Treatment of infectious disease

Abstract

The disclosure relates to treating and preventing coronavirus infections, in particular SARS-CoV-2. The disclosure also relates to treating or preventing cardiovascular symptoms associated with coronavirus infections, including blood clotting, heart attack, stroke, pulmonary embolism and / or thrombosis. The disclosure further relates to antibodies which may be used in such methods.

Description

[0001] TREATMENT OF INFECTIOUS DISEASE

[0002] Field of the Invention

[0003] The invention relates to methods of preventing and / or treating coronavirus infection which utilise CD91 as a therapeutic target, and agents for use in such methods. Specifically, the methods involve disrupting the interaction between CD91 and the coronavirus spike protein, by providing agents which either bind CD91 or reduce its expression. The invention also provides antibodies which bind CD91.

[0004] Background to the invention

[0005] The global health burden caused by coronavirus diseases has been appreciated in recent years, particularly with reference to the pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). The disease caused by SARS-CoV- 2 has become known as COVID-19, and has been primarily described as a lung disease (1). However, infection with SARS-CoV-2 can cause multi-organ dysfunction including acute kidney injury, acute cardiac injury, coagulopathy, thromboembolic complications including stroke and pulmonary embolism, and circulatory shock (3, 4). Cardiovascular complications are emerging as a significant threat in COVID-19 patients, with endothelial damage and clotting occurring in severe and critical cases of severe acute respiratory syndrome (SARS). It is now established that disseminated intravascular coagulation (DIC) and consumption coagulopathy(5) occur in severe cases of COVID-19, with a high risk of mortality in patients with underlying cardiovascular conditions (6).

[0006] The SARS-CoV-2 genome encodes four structural proteins known as the spike (S), envelope (E), nucleocapsid (N), and membrane (M) proteins, which are incorporated into virions. The S protein of SARS-CoV-2 contains a receptor-binding domain (S-RBD) which recognises the host’s receptor and mediates receptor recognition / membrane fusion (2). The presence of the spike protein responsible for host cell entry is characteristic of coronaviruses in general, therefore a means of preventing the spike protein from interacting with host cell receptors would have broad applicability in modifying coronavirus infection in general, not merely SARS-CoV-2.

[0007] SARS-CoV-2 is known to use Angiotensin Converting Enzyme II (ACE2) as an entry receptor (7) in the host cells of human tissues including the lungs, heart, kidneys, liver and gastrointestinal tract. ACE2 expression has been found undetectable in endothelial cells (8). Yet, the endothelium gets infected by SARS-CoV-2, which may severely impact the cardiovascular system, and lead to heart attack, stroke, pulmonary embolism, or thrombosis. The mechanisms of endothelial infection by SARS-CoV-2 were previously unknown (9).

[0008] CD91, also known as LRP1, is a large endocytic receptor known to be involved in several cellular processes, such as cell migration, survival, motility, proliferation and differentiation. CD91 is ubiquitously expressed in the cells including immune cells, hepatocytes, epithelial cells, neurons, adipocytes and endothelial cells. CD91 has been reported to play an important role in the endothelium by coordinating angiogenesis and maintaining the blood-brain barrier (17-19). Thus, CD91 is associated to different pathologies including thrombosis, fibrinolysis, atherosclerosis, and Alzheimer’s. So far, it has not been established whether CD91 controls cellular entry and binding of SARS-CoV- 2. Therefore, the role of CD91 in COVID-19 is unknown.

[0009] Treatment with aprotinin (23) or lactoferrin (24) improved COVID-19 patients’ outcomes. However, the mechanism behind the observed effects was not known. Indeed, the observed effects were attributed to various mechanisms: preventing interaction between the spike protein and the ACE2 receptor, inducing an interferon (IFN) response, inducing mediators of, or signalling factors associated with, innate immunity such as toll-like receptors (TLRs), or inhibiting inflammation and thereby preventing immunopathology.

[0010] The present inventors are the first to demonstrate that the CD91 receptor is potentially occupied by SARS-CoV-2, and coronavirus spike proteins in general, thus identifying CD91 as a useful target for treating or preventing coronavirus.

[0011] Brief Description of the Figures

[0012] Figure 1 : Protein-protein docking of SARS-CoV-2 spike protein to CD91. (A) 7L4Z (SARS-CoV-2-spike) structure with a molecular surface depicting lipophillicity, where hydrophilic and lipophilic areas are depicted in dark grey and neutral in light grey. 1CR8 (CD91 / LRP1) is pictured in its molecular form to show how the atoms fits into the SARS-CoV-2 spike protein. (B) 1CR8 (LRP1 / CD91) and 7L4Z (Covid-spike) with H- bond in the interaction site shown between CD91 CYS41 and CoV-spike ARG466. Alpha helices are shaded using horizontal line hatching. Beta sheets are shaded using circular dots. Loops are shaded using diagonal line hatching. (C) A bar chart showing the Best and Average London G binding scores of the unmutated CD91 protein (1CR8) and each mutated LRP1 protein through protein-protein docking with the Covid spike protein (7L4Z). (D) 1CR8 (LRP1) surface shown in light grey and 7L4Z (Spike) molecular surface shown in dark grey, at the interaction site. (E) Spike (dark grey) and LRP1 (light grey) shown in their best pose determined using protein-protein docking, performed as described in the Examples.

[0013] Figure 2: CD91 facilitates the cellular entry of SARS-CoV-2 pseudotyped particles. Figure shows Pseudovirus infection of HEK-293T cell transduced with constructs encoding the specified proteins. Infection data is expressed as mean relative luciferase units (RLU).

[0014] Figure 3: CD91 knockdown affects SARS-CoV-2 replication. Viral RNA levels measured by qPCR in HUVECs cells depleted for LRP1 Infected by SARS-CoV-2 at a multiplicity of infection (MOI) of 1 at 24 hrs post-infection.

[0015] Brief description of the sequence listing

[0016] SEQ ID NO: 1 shows the CD91 sequence from Uniprot ID Q07954

[0017] SEQ ID NO: 2 shows the sequence of PDB entry 1CR8

[0018] SEQ ID NO: 3 shows the sequence of the SARS CoV-2 spike protein receptor binding domain from PDB entry 7L4Z

[0019] SEQ ID NO: 4 shows the sequence of the SARS-CoV-2 spike protein from Uniprot ID P0DTC2

[0020] Summary of the invention

[0021] The inventors have discovered for the first time that the receptor binding domain of the coronavirus spike protein binds to CD91, and that CD91 contributes to coronavirus infection. The inventors have structurally characterised the binding of CD91 to the coronavirus spike protein, and identified the key amino acid residues which facilitate the interaction.

[0022] The invention provides a method of treating and / or preventing coronavirus infection, the method comprising administering a therapeutically effective amount of an agent to a subject, wherein the agent binds to CD91 and wherein the agent is not lactoferrin or aprotinin.

[0023] The invention also provides an agent that binds to CD91 for use in a method of treating and / or preventing coronavirus infection, wherein the method comprises administering a therapeutically effective amount of the agent to a subject, and wherein the agent is not lactoferrin or aprotinin.

[0024] The invention also provides use of an agent that binds to CD91 in the manufacture of a medicament for treating and / or preventing coronavirus infection, wherein the agent is not lactoferrin or aprotinin.

[0025] In another aspect, the invention also provides a method of treating or preventing coronavirus infection, the method comprising administering a therapeutically effective amount of an agent to a subject, wherein the agent reduces the expression of CD91.

[0026] The invention also provides an agent that reduces the expression of CD91 for use in a method of treating and / or preventing coronavirus infection, wherein the method comprises administering a therapeutically effective amount of the agent to a subject.

[0027] The invention also provides use of an agent that reduces the expression of CD91 in the manufacture of a medicament for treating and / or preventing coronavirus infection.

[0028] The invention also provides an antibody capable of binding to CD91. The antibody may bind to the same epitope of CD91 as the coronavirus spike protein wherein the epitope is determined by x-ray crystallography. The antibody may bind to an epitope comprising one or more amino acids selected from the group consisting of positions 1060, 1061, 1065- 1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090-1092, 1095-1099 of SEQ ID NO: 1. In addition to comprising one or more of these amino acids, the epitope may also comprise residues in other domains of CD91.

[0029] The invention also provides polynucleotides encoding the antibody, vectors comprising or encoding said polynucleotide, host cells and pharmaceutical compositions. The invention also provides a method of making an antibody of the invention, wherein the method comprises culturing the host cell comprising the polynucleotide or vector of the invention under conditions permitting production of the antibody and recovering the antibody so produced. Detailed Description of the Invention

[0030] It is to be understood that different applications of the disclosed products and methods may be tailored to the specific needs in the art. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments of the invention only, and is not intended to be limiting.

[0031] In addition, as used in this specification and the appended claims, the singular forms “a”, “an”, and “the” include plural referents unless the content clearly dictates otherwise. Thus, for example, reference to “an amino acid sequence” includes two or more such sequences, and the like.

[0032] A therapeutically effective dose of the agent for use as described herein will generally provide therapeutic benefit without causing substantial toxicity. Toxicity and therapeutic efficacy of an agent for use or antibody as described herein can be determined by standard pharmaceutical procedures in cell culture or experimental animals. Cell culture assays and animal studies can be used to determine the LD50 (the dose lethal to 50% of a population) and the ED50 (the dose therapeutically effective in 50% of a population). The dose ratio between toxic and therapeutic effects is the therapeutic index, which can be expressed as the ratio LD50 / ED50. An agent for use or antibody that exhibits large therapeutic indices is preferred. In one embodiment, the agent for use according to the invention, or the antibody of the invention, exhibits a high therapeutic index. The data obtained from cell culture assays and animal studies can be used in formulating a range of dosages suitable for use in humans. The dosage lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon a variety of factors, e.g., the dosage form employed, the route of administration utilized, the condition of the subject.

[0033] As used herein, the terms “polypeptide,” “peptide,” and “protein” are used interchangeably herein to refer to polymers of amino acids of any length. The polymer may be linear or branched, it may comprise modified amino acids, and it may be interrupted by non-amino acids. The terms also encompass an amino acid polymer that has been modified; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, pegylation, or any other manipulation, such as conjugation with a labeling component. As used herein, the term “amino acid” includes natural and / or unnatural or synthetic amino acids, including glycine and both the D or L optical isomers, and amino acid analogues, and peptidomimetics.

[0034] As used herein, the term “modulate” is meant to refer to any change in biological state, i.e., increasing or decreasing.

[0035] All methods described herein are performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. In regard to any of the methods provided, the steps of the method may occur simultaneously or sequentially. When the steps of the method occur sequentially, the steps may occur in any order, unless noted otherwise. In cases in which a method comprises a combination of steps, each and every combination or sub-combination of the steps is encompassed within the scope of the disclosure, unless otherwise noted herein.

[0036] “Sequence identity” as used herein in the context of two or more polypeptide or polynucleotide sequences, means that the sequences have a specified percentage of residues that are the same over a specified region that is determined using a standard algorithm, having regard to the extent that sequences are identical over a window of comparison. Typically, identity is assessed over the full length of the sequence. Identity or sequence identity may be determined using computer algorithms such as GAP, BESTFIT, FASTA and the BLAST family of programs as for example disclosed by Altschul et al., 1997, Nucl. Acids Res. 25:3389. A detailed discussion of sequence analysis can be found in Unit 19.3 of CURRENT PROTOCOLS IN MOLECULAR BIOLOGY Eds. Ausubel et al. (John Wiley & Sons Inc NY, 1995-1999).

[0037] A “conservative amino acid substitution” is one in which an amino acid residue is substituted by another amino acid residue having a side chain (R group) with similar chemical properties (e.g., charge or hydrophobicity). In general, a conservative amino acid substitution will not substantially change the functional properties of a protein. Accordingly, conservative substitutions in the amino acid sequences described herein may not significantly change the way in which said amino acids interact with other amino acids. In cases where two or more amino acid sequences differ from each other by conservative substitutions, the percent or degree of similarity may be adjusted upwards to correct for the conservative nature of the substitution. Means for making this adjustment are well known to those of skill in the art. See, e.g., Pearson (1994) Methods Mol. Biol. 24: 307-331, which is herein incorporated by reference. Reference to “significant” in the context of a difference, for example, a “significant increase”, “significant decrease” or “significant difference”, refers to a statistically significant difference, as determined using an appropriate statistical test. Appropriate statistical tests will be known to those skilled in the art.

[0038] The term “about”, as used herein, has its ordinary meaning of approximately. If the degree of approximation is not otherwise clear from the context, “about” means either within plus or minus 10% of the provided value, or rounded to the nearest significant figure. In some embodiments, the term “about” indicates the designated value ± up to 10%, up to ± 5%, or up to ± 1%.

[0039] All publications, patents and patent applications cited herein, whether supra or infra, are hereby incorporated by reference in their entirety.

[0040] Agents which bind CD91

[0041] CD91 is a multifunctional receptor that binds a wide variety of protein ligands - more than 75 molecules. CD91 is also known as A2MR, APOER, APR, IGFBP3R, LRP, LRP1, LRP1A, TGFBR5, low density lipoprotein receptor-related protein 1, LDL receptor related protein 1, KPA, IGFBP3R1 and IGFBP-3RLRP1. These names may be used interchangeably herein.

[0042] CD91 comprises an extracellular alpha-chain and an intracellular beta-chain. The alpha (a) chain contains four ligand-binding domains (clusters I-IV) consisting of 2, 8, 10, and 11 acidic cysteine-rich complement-type repeats (CRs) respectively, which are responsible for the majority of ligand binding to CD91. The clusters are followed by EGF homology domains, with cysteine rich EGF repeats. The intracellular beta chain contains two NPxY motifs that are required for endocytosis and multiple signalling pathways (15). Interestingly, the receptor-associated protein (RAP) inhibits binding of all known ligands of CD91 and acts too as a chaperone by preventing receptor aggregation and degradation in the endoplasmic reticulum (16).

[0043] The structure of the human CD91 protein is available, for example, as ID Q407954 in the Uniprot database. This sequence is shown in SEQ ID NO: 1. Entry 1CR8 in the protein data bank (PDB) provides the sequence of the complement repeat 8 (CR8). The 1CR8 sequence was used in the examples below. Moreover, the sequence of the human CD91 gene is available from the NUT GenBank with GenelD No: 4035. The invention provides a method of treating and / or preventing coronavirus infection, wherein the method comprises administering a therapeutically effective amount of an agent which binds CD91 to a subject. In preferred embodiments, CD91 in the context of the invention is human CD91. The CD91 may have the sequence of SEQ ID NO: 1. However, variants are also encompassed, for example naturally occurring variants. Such variants typically retain at least about 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94% or 95% identity to SEQ ID NO: 1 (or even about 96%, 97%, 98% or 99% identity). In other words, such variants may retain about 60% - about 99% identity to SEQ ID NO: 1, suitably about 80% - about 99% identity to SEQ ID NO: 1, more suitably about 90% - about 99% identity to SEQ ID NO: 1 and most suitably about 95% - about 99% identity to SEQ ID NO: 1.

[0044] The agent may be a protein ligand of CD91 (i.e. a protein that interacts with CD91), a small molecule or an antibody or antigen-binding fragment thereof.

[0045] Ligands of CD91 include A2 -Macroglobulin, P-amyloid precursor protein, APBB1, APOE, Aprotinin, CIS / Clq inhibitor, CALR, CD44, chylomicron, collectin, complement C3, CTGF, DLG4, elastase, factor IXa, factor Vila, fibronectin, gentamicin, GIPC1, heat shock proteins: gp96, hsp70, hsp90, heparin cofactor II, hepatic lipase, ITGB1BP1, lactoferrin, lipoprotein lipase, LPL, MAPK8IP1, MAPK8IP2, midkine, MMP13, MMP2, MMP9, neuroserpin, nexin-1, NOS1AP, PAI 2, PAI-1, PDGF, tPA, Upa, polymyxin B, protein C inhibitor, pseudomonas endotoxin A, receptor-associated protein (RAP), serpin- derived small peptide (SP16), SHC1, sphingolipid activator protein, SYNJ2BP, tat, thrombin, THBS1, thrombospondin 2, TEMPI, TIMP2, TIMP3, tissue factor pathway inhibitor, PLAT, transforming growth factor P, PLAUR and VLDL. The ligand may be one of these proteins, with the exception of lactoferrin and aprotinin. The agent could also be a non-naturally occurring protein ligand.

[0046] In preferred embodiments, the agent does not itself modulate signalling through the CD91 receptor. In such embodiments, binding of the agent does not alter the quantity or quality of signalling through the CD91 receptor, relative to its typical signalling pattern when the agent is not present. In other words, the agent does not interfere with the natural signalling pathway through the receptor. In these instances, the agent may not significantly modulate the binding of endogenous ligand to CD91. An endogenous ligand refers to a ligand present in the subject under normal physiological conditions, i.e. when the subject is healthy and uninfected. There may, for example, be less than a 10% increase or decrease in signalling through the receptor when the agent is present compared to when the agent is absent.

[0047] In some embodiments, the agent may itself activate signalling to the same or similar degree as an endogenous ligand of the receptor, such as lactoferrin. Once again, it is preferred that the overall degree of signalling through the receptor is the same, or there is less than a 10% increase or decrease in signalling, when the agent is present compared with when the agent is absent. Such changes in signalling may be determined by routine methods in the art. In other embodiments, the agent modulates signalling through the CD91 receptor.

[0048] In some embodiments, the agent is an antagonist of CD91. In such embodiments, the agent binds to CD91, but does not activate signalling through the receptor. In other embodiments, the agent is an agonist or a partial agonist of CD91. In such embodiments, the agent activates signalling through CD91 when binding the receptor. In preferred embodiments, the agent may activate signalling to the same or similar degree as an endogenous ligand of the receptor, such as lactoferrin. An endogenous ligand refers to a ligand present in the subject under normal physiological conditions, i.e. when the subject is healthy and uninfected. The agent may activate signalling to a lesser degree than an endogenous ligand of the receptor, for example lactoferrin. The agent may activate signalling to the greater degree than a natural or endogenous ligand of the receptor.

[0049] In preferred embodiments, the agent binds CD91 without affecting signalling.

[0050] The agent treats or prevents coronavirus infection by reducing the ability of the coronavirus spike protein to bind CD91 and enter host cells within the subject. In preferred embodiments, the agent prevents the receptor binding domain of the coronavirus spike protein from binding to CD91. In other instances, the agent inhibits the receptor binding domain of the coronavirus spike protein from binding to CD91. Typically, the agent competitively inhibits the binding of the coronavirus spike protein’s receptor binding domain to CD91. Inhibition may be determined using any technique known in the art. Experiments may for example be carried out at varying concentrations of the agent and / or the coronavirus spike protein. An agent of the invention is typically capable of inhibiting binding of the coronavirus spike protein to CD91 by at least 50%, at least 60%, at least 70%, at least 80% or at least 90%. In some embodiments, the agent will bind to the alpha chain of CD91. In some embodiments, the agent binds to an epitope comprising residues within cluster I, cluster II, cluster III or cluster IV of the alpha chain of CD91, or any combination thereof. In particular, the agent may bind to residues from more than one cluster. In particular embodiments the agent binds an epitope comprising the clusters of cysteine-rich, complement-type repeats (CRs). In preferred embodiments, the agent binds to an epitope comprising residues in Cluster II. Preferably, the agent binds to complement repeat 8, an epitope comprising complement repeat 8, or an epitope comprising residues within complement repeat 8.

[0051] Binding refers to the formation of a stable complex between multiple molecules. In some embodiments, a complex may be considered stable if it has a dissociation constant (KD) of 500 nM, less than 300 nM, less than 200 nM, less than 100 nM, less than 90 nM, less than 80 nM, less than 70 nM, less than 60 nM, less than 50 nM, less than 40 nM, less than 30 nM, less than 20 nM, less than 10 nM, less than 5 nM, less than 4 nM, less than 3 nM, less than 2 nM, less than 1 nM or less than 0.5 nM, as measured in a surface plasmon resonance assay.

[0052] Interaction of molecules or residues thereof refers to the formation or existence of intermolecular forces between said molecules or residues. Intermolecular forces may include ionic, covalent, co-ordinate or hydrogen bonds.

[0053] In some embodiments, the agent binds the same epitope on CD91 as the coronavirus spike protein, or binds to an overlapping epitope. Such binding may be determined by x-ray crystallography. In both cases, the agent prevents or inhibits binding of the spike protein to CD91 as described above.

[0054] In preferred embodiments, the agent binds the same epitope on CD91, or an overlapping epitope, as the receptor binding domain of the spike protein of SARS-CoV-2.

[0055] Therefore, wherein the agent is a protein or antibody, the agent may exhibit homology or sequence identity with the receptor binding domain of the coronavirus spike protein, and preferably, the binding region of the coronavirus spike protein. In some embodiments, the protein or antibody may have an amino acid sequence at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the receptor binding region of the coronavirus spike protein, preferably to the SARS-CoV-2 spike protein’s receptor binding region. The binding region of the coronavirus spike protein for CD91 may be identified by protein-protein docketing of the coronavirus spike protein with CD91. The crystal structures of the coronavirus spike protein and CD91 may be obtained from the protein data bank (PDB).

[0056] In preferred embodiments, the agent is capable of binding to residues of CD91 which interact with the binding region of the coronavirus spike protein. The examples of the present application show that amino acids 2, 3, 7-10, 12, 13, 20, 21, 23, 24, 32-34, 37- 41 of SEQ ID NO: 2 (corresponding to amino acids 1060, 1061, 1065-1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090-1092, 1095-1099 of SEQ ID NO: 1) are likely to be involved in the interaction of CD91 with the SARS-CoV-2 spike protein.

[0057] In particular embodiments, the agent may bind to or interact with at least one amino acid selected from 1060, 1061, 1065-1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090- 1092, 1095-1099, wherein the residue numbering corresponds to CD91 of SEQ ID NO: 1. In some instances, the agent may bind to or interact with at least two, at least three, at least four, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19 or all 20 of these residues. All combinations are contemplated.

[0058] In preferred embodiments, the agent binds to / interacts with at least Cl 097 of SEQ ID NO: 1. In some instances, the agent binds to / interacts with C1097 of SEQ ID NO: 1 and at least 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10 of the residues identified above.

[0059] Although these residues are provided for a particular sequence of human CD91, the skilled person could readily extrapolate the positions of these residues to other CD91 sequences, for example naturally occurring and non-naturally occurring variants, using routine techniques. The skilled person could also readily extrapolate the positions e.g. to the mouse or rat sequences of CD91. Agents binding to epitopes comprising the corresponding residues within these other CD91 sequences are therefore also provided by the invention.

[0060] The amino acid residues to which the agent binds may be determined by x-ray crystallography. For example residues may, in particular, be identified in this way by determining residues on CD91 within 5 A, preferably within 4 A of the agent. Binding residues may also be determined for example using mutational analysis, such as substitution of the residues with alanine. In some of any of the aforementioned embodiments, said interaction may be the formation or existence of ionic, covalent, co-ordinate or hydrogen bonds. In particular, binding of the agent may prevent the formation of hydrogen bonds between CD91 and the coronavirus spike protein, such as the hydrogen bond formed between Cys 1097 of CD91 (numbering according to SEQ ID NO: 1) and the coronavirus spike protein. Specifically, the agent may prevent or disrupt the formation of a hydrogen bond between Cys 1097 of CD91 and Arg466 of SARS-CoV-2 spike protein (numbering according to the sequence from UniProt ID P0DTC2, also set out in SEQ ID NO: 4), for example, by itself interacting with Cys 1097 of CD91.

[0061] In some embodiments, the agent comprises or consists of a protein. In some embodiments, the agent comprises or consists of a small molecule. In some embodiments, the agent comprises or consists of an antibody. In preferred embodiments, the agent comprises or consists of a protein or an antibody, and such embodiments are described in more detail herein. In some embodiments, the agent comprises or consists of a nucleic acid. In some embodiments, the nucleic acid comprises or consists of RNA. In some embodiments, the nucleic acid comprises or consists of DNA. In particular embodiments, the agent comprises or consists of an aptamer. In some embodiments, the aptamer comprises or consists of DNA. In some embodiments, the aptamer comprises or consists of RNA.

[0062] Nucleic acid aptamers binding CD91 may be prepared by systematic evolution of ligands by exponential enrichment (SELEX). Aptamers may be screened for binding to the residues described above using routine techniques in the art. Aptamers may also be screened for the prevent! on / inhibiti on of coronavirus spike protein binding to CD91 using routine methods.

[0063] In preferred embodiments, the agent selectively or specifically binds CD91. By “selectively” binding CD91, the agent has a greater affinity for CD91 than other molecules, such as ligands or receptors. Accordingly, said agent has a greater tendency to form stable complexes with CD91 than other molecules. However, the agent may still bind or interact with molecules which are not CD91.

[0064] By “specifically” binding CD91, the agent does not form stable complexes with molecules other than CD91 or fragments thereof, or molecules with extensive sequence similarity thereto. Agent which reduce the expression of CD91

[0065] In another aspect, the invention also provides a method of preventing or treating coronavirus infection, wherein the method comprises administering a therapeutically effective amount of an agent to a subject, and wherein the agent reduces the expression of CD91. The expression of CD91 may be reduced at the nucleic acid level, for example, by reducing the transcription of the CD91 gene, or promoting the degradation of mRNA encoding CD91. The agent may reduce the amount, level, or copy number of CD91 mRNA. The agent may be an siRNA which targets CD91 mRNA, by exhibiting complementarity to CD91 mRNA. In some embodiments, the agent may reduce the expression of CD91 at the protein level. For example, the agent may reduce expression at the protein level by reducing the translation efficiency of CD91 mRNA, or promoting degradation of the CD91 protein.

[0066] In preferred embodiments, the agent selectively or specifically reduces the expression of CD91 relative to other genes. By selectively reducing the expression of CD91, the agent reduces the expression of CD91 to a greater extent than it may modulate the expression of other genes. By specifically reducing the expression of CD91, the agent reduces the expression of CD91 without significantly modulating the expression of other genes. In preferred embodiments, the agent specifically reduces the expression of CD91 on the surface of cells. The agent may cause CD91 to be retained intracellularly. The agent may cause CD91 to be degraded. For example, the agent may promote the delivery of CD91 protein to a lysosome.

[0067] In some embodiments, the agent reduces the expression of CD91 in the endothelium or in endothelial cells. The agent may selectively or specifically reduce expression of CD91 in the endothelium or endothelial cells. By “selectively” reducing the expression of CD91 in the endothelium, the agent may reduce the expression of CD91 in the endothelium to a greater extent than in non-endothelial cells. Accordingly, the agent may reduce the expression of CD91 in tissues containing endothelial cells to a greater extent than tissues which do not contain, or contain a lower proportion of, endothelial cells. The agent may reduce the expression of CD91 in the lungs or in blood vessels, for example. By “specifically” reducing the expression in the endothelium, the agent may reduce the expression of CD91 only in endothelial cells, and not significantly decrease expression in other cell types. The agent may comprise or encode an inhibitor of CD91 expression, or specifically CD91 surface expression, which in under the control of a tissuespecific promoter, which causes the inhibitor to selectively or specifically be expressed or take effect in the endothelium.

[0068] In some embodiments, the agent is administered to the subject prophylactically, i.e. before they have become infected with the coronavirus. In particular embodiments, the agent is administered before the subject has come into contact, or is suspected to have come into contact, with the coronavirus. In some embodiments, the agent is administered after the subject has come into contact, or suspected to have come into contact with, the coronavirus.

[0069] Therapeutic effects

[0070] In some embodiments, treatment or prevention of coronavirus infection results in a decrease in viral replication or viral load in the subject. Viral load or replication can be estimated by quantifying the concentration of components of the virion in a sample obtained from a subject. In some embodiments, the sample may be obtained by a nasopharyngeal swab. In some embodiments, the sample obtained from the subject may be blood or plasma.

[0071] In some embodiments, treatment or prevention of coronavirus infection may result in a significantly reduced quantity of coronavirus RNA detected in the sample. In such embodiments, detection of coronavirus RNA may be by quantitative PCR, such as reverse transcription quantitative PCR (RT-qPCR). In some embodiments, treatment or prevention of coronavirus infection may result in a significantly reduced quantity of coronavirus proteins detected in the sample. In a particular embodiment, treatment or prevention may result in a decreased quantity of coronavirus spike protein which may be detected in a sample obtained from the subject. Said protein may be detected by ELISA, a lateral flow test or mass spectrometry, for example.

[0072] In some embodiments, treatment or prevention of coronavirus infection may result in a reduced number of infectious virions detected in a sample obtained from the subject. Methods of quantifying the number of infectious virions in a sample are known in the art, such as plaque assays (Mendoza, E. J., Manguiat, K., Wood, H., & Drebot, M. (2020). Two Detailed Plaque Assay Protocols for the Quantification of Infectious SARS-CoV-2. Current protocols in microbiology, 57(1), ecpmcl05).

[0073] In some embodiments, the treatment or prevention may result in the amelioration or the reduction in severity, duration and / or frequency of adverse effects associated with coronavirus infection. In some embodiments, the adverse effects may be signs or symptoms associated with the coronavirus infection. Symptoms refers to an effect of the coronavirus infection which is experienced only by the infected subject. Signs refer to an effect of the coronavirus infection which may be observed by an individual other than the infected subject, such as by a medical practitioner. Symptoms which may be alleviated by administration of the agent for use according to the invention may comprise fatigue, myalgia, abdominal pain, muscle ache, headache, nausea, shortness of breath, sore throat and loss of appetite. Signs which may be alleviated by administration of the agent for use according to the invention may comprise a continuous cough, vomiting, diarrhoea and fever.

[0074] In some embodiments, the adverse effects associated with coronavirus infection may be cardiovascular adverse effects, immunological adverse effects and / or respiratory adverse effects. Cardiovascular adverse effects refer to pathological effects caused by coronavirus infection of the cardiovascular system, and / or occurring within the cardiovascular system, namely, the heart and vasculature. The cardiovascular adverse effects may comprise disorders related to or caused by blood clotting or thrombosis. In particular embodiments, the cardiovascular adverse effects may be heart attack, stroke and / or pulmonary embolism. In some embodiments, the stroke is a haemorrhagic stroke. In some embodiments, the stroke is an ischaemic stroke. Cardiovascular adverse effects may be associated with markers indicating endothelial activation or thrombotic cascade. In some embodiments, administration of the agent may reduce the expression of Factor VIII, D-Dimer and / or von Willebrand factor on the endothelium, or reduce the increase in expression associated with coronavirus infection. Preventing coronavirus infection may prevent or reduce an increase in the expression of the expression of Factor VIII, D-Dimer and / or von Willebrand factor associated with coronavirus infection. Treating coronavirus may stabilise or reduce the expression of Factor VIII, D-Dimer and / or von Willebrand factor. In some embodiments, the adverse effects associated with coronavirus infection may be immunological adverse effects. Immunological adverse effects refer to anomalies or pathological effects within or caused by the immune system, which are caused by coronavirus infection. Typically, immunological adverse effects relate to an excessive immune response to coronavirus infection. In some embodiments, the immunological adverse effect is cytokine storm, sepsis or septic shock, or autoimmune sequelae to coronavirus infection, i.e. conditions in which the immune system attacks self-tissue following coronavirus infection. Cytokine storm refers to a systemic inflammatory syndrome associated with elevated levels of circulating cytokines and immune-cell hyperactivation that can be triggered by various therapies, pathogens, cancers, autoimmune conditions, and monogenic disorders. In some embodiments, administering the agent reduces the concentration of IL-ip, TNF-a and / or IL-6 which may be measured in the blood of a patient to whom the agent is administered. The method of preventing coronavirus infection may prevent or reduce an increase in the concentration of IL-ip, TNF-a and / or IL-6 in the blood of a patient to whom the agent is administered. The method of treating coronavirus infection may reduce the concentration of IL-ip, TNF-a and / or IL-6 in the blood of a patient to whom the agent is administered.

[0075] In some embodiments, the adverse effects associated with coronavirus infection may be respiratory adverse effects. Respiratory adverse effects refer to pathological effects caused by coronavirus infection of the respiratory system, and / or occurring within the respiratory system, namely the lungs, nasal passages and airways. In some embodiments, the respiratory adverse effects may be pneumonia, inflammation or fibrosis of the lungs, and / or acute respiratory stress. In some embodiments, the method of the invention may reduce the additional oxygen which a patient to whom the agent is administered may require due to reduced respiratory function, or prevent the patient from needing oxygen support. In some embodiments, the method may increase, or prevent a decrease, in the blood oxygen saturation which may be measured in a patient to whom the agent is administered.

[0076] The method may further comprise administering an agent which binds angiotensin converting enzyme 2 (ACE2), and / or transmembrane serine protease 2 (TMPRSS2). The agent which binds ACE2 preferably binds ACE2 selectively or specifically. In some embodiments, the method further comprises administering an agent which antagonises, or is a partial agonist of, the ACE2 receptor. In some embodiments, the method further comprises administering an agent which reduces the expression of ACE2. In particular embodiments, the agent may selectively or specifically reduce the expression of ACE2. The agent may selectively or specifically reduce the expression of ACE2 on the surface of cells, for example, by promoting intracellular retention of the receptor. In some embodiment, the method further comprises administering an agent which inhibits TMPRSS2 or reduces its expression. The agent preferably inhibits TMPRSS2, or reduces its expression, selectively or specifically.

[0077] As used herein, “coronavirus” is intended to refer to viruses of the family coronaviridae, which are particularly characterised by the presence of a spike protein. In some embodiments, the coronavirus is Middle East Respiratory Syndrome (MERS) coronavirus (MERS-CoV), Human coronavirus NL63 (HCoV-NL63), Human coronavirus 229E (HCoV-229E), Human coronavirus OC43 (HCoV-OC43) Severe Acute Respiratory Syndrome (SARS) coronavirus 1 or 2 (SARS-CoV-1 or SARS-CoV-2). In preferred embodiments, the coronavirus is SARS-CoV-2.

[0078] In some embodiments, the subject has come into contact with a coronavirus, such as SARS-CoV-2, or is suspected to have come into contact with a coronavirus, such as SARS-CoV-2. In some embodiments, the subject has been identified as being infected with coronavirus, preferably prior to commencing treatment. In some embodiments, the subject is identified as being infected with coronavirus by detection of coronavirus nucleic acids by polymerase chain reaction (PCR)-based techniques, particularly reverse transcription polymerase chain reaction (RT-PCR), quantitative PCR (qPCR), or most preferably, reverse transcription quantitative PCR (RT-qPCR). Alternatively, the subject may have been identified as being infected with SARS-CoV-2 by detection of coronavirus proteins such as the spike protein, such as by a lateral flow test. Accordingly, any of the methods of treating coronavirus infection described herein may further comprise a step of determining that the subject is infected with coronavirus. In some embodiments, the step of determining may comprise detecting the presence of the coronavirus or fragments thereof in a sample obtained from the subject. The step of determining that the subject is infected with coronavirus may comprise detection of coronavirus nucleic acids by PCR, such as RT-PCR, qPCR, or RT-qPCR. The step of determining that the subject is infected with coronavirus may alternatively or additionally comprise detection of coronavirus proteins such as the spike protein, such as by ELISA or a lateral flow test.

[0079] In some instances, the subject may be someone who is likely to come into contact with a coronavirus, in particular SARS-CoV-2. For example, the agent of the invention may be administered to health care workers.

[0080] In some embodiments, the SARS-CoV-2 is variant selected from: alpha, beta, gamma, delta, epsilon, eta, iota, kappa, zeta, mu and omicron. In some embodiments, the subject has been diagnosed as being infected with SARS-CoV-2 and the variant of SARS- CoV-2 has been identified.

[0081] Antibodies

[0082] The invention also provides an antibody capable of binding CD91. The antibody typically competes with the coronavirus (e.g. SARS-CoV-2) spike protein for binding to CD91, or binds to the same site (epitope) as the coronavirus (e.g. SARS-CoV-2) spike protein on CD91. In some instances, the antibody may not bind to exactly the same site as the coronavirus spike protein but may bind to an overlapping region (i.e. interacts with one or more of the same amino acids as the spike protein, but does not interact with a completely identical set of amino acids). As will be appreciated by the skilled person, an antibody that competes for binding may not necessarily bind to the same location on CD91, but may sterically block binding of the coronavirus spike protein to CD91.

[0083] To screen for antibodies that bind to a particular region, a routine cross-blocking assay can be performed. Other methods include alanine scanning mutants, peptide blots, or peptide cleavage analysis. In addition, methods such as epitope excision, epitope extraction and chemical modification of antigens can be employed. Such methods are well known in the art.

[0084] Binding may also be determined by x-ray crystallography analysis.

[0085] The antibody prevents or inhibits binding of the spike protein to CD91 as described above.

[0086] In preferred embodiments, the antibody’s epitope comprises one or more of the following amino acids of SEQ ID NO: 1 - 1060, 1061, 1065-1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090-1092, 1095-1099. In particular embodiments, the antibody’s epitope comprises at least two, at least three, at least four, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19 or all 20 of these residues. All combinations are contemplated.

[0087] In preferred embodiments, the antibody’s epitope comprises at least Cl 097 of SEQ ID NO: 1. In some instances, the epitope comprises C1097 of SEQ ID NO: 1 and at least 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 of the residues identified above.

[0088] The antibody’s epitope is typically determined by x-ray crystallography analysis. Antibodies of the present invention may therefore be assessed through x-ray crystallography analysis of the antibody bound to CD91. Epitopes may, in particular, be identified in this way by determining residues on CD91 within 5 A, preferably 4A, of an antibody paratope residue. As above, other methods include NMR, alanine scanning mutants, peptide blots, or peptide cleavage analysis. In addition, methods such as epitope excision, epitope extraction and chemical modification of antigens can be employed.

[0089] The antibody may possess any of the properties of the agent for use in the methods of the invention described herein.

[0090] Antibodies of the invention may be isolated antibodies. A composition consisting of an isolated antibody is substantially free of other antibodies having different antigenic specificities.

[0091] An antibody refers to a glycoprotein comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds, or an antigen-binding portion thereof. Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as HCVR or VH) and a heavy chain constant region. Each light chain is comprised of a light chain variable region (abbreviated herein as LCVR or VL) and a light chain constant region. The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen. The VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). In preferred embodiments, the antibody of the invention may be an antigen-binding fragment. As used herein, the terms “antigen-binding fragment” of an antibody, “antigenbinding portion” of an antibody, and the like, include any naturally occurring, enzymatically obtainable, synthetic, or genetically engineered polypeptide or glycoprotein that specifically binds an antigen to form a complex. Antigen-binding fragments of an antibody may be derived, e.g., from full antibody molecules using any suitable standard techniques such as proteolytic digestion or recombinant genetic engineering techniques involving the manipulation and expression of DNA encoding antibody variable and optionally constant domains. Such DNA is known and / or is readily available from, e.g., commercial sources, DNA libraries (including, e.g., phage-antibody libraries), or can be synthesized. The DNA may be sequenced and manipulated chemically or by using molecular biology techniques, for example, to arrange one or more variable and / or constant domains into a suitable configuration, or to introduce codons, create cysteine residues, modify, add or delete amino acids, etc. An antigen-binding fragment of the invention binds to the same epitope of the parent antibody, i.e. the antibody from which the antigenbinding fragment is derived. An antigen-binding fragment of the invention typically retains the parts of the parent antibody that interact with the epitope. The antigen-binding fragment typically comprise the complementarity-determining regions (CDRs) that interact with the antigen, such as one, two, three, four, five or six CDRs. The antigen-binding fragment may further comprise the structural scaffold surrounding the CDRs of the parent antibody, such as the variable region domains of the heavy and / or light chains. Typically, the antigen-binding fragment retains the same or similar binding affinity to the antigen as the parent antibody.

[0092] Non-limiting examples of antigen-binding fragments include: (i) Fab fragments; (ii) F(ab')2 fragments; (iii) Fd fragments; (iv) Fv fragments; (v) single-chain Fv (scFv) molecules; (vi) dAb fragments; and (vii) minimal recognition units consisting of the amino acid residues that mimic the hypervariable region of an antibody (e.g., an isolated complementarity determining region (CDR) such as a CDR3 peptide), or a constrained FR3-CDR3-FR4 peptide. Other engineered molecules, such as domain-specific antibodies, single domain antibodies, domain-deleted antibodies, chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies, tetrabodies, minibodies, nanobodies (e.g., monovalent nanobodies, bivalent nanobodies, etc.), small modular immunopharmaceuticals (SMIPs), and shark variable IgNAR domains, are also encompassed within the expression “antigenbinding fragment,” as used herein.

[0093] An antigen-binding fragment of an antibody will typically comprise at least one variable domain. The variable domain may be of any size or amino acid composition and will generally comprise at least one CDR adjacent to or in frame with one or more framework sequences. In some embodiments, an antigen-binding fragment of an antibody may contain at least one variable domain covalently linked to at least one constant domain. In any configuration of variable and constant domains, including any of the exemplary configurations listed above, the variable and constant domains may be either directly linked to one another or may be linked by a full or partial hinge or linker region. A hinge region may consist of at least 2 (e.g., 5, 10, 15, 20, 40, 60 or more) amino acids which result in a flexible or semi-flexible linkage between adjacent variable and / or constant domains in a single polypeptide molecule.

[0094] Moreover, an antigen-binding fragment of an antibody of the present disclosure may comprise a homo-dimer or hetero-dimer (or other multimer) of any of the variable and constant domain configurations listed above in non-covalent association with one another and / or with one or more monomeric VH or VL domain (e.g., by disulfide bond(s)).

[0095] An antibody of the invention may be a chimeric antibody, a CDR-grafted antibody, a nanobody, a human or humanised antibody. Preferably, the antibody is a human antibody. Fully human antibodies are those antibodies in which the variable regions and the constant regions (where present) of both the heavy and the light chains are all of human origin (i.e. derived from human germline immunoglobulin sequences), or substantially identical to sequences of human origin, but not necessarily from the same antibody.

[0096] By "specifically binds" or "immunoreacts with", it is meant that the antibody reacts with one or more antigenic determinants of the desired antigen and does not react with other polypeptides. In preferred embodiments, the antibody specifically binds CD91. In such embodiments, the antibody does not form a stable complex with molecules which are not CD91, fragments thereof or molecules with substantial sequence similarity thereto. Methods for determining whether an antibody specifically binds to an antigen are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, and the like. For example, an antibody that “specifically binds” human CD91, as used in the context of the present disclosure, includes antibodies that bind human CD91 or a portion thereof with a KD of less than about 500 nM, less than about 300 nM, less than about 200 nM, less than about 100 nM, less than about 90 nM, less than about 80 nM, less than about 70 nM, less than about 60 nM, less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 5 nM, less than about 4 nM, less than about 3 nM, less than about 2 nM, less than about 1 nM or less than about 0.5 nM, as measured in a surface plasmon resonance assay. An isolated antibody that specifically binds human CD91 may, however, have cross-reactivity to other antigens, such as CD91 molecules from other (non-human) species.

[0097] An antibody of the invention may be a monoclonal antibody. Monoclonal antibodies (mAbs) of the invention may be produced by a variety of techniques, including conventional monoclonal antibody methodology, for example those disclosed in “Monoclonal Antibodies: a manual of techniques” (Zola H, 1987, CRC Press) and in “Monoclonal Hybridoma Antibodies: techniques and applications” (Hurrell JGR, 1982 CRC Press).

[0098] An antibody of the invention may be multispecific, such as bispecific. A bispecific antibody of the invention binds two different epitopes. The epitopes may be in the same protein (e.g. two epitopes in CD91, preferably two epitopes of CD91 which interact with the receptor binding domain of the SARS-CoV-2 spike protein) or different proteins. One or more (e.g. two) antibodies of the invention can be coupled to form a multispecific (e.g. bispecific) antibody. Methods to prepare multispecific, e.g. bispecific, antibodies are well known in the art.

[0099] The antibody of the invention may be a full-length antibody, such as an IgA, IgD, IgE, IgG (IgGl, IgG2, IgG3 or IgG4) or IgM constant region. The constant regions of the antibodies may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system. Nonetheless, the antibodies of the invention function primarily to prevent coronavirus spike protein from binding CD91, therefore the constant region is not necessary. The antibodies may preferably lack a constant region to prevent adverse effects resulting from an autoimmune response directed against healthy endothelial cells.

[0100] Polynucleotides and vectors

[0101] The invention also provides one or more isolated polynucleotides encoding the agent for use according to the invention. In particular, the invention provides one or more polynucleotides encoding an antibody of the invention.

[0102] In some embodiments, the polynucleotide comprises or consists of DNA. In other embodiments, the polynucleotide comprises or consists of RNA, such as mRNA. The polynucleotide sequence may be collectively present on more than one polynucleotide, but collectively together they are able to encode an antibody or protein of the invention. For example, the polynucleotides may encode the heavy and / or light chain variable regions(s) of an antibody of the invention, or multiple domains of a protein of the invention.

[0103] Typically, one polynucleotide would encode each of the heavy and light chains of an antibody of the invention. Hence, the invention provides a first polynucleotide encoding the heavy chain variable domain of an antibody of the invention and a second polynucleotide encoding the light chain variable domain of said antibody. The invention also provides a single polynucleotide encoding both the heavy chain variable domain and the light chain variable domain of an antibody according to the invention, optionally wherein the sections of the polynucleotide encoding the heavy and light chains are separated by a linker.

[0104] Polynucleotides which encode an antibody or protein for use according to the invention, or an antibody of the invention, can be obtained by methods well known to those skilled in the art. For example, DNA sequences coding for part or all of the antibody heavy and light chains may be synthesised as desired from the corresponding amino acid sequences.

[0105] A polynucleotide of the invention may be provided in the form of an expression cassette, which includes control sequences operably linked to the inserted sequence, thus allowing for expression of the antibody of the invention in vivo. Accordingly, in some embodiments, cassettes may include a promoter, an open reading frame with or without introns, and a termination signal. Additional factors necessary or helpful in effecting expression may also be used, such as, for example, an inducible promoter. Hence, the invention also provides one or more expression cassettes comprising or encoding one or more polynucleotides encoding an antibody of the invention or an agent for use according to the invention, wherein the agent is a protein, nucleic acid or antibody. These expression cassettes, in turn, are typically provided within vectors (e.g. plasmids or recombinant viral vectors). Hence, the invention provides a vector encoding an antibody of the invention, or an agent for use according to the invention, wherein the agent is a protein, nucleic acid or antibody. The invention also provides vectors which collectively encode an antibody of the invention, or an agent for use according to the invention, wherein the agent is a protein, nucleic acid or antibody. The vectors may be cloning vectors or expression vectors. A suitable vector may be any vector which is capable of carrying a sufficient amount of genetic information, and allowing expression of a polypeptide of the invention.

[0106] The agent for use according to the invention may be encoded by a single nucleic acid molecule or multiple nucleic acid molecules. Accordingly, the polynucleotide of the invention may comprise a single molecule, or multiple molecules. For example, where the agent for use is an antibody, each chain may be encoded by a separate nucleic acid molecule.

[0107] In some embodiments, the polynucleotides, expression cassettes or vectors of the invention may be introduced into a host cell, e.g. by transfection. Hence, the invention also provides a host cell comprising the one or more polynucleotides, expression cassettes or vectors of the invention. The polynucleotides, expression cassettes or vectors of the invention may be introduced transiently or permanently into the host cell, allowing expression of an antibody from the one or more polynucleotides, expression cassettes or vectors. Such host cells include transient, or preferably stable higher eukaryotic cell lines, such as mammalian cells or insect cells, lower eukaryotic cells, such as yeast, or prokaryotic cells, such as bacteria cells. Particular examples of cells include mammalian HEK293, such as HEK293F, HEK293T, HEK293S or HEK Expi293F, CHO, HeLa, NSO and COS cells, or any other cell line used herein, such as the ones used in the Examples. Preferably the cell line selected will be one which is not only stable, but also allows for mature glycosylation.

[0108] The invention also provides a process for the production of a protein or an antibody of the invention, comprising culturing a host cell containing one or more vectors of the invention under conditions suitable for the expression of the antibody from the one or more polynucleotides of the invention, and isolating the protein or antibody from said culture. The antibodies can also be generated using various phage display methods known in the art and include those disclosed by Brinkman et al. (in J. Immunol. Methods, 1995, 182: 41- 50), Ames et al. (J. Immunol. Methods, 1995, 184: 177-186), Kettleborough et al. (Eur. J. Immunol. 1994, 24:952-958), Persic et al. (Gene, 1997 187 9-18), Burton et al. (Advances in Immunology, 1994, 57: 191-280) and WO 90 / 02809; WO 91 / 10737; WO 92 / 01047; WO 92 / 18619; WO 93 / 11236; WO 95 / 15982; WO 95 / 20401; and US 5,698,426; 5,223,409; 5,403,484; 5,580,717; 5,427,908; 5,750,753; 5,821,047; 5,571,698; 5,427,908; 5,516,637; 5,780,225; 5,658,727; 5,733,743 and 5,969,108. Pharmaceutical Compositions

[0109] The invention provides a pharmaceutical composition comprising the antibody of the invention or the agent for use according to the invention and a pharmaceutically acceptable carrier, salt and / or diluent. In some embodiments, the composition comprises a combination of agents for use according to the invention. The invention also provides a pharmaceutical composition comprising a polynucleotide or vector of the invention, and a pharmaceutically acceptable carrier or diluent.

[0110] In some embodiments, the pharmaceutical composition of the invention may comprise one or more pharmaceutically acceptable salts. A "pharmaceutically acceptable salt" refers to a salt that retains the desired biological activity of the parent compound and does not impart any undesired toxicological effects. Examples of such salts include acid addition salts and base addition salts.

[0111] In some embodiments, the pharmaceutical composition of the invention may comprise one or more pharmaceutically acceptable carriers. Suitable pharmaceutically acceptable carriers comprise aqueous carriers or diluents. Examples of suitable aqueous carriers include water, buffered water and saline. Other suitable pharmaceutically acceptable carriers include ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, and the like), and suitable mixtures thereof, vegetable oils, such as olive oil, and injectable organic esters, such as ethyl oleate. In many cases, it will be desirable to include isotonic agents, for example, sugars, polyalcohols such as mannitol, sorbitol, or sodium chloride in the composition.

[0112] Pharmaceutical compositions typically must be sterile and stable under the conditions of manufacture and storage. The composition can be formulated as a solution, microemulsion, liposome, or other ordered structure suitable to high drug concentration. Pharmaceutical compositions of the invention may comprise additional therapeutic agents, for example an additional anti-viral agent. The anti-viral agent may bind to coronavirus and thereby inhibit viral activity. Alternatively, the anti-viral agent may not bind directly to coronavirus but still affect viral activity / infectivity. The anti-viral agent could be a further anti -coronavirus antibody, which binds an epitope on a coronavirus such as SARS- CoV-2, e.g. the spike protein. Examples of an anti-viral agents which may be used in the method of the present invention include Remdesivir, Lopinavir, ritonavir, APN01, Favilavir, Nirmatrelvir, and / or molnupiravir.

[0113] The additional therapeutic agent may be an anti-inflammatory agent, such as a corticosteroid (e.g. Dexamethasone) or a non-steroidal anti-inflammatory drug (e.g. Tocilizumab). The additional therapeutic agent may be a further anti-coronavirus therapeutic. In some embodiments, the composition further comprises an agent which binds the ACE2 receptor. In preferred embodiments, the agent selectively or specifically binds the ACE2 receptor. By selectively binding the ACE2 receptor, agent has a greater affinity for ACE2 than other molecules, such as ligands or receptors. Accordingly, said agent has a greater tendency to form stable complexes with ACE2 than other molecules. However, the agent may still bind or interact with molecules which are not ACE2. By specifically binding ACE2, the agent does not form stable complexes with molecules other than ACE2 or fragments thereof, or molecules with extensive sequence similarity thereto.

[0114] The composition may further comprise an agent which reduces the expression of ACE2. In preferred embodiments, the agent reduces the expression of ACE2 on the surface of cells. In preferred embodiments, the agent selectively reduces the expression of ACE2 relative to other genes. By selectively reducing expression relative to other genes, the agent reduces the expression of ACE2 to a greater extent than it may modulate the expression of other genes. By specifically reducing the expression of ACE2, the agent reduces the expression of ACE2 without significantly modulating the expression of other genes.

[0115] The composition may further comprise an agent which inhibits TMPRSS2. The agent which inhibits TMPRSS2 may be nafamostat, camostat, or gabexate for example. The agent may also be an anti-TMPRSS2 antibody. In a preferred embodiment, the composition may comprise an agent which is capable of selectively or specifically binding CD91 for use in accordance with the invention, an agent which selectively or specifically binds ACE2, and an agent which inhibits TMPRSS2. In especially preferred embodiments, the agent which is capable of selectively or specifically binding CD91 is an antibody.

[0116] The composition may comprise a mixture of two or more antibodies, which may be referred to herein as a cocktail. The two or more antibodies may all bind CD91 in accordance with the invention, or comprise at least one antibody which binds CD91 in accordance with the invention, and one or more antibodies which bind epitopes on other proteins.

[0117] The additional therapeutic agent may be an anti-coronavirus vaccine.

[0118] The pharmaceutical composition may be administered subcutaneously, intravenously, intradermally, intramuscularly, intracranially, intranasally or orally. Typically, the antibodies, nucleic acids, pharmaceutical compositions and antibody cocktails compositions are administered intravenously or subcutaneously.

[0119] Also within the scope of the invention are kits comprising antibodies or other compositions of the invention and instructions for use. The kit may further contain one or more additional reagents, such as an additional therapeutic or prophylactic agent as discussed herein.

[0120] The dose of an agent, or specifically an antibody may vary depending on the age and size of a subject, as well as on the disease, conditions and route of administration. Antibodies may be administered at a dose of about 0.1 mg / kg body weight to a dose of about 100 mg / kg body weight, such as at a dose of about 5 mg / kg to about 10 mg / kg. Antibodies may also be administered at a dose of about 50 mg / kg, 10 mg / kg or about 5 mg / kg body weight.

[0121] The initial dose may be followed by administration of a second or plurality of subsequent doses. The second and subsequent doses may be separated by an appropriate time.

[0122] Screening methods

[0123] The invention also provides a method of screening an agent use in for treating and / or preventing coronavirus infections, said method comprising testing the binding of a coronavirus spike protein to CD91 in the presence of the agent; and identifying the agent as having utility in treating and / or preventing coronavirus infection if the agent inhibits or prevents the interaction of the coronavirus spike protein with CD91. As described above, the agent completely prevent the coronavirus spike protein from binding to CD91. Alternatively, the agent may provide a certain degree of inhibition as described above. Such prevent! on / inhibiti on of binding can be determined using routine methods known in the art. As explained above, the coronavirus may be any coronavirus. However, the coronavirus is typically SARS-CoV-2.

[0124] Screening methods of the invention may also involve determining the binding site of the agent on CD91. In these embodiments, an agent may be identified as having potential therapeutic activity if it binds to and / or interacts with any one of amino acids 1060, 1061, 1065-1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090-1092, 1095-1099 of SEQ ID NO: 1, (or a corresponding residue of a corresponding sequence), or any combination thereof. In preferred instances the agent binds to residue 1097 of SEQ ID NO: 1 (or corresponding sequence). Corresponding sequences are explained above. In particular embodiments, the antibody may recognise an epitope spanning multiple domains of CD91.

[0125] In some instances, the method comprises determining the binding site of the agent and then confirming that the agent prevents or inhibits the coronavirus spike protein binding to CD91.

[0126] Examples

[0127] Materials and Methods:

[0128] Protein X-ray crystal structure selection:

[0129] Using the protein data bank (PDB) a protein crystal structure was selected for each of the proteins of interest in this study.

[0130] For the low-density lipoprotein receptor related protein (LRP1) the protein with the code 1CR8 was selected from the PDB. 1CR8 corresponds to LRP1 protein complement repeat 8, which was thought to be the principal binding site for protein ligands. The sequence of 1CR8 is shown in SEQ ID NO: 2. The process used in the study that identified this crystal structure was an NMR method and the protein had 20 conformers. For the Covid-spike protein 7L4Z was selected from the PDB (SEQ ID NO: 3 - SARS- CoV-2 spike protein RBD sequence from 7L4Z). The process used in the study which identified the crystal structure was X-ray diffraction, the protein had a resolution of 3.96 A.

[0131] Computational preparation of proteins: Separately, the proteins were loaded into MOE (Molecular Operating Environment 2022’ developed by Chemical computing group ULC) as .PDB files. Using the QuickPrep feature in MOE the proteins were prepared. Default QuickPrep settings were used, apart fixing atoms further than 8 angstroms from the ligand. This option was toggled off as this is protein-protein docking, so naturally there is no ‘ligand’, this made the docking process to follow quicker and less computationally expensive.

[0132] Both 1CR8 and 7L4Z were complete in their primary structures so QuickPrep was not needed to fill in an gaps in their primary structures.

[0133] Protein-Protein Docking:

[0134] Two runs of the protein- protein docking were run with the first running 1CR8 as the receptor and 7L4Z as the ligand. The second docking run was then done in the opposite fashion to see if there was any difference in the binding site and binding scoring function of the best pose in the two runs.

[0135] As there was minimal difference in the binding scoring function, the binding site was determined. This allowed for two of the three chains of 7L4Z to be disregarded in order to make further docking quicker and less computationally expensive. The portion left of 7L4Z was then docked against 1CR8 again to establish whether the binding scores were changed by the deletion of the other subchains.

[0136] Protein mutagenesis:

[0137] Once the binding site had been established, the alanine scan feature in MOE was used to determine if the protein binding was specific to this confirmation of the protein. Being the smaller protein, of only 42 residues, 1CR8 was chosen to be mutated. Using the alanine scan, each of the 42 residues was mutated to an alanine residue and docked to 7L4Z as the ‘new’ protein.

[0138] Binding scoring functions were calculated in MOE for each of the 42 mutated proteins and compared against the original protein binding scoring function as both their best and average scoring function to see the specificity of the binding of the two proteins.

[0139] Protein stability: Using the residue scan function on MOE, the stability change for each residue was calculated by mutating each residue of 1CR8, again chosen due to the smaller nature of the protein compared to that of the covid-spike protein.

[0140] Each of the 42 residues were mutated by the software to each of the 20 amino acids and a change in stability value was calculated for each mutated protein.

[0141] Cell culture, transfection and virus:

[0142] HUVECs from pooled donors (Lonza) were cultured on 0.5 % gelatin (diluted from 2 % stock; in EGM2 media with supplements. HEK293T (human, kidney) were incubated in in DMEM supplemented with 10% FBS, 1% pen-strep and 100 pM L-glutamine. All cells were cultured in sterilised conditions at 37 °C with 5% of CO2. The infection experiments with SARS-CoV-2, were done under conditions of biosafety level 3. For siRNA transfection, HUVECs were cultured for up to six passages and transfected with Lipofectamine RNAiMAX. Media was replaced 24 hours after transfection. The following siRNAs were used to transfect cells: SMARTPool SiRNA targeting LRP1 (Dharmacon).

[0143] Plasmids:

[0144] HEK293T cells were transfected by using Fugene reagent following the manufacturer’s protocol with the plasmid DNA for ACE2, TMPRSS2 and LRP1. Pseudotyping:

[0145] SARS-CoV-2 pseudotyped viral particles (PV) contain Lentiviral vector incorporating luciferase reporter, packaging construct and SARS-CoV-2 spike expression plasmid. Cells were inoculated with SARS-CoV-2 PV and the transduction efficiency was quantified 24 hours posttransduction by measuring the activity of luciferase (RLU readings) in cell lysates using Bright Gio luciferase assay system and a luminometer.

[0146] Example 1 - Protein-protein docking of SARS-CoV-2 spike protein to CD91,

[0147] The protein sequences were retrieved from the protein databank (PDB). These crystal structures had the codes of 1CR8, 7L4Z, and 7LM9. These crystal structures corresponded to LRP1 (containing clusters of cysteine-rich complement-like repeats which are thought to be the principal binding sites for protein ligands), the covid spike protein (containing the spike protein receptor binding domain (RBD)) and the covid spike protein receptor-binding domain, respectively. Molecular operating environment (MOE) was used to prepare the crystal structures using the QuickPrep function. Once prepared, the first protein-protein dock was carried out using MOE between 7L4Z (SEQ ID NO: 3) and 7LM9 to establish a baseline docking score of the spike protein and the native receptor, the best docking score coming out at -99.62. The protein-protein dock was then carried out between 1CR8 (LRP1) and 7L4Z (Spike-RBD - SEQ ID NO: 3) to identify interactions between the spike protein and LRP1, achieving the highest docking score of -63.66 (Figure

[0148] 1 A, ID and IE).

[0149] The model showed that the residue numbered C41 (C39 of SEQ ID NO: 2) is located in the binding pocket and has been identified to be essential for interaction with Covid-spike. 1CR8 (LRP1) configuration forms several hydrogen bonds (H-bond) with 7L4Z (Covid-spike) (Figure IB).

[0150] In silico mutagenesis of single amino acids of 1CR8 (LRP1) in alanine residue was evaluated by scoring functions calculated in MOE for each of the 42 mutated proteins and compared against the original protein binding scoring function as both their best and average scoring function to see the specificity of the binding of the two proteins. Results are presented in Figure 1C (the residue numbering in Figure 1C differs from SEQ ID NO:

[0151] 2 by two amino acids, i.e. G4 of Figure 1C corresponds to G2 of SEQ ID NO 2, E42 of Figure 1C corresponds to E40 of SEQ ID NO: 2 etc). Thus, this is assessing the specificity of the binding with 7L4Z (Covid-spike) which is associated with the change in binding energy (AG) from wild type (WT) to the mutated LRP1 protein.

[0152] The determination of the stability of each of the 42 residues mutated was calculated for each mutated protein in Table 1, including the amino acids present in the binding pocket.

[0153] Table 1 :

[0154] Example 2 - CD91 facilitates the cellular entry of SARS-CoV-2 pseudotyped particles.

[0155] To further determine whether SARS-CoV-2 can use LRP1 for virus entry, lentiviral particles pseudotyped with the SARS-CoV-2 S protein were used in Hek-293-T. Pseudoviruses have been well used to perform virus entry assays, as they allow viral entry analysis.

[0156] HEK-293T cells were transfected with plasmids that encode the human ACE2, the transmembrane protease serine 2 (TMPRSS2), or LRP1. Results are presented in Figure 2. In cells overexpressing ACE2 and TMPRSS2 or LRP1 and TMPRSS2, the infection by SARS-CoV-2 pseudoviruses rendered cells susceptible to infection. Interestingly, transfection of ACE2, TMPRSS2 and LRP1 markedly enhanced infection.

[0157] Example 3 - LRP1 (CD91) functions contribute to replication of SARS-CoV-2 in endothelial cells

[0158] To characterize the impact of LRP1 deficiency in the replication of the virus, primary endothelial cells (ECs) such as human umbilical vein EC (HUVEC), were infected with the SARS-CoV-2 virus. Thus, LRP1, small interfering RNA (siRNA) knockdown was carried in ECs. Results are presented in Figure 3.

[0159] In these cells, data showed that LRP1 was affecting SARS-CoV-2 multiplication. In summary, the data discussed herein suggest that LRP1 (CD91) is an entry receptor which may be used by SARS-CoV-2 in endothelial cells.

[0160] References:

[0161] 1. C. C. Lai, T. P. Shih, W. C. Ko, H. J. Tang, P. R. Hsueh, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and coronavirus disease-2019 (COVID-19): The epidemic and the challenges. Int J Antimicrob Agents 55, 105924 (2020).

[0162] 2. X. Hu et al., The study of antiviral drugs targeting SARS-CoV-2 nucleocapsid and spike proteins through large-scale compound repurposing. Heliyon 7, e06387 (2021).

[0163] 3. C. Huang et al., Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Lancet 395, 497-506 (2020).

[0164] 4. A. Gupta et al., Extrapulmonary manifestations of COVID-19. Nat Med 26, 1017- 1032 (2020).

[0165] 5. M. Panigada et al., Hypercoagulability of COVID-19 patients in Intensive Care Unit. A Report of Thromboelastography Findings and other Parameters of Hemostasis. J Thromb Haemost, (2020).

[0166] 6. S. X. Gu et al., Thrombocytopathy and endotheliopathy: crucial contributors to COVID-19 thromboinflammation. Nat Rev Cardiol 18, 194-209 (2021).

[0167] 7. M. Hoffmann et al., SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 and Is Blocked by a Clinically Proven Protease Inhibitor. Cell 181, 271-280 e278 (2020).

[0168] 8. I. R. McCracken et al., Lack of Evidence of Angiotensin-Converting Enzyme 2 Expression and Replicative Infection by SARS-CoV-2 in Human Endothelial Cells. Circulation 143, 865-868 (2021).

[0169] 9. D. M. Smadja et al., COVID-19 is a systemic vascular hemopathy: insight for mechanistic and clinical aspects. Angiogenesis 24, 755-788 (2021).

[0170] 10. C. Grobler et al., Covid-19: The Rollercoaster of Fibrin(Ogen), D-Dimer, Von Willebrand Factor, P-Selectin and Their Interactions with Endothelial Cells, Platelets and Erythrocytes. Int J Mol Sci 21, (2020).

[0171] 11. E. E. Ladikou et al., Von Willebrand factor (vWF): marker of endothelial damage and thrombotic risk in COVID-19? Clin Med (Lond) 20, el78-el82 (2020).

[0172] 12. B. Horvath et al., Measurement of von Willebrand factor as the marker of endothelial dysfunction in vascular diseases. Exp Clin Cardiol 9, 31-34 (2004).

[0173] 13. G. Rastegarlari et al., Macrophage LRP1 contributes to the clearance of von Willebrand factor. Blood 119, 2126-2134 (2012). 14. H. Chun et al., Characterization of interaction between blood coagulation factor VIII and LRP1 suggests dynamic binding by alternating complex contacts. J Thromb Haemost 20, 2255-2269 (2022).

[0174] 15. A. P. Lillis, L. B. Van Duyn, J. E. Murphy-Ullrich, D. K. Strickland, LDL receptor- related protein 1 : unique tissue-specific functions revealed by selective gene knockout studies. Physiol Rev 88, 887-918 (2008).

[0175] 16. S. E. Williams, J. D. Ashcom, W. S. Argraves, D. K. Strickland, A novel mechanism for controlling the activity of alpha 2-macroglobulin receptor / low density lipoprotein receptor-related protein. Multiple regulatory sites for 39-kDa receptor- associated protein. J Biol Chem 267, 9035-9040 (1992).

[0176] 17. D. K. Strickland, D. T. Au, P. Cunfer, S. C. Muratoglu, Low-density lipoprotein receptor-related protein- 1 : role in the regulation of vascular integrity. Arterioscler Thromb Vase Biol 34, 487-498 (2014).

[0177] 18. J. Chen, Y. Su, S. Pi, B. Hu, L. Mao, The Dual Role of Low-Density Lipoprotein Receptor-Related Protein 1 in Atherosclerosis. Front Cardiovasc Med 8, 682389 (2021).

[0178] 19. H. Mao, P. Lockyer, W. H. Townley-Tilson, L. Xie, X. Pi, LRP1 Regulates Retinal Angiogenesis by Inhibiting PARP-1 Activity and Endothelial Cell Proliferation. Arterioscler Thromb Vase Biol 36, 350-360 (2016).

[0179] 20. M. Meilinger et al., Metabolism of activated complement component C3 is mediated by the low density lipoprotein receptor-related protein / alpha(2)-macroglobulin receptor. J Biol Chem 274, 38091-38096 (1999).

[0180] 21. F. Wrensch et al., Hepatitis C Virus (HCV)-Apolipoprotein Interactions and Immune Evasion and Their Impact on HCV Vaccine Design. Front Immunol 9, 1436 (2018).

[0181] 22. S. S. Ganaie et al., Lrpl is a host entry factor for Rift Valley fever virus. Cell 184, 5163-5178 e5124 (2021).

[0182] 23. F. J. Redondo-Calvo et al., Aprotinin treatment against SARS-CoV-2: A randomized phase III study to evaluate the safety and efficacy of a pan-protease inhibitor for moderate COVID-19. Eur J Clin Invest 52, el3776 (2022).

[0183] 24. E. Bolat et al., Lactoferrin for COVID-19 prevention, treatment, and recovery. Front Nutr 9, 992733 (2022). Sequence listing

[0184] SEQ ID NO: 1 Human LRP1 (Uniprot ID Q07954)

[0185] MLTPPLLLLLPLLSALVAAAIDAPKTCSPKQFACRDQITCISKGWRCDGERDCPDG SDEAPEICPQSKAQRCQPNEHNCLGTELCVPMSRLCNGVQDCMDGSDEGPHCREL QGNCSRLGCQHHCVPTLDGPTCYCNSSFQLQADGKTCKDFDECSVYGTCSQLCTN TDGSFICGCVEGYLLQPDNRSCKAKNEPVDRPPVLLIANSQNILATYLSGAQVSTIT PTSTRQTTAMDFSYANETVCWVHVGDSAAQTQLKCARMPGLKGFVDEHTINISLS LHHVEQMAIDWLTGNFYFVDDIDDRIFVCNRNGDTCVTLLDLELYNPKGIALDPA MGKVFFTDYGQIPKVERCDMDGQNRTKLVDSKIVFPHGITLDLVSRLVYWADAY LDYIEVVDYEGKGRQTIIQGILIEHLYGLTVFENYLYATNSDNANAQQKTSVIRVN RFNSTEYQVVTRVDKGGALHIYHQRRQPRVRSHACENDQYGKPGGCSDICLLANS HKARTCRCRSGFSLGSDGKSCKKPEHELFLVYGKGRPGIIRGMDMGAKVPDEHMI PIENLMNPRALDFHAETGFIYFADTTSYLIGRQKIDGTERETILKDGIHNVEGVAVD WMGDNLYWTDDGPKKTISVARLEKAAQTRKTLIEGKMTHPRAIVVDPLNGWMY WTDWEEDPKDSRRGRLERAWMDGSHRDIFVTSKTVLWPNGLSLDIPAGRLYWVD AFYDRIETILLNGTDRKIVYEGPELNHAFGLCHHGNYLFWTEYRSGSVYRLERGVG GAPPT VTLLRSERPPIFEIRMYD AQQQQ VGTNKCRVNNGGC S SLCL ATPGSRQC AC AEDQVLDADGVTCLANPSYVPPPQCQPGEFACANSRCIQERWKCDGDNDCLDNS DEAPALCHQHTCPSDRFKCENNRCIPNRWLCDGDNDCGNSEDESNATCSARTCPP NQFSCASGRCIPISWTCDLDDDCGDRSDESASCAYPTCFPLTQFTCNNGRCININW RCDNDNDCGDNSDEAGCSHSCSSTQFKCNSGRCIPEHWTCDGDNDCGDYSDETH ANCTNQATRPPGGCHTDEFQCRLDGLCIPLRWRCDGDTDCMDSSDEKSCEGVTH VCDPSVKFGCKDSARCISKAWVCDGDNDCEDNSDEENCESLACRPPSHPCANNTS VCLPPDKLCDGNDDCGDGSDEGELCDQCSLNNGGCSHNCSVAPGEGIVCSCPLG MELGPDNHTCQIQSYCAKHLKCSQKCDQNKFSVKCSCYEGWVLEPDGESCRSLD PFKPFIIFSNRHEIRRIDLHKGDYSVLVPGLRNTIALDFHLSQSALYWTDVVEDKIYR GKLLDNGALTSFEVVIQYGLATPEGLAVDWIAGNIYWVESNLDQIEVAKLDGTLR TTLLAGDIEHPRAIALDPRDGILFWTDWDASLPRIEAASMSGAGRRTVHRETGSGG

[0186] WPNGLTVDYLEKRILWIDARSDAIYSARYDGSGHMEVLRGHEFLSHPFAVTLYGG EVYWTDWRTNTLAKANKWTGHNVTVVQRTNTQPFDLQVYHPSRQPMAPNPCEA NGGQGPCSHLCLINYNRTVSCACPHLMKLHKDNTTCYEFKKFLLYARQMEIRGV DLDAPYYNYIISFTVPDIDNVTVLDYDAREQRVYWSDVRTQAIKRAFINGTGVETV VSADLPNAHGLAVDWVSRNLFWTSYDTNKKQINVARLDGSFKNAVVQGLEQPH GLVVHPLRGKLYWTDGDNISMANMDGSNRTLLFSGQKGPVGLAIDFPESKLYWIS SGNHTINRCNLDGSGLEVIDAMRSQLGKATALAIMGDKLWWADQVSEKMGTCSK ADGSGSVVLRNSTTLVMHMKVYDESIQLDHKGTNPCSVNNGDCSQLCLPTSETTR SCMCTAGYSLRSGQQACEGVGSFLLYSVHEGIRGIPLDPNDKSDALVPVSGTSLAV GIDFHAENDTIYWVDMGLSTISRAKRDQTWREDVVTNGIGRVEGIAVDWIAGNIY WTDQGFDVIEVARLNGSFRYVVISQGLDKPRAITVHPEKGYLFWTEWGQYPRIER SRLDGTERVVLVNVSISWPNGISVDYQDGKLYWCDARTDKIERIDLETGENREVV LSSNNMDMFSVSVFEDFIYWSDRTHANGSIKRGSKDNATDSVPLRTGIGVQLKDIK VFNRDRQKGTNVCAVANGGCQQLCLYRGRGQRACACAHGMLAEDGASCREYA GYLLYSERTILKSIHLSDERNLNAPVQPFEDPEHMKNVIALAFDYRAGTSPGTPNRI FFSDIHFGNIQQINDDGSRRITIVENVGSVEGLAYHRGWDTLYWTSYTTSTITRHTV DQTRPGAFERETVITMSGDDHPRAFVLDECQNLMFWTNWNEQHPSIMRAALSGA NVLTLIEKDIRTPNGLAIDHRAEKLYFSDATLDKIERCEYDGSHRYVILKSEPVHPF GLAVYGEHIFWTDWVRRAVQRANKHVGSNMKLLRVDIPQQPMGIIAVANDTNSC ELSPCRINNGGCQDLCLLTHQGHVNCSCRGGRILQDDLTCRAVNSSCRAQDEFEC ANGECINFSLTCDGVPHCKDKSDEKPSYCNSRRCKKTFRQCSNGRCVSNMLWCN GADDCGDGSDEIPCNKTACGVGEFRCRDGTCIGNSSRCNQFVDCEDASDEMNCSA TDCSSYFRLGVKGVLFQPCERTSLCYAPSWVCDGANDCGDYSDERDCPGVKRPR CPLNYFACPSGRCIPMSWTCDKEDDCEHGEDETHCNKFCSEAQFECQNHRCISKQ WLCDGSDDCGDGSDEAAHCEGKTCGPSSFSCPGTHVCVPERWLCDGDKDCADG ADESIAAGCLYNSTCDDREFMCQNRQCIPKHFVCDHDRDCADGSDESPECEYPTC GPSEFRCANGRCLSSRQWECDGENDCHDQSDEAPKNPHCTSQEHKCNASSQFLCS SGRCVAEALLCNGQDDCGDSSDERGCHINECLSRKLSGCSQDCEDLKIGFKCRCRP GFRLKDDGRTCADVDECSTTFPCSQRCINTHGSYKCLCVEGYAPRGGDPHSCKAV TDEEPFLIFANRYYLRKLNLDGSNYTLLKQGLNNAVALDFDYREQMIYWTDVTTQ GSMIRRMHLNGSNVQVLHRTGLSNPDGLAVDWVGGNLYWCDKGRDTIEVSKLN GAYRTVLVS SGLREPRALVVD VQNGYLYWTDWGDHSLIGRIGMDGS SRS VIVDT

[0187] KITWPNGLTLDYVTERIYWADAREDYIEFASLDGSNRHVVLSQDIPHIFALTLFEDY VYWTDWETKSINRAHKTTGTNKTLLISTLHRPMDLHVFHALRQPDVPNHPCKVN NGGCSNLCLLSPGGGHKCACPTNFYLGSDGRTCVSNCTASQFVCKNDKCIPFWW KCDTEDDCGDHSDEPPDCPEFKCRPGQFQCSTGICTNPAFICDGDNDCQDNSDEAN CDIHVCLPSQFKCTNTNRCIPGIFRCNGQDNCGDGEDERDCPEVTCAPNQFQCSITK RCIPRVWVCDRDNDCVDGSDEPANCTQMTCGVDEFRCKDSGRCIPARWKCDGED DCGDGSDEPKEECDERTCEPYQFRCKNNRCVPGRWQCDYDNDCGDNSDEESCTP RPCSESEFSCANGRCIAGRWKCDGDHDCADGSDEKDCTPRCDMDQFQCKSGHCIP LRWRCDADADCMDGSDEEACGTGVRTCPLDEFQCNNTLCKPLAWKCDGEDDCG DNSDENPEECARFVCPPNRPFRCKNDRVCLWIGRQCDGTDNCGDGTDEEDCEPPT AHTTHCKDKKEFLCRNQRCLS S SLRCNMFDDCGDGSDEEDC SIDPKLT SC ATNASI CGDEARCVRTEKAAYCACRSGFHTVPGQPGCQDINECLRFGTCSQLCNNTKGGHL CSCARNFMKTHNTCKAEGSEYQVLYIADDNEIRSLFPGHPHSAYEQAFQGDESVRI DAMDVHVKAGRVYWTNWHTGTISYRSLPPAAPPTTSNRHRRQIDRGVTHLNISGL I<MPRGIAIDWVAGNVYWTDSGRDVIEVAQMI<GENRI<TLISGMIDEPHAIVVDPLR GTMYWSDWGNHPKIETAAMDGTLRETLVQDNIQWPTGLAVDYHNERLYWADA KLSVIGSIRLNGTDPIVAADSKRGLSHPFSIDVFEDYIYGVTYINNRVFKIHKFGHSP LVNLTGGLSHASDVVLYHQHKQPEVTNPCDRKKCEWLCLLSPSGPVCTCPNGKR LDNGTCVPVPSPTPPPDAPRPGTCNLQCFNGGSCFLNARRQPKCRCQPRYTGDKCE LDQCWEHCRNGGTCAASPSGMPTCRCPTGFTGPKCTQQVCAGYCANNSTCTVNQ

[0188] GNQPQCRCLPGFLGDRCQYRQCSGYCENFGTCQMAADGSRQCRCTAYFEGSRCE VNKCSRCLEGACVVNKQSGDVTCNCTDGRVAPSCLTCVGHCSNGGSCTMNSKM MPECQCPPHMTGPRCEEHVFSQQQPGHIASILIPLLLLLLLVLVAGVVFWYKRRVQ GAKGFQHQRMTNGAMNVEIGNPTYKMYEGGEPDDVGGLLDADFALDPDKPTNF TNPVYATLYMGGHGSRHSLASTDEKRELLGRGPEDEIGDPLA

[0189] SEQ ID NO: 2 - PDB entry 1CR8 (low density lipoprotein receptor-related protein complement repeat 8)

[0190] PGGCHTDEFQCRLDGLCIPLRWRCDGDTDCMDSSDEKSCEGV

[0191] SEQ ID NO:3 - SARS-CoV-2 spike RBD sequence from PDB entry 7L4Z

[0192] RVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFS TFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTG CVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFN CYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFH HHHHH

[0193] SEQ ID NO: 4 - SARS-CoV-2 spike protein sequence (UniProt ID P0DTC2)

[0194] MFVFLVLLPLVS SQC VNLTTRTQLPP AYTNSFTRGVYYPDKVFRS S VLHSTQDLFL PFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSK TQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTF EYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALE PLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYN ENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPF GEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFT NVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGN YNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVG YQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKF LPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVN CTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICA SYQTQTNSPRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVS MTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQV KQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGD IAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFA MQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQ NAQALNTLVKQLSSNFGAISSVLNDILSRLDKVEAEVQIDRLITGRLQSLQTYVTQQ LIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVT YVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFV SGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNI QKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLC CMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT

Claims

CLAIMS1. A method of treating and / or preventing coronavirus infection, the method comprising administering a therapeutically effective amount of an agent to a subject, wherein the agent binds to CD91 and wherein the agent is not lactoferrin or aprotinin.

2. The method of claim 1, wherein the coronavirus is SARS-CoV-2.

3. The method of claim 1 or 2, wherein the agent the agent does not modulate signalling through the CD91 receptor.

4. The method of any one of the preceding claims, wherein the agent inhibits or prevents the receptor binding domain of the coronavirus spike protein from binding to CD91.

5. The method of any one of the preceding claims, wherein the agent competitively inhibits binding of the coronavirus spike protein receptor binding domain to CD91.

6. The method of any one of the preceding claims, wherein the agent is a protein ligand, a small molecule or a nucleic acid aptamer.

7. The method of any one of claims 1-5, wherein the agent is an antibody or antigenbinding fragment thereof.

8. The method of any one of the preceding claims, wherein the agent binds to the same epitope of CD91 as the spike protein of the coronavirus, or to an overlapping epitope.

9. The method of any one of the preceding claims, wherein the agent binds to and / or interacts with any one of amino acids 1060, 1061, 1065-1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090-1092, 1095-1099 of SEQ ID NO: 1, or any combination thereof.

10. The method of any one of the preceding claims, wherein the agent binds to and / or interacts with residue 1097 of SEQ ID NO: 1.

11. The method of any one of the preceding claims, wherein binding of the agent and / or interaction with the amino acids is determined by X ray crystallography.

12. A method of treating or preventing coronavirus infection, the method comprising administering a therapeutically effective amount of an agent to a subject, wherein the agent reduces the expression of CD91.

13. The method of claim 12, wherein the agent selectively or specifically reduces the expression of CD91 in the endothelium and / or on the surface of cells.

14. The method of claim 12 or 13, wherein the agent is a nucleic acid, optionally an antisense oligonucleotide or small interfering RNA.

15. The method of any of claims 12-14, wherein the agent promotes the intracellular retention and / or degradation of CD91.

16. The method of any one of the preceding claims, wherein the treating or preventing coronavirus infection comprises reducing viral load or viral replication in the subject.

17. The method of any one of the preceding claims, wherein the treating or preventing coronavirus infection comprises reducing adverse effects associated with coronavirus infection.

18. The method of claim 17, wherein the adverse effects are cardiovascular adverse effects, immunological adverse effects and / or respiratory adverse effects.

19. The method of claim 18, wherein the cardiovascular adverse effects are blood clotting, heart attack, stroke, pulmonary embolism and / or thrombosis.

20. The method of any one of the preceding claims, wherein the method further comprises administering an antagonist of Angiotensin Converting Enzyme (ACE2), and / or an inhibitor of transmembrane protease serine 2 (TMRPSS2).

21. An agent that binds to CD91 for use in a method of treating and / or preventing coronavirus infection, wherein the method comprises administering a therapeutically effective amount of the agent to a subject, and wherein the agent is not lactoferrin or aprotinin.

22. The agent for use according to claim 21, wherein:(a) the coronavirus is SARS-CoV-2; and / or(b) the agent is as defined in any one of claims 3-11; and / or(c) the treatment / prevention is as defined in any one of claims 16-19; and / or(d) the method further comprises administering an antagonist of Angiotensin Converting Enzyme (ACE2), and / or an inhibitor of transmembrane protease serine 2 (TMRPSS2).

23. Use of an agent that binds to CD91 in the manufacture of a medicament for treating and / or preventing coronavirus infection, and wherein the agent is not lactoferrin or aprotinin.

24. The use of claim 23, wherein:(a) the coronavirus is SARS-CoV-2; and / or(b) the agent is as defined in any one of claims 3-11; and / or(c) the treatment / prevention is as defined in any one of claims 16-19; and / or(d) the treatment and / or prevention further comprises administering an antagonist of Angiotensin Converting Enzyme (ACE2), and / or an inhibitor of transmembrane protease serine 2 (TMRPSS2).

25. An agent that reduces the expression of CD91 for use in a method of treating and / or preventing coronavirus infection, wherein the method comprises administering a therapeutically effective amount of the agent to a subject.

26. The agent for use according to claim 25, wherein:(a) the coronavirus is SARS-CoV-2; and / or(b) the agent is as defined in any one of claims 13-15; and / or(c) the treatment / prevention is as defined in any one of claims 16-19; and / or(d) the treatment and / or prevention further comprises administering an antagonist of Angiotensin Converting Enzyme (ACE2), and / or an inhibitor of transmembrane protease serine 2 (TMRPSS2).

27. Use of an agent that reduces the expression of CD91 in the manufacture of a medicament for treating and / or preventing coronavirus infection.

28. The use according to claim 27, wherein:(a) the coronavirus is SARS-CoV-2; and / or(b) the agent is as defined in any one of claims 13-15; and / or(c) the treatment / prevention is as defined in any one of claims 16-19; and / or(d) the treatment and / or prevention further comprises administering an antagonist of Angiotensin Converting Enzyme (ACE2), and / or an inhibitor of transmembrane protease serine 2 (TMRPSS2).

29. An anti-CD91 antibody which binds to an epitope comprising one or more amino acids selected from the group consisting of positions 1060, 1061, 1065-1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090-1092, 1095-1099 of SEQ ID NO: 1.

30. The antibody of claim 29, which binds to an epitope comprising position 1097 of SEQ ID NO: 1.

31. The antibody of claim 29 or 30, wherein the epitope is determined by x-ray crystallography, optionally wherein the epitope residues are within 4 A of an antibody paratope residue.

32. The antibody of any one of claims 29-31, which blocks the interaction of a coronavirus spike protein with CD91.

33. The antibody of claim 32, which blocks the interaction of the SARS CoV-2 spike protein with CD91.

34. A polynucleotide encoding an antibody according to any one of claims 29-33.

35. A vector comprising or encoding the polynucleotide of claim 34.

36. A host cell comprising the polynucleotide of claim 34 or the vector of claim 35.

37. A method of making the antibody of any one of claims 29-33, said method comprising culturing the host cell of claim 36 under conditions permitting production of the antibody and recovering the antibody so produced.

38. A pharmaceutical composition comprising the antibody of any one of claims 29-33.

39. A method of screening an agent for treating and / or preventing coronavirus infections, said method comprising:(a) testing the binding of a coronavirus spike protein to CD91 in the presence of the agent; and(b) identifying the agent as having utility in treating and / or preventing coronavirus infection if the agent inhibits or prevents the interaction of the coronavirus spike protein with CD91.

40. The method of claim 39, wherein the coronavirus is SARS-CoV-2.

41. The method of claim 39 or 40, which further comprises determining that the agent binds to and / or interacts with any one of amino acids 1060, 1061, 1065-1068, 1070, 1071, 1078, 1079, 1081, 1082, 1090-1092, 1095-1099 of SEQ ID NO: 1, or any combination thereof.

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