Compositions and methods for enhancing antimicrobial agents

Abstract

The present invention provides pharmaceutical compositions comprising an antimicrobial agent in combination with at least one cannabinoid and / or N-acylethanolamine and their use in the prevention or treatment of microbial infections and / or microbial biofilms.

Description

[0001] This application is a divisional application, the application date of the original application is July 13, 2017, the application number is 201780043787.1 (international application number PCT / IL2017 / 050800), and the name is “Compositions and methods for enhancing antimicrobial agents”. Technical Field

[0002] The present invention relates to compositions and methods for enhancing and / or minimizing the side effects of various antimicrobial agents. In particular, the present invention relates to pharmaceutical compositions comprising a combination of an antimicrobial agent, a cannabinoid, and optionally an N-acylethanolamine, and their use in treating and / or preventing microbial biofilm formation and microbially induced pathologies. Background Art

[0003] An antimicrobial is an agent that kills microorganisms or inhibits their growth. Antimicrobial drugs can be grouped according to the microorganisms they primarily act against. For example, antibiotics are used against bacteria, and antifungals are used against fungi.

[0004] Antibiotics, also called antimicrobials, are antimicrobial drugs used to treat and prevent bacterial infections. They can either kill or stop the growth of bacteria. While there are many different types of antibiotics, they all work in one of two ways. Bactericidal antibiotics (such as penicillin) kill bacteria. Bacteriostatics stop bacteria from multiplying. Because each type of antibiotic only works against certain types of bacteria or parasites, different antibiotics are used to treat different types of infections.

[0005] In the 20th century, antibiotics revolutionized medicine and effectively eradicated diseases such as tuberculosis in developed countries (together with vaccination). Their abundance and effectiveness have led to overuse and have prompted bacteria to develop resistance to some antibiotics and in some cases multi-drug resistance (MDR). This has led to a general problem of antimicrobial and antibiotic resistance, prompting the World Health Organization (WHO) to classify antimicrobial resistance as a "serious threat". In 2009, the European Center for Disease Prevention and Control (ECDC) reported that an estimated 25,000 people died of antibiotic-resistant bacterial infections in the European Union each year (ECDC / EMEA Joint Technical Report, "The bacterial challenge: time to react", published on September 17, 2009).

[0006] In addition, some antibiotics are associated with a series of adverse side effects. Depending on the antibiotics used, the target microbial organisms and individual patients, the range of side effects ranges from mild to severe. These side effects may reflect the pharmacological or toxicological properties of the antibiotics, or may involve hypersensitivity or allergies. The range of adverse reactions ranges from fever and nausea to major allergic reactions, including photosensitive dermatitis and allergies. For example, a common side effect is diarrhea caused by the destruction of the bacterial composition in the intestine, resulting in, for example, overgrowth of pathogenic bacteria such as Clostridium difficile. Antibacterial agents can also affect the vaginal flora and can cause yeast species of Candida to overgrow in the vulva-vaginal area. Interactions with other drugs may cause additional side effects, such as the administration of quinolone antibiotics and systemic corticosteroids, which may increase the risk of tendon injury. In addition, in humans and mouse models, exposure to antibiotics in the early stages of growth is associated with weight gain (Katrina Ray, Nature Reviews Endocrinology, 201, Vol. 28, p. 623).

[0007] In addition, some bacteria form pathogenic structures called biofilms. Biofilms are any group of microorganisms in which cells adhere to each other, and these cells often adhere to biological or abiotic surfaces. These adherent cells are usually embedded in the self-generated matrix of extracellular polymeric substances (EPS). The biofilm extracellular polymeric substance matrix is ​​usually a polymeric aggregate composed of extracellular DNA, proteins and polysaccharides. Microorganisms form biofilms due to many factors, which may include cell recognition, nutritional inducements of specific or nonspecific attachment sites on the surface, or in some cases by exposing cells to sub-inhibitory concentrations of antibiotics. When a cell is converted to a biofilm growth mode, it undergoes phenotypic changes in behavior, in which a large number of gene loci are differentially regulated.

[0008] Biofilms have been found to be involved in a variety of microbial infections in the body, accounting for an estimated 80% of all infections (Biel MA, Methods Mol. Biol, 2010, Vol. 635, pp. 175-194). Infectious processes in which biofilms are involved include bacterial vaginosis, urinary tract infections, catheter infections, middle ear infections, dental plaque formation, gingivitis, contact lens deposits, and more lethal processes such as endocarditis, cystic fibrosis infections, and permanent indwelling device infections such as joint prostheses and heart valves. Recently, it has been noted that bacterial biofilms can impair skin wound healing and reduce local antimicrobial efficacy in healing or treating infected skin wounds. Early detection of biofilms in wounds is critical to the successful management of chronic wounds. Although many technologies have been developed to identify bacteria in visible wounds, only a few can quickly and accurately identify bacterial biofilms.

[0009] The main problem with biofilm formation is that it allows the cells within the biofilm to be more resistant to the body's natural antimicrobials and antibiotics given in a standard manner. In fact, depending on the type of organism, antimicrobial, and experimental system, biofilm bacteria are up to a thousand times more resistant to antimicrobial stress than free-swimming bacteria of the same species. Examples of bacteria that can produce biofilms include Pseudomonas aeruginosa, Legionella, Staphylococci, Streptococci, and Candida. Streptococcus pneumoniae is the main cause of community-acquired pneumonia and meningitis in children and the elderly, and sepsis in HIV-infected patients. Legionella is known to grow in biofilms under certain conditions, where they are protected from the effects of disinfectants.

[0010] Cannabis is a genus of flowering plants belonging to the family Cannabaceae, order Rosales, which includes three different species: Cannabis sativa, Cannabis indica, and Cannabis ruderalis, which are native to Central and South Asia. Cannabis has long been used as hemp fiber, seeds, and seed oil, for medicinal purposes, and as a recreational drug. Pharmacologically, cannabis contains 483 known chemical compounds, including at least 85 different cannabinoids. Cannabinoids, terpenoids, and other compounds are secreted by glandular hairs that are present in large quantities on the calyx and bracts of the female plant.

[0011] Cannabinoids are a class of different chemical compounds that act on cannabinoid receptors on cells that inhibit the release of neurotransmitters in the brain. Cannabinoid receptors are a class of cell membrane receptors under the G protein-coupled receptor superfamily. As a typical G protein-coupled receptor, cannabinoid receptors contain seven transmembrane domains. As more and more evidence is found, there are currently two known cannabinoid receptor subtypes, called CB1 and CB2. CB1 receptors are mainly expressed in the brain (central nervous system), but are also expressed in the lungs, liver and kidneys. CB2 receptors are mainly expressed in the immune system and in hematopoietic cells.

[0012] Traditional cannabinoids are derived from their respective 2-carboxylic acids (2-COOH) by decarboxylation and catalyzed by heat, light or alkaline conditions. Phytocannabinoids (those derived from plants of the genus Cannabis) include, but are not limited to, tetrahydrocannabinol (THC), tetrahydrocannabinolic acid (THCA), cannabidiol (CBD), cannabinol (CBN), cannabigerol (CBG), cannabichromene (CBC), cannabichromene (CBL), cannabivarin (CBV), tetrahydrocannabinol (THCV), cannabichromene (CBDV), cannabichromevarin (CBCV), cannabigerovarin (CBGV) and cannabigerovarin monomethyl ether (CBGM).

[0013] The most notable cannabinoid is the phytocannabinoid Δ9-tetrahydrocannabinol (THC), which is the main psychoactive component of the cannabis plant. THC has approximately equal affinity for CB1 and CB2 receptors, and it has activity as a psychoactive agent, analgesic, muscle relaxant, antispasmodic, bronchodilator, neuroprotectant, antioxidant, and antipruritic. Dronabinol is the International Nonproprietary Name (INN) for the pure isomer of THC, (-)-trans-Δ9-tetrahydrocannabinol. Synthetic dronabinol is available as (registered trademark of Solvay Pharmaceuticals). In the United States, It is a Class III restricted drug available on prescription that is considered non-narcotic and has a low risk of physical or psychiatric dependence. It has been approved by the U.S. Food and Drug Administration (FDA) for the treatment of anorexia in AIDS patients and for refractory nausea and vomiting in patients undergoing chemotherapy. The dronabinol analog nabilone (a Schedule II restricted drug)—which has therapeutic uses as an antiemetic and as an adjunctive analgesic for neuropathic pain—is available in Canada under the trade name Commercially available. It has also been approved by the FDA and was launched in the United States in 2006.

[0014] Cannabidiol (CBD) is another major phytocannabinoid, accounting for 40% of plant extracts in selected cultivars. CBD is considered to have a wider range of medical applications than THC. An oral liquid containing CBD has been granted orphan drug status in the United States for the treatment of Dravet syndrome under the trade name Anandamide (N-arachidonoylethanolamine, AEA) - one of the main components of the endocannabinoid system - is a THC mimetic. Its effects can be central in the brain and peripheral in other parts of the body, mediated primarily by CB1 in the central nervous system and CB2 in the periphery. However, its short half-life due to the action of the enzyme fatty acid amide hydrolase (FAAH) is a disadvantage for potential therapeutic use.

[0015] N-acylethanolamines (NAEs) are lipid-derived signaling molecules formed when one of several types of acyl groups is attached to the nitrogen atom of ethanolamine.

[0016] Methods for synthesizing N-acetylethanolamine compounds are well known in the art. As described by Lambert et al. (Lambert DM, Vandevoorde S, Jonson KO, Fowler C J., Curr. Med. Chem., 2002; 9: 663-74; U.S. Patent No. 5,506,224 and U.S. Patent Application 2005 / 0054730), palmitoylethanolamide (PEA) was initially synthesized by refluxing ethanolamine with palmitic acid and producing white molten crystals at 98-99°C.

[0017] Palmitoylethanolamide (PEA, also known as N-(2-hydroxyethyl)hexadecanoamide, hydroxyethyl palmitamide, palmitol acetamide, and N-palmitoylethanolamine) is an endogenous fatty acid amide that belongs to the class of nuclear factor agonists. PEA has been shown to bind to receptors in the cell nucleus (nuclear receptors) and exert a variety of biological functions related to chronic pain and inflammation. Studies have shown that PEA interacts with different non-CB1 / CB2 receptors, indicating that PEA utilizes a unique "parallel" endocannabinoid signaling system. PEA has been shown to have anti-inflammatory, analgesic, neuroprotective, and anticonvulsant properties. PEA has also been shown to have anti-craving effects in marijuana-dependent patients, is effective in treating withdrawal symptoms, reducing marijuana use, and is effective in preventing marijuana-induced neurotoxicity and neuropsychiatric disorders.

[0018] The basic concept of the "partner effect" described by Mechoulam and co-workers in 1998 (Ben-Shabat et al., Eur. J. Pharmacol., 1998, Vol. 353(1), pp. 23-31) refers to the fact that cannabinoids and cannabis plants work together or synergistically and affect the body in a mechanism similar to the body's own endocannabinoid system.

[0019] This theory serves as the basis for a slightly controversial view in pharmacology that, in some cases, whole plant extracts are better therapeutic agents than cannabinoid extracts alone. Recently, Wagner and Ulrich-Merzenich expanded on the buddy effect theory (Wagner et al., Phytomedicine, 2009, Vol. 16 (2-3), pp. 97-110), who defined four basic mechanisms of synergistic action of whole plant extracts as: (a) the ability to affect multiple targets in the body, (b) the ability to improve the absorption of active ingredients, (c) the ability to overcome bacterial defense mechanisms, and (d) the ability to minimize adverse side effects.

[0020] PEA, in addition to being proposed to have a "partner effect" on cannabinoids such as THC, also has antimicrobial properties of its own. As early as the 1960s, the protective effect of PEA derived from egg yolk in streptococcal infection was noted (Keppel Hesselink JM, Int. J. Inflam., 2013). PEA exerts a protective effect and enhances anti-infective resistance by stimulating the phagocytosis of Escherichia coli K1 by macrophages (Redlich S, J. Neuroinflammation. June 14, 2014).

[0021] Currently, there are considerable challenges in treating infections caused by clinically relevant bacterial strains that exhibit multidrug resistance (MDR), such as methicillin-resistant Staphylococcus aureus (MRSA) and the more recent emergence of extremely drug-resistant Mycobacterium tuberculosis (XDR-TB). New classes of antimicrobial agents are therefore urgently needed, but only one new class has been introduced in the past 30 years.

[0022] There remains a need in the field of antimicrobial therapy for pharmaceutical combinations of antimicrobials with cannabinoids and other agents, particularly in humans, that can improve the efficacy of antimicrobials, reduce the minimum therapeutic dose of antimicrobials, thereby minimizing the development of resistance, reduce antimicrobial-associated side effects, prevent biofilm formation, and / or treat established biofilms. Summary of the invention

[0023] The present invention relates to compositions comprising an antimicrobial agent, a cannabinoid and optionally an N-acylethanolamine having enhanced antimicrobial therapeutic efficacy and / or reduced antimicrobial-related side effects. The present invention also relates to methods of using these compositions to treat diseases or conditions for which the antimicrobial agent is prescribed.

[0024] The present invention is based in part on the surprising experimental discovery that the combination of an antimicrobial agent and a cannabinoid enhances the antimicrobial activity of the antimicrobial agent. As exemplified below, the efficacy of the combination of an antibiotic and a cannabinoid is synergistic. It was also surprisingly found that N-acylethanolamine further enhances this effect.

[0025] In one aspect, the present invention provides a synergistic pharmaceutical composition comprising at least one antimicrobial agent, at least one cannabinoid, and a pharmaceutically acceptable carrier.

[0026] In some embodiments, the antibacterial efficacy of the pharmaceutical composition is similar to or better than the antibacterial efficacy of the same pharmaceutical composition comprising 2 to 150 times the amount of the antibacterial agent and without the cannabinoid.

[0027] In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 1000:1 and 1:1000, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 500:1 and 1:100, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 300:1 and 1:100, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 250:1 and 1:50, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 10:1 and 1:10, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 2:10 and 3:200, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 130:1 and 20:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 100:1 and 500:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 1:1 and 1:10, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 1:5 and 1:20, respectively.

[0028] In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 5:1 and 1:5, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 2:5 and 3:100, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 65:1 and 10:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is about 250:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is about 1:5, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is about 1:10, respectively.

[0029] In some embodiments, the weight ratio between gentamicin and THC is between about 5:1 and 1:5, respectively. In some embodiments, the weight ratio between gentamicin and THC is between about 2:5 and 3:100, respectively. In some embodiments, the weight ratio between gentamicin and THC is between about 65:1 and 10:1, respectively. In some embodiments, the weight ratio between ampicillin and THC is about 250:1, respectively. In some embodiments, the weight ratio between carbenicillin and THC is about 1:5, respectively. In some embodiments, the weight ratio between gentamicin and THC is about 1:10, respectively.

[0030] In some embodiments, the at least one antibacterial agent is selected from aminoglycosides, penicillins, cephalosporins, tetracyclines, macrolides, clindamycin, sulfonamides, metronidazole, quinolones, derivatives thereof, salts thereof, and any combination thereof.

[0031] In some embodiments, at least one antibacterial agent is an aminoglycoside or a salt thereof. In some embodiments, the aminoglycoside is gentamicin or a salt thereof. In some embodiments, the antibacterial efficacy of the pharmaceutical composition is similar to or better than the antibacterial efficacy of the same pharmaceutical composition comprising 2 to 64 times the amount of the antibacterial agent and without cannabinoids. In some embodiments, the antibacterial efficacy is measured against gentamicin-sensitive bacteria. In some embodiments, the gentamicin-sensitive bacteria are selected from non-resistant Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA).

[0032] In some embodiments, at least one antibacterial agent is a penicillin or a salt thereof. In some embodiments, the penicillin is ampicillin or a salt thereof. In some embodiments, the antibacterial efficacy of the pharmaceutical composition is similar to or better than the antibacterial efficacy of the same pharmaceutical composition comprising 2 to 16 times the amount of the antibacterial agent and without cannabinoids. In some embodiments, the antibacterial efficacy is measured against ampicillin-resistant bacteria. In some embodiments, the ampicillin-resistant bacteria is methicillin-resistant Staphylococcus aureus (MRSA). In some embodiments, the penicillin is carbenicillin or a salt thereof. In some embodiments, the antibacterial efficacy of the pharmaceutical composition is similar to or better than the antibacterial efficacy of the same pharmaceutical composition comprising 2 to 16 times the amount of the antibacterial agent and without cannabinoids. In some embodiments, the antibacterial efficacy is measured against Streptococcus pneumoniae.

[0033] In some embodiments, at least one cannabinoid is selected from tetrahydrocannabinol (THC), tetrahydrocannabinolic acid (THCA), cannabidiol (CBD), cannabinol (CBN), cannabigerol (CBG), cannabichromene (CBC), cannabichromene (CBL), cannabivarin (CBV), tetrahydrocannabinol (THCV), cannabichromena (CBDV), cannabichromevarin (CBCV), cannabigerovarin (CBGV) and cannabigerovarin monomethyl ether (CBGM), derivatives thereof, salts thereof, and any combination thereof.

[0034] In some embodiments, the cannabinoid is THC or a salt thereof. In some embodiments, the cannabinoid is CBD or a salt thereof. In some embodiments, the cannabinoid includes a mixture of THC or a salt thereof and CBD or a salt thereof.

[0035] In some embodiments, the pharmaceutical composition further comprises at least one N-acylethanolamine. In some embodiments, the N-acylethanolamine is selected from N-palmitoylethanolamine (PEA), Me-palmitoylethanolamine (Me-PEA), palmitoylcyclohexylamide, palmitoylbutylamide, palmitoylisopropylamide, oleoylethanolamine (OEA), palmitoylisopropylamide (PIA), derivatives thereof, salts thereof, and any combination thereof. In some embodiments, the N-acylethanolamine is PEA or a salt thereof.

[0036] In some embodiments, the antibacterial efficacy of a pharmaceutical composition and the same pharmaceutical composition without the cannabinoid is determined against the same bacteria, selected from gentamicin-sensitive Staphylococcus aureus ATCC 25923 strain, methicillin-resistant Staphylococcus aureus, and Streptococcus pneumoniae.

[0037] In some embodiments, the pharmaceutically acceptable carrier is suitable for a route of administration selected from the group consisting of oral, topical, mucosal, nasal, rectal, sublingual, parenteral, intravenous, intramuscular, and subcutaneous administration.

[0038] In another aspect, the present invention also provides a pharmaceutical composition as described above for use in treating or preventing bacterial infection or bacterial membrane.

[0039] In some embodiments, the use creates or expands the sensitivity of the bacteria to the antimicrobial agent as compared to the sensitivity of the bacteria to the antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the use is associated with a reduction in side effects as compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the side effects are selected from the group consisting of hypersensitivity to the at least one antimicrobial agent, allergic reaction to the at least one antimicrobial agent, fever, nausea, diarrhea, and any combination thereof. In some embodiments, the use is associated with an increase in antimicrobial activity as compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the use is associated with a reduction in the dosage of the at least one antimicrobial agent as compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the use is associated with a depletion of the therapeutic window of the at least one antimicrobial agent as compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid.

[0040] In some embodiments, the bacterial infection or bacterial biofilm is selected from Staphylococcus spp. infection or, Pseudomonas aeruginosa infection or biofilm, Porphyromonas spp. infection or biofilm, Moraxella spp. infection or biofilm, Peptostreptococcus spp. infection or biofilm, Enterococcus spp. infection or biofilm, Escherichia coli infection or biofilm, Klebsiella infection or biofilm, Streptococcal infection or biofilm, Treponema pallidum subspecies pallidum infection or biofilm, and Borrelia infection or biofilm.

[0041] In another aspect, the present invention also provides a method for treating or preventing a bacterial infection or bacterial biofilm in a subject in need thereof, the method comprising the step of administering to the subject a first pharmaceutical composition comprising at least one antibacterial agent and a second pharmaceutical composition comprising at least one cannabinoid.

[0042] In some embodiments, the above methods further comprise the step of administering to the subject a pharmaceutical composition comprising at least one N-acylethanolamine.

[0043] In some embodiments, the route of administration is selected from oral, topical, mucosal, nasal, rectal, sublingual, parenteral, intravenous, intramuscular, and subcutaneous administration. In some embodiments, the aminoglycoside is administered intravenously, intramuscularly, topically, orally, or in aerosolized form. In some embodiments, the penicillins are administered intravenously, intramuscularly, or orally. In some embodiments, at least one antibacterial agent is administered with at least one cannabinoid. In some embodiments, at least one antibacterial agent is administered separately from at least one cannabinoid. In some embodiments, the subject is a human.

[0044] In another aspect, the present invention also provides a kit comprising a first pharmaceutical composition comprising at least one antimicrobial agent and a second pharmaceutical composition comprising at least one cannabinoid.

[0045] In some embodiments, the above kit further comprises a third pharmaceutical composition comprising at least one N-acylethanolamine.

[0046] In some embodiments, the kits are for treating or preventing bacterial infections or bacterial biofilms.

[0047] In another aspect, the present invention also provides a method for increasing the efficacy of an antibacterial agent in a patient in need of antibacterial therapy, comprising administering to the patient at least one antibacterial agent in combination with at least one cannabinoid.

[0048] In some embodiments, the aminoglycoside is administered intravenously, intramuscularly, topically, orally, or in aerosolized form. In some embodiments, the penicillin is administered intravenously, intramuscularly, or orally. In some embodiments, the at least one antibacterial agent and the at least one cannabinoid are administered separately. In some embodiments, the at least one antibacterial agent and the at least one cannabinoid are administered together. In some embodiments, the patient is a human.

[0049] Other objects, features and advantages of the present invention will become apparent from the following description. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1 is a bar graph showing the CFU / mL concentration results for the treatments shown in Example 1.

[0051] Figure 2 TABLE 2 is a spreadsheet showing the OD value results for the treatments shown in Example 2.

[0052] Figure 3 TABLE 3 is a spreadsheet showing the OD value results for the treatments shown in Example 3.

[0053] Figure 4 TABLE 4 OD Values ​​for the Treatments ...

[0054] Figure 5 TABLE 5 OD Values ​​for the Treatments ...

[0055] Figure 6 TABLE 6 OD Values ​​for the Treatments Resulting from Example 6

[0056] Figure 7 TABLE 7 OD Values ​​for the Treatments ... DETAILED DESCRIPTION

[0057] The present invention relates to a combination comprising an antimicrobial agent and a cannabinoid. The present invention also relates to a combination comprising an antimicrobial agent, a cannabinoid and an N-acylethanolamine. The present invention also relates to methods of using these compositions in treating microbial-related infectious diseases or conditions that can be treated with an antimicrobial agent.

[0058] For example, the pharmaceutical composition or veterinary composition according to the present invention can be used to treat clinically relevant bacteria showing multidrug resistance (MDR), such as methicillin-resistant Staphylococcus aureus (MRSA) and extremely drug-resistant Mycobacterium tuberculosis XDR-TB, for which antibiotics have limited efficacy. In addition, these compositions can be used to prevent biofilm formation or help to decompose the biofilm structure that has been constructed, thereby treating the disease.

[0059] Typically, bacterial and fungal infections are treated with antimicrobial agents, which are often accompanied by various side effects that may reflect the pharmacological or toxicological properties of the antimicrobial agents. These side effects typically range from fever and nausea to major allergic reactions, including photosensitive dermatitis and anaphylaxis. Therefore, it is highly desirable to reduce the dosage of the antimicrobial agent in treatment, prolong and / or enhance its therapeutic effect, and / or reduce its associated side effects.

[0060] The pharmaceutical or veterinary compositions of the present invention provide improved medicaments compared to existing treatments, which exhibit increased therapeutic activity while minimizing the dose of antimicrobial agent administered and reducing associated adverse effects.

[0061] In one aspect, the invention provides a composition comprising at least one antimicrobial agent, at least one cannabinoid, and an acceptable carrier.

[0062] As used herein, the term "antimicrobial" is used to include substances, typically chemical substances, that kill microorganisms or retard the growth of microorganisms to a statistically significant degree. The term "antimicrobial" should be understood to include bactericides, fungicides and other such agents. The terms "antimicrobial," "bactericide," and "fungicide" are well known to those skilled in the art and their meanings will be easily discerned by the context in which each term is used.

[0063] The present invention discloses cannabinoid compounds - in the presence or absence of N-acylethanolamines - that exhibit an antimicrobial agent-sparing effect. As used herein, the term "antimicrobial agent-sparing" or "antimicrobial-sparing" refers to the ability to use a low dose of an antimicrobial agent where a moderate or high dose of an antimicrobial agent would normally be required. The cannabinoid and N-acylethanolamine compounds according to the present invention include pharmaceutically acceptable forms thereof, including isomers such as diastereomers and enantiomers, salts, solvates and polymorphs and racemic mixtures.

[0064] As used herein, the term "cannabinoid" generally refers to a class of different compounds that act on cannabinoid receptors on cells that inhibit the release of neurotransmitters in the brain. The ligands for these receptor proteins include endocannabinoids (naturally produced in the body by humans and animals), phytocannabinoids (found in cannabis and some other plants), and synthetic cannabinoids (man-made). There are at least 85 different cannabinoids isolated from cannabis, which exhibit different effects (El-Alfy et al., Pharmacology Biochemistry and Behavior, 2010, Vol. 95 (4), pp. 434-442).

[0065] In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 1000:1 and 1:1000, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 500:1 and 1:100, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 300:1 and 1:100, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 250:1 and 1:50, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 10:1 and 1:10, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 2:10 and 3:200, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 130:1 and 20:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 100:1 and 500:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 1:1 and 1:10, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 1:5 and 1:20, respectively.

[0066] In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 5:1 and 1:5, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 2:5 and 3:100, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is between about 65:1 and 10:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is about 250:1, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is about 1:5, respectively. In some embodiments, the weight ratio between the antimicrobial agent and the cannabinoid is about 1:10, respectively.

[0067] In some embodiments, the weight ratio of gentamicin to THC is between about 5:1 and 1:5, respectively. In some embodiments, the weight ratio of gentamicin to THC is between about 2:5 and 3:100, respectively. In some embodiments, the weight ratio of gentamicin to THC is between about 65:1 and 10:1, respectively. In some embodiments, the weight ratio of ampicillin to THC is about 250:1, respectively. In some embodiments, the weight ratio of carbenicillin to THC is about 1:5, respectively. In some embodiments, the weight ratio of gentamicin to THC is about 1:10, respectively.

[0068] In some embodiments, the antimicrobial efficacy of the composition is similar to or better than the antimicrobial efficacy of the same pharmaceutical composition comprising 2 to 150 times the amount of the antimicrobial agent and without the cannabinoid.

[0069] It should be understood that there are many alternative methods within the knowledge of those of ordinary skill in the art to test the antimicrobial or antibacterial efficacy of a composition, and the methods used in the Examples section are only representative methods for testing and determining the antimicrobial efficacy of a composition such as the composition of the present invention. The term "same composition" as used herein refers only to a corresponding composition identical to the original composition having only the differences described.

[0070] In some embodiments, the method for testing the antibacterial efficacy of the composition (or any test solution) comprises the following steps: (a) inoculating the bacteria (e.g., from a -80°C stock solution) into 3 mL of Muller Hinton (MH) medium; (b) culturing the inoculated medium at 37°C and 250 rpm for 18-20 hours to obtain a starting culture; (c) diluting the starting culture with saline to obtain 5×10 5 or 10 6(d) combining the starting culture and the composition to be tested in a test container (e.g., a 96-well plate); (e) culturing the test container at 37° C. with shaking (100 rpm) for 18-20 hours; (f) determining the viability of the bacteria in the test container to determine the antibacterial efficacy of the composition, for example, by determining the optical density (OD) of the bacterial culture. Routine calibration and / or modification of this method is well within the capabilities of any ordinary technician in the art.

[0071] As used herein, the term "minimum inhibitory concentration" or "MIC" relates to the lowest concentration of an agent or combination of agents, wherein the bacteria are substantially killed or at least substantially do not grow. The cutoff value for determining the MIC in the experimental part was set to an optical density (OD) value of ≤ 0.1 after incubating the bacteria overnight with the tested antimicrobial agents.

[0072] In some embodiments, the composition is a pharmaceutical composition and the carrier is a pharmaceutically acceptable carrier. In some embodiments, the composition is a veterinary composition and the carrier is a veterinarily acceptable carrier. In some embodiments, the composition is a cosmetic composition and the carrier is a cosmetically acceptable carrier.

[0073] As used herein, the term "pharmaceutical composition" refers to a preparation of an active agent described herein with other chemical components (such as physiologically suitable carriers and excipients). The purpose of a pharmaceutical composition is to facilitate administration of a compound to an organism. As used herein, the phrase "pharmaceutically acceptable carrier" refers to a carrier, excipient or diluent that does not cause significant irritation to an organism and does not destroy the biological activity and properties of the administered compound. Adjuvants are included in these phrases.

[0074] As used herein, the term "excipient" refers to an inert substance added to a pharmaceutical composition to further facilitate administration of an active ingredient. Examples of excipients include, but are not limited to, calcium carbonate, calcium phosphate, various sugars and various types of starch, cellulose derivatives, gelatin, vegetable oils, polyethylene glycols, and combinations thereof.

[0075] As used herein, the term "carrier" refers to a diluent, adjuvant, excipient or vehicle with which a compound is administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils. Water or aqueous solutions, saline solutions, aqueous glucose solutions and glycerol solutions can be used as carriers, particularly for injectable solutions. Suitable pharmaceutical carriers are described by EW Martin in "Remington's Pharmaceutical Sciences", 18th edition.

[0076] Suitable routes of administration may include, for example, oral, topical, rectal, nasal, transmucosal, enteral or parenteral delivery, including intramuscular, subcutaneous and intramedullary injections, as well as intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal or intraocular injections. Alternatively, the pharmaceutical composition may be administered in a local rather than systemic manner, for example, by injecting the pharmaceutical composition directly into a tissue area of ​​a patient.

[0077] As used herein, the term "pharmaceutically acceptable" refers to molecular entities and compositions that are physiologically tolerable and do not typically produce allergic or similar toxicity when administered to an individual. The term "pharmaceutically acceptable" may mean approved by a regulatory agency (e.g., the U.S. Food and Drug Administration) or listed in a generally recognized pharmacopoeia for use in animals (e.g., the U.S. Pharmacopeia).

[0078] The term "veterinary composition" includes compositions for internal administration and the entire range of feeds and beverages that can be consumed by animals. Typical veterinary dosage forms for internal administration are oral dosage forms, such as pastes, solutions, tablets, etc. However, injectable compositions are also contemplated. The composition of the present invention can also be a medicated feed, feed, nutrient, premix, drinking water and drinking water additive. Typically, for mixing in feed, the composition is provided as a powder, and for mixing in drinking water, the composition is provided as a fluid.

[0079] As used herein, the term "cosmetic composition" refers to a composition intended for application to a consumer's skin, particularly to the facial skin or to the facial skin around the eyes, to regulate the condition of the skin and / or improve the appearance of the skin.

[0080] The preparation technology and administration technology of medicine are well known in the art and can be found in "Remington's Pharmaceutical Sciences", Mack Publishing Co., Easton, Pa, for example. The pharmaceutical composition of the present invention can be prepared by methods well known in the art, for example, by conventional mixing, dissolving, granulating, sugar coating, grinding, emulsifying, encapsulating, embedding or lyophilizing methods. The pharmaceutical composition used according to the present invention can be prepared in a conventional manner using one or more physiologically acceptable carriers, including excipients and adjuvants, which help to process the active ingredient into a pharmaceutically usable preparation. Suitable formulations depend on the selected route of administration. For topical administration, the active ingredient of the pharmaceutical composition can be formulated into creams, ointments, solutions, patches, sprays, lotions, liniments, coatings, solid preparations such as silicone tablets, etc.

[0081] In some embodiments, the composition is non-liquid at room temperature. In some embodiments, the composition is solid or semi-solid at room temperature. In some embodiments, the composition is completely coated with an enteric coating.

[0082] In some embodiments, the antibiotic is selected from bactericidal antibiotics and bacteriostatic antibiotics. In some embodiments, the antibiotic is selected from, but not limited to, the following antibiotic classes: aminoglycosides, ansamycins, carbacephems, carbapenems, cephalosporins (first generation), cephalosporins (second generation), cephalosporins (third generation), cephalosporins (fourth generation), cephalosporins (fifth generation), glycopeptides, lincosamides (Bs), lipopeptides, macrolides (Bs), monolactams, nitrofurans, oxazolidinones (Bs), penicillins, penicillin combinations, peptide-based antibiotics, quinolones / fluoroquinolones, sulfonamides (Bs), tetracyclines (Bs), and drugs against mycobacteria.

[0083] In some embodiments, at least one antimicrobial agent is an antibacterial agent. In some embodiments, at least one antibacterial agent is selected from aminoglycosides, penicillins, cephalosporins, tetracyclines, macrolides, clindamycin, sulfonamides, metronidazole, quinolones, derivatives thereof, salts thereof, and any combination thereof. Each possibility represents a separate embodiment of the present invention.

[0084] In some embodiments, aminoglycoside antibiotics specifically list the following compounds: amikacin, gentamicin, kanamycin, neomycin, netilmycin, tobramycin, paromomycin and streptomycin. In some embodiments, ansamycin antibiotics specifically list the following compounds: geldanamycin, herbimycin and herbimycin. In some embodiments, carbacephem antibiotics specifically list the following compounds: cloracabib. In some embodiments, carbapenem antibiotics specifically list the following compounds: ertapenem, doripenem, imipenem / cilastatin, and meropenem. In some embodiments, cephalosporins (first generation) antibiotics specifically list the following compounds: cefoperazone, cefazolin, ceftriaxone, or ceftriaxone and cefalexin. In some embodiments, cephalosporins (second generation) antibiotics specifically list the following compounds: cefaclor, cefmandole, cefoxitin, cefuroxime and cefuroxime. In some embodiments, cephalosporins (third generation) antibiotics specifically list the following compounds: cefixime, cefdinir, cefditoren, cefoperazone, cefotaxime, cefpodoxime, ceftazidime, cefbutan, ceftizoxime and ceftriaxone. In some embodiments, cephalosporins (fourth generation) antibiotics specifically list the following compounds: cefopizine. In some embodiments, cephalosporins (fifth generation) antibiotics specifically list the following compounds: ceftaroline fosamil and ceftobiprole. In some embodiments, glycopeptide antibiotics specifically list the following compounds: teicoplanin, vancomycin, telavancin, dalbavancin and oritavancin. In some embodiments, lincosamide (Bs) antibiotics specifically list the following compounds: clindamycin and lincomycin. In some embodiments, lipopeptide antibiotics specifically list the following compounds: daptomycin. In some embodiments, macrolide antibiotics (Bs) specifically list the following compounds: azithromycin, clarithromycin, dirithromycin, erythromycin, roxithromycin, troleandomycin, telithromycin and spiramycin. In some embodiments, monolactam antibiotics specifically list the following compounds: aztreonam. In some embodiments, nitrofuran antibiotics specifically list the following compounds: furazolidone and nitrofurantoin (Bs). In some embodiments, oxazolidinone antibiotics (Bs) specifically list the following compounds: linezolid, posizolid, radezolid and torezolid. In some embodiments, penicillin antibiotics and combinations thereof specifically list the following compounds: amoxicillin, ampicillin, azlocillin, carbenicillin, cloxacillin, dicloxacillin, dicloxacillin, sulfaquinoxaline, methicillin, naphthamide penicillin, oxacillin, penicillin G, penicillin V, piperacillin, temocillin and ticarcillin.In some embodiments, polypeptide antibiotics specifically list the following compounds: bacitracin, colistin and polymyxin B. In some embodiments, quinolone / fluoroquinolone antibiotics specifically list the following compounds: ciprofloxacin, enoxacin, gatifloxacin, gemifloxacin, levofloxacin, lomefioxacin, moxifloxacin, nalidixic acid, norfloxacin, ofloxacin, trovafloxacin, grefloxacin, sparfloxacin and temafloxacin. In some embodiments, sulfonamide (Bs) antibiotics specifically list the following compounds: sulfamethoxazole, sulfacetamide, sulfadiazine, sulfadiazine, sulfadimethoxine, sulfamethoxazole, sulfamethoxazole, sulfa (archaic), sulfasalazine, sulfisoxazole, trimethoprim sulfamethoxazole and 2,4-diaminoazobenzene-4-sulfonamide (Sulfonamidochrysoidine) (archaic). In some embodiments, tetracycline (Bs) antibiotics specifically list the following compounds: demethylchlorotetracycline, deoxyoxytetracycline, minocycline, oxytetracycline and tetracycline. In some embodiments, antibiotics against mycobacteria specifically include the following compounds: clofazimine, dapsone, ciprofloxacin, cycloserine, ethambutol (Bs), ethionamide, isoniazid, pyrazinamide, rifamycin, rifabutin, rifapentine and streptomycin. In some embodiments, unclassified antibiotics specifically include the following compounds: arsphenamine, chloramphenicol (Bs), fosfomycin, fusidic acid, metronidazole, mupirocin, plate mycin, quinupristin / dalfopristin, methyl sulfone chloramphenicol, tigecycline (Bs), metronidazole and trimethoprim (Bs).

[0085] As used herein, the term "antibiotic" or "antibacterial agent" generally refers to any compound that kills, prevents or slows bacterial progression. These terms also include a class of different compounds that target bacterial cell walls (penicillins and cephalosporins) or cell membranes (polymyxins), or interfere with necessary bacterial enzymes (rifamycins, tiacyclines, quinolones and sulfonamides), have bactericidal activity, or eliminate protein synthesis (macrolides, lincosamides and tetracyclines), known as "bacterial inhibitors" (except bactericidal aminoglycosides). Further classification is based on its target specificity. "Narrow spectrum" antibacterial antibiotics target specific types of bacteria (e.g., Gram-negative or Gram-positive bacteria), while "broad-spectrum" antibiotics affect a variety of bacteria.

[0086] In some embodiments, at least one antimicrobial agent is an aminoglycoside or a salt thereof. In some embodiments, the aminoglycoside is gentamicin or a salt thereof. In some embodiments, the antimicrobial efficacy of the pharmaceutical composition is similar to or better than the antimicrobial efficacy of the same pharmaceutical composition comprising 2, 4, 8, 16, 32 or 64 times the amount of the antimicrobial agent and without cannabinoids. In some embodiments, the antimicrobial efficacy is measured against bacteria that are sensitive to gentamicin. In some embodiments, the bacteria that are sensitive to gentamicin are selected from non-resistant Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA). Each possibility represents a separate embodiment of the present invention.

[0087] In some embodiments, at least one antimicrobial agent is a penicillin or a salt thereof. In some embodiments, the penicillin is ampicillin or a salt thereof. In some embodiments, the antimicrobial efficacy of the pharmaceutical composition is similar to or better than the antimicrobial efficacy of the same pharmaceutical composition comprising 2, 4, 8 or 16 times the amount of the antimicrobial agent and without cannabinoids. In some embodiments, the antimicrobial efficacy is measured for ampicillin-resistant bacteria. In some embodiments, the ampicillin-resistant bacteria is methicillin-resistant Staphylococcus aureus (MRSA). In some embodiments, the penicillin is carbenicillin or a salt thereof. In some embodiments, the antimicrobial efficacy of the pharmaceutical composition is similar to or better than the antimicrobial efficacy of the same pharmaceutical composition comprising 2 or 4 times the amount of the antimicrobial agent and without cannabinoids. In some embodiments, the antimicrobial efficacy is measured for Streptococcus pneumoniae.

[0088] In some embodiments, the at least one cannabinoid is selected from tetrahydrocannabinol (THC), tetrahydrocannabinolic acid (THCA), cannabidiol (CBD), cannabinol (CBN), cannabigerol (CBG), cannabichromene (CBC), cannabichromene (CBL), cannabivarin (CBV), tetrahydrocannabinol (THCV), cannabidiol (CBDV), cannabichromevarin (CBCV), cannabigerovarin (CBGV), cannabigerovarin monomethyl ether (CBGM), derivatives thereof, salts thereof, and any combination thereof. Each possibility represents a separate embodiment of the present invention.

[0089] In some embodiments, the cannabinoid is THC or a salt thereof. In some embodiments, the cannabinoid is CBD or a salt thereof. In some embodiments, the cannabinoid comprises a mixture of THC or a salt thereof and CBD or a salt thereof.

[0090] In some embodiments, the pharmaceutical composition further comprises at least one N-acylethanolamine. In some embodiments, the N-acylethanolamine is selected from: N-palmitoylethanolamine (PEA), Me-palmitoylethanolamine (Me-PEA), palmitoylcyclohexylamide, palmitoylbutylamide, palmitoylisopropylamide, oleoylethanolamine (OEA), palmitoylisopropylamide (PIA), derivatives thereof, salts thereof, and any combination thereof. Each possibility represents a separate embodiment of the present invention. In some embodiments, the N-acylethanolamine is PEA or a salt thereof.

[0091] As used herein, the term "N-acylethanolamine" generally refers to a class of fatty acid amides - lipid-derived signaling molecules - formed when one of several types of acyl groups is attached to the nitrogen atom of ethanolamine. These amides can conceptually be formed by the release of a molecule of water from fatty acids and ethanolamine, but the known biosynthesis uses a specific phospholipase D to cleave the phospholipid unit from the N-acylphosphatidylethanolamine. The suffixes -amine and -amide in these names each refer to a single nitrogen atom of the ethanolamine that links the compound together: it is called "amine" in ethanolamine because it is considered to be the free terminal nitrogen in that subunit, and the nitrogen atom is called "amide" when the nitrogen atom is considered to be associated with the adjacent carbonyl group of the acyl subunit. In this application, the names of these compounds may be "amide" or "amine". The term "ethanolamine" is used in its usual sense and is intended to include monoethanolamine, diethanolamine, triethanolamine and mixtures thereof.

[0092] As used herein, the term "derivative" refers to a compound whose core structure is the same as or very similar to the core structure of the N-acylethanolamine compound, with chemical or physical modifications (eg, different or additional side groups).

[0093] As used herein, the term "salt" refers to any form of the active ingredient, wherein the active ingredient is in ionic form and coupled to a counterion (cation or anion) or in solution. This also includes complexes of the active ingredient with other molecules and ions, particularly complexes that are complexed by ionic interactions.

[0094] In some embodiments, the N-acylethanolamine includes a side chain with a length of 16 carbon atoms, called palmitoylethanolamine. In other embodiments, the N-acylethanolamine is selected from: caprylethanolamine (C10:0), laurylethanolamine (C12:0) and myristoylethanolamine (C14:0). Each possibility represents a separate embodiment of the present invention.

[0095] In some embodiments, N-acylethanolamine is a derivative of N-15 acylethanolamine (reference WO 2010 / 013240). In some embodiments, N-acylethanolamine is a derivative of N-palmitoylethanolamine. In some embodiments, the ethanolamide group of palmitoylethanolamine is substituted by a portion selected from butanamide, isopropylamide, cyclohexylamide and (2-methyl)ethanolamine in the derivative. Each possibility represents a separate embodiment of the present invention.

[0096] In some embodiments, the N-acylethanolamine is selected from: N-palmitoylethanolamine (PEA), Me-palmitoylethanolamine (Me-PEA), palmitoylcyclohexylamide, palmitoylbutylamide, palmitoylisopropylamide, oleoylethanolamine (OEA), palmitoylisopropylamide (PIA), derivatives thereof and salts thereof. Each possibility represents a separate embodiment of the present invention. In some embodiments, the N-acylethanolamine is PEA.

[0097] In some embodiments, the N-acylethanolamine is oleoylethanolamine (OEA) or a derivative thereof. In some embodiments, the ethanolamide group of OEA is replaced in the derivative by a moiety selected from butanamide, isopropylamide, cyclohexylamide and (2-methyl)ethanolamine. Each possibility represents a separate embodiment of the present invention. OEA, its derivatives and synthetic methods are well known in the art and are particularly described in U.S. Patent Nos. 6,656,972 and 7,348,338 and U.S. Patent Application Publication No. 2002 / 0173550.

[0098] In some embodiments, the N-acylethanolamine is selected from: N-palmitoylethanolamine (PEA), Me-palmitoylethanolamine (Me-PEA), palmitoylcyclohexylamide, palmitoylbutylamide, palmitoylisopropylamide, oleoylethanolamine (OEA), palmitoylisopropylamide (PIA), derivatives thereof, salts thereof, and any combination thereof. In some embodiments, the N-acylethanolamine is a salt of PEA.

[0099] In some embodiments, the antimicrobial efficacy of the pharmaceutical composition and the same pharmaceutical composition without cannabinoids is determined against the same bacteria, wherein the bacteria are selected from gentamicin-resistant Staphylococcus aureus ATCC 25923 strain, methicillin-resistant Staphylococcus aureus and Streptococcus pneumoniae. Each possibility represents a separate embodiment of the present invention.

[0100] In some embodiments, the pharmaceutically acceptable carrier is suitable for a route of administration selected from the group consisting of oral, topical, mucosal, nasal, rectal, sublingual, parenteral, intravenous, intramuscular, and subcutaneous administration. Each possibility represents a separate embodiment of the present invention. In some embodiments, the pharmaceutically acceptable carrier is suitable for intravenous administration. In some embodiments, the pharmaceutically acceptable carrier is suitable for intramuscular administration. In some embodiments, the pharmaceutically acceptable carrier is suitable for oral administration.

[0101] In another aspect, the present invention also provides a pharmaceutical composition as described above for use in treating or preventing infectious diseases that can be treated by antimicrobial agents.

[0102] The phrase "capable of being treated with an antimicrobial agent" refers to any infectious disease or condition that is susceptible to treatment with any antimicrobial agent, can be treated with any antimicrobial agent and / or is known to be treatable with any antimicrobial agent.

[0103] As used herein, the term "treat" includes, but is not limited to, any one or more of the following: eliminating, ameliorating, inhibiting, attenuating, blocking, suppressing, reducing, delaying, stopping, alleviating or preventing one or more symptoms or side effects of a disease or disorder.

[0104] The term "acute" refers to a disorder that has a relatively short, severe course.

[0105] As used herein, the term "chronic" means that the duration of the disease or condition of the present invention may be weeks, months or possibly years. The intensity of the disease or condition may be differentiated according to various conditions such as patient age, body temperature, season, disease type, etc.

[0106] In some embodiments, the disease is infection. In some embodiments, the microbial infection is in the form of a biofilm. In some embodiments, the infection is caused by bacteria, mycobacteria or fungi. In some embodiments, the microbial infection is a bacterial infection. In some embodiments, the infection is primary or opportunistic. In some embodiments, the infection can be classified by the anatomical location or organ system of the infection, including: urinary tract infection, skin infection, respiratory tract infection, odontogenic infection, vaginal infection and intra-amniotic infection. In addition, the inflammatory site where infection is the most common cause includes pneumonia, meningitis and salpingitis.

[0107] In some embodiments, the condition is a side effect associated with the use of antibiotics. In some embodiments, depending on the antibiotics used, the target microorganisms, and the individual patients, the side effects associated with the use of antibiotics range from mild to very severe. These side effects may reflect the pharmacological or toxicological properties of the antibiotics or may involve hypersensitivity or allergies. The range of adverse effects ranges from fever and nausea to major allergic reactions, including photosensitive dermatitis and allergies. Common side effects include diarrhea due to the destruction of the bacterial composition in the intestine, leading to, for example, overgrowth of pathogenic bacteria such as Clostridium difficile. Antibacterial agents can also affect the vaginal flora and can cause yeast species of the genus Candida to overgrow in the vulva-vaginal area. Additional side effects may be caused by interactions with other drugs, such as the administration of quinolone antibiotics and systemic corticosteroids, which increase the risk of tendon injury.

[0108] In another aspect, the present invention also provides a pharmaceutical composition as described above, for use in treating or preventing microbial infection or microbial biofilm.

[0109] As used herein, the term "biofilm" generally refers to a structure formed by any group of microorganisms on a biological or abiotic surface, in which cells adhere to each other and often adhere to the surface. These adherent cells are usually embedded in a self-generated matrix of an extracellular polymeric substance (EPS). The microbial cells grown in a biofilm are physiologically different from the cells of the same organism, on the contrary, they are single cells that can float or swim in a liquid culture medium. Infection processes involving biofilms include common problems such as bacterial vaginosis, urinary tract infections, catheter infections, middle ear infections, dental plaque formation, gingivitis, contact lens precipitation, and less common but more deadly diseases such as endocarditis, infections in cystic fibrosis, and infections such as permanent indwelling devices (such as joint prostheses and heart valves).

[0110] In some embodiments, the microbial infection or microbial biofilm is a bacterial infection or bacterial biofilm.

[0111] In some embodiments, the typical symptoms of bacterial infection are local redness, fever, edema, swelling and pain. One of the signs of bacterial infection is local pain, i.e. pain in a specific part of the body. For example, if an incision occurs and is infected by bacteria, pain is produced at the site of infection. Bacterial sore throats are usually characterized by more pain on one side of the throat. If pain occurs only in one ear, the ear infection is more likely to be diagnosed as bacterial. An incision that produces pus and milky white fluid is likely to be infected.

[0112] In some embodiments, the use creates or expands the sensitivity of a microorganism to an antimicrobial agent compared to the sensitivity of the microorganism to the antimicrobial agent in the absence of at least one cannabinoid. In some embodiments, the use creates or expands the sensitivity of a microorganism to an antimicrobial agent compared to the sensitivity of the microorganism to the antimicrobial agent in the absence of at least one cannabinoid and the absence of at least one N-acylethanolamine.

[0113] In some embodiments, the use is associated with a reduction in side effects compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the use is associated with a reduction in side effects compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid and the at least one N-acylethanolamine.

[0114] In some embodiments, the side effect is selected from the group consisting of hypersensitivity reaction to at least one antimicrobial agent, allergic reaction to at least one antimicrobial agent, fever, nausea, diarrhea, and any combination thereof. Each possibility represents a separate embodiment of the present invention.

[0115] In some embodiments, the use is associated with an increase in antimicrobial activity compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the use is associated with an increase in antimicrobial activity compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid and the at least one N-acylethanolamine.

[0116] In some embodiments, the use is associated with a reduced dose of the at least one antimicrobial agent compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the use is associated with a reduced dose of the at least one antimicrobial agent compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid and the at least one N-acylethanolamine.

[0117] In some embodiments, the use is associated with a depletion of the therapeutic window of the at least one antimicrobial agent compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid. In some embodiments, the use is associated with a depletion of the therapeutic window of the at least one antimicrobial agent compared to the use of the at least one antimicrobial agent in the absence of the at least one cannabinoid and the at least one N-acylethanolamine.

[0118] As used herein, the phrase "cannabinoids and cannabinoid-like compounds enhance the efficacy of antimicrobial agents" means that the antimicrobial effect of the antimicrobial agent is significantly improved when a cannabinoid is administered with or without N-acylethanolamine, compared to the therapeutic effect of the antimicrobial agent when the cannabinoid and / or N-acylethanolamine is not administered.

[0119] As used herein, the phrase "cannabinoids and cannabinoid-like compounds reduce the required dose of an antimicrobial agent" means that when a cannabinoid is administered with or without N-acetylethanolamine, a significantly lower dose of the antimicrobial agent is required to achieve the same antimicrobial effect of the antimicrobial agent than the dose of the antimicrobial agent required to achieve a certain antimicrobial effect when the antimicrobial agent is administered in the absence of the cannabinoid and / or N-acetylethanolamine.

[0120] As used herein, the phrase "cannabinoids and cannabinoid-like compounds reduce at least one side effect of an antimicrobial agent" means that the incidence or severity of at least one side effect of an antimicrobial agent is significantly lower when the antimicrobial agent is administered and a cannabinoid is administered with or without N-acylethanolamine, compared to the severity of the same side effect when the antimicrobial agent is administered in the absence of cannabinoids and / or N-acylethanolamine.

[0121] As used herein, the phrase "cannabinoids and cannabinoid-like compounds extend the therapeutic window of antimicrobial agents" means that when a cannabinoid is administered with or without N-acylethanolamine, the antimicrobial agent has an antimicrobial effect for a significantly longer period of time than when the cannabinoid and / or N-acylethanolamine is not administered, the antimicrobial agent has an antimicrobial effect for a significantly longer period of time.

[0122] In some embodiments, the microbial infection or microbial biofilm is selected from Staphylococcus spp. infection or biofilm (including Staphylococcus aureus infection or biofilm and S. epidermidis infection or biofilm), Pseudomonas aeruginosa infection or biofilm, Porphyromonas spp. infection or biofilm (including P. gingivalis infection or biofilm), Moraxella spp. infection or biofilm, Peptostreptococcus spp. infection or biofilm, Enterococcus spp. infection or biofilm, Escherichia coli infection or biofilm. coli infection or biofilm, Klebsiella infection or biofilm, Streptococcus infection or biofilm, Treponema pallidum subspecies spirochete infection or biofilm, and Borrelia infection or biofilm. Each possibility represents a separate embodiment of the present invention.

[0123] In another aspect, the present invention also provides a method for treating or preventing a microbial infection or a microbial biofilm in a subject in need thereof, the method comprising the step of administering to the subject a combination of a first pharmaceutical composition comprising at least one antimicrobial agent and a second pharmaceutical composition comprising at least one cannabinoid, wherein the antimicrobial efficacy of the combination is similar to or better than the antimicrobial efficacy of the same first pharmaceutical composition comprising 2 to 150 times the amount of antimicrobial agent in the absence of the second pharmaceutical composition comprising at least one cannabinoid, thereby treating or preventing the microbial infection or microbial biofilm.

[0124] In some embodiments, the method further comprises the step of administering to the subject a pharmaceutical composition comprising at least one N-acylethanolamine.

[0125] In some embodiments, the route of administration is selected from the group consisting of oral, topical, mucosal, nasal, rectal, sublingual, parenteral, intravenous, intramuscular, and subcutaneous administration. In some embodiments, the aminoglycoside is administered intravenously, intramuscularly, topically, orally, or in aerosolized form. In some embodiments, the penicillin is administered intravenously, intramuscularly, or orally. Each possibility represents a separate embodiment of the present invention.

[0126] In some embodiments, the subject is a human.

[0127] In some embodiments, the amount of the antimicrobial agent and the cannabinoid is sufficient to achieve a concentration of at least 0.0005 mg / mL gentamicin and at least 0.0001 mg / mL THC in the subject. In some embodiments, the amount of the antimicrobial agent and the cannabinoid is sufficient to achieve a concentration of at least 0.0001 mg / mL gentamicin and at least 0.0005 mg / mL THC in the subject.

[0128] In some embodiments, the amount of antimicrobial agent and cannabinoid is sufficient to achieve a concentration of at least 0.0002 mg / mL gentamicin and at least 0.0005 mg / mL THC in the subject. In some embodiments, the amount of antimicrobial agent and cannabinoid is sufficient to achieve a concentration of at least 0.00003 mg / mL gentamicin and at least 0.001 mg / mL THC in the subject.

[0129] In some embodiments, the amount of antimicrobial agent and cannabinoid is sufficient to achieve a concentration of at least 0.0313 mg / mL gentamicin and at least 0.0005 mg / mL THC in the subject. In some embodiments, the amount of antimicrobial agent and cannabinoid is sufficient to achieve a concentration of at least 0.0078 mg / mL gentamicin and at least 0.001 mg / mL THC in the subject.

[0130] In some embodiments, the amount of antimicrobial agent and cannabinoid is sufficient to achieve a concentration of at least 0.25 mg / mL ampicillin and at least 0.001 mg / mL THC in the subject.

[0131] In some embodiments, the amounts of the antimicrobial agent and the cannabinoid are sufficient to achieve a concentration of at least 0.0001 mg / mL carbenicillin and at least 0.0005 mg / mL THC in the subject.

[0132] In some embodiments, the amounts of the antimicrobial agent and the cannabinoid are sufficient to achieve a concentration of at least 0.0002 mg / mL gentamicin and at least 0.002 mg / mL THC in the subject.

[0133] On the other hand, the present invention also relates to a kit, the kit comprising

[0134] (a) a first pharmaceutical composition comprising at least one antimicrobial agent, and

[0135] (b) a second pharmaceutical composition comprising at least one cannabinoid.

[0136] In some embodiments, the kit further comprises a third pharmaceutical composition comprising at least one N-acylethanolamine.

[0137] In some embodiments, at least one antimicrobial agent and at least one cannabinoid are non-natural. In some embodiments, at least one antimicrobial agent, at least one cannabinoid, and at least one N-acylethanolamine are non-natural. In some embodiments, the kit is for treating or preventing microbial infection or biofilm formation.

[0138] In another aspect, the present invention also provides a pharmaceutical composition comprising at least one antimicrobial agent, a pharmaceutical composition comprising at least one cannabinoid, and a pharmaceutical composition comprising at least one N-acylethanolamine for use in treating or preventing microbial infection or biofilm formation.

[0139] In another aspect, the present invention also provides a method for treating or preventing an infectious condition or disease treatable by an antimicrobial agent in a subject in need thereof, the method comprising the step of administering to the subject a pharmaceutical composition comprising at least one antimicrobial agent and a pharmaceutical composition comprising at least one cannabinoid, thereby treating or preventing the infectious condition or disease.

[0140] As used herein, the terms "multidrug resistance (MDR) and "antimicrobial resistance (AMR)" generally refer to the evolution of a bacterial organism to become more or completely resistant to antimicrobial agents that were previously able to treat it. The term also includes antibiotic resistance, which applies to both bacteria and antibiotics. Resistance generally arises in one of three ways: natural resistance in some types of bacteria; genetic mutation; or through one species acquiring resistance from another. Resistance can arise spontaneously due to random mutations; or more commonly, gradually over time, and due to the misuse of antibiotics or antimicrobials.

[0141] As used herein, the term "topical" refers to applying a composition according to the present invention directly to at least a portion / area of ​​a subject's skin (human or non-human skin) to achieve a desired effect, for example, to treat a skin disorder as described herein.

[0142] For injection, the active ingredients of the pharmaceutical composition can be formulated in an aqueous solution, preferably in a physiologically compatible buffer such as Hank's solution, Ringer's solution or physiological saline buffer. For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.

[0143] The term "mucosal administration" relates to delivering the composition to a mucosal membrane, for example the oral or labial mucosa or a respiratory mucosa such as the nasal mucosa.

[0144] For oral administration, pharmaceutical compositions can be easily formulated by combining the active compound with a pharmaceutically acceptable carrier known in the art. Such carriers allow pharmaceutical compositions to be formulated into tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions, etc. for oral ingestion by patients. Pharmacological preparations for oral use can be prepared using solid excipients, optionally grinding the resulting mixture after adding suitable adjuvants as needed, and processing the granular mixture to obtain tablets or dragee cores. In particular, suitable excipients are fillers such as sugars, including lactose, sucrose, mannitol or sorbitol; cellulose preparations such as corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth gum, methylcellulose, hydroxypropyl methylcellulose and sodium carboxymethylcellulose; and / or physiologically acceptable polymers such as polyvinyl pyrrolidone (PVP). If necessary, disintegrants such as cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof, such as sodium alginate, may be added.

[0145] The term "oral administration" refers to any method of administration in which an active agent can be administered by swallowing, chewing, sucking or drinking an oral dosage form. Examples of solid dosage forms include conventional tablets, multilayer tablets, capsules, caplets, etc. that do not substantially release the drug in the mouth or oral cavity.

[0146] Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbomer gel, polyethylene glycol, titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures. Dyes or pigments may be added to the tablets or dragee coatings for identification or characterization of different combinations of active compound dosages.

[0147] Orally usable pharmaceutical compositions include hard or soft sealed capsules made of gelatin and plasticizers such as glycerol or sorbitol. Capsules can contain active ingredients mixed with fillers such as lactose, adhesives such as starch, lubricants such as talc or magnesium stearate and optional stabilizers. In soft capsules, the active ingredient can be dissolved or suspended in a suitable liquid, for example in fatty oil, liquid paraffin or liquid polyethylene glycol. In addition, stabilizers can be added. All preparations for oral administration should have a dosage suitable for the selected route of administration.

[0148] For buccal and sublingual administration, the compositions may take the form of tablets or lozenges formulated in conventional manner or in an adhesive carrier.

[0149] The pharmaceutical compositions described herein can be formulated for parenteral administration, for example by single injection or continuous infusion. The preparations for injection can be present in unit dose form, for example in ampoules or in multidose containers with optionally added preservatives. The composition can be a suspension, solution or emulsion in an oily or aqueous vehicle and can contain preparatants such as suspending agents, stabilizers and / or dispersants.

[0150] Pharmaceutical compositions for parenteral administration include aqueous solutions of active agents in water-soluble form. In addition, the suspension of active ingredient can be prepared as suitable oily or water-based injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters such as ethyl oleate, triglycerides or liposomes. Aqueous injection suspensions can contain materials that increase the viscosity of the suspension, such as sodium carboxymethylcellulose, sorbitol or dextran. Optionally, the suspension can also contain suitable stabilizers or agents that increase the solubility of the active ingredient to allow the preparation of highly concentrated solutions.

[0151] Alternatively, the active ingredient may be in powder form for constitution with a suitable vehicle, eg, a sterile, pyrogen-free water based solution, before use.

[0152] The compositions of the present invention can also be delivered using an in-situ formed depot (ISFD). Examples of in-situ formed depots include semisolid polymers that can be injected as a melt and form a depot when cooled to body temperature. Requirements for such ISFDs include a low melting point or glass transition temperature in the range of 25-658°C and an intrinsic viscosity in the range of 0.05-0.8dl / g. Below the viscosity threshold of 0.05dl / g, delayed diffusion cannot be observed, and above 0.8dl / g, the ISFD no longer uses a needle for injection. At temperatures above 378°C but below 658°C, these polymers behave like viscous fluids that solidify into high-viscosity depots. By incorporating the drug into the molten polymer without applying a solvent. Thermoplastic pastes (TPs) can be used to generate subcutaneous drug reservoirs from which they diffuse into the systemic circulation.

[0153] In situ cross-linked polymer systems utilize cross-linked polymer networks to control the diffusion of macromolecules over extended periods of time. The use of in situ cross-linked implants requires protection of the bioactive agent during the cross-linking reaction. This can be achieved by encapsulation into rapidly degradable gelatin microparticles.

[0154] ISFD can also be based on polymer precipitation. A water-insoluble and biodegradable polymer is dissolved in a biocompatible organic solvent, to which the drug is added and mixed to form a solution or suspension. When the formulation is injected into the body, the water-miscible organic solvent dissipates and water penetrates into the organic phase. This results in phase separation and precipitation of the polymer to form a reservoir at the injection site. An example of such a system is ATRIGELE TM (ARTIX Laboratories).

[0155] Thermally induced gelling systems can also be used as ISFDs. Many polymers show a sudden change in solubility with ambient temperature. The prototype of a thermosensitive polymer is poly(N-isopropylacrylamide), poly-NIPAAM, which exhibits a rather sharp lower critical solution temperature.

[0156] Thermoplastic pastes such as the new generation of poly(orthoesters) developed by AP Pharma can also be used for reservoir drug delivery. The pastes include polymers that are semisolid at room temperature, so heating is no longer required for drug incorporation and injection. Injection can be performed through a needle no larger than 22 gauge. The drug can be mixed into the system in a dry and therefore stable state. Shrinkage or swelling upon injection is believed to be unimportant, so the initial drug burst is expected to be lower than other types of ISFD. Autocatalytic degradation by surface erosion provides additional advantages.

[0157] The compositions of the present invention may also be delivered by medical devices, such as orthopedic implants, contact lenses, microneedle arrays, patches, and the like.

[0158] Sustained release (SR), extended release (ER, XR or XL), timed release or timed release, controlled release (CR) or continuous release (CR or Contin) pills are tablets or capsules formulated to dissolve slowly and release the drug over time. Sustained release tablets are formulated so that the active ingredient is embedded in a matrix of insoluble substances (e.g. acrylics, polysaccharides, etc.) so that the dissolved drug diffuses out through the pores in the matrix. In some SR formulations, the matrix physically swells to form a gel so that the drug first dissolves in the matrix and then leaves through the outer surface. The difference between controlled release and sustained release is that controlled release is a complete zero-order release, i.e. the drug is released over time regardless of concentration. On the other hand, sustained release means that the drug is released slowly over a period of time. It may or may not be a controlled release.

[0159] Pharmaceutical compositions suitable for use in the context of the present invention include compositions containing an effective amount of active ingredients to achieve the intended purpose. More specifically, a "therapeutically effective amount" refers to an amount of active ingredient that is effective to prevent, alleviate or ameliorate the symptoms or side effects of a disease or condition, or to prolong the survival of the subject being treated. Determining a therapeutically effective amount is well within the capabilities of those skilled in the art, especially based on the detailed disclosure provided herein.

[0160] For any formulation used in the methods of the invention, the dosage or therapeutically effective amount can be estimated initially from in vitro and cell culture assays. For example, the dosage can be formulated in an animal model to achieve a desired concentration or titer. This information can be used to more accurately determine useful dosages in humans.

[0161] The dosage of each compound of the claimed combination depends on several factors, including: the method of administration, the disease to be treated, the severity of the disease, whether the disease is to be treated or prevented, and the age, weight and health of the person to be treated. In addition, pharmacogenetic (the effect of genotype on the pharmacokinetic, pharmacodynamic or efficacy characteristics of a therapeutic agent) information about a particular patient may affect the dosage used.

[0162] Continuous daily dosing may not be required; the treatment regimen may require cycles during which no doses are given, or treatment may be provided as needed during acute exacerbations.

[0163] Dose escalation may or may not be required; treatment regimens may require reductions in drug doses.

[0164] The toxicity and therapeutic efficacy of the active ingredients described herein can be determined in vitro, in cell culture or in experimental animals by standard pharmacological methods. These data obtained from in vitro, cell culture analysis and animal studies can be used to formulate a range of doses for human use. The dosage can vary depending on the dosage form used and the route of administration utilized. The exact formulation, route of administration and dosage can be selected by an individual physician based on the patient's condition. (See, e.g., Fingl, E. et al., 1975, "The Pharmacological Basis of Therapeutics," Chapter 1, page 1.)

[0165] Depending on the severity and responsiveness of the condition to be treated, administration may be single or multiple administrations, with the course of treatment continuing from several days to several weeks, or until a cure or diminution of the disease state is achieved.

[0166] If desired, the composition of the present invention can be presented in a package or dispenser device, such as a kit approved by the FDA, which can contain one or more unit dosage forms containing active ingredients. The package can, for example, include metal or plastic foil, such as a blister package. The package or dispenser device can be accompanied by instructions for administration. The package or dispenser device can also be accompanied by a notice in the form prescribed by a government agency that manages the manufacture, use or sale of medicines, which reflects the form of the composition approved by the agency for human or veterinary administration. For example, the notice can include a label approved by the U.S. Food and Drug Administration (FDA) for prescription drugs or approved product inserts. The composition of the preparation of the present invention prepared in a pharmaceutically acceptable carrier can also be prepared, placed in a suitable container, and marked for the treatment of a specified inflammatory disorder, as further described above.

[0167] In some embodiments, the cannabinoid increases the effectiveness of the antibiotic compared to the same pharmaceutical composition without the antibiotic. In some embodiments, the cannabinoid reduces the required dose of the antibiotic compared to the same pharmaceutical composition without the cannabinoid. In some embodiments, the cannabinoid reduces at least one side effect of the antibiotic compared to the same pharmaceutical composition without the cannabinoid. In some embodiments, the cannabinoid extends the therapeutic window of the antibiotic compared to the same pharmaceutical composition without the cannabinoid. In some embodiments, the composition of cannabinoid and N-acylethanolamine increases the effectiveness of the antibiotic compared to the same pharmaceutical composition without the cannabinoid and N-acylethanolamine. In some embodiments, the composition of cannabinoid and N-acylethanolamine reduces the required dose of the antibiotic compared to the same pharmaceutical composition without the cannabinoid and N-acylethanolamine. In some embodiments, the composition of cannabinoid and N-acylethanolamine reduces at least one side effect of the antibiotic compared to the same pharmaceutical composition without the cannabinoid and N-acylethanolamine. In some embodiments, the composition of cannabinoid and N-acylethanolamine extends the therapeutic window of the antibiotic compared to the pharmaceutical composition without the cannabinoid and N-acylethanolamine.

[0168] In some embodiments, the route of administration is independently selected for each drug selected from oral, topical, mucosal, nasal, rectal, sublingual, parenteral, intravenous, intramuscular and subcutaneous administration. Each possibility represents a separate embodiment of the present invention.

[0169] In some embodiments of the above methods, the antibiotic and the cannabinoid are contained in the same pharmaceutical composition. In some embodiments of the above methods, the antibiotic, the cannabinoid, and the N-acylethanolamine are contained in the same pharmaceutical composition.

[0170] On the other hand, the present invention also provides a dosage unit, comprising or consisting of any one of the above-mentioned pharmaceutical compositions.

[0171] In some embodiments, the dosage unit comprises 10-500 μg of a cannabinoid. In some embodiments, the dosage unit comprises 0.5-50 mg of a cannabinoid. In some embodiments, the cannabinoid is THC. In some embodiments, the dosage unit is formulated for a route of administration selected from inhalation, topical, mucosal, nasal, oral, rectal, sublingual, parenteral, intravenous, intramuscular, and subcutaneous administration.

[0172] In another aspect, the present invention further provides a method for increasing the efficacy of an antibacterial agent in a patient in need of antibacterial therapy, comprising administering to the patient at least one antibacterial agent in combination with at least one cannabinoid.

[0173] The following examples are provided to more fully illustrate some embodiments of the present invention. However, they should not be interpreted as limiting the broad scope of the present invention in any way. Without departing from the scope of the present invention, those skilled in the art can easily design many variations and modifications of the principles disclosed herein.

[0174] Example

[0175] Example 1. PEA synergistically reduces the effective concentration of THC.

[0176] The goal of this experiment was to determine the minimum bactericidal concentration (MBC) of THC in the presence and absence of PEA on S. aureus ATCC 25923 strain (gentamicin-sensitive).

[0177] plan:

[0178] 1. Cultivate Staphylococcus aureus (from -80°C stock) in 3 ml MullerHinton (MH, Difco) medium at 37°C with stirring at 250 rpm for 18-20 hours (this culture was used as the starter for the experiment);

[0179] 2. Dilute the starter culture with saline to obtain 5 × 10 5 and 10 6 CFU / mL of working culture of bacteria.

[0180] 3. THC (0.125 mg / mL in saline) was diluted in a 96-well plate with or without PEA or gentamicin.

[0181] 4. Incubate the plate at 37°C with shaking (100 rpm) for 18-20 hours.

[0182] 5. On the following day, assess the CFU / mL of bacteria suspended in saline in the wells to determine the MBC.

[0183] Results: The MBC value of THC alone was found to be approximately 0.0078 mg / ml, which was 32 times lower than the MBC value of gentamicin alone (0.25 mg / ml), while the MBC value of THC combined with PEA was found to be approximately 0.0019 mg / ml, which was further reduced by 4 times compared to the MBC value of THC alone ( Figure 1 ).

[0184] Example 2. THC synergistically reduces the effective concentration of gentamicin.

[0185] The objective of this experiment was to determine the minimum inhibitory concentration (MIC) of gentamicin in the presence and absence of THC on Staphylococcus aureus ATCC 25923 strain (gentamicin-susceptible). The cutoff for determining the MIC was set at an optical density (OD) value < 0.1. The results are summarized in Figure 2 middle.

[0186] Results: The MIC value of THC alone was found to be about 0.001 mg / mL, while the MIC value of gentamicin alone was also found to be about 0.001 mg / mL. However, the MIC value of gentamicin combined with THC was synergistically reduced by 4 to 16 times compared to gentamicin alone (0.0005 mg / mL gentamicin + 0.0001 mg / mL THC; 0.0001 mg / mL gentamicin + 0.0005 mg / mL THC).

[0187] Example 3. THC synergistically reduces the effective concentration of gentamicin.

[0188] The objective of this experiment was to determine the minimum inhibitory concentration (MIC) of gentamicin in the presence and absence of THC on Staphylococcus aureus ATCC 25923 strain (gentamicin-susceptible). The cutoff for determining the MIC was set at an optical density (OD) value ≤ 0.1. The results are summarized in Figure 3 middle.

[0189] Results: The MIC value of THC alone was found to be about 0.002 mg / mL, while the MIC value of gentamicin alone was also found to be about 0.002 mg / mL. However, the MIC value of gentamicin combined with THC was synergistically reduced by 4 to 64 times compared to gentamicin alone (0.0002 mg / mL gentamicin + 0.0005 mg / mL THC; 0.00003 mg / mL gentamicin + 0.001 mg / mL THC).

[0190] Example 4. THC synergistically reduces the effective concentration of gentamicin.

[0191] The objective of this experiment was to determine the minimum inhibitory concentration (MIC) of gentamicin in the presence and absence of THC on a multidrug-resistant strain of Staphylococcus aureus, methicillin-resistant Staphylococcus aureus (MRSA). The cutoff for determining the MIC was set at an optical density (OD) value ≤ 0.1. The results are summarized in Figure 4 middle.

[0192] Results: The MIC value of THC alone was found to be about 0.002 mg / mL, while the MIC value of gentamicin alone was found to be about 0.125 mg / mL. However, the MIC value of gentamicin combined with THC was synergistically reduced by 4 to 16 times compared to gentamicin alone (0.0313 mg / mL gentamicin + 0.0005 mg / mL THC; 0.0078 mg / mL gentamicin + 0.001 mg / mL THC).

[0193] Table 1

[0194] THC Gentamycin THC + Gentamycin MRSA 0.001953 0.125 0.000977THC+0.0078125Gentamicin 0.000488THC+0.3125Gentamicin

[0195] Example 5. THC synergistically reduces the effective concentration of ampicillin.

[0196] The objective of this experiment was to determine the minimum inhibitory concentration (MIC) of ampicillin in the presence and absence of THC on methicillin-resistant Staphylococcus aureus (MRSA). The cutoff for determining the MIC was set at an optical density (OD) value ≤ 0.1. The results are summarized in Figure 5 middle.

[0197] Results: The MIC value of THC alone was found to be about 0.002 mg / mL, while the MIC value of ampicillin alone was found to be about 0.5 mg / mL. However, the MIC value of ampicillin combined with THC was synergistically reduced by 2 times compared to ampicillin alone (0.25 mg / mL ampicillin + 0.001 mg / mL THC).

[0198] Example 6. THC synergistically reduces the effective concentration of carbenicillin.

[0199] The goal of this experiment was to determine the minimum inhibitory concentration (MIC) of carbenicillin in the presence and absence of THC on S. pneumoniae. The cutoff for determining the MIC was set at an optical density (OD) value ≤ 0.1. The results are summarized in Figure 6 middle.

[0200] Results: The MIC value of THC alone was found to be about 0.002 mg / mL, while the MIC value of carbenicillin alone was found to be about 0.0002 mg / mL. However, the MIC value of carbenicillin combined with THC was synergistically reduced by 2 times compared to carbenicillin alone (0.0001 mg / mL carbenicillin + 0.0005 mg / mL THC).

[0201] Example 7. THC enhances antibiotics to treat biofilms.

[0202] The goal of this experiment was to determine the minimum inhibitory concentration (MIC) of gentamicin in the presence and absence of THC on established biofilms of Staphylococcus aureus ATCC 25923 strain (gentamicin-susceptible). The cutoff for determining the MIC was set at an optical density (OD) value ≤ 0.1. The results are summarized in Figure 7 middle.

[0203] Results: The MIC value of THC alone was found to be about 0.004 mg / mL, while the MIC value of gentamicin could not even be determined and was clearly much higher than 0.002 mg / mL. However, the MIC value of gentamicin was synergistically reduced to 0.0002 mg / mL compared to gentamicin alone and lower than the combination with THC (0.0002 mg / mL gentamicin + 0.002 mg / mL THC).

[0204] Example 8. Evaluation of combinations of antimicrobials, cannabinoids and / or N-acylethanolamines against non-resistant and resistant bacterial strains.

[0205] The objective of this study was to investigate whether the addition of cannabinoids and / or N-acylethanolamines to antimicrobial agents would reduce the minimum inhibitory concentration (MIC) of the antimicrobial agents against non-resistant and methicillin-resistant Staphylococcus aureus strains (MRSA).

[0206] A non-resistant standard Staphylococcus aureus strain (ATCC 25923) and a clinically resistant MRSA isolate (XU212) were used. Tetracycline and oxacillin were obtained from Sigma Chemical Co. Oxacillin was used instead of methicillin according to the recommendations of the National Committee for Clinical Laboratory Standards (NCCLS) (NCCLS, Summary Minutes, Subcommittee on Veterinary Antimicrobial Susceptibility Testing, January 18-19, 2017). Mueller-Hinton nutrient broth (MHB; Oxoid) was adjusted to contain 20 mg / L Ca 2+ and 10mg / L Mg 2+ .

[0207] An overnight culture of each strain was made into an inoculum density of 5 × 10 in 0.9% saline according to the MacFarland standard. 5Colony forming units (cfu). Tetracycline and oxacillin were dissolved directly in MHB, while norfloxacin and erythromycin were dissolved in DMSO and then diluted in MHB to obtain a starting concentration of 512 μg / mL. PEA and Δ9-tetrahydrocannabinol were dissolved in a vehicle comprising a mixture of ethanol, alkamuls-620 and MHB in a ratio of 1:1:18. Using a Nunc 96-well microtiter plate, 125 μL of MHB was distributed to wells 1-11. Then, 125 μL of test compound or appropriate antibiotic was distributed to one well and diluted continuously on the plate, leaving an empty well for growth control. The final volume was distributed to another well, which was used as a sterile control without MHB or inoculum. Finally, bacterial inoculum (125 μL) was added to the well, and the plate was cultured at 37°C for 18 hours. A DMSO control (3.125%) was also included. All MICs were measured in duplicate. The MIC was determined as the lowest concentration at which no growth was observed. Bacterial growth was detected by a color change from yellow to blue using a methanol solution (5 mg / mL) of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT; Lancaster).

[0208] Treatment groups consisted of the following: (1) control, (2) tetracycline, (3) oxacillin, (4) THC, (5) THC and PEA, (6) tetracycline and THC, (7) tetracycline, THC, and PEA, (8) oxacillin and THC, and (9) oxacillin, THC, and PEA.

[0209] Example 9. Evaluation of combinations of antimicrobial agents, cannabinoids, and / or N-acylethanolamines against biofilm formation.

[0210] The goal of this study was to investigate whether the addition of cannabinoids and / or N-acylethanolamines to antimicrobial agents enhances the antimicrobial agents in the prevention and / or eradication of biofilms.

[0211] Select a variety of pathogenic Staphylococcus targets, including non-resistant and MRSA clinical isolates. It has been previously shown that Staphylococcus pseudointermediates DK729, Staphylococcus pseudointermediates DSM21284 and Staphylococcus pseudointermediates DSM20373 form biofilms by crystal violet staining (Field et al., 2015). Staphylococcus aureus SA113 has also been shown to have the ability to form strong biofilms (Cramton et al., 1999). Staphylococcus aureus strains were grown in cation-regulated Mueller Hinton (CA-MH) (Oxoid) for minimum inhibitory concentration analysis or in tryptic soy nutrient broth (TSB) (Merck) supplemented with 1% glucose at 37°C for biofilm analysis.

[0212] Minimum inhibitory concentration assays were performed in triplicate in 96-well microtiter plates as previously described (Field et al., 2010, 2012, 2015). Briefly, the target strain was grown overnight in appropriate conditions and media, subcultured into fresh nutrient solution and grown to an OD of 600 0.5, diluted in a volume of 0.2 ml to a final concentration of 10 5 cfu / ml. Penicillin G, ampicillin, streptomycin, erythromycin and cefuroxime (Sigma) were resuspended in CA-MH medium to a stock concentration of 128 or 256 μg / ml. Antibiotics were adjusted to a starting concentration of 16, 32 or 64 μg / ml and two-fold serial dilutions of each compound were prepared in a 96-well plate for a total of 12 dilutions. The target strain was then added and after 16 hours of incubation at 37°C, the MIC was determined as the lowest peptide concentration that caused visible growth inhibition.

[0213] A static microtiter plate assay based on previous studies (Kelly et al., 2012) but modified to optimize the assay was used to study biofilm formation and combination treatments. Tryptic soy broth (TSB, Merck) supplemented with 1% D-(C)-glucose (Sigma Aldrich) (TSBg) was used for these assays, which facilitates biofilm formation. Briefly, 2 μl of logarithmic phase cells (10 7 cfu / ml of each culture) was added to 198 μl TSBg for a 1:100 dilution to obtain 10 5cfu / ml starting inoculum; 200ml TSBg was added to one set of wells as a negative control. All wells were inoculated in triplicate. The microtiter plates were then incubated at 37°C for 48 hours to allow biofilm formation. After the biofilm was constructed and washed once with phosphate buffered saline (PBS), the target compound was added to the microtiter plate wells at 1X, 2X, 4X, 8X and 16X. After 24 hours of incubation at 37°C, the plate was removed and gently washed once with PBS, then gently washed with 100mL of a solution containing 500mg XTT / L (2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazol-5-carboxanilide) (Sigma), and 10mM menadione (Sigma) was added to each well. The microtiter plates were incubated at 37°C for 2 hours in the dark. The absorbance was measured at 490 nm using a microtiter plate reader (Molecular Devices Spectramax M3, Sunnyvale CA, USA).The data were obtained in triplicate and calculated and expressed as mean ± SD.

[0214] Treatment groups consisted of the following: (1) control, (2) penicillin, (3) cefuroxime, (4) THC, (5) THC and PEA, (6) penicillin G and THC, (7) penicillin G, THC, and PEA, (8) cefuroxime and THC, and (9) cefuroxime, THC, and PEA.

[0215] Bacterial staining with crystal violet was performed based on the method disclosed in Merritt JH et al., Curr. Protoc. Microbiol., 2005. The following bacterial strains were used for experimental routine: Staphylococcus spp. (including Staphylococcus aureus and Staphylococcus epidermidis), Pseudomonas aeruginosa, Porphyromonas spp. (especially Porphyromonas gingivalis), Moraxella spp., Peptostreptococcus spp., and Enterococcus spp.

[0216] The cells are grown in the microtiter dish for the desired period of time, and then the wells are washed to remove planktonic bacteria. The cells still attached to the wells are then stained using a dye that allows the attachment pattern to be visualized. The surface-associated dye can also be dissolved for semi-quantitative assessment of the biofilm formed.

[0217] Each bacterium was inoculated in 3 to 5 mL culture and grown to stationary phase.

[0218] The culture was diluted 1:100 in medium. 100 μl of each diluted culture was added dropwise to each of four wells in a fresh microtiter plate that had not been treated with tissue culture. The plate was covered and incubated at the optimal growth temperature for the desired time. Then, 20 μl of 0.1% crystal violet solution (pre-filtered through a 0.44 μm filter) was added to each well and stained for 10 minutes at room temperature.

[0219] Example 10. Evaluation of cannabinoids and / or N-acylethanolamines on side effects associated with antimicrobial agents.

[0220] ICR male mice 8 weeks old at the start of the study were used. The mean animal weight at the start of the study was in the range of 24 ± 2 g. The minimum and maximum body weights in each group did not exceed ± 20% of the group mean body weight. The animals were randomly assigned to individual cages on the day of receipt. The animals were allowed to acclimate for seven to nine days.

[0221] The animals were divided into 5 experimental groups (6 animals in each group):

[0222] 1. Control / Vehicle

[0223] 2. Penicillin G

[0224] 3. Penicillin G and THC

[0225] 4. Penicillin G, THC, and PEA

[0226] 5.THC

[0227] During the study period (2 weeks), animals were treated orally by gavage. Mice were weighed at the beginning of the study and on the day of termination. Clinical signs were monitored daily, including, for example, observations for soft stools (diarrhea), body temperature, and the appearance of red or white rushes on the legs of the animals.

[0228] Before the experiment, the dose of antibiotics was titrated to give a tolerable dose that exhibited common antibiotic-related side effects but still had no respiratory depression. Common side effects of antibiotics in animals include diarrhea, itchy skin, fever, and weight gain.

[0229] Example 11. THC and / or THC+PEA combinations synergistically reduce the effective concentration of antibiotics in various bacterial strains.

[0230] The following strains of MRSA were evaluated: community-acquired MRSA (eg USA300), hospital-acquired MRSA, VISA-ATCC 700699, Staphylococcus aureus (strain Mu3 / ATCC 700698).

[0231] Among the other bacteria evaluated were: Enterococcus faecium (E. faecium), including clinically relevant strains and vancomycin-resistant enterococci (VRE).

[0232] The following antibiotics were studied: gentamicin, vancomycin, daptomycin, linezolid, clindamycin, cephalosporins, TMP / SMZ, and deoxycytopeniain.

[0233] Additionally, more than one cannabinoid was evaluated—CBD, CBN, CBG, THCA, and / or THCV.

[0234] The methods used were: micronutrient dilution MIC according to CLSI; synergy assessment using Checkerboard micronutrient dilution MIC and FICI (fractional inhibitory concentration index cutoff) calculation; evaluation of MBC values ​​of selected samples to assess kill time curves; and / or crystal violet staining in BKC.

[0235] The foregoing description of the specific embodiments will fully reveal the general nature of the invention, and others may be easily modified and / or adapted for various applications by applying current knowledge without undue experimentation and without departing from the general concepts, and therefore such adjustments and modifications should and are intended to be understood within the meaning and range of equivalents of the disclosed embodiments. It should be understood that the terms or terminology used herein are for descriptive and not limiting purposes. The tools, materials, and steps for performing the various disclosed functions may take various alternative forms without departing from the invention.

Claims

1. A pharmaceutical composition comprising at least one antimicrobial agent, at least one cannabinoid and a pharmaceutically acceptable carrier.

2. The pharmaceutical composition of claim 1, wherein the antibacterial efficacy of the pharmaceutical composition is similar to or better than the antibacterial efficacy of the same pharmaceutical composition comprising 2 to 150 times the amount of the antibacterial agent and without the cannabinoid.

3. The pharmaceutical composition of claim 1 or claim 2, wherein the weight ratio between the antimicrobial agent and the cannabinoid is from about 1050:1 to 8:1, respectively.

4. The pharmaceutical composition of any one of claims 1 to 3, wherein at least one antibacterial agent is selected from aminoglycosides, penicillins, cephalosporins, tetracyclines, macrolides, clindamycin, sulfonamides, metronidazole, quinolones, derivatives thereof, salts thereof, and any combination thereof.

5. The pharmaceutical composition of claim 4, wherein at least one antibacterial agent is an aminoglycoside or a salt thereof.

6. The pharmaceutical composition of claim 5, wherein the aminoglycoside is gentamicin or a salt thereof.

7. The pharmaceutical composition of claim 5 or claim 6, wherein the antibacterial efficacy of the pharmaceutical composition is similar to or better than the antibacterial efficacy of the same pharmaceutical composition comprising 2 to 64 times the amount of antibacterial agent and without cannabinoids.

8. The pharmaceutical composition of claim 7, wherein the antibacterial efficacy is determined against gentamicin-sensitive bacteria.

9. The pharmaceutical composition of any one of claims 1 to 8, wherein the at least one cannabinoid is tetrahydrocannabinol (THC) or a salt thereof.

10. Use of the pharmaceutical composition of any one of claims 1 to 9 for the preparation of a medicament for the treatment or prevention of bacterial infection or bacterial biofilm.

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