Antibodies to respiratory syncytial virus and other paramyxoviruses and methods of use thereof

Abstract

The present disclosure provides an antibody that can bind to a paramyxovirus and neutralize infection by the paramyxovirus. The paramyxovirus can be, for example, a respiratory syncytial virus, a metapneumovirus, or a pneumonia virus of mice. The antibody comprises a modification of the Fc region that improves the in vivo stability of the antibody, one or more effector functions of the antibody, or both. Also provided are antibody compositions, polynucleotides encoding the antibody, vectors, host cells, and methods of using the antibody for preventing and / or treating paramyxovirus infection.

Description

[Technical field]

[0001] SEQUENCE LISTING STATEMENT The sequence listing associated with this application is provided in text form in lieu of a paper copy and is incorporated herein by reference. The name of the text file containing the sequence listing is 930485_434WO_SEQUENCE_LISTING.txt. The text file is 29.7 KB, was created on February 7, 2022, and has been submitted electronically via EFS-Web. [Background technology]

[0002] background There is a need for means to prevent or treat infections by paramyxoviruses such as respiratory syncytial virus, metapneumovirus, or pneumonia virus of mice. [Brief description of the drawings]

[0003] [Figure 1] Figure 1 shows the design of a preclinical study evaluating the in vivo stability of human IgG1 antibodies RSV_Ab (HC: SEQ ID NO: 5; LC: SEQ ID NO: 4) and RSV_Ab_MLNS (HC: SEQ ID NO: 1; LC: SEQ ID NO: 4) in cynomolgus monkeys. Stability was determined by correlating neutralizing activity with total human IgG1 concentration in plasma (using a mouse monoclonal antibody (mAb) specific for human IgG1 CH2). [Diagram 2] Figure 2 shows the results of the preclinical study shown in Figure 1. Pharmacokinetics (PK) was calculated as the concentration of antibody in plasma captured by the coated Ds-Cav1F antigen, measured by ELISA. [Diagram 3]Figure 3 shows further results of the preclinical study shown in Figure 1. Data show blood PK of RSV_Ab (top) and RSV_Ab_MLNS (bottom). In 2 of 3 animals receiving RSV_Ab_MLNS, PK continued until day 113. Quantification was based on DsCav1-based antigen capture ELISA (animals were pre-screened to be immune to RSV): (re)analysis was performed in parallel on all samples. There was no obvious anti-drug antibody (ADA) response in any of the 12 animals (3 of 9 animals received engineered anti-FluA antibodies).

[0004] Detailed Description Provided herein is an antibody that can bind to paramyxovirus and neutralize infection by paramyxovirus. Paramyxovirus can be, for example, respiratory syncytial virus, metapneumovirus, or pneumonia virus of mice. The antibody comprises a modification in Fc that improves the in vivo stability of the antibody, one or more effector functions of the antibody, or both.

[0005] In some embodiments, after a single administration of the antibody to a subject (e.g., administered by slow intravenous infusion), the antibody is present in the subject's plasma at a concentration equal to or greater than the neutralization EC50 (concentration of antibody required for half-maximal neutralization) of the antibody for up to 100, up to 200, up to 300, up to 400, up to 500, up to 600, up to 700, up to 800, up to 900, up to 1,000, up to 1,100, up to 1,200, or up to 1,300 hours or more. The neutralization EC50 can be informed by the results of an in vitro assay and / or an in vivo assay. In some embodiments, the antibody optionally comprises a neutralization EC50 against RSV of about 20 micrograms (e.g., 20 micrograms) per ml in an in vitro neutralization assay and / or in cynomolgus monkeys.

[0006] For example, in some embodiments, following a single administration of the antibody at 15 mg / kg to a cynomolgus monkey (e.g., administered by slow intravenous infusion), the antibody is present in the subject's plasma at a concentration equal to or greater than the neutralization EC50 for up to 100, up to 200, up to 300, up to 400, up to 500, up to 600, up to 700, up to 800, up to 900, up to 1,000, up to 1,100, up to 1,200, or up to 1,300 hours or more.

[0007] In some embodiments, following a single administration of the antibody at 15 mg / kg to cynomolgus monkeys (e.g., administered by slow intravenous infusion), the antibody is present in plasma at a concentration of 20 micrograms per ml or greater for a period of more than 1000 hours, more than 1,050 hours, more than 1,100 hours, more than 1,150 hours, more than 1,200 hours, more than 1,250 hours, more than 1300 hours, or more, or up to about 1,250 hours, or up to 1,300 hours, or up to 1,200 hours, or up to 1,100 hours, or up to 1,000 hours. In certain embodiments, the antibody optionally comprises a neutralizing EC50 against RSV of about 20 micrograms per ml (such as, for example, 20 micrograms) in an in vitro neutralization assay and / or in cynomolgus monkeys.

[0008] In some embodiments, following a single administration of the antibody at 15 mg / kg to cynomolgus monkeys (e.g., administered by slow intravenous infusion), the antibody is present in the plasma at a concentration of 20 micrograms per ml or greater for a period of more than 10 days, more than 20 days, more than 30 days, more than 40 days, more than 50 days, or up to 55 days, or up to 50 days, or up to 60 days. In certain embodiments, the antibody optionally comprises a neutralizing EC50 against RSV of about 20 micrograms per ml (such as, for example, 20 micrograms) in an in vitro neutralization assay and / or in cynomolgus monkeys. Summary of the Invention

[0009] In some embodiments, 113 days after a single administration of the antibody composition at 15 mg / kg of antibody to cynomolgus monkeys (e.g., administered by slow intravenous infusion), the antibody is present in the plasma at a concentration of between 2 micrograms per ml and 10 micrograms per ml, or between 2 micrograms per ml and 7 micrograms per ml, or about 2, about 3, about 4, about 5, about 6, or about 7 micrograms per ml.

[0010] In some embodiments, following a single administration of an antibody or antibody composition at 15 mg / kg to a cynomolgus monkey (e.g., administered by slow intravenous infusion), the antibody is present in plasma at any one or more concentrations at predetermined times according to Table A.

[0011] [Table 1-1] [Table 1-2]

[0012] The plasma concentration of antibodies can be quantified, for example, using DsCav1-based antigen capture ELISA. The presence of virus or viral load (e.g., RSV in cynomolgus monkeys) in a subject can be assessed using known techniques and samples such as, for example, nasal swabs.

[0013] In some embodiments, the antibody does not induce an anti-drug antibody (ADA) response in cynomolgus monkeys, for example, as measured 56 days after a single dose of 15 mg / kg antibody or antibody composition administered by slow intravenous infusion.

[0014] Also provided are polynucleotides encoding the antibody vectors, host cells, and related compositions, and methods of using the antibodies, nucleic acids, vectors, host cells, and related compositions to prevent or treat (e.g., reduce, delay, eliminate, or prevent) a paramyxovirus infection in a subject and / or in the manufacture of a medicament for preventing or treating a paramyxovirus infection in a subject.

[0015] Before describing this disclosure in more detail, it is helpful to an understanding of the disclosure to provide definitions of certain terms used herein. Additional definitions are set forth throughout the disclosure.

[0016] Any concentration range, percentage range, ratio range, or integer range herein should be understood to include any integer value within the recited range, and fractions thereof (such as tenths and hundredths of integers) where appropriate, unless otherwise indicated. Also, any numerical range described herein for any physical characteristic, such as polymer subunits, size, or thickness, should be understood to include any integer within the recited range, unless otherwise indicated. As used herein, the term "about" means ±20% of the indicated range, value, or structure, unless otherwise indicated. The terms "a" and "an" as used herein should be understood to refer to "one or more" of the recited components. The use of alternatives (e.g., "or") should be understood to mean either, both, or any combination of the alternatives. As used herein, the terms "include," "having," and "comprise" are used interchangeably, and it is intended that the terms and variations thereof be interpreted in a non-limiting manner.

[0017] "Desired" or "desirably" means that the subsequently described element, component, event, or circumstance may or may not occur, and that the description includes circumstances in which the element, component, event, or circumstance occurs as well as circumstances in which they do not occur.

[0018] Moreover, it should be understood that individual constructs or groups of constructs derived from various combinations of the structures and subunits described herein are disclosed by this application to the same extent as if each construct or group of constructs was described individually, and thus selection of a particular structure or particular subunits is within the scope of this disclosure.

[0019] The term "consisting essentially of" is not equivalent to "comprising" and refers to a particular material or step of a claim or to something that does not substantially affect the basic characteristics of the claimed subject matter. For example, a domain, region, or module (such as a binding domain) of a protein or protein "consists essentially of" a particular amino acid sequence if the amino acid sequence of the domain, region, module, or protein includes extensions, deletions, mutations, or combinations thereof (e.g., amino- or carboxy-terminal or interdomain amino acids) that, in combination, contribute up to 20% (e.g., up to 15%, 10%, 8%, 6%, 5%, 4%, 3%, 2%, or 1%) of the length of the domain, region, module, or protein and do not substantially affect (i.e., do not reduce activity by more than 50%, such as 40%, 30%, 25%, 20%, 15%, 10%, 5%, or 1% or less) the activity of the domain, region, module, or protein (e.g., target binding affinity of a binding protein).

[0020] As used herein, "amino acid" refers to natural and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to natural amino acids. Naturally occurring amino acids include those encoded by the genetic code, as well as those that are subsequently modified, such as hydroxyproline, γ-carboxyglutamic acid, and O-phosphoserine. Amino acid analogs refer to compounds that have the same basic chemical structure as naturally occurring amino acids, i.e., an α-carbon bonded to a hydrogen, a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, and methionine methylsulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as naturally occurring amino acids. Amino acid mimetics refer to compounds that have a structure that is different from the general chemical structure of an amino acid, but function in a manner similar to naturally occurring amino acids.

[0021] As used herein, "mutation" refers to a change in the sequence of a nucleic acid molecule or polypeptide molecule as compared to a reference or wild-type nucleic acid molecule or polypeptide molecule, respectively. Mutations can cause several different types of changes in the sequence, such as nucleotide or amino acid substitutions, insertions, deletions, etc.

[0022] "Conservative substitution" refers to an amino acid substitution that does not significantly affect or alter the binding characteristics of a particular protein. Generally, a conservative substitution is one in which the substituted amino acid residue is replaced with an amino acid residue having a similar side chain. Conservative substitutions include those found in any of the following groups: Group 1: Alanine (Ala or A), Glycine (Gly or G), Serine (Ser or S), Threonine (Thr or T); Group 2: Aspartic acid (Asp or D), Glutamic acid (Glu or Z); Group 3: Asparagine (Asn or N), Glutamine (Gln or Q); Group 4: Arginine (Arg or R), Lysine (Lys or K), Histidine (His or H); Group 5: Isoleucine (Ile or I), Leucine (Leu or L), Methionine (Met or M), Valine (Val or V); and Group 6: Phenylalanine (Phe or F), Tyrosine (Tyr or Y), Tryptophan (Trp or W). Additionally or alternatively, amino acids can be classified into conservative substitution groups by similar function, chemical structure, or composition (e.g., acidic, basic, aliphatic, aromatic, or sulfur-containing). For example, the aliphatic group may include, for substitution purposes, Gly, Ala, Val, Leu, and Ile. Other conservative substitution groups include: sulfur-containing: Met and cysteine ​​(Cys or C); acidic: Asp, Glu, Asn, and Gln; small aliphatic, non-polar or slightly polar residues: Ala, Ser, Thr, Pro, and Gly; polar negatively charged residues and their amides: Asp, Asn, Glu, and Gln; polar positively charged residues: His, Arg, and Lys; large aliphatic, non-polar residues: Met, Leu, Ile, Val, and Cys; and large aromatic residues: Phe, Tyr, and Trp. Additional information can be found in Creighton (1984) Proteins, WH Freeman and Company.

[0023] As used herein, "protein" or "polypeptide" refers to a polymer of amino acid residues. Protein applies to naturally occurring amino acid polymers, as well as to amino acid polymers in which one or more amino acid residues are artificial chemical mimetics of the corresponding naturally occurring amino acids, and to non-naturally occurring amino acid polymers.

[0024] "Nucleic acid molecule" or "polynucleotide" or "polynucleic acid" refers to a polymeric compound containing covalently linked nucleotides that can be composed of natural subunits (e.g., purine or pyrimidine bases) or non-natural subunits (e.g., morpholine rings). Purine bases include adenine, guanine, hypoxanthine, and xanthine, while pyrimidine bases include uracil, thymine, and cytosine. Nucleic acid molecules include polyribonucleic acid (RNA), including mRNA, microRNA, siRNA, viral genomic RNA, and synthetic RNA, and polydeoxyribonucleic acid (DNA), including cDNA, genomic DNA, and synthetic DNA, any of which may be single- or double-stranded. If single-stranded, the nucleic acid molecule may be the coding strand or the non-coding (antisense) strand. A nucleic acid molecule that codes for an amino acid sequence includes all nucleotide sequences that code for the same amino acid sequence. Some versions of a nucleotide sequence may contain introns to the extent that the introns are removed during transcription or by post-transcriptional mechanisms. In other words, different nucleotide sequences may encode the same amino acid sequence as a result of redundancy or degeneracy in the genetic code, or by splicing.

[0025] In some embodiments, the polynucleotide (e.g., mRNA) comprises a modified nucleoside, a cap-1 structure, a cap-2 structure, or any combination thereof. In certain embodiments, the polynucleotide comprises pseudouridine, N6-methyladenosine, 5-methylcytidine, 2-thiouridine, or any combination thereof. In some embodiments, the pseudouridine comprises N1-methylpseudouridine. These features are known in the art, see, e.g., Zhang et al. Front. Immunol., DOI=10.3389 / fimmu.2019.00594 (2019); Eyler et al. PNAS 116(46): 23068-23071; DOI: 10.1073 / pnas.1821754116 (2019); Nance and Meier, ACS Cent. Sci. 2021, 7, 5, 748-756; doi.org / 10.1021 / acscentsci.1c00197 (2021), and van Hoecke and Roose, J. Translational Med 17:54 (2019); Please see https: / / doi.org / 10.1186 / s12967-019-1804-8.

[0026] The term "isolated" means that the material is removed from its original environment (e.g., the natural environment if it occurs in nature). For example, a naturally occurring nucleic acid or polypeptide present in a living animal is not isolated, but the same nucleic acid or polypeptide is isolated, separated from some or all of the coexisting materials of the natural system. Such a nucleic acid may be part of a vector, and / or such a nucleic acid or polypeptide may be part of a composition (e.g., a cell lysate), and is still isolated in that such a vector or composition is not part of the natural environment of the nucleic acid or polypeptide. "Isolated" can also refer in some embodiments to an antibody, polynucleotide, vector, host cell, or composition that is outside of a subject, e.g., outside of the human body.

[0027] The term "gene" refers to a segment of DNA or RNA involved in producing a polypeptide chain; in certain contexts, it includes the regions preceding and following the coding region (e.g., the 5' untranslated region (UTR) and the 3' UTR), and the intervening sequences (introns) between individual coding segments (exons).

[0028] As used herein, the terms "engineered," "recombinant," or "non-natural" refer to an organism, microorganism, cell, nucleic acid molecule, or vector that contains at least one genetic modification or that has been modified by gene transfer, where such modification or modification is introduced by genetic engineering (i.e., human intervention). Such an organism, microorganism, cell, nucleic acid molecule, or vector can also be described as "modified." Genetic modifications include, for example, modifications that introduce expressible nucleic acid molecules that code for functional RNA, proteins, fusion proteins, or enzymes, or the addition, deletion, substitution, or other functional disruption of other nucleic acid molecules, or the genetic material of a cell. Additional modifications include, for example, non-coding regulatory regions, where the modification alters expression of a polynucleotide, gene, or operon. A polypeptide encoded by an engineered non-natural nucleic acid molecule can be described as an "engineered" polypeptide.

[0029] As used herein, "heterologous" or "non-endogenous" or "exogenous" refers to a gene, protein, compound, nucleic acid molecule, or activity that is not native to a host cell or subject, or any gene, protein, compound, nucleic acid molecule, or activity that is native to a host cell or subject that has been modified. Heterologous, non-endogenous, or exogenous includes genes, proteins, compounds, or nucleic acid molecules that have been mutated or modified such that the structure, activity, or both differ between the native and modified gene, protein, compound, or nucleic acid molecule. In certain embodiments, a heterologous, non-endogenous, or exogenous gene, protein, or nucleic acid molecule (e.g., a receptor, ligand, etc.) is not endogenous to the host cell or subject, but may be a nucleic acid that encodes such a gene. The protein or nucleic acid molecule may have been added to the host cell by conjugation, transformation, transfection, electroporation, etc., and the added nucleic acid molecule may be integrated into the genome of the host cell or may exist as extrachromosomal genetic material (e.g., as a plasmid or other self-replicating vector). The term "homologous" or "homolog" refers to a gene, protein, compound, nucleic acid molecule, or activity found within or derived from a host cell, species, or strain. For example, a heterologous or exogenous polynucleotide or gene encoding a polypeptide may be homologous to a native polynucleotide or gene and encode a homologous polypeptide or activity, but the polynucleotide or polypeptide may have an altered structure, sequence, expression level, or any combination thereof. The non-endogenous polynucleotide or gene, as well as the encoded polypeptide or activity, may be derived from the same species, a different species, or a combination thereof.

[0030] In certain embodiments, a nucleic acid molecule or a portion thereof native to a host cell is considered heterologous to the host cell if it has been modified or mutated, or a nucleic acid molecule native to a host cell can be considered heterologous if it has been modified, modified with heterologous expression control sequences, or modified with endogenous expression control sequences that are not normally associated with the nucleic acid molecule native to the host cell. Furthermore, the term "heterologous" can refer to biological activities that are different, altered, or not endogenous to the host cell. As described herein, multiple heterologous nucleic acid molecules can be introduced into a host cell as separate nucleic acid molecules, as multiple genes that are individually controlled, as polycistronic nucleic acid molecules, as a single nucleic acid molecule encoding an antibody (or other polypeptide), or any combination thereof.

[0031] As used herein, the terms "endogenous" or "native" refer to a polynucleotide, gene, protein, compound, molecule, or activity that is normally present in a host cell or subject.

[0032] As used herein, the term "expression" refers to the process by which a polypeptide is produced based on a coding sequence of a nucleic acid molecule, such as a gene. The process may include transcription, post-transcriptional control, post-transcriptional modification, translation, post-translational control, post-translational modification, or any combination thereof. An expressed nucleic acid molecule is typically operably linked to an expression control sequence (e.g., a promoter).

[0033] The term "operably linked" refers to the association of two or more nucleic acid molecules on a single nucleic acid fragment such that the function of one is affected by the function of the other. For example, a promoter is operably linked to a coding sequence if it can affect the expression of that coding sequence (i.e., the coding sequence is under the transcriptional control of the promoter). "Unlinked" means that the associated genetic elements are not closely related to each other so that the function of one does not affect the other.

[0034] As described herein, two or more heterologous nucleic acid molecules can be introduced into a host (e.g., human) or host cell as separate nucleic acid molecules, as multiple genes that are individually controlled, as polycistronic nucleic acid molecules, as a single nucleic acid molecule encoding a protein (e.g., the heavy chain of an antibody), or any combination thereof. When two or more heterologous nucleic acid molecules are introduced into a host or host cell, it is understood that the two or more heterologous nucleic acid molecules can be introduced separately as a single nucleic acid molecule (e.g., on a single vector). The vector is integrated into the host chromosome at a single site or multiple sites, or any combination thereof. The number of heterologous nucleic acid molecules or protein activities referred to refers to the number of encoding nucleic acid molecules or protein activities, not the number of separate nucleic acid molecules introduced into the host or host cell.

[0035] The term "construct" refers to any polynucleotide that contains a recombinant nucleic acid molecule (or a protein of the present disclosure, if the context clearly indicates). A (polynucleotide) construct may be present in a vector (e.g., bacterial vector, viral vector) or integrated into a genome. A "vector" is a nucleic acid molecule capable of transporting another nucleic acid molecule. A vector may be, for example, a plasmid, a cosmid, a virus, an RNA vector, or a linear or circular DNA or RNA molecule that may include chromosomal, non-chromosomal, semisynthetic or synthetic nucleic acid molecules. Vectors of the present disclosure also include transposon systems (see, for example, Sleeping Beauty, e.g., Geurts et al., Mol. Ther. 8:108, 2003: Mates et al., Nat. Genet. 41:753, 2009). Exemplary vectors are those that can replicate autonomously (episomal vectors), deliver polynucleotides to a cellular genome (e.g., viral vectors), or express the nucleic acid molecule to which they are linked (expression vectors).

[0036] As used herein, "expression vector" or "vector" refers to a DNA construct containing a nucleic acid molecule operably linked to a suitable control sequence capable of effecting expression of the nucleic acid molecule in a suitable host. Such control sequences include a promoter to effect transcription, an optional operator sequence to control such transcription, a sequence encoding a suitable mRNA ribosomal binding site, and a sequence to control the termination of transcription and translation. A vector may be a plasmid, a phage particle, a virus, or simply a potential genome insert. Once transformed into a suitable host, a vector may replicate and function independently of the host genome, or in some cases may integrate into the genome itself or deliver a polynucleotide contained in the vector to the genome without vector sequences. In this specification, "plasmid", "expression plasmid", "virus", and "vector" are often used interchangeably.

[0037] The term "introduced" in the context of inserting a nucleic acid molecule into a cell means "transfection," "transformation," or "transduction," and includes reference to the incorporation of a nucleic acid molecule into a eukaryotic or prokaryotic cell. The nucleic acid molecule may be integrated into the genome of the cell (e.g., chromosome, plasmid, plastid, or mitochondrial DNA), converted into an autonomous replicon, or expressed transiently (e.g., transfected mRNA).

[0038] In certain embodiments, the polynucleotide of the present disclosure may be operably linked to certain elements of a vector. For example, the polynucleotide sequences necessary to affect the expression and processing of the linked coding sequence may be operably linked. Expression control sequences may include appropriate transcription initiation, transcription termination, promoter and enhancer sequences; efficient RNA processing signals such as splicing and polyadenylation signals; sequences that stabilize cytoplasmic mRNA; sequences that increase translation efficiency (i.e., Kozak consensus sequences); sequences that increase protein stability; and sequences that possibly enhance protein secretion. Expression control sequences may be operably linked if they are contiguous with the gene of interest and are expression control sequences that act in trans or at a distance to control the gene of interest.

[0039] In certain embodiments, the vector comprises a plasmid vector or a viral vector (e.g., a lentiviral vector or a gamma-retroviral vector). Viral vectors include negative-stranded RNA viruses such as retroviruses, adenoviruses, parvoviruses (e.g., adeno-associated viruses), coronaviruses, orthomyxoviruses (e.g., influenza viruses), rhabdoviruses (e.g., rabies virus and vesicular stomatitis virus), paramyxoviruses (e.g., measles and Sendai), positive-stranded RNA viruses such as picornaviruses and alphaviruses, and double-stranded DNA viruses such as adenoviruses, herpesviruses (e.g., herpes simplex virus types 1 and 2, Epstein-Barr virus, cytomegalovirus), and poxviruses (e.g., vaccinia, fowlpox, and canarypox). Other viruses include, for example, Norwalk virus, togaviruses, flaviviruses, reoviruses, papovaviruses, hepadnaviruses, and hepatitis viruses. Examples of retroviruses include avian leukosis sarcoma, mammalian types C, B and D viruses, the HTLV-BLV complex, lentiviruses and spumaviruses (Coffin, JM, Retroviridae: The viruses and their replication, In Fundamental Virology, Third Edition, BN Fields et al., Eds., Lippincott-Raven Publishers, Philadelphia, 1996).

[0040] A "retrovirus" is a virus with an RNA genome that is reverse transcribed into DNA using reverse transcriptase, and the reverse transcribed DNA is then integrated into the host cell genome. A "gammaretrovirus" refers to a genus of the Retroviridae family. Examples of gammaretroviruses include mouse stem cell virus, murine leukemia virus, feline leukemia virus, feline sarcoma virus, and avian reticuloendotheliosis virus.

[0041] "Lentiviral vector" includes HIV-based lentiviral vectors for gene delivery, which can be integrating or non-integrating, have a relatively large packaging capacity, and can transduce a variety of different cell types. Lentiviral vectors are usually generated after transient transfection of three (packaging, envelope, transfer) or more plasmids into producer cells. Like HIV, lentiviral vectors enter target cells through the interaction of viral surface glycoproteins with cell surface receptors. Once inside, viral RNA undergoes reverse transcription mediated by the viral reverse transcriptase complex. The product of reverse transcription is double-stranded linear viral DNA, which is the substrate for viral integration into the DNA of infected cells.

[0042] In certain embodiments, the viral vector can be a gamma retrovirus, such as Moloney Murine Leukemia Virus (MLV) derived vector. In other embodiments, the viral vector can be a more complex retrovirus derived vector, such as a lentivirus derived vector. HIV-1 derived vectors belong to this category. Other examples include lentivirus vectors derived from HIV-2, FIV, equine infectious anemia virus, SIV, and Maedi-Visna virus (ovine lentivirus). Methods of using retroviral and lentiviral viral vectors and packaging cells to transduce mammalian host cells with viral particles containing transgenes are known in the art and have been previously described, for example, in US Patent 8,119,772; Walchli et al., PLoS One 6:327930, 2011; Zhao et al., J. Immunol. 174:4415, 2005; Engels et al., Hum. Gene Ther. 14:1155, 2003; Frecha et al., Mol. Ther. 18:1748, 2010; and Verhoeyen et al., Methods Mol. Biol. 506:97, 2009. Retroviral and lentiviral vector constructs and expression systems are also commercially available. Other viral vectors can also be used for polynucleotide delivery, including, for example, DNA viral vectors, including adenovirus-based vectors and adeno-associated virus (AAV)-based vectors; amplicon vectors, vectors derived from herpes simplex virus (HSV), including replication-deficient HSV and attenuated HSV (Krisky et al., Gene Ther. 5:1517, 1998).

[0043] Other vectors that can be used with the compositions and methods of the present disclosure include vectors derived from baculovirus and alphavirus (Jolly, D J. 1999. Emerging Viral Vectors. pp 209-40 in Friedmann T. ed. The Development of Human Gene Therapy. New York: Cold Spring Harbor Lab), or plasmid vectors (such as sleeping beauty or other transposon vectors).

[0044] When the viral vector genome contains multiple polynucleotides that are expressed as separate transcripts in a host cell, the viral vector may also contain additional sequences between the two (or more) transcripts that allow for bicistronic or multicistronic expression. Examples of such sequences used in viral vectors include an internal ribosome entry site (IRES), a furin cleavage site, a viral 2A peptide, or any combination thereof.

[0045] Plasmid vectors, including plasmid vectors encoding DNA-based antibodies for direct administration to a subject, are further described herein.

[0046] As used herein, the term "host" refers to a cell or microorganism that is targeted for genetic modification with a heterologous nucleic acid molecule to produce a polypeptide of interest (e.g., an antibody of the present disclosure). "Host" can also refer to a subject to which an antibody-encoding nucleic acid is administered and / or a subject that has a paramyxovirus, e.g., respiratory syncytial virus, metapneumovirus, or pneumovirus of mice.

[0047] A host cell may include any individual cell or cell culture that can receive a vector, incorporate a nucleic acid, or express a protein. The term also encompasses the progeny of the host cell, whether genetically or phenotypically the same or different. Suitable host cells depend on the vector and may include mammalian cells, animal cells, human cells, monkey cells, insect cells, yeast cells, and bacterial cells. These cells may be induced to incorporate vectors or other materials by the use of viral vectors, transformation by calcium phosphate precipitation, DEAE-dextran, electroporation, microinjection, or other methods. See, e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual 2d ed. (Cold Spring Harbor Laboratory, 1989).

[0048] As used herein, "antigen" or "Ag" refers to an immunogenic molecule that elicits an immune response. This immune response may involve antibody production, activation of specific immunocompetent cells, activation of complement, antibody-dependent cellular cytotoxicity, or any combination thereof. An antigen (immunogenic molecule) may be, for example, a peptide, glycopeptide, polypeptide, glycopolypeptide, polynucleotide, polysaccharide, lipid, etc. It is readily apparent that an antigen may be synthetic, recombinantly produced, or derived from a biological sample. Exemplary biological samples that may contain one or more antigens include tissue samples, fecal samples, cells, bodily fluids, or combinations thereof. An antigen is produced by a cell that has been modified or genetically engineered to express the antigen. An antigen may also be present in a paramyxovirus (e.g., an F protein or a portion thereof), for example, in a virion, or expressed or presented on the surface of a cell infected with a paramyxovirus.

[0049] The term "epitope" or "antigenic epitope" includes any molecule, structure, amino acid sequence, or protein determinant that is recognized and specifically bound by a cognate binding molecule, such as an immunoglobulin, or other binding molecule, domain, or protein. Epitopic determinants generally contain chemically active surface groupings of molecules such as amino acids or sugar side chains and can have specific three-dimensional structural characteristics, as well as specific charge characteristics. When the antigen is or comprises a peptide or protein, the epitope can be composed of contiguous amino acids (e.g., a linear epitope), or can be composed of amino acids from different parts or regions of the protein that are brought into close proximity by protein folding (e.g., a discontinuous or conformational epitope), or noncontiguous amino acids that are brought into close proximity regardless of protein folding.

[0050] antibody In some embodiments, the disclosure provides an isolated antibody comprising: (i) two heavy chain polypeptides each having (i.e., comprising or consisting of) the amino acid sequence of SEQ ID NO:1; and (ii) two light chain polypeptides each having (i.e., comprising or consisting of) the amino acid sequence of SEQ ID NO:4.

[0051] In other embodiments, the disclosure provides an isolated antibody comprising: (i) two heavy chain polypeptides, each having (i.e., comprising or consisting of) the amino acid sequence of SEQ ID NO:2; and (ii) two light chain polypeptides, each having (i.e., comprising or consisting of) the amino acid sequence of SEQ ID NO:4.

[0052] In yet other embodiments, the disclosure provides an isolated antibody comprising: (i) two heavy chain polypeptides, each having (i.e., comprising or consisting of) the amino acid sequence of SEQ ID NO:3; and (ii) two light chain polypeptides, each having (i.e., comprising or consisting of) the amino acid sequence of SEQ ID NO:4.

[0053] In some embodiments, the disclosure provides an isolated antibody comprising: (i) two heavy chain polypeptides, each comprising the amino acid sequence of SEQ ID NO:6; and (ii) two light chain polypeptides, each comprising the amino acid sequence of SEQ ID NO:4.

[0054] In some embodiments, the disclosure provides an isolated antibody comprising: (i) two heavy chain polypeptides, each comprising the amino acid sequence of SEQ ID NO:7; and (ii) two light chain polypeptides, each comprising the amino acid sequence of SEQ ID NO:4.

[0055] In some embodiments, the disclosure provides an isolated antibody comprising: (i) two heavy chain polypeptides, each comprising the amino acid sequence of SEQ ID NO:8; and (ii) two light chain polypeptides, each comprising the amino acid sequence of SEQ ID NO:4.

[0056] The two heavy chain polypeptides of the antibody of the present disclosure combine to form a homodimer, one of the two heavy chain polypeptides further associated with one of the two light chain polypeptides, and the other of the two heavy chain polypeptides further associated with the other of the two light chain polypeptides.

[0057] The antibodies can bind to and neutralize infection by a paramyxovirus, which can be, for example, a respiratory syncytial virus, a metapneumovirus, or a pneumonia virus of mice.

[0058] As used herein, a "neutralizing antibody" is an antibody that can neutralize, i.e., prevent, inhibit, reduce, hinder, or interfere with the ability of a pathogen to initiate and / or perpetuate infection in a host. The terms "neutralizing antibody" and "neutralizing antibody" or "neutralizing antibody" are used interchangeably herein. In any of the embodiments disclosed herein, the antibody can prevent and / or neutralize paramyxovirus infection in an in vitro infection model and / or an in vivo animal infection model and / or in humans.

[0059] Terms understood by those skilled in the art of antibody technology are given their respective art-acquired meanings unless expressly defined otherwise herein.For example, the term "antibody" refers to an intact antibody comprising at least two heavy (H) chains (HC; also referred to as heavy chain polypeptides) and two light (L) chains (LC; also referred to as light chain polypeptides) connected by disulfide bonds (i.e., LC-HC-HC-LC as a bivalent tetrameric molecule).This term encompasses intact or full-length antibodies, including antibodies of any class or subclass, including IgG and its subclasses (IgG1, IgG2, IgG3, IgG4), IgM, IgE, IgA, and IgD.

[0060] The terms "VL" or "VL" and "VH" or "VH" refer to the variable binding regions from antibody light chains and antibody heavy chains, respectively. The variable binding regions contain distinct, well-defined subregions known as "complementarity determining regions" (CDRs) and "framework regions" (FRs). The terms "complementarity determining regions" and "CDRs" are synonymous with "hypervariable regions" or "HVRs" and generally refer to amino acid sequences within antibody variable regions that together confer antigen specificity and / or binding affinity to an antibody, where consecutive CDRs (i.e., CDR1 and CDR2, CDR2 and CDR3) are separated from each other in the primary structure by framework regions. There are three CDRs in each variable region (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, LCDR3, also called CDRH and CDRL, respectively). In certain embodiments, antibody VH comprises the following four FRs and three CDRs: FR1-HCDR1-FR2-HCDR2-FR3-HCDR3-FR4; antibody VL comprises the following four FRs and three CDRs: FR1-LCDR1-FR2-LCDR2-FR3-LCDR3-FR4. Generally, VH and VL together form an antigen binding site via their respective CDRs. However, in some cases, VH alone, VL alone, or 1, 2, 3, 4, or 5 CDRs are involved in antigen binding.

[0061] Numbering of the CDR and framework regions may be based on, for example, the Kabat, Chothia, EU, IMGT, and AHo numbering schemes (see, e.g., Kabat et al., "Sequences of Proteins of Immunological Interest, US Dept. Health and Human Services, Public Health Service National Institutes of Health, 1991, 5 th ed.; Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987); Lefranc et al., Dev. Comp. Immunol. 27:55, 2003; Honegger and Pluckthun, J. Mol. Bio. 309:657-670 (2001); and the antibody numbering method developed by the Chemical Computing Group (CCG) using, for example, the Molecular Operating Environment (MOE) software (www.chemcomp.com). Equivalent residue positions can be annotated and different molecules can be compared using the Antigen Receptor Numbering and Receptor Classification (ANARCI) software tool (2016, Bioinformatics 15:298-300). Thus, the identification of the CDRs of the exemplary variable domain (VH or VL) sequences provided herein according to one numbering scheme does not exclude antibodies that contain the CDRs of the same variable domain determined using a different numbering scheme.

[0062] The term "CL" refers to the "immunoglobulin light chain constant region" or "light chain constant region", i.e., the constant region from an antibody light chain. The term "CH" refers to the "immunoglobulin heavy chain constant region" or "heavy chain constant region", which can be further divided into CH1, CH2, and CH3 (IgA, IgD, IgG), or CH1, CH2, CH3, and CH4 domains (IgE, IgM), depending on the antibody isotype. The Fc region of an antibody heavy chain is further described herein. In any of the embodiments of the present disclosure, the antibody of the present disclosure comprises a CL, a CH1, a CH2, and a CH3.

[0063] "Fab" (Fragment Antigen Binding) is the portion of an antibody that binds to an antigen and comprises the variable region and CH1 of a heavy chain linked to a light chain via one or more interchain disulfide bonds. Each Fab fragment is monovalent with respect to antigen binding, i.e., it has a single antigen-binding site. Pepsin treatment of an antibody yields a single large F(ab')2 fragment, roughly corresponding to two disulfide-linked Fab fragments, that has bivalent antigen-binding activity and is still capable of cross-linking antigen. Both Fab and F(ab')2 are examples of "antigen-binding fragments". Fab' fragments differ from Fab fragments in that they have additional residues at the carboxy terminus of the CH1 domain, including one or more cysteines from the antibody hinge region. Fab'-SH is the designation herein for Fab' in which the cysteine ​​residues of the constant domains bear a free thiol group. F(ab')2 antibody fragments were originally produced as pairs of Fab' fragments with hinge cysteines between them. Other chemical bonds of antibody fragments are also known.

[0064] "Fv" is a small antibody fragment that contains a complete antigen recognition and binding site. This fragment usually consists of a dimer of one heavy-chain variable domain and one light-chain variable domain in tight non-covalent association. However, even a single variable domain (or half of an Fv containing only three CDRs specific for an antigen) has the ability to recognize and bind antigen, although sometimes with a lower affinity than the entire binding site.

[0065] The presently disclosed antibodies include an Fc polypeptide. An "Fc" dimer comprises the carboxy-terminal portions of both antibody H chains (i.e., the CH2 and CH3 domains of IgG) linked by disulfides. An antibody "effector function" refers to the biological activities attributable to the Fc region of an antibody (either a native sequence Fc region or an amino acid sequence variant Fc region) and varies with the antibody isotype. Examples of antibody effector functions include: C1q binding and complement-dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; downregulation of cell surface receptors (e.g., B cell receptor); and activation of B cells.

[0066] In some embodiments, the antibody comprises an Fc polypeptide comprising mutations selected from the following (EU numbering referring to human IgG1): G236A; S239D; A330L; and I332E; or a combination comprising any two or more of the same; for example, S239D / I332E; S239D / A330L / I332E; G236A / S239D / I332E; G236A / A330L / I332E (also referred to herein as "GAALIE"); or G236A / S239D / A330L / I332E. In some embodiments, the Fc polypeptide or fragment thereof does not comprise S239D. In some embodiments, the Fc polypeptide or fragment thereof comprises an S at position 239. GAALIE improves binding affinity to FcRγIIa and FcγRIIIa and reduces binding affinity to FcγRIIb.

[0067] In certain embodiments, the Fc polypeptide comprises one or more amino acid modifications that improve binding affinity to FcRn (e.g., enhance binding to FcRn) (e.g., at a pH of about 6.0), thereby extending in vivo half-life of a molecule comprising the Fc polypeptide (e.g., compared to a reference Fc polypeptide or antibody that is otherwise identical but does not comprise the modification). In certain embodiments, the Fc polypeptide comprises or is derived from an IgG (e.g., IgG1) Fc and half-life extending mutations, the half-life extending mutations comprising any one or more of the following: N434S; N434H; N434A; N434S; M252Y; S254T; T256E; T250Q; P257IQ311I; D376V; T307A; E380A (EU numbering referring to human IgG1). In certain embodiments, the half-life extending mutations include M428L / N434S (also referred to herein as "MLNS" or "LS"). In certain embodiments, the half-life extending mutations include M252Y / S254T / T256E. In certain embodiments, the half-life extending mutations include T250Q / M428L. In certain embodiments, the half-life extending mutations include P257I / Q311I. In certain embodiments, the half-life extending mutations include P257I / N434H. In certain embodiments, the half-life extending mutations include D376V / N434H. In certain embodiments, the half-life extending mutations include T307A / E380A / N434A.

[0068] In some embodiments, the antibody comprises an Fc polypeptide comprising substitution mutations M428L / N434S. In some embodiments, the antibody comprises an Fc polypeptide comprising substitution mutations G236A / A330L / I332E. In certain embodiments, the antibody comprises an Fc polypeptide comprising G236A, A330L, and I332E mutations (GAALIE) and no S239D mutation (e.g., comprising a natural S at position 239) (e.g., IgG1). In certain embodiments, the antibody comprises an Fc polypeptide comprising substitution mutations: M428L / N434S and G236A / A330L / I332E, optionally no S239D. In certain embodiments, the antibody comprises an Fc polypeptide comprising substitution mutations: M428L / N434S and G236A / S239D / A330L / I332E (EU numbering referring to human IgG1).

[0069] For example, it is understood that production in a mammalian cell line may result in the removal of one or more C-terminal lysines of an antibody heavy chain polypeptide (see, e.g., Liuetal.mAbs6(5):1145-1154(2014)). Thus, an antibody of the present disclosure can include a heavy chain polypeptide in which a C-terminal lysine residue is present or absent. In other words, embodiments in which the C-terminal residue of a heavy chain polypeptide is not a lysine and embodiments in which lysine is the C-terminal residue are encompassed. In certain embodiments, a composition comprises a plurality of antibodies of the present disclosure, wherein one or more antibodies do not include a lysine residue at the C-terminus of the heavy chain polypeptide, and wherein one or more antibodies include a lysine residue at the C-terminus of the heavy chain polypeptide.

[0070] Thus, in some embodiments, the disclosure provides an isolated antibody comprising: (i) two heavy chain polypeptides each comprising the amino acid sequence of SEQ ID NO:6, SEQ ID NO:7, or SEQ ID NO:8; and (ii) two light chain polypeptides each comprising the amino acid sequence of SEQ ID NO:4.

[0071] In certain embodiments, the antibody comprises a glycosylation-altering mutation, the glycosylation-altering mutation comprises N297A, N297Q, or N297G, and / or the antibody is partially or completely aglycosylated, and / or partially or completely afucosylated. Host cell lines and methods for making partially or completely aglycosylated or partially or completely afucosylated antibodies are known (see, e.g., PCT Publication No. WO 2016 / 181357; Suzuki et al. Clin. Cancer Res. 13(6):1875-82 (2007); Huang et al. MAbs 6:1-12 (2018)). Fucosylation of an antibody can be achieved by expressing the antibody in a host cell that has been genetically engineered to lack the ability to fucosylate the antibody (or have an inhibited or impaired ability) by introducing amino acid mutations to introduce or destroy fucosylation sites, by expressing the antibody under conditions in which the ability of the host cell to fucosylate the antibody is impaired (e.g., in the presence of 2-fluoro-L-fucose (2FF)), etc.

[0072] In certain embodiments, the antibody is: afucosylated; produced in a host cell that is unable to fucosylate the antibody or has an inhibited ability to fucosylate the antibody; produced under conditions that inhibit fucosylation by the host cell; or any combination thereof.

[0073] In certain embodiments, the antibody can induce continued protection in vivo in a subject even after the subject no longer finds detectable levels of the antibody (i.e., when the antibody is cleared from the subject after administration). Such protection is referred to herein as a vaccine effect. Without being bound by theory, it is believed that dendritic cells internalize the antibody-antigen complex and then induce or contribute to an endogenous immune response to the antigen. In certain embodiments, the antibody can activate dendritic cells that can induce T cell immunity to the antigen, for example, and includes one or more modifications, such as mutations in Fc, including G236A, A330L, and I332E.

[0074] In any of the embodiments disclosed herein, the antibody can be monoclonal. As used herein, the term "monoclonal antibody" (mAb) refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies that make up the population are identical except for naturally occurring mutations that may occasionally be present in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody preparations that contain different antibodies against different epitopes, each monoclonal antibody is directed against a single epitope of the antigen. In addition to their specificity, monoclonal antibodies have the advantage that they can be synthesized uncontaminated by other antibodies. The term "monoclonal" should not be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies useful in the present invention may be prepared by the hybridoma method first described by Kohler et al., Nature 256:495 (1975), or may be made using recombinant DNA methods in bacterial, eukaryotic, or plant cells (see, e.g., U.S. Pat. No. 4,816,567). Monoclonal antibodies may also be isolated from phage antibody libraries using the techniques described in, for example, Clackson et al., Nature, 352:624-628 (1991) and Marks et al., J. Mol. Biol., 222:581-597 (1991). Monoclonal antibodies may also be obtained using the methods disclosed in PCT Publication No. WO2004 / 076677A2.

[0075] A "human antibody" is an antibody that contains only sequences present in antibodies produced by humans. However, as used herein, a human antibody may contain residues or modifications that are not found in naturally occurring human antibodies (e.g., antibodies isolated from humans), including the modifications and mutant sequences described herein. These are typically created to further improve or enhance the performance of the antibody. In some cases, human antibodies are produced by transgenic animals. See, for example, U.S. Patent Nos. 5,770,429; 6,596,541 and 7,049,426.

[0076] Polynucleotides, Vectors, and Host Cells In another aspect, the disclosure provides an isolated polynucleotide encoding any of the antibodies or portions thereof (e.g., CDRs, VH, VL, heavy chains, or light chains) of the disclosure. The disclosed polynucleotides include those that encode, for example, an antibody heavy chain, an antibody light chain, an antibody heavy and light chains, or two antibody heavy chains and two antibody light chains.

[0077] In certain embodiments, the polynucleotide is codon-optimized for expression in a host cell. From a known coding sequence, codon optimization can be performed using known techniques and tools, such as the GenScript® OptimiumGene® tool or GeneArt® (ThermoFisher) Gene Synthesis; see also, for example, Scholten et al., Clin. Immunol. 119:135, 2006). Codon-optimized sequences include partially codon-optimized sequences (i.e., one or more codons are optimized for expression in a host cell) and fully codon-optimized sequences.

[0078] It is also understood that polynucleotides encoding an antibody of the present disclosure may have different nucleotide sequences, for example, due to the degeneracy of the genetic code, splicing, etc., while still encoding the same antibody.

[0079] In certain embodiments, it is understood that a polynucleotide encoding an antibody or portion thereof is included within a polynucleotide that includes other sequences and / or features for expression of the antibody or portion thereof, e.g., in a host cell. Exemplary features include promoter sequences, polyadenylation sequences, sequences encoding signal peptides (e.g., located at the N-terminus of the expressed antibody heavy or light chain), and the like.

[0080] In any of the embodiments disclosed herein, the polynucleotide can comprise deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). In some embodiments, the RNA comprises messenger RNA (mRNA).

[0081] In some embodiments, the polynucleotide comprises a modified nucleoside, a cap-1 structure, a cap-2 structure, or any combination thereof. In certain embodiments, the polynucleotide comprises pseudouridine, N6-methyladenosine, 5-methylcytidine, 2-thiouridine, or any combination thereof. In some embodiments, the pseudouridine comprises N1-methylpseudouridine.

[0082] Also provided is a vector that comprises or contains the polynucleotide disclosed herein. The vector can comprise any one or more of the vectors disclosed herein. In certain embodiments, a vector is provided that comprises a DNA plasmid construct that encodes an antibody or a portion thereof (e.g., so-called "DMAb"; see, for example, Muthumani et al., J Infect Dis. 214(3):369-378 (2016); Muthumani et al., Hum Vaccin Immunother 9:2253-2262 (2013); Flingai et al., Sci Rep. 5:12616 (2015); and Elliott et al., NPJ Vaccines 18 (2017) that are incorporated herein by reference for the DNA construct that encodes the antibody and related methods of use, including administration). In certain embodiments, the DNA plasmid construct comprises a single open reading frame encoding the heavy and light chains (or VH and VL) of the antibody, where the heavy chain encoding sequence and the light chain encoding sequence are optionally separated by a polynucleotide encoding a protease cleavage site and / or a polynucleotide encoding a self-cleaving peptide. In some embodiments, the replacement components of the antibody are encoded by polynucleotides contained in a single plasmid. In other embodiments, the replacement components of the antibody are encoded by polynucleotides contained in two or more plasmids (e.g., a first plasmid contains a polynucleotide encoding a heavy chain, VH, or VH+CH, and a second plasmid contains a polynucleotide encoding a cognate light chain, VL, or VL+CL). An exemplary expression vector is pVax1 available from Invitrogen®. The DNA plasmids of the present disclosure can be delivered to a subject, for example, by electroporation (e.g., intramuscular electroporation) or with an appropriate formulation (e.g., hyaluronidase). In some embodiments, the vectors of the present disclosure contain a nucleotide sequence encoding a signal peptide.The signal peptide may or may not be present on the mature antibody (e.g., it may be enzymatically cleaved). In some embodiments, the vectors of the disclosure include a polyadenylation signal sequence.

[0083] In some embodiments, a vector of the present disclosure comprises a CMV promoter.

[0084] In some embodiments, methods are provided that include administering to a subject a first polynucleotide (e.g., mRNA) encoding an antibody heavy chain, VH, or Fd (VH+CH1), and administering to a subject a second polynucleotide (e.g., mRNA) encoding a cognate antibody light chain, VL, or VL+CL.

[0085] In some embodiments, polynucleotides (e.g., mRNA) encoding heavy and light chains of the antibody are provided. In some embodiments, polynucleotides (e.g., mRNA) encoding two heavy and two light chains of the antibody are provided. See, for example, Li, JQ., Zhang, ZR., Zhang, HQ. et al. Intranasal delivery of replicating mRNA encoding neutralizing antibody against SARS-CoV-2 infection in mice. Sig Transduct Target Ther 6, 369 (2021). https: / / doi.org / 10.1038 / s41392-021-00783-1, whose antibody-encoding mRNA constructs, vectors, and related techniques are incorporated herein by reference. In some embodiments, the polynucleotide is delivered to the subject via an alphavirus replicon particle (VRP) delivery system. In some embodiments, the replicon comprises a modified VEEV replicon comprising two subgenomic promoters. In some embodiments, the polynucleotide or replicon is capable of co-translating the heavy (or VH, or VH+1) and light (or VL, or VL+CL) chains of an antibody. In some embodiments, methods are provided that include delivering such a polynucleotide or replicon to a subject.

[0086] In a further aspect, the present disclosure also provides a host cell that expresses an antibody according to the present disclosure; or that comprises or contains a vector or polynucleotide according to the present disclosure.

[0087] Examples of such cells include, but are not limited to, eukaryotic cells, such as yeast cells, animal cells, insect cells, plant cells; and prokaryotic cells, such as E. coli. In some embodiments, the cell is a mammalian cell. In certain such embodiments, the cell is a CHO cell (e.g., a DHFR-CHO cell (e.g., Urlaub et al., PNAS 77:4216 (1980)), human embryonic kidney cells (e.g., HEK293T cells), PER.C6 cells, Y0 cells, Sp2 / 0 cells, and other mammalian cell lines. NS0 cells, human liver cells, e.g., HepaRG cells, myeloma cells, or hybridoma cells. Other examples of mammalian host cell lines include mouse Sertoli cells (e.g., TM4 cells), SV40-transformed monkey kidney CV1 line (COS-7); baby hamster kidney cells (BHK); African green monkey kidney cells (VERO-76); monkey kidney cells (CV1); human cervical carcinoma cells (HELA); human lung cells (W138); human liver cells (HepG2); canine kidney cells (MDCK; buffalo rat liver cells (BRL3A); mouse mammary tumor (MMT060562); TRI cells; MRC5 cells; and FS4 cells. Suitable mammalian host cell lines for antibody production are described, for example, in Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003).

[0088] In certain embodiments, the host cell is a prokaryotic cell, such as E. coli. Expression of peptides in prokaryotic cells, such as E. coli, is well established (see, e.g., Pluckthun, A. Bio / Technology 9:545-551 (1991). For example, antibodies may be produced in bacteria, particularly in the case of glycosylation. For expression of antibody polypeptides in bacteria, see, e.g., U.S. Patent Nos. 5,648,237, 5,789,199, and 5,840,523.

[0089] In certain embodiments, cells may be transfected with the vectors of the invention using an expression vector. The term "transfection" refers to the introduction of a nucleic acid molecule, such as a DNA or RNA (e.g., mRNA) molecule, into a cell, such as a eukaryotic cell. In the present context, the term "transfection" encompasses any method known to those skilled in the art for introducing a nucleic acid molecule into a cell, such as a eukaryotic cell, including a mammalian cell. Such methods include, for example, electroporation, lipofection, such as cationic lipids and / or liposomes, calcium phosphate precipitation, nanoparticle-based transfection, virus-based transfection, or cationic polymers, such as DEAE-dextran or polyethyleneimine-based transfection, and the like. In certain embodiments, the introduction is non-viral.

[0090] Furthermore, the host cell of the present disclosure may be stably or transiently transfected with a vector, e.g., a vector according to the present disclosure. A method for expressing an antibody according to the present disclosure. In such an embodiment, the cell may be stably transfected with a vector described herein. Alternatively, the cell may be transiently transfected with a vector according to the present disclosure that encodes an antibody disclosed herein. In any of the embodiments disclosed herein, the polynucleotide may be heterologous to the host cell.

[0091] Thus, the present disclosure also provides recombinant host cells that heterologously express the antibodies of the present disclosure. For example, the cells may be of a species different from the species from which the antibody was derived, fully or partially (e.g., CHO cells expressing a human antibody or an engineered human antibody). In some embodiments, the cell type of the host cell does not naturally express the antibody. Furthermore, the host cell may impart post-translational modifications (PTMs; e.g., glycosylation or fucosylation) to the antibody that are not present in the native state of the antibody (or the native state of the parent antibody from which the antibody was engineered or derived). Such PTMs may cause functional differences (such as reduced immunogenicity). Thus, the antibodies of the present disclosure produced by the host cells disclosed herein may contain one or more post-translational modifications that differ from the antibody in its native state (or the parent antibody) (e.g., a human antibody produced by a CHO cell may contain more post-translational modifications that differ from the antibody when isolated from a human and / or when produced by a natural human B cell or plasma cell).

[0092] Insect cells useful for expressing the binding proteins of the present disclosure are known in the art and include, for example, Spodoptera frugipera Sf9 cells, Trichoplusia ni BTI-TN5B1-4 cells, and Spodoptera frugipera SfSWT01 "Mimic" cells. See, for example, Palmberger et al., J. Biotechnol. 153(3-4):160-166 (2011). Many baculovirus strains have been identified that may be used in combination with insect cells, particularly for transfection of Spodoptera fragiperda cells.

[0093] Eukaryotic microbes, such as filamentous fungi or yeast, are also suitable hosts for cloning or expressing protein-encoding vectors, and include fungal and yeast strains with "humanized" glycosylation pathways to produce antibodies with partially or fully human glycosylation patterns. See Gerngross, Nat. Biotech. 22:1409-1414 (2004); Li et al., Nat. Biotech. 24:210-215 (2006).

[0094] Plant cells can also be used as hosts to express the antibodies of the present disclosure. For example, the PLANTIBODIES® technology (described, for example, in U.S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429) uses transgenic plants to produce antibodies.

[0095] In certain embodiments, the host cell comprises a mammalian cell, hi certain embodiments, the host cell is a CHO cell, a HEK293 cell, a PER.C6 cell, a Y0 cell, a Sp2 / 0 cell, a NS0 cell, a human hepatocyte, a myeloma cell, or a hybridoma cell.

[0096] In a related embodiment, the disclosure provides a method for producing an antibody, the method comprising culturing a host cell of the disclosure under conditions and for a time sufficient to produce the antibody. A method useful for isolating and purifying a recombinantly produced antibody may, by way of example, comprise obtaining a supernatant from a suitable host cell / vector system that secretes the recombinant antibody into the medium, and then concentrating the medium using a commercially available filter. After concentration, the concentrate may be applied to a single suitable purification matrix, or to a series of suitable matrices, such as affinity matrices or ion exchange resins. One or more reverse-phase HPLC steps may be used to further purify the recombinant polypeptide. These purification methods may be used when isolating immunogens from their natural environment. One or more large-scale production methods of isolated / recombinant antibodies described herein include batch cell cultures that are monitored and controlled to maintain suitable culture conditions. Purification of soluble antibodies may be performed according to methods described herein and known in the art and compatible with the laws and guidelines of domestic and foreign regulatory authorities.

[0097] composition Also provided herein are compositions comprising any one or more of the antibodies, polynucleotides, vectors, or host cells of the present disclosure, alone or in any combination, and which may further comprise a pharma- ceutically acceptable carrier, excipient, or diluent, which are described in more detail herein.

[0098] In certain embodiments, a composition comprises a plurality of antibodies of the disclosure, wherein one or more of the antibodies of the plurality does not comprise a lysine residue at the C-terminus of a heavy chain polypeptide, and one or more of the antibodies of the plurality comprises a lysine residue at the C-terminus of a heavy chain polypeptide.

[0099] In certain embodiments, the composition comprises a first vector comprising a first plasmid, and a second vector comprising a second plasmid, where the first plasmid comprises a polynucleotide encoding the heavy chain, VH, or VH+CH, of the antibody, and the second plasmid comprises a polynucleotide encoding the cognate light chain, VL, or VL+CL, of the antibody. In certain embodiments, the composition comprises a polynucleotide (e.g., mRNA) bound to a suitable delivery vehicle or carrier. Exemplary vehicles or carriers for administration to a human subject include lipid or lipid-derived delivery vehicles, such as liposomes, solid lipid nanoparticles, oil suspensions, submicron lipid emulsions, lipid microbubbles, reverse lipid micelles, cochlear liposomes, lipid microtubules, lipid microcylinders, lipid nanoparticles (LNPs), or nanoscale platforms (see, e.g., Li et al. Wilery Interdiscip Rev. Nanomed Nanobiotechnol. 11(2):e1530 (2019)). Principles, reagents, and techniques for designing suitable mRNAs, formulating mRNA-LNPs, and delivering them have been described, for example, in Pardi et al. (J Control Release 217345-351 (2015)); Thess et al. (Mol Ther 23: 1456-1464 (2015)); Thran et al. (EMBO Mol Med 9(10):1434-1448 (2017); Kose et al. (Sci. Immunol. 4 eaaw6647 (2019); and Sabnis et al. (Mol. Ther. 26:1509-1519 (2019)). (2018)) and the techniques include capping, codon optimization, nucleoside modification, purification of the mRNA, incorporation of the mRNA into stable lipid nanoparticles (e.g., ionized cationic lipid / phosphatidylcholine / cholesterol / PEG lipid, ionized lipid; distearoyl PC:cholesterol:polyethylene glycol lipid), and subcutaneous, intramuscular, intradermal, intravenous, intraperitoneal, and intratracheal administration thereof, which are incorporated herein by reference.

[0100] Methods and Uses Also provided herein are methods of using the antibodies, nucleic acids, vectors, cells, or compositions of the disclosure in the diagnosis of paramyxoviruses (e.g., in a human subject, or in a sample obtained from a human subject).

[0101] Diagnostic methods (e.g., in vitro, ex vivo) may include contacting an antibody with a sample. Such a sample may be isolated from a subject, for example an isolated tissue sample taken from the nasal passages, paranasal sinuses, salivary glands, lungs, liver, pancreas, kidneys, ears, eyes, placenta, gastrointestinal tract, heart, ovaries, pituitary gland, adrenal gland, thyroid gland, brain, skin, blood. Diagnostic methods may also include detection of antigen / antibody complexes, particularly after contacting the antibody with the sample. Such detection steps can be performed on the bench, i.e. without contacting the human or animal body. Examples of detection methods are well known to those skilled in the art and include, for example, ELISA (enzyme-linked immunosorbent assay), including direct, indirect, and sandwich ELISA.

[0102] Also provided are methods of treating a subject using the disclosed antibodies or compositions comprising same, or the disclosed nucleic acids, vectors, or host cells, or compositions comprising same, where the subject is believed to be infected or at risk of infection with a paramyxovirus, such as, for example, respiratory syncytial virus, metapneumovirus, or mouse pneumonia virus. "Treating", "treatment", or "ameliorating" refers to the medical management of a disease, disorder, or condition in a subject (e.g., a human or a non-human mammal, such as a primate, horse, cat, dog, goat, mouse, or rat). In general, an appropriate dose or treatment regimen comprising the disclosed antibodies, nucleic acids, vectors, host cells, or compositions is administered in an amount sufficient to elicit a therapeutic or prophylactic benefit. Therapeutic or prophylactic / preventative benefits include improved clinical outcomes; reduction or alleviation of symptoms associated with the disease; reduced occurrence of symptoms; improved quality of life; longer disease-free status; reduction in the extent of the disease, stabilization of the disease state; delay or prevention of disease progression; remission; survival; extended survival; or any combination thereof. In certain embodiments, the therapeutic or prophylactic / preventive benefit includes a reduction or prevention of hospitalization for treatment of a paramyxovirus infection (i.e., in a statistically significant manner). In certain embodiments, the therapeutic or prophylactic / preventive benefit includes a reduction in the length of hospitalization for treatment of a paramyxovirus infection (i.e., in a statistically significant manner). In certain embodiments, the therapeutic or prophylactic / preventive benefit includes a reduction or elimination of the need for respiratory interventions, such as intubation and / or use of a ventilator device. In certain embodiments, the therapeutic or prophylactic / preventive benefit includes a reversal of late-stage disease symptoms and / or a reduction in mortality.

[0103] A "therapeutically effective amount" or "effective amount" of an antibody, polynucleotide, vector, host cell, or composition of the present disclosure refers to an amount of a composition or molecule sufficient to produce a therapeutic effect, including improved clinical outcome in a statistically significant manner; relief or alleviation of symptoms associated with a disease; reduced incidence of symptoms; improved quality of life; longer disease-free state; reduced extent of disease; stabilization of disease state; delayed progression of disease; remission; survival; or extended survival. When referring to an individual active ingredient administered alone, a therapeutically effective amount refers to the effect of that ingredient or cells expressing that ingredient alone. When referring to a combination, a therapeutically effective amount refers to the total amount of the active ingredient or supplementary active ingredients combined with the cells expressing the active ingredient that produce the therapeutic effect, regardless of whether the administration is performed sequentially, sequentially, or simultaneously.

[0104] Thus, in certain embodiments, a method is provided for treating a paramyxovirus (e.g., RSV, MPV, or PVM) infection in a subject, the method comprising administering to the subject an effective amount of an antibody, polynucleotide, vector, host cell, or composition disclosed herein.

[0105] The subjects that can be treated according to the present disclosure are generally human and other primate subjects for veterinary purposes, such as monkeys and apes.Other model organisms, such as mice and rats, can also be treated according to the present disclosure.In any of the above embodiments, the subject can be a human subject.The subject can be male or female, and can be of any suitable age, including infants, adolescents, young adults, adults, and geriatric subjects.

[0106] In certain embodiments, the human subject treated according to the present disclosure is an infant, a child, an adolescent, a young adult, a middle-aged adult, or an elderly person. In certain embodiments, the human subject treated according to the present disclosure is under 1 year old, or 1-5 years old, or 5-125 years old (e.g., 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, or 125 years old (including any age within or between the ranges). In certain embodiments, the human subject treated according to the present disclosure is 0-19 years old, 0-21 years old, 20 In some embodiments, the subject is a pediatric subject. Pediatric subjects include those who are 21 years of age or younger at the time of diagnosis or treatment. In certain embodiments, the human subject is 45-54 years of age, 55-64 years of age, 65-74 years of age, 75-84 years of age, or 85 years of age or older. In some embodiments, the human subject is male. In some embodiments, the human subject is female.

[0107] In certain embodiments, treatment is administered as a prophylaxis, either before or after exposure.

[0108] Thus, typical routes of administration of the compositions of the present disclosure include, but are not limited to, oral, topical, transdermal, inhalation, parenteral, sublingual, buccal, rectal, vaginal, and intranasal. The term "parenteral" as used herein includes subcutaneous injection, intravenous, intramuscular, intrasternal injection or infusion techniques. In certain embodiments, administering includes administering by a route selected from oral, intravenous, parenteral, intragastric, intrathoracic, intrapulmonary, intrarectal, intradermal, intraperitoneal, intratumoral, subcutaneous, topical, transdermal, intracisternal, intrathecal, intranasal, and intramuscular. In certain embodiments, the method includes orally administering the antibody, polynucleotide, vector, host cell, or composition to a subject.

[0109] Pharmaceutical compositions according to certain embodiments of the invention are formulated so that the active ingredients contained therein are bioavailable upon administration of the composition to a patient. The composition administered to a subject or patient can take the form of one or more dosage units, e.g., a tablet can be a single dosage unit, and a container of an antibody described herein in aerosol form can hold multiple dosage units. Actual methods of preparing such dosage forms will be known or apparent to those skilled in the art. See, e.g., Remington: The Science and Practice of Pharmacy, 20th Edition (Philadelphia College of Pharmacy and Science, 2000). The composition administered will, in any case, contain an effective amount of the antibody, polynucleotide, vector, host cell, or composition of the present disclosure to treat the disease or condition of interest according to the teachings herein.

[0110] The composition may be in solid or liquid form.In some embodiments, the carrier is particulate, so that the composition is in the form of, for example, a tablet or powder.The carrier may be liquid, so that the composition may be, for example, oral oil, injectable liquid, or aerosol, for example, useful for inhalation administration.When intended for oral administration, the pharmaceutical composition is preferably in either solid or liquid form, and the form considered as solid or liquid in this specification includes semi-solid, semi-liquid, suspension and gel form.

[0111] As a solid composition for oral administration, the pharmaceutical composition may be formulated into powder, granules, compressed tablets, pills, capsules, chewing gum, wafers, etc. Such solid compositions typically contain one or more inert diluents or edible carriers. In addition, one or more of the following may be present: binders such as carboxymethylcellulose, ethylcellulose, microcrystalline cellulose, tragacanth or gelatin; excipients such as starch, lactose or dextrin; disintegrants such as alginic acid, sodium alginate, primogel, corn starch; lubricants such as magnesium stearate or Sterotex; glidants such as colloidal silicon dioxide; sweeteners such as sucrose or saccharin; flavors such as peppermint, methyl salicylate or orange flavor; and colorants. When the composition is in the form of a capsule, for example a gelatin capsule, it may contain, in addition to the above-mentioned types of materials, liquid carriers such as polyethylene glycol or oil.

[0112] The composition may be in the form of liquid, for example, elixir, syrup, solution, emulsion, or suspension. The liquid may be for oral administration or for delivery by injection, as two examples. When intended for oral administration, the preferred composition contains one or more of sweeteners, preservatives, dyes / colorants, and flavor enhancers in addition to the compound of the present invention. The composition intended for administration by injection may contain one or more of surfactants, preservatives, wetting agents, dispersing agents, suspending agents, buffers, stabilizers, and isotonic agents.

[0113] Liquid pharmaceutical compositions may be in solution, suspension, or other similar form and may contain one or more of the following adjuvants: sterile diluents such as water for injection, saline, preferably saline, Ringer's solution, isotonic sodium chloride, synthetic mono- or diglycerides that can serve as a solvent or suspending medium, fixed oils such as polyethylene glycols, glycerin, propylene glycol, or other solvents; antibacterial agents such as benzyl alcohol or methylparabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates, or phosphates, and tonicity adjusters such as sodium chloride or glucose. Parenteral preparations can be enclosed in glass or plastic ampoules, disposable syringes, or multiple dose vials. Saline is the preferred adjuvant. Injectable pharmaceutical compositions are preferably sterile.

[0114] Liquid compositions intended for either parenteral or oral administration should contain an amount of the antibody disclosed herein such that a suitable dosage will be obtained. Typically, this amount is at least 0.01% of the antibody in the composition. When intended for oral administration, this amount may vary between 0.1 and about 70% by weight of the composition. Certain oral pharmaceutical compositions contain between about 4% and about 75% antibody. In certain embodiments, pharmaceutical compositions and formulations according to the invention are prepared such that a parenteral dosage unit contains 0.01 to 10% by weight of antibody before dilution.

[0115] The composition may be for topical administration, in which case the carrier may suitably comprise a solution, emulsion, ointment or gel base. For example, the base may comprise one or more of petrolatum, lanolin, polyethylene glycol, beeswax, mineral oil, diluents such as water and alcohol, and emulsifiers and stabilizers. A thickening agent may be present in the composition for topical administration. If intended for transdermal administration, the composition may comprise a transdermal patch or iontophoresis device. The pharmaceutical composition may be for rectal administration, for example, in the form of a suppository that melts in the rectum and releases the drug. The composition for rectal administration may contain an oily base as a suitable non-irritating excipient. Such bases include, but are not limited to, lanolin, cocoa butter, and polyethylene glycol.

[0116] The composition may include various materials that modify the physical form of the solid or liquid dosage unit. For example, the composition may include materials that form a coating shell around the active ingredient. The materials that form the coating shell are typically inert and may be selected from, for example, sugar, shellac, and other enteric coating agents. Alternatively, the active ingredient may be encapsulated in a gelatin capsule. The composition in solid or liquid form may include an agent that binds to the antibody of the present disclosure and thereby aids in the delivery of the compound. Suitable agents that may act in this capacity include monoclonal or polyclonal antibodies, one or more proteins, or liposomes. The composition may essentially consist of a dosage unit that can be administered as an aerosol. The term aerosol is used to refer to a variety of systems ranging from colloidal to systems consisting of pressurized packages. Delivery is by liquefied or compressed gas, or a suitable pump system that dispenses the active ingredient. The aerosol may be delivered in a single-phase, two-phase, or three-phase system to deliver the active ingredient. The delivery of the aerosol includes the necessary container, activator, valve, subcontainer, etc., which together may form a kit. One of ordinary skill in the art may determine the preferred aerosol without undue experimentation.

[0117] It is understood that the compositions of the present disclosure also encompass carrier molecules (e.g., lipid nanoparticles, nanoscale delivery platforms, etc.) for the polynucleotides described herein.

[0118] Pharmaceutical compositions may be prepared by methodologies well known in the pharmaceutical field. For example, compositions intended to be administered by injection can be prepared by combining a composition comprising the antibody described herein and, optionally, one or more of salts, buffers and / or stabilizers with sterile distilled water to form a solution. Surfactants may be added to facilitate the formation of a homogeneous solution or suspension. Surfactants are compounds that non-covalently interact with peptide compositions to facilitate the dissolution or homogeneous suspension of the antibody in an aqueous delivery system.

[0119] In general, appropriate dosages and treatment regimens provide a sufficient amount of the composition to provide therapeutic and / or prophylactic benefit (including improved clinical outcomes (e.g., longer disease-free and / or overall survival, or reduced severity of symptoms) as described herein). For prophylactic use, the dose should be sufficient to prevent, delay the onset of, or reduce the severity of disease associated with the disease or disorder. The prophylactic benefit of the compositions administered in accordance with the methods described herein can be determined by conducting preclinical and clinical studies (including in vitro and in vivo animal studies) and analyzing the data obtained therefrom by appropriate statistical, biological, and clinical methods and techniques, all of which can be readily performed by one of ordinary skill in the art.

[0120] The compositions are administered in an effective amount (e.g., to treat a paramyxovirus infection), which will vary depending on a variety of factors, including the activity of the particular compound used; the metabolic stability and duration of action of the compound; the subject's age, weight, general health, sex, diet; method and time of administration; excretion rate; drug combinations; the severity of the particular disorder or condition; and the subject being treated. In certain embodiments, administration of treatment with the formulations and methods of the present disclosure results in subjects exhibiting from about 10% up to about 99% reduction in one or more symptoms associated with the disease or disorder being treated, compared to placebo-treated or other suitable control subjects.

[0121] Generally, a therapeutically effective daily dose of an antibody is from about 0.001 mg / kg (i.e., 0.07 mg) to about 100 mg / kg (i.e., 7.0 g) (for a mammal weighing 70 kg); preferably, a therapeutically effective dose is from about 0.01 mg / kg (i.e., 0.7 mg) to about 50 mg / kg (i.e., 3.5 g) (for a mammal weighing 70 kg); more preferably, a therapeutically effective dose is from about 1 mg / kg (i.e., 70 mg) to about 25 mg / kg (i.e., 1.75 g) (for a mammal weighing 70 kg). For the polynucleotides, vectors, host cells, and related compositions of the present disclosure, the therapeutically effective amount may be different from that for antibodies.

[0122] In certain embodiments, the methods comprise administering the antibody, polynucleotide, vector, host cell, or composition to the subject 2, 3, 4, 5, 6, 7, 8, 9, 10 or more times.

[0123] In certain embodiments, the methods include administering an antibody, polynucleotide, vector, host cell, or composition to a subject multiple times, where a second or subsequent administration occurs about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 24, about 48, about 74, about 96 hours, or more, from the first or previous administration, respectively.

[0124] In certain embodiments, the methods involve administering the antibody, polynucleotide, vector, host cell, or composition at least once prior to the subject being infected with a paramyxovirus.

[0125] A composition comprising an antibody, polynucleotide, vector, host cell, or composition of the present disclosure may be administered simultaneously with, before, or after administration of one or more other therapeutic agents. Such combination therapy may include administration of a single pharmaceutical dosage formulation comprising a compound of the present invention and one or more additional active agents, as well as administration of a composition comprising an antibody of the present disclosure and each active agent in its own separate dosage formulation. For example, the antibody and other active agents described herein may be administered to a patient together in a single oral dosage composition, such as a tablet or capsule, or each agent may be administered in a separate oral dosage formulation. Similarly, the antibody and other active agents described herein may be administered to a subject together in a single parenteral dosage composition, such as saline or other physiologically acceptable solution, or each agent may be administered in a separate parenteral dosage formulation. When separate dosage formulations are used, the composition comprising the antibody and one or more additional active agents may be administered essentially simultaneously, i.e., simultaneously, or separately, staggered, i.e., sequentially and in any order. Combination therapy is understood to include all of these regimens.

[0126] In certain embodiments, combination therapy is provided that includes one or more antibodies (or one or more nucleic acids, host cells, vectors, or compositions) of the present disclosure and one or more anti-inflammatory agents and / or one or more antiviral agents. In certain embodiments, the one or more anti-inflammatory agents include corticosteroids, such as, for example, dexamethasone, prednisone, etc. In some embodiments, the one or more anti-inflammatory agents include cytokine antagonists, such as antibodies that bind to IL6 (such as siltuximab), or to IL-6R (such as tocilizumab), or to IL-1β, IL-7, IL-8, IL-9, IL-10, FGF, G-CSF, GM-CSF, IFN-γ, IP-10, MCP-1, MIP-1A, MIP1-B, PDGR, TNF-α, or VEGF. In some embodiments, anti-inflammatory agents such as ruxolitinib and / or anakinra are used. In some embodiments, the one or more antiviral agents include nucleotide analogs or nucleotide analog prodrugs, such as, for example, remdesivir, sofosbuvir, acyclovir, and zidovudine. In certain embodiments, the antiviral agent includes lopinavir, oseltamivir, peramivir, zanamivir, ritonavir, favipiravir, or any combination thereof. In some embodiments, the combination therapy includes leronlimab. The anti-inflammatory agent for use in the combination therapy of the present disclosure also includes nonsteroidal anti-inflammatory drugs (NSAIDS). It is understood that in such combination therapy, the one or more antibodies (or one or more nucleic acids, host cells, vectors, or compositions) and the one or more anti-inflammatory agents and / or the one or more antiviral agents can be administered in any order and sequence, or together.

[0127] In some embodiments, an antibody (or one or more nucleic acids, host cells, vectors, or compositions) is administered to a subject who has previously been administered one or more anti-inflammatory agents and / or one or more anti-viral agents. In some embodiments, one or more anti-inflammatory agents and / or one or more anti-viral agents are administered to a subject who has previously been administered an antibody (or one or more nucleic acids, host cells, vectors, or compositions).

[0128] In related aspects, uses of the antibodies, vectors, host cells, and compositions disclosed herein are provided.

[0129] In certain embodiments, an antibody, polynucleotide, vector, host cell, or composition is provided for use in a method of treating a paramyxovirus infection in a subject.

[0130] In certain embodiments, the antibodies, polynucleotides, vectors, host cells, or compositions are provided for use in methods of manufacturing or preparing a medicament for treating a Paramyxovirus infection in a subject.

[0131] [Table 2-1] [Table 2-2] [Table 2-3] [Table 2-4] [Table 2-5] [Table 2-6]

[0132] The present invention also provides the following non-limiting enumerated embodiments.

[0133] Embodiment 1. An antibody comprising: (i) two heavy chain polypeptides, each having the amino acid sequence of SEQ ID NO:1; and (ii) two light chain polypeptides, each having the amino acid sequence of SEQ ID NO:4. Embodiment 2: An antibody comprising: (i) two heavy chain polypeptides, each having the amino acid sequence of SEQ ID NO:2; and (ii) two light chain polypeptides, each having the amino acid sequence of SEQ ID NO:4. Embodiment 3: An antibody comprising: (i) two heavy chain polypeptides, each having the amino acid sequence of SEQ ID NO:3; and (ii) two light chain polypeptides, each having the amino acid sequence of SEQ ID NO:4. Embodiment 4: An antibody comprising: (i) two heavy chain polypeptides each comprising the amino acid sequence of SEQ ID NO:6; and (ii) two light chain polypeptides each comprising the amino acid sequence of SEQ ID NO:4. Embodiment 5: An antibody comprising: (i) two heavy chain polypeptides each comprising the amino acid sequence of SEQ ID NO:7; and (ii) two light chain polypeptides each comprising the amino acid sequence of SEQ ID NO:4. Embodiment 6: An antibody comprising: (i) two heavy chain polypeptides each comprising the amino acid sequence of SEQ ID NO:8; and (ii) two light chain polypeptides each comprising the amino acid sequence of SEQ ID NO:4. Embodiment 7: An isolated polynucleotide encoding: (i) the antibody of any one of embodiments 1-6; (ii) the heavy chain of the antibody of any one of embodiments 1-6; (iii) the light chain of the antibody of any one of embodiments 1-6; or (iv) the heavy chain of the antibody of any one of embodiments 1-6 and the light chain of the antibody of any one of embodiments 1-6. Embodiment 8. The polynucleotide of embodiment 7, wherein the polynucleotide comprises DNA or RNA, optionally wherein the RNA comprises mRNA, and optionally wherein the mRNA comprises modified nucleosides, a cap-1 structure, a cap-2 structure, or any combination thereof, and / or comprises pseudouridine, N6-methyladenosine, 5-methylcytidine, 2-thiouridine, or any combination thereof. Embodiment 9. A vector comprising the polynucleotide of embodiment 7 or 8. Embodiment 10. A host cell comprising the polynucleotide of embodiment 7 or 8, or the vector of embodiment 9. Embodiment 11. A composition comprising an antibody of any one of embodiments 1 to 6, a polynucleotide of embodiment 7 or 8, a vector of embodiment 9, and / or a host cell of embodiment 10, and a pharma- ceutically acceptable carrier, excipient, or diluent. Embodiment 12. A method for preventing a paramyxovirus infection in a subject, comprising administering to the subject an effective amount of an antibody of any one of embodiments 1 to 6; a polynucleotide of embodiment 7 or 8; a vector of embodiment 9; a host cell of embodiment 10; and / or a composition of embodiment 11. Embodiment 13. A method for treating a paramyxovirus infection in a subject, comprising administering to the subject an effective amount of an antibody of any one of embodiments 1 to 6; a polynucleotide of embodiment 7 or 8; a vector of embodiment 9; a host cell of embodiment 10; and / or a composition of embodiment 11. Embodiment 14 The method of embodiment 12 or 13, wherein the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof. Embodiment 15. The antibody of any one of embodiments 1-6, the polynucleotide of embodiment 7 or 8, the vector of embodiment 9, the host cell of embodiment 10, and / or the composition of embodiment 11 for use in a method of preventing a paramyxovirus infection in a subject, wherein optionally the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof. Embodiment 16. The antibody of any one of embodiments 1-6, the polynucleotide of embodiment 7 or 8, the vector of embodiment 9, the host cell of embodiment 10, and / or the composition of embodiment 11 for use in a method of treating a paramyxovirus infection in a subject, wherein optionally the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof. Embodiment 17. The antibody of any one of embodiments 1-6, the polynucleotide of embodiment 7 or 8, the vector of embodiment 9, the host cell of embodiment 10, and / or the composition of embodiment 11, for use in the manufacture of a medicament for preventing a paramyxovirus infection in a subject, wherein optionally the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof. Embodiment 18. The antibody of any one of embodiments 1-6, the polynucleotide of embodiment 7 or 8, the vector of embodiment 9, the host cell of embodiment 10, and / or the composition of embodiment 11, for use in the manufacture of a medicament for treating a paramyxovirus infection in a subject, wherein optionally the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof. Embodiment 19. A method of making the antibody of any one of embodiments 1-6, comprising culturing a host cell expressing the antibody under conditions and for a period of time sufficient to produce the antibody. Embodiment 20. The method of embodiment 19, further comprising isolating and / or purifying the antibody.

[0134] Working Example Example 1 Pharmacokinetic studies RSV_Ab neutralizes RSV, MPV, and PVM in vitro and in vivo in mice. In vivo studies were performed using cynomolgus monkeys to evaluate the stability of RSV_Ab (IgG1 with HC: SEQ ID NO:5; LC: SEQ ID NO:4) and RSV_Ab_MLNS (IgG1 with HC: SEQ ID NO:1; LC: SEQ ID NO:4). The study design is shown in Figure 1. The data are shown in Figures 2 and 3. As shown in Figure 2, after a dose of 15 mg / kg (slow intravenous infusion) RSV_Ab_MLNS, RSV_Ab_MLNS was present in plasma at concentrations greater than 20 micrograms per ml from 1000 hours post-dose onwards, including 1250 hours post-dose. In comparison, RSV_Ab was present at lower concentrations. As shown in Figure 3, RSV_Ab_MLNS showed superior stability over an extended period of time compared to RSV_Ab.

[0135] Example 2 Prophylactic and therapeutic use of RSV_AB_MLNS RSV_Ab_MLNS, RSV_Ab_GAALIE (HC: SEQ ID NO:2; LC: IgG1 comprising SEQ ID NO:4), or RSV_Ab_MLNS_GAALIE (HC: SEQ ID NO:3; LC: IgG1 comprising SEQ ID NO:4) are administered prophylactically to human subjects at risk of RSV infection and therapeutically to human subjects with RSV.

[0136] The various embodiments described above can be combined to provide further embodiments. All U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications, and non-patent publications mentioned herein and / or listed in the Application Data Sheet, including U.S. Patent Application No. 63 / 147,676, filed February 9, 2021, are incorporated herein by reference in their entirety. If necessary, aspects of the embodiments can be modified to use concepts from various patents, applications, and publications to provide further embodiments.

[0137] These and other changes can be made to the embodiments in light of the above detailed description. In general, the terms used in the following claims should not be construed to limit the claims to the specific embodiments disclosed in the specification and the claims, but rather to include all possible embodiments, along with the full scope of equivalents to which such claims are entitled. Accordingly, the claims are not limited by this disclosure.

Claims

1. Antibodies including: (i) two heavy chain polypeptides, each having the amino acid sequence of SEQ ID NO:1; and (ii) two light chain polypeptides, each having the amino acid sequence of SEQ ID NO:

4.

2. Antibodies including: (i) two heavy chain polypeptides, each having the amino acid sequence of SEQ ID NO:2; and (ii) two light chain polypeptides, each having the amino acid sequence of SEQ ID NO:

4.

3. Antibodies including: (i) two heavy chain polypeptides, each having the amino acid sequence of SEQ ID NO:3; and (ii) two light chain polypeptides, each having the amino acid sequence of SEQ ID NO:

4.

4. Antibodies including: (i) two heavy chain polypeptides each comprising the amino acid sequence of SEQ ID NO:6; and (ii) two light chain polypeptides each comprising the amino acid sequence of SEQ ID NO:

4.

5. Antibodies including: (i) two heavy chain polypeptides each comprising the amino acid sequence of SEQ ID NO:7; and (ii) two light chain polypeptides each comprising the amino acid sequence of SEQ ID NO:

4.

6. Antibodies including: (i) two heavy chain polypeptides each comprising the amino acid sequence of SEQ ID NO:8; and (ii) two light chain polypeptides each comprising the amino acid sequence of SEQ ID NO:

4.

7. An isolated polynucleotide encoding: (i) the antibody according to any one of claims 1 to 6; (ii) a heavy chain of the antibody according to any one of claims 1 to 6; (iii) the light chain of an antibody according to any one of claims 1 to 6; or (iv) The heavy chain of an antibody according to any one of claims 1 to 6 and the light chain of an antibody according to any one of claims 1 to 6.

8. 8. The polynucleotide of claim 7, wherein said polynucleotide comprises DNA or RNA, optionally wherein said RNA comprises mRNA, and further optionally wherein said mRNA comprises modified nucleosides, a cap-1 structure, a cap-2 structure, or any combination thereof, and / or comprises pseudouridine, N6-methyladenosine, 5-methylcytidine, 2-thiouridine, or any combination thereof.

9. A vector comprising the polynucleotide of claim 7 or 8.

10. A host cell comprising a polynucleotide according to claim 7 or 8 or a vector according to claim 9.

11. A composition comprising an antibody according to any one of claims 1 to 6, a polynucleotide according to claim 7 or 8, a vector according to claim 9, and / or a host cell according to claim 10, and a pharma- ceutically acceptable carrier, excipient, or diluent.

12. 13. A pharmaceutical composition comprising the antibody of any one of claims 1 to 6, the polynucleotide of claim 7 or 8, the vector of claim 9, the host cell of claim 10, and / or the composition of claim 11 for use in preventing a paramyxovirus infection in a subject, wherein optionally the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof.

13. 13. A pharmaceutical composition comprising the antibody of any one of claims 1 to 6, the polynucleotide of claim 7 or 8, the vector of claim 9, the host cell of claim 10, and / or the composition of claim 11 for use in treating a paramyxovirus infection in a subject, wherein optionally the paramyxovirus comprises respiratory syncytial virus, metapneumovirus, pneumonia virus of mice, or any combination thereof.

14. 13. Use of an antibody according to any one of claims 1 to 6, a polynucleotide according to claim 7 or 8, a vector according to claim 9, a host cell according to claim 10, and / or a composition according to claim 11, in the manufacture of a medicament for preventing a paramyxovirus infection in a subject, optionally wherein the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof.

15. 13. Use of an antibody according to any one of claims 1 to 6, a polynucleotide according to claim 7 or 8, a vector according to claim 9, a host cell according to claim 10, and / or a composition according to claim 11, in the manufacture of a medicament for treating a paramyxovirus infection in a subject, optionally wherein the paramyxovirus comprises a respiratory syncytial virus, a metapneumovirus, a pneumonia virus of mice, or any combination thereof.

16. A method of making an antibody according to any one of claims 1 to 6, comprising culturing a host cell expressing the antibody under conditions and for a period of time sufficient to produce the antibody.

17. 17. The method of claim 16, further comprising isolating and / or purifying the antibody.

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