Compositions and methods for enhancing the effectiveness of dicarboxylic acid esters and their uses

Abstract

Compositions and methods are provided for enhancing the effectiveness of dicarboxylic acid esters, such as diethyl azelaate, in treating pain, reducing blood glucose levels, inhibiting hemolysis and / or associated pain caused by PLA2, PLD, and other phospholipases typified by PLA2, as well as other pathologies and disorders. The present invention provides methods and compositions comprising a dicarboxylic acid ester and at least one solvent effective to enhance the therapeutic effect of the dicarboxylic acid ester. In some embodiments, the composition further comprises at least one terpene. Other embodiments include methods of making the compositions and using the compositions to treat various conditions for which the dicarboxylic acid ester is useful in treating.

Description

[Technical Field]

[0001] The present disclosure relates to methods and compositions comprising a dicarboxylic acid ester and at least one solvent effective to enhance the therapeutic effect of the dicarboxylic acid ester. In some embodiments, the composition further comprises at least one terpene. Other embodiments include methods of making the compositions and using the compositions to treat various conditions for which the dicarboxylic acid ester is useful in treating. [Background technology]

[0002] Diethyl azelaate (DEA), when administered orally and topically, exhibits unexpectedly diverse and useful medical activity for the treatment of insulin resistance, musculoskeletal pain, and envenomation by phospholipases present in animal venom. See, e.g., U.S. Patent Nos. 10,251,857 B2, 11,026,912 B2, 11,918,555, and 11,911,358, the disclosures of which are incorporated herein by reference in their entireties. See also Streeper RT and Izbicka E, "Diethyl azelate for the treatment of brown recluse spider bite, a neglected orphan indication," In Vivo 36(1): 86-93, (2022).

[0003] Anesthetics and analgesics are widely used to treat pain. Recently, Pavel et al. detailed the biochemical mechanisms by which general anesthetics exert their pharmacological effects. Pavel showed that inhaled anesthetics disrupt lipid raft order and induce anesthesia through this disruption. The most studied lipid rafts are cholesterol- and sphingomyelin-rich membrane domains (e.g., monosialotetrahexosylganglioside 1, also known as GM1) that bind to cholera toxin B subunit (CTxB). GM1 lipid rafts contain phospholipase D2 (PLD2). Inhaled anesthetics increase the fluidity of GM1 lipid rafts, disrupting them and allowing PLD2 to leave the lipid rafts and relocate near TREK1. Once there, PLD2 hydrolyzes phosphatidylcholine (PC) to generate phosphatidic acid (PA) and choline. PA then binds to and activates TREK1. Activation of TREK1 induces loss of consciousness and analgesia through the resulting potassium influx. Pavel et al. also show that PLD2 translocation also activates a second channel called TRAAK, an anesthesia-insensitive homolog of TREK-1 [Pavel MA, et al., PNAS, (2020) 117(24):13757-66].

[0004] The present inventors have discovered that the dicarboxylic acid esters of the present disclosure, when combined with at least one solvent and at least one terpene, are surprisingly effective in treating a variety of disorders, such as pain, diabetes, and exposure to brown recluse spider venom and bee venom. The present inventors have discovered that the dicarboxylic acid esters, when combined with at least one solvent and at least one terpene, produce a beneficial synergistic effect.

[0005] Dimethyl sulfoxide (DMSO) is an organosulfur compound that enhances the permeation of drugs, DNA, and other substances through plasma membranes. Increased DMSO concentrations reportedly cause plasma membrane thinning, subsequent pore formation, and ultimately bilayer collapse [de Menorval MA et al., “Effects of dimethyl sulfoxide in cholesterol-containing lipid membranes: A comparative study of experiments in silico and with cells,” PLoS One Vol 7(7): e41733, (2012)]. DMSO produces rapid, moderate, and short-term relief of joint pain [Swanson B, "Medical use of dimethyl sulfoxide (DMSO)," Rev Clin Basic Pharm, Vol. 5, pp. 1-33, (1985)] and has some utility in wound healing [Capriotti K et al., "Dimethyl sulfoxide: History, chemistry, and clinical utility in dermatology," J Clin Aesthet Dermatol, Vol. 5(9): 24-26, (2012)]. DMSO applied to the skin rapidly produces a distinctive garlic taste on the tongue, indicating that DMSO is rapidly transported throughout the body through tissues [Horita A, et al., "Skin penetrating property of drugs dissolved in dimethyl sulfoxide (DMSO) and other vehicles," Live Sciences, Vol. 3, pp. 1389-1395, (1964)]. Oral use of DMSO is limited to the treatment of systemic amyloid A amyloidosis, a complication of chronic inflammatory disease.

[0006] Terpenes also enhance skin penetration by interacting with intracellular lipids in the stratum corneum, the outermost layer of skin. Terpenes can enhance the activity of both hydrophilic and lipophilic drugs, even at low concentrations [Chen J, et al., “Natural terpenes as penetration enhancers for transdermal drug delivery,” Molecules, vol. 21, pg. 1709, (2016)]. Representative terpenes, such as limonene and menthol, exhibit low cytotoxicity in vivo and improve drug bioavailability in animal models and human skin. Limonene, when applied to the skin, exhibits anti-inflammatory activity and promotes wound healing in vivo [d'Alessio PA, et al., “Skin repair properties of d-limonene and perillyl alcohol in murine models,” Antiinflamm Antiallergy Agents Med Chem, Vol. 13(1): pp. 29-35, (2014)]. Menthol stimulates cutaneous nociceptors, elicits the release of vasodilatory peptides, and can increase skin temperature.

[0007] At its most basic level, all types of pain, except psychogenic pain, involve the transmission of aversive or noxious stimuli to the central nervous system via afferent nerves. The transmission of the resulting nerve impulse must cross the cell's plasma membrane at multiple points in the signaling chain [Basbaum A, et al., Cell. (2009)139(2):267-84]. The role of the plasma membrane in pain mechanisms has been demonstrated in vivo, where disruption of nerve signaling by increasing lipid raft fluidity via cyclodextrin-mediated cholesterol depletion reversibly reduced prostaglandin E2 (PGE2)-induced hyperalgesia [Ferrari, L, et al., J. Pain (2015)16(1):60-6].

[0008] Most pain conditions in humans and animal pain models result in significant changes in mechanosensation, with mechanical forces initially affecting the plasma membrane. Our understanding of this process was advanced by the discovery of a link between mechanical stimuli and signaling from membrane-associated phospholipase D2 (PLD2). PLD2 is a mechanosensitive enzyme present in membrane lipid sites composed of cholesterol, ganglioside GM1, and the mechanically activated ion channel TREK-1, which is responsible for downstream signaling [Petersen EN, et al., “Mechanical activation of TWIK-related potassium channel by nanoscopic movement and rapid second messenger signaling,” Elife, Vol. 12 (2024)]. A common experimental method for measuring the level of skin sensitivity induced by a stimulus uses a set of graded monofilaments known as von Frey fibers [Mills C, et al., “Estimating efficacy and drug ed50's using von Frey thresholds: Impact of Weber's law and log transformation,” J Pain, Vol. 13(6), pp. 519-523, (2012)].

[0009] Other members of the phospholipase family, represented by phospholipase A2 (PLA2), are expressed in a variety of species, from bacteria to humans. PLA2 causes pain and inflammation and, in some cases, exhibits hemolytic activity. Inhibition of PLA2 is considered a therapeutic target [Yedgar S, et al., “Inhibition of phospholipase A(2) as a therapeutic target,” Biochim Biophys Acta, Vol. 1488(1-2), pp. 182-187, (2000)]. DEA and related azelaic acid esters have been reported to inhibit the PLA2 enzyme and hemolytic activity of bee and snake venoms [Streeper RT and Izbicka E, “Diethyl azelate for the treatment of brown recluse spider bite, a neglected orphan indication,” In Vivo 36(1): 86-93, (2022)].

[0010] The present inventors have also demonstrated that dicarboxylic acid esters induce multiple changes in cellular and biological signaling in therapeutically useful ways. In addition to other pharmacological activities, dicarboxylic acid esters induce the cellular release and / or production of signaling molecules with analgesic properties. Representative cellular molecules include leptin, macrophage colony-stimulating factor (M-CSF), granulocyte colony-stimulating factor (G-CSF), and granulocyte-macrophage colony-stimulating factor (GM-CSF). Furthermore, the inventors have discovered that treatment of mammalian cells and tissues with dicarboxylic acid esters can suppress the production of pain-related markers such as inducible nitric oxide synthase (iNOS) [Kwok Y, et al., PLZhang J, et al., Int. Anesthesiol. Clin. (2007) 45(2):27-37], prostaglandin E2 (PGE2) [Ahlawat A, et al., Eur. J. Pharmacol. (2018) 818:419-28], and adenosine triphosphate (ATP) [Mense S, Dtsch Arztebl Int 2008; 105(12): 214-9].

[0011] There is a need in the art to enhance the effectiveness of dicarboxylic acid esters, such as diethyl azelaate, in treating pain, reducing blood glucose levels, inhibiting hemolysis and / or associated pain caused by PLA2, PLD, and other phospholipases typified by PLA2, as well as other conditions and disorders. The embodiments described herein address and satisfy these needs, as well as other needs that will be readily discernible by those of skill in the art upon review of the embodiments described below. Summary of the Invention

[0012] An embodiment of the present disclosure is a compound represented by Formula I: R2OOC-(CH2) n -COOR1 Dicarboxylic acid ester of wherein n is 4-10, and R1 and R2 are each independently lower alkyl, at least one solvent, and at least one terpene. Certain embodiments herein relate to eliciting an analgesic effect in a subject, thereby suppressing pain resulting in part from activation of pattern recognition receptors (PRRs), such as toll-like receptors (TLRs), nucleotide multimerization domain (NOD) receptors, and / or dectin receptors.

[0013] Certain embodiments herein relate to inducing an analgesic effect by promoting the secretion of leptin, macrophage colony-stimulating factor (M-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and / or granulocyte colony-stimulating factor (G-CSF), thereby suppressing localized pain in a subject.

[0014] In some aspects, embodiments herein relate to methods and pharmaceutical compositions for eliciting an analgesic effect by inhibiting the expression and secretion of prostaglandin E2 (PGE2) and / or inducible nitric oxide synthase (iNOS) and / or adenosine triphosphate (ATP), thereby suppressing localized pain in a subject, comprising a dicarboxylic acid ester of Formula I, at least one solvent, and at least one terpene.

[0015] In some aspects, embodiments herein relate to a method of eliciting an analgesic effect, thereby suppressing localized pain in a subject, comprising administering to said subject in need of treatment a composition comprising a dicarboxylic acid ester of formula I, at least one solvent, and at least one terpene, wherein administration results in relief of symptoms associated with a pain condition.

[0016] In some aspects, embodiments herein relate to methods of treating type 2 diabetes, insulin resistance, and other disorders caused by high blood glucose levels by administering to a subject in need thereof a composition comprising a dicarboxylic acid ester of formula I, at least one solvent, and at least one terpene, wherein the administration results in a reduction in blood glucose levels.

[0017] In some aspects, embodiments herein relate to methods for treating disorders, diseases, and conditions associated with PLA2 and / or PLD having pathological, endogenous, or exogenous phospholipase activity by administering to a subject in need thereof a composition comprising a dicarboxylic acid ester of formula I, at least one solvent, and at least one terpene, wherein the administration inhibits hemolysis caused by the PLA2 or PLD.

[0018] In some aspects, embodiments herein relate to the use of a dicarboxylic acid ester of Formula I, at least one solvent, and at least one terpene in the manufacture of a medicament for inducing an analgesic effect, thereby suppressing localized pain in a subject. In some aspects, embodiments herein relate to the use of a composition comprising a dicarboxylic acid ester of Formula I, at least one solvent, and at least one terpene for reducing blood glucose levels, thereby treating type 2 diabetes, insulin resistance, and other disorders, diseases, or conditions caused by elevated blood glucose levels.

[0019] In some aspects, embodiments herein relate to the use of a composition comprising a dicarboxylic acid ester of formula I, at least one solvent, and at least one terpene in the manufacture of a medicament for inhibiting hemolysis, such as hemolysis caused by PLA2 and / or PLD, in a subject, thereby treating diseases and conditions caused by or associated with PLD and / or PLA2. [Brief explanation of the drawings]

[0020] The accompanying drawings demonstrate certain aspects of the present disclosure, which may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein. [Figure 1] Figure 1 shows the variation in analgesic effect with increasing concentrations of DMSO in combination with DEA ​​on the duration of the sensitivity response in the cutaneous mechanosensitivity assay. [Figure 2] Figures 2A-2B show the dose-response of limonene in combination with DEA ​​and DMSO as assessed by the duration of sensitivity suppression in a skin mechanosensitivity assay. Figure 2A shows the dose-response of limonene for corrected time to baseline, and Figure 2B shows the dose-response of limonene for corrected AUC at time to baseline. [Figure 3] Figures 3A-3B show the effects of several related diesters of medium-chain fatty acids and DMSO (0.5 M each), alone and in combination with 2% limonene, on the duration of sensitivity suppression in a skin mechanosensitivity assay. Figure 3A shows the effects of several related diesters of medium-chain fatty acids and DMSO (0.5 M each), alone and in combination with 2% limonene, on the time to baseline, and Figure 3B shows the effects of several related diesters of medium-chain fatty acids and DMSO (0.5 M each), alone and in combination with 2% limonene, on the AUC at time to baseline. [Figure 4] FIG. 4 shows the effects of DEA, DMSO, limonene, and menthol as single agents and in combination on the duration of sensitivity suppression in a skin mechanical sensitivity assay. [Figure 5] FIG. 5 demonstrates the superiority of orally administered DEA / DMSO / limonene 77 / 21 / 2 mixture over DEA alone on blood glucose levels in diabetic patients. [Figure 6] FIG. 6 demonstrates the superiority of DEA, DMSO, and limonene over the 77 / 21 / 2 combination over each other in inhibiting phospholipase A2-induced hemolysis. DETAILED DESCRIPTION OF THE INVENTION

[0021] All applications, publications, patents, and other references cited herein are incorporated by reference in their entirety.

[0022] As used herein, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. The term "or" refers to one or a combination of two or more of the alternatively listed elements unless the context clearly dictates otherwise. As used herein, "comprises" means "includes." Thus, "including A or B" means "including A, B, or A and B," without excluding additional elements.

[0023] Ranges may be expressed herein as from "about" one particular value and / or to "about" another particular value. As used herein, the term "about" is intended to qualify the numerical value it modifies and to describe such value as a variable within a margin of error. When a specific margin of error, such as a standard deviation for an average value in a chart or table of data, is not given, the term "about" should be understood to mean a range that includes the stated value and also includes the range that would be included if rounded up or down to the nearest significant digit, taking into account significant digits.

[0024] All numerical values ​​or ranges used herein include integers within such ranges and fractions of integers within such values ​​or ranges, unless the context clearly dictates otherwise. Furthermore, such ranges are intended to encompass the exact numbers and any subranges therebetween. The ranges may be integral or continuous, including end values. Thus, for example, a reference to a range of about 3 hours to about 10 hours includes 3 hours, 4 hours, 5 hours, etc., as well as 3 hours 1 minute, 3 hours 2 minutes, 3 hours 4 minutes, etc., 4 hours 1 minute, 4 hours 2 minutes, 4 hours 4 minutes, etc. A reference to a range of 90-100% includes 92.2%-97.5%, 91.5-94.5%, etc. A reference to a range of 1-25 days includes subranges of 1 day to 5 days, or 3 days to 7 days, or 5 days to 25 days.

[0025] As used herein, the term "comprising" is intended to mean that the pharmaceutical compositions (or compositions) and methods include the recited elements without excluding others. The term "consisting essentially of," as used with respect to compositions of the present invention, means that the composition may include additional elements as long as the additional elements do not substantially alter the composition. The term "materially altered," as used with respect to compositions, refers to an increase or decrease in the therapeutic efficacy of the composition compared to the efficacy of a composition consisting of the recited elements. In other words, when used to define a composition, "consisting essentially of" means excluding other components that are critical to the composition. Thus, a composition consisting essentially of components defined herein does not exclude trace amounts of contaminants from separation and purification methods and pharmaceutically acceptable carriers. "Consisting of" means excluding more than trace amounts of other components and substantial method steps for administering the compositions of the present invention. Embodiments defined by each of these transitional phrases are within the scope of the present invention.

[0026] As used herein, the term "optional" or "optionally" means that the subsequently described event or circumstance may or may not occur, and that the specification includes cases where the event or circumstance occurs and cases where it does not occur.

[0027] As used herein, "subject" refers to a mammal, such as a primate, in one embodiment, a human. Non-human primates include marmosets, monkeys, chimpanzees, gorillas, orangutans, and gibbons, to name a few. The term "subject" also includes cats, dogs, ferrets, chinchillas, mice, rabbits, rats, gerbils, guinea pigs, cows, horses, pigs, sheep, goats, chickens, turkeys, ducks, pheasants, pigeons, doves, parrots, cockatoos, geese, and the like.

[0028] As used herein, a "subject in need" of the methods of the present disclosure may be a subject suffering from a pain condition.

[0029] As used herein, the term "therapeutically effective" amount refers to an amount of an active ingredient sufficient to induce a localized analgesic effect in a subject. A therapeutically effective amount of a pharmaceutical composition of the present disclosure may be effective for disrupting lipid rafts. A therapeutically effective amount of a pharmaceutical composition of the present disclosure may be effective for upregulating (e.g., stimulating) the expression and / or secretion of leptin, M-CSF, G-CSF, GM-CSF, etc. from cells of a subject. A therapeutically effective amount of a pharmaceutical composition of the present disclosure may be effective for downregulating (e.g., inhibiting) the expression and / or secretion of ATP, iNOS, PGE2, etc. from cells of a subject. In other words, a "therapeutically effective" amount is an amount that results in the relief, alleviation, reduction, or stabilization of at least one clinical symptom in a subject. Those skilled in the art will understand that the therapeutic effect need not be complete or curative, as long as a benefit is provided to the subject. The effective amount may vary depending on factors such as the wound or affected area, the disease or condition being treated, the particular targeting construct being administered, the size of the subject, or the severity of the disease or condition. One of ordinary skill in the art may empirically determine the effective amount of a particular composition without necessitating undue experimentation.

[0030] As used herein, the term "active ingredient" refers to a biologically active substance. In an embodiment, a dicarboxylic acid ester of the present disclosure is the active ingredient in a pharmaceutical composition. In an embodiment, a dicarboxylic acid ester of the present disclosure is the only active ingredient in a pharmaceutical composition. In an embodiment, an azelaic acid ester of the present disclosure is the active ingredient in a pharmaceutical composition. In an embodiment, diethyl azelate is the active ingredient in a pharmaceutical composition.

[0031] As used herein, the term "pharmaceutically acceptable carrier" refers to any suitable adjuvant, carrier, excipient, or stabilizer, which may be in solid or liquid form such as a tablet, capsule, powder, solution, suspension, or emulsion, that does not cause significant irritation to the subject and does not abrogate the biological activity and properties of the active ingredient being administered.

[0032] As used herein, the term "disease" is intended to be generally synonymous with, and is used interchangeably with, the terms "disorder" and "condition" (as in a medical condition), in that both reflect an abnormal state of the human or animal body, or one of its parts, that impairs normal function, is usually manifested by characteristic signs and symptoms, and results in a decrease in the lifespan or quality of life of the human or animal.

[0033] As used herein, "treating" or "treatment" of a disease or condition can refer to preventing the disease or condition, delaying the onset or rate of progression of the disease or condition, reducing the risk of developing the disease or condition, preventing or delaying the appearance of symptoms associated with the disease or condition, reducing or terminating symptoms associated with the disease or condition, causing complete or partial regression of the disease or condition, or some combination thereof. Treatment can also refer to prophylactic or preventative treatment of a disease or condition.

[0034] As used herein, the term "inhibiting" refers to decreasing, alleviating, limiting, and / or blocking a particular action, function, interaction, or manifestation. The terms "inhibiting," "decreasing," and "reducing" are used interchangeably herein. In embodiments, the term refers to reducing or preventing the level of a particular activity, function, interaction, or manifestation in a subject (e.g., but not limited to, the level of a biomarker such as a proteinaceous or non-proteinaceous molecule in tissues and body fluids) by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or less than the amount in a corresponding control. For example, pain is reduced, alleviated, or inhibited when an indicator of pain, e.g., blood levels of prostaglandin PGE2, is reduced by at least about 10%, 20%, 30%, 50%, 80%, or 100%. In embodiments, pain is reduced, alleviated, or inhibited by at least about 1-fold, 2-fold, 3-fold, 4-fold, or more compared to the pain before administration of the dicarboxylic acid ester of the present disclosure. The measurable change can be objective (e.g., measurable by some test or marker, e.g., in an in vitro or in vivo assay or test or observation) or subjective (e.g., the subject is indicative of or feels an effect).

[0035] As used herein, the term "pain" refers to an unpleasant sensory and emotional experience associated with or described in terms of actual or potential tissue damage, and includes more or less localized sensations of discomfort, distress, or agony resulting from stimulation of specialized nerve endings. There are many types of pain, including, but not limited to, nociceptive pain, neuropathic pain, or nociceptive pain, whether acute or chronic. The purpose of inducing an analgesic effect in a subject is to reduce the degree or severity of pain perceived by the subject.

[0036] As used herein, the phrase "localized pain" refers to non-generalized pain felt in a particular location or area in a particular part of a subject's body. In embodiments, the area of ​​pain is related to the primary lesion.

[0037] As used herein, the term "envenomation" refers to contact or injection of venom into a subject caused by a bite, fangs, teeth, or needle from a venomous organism. The term "envenomation" also refers to injection from a venom delivery organ. As used herein, the term "venom" refers to any poisonous or toxic substance delivered subcutaneously, intramuscularly, or locally by a bite or needle sting or upon contact with a subject by a venomous organism, which may include various toxins, such as, but not limited to, hematoxins, cytotoxins, myotoxins, and neurotoxins. As used herein, the term "venomous carrier" refers to any animal or organism that produces, secretes, or carries venom. Examples of venomous animals include, but are not limited to, insects, reptiles, amphibians, arthropods, mollusks, cnidarians, coelenterates, or other venomous vertebrates or invertebrates. Examples of venomous organisms include, but are not limited to, bacteria or fungi.

[0038] As used herein, the term "dermonecrotic arachnidism" is generally intended to be synonymous with, and used interchangeably with, the terms "necrotic arachnidism" or "necrosis" or "dermonecrosis" or "tissue necrosis," and refers to localized skin damage and tissue injury resulting from envenomation or contact with PLD toxins.

[0039] As used herein, the term "skin ulcer" or "ulcerosis" refers to an open wound or open wound on the skin where the epidermis is absent or intact. The underlying dermis or subcutaneous tissue may be exposed and visible. The surrounding skin may be reddened and inflamed. The cardinal symptoms and signs of any type of inflammatory process are pain, redness, heat, swelling, soreness, and loss of function. Such open wounds may be susceptible to infection by pathogens such as bacteria, fungi, and viruses. In advanced cases, the wound may exude pus. Pus (dead immune cells, skin cells, subcutaneous tissue cells, cellular fluids, and infectious agents) accumulates within the cavity of the skin ulcer, forming an abscess.

[0040] "Diabetes" refers to a group of metabolic diseases characterized by high blood sugar (glucose) levels resulting from defects in insulin secretion or action, or both. "Type 2 diabetes" or "T2D" refers to one of the two major types of diabetes in which beta cells in the pancreas produce insulin but the body cannot use it effectively because, at least in the early stages of the disease, the body's cells are resistant to insulin's action. In later stages of the disease, the beta cells may stop producing insulin. Type 2 diabetes is also known as insulin-resistant diabetes, non-insulin-dependent diabetes, and adult-onset diabetes.

[0041] "Prediabetes" refers to one or more early diabetes-related conditions such as impaired glucose utilization, abnormal or impaired fasting blood glucose levels, impaired glucose tolerance, impaired insulin sensitivity, and insulin resistance.

[0042] "Insulin resistance" refers to a condition in which cells are resistant to the effects of insulin, a hormone that controls the uptake of glucose into cells, or in which the amount of insulin produced is insufficient to maintain normal blood glucose levels. Cells become less able to respond to the action of insulin, which promotes the transport of the sugar glucose from the blood to muscles and other tissues (i.e., they become less sensitive to insulin). Eventually, the pancreas produces much more insulin than normal, and the cells remain resistant. As long as enough insulin is produced to overcome this resistance, blood glucose levels remain normal. When the pancreas can no longer keep up, blood glucose levels begin to rise, resulting in diabetes. Insulin resistance ranges from normal (insulin sensitive) to insulin resistant (IR).

[0043] "Obesity" refers to a chronic disease defined by an excessive amount of body fat. Normal body fat mass (expressed as a percentage of body weight) is 25-30% for women and 18-23% for men. Women with more than 30% body fat and men with more than 25% body fat are considered obese.

[0044] The vast literature on tissue penetration enhancers largely does not directly address enhancing the efficacy of therapeutic molecules; it typically focuses solely on increasing the tissue concentration of the drug, with the general assumption that increased concentration correlates with enhanced efficacy. The present inventors unexpectedly discovered that penetration and efficacy are not linearly correlated. For example, the efficacy of DEA in combination with a solvent such as DMSO follows a bell curve with a sharp peak at approximately 21% solvent, with significantly lower efficacy below and above that concentration. If penetration were the only driver of increased efficacy, such a pronounced peak would not be observed. While not intending to be bound by any theory of action or function, the present inventors believe that for DEA / solvent / terpene combinations, enhanced efficacy occurs within a narrow, specific range of DEA-to-solvent molar ratios, and that the efficacy of preferred DEA / solvent ratios is further enhanced by preferred amounts of at least one terpene (such as limonene and menthol). The embodiments herein demonstrate that enhanced penetration does not play a significant role in enhancing DEA efficacy.

[0045] Pharmaceutical Composition In embodiments, the present disclosure provides methods and pharmaceutical compositions for inducing an analgesic effect and thereby suppressing pain in a subject, comprising administering to the subject a compound of Formula I: R2OOC-(CH2) n -COOR1 The present invention provides a method and pharmaceutical composition comprising administering a pharmaceutical composition comprising a dicarboxylic acid ester of , at least one solvent, and at least one terpene. In embodiments, n is 4 to 10, 6 to 9, or 7 to 8. In embodiments, R1 and R2 are each independently lower alkyl. As used herein, the term "lower alkyl" refers to a C1 to C6 saturated straight-chain or branched alkyl group. Examples of suitable lower alkyl groups (R1 and R2) in Formula I include methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, t-butyl, and similar groups. In embodiments, R1 and R2 are the same. In embodiments, R1 and R2 are different.

[0046] In an embodiment, the dicarboxylic acid ester is an azelaic acid ester. In an embodiment, the azelaic acid ester is diethyl azelate, diisopropyl azelate, or a mixture thereof. In an embodiment, the azelaic acid ester is diethyl azelate.

[0047] In embodiments, the dicarboxylic acid ester of Formula I can be administered in an amount sufficient to induce an analgesic effect in a subject, such as by reducing a pain response. In embodiments, the dicarboxylic acid ester of Formula I can be administered in an amount sufficient to reduce a pain response, for example, by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95%. In embodiments, the dicarboxylic acid ester of Formula I can be administered in an amount sufficient to lower blood glucose levels in a subject and treat one or more disorders, diseases, or conditions caused by elevated blood glucose levels (e.g., insulin resistance, type II diabetes). In another embodiment, the dicarboxylic acid ester of Formula I can be administered in an amount sufficient to inhibit hemolysis caused by a PLD toxin, such that administration can treat disorders, diseases, and conditions associated with the toxin.

[0048] In embodiments, the dicarboxylic acid ester of Formula I is administered in an amount sufficient to provide the above-described effects, for example, about 10% to about 90%, about 20% to about 90%, about 30% to about 90%, about 40% to about 90%, about 50% to about 90%, about 60% to about 90%, about 70% to about 90%, about 70% to about 80%, or any value or range therebetween.

[0049] The solvent used with the dicarboxylic acid ester of the active ingredient, preferably diethyl azelate, can be any solvent that can enhance the efficacy of the active ingredient. The solvent can be dimethylformamide, trifluoroacetic acid, dimethyl sulfoxide (DMSO), dihydrolevoglucosenone (CYRENE), or the like. TMIn an embodiment, the solvent is DMSO.

[0050] In embodiments, the solvent may be administered in an amount sufficient to enhance the therapeutic effect of the dicarboxylic acid ester of Formula I. In certain embodiments, the solvent, e.g., DMSO, may be administered in an amount sufficient to reduce a pain response by, for example, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95%. In embodiments, the solvent, or DMSO, may be administered in an amount sufficient to enhance the therapeutic effect of the dicarboxylic acid ester of Formula I in lowering blood glucose levels in a subject and treating one or more disorders, diseases, or conditions caused by elevated blood glucose levels (e.g., insulin resistance, type 2 diabetes). In another embodiment, a solvent, or DMSO, can be administered in an amount sufficient to enhance the therapeutic effect of the dicarboxylic acid ester of Formula I in inhibiting toxin-induced hemolysis, such that administration can treat disorders, diseases, and conditions associated with PLD toxins.

[0051] In embodiments, the solvent may be administered in an amount sufficient to enhance the efficacy of the dicarboxylic acid ester to provide the above-described effects. One or more solvents may be used in an amount of, for example, about 1% to about 80%, about 5% to about 70%, about 10% to about 50%, about 15% to about 40%, about 20% to about 30%, or any value or range therebetween.

[0052] The at least one terpene or related terpene used with the dicarboxylic acid ester of the active ingredient, preferably diethyl azelate, can be any terpene that can enhance the efficacy of the active ingredient. The at least one terpene or related terpene can be a terpenoid, where the terpene or terpenoid has a monocyclic or bicyclic structure and / or the terpene or terpenoid has 10 carbon atoms in its chemical formula. The terpene or terpenoid may be selected from the group consisting of arnica extract, arnica oil, β-aryophyllene, bisabolol, α-cedrene, carene delta, caryophyllene acetate, caryophyllene, citronellol, cyclomethylenecitronol, decyl acetate, ethyl butyrate, ethyl linalool, hexenol cis-3, kalsec pepper extract, lemon terpenes, lime terpenes, D-limonene (also called limonene), mandarin terpenes, menthol, methyl salicylate, myrcene, nonyl acetate, octyl acetate, orange terpenes, α-pinene and β-pinene, tangerine terpenes, α-terpenol, α-terpinene, valencene, and combinations thereof. In embodiments, the terpene is limonene and / or D-limonene, optionally combined with menthol.

[0053] In embodiments, at least one terpene or related terpene may be administered in an amount sufficient to enhance the efficacy of the dicarboxylic acid ester in providing the above-described effects. One or more terpenes or terpenoids may be used in an amount of, for example, about 0.1% to about 20%, about 0.25% to about 10%, or about 1% to about 5%, or about 2% to about 4%, or any value or range therebetween.

[0054] method In an embodiment, the present disclosure provides a method for inducing an analgesic effect and thereby suppressing pain in a subject, comprising administering to the subject a pharmaceutical composition comprising an azelaic acid ester, at least one solvent, and at least one terpene. In an embodiment, the present disclosure provides a method for inducing an analgesic effect and thereby suppressing pain in a subject, comprising administering to the subject the pharmaceutical composition.

[0055] The induction of localized analgesic effects can be achieved by disrupting lipid rafts or increasing their fluidity. Lipid rafts (also known as lipid microdomains) are segregated lipid domains present in the outer leaflet of the plasma membrane. Lipid rafts are enriched in cholesterol, sphingomyelin, gangliosides, and sphingolipids such as monosialotetrahexosylganglioside 1 (GM1). Lipid rafts affect membrane fluidity and membrane protein trafficking, thereby regulating neurotransmission and receptor trafficking [Korade Z, et al., Neuropharmacology (2008) 55 (8): 1265-73].

[0056] In an embodiment, the analgesic effect is induced by the disruption of lipid rafts caused by contacting the dicarboxylic acid ester of Formula I with the lipid rafts. When the dicarboxylic acid ester of Formula I is topically applied to a subject (e.g., an open wound, wound periphery, healthy skin, mucosa, oral mucosal wound), the dicarboxylic acid ester contacts and disrupts lipid rafts located in the plasma membrane of the subject. In an embodiment, the lipid rafts contain GM1.

[0057] In embodiments, lipid rafts contain phospholipase D2 (PLD2). In embodiments, the dicarboxylic acid ester of Formula I disrupts lipid rafts containing GM1. In embodiments, disruption of lipid rafts by the dicarboxylic acid ester of Formula I causes PLD2 to leave the lipid rafts, thereby activating TREK-1 and subsequently promoting the production of the signaling lipid phosphatidic acid (PA).

[0058] Pain can be subdivided in various ways depending on its type. Based on time, pain can be classified as chronic or acute. Chronic pain is recognized as pain that persists beyond normal healing time, usually for more than three months. Chronic pain affects approximately 20% of people worldwide and accounts for approximately 20% of medical consultations [Treede Z, et al., Pain (2015) 156(6):1003-7]. Based on mechanism, pain can be nociceptive, neuropathic, or nociceptive. Nociceptive pain results from stimulation of pain receptors (nociceptors) in response to tissue injury by mechanical, chemical, or thermal stimuli. Neuropathic pain results from damage to components of the nervous system. Nociceptive pain results from altered nociceptive perception.

[0059] In embodiments, the subject is suffering from pain. In embodiments, the pain is acute or chronic. In embodiments, the pain is nociceptive pain, neuropathic pain, or nociceptive pain.

[0060] In embodiments, the pain is acute. In embodiments, the subject may require treatment for acute pain. Examples of acute pain include, but are not limited to, traumatic pain, procedural pain (such as postoperative pain, dental pain, skin pain, etc.), wound pain, musculoskeletal pain (such as back pain and neck pain), toothache, infection (such as wound infection), and toxins (such as insects, animals, bacteria, fungi, etc.).

[0061] In some embodiments, pain is chronic.In some embodiments, the subject may need treatment for chronic pain.Examples of chronic pain include but are not limited to fibromyalgia, arthralgia, iliotibial band syndrome pain, tennis elbow pain, cancer pain, musculoskeletal pain (for example, back pain, neck pain), temporomandibular joint disorder, trigeminal neuralgia, chronic headache, pain associated with neurological disease (such as MS, diabetic neuropathy), neuroma, pelvic inflammatory disease, endometriosis, shingles and postherpetic neuralgia, and chronic neuropathy associated with infection (for example, HIV), chemotherapy-induced neuropathic pain, surgical neuropathic pain, traumatic neuropathic pain, vulvodynia, atypical craniofacial pain, sciatica, phantom limb, odontalgia, and burning mouth syndrome.

[0062] In embodiments, the pain is nociceptive pain. In embodiments, the subject may need treatment for nociceptive pain. Nociceptive pain is usually acute and occurs in response to certain circumstances. Examples of nociceptive pain include, but are not limited to, the pain caused by sprains, burns, bruises, surgical operations, and fractures. Chronic nociceptive pain is caused by any condition that causes pain lasting more than 6 months. Examples of chronic nociceptive pain include the pain caused by cancer, rheumatoid arthritis, osteoarthritis, and musculoskeletal diseases (e.g., back pain).

[0063] In embodiments, the pain is neuropathic pain. In embodiments, the subject may need treatment for neuropathic pain. Neuropathic pain results from damage to components of the nervous system. Certain pathologies may be the underlying cause of neuropathic pain. For example, the subject may suffer from a metabolic disease (e.g., diabetic neuropathy), an autoimmune disease (e.g., multiple sclerosis), a viral infection (e.g., shingles and its sequelae, postherpetic neuralgia), a vascular disease (e.g., stroke), trauma, and / or cancer (Campbell JN et al., Neuron (2006) 52(1):77-92; Dworkin RH et al., Arch Neurol (2003) 60; 1524-34). In embodiments, the neuropathic pain is caused by nerve damage resulting from a metabolic disease, trauma, ischemia or hemorrhage, inflammation, neurotoxicity, neurodegeneration, paraneoplastic, vitamin deficiency, or cancer. Examples of neuropathic pain include, but are not limited to, post-herpetic (or post-shingles) neuralgia, reflex sympathetic dystrophy / burning pain (neurotrauma), the pain component of cancer pain, phantom limb pain, entrapment neuropathy (e.g., carpal tunnel syndrome), and peripheral neuropathy (widespread nerve damage).

[0064] In embodiments, the pain is nociceptive pain. In embodiments, the subject may need treatment for nociceptive pain. Nociceptive pain is pain caused by altered nociception, even though there is no clear evidence of actual or threatened tissue damage that causes peripheral nociceptor activation, or clear evidence of a disease or lesion in the somatosensory system that causes pain [Chimenti RL et al., Phys Ther. (2018) 98(5): 302-314].

[0065] In embodiments, the pain is selected from acute pain, chronic pain, nociceptive pain, neuropathic pain, algesic pain, traumatic pain, chemical pain, burning pain, ischemic pain, pain from an insect bite, tingling pain, musculoskeletal pain (e.g., back pain, neck pain), rheumatoid joint pain, post-operative pain, bone pain (e.g., osteoarthritis), pain from various skin diseases (e.g., acne, psoriasis, hidradenitis suppurativa, eczema, rosacea).

[0066] In embodiments, the reduction in pain (or the reduction in pain response) results from modulation of PRR activity. In embodiments, the reduction in pain response (or the reduction in pain response) results from modulation of NOD receptor activity. In embodiments, the reduction in pain response (or the reduction in pain response) results from modulation of dectin receptor activity.

[0067] Synthesis of dicarboxylic acid esters Dicarboxylic acid esters of Formula I are commercially available or can be prepared by various methods known in the art. In embodiments, dicarboxylic acid esters can be prepared by directly forming the ester from the required acid and alcohol. This condensation can be achieved by dehydrating the reaction mixture with a suitable agent or by heating a mixture of the acid and alcohol. In embodiments, dicarboxylic acid esters can be prepared by reacting the alcohol with an activated form of the acid. Activated forms of the acid include acid halides, acid anhydrides (including both homoanhydrides and heteroanhydrides), the reaction of the internal anhydride of the parent acid with the required alcohol, and esters and anhydrides of both the acid and alcohol formed by the reaction of the required acid or alcohol with p-toluenesulfonyl chloride to form the tosyl anhydride or ester, which is then reacted with the alcohol or acid, respectively, to produce the desired final ester. Similarly, a simple organic acid anhydride, such as acetic anhydride, can be used in place of p-toluenesulfonyl chloride. Additionally, starting with one ester selected from the desired composition of matter, dissolving the ester in the desired alcohol in the presence of a suitable acidic or basic catalyst can result in the conversion of the starting ester of the acid to the ester, where the alcohol becomes the proceeding alcohol, a process also known in the art as transesterification.

[0068] For example, starting with the dimethyl ester of the acid, one can conveniently form the diethyl ester of the acid by dissolving the ester in ethanol in the presence of an acid or a base. Furthermore, if mixed esters of acids are desired, appropriately constituted solutions of two or more desired alcohols can be utilized in any of the methods described herein.

[0069] Halogenated intermediates or components can be used to form the required esters. For example, thionyl chloride chlorinates both acids and alcohols, thereby resulting in acyl chlorides and alkyl chlorides. These acyl chlorides and alkyl chlorides can then be further reacted with the desired alcohol or acid, respectively, to produce the desired ester product. Other common halogenating agents include, for example, oxalyl chloride and phosphorus chlorides and bromides, such as phosphorus pentachloride or trichloride, phosphorus pentabromide or tribromide, or phosphorus oxychloride.

[0070] Ester formation is commonly achieved by the action of a strong base on a mixture of acid and alcohol, examples of which include lithium aluminum hydride and other metal hydrides, alkali metal alkoxides such as sodium ethoxide, and diisobutylaluminum hydride.

[0071] Administration method The pharmaceutical compositions of the present disclosure can be administered symptomatically in various ways. For example, the pharmaceutical compositions can be administered topically, transdermally, intravenously, subcutaneously, intramuscularly, or orally. The pharmaceutical compositions can be applied locally and / or around the target area or area where treatment is desired.

[0072] Treatment regimen can vary depending on various factors that those skilled in the art usually consider.These factors include route of administration, nature of formulation, nature of patient's illness, subject's size, weight, surface area, age, sex, other drugs administered to patient, and the judgment of the attending physician.The pharmaceutical composition can be administered together with or in addition to other treatments for inflammatory disease or pain relief.

[0073] The pharmaceutical composition may be administered in combination with one or more additional therapeutic agents for the treatment of pain and / or inflammation. The one or more additional therapeutic agents may be administered by the same or different administration routes. The one or more additional therapeutic agents may include analgesics, anti-inflammatory agents, anesthetics, antibiotics, and antifungals.

[0074] In embodiments, the pharmaceutical composition is administered topically to the subject's skin, e.g., to an area located at or at least near the area where treatment is desired. In embodiments, the pharmaceutical composition is administered to the subject topically by rubbing it into the skin, thereby allowing the composition (or at least the dicarboxylic acid ester) to be absorbed into the skin.

[0075] In embodiments, for prophylactic treatment, the pharmaceutical composition may be administered to a subject for a period of 96 hours to just before, 72 hours to just before, 48 hours to just before, 24 hours to just before, 12 hours to just before, 8 hours to just before, 4 hours to just before, 2 hours to just before, 1 hour to just before, or 0.5 hours to just before the procedure (or surgery), or any range derivable therein, up to just before the procedure. The pharmaceutical composition may be applied topically to an area of ​​healthy skin at any time, for example, in one embodiment, before hiking.

[0076] In embodiments, for prophylactic treatment, the pharmaceutical composition is administered locally or topically to a subject prior to a procedure such as venipuncture, injection, incision, hair removal, tattooing, and tattoo removal.

[0077] In embodiments, the pharmaceutical composition is administered locally or topically to a subject during or after a procedure such as venipuncture, injection, incision, hair removal, tattooing, and tattoo removal.

[0078] In embodiments, pharmaceutical composition is applied to the subject once or multiple times.For example, pharmaceutical composition can be administered at predetermined intervals.In embodiments, for example, pharmaceutical composition can be applied once a day, twice a day, three times a day, four times a day, or five or more times a day, or once every other day, once every three days, once every four days, etc.

[0079] In embodiments, the pharmaceutical composition is administered to a subject in a therapeutically effective dose. When administered to a subject, the therapeutically effective amount will vary depending on the particular condition being treated and the desired outcome.

[0080] The pharmaceutical composition may be administered to a subject in need of treatment about every 1 hour to about 24 hours, about every 1 hour to about 12 hours, about every 2 hours to about 8 hours, about every 2 hours to about 6 hours, about every 4 hours to about 6 hours, about every 4 hours to about 8 hours, about every 12 hours, about every 24 hours, about every 48 hours, or more frequently. In embodiments, the pharmaceutical composition may be administered once, twice, three times, four times, five times, six times, seven times, eight times, or more daily, or any combination thereof. In embodiments, the pharmaceutical composition may be administered daily, every other day, every two days, every three days, every four days, or less frequently, or any combination thereof. In embodiments, the pharmaceutical composition may be administered to a subject in need of treatment for 1 to 30 days, 1 to 25 days, 1 to 20 days, 1 to 15 days, or 1 to 10 days.

[0081] formulation In embodiments, the pharmaceutical compositions of the present disclosure may be formulated for delivery via any route of administration known in the art, including, but not limited to, topical, transdermal, intravenous, subcutaneous, intramuscular, or oral administration.

[0082] Appropriate formulations vary depending on the route of administration selected. Any well-known techniques, carriers, and excipients understood in the art can be used, for example, those disclosed in Remington's Pharmaceutical Sciences, 18th Edition (Mack Publishing Company, Easton, PA, 1990), which is incorporated herein by reference. The pharmaceutical compositions disclosed herein can be manufactured by any method known in the art, such as conventional mixing, dissolving, granulating, levigating, emulsifying, encapsulating, entrapping, or compressing processes.

[0083] Pharmaceutical compositions include those suitable for topical, transdermal, intravenous, subcutaneous, intramuscular, or oral administration, although the most suitable route may vary depending, for example, on the condition and disorder of the recipient.

[0084] In embodiments, the pharmaceutical composition is suitable for topical and transdermal administration. The topical and transdermal administration of the azelaic acid ester of the present disclosure can be in the form of a processed gel, cream, lotion, solution, ointment, suspension, or emulsion. These pharmaceutical compositions may further comprise one or more suitable excipients disclosed herein.

[0085] The dicarboxylic acid ester of Formula I for parenteral administration, such as intravenous, subcutaneous, or intramuscular administration, may be in the form of a solution, suspension, or emulsion. In one embodiment, these formulations are prepared in saline. These pharmaceutical compositions may further comprise one or more suitable excipients disclosed herein.

[0086] In embodiments, the pharmaceutical composition is suitable for oral administration. The pharmaceutical composition may be conveniently presented in unit dosage form and may be prepared by any method well known in the art of pharmacy. Generally, these methods comprise the step of mixing the dicarboxylic acid ester of formula I and the optional co-administered active ingredient with a carrier that constitutes one or more accessory ingredients. Generally, the pharmaceutical composition is prepared by uniformly and intimately mixing the active ingredient with a liquid carrier or a finely divided solid carrier, or both, and then, if necessary, shaping the product into the desired composition. Pharmaceutical compositions suitable for oral administration and the optional second active ingredient may be presented as discrete units, such as capsules, cachets, or tablets, each containing a predetermined amount of the active ingredient; as powders or granules; as a solution or suspension in an aqueous or non-aqueous liquid; or as an oil-in-water or water-in-oil liquid emulsion. The active ingredient may also be presented as a bolus, electuary, or paste. These pharmaceutical compositions may further comprise one or more suitable excipients disclosed herein.

[0087] excipients In embodiments, the pharmaceutical composition of the present disclosure may further comprise one or more excipients. The excipient may comprise a carrier, such as a water-insoluble polysaccharide or oligosaccharide. Examples of carriers include, but are not limited to, cellulose acetate, cellulose acetate butyrate, cellulose acetate propionate, cellulose acetate phthalate, chitosan, β-cyclodextrin, ethyl cellulose, hydroxypropylmethylcellulose phthalate (HPMCP), microcrystalline cellulose, starch, and any combination thereof.

[0088] The excipient may include a thickening agent, such as a water-soluble polysaccharide. Examples of thickening agents include, but are not limited to, hydroxypropylmethylcellulose (HPMC), acacia, alginic acid, colloidal silicon dioxide, calcium carboxymethylcellulose, gelatin, hydroxypropyl cellulose, hydroxypropylcellulose (hypromellose), methylcellulose, sucrose, sodium alginate, sodium carboxymethylcellulose, and any combination thereof.

[0089] In embodiments, the pharmaceutical compositions of the present disclosure may further comprise one or more pharmaceutical excipients, such as ascorbic acid, EDTA dihydrate, glycerin, citric acid monohydrate, sodium citrate dihydrate, sodium benzoate, sodium propionate, 70% sorbitol solution, sucralose, FD&C Yellow #6, artificial flavors (e.g., artificial peppermint flavor, artificial fruit flavor), purified water, or any combination thereof.

[0090] In embodiments, the pharmaceutical compositions of the present disclosure may contain a preservative.Suitable preservatives include, but are not limited to, mercury-containing substances such as phenylmercuric salts (for example, phenylmercuric acetate, phenylmercuric borate, and phenylmercuric nitrate) and thimerosal; stabilized chlorine dioxide; quaternary ammonium compounds such as benzalkonium chloride, cetyltrimethylammonium bromide, and cetylpyridinium chloride; imidazolidinyl urea; parabens such as methylparaben, ethylparaben, propylparaben, and butylparaben and their salts; phenoxyethanol; chlorophenoxyethanol; phenoxypropanol; chlorobutanol; chlorocresol; phenylethyl alcohol; EDTA disodium; benzoic acid, benzyl alcohol, and sorbic acid and their salts.

[0091] In embodiments, pharmaceutical compositions of the present disclosure may contain one or more acceptable pH adjusters and / or buffers, such as acids such as acetic acid, boric acid, citric acid, lactic acid, phosphoric acid, and hydrochloric acid; bases such as sodium hydroxide, sodium phosphate, sodium borate, sodium citrate, sodium acetate, sodium lactate, and tris(hydroxymethyl)aminomethane; and buffers such as citric acid / dextrose, sodium bicarbonate, and ammonium chloride. Such acids, bases, and buffers are included in amounts necessary to maintain the pH of the composition within a pharmaceutically acceptable range. In embodiments, the pH of pharmaceutical compositions of the present disclosure may be between pH 4 and pH 7.5.

[0092] In embodiments, the pharmaceutical compositions of the present disclosure may comprise a sugar alcohol. Examples of sugar alcohols include, but are not limited to, mannitol, glycerol, galactitol, fucitol, inositol, volemitol, maltotriitol, maltoetetraitol, polyglycitol, erythritol, threitol, ribitol, arabitol, xylitol, allitol, dulcitol, glucitol, sorbitol, altritol, iditol, maltitol, lactitol, isomalt, and any combination thereof.

[0093] In embodiments, the pharmaceutical composition of the present disclosure may contain additives. Examples of additives include, but are not limited to, diluents, binders, surfactants, lubricants, glidants, coating agents, plasticizers, colorants, flavoring agents, or pharmaceutically inactive substances. Examples of diluents include, for example, cellulose; cellulose derivatives such as microcrystalline cellulose and the like; starch; starch derivatives such as corn starch, cyclodextrin, and the like; sugars; sugar alcohols such as lactose, D-mannitol, and the like; inorganic diluents such as dried aluminum hydroxide gel, precipitated calcium carbonate, magnesium aluminum metasilicate, dibasic calcium phosphate, and the like. Examples of binders include, for example, hydroxypropyl cellulose, methylcellulose, hydroxypropylmethylcellulose, povidone, dextrin, pullulan, hydroxypropyl starch, polyvinyl alcohol, scacia, agar, gelatin, tragacanth, macrogol, and the like. Examples of surfactants include, for example, sucrose esters of fatty acids, polyoxyl stearate, polyoxyethylene hydrogenated castor oil, polyoxyethylene polyoxypropylene glycol, sorbitan sesquioleate, sorbitan trioleate, sorbitan monostearate, sorbitan monopalmitate, sorbitan monolaurate, polysorbate, glyceryl monostearate, sodium lauryl sulfate, lauromacrogol, quaternary ammonium salts (for example, benzyldimethyltetradecylammonium chloride hydrate, benzethonium chloride, benzylcetyldimethylammonium chloride hydrate, benzyldimethylstearylammonium chloride hydrate, benzyldodecyldimethylammonium chloride dihydrate, benzyldodecyldimethylammonium bromide), and the like.Examples of lubricants include, for example, stearic acid, calcium stearate, magnesium stearate, talc, and the like. Examples of glidants include, for example, dry aluminum hydroxide gel, magnesium silicate, and the like.Examples of coating agents include, for example, hydroxypropyl methylcellulose 2910, aminoalkyl methacrylate copolymer E, polyvinyl acetal diethylaminoacetate, macrogol 6000, titanium oxide, and the like. Examples of plasticizers include, for example, triethyl citrate, triacetin, macrogol 6000, and the like.

[0094] kit In embodiments, the present disclosure provides Kits are also provided that include a pharmaceutical composition of the present disclosure, particularly for topical administration, wipes for cleansing the site, swabs or brushes for spreading the substance to be applied, an adhesive dressing for covering the site, and optionally, the kit may include a disposal bag or container, optionally, the kit may include absorbent material, and optionally, the kit may include a pair of gloves (e.g., sterile nitrile or latex gloves). [Example]

[0095] material and method Chemicals: Diesters of azelaic acid were synthesized from azelaic acid and ethyl alcohol using acid-catalyzed esterification as described in Izbicka E, Streeper R and Louden C: Adaptive membrane fluidity modulation: A feedback regulated homeostatic system and target for pharmacological intervention. In Vivo 35 (3073-3095, 2021), followed by fractional distillation to 99% purity as determined by gas chromatography-mass spectrometry (GC-MS). Unless otherwise stated, chemicals were provided by Sigma-Aldrich (St. Louis, MO) and Thermo Fisher Scientific (Waltham, MA). Terpenes were provided by Vigon International (East Stroudsburg, PA).

[0096] The Touch Test Sensory Evaluator Kit (North Coast Medical Inc., Morgan Hill, CA) contains 20 monofilaments that apply desired loads between 0.25 and 512 millinewtons (mN), corresponding to 0.008 and 300 g, and are numerically coded on the logarithm of the applied load (milligrams): 1.65, 2.36, 2.44, and 2.83 (normal sensation); 3.22, 3.61, and 3.84 (reduced light touch sensation); 4.08, 4.17, and 4.31 (reduced defensive sensation); and 4.56, 4.74, 4.93, 5.07, 5.18, 5.46, 5.88, 6.10, 6.45, and 6.65 (defensive sensation loss). Notably, the logarithmic transformation makes variance more uniform across the different treatment groups. See Mills C, et al., “Estimating efficacy and drug ed50's using von Frey thresholds: Impact of Weber's law and log transformation,” J Pain 13(6): 519-523, 2012.

[0097] To test sensory levels and obtain data on sensitivity loss or recovery, a cutaneous mechanosensitivity assay (CMS) using monofilaments was performed according to a modified protocol of Rolke R et al., "Quantitative sensory testing: A comprehensive protocol for clinical trials," Eur J Pain 10(1): 77-88, 2006, and Kostek M et al., "A protocol of manual tests to measure sensation and pain in humans," J Vis Exp 118, 2016. Testing was performed at approximately the same time in the morning. Subjects were placed face-down on a flat surface, and a 10-cm-radius circular test area was marked behind the left and right popliteal fossa behind the knee. To establish a baseline for the minimum load eliciting tactile sensation, the area was randomly touched with a monofilament in a series of 10 measurements (five on each side) using monofilaments of increasing load. Each touch was recorded, and subjects reported any tactile sensation they experienced. A positive score for the monofilament was defined as reporting 6 or more tactile sensations.

[0098] Solutions of test substances were prepared in isopropyl alcohol. To test DEA and DMSO at a maximum equimolar concentration of approximately 3 M each, stock solutions were prepared without diluent and adjusted according to the densities of the two compounds to 78% v / v DEA and 22% v / v DMSO. These compounds were evaluated as single reagents and in equimolar stocks at various ratios: 78, 58, 39, and 19% v / v DEA and 22, 16, 11, and 5% v / v DMSO (rounded to the nearest integer). When additional components were present, the maximum concentrations of DEA and DMSO were 77% and 21%, respectively.

[0099] Following baseline assessment, the test substance solution was applied at a volume of 0.5 ml per test area and gently rubbed in for 15 seconds (Day 1, time = 0). Mechanical sensitivity testing was then performed at the earliest time point of 5 minutes (0.1 h), followed by 1, 4, 8, 24, 48, 72, and 96 hours. The assay was terminated when tactile sensation returned to baseline, which was designated the time to baseline (TBL). Unless otherwise specified, assays were performed with 10 replicates.

[0100] The area under the curve (AUC) for the course of each test from t = 0 to recovery to baseline sensitivity was determined using SigmaPlot version 14.5 (Inpixion, Palo Alto, CA). Statistical analysis was performed using Student's t test, with a p value < 0.05 considered significant and < 0.005 considered highly significant.

[0101] Hemolysis assays of human peripheral blood using a bee venom preparation containing the active hemolytic component PLA2 were performed in triplicate as described in Streeper RT and Izbicka E, "Diethyl azelate for the treatment of brown recluse spider bite, a neglected orphan indication," In Vivo 36(1): 86-93, 2022. Briefly, a stock solution of bee venom was diluted with phosphate-buffered saline (PBS) containing 0.5% Tween (diluent) to produce the same level of hemolysis as Triton-X, used as a positive control. All test substances were also prepared in the same diluent. The reagent was preincubated at room temperature for 5 minutes, and a 30-minute reaction was initiated by the addition of a red blood cell suspension.

[0102] To assess the effects of DEA alone and a mixture of DEA, DMSO, and limonene (77%, 21%, and 2%, respectively) on blood glucose levels, the solutions were taken orally in a volume of 12.5 μl (approximately 12.5 mg). Blood glucose levels (eight replicates per treatment) were quantified at time 0 before administration and then every 15 minutes for 2 hours using a OneTouch Ultra2 blood glucose meter (LifeScan, Malvern, PA) and UniStrip1 test strips (UniStrip Technologies, Charlotte, NC) as described by the manufacturer.

[0103] Example 1 - Evaluation of DEA and DMSO in a skin mechanical sensitivity assay This experiment demonstrates that DEA and DMSO used in combination exhibit unexpected synergistic activity and a non-linear concentration dependence in the cutaneous mechanosensitivity assay (CMS).

[0104] The CMS assay (n=20) established that the baseline sensitivity without treatment was monofilament number 2.36 with a bending load of 0.02 g. DEA and DMSO were tested as single agents or as a mixture of DEA and DMSO. The results, summarized in Table 1 (where a relative molar ratio of 1 corresponds to 78% DEA and 22% DMSO), show that increasing molar ratios did not clearly correlate with increasing time to baseline (TBL). The unusual relationship can be seen by comparing the data for DEA and DMSO, particularly at molar ratios of 0.75 and above. Table 1 below shows the relationship between the relative molar concentrations (Rm) of DEA and DMSO and the time to baseline in the skin mechanosensitivity assay. [Table 1]

[0105] Table 2 presents more extensive data in a concentration matrix of DEA and DMSO and endpoints expressed as time to baseline (TBL) for mixtures of the two compounds, along with values ​​corrected for the contribution of DMSO to the mixture (TBL-C) when the effect of DEA alone is subtracted from the TBL of the mixture. Because TBL values ​​were uniformly reproducible, standard deviations are not shown. Both AUC values ​​and standard deviations are shown. A highly significant maximum effect of TBL 72 h (TBL-C 68) was observed with DEA ​​and DMSO percentage ratios of 78 / 22 and 58 / 22. Subsequent tests used fibers with 4.74, 4.03, and 5.07 mm diameters, applying bending loads of 6–10 g. The results demonstrate a significant loss of defensive sensation compared to untreated controls. Table 2 below shows the relationship between DEA and DMSO concentrations, AUC, and time to baseline in the skin mechanosensitivity assay. [Table 2]

[0106] Figure 1 highlights the unexpected properties of DMSO in combination with DEA. While less than 10% DMSO was ineffective, the analgesic effect rapidly increased to a maximum at 22% DMSO and 78% DEA, followed by a rapid decrease with decreasing DEA concentrations. These data indicate that the enhanced analgesic effect of DEA is not solely due to enhanced tissue penetration by DMSO.

[0107] Example 2 - Screening of topical analgesics, turpentine, and terpenes These experiments demonstrated unexpectedly superior activity of turpentine, limonene, pinene, and menthol in certain combinations with DEA ​​and DMSO.

[0108] A CMS assay was first conducted to select topical analgesics. Time to baseline (TBL) values ​​for 4% lidocaine and Voltaren (1.16% diclofenac) were 4 h and 8 h, respectively. Turpentine (also known as turpentine) is a component of some analgesics and was therefore also tested in the CMS screening. Mixtures of 1% and 2% turpentine with 21% DEA and 77% DMSO yielded TBL values ​​of 24 h to 48 h compared to the vehicle control.

[0109] Turpentine is a highly variable mixture of components. Products made in the United States typically contain α-pinene (75-85%), varying amounts of β-pinene (up to 3%), camphene (4-15%), limonene (5-15%), and small amounts of 3-carene and terpinolene.

[0110] To assess the relative contribution of turpentine components in CMS testing, a screen was conducted in which all compounds were tested individually at 2% v / v using a mixture of 19% DEA and 5% DMSO plus the following turpentine components and related terpenes: amyl alcohol, arnica extract, arnica oil, β-ariophyllene, bisabolol, α-cedrene, carene delta, caryophyllene acetate, caryophyllene, citronellol, cyclomethylenecitronol, decyl acetate, ethyl butyrate, ethyl linalool, hexenol cis-3, kalsec pepper extract, lemon terpenes, lime terpenes, D-limonene (also known as limonene), mandarin terpenes, menthol, methyl salicylate, myrcene, nonyl acetate, octyl acetate, orange terpenes, α-pinene and β-pinene, tangerine terpenes, α-terpenol, α-terpinene, and valencene. Additionally, cannabinoid oil (30%) and the synthetic capsaicinoid pelargonic acid vanillylamide (PAVA) (0.25% to 4%) were tested. In all cases, the TBL for the test compounds was at least 1 h, but the differences between treatment and control were not statistically significant, with the exception of menthol, α-pinene, and D-limonene. These compounds were further evaluated in Example 3 below.

[0111] Example 3 - Evaluation of multi-component mixtures in a CMS assay This example illustrates the unexpected enhancement of TBL by a specific combination of DEA, DMSO, menthol, α-pinene, and limonene. Furthermore, among azelaic, suberin, and a diester of sebacic acid in the absence or presence of limonene, only DEA ± limonene significantly inhibited CMS.

[0112] Table 3 summarizes the effects of DEA and DMSO at various concentrations in combination with limonene. The last column shows TBL values ​​corrected for the contributions of DEA and DMSO without limonene (TBL-C). Bold TBL-C values ​​indicate values ​​greater than the contributions of DEA and DMSO. Table 3 below shows the relationship between the concentrations, AUC, and time to baseline in the skin mechanosensitivity assay for DEA, DMSO, and limonene. [Table 3]

[0113] As shown in Table 3, when 2% limonene was used, the longest TBL-C of 20 h was observed for equimolar DEA and DMSO (19% and 5%, respectively). The TBL-C value decreased to 16 h for equimolar DEA and DMSO (39% and 11%, respectively). However, 2% limonene did not further enhance the combined activity of equimolar DEA and DMSO, which was observed at the highest concentrations of 77% and 21%, respectively.

[0114] It is noteworthy that 2% limonene was synergistic with low rates of DEA and DMSO. As shown in Tables 2 and 3, a 39% DEA and 11% DMSO mixture produced a TBL-C of 7 h without limonene, but a TBL-C of 16 h in the presence of 1% or 2% limonene.

[0115] The dose response of limonene was evaluated with 20% DEA and 7% DMSO (Figures 2A and 2B). A maximum TBL of 16 h, corrected for the contributions of DEA and DMSO, was observed with 4% and 2% limonene. 10% limonene had no effect, while 0.25% limonene reduced the TBL below the level of DEA plus DMSO. Limonene also exhibited a bell curve dose response.

[0116] Given the significant enhancement of TBL by 2% limonene in the DEA / DMSO mixture, other medium-chain fatty acid diesters were also tested at a concentration of 39% in 11% DMSO ± 2% limonene. As shown in Figures 3A and 3B, DEA outperformed the other diesters, and limonene significantly enhanced the effects of DEA. Similarly, limonene showed peak analgesic activity at 1–2%, but its contribution decreased with increasing concentrations and did not add any benefit above 4% (Figures 2A and 2B).

[0117] In a direct comparison of pinene isomers (Table 4), 2% α-pinene was significantly superior to β-pinene in the 39% / 11% and 19% / 5% DEA / DMSO mixtures. The activity of 2% α-pinene was comparable to that of 2% limonene in the same equimolar concentrations of DEA / DMSO (see Table 2). However, the effects of mixtures of 2% limonene and 2% α-pinene in DEA / DMSO were mediocre. Table 4 below shows the relationship between the concentrations, AUC, and time to baseline in the CMS assay for DEA, DMSO, limonene, and pinene. [Table 4]

[0118] As shown in Table 5, menthol (0.1% to 4%) in combination with 39% DEA was ineffective. A surprising enhancement of analgesic efficacy was achieved by adding 0.1% menthol to DEA / DMSO mixtures in the percentage ratios of 39 / 11 and 19 / 5. The analgesic effect of DEA / DMSO / limonene in the ratios of 39 / 11 / 2 and 19 / 5 / 2 was significantly enhanced even further by 0.1% menthol. The 48-h TBL maximum observed with DEA / DMSO / limonene / menthol (39 / 11 / 2 / 0.1) was significantly greater than that of the DEA / DMSO / limonene mixture without menthol (TBL-C of 16 h, see Table 3). Table 5 below shows the effect of menthol on the activity of mixtures of DEA, DMSO, and limonene on the duration of sensitivity suppression in the skin mechanical sensitivity assay. [Table 5]

[0119] Figure 4 summarizes the effects of 39% DEA, 11% DMSO, 2% limonene, and 0.1% menthol as single agents and in combination on the duration of susceptibility suppression in the CMS assay. Both the quaternary and ternary mixtures showed superior activity to any single agent or binary mixture.

[0120] Example 4 - Comparison of Oral DEA with a DEA, DMSO, and Limonene Mixture on Blood Glucose Levels This experiment provides a comparison of oral DEA and a mixture of DEA, DMSO, and limonene on blood glucose levels.

[0121] Figure 5 shows blood glucose levels after a single oral dose of 12.5 μl of DEA (top dashed line), corresponding to 0.17 mg / kg DEA, and 12.5 μl of a 77% DEA, 21% DMSO, and 2% limonene mixture (bottom solid line), corresponding to 0.13 mg / kg DEA. The mean difference of 39% between the two formulations in mean blood glucose levels at 2 h is highly significant (p=0.001). Oral administration of DEA has previously been shown to exert optimal blood glucose control at 1 mg / kg. The present results suggest that the 77% DEA, 21% DMSO, and 2% limonene mixture is approximately 5.8 times more potent than DEA alone. Thus, embodiments include compositions containing about 70 to about 80% DEA, about 19 to about 22% DMSO, and about 1 to about 3% limonene, or 76 to 78% DEA, 20.5 to 21.5% DMSO, and 0.5 to 1.5% limonene.

[0122] Example 5 - Evaluation of mixture composition for PLA2-induced hemolysis This example demonstrates the unusual bell-shaped dose and concentration effects of a mixture of DEA, DMSO, and limonene compared with the single agents on the extent of hemolysis induced by PLA2 in bee venom.

[0123] In Figure 6, bars 1–5 show the bell-shaped hemolysis inhibitory activity of a mixture of DEA, DMSO, and limonene. Notably, maximum inhibition was measured at a concentration 50-fold lower than that effective in CMS. At concentrations corresponding to maximum inhibition, the individual components were ineffective in this assay (bars 6–9). Furthermore, comparison of three diethyl esters (C-8; suberate, C-9; azelaate, and C-10; sebacate; all at 5%) showed that diethyl sebacate significantly inhibited hemolysis by 60%.

[0124] Although the present embodiments have been described with particular reference to preferred features, examples, and preferred embodiments, those skilled in the art will understand that various changes can be made to these preferred embodiments without departing from the spirit and scope of the present invention.

Claims

1. Formula I: R ２ OOC-(CH ２ ) ｎ -COOR １ Dicarboxylic acid ester of [wherein n is 4 to 10, and R １ and R ２ are each independently a C1-C6 saturated straight-chain or branched alkyl group; at least one solvent; and At least one terpene A pharmaceutical composition comprising:

2. 2. The pharmaceutical composition of claim 1, wherein the dicarboxylic acid ester of formula I is an azelaic acid ester selected from diethyl azelate, diisopropyl azelate, and mixtures thereof.

3. 3. The pharmaceutical composition of claim 2, wherein the azelaic acid ester is diethyl azelate.

4. 10. The pharmaceutical composition of claim 1, wherein the dicarboxylic acid ester of formula I is present in an amount of about 30% to about 90% by weight of the composition.

5. 10. The pharmaceutical composition of claim 1, wherein the at least one solvent is selected from the group consisting of dimethylformamide, dimethyl sulfoxide (DMSO), dihydrolevoglucosenone (CYRENE™), N-methylpyrrolidone (NMP), halothane, isoflurane, enflurane, desulfurane, and sevoflurane, and mixtures and combinations thereof.

6. The pharmaceutical composition of claim 5 , wherein the solvent is DMSO.

7. 10. The pharmaceutical composition of claim 1, wherein the at least one solvent is present in an amount of about 10% to about 50% by weight of the composition.

8. 2. The pharmaceutical composition of claim 1, wherein the at least one terpene is a terpene or terpenoid selected from the group consisting of arnica extract, arnica oil, beta-aryophyllene, bisabolol, alpha-cedrene, carene delta, caryophyllene acetate, caryophyllene, citronellol, cyclomethylene citronol, decyl acetate, ethyl butyrate, ethyl linalool, hexenol cis-3, kalsec pepper extract, lemon terpenes, lime terpenes, D-limonene (also called limonene), mandarin terpenes, menthol, methyl salicylate, myrcene, nonyl acetate, octyl acetate, orange terpenes, alpha-pinene and beta-pinene, tangerine terpenes, alpha-terpenol, alpha-terpinene, valencene, and mixtures and combinations thereof.

9. 9. The pharmaceutical composition of claim 8, wherein the terpene is D-limonene.

10. 10. The pharmaceutical composition of claim 1, wherein the at least one terpene is present in an amount of about 1% to about 5% by weight of the composition.

11. about 70% to about 80% diethyl azelaate; DMSO in a proportion of about 19 to about 22%; and Approximately 1% to 3% limonene A pharmaceutical composition comprising:

12. 1. A method of inducing an analgesic effect and suppressing pain in a subject, comprising administering to the subject a compound of formula I: R ２ OOC-(CH ２ ) ｎ -COOR １ Dicarboxylic acid ester of [wherein n is 4 to 10, and R １ and R ２ are each independently a C1-C6 saturated straight-chain or branched alkyl group; at least one solvent; and At least one terpene A method comprising administering a therapeutically effective amount of a composition comprising:

13. 13. The method of claim 12, wherein the dicarboxylic acid ester of formula I is diethyl azelate.

14. The method of claim 12, wherein the dicarboxylic acid ester of formula I is present in an amount of from about 30% to about 90% by weight of the composition.

15. 13. The method of claim 12, wherein the at least one solvent is DMSO.

16. The method of claim 12, wherein the at least one solvent is present in an amount of from about 10% to about 50% by weight of the composition.

17. 13. The method of claim 12, wherein the at least one terpene is limonene.

18. 13. The method of claim 12, wherein the at least one terpene is present in an amount of from about 1% to about 5% by weight of the composition.

19. 1. A method of inducing an analgesic effect and suppressing pain in a subject, comprising administering to the subject: about 70% to about 80% diethyl azelaate; DMSO in a proportion of about 19 to about 22%; and Approximately 1% to 3% limonene A method comprising administering a therapeutically effective amount of a composition comprising:

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