Plasma kallikrein inhibitors for treating attacks of hereditary angioedema and use thereof

Abstract

To provide methods for treating hereditary angioedema (HAE) attacks or reducing the rate of HAE attacks.SOLUTION: A method includes administering to a subject in need thereof an antibody that binds to human plasma kallikrein at about 300 mg about every two weeks for a first treatment period of about four to nine months, monitoring the subject for HAE attacks during the first treatment period, and reducing the dose of the antibody to about 300 mg about every four weeks in the subject who does not have HAE attacks during the first treatment period, the antibody comprising the same heavy chain complementarity determining regions (CDRs) and the same light chain CDRs as DX-2930.SELECTED DRAWING: Figure 5B

Description

[Technical field]

[0001] Background technology Plasma kallikrein is a component of the serine proteases of the contact system and is a promising drug target for different inflammatory, cardiovascular, infectious (sepsis), and oncological diseases (Sainz IM et al., Thromb Haemost 98, 77-83, 2007). The contact system is activated on the endothelial cell surface by factor XIIa after exposure to foreign or negatively charged surfaces or by prolylcarboxypeptidase (Sainz IM et al., Thromb Haemost 98, 77-83, 2007). Activation of plasma kallikrein amplifies intrinsic coagulation via its feedback activation of factor XII and enhances inflammation via the production of the proinflammatory nonapeptide bradykinin. As the major kininogenase in the circulation, plasma kallikrein contributes significantly to the production of bradykinin in the vasculature. A genetic deficiency in the C1 inhibitor protein (C1-INH), the major natural inhibitor of plasma kallikrein, leads to hereditary angioedema (HAE). HAE patients suffer from acute attacks of painful edema, often precipitated by triggers of unknown origin (Zuraw BL et al., N Engl J Med 359, 1027-1036, 2008). Summary of the Invention

[0002] Provided herein are regimens for treating hereditary angioedema (HAE) attacks, reducing the rate of HAE attacks, or preventing HAE attacks using antibodies capable of binding to and inhibiting active human plasma kallikrein (pKal), such as an antibody having the same complementarity determining regions (CDRs) as DX-2930 (SHP643, also known as lanadelumab).

[0003] In an aspect, the disclosure provides a method for treating attacks of hereditary angioedema (HAE) or reducing the rate of HAE attacks, comprising: (i) administering (e.g., subcutaneously) to a subject in need thereof an antibody that binds human plasma kallikrein at about 300 mg about every two weeks for a first treatment period of about 4 to 9 months (e.g., 6 months); (ii) monitoring the subject for HAE attacks during the first treatment period; and (iii) reducing the dose of the antibody to about 300 mg about every four weeks in subjects who do not have HAE attacks during the first treatment period.

[0004] In some cases, the antibody used in the methods described herein comprises the same heavy chain complementarity determining regions (CDRs) and the same light chain CDRs as DX-2930. For example, the antibody comprises a heavy chain immunoglobulin variable domain (V) of SEQ ID NO:3. H ) and the light chain immunoglobulin variable domain of SEQ ID NO: 4 (V L ). Such an antibody may be a full-length antibody (e.g., an IgG1 molecule). Alternatively, the antibody may be an antigen-binding fragment thereof. In one example, the antibody may comprise a heavy chain of SEQ ID NO:1 and a light chain of SEQ ID NO:2.

[0005] Any of the antibodies described herein may be formulated into a pharmaceutical composition comprising a pharma- ceutically acceptable carrier. In some embodiments, the pharmaceutical composition may comprise sodium phosphate, citric acid, histidine, sodium chloride, and polysorbate 80. In some examples, the pharmaceutical composition comprises sodium phosphate at a concentration of about 30 mM, citric acid at a concentration of about 19 mM, histidine at a concentration of about 50 mM, sodium chloride at a concentration of about 90 mM, and polysorbate 80 at about 0.01%.

[0006] The subject treated with any of the methods described herein may be a human patient with low body weight, for example, a human patient weighing less than 35 kg. In some examples, the human patient is a pediatric patient. The subject may be a human patient having or suspected of having HAE (e.g., type I or type II), or at risk of HAE.

[0007] The details of one or more embodiments of the invention are set forth in the description below. Other features and advantages of the invention will be apparent from the following drawings and detailed description of certain embodiments, as well as from the appended claims. [Brief description of the drawings]

[0008] [Figure 1]Figure 1 is a Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) diagram that records systematic literature reviews used to identify studies for indirect treatment comparisons. Asterisks (*) represent studies that were excluded as inappropriate comparators and describe the use of therapeutic interventions that lacked real-world applicability, including attenuated androgens (danazol, stanozolol, methyltestosterone, fluoxymesterone, oxymetholone, and tibolone; n = 17), SC C1-INH / recombinant human hyaluronidase combination therapy (n = 1), and SC C1-INH (n = 2). Daggers (†) represent inappropriate studies that were excluded due to differences and / or lack of clarity in study design, study population, therapeutic intervention, endpoints, and / or patient characteristics, as summarized in Table 2. The double dagger (‡) represents the exclusion of four unnecessary nonrandomized controlled trials when one randomized controlled trial on IV C1-INH (Cinryze®) was identified as appropriate for NMA. Four nRCTs on this same intervention were excluded. C1-INH, C1 esterase inhibitor; IV, intravenous; NMA, network meta-analysis; nRCT, nonrandomized controlled trial; PRISMA, Preferred Reporting Items for Systematic Reviews and Meta-Analyses; RCT, randomized controlled trial; SC, subcutaneous. [Diagram 2] Figure 2 is the final network diagram for the indirect treatment comparison of lanadelumab (HELP trial; Banerji et al., JAMA. 320(20):2108-21, 2018.) and IV C1-INH (CHANGE trial; Zuraw et al., N Engl J Med. 363(6):513-22, 2010). The CHANGE trial is a crossover study. C1-INH, C1 esterase inhibitor; IV, intravenous; q2w, every 2 weeks; q4w, every 4 weeks; SC, subcutaneous. [Diagram 3]Figure 3 includes the rate ratios (95% confidence intervals, CrI) of attacks for all treatments compared to placebo. Rate ratios for lanadelumab vs placebo are from data from the HELP study and are based on 26 weeks of treatment. Rate ratios for IV C1-INH vs placebo are from data from the CHANGE study and are based on 12 weeks of treatment. C1-INH, C1 esterase inhibitor; CrI, confidence interval; IV, intravenous; q2w, every 2 weeks; q4w, every 4 weeks; SC, subcutaneous. [Figure 4] Figures 4A-4B contain hazard ratios (95% confidence intervals, CrI) for time to first seizure for all treatments compared to placebo after day 0 (Figure 4A) and after day 70 (Figure 4B). Data for IV C1-INH are from the CHANGE trial. C1-INH, C1 esterase inhibitor; CrI, confidence interval; IV, intravenous; q2w, every 2 weeks; q4w, every 4 weeks; SC, subcutaneous. [Diagram 5] Figures 5A-5B include predicted survival curves for time to first seizure after day 0 (Figure 5A) and after day 70 (Figure 5B). The data is projected as the median percentage (95% confidence interval) of patients who were seizure-free at 60 months. The data for IV C1-INH are from the CHANGE study. The data in Figure 5A are as follows: lanadelumab (300 SC q2w), 26.00%, lanadelumab (300 SC q4w), 13.70%, lanadelumab (150 SC q4w), 17.60%, IV C1 inhibitor, 6.50%, and placebo, 0.60%. Data in Figure 5B are as follows: lanadelumab (300 SC q2w), 46.20%, lanadelumab (300 SC q4w), 9.30%, lanadelumab (150 SC q4w), 16.50%, IV C1 inhibitor, 0.90%, and placebo, 0.00%. C1-INH, C1 esterase inhibitor; IV, intravenous; q2w, every 2 weeks; q4w, every 4 weeks; SC, subcutaneous. [Figure 6] Figure 6 is the evidence network for the comparison criteria of the HELP (HELP-03) and CHANGE trials. Cinryze® is a C1-INH, C1 esterase inhibitor. [Figure 7] FIG. 7 is a graph of predicted survival comparing time to first attack after 0 days of treatment with placebo in the HELP (HELP-03) and CHANGE trials. [Figure 8] FIG. 8 is a graph of predicted survival comparing time to first attack after 70 days of treatment with placebo in the HELP (HELP-03) and CHANGE trials. [Figure 9] FIG. 9 is a graph of predicted survival comparing time to first attack after 0 days of treatment with placebo or Cinryze® (C1-INH) in the CHANGE study. [Figure 10] FIG. 10 is a graph of predicted survival comparing time to first seizure after 0 days of treatment with placebo, lanadelumab 300 mg every 2 weeks (300 mg q2w), or lanadelumab 300 mg every 4 weeks (300 mg q4w) in the HELP (HELP-03) trial. [Figure 11] FIG. 11 is a graph of predicted survival comparing time to first attack after 70 days of treatment with placebo or Cinryze® (C1-INH) in the CHANGE study. [Figure 12] FIG. 12 is a graph of predicted survival comparing time to first seizure after 70 days of treatment with placebo, 300 mg lanadelumab every 2 weeks (300 mg q2w), or 300 mg lanadelumab every 4 weeks (300 mg q4w) in the HELP (HELP-03) trial. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0009] definition For convenience, before further description of the present invention, certain terms employed in the specification, examples, and appended claims are defined here. Other terms are defined as they appear herein.

[0010] The singular forms "a," "an," and "the" include the plural forms unless the context clearly dictates otherwise.

[0011] The term "about" or "approximately" means within an acceptable range of error for a particular value as determined by one of ordinary skill in the art, which depends in part on how the value is measured or determined, i.e., the limitations of the measurement system. For example, "about" can mean within a range of standard deviations accepted through practice in the art. Alternatively, "about" can mean within a range of up to ±20%, preferably up to ±10%, more preferably up to ±5%, and even more preferably ±1% of a given value. Alternatively, particularly with respect to biological systems or processes, the term can mean within an order of magnitude, preferably within a two-fold range, of a value. When a particular value is described in the present application and claims, unless otherwise specified, the term "about" is implicit and means within an acceptable range of error for the particular value in this context.

[0012] The term "antibody" refers to an immunoglobulin molecule capable of specifically binding to a target, such as a carbohydrate, polynucleotide, lipid, or polypeptide, through at least one antigen recognition site located in the variable region of the immunoglobulin molecule. Antibodies are made up of heavy chain immunoglobulin variable domains (V H At least one heavy (H) chain, light chain immunoglobulin variable domain (V L For example, an antibody can include at least one light chain comprising a heavy (H) chain variable region (referred to herein as V H or HV) and the light (L) chain variable region (abbreviated as V L or LV). In another example, the antibody comprises two heavy (H) chain variable regions and two light (L) chain variable regions.

[0013] As used herein, the term "antibody" encompasses not only intact (i.e., full-length) polyclonal or monoclonal antibodies, but also antigen-binding fragments thereof (Fab, Fab', F(ab')2, Fv, etc.), single chain (scFv), domain antibody (dAb) fragments (de Wildt et. al., Euro. J. Immunol. (1996) 26(3): 629-639), variants of any of these, fusion proteins comprising antibody portions, humanized antibodies, chimeric antibodies, diabodies, linear antibodies, single chain antibodies, multispecific antibodies (e.g., bispecific antibodies), and any other modified configuration of an immunoglobulin molecule that contains an antigen recognition site of the required specificity, including glycosylation variants of antibodies, amino acid sequence variants of antibodies, and covalently modified antibodies. Antibodies include antibodies of any class, such as IgD, IgE, IgG, IgA, or IgM (or subclasses thereof), and the antibody need not be of any particular class. Depending on the amino acid sequence of the constant domain of the antibody's heavy chain, immunoglobulins can be assigned to different classes. There are five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, some of which can be further divided into subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant domains corresponding to the different classes of immunoglobulins are called alpha, delta, epsilon, gamma, and mu, respectively. The structures and three-dimensional organization of the subunits of the different classes of immunoglobulins are well known. Antibodies can be from any source, although primate (human and non-human primate) and primatized antibodies are preferred.

[0014] V H Area and / or V LA region may include all or a portion of the amino acid sequence of a naturally occurring variable domain. For example, the sequence may exclude one, two or more N- or C-terminal amino acids, internal amino acids, may include one or more insertions or additional terminal amino acids, or may include other alterations. In one embodiment, a polypeptide comprising an immunoglobulin variable domain sequence may combine with another immunoglobulin variable domain sequence to form an antigen-binding site, e.g., a structure that preferentially interacts with plasma kallikrein.

[0015] V H Area and V L The regions can be further subdivided into regions of hypervariability called "complementarity determining regions" ("CDRs"), which incorporate regions that are more conserved than complementarity determining regions, called "framework regions" ("FRs"). The extent of the framework regions and CDRs has been defined (see Kabat, EA, et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, US Department of Health and Human Services, NIH Publication No. 91-3242 and Chothia, C. et al. (1987) J. Mol. Biol. 196:901-917). The Kabat definition is used herein. Each VH and VL is usually composed of three CDRs and four FRs, arranged from amino terminus to carboxy terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.

[0016] V H Area or V LIn addition to the variable region, the heavy or light chain of the antibody may further comprise all or a portion of a heavy or light chain constant region. In one embodiment, the antibody is a tetramer of two heavy and two light immunoglobulin chains, where the heavy and light immunoglobulin chains are linked to each other, for example, by disulfide bonds. In IgG, the heavy chain constant region comprises three immunoglobulin domains, CH1, CH2, and CH3. The light chain constant region comprises a CL domain. The variable regions of the heavy and light chains comprise a binding domain that interacts with an antigen. The constant region of the antibody typically mediates the binding of the antibody 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 light chain of the immunoglobulin may be of type kappa or lambda. In one embodiment, the antibody is glycosylated. The antibody may be functional for antibody-dependent cellular cytotoxicity and / or complement-mediated cytotoxicity.

[0017] One or more regions of the antibody may be human or effectively human. For example, one or more of the variable regions may be human or effectively human. For example, one or more CDRs may be human, e.g., HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3. The light chain (LC) and / or heavy chain (HC) CDRs may each be human. The HC CDR3 may be human. One or more framework regions may be human, e.g., FR1, FR2, FR3, and / or FR4 of the HC and / or LC. For example, the Fc region may be human. In one embodiment, all framework regions are human, e.g., derived from a human somatic cell, e.g., a hematopoietic cell or a non-hematopoietic cell that produces immunoglobulins. In one embodiment, the human sequences are germline sequences, e.g., encoded by a germline nucleic acid. In one embodiment, the framework (FR) residues of the selected Fab may be converted to the amino acid species of the corresponding residues in the most similar primate germline genes, particularly human germline genes. One or more constant regions can be human or effectively human, e.g., at least 70, 75, 80, 85, 90, 92, 95, 98, or 100% of an immunoglobulin variable domain, a constant region, a constant domain (CH1, CH2, CH3, and / or CL1), or an entire antibody can be human or effectively human.

[0018] Antibodies may be encoded by immunoglobulin genes or segments thereof. Exemplary human immunoglobulin genes include kappa, lambda, alpha (IgA1 and IgA2), gamma (IgG1, IgG2, IgG3, IgG4), delta, epsilon, and mu constant region genes, as well as many immunoglobulin variable region genes. Full-length immunoglobulin "light chains" (about 25 KDa or about 214 amino acids) are encoded by an NH2-terminal variable region gene (about 110 amino acids) and a COOH-terminal kappa or lambda constant region gene. Full-length immunoglobulin "heavy chains" (about 50 KDa or about 446 amino acids) are similarly encoded by a variable region gene (about 116 amino acids) and one of the other above constant region genes, such as gamma (encoding about 330 amino acids). The length of the human HC varies widely, with the HC CDR3 varying from about 3 amino acid residues to more than 35 amino acid residues.

[0019] The term "antigen-binding fragment" of a full-length antibody refers to one or more fragments of a full-length antibody that retain the property of specifically binding to a target of interest. Examples of binding fragments that are included within the term "antigen-binding fragment" of a full-length antibody and that retain functionality include (i) Fab fragments (V L Domain, V H Domain, C L (ii) F(ab')2 fragment (a bivalent fragment containing two Fab fragments linked by a disulfide bridge at the hinge region); (iii) V H (iv) an Fd fragment consisting of the V domain and the CH1 domain of a single arm of an antibody; L Domains and V H (v) an Fv fragment consisting of a V domain; H and (vi) isolated complementarity determining regions (CDRs). In addition, the two domains of the Fv fragment, the VL and V H Although the VFv and VFv are encoded by separate genes, they can be recombined using recombinant methods to form monovalent molecules known as single chain Fvs (scFvs). L Area and V H The antibody fragments can be linked by a synthetic linker that allows the regions to be produced as a single paired protein chain.See, for example, U.S. Patent Nos. 5,260,203, 4,946,778, and 4,881,175; Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883.Antibody fragments can be obtained using any suitable technique, including conventional techniques known to those skilled in the art.

[0020] The term "monospecific antibody" refers to an antibody that exhibits a single binding specificity and affinity for a particular target, e.g., epitope. This term includes "monoclonal antibody" or "monoclonal antibody composition" and is used herein to refer to a preparation of antibodies or fragments thereof of single molecular composition, regardless of how the antibody was made. An antibody may be "germlined" by reverting one or more non-germline amino acids in the framework regions to the germline amino acid of the corresponding antibody, so long as the binding characteristics are substantially retained.

[0021] Inhibition constant (K i ) is the unit of measure of inhibitory potency; it is the concentration of inhibitor required to reduce enzyme activity by half and is independent of the enzyme or substrate concentration. Apparent K i (K i,app ) is obtained at different substrate concentrations by measuring the inhibitory effect of different concentrations of an inhibitor (e.g., an inhibitory binding protein) on the rate of reaction (e.g., enzyme activity); the apparent K is determined by fitting the change in pseudo-first-order rate constant as a function of inhibitor concentration to the Morrison equation (Eq. 1). i We obtain an estimate of K i is Ki,app It is obtained from the y-intercept derived from a linear regression analysis of a plot of versus substrate concentration.

number

[0022] As used herein, "binding affinity" refers to the apparent binding constant or K A Represents K. A is the dissociation constant (K D ) is the reciprocal of the K for binding the second target. A binding antibody can have, for example, a binding affinity of at least 10, 10, 10, 10, 10, 10, and 10 M for a particular target molecule, e.g., plasma kallikrein. A higher affinity binding of a binding antibody for a first target compared to a second target is expressed as a K for binding the second target. A (or a number K D ) for binding of the first target A (or a small number K D In such cases, the binding antibody has specificity for a first target (e.g., a protein in a first conformation, or a mimic thereof) compared to a second target (e.g., the same protein in a second conformation, or a mimic thereof; or a second protein). The difference in binding affinity (e.g., with respect to specificity or other comparison) may be at least 1.5, 2, 3, 4, 5, 10, 15, 20, 30, 40, 50, 70, 80, 90, 100, 500, 1000, 10,000, or 10 5 It could be double.

[0023] Binding affinity can be determined by a variety of methods, including equilibrium dialysis, equilibrium binding, gel filtration, ELISA, surface plasmon resonance, or spectroscopy (e.g., using a fluorescence assay). Exemplary conditions for evaluating binding affinity are in HBS-P buffer (10 mM HEPES (pH 7.4), 150 mM NaCl, 0.005% (v / v) surfactant P20). These techniques can be used to measure the concentration of bound and free binding protein as a function of the binding protein (or target) concentration. The concentration of bound binding protein ([Bound]) is related to the concentration of free binding protein ([Free]) and the concentration of binding sites for the binding protein on the target, where (N) is expressed by the following formula:

number

[0024] However, in some cases, it is sufficient to obtain a quantitative measure of affinity, determined using methods such as, for example, ELISA or FACS analysis, which may be used to measure affinity, such as K A and can therefore be used in comparisons, e.g., to determine higher order affinities, e.g., 2-fold higher affinities, to obtain a qualitative measure of affinity, or to obtain an inference of affinity, e.g., by activity in a functional assay, e.g., in vitro or in vivo assay, K A It is not necessary to make an exact determination of.

[0025] The term "binding antibody" (or "binding protein" as used interchangeably herein) refers to an antibody that can interact with a target molecule. The term "target molecule" is used interchangeably with "ligand." A "plasma kallikrein binding antibody" refers to an antibody that can interact with (e.g., bind to) plasma kallikrein, and in particular includes antibodies that preferentially or specifically interact with and / or inhibit plasma kallikrein. An antibody inhibits plasma kallikrein if it causes a decrease in the activity of plasma kallikrein compared to the activity of plasma kallikrein in the absence of the antibody under the same conditions.

[0026] A "conservative amino acid substitution" is one in which an amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues having similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine).

[0027] The binding protein may include one or more mutations (e.g., substitutions (e.g., conservative or non-essential amino acid substitutions), insertions, or deletions) relative to the binding proteins described herein with respect to one or more framework and / or CDR amino acid residues of the binding protein. The plasma kallikrein binding protein may have mutations (e.g., substitutions (e.g., conservative or non-essential amino acid substitutions), insertions, or deletions) relative to the binding proteins described herein (e.g., at least 1, 2, 3, or 4, and / or less than 15, 12, 10, 9, 8, 7, 6, 5, 4, 3, or 2 mutations), e.g., mutations that have no substantial effect on the function of the protein. The mutations may be present in the framework regions, CDRs, and / or constant regions. In some embodiments, the mutations are present in the framework regions. In some embodiments, the mutations are present in the CDRs. In some embodiments, the mutations are present in the constant regions. Whether a particular substitution will be tolerated, i.e., will not adversely affect biological properties such as binding activity, can be predicted, for example, by assessing whether the mutation is conservative or by the methods of Bowie, et al. (1990) Science 247:1306-1310.

[0028] An "effectively human" immunoglobulin variable region is an immunoglobulin variable region that contains a sufficient number of human framework amino acid positions such that the immunoglobulin variable region does not elicit an immunogenic response in normal humans. An "effectively human" antibody is an antibody that contains a sufficient number of human amino acid positions such that the antibody does not elicit an immunogenic response in normal humans.

[0029] "Epitope" refers to a site on a target compound that is bound by a binding protein (e.g., an antibody such as a Fab or full-length antibody). When the target compound is a protein, the site may be composed entirely of amino acid building blocks, entirely of chemical modifications of the protein's amino acids (e.g., glycosyl sites), or a combination thereof. Overlapping epitopes contain at least one common amino acid residue, glycosyl group, phosphate group, sulfate group, or other molecular feature.

[0030] A "humanized" immunoglobulin variable region is an immunoglobulin variable region that has been modified to contain a sufficient number of human framework amino acid positions such that the immunoglobulin variable region does not elicit an immunogenic response in normal humans. Descriptions of "humanized" immunoglobulins include, for example, U.S. Patent Nos. 6,407,213 and 5,693,762.

[0031] An "isolated" antibody refers to an antibody that is at least 90% removed from at least one component of the natural sample from which the isolated antibody is obtained. An antibody can be "at least" a particular degree of purity if the species or population of species of interest is at least 5, 10, 25, 50, 75, 80, 90, 92, 95, 98, or 99% pure on a weight / weight basis.

[0032] The methods described herein include administering multiple doses of an antibody to a human subject in need thereof. The terms "patient," "subject," or "host" may be used interchangeably. The subject may be a subject who has undergone prior treatment for HAE, such as a treatment comprising an antibody described herein. In some embodiments, the subject is a pediatric subject (e.g., an infant, child, or adolescent subject). In some embodiments, the human subject is an adolescent less than 18 years of age. In some embodiments, the human subject is an adolescent between 12 and 18 years of age. In some embodiments, the subject is less than 40 to 65 years of age.

[0033] In some embodiments, the human subject is defined by gender, e.g., in some embodiments, the subject is female.

[0034] In some embodiments, the human subject is defined by weight. In some embodiments, the human subject is less than 50 kg. In some embodiments, the human subject is between 50 kg and 75 kg. In some embodiments, the human subject is between 75 kg and 100 kg. In some embodiments, the human subject is 100 kg or more.

[0035] In some embodiments, the human subject is defined by a history or absence of laryngeal attacks. In some embodiments, the subject has experienced at least one (e.g., 1, 2, 3, 4, 5, or more) laryngeal attack (i.e., laryngeal HAE attack) prior to administration of the antibody described herein. In some embodiments, the subject has not experienced a laryngeal attack prior to administration of the antibody described herein.

[0036] The terms "prekallikrein" and "pre-plasma kallikrein" are used interchangeably herein and refer to the zymogen form of active plasma kallikrein, also known as prekallikrein.

[0037] As used herein, the term "substantially identical" (or "substantially homologous") is used herein to refer to a first amino acid or nucleic acid sequence that contains a sufficient number of identical or equivalent (e.g., having similar side chains, e.g., conservative amino acid substitutions) amino acid residues or nucleotides as a second amino acid or nucleic acid sequence such that the first and second amino acid or nucleic acid sequences have (or encode proteins that have) a similar activity, e.g., binding activity, binding preference, or biological activity. In the case of antibodies, the second antibody has the same specificity and at least 50%, at least 25%, or at least 10% of the affinity for the same antigen.

[0038] Statistical significance may be determined by any method known in the art. Exemplary statistical tests include the Student's T-test, the non-parametric Mann-Whitney U-test, and the non-parametric Wilcoxon statistical test. Some statistically significant relationships have a P-value of less than 0.05 or 0.02. Certain binding proteins may, for example, exhibit differences in specificity or binding that are statistically significant (e.g., P-value < 0.05 or 0.02). For example, the terms "induce," "inhibit," "enhance," "enhance," "increase," "reduce," and the like, which represent a distinguishable qualitative or quantitative difference between two states, may represent a difference between two states, for example, a statistically significant difference.

[0039] A "therapeutically effective dose" preferably modulates a measurable parameter, such as plasma kallikrein activity, to a statistically significant degree, or at least about 20%, more preferably at least about 40%, even more preferably at least about 60%, and even more preferably at least about 80%, compared to untreated subjects. The properties of a compound to modulate a measurable parameter, such as a disease-related parameter, can be evaluated in animal model systems that are predictive of efficacy in human disorders and conditions. Alternatively, the properties of the composition can be evaluated by testing the properties of the compound to modulate the parameter in vitro.

[0040] The term "treating" as used herein refers to the application or administration of a composition comprising one or more active agents to a subject having HAE, symptoms of HAE, suspected of having HAE, predisposed to HAE, or at risk of having HAE, with the intent to cure, heal, alleviate, mitigate, alter, remedy, ameliorate, improve, or affect the disease, symptoms of the disease, or predisposition to the disease. "Preventative treatment," also known as "prophylactic treatment," refers to treatment aimed at protecting a subject from or reducing the risk of a disease to which the person is or may be exposed. In some embodiments, the treatment methods described herein are aimed at preventing the onset and / or recurrence of HAE.

[0041] The term "preventing" a disease in a subject refers to subjecting the subject to a pharmaceutical treatment, e.g., administration of a drug, to prevent at least one symptom of the disease, administered prior to clinical manifestation of the undesirable condition (e.g., a disease in a host animal or other undesirable condition) so that the host does not develop the undesirable condition. "Preventing" a disease can also be referred to as "prevention" or "prophylactic treatment."

[0042] A "prophylactically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired prophylactic result. Typically, a prophylactically effective amount is less than a therapeutically effective amount, since a prophylactic dose is used in a subject prior to or at an earlier stage of disease.

[0043] Binding of antibodies to plasma kallikrein (pKal) Plasma kallikrein binding antibodies (anti-pKal antibodies) for use in the methods described herein may be full-length (e.g., IgG (including IgG1, IgG2, IgG3, IgG4), IgM, IgA (including IgA1, IgA2), IgD, and IgE) or may comprise only an antigen-binding fragment (e.g., Fab, F(ab')2 or scFv fragments). The binding antibody may comprise two heavy chain immunoglobulins and two light chain immunoglobulins, or may be a single chain antibody. The plasma kallikrein binding antibody may be a recombinant protein, such as a humanized antibody, a CDR-grafted antibody, a chimeric antibody, a deimmunized antibody, or an in vitro generated antibody, and may optionally comprise a constant region derived from a human germline immunoglobulin sequence. In one embodiment, the plasma kallikrein binding antibody is a monoclonal antibody.

[0044] In one aspect, the disclosure features an antibody (e.g., an isolated antibody) that binds to plasma kallikrein (e.g., human plasma kallikrein and / or mouse kallikrein) and includes at least one immunoglobulin variable region. For example, the antibody includes a heavy chain (HC) immunoglobulin variable domain sequence and / or a light chain (LC) immunoglobulin variable domain sequence. In one embodiment, the antibody binds to and inhibits plasma kallikrein, e.g., human plasma kallikrein and / or mouse kallikrein.

[0045] In some embodiments, the antibodies described herein have the same CDR sequences as DX-2930, e.g., the heavy chain CDR sequences set forth as SEQ ID NOs:5-7 and the light chain CDR sequences set forth as SEQ ID NOs:8-10. In some embodiments, the antibodies comprise the same CDR sequences as DX-2930 and an LC immunoglobulin variable domain sequence that is at least 85, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical (e.g., in whole or in framework regions) to the LC variable domain described herein. In some embodiments, the antibodies comprise the same CDR sequences as DX-2930 and an HC immunoglobulin variable domain sequence that is at least 85, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical (e.g., in whole or in framework regions) to the HC variable domain described herein. In some embodiments, the antibody comprises the same CDR sequences as DX-2930 and an LC sequence that is at least 85, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical (e.g., entirely or in the framework regions) to an LC sequence described herein. In some embodiments, the antibody comprises the same CDR sequences as DX-2930 and an HC sequence that is at least 85, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical (e.g., entirely or in the framework regions) to an HC sequence described herein.

[0046] The plasma kallikrein binding protein can be an isolated antibody (e.g., at least 70, 80, 90, 95, or 99% free of other proteins). In some embodiments, the plasma kallikrein binding antibody or composition thereof is inactive or partially active (e.g., has a K of 5000 nM or greater) compared to the plasma kallikrein binding antibody. i,appThe plasma kallikrein-binding antibodies are isolated from antibody cleavage fragments (e.g., DX-2930) that bind to plasma kallikrein at a specific site. For example, the plasma kallikrein-binding antibodies are at least 70% free of such antibody cleavage fragments, and in other embodiments, the binding antibodies are at least 80%, at least 90%, at least 95%, at least 99%, or even 100% free of inactive or partially active antibody cleavage fragments.

[0047] The plasma kallikrein binding antibody may further inhibit plasma kallikrein, for example human plasma kallikrein.

[0048] In some embodiments, the plasma kallikrein binding antibody does not bind to prekallikrein (eg, human prekallikrein and / or mouse prekallikrein), but binds to the active form of plasma kallikrein (eg, human plasma kallikrein and / or mouse kallikrein).

[0049] In certain embodiments, the antibody binds to or near the active site of the catalytic domain of plasma kallikrein or a fragment thereof, or binds to an epitope that overlaps with the active site of plasma kallikrein.

[0050] The antibody has an affinity for plasma kallikrein, e.g., human plasma kallikrein, of at least 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 10 , and 10 11 M -1 In one embodiment, the antibody can bind human plasma kallikrein with a binding affinity of 1×10 -3 , 5×10 -4 s -1 , or 1 × 10 -4 s -1 Slower K off In one embodiment, the antibody binds to human plasma kallikrein at a concentration of 1×10 2 , 1×10 3 , or 5 × 103 M -1 s -1 Faster K on In one embodiment, the antibody binds to plasma kallikrein but does not bind to tissue kallikrein and / or plasma prekallikrein (e.g., the antibody binds to tissue kallikrein and / or plasma prekallikrein less efficiently than it binds to plasma kallikrein (e.g., 5-fold, 10-fold, 50-fold, 100-fold, or 1000-fold less, or not at all, compared to a negative control).

[0051] In one embodiment, the antibody inhibits human plasma kallikrein activity, e.g., with a Ki of less than 10-5, 10-6, 10-7, 10-8, 10-9, and 10-10 M. The antibody may have an IC50 of, e.g., less than 100 nM, 10 nM, 1, 0.5, or 0.2 nM. For example, the antibody may modulate plasma kallikrein activity and the production of factor XIIa (e.g., from factor XII) and / or bradykinin (e.g., from high molecular weight kininogen (HMWK)). The antibody may inhibit plasma kallikrein activity and / or the production of factor XIIa (e.g., from factor XII) and / or bradykinin (e.g., from high molecular weight kininogen (HMWK)). The affinity of the antibody for human plasma kallikrein may be characterized by a KD of less than 100 nM, less than 10 nM, less than 5 nM, less than 1 nM, less than 0.5 nM. In one embodiment, the antibody inhibits plasma kallikrein but not tissue kallikrein (e.g., the antibody inhibits tissue kallikrein less efficiently (e.g., 5-fold, 10-fold, 50-fold, 100-fold, or 1000-fold less, or not at all, compared to a negative control) compared to inhibiting plasma kallikrein).

[0052] In some embodiments, the antibodies have an apparent inhibition constant (K i,app ).

[0053] Plasma kallikrein binding antibodies can have their HC and LC variable domain sequences contained in a single polypeptide (eg, scFv) or in different polypeptides (eg, IgG or Fab).

[0054] In one embodiment, the variable domain sequences of the HC and LC are components of the same polypeptide chain. In another embodiment, the variable domain sequences of the HC and LC are components of different polypeptide chains. For example, the antibody is an IgG, e.g., IgG1, IgG2, IgG3, or IgG4. The antibody can be a soluble Fab. In other implementations, the antibody comprises a Fab2', scFv, minibody, scFv::Fc fusion, Fab::HSA fusion, HSA::Fab fusion, Fab::HSA::Fab fusion, or other molecule that comprises an antigen combining site of one of the binding proteins herein. The VH and VL regions of these Fabs may be provided as IgG, Fab, Fab2, Fab2', scFv, PEGylated Fab, PEGylated scFv, PEGylated Fab2, VH::CH1::HSA+LC, HSA::VH::CH1+LC, LC::HSA+VH::CH1, HSA::LC+VH::CH1, or other suitable constructs.

[0055] In one embodiment, the antibody is a human or humanized antibody, or is non-immunogenic in humans. For example, the antibody comprises one or more human antibody framework regions, such as all human framework regions, or framework regions that are at least 85, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99% identical to a human framework region. In one embodiment, the antibody comprises a human Fc domain, or an Fc domain that is at least 95, 96, 97, 98, or 99% identical to a human Fc domain.

[0056] In one embodiment, the antibody is a primate antibody or primatized antibody, or is non-immunogenic in humans. For example, the antibody comprises one or more framework regions of a primate antibody, such as all primate framework regions, or framework regions that are at least 85, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99% identical to a primate framework region. In one embodiment, the antibody comprises a primate Fc domain, or an Fc domain that is at least 95, 96, 97, 98, or 99% identical to a primate Fc domain. "Primates" include humans (Homo sapiens), chimpanzees (Pan troglodytes and Pan paniscus (bonobo)), gorillas (Gorilla gorilla), gibbons (gibons), monkeys, lemurs, aye-ayes (Daubentonia madagascariensis), and tarsiers. In some embodiments, the affinity of the primate antibody for human plasma kallikrein is less than 1000 nM, less than 500 nM, less than 100 nM, less than 10 nM, less than 5 nM, less than 1 nM, less than 0.5 nM, e.g., less than 10 nM, less than 1 nM, or less than 0.5 nM. D It is characterized by:

[0057] In certain embodiments, the antibody does not contain sequences of mouse or rabbit origin (eg, is not a mouse antibody or a rabbit antibody).

[0058] In some embodiments, the antibody used in the methods described herein may be DX-2930 or a functional variant thereof as described herein.

[0059] In one example, a functional variant of DX-2930 contains the same complementarity determining regions (CDRs) as DX-2930. In another example, a functional variant of DX-2930 contains the same complementarity determining regions (CDRs) as DX-2930. H and V L Compared to the FR of V H or V LThe functional variants may contain one or more mutations (e.g., conservative substitutions) in any of the FRs of DX-2930. Preferably, such mutations do not occur at residues predicted to interact with one or more CDRs, which can be determined by routine techniques. In other embodiments, the functional variants described herein contain one or more mutations (e.g., 1, 2, or 3 mutations) in one or more CDR regions of DX-2930. Preferably, such functional variants retain the same regions / residues that contribute to antigen binding as the parent. In yet other embodiments, the functional variants of DX-2930 contain one or more mutations (e.g., 1, 2, or 3 mutations) in one or more CDR regions of DX-2930. H V comprising an amino acid sequence that is at least 85% (e.g., 90%, 92%, 94%, 95%, 96%, 97%, 98%, or 99%) identical to H Chain and / or DX-2930 V L V having an amino acid sequence that is at least 85% (e.g., 90%, 92%, 94%, 95%, 96%, 97%, 98%, or 99%) identical to L These variants can bind to the active form of plasma kallikrein and preferably do not bind to prekallikrein.

[0060] The "percent identity" of two amino acid sequences is determined using the algorithm of Karlin and Altschul Proc. Natl. Acad. Sci. USA 87:2264-68, 1990, as modified, e.g., in Karlin and Altschul Proc. Natl. Acad. Sci. USA 90:5873-77, 1993. Such an algorithm is incorporated into the NBLAST and XBLAST programs (version 2.0) of Altschul, et al. J. Mol. Biol. 215:403-10, 1990. BLAST protein searches can be performed with the XBLAST program, score=50, wordlength=3, to obtain amino acid sequences homologous to the protein molecule of interest. When gaps exist between the two sequences, Gapped BLAST can be utilized as described in Altschul et al., Nucleic Acids Res. 25(17):3389-3402, 1997. When utilizing BLAST and Gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used.

[0061] In some embodiments, the antibody used in the methods and compositions described herein may be a DX-2930 antibody. The complete sequences of the heavy and light chains and the variable sequences of DX-2930 are provided below, with the signal sequence provided in italics. The CDRs are bold and underlined. [ka] [ka] [ka] [ka]

[0062] [Table 1]

[0063] Preparation of antibodies The antibodies described herein (e.g., DX-2930) can be produced by any method known in the art, see, e.g., Harlow and Lane, (1988) Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, New York and Greenfield, (2013) Antibodies: A Laboratory Manual, Second edition, Cold Spring Harbor Laboratory Press.

[0064] The sequence encoding the antibody of interest, e.g., DX-2930, may be maintained in a vector in a host cell, which may then be propagated and frozen for further use. Alternatively, the polynucleotide sequence may be used in genetic engineering to "humanize" the antibody or improve its affinity (affinity maturation) or other characteristics. For example, if the antibody is to be used in clinical trials and treatment in humans, the constant region may be engineered to be more similar to human constant regions to avoid immune responses. It may be desirable to genetically engineer the antibody sequence to obtain higher affinity for the target antigen and higher potency in inhibiting the activity of PKal. It is clear to one skilled in the art that one or more polynucleotide changes may be made to an antibody and still maintain its binding specificity for the target antigen.

[0065] Some antibodies, e.g., Fabs, can be produced in bacterial cells, e.g., E. coli cells (see, e.g., Nadkarni, A. et al., 2007 Protein Expr Purif 52(1):219-29). For example, if the Fab is encoded by a sequence in a phage display vector that contains a suppressible stop codon between the display entity and the bacteriophage protein (or a fragment thereof), the nucleic acid of the vector can be introduced into a bacterial cell that cannot suppress the stop codon. In this case, the Fab is not fused to the gene III protein and is secreted into the periplasm and / or medium.

[0066] Antibodies may also be produced in eukaryotic cells. In one embodiment, the antibodies (e.g., scFvs) are expressed in yeast cells, such as Pichia (see, e.g., Powers et al., 2001, J. Immunol. Methods. 251:123-35; Schoonooghe S. et al., 2009 BMC Biotechnol. 9:70; Abdel-Salam, HA. et al., 2001 Appl Microbiol Biotechnol 56(1-2):157-64; Takahashi K. et al., 2000 Biosci Biotechnol Biochem 64(10):2138-44; Edqvist, J. et al., 1991 J Biotechnol 20(3):291-300), Hansenula, or Saccharomyces.The skilled artisan will be familiar with the various aspects of the present invention, including, for example, oxygen conditions (see, e.g., Baumann K., et al. 2010 BMC Syst. Biol. 4:141), osmolarity (see, e.g., Dragosits, M. et al., 2010 BMC Genomics 11:207), temperature (see, e.g., Dragosits, M. et al., 2009 J Proteome Res. 8(3):1380-92), fermentation conditions (see, e.g., Ning, D. et al. 2005 J. Biochem. and Mol. Biol. 38(3): 294-299), yeast strains (see, e.g., Kozyr, AV et al. 2004 Mol Biol (Mosk) 38(6):1067-75; Horwitz, AH. et al., 1988 Proc Natl Acad Sci USA 85(22):8678-82; Yeast antibody production can be optimized by optimizing the following: (see Bowdish, K. et al. 1991 J Biol Chem 266(18):11901-8), overexpression of proteins to enhance antibody production (see, e.g., Gasser, B. et al., 2006 Biotechol. Bioeng. 94(2):353-61), culture acidity level (see, e.g., Kobayashi H., et al., 1997 FEMS Microbiol Lett 152(2):235-42), substrate and / or ion concentration (see, e.g., Ko JH. et al., 2996 Appl Biochem Biotechnol 60(1):41-8). Additionally, yeast systems can be used to produce antibodies with extended half-lives (see, e.g., Smith, BJ. et al. 2001 Bioconjug Chem 12(5):750-756).

[0067] In one preferred embodiment, the antibody is produced in mammalian cells. Preferred mammalian host cells for expressing cloned antibodies or antigen-binding fragments thereof include Chinese Hamster Ovary (CHO) cells (e.g., as described in Kaufman and Sharp, 1982, Mol. Biol. 159:601 621, including dhfr-CHO cells described in Urlaub and Chasin, 1980, Proc. Natl. Acad. Sci. USA 77:4216-4220, used with DHFR selectable marker), lymphocytic cell lines, such as NS0 myeloma cells and SP2 cells, COS cells, HEK293T cells (J. Immunol. Methods (2004) 289(1-2):65-80), and cells derived from transgenic animals, such as transgenic mammals. For example, the cells are mammalian epithelial cells.

[0068] In some embodiments, plasma kallikrein binding antibodies are produced in plants or cell-free based systems (see, eg, Galeffi, P., et al., 2006 J Transl Med 4:39).

[0069] In addition to the nucleic acid sequences encoding the various immunoglobulin domains, recombinant expression vectors may carry additional sequences, such as sequences that control replication of the vector in a host cell (e.g., origins of replication) and selectable marker genes. The selectable marker gene facilitates selection of host cells into which the vector has been introduced (see, e.g., U.S. Pat. Nos. 4,399,216, 4,634,665, and 5,179,017). For example, typically the selectable marker gene confers resistance to drugs, such as G418, hygromycin, or methotrexate, on a host cell into which the vector has been introduced. A preferred selectable marker gene is the dihydrofolate reductase (DHFR) gene (dhfr gene with methotrexate selection / amplification). - The gene includes the p53 gene (for use in a host cell) and the neo gene (for G418 selection).

[0070] In an exemplary system for recombinant expression of an antibody or antigen-binding portion thereof, a recombinant expression vector encoding both the antibody heavy chain and the antibody light chain is transfected with dhfr - The antibody heavy and light chain genes are introduced into CHO cells. In the recombinant expression vector, the antibody heavy and light chain genes are each operably linked to an enhancer / promoter regulatory element (e.g., enhancer / promoter regulatory element from SV40, CMV, adenovirus, etc., e.g., CMV enhancer / AdMLP promoter regulatory element or SV40 enhancer / AdMLP promoter regulatory element) to drive high levels of gene transcription. The recombinant expression vector also carries a DHFR gene, which allows for the selection of CHO cells transfected with the vector using methotrexate selection / amplification. The selected transformant host cells are cultured to allow expression of the antibody heavy and light chains, and intact antibody is recovered from the culture medium. Standard molecular biology techniques are used to prepare the recombinant expression vector, transfect the host cells, select the transformants, culture the host cells, and recover the antibody from the culture medium. For example, some antibodies can be isolated by affinity chromatography using a matrix bound to protein A or protein G.

[0071] For antibodies that include an Fc domain, the antibody production system may produce antibodies in which the Fc region is glycosylated. For example, the Fc domain of an IgG molecule is glycosylated at asparagine 297 in the CH2 domain. This asparagine is the site for modification with biantennary oligosaccharides. This glycosylation has been shown to be necessary for effector functions mediated by Fcg receptors and complement C1q (Burton and Woof, 1992, Adv. Immunol. 51:1-84; Jefferis et al., 1998, Immunol. Rev. 163:59-76). In one embodiment, the Fc domain is produced in a mammalian expression system that appropriately glycosylates the residue corresponding to asparagine 297. The Fc domain may also include other eukaryotic post-translational modifications.

[0072] Antibodies can also be produced by transgenic animals. For example, U.S. Patent No. 5,849,992 describes a method for expressing an antibody in the mammary gland of a transgenic mammal. A transgene is constructed that includes a milk-specific promoter and a nucleic acid encoding the antibody of interest and a signal sequence for selection. The milk produced by the female of such a transgenic mammal contains the antibody of interest secreted therein. The antibody can be purified from the milk or, in some applications, can be used directly.

[0073] Pharmaceutical Compositions The antibodies described herein (e.g., DX-2930) can be present in a composition, e.g., a pharma- ceutically acceptable composition or pharmaceutical composition. The antibodies described herein (e.g., DX-2930) can be formulated with a pharma- ceutically acceptable carrier. In some embodiments, 150 mg or 300 mg of DX-2930 antibody is present in a composition, e.g., a pharma- ceutically acceptable composition or pharmaceutical composition, optionally including a pharma- ceutically acceptable carrier.

[0074] Pharmaceutically acceptable carriers include any and all physiologically compatible solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc. Preferably, the carrier is suitable for subcutaneous, intravenous, intramuscular, parenteral, spinal or epidermal administration (e.g., by injection or infusion), although carriers suitable for inhaled and intranasal administration are contemplated as well.

[0075] The pharma- ceutically acceptable carrier in the pharmaceutical compositions described herein may include one or more buffers, amino acids, and osmolality modifiers. Any suitable buffer or combination of buffers may be used in the pharmaceutical compositions described herein to maintain or be useful in maintaining a suitable pH of the compositions. Non-limiting examples of buffers include sodium phosphate, potassium phosphate, citric acid, sodium succinate, histidine, Tris, and sodium acetate. In some embodiments, the buffer may be at a concentration of about 5-100 mM, 5-50 mM, 10-50 mM, 15-50 mM, or about 15-40 mM. For example, the one or more buffering agents can be at a concentration of about 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, 20 mM, 21 mM, 22 mM, 23 mM, 24 mM, 25 mM, 26 mM, 27 mM, 28 mM, 29 mM, 30 mM, 31 mM, 32 mM, 33 mM, 35 mM, 36 mM, 37 mM, 38 mM, 39 mM, or about 40 mM. In some examples, the pharma- ceutically acceptable carrier comprises sodium phosphate and citric acid, which can be at a concentration of about 30 mM and about 19 mM, respectively.

[0076] In some embodiments, the pharma- ceutically acceptable carrier comprises one or more amino acids, which may reduce aggregation of the antibody during storage prior to administration and / or increase the stability of the antibody. Exemplary amino acids for use in making the pharmaceutical compositions described herein include, but are not limited to, alanine, arginine, asparagine, aspartic acid, glycine, histidine, lysine, proline, or serine. In some examples, the concentration of the amino acid in the pharmaceutical composition may be about 5-100 mM, 10-90 mM, 20-80 mM, 30-70 mM, 40-60 mM, or about 45-55 mM. In some examples, the concentration of an amino acid (e.g., histidine) can be about 40 mM, 41 mM, 42 mM, 43 mM, 44 mM, 45 mM, 46 mM, 47 mM, 48 mM, 49 mM, 50 mM, 51 mM, 52 mM, 53 mM, 54 mM, 55 mM, 56 mM, 57 mM, 58 mM, 59 mM, or about 60 mM. In one example, the pharmaceutical composition includes histidine at a concentration of about 50 mM.

[0077] Any suitable osmolarity modifying agent may be used in the preparation of the pharmaceutical compositions described herein. In some embodiments, the osmolarity modifying agent is a salt or an amino acid. Examples of suitable salts include, but are not limited to, sodium chloride, sodium succinate, sodium sulfate, potassium chloride, magnesium chloride, magnesium sulfate, and calcium chloride. In some embodiments, the osmolarity modifying agent in the pharmaceutical composition may be at a concentration of about 10-150 mM, 50-150 mM, 50-100 mM, 75-100 mM, or about 85-95 mM. In some embodiments, the osmolality modifying agent can be at a concentration of about 80 mM, 81 mM, 82 mM, 83 mM, 84 mM, 85 mM, 86 mM, 87 mM, 88 mM, 89 mM, 90 mM, 91 mM, 92 mM, 93 mM, 94 mM, 95 mM, 96 mM, 97 mM, 98 mM, 99 mM, or about 100 mM. In one example, the osmolality modifying agent can be sodium chloride, which can be at a concentration of about 90 mM.

[0078] The pharma- ceutically acceptable carrier in the pharmaceutical composition described herein may further include one or more pharma- ceutically acceptable excipients. In general, pharma- ceutically acceptable excipients are pharmacologically inactive substances. Non-limiting examples of excipients include lactose, glycerol, xylitol, sorbitol, mannitol, maltose, inositol, trehalose, glucose, bovine serum albumin (BSA), dextran, polyvinyl acetate (PVA), hydroxypropyl methylcellulose (HPMC), polyethyleneimine (PEI), gelatin, polyvinylpyrrolidone (PVP), hydroxyethylcellulose (HEC), polyethylene glycol (PEG), ethylene glycol, glycerol, dimethylsulfoxide (DMSO), dimethylformamide (DMF), polyoxyethylene terephthalate (PEI ... Examples of suitable glycerols include diethylene sorbitan monolaurate (Tween-20), polyoxyethylene sorbitan monooleate (Tween-80), sodium dodecyl sulfate (SDS), polysorbates, polyoxyethylene copolymers, potassium phosphate, sodium acetate, ammonium sulfate, magnesium sulfate, sodium sulfate, trimethylamine N-oxide, betaine, zinc ions, copper ions, calcium ions, manganese ions, magnesium ions, CHAPS, sucrose monolaurate, and 2-O-β-mannoglycerate. In some embodiments, the pharma- ceutically acceptable carrier comprises about 0.001%-0.1%, 0.001%-0.05%, 0.005%-0.1%, 0.005%-0.05%, 0.008%-0.05%, 0.008%-0.03%, or about 0.009%-0.02% of an excipient. In some embodiments, the excipient is about 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or about 0.1%. In some embodiments, the excipient is polyoxyethylene sorbitan monooleate (Tween-80). In one example, the pharma- ceutically acceptable carrier comprises 0.01% Tween-80.

[0079] In some examples, the pharmaceutical compositions described herein include an anti-pKal antibody, also described herein (e.g., DX-2930), and one or more of sodium phosphate (e.g., disodium hydrogen phosphate dihydrate), citric acid (e.g., citric acid monohydrate), histidine (e.g., L-histidine), sodium chloride, and polysorbate 80. For example, the pharmaceutical composition may include the antibody, sodium phosphate, citric acid, histidine, sodium chloride, and polysorbate 80. In some examples, the antibody is formulated in about 30 mM sodium phosphate, about 19 mM citric acid, about 50 mM histidine, about 90 mM sodium chloride, and about 0.01% polysorbate 80. The concentration of the antibody (e.g., DX-2930) in the composition may be about 150 mg / mL or 300 mg / mL. In one example, the composition comprises or consists of about 150 mg of DX-2930 per mL of solution, about 30 mM disodium hydrogen phosphate dihydrate, about 19 mM (e.g., 19.6 mM) citric acid monohydrate, about 50 mM L-histidine, about 90 mM sodium chloride, and about 0.01% polysorbate 80. In another example, the composition comprises or consists of about 300 mg of DX-2930 per mL of solution, about 30 mM disodium hydrogen phosphate dihydrate, about 19 mM (e.g., 19.6 mM) citric acid monohydrate, about 50 mM L-histidine, about 90 mM sodium chloride, and about 0.01% polysorbate 80.

[0080] Pharmaceutically acceptable salts are salts that retain the desired biological activity of the compound and do not impart any undesired toxicological effects (see, for example, Berge, SM, et al., 1977, J. Pharm. Sci. 66:1-19). Examples of such salts include acid addition salts and base addition salts. Acid addition salts include salts derived from non-toxic inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, phosphorous acid, and the like, and salts derived from non-toxic organic acids such as aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxyalkanoic acids, aromatic acids, aliphatic and aromatic sulfonic acids, and the like. Base addition salts include salts derived from alkaline earth metals such as sodium, potassium, magnesium, calcium, and the like, and salts derived from non-toxic organic amines such as N,N'-dibenzylethylenediamine, N-methylglucamine, chloroprocaine, choline, diethanolamine, ethylenediamine, procaine, and the like.

[0081] The compositions may be in various forms. These include, for example, liquid, semi-solid, and solid dosage forms, such as liquid solutions (e.g., injectable and infusible solutions), dispersions or suspensions, tablets, pills, powders, liposomes, and suppositories. The form may vary depending on the intended mode of administration and therapeutic indication. Many compositions are in the form of injectable or infusible solutions, such as compositions similar to those used for human administration of antibodies. Exemplary modes of administration are parenteral (e.g., intravenous, subcutaneous, intraperitoneal, intramuscular). In one embodiment, the plasma kallikrein binding protein is administered by intravenous infusion or injection. In another embodiment, the plasma kallikrein binding protein is administered by intramuscular injection. In another embodiment, the plasma kallikrein binding protein is administered by subcutaneous injection. In another preferred embodiment, the plasma kallikrein binding protein is administered by intraperitoneal injection.

[0082] The terms "parenteral administration" and "administered parenterally" as used herein refer to modes of administration other than enteral and topical administration, usually by injection, and include, but are not limited to, intravenous, intramuscular, intraarterial, intrathecal, intraarticular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injection and infusion. In some embodiments, the antibody is administered subcutaneously.

[0083] The composition can be formulated as a solution, microemulsion, dispersion, liposome, or other ordered structure suitable for high drug concentration. Sterile injectable solutions can be prepared by incorporating the required amount of binding protein in a suitable solvent containing one or a combination of the above-listed ingredients, and optionally performing filtered sterilization. Generally, dispersions are prepared by incorporating the effective compound into a sterile vehicle containing a basic dispersion medium and other ingredients from those listed above as required. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred preparation method is vacuum drying and freeze-drying, which results in a powder of the active ingredient plus any additional desired ingredients from its previously sterile-filtered solution. The proper fluidity of the solution can be maintained, for example, by the use of a coating agent such as lecithin, by maintaining the required particle size in the case of dispersions, and by the use of surfactants. Prolonged absorption of injectable compositions can be achieved by including in the composition an agent that delays absorption, for example, monostearate salts and gelatin.

[0084] The antibodies described herein (e.g., DX-2930) can be administered by a variety of methods, including intravenous injection, subcutaneous injection, or infusion. For example, in some therapeutic applications, the antibodies can be administered at a dose of about 1-100 mg / m 2 or 7 to 25 mg / m 2It can be administered by intravenous infusion at a rate of less than 30, 20, 10, 5, or 1 mg / min to reach a dose of 100 mg / min. The route and / or form of administration varies depending on the desired results. In certain embodiments, the effective compound can be prepared with a carrier that protects the compound from rapid release, such as a sustained release formulation, including implants and microencapsulated delivery systems. Biodegradable, biocompatible polymers such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid can be used. Many methods are available for the preparation of such formulations. See, for example, Sustained and Controlled Release Drug Delivery Systems, JR Robinson, ed., 1978, Marcel Dekker, Inc., New York.

[0085] The pharmaceutical composition may be administered using a medical device. For example, in one embodiment, the pharmaceutical compositions disclosed herein may be administered using a device such as a needleless hypodermic device, a pump, or an implant.

[0086] In certain embodiments, the antibodies described herein (e.g., DX-2930) can be formulated to ensure proper distribution in vivo. For example, the blood-brain barrier (BBB) ​​excludes many highly hydrophilic compounds. To ensure that the therapeutic compounds disclosed herein cross the BBB (if necessary), they can be formulated, for example, in liposomes. For methods of formulating liposomes, see, for example, U.S. Patent Nos. 4,522,811; 5,374,548; and 5,399,331. Liposomes can enhance targeted drug delivery by including one or more moieties that are selectively transported to specific cells or organs (see, for example, VV Ranade, 1989, J. Clin. Pharmacol. 29:685).

[0087] The dosage regimen is adjusted to provide the optimum desired response (e.g., therapeutic response). For example, a single bolus may be administered, several divided doses may be administered over time, or the dose may be proportionally reduced or increased to follow the exigencies of the therapeutic situation. It is particularly preferred to formulate parenteral compositions into dosage unit forms for ease of administration and uniformity of dosage. As used herein, dosage unit form refers to physically discrete units suitable as a unitary dose to the subject to be treated; each unit contains a predetermined amount of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. The specifications for the dosage unit forms can be determined by or directly depend on (a) the unique characteristics of the active compound and the particular therapeutic effect to be achieved, and (b) the limitations inherent in the field of compounding the active compound for the treatment of an individual's susceptibility.

[0088] An exemplary, non-limiting range for a therapeutically or preventatively effective amount of an antibody (e.g., DX-2930) described herein is about 150 mg or 300 mg. As will be appreciated by one of skill in the art, a therapeutically or preventatively effective amount of an antibody for a pediatric subject may be less than an effective amount for an adult subject. In some embodiments, the effective amount administered to a pediatric subject is a fixed dose or a weight-based dose. In some embodiments, an effective amount of less than about 150 mg or 300 mg is administered to a pediatric subject. In some embodiments, a therapeutically or preventatively effective amount of an antibody is administered every two weeks or every four weeks during a first treatment period. In some embodiments, the antibody may be administered to a subject during a second treatment period. In some embodiments, the therapeutically or preventatively effective amount of an antibody in a first treatment period is different from the therapeutically or preventatively effective amount of an antibody in a second treatment period. In some embodiments, a therapeutically or preventatively effective amount of an antibody in a first treatment period is 150 mg and a therapeutically or preventatively effective amount of an antibody in a second treatment period is 300 mg. In some embodiments, the therapeutically or prophylactically effective amount of the antibody in the first treatment period is the same as the therapeutically or prophylactically effective amount of the antibody in the second treatment period. In one example, the therapeutically or prophylactically effective amount of the antibody in the first treatment period and the second treatment period is 300 mg.

[0089] In some embodiments, an exemplary, non-limiting range for a therapeutically or prophylactically effective amount of an antibody described herein (e.g., DX-2930) is about 300 mg. In some embodiments, a therapeutically or prophylactically effective amount of an antibody is administered in a single dose. If the subject experiences an HAE attack, the antibody may be further administered to the subject in multiple doses, for example, every two weeks at a dose of about 300 mg.

[0090] kit An antibody described herein (e.g., DX-2930) may be provided as a kit, e.g., a component of a kit. For example, the kit includes (a) a DX-2930 antibody, e.g., a composition (e.g., a pharmaceutical composition) comprising the antibody, and, optionally, (b) informational material. The informational material may be descriptive, instructional, marketing, or other material related to the methods described herein and / or use of an antibody described herein (e.g., DX-2930), e.g., with respect to the methods described herein. In some embodiments, the kit includes one or more doses of DX-2930. In some embodiments, the one or more doses are 150 mg or 300 mg.

[0091] The informational material of the kit is not limited in its form. In one embodiment, the informational material may include information about the production of the compound, the molecular weight of the compound, concentration, excretion rate, batch or production site information, etc. In one embodiment, the informational material relates to the use of the antibody to treat, prevent, or diagnose disorders and conditions, e.g., diseases or conditions associated with plasma kallikrein.

[0092] In one embodiment, the informational material can include instructions for administering an antibody described herein (e.g., DX-2930) in a suitable manner, e.g., in a suitable dose, dosage form, administration format, or administration schedule (e.g., a dose, dosage form, administration schedule, or administration format described herein), for carrying out a method described herein. In another embodiment, the informational material can include instructions for administering an antibody described herein (e.g., DX-2930) to a suitable subject, e.g., a human, e.g., a human having or at risk for a plasma kallikrein-associated disease or condition. For example, the material can include instructions for administering an antibody described herein (e.g., DX-2930) to a patient having a disorder or condition described herein, e.g., a plasma kallikrein-associated disease, e.g., according to an administration schedule described herein. The informational material of the subject kits is not limited in its form. In many cases, the informational material, e.g., instructions, is provided in printed form, but can also be in other formats, such as computer readable material.

[0093] The antibody (e.g., DX-2930) described herein can be provided in any form, such as liquid, dry or lyophilized. The antibody is preferably substantially pure and / or sterile. If the antibody is provided in a liquid solution, the liquid solution is preferably an aqueous solution, preferably a sterile aqueous solution. If the antibody is provided in a dry form, reconstitution is generally by the addition of a suitable solvent. A solvent, such as sterile water or buffer, can optionally be provided in the kit.

[0094] The kit may include one or more containers for the composition comprising an antibody described herein (e.g., DX-2930). In some embodiments, the kit includes separate containers, dividers, or compartments for the composition and the informational material. For example, the composition may be included in a bottle, vial, or syringe, and the informational material may be included in association with the container. In other embodiments, the separate elements of the kit are included in a single, undivided container. For example, the composition is included in a bottle, vial, or syringe having attached thereto the informational material in the form of a label. In some embodiments, the kit includes a plurality (e.g., a pack) of separate containers, each including one or more unit dosage forms (e.g., dosage forms described herein) of an antibody described herein (e.g., DX-2930). For example, the kit includes a plurality of syringes, ampoules, foil packets, or blister packs, each including a single unit dose of an antibody described herein (e.g., DX-2930). The containers of the kit may be airtight, waterproof (e.g., impermeable to changes in moisture or vapor), and / or light-tight.

[0095] The kit optionally includes a device suitable for administration of the composition, such as a syringe, or any such delivery device. In one embodiment, the device is an implantable device that dispenses a metered dose of the antibody. The disclosure also features methods of providing the kit, such as by combining the components described herein.

[0096] treatment In some aspects, the disclosure provides for the use of an antibody described herein (e.g., DX-2930) in the treatment of HAE. In particular, the disclosure provides treatment regimens that allow for a reduced dose and / or extended dosing schedule of DX-2930 (e.g., from once every two weeks to once every four weeks) in patients whose disease is well controlled (e.g., attack-free for a period of 4-9 months, etc.) and / or in patients who are underweight (e.g., pediatric patients).

[0097] (i) Hereditary angioedema Hereditary angioedema (HAE) is also known as "Quincke's edema," C1 esterase inhibitor deficiency, C1 inhibitor deficiency, and hereditary angioneurotic edema (HANE). HAE is characterized by unpredictable, recurrent attacks of severe subcutaneous or submucosal swelling (angioedema) that can affect, for example, the limbs, face, genitals, gastrointestinal tract, and airways (Zuraw, 2008). Symptoms of HAE include, for example, swelling of the arms, legs, lips, eyes, tongue, and / or pharynx; airway obstruction, which can include swelling of the pharynx (larynx), sudden hoarseness, and / or death from asphyxiation (Bork et al., 2012; Bork et al., 2000). Approximately 50% of all HAE patients will experience a laryngeal attack in their lifetime, and there is no way to predict whether a patient is at risk for a laryngeal attack (Bork et al., 2003; Bork et al., 2006). Symptoms of HAE also include recurrent episodes of abdominal cramping without a clear cause; and / or intestinal swelling, which may be severe and result in intestinal symptoms similar to an abdominal emergency, which may result in abdominal cramping, vomiting, dehydration, diarrhea, pain, shock, and / or unnecessary surgery (Zuraw, 2008). The swelling may last up to 5 days or more. Approximately one-third of individuals with this HAE develop a non-itchy rash called erythema marginatum during attacks. Most patients suffer from multiple attacks per year.

[0098] HAE is a rare disorder and the exact prevalence is not known, but current estimates range from 1 per 10,000 to 1 per 150,000, with most authors agreeing that 1 / 50,000 may be the most reliable estimate (Bygum, 2009; Goring et al., 1998; Lei et al., 2011; Nordenfelt et al., 2014; Roche et al., 2005).

[0099] Plasma kallikrein plays an important role in the pathogenesis of HAE attacks (Davis, 2006; Kaplan and Joseph, 2010). In normal physiology, C1-INH regulates the activity of plasma kallikrein as well as various other proteases, such as C1r, C1s, factor XIa, and factor XIIa. Plasma kallikrein regulates the release of bradykinin from high molecular weight kininogen (HMWK). Deficiency of C1-INH in HAE leads to dysregulation of plasma kallikrein activity, resulting in excessive production of bradykinin. Bradykinin is a vasodilator that is believed to contribute to the characteristic symptoms of HAE, such as localized swelling, inflammation, and pain (Craig et al., 2012; Zuraw et al., 2013).

[0100] Airway swelling can be life-threatening and results in death in some patients. Mortality is estimated to be 15-33%. HAE results in approximately 15,000-30,000 emergency department visits per year.

[0101] Trauma or stress, such as dental procedures, illness (e.g., viral illnesses such as colds and flu), menstruation, and surgery, can trigger attacks of angioedema. To prevent acute attacks of HAE, patients can try to avoid certain stimuli that have previously caused attacks. However, in many cases, attacks occur without a known trigger. Symptoms of HAE usually first appear in early childhood and worsen during adolescence. On average, untreated individuals have one attack every 1-2 weeks, with most episodes lasting about 3-4 days (ghr.nlm.nih.gov / condition / hereditary-angioedema). The frequency and duration of attacks vary significantly among people with hereditary angioedema, and even among members of the same family.

[0102] There are three types of HAE, known as types I, II, and III, all of which can be treated by the methods described herein. It is estimated that 1 in 50,000 people suffer from HAE, with type I accounting for approximately 85% of cases, type II accounting for approximately 15% of cases, and type III being very rare. Type III is the most recently characterized form, and was originally thought to affect only women, but families involving affected men have been identified.

[0103] HAE is inherited in an autosomal dominant pattern, so affected individuals may have inherited a mutation from one affected parent. De novo mutations in the gene can also occur, so HAE can develop in people with no family history of the disorder. It is estimated that 20-25% of cases result from de novo, spontaneous mutations.

[0104] Mutations in the SERPING1 gene cause hereditary angioedema types I and II. The SERPING1 gene provides instructions for making the C1 inhibitor protein, which is important in controlling inflammation. C1 inhibitor blocks the activity of certain proteins that promote inflammation. Mutations that cause hereditary angioedema type I result in reduced levels of C1 inhibitor in the blood. In contrast, mutations that cause type II result in the production of an abnormally functioning C1 inhibitor. Approximately 85% of patients have type I HAE, which is characterized by very low production of functionally normal C1-INH protein, while the remaining approximately 15% have type II HAE, which produces normal or high levels of impaired functioning C1-INH (Zuraw, 2008). Without adequate levels of functional C1 inhibitor, excessive amounts of bradykinin are produced from high molecular weight kininogen (HMWK), increasing vascular leakage mediated by the binding of bradykinin to B2 receptors (B2-R) on the surface of endothelial cells (Zuraw, 2008). Bradykinin promotes inflammation by increasing leakage of fluid through blood vessel walls into body tissues. Excessive accumulation of fluid in body tissues causes episodes of swelling seen in individuals with hereditary angioedema types I and II.

[0105] Mutations in the F12 gene are associated with some cases of hereditary angioedema type III. The F12 gene provides the instructions for making clotting factor XII. In addition to playing a key role in blood clotting (clotting), factor XII is also an important stimulator of inflammation and is involved in the production of bradykinin. Certain mutations in the F12 gene result in the production of factor XII with increased activity. As a result, more bradykinin is produced and blood vessel walls become leakier, leading to episodes of swelling. The cause of other cases of hereditary angioedema type III remains unknown. Mutations in one or more as yet unidentified genes may contribute to the disorder in these cases.

[0106] HAE may present similarly to other forms of angioedema resulting from allergies or other conditions, but differs significantly in cause and treatment. When hereditary angioedema is misdiagnosed as an allergy, it is most commonly treated with antihistamines, steroids, and / or epinephrine, which can be used in life-threatening reactions, despite the fact that these are usually ineffective in HAE. Misdiagnosis has also led to unnecessary exploratory laparotomy in patients with abdominal swelling, and in some HAE patients, abdominal pain has been inaccurately diagnosed as psychosomatic.

[0107] Like adults, children with HAE can suffer from recurrent and debilitating attacks. Symptoms can be present very early in infancy, with upper airway angioedema reported in patients with HAE as young as 3 years of age (Bork et al., 2003). In one case study of 49 pediatric patients with HAE, 23 had experienced at least one episode of airway angioedema by age 18 (Farkas, 2010). Because the disease generally worsens after puberty, there is a significant unmet medical need in children with HAE, particularly in adolescents (Bennett and Craig, 2015; Zuraw, 2008).

[0108] C1 inhibitor therapy, and other treatments for HAE, are described in Kaplan, AP, J Allergy Clin Immunol, 2010, 126(5):918-925.

[0109] Acute treatment of HAE attacks is provided to stop the progression of edema as quickly as possible. C1 inhibitor concentrate from donor blood administered intravenously is one of the acute treatments; however, this treatment is not available in many countries. In emergency situations where C1 inhibitor concentrate is not available, fresh frozen plasma (FFP) can be used as an alternative, as it also contains C1 inhibitor.

[0110] Purified C1 inhibitor derived from human blood has been used in Europe since 1979. Several C1 inhibitor treatments are currently available in the United States, and two C1 inhibitor products are currently available in Canada. Verinert P (CSL Behring), which is pasteurized, was approved by the FDA for acute attacks in 2009. Nanofiltered Cinryze (ViroPharma) was approved by the FDA in 2008 for prevention. Rhucin (Pharming) is an investigational recombinant C1 inhibitor that does not carry the risk of transmission of infectious diseases from human blood-borne pathogens.

[0111] Treatment of acute HAE attacks may also include medications and / or intravenous fluids for pain relief.

[0112] Other treatment modalities may stimulate the synthesis of C1 inhibitor or reduce the consumption of C1 inhibitor. Androgen medications such as danazol may reduce the frequency and severity of attacks by stimulating the production of C1 inhibitor.

[0113] Helicobacter pylori can cause abdominal attacks. Antibiotics to treat H. pylori reduce abdominal attacks.

[0114] Newer treatments attack the contact cascade. Ecallantide (KALBITOR®, DX-88, Dyax) inhibits plasma kallikrein and is approved in the United States. Icatibant (FIRAZYR®, Shire) inhibits the bradykinin B2 receptor and is approved in Europe and the United States.

[0115] Diagnosis of HAE may depend, for example, on family history and / or blood tests. Laboratory findings related to HAEI types, II and III are described, for example, in Kaplan, AP, J Allergy Clin Immunol, 2010, 126(5):918-925. In type I HAE, C1 inhibitor levels are decreased, as are C4 levels, and C1q levels are normal. In type II HAE, C1 inhibitor levels are normal or increased, but C1 inhibitor function is abnormal. C4 levels are decreased, and C1q levels are normal. In type III, C1 inhibitor, C4, and C1q levels may all be normal.

[0116] Symptoms of HAE can be assessed, for example, using a questionnaire, for example, a questionnaire completed by the patient, clinician, or family member. Such questionnaires are known in the art and include, for example, visual analog scales. See, for example, McMillan, CV et al. Patient. 2012;5(2):113-26. In some embodiments, the subject has HAEI or HAE type II. HAEI or HAE type II can be diagnosed by any method known in the art, for example, a clinical history consistent with HAE (e.g., episodes of non-pruritic swelling subcutaneously or mucosally) or diagnostic testing (e.g., functional testing of C1-INH and evaluation of C4 levels).

[0117] (ii) Lowering the dose of DX-2930 in the treatment of HAE The present disclosure provides a method of treating hereditary angioedema (HAE) (e.g., ameliorating, stabilizing, or eliminating one or more symptoms) by administering an anti-pKal antibody, such as DX-2930, to an HAE patient on an initial dosing schedule, e.g., 300 mg every 2 weeks, for a first treatment period (e.g., 4-9 weeks). The onset of HAE attacks is monitored in patients undergoing the first period of treatment following standard medical practice. If the patient does not have an HAE attack in the first treatment period, the dose of the antibody may be reduced and / or the interval between administrations of the antibody may be extended, e.g., reduced to 300 mg every 4 weeks or 300 mg every 6 weeks.

[0118] In some embodiments, the human patient may have low body weight. As used herein, low body weight, when applied to adults, refers to an adult body weight that is significantly lower than the average body weight of adults with matched physical characteristics such as height, age, and sex. For example, an adult patient with low body weight may have a body weight that is at least 20% (e.g., 30%, 40%, 50%, or more%) lower than the average body weight of adults with the above-mentioned matched physical characteristics. In some examples, the human patient is an adult HAE patient with a body weight of less than 40 kg (e.g., less than 35 kg, less than 30 kg, less than 25 kg). In other examples, the human patient with low body weight may be a pediatric patient (e.g., younger than 15 years old). Such a pediatric patient may have a body weight of less than 30 kg (e.g., less than 25 kg, less than 20 kg, less than 15 kg, or less than 10 kg, etc.).

[0119] In some embodiments, the subject may be defined by gender, e.g., in some embodiments, the subject is female, in other embodiments, the subject is male.

[0120] In some embodiments, the human subject is defined by a history or absence of laryngeal attacks. In some embodiments, the subject has experienced at least one (e.g., one, two, three, four, five, or more) laryngeal attack (i.e., laryngeal HAE attack) prior to administration of the antibody described herein. In some embodiments, the subject has not experienced a laryngeal attack prior to administration of the antibody described herein.

[0121] Treatment includes administering an amount effective to alleviate, ameliorate, alter, cure, ameliorate, improve, or affect the disorder, the symptoms of the disorder, or the predisposition to the disorder. Treatment can also delay the onset, e.g., prevent the onset or prevent the worsening of a disease or condition.

[0122] Methods for administering the DX-2930 antibody are also described in "Pharmaceutical Compositions". The appropriate dose of the antibody used may vary depending on the age and weight of the subject and the particular drug used. The antibody may be used as a competitive agent to inhibit or reduce undesirable interactions, such as between plasma kallikrein and its substrate (e.g., factor XII or HMWK). The dose of the antibody may be sufficient to block 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.9% of the activity of plasma kallikrein in the patient, particularly at the site of disease. In some embodiments, 150 mg or 300 mg of the antibody is administered every two weeks or every four weeks. In some embodiments, the antibody is administered to the subject in a first treatment period that includes administration of 150 mg or 300 mg of the antibody every two weeks or every four weeks. In some embodiments, the antibody is administered to the subject in a second treatment period following the first treatment period. In some embodiments, 300 mg of the antibody is administered as a single dose. If the subject experiences an HAE attack after a single dose, the antibody may be administered at 300 mg every two weeks in the first treatment period.

[0123] In one embodiment, the antibodies are used to inhibit the activity of plasma kallikrein (e.g., inhibit at least one activity of plasma kallikrein, e.g., reduce production of factor XIIa and / or bradykinin), e.g., in vivo. The binding proteins may be used by themselves or may be conjugated to an agent, e.g., a cytotoxic drug, a cytotoxic enzyme, or a radioisotope.

[0124] The antibodies can be used directly in vivo to eliminate cells expressing the antigen through natural complement-dependent cytotoxicity (CDC) or antibody-dependent cellular cytotoxicity (ADCC). The antibodies described herein can contain a complement-binding effector domain, such as the Fc portion from IgG1, 2, or 3, or the corresponding portion of IgM that binds complement. In one embodiment, a population of target cells is treated ex vivo with the antibodies described herein and appropriate effector cells. This treatment can be supplemented by adding complement or serum containing complement. Furthermore, phagocytosis of target cells coated with the antibodies described herein can be improved by binding of complement proteins. In another embodiment, target cells coated with antibodies containing a complement-binding effector domain are lysed by complement.

[0125] Methods for administering DX-2930 antibody are described in "Pharmaceutical Compositions". The appropriate dose of the molecule used varies depending on the age and weight of the subject and the specific drug used. The antibody can be used as a competitive agent to inhibit or reduce undesirable interactions, such as the interaction between natural or pathological agents and plasma kallikrein.

[0126] A therapeutically effective amount of an antibody described herein can be administered to a subject having, suspected of having, or at risk for HAE to treat the disorder (e.g., ameliorate or ameliorate symptoms or characteristics of the disorder, slow, stabilize, and / or halt progression of the disease).

[0127] The antibodies described herein may be administered in a therapeutically effective amount, which is an amount effective in treating the subject, e.g., curing, alleviating, reducing, or ameliorating at least one symptom of the subject's disorder to an extent greater than would be expected in the absence of said treatment, upon administration in one or more doses to the subject.

[0128] The dosage regimen can be adjusted to provide the optimum desired response (e.g., therapeutic response). For example, a single bolus can be administered, several divided doses can be administered over time, or the dose can be proportionally reduced or increased according to the exigencies of the therapeutic situation. In other examples, a bolus can be administered followed by several administrations over time, or the dose can be proportionally reduced or increased according to the exigencies of the therapeutic situation. In other examples, the dose can be divided into several doses and administered over time. It is particularly preferred to formulate parenteral compositions into dosage unit forms for ease of administration and uniformity of dose. As used herein, dosage unit form refers to physically discrete units suitable as a unitary dose to the subject to be treated; each unit contains a predetermined amount of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier.

[0129] Any of the subjects described herein may have undergone a prior treatment for HAE, such as a preventative or therapeutic treatment for HAE. Aspects of the present disclosure also provide a method of administering an antibody described herein (e.g., DX-2930) to a subject who has undergone one or more prior treatments for HAE. In some embodiments, the prior treatment for HAE is a treatment comprising an antibody described herein (e.g., DX-2930). In some embodiments, the subject has previously received multiple doses of DX-2930 every two weeks or every four weeks. In some embodiments, the subject has previously received 150 mg of DX-2930 every two weeks. In some embodiments, the subject has previously received 300 mg of DX-2930 every two weeks. In some embodiments, the subject has previously received 300 mg of DX-2930 every four weeks. In some embodiments, multiple administrations of the pretreatment antibody are administered at least 2 times, at least 3 times, at least 4 times, at least 5 times, at least 6 times, at least 7 times, at least 8 times, at least 9 times, at least 10 times, at least 11 times, at least 12 times, at least 13 times.

[0130] In some embodiments, the subject has undergone one or more prior treatments for HAE, which may include any agent known in the art for treating HAE. Exemplary anti-HAE agents include, but are not limited to, C1 inhibitors (e.g., Cinryze®, Verinert®, or Ruconest®), plasma kallikrein inhibitors (e.g., Kalbitor®), bradykinin receptor inhibitors (e.g., Firazyr®), annenuated androgens (e.g., danazol), and antifibrinolytics (e.g., traexamic acid). In some instances, the subject may undergo a tapering period before undergoing the anti-pKal antibody treatment described herein. The tapering period refers to a period prior to anti-pKal antibody treatment during which a subject who is midway through an anti-HAE treatment (e.g., C1-INH, oral androgens, and / or oral antifibrinolytics) gradually reduces the dose, frequency, or both of the anti-HAE agent such that the subject is gradually transitioned from the previous HAE treatment to the anti-pKal antibody treatment described herein. In some embodiments, tapering involves a gradual or stepwise method of reducing the dose of pretreatment and / or the frequency with which pretreatment is administered. The tapering period may last from 2-4 weeks and may vary based on individual patient factors. In some examples, pretreatment is terminated before anti-pKal antibody treatment begins. In other examples, pretreatment may be terminated within an appropriate time frame (e.g., 2 weeks, 3 weeks, or 4 weeks) after the subject is provided with the first dose of anti-pKal antibody.

[0131] Alternatively, subjects who are in the middle of a previous HAE treatment can be transitioned directly to the anti-pKal antibody treatment described herein without a tapering period.

[0132] In other embodiments, the subject has not received any prior treatment for HAE prior to the first treatment, first treatment period, and / or subsequent single and multiple dose treatments (second treatment period) described herein. In some embodiments, the subject has not received any treatment other than the antibody described herein during the first treatment period and / or during the second treatment period. In some embodiments, the subject has not received any prior treatment for HAE for at least 2 weeks (e.g., at least 2 weeks, 3 weeks, 4 weeks, 5 weeks, or more) prior to the first treatment or first treatment period, during the first treatment or first treatment period, and / or during the second treatment period. In some embodiments, the subject has not received any long-term preventative treatment for HAE (e.g., C1 inhibitor, attenuated androgen, antifibrinolytic agent) for at least 2 weeks prior to the first treatment or first treatment period, during the first treatment period, and / or during the second treatment period. In some embodiments, the subject has not received an HAE treatment comprising an angiotensin-converting enzyme (ACE) inhibitor for at least 4 weeks prior to the first treatment or first treatment period, during the first treatment period, and / or during the second treatment period. In some embodiments, the subject has not received a medication comprising an estrogen for at least 4 weeks prior to the first treatment or first treatment period, during the first treatment period, and / or during the second treatment period. In some embodiments, the subject has not received an androgen (e.g., stanozolol, danazol, oxandrolone, methyltestosterone, testosterone) for at least 2 weeks prior to the first treatment or first treatment period, during the first treatment period, and / or during the second treatment period.

[0133] Any of the methods described herein may further include monitoring the patient before, after, or during the course of treatment for side effects due to the antibody (e.g., serum or plasma concentration of the antibody or pKal activity levels) (e.g., elevated levels of creatine phosphatase) and / or inhibition levels of pKal. If one or more adverse effects are observed, the dose of the antibody may be reduced or treatment may be terminated. If inhibition levels fall below the minimum therapeutic level, additional doses of the antibody may be administered to the patient. The patient may also be assessed for production of antibodies against the administered antibody; activity of C1 inhibitor, C4, and / or C1q; quality of life; frequency of any HAE attacks, health-related quality of life, anxiety and / or depression (e.g., Hospital Anxiety and Depression Scale (HADS)), work productivity (e.g., Work Productivity and Activity Impairment Questionnaire (WPAI)), preference for subcutaneous administration of the antibody (e.g., D-2930) compared to other injectable substances, quality of life (e.g., angioedema-quality of life (AE-QOL), EuroQoL Group 5-dimension report).

[0134] In some embodiments, the plasma or serum concentration of the antibody (e.g., DX-2930) may be measured during the course of treatment (e.g., after the first administration) to assess efficacy of the treatment. If the plasma or serum concentration of the antibody is less than about 80 nM, a subsequent administration may be required, which may be the same or higher than the first dose. The plasma or serum concentration of the antibody may be measured by determining the protein level of the antibody in a plasma or serum sample obtained from the subject, for example, by an immunoassay or MS assay. The plasma or serum concentration of the antibody may also be measured by determining the inhibition level of pKal in a plasma or serum sample obtained from a subject treated with the antibody. Such assays may include synthetic substrate assays or Western blot assays to measure cleaved kininogen as described herein.

[0135] Alternatively or additionally, plasma or serum levels of creatine kinase and / or one or more coagulation parameters (e.g., activated partial thromboplastin time (aPTT), prothrombin time (PT), bleeding events) may be monitored during the course of treatment. If plasma or serum levels of creatine kinase are found to increase during treatment, the dose of the antibody may be reduced or treatment may be terminated. Similarly, if one or more coagulation parameters are found to be significantly affected during treatment, the dose of the antibody may be modified or treatment may be terminated.

[0136] In some embodiments, the optimal dose (e.g., optimal preventative dose or optimal therapeutic dose) of the antibody (e.g., DX-2930) can be determined as follows: The antibody is provided to a subject in need of treatment with an initial dose. The plasma concentration of the antibody in the subject is measured. If the plasma concentration is less than 80 nM, the dose of the antibody is increased with subsequent administrations. The dose of the antibody that maintains the plasma concentration of the antibody above about 80 nM can be selected as the optimal dose for the subject. The subject's creatine phosphokinase levels can be monitored over the course of treatment, and the optimal dose for the subject can be further adjusted based on the creatine phosphokinase levels, for example, the dose of the antibody can be reduced if an increase in creatine phosphokinase is observed during treatment.

[0137] (iii) Combination therapy The antibodies described herein (e.g., DX-2930) can be administered in combination with one or more other therapies to treat a disease or condition associated with plasma kallikrein activity, such as a disease or condition described herein. For example, the antibodies described herein (e.g., DX-2930) can be used therapeutically or prophylactically (e.g., before, during, or after a course of treatment) with another anti-plasma kallikrein Fab or IgG (e.g., another Fab or IgG described herein), another plasma kallikrein inhibitor, a peptide inhibitor, a small molecule inhibitor, or surgery. Examples of plasma kallikrein inhibitors that can be used in combination therapy with the plasma kallikrein binding antibodies described herein include, for example, the plasma kallikrein inhibitors described in International Patent Publication No. WO 95 / 21601 or WO 2003 / 103475.

[0138] One or more plasma kallikrein inhibitors may be used in combination with the antibodies described herein (e.g., DX-2930). For example, this combination may result in lower doses of the inhibitor being required, thereby reducing side effects.

[0139] The antibodies described herein (e.g., DX-2930) may be administered in combination with one or more current therapies for treating HAE. For example, the DX-2930 antibody may be used simultaneously with a second anti-HAE therapeutic agent, such as ecallantide, a C1 esterase inhibitor (e.g., CINRYZE™), aprotinin (TRASYLOL®), and / or a bradykinin B2 receptor inhibitor (e.g., icatibant (Firazyr®)).

[0140] The term "combination" refers to the use of two or more agents or therapies to treat the same patient, where the use or action of the agents or therapies overlap in time. The agents or therapies can be administered simultaneously (e.g., as a single formulation administered to the patient or as two separate formulations administered simultaneously) or sequentially in any order. Sequential administrations are administrations provided at different times. The time between administration of one agent and administration of another agent can be minutes, hours, days, or weeks. The use of plasma kallikrein binding antibodies described herein can also be used to reduce the dose of another therapy, for example, to reduce side effects associated with another agent being administered. Thus, combinations can include administering the second agent at a dose at least 10, 20, 30, or 50% lower than would be used in the absence of the plasma kallikrein binding antibody. In some embodiments, a subject can be provided with C1 inhibitor as a loaded IV or SC administration at the same time as the first administration of an anti-pKal antibody (e.g., DX-2930) described herein. Thereafter, the subject may continue anti-pKal antibody treatment (without further administration of C1 inhibitor).

[0141] The combination therapy may include administering an agent that reduces the side effects of the other therapy. The agent may be an agent that reduces the side effects of treating a plasma kallikrein-associated disease.

[0142] (iv) Assays for evaluating treatment regimens Also within the scope of this disclosure are assay methods for assessing the efficacy of any of the treatment methods described herein. In some embodiments, plasma or serum concentrations of one or more biomarkers associated with HAE (e.g., 2-chain HMWK) may be measured before and / or during the course of treatment (e.g., after the first dose) to assess efficacy of the treatment. In some embodiments, plasma or serum concentrations (levels) of one or more biomarkers associated with HAE obtained at a time point after administration of a dose are compared to concentrations of the biomarkers in samples obtained at an earlier time point after administration of a dose or before the first dose. In some embodiments, the biomarker is 2-HMWK.

[0143] The level of the biomarker can be measured by detecting the biomarker in a plasma or serum sample obtained from a subject using an antibody that specifically detects the biomarker, for example, by immunoassay such as a Western blot assay or ELISA. In some embodiments, the level of 2-HWMK in a plasma or serum sample obtained from a subject is assessed by immunoassay. Antibodies used in immunoassays for the detection of 2-HWMK are used in the art, and the selection of such antibodies for use in the methods described herein will be apparent to one of skill in the art.

[0144] Without further elaboration, it is believed that a person skilled in the art can utilize the present invention to the extent that it is safe based on the above description. Therefore, the following specific embodiments are to be construed as merely illustrative, and not limiting in any way to the remainder of the disclosure. All publications mentioned herein are incorporated by reference for the purposes or subject matter mentioned herein.

[0145] Working Example Example 1: Comparison of indirect treatment with DX-2930 and intravenous C1 esterase inhibitor treatment for long-term prevention of attacks of hereditary angioedema Overview of the study A systematic literature review was performed to identify and summarize existing evidence for efficacy and safety of long-term preventive treatments in patients with type I and type II hereditary angioedema (HAE) for indirect treatment comparison (ITC). Additionally, data from the HELP trial (Banerji et al., JAMA 320(20):2108-21, 2018) were analyzed to establish optimal parametric survival models for use in ITC with evidence identified by the systematic literature review. All analyses were performed using R (R Core Team, 2018, www.R-project.org).

[0146] The HELP trial evaluated the efficacy and safety of subcutaneous lanadelumab (DX-2930) in preventing acute angioedema attacks in patients with type I and type II HAE and investigated three lanadelumab dosing regimens (300 mg every 2 weeks [q2w], 300 mg every 4 weeks [q4w], and 150 mg q4w) and compared them with placebo (Banerji et al., JAMA 320(20):2108-21, 2018).

[0147] material and method A systematic literature search up to June 2017 was performed using MEDLINE, Embase, MEDLINE In-Process, and Cochrane Library databases. A search of conference proceedings was performed in 2016 and 2017, including EAACI; American College of Allergy, Asthma and Immunology; World Allergy Congress; and European Society for Immunodeficiency. Further searches of key health technology assessment organizations included NICE (National Institute for Health and Care Excellence), Canadian Agency for Drugs and Technologies in Health Common Drug Review, Scottish Medicines Consortium, and All Wales Medicines Strategy Group. The search focused on trials for long-term prevention of angioedema attacks in patients aged 12 years or older with type I or type II HAE based on this criteria is listed in Table 1B.

[0148] [Table 2] HAE, hereditary angioedema; RCT, randomized controlled trial a Systematic reviews and meta-analyses of RCTs and non-RCTs were included and flagged for screening of bibliographies to determine whether the literature search missed any potentially relevant studies. b Publications in languages ​​other than English were included and investigated if sufficient evidence was not identified from publications written in English.

[0149] Development and reporting of this analysis adhered to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines (Moher et al., PLoS Med. 6(7):e1000097, 2009). Randomized controlled trials were considered the highest measure of evidence for ITC, and data from nonrandomized trials were considered when randomized controlled trial data were not available.

[0150] Abstracts and full-text articles were assessed for inclusion by two independent reviewers, and data extraction from selected articles was performed by one reviewer and independently verified by a second reviewer: any uncertainties were resolved by a third reviewer. Assessment of the descriptions of included randomized controlled trials was performed by two independent reviewers using comprehensive appraisal criteria based on recommendations in NICE's manufacturer evidence submission template (Single technology appraisal (STA): user guide for company evidence submission template. UK: National Institute for Health and Care Excellence; www.nice.org.uk / process / pmg24 / chapter / instructions-for-companies#quality-assessment-of-the-relevant-randomised-controlled-trials, 2015).

[0151] Parametric survival models Kaplan-Meier curves were derived for time to first HAE attack (defined as any investigator-confirmed attack) after day 0 and after day 70 for each lanadelumab dosing regimen and placebo using patient-level data from the HELP trial (Banerji et al., JAMA 320(20):2108-21, 2018). Univariate Cox proportional hazards models with treatment as the only covariate were fitted to estimate the relative treatment effect (i.e., HR) of each lanadelumab dose compared with placebo.

[0152] To extrapolate the long-term survival curves used in the NMA, standard parametric survival models (Exponential, Weibull, log-normal, log-logistic, Gompertz, and generalized gamma survival models) (Latimer, Decision Support Unit, National Institute for Health and Clinical Excellence; nicedsu.org.uk / wp-content / uploads / 2016 / 03 / NICE-DSU-TSD-Survival-analysis.updated-March-2013.v2.pdf, 2011) were fitted to the Kaplan-Meier data. When standard models did not fit the observed data, more flexible proportional hazards spline models were considered, which are an extension of the standard Weibull model that models the log cumulative hazard as a function of a natural cubic spline of log time (Royston et al., Stat Med. 21(15):2175-97, 2002). Methods used to assess model fit were the clinical relevance of the extrapolation, measures of goodness of fit, and visual inspection by overlaying the fitted survival curves with the observed survival data.

[0153] Attack rate and time to first HAE attack The outcomes of interest in this ITC were the rate of events (HAE attacks) and time to event (first HAE attack). Attack rate was defined as the number of attacks experienced in a 28-day cycle, where the relative treatment effect was estimated as a rate ratio (RR). Time to first attack was defined as the time that patients with HAE had their first attack after day 0 (the day of first preventive therapy) or after day 70 (the approximate day when steady-state plasma lanadelumab concentrations were achieved). For time to first attack, the relative treatment effect was estimated as a hazard ratio (HR).

[0154] A Bayesian NMA relying on Markov chain Monte Carlo methods was developed to assess the outcomes of attack rate and time to first attack. Relative efficacy was estimated using a treatment effect model to allow synthesis of direct and indirect evidence in one analysis while accounting for correlations arising from multiple arm trials (Dias et al., Decision Support Unit, National Institute for Health and Clinical Excellence; nicedsu.org.uk / wp-content / uploads / 2017 / 05 / TSD2-General-meta-analysis-corrected-2Sep2016v2.pdf, 2011). Some trials in the evidence network did not report the time to first attack. However, because the proportion of patients who did not experience HAE attacks was reported in all trials, an extension of the NMA method was used to synthesize the data of measures and time to event data in the same analysis (Woods et al., BMC Med Res Methodol. 10:54, 2010). Treatment effects were compared using confidence intervals (CrIs); in this case, the CrI for treatment vs. placebo did not include the value 1, and this result was considered statistically significant. Both HELP and CHANGE were randomized, double-blind studies in patients with confirmed HAE (Banerji et al., JAMA. 320(20):2108-21, 2018; Zuraw et al., N Engl J Med. 363(6):513-22, 2010). Fixed-effects and random-effects models (REM) were considered in the NMA, and the choice of model was based on the evaluation of study design, inclusion criteria, and patient characteristics.

[0155] Predicted survival curves Projected survival curves were derived separately for each treatment arm for time to first seizure after day 0 and after day 70 by combining spline survival curves fitted to the HR from the ITC and placebo data from the HELP trial. These curves were used to estimate the proportion of patients who were seizure-free 60 months after the start of preventive treatment (day 0 or day 70) and 95% confidence intervals (CI) for each comparator in the ITC.

[0156] result Systematic literature review A systematic literature review was performed. An overview of the identified records is presented in Figure 1. The search identified 1299 records, of which 52 (22 records from 7 randomized controlled trials [RCTs] and 30 records from 23 nonrandomized controlled trials [nRCTs]) were selected for detailed feasibility assessment that matched the eligibility criteria and involved lanadelumab results from the HELP trial and could be included in the ITC. Twenty-two records were excluded because their initial publication was a secondary publication already identified among the 52 records that matched the eligibility criteria. Of the remaining records for feasibility assessment, 20 records lacked real-world applicability and were therefore excluded as inappropriate comparison criteria. These include two RCTs (Gelfand et al., N Engl J Med. 295(26):1444-8, 1976; Sheffer et al., Ann Int Med. 86(3):306-8, 1977) and 15 nRCTs (Fust et al., Eur J Clin Invest. 41(3):256-62, 2011; Agostoni et al., Medicine (Baltimore).71(4):206-15, 1992; Bork et al., Ann Allergy Asthma Immunol. 100(2):153-61, 2008; Caminoa et al., Allergy. 68(suppl 97):61, 2013; Cicardi et al., J Allergy Clin Immunol. 99(2):194-6, 1997; Davis et al., Johns Hopkins Med J. 135(6):391-8, 1974; Farkas et al., J Oral Maxillofac Surg. 57(4):404-8, 1999; Kreuz et al., Transfusion.49(9):1987-95, 2009; Obtulowicz et al., Int Rev Allergol Clin Immunol. 3(3):163-6, 1997; Ott et al., Clin Endocrinol (Oxf).66(2):180-4, 2007;Psarros et al., Int Arch Allergy Immunol. 164(4):326-32, 2014;Rosen et al., Birth Defects: Orig Artic Ser. 16(1):499-507; 1980;Steiner et al., Orphanet J Rare Dis. 11:43; 2016;Winnewisser et al., J Int Med. 241(1):39-46, 1997;Zotter et al., Orphanet J Rare Dis. 9:205, 2014), as well as one RCT describing the use of SC C1-INH / recombinant human hyaluronidase combination therapy in patients with HAE (Riedl et al., Allergy Asthma Proc. 37(6):489-500, 2016) were included because these interventions are not recommended by the WAO / EAACI guidelines for the maintenance of HAE as long-term prevention (Maurer et al., Allergy. 73(8):1575-96, 2018); two RCTs describing the use of SC C1-INH (Longhurst et al., N Engl J Med. 376(12):1131-40, 2017; Zuraw et al., Allergy. 70(10):1319-28, 2015) were excluded because this product is not commercially available outside the United States and Canada (CSL Behring. Global product list; www.cslbehring.com / products / global-products-list; retrieved 2019 Feb. 20), limiting the real-world applicability of all comparisons.

[0157] The one reported record comes from a phase 3 trial of IV C1-INH (Zuraw et al., N Engl J Med. 363(6):513-22, 2010), which was deemed appropriate for ITC with lanadelumab results from the HELP trial. Because this RCT was relevant for ITC and because nRCTs were only considered if no RCTs were identified for a given intervention, four nRCTs for this intervention (Bernstein et al., J Allergy Clin Immunol Pract. 2(1):77-84 2014; Aygoren-Pursun et al., J Allergy Clin Immunol. 137(2):AB251, 2016; Rasmussen et al., Ann Allergy Asthma Immunol. 116(5):476-7, 2016; Zuraw et al., Am J Med. 125(9):938.e1-.e7, 2012) were subsequently excluded. The remaining five studies (Banerji et al., N Engl J Med. 376(8):717-28, 2017; Bork et al., Oral Surg Oral Med Oral Pathol Oral Radiol Endod. 112(1):58-64, 2011; Bouillet et al., Allergy. 72(Suppl 103):593, 2017; Farkas et al., Allergy. 67(12):1586-93, 2012; Levi et al., J Allergy Clin Immunol. 117(4):904-8, 2006) were excluded due to differences and / or lack of clarity in study design, study population, treatment intervention, endpoints, and / or patient characteristics (Table 2). A network diagram of the two studies included in the ITC is shown in Figure 2. The design of each study is summarized in Table 3 , and the baseline demographic characteristics and primary endpoint results for both studies are presented in Table 4 .

[0158] [Table 3] TIFF2025061116000011.tif38161C1-INH, C1 esterase inhibitor; IV, intravenous; PD, pharmacodynamics; PK, pharmacokinetics; q2w, every 2 weeks; SC, subcutaneous a It does not include feasibility assessment of records that were included in the ITC (n = 1) or excluded as secondary publications (n ​​= 22), inappropriate comparison criteria (n = 20), or unnecessary nRCTs (n = 4). b No detailed dosing information was reported.

[0159] [Table 4] C1-INH, C1 esterase inhibitor; HAE, hereditary angioedema a The CHANGE trial included two parts: Part A, which evaluated the use of IV C1-INH in acute attacks, and Part B, which evaluated its use in prevention. Only Part B is summarized here, which was a crossover study consisting of two 12-week treatment periods in which patients were randomly assigned to receive either IV C1-INH or placebo during the first treatment period, followed by crossover in the second treatment period to treatment not administered during the first treatment period (Zuraw et al., N Engl J Med. 363(6):513-22, 2010). b Banerji et al., JAMA 320(20):2108-21, 2018

[0160] [Table 5] C1-INH, C1 esterase inhibitor; CI, confidence interval; HAE, hereditary angioedema; IV, intravenous; LS, least squares; NA, not applicable; q2w, every 2 weeks; q4w, every 4 weeks; SD, standard deviation aA bout was defined as a discrete episode during which a subject progressed from no angioedema to symptoms of angioedema; a bout progressing from one site to another, or a bout that began to regress and then worsen before full recovery, was considered a single bout (Zuraw et al., N Engl J Med. 363(6):513-22, 2010). b Banerji et al., JAMA 320(20):2108-21, 2018 *P<0.001 vs placebo.

[0161] Time to first seizure (HELP study) As shown in the HELP trial, the time to first HAE attack increased after each time point for all three lanadelumab dosing regimens compared with placebo, corresponding to a reduced risk of experiencing a first attack after day 0 and after day 70 since treatment began. After day 0, the median number of days to first attack was 59 (95% CI: 28-not estimable [NE]) for patients receiving lanadelumab 300 mg q2w; 28 (95% CI: 10-101) for lanadelumab 300 mg q4w; and 26 (95% CI: 11-NE) for lanadelumab 150 mg q4w, compared with 8 (95% CI: 6-18) for patients receiving placebo. After day 70, more than 50% of patients receiving lanadelumab 300 mg q2w and 150 mg q4w were seizure-free by the end of the 6-month treatment period; thus, the median number of days to first seizure was NE. The median time to first seizure after day 70 in patients receiving lanadelumab 300 mg q4w was 61 days (95% CI: 25-NE) compared with 12 days (95% CI: 6-16) in patients receiving placebo.

[0162] As observed in the HELP study, 2.4% of patients receiving placebo were seizure-free from day 0 to the end of the treatment period compared with 44%, 31%, and 39% of patients receiving lanadelumab 300 mg q2w, 300 mg q4w, and lanadelumab 150 mg q4w, respectively (Table 5). Similar or greater percentages of seizure-free patients were observed with lanadelumab treatment from day 70 to the end of the treatment period. The proportion of lanadelumab-treated patients who were seizure-free was also derived from Kaplan-Meier estimates of time to first seizure after treatment to validate the Kaplan-Meier survival analysis. These estimated percentages of seizure-free patients were generally consistent with those observed from day 0 and day 70 to the end of the 6-month treatment period (Table 5).

[0163] [Table 6] CI, confidence interval; q2w, every 2 weeks; q4w, every 4 weeks a Kaplan-Meier estimates of time to first seizure after treatment were used to derive the proportion of patients who remained seizure-free. b Excluding patients who discontinued before day 70: lanadelumab 300 mg q2w, n=26; lanadelumab 300 mg q4w, n=29; lanadelumab 150 mg q4w, n=28; placebo, n=37.

[0164] Selection of a model for the time-to-first-seizure endpoint Various parametric survival models were applied to the data on time to first seizure after day 0 and after day 70. Of six standard parametric models, the Akaike Information Criterion (AIC) values ​​indicated that the Gompertz model best fit the Kaplan-Meier data for the study period (i.e., after day 0 and after day 70). The Gompertz model predicted that patients who were seizure-free from day 0 to the end of the study would remain seizure-free indefinitely after the study ended. However, based on the possibility of pharmacological drug resistance and loss of response, where the therapeutic effect may wane with prolonged exposure, to put it conservatively (Roda et al., Clin Transl Gastroenterol. 7:e135, 2016; Salva Lacombe et al., Drugs. 51(4):552-70, 1996), the Gompertz model was deemed clinically implausible.

[0165] Thus, because the standard distribution for the parametric survival analysis did not fit the observed data well, a more flexible proportional hazards spline parametric model was fitted to the data. A spline model with one internal knot (k=1) was found to fit the data best, which was confirmed based on visual comparison with the Kaplan-Meier data and the clinical relevance of the reduction over time in the relative effect of each lanadelumab dose compared to placebo. The addition of additional internal knots (k=2-5) to the spline model did not result in a corresponding improvement in model fit.

[0166] Indirect treatment comparison via network meta-analysis A fixed-effects model was chosen for the NMA because no systematic differences were identified between the patient populations of the HELP and CHANGE trials, which were similar in age, sex, and percentage of patients with type I or type II HAE (Tables 3 and 4 above). Thus, we assumed that each study estimated the same treatment effect and that any variation between studies was due to sampling variation only.

[0167] Attack rate RRs showed a significant reduction in attack rate in patients receiving lanadelumab (all dosing regimens) or IV C1-INH compared with placebo (Figure 3). Reductions in attack rate (per 4-week cycle) were 87%, 73%, and 76% (corresponding to median RRs [95% CrI] of 0.13 [0.07-0.24], 0.27 [0.18-0.40], and 0.24 [0.15-0.39]) in the lanadelumab 300 mg q2w, 300 mg q4w, and 150 mg q4w treatment arms, respectively, and 51% (corresponding to a median RR [95% CrI] of 0.49 [0.40-0.60]) for IV C1-INH compared with placebo.

[0168] Time to first seizure When comparing treatment with placebo, a significantly lower risk of first seizure after day 0 was observed for all lanadelumab dosing regimens (median HR [95% CrI]: 300 mg q2w, 0.27 [0.13-0.55]; 300 mg q4w, 0.39 [0.21-0.74]; 150 mg q4w, 0.34 [0.17-0.66]), corresponding to a 61% to 73% reduction in the risk of first seizure after day 0 (Figure 4A). Results regarding the risk of first seizure after day 70 were similar (median HR [95% CrI]: 300 mg q2w, 0.09 [0.04-0.22]; 300 mg q4w, 0.27 [0.14-0.53]; 150 mg q4w, 0.20 [0.10-0.41]), corresponding to a reduction of 73% to 91% (Figure 4B). In contrast, reduced risk of first seizure with IV C1-INH treatment was observed compared with placebo, although these reductions were not statistically significant (median HR [95% CrI]: day 0, 0.54 [0.23-1.19]; day 70, 0.53 [0.23-1.19]).

[0169] 60-month predicted survival curve Projected survival curves for 60 months based on time to first seizure were derived for each treatment arm after day 0 and after day 70 by combining spline survival curves fitted to the HR from the ITC and placebo data from the HELP study (Figures 5A-5B). The highest predicted proportion of seizure-free patients was observed with lanadelumab 300 mg q2w across both time points. When the time to first seizure data after day 0 were extrapolated outside the 60-month range, the predicted percentages of seizure-free patients were 26%, 6.5%, and 0.6% for treatment with lanadelumab 300 mg q2w, IV C1-INH, and placebo, respectively. When the time-to-first-seizure data were extrapolated outside the 60-month range, the predicted percentages of seizure-free patients were 46%, 0.9%, and 0% after day 70 for treatment with lanadelumab 300 mg q2w, IV C1-INH, and placebo, respectively.

[0170] A proportional hazards spline model with one internal knot (k=1) was found to best fit the observed time to first attack data from the HELP trial. These data showed that the key benefit of administering lanadelumab compared to placebo in preventing HAE attacks was the increased attack-free interval. The highest predicted proportion of attack-free patients was observed with lanadelumab 300 mg q2w across all time points, and the lowest predicted proportion of attack-free patients was observed with IV C1-INH and placebo. Based on a half-life of lanadelumab of approximately 14 days (range 13.8-15.0 days), the time to steady-state concentrations of lanadelumab was estimated to be approximately 70 days. Thus, after day 70, when lanadelumab reached steady-state concentrations, more than 50% of patients in the lanadelumab 300 mg q2w treatment arm were seizure-free for the remainder of the 6-month study period, and the median number of days to first seizure after day 70 in the 300 mg q4w arm was 61 compared with 12 in the placebo arm. Based on the projected survival curves, the proportion of seizure-free patients after the first 6 months of treatment with lanadelumab 300 mg q2w increased as the time to first seizure (45% and 72%) was assessed later (after day 0 and after day 70, respectively); a similar pattern was observed in the lanadelumab 300 mg q4w (31% and 37%, respectively) and 150 mg q4w (36% and 47%, respectively) treatment arms, whereas patients receiving C1-INH showed the opposite pattern of decline (20% and 13%, respectively) at subsequent evaluations. These findings generally remained consistent when the data were extrapolated beyond the 60-month period.

[0171] Thus, HAE patients who have been treated with lanadelumab for a period of time (e.g., 6 months) and have no HAE attacks may be administered a reduced dose of lanadelumab, e.g., from 300 mg / 2 weeks to 300 mg / 4 weeks.

[0172] Example 2: Indirect Treatment Comparison of DX-2930 and Intravenous C1 Esterase Inhibitor Treatment Using Individual Patient Data from the HELP-03 and CHANGE Trials the purpose The objective was to perform an indirect treatment comparison (ITC) of lanadelumab (300 mg subcutaneous every 2 weeks (300 mg SC q2w), 300 mg subcutaneous every 4 weeks (300 mg SC q 2w)) and Cinryze® (C1-INH) 1000 units intravenously (1000U IV) using individual patient data (IPD) from the HELP-03 (HELP) and CHANGE trials. The use of IPD instead of pooled data is preferred for several reasons. First, IPD provides more information, reducing heterogeneity throughout the network and resolving potential inconsistencies. In addition, IPD facilitates estimation of subgroup effects, aiding convergence and generally resulting in more accurate estimates. Furthermore, IPD allows for the distinction of within-study associations from across-study associations. Furthermore, using IPD from only a few studies may also reduce ecological bias, which is often a problem when evidence is sparse and sample sizes are small.

[0173] Desired outcome Both the HELP and CHANGE trials referred to the number of HAE attacks per 28 days. A previous study by BresMed reported the time to first HAE attack after treatment on days 0 and 70 in the HELP trial. This outcome measure was estimated using the IPD reported in the CHANGE trial. Only the HELP trial disclosed the proportion of patients experiencing a 50% or greater reduction in the number of attacks. The CHANGE trial did not report this outcome measure and was not considered relevant for the ITC.

[0174] conclusion Overall, the assumption of similarity was not deemed to hold between the HELP and CHANGE studies with respect to baseline HAE attack rates per 28 days. This was confirmed by expert clinician opinion. Other covariates of interest (e.g., age, sex, and weight) also differed between the studies, but these were deemed to have less impact on the results. Thus, it was decided that this base case analysis would consider HAE attack rates per 28 days at baseline, but the regression models for the sensitivity analysis would include other important covariates that could potentially modify the treatment effect. Any differences between this base case and sensitivity analysis findings may indicate that the base case results are not robust and may possibly be driven by study differences. In this case, a second sensitivity analysis is planned to assess whether the assessment of attacks per patient or investigator influences the attack rate per 28 days results. To account for this, the analysis will include only patient-reported HAE attacks in both studies.

[0175] A common standard of comparison As shown in the evidence network in Figure 6, the common comparison standard for Cinryze® (C1-INH) and lanadelumab in the HELP and CHANGE studies was placebo. The HELP study compared three different lanadelumab dosing regimens with placebo, and the CHANGE study was a crossover study comparing Cinryze® with placebo. Excluding the 150 mg dose of lanadelumab (not approved for this indication) reduced the number of comparisons and avoided multiplicity problems (e.g., testing multiple hypotheses). When multiple hypotheses are tested simultaneously, the alpha error increases, and therefore the significance level requires adjustment (e.g., Bonferroni). It can be difficult to identify statistically significant differences at a significance level of less than 5%. Thus, it is recommended to avoid testing multiple hypotheses.

[0176] method Statistical methods for proving hypotheses can be characterized first as frequentist or Bayesian. Frequentist methods, such as adjusted indirect comparisons (e.g. Bucher method, matched-adjusted indirect comparison (MAIC), and simulated treatment comparison (STC)), allow the indirect comparison of two interventions in a single step. The term network meta-analysis (NMA) stands for the simultaneous comparison of a larger network of interventions. In Bayesian methods, the minimal informative priors are usually the assigned treatment effects.

[0177] For this ITC, a frequentist approach was preferred over a Bayesian one due to the limited evidence base (N = 2 studies). Currently, there are various frequentist approaches to include IPD in statistical models for evidence synthesis. These methods differ mainly based on the criteria for inclusion or exclusion of arms ("anchors") of a common comparator.

[0178] As a second step, the Bucher method is often used to combine the relative treatment effects in fixed comparisons to obtain the final ITC result. Only two treatment interventions can be compared at one time. If multiple comparisons are of interest, multiple analyses must be performed. Corresponding point estimates are expected to be very similar to those obtained via Bayesian analysis. However, the 95% confidence intervals are usually not as wide as the confidence intervals of the Bayesian analysis.

[0179] With detailed feasibility evaluation, the ITC of the present invention was performed using a frequentist-based fixed approach. Importantly, this approach preserved randomization. In the base case analysis, the regression model included the HAE attack rate per 28 days at baseline. Other covariates of interest (e.g., age, sex, and weight) were added in a sensitivity analysis to explore the overall impact of these variables on the outcome. A second sensitivity analysis evaluated the impact of patient-reported HAE attacks that were not confirmed by the investigator on the base case outcome.

[0180] Poisson regression - HAE attack rate per 28 days Poisson regression analyses were performed to estimate rate ratios (RRs) for HAE attack rates per 28 days. Poisson regression requires that the following assumptions be met: (1) observations are independent, (2) measurements follow a Poisson distribution, and (3) the conditional means and variances of the models are identical.

[0181] Given that the outcome of interest (rate of HAE attacks per 28 days) was expressed as a measure of data, Poisson regression was deemed appropriate, and the number of attacks per treatment period was taken into account in the model. According to Banjeri et al., the model included an offset variable to adjust for differences in follow-up periods. The offset was defined as the logarithm of the number of days that a patient was observed during the treatment period.

[0182] The measure of treatment effect (input of the model) was expressed as the estimated RR of attacks per 28 days. For the lanadelumab regimens (300 mg q2w and 300 mg q4w), this measure was expressed as the ratio of attacks per 28 days during each treatment compared to treatment with placebo. For Cinryze® (C1-INH), the RR represented the ratio of attacks per 28 days during treatment with Cinryze® compared to treatment with placebo. The estimated RRs generated by the Poisson model were then entered into the ITC.

[0183] Nonparametric Estimation and Cox Regression For the outcome "time to first seizure," Cox regression analysis was performed to estimate the hazard ratio (HR) of first seizure for each lanadelumab regimen (300 mg q2w and 300 mg q4w) compared to placebo, and the HR of first seizure for Cinryze® compared to placebo. The estimated HRs were then entered (via Bucher's method) into an ITC to compare lanadelumab dosing regimens and Cinryze®, as described. Cox regression is semiparametric and therefore does not specify any requirements regarding the shape of the baseline hazard, but the following assumptions must be met: (1) proportional hazards, (2) linear relationship between covariates, and (3) independent observations.

[0184] Kaplan-Meier curves were derived via nonparametric estimation to model the outcomes of "time to first attack after day 0 and after day 70." Patients were censored at the date of their last data capture. This graph showed that the hazard function was proportional over time, and therefore no statistical measures were necessary to account for nonproportional hazards.

[0185] In the base case, the models were adjusted for the number of HAE attacks per 28 days at baseline. In a sensitivity analysis, the models also included relevant covariates (age, sex, and weight). Due to the small sample size of the CHANGE study, formal variable selection was not performed. Estimates obtained from these adjusted models were compared with unadjusted estimates from models excluding the covariates.

[0186] Because the CHANGE study was a crossover study, the individuals observed twice (under placebo and Cinryze treatment) were not independent. As described, a mixed model including fixed effects for treatment, period, and sequence, and a random effect for study subject was used to account for repeated measurements in the study.

[0187] result 1. HAE attack rate per 28 days Table 6 shows that the baseline attack rates for placebo and Cinryze treatment were the same. The CHANGE crossover study did not include a run-in period. During the treatment period, the HAE attack rate per 28 days for placebo was higher compared to the baseline attack rate for Cinryze treatment. The mean number of attacks per treatment period was significantly lower for Cinryze than for placebo. The treatment period was comparable between the two interventions. The time to first attack after 0 and 70 days of treatment for placebo was shorter than for Cinryze treatment.

[0188] [Table 7]

[0189] [Table 8] TIFF2025061116000017.tif57162

[0190] As shown in Table 7, both lanadelumab 300 mg q2w and 300 mg q4w reduced the mean number of seizures per treatment period compared to placebo. The mean baseline seizure rates were comparable between the three study arms. The placebo arm had the shortest treatment period, and the time to first seizure with lanadelumab treatment (300 mg q2w and 300 mg q4w) was longer compared to placebo.

[0191] Rate of HAE attacks per 28 days - Poisson regression Furthermore, the estimated mean HAE attack rate per 28 days was 1.99 (95% CI: 1.56; 2.52) for Cinryze® treatment and 4.03 (95% CI: 3.31; 4.91) for placebo. The corresponding RR (0.49) indicated that patients treated with Cinryze® had 0.49 times the rate of HAE attacks compared to patients treated with placebo. This difference was statistically significant (as the corresponding 95% CI does not include 1). The resulting percent change in the mean attack rate for patients treated with Cinryze® compared to placebo was -51%.

[0192] [Table 9]

[0193] As shown in Table 9, the estimated mean rates of HAE attacks per 28 days were 0.26 (95% CI: 0.15; 0.45) for lanadelumab 300 mg q2w, 0.54 (95% CI: 0.37, 0.77) for lanadelumab q2w, and 2.0 (95% CI: 1.69; 2.38) for placebo treatment. The corresponding RRs indicated that patients treated with lanadelumab 300 mg q2w and lanadelumab q4w had 0.13- and 0.27-fold higher rates of HAE attacks, respectively, compared with patients treated with placebo. These estimates were statistically significant as the corresponding 95% CIs did not include the value "1." The resulting percentage change in mean seizure rate compared to placebo was -87% in patients treated with lanadelumab 300 mg q2w and -73% in patients treated with lanadelumab 300 mg q2w.

[0194] [Table 10]

[0195] Indirect comparisons using the Bucher method (Table 10) showed that patients treated with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w had 0.27-fold and 0.54-fold higher rates of HAE attacks compared to patients treated with Cinryze®. These results were statistically significant with the corresponding 95% CIs not including the value "1". The resulting percent changes in mean attack rates compared to Cinryze® were -73% and -46% for patients treated with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w, respectively. The ITC-generated estimates and confidence / confidence intervals for this IPD appeared to be consistent with the Bayesian NMA findings.

[0196] [Table 11]

[0197] Sensitivity analysis considering patient-reported HAE attacks in the HELP study As shown in Table 11, the estimated mean rates of HAE attacks per 28 days were 0.27 (CI: 0.16; 0.46) for lanadelumab 300 mg q2w, 0.55 (CI: 0.38; 0.78) for lanadelumab 300 mg q4w, and 2.02 (95% CI: 1.71; 2.39) for placebo. The corresponding RRs indicated that patients treated with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w had 0.13- and 0.27-fold higher rates of HAE attacks compared with patients treated with placebo, respectively. These estimates were statistically significant because the corresponding 95% CIs did not include 1. The percent change in mean seizure rate compared to placebo was -87% and -73% in patients treated with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w, respectively.

[0198] [Table 12]

[0199] In the base case, indirect comparison via Bucher's method showed that patients treated with lanadelumab 300 mg q2w had a 0.27-fold rate of HAE attacks compared to patients treated with Cinryze® (Table 12), and patients treated with lanadelumab 300 mg q4w experienced a 0.55-fold rate of HAE attacks compared to patients treated with Cinryze®. These estimates were statistically significant because the corresponding 95% CIs did not include 1. The percent change in mean attack rate compared to Cinryze® was −73% and −45% for treatment with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w, respectively. Comparable sensitivity analyses were not performed with Bayesian NMA.

[0200] [Table 13]

[0201] Sensitivity analysis considering all covariates (normalized baseline seizure rate, age, sex, weight) As shown in Table 13, the estimated mean rate of HAE attacks per 28 days was 2.21 (95% CI: 1.43; 3.41) for Cinryze® and 4.37 (95% CI: 2.89; 6.60) for placebo. The corresponding RR of 0.51 indicated that patients treated with Cinryze® had a mean rate of HAE attacks that was 0.51 times higher than patients treated with placebo. This result was statistically significant because the corresponding 95% CI did not include 1. The resulting percent change in mean attack rate with Cinryze® treatment compared to placebo was -49%.

[0202] [Table 14]

[0203] As shown in Table 14, the estimated mean rates of HAE attacks per 28 days were 0.27 (95% CI; 0.16; 0.46) for lanadelumab 300 mg q2w, 0.54 (95% CI; 0.37; 0.78) for lanadelumab 300 mg q4w, and 2.00 (95% CI; 1.63; 2.45) for placebo. The corresponding RRs showed that patients treated with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w had 0.14- and 0.27-fold higher rates of HAE attacks compared with patients receiving placebo, respectively. These results were statistically significant because the corresponding 95% CIs did not include 1. The resulting percentage changes in mean seizure rate compared to placebo were -86% and -73% for lanadelumab 300 mg q2w and lanadelumab 300 mg q4w, respectively.

[0204] [Table 15]

[0205] In the base case, indirect comparison via Bucher's method showed that patients treated with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w experienced 0.27-fold and 0.53-fold rates of HAE attacks, respectively, compared to patients treated with Cinryze® (Table 15). These results were statistically significant with the corresponding 95% CIs not including the value "1". The resulting percent changes in mean attack rates compared to Cinryze® were -73% and -47% for patients treated with lanadelumab 300 mg q2w and lanadelumab 300 mg q4w, respectively. Bayesian NMA results were not available for this sensitivity analysis.

[0206] [Table 16]

[0207] This section summarizes the ITC results of lanadelumab (300 mg q2w and 300 mg q4w) and Cinryze®. Base case and two sensitivity analyses demonstrated statistically significant differences in HAE attack rates per 28 days between lanadelumab (300 mg q2w and 300 mg q4w) and Cinryze®.

[0208] Two independent Poisson regression models generated RR estimates identifying the relative treatment effects of lanadelumab (300 mg q2w and 300 mg q4w) vs placebo and Cinryze® vs placebo. The base case considered the normalized HAE attack rate per 28 days at baseline, and a mixed model was used to account for the crossover design of the CHANGE trial. The corresponding results were applied as input to the ITC using the Bucher method.

[0209] The first sensitivity analysis included all HAE attacks reported in the HELP study, regardless of whether the attacks were confirmed by the investigator. The regression model for the second sensitivity analysis initially included age, sex, and weight. The two sensitivity analyses yielded results consistent with the base case and did not change the conclusions. These findings were also confirmed by the lack of statistically significant effects for covariates (apart from normalized baseline attack rate) on the outcome.

[0210] 2.Comparability of placebo effects in the CHANGE and HELP studies The power of non-parametric estimation showed that the mean time (in days) to first seizure after 0 days of placebo was significantly shorter in the CHANGE trial than in the HELP trial (4.773 days [SE 1.673] vs. 19.634 days [SE 4.212]). In Figure 7, the probability of being "free from first seizure" after 0 days of treatment is plotted on the y-axis and the observation period is plotted on the x-axis. The placebo treatments in the CHANGE and HELP trials were not equally effective, with patients in the CHANGE trial experiencing their first seizure significantly sooner than patients in the HELP trial, and the corresponding observation period being significantly shorter.

[0211] The output of non-parametric estimation showed that the mean time (in days) to first seizure after 70 days of placebo was significantly shorter in the CHANGE trial than in the HELP trial (3.143 days [SE 0.563] vs. 21.378 days [SE 4.687]). In Figure 8, the probability of being "free from a first seizure" after 70 days of treatment is plotted on the y-axis and the observation period is plotted on the x-axis. The placebo treatments in the CHANGE and HELP trials were not equally effective, with patients in the CHANGE trial experiencing their first seizure significantly sooner than patients in the HELP trial, and the corresponding observation period being significantly shorter.

[0212] This section summarizes the findings of the analysis comparing the placebo effect between the CHANGE and HELP trials. The placebo effect in terms of time to first seizure after 0 days of treatment and time to first seizure after 70 days of treatment was not comparable between the two trials. This was assessed via Kaplan-Meier graphs, various statistical tests (log-rank, Wilcoxon, likelihood ratio tests), and Cox regression models that included baseline seizures, age, sex, and weight normalized as covariates and accounted for differences in study design. There was no placebo effect in the CHANGE trial, and individuals experienced few seizures at baseline and during subsequent placebo treatment. A placebo effect was observed in the HELP trial.

[0213] In a randomized controlled trial, patients are randomly assigned to treatment arms to ensure comparability of relevant covariates between groups. ITCs are performed in the event of a lack of direct treatment comparisons. The assumption of similarity between studies must be held to ensure that the results of the ITC are valid. Thus, covariates of interest must be comparable between corresponding study arms. This is usually established during the process of feasibility assessment.

[0214] Placebo effects are often not comparable between the studies included in the ITC. However, it is not easy to evaluate the meaning of these differences. In the ITC of the present invention, the differences between placebo effects could favor either Cinryze® or lanadelumab. The absence of placebo effects in the CHANGE study implied that patients during C1-INH treatment could only benefit from the effect of the effective treatment without additional placebo effects. This is expected to occur even if Cinryze® is not effective. In contrast, placebo effects were observed in the HELP study. Thus, patients treated with lanadelumab may have benefited from the efficacy of the effective treatment and additional placebo effects. This difference may favor lanadelumab over Cinryze® in the analysis of the present invention.

[0215] For linear regression of the change from baseline in HAE attack rate per 28 days, the outcome measure of interest is the rate difference. The rate difference in change from baseline compared to placebo was higher in the CHANGE study than in the HELP study because no placebo effect was observed in the CHANGE study, resulting in ITC results in favor of Cinryze®.

[0216] However, in Poisson regression, the relative treatment effect is defined as the RR at follow-up, and baseline seizure rate is included as a covariate. Compared to linear regression, this analysis uses different outcome measures. As a result, the differences regarding the placebo effect between the two studies have little impact on the analysis, indicating a high probability of a statistically significant difference in favor of lanadelumab over Cinryze®.

[0217] For times to first seizure after 0 and 70 days of treatment, seizures occurred more frequently and significantly sooner during placebo treatment in the CHANGE trial than in the HELP trial. As discussed, the meaning of these findings is not easy to assess.

[0218] 3. Time to first seizure – nonparametric estimation and Cox regression Nonparametric estimates showed that the mean time to first seizure (in days) after 0 days of placebo treatment in the CHANGE trial was significantly shorter than in the HELP trial (4.773 days [SE 1.673] vs. 19.634 days [SE 4.212]).

[0219] As shown in Figure 9, the probability of being "free from first seizure" after 0 days of treatment is plotted on the y-axis and the observation period is plotted on the x-axis. Placebo was not more effective than Cinryze treatment in the CHANGE study. Patients who received placebo experienced their first seizure significantly sooner than those during Cinryze treatment. If the two curves do not cross, the proportional hazards assumption can be considered to hold. This confirms that Cox regression is a valid method to estimate hazard ratios (HRs) for ITC.

[0220] As shown in Figure 10, the probability of being "free from first attack" after 0 days of treatment is plotted on the y-axis and the observation period is plotted on the x-axis. In the HELP study, placebo was not more effective than the two lanadelumab dosing regimens. Patients who received placebo experienced their first attack significantly sooner than patients during lanadelumab treatment. If the curves do not cross, the proportional hazards assumption can be considered to hold. This confirms that Cox regression is a valid method to estimate hazard ratios for ITC.

[0221] Different analytical methods used may contribute to the contradictory findings. The ITC of the present invention used a frequentist Cox regression model that considered repeated measures (including fixed effects of treatment period and sequence and random effects of test subjects) and normalized the baselined HAE attack rate. Previous NMA using Bayesian methods did not take these measures and only reported the results of the random effects model used for comparison.

[0222] Comparing lanadelumab 300 mg q2w and lanadelumab 300 mg q4w with placebo, Cox regression produced HRs of 0.27 (95% CI: 0.15; 0.50) and 0.38 (95% CI: 0.22; 0.67), respectively (Table 16). Bayesian NMA produced comparable HRs of 0.27 with corresponding 95% confidence intervals (Crl) of (0.13; 0.55) and 0.39 with 95% Crl of (0.21; 0.74), respectively.

[0223] [Table 17]

[0224] [Table 18]

[0225] Indirect comparison via Bucher's method showed that individuals treated with lanadelumab 300 mg q2w had a 0.73-fold hazard of first HAE attack after treatment day 0 compared to individuals treated with Cinryze® (Table 18). Treatment with lanadelumab 300 mg q4w increased the hazard of attacks by 1.05-fold compared to Cinryze®. This finding was not statistically significant, as evidenced by the inclusion of the value "1" in the corresponding 95% CI. The estimates made with ITC using IPD differed from the results of Bayesian NMA, mainly due to the different methodology applied (as described above). The conclusions did not change, as the boundaries of the intervals appeared similar.

[0226] [Table 19]

[0227] When comparing Cinryze® and placebo, Cox regression produced a HR of 0.37 (95% CI; 0.18; 0.76) (Table 19). These results were consistent with the base case.

[0228] [Table 20]

[0229] When comparing lanadelumab 300 mg q2w and lanadelumab 300 mg q4w with placebo, Cox regression produced HRs of 0.27 (95% CI: 0.14; 0.51) and 0.37 (95% CI: 0.21; 0.66), respectively (Table 20). These estimates were consistent with previous findings. The inclusion of additional covariates did not affect the primary findings.

[0230] [Table 21]

[0231] Indirect comparison via the Bucher method estimated that patients treated with lanadelumab 300 mg q2w had a 0.72-fold hazard of first HAE attack after treatment on day 0 compared with patients treated with Cinryze treatment (Table 21). Lanadelumab 300 mg q4w treatment resulted in approximately the same hazard of HAE attack when compared with Cinryze. This finding was not statistically significant, as evidenced by the inclusion of the value "1" in the corresponding 95% CI. This result was consistent with the base case.

[0232] [Table 22]

[0233] Nonparametric estimates showed that the mean time to first seizure (in days) after 70 days of placebo treatment in the CHANGE trial was significantly shorter than in the HELP trial (3.1429 days [SE 0.5625] vs. 21.378 days [SE 4.687]).

[0234] The probability of being "free from first seizure" after 70 days of treatment is plotted on the y-axis of the graph in Figure 11, and the observation period is plotted on the x-axis. In the CHANGE study, placebo was not more effective than Cinryze treatment. Patients treated with placebo experienced their first seizure significantly sooner than during treatment with Cinryze. If the two curves do not cross, it can be concluded that Cox regression is a valid method for estimating the HR for ITC.

[0235] As shown in Figure 12, the probability of being "free from the first attack" after 70 days of treatment is plotted on the y-axis and the observation period is plotted on the x-axis. Placebo treatment was not more effective than lanadelumab treatment (300 mg q2w and 300 mg q4w) in the HELP study. Patients treated with placebo experienced their first attack significantly sooner than patients treated with lanadelumab. If the curves do not cross, it can be concluded that Cox regression is a valid method for estimating the HR for ITC.

[0236] When comparing Cinryze® and placebo, Cox regression yielded a HR of 0.34 (95% CI: 0.16; 0.72). In Bayesian NMA, the corresponding HR was 0.53 (0.00; 318.88) (Table 22).

[0237] [Table 23]

[0238] When comparing lanadelumab 300 mg q2w and lanadelumab 300 mg q4w with placebo, Cox regression yielded HRs of 0.07 (95% CI; 0.03; 0.16) and 0.21 (95% CI; 0.11; 0.40), respectively (Table 23). In Bayesian NMA, the corresponding HRs were 0.09 (95% CI; 0.04; 0.22) and 0.27 (95% CI; 0.14; 0.53), respectively.

[0239] [Table 24]

[0240] Indirect comparison via Bucher's method showed that patients treated with lanadelumab 300 mg q2w had a 0.19-fold hazard of a first HAE attack after 70 days of treatment compared to patients treated with Cinryze® (Table 24). These results were statistically significant by not including the value "1" in the corresponding 95% CI (95% CI: 0.0584; 0.6200). The hazard of an HAE attack during treatment with lanadelumab 300 mg q4w was 0.62-fold the hazard of an attack during Cinryze® treatment. These results comparing treatment with lanadelumab 300 mg q4w to treatment with Cinryze® were not statistically significant (95% CI: 0.2336, 1.6381), as evidenced by the inclusion of "1" in the corresponding 95% CI. The estimates produced by the ITC using IPD differed from the results of the Bayesian NMA, but the interval boundaries appeared similar and so the conclusions remained unchanged.

[0241] [Table 25]

[0242] When comparing lanadelumab 300 mg q2w and lanadelumab 300 mg q4w with placebo, Cox regression yielded HRs of 0.05 (95% CI; 0.02; 0.14) and 0.19 (95% CI; 0.10; 0.38), respectively (Table 25). The inclusion of additional covariates had no effect.

[0243] [Table 26]

[0244] When comparing Cinryze® and placebo, Cox regression yielded a HR of 0.32 (95% CI: 0.14; 0.71) (Table 26). These results were consistent with base case findings.

[0245] [Table 27]

[0246] Indirect comparison via Bucher's method showed that patients treated with lanadelumab 300 mg q2w had a 0.15-fold hazard of a first HAE attack after 70 days of treatment compared to patients treated with Cinryze® (Table 27). These results were statistically significant by not including the value "1" in the corresponding 95% CI (95% CI: 0.0431; 0.5286). Treatment with lanadelumab 300 mg q4w resulted in a 0.56-fold hazard of a HAE attack when compared to Cinryze®. These results comparing treatment with lanadelumab 300 mg q4w to treatment with Cinryze® were not statistically significant as evidenced by the inclusion of "1" in the corresponding 95% CI (95% CI: 0.2086, 1.5262).

[0247] [Table 28]

[0248] conclusion Consistent with the results of the Bayesian NMA using pooled data, our regression-based frequentist ITC showed that lanadelumab 300 mg q2w and lanadelumab 300 mg q4w were statistically more effective than Cinryze® in reducing the rate of HAE attacks per 28 days (RR 0.27, 95% CI [0.15; 0.49] and RR 0.54, 95% CI [0.35; 0.85], respectively). A sensitivity analysis including patient-reported HAE attacks in the HELP study confirmed these findings, yielding RR estimates and 95% Ci consistent with the primary results. A second sensitivity analysis including the covariates age, sex, and weight yielded similar findings, further supporting the outcomes of the base case analysis.

[0249] With regard to time to first seizure after 0 days of treatment and time to first seizure after 70 days of treatment, the ITC of the present invention reached conclusions similar to those described in the Bayesian NMA: lanadelumab 300 mg q2w and lanadelumab 300 mg q4w were not statistically effective compared to Cinryze® in extending the time to first seizure after 0 days of treatment (HR 0.73, 95% CI [0.29; 1.84] and HR 1.05, 95% CI [0.43, 2.52], respectively). However, lanadelumab 300 mg q2w and lanadelumab 300 mg q4w appeared to exhibit greater efficacy compared with Cinryze® in extending the time to first seizure after 70 days of treatment (HR 0.19, 95% CI [0.06; 0.62] and HR 0.62, 95% CI [0.23; 1.64], respectively). Only the difference between lanadelumab 300 mg q2w and Cinryze® was statistically significant. In sensitivity analyses for time to first seizure after 0 and 70 days of treatment, models including age, sex, and weight yielded lower HR estimates; however, this finding did not change any of the base-case conclusions.

[0250] The conclusions of our ITC were consistent with those of the Bayesian NMA, although the corresponding estimates and uncertainty intervals differed, likely due to the lack of consideration of repeated measures in the Bayesian NMA (for the CHANGE crossover trial) and differences in the types of data entry.

[0251] Other embodiments All features disclosed herein may be combined in any combination. Each feature disclosed herein may be replaced with another feature serving the same, equivalent, or similar purpose. Thus, unless otherwise expressly stated, each feature disclosed is merely an example of a generic series of equivalent or similar features.

[0252] From the above description, those skilled in the art can easily ascertain the important features of the present invention, and can make various changes and modifications to the present invention to suit various usages and conditions without departing from the spirit and scope thereof. Therefore, other embodiments are also within the scope of the claims.

[0253] Doctrine of Equivalents Although several embodiments of the present invention have been described and illustrated herein, those skilled in the art will readily envision various other means and / or structures for performing the functions and / or obtaining the results and / or one or more advantages described herein, and each such variation and / or modification is deemed to be within the scope of the embodiments of the present invention described herein. More generally, those skilled in the art will readily appreciate that all parameters, dimensions, materials, and configurations described herein are meant to be exemplary, and that the actual parameters, dimensions, materials, and / or configurations will vary based on the particular application in which the teachings are used. Those skilled in the art will recognize or be able to ascertain using no more than routine experimentation many equivalents to the specific embodiments of the present invention described herein. Thus, it will be understood that the above embodiments are presented by way of example only, and that within the scope of the appended claims and their equivalents, the embodiments of the present disclosure are directed to each individual feature, system, article, material, kit, and / or method described herein. Furthermore, any combination of two or more of such features, systems, articles, materials, kits, and / or methods is within the inventive scope of the present disclosure, where such features, systems, articles, materials, kits, and / or methods are not mutually exclusive.

[0254] All definitions and those used herein should be understood to control over dictionary definitions, definitions in documents incorporated by reference, and / or ordinary meanings of the defined terms.

[0255] The indefinite articles "a" and "an," as used herein in the specification and claims, unless expressly stated to the contrary, should be understood to mean "at least one."

[0256] The term "and / or" in the specification and claims, as used herein, should be understood to mean "either or both" of the coordinated elements, i.e., elements that are present conjunctively in some cases and disjunctively in other cases. Multiple elements listed with "and / or" are interpreted in the same manner, i.e., "one or more" of the coordinated elements. Optionally, other elements may be present other than the elements specifically identified by the "and / or" clause, whether related or unrelated to the specifically identified elements. Thus, as a non-limiting example, a reference to "A and / or B," when used with open-ended language such as "comprising," may refer in one embodiment to A only (optionally including elements other than B); in another embodiment to B only (optionally including elements other than A); in yet another embodiment to both A and B (optionally including other elements); and so forth.

[0257] As used herein in the specification and claims, "or" is understood to have the same meaning as "and / or" as defined above. For example, when separating items in a list, "or" or "and / or" is to be construed as inclusive, i.e., the inclusion of at least one (but also including more than one) of a number or list of elements, and optionally additional unlisted items. Only terms clearly stated to the contrary, such as "only one of" or "exactly one of," or, when used in the claims, "consisting of," refer to the inclusion of exactly one element of a number or list of elements. In general, the term "or," when used herein, when preceded by an exclusive term, such as "either," "one of," "only one of," or "exactly one of," is to be construed merely to represent exclusive alternatives (i.e., "one or the other, but not both"). "Consisting essentially of," when used in the claims, has its ordinary meaning as used in the field of patent law.

[0258] As used herein in the specification and claims, the phrase "at least one" in connection with a list of one or more elements is understood to mean at least one element selected from any one or more of the elements in the list of elements, but does not necessarily include at least one of every element specifically listed in the list of elements, and does not exclude any combination of elements in the list of elements. This definition also allows for the optional presence of elements other than those specifically identified in the list of elements referred to by the phrase "at least one," whether related or unrelated to the specifically identified elements. Thus, as a non-limiting example, "at least one of A and B (or, equivalently, "at least one of A or B" or, equivalently, "at least one of A and / or B") may represent, in one embodiment, at least one A, optionally including more than one, (and optionally including elements other than B), where B is absent; in another embodiment, at least one B, optionally including more than one, (and optionally including elements other than A); in a further embodiment, at least one A, optionally including more than one, and at least one B, optionally including more than one, (and optionally including other elements); etc.

[0259] Also, unless expressly stated to the contrary, in any method claimed herein that includes more than one step or action, it is understood that the order of the method steps or actions is not necessarily limited to the order in which the method steps or actions are described.

[0260] In the claims and the above specification, all transitional phrases such as "comprising," "including," "carrying," "having," "containing," "involving," "holding," "composed of," and the like, are to be understood to be open-ended, i.e., to mean including, but not limited to. Only the transitional phrases "consisting of" and "consisting essentially of" are closed or semi-closed transitional phrases, respectively, as set forth in Section 2111.03 of the United States Patent Office Manual of Examining Procedures.

Claims

1. 1. A method for treating or reducing the rate of hereditary angioedema (HAE) attacks, comprising: administering to a subject in need thereof an antibody that binds to human plasma kallikrein at about 300 mg about every two weeks for a first treatment period of about 4-9 months; monitoring the subject for HAE attacks during the first treatment period; reducing the dose of the antibody to about 300 mg about every 4 weeks in subjects who are not having HAE attacks during the first treatment period; Including, the antibody comprises the same heavy chain complementarity determining regions (CDRs) and the same light chain CDRs as DX-2930; method.

2. 10. The method of claim 1, wherein the first treatment period is for about six months.

3. The method of claim 1 or 2, wherein the patient has an underweight.

4. The method of claim 3, wherein the patient has a body weight of less than 35 kg.

5. The method of any one of claims 1 to 4, wherein the patient is a pediatric patient.

6. The antibody comprises a heavy chain immunoglobulin variable domain of SEQ ID NO: 3 (V Ｈ ) and the light chain immunoglobulin variable domain of SEQ ID NO: 4 (V Ｌ The method according to any one of claims 1 to 3, comprising:

7. The method of any one of claims 1 to 6, wherein the antibody is a full-length antibody or an antigen-binding fragment thereof.

8. The method of any one of claims 1 to 7, wherein the antibody is an IgG1 molecule.

9. The method of claim 8 , wherein the antibody comprises a heavy chain of SEQ ID NO:1 and a light chain of SEQ ID NO:

2.

10. The method of any one of claims 1 to 9, wherein the antibody is formulated in a pharmaceutical composition comprising a pharma- ceutically acceptable carrier.

11. 11. The method of claim 10, wherein the pharmaceutical composition comprises sodium phosphate, citric acid, histidine, sodium chloride, and polysorbate 80.

12. 12. The method of claim 11, wherein the sodium phosphate is at a concentration of about 30 mM, the citric acid is at a concentration of about 19 mM, the histidine is at a concentration of about 50 mM, the sodium chloride is at a concentration of about 90 mM, and the polysorbate 80 is at about 0.01%.

13. The method of any one of claims 1 to 12, wherein the antibody is administered subcutaneously.

14. The method of any one of claims 1 to 13, wherein the subject is a human patient having, suspected of having, or at risk for HAE.

15. 15. The method of claim 14, wherein the subject has HAEI type I or II.

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