Compositions and methods for improving systemic delivery, tolerability and efficacy of cationic macrocyclic peptides

Abstract

Compositions are provided for the formulation of theta defensins and / or theta defensin analogs that are highly suitable for parenteral administration. Compositions are provided for the formulation of theta defensins and / or theta defensin analogs that are highly suitable for parenteral administration.SOLUTION: Such formulations provide the theta defensin and / or theta defensin analog in a slightly acidic buffer containing propylene glycol. Surprisingly, the inventors have found that such formulations increase the bioavailability of θ-defensins and / or θ-defensin analogues so provided by at least a factor of 10 compared to conventional isotonic saline solutions, and dramatically improve bioavailability in human subjects compared to animal models. The inventors have also found that such formulations advantageously exhibit low viscosity at high peptide concentrations, allowing reduced injection volumes and sterilization by simple filtration.SELECTED DRAWING: Figure 2

Description

[Technical Field]

[0001] The field of the invention is improving the systemic delivery or pharmacological efficacy of cationic peptide drugs, particularly macrocyclic theta-defensins and / or analogs thereof. [Background technology]

[0002] The following description contains information that may be useful in understanding the present invention. No admission is made that any of the information provided herein is prior art or relevant to the presently claimed invention, or that any publication referenced specifically or implicitly is prior art.

[0003] Inflammation is a complex defensive response to pathogens, tissue injury, and exposure to irritants, involving vascular changes, immune cell recruitment, and the release of various chemical and peptide mediators. While inflammation helps eliminate the initial cause of cellular injury and clear necrotic cells from damaged tissue, the inflammatory response itself can be harmful. For example, chronic inflammation resulting from autoimmune disease can contribute to damage to affected tissues. Similarly, inflammation resulting from acute processes such as viral or bacterial infections can result in tissue damage and septic shock, which is exacerbated by chronic inflammation if the initial injury results in prolonged, unresolved inflammation. Chronic inflammation often leads to tissue scarring, fibrosis, and loss of function. Additionally, the pain and swelling associated with inflammation can be debilitating, especially when it is the result of a chronic condition.

[0004] Unfortunately, current methods for treating inflammation suffer from many drawbacks. For example, traditional pharmaceutical approaches (e.g., treatment with steroidal or nonsteroidal anti-inflammatory drugs) only provide short-term relief, often at the expense of significant side effects that limit the use of such drugs. More recently, "biologics" (e.g., humanized monoclonal antibodies against pro-inflammatory cytokines) have been used to treat certain chronic conditions characterized by inflammation, but such approaches necessarily target only a single inflammatory mechanism and can lead to immunosuppression or even an immunocompromised state in the treated individual. In addition, antibody-based biologics can require the intravenous administration of relatively large volumes of fluid, which are generally administered by infusion under the care of trained medical personnel.

[0005] Mammalian defensins are cationic, tridisulfide-containing peptides comprising three structurally distinct, distinct subfamilies. α- and β-defensins are linear peptides ranging in length from 29 to approximately 60 amino acids, with free amino- and carboxyl-terminal amino acids. α- and β-defensins have similar three-dimensional topologies but differ in disulfide bonds (Non-Patent Document 1). In contrast, naturally occurring θ-defensins are backbone-cyclized 18-amino acid peptides that are distinct from α- and β-defensins both structurally and functionally. Defensins contribute to host defense as antibacterial agents (Non-Patent Document 2) and by regulating inflammation (Non-Patent Document 3) and adaptive immune responses (Non-Patent Document 4). All publications herein are incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference. If the definition or use of a term in an incorporated reference contradicts or is contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.

[0006] θ-Defensins are expressed in Old World monkeys (e.g., macaques and baboons) and are the only known cyclic proteins in animals (Non-Patent Document 5). The basic θ-defensin backbone structure is generated by head-to-tail splicing of two nonapeptide precursors. In rhesus monkeys, alternative binary splicing of nonapeptides encoded by three precursor genes provides six θ-defensin isoforms, rhesus theta-defensins RTD-1 to RTD-6 (SEQ ID NOS: 1 to 6) (Non-Patent Document 6, Non-Patent Document 7). In baboons, alternative nonapeptide splicing generates ten θ-defensin isoforms, baboon theta-defensins BTD-1 to BTD-10 (SEQ ID NOS: 7 to 16) (Non-Patent Document 8). θ-Defensins are expressed at high levels in neutrophil granules and monocytes of these species. These θ-defensins play a major role in the antibacterial activity of rhesus monkey neutrophil granule extracts. The RTD-1 isoform is the most abundant θ-defensin in macaques, constituting approximately 55% of the total θ-defensin content of rhesus monkey neutrophils (Non-Patent Document 9).

[0007] Humans and other apes lack θ-defensins due to the presence of a stop codon mutation within the pre-coding sequence of the θ-defensin gene in these species (Non-Patent Document 10). It has been suggested that the expression of θ-defensins in Old World monkeys is related to differences in the immune and inflammatory responses of these non-human primates compared with those of humans (Non-Patent Document 5).

[0008] Although α-, β-, and θ-defensins were initially identified based on their broad-spectrum antibacterial properties, subsequent studies have revealed distinct immunoregulatory roles (Non-Patent Document 11). For example, some α- and β-defensins mediate chemotaxis for T cells, neutrophils, dendritic cells, and monocytes (Non-Patent Document 4, Non-Patent Document 12, Non-Patent Document 13, Non-Patent Document 14), and induce the secretion of proinflammatory cytokines from activated dendritic cells, peripheral blood mononuclear cells, and epithelial cells (Non-Patent Document 3, Non-Patent Document 15, Non-Patent Document 16, Non-Patent Document 17, Non-Patent Document 18, Non-Patent Document 19, Non-Patent Document 20).

[0009] In contrast to such proinflammatory activity, θ-defensins have recently been reported to have anti-inflammatory properties both in vitro and in vivo. For example, RTD-1 was found to be a potent inhibitor of cytokine secretion by human peripheral blood leukocytes stimulated with various Toll-like receptor (TLR) agonists (Non-Patent Document 21). Naturally occurring θ-defensin isoforms (RTD1-6) have varying efficacy in reducing TNF in lipopolysaccharide- or E. coli-stimulated leukocytes (Non-Patent Document 21). RTD-1 has also been shown to reduce inflammatory cytokines, including TNF-α, IL-1β, and several chemokines, in a mouse model of SARS coronavirus infection (Non-Patent Document 22), as well as in E. coli peritonitis and polymicrobial sepsis (Non-Patent Document 21).

[0010] Recently, Selsted and Tran (Patent Document 1) demonstrated that θ-defensins and θ-defensin analogs that retain the θ-defensin core structure are effective in treating chronic inflammatory conditions such as rheumatoid arthritis. However, it is unclear which formulations of these small cyclic peptides are optimal for achieving therapeutic efficacy at sufficiently low drug exposure to limit potential adverse events, including those associated with injection site reactions after subcutaneous administration.

[0011] Jeong et al. (Patent Document 2) describes a method for improving the pharmacodynamic effects of some peptide drugs by forming an emulsion in the presence of a surfactant and an organic solvent in an aqueous solution, followed by removal of the organic solvent before administration. Various alcohols and polyols, including propylene glycol, are listed as suitable organic solvents that are removed before administration. However, it is unclear whether this approach would be effective for small, basic, cyclic peptides such as theta-defensins.

[0012] Patent Document 3 (Engelud et al.) and Patent Document 4 (Knudsen et al.) describe the use of various polyols as isotonicity agents in peptide pharmaceutical formulations for parenteral administration. Similarly, Patent Document 5 (Pedersen et al.) describes the use of propylene glycol instead of various sugars as an isotonicity agent to prevent the formation of deposits that may interfere with the function of equipment and needles used to administer various specific peptide drugs. However, this reference does not describe any effects other than the reduction in solid residue provided by such formulations, and there is no evidence or suggestion that the use of such compounds has any effect on the bioavailability or pharmacodynamic effects of the peptide drugs so prepared.

[0013] Patent document 6 (Sonavaria et al.) describes an injectable formulation for peptide drugs containing high concentrations (up to 99%) of organic solvents (including polyols), which are said to improve peptide stability. However, injection of such high concentrations of organic solvents is associated with significant negative effects, including pain and swelling. [Prior art documents] [Patent documents]

[0014] [Patent Document 1] US Patent Application Publication No. 2013 / 0157964 [Patent Document 2] International Publication No. 02 / 064166 [Patent Document 3] US Patent Application Publication No. 2006 / 084605 [Patent Document 4] US Patent Application Publication No. 2006 / 0287221 [Patent Document 5] US Patent Application Publication No. 2007 / 0010424 [Patent Document 6] International Publication No. 2016 / 059593 [Non-patent literature]

[0015] [Non-Patent Document 1] Selsted and Ouellette, Nat Immunol 6:551-557 (2005) [Non-patent document 2] Ericksen et al., Antimicrob Agents Chemother 49:269-275 (2005) [Non-patent document 3] Khine et al., Blood 107:2936-2942 (2006) [Non-patent document 4] Chertov et al., J Biol Chem 271:2935-2940 (1996) [Non-Patent Document 5] Lehrer et al., J Biol Chem 287:27014-27019 (2012) [Non-patent document 6] Tang et al., Science 286:498-502 (1999) [Non-Patent Document 7] Leonova et al., J Leukoc Biol 70:461-464 (2001) [Non-patent document 8] Garcia et al., Infect Immun 76:5883-5891 (2001) [Non-Patent Document 9] Tongaonkar et al., J Leukoc Biol 89:283-290 (2011) [Non-Patent Document 10] Nguyen et al., Peptides 24:1647-1654 (2003) [Non-Patent Document 11] Yang et al., Annu Rev Immunol 22:181-215 (2004) [Non-Patent Document 12] Yang et al., Science 286:525-528 (1999) [Non-Patent Document 13] Grigat et al., J Immunol 179:3958-3965 (2007) [Non-Patent Document 14] Soruri et al., Eur J Immunol 37:2474-2486 (2007) [Non-Patent Document 15] Boniotto et al., Antimicrob Agents Chemother 50:1433-1441 (2006) [Non-Patent Document 16] Ito et al., Tohoku J Exp Med 227:39-48 (2012) [Non-Patent Document 17] Yin et al., Immunol 11:37(2010) [Non-Patent Document 18] Niyonsaba et al., J Immunol 175:1776-1784 (2005) [Non-Patent Document 19] Li et al., Invest Ophthalmol Vis Sci 50:644-653 (2009) [Non-Patent Document 20] Syeda et al., J Cell Physiol 214:820-827 (2008) [Non-Patent Document 21] Schaal et al., PLoS One 7, e51337 (2012) [Non-Patent Document 22] Wohlford-Lenane et al. J Virol 83:11385-11390 (2009) Summary of the Invention [Problem to be solved by the invention]

[0016] Thus, there remains a need for methods and compositions that provide the pharmacodynamic effects of administered defensins suitable to achieve a sufficient therapeutic effect. [Means for solving the problem]

[0017] The present subject matter provides compositions and methods for providing injectable formulations of theta-defensins and theta-defensin analogs that offer improved bioavailability compared to conventional isotonic saline solutions. Such formulations also have low viscosity, allowing for scalable sterilization by filtration.

[0018] One embodiment of the inventive concept is a method of treating an individual with a chronic inflammatory condition (e.g., rheumatoid arthritis, inflammatory bowel disease, cancer-related inflammation, diabetes, and / or a chronic disease characterized by dysregulated or unresolved chronic inflammation) by providing the θ-defensin or θ-defensin analog in an aqueous solution comprising the θ-defensin or θ-defensin analog and 0.5% to 1.5% v / v propylene glycol and having a pH of 5.0 to 7.0, and administering the aqueous solution by subcutaneous injection to the individual in need of treatment. In some embodiments, the aqueous solution comprises acetate. The aqueous solution provides increased pharmacological potency or therapeutic effect of the θ-defensin or θ-defensin analog compared to a solution of the θ-defensin or θ-defensin analog in normal saline. The pharmacodynamic effect of a θ-defensin or θ-defensin analog can be increased by at least 10 to 40 times compared to a similar concentration of theta-defensin or θ-defensin analog provided in normal saline solution. -1The θ-defensin analog utilized in the method may be a cyclic icosipeptide, cyclic eneadecapeptide, cyclic octadecapeptide, cyclic heptadecapeptide, cyclic hexadecapeptide, cyclic pentadecapeptide, cyclic tetradecapeptide, cyclic tridecapeptide, cyclic dodecapeptide, cyclic hendecapeptide, or cyclic decapeptide. In a preferred embodiment, the aqueous solution comprises 1% v / v propylene glycol and 20 mM acetate, and has a pH of 6.0.

[0019] Another embodiment of the inventive concept is a method for treating a patient with a steroid hormone, comprising administering to a patient a steroid hormone ... -1 to 50 mg mL -1 (or greater) and 0.5% to 1.5% v / v propylene glycol, and passing the aqueous buffer through a filter having a pore size of 0.2 μm or less. The aqueous buffer may contain acetate, and the resulting θ-defensin preparation may have a pH of 5.0 to 7.0.

[0020] Another embodiment of the inventive concept is a pharmaceutical composition for the treatment of chronic inflammatory conditions (such as rheumatoid arthritis, inflammatory bowel disease, cancer-related inflammation, diabetes, and / or chronic diseases characterized by dysregulated or unresolved chronic inflammation), wherein the pharmaceutical composition is administered in an amount of 50 mg / mL. -1The pharmaceutical composition may contain up to 1000 mg of a θ-defensin or θ-defensin analog and 0.5% to 1.5% v / v propylene glycol, and may contain acetate. The pharmaceutical composition may have a pH of 5.0 to 7.0 and, upon parenteral administration, provides an increased pharmacodynamic effect of the θ-defensin or θ-defensin analog compared to a corresponding pharmaceutical composition prepared without propylene glycol. The pharmacological potency of the θ-defensin or θ-defensin analog is increased by at least 10- to 40-fold compared to the same concentration of the θ-defensin or θ-defensin analog provided in normal saline solution. Such pharmaceutical compositions may be formulated for subcutaneous, intramuscular, and / or intravenous injection. Suitable θ-defensin analogs may be cyclic icosipeptides, cyclic eneadecapeptides, cyclic octadecapeptides, cyclic heptadecapeptides, cyclic hexadecapeptides, cyclic pentadecapeptides, cyclic tetradecapeptides, cyclic tridecapeptides, cyclic dodecapeptides, cyclic hendecapeptides, or cyclic decapeptides. In one preferred embodiment, the pharmaceutical composition contains 1% v / v propylene glycol and 20 mM acetate, and has a pH of 6.0.

[0021] Another embodiment of the inventive concept is a 50 mg / mL steroidal anti-inflammatory drug for the treatment of chronic inflammatory conditions (such as rheumatoid arthritis, inflammatory bowel disease, cancer-related inflammation, diabetes, and / or chronic diseases characterized by dysregulated or unresolved chronic inflammation). -1The present invention relates to a method for the use of a θ-defensin or a θ-defensin analog in a solution containing 0.5% to 1.5% v / v propylene glycol at a concentration of up to 5.0%. The solution may contain acetate. The solution is formulated for parenteral administration and may have a pH of 5.0 to 7.0, providing an increased pharmacodynamic effect of the θ-defensin or a θ-defensin analog compared to a corresponding amount of the θ-defensin or a θ-defensin analog prepared without propylene glycol. The pharmacological potency of the θ-defensin or a θ-defensin analog may be increased by at least 10- to 40-fold upon parenteral administration compared to a similar concentration of the θ-defensin or a θ-defensin analog provided in normal saline solution. Suitable routes of parenteral administration include subcutaneous, intramuscular, and / or intravenous injection. Suitable theta-defensin analogs include cyclic icosipeptides, cyclic eneadecapeptides, cyclic octadecapeptides, cyclic heptadecapeptides, cyclic hexadecapeptides, cyclic pentadecapeptides, cyclic tetradecapeptides, cyclic tridecapeptides, cyclic dodecapeptides, cyclic hendecapeptides, and / or cyclic decapeptides. In one preferred embodiment, the composition comprises 1% v / v propylene glycol and 20 mM acetate and has a pH of 6.0.

[0022] Another embodiment of the inventive concept is a 50 mg mL dose of 100 mg ... -1The present invention relates to a method for administering a θ-defensin or a θ-defensin analog in a solution containing 0.5% to 1.5% propylene glycol. The solution may contain acetate. The composition is formulated for parenteral administration, may have a pH of 5.0 to 7.0, and provides an increased pharmacodynamic effect of the θ-defensin or a θ-defensin analog compared to a corresponding amount of the θ-defensin or a θ-defensin analog prepared without propylene glycol. The pharmacological potency of the θ-defensin or a θ-defensin analog may be increased by at least 10- to 40-fold upon parenteral administration compared to a similar concentration of the θ-defensin or a θ-defensin analog provided in normal saline solution. Suitable routes of parenteral administration include subcutaneous, intramuscular, and / or intravenous injection. Suitable theta-defensin analogs include cyclic icosipeptides, cyclic enneadecapeptides, cyclic octadecapeptides, cyclic heptadecapeptides, cyclic hexadecapeptides, cyclic pentadecapeptides, cyclic tetradecapeptides, cyclic tridecapeptides, cyclic dodecapeptides, cyclic hendecapeptides, and / or cyclic decapeptides. In one preferred embodiment, the composition comprises 1% v / v propylene glycol and 20 mM acetate and has a pH of 6.0.

[0023] Various objects, features, aspects and advantages of the present subject matter will become more apparent from the following detailed description of preferred embodiments, taken in conjunction with the accompanying drawing figures in which like numerals represent like elements. [Brief explanation of the drawings]

[0024] [Figure 1] FIG. 1 shows representative data showing the effect of subcutaneous injection of RTD-1 in saline in an animal model of rheumatoid arthritis. [Figure 2] FIG. 1 shows typical effects of subcutaneous injection of RTD-1 in saline containing 1% (v / v) propylene glycol in an animal model of rheumatoid arthritis. [Figure 3]FIG. 1 shows typical effects of subcutaneous injection of RTD-1 in saline containing 1% (v / v) propylene glycol or in 20 mM Na acetate containing 1% (v / v) propylene glycol in an animal model of rheumatoid arthritis. [Figure 4A] FIG. 1 shows a graph of plasma theta-defensin concentrations (ng / mL) versus time (hours) in male rats on day 1 of a study in which a theta-defensin formulation of the present inventive concept was injected subcutaneously. [Figure 4B] FIG. 1 shows a graph of plasma theta-defensin concentrations (ng / mL) versus time (hours) in female rats on day 1 of a study in which theta-defensin formulations of the present inventive concepts were injected subcutaneously. [Figure 5A] FIG. 1 shows a graph of plasma theta-defensin concentrations (ng / mL) versus time (hours) in male rats on day 13 of a study in which a theta-defensin formulation of the present inventive concept was injected subcutaneously. [Figure 5B] FIG. 1 shows a graph of plasma theta-defensin concentrations (ng / mL) versus time (hours) in female rats on day 13 of a study in which a theta-defensin formulation of the present inventive concept was injected subcutaneously. [Figure 6A] FIG. 1 shows a graph of plasma theta-defensin concentrations (ng / mL) versus time (hours) in male rats on day 41 of a study in which a theta-defensin formulation of the present inventive concept was injected subcutaneously. [Figure 6B] FIG. 1 shows a graph of plasma theta-defensin concentrations (ng / mL) versus time (hours) in female rats on day 41 of a study in which a theta-defensin formulation of the present inventive concept was injected subcutaneously. [Figure 7A] FIG. 10 is a graph showing the results of a linearity study of Cmax versus theta-defensin dose in male and female rats on day 1 of the study in which a theta-defensin formulation of the present invention was injected subcutaneously. [Figure 7B] FIG. 10 is a graph showing the results of a linearity study of Cmax versus theta-defensin dose in male and female rats on day 41 of the study in which a theta-defensin formulation of the present invention was injected subcutaneously. [Figure 8A]FIG. 10 shows a graph depicting the results of a linearity study of AUC0-TLast versus theta-defensin dose in male and female rats on day 1 of the study in which a theta-defensin formulation of the present invention was injected subcutaneously. [Figure 8B] FIG. 10 is a graph showing the results of a linearity study of AUC0-TLast versus theta-defensin dose in male and female rats on day 41 of the study in which a theta-defensin formulation of the present invention was injected subcutaneously. [Figure 9] FIG. 1 shows typical results of measuring plasma θ-defensin concentrations (ng / mL) over time in different treatment groups in a human clinical trial in which a θ-defensin formulation of the inventive concept was injected subcutaneously. [Figure 10A] FIG. 1 shows a typical dependence of Cmax (ng / mL) on dose (μg / kg) of a theta-defensin formulation of the present inventive concept injected subcutaneously in a human clinical trial. [Figure 10B] FIG. 1 shows a typical dependence of AUC0-TLast on dose (μg / kg) of a theta-defensin formulation of the present inventive concept injected subcutaneously in a human clinical trial. [Figure 11A] FIG. 1 shows a typical dependence of Cmax (ng / mL) on total theta-defensin dose (mg) for a theta-defensin formulation of the present invention injected subcutaneously in a human clinical trial. [Figure 11B] FIG. 1 shows the typical dependence of AUC0-TLast on total theta-defensin dose (mg) for a theta-defensin formulation of the present invention injected subcutaneously in a human clinical trial. DETAILED DESCRIPTION OF THE INVENTION

[0025] The following discussion provides a number of exemplary embodiments of the inventive subject matter. While each embodiment represents a single combination of the inventive elements, the inventive subject matter is considered to include all possible combinations of the elements of the present disclosure. Thus, if one embodiment comprises elements A, B, and C, and a second embodiment comprises elements B and D, the inventive subject matter is also considered to include other remaining combinations of A, B, C, or D, even if not explicitly disclosed.

[0026] The present subject matter provides devices, systems, and methods in which 0.5% to 1.5% propylene glycol in a slightly acidic (e.g., pH 5-7) aqueous solution has surprisingly been found to increase the preclinical efficacy of parenterally administered θ-defensins and / or θ-defensin analogs by at least 20-fold compared to conventional neutral isotonic saline solutions. This allows for convenient subcutaneous administration of relatively small volumes (e.g., about 1 mL) of θ-defensins in amounts effective for treating chronic inflammatory conditions such as rheumatoid arthritis, inflammatory bowel disease, or diabetes, or other conditions resulting from a dysregulated inflammatory response in humans. The addition of propylene glycol at these concentrations unexpectedly results in the production of concentrated (e.g., 10 mg mL) doses of θ-defensins. -1 It was also found to dramatically reduce the viscosity of the solution, allowing for simple, convenient, and scalable sterilization by filtration. Furthermore, injection site reactions of the θ-defensin formulations were significantly reduced compared to those observed when isotonic saline was used as the formulation vehicle.

[0027] Suitable theta-defensins include those peptides found in mammals that express natural theta-defensins, and may include one or more theta-defensins derived from untranslated genes present in some primate species (e.g., Homo sapiens). In some embodiments of the inventive concept, the one or more theta-defensins may correspond to those found in rhesus monkeys (Macaca mulatta), e.g., RTD-1 (SEQ ID NO: 1), RTD-2 (SEQ ID NO: 2), RTD-3 (SEQ ID NO: 3), RTD-4 (SEQ ID NO: 4), RTD-5 (SEQ ID NO: 5), and / or RTD-6 (SEQ ID NO: 6). In other embodiments of the inventive concept, the one or more theta-defensins may correspond to those found in the Anubis baboon (Papio anubis), e.g., BTD-1 (SEQ ID NO:7), BTD-2 (SEQ ID NO:8), BTD-3 (SEQ ID NO:9), BTD-4 (SEQ ID NO:10), BTD-5 (SEQ ID NO:11), and / or BTD-6 (SEQ ID NO:12), BTD-7 (SEQ ID NO:13), BTD-8 (SEQ ID NO:14), BTD-9 (SEQ ID NO:15), and / or BTD-10 (SEQ ID NO:16).

[0028] Several embodiments of the present application that describe and utilize θ-defensins include θ-defensin analogs. The term θ-defensin analog refers to a cyclic peptide having about 40%, 50%, 60%, 70%, 80%, 90% or more sequence identity with a naturally occurring θ-defensin peptide sequence. The θ-defensin analog may incorporate one, two, three, or more of the core features of naturally occurring θ-defensins. Exemplary core features include a cyclic structure, the presence of one, two, three, or more disulfide bonds within the peptide (e.g., between cysteine ​​pairs in the analog), a positive charge in solution under physiological conditions, and the presence of a beta-pleated sheet secondary structure. Such θ-defensin analogs may contain 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more than 20 amino acids, and in some embodiments, may incorporate non-naturally occurring amino acids. Analogs of theta-defensins may contain one or more L-amino acids, one or more D-amino acids, and / or a mixture of L- and D-amino acids. In some embodiments, non-peptide bonds may be utilized between adjacent amino acid residues of theta-defensin analogs. Theta-defensin analogs may exhibit one or more deletions or substitutions of amino acids from the native theta-defensin sequence. Such substitutions may be conservative (e.g., the substituted amino acids retain charge, hydrophobicity, hydrophilicity, and / or steric properties). In some embodiments, theta-defensin analogs may include grafting or attachment of non-peptide moieties, such as polyethylene glycol and / or other hydrophilic polymers, cell receptor targeting moieties, and / or moieties that aid in processing / purification. Examples of suitable theta-defensin analogs based on RTD-1 (SEQ ID NO: 1) are provided as SEQ ID NO: 17 (RTD-1-27), SEQ ID NO: 18 (RTD-1-28), and SEQ ID NO: 19 (RTD-1-29).

[0029] Provided herein are compositions and methods for enhancing the pharmacological effects of θ-defensins and / or θ-defensin analogs utilized in the treatment of chronic inflammatory conditions (e.g., rheumatoid arthritis and / or diabetes). Such compositions comprise about 0.5% to about 1.5% propylene glycol in a weakly acidic aqueous solution (e.g., about pH 5-7), and contain 50 mg mL -1 The claimed compositions are suitable for subcutaneous injection of θ-defensins at concentrations up to 1000 mg / mL. Such formulations have surprisingly been found to dramatically (e.g., at least 10-fold greater) increase the pharmacological effect or potency of the administered θ-defensins compared to θ-defensins provided at the same or similar concentrations in conventional saline solutions and / or solutions without propylene glycol. This is shown in Figures 1, 2, and 3 below. Such formulations have also been found to have significantly reduced viscosity compared to θ-defensins provided at similar concentrations in conventional normal saline solutions (e.g., phosphate-buffered saline, pH 7-7.5), allowing for sterilization by filtration. Furthermore, the claimed compositions have significantly reduced injection site reactions compared to isotonic saline vehicles.

[0030] Figures 1, 2, and 3 show the results of treatment with subcutaneous injections of the theta-defensin RTD-1 in an animal model of rheumatoid arthritis, where the theta-defensin was provided either in conventional normal saline vehicle or in normal saline containing a low concentration (1% v / v) of propylene glycol. Rats with established pristane-induced arthritis were treated daily with subcutaneous injections of RTD-1 formulated in normal saline (Figure 1) or in saline solution containing 1% propylene glycol (Figure 2) at the doses indicated. The lowest effective dose of RTD-1 in normal saline diluent was 1 mg / kg, and the maximal pronounced effect was at 3 mg / kg (see Figure 1). The inclusion of 1% propylene glycol in the vehicle significantly reduced the effective RTD-1 dose (to 0.08 mg / kg, see Figure 2), with 0.08 mg / kg (the lowest dose tested) producing a maximal anti-arthritic effect equivalent to a 3 mg / kg dose of RTD-1 in saline (i.e., the maximal effect observed). A comparable improvement in pharmacological effect was obtained when RTD-1 was formulated in a buffered (20 mM sodium acetate) aqueous diluent containing 1% propylene glycol (Figure 3). Applicants believe that similar effects may be achieved with lower doses of theta-defensin.

[0031] Surprisingly, such formulations were also found to have significantly reduced viscosity compared to θ-defensins provided at similar concentrations in conventional normal saline solutions (e.g., isotonic phosphate buffered saline, pH 7-7.5), allowing for sterilization by filtration.

[0032] We have previously demonstrated that θ-defensins and θ-defensin analogs (e.g., cyclic octadecapeptides and / or cyclic tetradecapeptides) have significant effects in reducing inflammation in animal models of chronic inflammatory diseases, such as pristane-treated rats. θ-defensins are readily soluble in aqueous solutions. In initial studies, θ-defensins were prepared in conventional normal saline solutions and readily administered by subcutaneous injection in mice without apparent systemic adverse effects; however, at high concentrations of θ-defensins, fat necrosis in the subcutaneous tissue was observed. Similarly, in rat and dog animal models, low concentrations of θ-defensins were well tolerated systemically and locally. However, at high concentrations, injection of θ-defensins in normal saline resulted in local inflammation and swelling at the injection site in a dose-dependent manner. The swelling and inflammation were also found to persist for several weeks. However, relatively high concentrations of θ-defensins (e.g., approximately 10–50 mg mL) -1 ) may be necessary in human therapy to provide the necessary θ-defensin dosage while maintaining a volume suitable for subcutaneous injection.

[0033] Microscopic studies of injection sites where θ-defensins were administered in conventional saline solution showed localized inflammation and necrotic changes at the injection site in canine and porcine test subjects. Without wishing to be bound by theory, the inventors believe that θ-defensins may interact with components of the extracellular matrix found in the skin of some animals at the subcutaneous injection depth, causing θ-defensin precipitation and / or the formation of precipitation complexes within this tissue layer. Studies of the interaction of RTD-1 with whole blood, plasma collected after treatment with anticoagulants, and serum have shown that θ-defensins interact with fibrinogen to form insoluble complexes. Therefore, the inventors believe that θ-defensins may interact with fibrinogen and / or fibrinogen-like proteins associated with the extracellular matrix.

[0034] The inventors have found that the use of hypotonic saline supplemented with poloxamer and other conventional excipients does not reduce or eliminate the injection site inflammation and tissue injury induced by this θ-defensin. Surprisingly, low concentrations of propylene glycol are comparable to high concentrations (e.g., 10 mg mL -1 Propylene glycol has also been effective in providing solubility for θ-defensins (e.g., 0.1% or more) and reducing or preventing swelling and / or inflammation upon subcutaneous injection of high concentrations of θ-defensins into susceptible species. Propylene glycol is freely miscible with water, available as a pharmaceutical-grade sterile liquid, and generally recognized as safe. A relatively narrow range of propylene glycol concentrations has been found to be effective in reducing and / or eliminating side effects upon injection of high concentrations of θ-defensins. In some embodiments, an effective concentration of propylene glycol in water has been found to be between about 0.4% and about 1.6% (v / v). In other embodiments, an effective concentration of propylene glycol in water has been found to be between about 0.5% and about 1.5% (v / v). In one preferred embodiment, an effective concentration of propylene glycol in water is about 1% (v / v).

[0035] Further research surprisingly demonstrated that using a propylene glycol / water solvent with a weakly acidic pH (e.g., pH about 5-7) further improved results. In a preferred embodiment, the propylene glycol / water solvent has a pH of about 6.0 and a final osmolality of about 180-230 mOsm. The pH of the propylene glycol / water solvent can be maintained by using a buffer species at a low molarity (e.g., less than about 50 mM). Suitable buffer concentrations can be, for example, in the range of 1 mM to 50 mM, 5 mM to 35 mM, 10 mM to 30 mM, or about 20 mM. Suitable buffer species can be salts of organic acids such as acetic acid, citric acid, malic acid, tartaric acid, HEPES, MES, etc. In other embodiments, the buffer species can be zwitterionic species with a suitable pKa. In a preferred embodiment, the buffer species is an acetate salt (e.g., sodium acetate) at a concentration of about 20 mM, providing a pH of about 6.0. Surprisingly, the inventors have found that the use of even small amounts (e.g., less than 50 mM) of saline (e.g., NaCl) results in adverse reactions to subcutaneous injection of theta-defensins. Preferred embodiments of aqueous solvent systems used for subcutaneous administration may exclude NaCl and similar salts.

[0036] Surprisingly, the inventors found that theta-defensins are freely soluble in such acidic propylene glycol / water solvent systems and that they are readily soluble in solutions containing 12.5 mg mL of θ-defensin after subcutaneous injection of RTD-1 in rats and dogs. -1 Only transient skin reactions were observed at concentrations up to 50 mg / mL -1 This is the estimated 10 mg mL of θ-defensin that is expected to be required to provide a dosage suitable for human therapy. -1 Supports concentration.

[0037] As described above, the inventors unexpectedly discovered that the use of a propylene glycol / water solvent system dramatically increases the pharmacological potency of subcutaneously administered theta-defensins compared to conventional saline formulations, in addition to eliminating the inflammation associated with saline-based formulations. The preclinical efficacy enhancement of the weakly acidic propylene glycol / water in established pristane-induced arthritis can be at least 5-fold, 10-fold, 15-fold, 20-fold, 30-fold, 40-fold, 50-fold, or even greater than a similar amount provided in conventional saline solution. For example, subcutaneous injection of RTD-1 provided at 0.08 mg / kg in 1% propylene glycol + 20 mM acetate, pH 6, was found to be equivalently effective in alleviating the symptoms of rheumatoid arthritis as administration of RTD-1 at 3 mg / kg in isotonic saline. This suggests at least a 37-fold increase in pharmacological potency, however, because the symptomatic relief provided is an upper limit of the benefit conferred by theta-defensin administered in isotonic saline, the actual increase in pharmacodynamic effect may be greater than 37-fold (e.g., 40-fold, 50-fold, 70-fold, 100-fold, or greater), which advantageously reduces the amount of theta-defensin required to provide adequate therapy.

[0038] Sterilization of conventional solutions used for injection is typically accomplished by filtration, using filters with pore sizes of 0.2 μm or less. However, this sterilization by filtration process is problematic for protein drug solutions because they are typically too viscous for efficient sterile filtration at the high protein concentrations desired to provide low injection volumes. As used herein, such viscous solutions refer to solutions or dispersions that have such high internal resistance to flow that filtration is difficult or impossible (e.g., requiring pressures in excess of the burst pressure of the filtration membrane). The viscosity of such viscous solutions can be 105 mPa·s (105 centipoise (cp)) or greater. In one embodiment, the viscosity of such viscous solutions is at least about 90 to about 95 mPa·s (about 90 to about 95 cp). In another embodiment, the viscosity of such viscous solutions is at least about 40 mPa·s (about 40 cp). In yet another embodiment, the viscosity of such a viscous solution is at least greater than the viscosity of water (ie, greater than about 1.0 mPa·s (about 1.0 cP)).

[0039] During the course of their research, the inventors discovered that although theta-defensins are highly soluble, aqueous solutions can become very viscous as the protein concentration increases. Indeed, conventional saline solutions of theta-defensins at concentrations of 2% w / v or greater were found to be too viscous to allow sterilization by filtration on a reasonable and / or manufacturable scale. Surprisingly, similar solutions prepared in propylene glycol / water solvents had low viscosities at high theta-defensin concentrations (e.g., 2% w / v, 5% w / v, 10% w / v, or higher). This allows for simple and scalable sterilization of such preparations using conventional filtration through media of approximately 0.2 μm or less.

[0040] Embodiments of the present concept include a method for treating conditions associated with chronic inflammation by subcutaneous injection of a pharmaceutical composition containing one or more θ-defensins in an acidic aqueous solution containing 0.5% to 1.5% propylene glycol. Suitable conditions include rheumatoid arthritis, inflammatory bowel disease, diabetes, and other conditions resulting from a dysregulated inflammatory response. The pharmaceutical composition may contain a single species of θ-defensin or two or more species of θ-defensins. The pharmaceutical composition may be administered in a volume ranging from about 0.1 mL to about 2.5 mL, about 0.25 mL to about 2 mL, 0.5 mL to about 1.5 mL, or about 1 mL. The concentration of θ-defensin or the total concentration of θ-defensin species in the pharmaceutical composition is about 1 mg / mL. -1 ~about 50mgmL -1 and preferably about 12.5 mg mL -1 The concentration and / or injection volume of the theta-defensin can be adjusted to provide a dose of about 0.001 mg / kg to about 3 mg / kg, about 0.01 to about 1 mg / kg, or about 0.08 mg / kg.

[0041] In some embodiments, the pharmaceutical composition may include, in addition to one or more theta-defensins and / or theta-defensin analogs and propylene glycol, additional therapeutic compounds, such as one or more steroids with anti-inflammatory effects, nonsteroidal anti-inflammatory drugs, antibodies or antibody fragments directed against pro-inflammatory cytokines, and / or one or more analgesic compounds.

[0042] Such theta-defensin preparations can be administered using any suitable schedule. Subcutaneous injections can be provided once a week, twice a week, three times a week, every other day, daily, every 12 hours, every 8 hours, or every 6 hours, as needed to establish or maintain the desired therapeutic effect. The duration of treatment can range from about 1 week, about 2 weeks, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 6 months, about 12 months, about 18 months, about 24 months, or more than 24 months, as needed to establish or maintain the desired therapeutic effect. In some embodiments, theta-defensin dosing can be higher and / or more frequent during the initial phase of treatment to establish symptomatic remission or partial remission, and then reduced to maintain symptomatic remission, either in terms of theta-defensin dose and / or frequency of administration, or both.

[0043] Example Pharmacokinetic studies in rats Theta-defensin RTD-1 was formulated in a weakly acidic buffer containing propylene glycol as described above and administered subcutaneously to male and female Sprague-Dawley rats, after which the plasma concentrations of theta-defensin were determined. Rats were given doses ranging from 1 to 4 mg / kg. The dose volume was kept constant at 0.32 mL / kg; therefore, theta-defensin concentrations ranged from 3.125 to 12.5 mg / mL. Doses were administered three times weekly for 6 weeks. Exemplary results are shown in Table 1 (providing results for male rats) and Table 2 (providing results for female rats). Table 1 and the following tables use the following acronyms:

[0044] AUC 0-TLast Area under the plasma concentration versus time curve from time 0 (pre-dose) to the time of the last measurable concentration C Last Final plasma concentrations measured above the limit of quantification C max maximum plasma concentration MRT Last Mean residence time to the last time point at which the plasma concentration of the analyte was measured TLast Time at which the final plasma concentration above the limit of quantification was measured T max Time of maximum concentration.

[0045] [Table 1]

[0046] [Table 2]

[0047] Tables 3, 4, and 5 show typical results from a kinetic study of plasma concentrations (ng / mL) of θ-defensin in male rats given 1 mg / kg, 2 mg / kg, and 4 mg / kg, respectively. Tables 6, 7, and 8 show typical results from a similar study conducted in female rats. Figures 4A and 4B show graphs of plasma θ-defensin concentrations (ng / mL) versus time (hours) on day 1 of such a study for male and female rats, respectively. Figures 5A and 5B show graphs of plasma θ-defensin concentrations (ng / mL) versus time (hours) on day 13 of such a study for male and female rats, respectively. Figures 6A and 6B show graphs of plasma θ-defensin concentrations (ng / mL) versus time (hours) on day 41 of such a study for male and female rats, respectively.

[0048] [Table 3]

[0049] [Table 4]

[0050] [Table 5]

[0051] [Table 6]

[0052] [Table 7]

[0053] [Table 8]

[0054] Table 9 shows the C of male and female rats. max and AUC 0-TLast Typical results of dose proportionality for

[0055] [Table 9]

[0056] Figure 7A shows the C of male and female rats on day 1. max Figure 7B provides graphs showing the results of a linearity study of θ-defensin dose versus C in male and female rats at day 41. max 8A provides graphs showing the results of a linearity study of θ-defensin versus dose. FIG. 8B shows the AUC 0-TLast Figure 8B provides graphs showing the results of a linearity study of theta-defensin versus dose. Figure 8B shows the AUC 0-TLast 1 provides a graph showing the results of a linearity study versus theta-defensin dose.

[0057] Human clinical trials Theta-defensin RTD-1 was formulated in a weakly acidic buffer containing propylene glycol as described above and administered subcutaneously to male and female subjects ranging in age from 23 to 73 years, and plasma concentrations of theta-defensin were determined. Subjects received 20 μg / kg, 40 μg / kg, 80 μg / kg, 160 μg / kg, or 325 μg / kg of theta-defensin via subcutaneous injection. It should be understood that these doses are substantially lower than those used in the animal studies described above. Subjects receiving 20 to 80 μg / kg had the dose delivered at a single site. Subjects receiving larger doses had the dose distributed between two sites. Figure 9 shows typical results of measuring plasma theta-defensin concentrations (ng / mL) over time for different treatment groups. As shown, bioavailability following subcutaneous injection was dramatically increased in humans compared to rats. Similarly, improved results were shown in human subjects compared to subcutaneous injection of RTD-1 in canine and porcine animal models. max The dependence of AUC (ng / mL) on RTD-1 dose (μg / kg) is shown in Figure 10A. 0-TLast The results of a similar study of the dependence of RTD-1 dose (μg / kg) on ​​C are shown in Figure 10B. max The dependence of AUC (ng / mL) on total RTD-1 administered (mg) is shown in Figure 11A. 0-TLast The results of a similar study of the dependence of AUC on total RTD-1 (mg) administered are shown in Figure 10B. 0-TLast and C max Both were approximately linear functions of theta-defensin dose. Surprisingly, much lower doses of theta-defensin were required in humans to achieve similar plasma concentrations than those demonstrated in test animals. The inventors anticipate that similar or improved results will be observed with analogs of theta-defensin.

[0058] It should be apparent to those skilled in the art that many more modifications besides those already described are possible without departing from the inventive concepts herein. Accordingly, the inventive subject matter is not to be limited except by the spirit of the appended claims. Moreover, in interpreting both the specification and the claims, all terms should be interpreted in the broadest possible manner consistent with the context. In particular, the terms "comprises" and "comprising" should be interpreted as referring to elements, components, or steps in a non-exclusive manner, indicating that the referenced element, component, or step may be present, utilized, or combined with other elements, components, or steps not expressly mentioned. When a specification claim refers to at least one selected from the group consisting of A, B, C, ..., and N, the statement should be interpreted as requiring only one element from that group, rather than A plus N, or B plus N, etc.

[0059] [Appendix 1] A method for treating an individual having a chronic inflammatory condition, comprising providing an amount of a theta-defensin or a theta-defensin analog in a first aqueous solution comprising the theta-defensin or a theta-defensin analog and 0.5% to 1.5% v / v propylene glycol, the first aqueous solution having a pH of 5.0 to 7.0, and administering the first aqueous solution by subcutaneous injection to an individual in need of treatment, the first aqueous solution being formulated to provide an increase in the pharmacological potency or therapeutic effect of the theta-defensin or theta-defensin analog compared to a second aqueous solution comprising the amount of theta-defensin or theta-defensin analog in normal saline solution.

[0060] [Appendix 2] The method according to Appendices 1, wherein the aqueous solution further contains an acetate salt. [Appendix 3] The method described in Appendix 1 or 2, wherein the pharmacodynamic effect of the amount of theta-defensin or theta-defensin analog is increased by at least 10-fold compared to the amount of theta-defensin or theta-defensin analog provided in normal saline solution.

[0061] [Appendix 4] The method of any one of Appendices 1 to 3, wherein the pharmacodynamic effect of said amount of said θ-defensin or θ-defensin analog is increased by at least 40-fold compared to said amount of said θ-defensin or θ-defensin analog provided in normal saline solution.

[0062] [Appendix 5] The method of any one of Appendices 1 to 4, wherein the chronic inflammatory condition is selected from the group consisting of rheumatoid arthritis, inflammatory bowel disease, cancer-related inflammation, diabetes, and chronic diseases characterized by dysregulated or unresolved chronic inflammation.

[0063] [Appendix 6] The first aqueous solution is 50 mg / mL -1 6. The method of any one of claims 1 to 5, comprising the θ-defensin or θ-defensin analog at a concentration of up to 1000 mg / mL. [Appendix 7] The method described in any one of Appendices 1 to 6, wherein the θ-defensin analog is selected from the group consisting of cyclic icosipeptide, cyclic eneadecapeptide, cyclic octadecapeptide, cyclic heptadecapeptide, cyclic hexadecapeptide, cyclic pentadecapeptide, cyclic tetradecapeptide, cyclic tridecapeptide, cyclic dodecapeptide, cyclic hendecapeptide, and cyclic decapeptide.

[0064] [Appendix 8] The method according to any one of Appendices 1 to 7, wherein the first aqueous solution contains 1% v / v propylene glycol and 20 mM acetate and has a pH of 6.0. [Appendix 9] A method for sterilizing an aqueous theta-defensin preparation, comprising providing a theta-defensin or a theta-defensin analog in an aqueous buffer solution containing theta-defensin at a concentration of at least 1 mg mL-1 and 0.5% to 1.5% v / v propylene glycol, and passing the aqueous buffer solution through a filter having a pore size of 0.2 μm or less.

[0065] [Appendix 10] The method according to Appendix 9, wherein the aqueous buffer further contains acetate. [Appendix 11] The method of appendix 9 or 10, wherein the aqueous θ-defensin preparation has a pH of 5.0 to 7.0.

[0066] [Supplementary Note 12] The θ-defensin or θ-defensin analog is 50 mg / mL -1 The method according to any one of appendices 9 to 11 provided above. [Appendix 13] 50 mg mL in an aqueous solution containing 0.5% to 1.5% v / v propylene glycol -1 A pharmaceutical composition for the treatment of a chronic inflammatory condition comprising a theta-defensin or a theta-defensin analog at a concentration of up to 1000 mg / kg, said pharmaceutical composition having a pH of 5.0 to 7.0 and formulated for parenteral administration.

[0067] [Appendix 14] The pharmaceutical composition according to Appendices 13, further comprising an acetate salt. [Appendix 15] A pharmaceutical composition described in Appendix 13 or 14, wherein the concentration of theta-defensin or theta-defensin analog is selected so as to increase the pharmacological potency of theta-defensin or theta-defensin analog by at least 10-fold compared to a similar concentration of theta-defensin or theta-defensin analog provided in normal saline solution.

[0068] [Appendix 16] A pharmaceutical composition described in any one of Appendices 13 to 15, wherein the concentration of theta-defensin or theta-defensin analog is selected so as to increase the pharmacological potency of theta-defensin or theta-defensin analog by at least 40-fold compared to a similar concentration of theta-defensin or theta-defensin analog provided in normal saline solution.

[0069] [Appendix 17] The pharmaceutical composition according to any one of Appendices 13 to 16, wherein the chronic inflammatory condition is selected from the group consisting of rheumatoid arthritis, inflammatory bowel disease, cancer-related inflammation, diabetes, and chronic diseases characterized by dysregulated or unresolved chronic inflammation.

[0070] [Appendix 18] The pharmaceutical composition according to any one of Appendices 13 to 17, wherein the parenteral administration is selected from the group consisting of subcutaneous injection, intramuscular injection, and intravenous injection. [Supplementary Note 19] The θ-defensin analogue is a cyclic icosyl peptide. icosipeptide), cyclic eneadecapeptide, cyclic octadecapeptide, cyclic heptadecapeptide, cyclic hexadecapeptide, cyclic pentadecapeptide, cyclic tetradecapeptide, cyclic tridecapeptide, cyclic dodecapeptide, cyclic hendecapeptide, and cyclic decapeptide.

[0071] [Appendix 20] The pharmaceutical composition according to any one of Appendices 13 to 19, wherein the pharmaceutical composition comprises 1% v / v propylene glycol and 20 mM acetate, and has a pH of 6.0. [Appendix 21] 50 mg mL in an aqueous solution containing 0.5% to 1.5% v / v propylene glycol for the treatment of chronic inflammatory conditions. -1 A method for using a composition comprising a theta-defensin or a theta-defensin analog, wherein the composition is formulated for parenteral administration, has a pH of 5.0 to 7.0, and provides an increased pharmacodynamic effect of the theta-defensin compared to a corresponding pharmaceutical composition prepared without propylene glycol.

[0072] [Appendix 22] The method of use described in Appendix 21, wherein the composition further comprises an acetate salt. [Appendix 23] The use of Appendices 21 or 22, wherein the pharmacological potency of the theta-defensin or theta-defensin analog is increased by at least 10-fold compared to the theta-defensin or theta-defensin analog provided in normal saline solution.

[0073] [Appendix 24] The use of any one of Appendices 21 to 23, wherein the pharmacological potency of the theta-defensin or theta-defensin analog is increased by at least 40-fold compared to the theta-defensin or theta-defensin analog provided in normal saline solution.

[0074] [Appendix 25] The use of any one of Appendices 21 to 24, wherein the chronic inflammatory condition is selected from the group consisting of rheumatoid arthritis, inflammatory bowel disease, cancer-related inflammation, diabetes, and chronic diseases characterized by dysregulated or unresolved chronic inflammation.

[0075] [Appendix 26] The method of any one of Appendices 21 to 25, wherein the parenteral administration is selected from the group consisting of subcutaneous injection, intramuscular injection, or intravenous injection. [Supplementary Note 27] The θ-defensin analogue is a cyclic icosyl peptide. 27. The use of any one of claims 21 to 26, wherein the cyclic cyclohexyl 1-hydroxybenzoate is selected from the group consisting of cyclic cyclohexyl 1-hydroxybenzoate, ...

[0076] [Appendix 28] The use of any one of Appendices 21 to 27, wherein the composition contains 1% v / v propylene glycol and 20 mM acetate and has a pH of 6.0. [Appendix 29] 50 mg mL in an aqueous solution containing 0.5% to 1.5% v / v propylene glycol for the preparation of a composition useful for the treatment of chronic inflammatory conditions. -1 A method for use of a theta-defensin or a theta-defensin analog up to 100 mg / kg, wherein the composition is formulated for parenteral administration, has a pH of 5.0 to 7.0, and provides an increased pharmacodynamic effect of the theta-defensin compared to a corresponding pharmaceutical composition prepared without propylene glycol.

[0077] [Appendix 30] The use of Appendices 29, wherein the composition further comprises an acetate salt. [Appendix 31] The use of Appendices 29 or 30, wherein the pharmacological potency of the theta-defensin or theta-defensin analog is increased by at least 10-fold compared to the theta-defensin or theta-defensin analog provided in normal saline solution.

[0078] [Appendix 32] The use of any one of Appendices 29 to 31, wherein the pharmacological potency of the theta-defensin or theta-defensin analog is increased by at least 40-fold compared to the theta-defensin or theta-defensin analog provided in normal saline solution.

[0079] [Appendix 33] The use of any one of Appendices 29 to 32, wherein the chronic inflammatory condition is selected from the group consisting of rheumatoid arthritis, inflammatory bowel disease, cancer-related inflammation, diabetes, and chronic diseases characterized by dysregulated or unresolved chronic inflammation.

[0080] [Appendix 34] The method of any one of Appendices 29 to 33, wherein the parenteral administration is selected from the group consisting of subcutaneous injection, intramuscular injection, or intravenous injection. [Supplementary Note 35] The θ-defensin analogue is a cyclic icosyl peptide. icosipeptide), cyclic eneadecapeptide, cyclic octadecapeptide, cyclic heptadecapeptide, cyclic hexadecapeptide, cyclic pentadecapeptide, cyclic tetradecapeptide, cyclic tridecapeptide, cyclic dodecapeptide, cyclic hendecapeptide, and cyclic decapeptide.

[0081] [Appendix 36] The use of any one of Appendices 29 to 35, wherein the composition comprises 1% v / v propylene glycol and 20 mM acetate and has a pH of 6.0.

Claims

1. 1. Use of propylene glycol in an injectable θ-defensin formulation for treating an individual having a chronic inflammatory condition, wherein the θ-defensin formulation comprises a first concentration of a θ-defensin or a θ-defensin analog, wherein the inclusion of propylene glycol increases the pharmacological potency of the injectable θ-defensin formulation by at least 10-fold compared to a corresponding formulation provided in a normal saline solution comprising the first concentration of the θ-defensin or a θ-defensin analog, and wherein the θ-defensin or θ-defensin analog is RTD-1 (SEQ ID NO: 1).

2. The use of claim 1, wherein the pharmacological potency of the injectable θ-defensin formulation is increased by at least 40 times compared to a corresponding formulation provided in normal saline solution containing the first concentration of the θ-defensin or θ-defensin analog.

3. The use of claim 1, wherein the θ-defensin or θ-defensin analog is effective in treating a chronic inflammatory condition selected from the group consisting of rheumatoid arthritis, inflammatory bowel disease, cancer-associated inflammation, diabetes, and chronic diseases characterized by dysregulated or unresolved chronic inflammation.

4. 2. The use of claim 1, wherein propylene glycol is provided at 0.5% v / v to 1.5% v / v.

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