Decreasing staphylococcus aureus infection in colonized patients

Abstract

To provide a method for treating a subject colonized by Staphylococcus aureus with an anti-α toxin antibody or antigen binding fragment thereof.SOLUTION: Disclosed is a method for treating a subject colonized by Staphylococcus aureus (S. aureus), the method comprising a step of administering an antibody or antigen binding fragment thereof that binds to α toxin (AT) of S. aureus to the subject, where a polymerase chain reaction (PCR) is used to detect the level of S. aureus in a sample taken from the subject.SELECTED DRAWING: None

Description

[Background technology]

[0001] Bacterial pneumonia occurring within hospitalized or intensive care unit (ICU) patient populations is a clinically significant and serious disease that contributes significantly to morbidity and mortality. It constitutes the second leading type of hospital-acquired infection and the leading cause of death due to hospital-acquired infections in the United States (Non-Patent Document 1). Staphylococcus aureus is the leading cause of hospital-acquired pneumonia. A recent study of European ICUs reported that 23% of mechanically ventilated ICU patients developed pneumonia caused by S. aureus, and more than half of these were caused by methicillin-resistant Staphylococcus aureus (MRSA) (Non-Patent Document 2).

[0002] Staphylococcus aureus also causes a wide range of additional diseases, including skin and soft tissue infections, endocarditis, osteomyelitis, pneumonia, and bacteremia (Non-Patent Document 3). During infection, S. aureus releases a number of toxic substances, including alpha-toxin (AT) as the most predominant virulence factor that causes tissue invasion and necrosis (Non-Patent Document 4). The crucial role played by AT in the pathogenesis of S. aureus is supported by animal models (skin necrosis, pneumonia, sepsis, endocarditis, and mastitis) and by analytical studies in humans, where the presence of anti-AT antibodies during severe infections was associated with improved outcomes.

[0003] Preclinical studies have shown that monoclonal antibody-based approaches are promising for prophylactic and adjuvant therapy against S. aureus infections (see, e.g., Non-Patent Document 5; Non-Patent Document 6; Non-Patent Document 7; and Non-Patent Document 8). Anti-AT antibodies have shown promising results in their ability to treat and prevent S. aureus infections. MEDI4893, or subratoxumab, is a human monoclonal antibody with a long half-life that binds with high affinity to AT and effectively blocks AT pore formation in target cell membranes. Preclinical results have demonstrated that prophylaxis with anti-AT antibodies containing the MEDI4893 binding region reduced disease severity in mouse infection models of dermonecrosis, pneumonia, and fatal bacteremia / sepsis (see, e.g., Patent Documents 1 and 2, each of which is incorporated herein by reference in its entirety). [Prior art documents] [Patent documents]

[0004] [Patent Document 1] International Publication No. 2012 / 109285 Brochure [Patent Document 2] International Publication No. 2014 / 074540 Brochure [Non-patent literature]

[0005] [Non-Patent Document 1] Spellberg and Talbot, 2010 [Non-Patent Document 2] Esperatti et al., 2010 [Non-Patent Document 3] Lowy, FD, N. Engl. J. Med., 339(8):520-32(1998) [Non-Patent Document 4] Wilke and Bubeck Wardenburg,2010 [Non-Patent Document 5] Hazenbos et al.,PLoS Pathog.,9(10):e1003653.doi:10.1371 / journal.ppat.10036532013(2013) [Non-Patent Document 6] Rouha, H., MAbs,7(1):243-254(2015);Foletti et al.,J.Mol.Biol.,425(10):1641-1654(2013) [Non-Patent Document 7] Karauzum et al., J Biol Chem.,287(30):25203-15(2012) [Non-Patent Document 8] Hua et al., Antimicrob Agents Chemother.,58(2):1108-17(2014) Summary of the Invention [Problem to be solved by the invention]

[0006] However, S. aureus infections, such as pneumonia, can develop very rapidly in patients colonized with S. aureus, so methods are needed to identify at-risk patients who will achieve the greatest benefit from anti-AT antibodies. [Means for solving the problem]

[0007] Staphylococcus aureus (S. aureus) pneumonia is an early and life-threatening complication in mechanically ventilated intensive care unit (ICU) patients despite infection management and antibiotics. As demonstrated herein, anti-alpha toxin (AT) antibodies have been evaluated for the prevention of S. aureus pneumonia and have been shown to be associated with clinically meaningful efficacy (≥25% reduction in associated risk) and acceptable safety. In particular, a 32% reduction in S. aureus pneumonia was observed in patients receiving anti-AT antibodies, without any safety concerns. Furthermore, even greater efficacy was observed in certain subgroups of patients. Thus, provided herein is a method for identifying patients at high risk of developing S. aureus infections who would benefit from receiving anti-AT antibodies.

[0008] Provided herein is a method of treating a subject colonized with Staphylococcus aureus (S. aureus), comprising administering to the subject an antibody or antigen-binding fragment thereof that binds to S. aureus alpha-toxin (AT), wherein polymerase chain reaction (PCR) is used to detect levels of S. aureus in a sample obtained from the subject. In certain examples, the sample obtained from the subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with a polymerase chain reaction (PCR) cycle threshold (Ct) value. In certain examples, the method reduces the incidence of infection associated with the presence of S. aureus in the subject.

[0009] Provided herein is a method for preventing a Staphylococcus aureus (S. aureus) infection in a subject, comprising administering to the subject an antibody or antigen-binding fragment thereof that binds to S. aureus AT, wherein a sample obtained from the subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with the PCR Ct value.

[0010] Provided herein is a method comprising administering subratoxumab to a subject to reduce the incidence of S. aureus pneumonia in the subject, the reduction being determined by clinical, microbiological and radiographic means, optionally resulting in a reduction in incidence of about 30%.

[0011] Provided herein are methods that include administering subratoxumab to a subject to reduce the incidence of pneumonia of any cause in the subject, wherein the reduction is determined by clinical, microbiological and radiographic means, optionally resulting in a reduction in incidence of about 30%.

[0012] In certain instances, infection is determined by clinical, microbiological and radiographic means.

[0013] In certain examples, clinical measures include abnormal temperature, abnormal white blood cell count, cough, purulent sputum, bronchial breath sounds, dyspnea, tachypnea (respiratory rate >30 breaths / min), hypoxia, or any combination thereof.

[0014] In certain examples, microbiological means include respiratory specimens, blood cultures, pleural fluid aspirates, or lung tissue cultures that are positive for S. aureus.

[0015] In the given example, radiographic measures include new or worsening infiltrate on a chest x-ray.

[0016] In certain examples, PCR has been used to detect levels of S. aureus in a sample obtained from a subject. In certain examples, the sample obtained from the subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with a polymerase chain reaction (PCR) cycle threshold (Ct) value.

[0017] In certain examples, the methods provided herein further include detecting a level of S. aureus in a sample obtained from the subject.

[0018] In certain examples, a sample obtained from a subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with a PCR Ct value of greater than or equal to 29. In certain examples, a PCR Ct value of 29 corresponds to a concentration of about 1,600 to about 1,700 colony forming units (CFU) / mL of S. aureus.

[0019] Provided herein is a method of treating a subject colonized with a S. aureus infection, comprising administering to the subject an antibody or antigen-binding fragment thereof that binds to S. aureus AT, wherein a sample obtained from the subject has a concentration of S. aureus not exceeding 1,700 CFU / mL. In certain examples, the method reduces the incidence of infection associated with the presence of S. aureus in the subject.

[0020] Provided herein is a method for preventing a Staphylococcus aureus (S. aureus) infection in a subject, the method comprising administering to the subject an antibody or antigen-binding fragment thereof that binds to S. aureus AT, wherein a sample obtained from the subject has a concentration of S. aureus not exceeding 1,700 CFU / mL.

[0021] In the example given, the concentration of S. aureus AT was measured using PCR.

[0022] In certain instances, the subject is colonized with S. aureus.

[0023] In certain examples, a sample obtained from a subject has a level of S. aureus that correlates with a PCR Ct value of at least 3. In certain examples, a sample obtained from a subject has a level of S. aureus that correlates with a PCR Ct value of at least 3.

[0024] In certain examples, levels of S. aureus are detected within 3 hours, optionally within 2 hours.

[0025] In the example given, the PCR detects S. aureus Protein A.

[0026] In certain instances, the subject is ventilated, and optionally the subject is on a respirator. In certain instances, the subject is taking antibiotics.

[0027] In certain instances, the sample is a skin or soft tissue sample. In certain instances, the sample is obtained from the lower respiratory tract of the subject. In certain instances, the sample is an endotracheal aspirate. In certain instances, the sample is a tracheal sample. In certain instances, the sample is a bronchial sample.

[0028] In certain examples, the sample contains bacteria that would not grow in culture to identify S. aureus. In certain examples, the sample contains bacteria that are not Staphylococcus. In certain examples, the S. aureus is antibiotic resistant. In certain examples, the methods provided herein further include determining whether the S. aureus is antibiotic resistant.

[0029] In certain examples, the S. aureus is methicillin resistant. In certain examples, the methods provided herein further include determining whether the S. aureus is methicillin resistant.

[0030] In certain instances, resistance is determined using PCR.

[0031] In certain instances, the infection is pneumonia. In certain instances, the infection is ICU (intensive care unit) pneumonia.

[0032] In certain instances, the subject is a human.

[0033] In certain examples, an antibody or antigen-binding fragment thereof that binds to the S. aureus AT binds to the same S. aureus AT epitope as an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 7 and a VL comprising the amino acid sequence of SEQ ID NO: 8. In certain examples, an antibody or antigen-binding fragment thereof that binds to the S. aureus AT competitively inhibits binding to the S. aureus AT of an antibody comprising a VH comprising the amino acid sequence of SEQ ID NO: 7 and a VL comprising the amino acid sequence of SEQ ID NO: 8.

[0034] In certain examples, an antibody or antigen-binding fragment thereof that binds to S. aureus AT comprises a VH CDR1, a VH CDR2, a VH CDR3, a VL CDR1, a VL CDR2, and a VL CDR3 of MEDI4893. In certain examples, an antibody or antigen-binding fragment thereof that binds to S. aureus AT comprises a variable heavy chain (VH) complementarity determining region (CDR)1 comprising the amino acid sequence of SEQ ID NO:1, a VH CDR2 comprising the amino acid sequence of SEQ ID NO:2, a VH CDR3 comprising the amino acid sequence of SEQ ID NO:3, a variable light chain (VL) CDR1 comprising the amino acid sequence of SEQ ID NO:4, a VL CDR2 comprising the amino acid sequence of SEQ ID NO:5, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO:6.

[0035] In certain examples, an antibody or antigen-binding fragment thereof that binds to the S. aureus AT comprises a VH comprising the amino acid sequence of SEQ ID NO: 7. In certain examples, an antibody or antigen-binding fragment thereof that binds to the S. aureus AT comprises a VL comprising the amino acid sequence of SEQ ID NO: 8.

[0036] In certain examples, an antibody or antigen-binding fragment thereof that binds to the S. aureus AT comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 9. In certain examples, an antibody or antigen-binding fragment thereof that binds to the S. aureus AT comprises a light chain comprising the amino acid sequence of SEQ ID NO: 10.

[0037] In some instances, the antibody or antigen-binding fragment that binds to S. aureus AT further comprises a heavy chain constant region. In some instances, the heavy chain constant region is selected from the group consisting of human immunoglobulin IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2 heavy chain constant regions. In some instances, the heavy chain constant region is a human IgG1 constant region.

[0038] In certain instances, the antibody or antigen-binding fragment that binds to S. aureus AT further comprises a light chain constant region. In certain instances, the light chain constant region is selected from the group consisting of human immunoglobulin IgGκ and IgGλ light chain constant regions. In certain instances, the light chain constant region is a human IgGκ light chain constant region.

[0039] In certain examples, the antibody or antigen-binding fragment thereof that binds to S. aureus AT is an IgG antibody or antigen-binding fragment thereof.

[0040] In certain examples, the antibody or antigen-binding fragment thereof that binds to the S. aureus AT comprises an Fc region that has been genetically engineered to improve half-life. In certain examples, the antibody or antigen-binding fragment thereof that binds to the S. aureus AT comprises an Fc region that includes a YTE mutation.

[0041] In certain examples, the antibody or antigen-binding fragment that binds to S. aureus AT is a monoclonal antibody or antigen-binding fragment.

[0042] In certain examples, the antibody or antigen-binding fragment that binds to the S. aureus AT is a full-length antibody. In certain examples, the antibody or antigen-binding fragment that binds to the S. aureus AT is an antigen-binding fragment. In certain examples, the antigen-binding fragment is a Fab, Fab', F(ab')2, single chain Fv (scFv), disulfide-linked Fv, intrabody, IgGΔCH2, minibody, F(ab')3, tetrabody, triabody, diabody, DVD-Ig, Fcab, mAb, or a combination thereof. 2 , (scFv)2 or scFv-Fc.

[0043] In certain examples, an antibody or antigen-binding fragment thereof that binds to S. aureus AT has an affinity for S. aureus AT of between 80 and 100 pM.

[0044] In certain examples, the antibody or antigen-binding fragment thereof is subratoxumab.

[0045] In certain instances, 2,000 mg of the antibody or antigen-binding fragment is administered. In certain instances, 5,000 mg of the antibody or antigen-binding fragment is administered.

[0046] In certain examples, preventing S. aureus infection includes neutralizing toxins, inducing opsonophagocytosis, inhibiting thromboembolic lesion formation, inhibiting S. aureus-associated sepsis, or any combination of the above.

[0047] Provided herein is an antibody or antigen-binding fragment that binds to a Staphylococcus aureus (S. aureus) AT for use in treating a subject colonized with S. aureus, wherein a sample obtained from the subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with a polymerase chain reaction (PCR) cycle threshold (Ct) value. In certain examples, the treating reduces the incidence of infection associated with the presence of S. aureus in the subject.

[0048] Provided herein is an antibody or antigen-binding fragment thereof that binds to S. aureus AT for use in preventing S. aureus infection in a subject colonized with S. aureus, wherein a sample obtained from the subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with the PCR Ct value.

[0049] In certain examples, a sample obtained from a subject has a level of S. aureus that does not exceed the level of S. aureus that correlates with a PCR Ct value of 29. In certain examples, an antibody or antigen-binding fragment thereof is not administered to a subject in instances in which the level of S. aureus in a sample obtained from a subject is detected that does not exceed the level of S. aureus that correlates with the PCR Ct value.

[0050] Provided herein is an in vitro method for identifying a subject colonized with S. aureus that is responsive to an antibody or antigen-binding fragment thereof that binds to S. aureus AT, comprising detecting a level of S. aureus in a sample obtained from the subject, wherein a level of S. aureus that does not exceed a level of S. aureus that correlates with a PCR Ct value indicates responsiveness to the antibody or antigen-binding fragment thereof. In certain examples, the PCR Ct value is 29. [Brief description of the drawings]

[0051] [Figure 1] FIG. 1 is a diagram illustrating the use of S. aureus PCR testing to rapidly identify patients. The PCR test generates a cycle threshold (Ct) value that represents the number of PCR cycles required to reach a threshold signal (represented by the horizontal line in the graph on the right side of the figure). The Ct value is inversely proportional to the bacterial load: the more S. aureus in a sample, the fewer cycles required to reach the threshold level and therefore have a lower Ct value (curve on the left side of the graph), whereas the less S. aureus in a sample, the more cycles required to reach the threshold level and therefore have a higher Ct value (curve on the right side of the graph). (See Example 1.) [Diagram 2] 1 is a histogram showing Ct values ​​obtained from patients at screening in a Phase II clinical trial. A significant number of patients had low Ct values ​​(high bacterial load) at screening. (See Example 2.) [Diagram 3] Graph showing variability of Ct values ​​over time on the ventilator. Even during the first few days, Ct values ​​had a wide range of distribution. (See Example 2.) [Figure 4]FIG. 1 shows results from three types of culture assays: a quantitative assay measuring CFU / mL, a semi-quantitative assay (culture score, where mild=+ or ++; moderate=+++ and severe=+++), and a qualitative assay (present / absent). Over half of the subjects tested had a high bacterial load (defined by either at least 105 CFU / mL (in the quantitative assay) or a moderate or severe culture score (sure) (in the semi-quantitative assay). (See Example 3.) [Diagram 5] Graph showing Ct values ​​obtained from samples with mild, moderate or severe scores in a semi-quantitative assay (see Example 3). [Figure 6] 1 is a graph showing concordance and discordance between PCR and culture assays for S. aureus. Of 209 samples that tested positive in the PCR assay, 162 also tested positive in the culture assay (77.5% concordance rate), while 47 of them were negative in the culture assay (22.5% discordance rate). All discordant results were from samples that tested negative in the culture assay and positive in the PCR assay. Thus, the PCR assay is much more sensitive than the culture assay. (See Example 3.) [Figure 7] 1 is a graph showing that antibiotic use adversely affects culture results. The percentage of patients taking antibiotics was significantly higher in the group of patients who tested positive for S. aureus by PCR, while negative test results for S. aureus by culture were comparable to the group of patients who tested positive for S. aureus by both PCR and culture. (See Example 3.) [Figure 8] 1 is a graph showing that antibiotic history did not affect culture results (see Example 3). [Figure 9]1 is a graph showing the distribution of culture positive and culture negative samples by PCR Ct value. 85% of the culture negative samples have a Ct value higher than 29, and about 80% of the culture positive samples have a Ct value lower than 29. Thus, a Ct value of 29 effectively distinguishes between culture positive and culture negative samples. (See Example 4.) [Figure 10] 1 is a graph showing the relationship between Ct value and the concentration of S. aureus (colony forming units (CFU) / mL) in samples measured in a Phase 2 clinical trial. There was a statistically significant inverse correlation between Ct value and the number of CFU / mL. The majority of samples with Ct values ​​greater than 29 had S. aureus loads of 103 CFU / mL or less. The majority of samples with Ct values ​​less than 29 had S. aureus loads of more than 103 CFU / mL. A Ct value of 29 corresponds to approximately 3.2 log10 CFU / mL (approximately 1,600-1,700 CFU / mL), which effectively distinguishes between low and high colony formers. (See Example 4.) [Figure 11] Graph showing the correlation between sample Ct value and the concentration of S. aureus (colony forming units (CFU) / mL) in tracheal (left graph) and bronchial (right graph) samples. These results demonstrate that Ct values ​​are inversely correlated with S. aureus loading in both tracheal and bronchial samples. (See Example 4.) [Figure 12]Graph showing correlation between Ct value in samples and concentration of S. aureus (colony forming units (CFU) / mL) in samples with non-Staphylococcus growth (left graph) and without non-Staphylococcus growth (right graph). These results show that Ct value is inversely correlated with S. aureus loading, independent of the presence or absence of non-Staphylococcus bacteria in the sample, proving that the PCR test is specific for S. aureus. (See Example 4.) DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0052] The present disclosure is directed to methods for preventing S. aureus infections in patients with low levels of S. aureus colonization, as well as methods for identifying patients who are colonized with S. aureus and who would benefit from alpha-toxin antibodies.

[0053] I. Definition In the context of describing the present invention (particularly in the context of the claims which follow), use of the terms "a," "an," "the," and "at least one," and similar referents, should be construed to encompass both the singular and the plural, unless otherwise indicated in the specification or clearly contradicted by context.

[0054] Use of the term "at least one" following a list of one or more items (e.g., "at least one of A and B") should be construed to mean one item selected from the listed items (A or B), or any combination of two or more of the listed items (A and B), unless otherwise indicated in the specification or clearly contradicted by context.

[0055] The terms "comprise," "have," "include," and "contain" should be construed as open-ended terms (i.e., meaning "including, but not limited to"), unless otherwise noted.

[0056] The term "alpha toxin" or "AT" as used herein refers to bacterial alpha toxin polypeptides, including, but not limited to, native alpha toxin polypeptides and isoforms of alpha toxin polypeptides. "Alpha toxin" includes full-length unprocessed alpha toxin polypeptides as well as forms of alpha toxin polypeptides that arise as a result of processing within the cell. As used herein, the term "S. aureus alpha toxin" refers to: [ka] It refers to a polypeptide comprising the amino acid sequence of

[0057] The S. aureus alpha-toxin H35L mutant has the sequence [ka] has.

[0058] "Alpha toxin polynucleotide," "alpha toxin nucleotide," or "alpha toxin nucleic acid" refers to a polynucleotide that encodes an alpha toxin.

[0059] The term "antibody" refers to an immunoglobulin molecule that recognizes and specifically binds to a target, such as a protein, polypeptide, peptide, carbohydrate, polynucleotide, lipid, or a combination of the above, through at least one antigen recognition site within the variable region of the immunoglobulin molecule. As used herein, the term "antibody" encompasses intact polyclonal antibodies, intact monoclonal antibodies, chimeric antibodies, humanized antibodies, human antibodies, fusion proteins containing antibodies, and any other modified immunoglobulin molecule, so long as the antibody exhibits the desired biological activity. Antibodies can be any of the five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, or their subclasses (isotypes) (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), based on the identity of their heavy chain constant domains, called α (alpha), δ (delta), ε (epsilon), γ (gamma), and μ (mu), respectively. The different classes of immunoglobulins have different well-known subunit structures and three-dimensional configurations. Antibodies can be naked or can be conjugated to other molecules, such as toxins, radioisotopes, etc.

[0060] The term "monoclonal antibody" as used herein refers to an antibody produced by a single clone of B cells and that binds to the same epitope. In contrast, the term "polyclonal antibody" refers to a population of antibodies produced by different B cells and that bind to different epitopes of the same antigen.

[0061] The term "antibody fragment" refers to a portion of an intact antibody. An "antigen-binding fragment", "antigen-binding domain" or "antigen-binding region" refers to a portion of an intact antibody that binds to an antigen. An antigen-binding fragment may include an antigen-determining region of an intact antibody (e.g., a complementarity-determining region (CDR)). Examples of antigen-binding fragments of antibodies include, but are not limited to, Fab, Fab', F(ab')2 and Fv fragments, linear antibodies and single-chain antibodies. Antigen-binding fragments of antibodies can be obtained from any animal species, such as rodents (e.g., mice, rats or hamsters) and humans, or can be artificially generated.

[0062] A whole antibody typically consists of four polypeptides: two identical copies of a heavy (H) chain polypeptide and two identical copies of a light (L) chain polypeptide. Each heavy chain contains one N-terminal variable (VH) region and three C-terminal constant (CH1, CH2 and CH3) regions, and each light chain contains one N-terminal variable (VL) region and one C-terminal constant (CL) region. The variable regions of each pair of light and heavy chains form the antigen-binding site of the antibody. The VH and VL regions have the same general structure, and each region contains four framework regions whose sequences are relatively conserved. As used herein, the term "framework region" refers to the relatively conserved amino acid sequences within the variable regions located between the hypervariable regions or complementarity determining regions (CDRs). There are four framework regions in each variable domain, designated FR1, FR2, FR3 and FR4. These framework regions form a β-sheet that provides the structural framework for the variable region (see, e.g., CA Janeway et al. (eds.), Immunobiology, 5th Ed., Garland Publishing, New York, NY (2001)). The three CDRs, known as CDR1, CDR2 and CDR3, form the "hypervariable region" of an antibody, which is responsible for antigen binding.

[0063] The terms "VL" and "VL domain" are used interchangeably to refer to the light chain variable region of an antibody.

[0064] The terms "VH" and "VH domain" are used interchangeably to refer to the heavy chain variable region of an antibody.

[0065] The term "Kabat numbering" and similar terms are art-recognized and refer to a system for numbering amino acid residues in the heavy and light chain variable regions of an antibody or antigen-binding fragment thereof. In certain embodiments, the CDRs may be determined according to the Kabat numbering system (see, e.g., Kabat EA & Wu TT (1971) Ann NY Acad Sci 190:382-391 and Kabat EA et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, USDepartment of Health and Human Services, NIH Publication No. 91-3242). Using the Kabat numbering system, the CDRs in an antibody heavy chain molecule are typically located at amino acid positions 31-35 (CDR1), 50-65 (CDR2), and 95-102 (CDR3), which may optionally include one or two additional amino acids following 35 (referred to as 35A and 35B in the Kabat numbering scheme). Using the Kabat numbering system, the CDRs in an antibody light chain molecule are typically located at amino acid positions 24-34 (CDR1), 50-56 (CDR2), and 89-97 (CDR3). In certain embodiments, the CDRs of the antibodies described herein have been determined according to the Kabat numbering scheme.

[0066] Chothia refers instead to the location of the structural loops (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)). The ends of the Chothia CDR-H1 loop when numbered using the Kabat numbering convention vary from H32 to H34 depending on the length of the loop (this is because the Kabat numbering scheme places insertions at H35A and H35B; if neither 35A nor 35B are present, the loop ends at 32, if only 35A is present, the loop ends at 33, and if both 35A and 35B are present, the loop ends at 34). The AbM hypervariable regions represent a compromise between the Kabat CDRs and the Chothia structural loops and are used by Oxford Molecular's AbM antibody modeling software.

[0067] [Table 1]

[0068] The terms "constant region" or "constant domain" as used herein are interchangeable and have the usual meaning in the art. The constant region is the portion of an antibody, e.g., the carboxyl-terminal portion of the light and / or heavy chain, that is not directly involved in binding the antibody to an antigen, but may exhibit various effector functions, such as interaction with Fc receptors. The constant region of an immunoglobulin molecule generally has a more conserved amino acid sequence compared to the immunoglobulin variable domain.

[0069] As used herein, the term "heavy chain," when used in reference to an antibody, refers to any distinct type, e.g., α (alpha), δ (delta), ε (epsilon), γ (gamma), and μ (mu), which give rise to the IgA, IgD, IgE, IgG, and IgM classes of antibodies, respectively, including the IgG, e.g., IgG1, IgG2, IgG3, and IgG4 subclasses, based on the amino acid sequence of the constant domain. Heavy chain amino acid sequences are well known in the art. In certain embodiments, the heavy chain is a human heavy chain.

[0070] As used herein, the term "light chain," when used in reference to an antibody, can refer to any distinct type, e.g., κ (kappa) or λ (lambda), based on the amino acid sequence of the constant domain. Light chain amino acid sequences are well known in the art. In certain embodiments, the light chain is a human light chain.

[0071] A "chimeric" antibody refers to an antibody or fragment thereof that contains both human and non-human regions. A "humanized" antibody is an antibody that contains a human antibody framework and at least one CDR obtained or derived from a non-human antibody. Non-human antibodies include antibodies isolated from any non-human animal, such as rodents (e.g., mice or rats). A humanized antibody may contain one, two, or three CDRs obtained or derived from a non-human antibody. A fully human antibody does not contain any amino acid residues obtained or derived from a non-human animal. It will be understood that fully human and humanized antibodies have a lower risk of eliciting an immune response in humans compared to murine or chimeric antibodies (see, e.g., Harding et al., mAbs, 2(3):256-26 (2010)).

[0072] As used herein, "epitope" is a term of the art and refers to a localized region of an antigen to which an antibody or antigen-binding fragment thereof can specifically bind. An epitope can be, for example, consecutive amino acids of a polypeptide (linear or continuous epitope), or an epitope can be, for example, joined together from two or more non-contiguous regions of one or more polypeptides (conformational, non-linear, discontinuous or non-contiguous epitope). In certain embodiments, the epitope to which an antibody or antigen-binding fragment thereof binds can be determined, for example, by NMR spectroscopy, X-ray diffraction crystallography, ELISA assay, hydrogen / deuterium exchange with mass spectrometry (e.g., liquid chromatography liquid spray mass spectrometry), array-based oligopeptide scanning assay, and / or mutagenesis mapping (e.g., site-directed mutagenesis mapping). In the case of X-ray crystallography, crystallization can be achieved using any of the methods known in the art (e.g., Giege R et al., (1994) Acta Crystallogr D Biol Crystallogr 50(Pt 4):339-350; McPherson A (1990) Eur J Biochem 189:1-23; Chayen NE (1997) Structure 5:1269-1274; McPherson A (1976) J Biol Chem 251:6300-6303).Crystals of the antibody / antigen-binding fragment thereof:antigen can be studied using known X-ray diffraction techniques and refined using computer software such as X-PLOR (Yale University, 1992, distributed by Molecular Simulations, Inc.; see e.g., Meth Enzymol (1985) volumes 114&115, eds Wyckoff HW et al.,; U.S. Patent Application Publication No. 2004 / 0014194) and BUSTER (Bricogne G (1993) Acta Crystallogr D Biol Crystallogr 49(Pt 1):37-60; Bricogne G (1997) Meth Enzymol 276A:361-423, ed Carter CW; Roversi P et al., (2000) Acta Crystallogr D Biol Crystallogr 56(Pt 10):1316-1323). Mutagenesis mapping studies can be accomplished using any method known to those of skill in the art, e.g., for a description of mutagenesis techniques such as alanine scanning mutagenesis techniques, see Champe M et al., (1995) J Biol Chem 270:1388-1394 and Cunningham BC & Wells JA (1989) Science 244:1081-1085.

[0073] An antibody that "binds to the same epitope" as a reference antibody refers to an antibody that binds to the same amino acid residue as the reference antibody. The ability of an antibody to bind to the same epitope as a reference antibody can be determined by hydrogen / deuterium exchange assays (see Coales et al. Rapid Commun. Mass Spectrom. 2009;23:639-647) or X-ray crystallography.

[0074] As used herein, the terms "immunospecifically binds," "immunospecifically recognizes," "specifically binds," and "specifically recognizes" are analogous terms in the context of an antibody or antigen-binding fragment thereof. These terms mean that the antibody or antigen-binding fragment thereof binds to an epitope via its antigen-binding domain and that binding requires some complementarity between the antigen-binding domain and the epitope. Thus, an antibody that "specifically binds" to a first S. aureus leukotoxin may also bind to other S. aureus leukotoxins, but the extent of binding to unrelated, non-leukotoxin proteins is less than about 10% of the binding of the antibody to the first S. aureus leukotoxin as measured, for example, by radioimmunoassay (RIA), enzyme-linked immunosorbent assay (ELISA), BiaCore, or octet binding assays.

[0075] An antibody is said to "competitively inhibit" the binding of a reference antibody to a given epitope if it preferentially binds to that epitope or an overlapping epitope to the extent that it blocks the binding of the reference antibody to that epitope to some extent. Competitive inhibition can be determined by any method known in the art, such as a competitive ELISA assay. An antibody can be said to competitively inhibit the binding of the reference antibody to a given epitope by at least 90%, at least 80%, at least 70%, at least 60%, or at least 50%.

[0076] The term "nucleic acid sequence" is intended to encompass polymers of DNA or RNA, i.e., polynucleotides that may be single-stranded or double-stranded and may contain non-natural or modified nucleotides. As used herein, the terms "nucleic acid" and "polynucleotide" refer to polymeric forms of nucleotides of any length, either ribonucleotides (RNA) or deoxyribonucleotides (DNA). These terms refer to the primary structure of the molecule and thus include double-stranded and single-stranded DNA as well as double-stranded and single-stranded RNA. These terms include, as equivalents, nucleotide analogs and modified polynucleotides, such as, but not limited to, analogs of either RNA or DNA made from methylated and / or capped polynucleotides. Nucleic acids are typically linked via phosphate bonds to form nucleic acid sequences or polynucleotides, although many other linkages are known in the art (e.g., phosphorothioates, boranophosphates, etc.).

[0077] As used herein, "transfection," "transformation," or "transduction" refers to the introduction of one or more exogenous polynucleotides into a host cell by using physical or chemical methods. Many transfection techniques are known in the art, including calcium phosphate DNA coprecipitation (see, e.g., Murray EJ (ed.), Methods in Molecular Biology, Vol. 7, Gene Transfer and Expression Protocols, Humana Press (1991)); DEAE-dextran; electroporation; cationic liposome-mediated transfection; tungsten particle-facilitated microparticle bombardment (Johnston, Nature, 346:776-777 (1990)); and strontium phosphate DNA coprecipitation (see Brash et al, Mol. Cell Biol., 7:2031-2034 (1987)). Phage or viral vectors can be introduced into host cells following propagation of infectious particles in suitable packaging cells, many of which are commercially available.

[0078] As used herein, the terms "treatment," "treating," and the like refer to measures (e.g., administration of an antibody or antigen-binding fragment thereof provided herein to a subject) that cures, slows, alleviates symptoms, and / or halts progression of a diagnosed pathological condition or disorder. Thus, subjects in need of treatment include those already diagnosed with the disorder or suspected of having the disorder. A "therapeutically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve a desired therapeutic result (e.g., treatment of a S. aureus infection).

[0079] Prophylactic or preventative measures refer to measures (e.g., administration of an antibody or antigen-binding fragment thereof provided herein to a subject) that prevent and / or slow the onset of a targeted pathological condition or disorder. Subjects in need of prophylactic or preventative measures thus include subjects susceptible to a disorder and subjects in which a disorder is to be prevented. A "prophylactically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired prophylactic result (e.g., prevention of S. aureus infection or onset of disease).

[0080] A subject "colonized" with S. aureus refers to a subject having S. aureus present in or on the body. Colonization can be determined, for example, by detecting S. aureus in a sample obtained from the subject. S. aureus can be detected, for example, by culturing or polymerase chain reaction (PCR). Infections resulting from or associated with the presence of S. aureus in or on the subject's body show radiographic and / or clinical signs of the bacteria. S. aureus infections can occur, for example, as skin or soft tissue infections (SSTIs) or bacteremia. S. aureus bacteria can travel through the bloodstream and infect sites within the body, resulting in pneumonia, ICU pneumonia, bone or joint infections, device infections, wound infections, surgical site infections, or osteomyelitis. Radiographic signs include, for example, x-rays showing infiltrates. Clinical signs include, for example, abnormal temperature, abnormal white blood cell count, cough, purulent sputum, bronchial breath sounds, dyspnea, tachypnea (respiratory rate >30 breaths / min), and / or hypoxia.

[0081] As used herein, the terms "administer," "administering," "administration," and the like refer to methods (e.g., intravenous administration) that can be used to enable delivery of a drug, such as an anti-S. aureus antibody or combination of antigen-binding fragments thereof, to a desired site of biological action. Administration techniques that can be used with the agents and methods described herein can be found, for example, in: Goodman and Gilman, The Pharmacological Basis of Therapeutics, current edition, Pergamon; and Remington's, Pharmaceutical Sciences, current edition, Mack Publishing Co., Easton, Pa.

[0082] Administration "in combination with" one or more further therapeutic agents includes simultaneous (concurrent) and consecutive administration in any order.

[0083] As used herein, unless otherwise stated or clear from the context, the term "or" is understood to be inclusive. As used herein in phrases such as "A and / or B," the term "and / or" is intended to include both "A and B," "A or B," and "A" and "B." Similarly, as used in phrases such as "A, B, and / or C," the term "and / or" is intended to include each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B, or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0084] II. Anti-Staphylococcus aureus α-toxin antibody As provided herein, antibodies and antigen-binding fragments thereof (e.g., monoclonal antibodies and fragments) that bind to S. aureus alpha-toxin can be used to avoid S. aureus in subjects with low levels of S. aureus colonization.

[0085] Alpha-toxin (AT) is an important virulence factor for several S. aureus conditions, including pneumonia, skin and soft tissue infections (SSTIs), bacteremia, etc. (Bubeck Wardenburg, J. and O. Schneewind, J. Exp. Med., 205:287-294 (2008); Inoshima et al., J. Invest. Dermatol., 132:1513-1516 (2012); and Foletti et al., supra). Passive immunization with anti-AT monoclonal antibodies reduced disease severity in pneumonia and dermonecrosis models (Hua et al., Antimicrob. Agents Chemother., 58:1108-1117 (2014); Tkaczyk et al., Clin. Vaccine Immunol., 19:377-385 (2012); and Ragle, BE and J. Wardenburg Bubeck, Infect. Immun., 77:2712-2718 (2009)), and vaccination with AT toxoids containing the H35L mutation (ATH35L) prevented death in mouse lethal bacteremia and pneumonia models (Bubeck Wardenburg, supra; Foletti et al., supra; Hua et al., supra; Ragle, supra; Menzies, BE and DS Kernodle, Infect. Immun., 77:2712-2718 (2009); and Adhikari et al., PLoS One, 7:e38567 (2012)). AT contributes to multiple aspects of S. aureus pathogenesis during bacteremia and sepsis, including stimulation of the hyperinflammatory response characteristic of sepsis and activation of ADAM10-mediated endothelial tight junction scission, leading to loss of vascular integrity (Powers et al., J Infect. Dis., 206:352-356 (2012); Wilke, GA and J. Bubeck Wardenburg, Proc. Natl. Acad. Sci. USA, 107:13473-13478 (2010); and Becker et al., J Innate Immun., 6:619-631 (2014)).AT has also been demonstrated to target platelets, which prevents the repair of damaged endothelial barriers and promotes organ dysfunction through the formation of platelet-neutrophil aggregates (Powers et al., Cell Host Microbe, 17:775-787 (2015)). The structure and function of alpha-toxin have been described in detail, for example, in Bhakdi, S. and J. Tranum-Jensen, Microbiol. Mol. Biol. Rev., 55(4):733-751 (1991).

[0086] Monoclonal and polyclonal antibodies that bind to AT are also known in the art (see, e.g., Hua et al., Antimicrob. Agents Chemother., 58(2):1108-1117 (2014); and Oganesyan et al., J. Biol. Chem., 289:29874-29880 (2014)) and are commercially available from sources such as, for example, Sigma Aldrich (St. Louis, MO) and AbCam (Cambridge, MA). Exemplary antibodies that bind to AT are disclosed in WO 2012 / 109285 and WO 2014 / 074540, both of which are incorporated by reference in their entireties.

[0087] In one example, an antibody or antigen-binding fragment (e.g., a monoclonal antibody or fragment) that specifically binds to S. aureus alpha-toxin (AT) comprises, consists essentially of, or consists of (i) a heavy chain polypeptide comprising a CDR1 amino acid sequence of SEQ ID NO: 1, a CDR2 amino acid sequence of SEQ ID NO: 2, and a CDR3 amino acid sequence of SEQ ID NO: 3, and (ii) a light chain polypeptide comprising a CDR1 amino acid sequence of SEQ ID NO: 4, a CDR2 amino acid sequence of SEQ ID NO: 5, and a CDR3 amino acid sequence of SEQ ID NO: 6. In another example, an antibody or antigen-binding fragment heavy chain polypeptide (e.g., a monoclonal antibody or fragment) that specifically binds to S. aureus AT comprises, consists essentially of, or consists of a variable region amino acid sequence of SEQ ID NO: 7. In another example, an antibody or antigen-binding fragment light chain polypeptide (e.g., a monoclonal antibody or fragment) that specifically binds to S. aureus AT comprises, consists essentially of, or consists of a variable region amino acid sequence of SEQ ID NO: 8. In another example, an antibody or antigen-binding fragment (e.g., a monoclonal antibody or fragment) that specifically binds to S. aureus AT comprises, consists essentially of, or consists of a variable heavy chain that comprises, consists essentially of, or consists of the amino acid sequence of SEQ ID NO: 7 and a light chain variable region that comprises, consists essentially of, or consists of the amino acid sequence of SEQ ID NO: 8. In another example, an antibody or antigen-binding fragment (e.g., a monoclonal antibody or fragment) that specifically binds to S. aureus AT comprises, consists essentially of, or consists of a heavy chain that comprises, consists essentially of, or consists of the amino acid sequence of SEQ ID NO: 9 and / or a light chain variable region that comprises, consists essentially of, or consists of the amino acid sequence of SEQ ID NO: 10.

[0088] Representative anti-AT antibody sequences are provided below. Additional anti-AT antibodies are provided, for example, in U.S. Patent No. 9,527,905, which is incorporated herein by reference. In certain examples, the antibodies or antigen-binding fragments thereof described herein bind to AT and have six CDRs of the antibodies listed in the two tables below (i.e., the three VH CDRs of the antibodies listed in the first table and the three VL CDRs of the same antibodies listed in the second table).

[0089] The anti-AT antibody MEDI4893 (also known as subratoxumab) is a human monoclonal antibody with a long half-life that binds with high affinity to AT and effectively blocks AT pore formation in target cell membranes. The half-life extension was achieved by introducing three amino acid substitutions (M252Y / S254T / T256E; termed YTE) in the Fc domain to increase binding to the neonatal Fc receptor (FcRn) and consequently increase serum half-life. The same version of MEDI4893 is designated "MEDI4893" because the YTE mutations reduce antibody half-life and exposure in mice. * Except for the YTE modification, referred to as "LC10," MEDI4893 (or subratoxumab) has been utilized to demonstrate efficacy in preclinical model studies previously described in WO 2012 / 109285 and WO 2014 / 074540, both of which are incorporated by reference herein in their entireties. MEDI4893 (or subratoxumab) contains a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 9 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 10. MEDI4893 (subratoxumab) and MEDI4893 * The identical CDR, VH and VL sequences are provided in the table below.

[0090] [Table 2]

[0091] In certain examples, the antibodies or antigen-binding fragments thereof described herein bind to AT and comprise the VH of an antibody listed in the table below in combination with, for example, a VL.

[0092] [Table 3]

[0093] In certain examples, the antibodies or antigen-binding fragments thereof described herein bind to AT and comprise the VL of an antibody listed in the table below, e.g., in combination with a VH, optionally in combination with a VH of the same antibody listed in the previous table.

[0094] [Table 4]

[0095] In certain examples, the antibodies or antigen-binding fragments thereof described herein bind to AT and comprise a heavy chain of an antibody listed in the table below, in combination with, for example, a light chain.

[0096] [Table 5]

[0097] In certain examples, the antibodies or antigen-binding fragments thereof described herein bind to AT and comprise a light chain of an antibody listed in the table below, e.g., in combination with a heavy chain, optionally a heavy chain of the same antibody listed in the previous table.

[0098] [Table 6]

[0099] In certain embodiments, the CDRs of an antibody or antigen-binding fragment thereof can be determined according to the Chothia numbering scheme, which refers to the location of immunoglobulin structural loops (see, e.g., Chothia C & Lesk AM, (1987), J Mol Biol 196:901-917; Al-Lazikani B et al., (1997) J Mol Biol 273:927-948; Chothia C et al., (1992) J Mol Biol 227:799-817; Tramontano A et al., (1990) J Mol Biol 215(1):175-82; and U.S. Pat. No. 7,709,226). Typically, when using the Kabat numbering convention, a Chothia CDR-H1 loop is located at heavy chain amino acids 26-32, 33 or 34, a Chothia CDR-H2 loop is located at heavy chain amino acids 52-56 and a Chothia CDR-H3 loop is located at heavy chain amino acids 95-102, while a Chothia CDR-L1 loop is located at light chain amino acids 24-34, a Chothia CDR-L2 loop is located at light chain amino acids 50-56 and a Chothia CDR-L3 loop is located at light chain amino acids 89-97. The ends of the Chothia CDR-H1 loop when numbered using the Kabat numbering convention vary from H32 to H34 depending on the length of the loop (this is because the Kabat numbering scheme places insertions at H35A and H35B; if neither 35A nor 35B are present, the loop ends at 32, if only 35A is present, the loop ends at 33, and if both 35A and 35B are present, the loop ends at 34).

[0100] In certain aspects, provided herein is a combination of antibodies and antigen-binding fragments thereof that comprise the Chothia VH and VL CDRs of the MEDI4893 antibody. In certain embodiments, the antibodies or antigen-binding fragments thereof comprise one or more CDRs, wherein the Chothia CDRs and the Kabat CDRs have identical amino acid sequences. In certain embodiments, provided herein is an antibody or antigen-binding fragment thereof that comprises a combination of Kabat and Chothia CDRs.

[0101] In certain embodiments, the CDRs of an antibody or antigen-binding fragment thereof can be determined according to the IMGT numbering system as described in Lefranc MP, (1999) The Immunologist 7:132-136 and Lefranc MP et al., (1999) Nucleic Acids Res 27:209-212. According to the IMGT numbering scheme, VH-CDR1 is at positions 26-35, VH-CDR2 is at positions 51-57, VH-CDR3 is at positions 93-102, VL-CDR1 is at positions 27-32, VL-CDR2 is at positions 50-52, and VL-CDR3 is at positions 89-97. In certain embodiments, provided herein are combinations of antibodies and antigen-binding fragments thereof comprising the IMGT VH and VL CDRs of MEDI4893, e.g., as described in Lefranc MP (1999) supra and Lefranc MP et al., (1999) supra).

[0102] In certain aspects, the CDRs of an antibody or antigen-binding fragment thereof may be determined according to MacCallum RM et al., (1996) J Mol Biol 262:732-745. See also, e.g., Martin A. "Protein Sequence and Structure Analysis of Antibody Variable Domains," in Antibody Engineering, Kontermann and Duebel, eds., Chapter 31, pp. 422-439, Springer-Verlag, Berlin (2001). In certain embodiments, provided herein are combinations of antibodies or antigen-binding fragments thereof that comprise the VH and VL CDRs of the MEDI4893 antibody determined by the method in MacCallum RM et al.

[0103] In certain aspects, the CDRs of an antibody or antigen-binding fragment thereof can be determined according to the AbM numbering scheme, which represents a compromise between the Kabat CDRs and Chothia structural loops and refers to the AbM hypervariable regions used by Oxford Molecular's AbM antibody modeling software (Oxford Molecular Group, Inc.). In certain embodiments, provided herein is a combination of an antibody or antigen-binding fragment comprising the VH and VL CDRs of the MEDI4893 antibody as determined by the AbM numbering scheme.

[0104] In another aspect, the antibodies or antigen-binding fragments thereof (e.g., monoclonal antibodies or fragments) described herein can comprise a constant region (Fc) of any suitable class (e.g., IgG, IgA, IgD, IgM, and IgE) that has been modified to improve the half-life of the antibody or antigen-binding fragment (e.g., monoclonal antibody or fragment). For example, the antibodies or antigen-binding fragments thereof (e.g., monoclonal antibodies or fragments) described herein can comprise an Fc that includes a mutation that extends the half-life relative to the same antibody that is not mutated.

[0105] Genetic engineering of Fc regions has been widely used in the art to extend the half-life of therapeutic antibodies and protect them from in vivo degradation. In some embodiments, the Fc region of an IgG antibody or antigen-binding fragment may be modified to increase the affinity of the IgG molecule to the fetal Fc receptor (FcRn), which mediates IgG catabolism and protects the IgG molecule from degradation. Suitable Fc region amino acid substitutions or modifications are known in the art, including, for example, the triple substitution M252Y / S254T / T256E (referred to as "YTE") (see, for example, U.S. Pat. No. 7,658,921; U.S. Patent Application Publication No. 2014 / 0302058; and Yu et al., Antimicrob. Agents Chemother., 61(1):e01020-16 (2017)). In certain embodiments, an antibody or antigen-binding fragment (eg, a monoclonal antibody or fragment) that binds to S. aureus AT comprises an Fc region that includes a YTE mutation.

[0106] The antibodies or antigen-binding fragments (e.g., monoclonal antibodies or fragments) described herein may be or may be derived from human, humanized, non-human, or chimeric antibodies. In one aspect, the antibodies or antigen-binding fragments thereof described herein are fully human antibodies.

[0107] Human, non-human, chimeric or humanized antibodies can be obtained by any means, including in vitro sources (e.g., hybridomas or cell lines recombinantly producing antibodies) and in vivo sources (e.g., rodents, human tonsils). Methods of producing antibodies are known in the art and are described, for example, in Koehler and Milstein, Eur. J. Immunol., 5:511-519 (1976); Harlow and Lane (eds.), Antibodies: A Laboratory Manual, CSH Press (1988); and Janeway et al. (eds.), Immunobiology, 5th Ed., Garland Publishing, New York, NY (2001)). In certain embodiments, human or chimeric antibodies can be produced using transgenic animals (e.g., mice) in which one or more endogenous immunoglobulin genes have been replaced with one or more human immunoglobulin genes. Examples of transgenic mice in which endogenous antibody genes have been effectively replaced with human antibody genes include, but are not limited to, the Medarex HUMAB-MOUSE™, the Kirin TC MOUSE™, and the Kyowa Kirin KM-MOUSE™ (see, e.g., Lonberg, Nat. Biotechnol., 23(9):1117-25 (2005), and Lonberg, Handb. Exp. Pharmacol., 181:69-97 (2008)). Humanized antibodies can be produced using any suitable method known in the art (see, e.g., An, Z. (ed.), Therapeutic Monoclonal Antibodies: From Bench to Clinic, John Wiley & Sons, Inc., Hoboken, NJ (2009)), including, for example, the grafting of non-human CDRs into a human antibody framework (see, e.g., Kashmiri et al., Methods, 36(1):25-34 (2005); and Hou et al., J. Biochem., 144(1):115-120 (2008)).In one embodiment, humanized antibodies can be generated using the methods described, for example, in US Patent Application Publication No. 2011 / 0287485A1.

[0108] III. Nucleic Acids, Vectors and Host Cells Also provided herein are one or more isolated nucleic acid sequences encoding an antibody or antigen-binding fragment thereof that binds to AT (optionally, the antibody or antigen-binding fragment thereof is a monoclonal antibody or fragment).

[0109] The disclosure further provides one or more vectors comprising one or more nucleic acid sequences encoding an antibody or antigen-binding fragment thereof that binds to AT (optionally, the antibody or antigen-binding fragment thereof is a monoclonal antibody or fragment). The vector can be, for example, a plasmid, an episome, a cosmid, a viral vector (e.g., a retrovirus or adenovirus), or a phage.

[0110] A vector containing a nucleic acid sequence encoding an antibody or antigen-binding fragment thereof that binds to AT (optionally, the antibody or antigen-binding fragment thereof is a monoclonal antibody or fragment) can be introduced into a host cell capable of expressing the encoded polypeptide, including any suitable prokaryotic or eukaryotic cell. Thus, the present disclosure provides an isolated cell containing the vector. Host cells that can be used include cells that can be grown easily and reliably, have a reasonably fast growth rate, have a well-characterized expression system, and can be easily and efficiently transformed or transfected.

[0111] A nucleic acid sequence encoding the amino acids of any of the antibodies or antigen-binding fragments described herein (optionally, monoclonal antibodies or fragments) can be introduced into a cell by "transfection," "transformation," or "transduction."

[0112] IV. Pharmaceutical Compositions and Methods of Administering Anti-AT Antibodies The disclosure provides compositions comprising an effective amount of any of the AT antibodies or antigen-binding fragments thereof described herein and a pharma- ceutically acceptable carrier, which amount can be effective to reduce the risk of infection in a subject colonized with S. aureus.

[0113] In another embodiment, the composition may comprise a nucleic acid sequence encoding an AT-binding antibody or antigen-binding fragment. The nucleic acid sequence may be present in a vector or a combination of vectors.

[0114] In one aspect, the composition is a pharma- ceutically acceptable (e.g., physiologically acceptable) composition comprising a carrier, e.g., a pharma- ceutically acceptable (e.g., physiologically acceptable) carrier and an AT-binding antibody or antigen-binding fragment nucleic acid sequence or vector.

[0115] Any suitable carrier can be used within the context of the present disclosure, and such carriers are well known in the art. The choice of carrier will be determined in part by the intended site to which the composition can be administered and the intended method used to administer the composition. The composition can be optionally sterile. The composition can be frozen or lyophilized for storage, and can be reconstituted with suitable sterile carrier before use. The composition can be produced according to conventional techniques, for example, as described in Remington: The Science and Practice of Pharmacy, 21st Edition, Lippincott Williams & Wilkins, Philadelphia, PA (2001).

[0116] The compositions desirably contain an AT-binding antibody or antigen-binding fragment in an amount effective to reduce the risk of S. aureus infection in a patient colonized with S. aureus. To this end, the methods of the disclosure comprise administering a therapeutically or prophylactically effective amount of an AT-binding antibody or antigen-binding fragment thereof, or a composition of the above-described antibodies or fragments (including monoclonal antibodies or fragments).

[0117] In the case of repeated administration over several days or more depending on the condition, the treatment can be repeated until the desired suppression of disease symptoms occurs. However, other administration regimes may be useful and are included within the scope of the present disclosure. The desired dosage can be delivered by administering a single bolus of the composition, multiple boluses of the composition, or continuous infusion of the composition.

[0118] An effective amount of an anti-AT antibody or antigen-binding fragment thereof can be administered to a subject, such as a human, using standard administration techniques, including intravenous, intraperitoneal, subcutaneous, and intramuscular routes of administration. The anti-AT antibody or antigen-binding fragment thereof can be suitable for parenteral administration. The term "parenteral" as used herein includes intravenous, intramuscular, subcutaneous, and intraperitoneal administration. In some embodiments, the anti-AT antibody or antigen-binding fragment thereof is administered to a subject using peripheral systemic delivery by intravenous, intraperitoneal, or subcutaneous injection.

[0119] The AT-binding antibody or antigen-binding fragment or composition comprising the same may be administered alone or in combination with other drugs (e.g., adjuvants) conventionally used to treat S. aureus infections. The AT-binding antibody or antigen-binding fragment comprising the composition may be used in combination with one or more antibiotics, such as penicillinase-resistant β-lactam antibiotics (e.g., oxacillin or flucloxacillin). Gentamicin can be used to treat serious infections, such as endocarditis. However, most strains of S. aureus are now resistant to penicillin, and two out of every 100 people carry methicillin-resistant S. aureus (MRSA). MRSA infections are typically treated with vancomycin, and mild skin infections can be treated with a triple antibiotic ointment.

[0120] Compositions containing AT-binding antibodies or antigen-binding fragments can be used, for example, in combination with one or more anti-Staphylococcus aureus antibiotics.

[0121] V. Methods for Identifying Patients Who Would Benefit from Anti-AT Antibodies As demonstrated herein, administration of an anti-AT antibody or antigen-binding fragment thereof to a subject colonized with low levels of S. aureus, e.g., levels of S. aureus that do not exceed a threshold level of S. aureus, can reduce the incidence of infection associated with the presence of S. aureus in the subject.

[0122] The amount of S. aureus in a subject can be determined based on the amount of S. aureus in a sample obtained from the subject. The sample can be, for example, a skin or soft tissue sample. The sample can be, for example, obtained from the lower respiratory tract of the subject. The sample can be, for example, an endotracheal aspirate, a tracheal sample, or a bronchial sample.

[0123] In certain embodiments, an anti-AT antibody or antigen-binding fragment thereof is administered to a subject, wherein a sample obtained from the subject has a concentration of S. aureus that does not exceed a predetermined threshold estimated to be 3.2 log10 CFU / mL (approximately 1,600-1,700 CFU / mL).

[0124] The amount of S. aureus in a sample obtained from a subject can be quantified using polymerase chain reaction (PCR). For example, the amount of S. aureus in a sample obtained from a subject can be a quantifiable number based on the number of PCR cycles required to reach a threshold signal, referred to herein as "cycle threshold" or "Ct value". A high Ct value (i.e., a large number of PCR cycles required to reach a threshold signal) indicates a low level of S. aureus, while a low Ct value (i.e., a small number of PCR cycles required to reach a threshold signal) indicates a high level of S. aureus.

[0125] PCR is a particularly useful method for quantifying the amount of S. aureus in a sample because it can be performed quickly and uniformly. For example, PCR can detect the amount of S. aureus in a sample within two hours. Laboratories vary greatly in their methods for bacterial culture, but laboratories will perform PCR using the same type of equipment which creates uniform PCR output across different laboratories.

[0126] PCR can be used to determine the amount of S. aureus in a sample based on the amplification of a single S. aureus gene or a combination of S. aureus genes. For example, PCR can be used to detect the S. aureus protein A gene. Detection of S. aureus protein A detects all S. aureus, regardless of their methicillin susceptibility. However, PCR can also be used to determine whether S. aureus is antibiotic resistant, e.g., methicillin resistant. For example, PCR can be used to detect the presence of the methicillin resistance determinant (mecA) and the staphylococcal chromosomal cassette (SCCmec). Detection of both mecA and SCCmec indicates that S. aureus is methicillin resistant.

[0127] In certain embodiments, an anti-AT antibody or antigen-binding fragment thereof is administered to a subject, where a sample obtained from the subject has a level of S. aureus that does not exceed the highest level of S. aureus that correlates with a PCR Ct value. In certain embodiments, an anti-AT antibody or antigen-binding fragment thereof is administered to a subject, where a sample obtained from the subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with a PCR Ct value of about 29. In certain embodiments, an anti-AT antibody or antigen-binding fragment thereof is administered to a subject, where a sample obtained from the subject has a level of S. aureus that does not exceed a level of S. aureus that correlates with a PCR Ct value of 29-36.

[0128] In certain embodiments, an anti-AT antibody or antigen-binding fragment thereof is administered to a subject, where a sample obtained from the subject has a level of S. aureus that is at least at the lowest level of S. aureus that correlates with a PCR Ct value of about 3 to about 29. In certain embodiments, an anti-AT antibody or antigen-binding fragment thereof is administered to a subject, where a sample obtained from the subject has a level of S. aureus that correlates with a PCR Ct value of 3 to 29.

[0129] A PCR Ct value of 29 is estimated to correspond to a concentration of approximately 1,600-1,700 colony forming units (CFU) / mL of S. aureus.

[0130] The method for identifying subjects who would benefit from receiving anti-AT antibodies or antigen-binding fragments thereof provided herein may be particularly useful in subjects on mechanical ventilation. Thus, the subject may be a subject on a mechanical ventilation. Although the overall disease incidence is fairly low at about 1-2%, the incidence of Staphylococcus aureus pneumonia is much higher (>20%) in patients colonized with Staphylococcus aureus. In subjects on mechanical ventilation, Staphylococcus aureus pneumonia is an early event that generally occurs within the first week after initiation of mechanical ventilation, so earlier administration of anti-AT antibodies or antigen-binding fragments thereof may be crucial in avoiding pneumonia. PCR-based techniques for assessing the level of S. aureus in patients can be completed within about 2 hours, whereas culture techniques are much slower (and further lack the precise quantitative nature of PCR-based techniques).

[0131] PCR-based techniques are also advantageous compared to culture-based techniques for S. aureus colonization because PCR-based techniques may be more sensitive in detecting S. aureus, especially in subjects taking antibiotics. Thus, the subject may take antibiotics. Furthermore, in certain embodiments of the methods provided herein, PCR analysis of a sample obtained from a subject indicates that the subject would benefit from taking an anti-AT antibody or antigen-binding fragment thereof, while the sample does not contain bacteria that would grow in a culture assay used to determine the presence of S. aureus.

[0132] The following examples further illustrate the invention but, of course, should not be construed as in any way limiting its scope. EXAMPLES

[0133] Example 1 Staphylococcus aureus (S. aureus) pneumonia in mechanically ventilated patients is an early event that typically occurs within the first week after mechanical ventilation. Thus, rapid identification of patients at risk for developing this infection can be lifesaving. Although the overall disease incidence is fairly low, approximately 1-2%, the incidence is much higher (>20%) in patients colonized with S. aureus.

[0134] Rapid polymerase chain reaction (PCR) was analyzed as an assay to identify patients at risk for colonization with Staphylococcus aureus (S. aureus) in the lower respiratory tract (LRT) in a phase 2 clinical trial of mechanically ventilated patients in an intensive care unit (ICU). The Xpert® MRSA / SA SSTI assay (Cepheid, Sunnyvale, CA) was used according to the product protocol for performing PCR. The assay is a rapid, fully automated DNA test for the simultaneous detection of Methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus aureus (SA) directly from skin and soft tissue specimens. Specimens are collected on duplicate swabs that are placed into tubes containing elution reagent. After a brief vortex mixer run, the elution material and two single-use reagents (Reagent 1 and Reagent 2) provided with the assay are transferred to the various uniquely labeled chambers of a disposable fluidic cartridge (Xpert MRSA / SA cartridge). The cartridge is placed into the GeneExpert® DX System instrument platform, which performs automated real-time multiplex PCR to detect DNA, where additional sample preparation, amplification and real-time detection are fully automated and fully integrated.

[0135] The primers and probes in the Xpert MRSA / SA assay detect nucleic acid sequences of S. aureus protein A (spa), the gene for MecA-mediated oxacillin resistance (mecA) and the Staphylococcus aureus cassette chromosome (SCCmec) inserted into the SA chromosome attB site. The assay includes sample processing controls to control proper processing of the target bacteria and to monitor the presence of inhibitors in the PCR assay. The Probe Check control verifies rehydration of reagents, PCR tube filling in the cartridge, probe integrity and dose stability.

[0136] The assay is depicted in Figure 1: An endotracheal aspirate (ETA) sample was obtained from the patient and a swab of the sample was inserted into the elution reagent. The mixture was vortexed and dispensed into the cartridge. The cartridge was then inserted into the PCR instrument to begin the test. The total test run time is less than 2 minutes, with the entire test being completed within 75 minutes.

[0137] PCR tests generate a cycle threshold (Ct) value, which represents the number of PCR cycles required to reach a threshold signal. The Ct value is inversely proportional to the bacterial load: the more S. aureus in a sample, the fewer cycles required to reach the threshold level and therefore have a lower Ct value, whereas the less S. aureus in a sample, the more cycles required to reach the threshold level and therefore have a higher Ct value.

[0138] Example 2 The rapid PCR test discussed in Example 1 was used to detect the presence or absence of S. aureus and determine if the S. aureus is methicillin-resistant S. aureus (MRSA) by looking at three molecular targets. The first target detects S. aureus protein A (SPA), which detects all S. aureus, regardless of their methicillin susceptibility status. The second and third targets are the methicillin-resistance determinant determinant (mecA) and the S. aureus chromosomal cassette (SCCmec), which is used to detect MRSA.

[0139] Rapid PCR testing was performed on 720 patients at screening in the Phase 2 clinical trial. Of these patients, 299 (41.5%) were colonized with S. aureus. This number is higher than the S. aureus colonization rate in the general public (25-30%), but is reasonable for mechanically ventilated patients in an ICU. Of the 299 patients colonized with S. aureus, 277 (92.6%) had methicillin-susceptible S. aureus (MSSA) and 22 (7.4%) had MRSA.

[0140] Ct values ​​for 295 of 299 patients colonized with S. aureus are shown in Figure 2. (Patients with 4 PCR + zero or missing Ct values ​​are not shown.) The average Ct value in these patients was 25.7, with a substantial number of patients having low Ct values ​​(high bacterial load) at screening.

[0141] Screening was performed at various time points after initiation of mechanical ventilation. Thus, Ct values ​​were compared with the number of days on mechanical ventilation. The results shown in Figure 3 demonstrate that Ct values ​​have a wide distribution independent of the number of days on mechanical ventilation. Thus, the number of days on mechanical ventilation could not be used to predict S. aureus colonization or Ct values.

[0142] Example 3 In this example, we demonstrate that rapid PCR is more sensitive than culture assays in detecting S. aureus colonization. Culture assays usually involve plating or streaking either undiluted samples or serial dilutions on agar containing plates and vary widely among different laboratories. After incubating the plates for 24-48 hours, either the scattered colonies of S. aureus were counted (quantitative culture methods) or the number of quadrants of S. aureus growth and growth density (semi-quantitative culture methods) were assessed. Qualitative cultures do not provide quantification and only assess the presence or absence of S. aureus. (See FIG. 4.) Thus, when different culture methods are used, the status of the culture (positive or negative for S. aureus infection) may differ and may adversely bias treatment regimens if not accurate.

[0143] Cultures were performed on a randomized subgroup of 299 patients (N=209). Only 162 of these 209 patients (77.5%) produced positive culture results, while 47 (22.5%) produced negative culture results. Thus, the culture assay would miss 22.5% of subjects colonized with S. aureus. In other words, there was a 77.5% concordance rate and a 22.5% discordance rate between the rapid PCR test and the culture assay. All of this discordance was due to samples that tested positive by rapid PCR and negative by culture. (See Figure 6.)

[0144] Further inspection of the results demonstrated that a significantly higher percentage of patients with samples that tested positive by PCR and negative by culture were using antibiotics concomitantly than patients with samples that tested positive by PCR and culture. (See Figure 7.) These data suggest that antibiotic use negatively impacts culture results. However, prior antibiotic use had no impact on culture results. (See Figure 8.)

[0145] Furthermore, the sensitivity of the culture assay varied due to differences in plating (e.g., sample dilution, plating volume, and plate number) among the various laboratories. The laboratory surveys are summarized in the table below.

[0146] [Table 7]

[0147] Among these nine laboratories, the detection limits ranged from 3.3 to 10 5 CFU / mL. Thus, low culture sensitivity in some laboratories further contributed to the discrepancy between PCR and culture assays.

[0148] The Ct cutoff values ​​for whether a culture was classified as mild, moderate, or severe were examined. The majority of cultures classified as severe fell below a PCR Ct value of 29. However, a significant portion of cultures classified as moderate also fell below a PCR Ct value of 29. In fact, even some cultures classified as mild also fell below a PCR Ct value of 29. Thus, the assessment of these cultures was inconsistent. (See FIG. 5.)

[0149] Methicillin susceptibility determinations were concordant using either the rapid PCR or culture assay.Of 162 samples tested by both PCR and culture, 9 (5.6%) were identified as MRSA and 153 (94.4%) were identified as MSSA, regardless of method.

[0150] These results demonstrate that the culture and PCR assays are equally effective in detecting methicillin susceptibility, but the PCR assay is more sensitive for S. aureus colonization.

[0151] Example 4 Ct values ​​were analyzed in culture positive and culture negative samples. The distribution of culture positive and culture negative samples by Ct value is shown in Figure 9 and the results are also summarized in the table below.

[0152] [Table 8]

[0153] Based on these results, a Ct value of 29 does the best job of discriminating between culture-positive and culture-negative samples.

[0154] The Ct value also correlated with the S. aureus concentration. The results shown in FIG. 10 demonstrate that there is a statistically significant inverse linear correlation between the Ct value and the number of CFU / mL. The majority of samples with Ct values ​​greater than 29 had low S. aureus loads (≦10 3 CFU / mL), and the majority of samples with Ct values ​​below 29 had high S. aureus loads (>10 4 The Ct value of 29 corresponded to approximately 3.2 log10 CFU / mL (approximately 1,600 to 1,700 CFU / mL).

[0155] A similar correlation between Ct values ​​and S. aureus concentrations was detected in bronchial and tracheal cultures (see FIG. 11).

[0156] A similar correlation between Ct value and S. aureus concentration was also detected whether non-Staphylococcus growth was observed or not (see FIG. 12).

[0157] These results demonstrate that PCR is a robust method for quantifying S. aureus colonization and that a Ct cutoff value of 29 effectively discriminated between low and high S. aureus colonization.

[0158] Example 5 The efficacy of MEDI4893, an anti-S. aureus antibody, in preventing S. aureus pneumonia in mechanically ventilated ICU patients was compared in patients with low and high levels of S. aureus colonization using a Ct cutoff value of 29.

[0159] Subjects were considered to be mechanically ventilated if they (i) had an endotracheal or nasotracheal tube intubated and were receiving positive pressure ventilatory support, or (ii) were not intubated with either an endotracheal or nasotracheal tube but had required positive pressure ventilation for 8 hours or more within the last 24 hours (e.g., patients with tracheostomy, continuous positive airway pressure [CPAP], etc.).

[0160] S. aureus pneumonia was diagnosed in mechanically ventilated patients who met the following radiographic, clinical, and microbiological criteria that were not attributable to any apparent noninfectious cause at the time of diagnosis:

[0161] Radiographic criteria: new or worsening infiltrate consistent with pneumonia on a chest x-ray obtained within 24 hours of the event (diagnosed by a board-certified radiologist), and

[0162] Clinical criteria: At least two of the following minor or one major respiratory sign or initial symptom: ·Small criteria: Systemic signs of infection (one or more of the following): Abnormal body temperature (oral or tympanic temperature >38°C, or core temperature ≥38.3°C, or hypothermia defined as a core temperature <35°C) and / or abnormal white blood cell count (WBC count >10,000 cells / mm3, WBC count <4,500 cells / mm3, or >15% band neutrophils) Production of purulent intratracheal secretions New physical exam findings consistent with pneumonia / cirrhosis (e.g., crackles, rhonchi, bronchial breath sounds), dullness to percussion Major criteria: Acute changes in the ventilatory support system to improve oxygen delivery, determined by: A PaO2 / FiO2 ratio of <240 mmHg maintained for at least 4 hours, or A reduction in PaO2 / FiO2 of ≥ 50mmHg that is sustained for at least 4 hours and

[0163] Microbiological confirmation (obtained within 24 hours after onset of the event) of at least one of the following: Respiratory specimens positive for S. aureus by culture. Includes respiratory secretion specimens obtained by endotracheal aspirate in intubated subjects or by bronchoscopy with bronchoalveolar lavage (BAL) or protected specimen brush (PSB) sampling. Sputum specimens were acceptable in subjects who were not intubated but met the protocol definition of mechanical ventilation. Blood cultures positive for S. aureus (and no obvious primary source of infection outside the lungs) Positive pleural fluid aspirate or lung tissue culture for S. aureus during an episode of pneumonia

[0164] The results are shown in the table below.

[0165] [Table 9]

[0166] Relative risk reduction (5,000 mg MEDI4893 vs. placebo; 90% confidence intervals (CI) and p-values ​​based on Poisson regression with robust variance.

[0167] An 82.6% relative risk reduction of S. aureus isolated pneumonia (90% CI: -1.0%, 97.0%), a 30.6% relative risk reduction of all-cause pneumonia (90% CI: -4.9%, 54.0%), and a 23.1% relative risk reduction of all-cause pneumonia or death (90% CI: 23.1% to 4.9%, 43.6%) were also observed.

[0168] [Table 10]

[0169] P-values ​​for interactions are derived from Poisson regression analysis with robust variance, including treatment group, subgroup under study, and treatment by subgroup interactions. Relative risk reductions (5,000 mg MEDI4893 vs. placebo) and 90% confidence intervals (CIs) were based on unconditional confidence intervals for proportions.

[0170] Patients with a Ct value of at least 29 (low S. aureus) had an attack rate of 33.3% and MEDI4893 produced a statistically significant relative reduced risk (RRR) of approximately 67%. In contrast, the total population had an attack rate of 26% and MEDI4893 produced a statistically insignificant RRR of 32%. Patients with a Ct value less than 29 (high S. aureus) had an attack rate of 22.2% and a statistically insignificant RRR of approximately 2.5%.

[0171] These results demonstrate that MEDI4893, an anti-alpha toxin antibody, showed increased efficacy in patients with low S. aureus colonization.

[0172] Example 6 The effect of anti-S. aureus antibodies on savings in healthcare resource utilization was also analyzed. The main results are summarized in the table below.

[0173] [Table 11]

[0174] These results demonstrate that MEDI4893 reduced length of hospital stay, ICU stay, and length of time on mechanical ventilation.

[0175] All references cited in this specification, including publications, patent applications, and patents, are hereby incorporated by reference to the same extent as if each reference was individually and specifically indicated to be incorporated by reference and was set forth in its entirety herein.

[0176] The recitation of ranges of values ​​herein is merely intended to serve as a shorthand method of individually referring to each individual value within the range, unless otherwise indicated herein, and each individual value is incorporated herein as if it were individually recited herein. All methods described herein can be performed in any suitable order, unless otherwise indicated herein or otherwise clearly contradicted by context. Any and all examples provided herein, or the use of exemplary language (e.g., "such as"), are merely intended to further clarify the invention, and do not pose limitations on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0177] Preferred embodiments of the invention are described herein, including the best mode known to the inventors for carrying out the invention. Variations of those preferred embodiments may become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventors expect that such variations will be employed by those of ordinary skill in the art as appropriate, and the inventors do not intend to practice the invention otherwise than as specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

Claims

1. 1. A pharmaceutical composition for treating or preventing Staphylococcus aureus (S. aureus) infection in a subject colonized with S. aureus, wherein the pharmaceutical composition comprises an antibody or antigen-binding fragment thereof that binds to S. aureus alpha-toxin (AT), and wherein polymerase chain reaction (PCR) is used to detect levels of S. aureus in a sample obtained from the subject.

2. 1. A pharmaceutical composition for reducing the incidence of Staphylococcus aureus (S. aureus) pneumonia in a subject, said pharmaceutical composition comprising subratoxumab, said reduction being determined by clinical, microbiological and radiographic means.

3. The pharmaceutical composition of claim 2, wherein the incidence is reduced by about 30%.

4. 1. A pharmaceutical composition for reducing the incidence of pneumonia of any cause in a subject, said pharmaceutical composition comprising subratoxumab, said reduction being determined by clinical, microbiological and radiographic means.

5. The pharmaceutical composition of claim 4, wherein the incidence is reduced by about 30%.

6. 6. The pharmaceutical composition of any one of claims 2 to 5, wherein (a) the clinical measures include abnormal body temperature, abnormal white blood cell count, cough, purulent sputum, bronchial breath sounds, dyspnea, tachypnea (respiratory rate >30 breaths / min), hypoxia, or any combination thereof; (b) the microbiological measures include a respiratory specimen, blood culture, pleural fluid aspirate, or lung tissue culture that is positive for S. aureus; and / or (c) the radiographic measures include a new or worsening infiltrate on a chest radiograph.

7. The pharmaceutical composition of any one of claims 2 to 5, wherein PCR is used to detect the level of S. aureus in a sample obtained from the subject.

8. A pharmaceutical composition for treating or preventing a Staphylococcus aureus (S. aureus) infection in a subject colonized with the infection, wherein the pharmaceutical composition comprises an antibody or antigen-binding fragment thereof that binds to S. aureus AT, and wherein a sample obtained from the subject has a concentration of S. aureus not exceeding 1,700 CFU / mL.

9. The pharmaceutical composition of claim 1 or 8, wherein the antibody or antigen-binding fragment thereof that binds to Staphylococcus aureus (S. aureus) AT comprises a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO:

8.

10. An in vitro method for identifying a subject colonized with Staphylococcus aureus (S. aureus) that is responsive to an antibody or antigen-binding fragment thereof that binds to S. aureus AT, the method comprising a step of detecting the level of S. aureus in a sample obtained from the subject, wherein a level of S. aureus that does not exceed a level of S. aureus that correlates with a PCR Ct value is the criterion for determining that the subject is responsive to the antibody or antigen-binding fragment thereof.

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