CD3 antibodies for treating coronavirus

Abstract

The present invention provides methods of treating and preventing symptoms of coronavirus infection using antibodies that recognize CD3. The present invention further provides routes of administration and formulations for use in the methods.

Description

[0001] Cross Reference to Related Applications

[0002] This application claims priority to U.S. Provisional Patent Application Serial No. 63 / 058,978, filed July 30, 2020, the contents of which are hereby incorporated by reference in their entirety.

[0003] Reference to a Sequence Listing

[0004] This application is being filed electronically via EFS-Web and includes an electronically submitted sequence listing in.txt format. The.txt file contains a sequence listing entitled “TIZI_028_001WO_SeqList_ST25.txt” created on July 29, 2021, and having a size of 33 kilobytes. The sequence listing contained in the.txt file is part of the specification and is herein incorporated by reference in its entirety. TECHNICAL FIELD

[0005] The present invention relates generally to compositions and methods for treating coronavirus infections and variants thereof (e.g., COVID-19, SARS, and MERS) by administering anti-CD3 antibodies alone or in combination with a steroid such as dexamethasone or an antiviral therapy. BACKGROUND

[0006] Coronaviruses are enveloped, non-segmented, positive-sense RNA viruses belonging to the Coronaviridae family. Coronaviruses can cause multi-system infections, primarily respiratory tract infections in humans, such as severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS). The novel coronavirus (2019-nCoV, COVID-19) has brought about a respiratory viral pandemic to the world population. Current efforts are focused on quarantining and isolating infected individuals. This outbreak can be controlled with a protective vaccine to prevent COVID-19 infection, but no vaccine to treat this virus has been available to date.

[0007] The human CD3 antigen is composed of a minimum of four invariant polypeptide chains that non-covalently associate with the T cell receptor on the surface of a T cell, and the antigen is now commonly referred to as the CD3 antigen complex. It is intimately involved in the process of T cell activation in response to antigen recognition by the T cell receptor.

[0008] Because of the fundamental nature of CD3 in initiating an anti-antigen response, monoclonal antibodies directed against this receptor are capable of blocking or at least modulating the immune process and thereby can be used as agents for disease.

[0009] Accordingly, there is a need for therapies that neutralize CD3 biological activity to treat and prevent coronavirus infections such as COVID-19 and associated symptoms. SUMMARY

[0010] In an aspect, the present disclosure provides a method of treating, preventing, or reducing symptoms of a coronavirus infection in a subject in need thereof, comprising administering to the subject a composition comprising an anti-CD3 antibody. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is fully human or humanized.

[0011] In some embodiments, the anti-CD3 antibody comprises a heavy chain complementarity determining region 1 (CDRH1) comprising the amino acid sequence GYGMH (SEQ ID NO: 42), a heavy chain complementarity determining region 2 (CDRH2) comprising the amino acid sequence VIWYDGSKKYYVDSVKG (SEQ ID NO: 43), a heavy chain complementarity determining region 3 (CDRH3) comprising the amino acid sequence QMGYWHFDL (SEQ ID NO: 44), a light chain complementarity determining region 1 (CDRL1) comprising the amino acid sequence RASQSVSSYLA (SEQ ID NO: 45), a light chain complementarity determining region 2 (CDRL2) comprising the amino acid sequence DASNRAT (SEQ ID NO: 46), and a light chain complementarity determining region 3 (CDRL3) comprising the amino acid sequence QQRSNWPPLT (SEQ ID NO: 47). In some embodiments, the anti-CD3 antibody comprises a variable heavy chain amino acid sequence comprising the amino acid sequence of SEQ ID NO: 48 and a variable light chain amino acid sequence comprising the amino acid sequence of SEQ ID NO: 49. In some embodiments, the anti-CD3 antibody comprises a heavy chain amino acid sequence comprising the amino acid sequence of SEQ ID NO: 50 and a light chain amino acid sequence comprising the amino acid sequence of SEQ ID NO: 51.

[0012] In some embodiments, the coronavirus is SARS-CoV, SARS-CoV-2, MERS-CoV, or a mutant or variant thereof. In some embodiments, the symptom of a coronavirus infection is one or more of: an overactive immune response, fever, gastrointestinal symptoms, respiratory symptoms, anosmia (loss of smell), dysgeusia (loss of taste), cough, headache, sore throat, pain when swallowing, dyspnea, difficulty breathing, tachypnea, nausea, vomiting, decreased O2 saturation, diarrhea, rhinorrhea, abdominal pain, myalgia, fever, conjunctivitis, and loss of appetite. In some embodiments, the overactive immune response comprises increased levels of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), and D-dimer. In some embodiments, the levels of at least one of IL-6, CRP, and D-dimer are reduced.

[0013] In some embodiments, the subject has or is suspected of having a coronavirus infection. In some embodiments, the subject has been or is believed to have been exposed to a coronavirus and has not yet developed symptoms of a coronavirus infection.

[0014] In some embodiments, the method further comprises administering to the subject a composition comprising dexamethasone.

[0015] In some embodiments, the composition is administered orally, mucosally, by inhalation, nasally, intravenously, or any combination thereof. In some embodiments, the inhalation administration is by inhaler or nebulizer.

[0016] In some embodiments, the method further comprises administering an anti-TNFa antibody, an anti-CD20 antibody, an anti-IFNy antibody, an anti- granulocyte-macrophage colony-stimulating factor antibody, or an anti-IL-6R antibody. In some embodiments, the method further comprises administering to the subject an antiviral drug, an immunopotentiating drug, vitamin C, vitamin D, vitamin E, or any combination thereof. In some embodiments, the antiviral drug is azidothymidine, remdesivir, or actinomycin D.

[0017] In some embodiments, the anti-CD3 is administered nasally at a daily dose of 50 pg to 100 pg. In some embodiments, the daily dose is administered once daily. In some embodiments, the daily dose is administered at least for 10 consecutive days. In some embodiments, the anti-CD3 is administered orally at a daily dose of 1.0 to 2.5 mg.

[0018] In some embodiments, the subject is further administered dexamethasone. In some embodiments, the dexamethasone is administered by inhalation. In some embodiments, the dexamethasone is administered by inhalation by metered dose inhaler. In one aspect, the disclosure provides a nasal or inhalation formulation comprising an anti-CD3 antibody and dexamethasone.

[0019] In one aspect, the disclosure provides a method of treating, preventing, or alleviating symptoms of a disease or condition in a subject in need thereof, comprising administering to the subject a composition comprising an anti-CD3 antibody.

[0020] In some embodiments, the subject has an inflammatory disease or condition. In some embodiments, the inflammatory disease or condition is autoimmune encephalomyelitis, lupus, or arthritis. In some embodiments, the subject has a pulmonary disease or condition. In some embodiments, the pulmonary disease or condition is acute respiratory distress syndrome (ARDS). In some embodiments, the subject has a neurodegenerative disease. In some embodiments, the subject has a neurodegenerative disease or condition. In some embodiments, the neurodegenerative disease or condition is multiple sclerosis. In some embodiments, the multiple sclerosis is secondary progressive multiple sclerosis.

[0021] In one aspect, the present disclosure provides a method of treating, preventing, or reducing symptoms of a disease or condition in a subject in need thereof, comprising: a) collecting a sample from the subject; b) measuring a marker of overactive immune response; and c) administering to the subject a composition comprising an anti-CD3 antibody based on the level of the marker. In some embodiments, the disease or condition is a coronavirus infection, an inflammatory disease or condition, a pulmonary disease or condition, or a neurodegenerative disease or condition. In some embodiments, the marker of overactive immune response is at least one of interleukin 6 (IL-6), C-reactive protein (CRP), D-dimer, interferon (IFN), interferon alpha (IFN-alpha), interferon gamma (IFN-gamma), interleukin 1 beta (IL-1 beta), and / or CXCL10. In some embodiments, the level of the marker is increased or higher compared to a healthy subject. In some embodiments, the anti-CD3 antibody is Foselumab.

[0022] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice of the present application, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are expressly incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples described herein are illustrative only and are not intended to be limiting.

[0023] Other features and advantages of the present application will be apparent from the following detailed description, and from the claims, and will be encompassed by the appended claims. BRIEF DESCRIPTION OF DRAWINGS

[0024] Figure 1 is a diagram showing the various stages of COVID-19 pathogenesis progression. Through the use of therapeutic interventions as shown, as well as combination therapy, it is expected that the progression of the disease is delayed. Dexamethasone treatment is used to suppress the underlying inflammatory mechanisms, while anti-CD3 treatment is to activate the mucosal immune system to suppress the cytokine storm.

[0025] Figure 2 is a graphic showing a brief overview of the clinical design. The patient population selected for the study will be early stage patients who are symptomatic but have not progressed to the severe stage requiring the use of a ventilator. The clinical study consists of three arms of 6 to 10 patients randomized. The doses of dexamethasone using pMDI administration and Foselumab administered through a nasal spray device are as shown. The clinical endpoints and study duration are the same as shown in the figure.

[0026] Figure 3is an illustration showing a brief overview of the clinical design. The patient population selected for the study will be early-stage patients who are symptomatic but have not progressed to the severe stage requiring the use of a ventilator. The clinical study consists of three groups of 6 to 10 patients randomized. The doses of azidothymidine administered and foralumab administered by a nasal spray device are as shown. The clinical endpoints and study duration are the same as shown in the figure.

[0027] Figure 4 is a photograph of: A) a hand-held metered dose inhaler (pMDI) and B) a nasal spray device for nasal administration of foralumab. The device can also be used for nasal administration of anti-IL-6R.

[0028] Figure 5 is a graph showing a clinical study design for a method of treating COVID-19 with an anti-CD3 antibody (foralumab).

[0029] Figure 6A Plots and charts showing reduction of IL-6 in subjects with COVID-19 treated with an anti-CD3 antibody (foralumab) and dexamethasone, anti-CD3 antibody (foralumab) alone, or placebo (control). IL = interleukin; CRP = C-reactive protein; Dexa = dexamethasone.

[0030] Figure 6B Plots and charts showing reduction of C-reactive protein in subjects with COVID-19 treated with an anti-CD3 antibody (foralumab) and dexamethasone, anti-CD3 antibody (foralumab) alone, or placebo (control).

[0031] Figure 7 A series of lung CT scan images of subjects with COVID-19 treated with an anti-CD3 antibody (foralumab) and dexamethasone (Dexa), anti-CD3 antibody (foralumab) alone, or placebo (control). DETAILED DESCRIPTION

[0032] The present invention provides monoclonal antibodies that specifically bind to human CD3 for use in treating, preventing, or alleviating symptoms of a coronavirus infection, such as SARS-CoV-2 (i.e., COVID-19), SARS-CoV-1, and Middle East Respiratory Syndrome Coronavirus (MERS-CoV). These antibodies are collectively referred to herein as “huCD3” antibodies. The antibodies can be, for example, fully human.

[0033] The exact mechanism of SARS-CoV-2 pathogenesis remains to be discovered, suggesting that the features of the clinical presentation of severe SARS-CoV-2 infection are also characterized by an overexuberant immune response accompanied by lung lymphomononuclear cell infiltration and proliferation, which can contribute more to tissue damage than the direct effects of viral replication.

[0034] SARS-CoV-2 involves a two-step process: a virus-mediated process and an immune-mediated process. The first step of the virus-mediated process is characterized mainly by acute viral symptoms, fever, myalgia, cough, and is a direct result of viral infection of cells. During the viral phase, cell infection occurs in various organs via virus-specific receptors and is followed by clinical deterioration. The viral phase is followed by a second phase characterized by an inappropriate exuberant immune response involving multiple cytokines and immune cells, which causes immune-mediated end-organ damage. The second immune-mediated phase is associated with life-threatening complications, most commonly acute respiratory distress syndrome (ARDS), which is caused by an overexuberant host response and is referred to as cytokine storm.

[0035] Given the overexuberant inflammatory role of coronavirus infection (such as SARS-CoV-2 (COVID19)), drugs that modulate the immune response are being explored as adjunctive therapy for the management of moderate to severe COVID-19.

[0036] As disclosed herein, modulation of regulatory T cells (Tregs) by administration of anti-CD3 antibodies can be a treatment for overexuberant immune responses. Tregs migrate into inflamed tissues, suppress inflammatory responses, and accelerate tissue repair. Without being bound by theory, the present inventors believe that induction of regulatory T cells by administration of anti-CD3 antibodies can be used as a treatment for diseases and conditions associated with overexuberant inflammatory responses, including but not limited to COVID-19, ARDS, inflammatory conditions, and neurodegenerative diseases.

[0037] Surprisingly, the present inventors have demonstrated that mucosal (oral and nasal) administration of anti-CD3 monoclonal antibodies is immunomodulatory and suppresses overexuberant immune responses in a number of inflammatory and autoimmune diseases, including models of multiple sclerosis, diabetes, arthritis, lupus, colitis, and graft rejection. The mechanism of action includes induction of regulatory T cells, downregulation of Th1 and Th17 cells, and downregulation of CD8 cells.

[0038] Further, the present inventors have shown in human studies that full humanized anti-CD3 Mab (Fresolimumab) was administered nasally to healthy volunteers at a dose range of 10 ug, 50 ug and 250 ug per dose, administered for 5 consecutive days. Fresolimumab administered nasally at a dose of 50 ug was well tolerated and suppressed cytotoxic CD8+ as well as CD8+ cells secreting perforin and the treatment also suppressed the pro-inflammatory cytokine IFN-g. Thus, oral or nasal administration of Fresolimumab has immunomodulatory properties which would be beneficial in treating the hyperactive immune response that accompanies COVID infection.

[0039] Thus, in various aspects, the present invention provides methods of treating immune activation in Covid-19 by administering anti-CD3 mAb alone or in combination with dexamethasone. The treatment can be administered a) anti-CD3 administered nasally; b) anti-CD3 administered orally; c) anti-CD3 administered in combination nasally and orally and d) anti-CD3 administered nasally or orally in combination with dexamethasone. In particular, anti-CD3 mAb at a suitable dose can be administered nasally by a hand-held spray device alone or in combination with oral administration of anti-CD3. The present invention also relates to direct lung delivery of anti-CD3 mAb and dexamethasone which is delivered by a hand-held inhaler for the treatment of COVID-19 patients.

[0040] Direct delivery of drugs by inhalation is advantageous because when compared to other routes of administration, the same level of efficacy can be achieved at much lower doses and also minimizes side effects. Inhalation therapy is an effective treatment for lung diseases because the drug is delivered directly into the lungs. Dry powder inhalers (DPI) are portable solid powder delivery units without propellants. DPIs can target the drug directly to deep sites in the lungs. Metered dose inhalers (MDI) are small devices that deliver a measured amount of medication to your lungs. When you inhale, a calibrated dose of medication is delivered with each puff (breath).

[0041] Anti-CD3 antibodies

[0042] Antibodies specific for the CD3 epsilon chain (CD3 epsilon) and antigen-binding fragments thereof are referred to herein as "anti-CD3 antibodies" or "CD3 antibodies," and compositions are referred to herein as "anti-CD3 antibody compositions." Any anti-CD3 antibody known in the art is suitable for use in the present disclosure. Anti-CD3 antibodies are monoclonal antibodies.

[0043] The anti-CD3 antibody can be any antibody specific for CD3. The anti-CD3 antibody can be polyclonal, monoclonal, recombinant (e.g., chimeric, de-immunized, or humanized), fully human, non-human (e.g., murine) single-chain, or single-domain antibody. In some embodiments, the antibody has effector function and can fix complement. In some embodiments, the antibody has reduced or no ability to bind an Fc receptor. For example, the anti-CD3 antibody can be of an isotype or subtype, fragment, or other mutant that does not support binding to an Fc receptor, e.g., it has a mutagenized or deleted Fc receptor binding region. The antibody can be conjugated to a toxin or an imaging agent.

[0044] A variety of anti-CD3 antibodies are known, including but not limited to OKT3 (muromonab / Orthoclone OKT3.TM., Ortho Biotech, Raritan, N.J.; U.S. Patent No. 4,361,549); hOKT3 (Herold et al., N.E.J.M. 346(22): 1692-1698 (2002); HuM291 (Nuvion.TM., Protein Design Labs, Fremont, Calif.); gOKT3-5 (Alegre et al., J. Immunol. 148(11):3461-8 (1992); 1F4 (Tanaka et al., J. Immunol. 142:2791-2795 (1989)); G4.18 (Nicolls et al., Transplantation 55:459-468 (1993)); 145-2C11 (Davignon et al., J. Immunol. 141(6): 1848-54 (1988)); and as described in Frenken et al., Transplantation 51(4):881-7 (1991); U.S. Patent Nos. 6,491,9116, 6,406,696, and 6,143,297.

[0045] Methods of making such antibodies are also known. Full-length CD3 protein or antigenic peptide fragments of CD3 can be used as immunogens, or can be used to identify anti-CD3 antibodies made with other immunogens (e.g., cells, membrane preparations, etc., e.g., E-rosette-positive purified normal human peripheral T cells as described in U.S. Patent Nos. 4,361,549 and 4,654,210). The anti-CD3 antibody can bind an epitope on any domain or region on CD3.

[0046] Chimeric, humanized, de-immunized, or fully human antibodies are desirable for applications involving repeated administration, e.g., therapeutic treatment of a human subject.

[0047] Chimeric antibodies contain portions of two different antibodies, typically of two different species. Generally, such antibodies contain human constant regions and variable regions from another species, such as murine variable regions. For example, mouse / human chimeric antibodies have been reported that exhibit the binding properties of the parent mouse antibody, as well as effector functions associated with the human constant regions. See, e.g., Cabilly et al., U.S. Patent No. 4,816,567; Shoemaker et al., U.S. Patent No. 4,978,745; Beavers et al., U.S. Patent No. 4,975,369; and Boss et al., U.S. Patent No. 4,816,397, all of which are incorporated herein by reference. Typically, these chimeric antibodies are constructed by preparing genomic gene libraries from DNA extracted from pre-existing murine hybridomas (Nishimura et al., Cancer Research, 47:999 (1987)). Variable region genes are then screened from the libraries for both heavy and light chains that exhibit the correct pattern of antibody fragment rearrangement. Alternatively, cDNA libraries are prepared from RNA extracted from hybridomas and screened, or variable regions are obtained by polymerase chain reaction. The cloned variable region genes are then ligated into expression vectors in cloning cassettes containing appropriate heavy or light chain human constant region genes. The chimeric genes can then be expressed in selected cell lines, such as murine myeloma lines. Such chimeric antibodies have been used in human therapy.

[0048] Humanized antibodies are known in the art. Generally, "humanization" produces an antibody with reduced immunogenicity in humans, while retaining all the antigen-binding properties of the original molecule. To retain all the antigen-binding properties of the original antibody, the structure of its combining site must be faithfully reproduced in the "humanized" version. This can potentially be achieved by grafting the combining site of a non-human antibody onto a human framework by: (a) grafting the entire non-human variable domain onto a human constant region to produce a chimeric antibody (Morrison et al., Proc. Natl. Acad. Sci., USA 81 :6801 (1984); Morrison and Oi, Adv. Immunol. 44:65 (1988)) (which preserves the ligand-binding properties, but also retains the immunogenicity of the non-human variable domain); (b) grafting only the non-human CDRs onto a human framework and constant region, with or without retention of key framework residues (Jones et al., Nature, 321 :522 (1986); Verhoeyen et al., Science 239:1539 (1988)); or (c) grafting the entire non-human variable domain (to preserve the ligand-binding properties), but also "cloaking" them with a human-like surface by judiciously replacing exposed residues (to reduce antigenicity) (Padlan, Molec. Immunol. 28:489 (1991)).

[0049] Humanization by CDR grafting generally involves grafting only the CDRs onto a human framework and constant region of a human fragment. In theory, this should substantially eliminate immunogenicity (unless there are allotype or idiotype differences). However, it has been reported that some framework residues of the original antibody also need to be preserved (Riechmann et al., Nature 332:323 (1988); Queen et al., Proc. Natl. Acad. Sci. USA 86:10,029 (1989)). Framework residues that need to be preserved can be identified by computer modeling. Alternatively, key framework residues can be potentially identified by comparing known antibody combining site structures (Padlan, Molec. Immun. 31(3): 169-217 (1994)). The compositions and methods of the present disclosure also include partially humanized antibodies, in which the six CDRs of the heavy and light chains of a murine monoclonal antibody, as well as a limited number of structural amino acids, are grafted onto a CDR-depleted human IgG scaffold by recombinant technology (Jones et al., Nature 321 :522-525 (1986)).

[0050] Deimmunized antibodies are made by replacing immunogenic epitopes in the murine variable domains with benign amino acid sequences to produce deimmunized variable domains. The deimmunized variable domains are ligated to human IgG constant domain genes to produce deimmunized antibodies (Biovation, Aberdeen, Scotland).

[0051] The anti-CD3 antibody can also be a single chain antibody. Single chain antibodies (scFV) can be engineered (see, e.g., Colcher et al., Ann. N. Y. Acad. Sci. 880:263-80 (1999); and Reiter, Clin. Cancer Res. 2:245-52 (1996)). Single chain antibodies can be dimerized or multimerized to produce multivalent antibodies that are specific for different epitopes of the same target CD3 protein. In some embodiments, the antibody is monovalent, e.g., as described in Abbs et al., Ther. Immunol. 1(6):325-31 (1994), incorporated herein by reference.

[0052] Exemplary anti-CD3 antibodies include a heavy chain complementarity determining region 1 (CDRH1) comprising the amino acid sequence GYGMH (SEQ ID NO: 42), a heavy chain complementarity determining region 2 (CDRH2) comprising the amino acid sequence VIWYDGSKKYYVDSVKG (SEQ ID NO: 43), a heavy chain complementarity determining region 3 (CDRH3) comprising the amino acid sequence QMGYWHFDL (SEQ ID NO: 44), a light chain complementarity determining region 1 (CDRL1) comprising the amino acid sequence RASQSVSSYLA (SEQ ID NO: 45), a light chain complementarity determining region 2 (CDRL2) comprising the amino acid sequence DASNRAT (SEQ ID NO: 46), and a light chain complementarity determining region 3 (CDRL3) comprising the amino acid sequence QQRSNWPPLT (SEQ ID NO: 47).

[0053] In some embodiments, the anti-CD3 antibody comprises a variable heavy chain amino acid sequence comprising QVQLVESGGGVVQPGRSLRLSCAASGFKFSGYGMHWVRQAPGKGLEWVAVIWYDGSKKYYVDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARQMGYWHFDLWGRGTLVTVSS (SEQ ID NO: 48) and a variable light chain amino acid sequence comprising EIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPPLTFGGGTKVEIK (SEQ ID NO: 49).

[0054] Preferably, the anti-CD3 antibody comprises a heavy chain amino acid sequence comprising: QVQLVESGGGVVQPGRSLRLSCAASGFKFSGYGMHWVRQAPGKGLEWVAVIWYDGSKKYYVDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARQMGYWHFDLWGRGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 50), and a light chain amino acid sequence comprising: EIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 51). Such an anti-CD3 antibody is referred to herein as NI-0401, foralumab, or 28F11-AE. (See, e.g., Dean Y, Dépis F, Kosco-Vilbois M. “Combination therapies in the context of anti-CD3 antibodies for the treatment of autoimmune diseases”, Swiss Med Wkly. (2012) (the contents of which are incorporated by reference in their entirety).

[0055] In some embodiments, the anti-CD3 antibody is a fully human or humanized antibody. In some embodiments, the anti-CD3 antibody formulation includes a full-length anti-CD3 antibody. In some embodiments, the anti-CD3 antibody formulation includes an antibody fragment that specifically binds CD3. In some embodiments, the anti-CD3 antibody formulation includes a combination of a full-length anti-CD3 antibody and an antigen-binding fragment that specifically binds CD3.

[0056] In some embodiments, the antibody or antigen-binding fragment thereof that binds CD3 is a monoclonal antibody, a domain antibody, a single chain, a Fab fragment, a F(ab’)2 fragment, a scFv, a scAb, a dAb, a single domain heavy chain antibody, or a single domain light chain antibody. In some embodiments, such antibody or antigen-binding fragment thereof that binds CD3 is a mouse antibody, other rodent antibody, a chimeric antibody, a humanized antibody, or a fully human monoclonal antibody.

[0057] Optionally, the anti-CD3 antibody or antigen-binding fragment thereof for use in the formulations of the disclosure includes at least one amino acid mutation. Typically, the mutation is in the constant region. The mutation results in an antibody with altered effector function. Effector function of an antibody is altered by changing (i.e., enhancing or decreasing) the affinity of the antibody for an effector molecule, such as an Fc receptor or a complement component. For example, the mutation results in an antibody that is capable of reducing the release of cytokines from T cells. For example, the mutation is at amino acid residue 234, 235, 265, or 297, or a combination thereof, in the heavy chain. Preferably, the mutation results in an alanine residue at position 234, 235, 265, or 297, or an glutamic acid residue at position 235, or a combination thereof.

[0058] Preferably, the anti-CD3 antibodies provided herein contain one or more mutations that prevent heavy chain constant region-mediated release of one or more cytokines in vivo.

[0059] In some embodiments, the anti-CD3 antibody or antigen-binding fragment thereof for use in the formulations of the disclosure is a fully human antibody. A fully human CD3 antibody as used herein includes, for example, L234A and L235E mutations in the Fc region such that cytokine release upon exposure to the anti-CD3 antibody is significantly reduced or eliminated. The L234A and L235E mutations in the Fc region of the anti-CD3 antibodies provided herein reduce or eliminate cytokine release upon exposure of the anti-CD3 antibody to human leukocytes, while the following mutations maintain a significant cytokine release capacity. Significant reduction in cytokine release is defined, for example, by comparing the level of cytokine release upon exposure to an anti-CD3 antibody with L234A and L235E mutations in the Fc region to the level of cytokine release upon exposure to another anti-CD3 antibody with one or more of the following mutations. Other mutations in the Fc region include, for example, L234A and L235A, L235E, N297A, D265A, or a combination thereof.

[0060] The term "cytokine" refers to all human cytokines known in the art that bind to extracellular receptors expressed on the surface of cells and thereby modulate cell function, including but not limited to IL-2, IFN-g, TNF-a, IL-4, IL-5, IL-6, IL-9, IL-10, and IL-13.

[0061] The anti-CD3 formulation comprises a unit dose of anti-CD3 antibody in the range of about 0.01 mg to about 25 mg; or 0.01 mg to about 10 mg. For example, the unit dose is about 0.01, 0.02, 0.03, 0.04, 0.50, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9, 9.5, 10 mg or more. Preferably, the unit dose is 0.05 mg, 0.1 mg, 0.5 mg, 1.0 mg, 2.5 mg, 5.0 mg, or 10 mg.

[0062] The anti-CD3 antibody formulation includes one or more salts (buffered salts), one or more polyols, and one or more excipients. The formulations of the present disclosure can also contain a buffer or a preservative. The anti-CD3 antibody formulation is buffered in a solution at a pH in the range of about 4 to 8; in the range of about 4 to 7; in the range of about 4 to 6; in the range of about 5 to 6; or in the range of about 5.5 to 6.5. Preferably, the pH is 5.5.

[0063] Examples of salts include those prepared from hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, maleic acid, acetic acid, salicylic acid, citric acid, boric acid, formic acid, malonic acid, succinic acid, and the like. Such salts can also be prepared as alkali metal or alkaline earth metal salts, such as sodium, potassium, or calcium salts. Examples of buffers include phosphate, citrate, acetate, and 2-(N-morpholino)ethanesulfonic acid (MES).

[0064] The formulations of the present disclosure can include a buffer system. As used in this application, the term "buffer" or "buffer system" refers to a compound, usually in combination with at least one other compound, that provides a buffer system in solution that exhibits a buffering capacity, i.e., the ability to neutralize an acid or a base (alkali) within certain limits, while changing the original pH relatively little or not at all.

[0065] Buffering agents include borate buffers, phosphate buffers, calcium buffers, and combinations and mixtures thereof. Borate buffers include, for example, boric acid and salts thereof, such as sodium or potassium borate. Borate buffers also include compounds that generate boric acid or salts thereof in solution, such as potassium or potassium metaborate.

[0066] Phosphate buffer systems include one or more of an acid phosphate, a dibasic phosphate, and the like. Particularly useful phosphate buffers are those selected from the group consisting of alkali and / or alkaline earth metal phosphates. Examples of suitable phosphate buffers include one or more of disodium hydrogen phosphate (Na2HP04), sodium dihydrogen phosphate (NaH2P04), and potassium dihydrogen phosphate (KH2P04). Phosphate buffer components are often used in amounts from 0.01% or to 0.5% (w / v) (calculated as phosphate ion).

[0067] Other known buffering compounds, such as citrate, sodium bicarbonate, TRIS, and the like, can optionally be added to CD3 formulations, depending on the formulation. Other ingredients in the solution, while serving other functions, can also affect the buffering capacity. For example, EDTA, often used as a complexing agent, can have a significant effect on the buffering capacity of the solution.

[0068] Preferred salts for use in formulations of the present disclosure include sodium chloride, sodium acetate, sodium acetate trihydrate, and sodium citrate.

[0069] The concentration of salt in formulations according to the present disclosure is between about 10 mM and 500 mM, between about 25 mM and 250 mM, between about 25 mM and 150 mM.

[0070] The concentration of sodium acetate trihydrate ranges from about 10 mM to 100 mM. For example, the sodium acetate trihydrate is about 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 mM. Preferably, the sodium acetate trihydrate is 25 mM.

[0071] The concentration of sodium chloride ranges from about 50 mM to 500 mM. For example, the sodium chloride is about 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, or 500 mM. Preferably, the concentration of sodium chloride is about 125 mM.

[0072] The concentration of sodium citrate ranges from about 10 mM to 100 mM. For example, the sodium citrate is about 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 mM. Preferably, the sodium citrate ranges from about 25 to 50 mM.

[0073] In some embodiments, the salt is sodium acetate trihydrate at a concentration ranging from about 25 mm to 100 mm and sodium chloride at a concentration ranging from about 150 mm to 500 mm.

[0074] Preferably, the formulation includes about 25 mM sodium acetate trihydrate and about 150 mM sodium chloride.

[0075] The formulation includes one or more polyols as a bulking agent and / or a stabilizing excipient. The polyol includes, for example, trehalose, mannitol, maltose, lactose, sucrose, sorbitol, or glycerol. The concentration of the polyol ranges from about 0.1% to 50% or 5% to 25%. For example, the polyol is about 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50%.

[0076] In some embodiments, the polyol is trehalose at a concentration ranging from about 1% to 50% or 5% to 25%. For example, the trehalose is about 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50%. Preferably, the concentration of trehalose is about 10% or about 20%. Most preferably, the concentration of trehalose is about 20%.

[0077] In some embodiments, the polyol is sorbitol at a concentration ranging from about 1% to about 10%. In some embodiments, the polyol is glycerol at a concentration ranging from about 1% to about 10%.

[0078] In some embodiments, the polyol is mannitol at a concentration ranging from about 0.1% to about 10%. In some embodiments, the polyol is maltose at a concentration ranging from about 1% to about 10%.

[0079] Formulations include one or more excipients and / or surfactants to inhibit or otherwise reduce antibody aggregation. Suitable excipients that reduce antibody aggregation include, by way of non-limiting example, surfactants such as, by way of non-limiting example, polysorbate 20 or polysorbate 80. In some embodiments, polysorbate 20 or polysorbate 80 is present at a concentration ranging from about 0.01 to 1% or about 0.01 to 0.05%. For example, the concentration of polysorbate 20 or polysorbate 80 is about 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1.0%.

[0080] Preferably, the surfactant is polysorbate 80 at a concentration ranging from about 0.01 to 0.05%. More preferably, the polysorbate 80 is 0.02%.

[0081] Formulations include one or more excipients to reduce antibody oxidation. Suitable excipients that reduce antibody oxidation include, by way of non-limiting example, antioxidants. Antioxidants include, for example, methionine, D-arginine, BHT, or ascorbic acid. Antioxidants are present at a concentration ranging from about 0.01% to 1%; 0.1% to 1%; or 0.1% to 0.5%. In some embodiments, the antioxidant is methionine. In some embodiments, methionine is present at a concentration ranging from about 0.01% to 1%; 0.1% to 1%; or 0.1% to 0.5%. For example, methionine is present at a concentration of about 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1.0%. Preferably, the methionine is about 0.1%.

[0082] Formulations include one or more chelating agents such as, for example, ethylenediaminetetraacetic acid (EDTA). Chelating agents are present at a concentration ranging from 0.01% to 1%; 0.1% to 1%; or 0.1% to 0.5%. For example, chelating agents are present at a concentration of about 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1.0%. Preferably, the chelating agent is EDTA at a concentration of about 0.1%.

[0083] In some embodiments, formulations include one or more excipients to increase stability. In some embodiments, the excipient that increases stability is human serum albumin. In some embodiments, human serum albumin is present in a range of about 1 mg to about 5 mg.

[0084] In some embodiments, the formulation includes magnesium stearate (Mg stearate), an amino acid, or both magnesium stearate and an amino acid. Suitable amino acids include, for example, leucine, arginine, histidine, or combinations thereof.

[0085] In some embodiments, the one or more additional excipients is low moisture microcrystalline cellulose (such as Avicel), polyethylene glycol (PEG), or starch.

[0086] Other examples of pharmaceutically acceptable carriers and excipients that can be used in the formulations of the present disclosure include, but are not limited to, binders, fillers, disintegrants, lubricants, antimicrobials, antioxidants, and coating agents such as: binders: corn starch, potato starch, other starches, gelatin, natural and synthetic gums such as acacia, xanthan gum, sodium alginate, alginic acid, other alginates, powdered tragacanth, guar gum, cellulose and its derivatives (e.g., ethyl cellulose, cellulose acetate, carboxymethyl cellulose calcium, sodium carboxymethyl cellulose), polyvinylpyrrolidone (e.g., povidone, cross-linked povidone, copovidone, etc.), methyl cellulose, Methocel, pregelatinized starch (e.g., STARCH 1500® and STARCH 1500 LM® sold by Colorcon, Ltd.), hydroxypropyl methylcellulose, microcrystalline cellulose (FMC Corporation, Marcus Hook, PA, USA), Emdex, Plasdone, or mixtures thereof, fillers: talc, calcium carbonate (e.g., granular or powdered), dibasic calcium phosphate, tribasic calcium phosphate, calcium sulfate (e.g., granular or powdered), microcrystalline cellulose, powdered cellulose, dextrates, kaolin, mannitol, silicic acid, sorbitol, starch, pregelatinized starch, dextrose, fructose, honey, lactose anhydrous, lactose monohydrate, lactose and aspartame, lactose and cellulose, lactose and microcrystalline cellulose, maltodextrin, maltose, mannitol, microcrystalline cellulose and guar gum, molasses, sucrose, or mixtures thereof, disintegrants: agar-agar, alginic acid, calcium carbonate, microcrystalline cellulose, croscarmellose sodium, crospovidone, polacrilin potassium, sodium starch glycolate (such as Explotab), potato or tapioca starch, other starches, pregelatinized starch, clays, other alginates, other celluloses, gums (such as gellan gum), low-substituted hydroxypropyl cellulose, cross-linked polyplasdone, or mixtures thereof, lubricants: calcium stearate, magnesium stearate, mineral oil, light mineral oil, glycerin, sorbitol, mannitol, polyethylene glycol, other glycols, compritol, stearic acid, sodium lauryl sulfate, sodium stearyl fumarate (such as Pruv), vegetable-based fatty acid lubricants, talc, hydrogenated vegetable oil (e.g., peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate, ethyl laurate, agar, syloid silica aerogel (AEROSIL 200, W.R. Grace Co., Baltimore, MD USA), an agglomerated aerosol of synthetic silica (Deaussa Co., Piano, TX USA), fumed silica (CAB-O-SIL, Cabot Co.BHA, BHT, EDTA, or mixtures thereof, and coating agents: sodium carboxymethylcellulose, cellulose acetate phthalate, ethyl cellulose, gelatin, a pharmaceutical glaze, hydroxypropyl cellulose, hydroxypropyl methyl cellulose (hypromellose), hydroxypropyl methyl cellulose phthalate, methyl cellulose, polyethylene glycol, polyvinyl acetate phthalate, shellac, sucrose, titanium dioxide, carnauba wax, microcrystalline wax, gellan gum, maltodextrin, methacrylate ester, microcrystalline cellulose, and carrageenan, or mixtures thereof.

[0087] Formulations can also include other excipients and classes thereof including, but not limited to, Pluronic®, poloxamer (such as Lutrol® and poloxamer 188), ascorbic acid, glutathione, protease inhibitors (e.g., soybean trypsin inhibitor, organic acids), pH lowering agents, creams and lotions (e.g., maltodextrin and carrageenan); materials for chewable tablets (e.g., dextrose, fructose, lactose monohydrate, lactose and aspartame, lactose and cellulose, maltodextrin, maltose, mannitol, microcrystalline cellulose and guar gum, crystalline sorbitol); parenterals (e.g., mannitol and povidone); plasticizers (e.g., dibutyl sebacate, plasticizers for coating, polyvinyl acetate phthalate); powder lubricants (e.g., glyceryl behenate); soft gelatin capsules (e.g., sorbitol special solution); spheres for coating (e.g., sucrose spheres); spheronization agents (e.g., glyceryl behenate and microcrystalline cellulose); suspending agents / gelling agents (e.g., carrageenan, gellan gum, mannitol, microcrystalline cellulose, povidone, sodium starch glycolate, xanthan gum); sweeteners (e.g., aspartame, aspartame and lactose, dextrose, fructose, honey, maltodextrin, maltose, mannitol, molasses, crystalline sorbitol, sorbitol special solution, sucrose); wet granulation agents (e.g., calcium carbonate, lactose anhydrous, lactose monohydrate, maltodextrin, mannitol, microcrystalline cellulose, povidone, starch), caramel, carboxymethylcellulose sodium, cherry cream flavor and cherry flavor, citric acid anhydrous, citric acid, confectioner’s sugar, D&C Red No. 33, D&C Yellow #10 Aluminum Lake, disodium edetate, 15% ethanol, FD&C Yellow No. 6 Aluminum Lake, FD&C Blue #1 Aluminum Lake, FD&C Blue No. 1, FD&C Blue No. 2 Aluminum Lake, FD&C Green No. 3, FD&C Red No. 40, FD&C Yellow No. 6 Aluminum Lake, FD&C Yellow No. 6, FD&C Yellow No. 10, glyceryl palmitostearate, glyceryl monostearate, indigotine, lecithin, mannitol, methyl and propyl parabens, monoammonium glycyrrhizinate, natural and artificial orange flavor, pharmaceutical glaze, poloxamer 188, polydextrose, polysorbate 20, polysorbate 80, povidone, pregelatinized corn starch, pregelatinized starch, red iron oxide, sodium saccharin, sodium carboxymethyl ether, sodium chloride, sodium citrate, sodium phosphate, strawberry flavor, synthetic black iron oxide, synthetic red iron oxide, titanium dioxide, and white wax.

[0088] The CD3 antibodies are formulated for enteral, parenteral, or nasal administration. For example, the CD3 antibodies are formulated for nasal, oral, inhalational, subcutaneous, or intravenous administration.

[0089] For enteral administration (i.e., oral), the formulation can be a capsule or tablet. Parenteral administration includes intravenous, subcutaneous, intramuscular, and intra-articular administration, and can be a liquid or lyophilized powder in a sealed vial or other container. The preferred oral dosage range is 0.1 mg to 5 mg per day. For example, a dosage of 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg, 0.6 mg, 0.7 mg, 0.8 mg, 0.9 mg, 1.0 mg, 1.5 mg, 2.0 mg, 2.5 mg, 3.0 mg, 3.5 mg, 4.0 mg, or 5.0 mg per day is administered. The dosage is administered once per day or twice per day.

[0090] For nasal administration, the formulation can be an aerosol in a sealed vial or other suitable container. The preferred nasal dosage range is 0.05 mg to 1 mg per day. For example, a dosage of 0.05 mg, 0.06 mg, 0.07 mg, 0.08 mg, 0.09 mg, 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg, 0.6 mg, 0.7 mg, 0.8 mg, 0.9 mg, or 1.0 mg per day is administered. The dosage is divided equally between each nostril. The dosage is administered once per day or twice per day.

[0091] In some embodiments, the anti-CD3 antibody formulation is a subcutaneous formulation. In some embodiments, the subcutaneous anti-CD3 antibody formulation is packaged in a sealed vial or other container. The preferred subcutaneous dosage range is 0.2 mg to 5 mg per day. For example, a dosage of 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg, 0.6 mg, 0.7 mg, 0.8 mg, 0.9 mg, 1.0 mg, 1.5 mg, 2.0 mg, 2.5 mg, 3.0 mg, 3.5 mg, 4.0 mg, or 5.0 mg per day is administered. The dosage is administered once per day or twice per day. A preferred formulation for subcutaneous administration is the preferred dosage of anti-CD3 antibody in 25 mM sodium acetate buffer, 125 mM sodium chloride, and 0.02% polysorbate 80 at pH 5.5.

[0092] In some embodiments, the anti-CD3 antibody formulation is an inhalation formulation. For inhalation administration, the formulation can be an aerosol in a sealed vial or other suitable container. Administration by inhalation can be in the form of an inhaler or nebulizer. The nebulizer and / or inhaler is handheld. Optionally, the nebulizer and / or inhaler can have different sizes to fit children and / or adults.

[0093] Preferred inhaled doses range from 0.1 mg to 5 mg per day. For example, a dose of 0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg, 0.6 mg, 0.7 mg, 0.8 mg, 0.9 mg, 1.0 mg, 1.5 mg, 2.0 mg, 2.5 mg, 3.0 mg, 3.5 mg, 4.0 mg, or 5.0 mg per day is administered. The dose is administered once daily or twice daily.

[0094] The particles of the granular formulation have a diameter of between about 1 mm to about 5 mm, for example, less than 5 mm in diameter, less than 4 mm in diameter, less than 3 mm in diameter, less than 2 mm in diameter, and about 1 mm in diameter.

[0095] The particles of the granular formulation comprising an anti-CD3 antibody or antigen-binding fragment thereof have an average diameter of between about 0.1 mm to about 50 mm. The particles of the granular formulation comprising an anti-CD3 antibody or antigen-binding fragment thereof have an average diameter of between about 1 mm to about 10 mm, for example, less than 10 mm in average diameter, less than 9 mm in average diameter, less than 8 mm in average diameter, less than 7 mm in average diameter, less than 6 mm in average diameter, less than 5 mm in average diameter, less than 4 mm in average diameter, less than 3 mm in average diameter, and about 2 mm in average diameter. In some embodiments, the particles have an average diameter of between about 2 mm and 5 mm. In some embodiments, the particles have an average diameter of between 2 mm and 5 mm, wherein each particle has a diameter of less than about 50 mm.

[0096] In some embodiments, the CD3 antibody is a prolonged release and controlled release formulation. Methods of producing prolonged release and controlled release formulations are known in the art and include, for example, the use of macroporous beads.

[0097] In some embodiments, the anti-CD3 antibody formulation comprises a full-length anti-CD3 antibody. In some embodiments, the anti-CD3 antibody formulation comprises an antibody fragment that specifically binds CD3. In some embodiments, the anti-CD3 antibody formulation comprises a combination of a full-length anti-CD3 antibody and an antigen-binding fragment that specifically binds CD3.

[0098] Methods of treatment, prevention, and alleviation of symptoms

[0099] The present disclosure provides methods of treating, preventing, or reducing symptoms of a coronavirus infection, cytokine release syndrome (Shimabukuro-Vornhagen et al., J Immunother Cancer 6(1): 56 (2018), which is incorporated herein by reference), an inflammatory disease, acute respiratory distress syndrome (ARDS), a neurodegenerative or pulmonary disease in a subject (i.e., a patient) in need thereof, comprising administering to the subject a composition comprising a CD3 antibody (anti-CD3).

[0100] The present disclosure further provides methods of treating, preventing, or reducing symptoms of a coronavirus infection, an inflammatory disease, acute respiratory distress syndrome (ARDS), a neurodegenerative or pulmonary disease in a subject in need thereof, comprising administering to the subject: a composition comprising a CD3 antibody and a composition comprising dexamethasone, wherein administration of the composition comprising a CD3 antibody and administration of the composition comprising dexamethasone can occur in any order or simultaneously.

[0101] The CD3 antibodies described herein can be used as therapeutic agents. Such agents will generally be used to treat, reduce, and / or prevent a disease or abnormal state associated with a coronavirus infection, an inflammatory disease, acute respiratory distress syndrome (ARDS), cytokine release syndrome, a neurodegenerative or pulmonary disease in a subject (e.g., a human patient). The treatment regimen is carried out by identifying a subject (e.g., a human patient) having (or at risk of developing) a disease or condition described herein using standard methods. A CD3 antibody formulation, preferably a CD3 antibody formulation having high specificity and high affinity for its target antigen, is administered to the subject, and the CD3 antibody formulation generally has an effect due to its binding to the target.

[0102] Administration of the antibody eliminates or inhibits, interferes with signaling or otherwise modulates the function of the target (e.g., CD3), and can modulate cellular behavior. Without being bound by theory, administration of an anti-CD3 antibody can, for example, induce T regulatory cells (Tregs) and suppress inflammation in autoimmune models. In some embodiments, the methods described herein comprise inducing T regulatory cells. In another example, intranasal administration of anti-CD3 induces IL-10-dependent Tregs that suppress inflammation and disease progression in inflammatory disease models such as experimental autoimmune encephalomyelitis, lupus, and arthritis. In some embodiments, the methods described herein comprise inducing IL-10-dependent T regulatory cells. In some embodiments, the methods described herein comprise suppressing inflammation. In another example, administration of anti-CD3 to subjects with secondary progressive multiple sclerosis, a neurodegenerative disease, is immunologically active as measured by suppression of CD8+ T cell responses and induction of CD4+ IL-10 responses.

[0103] In some embodiments, the methods provided herein treat, prevent, or reduce symptoms. In some embodiments, the symptoms are at least one acute symptom. An acute symptom is a symptom that resolves in less than one month, or a symptom that resolves within the expected timeframe for a given disease or condition. For example, a subject having a coronavirus infection (e.g., COVID-19) or a variant thereof can experience acute symptoms that resolve within 2 to 4 weeks, such as fever, sore throat, cough, shortness of breath, and chest pain.

[0104] In some embodiments, the symptoms are at least one chronic symptom. A chronic symptom is a symptom that persists in a subject having a disease or condition after a time considered normal for that disease or condition. For example, a subject having a coronavirus infection (e.g., COVID-19) or a variant thereof can experience chronic symptoms that can persist for more than 1 month, more than 2 months, more than 3 months, more than 4 months, more than 5 months, or more than 6 months after infection. A long-term inflammatory response can lead to chronic symptoms. In some embodiments, the chronic symptoms are at least one of fatigue, muscle aches and pains, poor sleep, cough, shortness of breath, orthopnea, leg swelling, exercise intolerance due to COVID-19 induced heart failure, pulmonary embolism, pulmonary fibrosis, ARDS associated with COVID-19, palpitations with mild exertion, night sweats, organ damage (e.g., heart or respiratory organ damage), and poor temperature control.

[0105] In some embodiments, the symptoms are an overactive immune response. An overactive immune response can include, for example, cytokine release syndrome, “cytokine storm,” and the like. In some embodiments, the overactive immune response comprises an increase in the level of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), D-dimer, interferon (IFN), interferon alpha (IFN-a), interferon gamma (IFN-g), interleukin 1 beta (IL-1b), and / or CXCL10. In some embodiments, the overactive immune response comprises an increase in the level of at least IL-6 and CRP. In some embodiments, the overactive immune response comprises an increase in the level of at least IL-6. In some embodiments, the overactive immune response comprises an increase in the level of at least CRP.

[0106] In some embodiments, the methods of treating, preventing, or reducing symptoms provided herein comprise measuring a symptom. In some embodiments, the symptom is an overactive immune response. In some embodiments, measuring an overactive immune response comprises determining a level of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), D-dimer, interferon (IFN), interferon alpha (IFN-a), interferon gamma (IFN-g), interleukin 1 beta (IL-1 beta), or CXCL10. In some embodiments, measuring an overactive immune response comprises determining a level of at least one of IL-6, CRP, and D-dimer. In some embodiments, measuring an overactive immune response comprises determining a level of at least one of IL-6 and CRP. In some embodiments, measuring an overactive immune response comprises determining a level of at least IL-6. In some embodiments, measuring an overactive immune response comprises determining a level of at least CRP.

[0107] In some embodiments, the methods of treating, preventing, or reducing symptoms comprise administering an anti-CD3 antibody to a subject based on an overactive immune response in the subject. In some embodiments, an anti-CD3 antibody is administered to a subject if the overactive immune response comprises a high level of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), D-dimer, interferon (IFN), interferon alpha (IFN-a), interferon gamma (IFN-g), interleukin 1 beta (IL-1 beta), or CXCL10. In some embodiments, an anti-CD3 antibody is administered to a subject if the overactive immune response comprises a high level of at least one of IL-6, CRP, or D-dimer. In some embodiments, an anti-CD3 antibody is administered to a subject if the overactive immune response comprises a high level of at least one of IL-6 and CRP. In some embodiments, an anti-CD3 antibody is administered to a subject if the overactive immune response comprises a high level of at least IL-6. In some embodiments, an anti-CD3 antibody is administered to a subject if the overactive immune response comprises a high level of at least CRP.

[0108] A therapeutically effective amount of an active ingredient (e.g., an anti-CD3 antibody of the application) generally involves an amount necessary to achieve the intended purpose. As indicated above, this can be a binding interaction between the antibody and its target antigen that, in some cases, interferes with the function of the target. The amount required to be administered will also depend on the binding affinity of the antibody for its specific antigen, and will also depend on the rate at which the administered antibody is depleted from the free volume of other subjects to which it is administered. As a non-limiting example, a common range for a therapeutically effective administration of an antibody or antibody fragment of the application can be from about 0.1 mg / kg body weight to about 50 mg / kg body weight. A common range for frequency of administration can be, for example, from once daily to once weekly.

[0109] The methods provided herein comprise administering anti-CD3 as a dose or unit dose. The dose should be sufficient to result in a reduction of symptoms associated with the disease or condition described herein (i.e., an effective amount). The dose selected should be sufficient to constitute effective treatment, but not so high as to cause unacceptable side effects (e.g., mucositis or anaphylactic shock). Preferably, the patient’s health status and the status of the disease or condition should be closely monitored during and for a reasonable period of time after treatment.

[0110] In some embodiments, the anti-CD3 is administered as a daily dose. In some embodiments, the anti-CD3 is administered nasally. In some embodiments, the daily dose administered nasally is from 50 µg to 100 µg. In some embodiments, the daily dose is about 20 µg, about 25 µg, about 30 µg, about 35 µg, about 40 µg, about 45 µg, about 50 µg, about 55 µg, about 60 µg, about 65 µg, about 70 µg, about 75 µg, about 80 µg, about 85 µg, about 90 µg, about 95 µg, or about 100 µg. In some embodiments, the daily dose is about 50 µg.

[0111] In some embodiments, the anti-CD3 is administered as a daily dose. In some embodiments, the anti-CD3 is administered orally. In some embodiments, the daily dose administered orally is from about 1.0 mg to about 2.5 mg. In some embodiments, the daily dose administered orally is about 0.5 mg, about 0.75 mg, about 1.0 mg, about 1.25 mg, about 1.5 mg, about 1.75 mg, about 2.0 mg, about 2.25 mg, about 2.5 mg, about 2.75 mg, about 3.0 mg, about 3.25 mg, about 3.5 mg, about 3.75 mg, about 4.0 mg, about 4.25 mg, about 4.5 mg, about 4.75 mg, or about 5.0 mg.

[0112] In some embodiments, the anti-CD3 is administered for at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 11 days, at least 12 days, at least 13 days, at least 14 days, at least 15 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, at least 20 days, at least 21 days, at least 22 days, at least 23 days, at least 24 days, at least 25 days, at least 26 days, at least 27 days, at least 28 days, at least 29 days, or at least 30 days. In some embodiments, the anti-CD3 is administered for at least 10 consecutive days.

[0113] The anti-CD3 can be administered by nasal drops, by nasal inhalation, by oral inhalation, intravenously, orally, any combination thereof, or any other route of administration described herein. Alternatively, the active compound is administered orally via enteric-coated capsules. Administration by inhalation can be in the form of an inhaler or an atomizer. The atomizer and / or inhaler are hand-held. Optionally, the atomizer and / or inhaler can have different sizes to fit children and / or adults.

[0114] The effectiveness of the treatment is determined in conjunction with any known method for diagnosing or treating a disease or disorder described herein. Alleviation of one or more symptoms of the disease or disorder indicates that the antibody provides a clinical benefit.

[0115] Co-administration of anti-CD3 and other agents

[0116] The present disclosure provides a method of treating, preventing, or alleviating symptoms of a disease or disorder in a subject in need thereof, comprising co-administering to the subject a composition comprising a CD3 antibody (anti-CD3) and another active agent. The active agent can be any drug or supplement used to treat the disease or disorder. The disease or disorder can be any disease or disorder provided herein. In some embodiments, the disease or disorder is a coronavirus infection, an inflammatory disease, acute respiratory distress syndrome (ARDS), a neurodegenerative or a pulmonary disease in the subject. The co-administration of the anti-CD3 and the other active agent can be in any order. For example, the anti-CD3 can be administered before, simultaneously with, or after the other active agent. The co-administration occurs at any time during a treatment period in which the subject in need thereof receives treatment for the disease. The co-administration of the anti-CD3 and the other active agent can be on the same day, on different days, on the same week, or on different weeks.

[0117] In some embodiments, the methods of treating, preventing, or alleviating symptoms provided herein comprise co-administering an anti-CD3 antibody with another therapeutic agent. In some embodiments, the anti-CD3 antibody is co-administered with dexamethasone. In some embodiments, the anti-CD3 antibody of the present disclosure is administered before, simultaneously with, or after the administration of dexamethasone. When administered simultaneously, the anti-CD3 antibody and dexamethasone can be formulated together or separately and administered as described herein.

[0118] In some embodiments, the methods of treatment, prevention, or alleviation of symptoms provided herein comprise co-administration of an anti-CD3 antibody with a monoclonal antibody. In some embodiments, the monoclonal antibody is an anti-IL-6R antibody. In some embodiments, the antibody is tocilizumab. In some embodiments, the IL-6R antibody comprises a VH CDR1 region comprising the amino acid sequence of SEQ ID NO: 15, a VH CDR2 region comprising the amino acid sequence of SEQ ID NO: 37, a VH CDR3 region comprising the amino acid sequence of SEQ ID NO: 35, a VL CDR1 region comprising the amino acid sequence of SEQ ID NO: 24, a VL CDR2 region comprising the amino acid sequence of SEQ ID NO: 25, and a VL CDR3 region comprising the amino acid sequence of SEQ ID NO: 26. In some embodiments, the methods of treatment, prevention, or alleviation of symptoms provided herein comprise co-administration of intranasally delivered anti-CD3 and anti-IL6R antibodies. In some embodiments, the anti-IL6R antibody is delivered intravenously. In some embodiments, the anti-IL6R antibody is delivered by inhalation. In some embodiments, the anti-IL6R antibody is delivered orally. In some embodiments, the anti-IL6R antibody is delivered subcutaneously. In some embodiments, the co-administration of anti-IL-6R mAb and anti-CD3 produces a synergistic effect that reduces symptoms associated with a disease or condition described herein, such as a coronavirus disease (e.g., COVID-19, SARS, or MERS).

[0119] In some embodiments, the methods of treatment, prevention, or alleviation of symptoms provided herein comprise co-administration of an anti-CD3 antibody and an anti-TNFa antibody, an anti-CD20 antibody, an anti-IFNg antibody, an anti- granulocyte-macrophage colony-stimulating factor antibody, an anti-IL6R antibody, or a combination thereof. In some embodiments, the anti-CD3 is co-administered with casirivimab and imdevimab. In some embodiments, the anti-CD3 is co-administered with bamlanivimab and etesevimab. In some embodiments, the anti-CD3 is co-administered with sotrovimab.

[0120] In some embodiments, the methods of treating, preventing, or reducing symptoms provided herein comprise co-administration of an anti-CD3 antibody and an antiviral agent. In some embodiments, the antiviral agent is azidothymidine (also known as zidovudine or AZT), remdesivir, or dactinomycin. In some embodiments, the methods of treating, preventing, or reducing symptoms provided herein comprise co-administration of an anti-CD3 antibody and an immune-enhancing drug, vitamin C, vitamin D, vitamin E, or any combination thereof. In some embodiments, the methods of treating, preventing, or reducing symptoms provided herein comprise co-administration of an anti-CD3 antibody and an anticoagulant (i.e., a blood-thinning agent). In some embodiments, the anticoagulant is low-dose heparin or enoxaparin.

[0121] In some embodiments, the methods of treating, preventing, or reducing symptoms provided herein comprise co-administration of an anti-CD3 antibody and a corticosteroid. In some embodiments, the corticosteroid is dexamethasone, prednisone, or methylprednisolone. In some embodiments, the corticosteroid is dexamethasone. In some embodiments, dexamethasone is administered orally as a unit dose. In some embodiments, dexamethasone is administered once per week, twice per week, three times per week, or four times per week during treatment. In some embodiments, dexamethasone is administered orally as a unit dose, wherein the unit dose is about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, or about 10 mg. In some embodiments, dexamethasone is administered orally as a unit dose, wherein the unit dose is about 6 mg.

[0122] In some embodiments, the methods of treating, preventing, or reducing symptoms provided herein comprise co-administration of an anti-CD3 antibody and convalescent plasma. In some embodiments, the convalescent plasma is derived from a subject who has recovered from a coronavirus.

[0123] Coronavirus infection

[0124] The present disclosure provides methods of treating, preventing, or reducing symptoms of a coronavirus infection in a subject in need thereof, comprising administering to the subject a composition comprising an anti-CD3 antibody.

[0125] In some embodiments, the subject has a disease or abnormal state associated with a coronavirus infection. For example, the coronavirus can be SARS-CoV (i.e., SARS), SARS-CoV-2 (i.e., COVID-19), MERS-CoV (i.e., MERS), or mutants and / or variants thereof. In some embodiments, the subject has a disease or abnormal state associated with MERS and / or variants thereof. In some embodiments, the subject has a disease or abnormal state associated with SARS and / or variants thereof. In some embodiments, the subject has a disease or abnormal state associated with SARS-CoV-2 and / or variants thereof. By “variant” is meant a genetic variant of a coronavirus, such that new genetic mutations occur in the variant relative to one or more known strains of the coronavirus. The mutations (e.g., substitutions or deletions) can be to any nucleotide in the coronavirus genome. The variant can be a variant of interest, a variant of concern, or a variant of severe consequence. For example, B.1.1.7 (alpha), B.1.351 (beta), B.1.617 (delta), and P.1 (gamma), B.1.526 (iota), B.1.427 (epsilon), B.1.429 (epsilon), B.1.1.7 (alpha), P.2 (zeta), and their lineages are currently classified as variants of SARS-CoV-2. It is to be understood that new variants of the coronavirus with new mutations or sets of mutations can arise, and these are also covered by the term “coronavirus” as described herein.

[0126] The methods provided herein comprise administering anti-CD3 as a dose or unit dose. The dose should be sufficient to result in a reduction of symptoms associated with a coronavirus infection (e.g., an overactive immune response or cytokine storm) or a slowing of replication of the coronavirus within the host, and preferably also prevent or reduce symptoms of a coronavirus-associated disease (e.g., COVID-19, SARS, or MERS). The dose selected should be sufficient to constitute effective treatment, but not so high as to cause unacceptable side effects (e.g., mucositis or anaphylactic shock). Preferably, the status of the disease condition (e.g., SERS, MERS, or COVID-19) and the health of the patient should be closely monitored during and for a reasonable period of time after treatment.

[0127] In some embodiments, the anti-CD3 is administered as a daily dose. In some embodiments, the anti-CD3 is administered nasally. In some embodiments, the daily dose administered nasally is from 50 μg to 100 μg. In some embodiments, the daily dose is about 20 μg, about 25 μg, about 30 μg, about 35 μg, about 40 μg, about 45 μg, about 50 μg, about 55 μg, about 60 μg, about 65 μg, about 70 μg, about 75 μg, about 80 μg, about 85 μg, about 90 μg, about 95 μg, or about 100 μg. In some embodiments, the daily dose is about 50 μg.

[0128] In some embodiments, the anti-CD3 is administered as a daily dose. In some embodiments, the anti-CD3 is administered orally. In some embodiments, the daily dose administered orally is from about 1.0 mg to about 2.5 mg. In some embodiments, the daily dose administered orally is about 0.5 mg, about 0.75 mg, about 1.0 mg, about 1.25 mg, about 1.5 mg, about 1.75 mg, about 2.0 mg, about 2.25 mg, about 2.5 mg, about 2.75 mg, about 3.0 mg, about 3.25 mg, about 3.5 mg, about 3.75 mg, about 4.0 mg, about 4.25 mg, about 4.5 mg, about 4.75 mg, or about 5.0 mg.

[0129] In some embodiments, the anti-CD3 is administered for at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 11 days, at least 12 days, at least 13 days, at least 14 days, at least 15 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, at least 20 days, at least 21 days, at least 22 days, at least 23 days, at least 24 days, at least 25 days, at least 26 days, at least 27 days, at least 28 days, at least 29 days, or at least 30 days. In some embodiments, the anti-CD3 is administered for at least 10 consecutive days.

[0130] In some embodiments, the anti-CD3 antibody is co-administered with a corticosteroid. In some embodiments, the corticosteroid is dexamethasone, prednisone, or methylprednisolone. In some embodiments, the corticosteroid is dexamethasone. In some embodiments, the dexamethasone is administered orally as a unit dose. In some embodiments, the dexamethasone is administered once per week, twice per week, three times per week, or four times per week during the treatment period. In some embodiments, the dexamethasone is administered orally. In some embodiments, the dexamethasone is administered nasally. In some embodiments, the dexamethasone is administered as a unit dose, wherein the unit dose is about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, or about 10 mg. In some embodiments, the dexamethasone is administered orally as a unit dose, wherein the unit dose is about 6 mg.

[0131] In some embodiments, the anti-CD3 antibody is co-administered with an antiviral agent. In some embodiments, the antiviral agent is zidovudine. In some embodiments, the zidovudine is administered orally as a unit dose. In some embodiments, the unit dose is about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg.

[0132] In some embodiments, the subject is a human subject. In some embodiments, the subject has or is suspected of having a coronavirus infection. In some embodiments, the subject has been or is believed to have been exposed to a coronavirus and has not yet developed symptoms of a coronavirus infection. In some embodiments, the coronavirus is, for example, the virus that causes COVID-19, SARS, or MERS. In some embodiments, a sample from the subject is tested for active coronavirus, evidence of coronavirus exposure, or evidence of coronavirus immunity. The sample from the subject can be, for example, a blood sample, a serum or plasma sample, a lavage sample, a urine sample, or any sample that can be used to determine an overactive immune response. In some embodiments, the subject has tested positive in a coronavirus diagnostic test. In some embodiments, the subject has an active coronavirus infection. In some embodiments, the diagnostic test measures the presence of a coronavirus genome. In some embodiments, the diagnostic test is a quantitative polymerase chain reaction test. In some embodiments, the diagnostic test measures the presence of an antibody that binds to a coronavirus epitope. In some embodiments, the coronavirus epitope is on a coronavirus spike protein.

[0133] Signs and symptoms of a coronavirus infection can be determined by a medical professional or self-reported in a symptomology survey as a patient reported outcome survey. In some embodiments, the symptom is one or more of fever, gastrointestinal symptoms, respiratory symptoms, loss of smell (anosmia), taste disorder (ageusia), cough, headache, sore throat, pain when swallowing, shortness of breath, difficulty breathing, rapid breathing, nausea, vomiting, decreased O2 saturation, diarrhea, runny nose, abdominal pain, myalgia, fever, conjunctivitis, and loss of appetite.

[0134] A coronavirus infection can cause an overactive immune response as a symptom, sometimes referred to as a cytokine storm, hyperinflammatory response, and the like. An overactive immune response is characterized by a rapid increase and elevated levels (i.e., high levels) of proinflammatory molecules, such as cytokines and chemokines. The rapid increase and high levels of proinflammatory molecules can include interleukin 6 (IL-6), C-reactive protein, D-dimer, interferon (IFN), interferon alpha (IFN-a), interferon gamma (IFN-g), interleukin 1 beta (IL-1b), and / or CXCL10. Without being bound by theory, it is believed that an overactive immune response leads to severe and fatal cases of coronavirus infection.

[0135] In some embodiments, the symptom is an overactive immune response. In some embodiments, the overactive immune response comprises an elevated level of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), D-dimer, interferon (IFN), interferon alpha (IFN-a), interferon gamma (IFN-g), interleukin 1 beta (IL-1b), and / or CXCL10. In some embodiments, the overactive immune response comprises an elevated level of at least IL-6 and CRP. In some embodiments, the overactive immune response comprises an elevated level of at least IL-6. In some embodiments, the overactive immune response comprises an elevated level of at least CRP.

[0136] An overactive immune response can be measured by determining the level of proinflammatory molecules, such as cytokines and chemokines, in a sample collected from a subject. The sample from the subject can be, for example, a blood sample, a serum or plasma sample, a lavage sample, a urine sample, or any sample that can be used to determine an overactive immune response. Any method known in the art for measuring cytokines and chemokines in a sample collected from a subject can be used. For example, the method can be, but is not limited to, flow cytometry microsphere array, quantitative polymerase chain reaction, immunoassay (e.g., ELISA or immunoturbidimetry assay), electrochemiluminescence immunoassay, and the like. Commercially available laboratory developed tests and diagnostic kits can be used to determine the level of cytokines or chemokines. General guidelines available to one of skill in the art can be used to determine whether the level of a cytokine or chemokine (e.g., IL-6 or CRP) is low, normal, or elevated (i.e., high). It will be appreciated that the data provided by the method used to determine the level of one or more cytokines or chemokines is interpreted by the skilled person (e.g., a medical practitioner) in the context of the particular patient's condition.

[0137] In some embodiments, the methods of treating, preventing, or reducing symptoms provided herein comprise measuring a symptom. In some embodiments, the symptom is an overactive immune response. In some embodiments, measuring an overactive immune response comprises determining the level of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), D-dimer, interferon (IFN), interferon alpha (IFN-alpha), interferon gamma (IFN-gamma), interleukin 1 beta (IL-1 beta), or CXCL10. In some embodiments, measuring an overactive immune response comprises determining the level of at least one of IL-6, CRP, and D-dimer. In some embodiments, measuring an overactive immune response comprises determining the level of at least one of IL-6 and CRP. In some embodiments, measuring an overactive immune response comprises determining the level of at least IL-6. In some embodiments, measuring an overactive immune response comprises determining the level of at least CRP.

[0138] In some embodiments, the method of treating, preventing, or reducing symptoms comprises administering to the subject an anti-CD3 antibody based on an overactive immune response in the subject. In some embodiments, the anti-CD3 antibody is administered to the subject if the overactive immune response comprises a high level of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), D-dimer, interferon (IFN), interferon alpha (IFN-alpha), interferon gamma (IFN-gamma), interleukin 1 beta (IL-1 beta), or CXCL10. In some embodiments, the anti-CD3 antibody is administered to the subject if the overactive immune response comprises a high level of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), or D-dimer. In some embodiments, the anti-CD3 antibody is administered to the subject if the overactive immune response comprises a high level of at least one of interleukin 6 (IL-6) and C-reactive protein (CRP). In some embodiments, the anti-CD3 antibody is administered to the subject if the overactive immune response comprises a high level of at least interleukin 6 (IL-6). In some embodiments, the anti-CD3 antibody is administered to the subject if the overactive immune response comprises a high level of at least C-reactive protein (CRP).

[0139] Other diseases and disorders

[0140] Provided herein are methods of treating, preventing, or reducing symptoms of a disease or disorder in a subject in need thereof, comprising administering to the subject a composition comprising a CD3 antibody (anti-CD3). In some embodiments, the subject has an inflammatory disease or disorder. In some embodiments, the inflammatory disease is autoimmune encephalomyelitis. In some embodiments, the inflammatory disease is lupus. In some embodiments, the inflammatory disease is arthritis. In some embodiments, the subject has acute respiratory distress syndrome (ARDS). In some embodiments, the subject has a disease or abnormal state associated with a neurodegenerative disease or disorder. In some embodiments, the neurodegenerative disorder is multiple sclerosis. In some embodiments, the neurodegenerative disorder is secondary progressive multiple sclerosis. In some embodiments, the subject has a disease or abnormal state associated with a pulmonary disease or disorder. Examples of pulmonary inflammatory diseases include ARDS (acute respiratory distress syndrome) and systemic pulmonary sclerosis.

[0141] Pharmaceutical compositions

[0142] The anti-CD3 antibodies described herein can be incorporated into pharmaceutical compositions suitable for oral or mucosal administration, e.g., by ingestion, inhalation, or absorption, for example, via nasal, intranasal, pulmonary, buccal, sublingual, rectal, or vaginal administration. Such compositions can include inert diluents or an edible carrier. For purposes of oral therapeutic administration, the active compound (e.g., anti-CD3 antibody) can be mixed with excipients and used in the form of tablets, capsules, and the like. Oral anti-CD3 antibody compositions can also be prepared using a fluid carrier, such as an oral syrup or elixir. Pharmaceutically compatible binders and / or adjuvants can be included as part of the composition. Provided are oral dosage forms comprising an anti-CD3 antibody, wherein the dosage form provides therapeutically effective blood levels of the anti-CD3 antibody to a subject following oral administration. Also provided are mucosal dosage forms comprising an anti-CD3 antibody, wherein the dosage form provides therapeutically effective blood levels of the anti-CD3 antibody to a subject following mucosal administration. For purposes of mucosal therapeutic administration, the active compound (e.g., anti-CD3 antibody) can be mixed with excipients or carriers suitable for administration by inhalation or absorption, e.g., via nasal sprays or drops, or rectal or vaginal suppositories.

[0143] Solid oral dosage forms include, but are not limited to, tablets (e.g., chewable tablets), capsules, caplets, powders, pellets, granules, powder in a sachet, enteric-coated tablets, enteric-coated beads, and enteric-coated soft capsules. Also included are multi-layered tablets, where different layers can contain different drugs. Solid dosage forms also include encapsulated powders, pellets, and granules. Powders, pellets, and granules can be coated, e.g., with suitable polymers or conventional coating materials to achieve, e.g., greater stability in the gastrointestinal tract, or to achieve a desired release rate. In addition, capsules containing powders, pellets, or granules can be further coated. Tablets or caplets can be scored to facilitate division, to allow for adjustment of dosage as needed. Dosage forms of the present application can be unit dosage forms, where the dosage form is intended to deliver one therapeutic dose per administration, e.g., one tablet equals one dose. Such dosage forms can be prepared by pharmaceutical methods well known to those skilled in the art (see Remington's Pharmaceutical Sciences, 18th Ed., Mack Publishing, Easton Pa. (1990)).

[0144] Typical oral dosage forms can be prepared by combining the active ingredient(s) in an intimate admixture with at least one excipient according to conventional pharmaceutical compounding techniques. Excipients can take a wide variety of forms depending on the form of preparation desired. For example, excipients suitable for use in solid oral dosage forms (e.g., powders, tablets, capsules, and caplets) include, but are not limited to, starches, sugars, microcrystalline cellulose, diluents, granulating agents, lubricants, binders, and disintegrating agents. Examples of excipients suitable for use in oral liquid dosage forms include, but are not limited to, water, glycols, oils, alcohols, flavoring agents, preservatives, and coloring agents.

[0145] Tablets and capsules represent convenient pharmaceutical compositions and oral dosage forms, in which solid excipients are employed. If desired, tablets can be coated by standard aqueous or nonaqueous techniques. Such dosage forms can be prepared by any of the methods of pharmaceutical formulation. Typically, a pharmaceutical composition and dosage form is prepared by uniformly and intimately mixing, or bringing into association, the active ingredient with liquid carriers, finely divided solid carriers, or both, and then, if necessary, shaping the product into the desired presentation.

[0146] As an example, tablets can be prepared by compression or molding. Compressed tablets can be prepared by either (1) directly compressing the active ingredient(s) in a free-flowing form (e.g., a powder or granules), optionally with an excipient, in a suitable machine, or (2) preparing a powder or granular mixture incorporating the active ingredient(s) and then compacting the mixture in a suitable machine. Molded tablets can be made by molding a mixture of the powdered active ingredient(s) moistened with a liquid diluent.

[0147] Excipients which can be used in oral dosage forms of the present application include, but are not limited to, binders, fillers, disintegrants, and lubricants. Binders suitable for use in pharmaceutical compositions and dosage forms include, but are not limited to, corn starch or other starches, gelatin, natural and synthetic gums such as acacia, sodium alginate, alginic acid, other alginates, powdered tragacanth, guar, cellulose and its derivatives (e.g., ethyl cellulose, cellul acetate, carboxymethyl cellulose calcium, sodium carboxymethyl cellulose), polyvinyl pyrrolidine, methyl cellulose, pregelatinized starch, hydroxypropyl methyl cellulose (e.g., Nos. 2208, 2906, 2910), microcrystalline cellulose, and mixtures thereof.

[0148] Microcrystalline cellulose in a suitable form includes, but is not limited to, materials sold as AVICEL™ PH-101, AVICEL™ PH-103, AVICEL™ RC-581, AVICEL™ PH-105 (available from FMC Corporation, American Viscose Division, Avicel Sales, Marcus Hook, Pa.) and mixtures thereof. A particular binder is a mixture of microcrystalline cellulose and sodium carboxymethyl cellulose, which is sold as AVICEL™ RC-581. Suitable anhydrous or low moisture excipients or additives include AVICEL™ PH-103 and Starch 1500™ LM.

[0149] Examples of fillers suitable for use in the pharmaceutical compositions and dosage forms disclosed herein include, but are not limited to, talc, calcium carbonate (e.g., granular or powdered), microcrystalline cellulose, powdered cellulose, dextrates, kaolin, mannitol, silicic acid, sorbitol, starch, pre-gelatinized starch, and mixtures thereof. Binders or fillers in the pharmaceutical compositions and dosage forms of the application are typically present with about 50 to about 99 weight % of the pharmaceutical composition or dosage form.

[0150] Disintegrants can be used in the pharmaceutical compositions and oral or mucosal dosage forms of the application to provide tablets that disintegrate when exposed to an aqueous environment. Tablets containing too much disintegrant can disintegrate upon storage, while tablets containing too little disintegrant can not disintegrate at the desired rate or under the desired conditions. As such, a sufficient amount of disintegrant to neither too much nor too little to adversely affect the release of the active ingredient should be used to form the pharmaceutical compositions and solid oral dosage forms described herein. The amount of disintegrant used varies based on the type of formulation and is readily discernible to one of ordinary skill in the art. Typically, the pharmaceutical compositions and dosage forms contain about 0.5 to about 15 weight % of a disintegrant, preferably about 1 to about 5 weight % of a disintegrant.

[0151] Disintegrants that can be used in the pharmaceutical compositions and oral or mucosal dosage forms of the application include, but are not limited to, agar, alginic acid, calcium carbonate, Primogel, microcrystalline cellulose, sodium croscarmellose, cross-linked crospovidone, polacrilin potassium, sodium starch glycolate, corn, potato or tapioca starch, other starches, pre-gelatinized starch, other starches, clays, other alginates, other celluloses, gums, and mixtures thereof.

[0152] Lubricants which can be used in the pharmaceutical compositions and dosage forms of the application include, but are not limited to, calcium stearate, magnesium stearate, or Sterotes, mineral oil, light mineral oil, glycerin, sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid, sodium lauryl sulfate, talc, hydrogenated vegetable oil (e.g., peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate, ethyl laureate, agar, and mixtures thereof. Additional lubricants include, for example, syloid silica aerogel (AEROSIL™ 200, manufactured by W.R. Grace Co. of Baltimore, Md.), an agglomerate of synthetic silica (sold by the Degussa Co. of Plano, Tex.), CAB-O-SIL™ (a pyrogenic silica product sold by the Cabot Co. of Boston, Mass.), and mixtures thereof. If used at all, lubricants are typically used in an amount of less than about 1 weight % of the pharmaceutical compositions or dosage forms in which they are incorporated. Glidants, such as colloidal silicon dioxide, can also be used.

[0153] The pharmaceutical compositions and oral or mucosal dosage forms can further comprise one or more compounds that reduce the rate by which the active ingredient decomposes. Thus, the oral dosage forms described herein can be processed to be immediate release dosage forms or sustained release dosage forms. Immediate release dosage forms can release the anti-CD3 antibody within a relatively short period of time, e.g., within a few minutes to a few hours. Sustained release dosage forms can release the anti-CD3 antibody over a period of several hours, e.g., up to 24 hours or more if desired. In either case, during delivery, the delivery can be controlled to proceed at substantially a certain predetermined rate. In some embodiments, the solid oral dosage forms can be coated with a polymeric coating material or other known coating material to achieve, e.g., greater stability on the shelf or in the gastrointestinal tract, or to achieve control over the release of the drug. Such coating techniques and the materials used therein are known in the art. Such compounds, referred to herein as "stabilizers," include, but are not limited to, antioxidants such as ascorbic acid and salt buffers. Cellulose acetate phthalate, polyvinyl acetate phthalate, hydroxypropyl methyl cellulose phthalate, methacrylic acid-methacrylic acid ester copolymers, cellulose acetate trimellitate, carboxymethyl ethyl cellulose, and hydroxypropyl methyl cellulose acetate succinate, among others, are particularly useful to achieve enteric coatings. Mixtures of waxes, shellac, zein, ethyl cellulose, acrylic resins, cellulose acetate, silicone elastomers can be used to achieve sustained release coatings. See, e.g., Remington, supra, Chapter 93, for other types of coatings, techniques and apparatus.

[0154] Liquid for oral or mucosal administration represents another convenient dosage form, in which case a solvent can be used. In some embodiments, the solvent is a buffered liquid, such as phosphate buffered saline (PBS). Liquid oral dosage forms can be prepared by mixing the active ingredient in a suitable solvent to form a solution, suspension, syrup, or elixir of the active ingredient in the liquid. The solution, suspension, syrup, and elixir can optionally include other additives, including but not limited to glycerin, sorbitol, propylene glycol, a sugar or other sweetener, a flavoring agent, and a stabilizer. Flavoring agents can include, but are not limited to, peppermint, methyl salicylate, or orange flavoring. Sweeteners can include sugar, aspartame, saccharin, sodium cyclamate, and xylitol.

[0155] To reduce the extent of inactivation of an anti-CD3 antibody administered orally in the stomach of a treated subject, an antacid can be administered concurrently with the immunoglobulin, which neutralizes the otherwise acidic nature of the intestine. Thus, in some embodiments, an anti-CD3 antibody is administered orally with an antacid (e.g., aluminum or magnesium hydroxide, such as MAALOX™ antacid or MYLANTA™ antacid), or an H2 blocker (such as cimetidine or ranitidine). One skilled in the art will appreciate that the dosage of antacid administered in conjunction with an anti-CD3 antibody depends on the particular antacid used. When the antacid is MYLANTA™ antacid in liquid form, between 15 ml and 30 ml, e.g., about 15 ml, can be administered. When cimetidine H2 blocker is used, between about 400 and 800 mg per day can be used.

[0156] The kits described herein can include an anti-CD3 antibody composition as a prepared liquid oral or mucosal dosage form ready for administration, or alternatively, can include an anti-CD3 antibody composition as a solid pharmaceutical composition that can be reconstituted with a solvent to provide a liquid oral or mucosal dosage form. When the kit includes an anti-CD3 antibody composition as a solid pharmaceutical composition that can be reconstituted with a solvent to provide a liquid dosage form (e.g., for oral or nasal administration), the kit can optionally include the reconstitution solvent. In this case, the solvent or reconstitution solvent is mixed with the active ingredient to provide a liquid oral dosage form of the active ingredient. Typically, the active ingredient is soluble in the solvent and forms a solution. The solvent can be, for example, water, a non-aqueous liquid, or a combination of a non-aqueous component and an aqueous component. Suitable non-aqueous components include, but are not limited to, an oil; an alcohol, such as ethanol; glycerol; and a glycol, such as polyethylene glycol and propylene glycol. In some embodiments, the solvent is phosphate buffered saline (PBS).

[0157] For administration via inhalation, the mucosal anti-CD3 antibody compound can be delivered in the form of an aerosol spray from pressurized containers or nebulizers, or from a bomb, with suitable propellants, e.g., gases such as carbon dioxide. Such methods include those described in U.S. Patent No. 6,468,798.

[0158] Systemic administration can also be transmucosal. For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives. Transmucosal administration can be accomplished through the use of nasal sprays or suppositories, or rectal or vaginal delivery.

[0159] The anti-CD3 antibody compound can also be prepared in a form suitable for rectal administration such as with standard rectal suppository formulations, or retention enemas.

[0160] In one embodiment, oral or mucosal anti-CD3 antibody compositions are prepared with carriers that protect the anti-CD3 antibody against rapid elimination from the body, such as those designed to slowly release a drug into the body. Biodegradable, biocompatible polymers can be used, such as ethylene vinyl acetate, poly anhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. These can be used to prepare compositions that can be administered in standard fashion. Materials also can be obtained commercially, e.g., from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensions (including liposomes targeted to infected cells and monoclonal antibody-coated liposomes) can also be used as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in U.S. Patent No. 4,522,811.

[0161] The dosage, toxicity and therapeutic efficacy of such anti-CD3 antibody compositions can be determined by standard pharmaceutical procedures in cell cultures, or experimental animals, e.g., cells harvested from animals after mucosal administration of the anti-CD3 antibody. For example, for determining LD50 (the dose lethal to 50% of the population) and ED50 (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD50 / ED50. Compositions that exhibit large therapeutic indices are preferred. While anti-CD3 antibody compositions that exhibit toxic side effects can be used, care should be taken to design a delivery system that targets such compounds to the site of affected tissue in order to minimize potential damage to uninfected cells and, thereby, reduce side effects.

[0162] Data obtained from cell culture and animal studies (e.g., cells taken from animals after mucosal administration of an anti-CD3 antibody) can be used in formulating a range of dosage for use in humans. The dosage of an anti-CD3 antibody composition lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage can vary within this range depending upon the dosage form employed and the route of administration utilized. For any oral or mucosal anti-CD3 antibody composition used in the methods described herein, the therapeutically effective dose can be estimated initially from cell culture assays (e.g., cells taken from animals after mucosal administration of an anti-CD3 antibody). A dose can be formulated in an animal model to achieve a desired IL-10 or TGFβ, or modulatory cell, circulating plasma concentration that is within the range that includes the IC50 (i.e., the concentration of the test compound that achieves a half-maximal inhibition of symptoms) as determined in cell culture. Such information can be used to more accurately determine useful doses in humans. Levels of IL-10 or TGFβ in plasma can be measured by methods known in the art, e.g., by ELISA. Levels of modulatory cells can be measured by methods known in the art, e.g., by flow cytometry-based methods.

[0163] A therapeutically effective amount (i.e., effective dose) of an anti-CD3 antibody, as defined herein, depends on the antibody chosen, the mode of delivery, and the condition to be treated. For example, a single dose amount in the range of about 1: g / kg to 1000 g / kg can be administered; in some embodiments, about 5, 10, 50, 100, or 500: g / kg can be administered. In some embodiments (e.g., pediatric subjects), about 1 to 100: g / kg (e.g., about 25 or 50: g / kg) of an anti-CD3 antibody can be administered. Anti-CD3 antibody compositions can be administered from once or more times per day to once or more times per week, including every other day. Oral or mucosal anti-CD3 antibody compositions can be administered, for example, for about 10 to 14 days or longer. Those of skill in the art will appreciate that certain factors can influence the dosage and timing required to effectively treat a subject, including but not limited to the severity of the disease or condition, previous treatments, the general health and / or age of the subject, and the

[0164] Oral or mucosal anti-CD3 antibody compositions can also include one or more therapeutic agents for treating a coronavirus infection. Such therapeutic agents can include, for example, NSAIDs (including COX-2 inhibitors); other antibodies, e.g., anti-cytokine antibodies (e.g., antibodies against IFN-a, IFNy, and / or TNFa); gold-containing compounds; immunosuppressive drugs (such as corticosteroids, e.g., dexamethasone, prednisolone, and methylprednisolone).

[0165] For example, a combination therapy can include one or more antibodies of the application co-formulated and / or co-administered with one or more additional therapeutic agents, such as a corticosteroid (e.g., dexamethasone, prednisolone, and methylprednisolone), an antiviral, an immunopotentiator, vitamin C, vitamin D, vitamin E. Advantageously, such combination therapies can use lower doses of the administered therapeutic agents, thereby avoiding possible toxicities or complications associated with various monotherapies.

[0166] The antiviral is azidothymidine, remdesivir, or dactinomycin.

[0167] In some embodiments, an anti-CD3 antibody is administered to a subject orally, nasally, or both orally and nasally.

[0168] In some embodiments, an anti-CD3 antibody is administered to a subject orally, nasally, or both orally and nasally.

[0169] In some embodiments, an anti-CD3 antibody is administered to a subject nasally, and a corticosteroid, such as dexamethasone, is delivered orally, such as, for example, by inhalation.

[0170] In some embodiments, an anti-CD3 antibody is administered to a subject nasally, and a corticosteroid, such as dexamethasone, is delivered orally, such as, for example, by inhalation.

[0171] The pharmaceutical composition can be included in a container, pack, or dispenser together with instructions for administration.

[0172] The method of treatment or prevention generally includes administering to the subject an oral or mucosal anti-CD-3 antibody composition sufficient to stimulate the mucosal immune system. In some embodiments, the method includes administering an oral or mucosal anti-CD3 antibody composition sufficient to increase IL-10 and / or TGF-β production by T cells (e.g., regulatory T cells) in the peripheral blood, for example, by about 100%, 200%, 300%, or more. In some embodiments, the method includes administering an oral anti-CD3 antibody composition sufficient to reduce T cell proliferation in the peripheral blood, for example, by about 20% (e.g., in some embodiments, by at least about 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or more).

[0173] In some embodiments, the method can include administering dexamethasone to the subject prior to, concurrently with, or after administering the oral or mucosal anti-CD3 composition.

[0174] Definitions

[0175] Unless otherwise defined, scientific and technical terms used in connection with the present disclosure shall have the meanings that are commonly understood by those of ordinary skill in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular. Generally, the nomenclature used herein and the technique described herein in connection with cell and tissue culture, molecular biology, and protein and oligonucleotide or polynucleotide chemistry and hybridization are those well-known and commonly used in the art. Standard techniques are used for recombinant DNA, oligonucleotide synthesis, and tissue culture and transformation (e.g., electroporation, lipofection). Enzymatic reactions and purification techniques are performed according to manufacturer’s specifications or as commonly accomplished in the art or as described herein. The foregoing techniques and procedures are generally performed according to conventional methods well-known in the art and as described in various general and more specific references that are cited and discussed throughout the present specification. See e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989)). The nomenclature used herein and the laboratory procedures in analytical chemistry, synthetic organic and medicinal chemistry, and pharmacology described herein are those well-known and commonly used in the art. Standard techniques are used for chemical syntheses, chemical analyses, formulation and delivery of pharmaceuticals, and treatment of patients.

[0176] As used in accordance with the present disclosure, the following terms are to be understood as having the following meanings, unless otherwise indicated:

[0177] As used herein, the term “antibody” refers to immunoglobulin molecules and immunologically active portions of immunoglobulin (Ig) molecules, i.e., molecules that contain an antigen-binding site that specifically binds (immunoreacts with) an antigen. Such antibodies include, but are not limited to, polyclonal, monoclonal, chimeric, single-chain, Fab, Fab’ and F(ab’)2 fragments, as well as Fab expression libraries. By “specifically binds” or “immunoreacts with” is meant that the antibody reacts with one or more antigenic determinants of the desired antigen and does not react with other polypeptides (i.e., binds) or binds other polypeptides with a much lower affinity (Kd> 10 -6 ) than the desired antigen.

[0178] The basic antibody structural unit is known to comprise a tetramer. Each tetramer is composed of two identical pairs of polypeptide chains, each pair having one "light" (about 25 kDa) and one "heavy" chain (about 50-70 kDa). The amino-terminal portion of each chain includes a variable region of about 100 to 110 or more amino acids primarily responsible for antigen recognition. The carboxy-terminal portion of each chain defines a constant region primarily responsible for effector function. Human light chains are classified as kappa and lambda light chains. Heavy chains are classified as mu, delta, gamma, alpha, or epsilon, and define a class of immunoglobulin, IgM, IgD, IgA, and IgE, respectively. Within light chains and heavy chains, the variable and constant regions are joined by a "J" region of about 12 or more amino acids, with the heavy chain also including a "D" region of about 10 more amino acids. See generally, Fundamental Immunology Ch. 7 (Paul, W. et al, 2nd ed., Raven Press, N.Y. (1989)). The variable regions of each light / heavy chain pair interact to form the antibody binding site.

[0179] As used herein, the term "monoclonal antibody" (MAb) or "monoclonal antibody composition" refers to a population of antibody molecules that contain only one species of antibody molecule consisting of a unique light chain gene product and a unique heavy chain gene product. In particular, the complementarity determining regions (CDRs) of a monoclonal antibody are identical in all the molecules of the population. MAbs contain an antigen binding site that is capable of immunoreacting with a specific epitope of an antigen, characterized by a unique binding affinity for that epitope.

[0180] Generally, antibody molecules obtained from humans relate to any of the classes IgG, IgM, IgA, IgE, and IgD, which differ from one another in the nature of the heavy chain present in the molecule. Certain classes have subclasses, such as IgGl, IgG2, etc. Furthermore, in humans, the light chain can be a kappa chain or a lambda chain.

[0181] As used herein, the term "epitope" includes any protein determinant capable of specific binding to an immunoglobulin, scFv, or T-cell receptor. The term "epitope" includes any protein determinant capable of specific binding to an immunoglobulin or T-cell receptor. Epitopic determinants usually consist of chemically active surface grouping of molecules, such as amino acids or sugar side chains, and usually have specific three-dimensional structural characteristics, as well as specific charge characteristics. An antibody specifically binds an antigen when the dissociation constant is < 1 μM, preferably < 100 nM, and most preferably < 10 nM.

[0182] As used herein, the terms "immunobind" and "immunobinding property" and "specific binding" refer to the type of noncovalent interaction that occurs between an immunoglobulin molecule and the antigen to which the immunoglobulin is specific. The strength or affinity of an immunobinding interaction can be expressed in terms of the dissociation constant (Kd) of the interaction, where a smaller Kd represents a greater affinity. The immunobinding properties of a selected polypeptide are quantified using methods well known in the art. One such method entails measuring the rates of antigen binding site / antigen complex formation and dissociation, where these rates depend on the concentrations of the complex partners, the affinity of the interaction, and geometric parameters that influence the rates equally in both directions. From this, both the "association rate constant" (Kon) and the "dissociation rate constant" (Koff) can be determined by calculating the concentrations and actual rates of association and dissociation (see Nature 361 : 186-87 (1993)). The ratio Koff / Kon enables all parameters independent of affinity to be eliminated, and equals the dissociation constant Kd (see generally Davies et al. (1990) Annual Rev Biochem 59:439-473). An antibody of the present disclosure can be said to specifically bind to a CD3 epitope when the equilibrium binding constant (Kd) is about 1 μΜ (preferably about 100 nM, more preferably about 10 nM, and most preferably about 100 pM to about 1 pM), as measured by an assay such as a radioligand binding assay or similar assay known to one of skill in the art.

[0183] Conservative amino acid substitutions refer to the interchangeability of residues having similar side chains. For example, one group of amino acids having aliphatic side chains is glycine, alanine, valine, leucine, and isoleucine; a second group of amino acids having aliphatic-hydroxyl side chains is serine and threonine; a third group of amino acids having amide-containing side chains is asparagine and glutamine; a fourth group of amino acids having aromatic side chains is phenylalanine, tyrosine, and tryptophan; a fifth group of amino acids having basic side chains is lysine, arginine, and histidine; and a sixth group of amino acids having sulfur-containing side chains is cysteine and methionine. Preferred conservative amino acid substitution groups are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine, alanine valine, glutamate-aspartate, and asparagine-glutamine.

[0184] As discussed herein, minor variations in the amino acid sequence of an antibody or immunoglobulin molecule are contemplated to be encompassed by the present disclosure, provided that the variations in the amino acid sequence maintain at least 75%, more preferably at least 80%, 90%, 95%, and most preferably 99%. In particular, conservative amino acid substitutions are contemplated. Conservative substitutions are those that take place within a family of amino acids that are related in their side chain. Genetically encoded amino acids are generally divided into the following families: (1) acidic amino acids are aspartic acid, glutamic acid; (2) basic amino acids are lysine, arginine, histidine; (3) non-polar amino acids are alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan, and (4) uncharged polar amino acids are glycine, asparagine, glutamine, cysteine, serine, threonine, tyrosine. Hydrophilic amino acids include arginine, asparagine, aspartic acid, glutamic acid, glutamine, histidine, lysine, serine, and threonine. Hydrophobic amino acids include alanine, cysteine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, and valine. Other families of amino acids include (i) serine and threonine, which are the aliphatic-hydroxyl family; (ii) asparagine and glutamine, which are the amide-containing family; (iii) alanine, valine, leucine and isoleucine, which are the aliphatic family; and (iv) phenylalanine, tryptophan, and tyrosine, which are the aromatic family.

[0185] The term "agent" is used herein to denote a chemical compound, a mixture of chemical compounds, a biological macromolecule, or an extract from a biological material.

[0186] The term patient includes human and veterinary subjects.

[0187] The present disclosure also includes Fv, Fab, Fab' and F(ab')2 anti-CD3 antibody fragments, single-chain anti-CD3 antibodies, bispecific anti-CD3 antibodies, heteroconjugate anti-CD3 antibodies, trispecific antibodies, immunoconjugates, and fragments thereof.

[0188] Bispecific antibodies are antibodies that have binding specificities for at least two different antigens. In the present case, one of the binding specificities is for CD3. The second binding target is any other antigen, and advantageously is a cell surface protein or receptor or receptor subunit.

[0189] All publications and patent documents cited in this document are hereby incorporated by reference as if each such publication or document was specifically and individually indicated to be incorporated by reference herein. Citation of the publications and patent documents is not intended as an admission that any is pertinent prior art, nor does it constitute any admission as to the contents or date of the same. Having now described the present disclosure in detail, the same will be understood to be one preferred embodiment of the application. Certain modifications of the above-described embodiments of the present application will be apparent to those skilled in the art, and this application is intended to cover and embrace such modifications, and all reasonable equivalents thereof. The foregoing description and examples have been set forth to illustrate the application and are not intended to limit the scope thereof. Numerous variations, changes and substitutions will now occur to those skilled in the art without departing from the application. It is therefore intended that the application not be limited, to the extent that the spirit and scope of the application are set forth in the following claims. Example

[0190] Example 1: Bioavailability study of foralumab administered by subcutaneous delivery in a mouse model

[0191] The objective of this study is to compare the pharmacokinetic (PK) profiles of foralumab administered intravenously and subcutaneously in a mouse model.

[0192] The results of this study will provide the feasibility of administering foralumab via subcutaneous injection, which represents a possible human treatment route. The intravenous administration route will be used as a comparator, as it was validated in previous preclinical repeat-dose toxicity studies and early clinical studies of foralumab administration.

[0193] Study Design

[0194] The mouse model used for this study is a human CD3 epsilon transgenic mouse model, which contains a humanized CD3 epsilon chain of the CD3 co-receptor within a functional mouse immune system. This model is used to determine the in vivo efficacy of human-specific immunotherapies targeting the human CD3 epsilon chain. A total of 132 mice (66 male and 66 female) were used. Groups were dosed intravenously (IV) or by subcutaneous injection (SC). Study groups 1, 2, and 3 contained 18 male and 18 female mice, divided into 3 groups per time point. Intravenous (IV) group 4 had 3 male and 3 female mice, while subcutaneous placebo group 5 had 9 male and 9 female mice. For each time point, 3 male and 3 female mice were used as described below (Table 1.1).

[0195] Table 1.1. Study groups

[0196]

[0197] Dose Formulations

[0198] Forelumab (NI-0401) was formulated in 25 mM sodium acetate buffer, 125 mM sodium chloride, and 0.02% polysorbate 80, pH 5.5. The vehicle control (placebo) used was 25 mM sodium acetate buffer, 125 mM sodium chloride, and 0.02% polysorbate 80, pH 5.5.

[0199] Dose Levels and Volumes

[0200] A single dose of 0.3 mg / kg was chosen, which was previously shown to cause up to 70% T cell reduction in the peripheral blood and 80% modulation of human CD3 epsilon molecules at the T cell membrane in LCD3 transgenic mice. The dose volume was 2.5 mL / kg, injected manually in a bolus.

[0201] Test Article Administration

[0202] A single dose of forelumab formulation or placebo was administered subcutaneously on the ventral abdominal wall or otherwise intravenously via the posterior orbital sinus.

[0203] Blood Sampling

[0204] Blood samples were collected from the mice by intracardiac puncture under terminal anaesthesia at the indicated time points after administration. The samples were collected in Plasma Separator Tubes (BD) and the plasma was separated by centrifugation. Aliquots of 40-50 μL plasma were frozen and stored at -50°C.

[0205] Results and Conclusions

[0206] Subcutaneous administration of forelumab resulted in efficient delivery to the blood and was pharmacologically active (Table 1.2). The blood levels of subcutaneously delivered forelumab peaked between 6 and 24 hours after delivery (Table 1.3). The bioavailability of forelumab delivered subcutaneously at 0.3 mg / kg was approximately 55% (Table 1.2). Increasing the dose of forelumab to 0.6 mg / kg in the subcutaneous delivery increased the bioavailability to 92% (Table 1.2). Since the Cmax achieved by subcutaneous administration is reduced by 50% compared to intravenous administration, infusion related reactions can be reduced. Together these results indicate that delivery of forelumab by subcutaneous administration is an efficient method of dosing forelumab and related antibodies to a subject.

[0207] Table 1.2. Pharmacokinetic parameters

[0208]

[0209] Table 1.3. Plasma concentrations from different routes of administration

[0210] PK Time (h) 0.3 mg / kg IV 1x 0.3 mg / kg SC 1x 0.6 mg / kg SC 2x 0 7306 0 0 0.5 7040 70 913 2 6283 950 3467 6 4660 2167 4183 24 3050 1733 5117 48 2033 1567 5000 120 1400 802 2683

[0211] Example 2: Pharmacokinetics, pharmacodynamics, and safety study of foralumab (NI-0401) administered intravenously

[0212] The primary objectives of these studies were to assess the safety and tolerability of foralumab in human subjects. The studies included assessment of the pharmacokinetic profile, immunogenicity, and pharmacodynamic effects of foralumab delivered over five days of intravenous administration.

[0213] Test Product, Dose, and Mode of Administration

[0214] Foralumab human monoclonal antibody was supplied in 3 mL vials, each containing 2 mL of foralumab formulation at a concentration of 2.0 mg / mL. Each vial contained 4.0 mg of foralumab. Foralumab doses were administered over 2 hours by intravenous infusion. Dosing regimens of foralumab were provided for eight different cohorts according to Table 2.1.

[0215] Table 2.1. Dosing regimens

[0216] Cohorts Day 1 (pg / m 2 Body surface area) Day 2 (pg / m 2 Body surface area) Day 3 (μg / m 2 Body surface area) Day 4 (ug / m 2 Body surface area) Day 5 (ug / m 2 Body surface area) Cohort 1 500 500 500 500 500 Cohort 2 500 500 650 650 650 Cohort 3 650 650 650 650 650 Cohort 4 650 650 1000 1000 1000 Cohort 5 1000 1000 1000 1000 1000 Cohort 6 1250 1250 1250 1250 1250 Cohort 7 1500 1500 1500 1500 1500 Cohort 8 1750 1750 1750 1750 1750

[0217] The pharmacokinetic profile was determined by measuring foralumab plasma levels at specified times after administration. Foralumab plasma levels were measured using a ligand binding assay. A specific anti-foralumab antibody was used as the capture reagent, and a fluorescently labeled anti-human IgGl. The sensitivity of the assay was 20 ng / mL (lower limit of detection).

[0218] Pharmacokinetic Results

[0219] Foralumab plasma levels were measured using a ligand binding assay. A specific anti-foralumab antibody was used as the capture reagent, and a fluorescently labeled anti-human IgGl. The sensitivity of the assay was 20 ng / mL (lower limit of detection).

[0220] For the study design in Table 2.1, the Cmax and AUCo-t parameters on study day 5 produced mean values of 584.4 ng / mL (range 28.43 to 1860.1 ng / mL) and 28570 ng.h / mL (range 1453 to 106494 ng.h / mL), respectively, over the dose range of 0.73 to 3.73 mg.

[0221] The following results show representative sampling of pharmacokinetic curves from three individuals from independent studies. These subjects were treated with five 1.0 mg doses (approximately + / - 500 μg / m 2 ), two 2.0 mg doses (approximately + / - 1000 μg / m 2 ), or a single 10.0 mg dose (approximately + / - 5000 μg / m 2 ). For these 3 subjects, the concentration curves were sufficient to estimate apparent Cmax (1 hour post-infusion) and AUCo-6, both of which showed a clear increase with dose. The apparent Cmax values for the 1.0 mg, 2.0 mg, and 10.0 mg doses were 110 ng / mL, 350 ng / mL, and 2800 ng / mL, respectively. The AUC values for the 1.0 mg, 2.0 mg, and 10.0 mg doses were 440 ng.h / mL, 1700 ng.h / mL, and 11800 ng.h / mL, respectively.

[0222] The following results present the foralumab PK data obtained following a single 2h infusion of 10.0 mg (subject 001-0001). Following a 2-hour intravenous infusion of 10.0 mg study drug, the plasma concentration of foralumab increased rapidly during the infusion period as expected. Following the infusion, the concentration values declined in a substantially mono-exponential manner. The early rapid decline in plasma concentration is due to the binding of the drug to its target on circulating T cells and the distribution of the drug. Following a single 10.0 mg foralumab dose, the estimated plasma terminal half-life was approximately 13 hours, although with more sensitive assays, the terminal half-life can be much longer (theoretically it would take approximately 78 h for elimination from the systemic circulation, which is 6 times the terminal half-life). The measured half-life of foralumab is shorter than expected for an IgGl molecule (typically approximately 3 weeks) due to the rapid uptake of the drug by the target cells.

[0223] Exposure based on AUCo-t on study days 1-5 indicates accumulation of foralumab in plasma. Plasma drug concentrations increased during the treatment period for one subject (030-0003) exposed to 5 doses of 1.0 mg foralumab, which corresponds to approximately 21 μg / kg / dose (+ / - 500 μg / m 2 / dose). When superimposed, this subject's PK and PD curves are correlated, both showing a peak at the end of the treatment period. This can be due to the effect of the frequency of dosing and / or depletion of the target on the disposition of the mAb decreasing.

[0224] In summary, these observations suggest that at 1.0 mg (i.e. 21 μg / kg or 500 μg / m 2The above doses of foralumab administered intravenously can result in drug accumulation over the 5-day dosing period. However, drug is expected to be rapidly eliminated and eliminated within approximately 3-4 days of the last dose.

[0225] Pharmacodynamic Results

[0226] For all pharmacodynamic analyses, foralumab doses were not distinguished in their expected pharmacology. All foralumab dose levels had an effect on TCR-CD3 complex and cell populations over the time course observed.

[0227] Modulation of TCR-CD3 complex was measured in CD8+ or CD4+ T cells starting at the designated time point after initiation of treatment. The maximum modulation of TCR-CD3 complex was observed in all treatment groups at the end of the treatment period on study day 5. The mean modulation in all treatment groups on study day 5 was 81.1% and the highest mean TCR-CD3 complex modulation at this point was observed in treatment cohort 8 (94%). Following the end of the treatment period, TCR-CD3 modulation gradually decreased. All patients in all treatment cohorts for which CD3 modulation data were available achieved CD3 modulation above 50%. CD3 modulation in all treatment groups remained above 50% for an average duration of 8.7 days and above 30% for an average duration of 12.9 days.

[0228] The preliminary kinetic profile of TCR-CD3 modulation for each cohort of 3 patients (doses from 500 to 1500 μg / m 2 The preliminary kinetic profile of TCR-CD3 modulation for each cohort of 3 patients (doses from 500 to 1500 μg / m

[0229] Circulating leukocyte counts and subpopulation counts were generated during and after the time course of foralumab administration. On study day 1, there was a transient increase in CD45+ leukocyte counts in all foralumab treatment cohorts (overall mean 68.7% increase) 6 hours after dosing, followed by a return to near or below baseline levels in most cohorts. By week 3, CD45+ leukocyte counts were below their baseline values in all cohorts except 1 and 4. In all treatment groups, CD45+ lymphocytes, CD3+ T cells, CD3+CD4+ T cells (helper T cells), and CD3+CD8+ T cells (cytotoxic T cells) rapidly and almost completely disappeared (>90%) from circulation within 24 h of the first infusion, followed by a return to near baseline values by week 3. These results strongly suggest that foralumab can induce lymphocyte depletion in humans. There was no apparent dose response in the reduction or recovery, although recovery was faster in cohort 1 than in all other cohorts except CD8+ cell counts. On study day 1, there was also a rapid decrease in CD3-CD19+ (B cells) and CD3-CD16+CD56+ cells (natural killer cells) counts in all treatment groups 6 hours after dosing, with different (non-dose dependent) recovery of these cell counts by study day 3, and higher than baseline values in week 3 in most cases.

[0230] Cytokine levels were assessed in subjects administered foralumab. Substantial variation between subjects in their cytokine release was observed following treatment with foralumab, which did not appear to be dose related. In patients with a significant increase in pro-inflammatory cytokines, this was accompanied by symptoms suggestive of an infusion-related reaction (IRR), although most symptoms were mild and of short duration. Most patients had little or no evidence of pro-inflammatory cytokine release on subsequent treatment days.

[0231] Safety Results

[0232] Adverse events (AEs) were assessed in human subjects administered foratumumab by intravenous administration. Nineteen of 24 patients (79%) experienced a total of 94 AEs, of which 3 (3%) were serious adverse events (SAEs). Fifty-eight AEs (62%) were of mild severity, 32 (34%) were of moderate severity, and 4 (4.3%) were severe. Sixty-eight of 91 non-serious AEs (75%) were considered by the investigator to have a reasonable likelihood of being drug-related. AEs occurring during the 5-day treatment period were predominantly defined as infusion-related reactions (IRRs) (61%) as they were reported during or within 24 hours after infusion of foratumumab. The most common IRRs were chills (9 events in 6 patients), fever (8 events in 7 patients), headache (8 events in 6 patients), hypotension (5 events in 3 patients), and ALT elevation (3 events in 3 patients).

[0233] One IRR was reported as a SAE: a transient ALT elevation in one patient in Cohort 5 on study Day 2 resulted in discontinuation of study drug treatment. Visual inspection indicated a clear dose response in the reporting of treatment-related AEs, with more drug-related AEs reported in the higher dose cohorts; all hematologic and lymphatic system disorders were reported by the two highest dose cohorts.

[0234] Three SAEs were reported; two were considered unrelated to study drug: one patient experienced a relapse of Crohn's disease 8 days after completion of the 5-day treatment, resulting in an extended hospitalization, while a second patient sustained a multi-segmental fracture of the left humerus with displacement of bone fragments after a fall on the street one and a half months after study treatment ended. One SAE was considered related to study drug: a transient elevation of ALT on study Day 2 resulted in discontinuation of study drug treatment. During the study, a small number of AEs (18) were reported after the 5-day treatment period. There were no deaths and no AEs that resulted in study discontinuation.

[0235] Hematologic and biochemical analyses of subjects receiving foratumumab indicated that, in general, mean hemoglobin, hematocrit, and red blood cell count values remained stable over time in all treatment cohorts, and there was no substantial change in mean platelet count over time across treatment cohorts. Mean total white blood cell count decreased from baseline to -2.98 10E9 / L in all treatment cohorts by study Day 5, with no evidence of a dose response. White blood cell counts recovered by Week 12. Mean neutrophil and monocyte counts showed the same pattern, with mean decreases by study Day 5 and recovery by Week 12 and Week 3, respectively.

[0236] Liver function was assessed in subjects receiving foralumab and some transient and non-serious liver test abnormalities were detected. Five patients (15%), in two cases starting at day 5 and in the other cases at weeks 2, 3, and 4, respectively, had isolated and transient elevations of ALP above the upper limit of the normal range. One patient had a pre-existing abnormal ALP level. One patient had a transient and isolated elevation of AST / ALT to 3.5 times the upper limit of the normal range at day 5, which normalized at week 2. Six patients (18%) had liver test abnormalities that described mild severity and transient duration of cholestatic liver injury. Except in one patient, there were no associated elevations of serum bilirubin. Of these six patients, three already had pre-existing liver test abnormalities of the same magnitude. Elevations of liver enzymes occurred mainly at day 5 and returned to baseline within one week. One patient had mild jaundice (bilirubin elevated at day 2 and resolved at day 4), while another had hepatomegaly (a second elevation of liver enzymes in this patient occurred again at week 4, who also had pre-existing liver test abnormalities). No other signs of liver insufficiency were noted in these cases, and no etiology was identified.

[0237] Conclusions

[0238] From the safety data it can be concluded that the dose-limiting toxicity dose was not reached in these studies. However, by steroid premedication, the safety profile of foralumab was extended from 500 mg / m 2 (~1.00 mg) to 1750 mg / m 2 (~3.5 mg) daily dose. The study has shown that foralumab is pharmacologically active. Foralumab has an effect on the TCR-CD3 complex and T cell subsets, which reflects the expected pharmacology of the drug and its target. After foralumab treatment, lymphocyte counts decreased below the normal range in all patients. The foralumab dose had no apparent effect on the duration of lymphopenia. By week 4, mean lymphocyte counts returned close to pre-treatment values. Lymphopenia is an expected effect of administration of anti-CD3 antibodies.

[0239] Example 3: Treatment of COVID-19 with an anti-CD3 antibody in human subjects

[0240] Study Design

[0241] Human subjects with flu-like symptoms consistent with COVID-19 infection were evaluated and screened for the study. Inclusion criteria included a positive RT-PCR COVID-19 test, and 39 subjects were enrolled.

[0242] Subjects were randomized into three groups: no fremanezumab treatment (n=16), intranasal fremanezumab and dexamethasone (n=11), and intranasal fremanezumab alone (n=12). 50 μg of fremanezumab was administered by nasal drops into each nostril (total of 100 ug). Subjects who received dexamethasone received 6 mg of oral dexamethasone on days 1-3. Subjects in the control group did not receive fremanezumab.

[0243] Laboratory Tests

[0244] Nasopharyngeal swabs were used to screen for COVID-19 by RT-PCR. Clinical laboratory tests included complete blood count, IL-6, D-dimer, CRP, COVID-19 serology, HIV, syphilis, pregnancy, hepatitis, and glycosylated hemoglobin. Leukocyte counts were measured by flow cytometry and impedance. C-reactive protein (CRP) and glycosylated hemoglobin were measured by nephelometry, D-dimer was measured by immunonephelometry, and IL-6 was measured by chemiluminescence.

[0245] Serum levels of IL-6, CRP, and D-dimer were quantified on days -2, 5, and 10. As shown in Figure 6A and Figure 6B Fremanezumab resulted in a 69% reduction in IL-6 levels at day 10 (p=0.031) and an 85% reduction in CRP at day 10 (p=0.032), as shown in Table 3.1. Comparison of the three groups showed differences between control vs. fremanezumab at day 5 (p=0.01) and day 10 (p=0.031) and CRP at day 10 (p=0.032), as shown in Table 3.1. These results indicate that treatment with fremanezumab can reduce hyperinflammation and adverse inflammatory responses associated with coronavirus infection.

[0246] Table 3.1 Levels of IL-6, CRP, and D-dimer

[0247]

[0248] Lung and CT Analysis

[0249] Pulmonary CT scans were performed using a 16-channel (Toshiba-Alexion) CT scanner. No contrast was used. The scan coverage was from the lung apex to the level of the bilateral adrenal glands. The tube voltage was between 100-120 Kv. The thin wall tissue slide thickness was 1 mm.

[0250] Pulmonary injury consisted of patchy opacities and ground-glass appearance, which were graded on a scale of 0 to 4 as follows: 0 = no detectable abnormalities or lung involvement < 5%; Stage 1 = mild lung involvement involving approximately 10% of the lung area, Stage 2 = moderate lung involvement involving approximately 25% of the lung area with patchy opacities and ground-glass lesions, Stage 3 = severe confluent ground-glass lesions and consolidation involving 25% to 50% of the lung area; Stage 4 involved more than half of the lung area with very severe ground-glass lesions and consolidation.

[0251] Treatment with fremanezumab was given for 10 consecutive days. Computed tomography (CT) scans of the lungs were obtained and analyzed at -2 days prior to treatment and at study completion on day 13. Baseline lung CT scans were compared to scans obtained on day 13. Subjects were classified as worsening if they increased by one or more stages, as improved if they decreased by one stage, and as having a significant improvement if they decreased by 2 or more stages. Subjects were stable if they did not change in stage. Lung CT analysis was performed in a blinded fashion by three radiologists.

[0252] Results from the lung CT scans are shown in Figure 7 Axial images of a control patient show extensive ground-glass opacities (bilateral upper lobes and anterior and posterior segments of the right middle lobe) two days prior to treatment (I), showing significant progression on the 13-day follow-up (II). III-IV: Axial images in a patient treated with fremanezumab and dexamethasone (fremanezumab / dexamethasone) show extensive ground-glass opacities in the anterior and posterior segments and consolidation in both lower lobes (III), showing partial resolution on the 13-day follow-up scan (IV). V-VI: Axial images in a patient receiving fremanezumab show ground-glass opacities in the posterior segments of the lung (V), showing interval resolution on the 13-day follow-up scan (VI).

[0253] Each patient was classified as worsening, stable, improved, or significantly improved. As shown in Table 3.2, 1 / 10 of the subjects treated with foralumab + dexamethasone worsened. 10 / 14 control subjects, 2 / 10 foralumab + dexamethasone subjects, and 2 / 12 foralumab subjects remained stable. Regarding improvement, since 6 patients in the control group and 2 in the foralumab + dexamethasone group did not have lung involvement at day -2, they could not improve. Improvement occurred in 3 / 8 of the control group, 1 / 8 of the foralumab + dexamethasone group, and 5 / 12 of the foralumab group. Significant improvement was observed in 1 / 8 of the control group, 6 / 8 of the foralumab + dexamethasone group, and 5 / 12 of the foralumab group. From this, significant improvement was observed primarily in subjects who received foralumab + dexamethasone or foralumab alone. Control vs foralumab + dexamethasone, p = 0.01 and control vs foralumab / dexamethasone + foralumab, p = 0.04 (Chi-square analysis).

[0254] Table 3.2 Lung injury assessment

[0255] Control Fresolimumab + Dexamethasone Fresolimumab Worsened 0 / 14 1 / 10 0 / 12 Stable 10 / 14 2 / 10 2 / 12 Improved 3 / 8 1 / 8 5 / 12 Significantly Improved 1 / 8 6 / 8 5 / 12

[0256] Patient-Reported Outcomes (PROs) and Medical-Reported Outcomes

[0257] The PRO consisted of 15 questions with the following response systems: (1) Loss of smell (anosmia): 0 = normal, 3 = decreased, 5 = completely lost. (2) Taste disorder (ageusia): 0 = normal, 3 = decreased, 5 = completely lost. (3) Cough: 0 = not present, 3 = present for a short time, 5 = present for more than half a day. (4) Headache: 0 = not present, 3 = present for a short time, 5 = present for more than half a day; (5) Sore throat: 0 = not present, 3 = moderate, painful on swallowing, 5 = intense, severe pain on swallowing. (6) Difficulty breathing: 0 = not present, 3 = moderate, some lack of air, 5 = intense, difficulty breathing. (7) Nausea / vomiting: 0 = not present, 3 = nausea without vomiting, 5 = vomiting. 8) O2 saturation: 0 => 95, 3 = 94-95%, 5 = 91-93%. (9) Diarrhea assessed according to the Bristol scale (reference): 0 = 0-4 types, 3 = 5 or 6 types, 5 = 7 types. (10) Runny nose: 0 = not present, 3 = mucus in the nose, 5 = rhinorrhea (liquid); (11) Abdominal pain: 0 = not present, 3 = moderate, 5 = intense. (12) Myalgia: 0 = not present, 3 = moderate, 5 = intense (generalized). (13) Fever: 0 = not present, 3 = 37-38C, 5 => 38.0C. (14) Conjunctivitis: 0 = not present, 5 = present. (15) Appetite: 0 = normal, 3 = decreased, 5 = completely lost. General health (how do you feel today) was assessed using the Baker Wong scale for pain assessment (0-10). The maximum possible score was 85.

[0258] COVID-19 symptoms reported on Day -2 were compared to total symptoms on Day 13. Symptoms reported by subjects were graded according to the following domains: Domain 1 (weakness, fatigue, loss of appetite, body aches, back pain); Domain 2 (fever, chills, sweating); Domain 3 (nausea, diarrhea, upper abdominal pain); Domain 4 (ageusia); Domain 5 (anosmia); Domain 6 (runny nose, sore throat, sneezing); Domain 7 (headache, anxiety, eye pain, dizziness); Domain 8 (cough, difficulty breathing, chest pain). Each subject scored one symptom in each domain.

[0259] At Day -2, subjects had experienced symptoms in an average of 5 domains for an average of 6 days. Most subjects improved during the course of the study, with no large differences between treatment groups. At the end of the study, 23 of 39 subjects (58.9%) were asymptomatic; 8 of 16 (50%) in the control group, 6 of 11 (54.5%) in the fremanezumab + dexamethasone group, and 9 of 12 (75%) in the fremanezumab group. Of the 16 subjects who remained symptomatic at the end of the study, anosmia (domain 5) and cough (domain 8) were the most common symptoms. There were no controls reporting rapid recovery from anosmia and ageusia in either fremanezumab treatment group. These results suggest that intranasal administration of fremanezumab can improve COVID-19 symptoms.

[0260] Other embodiments

[0261] While the present disclosure has been described in connection with specific embodiments thereof, the foregoing description is intended to illustrate and not limit the scope of the disclosure, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims. SEQUENCE LISTING <110> Tiziana Life Sciences PLC <120> CD3 antibody for treating coronavirus <130> TIZI-028 / 001WO 322161-2459 <150> 63 / 058,978 <151> 2020-07-30 <160> 53 <170> PatentIn version 3.5 <210> 1 <211> 375 <212> DNA <213> Artificial sequence <220> <223> 3B9 VL1-VH nucleic acid sequence <400> 1 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctgggtcctc ggtgaaggtc 60 tcctgcaagg cttctggagg caccttcagc agctatgcta tcagctgggt gcgccaggcc 120 cctggacaag ggcttgagtg gatgggaggg atcatccctc tctttgatac aacaaagtac 180 gcacagcagt tccagggcag agtcacgatt accgcggacg aatccacgag cacagcctac 240 atggagctga gcagcctgag atctgaggac acggccgtat tttactgtgc gagagatcgg 300 gatattttga ctgattatta tcccatgggc ggtatggacg tctggggcca agggaccacg 360 gtcaccgtct cctca 375 <210> 2 <211> 125 <212> PRT <213> Artificial Sequence <220> <223> 3B9 VL1-VH Amino Acid Sequence <400> 2 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Gly Thr Phe Ser Ser Tyr 20 25 30 Ala Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Leu Phe Asp Thr Thr Lys Tyr Ala Gln Gln Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Ala Asp Glu Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Phe Tyr Cys 85 90 95 Ala Arg Asp Arg Asp Ile Leu Thr Asp Tyr Tyr Pro Met Gly Gly Met 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser Ser 115 120 125 <210> 3 <211> 324 <212> DNA <213> Artificial Sequence <220> <223> 39B9 VL1-VL nucleic acid sequence <400> 3 gccatccagt tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60 atcacttgcc gggcaagtca gggcattagc agtgttttag cctggtatca gcagaaacca 120 gggaaagctc ctaagctcct gatctatgat gcctccagtt tggaaagtgg ggtcccatca 180 aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240 gaagattttg caacttatta ctgtcaacag tctaatagtt acccgctcac tttcggcgga 300 gggaccaagg tggagatcaa acgt 324 <210> 4 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> 39B9 VL1-VL amino acid sequence <400> 4 Ala Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Ser Ser Val 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Ser Leu Glu Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Asn Ser Tyr Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg 100 105 <210> 5 <211> 324 <212> DNA <213> Artificial sequence <220> <223> 39B9 VL5-VL nucleic acid sequence <400> 5 gacatcctga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60 atcacttgtc gggcgagtca ggatattagc agctggttag cctggtatca gcagaaacca 120 gggaaagctc ctaagctcct gatctatgat gcctccagtt tggaaagtgg ggtcccatca 180 aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240 gaagattttg caacttatta ctgtcaacag tctaatagtt acccgctcac tttcggcgga 300 gggaccaagg tggagatcaa acga 324 <210> 6 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> 39B9 VL5-VL amino acid sequence <400> 6 Asp Ile Leu Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Ser Trp 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Ser Leu Glu Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Ser Asn Ser Tyr Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg 100 105 <210> 7 <211> 357 <212> DNA <213> Artificial Sequence <220> <223> 12A VH nucleic acid sequence <400> 7 caggtgcagc tggtggagtc ttggggaggc gtggtccagc ctgggaggtc cctgagactc 60 tcctgtgcag cgtctggatt caccttcagt aactatgaca tgtactgggt ccgccaggct 120 ccaggcaagg ggctggagtg ggtggcagtt atattagatg atggaaataa taattactac 180 gcagactccg tgaagggccg attcaccatc tccagagaca attccaagaa aaaggtgtat 240 ctgcaaatga atagcctgag agctgaggac acggctgtgt attactgtgt gagagcgtcc 300 cctaactggg gtcttcttga cttctggggc cagggaaccc tggtcaccgt ctcgagt 357 <210> 8 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> 12A VH amino acid sequence <400> 8 Gln Val Gln Leu Val Glu Ser Trp Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Asp Met Tyr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Leu Asp Asp Gly Asn Asn Asn Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Lys Lys Val Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val Arg Ala Ser Pro Asn Trp Gly Leu Leu Asp Phe Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 9 <211> 324 <212> DNA <213> artificial sequence <220> <223> 12A VL nucleic acid sequence <400> 9 gaaattgtgt tgacacagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60 atcacttgtc gggcgagtca gggtattagc agctggttag cctggtatca gcagaaacca 120 gggaaagctc ctaagctcct gatctatgat gcctccagtt tggaaagtgg ggtcccatca 180 aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240 gaagattttg caacttatta ctgtcaacag tttaatagtt acccgatcac cttcggccaa 300 gggacacgac tggagattaa acgt 324 <210> 10 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> 12A VL amino acid sequence <400> 10 Glu Ile Val Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Ser Ser Trp 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Ser Leu Glu Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Phe Asn Ser Tyr Pro lie 85 90 95 Thr Phe Gly Gin Gly Thr Arg Leu Glu He Lys Arg 100 105 <210> 11 <211> 357 <212> DNA <213> Artificial Sequence <220> <223> 5C VH nucleic acid sequence <400> 11 caggtgcagc tggtgcagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60 tcctgtgcag cctctggatt catcttcagt agctatgaca tgtactgggt ccgccaggct 120 ccaggcaagg ggctggagtg ggtggcagtt atattatatg atggaaataa taaatactac 180 gcagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacggtgtat 240 ctgcaaatga acagcctgag agctgaggac acggctgtgt attactgtgt gagagcgtcc 300 cctaactggg gtctttttga cttctggggc cagggaaccc tggtcaccgt ctcgagt 357 <210> 12 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> 5C VH amino acid sequence <400> 12 Gln Val Gln Leu Val Gln Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Ile Phe Ser Ser Tyr 20 25 30 Asp Met Tyr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Leu Tyr Asp Gly Asn Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Val Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val Arg Ala Ser Pro Asn Trp Gly Leu Phe Asp Phe Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 13 <211> 324 <212> DNA <213> artificial sequence <220> <223> 5C VL nucleic acid sequence <400> 13 gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60 atcacttgcc gggcaagtca gggcattagc agtgatttag cctggtatca gcagaaacca 120 gggaaagctc ctaagctcct gatgtatgat gcctccagtt tggaaagtgg ggtcccatca 180 aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240 gaagattttg caacttatta ctgtcaacag tttaatagtt acccgatcac cttcggccaa 300 gggacacgac tggagattaa acgt 324 <210> 14 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> 5C VL amino acid sequence <400> 14 Asp Ile Gin Met Thr Gin Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gin Gly Ile Ser Ser Asp 20 25 30 Leu Ala Trp Tyr Gin Gin Lys Pro Gly Lys Ala Pro Lys Leu Leu Met 35 40 45 Tyr Asp Ala Ser Ser Leu Gin Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gin Pro Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gin Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Asn Ser Tyr Pro Ile 85 90 95 Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys Arg 100 105 <210> 15 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> 39B9 VH CDR1 <400> 15 Ser Tyr Ala Ile Ser 1 5 <210> 16 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> 39B9 VH CDR2 <400> 16 Gly Ile Ile Pro Leu Phe Asp Thr Thr Lys Tyr Ala Gln Gln Phe Gln 1 5 10 15 Gly <210> 17 <211> 19 <212> PRT <213> Artificial Sequence <220> <223> 39B9 VH CDR3 <400> 17 Cys Ala Arg Asp Arg Asp Ile Leu Thr Asp Tyr Tyr Pro Met Gly Gly 1 5 10 15 Met Asp Val <210> 18 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> 12A VH CDR1 <400> 18 Asn Tyr Asp Met Tyr 1 5 <210> 19 <211 > 17 <212> PRT <213> Artificial Sequence <220> <223> 12A VH CDR2 <400> 19 Val lie Leu Asp Asp Gly Asn Asn Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> 20 <211 > 13 <212> PRT <213> Artificial Sequence <220> <223> 12A VH CDR3 <400> 20 Cys Val Arg Ala Ser Pro Asn Trp Gly Leu Leu Asp Phe 1 5 10 <210> 21 <211 > 5 <212> PRT <213> Artificial Sequence <220> <223> 5C VH CDR1 <400> 21 Ser Tyr Asp Met Tyr 1 5 <210> 22 <211 > 17 <212> PRT <213> Artificial Sequence <220> <223> 5C VH CDR2 <400> twenty two Val Ile Leu Tyr Asp Gly Asn Asn Lys Tyr Tyr Ala Asp Ser Val Lys 1 5 10 15 Gly <210> twenty three <211> 13 <212> PRT <213> Artificial sequence <220> <223> 5C VH CDR3 <400> twenty three Cys Val Arg Ala Ser Pro Asn Trp Gly Leu Phe Asp Phe 1 5 10 <210> twenty four <211> 11 <212> PRT <213> Artificial sequence <220> <223> 39B9 VL1 VL CDR1 <400> twenty four Arg Ala Ser Gln Gly Ile Ser Ser Val Leu Ala 1 5 10 <210> 25 <211> 7 <212> PRT <213> Artificial sequence <220> <223> IL-6 antibody VL CDR2 <400> 25 Asp Ala Ser Ser Leu Glu Ser 1 5 <210> 26 <211> 9 <212> PRT <213> Artificial sequence <220> <223> 39B9 VL1 VL CDR3 <400> 26 Gln Gln Ser Asn Ser Tyr Pro Leu Thr 1 5 <210> 27 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 39B9 VL5 VL CDR1 <400> 27 Arg Ala Ser Gln Asp Ile Ser Ser Trp Leu Ala 1 5 10 <210> 28 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 12A VL CDR1 <400> 28 Arg Ala Ser Gln Gly Ile Ser Ser Trp Leu Ala 1 5 10 <210> 29 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> 12A VL CDR3 <400> 29 Gln Gln Ser Asn Ser Tyr Pro Ile Thr 1 5 <210> 30 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 5C VL CDR1 <400> 30 Arg Ala Ser Gln Gly Ile Ser Ser Val Asp Ala 1 5 10 <210> 31 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> VL CDR3 Motif <220> <221> MISC_FEATURE <222> (4)..(4) <223> Xaa at position 4 is Asparagine or Glutamine <400> 31 Gln Gln Ser Xaa Ser Tyr Pro Leu Thr 1 5 <210> 32 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> VH CDR2 Motif <220> <221> MISC_FEATURE <222> (5)..(5) <223> Xaa at position 5 can be Leucine or Alanine <220> <221> MISC_FEATURE <222> (7)..(7) <223> Xaa at position 7 can be Aspartic Acid or Glutamic Acid <220> <221> MISC_FEATURE <222> (14)..(14) <223> Xaa at position 14 can be Glutamine or Lysine <400> 32 Gly Ile Ile Pro Xaa Phe Xaa Thr Thr Lys Tyr Ala Gln Xaa Phe Gln 1 5 10 15 Gly <210> 33 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> VH CDR3 motif <220> <221> MISC_FEATURE <222> (11)..(11) <223> Xaa at position 11 is methionine or leucine <400> 33 Asp Arg Asp Ile Leu Thr Asp Tyr Tyr Pro Xaa Gly Gly Met Asp Val 1 5 10 15 <210> 34 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> FRW3 motif <220> <221> MISC_FEATURE <222> (4)..(4) <223> Xaa at position 4 is phenylalanine or tyrosine <400> 34 Thr Ala Val Xaa Tyr Cys Ala Arg 1 5 <210> 35 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> NI-1201-WT VH CDR3 <400> 35 Asp Arg Asp Ile Leu Thr Asp Tyr Tyr Pro Met Gly Gly Met Asp Val 1 5 10 15 <210> 36 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> NI-1201-WT FRW3 <400> 36 Thr Ala Val Phe Tyr Cys Ala Arg 1 5 <210> 37 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> NI-1201-WT VH CDR2 <400> 37 Gly Ile Ile Pro Leu Phe Asp Thr Thr Lys Tyr Ala Gln Lys Phe Gln 1 5 10 15 Gly <210> 38 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> NI-1201-A VL FRW3 <400> 38 Thr Ala Val Tyr Tyr Cys Ala Arg 1 5 <210> 39 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> NI-1201-B VH CDR3 <400> 39 Asp Arg Asp Ile Leu Thr Asp Tyr Tyr Pro Leu Gly Gly Met Asp Val 1 5 10 15 <210> 40 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> NI-1201-C VH CDR2 <400> 40 Gly Ile Ile Pro Ala Phe Glu Thr Thr Lys Tyr Ala Gln Lys Phe Gln 1 5 10 15 Gly <210> 41 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> NI-1201-D VL CDR3 <400> 41 Gln Gln Ser Gln Ser Tyr Pro Leu Thr 1 5 <210> 42 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 CDRH1 <400> 42 Gly Tyr Gly Met His 1 5 <210> 43 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 CDRH2 <400> 43 Val Ile Trp Tyr Asp Gly Ser Lys Lys Tyr Tyr Val Asp Ser Val Lys 1 5 10 15 Gly <210> 44 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 CDRH3 <400> 44 Gln Met Gly Tyr Trp His Phe Asp Leu 1 5 <210> 45 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 CDRL1 <400> 45 Arg Ala Ser Gln Ser Val Ser Ser Tyr Leu Ala 1 5 10 <210> 46 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 CDRL2 <400> 46 Asp Ala Ser Asn Arg Ala Thr 1 5 <210> 47 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 CDRL3 <400> 47 Gln Gln Arg Ser Asn Trp Pro Pro Leu Thr 1 5 10 <210> 48 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 VH <400> 48 Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Lys Phe Ser Gly Tyr 20 25 30 Gly Met His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val He Trp Tyr Asp Gly Ser Lys Lys Tyr Tyr Val Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr He Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gin Met Gly Tyr Trp His Phe Asp Leu Trp Gly Arg Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 49 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 VL <400> 49 Glu He Val Leu Thr Gin Ser Pro Ala Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gin Ser Val Ser Ser Tyr 20 25 30 Leu Ala Trp Tyr Gin Gin Lys Pro Gly Gin Ala Pro Arg Leu Leu He 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly He Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr He Ser Ser Leu Glu Pro 65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gin Gin Arg Ser Asn Trp Pro Pro 85 90 95 Leu Thr Phe Gly Gly Gly Thr Lys Val Glu He Lys 100 105 <210> 50 <211> 448 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 HC <400> 50 Gln Val Gin Leu Val Glu Ser Gly Gly Gly Val Val Gin Pro Gly Arg 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Lys Phe Ser Gly Tyr 20 25 30 Gly Met His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val He Trp Tyr Asp Gly Ser Lys Lys Tyr Tyr Val Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr lie Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gin Met Gly Tyr Trp His Phe Asp Leu Trp Gly Arg Gly Thr 100 105 110 Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro 115 120 125 Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly 130 135 140 Cys Leu Val Lys Asp Tyr Phe Pro Gin Pro Val Thr Val Ser Trp Asn 145 150 155 160 Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin 165 170 175 Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser 180 185 190 Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Val Asn His Lys Pro Ser 195 200 205 Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr 210 215 220 His Thr Cys Pro Pro Cys Pro Ala Pro Glu Ala Glu Gly Gly Pro Ser 225 230 235 240 Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg 245 250 255 Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro 260 265 270 Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala 275 280 285 Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val 290 295 300 Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr 305 310 315 320 Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr 325 330 335 Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu 340 345 350 Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys 355 360 365 Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser 370 375 380 Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp 385 390 395 400 Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser 405 410 415 Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala 420 425 430 Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 435 440 445 <210> 51 <211> 215 <212> PRT <213> Artificial Sequence <220> <223> Anti-CD3 LC <400> 51 Glu Ile Val Leu Thr Gln Ser Pro Ala Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr lie Ser Ser Leu Glu Pro 65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gin Gin Arg Ser Asn Trp Pro Pro 85 90 95 Leu Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys Arg Thr Val Ala 100 105 110 Ala Pro Ser Val Phe lie Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser 115 120 125 Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu 130 135 140 Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser 145 150 155 160 Gln Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu 165 170 175 Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val 180 185 190 Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys 195 200 205 Ser Phe Asn Arg Gly Glu Cys 210 215 <210> 52 <211> 454 <212> PRT <213> Artificial sequence <220> <223> TZLS-501 heavy chain <400> 52 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ser 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Gly Thr Phe Ser Ser Tyr 20 25 30 Ala Ile Ser Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Gly Ile Ile Pro Leu Phe Asp Thr Thr Lys Tyr Ala Gln Lys Phe 50 55 60 Gln Gly Arg Val Thr Ile Thr Ala Asp Glu Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Asp Arg Asp Ile Leu Thr Asp Tyr Tyr Pro Met Gly Gly Met 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser Ser Ala Ser Thr 115 120 125 Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser 130 135 140 Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu 145 150 155 160 Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His 165 170 175 Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser 180 185 190 Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys 195 200 205 Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu 210 215 220 Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro 225 230 235 240 Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys 245 250 255 Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val 260 265 270 Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp 275 280 285 Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr 290 295 300 Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp 305 310 315 320 Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu 325 330 335 Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gin Pro Arg 340 345 350 Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys 355 360 365 Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp 370 375 380 Ile Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys 385 390 395 400 Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser 405 410 415 Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser 420 425 430 Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser 435 440 445 Leu Ser Leu Ser Pro Gly 450 <210> 53 <211> 214 <212> PRT <213> Artificial sequence <220> <223> TZLS-501 heavy chain <400> 53 Ala Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Gly Ile Ser Ser Val 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Ser Leu Glu Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Asn Ser Tyr Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg Thr Val Ala Ala 100 105 110 Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala 130 135 140 Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin 145 150 155 160 Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser 165 170 175 Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr 180 185 190 Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser 195 200 205 Phe Asn Arg Gly Glu Cys 210

Claims

1. Use of an anti-CD3 antibody or a composition comprising an anti-CD3 antibody in the manufacture of a medicament for treating, preventing, or reducing symptoms of a coronavirus infection in a subject in need thereof, wherein the anti-CD3 antibody comprises a heavy chain complementarity determining region 1 (CDRH1) consisting of the amino acid sequence GYGMH (SEQ ID NO: 42), a heavy chain complementarity determining region 2 (CDRH2) consisting of the amino acid sequence VIWYDGSKKYYVDSVKG (SEQ ID NO: 43), a heavy chain complementarity determining region 3 (CDRH3) consisting of the amino acid sequence QMGYWHFDL (SEQ ID NO: 44), a light chain complementarity determining region 1 (CDRL1) consisting of the amino acid sequence RASQSVSSYLA (SEQ ID NO: 45), a light chain complementarity determining region 2 (CDRL2) consisting of the amino acid sequence DASNRAT (SEQ ID NO: 46), and a light chain complementarity determining region 3 (CDRL3) consisting of the amino acid sequence QQRSNWPPLT (SEQ ID NO: 47), and wherein the coronavirus infection is COVID-19.

2. The use of claim 1, wherein the anti-CD3 antibody is a monoclonal antibody.

3. The use of claim 1 or 2, wherein the anti-CD3 antibody is fully human or humanized.

4. The use of claim 1 or 2, wherein the anti-CD3 antibody comprises a variable heavy chain amino acid sequence comprising the amino acid sequence of SEQ ID NO: 48 and a variable light chain amino acid sequence comprising the amino acid sequence of SEQ ID NO:

49.

5. The use of claim 1 or 2, wherein the anti-CD3 antibody comprises a heavy chain amino acid sequence comprising the amino acid sequence of SEQ ID NO: 50 and a light chain amino acid sequence comprising the amino acid sequence of SEQ ID NO:

51.

6. The use of claim 1 or 2, wherein the coronavirus is SARS-CoV-2 or a variant thereof.

7. The use of claim 1 or 2, wherein the symptom of the coronavirus infection is one or more of the following: an overactive immune response, loss of smell, taste disorder, cough, headache, sore throat, difficulty breathing, shortness of breath, nausea, vomiting, decreased O2 saturation, diarrhea, rhinorrhea, abdominal pain, myalgia, fever, conjunctivitis, and loss of appetite.

8. The use of claim 1 or 2, wherein the symptom of the coronavirus infection is a gastrointestinal symptom or a respiratory symptom.

9. The use of claim 7, wherein the overactive immune response comprises increased levels of at least one of interleukin 6 (IL-6), C-reactive protein (CRP), and D-dimer.

10. The use of claim 9, wherein the medicament decreases the levels of at least one of IL-6, CRP, and D-dimer.

11. The use of claim 1 or 2, wherein the subject has or is suspected of having a coronavirus infection.

12. The use of claim 1 or 2, wherein the subject has or is believed to have been exposed to a coronavirus and has not yet developed symptoms of a coronavirus infection.

13. The use of claim 1 or 2, the medicament further comprising a composition comprising dexamethasone.

14. The use of claim 1 or 2, wherein the composition is administered via mucosa, intravenously, or a combination thereof.

15. The use of claim 14, wherein the composition is administered orally, nasally, or by inhalation.

16. The use of claim 15, wherein the inhalation is by inhaler or nebulizer.

17. The use of claim 1 or 2, wherein the anti-CD3 antibody is administered nasally at a daily dose of 50 pg to 100 pg.

18. The use of claim 1 or 2, wherein the anti-CD3 antibody is administered orally at a daily dose of 1.0 to 2.5 mg.

19. The use of claim 1 or 2, wherein the medicament further comprises dexamethasone.

20. The use of claim 19, wherein the dexamethasone is administered by inhalation.

21. The use of claim 20, wherein the administration by inhalation is by metered dose inhaler.

22. The use of claim 1 or 2, wherein the symptom of a coronavirus infection is pain on swallowing or difficulty breathing.

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