10 LCDR3 GATDLSPWSIV

(Kabat)

11 VL DIELTQPPSVSVSPGETASITCSGDKLGDYYVHWYQQKPGQAPVLVIYQDSKRP

SGIPERFSGSNSGNTATLTISGTQAEDEADYYCGATDLSPWSIVFGGGTKLTVL

12 DNA VL GATATCGAGCTGACTCAGCCCCCTAGCGTCAGCGTCAGCCCTGGCGAGACAGCC

TCTATCACCTGTAGCGGCGATAAGCTGGGCGACTACTACGTGCACTGGTATCAG CAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTATCAGGACTCTAAGCGGCCT AGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCGCTACCCTG ACTATTAGCGGCACTCAGGCCGAGGACGAGGCCGACTACTACTGCGGCGCTACC GACCTGAGCCCCTGGTCTATCGTGTTCGGCGGAGGCACTAAGCTGACCGTGCTG

13 Light DIELTQPPSVSVSPGETASITCSGDKLGDYYVHWYQQKPGQAPVLVIYQDSKRP

Chain SGIPERFSGSNSGNTATLTISGTQAEDEADYYCGATDLSPWSIVFGGGTKLTVL

GQPKAAPSVTLFPPSSEELQANKATLVCLI SDFYPGAVTVAWKADSSPVKAGVE TTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

14 DNA Light GATATCGAGCTGACTCAGCCCCCTAGCGTCAGCGTCAGCCCTGGCGAGACAGCC

Chain TCTATCACCTGTAGCGGCGATAAGCTGGGCGACTACTACGTGCACTGGTATCAG

CAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTATCAGGACTCTAAGCGGCCT AGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCGCTACCCTG ACTATTAGCGGCACTCAGGCCGAGGACGAGGCCGACTACTACTGCGGCGCTACC GACCTGAGCCCCTGGTCTATCGTGTTCGGCGGAGGCACTAAGCTGACCGTGCTG GGTCAGCCTAAGGCTGCCCCCAGCGTGACCCTGTTCCCCCCCAGCAGCGAGGAG CTGCAGGCCAACAAGGCCACCCTGGTGTGCCTGATCAGCGACTTCTACCCAGGC GCCGTGACCGTGGCCTGGAAGGCCGACAGCAGCCCCGTGAAGGCCGGCGTGGAG ACCACCACCCCCAGCAAGCAGAGCAACAACAAGTACGCCGCCAGCAGCTACCTG AGCCTGACCCCCGAGCAGTGGAAGAGCCACAGGTCCTACAGCTGCCAGGTGACC CACGAGGGCAGCACCGTGGAAAAGACCGTGGCCCCAACCGAGTGCAGC

NOV0308

15 HCDR1 SYAIS

(Kabat)

16 HCDR2 GI I PVLGTANYAQKFQG

(Kabat)

17 HCDR3 VPTDYFDY

(Kabat)

18 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPGQGLEWMGGIIP

VLGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARVPTDYFDY WGQGTLVTVSS

19 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTCTCTAGCTACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GTGCTGGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGTGCCTACCGACTACTTCGACTAC TGGGGTCAGGGCACCCTGGTCACCGTGTCTAGC

20 Heavy QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPGQGLEWMGGIIP

Chain VLGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARVPTDYFDY

WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN SGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVD KRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWD VSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKE YKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFS CSVMHEALHNHYTQKSLSLSPGK 21 DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTCTCTAGCTACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GTGCTGGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGTGCCTACCGACTACTTCGACTAC TGGGGTCAGGGCACCCTGGTCACCGTGTCTAGCGCTAGCACTAAGGGCCCAAG TGTGTTTCCCCTGGCCCCCAGCAGCAAGTCTACTTCCGGCGGAACTGCTGCCC TGGGTTGCCTGGTGAAGGACTACTTCCCCGAGCCCGTGACAGTGTCCTGGAAC TCTGGGGCTCTGACTTCCGGCGTGCACACCTTCCCCGCCGTGCTGCAGAGCAG CGGCCTGTACAGCCTGAGCAGCGTGGTGACAGTGCCCTCCAGCTCTCTGGGAA CCCAGACCTATATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGAC AAGAGAGTGGAGCCCAAGAGCTGCGACAAGACCCACACCTGCCCCCCCTGCCC AGCTCCAGAACTGCTGGGAGGGCCTTCCGTGTTCCTGTTCCCCCCCAAGCCCA AGGACACCCTGATGATCAGCAGGACCCCCGAGGTGACCTGCGTGGTGGTGGAC GTGTCCCACGAGGACCCAGAGGTGAAGTTCAACTGGTACGTGGACGGCGTGGA GGTGCACAACGCCAAGACCAAGCCCAGAGAGGAGCAGTACAACAGCACCTACA GGGTGGTGTCCGTGCTGACCGTGCTGCACCAGGACTGGCTGAACGGCAAAGAA TACAAGTGCAAAGTCTCCAACAAGGCCCTGCCAGCCCCAATCGAAAAGACAAT CAGCAAGGCCAAGGGCCAGCCACGGGAGCCCCAGGTGTACACCCTGCCCCCCA GCCGGGAGGAGATGACCAAGAACCAGGTGTCCCTGACCTGTCTGGTGAAGGGC TTCTACCCCAGCGATATCGCCGTGGAGTGGGAGAGCAACGGCCAGCCCGAGAA CAACTACAAGACCACCCCCCCAGTGCTGGACAGCGACGGCAGCTTCTTCCTGT ACAGCAAGCTGACCGTGGACAAGTCCAGGTGGCAGCAGGGCAACGTGTTCAGC TGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAG CCTGAGCCCCGGCAAG

22 LCDR1 SGDNLGSKYVH

(Kabat)

23 LCDR2 DDNKRPS

(Kabat)

24 LCDR3 QSWTLGNWV

(Kabat)

25 VL DIELTQPPSVSVSPGQTASITCSGDNLGSKYVHWYQQKPGQAPVLVIYDDNKR

PSGIPERFSGSNSGNTATLTISGTQAEDEADYYCQSWTLGNWVFGGGTKLTVL

26 DNA VL GATATCGAGCTGACTCAGCCCCCTAGCGTCAGCGTCAGCCCTGGTCAGACCGC

CTCTATCACCTGTAGCGGCGATAACCTGGGCTCTAAATACGTGCACTGGTATC AGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGATAACAAGCGG CCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCGCTAC CCTGACTATTAGCGGCACTCAGGCCGAGGACGAGGCCGACTACTACTGTCAGT CCTGGACCCTGGGCAACTGGGTGTTCGGCGGAGGCACTAAGCTGACCGTGCTG

27 Light DIELTQPPSVSVSPGQTASITCSGDNLGSKYVHWYQQKPGQAPVLVIYDDNKR

Chain PSGIPERFSGSNSGNTATLTISGTQAEDEADYYCQSWTLGNWVFGGGTKLTVL

GQPKAAPSVTLFPPSSEELQANKATLVCLI SDFYPGAVTVAWKADSSPVKAGV ETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS

28 DNA Light GATATCGAGCTGACTCAGCCCCCTAGCGTCAGCGTCAGCCCTGGTCAGACCGC

Chain CTCTATCACCTGTAGCGGCGATAACCTGGGCTCTAAATACGTGCACTGGTATC

AGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGATAACAAGCGG CCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCGCTAC CCTGACTATTAGCGGCACTCAGGCCGAGGACGAGGCCGACTACTACTGTCAGT CCTGGACCCTGGGCAACTGGGTGTTCGGCGGAGGCACTAAGCTGACCGTGCTG GGTCAGCCTAAGGCTGCCCCCAGCGTGACCCTGTTCCCCCCCAGCAGCGAGGA GCTGCAGGCCAACAAGGCCACCCTGGTGTGCCTGATCAGCGACTTCTACCCAG GCGCCGTGACCGTGGCCTGGAAGGCCGACAGCAGCCCCGTGAAGGCCGGCGTG GAGACCACCACCCCCAGCAAGCAGAGCAACAACAAGTACGCCGCCAGCAGCTA CCTGAGCCTGACCCCCGAGCAGTGGAAGAGCCACAGGTCCTACAGCTGCCAGG TGACCCACGAGGGCAGCACCGTGGAAAAGACCGTGGCCCCAACCGAGTGCAGC

NOV0563

29 HCDR1 DNAIS

(Kabat)

30 HCDR2 GINPDFGWANYAQKFQG

(Kabat) HCDR3 DSSGMGY

(Kabat)

VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFSDNAISWVRQAPGQGLEWMGGINP DFGWANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARDSSGMGYW GQGTLVTVSS

DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGCGATAACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGGATTAACCCC GACTTCGGCTGGGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGGGACTCTAGCGGAATGGGCTACTGG GGTCAGGGCACCCTGGTCACCGTGTCTAGC

Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSDNAISWVRQAPGQGLEWMGGINP

DFGWANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARDSSGMGYW GQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNS GALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDK RVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDV SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKEY KCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGF YPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSC SVMHEALHNHYTQKSLSLSPGK

DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGCGATAACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGGATTAACCCC GACTTCGGCTGGGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGGGACTCTAGCGGAATGGGCTACTGG GGTCAGGGCACCCTGGTCACCGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGT GTTTCCCCTGGCCCCCAGCAGCAAGTCTACTTCCGGCGGAACTGCTGCCCTGG GTTGCCTGGTGAAGGACTACTTCCCCGAGCCCGTGACAGTGTCCTGGAACTCT GGGGCTCTGACTTCCGGCGTGCACACCTTCCCCGCCGTGCTGCAGAGCAGCGG CCTGTACAGCCTGAGCAGCGTGGTGACAGTGCCCTCCAGCTCTCTGGGAACCC AGACCTATATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAG AGAGTGGAGCCCAAGAGCTGCGACAAGACCCACACCTGCCCCCCCTGCCCAGC TCCAGAACTGCTGGGAGGGCCTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGG ACACCCTGATGATCAGCAGGACCCCCGAGGTGACCTGCGTGGTGGTGGACGTG TCCCACGAGGACCCAGAGGTGAAGTTCAACTGGTACGTGGACGGCGTGGAGGT GCACAACGCCAAGACCAAGCCCAGAGAGGAGCAGTACAACAGCACCTACAGGG TGGTGTCCGTGCTGACCGTGCTGCACCAGGACTGGCTGAACGGCAAAGAATAC AAGTGCAAAGTCTCCAACAAGGCCCTGCCAGCCCCAATCGAAAAGACAATCAG CAAGGCCAAGGGCCAGCCACGGGAGCCCCAGGTGTACACCCTGCCCCCCAGCC GGGAGGAGATGACCAAGAACCAGGTGTCCCTGACCTGTCTGGTGAAGGGCTTC TACCCCAGCGATATCGCCGTGGAGTGGGAGAGCAACGGCCAGCCCGAGAACAA CTACAAGACCACCCCCCCAGTGCTGGACAGCGACGGCAGCTTCTTCCTGTACA GCAAGCTGACCGTGGACAAGTCCAGGTGGCAGCAGGGCAACGTGTTCAGCTGC AGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCTGAGCCT GAGCCCCGGCAAG

LCDR1 RASQDISSYLN

(Kabat)

LCDR2 DASTLQS

(Kabat)

LCDR3 QQSGHWLSKT

(Kabat)

VL DIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYDAST LQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSGHWLSKTFGQGTKVE IK 40 DNA VL GATATTCAGATGACTCAGTCACCTAGTAGCCTGAGCGCTAGTGTGGGCGATAG

AGTGACTATCACCTGTAGAGCCTCTCAGGATATCTCTAGCTACCTGAACTGGT ATCAGCAGAAGCCCGGTAAAGCCCCTAAGCTGCTGATCTACGACGCCTCTACC CTGCAGTCAGGCGTGCCCTCTAGGTTTAGCGGTAGCGGTAGTGGCACCGACTT CACCCTGACTATCTCTAGCCTGCAGCCCGAGGACTTCGCTACCTACTACTGTC AGCAGTCAGGCCACTGGCTGTCTAAGACCTTCGGTCAGGGCACTAAGGTCGAG ATTAAG

41 Light Chain DIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYDAST

LQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSGHWLSKTFGQGTKVE IKRTVAAPSVFIFPPSDEQLKSGTASWCLLNNFYPREAKVQWKVDNALQSGN SQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNR GEC

42 DNA Light GATATTCAGATGACTCAGTCACCTAGTAGCCTGAGCGCTAGTGTGGGCGATAG

Chain AGTGACTATCACCTGTAGAGCCTCTCAGGATATCTCTAGCTACCTGAACTGGT

ATCAGCAGAAGCCCGGTAAAGCCCCTAAGCTGCTGATCTACGACGCCTCTACC CTGCAGTCAGGCGTGCCCTCTAGGTTTAGCGGTAGCGGTAGTGGCACCGACTT CACCCTGACTATCTCTAGCCTGCAGCCCGAGGACTTCGCTACCTACTACTGTC AGCAGTCAGGCCACTGGCTGTCTAAGACCTTCGGTCAGGGCACTAAGGTCGAG ATTAAGCGTACGGTGGCCGCTCCCAGCGTGTTCATCTTCCCCCCCAGCGACGA GCAGCTGAAGAGCGGCACCGCCAGCGTGGTGTGCCTGCTGAACAACTTCTACC CCCGGGAGGCCAAGGTGCAGTGGAAGGTGGACAACGCCCTGCAGAGCGGCAAC AGCCAGGAGAGCGTCACCGAGCAGGACAGCAAGGACTCCACCTACAGCCTGAG CAGCACCCTGACCCTGAGCAAGGCCGACTACGAGAAGCATAAGGTGTACGCCT GCGAGGTGACCCACCAGGGCCTGTCCAGCCCCGTGACCAAGAGCTTCAACAGG GGCGAGTGC

NOV1216

43 HCDR1 DYAIS

(Kabat)

44 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

45 HCDR3 EQDPEYGYGGYPYEAMDV

(Kabat)

46 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGII

PAFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEY GYGGYPYEAMDVWGQGTLVTVSS

47 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCG

TGAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAG CTGGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATC CCCGCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTA TCACCGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAG ATCAGAGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTAC GGCTACGGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCC TGGTCACCGTGTCTAGC

48 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGII

PAFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEY GYGGYPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLV KDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTY ICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDT LMI SRTPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRV VSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPS REEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCG Chain TGAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAG

CTGGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATC CCCGCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTA TCACCGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAG ATCAGAGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTAC GGCTACGGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCC TGGTCACCGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGTGTTTCCCCTGGC CCCCAGCAGCAAGTCTACTTCCGGCGGAACTGCTGCCCTGGGTTGCCTGGTG AAGGACTACTTCCCCGAGCCCGTGACAGTGTCCTGGAACTCTGGGGCTCTGA CTTCCGGCGTGCACACCTTCCCCGCCGTGCTGCAGAGCAGCGGCCTGTACAG CCTGAGCAGCGTGGTGACAGTGCCCTCCAGCTCTCTGGGAACCCAGACCTAT ATCTGCAACGTGAACCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTGG AGCCCAAGAGCTGCGACAAGACCCACACCTGCCCCCCCTGCCCAGCTCCAGA ACTGCTGGGAGGGCCTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGGACACC CTGATGATCAGCAGGACCCCCGAGGTGACCTGCGTGGTGGTGGACGTGTCCC ACGAGGACCCAGAGGTGAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCA CAACGCCAAGACCAAGCCCAGAGAGGAGCAGTACAACAGCACCTACAGGGTG GTGTCCGTGCTGACCGTGCTGCACCAGGACTGGCTGAACGGCAAAGAATACA AGTGCAAAGTCTCCAACAAGGCCCTGCCAGCCCCAATCGAAAAGACAATCAG CAAGGCCAAGGGCCAGCCACGGGAGCCCCAGGTGTACACCCTGCCCCCCAGC CGGGAGGAGATGACCAAGAACCAGGTGTCCCTGACCTGTCTGGTGAAGGGCT TCTACCCCAGCGATATCGCCGTGGAGTGGGAGAGCAACGGCCAGCCCGAGAA CAACTACAAGACCACCCCCCCAGTGCTGGACAGCGACGGCAGCTTCTTCCTG TACAGCAAGCTGACCGTGGACAAGTCCAGGTGGCAGCAGGGCAACGTGTTCA GCTGCAGCGTGATGCACGAGGCCCTGCACAACCACTACACCCAGAAGTCCCT GAGCCTGAGCCCCGGCAAG

LCDR1 SGDNIPQHSVH

(Kabat)

LCDR2 DDTERPS

(Kabat)

LCDR3 SSWDSSMDSW

(Kabat)

VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTE RPSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTK LTVL

DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCG CTAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTA TCAGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGACACCGAG CGGCCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCG CTACCCTGACTATCTCTAGGGCTCAGGCCGGCGACGAGGCCGACTACTACTG CTCTAGCTGGGATAGCTCTATGGATAGCGTGGTGTTCGGCGGAGGCACTAAG CTGACCGTGCTG

Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTE

RPSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTK LTVLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSP VKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTV APTECS

DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCG Chain CTAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTA

TCAGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGACACCGAG CGGCCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCG CTACCCTGACTATCTCTAGGGCTCAGGCCGGCGACGAGGCCGACTACTACTG CTCTAGCTGGGATAGCTCTATGGATAGCGTGGTGTTCGGCGGAGGCACTAAG CTGACCGTGCTGGGTCAGCCTAAGGCTGCCCCCAGCGTGACCCTGTTCCCCC CCAGCAGCGAGGAGCTGCAGGCCAACAAGGCCACCCTGGTGTGCCTGATCAG CGACTTCTACCCAGGCGCCGTGACCGTGGCCTGGAAGGCCGACAGCAGCCCC GTGAAGGCCGGCGTGGAGACCACCACCCCCAGCAAGCAGAGCAACAACAAGT ACGCCGCCAGCAGCTACCTGAGCCTGACCCCCGAGCAGTGGAAGAGCCACAG GTCCTACAGCTGCCAGGTGACCCACGAGGGCAGCACCGTGGAAAAGACCGTG GCCCCAACCGAGTGCAGC NOV1218

57 HCDR1 THGLH

(Kabat)

58 HCDR2 AISYDASETNYADSVKG

(Kabat)

59 HCDR3 ESIGGYFDY

(Kabat)

60 VH QVQLLESGGGLVQPGGSLRLSCAASGFTFPTHGLHWVRQAPGKGLEWVSAIS

YDASETNYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARESIGGY FDYWGQGTLVTVSS

61 DNA VH CAGGTGCAGCTGCTGGAATCAGGCGGCGGACTGGTGCAGCCTGGCGGTAGCC

TGAGACTGAGCTGCGCTGCTAGTGGCTTCACCTTCCCTACTCACGGCCTGCA CTGGGTCAGACAGGCCCCTGGTAAAGGCCTGGAGTGGGTCAGCGCTATTAGC TACGACGCTAGTGAAACTAACTACGCCGATAGCGTGAAGGGCCGGTTCACTA TCTCTAGGGATAACTCTAAGAACACCCTGTACCTGCAGATGAATAGCCTGAG AGCCGAGGACACCGCCGTCTACTACTGCGCTAGAGAGTCTATCGGCGGCTAC TTCGACTACTGGGGTCAGGGCACCCTGGTCACCGTGTCTAGC

62 Heavy Chain QVQLLESGGGLVQPGGSLRLSCAASGFTFPTHGLHWVRQAPGKGLEWVSAIS

YDASETNYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARESIGGY FDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVT VSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHKPS NTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV TCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKS RWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

63 DNA Heavy CAGGTGCAGCTGCTGGAATCAGGCGGCGGACTGGTGCAGCCTGGCGGTAGCC

Chain TGAGACTGAGCTGCGCTGCTAGTGGCTTCACCTTCCCTACTCACGGCCTGCA

CTGGGTCAGACAGGCCCCTGGTAAAGGCCTGGAGTGGGTCAGCGCTATTAGC TACGACGCTAGTGAAACTAACTACGCCGATAGCGTGAAGGGCCGGTTCACTA TCTCTAGGGATAACTCTAAGAACACCCTGTACCTGCAGATGAATAGCCTGAG AGCCGAGGACACCGCCGTCTACTACTGCGCTAGAGAGTCTATCGGCGGCTAC TTCGACTACTGGGGTCAGGGCACCCTGGTCACCGTGTCTAGCGCTAGCACTA AGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCAGCAAGTCTACTTCCGGCGG AACTGCTGCCCTGGGTTGCCTGGTGAAGGACTACTTCCCCGAGCCCGTGACA GTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGTGCACACCTTCCCCGCCG TGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCGTGGTGACAGTGCCCTC CAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTGAACCACAAGCCCAGC AACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTGCGACAAGACCCACA CCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGCCTTCCGTGTTCCT GTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGGACCCCCGAGGTG ACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGTGAAGTTCAACT GGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGCCCAGAGAGGA GCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTGCTGCACCAG GACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAAGGCCCTGC CAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCACGGGAGCC CCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAACCAGGTG TCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGTGGAGT GGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAGTGCT GGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCC AGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGC ACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

64 LCDR1 SGDALGKNTVS

(Kabat)

65 LCDR2 DDTDRPS

(Kabat)

66 LCDR3 SSTDLSTW

(Kabat)

67 VL SYELTQPLSVSVALGQTARITCSGDALGKNTVSWYQQKPGQAPVLVIYDDTD

RPSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSTDLSTWFGGGTKLT VL 68 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCG

CTAGAATCACCTGTAGCGGCGACGCCCTGGGTAAAAACACCGTCAGCTGGTA TCAGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGACACCGAT AGACCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCG CTACCCTGACTATCTCTAGGGCTCAGGCCGGCGACGAGGCCGACTACTACTG CTCTAGCACCGACCTGAGCACCGTGGTGTTCGGCGGAGGCACTAAGCTGACC GTGCTG

69 Light Chain SYELTQPLSVSVALGQTARITCSGDALGKNTVSWYQQKPGQAPVLVIYDDTD

RPSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSTDLSTWFGGGTKLT VLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVK AGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAP TECS

70 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCG

Chain CTAGAATCACCTGTAGCGGCGACGCCCTGGGTAAAAACACCGTCAGCTGGTA

TCAGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGACACCGAT AGACCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCG CTACCCTGACTATCTCTAGGGCTCAGGCCGGCGACGAGGCCGACTACTACTG CTCTAGCACCGACCTGAGCACCGTGGTGTTCGGCGGAGGCACTAAGCTGACC GTGCTGGGTCAGCCTAAGGCTGCCCCCAGCGTGACCCTGTTCCCCCCCAGCA GCGAGGAGCTGCAGGCCAACAAGGCCACCCTGGTGTGCCTGATCAGCGACTT CTACCCAGGCGCCGTGACCGTGGCCTGGAAGGCCGACAGCAGCCCCGTGAAG GCCGGCGTGGAGACCACCACCCCCAGCAAGCAGAGCAACAACAAGTACGCCG CCAGCAGCTACCTGAGCCTGACCCCCGAGCAGTGGAAGAGCCACAGGTCCTA CAGCTGCCAGGTGACCCACGAGGGCAGCACCGTGGAAAAGACCGTGGCCCCA ACCGAGTGCAGC

NOV1219

71 HCDR1 THGLH

(Kabat)

72 HCDR2 AISYEGSETNYADSVKG

(Kabat)

73 HCDR3 ESIGGYFDY

(Kabat)

74 VH QVQLLESGGGLVQPGGSLRLSCAASGFTFPTHGLHWVRQAPGKGLEWVSAIS

YEGSETNYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARESIGGY FDYWGQGTLVTVSS

75 DNA VH CAGGTGCAGCTGCTGGAATCAGGCGGCGGACTGGTGCAGCCTGGCGGTAGCC

TGAGACTGAGCTGCGCTGCTAGTGGCTTCACCTTCCCTACTCACGGCCTGCA CTGGGTCAGACAGGCCCCTGGTAAAGGCCTGGAGTGGGTCAGCGCTATTAGC TACGAGGGTAGCGAGACTAACTACGCCGATAGCGTGAAGGGCCGGTTCACTA TCTCTAGGGATAACTCTAAGAACACCCTGTACCTGCAGATGAATAGCCTGAG AGCCGAGGACACCGCCGTCTACTACTGCGCTAGAGAGTCTATCGGCGGCTAC TTCGACTACTGGGGTCAGGGCACCCTGGTCACCGTGTCTAGC

76 Heavy Chain QVQLLESGGGLVQPGGSLRLSCAASGFTFPTHGLHWVRQAPGKGLEWVSAIS

YEGSETNYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARESIGGY FDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVT VSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHKPS NTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV TCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKS RWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 77 DNA Heavy CAGGTGCAGCTGCTGGAATCAGGCGGCGGACTGGTGCAGCCTGGCGGTAGCC Chain TGAGACTGAGCTGCGCTGCTAGTGGCTTCACCTTCCCTACTCACGGCCTGCA

CTGGGTCAGACAGGCCCCTGGTAAAGGCCTGGAGTGGGTCAGCGCTATTAGC TACGAGGGTAGCGAGACTAACTACGCCGATAGCGTGAAGGGCCGGTTCACTA TCTCTAGGGATAACTCTAAGAACACCCTGTACCTGCAGATGAATAGCCTGAG AGCCGAGGACACCGCCGTCTACTACTGCGCTAGAGAGTCTATCGGCGGCTAC TTCGACTACTGGGGTCAGGGCACCCTGGTCACCGTGTCTAGCGCTAGCACTA AGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCAGCAAGTCTACTTCCGGCGG AACTGCTGCCCTGGGTTGCCTGGTGAAGGACTACTTCCCCGAGCCCGTGACA GTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGTGCACACCTTCCCCGCCG TGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCGTGGTGACAGTGCCCTC CAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTGAACCACAAGCCCAGC AACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTGCGACAAGACCCACA CCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGCCTTCCGTGTTCCT GTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGGACCCCCGAGGTG ACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGTGAAGTTCAACT GGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGCCCAGAGAGGA GCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTGCTGCACCAG GACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAAGGCCCTGC CAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCACGGGAGCC CCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAACCAGGTG TCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGTGGAGT GGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAGTGCT GGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAGTCC AGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCTGC ACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

78 LCDR1 SGDALGKNTVS

(Kabat)

79 LCDR2 DDTDRPS

(Kabat)

80 LCDR3 SSTDLSTW

(Kabat)

81 VL SYELTQPLSVSVALGQTARITCSGDALGKNTVSWYQQKPGQAPVLVIYDDTD

RPSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSTDLSTWFGGGTKLT VL

82 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCG

83 Light Chain SYELTQPLSVSVALGQTARITCSGDALGKNTVSWYQQKPGQAPVLVIYDDTD

84 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCG

Chain CTAGAATCACCTGTAGCGGCGACGCCCTGGGTAAAAACACCGTCAGCTGGTA

NOV2106 85 HCDR1 DYAIS

(Kabat)

86 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

87 HCDR3 EQDPEFGYGGYPYEAMDV

(Kabat)

88 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEFGY GGYPYEAMDVWGQGTLVTVSS

89 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTTCGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGC

90 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEFGY GGYPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

91 DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTTCGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCA GCAAGTCTACTTCCGGCGGAACTGCTGCCCTGGGTTGCCTGGTGAAGGACTAC TTCCCCGAGCCCGTGACAGTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGT GCACACCTTCCCCGCCGTGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCG TGGTGACAGTGCCCTCCAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTG AACCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTG CGACAAGACCCACACCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGC CTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGG ACCCCCGAGGTGACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGT GAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGC CCAGAGAGGAGCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTG CTGCACCAGGACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAA GGCCCTGCCAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCAC GGGAGCCCCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAAC CAGGTGTCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGT GGAGTGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAG TGCTGGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAG TCCAGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCT GCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

92 LCDR1 SGDNIPQHSVH

(Kabat)

93 LCDR2 DDTERPS

(Kabat)

94 LCDR3 SSWDSSMDSW

(Kabat)

95 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL 96 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

TAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTATC AGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGACACCGAGCGG CCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCGCTAC CCTGACTATCTCTAGGGCTCAGGCCGGCGACGAGGCCGACTACTACTGCTCTA GCTGGGATAGCTCTATGGATAGCGTGGTGTTCGGCGGAGGCACTAAGCTGACC GTGCTG

97 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKA GVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTE CS

98 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

Chain TAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTATC

AGCAGAAGCCCGGTCAGGCCCCCGTGCTGGTGATCTACGACGACACCGAGCGG CCTAGCGGAATCCCCGAGCGGTTTAGCGGCTCTAATAGCGGTAACACCGCTAC CCTGACTATCTCTAGGGCTCAGGCCGGCGACGAGGCCGACTACTACTGCTCTA GCTGGGATAGCTCTATGGATAGCGTGGTGTTCGGCGGAGGCACTAAGCTGACC GTGCTGGGTCAGCCTAAGGCTGCCCCCAGCGTGACCCTGTTCCCCCCCAGCAG CGAGGAGCTGCAGGCCAACAAGGCCACCCTGGTGTGCCTGATCAGCGACTTCT ACCCAGGCGCCGTGACCGTGGCCTGGAAGGCCGACAGCAGCCCCGTGAAGGCC GGCGTGGAGACCACCACCCCCAGCAAGCAGAGCAACAACAAGTACGCCGCCAG CAGCTACCTGAGCCTGACCCCCGAGCAGTGGAAGAGCCACAGGTCCTACAGCT GCCAGGTGACCCACGAGGGCAGCACCGTGGAAAAGACCGTGGCCCCAACCGAG TGCAGC

NOV2107

99 HCDR1 DYAIS

(Kabat)

100 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

101 HCDR3 EQDPEAGYGGYPYEAMDV

(Kabat)

102 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEAGY GGYPYEAMDVWGQGTLVTVSS

103 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGGCCGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGC

104 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEAGY GGYPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 105 DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGGCCGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCA GCAAGTCTACTTCCGGCGGAACTGCTGCCCTGGGTTGCCTGGTGAAGGACTAC TTCCCCGAGCCCGTGACAGTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGT GCACACCTTCCCCGCCGTGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCG TGGTGACAGTGCCCTCCAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTG AACCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTG CGACAAGACCCACACCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGC CTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGG ACCCCCGAGGTGACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGT GAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGC CCAGAGAGGAGCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTG CTGCACCAGGACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAA GGCCCTGCCAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCAC GGGAGCCCCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAAC CAGGTGTCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGT GGAGTGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAG TGCTGGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAG TCCAGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCT GCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

106 LCDR1 SGDNIPQHSVH

(Kabat)

107 LCDR2 DDTERPS

(Kabat)

108 LCDR3 SSWDSSMDSW

(Kabat)

109 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL

110 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

111 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

112 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

Chain TAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTATC

NOV2108 113 HCDR1 DYAIS

(Kabat)

114 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

115 HCDR3 EQDPESGYGGYPYEAMDV

(Kabat)

116 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPESGY GGYPYEAMDVWGQGTLVTVSS

117 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTCCGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGC

118 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPESGY GGYPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

119 DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTCCGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCA GCAAGTCTACTTCCGGCGGAACTGCTGCCCTGGGTTGCCTGGTGAAGGACTAC TTCCCCGAGCCCGTGACAGTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGT GCACACCTTCCCCGCCGTGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCG TGGTGACAGTGCCCTCCAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTG AACCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTG CGACAAGACCCACACCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGC CTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGG ACCCCCGAGGTGACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGT GAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGC CCAGAGAGGAGCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTG CTGCACCAGGACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAA GGCCCTGCCAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCAC GGGAGCCCCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAAC CAGGTGTCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGT GGAGTGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAG TGCTGGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAG TCCAGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCT GCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

120 LCDR1 SGDNIPQHSVH

(Kabat)

121 LCDR2 DDTERPS

(Kabat)

122 LCDR3 SSWDSSMDSW

(Kabat)

123 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL 124 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

125 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

126 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

Chain TAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTATC

NOV2109

127 HCDR1 DYAIS

(Kabat)

128 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

129 HCDR3 EQDPEYGFGGYPYEAMDV

(Kabat)

130 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEYGF GGYPYEAMDVWGQGTLVTVSS

131 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTACGGCTTC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGC

132 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEYGF GGYPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 133 DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCGAGTACGGCTTC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCA GCAAGTCTACTTCCGGCGGAACTGCTGCCCTGGGTTGCCTGGTGAAGGACTAC TTCCCCGAGCCCGTGACAGTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGT GCACACCTTCCCCGCCGTGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCG TGGTGACAGTGCCCTCCAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTG AACCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTG CGACAAGACCCACACCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGC CTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGG ACCCCCGAGGTGACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGT GAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGC CCAGAGAGGAGCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTG CTGCACCAGGACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAA GGCCCTGCCAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCAC GGGAGCCCCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAAC CAGGTGTCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGT GGAGTGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAG TGCTGGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAG TCCAGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCT GCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

134 LCDR1 SGDNIPQHSVH

(Kabat)

135 LCDR2 DDTERPS

(Kabat)

136 LCDR3 SSWDSSMDSW

(Kabat)

137 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL

138 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

139 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

140 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

Chain TAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTATC

NOV2110_N297A 141 HCDR1 DYAIS

(Kabat)

142 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

143 HCDR3 EQDPEYGYGGFPYEAMDV

(Kabat)

144 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEYGY GGFPYEAMDVWGQGTLVTVSS

145 DNA VH CAGGTGCAATTGGTGCAGAGCGGTGCCGAAGTGAAAAAACCGGGCAGCAGCGT

GAAAGTTAGCTGCAAAGCATCCGGAGGGACGTTTCGTGACTACGCTATCTCTT GGGTGCGCCAGGCCCCGGGCCAGGGCCTCGAGTGGATGGGCGGTATCATCCCG GCTTTCGGCACTGCGAACTACGCCCAGAAATTTCAGGGCCGGGTGACCATTAC CGCCGATGAAAGCACCAGCACCGCCTATATGGAACTGAGCAGCCTGCGCAGCG AAGATACGGCCGTGTATTATTGCGCGCGTGAACAGGACCCGGAATACGGTTAC GGTGGTTTCCCGTATGAAGCTATGGATGTTTGGGGCCAAGGCACCCTGGTGAC TGTTAGCTCA

146 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEYGY GGFPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYASTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

147 DNA Heavy CAGGTGCAATTGGTGCAGAGCGGTGCCGAAGTGAAAAAACCGGGCAGCAGCGT

Chain GAAAGTTAGCTGCAAAGCATCCGGAGGGACGTTTCGTGACTACGCTATCTCTT

GGGTGCGCCAGGCCCCGGGCCAGGGCCTCGAGTGGATGGGCGGTATCATCCCG GCTTTCGGCACTGCGAACTACGCCCAGAAATTTCAGGGCCGGGTGACCATTAC CGCCGATGAAAGCACCAGCACCGCCTATATGGAACTGAGCAGCCTGCGCAGCG AAGATACGGCCGTGTATTATTGCGCGCGTGAACAGGACCCGGAATACGGTTAC GGTGGTTTCCCGTATGAAGCTATGGATGTTTGGGGCCAAGGCACCCTGGTGAC TGTTAGCTCAGCCTCCACCAAGGGTCCATCGGTCTTCCCCCTGGCACCCTCCT CCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTAC TTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGT GCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCG TGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTG AATCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTTGAGCCCAAATCTTG TGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGAC CGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGG ACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGT CAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGC CGCGGGAGGAGCAGTACGCCAGCACGTACCGGGTGGTCAGCGTCCTCACCGTC CTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAA AGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCC GAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAAC CAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGT GGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCG TGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAG AGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCT GCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA

148 LCDR1 SGDNIPQHSVH

(Kabat)

149 LCDR2 DDTERPS

(Kabat)

150 LCDR3 SSWDSSMDSW

(Kabat)

151 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL 152 DNA VL AGCTACGAACTGACCCAGCCGCTGAGCGTGAGCGTGGCCCTGGGCCAGACCGC

GAGGATTACCTGTAGCGGCGATAACATCCCGCAGCATTCTGTTCATTGGTACC AGCAGAAACCGGGCCAGGCGCCGGTGCTGGTGATCTACGACGACACTGAACGT CCGAGCGGCATCCCGGAACGTTTTAGCGGATCCAACAGCGGCAACACCGCGAC CCTGACCATTAGCAGGGCCCAGGCGGGCGACGAAGCGGATTATTACTGCTCTT CTTGGGACTCTTCTATGGACTCTGTTGTGTTTGGCGGCGGCACGAAGTTAACC GTCCTA

153 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

154 DNA Light AGCTACGAACTGACCCAGCCGCTGAGCGTGAGCGTGGCCCTGGGCCAGACCGC

Chain GAGGATTACCTGTAGCGGCGATAACATCCCGCAGCATTCTGTTCATTGGTACC

AGCAGAAACCGGGCCAGGCGCCGGTGCTGGTGATCTACGACGACACTGAACGT CCGAGCGGCATCCCGGAACGTTTTAGCGGATCCAACAGCGGCAACACCGCGAC CCTGACCATTAGCAGGGCCCAGGCGGGCGACGAAGCGGATTATTACTGCTCTT CTTGGGACTCTTCTATGGACTCTGTTGTGTTTGGCGGCGGCACGAAGTTAACC GTCCTAGGTCAGCCCAAGGCTGCCCCCTCGGTCACTCTGTTCCCGCCCTCCTC TGAGGAGCTTCAAGCCAACAAGGCCACACTGGTGTGTCTCATAAGTGACTTCT ACCCGGGAGCCGTGACAGTGGCCTGGAAGGCAGATAGCAGCCCCGTCAAGGCG GGAGTGGAGACCACCACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAG CAGCTATCTGAGCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCT GCCAGGTCACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAA TGTTCA

NOV2111 _N297A

155 HCDR1 DYAIS

(Kabat)

156 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

157 HCDR3 EQDPEYGYGGYPFEAMDV

(Kabat)

158 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEYGY GGYPFEAMDVWGQGTLVTVSS

159 DNA VH CAGGTGCAATTGGTGCAGAGCGGTGCCGAAGTGAAAAAACCGGGCAGCAGCGT

GAAAGTTAGCTGCAAAGCATCCGGAGGGACGTTTCGTGACTACGCTATCTCTT GGGTGCGCCAGGCCCCGGGCCAGGGCCTCGAGTGGATGGGCGGTATCATCCCG GCTTTCGGCACTGCGAACTACGCCCAGAAATTTCAGGGCCGGGTGACCATTAC CGCCGATGAAAGCACCAGCACCGCCTATATGGAACTGAGCAGCCTGCGCAGCG AAGATACGGCCGTGTATTATTGCGCGCGTGAACAGGACCCGGAATACGGTTAC GGTGGTTACCCGTTCGAAGCTATGGATGTTTGGGGCCAAGGCACCCTGGTGAC TGTTAGCTCA

160 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPEYGY GGYPFEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYASTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 161 DNA Heavy CAGGTGCAATTGGTGCAGAGCGGTGCCGAAGTGAAAAAACCGGGCAGCAGCGT Chain GAAAGTTAGCTGCAAAGCATCCGGAGGGACGTTTCGTGACTACGCTATCTCTT

GGGTGCGCCAGGCCCCGGGCCAGGGCCTCGAGTGGATGGGCGGTATCATCCCG GCTTTCGGCACTGCGAACTACGCCCAGAAATTTCAGGGCCGGGTGACCATTAC CGCCGATGAAAGCACCAGCACCGCCTATATGGAACTGAGCAGCCTGCGCAGCG AAGATACGGCCGTGTATTATTGCGCGCGTGAACAGGACCCGGAATACGGTTAC GGTGGTTACCCGTTCGAAGCTATGGATGTTTGGGGCCAAGGCACCCTGGTGAC TGTTAGCTCAGCCTCCACCAAGGGTCCATCGGTCTTCCCCCTGGCACCCTCCT CCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTAC TTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGT GCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCG TGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTG AATCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTTGAGCCCAAATCTTG TGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGAC CGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGG ACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGT CAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGC CGCGGGAGGAGCAGTACGCCAGCACGTACCGGGTGGTCAGCGTCCTCACCGTC CTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAA AGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCC GAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAAC CAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGT GGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCG TGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAG AGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCT GCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA

162 LCDR1 SGDNIPQHSVH

(Kabat)

163 LCDR2 DDTERPS

(Kabat)

164 LCDR3 SSWDSSMDSW

(Kabat)

165 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL

166 DNA VL AGCTACGAACTGACCCAGCCGCTGAGCGTGAGCGTGGCCCTGGGCCAGACCGC

167 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

168 DNA Light AGCTACGAACTGACCCAGCCGCTGAGCGTGAGCGTGGCCCTGGGCCAGACCGC

Chain GAGGATTACCTGTAGCGGCGATAACATCCCGCAGCATTCTGTTCATTGGTACC

NOV2112 169 HCDR1 DYAIS

(Kabat)

170 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

171 HCDR3 EQDPSYGYGGYPYEAMDV

(Kabat)

172 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPSYGY GGYPYEAMDVWGQGTLVTVSS

173 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCTCCTACGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGC

174 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQDPSYGY GGYPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

175 DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGGACCCCTCCTACGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCA GCAAGTCTACTTCCGGCGGAACTGCTGCCCTGGGTTGCCTGGTGAAGGACTAC TTCCCCGAGCCCGTGACAGTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGT GCACACCTTCCCCGCCGTGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCG TGGTGACAGTGCCCTCCAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTG AACCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTG CGACAAGACCCACACCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGC CTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGG ACCCCCGAGGTGACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGT GAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGC CCAGAGAGGAGCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTG CTGCACCAGGACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAA GGCCCTGCCAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCAC GGGAGCCCCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAAC CAGGTGTCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGT GGAGTGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAG TGCTGGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAG TCCAGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCT GCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

176 LCDR1 SGDNIPQHSVH

(Kabat)

177 LCDR2 DDTERPS

(Kabat)

178 LCDR3 SSWDSSMDSW

(Kabat)

179 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL 180 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

181 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

182 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

Chain TAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTATC

NOV2113

183 HCDR1 DYAIS

(Kabat)

184 HCDR2 GI I PAFGTANYAQKFQG

(Kabat)

185 HCDR3 EQSPEYGYGGYPYEAMDV

(Kabat)

186 VH QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQSPEYGY GGYPYEAMDVWGQGTLVTVSS

187 DNA VH CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT

GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGTCCCCCGAGTACGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGC

188 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFRDYAISWVRQAPGQGLEWMGGIIP

AFGTANYAQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREQSPEYGY GGYPYEAMDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDY FPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNV NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISR TPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDK SRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 189 DNA Heavy CAGGTGCAGCTGGTGCAGTCAGGCGCCGAAGTGAAGAAACCCGGCTCTAGCGT Chain GAAAGTCAGCTGTAAAGCTAGTGGCGGCACCTTTAGAGACTACGCTATTAGCT

GGGTCAGACAGGCCCCAGGTCAGGGCCTGGAGTGGATGGGCGGAATTATCCCC GCCTTCGGCACCGCTAACTACGCTCAGAAATTTCAGGGTAGAGTGACTATCAC CGCCGACGAGTCTACTAGCACCGCCTATATGGAACTGTCTAGCCTGAGATCAG AGGACACCGCCGTCTACTACTGCGCTAGAGAGCAGTCCCCCGAGTACGGCTAC GGCGGCTACCCCTACGAGGCTATGGACGTGTGGGGTCAGGGCACCCTGGTCAC CGTGTCTAGCGCTAGCACTAAGGGCCCAAGTGTGTTTCCCCTGGCCCCCAGCA GCAAGTCTACTTCCGGCGGAACTGCTGCCCTGGGTTGCCTGGTGAAGGACTAC TTCCCCGAGCCCGTGACAGTGTCCTGGAACTCTGGGGCTCTGACTTCCGGCGT GCACACCTTCCCCGCCGTGCTGCAGAGCAGCGGCCTGTACAGCCTGAGCAGCG TGGTGACAGTGCCCTCCAGCTCTCTGGGAACCCAGACCTATATCTGCAACGTG AACCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTGGAGCCCAAGAGCTG CGACAAGACCCACACCTGCCCCCCCTGCCCAGCTCCAGAACTGCTGGGAGGGC CTTCCGTGTTCCTGTTCCCCCCCAAGCCCAAGGACACCCTGATGATCAGCAGG ACCCCCGAGGTGACCTGCGTGGTGGTGGACGTGTCCCACGAGGACCCAGAGGT GAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCACAACGCCAAGACCAAGC CCAGAGAGGAGCAGTACAACAGCACCTACAGGGTGGTGTCCGTGCTGACCGTG CTGCACCAGGACTGGCTGAACGGCAAAGAATACAAGTGCAAAGTCTCCAACAA GGCCCTGCCAGCCCCAATCGAAAAGACAATCAGCAAGGCCAAGGGCCAGCCAC GGGAGCCCCAGGTGTACACCCTGCCCCCCAGCCGGGAGGAGATGACCAAGAAC CAGGTGTCCCTGACCTGTCTGGTGAAGGGCTTCTACCCCAGCGATATCGCCGT GGAGTGGGAGAGCAACGGCCAGCCCGAGAACAACTACAAGACCACCCCCCCAG TGCTGGACAGCGACGGCAGCTTCTTCCTGTACAGCAAGCTGACCGTGGACAAG TCCAGGTGGCAGCAGGGCAACGTGTTCAGCTGCAGCGTGATGCACGAGGCCCT GCACAACCACTACACCCAGAAGTCCCTGAGCCTGAGCCCCGGCAAG

190 LCDR1 SGDNIPQHSVH

(Kabat)

191 LCDR2 DDTERPS

(Kabat)

192 LCDR3 SSWDSSMDSW

(Kabat)

193 VL SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

PSGIPERFSGSNSGNTATLTISRAQAGDEADYYCSSWDSSMDSWFGGGTKLT VL

194 DNA VL AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

195 Light Chain SYELTQPLSVSVALGQTARITCSGDNIPQHSVHWYQQKPGQAPVLVIYDDTER

196 DNA Light AGCTACGAGCTGACTCAGCCCCTGAGCGTCAGCGTGGCCCTGGGTCAGACCGC

Chain TAGAATCACCTGTAGCGGCGATAATATCCCTCAGCACTCAGTGCACTGGTATC

2B6 (WO2004/016750) 197 HCDR1 DYPFTNYWIH

(Kabat)

198 HCDR2 VIDPSDTYPNYNKKFKG

(Kabat)

199 HCDR3 NGDSDYYSGMDY

(Kabat)

200 VH QVQLQQPVTELVRPGASVMLSCKASDYPFTNYWIHWVKQRPGQGLEWIGVIDP

SDTYPNYNKKFKGKATLTVWSSSTAYMQLSSLTSDDSAVYYCARNGDSDYYS GMDYWGQGTSVTVSS

201 DNA VH CAGGTCCAATTGCAGCAGCCTGTGACTGAGCTGGTGAGGCCGGGGGCTTCAGT

GATGTTGTCCTGCAAGGCTTCTGACTACCCCTTCACCAACTACTGGATACACT GGGTAAAGCAGAGGCCTGGACAAGGCCTGGAGTGGATCGGAGTGATTGATCCT TCTGATACTTATCCAAATTACAATAAAAAGTTCAAGGGCAAGGCCACATTGAC TGTAGTCGTATCCTCCAGCACAGCCTACATGCAGCTCAGCAGCCTGACATCTG ACGATTCTGCGGTCTATTACTGTGCAAGAAACGGTGATTCCGATTATTACTCT GGTATGGACTACTGGGGTCAAGGAACCTCAGTCACCGTCTCCTCA

202 LCDR1 RTSQSIGTNIH

(Kabat)

203 LCDR2 NVSESIS

(Kabat)

204 LCDR3 QQSNTWPFT

(Kabat)

205 VL DILLTQSPAILSVSPGERVSFSCRTSQSIGTNIHWYQQRTNGFPRLLIKNVSE

SISGIPSRFSGSGSGTDFILSINSVESEDIADYYCQQSNTWPFTFGGGTKLEI

K

206 DNA VL GACATCTTGCTGACTCAGTCTCCAGCCATCCTGTCTGTGAGTCCAGGAGAGAG

AGTCAGTTTTTCCTGCAGGACCAGTCAGAGCATTGGCACAAACATACACTGGT ATCAGCAAAGAACAAATGGTTTTCCAAGGCTTCTCATAAAGAATGTTTCTGAG TCTATCTCTGGGATCCCTTCCAGGTTTAGTGGCAGTGGATCAGGGACAGATTT TATTCTTAGCATCAACAGTGTGGAGTCTGAAGATATTGCAGATTATTATTGTC AACAAAGTAATACCTGGCCGTTCACGTTCGGAGGGGGGACCAAGCTGGAAATA AAA

AB 20 (WO2009/083009)

207 HCDR1 SYGIS

(Kabat)

208 HCDR2 WI SAYNGNTKYAQKLQG

(Kabat)

209 HCDR3 DSAAHGMDV

(Kabat)

210 VH QVQLVQSGGEVKKPGASVKVSCKTSGYTFTSYGISWVRQAPGQGLEWMGWISA

YNGNTKYAQKLQGRLTMTTDTSTTTAYMELRSLRSDDTAVYYCARDSAAHGMD VWGQGTTVTVSS

211 LCDR1 RASQGISSWLA

(Kabat)

212 LCDR2 AASSLQS

(Kabat)

213 LCDR3 QQYNSYPYT

(Kabat)

214 VL DIQMTQSPSSLSASVGDRVTITCRASQGISSWLAWYQQKPEKAPKSLIYAASS

LQSGVPSRFRGSGSGTDFTLTISSLQPEDFATYYCQQYNSYPYTFGQGTKLEI KR

Ab 24 (WO2009/083009)

215 HCDR1 SYGLS

(Kabat)

216 HCDR2 WISPYNGNTHYAQKLQG

(Kabat)

217 HCDR3 DSAAHGMDV

(Kabat)

218 VH QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYGLSWVRQAPGQGLEWMGWISP

YNGNTHYAQKLQGRVTMTTDTSTSTAYMDLRSLRSDDTAVYYCARDSAAHGMD VWGQGTTVTVSS 219 LCDR1 RASQGISSWLA

(Kabat)

220 LCDR2 AASSLQS

(Kabat)

221 LCDR3 QQYNSYPYT

(Kabat)

222 VL DIQMTQSPSSLSASVGDRVTITCRASQGISSWLAWYQQKPEKAPKSLIYAASS

LQSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQYNSYPYTFGQGTKLEI KR

AB 26 (WO2009/083009)

223 HCDR1 SYGLS

(Kabat)

224 HCDR2 WI SAYNGNTNYAQKLQG

(Kabat)

225 HCDR3 DSAAHGMDV

(Kabat)

226 VH QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYGLSWVRQAPGQGLEWMGWISA

YNGNTNYAQKLQGRVTMTTDTSTSTAYMDLRSLRSDDTAVYYCARDSAAHGMD VWGQGTTVTVSS

227 LCDR1 RASQGISSWLA

(Kabat)

228 LCDR2 AASSLQS

(Kabat)

229 LCDR3 QQYNSYPYT

(Kabat)

230 VL DIQMTQSPSSLSASVGDRVTITCRASQGISSWLAWYQQKPEKAPKSLIYAASS

LQSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQYNSYPYTFGQGTKLEI KR

AB 28 (WO2009/083009)

231 HCDR1 SYGIS

(Kabat)

232 HCDR2 WI SAYNGNTKYAQKLQG

(Kabat)

233 HCDR3 DSAAHGMDV

(Kabat)

234 VH QVQWQSGAEVKKPGASVKVSCKTSGYTFTSYGISWVRQAPGQGLEWMGWISA

YNGNTKYAQKLQGRLTMTTDTSTTTAYMELRSLRSDDTAVYYCARDSAAHGMD VWGQGTTVSVSS

235 LCDR1 RASQGISSWLA

(Kabat)

236 LCDR2 AASSLQS

(Kabat)

237 LCDR3 QQYNSYPYT

(Kabat)

238 VL DIQMTQSPSSLSASVGDRVTITCRASQGISSWLAWYQQKPEKAPKSLIYAASS

LQSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQYNSYPYTFGQGTKLEI KR

6G11 (WO2012/022985)

239 HCDR1 SYGMH

(Kabat)

240 HCDR2 VI SYDGSNKYYADSVKG

(Kabat)

241 HCDR3 ELYDAFDI

(Kabat)

242 VH EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWMAVISY

DGSNKYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARELYDAFDI WGQGTLVTVSS 243 LCDR1 TGSSSNIGAGYDVH

(Kabat)

244 LCDR2 ADDHRPS

(Kabat)

245 LCDR3 ASWDDSQRAVI

(Kabat)

246 VL QSVLTQPPSASGTPGQRVTISCTGSSSNIGAGYDVHWYQQLPGTAPKLLIYAD

DHRPSGVPDRFSGSKSGTSASLAISGLRSEDEADYYCASWDDSQRAVIFGGGT KLTVLG

CD32b Related Sequences

247 H167 CD32a MTMETQMSQNVCPRNLWLLQPLTVLLLLASADSQAAAPPKAVLKLEPPWINVL

UniProtKB QEDSVTLTCQGARSPESDSIQWFHNGNLIPTHTQPSYRFKANNNDSGEYTCQT

P12318 GQTSLSDPVHLTVLSEWLVLQTPHLEFQEGETIMLRCHSWKDKPLVKVTFFQN

GKSQKFSHLDPTFSIPQANHSHSGDYHCTGNIGYTLFSSKPVTITVQVPSMGS SSPMGI IVAWIATAVAAIVAAWALIYCRKKRI SANSTDPVKAAQFEPPGRQ MIAIRKRQLEETNNDYETADGGYMTLNPRAPTDDDKNIYLTLPPNDH SNN

248 CD32b MGILSFLPVLATESDWADCKSPQPWGHMLLWTAVLFLAPVAGTPAAPPKAVLK variant 1 LEPQWINVLQEDSVTLTCRGTHSPESDSIQWFHNGNLIPTHTQPSYRFKANNN

UniProtKB DSGEYTCQTGQTSLSDPVHLTVLSEWLVLQTPHLEFQEGETIVLRCHSWKDKP

P31994-1 LVKVTFFQNGKSKKFSRSDPNFSIPQANHSHSGDYHCTGNIGYTLYSSKPVTI

TVQAPSSSPMGI IVAWTGIAVAAIVAAWALIYCRKKRI SALPGYPECREMG ETLPEKPANPTNPDEADKVGAENTITYSLLMHPDALEEPDDQNRI

249 CD32b MGILSFLPVLATESDWADCKSPQPWGHMLLW AVLFLAPVAGTP APPK variant 2 AVLKLEPQWINVLQEDSVTLTCRGTHSPESDSIQWFHNGNLIPTHTQPS

UniProtKB YRFKA NNDSGEYTCQTGQTSLSDPVHLTVLSEWLVLQTPHLEFQEGET P31994-2 IVLRCHSWKDKPLVKVTFFQNGKSKKFSRSDPNFSIPQA HSHSGDYHC

TGNIGYTLYSSKPVTITVQAPSSSPMGI IVAWTGIAVAAIVAAWALI YCRKKRISA PTNPDEADKVGAENT ITYSLLMHPDALEEPDDQNRI

250 NOV2108 VLRCHSWKDKPLVKVTF

binding

epitope

IL-15 & IL-15Ra Related Sequences

251 IL-15 (with MRISKPHLRSISIQCYLCLLLNSHFLTEAGIHVFILGCFSAGLPKTEAJV!VV signal ISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISLESGDAS peptide) aa IHDTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLQSFVHIVQMFI human NTS

252 Coding atgagaattt cgaaaccaca tttgagaagt atttccatcc

sequence of agtgctactt gtgtttactt ctaaacagtc attttctaac

immature/ tgaagctggc attcatgtct tcattttggg ctgtttcagt

precursor gcagggcttc ctaaaacaga agccaactgg gtgaatgtaa

form of taagtgattt gaaaaaaatt gaagatctta ttcaatctat

human IL-15 gcatattgat gctactttat atacggaaag tgatgttcac

including cccagttgca aagtaacagc aatgaagtgc tttctcttgg

signal agttacaagt tatttcactt gagtccggag atgcaagtat

peptide DNA tcatgataca gtagaaaatc tgatcatcct agcaaacaac

agtttgtctt ctaatgggaa tgtaacagaa tctggatgca

aagaatgtga ggaactggag gaaaaaaata ttaaagaatt

tttgcagagt tttgtacata ttgtccaaat gttcatcaac acttcttga IL-15 atgtggctcc agagcctgct actcctgggg acggtggcct

expression gcagcatctc gaactgggtg aacgtgatct cggacctgaa

construct gaagatcgag gacctcatcc agtcgatgca catcgacgcg

DNA (human acgctgtaca cggagtcgga cgtccacccg tcgtgcaagg

IL-15 with tcacggcgat gaagtgcttc ctcctggagc tccaagtcat

GMCSF ctcgctcgag tcgggggacg cgtcgatcca cgacacggtg

signal gagaacctga tcatcctggc gaacaactcg ctgtcgtcga

peptide ) acgggaacgt cacggagtcg

ggctgcaagg agtgcgagga gctggaggag aagaacatca

aggagttcct gcagtcgttc gtgcacatcg tccagatgtt

catcaacacg tcgtga

I IL-15 codon cctggccatt gcatacgttg tatccatatc ataatatgta

optimized catttatatt ggctcatgtc caacattacc gccatgttga

DNA cattgattat tgactagtta ttaatagtaa tcaattacgg

ggtcattagt tcatagccca tatatggagt tccgcgttac

ataacttacg gtaaatggcc cgcctggctg accgcccaac

gacccccgcc cattgacgtc aataatgacg tatgttccca

tagtaacgcc aatagggact ttccattgac gtcaatgggt

ggagtattta cggtaaactg cccacttggc agtacatcaa

gtgtatcata tgccaagtac gccccctatt gacgtcaatg

atggtaaatg gcccgcctgg cattatgccc agtacatgac

cttatgggac tttcctactt ggcagtacat ctacgtatta

gtcatcgcta ttaccatggt gatgcggttt tggcagtaca

tcaatgggcg tggatagcgg tttgactcac ggggatttcc

aagtctccac cccattgacg tcaatgggag tttgttttgg

caccaaaatc aacgggactt tccaaaatgt cgtaacaact

ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg

ggaggtctat ataagcagag ctcgtttagt gaaccgtcag

atcgcctgga gacgccatcc acgctgtttt gacctccata

gaagacaccg ggaccgatcc agcctccgcg ggcgcgcgtc

gacaagaaat gcggatctcg aagccgcacc tgcggtcgat

atcgatccag tgctacctgt gcctgctcct gaactcgcac

ttcctcacgg aggccggtat acacgtcttc atcctgggct

gcttctcggc ggggctgccg aagacggagg cgaactgggt

gaacgtgatc tcggacctga agaagatcga ggacctcatc

cagtcgatgc acatcgacgc gacgctgtac acggagtcgg

acgtccaccc gtcgtgcaag gtcacggcga tgaagtgctt

cctcctggag ctccaagtca tctcgctcga gtcgggggac

gcgtcgatcc acgacacggt ggagaacctg atcatcctgg

cgaacaactc gctgtcgtcg aacgggaacg tcacggagtc

gggctgcaag gagtgcgagg agctggagga gaagaacatc

aaggagttcc tgcagtcgtt cgtgcacatc gtccagatgt

tcatcaacac gtcgtgaggg cccggcgcgc cgaattcgcg

gatatcggtt aacggatcca gatctgctgt gccttctagt

tgccagccat ctgttgtttg cccctccccc gtgccttcct

tgaccctgga aggtgccact cccactgtcc tttcctaata

aaatgaggaa attgcatcgc attgtctgag taggtgtcat

tctattctgg ggggtggggt ggggcaggac agcaaggggg

aggattggga agacaatagc aggcatgctg gggatgcggt

gggctctatg ggtacccagg tgctgaagaa ttgacccggt

tcctcctggg ccagaaagaa gcaggcacat ccccttctct

gtgacacacc ctgtccacgc ccctggttct tagttccagc

cccactcata ggacactcat agctcaggag ggctccgcct

tcaatcccac ccgctaaagt acttggagcg gtctctccct

ccctcatcag cccaccaaac caaacctagc ctccaagagt

gggaagaaat taaagcaaga taggctatta agtgcagagg

gagagaaaat gcctccaaca tgtgaggaag taatgagaga aatcata

I IL-15 codon MRISKPHLRSISIQCYLCLLLNSHFLTEAGIHVFILGCFSAGLPKTEANW V optimized ISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISLESGDAS aa IHDTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLQSFVHIVQMFI

NTS Immature MAPRRARGCRTLGLPALLLLLLLRPPATRGJTCPPP SVEi¾.DJiVVXSYS.LYS

IL-15Ra RERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRDPALVHQRPAP

(full PSTVTTAGVTPQPESLSPSGKEPAASSPSSNNTAATTAAIVPGSQLMPSKSPS length TGTTEISSHESSHGTPSQTTAKNWELTASASHQPPGVYPQGHSDTTVAISTST human) aa VLLCGLSAVSLLACYLKSRQTPPLASVEMEAMEALPVTWGTSSRDEDLENCSH with signal HL

peptide

I Immature MAPRRARGCRTLGLPALLLLLLLRPPATRGJTCPPP SVEi¾.DJiVVXSYS.LYS IL-15Ra RERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRDPALVHQRPAP ( soluble PSTVTTAGVTPQPESLSPSGKEPAASSPSSNNTAATTAAIVPGSQLMPSKSPS human PQG TGTTEISSHESSHGTPSQTTAKNWELTASASHQPPGVYPQG

termination

with signal

peptide )

I Coding atggccccgc ggcgggcgcg cggctgccgg accctcggtc

sequence of tcccggcgct gctactgctg ctgctgctcc ggccgccggc

immature gacgcggggc atcacgtgcc ctcccccca t gtccgtggaa

form of cacgcagaca tctgggtcaa gagctacagc ttgtactcca

full length gggagcggta catttgtaac tctggtttca agegtaaage

human IL- cggcacgtcc agectgaegg agtgcgtgtt gaacaaggee

15Ra DNA acgaatgtcg cccactggac aacccccagt etcaaatgea

ttagagaccc tgccctggtt

caccaaaggc cagcgccacc ctccacagta aegaeggcag

gggtgacccc acagecagag agcctctccc cttctggaaa

agagcccgca gcttcatctc ccagctcaaa caacacagcg

gccacaacag cagctattgt cccgggctcc cagctga tgc

cttcaaaatc accttccaca ggaaccacag aga taagcag

tea tgagtcc tcccacggca ccccctctca gacaacagcc

aagaactggg aactcacagc atccgcctcc caccagccgc

caggtgtgta tccacagggc

cacagcgaca ccactgtggc tatctccacg tccactgtcc

tgctgtgtgg gctgageget gtgtctctcc tggca tgcta

cctcaagtca aggcaaactc ccccgctggc cagcgttgaa

a tggaageca tggaggctct geeggtgact tgggggacca

gcagcagaga tgaagacttg gaaaactget ctcaccacct

I Coding atggccccgc ggcgggcgcg cggctgccgg accctcggtc

sequence tcccggcgct gctactgctg ctgctgctcc ggccgccggc

immature gacgcggggc atcacgtgcc ctcccccca t gtccgtggaa

form of cacgcagaca tctgggtcaa gagctacagc ttgtactcca

soluble gggagcggta catttgtaac tctggtttca agegtaaage

human IL- cggcacgtcc agectgaegg agtgcgtgtt gaacaaggee

15Ra DNA acgaatgtcg cccactggac aacccccagt etcaaatgea

ttagagaccc tgccctggtt

caccaaaggc cagcgccacc ctccacagta aegaeggcag

gggtgacccc acagecagag agcctctccc cttctggaaa

agagcccgca gcttcatctc ccagctcaaa caacacagcg

gccacaacag cagctattgt cccgggctcc cagctga tgc

cttcaaaatc accttccaca ggaaccacag aga taagcag

tea tgagtcc tcccacggca ccccctctca gacaacagcc

aagaactggg aactcacagc atccgcctcc caccagccgc

caggtgtgta tccacagggc

I IL-15Ra w/o ITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNV signal AHWTTPSLKCIRDPALVHQRPAPPSTVTTAGVTPQPESLSPSGKEPAASSPSS peptide NNTAATTAAIVPGSQLMPSKSPSTGTTEISSHESSHGTPSQTTAKNWELTASA with PQG SHQPPGVYPQG

termination 261 IL-15Ra cctggccatt gcatacgttg tatccatatc ataatatgta codon catttatatt ggctcatgtc caacattacc gccatgttga optimized cattgattat tgactagtta ttaatagtaa tcaattacgg DNA ggtcattagt tcatagccca tatatggagt tccgcgttac

ataacttacg gtaaatggcc cgcctggctg accgcccaac

gacccccgcc cattgacgtc aataatgacg tatgttccca

tagtaacgcc aatagggact ttccattgac gtcaatgggt

ggagtattta cggtaaactg cccacttggc agtacatcaa

gtgtatcata tgccaagtac gccccctatt gacgtcaatg

atggtaaatg gcccgcctgg cattatgccc agtacatgac

cttatgggac tttcctactt ggcagtacat ctacgtatta

gtcatcgcta ttaccatggt gatgcggttt tggcagtaca

tcaatgggcg tggatagcgg tttgactcac ggggatttcc

aagtctccac cccattgacg tcaatgggag tttgttttgg

caccaaaatc aacgggactt tccaaaatgt cgtaacaact

ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg

ggaggtctat ataagcagag ctcgtttagt gaaccgtcag

atcgcctgga gacgccatcc acgctgtttt gacctccata

gaagacaccg ggaccgatcc agcctccgcg ggcgcgcgtc

gacgctagca agaaatggcc ccgaggcggg cgcgaggctg

ccggaccctc ggtctcccgg cgctgctact gctcctgctg

ctccggccgc cggcgacgcg gggcatcacg tgcccgcccc

ccatgtccgt ggagcacgca gacatctggg tcaagagcta

cagcttgtac tcccgggagc ggtacatctg caactcgggt

ttcaagcgga aggccggcac gtccagcctg acggagtgcg

tgttgaacaa ggccacgaat gtcgcccact ggacgacccc

ctcgctcaag tgcatccgcg acccggccct ggttcaccag

cggcccgcgc caccctccac cgtaacgacg gcgggggtga

ccccgcagcc ggagagcctc tccccgtcgg gaaaggagcc

cgccgcgtcg tcgcccagct cgaacaacac ggcggccaca

actgcagcga tcgtcccggg ctcccagctg atgccgtcga

agtcgccgtc cacgggaacc acggagatca gcagtcatga

gtcctcccac ggcaccccct cgcaaacgac ggccaagaac

tgggaactca cggcgtccgc ctcccaccag ccgccggggg

tgtatccgca aggccacagc gacaccacgg tggcgatctc

cacgtccacg gtcctgctgt gtgggctgag cgcggtgtcg

ctcctggcgt gctacctcaa gtcgaggcag actcccccgc

tggccagcgt tgagatggag gccatggagg ctctgccggt

gacgtggggg accagcagca gggatgagga cttggagaac

tgctcgcacc acctataatg agaattcgat ccagatctgc

tgtgccttct agttgccagc catctgttgt ttgcccctcc

cccgtgcctt ccttgaccct ggaaggtgcc actcccactg

tcctttccta ataaaatgag gaaattgcat cgcattgtct

gagtaggtgt cattctattc tggggggtgg ggtggggcag

gacagcaagg gggaggattg ggaagacaat agcaggcatg

ctggggatgc ggtgggctct atgggtaccc aggtgctgaa

gaattgaccc ggttcctcct gggccagaaa gaagcaggca

catccccttc tctgtgacac accctgtcca cgcccctggt

tcttagttcc agccccactc ataggacact catagctcag

gagggctccg ccttcaatcc cacccgctaa agtacttgga

gcggtctctc cctccctcat cagcccacca aaccaaacct

agcctccaag agtgggaaga aattaaagca agataggcta

ttaagtgcag agggagagaa aatgcctcca acatgtgagg

aagtaatgag agaaatcata

262 IL-15Ra MAPRRARGCRTLGLPALLLLLLLRPPATRGITCPPPMSVEHADIWVKSYSLYS codon RERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRDPALVHQRPAP optimized PSTVTTAGVTPQPESLSPSGKEPAASSPSSNNTAATTAAIVPGSQLMPSKSPS aa TGTTEISSHESSHGTPSQTTAKNWELTASASHQPPGVYPQGHSDTTVAISTST

VLLCGLSAVSLLACYLKSRQTPPLASVEMEAMEALPVTWGTSSRDEDLENCSH HL 263 IL-15Ra cctggccatt gcatacgttg tatccatatc ataatatgta codon catttatatt ggctcatgtc caacattacc gccatgttga optimized cattgattat tgactagtta ttaatagtaa tcaattacgg DNA ggtcattagt tcatagccca tatatggagt tccgcgttac

ataacttacg gtaaatggcc cgcctggctg accgcccaac

gacccccgcc cattgacgtc aataatgacg tatgttccca

tagtaacgcc aatagggact ttccattgac gtcaatgggt

ggagtattta cggtaaactg cccacttggc agtacatcaa

gtgtatcata tgccaagtac gccccctatt gacgtcaatg

atggtaaatg gcccgcctgg cattatgccc agtacatgac

cttatgggac tttcctactt ggcagtacat ctacgtatta

gtcatcgcta ttaccatggt gatgcggttt tggcagtaca

tcaatgggcg tggatagcgg tttgactcac ggggatttcc

aagtctccac cccattgacg tcaatgggag tttgttttgg

caccaaaatc aacgggactt tccaaaatgt cgtaacaact

ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg

ggaggtctat ataagcagag ctcgtttagt gaaccgtcag

atcgcctgga gacgccatcc acgctgtttt gacctccata

gaagacaccg ggaccgatcc agcctccgcg ggcgcgcgtc

gacgctagca agaaatggcc ccgaggcggg cgcgaggctg

ccggaccctc ggtctcccgg cgctgctact gctcctgctg

ctccggccgc cggcgacgcg gggcatcacg tgcccgcccc

ccatgtccgt ggagcacgca gacatctggg tcaagagcta

cagcttgtac tcccgggagc ggtacatctg caactcgggt

ttcaagcgga aggccggcac gtccagcctg acggagtgcg

tgttgaacaa ggccacgaat gtcgcccact ggacgacccc

ctcgctcaag tgcatccgcg acccggccct ggttcaccag

cggcccgcgc caccctccac cgtaacgacg gcgggggtga

ccccgcagcc ggagagcctc tccccgtcgg gaaaggagcc

cgccgcgtcg tcgcccagct cgaacaacac ggcggccaca

actgcagcga tcgtcccggg ctcccagctg atgccgtcga

agtcgccgtc cacgggaacc acggagatca gcagtcatga

gtcctcccac ggcaccccct cgcaaacgac ggccaagaac

tgggaactca cggcgtccgc ctcccaccag ccgccggggg

tgtatccgca aggccacagc gacaccacgt aatgagaatt

cgcggatatc ggttaacgga tccagatctg ctgtgccttc

tagttgccag ccatctgttg tttgcccctc ccccgtgcct

tccttgaccc tggaaggtgc cactcccact gtcctttcct

aataaaatga ggaaattgca tcgcattgtc tgagtaggtg

tcattctatt ctggggggtg gggtggggca ggacagcaag

ggggaggatt gggaagacaa tagcaggcat gctggggatg

cggtgggctc tatgggtacc caggtgctga agaattgacc

cggttcctcc tgggccagaa agaagcaggc acatcccctt

ctctgtgaca caccctgtcc acgcccctgg ttcttagttc

cagccccact cataggacac tcatagctca ggagggctcc

gccttcaatc ccacccgcta aagtacttgg agcggtctct

ccctccctca tcagcccacc aaaccaaacc tagcctccaa

gagtgggaag aaattaaagc aagataggct attaagtgca

gagggagaga aaatgcctcc aacatgtgag gaagtaatga

gagaaatcat a

264 IL-15Ra MAPRRARGCRTLGLPALLLLLLLRPPATRGITCPPPMSVEHADIWVKSYSLYS codon RERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRDPALVHQRPAP optimized PSTVTTAGVTPQPESLSPSGKEPAASSPSSNNTAATTAAIVPGSQLMPSKSPS aa TGTTEISSHESSHGTPSQTTAKNWELTASASHQPPGVYPQGHSDTT

265 C-terminal PQGHSDTT

end of the

soluble

form of

human IL- 15Ra 266 C-terminal PQGHSDT

end of the

soluble

form of

human IL- 15Ra

267 C-terminal PQGHSD

end of the

soluble

form of

human IL- 15Ra

268 C-terminal PQGHS

end of the

soluble

form of

human IL- 15Ra

269 C-terminal PQGH

end of the

soluble

form of

human IL- 15Ra

270 C-terminal PQG

end of the

soluble

form of

human IL- 15Ra

271 Soluble ITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNV form of AHWTTPSLKCIRDPALVHQRPAPPSTVTTAGVTPQPESLSPSGKEPAASSPSS human IL- NNTAATTAAIVPGSQLMPSKSPSTGTTEISSHESSHGTPSQTTAKNWELTASA 15Ra SHQPPGVYPQGHSDTT

272 IL-15Ra 0- NWELTASASHQPPGVYPQG

glycosylati

on on Thr5

273 IL-15Ra N- ITCPPPMSVEHADIWVK

glycosylati

on on Ser 8

274 IL-15Ra N- ITCPPPMSVEHADIWVKSYSLYSRERYICNS

glycosylati

on on Ser

18, 20, 23

or 31

275 IL-15Ra RXXR

heterologou

s protease

cleavage

site

recognized

by furin

protease

Arg-X-X-Arg

Xaa = any

amino acid Il-15Ra XXPRXX

heterologou

s protease

cleavage

site A-B- Pro-Arg-X-Y

1,2 Xaa =

hydrophobic

amino acids

5 , 6 Xaa =

nonacidic

amino acids

IL-15RaFc MAPRRARGCRTLGLPALLLLLLLRPPATRGITCPPPMSVEHADIWVKSYSLYS fusion RERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRDPALVHQRPAP protein PSTVTTAGVTPQPESLSPSGKEPAASSPSSNNTAATTAAIVPGSQLMPSKSPS

TGTTEISSHESSHGTPSQTTAKNWELTASASHQPPGVYPQGHSDTTPKSCDKT HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSHEDPEVKFN WYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALP APIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWE SNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNH YTQKSLSLSPGK

TGTTEISSHESSHGTPSQTTAKNWELTASASHQPPGVYPQGPKSCDKTHTCPP CPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSHEDPEVKFNWYVDG VEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQP ENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKS LSLSPGK

Claims

1. A combination comprising:

i) an anti- human CD32b antibody molecule; and

ii) an interleukin-15 (IL-15)/IL-15 receptor alpha (IL-15Ra) complex.

2. The combination of claim 1, wherein the anti-human CD32b antibody molecule comprises heavy and light chain variable domain CDRs comprising:

a. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 1, 2, and 3, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 8, 9, and 10, respectively;

b. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 15, 16, and 17, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 22, 23, and 24, respectively;

c. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 29, 30, and 31 respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 36, 37, 38, respectively;

d. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 43, 44, and 45, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 50, 51, 52, respectively;

e. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 57, 58, and 59, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 64, 65, and 66, respectively;

f. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 71, 72, and 73, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 78, 79, and 80, respectively;

g. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 85, 86, and 87, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 92, 93, and 94, respectively;

h. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 99, 100, and 101, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 106, 107, and 108, respectively;

i. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 1 13, 114, and 115, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 120, 121, and 122, respectively;

j . the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 127, 128, and 129, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 134, 135, and 136, respectively;

k. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 141, 142, and 143, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 148, 149, and 150, respectively;

1. the HCDRl, HCDR2, and HCDR3 sequences of SEQ ID NOs: 155, 156, and 157, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 162, 163, and 164, respectively; m. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 169, 170, and 171, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 176, 177, and 178, respectively;

n. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 183, 184, and 185, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 190, 191, and 192, respectively;

o. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 197, 198, and 199, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 202, 203, and 204, respectively;

p. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 207, 208, and 209, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 211, 212, and 213, respectively;

q. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 215, 216, and 217, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 219, 220, and 221, respectively;

r. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 223, 224, and 225, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 227, 228, and 229, respectively;

s. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 231, 232, and 233, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 235, 236, and 237, respectively; or

t. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 239, 240, and 241, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 243, 244, and 245, respectively;

wherein the CDRs are numbered according to Kabat numbering.

The combination of claim 1 or claim 2, wherein the IL-15/IL-15Ra complex is a heterodimeric complex of human IL-15 and human soluble IL-15Ra.

The combination of claim 3, wherein the human IL-15 comprises residues 49 to 162 of the amino acid sequence of SEQ ID NO: 251 and the human soluble IL-15Ra comprises the amino acid sequence of SEQ ID NO: 260.

The combination of any one of the preceding claims, wherein the anti-CD32b antibody molecule comprises a heavy chain variable domain (VH) comprising an amino acid sequence at least 90% identical to any of SEQ ID NO: 4, 18, 32, 46, 60, 74, 88, 102, 116, 130, 144, 158, 172, 186, 200, 210, 218, 226, 234 or 242.

The combination of any one of the preceding claims, wherein the anti-CD32b antibody molecule comprises a light chain variable domain (VL) comprising an amino acid sequence at least 90% identical to any of SEQ ID NO: 11, 25, 39, 53, 67, 81, 95, 109, 123, 137, 151, 165, 179, 193, 205, 214, 222, 230, 238 or 246.

The combination of any one of the preceding claims, wherein the anti-CD32b antibody molecule comprises a VH and a VL amino acid sequence comprising:

a. the VH amino acid sequence of SEQ ID NO: 4 and the VL amino acid sequence of SEQ ID NO:

11;

b. the VH amino acid sequence of SEQ ID NO: 18 and the VL amino acid sequence of SEQ ID NO: 25;

c. the VH amino acid sequence of SEQ ID NO: 32 and the VL amino acid sequence of SEQ ID NO: 39;

d. the VH amino acid sequence of SEQ ID NO: 46 and the VL amino acid sequence of SEQ ID NO: 53;

e. the VH amino acid sequence of SEQ ID NO: 60 and the VL amino acid sequence of SEQ ID NO: 67;

f. the VH amino acid sequence of SEQ ID NO: 74 and the VL amino acid sequence of SEQ ID NO: 81;

g. the VH amino acid sequence of SEQ ID NO: 88 and the VL amino acid sequence of SEQ ID NO: 95;

h. the VH amino acid sequence of SEQ ID NO: 102 and the VL amino acid sequence of SEQ ID NO: 109;

i. the VH amino acid sequence of SEQ ID NO: 116 and the VL amino acid sequence of SEQ ID NO: 123;

j . the VH amino acid sequence of SEQ ID NO: 130 and the VL amino acid sequence of SEQ ID NO: 137;

k. the VH amino acid sequence of SEQ ID NO: 144 and the VL amino acid sequence of SEQ ID NO: 151;

1. the VH amino acid sequence of SEQ ID NO: 158 and the VL amino acid sequence of SEQ ID NO: 165;

m. the VH amino acid sequence of SEQ ID NO: 172 and the VL amino acid sequence of SEQ ID NO: 179;

n. the VH amino acid sequence of SEQ ID NO: 186 and the VL amino acid sequence of SEQ ID NO: 193;

0. the VH amino acid sequence of SEQ ID NO: 200 and the VL amino acid sequence of SEQ ID NO: 205;

p. the VH amino acid sequence of SEQ ID NO: 210 and the VL amino acid sequence of SEQ ID NO: 214;

q. the VH amino acid sequence of SEQ ID NO: 218 and the VL amino acid sequence of SEQ ID NO: 222;

r. the VH amino acid sequence of SEQ ID NO: 226 and the VL amino acid sequence of SEQ ID NO: 230;

s. the VH amino acid sequence of SEQ ID NO: 234 and the VL amino acid sequence of SEQ ID NO: 238; or

t. the VH amino acid sequence of SEQ ID NO: 242 and the VL amino acid sequence of SEQ ID NO: 246.

The combination of any one of the preceding claims, wherein the anti-CD32b antibody molecule comprises a heavy chain and a light chain amino acid sequence comprising:

a. the heavy chain amino acid sequence of SEQ ID NO: 6 and the light chain amino acid sequence of SEQ ID NO: 13;

b. the heavy chain amino acid sequence of SEQ ID NO: 20 and the light chain amino acid sequence of SEQ ID NO: 27;

c. the heavy chain amino acid sequence of SEQ ID NO: 34 and the light chain amino acid sequence of SEQ ID NO: 41;

d. the heavy chain amino acid sequence of SEQ ID NO: 48 and the light chain amino acid sequence of SEQ ID NO: 55;

e. the heavy chain amino acid sequence of SEQ ID NO: 62 and the light chain amino acid sequence of SEQ ID NO: 69;

f. the heavy chain amino acid sequence of SEQ ID NO: 76 and the light chain amino acid sequence of SEQ ID NO: 83;

g. the heavy chain amino acid sequence of SEQ ID NO: 90 and the light chain amino acid sequence of SEQ ID NO: 97;

h. the heavy chain amino acid sequence of SEQ ID NO: 104 and the light chain amino acid sequence of SEQ ID NO: 111;

1. the heavy chain amino acid sequence of SEQ ID NO: 118 and the light chain amino acid sequence of SEQ ID NO: 125;

j . the heavy chain amino acid sequence of SEQ ID NO: 132 and the light chain amino acid sequence of SEQ ID NO: 139;

k. the heavy chain amino acid sequence of SEQ ID NO: 174 and the light chain amino acid sequence of SEQ ID NO: 181; or

1. the heavy chain amino acid sequence of SEQ ID NO: 188 and the light chain amino acid sequence of SEQ ID NO: 195.

9. The combination of any one of the preceding claims, wherein the anti-CD32b antibody molecule is afucosylated.

10. The combination according to claim 1 comprising the anti-CD32b antibody molecule and IL-15/IL- 15Ra complex separately or together.

11. The combination according to claim 1 for use as a medicament, wherein anti-CD32b antibody molecule and the IL-15/IL-15Ra complex are administered simultaneously or sequentially.

12. The combination according to any of the preceding claims comprising a quantity of the anti-CD32b antibody molecule and IL-15/IL-15Ra complex which is therapeutically effective for the treatment of B cell malignancies, Hodgkins lymphoma, Non-Hodgkins lymphoma, multiple myeloma, diffuse large B cell lymphoma, acute lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse small cleaved cell lymphoma, MALT lymphoma, mantel cell lymphoma, marginal zone lymphoma, follicular lymphoma, systemic light chain amyloidosis or haematological malignancies.

13. The combination according to any of claims 1 to 12 comprising a quantity of the anti-CD32b antibody molecule and IL-15/IL-15Ra complex which is therapeutically effective for the treatment of a cancer that has become resistant or refractory to therapy with an anti-CD32b antibody molecule.

14. Use of a combination according to claim 1 for the manufacture of a medicament for the treatment of cancer.

15. The use according to claim 14, wherein the cancer is a B cell malignancy, Hodgkins lymphoma, Non- Hodgkins lymphoma, multiple myeloma, diffuse large B cell lymphoma, acute lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse small cleaved cell lymphoma, MALT lymphoma, mantel cell lymphoma, marginal zone lymphoma, follicular lymphoma, systemic light chain amyloidosis or haematological malignancies.

16. A method for the treatment of multiple myeloma, chronic lymphocytic leukemia (CLL), non- Hodgkins lymphoma (NHL) or B-cell malignancies, the method comprising administering an effective amount of the combination of any one of claims 1 to 11 to a subject in need thereof.

17. A method for the treatment of a cancer, the method comprising administering an effective amount of the combination of any of claims 1 to 11 to a subject in need thereof, wherein the cancer is resistant or refractory therapy with an anti-CD32b antibody molecule.

18. The method of claim 16 or claim 17, wherein the anti-CD32b antibody molecule and the IL-15/IL- 15Ra complex are administered simultaneously or sequentially.

19. The use of claims 14-15 or the method of claims 16-18, further comprising administering a chemotherapy agent.

20. A method of enhancing ADCC activity of an anti-CD32b antibody comprising administering an effective amount of an IL-15/IL-15Ra complex in combination with an anti-CD32b antibody.

21. The method of claim 20, wherein the anti-CD32b antibody comprises heavy and light chain variable domain CDRs comprising:

a. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 1, 2, and 3, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 8, 9, and 10, respectively;

b. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 15, 16, and 17, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 22, 23, and 24, respectively;

c. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 29, 30, and 31 respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 36, 37, 38, respectively;

d. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 43, 44, and 45, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 50, 51, 52, respectively;

e. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 57, 58, and 59, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 64, 65, and 66, respectively;

f. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 71, 72, and 73, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 78, 79, and 80, respectively;

g. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 85, 86, and 87, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 92, 93, and 94, respectively;

h. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 99, 100, and 101, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 106, 107, and 108, respectively;

i. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 1 13, 114, and 115, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 120, 121, and 122, respectively;

j . the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 127, 128, and 129, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 134, 135, and 136, respectively; k. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 141, 142, and 143, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 148, 149, and 150, respectively;

1. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 155, 156, and 157, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 162, 163, and 164, respectively;

m. the HCDR1, HCDR2, and HCDR3 sequences of SEQ ID NOs: 169, 170, and 171, respectively, and the LCDR1, LCDR2, and LCDR3 sequences of SEQ ID NOs: 176, 177, and 178, respectively;

wherein the CDRs are numbered according to Kabat numbering.

22. The method of claim 20 or claim 21, wherein the IL-15/IL-15Ra complex is a heterodimeric complex of human IL-15 and human soluble IL-15Ra.

23. The method of claim 22, wherein the human IL-15 comprises residues 49 to 162 of the amino acid sequence of SEQ ID NO: 251 and the human soluble IL-15Ra comprises the amino acid sequence of SEQ ID NO: 260.

24. The method of any one of claims 20-23, wherein the anti-CD32b antibody molecule comprises a heavy chain variable domain comprising an amino acid sequence at least 90% identical to any of SEQ ID NO: 4, 18, 32, 46, 60, 74, 88, 102, 116, 130, 144, 158, 172, 186, 200, 210, 218, 226, 234 or 242.

25. The method of any one of claims 20-24, wherein the anti-CD32b antibody molecule comprises a light chain variable domain comprising an amino acid sequence at least 90% identical to any of SEQ ID NO: 11, 25, 39, 53, 67, 81, 95, 109, 123, 137, 151, 165, 179, 193, 205, 214, 222, 230, 238 or 246.

26. The method of any one of claims 20-25, wherein the anti-CD32b antibody molecule comprises a VH and a VL amino acid sequence comprising:

11;

m. the VH amino acid sequence of SEQ ID NO: 172 and the VL amino acid sequence of SEQ ID NO: 179; n. the VH amino acid sequence of SEQ ID NO: 186 and the VL amino acid sequence of SEQ ID NO: 193;

27. The method of any one of claims 20-26, wherein the anti-CD32b antibody molecule comprises a heavy chain and a light chain amino acid sequence comprising:

j . the heavy chain amino acid sequence of SEQ ID NO: 132 and the light chain amino acid sequence of SEQ ID NO: 139; k. the heavy chain amino acid sequence of SEQ ID NO: 174 and the light chain amino acid sequence of SEQ ID NO: 181; or

the heavy chain amino acid sequence of SEQ ID NO: 188 and the light chain amino acid sequence of SEQ ID NO: 195..

28. The method of any one of claims 20-27, wherein the anti-CD32b antibody molecule is aiucosylated.

29. The method of claim 20 comprising administering the anti-CD32b antibody molecule and IL-15/IL- 15Ra complex separately or together.

30. The method of claim 20 comprising administering the anti-CD32b antibody molecule and IL-15/IL- 15Ra complex simultaneously or sequentially.

31. The method according to any one of claims 20-30, wherein enhancement of ADCC is therapeutically effective for the treatment of B cell malignancies, Hodgkins lymphoma, Non-Hodgkins lymphoma, multiple myeloma, diffuse large B cell lymphoma, acute lymphocytic leukemia, chronic lymphocytic leukemia, small lymphocytic lymphoma, diffuse small cleaved cell lymphoma, MALT lymphoma, mantel cell lymphoma, marginal zone lymphoma, follicular lymphoma, systemic light chain amyloidosis or haematological malignancies.