Methods of use for rhamm-derived peptides and rhamm-derived peptide mimetics

Abstract

The present invention provides methods for treating a disease or disorder associated with inflammation and / or with TLR signaling and / or with NLRP3 inflammasome activation in a subject. The methods involve administering to the subject a pharmaceutical composition comprising a RHAMM-derived peptide or a RHAMM-derived peptide mimetic.

Description

049648- / 618931 T19083WO001 METHODS OF USE FOR RHAMM-DERIVED PEPTIDES AND RHAMM-DERIVED PEPTIDE MIMETICS CROSS REFERENCE TO RELATED APPLICATION

[0001] This application claims the benefit of US Provisional Application No.63 / 582,799, filed September 14, 2023, which is incorporated by reference in its entirety for all purposes. REFERENCE TO A SEQUENCE LISTING

[0002] The application includes sequences in an electronic sequence listing named 618931SEQLST.XML of size 11.9 KB, created September 10, 2024, which is incorporated by reference. BACKGROUND OF THE INVENTION

[0003] Tissue injury results in the elaboration of endogenous danger signals that stimulate Toll-Like Receptors (TLRs). Invading pathogens are also sensed by the TLRs. TLR signaling activates several signaling pathways, including NFκB, and increases the expression of pro-IL1ß [1]. The second signal is the activation of the purinergic receptor P2X7 by extracellular ATP that results in the formation of the NLRP3 inflammasome, consisting of NLRP3, ASC, and pro- caspase1 [2,3], a response that has been demonstrated after hyperoxia exposure to adult mice [3] and in studies of neonatal mice exposed to hyperoxia [4]. Activation of this complex cleaves pro- caspase1 to caspase1 (p20), which in turn proteolytically cleaves pro-IL1ß to produce mature IL1ß (also referred to as IL-1ß). Released IL1ß interacts with its receptor, IL1R, to initiate and amplify inflammatory pathways. This system is a universal activator of the innate immune response to external challenges by foreign organisms as well as endogenous danger signals.

[0004] Hyaluronan (hyaluronic acid, HA), a glycosaminoglycan component of the extracellular matrix, is composed of repeating disaccharide units of glucuronic acid and N-acetyl glucosamine, and its accumulation as fragments is an early endogenous danger signal that mediates inflammation [5,6]. HA regulates inflammation in a dose- and molecular size- dependent manner through receptors, in particular RHAMM (also known as Hyaluronan mediated motility receptor) and Toll-like receptors (TLRs) [7]. It has been demonstrated that alveolar macrophages from bleomycin-injured animals are more motile than those from control animals and that a RHAMM peptide mimetic of the HA-binding region inhibits this increased049648- / 618931 T19083WO001 motility [8]. Further, systemic administration of this RHAMM peptide mimetic to bleomycin- injured animals resulted in decreased macrophage accumulation and fibrosis [8]. These data suggest that HA is upstream of and critical for the inflammatory response to lung injury.

[0005] Taking advantage of the known pro-inflammatory signals generated by HA fragment:HA receptor interactions, classes of peptide mimetics have been generated that dampen inflammation and control consequent fibrosis in animal models in the absence of apparent toxicity. Analysis of the HA-binding domains highlighted the helical structure of this sequence which provides a large surface area for interaction and is a requirement for protein:carbohydrate interactions [9]. Stapling either of the two domains using lactam bridges stabilizes the helix and increases the binding affinity of peptides [9]. A peptide mimetic (Peptide 152, of amino acid sequence SEQ ID NO:3 VSKLREQLVKRKQN, wherein glutamic acid (E) at position 6 is covalently bonded to lysine (K) at position 10, also referred to as Ac-(cyclo-6,10) – VSKLREQLVKRKQN-NH2) is reported that mimics the carboxy terminal HA-binding domain of RHAMM and that blocks macrophage chemotaxis to fetal bovine serum (FBS) or HA fragments (U.S. Patent 10,562,935 B2). SUMMARY OF THE INVENTION

[0006] In one aspect, the invention provides a method of treating or effecting prophylaxis of a disease or disorder associated with inflammation in a subject, comprising administering to the subject a pharmaceutical composition comprising a RHAMM-derived peptide mimetic comprising an amino acid sequence of SEQ ID NO:3, wherein glutamic acid (E) at position 6 is covalently bonded to lysine (K) at position 10.

[0007] In some methods, the disease or disorder associated with inflammation is selected from the group consisting of sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome, TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2-mediated pneumonia, and a disorder associated with TLR activation, activation of the NLRP3 inflammasome and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells.049648- / 618931 T19083WO001

[0008] In some methods, the disease or disorder associated with inflammation is sepsis. In some methods, the disease or disorder associated with inflammation is gram-negative pneumonia. In some methods, the disease or disorder associated with inflammation is acute lung injury. In some methods, the disease or disorder associated with inflammation is neonatal hyperoxia. In some methods, the disease or disorder associated with inflammation is bronchopulmonary dysplasia. In some methods, the disease or disorder associated with inflammation is acute respiratory distress syndrome. In some methods, the disease or disorder associated with inflammation is TLR7-mediated pneumonia. In some methods, the disease or disorder associated with inflammation is viral mediated inflammation. In some methods, the disease or disorder associated with inflammation is influenza A inflammation. In some methods, the disease or disorder associated with inflammation is influenza A pneumonia. In some methods, the disease or disorder associated with inflammation is COVID-19 pneumonia. In some methods, the disease or disorder associated with inflammation is SARS-CoV2 inflammation. In some methods, the disease or disorder associated with inflammation is SARS- CoV-2-mediated pneumonia.

[0009] In some methods, the disorder associated with TLR activation, activation of the NLRP3 inflammasome and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells is acute kidney injury, acute liver injury, immune-mediated nervous system injury, necrotizing enterocolitis, inflammatory bowel disease, gout, traumatic brain injury, white matter brain injury, and wound healing.

[0010] In some methods, the immune-mediated nervous system injury is multiple sclerosis. In some methods, the inflammatory bowel disease is Crohn’s disease or ulcerative colitis.

[0011] In another aspect, the invention provides a method of treating or effecting prophylaxis of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation in a subject, comprising administering to the subject a pharmaceutical composition comprising a RHAMM-derived peptide mimetic comprising an amino acid sequence of SEQ ID NO:3, wherein glutamic acid (E) at position 6 is covalently bonded to lysine (K) at position 10. In some methods, the NLRP3 inflammasome activation results in elevated hyaluronan concentrations and inflammation in the subject.049648- / 618931 T19083WO001

[0012] In some methods, the disease or disorder associated with TLR signaling or NLRP3 inflammasome activation is selected from the group consisting of sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome, TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2-mediated pneumonia, acute kidney injury, acute liver injury, immune-mediated nervous system injury, necrotizing enterocolitis, inflammatory bowel disease, gout, traumatic brain injury, white matter brain injury, and wound healing.

[0013] In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is sepsis. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is gram-negative pneumonia. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is acute lung injury. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is neonatal hyperoxia. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is bronchopulmonary dysplasia. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is acute respiratory distress syndrome. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is TLR7-mediated pneumonia. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is viral mediated inflammation. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is influenza A inflammation. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is influenza A pneumonia. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is COVID-19 pneumonia. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is SARS-CoV2 inflammation. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is SARS-CoV-2-mediated pneumonia.

[0014] In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is acute kidney injury. In some methods, the disease or disorder049648- / 618931 T19083WO001 associated with TLR signaling and / or NLRP3 inflammasome activation is acute liver injury. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is immune-mediated nervous system injury. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is necrotizing enterocolitis. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is inflammatory bowel disease. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is gout. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is traumatic brain injury. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is white matter brain injury. In some methods, the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is wound healing. In some methods, the immune- mediated nervous system injury is multiple sclerosis. In some methods, the inflammatory bowel disease is Crohn’s disease or ulcerative colitis.

[0015] In some methods, TLR signaling and / or NLRP3 inflammasome activation is blocked. In some methods, blockage of TLR signaling and / or NLRP3 inflammasome activation is measured by lack of production of pro-IL1ß, lack of production of mature IL1ß, inhibition of inflammation, and / or inhibition of cytokine elaboration. In some methods, TLR4 signaling and / or NLRP3 inflammasome activation is blocked. In some methods, TLR7 signaling and / or NLRP3 inflammasome activation is blocked.

[0016] In some methods, the subject is a mammal. In some methods, the mammal is a primate. In some methods, the mammal is a non-human primate. In some methods, the mammal is a human.

[0017] In some methods, the human is a neonate, child, or adult. In some methods, the human is a neonate. In some methods, the human is a child. In some methods, the human is an adult.

[0018] In some methods, the mammal is a rodent. In some methods, the rodent is a mouse, rat, guinea pig, hamster, or gerbil.049648- / 618931 T19083WO001

[0019] In some methods, the mammal is selected from the group consisting of human, baboon, chimpanzee, monkey, cynomolgus, marmoset, rhesus, mouse, rat, rabbit, cat, dog, horse, cow, sheep, goat, pig, ferret, guinea pig, hamster, gerbil, and bat. In some methods, the subject is a bird. In some methods, the bird is a chicken.

[0020] In some methods, the pharmaceutical composition is administered intravenously. In some methods, the pharmaceutical composition is administered subcutaneously. In some methods, the pharmaceutical composition is administered by inhalation.

[0021] In some methods, the pharmaceutical composition is administered at a dosage of about 0.0002 mg / kg of subject body weight to about 20 mg / kg of subject weight. In some methods, the pharmaceutical composition is administered at a dosage of about 0.02 mg / kg of subject body weight. In some methods, the pharmaceutical composition is administered at a dosage of about 0.2 mg / kg of subject body weight. In some methods, the pharmaceutical composition is administered at a dosage of about 20 mg / kg of subject body weight.

[0022] A further understanding of the nature and advantages of the present invention may be realized by reference to the remaining portions of the specification and claims. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 depicts the experimental design for the short- and long-term effects of neonatal hyperoxia exposure in mice.

[0024] Figures 2A-C depict results of studies showing that RHAMM KO mice do not activate the NLRP3 inflammasome pathway, where IL1β mRNA (2A), IL1β protein (2B), and caspase-1 (2C) are measured. Top panel of 2C depicts images of western blots cleaved caspase-1 and ß- actin levels, and bottom panel of 2C depicts plot of quantified data from top panel of 2C.

[0025] Figures 3A-C depict results of studies showing dose-dependent blockade of NLRP3 inflammasome activity and inflammation with SQ Peptide 152, where IL-1β protein (3A), MPO activity (3B) and NAG activity (3C) are measured.049648- / 618931 T19083WO001

[0026] Figures 4A-B depict results of studies showing hyperoxia decreases alveolarization and a single SQ dose of Peptide 152 rescues this outcome.4A: Microscopy images: 21% O2 and saline (top left); 21% O2and Peptide 152, 10 μmole / kg of subject body weight (top right); 95% O2 and saline (bottom left); 95% O2 and Peptide 152, 10 μmole / kg of subject body weight (bottom right). 4B: Plot of radial alveolar counts as a measure of alveolarization from images in 4A.

[0027] Figures 5A-D depict results of studies showing neonatal hyperoxia results in chronic activation of the NLRP3 inflammasome and inflammation. Peptide 152 given at 10 weeks of age inhibits this activation and inflammation. Lung IL1β mRNA (5A), lung IL1β protein (5B), MPO activity (5C), NAG activity (5D).

[0028] Figures 6A-D depict results of studies showing neonatal hyperoxia exposure results in exaggerated NLRP3 inflammasome and inflammation responses to acute lung injury at 10 weeks of age (increased lung IL1ß mRNA, protein, neutrophils, and macrophages). A single SQ dose of Peptide 152 given at the time of IN bleomycin blocked the inflammasome and inflammation responses. Lung IL1β mRNA (6A), lung IL1β protein (6B), MPO activity (6C), NAG activity (6D).

[0029] Figures 7A-B depicts results of studies in mouse models of LPS-induced systemic (intraperitoneal; IP) sepsis and pneumonia (intranasal).7A depicts results of dosage studies on effect of Peptide 152 on systemic effect of IP LPS. Figure 7B depicts results of flow cytometry studies on effect of Peptide 152 on lung effects of IN LPS.

[0030] Figures 8A-B depict results of studies in a non-infectious model of viral infection, where MPO activity (8A) and NAG activity (8B) in cells obtained by BAL over time are measured.

[0031] Figure 9A-C depict results of studies in a non-infectious model of viral infection, where IL1ß mRNA (9A) and IL1ß protein (9B).and IFN-γ protein (9C) are measured.

[0032] Figures 10A-E depict results of studies in a non-infectious model of viral infection, where HAS1 mRNA (10A), HAS2 mRNA (10B), HAS3 mRNA (10C), plasma HA content (10D), and BAL HA content (10E) are measured.049648- / 618931 T19083WO001

[0033] Figures 11A-C depict results of studies in a non-infectious model of viral infection, where caspase 1 activity (11A), plasma HA (11B), and BAL IL1ß (11C) are measured.

[0034] Figures 12A-B depict results of studies in a non-infectious model of viral infection, where MPO activity (12A) and NAG activity (12B) in lavage cells are measured.

[0035] Figures 13A-B depict results of studies in RHAMM WT and RHAMM KO mice where MPO activity (13A) and N-acetyl glucosaminidase activity (13B) are measured.

[0036] Figures 14A-B depict results of studies in a mouse model of influenza A viral pneumonia, where lung IL1ß protein with increasing viral dose (14A) and lung IL1ß protein over 72 hours after infection (14B) are measured.

[0037] Figures 15A-B depict results of studies in a mouse model of influenza A viral pneumonia, where HA concentrations in the BAL with varying dose (15A) and HA concentrations in the BAL over time after infection (15B) are measured.

[0038] Figures 16A-B depict results of studies in a mouse model of influenza A viral pneumonia, where MPO activity with varying dose (16A) and MPO activity over time (16B) are measured.

[0039] Figure 17 depicts results of studies in a mouse model of influenza A viral pneumonia, where MPO activity was measured. Peptide 152 was given SQ 48 hours after IN Influenza A in mice and lung lavage cells were examined for their MPO activity on day 7. Peptide 152 decreased neutrophil accumulation due to Influenza A viral infection.

[0040] Figures 18A-F depict results of studies in a mouse model of SARS-CoV-2 viral pneumonia, where TLR7 mRNA (18A), IL1ß mRNA (18B), IFNγ mRNA (18C) HAS2 mRNA (18D), TLR2 mRNA (18E), and Hmmr mRNA (18F) are measured. SARS-CoV-2 infection in K18-hACE2R mice results in increased TLR7, IL1ß, IFN γ, HAS2 mRNA, TLR2 mRNA, and Hmmr mRNA at 6 days after infection.

[0041] Figure 19A depicts microscopy images. Left panel (Uninfected); middle panel (Infected, PBS control); right panel (Infected, Peptide 152 on day 3). Figure 19B depicts lung score quantified from Figure 19A images.049648- / 618931 T19083WO001

[0042] Figures 20A-B depict results of time course studies after SARS-CoV-2 infection in K18-hACE2R mice. MPO activity (20A) and NAG activity (20B). BRIEF DESCRIPTION OF THE SEQUENCES

[0043] SEQ ID NO:1 sets forth the amino acid sequence of human RHAMM protein UniProt O75330-1.

[0044] SEQ ID NO:2 sets forth the amino acid sequence of mouse RHAMM protein UniProt Q00547-1.

[0045] SEQ ID NO:3 sets forth the amino acid sequence of RHAMM peptide mimetic P152.

[0046] SEQ ID NO:4 sets forth the amino acid sequence of human Toll-Like Receptor Protein 4 UniProt O00206.

[0047] SEQ ID NO:5 sets forth the amino acid sequence of mouse Toll-Like Receptor Protein 4 UniProt Q9QUK6.

[0048] SEQ ID NO:6 sets forth the amino acid sequence of human Toll-Like Receptor Protein 7 UniProt Q9NYK1.

[0049] SEQ ID NO:7 sets forth the amino acid sequence of mouse Toll-Like Receptor Protein 7 UniProt P58681. DEFINITIONS

[0050] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which this invention pertains. The following references provide one of skill with a general definition of many of the terms used in this invention: Academic Press Dictionary of Science and Technology, Morris (Ed.), Academic Press (1sted., 1992); Oxford Dictionary of Biochemistry and Molecular Biology, Smith et al. (Eds.), Oxford University Press (revised ed., 2000); Encyclopaedic Dictionary of Chemistry, Kumar (Ed.), Anmol Publications Pvt. Ltd. (2002); Dictionary of Microbiology and Molecular Biology, Singleton et al. (Eds.), John Wiley & Sons (3rded., 2002); Dictionary of Chemistry, Hunt (Ed.), Routledge (1sted., 1999); Dictionary of Pharmaceutical049648- / 618931 T19083WO001 Medicine, Nahler (Ed.), Springer-Verlag Telos (1994); Dictionary of Organic Chemistry, Kumar and Anand (Eds.), Anmol Publications Pvt. Ltd. (2002); and A Dictionary of Biology (Oxford Paperback Reference), Martin and Hine (Eds.), Oxford University Press (4thed., 2000). In addition, the following definitions are provided to assist the reader in the practice of the invention.

[0051] The terms “protein,” “polypeptide,” and “peptide,” used interchangeably herein, refer to polymeric forms of amino acids of any length, including coded and non-coded amino acids and chemically or biochemically modified or derivatized amino acids. The terms include natural full length proteins, fragments and synthetic peptides. The terms include polymers that have been modified, such as polypeptides having modified peptide backbones.

[0052] Proteins are said to have an “N-terminus” and a “C-terminus.” The term “N-terminus” relates to the start of a protein or polypeptide, terminated by an amino acid with a free amine group (-NH2). The term “C-terminus” relates to the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH).

[0053] The terms “nucleic acid” and “polynucleotide,” used interchangeably herein, refer to polymeric forms of nucleotides of any length, including ribonucleotides, deoxyribonucleotides, or analogs or modified versions thereof. The terms include single-, double-, and multi-stranded DNA or RNA, genomic DNA, cDNA, DNA-RNA hybrids, and polymers comprising purine bases, pyrimidine bases, or other natural, chemically modified, biochemically modified, non- natural, or derivatized nucleotide bases.

[0054] Nucleic acids are said to have “5’ ends” and “3’ ends” because mononucleotides are reacted to make oligonucleotides in a manner such that the 5’ phosphate of one mononucleotide pentose ring is attached to the 3’ oxygen of its neighbor in one direction via a phosphodiester linkage. An end of an oligonucleotide is referred to as the “5’ end” if its 5’ phosphate is not linked to the 3’ oxygen of a mononucleotide pentose ring. An end of an oligonucleotide is referred to as the “3’ end” if its 3’ oxygen is not linked to a 5’ phosphate of another mononucleotide pentose ring. A nucleic acid sequence, even if internal to a larger oligonucleotide, also may be said to have 5’ and 3’ ends. In either a linear or circular DNA molecule, discrete elements are referred to as being “upstream” or 5’ of the “downstream” or 3’ elements.049648- / 618931 T19083WO001

[0055] “Sequence identity” or “identity” in the context of two polynucleotide or polypeptide sequences makes reference to the residues in the two sequences that are the same when aligned for maximum correspondence over a specified comparison window. When percentage of sequence identity is used in reference to proteins it is recognized that residue positions which are not identical often differ by conservative amino acid substitutions, where amino acid residues are substituted for other amino acid residues with similar chemical properties (e.g., charge or hydrophobicity) and therefore do not change the functional properties of the molecule. When sequences differ in conservative substitutions, the percent sequence identity may be adjusted upwards to correct for the conservative nature of the substitution. Sequences that differ by such conservative substitutions are said to have “sequence similarity” or “similarity.” Means for making this adjustment are well known to those of skill in the art. Typically, this involves scoring a conservative substitution as a partial rather than a full mismatch, thereby increasing the percentage sequence identity. Thus, for example, where an identical amino acid is given a score of 1 and a non-conservative substitution is given a score of zero, a conservative substitution is given a score between zero and 1. The scoring of conservative substitutions is calculated, e.g., as implemented in the program PC / GENE (Intelligenetics, Mountain View, California).

[0056] “Percentage of sequence identity” refers to the value determined by comparing two optimally aligned sequences (greatest number of perfectly matched residues) over a comparison window, wherein the portion of the polynucleotide sequence or the polypeptide sequence in the comparison window may comprise additions or deletions (i.e., gaps) as compared to the reference sequence (which does not comprise additions or deletions) for optimal alignment of the two sequences. The percentage is calculated by determining the number of positions at which the identical nucleic acid base or amino acid residue occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison, and multiplying the result by 100 to yield the percentage of sequence identity. Unless otherwise specified (e.g., the shorter sequence includes a linked heterologous sequence), the comparison window is the full length of the shorter of the two sequences being compared.

[0057] Unless otherwise stated, sequence identity / similarity values refer to the value obtained using GAP Version 10 using the following parameters: % identity and % similarity for a nucleotide sequence using GAP Weight of 50 and Length Weight of 3, and the nwsgapdna.cmp049648- / 618931 T19083WO001 scoring matrix; % identity and % similarity for an amino acid sequence using GAP Weight of 8 and Length Weight of 2, and the BLOSUM62 scoring matrix; or any equivalent program thereof. “Equivalent program” includes any sequence comparison program that, for any two sequences in question, generates an alignment having identical nucleotide or amino acid residue matches and an identical percent sequence identity when compared to the corresponding alignment generated by GAP Version 10.

[0058] The term “conservative amino acid substitution” refers to the substitution of an amino acid that is normally present in the sequence with a different amino acid of similar size, charge, or polarity. Examples of conservative substitutions include the substitution of a non-polar (hydrophobic) residue such as isoleucine, valine, or leucine for another non-polar residue. Likewise, examples of conservative substitutions include the substitution of one polar (hydrophilic) residue for another such as between arginine and lysine, between glutamine and asparagine, or between glycine and serine. Additionally, the substitution of a basic residue such as lysine, arginine, or histidine for another, or the substitution of one acidic residue such as aspartic acid or glutamic acid for another acidic residue are additional examples of conservative substitutions. Examples of non-conservative substitutions include the substitution of a non-polar (hydrophobic) amino acid residue such as isoleucine, valine, leucine, alanine, or methionine for a polar (hydrophilic) residue such as cysteine, glutamine, glutamic acid or lysine and / or a polar residue for a non-polar residue. Typical amino acid categorizations are summarized below (Table 1).

[0059] Table 1. Amino Acid Categorizations Alanine Ala A Nonpolar Neutral 1.8 Arginine Arg R Polar Positive -4.5 Asparagine Asn N Polar Neutral -3.5 Aspartic acid Asp D Polar Negative -3.5 Cysteine Cys C Nonpolar Neutral 2.5 Glutamic acid Glu E Polar Negative -3.5 Glutamine Gln Q Polar Neutral -3.5 Glycine Gly G Nonpolar Neutral -0.4 Histidine His H Polar Positive -3.2 Isoleucine Ile I Nonpolar Neutral 4.5 Leucine Leu L Nonpolar Neutral 3.8 Lysine Lys K Polar Positive -3.9 Methionine Met M Nonpolar Neutral 1.9 Phenylalanine Phe F Nonpolar Neutral 2.8 Proline Pro P Nonpolar Neutral -1.6 Serine Ser S Polar Neutral -0.8 Threonine Thr T Polar Neutral -0.7 Tryptophan Trp W Nonpolar Neutral -0.9 Tyrosine Tyr Y Polar Neutral -1.3 Valine Val V Nonpolar Neutral 4.2

[0060] A “homologous” sequence (e.g., nucleic acid sequence or amino acid sequence) refers to a sequence that is either identical or substantially similar to a known reference sequence, such049648- / 618931 T19083WO001 that it is, for example, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the known reference sequence.

[0061] The term “fragment” when referring to a protein means a protein that is shorter or has fewer amino acids than the full-length protein. The term “fragment” when referring to a nucleic acid means a nucleic acid that is shorter or has fewer nucleotides than the full-length nucleic acid. A fragment can be, for example, an N-terminal fragment (i.e., removal of a portion of the C-terminal end of the protein), a C-terminal fragment (i.e., removal of a portion of the N- terminal end of the protein), or an internal fragment. A fragment can also be, for example, a functional fragment or an immunogenic fragment.

[0062] The term “variant” as used herein includes modifications, derivatives, or chemical equivalents of the amino acid and nucleic acid sequences disclosed herein that perform substantially the same function as the polypeptides or nucleic acid molecules disclosed herein in substantially the same way. In one embodiment, variants of polypeptides disclosed herein include, without limitation, conservative amino acid substitutions. Variants of polypeptides also include additions and deletions to the polypeptide sequences disclosed herein. In addition, variant nucleotide sequences and polypeptide sequences include analogs and derivatives thereof.

[0063] A "vector" is a replicon, such as plasmid, phage or cosmid, to which another polynucleotide segment may be attached so as to bring about the replication of the attached segment. Vectors capable of directing the expression of genes encoding for one or more polypeptides are referred to as "expression vectors".

[0064] The term "naked polynucleotide" refers to a polynucleotide not complexed with colloidal materials. Naked polynucleotides are sometimes cloned in a plasmid vector.

[0065] A "peptidomimetic" refers to a synthetic chemical compound which has substantially the same structural and / or functional characteristics of a peptide consisting of natural amino acids. The peptidomimetic can contain entirely synthetic, non-natural analogues of amino acids, or can be a chimeric molecule of partly natural peptide amino acids and partly non-natural analogs of amino acids. The peptidomimetic can also incorporate any amount of natural amino acid conservative substitutions as long as such substitutions also do not substantially alter the049648- / 618931 T19083WO001 mimetic's structure and / or inhibitory or binding activity. Polypeptide mimetic compositions can contain any combination of nonnatural structural components, which are typically from three structural groups: a) residue linkage groups other than the natural amide bond ("peptide bond") linkages; b) non-natural residues in place of naturally occurring amino acid residues; or c) residues which induce secondary structural mimicry, i.e., to induce or stabilize a secondary structure, e.g., a beta turn, gamma turn, beta sheet, alpha helix conformation, and the like.

[0066] A “RHAMM-derived peptide” is a peptide that comprises an amino acid sequence of a natural RHAMM polypeptide. Amino acid residues of a RHAMM-derived peptide may be modified to enhance activity of the RHAMM derived peptide. A “RHAMM-derived peptide mimetic” is a peptide that mimics the properties of a natural RHAMM polypeptide, such as charge based residues.

[0067] Exemplary natural RHAMM polypeptide sequences are provided as SEQ ID NO:1 and SEQ ID NO:2. An exemplary RHAMM-derived peptide and RHAMM-derived peptide mimetic is Peptide 152, of amino acid sequence SEQ ID NO:3 VSKLREQLVKRKQN, wherein glutamic acid (E) at position 6 is covalently bonded to lysine (K) at position 10.

[0068] “Binding affinity” is generally expressed in terms of equilibrium association or dissociation constants (KAor KD, respectively), which are in turn reciprocal ratios of dissociation and association rate constants (koff and kon, respectively). Thus, equivalent affinities may correspond to different rate constants, so long as the ratio of the rate constants remains the same.

[0069] As used herein, the term "binding specificity" refers to the selective affinity of one molecule for another such as the binding of a peptide to HA, receptors to ligands, and enzymes to substrates.

[0070] The term "specific binding" refers to binding between two molecules, for example, a ligand and a receptor, characterized by the ability of a molecule (ligand) to associate with another specific molecule (receptor) even in the presence of many other diverse molecules, i.e., to show preferential binding of one molecule for another in a heterogeneous mixture of molecules. Specific binding of a ligand to a receptor is also evidenced by reduced binding of a detectably labeled ligand to the receptor in the presence of excess unlabeled ligand (i.e., a binding competition assay).049648- / 618931 T19083WO001

[0071] The term “patient” includes human and other mammalian subjects that receive either prophylactic or therapeutic treatment.

[0072] The term "subject" refers to human and non-human animals (especially non-human mammals). The term "subject" is used herein, for example, in connection with therapeutic and diagnostic methods, to refer to human or animal subjects. Animal subjects include, but are not limited to, animal models, such as, mammalian models of conditions or disorders associated with inflammation or associated with TLR signaling and / or NLRP3 inflammasome activation such as sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome (ARDS), TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2-mediated pneumonia, any disorder associated with TLR activation, activation of the NLRP3 inflammasome, and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells, including but not limited to acute kidney injury, acute liver injury, immune-mediated nervous system injury (e.g., multiple sclerosis), necrotizing enterocolitis, inflammatory bowel disease (Crohn’s disease and ulcerative colitis), gout, traumatic brain injury, white matter brain injury, and wound healing.. Other specific examples of non-human subjects include, e.g., non-human primates, primates, baboons, chimpanzees, monkeys, cynomolgus, marmoset, rhesus, rodents (e.g., mice, rats, guinea pigs, hamsters, gerbils), rabbits, cats, dogs, horses, cows, sheep, goats, pigs, ferrets, bats, birds (e.g., chickens) etc.

[0073] The term "individual" or “subject” refers to any animal, including humans, non-human primates, primates, baboons, chimpanzees, monkeys, cynomolgus, marmoset, rhesus, rodents (e.g., mice, rats, guinea pigs, hamsters, gerbils), rabbits, cats, dogs, horses, cows, sheep, goats, pigs, ferrets, bats, birds (e.g., chickens) etc. Some individuals or subjects are mammals, including any animal classified as such, including humans, non-human primates, primates, baboons, chimpanzees, monkeys, cynomolgus, marmoset, rhesus, rodents (e.g., mice, rats, guinea pigs, hamsters, gerbils), rabbits, cats, dogs, horses, cows, sheep, goats, pigs, ferrets, and bats, etc.049648- / 618931 T19083WO001

[0074] The term “disease” refers to any abnormal condition that impairs physiological function. The term is used broadly to encompass any disorder, illness, abnormality, pathology, sickness, condition, or syndrome in which physiological function is impaired, irrespective of the nature of the etiology.

[0075] The term “symptom” refers to subjective evidence of a disease as perceived by the subject. A "sign" refers to objective evidence of a disease as observed by a physician.

[0076] Examples of conditions associated with inflammation include sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome (ARDS), TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2-mediated pneumonia, any disorder associated with TLR activation, activation of the NLRP3 inflammasome, and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells, including but not limited to acute kidney injury, acute liver injury, immune-mediated nervous system injury (e.g., multiple sclerosis), necrotizing enterocolitis, inflammatory bowel disease (Crohn’s disease and ulcerative colitis), gout, traumatic brain injury, white matter brain injury, and wound healing. Examples of a condition associated with TLR signaling and / or NLRP3 inflammasome activation include sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome (ARDS), TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, and SARS-CoV-2-mediated pneumonia. Examples of a condition associated with TLR4 signaling and / or NLRP3 inflammasome activation include sepsis gram-negative pneumonia, and conditions that have elevated HA and inflammation. Examples of conditions associated with TLR7 signaling and / or NLRP3 inflammasome activation are viral mediated inflammation, including but not limited to influenza A and SARS-CoV2 inflammation and pneumonia.

[0077] The terms "treat," "treating," "treatment," and "therapeutically effective" used herein do not necessarily imply 100% or complete treatment. Rather, there are varying degrees of treatment recognized by one of ordinary skill in the art as having a potential benefit or049648- / 618931 T19083WO001 therapeutic effect. In this respect, the inventive method can provide any amount of any level of treatment. Furthermore, the treatment provided by the inventive method can include the treatment of one or more conditions or symptoms of the disease being treated.

[0078] The terms "therapeutically effective dose," or "therapeutically effective amount," refer to that amount of a compound that results in prevention, delay of onset of symptoms, or amelioration of symptoms of inflammation or infectious disease. A therapeutically effective amount will, for example, be sufficient to treat, prevent, reduce the severity, delay the onset, or reduce the risk of occurrence of one or more symptoms of inflammation or infectious disease. The effective amount can be determined by methods well known in the art and as described in subsequent sections of this description.

[0079] The terms "treatment," "therapeutic method," and their cognates refer to treatment and prophylactic / preventative measures. Those in need of treatment may include individuals already having a particular medical disorder, as well as those who may ultimately acquire the disorder. The need for treatment is assessed, for example, by the presence of one or more risk factors associated with the development of a disorder, the presence or progression of a disorder, or likely receptiveness to treatment of a subject having the disorder. Treatment may include slowing or reversing the progression of a disorder.

[0080] Compositions or methods “comprising” or “including” one or more recited elements may include other elements not specifically recited. For example, a composition that “comprises” or “includes” a RHAMM-derived peptide may contain a RHAMM-derived peptide alone or in combination with other ingredients. For example, a composition that “comprises” or “includes” a RHAMM-derived peptide mimetic may contain a RHAMM-derived peptide mimetic alone or in combination with other ingredients. When the disclosure refers to a feature comprising specified elements, the disclosure should alternative be understood as referring to the feature consisting essentially of or consisting of the specified elements. Moreover, elements that are shown or described as being combined with other elements, can, in various embodiments, exist as stand-alone elements.

[0081] Designation of a range of values includes all integers within or defining the range, and all subranges defined by integers within the range.049648- / 618931 T19083WO001

[0082] Unless otherwise apparent from the context, the term “about” encompasses insubstantial variations, such as values within a standard margin of error of measurement (e.g., SEM) of a stated value. Unless otherwise apparent from the context, the term “about” encompasses values within ±5% or ±10% of a stated value.

[0083] The singular forms of the articles “a,” “an,” and “the” include plural references unless the context clearly dictates otherwise.

[0084] Abbreviations used herein include Myeloperoxidase (MPO), N-acetylglucosaminidase (NAG), Bronchopulmonary Dysplasia (BPD), Knockout (KO); bronchoalveolar lavage (BAL); hyaluronic acid (HA), HA Synthase 1 (HAS1), HA Synthase 2 (HAS2), HA Synthase 3 (HAS3); fetal bovine serum (FBS), subcutaneous (SQ); intraperitoneal (IP); intranasal (IN), lipopolysaccharide (LPS); acute respiratory distress syndrome (ARDS). DETAILED DESCRIPTION

[0085] I. Overview

[0086] The invention provides methods of treating or effecting prophylaxis of a disease or disorder associated with inflammation or of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation in a subject, using a RHAMM-derived peptide or a RHAMM-derived peptide mimetic. In some embodiments, the disease or disorder associated with inflammation or disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome (ARDS), TLR7- mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2-mediated pneumonia, any disorder associated with TLR activation , activation of the NLRP3 inflammasome, and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells, including but not limited to acute kidney injury, acute liver injury, immune- mediated nervous system injury (e.g., multiple sclerosis), necrotizing enterocolitis, inflammatory bowel disease (Crohn’s disease and ulcerative colitis), gout, traumatic brain injury, white matter brain injury, and wound healing. In some methods, the RHAMM-derived peptide or RHAMM- derived peptide mimetic is Peptide 152 (also referred to as P152, p.152, p152), of amino acid049648- / 618931 T19083WO001 sequence SEQ ID NO:3 (VSKLREQLVKRKQN, wherein E at position 6 is covalently bonded to K at position 10, also referred to as Ac-(cyclo-6,10) – VSKLREQLVKRKQN-NH2).

[0087] The invention provides methods of treating a disease or disorder associated with associated with inflammation or disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation, for example sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome (ARDS), TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2- mediated pneumonia, any disorder associated with TLR activation, activation of the NLRP3 inflammasome, and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells, including but not limited to acute kidney injury, acute liver injury, immune- mediated nervous system injury (e.g., multiple sclerosis), necrotizing enterocolitis, inflammatory bowel disease (Crohn’s disease and ulcerative colitis), gout, traumatic brain injury, white matter brain injury, and wound healing in a subject using a RHAMM-derived peptide or a RHAMM- derived peptide mimetic.

[0088] Peptide 152 at concentrations of nanomoles / kg of subject body weight is highly effective in blocking inflammation in a variety of proof-of-concept mouse injury models, including a neonatal mouse hyperoxia model of BPD, intraperitoneal LPS-induced systemic sepsis, intranasal LPS-induced gram-negative pneumonia and intranasal bleomycin-induced acute lung injury. Peptide 152 decreases the inflammatory response in infectious mouse models of Influenza A and SARS-CoV-2.

[0089] Peptide 152 has widespread activity in acute non-infectious and infectious models, including those that are mediated by TLR4 (for example but not limited to sepsis, gram negative pneumonia, conditions that have elevated HA and inflammation) and TLR7 (viral mediated inflammation, including but not limited to influenza A and SARS-CoV2 inflammation and pneumonia). Peptide 152 is useful in blocking all TLR signaling.

[0090] II. RHAMM049648- / 618931 T19083WO001

[0091] Receptor for HA-mediated motility (RHAMM, also referred to as Hyaluronan mediated motility receptor, HMMR, IHABP, CD168) is a receptor for hyaluronic acid (HA). RHAMM is an HA receptor expressed at the cell surface and intracellularly that regulates cell locomotion and proliferation (Zaman A. et al., Am J Respir Cell Mol Biol 33:447-454, 2005). The mature protein comprises 724 amino acids of which residues 1-22 are disordered, residues 40-81 are disordered, residues 365-546 are required for interaction with FAM83D, residues 635-645 are hyaluronic acid-binding, and residues 657-666 are hyaluronic acid-binding. An exemplary human RHAMM amino acid sequence is UniProt O75330-1, provided as SEQ ID NO:1. An exemplary mouse RHAMM amino acid sequence is UniProt Q00547-1, provided as SEQ ID NO:2.

[0092] Reference to RHAMM includes known natural variations which are listed in the UniProtKB / Swiss-Prot database and permutations thereof. Additionally, reference to RHAMM includes RHAMM with known post-translational modifications. Examples of known post- translational modifications are listed in the UniProtKB / Swiss-Prot database. Unless otherwise apparent from context, reference to RHAMM, or its fragments includes the natural human amino acid sequences including isoforms, mutants, and allelic variants thereof. Unless otherwise apparent from the context, reference to RHAMM means a natural human form of RHAMM including all isoforms, including soluble forms, irrespective of whether posttranslational modification (e.g., phosphorylation, glycation, or acetylation) is present.

[0093] III. RHAMM-derived peptides and RHAMM-derived peptide mimetics

[0094] Exemplary RHAMM-derived peptides and RHAMM-derived peptide mimetics are provided in U.S. Patent 10,562,935 B2, US Patent 8044022, US Patent Publication 20230135239 and in references [38-41]. For example, Peptide 152 is prepared as in U.S. Patent 10,562,935 B2.

[0095] IV. Toll-Like Receptors

[0096] Toll-like receptors are pattern recognition receptors (PRRs) that recognize specific molecular patterns. TLRs recognize pathogen-associated (PAMPs) or endogenous or “danger- associated” molecular patterns (DAMPs). PRRs are restricted to specific cellular compartments.049648- / 618931 T19083WO001 For example, TLR1, 2, 4, 5, 6, and 11 are located on the cell surface, and TLR3, 7, 8, 9, and 10 are in endosomes. Cell surface TLRs recognize bacterial components and extracellular DAMPs, and endosomal TLRs recognize single- and double-stranded RNA and CpG-DNA or intracellular DAMPs. (Garantziotis, S. and Savani, R. Am J. Physiol Cell Physiol 323:C202-C214, 2022).

[0097] A. Toll-like receptor 4

[0098] Human Toll-like receptor 4 (TLR4) is a single-pass type 1 membrane protein synthesized as an 839 amino acid protein, with residues 1-23 being a signal sequence that is cleaved in the mature protein. Residues 24-31 comprise the extracellular domain, residues 632- 652 comprise the transmembrane domain, and residues 653-839 comprise the cytoplasmic domain. An exemplary human TLR4 amino acid sequence is UniProt O00206 TLR4_HUMAN, provided as SEQ ID NO:4. An exemplary mouse TLR4 amino acid sequence is UniProt Q9QUK6, provided as SEQ ID NO:5.

[0099] Reference to TLR4 includes known natural variations which are listed in the UniProtKB / Swiss-Prot database and permutations thereof. Additionally, reference to TLR4 includes TLR4 with known post-translational modifications. Examples of known post- translational modifications are listed in the UniProtKB / Swiss-Prot database. Unless otherwise apparent from context, reference to TLR4, or its fragments includes the natural human amino acid sequences including isoforms, mutants, and allelic variants thereof. Unless otherwise apparent from the context, reference to TLR4 means a natural human form of TLR4 including all isoforms, including soluble forms, irrespective of whether posttranslational modification (e.g., phosphorylation, glycation, or acetylation) is present.

[0100] B. Toll-like receptor 7

[0101] Human Toll-like receptor 7 (TLR7) is a single-pass type 1 membrane protein synthesized as a 1049 amino acid protein, with residues 1-26 being a signal sequence that is cleaved in the mature protein. Residues 27-839 comprise the extracellular domain, residues 840- 860 comprise the transmembrane domain, and residues 861-1049 comprise the cytoplasmic domain. An exemplary human TLR7 amino acid sequence is UniProt Q9NYK1049648- / 618931 T19083WO001 TLR7_HUMAN, provided as SEQ ID NO:6. An exemplary mouse TLR7 amino acid sequence is UniProt P58681 ^ TLR7_MOUSE, provided as SEQ ID NO:7.

[0102] Reference to TLR7 includes known natural variations which are listed in the UniProtKB / Swiss-Prot database and permutations thereof. Additionally, reference to TLR7 includes TLR7 with known post-translational modifications. Examples of known post- translational modifications are listed in the UniProtKB / Swiss-Prot database. Unless otherwise apparent from context, reference to TLR7, or its fragments includes the natural human amino acid sequences including isoforms, mutants, and allelic variants thereof. Unless otherwise apparent from the context, reference to TLR7 means a natural human form of TLR7 including all isoforms, including soluble forms, irrespective of whether posttranslational modification (e.g., phosphorylation, glycation, or acetylation) is present.

[0103] V. Mouse Models of Human Disease

[0104] RHAMM-derived peptides or RHAMM-derived peptide mimetics are administered in mouse models of a disease or disorder associated with inflammation or of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation. Examples of mouse models are a neonatal mouse hyperoxia model of bronchopulmonary dysplasia (Examples 1 and 7), intraperitoneal lipopolysaccharide (LPS) mouse model of systemic sepsis (Examples 2 and 7), mouse model of intratracheal (IT) SM345405 (TLR agonist)-induced inflammation (Examples 3, 4, and 7), mouse model of intranasal (IN) Influenza A viral pneumonia (Examples 3, 5, and 7), and mouse model of SARS-CoV-2 viral pneumonia (Examples 3 and 6-7). Other mouse models to which RHAMM-derived peptides or RHAMM-derived peptide mimetics can be administered include any disorder associated with TLR activation, activation of the NLRP3 inflammasome, and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells, which include but are not limited to acute kidney injury, acute liver injury, immune-mediated nervous system injury (e.g., multiple sclerosis), necrotizing enterocolitis, inflammatory bowel disease (Crohn’s disease and ulcerative colitis), gout, traumatic brain injury, white matter brain injury, and wound healing.

[0105] VI. Therapeutic Methods049648- / 618931 T19083WO001

[0106] RHAMM-derived peptides or RHAMM-derived peptide mimetics can be used to treat or effect prophylaxis of a disease or disorder associated with inflammation or of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation in a subject. Examples of a disease or disorder associated with inflammation or disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation are sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome (ARDS), TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2-mediated pneumonia, and any disorder associated with TLR activation, activation of the NLRP3 inflammasome, and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells, including but not limited to acute kidney injury, acute liver injury, immune-mediated nervous system injury (e.g., multiple sclerosis), necrotizing enterocolitis, inflammatory bowel disease (Crohn’s disease and ulcerative colitis), gout, traumatic brain injury, white matter brain injury, and wound healing. RHAMM- derived peptides or RHAMM-derived peptide mimetics can be used to treat or effect prophylaxis of sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome (ARDS), TLR7-mediated pneumonia, viral- mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2-mediated pneumonia, and any disorder associated with TLR activation, activation of the NLRP3 inflammasome, and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells, including but not limited to acute kidney injury, acute liver injury, immune-mediated nervous system injury (e.g., multiple sclerosis), necrotizing enterocolitis, inflammatory bowel disease (Crohn’s disease and ulcerative colitis), gout, traumatic brain injury, white matter brain injury, and wound healing. In some methods, blockage of TLR signaling and / or NLRP3 inflammasome activation is measured by lack of production of pro-IL1ß, lack of production of mature IL1ß, inhibition of inflammation, and / or inhibition of cytokine elaboration. In some methods, blockade of TLR4 signaling and / or NLRP3 inflammasome activation is measured by lack of production of pro- IL1ß, lack of production of mature IL1ß, inhibition of inflammation, and / or inhibition of cytokine elaboration. In some methods, blockade of TLR7 signaling and / or NLRP3 inflammasome activation is measured by lack of production of pro-IL1ß, lack of production of049648- / 618931 T19083WO001 mature IL1ß, inhibition of inflammation, and / or inhibition of cytokine elaboration. In some methods, a RHAMM-derived peptide or RHAMM-derived peptide mimetic blocks LPS-induced sepsis and pneumonia, as well as influenza A and SARS-CoV-2-mediated pneumonia (TLR7). In some methods, Peptide 152 blocks LPS-induced sepsis and pneumonia, as well as influenza A and SARS-CoV-2-mediated pneumonia (TLR7).

[0107] RHAMM-derived peptides or RHAMM-derived peptide mimetics can be used to treat a subject infected with a virus, bacterium, fungus, worm, or protozoan. In some embodiments, the subject is infected with SARS-CoV-2, influenza A, or a gram negative bacteria. In some methods, the RHAMM-derived peptide or RHAMM-derived peptide mimetic is Peptide 152.

[0108] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 0.0002 mg / kg to about 20 mg / kg of subject body weight.

[0109] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 0.0002 mg / kg to about 0.002 mg / kg of subject body weight. In some methods, the dose is about 0.0002 mg / kg, about 0.0003 mg / kg, about 0.0004 mg / kg, about 0.0005 mg / kg, about 0.0006 mg / kg, about 0.0007 mg / kg, about 0.0008 mg / kg, about 0.0009 mg / kg, about 0.001 mg / kg, about 0.0011 mg / kg, about 0.0012 mg / kg, about 0.0013 mg / kg, about 0.0014 mg / kg, about 0.0015 mg / kg, about 0.0016 mg / kg, about 0.0017 mg / kg, about 0.0018 mg / kg, about 0.0019 mg / kg, or about 0.002 mg / kg of subject body weight.

[0110] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 0.002 mg / kg to about 0.02 mg / kg of subject body weight. In some methods, the dose is about 0.002 mg / kg, about 0.003 mg / kg, about 0.004 mg / kg, about 0.005 mg / kg, about 0.006 mg / kg, about 0.007 mg / kg, about 0.008 mg / kg, about 0.009 mg / kg, about 0.01 mg / kg, about 0.011 mg / kg, about 0.012 mg / kg, about 0.013 mg / kg, about 0.014 mg / kg, about 0.015 mg / kg, about 0.016 mg / kg, about 0.017 mg / kg, about 0.018 mg / kg, about 0.019 mg / kg, or about 0.02 mg / kg of subject body weight.

[0111] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 0.02 mg / kg to about 0.2 mg / kg of subject body weight. In some methods, the dose is about 0.02 mg / kg, about 0.03 mg / kg, about 0.04 mg / kg, about 0.05049648- / 618931 T19083WO001 mg / kg, about 0.06 mg / kg, about 0.07 mg / kg, about 0.08 mg / kg, about 0.09 mg / kg, about 0.1 mg / kg, about 0.11 mg / kg, about 0.12 mg / kg, about 0.13 mg / kg, about 0.14 mg / kg, about 0.15 mg / kg, about 0.16 mg / kg, about 0.17 mg / kg, about 0.18 mg / kg, about 0.19 mg / kg, or about 0.2 mg / kg of subject body weight.

[0112] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 0.2 mg / kg to about 2.0 mg / kg of subject body weight. In some methods, the dose is about 0.2 mg / kg, about 0.3 mg / kg, about 0.4 mg / kg, about 0.5 mg / kg, about 0.6 mg / kg, about 0.7 mg / kg, about 0.8 mg / kg, about 0.9 mg / kg, about 1.0 mg / kg, about 1.1 mg / kg, about 1.2 mg / kg, about 1.3 mg / kg, about 1.4 mg / kg, about 1.5 mg / kg, about 1.6 mg / kg, about 1.7 mg / kg, about 1.8 mg / kg, about 1.9 mg / kg, or about 2.0 mg / kg of subject body weight.

[0113] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 2 mg / kg to about 20 mg / kg of subject body weight. In some methods, the dose is about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 8 mg / kg, about 9 mg / kg, about 10 mg / kg, about 11 mg / kg, about 12 mg / kg, about 13 mg / kg, about 14 mg / kg, about 15 mg / kg, about 16 mg / kg, about 17 mg / kg, about 18 mg / kg, about 19 mg / kg, or about 20 mg / kg of subject body weight.

[0114] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 0.1 nmole / kg to about 10,000 nmole / kg of subject body weight.

[0115] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 0.1 nmole / kg to about 1.0 nmole / kg of subject body weight. In some methods, the dose is about 0.1 nmole / kg, about 0.2 nmole / kg, about 0.3 nmole / kg, about 0.4 nmole / kg, about 0.5 nmole / kg, about 0.6 nmole / kg, about 0.7 nmole / kg, about 0.8 nmole / kg, about 0.9 nmole / kg, or about 1.0 nmole / kg of subject body weight.

[0116] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 1 nmole / kg to about 10 nmole / kg of subject body weight. In some methods, the dose is about 1 nmole / kg, about 2 nmole / kg, about 3 nmole / kg, about 4049648- / 618931 T19083WO001 nmole / kg, about 5 nmole / kg, about 6 nmole / kg, about 7 nmole / kg, about 8 nmole / kg, about 9 nmole / kg, or about 10 nmole / kg of subject body weight.

[0117] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 10 nmole / kg to about 100 nmole / kg of subject body weight. In some methods, the dose is about 10 nmole / kg, about 20 nmole / kg, about 30 nmole / kg, about 40 nmole / kg, about 50 nmole / kg, about 60 nmole / kg, about 70 nmole / kg, about 80 nmole / kg, about 90 nmole / kg, or about 100 nmole / kg of subject body weight.

[0118] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 100 nmole / kg to about 1000 nmole / kg of subject body weight. In some methods, the dose is about 100 nmole / kg, about 200 nmole / kg, about 300 nmole / kg, about 400 nmole / kg, about 500 nmole / kg, about 600 nmole / kg, about 700 nmole / kg, about 800 nmole / kg, about 900 nmole / kg, or about 1000 nmole / kg of subject body weight.

[0119] In some methods, a RHAMM-derived peptide or a RHAMM-derived peptide mimetic is administered at a dose of about 1000 nmole / kg to about 10,000 nmole / kg of subject body weight. In some methods, the dose is about 1000 nmole / kg, about 2000 nmole / kg, about 3000 nmole / kg, about 4000 nmole / kg, about 5000 nmole / kg, about 6000 nmole / kg, about 7000 nmole / kg, about 8000 nmole / kg, about 9000 nmole / kg, or about 10,000 nmole / kg of subject body weight.

[0120] It will be appreciated that dosages or amounts incrementally between those described above are within the scope of the present disclosure.

[0121] Exemplary timing of dosages being administered to a subject includes once every 12 hours, once every day, once every 2 days, once every 3 days, once every 4 days, once every 5 days, once every 6 days, once every 7 days, once every week, once every month, once 12 months, once a year, or according to any other schedule determined by empirical analysis. Alternatively, administration of a RHAMM-derived peptide or a RHAMM-derived peptide mimetic can be on an irregular basis as indicated by monitoring of symptoms of a disease or disorder associated with inflammation or of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation.049648- / 618931 T19083WO001

[0122] In some methods of treating a disease or disorder associated with inflammation or of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation in a subject, the subject is administered multiple doses of the RHAMM-derived peptide or RHAMM- derived peptide mimetic. In some methods of treating a viral illness in a subject, the subject is administered multiple doses of the RHAMM-derived peptide or RHAMM-derived peptide mimetic. In some methods, the multiple doses are started as early as six hours after infection, and continuing until at least 96 hours after infection.

[0123] Exemplary dosages in units of mg / kg of subject body weight, nmoles / kg of subject body weight, and μmoles / kg of subject body weight are: 0.0002 mg / kg = 0.1 nmoles / kg 0.002 mg / kg = 1 nmole / kg 0.02 mg / kg = 10 nmole / kg 0.2 mg / kg = 100 nmole / kg 2 mg / kg = 1 µmole / kg (1,000 nmole / kg) 6 mg / kg = 3 µmole / kg (3,000 nmole / kg) 20 mg / kg = 10 µmole / kg (10,000 nmole / kg)

[0124] An exemplary dosage is 0.02 mg / kg of subject body weight. An exemplary dosage is 0.2 mg / kg of subject body weight. An exemplary dosage is 20 mg / kg of subject body weight.

[0125] Exemplary dosage of P152 in a human neonate is 0.2 mg / kg of subject body weight. Exemplary dosage for treating a disease that involves severe inflammation (such as SARS-CoV- 2 or ARDS) in children and adults is 20 mg / kg of subject body weight. For example, an exemplary dosage for treating a disease that involves severe inflammation (such as SARS-CoV-2 or ARDS) in both children and adults is administered in 1-3 dosages over the first seven days after clinical presentation, where each dose is 20 mg / kg of subject body weight.

[0126] The RHAMM-derived peptide or RHAMM-derived peptide mimetic can be administered in combination with another a second therapeutic agent effective in treatment or049648- / 618931 T19083WO001 prophylaxis of a disease or disorder associated with inflammation or of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation before, during or after the administration of RHAMM-derived peptide to the subject. The RHAMM-derived peptide or RHAMM-derived peptide mimetic may be co-administered with one or more other suitable second therapeutic agents, or one or more such second therapeutic agents may be incorporated into the RHAMM-derived peptide or RHAMM-derived peptide mimetic. Exemplary second therapeutic agents include, but not limited to high molecular weight hyaluronan, corticosteroids (systemic or topical), anti-viral agents, anti-bacterial agents, anti-fungal agents, anti-worm agents, anti-protozoan agents, cytotoxic agents, and gene therapy vectors or constructs.

[0127] In some methods, a RHAMM-derived peptide or RHAMM-derived peptide mimetic is administered in combination with or is conjugated to a cytotoxic agent.

[0128] A gene therapy vector or construct co-administered with a RHAMM-derived peptide or RHAMM-derived peptide mimetic may comprise a polynucleotide encoding a polypeptide (and optionally a carrier fused to the polypeptide). The polypeptide encoded by the polynucleotide is expressed in situ in a patient. Such polynucleotides can be DNA or RNA. A polynucleotide segment encoding a polypeptide is typically linked to regulatory elements, such as a promoter and enhancer that allow expression of the DNA segment in the intended target cells of a patient. For expression in blood cells, promoter and enhancer elements from light or heavy chain immunoglobulin genes or the CMV major intermediate early promoter and enhancer are suitable to direct expression. The linked regulatory elements and coding sequences are often cloned into a vector.

[0129] The polynucleotide can be delivered in naked form (i.e., without colloidal or encapsulating materials). Alternatively, a number of viral vector systems can be used including retroviral systems (see, e.g., Lawrie and Tumin, Cur. Opin. Genet. Develop.3, 102-109 (1993)); adenoviral vectors {see, e.g., Bett et al, J. Virol.67, 5911 (1993)) including retrovirus derived vectors such MMLV, HIV-1, and ALV; adeno-associated virus vectors {see, e.g., Zhou et al., J. Exp. Med.179, 1867 (1994)), lentiviral vectors such as those based on HIV or FIV gag sequences, viral vectors from the pox family including vaccinia virus and the avian pox viruses, viral vectors from the alpha virus genus such as those derived from Sindbis and Semliki Forest049648- / 618931 T19083WO001 Viruses (see, e.g., Dubensky et al., J. Virol.70, 508-519 (1996)), Venezuelan equine encephalitis virus (see US 5,643,576) and rhabdoviruses, such as vesicular stomatitis virus (see WO 96 / 34625) and papillomaviruses (Ohe et al., Human Gene Therapy 6, 325-333 (1995); Woo et al, WO 94 / 12629 and Xiao & Brandsma, Nucleic Acids. Res.24, 2630-2622 (1996)).

[0130] A polynucleotide encoding a polypeptide, or a vector containing the same, can be packaged into liposomes. Suitable lipids and related analogs are described by US 5,208,036, US 5,264,618, US 5,279,833, and US 5,283,185. Vectors, DNA, or RNA encoding an immunogen, can also be adsorbed to or associated with particulate carriers, examples of which include polymethyl methacrylate polymers and polylactides and poly(lactide-co-glycolides), (see, e.g., McGee et al., J. Micro Encap.1996).

[0131] In pharmacology, a route of administration is the path by which a drug, fluid, or other substance is brought into contact with the body. In general, methods of administering drugs and nucleic acids for treatment of a mammal are well known in the art and can be applied to administration of the described RHAMM-derived peptides and RHAMM-derived peptide mimetics and compositions. The described RHAMM-derived peptides and RHAMM-derived peptide mimetics and compositions can be administered via any suitable route in a preparation appropriately tailored to that route. In general, any suitable method recognized in the art for delivering a therapeutic drug, protein, or nucleic acid can be adapted for use with a herein described RHAMM-derived peptides and RHAMM-derived peptide mimetics and compositions. Routes of administration include, but are not limited to, parenteral, local, direct injection, intraparenchymal, intratumoral, intramuscular, implantation, topical, systemic, intravascular, intravenous, intra-arterial, intraventricular, intralymphatic, transdermal, intracutaneous, intradermal, subdermal, subcutaneous, intraperitoneal, intracranial, subdural, intrathecal, epidural, rectal, airway (aerosol, inhalation), nasal, oral, buccal (mouth / cheek), and sublingual (under the tongue) administration. In some embodiments, the compositions are administered by subcutaneous or intravenous infusion or injection.

[0132] Pharmaceutical compositions for parenteral administration are preferably sterile and substantially isotonic and manufactured under GMP conditions. Pharmaceutical compositions can be provided in unit dosage form (i.e., the dosage for a single administration).049648- / 618931 T19083WO001 Pharmaceutical compositions can be formulated using one or more physiologically acceptable carriers, diluents, excipients, or auxiliaries. The formulation depends on the route of administration chosen.

[0133] Unless otherwise specified herein, the materials and reagents required for practicing the invention, e.g., the RHAMM-derived peptides or RHAMM-derived peptide mimetics, as well as the related therapeutic methods, can all be generated or performed in accordance with the procedures exemplified herein or routinely practiced protocols well known in the art. See, e.g., Methods in Enzymology, Volume 289: Solid-Phase Peptide Synthesis, J. N. Abelson, M. I. Simon, G. B. Fields (Editors), Academic Press; 1st edition (1997) (ISBN-13: 978-0121821906); U.S. Pat. Nos.4,965,343, and 5,849,954; Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press, N.Y., (3rded., 2000); Brent et al., Current Protocols in Molecular Biology, John Wiley & Sons, Inc. (Ringbou ed., 2003); Davis et al., Basic Methods in Molecular Biology, Elsevier Science Publishing, Inc., New York, USA (1986); or Methods in Enzymology: Guide to Molecular Cloning Techniques Vol.152, S. L. Berger and A. R. Kimmerl Eds., Academic Press Inc., San Diego, USA (1987); Current Protocols in Protein Science (CPPS) (John E. Coligan, et. al., ed., John Wiley and Sons, Inc.), Current Protocols in Cell Biology (CPCB) (Juan S. Bonifacino et. al. ed., John Wiley and Sons, Inc.), and Culture of Animal Cells: A Manual of Basic Technique by R. Ian Freshney, Publisher: Wiley-Liss; 5th edition (2005), Animal Cell Culture Methods (Methods in Cell Biology, Vol.57, Jennie P. Mather and David Barnes editors, Academic Press, 1st edition, 1998). The following sections provide additional guidance for practicing the compositions and methods of the present invention.

[0134] In some embodiments, compositions useful in methods of the invention contain a carrier for the RHAMM-derived peptide or RHAMM-derived peptide mimetic, desirably a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier can be any suitable pharmaceutically acceptable carrier. It can be one or more compatible solid or liquid fillers, diluents, other excipients, or encapsulating substances which are suitable for administration into a human or veterinary patient (e.g., a physiologically acceptable carrier or a pharmacologically acceptable carrier). The term "carrier" denotes an organic or inorganic ingredient, natural or synthetic, with which the active ingredient is combined to facilitate the use of the active ingredient, e.g., the administration of the active ingredient to a subject. The pharmaceutically049648- / 618931 T19083WO001 acceptable carrier can be co-mingled with one or more of the active components, e.g., a hybrid molecule, and with each other, when more than one pharmaceutically acceptable carrier is present in the composition, in a manner so as not to substantially impair the desired pharmaceutical efficacy. Pharmaceutically acceptable materials typically are capable of administration to a subject, e.g., a patient, without the production of significant undesirable physiological effects such as nausea, dizziness, rash, or gastric upset. It is, for example, desirable for a composition comprising a pharmaceutically acceptable carrier not to be immunogenic when administered to a human patient for therapeutic purposes.

[0135] Pharmaceutical compositions useful in methods of the invention can additionally contain suitable buffering agents, including, for example, acetic acid in a salt, citric acid in a salt, boric acid in a salt, and phosphoric acid in a salt. The compositions can also optionally contain suitable preservatives, such as benzalkonium chloride, chlorobutanol, parabens, and thimerosal. Pharmaceutical compositions useful in methods of the invention can be presented in unit dosage form and can be prepared by any suitable method, many of which are well-known in the art of pharmacy. Such methods include the step of bringing the RHAMM-derived peptide or RHAMM-derived peptide mimetic into association with a carrier that constitutes one or more accessory ingredients. In general, the composition is prepared by uniformly and intimately bringing the active agent into association with a liquid carrier, a finely divided solid carrier, or both, and then, if necessary, shaping the product.

[0136] A composition suitable for parenteral administration conveniently comprises a sterile aqueous preparation of the RHAMM-derived peptide or RHAMM-derived peptide mimetic, which preferably is isotonic with the blood of the recipient. This aqueous preparation can be formulated according to known methods using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation also can be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1,3-butane diol. Among the acceptable vehicles and solvents that can be employed are water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil can be employed, such as synthetic mono-or di-glycerides. In addition, fatty acids such as oleic acid can be used in the preparation of injectables. Carrier formulations suitable for oral,049648- / 618931 T19083WO001 subcutaneous, intravenous, intramuscular, etc. administrations can be found in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA.

[0137] Preparation of pharmaceutical compositions useful in methods of the invention and their various routes of administration can be carried out in accordance with methods well known in the art. See, e.g., Remington: The Science and Practice of Pharmacy, Mack Publishing Co., 20thed., 2000; and Sustained and Controlled Release Drug Delivery Systems, J.R. Robinson, ed., Marcel Dekker, Inc., New York, 1978. The delivery systems useful in the context of the invention include time-released, delayed release, and sustained release delivery systems such that the delivery of the inventive composition occurs prior to, and with sufficient time to cause, sensitization of the site to be treated. The inventive composition can be used in conjunction with other therapeutic agents or therapies. Such systems can avoid repeated administrations of the inventive composition, thereby increasing convenience to the subject and the physician, and may be particularly suitable for certain compositions useful in methods of the invention.

[0138] Many types of release delivery systems are available and known to those of ordinary skill in the art. Suitable release delivery systems include polymer base systems such as poly(lactide-glycolide), copolyoxalates, polycaprolactones, polyesteramides, polyorthoesters, polyhydroxybutyric acid, and polyanhydrides. Microcapsules of the foregoing polymers containing drugs are described in, for example, U.S. Patent 5,075,109. Delivery systems also include non-polymer systems that are lipids including sterols such as cholesterol, cholesterol esters, and fatty acids or neutral fats such as mono-di-and triglycerides; hydrogel release systems; sylastic systems; peptide based systems; wax coatings; compressed tablets using conventional binders and excipients; partially fused implants; and the like. Specific examples include, but are not limited to: (a) erosional systems in which the active composition is contained in a form within a matrix such as those described in U.S. Patents 4,452,775, 4,667,014, 4,748,034, and 5,239,660 and (b) diffusional systems in which an active component permeates at a controlled rate from a polymer such as described in U.S. Patents 3,832,253 and 3,854,480. In addition, pump-based hardware delivery systems can be used, some of which are adapted for implantation.

[0139] VII. Kits049648- / 618931 T19083WO001

[0140] The invention also provides kits suitable for carrying out the methods of the invention. Typically, the kits contain two or more components required for performing the therapeutic methods of the invention. Kit components include, but are not limited to, one or more RHAMM- derived peptides or RHAMM-derived peptide mimetics useful in methods of the invention, appropriate reagents, and / or equipment. Some kits include a second therapeutic agent effective in treatment or prophylaxis of a disease or disorder associated with inflammation or of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation. Exemplary second therapeutic agents include high molecular weight hyaluronan, corticosteroids (systemic or topical), anti-viral agents, anti-bacterial agents, anti-fungal agents, anti-worm agents, anti- protozoan agents, cytotoxic agents, and gene therapy vectors or constructs. Some kits include a second therapeutic agent incorporated into the RHAMM-derived peptide or RHAMM-derived peptide mimetic. Some kits include a RHAMM-derived peptide or RHAMM-derived peptide mimetic conjugated to a cytotoxic agent.

[0141] Generally, the RHAMM-derived peptide or RHAMM-derived peptide mimetic in a kit is suitably packaged, e.g., in a vial, pouch, ampoule, and / or any container appropriate for a therapeutic method. Kit components can be provided as concentrates (including lyophilized compositions), which may be further diluted prior to use, or they can be provided at the concentration of use. For use of the RHAMM-derived peptide or RHAMM-derived peptide mimetic in vivo, single dosages may be provided in sterilized containers having the desired amount and concentration of components. EXAMPLES

[0142] The following examples are provided to further illustrate the invention but not to limit its scope. Other variants of the invention will be readily apparent to one of ordinary skill in the art and are encompassed by the appended claims.

[0143] Example 1. Neonatal Hyperoxia Model of Bronchopulmonary Dysplasia (BPD)

[0144] Bronchopulmonary Dysplasia (BPD), a devastating pulmonary complication in preterm infants with respiratory failure [10-13], develops in 30-50% of infants less than 1000-gram birth weight and is thought to involve ventilator- and oxygen-induced damage to an immature lung049648- / 618931 T19083WO001 that results in an inflammatory response and ends in aberrant lung development with decreased alveolarization [14,15]. Approximately12,000-15,000 infants develop BPD each year in the United States. In addition to the direct impacts on the infant, and the clear toll that this disease takes on families, healthcare expenditure has been estimated at over $5B per year

[0013] . Additionally, infants with BPD have long-term morbidity and exaggerated responses to lung injury into adolescence and adulthood [16-18]. Thus, therapies that decrease the incidence and / or severity of this condition would have significant impact on morbidity, mortality, human costs, healthcare expenditure, and long-term outcomes. BPD is a complex medical condition with a multifactorial pathogenesis. Key contributors include oxygen toxicity, ventilator-induced lung injury, inflammation, and an arrest of lung vascular development [19,20]. An inflammatory response and the elaboration of growth factors and cytokines are associated with the development of BPD [21,22]. The innate immune system is an early, non-specific, yet robust system that senses endogenous danger signals and mobilizes inflammatory cells as a response. This system has been extensively characterized in both rodent and non-human primate models of BPD, as well as in tracheal aspirates from preterm infants with respiratory failure [4]. The regulation of the innate immune system by hyaluronan (hyaluronic acid, HA) has been examined. (HA) fragments have been identified as an endogenous danger signal that interact with HA receptors such as RHAMM (Receptor for HA-Mediated Motility, CD168) and cooperate with Toll-Like Receptors (TLRs) to activate the innate immune system to produce IL1ß, a key regulator of inflammation [23,24].

[0145] Neonatal mice are born in the saccular stage of lung development and exposure to hyperoxia during the initial five days of life mimics what preterm infants are subjected to in the Neonatal Intensive Care Unit. This model has been used to demonstrate that the NLRP3 inflammasome is critical to the development of BPD (Liao, J. et al., Nat Comm, 2015 Nov 27;6:897).

[0146] This model has been extended to determine the long-term effects of neonatal hyperoxia exposure at 8-10 weeks of age (young adulthood). Figure 1 shows the experimental design.

[0147] The importance of RHAMM in this model was established by using RHAMM KO mice. Figures 2A-C show that RHAMM KO mice do not activate the NLRP3 inflammasome049648- / 618931 T19083WO001 pathway and do not increase IL1ß mRNA (2A) or IL1ß protein (2B), and do not activate caspase 1 (2C) after neonatal hyperoxia exposure.

[0148] A single subcutaneous dose of Peptide 152 was given to neonatal mice on postnatal day 1 (PN1) of life at the start of hyperoxia exposure and lung lavage inflammatory cells and IL1ß protein were measured at PN7. A dose response study (dosing from 0.0002 mg / kg of subject body weight to 20 mg / kg of subject body weight) was developed to determine the optimal dosing for Peptide 152. Figures 3A-C show dose-dependent blockade of NLRP3 inflammasome activity and inflammation with SQ Peptide 152. Figure 3A (IL1ß protein), Figure 3B (MPO = neutrophil content); Figure 3C (NAG = macrophage content). The optimal dose determined from Figures 3A-C was 0.02 mg / kg of subject body weight.

[0149] Since alveolarization is inhibited in BPD and in the mouse model of BPD, a single SQ dose of Peptide 152 [dosage of 10 µmoles / kg of subject body weight (20 mg / kg of subject body weight)] was examined to determine if it would rescue alveolarization due to neonatal hyperoxia exposure on PN15. Figures 4A-B show that neonatal hyperoxia in mice is associated with decreased alveolarization (decreased Radial Alveolar Count, RAC) and that Peptide 152 rescues this phenotype. Figures 4A-B: Hyperoxia decreases alveolarization and a single SQ dose of Peptide 152 rescues this outcome. Figure 4A microscope images; Figure 4B Plot of radial alveolar counts determined from images in 4A.

[0150] Next, the long-term effects of neonatal hyperoxia on NLRP3 inflammasome activation were determined and whether Peptide 152 administration [dosage of 10 µmoles / kg of subject body weight (20 mg / kg of subject body weight)] at the later timepoint affects these outcomes. Mice were exposed to either 21% or 95% O2from PN1 to PN5 and then allowed to remain in 21% O2until 10 weeks of age. The activation status of the NLRP3 inflammasome at the 10-week timepoint was determined. Figures 5A-D: Neonatal hyperoxia results in chronic activation of the NLRP3 inflammasome and inflammation. Peptide 152 given at 10 weeks of age inhibits this activation and inflammation. Figures 5A-D show that lung IL1ß mRNA (5A) and protein (5B), as well as neutrophil (MPO; 5C) and macrophage (NAG; 5D) contents were elevated in mice that had previously been exposed to neonatal hyperoxia from PN1 to PN5, suggesting chronic049648- / 618931 T19083WO001 activation of the NLRP3 inflammasome. One SQ dose of Peptide 152 inhibited this activation and inflammation.

[0151] It is known that neonatal hyperoxia exposure in mice results in exaggerated responses to acute lung injury in adulthood. The effects of neonatal hyperoxia exposure on intranasal bleomycin injury at 10 weeks of age and the effects of Peptide 152 on these responses were determined. Studies in Figures 6A-D confirmed the exaggerated response to acute lung injury in mice previously exposed to neonatal hyperoxia and showed that Peptide 152 given at 10 weeks [dosage of 10 µmoles / kg of subject body weight (20 mg / kg of subject body weight)] prevents this exaggerated response. Figures 6A-D: Neonatal hyperoxia exposure results in exaggerated NLRP3 inflammasome and inflammation responses to acute lung injury at 10 weeks of age (increased lung IL1ß mRNA (6A), protein (6B), neutrophils (6C), and macrophages (6D)). A single SQ dose of Peptide 152 given at the time of IN bleomycin blocked the inflammasome and inflammation responses. Peptide 152 given at the time of neonatal hyperoxia (Figures 3 and 4) prevents the long-term activation of the NLRP3 inflammasome and the later exaggerated responses to acute lung injury.

[0152] These data also demonstrate the efficacy of Peptide 152 in blocking NLRP3 activation and inflammation in the bleomycin model of acute lung injury (room air-exposed mice).

[0153] Example 2. Mouse models of LPS-induced systemic (intraperitoneal) sepsis and pneumonia (intranasal)

[0154] Both foreign organisms and endogenous danger signals are sensed by pattern recognition receptors such as Toll-Like Receptors (TLRs). TLR4 senses lipopolysaccharide (LPS) and, when LPS is administered intraperitoneally to adult mice, results in activation of the NLRP3 inflammasome, increased plasma cytokines and evidence of systemic sepsis. In addition, intranasal administration of LPS, a model of gram-negative pneumonia, results in an influx of inflammatory cells, mostly neutrophils, at 24 hours after challenge. Studies were performed to determine if Peptide 152 was able to block the systemic and lung effects of IP and IN LPS respectively. Figure 7A shows a dose-dependent decrease in plasma IL1ß 3 hours after IP LPS and a single SQ dose of Peptide 152 [dosage at 1 nmole / kg of subject body weight, 10 nmole / kg of subject body weight, 100 nmole / kg of subject body weight]. Flow cytometry of a single cell049648- / 618931 T19083WO001 preparation of whole lung after IN saline or LPS shows that IN LPS results in increased white blood cells, largely neutrophils, but also macrophages and that a single SQ dose of Peptide 152 of 10 µmoles / kg of subject body weight (20 mg / kg of subject body weight) decreases the accumulation of these cells in the lung 24 hours later (7B).

[0155] Example 3. The effects of Peptide 152 in viral-mediated inflammation – Mouse models of TLR7-, influenza A-, and SARS-CoV-2-mediated pneumonia

[0156] Coronaviruses are a diverse group of viruses that cause human diseases and affect multiple organ systems with wide variation in severity ranging from asymptomatic to severe. The most common severe presentation of coronavirus disease 2019 (COVID-19) is respiratory distress due to a viral pneumonia and the consequences of the activation of host immune responses. The single stranded RNA Betacoronavirus SARS-CoV-2 that causes COVID-19 consists of a viral pneumonia, the elaboration of cytokines, and respiratory failure in those vulnerable

[0025] . Survivors of COVID-19-induced respiratory failure often develop progressive fibrosis leading to chronic respiratory failure and the need for lung transplantation [26,27]. In addition, this disease affects multiple other organs including the brain, heart, kidney, pancreas and muscles

[0028] . The course of the COVID-19 pandemic has placed enormous pressure on health care systems and public health infrastructure, causing collateral economic and social damage worldwide.

[0157] SARS-CoV-2 is highly transmissible, with a frequent rise of variants that make the virus more infectious

[0029] . Despite the remarkable development of apparently effective vaccines against the original SARS-COV-2, it is becoming clear that emerging viral variants exhibit increasing evasion of existing convalescent or vaccine-induced immune protection. Vaccine hesitancy has also encouraged the spread and severity of the disease. Therefore, unraveling the biology of this disease and development of other therapeutics to combat severe COVID-19 remain essential.

[0158] Although classically, SARS-CoV-2 infection occurs via airway epithelial cells, several key observations have demonstrated the ability of this virus to infect inflammatory cells, in particular macrophages. Infection of PBMCs depends on FcγR uptake

[0030] and, in a humanized ACE2R mouse model, is associated with NLRP3 inflammasome activation, the049648- / 618931 T19083WO001 elaboration of IL1ß and hyperinflammation. Blockade of the NLRP3 inflammasome resulted in reversal of lung pathology

[0031] . Further, while infection in epithelial cells does increase IL1ß, the level of expression is not high suggesting that the major inflammatory response is not driven by epithelial cells

[0032] . Lastly, studies show that Spike protein directly activates NF ^B in human macrophages via TLR2 to induce inflammatory cytokine expression

[0033] .

[0159] Similarly, influenza A virus (IAV), the only species of the genus Alphainfluenzavirus, causes a viral pneumonia and respiratory distress that is lethal in vulnerable people, and can occasionally cause pandemics with widespread severe illness and death. Influenza A viruses are also single-stranded RNA viruses. Two surface molecules define the nomenclature, namely hemagglutinin (H) and neuraminidase (N). The principal strains currently active are H1N1 (Spanish flu pandemic), H1N2 (Asian flu pandemic) and H3N2 (Hong Kong flu pandemic). Influenza A / PR8 / 34 is a mouse adapted strain that is widely used to interrogate the pathogenesis of influenza infection, and to compare against other viral pneumonias, including SARS-CoV-2.

[0160] Several pattern recognition receptors act as sensors for viral RNA. TLR3 senses double stranded (ds) RNA, TLR7 and TLR8 sense single stranded (ss) RNA, RIG1 senses short dsRNA and ssRNA motifs and MDA5 senses long dsRNA

[0034] . These danger sensors drive both interferon and cytokine production by the activation of Interferon Regulatory Factors (IRFs), NF ^B and the NLRP3 inflammasome

[0035] . Two TLR7 / 8 agonists, R848 and SM324405, have been studied. R848 stimulates both TLR7 / 8 whereas SM324405 is specific to TLR7

[0036] . SM324405 is an antedrug that is rapidly metabolized to an inactive form and has a plasma half- life of about 2.5 minutes. SM324405 has few to no systemic side effects. SM3244045 stimulates multiple cytokines in human PBMCs and stimulates IFN production in both human and mouse cells. [36, 44] In order to mimic SARS-CoV-2 responses specifically through a single TLR that drives ssRNA responses, a non-infectious lung injury model was established in which intratracheal (IT) administration of SM324405, the specific TLR7 agonist, results in elaboration of interferons α, ß and γ, as well as IL6 and IL1ß.

[0161] Example 4. Non-infectious model of viral infection using IT administration of a pure TLR7 agonist049648- / 618931 T19083WO001

[0162] Peptide 152 was tested in a non-infectious model of viral infection using IT administration of a pure TLR7 agonist (SM324405).

[0163] Figures 8A-B show that administration of IT SM324405 results in increased MPO and NAG activities in cells obtained by BAL over time, indicating lung accumulation of neutrophils (8A) and macrophages (8B), respectively.

[0164] Figures 9A-C show that lung IL1ß mRNA (9A) and protein (9B), as well as IFN-γ (9C) are elevated after IT SM324405 suggesting that activation of TLR7 results in activation of the NLRP3 inflammasome and Interferon Regulatory Factors in a manner similar to viral infection.

[0165] Figures 10A-E show that similar to responses previously reported for human Influenza A and SARS-CoV-2 infections, IT SM324405 results in increase expression of HA synthases, as well as in plasma and lung HA. HAS1 mRNA (10A), HAS2 mRNA (10B), HAS3 mRNA (10C), plasma HA content (10D), and BAL HA content (10E).

[0166] Next, studies determined the effect of one SQ dose of Peptide 152 [dosage of 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight)] SQ peptide on the responses to IT SM324405. The inventors were able to show that Peptide 152 was able to block caspase-1 activity, plasma HA, and lung IL1ß protein elaboration after IT SM324405 (Figures 11A-C).

[0167] Figures 11A-C show that Peptide 152 given SQ abrogated the increased caspase-1 activity, plasma HA and lung IL1ß protein associated with IT SM324405. caspase 1 activity (11A), plasma HA (11B), and BAL IL1ß (11C). Mice were treated with 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight) SQ peptide.

[0168] Blockade of the NLRP3 pathway would predict that inflammation would also be blocked. Lavage cells were examined for their neutrophil (MPO) and macrophage (NAG) contents after IT SM324405 and Peptide 152 treatment.

[0169] Figures 12A-B show that Peptide 152 was able to block the inflammatory response to IT SM324405, suggesting that the inflammatory response to TLR7 activation can be inhibited by Peptide 152. MPO activity (neutrophils) (12A) and NAG activity (macrophages) (12B). Mice049648- / 618931 T19083WO001 were treated with 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight) SQ peptide.

[0170] Studies then determined the response to IT SM324405 in WT and RHAMM KO mice. Figures 13A-B show that RHAMM KO mice do not develop inflammation after TLR7 activation.

[0171] Figures 13A-B show that RHAMM KO mice do not have any increase in lung neutrophil or macrophage accumulation after IT administration of SM 324405. MPO activity (13A) and N-acetyl glucosaminidase activity (13B).

[0172] Example 5. Mouse model of Influenza A viral pneumonia

[0173] To test the efficacy of Peptide 152 in viral models of lung inflammation, the mouse- adapted strain of Influenza A (PR8 / 34) was used to establish the mouse model of Influenza A pneumonia using dose- and time-dependent studies after intranasal (IN) administration of the mouse-adapted virus (Figures 14A-B, 15A-B and 16A-B).

[0174] Figures 14A-B depict results of studies in a mouse model of influenza A viral pneumonia, where lung IL-1ß protein (14A) and lung IL-1ß protein after 7 days (14B) are measured. Figures 14A-B: IN Influenza A / PR8 / 34 increased lung IL1ß in a dose-dependent manner and was maximally increased 48 hours after infection with 20k pfu.

[0175] Figures 15A-B depict results of studies in a mouse model of influenza A viral pneumonia, where dose (15A) and time (15B)-dependent increases in HA in the BAL are measured. Figures 15A-B: IN Influenza A / PR8 / 34 resulted in a dose- and time-dependent increase in lung HA content with maximal increase at 7 days after infection.

[0176] Figures 16A-B depict results of studies in a mouse model of influenza A viral pneumonia, where MPO activity with varying dose (16A) and MPO activity over time (16B) are measured. Figures 16A-B: IN Influenza A / PR8 / 34 resulted in a dose- and time-dependent increase in neutrophil accumulation in the lung with maximal inflammation at 7 days after infection.049648- / 618931 T19083WO001

[0177] A single SQ dose of Peptide 152 [dosage of 20 mg / kg of subject body weight] was administered at 48 hours after IN Influenza virus administration and the MPO activity of cells obtained by lung lavage on day 7 after infection was determined. Peptide 152 was able to decrease neutrophil accumulation after Influenza A infection (Figure 17). Figure 17: Peptide 152 was given SQ 48 hours after IN Influenza A in mice and lung lavage cells were examined for their MPO activity on day 7. Peptide 152 decreased neutrophil accumulation due to Influenza A viral infection. 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight) Peptide 152 was given SQ at 48 hours after influenza infection (Figure 17).

[0178] These data show that the TLR7 agonist model accurately mimics Influenza A viral infection and that Peptide 152 can inhibit the inflammatory response to the virus.

[0179] Example 6. Mouse model of SARS-CoV-2 viral pneumonia

[0180] Mouse ACE2 receptor does not bind to SARS-CoV-2. Transgenic mice that express human ACE2 on the K18 keratinocyte promoter and human ACE2 receptor knock-in mice have been developed and used to establish mouse models of COVID-19

[0037] . SARS-CoV-2 was studied in mice. This infection is associated with increased expression of TLR7, IL1ß, IFNγ, and HAS2 mRNA. Figures 18A-F depict results of studies in a mouse model of SARS-CoV-2 viral pneumonia, where TLR7 mRNA (18A), IL1ß mRNA (18B), IFNγ mRNA (18C) HAS2 mRNA (18D), TLR2 mRNA (18E), and Hmmr mRNA (18F) are measured. SARS-CoV-2 infection in K18-hACE2R mice results in increased TLR7, IL1ß, IFN γ, HAS2 mRNA, TLR2 mRNA, and Hmmr mRNA at 6 days after infection. Figures 18A-F: SARS-CoV-2 infection in K18-hACE2R mice results in increased TLR7, IL1ß, IFNγ, HAS2 mRNA, TLR2 mRNA, and Hmmr mRNA at 6 days after infection.

[0181] A single SQ dose of Peptide 152 [dosage of 20 mg / kg of subject body weight] was given to mice three days after infection with SARS-CoV-2 and showed efficacy in decreasing inflammation associated with viral infection. Figure 19A depicts microscopy images. Left panel (Uninfected); middle panel (Infected, PBS control); right panel (Infected, Peptide 152 on day 3). Figure 19B depicts lung score quantified from Figure 19A images. Figures 19A-B: SARS-CoV- 2 infection in mice is associated with an extensive inflammatory response. A single SQ dose of Peptide 152 on day 3 after infection decreased the inflammatory response to infection. To049648- / 618931 T19083WO001 determine the optimum timing of peptide dosing, time courses of inflammation after SARS- CoV-2 infection were examined using MPO activity (as a measure of neutrophils) and NAG activities (as a measure of macrophages) of cells obtained by bronchoalveolar lavage. Maximum neutrophil and macrophage accumulation in the lung was seen at 24 hours after infection. Figures 20A-B depict results of time course studies after SARS-CoV-2 infection in K18- hACE2R mice. MPO activity (20A) and NAG activity (20B). Figures 20A-B: Time course of inflammation after SARS-CoV-2 infection in K18-hACE2R mice shows maximum neutrophil (MPO) and macrophages (NAG) at 24 hours after infection. For studies depicted in Figures 20A- B, 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight) peptide was given SQ 72h after IN SARS-CoV-2 infection. Based on these data, an optimum dosing for severe viral illnesses is multiple doses, starting as early as six hours after infection, and then continuing until at least 96 hours after infection.

[0182] Example 7. Materials and Methods –

[0183] Neonatal Mouse Hyperoxia Model of Bronchopulmonary Dysplasia

[0184] Neonatal mice were used in the first fourteen postnatal (PN) days, with a smaller cohort allowed to reach PN30 for non-sedated respiratory physiology and mechanics measurements. All WT mice were in a pure C57Bl / 6 background obtained from Jackson Laboratories, Massachusetts. Rhamm- / -mice, also on a pure C57Bl / 6 background, were the kind gift of Dr. Eva A. Turley (Lawson Research Institute, University of Western Ontario, London, ON, Canada). Breeder cages were established consisting of two females with one male. Once pregnancy was established, the pregnant mice were separated into individual cages where birth occurred. All cages were checked daily, and the date of birth recorded. Litter sizes were adjusted to 6 pups per litter and 12 pups (2 litters) per treatment group, with equal numbers of males and females, to normalize the effects of nutrition on lung development and randomized to either 21% oxygen(O2) or 95% O2from PN1 to PN5 to provide hyperoxia exposure during the saccular phase oflung development. All mice were then maintained in 21% O2until harvest at PN7, PN14 or PN30. The oxygen environment was regulated using the Biospherix-controlled atmosphere animal chamber (Biospherix, Redfield, NY). This system accurately maintained the oxygen content of experimental chambers to ± 2%. For all studies, the mice were observed every 12049648- / 618931 T19083WO001hours to monitor well-being and mortality. Mothers were rotated between 21% O2and 95% O2cages every 24 hours during the first five days to prevent acute lung injury from hyperoxia. The mothers’ noses were dabbed with vanilla oil at each rotation to prevent cannibalization of the litters.

[0185] Long-term outcomes of neonatal hyperoxia exposure in mice

[0186] Infants with BPD have long-term morbidity and poorer outcomes with lung injury into adolescence and adulthood. A number of studies have demonstrated long-term impacts of neonatal hyperoxia exposure in mice. Thus, neonatal hyperoxia is associated with long-term impaired alveolarization, pulmonary vascular disease and a shortened life span. In addition, neonatal hyperoxia is associated with increased lung inflammation, fibrosis and TGFß1 when these mice are exposed to intratracheal bleomycin as a “second hit” in adulthood. The studies examined the effects of RHAMM-derived peptides given in the first week after birth on the late outcomes of decreased alveolarization and increased IL1ß, as well as to the response to intranasal (IN) bleomycin (2U / kg) as a model of acute lung injury at eight weeks of age. In addition, 10-week-old mice with and without neonatal hyperoxia exposure were given a single subcutaneous injection of peptide to determine whether this confers protection when used in adulthood. It has previously been shown that RHAMM-derived peptide inhibits inflammation and fibrosis in a rodent intratracheal bleomycin model of acute lung injury [8]. Mice given either 21% or 95% O2from PN1 to PN5 were allowed to feed ad libitum until young adulthood at 10 weeks of age. One cohort of animals was sacrificed at the 10-week mark to determine the long- term effects of neonatal hyperoxia. Other cohorts (n=12 / group, equal numbers of males and females) were given either IN bleomycin or an equivalent volume of IN saline. Mice were examined at 7 and 14 days after IN treatments for an assessment of loss of body weight as an indicator of the severity of disease, neutrophil and macrophage influx into the lung, and biochemical studies of the Nlrp3 inflammasome (caspase 1 activity and western blot), cytokine elaboration (IL1ß mRNA and protein), and inflammatory cell accumulation (flow cytometry, MPO and NAG activity assays). In addition, 10-week-old mice previously exposed to either 21% or 95% O2were given SQ peptide prior to IN bleomycin or saline to determine if this treatment ameliorates the adverse outcome of acute lung injury.049648- / 618931 T19083WO001

[0187] Animals were harvested at designated time points (PN7, PN14) in order to assess inflammation, alveolarization, cytokine / growth factor elaboration, and activation of the Nlrp3 inflammasome and at 10 weeks to determine the late outcomes after neonatal hyperoxia exposure. The next harvest was at 7 and 14 days after IN bleomycin or saline in 10-week old mice. At the end of the exposures, mice were given intraperitoneal ketamine / xylazine (75 / 3 mg / kg of subject body weight) for anesthesia. For tissue harvest, the lung was cleared of all blood by pulmonary artery infusion with saline. Bronchoalveolar lavage (BAL) was obtained using 36 ml / kg (= total lung volume) of LPS-negative, sterile saline to normalize for differences in the size of animals as previously described [8]. BAL was spun to separate cells from supernatant. Cells were examined for MPO and NAG activities for neutrophil and macrophage content. Supernatant was used to determine concentrations of IL1ß and hyaluronan (HA). The left lung was inflated and processed for histologic and morphometric analysis. The right lung was snap frozen for later protein / RNA studies, including caspase-1 activity assays.

[0188] Peptide 152 Dosing Conversion of doses: 0.0002 mg / kg of subject body weight = 0.1 nmoles / kg of subject body weight 0.002 mg / kg of subject body weight = 1 nmole / kg of subject body weight 0.02 mg / kg of subject body weight = 10 nmole / kg of subject body weight 0.2 mg / kg of subject body weight = 100 nmole / kg of subject body weight 2 mg / kg of subject body weight = 1 µmole / kg of subject body weight (1,000 nmole / kg of subject body weight) 6 mg / kg of subject body weight = 3 µmole / kg of subject body weight (3,000 nmole / kg of subject body weight) 20 mg / kg of subject body weight = 10 µmole / kg of subject body weight (10,000 nmole / kg of subject body weight)049648- / 618931 T19083WO001

[0189] Intraperitoneal lipopolysaccharide (LPS) mouse model of systemic sepsis

[0190] Adult C57Bl / 6 mice were given intraperitoneal (IP) LPS and blood samples were obtained three hours later. Plasma was separated by centrifugation and an ELISA kit was used to determine the concentration of IL1ß in the samples. Peptide was administered SQ at doses ranging from 0.002 mg / kg of subject body weight to 0.2 mg / kg of subject body weight (Figures 7A-B), where the optimal dose was 0.2 mg / kg of subject body weight (100 nmoles / kg of subject body weight).

[0191] Adult C57Bl / 6 mice were given intranasal (IN) LPS and lungs were harvested and processed as single cell preparations for flow cytometry at 24 hours later. Flow cytometry analysis of inflammatory cells showed that IN LPS resulted in accumulation of neutrophils and macrophages. SQ treatment with one 0.2 mg / kg of subject body weight dose of Peptide 152 at the time of IN LPS resulted in decreased inflammation in the lung at 24 hours (Figure 7B)

[0192] Mouse model of intratracheal (IT) SM345405 (TLR7 agonist)-induced inflammation

[0193] Preliminary dose-response studies of intratracheal (IT) SM324405 in wild type young adult mice aged 8-12 weeks were conducted to determine the optimal dose (data not shown). It was concluded that the optimal dose of SM324405 IT, which caused injury without any mortality, was 1 mg / kg of subject body weight. An equal volume of vehicle (normal saline with DMSO) was used as a vehicle control. Further, using sham-operated mice where the trachea was exposed but no injections were given IT, it was ensured that surgery alone was not associated with any of the changes seen with IT administration of SM324405. Mice were harvested as described above (under “Long-term outcomes of neonatal hyperoxia exposure in mice”) between 4 and 72 hours as a time course to determine changes in MPO, NAG, IL1ß, HA, and protein / mRNA studies. For peptide studies, mice were treated with 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight) SQ peptide (Figures 11 and 12).

[0194] Mouse model of intranasal (IN) Influenza A viral pneumonia

[0195] All influenza A experiments were performed under BSL-2 protocols with trained personnel. Mice (8-10 weeks of age) were anesthetized using isofluorane and an inoculum of the mouse-adapted A / PR8 / 34 strain of influenza A between 5,000 and 20,000 PFU was given049648- / 618931 T19083WO001 intranasally (IN) to WT and Rhamm- / -mice (both C57Bl / 6). Mice were allowed to recover and were observed regularly for signs of illness. Mice were sacrificed at 2, 4, and 6 days after infection, and tissues were processed as described above in section “Long-term outcomes of neonatal hyperoxia exposure in mice.”

[0196] . For peptide studies, 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight) peptide was given SQ at 48 hours after influenza infection (Figure 17).

[0197] Mouse model of SARS-CoV-2 viral pneumonia

[0198] All SARS-CoV-2 experiments were performed in the BSL3 facility at Cornell University (Dr. Hector Aguilar-Carreño / David Gludish), following containment procedures required by the Cornell University Institutional Biosafety Committee. WT mice are resistant to SARS-CoV-2 infection [5]. Several genetic models have been developed and include both transgenic expression of hACE2 receptor under the control of the human cytokeratin 18 promoter (K18-hACE2 receptor mice)

[0044] or mice that have had the human ACE2 receptor knocked into the mouse locus (KO / KI mice)

[0045] . Mice were anesthetized using isoflurane and an intranasal inoculum of 1 x 104viral particles was given intranasally to hACE2 receptor transgenic mice. Mice were allowed to recover and were observed regularly and at least twice daily for signs of illness. Mice were sacrificed at 2, 4 and 6 days after infection. Tissues were harvested and processed as described above in section “Long-term outcomes of neonatal hyperoxia exposure in mice.”

[0199] In peptide treatment studies, 20 mg / kg of subject body weight (10 µmoles / kg of subject body weight) peptide was given SQ 72h after IN SARS-CoV-2 infection (Figure 19A-B).

[0200] Example 8. Human dosing of P152

[0201] An exemplary dosing of P152 in human neonates is 0.2 mg / kg of subject body weight.

[0202] An exemplary dosing for diseases that involve severe inflammation (such as SARS- CoV-2 or ARDS) in both human children and human adults is 20 mg / kg of subject body weight. In some treatments for diseases that involve severe inflammation (such as SARS-CoV-2 or049648- / 618931 T19083WO001 ARDS) in both human children and human adults is 20 mg / kg of subject body weight, with 1-3 doses over the first seven days after clinical presentation.

[0203] An exemplary dosing for severe viral illnesses in humans is multiple doses, starting as early as six hours after infection, and then continuing until at least 96 hours after infection.

[0204] In exemplary dose escalation studies in Phase 1 and 2 clinical trials, safety, tolerability and optimal dosing are determined in healthy human subjects either with and without IV LPS administration (2ng / kg of subject body weight), an established and accepted human model of systemic sepsis.049648- / 618931 T19083WO001

[0205] Sequence Listing

[0206] SEQ ID NO:1 amino acid sequence of human RHAMM UniProt O75330-1 HMMR_HUMAN MSFPKAPLKRFNDPSGCAPSPGAYDVKTLEVLKGPVSFQKSQRFKQQKESKQNLNVDKDTTLPA SARKVKSSESKESQKNDKDLKILEKEIRVLLQERGAQDRRIQDLETELEKMEARLNAALREKTS LSANNATLEKQLIELTRTNELLKSKFSENGNQKNLRILSLELMKLRNKRETKMRGMMAKQEGME MKLQVTQRSLEESQGKIAQLEGKLVSIEKEKIDEKSETEKLLEYIEEISCASDQVEKYKLDIAQ LEENLKEKNDEILSLKQSLEENIVILSKQVEDLNVKCQLLEKEKEDHVNRNREHNENLNAEMQN LKQKFILEQQEREKLQQKELQIDSLLQQEKELSSSLHQKLCSFQEEMVKEKNLFEEELKQTLDE LDKLQQKEEQAERLVKQLEEEAKSRAEELKLLEEKLKGKEAELEKSSAAHTQATLLLQEKYDSM VQSLEDVTAQFESYKALTASEIEDLKLENSSLQEKAAKAGKNAEDVQHQILATESSNQEYVRML LDLQTKSALKETEIKEITVSFLQKITDLQNQLKQQEEDFRKQLEDEEGRKAEKENTTAELTEEI NKWRLLYEELYNKTKPFQLQLDAFEVEKQALLNEHGAAQEQLNKIRDSYAKLLGHQNLKQKIKH VVKLKDENSQLKSEVSKLRCQLAKKKQSETKLQEELNKVLGIKHFDPSKAFHHESKENFALKTP LKEGNTNCYRAPMECQESWK

[0207] SEQ ID NO:2 amino acid sequence of mouse RHAMM UniProt Q00547-1·HMMR_MOUSE MSFPKAPLKRFNDPSGCAPSPGAYDVKTSEATKGPVSFQKSQRFKNQRESQQNLNIDKDTTLLA SAKKAKKSVSKKDSQKNDKDVKRLEKEIRALLQERGTQDKRIQDMESELEKTEAKLNAAVREKT SLSASNASLEKRLTELTRANELLKAKFSEDGHQKNMRALSLELMKLRNKRETKMRSMMVKQEGM ELKLQATQKDLTESKGKIVQLEGKLVSIEKEKIDEKCETEKLLEYIQEISCASDQVEKCKVDIA QLEEDLKEKDREILSLKQSLEENITFSKQIEDLTVKCQLLETERDNLVSKDRERAETLSAEMQI LTERLALERQEYEKLQQKELQSQSLLQQEKELSARLQQQLCSFQEEMTSEKNVFKEELKLALAE LDAVQQKEEQSERLVKQLEEETKSTAEQLTRLDNLLREKEVELEKHIAAHAQAILIAQEKYNDT AQSLRDVTAQLESVQEKYNDTAQSLRDVTAQLESEQEKYNDTAQSLRDVTAQLESEQEKYNDTA QSLRDVTAQLESVQEKYNDTAQSLRDVTAQLESYKSSTLKEIEDLKLENLTLQEKVAMAEKSVE DVQQQILTAESTNQEYARMVQDLQNRSTLKEEEIKEITSSFLEKITDLKNQLRQQDEDFRKQLE EKGKRTAEKENVMTELTMEINKWRLLYEELYEKTKPFQQQLDAFEAEKQALLNEHGATQEQLNK049648- / 618931 T19083WO001 IRDSYAQLLGHQNLKQKIKHVVKLKDENSQLKSEVSKLRSQLVKRKQNELRLQGELDKALGIRH FDPSKAFCHASKENFTPLKEGNPNCC

[0208] SEQ ID NO:3 amino acid sequence of Peptide 152 RHAMM- derived peptide VSKLREQLVKRKQN, wherein E at position 6 is covalently bonded to K at position 10.

[0209] SEQ ID NO:4 amino acid sequence of human TLR4_HUMAN UniProt O00206-1 MMSASRLAGTLIPAMAFLSCVRPESWEPCVEVVPNITYQCMELNFYKIPDNLPFSTKNLDLSFN PLRHLGSYSFFSFPELQVLDLSRCEIQTIEDGAYQSLSHLSTLILTGNPIQSLALGAFSGLSSL QKLVAVETNLASLENFPIGHLKTLKELNVAHNLIQSFKLPEYFSNLTNLEHLDLSSNKIQSIYC TDLRVLHQMPLLNLSLDLSLNPMNFIQPGAFKEIRLHKLTLRNNFDSLNVMKTCIQGLAGLEVH RLVLGEFRNEGNLEKFDKSALEGLCNLTIEEFRLAYLDYYLDDIIDLFNCLTNVSSFSLVSVTI ERVKDFSYNFGWQHLELVNCKFGQFPTLKLKSLKRLTFTSNKGGNAFSEVDLPSLEFLDLSRNG LSFKGCCSQSDFGTTSLKYLDLSFNGVITMSSNFLGLEQLEHLDFQHSNLKQMSEFSVFLSLRN LIYLDISHTHTRVAFNGIFNGLSSLEVLKMAGNSFQENFLPDIFTELRNLTFLDLSQCQLEQLS PTAFNSLSSLQVLNMSHNNFFSLDTFPYKCLNSLQVLDYSLNHIMTSKKQELQHFPSSLAFLNL TQNDFACTCEHQSFLQWIKDQRQLLVEVERMECATPSDKQGMPVLSLNITCQMNKTIIGVSVLS VLVVSVVAVLVYKFYFHLMLLAGCIKYGRGENIYDAFVIYSSQDEDWVRNELVKNLEEGVPPFQ LCLHYRDFIPGVAIAANIIHEGFHKSRKVIVVVSQHFIQSRWCIFEYEIAQTWQFLSSRAGIIF IVLQKVEKTLLRQQVELYRLLSRNTYLEWEDSVLGRHIFWRRLRKALLDGKSWNPEGTVGTGCN WQEATSI

[0210] SEQ ID NO:5 amino acid sequence of mouse TLR4 UniProt Q9QUK6049648- / 618931 T19083WO001 MMPPWLLARTLIMALFFSCLTPGSLNPCIEVVPNITYQCMDQKLSKVPDDIPSSTKNIDLSFNP LKILKSYSFSNFSELQWLDLSRCEIETIEDKAWHGLHHLSNLILTGNPIQSFSPGSFSGLTSLE NLVAVETKLASLESFPIGQLITLKKLNVAHNFIHSCKLPAYFSNLTNLVHVDLSYNYIQTITVN DLQFLRENPQVNLSLDMSLNPIDFIQDQAFQGIKLHELTLRGNFNSSNIMKTCLQNLAGLHVHR LILGEFKDERNLEIFEPSIMEGLCDVTIDEFRLTYTNDFSDDIVKFHCLANVSAMSLAGVSIKY LEDVPKHFKWQSLSIIRCQLKQFPTLDLPFLKSLTLTMNKGSISFKKVALPSLSYLDLSRNALS FSGCCSYSDLGTNSLRHLDLSFNGAIIMSANFMGLEELQHLDFQHSTLKRVTEFSAFLSLEKLL YLDISYTNTKIDFDGIFLGLTSLNTLKMAGNSFKDNTLSNVFANTTNLTFLDLSKCQLEQISWG VFDTLHRLQLLNMSHNNLLFLDSSHYNQLYSLSTLDCSFNRIETSKGILQHFPKSLAFFNLTNN SVACICEHQKFLQWVKEQKQFLVNVEQMTCATPVEMNTSLVLDFNNSTCYMYKTIISVSVVSVI VVSTVAFLIYHFYFHLILIAGCKKYSRGESIYDAFVIYSSQNEDWVRNELVKNLEEGVPRFHLC LHYRDFIPGVAIAANIIQEGFHKSRKVIVVVSRHFIQSRWCIFEYEIAQTWQFLSSRSGIIFIV LEKVEKSLLRQQVELYRLLSRNTYLEWEDNPLGRHIFWRRLKNALLDGKASNPEQTAEEEQETA TWT

[0211] SEQ ID NO:6 amino acid sequence of human TLR7 UniProt Q9NYK1 MVFPMWTLKRQILILFNIILISKLLGARWFPKTLPCDVTLDVPKNHVIVDCTDKHLTEIPGGIP TNTTNLTLTINHIPDISPASFHRLDHLVEIDFRCNCVPIPLGSKNNMCIKRLQIKPRSFSGLTY LKSLYLDGNQLLEIPQGLPPSLQLLSLEANNIFSIRKENLTELANIEILYLGQNCYYRNPCYVS YSIEKDAFLNLTKLKVLSLKDNNVTAVPTVLPSTLTELYLYNNMIAKIQEDDFNNLNQLQILDL SGNCPRCYNAPFPCAPCKNNSPLQIPVNAFDALTELKVLRLHSNSLQHVPPRWFKNINKLQELD LSQNFLAKEIGDAKFLHFLPSLIQLDLSFNFELQVYRASMNLSQAFSSLKSLKILRIRGYVFKE LKSFNLSPLHNLQNLEVLDLGTNFIKIANLSMFKQFKRLKVIDLSVNKISPSGDSSEVGFCSNA RTSVESYEPQVLEQLHYFRYDKYARSCRFKNKEASFMSVNESCYKYGQTLDLSKNSIFFVKSSD FQHLSFLKCLNLSGNLISQTLNGSEFQPLAELRYLDFSNNRLDLLHSTAFEELHKLEVLDISSN SHYFQSEGITHMLNFTKNLKVLQKLMMNDNDISSSTSRTMESESLRTLEFRGNHLDVLWREGDN RYLQLFKNLLKLEELDISKNSLSFLPSGVFDGMPPNLKNLSLAKNGLKSFSWKKLQCLKNLETL DLSHNQLTTVPERLSNCSRSLKNLILKNNQIRSLTKYFLQDAFQLRYLDLSSNKIQMIQKTSFP ENVLNNLKMLLLHHNRFLCTCDAVWFVWWVNHTEVTIPYLATDVTCVGPGAHKGQSVISLDLYT049648- / 618931 T19083WO001 CELDLTNLILFSLSISVSLFLMVMMTASHLYFWDVWYIYHFCKAKIKGYQRLISPDCCYDAFIV YDTKDPAVTEWVLAELVAKLEDPREKHFNLCLEERDWLPGQPVLENLSQSIQLSKKTVFVMTDK YAKTENFKIAFYLSHQRLMDEKVDVIILIFLEKPFQKSKFLQLRKRLCGSSVLEWPTNPQAHPY FWQCLKNALATDNHVAYSQVFKETV

[0212] SEQ ID NO:7 amino acid sequence of mouse TLR7_MOUSE UniProt P58681 MVFSMWTRKRQILIFLNMLLVSRVFGFRWFPKTLPCEVKVNIPEAHVIVDCTDKHLTEIPEGIP TNTTNLTLTINHIPSISPDSFRRLNHLEEIDLRCNCVPVLLGSKANVCTKRLQIRPGSFSGLSD LKALYLDGNQLLEIPQDLPSSLHLLSLEANNIFSITKENLTELVNIETLYLGQNCYYRNPCNVS YSIEKDAFLVMRNLKVLSLKDNNVTAVPTTLPPNLLELYLYNNIIKKIQENDFNNLNELQVLDL SGNCPRCYNVPYPCTPCENNSPLQIHDNAFNSLTELKVLRLHSNSLQHVPPTWFKNMRNLQELD LSQNYLAREIEEAKFLHFLPNLVELDFSFNYELQVYHASITLPHSLSSLENLKILRVKGYVFKE LKNSSLSVLHKLPRLEVLDLGTNFIKIADLNIFKHFENLKLIDLSVNKISPSEESREVGFCPNA QTSVDRHGPQVLEALHYFRYDEYARSCRFKNKEPPSFLPLNADCHIYGQTLDLSRNNIFFIKPS DFQHLSFLKCLNLSGNTIGQTLNGSELWPLRELRYLDFSNNRLDLLYSTAFEELQSLEVLDLSS NSHYFQAEGITHMLNFTKKLRLLDKLMMNDNDISTSASRTMESDSLRILEFRGNHLDVLWRAGD NRYLDFFKNLFNLEVLDISRNSLNSLPPEVFEGMPPNLKNLSLAKNGLKSFFWDRLQLLKHLEI LDLSHNQLTKVPERLANCSKSLTTLILKHNQIRQLTKYFLEDALQLRYLDISSNKIQVIQKTSF PENVLNNLEMLVLHHNRFLCNCDAVWFVWWVNHTDVTIPYLATDVTCVGPGAHKGQSVISLDLY TCELDLTNLILFSVSISSVLFLMVVMTTSHLFFWDMWYIYYFWKAKIKGYQHLQSMESCYDAFI VYDTKNSAVTEWVLQELVAKLEDPREKHFNLCLEERDWLPGQPVLENLSQSIQLSKKTVFVMTQ KYAKTESFKMAFYLSHQRLLDEKVDVIILIFLEKPLQKSKFLQLRKRLCRSSVLEWPANPQAHP YFWQCLKNALTTDNHVAYSQMFKETV049648- / 618931 T19083WO001

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Hauser-Kawaguchi, A. “The Development of Platforms for Inhibiting RHAMM-HA Interactions & The Development Of An Optical Probe For Measuring Glomerular Filtration Rate” (2018) Electronic Thesis and Dissertation Repository.5576. 40. Ibrahim, J.et al., The Inflammation Superhighway: Tolls, Signals, and Pathways to Bronchopulmonary Dysplasia Updates on Neonatal Chronic Lung Disease, Chapter 9, pages 131-150 2020 41. Savani, R.C. Modulators of inflammation in Bronchopulmonary Dysplasia Seminars in Perinatology 42 (2018) 459-470049648- / 618931 T19083WO001 42. Garantziotis, S. and Savani, R. Am J. Physiol Cell Physiol 323:C202- C214, 2022 43. Kurimoto A, Hashimoto K, Nakamura T, et al. Synthesis and biological evaluation of 8-oxoadenine derivatives as toll-like receptor 7 agonists introducing the antedrug concept. J Med Chem.2010;53(7):2964-2972. 44. McCray PB, Jr., Pewe L, Wohlford-Lenane C, et al. 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[0214] Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be readily apparent to one of ordinary skill in the art in light of the teachings of this invention that certain changes and modifications may be made thereto without departing from the spirit or scope of the appended claims.

[0215] All publications, databases, GenBank and UniProt sequences, patents, and patent applications cited in this specification are herein incorporated by reference as if each was specifically and individually indicated to be incorporated by reference.

Claims

049648- / 618931 T19083WO001 WHAT IS CLAIMED IS:

1. A method of treating or effecting prophylaxis of a disease or disorder associated with inflammation in a subject, comprising administering to the subject a pharmaceutical composition comprising a RHAMM-derived peptide mimetic comprising an amino acid sequence of SEQ ID NO:3, wherein glutamic acid (E) at position 6 is covalently bonded to lysine (K) at position 10.

2. The method of claim 1, wherein the disease or disorder associated with inflammation is selected from the group consisting of sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome, TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS-CoV2 inflammation, SARS-CoV-2- mediated pneumonia, and a disorder associated with TLR activation, activation of the NLRP3 inflammasome and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells.

3. The method of claim 2, wherein the disease or disorder associated with inflammation is sepsis.

4. The method of claim 2, wherein the disease or disorder associated with inflammation is gram-negative pneumonia.

5. The method of claim 2, wherein the disease or disorder associated with inflammation is acute lung injury.

6. The method of claim 2, wherein the disease or disorder associated with inflammation is neonatal hyperoxia.

7. The method of claim 2, wherein the disease or disorder associated with inflammation is bronchopulmonary dysplasia.

8. The method of claim 2, wherein the disease or disorder associated with inflammation is acute respiratory distress syndrome.049648- / 618931 T19083WO001 9. The method of claim 2, wherein the disease or disorder associated with inflammation is TLR7-mediated pneumonia.

10. The method of claim 2, wherein the disease or disorder associated with inflammation is viral mediated inflammation.

11. The method of claim 2, wherein the disease or disorder associated with inflammation is influenza A inflammation.

12. The method of claim 2, wherein the disease or disorder associated with inflammation is influenza A pneumonia.

13. The method of claim 2, wherein the disease or disorder associated with inflammation is COVID-19 pneumonia.

14. The method of claim 2, wherein the disease or disorder associated with inflammation is SARS-CoV2 inflammation.

15. The method of claim 2, wherein the disease or disorder associated with inflammation is SARS-CoV-2-mediated pneumonia.

16. The method of claim 2, wherein the disorder associated with TLR activation, activation of the NLRP3 inflammasome and / or increased hyaluronan concentrations, inflammatory cytokines and inflammatory cells is acute kidney injury, acute liver injury, immune-mediated nervous system injury, necrotizing enterocolitis, inflammatory bowel disease, gout, traumatic brain injury, white matter brain injury, and wound healing.

17. The method of claim16, wherein the immune-mediated nervous system injury is multiple sclerosis.

18. The method of claim16, wherein the inflammatory bowel disease is Crohn’s disease or ulcerative colitis.

19. A method of treating or effecting prophylaxis of a disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation in a subject, comprising049648- / 618931 T19083WO001 administering to the subject a pharmaceutical composition comprising a RHAMM-derived peptide mimetic comprising an amino acid sequence of SEQ ID NO:3, wherein glutamic acid (E) at position 6 is covalently bonded to lysine (K) at position 10.

20. The method of claim 19, wherein the NLRP3 inflammasome activation results in elevated hyaluronan concentrations and inflammation in the subject.

21. The method of claim 19, wherein the disease or disorder associated with TLR signaling or NLRP3 inflammasome activation is selected from the group consisting of sepsis, gram-negative pneumonia, acute lung injury, neonatal hyperoxia, bronchopulmonary dysplasia, acute respiratory distress syndrome, TLR7-mediated pneumonia, viral mediated inflammation, influenza A inflammation, influenza A pneumonia, COVID-19 pneumonia, SARS- CoV2 inflammation, SARS-CoV-2-mediated pneumonia, acute kidney injury, acute liver injury, immune-mediated nervous system injury, necrotizing enterocolitis, inflammatory bowel disease, gout, traumatic brain injury, white matter brain injury, and wound healing.

22. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is sepsis.

23. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is gram-negative pneumonia.

24. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is acute lung injury.

25. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is neonatal hyperoxia.

26. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is bronchopulmonary dysplasia.

27. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is acute respiratory distress syndrome.049648- / 618931 T19083WO001 28. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is TLR7-mediated pneumonia.

29. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is viral mediated inflammation.

30. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is influenza A inflammation.

31. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is influenza A pneumonia.

32. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is COVID-19 pneumonia.

33. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is SARS-CoV2 inflammation.

34. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is SARS-CoV-2-mediated pneumonia.

35. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is acute kidney injury.

36. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is acute liver injury, 37. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is immune-mediated nervous system injury.

38. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is necrotizing enterocolitis.

39. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is inflammatory bowel disease.049648- / 618931 T19083WO001 40. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is gout.

41. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is traumatic brain injury.

42. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is white matter brain injury.

43. The method of claim 21, wherein the disease or disorder associated with TLR signaling and / or NLRP3 inflammasome activation is wound healing.

44. The method of claim 21, wherein the immune-mediated nervous system injury is multiple sclerosis.

45. The method of claim 21, wherein the inflammatory bowel disease is Crohn’s disease or ulcerative colitis.

46. The method of any one of claims 19-45, wherein TLR signaling and / or NLRP3 inflammasome activation is blocked 47. The method of claim 46, wherein blockage of TLR signaling and / or NLRP3 inflammasome activation is measured by lack of production of pro-IL1ß, lack of production of mature IL1ß, inhibition of inflammation, and / or inhibition of cytokine elaboration.

48. The method of claim 19, wherein TLR4 signaling and / or NLRP3 inflammasome activation is blocked.

49. The method of claim 19, wherein TLR7 signaling and / or NLRP3 inflammasome activation is blocked.

50. The method of any preceding claim, wherein the subject is a mammal.

51. The method of claim 50, wherein the mammal is a primate.

52. The method of claim 50, wherein the mammal is a non-human primate.049648- / 618931 T19083WO001 53. The method of claim 50, wherein the mammal is a human.

54. The method of claim 53, wherein the human is a neonate, child, or adult.

55. The method of claim 54, wherein the human is a neonate.

56. The method of claim 54, wherein the human is a child.

57. The method of claim 54, wherein the human is an adult.

58. The method of claim 50, wherein the mammal is a rodent.

59. The method of claim 58, wherein the rodent is a mouse, rat, guinea pig, hamster, or gerbil.

60. The method of claim 50, wherein the mammal is selected from the group consisting of human, baboon, chimpanzee, monkey, cynomolgus, marmoset, rhesus, mouse, rat, rabbit, cat, dog, horse, cow, sheep, goat, pig, ferret, guinea pig, hamster, gerbil, and bat.

61. The method of any one of claims 1-49, wherein the subject is a bird.

62. The method of claim 61, wherein the bird is a chicken.

63. The method of any preceding claim, wherein the pharmaceutical composition is administered intravenously.

64. The method of any one of claims 1-62, wherein the pharmaceutical composition is administered subcutaneously. 65 The method of any one of claims 1-62, wherein the pharmaceutical composition is administered by inhalation.

66. The method of any preceding claim, wherein the pharmaceutical composition is administered at a dosage of about 0.0002 mg / kg of subject body weight to about 20 mg / kg of subject weight.049648- / 618931 T19083WO001 67. The method claim 66, wherein the pharmaceutical composition is administered at a dosage of about 0.02 mg / kg of subject body weight.

68. The method of claim 66, wherein the pharmaceutical composition is administered at a dosage of about 0.2 mg / kg of subject body weight.

69. The method of claim 66, wherein the pharmaceutical composition is administered at a dosage of about 20 mg / kg of subject body weight.

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