Materials and methods for the prevention and treatment of viral diseases

Abstract

This disclosure provides peptides and use thereof to prevent or treat diseases caused by viruses.

Description

[0001] MATERIALS AND METHODS FOR THE PREVENTION

[0002] AND TREATMENT OF VIRAL DISEASES

[0003] FIELD OF INVENTION

[0004]

[0001] This invention is generally in the field of delivery of peptides to treat or prevent disease or disorders.

[0005] STATEMENT OF GOVERNMENT SUPPORT

[0006] [2] This invention was made with government support under grant number 7200AA22CA00002 awarded by the United States Agency for International Development. The government has certain rights in the invention.

[0007] INCORPORATION BY REFERENCE OF MATERIAL SUBMITTED ELECTRONICALLY

[0008] [3] This application contains, as a separate part of the disclosure, a Sequence Listing in computer-readable form which is incorporated by reference in its entirety and identified as follows: 55885_SeqListing.xml; Size: 419,212 bytes; Created: June 24, 2025.

[0009] BACKGROUND

[0010] [4] The global threat posed by existing and emerging viral epidemics represents one of the most critical global public health concerns. While the field of viral therapeutics has advanced in response to this pressing demand, new safe and effective antiviral agents for the prevention and treatment of viral infections are needed urgently.

[0011] [5] Of particular concern are enveloped viruses. An enveloped virus is made up of a nucleoprotein core, surrounded by a lipoprotein envelope having a closed lipid bilayer. The lipid is derived from the host cell's membrane(s), with glycoprotein on the outside and matrix protein or nucleocapsid protein on the inside. Exemplary enveloped virus families include Togaviridae, Flaviviridae, Coronaviridae, Rhabdoviridae, Filoviridae, Paramyxoviridae, Orthomyxoviridae, Bunyaviridae, Arenaviridae, Retroviridae, Herpesviridae, Poxviridae and some members of the Iridoviridae. These viruses are responsible for a range of diseases, including: encephalitis, intestinal infections, immunosuppressive disease, respiratory disease, hepatitis, and pox infections. Examples of enveloped viruses include: human immunodeficiency virus (HIV), herpes simplex viruses (HSV), hepatitis B virus (HBV), hepatitis C virus (HCV), dengue virus, West Nile virus, measles virus, and Ebola virus, among others.

[0012] [6] Non-enveloped viruses also represent a significant concern. For example, the Picornaviridae family include polioviruses, hepatitis A virus, and coxsackieviruses that may cause hand, foot, and mouth disease (HFMD), as well as disease of muscles, lungs, and heart. Papi llomavi ridae is a family of enveloped viruses that contains over 100 human papillomaviruses (HPV), the most common sexually transmitted infection.

[0013] [7] Most antiviral agents are specific to one pathogen, making the development of a drug stockpile for the prevention and treatment of emerging new viruses a significant challenge. There is also a need to prevent and / or treat multiple, intersecting viral pathogens, such as sexually transmitted infection that can comprise HIV, HSV, and HPV. In this regard, new classes of antiviral agents, to be used alone or in combination with existing antiviral agents and therapeutics, are needed. Ideally, the new agent(s) would meet one or more of the following criteria: be safe locally and systemically; act as early as possible in the viral life cycle; be as virus-specific as possible (i.e., attack a target specific to the virus but not the host); render the intact virus noninfectious; prevent the death or dysfunction of virus-infected cells; prevent further production of virus from infected cells; prevent spread of virus infection to uninfected cells; be potent and active against the broadest possible range of strains and isolates of a given virus; be efficacious against emerging new strains (i.e., the agent should not lead to viral resistance); be resistant to degradation under physiological and rigorous environmental conditions; and / or be readily and inexpensively produced.

[0014] [8] In view of the foregoing, there is a need in the art for new methods and compositions for inhibiting viral infection. The disclosure provides such methods and compositions. These and other advantages, as well as additional inventive features, will become apparent from the description provided herein.

[0015] SUMMARY

[0016] [9] The disclosure provides a peptide comprising the amino acid sequence SWLRX5IWEWIX11EVLX15EX17X18 (SEQ ID NO: 147), wherein X5 is A, V, or L; Xu is S, D-serine, or phospho-serine; X15 is S, D-serine, or phosphoserine; X17 is F or phospho-tyrosine; and X18-X20 are independently selected from R or phospho-tyrosine. In various aspects, X5 is A. Optionally, Xu is S and X15 is D-serine or Xu is D-serine and X15 is S; in various aspects, X11 is S and X15 is D-serine. X17 is optionally F. Xis is optionally R or K. In various aspects, the peptide further comprises one or more C-terminal amino acids. In this regard, the peptide optionally comprises X19 which may be R, and may further comprise X20 which is optionally R. In an exemplary aspect, the peptide comprises the amino acid sequence SWLRAIWEWISEVLsEFRRR (SEQ ID NO: 149). In various aspects, the peptide comprises an amide group at the C-terminus. In this regard, the disclosure provides a peptide comprising the amino acid sequence SWLRAIWEWISEVLsEFRRR-NF (SEQ ID NO: 150). Also provided herein are methods of treating or preventing a viral disease, the methods comprising administering the peptide to a subject in need thereof. Also provided are compositions comprising the peptide and a drug delivery device (e.g., an implantable drug delivery device) comprising the peptide.

[0017]

[0010] The disclosure also contemplates use of the peptide described herein in treating or preventing a viral disease, as well as use of the peptide described herein in the preparation of a medicament for treating or preventing a viral disease in a subject in need thereof. Additionally provided is the peptide described herein for use in treating or preventing a viral disease in a subject in need thereof.

[0018] DESCRIPTION OF THE FIGURES

[0019]

[0011] FIG. 1 illustrates exemplary peptide conjugates based on the peptide 346-001 backbone. Conj, denotes conjugate. Peptide 346-001 backbone is illustrated merely to provide reference for different conjugates, which may be based on any amino acid sequence provided herein.

[0012] FIGs. 2A-C illustrate exemplary peptide N-conjugates via a lysine linker based on the peptide 346-001 backbone (SEQ ID NO: 147). Peptide 346-001 backbone is illustrated merely to provide reference for different conjugates, which may be based on any amino acid sequence provided herein.

[0020]

[0013] FIGs. 3A-C illustrate exemplary peptide O-conjugates via a serine linker based on the peptide 346-001 backbone (SEQ ID NO: 148). Peptide 346-001 backbone is illustrated merely to provide reference for different conjugates, which may be based on any amino acid sequence provided herein.

[0021]

[0014] FIGs. 4A-G illustrate exemplary peptide S-conjugates via a cysteine linker based on the peptide 346- 001 backbone (SEQ ID NO: 149). Peptide 346-001 backbone is illustrated merely to provide reference for different conjugates, which may be based on any amino acid sequence provided herein.

[0022]

[0015] FIG. 5 is a bar graph illustrating the EC50 (micromolar) of various peptide variants against HIV-1 HIB in TZM-bl-FcRI cells. The peptide variants screened were based on SEQ ID NO: 149. A peptide comprising the amino acid sequence of SEQ ID NO: 149 was examined alongside a series of variants wherein the amino acid at position 5 (X5) was substituted with the amino acids set forth on the x-axis of the graph; the rest of the amino acid sequence of SEQ ID NO: 149 was unchanged. The peptide comprising SEQ ID NO: 149 exhibited the highest degree of potency, and lipophilic groups demonstrated a beneficial impact on potency (e.g., 5-Val and 5- Leu). A higher degree of lipophilicity introduced at position 5 (e.g., lie at position 5) reduced potency compared to SEQ ID NO: 149.

[0023]

[0016] FIGs. 6A-6C are graphs illustrating the stability of antiviral peptides in human cervicovaginal fluids from multiple donors (mean ± SEM, N = 3 per group); dotted horizontal line indicates 50% degradation. (A) Grey circles, peptide 346-203 (H-swlraiwewisevlsefrrr-NH2, TFA salt); black squares, peptide 346-192 (H- SWLRAIWEWISEVLSEFRRR-NH2, acetate salt) (SEQ ID NO: 236). (B) Grey circles, peptide 346-203 (H- swlraiwewisevlsefrrr-NH2, TFA salt) (SEQ ID NO: 248); black triangles, peptide 346-232 (H- SWLRAIWEWISEVLSEFRRR-NH2, acetate salt) (SEQ ID NO: 277), two combined, separate experiments. (C) Grey circles, peptide 346-203 (H-swlraiwewisevlsefrrr-NH2, TFA salt) (SEQ ID NO: 248); black diamonds, peptide 346-212 (H-SWLRAIWEWIsEVLsEFRRR-NH2, TFA salt) (SEQ ID NO: 257).

[0024]

[0017] FIG. 7 is a line graph illustrating pharmacokinetics of single bolus dose IV administration of peptide 346- 232 (H-SWLRAIWEWISEVLSEFRRR-NH2, acetate salt)(SEQ ID NO: 277) in female beagle dogs (N = 4 per group). Grey circles represent peptide 346-232 plasma concentrations following IV dosing at 15 mg kg-1 and black circles represent dosing at 44 mg kg-1 .

[0025] DETAILED DESCRIPTION

[0026]

[0018] The disclosure provides peptides, devices, systems and methods for treating, preventing, reducing the likelihood of having, reducing the severity of and / or slowing the progression of a condition in a subject. In various aspects, the disclosure provides materials and methods designed for the prevention and treatment of viral disease, specifically a viral disease caused by a virus. Aspects of the materials and methods include, but are not limited to:

[0027] • Peptides, or peptide prodrugs, or peptide conjugates, comprising 5-50 amino acids with antimicrobial properties;

[0028] • Optionally, one or more other active pharmaceutical ingredients (APIs) (also referred to herein as "complementary agents”) used in combination with the above peptide; and / or

[0029] • Drug delivery systems that result in a pharmacologically desirable amount of the peptide(s) (and, optionally API (s)) in the target anatomic compartment.

[0030]

[0019] Thus, in various aspects, the disclosure provides a peptide comprising the sequence SWLRX5IWEWIX11EVLXI5EX17X18 (SEQ ID NO: 147), wherein X5is A, V, or L; Xu is S, D-serine, or phosphoserine; X15 is S, D-serine, or phospho-serine; X17 is F or phospho-tyrosine; and X18-X20 are independently selected from R or phospho-tyrosine. In various aspects, X5 is A. Optionally, Xu is S and X15 is D-serine or Xu is D-serine and X15 is S; in various aspects, Xu is S and X15 is D-serine. X17 is optionally F and Xis is optionally R or K. In various aspects, the peptide further comprises one or more additional amino acids at the C-terminus, such as one or two additional amino acids at the C-terminus. For instance, the peptide may comprise the sequence SWLRX5lWEWIXnEVLX15EXi7Xi8Xi9X2o (SEQ ID NO: 148). In this respect, the peptide optionally comprises X19 which is R and may further comprise X20 which is optionally R. In an exemplary aspect, the peptide comprises the amino acid sequence SWLRAIWEWISEVLsEFRRR (SEQ ID NO: 149). In various aspects, the peptide comprises an amide group at the C-terminus. Optionally, the peptide comprises the amino acid sequence of SEQ ID NO: 150.

[0031]

[0020] The disclosure further provides methods of treating or preventing a viral disease. The methods comprise administering to a subject in need thereof a peptide comprising (or consisting essentially of, or consisting of) the sequence SWLRX5IWEWIX11EVLXI5EX17X18 (SEQ ID NO: 147) or the sequence SWLRX5IWEWIX11EVLXI5EX17X18X19X20 (SEQ ID NO: 148). The disclosure further provides compositions comprising any one or more of the peptides described herein. It will be appreciated that description of peptides herein is applicable to both descriptions of compositions and methods of treating or preventing a viral disease.

[0032]

[0021] The administration can be achieved by a range of parenteral routes of administration, including but not limited to: intravenous, subdermal, intramuscular, buccal (i.e., via the buccal mucosa), intranasal, pulmonary, and topical, including but not limited applied to the skin, vaginally, and rectally. Optionally, the method comprises further administering one or more complementary agents (i.e., other APIs) suitable for, e.g., the treatment or prevention of a medical condition, such as a disease caused by viruses or related illness or symptom.

[0033]

[0022] It will be appreciated that "treating” a disease or disorder does not require 100% abolition of the disease or disorder (i.e., complete reversal of the disease). Any degree of "treatment” is contemplated by the disclosure, including lessening of one or more symptoms of the disease, reduction in viral load, improvement in quality of life, and the like. Similarly, it will be appreciated that "preventing” a disease, in the context of the disclosure, does not require 100% inhibition of appearance of the disease. Any degree of reduction in the initial appearance of symptoms, inhibition of viral infection, prolonging relapse, inhibiting an initial surge of viral load, and the like, are contemplated. In various aspects, the viral disease is an HIV infection, e.g., an HIV-1 or HIV-2 infection. Alternatively, the viral disease may be an HSV infection, such as an HSV-1 or HSV-2 infection. Alternatively, viral disease is a viral respiratory disease, such as a coronavirus infection (e.g., SARS-CoV-2 infection), an influenza infection, or a respiratory syncytial virus (RSV) infection.

[0034]

[0023] In the context of the disclosure, the subject is a mammal, which refers to any member of the class Mammalia, including, without limitation, humans and nonhuman primates such as chimpanzees and other apes and monkey species; farm animals such as cattle, sheep, pigs, goats and horses; domesticated mammals, such as dogs and cats; laboratory animals including rodents such as mice, rats and guinea pigs, and the like. The term does not denote a particular age or sex. Thus, adults (i.e. , human subjects aged 18 years or more), children (i.e., human subjects aged one to eighteen years) and newborns (i.e., human subjects aged one year or less), whether male or female, are intended to be included within the scope of ''subject.”

[0035] Antiviral Peptides

[0036]

[0024] The disclosure provides, e.g., peptides with antiviral properties and methods of use thereof. Optionally, the peptides have a length of five to fifty amino acid residues (e.g., five to eight amino acid residues; or eight to twelve amino acid residues; or twelve to twenty amino acid residues; or twenty to thirty five amino acid residues; or thirty five to fifty amino acid residues). In various aspects, the amino acid sequence is a virucidal amphipathic o-helical peptide.

[0037]

[0025] The disclosure contemplates a peptide derived from a peptide comprising the amino acid sequence SWLRAIWEWISEVLsEFRRR (SEQ ID NO: 149). In this regard, the disclosure contemplates, e.g., a variant of a peptide comprising the amino acid sequence of SEQ ID NO: 149, such as peptide comprising, e.g., one, two, or three amino acid substitutions (such as conservative substitutions) compared to SEQ ID NO: 149. In other aspects, the disclosure contemplates variants of a peptide comprising the amino acid sequence of SEQ ID NO: 149 comprising four or five amino acid substitutions. Optionally, the peptide comprises no more than seven, no more than six, no more than five, no more than four, no more than three, no more than two, or only one amino acid substitute compared SEQ ID NO: 149. For example, substitutions (e.g., conservative substitutions), deletions and additions may be made at non-critical residue positions within the selected peptide without substantially adversely affecting its biological activity. Modifications can be made at the receptor binding site(s) to adjust binding efficiency and specificity. In addition, changes may be made to select residues to increase peptide stability (e.g., replacing a cysteine residue with another amino acid, such as serine). The peptide can be optionally flanked and / or modified at one or both of the N- and C-termini, as desired. For instance, the peptide may lack the C-terminal two amino acids of SEQ ID NO: 149.

[0038]

[0026] Optionally, the peptide comprises the amino acid sequence of SEQ ID NO: 149 comprising one, two, or three substitutions (or more) selected from the following: the S at position 1 (Xi) is substituted with K, D, A, N, M, T, or V; the W at position 2 (X2) is substituted with I, Y, F, P, L, V, H, M, A, T, or S; the L at position 3 (X3) is substituted with W, I, M, V, F, M, A, or T; the R at position 4 (X4) is substituted with C, K, Q, H, P, or C; the A at position 5 (X5) is substituted with V or L; the I at position 6 (Xs) is substituted with W, V, L, or F; the W at position 7 (X7) is substituted with V, Y, F, P, L, V, H, or D; the E at position 8 (Xs) is substituted with D, N, Y, or S; the W at position 9 (Xg) is substituted with L, Y, F, P, L, V, or H; the I at position 10 (X10) is substituted with V, L, or F; the S at position 11 (Xu) is substituted with C, A, P, M, H, or T; the E at position 12 (X12) is substituted with D, S, Q, Y, or T; the V at position 13 (X13) is substituted with I, L, A, F, or M; the L at position 14 (X14) is substituted with V, I, M, or F; the s (D-serine) at position 15 (X15) is substituted with S, D, A, N, T, Y, or P; the E at position 16 (Xie) is substituted with D, N, Y, phospho-tyrosine, or S; the F at position 17 (X17) is substituted with W, Y, L, P, phosphor-tyrosine, or H; or the R at position 18 (Xis) is substituted with E, H, K, P, phospho-tyrosine, or Q; the R at position 19 (X19) is substituted with phospho-tyrosine; and the R at position 19 (X19) is substituted with phospho-tyrosine. In various aspects, X5 is A; Xu is S, D-serine, or phospho-serine (e.g., S); X15 is S, D-serine, or phospho-serine (e.g., D-serine); X17 is F or phospho-tyrosine (e.g., F); Xis is R or phospho-tyrosine (e.g., R); X19 is R or phospho-tyrosine (e.g., R); and X20 is R or phospho-tyrosine (e.g., R). Also optionally, the peptide includes D-forms of any of the amino acids described herein (e.g., all or a subset of the amino acids may be D- amino acids). Also optionally, the amino acids may be phosphorylated; for instance, S, T, Y, or H may be the phosphorylated versions of the amino acids, phospho-serine, phosphor-threonine, phosphor-tyrosine, or phosphor-histidine.

[0039]

[0027] In various aspects, the peptide has a sequence of Formula 1 : X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16-X17-X18-X19-X20 (Formula 1 ; SEQ ID NO: 151), wherein: Xi is S (including D-serine or phospho-serine), K, D, A, N, M, T (including phospho-threonine), or V; X2 is W, I, Y (including phospho-tyrosine), F, P, L, V, H (including phospho-histidine), M, A, T, or S (including D-serine or phospho-serine); X3 is L, W, I, M, V, F, M, A, or T (including phospho-threonine); X4 is R, C, K, Q, H (including phospho-histidine), P, or C; X5 is A, V, or L; Xe is I, W, V, L, or F; X7 is W, V, Y (including phospho-tyrosine), F, P, L, V, H (including phospho-histidine), or D; Xs is E, D, N, Y (including phospho-tyrosine), or S (including D-serine or phospho-serine); Xg is W, L, Y (including phospho-tyrosine), F, P, L, V, or H (including phospho-histidine); X10 is I, V, L, or F; Xu is S (including D-serine or phospho-serine), A, P, M, H (including phospho-histidine), C, or T (including phospho-threonine); X12 is E, D, S (including D-serine or phospho-serine), Q, Y (including phosphotyrosine), or T (including phospho-threonine); X13 is V, I, L, A, F, or M; X14 is L, V, I, M, or F; X15 is S (including D- serine or phospho-serine), D, A, N, T (including phospho-threonine), Y (including phospho-tyrosine), or P; X16 is E, D, N, Y, or S (including D-serine or phospho-serine); X17 is F, W, Y (including phospho-tyrosine), L, P, or H (including phospho-histidine); Xis is R, Y (including phospho-tyrosine), K, E, H (including phospho-histidine), P, or Q; X19 is R or Y (including phospho-tyrosine); and X20 is R or Y (including phospho-tyrosine). Optionally, Xi is S (or D-serine or phospho-serine); X2 is W; X3 is L; Xg is W; X12 is E; X14 is L; and X15 is S (or D-serine or phospho-serine).

[0028] Other peptides disclosed herein include a peptide comprising the amino acid sequence S-W-L-X4-X5-X6- X7-X8-W-X10-X11-E-X13-L-S-D-X17-X18 (Formula 2; SEQ ID NO: 2), wherein X4is R, G, A, I, L, V, or F; X5A, V, or L; Xe is I, A, W, V, L, or F; X7 is W, A, V, Y (including phospho-tyrosine), F, L, V, H (including phospho-histidine), or D; Xs is D, A, S (including phospho-serine), E, or Y; X10 is I, A, V, L, or F; Xu is C, S (including phosphoserine), A, W, M, H (including phospho-histidine), W, R, V, or T (including phospho-threonine); X13 is V, F, I, L, A, or M; X17 is F, W, Y (including phospho-tyrosine), L, or H (including phospho-histidine); and Xis is K, R, H (including phospho-histidine), A, P, or G. With respect to the peptide of Formula 2, when X4is not R, it is G, A, I, L, V, or F. When Xs is not I, it is A, W, V, L, or F, optionally W, V, L, or F. When X7 is not W, it is A, V, Y, F, L, V, H, or D; optionally V, Y, F, L, V, H, or D, optionally A or F. When Xs is not D, it is A, S, E, or Y, optionally, E, Y, or S. When X10 is not I, it is A, V, L, or F, optionally V, L, or F. When Xu is not C, it is S, A, W, M, H, W, R, V, or T, optionally S, A, M, H, or T. When X13 is not V, it is F, I, L, A, or M. When X17 is not F, it is W, Y, L, or H. When Xis is not K, it is R, H, A, P, or G, optionally R, H, or P. The sequence may include L- or D-forms of the amino acids, or any combination thereof. The sequence may include phosphorylated amino acids.

[0040]

[0029] In a preferred aspect, the disclosure provides a peptide comprising the amino acid sequence SWLRX5IWEWIX11EVLX15EX17X18 (SEQ ID NO: 147), wherein X5is A, V, or L; Xu is S, D-serine, or phosphoserine; X15 is S, D-serine, or phospho-serine; X17 is F or phospho-tyrosine; and Xis is R or phospho-tyrosine. In various aspects, X5 is A. Optionally, Xu is S and X15 is D-serine or Xu is D-serine and X15 is S. In various aspects, X11 is S and X15 is D-serine. X17 is optionally F, and Xis is optionally R, K, or phospho-tyrosine. In another preferred aspect, the disclosure provides a peptide comprising the amino acid sequence SWLRX5IWEWIX11EVLX15EX17X18X19X20 (SEQ ID NO: 148), wherein Xig-X2o are each optionally R or phosphotyrosine. In an exemplary aspect, the peptide comprises the amino acid sequence SWLRAIWEWISEVLsEFRRR (SEQ ID NO: 149). In various aspects, the peptide comprises an amide group at the C-terminus. Provided herein is a peptide comprising the amino acid sequence SWLRAIWEWISEVLsEFRRR-NFfe (SEQ ID NO: 150).

[0041]

[0030] The disclosure also contemplates peptides wherein the sequence set forth in herein is reversed (i.e., a retropeptide, such as a peptide comprising the reversed sequence of SEQ ID NO: 149 such that the N-terminus is the C terminus of the retropeptide).

[0042]

[0031] Peptide sequences derived from SEQ ID NO: 149 (including peptides comprising SEQ ID NO: 147 or SEQ ID NO: 148), and modifications thereof, may possess different, non-obvious pharmacologic properties relative to the parent sequence. These may consist of higher or lower in vitro and / or in vivo efficacy against one or more viruses that cause diseases. Other, exemplary properties that may be affected by modifying the peptide sequence include: in vitro stability; in vivo stability; in vivo half-life; protein binding; and cellular penetration.

[0043]

[0032] In non-limiting aspects of the disclosure, the peptide contains overall features that impact its efficacy against viruses. One exemplary, non-limiting feature is associated with the peptide's o-helical structure. Without wishing to be bound by any particular theory, the o-helical structure can allow the peptide to interact with the viral membrane and disrupt its integrity, thereby releasing viral components such as capsids and exposing the viral genetic material to exonuclease degradation. The charge distribution along the peptide's o-helix, determined by the nature of the amino acid side-groups in the sequence, is an exemplary feature that may determine how the peptide associates, binds, and / or disrupts the viral particle. Another exemplary, non-limiting feature concerns the peptide's amphipathic nature, defined commonly in the art as possessing both hydrophilic and lipophilic properties. Without wishing to be bound by any particular theory, the peptide's amphipathicity can disrupt the ligand-host receptor interaction and viral escape from endosomes. The overall peptide hydrophobicity is another exemplary feature that can impact its antiviral properties by determining how it recognizes host-cellular components of virus membranes, possibly by their lipid composition. Another exemplary, non-limiting feature is associated with specific peptide-membrane protein interaction, such that retropeptides, peptides with scrambled hydrophobic or hydrophilic amino acids, or peptides comprised containing D-amino acids, but all based on the sequence of the parent, active peptide (e.g., peptide comprising SEQ ID NO: 149), display antiviral properties. In summary, there are a number of features that can, in certain embodiments, affect the antiviral peptide's interaction with the viral particle, specifically the viral membrane or another feature on the virus surface, causing disruption and leading to virus inactivation. In some cases, destabilization of physical linkages between the mature conical capsid core and the viral membrane, i.e., the core-membrane linkage can occur. In other cases, the shedding of the viral surface protein(s) responsible for entry into cells can be altered or shed.

[0044]

[0033] Peptides are a series of amino acids connected one to the other by peptide bonds between the o-amino and o-carboxy groups of adjacent amino acids. Peptides can be a variety of lengths, either in their neutral (uncharged) forms or in forms which are salts, and either free of modifications such as glycosylation, side chain oxidation, or phosphorylation or containing these modifications, subject to the condition that the modification not destroy the biological activity of the polypeptides as herein described. Although the peptide, in various aspects, is substantially free of other naturally occurring proteins and fragments thereof, in some embodiments the peptides can be synthetically conjugated to native fragments or particles.

[0045]

[0034] In various aspects, the peptides are optionally polymerized, each to itself, to form larger homopolymers, or with different peptides to form heteropolymers. In some instances, peptides will be combined in a composition as an admixture and will not be linked. The peptide can also be conjugated to lipid-containing molecules or to different peptides.

[0046]

[0035] In various aspects of the disclosure, the peptide is chemically conjugated with a moiety that provides a functional or practical advantage. Non-limiting examples of such linked species, known in the art, include: polyethylene glycol, or other polymers to increase the in vivo residence time or half-life of the peptide; a small, biocompatible linker group that is amenable to high yielding bioconjugation reactions, so-called "click chemistry” linkers well known in the art (5-8) and fully incorporated herein; a biotin linker on the peptide that binds to streptavidin on another moiety, or other similar affinity-based linker systems known in the art, to facilitate in vivo detection or imaging of the peptide.

[0036] Linkages for homo- or hetero-polymers or for coupling to carriers can be provided in a variety of ways known in the art. For example, cysteine residues can be added at both the amino- and carboxy-termini, where the peptides are covalently bonded via controlled oxidation of the cysteine residues. Also useful are a large number of heterobifunctional agents which generate a disulfide link at one functional group end and a peptide link at the other, including N-succidimidyl-3-(2-pyridyl-dithio) proprionate (SPDP). This reagent creates a disulfide linkage between itself and a cysteine residue in one protein and an amide linkage through the amino on a lysine or other free amino group in the other.

[0047]

[0037] In various aspects, the pharmacology of the peptide is enhanced by synthesizing a suitable prodrug; methods of prodrug synthesis are known in the art (9, 10). Prodrugs of the peptide can involve the C-terminal carboxy group, the tyrosine phenol group in tyrosyl peptides, and the N-terminal amino group. The peptide prodrug can have improved pharmacological properties when compared to the parent drug, such as improved bioavailability to the target compartment.

[0048]

[0038] In some instances, it is desirable to administer the antimicrobial peptides disclosed herein as enzyme- cleavable polymeric prodrugs with tailored release profiles, such as the "drugamer” class (60) described in the art for pulmonary- {61, 62) and liver-targeted delivery (63) as nonlimiting examples. In one embodiment, peptide 346-232 (H-SWLRAIWEWISEVLsEFRRR-NF ) is conjugated to a drugamer as an injectable, liver-targeting enzyme-cleavable polymeric prodrug as a long-acting therapeutic for the treatment of HBV infection. In another embodiment, the antimicrobial peptide is conjugated as a prodrug to a targeted drug delivery system aimed at concentrating said peptide in a predetermined anatomic compartment {e.g., liver, lung, kidney, brain, etc.). Targeted drug delivery using a prodrug approach is well-known in the art and can be leveraged for the embodiment described here. For example, prodrugs for liver-targeted drug delivery have been described in Erion (64). In another embodiment, targeted drug delivery is achieved by conjugating the antimicrobial peptide to a nanoparticle with the appropriate size and / or surface features, as known in the art.

[0049]

[0039] In various aspects of the disclosure, the pharmacology of the peptide is modified (e.g., enhanced) by synthesizing a suitable conjugate with a lipophilic moiety, or an acceptable salt thereof, as shown schematically in TABLE 2 using peptide 346-001 as an exemplary backbone. It is understood that the approaches described herein illustrated using peptide 346-001 also apply to other peptides described herein (e.g., a peptide comprising SEQ ID NO: 147 or SEQ ID NO: 149). In accordance with TABLE 2, peptide conjugation can occur in a variety of non-limiting strategies. Conjugation can be beneficial at the N-terminus, at the C-terminus, or at both termini. Conjugation can also be beneficial at the reactive side chains of the peptide, as shown in FIG 1 using peptide 346-001 as an exemplary backbone. Any combination of these conjugation strategies can be used to enhance the properties of the parent peptide backbone. The conjugates are prepared using organic synthesis strategies, methods, and processes, many of which are known in the art. In various aspects, the disclosure provides the peptide described herein which is modified with a lipophilic moiety via a linker bonded to an S or E within the amino acid sequence.

[0040] The disclosure further contemplates peptides which are not conjugated with a lipophilic moiety.

[0050]

[0041] In various aspects of the disclosure, the pharmacology of the peptide is modified (e.g., enhanced) by synthesizing a suitable conjugate with a second, complementary peptide. Conjugation can be beneficial at the Interminus, at the C-terminus, or at both termini. Conjugation can also be beneficial at the reactive side chains of the peptide, as shown in FIG 1 using peptide 346-001 as an exemplary backbone. Any combination of these conjugation strategies can be used to enhance the properties of the parent peptide backbone. The conjugates are prepared using organic synthesis strategies, methods, and processes, many of which are known in the art.

[0051]

[0042] In various aspects, conjugation is achieved directly to the peptide. In various aspects, conjugation is achieved via one or more linker groups. Optionally, the linker, or spacer, consists of a single amino acid. Alternatively, the linker optionally comprises two or more amino acids, such as GSG, multiples thereof (GSG)n, or GSGSGC (SEQ ID NO: 150), known in the art. As described herein, linkers can comprise peptides, polyether compounds (e.g., PEG), and / or combinations thereof. In applications where it is beneficial to have longer distance between the conjugate and the antiviral peptide, polyethylene glycol (PEG) chains of varying lengths may be employed.

[0052]

[0043] In non-limiting examples, the complementary peptide moiety conjugated with the peptide described herein is a cell-targeting peptide, such as a cell-targeting peptide known in the art. Non-limiting examples of celltargeting peptides include peptides derived from the V3 loop of gp120, the surface glycoprotein of HIV-1, such as the following nonlimiting exemplary sequences:

[0053] HIV-1 gp120 V3-A, RKSIHIGPGRAFYTTG (SEQ ID NO: 117)

[0054] HIV-1 gp120 V3-B, RKGIRIGPGRAVYAAE (SEQ ID NO: 118)

[0055]

[0044] In another non-limiting example, the cell-targeting peptide comprises (or consists of) Tuftsin (TKPR (SEQ ID NO: 120)) or the canine analogs (TKPK (SEQ ID NO: 121), TKPKG (SEQ ID NO: 119)). In another nonlimiting embodiment, the Tuftsin sequence is incorporated into the peptide 346 amino acid backbone. Nonlimiting examples include:

[0056] X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-T-K-P-R (SEQ ID NO: 122) Xi-X2-X3-X4-X5-X6-X7-X8-X9-Xio-T-K-P-R-Xi5-Xi6-Xi7-Xi8 (SEQ ID NO: 123) T-K-P-R-X5-X6-X7-X8-X9-Xio-Xii-Xi2-Xi3-Xi4-Xi5-Xi6-Xi7-Xi8 (SEQ ID NO: 124)

[0057]

[0045] Another non-limiting example, the complementary peptide moiety comprises a cell-penetrating peptide, such as a cell-penetrating peptide known in the art, such as the following nonlimiting exemplary sequences:

[0058] Tat48-60, GRKKRRQRRRPPQ (SEQ ID NO: 125)

[0059] HIV-1 Tat-(49-57), RKKRRQRRR (SEQ ID NO: 126)

[0060] HIV-1 Rev-(34-50), TRQARRNRRRRWRERQR (SEQ ID NO: 127) FHV Coat-(35-49), RRRRNRTRRNRRRVR (SEQ ID NO: 128) Penetratin, RQIKIWFQNRRMKWKK (SEQ ID NO: 129) DPV3, RKKRRRESRKKRRRES (SEQ ID NO: 130)

[0061] CADY, GLWRALWRLLRSLWRLLWRA (SEQ ID NO: 131)

[0062] MAP, KLALKLALKALKAALKA (SEQ ID NO: 132)

[0063] Maurocalcine, GDCLPHLKLCKENKDCCSKKCKRRGTNIEKRCR (SEQ ID NO: 133) pVEC, LLIILRRRIRKQAHAHSK (SEQ ID NO: 134) TP10, AGYLLGKINLKALAALAKKIL (SEQ ID NO: 135) TP2, PLIYLRLLRGQF (SEQ ID NO: 136) Pep-1, KETWWETWWTEWSQPKKKRKV (SEQ ID NO: 137) Pep-7, SDLWEMMMVSLACQY (SEQ ID NO: 138) p28, LSTAADMQGWTDGMASGLDKDYLKPDD (SEQ ID NO: 139) Transportan, GWTLNSAGYLLGKINLKALAALAKKIL (SEQ ID NO: 140) C105Y, CSIPPEVKFNKPFVYLI (SEQ ID NO: 141) KFGF, AAVLLPVLLAAP (SEQ ID NO: 142) Polyarginine, (R)n4 < n < 12

[0064]

[0046] In non-limiting examples, the sequence of the complementary peptide moiety is reversed (i.e., a retropeptide, such that the N-terminus of the complementary peptides listed above is the C-terminus of the retropeptide). In other, nonlimiting examples, one or more amino acids in the complementary peptide sequence are D-amino acids.

[0065]

[0047] In one various aspects, conjugation is achieved directly to the peptide. In various aspects, conjugation is achieved via one or more linker groups. Optionally, the linker, or spacer, consists of a single amino acid.

[0066] Alternatively, the linker optionally comprises two or more amino acids, such as GSG, multiples thereof (GSG)n, or GSGSGC (SEQ ID NO: 150), known in the art. As described herein, linkers can comprise peptides, polyether compounds (i.e., PEG), and / or combinations thereof. In applications where it is beneficial to have longer distance between the conjugate and the antiviral peptide, polyethylene glycol (PEG) chains of varying lengths. PEG is a typically biologically inert chemical that confers greater water solubility to peptides with which it is incorporated as constituent chemical group. PEG is non-toxic and non-immunogenic, hydrophilic, and highly flexible. In additional embodiments, the linker comprises one or more polyethylene glycol oligomer moieties having a formula of -(OCFhCFDm-, wherein m is an integer 1 to 24. In one exemplary embodiment, m is 24-1,000. In another embodiment, m is 1,000-5,000.

[0067]

[0048] In accordance with this aspect of the present disclosure, linkers may be an amino acid, such as, but not limited to, lysine, serine, aspartic acid, glutamic acid, or cysteine.

[0068]

[0049] When lysine is used as a linker, either as a single amino acid or as part of a larger linker comprising two or more amino acids, synthetic spacers (e.g., PEG) or a combination thereof, the amino side-chain can be used for conjugation as shown in FIG 2 using the peptide 346-001 backbone as a non-limiting example for illustrative purposes only. Suitable conjugates can be carbonyl compounds, as depicted in FIG 2 (A) and (B). In such non- limiting embodiments, Y may be 0, S, NRi (where Ri is H, alkyl, or aryl), P(0)R2 [where R2 is alkyl, alkyl-aryl (e.g., benzyl), aryl, OH, O-alkyl, O-alkyl-aryl, O-aryl, NRi-alkyl, NRi-alkyl-aryl, or NRi-aryl], or CR2R3 [where R2 and or R3 are alkyl, alkyl-aryl (e.g., benzyl), aryl, O-alkyl, O-alkyl-aryl, O-aryl, or NRi-alkyl, NRi-alkyl-aryl, or NRi- aryl], In other words, as described herein, exemplary, non-limiting conjugates as shown in FIG 2 can be classified as amides, carbamates, ureas, thiocarbamates, phosphoramidates, and phosphonates. The alkyl chain length, n, is 1-50 and R is alkyl, aryl, or derivatives thereof, such as PEG. In one non-limiting example, palmitic acid is conjugated with the lysine linker; i.e., Y = CH2, n = 13, R = CH3. In non-limiting embodiments, the conjugation moiety is unsaturated, such as oleic acid (R = CH3) as shown in FIG 2 (B), or linoleic acid, and comprises one or more double bonds. In other non-limiting examples, the lysine NH2 side chain is conjugated via a methylene group, as shown in FIG 2 (C), and Y can be 0, S, NR1, P(O)R2, or CR2R3.

[0069]

[0050] In non-limiting examples the moiety conjugated to X via the 0=C-Y- group as described above and in FIG 2 (A), or via the CH2-Y- group as described above in FIG 2 (C) comprises one or more cholesterol groups, or derivatives thereof. In further non-limiting examples, the moiety comprises of one or more tocopherol groups, or derivatives thereof. Tocopherols constitute a series of related benzopyranols (or methyl tocols) that occur in plant tissues and vegetable oils and are powerful lipid-soluble antioxidants. These compounds are produced by plants and other oxygenic photosynthetic organisms, such as algae and some cyanobacteria, and are essential components of the diet of animals, and collectively they are termed "vitamin E”. The antiviral peptide conjugate of the present disclosure includes tocopherol, a tocopherol derivative, or pharmaceutically acceptable salt thereof. Examples of tocopherol that may be used in the present aspect include, but are not limited to, o-tocopherol, |3- tocopherol, y-tocopherol, and 5-tocopherol. Examples of useful tocopherol derivatives or pharmaceutically acceptable salts thereof include, but are not limited to, tocotrienols, tocopheroxyl radical, 8o- alkyldioxytocopherone, tocopherol quinone, tocopherol hydroquinone, D, / .-tocopherol, D, 1-tocopheryl acetate, o- tocopheryl acetate, o-tocopheryl nicotinate, and o-tocopheryl succinate. In further non-limiting examples, the moiety comprises one or more dihydrosphingosine groups, one or more sphingosine groups, or derivatives thereof. In further non-limiting examples, the moiety comprises one or more phospholipid groups, or derivatives thereof.

[0070]

[0051] When serine is used as a linker, either as a single amino acid or as part of a larger linker comprising two or more amino acids, synthetic spacers (e.g., PEG) or a combination thereof, the hydroxy side-chain can be used for conjugation as shown in FIG 3 using the peptide 346-001 backbone as a non-limiting example for illustrative purposes only. Suitable conjugates include carbonyl compounds, as depicted in FIG 3 (A) and (B). In such nonlimiting embodiments, Y may be 0, S, NR1 [where R1 is H, alkyl, alkyl-aryl (e.g., benzyl), or aryl], P(O)R2 [where R2 is alkyl, alkyl-aryl (e.g., benzyl), aryl, OH, O-alkyl, O-alkyl-aryl, O-aryl, NRi-alkyl, NRi-alkyl-aryl, or NRi-aryl], or CR2R3 [where R2 and or R3 are H, alkyl, alkyl-aryl (e.g., benzyl), aryl, O-alkyl, O-alkyl-aryl, O-aryl, or NRi-alkyl, NRi-alkyl-aryl, or NRi-aryl], In other words, as described herein, exemplary, non-limiting conjugates as shown in FIG 2 can be classified as esters, carbonates, carbamates, thiocarbonates, phosphoramidates, and phosphonates. The alkyl chain length, n, is 1-50 and R is alkyl, aryl, or derivatives thereof, such as PEG. In one non-limiting example, palmitic acid is conjugated with the serine linker; i.e., Y = CH2, n = 13, R = CH3. In nonlimiting embodiments, the conjugation moiety is unsaturated, such as oleic acid (R = CH3) as shown in FIG 3 (B), or linoleic acid, and comprises or more double bonds. In other non-limiting examples, the serine OH side chain is conjugated via a methylene group, as shown in FIG 3 (C), and Y can be 0, S, NR1, P(0)R2, or CR2R3.

[0071]

[0052] In non-limiting examples the moiety conjugated to X via the 0=C-Y- group as described above and in FIG 3 (A), or via the CH2-Y- group as described above in FIG 3 (C) comprises one or more cholesterol groups, or derivatives thereof. In further non-limiting examples, the moiety comprises one or more tocopherol groups, or derivatives thereof. In further non-limiting examples, the moiety comprises one or more dihydrosphingosine groups, one or more sphingosine groups, or derivatives thereof. In further non-limiting examples, the moiety comprises one or more phospholipid groups, or derivatives thereof.

[0072]

[0053] In general, thiol groups present in cysteine (or cysteine derivative) side-chains or terminal groups can be reacted with reagents possessing thiol-reactive functional groups using reaction schemes known in the art. Exemplary thiol-reactive functional groups include, without limitation, iodoacetamides, maleimides, and alkyl halides. Non-limiting examples of such conjugation schemes using a terminal cysteine linker are shown in FIG 4. Cysteine can be used as a linker, either as a single amino acid or as part of a larger linker comprising two or more amino acids, synthetic spacers (e.g., PEG) or a combination thereof. FIG 4 uses the peptide 346-001 backbone as a non-limiting example for illustrative purposes only. Exemplary, non-limiting embodiments shown in FIG 4 include: (A) derivatives of cholesterol, or pharmaceutically acceptable salts there-of; (B) maleimide derivatives of PEG, or pharmaceutically acceptable salts there-of, where m is 1-12, n is 1-12, and R is H, alkyl, or aryl; (C) maleimide derivatives of cholesterol, or pharmaceutically acceptable salts there-of, where m is 1-12, n is 1-12, and p is 1-12; (D) maleimide derivatives of tocopherol, or pharmaceutically acceptable salts there-of, where m is 1-12, n is 1-12, and p is 1-12; (E) maleimide derivatives of dihydrosphingosine, or pharmaceutically acceptable salts there-of, where m is 1-12, n is 1-12, and R is H, alkyl, or aryl; (F) maleimide derivatives of sphingosine, or pharmaceutically acceptable salts there-of, where m is 1-12, and n is 1-12; (G) maleimide derivatives of phospholipids, or pharmaceutically acceptable salts thereof, where R is H, alkyl, or aryl.

[0073]

[0054] The peptides of the disclosure can be prepared in a wide variety of ways. In various aspects, the peptide is desirably small while maintaining substantially all of the virucidal activity of the large peptide. The peptides can be prepared synthetically or by recombinant DNA technology using, e.g., methods known in the art. The peptides can be synthesized in solution or on a solid support in accordance with conventional techniques. Various automatic synthesizers are commercially available and can be used in accordance with known protocols.

[0074]

[0055] Compositions are also provided that comprise a peptide (or multiple peptides) of the disclosure. For example, in various aspects, the disclosure provides a composition comprising a peptide comprising the sequence SWLRX5IWEWIX11EVLX15EX17X18 (SEQ ID NO: 147), wherein X5is A, V, or L; Xu is S, d-serine, or phospho-serine; X15 is S, d-serine, or phospho-serine; X17 is F or phospho-tyrosine; and Xis is selected from R or phospho-tyrosine. In various aspects, X5 is A. Optionally, Xu is S and X15 is D-serine or Xu is D-serine and X15 is S. In various aspects, Xu is S and X15 is D-serine. X17 is optionally F and Xis is optionally R. The disclosure provides a composition comprising a peptide comprising the sequence SWLRX5IWEWIX11EVLX15EX17X18X19X20 (SEQ ID NO: 148) wherein X19-X20 are each optionally R or phospho-tyrosine (e.g., R). In an exemplary aspect, the peptide comprises the amino acid sequence SWLRAIWEWISEVLsEFRRR (SEQ ID NO: 149). In various aspects, the peptide comprises an amide group at the C-terminus. Optionally, one or more amino acids are D- amino acids. In various aspects, the peptide is formulated with an additional peptide, a liposome, an adjuvant and / or a pharmaceutically acceptable carrier. Liposomes can also be used to increase the half-life of the peptide composition. Liposomes useful in the context of the present disclosure include emulsions, foams, micelles, insoluble monolayers, liquid crystals, phospholipid dispersions, lamellar layers and the like. In these preparations the peptide to be delivered is incorporated as part of a liposome, alone or in conjunction with a molecule that binds to a suitable receptor, or with other therapeutic or immunogenic compositions. Thus, liposomes filled with a desired peptide of the disclosure can be directed to the site of cells infected with the virus, where the liposomes then deliver the selected therapeutic / immunogenic peptide compositions. Liposomes for use in the context of the disclosure may be formed from standard vesicle-forming lipids, which generally include neutral and negatively charged phospholipids and a sterol, such as cholesterol. The selection of lipids is generally guided by consideration of, e.g., liposome size and stability of the liposomes in vivo. A variety of methods are available for preparing liposome, many of which are described in the art.

[0075]

[0056] In another aspect, the peptide is chemically or physically attached to polymeric micro- or nanoparticles, a number of which are known in the art. Nonlimiting examples include biodegradable, biocompatible microspheres and nanospheres, where nonlimiting examples of resorbable synthetic polymers include poly (lactic acids), poly (glycolic acids), poly (lactic-co-glycolic acids), poly (caprolactones) (POLs), and mixtures thereof. In some embodiments, the peptide is dispersed inside the polymer particles using emulsion techniques well known in the art. In other embodiments, the N-terminus of the peptide is conjugated to the surface of the polymer particles via free carboxy groups. Alternatively, the peptide C-terminus is coupled to free carboxy groups on the surface of the polymer particles via a suitable linker group using synthetic chemistry approaches well-known in the art. The purposes of the polymer micro- or nanoparticles include, but are not limited to: monodispersity for efficient delivery to target anatomic regions, and mucoadhesive properties to affect retention times of the agent.

[0076] Complementary Agents

[0077]

[0057] In addition to the peptide disclosed above, one or more complementary agents are optionally administered to the subject. "Complementary” is intended to mean that the agents are supportive in achieving the desired pharmacological or biological effect (e.g., alleviating one or more symptoms of a viral (e.g., viral non- respiratory) disease, alleviating one or more side-effects of the disease, addressing a separate illness or disorder, or addressing an associated disruption in normal functioning of the subject).

[0078]

[0058] In various aspects, the complementary agent(s) possesses antimicrobial properties. Nonlimiting examples of complementary antimicrobial agents include: • Antiretroviral agents, including, but not limited to nucleoside reverse transcriptase inhibitors (NRTIs, e.g., tenofovir disoproxil fumarate, tenofovir alafenamide, emtricitabine, lamivudine), nucleoside reverse transcriptase translocation inhibitors (NRTTI, e.g., islatravir), non-NRTIs (e.g., rilpivirine, etravirine, doravirine), integrase inhibitors (INSTIs, e.g., raltegravir, dolutegravir, cabotegravir, elvitegravir), protease inhibitors (Pls, e.g., tipranavir, darunavir, atazanavir), and capsid inhibitors (Cis, e.g., lenacapavir),

[0079] • Antiviral agents, including, but not limited to valaciclovir, acyclovir, famciclovir, ganciclovir, and remdesivir,

[0080] • Broad-spectrum antimicrobial agents, including, but not limited to / .-lactic acid and D-lactic acid, as well as probiotic bacteria, such as lactobacilli,

[0081] • Antibacterial agents, including, but not limited to metronidazole, clindamycin, and tinidazole,

[0082] • Antifungal agents, including, but not limited to fluconazole, voriconazole, and amphotericin B,

[0083] • Antiparasitic agents, including, but not limited to ivermectin, and

[0084] • Transition metal salts and complexes (e.g., zinc complexes, silver complexes, copper complexes).

[0085]

[0059] In various aspects, the complementary microbicide component is a small molecule therapeutic. For example, the complementary agent may be a CD4-mimetic compound that binds HIV vaginally, as described by Madani et al. (f 1) incorporated herein in its entirety. In another aspect of the disclosure, the complementary microbicidal compound is a biologic therapeutic (e.g., a protein therapeutic or a nucleic acid-based therapeutic).

[0086]

[0060] In another aspect, the complementary antiviral agent comprises a protein. A non-limiting example of an antiviral protein is an antiviral protein belonging to the lectin family, such as griffithsin (GRFT), cyanovirin-N (CV- N), and scytovirin (SVN), described in (12), which is incorporated by reference herein in its entirety. In another nonlimiting example, the antiviral agent is a neutralizing antibody (bNAb), such as a broadly neutralizing antibody, a non-limiting example of which is VRC01 that possesses activity against various HIV-1 isolates {13). Other, next generation bNAbs that are more potent against HIV or neutralize other viruses such as HSV or HPV, or bacteria, such as multidrug-resistant Neisseria gonorrhoeae are also included herein as part of the disclosure.

[0087]

[0061] In some cases, the complementary pharmaceutically active substance is an antibacterial agent. In some cases, the antibacterial agent is a biologic with activity against multidrug-resistant Neisseria gonorrhoeae, such as those described in the art (e.g., 14-17, incorporated herein by reference).

[0088]

[0062] In some cases, the complementary agent is an agent that affects immune and fibrotic processes. Nonlimiting examples of agents that affect immune and fibrotic processes include, but are not limited to, inhibitors of Rho-associated coiled-coil kinase 2 (ROCK2), for example, KD025 (Kadmon) and didemnins (e.g., plitidepsin).

[0089]

[0063] In some cases, the complementary agent is a sirtuin (SIRT1-7) inhibitor. In some cases, the sirtuin inhibitor is EV-100, EV-200, EV-300, or EV-400 (Evrys Bio). In some cases, administration of a sirtuin inhibitor restores a human host's cellular metabolism and immunity.

[0064] In various aspects, the complementary agent is a contraceptive. As used herein, the term "contraceptive" refers to an active agent that prevents conception or pregnancy. In various aspects, the contraceptive is a hormonal contraceptive or a non-hormonal contraceptive. In various aspects, the complementary agent is a hormonal contraceptive. Non-limiting examples of hormonal contraceptives include estrogens or progestins, e.g., estradiol, etonogestrel, levonorgestrel, medroxyprogesterone acetate, segesterone acetate, norethindrone, and progesterone. In various aspects, the complementary agent is a non-hormonal contraceptive. In various aspects, the non-hormonal contraceptive comprises multivalent antibodies (e.g., IgGs) with high agglutination potencies for trapping vigorously motile sperm, or a small molecule, such as an inhibitor of soluble adenylyl cyclase (sAC:ADCY10), essential for male fertility.

[0090]

[0065] The complementary agents described herein can be administered alone (as part of a therapeutic regimen including administration of the peptide) or in combination with other complementary agents (and administration of the peptide). In some cases, the formulations described herein comprise more than one pharmaceutically active substance. In some cases, the formulations described herein comprise a combination of pharmaceutically active substances.

[0091]

[0066] In some cases, a combination of complementary agents is provided which includes chloroquine and azithromycin, hydroxychloroquine and azithromycin, lopinavir and ritonavir, KD025 and ribavirin, KD025 and remdesivir, EV-100 and ribavirin, or EV-100 and remdesivir. In certain embodiments, an antiviral peptide described herein is combined with one or more complementary agents disclosed above.

[0092]

[0067] In certain aspects of the disclosure, a complementary agent is chemically linked with the peptide to generate peptide-drug conjugates. This strategy is an effective conjugation strategy for targeted delivery and improved pharmacological outcomes as described, for example, in the review by Wang et al. {18), incorporated herein by reference in its entirety and in particularly in regard to its discussion of conjugates.

[0093] Multipurpose Prevention Technologies

[0094]

[0068] The disclosure contemplates use of the peptide described herein in the context of multipurpose prevention technologies, including delivery of the peptide using the delivery devices and methods described below. The oral, subdermal, intramuscular, rectal, and vaginal delivery of antiviral (e.g., antiretroviral (ARV)) drugs has shown clinical promise in protecting women from HIV infection. However, the effectiveness of this preexposure prophylaxis (PrEP) strategy has been severely limited by adherence to the dosing regimen. Multipurpose prevention technologies (MPTs) {19) are an integrated biomedical approach that provides dual (or more) protection (20-22), usually from HIV infection, along with other sexually transmitted infections (STIs), and unintended pregnancy. The MPT strategy exploits synergies in contraceptive demand for STI protection (23), significantly improving user compliance relative to single-purpose antiviral products (24).

[0095]

[0069] The inclusion of a contraceptive feature in MPTs can motivate product uptake and use for HIV PrEP (22, 23, 25-27), thereby improving effectiveness. Women can receive dual protection discreetly, even if their stated intention is to address just one health need, because of pressures from their sociocultural context (e.g., HIV stigma) or relationships. The inclusion of a contraceptive feature in MPT intravaginal rings (IVRs) can strongly motivate product uptake and use for HIV PrEP, thereby improving effectiveness. When asked about preference for single indication products versus MPTs, the overwhelming majority of reproductive age women in various settings, geographical locations, and demographic groups expressed a preference for an MPT. Providing access to MPTs therefore may indirectly improve both contraception and HIV / STI prevention coverage (28, 29).

[0096]

[0070] All MPTs disclosed in the art share a number of common features. At least one of the agents delivered by the MPT via IVR acts in the cervicovaginal tissues or systemic circulation. For example, the two most prominent IVR MPTs under development involve the delivery of levonorgestrel (LNG) as the contraceptive agent and either dapivirine (DPV) (30) or tenofovir (TFV) (31) as the antiviral agent against HIV. Both products are undergoing clinical evaluation in sub-Saharan Africa (32-34). In these MPT IVRs the contraceptive agent acts systemically and possibly in the cervicovaginal tissues, while the antiviral drugs (DPV and TFV) are believed to act in the cervicovaginal tissues. None of the drugs are active in the cervicovaginal fluids.

[0097]

[0071] A number of other vaginal MPT products are under development and aim to prevent unwanted pregnancy using hormonal contraceptives such as a progestin only (e.g., LNG) or a progestin-estrogen combination, such as etonogestrel and ethinyl estradiol. Hormonal contraception is known in the art to result in real and perceived side effects, including altered bleeding patterns, weight gain, mood swings and depression, headaches and nausea (35). Many women would prefer a non-hormonal method that does not require use immediately before or after sex. In the context of an MPT that protects against sexual HIV acquisition and unintended pregnancy, there is growing concern, largely based on injectable depot medroxyprogesterone acetate (DMPA) (36, 37), that a LNG-only regimen as part of an MPT may lead to increased risk of HIV-1 acquisition (38-42) The use of a vaginally active non-hormonal contraceptive is desirable as it avoids these potential risks associated with exogenous hormones. The disclosure contemplates use of the peptide described herein in the context of vaginal MPT products.

[0098]

[0072] When drugs are delivered from a vaginal product, they partition into the cervicovaginal fluids and, from there, into the tissues and finally systemic circulation. There are considerable drug concentration gradients across these anatomic compartments, with the cervicovaginal fluid having drug concentrations order(s) of magnitude higher than the other compartments, depending on the agent's physicochemical properties. It therefore is theoretically beneficial to use an agent against HIV or other STIs that is active in the cervicovaginal fluids.

[0099]

[0073] The disclosure contemplates use of a vaginal MPT product, such as a product disclosed herein, to deliver the peptide described herein, optionally with one or more other agents, all of which are active in the cervicovaginal fluids. The agents are optionally microbicidal against HIV and other microorganisms leading to STIs and provide non-hormonal contraception.

[0074] An MPT drug formulation can include essentially any therapeutic, prophylactic, or diagnostic agent that would be useful for delivery to an anatomic compartment. In the disclosure, the MPT drug product must contain at least one or more antiviral peptides disclosed herein and one or more contraceptive agents. One or more additional complementary agents, as described above, can be delivered in combination with the antiviral peptide, as determined by the application.

[0100]

[0075] In one preferred aspect, the non-hormonal contraceptive is active against sperm. For example, ferrous gluconate causes spermiostasis (43, 44). In another, non-limiting example, the non-hormonal contraceptive is a small molecule, such as an inhibitor of soluble adenylyl cyclase (sAC:ADCY10), essential for male fertility (45). Other non-limiting examples of male, non-hormonal contraceptives known in the art target EPPIN, a surface protein on human spermatozoa that has an essential function in reproduction (46, 47), and cyclin-dependent kinase 2 (CDK2) (48), included herein in their entirety. In another embodiment, the non-hormonal contraceptive consists of multivalent IgGs with high agglutination potencies for trapping vigorously motile sperm (49, 50).

[0101] Administration Methods

[0102]

[0076] The active agent(s) described above (i.e., the peptide and complementary agents, when present) are administered to the mammalian subject using any suitable route of delivery. Pharmaceutical compositions of the present disclosure can be administered orally, by inhalation, via a pulmonary route of administration, I ntranasally (including intranasal instillation), topically, transdermally, intraplurally, intraperitoneally, by application to a mucous membrane, parenterally, topically, intravenously, subcutaneously, intraperitoneally, or by intramuscular administration.

[0103]

[0077] Delivery of the peptide(s) and complementary agents, if applicable, is optionally achieved via a pulsatile means as known in the art; e.g., oral tablet, intravenous injection, subcutaneous injection, intramuscular injection, intravitreal injection, topical cream, vaginal tablet, vaginal film, vaginal insert, vaginal gel, vaginal cream, rectal suppository, rectal enema, or rectal gel.

[0104]

[0078] Alternatively, delivery of the peptide(s) and complementary agents may be achieved using a sustained release or long-acting system or drug delivery device as known in the art; e.g., injectable nanoparticles (subcutaneous, intramuscular, intravitreal), injectable microparticles (subcutaneous, intramuscular, intravitreal), microneedles and microneedle arrays, subdermal implants, intramuscular implants, intravaginal rings. Nonlimiting examples of devices suitable for delivery of the antiviral peptide(s) and complementary agents include those disclosed in International Patent Publication Nos. WO 2022115599, WO 2021 / 108722, and WO 2021 / 071974, the disclosure of each of which is incorporated herein by reference. The disclosure contemplates an implantable drug delivery device comprising the peptide described herein, including intravaginal rings or buccal implants.

[0105]

[0079] For example, a suitable drug delivery device can comprise a scaffold comprising one or more biocompatible materials, one or more chambers containing a plurality of cells, one or more membranes, and one or more nutrient supplementation systems. In the context of the disclosure, the cells may produce the peptide described herein. In some cases, the drug delivery device is adapted for intravaginal use. In some cases, the one or more biocompatible materials comprise one or more thermoplastic polymers, one or more elastomers, one or more biocompatible metals, or combinations thereof. Non-limiting examples of suitable biocompatible materials include silicone, polyurethane, poly (ethylene-co-vinyl acetate) (EVA), or a combination thereof. Nonlimiting examples of suitable cells include bacterial cells, fungal cells, mammalian cells, or a combination thereof. The cells may produce the peptide described herein, or may produce a different active agent suitable for use with the peptide described herein. Non-limiting examples of suitable materials for the membrane or membranes include polyester, polypropylene, polycarbonate, polyethylene terephthalate (PET), anisotropic materials, polysulfone (PSF), microfiber or nanofiber mats, polyimide, tetrafl uoroethylene / polytetrafluoroethylene (PTFE), expanded polytetrafluoroethylene (ePTFE), poly(ethylene-co-vinyl acetate) (EVA), polyacrylonitrile, polyethersulfone, acrylic resin, cellulose acetate, cellulose nitrate, polyamide, hydroxylpropyl methyl cellulose (HPMC), or a combination thereof. Non-limiting examples of suitable nutrient supplementation systems comprise nutrients, growth factors, hormones, vitamins, O2-generating agents, pH buffering agents, cell culture media, antibiotics, or a combination thereof.

[0106]

[0080] For example, a suitable drug delivery device can comprise (a) one or more kernels comprising one or more active pharmaceutical ingredients (APIs), such as the peptide described herein; and (b) one or more skins comprising a continuous membrane; wherein the one or more kernels and / or the skin comprises defined pores, and wherein the pores are not produced mechanically. In some embodiments, the reservoir kernel comprises a paste comprising one or more APIs. In some embodiments, the kernel comprises a fiber-based carrier and / or a porous sponge. In some embodiments, the device further comprises a shape adapted to be disposed within the body of a patient. For example, the device can be capsule-shaped, or can be in the shape of a torus. In some embodiments, the device comprises one or more cylindrical core elements disposed within a first skin, wherein the core elements comprise a kernel and optionally a second skin. In some embodiments, the skin comprises a rate-limiting skin. Non-limiting examples of suitable materials for the skin include poly (dimethyl siloxane), silicone, one or more polymers, and / or metal, where the polymer can be, without limitation, poly(ether), poly (acrylate), poly(methacrylate), poly(vinyl pyrolidone), poly(vinyl acetate), poly(urethane), cellulose, cellulose acetate, poly (siloxane), poly (ethylene), poly (tetrafluoroethylene) (such as expanded poly (tetrafluoroethylene) or ePTFE) and other fluorinated polymers, poly(siloxanes), copolymers thereof, or combinations thereof, poly(amides), poly(esters), poly (ester amides), poly(anhydrides), poly(orthoesters), polyphosphazenes, pseudo poly(amino acids), poly(glycerol-sebacate), poly (lactic acids), poly (glycolic acids), poly (lactic-co-glycolic acids), poly(caprolactones) (PCLs), PCL derivatives, amino alcohol-based poly (ester amides) (PEA), poly (octane-diol citrate) (POC), copolymers thereof, or mixtures thereof.

[0107]

[0081] For example, a suitable drug delivery device can comprise a buccal implant device configured to provide sustained drug delivery, the buccal implant device comprising: a body having a non-cylindrical geometry, the body adapted to be disposed within the oral mucosa of a patient; and one or more active pharmaceutical ingredients (e.g., peptide described herein) disposed within the body. In some embodiments, the buccal implant device comprises: a body adapted to be disposed within the oral mucosa of a patient; means for guiding the body into or out of the oral cavity of a patient; and one or more active pharmaceutical ingredients disposed within the body. In some embodiments, the device comprises a matrix, reservoir, or hybrid design comprising one or more thermoplastic polymers, elastomer materials, or metals suitable for pharmaceutical use and a therapeutically effective amount of one or more active pharmaceutical ingredients. For example, the device can be a buccal implant device configured to provide sustained drug delivery, the buccal implant device comprising: a body adapted to be disposed within the oral mucosa of a patient; means for guiding the body into or out of the oral cavity of a patient; and one or more active pharmaceutical ingredients disposed within the body. Nonlimiting examples of suitable shapes for the device include a saw shape, a saber shape, and a ribbed shape. In some embodiments, the body is tapered, e.g., having a single taper, two tapers, or three tapers. In some embodiments, the body has one or more curved edges configured to direct the implant into a specific orientation within the oral cavity. In some embodiments, the one or more curved edges form a point at an end of the body. In some embodiments, the device further comprises a hole formed in the body and / or a loop coupled to the hole to guide the body into or out of the oral cavity. In some embodiments, the device comprises expanded polytetrafluoroethylene (ePTFE) having microscopic pores. In some embodiments, the device comprises a ratelimiting skin comprising, without limitation, expanded polytetrafluoroethylene (ePTFE).

[0108]

[0082] In some embodiments, the API is formulated as an insert, a suppository, a film, a gel, a douche, a cream, an ointment, or a rinse. In some embodiments, an API is formulated as a long-acting, fast-dissolving, slow-releasing, and / or mucoadhesive composition. In some embodiments, a long-acting composition is one that lasts 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 1 day, 2 days, 7 days, 14 days, 21, days, 28 days, 1 month, 2 months, 6 months, 1 year, 2 years, or 5 years. In some embodiments, a fast-dissolving composition is one that dissolves in less than 5 seconds, 30 seconds, 1 minute, 2 minutes, 5 minutes, 10 minutes or 15 minutes. In some embodiments a slow-releasing composition is one that releases over 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 1 day, 2 days, 7 days, 14 days, 21, days, 28 days, 1 month, 2 months, 6 months, 1 year, 2 years, or 5 years. In some embodiments, a mucoadhesive composition comprises a composition that interacts with the mucus layer covering the mucosal epithelial surface, and mucin molecules and increases the residence time of the dosage form at the site of absorption.

[0109] Respiratory Drug Delivery Systems

[0110]

[0083] The active agent(s) described herein (i.e., the peptide and complementary agents, when present) may be administered to the respiratory system using any suitable delivery system. Suitable delivery systems include, but are not limited to, metered-dose aerosol systems, dry-powdered inhalation systems, and nasal inhalation systems. An illustrative nonlimiting summary of the various drug delivery embodiments is given below. Nasal Drug Delivery

[0111]

[0084] Nasal delivery of liquid and solid formulations of the agent(s) disclosed above (i.e. , the antiviral peptide and / or complementary agent) can be achieved by the following non-limiting devices:

[0112] • Pipettes for drop or vapor delivery, which may be breath powered or hand-actuated;

[0113] • Vapor inhaler, such as commercial menthol inhalers for rhinitis;

[0114] • Mechanical liquid spray pumps and squeeze bottles, which may be breath powered or hand-actuated;

[0115] • Gas-driven spray systems and atomizers, including pressurized metered-dose inhalers (pMDIs);

[0116] • Electrically powered nebulizers and atomizers;

[0117] • Hand-actuated, single dose mechanical powder sprayers;

[0118] • Breath-actuated solid inhalers; and

[0119] • Single-dose mechanical solid insufflators.

[0120] Pulmonary Drug Delivery

[0121]

[0085] The mass median aerodynamic diameter and geometric standard deviation (GSD) are parameters that determine the site of particle deposition in the respiratory tract. Large particles or droplets deposit by impaction in the upper respiratory tree of the lung (oropharyngeal and tracheo-bronchial region), where air velocity is high and the air flow is turbulent. Particles in the size range of 0.5-5 m deposit by sedimentation in the terminal bronchioles and alveolar regions. The larger the GSD, the more sites the aerosol will be deposited in the respiratory tract. In general, aerosols with GSD < 2 are desirable and, ideally, aerosol particle size distributions should be as close as possible to monodispersity to increase deposition at the desired site of action and increase the efficacy of the treatment.

[0122]

[0086] Pulmonary drug delivery strategies have been described and are well-known in the art. Dry powder inhalers are popular devices used to deliver drugs, especially proteins, to the lungs. Some of the commercially available dry powder inhalers include Spinhaler (Fisons Pharmaceuticals, Rochester, NY) and Rotahaler (GSK, Research Triangle Park, NC). Several types of nebulizers are available, namely jet nebulizers, ultrasonic nebulizers, and vibrating mesh nebulizers. Jet nebulizers are driven by compressed air. Ultrasonic nebulizers use a piezoelectric transducer to create droplets from an open liquid reservoir. Vibrating mesh nebulizers use perforated membranes actuated by an annular piezo-element to vibrate in resonant bending mode. The holes in the membrane have a large cross-section size on the liquid supply side and a narrow cross-section size on the side from where the droplets emerge. Depending on the therapeutic application, the hole sizes and number of holes can be adjusted. Selection of a suitable device depends on parameters, such as nature of the drug and its formulation, the site of action, and pathophysiology of the lung. Aqueous suspensions and solutions are nebulized effectively. Aerosols based on mechanically generated vibration mesh technologies also have been used successfully to deliver proteins to lungs.

[0123] Excipients and Manufacturing Considerations

[0087] The disclosure contemplates a composition comprising one or more of the peptides described herein and one or more pharmaceutically acceptable excipients. Pharmaceutically acceptable excipients are known in the art and may include, e.g., viscosity modifiers, bulking agents, surface active agents, dispersants, disintegrants, osmotic agents, diluents, binders, anti-adherents, lubricants, glidants, pH modifiers, antioxidants and preservants, and other non-active ingredients of the formulation intended to facilitate handling and / or affect the release kinetics of the drug.

[0124]

[0088] In some aspects, the binders and / or disintegrants may include, but are in no way limited to, starches, gelatins, carboxymethylcellulose, croscarmellose sodium, methyl cellulose, ethyl cellulose, hydroxymethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropylmethyl cellulose, hydroxypropylethyl cellulose, hydroxypropylmethyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, polyethylene glycol, sodium starch glycolate, lactose, sucrose, glucose, glycogen, propylene glycol, glycerol, sorbitol, polysorbates, and / or colloidal silicon dioxide. In certain embodiments, the anti-adherents or lubricants may include, but are in no way limited to, magnesium stearate, stearic acid, sodium stearyl fumarate, and / or sodium behenate. In some embodiments, the glidants may include, but are in no way limited to, fumed silica, talc, and / or magnesium carbonate. In some embodiments, the pH modifiers may include, but are in no way limited to, citric acid, lactic acid, and / or gluconic acid. In some embodiments, the antioxidants and preservants may include, but are in no way limited to ascorbic acid, butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), cysteine, methionine, vitamin A, vitamin E, sodium benzoate, and / or parabens.

[0125]

[0089] In some aspects, excipients can stabilize biomolecules with respect to degradation or loss of biological activity using approaches known to those skilled in the art {51). Certain excipients stabilize biomolecules by creating a "water-like” environment in the dry state through hydrogen bonding interactions, e.g., sugars (52) and amino acids (53). Other excipients create a glassy matrix that provides hydrogen bonding and immobilized the biomolecules to prevent aggregation that leads to loss of biologic activity (e.g., trehalose, inulin). Still other excipients can stabilize the pH in the implant formulation (e.g., buffer salts). Finally, surfactants can reduce the concentration of the biomolecules at the air-water interface during drying processes of formulation, decreasing shear stress and insoluble aggregate formation, and allowing the previously described stabilization mechanisms to occur throughout the drying process.

[0126]

[0090] Solid formulations for dry powder inhalers (DPIs) can be prepared by a variety of methods, many of which are well known in the art. These include, but are not limited to spray drying, spray-freeze drying, supercritical fluid technology, solvent precipitation method, double emulsion / solvent evaporation technique, particle replication in nonwetting templates, and lyophilization. The resulting polydisperse mixtures can be refined further by specialized milling techniques. Jet-milling of drugs and excipients under nitrogen gas with a nano-jet milling machine is one non-limiting exemplary method known in the art for creating nanoparticles meant for pulmonary drug delivery. Methods

[0127]

[0091] The disclosure provides materials and methods for delivery of an antiviral peptide (and, optionally, one or more complementary agents) to a subject for the purposes of treating, preventing, reducing the likelihood of having, reducing the severity of and / or slowing the progression of a medical condition in a subject, in a preferred aspect, the medical condition is a viral infection, or exposure to a virus.

[0128]

[0092] In some cases, a subject in need of treatment for a disease or disorder disclosed herein, such as an infectious disease, is symptomatic for the disease or disorder. In some cases, a subject in need of treatment for a disease or disorder disclosed herein, such as an infectious disease, is asymptomatic for the disease or disorder. A subject in need of treatment for a disease or disorder disclosed herein can be identified by a skilled practitioner, such as without limitation, a medical doctor or a nurse. In some cases, the antiviral peptide is used to prevent a viral disease in the subject.

[0129]

[0093] In various aspects, the peptide is delivered to a subject using an implant system which delivers the peptide (and one or more other APIs) to a body compartment, thereby treating, preventing, reducing the likelihood of having, reducing the severity of and / or slowing the progression of a viral disease in a subject. In some cases, the anatomic compartment is the vagina. In other cases, the target body compartment is systemic circulation. A long-acting drug delivery system reduces problems associated with patient adherence to regimens that involve frequent dosing from, for example, the subcutaneous, intramuscular, or buccal compartments,

[0130]

[0094] in various aspects, the disclosure provides a method of treating or preventing a viral non-respiratory disease. By "viral non-respiratory disease” is meant a disease whose primary etiology is not a viral infection of the respiratory system of a patient, e.g., an infection of the upper respiratory system and / or the lower respiratory system. In particular, the acquisition of the virus is not predominantly through the respiratory system, e.g., through the nasal passages, larynx, trachea, lungs, and / or bronchi.

[0131]

[0095] Illustrative, non-restrictive examples of non-respiratory viral diseases contemplated by the disclosure are provided below in summary form. Based on these examples, one skilled in the art can adapt the disclosed technology to other applications. One skilled in the art would recognize whether such applications involve topical drug delivery (e.g., certain vaginal implant devices such as IVRs) or systemic drug delivery (e.g., subdermal or intramuscular implant devices).

[0132] • HIV prevention, wherein the method optionally comprises administration of a complementary agent which is a suitable antiretroviral agent (e.g., a biologic), or a vaccine and / or adjuvant;

[0133] • HIV treatment, wherein the method optionally comprising administering using a complementary agent which is a suitable antiretroviral agents (e.g., a biologic);

[0134] • Sexually transmitted infections (STIs), including but not limited to prevention or treatment, both active and chronic active, wherein the method optionally comprises administering a complementary agent which is a suitable antimicrobial agent. Illustrative, but not limiting examples of STIs include: gonorrhea, chlamydia, lymphogranuloma venereum, syphilis, including multidrug-resistant (MDR) organisms, hepatitis C virus, and herpes simplex virus;

[0135] • Hepatitis B virus (HBV) prevention or treatment, both active and chronic active;

[0136] • Herpes simplex virus (HSV) and varicella-zoster virus (shingles) Zoster / Shingles, prevention or treatment, both active and chronic active;

[0137] • Cytomegalovirus (CMV) and congenital CMV infection, prevention or treatment, both active and chronic active.

[0138]

[0096] In some embodiments, the prevention of a viral infection is complemented by the prevention of unintended pregnancy (i.e., contraception), optionally using an MPT approach as described above. In an MPT approach, the treatment or prevention of a viral infection can be complemented by the treatment or prevention of a non-viral infection, for example, bacterial vaginosis (BV), as well as other microbial dysbiotic vaginal states, including both active and chronic active.

[0139]

[0097] Thus, in various aspects, the disclosure provides a method of treating or preventing a viral respiratory disease. In particular, the disclosure provides a method of treating or preventing a disease caused by a virus. Viral respiratory diseases are generally illnesses caused by viruses having similar traits and which affect the upper respiratory tract. Viral respiratory diseases include, e.g., infectious diseases (e.g., a coronavirus (CoV) infection (e.g., SARS-CoV-2 infection), an influenza infection, a parainfluenza infection, a respiratory syncytial virus (RSV) infection, a respiratory adenovirus infection, and the like), and analogous conditions in non-human mammals. The viral respiratory disease may be, for example, COVID-19, severe acute respiratory syndrome (SARS), or Middle East respiratory syndrome (MERS). Symptoms of the viral respiratory disease may include, e.g., fever, headache, body aches, dry cough, hypoxia (oxygen deficiency), and / or pneumonia. In some cases, the viral respiratory disease is COVID-19 (i.e., SARS-CoV-2 infection).

[0140]

[0098] Influenza spreads around the world in seasonal epidemics, resulting in the deaths of hundreds of thousands annually-mi llions in pandemic years. For example, three influenza pandemics occurred in the 20th century and killed tens of millions of people, with each of these pandemics being caused by the appearance of a new strain of the virus in humans. Often, these new strains result from the spread of an existing influenza virus to humans from other animal species. Influenza viruses are RNA viruses of the family Orthomyxoviridae, which comprises five genera: influenza virus A, influenza virus B, influenza virus C, isavirus, and Thogoto virus. The influenza A virus can be subdivided into different serotypes based on the antibody response to these viruses. The serotypes that have been confirmed in humans, ordered by the number of known human pandemic deaths, are: H1 N1 (the strain that caused Spanish influenza in 1918), H2N2 (caused Asian Influenza in 1957), H3N2 (caused Hong Kong Flu in 1968), H5N1 (a pandemic threat in the 2007-08 influenza season), H7N7 (has unusual zoonotic potential), H1 N2 (endemic in humans and pigs), H9N2, H7N2, H7N3 and H10N7. Influenza B causes seasonal flu and influenza C causes local epidemics, and both influenza B and C are less common than influenza A.

[0099] Coronaviruses are a family of common viruses that cause a range of illnesses in humans from the common cold to severe acute respiratory syndrome (SARS). Coronaviruses can also cause a number of diseases in animals. Coronaviruses are enveloped, positive-stranded RNA viruses whose name derives from their characteristic crown-like appearance in electron micrographs. Coronaviruses are classified as a family within the Nidovirales order, viruses that replicate using a nested set of mRNAs. The coronavirus subfamily is further classified into four genera: alpha, beta, gamma, and delta coronaviruses. The human coronaviruses (HCoVs) are in two of these genera: alpha coronaviruses (including HCoV-229E and HCoV-NL63) and beta coronaviruses (including HCoV-HKU1, HCoV-OC43, Middle East respiratory syndrome coronavirus (MERS- CoV), the severe acute respiratory syndrome coronavirus (SARS-CoV and SARS-CoV-2).

[0141]

[0100] As used herein "detecting” refers to the identification of a viral infection. Various methods for detecting a viral infection are known in the art. In some embodiments, a viral infection is detected by PCR analysis. In some embodiments, a viral infection is detected by antigen analysis. In some embodiments, the detecting is performed by the subject. In some embodiments, the detecting is not performed by the subject.

[0142]

[0101] Another embodiment exploits the synergy between fully integrated point-of-care testing (POCT) medical diagnostics with a therapy based on self-administration of the antimicrobial peptide ( / .e., companion diagnostic). In a non-limiting embodiment, women infected with HPV vaginally self-collect, test for HPV, and, based on the results treat with the antimicrobial peptide disclosed herein using a suitable vaginal product (e.g., insert, gel, suppository, film, or IVR). In one embodiment, the self-collection device is sent to a laboratory for analysis (e.g., Teal Wand™ for cervical cancer screening). In another embodiment, the self-collected sample is tested (e.g., antigen, PCR) by the patient followed to self-treatment. In another, non-limiting example, the vaginally collected, tested, and treated virus consists of HSV, dengue virus, Zika virus, etc. In another embodiment, the self-testing and product administration is paired with telemedicine support. Companion testing can be used to tailor the antimicrobial peptide dosing regimen, including dosing frequency and dose size, under appropriate medical supervision.

[0143]

[0102] The disclosure contemplates veterinary application of the materials and methods described herein, involving all mammals, including, but not limited to dogs, cats, horses, pigs, sheep, goats, and cows.

[0144]

[0103] In one embodiment of the disclosure, the system serves multiple purposes, where more than one medical condition is targeted simultaneously. An example of such a multipurpose drug delivery system involves the treatment of a SARS-CoV-2 infection along with concomitant dampening of the host immune response.

[0145] EXAMPLES

[0146] EXAMPLE 1 - Formulation 1 (Intravaginal Administration - On Demand)

[0147]

[0104] For on-demand intravaginal administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147 or SEQ ID NO: 149) is formulated as a vaginal film product. The films are produced by the solvent casting method, known in the art, and consist of poly (vinyl alcohol) (PVA, 67 kDa). To a solution of PVA (25% w / w) in deionized water is added slowly with magnetic stirring the antiviral peptide 0.01-100 mM, preferably 0.5-10 mM and preservatives known in the art to stabilize peptides (e.g., histidine and polysorbate 20). The pH of the solution is adjusted to an optimal pH for peptide stability, typically in the 6.0-7.5 range. The solution is stirred magnetically to ensure dissolution of all components and homogenous distribution, while removing entrapped air bubbles. The solution then is cast onto a plastic substrate (e.g., polyester) on a glass plate using a die press. The film is allowed to dry at room temperature and then is cut into sections of predetermined dimensions using a scalpel.

[0148]

[0105] In another example, a non-hormonal contraceptive from the soluble adenylyl cyclase (sAC) inhibitor class is included in the film formulation at 0.1-1,000 piM, preferably 10-100 piM, in combination with the antiviral peptide to form a multipurpose prevention technology.

[0149] EXAMPLE 2 - Formulation 2 (Intravaginal Administration - Long-acting)

[0150]

[0106] For long-acting intravaginal administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147 or SEQ ID NO: 149) is formulated as an intravaginal ring product, using designs known in the art to be suitable for the delivery of such agents. The device is engineered to deliver the antiviral peptide at daily release rates in the 0.01-10 mg d1range, preferably in the 0.2-5 mg d1range, such as 0.5-4 mg d1(e.g., 1 mg d1, 2 mg d1, 3 mg d1, or 4 mg d1). Preferably, the antiviral peptide is delivered in combination with a non-hormonal contraceptive from the sAC inhibitor class controlled independently to deliver the contraceptive at daily release rates in the 0.01-10 mg d1range, preferably in the 0.1-4 mg d1(e.g., 1 mg d1, 2 mg d1, 3 mg d1, or 4 mg d1). The combination forms an example of a form a multipurpose prevention technology.

[0151] EXAMPLE 3 - Formulation 3 (Intrarectal Administration - On Demand)

[0152]

[0107] For on-demand intrarectal administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147 or SEQ ID NO: 149) is formulated as rectal enema product. The peptide is dissolved in normal saline, or 50% v / v normal saline in deionized water. The concentration of the peptide is 0.01-100 mM, preferably 0.1-5 mM. Sodium hydroxide (5 M) or hydrochloric acid (5 M) is added to bring the solution near neutral pH. Alternately, the peptide powder is added to a sodium bicarbonate solution (1 mg mL1) and sodium chloride powder is added at a final concentration of 0.9% w / v to produce a saline-based enema. The final osmolarity of the formulation is 50-500 mOsm kg1, preferably 100-300 mOsm kg1.

[0153]

[0108] In another example, the antiretroviral drug tenofovir (TFV), or a suitable prodrug of tenofovir, also is dissolved in the rectal enema product along with the peptide as described above to create a combination product. The concentration of TFV is 0.5-50 mg mL1, preferably 0.5-5 mg mL1. Prodrugs of TFV are formulated at the same molar equivalent concentration.

[0109] In another example, the above ingredients are premixed and stored in solid form to produce the enema formulation on demand by the addition of water. This can hold the advantage of a longer shelf-life in smaller product package volume and mass.

[0154] EXAMPLE 4 - Formulation 1 (Pulmonary Administration)

[0155]

[0110] For aerosol administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147 or SEQ ID NO: 149) may be used as a liquid solution along with a surfactant and propellant. Typical percentages of Peptide 346-001 are 0.01 %-20% w / w, preferably 1 %-10% w / w. The surfactant is nontoxic, and preferably soluble in the propellant. Representative of such agents are the esters or partial esters of fatty acids containing from 6 to 22 carbon atoms, such as caproic, octanoic, lauric, palmitic, stearic, linoleic, linolenic, olesteric and oleic acids with an aliphatic polyhydric alcohol or its cyclic anhydride. Mixed esters, such as mixed or natural glycerides may be employed. The surfactant may constitute 0.1 %-20% w / w of the composition, preferably 0.25-5% w / w. The balance of the composition is ordinarily propellant. A carrier can also be included as desired, e.g., lecithin for intranasal delivery.

[0156] EXAMPLE 5 - Formulation 2 (Nasal Administration)

[0157]

[0111] Aqueous solutions of a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147 or SEQ ID NO: 149) and bulking agents (mannitol or lactose) are prepared in Dulbecco's phosphate-buffered saline (pH 8.0, adjusted with aqueous ammonia, 28% w / w). The resulting solutions are filtered through a 0.2 m PES filter (Celltreat Scientific Products, Shirley, MA) and aliquots (2 mL) of the solution are filled into sterile 5 mL borosilicate glass vials. Rubber stoppers are placed on top of the vials prior lyophilization. At the completion of the cycle, nitrogen is introduced into the chamber. Vials are sealed with rubber stoppers and aluminum crimp-top closure.

[0158] EXAMPLE 6

[0159]

[0112] In Vitro Substitution Analysis'. Twenty (20) peptides with sequence (H-SWLRX5IWEWISEVLSEFRRR- NH2, acetate salts, where X5 is one of the 20 natural amino acids; OCIS-346-232, OCIS-346-235 to -253) were evaluated for inhibition of HIV-1 IIIB in TZM-bl-FcRI cells. The peptides were evaluated at 10 concentrations with 8 replicates each for efficacy for 48 hours. Virus was diluted in assay media at 1 : 128 (based on a previous titration to an MOI of approximately 0.001-0.01) and compound (diluted in PBS) in the appropriate volume based on the titration data were incubated for 30 minutes at 37°C / 5% CO2. Following the incubation, the virus / compound mixture was further diluted 1 :8.5 and 180 L was added to pre-plated TZM-bl-FcRI cells. The cultures were incubated for 6 hours at the above conditions and then washed to remove residual virus and compound. The cultures were incubated for an additional 48 hours at 37°C / 5% CO2 before measuring D- galactosidase in the wells using a chemilluminescent endpoint. The in vitro half-maximal effective concentrations (EC50) against HIV-1 were calculated from dose-response plots in Prism (version 10.2.2, GraphPad Software LLC, Boston, MA) using variable slope model (4 parameter) that does not assume a Hill slope of unity.

[0113] In Vitro Peptide Stability in Cervicovaginal Fluids: Test peptide (10 mg) was dissolved in DMSO (100 piL) and the resulting solution added to acetate buffer (100 mM, pH 3.8, 9.9 mL). The mixture was agitated vigorously on a horizontal shaker for 15 min followed by centrifugation at 1,000 xg for 3 min. An aliquot (50 piL) of the resulting supernatant was mixed with human cervicovaginal fluid pooled from multiple donors, diluted (1 :1, v / v) with acetate buffer, pH 3.8, (50 piL, final volume 100 piL); three replicates per group, one sample per replicate per timepoint. The samples were maintained at 37°C and pulled for analysis at predetermined timepoints. To the sample (100 piL) was added acetonitrile (300 piL) and the cloudy mixture centrifuged at 13,000 RPM for 15 min. An aliquot (200 piL) of the supernatant was dried under vacuum in a SpeedVac. The dried sample was reconstituted in formic acid (0.1% v / v, 200 piL) in acetonitrile-water (30-70 v / v) and analyzed by LC- QTOF-MS.

[0160]

[0114] In Vivo Peptide Stability in Blood Following IV Dosing in Beagle Dogs: Non-nai've, female beagle dogs (8-12 months, 7-12 kg, N = 4 per group) were administered OCIS-346-232 via slow, bolus IV injection. The dose level was 15 mg kg d-1 (7.5 mg mL-1) in the first group and 44 mg kg d-1 (22 mg mL-1) in the second group. Blood samples (ca. 1 mL) were collected at predetermined timepoints, converted to plasma (K2EDTA), and stored at -75 ± 15°C. Concentrations of OCIS-346-232 in the plasma samples were analyzed by LC-MS / MS.

[0161]

[0115] Discussion: The present example illustrates multiple, unexpected findings with respect to the peptide of the disclosure. Data from an orthogonal amino acid scanning experiment focused on position 5 of the peptide sequence (the in vitro substitution analysis) are provided herein. Scan results are summarized in FIG 5. The in vitro half-maximal effective concentration (EC50) against HIV-1 varied widely depending on the nature of the amino acid residue at position 5 ("5-amino acid residue”). The lipophilicity of the 5-amino acid residue appeared to have an important effect on potency. Residues with charged side chains (e.g., Glu, Pro, Arg) or low lipophilicity (e.g., Gly, Gin) led to high EC50 values, i.e. , low potency. The lowest EC50 values, highest potencies, were obtained with Ala, Vai, and Leu at amino acid position 5. However, there appeared to be an inflection point, as more lipophilic residues (e.g., lie) and aromatic, lipophilic residues (e.g., Phe, Trp) did not have a beneficial impact on potency.

[0162]

[0116] Stability of the peptides in biological fluids is an important attribute. FIGs 6A-6C show illustrative peptide stability data in human cervicovaginal fluids (CVF, pooled from multiple donors). All peptides shared the same sequence, but natural, L-amino acids were replaced by their D-isomer counterparts at key locations in the sequence. The all-D-amino acid peptide provided a benchmark for maximum stability to enzymatic degradation. As expected, the all-D-amino acid peptide (peptide 346-203; H-swlraiwewisevlsefrrr-NH2, TFA salt) was more stable in CVF than the corresponding all-L-amino acid peptide (peptide 346-192; H- SWLRAIWEWISEVLSEFRRR-NH2, acetate salt) (FIG 6A). Surprisingly, only replacing the 15-Ser L-isomer with its D-isomer (peptide 346-232; H-SWLRAIWEWISEVLSEFRRR-NH2, acetate salt) led to dramatically enhanced stability compared to the all L-amino acid peptide (FIGs 6A and B). Replacing both the 11 -Ser and 15-Ser L- isomers with their D-isomers (peptide 346-212; H-SWLRAIWEWIsEVLsEFRRR-NH2, TFA salt) increased the peptide stability in CVF and made it equivalent to the all-D-amino acid peptide (peptide 346-203; H- swiraiwewisevlsefrrr-NF , TFA salt) (FIG 60). There was not much difference in stability between peptide 346- 232 (15-D-ser) and peptide 346-212 (11- and 15-D-ser), another unexpected result.

[0163]

[0117] The data illustrated in FIG 7 show that peptide 346-232, containing only one unnatural amino acid (15- D-ser) persisted in vivo, in blood (analyzed as plasma) over 24, and beyond following IV bolus dosing in female beagle dogs. This remarkable stability can be attributed to the substitution at the 15-position limiting enzymatic degradation, an unexpected result as linear, short peptides are unstable in systemic circulation. Optionally, D- serine at positions 11- and / or 15- is replaced with the related unnatural amino acid phospho-serine to, e.g., increase stability and solubility in biological fluids compared to the parent sequence containing only natural amino acids. Additionally, the C-terminus of the peptides disclosed herein can be modified without compromising their antiviral properties. Substitutions at the 17-, 18-, 19-, and 20- positions within the sequence (with reference to SEQ ID NO: 147) may increase peptide solubility in biological fluids. It will be appreciated that systemic stability and long half-lives of the antiviral peptides means the antiviral peptides disclosed herein can be used to treat systemic infections {e.g., HBV, HCV, West Nile virus), in addition to local infections.

[0164] TABLE 1

[0165] Peptide SEQ ID NO: Peptide Sequence and Modifications

[0166] 346-001 3 SWLRDIWDWICEVLSDFK

[0167] 346-002 4 swlrdiwdwicevlsdfk

[0168] 346-003 5 GSWLRDIWDWICEVLSDFK

[0169] 346-004 6 SWLRDIWDWICEVLSDFKTW

[0170] 346-005 7 kfdslveciwdwidrlws

[0171] 346-006 8 KWLCRIWSWISDVLDDFE

[0172] 346-007 9 SIWRDWVDLICEFLSDWK

[0173] 346-008 10 DWLRII DWVCSWSDFK

[0174] 346-010 11 SWLRDIWDWISEVLSDFK

[0175] 346-011 12 swlrdiwdwisevlsdfk

[0176] 346-012 13 SWLRDIWDWICEVLSDFR

[0177] 346-013 14 SYLRDIWDYICEVLSDFK

[0178] 346-014 15 SYLRDIWDYISEVLSDFK

[0179] 346-015 16 SYLREIWDYISEVLSDFR

[0180] 346-016 17 SWLREIWDWICEVLSDFK

[0181] 346-017 18 Ac-SWLRDIWDWICEVLSDFK-NH2

[0182] 346-018 19 Ac-swlrdiwdwicevlsdfk-NH2

[0183] 346-019 20 AC-GS LRDI D ICEVLSDFK-NH2

[0184] 346-020 21 Ac-SWLRDIWDWICEVLSDFKTW-NH2

[0185] 346-021 22 Ac-kfdslveciwdwidrlws-NH2 Peptide SEQ ID NO: Peptide Sequence and Modifications

[0186] 346-022 23 Ac-KWLCRIWSWISDVLDDFE-NH2

[0187] 346-023 24 AC-SIWRDWVDLICEFLSDWK-NH2

[0188] 346-024 25 AC-DWLRIIWDWVCSWSDFK-NH2

[0189] 346-025 26 AC-SWLRDIWDWISEVLSDFK-NH2

[0190] 346-026 27 Ac-swlrdiwdwisevlsdfk-NH2

[0191] 346-027 28 AC-SWLRDIWDWICEVLSDFR-NH2

[0192] 346-028 29 AC-SYLRDIWDYICEVLSDFK-NH2

[0193] 346-029 30 AC-SYLRDIWDYISEVLSDFK-NH2

[0194] 346-030 31 AC-SYLREIWDYISEVLSDFR-NH2

[0195] 346-031 32 AC-SWLREIWDWICEVLSDFK-NH2

[0196] 346-032 33 SWLRDIWDWISEVLSDFR

[0197] 346-033 34 SWLRDIWDYISEVLSDFR

[0198] 346-034 35 swlrdiwdyisevlsdfr

[0199] 346-035 36 SWLRDIWDYICEVLSDFR

[0200] 346-036 37 AC-SWLRDIWDWISEVLSDFR-NH2

[0201] 346-037 38 AC-SWLRDIWDYISEVLSDFR-NH2

[0202] 346-038 39 Ac-swlrdiwdyisevlsdfr-NH2

[0203] 346-039 40 AC-SWLRDIWDYICEVLSDFR-NH2

[0204] 346-040 41 AWLRDIWDWICEVLSDFK

[0205] 346-041 42 SALRDIWDWICEVLSDFK

[0206] 346-042 43 SWARDIWDWICEVLSDFK

[0207] 346-043 44 SWLADIWDWICEVLSDFK

[0208] 346-044 45 SWLRAIWDWICEVLSDFK

[0209] 346-045 46 SWLRDAWDWICEVLSDFK

[0210] 346-046 47 SWLRDIADWICEVLSDFK

[0211] 346-047 48 SWLRDIWAWICEVLSDFK

[0212] 346-048 49 SWLRDIWDAICEVLSDFK

[0213] 346-049 50 SWLRDIWDWACEVLSDFK

[0214] 346-050 51 SWLRDIWDWIAEVLSDFK

[0215] 346-051 52 SWLRDIWDWICAVLSDFK

[0216] 346-052 53 SWLRDIWDWICEALSDFK

[0217] 346-053 54 SWLRDIWDWICEVASDFK

[0218] 346-054 55 SWLRDIWDWICEVLADFK

[0219] 346-055 56 SWLRDIWDWICEVLSAFK

[0220] 346-056 57 SWLRDIWDWICEVLSDAK Peptide SEQ ID NO: Peptide Sequence and Modifications

[0221] 346-057 58 SWLRDIWDWICEVLSDFA

[0222] 346-058 59 Ac-SWLRDIWDWICEVLSDFKK

[0223] 346-059 60 KSWLRDIWDWICEVLSDFK-NH2

[0224] 346-060 61 Ac-K(palmitoyl)-SWLRDIWDWICEVLSDFK-NH2

[0225] 346-061 62 Ac-SWLRDIWDWICEVLSDFK-K(palmitoyl)-NH2

[0226] 346-062 63 Ac-SWLRDIWDWICEVLSDFK-amino-PEG12-propionyl-NH2

[0227] 346-063 64 Ac-amino-PEG12-propionyl-SWLRDIWDWICEVLSDFK-NH2

[0228] 65 Ac-SWLRDIWDWICEVLSDFKC(succinimido-propionylaminoethyl-mPEG2K)-

[0229] 346-064

[0230] NH2

[0231] 346-065 66 SWLRSIWDWICEVLSDFK

[0232] 346-066 67 SWLRLIWDWICEVLSDFK

[0233] 346-067 68 SWLRFIWDWICEVLSDFK

[0234] 346-068 69 SWLRDIWDWIIEVLSDFK

[0235] 346-069 70 SWLRDIWDWIWEVLSDFK

[0236] 346-070 71 SWLRDIWDWIREVLSDFK

[0237] 346-071 72 SWLRDIWDWIPEVLSDFK

[0238] 346-072 73 SWLRDIWDWIR

[0239] 346-073 74 SIWRDIWDLAVEAVSDWK

[0240] 346-074 75 SILRDIWDWICEVLSDFK

[0241] 346-075 76 SLWRDIWDWICEVLSDFK

[0242] 346-076 77 SWLRDIWDLICEVLSDFK

[0243] 346-077 78 SWLRDIWDWIVEVLSDFK

[0244] 346-078 79 SWLRDIWDWICEAVSDFK

[0245] 346-079 80 SWLRDIWDWICEVLSDWK

[0246] 346-080 81 SWLQRIWQWISDVLSEFE

[0247] 346-081 82 SWLQDIWDWICEVLSDFK

[0248] 346-082 83 SWLRRIWDWICEVLSDFK

[0249] 346-083 84 SWLRDIWQWICEVLSDFK

[0250] 346-084 85 SWLRDIWDWICDVLSDFK

[0251] 346-085 86 SWLRDIWDWICEVLSEFK

[0252] 346-086 87 SWLRDIWDWICEVLSDFE

[0253] 346-087 88 SWLRAIWDWISEVLSDFR

[0254] 346-088 89 SWLRalWDWISEVLSDFR

[0255] 346-089 90 SWLRAIWDWIsEVLSDFR

[0256] 346-090 91 SWLRAIWDWISEVLSDFr Peptide SEQ ID NO: Peptide Sequence and Modifications

[0257] 346-091 92 Ac-SWLRDIWDWICEVLSDFK-K(oleoyl)-NH2

[0258] 346-092 93 Ac-K(succinyl-D-o-tocopherol)-SWLRDIWDWICEVLSDFK-NH2

[0259] 346-093 94 Ac-SWLRDIWDWICEVLSDFK-K(succinyl-D-o-tocopherol)-NH2

[0260] 346-094 95 SWLRDIWDWICEVLSDFK-K(palmitoyl)-NH2

[0261] 346-095 96 Ac-C(succinimido-propionylaminoethyl-mPEG2K)-SWLRDIWDWICEVLSDFK-

[0262] NH2

[0263] 346-096 97 SWLRDIWDWICEVLSDFK-C(cholesteryloxycarbonylmethyl)-NH2

[0264] 346-097 98 SWLRDIADWIAEVLSDFA

[0265] 346-098 99 SWLRAIADWIAEVLSDFK

[0266] 346-099 100 AC-SWLRDIADWIAEVLSDFA-NH2

[0267] 346-100 101 AC-SWLRAIADWIAEVLSDFK-NH2

[0268] 346-101 102 AC-SWLRDIADWICEVLSDFK-NH2

[0269] 346-102 103 AC-SWLRAIWDWISEVLSDFR-NH2

[0270] 346-103 104 SWLRDIWDWIGEVLSDFK

[0271] 346-104 105 SWLRDIADWIAEVLSDFK

[0272] 346-105 106 SWLRDIGDWIAEVLSDFK

[0273] 346-106 107 SWLRDIADWIAEVLSDFR

[0274] 346-107 108 SWLRDIADWICEVLSDWK

[0275] 346-108 109 SWLRDIADWICEALSDFK

[0276] Upper case single letter amino acid abbreviations indicate / .-isomer; lower case single letter amino acid abbreviations indicate D-isomer.

[0277] TABLE 2. Examples of peptide conjugates based on Peptide 346-001 backbone.

[0278] Conj-Ser-Trp-Leu-Arg-Asp-lle-Trp-Asp-Trp-lle-Cys-Glu-Val-Leu-Ser-Asp-Phe-Lys-NH2 (SEQ ID NO: 110)

[0279] Ac-Ser-Trp-Leu-Arg-Asp-lle-Trp-Asp-Trp-lle-Cys-Glu-Val-Leu-Ser-Asp-Phe-Lys-Conj (SEQ ID NO: 111)

[0280] Conj-Ser-Trp-Leu-Arg-Asp-lle-Trp-Asp-Trp-lle-Cys-Glu-Val-Leu-Ser-Asp-Phe-Lys-Conj (SEQ ID NO: 112) Conj, denotes conjugate

[0281] TABLE 3. Examples of peptides

[0282]

[0283]

[0284]

[0285] Upper case single letter amino acid abbreviations indicate L-isomer; lower case single letter amino acid abbreviations indicate D-isomer.

[0286] EXAMPLE 7 - Formulation 4 (Intravaginal Administration - Therapy)

[0287]

[0118] For intravaginal therapeutic administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147, for example SEQ ID NO: 276, SWLRAIWEWIsEVLsEFRRR-NH2) is formulated as a vaginal insert product for the treatment of HPV infection.

[0288]

[0119] The insert is produced by methods known in the art (65). The inserts typically are 0.1 -2.0 g, preferably 0.5-1 .0 g, in mass, and can have a number of geometries from round to more complex as described in (65). In one embodiment, it may be advantageous to maintain therapeutically active drug concentrations in the vagina for the time period spanning administered doses. One preferred approach to achieving this PK profile lies with using mucoadhesive excipients in the insert, as described below in non-limiting examples.

[0289]

[0120] Hydroxypropyl methylcellulose (molecular weight: 10, 26, 86, 90 and 120 kDa) or Kollidon ® (Sigma- Aldrich) are blended with the antiviral peptide (0.1-20% w / w / , preferably 0.5, 1, 2, and 5% w / w) in a high efficiency mixing device (e.g., speed mixer, 1 min at 3,500 rpm). The mixture is then compressed using a tablet press and an appropriate die to fabricate tablets that are 0.1, 0.25, 0.5, and 1.0 g in mass.

[0290]

[0121] In another embodiment, the following excipients are used instead of hydroxypropyl methylcellulose or Kollidon ©: chitosan and / or locust bean gum and / or pectin (exemplary total concentration, 60-90% w / w), magnesium stearate (exemplary concentration, 0.25-1% w / w).

[0291]

[0122] In another embodiment, the insert is fabricated by granulation with hydrophobic polymers as described in the art (66).

[0292]

[0123] In another embodiment, the insert consists of an osmotic pump, as described in the art (67). Here, the inserts are coated with a semi-permeable, non-swellable outer membrane with a small (e.g., laser-drilled) orifice. The systems are formulated with osmoattractants in the tablet core to promote water diffusion through the outer membrane, leading to a buildup in internal pressure that drives the sustained release of the antiviral peptide through the orifice.

[0293]

[0124] In another example, the self-administration of the antiviral vaginal insert is paired with a companion diagnostic for HPV. The diagnostic in combination with optional telemedicine is used to more efficiently treat the HPV infection.

[0294]

[0125] In another example, a non-hormonal contraceptive from the sAC inhibitor class is included in the insert formulation (0.02-5% w / w / , preferably 0.1, 0.25, 0.5, 1, and 2.5% w / w) in combination with the antiviral peptide to form a multipurpose technology.

[0295] EXAMPLE 8 - Formulation 5 (Intravaginal Administration - On Demand)

[0296]

[0126] For intravaginal on-demand administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147, for example SEQ ID NO: 276, SWLRAIWEWIsEVLsEFRRR-NH2) is formulated as a vaginal insert product for the prevention of HSV infection. The insert is self-administered before, and / or after vaginal intercourse. In some embodiments, after vaginal intercourse is 1 minute, 2 minutes, 5 minutes, 10 minutes, 30 minutes, 1 hour, 2 hours, 3 hours, 6 hours, 12 hours, or 24 hours after vaginal intercourse.

[0297]

[0127] The insert is produced by methods known in the art (65). The inserts typically are 0.5-1.0 g in mass, and can have a number of geometries such as round, as described in (65). In one embodiment it may be advantageous to rapidly obtain therapeutically active drug concentrations in the vagina prior to sex. One preferred approach to achieving this PK profile lies with using fast-dissolving excipients in the insert, as described below in the non-limiting example.

[0298]

[0128] Disintegrants (superdisintegrants), diluents, binding agents, dispersing agents, glidants, osmoattractants, buffering agents, lubricating, and pore formers or channel creators are blended in proportions known in the art to achieve rapid dissolution.

[0129] In another example, a non-hormonal contraceptive from the sAC inhibitor class is included in the insert formulation (0.02-5% w / w / , preferably 0.1, 0.25, 0.5, 1, and 2.5% w / w) in combination with the antiviral peptide to form a multipurpose prevention technology.

[0299] EXAMPLE 9 - Formulation 6 (Intravaginal Administration - Therapy)

[0300]

[0130] For intravaginal therapeutic administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147, for example SEQ ID NO: 276, SWLRAIWEWIsEVLsEFRRR-NH2) is formulated as a vaginal insert product for the treatment of HSV infection.

[0301]

[0131] The insert is produced by methods known in the art (65). The inserts typically are 0.1 -2.0 g, preferably 0.5-1 .0 g, in mass, and can have a number of geometries from round to more complex as described in (65). In one embodiment it may be advantageous to maintain therapeutically active drug concentrations in the vagina for the time period spanning administered doses. One preferred approach to achieving this PK profile lies with using mucoadhesive excipients in the insert, as described below in non-limiting examples.

[0302]

[0132] Hydroxypropyl methylcellulose (molecular weight: 10, 26, 86, 90 and 120 kDa) or Kollidon ® (Sigma- Aldrich) are blended with the antiviral peptide (0.1-20% w / w, preferably 0.5, 1, 2, and 5% w / w) in a high efficiency mixing device (e.g., speed mixer, 1 min at 3,500 rpm). The mixture is then compressed using a tablet press and an appropriate die to fabricate tablets that are 0.1, 0.25, 0.5, and 1.0 g in mass.

[0303]

[0133] In another embodiment, the following excipients are used instead of hydroxypropyl methylcellulose or Kollidon ®: chitosan and / or locust bean gum and / or pectin (exemplary total concentration, 60-90% w / w), magnesium stearate (exemplary concentration, 0.25-1% w / w).

[0304]

[0134] In another embodiment, the insert is fabricated by granulation with hydrophobic polymers as described in the art (66).

[0305]

[0135] In another embodiment, the insert consists of an osmotic pump, as described in the art (67). Here, the inserts are coated with a semi-permeable, non-swellable outer membrane with a small (e.g., laser-drilled) orifice. The systems are formulated with osmoattractants in the tablet core to promote water diffusion through the outer membrane, leading to a buildup in internal pressure that drives the sustained release of the antiviral peptide through the orifice.

[0306]

[0136] In another example, the self-administration of the antiviral vaginal insert is paired with a companion diagnostic for HSV. The diagnostic in combination with optional telemedicine is used to more efficiently treat the HSV infection.

[0307]

[0137] In another example, a non-hormonal contraceptive from the sAC inhibitor class is included in the insert formulation (0.02-5% w / w / , preferably 0.1, 0.25, 0.5, 1, and 2.5% w / w) in combination with the antiviral peptide to form a multipurpose technology.

[0308] EXAMPLE 10 - Formulation 7 (Oral Administration - Therapy)

[0138] For oral therapeutic administration, a peptide described herein (e.g., a peptide comprising the amino acid sequence of SEQ ID NO: 147, for example SEQ ID NO: 276, SWLRAIWEWIsEVLsEFRRR-NH2) is formulated as a cream product for the treatment of HSV infection, including herpes labialis (also known as cold sores) and herpes zoster (also known as shingles) in adults. It is also used to treat chickenpox and cold sores in children.

[0309]

[0139] In a non-limiting example, the cream or ointment contains 0.1-25% w / w of the antimicrobial peptide, preferably 0.25-2.5 % w / w. Many oral creams, gels, and ointments are known in the art. In one embodiment, the ointment consists of a polyethylene glycol (PEG) base.

[0310]

[0140] All references cited herein are incorporated by reference in their entirety as though fully set forth herein. Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. Allen et al., Remington: The Science and Practice of Pharmacy 22nded., Pharmaceutical Press (September 15, 2012); Hornyak etal., Introduction to Nanoscience and Nanotechnology, CRC Press (Boca Raton, FL, 2008); Oxford Textbook of Medicine, Oxford Univ. Press (Oxford, England, UK, May 2010, with 2018 update); Harrison's Principles of Internal Medicine, Vol .1 and 2, 20thed., McGraw-Hill (New York, NY, 2018); Singleton and Sainsbury, Dictionary of Microbiology and Molecular Biology, 3rded., revised ed., J. Wiley & Sons (New York, NY, 2006); Smith, March’s Advanced Organic Chemistry Reactions, Mechanisms and Structure 7thed, J. Wiley & Sons (New York, NY, 2013); and Singleton, Dictionary of DNA and Genome Technology, 3rded., Wiley-Blackwell (Hoboken, NJ, 2012), provide one skilled in the art with a general guide to many of the terms used in the present application.

[0311]

[0141] One skilled in the art will recognize many methods and materials similar or equivalent to those described herein, which could be used in the practice of the present invention. Indeed, the present invention is in no way limited to the methods and materials described. The entire document is intended to be related as a unified disclosure, and it should be understood that all combinations of features described herein (even if described in separate sections) are contemplated, even if the combination of features is not found together in the same sentence, or paragraph, or section of this document.

[0312]

[0142] It should be understood that, while various embodiments in the specification are presented using "comprising” language, under various circumstances, a related embodiment may also be described using "consisting of' or "consisting essentially of” language. The disclosure contemplates embodiments described as "comprising” a feature to include embodiments which "consist of” or "consist essentially of” the feature. The term "a” or "an” refers to one or more, and the terms "a” (or "an”), "one or more,” and "at least one” can be used interchangeably herein. The term "or” should be understood to encompass items in the alternative or together, unless context unambiguously requires otherwise.

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Claims

WHAT IS CLAIMED IS:

1. A peptide comprising the amino acid sequence SWLRXsIWEWIXnEVLXisEXizX^SEQ ID NO: 147), whereinX5 is A, V, or L;X11 is S, D-serine, or phospho-serine;X15 is S, D-serine, or phospho-serine;X17 is F or phospho-tyrosine; andXis is R or phospho-tyrosine.

2. The peptide of claim 1 , wherein X5 is A.

3. The peptide of claim 1 or claim 2, wherein (i) Xu is S and X15 is D-serine or (ii) Xu is D-serine and X15 is S.

4. The peptide of claim 3, wherein Xu is S and X15 is D-serine.

5. The peptide of any one of claims 1-4, wherein X17 is F.

6. The peptide of any one of claims 1-5, wherein Xis is R.

7. The peptide of any one of claims 1-6 comprising the amino acid sequenceSWLRX5lWEWIXiiEVLXi5EXizXi8Xi9X2o (SEQ ID NO: 148), wherein Xig-X2o are independently selected from R or phospho-tyrosine.

8. The peptide of claim 7, wherein X19-X20 are R.

9. The peptide of claim 1 , wherein the peptide comprises the amino acid sequence SWLRAIWEWISEVLsEFRRR (SEQ ID NO: 149).

10. The peptide of any one of claims 1-9, comprising an amide group at the C-terminus.

11. A peptide comprising the amino acid sequence SWLRAIWEWISEVLsEFRRR-NF (SEQ ID NO: 150).

12. The peptide of any one of claims 1-11, wherein the peptide is modified with a lipophilic moiety via a linker bonded to an S or E within the amino acid sequence.

13. The peptide of any one of claims 1-12, wherein the peptide comprises one or more D-amino acids.

14. A targeting peptide comprising the peptide of any one of claims 1-13.

15. The targeting peptide of claim 14, wherein the targeting peptide is a cell-targeting peptide.

16. A prodrug comprising the peptide of any one of claims 1-13 or the targeting peptide of claim 14 or 15.

17. The prodrug of claim 16, wherein the prodrug is a drugamer prodrug.

18. A composition comprising the peptide of any one of claims 1-13, the targeting peptide of claim 14 or 15, or the prodrug of claim 16 or 17.

19. The composition of claim 18, further comprising a contraceptive.

20. The composition of claim 19, wherein the contraceptive is a non-hormonal contraceptive.21 . The composition of claim 19 or 20, wherein the contraceptive is a soluble adenylyl cyclase (sAC) inhibitor or an antibody.

22. The composition of any one of claims 18-21, formulated as an insert, a suppository, a film, a gel, a douche, a cream, an ointment, or a rinse.

23. The composition of any one of claims 18-22, formulated as a long-acting, fast-dissolving, slow- releasing, and / or mucoadhesive composition.

24. An implantable drug delivery device comprising the peptide of any one of claims 1-13, the targeting peptide of claim 14 or 15, the prodrug of claim 16 or 17, or the composition of any one of claims 18-23.

25. The implantable drug delivery device of claim 24, which is an intravaginal ring.

26. The implantable drug delivery device of claim 24, which is a buccal implant.

27. The implantable drug delivery device of any one of claims 24-26, wherein the drug delivery device is an osmotic pump drug delivery device.

28. A method of treating or preventing a viral disease, the method comprising administering to a subject in need thereof the peptide of any one of claims 1-13, the targeting peptide of claim 14 or 15, the prodrug of claim 16 or 17, the composition of any one of claims 18-23, or the drug delivery device of any one of claims 24-27.

29. The method of claim 28, wherein the viral disease is a human immunodeficiency virus (HIV), herpes simplex virus (HSV), human papillomavirus (HPV), hepatitis A virus, hepatitis B virus, hepatitis C virus, dengue virus, West Nile virus, Zika virus, or respiratory virus infection.

30. The method of claim 29, wherein the viral disease is an HIV infection.31 . The method of claim 30, wherein the viral disease is an HIV-1 or HIV-2 infection.

32. The method of claim 29, wherein the viral disease is an HSV infection.

33. The method of claim 32, wherein the viral disease is an HSV-1 or HSV-2 infection.

34. The method of claim 29, wherein the viral disease is a hepatitis virus infection.

35. The method of claim 34, wherein the viral disease is a hepatitis B virus (HBV) infection.

36. The method of claim 29, wherein the viral disease is respiratory virus infection.

37. The method of claim 36, wherein the respiratory virus infection is a coronavirus infection, an influenza infection, or a respiratory syncytial virus (RSV) infection.

38. The method of claim 37, wherein the respiratory virus infection is SARS-CoV-2 infection.

39. The method of any one of claims 28-38, further comprising administering one or more complementary agents.

40. The method of claim 39, wherein the one or more complementary agents comprises a contraceptive or an antibody.41 . The method of claim 40, wherein the contraceptive is a non-hormonal contraceptive.

42. The method of any one of claims 28-41 , wherein the administering comprises systemic administration.

43. The method of claim 42, wherein the systemic administration comprises injection, implantation, or oral administration.

44. The method of claim 42 or claim 43, wherein the systemic administration comprises intramuscular, subcutaneous, intravenous, or buccal administration.

45. The method of any one of claims 28-41 , wherein the administering comprises topical administration.

46. The method of claim 45, wherein the topical administration comprises vaginal, rectal, oral, nasal, or pulmonary administration.

47. The method of claim 46, wherein the topical administration comprises nasal or pulmonary administration.

48. The method of any one of claims 28-47, further comprising detecting the presence of a viral infection.

49. The method of claim 48, wherein detecting the presence of a viral infection comprises performing PCR analysis or antigen analysis on a sample obtained from the subject.

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