Composition and methods for treating system lupus erythematosus

Abstract

A composition for treating system lupus erythematosus (SLE) includes a plant virus-based nanoparticle engineered to express at least a tolerogenic peptide associated with SLE and a buffer, of which a method of manufacturing includes infecting a plant with a virus to produce the plant virus-based nanoparticle, sampling symptomatic leaves from the plant, homogenizing the plant virus-based nanoparticle, incubating the plant virus-based nanoparticle, centrifuging the plant virus-based nanoparticle, and filtrating the plant virus-based nanoparticle.

Description

[0001]COMPOSITION AND METHODS FOR TREATING SYSTEM LUPUS ERYTHEMATOSUS CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority of U.S. Provisional Application Serial No. 63 / 530,125, filed on August 1, 2023, and entitled “COMPOSITION AND METHODS OF AN 5 ENGINEERED VIRUS-BASED NANOPARTICLE FOR THE TREATMENT OF LUPUS”, which is incorporated by reference herein in its entirety. FIELD OF THE INVENTION The present invention generally relates to the field of nanomedicine. In particular, the present invention is directed to a composition and methods of an engineered virus-based 10 nanoparticle for the treatment of system lupus erythematosus (SLE). REFERENCE TO SEQUENCE LISTING This specification includes a sequence listing submitted herewith, which includes the file entitled 1266-003PCT1.xml having the following size: 1,716 bytes which was created July 19, 2024, the contents of which are incorporated by reference herein. 15 BACKGROUND System lupus erythematosus (SLE) is an autoimmune disease wherein the body’s immune system erroneously attacks its own healthy tissues, leading to systemic inflammation and damage. Current therapeutic strategies often fail to fully control SLE or may induce significant side effects. Meanwhile, plant viruses, despite their simplicity, are intricate entities 20 capable of robust self-assembly, making them potentially useful templates in nanobiotechnology. SUMMARY OF THE DISCLOSURE In an aspect, a composition for treating system lupus erythematosus (SLE) is described. Composition includes a plant virus-based nanoparticle engineered to express at least a tolerogenic peptide associated with SLE and a buffer. 25 In another aspect, a method of manufacturing composition for treating SLE is described. Method includes infecting a plant with a virus to produce a plant virus-based nanoparticle, sampling symptomatic leaves from the plant, homogenizing the plant virus-based nanoparticle, incubating the plant virus-based nanoparticle, centrifuging the plant virus-based nanoparticle, and filtrating the plant virus-based nanoparticle. 30 In another aspect, another method of manufacturing composition for treating SLE is described. Method includes infecting a plant with a virus to produce a plant virus-based 1 Attorney Docket No.1266-003PCT1 nanoparticle, sampling symptomatic leaves from the plant, homogenizing the plant virus-based nanoparticle, incubating the plant virus-based nanoparticle, and filtrating the plant virus-based nanoparticle. In another aspect, another method of manufacturing composition for treating SLE is 5 described. Method includes agroinfiltrating a plant with a virus to produce a plant virus-based nanoparticle, sampling symptomatic leaves of the plant, homogenizing the plant virus-based nanoparticle, centrifuging the plant virus-based nanoparticle, and filtrating the plant virus-based nanoparticle. The details of one or more variations of the subject matter described herein are set forth 10 in the accompanying drawings and the description below. Other features and advantages of the subject matter described herein will be apparent from the description and drawings, and from the claims. DESCRIPTION OF DRAWINGS For the purpose of illustrating the invention, the drawings show aspects of one or more 15 embodiments of the invention. However, it should be understood that the present invention is not limited to the precise arrangements and instrumentalities shown in the drawings, wherein: FIG.1 is a schematic illustration of an exemplary embodiment of a composition for treating system lupus erythematosus (SLE). FIGS.2A-B are schematic illustrations of exemplary embodiments of a plant virus-based 20 nanoparticle; FIG.3A is a schematic illustration of a Tomato bushy stunt virus (TBSV); FIG.3B is a schematic illustration of a Cowpea mosaic virus (CPMV); FIG.4 is a flow diagram illustrating an exemplary embodiment of a method of manufacturing the composition for treating SLE; 25 FIG.5 is a flow diagram illustrating an exemplary embodiment of another method of manufacturing the composition for treating SLE; FIG.6 is an exemplary embodiment of a method of using plant virus-based nanoparticle for treating SLE; and FIG.7 is an exemplary embodiment of an extraction process of a plant virus-based nanoparticle. 30 The drawings are not necessarily to scale and may be illustrated by phantom lines, diagrammatic representations, and fragmentary views. In certain instances, details that are not 2 Attorney Docket No.1266-003PCT1 necessary for an understanding of the embodiments or that render other details difficult to perceive may have been omitted. Like reference symbols in the various drawings indicate like elements. DETAILED DESCRIPTION 5 At a high level, aspects of the present disclosure are directed to compositions and methods for treating system lupus erythematosus (SLE). In one or more embodiments, composition may include a plant virus-based nanoparticle engineered to express at least a tolerogenic peptide associated with SLE and a buffer. In one or more embodiments, method may include identifying tolerogenic peptide associated with SLE, modifying plant-virus based 10 nanoparticle to express the identified tolerogenic peptide, and synthesizing the plant virus-based nanoparticle containing the identified tolerogenic peptide. Aspects of the present disclosure can be used to prevent and treat autoimmune diseases such as SLE using plant virus-based nanoparticles. Referring now to FIG.1, an exemplary embodiment of a composition 100 for treating 15 SLE is illustrated. For the purposes of this disclosure, “system lupus erythematosus (SLE)” or simply “lupus” is a complex, multisystemic autoimmune disease that is often characterized by a production of autoantibodies that target a body’s own tissues, dysregulation of cytokines, apoptosis, and B- and T-cell functions, and / or systemic clinical manifestations that include inflammation and tissue damage. This complex disorder involves intricate interactions between 20 genetic, hormonal, immunological, and environmental factors. Clinical manifestations of SLE may range from milder forms such as skin rash and arthritis, to severe life-threatening systemic diseases involving the kidneys, heart, lungs, and nervous system. Current therapeutic strategies often fail to fully control SLE or may induce significant side effects, necessitating the development of new treatment approaches. 25 With continued reference to FIG.1, it is worth noting that the foundation of composition 100 lies in harnessing the immunomodulatory effect of a virus-based nanoparticle, which carries a tolerogenic peptide designed to recalibrate aberrant immune responses observed in SLE, as described below. For the purposes of this disclosure, an “immunomodulatory effect” is an effect of modulating the immune system. As a nonlimiting example, a plant virus-based nanoparticle 30 may modulate an immune system to induce tolerance and therefore prevent or treat a case of autoimmune disease, such as without limitation SLE. 3 Attorney Docket No.1266-003PCT1 With continued reference to FIG.1, composition 100 comprises a plant virus-based nanoparticle 104. For the purposes of this disclosure, a “nanoparticle” is a particle with a size that ranges between 1 and 500 nanometers. For the purposes of this disclosure, a “plant virus- based nanoparticle” is a virus nanoparticle that is capable of infecting and replicating in plants. 5 In one or more embodiments, plant virus-based nanoparticle 104 may include at least a unit or subunit of a plant viral protein. For the purposes of this disclosure, a “plant virus protein” is a protein that constitutes or is produced by a plant virus. Since plant viruses are unable to replicate in mammals, plant virus proteins may be a safer option for medical applications in human hosts. Nanoparticles created from subunits of plant virus proteins may be genetically engineered to 10 express, on their external surface, an antigen-specific peptide related to an autoimmune disease and then be grown in a plant host, as described in detail below in this disclosure. For the purposes of this disclosure, a “virus nanoparticle” is a proteinaceous and often infectious nanoscale structure that is capable of delivering its nucleic acid efficiently into a host cell, enabling production of new viruses therein. “Virus” and “virus nanoparticle” may be used 15 interchangeably throughout this disclosure. Similarly, “plant virus” and “plant virus-based nanoparticle” may be used interchangeably as well. Virus includes a capsid. For the purposes of this disclosure, a “capsid” is a protein shell exposed at the exterior of a virus that possesses a specific geometric pattern. As a nonlimiting example, capsid may possess an icosahedral shape. For the purposes of this disclosure, an “icosahedron” is a geometric shape with 20 sides, each 20 composed of an equilateral triangle. As another nonlimiting example, capsid may include a filamentous structure. For the purposes of this disclosure, a “filamentous structure” is an elongated, thread-like formation that makes up a capsid of certain viruses. As another nonlimiting example, capsid may include a rod-shaped structure. As another nonlimiting example, capsid may include a helical structure. For the purposes of this disclosure, a “helical 25 structure” is a type of structure characterized by a cylindrical, elongated shape with a helical symmetry. This structure may be formed by the regular, repeating arrangement of protein subunits around a viral nucleic acid, providing protection, structural integrity, and aiding in the infectivity of a virus. As another nonlimiting example, capsid may include a spherical structure. Additionally, and / or alternatively, capsid may adopt any geometry not disclosed herein yet 30 deemed possible by a person of ordinary skill in the art upon reviewing the entirety of this disclosure. 4 Attorney Docket No.1266-003PCT1 With continued reference to FIG.1, in one or more embodiments, plant virus protein may be produced through a process of molecular farming. For the purposes of this disclosure, “molecular farming” is a process of producing pharmaceutically important and commercially valuable proteins in plants. Once extracted, plant virus proteins may be used to create plant 5 virus-based nanoparticle 104. Plant virus-based nanoparticle 104 and / or plant virus protein may be sourced from a variety of plant hosts. Suitable host plants for such purpose may include a Nicotiana benthamiana plant, a Nicotiana tabacum plant, a Solanum lycopersicum or Lycopersicon esculentum plant, a Cycorium intybus plant, a Brassica oleracea var. capitata plant, a Beta vulgaris var cicla plant, a Ocimum basilicum plant, a red beet plant, a spinach plant, 10 or the like. For the purposes of this disclosure, a “Nicotiana benthamiana plant” is a close relative of tobacco and a species of Nicotiana indigenous to Australia. It may be used for farming monoclonal antibodies and other recombinant proteins. For the purposes of this disclosure, a “Nicotiana tabacum plant”, commonly known as cultivated tobacco, is a plant species that belongs to the Nicotiana genus in the Solanaceae family. It is a widely grown herbaceous plant 15 primarily used to produce tobacco products. This species is characterized by large, broad leaves and is cultivated in various climates worldwide. In some cases, a Nicotiana tabacum plant may serve as a model organism and a host for genetic engineering, enabling the expression and study of recombinant proteins, vaccines, and other biologically significant compounds. For the purposes of this disclosure, a “Solanum lycopersicum plant” or “Lycopersicon esculentum plant”, 20 commonly known as the tomato plant, is a plant species that belongs to the Solanaceae family and characterized by its production of edible, fleshy fruits. Solanum lycopersicum is noted for its agricultural significance and its utility in genetic engineering and plant breeding. This species may serve as a model organism for studying plant genetics, disease resistance, and metabolic pathways. It may also be utilized in biotechnological applications for the production of 25 recombinant proteins, novel traits, and improved cultivars through genetic modification techniques. For the purposes of this disclosure, a “Cycorium intybus plant” is a hardy plant widely used in folklore medicine to treat various ailments ranging from wounds to diabetes. It is believed to have antimicrobial, anthelmintic, antimalarial, hepatoprotective, antidiabetic, gastroprotective, anti-inflammatory, analgesic, antioxidant, tumor-inhibitory, and antiallergic 30 activities across several different cultures. For the purposes of this disclosure, a “Brassica oleracea var. capitata plant” is a is a biennial plant grown as an annual for its dense-leaved 5 Attorney Docket No.1266-003PCT1 heads and characterized by a short stem and a rosette of green, purple, or white leaves that form a tight, globular, and compact head. It is known as wild cabbage in its uncultivated form. Some cultivated forms of Brassica oleracea var. capitata plant include cabbage, broccoli, cauliflower, kale, brussels sprouts, collard greens, savoy cabbage, kohlrabi and gai lan. It is native to costal 5 southern and western Europe. For the purposes of this disclosure, a “Beta vulgaris var cicla plant” is a plant that is more commonly known as chard or spinach beet. It originates from the Mediterranean and has some medicinal properties, mainly in boosting the immune system and lowering blood pressure. For the purposes of this disclosure, an “Ocimum basilicum plant” is a member of the Lamiaceae (mint) family and is more commonly referred to as basil. For the 10 purposes of this disclosure, a “red beet plant” is a biennial plant grown as an annual for its edible root and leafy greens and characterized by its swollen root, which is typically deep red or purple in color. It is a promising candidate for some medicinal uses. As a nonlimiting example, the phytochemicals present in red beet may provide protection against diseases including cancer and cardiovascular diseases. For the purposes of this disclosure, a “spinach plant” is a leafy green 15 that belongs to the amaranth family and is closely related to beets and quinoa. With continued reference to FIG.1, in one or more embodiments, plant virus-based nanoparticle 104 may contain one or more viruses, such as without limitation Tomato bush stunt virus (TBSV), Cowpea mosaic virus (CPMV), and / or the like. For the purposes of this disclosure, a “Tomato bush stunt virus (TBSV)” is a plant virus from the Tombusvirus group that 20 primarily infects vegetable crops and causes stunting of growth, leaf mottling, and deformed or absent fruit in an infected plant. TBSV is a prototypic member of the Tombusviridae family. TBSV may provide a scaffold for plant virus-based nanoparticle 104 to enable a stable C- terminal display of peptides. The structure of TBSV has been resolved at an atomic resolution with a 30-nm capsid composed of 180 identical copies of a single coat protein (CP / p41) arranged 25 in T = 3 symmetry. CP may include an RNA binding domain (R), a shell domain (S) forming a capsid backbone, and a C-terminal protruding (P) domain that can accommodate exogenous peptides for display. The structure of TBSV may include a single-stranded RNA with a linear genome of about 4,800 nucleotides. TBSV may also possess three symmetrically distinct coat protein monomers. TBSV may replicate, without limitation, using cytoplasmic replication. This 30 virus may penetrate a host cell by uncoating and releasing viral RNA into cytoplasm. TBSV may also spread indirectly, such as without limitation through water, soil, and / or infected seeds. 6 Attorney Docket No.1266-003PCT1 When engineered to express peptides, the behavior of TBSV may experience one or more modifications, such as without limitation a decrease in virus concentration, infection percentage, and / or disease severity. With continued reference to FIG.1, for the purposes of this disclosure, a “Cowpea 5 mosaic virus (CPMV)” or “Sunn-hemp mosaic virus” is a non-enveloped plant virus of the Comovirus group. Infection of a susceptible cowpea leaf may cause a "mosaic" pattern in the leaf and result in high virus yields. CPMV genome may include 2 strands of RNA which are separately encapsulated. CPMV may be approximately 30 nanometers in diameter and contain 60 copies each of a Large (L) and Small (S) coat proteins, as explained below in this disclosure. 10 The structure of CPMV is well-characterized to an atomic resolution. CPMV nanoparticles may be thermostable and readily isolated from plants. There are many stable mutants of CPMV already prepared that allow specific modification of their capsid surfaces. In some cases, CPMV may include an icosahedral capsid, as described above. With continued reference to FIG.1, plant virus-based nanoparticle 104 is configured to 15 express at least a tolerogenic peptide 108. For the purposes of this disclosure, a “tolerogenic peptide” is a short chain of amino acids designed to induce immune tolerance, particularly in the context of autoimmune diseases, allergies, and transplant rejection. By promoting tolerance, a tolerogenic peptide may prevent an immune system from attacking a body's own tissues or introduced antigens without broadly suppressing immune function. In one or more embodiments, 20 tolerogenic peptide 108 may include hCDR1. For the purposes of this disclosure. “hCDR1” is a tolerogenic peptide with a sequence of HGYYWSWIRQPPGKGEEWI (SEQ ID NO: 1) that may significantly down-regulate the mRNA expression of pathogenic cytokines and pro- apoptotic molecules, while up-regulating the expression of TGF-beta and FoxP3. This may result in a significant decrease in SLEDAI-2K and BILAG scores, suggesting that hCDR1 may be a 25 disease-specific treatment for SLE patients. hCDR1 has been used on mice and has shown potential beneficial effect in human lupus. As a nonlimiting example, a treatment with hCDR1 may result in significant reduction in the gene expression of inflammatory cytokines IL-1b, TNF- a, IFN-g, and IL-10, and B Lymphocyte Stimulator (BLyS), a protein believed to play a role in the development of SLE. This reduction was observed in all patients treated with hCDR1. 30 hCDR1 is also known as edratide and may be designed based on complementarity determining regions 1 (CDR1) of a human ant-DNA autoantibody. 7 Attorney Docket No.1266-003PCT1 With continued reference to FIG.1, in one or more embodiments, tolerogenic peptide 108 may include an immunodominant peptide. For the purposes of this disclosure, an “immunodominant peptide” is a peptide that is a representative epitope of a given protein antigen to an immune system in the context of a specific autoimmune disease. In one or more 5 embodiments, immunodominant peptide may include a cytoplasmically located protein. For the purposes of this disclosure, a “cytoplasmical location” is a location inside the cytoplasm of a cell. In one or more embodiments, a coat protein of plant virus-based nanoparticle 104 may be engineered to display immunodominant peptides. For the purposes of this disclosure, a “coat protein” is a protein that is a constituent of the capsid of a virus, as described above. In one or 10 more embodiments, at least an immunodominant peptide may be fused to a protein of CPMV and / or TBSV. In one or more embodiments, the presence of immunodominant peptide may reduce the yield of plant virus-based nanoparticles 104 in plants. As a nonlimiting example, the presence of hCDR1 may reduce the yield of CPMV and / or TBSV nanoparticles. In some embodiments, the presence of immunodominant peptide may improve the yield of plant virus- 15 based nanoparticles 104 in plants. As a nonlimiting example, the presence of hCDR1 may improve the yield of CPMV and / or TBSV. With continued reference to FIG.1, composition 100 further comprises a buffer 112. Buffer may stabilize plant virus-based nanoparticle 104 and maintain a pH for increased stability and functionality. For the purposes of this disclosure, a “buffer” is a solution or mixture that 20 contains at least a pair of weak acid, HA, and its conjugate base, A-, (i.e., the weak acid minus one proton) in a molar ratio between 10:1 and 1:10, wherein the solution maintains a stable pH close to the pKa(i.e., the negative log of the acid dissociation constant, Ka) of the weak acid, against addition of acidic or basic chemical species. For simplicity, a buffer containing a pair of conjugate base and acid may be written as A- / HA. Additional examples will be provided below. 25 The pH of a buffer solution may be calculated using the Henderson Hasselbalch equation: ^ [ log ^ ^^ି] ^^ ^^ = ^^ ^^ + [ ^^ ^^]^ With continued reference to FIG.1, buffer may include any type of buffer deemed suitable by a person of ordinary skill in the art upon reviewing the entirety of this disclosure. As another nonlimiting example, buffer may include an acetate buffer (i.e.,30CH3COONa / CH3COOH). As another nonlimiting example, buffer may include a borate buffer 8 Attorney Docket No.1266-003PCT1 (i.e., Na2B4O7·10H2O / H3BO3). As another nonlimiting example, buffer may include a bicarbonate buffer (i.e., NaHCO3 / H2CO3or Na2CO3 / NaHCO3, depending on the desired pH). As another nonlimiting example, composition 100 may include a cacodylate buffer (i.e., NaC2H6AsO2 / HC2H6AsO2). As another nonlimiting example, buffer may include a Good’s buffer. For the purposes of this disclosure, “Good’s buffers” are a group of more than 20 conjugate acid / base pairs selected and described by Norman Good and colleagues for biochemical and biological research during 1966–1980. For simplicity, only the conjugate acid may be shown for each conjugate acid / base pair. Good’s buffers include MES (C₆H₁₃NO₄S), ACES (C₄H₉NO₄S), PIPES (C₈H₁₈N₂O₆S₂), MOPS (C₇H₁₅NO₄S), TES (C₆H₁₅NO₆S), HEPES (C₈H₁₈N₂O₄S), Tricine (C₆H₁₃NO₅), TRIS (C₄H₁₁NO₃), Bicine (C₆H₁₃NO₄), TAPS (C₇H₁₇NO₆S), CHES (C₈H₁₇NO₃S), CAPS (C₉H₁₉NO₃S), AMPSO (C₉H₁₉NO₄S), Gly-Gly (C₄H₈N₂O₃), ADA (C₄H₇NO₄), BES (C₆H₁₅NO₅S), MOPSO (C₇H₁₅NO₅S), EPPS (C₉H₂₀N₂O₄S), HEPPS (C₁₁H₂₄N₂O₄S), CAPSO (C₉H₁₉NO₄S), HEPPSO (C₉H₂₀N₂O₅S), CABS (C₁₀H₁₉NO₃S), ACESO (C₄H₉NO₅S), TES-Na (C₆H₁₄NO₆SNa), BICINE-Na (C₆H₁₂NO₄Na), TRICINE-Na (C₆H₁₂NO₅Na), MES-Na (C₆H₁₂NO₄SNa), HEPES-Na (C₈H₁₇N₂O₄SNa), MOPS-Na (C₇H₁₄NO₄SNa), and PIPES-Na (C₈H₁₇N₂O₆S₂Na). As a nonlimiting example, buffer may include a phosphate buffer (i.e., NaH2PO4 / H3PO4, Na2HPO4 / NaH2PO4, or Na3HPO4 / Na2HPO4, depending on the desired pH). As another nonlimiting example, buffer may include a phosphate- buffered saline (PBS) solution, a commonly used buffer in biological research and pharmaceutical formulations that typically contains 137 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, and 1.8 mM KH2PO4. These buffer components may help to preserve the structural integrity of plant virus-based nanoparticle 104 and its associated tolerogenic peptide 108, ensuring their effective delivery at a target site in a body. A person of ordinary skill in the art, upon reviewing the entirety of this disclosure, will be able to recognize suitable buffers 112 for composition 100. With continued reference to FIG.1, in one or more embodiments, composition 100 may include sodium acetate. For the purposes of this disclosure, “sodium acetate” is organic sodium salt with a chemical formula of CH3COONa. It may also be known as acetic acid, sodium salt, sodium acetate anhydrous, or acetic acid sodium salt. Sodium acetate is the anhydrous, sodium salt form of acetic acid. Sodium acetate may be a hygroscopic powder that absorbs moisture from air. Sodium acetate is a strong electrolyte that fully dissolves and disassociates in water to 9 Attorney Docket No.1266-003PCT1 form sodium ions (Na+) and acetate ions (CH3COO-). Na+is the principal cation of extracellular fluid and plays a large part in fluid and electrolyte replacement therapies. Sodium acetate may be used as an electrolyte replenisher in isosmotic solution for parenteral replacement of acute losses of extracellular fluid without disturbing normal electrolyte balance. A solution of sodium acetate 5 and acetic acid may act as a buffer to maintain a relatively constant pH. This may be useful especially in biochemical applications where reactions are pH-dependent in a mildly acidic range. In this case, sodium acetate is used to titrate a composition to adjust its pH. With continued reference to FIG.1, composition 100 may be taken in any suitable form of dosage and / or delivery, including without limitation oral dosage and intravenous dosage. For 10 the purposes of this disclosure, an “oral dosage” is an ingestion of a composition through the mouth. Oral dosage of composition 100 may include use of pills, syrup, tablets, thin film, liquid solution, powder, solid crystals, natural or herbal plants, seeds, or food, pastes, or the like. For the purposes of this disclosure, an “intravenous dosage” is an administration of a composition using injection. Intravenous dosage may include without limitation intravenous injection. 15 Composition 100 may be given intravenously in any suitable manner, including as a bolus and / or as an infusion. Alternatively, other injection methods such as intramuscular injection, intraperitoneal injection, subcutaneous injection, and / or transcutaneous injection may be used. With continued reference to FIG.1, in one or more embodiments, composition 100 may include one or more anti-lupus ingredients. For the purposes of this disclosure, an “anti-lupus20 ingredient” is a medically active chemical that may be used to treat and / or prevent SLE. Anti- lupus ingredient may include a non-steroidal anti-inflammatory drug (NSAID), a corticosteroid, methotrexate, hydroxychloroquine, sulfasalazine, leflunomide, a tumor necrosis factor inhibitor, a T-cell costimulatory blocking agent, a B-cell depleting agent, an Interleukin-1 receptor antagonist therapy, and / or any other immunomodulatory and / or cytotoxic agent. 25 Referring now to FIG.2A, an exemplary embodiment 200a of plant virus-based nanoparticle 104 is illustrated. Embodiment 200a may be a structural representation of CPMV. In one or more embodiments, plant virus-based nanoparticle 104 may include a large coat protein subunit 204. As a nonlimiting example, for CPMV, plant virus-based nanoparticle 104 may include 60 copies of large coat protein subunit 204. In one or more embodiments, plant virus- 30 based nanoparticle 104 may include a small coat protein subunit 208. As a nonlimiting example, plant virus-based nanoparticle 104 may include 60 copies of small coat protein subunit 208. In 10 Attorney Docket No.1266-003PCT1 one or more embodiments, plant virus-based nanoparticle 104 may include at least a tolerogenic peptide 108. In one or more embodiments, at least a tolerogenic peptide 108 may include immunodominant peptide 212. In one or more embodiments, immunodominant peptide 212 may be embedded within coat protein. In one or more embodiments, immunodominant peptide 212 5 may be located outside of coat protein. In one or more embodiments, immunodominant peptide 212 may be exposed on the surface of large coat protein subunit 204. In one or more embodiments, immunodominant peptide 212 may be located inside of a nanoparticle. In one or more embodiments, immunodominant peptide 212 may be placed inside of coat protein. In one or more embodiments, plant virus-based nanoparticle 104 may include an icosahedral structure. 10 In one or more embodiments, plant virus-based nanoparticle 104 may include a filamentous structure. In one or more embodiments, plant virus-based nanoparticle 104 may include a rod- shaped structure. In one or more embodiments, plant virus-based nanoparticle 104 may include a helical structure. In one or more embodiments, plant virus-based nanoparticle 104 may include a spherical structure. In one or more embodiments, plant virus-based nanoparticle 104 may be 15 homogeneous or uniform in size. In some embodiments, plant virus-based nanoparticle 104 may be homogeneous or uniform in shape. Additionally, and / or alternatively, plant virus-based nanoparticle 104 may adopt any geometry not disclosed herein yet deemed possible by a person of ordinary skill in the art upon reviewing the entirety of this disclosure. Referring now to FIG.2B, an exemplary embodiment 200b of plant virus-based 20 nanoparticle 104 is illustrated. Plant virus-based nanoparticle 104 disclosed herein may be consistent with details described with respect to FIG.2A. Embodiment 200b may be a structural representation of TBSV. In one or more embodiments, plant virus-based nanoparticle 104 may include TBSV, as described above. In one or more embodiments, plant virus-based nanoparticle 104 may include a single protein subunit 216. As a nonlimiting example, for TBSV, plant virus- 25 based nanoparticle 104 may include 180 copies of a single protein subunit 216. In one or more embodiments, plant virus-based nanoparticle 104 may include at least a tolerogenic peptide 108. In one or more embodiments, at least a tolerogenic peptide 108 may include immunodominant peptide 212. In one or more embodiments, immunodominant peptide 212 may be exposed on the surface of a single protein subunit 216. 30 Referring now to FIGS.3A-B, exemplary structures of TBSV (300a) and CPMV (300b) are illustrated in FIG.3A and 3B, respectively. 11 Attorney Docket No.1266-003PCT1 Referring now to FIG.4, a flow diagram illustrating an exemplary embodiment of method 400 for manufacturing plant virus-based nanoparticle 104 is illustrated. Plant virus-based nanoparticle 104 may include any type of plant virus-based nanoparticle described in this disclosure without limitation. At step 405, method 400 includes infecting a plant with a virus to 5 produce plant virus-based nanoparticle 104. For the purposes of this disclosure, “infection” is a process of delivering viral genes into a host, wherein the viral genes are capable of replication to produce new copies of the corresponding virus. Infection allows a plant to produce plant virus- based nanoparticle 104. In one or more embodiments, infection may be done by spontaneous infiltration. For the purposes of this disclosure, “spontaneous infiltration” is a type of infiltration 10 that is associated with a negative Gibbs free energy change and occurs naturally without energy input or intervention. For the purposes of this disclosure, “infiltration” is a process through which one or more substances penetrate or permeate from the surface of a plant into its tissues. As a nonlimiting example, spontaneous infiltration may include spraying infectious viral genes onto a plant, spraying viral solution obtained from previously infected leaves, immersion of 15 plants into a solution obtained from previously infected leaves and the like. As another nonlimiting example, infection may be done by infiltration of viral genes into a plant. As another nonlimiting example, infection may be done by infiltration of viral solution obtained from previously infected leaves. In one or more embodiments, infection may be done by forced infiltration. For the purposes of this disclosure, “forced infiltration” is a type of infiltration that 20 requires a force, a pressure, an energy input, or a similar form of intervention to be applied. As a nonlimiting example, forced infiltration may include syringe infiltration, vacuum infiltration, and the like. As another nonlimiting example, vacuum infiltration may include vacuum infiltration using a vacuum pump, vacuum infiltration using a syringe, and the like. A person of ordinary skill in the art, upon reviewing the entirety of this disclosure, will be aware of various infiltration 25 techniques that may be applied to generate plant virus-based nanoparticle 104 as described in this disclosure. With continued reference to FIG.4, at step 410, method 400 includes sampling symptomatic leaves from the plant. For the purposes of this disclosure, “sampling” is an action of taking samples from the leaves of plant to inspect whether they have produced plant virus- 30 based nanoparticles 104. This may happen after a certain time period subsequent to a successful infection step, as described above. Samples may be taken from leaves that are symptomatic. For 12 Attorney Docket No.1266-003PCT1 the purposes of this disclosure, “symptomatic” is a descriptor that describes one or more characteristics of an object that appear different due to viral infection. As a nonlimiting example, it may take three to six days for a CPMV-based nanoparticle and / or for a TBSV-based nanoparticle to form and accumulate in a plant. As a result, leaves may appear symptomatic in 5 several possible ways, such as without limitation, light and dark green patches or irregular mottling, a stunted, curled, or puckered appearance, and veins of leaves appearing lighter than normal or banded with dark green or yellow. Once plant virus-based nanoparticles 104 are detected, they may be extracted following steps described below. With continued reference to FIG.4, at step 415, method 400 includes homogenizing the 10 plant virus-based nanoparticle 104. For the purposes of this disclosure, “homogenizing” is a process of blending elements into a uniform mixture with a consistent or substantially consistent composition across its entirety. In one or more embodiments, a homogenization process may include combining a tissue of plant containing plant virus-based nanoparticles 104 with an extraction buffer solution. For the purposes of this disclosure, an “extraction buffer solution” is 15 an aqueous solution capable of breaking open cells and releasing elements therein. In one or more embodiments, extraction buffer solution may include salts to regulate its acidity. In the case of the TBSV, a sodium acetate solution may be used as extraction buffer solution. With continued reference to FIG.4, at step 420, method 400 includes incubating plant virus-based nanoparticle 104. A homogenous mixture of plant leaves and extraction buffer is 20 incubated in ice for a period of time. For the purposes of this disclosure, “incubating” is a process of subjecting an item to a hot or cold temperature for a certain period of time, until a certain goal is accomplished. With continued reference to FIG.4, in some cases, at step 425, method 400 may include centrifuging plant virus-based nanoparticle 104. For the purposes of this disclosure, 25 “centrifuging” is a process of separating multiple components in a mixture based on their difference in density by applying a centrifugal force to the mixture using a centrifuge device. For the purposes of this disclosure, a “centrifuge device” is a device comprising a rotating element attached to a stationary axis and configured to a spin sample under a high rotational speed in order to achieve separation between various elements therein. The amount of time to apply for 30 this centrifuging step may vary from one type of plant virus-based nanoparticle 104 to another. 13 Attorney Docket No.1266-003PCT1 In some cases, this step may be repeated multiple times to achieve an improved yield. As a nonlimiting example, for TBSV, centrifuging step may be repeated two or three times. With continued reference to FIG.4, at step 430, method 400 includes filtrating plant virus-based nanoparticle 104. In some cases, filtrating plant virus-based nanoparticle 104 may 5 include filtrating the plant virus-based nanoparticle 104 using tangential flow filtration (TFF), nanofiltration (NF), and / or gel-filtration chromatography, among other similar separation / purification techniques. For the purposes of this disclosure, tangential flow filtration (TFF), also known as cross-flow filtration, is a separation technique utilized to filter and concentrate biomolecules in solution. In TFF, a feed solution may flow tangentially across the 10 surface of a filter membrane, while an applied pressure forces some of the fluid through the membrane as filtrate (permeate). A tangential motion may help reduce membrane fouling and allow continuous filtration. TFF is commonly employed in bioprocessing for concentrating proteins, clarifying cell lysates, and purifying biopharmaceuticals, providing an efficient method for separating components based on size and molecular weight. For the purposes of this 15 disclosure, “nanofiltration (NF)” is a membrane filtration process that separates particles and solutes in the nanometer range, typically between 1 and 10 nanometers. NF may utilize a semi- permeable membrane to selectively allow certain molecules, such as monovalent ions and small organic molecules, to pass through while rejecting larger molecules, multivalent ions, and contaminants. NF may operate under moderate pressure and is commonly employed in water 20 treatment, pharmaceutical purification, and food processing, providing a means for removing impurities, softening water, and concentrating valuable substances with high efficiency. For the purposes of this disclosure, “gel-filtration chromatography” is a type of separation and purification technique based on a differing ability of chemical species to retain in pores of a gel- filtration medium. Gel-filtration chromatography is also known as size-exclusion 25 chromatography. A column used for gel-filtration chromatography may be packed with fine, porous beads composed of dextran polymers, agarose, polyacrylamide, and / or the like. The pore sizes of these beads are used to estimate the dimensions of macromolecules and separate them accordingly. Generally, chemical species of smaller sizes tend to retain in gel-filtration medium for a longer period of time (i.e., separates from a column later), whereas chemical species of 30 larger sizes tend to retain in gel-filtration medium for a shorter period of time (i.e., separates from a column earlier). The main application of gel-filtration chromatography is the fractionation 14 Attorney Docket No.1266-003PCT1 of proteins and other water-soluble polymers. Gel-filtration chromatography may be contrasted with gel permeation chromatography, which is a similar separation and purification technique often used to analyze the molecular weight distribution of organic-soluble polymers. Referring now to FIG.5, a flow diagram illustrating another exemplary embodiment of 5 method 400 for manufacturing plant virus-based nanoparticle 104 is illustrated. Plant virus-based nanoparticle 104 may include any type of plant virus-based nanoparticle described in this disclosure without limitation. At step 505, method 500 includes agroinfiltrating a plant with a virus to produce plant virus-based nanoparticle 104. For the purposes of this disclosure, “agroinfiltration” is a method used in plant biology to induce transient expression of genes in a 10 plant in order to produce a desired protein. As a nonlimiting example, agroinfiltration may be used for CPMV and / or for TBSV. As a nonlimiting example, to perform agroinfiltration, Agrobacterium tumefaciens may be directly injected into a plant leaf or brought into association with plant cells immobilized on a porous support. Subsequently, bacteria may transfer a desired gene into plant cells via transfer of T-DNA. Agroinfiltration may be beneficial when compared 15 to more traditional plant transformations due to its speed and convenience, although yields of the recombinant protein may generally also be higher and more consistent. Additionally, and / or alternatively, other production processes may also be used to produce plant virus-based nanoparticle 104. Once a plant has undergone such production processes, it may produce plant virus-based nanoparticles 104 to be used in composition 100. 20 With continued reference to FIG.5, at step 510, method 500 includes sampling leaves of the plant. This step may be performed utilizing any process of sampling consistent with details described above. With continued reference to FIG.5, at step 515, method 500 includes homogenizing the plant virus-based nanoparticle. This step may be performed utilizing any process of 25 homogenization as explained above. As a nonlimiting example, for CPMV and TBSV, a phosphate buffer or a PBS buffer may be used, consistent with details described above. With continued reference to FIG.5, at step 520, method 500 includes centrifuging the plant virus-based nanoparticle 104. This step may be performed utilizing any process of centrifugation as described above. It is worth noting that production of CPMV-based and TBSV- 30 based nanoparticles may not necessarily involve an incubation step described above for method 400. 15 Attorney Docket No.1266-003PCT1 With continued reference to FIG.5, at step 525, method 500 includes filtrating plant virus-based nanoparticle 104. This step may be performed utilizing any process of filtration as described above. Referring now to FIG.6, an exemplary embodiment of a method 600 of using plant virus- 5 based nanoparticle 104 for treating SLE is illustrated. Genetic sequences that encode tolerogenic peptide 108, such as hCDR1 (HGYYWSWIRQPPGKGEEWI, SEQ ID NO: 1) may be cloned into an expression vector harboring a viral genome or CP gene. For the purposes of this disclosure, an “expression vector” is typically a plasmid or virus designed for gene expression in cells. Expression vector is also known as expression construct. As nonlimiting examples, hCDR1 10 genes may be isolated from TBSV, an expression vector that is infective, and incorporated into CPMV, an expression vector that is non-infective. The obtained expression vector may then undergo either in vitro retro-transcription or an Agrobacterium tumefaciens transformation. For in vitro retro-transcription, plant may undergo infection using an infective RNA. For Agrobacterium tumefaciens transformation, plant may undergo agroinfiltration, as previously 15 described. Referring now to FIG.7, an exemplary embodiment of a production process 700 of a plant virus-based nanoparticle 104 is presented. Once plant virus-based nanoparticle 104 (pVNP) infects plant, the plant may start to produce genetically modified plant viruses. In other words, plant may function as a bioreactor. Upon detection of plant virus-based nanoparticles 104, 20 samples containing such plant virus-based nanoparticles 104 may be homogenized with extraction buffer solution, as described above. Upon homogenization, samples may be centrifuged and filtrated following procedures described above to yield plant virus-based nanoparticles 104, which may be used for downstream therapeutic applications, such as treatment of SLE. 25 The foregoing has been a detailed description of illustrative embodiments of the invention. Various modifications and additions can be made without departing from the spirit and scope of this invention. Features of each of the various embodiments described above may be combined with features of other described embodiments as appropriate in order to provide a multiplicity of feature combinations in associated new embodiments. Furthermore, while the 30 foregoing describes a number of separate embodiments, what has been described herein is merely illustrative of the application of the principles of the present invention. Additionally, 16 Attorney Docket No.1266-003PCT1 although particular methods herein may be illustrated and / or described as being performed in a specific order, the ordering is highly variable within ordinary skill to achieve embodiments as disclosed herein. Accordingly, this description is meant to be taken only by way of example, and not to otherwise limit the scope of this invention. 5 In the descriptions above and in the claims, phrases such as “at least one of” or “one or more of” may occur followed by a conjunctive list of elements or features. The term “and / or” may also occur in a list of two or more elements or features. Unless otherwise implicitly or explicitly contradicted by the context in which it is used, such a phrase is intended to mean any of the listed elements or features individually or any of the recited elements or features in 10 combination with any of the other recited elements or features. For example, the phrases “at least one of A and B;” “one or more of A and B;” and “A and / or B” are each intended to mean “A alone, B alone, or A and B together.” A similar interpretation is also intended for lists including three or more items. For example, the phrases “at least one of A, B, and C;” “one or more of A, B, and C;” and “A, B, and / or C” are each intended to mean “A alone, B alone, C alone, A and B 15 together, A and C together, B and C together, or A and B and C together.” In addition, use of the term “based on,” above and in the claims is intended to mean, “based at least in part on,” such that an unrecited feature or element is also permissible. The subject matter described herein can be embodied in systems, apparatus, methods, and / or articles depending on the desired configuration. The implementations set forth in the 20 foregoing description do not represent all implementations consistent with the subject matter described herein. Instead, they are merely some examples consistent with aspects related to the described subject matter. Although a few variations have been described in detail above, other modifications or additions are possible. In particular, further features and / or variations can be provided in addition to those set forth herein. For example, the implementations described above 25 can be directed to various combinations and sub-combinations of the disclosed features and / or combinations and sub-combinations of several further features disclosed above. In addition, the logic flows depicted in the accompanying figures and / or described herein do not necessarily require the particular order shown, or sequential order, to achieve desirable results. Other implementations may be within the scope of the following claims. 30 17 Attorney Docket No.1266-003PCT1

Claims

WHAT IS CLAIMED IS:

1. A composition for treating system lupus erythematosus (SLE), the composition comprising: 5 a plant virus-based nanoparticle engineered to express at least a tolerogenic peptide associated with SLE; and a buffer.

2. The composition of claim 1, wherein the buffer comprises one or more members selected from a list consisting of sodium acetate, acetate buffer, borate buffer, bicarbonate buffer, cacodylate buffer, Good’s buffer, phosphate buffer, bicarbonate buffer, and PBS buffer.

3. The composition of claim 1, wherein the at least a tolerogenic peptide includes hCDR1.

4. The composition of claim 1, wherein the at least a tolerogenic peptide includes a peptide associated with SLE.

5. The composition of claim 1, wherein the at least a tolerogenic peptide is engineered using a Tomato bushy stunt virus (TBSV).

6. The composition of claim 5, wherein the TBSV comprises 180 copies of a protein subunit.

7. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a Nicotiana benthamiana plant.

8. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a Nicotiana tabacum plant.

9. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a Lycopersicon esculentum plant.

10. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a Cycorium intybus plant.

11. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a Brassica oleracea var. capitata plant.

12. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a Beta vulgaris var cicla plant.

13. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from an Ocimum basilicum plant. 18 Attorney Docket No.1266-003PCT114. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a spinach plant.

15. The composition of claim 1, wherein the plant virus-based nanoparticle is sourced from a red beet plant. 5 16. The composition of claim 1, wherein the composition comprises an oral dosage form.

17. The composition of claim 1, wherein the composition comprises an intravenous dosage form.

18. The composition of claim 1, wherein the at least a tolerogenic peptide is fused to a protein of a TBSV.

19. The composition of claim 1, wherein the plant virus-based nanoparticle comprises a Cowpea mosaic virus (CPMV).

20. The composition of claim 19, wherein the CPMV comprises 60 copies of a protein subunit.

21. The composition of claim 20, wherein the protein subunit comprises a large coat protein.

22. The composition of claim 20, wherein the protein subunit comprises a small coat protein.

23. The composition of claim 19, wherein the CPMV comprises an icosahedral capsid.

24. A method of manufacturing a composition for treating SLE, wherein the method comprises: infecting a plant with a virus to produce a plant virus-based nanoparticle; sampling symptomatic leaves from the plant; homogenizing the plant virus-based nanoparticle; incubating the plant virus-based nanoparticle; centrifuging the plant virus-based nanoparticle; and filtrating the plant virus-based nanoparticle.

25. The method of claim 24, wherein filtrating the plant virus-based nanoparticle comprises filtrating the plant virus-based nanoparticle using tangential flow filtration (TFF), nanofiltration (NF), or gel-filtration chromatography.

26. A method of manufacturing a composition for treating SLE, wherein the method comprises: agroinfiltrating a plant with a virus to produce a plant virus-based nanoparticle; sampling symptomatic leaves of the plant; 19 Attorney Docket No.1266-003PCT1homogenizing the plant virus-based nanoparticle; centrifuging the plant virus-based nanoparticle; and filtrating the plant virus-based nanoparticle.

27. The method of claim 26, wherein filtering the plant virus-based nanoparticle comprises 5 filtrating the plant virus-based nanoparticle using tangential flow filtration (TFF), nanofiltration (NF), or gel-filtration chromatography.

28. A method of manufacturing a composition for treating SLE, wherein the method comprises: infecting a plant with a virus to produce a plant virus-based nanoparticle; sampling symptomatic leaves from the plant; homogenizing the plant virus-based nanoparticle; incubating the plant virus-based nanoparticle; and filtrating a solution comprising the incubated plant virus-based nanoparticle.

29. The method of claim 28, wherein filtering the solution comprises filtrating the solution using tangential flow filtration (TFF), nanofiltration (NF), or gel-filtration chromatography. 20 Attorney Docket No.1266-003PCT1

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